i. HOST/GUEST COMPLEXES FOR ABSOLUTE STEREOCHEMICAL DETERMINATION OF ORGANIC MOLECULES; ii. TOTAL SYNTHESIS OF (–)-SALINOSPORAMIDE A; iii. STEREOCONTROLLED SPIROKETALIZATIONS, AND EFFORTS TOWARDS THE TOTAL SYNTHESIS OF OBTUSIN By Hadi Gholami A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of Chemistry – Doctor of Philosophy 2018 ABSTRACT By Hadi Gholami i. HOST/GUEST COMPLEXES FOR ABSOLUTE STEREOCHEMICAL DETERMINATION OF ORGANIC MOLECULES; ii. TOTAL SYNTHESIS OF (–)-SALINOSPORAMIDE A; iii. STEREOCONTROLLED SPIROKETALIZATIONS, AND EFFORTS TOWARDS THE TOTAL SYNTHESIS OF OBTUSIN This dissertation consists of two parts. In the first part (Chapters I and II), the development of new molecular sensors for detection of asymmetry of organic molecules is disclosed. These sensors, also referred to as “host molecules”, are capable of forming stable complexes with chiral organic molecules. The design of the hosts are such that the point chirality of organic molecules is transferred into a helical twist of the molecular receptor, leading to axial chirality that can be detected as Exciton Coupled Circular Dichroism (ECCD). The absolute configuration of a few classes of organic molecules such as chiral monoamines, cyanohydrins, sulfoxides, phosphine oxides, and mono alcohols is assigned in a microscale, rapid and unambiguous manner. In the second part of this thesis (Chapters III and IV), the development of new organic reactions and their further application towards the synthesis of natural products is discussed. In Chapter III, the successful completion of the total synthesis of (–)- Salinosporamide A is described. During the synthesis of this molecule, new reactions are developed that enabled the completion of this synthesis. In Chapter IV, the development of a halenium initiated spiroketalization and application of this methodology towards the total synthesis of Obtusin is disclosed. The enabling role of the protecting group on the nucleophilic oxygen atom stands as a unique discovery that can improve the reaction outcome. Dedicated to my beloved family for their love and support. iv ACKNOWLEDGEMENTS I would like to express my most sincere gratitude and appreciation to my Ph. D. advisor, Professor Babak Borhan, for his endless support, continuing encouragement, and kind guidance and advise. I cannot possibly be more thankful to him for welcoming me into his research group and providing me with the absolute freedom in thinking to enjoy the research that I was doing. I could not have asked for a better advisor and he for sure has exceeded my expectation. I am greatly thankful to my committee member, Professor William Wulff, Professor James Jackson and Professor Milton Smith, for their helpful suggestions and comments and their support during the past few years. I would like to thank all the members of the Borhan group for their invaluable help and support and friendship during my graduate school. I especially would like to thank Dr. Chrysoula Vasileiou for her limitless support and help. I am immensely thankful to Chrysoula for her patience in proofreading my dissertation and manuscripts. The past members of Borhan group, especially Dr. Kumar Ashtekar, Dr. Mercy Anyika, Dr. Arvind Jaganathan, Dr. Roozbeh Yousefi, and Dr. Elizabeth Santos are acknowledged for their encouragement, friendship and support during my graduate studies. I have had the pleasure and privilege of working with most of the current and former members of Borhan group, to that extend, I am thankful to all of them. I specially thank Jun and Debarshi for helping me with the spectroscopy projects and Yi, Saeedeh and Xinliang for working with v me on the synthetic projects. I am very thankful to all my talented undergraduate students, Olivia, Matt, Katie and Lindsey for helping me with the research. I would like to extent my sincere gratitude to all my friends in the chemistry department for their support and encouragement. I felt welcome by all of them and they will be missed. I am especially thankful to Tayeb Kakeshpour for cheering me up whenever things get tough and my little friend, Mona Borhan for making Saturdays as a special day of week. Saving the best for last, I have no word to describe how thankful I am to my wife, Mina who has been supporting me since the first day we have met. Her unconditional love and support have been my driving force in my daily life. I definitely would not be standing here if it was not because of her support and encouragements. She is my best friend and my better half. I wish I could live up to all her support. I would like to thank our families, specially our parents for their endless love and support. They have given me everything and have patiently supported me during my educations. vi TABLE OF CONTENTS I-3 I-4 I-5 I-6 I-7 I-8 LIST OF TABLES ............................................................................................................. x LIST OF FIGURES ......................................................................................................... xii KEY TO SYMBOLS AND ABBREVIATIONS .............................................................. xxiv Chapter I: Determination of absolute configuration of chiral organic molecules via exciton coupled circular dichroism ................................................. 1 Introduction ..................................................................................... 2 I-1 I-2 Methods for the absolute stereochemical determination of chiral molecules ........................................................................................ 4 Application of chiroptical spectroscopy for determination of absolute configuration of chiral molecules ..................................................... 7 Chemical derivatization of chiral molecules with interacting chromophores for determination of absolute configuration ............ 13 Porphyrin tweezers methodology: host:guest complex formation with chiral molecules for determination of absolute configuration ......... 20 Design of a new host system for determination of absolute configuration of chiral molecules with one sites of attachment ...... 26 Determination of the absolute configuration of chiral monoamines ....................................................................................................... 31 Experimental section ...................................................................... 38 I-8-1 Job’s continuous plot analysis to determine complex stoichiometry ....................................................................... 38 I-8-2 Synthesis of MAPOL I-31 .................................................... 40 REFERENCES ......................................................................................... 46 Chapter II: Determination of absolute configuration of chiral molecules (chiral cyanohydrins, sulfoxides, phosphine oxides and alcohols) .............. 54 II-1 Determination of absolute configuration of chiral cyanohydrins ..... 55 II-2 Determination of absolute configuration of chiral sulfoxides .......... 76 II-3 Determination of absolute configuration of chiral phosphine Oxides ....................................................................................................... 98 II-4 Tuning electronic properties of MAPOL scaffold ......................... 115 II-5 Experimental section ................................................................... 119 II-5-1 Materials and general instrumentations ........................... 119 II-5-2 Procedure for UV-vis and CD titration of Zn-MAPOL with chiral guest molecules (cyanohydrins, sulfoxides or phosphine oxides) .............................................................................. 119 vii II-5-2-1 General procedure for UV-vis measurement ..... 119 II-5-2-2 General procedure for CD measurement .......... 120 II-5-3 Job’s continuous plot analysis to determine the stoichiometry of complex between Zn-MAPOL and chiral guest molecules and binding constant measurements of the complex ....... 121 II-5-4 Synthesis of the host systems .......................................... 122 II-5-4-1 Synthesis of Zn-MAPOL .................................... 122 II-5-4-2 Synthesis of Zn-MAPOL-OMe II-40 ................... 123 II-5-4-3 Synthesis of F4-Zn-MAPOL ............................... 124 II-5-4-4 Synthesis of F30-Zn-MAPOL .............................. 127 II-5-5 Synthesis of chiral cyanohydrins ...................................... 131 II-5-6 Synthesis of chiral sulfoxides ........................................... 132 II-5-7 Conformational modeling studies ..................................... 139 REFERENCES ...................................................................................... 140 Chapter III: Total synthesis of (–)-Salinosporamide A ......................................... 147 III-1 Introduction ................................................................................. 148 III-2 Proteasome Inhibition by Salinosporamide A ............................. 149 III-3 A survey of the previous total syntheses of Salinosporamide A . 150 III-4 Our synthetic approach towards the total synthesis of Salinosporamide A ...................................................................... 153 III-5 Synthesis of Salinosporamide A ................................................. 155 III-5-1 A Survey of anionic approaches for C-5 functionalization 159 III-5-2 Cationic approaches for C-5 functionalization .................. 161 III-5-3 Radical based approaches for C-5 functionalization ........ 164 III-5-4 Neutral based strategies for C-5 functionalization ........... 167 III- 6 The total synthesis of (–)-Salinosporamide A ............................. 190 III- 7 Experimental section ................................................................... 197 III-7- 1 General remarks ............................................................. 197 III-7- 2 Total synthesis of (–)-Salinosporamide A ....................... 198 III-7- 3 Crystal data ..................................................................... 229 III-7-3-1 Crystal data for III-91b (BB817C) ...................... 229 III-7-3-2 Crystal data for (–)-III-25 (BB817D) .................. 244 REFERENCES ...................................................................................... 259 Chapter IV: Diastereoselective halonium initiated spiroketalization; towards the total synthesis of Obtusin .................................................................. 267 IV- 1 Introduction; structural features of spiroketal moiety .................. 268 IV- 2 Chemical synthesis of spiroketals ............................................... 271 IV- 3 Development of halenium initiated spiroketalization ................... 276 IV- 4 Development of mono and di-brominated spiroketalization reaction .................................................................................................... 280 IV- 5 Towards the total synthesis of Obtusin ....................................... 291 IV-5- 1 Introduction ..................................................................... 291 viii IV-5- 2 progress towards the total synthesis of Obtusin ............. 293 IV- 6 Experimental section ................................................................... 308 IV-6-1 General remarks ............................................................ 308 IV-6-2 General procedure for bromo spiroketalization of THP- protected keto-alkenols ................................................. 309 IV-6-3 General procedure for dibromo spiroketalization of THP- protected keto-alkenols ................................................. 315 IV-6-4 Synthesis of ketones IV-116 to IV-121 .......................... 321 IV-6-5 General procedure for synthesis of ketone IV-131 to IV-133 ....................................................................................... 335 IV-6-6 Procedure for the synthesis of THP-protected keto alkenols IV-136 ............................................................................ 345 IV-7 Experimental date for the synthesis of Obtusin .......................... 347 REFERENCES ...................................................................................... 363 ix LIST OF TABLES Table II-1: Predicted and observed ECCD for cyanohydrins complexed with Zn- MAPOL ..................................................................................................... 65 Table II-2: Predicted and observed ECCD for sulfoxides complexed with ZnMAPOL .................................................................................................................. 85 Table III-1: Optimization of the C-H insertion reaction at the C-5 center ................. 173 Table III-2: Optimization of olefin cleavage towards installation of carbonyl on C-5 183 Table III-3: Selective deprotection of the TBS ether in the presence of Boc protecting group ...................................................................................................... 186 Table III-4: Cardova’s reported asymmetric aziridination of a,b-unsaturated aldehyde ............................................................................................................... 191 Table III-5: Fractional atomic coordinates (×104) and equivalent isotropic displacement parameters (Å2×103) for III-91b. Ueq is defined as 1/3 of the trace of the orthogonalised Uij ................................................................................... 234 Table III-6. Anisotropic displacement parameters (×104) III-91b. the anisotropic displacement factor exponent takes the form: -2p2[h2a*2 × U11+ ... +2hka* × b* × U12] ............................................................................................. 235 Table III-7: Bond lengths in Å for III-91b .................................................................. 237 Table III-8: Bond angles in ° for III-91b ..................................................................... 239 Table III-9: Hydrogen fractional atomic coordinates (×104) and equivalent isotropic displacement parameters (Å2×103) for III-91b. Ueq is defined as 1/3 of the trace of the orthogonalised Uij ................................................................ 241 Table III-10: Fractional atomic coordinates (×104) and equivalent isotropic displacement parameters (Å2×103) for (–)-III-25. Ueq is defined as 1/3 of the trace of the orthogonalised Uij ................................................................................... 249 Table III-11: Anisotropic Displacement Parameters (×104) (–)-III-25. The anisotropic displacement factor exponent takes the form: -2p2[h2a*2 × U11+ ... +2hka* × b* × U12] ................................................................................................. 251 x Table III-12: Bond lengths in Å for (–)-III-25 ............................................................... 252 Table III-13: Bond angles in ° for (–)-III-25 ................................................................. 254 Table III-14: Hydrogen fractional atomic coordinates (×104) and equivalent isotropic displacement parameters (Å2×103) for (–)-III-25. Ueq is defined as 1/3 of the trace of the orthogonalised Uij ................................................................ 256 Table III-15: Hydrogen bond information for (–)-III-25 ............................................... 258 Table IV-1: Substrate scope initiated diastereoselective spiroketalization ..................................................................................... 286 Table IV-2: Diastereoselective dibromo spiroketalization ......................................... 289 Table IV-3: Bromo etherification of alcohol IV-86 ..................................................... 296 Table IV-4: Bromo etherification of protected alcohol IV-101 ................................... 301 for halenium atom xi LIST OF FIGURES Figure I-1. Representative enantiomers with drastically different biological activities . 3 Figure I-2. Assignment of the absolute configuration of a secondary alcohol using Mosher method. Structure of the stable conformer that leads to the change in the chemical shift of the derivatized alcohol and the 1H-NMR traces of both diastereomers, as well as the proposed mnemonic (dashed box) are demonstrated. ............................................................................................ 5 Figure I-3. a. Plane-polarized light composed of two circularly polarized components, lcp and rcp. As long as the phases of intensities of these two components remain the same, the resultant oscillating vector will be in the same plane and magnitude. b. If the rcp component become less intense (due to interaction with chiral media) the electric vector of light follows an elliptical path (elliptically polarized light, green oval). c. The degree of ellipticity is define as y, which is proportional to the circular dichroism. The rotation of ellipse (a) corresponds to optical rotation and ORD .................................. 8 Figure I-4. Positive or negative CD spectra or Cotton effects ..................................... 9 Figure I-5. Schematic exciton band energy diagram of system with two interacting chromophores in various geometries. The ovals indicate chromophores with their electric dipole transition moments. The magnitude of the net electric dipole transition moments (µnet) is deduced by employing a simple vector model analysis. ......................................................................................... 12 Figure I-6. Schematic demonstration of two independently conjugated chromophores arranged in a helical twist. The electric dipole transition moments (blue line) are along the long axis of the benzoate moieties. The side view of interacting chromophores indicates clockwise or counter-clockwise arrangement of chromophores depending on the chirality of the diol. The clockwise helicity generates a positive ECCD spectrum (shown on tap right) .................................................................................................................. 14 Figure I-7. Dioxolane formation of chiral diols with I-6 leads to CD active complexes. The highlighted diastereomeric complexes (box) depict equilibrating dioxolanes that favor M conformation due to the chirality of the bound diol. .................................................................................................................. 15 xii Figure I-8. Development of an azepine probe (I-8) for sensing the absolute configuration of chiral carboxylic acids ..................................................... 16 Figure I-9. Development of stereodynamic molecular probe I-10 for determination of enantiomeric composition and absolute configuration of chiral amino alcohols through imine formation .............................................................. 17 Figure I-10. Structure of propeller-like Cu(II) complex with chirality on the backbone of one of the quinoline based ligand ............................................................. 18 Figure I-11. Induced helical twist of the achiral quinoline based ligand on Cu(II) due to stereochemistry of the bound amino acid (methionine) ............................ 19 Figure I-12. Structure of a copper based molecular probe for sensing the chirality of carboxylic acids......................................................................................... 19 Figure I-13. Stereodynamic assembly of chiral alcohol with in situ generated iminium I-19 into a trigonal bipyramidal scaffold leading to P or M helicities depending on the chirality of alcohol ........................................................ 20 Figure I-14. (a) Structure of C5-TPP tweezer; (b) model for complexation of chiral diamine with C5-TPP tweezer (a bidentate coordination of diamines with the porphyrin tweezer leads to an observed helical twist); (c) Experimentally measured ECCD signal and proposed model for the preferred helical twist (left), counter-clockwise arrangement of interacting dipole transition moments leads to a negative ECCD signal. ............................................. 22 Figure I-15. Frontier orbital energy and charges of zinc porphyrins. Bonding affinity of iso-propanol and iso-propylamine with Zn-TPP-monoester and Zn-TPFP- monoester as analogues of Zn-TPP tweezer and Zn-TPFP tweezer (dashes box depicts structure of C5-Zn-TPFP tweezer). ....................................... 24 Figure I-16. Pictorial representation of functional group families with methods for assignment of their stereochemistry. Chiral molecules with single site are derivatized with a carrier to form a bidentate guest molecule (left panel) prior to complex formation with tweezers; Structure of host systems, with various linker size as well as tuned steric and electronic profiles that enables determination of absolute configuration of the latter functionalities are shown at the bottom of this figure ............................................................. 25 Figure I-17. Rational design of a new host system capable of sensing chirality of molecules with one site of attachment. Linker modification and change in the binding paradigm are notable adjustments ......................................... 27 xiii Figure I-18. Structure of newly designed host system. Biphenyl back bone, capable of adopting P or M helicity, renders equilibrating atropisomers. Upon introduction of chiral guest, that rests in the highlighted binding sites, one of the helicities become more prominent leading to the corresponding ECCD signal......................................................................................................... 28 Figure I-19. Successful synthesis of MAPOL I-31 ......................................................... 29 Figure I-20. Complexation of MAPOL with chiral monoamine II-37S leads to a strong ECCD signal with a negative sign ............................................................. 31 Figure I-21. 1H-NMR spectra of titration of MAPOL 1 with S-(+)-1-cyclohexylethylamine I-37S. Highlighted with blue indicates the changes in the chemical shift of the methyl group of the amine substrate and the red area shows the changes in the chemical shift of the MAPOL backbone upon complexation with amine I-37S ....................................................................................... 33 Figure I-22. Job’s continuous plot of MAPOL I-31 with I-37S: following the changes in chemical shift of MAPOL at 7.55 ppm (left) and chemical shift of amine I- 37S at 0.99 ppm ....................................................................................... 34 Figure I-23. a) Complexation of I-37S with MAPOL leads to a strong negative ECCD signal. b) Binding of amine I-37S, with the P-helicity of MAPOL leads to steric clash of the methyl group with the porphyrin ring. Thus, positive ECCD with I-37S bound to MAPOL is not anticipated. c) The same binding of I- 37S with the M-helicity of MAPOL places the medium size methyl group in the most sterically avaiable position. This favorable arrangement leads to the anticipated positive ECCD spectrum .................................................. 35 Figure II-1. Synthetic utility of cyanohydrin moiety ...................................................... 55 Figure II-2. Mosher’s method determination of absolute configuration of chiral cyanohydrins ............................................................................................. 56 Figure II-3. Application of chiral solvating agents for determining the absolute configuration of chiral cyanohydrins ......................................................... 58 Figure II-4. Porphyrin their envisioned complexation with chiral cyanohydrins for determination of absolute configuration ........................ 59 Figure II-5. Structure of MAPOL and potential modifications to improve binding interaction ................................................................................................. 60 tweezers and xiv Figure II-6. UV-vis spectra of Zn-MAPOL (1 μM) in different solvents at room temperature .............................................................................................. 61 Figure II-7. ECCD spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl) acetonitrile II-6R ........................................................................................ 62 Figure II-8. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-2-hydroxy-2-(o- tolyl) acetonitrile II-6R (50 equivalents) in different solvents at 0 ºC ......... 63 Figure II-9. ECCD spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl) acetonitrile II-6R (50 equivalents) at different temperatures ..................... 64 Figure II-10. UV-vis spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl) acetonitrile II-9R ........................................................................................ 67 Figure II-11. Jobs’ plot of Zn-MAPOL with (R)-4-methylmandelonitrile II-9R absorbance at 410 nm .............................................................................. 68 Figure II-12. Quenching the ECCD signal of II-6R in complex with Zn-MAPOL by addition of methanol ............................................................................................... 70 Figure II-13. Quenching the ECCD signal of II-6R in complex with Zn-MAPOL by addition of acetonitrile (MeCN) ............................................................................... 71 Figure II-14. a) Bidentate binding of II-6R, with the P-helicity of Zn-MAPOL, places the large aryl group in the most sterically available position. This favorable arrangement leads to the anticipated positive ECCD spectrum. b) Complexation of 5R with Zn-MAPOL leads to a strong positive ECCD signal. c) The same bidentate binding with the M-helicity leads to steric clash of the large aryl group with the porphyrin ring. Thus, negative ECCD with II-6R bound to Zn-MAPOL is not anticipated ..................................................... 72 Figure II-15. Side and top view of geometry optimized (DFT-B3LYP/6-31G*) complex of II-10R bound to Zn-MAPOL. The model shows the bulky t-butyl group in the open quadrant, the hydrogen bonding of the alcohol with the biphenol core, and the interaction of the cyano group with the metal center. The optimized geometry predicts a positive ECCD, as observed .................................... 73 Figure II-16. ECCD spectra of Zn-MAPOL (1 μM) complexed with 4-chloromandelonitrile II-7 (50 equiv) at different enantio excess (left). The plot of ECCD amplitude versus %ee ............................................................................................... 75 Figure II-17. Example of bioactive drugs with chirality at the sulfur atom ..................... 76 xv Figure II-18. Chemical modification and derivatization of chiral sulfoxides for determination of absolute configuration .................................................... 77 Figure II-19. ECCD spectrum of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in hexane at 0 ºC ........................................... 79 Figure II-20. UV-vis spectra of Zn-MAPOL (1 μM) and Zn-MAPOL complexed with (R)- p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC (solvents that yield ECCD signal) ............................................................................. 79 Figure II-21. UV-vis spectra of Zn-MAPOL (1 μM) and Zn-MAPOL complexed with (R)- p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC (solvents that do not yield ECCD signal) .................................................................. 80 Figure II-22. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC ............................ 81 Figure II-23. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC ............................ 82 Figure II-24. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane ............................................................ 82 Figure II-25. UV-vis spectral change upon titration of Zn-MAPOL (1 μM) with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane .................................................... 83 Figure II-26. Job’s plot of Zn-MAPOL with (S)-p-tolyl vinyl sulfoxide II-20R absorbance at 409 nm .................................................................................................. 84 Figure II-27. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC .............. 84 Figure II-28. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane ............................................................ 88 Figure II-29. X-ray crystal structure of Zn-TPP in complex with dimethyl sulfoxide (hydrogens are omitted for clarity) ............................................................ 89 Figure II-30. a) Bidentate binding of II-20R, with the P-helicity of Zn-MAPOL, places the large aryl group in the most sterically available position. This favorable arrangement leads to the anticipated positive ECCD spectrum. b) Complexation of II-20R with Zn-MAPOL leads to a strong positive ECCD signal. c) The same bidentate binding with the M-helicity leads to steric xvi clash of the large aryl group with the porphyrin ring. Thus, negative ECCD with II-20R bound to Zn-MAPOL is not anticipated ................................... 90 Figure II-31. Energy minimized (B3LYP/6-31G*) structures of the P and M helical Zn- MAPOL complexed with II-22R. The length of the substituents, measured as L1 and L2, dictate the preferred helicity of the host system. The shorter L2 of the phenyl group leads to smaller steric interactions with the porphyrin ring that is not bound to the sulfoxide ....................................................... 91 Figure II-32. Sterioml L parameter (total length of the substituent along the primary bond axis) and the measure length (L1, length of substituent to the furthest heavy atom) are illustrated for 1,2-dimethylpropyl group .................................... 92 Figure II-33. Negative ECCD spectrum obtained with Zn-MAPOL complexed with 50 equivalents of (S)-p-toluene sulfinamide II-29S at 0 ºC in hexane ........... 94 Figure II-34. UV-vis spectral change upon titration of Zn-MAPOL (1 μM) with (S)-p- toluene sulfinamide S- II-29S at 0 ºC in hexane ....................................... 94 Figure II-35. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (S)-p-toluene sulfinamide II-29S at 0 ºC ............. 95 Figure II-36. X-ray crystal structure of Zn-TPP complexed with sulfinamide II-34S (hydrogens are omitted for clarity) ............................................................ 96 Figure II-37. Complexation of esomeprazole (50 µM) with Zn-MAPOL (1 µM) yields a positive ECCD spectrum, correctly predicting the S stereochemistry of the molecule. The assignment is based on considering the benzoimidazole side chain as the larger substituent (longer linear length as compared to the pyridyl side chain) ..................................................................................... 97 Figure II-38. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (R)-methyl phenyl propyl phosphine oxide II-30R at 0 ºC in hexane .......................... 99 Figure II-39. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (S)-methyl- phenyl-propyl phosphine oxide II-30S at 0 ºC in hexane ....................... 100 Figure II-40. UV-vis spectral change upon titration of Zn-MAPOL (1 μM) with (R)-methyl- phenyl-propyl phosphine oxide II-30R ................................................... 100 Figure II-41. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (R)-methyl-phenyl-propyl phosphine oxide II-30R ............................................................................................................... 101 xvii Figure II-42. Structures of Sofosbuvir and epi-Sofosbuvir ......................................... 104 Figure II-43. ECCD spectra of epi-Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, DCM was added ................................. 105 Figure II-44. ECCD spectra of epi-Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, NO dichloromethane was added (a), 40 µL dichloromethane was added (b) and 100 µL dichloromethane was added (c) ........................................................................................................... 106 Figure II-45. ECCD spectra of Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, DCM was added ..................................... 107 Figure II-46. ECCD spectra of Sofosbuvir complexed with ZnMAPOL (1 µM) in dichloromethane .................................................................................... 108 Figure II-47. ECCD spectra of epi-Sofosbuvir complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C ........................................................................ 109 Figure II-48. ECCD spectra of epi-Sofosbuvir complexed with ZnMAPOL (1 µM) in chloroform at 0 °C .................................................................................. 110 Figure II-49. ECCD spectra of Sofosbuvir complexed with ZnMAPOL (1 µM) in chloroform at 0 °C .................................................................................. 110 Figure II-50. ECCD spectra of Sofosbuvir (50 equiv) complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C, effect of addition of acetone ...................... 111 Figure II-51. ECCD spectra of epi-Sofosbuvir (50 equiv) complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C, effect of addition of acetone ............... 112 Figure II-52. Coordination and the working model of Sofosbuvir complexed with ZnMAPOL. P-helicity is favored and positive ECCD signal was observed ............................................................................................................... 113 Figure II-53. Coordination and the working model of epi-Sofosbuvir complexed with ZnMAPOL. M-helicity is favored and negative ECCD signal was observed ............................................................................................................... 114 Figure II-54. Structural features of F6-backbone ........................................................ 115 Figure II-55. Structure and UV-vis spectrum of F4-MAPOL II-32 (1 mM solution) in hexane at room temperature. ............................................................................. 116 xviii Figure II-56. Structure and UV-vis spectrum of F30-Zn-MAPOL II-34 (1 mM solution) in hexane at room temperature. ................................................................ 117 Figure II-57. ECCD spectrum of F30-Zn-MAPOL (1 μM) complexed with (R)-1- phenylethanol II-33R (100 equiv) in hexane at 0 ºC .............................. 118 Figure III-1. Structure of Salinosporamide A and its closely related natural products ............................................................................................................... 149 Figure III-2. Proposed inhibitory mechanism of Salinosporamide A in the proteasome active site ............................................................................................... 150 Figure III-3. First total synthesis of Salinosporamide A with the highlighted Baylis- Hillman cyclization as the key reaction of the synthesis ........................ 151 Figure III-4. Danishefsky’s total synthesis of Salinosporamide A .............................. 151 Figure III-5. Biosynthetically inspired total syntheses of Salinosporamde A ............. 152 Figure III-6. Conia-Ene type reaction to assemble the core of Salinosporamide A ... 153 Figure III-7. Our developed methodology that give access to pyrrolidinone core and its exploitation for the synthesis of Salinosporamide A .............................. 154 Figure III-8. Synthesis of aziridine-alcohol III-25 for implementation of the tandem aza/Payne/hydroamination reaction ...................................................... 156 Figure III-9. Tandem aza/Payne/hydroamination and selective epoxide opening, access to decorated 2-pyrrolidinone core .......................................................... 157 Figure III-10. Installation of the carbon side chain on C-3 and deprotection of the amide moiety of the 2-pyrrolidinone core ......................................................... 158 Figure III-11. Functionalization of C-5, an anionic route ............................................. 160 Figure III-12. Anionic functionalization of C-5 and the challenges facing this strategy ............................................................................................................... 161 Figure III-13. Cationic approaches towards C-5 functionalization ............................... 162 Figure III-14. Functionalization of C-5, a radical initiated oxidative strategy ............... 163 Figure III-15. Radical based strategies towards installation of functional group at C-5 ............................................................................................................... 165 xix Figure III-16. General reactivity profile in comparison to the direct C-H functionalization; controlling factor in the C-H insertion reactions ..................................... 168 Figure III-17. Application of C-H insertion strategy in the total synthesis of natural products and complex molecules .......................................................... 169 Figure III-18. Functionalization at C-5 center employing metal mediated decomposition of a-diazo carbonyl, a C-H insertion strategy ............................................ 171 Figure III-19. Envisioned functionalities to deliver the active carbene to the C-5 center for the C-H insertion reaction ...................................................................... 172 Figure III-20. Installation of the carbonyl moiety on the C-5 center ............................. 176 Figure III-21. Selective oxidative cleavage of the terminal olefin ................................ 177 Figure III-22. Oxidative cleavage of the internal olefin, b-lactone ring formation ........ 178 Figure III-23. Oxidative cleavage of the internal olefin towards deprotection of the pyrrolidinone nitrogen ............................................................................ 179 Figure III-24. Strategies for the removal of the group on the pyrrolidinone nitrogen ... ............................................................................................................... 180 Figure III-25. An envisioned enone moiety for the facile removal of the group on the pyrrolidinone nitrogen after oxidative cleavage of the internal olefin ..... 181 Figure III-26. Synthesis of the envisioned enone core and its X-ray crystal structure 182 Figure III-27. Access to the advanced bicyclic core III-94b, similar to Fukuyama’s intermediate towards Salinosporamide A .............................................. 185 Figure III-28. Completion of the racemic synthesis of Salinosporamide A .................. 188 Figure III-29. Synthesis of a,b-unsaturated aldehyde III-107 for implementation of the asymmetric aziridination reaction .......................................................... 192 Figure III-30. Asymmetric aziridination of a,b-unsaturated aldehyde, access to aziridine- alcohol (–)-III-25 ..................................................................................... 193 Figure III-31. Total synthesis of (–)-Salinosporamide A .............................................. 195 Figure III-32. X-ray structure of III-91b (view 1) .......................................................... 232 xx Figure III-33. X-ray structure of III-91b (view 2) .......................................................... 233 Figure III-34. X-ray structure of (–)-III-25 (view 1) ...................................................... 248 Figure III-35: The Model has Chirality at C1 (Chiral SPGR) S Verify; The Model has Chirality at C2 (Chiral SPGR) S Verify; The Model has Chirality at C3 (Chiral SPGR) S Verify ...................................................................................... 248 Figure III-36: The following hydrogen bonding interactions with a maximum D-D distance of 2.9 Å and a minimum angle of 120 ° are present in (–)-III-25: O1–O3: 2.865 Å .................................................................................................. 249 Figure IV-1. Spiroketal moiety embedded in the structure of natural products .......... 268 Figure IV-2. Anomeric effect, principle and structural requirements .......................... 269 Figure IV-3. The anomeric arrangements in 6,5- and 6,6-spiroketal frameworks ..... 270 Figure IV-4. Different approaches to access the spiroketal moiety ........................... 272 Figure IV-5. List’s approach to access spiroketal moiety in an enantioselective fashion ............................................................................................................... 273 Figure IV-6. Nagorny’s approach to access the spiroketal in an enantioselective fashion ................................................................................................................ 274 Figure IV-7. Carreira’s approach to access spiroketal in an enantioselective fashion ............................................................................................................... 275 Figure IV-8. Examples of halogenated natural products. The halogenated spiroketals are highlighted in blue ............................................................................ 276 Figure IV-9. Some of the developed methodologies by our group in the field of halofunctionalization of olefins ............................................................... 277 Figure IV-10. Experimental validation of the halenium affinity (HalA) ........................ 278 Figure IV-11. The effect of nucleophile on the olefin halofunctionalization, demonstration of nucleophile-assisted alkene activation (NAAA) ................................. 279 Figure IV-12. Our approach to access halogenated spiroketals ................................. 280 Figure IV-13. Bromo spiroketalization of ketone tethered to an unprotected alcohol . 281 xxi to access Obtusin IV-22. Retrosynthetic approach Figure IV-14. X-ray crystal structure of macrocyclic IV-45 ......................................... 282 Figure IV-15. Bromo spiroketalization of ketone with a protected tethered alcohol .... 283 Figure IV-16. Computed HalA values for ketone bearing different groups ................. 284 Figure IV-17. X-ray crystal structure of product IV-54 and NOESY of IV-45 .............. 285 Figure IV-18. Synthesis of 5,6- and 5,5-spiroketals .................................................... 287 Figure IV-19. Dibromospiroketal formation by excluding ethanol from the reaction mixture ............................................................................................................... 288 Figure IV-20. X-ray crystal structure of dibromo-spiroketal IV-67 ............................... 290 Figure IV-21. Structure of Obtusin and its proposed biogenesis ................................ 291 Figure featuring dibromo- spiroketalization and bromo-etherification steps .................................... 292 Figure IV-23. Installation of the cis olefin, access to epoxy alcohol IV-91 .................. 294 Figure IV-24. Installation of the ketone surrogate, towards bromo-etherification reaction ............................................................................................................... 295 Figure IV-25. Assignment of the relative stereochemistry of the diastereomers IV-96 and IV-97 ...................................................................................................... 297 Figure IV-26. On the origin of the diastereoselectivity leading to IV-87 and IV-88 ..... 299 Figure IV-27. Rational design to improve the diastereoselectivity of the bromo- etherification reaction ............................................................................. 300 Figure IV-28. Comparison of the bromo-etherification with and without protection group on the nucleophilic oxygen .................................................................... 303 Figure IV-29. Direct addition of alkyl lithium to the nitrile moiety towards installation of the ketone functionality ................................................................................ 304 Figure IV-30. Approaches to improve the efficiency of the conversion of nitrile to ketone ............................................................................................................... 305 xxii Figure IV-31. A different approach to convert epoxy alcohol IV-90 to advanced intermediate IV-109 ............................................................................... 306 xxiii KEY TO SYMBOLS AND ABBREVIATIONS Å cm-1 M mM mg mmol > < CHCl3 DMF ESI Et3N EtOAc HOMO HRMS LUMO mp mbar MgSO4 NaOH angstrom wavenumber molar millimolar milligram millimole larger than less than chloroform dimethylformamide electrospray Ionization triethylamine ethyl acetate highest occupied molecular orbital high resolution mass spectrometry lowest unoccupied molecular orbital melting point milibarr magnesium sulfate sodium hydroxide xxiv NMR IR α [a]D Å A ACN AcOH nuclear magnetic resonance infrared angle of rotation specific rotation angstrom CD amplitude acetonitrile acetic acid aromatic Ar BF3•OEt2 boron trifluoride diethyl ether BINOL Bn BnBr BnCl CD CE cm d DCM DDQ DET DFT 1,1’-Bi-2-naphthol benzyl benzyl bromide benzyl chloride circular dichroism Cotton effect centimeter doublet dichloromethane 2,3-dichloro-5,6-dicyano-1,4-benzoquinone diethyl tartrate density functional theory xxv DIPT DMAP DMF ee Et3N ε ECCD Et2O EtOAc eq g h HPLC Hz iPr IR J Kassoc LAH m m mCPBA diisobutylaluminum hydride 4-diaminopyridine triethylamine N,N-dimethylformamide enantiomeric excess molar absorption coefficient Exciton Coupled Circular Dichroism diethyl ether ethyl acetate equivalents gram(s) hour(s) high pressure liquid chromatography hertz NMR coupling constant association constant magnetic dipole transition moment multiplet lithium aluminum hydride isopropyl infrared 3-chloroperoxybenzoic acid xxvi MeOH min methanol minute µg MHz M μM MS m/z n NaOH nm NMR ORD PCC Ph PMB q R s rt t TBAF micro gram megahertz molar micro molar mass spectrometry mass to charge ratio refractive index sodium hydroxide nanometer nuclear magnetic resonance optical rotatory dispersion pyridinium chlorochromate phenyl para-methoxybenzyl quartet singlet room temperature triplet tetrabutylammonium fluoride rotational strength xxvii tert-butyl dimethylsilyl trifluoroacetic acid tetrahydrofuran ultraviolet-visible spectroscopy 5-(4-carboxyphenyl)-10,15,20-triphenylporphyrin tweezer zinc tetraphenylporphyrin TBS TFA THF UV-vis TPP-tz Zn-TPP Zn-TPP-tz zinc 5-(4-carboxyphenyl)-10,15,20- triphenylporphyrin tweezer Zn-TPFP zinc 5-(4-carboxyphenyl)-10,15,20-tri(pentafluorophenyl)porphyrin tweezer Zn(OAc)2 lcp rcp VCD ECD edtm, µ UV InCl3 TFA BBr3 Dd CDCl3 X TMEDA zinc acetate left circularly polarized light right circularly polarized light vibrational circular dichroism electronic circular dichroism electric dipole transition moment ultraviolet spectroscopy indium(III) chloride trifluoroacetic acid boron tribromide difference in chemical shift deuterated chloroform mole fraction tetramethylethylenediamine xxviii NH4Cl TLC Na2SO4 CH2Cl2 NH4HCO3 MPA equiv CaH2 THF EtOH SeO2 tBuOOH NBS DMP DMS MgBr2 AIBN DBU dr PPh3 TMSCHN2 LDA ammonium chloride thine layer chromatography sodium sulfate dichloromethane ammonium bicarbonate methoxyphenyl acetic acid equivalents calcium hydride tetrahydrofuran ethanol Selenium dioxide tert-butyl hydroperoxide N-bromosuccinimide Dess-Martin periodinane dimethyl sulfide magnesium bromide 2,2′-azobis(2-methylpropionitrile) 1,8-diazabicyclo(5.4.0)undec-7-ene diastereomeric ratio triphenylphosphine trimethylsilyldiazomethane lithium diisopropylamide xxix chromium trioxide 4-toluenesulfonyl chloride sodium iodide potassium tert-butoxide hydrochloric acid CrO3 TsCl NaI KOtBu HCl NaHMDS sodium bis(trimethylsilyl)amide KHMDS MMPP Boc TBSCl KOH NaOAc DMSO Me3SOI OsO4 CSA ZnCl2 Me3Al Ph3PCl2 BOPCl NAAA n.a. potassium bis(trimethylsilyl)amide magnesium monoperoxyphthalate tert-butyloxycarbonyl tert-Butyldimethylsilyl chloride potassium hydroxide sodium acetate dimethyl sulfoxide trimethylsulfoxonium iodide osmium tetroxide camphorsulfonic acid zinc chloride trimethylaluminium triphenylphosphine dichloride bis(2-oxo-3-oxazolidinyl) phosphinic chloride nucleophile assisted alkene activation not applicable xxx n.d. Na2SO4 NaH HalA MOM TES PMB TBDMH DCDMH NOSEY THP BF3•Et2O TBCO HMPA nOe TMSCl TMS DHP tBuLi nBuLi PPTS DIBAL-H not determined sodium sulfate sodium hydride halenium affinity methoxymethyl acetal 4-methoxybenzyl ether triethylsilane 1,3-dibromo-5,5-dimethylhydantoin 1,3-dichloro-5,5-dimethylhydantoin nuclear Overhauser effect spectroscopy tetrahydropyran boron trifluoride diethyl etherate 2,4,4,6-tetrabromo-2,5-cyclohexadienone hexamethylphosphoramide nuclear Overhauser effect trimethylsilyl chloride trimethylsilyl dihydropyran tert-butyllithium n-butyllithium pyridinium p-toluenesulfonate diisobutylaluminium hydride xxxi Chapter I: Determination of absolute configuration of chiral organic molecules via exciton coupled circular dichroism 1 I-1 Introduction Life on earth, as we know it, is built with chiral scaffolds. From simple amino acids and sugars that are the foundation of biological life, to hormones, antibiotics, natural products, etc., all are examples of chiral objects. Chirality has been the subject of ongoing research in different branches of science, and understanding of chirality and its impact on chemistry and life has intrigued scientists since the introduction of this concept over a century ago.1 A chiral molecule is capable of existing in two enantiomeric states. Enantiomers refer to non-superimposable, yet mirror images of a structure. Much akin to the left and right hand that are mirror images of each other but cannot be superimposed, a chiral molecule cannot be superimposed on its mirror image either. At the molecular level, a structure is chiral if there are no elements of symmetry. When a molecule is chiral, there are two different ways to arrange the atoms. The two enantiomers of a molecule can have different behaviors as they interact with an asymmetric environment such as the active site of a receptor. One of the areas where the different actions of the two enantiomers is significantly important is when these molecules interact with a living organism such as the human body.3 As such, the different enantiomers of a chiral molecule can have different taste or smell due to their different interactions with the sensory receptors. For instance, the two enantiomers of mercaptohexanal, despite their similar physical and chemical characteristic have different smells (Figure I-1). The S enantiomer has a fruity odor while the R enantiomer has a pungent sulfur smell. Another example where chirality has led to different 2 properties, is observed in limonene. (R)-Limonene is the isomer that is found in orange and is the origin of its smell. The S enantiomer of this compound is the fragrant ingredient of lemon (Figure I-1). The impact of stereochemistry is far more pronounced when chirality is embedded in pharmaceutical products. Different enantiomers of a drug can interact differently with the chiral environment of a living organisms. Therefore , the mirror image of an active drug molecule can have devastatingly deleterious effects.1 SH O SH O (R)-3-mercaptohexanal (I-1R) H (S)-3-mercaptohexanal (I-1S) H (R)-Limonene (I-2R) (S)-Limonene (I-2S) O N O O O NH O N O O O NH (R)-Thalidomide (I-3R) (S)-Thalidomide (I-3S) Figure I-1. Representative enantiomers with drastically different biological activities. Thalidomide is an example where the two enantiomers lay at the two opposite ends of the spectrum with regards to the human health (Figure I-1). While the R enantiomer has antiemetic properties, the S enantiomer causes fetal abnormalities in infants. In 1960s, this drug was used as its racemic mixture and when it was prescribed to pregnant women, causing terrible birth defects for some babies.4 Catastrophic events such as this case and many other instances, have led to strict regulations with respect to the effect of the each enantiomers of a drug molecule. Currently, chiral drugs are only 3 approved as a single enantiomer, which makes the determination of absolute stereochemistry of a molecule an important task. Thus, introducing simple, efficient and reliable techniques for the assignment of absolute configuration of chiral molecule are in high demand. Moreover, the assignment of stereochemistry is central to the progress of chemistry in any of its sub-disciplines that require the use of molecule for function, whether it is a pharmaceutical agent, synthetic intermediate, identification of new natural products, etc. The aim of our research program is to develop methodologies that could be used for the absolute stereochemical determination of chiral compounds. The goals are to provide simple, the stereochemistry of chiral centers. rapid, efficient, and non-ambiguous procedures for establishing for function, whether it that require the use of molecules I-2 Methods for the absolute stereochemical determination of chiral molecules Assignment of stereochemistry is crucial to the progress of chemistry in any of its sub-disciplines is pharmaceutical agent, synthetic intermediate, identification of new natural products, etc. To this end, many different approaches have been reported to establish the absolute configuration of chiral molecules. One of the most widely utilized techniques for his purpose is Mosher ester analysis of chiral molecules.5-9 According to this method, enantiomeric structures are derivatized with a chiral aryl carboxylic acids, with known configuration, to form two diastereomers. Derivatization can be performed on chiral molecules that contain hydroxyl and amine functionalities to form the corresponding 4 ester or amide derivatives. NMR analysis of the diastereomeric derivatives can lead to the elucidation of absolute configuration of the derivatized molecule. The anisotropic effect of the aryl substituents on the derivatization agent (aryl carboxylic acid) causes changes in the chemical shift of the substituents on the chiral target molecule. Interpretation of the changes in the chemical shifts can lead to determination of absolute configuration of the target molecule. There are a few assumptions that need to be taken into consideration before utilizing Mosher ester analysis for determination of absolute configuration. MeO OH O * Ph MPA = methoxy phenyl acetic acid (I-4) (R)-MPA(I-4R) (S)-MPA (I-4S) OH 1' 3' I-3R 4' 5' H ΔδRS> 0 RO L2L1 ΔδRS< 0 ΔδRS = δ(R) - δ(S) MeO MeO H syn O H 1' 3' 5' 4' O s-trans H shielding I-3R-R syn O H 1' shielding 5' O s-trans 3' 4' I-3R-S Figure I-2. Assignment of the absolute configuration of a secondary alcohol using Mosher method. Structure of the stable conformer that leads to the change in the chemical shift of the derivatized alcohol and the 1H-NMR traces of both diastereomers, as well as the proposed mnemonic (dashed box) are demonstrated. As illustrated in Figure I-2, structural features of Mosher ester i.e. the s-trans conformation of the ester group, the syn alignment of the ester carbonyl with the smallest group on the chiral center as well as the anti-periplanar alignment of methoxy 5 C-O bond with ester C-O bond are crucial in order to conceive meaningful results from changes in the chemical shifts. Deviation from any of these assumptions causes discrepancies on the results of Mosher ester analysis.8-9 Although this method has been extensively used for determination of absolute configuration, the requirement of milligram quantity of chiral sample for chemical derivatization and the uncertainty observed in some of the results are the limitations of this method. Furthermore, Mosher ester analysis can only be performed on molecules that contain a single derivatization group. Another NMR based technique that can be used for the determination of absolute configuration is application of chiral solvating agents. In this method, the chiral solvating agent, non-covalently interacts with the target molecule and the anisotropic effect of the solvating agent on the substituents of the target molecule can be informational towards determination of the absolute configuration. Elimination of chemical reaction can be considered as advantage of these techniques as compared with the Mosher methods. However, lack of predictability and ambiguous results are by far the most notable limitations associated with the aforementioned methods for determination of the absolute configuration of chiral molecules.10-11 On the other end of the spectrum, X-ray crystallographic techniques present a different solution to this issue. X-ray structure can provide unambiguous determination of the absolute configuration of asymmetry. The necessity of high quality single crystal hampers the application of this technique as a routine procedure for determination of absolute configuration. Nonetheless, X-ray structure elucidation is one of the most 6 powerful techniques for determination of the relative stereochemistry of organic molecules. Application of chiroptical spectroscopy for determination of absolute I-3 configuration of chiral molecules Interaction between plane polarized light and chiral molecules (chiroptical spectroscopy) is one of the earliest observations made with asymmetric molecules. Chiroptical data, as a unique physical property of chiral molecules, are measured as early as mid-nineteenth century.1 Light waves consist of electric and magnetic vectors that oscillate perpendicular to one another and to the direction of the propagation of light. Unpolarized light in which the light waves propagate in all directions is the characteristic of most of the ordinary light sources. If an unpolarized light passes through a polarizing filter only small fraction of light waves which oscillate in parallel with the direction of the filter can make it through, thus leading to linearly polarized light (Figure I-3-a). Linearly polarized light is comprised of two circularly polarized light components i.e. left circularly polarized light (lcp) and right circularly polarized light (rcp). The phase and intensity of lcp and rcp components of light are the same, thus the projection of their combined amplitudes yields a straight line perpendicular to the propagation axis of light.1, 12 7 a. lcp E rcp b. lcp E rcp c. a/2 α ψ b/2 Figure I-3. a. Plane-polarized light composed of two circularly polarized components, lcp and rcp. As long as the phases of intensities of these two components remain the same, the resultant oscillating vector will be in the same plane and magnitude. b. If the rcp component become less intense (due to interaction with chiral media) the electric vector of light follows an elliptical path (elliptically polarized light, green oval). c. The degree of ellipticity is define as y, which is proportional to the circular dichroism. The rotation of ellipse (a) corresponds to optical rotation and ORD. the characteristics of the right and left components of light, such as velocity and through an achiral or racemic solution, When polarized light passes absorbance, undergo changes to the same extent. As a result, the oscillating plan of polarized light remains unchanged. On the other hand, when light passes through an absorbing optically active substance, the velocity and absorbance of the lcp and rcp components of incident polarized light undergo changes with different extents, leading to a Cotton effect that is named after the French scientist, Amie Cotton. Measurement of alteration of velocity of left and right components of light results in Optical Rotatory Dispersion.13 Optical rotatory dispersion is based on the difference in the reflective indices of the two components of light, thus it can be recorded over all wavelengths. A derivative of ORD that is commonly utilized by scientists is specific optical rotation ([a]D) that is indicative of the optical activity at a specific wavelength. Sodium D-line (589 nm) is the wavelength commonly used as reference of optical rotation leading to [a]D. 8 Furthermore, these two components of light are absorbed by chiral media to a different extent due to difference in absorption coefficient of lcp and rcp, eL ¹ eR. The difference De º eL – eR is called Circular Dichroism (CD). Recording differences in the absorption of lcp and rcp over wavelengths that the molecule absorbs light leads to a CD spectrum. CD spectra are similar to UV-vis spectra with the major difference that UV-vis is always a positive curve whereas the CD spectrum can be positive or negative depending on the chirality of the sample (Figure I-4). positive CD negative CD Δε Δε wavelength (nm) wavelength (nm) Figure I-4. Positive or negative CD spectra or Cotton effects. This unique property based on the interaction of polarized light with a chiral compound that leads to two distinct parameters,13 ORD and CD, has found increasing usage in the assignment of absolute configuration. It should be noted that ORD can be observed in all wavelengths since the technique originates from changes is reflective indices of lcp and rcp. On the other hand, CD can only be observed in the wavelengths that the molecule exhibits absorption. The application of chiroptical spectroscopy for determination of absolute configuration of chiral molecules is one of the most widely utilized methods. Measuring optical rotation can provide an empirical method for determination of absolute configuration. Comparison of the measured optical rotation (aD) with the reported value 9 for the exact same compound can give access to the absolute configuration of a molecule. Nonetheless, if the optical rotation of a molecule is not known, measurement of [a]D cannot give any information with respect to the absolute configuration of a given compound. With recent advancements in computational modeling, optical rotation of a chiral molecule can be calculated and upon comparison with the measured value a platform for determination of the absolute stereochemistry can be exploited.14-16 Optical rotatory dispersion13 is another tool that can be utilized for determination of absolute stereochemistry.17-19 This method needs to be paired with computationally extracted ORD spectra in order to provide a reference spectrum for comparison with the experimentally measure ORD. Flexibility of molecular structure and presence of close lying conformers can complicate the computational analysis and applicability of this technique. Another area where chiroptical techniques are utilized extensively for determination of chirality is application of methods based on circular dichroism (CD) spectroscopy.20 Among various aspects of circular dichroism spectroscopy, vibrational circular dichroism (VCD), based on absorption at the infrared region and electronic circular dichroism (ECD) for absorption at UV-Vis region have found widespread applications in assignment of the absolute configuration of chiral molecules. In the case of VCD, the experimental spectrum of the target molecule must be compared with the calculated VCD spectra for both of the enantiomers.21-25 This technique has shown promising utility for determination of chirality if the calculated spectra are performed with high-level 10 quantum chemical calculation. Due to weak absorption in VCD spectroscopy, long acquisition time and concentrated samples (large sample size) are usually required to access reliable results. framework upon light. Much akin interaction with polarized Electronic CD (ECD) originates from electronic transition in the UV-vis region within a chiral to VCD spectroscopy, this method can be merged with computational techniques to exploit a comparative tool for the assignment of absolute configuration. A unique variant of this technique is the Exciton Coupled Circular Dichroism (ECCD) pioneered by Harada and Nakanishi in 1980s.26-27 ECCD utilizes the excited state coupling of two or more close- lying and strongly absorbing chromophores embedded in the structure of a chiral molecule. Presence of similar chromophores or chromophores with close UV-vis absorption profiles is crucial for a strong exciton couplet. These chromophores can pre- exist or be introduced into the chiral molecules via derivatization. For ECCD to arise, the electric dipole transition moments of the close by chromophores should interact with each other, that would lead in a splitting of the excited state energies. An electric dipole transition moment (edtm, µ) can be described for each transition. The edtm for each transition defines the net displacement of charge during the transition.12 In most common systems that utilize ECCD, two chromophores are interacting with each other. In this paradigm, the transition dipoles can have in-phase or out-of-phase arrangements that result in two exciton states. The splitting energy of the two states (2V12) is referred to as Davydov or exciton splitting (Figure I-5). The relative arrangements of the interacting chromophores determine the magnitude and the energy of splitting. For 11 systems with two chromophores, a face-to-face, head-to-tail or oblique arrangement of dipoles can be described. In the face-to-face arrangement, interaction of the two chromophores leads to increase in the excitation state energy due to electrostatic attraction and repulsion, as a results a blue shift in absorption is predicted (Figure I-5-a). E G E G E monomer monomer a. face-to-face R12 µ1 µ1 y g r e n E a. head-to-tail R12 µ1 y g r µ1 e n E c. oblique µ1 R12 y g r e n E µ2 2V12 E" E' µnet µnet = 0 dimer 2V12 blue shift E" E' µnet = 0 µnet red shift dimer 2V12 E" E' µnet µnet band-splitting G monomer dimer Figure I-5. Schematic exciton band energy diagram of system with two interacting chromophores in various geometries. The ovals indicate chromophores with their electric dipole transition moments. The magnitude of the net electric dipole transition moments (µnet) is deduced by employing a simple vector model analysis. 12 This arrangement is also referred to as H-aggregation. In the case of head-to-tail arrangement (also known as J-aggregation) a red shift in absorption is expected since the electronic transition to the excited state with higher energy is dipole forbidden (Figure I-5-b). As illustrated in Figure I-5-c, if the two chromophores have an oblique arrangement, absorption to both of the excitation states will take place and a splitting of the absorption band into two components will be observed.12 This arrangement is the most general type of aggregation of chromophore that is observed commonly for interacting chromophores and can provide useful information with regards to molecular chirality. I-4 for determination of absolute configuration Chemical derivatization of chiral molecules with interacting chromophores The coupling of asymmetrically disposed chromophores leads to an interpretable signal which is the direct consequence of the helicity of the coupling transitions of the two chromophores. Since not every chiral compound has asymmetrically arranged strong absorbing chromophores in their skeleton, enormous efforts have been directed towards design and introduction of interacting chromophores in chiral molecules. One of the early approaches was to derivatize chiral molecules with two chromophores for the purpose of absolute stereochemical determination. In this regards, derivatization of chiral diols with 4-substituted benzoate as chromophores was envisioned (Figure I-6).28 The stereochemistry of the derivatized diol dictates the relative arrangement of the newly installed benzoate chromophores. 1,2- 13 diol derivatized with 4-substituted benzoate demonstrates the oblique alignment of interacting chromophore in a helical arrangement (Figure I-6). Coupling of the edtm of the benzoate chromophores (indicated with blue solid lines) leads to the observed bisignate CD spectrum. X X X X Me O O s-trans H O H syn O O O s-trans H O H syn O positive couplet Me Δε Α nm counter-clockwise helicity negative ECCD couplet clockwise helicity positive ECCD couplet Figure I-6. Schematic demonstration of two independently conjugated chromophores arranged in a helical twist. The electric dipole transition moments (blue line) are along the long axis of the benzoate moieties. The side view of interacting chromophores indicates clockwise or counter-clockwise arrangement of chromophores depending on the chirality of the diol. The clockwise helicity generates a positive ECCD spectrum (shown on tap right). The sign of the ECCD spectrum is denoted as positive if the positive peak of the couplet appears at higher wavelength (as shown in Figure I-6) or negative if the negative couplet appears at higher wavelength. The sign of the ECCD spectrum is the direct demonstration of the helicity of the interacting chromophores, which in turn originates form the absolute configuration of the chiral diol. Thus, a clockwise helicity of 14 interacting chromophores leads to a positive ECCD signal and counterclockwise helicity generates a negative ECCD spectrum, thus enabling a none-empirical determination of the orientation of chromophores.12, 26-27 Another approach for installation of chromophores into chiral molecules for determination of the absolute configuration has been reported by Rosini and coworkers.29-30 They utilized a bridged biphenyl system bearing a ketal moiety to form spiro dioxolanes with chiral 1,2- and 1,3-diols (Figure I-7). In this manner, the chirality of the diol induces axial chirality into the backbone of the bridge biphenyl system and leads to the appropriate ECCD response. The CD signal can be directly correlated into the absolute configuration of the derivatized diol. This method reveals a simple procedure for introducing absorbing chromophore into UV transparent diols in order to observe predicable CD signals that give access to the absolute stereochemical free rotation OMe OMe I-6 25 °C, MS OH * R2 OH * ( )n I-5 R1 H R H R H H O O H (R,R,P)-I-7 CD response R1 O ( )n O R2 I-7 H R H R H H O O H (R,R,M)-I-7 Figure I-7. Dioxolane formation of chiral diols with I-6 leads to CD active complexes. The highlighted diastereomeric complexes (box) depict equilibrating dioxolanes that favor M conformation due to the chirality of the bound diol. 15 determination. This backbone has been modified to be applicable for derivatization of chiral carboxylic acids (Figure I-8).2, 31 In the same paradigm, the chirality of the carboxylic acid induces a twist into the biaryl backbone of the azepine I-18 that leads to the observed CD signal. Figure I-8 (box) highlights the most stable conformation of the derivatized acid that features co-planarity of the smallest group on the chiral acid with the methyl moiety of the azepine in order to minimize 1,3- strain. P helicity is more stable for the shown configuration due to alleviation of steric repulsion between the large group on the chiral center and the azepine. R2 R1 R3 CO2H + 25 °C, MS (P) NH I-8 (P) O S N HH M L favored conformation (M) O R1 R3 R2 O R1 R3 R2 N I-9P N I-9M CD response Figure I-8. Development of an azepine probe (I-8) for sensing the absolute configuration of chiral carboxylic acids. In another elegant design, Wolf and coworkers introduced triaryl I-10 as a molecular probe for determination of the absolute configuration of amino alcohols (Figure I-9).32-36 This foldamer features two cofacial salicylaldehyde rings on a naphthalene backbone. They have shown that salicylaldehyde rings can freely rotate around the C-C bond of the naphthalene ring leading to C2-symmetric M,M-I-10 and P,P-I-10 rotamers that 16 interconvert through the meso form. Schiff base formation of the aldehyde moieties on the sensor’s backbone with chiral amino alcohols would result in a population imbalance between the M,M and P,P rotamers. The rotamer with minimized steric repulsion will be dominating and generates the corresponding CD signal. The intensity and sign of the CD signal can be used to elucidate the absolute configuration of the bound chiral molecules as well as the enantiomeric excess of the sample. O HO HO O O O HO OH O HO OH O HO H2N R H I-11 (P,P)-I-10 I-10 meso form (M,M)-I-10 OH NH2 R H I-11 R H HO OH HO OH N N R H R H OH HO N HO OH N R H CD response abolute configuration, enantiomeric composition (P,P,S,S)-enantiomer I-12 (M,M,R,R)-enantiomer I-12 Figure I-9. Development of stereodynamic molecular probe I-10 for determination of enantiomeric composition and absolute configuration of chiral amino alcohols through imine formation. Another example that utilizes three interacting chromophores for the determination of the absolute configuration of chiral molecules is reported by Canary and coworkers (Figure I-10). They take advantage of complexation of Zn(II) and Cu(II) with quinoline based ligands to arrange the chromophores around the metal center. 17 The chirality on linker to one of the quinoline ligands controls the helical arrangement of the ligands around metal center leading to a strong CD signal. In a similar manner, the helical twist of quinoline based ligands around Cu(II) is controlled by the stereochemistry of chiral methionine (Figure I-11).37-39 The R amino acid induces a P twist the ligands and generates positive CD couplet. On the other hand, the S enantiomer leads to M twist and a negative ECCD signal as output. side view top view N N X Cu N Me I-13 N H N N H Cu N Me Figure I-10. Structure of propeller-like Cu(II) complex with chirality on the backbone of one of the quinoline based ligand. Alternatively, in order to access a stereo-dynamic system, an achiral ligand can be used as a chromophore and chiral molecules can be introduced to the complex through ligand exchange with water molecule on the copper complex (Figure I-12).40-41 The chirality of the new coming ligand (chiral carboxylic acid) then dictates the helicity of the interacting chromophores. The helical twist of the ligands provides strong ECCD signal that is consequence of the absolute configuration of the bound chiral carboxylic acid. 18 X O N Cu N N N X O N Cu N O S I-14(S) (S)-methionine complex negative CD couplet I-14(R) (R)-methionine complex positive CD couplet O S Figure I-11. Induced helical twist of the achiral quinoline based ligand on Cu(II) due to stereochemistry of the bound amino acid (methionine). N N H2O Cu N I-15 CD inactive complex N Ph I-16 CO2H CD active complex positive CD couplet for complex with I-16S negative CD couplet for complex with I-16R Figure I-12. Structure of a copper based molecular probe for sensing the chirality of carboxylic acids. In another elegant strategy, Anslyn and coworkers described a dynamic covalent assembly between pyridine-2-carboxaldehyde, di(2-picolyl)amine, a chiral alcohol and zinc triflate that proceeds via an activated iminium intermediate I-19 (Figure I-13).42-44 The chirality of the alcohol determines the chirality of the newly formed N,O-acetal center in I-20 and consequently the helical twist of the pyridine ligands is the outcome of the latter stereo-induction. This multi-component association gives access to stereochemical determination as well as estimation of the enantiomeric excess of the chiral secondary alcohols. 19 + O N I-17 H N I-18 M + R*OH Zn(OTf)2 N TfO N I-19 N N P CD signal determination of absolute configuration and enantiomeric excess of secondary alcohols. N N Cu N OR* H *RO H N N Cu N N I-20M I-20P X Zn N OR* N N I-20 Figure I-13. Stereodynamic assembly of chiral alcohol with in situ generated iminium I-19 into a trigonal bipyramidal scaffold leading to P or M helicities depending on the chirality of alcohol. Porphyrin tweezers methodology: host:guest complex formation with chiral I-5 molecules for determination of absolute configuration Another strategy that is applicable for organic compounds that lack the presence of two strongly absorbing chromophores is host-guest complex formation of the chiral organic molecule (guest) and an achiral system containing two or more chromophores (host). In this approach, chiral molecules are not necessarily chemically derivatized with the chromophoric host systems, yet their chirality is transferred to interacting chromophores of the host system through metal coordination. The porphyrin tweezers methodology pioneered by Nakanishi and co-workers is one of the most utilized host system that has been applied to the assignment of absolute configuration of different families of organic molecules.45 In this approach, porphyrins as unique chromophores form host-gust complexes with chiral organic molecules. In the complex formation 20 event, the chirality of the bound organic compound leads to a helical twist of the porphyrin tweezer resulting in a detectable ECCD signal. The sign of the ECCD signal can be non-empirically correlated to the helical twist of the interacting porphyrin chromophores. As illustrated in Figure I-14, the host guest complexation of diamine I- 22, with stereogenic center on one of the amine groups with porphyrin tweezer I-21 gives rise to the latter helical arrangement of the two porphyrins. In order to minimize the steric repulsion of the complex, the porphyrin bound to the stereogenic nitrogen slides towards the small group on the chiral center and moves away from the larger group on this carbon. 21 a Ph Ph N N Zn N N Ph Ph Ph N N Zn N N Ph O O C5-TPP tweezer I-21 O O Zn Zn Zn Zn O O O O b SL H2N NH2 I-22 favored c Zn negative CD couplet I-23M C5-TPP tweezer Zn N H2 Zn N H2 100 50 0 -50 -100 350 O O I-23 O O unfavored Zn 400 450 wavelength (nm) 500 positive CD couplet I-23P Figure I-14. (a) Structure of C5-TPP tweezer; (b) model for complexation of chiral diamine with C5- TPP tweezer (a bidentate coordination of diamines with the porphyrin tweezer leads to the observed helical twist); (c) Experimentally measured ECCD signal and proposed model for the preferred helical twist (left), counter-clockwise arrangement of interacting dipole transition moments leads to a negative ECCD signal. 22 The latter arrangement leads to a counter-clockwise helical twist, and a negative ECCD signal for this complex is predicted. The tweezer methodology is competent to repot the chirality of host molecules with two bonding sites. This ECCD based approach has been applied to the absolute stereochemical deamination of a wide range of compound including polyols,46 carbohydrates,47-49 quinuclidines,50-51 amines,52-53 amino acids amino alcohols,38, 54 and hydroxyl acids.55-56 The requirement of two coordinating sites on the guest molecules preclude complex formation of the porphyrin twezeers with chiral molecules with only one binding site such as chiral mono-alcohol, mono-amines, carboxylic acids, etc. Nonetheless, this issue has been circumvented to some extent via derivatization of the chiral mono dentate molecules with a carrier that provides a second coordinating site. Following this approach, chiral alcohols, amines and carboxylic acids can be converted to their corresponding ester or amide derivatives prior to complexation with the porphyrin tweezer.57-61 The zinc centered porphyrin is Lewis acidic enough for coordination with amine containing chiral molecules, however hydroxyl groups suffer from low binding affinity for complex formation with the zinc porphyrin. Our group has included significant modifications to the porphyrin tweezer methodology, providing a platform for sensing the chirality of chiral molecules with hydroxyl groups as coordinating site. The Lewis acidity of the zinc center was greatly improved by employing perfluorinated phenyl rings substituted on the porphyrin meso positions. Zn-TPFP I-26 porphyrin tweezer is capable of ECCD active complex formation with chiral 1,2-diols,62 epoxy alcohols63 and distal diols64 for absolute stereochemical determination (Figure I-16). The computed HOMO 23 and LUMO energies of zinc-tetraphenyl porphyrin (Zn-TPP) and zincated perfluorinatedphenyl porphyrin (Zn-TPFP) confirmed the increased Lewis acidity of the metallocenter in the fluorinated porphyrin tweezer system as compared to the zinc in the non-fluorinated host systems I-15). These computational data were corroborated with experimentally measured binding constants for complexation of alcohols and amines with zinc porphyrins of various electronic profiles. The measured binding constants for Zn-TPFP are roughly 40 times higher than that for Zn-TPP for complexation with amines or alcohols (Figure I-15).62 (Figure Ar N N N Zn N Ar Ar PhF5 N N Zn N N F5Ph PhF5 F5Ph PhF5 N N Zn N N PhF5 Ar = C6H5, Zn-TPP-monoester I-24 Ar = C6F5, Zn-TPFP-monoester I-25 O O Me O O porphyrin chromophore ELUMO (eV) Mullikan charge electrostatice charge of Zn2+ O O C5-TPFP tweezer I-26 Kassoc (iPrOH) M-1 Kassoc (iPrNH2) M-1 of Zn2+ 0.961 0.942 1.387 1.337 2170 ± 140 49 ± 2 -2.818 -2.211 Zn-TPFP-monoester I-24 Zn-TPP-monoester I-25 Figure I-15. Frontier orbital energy and charges of zinc porphyrins. Bonding affinity of iso-propanol and iso-propylamine with Zn-TPP-monoester and Zn-TPFP-monoester as analogues of Zn-TPP tweezer and Zn-TPFP tweezer (dashes box depicts structure of C5-Zn-TPFP tweezer). The development of TPFP tweezer methodology provided a unique platform for assignment of absolute configuration of many classes of organic molecules. The 473000 ± 8700 11400 ± 950 24 following pictorial summary illustrates our major contributions to this area (Figure I- 16).62-67 α-chiral carboxylic acids 1. EDC, DMAP carrier erythro and threo diols, amino alcohols and diamines HO O iPr H S CH3 O HO R X X = F, Cl, Br H H2N NHBoc H2N 2. TFA Zn-TPP-C5 and Zn-TPP-C3 Org. Lett. 2002, 4, 3423-3426. chiral α-halo carboxylic acids H2N installation of the carrier Zn-TPP-C5 and Zn-TPP-C3 Eur. J. Org. Chem. 2009, 4242-4253. O iPr H S CH3 N H O H N H R X O HO remote chirality, β−, γ−, δ−chiral carboxylic acids R * installation of the carrier Zn-TBP-C5 and Zn-TBP-C3 H2N R n N H Angew Chem. Int. Ed. 2015, 54, 4274-4278 O R * R' n R1 R1 OH OH NH2 R2 HO Fluorinated Porphyrin Tweezer (Zn-TPP-C5) H2N H2N R2 R2 R1 * * * * * * J. Am. Chem. Soc. 2008, 130, 1885-1893 chiral epoxy Alcohols Fluorinated Porphyrin Tweezer (Zn-TPP-C5) J. Am. Chem. Soc. 2008, 130, 16126-16127. long-range 1,n-Glycols R2 O * * R1 R3 OH OH OH * n = 0 - 14 * n Zn-TPFP-C5 and Zn-TPFP-C3 J. Am. Chem. Soc. 2012, 134, 9026-9029. Ar N N Zn NN Ar Ar N N Zn NN Ar Ar Ar O O O n O porphyrin tweezers: n = 1 : Zn-TPP-C3 (Ar = Ph) I-27 n = 3 : Zn-TPP-C5 (Ar = Ph) I-21 n = 1 : Zn-TBP-C3 (Ar = 3, 5-di-tBuPh) I-28 n = 3 : Zn-TBP-C5 (Ar = 3, 5-di-tBuPh) I-29 fluorinated porphyrin tweezers: n = 1 : Zn-TPFP-C3 (Ar = F5Ph) I-30 n = 3 : Zn-TPFP-C5 (Ar = F5Ph) I-26 Figure I-16. Pictorial representation of functional group families with methods for assignment of their stereochemistry. Chiral molecules with single site are derivatized with a carrier to form a bidentate guest molecule (left panel) prior to complex formation with tweezers; Structure of host systems, with various linker size as well as tuned steric and electronic profiles that enables determination of absolute configuration of the latter functionalities are shown at the bottom of this figure. Modifications of porphyrin tweezer, in terms of electronic modulation of the host system as well as steric adjustments and linker size variations, have led to successful 25 application of this methodology for the determination of the absolute configuration of the represented functionalities. Nevertheless, the major limitation associated with the porphyrin tweezer methodology, as illustrated in Figure I-16, is the prerequisite of needing two coordinating sites on the chiral molecule to induce a helical arrangement of the interacting porphyrin chromophores. Designing a host system that eliminate the requirement of two binding site that limits the application of this methodology, became our group’s focus. Design of a new host system for determination of absolute configuration of I-6 chiral molecules with one sites of attachment As described above, application of the porphyrin tweezers methodology, with all its potential, is limited to molecules with two coordination sites. If a molecule with sing site of attachment is to be analyzed via porphyrin tweezers, a drivatization step is necessary that implement a second binding site in order to derive the complex formation. As derivatization is not desirable it also cannot be implemented to many classes of molecules with one site of attachment. We set out to design a new host system to remedy this shortcoming. The envisioned host system should be capable of forming ECCD active complexes with chiral molecules with only one site of attachment without the necessity for derivatization. 26 chromophore structural modification Linker adjustment chromophore O O ( )1, 3 O O porphyrin tweezer host system chro m ophore free rotation chromophore structural modification binding inclusion site chro m ophore OH OH binding site chromophore new host system with change in binding paradigm Figure I-17. Rational design of a new host system capable of sensing chirality of molecules with one site of attachment. Linker modification and change in the binding paradigm are notable adjustments. Approaching this problem from a rational design perspective, we realized that the porphyrin tweezer methodology with its flexible linker (alkyl chain as linker) necessitates the concurrent coordination of two sites form a guest molecule to the metalloporphyrin in order to induce helical twist and generate CD active complexes. Our approach was to design a molecular framework that places the porphyrin moieties in a face-to-face arrangement even in the absence of a chiral molecules. As such, the envisioned system would be helical, but racemic, and can adopt either P or M helicity. The chiral molecule would then solely be responsible to induce a preference over the existing and equilibrating helicities (Figure I-17). To this end, a 2,2’-biphenol functionality was envisioned as a backbone for the interacting chromophoric porphyrin rings (Figure I-17). 2,2’-Biphenol systems can exist as equal P and M atropisomers due to the free rotation around the bi-aryl C-C bond (in the absence of bulky substituent at the 6,6’ positions). In this hypothetical design the porphyrin ring would be placed on the 3,3’ position of the biphenol core. The free 27 rotation along the axial chirality of the biphenyl moiety leads to equally populated P an M atropisomers.68 The P atrop-isomer lead to a clockwise helical twist of the chromophoric porphyrin rings, yielding to a positive ECCD signal, while a negative ECCD signal originates from the M helicity. In the absence of a chiral guest, the host molecule is ECCD silent since the population of the equlibrating helical twists (P and M) leading to opposite ECCD signals is equal. Upon introduction of a chiral molecule that can form a stable complex with the host system, an imbalance in the population of the P and M helicities is anticipated. As a result, the interaction between the host system and a chiral molecule, the point chirality of the chiral guest molecule is transferred to the axial chirality of the host system. Thus, the favored atrop-isomer with higher population will result in a ECCD signal. Helical chirality reporters (through ECCD) Ph N Ph Ph N H NH N Ph N H N H N binding site OH OH Ph N Ph Axial chirality: can adopt P or M helicity depending on chirality of host MAPOL I-31 Figure I-18. Structure of newly designed host system. Biphenyl back bone, capable of adopting P or M helicity, renders equilibrating atropisomers. Upon introduction of chiral guest, that rests in the highlighted binding sites, one of the helicities become more prominent leading to the corresponding ECCD signal. 28 MAPOL (I-31)68 was envisioned as the potential host molecule able to deliver on the latter prerequisites (Figure I-18). The synthesis of this molecule was the first challenge to overcome. Typically, synthesis of mono-porphyrin compounds proceeds with average yield of 10-20%. For our target molecule with two porphyrin rings embedded in the structure of the host system, the theoretical yield was not expected to be high. Many different routes for the synthesis of MAPOL were screened. Eventually a disconnective approach was chosen to forge the porphyrin moieties. 1. nBuLi, TMEDA -78 °C to rt, 2.5 h 2. DMF, -78 °C 63% O O I-32 CHO O O CHO I-33 InCl3 Pyrrole 83% 1. NaBH4, THF/MeOH 2. I-34, MeCN, TFA 3. DDQ, 8-10% NH HN O I-35 O I-34 N HN Ph Ph Ph NH N OR OR N NH HN N O O NH H N N H NH Ph Ph Ph BBr3, CH2Cl2 rt, 2 h, 85% Figure I-19. Successful synthesis of MAPOL I-31. 29 I-36, R = CH3 I-31, R = H As depicted in Figure I-18, the synthesis of MAPOL commenced with commercially available 2,2’-bisphenol I-32 that was underwent a directed ortho formylation leading to the bisaldehyde I-33. Condensation of this bisaldehyde with pyrrole in the presence of InCl3 led to the formation of dipyrromethane derivative I-34. To prepared the other half of the porphyrin rings, benzaldehyde was condensed with pyrrole followed by dibenzylation to give I-35. These two components (I-34 and I-35) were married together through a 2+2 porphyrin formation reaction. Diketone I-35 was reduced to its corresponding diol that was directly reacted with dipyrromethane I-34 in the presence of TFA as a Lewis acid in acetonitrile. Oxidation of the condensed product with DDQ furnished the porphyrin ring to yield I-36. The dimethyl MAPOL I-36 was readily isolated in a relatively good yield of ~10% considering two porphyrin rings were assembled in one step. The ease of isolation was the major advantage of this synthesis. Since the only possible porphyrin in this reaction is the expected product, the higher order oligomers were easily removed form organic products with a simple filtration. MAPOL was obtained via demethylation of I-36 in the presence of BBr3. 30 I-7 Determination of the absolute configuration of chiral monoamines With MAPOL in hand, we set out to test the applicability of this host system with chiral monoamines as proof of concept. The classical porphyrin tweezers were incompetent to form an ECCD active complex with monoamines due to lack of a second binding element. In this newly designed host system a change in the binding paradigm was implemented. As such, MAPOL, bearing a biphenol core, was envisioned to form a hydrogen bond network with the amino group of chiral monoamines. This binding mode places the chiral carbon in the center of the host molecule, forcing interactions with the pseudo face-to-face arranged porphyrins. Two diastereomeric complexes are thus possible, where the fixed chirality of chiral amine (R or S) induces the flexible axial chirality of the host system (P or M). Ph NH N OH OH N HN Ph Ph N NH HN N Ph NH2 I-37S (20 equiv) Ph Ph MAPOL I-31 (1 µM, in hexane) Figure I-20. Complexation of MAPOL with chiral monoamine II-37S leads to a strong ECCD signal with a negative sign. 31 The equilibrating diastereomers are expected to have different energies, with the more stable complex that has alleviated steric strain, having a higher population. The observed ECCD signal would be the result of the higher populated helicity. With this hypothetical analogy, chiral amine II-37S was complex with MAPOL in hexane (Figure I- 20). Gratifyingly, the complex was CD active and produced a negative ECCD signal. The generation of ECCD signal confirms the formation of a stable chiral host:guest complex. To further verify the complex formation between chiral amine and MAPOL, the complexation was followed by 1H-NMR. Addition of amine I-37S to MAPOL in chloroform showed dramatic changes in the chemical shift of the guest molecule along with notable changes of the protons on the host molecule. In brief, protons on the chiral amine I-37S undergo significant up field shifts at a lower host:guest ratio (Figure I-21). Upon increasing the host:guest ratio, the chemical shifts of the amine protons increase towards those of the free amine. At low equivalents of free amine, most of the guest molecules are in the cavity of MAPOL, thus experiencing strong anisotropic effect of the porphyrin rings that results in up-field shift of the amine’s proton resonances. At higher equivalents of the amine however, the chemical shifts are the average chemical shift of the bound and free amine. The region highlighted with blue background indicates the changes in the chemical shifts of the methyl group on the chiral amine I-37S. This methyl group in the free amine appears at ~1.0 ppm, while in the 1:4 complexation ratio with MAPOL (0.25 equiv) is located at ~0.0 ppm. 32 H H Porphyrin OH OH Porphyrin I-31 S-(+)-1-cyclohexylethylamine II-37S NH2 Me I-37S Figure I-21. 1H-NMR spectra of titration of MAPOL 1 with S-(+)-1-cyclohexylethylamine I-37S. Highlighted with blue indicates the changes in the chemical shift of the methyl group of the amine substrate and the red area shows the changes in the chemical shift of the MAPOL backbone upon complexation with amine I-37S. Job’s plot analysis of the changes in the chemical shift of both amine and MAPOL as function of the equivalents of added amine clearly indicates a 1:1 stoichiometry of the complex (Figure I-21). The fact that following the chemical shift changes of either MAPOL I-31 or amine I-37S in the 1H-NMR titration leads to the same binding stoichiometry is self-consistent and supports the proposed binding paradigm. 33 Figure I-22. Job’s continuous plot of MAPOL I-31 with I-37S: following the changes in chemical shift of MAPOL at 7.55 ppm (left) and chemical shift of amine I-37S at 0.99 ppm. The generality of the host system for complex formation with a variety of chiral monoamines was tested. Of note is the opposite ECCD signals for enantiomeric amines that points to the fact that one enantiomer of the guest molecule favors M helicity and the opposite enantiomer favors P helicity. To correlate the chirality of the bound amine with the helicity of the preferred atrop-isomer of MAPOL, the following mnemonic is proposed. The binding element that derives the complex formation is hydrogen bonding between amine group of the guest molecule and the hydroxyl groups of the host system. This binding paradigm forces the amine group to point towards the center of MAPOL and places the stereogenic carbon in the cavity of MAPOL. Minimization of the steric repulsion between groups on the asymmetric carbon and the porphyrin rings were thought to be the determinant factor for the preferred helicity. In Figure I-22, the S enantiomer of chiral amine I-37S is complexed with MAPOL. The groups attached to the chiral center have different sizes. These groups can be generally classified as small (hydrogen), medium (methyl) and large (cyclohexyl) based on their relative size. 34 a H Me (S) NH2 I-37S (20 equiv) b Ph N HN Ph H Ph NNH H O ECCD measurement MAPOL (1 µM) Hexane, 0 °C Ph Ph Me NNH NHN Ph S NH H O H P-helicity positive ECCD c Ph NHN Ph H Ph NNH Me S NH H H O H O Ph Ph NNH NHN Ph M-helicity negative ECCD D C . l o M 300 200 100 0 -100 -200 -300 negative ECCD M helicity 350 400 450 Wavelength (nm) 500 top view H Cy Me P-(R) complex disfavored top view Cy H Me M-(R) complex favored Figure I-23. a) Complexation of I-37S with MAPOL leads to a strong negative ECCD signal. b) Binding of amine I-37S, with the P-helicity of MAPOL leads to steric clash of the methyl group with the porphyrin ring. Thus, positive ECCD with I-37S bound to MAPOL is not anticipated. c) The same binding of I-37S with the M-helicity of MAPOL places the medium size methyl group in the most sterically available position. This favorable arrangement leads to the anticipated positive ECCD spectrum. 35 A-value (or other steric parameters) can be used for assignment of the relative size of the substituents. In the case of chiral monoamines with MAPOL we sorted the substituents on the stereogenic center based on their A-values. Both possible stereogenic complexes of the chiral amine with MAPOL (S-M and S-P diastereomeric complexes) will be considered for relative stability (Figure I-23, b versus c). Among the substituents attached to the chiral center, the cyclohexyl group is the most sterically demanding group, thus occupying the most sterically accessible position. Due to the tetrahedral geometry of the chiral carbon, the remaining two groups (medium and small) will be projected towards specific quadrants. Considering the P helicity, the medium group points towards one of the porphyrin rings, while in the M helicity the smaller group is placed in the same position. This simple analysis reveals that the M helicity is more stable than P helicity. As a result, the sign of the ECCD signal is expected for this favored complex. Indeed, the experimental measurements corroborated the negative ECCD signal. The same analogy can be utilized for all of the amines, with the measured ECCD signal in full agreement with the predicted sign obtained from the proposed mnemonic. Furthermore, the measurements are in micro scale regime and assessment of absolute configuration can be obtained in minutes, which adds to the practicality of this technique. In summary, we have designed a new host system that can give access to the absolute configuration of chiral monoamines in a non-empirical, micro scale and rapid fashion. In this new design, chiral monoamines form complexes with MAPOL as the host system through hydrogen bonding. Metal-free porphyrin rings act solely as 36 chromophores and reporter of helicity as opposed to the porphyrin tweezers methodology where the porphyrin rings are in fact the binding site as well as the interacting chromophores. This design holds promises for assignment of absolute configuration of other mono-dentate chiral molecules. to deliver methods 37 I-8 Experimental section Job’s continuous plot analysis to determine complex stoichiometry 1-8-1 The complex stoichiometry for the host-guest complexes were determined through 1H-NMR titration of MAPOL (host) with chiral amines S-(+)-1-cyclohexylethylamine I- 37S as the guest molecule. The following describes the experimental detail for binding titration of I-37S with MAPOL. Upon complexation of amine I-37S with MAPOL, 1H-NMR spectra of MAPOL as well as amine undergoes changes in chemical shift (Dd) due to the large anisotropic effect of the porphyrin rings. These changes in chemical shifts can be used as a read-out for determining the composition of host:guest complexation in solution. For titration, MAPOL (5.0 mg, 0.004 mmol) was dissolved in CDCl3 (1 mL), and the NMR spectrum was recorded (tert-butyl methyl ether was used as internal standard). Following this, 0.1, 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 5, … up to 100 equivalents of amine [dissolved in CDCl3 (0.1M amine stock solution for up to 2 equivalents and 1M amine stock solution for 3 equivalents and above)] was added to the NMR tube; the NMR spectra was recorded after each addition. Job’s plot based on changes in chemical shift of MAPOL (DdMAPOL), was obtained with respect to changes in chemical shift (Dd) of one of the MAPOL protons (peak at 7.55 ppm). The molar fraction of MAPOL (XMAPOL) was multiplied by the changes in 1H- NMR chemical shift of MAPOL (DdMAPOL) for each titration data point. The resulting value was plotted against the molar fraction of MAPOL. Peaking at 0.5 mol fraction corresponds to a 1:1 host:guest complex. 38 Alternatively, Job’s plot analysis based on the chemical shift changes of the amine (Ddamine) was pursued. The chemical shift changes of the methyl peak (doublet centered at 0.99 ppm) of amine I-37S, was chosen for analysis. Changes in chemical shift of the methyl (Ddamine) was multiplied the by mole fraction of amine (Xamine) at each titration data point and the result was plotted against the mole fraction of amine (Xamine). 39 I-8-2 Synthesis of MAPOL I-31: NH HN O I-35 O 1,9-Bis(benzoyl)-5-phenyldipyrromethane I-35 was prepared according to previously reported procedures.69 1H-NMR (CDCl3, 500 MHz); d 5.66 (s, 1H), 5.98 (m, 1H), 6.58 (m, 2H), 7.32 (t, J = 7.5 Hz, 1H), 7.38 (t, J = 7.5 Hz, 6H), 7.48 (m, 4H), 7.76 (d, J = 7.5 Hz, 4H), 11.12 (br 2H) ppm. 13C-NMR (CDCl3, 125 MHz); δ 44.9, 111.2, 120.9, 127.3, 127.9, 128.7, 128.8, 129.6, 130.9, 131.5, 138.1, 140.3, 141.2, 184.4. TOF MS ES+ (C29H22N2O2): Calc. [M + H]+: 431.1760, Found [M + H]+: 431.1757. CHO O O CHO I-33 Synthesis of 2,2'-simethoxy-[1,1'-biphenyl]-3,3'-dicarbaldehyde I-33: Freshly distilled ether (85 mL) was added to a flame-dried round bottom flask equipped with a magnetic stirring bar, and cooled to -78 °C. To this flask was added 2,2’-dimethoxy-biphenyl I-32 (3.0 g, 14.0 mmol), n-BuLi (2.5 M solution in hexane, 16.8 40 mL, 42 mmol, 3.0 equiv) and TMEDA (4.8 mL, 42 mmol, 3.0 equiv) dropwise. The reaction mixture was kept at -78 °C for 5 min, and then allowed to warm to room temperature, and stirred for an additional 2.5 h. The solution was once again cooled to - 78 °C, and DMF (6.2 mL, 80 mmol) was added slowly over a 10 min period. The reaction mixture was allowed to warm to room temperature, and was immediately quenched with aqueous NH4Cl solution. The organic layer was extracted with dichloromethane, and solvents were removed under reduced pressure. The crude mixture was purified by column chromatography (silica gel, 10% - 15% EtOAc in hexane) to give I-33 (2.4 g, 63%) as a white solid. 1H-NMR (CDCl3, 300 MHz); d 3.56 (s, 6H), 7.30 (t J = 7.5, 2H), 7.61 (dd, J1 = 7.0 Hz J2 = 1.5 Hz, 2H), 7.89 (dd, J1 = 7.5 Hz J2 = 1.5 Hz, 2H), 10.43 (s, 2H). 13C-NMR (CDCl3, 75 MHz); d 63.1, 124.4, 128.7, 129.5, 131.5, 137.5, 160.9, 189.8. TOF MS ES+ (C16H14O4): Calc. [M + H]+: 271.0970, Found [M + H]+: 271.0967. NH H N O O I-34 N H NH Synthesis of 2,2'-dimethoxy-3,3'-dipyrromethane-1,1'-biphenyl I-34: Following a standard procedure,70 bis-aldehyde I-33 (0.54 g, 2.0 mmol) was dissolved in pyrrole (27.0 mL, 400 mmol). After degassing with a stream of argon for 10 41 min, InCl3 (0.40 mmol, 88 mg) was added. The mixture was stirred for 1.5 h at room temperature under atmosphere of argon. The reaction was quenched by addition of NaOH (480 mg, 12.0 mmol) with stirring for an additional 45 min. The reaction mixture was filtered by a Buchner funnel to remove insoluble byproducts. The excess pyrrole was recovered using vacuum distillation. The viscous residue was triturated with hexanes (2 x 15 mL), and the volatiles were evaporated using rotary evaporator. The resultant chromatography (hexanes/dichloromethane/ethyl acetate = 7:2:1). The product was isolated as a light subjected column to yellow solid was yellow solid (840 mg, 83%). 1H-NMR (500 MHz, CDCl3); δ 2.97 (s, 3H), 5.79 (s, 2H), 5.89 (m, 4H), 6.11 (dd, J1 = 6.0 Hz, J2 = 3.0 Hz, 4H), 6.67 (m, 4H), 7.08 (dd, J1 = 8.0 Hz, J2 = 7.0 Hz, 2H), 7.15 (dd, J1 = 8.0 Hz, J2 = 2.0 Hz, 2H), 7.27 (dd, J1 = 7.0 Hz, J2 = 2.0 Hz, 2H), 8.27 (br, 4H, NH). 13C- NMR (125 MHz, CDCl3); δ 155.1, 136.1, 132.1, 130.4, 129.3, 124.0, 116.8, 108.3, 106.8, 60.4, 39.0. TOF MS ES+ (C32H30N4O2): Calc. [M + H]+: 503.2447, Found [M + H]+: 503.2444. 42 Ph NH N OMe OMe N HN Ph Ph N NH HN N Ph Methoxy-MAPOL I-36 Ph Ph Synthesis of Methoxy-MAPOL I-36: Following a standard procedure,71 to a solution of freshly prepared 1,9-bis(benzoyl)- 5-phenyldipyrromethane I-35 (0.43 g, 1.0 mmol) in dry THF/methanol (10:1, 44 mL) was added NaBH4 (766 mg, 20.0 mmol) in several portions. After completion of the reduction (TLC 40% EtOAc/Hexanes), saturated NH4Cl (70 mL) and CH2Cl2 (130 mL) were added to the reaction mixture. The organic layer was separated, washed twice with water, dried (Na2SO4), and placed in a 1000-mL round-bottomed flask. Removal of the solvent by rotary evaporation under vacuum yielded the dicarbinol as a foamlike yellow solid. To this crude product, dipyrromethane I-34 (0.23 g, 0.50 mmol) and acetonitrile (400 mL) were added, and the mixture was stirred to achieve a homogeneous yellow solution. TFA (0.93 mL, 12.2 mmol) was added with rapid stirring. After 5 min, DDQ (0.67 g, 3.7 mmol) was added. After stirring for 1 h at room temperature, triethylamine (1.69 mL, 12.2 mmol) was added, and the entire reaction mixture was filtered through a pad of alumina (4 x 8 cm) and washed with CH2Cl2 (~ 500 mL) until the eluant was no longer colored. The resulting porphyrin solution was concentrated, redissolved in CH2Cl2 (50 43 mL), and passed through a pad of silica (4 x 8 cm) with CH2Cl2 to remove nonporphyrinic pigments. The purple fractions were combined and concentrated to give a purple solid. Precipitation from CH2Cl2/methanol gave the pure product I-36 (62 mg, 9.6%). 1H-NMR (500 MHz, CDCl3); δ -2.68 (br, 4H), 3.07 (s, 6H), 7.64 (dd, J1 = 8.0 Hz, J2 = 7.0 Hz, 2H), 7.70-7.85 (m, 18 H), 8.16-8.20 (m, 4H), 8.25-8.31 (m, 12H), 8.93 (m, 12H), 9.08 (m, 4H). 13C-NMR (CDCl3); δ 61.1, 115.9, 120.0, 120.4, 122.0, 126.62, 126.66, 127.6, 127.7, 132.0, 132.3, 134.4, 134.5, 135.3, 135.9, 142.0, 142.2, 158.2. TOF MS ES+ (C90H62N8O2): Calc. [M + H]+: 1287.5074, Found [M + H]+: 1287.5117. Ph NH N OH OH N HN Ph Ph N NH HN N Ph MAPOL I-31 Ph Ph Synthesis of MAPOL I-31: To a sample of methoxy MAPOL I-36 (50 mg, 0.038 mmol) in dry methylene chloride (100 mL) was added BBr3 (1.0 M solution in dichloromethane, 0.2 mmol, 0.2 mL) drop-wise under atmosphere of argon at 0 ℃. The reaction was stirred at room temperature for 2 h before it was quenched with saturated solution of ammonium 44 bicarbonate (NH4HCO3). The reaction was extracted with methylene chloride, dried (Na2SO4), passed through a pad of silica and solvent removed to give a purple solid, which was purified further by precipitation from CH2Cl2/MeOH to give the pure MAPOL I-31 (42 mg, 85%). 1H-NMR (500 MHz, CDCl3); δ -2.78 (br, 4H, NH), 6.12 (s, 2H, OH), 7.57 (m, 6H), 7.66 (m, 8H), 7.70-7,75 (m, 6H), 8.06 (d, J = 7.0 Hz, 4H), 8.10-8.13 (m, 8H), 8.18 (d, J = 7.0 Hz, 4H), 8.74 (d, J = 4.5 Hz, 4H), 8.78-8.81 (dd, J1 = 11.5 Hz, J2 = 4.5 Hz), 8.91 (d, J = 4.5 Hz, 4H). 13C-NMR (125 MHz, CDCl3); δ 112.7, 120.2, 120.3 120.8, 125.8, 126.6, 126.6, 127.6, 127.7, 129.7, 132.7, 134.5, 134.9, 141.8, 142.0, 152.7); IR (film) 3319, 2924, 2853, 1716, 1473, 1441, 1350, 1219, 1176, 1072, 966, 800, 731. TOF MS ES+ (C88H58N8O2): Calc. [M + H]+: 1259.4761, Found [M + H]+: 1259.4816. 45 REFERENCES 46 REFERENCES 1. Berova, N. N., K.; Woody, R., Circular Dichroism Principles and Applications. Wiley-VCH: 2000. 2. 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Journal of Organic Chemistry 2000, 65 (22), 7323-7344. 53 Chapter II: Determination of absolute configuration of chiral molecules (chiral cyanohydrins, sulfoxides, phosphine oxides and alcohols) 54 II-1 Determination of absolute configuration of chiral cyanohydrins With the successful utilization of MAPOL for determining the absolute configuration of chiral monoamines,1 we sought to expand the applicability of this host system to determine the absolute configuration of other functionalities. Specifically, we set out to target tweezers methodologies, have failed to provide a robust and reliable means for determination of absolute configuration. For this purpose, chiral cyanohydrins were selected as a target functionality. Chiral cyanohydrins are versatile intermediates in organic synthesis since techniques, specifically porphyrin functionalities that chiroptical they can be easily converted into a variety of other chiral functional groups such as a- O R O R1 O OH O R OH NH2 CN R O R CN OH R1 n N Cbz n N Cbz OH R H2N OTMS H R1 O R1 O R OH O O OH R O R1 OH H N R1 R O O S R1 O CN R OH R CN F R CN Figure II-1. Synthetic utility of cyanohydrin moiety. 55 hydroxyacids, a-hydroxyketones, hydroxyamines, a-aminonitriles, and a-hydroxy esters (Figure II-1).3-4 Chiral cyanohydrins can be readily accessed through asymmetric addition of cyanide to aldehydes and ketones. Over the past few decades many different catalytic systems have been reported for the synthesis of this chiral moiety. Despite enormous advancements in the synthesis of chiral cyanohydrins, a direct, general and practical method for the absolute stereochemical determination of this functionality is lacking. The current state of the art in the determination of absolute configuration of chiral cyanohydrins is based on NMR analysis of their diastereomeric Mosher ester derivatives.5-8 Following typical assumptions in the Mosher ester analysis, models for the most stable conformer of the diastereomeric derivatives are utilized to can explain the chemical shift changes and provide a route for the assignment of stereochemistry. For cyanohydrins derived from aldehydes, the co-planarity of the hydrogen atom on the chiral carbon with the carbonyl group of the Mosher ester is the key feature in the proposed a. Cyanohydrins derived form aldehydes MeO O H O R CN H O H O R CN MeO H (S)-MPA ΔδRS> 0 (1H-NMR) H R CN O R ΔδRS< 0 (13C-NMR) b. Cyanohydrins derived form ketones CN O (R)-MPA MeO O R H R2 R1 ΔδRS< 0 (1H-NMR) O CN O R2 R1 S MeO H ΔδRS> 0 (1H-NMR) Figure II-2. Mosher’s method determination of absolute configuration of chiral cyanohydrins. 56 model. With this alignment and the Mosher method assumption of anti-alignment of the methoxy group and the C-O ester bond of the MPA acid, the groups on the chiral carbon either will eclipse the phenyl group of the Mosher ester or be on the opposite face. The anisotropic effect of the phenyl group results in shielding of the nuclei that are eclipse (Figure II-2). Interpretation of the shielding or deshielding of the substituent provides the absolute configuration of the chiral cyanohydrin center. In this particular case the effect of the Mosher ester on the 1H-NMR of the group attached to the cyanohydrin center as well as the changes on the 13C-NMR of the nitrile group are used for assignment of absolute configuration. The same analogy can be used for cyanohydrins derived from ketones. In these substrates, upon derivatization with Mosher acid, the nitrile group as the smallest group on the stereogenic carbon, will be co-planer with the ester carbonyl. In cyanohydrins derived from ketones, change in the 1H-NMR of both substituents attached to the chiral center will be used as a probe for the determination of absolute configuration. These techniques rely heavily on predicting conformational distribution of populations that affect the resultant NMR spectra. Another NMR-based approach for determining the absolute configuration of chiral cyanohydrins is the use of chiral solvating agents.9-10 Chiral solvating agents function non- covalently, i.e. via hydrogen bonding, interact with chiral cyanohydrins. Similar to Mosher ester analysis, the anisotropic effect of the substituents on the chiral solvating agent causes a shift in the 1H-NMR of the groups on the chiral cyanohydrin. The absolute configuration of cyanohydrins can be assessed utilizing the proposed models (Figure II- 3). The advantage of chiral solvating agents over Mosher method is the elimination of the 57 derivatization step. Nonetheless, the non-covalent interaction between solvating agent and the guest molecule can complicate the results as the nature of interactions is heavily depending on the structure of the target molecule, solvent composition, temperature, etc. H −δ O O H H S +δ N −δ O H O R R H N H −δ O O H H S +δ N −δ O H O S H R N +δ N +δ N Figure II-3. Application of chiral solvating agents for determining the absolute configuration of chiral cyanohydrins. On the other hand, chiroptical techniques, despite their intrinsic sensitivity to asymmetry as well as their propensity for rapid and micro-scale analysis, have not provided a routine method for the analysis of this functional group. The main shortcoming in this area has been devising a host system that can form stable and ECCD active complexes with chiral cyanohydrins. We set out to remedy the shortcoming in this area utilizing our MAPOL-based host:guest complexation methodology. This host system is capable of sensing the chiral information from the bound guest molecules and function as a reporter of chirality. Our goal was set to sense the absolute configuration of chiral cyanohydrins in a reliable, predictable and readable fashion. In addition, we planned to take advantage of the high sensitivity of the chiroptical techniques to devise a rapid, microscale and non-empirical methodology for assignment of absolute configuration of cyanohydrins. 58 At first glance, the cyanohydrin moiety has two potential binding sites, the hydroxyl and the nitrile groups, that are located on the chiral carbon with tetrahedral geometry. Our initial approach was to exploit both of these potential binding sites to drive a complex formation with perfluorinated porphyrin tweezers. As such, porphyrin tweezers such as C3-Zn-TPFP-tz (II-1) and C5-Zn-TPFP-tz (II-2)11 were complexed with the synthesized chiral cyanohydrin II-6R (Figure II-4). OH CN II-6R PhF5 = F F F F F PhF5 N N Zn NN F5Ph sites of attachment PhF5 F5Ph PhF5 N N Zn NN PhF5 OO n O O n = 3, Zn-TPFP-C5-tz II-1 n = 1, Zn-TPFP-C3-tz II-2 Figure II-4. Porphyrin tweezers and their envisioned complexation with chiral cyanohydrins for determination of absolute configuration. Not surprisingly, under various complexation conditions, no ECCD signal was observed. This can be attributed to multiple reasons, with employing chiral cyanohydrins as a bidentate guest molecule. The fact that both of the coordinating sites (nitrile and hydroxyl group) are located on the same tetrahedral carbon might preclude concurrent coordination of both groups to the zinc of the host system. Furthermore, Zn-TPFP porphyrins are Lewis acidic enough for coordination of oxygen based nucleophiles, 59 however the nitrile group might be a weak coordinating group due to the sp hybridization on the nitrogen, thus a concurrent binding might not be a viable scenario. Based on the latter results, we sought to consider only the hydroxyl group as a mono- dentate binding partner. Our approach was to use MAPOL (II-3) as a host system to drive complex formation with cyanohydrins. As we have shown previously, MAPOL was competent to form stable and ECCD active host:guest complex with chiral monoamines via hydrogen bonding. We envisaged a similar hydrogen bonding network forming between the hydroxyl group of cyanohydrins and hydroxyl groups on MAPOL to generate host:guest complexes. Nonetheless, complexation of cyanohydrin II-6R with MAPOL did not yield an ECCD active complex. Routine optimization procedure failed to give a detectable ECCD signal. The weaker hydrogen bonding ability of hydroxyl group as compared to chiral mono-amines is attributed to the latter observation. At this point we set out to include modification of the host system with the goal to increase binding interactions for complex formation (Figure II-5). Increase in the hydrogen bonding affinity Stronger hydrogen bonding donor Dual binding host systme Ph Ph Zn-MAPOL II-4 MAPOL II-3 Figure II-5. Structure of MAPOL and potential modifications to improve binding interaction. Ph Ph 60 Ph N H N OH OH N NH Ph N H N Modifications Ph binding site Ph HN N Ph N H N OH OH N NH EWG EWG Ph N H N Ph binding site Ph HN N Ph N N Zn N Ph N Ph binding site OH OH N Ph Ph N N Zn N Ph via introducing electron-withdrawing groups (EWG) on the backbone of MAPOL could potentially facilitate hydrogen bonding interaction by generating more acidic hydroxyl groups. However, as it will be discussed in detail in section II-4, MAPOL with fluorinated backbone proved to be incompetent for complex formation with alcohols. Figure II-6. UV-vis spectra of Zn-MAPOL (1 μM) in different solvents at room temperature. Alternately, porphyrin rings in MAPOL can be potentially utilized as binding sites for complex formation with chiral cyanohydrins. We sought inclusion of metal in these porphyrin rings in order to provide additional coordinating sites to enable ECCD active complex formation with chiral cyanohydrins. MAPOL was zincated with zinc acetate to form Zn-MAPOL (II-4) and the presence of zinc at the center of the porphyrin ring was confirmed using 1H NMR (disappearance of pyrrolic NH peaks). Zn-MAPOL has a strong UV absorption with lmax at 413 nm in hexanes. Figure II-6 illustrates the UV spectra of Zn-MAPOL in different solvents. A solution of Zn-MAPOL in 61 hexane was complexed with chiral cyanohydrin II-6R. Gratifyingly, strong ECCD signal was observed (Figure II-7). ECCD titration of Zn-MAPOL with different equivalents of cyanohydrin II-6R revealed that the amplitude of ECCD signals increases with higher equivalents of cyanohydrins. Saturation of the signal was observed at around 300 equivalents of cyanohydrin (Figure II-7, right). 5 equiv 7.5 equiv 10 equiv 30 equiv 15 equiv 50 equiv 75 equiv 100 equiv 250 equiv 500 equiv 750 equiv 1000 equiv D C . l o M 300 200 100 0 -100 -200 -300 380 420 400 Wavelength (nm) 440 700 600 500 400 300 200 100 0 e d u t i l p m A D C C E 460 0 100 200 300 400 500 Equivalence Figure II-7. ECCD spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl)acetonitrile II-6R. The ECCD signal can be detected after addition of 5 equivalents of chiral cyanohydrin. This accounts for less than 1 µg of cyanohydrin that is required for determination of its absolute configuration. Furthermore, the method eliminates preparative chemistries and ECCD signal can be obtained in minutes. Traditionally, hexane has been utilized as the solvent of choice for host:guest complex formation and ECCD measurements using the porphyrin tweezers methodology. The low polarity of hexane and other nonpolar solvents minimizes solvent-solute 62 interactions and consequently facilitates the host:guest complex formation. Indeed, only non-polar solvents result in formation of ECCD active complex of cyanohydrin and Zn- MAPOL (Figure II-8). The complex of Zn-MAPOL and cyanohydrin II-6R in hexane and pentane yield high ECCD amplitudes, while signal with lower intensity was observed when methyl cyclohexane was used as solvent. Nonetheless, ECCD signal was not observed in polar solvents such as dichloromethane, acetone, methanol, etc. D C . l o M 200 100 0 -100 -200 Hexane nPentane Methylcyclohexane Chloroform Dichloromethane Acetone Acetonirile Methanol 380 400 420 440 460 Figure II-8. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-2-hydroxy-2-(o-tolyl)acetonitrile II- 6R (50 equivalents) in different solvents at 0 ºC The solvent-solute interaction can be observed in the UV spectrum of Zn-MAPOL in different solvents (Figure II-6). Zn-MAPOL in hexane and pentane has similar UV-vis spectra with lmax at 413 nm. In more polar solvents, the UV-vis spectrum of Zn-MAPOL red shifts, along with increase in the absorption. This can be due to coordination to the zinc centered porphyrin or conformational changes of the host system. The ECCD of the host:guest complex is sensitive to the temperature of CD measurement. Stronger ECCD signals (higher amplitude of the signal) were observed at 63 lower temperatures. Upon increasing temperature, the intensity of the ECCD signal decreases, and above 40 °C the system becomes ECCD silent (Figure II-9). The trend shows a decay in the signal amplitude as a function of temperature. The decrease in the ECCD signal can be explained considering the fact that the low energy differences between the equilibrating diastereomeric atrop-isomers can be overcome at higher temperatures. D C . l o M 200 100 0 -100 -200 –5 C 0 C 5C 10 C 15 C 20 C 30 C 25 C 35 C 40 C 45 C 400 350 300 250 200 150 100 50 0 e d u t i l p m A D C C E 380 400 420 440 460 Wavelength (nm) 0 10 20 30 40 50 Temperature (°C) Figure II-9. ECCD spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl) acetonitrile II-6R (50 equivalents) at different temperatures. Next, we turn our attention to the generality of the method for complex formation with different chiral cyanohydrins. A diverse set of chiral cyanohydrins was synthesized following reported procedures.12-15 Hexane was used as solvent and ECCD measurements were performed at 0 °C with 1 µM Zn-MAPOL and 50 equivalents of chiral 64 cyanohydrins. As depicted in Table II-1, all surveyed cyanohydrins, derived from both aldehydes and ketones, gave strong ECCD signals. Table II-1. Predicted and observed ECCD for cyanohydrins complexed with Zn- MAPOL. sulfoxide OH CN II-5R entrya 1 2 3 4 5 6 7 8 9 OH CN Me OH CN OH CN OH CN OH CN OH CN OH CN OH CN Cl Cl MeO Me Pred. pos neg pos pos l nm (De) 424, +142 415, –101 424, –14 414, +25 423, +151 416, –130 423, +97 414, –107 423, –51 414, +55 A/Acorr c +243/+282 –39/–278 +281/+312 +204/+211 –106/–200 II-5S II-6R II-7R II-7S neg II-8R II-9R II-10R II-11R pos pos pos pos 423, + 124 415, –85 +209/+214 423, +92 415, –93 425, +43 416, –23 424, +99 415, –97 +185/+190 +66/+150 +196/+242 65 Table II-1 (cont’d) entrya 10 11 12 13 14 15 16 17 18 sulfoxide Pred. l nm (De) A/Acorr c Me Me Me S Me S Me Me Me Me Me OH CN OH CN OH CN OH CN OH CN OH CN OH CN OH CN OH CN II-12S pos II-12S II-13S II-13R II-14R neg pos neg pos II-14S neg II-15R pos II-15S neg II-16R pos 424, +46 415, –56 425, –54 415, +39 425, +49 415, –29 423, –45 414, +42 425, +42 416, –23 425, –32 414, +39 425, +45 415, –30 425, +44 416, –42 425, +28 418, –38 +102/+112 –93/–109 +78/+96 –87/–98 +65/+75 –71/–76 +75/+95 –86/–97 +66/+146 [a] All CD measurements were performed with 1 µM Zn-MAPOL in hexane at 0 °C; 50 equiv of cyanohydrin were used to record the data. [b] Corrected amplitude based on the ee of the cyanohydrins. [c] For ketones, 2 µM Zn-MAPOL was used for analysis. [d] Due to the low ee of the substrate, 100 equiv was used for complexation. 66 Of note are the enantiomers that give opposite signals with comparable amplitude when the enantiopurity of the cyanohydrin sample is taken into account (Acorr.). Furthermore, the amplitude of the ECCD signal of cyanohydrins derived from aldehydes is generally higher than that of cyanohydrins derived from ketones. A better stereo- differentiation of substituents on the chiral center (hydrogen versus R group) or stronger binding interaction (less steric repulsion) is most probably responsible for the higher ECCD amplitude of cyanohydrins derived from aldehydes. Furthermore, since the predicted sign arises from the relative size of the substituents on the chiral center, thus the predicted sign of ECCD signal is independent of the assignment of configuration (R or S). The R/S assignment is derived from Cahn-Ingold-Prelog rules that only considers the atomic number of substituents. e c n a b r o s b A 0.8 0.6 0.4 0.2 0 0 equiv 0.25 equiv 0.5 equiv 0.75 equiv 1.0 equiv 2.0 equiv 3.5 equiv 5.0 equiv 7.5 equiv 10.0 equiv 20.0 equiv 30.0 equiv 40.0 equiv 50.0 equiv 75.0 equiv 100.0 equiv 0.9 0.85 0.8 0.75 0.7 380 400 420 Wavelength (nm) 440 410 412 414 416 418 420 422 Figure II-10. UV-vis spectra of Zn-MAPOL complexed with (R)-2-hydroxy-2-(o-tolyl)-acetonitrile II-9R. 67 In Table II-1 we report the ECCD amplitude at 50 equivalents (50 µM) of added cyanohydrins. It should be noted that the signal could be observed at lower equivalents. Generally, aldehyde derived cyanohydrins give clear signal at around 5-10 equivalents while for the ketone derived cyanohydrins 10-20 equivalents of the guest molecule is required to have a reliable signal. Figure II-11. Job’s plot of Zn-MAPOL with (R)-4-methylmandelonitrile II-9R absorbance at 410 nm. The stoichiometry of the complex between cyanohydrins and Zn-MAPOL was determined employing Job’s plot analysis. Job’s plot was derived by following the changes in the UV-vis absorption (at 410 nm) of the complex upon increasing the equivalents of cyanohydrin (Figure II-11). A peak at 0.5 mole fraction of cyanohydrin is indicative of 1:1 host/guest complex formation. With this working methodology in hand, we sought to propose a binding model that can provide a mean for prediction of the sign of ECCD signals, enabling a solution for the 68 determination of absolute configuration of cyanohydrins. As such, we required a better understanding of the bonding paradigm involved in the complex formation between cyanohydrins and Zn-MAPOL. Both components of the complex (Zn-MAPOL and cyanohydrin) are capable of forming a bidentate interaction. Zn-MAPOL consists of hydrogen bonding elements (via hydroxyl groups) and coordination interaction (via the zinc centered porphyrin rings). Cyanohydrins, on the other hand, have hydroxyl group and nitrile moiety that both can be potentially involved in the hydrogen bonding or coordination interactions. A dual bonding host system was required to give rise to the observed ECCD signal. As discussed at the beginning of this chapter, MAPOL with only the hydrogen bonding element failed to produce an ECCD active complex with chiral cyanohydrins. Furthermore, weakening the hydrogen bonding capability of the host system, by replacing the hydroxyl groups of the host (Zn-MAPOL) with methoxy groups (Zn-MAPOL-OMe), failed to produce consistent ECCD signals with chiral cyanohydrins. Nonetheless, some of the chiral cyanohydrins did yield ECCD signals upon complexation with Zn-MAPOL-OMe, however the intensity of the signals was low. It should be noted that Zn-MAPOL-OMe can still act as hydrogen bond acceptor through the oxygen atoms and similar bonding regime as Zn- MAPOL can be anticipated. As a result, generation of ECCD active complex between some of the cyanohydrins and this host system is not unexpected. 69 In order to delineate the effect of each of the binding elements on the generation of host:guest complex, the chiroptical behavior of the ECCD active complex of cyanohydrin II-6R (50 equiv) with Zn-MAPOL in the presence of binding competitors was interrogated. The objectives were to introduce molecules that have similar binding moieties as cyanohydrins to the ECCD active complex and follow the effects on the ECCD signal. Acetonitrile was used to compete with the nitrile group of cyanohydrins and methanol was used as competitor of the hydroxyl group of cyanohydrins. These were added to the ECCD active II-6R:Zn-MAPOL complex individually. The amplitude of the signal decreases upon addition of these bonding competitors and complete eradication of the ECCD signal was observed at around 2000 equivalent of methanol (Figure II-12) and 4000 equivalents of acetonitrile (Figure II-13). It can be noted that the decay of the signal is much faster when methanol is added to the ECCD active complex as compared to 200 100 D C . l o M 0 -100 -200 0 equiv MeOH 25 equiv MeOH 50 equiv MeOH 100 equiv MeOH 500 equiv MeOH 1000 equiv MeOH 1500 equiov MeOH 2000 equiv MeOH 380 400 420 440 Wavelength (nm) e d u t i l p m A 350 300 250 200 150 100 50 0 460 D C C E 0 500 100015002000 Equiv of MeOH Figure II-12. Quenching the ECCD signal of II-6R in complex with Zn-MAPOL by addition of methanol. 70 addition of acetonitrile. This can potentially indicate that the hydroxyl group of cyanohydrin is the main binding element in the complex formation and disrupting this bonding element has a more profound effect on the strength of the ECCD signal. Additionally, chemical intuition suggests that hydroxyl groups are better hydrogen bonding donors and acceptors than nitrile groups. We envisioned the bonding paradigm in which the hydroxyl group of the cyanohydrin is involved in hydrogen bonding with the biphenol core while the nitrile is coordinated with the metallo center. 200 100 D C . l o M 0 -100 -200 0 equiv MeCN 250 equiv MeCN 500 equiv MeCN 1000 equiv MeCN 1500 equiv MeCN 2000 equiv MeCN 3000 equiv MeCN 4000 equiv MeCN 380 400 420 440 Wavelength (nm) 350 300 250 200 150 100 50 0 e d u t i l p m A D C C E 0 460 30 40 20 10 Equiv of MeCN Figure II-13. Quenching the ECCD signal of II-6R in complex with Zn-MAPOL by addition of acetonitrile (MeCN). With this proposed binding mode, the following mnemonic is presented (Figure II-14). Hydroxyl group of cyanohydrin is pointing towards the center of Zn-MAPOL to form a hydrogen-bonding network with the backbone of the host system. The nitrile group concurrently points towards the zinc in the center of one of the porphyrin rings. These two 71 binding interactions place the chiral tetrahedral carbon in the center of the host system and the induction of point chirality of cyanohydrin to the axial chirality of the host system can take place. Much akin to the discussion that was presented for complexation of chiral amine with MAPOL, minimization of steric repulsion is the determinant factor that leads to the preferred helicity. As shown in Figure II-14, in the R, M complex the larger group (Ar) based on A-value as the operating steric parameter,16 is in the sterically congested position. The repulsion between the porphyrin ring and the aryl group causes this complex to be less favorable. On the other hand, in the R, P complex, the large substituent on the chiral center is positioned in the open quadrant and the smaller group (hydrogen) is a Ph b Ph N N Zn NN Ph Ph H H R O H O Ar C Ph N H O NN Zn N N Ph P-helicity positive ECCD top view Zn Ar H N Zn P-(R) complex favored e d u t i l p m A 150 100 50 0 -50 -100 -150 c Ph N N Zn NN Ph Me OH R CN ECCD measurement Zn-MAPOL (1 µM) Hexane, 0 °C Ph Ph NN Zn N N Ph Ar R O H C Ph N HHH O O M-helicity negative ECCD top view H Zn Ar 380 400 420 440 Wavelength (nm) N Zn 460 M-(R) complex disfavored Figure II-14. a) Bidentate binding of II-6R, with the P-helicity of Zn-MAPOL, places the large aryl group in the most sterically available position. This favorable arrangement leads to the anticipated positive ECCD spectrum. b) Complexation of 5R with Zn-MAPOL leads to a strong positive ECCD signal. c) The same bidentate binding with the M-helicity leads to steric clash of the large aryl group with the porphyrin ring. Thus, negative ECCD with II-6R bound to Zn-MAPOL is not anticipated. 72 placed in close proximity of the porphyrin ring. One would expect this complex to be more stable. Since the helicity of this complex (P) generates a clockwise twist of the interacting chromophores, a positive ECCD signal is expected. Indeed, the experimental measurement confirmed the positive sign of the ECCD signal. The same analogy can be used for cyanohydrins in Table II-1 and the predicted ECCD signals are in agreement with the experimental ECCD signals. This working model can be used to determine the absolute configuration of chiral cyanohydrin with only considering the relative size of the substituents on the chiral carbon. The relative size of the substituents is used to determine the preferred helicity. A-values as thermodynamic steric parameters, are used as a general guiding tool for assignment of relative size (small versus large) of substituents on the chiral carbon. side view 2.3Å 2.0Å 1.8Å top view top view side view 2.3Å 2.0Å 1.8Å Figure II-15. Side and top view of geometry optimized (DFT-B3LYP/6-31G*) complex of II-10R bound to Zn-MAPOL. The model shows the bulky t-butyl group in the open quadrant, the hydrogen bonding of the alcohol with the biphenol core, and the interaction of the cyano group with the metal center. The optimized geometry predicts a positive ECCD, as observed. 73 Figure II-15 illustrates an energy minimized model for II-10R bound to Zn-MAPOL in the proposed orientation that leads to the observed ECCD. Of note are the structural features such as the ordinary bond angles and lengths for hydrogen bonding and coordination to the metallocenter that corroborate the proposed model in Figure II-15. In addition to the application of the technique for determination of the absolute configuration of chiral cyanohydrins, the method can be utilized for determining the enantiopurity of the cyanohydrin sample. The amplitude of ECCD signal is linearly correlated to the enantiopurity of the chiral samples. This in fact is one of the most from important properties of chiroptical techniques that enables analysis of samples that are not necessarily enantio-pure. The diastereomeric complexes arising two enantiomers of a molecule that are bound to the host system would have opposite ECCD signals and the excess enantiomer would lead to the observed signal. Samples with low enantio-purity can be an issue if Mosher ester or any other techniques are used for determination of the absolute configuration. In those techniques, sample with low ee would lead to the generation of disatereomers that need to be separated before analysis and it can complicate the assignment of configuration. We further take advantage of this unique property of chiroptical techniques to assess the enantio purity of the cyanohydrin sample. To this end, a standard cure was derived from correlating the ECCD amplitude with the enantiopurity of the cyanohydrin samples (Figure II-16). A linear fit with high regression (R2 = 0.998) was accessed upon complexation of cyanohydrin II-7 with Zn- MAPOL. The derived standard curve could be used to measure the ee of unknown samples. 74 150 100 D C . l o M 50 0 -50 -100 97% ee (R) 90% ee (R) 83% ee (R) 72% ee (R) 62% ee (R) 54% ee (R) 40% ee (R) 30% ee (R) 15% ee (R) 0% ee 18% ee (S) 39% ee (S) 53% ee (S) R2 = 0.995 D C 200 150 100 50 0 -50 -100 -150 -200 . l o M 380 420 400 Wavelength (nm) 440 -100-75 -50 -25 0 25 50 75 100 460 %ee (R) Figure II-16. ECCD spectra of Zn-MAPOL (1 μM) complexed with 4-chloromandelonitrile II-7 (50 equiv) at different enantio excess (left). The plot of ECCD amplitude versus %ee. In summary, we report on the use of a new host system for the absolute stereochemical determination of cyanohydrins in a non-empirical fashion. This constitutes the first direct methodology that enables a reliable and rapid assignment for the absolute stereochemistry of cyanohydrins. This is a microscale method that requires no derivatization and yields the results in a matter of minutes. The stereochemistry is reliably extracted from the CD spectra by assuming hydrogen bonding of the alcohol to the biphenol core, coordination of the cyano group with the zincated porphyrin, and minimizing steric interactions by placing the large group in the most unhindered quadrant. 75 II-2 Determination of absolute configuration of chiral sulfoxides The ubiquitous nature of the chiral carbon atom, in every facet of science as it relates to the structure of an organic molecule, has led to tremendous activity for assigning its absolute stereochemistry. The chiral sulfur atom has enjoyed much less attention, although one can argue it is no less important. In its oxide form, the sulfur atom can be asymmetric, with similarly profound impact on its chemical and spectroscopic characteristics, as well as its biological activity. From a substantial list of biologically active sulfoxide containing molecules,17 Nexium® (one of the world’s highest selling drugs), oxisurane, and armodafinil are examples of important pharmaceuticals that share the same commonality; the sulfoxide is the only asymmetric center in these molecules (Figure II-17).18-19 H N N O N S (S) O O Esomeprazole (Nexium®) $4.7 billion sales in 2015 Ph N (S) S O Oxisurane O O NH2 O S (R) Ph Armodafinil Figure II-17. Example of bioactive drugs with chirality at the sulfur atom. Chiral sulfoxides as ligands and auxiliaries are also important contributors in asymmetric synthesis, and are used often in both academic and industrial settings.20-22 The importance of this functionality necessitates a robust, reliable, efficient, preferably micro-scale and direct method for the assignment of absolute configuration. Nonetheless, the structural features of sulfoxides (lack of a suitable derivatizing site) has posed a challenge to port over the prototypical procedures used for the absolute stereochemical 76 O R1 S R2 MSH NH2 O R1 S R2 Mes-SO3– KOH O R1 S NH R2 (S)-MPA coupling reagent (R)-MPA O R1 S O R1 S N (S)-MPA R2 N (R)-MPA R2 MSH = Mes-SO2-O-NH2 MPA = Ph MeO H CO2H N SO R1 R2 H O OMe Figure II-18. Chemical modification and derivatization of chiral sulfoxides for determination of absolute configuration. assignment of carbon substituted functionalities. A modified Mosher ester analysis was an early attempt in this regard. Typical ester derivatization of the sulfoxide oxygen results in racemization on the chiral sulfur. Yabuuchi and Kusumi circumvented this problem via an initial oxidation of the sulfoxide to generate corresponding iminosulfanone, which was then derivatized as its MPA-amide (Figure II-18).23 NMR analysis of the Mosher ester derivative employing the proposed mnemonic (Figure II-18, shadow box) could lead to assignment of stereochemistry on the chiral sulfur. Limited use of chiral solvating agents for NMR analysis has been reported, although this strategy has not delivered a comprehensive solution.24-26 An electronic CD method to address aryl sulfoxides has been reported by Rosini and coworkers.24 Alternatively, the most successful chiroptical technique for determination of absolute configuration of sulfoxides has been vibrational circular dichroism (VCD),27-31 although this has not yet become a routine and common methodology for the synthetic community. 77 Our goal was to develop a simple and micro-scale procedure that enables the facile determination of the absolute stereochemistry of chiral sulfoxides using Exciton Coupled Circular Dichroism (ECCD). As previously discussed, our approach to this problem originates from our interest in utilizing host systems capable of adopting either the P or M helicity as the consequence of predictable interactions of a bound chiral guest molecule. The host:guest interactions are such that the point chirality of the guest is translated to the axial chirality of the host, which in turn leads to an observable ECCD signal. To this end, we envisioned the use of our recently developed host molecule, MAPOL II-3,1 as a receptor that binds sulfoxides. Our constraints for success were to develop a methodology that is sensitive, highly reproducible, and operationally simple for chemists not necessarily versed in spectroscopic techniques. Herein, we demonstrate the utility of Zn-MAPOL II-432 to address a long-standing problem in the assignment of absolute stereochemistry of chiral sulfoxides with the specific goals addressed above. Although sulfoxides have putatively two sites of coordination (the oxygen atom and the sulfur lone pair), the geometric constraints of the host system would preclude bidentate binding. We first hypothesized that sulfoxides could bind via hydrogen bonding to the biphenol, and thus provide a mechanism for interaction with MAPOL. Nonetheless, binding of a number of chiral sulfoxides with MAPOL failed to produce a discernable ECCD signal. We had also envisioned the possibility of a different binding paradigm with the zincated analogue of MAPOL (Zn-MAPOL). One could propose chiral sulfoxides putatively coordinate to the zinc center of the porphyrin ring. Gratifyingly, complexation of 78 model sulfoxide II-20R with Zn-MAPOL led to a strong ECCD signal centered on the Soret band (420 nm) of the host system (Figure II-19). D C . l o M 200 100 0 -100 -200 380 O S II-20R 460 400 420 440 Wavelength (nm) Figure II-19. ECCD spectrum of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in hexane at 0 ºC. Figure II-20. UV-vis spectra of Zn-MAPOL (1 μM) and Zn-MAPOL complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC (solvents that yield ECCD signal). 79 As depicted in Figure II-22, when nonpolar solvents (hexane, pentane and methyl cyclohexane) were used for complex formation of II-20R with Zn-MAPOL, strong ECCD signals were observed. On the other hand, polar solvents failed to give any detectable ECCD signal for II-20R:Zn-MAPOL complex (Figure II-22). The UV-Vis spectra of II- 20R;ZnMAPOL complex in different solvents also revealed complex formation between sulfoxide II-20R and Zn-MAPOL. The UV-Vis spectra in solvents that the complex yield ECCD signal, show characteristic red shift upon addition of sulfoxide to Zn-MAPOL solution (Figure II-20). In sharp contrast, in solvents that the complex was ECCD silent, addition of sulfoxide did not change the UV spectrum (Figure II-21). It can be summarized that in the case of polar solvents, the interaction between the solvent and the host system out competes the interaction between the sulfoxide and Zn-MAPOL. Figure II-21. UV-vis spectra of Zn-MAPOL (1 μM) and Zn-MAPOL complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC (solvents that do not yield ECCD signal). 80 D C . l o M 200 100 0 -100 -200 -300 hexane pentane methyl cylcohexane isopropyl alochol acetone acetonitrile dichloromethane chloroform 380 400 420 440 460 Wavelength (nm) Figure II-22. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC. In addition, the II-20R:Zn-MAPOL showed the expected temperature dependence for the observed signal (Figure II-23). At lower temperatures the ECCD amplitude is higher, and at above 40 °C the ECCD signal was not observed. Interestingly, upon lowering the temperature the ECCD signal can be restored. The latter observation consistent the fact that at the lower temperatures the energy difference between diastereomeric rotomers is more pronounced, thus leading to a larger population difference. Next, the dependence of the ECCD amplitude to the equivalents of sulfoxide was investigated (Figure II-24). ECCD signal can be observed upon addition of 5 equivalents of sulfoxide to Zn-MAPOL. Increasing the equivalents of sulfoxide increases the ECCD amplitude while preserving the sign of the ECCD signal, reaching saturation at 100 equivalents in most cases. 81 –5 C 0 C 5C 10 C 15 C 20 C 25 C 30 C 40 C back to 0 C 300 200 100 0 -100 -200 -300 D C . l o M 380 400 420 440 460 Wavelength (nm) Figure II-23. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R (50 equiv) in different solvents at 0 ºC. 5 equiv. 10 equiv. 20 equiv. 30 equiv. 40 equiv. 50 equiv. 60 equiv. 80 equiv. 100 equiv. 150 equiv. 200 equiv. 300 200 100 0 -100 -200 -300 D C . l o M 380 420 400 Wavelength (nm) 440 460 Figure II-24. ECCD spectra of Zn-MAPOL (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane. 82 The saturation of the ECCD amplitude is due to the fact that the host:guest complexation is a dynamic process, while the ECCD amplitude is proportional to the population of the bound complex that is favored by a larger excess of the sulfoxide. As illustrated in Figure II-25, the binding affinity can be measured at 409 nm upon addition of sulfoxide to the solution of Zn-MAPOL (1 µM). Kassoc of the complex is measured in excess of 14,500 M-1 (Figure II-27). Furthermore, the stoichiometry of the complex between sulfoxide and Zn-MAPOL is extracted from the UV titration spectra. Job’s plot analysis indicates a 1:1 stoichiometry of the complex as evident from the maxima obtained at 0.5 mole fraction (Figure II-26). e c n a b r o s b A 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 ZnMAPOL 0.1 equiv. 0.25 equiv. 0.5 equiv. 0.75 equiv. 1 equiv. 2.5 equiv. 5 equiv. 7.5 equiv. 10 equiv. 20 equiv. 30 equiv. 40 equiv. 50 equiv. 75 equiv. 100 equiv. 150 equiv. 200 equiv. 300 equiv. 500 equiv. 460 380 400 420 440 Wavelength (nm) Figure II-25. UV-vis spectral change upon titration of Zn-MAPOL (1 μM) with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane. 83 Figure II-26. Job’s plot of Zn-MAPOL with (S)-p-tolyl vinyl sulfoxide II-20R absorbance at 409 nm. m n 9 0 4 @ s b A Δ 0.4 0.35 0.3 0.25 0.2 0.15 0.1 0.05 0 = 14672 M-1 K assoc R2 = 0.993 0 0.0001 0.0002 0.0003 0.0004 0.0005 Con.p-tolyl vinyl sulfoxide (molar) Figure II-27. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC. 84 With the initial results in hand, a number of chiral sulfoxides, with various alkyl and aryl substituents were synthesized via previously reported procedures (Table II-2).29, 33-37 Using a standard protocol (1 µM Zn-MAPOL and 50 equivalents of sulfoxide dissolved in hexane), strong ECCD signals were obtained for host/guest complexes in all cases. Not surprisingly, enantiomers yield ECCD signals with opposite sign albeit with equal intensities when correction for enantiopurity is applied (Acorr). Table II-2. Predicted and observed ECCD for sulfoxides complexed with ZnMAPOL. entrya 1 2 3 4 5 6 7 sulfoxide O S Me O S Me O S O S O S O S O S Me Me 85 L1(Å)/L2 (Å)b Pred. II-17R 4.6/1.8 neg II-17S 4.6/1.8 pos II-18R 6.1/1.8 neg II-18S 6.1/1.8 pos II-19R 6.1/2.7 neg II-19S 6.1/2.7 pos II-20R 6.1/2.7 neg l nm (De) 426, –166 418, +173 425, +168 416, –116 427, –252 419, +219 427, +199 419, –137 427, –248 419, +240 427, + 218 419, –135 427, –241 419, +178 A/Acorr c –339/–365 +284/+351 –471/–611 +336/+589 –488/–574 +353/+588 –419/–445 Table II-2 (cont’d) entrya sulfoxide O S L1(Å)/L2 (Å)b Pred. l nm (De) A/Acorr c II-21R 6.1/2.8 neg II-21S 6.1/2.8 pos II-22R 6.1/4.6 neg II-22S 6.1/4.6 pos II-23R 5.3/1.8 neg II-24R 2.8/1.8 neg II-24S 2.8/1.8 pos II-25R 4.6/2.8 neg O S O S O S O S Me O S O S O S Me Me 428, –259 420, +140 427, +154 419, –84 428, –299 420, +262 428, +257 419, –201 426, –97 418, +81 426, –168 418, +137 426, +122 418, –155 428, –270 419, +155 425, +48 417, –29 426, +65 420, –30 425, +17 417, –9 –399/–463 +238/+456 –561/–738 +458/+750 –178/–212 –305/–406 +277/+420 –425/–559 +77d +95/+271 +26d 8 9 10 11 12 13 14 15 16 17 18 O O S O S O S D5 II-26S 6.1/5.8 pos II-27S 2.8/2.8 pos II-28S 2.8/2.8 pos 86 sulfoxide O S NH2 L1(Å)/L2 (Å)b Pred. l nm (De) A/Acorr c II-29S 6.1/1.6 neg 426, –709 418, +624 –1333 d Table II-2 (cont’d) entrya 19 [a] Zn-MAPOL (1 µM) and 50 equivalents of the sulfoxide were dissolved in hexane, and CD measurements were performed at 0 ºC. [b] L1 and L2 correspond to the substituents on the left and right hand side of the structures as drawn, respectively. [c] Acorr values are calculated by dividing the measured A by the %ee. [d] These samples were enantiomerically pure. Deriving a simple mnemonic for correlating the sign of the observed ECCD spectrum to the absolute stereochemistry at the sulfur center requires a structurally detailed picture of the geometric constraints that lead to the observed helicities reported in Table II-2. To begin, it is important to point out a number of facts: 1. Zincation of the porphyrin rings was obligatory to observe ECCD with bound chiral sulfoxides. 2. The bis-methoxylated Zn- MAPOL (Zn-MAPOL-OMe) did yield ECCD signals of the same sign, albeit with reduced amplitudes, thus suggesting that coordination to the zinc center is the main binding interaction responsible for induced helicity. Figure II-27 illustrates, the CD titration of sulfoxide II-20R with Zn-MAPOL-OMe. The sign of the ECCD signal is same as when this substrate is complexed with Zn-MAPOL (Figure II-19) however the amplitudes of signals are lower at any given equivalents of sulfoxide. 3. Hard-soft binding principles dictate the tendency for sulfoxides to coordinate to metals either through oxygen or sulfur. Oxygen atom binding to metal centers is most often observed as a result of its greater polarization, and thus higher electron density. Soft metals (late transition metals such as platinum) tend to coordinate to sulfur, while harder metals prefer oxygen as the coordinating site.22 Fortuitously, the crystal structure of Zn-TPP in complex with dimethyl sulfoxide with Zn- 87 TPP is reported and indeed reveals the anticipated coordination of oxygen to the zinc center.38 Furthermore, we obtained a high resolution crystal structure of Zn-TPP in complex with dimethyl sulfoxide that confirms the coordination of oxygen atom of sulfoxide to the zinc center of the porphyrin (Figure II-28). Unfortunately, countless efforts to obtain crystal structure of Zn-MAPOL or its analogue with or without bound substrate were unsuccessful The latter three points argue strongly for the coordination of the sulfoxide oxygen atom to the metallated porphyrin. 100 50 0 -50 -100 -150 D C . l o M 50 equiv. 100 equiv. 150 equiv. 200 equiv. 300 equiv. 400 equiv. 380 420 400 Wavelength (nm) 440 460 Figure II-28. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (R)-p-tolyl vinyl sulfoxide II-20R at 0 ºC in hexane. Figure II-29 depicts our proposed mode of binding and conformational arrangements that lead to the preferred helicity and consequently the observed ECCD. This was derived based on the constraints and observations about complexation described above. Coordination of the oxygen atom lone pairs of the sulfoxide with the metalloporphyrin occurs in a manner to minimize steric interactions. The geometry of the sulfoxide, in 88 particular the lone pair orbital bound to Zn2+, necessitates the projection of one of the sulfur atom substituents towards the bound porphyrin ring. This is apparent from the crystal structure of DMSO bound to Zn-TPP with a Zn-O-S bond angle of 128°. Of the three substituents on the sulfur atom, the lone pair presumably would be the least sterically intrusive group, and thus would occupy this space. Our own high resolution crystal structure of DMSO, bound to Zn-TPP, also demonstrates the same conformational preference, with the lone pair pointing towards the plane of the porphyrin ring (Figure II- 29). Figure II-29. X-ray crystal structure of Zn-TPP in complex with dimethyl sulfoxide (hydrogens are omitted for clarity). As illustrated with substrate II-20R in Figure II-30, the next choice would be to minimize steric interactions of the two remaining groups as they project outwards from the center of the porphyrin ring they are bound to, by placing the largest group, in this case the aryl substituent, in the least sterically demanding space. Considering both P and M helicities of the host, the M-(R) complex orients the aryl group away from the 89 second porphyrin ring, while in the P-(R) complex, the aryl group suffers from larger, undesired interactions. One would thus suggest that the M helicity is energetically favored. The prediction based on this model leads to a negative induced helicity in Zn- MAPOL, which indeed fits the observed ECCD spectrum for II-20R bound to Zn-MAPOL. This simple mnemonic predicts the correct ECCD for all substrates listed in Table II-2. a Ph b Ph N N Zn NN Ph Ph Ar Ph S (R) O H H O O NN Zn N N Ph P-helicity positive ECCD top view Zn Ar (R) S O Zn D C . l o M 200 100 0 -100 -200 380 O S (R) (50 equiv.) ECCD measurement ZnMAPOL (1 µM) Hexane, 0 °C c Ph N N Zn NN Ph Ph Ar S (R) O HH O O Ph Ph NN Zn N N Ph M-helicity negative ECCD top view Zn Ar (R) S O 400 420 Wavelength (nm) Zn 440 P-(R) complex disfavored M-(R) complex favored Figure II-30. a) Bidentate binding of II-20R, with the P-helicity of Zn-MAPOL, places the large aryl group in the most sterically available position. This favorable arrangement leads to the anticipated positive ECCD spectrum. b) Complexation of II-20R with Zn-MAPOL leads to a strong positive ECCD signal. c) The same bidentate binding with the M-helicity leads to steric clash of the large aryl group with the porphyrin ring. Thus, negative ECCD with II-20R bound to Zn-MAPOL is not anticipated. The critical choice to be made for applying the latter mnemonic rests with determining which of the two groups attached to the asymmetric sulfur is larger. In systems described previously (amines and cyanohydrins), A-values, a thermodynamic measure of steric size, were used as determinants of size. A careful analysis of the data in Table II-2, 90 specifically, correlation of the strength of the ECCD signal with the difference in steric size of the substituents, suggests that A-values alone are not sufficient to rationalize the data. For example, although the A-values of phenyl and 4-Me-Ph are expectedly similar, the ECCD observed for II-22 (Table II-2, entries 10 and 11) is unexpectedly large (Acorr = – 738). The greatest difference between the latter two substituents is in their length, i.e., measuring the distance L1 from the sulfur atom to the furthest heavy atom on 4-Me-Ph is 6.1 Å, while L2, measured from the sulfur atom to the furthest heavy atom on Ph is 4.6 Å (Figure II-31). The geometry dictated by the coordination of the sulfoxide with the zincated porphyrin projects the two substituents towards the plane of the opposite porphyrin ring. Thus, the difference in length of the two substituents is more significant than their bulk, leading to a larger P/M population difference. L 1 = 6.1 Å L 2 = 4.6 Å P-(R) complex M-(R) complex Figure II-31. Energy minimized (B3LYP/6-31G*) structures of the P and M helical Zn-MAPOL complexed with II-22R. The length of the substituents, measured as L1 and L2, dictate the preferred helicity of the host system. The shorter L2 of the phenyl group leads to smaller steric interactions with the porphyrin ring that is not bound to the sulfoxide. As depicted in Figure II-31, the length of the substituents is measured in an analogous fashion to the L parameter for sterimol analysis (Figure II-32).39-42 A note of caution is 91 warranted; the latter criterion applies only if the length of the substituent can be predicted in a reasonable manner. In cases such as flexible long chain alkyl groups, where a number of different energetically close lying conformations can be present, the prediction for length can be difficult, and thus, could lead to erroneous predictions. L L1 L : Sterimol parameter L1 : Measured length Figure II-32. Sterioml L parameter (total length of the substituent along the primary bond axis) and the measure length (L1, length of substituent to the furthest heavy atom) are illustrated for 1,2-dimethylpropyl group. Prioritization of the longer group over the shorter substituent, as described above, leads to a predictable assignment of absolute stereochemistry for all substrates listed in Table II-2. Note that the table lists L1 and L2 values for each substituent, which are used to derive the predicted sign of the ECCD spectrum. The most striking example that validates the use of length, as the primary consideration to distinguish the relative size of the substituents on the sulfur center is the ECCD obtained with II-25R (Table II-2, entry 15). Considering A-values, one would expect the opposite sign for the ECCD spectrum as the t-butyl group has a larger A-value (>4.5) than the phenyl substituent (3.0). Nonetheless, the length of the phenyl group is longer (4.6 Å for Ph, 2.8 Å for t-butyl), thus dictating the population difference that leads to the observed negative ECCD. In cases where the measured L is identical for both groups, then A-values are used as the secondary consideration to decide size priority. An example of this is II-27S, where L1 and L2 for t-butyl and iso-propyl are the same, yet the A-value for t-butyl (>4.5) is 92 substantially larger than the A-value for iso-propyl (2.15). Considering that the A-value is a predictor of ‘volume’, it stands to reason that when the lengths of the substituents are the same, the volume of the substituents can exert a stereodefining role. Indeed, the predicted positive ECCD is observed experimentally (Table 1, entry 17). Notably, the same analysis can be used for the absolute determination of II-28S. The slightly larger per-deuterated benzene ring leads to the anticipated ECCD signal, albeit in small amplitude, reflective of the slightly larger volume of the perdeuterated phenyl vs. phenyl. Nonetheless, the latter observation demonstrates the sensitivity of the current system. The size difference of phenyl versus perdeuterated phenyl is very minimal. Deuterium with A-value of 0.006 is assumed to be slightly larger than hydrogen (A-value of 0.000). In a seminal analysis Dunitz and Ibberson have studied the molecular volume of C6H6 versus C6D6 in the crystal structure at different temperature.43 They realized that at lower temperatures benzene is slightly larger than deuterated benzene due to stronger vibrations of C–H versus C–D. However, at temperature above 170 K deuterated benzene occupies more space than C6H6. The sulfinamide II-29S (Table II-2, entry 19) produced II-33 the demonstrates the ECCD titration of ZnMAPOL with sulfinamide II-29S. Much akin to sulfoxides, saturation of the ECCD amplitude was observed at about 100 equivalents of sulfinamide (Figure II-33, right). Strong ECCD amplitude observed for sulfinamide in complex with Zn-MAPOL can be indicative of high binding affinity of this moiety as compared to sulfoxides. Binding affinity of sulfinamide in complex formation with Zn- largest ECCD amplitude upon complexation with ZnMAPOL. Figure 93 MAPOL was measured by following the changes in the UV-Vis spectra of Zn-MAPOL upon addition of different equivalents of sulfinamide (Figure II-34). 800 600 400 200 0 -200 -400 -600 -800 D C . l o M 5 equiv. 10 equiv. 15 equiv. 20 equiv. 30 equiv. 25 equiv. 40 equiv. 50 equiv. 75 equiv. 100 equiv. 150 equiv. 200 equiv. 380 420 400 Wavelength (nm) 440 460 Figure II-33. Negative ECCD spectrum obtained with Zn-MAPOL complexed with 50 equivalents of (S)- p-toluene sulfinamide II-29S at 0 ºC in hexane. e c n a b r o s b A 0.8 0.6 0.4 0.2 0 ZnMAPOL 0.1 equiv. 0.25 equiv. 0.5 equiv. 0.75 equiv. 2.5 equiv. 1 equiv. 5 equiv. 7.5 equiv. 10 equiv. 25 equiv. 40 equiv. 50 equiv. 75 equiv. 100 equiv. 380 400 420 440 460 Figure II-34. UV spectral change upon titration of Zn-MAPOL (1 μM) with (S)-p-toluene sulfinamide S- II-29S at 0 ºC in hexane. Wavelength (nm) 94 A superior binding affinity (Kd = 35,236 M-1) was measured that can presumably be indicative of the highly polarized nature of sulfur–oxygen bond (Figure II-35). Figure II-35. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (S)-p-toluene sulfinamide II-29S at 0 ºC. In the sulfinamide substrate there is an –NH2 group that can coordinate to the zinc center and potentially change the binding mode for complex formation. The crystal structure of sulfinamide II-34S with Zn-TPP provides clear evidence for the anticipated binding of the oxygen atom of this substrate to the metallocenter (Figure II-36). 95 Figure II-36. X-ray crystal structure of Zn-TPP complexed with sulfinamide II-34S (hydrogens are omitted for clarity). Having demonstrated the efficacy of the methodology with a variety of chiral sulfoxides, we next challenged the system with a more complex molecular structure containing ancillary groups that could alter or abolish coordination of the sulfoxide moiety with the metalloporphyrin. To address the latter concern, esomeprazole (Nexium®), a highly effective proton pump inhibitor, with a number of potential coordinating functional groups was complexed with Zn-MAPOL (Figure II-37). Gratifyingly, a positive ECCD was observed, consistent with what was expected, considering the longer benzo-imidazole substituent as the determinant for helicity. 96 Zn-MAPOL 2 (1 µM) Hexane 0 °C small 7 . 9 Å O Large H N N O S (S) N esomeprazole (Nexium®) 6.7 Å O D C . l o M 200 150 100 50 0 -50 -100 380 420 400 440 Wavelength (nm) 460 Figure II-37. Complexation of esomeprazole (50 µM) with Zn-MAPOL (1 µM) yields a positive ECCD spectrum, correctly predicting the S stereochemistry of the molecule. The assignment is based on considering the benzoimidazole side chain as the larger substituent (longer linear length as compared to the pyridyl side chain). In summary, we have reported the first direct chiroptical method for the absolute stereochemical determination of chiral sulfoxides without the need for derivatizations or chemical transformations. The procedure is reliable, requiring only microgram quantity of chiral sulfoxides. Furthermore, the absolute stereochemistry is derived in a matter of minutes, with an operationally simple method that is amenable for researchers with little experience in chiroptical techniques. The mnemonic for correlating the observed ECCD sign to the actual stereochemistry at the sulfur center is straightforward, requiring the operator to only make a choice on the relative length and size of the substituents attached to the sulfur center. 97 II-3 Determination of absolute configuration of chiral phosphine oxides With the successful demonstration of Zn-MAPOL as a molecular receptor for the determination of absolute configuration of chiral sulfoxides, we were interested to further expand the utility of this host system for other functionalities. In this regards, we envisioned application of Zn-MAPOL for determination of absolute configuration of phosphine oxides as close analogues of chiral sulfoxides. The lack of a reliable methodology to detect the absolute configuration of chiral phosphine oxides was our motivation to pursue the feasibility of complex formation of phosphine oxide with our for determination of absolute configuration newly designed host system for the purpose of sensing the chirality on the chiral phosphorus atom. Current state of the art for determination of absolute configuration of chiral phosphine oxide is heavily reliant on X-ray crystallography. Nonetheless, tert-butyl-1-(2- application of VCD methylnaphthyl) phosphine oxide is reported.44 Chiral phosphine oxides are widely used in transition metal chemistry as ligands.45-46 Upon reduction, phosphine oxides can be converted into chiral phosphines which are equally important as chiral phosphine oxides. Additionally, chiral phosphine oxide moiety can be found in a number of pharmaceutically important products. Our goal was to leverage the inherent sensitivity of chiroptical techniques utilizing our host system to devise a methodology that can lead to the absolute stereochemical determination of chiral phosphine oxides in a rapid, micro-scale, derivatization-free and non-empirical fashion. Mr. Debarshi Chakraborty (Borhan group, MSU) has synthesized a diverse set of chiral phosphine oxides utilizing reported procedures.47-50 Complex formation between 98 various phosphine oxides and Zn-MAPOL, utilizing the optimized condition for complexation of sulfoxides, leads to detectable and reliable ECCD signals. As an example, the ECCD signal of (R)-methyl phenyl propyl phosphine oxide with Zn-MAPOL is shown (Figure II-38). O P II-30R Ph Me (R)-methyl-phenyl-propyl-phosphine oxide Figure II-38. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (R)-methyl phenyl propyl phosphine oxide II-30R at 0 ºC in hexane. As expected, the enantiomer of this phosphine oxide leads to ECCD signal with opposite sign (Figure II-39). Interestingly, the observed amplitudes for ECCD of chiral phosphine oxides are generally higher than chiral sulfoxides. 99 O P II-30S Me Ph Figure II-39. ECCD spectra of Zn-MAPOL-OMe (1 μM) complexed with (S)-methyl-phenyl-propyl phosphine oxide II-30S at 0 ºC in hexane. Figure II-40. UV-vis spectral change upon titration of Zn-MAPOL (1 μM) with (R)-methyl-phenyl-propyl phosphine oxide II-30R. The large amplitudes for phosphine oxides proved instrumental for our 100 measurements, since the ECCD signals could be measured with as low as only one equivalent of phosphine oxide when complexed with Zn-MAPOL (1 µM). This set an extremely low detection limit that has enormous advantages for analysis of samples with limited available quantity. The exceptionally high intensity of the ECCD signals finds its origin in high bonding affinity of phosphine oxides to Zn-MAPOL that is the result of the highly polarized phosphorous–oxygen bond. Binding affinity was measured following changes in the UV-vis absorption of Zn-MAPOL at 409 nm upon addition of chiral phosphine oxide (Figures II-40 and II-41). This bonding affinity is roughly twenty times higher than the affinity measured for sulfoxides. Figure II-41. Change in the absorbance at 409 nm for evaluating Kassoc upon titration of Zn-MAPOL (1 μM) with (R)-methyl-phenyl-propyl phosphine oxide II-30R. at 0 ºC. We envisioned the binding interaction of chiral phosphine oxides with Zn-MAPOL to 101 be similar to what we proposed for chiral sulfoxides. As such, coordination of the highly polarized oxygen atom of P–O bond to the metalocentered porphyrin drives the complex formation and places the stereogenic phosphorus atom at the center of the host system. Minimization of steric repulsions dictates the preferred helicity that leads to the corresponding ECCD signal. In a similar arrangement of substituents on the chiral center as proposed for chiral sulfoxides, the smallest group on the phosphine oxide would point towards the bound porphyrin and the remaining medium and large groups project towards the second porphyrin ring. The relative size of medium and large substituents induce preference on the orientation of the second porphyrin ring, leading to the corresponding twist on the backbone of the host system. The preferred helicity is obtained such that the second porphyrin reside closer to the medium size group and further away from the larger group to minimize steric repulsion. The mnemonic shown in Figure II-52 and II-53 can be used to predict the sign of the ECCD signal by considering the relative size of the substituents on the chiral center. As stated earlier, a large number of pharmaceutical products contain embedded chiral phosphine oxides in their skeleton. A research team from Merck Chemical Company showed interest in employing our Zn-MAPOL methodology to sense the absolute configuration of chiral phosphine oxides within drug molecules. They have recently reported a new methodology that gives access to a family of prodrugs with chiral phosphorous center in the form of phosphoramidates.51 Zn-MAPOL proved to be efficient for formation of ECCD active complexes with chiral phosphine oxides bearing carbon based substituents. We sought to verify the applicability 102 of our host molecule with complex systems that contain other coordinating sites and chiral centers along with a chiral phosphine oxide moiety. At the outset of this endeavor, we were interested in developing a reliable method that can be used by scientists at Merck for the determination of the absolute configuration of stereogenic phosphorous as part of pharmaceutical products. In this regards, determination of the absolute configuration of Sofosbuvir as a close analogue of most of pharmaceutical compounds containing stereogenic phosphorous was targeted (Figure II-42). Sofosbuvir is currently used for treatment of hepatitis C. This molecule has multiple chiral centers as well as different functionalities such as ester, amide, hydroxyl, etc. The only difference between the two stereoisomers of Sofosbuvir is the configuration on the stereogenic phosphorous atom. In an ideal scenario, the complex formation between the host system and these pseudo enantiomers (with regards to the phosphorous center) should yield ECCD signals with opposite sign. In this envisioned setting, the preference of the helical twists solely originates from the chirality on phosphorous and the substituents attached to this moiety. Using hexane, the optimal solvent for most of our host:guest complex formations, both epimers of Sofosbuvir resulted in a positive ECCD signal, albeit with different intensities. The intensity of the ECCD signal for Sofosbuvir was higher than the ECCD signal for epi-Sofosbuvir. This observation was not unexpected, since these two epimers are diastereomers of one another. 103 O NH O N O P O O O N H O HO F O Ph Sofosbuvir Figure II-42. Structures of Sofosbuvir and epi-Sofosbuvir. O O Ph Epi-sofosbuvir O P O N H O N HO F O NH We postulated that this inconsistency is the result of possible secondary interactions along with the primary coordination of the phosphorous oxygen moiety to the zinc centered porphyrin. These secondary interactions can exist as a result of hydrogen bonding or possibly coordination of heteroatoms of the substrate to the second zinc center. In other word, the two stereoisomers behave like two diastereomers (as they are) in complex formation with Zn-MAPOL and the opposite configuration on the phosphorous center is obscured by other parameters. Based on our previous findings with this host system, the phosphine oxide binding with zinc is the strongest interaction amongst other functionalities that are present in the molecule. Our approach to this issue was to take advantage of the superior binding affinity of phosphine oxide in comparison to the other moieties and pursue strategies that weaken the overall interaction. We hypothesized that the weak interactions can be deteriorated or eliminated, while preserving the stronger interaction (phosphine oxide). In this manner, we investigated complex formation in the presence of different additives in order to weaken the putative secondary interactions that originate from other functionalities embedded in the molecule. It was noticed that different additives have a similar behavior, resulting in decreasing the ECCD intensity of both epimers. 104 Based on our previous investigations, the interaction between the host system and the guest molecule is maximized in hexane. As a result, the ECCD intensity in this is maximum for any given host-guest system. Hexane was chosen as solvent and the additives were used from their stock solutions in DCM. We noticed that after addition of about 50-100 equivalents of additive (butyl amine for instance) to the complex of Sofosbuvir and ZnMAPOL (1 µM each), the ECCD signal of both epimers of Sofosbuvir decreases and the ECCD signal for epi-Sofosbuvir switches from postivie to negative. We hypothesized that the additives either occupy the second zinc centered porphyrin or disrupt the possible hydrogen bonding network. As a result, the interaction between Sofosbuvir and ZnMAPOL becomes mono-dentate (as desired), thus the host/guest complex would follow the same mnemonic as a simple phosphine oxide substrate. D C . l o M 200 100 0 -100 -200 0 µL DCM 10 µL DCM 10 µL DCM 20 µL DCM 40 µL DCM 60 µL DCM 80 µL DCM 100 µL DCM 380 400 420 440 460 Figure II-43. ECCD spectra of epi-Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, DCM was added. Wavelength (nm) 105 The observed trend was similar for various additives. We realized that the effect is related to the change in solvent composition as opposed to the effect of additives (addition of DCM). Next, addition of DCM to a solution of Sofosbuvir and ZnMAPOL (1 µM each) in hexane at room temperature was investigated (Figure II-43). a b c Figure II-44. ECCD spectra of epi-Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, NO dichloromethane was added (a), 40 µL dichloromethane was added (b) and 100 µL dichloromethane was added (c). As observed before, the amplitude of the initial positive ECCD signal of epi-Sofosbuvir in complex with ZnMAPOL (Figure II-44-a) decreased by addition of DCM. A clear switch in the ECCD signal was observed upon addition of 40 µL DCM to the complex in 1 mL hexane (Figure II-44-b) and followed by a clean negative signal after addition of 100 µL of DCM (Figure II-44-c). Nevertheless, the ECCD signal for Sofosbuvir showed a lower degree of dependence to the addition of DCM. After addition of 150 µL DCM, the signal 106 decreased to ~80% of its initial intensity and the sign of the ECCD signal remained positive (Figure II-44). 0 µL DCM 50 µL DCM 10 µL DCM 100 µL DCM 150 µL DCM 600 400 200 0 -200 -400 D C . l o M 380 400 420 440 460 Wavelength (nm) Figure II-45. ECCD spectra of Sofosbuvir (1 equiv) complexed with ZnMAPOL (1 µM) in hexane at room temperature, DCM was added. 107 Figure II-46. ECCD spectra of Sofosbuvir complexed with ZnMAPOL (1 µM) in dichloromethane. Next we measured the ECCD spectra for both epimers of Sofosbuvir in DCM. Sofosbuvir (0.001 M in DCM) was added to a 1 µM solution of ZnMAPOL in DCM at 0 °C. Addition of Sofosbuvir gave a positive ECCD signal (Figure II-45) while epi-Sofosbuvir produced a negative ECCD signal (Figure II-46). These data are consistent with the data in mixed hexane-DCM solvent system. Of note is the lower intensity of the ECCD signal in DCM, as compared to the signal for both epimers in hexane-DCM (compare Figures II- 43 and II-45 to Figures II-46 and II-47). 108 Figure II-47. ECCD spectra of epi-Sofosbuvir complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C. Furthermore, a variety of solvents were screened. Most of the solvents did not yield any detectable ECCD signal. However, chloroform gave similar results as to what was observed in DCM with slightly lower intensities for both epimers of Sofosbuvir (Figures II- 48 and II-49). It should be noted that none of the previously studied functional groups form an ECCD active complex with porphyrin tweezers or MAPOL. Under this conditions other possible binding events were eradicated and the coordination of phosphine oxide was preserved, as it has by far the strongest interaction with the host system. 109 Figure II-48. ECCD spectra of epi-Sofosbuvir complexed with ZnMAPOL (1 µM) in chloroform at 0 °C. Figure II-49. ECCD spectra of Sofosbuvir complexed with ZnMAPOL (1 µM) in chloroform at 0 °C. 110 0 µL Acetone 10 µL Acetone 20 µL Acetone 30 µL Acetone 50 µL Acetone 75 µL Acetone 100 µL Acetone 600 400 200 0 -200 -400 D C . l o M 380 400 420 440 460 Wavelength (nm) Figure II-50. ECCD spectra of Sofosbuvir (50 equiv) complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C, effect of addition of acetone. Since most chiral phosphine oxides embedded in pharmaceutical products are relatively polar, acetone is normally used to prepared the stock solution of these compounds. Thus, the effect of addition of acetone on the ECCD spectra of both epimers of Sofosbuvir was screened. To this end, to a solution of ZnMAPOL (1µM in DCM) Sofosbuvir (50 equiv) at 0 °C was added. CD spectra of the sample was collected after addition of 0,10, 20, 30, 50, 75 and 100 µL acetone. Interestingly, addition of acetone resulted in a decrease of the intensity of the ECCD signal (Figure II-50). After 100 µL of acetone the signal was completely lost. The same erosion of ECCD signal was observed for epi-Sofosbuvir upon addition of acetone (Figure II-51). As a result the stock solutions should be prepared in either DCM or chloroform. 111 D C . l o M 300 200 100 0 -100 -200 -300 0 µL Acetone 10 µL Acetone 20 µL Acetone 30 µL Acetone 50 µL Acetone 75 µL Acetone 100 µL Acetone 380 400 420 440 460 Wavelength (nm) Figure II-51. ECCD spectra of epi-Sofosbuvir (50 equiv) complexed with ZnMAPOL (1 µM) in dichloromethane at 0 °C, effect of addition of acetone. To propose a working model, the relative sizes of the substituents on the phosphorous atom should be assigned. The O-phenyl is assigned as the smallest group, the substituent on the nitrogen atom as the medium size group and the uracil group as the largest group. Upon binding of the oxygen atom to the zinc, the smallest group (O- phenyl) on the phosphorous atom points towards the bound porphyrin. The remaining medium and large substituents project toward the second porphyrin ring. For the Sofosbuvir the P-helicity is favored as the large group is positioned away from the second porphyrin ring. This arrangement leads to a clockwise twist of the interacting chromophores, yielding a positive ECCD signal, which is in agreement with the measured spectrum (Figure II-52). Following the same analogy, M-helicity is preferred and as a result a negative ECCD signal is predicted for epi-Sofosbuvir, which is confirmed by the recorded ECCD signal (Figure II-53). 112 O O N H HO F O NH O N O P O O Ph Sofosbuvir S: Small M: Medium L: Large O P s M L ECCD measurement ZnMAPOL (1 µM) DCM, 0 °C D C . l o M 200 100 0 -100 -200 380 400 420 440 460 Wavelength (nm) Ph Ph N N Zn NN Ph Ph Ph NN Zn N N Ph PM L S O HH O O M-helicity negative ECCD top view Zn M L P O Zn M-(R) complex disfavored Ph Ph N N Zn NN Ph Ph NN Zn N N Ph PM L S O Ph H H O O P-helicity positive ECCD top view Zn M L P O Zn P-(R) complex favored Figure II-52. Coordination and the working model of Sofosbuvir complexed with ZnMAPOL. P-helicity is favored and positive ECCD signal was observed. 113 O NH O O O P O O Ph N H O N HO F epi-Sofosbuvir S: Small M: Medium L: Large O P s M L ECCD measurement ZnMAPOL (1 µM) DCM, 0 °C D C . l o M 200 100 0 -100 -200 380 400 420 440 460 Wavelength (nm) Ph Ph N N Zn NN Ph Ph Ph NN Zn N N Ph PL M S O HH O O M-helicity negative ECCD top view Zn L M P O Zn M-(R) complex favored Ph Ph N N Zn NN Ph Ph NN Zn N N Ph PL M S O Ph H H O O P-helicity positive ECCD top view Zn L M P O Zn P-(R) complex disfavored Figure II-53. Coordination and the working model of epi-Sofosbuvir complexed with ZnMAPOL M- helicity is favored and negative ECCD signal was observed. 114 II-4 Tuning electronic properties of MAPOL scaffold MAPOL and its zincated analogue, Zn-MAPOL, are powerful hosts for the determination of absolute configuration of multiple functionalities. We sought to further investigate the binding capability of this scaffold by modulation of the electronic properties on the binding site. Namely, the hydrogen bonding affinity was thought to improve by placing fluorine atoms on the biphenol backbone. There are six hydrogens on the biphenyl moiety, and replacing them with fluorine atoms would result in more electron deficient aromatic rings that subsequently leads to more acidic hydroxyl groups. As a result, a stronger hydrogen binding network would be expected. Nonetheless, introducing fluorine in the 6,6’ position of the host backbone results in elevating the rotational barrier of the diaryl bond, thus II-31 was not a suitable backbone for host synthesis. We envisioned a backbone with only four fluorine atoms with hydrogens at 6,6’ positioned to facilitate the free rotation along the diaryl bond (highlighted in bold, Figure II-54). hindered rotation F F F F F F II-31 R OR OR R Figure II-54. Structural features of F6-backbone. The F4-MAPOL II-32 was synthesized in a similar route described for MAPOL. We sought to utilize the improved hydrogen bonding affinity of this host system to form CD active complexes with alcohols as proof of concept. (R)-1-phenylethanol II-33 as a model 115 alcohol was complexed with F4-MAPOL, nonetheless the complex did not lead to an observable ECCD signal. This could be due to weak hydrogen bonding interaction between the host system and the alcohol or low energy difference of the diastereomeric rotomers. With this unsuccessful trial, we resorted to a different modification of the host system. F F F F Ph N H N OH OH N NH Ph N H N Ph Ph HN N Ph Ph F4-MAPOL II-32 Figure II-55. Structure and UV-vis spectrum of F4-MAPOL II-32 (1 µM solution) in hexane at room temperature. Our previous studies with porphyrin tweezer system demonsterated that the high Lewis acidity of the zinc center on porphyrins is substantially influential to derive complex formation with alcohols. TPFP tweezers were successful in formation of ECCD active complexes with diols and epoxy alcohols. To this end, we pursued incorporation of fluorine atoms on the porphyrin ring in order to improve Lewis acidity on the zinc center. In analogy, F30-Zn-MAPOL II-34 was synthesized. This host system was complexed with 116 chiral (R)-1-phenylethanol. Gratifyingly, an ECCD signal was observed for this chiral alcohol in complex with F30-Zn-MAPOL (Figure II-56). The latter modification presents a unique opportunity to further explore the applicability of this molecular receptor for molecules that lack the presence of strong coordinating sites. Further investigation into the generality of this methodology for determination of absolute configuration of chiral alcohols is ongoing. PhF5 F5Ph N Zn N N N OH OH N N Zn N PhF5 PhF5 N PhF5 F5Ph F30-Zn-MAPOL II-34 Figure II-56. Structure and UV-vis spectrum of F30-Zn-MAPOL II-34 (1 µM solution) in hexane at room temperature. 117 OH II-33R Figure II-57. ECCD spectrum of F30-Zn-MAPOL (1 μM) complexed with (R)-1-phenylethanol II-33R (100 equiv) in hexane at 0 ºC. 118 Experimental section II-5 II-5-1 Materials and general instrumentations Anhydrous solvents used for CD measurements were purchased from Aldrich and were spectra grade. Unless otherwise mentioned, solvents were purified as follows. Toluene and CH2Cl2 were dried over CaH2 whereas THF and Et2O were dried over sodium (dryness was monitored by colorization of benzophenone ketyl radical); they were freshly distilled prior to use. NMR spectra were obtained using 500 MHz Varian NMR spectrometers and referenced using the residual 1H peak from the deuterated solvent. Column chromatography was performed using Silicycle 60Å, 35-75 µm silica gel. Pre- coated 0.25 mm thick silica gel 60 F254 plates were used for analytical TLC and visualized using UV light, iodine, potassium permanganate stain, p-anisaldehyde stain or phosphomolybdic acid in EtOH stain. CD spectra were recorded on a JASCO J-810 spectropolarimeter, equipped with a temperature controller (Neslab 111) for low temperature studies, and are reported as Mol. CD / l [nm]. UV-vis spectra were recorded on an Agilent, Cary 100 UV-visible spectrophotometer with scan rate of 100nm/min. Procedure for UV-vis and CD titration of Zn-MAPOL with chiral guest II-5-2 molecules (cyanohydrins, sulfoxides or phosphine oxides) II-5-2-1 General procedure for UV-vis measurement Zn-MAPOL II-4 (1 μL of a 0.001 M solution in anhydrous CH2Cl2) was added to hexane (> 95% n-Hexane) (1 mL) in a 1.0 cm UV-cell to obtain a 1 μM solution. The background spectrum was recorded from 350 nm to 500 nm with a scan rate of 100 119 nm/min. Chiral guest (1 up to 5000 equivalents) from a stock solution (in anhydrous dichloromethane) with different concentration was added as follows: For less than 20 equivalents a stock solution with concentration of 1 µM was used; A 0.01 M solution was used for up to 100 equivalents, a 0.1 M solution was used for up to 100 equivalents, and for more than 1000 equivalents if needed a 1 M solution was used. As a result, the maximum dilution of the spectroscopic sample was kept to less than 0.5% of the initial volume. The UV spectrum was measured after each addition. In order to compensate for the cyanohydrin contribution on the observed UV spectrum, same amount of cyanohydrin was added to the reference UV sample for each titration point. II-5-2-2 General procedure for CD measurement Zn-MAPOL (1 μL of a 0.01 M solution in anhydrous CH2Cl2) was added to hexane (> 95% n-Hexane) (1 mL) in a 1.0 cm CD cell (equipped with cooling water circulation) to obtain a 1 μM solution (for cyanohydrins derived from ketone (11-15) 2 μL Zn-MAPOL was used, resulting in a 2 μM solution for CD measurements). The background spectrum was recorded from 350 nm to 480 nm with a scan rate of 100 nm/min at 0 ºC. Chiral guest (up to 1000 equivalents of a 0.01 M or 0.1 M solution in anhydrous CH2Cl2) was added into the prepared host solution to afford the host/guest complex. The CD spectra were measured immediately (10 accumulations). The resultant ECCD spectra recorded in millidegrees were normalized based on the host concentration (1 μM or 2 μM) to obtain the molecular CD (Mol. CD). 120 II-5-3 Job’s continuous plot analysis to determine the stoichiometry of complex between Zn-MAPOL and chiral guest molecules and binding constant measurements of the complex Job’s plot analysis was achieved from changes in the UV-vis absorbance of Zn- MAPOL upon addition of chiral molecules according to the following procedure: To 1 mL of hexanes in a 1 cm cuvette, Zn-MAPOL (1 μL of a 1 mM stock solution) was added. UV- vis spectrum was collected (350 nm-500 nm). To this Zn-MAPOL solution, chiral guest at (0.25, 0.5, 0.75, 1, 2, 5, 7.5, 10, 20, 30, 40, 50, and 75 equivalents) was added and UV- vis spectra (350 nm-500 nm) were recorded after each addition. Changes in the UV-vis absorbance (∆Aabs) were calculated by subtracting the absorbance at each titration point MAPOL (XZn-MAPOL) multiplied by the change in UV-vis absorbance (∆Aabs) for each from the absorbance of free Zn-MAPOL at 410 or 409 nm. The molar fraction of Zn- titration point was determined and was plotted against the molar fraction of Zn-MAPOL. Binding constants of the complex between Zn-MAPOL and chiral guest molecules were also extracted from changes in the UV-vis spectrum of host system upon addition of chiral guest molecules. Plotting the changes at 410 or 409 nm against the concentration of guest molecule lead to the corresponding curve that the Kassoc values were evaluated employing the following equation reported by Aida and co-workers:52 ΔAbs = (L(1+ KassocX+ KassocA) – (L2(KassocX+ KassocA+1)2 – 4 Kassoc where X and A represent [Guest]total and [Host]total, respectively; L denotes ΔAbs at 100% complexation; L and Kassoc are parameters. 2AXL2)0.5)/2KassocA 121 II-5-4 Synthesis of the host systems Synthesis of Zn-MAPOL II-5-4-1 Ph N N Zn N Ph N Ph OH OH N Ph Ph N N Zn N Zn-MAPOL II-4 Ph Following a standard procedure, to a solution of MAPOL (12.6 mg, 0.010 mmol) in dry dichloromethane (2 mL) was added 5 drops of a saturated solution of zinc acetate [Zn(OAc)2] in methanol (saturated solution of zinc acetate in methanol was prepared from a zinc acetate sample that was washed with dichloromethane, ethyl acetate, and diethyl ether successively, followed by drying under vacuum). Reaction was stirred at room temperature for 12 h. The reaction mixture was passed through a plug of celite. The filtrate was deposited on silica gel and place on a pipet column. DCM was used as eluent to deliver pure zincated MAPOL (Zn-MAPOL 3) (12 mg, 86 %). 1H-NMR (500 MHz, CDCl3); δ 6.22 (s, 2H, OH), 7.56 (m, 6H), 7.64 (m, 8H), 7.68-7.74 (m, 6H), 8.02 (d, J = 7.0 Hz, 4H), 8.09-8.14 (m, 8H), 8.16-8.17 (m, 4H), 8.79 (d, J = 4.0 Hz, 4H), 8.85 (d, J = 5.0, 4H), 8.88 (d, J = 4.0 Hz, 4H), 8.96 (d, J = 5.0 Hz, 2H) ppm. 122 13C-NMR (125 MHz, CDCl3); δ 113.7, 120.3, 121.2, 125.8, 126.4, 126.5, 127.4, 127.5, 130.4, 131.2, 131.9, 132.0, 132.7, 134.2, 134.3, 134.8, 142.5, 142.6, 150.1, 150.2, 150.3, 150.4, 152.6 ppm. II-5-4-2 Synthesis of Zn-MAPOL-OMe II-40 Ph Ph N N Zn N O O Me Me N Zn N N Ph N Ph Ph N Zn-MAPOL-OMe II-35 Ph MAPOL-OMe (10.0 mg, 0.008 mmol) was converted into Zn-MAPOL-OMe (10.0 mg, 90%) according to the procedure described for preparation of Zn-MAPOL II-4. 1H-NMR (500 MHz, CDCl3); δ 2.95 (s, 6H), 7.57 (dd, J1 = 8 Hz, J2 = 7 Hz, 2H), 7.66-7.76 (m, 18H), 8.10 (m, 4H), 8.15 (d, J = 7 Hz, 4H), 8.18 (d, J = 5 Hz, 6H), 8.22 (d, J = 6 Hz 2H), 8.87 (m, 4H), 8.91 (m, 8H), 9.04 (m, 4H). 13C-NMR (125 MHz, CDCl3); δ 61.0, 116.8, 121.0, 121.3, 121.9, 126.4, 126.5, 127.4, 131.7, 131.8, 131.9, 132.1, 132.2, 134.3, 134.4, 135.5, 135.9, 142.7, 142.8, 150.0, 150.1, 150.2, 150.4, 158.2 ppm 123 II-5-4-3 Synthesis of F4-Zn-MAPOL F F CHO OMe OMe CHO F F II-41 BBr3 DCM F F CHO OH OH CHO F F II-42 InCl3 Pyrrole F F F F NH H N OH OH N H NH II-43 Synthesis of F4-Bisphenol-dipyrromethan II-43: To a sample of dimethoxy bisaldehyde II-41 (0.34 g, 1.0 mmol) in dry methylene chloride (10 mL) was added BBr3 [5.0 mmol, 5.0 mL (1.0 M solution in methylene chloride)] drop-wise under atmosphere of argon at 0 ℃. The reaction was stirred at room temperature for 12 h before it was quenched with saturated solution of ammonium bicarbonate (NH4HCO3). The reaction was extracted with methylene chloride, dried (Na2SO4) and volatiles were removed. The crude was taken up in pyrrole (200 mmol, 14.0 mL). To this solution was added InCl3 (0.1 equiv, 0.1 mmol) at room temperature under atmosphere of argon. Reaction was stirred at this temperature for 90 min. NaOH (0.3 g) was added to the reaction and stirring was continued for another hour. Reaction mixture was filtered; excess pyrrole was isolated by distillation 1 at 45-50 °C under vacuum (~0.1 torr). and crude product was purified by column chromatography (1:1 ethyl acetate/hexanes). 300 mg dipyrromethane product II- 43 was isolated, 54 % yield over two steps. 1H NMR (500 MHz, CDCl3) δ 8.39 (s, 4H, NH), 6.79 (m, 4H), 6.71 (m, 2H), 6.21 (m, 4H), 6.14 (m, 4H), 5.97 (s, 2H), 5.57 (s, 2H). 124 F F F F NH H N OH OH N H NH II-43 Ph NH HN O O II-44 Ph 1) NaBH4, THF/MeOH 2) MeCN, TFA 3) DDQ Ph F F F F Ph NH N OH OH N HN Ph Ph N NH HN N Ph F4-MAPOL II-37 Ph Ph F4-MAPOL II-37 was synthesis from II-43 following the procedure described for the synthesis of MAPOL.1 To a solution of freshly prepared 1,9-bis(benzoyl)-5- phenyldipyrromethane II-44 (0.43 g, 1.0 mmol) in dry THF/methanol (10:1, 44 mL) was added NaBH4 (0.766 g, 20.0 mmol) in several portions. After completion of the reduction (TLC 40% EtOAc/Hexanes), saturated NH4Cl (70 mL) and CH2Cl2 (130 mL) were added to the reaction mixture. The organic layer was separated, washed twice with water, dried (Na2SO4), and placed in a 1000-mL round-bottomed flask. Removal of the solvent by rotary evaporation under vacuum yielded the dicarbinol as a foamlike yellow solid. To this crude product, dipyrromethane II-43 (0.27 g, 0.5 mmol) and acetonitrile (400 mL) were added, and the mixture was stirred to achieve a homogeneous yellow solution. TFA (0.930 mL, 12.2 mmol) was added with rapid stirring. After 5 min, DDQ (0.67 g, 3.7 mmol) was added. After stirring for 1 h at room temperature, triethylamine (1.69 mL, 12.2 mmol) was added, and the entire reaction mixture was filtered through a pad of alumina (4 x 8 cm) and washed with CH2Cl2 (~ 500 mL) until the eluant was no longer colored. The 125 resulting porphyrin solution was concentrated, redissolved in CH2Cl2 (50 mL), and passed through a pad of silica (4 x 8 cm) with CH2Cl2 to remove nonporphyrinic pigments. The purple fractions were combined and concentrated to give a purple solid. Precipitation from CH2Cl2/methanol gave the pure product II-37 (35 mg, 5.2% yield). 1H NMR (500 MHz, CDCl3) δ 9.06 (br, 4H), 9.2 (br, 4H), 8.89 (br, 8H), 8.25 (m, 12H), 7.81 (m, 20H), 7.55 (br, 2H), –2.61 (br, 4H). 126 II-5-4-4 Synthesis of F30-Zn-MAPOL F5 F5Ph NH HN O II-44 PhF5 O (((perfluorophenyl)methylene)bis(1H-pyrrole-5,2-diyl))bis ((perfluorophenyl)- methanone) II-44: This compound was synthesized according to previously reported procedures.53 1H NMR (500 MHz dmso-d6) δ 12.70 (s, 2H, NH), 6.87 (br, 2H), 6.03 (s, 1H), 6.01(br, 2H). F5Ph N H N O O Me Me N NH PhF5 N H N PhF5 PhF5 HN N PhF5 F5Ph F30-MAPOL-OMe II-45 F30-MAPOL-OMe II-45 was synthesized using F30-diacyl dipyrromethan II-44 following the standard procedure described for the synthesis of MAPOL.1 To a solution of 127 freshly prepared F30-diacyl dipyrromethan II-44 (0.7 g, 1.0 mmol) in dry THF/methanol (10:1, 44 mL) was added NaBH4 (0.766 g, 20.0 mmol) in several portions. After completion of the reduction (TLC 40% EtOAc/Hexanes), saturated NH4Cl (70 mL) and CH2Cl2 (130 mL) were added to the reaction mixture. The organic layer was separated, washed twice with water, dried (Na2SO4), and placed in a 1000-mL round-bottomed flask. Removal of the solvent by rotary evaporation under vacuum yielded the dicarbinol as a foamlike yellow solid. To this crude product, dipyrromethane I-34 (0.23 g, 0.50 mmol) and acetonitrile (400 mL) were added, and the mixture was stirred to achieve a homogeneous yellow solution. TFA (0.93 mL, 12.2 mmol) was added with rapid stirring. After 5 min, DDQ (0.67 g, 3.7 mmol) was added. After stirring for 1 h at room temperature, triethylamine (1.69 mL, 12.2 mmol) was added, and the entire reaction mixture was filtered through a pad of alumina (4 x 8 cm) and washed with CH2Cl2 (~ 500 mL) until the eluant was no longer colored. The resulting porphyrin solution was concentrated, redissolved in CH2Cl2 (50 mL), and passed through a pad of silica (4 x 8 cm) with CH2Cl2 to remove nonporphyrinic pigments. The purple fractions were combined and concentrated to give a purple solid. Precipitation from CH2Cl2/methanol gave the pure product II-45 (20 mg, 2.1%). 1H NMR (500 MHz, CDCl3) δ 9.17 (br, 4H), 8.96 (s, 8H), 8.84 (s, 4H), 8.14 (m, 4H), 7.66 (dd, J1 =J2 = 7.6 Hz, 2H), 2.97 (s, 6H), –2.81 (br, 4H). 128 PhF5 F5Ph N H N N H N OH OH N NH PhF5 PhF5 HN N F5Ph F30-MAPOL II-46 PhF5 F30-MAPOL II-46 Following the standard procedure described for the synthesis of MAPOL,1 F30-MAPOL-OMe was subjected demethylation of the methoxyl group through reaction with BBr3. The crude was used in the next step without purification. 1H NMR (500 MHz, CDCl3) δ 9.03 (br, 4H), 8.88 (br, 8H), 8.70 (br, 4H), 8.19 (dd, J = 7.5, 1.8 Hz, 2H), 8.12 (dd, J = 7.5, 1.8 Hz, 2H), 7.63 (dd, J = 7.5 Hz, 2H), -2.91 (s, 4H). PhF5 F5Ph N Zn N N N PhF5 PhF5 N OH OH N N Zn N PhF5 F5Ph F30-Zn-MAPOL II-39 129 F30-Zn-MAPOL II-39 was prepared from F30-MAPOL (20 mg) following the procedure described for Zn-MAPOL32 synthesis. 15 mg pure product was prepared 71% yield over two steps. 1H NMR (500 MHz, CDCl3) δ 9.11 (d, J = 4.7 Hz, 4H), 8.92 (q, J = 4.7 Hz, 8H), 8.79 (d, J = 4.7 Hz, 4H), 8.20 (dd, J = 7.5, 1.7 Hz, 2H), 8.14 (dd, J = 7.5, 1.7 Hz, 2H), 7.65 (dd, J1 = J2 = 7.5 Hz, 1H), 6.14 (s, 2H, OH). 130 II-5-5 Synthesis of chiral cyanohydrins Asymmetric silylated cyanohydrines derived from aldehydes and ketones were converted into their corresponding chiral cyanohydrins according to the reported procedure.14 Cyanohydrins II-5R,14 II-5S,12 II-6R,14 II-7S,12 II-10R,14 II-11R,14 II-12R,13 II- 12S,13 II-13R,13 II-13S,13 II-14R,13 II-14S,13 II-15R,13 II-15S13 and II-16R15 were synthesized according to reported procedures. Cyanohydrins derived from ketones: II-12R, II-12S, II-13R, II-13S, II-14R, II-14S, II- 15R, II-15S were synthesized according to method D in the referenced study.13 L- Phenylglycine was applied as catalyst to access the R cyanohydrins and cyanohydrin with S configuration were synthesized using R-phenylglycine as chiral catalyst. 131 II-5-6 Synthesis of chiral sulfoxides Chiral sulfoxides with known configuration on the stereogenic sulfur were synthesized according to reported procedures. Enantiomeric purity of the synthesized chiral sulfoxides was measured using optical rotation and comparison with the known literature values. O S Me II-17R Methyl phenyl sulfoxide (II-17R and II-17S): R enantiomer was synthesized according to the reported procedure.35 [#]% = +138.8 (c = 2.0, EtOH), ee = 85%; Lit.35 +149 (c = 2.0, EtOH); [#]% = –116.5 (c = 1.0, acetone), ee = 81%; Lit.35 –143 (c = 1.0, acetone); S enantiomer was synthesized according to the reported procedure.35 1H NMR (500 MHz CDCl3) δ 7.74 – 7.54 (m, 2H), 7.54 – 7.46 (m, 3H), 2.70 (s, 3H). 13C NMR (126 MHz, CDCl3) δ 145.55, 130.98, 129.30, 123.46, 123.42, 43.88. O S Me II-18R Methyl p–tolyl sulfoxide (II-18R and II-18S): R enantiomer was synthesized according to the reported procedure.37 [#]% = +116 (c = 1.0 ethanol), ee = 77%; Lit.54 +151 (c = 1.0, ethanol) [#]% = –86 (C = 1.0), ee = 57%; Lit.55 –150 (c = 2.0, acetone) S enantiomer was synthesized according to the reported procedure.33 132 1H NMR (500 MHz, CDCl3) δ 7.61 – 7.44 (m, 2H), 7.37 – 7.28 (m, 2H), 2.69 (s, 3H), 2.40 (s, 3H). 13C NMR (126 MHz, CDCl3) δ 142.49, 141.49, 130.02, 123.52, 44.02, 21.41. O S II-19R Ethyl p–tolyl sulfoxide (II-19R and II-19S): R enantiomer was synthesized according to the reported procedure.37 [#]% = +168 (c = 1.0 acetone); Lit.56 +198 (c = 1.0, acetone); ee = 85% [#]% = –119 (c = 1.0 acetone); Lit.57 –202 (c = 1.0, acetone); ee = 60% S enantiomer was synthesized according to the reported procedure.33 1H NMR (500 MHz, CDCl3) δ 7.59 – 7.38 (m, 2H), 7.37 – 7.25 (m, 2H), 2.85 (dq, J = 13.2, 7.4 Hz, 1H), 2.73 (dt, J = 13.2, 7.4 Hz, 1H), 2.39 (s, 3H), 1.16 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 141.34, 140.06, 129.81, 124.19, 50.35, 21.40, 6.04. O S II-20R (R)-p–Tolyl vinyl sulfoxide (II-20R): [#]% = +376 (c = 1.0, acetone) Lit.58 +400 (c = 1.0, acetone); ee = 94% R enantiomer was synthesized according to the reported procedure.37 1H NMR (500 MHz, CDCl3) δ 7.50-7.48 (m, 2H), 7.30-7.28 (m, 2H), 6.56 (dd, J = 16.5, 9.6 Hz, 1H), 6.17 (d, J = 16.5 Hz, 1H), 5.86 (d, J = 9.6 Hz, 1H), 2.39 (s, 3H). 13C NMR (126 MHz, CDCl3) δ 143.03, 141.87, 140.07, 130.13, 124.87, 120.38, 21.43. 133 O S II-21R i–Propyl p–tolyl sulfoxide (II-21R and II-21S): R enantiomer was synthesized according to the reported procedure.37 [#]% = +164 (c = 1.0, acetone); Lit.35 +194 (c = 0.9, acetone); ee = 85% [#]% = –101 (c = 1.0, acetone); Lit.59 –194 (c = 1.0, acetone); ee = 52% S enantiomer was synthesized according to the reported procedure.33 1H NMR (500 MHz, CDCl3) δ 7.51 – 7.38 (m, 2H), 7.31 – 7.27 (m, 2H), 2.78 (h, J = 6.9 Hz, 1H), 2.39 (s, 3H), 1.17 (d, J = 6.9 Hz, 3H), 1.12 (d, J = 6.9 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 141.36, 138.44, 129.55, 125.03, 54.50, 21.41, 15.71, 14.12. O S II-22R Phenyl p–tolyl sulfoxide (II-22R and II-22S): R enantiomer was synthesized according to the reported procedure.37 [#]% = +16.0 (c = 1.0, acetone); Lit.60 +21 (c = 2.0, acetone); ee = 76% [#]% = –12.6 (c = 1.0, acetone); Lit.60 –21 (c = 2.0, acetone); ee = 61% S enantiomer was synthesized according to the reported procedure.33 1H NMR (500 MHz, CDCl3) δ 7.61 – 7.59 (m, 2H), 7.51 – 7.49 (m, 2H), 7.44 – 7.38 (m, 3H), 7.26 – 7.22 (m, 3H), 2.34 (s, 3H). 13C NMR (126 MHz, CDCl3) δ 145.77, 142.44, 141.62, 130.85, 130.01, 129.23, 124.96, 124.66, 21.40. 134 O S Me II-23R (R)-Benzyl methyl sulfoxide (II-23R): R enantiomer was synthesized according to the reported procedure.35 [#]% = –88.1 (c = 1.0, EtOH), ee = 84%; Lit.35(Fernandez, 1992 #22) –105 (c = 6.0, EtOH); 1H NMR (500 MHz, CDCl3) δ 7.38 – 732 (m, 3H), 7.30 – 7.25 (m, 2H), 4.04 (d, J = 12.8 Hz, 1H), 3.91 (d, J = 12.8 Hz, 1H), 2.43 (s, 3H).13C NMR (126 MHz, CDCl3) δ 129.98, 129.61, 128.96, 128.41, 60.30, 37.26. O S Me II-24R t–Butyl methyl sulfoxide (II-24R and II-24S): R enantiomer was synthesized according to the reported procedure.35 [#]% = –3.2 (c = 1.0, acetone), ee = 75%; Lit.35 –4.3 (c = 3.83, acetone) [#]% = +2.8 (c = 1.0, acetone), ee = 66%; Lit.35 +4.3 (c = 1.64, acetone) S enantiomer was synthesized according to the reported procedure.35 1H NMR (500 MHz, CDCl3) δ 2.33 (s, 3H), 1.20 (s, 9H). 13C NMR (126 MHz, CDCl3) δ 52.51, 31.50, 22.43. O S II-25R 135 t–Butyl phenyl sulfoxide (II-25R): R enantiomer was synthesized according to the reported procedure.37 [#]% = +123 (c = 1.0, acetone); Lit.37 +161 (c = 1.0, acetone); ee = 76% 1H NMR (500 MHz, CDCl3) δ 7.66 – 7.53 (m, 2H), 7.50-7.47 (m, 3H), 1.17 (s, 9H). 13C NMR (126 MHz, CDCl3) δ 139.98, 131.10, 128.33, 126.28, 55.74, 22.77. O S O II-26S (S)-Menthyl p-tolyl sulfoxide (II-25S):33 1H NMR (500 MHz, CDCl3) δ 7.58 (d, J = 8.3 Hz, 2H), 7.30 (d, J = 7.9 Hz, 2H), 4.10 (td, J = 10.8, 4.5 Hz, 1H), 2.39 (s, 3H), 2.26 (dd, J = 12.0, 3.1 Hz, 1H), 2.17 – 2.06 (m, 1H), 1.66 (dt, J = 12.0, 2.8 Hz, 2H), 1.50 – 1.41 (m, 1H), 1.38 – 1.28 (m, 1H), 1.25 – 1.14 (m, 1H), 1.08 – 0.96 (m, 1H), 0.94 (d, J = 6.5 Hz, 3H), 0.90 – 0.78 (m, 4H), 0.69 (d, J = 7.0 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 143.07, 142.39, 129.58, 124.95, 80.10, 77.25, 77.00, 76.75, 47.81, 42.91, 33.98, 31.70, 25.18, 23.10, 22.06, 21.49, 20.83, 15.43. O S II-27S (R)-t–Butyl i–propyl sulfoxide (II-27S): R enantiomer was synthesized according to the reported procedure.37 ee = 35% (HPLC, according to the reported method). 136 1H NMR (500 MHz, CDCl3) 2.85 (m, 1H), 1.29 (d, J = 7.1 Hz, 1H), 1.24 (s, 9H), 1.21 (d, J = 7.0 HZ, 1H). 13C NMR (125 MHz, CDCl3) δ 54.05, 44.45, 23.40, 20.43, 15.24. O S S-4l D5 (S)-Phenyl perdeuteriophenyl sulfoxide (S-4l) S enantiomer was synthesized from following precursor according to the reported procedure. O S O O-(1R,2S,5R)-(–)-menthyl-(S)-benzenesulfinate (–)-(S) This compound was synthesized according the reported procedure.29 The diastereomeric mixture product was purified to the titled pure diastereomer according the procedure described by Herbrandson and Dickerson.61 [#]% = –210 (c = 1.45, acetone); Lit. –209.3 (c = 1.45, acetone) to 1H NMR (500 MHz, CDCl3) δ 7.77 – 7.66 (m, 2H), 7.56 – 7.46 (m, 3H), 4.12 (td, J = 10.8, 4.5 Hz, 1H), 2.31 – 2.23 (m, 1H), 2.13 – 2.07 (m, 1H), 1.71 – 1.64 (m, 2H), 1.53 – 1.43 (m, 1H), 1.38 – 1.30 (m, 1H), 1.22 (q, J = 11.8 Hz, 1H), 1.03 (m, 1H), 0.95 (d, J = 6.6 Hz, 3H), 0.85 (d, J = 7.2 Hz, 4H), 0.69 (d, J = 6.9 Hz, 3H). 137 13C NMR (126 MHz, CDCl3) δ 146.00, 131.84, 128.94, 125.01, 80.38, 47.82, 42.93, 33.96, 31.72, 25.19, 23.10, 22.06, 20.84, 15.42. O S D5 II-28S II-28S was synthesized from O-(1R,2S,5R)-(–)-menthyl-(S)-benzenesulfinate (–)-(S) (Shown above) according to the reported procedure.29 1H NMR (500 MHz, CDCl3) δ 7.63 (dt, J = 7.6, 1.6 Hz, 2H), 7.48 – 7.29 (m, 3H). 13C NMR (126 MHz, CDCl3) δ 145.55 (s), 145.36 (s), 131.00, 130.49 (t, J13C–D = 24.7 Hz), 129.27 (s), 128.76 (t, J13C–D = 24.7 Hz), 124.72 (s), 124.33 (t, J13C–D = 24.7 Hz). O S NH2 II-29S (S)-p–toluene sulfinamide II-29S was purchased from Sigma-Aldrich and was used as received. 138 II-5-7 Conformational modeling studies Full optimizations on all conformations of the complexes studied were performed at the semi-empirical Parameterized Model (PM6) using the Spartan-14 software running on a Linux platform. The local minima were then subjected to geometry optimization at the B3LYP/6-31G* (gas phase) level of theory. 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Chemists, particularly organic chemists have aligned their research goals with nature’s creatively designed molecules to pursue a few fundamental objectives: First, they try to provide a synthetic route that can give access to large quantity of naturally occurring molecules that in most cases are isolated in minute quantities. Secondly, nature has created these molecules through precise biochemical transformation, while organic chemists can exercise their creativity by developing new reactions and methodologies. Furthermore, inspired by the structure and chemical composition of natural compounds, a rich platform is available for drug discovery and formulation of new therapeutics and materials.1 Starting with Wohler’s total synthesis of urea, almost two centuries ago, chemical synthesis of naturally occurring molecules has led chemists to make important contributions in areas ranging from drug development to materials science. As one would expect, there are infinite number of possible syntheses for a natural product. Thus, these fascinating molecules have and will provide limitless source of inspiration for chemists. As a result, regardless of how many times a naturally occurring molecule has been targeted for chemical synthesis, they continue to fuel creativity in the field of chemistry.1 One such molecule, which has proved to be a test bed for reaction discovery in the past few years, is Salinosporamide A (III-1, Figure III-1). This molecule with its intriguing chemical structure with multiple fascinating features has led to development of many new 148 methodologies that are not only utilized to access this molecule, but also continue to benefit the field in a broader scope. Salinosporamide A that is structurally related to Omuralide III-3 and Lactacystin III-4,2 was first isolated form a novel microbial source, Salinispora strains, by Fenical and co-worker in 2003.3 Since its isolation, the chemical synthesis of Salinosporamide A has been targeted by many research groups.4-5 Chemist’s fascination with the synthesis of this compound lent itself to the presence of densely functionalized g-lactone core with five contiguous stereocenters in this structure along with the unique biological properties of this compound. Salinosporamide A is a superbly effective proteasome inhibitor against many tumor cell lines. H H N O HO O O Cl Salinosporamide A III-1 H H N O HO O O H N O HO O O Salinosporamide B III-2 Omuralide III-3 HO O H N O S OH AcHN HO2C Lactacystin III-4 Figure III-1. Structure of Salinosporamide A and its closely related natural products. III- 2 Proteasome inhibition by Salinosporamide A The mode of action of Salinosporamide A, related to the inhibition of proteasome, has been studied using yeast 20S proteasome core in complex with the natural product. These investigations are empowered by the availability of the crystal structure of the Salinosporamide A in complex with 20S proteasome.6 The b-lactone moiety of Salinosporamide A (the chemical warhead of this compound) undergoes opening via attack originating from Thr1Og residue (Figure III-2). The tertiary alcohol III-5, revealed 149 after the b-lactone opening event, further acts as a nucleophile to displace the chlorine atom and furnishes the substituted tetrahydrofuran ring III-6. The latter transformations R1 O H N O R3 R2 O OH NH2 H3C H N O R1 O O HO H N O R3 H3C H N N H2 III-5 O Cl inhibitor-proteasome complex H3C H N O R1 H N O R3 O O O N H2 III-6 Figure III-2. Proposed inhibitory mechanism of Salinosporamide A in the proteasome active site. exploit a nonreversible process that covalently links the compound with the proteasome. The high IC50 of Salinosporamide A, 2.6 Kass (M-1s-1) for chymotrypsin and 4.1 Kass (M-1s- 1) for trypsin like proteasomes, indeed finds its origin in the latter strong complex formation with the proteasome active site. III- 3 A survey of the previous total syntheses of Salinosporamide A The highly captivating structural architecture of Salinosporamide A, along with its unique cytotoxicity profile has made this natural product an attractive synthetic target for many research groups in the past few years. Namely, five contiguous stereogenic centers along with a highly functionalized bicyclic framework provides a unique opportunity for reaction discovery and has fueled the inspiration of many chemists to assemble a diverse set of steps to give access to this molecule.4-5 After its isolation in 2003, Salinosporamide A was first synthesized by Corey’s group in 2004.7 Their synthesis commenced with (S)-threonine methyl ester III-7, followed by an elegant set of chemical transformations that gave access to Salinosporamide A (Figure 150 III-3). The key reaction in their synthesis is a Baylis-Hillman cyclization reaction on III-8 to assemble the 2-pyrrolidinone core III-9. Throughout their synthesis, they have developed many different reactions that are utilized and continue to be useful for researchers in the field.8 OMe 6 steps MeO2C H N OOH III-7 MeO2C BnO PMB N O O III-8 Quinuclidine III-9a:III-9b 9:1 Salinosporamide A III-1 10 steps PMB N O PMB N O MeO2C BnO HO III-9a Me + MeO2C BnO Me OH III-9b Figure III-3. First total synthesis of Salinosporamide A with the highlighted Baylis-Hillman cyclization as the key reaction of the synthesis. The second total synthesis of Salinosporamide A is reported by Danishefsky’s group in 2005 (Figure III-4).9 Their synthesis leveraged the stereo-defined face selectivity of pyroglutamate derivative III-10 to introduce substituents on the 2-pyrrolidinone core leading to III-11, which is converted to the natural product in 23 steps. Ph N O O H III-10 HO H N O 5 steps EtO2CO 23 steps Salinosporamide A III-1 III-11 OBn Figure III-4. Danishefsky’s total synthesis of Salinosporamide A. The first biosynthetically inspired synthesis of Salinosporamide A is reported by Pattenden and co-workers in 2006 (Figure III-4, a).10 The key transformation in their 151 synthesis is a nucleophilic attack of amidomalonate moiety in III-13 into the tethered methyl ketone to furnish the substituted pyrrolidinone core III-14. a. Pattenden's biosynthetically inspired synthesis O MeO Me 3 steps O III-12 OBn b. Romo's synthesis H N O MeO2C MeO2C Me OO III-13 AcOH H2O 71% OBn O O BnO CO2H steps NH2 III-15 HO2C BnO N PMB III-16 Cl MeO2C MeO2C Me H N OH O 10 steps Salinosporamide A III-1 OBn III-14 Br N n-Pr OTf 4-pyrrolidinopyridine, i-Pr2NEt, DCM, -10 °C, 6h 25-35% III-17a:III-17b 1:2-3 BnO O BnO O O O PMB N O III-17a Cl + PMB N O III-17b Cl H N MeO2C MeO2C Me OH III-20 Salinosporamide A III-1 steps c. Fukuyama's synthesis CO2H steps III-18 CO2Me CO2Me O O N H III-19 HCO2H, Ac2O THF, O °C aq HCl, acetone 0 °C to rt then 71% crystallization Salinosporamide A III-1 steps Figure III-5. Biosynthetically inspired total syntheses of Salinosporamdie A. This core (III-14) can eventually lead to the natural product with a few synthetic transformations. The latter biosynthetically inspired reaction of amidomalonate into methyl ketone has been utilized by many other research groups. Figure III-5, b and c, 152 highlights the reported total synthesis of Salinosporamide A by Romo11-12 and Fukuyama.13 They have used the biosynthetically inspired route to forge the pyrrolidine core towards the total synthesis of this molecule. In a similar manner, Hatakeyama and coworkers have utilized indium catalyzed Conia-Ene type reaction on III-21 to install the III-22 (Figure 6).14 A sequence of pyrrolidinone core of diastereoselective reactions ultimately converts III-22 to Salinosporamide A. Additionally, many other total and formal syntheses of this natural product have also been reported in the past few years.15-20 21-22 the natural product O AcO PMB N CO2Me CO2Me In(OTf)3 (5 mol%) toluene 110 °C MeO2C MeO2C III-21 PMB N O III-22 OAc steps Salinosporamide A III-1 Figure III-6. Conia-Ene type reaction to assemble the core of Salinosporamide A. III- 4 Our synthetic approach towards the total synthesis of Salinosporamide A Our total synthesis of Salinosporamide A was initiated by Dr. Aman Kulshrestha, a former graduate student in our group.23 Our synthetic approach stems from a methodology that was developed in our lab few years ago.24 In this methodology, an aza- Payne/hydroamination rearrangement of aziridine alcohol III-23 delivers the functionally decorated 2-pyrrolidinone core III-24 that can serve as a versatile platform to give access to the skeleton of Salinosporamide A (Figure III-7, a). As illustrated in Figure III-7, b, deprotonation of aziridinalcohol III-25 triggers the aza/Payne rearrangement via anti opening of the aziridine by the neighboring oxygen nucleophile, leading to the formation 153 of the corresponding epoxide III-25b. This action reveals a nitrogen anion that is stabilized by the tosyl group. The nucleophilic nitrogen consequently adds across the alkyne moiety, aided by protonation of the emerging vinyl anion, yielding the pyrrolidine core III-26 (Figure III-7, b). We surmised that III-26 can be utilized to access Salinosporamide A. a. tandem aza-Payne/hydroamination reaction O S R1 Ts N R5 R1 R2 R4 OR3 III-24 R2 Ts N OHR4 R3 III-23 DMSO R5 b. access to the core of salinosporamide A through aza-Payne/hydroamination OH Ts N III-25 TBSO Ts N TBSO O III-25b O S TBSO TBSO OH S Ts N O III-25a Ts N O III-26 c. our retrosynthetic approach for the synthesis of Salinosporamide A H H N O steps HO O O Cl Salinosporamide A III-1 TBSO 5 Ts N 3 O III-26 C-3 alkylation R N 3 OR 5 H HO III-26-a O C-5 functionalization X 5 R N 3 RO RO III-26-b X = carbonyl surrogate O OR Figure III-7. Our developed methodology that give access to pyrrolidinone core and its exploitation for the synthesis of Salinosporamide A. 154 Our generalized retrosynthesis of Salinosporamide A is illustrated in Figure III-7, c. We envisioned an advanced intermediate such as III-26b that can be converted to the natural product in few steps. The carbonyl surrogate on III-26b is thought to be accessed via C-5 functionalization from III-26a. The epoxide moiety in III-26 would provide a path for C-3 alkylation. III- 5 Synthesis of Salinosporamide A for the diastereoselective aza-Payne/hydroamination reaction (Figure Our generalized retrosynthetic analysis illustrated in Figure III-7. Our synthesis commenced with the preparation of the appropriately functionalized aziridine alcohol III- 25 III-8). Commercially available 3-methyl-2-butanol III-27 was protected as its TBS ether followed by a selective allylic oxidation of the less sterically encumbered methyl group with SeO2 and tBuOOH that delivers allyl alcohol III-28. The allylic oxidation proceeded with partial over oxidation of the alcohol to the corresponding aldehyde, but reduction of the crude mixture delivers the III-28 in excellent yield. Racemic aziridination of the allyl alcohol III- 28 with Chloramine-T in the presence of catalytic bromine source (NBS) gave access to aziridine alcohol III-29.25 Dess-Martin periodinane oxidation of this alcohol yielded the corresponding aziridine aldehyde III-30 with good yield from III-27. Following a methodology developed in our lab, addition of ethynyl magnesium bromide to the latter aldehyde III-30 resulted in the formation of propargyl alcohol III-25 with excellent yield and complete diastereoselectivity.26 Access to III-25 sets the stage for implementation of the key tandem aza-Payne/hydroamination reaction. 155 HO III-27 1.TBSCl, Imid. DMF, rt 2. SeO2, H2O2 then NaBH4 DCM, rt MeOH TBSO III-28 Ts N O OH Chloramine-T (1.0 equiv) NBS (0.2 equiv), ACN, 12 h TBSO MgBr (2.0 equiv) DCM, -78 °C 85%, >99:1 dr TBSO Ts N OH III-29 OH Ts N III-25 DMP, NaHCO3 DCM, 0 °C to rt TBSO III-30 (54%, 4 steps) crystalization Figure III-8. Synthesis of aziridine-alcohol III-25 for implementation of the tandem aza/Payne/hydroamination reaction. Fortunately, the key tandem aza-Payne/hydroamination reaction on III-25 took place with high yield and selectivity and afforded the corresponding N-Ts enamide III-26, which was used in the following step without purification (Figure III-9). Ozonolysis of the resultant enamide III-26 followed by reductive work up of the ozonide complex utilizing dimethyl sulfide (DMS) leads to the 2-pyrrolidinone intermediate III-31 in excellent yield, without the need for purification. The crude product from ozonolysis reaction was directly subjected to the following reaction. C-3 in Salinosporamide A contains a side chain bearing a chlorine atom. Thus we envisioned that a regioselective opening of the epoxide III-31 could give access to the appropriate moiety for installation of chloride at a later stage. In this paradigm, nucleophilic opening of epoxide with bromide was thought to be a suitable strategy, facilitating the installation of the carbon side chain on C-3. Treatment of crude epoxide III-31 with MgBr2 in dry diethyl ether at 0 °C leads to a highly regioselective opening of 156 the epoxide, affording bromohydrin III-32. The anti-opening of the epoxide with bromide and the regioselectivity of the reaction were confirmed based on X-ray crystal structure of this compound (Figure III-9). The reaction proved to be sensitive to the quality of MgBr2, requiring a freshly purchased MgBr2 that was dried under vacuum overnight. OH Ts N TBSO III-25 NaH (2.0 equiv), TMSOI (2.0 equiv) TBSO DMSO rt Ts N O III-26 O3, DCM -78 °C then DMS (5 equiv) TBSO Ts N O O III-31 MgBr2 (3 equiv) Et2O, 0 °C >99:1 dr TBSO Ts N O HO Br III-32 (82%, 3 steps) X-ray stucture of III-32 Figure III-9. tandem aza/Payne/hydroamination and selective epoxide opening, access to decorated 2-pyrrolidinone core. We next turned our attention to the installation of the substituent on C-3. To this end, various metal catalyzed enolization/alkylation sequence of the a-bromo amide in III-32 were envisioned (Figure III-10, a). However, undesired intramolecular displacement of the bromide by the tertiary hydroxyl group led to the formation of epoxide III-31, hampering the introduction of the alkyl group on C-3 under basic conditions. We then resorted to a radical mediated alkylation of a-bromo amide III-32. To that end, III-32 was treated with allyl-tri-n-butyl stannane in the presence of AIBN as a radical initiator. 157 a. envisioned functionalization of C–3 through enolization Ts TBSO N metal mediated TBSO Ts N O enolization HO III-32 Br HO III-33 b. successful radical mediated side chain intallation on C–3 O M alkylation TBSO Ts N HO III-34 O R TBSO Ts N O SnBu3 (1.1 equiv) TBSO HO III-32 Br AIBN (0.15 equiv), PhCH3 dr 75:25 (III-33b:III-33a) 75% Ts N TBSO O + Ts N O OH III-33a OH III-33b DBU (0.1 equiv) DCM, rt dr >99:1 TBSO Ts N O OH III-33b Mg°, MeOH sonication, 1h (two steps) 90% TBSO H N O OH III-34 X-ray stucture of III-34 Figure III-10. Installation of the carbon side chain on C-3 and deprotection of the amide moiety of the 2-pyrrolidinone core. The reaction delivers the a-allylated product III-33 as a ~3:1 diastereomeric mixture. The presence of sterically intrusive substituents at C-4 and C-5 dictates the diastereomeric ratio of the allylation reaction on C-3. We leveraged the steric bias around C-3 to syphon the diastereomeric mixture (III-33a and III-33b) into pure diastereomer III- 33b. Epimerization of C-3 in the presence of catalytic amount of DBU exclusively give access to the thermodynamic diastereomer III-33b. By having the substitution pattern on the C-3 and C-4 aligned with the skeleton of the natural product, manipulation of C-5 was tackled. In this vein, the tosyl protecting group was removed utilizing magnesium metal in 158 methanol with sonication, yielding III-34 in high yield.27 The X-ray crystal structure of III- 34 confirmed the relative stereochemistry of the substituents on the pyrrolidinone core (Figure III-10, dashed box). The reactions described above (from alcohol III-27 to pyrrolidinone III-34) were developed by Dr. Aman Kulshrestha,23 however these reaction were further optimized and performed on large scale as part of contributions of this thesis towards the total syntheis of Salinosporamide A. The C-5 center in the natural product is a quaternary center bearing the b-lactone, thus installation of a carbonyl group on this carbon was targeted. We faced enormous issues with functionalization of this center. We envisioned utilization of anionic, cationic or radical based strategies to alkylate C-5. To pursue any of these strategies, the advance intermediate III-34 or its analogues had to be modified accordingly. For instance, to implement anionic based approaches, C-5 must bear an acidic hydrogen, thus the carbinol carbon on C-5 was elaborated to a variety of carbonyl moieties. Below, we will discuss some of the anionic approaches that were pursued. III-5- 1 A Survey of anionic approaches for C-5 functionalization Anionic approaches, where C-5 acts a nucleophile, were pursued via functional group interconversion of the substituents on C-5 to expose an acidic a hydrogen for deprotonation. To this end, the oxidation state of the carbinol carbon on C-5 was adjusted to ester III-37 or aldehyde III-36 in order to facilitate deprotonation of this center (Figure III-11). Many different reaction strategies were tested with no success to functionalize this carbon to give III-39. In summary, there are two major challenges that preclude 159 functionalization of C-5 center under basic conditions. First, the presence of a tertiary alcohol on C-4 and its facile elimination assisted by the presence of acidic hydrogens on C-3, posed a major challenge for the latter functionalization. Secondly, the kinetic acidity of the C-3 proton prevents deprotonation of the more acidic, albeit sterically hindered, C- 5 proton (Figures III-11 and III-12). TBSO Ts N O OH III-33b HF·Pyr, ACN 15% epimer 6h, 93% on C-3 HO Ts N O OH III-35 DMP, NaHCO3 DCM, 5h, 75% O Ts N O OH III-36 NaH2PO4, NaClO2 tBuOH, H2O, 12h TMSCN, MeOH 12h, 0 °C, 50% then O MeO Ts N O OH III-37 LiHMDS (1.67 equiv) -78 °C then mtheyl chloroformate (1.9 equiv) –78 °C, 28% conv. O Ts N O MeO III-38 Ts N O MeO2C MeO2C OH III-39 target molecule Figure III-11. Functionalization of C-5, an anionic route. As a result, formation of many different side products such as dehydration, functionalization on C-3, retroaldol reaction and degradation of pyrrolidinone core are the unavoidable issues associated with the C-5 functionalization (Figures III-11 and III-12). To remedy this issue, a slight modification of the pyrrolidinone core was considered. In this respect, tertiary alcohol at C-4 was masked as a lactone in III-41. Nonetheless, this did not lead to the desired functionalization on C-5 under various conditions. As a small sample of the reactions that were investigated, formylation of the C-5 with LDA is illustrated in Figure III-12. Undesired deprotonation of C-3 leading to III-45 and elimination to furnish III-44 represent major side products that hampered functionalization at C-5. 160 These results point to the fact that deprotonation at C-3 is an issue, while deprotonation at C-5 suffers from unintended b-elimination, even with a hydroxide present at C-4 (Figure III-12). TBSO Ts N O OH III-33b O3, DCM -78 ∞C then 75% PPh3, -78 ∞C TBSO Ts N O O III-40 OH PCC, MgSO4 0 ∞C, 95 % TBAF, THF 0 °C, 60% HO Ts N O O III-42 O CrO3, H2SO4 H2O, 12h, 0 ∞C to rt then TMSCHN2, MeOH 12 h, 0 ∞C, 60% Ts N MeO2C O III-43 O O LDA –78 °C then HCHO Ts N O Ts N O O TBSO O III-41 HO MeO2C O III-46 O target molecule MeO2C Ts N III-44 O + CO2H (1:1) Ts N MeO2C O III-45 O O OH The group on C-4 poses a challenge towards C-5 functionalization via anionic strategies; Elimination, retroaldol, etc. are the main culprits that impede further functionalization. RO Ts N O OR R Figure III-12. Anionic functionalization of C-5 and the challenges facing this strategy. III-5- 2 Cationic approaches for C-5 functionalization In this strategy, C-5 was envisioned to act as an electrophilic center. To this end, the pyrrolidinone core must be modified to expose an enamide moiety such as III-46 in which C-5 could act as an electrophilic center (Figure III-13). An envisioned strategy was the 161 epoxidation of III-46 that can provide a path towards functionalization of C-5. One possible route could be acid mediated opening of epoxide in III-47 and interception of the intermediate with cyanide (III-48), ultimately leading to functionalization of this center. Towards this goal, III-33b was deprotected under acidic conditions. The alcohol III-49 was then subjected to dehydration condition with Burgess28 and Grieco29 reagents, however, enamide III-46 was not obtained. TBSO Ts N OH III-33b O steps Ts N OH III-46 O O [O] Ts N OH III-47 cationic center NC TMSO Ts N O O TMSCN HO Ts N O O O S NEt3 N O MeO OH III-48 or other dehydrating agents complex mixture of products Ts N O OH III-46 target molecule toluene, rt O KOtBu, THF NaI, DBU DMF O DBU or Ag2(CO3) DCM, rt or 0 °C Ts N O III-52 this product was isolated in nearly qunatitative yield TBSO Ts N O OH III-33b HCl (con.) MeOH TsCl Et3N DCM 64% NaI acetone reflux quant. OH III-49 TsO Ts N OH III-50 Ts N I OH III-51 Figure III-13. Cationic approaches towards C-5 functionalization. 162 In most cases a complex reaction mixture of products was observed. In another approach, an indirect dehydration reaction was pursued. Conversion of alcohol III-49 into a good leaving group followed by elimination was thought to deliver enamide III-46. Nonetheless, treatment of the tolylated analogue III-50 under various basic condition did not lead to the desired elimination to form enamide III-46. In another route, the iodide analogue III-51 was subjected to the elimination reaction.30 Unfortunately, enamide III-46 was not achieved, however, clean conversion of the iodide analogue to III-52 was witnessed, demonstrating once again the facile dehydration of the tertiary alcohol at C-4 (Figure III-13). Reaction at lower temperatures or using different bases did not prevent the undesired elimination. Overman's directed oxidation Me O Si H O Me Br Me N OAc III-53 N O Implementation of the directed oxidation Ts N O MeO2C (TMS)3SiH, AIBN Cu(OAc)2•H2O wet (CH2Cl2) 80 °C N O Me O Si OR O Me Me Me N OAc III-54 OR = OH or OAc (79%) Si Br Cl Et3N, CH2Cl2 MeO2C H O Si Ts N O same condition as described above did not lead to C-5 functionalization MeO2C TMS Br rt, 55% O III-56 Figure III-14. Functionalization of C-5, a radical initiated oxidative strategy. III-55 center with cationic nature RO Ts N O 163 HO III-37 In a different approach, we were inspired by the radical chain oxidation reaction to functionalize the angular carbon on III-53 to access III-54 reported by Overman and coworkers.31 In this transformation, the generated radical results in the homolytic cleavage of the C–H bond, initiating the oxidation of this center that leads to the formation of the corresponding N,O-acetal III-54. It was envisioned that such a transformation could lead to oxidation at C-5 of III-55, which ultimately can deliver the intended alkylation of C- 5. To this end, the tertiary alcohol in III-37 was functionalized with (bromomethyl)- dimethylsilyl as a radical-translocating group leading to III-55 (Figure III-14). Upon subjecting III-55 to the reaction conditions described in the reported procedure.31 the desired functionalization/oxidation of C-5 was not observed and instead a complex mixture of products was generated. The lack of the angular geometry on C-5, as compared to the framework reported by Overman in structure III-53, could be a possible reason for the failure of the latter reaction. III-5- 3 Radical based approaches for C-5 functionalization Having failed to alkylated C-5 via anionic or cationic strategies, we envisioned a homolytic C–H cleavage to deliver the resulting radical intermediate that can be trapped by appropriate group, ultimately leading to the functionalization of this center. One of our early approaches was inspired by Overman’s strategy that is outlined above. 31 In this regard, III-57 was envisioned to undergo translocation of the generated radical in III-57a to that of III-57b which in turn can be trapped by various functionalities such as alkyne moieties to give C-5 functionalized III-58.32 Nonetheless, we could not observe any 164 functionalization at the C-5 center under various conditions with different trapping groups. Removal of the bromide under radical condition and formation of a complex mixture of products was observed (Figure III-15). Ts N O TBSO H TBSO H Ts N O TBSO Ts N O radical initiator Si O Me Me III-57a C5–H abstraction O TMS III-57b O Cl Si Br OTBS Ts N OH III-33b O Si Me Me Br III-57 radical conditions with Various different trapping groups did not result in any functionalization at C-5. OTBS H N O OH III-33b Br I I N OTBS OH III-59 O radical initiator TBSO H N O OH III-59a radical trap Ts N O R R TBSO O TMS III-58 R TBSO radical trap Bn N O C5–H abstraction HO III-59b Figure III-15. Radical based strategies towards installation of functional group at C-5. Alternatively, an aryl iodide was envisioned to deliver the radical on C-5 (Figure III- 15). 2-Iodo benzyl was installed on the amide nitrogen of III-33b to deliver III-59.33 Upon subjecting III-59 to radical initiators it was speculated that the aryl radical III-59a generated from the homolytic cleavage of the iodo group, would undergo hydrogen abstraction from C-5 (6 six atoms away) to give access to the radical species on C-5 (III- 165 59b), which can be trapped by functionalities that can deliver a carbonyl group on this center. Nevertheless, subjecting III-59 to radical mediated reaction conditions did not lead to the desired functionalization on C-5. Instead, a complex mixture of products was observed based on crude NMR analysis. Having faced all the above mentioned unsuccessful efforts, and many other strategies that are not discussed here, to functionalize the C-5 center under anionic, cationic and radical conditions, we resorted to neutral based strategies for the latter functionalization. 166 III-5- 4 Neutral based strategies for C-5 functionalization Since none of the charge or radical based strategies gave access to any functionalization of C-5, the effort resorted to approaches during which this center would stay neutral in order to prevent prevalent side reactions. Direct C–H functionalization can provide a route to avoid generating a reactive center at C-5.34-38 These types of reactions are orthogonal to the general reactivity of molecules that are driven by the polarization due to the presence of functionalities in the molecules or radical centers that are generated therein. As illustrated in Figure III-16, in the shown structure with multiple reactive sites, the blue arrows are indicative of sites that are reactive based on the polarization of the functional groups in the molecule while the red arrows point to the centers that are reactive towards direct C–H functionalization.37-38 One category of direct C–H functionalization is that of active carbene species that insert into carbon-hydrogen bonds, leading to functionalization at the centers highlighted red arrows in Figure (III-16). There are a few factors that control the reactivity of the hydrogen bond for C–H insertion reaction. Among those, the steric and electronic profiles are the most pronounced factors that determine the fate of the active carbene species.37-38 Less sterically hindered and more electron rich C–H bonds are more reactive for insertion reactions (Figure III-16, bottom). Nevertheless, extra factors such as conformational preferences and the distance between the reactive carbene and C–H bond can affect the course of an insertion reaction. In this regard, intramolecular C–H insertion reactions are generally preferred to take place on the hydrogen atoms that are six atoms away from the active carbene. Thus, 167 five membered ring products are expected in most cases where intramolecular C-H insertion reactions ensue. a. sites of reaction based on functional groups vs C-H functionalization O O N Br blue arrows: traditional sites for modification (rely on polarizable functionalities) red arrows: sites for direct C–H functionalization a. controling factor of C–H isertion 1° C–H; sterically favored electronically disfavored 2° C–H; sterically electronically favored favored CH3 EWG EDG 2° C–H; sterically favored electronically disfavored 3° C–H; sterically electronically favored disfavored Figure III-16. General reactivity profile in comparison to the direct C-H functionalization; controlling factor in the C-H insertion reactions. A survey of the reported procedures shows the frequent application of C-H insertion strategies to assemble the skeleton of natural products.34-35, 39 In general, there are two different types of motifs that provide carbene species for the latter functionalization. Reaction of diazo carbonyl compounds in the presence of transition metals40 and deprotonation of vinyl halide followed by subsequence a elimination are common approaches to generate the carbene intermediate.41-43 168 Du Bois and coworkers have employed the reaction of a-diazo carbonyl III-60, in the presence of a rhodium complex for intramolecular C-H insertion on the carbinol center to deliver III-61.39 III-61 is further elaborated to give access to (–)-tetrodotoxin III-62 (Figure III-17, a). The use of vinyl halide for generating active carbenes for C–H insertion is well explored. As an example, depicted in Figure III-17b and c is the deprotonation of vinyl halides III-63 and III-66 to provide the corresponding carbene intermediates for subsequent insertion; the five membered ring products of the latter reaction are further used towards target complex skeletons.42 44 a. Du Bois's synthesis of tetrodotoxin Me Me O O N2 H O PivO III-60 OTBS Rh2(HNCOCPh3)4 O O CCl4 O O PivO O III-61 O Me O Me OH OTBS steps O HO OHO OH NH OH HO HN H2N (–)-tetrodotoxin III-62 b. Grainger's synthesis of AB ring system of ingenol Cl H O TMSO III-63 NaHMDS Et2O, rt 63% O TMSO III-64 O HO HO HO ingenol III-65 H OH c. Wardro's synthesis of lactacysitn Br H iPr III-66 Ph O O KHMDS Et2O, rt N H Ph iPr O O III-67 N H steps (+)-lactacystin III-4 Figure III-17. Application of C-H insertion strategy in the total synthesis of natural products and complex molecules. 169 With the wealth of reports on the direct C–H insertion reactions, and the fact that most of the charge containing strategies failed to deliver C-5 functionalization for our synthesis, the idea was conceived of leveraging C-H insertion strategies to functionalize this center. The approach was to utilize one of the neighboring groups near C-5 as the handle to deliver the active carbene intermediate for insertion into the target C–H bond. It was envisioned that the amide moiety, or the tertiary alcohol on C-4, or the carbinol group on C-5 or the side chain on the C-3 can be utilized as an ancillary group to deliver the active carbene species for functionalization of the C-5 center. One of the early approaches was to utilize the substituents on C-3 to generate a carbene from a-diazo carbonyl (III-72) for C-5 functionalization (Figure III-18). The plan was to convert the advanced skeleton III- 71 into the a-diazo compound III-72. To this end, deprotection of the primary alcohol in III-34 was carried out using HCl (2N aq.) in THF.45 Diol protection of primary and tertiary alcohol in III-68 gave the corresponding acetal product III-69. The allyl group on C-3 was then oxidized to carboxylic acid III-71, followed by acid chloride formation. As reported in the literature,46 treatment of acid chlorides with (trimethylsilyl)diazomethane could give access to the a-diazo carbonyl compounds. In our case however, this reported methodology repeatedly failed to deliver the desire transformation to access III-72. 170 TBSO H N O OH III-34 HCl (2N) THF quant. HO H N OH III-68 O MeO OMe PTSA acetone 75% Boc N O O CO2H O III-71 H N O O O III-69 Boc2O DMAP acetonitrile 91% RuCl3, NaIO4 ACN:CCl4:H2O (1:1:1.5) rt, quant. O Boc N O O III-70 Boc N O N2 O O O III-72 Me Me O O Me H O Boc N O III-72 N2 (COCl)2, DMF DCM then TMSCHN2 THF/ACN different conditions to access compound this diazocarbonyl failed to yield the desired product. metal mediated decomposition of diazocarbonyl, C–H insertion on C-5 Me Me Me O N O O III-73 O target molecule Boc Figure III-18. Functionalization at C-5 center employing metal mediated decomposition of a-diazo carbonyl, a C-H insertion strategy. Next we turned our attention to engage the tertiary alcohol on C-4, group on the C-5 or the amide nitrogen as a handle to deliver the carbene species. Many trials to use the tertiary alcohol as a handle for carbene insertion failed presumably due to the sterically congested hydroxyl group. Difficulties associated with the functionalization of this hindered alcohol precludes installation of any surrogates that would deliver the active carbene intermediate. 171 a. envisioned moieties to deliver carbene for C-5 functionalization R N H O O OH III-74 R N O RO H O III-75 b. lactam nitrogen to deliver carbene for C–H isertion H N O RO OH III-76 C–H insertion H RO N O OH III-77 Br TBSO H N O OH III-78 carbene generation H N O TBSO H TBSO C–H insertion N O OH III-79 target molecule OH III-78a H N O OH III-34 1. NaH (2.5 equiv) DMF TBSO 2. Br Br (1.0 equiv) 0 °C, 89% Figure III-19. Envisioned functionalities to deliver the active carbene to the C-5 center for the C- H insertion reaction. The last resort was to install a carbene surrogate on the amide nitrogen (Figure III- 19, a). The deprotected pyrrolidonone III-34 was derivatized as the vinyl bromide III-78 in high yield. This vinyl bromide was to be converted to the corresponding carbene upon deprotonation of the vinyl proton. We were optimistic that the carbene would only insert into the targeted C–H at C-5, since this bond is fairly electron rich and is in close proximity of the generated carbene (six atoms away) (Figure III-19, b). As such, conditions to 172 generate the requisite carbene III-78a from vinyl bromide III-78 were examined (see Table III-1). Table III-1: Optimization of the C-H insertion reaction at the C-5 center. Br TBSO H N O OH III-78 conditions H TBSO entrya solvent KHMDS (equiv)b temp. (°C)c time (hour) N O OH III-79 conversion (%)d THF THF THF THF THF toluene toluene toluene toluene toluene toluene toluene toluene 3.0 6.0 5.0 2.0 5.0 3.0 3.0 3.0 2.0 5.0 5.0 5.0 3.0 12 12 12 14 6 12 12 12 20 4 6 6 15 –40 to 4 –40 to rt –40 to rt –78 to –30 –78 to rt –78 to 0 –78 to 0 –40 to 0 –78 to 0 –40 to 0 –40 to 0 0 to rt –78 173 40 100 100 0 0 60 10 100 25 100 80 90 85 1 2f 3 4 5g 6 7h 8 9 10 11i 12 13 yield (%) III- 79/III-78 25/53 25/– 30/– –/90 –/85 27/30 –/80 33/– 14/74 43/– 10/36 15/9 53/13 Table III-1 (cont’d). entry solvent KHMDS (equiv)b 14 15 16 17 18 19 20 k21 toluene toluene toluene toluene toluene toluene toluene toluene 3.0 3.0 5.0 6.0 3.0 3.0 3.0 5.0 temp. (°C)c –40 to –20 –40 to –20 –40 to –20 –40 to –20 –40 to –20 –40 to –20 –40 to –20 –40 to –20 time (hour) 12 3.5 3.5 3.5 12 1 2 3.5 conversion (%)d 70 87 90 90 70 75 80 90 yield (%) III- 79/III-78 60/17 58/14 75/10 71/10 51/20 50/25i 53/30i 73/10 [a] Reactions were performed on a 0.1 mmol scale. [b] KHMDS (0.5 M in toluene) was used. [c] The base was added at the first temperature and the reaction was stirred at the second temperature. [d] Conversions are estimated from 1H-NMR using triphenylmethane as an internal standard. [e] Isolated yield unless otherwise noted. [f] KHMDS was added in two portions with 6-hour interval. [g] KHMDS (0.5 M in THF) was used. [h] LiHMDS (1.0 M in hexane) was used. [i] KHMDS was added over 45 minutes at –40 °C. [j] Yields are estimated from 1H-NMR using triphenylmethane as internal standard. [k] Reaction was performed on a 1 mmol scale. Vinyl bromide III-78 was reacted with potassium bis(trimethylsilyl)amide (KHMDS, 0.5 M in toluene) under various conditions. To our delight, the C-H inserted product III-79 was the only isolable product of the reaction along with the remaining starting material III-78. The reaction proved to be sensitive to multiple variables. Reaction time, equivalents of KHMDS, initial reaction temperature, and the temperature during the reaction progress were crucial for the optimal yield of the C–H inserted product. With THF as the solvent, 174 KHMDS as the base and the initial reaction temperature of –40 °C followed by stirring at 4 °C, only 40% conversion was observed based on 1H NMR analysis of the crude mixture. In the same reaction, ~25% product formation was also observed based on 1H NMR analysis (entry 1, Table 1). Increasing the temperature and the equivalents of the base leads to complete consumption of the vinyl bromide, however the yield of the product did not show significant improvement (entry 2-3, Table 1). To our surprise, KHMDS solution in THF instead of toluene did not give any product formation and complete recovery of the vinyl bromide was observed (entry 5, Table 1). Keeping this anomaly in mind, toluene was examined as the reaction solvent. Generally, the conversions and yields improved in comparison to reactions carried out in THF as solvent. Ultimately, we realized that at higher reaction temperatures, despite improved conversions, the yield of the product does not improve (entry 12, Table 1). It should be noted that both starting vinyl bromide and the product are prone to decomposition and side product formation due to the presence of hydroxyl group on C-4 and the acidic hydrogen on C-3. Thus, the reaction should give better results at lower temperatures. The reaction at –78 °C gave good conversion and moderate yield of the product (entry 13, Table 1). The mass balance of the reaction can be improved by further optimization. Using 5.0 equivalents of the base with an initial temperature of –40 °C and stirring at –20 °C for 3.5 hours followed by quenching at –20 °C gave the best conversion and yield of the product along with good mass balance (entry 16, Table 1). The reaction on a larger scale gave similar results with 73% yield of the desired product (83% brsm) (entry 21, Table 1). 175 With this methodology in place, the newly installed olefin on C–5 can be converted to carbonyl group through oxidative cleavage. Additionally, the terminal olefin side chain on C–3 must be oxidatively cleaved to provide the required functionality for installation of the chlorine atom. One of the early efforts was to oxidatively cleave both the terminal and internal double bonds in III-79 in order to shorten the number of steps. It was hypothesized that the aldehyde obtained from the oxidation of the terminal olefin in III-80 will be trapped as its hemiacetal III-81 by the neighboring tertiary alcohol on C-4, opening an avenue for dissecting the bis-aldehyde product for following transformations (Figure III-20). Similar hemiacetal between the aldehyde on C-5 and the hydroxyl on C-4 in III-82 would be less favored due to the smaller ring size. To this end, compound III-80 was subjected to ozonolysis conditions. Unfortunately, a complex mixture of multiple products was observed. This outcome can be due to the fact that the aldehydes can undergo hemiacetal formation with the neighboring tertiary alcohol and the newly formed stereogenic acetal center exist as mixture of stereoisomers. N O TBSO OH III-79 ozonolysis reductive quench O TBSO O N 4 OH III-80 O O O O TBSO HO or O O N O OHC N TBSO O III-81 more favored OH III-82 O less favored a possible pathway for dissecting the two aldehyde groups Figure III-20. Installation of the carbonyl moiety on the C-5 center. 176 To avoid this complication, a sequential oxidative cleavage of the olefin was pursued. We leverage the reactivity differences of the olefins to selectively cleave the terminal in the presence of the internal, tri-substituted double bond in III-79. Osmium tetroxide mediated oxidation led to selective cleavage of the terminal double bond and the resultant aldehyde was trapped as the hemiacetal by the neighboring tertiary alcohol leading to a diastereomeric mixture III-83. This diastereomeric mixture of hemiacetals can be reduce to the diol III-84 or alternately they can be converted to the acetals III-85a/b via acid catalyzed reaction with methanol. TBSO N O HO III-79 OsO4 (0.05 equiv) NaIO4 (3 equiv) THF:H2O (1:1) TBSO N O O NaBH4 (5 equiv) MeOH 0 °C (88%, 2 steps) ~ 1:1 dr OH III-83 MeOH (10.0 equiv) CSA, DCM, rt III-85a/III-85b (1:1) 87% from III-79 TBSO N O OH III-84 OH N O O OMe III-85b O TBSO + TBSO N O OMe III-85a Figure III-21. Selective oxidative cleavage of the terminal olefin. To our surprise, the acetal formation proceeded without any preferences for either diastereomers III-85a or III-85b. One would expect that the geometry of the bicyclic backbone would favor addition of methanol to the oxocarbenium intermediate, generated form reaction of III-83 with acid, from the convex face leading to III-85a as the major product. Nevertheless, the results did not follow this hypothetical speculation. However, 177 this poor diastereoselectivity will not be an issue since the two diastereomers are separable via column chromatography, and furthermore, the stereochemistry of the acetal center is inconsequential since it will eventually be converted into a primary alcohol. The cleavage of the internal olefin to install the carbonyl moiety at C-5 was the next target. It was envisioned that there could be multiple paths to complete the total syntheis of Salinosporamide from III-84 or III-85. One approache is depicted in Figure III-22 ans was the first to be explored. The internal olefin in III-84 subjected to ozonolysis followed by oxidative workup. The resulting acid III-86 was then used to form b-lactone III-86 along with the conversion of the primary alcohol into a chloride to access the advanced intermediate III-87 in 23% yield form III-84.7 From this point the removal of the group on the nitrogen and formation of the carbinol on C-5 in three steps would lead to completion of the syntheis of Salinosporamide A. Among the remaining four steps, the removal of the group on the nitrogen atom is the most untested and could pose a challenge for the completion of the synthesis. Thus, removal of this group on a more readily available analogue was first investigated. O TBSO N O OH III-84 OH 1. O3, H2O, ACN 0 °C 2. NaClO2 H2O:ACN, rt HO2C N O TBSO OH III-86 OH 1. BOPCl, Pyr DCM, rt 2. PPh3Cl2, Pyr ACN, rt ~25% from III-84 TBSO O O O N O III-87 Cl Figure III-22. Oxidative cleavage of the internal olefin, b-lactone ring formation. 178 Acetal III-85 was anticipated to undergo oxidative cleavage to give methyl ester III- 88. This ester is a simpler analogue of III-87 for dealkylation studies of the nirtrogen atom. Direct conversion of the olefin to methyl ester by ozonolysis in methanol followed by treatment of the reaction mixture with acetic anhydride failed to yield III-88.47 Alternatively, a sequential ozonolysis/oxidation-esterification led to the formation of methyl ester III-86 in good yield (Figure III-23). TBSO N O O OMe III-85b 1) O3, DCM then, DMS 73% O O OHC N TBSO O OMe III-89 1.O3, MeOH/DCM NaHCO3, –78 °C 2. Ac2O, Et3N, DCM, rt O O N MeO2C TBSO O OMe III-88b was not observed (complex mixture of products) 1. NaClO2, NaH2PO4 2-methyl-2-butene tBuOH, H2O 2. TMSCHN2 MeOH, 70% O O N MeO2C TBSO O OMe III-88b Figure III-23. Oxidative cleavage of the internal olefin towards deprotection of the pyrrolidinone nitrogen. Our first trial to free the nitrogen atom in III-88 was a radical mediated a-cleavage of the ketone utilizing samarium (II) iodide. This reagent is well known to cleave carbon heteroatom bonds on the alpha position of carbonyl compounds.48-52 However, the 179 reaction proved incompetent for the cleavage of the group on the nitrogen in III-86. The reaction with samarium (II) iodide under various conditions only resulted in the recovery of the starting material III-86 (Fgure III-24). In another approach, we resorted to different strategies that convert the methyl ketone to a functionality that can be easily cleaved from the nitrogen atom. prone to reductive cleavage O O N SmI2 –78 °C MeOH/THF H N O MeO2C TBSO O MeO2C TBSO O OMe III-88b this reduction in various conditions (different equiv., temp., solvent composition, etc.) did not yield the desired product. prone to hydrolytic cleavage OMe III-89b Baeyer-Villiger oxidation O N O O MeO2C TBSO O OMe III-90b mCPBA, DCM: slow reaction, NMR showed only ~20% conversion in 48 h. MMPP, H2O: conversion can be improved to ~ 40% in 48 h. Figure III-24. Strategies for the removal of the group on the pyrrolidinone nitrogen. In this regard, Baeyer-Villiger oxidation of III-88 to give access to the acetate product III-90 was perfomed. It is anticipated that this oxidation would occur next to the methylene group, leading to an acetate group that is prone to hydrolytic cleavage. After screening various oxidizing reagents known to perform the Baeyer-Villiger reaction, we realized that meta-chloroperbenzoic acid can give access to the desired product III-90, but only with 180 low conversion.53 The reaction conversion can be improved to some extent utilizing magnesium monoperoxyphthalate (MMPP) in water.54 However, complete conversion of the starting material was not achieved and the reaction in the presence of a multifold excess of the oxidant and elongated reaction time did not improve the results. Next, we envisioned modification of the olefin in III-85 in such a way that the oxidation of the olefin would lead to a cleavable group remaining on the pyrrolidone nitrogen. To this end, oxidative cleavage of the enone in III-91 was speculated to provide an intermediate that would be capable of being easily transformed to III-93 (Figure III-25). O N O TBSO O OMe III-91 prone to oxidative cleavage oxidative cleavage O O N O O TBSO O III-92 OMe prone to hydrolysis O H N TBSO O O OMe III-93 Figure III-25. An envisioned enone moiety for the facile removal of the group on the pyrrolidinone nitrogen after oxidative cleavage of the internal olefin. The allylic methylene in III-85 was oxidized to the corresponding carbonyl with CrO3 and pyridine, yielding enone III-91 in high yield.55 The X-ray crystal structure of III-91b confirmed the structure as predicted (Figure III-26). 181 two different views TBSO N O O OMe III-85b TBSO CrO3 (20 equiv) pyridine (140 equiv) DCM, reflux up to 98 % O N O O OMe III-91b Figure III-26. Synthesis of the envisioned enone core and its X-ray crystal structure. X-ray stucture of III-91b The oxidative cleavage of III-91 proved challenging. A diverse set of oxidative conditions were tried with no success, indicating the low reactivity of the olefin. The low reactivity of this double bond for oxidation is not surprising since the olefin is conjugated. Furthermore, this olefin is hindered by the bicyclic framework, as well as the neighboring quaternary carbon. As listed in Table 2, most trials resulted in the recovery of the unsaturated imide III-91. Most of the metal mediated conditions failed to give access to the cleaved product.56-57 Ozonolysis, at low temperature after 30 minutes did not give any cleaved products. This strong oxidizing condition at higher temperatures resulted in a complex mixture of decomposed products (entry 7, Table 2). Generally, ozonolysis results in a blue solution, indicating the completion of the reaction and accumulation of excess unreacted ozone. As indicated in the entry 6 Table 2, ozonolysis reaction at –78 °C after 30 minutes (reaction was blue) leads to almost complete recovery of starting materials III-91. However, the presence of the blue color was misleading since with longer reaction time the formation of the cleaved products was observed. 182 Table III-2: Optimization of olefin cleavage towards installation of carbonyl on C-5. O N O TBSO O OMe III-91 oxidative cleavage O OHC N O O TBSO O III-92 OMe OHC H N O TBSO O III-93 OMe entry conditions results (notes) KMnO4, 18-crown-6 then NaIO4, LiOH complex mixture of products OsO4, Oxone, DMF, rt III-91 was recovered (>98%) OsO4, Oxone, DMF, 65 °C III-91 was recovered (>98%) RuCl3, NaClO4 III-91 was recovered (>98%) RuO2, NaIO4, EtOH/H2O/TFA, rt III-91 was recovered (>98%) O3, DCM, –78 °C, 30 minutes III-91 was recovered (>98%) O3, acetone/H2O (95:5), 0 °C complex mixture of products O3, DCM, –78 °C, 90 minutes then DMS 40 conv. (III-92: III-93 ~3:1) basic workup O3, DCM, –78 °C, 3-5 hours then DMS 90 conv. (III-92 85%, III-93 8%) acidic workup O3, MeOH, NaHCO3, –78 °C, 3-5 hours then Ac2O, Et3N 183 complex mixture of products 1 2 3 4 5 6 7 8 9 10 After a number of these trials, the bubbling of ozone for 90 min into the DCM solution of III-91at –78 °C gave a ~40% conversion (entry 8, Table 2). Elongating the reaction time to 3 h led to >90% cleavage of III-91 (entry 9, Table 2). Reductive workup of the ozonolysis reaction with dimethyl sulfide provide the desire aldehyde functionality on C-5. It was noted that the basic workup of the reaction leads to partial cleavage of the di-carbonyl on the pyrrolidone nitrogen delivering III-93. Nevertheless, when the reductive ozonolysis product was quenched with saturated ammonium chloride, the dicarbonyl product III-92 was isolated. Formation of III-93 under basic workup condition demonstrates the susceptibility of the dicarbonyl moiety on III-92 for hydrolytic removal of this group from pyrrolidine nitrogen. Due to the ease of purification, dicarbonyl III-92 was preferred over III-93. With aldehyde III-92 in hand, we postulated the use of our methodology to convert the aldehyde to methyl ester from the Borhan laboratories by treatment of III-92 with oxone in methanol (Figure III-27, top).58-59 Unfortunately, this condition did not result in the formation of methyl ester III-89 (Figure III-27, top). In another approach, III-92 was oxidized under Kraus-Pinnick conditions to deliver that corresponding carboxylic acid III- 94. To our delight, the di-carbonyl on the imide nitrogen was removed under the basic conditions of the Kraus-Pinnick oxidation. This result is in agreement with the product distribution that was observed under basic work up of the ozonlysis reaction (entry 9, Table 2). The crude carboxylic acid III-94 was directly converted to the methyl ester III- 89b with trimethyldiazomethane in methanol in near quantitative yield, and was used in the following Boc-protection reaction without purification (Figure III-27). 184 O OHC N O O TBSO O III-92b OMe oxone (2.5 equiv.) MeOH rt H N O MeO2C TBSO O III-89b complex mixture of products was observed OMe O OHC N O O NaClO2, NaH2PO4 2-methyl-2-butene TBSO tBuOH, H2O O III-92b Fukuyama's synthesis of Salinosporamide A OMe HO2C TBSO O III-94b H N O OMe TMSCHN2 MeOH MeO2C TBSO O III-89b H N O OMe Boc2O, DMAP acetonitrile, rt 67% H H N O HO O O Salinosporamide A (III-1) Cl few steps high overall yield BnO2C TMSO O III-96 Boc N O OMe MeO2C TBSO O III-95b Boc N O OMe Figure III-27. Access to the advanced bicyclic core III-94b, similar to Fukuyama’s intermediate towards Salinosporamide A. The tert-butyloxycarbonyl protection of the crude amide III-89b was carried out following the reported protocol,13 leading to III-95b in good yield (67% over three steps). This advanced scaffold closely resembles III-96, an intermediate in Fukyuama and coworkers’ synthesis of Salinosporamide A (Figure III-27, highlighted box). To that end, a selective de-protection of the TBS-ether in the presence of the Boc protecting group was pursued. It was realized that the Boc group is labile and is removed before TBS deprotection, thus leading to amide III-99. 185 Table III-3: Selective deprotection of the TBS ether in the presence of Boc protecting group. MeO2C TBSO O III-95 Boc N O OMe deprotection condition MeO2C HO O III-97 Boc N OMe O + MeO2C HO O III-98 H N OMe O + MeO2C TBSO O III-99 H N O OMe entrya solvent reagent condition results (notes)b MeOH CSA (1.1 equiv) 0 °C, 3h III-95 was recovered (>98%) THF HCl (2N) 0 °C to rt, 3h III-95 was recovered (>98%) MeOH PTSA (1.1 equiv) rt, 3h MeOH PTSA (1.1 equiv) 0 °C III-98 was the major product (>80%) ~20 % conversion, III-99 as major product THF HF•Pyr 0 °C to rt, 3h complex mixture of products MeOH AcCl 4 °C, 12h ~30 % conversion, III-98 as major product AcOH:THF:H2O (3:1:1) 0 °C III-95 was recovered (>98%) AcOH:THF:H2O (3:1:1) 45 °C, 12h III-95 was recovered (>98%) 1 2 3 4 5 6 7 8 9 HCO2H:THF:H2O (3:6:1) rt rt III-95 was recovered (>98%) III-98 and III-99 (~2:1) as products 10 DCM BF3•Et2O 186 Table III-3 (cont’d) entry solvent reagent condition results (notes)b 11 12 13 14 15 16 17 THF TBAF (5.0 equiv) 0 °C to rt, 5h complex mixture of products MeOH CCl4, ultrasonication 40 °C, 2h III-95 was recovered (>98%) MeOH PPTS (1.1 equiv) rt, 2h III-95 was recovered (>98%) MeOH H2SO4 0 °C to rt, 2h III-95 was recovered (>98%) ACN HF (aq) 0 °C to rt, 3h ~ 50% conv., III-99 as major product EtOH HCl (con.) 1% 0 °C to rt, 3h III-95 was recovered (>98%) DCM Quinolinium fluorochromate rt, 5h III-95 was recovered (>98%) 18 THF 19 THF 20 THF TBAF (5.0 equi), CSA (5.0 equiv) TBAF (2.5 equi), AcOH (7.5 equiv) TBAF (5.0 equi), AcOH (10.0 equiv) 0 °C to rt, 3h ~ 20% conv., III-97 as major product rt, 72h ~75% con., 60% III-97 was isolated 45 °C, 48h >95% con., 85% III-97 was isolated [a] Reaction were performed in a 0.1 mmol scale. [b] Yield and the ratio of the products were estimated based on 1H-NMR analysis of the crude mixture. As illustrated in Table 3, various reported conditions failed to give access to the de- protected alcohol III-99. The main problems were the removal of the Boc group yielding 187 III-99, or the concurrent removal of the TBS and Boc groups leading to III-98. Most of the reported acid catalyzed strategies to remove TBS either resulted in the recovery of III-98, or the dual deprotection of TBS and the Boc group. Ultimately, treatment of III-95 with TBAF in the presence of acetic acid gave access to the desired product III-97 in good yield (entry 20, Table 3).60 Buffered reaction with acid was necessary to prevent removal of the Boc group or transposition of the Boc from the amide nitrogen to the alcohol on C- 5. Deprotected alcohol III-97 was readily oxidized to the aldehyde III-100b employing Dess–Martin periodinane in DCM. Following reported procedures, aldehyde III-100b was reacted with in situ generated cyclohexenyl zinc chloride at –78 °C to give access to the alcohol III-101b with complete stereochemical control in excellent yield.7, 13 Boc N O MeO2C HO O OMe III-97b DMP DCM 0 °C to rt 12 hours 85% H H N HO O O developed methodology by Corey O Salinosporamide A (III-1) Cl Boc N O O MeO2C O OMe III-100b ZnCl THF, –78 °C 82% H Boc N O HO MeO2C O III-101b OMe TFA/H2O THF, 0 °C to 60 °C then NaBH4 H2O/THF 68% H H N O HO MeO2C OH III-102b OH Figure III-28. Completion of the racemic synthesis of Salinosporamide A. The Boc protecting group was removed with exposure to trifluoro acetic acid, resulting in the hydrolysis of the acetal group as well. The crude hemiacetal was treated with 188 NaBH4, leading to diol III-102b in good yield.13 The advanced intermediate III-102b can be converted to Salinosporamide A following the procedure reported by Corey and co- workers.7 Having in hand the synthesis that leads to the racemic total synthesis of Salinosporamide A, we pursued the asymmetric synthesis of this molecule. The following section highlights our total synthesis of (–)-Salinosporamide A. 189 III- 6 The total synthesis of (–)-Salinosporamide A After successful implementation of the racemic synthesis of Salinosporamide A, we sought to carry out the enantiopure synthesis of this natural product. The highly diastereoselective nature of the racemic synthetic route provides a routine strategy to implement the enantiopure synthesis. As such, we envisioned utilization of the enantiopure aziridine III-30 followed by a sequence of diastereoselective reactions, as outlined above, to give access to (–)-Salinosporamide A. Asymmetric aziridination reactions have witnessed tremendous attention in the past few years.61-62 A highly enantioselective aziridination reaction that gives access to tri-substituted in gram scale was desirable as the starting point for the synthesis of (—)-Salinosporamide A. After a thorough survey of the reported protocols for the synthesis of tri-substituted aziridines,63- 67 we chose to use the reported procedure by Cardova and co-workers.67 They have used commercially available proline based chiral catalyst III-105 to synthesize various tri- substituted aziridines from unsaturated aldehydes (Table 4). Fortunately, this reported procedure utilizes unsaturated aldehydes as starting materials and the resulting tosyl protected aziridines are the starting materials for our racemic synthetic route. Cardova and coworkers have demonstrated the applicability of their methodology for various substrates and the aziridine products are isolated in good yield and excellent enantiopurity (Table 4).67 In particular, substrate III-103c, highlighted in Table 4 (entry 3), is similar to the unsaturated aldehyde III-107, requiring for the total synthesis of (–)- Salinosporamide A (Figure III-29). 190 Table III-4: Cardova’s reported asymmetric aziridination of a,b-unsaturated aldehyde. O cat. (20 mol%) TsNHOTs (1.2 equiv) NaOAc (1.5 equiv) Toluene, rt Ts N O R2 R1 III-104 R2 R1 III-103 catalyst N H Ph Ph OTMS III-105 entry aldehydes t (hours) yield (%) 1 2 3 4 5 6 Ph hex Ph O O O O O O III-103a III-103b III-103c III-103e III-103f III-103g 74 68 66 72 66 65 81 77 84 79 83 55 dr 8:1 10:1 >25:1 >25:1 17:1 >25:1 ee (%) 99 99 99 99 99 98 Aldehyde III-107 was synthesized to implement the asymmetric aziridination as the starting point for the synthesis of (–)-Salinosporamide A. For this purpose, protection of commercially available 2-methyl 2-butenol followed by a highly selective oxidation of the less sterically hindered allylic methyl group gives access to aldehyde III-107 in high yield (Figure III-29). 191 TBSCl, Imid. DMF, 0 °C to rt TBSO HO III-27 III-106 nOe H H SeO2, H2O2 DCM, rt 71% (two steps) O TBSO III-107 n O e O TBSO CH2 CH3 nOe III-107 Figure III-29. Synthesis of a,b-unsaturated aldehyde III-107 for implementation of the asymmetric aziridination reaction. With aldehyde III-107 in hand, the enantioselective aziridination reaction, following the procedure reported by Cardova,67 was carried out. Fortunately, the aziridinal (+)-III- 30 was obtained in excellent yield and enantioselectivity (>98% ee). Asymmetric aziridination was carried out in 15 mmol scale (~5 g) without significant deterioration of the reaction efficiency. The reaction, in most cases, was completed within 12 h. To our surprise, decreasing the catalytic loading to 5 mol% (instead of the reported 20 mol%), showed only about 25% conversion of starting material in 24 h. Chiral aziridine aldehyde (+)-III-30 was reacted with ethynyl magnesium bromide at low temperature. Complete diastereoselective reaction was observed with excellent yield. The crystal structure of (+)-III-25 confirmed the absolute configuration of the asymmetric centers (Figure III-30). 192 TBSO III-107 O cat (20 mol%) TsNHOTs (1.2 equiv) NaOAc (1.5 equiv) Toluene, rt, ON Ts N O TBSO 85% yield, >98% ee (+)-III-30 5 g scale Ts N O TBSO (+)-III-30 MgBr (2.2 equiv) DCM, -78 °C >99:1 dr, 88 % TBSO OH Ts N (–)-III-25 X-ray structure of (–)-III-25 Figure III-30. Asymmetric aziridination of a,b-unsaturated aldehyde, access to aziridine-alcohol (–)-III-25. the completion of the Figure III-31 demonstrates total synthesis of (–)- Salinosporamide A. The key tandem aza/Payne hydroamination reaction proceeded with high yield and the resultant enamide product was subjected to ozonolysis followed by the regio- and diastereoselective epoxide opening with MgBr2 to deliver bromohydrin (+)-III- 32 in high yield over three steps. With bromohydrin (+)-III-32 in hand, radical mediated functionalization of C-3 was pursued. Following the optimized methodology for the racemic synthesis, exposure of bromohydrin (+)-III-32 to AIBN as a radical initiator followed by addition of tri-n-butyl allyl stannane in refluxing toluene, resulted in (–)-III- 33a/b as a 3:1 mixture of diastereomers at C-3. This diastereomeric mixture was then epimerized to the thermodynamic product (–)-III-33b in the presence of catalytic DBU. The tosyl protecting group was removed with magnesium powder in methanol under sonication to deliver the pyrrolidinone (–)-III-34. 193 The amide nitrogen of the pyrrolidinone (–)-III-34 was functionalized with vinyl bromide en route to the C-H insertion reaction. The C–H insertion took place in good yield delivering (+)-III-79. Oxidative cleavage of the terminal olefin followed by acetalization gave access to III-85a and (–)-III-85b. From this step the synthesis is shown for the beta diastereomer III-85b, same reactions sequence are carried out with III-85a with comparable results to that of III-85b. Oxidation of the internal olefin led to aldehyde (–)- III-92b. A sequence of Kraus-Pinnick oxidation, esterification of the resultant acid and Boc protection of the amide resulted in (–)-III-95b. The TBS group was removed in the presence of Boc protecting group and the resulting primary alcohol was oxidized to aldehyde (–)-III-100b. Following reported procedures,13 addition of cyclohexenyl zinc chloride to this aldehyde led to (–)-III-101b with complete stereochemical control for the newly formed stereocenters. Treatment of (–)-III-101b with TFA followed by sodium borohydride gave access to diol (–)-III-102b. The methyl ester in (–)-III-102 was hydrolyzed with dimethylaluminium methyltellurolate and the resulting carboxylic acid was converted to the b-lactam (–)-III-108.2, 14 Completion of the synthesis was achieved by chlorination of the primary alcohol in (–)-III-108 to yield (–)-Salinosporamide A (Figure III- 31).7, 13 The NMRs and optical rotation of the synthesized (–)-Salinosporamide A was same as the reported data. 194 1. Me3SOI, NaH, DMSO 2. O3, DCM, –78 °C then DMS, rt 3. MgBr2 (3 equiv), Et2O 86% (over three steps) 0 °C, TBSO Ts N OH Mg°, MeOH sonication, 1h (two steps) 90% (–)-III-33b Br KHMDS (3.0 equiv) (0.5 M in toluene) toluene, –20 °C 72 % (87% brsm) H TBSO HO Br (+)-III-32 O DBU (0.1 equiv) DCM, rt dr >99:1 N O OH (+)-III-79 TBSO Ts N O SnBu3 (1.1 equiv) AIBN (0.15 equiv), PhCH3 dr 75:25 (III-33b:III-33a) 75% TBSO Ts N O TBSO + Ts N O OH III-33a OH (–)-III-33b OsO4 (0.05 equiv) NaIO4 (3 equiv) THF:H2O (1:1) then MeOH (10.0 equiv.) CSA, DCM, rt 85%, (1:1 dr) TBSO N O O α-OMe III-85a β-OMe III-85b OMe CrO3 (20 equiv) pyridine (140 equiv) DCM, reflux 95 % O TBSO N O OH Ts N TBSO (–)-III-25 TBSO H N O OH (+)-III-34 2. Br 1. NaH DMF Br TBSO N O OH (+)-III-78 Boc N O OMe THF, 45 °C 84% Boc N O MeO2C TBSO O (–)-III-95b TBAF AcOH HO MeO2C O (–)-III-97b OMe H H N O HO O O (–)-Salinosporamide A (III-1) Cl 1. NaClO2, NaH2PO4 2-methyl-2-butene tBuOH, H2O 2. TMSCHN2, MeOH 3. Boc2O, DMAP acetonitrile, rt 82% (3 steps) O OHC N O O TBSO O (–)-III-92b OMe O3, –78 °C DCM DMS (5.0 equiv) rt, 68% 85% (brsm) DMP DCM 0 °C to rt 12 h, 80% Boc N O O MeO2C O (–)-III-100b OMe ZnCl THF, –78 °C 92% H HO MeO2C O (–)-III-101b PPh3Cl2 Pyr:ACN 64% HO O H H N O O (–)-III-108 OH [Me2AlTeMe]2 toluene, rt BOPCl Pyridine DCM 68% H HO MeO2C O (–)-III-91b OMe Boc N O TFA/H2O THF, 0 °C to 60 °C then NaBH4 H2O/THF 65% O OMe Boc N OH (–)-III-102 OH Figure III-31. Total synthesis of (–)-Salinosporamide A. 195 In summary, we report the asymmetric total synthesis of (–)-Salinosporamide A. Our synthesis highlights the development of a few key reactions that enabled as to access this natural product. The synthesis is built on a tandem aza/Payne reaction that give access to the pyrrolidine core of the natural product. A C–H insertion reaction stands as one of the key reactions that facilitate a functionalization that most of the routine procedures failed to accomplish. Throughout the synthesis many selective reactions are developed that can be useful in the field 196 III-7 Experimental section III-7-1 General remarks Molecular sieves (4Å) were dried at 160 °C under 0.25 mtorr pressure prior to use. Unless otherwise mentioned, solvents were purified as follows. CHCl3 (amylene stabilized) was purchased from Sigma Aldrich and incubated over 4Å MS for 48 h prior to use. Toluene and CH2Cl2 were dried over CaH2 whereas THF and Et2O were dried over sodium (dryness was monitored by colorization of benzophenone ketyl radical); they were freshly distilled prior to use. NMR spectra were obtained using a 500 MHz and 600 MHz Varian NMR spectrometers and referenced using the residual 1H peak from the deuterated solvent. Waters 2795 (Alliance HT) instrument was used for HRMS 68 analysis with polyethylene glycol (PEG-400-600) as a reference. Column chromatography was performed using Silicycle 60Å, 35-75 µm silica gel. Pre- coated 0.25 mm thick silica gel 60 F254 plates were used for analytical TLC and visualized using UV light, iodine, potassium permanganate stain, p-anisaldehyde stain or phosphomolybdic acid in EtOH stain. 197 III-7-2 Total synthesis of (–)-Salinosporamide A Synthesis of aldehyde III-107: TBSCl, Imid. DMF, 0 °C to rt HO III-27 TBSO III-106 SeO2, H2O2 DCM, rt 71% (two steps) TBSO III-107 O Synthesis of III-106: Commercially available 3-methyl-2-butenol III-27 (12.0 g, 139 mmol) was dissolved in DMF (70 mL). To this solution imidazole (23.7 g, 348 mmol) and TBSCl (23.1 g, 153 mmol) were added at 0 ºC. Reaction was stirred at room temperature for 5 h before it was quenched by addition of water (100 mL). The aqueous layer was extracted with ethyl acetate (3 x 100 mL). Combined organic layers were washed with brine (100 mL) and dried over sodium sulfate. Volatiles were removed under vacuum and the residue was used in the following step without purification. An analytical sample of the crude was passed through silica plug (ethyl acetate/hexane = 5%) to give pure compound (III-106) as a clear oil. 1H-NMR (500 MHz, CDCl3) δ 5.30 (t, 1H, J = 6.5 Hz), 4.22 (d, 2H, J = 6.5 Hz), 1.68 (s, 3H), 1.60 (s, 3H), 0.90 (s, 9H), 0.07 (s, 6H); 13C-NMR (125 MHz, CDCl3) δ 133.9, 124.9, 60.6, 26.4, 26.1, 18.8, 18.2, -4.7. TBSO III-107 O Synthesis of aldehyde 107: SeO2 (9.0 g, 82 mmol, 1.0 equiv) was dissolved in DCM (200 mL) at room temperature. To this mixture was added tBuOOH (70 wt.% in water, 198 30.0 mL, 217 mmol) dropwise at 0 °C. The reaction was stirred at this temperature for 30 min. To this mixture at 0 °C was added crude III-106 (100 mmol) in DCM (100 mL) dropwise. The reaction mixture was stirred at room temperature for 15 h before it was quenched by addition of water (50 mL). The organic layer was separated and the aqueous layer was extracted with DCM (3 x100 mL). The combined organic extract was washed with 10% KOH (4 x 50 mL), sat. NaHCO3 (1 X 100 mL) and water (1 x 50mL). The organic solvent was evaporated under reduced pressure. Crude aldehyde was purified on column chromatography (silica, ethyl acetate/hexane = 5%) to yield the pure aldehyde as a light yellow oil (15.2 g, 71.0 mmol, 71% yield). 1H-NMR (500 MHz, CDCl3) δ 9.40 (s, 1H), 6.50 (m, 1H), 4.48 (dq, J = 5.3, 1.2 Hz, 2H), 1.71 (q, J = 1.3 Hz, 3H), 0.90 (s, 9H), 0.08 (s, 6H). 13C-NMR (125 MHz, CDCl3) δ 194.59, 153.16, 137.71, 60.48, 25.82, 18.28, 9.37, -5.30. TOF MS ES+ (C11H23O2Si): Calc. [M + H]+: 215.1467, Found [M + H]+: 215.1463. O cat (20 mol%) Ts N TBSO III-107 TsNHOTs (1.2 equiv) NaOAc (1.5 equiv) Toluene, rt, ON TBSO (+)-III-30 O in toluene Synthesis of Aziridine (+)-III-30: To a stirred solution of aldehyde III-107 (3.2 g, 15 (S)-(–)-α,α-diphenyl-2- mml, 1.0 equiv) pyrrolidinemethanol trimethylsilyl ether (1.0 g, 3.0 mmol, 0.2 equiv), 4-methyl-N- (tosyloxy)benzenesulfonamide (6.1 g, 18 mmol, 1.2 equiv) and NaOAc (1.8 g, 22 mmol, 1.5 equiv) at room temperature. The resulting reaction mixture was vigorously stirred at (30 mL) were added 199 room temperature for 12 h. After completion of the reaction, the insoluble salts were removed by filtration and the cake was washed with ethyl acetate. After removal of the volatiles the crude product was purified on silica column chromatography (ethyl acetate/hexane = 10%) yielding the aziridine product as light yellow oil (5.1 g, 12 mmol 88%). The enantiomeric excess was determined by HPLC analysis in comparison with racemic material (ODH-column, n-hexane/iPrOH = 98/2, 0.5 mL/min, 250 nm); RT (major enantiomer) = 20.1 min, RT (minor enantiomer) = 21.6 min. ee = >98% 23 = +31.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 9.51 (s, 1H), 7.82 (d, J = 8.4 Hz, 2H), 7.31 (d, J = 8.4 Hz, 2H), 3.73 – 3.63 (m, 3H), 2.43 (s, 3H), 1.40 (s, 3H), 0.82 (s, 9H), -0.02 (s, 3H), -0.04 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 194.50, 144.63, 136.20, 129.69, 127.51, 60.33, 56.80, 50.98, 25.68, 21.64, 18.16, 11.90, -5.51, -5.57. TOF MS ES+ (C18H30NO4SSi): Calc. [M + H]+: 384.1665, Found [M + H]+: 384.1667. Ts N O TBSO (+)-III-30 MgBr (2.2 equiv) DCM, -78 °C >99:1 dr, 88 % OH Ts N TBSO (–)-III-25 Synthesis of aziridine alcohol III-25: To aldehyde III-30 (5.0 g, 13 mmol, >98% ee) in dry DCM (100 mL), at –78 °C under argon was added ethynylmagnesium bromide (0.5 M in THF, 2.1 equiv, 27 mmol, 55 mL) dropwise. The reaction mixture was stirred at this 200 temperature and the reaction progress was monitored by TLC. After 30 min, TLC showed complete consumption of starting material. The reaction was quenched by addition of sat. ammonium chloride (50 mL) at –78 °C. The reaction mixture was warmed up to room temperature. The organic layer was isolated and the aqueous layer was extracted with DCM (3 x 50 mL). The combined organic extract was washed with brine, dried over sodium sulfate, and concentrated under reduced pressure. The crude product was isolated by column chromatography (silica, ethyl acetate/hexane = 10%). Alcohol III-25 was isolated as a white crystalline solid (4.7 g, 88 %, 11.5 mmol, dr >99:1). mp = 82–84° 23 = –11.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 7.81 (d, J = 8.4 Hz, 2H), 7.29 (d, J = 8.4 Hz, 2H), 4.85 (dd, J = 3.6, 2.3 Hz, 1H), 3.67 (dd, J = 11.2, 5.3 Hz, 1H), 3.60 – 3.50 (m, 2H), 3.22 (dd, J = 6.8, 5.3 Hz, 1H), 2.50 (d, J = 2.3 Hz, 1H), 2.41 (s, 3H), 1.52 (s, 3H), 0.81 (s, 9H), -0.04 (s, 3H), -0.06 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 144.28, 137.09, 129.56, 127.26, 80.32, 74.42, 65.25, 60.42, 58.16, 50.88, 25.68, 21.62, 18.11, 12.89, -5.52, -5.60. TOF MS ES+ (C20H32NO4SSi): Calc. [M + H]+: 410.1821, Found [M + H]+: 410.1819. 201 OH Ts N TBSO (–)-III-25 NaH (2.0 equiv), Me3SOI (2.0 equiv) DMSO rt TBSO Ts N O (+)-III-26 O3, DCM -78 °C then DMS (5 equiv) Ts N TBSO TBSO Ts N O O (–)-III-31 O MgBr2 (3 equiv) Et2O, 0 °C >99:1 dr (86%, 3 steps) HO (+)-III-32 Br Synthesis of pyrrolidine (+)-III-26: To trimethylsulfoxonium iodide (Me3SOI, 9.5 g, 43 mmol, 2.0 equiv, dried under vacuum for 12 h) in dry DMSO (100 mL) was added sodium hydride (1.8 g, 43 mmol, 2.0 equiv, 60% in mineral oil) portion-wise at room temperature under argon. The reaction mixture was stirred under argon for 30 min. To this mixture was added alcohol (–)-III-25 (8.8 g, 21 mmol) in dry DMSO (50 mL) drop- wise. The reaction was stirred for 12 h at room temperature before it was quenched with sat. NH4Cl (100 mL). The aqueous layer was extracted with ethyl acetate (3 x 100 mL). The combined organic extract was washed with brine (100 mL), dried over sodium sulfate and concentrated under reduced pressure. Crude product was pure enough and utilized in the following step without further purification. An analytical sample of the crude was passed through a plug of silica (ethyl acetate/hexane = 20%) to give pure (+)-III-26 as a white solid. mp = 74–76° 23 = +81.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 7.60 (d, J = 8.2 Hz, 2H), 7.24 (d, J = 8.2 Hz, 2H), 5.36 (d, J = 1.0 Hz, 1H), 4.74 (d, J = 1.0 Hz, 1H), 4.03 (dd, J = 11.1, 3.4 Hz, 1H), 3.95 – 3.86 (m, 202 2H), 3.39 (d, J = 0.9 Hz, 1H), 2.38 (s, 3H), 1.47 (s, 3H), 0.85 (s, 9H), 0.09 (s, 3H), 0.08 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 143.88, 142.74, 134.62, 129.22, 127.66, 98.50, 66.86, 63.92, 62.72, 62.35, 25.73, 21.64, 17.97, 14.24, -5.55, -5.68. TOF MS ES+ (C20H32NO4SSi): Calc. [M + H]+: 410.1821, Found [M + H]+: 410.1837. TBSO Ts N O O (–)-III-31 Synthesis of pyrrolidinone (–)-III-31: The crude enamide (+)-III-26 (9.0 g, from the above reaction) was dissolved in anhydrous DCM (250 mL). Ozone gas was bubbled through this solution at –78 °C, until the reaction turn to a persistent dark blue color (approximately 30 min). At this point the, bubbling of ozone was ceased and the excess ozone was removed by bubbling nitrogen through the reaction –78 °C until the blue color disappeared. Dimethyl sulfide (DMS, 7 mL) was added to the mixture and the reaction was allowed to warm up to room temperature. Evaporation of the volatiles gave the crude product (–)-III-31, which was used in the next step without purification. An analytical sample of the crude was passed through a plug of silica (ethyl acetate/hexane = 10% to 20%) to afford pure (–)-III-31 as a white solid. mp = 109–111° 23 [a]D = –62.0 (c 1.0, CHCl3). 203 1H-NMR (500 MHz, CDCl3) δ 7.83 (d, J = 8.1 Hz, 2H), 7.29 (d, J = 8.0 Hz, 2H), 4.36 – 4.29 (m, 1H), 4.13 (dd, J = 11.5, 2.5 Hz, 1H), 3.88 (dt, J = 11.5, 1.2 Hz, 1H), 3.33 (s, 1H), 2.40 (s, 3H), 1.58 (s, 3H), 0.78 (s, 7H), 0.05 (s, 3H), -0.02 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 169.09, 145.13, 135.23, 129.59, 128.05, 64.51, 61.36, 59.38, 57.35, 25.54, 21.67, 17.82, 13.51, -5.77, -5.85. TOF MS ES+ (C19H30NO5SSi): Calc. [M + H]+: 412.1614, Found [M + H]+: 412.1624. TBSO Ts N O HO (+)-III-32 Br Synthesis of bromo alcohol (+)-III-32: To the crude epoxide (–)-III-31 from the above reaction (dried under vacuum overnight) under argon was added dry diethyl ether (300 mL) and the reaction flask was place at 0 °C. Anhydrous MgBr2 (dried overnight under vacuum, 3.0 equiv, 17 g) was added to the reaction in portions. The reaction was stirred at this temperature for 3 h, after which TLC showed complete consumption of the epoxide (–)-III-31. The reaction was quenched by slow addition of saturated ammonium chloride (100 mL) at 0 °C. The organic layer was separated and the aqueous layer was extracted with ethyl acetate (3 x 100 mL). The combined organic extract was washed with water and brine, then dried over sodium sulfate. Volatiles were evaporated and the crude product was purified with column chromatography (ethyl acetate/hexane = 10% to 20%). Pure product (+)-III-32 was isolated as a white solid (9.0 g, 18 mmol, 86% yield over 3 steps). 204 mp = 143–145° 23 = +31.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 7.90 (d, J = 8.3 Hz, 1H), 7.31 (d, J = 8.1 Hz, 1H), 4.20 – 4.11 (m, 1H), 3.96 (s, 0H), 2.41 (s, 1H), 1.61 (s, 1H), 0.90 (s, 3H), 0.13 (s, 1H), 0.10 (s, 1H). 13C-NMR (125 MHz, CDCl3) δ 169.09, 145.13, 135.23, 129.59, 128.05, 64.52, 61.36, 59.38, 57.35, 25.54, 21.67, 17.82, 13.51, -5.77, -5.85. TOF MS ES+ (C19H31BrNO5SSi): Calc. [M + H]+: 492.0876, Found [M + H]+: 492.0872. TBSO Ts N O HO Br (+)-III-32 SnBu3 (1.1 equiv) TBSO Ts N AIBN (0.15 equiv), PhCH3 dr 75:25 (III-33b:III-33a) 75% O + TBSO Ts N O OH (–)-III-33b OH III-33a Synthesis of III-33: Bromo pyrrolidinone (+)-III-32 (5.5 g, 11 mmol, 1.0 equiv) was dissolved in toluene (100 mL). The solution was degassed for 10 minutes at 100 °C under argon. AIBN (0.15 mmol, 1.65 mmol, 270 mg) was added to this mixture followed by addition of alllyltributyl stannane (1.1 equiv, 12 mmol, 4.0 mL). The reaction mixture was heated to reflux for 3 h (TlC confirmed completion of the reaction). The solvent was evaporated (in a well ventilated hood) and the crude was purified via column chromatography (ethyl acetate/hexanes = 10 to 20%) yielding an inseparable mixture of distereomers as a 1:3 ratio of III-33a and III-33b (3.8 g, 8.3 mmol, 75% combined yield). 205 TBSO O Ts N OH O DBU (0.1 equiv) DCM, rt dr >99:1 TBSO H N O Mg°, MeOH sonication, 1h (two steps) 90% OH (+)-III-34 TBSO Ts N O + OH III-33a TBSO Ts N OH (–)-III-33b (–)-III-33b The distereomeric mixture of III-33a and III-33b (5.9 g, 13 mmol) was dissolved in dry DCM (150 mL) at room temperature and DBU (0.20 mL, 1.3 mmol, 0.10 equiv) was added. The reaction was stirred at room temperature for 5 h. Solvent was evaporated and the residue was used in the following step without further purification. An analytical sample of the crude was passed through a plug of silica (ethyl acetate/hexane = 20%) to give pure (–)-III-33b as a white solid. mp = 128–130° 23 = –16.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 7.93 – 7.88 (m, 2H), 7.28 (d, J = 8.1 Hz, 2H), 5.88 – 5.76 (m, 1H), 5.07 (dp, J = 16.9, 1.5 Hz, 1H), 5.00 (dq, J = 10.1, 1.3 Hz, 1H), 4.10 (ddd, J = 11.7, 2.9, 1.0 Hz, 1H), 4.00 (dd, J = 2.9, 1.5 Hz, 1H), 3.89 (dt, J = 11.7, 1.3 Hz, 1H), 2.84 (ddd, J = 6.8, 5.7, 1.0 Hz, 1H), 2.48 – 2.41 (m, 1H), 2.40 (s, 3H), 2.21 (dtd, J = 15.2, 8.2, 1.2 Hz, 1H), 1.62 (br, 1H), 1.45 (d, J = 1.1 Hz, 3H), 0.79 (d, J = 1.1 Hz, 9H), 0.06 (d, J = 1.0 Hz, 3H), 0.01 (d, J = 1.0 Hz, 3H). 13C-NMR (125 MHz, CDCl3) δ 174.02, 144.74, 136.59, 135.89, 129.46, 128.14, 116.65, 77.06, 70.69, 62.11, 52.08, 27.95, 25.67, 22.38, 21.67, 17.94, -5.73, -5.90. TOF MS ES+ (C22H36NO5SSi): Calc. [M + H]+: 454.2083, Found [M + H]+: 454.2087. 206 TBSO H N O OH (+)-III-34 Synthesis of amide (+)-III-34: To the crude lactam (–)-III-33b (5.8 g) in methanol (100 mL) was added magnesium powder (5.0 equiv, 65 mmol 1.6 g). The solution was sonicated in an ultra-sonicator bath for 1 h to afford detosylated compound (+)-III-34 (3.50 g, 11.7 mmol, 90% in two steps) as a white solid. mp = 102–104° 23 = +65.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 6.04 – 5.90 (m, 1H), 5.80 (br, 1H), 5.22 – 5.11 (d, J = 17.5 Hz, 1H), 5.09 – 5.03 (d, J = 11.5 Hz, 1H), 3.63 (dd, J = 10.6, 3.8 Hz, 1H), 3.59 (dd, J = 10.6, 5.4 Hz, 1H), 3.38 (ddd, J = 5.1, 3.8, 1.2 Hz, 1H), 2.57 (dddd, J = 11.8, 9.5, 7.9, 4.4 Hz, 1H), 2.47 – 2.35 (m, 2H), 1.37 (s, 3H), 0.86 (s, 10H), 0.03 (s, 3H), 0.03 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 177.25, 137.34, 116.45, 77.57, 65.03, 62.75, 50.80, 28.80, 25.77, 23.06, 18.08, -5.57, -5.63. TOF MS ES+ (C15H30NO3Si): Calc. [M + H]+: 300.1995, Found [M + H]+: 300.1999. TBSO H N O OH (+)-III-34 Br TBSO N O OH (+)-III-78 1. NaH, DMF 0 °C 2. Br Br 207 Synthesis of vinyl bromide (+)-III-78: To a solution of amide (+)-III-34 (2.4 g, 8.0 mmol, 1.0 equiv) in dry DMF (50 mL) at 0 °C was added sodium hydride (0.68 g, 17 mmol, 2.1 equiv). The reaction mixture was stirred at this temperature for 30 min before a solution of ally bromide69 (1.79 g, 8.40 mmol, 1.05 equiv) in dry DMF (10 mL) was added dropwise. The reaction was stirred at 0 °C for 2 h, quenched by the addition of saturated NH4Cl (50 mL) and the aqueous layer was extracted with ethyl acetate (3 x 10 mL). The combined organic extract was washed with brine (50 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate/hexane = 20%), yielding pure product (+)-III-78 as an off white solid (2.9 g, 85%). mp = 64–66° 23 = +59.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 6.12 (q, J = 1.3 Hz, 1H), 5.96 (dddd, J = 17.1, 10.0, 8.5, 5.5 Hz, 1H), 5.15 (dq, J = 17.1, 1.7 Hz, 1H), 5.04 (dtd, J = 10.1, 1.8, 0.8 Hz, 1H), 4.45 – 4.38 (m, 1H), 3.73 – 3.63 (m, 2H), 3.42 (dt, J = 15.2, 1.0 Hz, 1H), 3.14 (t, J = 2.7 Hz, 1H), 2.63 (dddd, J = 14.7, 7.2, 3.6, 1.8 Hz, 1H), 2.59 – 2.54 (m, 1H), 2.41 – 2.31 (m, 1H), 1.76 – 1.73 (m, 3H), 1.62 (br, 1H), 1.43 (s, 3H), 0.84 (s, 9H), 0.02 (s, 3H), 0.01 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 174.74, 137.73, 137.25, 116.28, 104.67, 76.76, 67.92, 59.10, 50.94, 46.29, 29.01, 25.73, 23.26, 17.96, 17.24, -5.63, -5.75. TOF MS ES+ (C19H35BrNO3Si): Calc. [M + H]+: 432.1570, Found [M + H]+: 432.1574. 208 Br TBSO N O OH (+)-III-78 KHMDS (3.0 equiv) (0.5 M in toluene) toluene, –20 °C 73% (87% brsm) N O H TBSO OH (+)-III-79 Synthesis of bicyclic (+)-III-79: To a solution of vinyl bromide (+)-III-78 (0.43 g, 1.0 mmol, 1.0 equiv) in dry toluene (20 mL) at 40 °C was added KHMDS (0.5 M in toluene, 10 mL, 5.0 mmol, 5.0 equiv) dropwise. The reaction mixture was cooled to–20 °C and was stirred at this temperature for 3.5 h. The reaction was then quenched at this temperature with sat. NH4Cl (20 mL). Reaction mixture was brought to room temperature and the organic layer was separated. The aqueous layer was extracted with ethyl acetate (3 x 20 mL). The combined organic layers were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified on column chromatography (ethyl acetate/hexane = 20% to 40%). Pure product (+)-III-79 was isolated as a light yellow oil (257 mg, 0.73 mmol, 73% yield). 23 = +76.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 5.97 (ddd, J = 24.2, 9.4, 5.6 Hz, 1H), 5.33 (s, 1H), 5.14 – 5.06 (m, 1H), 4.99 (d, J = 10.1 Hz, 1H), 4.27 (d, J = 15.5 Hz, 1H), 3.70 (d, J = 10.7 Hz, 1H), 3.66 – 3.59 (m, 1H), 3.49 (d, J = 10.7 Hz, 1H), 2.94 (t, J = 6.7 Hz, 1H), 2.57 – 2.48 (m, 1H), 2.34 – 2.24 (m, 1H), 1.81 (d, J = 1.6 Hz, 3H), 1.43 (s, 3H), 1.42 (br, 1H), 0.85 (d, J = 0.9 Hz, 9H), 0.01 (s, 3H), 0.00 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 177.71, 142.49, 137.89, 120.33, 115.56, 83.34, 80.79, 65.17, 53.43, 53.29, 27.54, 25.73, 21.39, 18.08, 14.68, -5.59, -5.76. 209 TOF MS ES+ (C19H34NO3Si): Calc. [M + H]+: 352.2308, Found [M + H]+: 352.2325. H TBSO N O OH (+)-III-79 OsO4 (0.05 equiv) NaIO4 (3 equiv) THF:H2O (1:1) then MeOH (10.0 equiv.) CSA, DCM, rt 85%, (1:1 dr) TBSO N O TBSO N O + O OMe (+)-III-85a O OMe (–)-III-85b Synthesis of III-85 a and b: To a solution of (+)-III-79 (3.0 mmol, 1.0 g, 1.0 equiv) in THF:H2O (1:1, 60 mL) at room temperature was added OsO4 (0.05 M solution in water, 3.0 mL, 0.15 mmol, 0.05 equiv). To this mixture sodium periodate (1.9 g, 9.0 mmol, 3.0 equiv) was added after 5 min (during which the reaction turns dark). The reaction mixture was stirred at room temperature for 1 h (reaction turns to a milky suspension) during which, TLC indicated complete consumption of starting material. The reaction was quenched by addition of sodium thiosulfate (10 mL, sat. sol.). The reaction mixture was transferred to separatory funnel and extracted with ethyl acetate (4 x 20 mL). The organic layers were combined, washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. To the resulting diastereomeric hemiacetal mixture in DCM (150 mL) were added methanol (2.0 equiv, 6.0 mmol, 0.25 mL) and CSA (0.15 mmol, 35 mg, 0.05 equiv) at room temperature. The reaction mixture was stirred at this temperature for 12 h after which solvent was evaporated and the crude product was purified via column chromatography (silica, ethyl acetate/hexane = 20% to 40%). Two diastereomers were isolated in a 1:1 ratio with a combined yield of 85%. 210 TBSO N O O OMe (+)-III-85a Data for the less polar diastereomer; (+)-III-85a: 450 mg, 1.22 41% yield as a clear oil. 23 = +48.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 5.38 (q, J = 1.5 Hz, 1H), 4.90 (dd, J = 6.3, 3.9 Hz, 1H), 4.12 (d, J = 15.3 Hz, 1H), 3.70 (d, J = 10.8 Hz, 1H), 3.56 (ddt, J = 15.3, 2.5, 1.2 Hz, 1H), 3.36 (d, J = 10.8 Hz, 1H), 3.30 (s, 3H), 3.11 (d, J = 8.6 Hz, 1H), 2.71 (ddd, J = 14.0, 6.2, 1.2 Hz, 1H), 2.00 (ddd, J = 14.0, 8.6, 3.9 Hz, 1H), 1.77 (d, J = 1.5 Hz, 3H), 1.51 (s, 3H), 0.85 (s, 9H), 0.01 (s, 3H), -0.00 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 177.68, 138.58, 121.30, 105.15, 89.60, 82.22, 65.32, 55.47, 54.60, 52.79, 34.33, 25.78, 19.59, 18.15, 14.76, -5.51, -5.78. TOF MS ES+ (C19H34NO4Si): Calc. [M + H]+: 368.2257, Found [M + H]+: 368.2267. TBSO N O O OMe (–)-III-85b Data for the more polar diastereomer; (–)-III-85b: 480 mg, 1.30 mmol, 44% yield as a clear oil. 23 = –77.0 (c 1.0, CHCl3). 211 [a]D 1H-NMR (500 MHz, CDCl3) δ 5.47 (q, J = 1.8 Hz, 1H), 4.88 (d, J = 5.0 Hz, 1H), 4.09 – 4.04 (m, 1H), 3.70 (d, J = 10.7 Hz, 1H), 3.68 – 3.63 (m, 1H), 3.37 (d, J = 10.7 Hz, 1H), 3.10 (s, 3H), 2.92 (d, J = 8.0 Hz, 1H), 2.43 (d, J = 13.2 Hz, 1H), 2.07 (ddd, J = 13.2, 8.1, 5.0 Hz, 1H), 1.77 (d, J = 1.5 Hz, 3H), 1.44 (s, 3H), 0.84 (s, 9H), 0.01 (s, 3H), 0.00 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 177.19, 138.58, 122.36, 104.16, 90.74, 83.16, 65.51, 54.26, 53.77, 53.66, 34.70, 25.76, 20.88, 18.09, 14.48, -5.51, -5.87. TOF MS ES+ (C19H34NO4Si): Calc. [M + H]+: 368.2257, Found [M + H]+: 368.2267. TBSO N O O OMe (–)-III-85b CrO3 (20 equiv) pyridine (140 equiv) DCM, reflux up to 98% O N O TBSO O OMe (–)-III-91b Synthesis of enone (–)-III-91b: To CrO3 (22 mmol, 2.2 g, 20 equiv) in dry DCM (30 mL) at 0 °C was added pyridine (154 mmol, 12.4 mL, 140 equiv) dropwise under argon. The reaction mixture was stirred at this temperature for 10 min followed by stirring at room temperature for 30 min. To this mixture at 0 °C was added (–)-III-85b (1.1 mmol, 0.40 g, 1.0 equiv) in DCM (30 mL). the reaction mixture was refluxed for 24 h. After this period of time, the reaction mixture was cooled to room temperature and volatiles were removed under pressure. The residue was re-dissolved in ethyl acetate (100 mL) and the precipitates were removed by filtration through celite. The product was purified via column chromatography (silica, ethyl acetate/hexane = 20%), yielding (–)-III-91b as a white solid (305 mg, 0.8 mmol, 73%). 212 mp = 161–164° 25 = –210.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 6.78 (q, J = 1.7 Hz, 1H), 4.87 (d, J = 4.8 Hz, 1H), 3.92 (d, J = 10.5 Hz, 1H), 3.39 (d, J = 10.5 Hz, 1H), 3.12 (d, J = 8.1 Hz, 1H), 3.05 (s, 2H), 2.46 (d, J = 13.2 Hz, 1H), 2.16 (ddd, J = 13.2, 8.1, 4.8 Hz, 1H), 1.89 (d, J = 1.7 Hz, 4H), 1.47 (s, 3H), 0.85 (s, 7H), 0.03 (s, 3H), 0.03 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 172.51, 168.68, 141.31, 136.12, 104.01, 88.66, 76.41, 66.80, 56.73, 54.67, 35.61, 25.75, 20.42, 18.13, 11.13, -5.64, -5.92. TOF MS ES+ (C19H32NO7Si): Calc. [M + H]+: 382.2050, Found [M + H]+: 382.2060. O N O TBSO O OMe (–)-III-91a (–)-III-91a (282 mg, 0.74 mmol, 74% yield) was synthesized as an off white solid from (+)- III-85a (386 mg, 1.0 mmol), according to the procedure described above for the synthesis of (–)-III-91b. mp = 75–78° 23 = –12.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 6.74 (q, J = 1.6 Hz, 1H), 4.89 (dd, J = 6.3, 4.0 Hz, 1H), 3.92 (d, J = 10.7 Hz, 1H), 3.39 (d, J = 10.7 Hz, 1H), 3.33 (d, J = 9.2 Hz, 1H), 3.29 (s, 3H), 2.71 (ddd, J = 14.3, 6.3, 1.1 Hz, 1H), 2.14 (ddd, J = 14.2, 9.0, 4.0 Hz, 1H), 1.88 (d, J = 1.6 Hz, 3H), 1.53 (s, 3H), 0.85 (s, 9H), 0.03 (s, 3H), 0.02 (s, 3H). 213 13C-NMR (125 MHz, CDCl3) δ 173.40, 169.09, 141.66, 136.15, 105.16, 87.18, 75.69, 66.48, 57.90, 55.76, 35.44, 25.72, 19.05, 18.14, 11.23, -5.64, -5.85. TOF MS ES+ (C19H32NO5Si): Calc. [M + H]+: 382.2050, Found [M + H]+: 382.2051. O N O TBSO O OMe (–)-III-91b O3, –78 °C DCM DMS (5.0 equiv) rt, 68% 85% (brsm) O OHC N O O TBSO O (–)-III-92b OMe Synthesis of dicarbonyl (–)-III-92b: (–)-III-91b (0.25 g, 0.65 mmol) was dissolved in DCM (50 mL). To this mixture at –78 °C was bubbled ozone for 3 h. The excess ozone was removed via bubbling nitrogen at this temperature. To the reaction mixture was added dimethyl sulfide (0.25 mL, 3.3 mmol, 5.0 equiv) at –78 °C. The reaction mixture was warmed up to room temperature and stirred for 3 h. Saturated aqueous ammonium chloride (20 mL) was added to the reaction mixture and the organic layer was separated. The aqueous layer was extracted with DCM (3 x 20 mL). Organic layers were combined, washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified via column chromatography (ethyl acetate/hexane = 10% to 20%) to give the pure product (–)-III-92b as an off white solid (180 mg, 0.44 mmol, 68% yield). Starting material (–)-III-91b (64 mg, 17%) was recovered. mp = 79–81° 23 = –96.0 (c 1.0, CHCl3). 214 [a]D 1H-NMR (500 MHz, CDCl3) δ 9.50 (s, 1H), 4.99 (d, J = 4.7 Hz, 1H), 4.18 (d, J = 11.3 Hz, 1H), 4.11 (d, J = 11.4 Hz, 1H), 3.31 (s, 3H), 2.93 (dd, J = 8.2, 0.7 Hz, 1H), 2.45 (dd, J = 13.4, 0.7 Hz, 1H), 2.39 (s, 3H), 2.21 (ddd, J = 13.2, 8.2, 4.8 Hz, 1H),1.42 (s, 3H), 0.83 (s, 9H), 0.04 (s, 3H), 0.02 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 196.3, 193.8, 176.4, 168.2, 104.0, 89.8, 75.1, 60.1, 55.0, 51.3, 35.1, 26.2, 25.7, 25.6, 20.0, 17.9, -5.8, -5.9. TOF MS ES+ (C19H32NO7Si): Calc. [M + H]+: 414.1948, Found [M + H]+: 414.1950. O OHC N O O TBSO O (+)-III-92a OMe (+)-III-92a (170 mg, 0.40 mmol, 82%) was synthesized from (–)-III-91a (191 mg, 0.50 mmol), as a white solid according to the procedure described above for the synthesis of (+)-III-92b. mp = 71–74° 23 =+80.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 9.47 (s, 1H), 5.10 – 5.03 (m, 2H), 4.18 (d, J = 11.4 Hz, 1H), 4.06 (d, J = 11.4 Hz, 1H), 3.30 (s, 3H), 3.09 (ddd, J = 9.5, 1.5, 0.8 Hz, 1H), 2.64 (ddd, J = 14.4, 6.2, 1.6 Hz, 1H), 2.40 (s, 3H), 2.25 (ddd, J = 14.4, 9.4, 3.8 Hz, 1H), 1.53 (s, 3H), 0.84 (s, 9H), 0.05 (s, 3H), 0.02 (s, 3H). 215 13C-NMR (125 MHz, CDCl3) δ 196.8, 193.9, 177.5, 168.4, 105.2, 89.8, 75.0, 59.4, 55.8, 52.7, 35.4, 26.3, 25.6, 19.0, 17.9, -5.8. TOF MS ES+ (C19H32NO7Si): Calc. [M + H]+: 414.1948, Found [M + H]+: 414.1936. O OHC N O O TBSO O (–)-III-92b OMe NaClO2, NaH2PO4 2-methyl-2-butene tBuOH, H2O HO2C TBSO O III-94b Boc N O OMe TMSCHN2 MeOH 0 °C to rt MeO2C TBSO O III-89b Boc N O OMe Boc N O Boc2O, DMAP acetonitrile, rt 82% (3 steps) MeO2C TBSO O (–)-III-95b Synthesis of (–)-III-95b: Dicarbonyl (–)-III-92b (0.17 g, 0.41 mmol, 1.0 equiv) was dissolved in tert-butyl alcohol (7 mL). To this solution at room temperature was added 2- methyl 2-butene (0.1 mL, 1.0 mmol) followed by addition of sodium chlorite (1.7 mmol, OMe 3.5 mL of 0.5 M aqueous solution) and sodium phosphate mono basic (3.5 mmol, 3.5 mL of 1.0 M aqueous solution). The reaction was stirred at room temperature for 24 h. aqueous saturated NH4Cl (10 mL) was added to the reaction mixture and the solution was transferred to a separatory funnel. The aqueous layer was extracted with ethyl acetate (5 x 20 mL). The combined organic extracts were dried over sodium sulfate, filtered and concentrated under reduced pressure. Crude acid (–)-III-94b was used in the following step without further purification. 216 The crude carboxylic acid (–)-III-94b was dissolved in methanol (10 mL). To this solution at 0 °C trimethylsilyl diazomethane (2.0 M in diethyl ether, 10 equiv, 4.0 mmol, 2.0 mL) was added dropwise under nitrogen. The reaction mixture was stirred for 12 h, during which the reaction mixture was allowed to warm up to room temperature. Aqueous NaHCO3 (sat., 20 mL) was added to the reaction mixture and the aqueous layer was extracted with ethyl acetate (4 x 10 mL). The combined organic extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. The resulting methyl ester (–)-III-89b was used in the next step without further purification. The crude methyl ester (–)-III-89b was dissolved in dried acetonitrile (10 mL). To this mixture under nitrogen at 0 °C were added Boc-anhydride (0.72 g, 3.2 mmol, 8.0 equiv) and DMAP (49 mg, 0.4 mmol, 1.0 equiv). The reaction was stirred at 0 °C for another 5 h upon which, 1H-NMR confirmed complete consumption of the starting material. Volatiles were removed under pressure and the residue was directly loaded on to a column (basified silica with Et3N, ethyl acetate/hexane = 10% to 20% as eluent) to yield the Boc protected product (–)-III-95b as a light yellow oil (160 mg, 0.34 mmol, 82% yield from (– )-III-92b). Data for (–)-III-95b: 23 = –95.0 (c 0.5, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 4.89 (d, J = 5.0, 1H), 4.37 (d, J = 10.8 Hz, 1H), 4.12 (d, J = 10.8 Hz, 1H), 3.74 (s, 3H), 3.21 (s, 3H), 2.76 (d, J = 8.1 Hz, 1H), 2.43 (d, J = 13.3 Hz, 1H), 2.16 (ddd, J = 13.3, 8.1, 5.0 Hz, 1H), 1.48 (s, 9H), 1.44 (s, 3H), 0.82 (s, 9H), 0.02 (s, 3H), -0.01 (s, 3H). 217 13C-NMR (125 MHz, CDCl3) δ 173.6, 167.6, 150.0, 103.9, 86.9, 82.9, 62.2, 55.0, 52.1, 51.6, 35.5, 28.2, 28.0, 25.7, 20.5, 18.0, -5.6, -5.8. TOF MS ES+ (C22H40NO8Si): Calc. [M + H]+: 474.2523, Found [M + H]+: 474.2519. MeO2C TBSO O (+)-III-95a Boc N O OMe (+)-III-95a (120 mg, 0.25 mmol, 77%) was synthesized from the corresponding dicarbonyl (+)-III-92a (140 mg, 0.33 mmol) according to the procedure described above for the synthesis of (–)-III-95b as a light red colored oil. 23 = +28.0 (c 0.5, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 4.97 (dd, J = 6.0, 4.3 Hz, 1H), 4.30 (d, J = 10.8 Hz, 1H), 4.14 (d, J = 10.8 Hz, 1H), 3.71 (s, 3H), 3.26 (s, 3H), 2.90 (dd, J = 9.3, 1.5 Hz, 1H), 2.65 (ddd, J = 14.0, 6.0, 1.5 Hz, 1H), 2.15 (ddd, J = 14.0, 9.3, 4.3 Hz, 1H), 1.55 (s, 3H), 1.47 (s, 9H), 0.82 (s, 9H), 0.02 (s, 3H), 0.01 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 174.75, 167.76, 149.65, 105.10, 85.89, 83.45, 61.57, 55.61, 52.66, 52.01, 35.91, 27.99, 27.87, 25.65, 19.85, 17.94, -5.65, -5.71. TOF MS ES+ (C22H40NO8Si): Calc. [M + H]+: 474.2523, Found [M + H]+: 474.2522. 218 Boc N O TBSO MeO2C O (–)-III-95b OMe TBAF, AcOH THF, 45 °C 84% HO MeO2C O (–)-III-97b Boc N O OMe Synthesis of (–)-III-97b: To silyl protected alcohol (–)-III-95b (1.0 equiv, 0.30 mmol, 0.14 g) in dry THF (10 mL) under argon at 0 °C were added acetic acid (20.0 equiv, 6.00 mmol, 0.33 mL) and TBAF (1.0 M in THF, 10 equiv, 3.0 mmol, 3.0 mL) dropwise. The reaction mixture was placed in a pre-heated oil bath at 45 °C. The mixture was stirred at this temperature for 48 h, upon which TLC showed completion of reaction. The reaction mixture was cooled to room temperature and was quenched by the addition of sat. NaHCO3 (10 mL). The aqueous layer was extracted with ethyl acetate (4 x 10 mL). The combined organic extract was washed with brine (10 mL), dried over sodium sulfate and concentrated under reduced pressure. The resulting crude product was purified on column chromatography (ethyl acetate/hexane = 50%) to yield the pure alcohol (–)-III- 97b as a white sticky solid (90 mg, 0.25 mmol, 84%). 23 = –115.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 4.89 (d, J = 4.9 Hz, 1H), 4.41 (d, J = 12.2 Hz, 1H), 4.11 (d, J = 12.2 Hz, 1H), 3.75 (s, 3H), 3.20 (s, 3H), 2.96 (d, J = 8.0 Hz, 1H), 2.40 (d, J = 13.2 Hz, 1H), 2.13 (ddd, J = 13.2, 8.0, 4.9 Hz, 1H), 1.98 (br, 1H), 1.48 (s, 12H). 13C-NMR (125 MHz, CDCl3) δ 173.94, 167.42, 150.58, 103.89, 86.93, 83.62, 62.33, 55.00, 52.28, 51.21, 35.45, 27.98, 22.65, 20.73. TOF MS ES+ (C16H26NO8): Calc. [M + H]+: 360.1658, Found [M + H]+: 360.1671. 219 HO MeO2C O (+)-III-97a Boc N O OMe (+)-III-97a (61 mg, 0.17 mmol, 81%) was synthesized according to the procedure described above for the synthesis of (–)-III-97b as a white sticky solid. 23 = +65.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 5.03 – 4.96 (m, 1H), 4.38 (dd, J = 11.9, 5.3 Hz, 1H), 4.18 (dd, J = 11.9, 6.3 Hz, 1H), 3.74 (s, 3H), 3.28 (s, 3H), 3.01 (ddd, J = 9.3, 1.6, 0.7 Hz, 1H), 2.68 (ddd, J = 14.1, 6.0, 1.6 Hz, 1H), 2.18 (ddd, J = 14.1, 9.3, 4.1 Hz, 1H), 1.89 (br, 1H), 1.61 (s, 3H), 1.50 (s, 9H). 13C-NMR (125 MHz, CDCl3) δ 174.86, 167.54, 150.03, 105.10, 86.00, 84.01, 61.48, 55.61, 52.46, 52.19, 35.79, 27.91, 27.63, 20.14. TOF MS ES+ (C16H26NO8): Calc. [M + H]+: 360.1658, Found [M + H]+: 360.1671. Boc N O HO MeO2C O (–)-III-97b OMe DMP DCM 0 °C to rt 12 h, 80% O MeO2C O (–)-III-100b Boc N O OMe Synthesis of aldehyde (–)-III-100b: Alcohol (–)-III-95b (1.0 equiv, 0.24 mmol, 85 mg) was dissolved in dry DCM (5 mL). To this solution at 0 °C were added Dess-Martin periodinane (5.0 equiv, 1.1 mmol, 400 mg) and sodium bicarbonate (5.0 equiv, 1.1 mmol, 85 mg). The reaction mixture was allowed to warm up to room temperature and stirred 220 for 12 h. The reaction was quenched by addition of sodium thiosulfate (5 mL, sat.), sodium bicarbonate (5 mL, sat.) and water (5 mL). The mixture was extracted with DCM (4 x 10 mL). The combined organic extract was washed with brine, dried over sodium sulfate and concentrated under reduced pressure. The crude aldehyde was purified by column chromatography (silica, ethyl acetate/hexane = 20% to 50%) to yield pure aldehyde (–)- III-100b (68 mg, 0.19 mmol, 80%) as a white sticky gel. 23 = –122.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3) δ 10.08 (s, 1H), 4.96 (d, J = 5.0 Hz, 1H), 3.89 (s, 3H), 3.23 (s, 3H), 2.82 (dd, J = 8.2, 0.7 Hz, 1H), 2.49 (dd, J = 13.4, 0.7 Hz, 1H), 2.16 (ddd, J = 13.4, 8.2, 5.0 Hz, 1H), 1.43 (s, 9H), 1.37 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 196.8, 172.1, 166.7, 105.0, 84.4, 55.5, 52.8, 50.7, 34.8, 27.8, 21.5. TOF MS ES+ (C16H24NO8): Calc. [M + H]+: 358.1502, Found [M + H]+: 358.1504. O MeO2C O (–)-III-100a Boc N O OMe (–)-III-100a was synthesis according to the procedure described above for the synthesis of (–)-III-100b. 50 mg pure aldehyde was isolated (0.14 mmol, 85%) as a white gluey solid. 23 [a]D = +48.0 (c 1.0, CHCl3). 221 1H-NMR (500 MHz, CDCl3) δ 10.11 (s, 1H), 5.03 (dd, J = 5.8, 3.9 Hz, 1H), 3.87 (s, 3H), 3.32 (s, 3H), 2.99 (dd, J = 9.5, 2.6 Hz, 1H), 2.66 (ddd, J = 14.0, 5.8, 2.6 Hz, 1H), 2.20 (ddd, J = 14.0, 9.5, 3.9 Hz, 1H), 1.51 (s, 3H), 1.43 (s, 9H). 13C-NMR (125 MHz, CDCl3) δ 196.3, 172.8, 166.4, 166.2, 106.1, 85.4, 84.7, 55.8, 52.7, 51.9, 35.4, 27.7, 21.6. TOF MS ES+ (C16H23NO8): Calc. [M + H]+: 358.1502, Found [M + H]+: 358.1502. Boc N O O MeO2C O (–)-III-100b OMe ZnCl THF, –78 °C 92% H HO MeO2C O (–)-III-101b Boc N O OMe Synthesis of (–)-III-101b: Following reported procedures.13 tributyl(2-cyclohexenyl) stannane (10 equiv, 1.8 mmol, 0.72 g) was dissolved in dry THF (2 mL). To this mixture at –78 °C under argon was added n-butyllithium (10 equiv, 1.8 mmol, 1.0 mL of 1.8 M sol. in hexane) dropwise. After stirring for 1 h at this temperature, to this mixture was added ZnCl2 (10 equiv, 1.8 mmol, 1.8 mL of 1.0 M sol. in diethyl ether). The reaction mixture was stirred at –78 °C for another 30 min, after which aldehyde (–)-III-100b (1.0 equiv, 0.18 mmol, 65 mg) in dry THF (2 mL) was added dropwise. The reaction mixture was further stirred at this temperature for 5 h. Aqueous NH4Cl (10 mL, sat.) was added at –78 °C, the reaction mixture was warmed to room temperature and the organic layer was separated. The aqueous layer was extracted with ethyl acetate (3 x 10 mL). The combined organic extracts were washed with brine (10 mL), dried over sodium sulfate and concentrated 222 25 under reduced pressure. The crude product was purified by column chromatography (silica, ethyl acetate/hexane = 50%) yielding the pure product (–)-III-101b as a white foam (72 mg, 0.16 mmol, 92% yield). = –149.0 (c 1.0, CHCl3). [a]D 1H-NMR (500 MHz, CDCl3). δ 5.98 (dq, J = 10.7, 3.3, 2.6 Hz, 1H), 5.49 (dd, J = 10.7, 2.9 Hz, 1H), 4.90 (d, J = 4.8 Hz, 1H), 4.25 – 4.17 (m, 1H), 3.78 (s, 3H), 3.25 (s, 2H), 2.99 (d, J = 7.9 Hz, 1H), 2.93 (s, 1H), 2.44 (d, J = 13.1 Hz, 1H), 2.16 (ddd, J = 13.1, 7.9, 4.9 Hz, 1H), 1.98 (m, 2H), 1.84 (d, J = 6.9 Hz, 1H), 1.79 (td, J = 10.1, 9.5, 6.0 Hz, 1H), 1.75 – 1.67 (m, 1H), 1.56 (d, J = 8.8 Hz, 3H), 1.51 (s, 9H), 1.47 (s, 3H). 13C-NMR (125 MHz, CDCl3)175.47, 167.08, 150.43, 134.84, 125.17, 103.63, 89.17, 83.11, 76.02, 54.88, 51.94, 51.03, 37.84, 35.81, 30.23, 27.84, 25.07, 20.75, 20.66. TOF MS ES+ (C22H34NO8): Calc. [M + H]+: 440.2284, Found [M + H]+: 440.2286. Boc N O H O HO MeO2C OMe (–)-III-101a Synthesis of (–)-III-101a: (–)-III-101a was synthesized according to the procedure described above for the synthesis of (–)-III-101b. (–)-III-101a (51 mg, 0.12 mmol, 92% yield) was isolated as a white foam. 23 [a]D = –10.0 (c 1.0, CHCl3). 223 1H-NMR (500 MHz, CDCl3) δ 5.98 (ddt, J = 10.0, 3.9, 1.9 Hz, 1H), 5.44 (dd, J = 10.0, 2.9 Hz, 1H), 4.99 (dd, J = 6.0, 4.5 Hz, 1H), 4.25 (d, J = 6.4 Hz, 1H), 3.75 (s, 3H), 3.28 (s, 3H), 3.12 (d, J = 9.1 Hz, 1H), 2.90 (br, 1H), 2.65 (ddd, J = 14.0, 6.0, 1.4 Hz, 1H), 2.16 (ddd, J = 14.0, 9.1, 4.5 Hz, 1H), 1.98 (m, 2H), 1.88 – 1.68 (m, 4H), 1.60 (s, 3H), 1.57 – 1.53 (m, 2H), 1.49 (s, 9H). 13C-NMR (125 MHz, CDCl3) δ 176.29, 167.18, 150.16, 134.62, 125.33, 104.88, 88.02, 83.85, 75.34, 55.60, 52.29, 51.88, 37.75, 36.05, 30.26, 27.77, 25.05, 20.82, 20.16. TOF MS ES+ (C22H34NO8): Calc. [M + H]+: 440.2284, Found [M + H]+: 440.2282. H HO MeO2C O (–)-III-101b Boc N O OMe TFA/H2O/THF, 0 °C to 60 °C then NaBH4 H2O/THF 65% H Boc N O HO MeO2C OH (–)-III-102 OH Synthesis of diol (–)-III-102: Following the reported procedures,13 to a solution of (– )-III-101b (1.0 equiv, 0.13 mmol, 60 mg) in distilled THF (0.5 mL) at 0 °C was added a mixture of trifluoroacetic acid and water (1:1, v:v, 2 mL) dropwise. The reaction flask was placed in a preheated oil bath at 60 °C and stirring was continued at this temperature for 2 h. To the mixture at 0 °C was added ice (1.0 g) and toluene (3 mL), and the volatiles were removed under vacuum at room temperature. During the concentration period, toluene was added to the mixture to keep the concentration of TFA low. The resulting residue was dissolved in distilled THF (1 mL) and water (1 mL). To this mixture at 0 °C was added sodium borohydride (25 mg, 0.65 mmol, 5.0 equiv) and the mixture was stirred 224 for 1 h. The reaction was quenched with saturated solution of ammonium chloride (5 mL) and the mixture was extracted with ethyl acetate (5 x 10 mL). The combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced pressure to give the crude product as a white solid. The crude product was purified on silica column chromatography (silica, ethyl acetate/hexane = 20% then methanol/ethyl acetate = 5%) to give (–)-III-102 as a pure white solid (28 mg, 0.08 mmol, 65%). 23 = –55.0 (c 1.0, CHCl3) [a]D 1H-NMR (500 MHz, CDCl3). δ 8.37 (brs, 1H), 6.00 (m, 1H), 5.72 (m, 1H), 5.30 (brs, 1H), 4.10 – 4.03 (m, 1H), 3.80 (s, 3H), 3.78 – 3.72 (m, 2H), 3.70 – 3.65 (m, 1H), 2.83 (dd, J = 9.8, 3.3 Hz, 1H), 2.78 – 2.68 (m, 1H), 2.17 (d, J = 7.2 Hz, 1H), 1.97 (m, 3H), 1.90 (m, 2H), 1.60 – 1.70 (m, 2H), 1.52 (s, 3H). 13C-NMR (125 MHz, CDCl3) δ 180.4, 172.5, 133.8, 124.1, 81.8, 79.4, 75.3, 61.9, 52.8, 51.2, 38.8, 28.0, 26.1, 24.8, 20.7, 19.6. TOF MS ES+ (C16H26NO6): Calc. [M + H]+: 328.1760, Found [M + H]+: 328.1760. H Boc N O HO MeO2C OH (–)-III-102 OH [Me2TeAlMe]2 toluene, rt then BOPCl Pyridine, DCM 68% H H N O HO O O (–)-III-108 OH Synthesis of b-lactone (–)-III-108: To a suspension of tellurium (0.72 g, 5.60 mmol) in toluene (2.5 mL) was added Me3Al (2.0 M in toluene, 2.5 mL, 5.0 mmol). The mixture 225 was refluxed for 6 h and cooled to room temperature. The resulting [Me2AlTeMe]2 (ca 0.8 M in toluene, 2.0 mL, 8.0 mmol, 10 equiv) was added to (–)-III-102 (25.0 mg, 0.08 mmol). After stirring at room temperature for 12 h, the reaction mixture was diluted with ethyl acetate (5 mL). HCl (1.0 M, 3 mL) was added slowly to the mixture at 0 °C. The reaction mixture was further stirred at room temperature for 2 h. The acidified aqueous layer (pH = 1) was extracted with ethyl acetate/ethanol (20:1 v:v, 20 ml). The combined organic extracts were dried over sodium sulfate and concentrated under reduced pressure to yield the crude carboxylic acid as an off white solid, which was used in the following step without purification. To a solution of the carboxylic acid (20 mg) in dichloromethane (2 mL) were added pyridine (0.3 mL) and BOPCl (100 mg, 0.25 mmol), and the mixture was stirred at room temperature for 3.5 h. The reaction mixture was diluted with ethyl acetate (5 mL) and saturated NH4Cl (5 mL). The organic layer was separated and the aqueous layer was extracted with ethyl acetate (4 x 5 mL). The combined organic extracts were washed with brine (5 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified by column chromatography (silica, ethyl acetate/hexane = 20% to 50%) to give the β-lactone (–)-III-108 (12.0 mg, 0.04 mmol, 68%) as a colorless solid: 23 = –72.0 (c 0.5, MeOH) [a]D 1H-NMR (500 MHz, pyridine-d5) δ 10.60 (s, 1H), 6.46 (d, J = 10.3 Hz, 1H), 5.91 (ddd, J = 10.3, 4.9, 2.5 Hz, 1H), 5.08 (br, 2H), 4.30 (m, 2H), 4.20 (dt, J = 8.8, 6.4 Hz, 1H), 3.41 (t, J = 7.0 Hz, 1H), 2.90 (m, 1H), 2.56 (dq, J = 13.4, 7.0 Hz, 1H), 2.34 (dq, J = 13.3, 6.4 Hz, 3H), 2.19 (s, 3H), 1.99 – 1.90 (m, 2H), 1.74 (m, 2H), 1.40 (m, 1H). 226 13C-NMR (125 MHz, pyridine-d5) δ 178.66, 170.31, 129.40, 129.20, 87.49, 80.60, 71.48, 60.24, 46.76, 39.77, 29.62, 26.90, 25.78, 22.18, 20.76. TOF MS ES+ (C15H22NO5): Calc. [M + H]+: 296.1498, Found [M + H]+: 296.1497. H H N O HO O O (–)-III-108 OH PPh3Cl2 Pyr:ACN 65% H H N O HO O O (–)-III-1 Cl Synthesis of (–)-Salinosporamide A (–)-III-1: β-lactone (–)-III-108 (8.0 mg, 0.03 mmol) was dissolved in acetonitrile (0.5 mL) and anhydrous pyridine (0.5 mL) at room temperature under argon. To this solution was added Ph3PCl2 (5.00 equiv, 0.16 mmol, 55.0 mg), and stirring was continued for 3.5 h. The reaction was quenched by addition of water (5 mL) followed by addition of ethyl acetate (5 mL). The aqueous layer was extracted with ethyl acetate (4 x 5 mL), combined extracts were washed with brine (5 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified by column chromatography (silica, ethyl acetate = 20% to 40%) to yield (–)- Salinosporamide A as a white solid (6.0 mg, 0.02 mmol, 64%). 23 = –69.0 (c 0.4, MeOH) [a]D 1H-NMR (500 MHz, pyridine-d5) δ 10.67 (s, 1H), 6.45 (d, J = 10.3 Hz, 1H), 5.91 (d, J = 10.3 Hz, 1H), 5.02 (br, 1H), 4.28 (t, J = 9.1 Hz, 1H), 4.21 – 4.11 (m, 1H), 4.05 (m, 1H), 3.21 (t, J = 7.2 Hz, 1H), 2.94 – 2.82 (m, 1H), 2.56 – 2.47 (m, 1H), 2.41 – 2.30 (m, 2H), 227 2.10 (d, J = 2.3 Hz, 3H), 1.99 – 1.90 (m, 2H), 1.72 (t, J = 11.0 Hz, 2H), 1.42 – 1.36 (m, 1H). 13C-NMR (125 MHz, pyridine-d5) δ 177.34, 169.84, 129.48, 129.09, 86.72, 80.76, 71.39, 46.57, 43.70, 39.73, 29.42, 26.87, 25.77, 22.14, 20.41. TOF MS ES+ (C15H21ClNO4): Calc. [M + H]+: 314.1159, Found [M + H]+: 314.1165. 228 Compound BB817C Formula C19H31NO5Si Dcalc./ g cm-3 µ/mm-1 1.215 0.140 Formula Weight 381.54 Color Shape colorless plate Size/mm3 0.38×0.15×0.06 T/K Crystal System Space Group a/Å b/Å c/Å a/° b/° g/° V/Å3 Z 173(2) triclinic P-1 7.1927(8) 7.5787(8) 21.102(2) 83.7720(10) 82.5490(10) 66.3970(10) 1043.13(19) 2 III-7-3 Crystal data III-7-3-1 Crystal data for III-91b (BB817C) A suitable Experimental. Single colorless plate-shaped crystals of III-91b (BB817C) were used as received. crystal (0.38×0.15×0.06) mm3 was selected and mounted on a nylon loop with paratone oil on a Bruker APEX-II CCD diffractometer. The crystal was kept at T = 173(2) K during data collection. Using Olex2,70 the structure was solved with the ShelXT71 structure solution program, using the Intrinsic Phasing solution method. The model 229 was refined with version of XL71 using Least Squares minimisation. Crystal Data. C19H31NO5Si, Mr = 381.54, triclinic, P-1 (No. 2), a = 7.1927(8) Å, b = 7.5787(8) Å, c = 21.102(2) Å, a = 83.7720(10)°, b = 82.5490(10)°, g = 66.3970(10)°, V = 1043.13(19) Å3, T = 173(2) K, Z = 2, Z' = 1, µ(MoKa) = 0.140, 15883 reflections measured, 4145 unique (Rint = 0.0465) which were used in all calculations. The final wR2 was 0.1580 (all data) and R1 was 0.0547 (I > 2(I)). Z' Wavelength/Å Radiation type Qmin/° Qmax/° Measured Refl. Independent Refl. Reflections Used Rint Parameters Restraints Largest Peak Deepest Hole GooF wR2 (all data) wR2 R1 (all data) R1 1 0.710730 MoKa 1.950 26.142 15883 4145 2893 0.0465 243 0 0.743 -0.271 1.063 0.1580 0.1404 0.0796 0.0547 230 Structure Quality Indicators Reflection: Refinement: A colorless plate-shaped crystal with dimensions 0.38×0.15×0.06 mm3 was mounted on a nylon loop with paratone oil. Data were collected using a Bruker APEX-II CCD diffractometer equipped with an Oxford Cryosystems low-temperature device, operating at T = 173(2) K. Data were measured using w of -0.50° per frame for 299.34 s using MoKa radiation (sealed tube, 50 kV, 40 mA). The total number of runs and images was based on the strategy calculation from the program COSMO (BRUKER, V1.61, 2009). The actually achieved resolution was Q = 26.142. Cell parameters were retrieved using the SAINT72 software and refined using SAINT72 on 5215 reflections, 33% of the observed reflections. Data reduction was performed using the SAINT72 software which corrects for Lorentz polarisation. The final completeness is 99.90 out to 26.142 in Q. A multi-scan absorption correction was performed using SADABS-2014/5 (Bruker,2014/5) was used for absorption correction. wR2 73 was 0.0606 before and 0.0512 after correction. The Ratio of minimum to maximum transmission is 0.9066. The l/2 correction factor is 0.00150. The absorption coefficient µ of this material is 0.140 mm-1 at this wavelength (l = 0.71073Å) and the minimum and maximum transmissions are 0.6757 and 0.7453. SADABS-2014/5 (Bruker,2014/5) was used for 231 absorption correction. wR2 minimum to maximum transmission is 0.9066. The l/2 correction factor is 0.00150. 73 was 0.0606 before and 0.0512 after correction. The Ratio of The structure was solved in the space group P-1 (# 2) by Intrinsic Phasing using the ShelXT71 structure solution program. The structure was refined by Least Squares using version 2014/6 of XL71 incorporated in Olex2.70 All non-hydrogen atoms were refined anisotropically. Hydrogen atom positions were calculated geometrically and refined using the riding model. There is a single molecule in the asymmetric unit, which is represented by the reported sum formula. In other words: Z is 2 and Z' is 1. Figure III-32. X-ray structure of III-91b (view 1). 232 Figure III-33. X-ray structure of III-91b (view 2). 233 Table III-5: Fractional Atomic Coordinates (×104) and Equivalent Isotropic Displacement Parameters (Å2×103) for III-91b. Ueq is defined as 1/3 of the trace of Atom the orthogonalised Uij. x y z Ueq Si1 O1 O2 O3 O4 O5 N1 C1 C2 C3 C4 C5 C6 C7 C8 C9 C10 C11 5874.7(11) 9017(2) 12240(2) 11576(2) 10562(3) 6037(2) 8731(3) 11146(3) 11416(3) 9658(3) 7955(3) 6991(3) 6290(3) 7428(4) 9118(4) 10180(3) 12161(4) 6461(3) 7679.7(12) 4166(2) 4302(2) 5871(2) 9004(2) 6652(2) 7323(2) 3114(3) 2408(3) 4025(3) 4652(3) 6884(3) 8085(3) 9081(3) 8541(3) 5781(3) 5057(4) 3687(3) 234 3600.4(3) 1153.2(7) 1025.7(7) 2472.6(8) 1330.2(9) 2936.2(7) 1907.3(9) 1188.5(11) 1882.7(11) 2229.3(11) 1783(1) 1800.4(10) 1199.8(11) 990.3(11) 1406.3(12) 2236.8(10) 378.5(12) 1921.4(12) 45.6(2) 30.7(4) 34.8(4) 42.2(4) 46.7(5) 37.8(4) 29.0(4) 31.6(5) 33.6(5) 30.1(5) 29.2(5) 28.2(5) 33.0(5) 35.5(6) 34.4(5) 31.1(5) 43.5(6) 36.3(6) Table III-5 (cont’d) Atom C12 C13 C14 C15 C16 C17 C18 C19 x 7228(4) 5312(3) 6444(7) 7853(5) 3272(4) 1671(5) 3191(6) 2843(7) y 10534(4) 7603(3) 9874(6) 5821(5) 8301(6) 9814(8) 9092(8) 6460(8) z 440.4(13) 2348.5(10) 3406.9(18) 4060.5(15) 4033.4(14) 3628.5(19) 4680.3(17) 4149.4(19) Ueq 50.7(7) 32.9(5) 81.9(11) 70(1) 68.6(10) 107.9(17) 106.1(17) 110.5(17) displacement factor exponent takes the form: -2p2[h2a*2 × U11+ ... +2hka* × b* × U12] Table III-6: Anisotropic Displacement Parameters (×104) III-91b. The anisotropic Atom Si1 O1 O2 O3 O4 O5 N1 C1 U11 39.2(4) 27.4(8) 32.2(8) 32.5(9) 47.8(10) 37.0(9) 27.5(9) 26.5(11) U22 62.8(5) 32.2(8) 39.4(9) 51.5(11) 36.7(9) 39.6(9) 28.1(9) 27.4(11) U23 -9.3(3) -6.5(6) -2.6(7) -11.6(8) -7.6(8) -2.1(7) -6.7(8) -6.4(9) U13 0.7(3) -0.5(6) -0.1(7) -7.8(7) 6.6(9) 1.2(7) -0.1(8) -0.4(9) U12 -20.2(4) -9.8(6) -17.2(7) -16.4(8) -27.3(8) -9.5(7) -11.6(8) -7.9(9) U33 35.3(4) 31.0(8) 35.1(9) 45.7(10) 63.5(12) 30.4(9) 32.6(10) 38.6(13) 235 Table III-6 (cont’d) U11 Atom C2 C3 C4 C5 U22 U33 27.6(11) 32.3(12) 28.4(11) 28.0(11) 42.3(14) 29.0(12) 29.9(12) 32.0(12) U23 0.3(10) -0.4(9) -3.2(9) -2.3(9) U13 -3.5(10) -0.6(9) 0.1(9) -3.3(9) U12 -7.5(9) -11.1(9) -10.2(9) -11.3(9) 27.2(11) 27.4(11) 28.1(11) 25.5(11) 31.7(12) 31.5(12) 31.6(12) -4.6(10) -4.7(9) -6.9(10) -3.3(10) -10.5(10) -8.8(10) 0.8(11) -5.6(10) 5.2(12) -3.1(10) -23(2) 1.4(17) -28.9(18) -39(3) -65(3) -10(3) 3.9(10) 6.3(10) 2.0(9) 2.9(11) -0.1(10) 2.4(13) -1.3(10) -6(2) -11.8(14) 6.0(13) 2.4(17) 19.5(18) 15(2) -7.9(10) -11.5(10) -11.9(10) -19.7(12) -15.4(10) -16.2(13) -8.8(10) -67(2) -22.4(18) -28.1(18) 4(2) -46(3) -114(4) 40.0(13) 36.4(13) 26.2(11) 42.3(14) 32.9(12) 59.6(17) 30.8(12) 113(3) 53.0(18) 39.0(16) 42.7(19) 65(2) 95(3) 25.5(11) 23.9(11) 37.8(13) 47.5(15) 33.5(12) 40.1(14) 31.9(12) 93(3) 103(3) 125(3) 167(4) 197(5) 215(6) 34.5(13) 41.6(14) 28.9(12) 39.8(14) 44.6(14) 45.2(16) 32.5(12) 68(2) 46.4(18) 40.6(17) 76(3) 58(2) 66(2) 236 C6 C7 C8 C9 C10 C11 C12 C13 C14 C15 C16 C17 C18 C19 Atom Si1 Si1 Si1 Si1 O1 O1 O2 O2 O3 O4 O5 N1 N1 N1 C1 C2 C3 C3 C4 C4 Table III-7: Bond Lengths in Å for III-91b. Atom O5 C14 C15 C16 C1 C4 C1 C10 C9 C8 C13 C5 C8 C9 C2 C3 C4 C9 C5 C11 Length/Å 1.6449(17) 1.859(4) 1.846(3) 1.872(3) 1.423(3) 1.450(3) 1.406(3) 1.422(3) 1.205(3) 1.210(3) 1.415(3) 1.466(3) 1.401(3) 1.404(3) 1.513(3) 1.533(3) 1.533(3) 1.523(3) 1.553(3) 1.509(3) 237 Table III-7 (cont’d) Atom C5 C5 C6 C7 C7 C16 C16 C16 Atom C6 C13 C7 C8 C12 C17 C18 C19 Length/Å 1.495(3) 1.530(3) 1.328(3) 1.488(3) 1.489(3) 1.535(5) 1.535(4) 1.534(6) 238 Table III-8: Bond Angles in ° for III-91b. Atom Si1 Si1 Si1 Si1 Si1 Si1 O1 O2 O5 N1 N1 N1 C1 C1 C1 C2 C3 C3 C3 C4 Atom O5 O5 O5 C14 C15 C15 C1 C1 C13 C8 C8 C9 O1 O2 O2 C1 C4 C9 C9 O1 Atom C14 C15 C16 C16 C14 C16 C4 C10 Si1 C5 C9 C5 C2 O1 C2 C3 C2 C2 C4 C3 239 Angle/° 109.58(13) 103.70(13) 110.02(12) 110.77(18) 110.93(18) 111.60(15) 111.33(16) 113.84(18) 126.46(14) 111.01(18) 126.95(18) 111.81(17) 106.30(17) 111.67(17) 107.13(18) 101.61(17) 103.59(17) 110.52(18) 105.08(17) 104.42(16) Table III-8 (cont’d) Atom O1 O1 C3 C11 C11 N1 N1 N1 C6 C6 C13 C7 C6 C6 C8 O4 O4 N1 O3 O3 Atom C4 C4 C4 C4 C4 C5 C5 C5 C5 C5 C5 C6 C7 C7 C7 C8 C8 C8 C9 C9 Atom C5 C11 C5 C3 C5 C4 C6 C13 C4 C13 C4 C5 C8 C12 C12 N1 C7 C7 N1 C3 240 Angle/° 108.23(17) 108.55(18) 103.74(17) 116.22(19) 114.95(18) 102.37(16) 102.37(17) 109.52(17) 119.14(18) 109.09(17) 113.21(18) 111.5(2) 109.1(2) 130.3(2) 120.6(2) 126.2(2) 128.1(2) 105.71(19) 125.0(2) 127.6(2) Table III-8 (cont’d) Atom O5 C17 C18 C18 C19 C19 C19 Atom C13 C16 C16 C16 C16 C16 C16 Atom C5 Si1 Si1 C17 Si1 C17 C18 Angle/° 110.00(17) 109.9(2) 109.7(2) 109.8(3) 108.5(3) 109.7(3) 109.2(3) Table III-9: Hydrogen Fractional Atomic Coordinates (×104) and Equivalent Isotropic Displacement Parameters (Å2×103) for III-91b. Ueq is defined as 1/3 of the trace of the Atom H1 H2A H2B H3 H6 H10A H10B H10C orthogonalised Uij. x y z Ueq 11670 12753 11280 9240 5171 12523 13126 10779 2000 2283 1154 3581 8129 4004 5688 5999 241 908 2001 1976 2668 994 94 279 316 38 40 40 36 40 65 65 65 Table III-9 (cont’d) Atom H11A H11B H11C H12A H12B H12C H13A H13B H14A H14B H14C H15A H15B H15C H17A H17B H17C H18A H18B H18C x 5450 5772 7193 8335 7303 5914 4877 4116 7693 6629 5308 7581 7834 9193 1649 2017 326 4295 1877 3348 z 1608 2353 1893 105 585 270 2370 2269 3115 3802 3202 4127 4476 3822 3232 3525 3870 4921 4925 4608 y 4184 3958 2290 9987 11687 10883 9013 7350 9564 10330 10885 4644 6296 5539 9264 10946 10200 8171 9285 10326 242 Ueq 54 54 54 76 76 76 39 39 123 123 123 105 105 105 162 162 162 159 159 159 Table III-9 (cont’d) Atom H19B H19C y 6756 5489 z 4384 4401 Ueq 166 166 x 1495 3883 243 III-7-3-2 Crystal data for (–)-III-25 (BB817D) Compound BB817D Formula C20H31NO4SSi Dcalc./ g cm-3 µ/mm-1 1.194 0.218 Formula Weight 409.61 Color Shape colorless plate Size/mm3 0.46×0.32×0.05 T/K 173(2) Crystal System orthorhombic Flack Parameter Hooft Parameter Space Group a/Å b/Å c/Å a/° -0.02(2) -0.02(2) P212121 10.307(2) 12.283(3) 17.991(4) 90 Experimental. Single colorless plate-shaped crystals of (–)-III-25 (BB817D) were used as received. crystal (0.46×0.32×0.05) mm3 was selected and suitable A mounted on a nylon loop with paratone oil on a Bruker APEX-II CCD diffractometer. The crystal was kept at T = 173(2) K during data collection. Using Olex2,70 the structure was solved with the XT structure solution program, using the Intrinsic Phasing solution method. The model was refined with version of XL71 using Least Squares 244 minimisation. Crystal Data. C20H31NO4SSi, Mr = 409.61, orthorhombic, P212121 (No. 19), a = 10.307(2) Å, b = 12.283(3) Å, c = 17.991(4) Å, a = b = g = 90°, V = 2277.7(8) Å3, T = 173(2) K, Z = 4, Z' = 1, µ(MoKa) = 0.218, 43882 reflections measured, 5640 unique (Rint = 0.0420) which were used in all calculations. The final wR2 was 0.0993 (all data) and R1 was 0.0372 (I > 2(I)). 245 b/° g/° V/Å3 Z Z' 90 90 2277.7(8) 4 1 Wavelength/Å 0.710730 Radiation type Qµin/° Qµax/° Measured Refl. Independent Refl. Reflections Used Rint Parameters Restraints Largest Peak Deepest Hole GooF wR2 (all data) MoKa 2.007 28.441 43882 5640 5095 0.0420 255 0 0.310 -0.217 1.050 0.0993 Structure Quality Indicators Reflection: Refinemen: A colorless plate-shaped crystal with dimensions 0.46×0.32×0.05 mm3 was mounted on a nylon loop with paratone oil. Data were collected using a Bruker APEX-II CCD diffractometer equipped with an Oxford Cryosystems low-temperature device, operating at T = 173(2) K. Data were measured using w of -0.50° per frame for 200.84 s using MoKa radiation (sealed tube, 50 kV, 40 mA). The total number of runs and images was based on the strategy calculation from the program COSMO (BRUKER, V1.61, 2009). The actually achieved resolution was Q = 28.441. Cell parameters were retrieved using the SAINT72 software and refined using SAINT72 on 9967 reflections, 23 % of the observed reflections. Data reduction was performed using the SAINT72 software which corrects for Lorentz polarisation. The final completeness is 100.00 out to 28.441 in Q. A multi-scan absorption correction was performed using SADABS-2014/5 (Bruker,2014/5) was used for absorption correction. wR2 73 was 0.0660 before and 0.0554 after correction. The Ratio of minimum to maximum transmission is 0.9136. The l/2 correction factor is 0.00150. The absorption coefficient m of this material is 0.218 mm-1 at this wavelength (l = 0.71073Å) and the minimum and maximum transmissions are 0.6813 and 0.7457. SADABS-2014/5 246 (Bruker,2014/5) was used for absorption correction. wR2 73 was 0.0660 before and 0.0554 after correction. The Ratio of minimum to maximum transmission is 0.9136. The l/2 correction factor is 0.00150. The structure was solved in the space group P212121 (# 19) by Intrinsic Phasing using the XT (Sheldrick, 2015) structure solution program. The structure was refined by Least Squares using version 2014/6 of XL71 incorporated in Olex2.70 All non-hydrogen atoms were refined anisotropically. Hydrogen atom positions were calculated geometrically and refined using the riding model, except for the Hydrogen atom on the oxygen atom which was found by difference Fourier methods and refined isotropically. There is a single molecule in the asymmetric unit, which is represented by the reported sum formula. In other words: Z is 4 and Z' is 1. The Flack parameter was refined to -0.02(2). Determination of absolute structure using Bayesian statistics on Bijvoet differences using the Olex2 results in -0.02(2). Note: The Flack parameter is used to determine chirality of the crystal studied, the value should be near 0, a value of 1 means that the stereochemistry is wrong and the model should be inverted. A value of 0.5 means that the crystal consists of a racemic mixture of the two enantiomers. 247 Figure III-34. X-ray structure of (–)-III-25 (view 1). Figure III-35: The Model has Chirality at C1 (Chiral SPGR) S Verify; The Model has Chirality at C2 (Chiral SPGR) S Verify; The Model has Chirality at C3 (Chiral SPGR) S Verify. 248 Figure III-36: The following hydrogen bonding interactions with a maximum D-D distance of 2.9 Å and a minimum angle of 120 ° are present in (–)-III-25: O1–O3: 2.865 Å. Table III-10: Fractional Atomic Coordinates (×104) and Equivalent Isotropic Displacement Parameters (Å2×103) for (–)-III-25. Ueq is defined as 1/3 of the trace of the orthogonalised Uij. x y 828.6(5) 5881.5(7) 429.0(18) 4872.6(17) -385.9(17) 1622.2(18) 1616.0(18) 6886.9(5) 5337.9(5) 4081.6(17) 5517.8(16) 6311.0(15) 6996.9(16) 6305.4(15) 249 z 4781.5(3) 2770.5(4) 4822.9(13) 3472.8(11) 4848.5(11) 5431.6(10) 4079.7(11) Ueq 29.27(14) 30.61(16) 44.0(5) 41.2(4) 41.4(4) 38.5(4) 27.4(4) Atom S1 Si1 O1 O2 O3 O4 N1 Table III-10 (cont’d) Atom C1 C2 C3 C4 C5 C6 C7 C8 C9 C10 C11 C12 C13 C14 C15 C16 C17 C18 C19 C20 x 2001(2) 3003(2) 1748(2) 2615(3) 3375(3) 1730(3) 3886(2) 6873(3) 4948(4) 6914(2) 7723(4) 7836(3) 6056(3) 521(2) -561(2) -817(3) -11(3) 1065(3) 1340(3) -280(4) z 4083.4(13) 4150.1(13) 4789.7(15) 4781.9(15) 4767.9(18) 3359.2(14) 3517.2(15) 2629(2) 1911.7(18) 3075.2(14) 3752.3(19) 2447.9(18) 3284(2) 4413.2(12) 3964.5(14) 3693.2(15) 3855.4(15) 4305.9(16) 4588.7(16) 3534(2) y 5125.6(18) 5993.0(19) 4458(2) 3501(2) 2779(2) 4554(2) 6318(2) 6589(2) 5015(3) 4152(2) 4472(3) 3807(3) 3185(2) 8191.0(19) 8341(2) 9376(2) 10251(2) 10074(2) 9046(2) 11368(3) 250 Ueq 27.8(5) 28.3(5) 32.6(5) 37.8(5) 47.0(7) 38.3(6) 35.2(5) 55.2(8) 56.2(8) 33.1(5) 58.3(9) 53.8(8) 53.9(8) 29.4(5) 37.1(6) 43.3(6) 44.2(7) 47.4(7) 39.7(6) 62.0(9) Table III-11: Anisotropic Displacement Parameters (×104) (–)-III-25. The anisotropic displacement factor exponent takes the form: -2p2[h2a*2 × U11+ ... +2hka* × b* × U12] U12 Atom 3.6(2) S1 Si1 1.5(3) -7.0(8) O1 11.4(8) O2 O3 -0.5(8) 8.9(8) O4 N1 3.5(7) 2.9(9) C1 C2 2.5(9) -1.5(9) C3 -2.6(11) C4 C5 7.6(12) 2.2(11) C6 C7 3.8(10) U23 -1.1(2) -0.5(2) 9.2(10) 5.9(9) 2.3(9) -2.6(7) -1.1(8) 0.1(9) -0.3(9) 4.5(10) 9.8(11) 11.9(12) -5.3(10) 3.9(10) U11 25.2(3) 25.7(3) 31.3(9) 33.2(9) 29.4(8) 40.7(9) 24.6(9) 26.1(10) 23.9(10) 27.0(11) 37.1(12) 46.8(15) 42.7(14) 33.5(12) U22 31.9(3) 33.6(3) 36.6(10) 45.3(11) 42.2(10) 43.3(10) 26.0(9) 26.4(11) 27.3(11) 32.2(12) 34.5(12) 39.4(14) 34.4(13) 29.4(12) U33 30.8(3) 32.6(3) 64.1(13) 45.2(10) 52.7(11) 31.4(8) 31.5(9) 31.1(11) 33.7(11) 38.5(12) 41.6(13) 54.9(16) 38.0(13) 42.8(13) U13 3.4(2) 3.0(3) 4.3(9) 13.2(8) 8.7(8) -1.6(7) 2.6(8) -1.9(9) 1.8(9) 0.3(10) -3.5(11) 1.4(14) -5.8(11) 10.8(10) C8 C9 C10 C11 C12 39.4(15) 58.7(19) 29.3(11) 64(2) 54.7(18) 40.2(16) 64(2) 33.0(12) 55.6(19) 53.3(18) 86(2) 46.4(16) 36.8(12) 55.5(19) 53.3(17) 251 13.1(15) -4.5(15) -1(1) -5.2(15) 6.8(14) 7.9(16) -16.0(14) 5.1(10) -21.7(16) 20.5(15) -2.0(13) 11.4(16) 2.9(10) 12.4(16) 23.5(15) Table III-11 (cont’d) U11 Atom C13 46.3(16) 37.7(15) U22 U33 78(2) U23 7.0(14) U13 10.0(15) U12 1.2(13) 6.8(9) 3.6(10) 16.1(13) 18.6(13) 2.3(13) 3.7(11) 25.6(17) C14 C15 C16 C17 C18 C19 C20 27.8(11) 30.0(11) 30.3(11) -3.5(9) 3.6(8) 28.3(12) 34.0(13) 53.8(17) 59.6(18) 41.8(14) 75(2) 48.1(15) 59.1(17) 42.8(15) 33.2(13) 34.1(13) 52.0(18) 34.8(12) 36.9(13) 36.0(13) 49.4(15) 43.2(14) 58.9(19) 1.3(11) 10.8(12) 2.3(12) -6.1(11) -7.4(11) 15.3(16) 0.9(9) 2.8(11) 11.1(12) -0.9(14) -9.0(11) 17.9(18) Table III-12: Bond Lengths in Å for (–)-III-25 Atom S1 S1 S1 S1 Si1 Si1 Si1 Si1 Atom O3 O4 N1 C14 O2 C8 C9 C10 252 Length/Å 1.4428(19) 1.4336(19) 1.6622(19) 1.762(2) 1.6512(18) 1.864(3) 1.863(3) 1.886(3) Table III-12 (cont’d) Atom O1 O2 N1 N1 C1 C1 C1 C2 C3 C4 C10 C10 C10 C14 C14 C15 C16 C17 C17 C18 Atom C3 C7 C1 C2 C2 C3 C6 C7 C4 C5 C11 C12 C13 C15 C19 C16 C17 C18 C20 C19 253 Length/Å 1.437(3) 1.417(3) 1.503(3) 1.486(3) 1.489(3) 1.535(3) 1.506(3) 1.511(3) 1.477(3) 1.183(4) 1.528(4) 1.535(4) 1.528(4) 1.388(3) 1.384(4) 1.387(4) 1.390(4) 1.391(4) 1.514(4) 1.390(4) Table III-13: Bond angles in ° for (–)-III-25. Atom S1 S1 S1 S1 S1 S1 Si1 Si1 Si1 Si1 Si1 Si1 O2 N1 N1 N1 C1 C1 C1 C1 Atom N1 C14 O3 N1 C14 C14 C8 C9 C10 C10 C8 C10 Si1 S1 S1 C1 C3 C6 N1 C3 Angle/° 106.05(11) 108.71(11) 118.23(12) 112.43(10) 108.89(12) 101.13(10) 109.85(14) 109.75(14) 103.68(10) 111.55(13) 110.21(17) 111.62(14) 126.02(16) 122.69(15) 121.07(16) 59.75(14) 118.29(19) 113.37(19) 59.57(14) 115.71(19) Atom O3 O3 O4 O4 O4 N1 O2 O2 O2 C8 C9 C9 C7 C1 C2 C2 N1 N1 C2 C2 254 Table III-13 (cont’d) Atom C2 C6 N1 N1 C1 O1 O1 C4 C5 O2 C11 C11 C11 C12 C13 C13 C15 C19 C19 C16 Atom C1 C1 C2 C2 C2 C3 C3 C3 C4 C7 C10 C10 C10 C10 C10 C10 C14 C14 C14 C15 255 Atom C6 C3 C1 C7 C7 C1 C4 C1 C3 C2 Si1 C12 C13 Si1 Si1 C12 S1 S1 C15 C14 Angle/° 122.1(2) 115.8(2) 60.68(14) 116.53(19) 123.1(2) 111.6(2) 108.4(2) 108.3(2) 175.7(3) 106.93(19) 109.93(19) 108.6(2) 108.6(3) 110.44(18) 110.21(18) 108.9(2) 118.95(19) 119.61(18) 121.4(2) 118.6(3) Table III-13 (cont’d) Atom C16 C16 C18 C19 C14 Atom C17 C17 C17 C18 C19 Atom C18 C20 C20 C17 C18 Angle/° 118.5(3) 120.7(3) 120.8(3) 121.2(3) 118.8(3) Table III-14: Hydrogen fractional atomic coordinates (×104) and equivalent isotropic displacement parameters (Å2×103) for (–)-III-25. Ueq is defined as 1/3 of the trace of the x 20(30) 3382 1939 3985 831 2320 1863 3393 4270 7671 orthogonalised Uij. y z 4610(30) 4850(20) Ueq 51(10) 4658 5237 4757 3355 3305 2946 3045 3613 2361 34 39 56 58 58 58 42 42 83 6084 4914 2200 4295 3933 5062 6350 7044 6402 256 Atom H1 H2 H3 H5 H6A H6B H6C H7A H7B H8A Table III-14 (cont’d) Atom H8B H8C H9A H9B H9C H11A H11B H11C H12A H12B H12C H13A H13B H13C H15 H16 H18 H19 H20A H20B x 7096 6375 4432 5551 4369 8274 8269 7145 7330 8346 8421 5508 6604 5506 -1114 -1560 1622 2077 -6 203 z 3113 2339 1992 1498 1793 3627 3902 4163 2006 2609 2328 3709 3418 2860 3846 3390 4423 4897 3012 3815 y 6905 7120 4356 4897 5624 5096 3857 4669 3613 3176 4411 3382 2563 2989 7747 9489 10665 8933 11385 11918 257 Ueq 83 83 84 84 84 87 87 87 81 81 81 81 81 81 45 52 57 48 93 93 Table III-14 (cont’d) Atom H20C Atom -1211 Atom 11523 Atom 3565 Atom 93 Table III-15: Hydrogen bond information for (–)-III-25. 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Mol Cell Endocrinol 2008, 287 (1-2), 65-71. 266 Chapter IV: Diastereoselective halonium initiated spiroketalization; towards the total synthesis of Obtusin 267 IV-1 Introduction; structural features of spiroketal moiety A large number of structurally diverse bioactive natural products found in insects, plants, bacteria and marine sources contain a spiroketal moiety. Spiroketals are often the source of bioactivity for molecules bearing this functionality.2 Furthermore, the advantageous pharmacophoric properties of this functional group for drug discovery have led to an interest in the study and synthesis of spiroketals.2-10 H H OH Me Me H O O O O (R)-Olean IV-1R active on male olive fruitfly (S)-Olean IV-1S active on female olive fruitfly Me OH O OO H OH HO O O H Me H Me O O Okadaic acid IV-2 O OH OH Me OH Me Me O Me O O MeH NH OH OH Me Me H O H H O Me HO HO H H Me O HO HO O Me O O Me Spirofungin B IV-3 Me H O HH O OH H N Me O N H O Me Me Me OH Bistramide A IV-4 Integramycin IV-5 Figure IV-1. Spiroketal moiety embedded in the structure of natural products. Most spiroketal frameworks found in natural products belong to the 6,6-, 5,6- or 5,5- spiroketal structural categories (Figure IV-1). In fact, the simplest 6,6-spiroketal, Olean IV-1, is the female-produced sex pheromone of the olive fruit fly.10 Okadaic acid IV-2, 268 Spirofungin IV-3, Bistramide A IV-4 and Integramycin IV-5 are just a few examples of natural products that contain spiroketal moieties in their highly functionalized skeletons (Figure IV-1). From the structural standpoint, spiroketals are members of the acetal containing molecules in which the carbon bearing the two carbon-oxygen single bonds (acetal carbon) is in the junction of two rings. O 1 O antiperiplanar alignment of axial C–O bond with the oxygen lone pair n–σ∗ O σ∗C–O O O O equatorial C–O bond O O σ∗C–O n–σ∗ interaction not possible due to orbital misalignment Figure IV-2. Anomeric effect, principle and structural requirements. The structural feature of spiroketals leads to its special reactivity as the result of the anomeric effects. The term “anomeric effect” refers to the tendency of an electronegative group at the anomeric center (C1) of a pyranose ring to occupy an axial rather than an equatorial position, despite unfavorable steric repulsions (Figure IV-2). As illustrated in Figure IV-2, anomeric stabilization finds its origin in the interaction between one of the lone pairs on the oxygen atom and the s* orbital of the C–O bond. The antiperiplanar alignment of the interacting orbitals can take place when a group is the axial position. The extent of anomeric stabilization is estimated to be in the range of 1.4 – 2.4 kcal/mol. Nonetheless the steric parameter or other molecular forces can obscure the anomeric 269 stabilization. In a 6,6-spiroketal framework four different arrangements can be identified with regards to the number of anomeric stabilization. As such, one structure with two anomeric interactions, two forms with one anomeric relationship and one with no anomeric alignments are possible (Figure IV-3). The structure with two anomeric relationships is the thermodynamic spiroketal if the anomeric stabilizations are not compromised by other steric parameters.3 [6,5]-spiroketal systems O O anomeric alignment O O nonanomeric alignment [6,6]-spiroketal systems doubly anomeric isomer nonanomeric isomer doubly nonanomeric isomer O O O O O O O O two anomeric relationships one anomeric relationship relationship one nonanomeric two nonanomeric relationships Figure IV-3. The anomeric arrangements in 6,5- and 6,6-spiroketal frameworks. Not surprisingly, the biochemical applications and structural architecture of spiroketals have attracted vast synthetic efforts over the past few decades. To date, several elegant strategies towards stereoselective construction of these carbocyclic cores 270 have been reported. 2-4 However, the synthesis of halogenated spiroketal still remains ill explored.11-15 Encouraged by our endeavors in halofunctionalization of olefins, we set out to develop an efficient and robust procedure to access halogenated spiroketals in a diastereoselective fashion. IV-2 Chemical synthesis of spiroketals The anomeric carbon of the spiroketal moiety is at the ketone oxidation level; hence, the most general approach to access these motifs involves acid catalyzed acetal formation by a pendant diol onto the incipient ketone in IV-6 (Figure IV-4a).2 In this acetalization reaction, the most stable spiroketal product, which is usually the double anomeric spiroketal, can be readily obtained. Since most of the naturally occurring compounds contain a double anomeric spiroketal moiety, the aforementioned acid catalyzed dehydration approach has been extensively utilized to access numerous natural products. Furthermore, the selectivity of the acid catalyzed spiroketalization can be changed by the stereo-defined functionalities along the acyclic chain of the dihydroxy ketone.3 As illustrated in Figure IV-4b, Schreiber has demonstrated that beta diketone IV- 7 with three epimerizable stereocenters can lead to the formation of spiroketal IV-8 with complete diastereoselectivity.16 In this reaction the stereodefined centers (highlighted in blue in IV-7), determine the course of the reaction. The methyl on the alpha position to carbonyl can be epimerized to give access to a single diastereomer IV-8. Another approach that allows control over the stereoselectivity in the cyclization event, involves protonation of a dihydropyranyl moiety IV-9, leading to the oxocarbenium 271 intermediate (IV-10) followed by interception of this intermediate by a tethered alcohol. In this strategy, the well-defined behaviors of oxonium embedded in a six-membered ring (axial attack) guides the trajectory of the incoming alcohol nucleophile to furnish spiroketal formation (Figure IV-4c).2-3 O O IV-1 O O Me Me O Me O IV-8 O Me a. general spiroketalization reaction of keto-diols O HO OH acid catalyzed IV-6 b. Schreiber's dynamic diastereoselective spiroketalization OH O O TBSO Me Me Me Me IV-7 OH CSA CCl4/MeOH HO c. spiroketalization via protonation of DHP core O OH IV-9 via: X* O Bronsted acid (HX) HO IV-10 OH O axial approach d. our approach, halenium initiated spiroketalization O O IV-1 * RBr HO R O Br IV-11 Halenium initiated spiroketalization HO via: O R Br * * IV-12 O O * * IV-13 * denotes the newly formed stereocenters Figure IV-4. Different approaches to access the spiroketal moiety. We envisioned arriving at a similar oxonium intermediate IV-12 via activation of an olefin with a halenium, and using a ketone as the nucleophile in IV-11 (Figure IV-4d). The 272 well-defined behaviors of the oxonium embedded in IV-12 would then lead to the diastereoselective formation of spiroketal IV-13. The latter oxacrbinium intermediate embedded in a six-membered ring has been utilized by a few researchers to successfully develop enantioselective syntheses of spiroketals. In this respect, List and coworkers have reported the first direct synthesis of spiroketals in a catalytic enantioselective manner via protonation of dihydropyranyl IV-9 (Figure IV-5).10 They employed the Brønsted acid catalyst IV-14, which contains a highly constrained binding pocket to give rise to the desired enantioselectivity during the spiroketalization event. The latter methodology gives access to various spiroketal frameworks with high enantioselectivity. HX* : O OH IV-9 Ar OH O P O O N P Ar Et Ar = Ar O O Ar Et Chiral Bronsted acid (HX*) HO via: X* O IV-10 O O IV-1 acidic site Electrophile H O O P O Nucleophile H basic site O N P O O general features of the caralyst that contains acidic and basic sites to impose a better selectivity on the spiroketalization reaction. Et Figure IV-5. List’s approach to access the spiroketal moiety in an enantioselective fashion. In a seminal report, the Nagorny group has used a similar dihyropyranyl moiety as a precursor for enantioselective spiroketalization utilizing a BINOL-based chiral phosphoric acid (Figure IV-6).17 In their procedure, a tertiary alcohol intercepts the oxocarbenium 273 intermediate en route to the spiroketal formation (Figure IV-6). They have further investigated the mechanism of their spiroketalization reaction as well as the origin of the selectivity through experimental and computational studies. Their results revealed a highly diastereoselctive syn-selective protonation and nucleophilic addition mechanism in the course of spiroketalization (Figure IV-6).18 O R1 ( )n IV-15 RR OH cat. (5 mol%) pentane RR ( )n O O IV-16 R1 O D HO Bn Bn IV-18 D cat. (5 mol%) pentane O H O Bn Bn IV-19 + H O D O Bn Bn IV-20 99:1 syn- H–O delivery iPr iPr iPr O P O OH O iPr iPr IV-17 iPr Figure IV-6. Nagorny’s approach to access spiroketals in an enantioselective fashion. Another elegant strategy to forge the spiroketal framework is reported recently by Carreira and coworkers.19 In this orthogonal approach, the hydroxyketone derivative IV- 22 undergoes formation of p-allyl complex in the presence of iridium–(P, olefin) complex (Figure IV-7). The ketone which is in equilibrium with its hemiacetal moiety via the pendent hydroxyl group, exposes a nucleophile for interception of the metal-allyl complex, consequently forming the spiroketal framework. The chirality of the phosphoramidite based ligands determine the face selectivity of asymmetric allylic substitution. Mutarotation of the equilibrating hemiacetal intermediate provides a mechanism to set the 274 stereochemistry on the acetal center under the influence of the chiral ligand (Figure IV- 7). OBoc O OH IV-22 O P O (S)-25 (4 mol%) [{Ir(cod)Cl}2] (1 mol%) Zn(OTf)2 (2 mol%) acetone (0.25 M) 4 °C, 24 h O O IV-24 IrL* ring-chain tautomerism IrL* O O (R) HO IV-23c Figure IV-7. Carreira’s approach to access spiroketals in an enantioselective fashion. (S) HO IV-23b Mutarotation OH O IV-23a IrL* is not well explored. Halogenated natural products are Despite excellent advances in the spiroketalization reaction, synthesis of halogenated spiroketals frequently encountered. In fact, more than 5000 halogenated natural products had been discovered as of 2011.20 Some of these halogenated natural products contain the spiroketal moiety in their skeleton (Figure IV-8).21 A robust and diastereoselective synthesis of halogenated spiroketals can provide a route towards the total synthesis of these compounds. 275 Me Me Br Me OH Me Me Me Br pinnatol (IV-27) dectylone (IV-26) OH Me O Me Me Br Cl Cl Me Br Cl (+)-halomon (IV-28) Br H Br O H O H O obtusin (IV-29) H Br H Br O H H O O H C chinzallene (IV-30) Br H Br Br O O Cl H O Me C obtusallene V (IV-31) Br Figure IV-8. Examples of halogenated natural products. The halogenated spiroketals are highlighted in blue. IV-3 Development of halenium initiated spiroketalization Our approach for the synthesis of halogenated spiroketals stems from our group’s interest in the halofunctionalization of olefins. So far, we have disclosed several methodologies for functionalization of olefins.1, 22-25 These reactions are generally initiated by an active halenium source that adds to olefins, initiating further functionalization to the this field.24 This report stands as (Figure IV-9). Enantioselective chlorofunctionalization of the unsaturated caboxylic acids IV-32 that leads to the synthesis of chiral chlorolactones IV-33 was our introductory methodology first asymmetric chlorofunctionalization of olefin that was empowered by the unique catalytic effect of the dimer cinchona alkaloid chiral catalyst [DHQD2PHAL].26 In a similar strategy, the halofunctionalization of unsaturated amides gave access to chiral halogenated oxazoline products.22-23 Our mechanistic studies led to the interrogation of the face selectivity of the halenium and the nucleophile addition via extensive deuterium labeling experiments. In this investigation, the relative and absolute selectivity of the incoming halenium and 276 nucleophile were investigated. These studies revealed that syn or anti addition across the olefin depends on the nature of the nucleophile and all halocyclization reactions investigated, the chlorenium face selectivity was the same.23 R/Ar Cl O O IV-33 CO2H IV-32 Ar/R N O N O N N O N N (DHQD)2PHAL (IV-34) n-PrOH R/Ar O O R/Ar H N IV-39 O O NH IV-40 Cl O Cl N R R O N Cl O R = Ph: DCDPH R = Me: DCDMH CHCl3-Hex O O Ar/R NH IV-41 R2 O R1 IV-35 N H Ar Ar O Ar NH IV-37 O R2 R1 IV-36 Ar N Cl Ar Cl O N Ar IV-38 Figure IV-9. Some of the developed methodologies by our group in the field of halofunctionalization of olefins. During the development of halofunctionalization reactions, we realized the need to introduce a parameter that define the affinity of various functional groups for absorbing a halogen. Therefore, a new scale that defines the thermodynamic tendency of a given nucleophile to capture the halenium ion was introduced.1 The halenium affinity (HalA) scale ranks various acceptors (olefins for instance) embedded in a molecule based on their ability to stabilize a “free halenium ion”, thus providing a mean for prediction of the path of reactions and product distributions. 277 HalA=127.2 Kcal/mol t-Bu N t-Bu t-Bu N t-Bu HalA=126.1 Kcal/mol IV-42 1.0 equiv Me IV-43 1.0 equiv S Cl CDSC 1.0 equiv acetone-d6 -90 ºC SbCl6 t-Bu N Cl SbCl6 t-Bu IV-42-Cl Me SbCl6 t-Bu N Cl t-Bu IV-43-Cl ΔHalA = 1.1 kcal/mol B3LYP/6-31G*/SM8 (acetone) predicts 1:7 ratio integral ratio of A-Cl:B-Cl =12:88 ~ 1:7.3 IV-43-Cl B-Cl (C3-H) IV-42-Cl A-Cl (C4-H) IV-42-Cl A-Cl (C3-H) IV-42 A (C4-H) IV-42 A (C3-H) B IV-43 (C3-H) Figure IV-10. Experimental validation of the halenium affinity (HalA).1 The direct implication of the halenium affinity can be experimentally monitored.1 It was noted that when equimolar ratio of two pyridines with different halenium affinities are subjected to a competitive reaction with one equivalent of chlorenium ion, the distribution of the chlorinated pyridines can be observed with H-NMR at low temperature (Figure IV- 10). The ratio of the two pyridinium products is in agreement with the predicted ratio from the computationally calculated HalA values. In this study the pyridine with a methyl group at the para position IV-43 has higher tendency to capture the chlorenium leading to IV- 43-Cl. This provides a predictive tool for the course of a reaction, as well as it can be 278 utilized to design new reactions and interrogate the mechanism of halogenation reactions. Parametrization of the affinity of olefins for halenium capture led to a revised mechanistic understanding of halofunctionalization of olefins. the classical assumptions the role of the nucleophile in the halofunctionalization of olefins is neglected. However, empowered by the HalA, our group was able to demonstrate that the halenium affinity of an olefin can dramatically affect the role pendent nucleophile in the course of the reaction.27 We noted a direct and strong correlation between the rate of halofunctionalization and the strength of the nucleophile (Figure IV-11). Thus, the In nucleophile is involved in the halenium capture by the olefin. We proposed nucleophile- assisted alkene activation (NAAA), in which the preorganization of the olefin with the tethered nucleophile facilitates the halofunctionalization reaction.27 As such, a stronger nucleophile (carboxylate anion in this instance) exposes a more reactive olefin for halofunctionalization reaction (Figure IV-11). Ph Nuc O O O Nuc DCDMH CHCl3, rt rxn time 72 h 12 h 20 min <2 min OH OH O H N O NBu4 Nuc Ph Cl energy (kcal/mol) activation 27.7 16.7 14.3 8.8 Aδ- δ+ Cl δ- δ+ Nuc nucleophile induced pre-polarization Aδ- Cl δ+ Nuc nucleophile participation in RDS Figure IV-11. The effect of nucleophile on the olefin halofunctionalization, demonstration of nucleophile-assisted alkene activation (NAAA).27 279 IV-4 Development of mono and di-brominated spiroketalization reaction We sought to investigate the halofunctionalization with a ketone as the tethered nucleophile. The latter reaction would result in formation of oxocarbinium species that can lead to spiroketals if intercepted by a tethered alcohol moiety. Ketone IV-11 bearing both alcohol and olefin was envisioned to undergo halenium (bromenium) capture with the aid of the ketone functioning as a weak nucleophile (Figure IV-12). The latter event exposes a oxocarbenium intermediate that can be intercepted by the tethered alcohol to furnish the spiroketal. This would result in the generation of three stereocenters, nonetheless, with complete diastereoselectivity control (Figure IV-12). R X O IV-11 X = Br, I halenium initiated spirocyclization OH O R X * * * O newly formed stereocenters O O * IV-13 * RX * Figure IV-12. Our approach to access halogenated spiroketals. Ketone IV-44 was synthesized and subjected to the reaction with the bromenium source (Figure IV-13). To our surprise, we isolated spiroketal IV-45 only as a minor product. The major product of the reaction was identified as the macrocycle IV-46 structure was confirmed by X-ray crystallography (Figure IV-14). Despite the fact that synthesis of the macro-cyclic side product is not the main aim of our program, the current methodology can potentially be tuned to access this class of molecules that are challenging to synthesized by the known methods.28-31 The product distribution of this reaction points to the fact that the ketone is most likely in a pre-equilibrium with the its hemiacetal IV-44a and IV-44b. Hemiacetal IV-44a/b exposes two nucleophilic oxygens 280 that can act as a nucleophile for interception of the activated olefin via the bromenium ion. Ph O IV-44 DCDMH, DBDMH (1 equiv each) toluene, 4 h, rt 89% overall yield OH Br Ph O O IV-45 Br Ph IV-46 O O + 1:3 Halenium capture Halenium capture Br Ph IV-47 O OH Br Ph O OH IV-48a spiroketalization macrocyclization Br Ph O OH IV-48b Ph IV-44a O OH minor Ph IV-44b O HO major mixture of hemiketals Ph IV-49 DCDMH, DBDMH (1 equiv each) toluene, >98% conversion OH PhBr O IV-49b this product was not observed Br Ph O IV-49b Figure IV-13. Bromo spiroketalization of ketone tethered to an unprotected alcohol. In such a manner, if the hydroxyl group of the equilibrating hemiacetal IV-44a acts as the nucleophile, bromo spiroketal IV-45 would form as product of the reaction. However, if the other oxygen of hemiacetal, which is imbedded in the six membered ring in IV-44b is the nucleophile, macrocyclic IV-46 will form. To confirm that macrocycle IV-46 originates from hemiacetal intermediate IV-44a/b, unsaturated alcohol IV-49 was 281 synthesized and subjected to the same reaction conditions, however, macrocycle IV-49b was not obtained. The only difference between the alcohol and the keto-alcohol is the presence of the ketone moiety on the alkyl chain of keto-alcohol IV-44 (Figure IV-13). The ketone functionality in IV-44 can provide a route to facilitate the entropically less favorable pathway that leads to macrocycle IV-46. The presence of the equilibrating hemiacetal reduces the entropic barrier in comparison to the direct macrocyclization of IV-49. Presence of the hemiacetal is in agreement with Carreira’s report shown in Figure IV-7. In order to circumvent this side product, we sought to protect the tethered alcohol, to prevent the formation of hemiacetal intermediates that lead to the macrocyclic product. Br IV-46 O O IV-46 Figure IV-14. X-ray crystal structure of macrocyclic IV-46. In pursuit of the appropriate agent, various protecting groups were screened. Most of the protecting groups did not allow the protected alcohol to form a spirokatal (Figure IV- 15). When alcohol in IV-50 was protected with MOM group, low conversion of the ketone to the spiroketal IV-45 was observed. Employing THP as a protecting group improved the conversion of the reaction and the spiroketal was isolated in excellent yield and diastereoselectivity (Figure IV-15). Addition of ethanol was crucial for this reaction as it facilitates removal of the THP group from intermediate IV-51, furnishing spiroketal formation. 282 Ph O IV-50 O Pg DCDMH, DBDMH (1 equiv each) toluene, rt EtOH (10.0 equiv) Br Ph O O IV-45 + recovered starting compound IV-50 Br Ph halenium capture O O O IV-51 EtOH IV-50a Pg = TBS, TES, PMB <2% IV-50b Pg = MOM IV-50c Pg = THP 20-35% >99% (>98% dr) bromo spiroketal IV-45 >95% 10-18% <2% Figure IV-15. Bromo spiroketalization of ketone with a protected tethered alcohol. This result was justified by the fact that the ketone is a weak nucleophile, thus it is incompetent to aid is the halo-functionalization of the olefin. However, introduction of an alcohol can improve the nucleophilicity of the ketone towards halo-functionalization of olefin. Of note, the putative NAAA pathway facilitate by the hydroxyl of the hemiacetal activates the olefin more than the parent ketone. The protection of the alcohol is required in order to avoid macrocyclization, as discussed above. The latter experimental results are in full agreement with the computationally obtained halenium affinities of the olefin in the presence of a ketone engaged as a hemiacetal with the pendent hydroxyl, in its free and protected forms (Figure IV-16). 283 Br -OH attack Br R-O-R attack Br Br O O IV-47 H PhBr H O PhBr O IV-48a O IV-52a PhBr O O O IV-51 PhBr HalA (Br) kcal/mol 148.4 142.9 140.7 141.6 Ph Ph Ph HO O IV-44a/b O IV-52 O THPO IV-50c Figure IV-16. Computed HalA values for ketones bearing different groups. As illustrated in Figure IV-16, ketone bearing a free alcohol, which is in equilibrium with its hemiacetal form, exposes two nucleophilic oxygens for halofunctionalization of the olefin. IV-47 and IV-48a have higher HalA value as compared to methyl ketone IV-52. The methyl ketone that shows the lowest HalA value resembles substrates with protecting groups such as TES, TBS and PMB (Figure IV-15) that were incompetent to furnish spiroketalization. Ketone with THP as the alcohol’s protecting group showed higher HalA as compared to the methyl ketone yet lower than IV-47 and IV-48a. These data are in full agreement with the experimental results depicted in Figure IV-13 and IV-15. It must be noted that prototypical reagents such as N-bromosuccinimide and 1,3-dibromo-5,5- dimethylhydantoin, proved to be slow and less efficient for this transformation. Hence, we i. 3-bromo-1-chloro-5,5- resorted to a more active bromenium source e. 284 dimethylhydantoin, which was generated in situ by combining equimolar amounts of DBDMH and DCDMH. To demonstrate the generality of this reaction, keto-alkenols shown in Table IV-1, incorporating the tetrahydropyranyl protection were subjected to the optimized conditions. Table IV-1 summarizes the results of halenium atom initiated cascade spiroketalizations that furnished the substituted 3-halo-1,7-dioxaspiro[5,5]undecanes (45, 53-60) in a diastereoselective fashion. Modulating the electronic or steric profile of the substituents attached to the olefins in Table IV-1 did not make any significant difference to the overall reaction outcome. The corresponding [6,6]-spiroketals were obtained in excellent yields and diastereoselectivity. The relative stereochemistry in these halogenated spiroketals was established unequivocally by the crystal structure of product IV-54 and NOESY analysis of IV-45 (Figure IV-17). H H nOe H Br O H O H O H IV-45 Figure IV-17. X-ray crystal structure of product IV-54 and NOESY of IV-45. H O H IV-54 Br As with the bromenium source, an efficient diastereoselective cascade spiroketalization could also be initiated using N-iodosuccinimide as an iodenium source (entry 2, Table IV-1). Several attempts to initiate the corresponding fluorofunctionalization led only to products of a-fluorination of the ketone, whereas chlorofunctionalization using 285 trichloroisocyanuric acid gave a complex mixture of undesired alcohol oxidation products resulting from the in situ oxidative deprotection of the -OTHP group. Table IV-1. Substrate scope for halenium atom initiated diastereoselective R2 O R1 entrya 1d 2e 3 4 5 6f 7g 8 9 10 R1 Ph Ph b-naphthyl a-naphthyl p-CH3-C6H4 o-CH3-C6H4 p-CH3O-C6H4 p-F-C6H4 p-Ph-C6H4 Ph spiroketalization. DCDMH, DBDMH (1 equiv each) OTHP toluene, rt, 3-4 hours EtOH (10.0 equiv) O R2 R1Br O R2 H H H H H H H H H CH3 product IV-45 IV-45-I IV-53 IV-54 IV-55 IV-56 IV-57 IV-58 IV-59 IV-60 Yieldb >99% 82% 93% 88% 87% 86% 65% 84% 83% 77% drc >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 [a[a] Reactions were performed using 0.1 mmol of substrate. [b] Isolated yields. [c] drs were estimated by 1H NMR (500 MHz) analysis. [d] Reaction was performed on a 1.0 mmol scale. [e] Reaction was performed for 12 h at rt using 1.1 equiv N-iodosuccinimide as a halenium source. [f] After 3 h, conc. HCl (~10 mol%) was added and the reaction was continued for another 20 min prior to quench. [g] 1.1 equiv of NBS was employed as the bromenium source. 286 Substrate IV-61 that undergoes a [6,5]-spiroketalization via the intermediacy of a similar six-membered oxocarbenium, furnishes the corresponding product as a single diastereomer (Figure 18). In contrast, a [5,5]-spiroketalization of the keto-alkenol 21 results in a 2:1 mixture of diastereomers. The latter observation is due to the skewed framework of [5,5]-spiroketals that unlike the [6,6]-spiroketals, does not benefit fully from the anomeric effect. Hence, during the formation of [5,5]-spiroketals, the diastereomeric transition states are not energetically biased enough to promote a significant product selectivity, without any external aid. Ph THPO IV-61 O DCDMH, DBDMH (1 equiv. each) PhBr O O toluene, rt, 3 h EtOH (10.0 equiv) 82% IV-62 >98:2 dr crystal structure of IV-62 n-Bu O DCDMH, DBDMH (1 equiv. each) O O OTHP IV-63 toluene, rt, 3 h EtOH (10.0 equiv) H IV-64 1:1.6 dr Figure IV-18. Synthesis of 5,6- and 5,5-spiroketals. 63% Br n-Bu We also exploited the stereoelectronics associated with the 6-membered oxocarbenium towards a dual functionalization of the above keto-alkenols. As depicted in Figure IV-19, when ethanol was excluded from the reaction mixture, the oxocarbenium intermediate putatively undergoes a-deprotonation to generate vinyl ether IV-61. This intermediate can further proceed through bromenium capture leading to dibrominated IV- 62 that eventually furnishes product IV-64. The second bromenium is captured from a 287 pseudo-axial approach that leads to the high diastereoselectivity of the product. This dibromo-spiroketal scaffold is unique to the bioactive natural products derived from the family of Laurencia obtusa and others.13, 32-34 Ph Br Ph O IV-50c halenium capture H O O O IV-51 OTHP NBS (2.5 equiv) 12 M HCl (cat) CHCl3, 2 h, rt A Br A–Br = NBS Br Br Ph O no EtOH O Ph pseudo-axial IV-61 attack O THP IV-62 Br O Br O Ph IV-64 >98:2 dr Br Br Br Ph O O THP IV-63 EtOH bromo spiroketal IV-45 Figure IV-19. Dibromospiroketal formation by excluding ethanol from the reaction mixture. Table IV-2 summarizes the results for a handful of keto-alkenols that were subjected to the optimized conditions for di-bromospiroketalization. Catalytic amount of HCl was necessary to furnish the desired transformation. Acid can potentially activate N- bromosuccinimide for the latter transformation. Moderate to excellent yields were obtained with various substituted keto-alkenols. As in the monobromo-spiroketalization, the sterics and electronics of the substituted olefins did not have a significant effect on the overall outcome. The relative stereochemistry was established by the crystal structure of IV-67 (Figure IV-20). The [6,5]-spiroketalization was equally efficient as the [6,6]- spiroketalization (compare entries 1 and 2, Table IV-2), however the [5,5]-ketalization was less selective owing to the skewed stereoelectronics of the oxocarbenium embedded in 288 a 5-membered ring (entry 9). Table IV-2. Diastereoselective dibromo spiroketalization R2 R1 O ( )m (n,m = 0 or 1) ( )n OTHP NBS (2.5 equiv) 12 M HCl (cat) CHCl3, 2 h, rt R2 R1Br O ( ) O n ( )m Br entrya 1d 2e 3 4 5 6f 7g 8 9 10 R1 Ph Ph b-naphthyl a-naphthyl p-CH3-C6H4 o-CH3-C6H4 p-F-C6H4 p-Ph-C6H4 H CH3 R2 H H H H H H H H n-Bu CH3 (n, m) product Yieldb (2, 2) (1, 2) (2, 2) (2, 2) (2, 2) (2, 2) (2, 2) (2, 2) (1, 1) (2, 2) IV-64 IV-65 IV-66 IV-67 IV-68 IV-69 IV-70 IV-71 IV-72 IV-73 87% 73% 88% 61% 74% 85% 65% 59% 88% 70% drc >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 >98:2 2:1 1:1 [a] Reactions were performed using 0.1 mmol of substrate. [b] Isolated yields. [c] dr’s were estimated by 1H NMR (500 MHz) analysis. [d] Reaction was performed on a 1.0 mmol scale. [e] Reaction was performed for 12 h at rt using 1.1 equiv N-iodosuccinimide as a halenium source. [f] After 3 h, conc. HCl (~10 mol%) was added and the reaction was continued for another 20 min prior to quench. [g] 1.1 equiv of NBS was employed as a bromenium source. 289 The [6,6]-spiroketalization of the tri-substituted olefin in 19 furnished a 1:1 diastereomeric mixture as the energetics of the two conformers (ring flipping) in the corresponding enol-ether intermediate (similar to IV-61 in Figure IV-19) are not biased due to the presence of the gem-dimethyl substitution. O H O H Br IV-67 Br Figure IV-20. X-ray crystal structure of dibromo-spiroketal IV-67. In summary, we have demonstrated the application of Halenium Affinity (HalA) as a guiding tool towards modulation of nucleophilic strength, leading to a diastereoselective spiroketalization. Unaided ketones were found to be inefficient nucleophiles towards olefin activation; however, the tethered alcohol provided a handle to modulate the ketone’s nucleophilic strength. Calculation of the HalA values revealed the effect of tethered alcohol on the halogen affinity of the olefin. THP protected tethered alcohol proved to be optimal for the spiroketalization reaction. Additionally, the spiroketalization was rerouted to give access to the dibrominated spiroketals in a diastereoselective manner. 290 IV-5 Towards the total synthesis of Obtusin IV-5-1 Introduction With the successful development of bromo spiroketalization, especially the diastereoselective dibromo spiroketalization, we sought to utilized this methodology to access Obtusin IV-74, a natural product isolated in 1979 from the Mediterranean alga Laurencia obtusa.33-34 From the structural standpoint, Obtusin contains eight contiguous stereocenters and a skeleton featuring three five-membered ether rings, two of which are part of a spiroketal moiety (Figure IV-21). This molecule poses a synthetic challenge and its chemical synthesis has not been reported thus far. Our strategy towards Obtusin takes advantage of the crucial rule of protecting groups to access the desirable selectivity in forging the skeleton of this molecule. We will begin with a molecule from chiral pool followed by sequential diastereoselective synthetic steps towards obtaining the target. Br O 3 4 Br 13 12 H 7 6 H O 9 10 Br O Obtusin (IV-74) bromination via enolization 12 13 9 10 HO O IV-75 HO 7 6 4 3 R Br bromo etherification; anti addition to cis double bond Br bromo spiroketalization; anti addition to trans double bond Figure IV-21. Structure of Obtusin and its proposed biogenesis. 291 Our retrosynthetic approach to Obtusin is inspired by the proposed biosynthesis of this molecule. Obtusin can be unraveled to the linear polyketide structure IV-75 (Figure 22).35 The stereochemistry at the chiral centers indicates anti addition across a trans C12- C13 double bond and a cis C3-C4 double bond. Based on our retrosynthetic approach, IV-76 can be obtained from IV-77 using our dibromo spiroketalization. A bromo etherification reaction can be envisioned to construct the THF ring in IV-77 from the linear carbon chain of IV-75. The carbon chain skeleton of IV-75 with the central chiral diol will originate from the addition of the appropriate carbon chains to the chiral diepoxide IV-78 (Figure 22). Br O 3 4 Br 13 12 O 9 H 7 6 H O 10 Br Obtusin (74) O 3 4 Br R H 7 6 H O R = CH2OTBDPS Br 13 12 IV-76 O 9 10 Br dibromo spiroketalization installation of alkyl chains O 6 7 IV-78 O Br R 3 4 O 7 6 IV-77 9 O O THP 12 13 bromo etherification R 3 4 OH O 6 7 9 OTHP IV-75 13 12 Figure IV-22. Retrosynthetic approach to access Obtusin featuring dibromo-spiroketalization and bromo-etherification steps. 292 IV-5-2 Progress towards the total synthesis of Obtusin As illustrated in Figure IV-23, our synthesis of Obtusin commences with enantio-pure diepoxide IV-78 that contains the correct stereochemistry on the central di-carbinol centers (C6 and C7). Synthesis of diepoxide IV-78 is reported and amenable to multi gram scale starting from D-(–)-tartaric acid.36-37 As illustrated in Figure IV-23, D-(–)- tartaric acid is converted to the enantio-enriched diepoxide IV-78 in five steps with high overall yield.36-37 This epoxide is the lynchpin for installation of the rest of the carbon atoms of the natural product. To this end, a sequential selective opening of the diepoxide was planned. Alkynyl anion of IV-79, as the surrogate for the cis olefin, was added to diepoxide IV-78.38 Mono-epoxide IV-80 was isolated in moderate yield. Competing diol formation that originated from double addition of the alkynyl anion to the diepoxide IV-78 was the unavoidable side product in this reaction. The alkyne functionality in IV-80 was then partially reduced to the corresponding cis double bond in IV-81 in nearly quantitative yield. The optimization of the reduction reaction showed that longer reaction time would result in the formation of a complex mixture (possibly over reduced product). Thus, a 30- minute reaction time was identified as optimal where the reduction is complete and side product formation is minimal. The alcohol of IV-81 was then protected as a THP. As outlined in the previous section, this protecting group is essential for the dibromo spiroketalization reaction. This protection led to a separable 2:1 diastereomeric mixture. The major diastereomer was carried in the following steps to avoid complications in NMR assignment, although the stereochemistry if inconsequential, as it is not present in the final target (Figure IV-23). 293 TBDPS O IV-79 1) nBuLi 2) BF3 •OEt2 3) O O IV-78 5 h, 60% O TBDPS O O TBDPS OH IV-80 Lindlar's Cat. H2 (gas) EtOAc 30 min, quant. O IV-82 O THP DHP PPTS, DCM 12 h, 85% ~2:1 dr on the THP group O TBDPS O IV-81 OH HO OH O OH O OH D-tartaric acid 5 steps ca 45% O R R IV-78 O Figure IV-23. Installation of the cis olefin, access to epoxy alcohol IV-91. We then turned our attention to opening of the epoxide in IV-82 in order to install the ketone moiety along with introducing the rest of the alkyl chain (Figure IV-24). Our first was to use dithiane IV-83. However, under various reaction conditions, the target alcohol IV-84 was not formed. We realized that the reaction of dithiane with epoxide IV-82 requires addition Lewis acids (such as BF3•Et2O) to catalyze the reaction. Nonetheless, using Lewis acids to facilitate the latter transformation resulted in the removal of the dithiane group and formation of other side products. We next took a different approach for opening of this epoxide as a path for installation of the ketone moiety, namely, to use a nitrile as the nucleophile. Epoxide IV-82 was readily reacted with potassium cyanide in methanol, yielding alcohol IV-86 in good yield (Figure IV-24).39 294 S S IV-83 1) nBuLi 2) TBDPS O O TBDPS O OH S S O THP IV-84 IV-82 O THP many different conditions failed to give access to epoxide opening using dithiane anion. O TBDPS OH O O TBDPS O THP O IV-82 KCN Methanol 87% O TBDPS O THP IV-85 OH CN O THP IV-86 Figure IV-24. Installation of the ketone surrogate, towards bromo-etherification reaction. With IV-86 in hand, bromo-etherification of the cis olefin was targeted. As illustrated in Table IV-3, bromo etherification of IV-86 resulted in the formation of diastereomeric tetrahydrofuran rings IV-87 and IV-88. The latter result stems from the opposing face selectivity of the cis olefin in the bromo etherification step (re face attack leading to IV-87 while si face attack yields IV-88). Reaction in DCM as the solvent and 2,4,4,6-tetrabromo- 2,5-cyclohexadienone (TBCO) as the bromine source led to the formation of undesired diastereomer IV-88 as the major diastereomer (>20:1 dr), (entry 1, Table IV-3). 295 TBDPS O THP O IV-86 entrya Table IV-3: Bromo etherification of alcohol IV-86. OH Br conditions CN Br TBDPSO OTHP CNH O H IV-87 desired diastereomer OTHP O H CNH + Br TBDPSO IV-88 undesired diastereomer solvent DCM toluene DMF methanol acetonitrile acetonitrile/TFE (1:1) THF:HMPA (50:1) THF:HMPA (50:1)e THF:HMPA (50:1)f THF:HMPA (50:1)g 1 2 3 4 5 6 7 8 9 10 conversionb dr (IV-96:IV-97)c >99 >99 >99 >99 >99 >99 >99d >99 30 >99 1:20 1:1.6 1:1.6 1:1.5 1:1.6 1:1.7 (1:1) (1:1) (1:1) (1:1) [a] Reactions are performed in 0.1 mmol scale unless otherwise noted. [b] Conversions are estimated from 1H-NMR using tert-butyl methyl ether as internal standard. [c] Diastereomeric ratios are estimated from 1H- NMR using tert-butyl methyl ether as internal standard. [d] Isolated yields for IV-87 and IV-88 are 45% and 49% respectively. [e] NBS was used instead of TBCO. [f] Reaction was performed at room temperature. [g] Reaction was performed at –78 °C. 296 Further optimization of the reaction revealed that more polar solvents can improve that ratio of the desire diastereomer. As such, when THF with HMPA as co-solvent was used ~1:1 ratio of the diastereomeric products was isolated. TBCO, a commonly used as a mild bromine source for bromo etherification reaction,40-41 gave poor diastereoselectivity and so did other bromine sources such as NBS. (Table IV-3, entry 8). Reactions at different temperatures also did not lead to any improved selectivity for the desired product (entries 9 and 10, Table IV-3). The relative stereochemistries of the diastereomers IV-87 and IV-88 were established using extensive 2D NMR analysis. As illustrated in Figure IV-25, for the desired diastereomer IV-87, nOe correlations between hydrogen atoms on both ethereal carbon atoms confirm their syn geometry. However, for the undesired diastereomer IV-88, nOe between these two hydrogen atoms was not observed, while a series of the other nOe correlations confirmed the illustrated relative stereochemistry. H H Br OTHP CN TBDPSO O H H nOe IV-87 desired diastereomer nOe nOe nOe H Br H H OTHP CN nOe O H H TBDPSO NO nOe IV-88 undesired diastereomer Figure IV-25. Assignment of the relative stereochemistry of the diastereomers IV-96 and IV-97. 297 With these results in hand, we sought to identify the intrinsic factors that govern the latter selectivity. To better understand the origin of the observed selectivity in this reaction, we turned our attention to the approaches of the nucleophilic oxygen and the bromenium for the reaction with the cis olefin. As illustrated in Figure IV-26, since the addition across olefin is anti, the face selectivity of the olefin would result in the formation of the two diastereomers (IV-87 and IV-88). In return the face selectivity of the cis double bond is dictated by the stereocenters on IV-86. Thus, the pre-organized structure IV-86a would lead to the desired diastereomer, and IV-86b would lead to the undesired distereomer IV- 88. In this putative arrangement, the allylic groups on the cis double bond would have a pseudo syn-pentane interaction with the group on C-7 that is bearing the nucleophilic oxygen atom. This steric repulsion would disfavor IV-86-a, and as a result the desired diastereomer will form as the minor product. On the other hand, in IV-86-b, which leads to the undesired diastereomer, the bulky allylic group on C-3 points to the opposite side of the groups on C-7 and C-6, thus a facile bromo etherification ensues, leading to IV-88 as the major product. In order to reverse the intrinsic selectivity, the protecting group on of C-6 oxygen was removed. The hypothesis was that with a smaller group on C-6, the carbon chain may adopt a different conformation as compared to IV-86, thus giving access to the desired diastereomer as the major product. To this end, IV-89 in which the THP group was removed, was subjected to the bromination conditions (dashed box, Figure IV-26). The reaction resulted in a 3:1 diastereomeric ratio in favor of the undesired diastereomer IV-91 (Figure IV-26). 298 THP O TBDPS O IV-86 HO CN Br TBDPS O Br H 3 4 H 6 HO THP CN O H IV-86-a1 7 + Br H H 4 3 6 THP CN O 7 H IV-86-b1 HO O TBDPS TBDPS O OH CN HO IV-89 THF:HMPA IV-90:IV-91 0°C (1:3) OH CN IV-90 O H H + OH O H H CN IV-91 TBCO Br TBDPSO Br TBDPSO steric repulsion δ+ Br H 3 4 H TBDPS O THP O 7 6 CN H O H δ+ IV-85-a2 H δ+ Br H 4 3 O THP O 7 6 CN H δ+ O H TBDPS IV-86-b2 disfavored trajectory for addition to the olefin favored trajectory for addition to the olefin OTHP CN H O H Br TBDPSO + Br TBDPSO OTHP CN H O H IV-87 desired diastereomer IV-88 undesired diastereomer Figure IV-26. On the origin of the diastereoselectivity leading to IV-87 and IV-88. Since going small on C-6 did not solve the undesired stereochemical problem, we decided to approach the solution in a complete opposite manner. We proposed to impose an artificial bulkiness on the C-7 hydroxyl (the OH involved in the bromo etherification) to disfavor the approach that leads to the undesired diastereomer IV-88. We hypothesized a group capable of templating, yet easily removable on this oxygen atom would provide a path to reverse the intrinsic selectivity. The hypothetical group on the nucleophilic oxygen would suffer from a syn-pentane interaction with the groups on the allylic position 299 of the cis olefin, thus the approach highlighted in IV-86-b would suffer, leading to switch in the diastereoselectivity. THP O TBDPS O IV-92 O Pg TBDPS O CN Br Br H δ+ Br H H 4 H O 3 3 4 THP CN O 7 H + Br IV-92-a1 6 O Pg H H 4 3 O THP CN O 7 H IV-92-b1 6 O Pg TBDPS TBDPS THP O 7 CN H IV-92-a2 6 O δ+ Pg H 4 δ+ Br H 3 THP O 7 CN H IV-92-b2 6 O δ+ Pg favored trajectory O TBDPS disfavored trajectory steric repulsion OTHP Br TBDPSO O H H CN + Br TBDPSO OTHP CN H O H IV-87 desired diastereomer IV-88 undesired diastereomer TBDPS O THP O CN HO IV-86 TMSCl Et3N, DMAP (cat.) DCM, r.t. quant. TBDPS O THP O CN IV-92 O TMS Figure IV-27. Rational design to improve the diastereoselectivity of the bromo-etherification reaction. To test this hypothesis, trimethylsilyl group as a large, yet easily removable group during the bromo etherification, was chosen (Figure IV-26). Alcohol IV-86 was reacted with TMSCl to provide the protected alcohol IV-92 in nearly quantitative yield. With IV-92 300 in hand, bromo etherification was implemented (Figure IV-27). To our delight, employing THF/HMPA as the solvent system leads to the desired diastereomer IV-87 as the major product with >10:1 selectivity (entry 1, Table IV-4). Of note is that under similar conditions, the unprotected alcohol IV-86 led to a 1:1 diastereomeric mixture of IV-87 and IV-88 (compare entry 7, Table IV-4 with entry 1, Table IV-3). As illustrated in Table IV-4, further optimization of the reaction led to DCM as the solvent of choice, with methanol (10.0 equiv), as the scavenger of TMS group (entry 9). The desired product IV-87 was isolated in >85% yield along with small amount of the undesired diastereomer (<5%). Table IV-4: Bromo etherification of protected alcohol IV-101. THP O TBDPS O CN Br condtion O IV-92 TMS solvent THF DMC THF DCM DCM chloroform entrya 1 2 3 4 5 6 7 OTHP Br TBDPSO + O H H CN Br TBDPSO OTHP O H H CN IV-87 desired diastereomer IV-88 undesired diastereomer additive conversionb dr (IV-96:IV-97)c HMPA (2% v:v) none none water (10% v:v) MeOH (10% v:v) MeOH (10% v:v) >99d 25 90 40 >99 >99 >99 >10:1 >10:1 2:1 >10:1 14:1 >15:1 15:1 DCM MeOH (10% v:v)e 301 Table IV-4 (cont’d) entrya 8 9 10f solvent DCM DCM DCM additive conversionb dr (IV-96:IV-97)c EtOH (10% v:v) MeOH (10 equiv) MeOH (10 equiv) >99 >99 >99g >10:1 >15:1 >15:1 [a] Reactions are performed on a 0.1 mmol scale unless otherwise noted. [b] Conversions are estimated from 1H-NMR using tert-butyl methyl ether as internal standard. [c] Diastereomeric ratios are estimated from 1H-NMR using tert-butyl methyl ether as internal standard. [d] Isolated yields for IV-87 and IV-88 were 75% and 8%, respectively. [e] Reaction was performed at 0 °C. [f] Reaction was performed on a 1.0 mmol scale. g Isolated yields for IV-87 and IV-88 were 85% and 4%, respectively The results of TMS protected and unprotected alcohol for bromo-etherification in DCM highlights the significant effect of the protecting group to improve the dr. As depicted in Figure IV-28, the 1H-NMR traces of the crude reaction mixtures with and without the protecting group on the nucleophilic oxygen atom under similar conditions result in the formation of the opposite diastereomer in nearly exclusive manner. 302 THP O TBDPS O CN O R R = H (IV-86) R = TMS (IV-92) TBCO DCM, rt additive OTHP Br TBDPSO + O H H CN Br TBDPSO OTHP O H H CN IV-87 desired diastereomer IV-88 undesired diastereomer 1. R = H; No additve 2. R = TMS; methanol (10 equiv) IV-87:IV-88 dr: 1:20 IV-87:IV-88 dr: >15:1 Figure IV-28. Comparison of the bromo-etherification with and without protection group on the nucleophilic oxygen. With the decorated THF ring IV-87 in hand, further modification of the nitrile group to install the ketone moiety along with installation of the remaining of the carbon chain to access IV-93 was pursued. One of our early approaches was to subject the nitrile in IV- 87 to direct nucleophilic addition. For this purpose, in situ generated 2-hexenyl lithium was added to IV-87 at low temperature (Figure 29). The reaction afforded the ketone IV- 93 only in small quantity (<10% yield), returning the nitrile starting material as the major isolated compound. Reaction at higher temperatures or longer reaction times, resulted in complex mixture of products. Products originating from decomposition or degradation of the ring were the major side products of the reaction. Further optimization of this 303 reaction is necessary to achieve large quantities of ketone IV 103 in order to implement the key dibromo-spiroketalization. THP O H O H IV-87 CN Br TBDPSO Li 7% THF, –78 °C 65% IV-96 was recovered THP O O H O H IV-93 Br TBDPSO Figure IV-29. Direct addition of alkyl lithium to the nitrile moiety towards installation of the ketone functionality. Presence of the labile bromide on IV-87 could be the major issue that hampers the ketone formation. To remedy this issue, the nitrile was converted to the ketone prior to the bromo etherification step. However, the reaction of nitrile IV-92 with in situ generated 2-hexenyl lithium resulted in ketone IV-94 with only 30% yield (Figure IV-30, top). In order to improve the yield of ketone formation, an indirect approach was pursued. The nitrile group in IV-95 was reduced to the corresponding aldehyde IV-96 followed by addition of 2-hexeneyl lithium. The corresponding diastereomeric alcohol was then oxidized to install the ketone functionality leading to IV-98. Unfortunately, a complex reaction mixture was observed, hampering the identification of the product. A small fraction of the complex reaction mixture was isolated and identified as the furan derivative IV-99 (Figure IV-30). This product might have formed under oxidation condition via condensation/dehydration 304 TBDPS O THP O CN IV-92 O TMS Li THF, –78 °C 30% O TBDPS TMS O O O THP IV-94 TBDPS O TMS O CN IV-95 O TMS TMS O OH TBDPS O O TMS IV-97 DIBAL-H (3.0 equiv) DCM, -78 °C 71% TBDPS O TMS O IV-96 O TMS O Li THF, –78 °C 30% PCC (1.1 equiv) NaOAc DCM, rt TMS O O TBDPS O this product was not observed O TMS IV-98 This compound was the major product of the reaction O IV-99 Figure IV-30. Approaches to improve the efficiency of the conversion of nitrile to ketone. TBDPSO With ketone IV-94 in hand, the desired bromo functionalization reaction was attempted. However, due to the presence of multiple reactive sites (two double bonds, and nucleophilic oxygen atoms), complex mixture of products was observed. This results assured us that a sequential and stepwise implementation of the bromo etherification and dibromo spiroketalization reactions should be followed in order to minimize the complexity of the synthesis. Since protection of the epoxy alcohol IV-81 with the THP protecting group resulted in diastereomeric mixture, a parallel pathway was pursued in which the installation of this protecting group was delayed to a later stage of the synthesis (Figure IV-31). Epoxy 305 alcohol IV-81 was subjected to epoxide opening with cyanide to give rise to diol IV-89 which was immediately protected as TMS ether leading to the di-protected analogue IV- 95 in good yield. Bromo etherification of IV-95 under optimized condition led to IV-90 in high yield and excellent diastereoselectivity (>95:5 dr), after acidic workup of the reaction to remove the TMS group. This alcohol was then protected as a THP acetal (IV-100) in high yield as a ~1:1 diastereomeric mixture on the acetal stereocenter. Future plan is to use this compound to complete the total synthesis of Obtusin. TBDPS O IV-81 OH O KCN MeOH TBDPS O OH CN TMSCl Et3N DCM 75% (two steps) TMS O TBDPS O CN IV-95 O TMS IV-89 HO Br TBDPSO OTHP O H H IV-100 CN DHP (1.2 equiv) PPTS (cat.) 91%, 1:1 dr DCM Br TBDPSO OH H O H IV-90 CN TBCO (1.2 equiv) MeOH (10 equiv) DCM then HCl (cat.), MeOH 87%, >95:5 dr Figure IV-31. A different approach to convert epoxy alcohol IV-90 to advanced intermediate IV-109. At this point, we believe that the most hopeful strategy is the direct addition of the alkenyl group to the nitrile moiety in IV-87 or IV-100 to reveal the desired ketone. There are reported procedures that utilize organo zinc reagents as milder carbon nucleophiles for addition to nitriles. We are considering optimization of the direct addition of the alkyl group to the nitrile to give provide the ketone, setting the stage for the key dibromo spiroketalization. 306 In summary, we have demonstrated a streamlined synthesis that give access to the advance THF core of Obtusin. During the synthesis we uncovered the key role of the protecting group to control diastereoselectivity of the bromo etherification reaction to that installs the THF core of the natural product. Conversion of the nitrile to the ketone moiety proved to be challenging. Current efforts focus on the optimization of the latter transformation in order to implement the diastereoselective dibromo spiroketalization reaction. 307 IV-6 Experimental section IV-6-1 General remarks: Molecular sieves (4Å) were dried at 160 °C under 0.25 mtorr vacuum prior to use. Unless otherwise mentioned, solvents were purified as follows. CHCl3 (amylene stabilized) was purchased from Sigma Aldrich and incubated over 4Å MS for 48 h prior to use. Toluene and CH2Cl2 were dried over CaH2 whereas THF and Et2O were dried over sodium (dryness was monitored by colorization of benzophenone ketyl radical); they were freshly distilled prior to use. NMR spectra were obtained using a 500 MHz and 600 MHz Varian NMR spectrometers and referenced using the residual 1H peak from the deuterated solvent. Waters 2795 (Alliance HT) instrument was used for HRMS (ESI) analysis with polyethylene glycol (PEG-400-600) as a reference. Column chromatography was performed using Silicycle 60Å, 35-75 µm silica gel. Pre- coated 0.25 mm thick silica gel 60 F254 plates were used for analytical TLC and visualized using UV light, iodine, potassium permanganate stain, p-anisaldehyde stain or phosphomolybdic acid in EtOH stain. Halofunctionalization reactions were performed in the absence of light. N- chlorosuccinimide (NCS), N-bromosuccinimide (NBS), N-iodosuccinimide (NIS), 1,3- dichloro-5,5-dimethylhydantoin (DCDMH), and 1,3-dibromo-5,5-dimethylhydantoin (DBDMH) were re-crystallized prior to use. All other commercially available reagents and solvents were used as received unless otherwise mentioned. 308 IV-6-2 General procedure for bromo spiroketalization of THP-protected keto- alkenols R2 R1 O OTHP DCDMH, DBDMH (1 equiv. each) Toluene, rt, 3-4 h EtOH (10.0 equiv) O R2 O R1 Br To the THP-protected keto-alkenol (0.1 mmol) in dry toluene (1 mL) was added dry ethanol (10.0 mmol, 10.0 equiv) and a mixture of 1,3-dibromo-5,5-dimethylhydantoin, DBDMH (1.0 mmol, 1.0 equiv) and 1,3-dichloro-5,5-dimethylhydantoin, DCDMH (1.0 mmol, 1.0 equiv). The reaction mixture was stirred at room temperature and followed by TLC. The initial colorless solution changed to orange, and become clear along with precipitation of hydantoin. This clear solution was the indication for the completion of the reaction, which took 3-4 h. The reaction mixture was diluted with hexanes (2 mL) followed by addition of sodium sulfite (2 ml of a 10% aqueous solution). The organic layer was separated and the aqueous layer was extracted with hexanes (3 x 2 mL). Combined organic extracts were concentrated under reduced pressure at room temperature to give the crude product. Crude 1H NMR was used to determine the diastereoselectivity of the reaction. The crude product was then purified by column chromatography (silica, ethyl acetate/hexanes = 3%) leading to the pure bromo spiroketal products (65 – 99% average yield). O O Br IV-45 309 Spiroketalization reaction to produce IV-45 was performed on a 1.0 mmol of ketone IV- 116, leading to the IV-45 in 99% yield (307 mg). Reaction with 0.1 mmol scale yielded a similar result. 1H NMR (500 MHz, CDCl3) δ 7.44 – 7.39 (m, 2H), 7.38 – 7.30 (m, 3H), 4.66 (d, J = 10.4 Hz, 1H), 4.00 (ddd, J = 12.0, 10.3, 4.5 Hz, 1H), 3.72 – 3.60 (m, 2H), 2.50 (tdd, J = 12.9, 12.0, 5.0 Hz, 1H), 2.26 (dtd, J = 13.0, 4.5, 2.9 Hz, 1H), 1.82 – 1.63 (m, 4H), 1.63 – 1.54 (m, 1H), 1.52 – 1.41 (m, 3H). 13C NMR (125 MHz, CDCl3) δ 139.66, 128.31, 128.07, 127.97, 95.98, 76.33, 60.79, 52.39, 37.64, 34.88, 31.25, 25.01, 18.34. TOF MS ES+ (C15H20BrO2): Calc. [M + H]+: 311.0647, Found [M + H]+: 311.0645. O O Br IV-53 Spiroketalization of ketone IV-132 (35 mg, 0.1 mmol) produced IV-53 (33.5 mg, 93% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.89 – 7.79 (m, 4H), 7.54 (dd, J = 8.4, 1.7 Hz, 1H), 7.51 – 7.42 (m, 2H), 4.83 (d, J = 10.3 Hz, 1H), 4.12 (ddd, J = 12.1, 10.3, 4.5 Hz, 1H), 3.75 – 3.64 (m, 2H), 2.55 (qd, J = 12.5, 5.5 Hz, 1H), 2.30 (dtd, J = 13.0, 4.4, 3.0 Hz, 1H), 1.87 – 1.78 (m, 2H), 1.75 – 1.66 (m, 2H), 1.62 – 1.55 (m, 1H), 1.53 – 1.42 (m, 3H). 310 13C NMR (125 MHz, CDCl3) δ 137.04, 133.40, 132.98, 128.15, 127.89, 127.71, 127.55, 126.04, 125.38, 96.08, 76.56, 60.88, 52.27, 37.70, 34.91, 31.30, 25.02, 18.37. O O Br IV-54 Spiroketalization of ketone IV-131 (36 mg, 0.1 mmol) produced IV-54 (32.8 mg, 88% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 8.37 (d, J = 8.6 Hz, 1H), 7.85 (dd, J = 16.0, 8.1 Hz, 2H), 7.67 – 7.56 (m, 1H), 7.49 (dq, J = 15.4, 7.4 Hz, 3H), 5.49 (br, 1H), 4.36 (br, 1H), 3.82 – 3.54 (m, 2H), 2.62 (qd, J = 12.1, 7.5 Hz, 1H), 2.34 (dq, J = 12.6, 4.0 Hz, 1H), 1.92 – 1.83 (m, 2H), 1.75 – 1.61 (m, 2H), 1.54 (pd, J = 12.3, 5.8 Hz, 2H), 1.43 (dd, J = 26.1, 9.7 Hz, 2H). 13C NMR (125 MHz, CDCl3) δ 139.15, 133.76, 131.84, 128.87, 128.84, 125.83, 125.38, 125.21, 96.18, 60.94, 52.47, 37.77, 35.01, 31.56, 24.98, 18.33. O O Br IV-55 CH3 Spiroketalization of ketone IV-117 (33 mg, 0.1 mmol) produced IV-55 (28.2 mg, 87%yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.30 (d, J = 8.0, 2H), 7.16 (d, J = 8.0 Hz, 2H),, 4.63 (d, J = 10.3 Hz, 1H), 4.00 (ddd, J = 12.0, 10.3, 4.5 Hz, 1H), 3.72 – 3.59 (m, 2H), 2.56 – 2.43 (m, 311 1H), 2.35 (s, 3H), 2.26 (dtd, J = 12.9, 4.4, 2.9 Hz, 1H), 1.81 – 1.62 (m, 4H), 1.60 – 1.41 (m, 4H). 13C NMR (125 MHz, CDCl3) δ 138.0, 136.7, 128.8, 127.8, 95.9, 76.1, 60.74, 52.5, 37.6, 34.8, 31.2, 25.0, 21.2, 18.3 ppm. O H3C O Br IV-56 Spiroketalization of ketone IV-118 (49 mg, 0.15 mmol) produced IV-56 (39.0 mg, 80% yield) as a white foam. mp = 47 – 50° 1H NMR (500 MHz, CDCl3) δ 7.46 – 7.39 (m, 1H), 7.32 – 7.15 (m, 3H), 5.06 (d, J = 10.4 Hz, 1H), 4.11 (ddd, J = 12.1, 10.4, 4.4 Hz, 1H), 3.76 – 3.63 (m, 2H), 2.59 – 2.52 (m, 1H), 2.51 (s, 3H), 2.29 (dtd, J = 13.0, 4.3, 2.8 Hz, 1H), 1.87 – 1.65 (m, 4H), 1.62 – 1.42 (m, 4H). 13C NMR (125 MHz, CDCl3) δ 13C NMR (126 MHz, cdcl3) δ 138.36, 136.46, 130.14, 127.96, 127.06, 126.11, 95.96, 71.79, 60.72, 53.10, 37.76, 34.95, 31.36, 25.03, 20.22, 18.33. O O Br IV-57 OCH3 Spiroketalization of ketone IV-121 (34 mg, 0.1 mmol) produced IV-57 (25.0 mg, 73% yield) as a white foam. 312 1H NMR (500 MHz, CDCl3) δ 7.33 (d, J = 8.7 Hz, 2H), 6.88 (d, J = 8.6 Hz, 2H), 4.61 (d, J = 10.4 Hz, 1H), 3.99 (ddd, J = 12.0, 10.3, 4.5 Hz, 1H), 3.80 (s, 3H), 3.71 – 3.60 (m, 2H), 2.49 (tdd, J = 12.9, 12.0, 4.9 Hz, 1H), 2.25 (dtd, J = 12.9, 4.4, 2.9 Hz, 1H), 1.80 – 1.62 (m, 4H), 1.60 – 1.42 (m, 4H). 13C NMR (125 MHz, CDCl3) δ 159.44, 131.98, 129.03, 113.44, 95.98, 75.82, 60.76, 55.20, 52.77, 37.64, 34.87, 31.27, 25.01, 18.34, 14.13. TOF MS ES+ (C16H22BrO3): Calc. [M + H]+: 341.0752, Found [M + H]+: 341.0741. O O Br IV-58 F Spiroketalization of ketone IV-119 (49 mg, 0.15 mmol) produced IV-58 (41.0 mg, 84% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.43 – 7.34 (m, 2H), 7.09 – 6.99 (m, 2H), 4.64 (d, J = 10.3 Hz, 1H), 3.93 (ddd, J = 12.1, 10.3, 4.5 Hz, 1H), 3.68 (ddt, J = 11.0, 5.0, 1.8 Hz, 1H), 3.64 – 3.57 (m, 1H), 2.49 (tdd, J = 13.1, 12.0, 4.8 Hz, 1H), 2.25 (dtd, J = 13.0, 4.4, 2.8 Hz, 1H), 1.83 – 1.62 (m, 4H), 1.60 – 1.43 (m, 4H). 13C NMR (125 MHz, CDCl3) δ 162.55 (d, JC-F = 246.1 Hz), 135.57 (d, JC-F = 3.3 Hz), 129.55 (d, JC-F = 8.1 Hz), 114.94 (d, JC-F = 21.4 Hz), 96.04, 75.63, 60.81, 52.54, 37.59, 34.82, 31.17, 24.96, 18.33. 19F NMR (470 MHz, CDCl3) δ -113.86 – -113.95 (m). TOF MS ES+ (C15H19BrFO2): Calc. [M + H]+: 329.0552, Found [M + H]+: 329.0544. 313 O O Br IV-59 Spiroketalization of ketone IV-120 (39 mg, 0.1 mmol) produced IV-59 (32.0 mg, 83% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.61 – 7.56 (m, 4H), 7.50 – 7.46 (m, 2H), 7.42 (dd, J = 9.3, 5.9 Hz, 2H), 7.34 – 7.31 (m, 1H), 4.71 (d, J = 10.4 Hz, 1H), 4.03 (ddd, J = 10.5, 8.0, 5.1 Hz, 1H), 3.74 – 3.63 (m, 2H), 2.52 (qd, J = 12.7, 5.2 Hz, 1H), 2.32 – 2.26 (m, 1H), 1.83 – 1.71 (m, 3H), 1.71 – 1.65 (m, 1H), 1.61 – 1.43 (m, 4H). 13C NMR (125 MHz, CDCl3) δ 140.86, 138.67, 128.76, 128.70, 128.36, 127.26, 127.16, 126.86, 96.03, 76.06, 60.83, 52.34, 37.66, 34.88, 31.25, 25.01, 18.37. O O Br IV-62 Spiroketalization of ketone IV-136 (30 mg, 0.1 mmol) produced IV-63 (24.3 mg, 82% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.40 – 7.28 (m, 5H), 4.82 (d, J = 10.4 Hz, 1H), 4.01 (ddd, J = 11.9, 10.4, 4.6 Hz, 1H), 3.95 (td, J = 8.1, 4.9 Hz, 1H), 3.89 (td, J = 8.1, 6.8 Hz, 1H), 2.54 – 2.45 (m, 1H), 2.38 (dtd, J = 12.7, 4.6, 2.9 Hz, 1H), 2.07 (td, J = 13.6, 4.6 Hz, 1H), 2.03 – 1.92 (m, 2H), 1.88 – 1.78 (m, 2H), 1.76 – 1.67 (m, 1H). 13C NMR (125 MHz, CDCl3) δ 139.71, 128.31, 128.12, 127.91, 106.14, 77.48, 67.36, 52.06, 37.18, 35.20, 32.71, 23.63. 314 TOF MS ES+ (C14H18BrO2): Calc. [M + H]+: 297.0490, Found [M + H]+: 297.0482. O Ph Br IV-46 O Analytical data for bromo macrocyclic product IV-46: 1H NMR (500 MHz, CDCl3) δ 7.32 – 7.24 (m, 5H), 4.04 – 3.99 (m 2H), 3.11 – 3.10 (m, 1H), 2.80 (t, J = 10.0, 1H), 2.76 – 2.67 (m, 3H), 2.51 – 2.49 (m, 1H), 2.44 – 2.39 (m, 1H), 2.50 – 2.20 (m, 1H), 2.12 –2.04 (m, 1H), 1. 61 – 1.58 (m, 1H), 1.51 – 1.38 (m, 2H). 13C NMR (125 MHz, CDCl3) δ 208.15, 139.78, 128.30, 128.15, 127.80, 85.39, 69.49, 55.65, 42.03, 41.97, 32.87, 29.31, 22.07. IV-6-3 General procedure for dibromo spiroketalization of THP-protected keto- alkenols O R OTHP NBS (2.5 equiv) cat. HCl CHCl3, rt O Br O R Br To the THP-protected keto-alkenols (0.1 mmol) dissolved in chloroform (amylene stabilized, 1 mL) was added N-bromosuccinimide (44.5 mg, 0.25 mmol, 2.5 equiv) at room temperature. To this mixture was then added concentrated HCl (~10 µL). The reaction was continued to stir at room temperature (protected from light) and was monitored by TLC. After completion of the reaction (~4 h), sodium sulfite (2 mL of 10% aqueous solution) was added to the reaction. The organic layer was separated and the aqueous 315 layer was extracted with hexanes (3 x 2 mL). The combined organic extracts were dried over sodium sulfate and concentrated under reduced pressure at room temperature to give the crude product. Crude 1H NMR was used to determine the diastereoselctivity of the reaction. The crude product was then purified by column chromatography (silica, ethyl acetate/hexanes = 3%) leading to the pure dibromo-spiroketal products. O Br O Br IV-64 Dibromo spiroketalization of ketone IV-116 (31 mg, 0.1 mmol) produced IV-64 (337 mg product was isolated, 87% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.49 – 7.42 (m, 2H), 7.39 – 7.33 (m, 3H), 4.68 (d, J = 10.5 Hz, 1H), 4.46 (td, J = 11.2, 4.3 Hz, 1H), 4.03 (t, J = 3.2 Hz, 1H), 3.76 (dd, J = 11.7, 3.9 Hz, 1H), 3.65 (td, J = 11.5, 11.1, 2.6 Hz, 1H), 3.11 (ddd, J = 14.2, 11.9, 3.4 Hz, 1H), 2.60 (ddd, J = 14.0, 4.2, 2.9 Hz, 1H), 2.20 (d, J = 13.7 Hz, 1H), 1.65 (td, J = 8.9, 4.5 Hz, 1H), 1.55 (s, 1H), 1.52 – 1.48 (m, 1H), 1.48 – 1.40 (m, 1H), 1.32 (td, J = 13.4, 4.5 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 128.60, 128.18, 128.04, 127.88, 96.29, 88.84, 61.97, 55.17, 48.12, 39.55, 33.86, 24.42, 18.61. O Br O Br IV-65 316 Dibromo spiroketalization of ketone IV-136 (30 mg, 0.1 mmol) produced IV-65 (27.3 mg, 73% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.47 – 7.40 (m, 2H), 7.40 – 7.30 (m, 3H), 4.86 (d, J = 10.5 Hz, 1H), 4.45 (ddd, J = 11.8, 10.5, 4.4 Hz, 1H), 4.12 (t, J = 3.1 Hz, 1H), 4.10 – 4.00 (m, 2H), 3.00 (ddd, J = 14.3, 11.9, 3.3 Hz, 1H), 2.69 (ddd, J = 14.2, 4.4, 2.9 Hz, 1H), 2.19 (ddd, J = 12.2, 7.5, 2.8 Hz, 1H), 2.04 – 1.82 (m, 3H). 13C NMR (125 MHz, CDCl3) δ 128.58, 128.21, 128.00, 127.42, 101.53, 77.68, 69.83, 53.24, 47.88, 40.54, 38.36, 23.88. O Br O Br IV-66 Dibromo spiroketalization of ketone IV-132 (35 mg, 0.1 mmol) produced IV-66 (38.5 mg, 88% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.89 – 7.82 (m, 4H), 7.61 (dd, J = 8.5, 1.9 Hz, 1H), 7.51 – 7.46 (m, 2H), 4.86 (d, J = 10.5 Hz, 1H), 4.65 – 4.54 (m, 1H), 4.07 (t, J = 3.2 Hz, 1H), 3.80 (ddd, J = 10.9, 4.6, 2.2 Hz, 1H), 3.70 (td, J = 11.7, 2.6 Hz, 1H), 3.16 (ddd, J = 14.8, 12.0, 3.4 Hz, 1H), 2.65 (ddd, J = 14.1, 4.4, 2.8 Hz, 1H), 2.24 (dt, J = 13.7, 3.3 Hz, 1H), 1.71 – 1.50 (m, 2H), 1.47 – 1.40 (m, 1H), 1.35 (td, J = 13.4, 4.5 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 136.19, 133.51, 132.89, 128.15, 128.10, 127.73, 127.68, 126.19, 126.12, 125.26, 96.36, 76.95, 62.04, 55.20, 47.96, 39.58, 33.86, 24.42, 18.63. 317 O Br O Br IV-67 Dibromo spiroketalization of ketone IV-131 (35 mg, 0.1 mmol) produced IV-67 (26.7 mg, 61% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.93 – 7.80 (m, 2H), 7.53 (m, 3H), 5.80 – 5.20 (br,1H), 5.10 – 4.60 (br,1H), 4.14 (t, J = 3.2 Hz, 1H), 3.84 (d, J = 11.0 Hz, 1H), 3.72 (ddd, J = 13.6, 11.2, 2.6 Hz, 1H), 3.25 (ddd, J = 14.8, 11.7, 3.4 Hz, 1H), 2.71 (ddd, J = 14.1, 4.1, 2.9 Hz, 1H), 2.27 (d, J = 13.8 Hz, 1H), 1.61 – 1.48 (m, 4H), 1.39 (td, J = 13.2, 4.5 Hz, 2H). 13C NMR (125 MHz, CDCl3) δ 134.75, 133.27, 128.85, 128.59, 128.37, 125.99, 125.94, 125.50, 125.20, 96.64, 82.47, 62.09, 55.36, 49.49, 40.00, 33.93, 24.39, 18.60. O O Br Br IV-68 CH3 Dibromo spiroketalization of ketone IV-117 (33 mg, 0.1 mmol) produced IV-68 (30.0 mg, 74% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.34 (d, J = 7.8 Hz, 2H), 7.18 (d, J = 7.6 Hz, 2H), 4.65 (d, J = 10.5 Hz, 1H), 4.46 (td, J = 11.3, 4.3 Hz, 1H), 4.03 (t, J = 3.1 Hz, 1H), 3.75 (dd, J = 11.4, 4.4 Hz, 1H), 3.65 (td, J = 11.6, 2.5 Hz, 1H), 3.10 (ddd, J = 14.8, 12.0, 3.4 Hz, 1H), 2.60 318 (dt, J = 14.1, 3.5 Hz, 1H), 2.35 (s, 3H), 2.22 – 2.15 (m, 1H), 1.65 (tt, J = 13.9, 4.1 Hz, 2H), 1.48 (dt, J = 12.4, 4.2 Hz, 1H), 1.41 (m, 1H), 1.31 (td, J = 13.4, 4.5 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 138.39, 135.95, 128.92, 127.90, 96.29, 76.49, 61.93, 55.23, 48.26, 39.58, 33.86, 24.43, 21.31, 18.60. ppm. O H3C O Br IV-69 Br Dibromo spiroketalization of ketone IV-118 (33 mg, 0.1 mmoll) produced IV-69 (34.3 mg, 85% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.50 (d, J = 7.3 Hz, 1H), 7.27 – 7.20 (m, 2H), 7.16 (d, J = 7.4 Hz, 1H), 5.05 (d, J = 10.6 Hz, 1H), 4.57 (td, J = 11.5, 4.1 Hz, 1H), 4.04 (d, J = 3.3 Hz, 1H), 3.81 – 3.73 (m, 1H), 3.66 (td, J = 11.6, 2.5 Hz, 1H), 3.12 (ddd, J = 14.9, 12.0, 3.4 Hz, 1H), 2.61 (dt, J = 14.2, 3.6 Hz, 1H), 2.48 (s, 3H), 2.18 (dd, J = 13.6, 4.0 Hz, 1H), 1.72 – 1.57 (m, 2H), 1.41 (d, J = 13.1 Hz, 1H), 1.32 (td, J = 13.5, 4.5 Hz, 2H). 13C NMR (125 MHz, CDCl3) δ 136.47, 136.36, 130.16, 128.31, 128.25, 126.26, 96.31, 77.20, 61.88, 55.34, 51.75, 39.65, 33.91, 24.43, 20.14, 18.59. O Br O Br IV-70 F Dibromo spiroketalization of ketone IV-119 (33 mg, 0.1 mmol) produced IV-70 (26.5 mg, 65% yield) as a white foam. 319 1H NMR (500 MHz, CDCl3) δ 7.43 (dd, J = 8.6, 5.5 Hz, 2H), 7.08 – 7.01 (m, 2H), 4.66 (d, J = 10.5 Hz, 1H), 4.39 (td, J = 11.3, 4.4 Hz, 1H), 4.02 (t, J = 3.2 Hz, 1H), 3.76 (dd, J = 11.1, 4.5 Hz, 1H), 3.62 (td, J = 11.7, 11.1, 2.7 Hz, 1H), 3.09 (ddd, J = 14.1, 11.9, 3.4 Hz, 1H), 2.59 (ddd, J = 14.1, 4.3, 2.9 Hz, 1H), 2.19 (d, J = 13.7 Hz, 1H), 1.64 (tdt, J = 13.1, 8.3, 4.0 Hz, 1H), 1.49 (dt, J = 12.6, 4.3 Hz, 1H), 1.46 – 1.55 (m, 2H), 1.32 (td, J = 13.3, 4.7 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 129.69 (d, JC-F = 8.4 Hz), 115.10 (d, JC-F = 21.5 Hz), 96.36, 90.89, 62.03, 55.07, 48.27, 39.48, 33.82, 29.71, 22.70, 18.62. O Br O Br IV-71 Dibromo spiroketalization of ketone IV-120 (39 mg, 0.1 mmol) produced IV-71 (23.0 mg, 59% yield) as a white foam. 1H NMR (500 MHz, CDCl3) δ 7.59 (d, J = 8.0 Hz, 4H), 7.53 (d, J = 7.9 Hz, 2H), 7.43 (t, J = 7.6 Hz, 2H), 7.33 (t, J = 7.3 Hz, 1H), 4.74 (d, J = 10.5 Hz, 1H), 4.50 (td, J = 11.1, 4.1 Hz, 1H), 4.05 (d, J = 3.0 Hz, 1H), 3.82 – 3.74 (m, 1H), 3.68 (t, J = 11.4 Hz, 1H), 3.13 (td, J = 12.9, 11.7, 3.3 Hz, 1H), 2.63 (dt, J = 14.3, 3.5 Hz, 1H), 2.22 (d, J = 13.7 Hz, 1H), 1.69 (m, 1H), 1.59 – 1.54 (m, 1H), 1.49 (s, 1H), 1.43 (m, 1H), 1.34 (td, J = 13.4, 4.5 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 128.74, 128.44, 127.34, 127.18, 126.98, 96.35, 76.45, 62.02, 55.17, 48.08, 39.56, 33.87, 24.43, 18.65. 320 IV-6-4 Synthesis of ketones IV-116 to IV-121 These substrates were synthesized according to the following general scheme: R O Ph3P LiHMDS, THF OH R I R 1. tBuLi, Et2O –100 °C O 2. Me N OMe R OTHP OH PPh3, I2 Imidazole DCM O OTHP Synthesis of THP-protected Weinreb amide IV-103: O O IV-101 O H N •HCl Et3Al DCM OMe N O Me IV-102 OH THP PPTS DCM Me OMe N O IV-103 OTHP Following a reported procedure,42 to a suspention of N,O-dimethyl hydroxyl amine hydrochloride (6.0 g, 61.5 mmol, 3.0 equiv) in dry DCM (60 mL) was added diethyl aluminum (1.0 M in heptane, 60.0 mL, 60.0 mmol) dropwise at –10 °C. After addition was completed, the mixture was stirred at room temperature for 30 min. To this mixture at 0 °C was added a solution of d-lactone (2.0 g, 20.0 mmol, 1.0 equiv) in DCM (10 mL). The reaction was further stirred at room temperature for 20 h, before it was quenched by careful addition sodium bicarbonate (1.0 M, 100 mL). The aqueous layer was extracted with DCM (5 x 50 mL), and the combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced pressure to give the crude alcohol IV-102, which was used in the next step without further purification. 321 To the crude alcohol (ca. 2.6 g) in anhydrous DCM (100 mL) were added PPTS (100 mg) and DHP (2.0 mL, 22.0 mmol, 1.1 equiv) at 0 °C. The reaction was continued to stirred at room temperature for 12 h. The volatiles were removed under reduced pressure and the crude product was purified on column chromatography (silica, ethyl acetate/hexanes 30%) to yield the pure THP-protected Weinreb amide IV-103 (2.6 g, 53% yield) as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 4.55 (dd, J = 4.4, 2.8 Hz, 1H), 3.87 – 3.79 (m, 1H), 3.74 (dt, J = 9.6, 6.5 Hz, 1H), 3.65 (s, 3H), 3.51 – 3.42 (m, 1H), 3.38 (dt, J = 9.7, 6.3 Hz, 1H), 3.15 (s, 3H), 2.43 (t, J = 7.5 Hz, 2H), 1.90 – 1.74 (m, 2H), 1.73 – 1.58 (m, 4H), 1.57 – 1.44 (m, 4H). 13C NMR (126 MHz, CDCl3) δ 98.77, 69.43, 67.19, 62.23, 61.17, 30.70, 29.41, 25.45, 22.31, 21.44, 19.56, 19.11. TOF MS ES+ (C12H23NNaO4): Calc. [M + Na]+: 268.1525, Found [M + H]+: 268.1548. Synthesis of but-3-en-1-ol derivatives R O Ph3P OH LiHMDS, THF R OH Following the reported procedure,43 lithium bis(trimethylsilyl)amide (1.0 M in THF, 14.0 mL, 14.0 mmol) was added at –20 °C (3-propan-1-ol) triphenylphosphonium bromide (2.4 g, 6.0 mmol, 1.2 equiv) in dry THF (14.0 mL). The solution was stirred at this temperature for 1 h. To this solution at –20 °C was added aldehyde (5.0 mmol, 1.0 equiv). The reaction was stirred at this temperature for another to a suspension 322 2 h, then allowed to warm up to room temperature and was stirred for 12 h. Saturated ammonium chloride (20 mL) was added to the reaction mixture, the organic layer was separated and the aqueous layer was extracted with ethyl acetate (3 x 20 mL). The combined organic extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure to yield the crude alcohol that was purified via column chromatography (silica, ethyl acetate/hexanes 10 to 20%) to afford the corresponding pure alcohol. OH IV-104 Following the above procedure, alcohol IV-104 (350 mg, 47% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.37 – 7.32 (m, 2H), 7.31 – 7.26 (m, 2H), 7.23 – 7.16 (m, 1H), 6.49 (dt, J = 15.9, 1.5 Hz, 1H), 6.19 (dt, J = 15.8, 7.1 Hz, 1H), 3.74 (t, J = 6.3 Hz, 2H), 2.48 (dtd, J = 7.6, 6.3, 1.4 Hz, 2H). 13C NMR (126 MHz, CDCl3) δ: 137.1, 132.7, 128.5, 127.2, 126.1, 126.0, 62.0, 36.3 ppm. OH IV-105 Following the above procedure, alcohol IV-105 (460 mg, 57% yield) was synthesized as a white solid. mp = 49 – 51° 323 1H NMR (500 MHz, Chloroform-d) δ: 7.24 (d, J = 7.9 Hz, 2H), 7.09 (d, J = 7.9 Hz, 2H), 6.45 (d, J = 15.9 Hz, 1H), 6.13 (dt, J = 15.9, 7.2 Hz, 1H), 3.73 (t, J = 6.5 Hz, 2H), 2.46 (td, J = 7.2, 6.5 Hz, 2H), 2.31 (s, 3H), 1.47 (s, 1H) ppm. 13C NMR (126 MHz, CDCl3) δ: 137.03, 134.38, 132.73, 129.20, 125.94, 125.15, 62.03, 36.40, 21.15 ppm. OH IV-106 Following the above procedure, alcohol IV-106 (380 mg, 47% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ: 7.42 (m, 1H), 7.16 – 7.11 (m, 3H), 6.69 (d, J = 15.5 Hz, 1H), 6.06 (dt, J = 15.5, 7.5 Hz, 1H), 3.75 (t, J = 6.3 Hz, 2H), 2.50 (dt, J = 7.5, 6.3 Hz, 2H), 2.33 (s, 3H), 1.51 (s, 1H). 13C NMR (126 MHz, CDCl3) δ 136.31, 135.02, 130.72, 130.24, 127.59, 127.21, 126.05, 125.43, 62.06, 36.71, 19.84 ppm. F IV-107 OH Following the above procedure, alcohol IV-107 (420 mg, 50% yield) was synthesized as a colorless oil. 324 1H NMR (500 MHz, Chloroform-d) δ 7.33 – 7.25 (m, 2H), 7.01 – 6.92 (m, 2H), 6.44 (d, J = 15.7 Hz, 1H), 6.10 (dt, J = 15.8, 7.2 Hz, 1H), 3.74 (t, J = 6.3 Hz, 2H), 2.49 – 2.39 (m, 2H). 13C NMR (126 MHz, cdcl3) δ 162.08 (d, JC-F = 246.1 Hz), 133.37 (d, JC-F = 3.3 Hz), 131.57, 127.50 (d, JC-F = 8.0 Hz), 126.09 (d, JC-F = 2.3 Hz), 61.99, 36.32. Ph IV-108 OH Following the above procedure, alcohol IV-108 (600 mg, 54% yield) was synthesized as a white solid. mp = 132–133° 1H NMR (500 MHz, Chloroform-d) δ 7.60 – 7.56 (d, J = 8.0 Hz, 2H), 7.53 (d, J = 8.0 Hz, 2H), 7.43 – 7.40 (m, 4H), 7.34 – 7.30 (m, 1H), 6.53 (d, J = 15.9 Hz, 1H), 6.28 – 6.20 (dt, J = 15.9, 7.5 Hz, 1H), 3.76 (q, J = 6.0 Hz, 2H), 2.53 – 2.48 (m, 2H), 1.43 (t, J = 5.7 Hz, 1H). 13C NMR (126 MHz, cdcl3) δ 140.70, 140.01, 136.24, 132.39, 128.76, 127.24, 127.22, 126.89, 126.48, 62.03, 36.49. O IV-109 OH Following the above procedure, alcohol IV-109 (350 mg, 40% yield) was synthesized as a white solid. 325 mp = 73–75° 1H NMR (500 MHz, Chloroform-d) δ 7.27 (d, J = 6.5 Hz, 2H), 6.82 (d, J = 6.5 Hz, 2H), 6.43 (d, J = 16.0 Hz, 1H), 6.04 (dt, J = 16.0, 7.2 Hz, 1H), 3.79 (s, 3H), 3.72 (t, J = 6.5 Hz, 2H), 2.45 (td, J = 7.2, 6.5 Hz, 2H), 1.44 (s, 1H). 13C NMR (126 MHz, cdcl3) δ 158.94, 132.30, 130.01, 127.18, 123.96, 113.92, 62.08, 55.28, 36.40. Synthesis of homoallylic iodide R OH PPh3, I2 Imidazole DCM R I Homoallylic alcohols (~2.5 mmol) from above reactions were dissolved in DCM (~25 mL). To these solutions at room temperature were added triphenyl phosphine (1.2 equiv), iodine (1.2 equiv) and imidazole (1.2 equiv). The reaction mixtures were stirred at room temperature and followed by TLC. The reactions were completed within 1 h, after which water (25 mL) was added to quench the reaction. The organic layer was separated and the aqueous layer was extracted with DCM (3 x 20 mL). The combined organic extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure to yield the crude alcohol that was purified via column chromatography (silica, ethyl acetate/hexanes 5%) to afford the corresponding pure products. I IV-110 326 Following the above procedure, homoallylic iodide IV-110 (600 mg, 83% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.39 – 7.33 (m, 2H), 7.29 (dd, J = 8.5, 6.8 Hz, 2H), 7.24 – 7.19 (m, 1H), 6.45 (dt, J = 15.8, 1.5 Hz, 1H), 6.13 (dt, J = 15.8, 7.0 Hz, 1H), 3.23 (t, J = 7.3 Hz, 2H), 2.77 (qd, J = 7.2, 1.4 Hz, 2H). 13C NMR (126 MHz, cdcl3) δ 137.03, 132.26, 130.94, 128.57, 126.99, 126.17, 36.99, 5.06. I IV-111 Following the above procedure, homoallylic iodide IV-111 (665 mg, 88% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ: 7.25 (d, J = 8.5 Hz, 2H), 7.10 (d, J = 8.5 Hz, 2H), 6.42 (d, J = 15.8, 1.5 Hz, 1H), 6.08 (dt, J = 15.8, 7.0 Hz, 1H), 3.22 (t, J = 7.3 Hz, 2H), 2.78 – 2.73 (dtd, J = 7.3, 7.0, 1.5 Hz, 2H), 2.32 (s, 3H) ppm. 13C NMR (126 MHz, cdcl3) δ: 137.21, 134.22, 132.08, 129.25, 127.42, 126.04, 37.04, 21.18, 5.23 ppm. I IV-112 Following the above procedure, homoallylic iodide IV-112 (500 mg, 85% yield) was synthesized as a colorless oil. 327 1H NMR (500 MHz, Chloroform-d) δ 7.42 – 7.39 (m, 1H), 7.14 (m, 3H), 6.66 (d, J = 15.6 Hz, 1H), 5.99 (dt, J = 15.6, 7.0 Hz, 1H), 3.25 (t, J = 7.2 Hz, 2H), 2.78 (dt, J = 7.2, 7.0 Hz, 2H), 2.33 (s, 3H) ppm. 13C NMR (126 MHz, cdcl3) δ 136.26, 135.21, 130.30, 130.23, 129.80, 127.35, 126.09, 125.60, 37.18, 19.87, 5.40 ppm. F IV-113 I Following the above procedure, homoallylic iodide IV-113 (610 mg, 93% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.35 – 7.27 (m, 2H), 7.04 – 6.95 (m, 2H), 6.41 (dt, J = 15.8, 1.4 Hz, 1H), 6.04 (dt, J = 15.8, 6.9 Hz, 1H), 3.22 (t, J = 7.2 Hz, 2H), 2.75 (qd, J = 7.1, 1.4 Hz, 2H). 13C NMR (126 MHz, Chloroform-d) δ 162.20 (d, JC-F = 246.5 Hz), 133.19 (d, JC-F = 3.3 Hz), 131.10, 128.23 (d, JC-F = 2.3 Hz), 127.65 (d, JC-F = 8.1 Hz), 115.47 (d, JC-F = 21.7 Hz), 36.87, 5.06. Ph IV-114 I Following the above procedure, homoallylic iodide IV-114 (850 mg, 98% yield) was synthesized as an off white solid. mp = 93–95° 328 1H NMR (500 MHz, Chloroform-d) δ: 7.59 – 7.52 (m, 4H), 7.43 – 7.40 (m, 4H), 7.35 – 7.29 (m, 1H), 6.49 (d, J = 15.8 Hz, 1H), 6.18 (dt, J = 15.8, 7.0 Hz, 1H), 3.24 (t, J = 7.2 Hz, 2H), 2.79 (dt, J = 7.2, 7.0 Hz, 2H). 13C NMR (126 MHz, cdcl3) δ: 140.70, 140.20, 136.06, 131.82, 128.77, 128.62, 127.28, 126.90, 126.59, 37.04, 5.06 ppm. O IV-115 I Following the above procedure, homoallylic iodide IV-115 (500 mg, 94% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.28 (m, 2H), 6.89 – 6.78 (m, 2H), 6.39 (d, J = 15.8 Hz, 1H), 5.99 (dtd, J = 15.7, 7.0, 0.8 Hz, 1H), 3.79 (s, 3H), 3.21 (td, J = 7.3, 0.8 Hz, 2H), 2.74 (qt, J = 7.2, 1.0 Hz, 2H). 13C NMR (126 MHz, cdcl3) δ 159.07, 131.65, 129.84, 127.31, 126.29, 113.98, 55.31, 37.08, 5.46. 329 Synthesis of ketones IV-116 to IV-121 I IV-110 –100 °C OMe N 2. Me 1. tBuLi, Et2O O IV-103 OTHP O IV-116 OTHP General procedure for synthesis of ketones from the corresponding homoallylic iodide: To a flame dried flask containing diethyl ether (10.0 mL) under argon at –100 °C (liquid nitrogen in methanol bath) was added tBuLi (1.7 M in pentane, 3.0 mL, 5.25 mmol, 2.5 equiv) dropwise. To this solution was added alkyl iodide IV-110 (560 mg, 2.1 mmol, 1.0 equiv) in diethyl ether (3.0 mL) dropwise at the same temperature. The reaction stirred at this temperature for 30 min. To this solution at –78 °C was added a solution of the Weinreb amide IV-103 (514 mg, 2.1 mmol) in diethyl ether (3 mL). The reaction was slowly warmed up to room temperature over 2 h. TLC showed completion of the reaction. Saturated ammonium chloride (10 mL) was added to this solution, the organic layer was separated and the aqueous layer was extracted with ethyl acetate (3 x 10 mL). The combined organic extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure to yield the crude alcohol that was purified via column chromatography (silica, ethyl acetate/hexanes 10 – 20%) to afford the corresponding pure IV-116 as a clear oil (510 mg, 82%). 1H NMR (500 MHz, Chloroform-d) δ 7.31 – 7.28 (m, 2H), 7.28 – 7.24 (m, 2H), 7.19 – 7.13 (m, 1H), 6.39 – 6.35 (m, 1H), 6.17 (dt, J = 15.8, 6.8 Hz, 1H), 4.51 (dd, J = 4.7, 2.7 Hz, 1H), 3.80 (dddd, J = 11.1, 7.7, 4.3, 2.9 Hz, 1H), 3.71 (dt, J = 9.7, 6.4 Hz, 1H), 3.49 – 3.43 330 (m, 1H), 3.38 (dt, J = 9.7, 6.1 Hz, 1H), 2.62 – 2.56 (m, 2H), 2.55 – 2.50 (m, 2H), 2.49 – 2.42 (m, 2H), 1.89 – 1.83 (m, 2H), 1.80 – 1.70 (m, 2H), 1.57 – 1.43 (m, 5H). 13C NMR (126 MHz, cdcl3) δ 209.79, 137.41, 130.66, 128.99, 128.48, 127.05, 125.98, 98.90, 66.57, 62.45, 42.23, 39.72, 30.69, 27.13, 25.43, 23.94, 19.69. TOF MS ES+ (C20H28NaO3): Calc. [M + H]+: 339.1936, Found [M + H]+: 339.1923. O IV-117 OTHP Following the above procedure, ketone IV-117 (320 mg, 91% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ: 7.20 (d, J = 8.0 Hz, 2H), 7.07 (d, J = 8.0 Hz, 2H), 6.34 (d, J = 15.7 Hz, 1H), 6.11 (dt, J = 15.7, 6.8 Hz, 1H), 4.54 (dd, J = 4.4, 2.8 Hz, 1H), 3.83 (ddd, J = 11.0, 7.7, 3.2 Hz, 1H), 3.72 (dt, J = 9.5, 6.4 Hz, 1H), 3.51 – 3.43 (m, 1H), 3.36 (dt, J = 9.6, 6.2 Hz, 1H), 2.55 (t, J = 7.5 Hz, 2H), 2.45 (td, J = 7.5, 2.2 Hz, 4H), 2.30 (s, 3H), 1.83 – 1.74 (m, 1H), 1.73 – 1.61 (m, 3H), 1.61 – 1.45 (m, 6H) ppm. 13C NMR (126 MHz, cdcl3) δ 210.15, 136.77, 134.59, 130.48, 129.15, 127.86, 125.85, 98.84, 67.13, 62.32, 42.63, 42.31, 30.71, 29.21, 27.13, 25.44, 21.12, 20.62, 19.63 ppm. TOF MS ES+ (C21H30NaO3): Calc. [M + H]+: 353.2093, Found [M + H]+: 353.2107. 331 O IV-118 OTHP Following the above procedure, ketone IV-118 (300 mg, 90% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.36 – 7.34 (m, 1H), 7.14 – 7.08 (m, 3H), 6.58 (d, J = 15.6 Hz, 1H), 6.03 (dt, J = 15.6, 6.8 Hz, 1H), 4.54 (dd, J = 4.4, 2.8 Hz, 1H), 3.83 (ddd, J = 11.0, 7.7, 3.0 Hz, 1H), 3.73 (dt, J = 9.6, 6.4 Hz, 1H), 3.54 – 3.42 (m, 1H), 3.36 (dt, J = 9.6, 6.3 Hz, 1H), 2.60 – 2.54 (m, 2H), 2.53 – 2.43 (m, 4H), 2.30 (s, 3H), 1.85 – 1.73 (m, 1H), 1.73 – 1.61 (m, 3H), 1.63 – 1.45 (m, 6H). 13C NMR (126 MHz, cdcl3) δ 210.09, 136.52, 134.97, 130.28, 130.15, 128.56, 127.00, 126.00, 125.42, 98.87, 67.15, 62.35, 42.66, 42.38, 30.73, 29.23, 27.48, 25.46, 20.64, 19.81, 19.65. TOF MS ES+ (C21H30NaO3): Calc. [M + H]+: 353.2093, Found [M + H]+: 353.2093. O IV-119 OTHP F Following the above procedure, ketone IV-119 (325 mg, 95% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.30 – 7.20 (m, 2H), 6.99 – 6.91 (m, 2H), 6.34 (dd, J = 15.8, 1.5 Hz, 1H), 6.08 (dt, J = 15.8, 6.9 Hz, 1H), 4.53 (dd, J = 4.5, 2.8 Hz, 1H), 3.83 (ddd, J = 11.0, 7.7, 3.2 Hz, 1H), 3.72 (dt, J = 9.6, 6.4 Hz, 1H), 3.52 – 3.43 (m, 1H), 3.36 332 (dt, J = 9.6, 6.3 Hz, 1H), 2.56 (t, J = 7.4 Hz, 2H), 2.50 – 2.39 (m, 4H), 1.84 – 1.74 (m, 1H), 1.73 – 1.62 (m, 3H), 1.62 – 1.45 (m, 6H). 13C NMR (126 MHz, Chloroform-d) δ 210.01, 161.98 (d, JC-F = 246.0 Hz), 133.55 (d, JC-F = 3.3 Hz), 129.52, 128.70 (d, JC-F = 2.2 Hz), 127.41 (d, JC-F = 7.7 Hz), 115.34 (d, JC-F = 21.5 Hz), 98.89, 67.15, 62.38, 42.65, 42.18, 30.73, 29.23, 27.04, 25.46, 20.64, 19.67. TOF MS ES+ (C20H27FNaO3): Calc. [M + H]+: 357.1842, Found [M + H]+: 357.1869. O IV-120 OTHP Ph Following the above procedure, ketone IV-120 (250 mg, 65% yield) was synthesized as an off white solid. mp = 41– 44° 1H NMR (500 MHz, Chloroform-d) δ 7.59 – 7.54 (m, 2H), 7.54 – 7.47 (m, 2H), 7.46 – 7.35 (m, 3H), 7.35 – 7.27 (m, 1H), 6.46 – 6.38 (m, 1H), 6.22 (dt, J = 15.8, 6.8 Hz, 1H), 4.54 (dd, J = 4.5, 2.7 Hz, 1H), 3.83 (ddd, J = 11.1, 7.6, 3.4 Hz, 1H), 3.73 (dt, J = 9.6, 6.4 Hz, 1H), 3.52 – 3.42 (m, 1H), 3.37 (dt, J = 9.6, 6.2 Hz, 1H), 2.63 – 2.54 (m, 2H), 2.53 – 2.43 (m, 3H), 1.85 – 1.74 (m, 1H), 1.74 – 1.63 (m, 3H), 1.63 – 1.42 (m, 6H). 13C NMR (126 MHz, cdcl3) δ 210.08, 140.75, 139.81, 136.46, 130.25, 129.17, 129.15, 128.77, 128.75, 127.24, 127.19, 127.14, 127.09, 126.98, 126.92, 126.88, 126.46, 126.41, 98.88, 67.17, 62.37, 42.67, 42.24, 30.74, 29.24, 27.20, 25.47, 20.66, 19.66. TOF MS ES+ (C26H32NaO3): Calc. [M + H]+: 415.2249, Found [M + H]+: 415.2264. 333 O IV-121 OTHP O Following the above procedure, ketone IV-121 (290 mg, 95% yield) was synthesized as a colorless oil. 1H NMR (500 MHz, Chloroform-d) δ 7.26 – 7.20 (m, 2H), 6.84 – 6.77 (m, 2H), 6.32 (dt, J = 15.8, 1.5 Hz, 1H), 6.02 (dt, J = 15.8, 6.9 Hz, 1H), 4.54 (dd, J = 4.4, 2.8 Hz, 1H), 3.83 (ddd, J = 11.0, 7.5, 3.4 Hz, 1H), 3.77 (s, 3H), 3.72 (dt, J = 9.6, 6.4 Hz, 1H), 3.53 – 3.43 (m, 1H), 3.36 (dt, J = 9.6, 6.2 Hz, 1H), 2.55 (t, J = 7.5 Hz, 2H), 2.43 (tdd, J = 8.0, 7.3, 4.7, 1.5 Hz, 4H), 1.86 – 1.74 (m, 1H), 1.72 – 1.61 (m, 3H), 1.61 – 1.45 (m, 6H). 13C NMR (126 MHz, cdcl3) δ 210.22, 158.79, 130.23, 130.03, 127.08, 126.74, 113.89, 98.87, 67.16, 62.35, 55.27, 42.65, 42.42, 30.73, 29.23, 27.15, 25.46, 20.64, 19.65. TOF MS ES+ (C21H30NaO4): Calc. [M + H]+: 369.2042, Found [M + H]+: 369.2060. 334 IV-6-5 General procedure for synthesis of ketone IV-131 to IV-133 R1 O R2 R1 R2 EtO O P EtO O OEt R1 OEt LiCl, 0 °C then DBU rt, 12 h R2 O 1. DIBAL-H, DCM, –78 °C 2. PBr3, Et2O 3. NaTs, DMF Ts R1 R2 nBuLi, THF, –78 °C O then OTHP R1 R2 PCC NaOAc DCM O OTHP R1 Pd(PPh3)4 NaBH4 THF:IPA OH OTHP R2 Ts OH OTHP Synthesis of substituted cinnamyl alcohol: R1 O R2 O EtO O P EtO LiCl, 0 °C then DBU rt, 12 h OEt R1 OEt R2 O Following a reported procedure,44 to a solution of lithium chloride (dried under vacuum, 466 mg, 11 mmol, 1.1 equiv) in dry acetonitrile at 0 °C, aldehyde or ketone (10.0 mmol, 1.0 equiv) and triethylphosphonoacetate (2.25 g, 10.0 mmol, 1.0 equiv) were added. After 4 min, DBU (1.5 g, 10.0 mmol,1.0 equiv) was added. The reaction mixture was warmed to room temperature and stirred for 12 h. The reaction mixture was diluted with ethyl acetate (50 mL), washed with saturated ammonium chloride solution and brine. The volatiles were removed under reduced pressure and the resultant crude esters were purified on column chromatography (silica, ethyl acetate/hexanes = 20 – 30%). The resulting esters were reduced with DIBAL-H to yield the allyl alcohols as described below. To a solution of esters (5.0 mmol) in dry DCM (20 mL) at –78 °C was added DIBAL-H (1.0 M in hexane, 10.0 mL, 2.0 equiv). The reaction mixture was warmed to room 335 temperature over 2 h, upon which TLC showed completion of reaction. The reaction mixtures were quenched by addition of methanol (5.0 mL) at 0 °C. Saturated potassium sodium tartrate solution (20.0 mL) was added to the solution and the mixture was vigorously stirred at room temperature for 2 h. The aqueous layer was extracted with DCM (3 x 30 mL). The combined organic extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under vacuum to yield the corresponding allyl alcohols. OH IV-122 Analytical data for alchol IV-122; 1H NMR (500 MHz, CDCl3) δ 7.81 – 7.75 (m, 3H), 7.72 (s, 1H), 7.59 (dd, J = 8.6, 1.8 Hz, 1H), 7.48 – 7.39 (m, 2H), 6.77 (dt, J = 15.9, 1.6 Hz, 1H), 6.48 (dt, J = 15.9, 5.7 Hz, 1H), 4.37 (d, J = 5.6 Hz, 2H), 1.46 (s, 1H). 13C NMR (125 MHz, CDCl3) δ 134.0, 133.5, 133.0, 131.2, 128.8, 128.2, 127.9, 127.6, 126.4, 126.2, 125.9, 123.5, 63.8 ppm. OH IV-123 Analytical data alcohol IV-123; 1H NMR (500 MHz, CDCl3) δ 8.11 (d, J = 8.0 Hz, 1H), 7.84 (dd, J1 = 7.5 Hz, J2 = 2 Hz, 1H), 7.77 (d, J = 8.0 Hz, 1H), 7.57 (d, J = 7.0 Hz, 1H), 7.51-7.46 (m, 2H), 7.43 (dd, J1 = 336 7.5 Hz, J2 = 7.5 Hz, 1H), 7.37 (d, J = 15.5 Hz, 1H), 6.38 (ddd, J1 = 15.5 Hz, J2 = 5 Hz, J3 = 5 Hz, 1H), 4.42 (dd, J1 = 5 Hz, J2 = 1.5 Hz, 2H), 1.66 (br, 1H); 13C NMR (125 MHz, CDCl3) δ 134.4, 133.5, 131.7, 131.1, 128.5, 128.1, 127.9, 126.0, 125.7, 125.5, 123.8, 123.6, 63.9 ppm. Ph IV-124 OH Analytical data for alcohol IV-124; 1H NMR (500 MHz, CDCl3) δ 7.41 – 7.38 (m, 2H), 7.34 – 7.29 (m, 2H), 7.27 – 7.23 (m, 1H), 5.98 – 5.94 (m, 1H), 4.3 (d, J = 6.5 Hz, 2H), 2.08 – 2.06 (m, 3H), 1.73 (br, 1H). 13C NMR (125 MHz, CDCl3) δ 142.79, 137.73, 128.08, 127.21, 126.45, 125.71, 59.86, 15,95. General procedure for substituted allyl sulfonyl substrates: R2 OH R1 1) PBr3, Et2O 2) TsNa, DMF R2 Ts R1 To allyl alcohols (16 mmol) in dry diethyl ether (10 mL) at room temperature was added phosphorous tribromide (6.15 mmol, 0.3 equiv) drop-wise at 0 °C. Reaction was completed in 1 h, as judged by TLC analysis. The reaction mixture was quenched by slow addition sodium bicarbonate (saturated solution, 10 mL). Layers were separated and the aqueous layer was extracted with diethyl ether (3 x 20 m). The combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced 337 pressure at 0 °C (protected from light). The resulting crude allyl bromide was quickly reacted with sodium p-toluenesulfinate (1.5 equiv) in DMF (10 mL) at room temperature. The reaction was stirred for 30 h, and then quenched with saturated ammonium chloride (20 mL). The aqueous layer was extracted with ethyl acetate (3 x 20 m). The combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced pressure. Crude allyl sulfonyl products were purified by crystallization from ethyl acetate. Ts IV-125 Analytical data for sulfonyl IV-125; 1H NMR (500 MHz, CDCl3) δ 7.75-7.79 (m, 5H), 7.63 (s, 1H), 7.48 (d, J = 8.5 Hz, 1H), 7.42-7.47 (m, 2H), 7.30 (d, J = 7.5 Hz, 2H), 6.53 (d, J = 16.0 Hz, 1H), 6.21 (ddd, J1 = 16.0 Hz, J2 = 7.5 Hz, J3 = 7.5 Hz, 1H), 3.97 (d, J = 7.5 Hz, 2H), 2.41 (s, 3H). 13C NMR (125 MHz, CDCl3) δ 144.7, 139.0, 135.4, 133.3, 133.28, 133.21, 129.7, 128.5, 128.3, 128.0, 127.6, 127.0, 126.4, 126.3, 123.2, 115.5, 60.6, 21.6 ppm. IR (film) 3052, 2959, 2917, 1652, 1596, 1510, 1406, 1314 (s), 1288,, 1151, 1136 (s), 1085, 963, 812, 752, 724 cm-1. Ts IV-126 338 Analytical data for sulfonyl IV-126; 1H NMR (500 MHz, CDCl3) δ 7.81 (d, J = 8.0 Hz, 1H), 7.77 (d, J = 8.5 Hz, 3H), 7.66 (d, J = 8.5 Hz, 1H), 7.48-7.40 (m, 4H), 7.31 (d, J = 8.0, 2H), 7.05 (d, J = 15.0 Hz, 1H), 6.11 (ddd, J1 = 15.0 Hz, J2 = 7.5 Hz, J3 = 7.5 Hz, 1H), 4.03 (d, J = 7.5 Hz, 2H), 2.42 (s, 3H); 13C NMR (125 MHz, CDCl3) δ 144.7, 136.7, 135.2, 133.5, 133.4, 130.7, 129.7, 128.7, 128.6, 128.5, 126.1, 125.9, 125.5, 124.2, 123.4, 118.7, 60.7, 21.6 ppm. IR (film) 3047, 2976, 2925, 1, 1595, 1508, 1398, 1316 (s), 1292, 1148 (s), 1088, 967, 799, 778, 726, 669 cm-1. Ph IV-127 Ts Analytical data for sulfonyl IV-127; 1H NMR (500 MHz, CDCl3) δ 7.76 – 7.74 (m, 2H), 7.31 – 7.24 (m, 7H), 5.71 – 5.69 (td, J = 8.0, 1.5 Hz, 1H), 3.97 (d, J = 8.0 Hz, 2H), 2.42 (s, 3H), 1.69 (d, J = 1.5 Hz, 1H). 13C NMR (125 MHz, CDCl3) δ 144.65, 144.34, 142.24, 135.69, 129.67, 128.51, 128.33, 127.83, 125.81, 113.40, 56.72, 21.63, 15.98. 339 General procedure for synthesis of substituted sulfonyl alcohol substrates: R2 Ts R1 O 2) 1) nBuLi THF OTHP R1 R2 Ts OH OTHP In a typical procedure, allyl sulfonyl substrates (2.0 mmol) were dissolved in dry THF (3 mL) under argon. To this solution at –78 °C was added nBuLi (1.8 M in hexane, 1.2 mL, 2.1 mmol, 1.05 equiv) dropwise. The reaction was stirred at this temperature for 5 min, then it was warmed to room temperature and stirring was continued for another 10 min. To this mixture at –78 °C was added neat THP-protected epoxy alcohol (420.2 mg, 2.1 mmol, 1.05 equiv) via syringe over 5 minutes. The reaction mixture was warmed to room temperature and was followed by TLC. After completion of the reaction (~1 h), saturated ammonium chloride (10 mL) was added to the mixture. The aqueous layer was extracted with ethyl acetate (3 x 10 m). The combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced pressure. The crude mixture was purified via column chromatography yielding a ~1:1 diastereomeric mixture which was subjected to the next step without further separating the diastereomers. General procedure for detosylation of substituted sulfonyl alcohol substrates: R2 Ts OH R1 OTHP Pd(PPh3)4 NaBH4 THF:IPA 3:2 R2 OH R1 OTHP Diastereomeric sulfonyl alcohol (1.5 mmol) was dissolved in a mixture of THF:IPA (3:2, 20 mL) under an argon atmosphere. To this mixture at room temperature was added 340 tetrakis(triphenylphosphine)palladium (86 mg, 0.075 mmol, 0.05 equiv). The reaction mixture was placed in an ice bath at 0 ºC. To this cold mixture sodium borohydride (582 mg, 15.0 mmol, 10.0 mmol) was added in a few portions. The reaction was stirred at room temperature for 12 h, then it was quenched at 0 °C by careful addition of water (10 mL). The aqueous layer was extracted with ethyl acetate (3 x 20 mL). The combined organic layers were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude product was purified via column chromatography yielding alcohol products in good yield. OH IV-128 OTHP Analytical data for alcohol IV-128: 1H NMR (500 MHz, CDCl3) δ 7.78 – 7.73 (m, 3H), 7.65 (d, J = 1.6 Hz, 1H), 7.55 (dd, J = 8.5, 1.8 Hz, 1H), 7.44 – 7.37 (m, 2H), 6.60 – 6.53 (m, 1H), 6.35 (dt, J = 15.8, 6.9 Hz, 1H), 4.55 (dd, J = 4.6, 2.8 Hz, 1H), 3.89 – 3.81 (m, 1H), 3.77 – 3.66 (m, 2H), 3.54 – 3.43 (m, 2H), 3.39 (dtd, J = 9.7, 6.4, 1.5 Hz, 1H), 2.38 (m, 2H), 1.88 – 1.75 (m, 2H), 1.73 – 1.37 (m, 12H). OH IV-129 OTHP Analytical data for alcohol IV-129: 341 1H NMR (500 MHz, CDCl3) δ 8.14 – 8.07 (m, 1H), 7.83 – 7.79 (m, 1H), 7.75 – 7.70 (m, 1H), 7.53 (dt, J = 7.1, 1.0 Hz, 1H), 7.50 – 7.38 (m, 3H), 7.21 – 7.10 (m, 1H), 6.23 (dt, J = 15.5, 6.9 Hz, 1H), 4.58 – 4.51 (m, 1H), 3.89 – 3.82 (m, 1H), 3.79 – 3.65 (m, 2H), 3.52 – 3.44 (m, 1H), 3.38 (m, 1H), 2.55 – 2.32 (m, 2H), 1.81 (qt, J = 8.7, 2.5 Hz, 1H), 1.75 – 1.39 (m, 14H). OH IV-130 OTHP Analytical data for alcohol VI-130: 1H NMR (500 MHz, CDCl3) δ 7.36 – 7.34 (m, 2H), 7.29 – 7.26 (m, 2H), 7.20 – 7.18 (m, 1H), 5.78(t, J = 7.5 Hz, 1H), 4.56 (m, 1H), 3.86 – 3.83 (m, 1H), 3.77 – 3.72 (m, 1H), 3.66 (m, 1H), 3.50 – 3.47 (m, 1H), 3.41 – 3.37 (m, 1H), 2.34 – 2.27 (m, 2H), 2.04 (s, 3H), 1.81 – 1.78 (m, 1H), 1.71 – 1.66 (m, 1H), 1.63 – 1.43 (m, 12H). 13C NMR (126 MHz, cdcl3) δ 143.81, 135.15, 128.12, 127.88, 126.52, 125.56, 98.92, 71.46, 67.48, 62.37, 37.31, 37.14, 30.75, 29.66, 25.46, 25.06, 22.40, 19.66, 15.78. General procedure for oxidation of the alcohol substrates into olefinic ketones: OH R2 R1 OTHP PCC NaOAc DCM O R2 R1 OTHP The olefinic alcohol (1.0 mmol) was dissolved in DCM (5 mL) in a 20 mL screw-top vial. To this solution at room temperature were added pyridinium chlorochromate (323.4 mg, 342 1.5 mmol, 1.5 equiv) and sodium acetate (24.6 mg, 0.3 mmol, 0.3 equiv). The reaction was stirred at this temperature until TLC showed complete consumption of the alcohol (ca. 1 h). The crude mixture was diluted with ethyl acetate and the precipitates were removed by passing the mixture through celite. Volatiles were removed and the resulting crude ketone was purified via column chromatography (ethyl acetate/hexanes = 20%). O IV-131 OTHP Analytical data for ketone IV-131: 1H NMR (500 MHz, CDCl3) δ 7.77 – 7.72 (m, 3H), 7.65 (d, J = 1.6 Hz, 1H), 7.53 (dd, J = 8.5, 1.8 Hz, 1H), 7.44 – 7.37 (m, 2H), 6.54 (d, J = 15.8 Hz 1H), 6.31 (dt, J = 15.8, 6.8 Hz, 1H), 4.54 (dd, J = 4.4, 2.8 Hz, 1H), 3.83 (ddd, J = 11.1, 7.6, 3.2 Hz, 1H), 3.73 (dt, J = 9.6, 6.4 Hz, 1H), 3.49 – 3.44 (m, 1H), 3.37 (dt, J = 9.7, 6.2 Hz, 1H), 2.61 (dd, J = 8.2, 6.8 Hz, 2H), 2.55 – 2.49 (m, 2H), 2.46 (td, J = 7.3, 3.7 Hz, 2H), 1.79 (qd, J = 8.0, 3.7 Hz, 1H), 1.68 (dddd, J = 12.6, 9.1, 5.7, 2.4 Hz, 3H), 1.62 – 1.45 (m, 6H). 13C NMR (125 MHz, CDCl3) δ 210.04, 134.82, 133.58, 133.27, 132.69, 130.76, 129.39, 128.04, 127.80, 127.57, 126.12, 125.54, 123.41, 98.84, 67.12, 62.32, 42.63, 42.20, 30.69, 29.20, 27.21, 25.42, 20.62, 19.62. TOF MS ES+ (C24H30NaO3): Calc. [M + H]+: 389.2093, Found [M + H]+: 389.2119. 343 O IV-132 OTHP Analytical data for ketone IV-132: 1H NMR (500 MHz, Chloroform-d) δ 8.13 – 8.07 (m, 1H), 7.85 – 7.81 (m, 1H), 7.75 (d, J = 8.0 Hz, 1H), 7.54 – 7.51 (m, 1H), 7.49 (td, J = 8.0, 1.5 Hz, 2H), 7.44 – 7.40 (m, 1H), 7.15 (d, J = 15.5 Hz, 1H), 6.20 (dt, J = 15.5, 6.6 Hz, 1H), 4.55 (dt, J = 5.7, 2.7 Hz, 1H), 3.84 (td, J = 8.3, 3.9 Hz, 1H), 3.75 (dt, J = 9.6, 6.4 Hz, 1H), 3.54 – 3.45 (m, 1H), 3.38 (dt, J = 9.8, 6.4 Hz, 1H), 2.71 – 2.64 (m, 2H), 2.65 – 2.58 (m, 2H), 2.51 (t, J = 7.3 Hz, 2H), 1.85 – 1.77 (m, 1H), 1.74 – 1.66 (m, 3H), 1.66 – 1.44 (m, 6H). TOF MS ES+ (C24H30NaO3): Calc. [M + H]+: 389.2093, Found [M + H]+: 389.2121. O IV-133 OTHP Analytical data for ketone IV-133: 1H NMR (500 MHz, Chloroform-d) δ 7.36 – 7.32 (m, 2H), 7.29 – 7.24 (m, 2H), 7.20 – 7.18 (m, 1H), 5.67 (t, J = 6.5 Hz, 1H), 4.54 (m, 1H), 3.84 – 3.80 (m, 1H), 3.74 – 3.70 (m, 1H), 3.49 – 3.46 (m, 1H), 3.38 – 3.33 (m, 1H), 2.53 (t, J = 6.5 Hz, 2H), 2.46 – 2.43 (m, 4H), 2.02 (s, 3H), 1.80 – 1.76 (m, 1H), 1.70 – 1.63 (m, 3H), 1.60 – 1.51 (m, 6H). 13C NMR (126 MHz, cdcl3) δ 210.36, 143.61, 135.80, 128.14, 126.65, 126.46, 125.59, 98.85, 67.13, 62.32, 42.61, 42.35, 30.71, 29.22, 25.44, 23.12, 20.64, 19.62, 15.79. 344 IV-6-6 Procedure for the synthesis of THP-protected keto alkenols IV-136 I OTHP IV-134 2) Ph 1) tBuLi O N O IV-135 Ph O IV-136 OTHP To a flame dried round bottom flask (10 mL) containing dry diethyl ether (4 mL) at – 100 ºC (liquid nitrogen-methanol bath) was added tBuLi (1.7 M solution in pentane, 1.5 mL) dropwise. The THP protected alkyl iodide IV-134 (1 mmol) in dry diethyl ether (1 mL) was added over 5 min. The reaction was stirred at –100 °C for 10 min, then slowly brought to room temperature and kept at that temperature for 10 min. After this period of time (~30 min), the reaction mixture was placed in to a liquid nitrogen-methanol bath (–100 ºC) and a solution of the corresponding morpholine amide IV-135 (0.5 mmol) dry ether (1 mL) was added over 5 min. The reaction mixture stirred at –78 ºC and was monitored by TLC. After complete consumption of the amide starting material, the reaction was quenched at –78 °C with NaCl solution (sat.). The organic phase was separated and the aqueous layer was washed with ethyl acetate (3 x 5 mL). The combined organics were concentrated under reduced pressure to yield the crude product. Column chromatography of the crude mixture (ethyl acetate/hexanes = 20%) led to the isolation of the pure THP-protected keto alkenol IV-136 (80 mg, 60% yield) as a clear oil. 345 1H NMR (500 MHz, Chloroform-d) δ 7.37 – 7.25 (m, 4H), 7.25 – 7.15 (m, 1H), 6.48 – 6.36 (m, 1H), 6.20 (dt, J = 15.8, 6.8 Hz, 1H), 4.54 (dd, J = 4.7, 2.7 Hz, 1H), 3.88 – 3.79 (m, 1H), 3.73 (tt, J = 8.6, 6.3 Hz, 1H), 3.53 – 3.45 (m, 1H), 3.41 (dt, J = 9.7, 6.1 Hz, 1H), 2.66 – 2.58 (m, 2H), 2.58 – 2.52 (m, 2H), 2.52 – 2.46 (m, 2H), 1.94 – 1.86 (m, 2H), 1.83 – 1.76 (m, 1H), 1.73 – 1.65 (m, 2H), 1.59 – 1.48 (m, 4H). 13C NMR (126 MHz, cdcl3) δ 209.70, 137.31, 130.57, 128.89, 128.39, 126.96, 125.89, 98.80, 66.47, 62.36, 42.14, 39.63, 30.60, 27.04, 25.34, 23.85, 19.60. 346 IV-7 Experimental date for the synthesis of Obtusin O R R IV-78 O Synthesis of Diepoxide IV-78: Following the reported procedure,36-37 7.5-gram IV-78 as a clear oil (solidifies at –20 °C to a white solid) was synthesized from D-tartaric acid. 1H NMR (500 MHz, CDCl3) δ 2.89 – 2.84 (m, 2H), 2.80 (m, 2H), 2.71 (m, 2H). 13C NMR (125 MHz, CDCl3) δ 77.3, 77.0, 76.7, 51.1, 44.3. TBDPS O IV-79 1) nBuLi 2) BF3 •OEt2 3) O O IV-78 60% O TBDPS OH IV-80 O Synthesis of epoxy alcohol IV-80: Following a reported procedure,38 to IV-79 (1.0 equiv, 10.0 mmol, 2.9 g) in dry THF (90 mL) at –78 °C nBuLi (2.0 M in hexanes, 1.0 equiv, 10.0 mmol, 5 mL) was added dropwise and the reaction was stirred at this temperature for 30 min. To this mixture, was added BF3•OEt2 (1.0 equiv, 10.0 mmol, 1.3 mL), followed by rapid addition of IV-78 (1.0 equiv, 10.0 mmol, 860 mg) in dry THF (10 mL) at –78 °C. The reaction was stirred at this temperature for 5 h, before it was quenched by addition of saturated aqueous NH4Cl (50 mL). Layers were separated and the aqueous layer was extracted with ethyl acetate (3 x 50 mL). The combined extracts were washed with brine (50 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude 347 residue was purified via column chromatography (silica, ethyl acetate = 20 – 30%) to give pure epoxy alcohol IV-80 as a clear oil (2.2 g, 60%). 1H NMR (500 MHz, CDCl3) δ 7.72 – 7.64 (m, 4H), 7.44 – 7.34 (m, 5H), 4.31 (t, J = 2.2 Hz, 2H), 3.56 (ddd, J = 10.5, 7.5, 4.4 Hz, 1H), 3.05 (td, J = 4.2, 2.7 Hz, 1H), 2.77 (dd, J = 4.9, 4.1 Hz, 1H), 2.71 (dd, J = 4.9, 2.7 Hz, 1H), 2.53 – 2.40 (m, 2H), 1.03 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.59, 133.16, 129.83, 129.81, 129.79, 127.69, 127.68, 81.16, 80.50, 72.20, 70.88, 69.41, 64.69, 53.95, 52.80, 44.86, 26.68, 26.67, 25.07, 24.27, 19.15. TOF MS ES+ (C23H29O3Si): Calc. [M + H]+: 381.1886, Found [M + H]+: 381.1875. O TBDPS OH IV-80 O Lindlar's Cat. H2 (gas) EtOAc quant. O TBDPS O IV-81 OH Synthesis of IV-81: To epoxy alcohol IV-80 (900 mg, 2.4 mmol) in ethyl acetate (100 mL) was added Lindlar’s catalyst (100 mg) at room temperature. The reaction flask was placed under vacuum and backfilled with hydrogen gas (balloon) three times. The reaction was stirred vigorously at room temperature for 30 min, upon which 1H NMR showed complete consumption of starting material. The mixture was passed through a pad of celite to remove the insoluble material and the crude was used in the following step without further purification. A small sample of the crude was passed through a plug of silica to afford the pure alcohol IV-81. 1H NMR (500 MHz, CDCl3) δ 7.72 – 7.68 (m, 4H), 7.47 – 7.38 (m, 6H), 5.81 (dtt, J = 11.0, 6.3, 1.5 Hz, 1H), 5.55 (dtt, J = 10.9, 7.7, 1.6 Hz, 1H), 4.30 – 4.17 (m, 2H), 3.45 (qd, J = 348 6.3, 4.5 Hz, 1H), 2.93 (td, J = 4.3, 2.7 Hz, 1H), 2.75 (dd, J = 5.0, 4.1 Hz, 1H), 2.68 (dd, J = 5.0, 2.7 Hz, 1H), 2.30 – 2.23 (m, 2H), 1.98 (d, J = 6.1 Hz, 1H), 1.05 (s, 10H). 13C NMR (125 MHz, CDCl3) δ 135.59, 135.56, 133.56, 133.54, 132.30, 129.67, 129.55, 127.69, 127.68, 127.66, 127.65, 127.61, 125.66, 70.54, 59.97, 54.69, 44.79, 32.78, 26.77, 19.13. TOF MS ES+ (C23H31O3Si): Calc. [M + H]+: 383.2042, Found [M + H]+: 383.2039. O TBDPS O IV-81 OH DHP O TBDPS PPTS, DCM 85% ~2:1 dr on the THP group IV-82 O O THP Synthesis of THP protected epoxy alcohol IV-91: To alcohol IV-81 (1.53 g, 4.0 mmol) in dry DCM (20 mL) at 0 °C were added 3,4-dihydro-2H-pyran (DHP, 1.1 equiv, 4.4 mmol, 0.41 mL) and PPTS (0.1 equiv, 0.4, 100 mg). The reaction was stirred for 12 h, allowing it to warm up to room temperature. Solvent was removed under reduced pressure and the crude product was purified using column chromatography (silica, ethyl acetate/ hexanes = 5, to 10%) to give two diastereomers (1.6 g, 85% combined yield). Data for the less polar diastereomer (F1): 1H NMR (500 MHz, CDCl3) δ 7.70 – 7.61 (m, 4H), 7.44 – 7.32 (m, 6H), 5.71 (dtt, J = 11.1, 6.1, 1.6 Hz, 1H), 5.50 (dtt, J = 11.0, 7.5, 1.7 Hz, 1H), 4.89 (dd, J = 4.5, 2.8 Hz, 1H), 4.29 – 4.23 (m, 2H), 3.78 (ddd, J = 11.3, 8.4, 3.3 Hz, 1H), 3.41 (dddd, J = 10.8, 4.9, 3.9, 1.4 Hz, 1H), 3.31 (q, J = 6.8 Hz, 1H), 2.84 (ddd, J = 7.1, 4.2, 2.7 Hz, 1H), 2.62 (dd, J = 4.9, 349 4.2 Hz, 1H), 2.37 (dd, J = 4.8, 2.7 Hz, 1H), 2.30 – 2.14 (m, 2H), 1.82 – 1.64 (m, 2H), 1.58 – 1.41 (m, 5H), 1.02 (s, 10H). 13C NMR (125 MHz, CDCl3) δ 135.54, 133.69, 133.68, 131.67, 129.60, 127.64, 125.53, 97.29, 76.45, 62.43, 60.29, 54.55, 43.54, 31.08, 30.72, 26.78, 26.76, 25.40, 19.40, 19.13. TOF MS ES+ (C28H39O4Si): Calc. [M + H]+: 467.2618, Found [M + H]+: 467.2611. Data for the more polar diastereomer (F2): 1H NMR (500 MHz, CDCl3) δ 7.70 – 7.63 (m, 4H), 7.44 – 7.34 (m, 6H), 5.70 (dtt, J = 12.3, 6.1, 1.6 Hz, 1H), 5.44 (dtt, J = 11.0, 7.6, 1.7 Hz, 1H), 4.61 (dd, J = 4.3, 2.9 Hz, 1H), 4.24 (dd, J = 6.3, 1.7 Hz, 2H), 3.92 (ddd, J = 11.5, 8.1, 3.4 Hz, 1H), 3.48 – 3.42 (m, 1H), 3.35 (q, J = 6.3 Hz, 1H), 2.97 (ddd, J = 6.2, 4.1, 2.7 Hz, 1H), 2.68 (dd, J = 5.0, 4.1 Hz, 1H), 2.55 (dd, J = 5.0, 2.7 Hz, 1H), 2.19 – 2.11 (m, 2H), 1.77 (tdd, J = 15.9, 8.4, 4.3 Hz, 1H), 1.68 – 1.59 (m, 1H), 1.58 – 1.44 (m, 4H), 1.02 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.60, 135.57, 135.55, 133.67, 131.90, 129.64, 127.67, 125.32, 97.39, 76.53, 62.37, 60.21, 54.00, 44.54, 30.64, 29.71, 26.79, 25.34, 19.46, 19.15. TOF MS ES+ (C28H39O4Si): Calc. [M + H]+: 467.2618, Found [M + H]+: 467.2614. 350 O TBDPS O THP O IV-82 KCN Methanol 87% O TBDPS OH CN O THP IV-86 Synthesis of nitrile IV-95: To epoxide IV-82 (200 mg, 0.4 mmol) in methanol (10 mL), potassium cyanide (10.0 equiv, 4.0 mmol, 260 mg) was added at room temperature. Stirring was continued for 48 h upon which TLC showed complete consumption of epoxide IV-82. Saturated ammonium chloride (20 mL) was added to the reaction mixture and the aqueous layer was extracted with ethyl acetate (3 x 20 ml). The combined organic extracts were washed with brine (50 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude residue was purified via column chromatography (silica, ethyl acetate = 20 – 30%) to give nitrile IV-86 as a clear oil (170 mg, 87%). 1H NMR (500 MHz, CDCl3) δ 7.69 – 7.64 (m, 4H), 7.45 – 7.35 (m, 6H), 5.83 – 5.64 (m, 1H), 5.43 (dtt, J = 10.8, 7.6, 1.6 Hz, 1H), 4.38 (dd, J = 7.3, 2.5 Hz, 1H), 4.27 – 4.21 (m, 1H), 4.23 – 4.14 (m, 1H), 3.99 (d, J = 4.2 Hz, 1H), 3.92 (dtd, J = 11.5, 3.7, 1.4 Hz, 1H), 3.70 (tdd, J = 6.0, 4.9, 4.2 Hz, 1H), 3.54 (dt, J = 7.5, 5.5 Hz, 1H), 3.46 (dddd, J = 11.5, 8.3, 5.7, 3.1 Hz, 1H), 2.56 (dd, J = 16.8, 4.9 Hz, 1H), 2.40 (ddd, J = 16.7, 6.1, 0.8 Hz, 1H), 2.18 – 2.05 (m, 2H), 1.82 – 1.73 (m, 1H), 1.73 – 1.65 (m, 1H), 1.52 – 1.37 (m, 5H), 1.03 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.59, 135.57, 135.55, 135.48, 133.51, 133.50, 132.17, 129.71, 127.73, 127.71, 127.70, 127.67, 127.62, 125.33, 117.48, 100.88, 80.76, 68.71, 65.35, 60.02, 31.11, 29.42, 26.78, 24.92, 22.45, 21.23, 19.13. TOF MS ES+ (C29H40NO4Si): Calc. [M + H]+: 494.2727, Found [M + H]+: 494.2722. 351 TBDPS O THP O CN HO IV-86 TMSCl Et3N, DMAP (cat.) DCM, r.t. 95% TBDPS O THP O CN IV-92 O TMS Synthesis of IV-101: Alcohol IV-95 (7.5 mg, 0.015) was dissolved in dry DCM (1 mL), and to this solution at 0 °C were added triethyl amine (3.0 equiv, 0.045 mmol, 7 µL), DMAP (1 mg) and trimethylsilyl chloride (3.0 equiv, 0.045 mmol, 6 µL). After stirring at this temperature for 30 min, TLC showed completion of reaction. Solvent was removed under reduced pressure, and the crude product was passed through a plug of silica (basified with triethyl amine) Ethyl acetate in hexanes (5%) was used as eluent to provide the pure product in >95% yield (8 mg product was isolated). Note, TMS can fall off if the silica is not basified with triethyl amine. 1H NMR (500 MHz, CDCl3) δ 7.69 – 7.62 (m, 4H), 7.43 – 7.33 (m, 6H), 5.72 – 5.63 (m, 1H), 5.40 (dddd, J = 9.4, 7.7, 6.3, 3.7 Hz, 1H), 4.41 (dd, J = 5.9, 2.7 Hz, 1H), 4.24 – 4.19 (m, 2H), 4.13 – 4.08 (m, 1H), 3.80 (dt, J = 10.6, 4.8 Hz, 1H), 3.46 – 3.38 (m, 2H), 2.58 (dd, J = 16.6, 3.1 Hz, 1H), 2.27 (dd, J = 16.6, 9.3 Hz, 1H), 2.18 – 2.11 (m, 1H), 1.88 (dt, J = 15.7, 8.7 Hz, 1H), 1.78 – 1.68 (m, 1H), 1.60 (ddd, J = 12.7, 6.2, 2.9 Hz, 1H), 1.50 – 1.35 (m, 4H), 1.02 (s, 9H), 0.10 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.57, 133.67, 131.71, 129.64, 129.63, 127.67, 127.65, 126.45, 119.06, 101.01, 79.61, 70.51, 63.83, 60.23, 30.83, 26.79, 26.69, 25.19, 21.13, 20.45, 19.15, 0.17, 0.08. 352 TOF MS ES+ (C32H48NO4Si2): Calc. [M + H]+: 566.3122, Found [M + H]+: 566.3119. General procedure for the bromo-etherification of alcohol IV-95 (Table IV-3): TBDPS O THP O HO IV-86 CN TBCO THF/HMPA OTHP Br TBDPSO + O H H CN Br TBDPSO OTHP O H H CN IV-87 desired IV-88 undesired diastereomer diastereomer To alcohol IV-95 (0.06 mmol, 30 mg) in THF (1 mL) and HMPA (40 µL) was added TBCO (1.2 equiv, 0.07, 30 mg) at room temperature. The reaction was stirred at this temperature and monitored by TLC. After 30 min, TLC showed complete consumption of alcohol IV- 95. Aqueous sodium thiosulfate (10%, 1 mL) was added to the reaction mixture and the aqueous layer was extracted with ethyl acetate (3 x 2 mL). The combined organic extracts were washed with brine (2 mL), dried over sodium sulfate and concentrated under reduced pressure. 1H NMR analysis of the crude mixture showed ~1:1 ratio of diastereomers. The crude residue was purified via column chromatography (silica, ethyl acetate = 10 – 20%) to give both diastereomers (15 mg, 45% IV-96 and 17 mg, 49% IV- 97 were isolated; 94% combined yield). 353 General procedure for the bromo-etherification of TMS-protected alcohol IV-101 (Table IV-4): TBDPS O THP O CN TBCO DCM, MeOH O TMS IV-92 OTHP Br TBDPSO + O H H CN Br TBDPSO OTHP O H H CN IV-87 desired IV-88 undesired diastereomer diastereomer To TMS-protected alcohol IV-101 (0.5 mmol, 280 mg) in anhydrous DCM (50 mL) were added methanol (10.0 equiv, 5.0 mmol, 0.2 mL) and TBCO (1.2 equiv, 0.6, 246 mg) at room temperature. The reaction was stirred at this temperature and monitored by TLC. After 1 h, TLC showed complete consumption of IV-101. The solvent was removed under reduced pressure and 1H NMR analysis of the crude mixture showed >15:1 ratio of diastereomers. The crude residue was purified via column chromatography (silica, ethyl acetate = 10 – 20%) to give both diastereomers (240 mg, 85% IV-96 and 10 mg, 4% IV- 97 were isolated; 89% combined yield). 354 OTHP O H H CN Br TBDPSO IV-88 undesired diastereomer Data for the less polar diastereomer (undesired diastereomer IV-88): 1H NMR (500 MHz, CDCl3) δ 7.69 – 7.61 (m, 4H), 7.46 – 7.32 (m, 6H), 4.71 – 4.65 (m, 1H), 4.54 (ddd, J = 8.8, 6.5, 2.2 Hz, 1H), 4.49 (td, J = 4.7, 2.1 Hz, 1H), 4.35 (ddd, J = 6.9, 6.1, 4.2 Hz, 1H), 4.02 – 3.92 (m, 3H), 3.90 – 3.83 (m, 1H), 3.57 – 3.48 (m, 1H), 2.78 (dd, J = 16.5, 7.0 Hz, 1H), 2.70 – 2.61 (m, 1H), 2.18 (ddd, J = 13.5, 6.5, 2.1 Hz, 1H), 2.08 (ddd, J = 13.7, 9.0, 5.0 Hz, 1H), 1.81 (tdd, J = 9.2, 7.5, 6.3, 4.0 Hz, 1H), 1.77 – 1.68 (m, 1H), 1.55 (pd, J = 9.3, 7.5, 5.6 Hz, 4H), 1.04 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.59, 135.53, 135.52, 133.09, 132.86, 129.85, 127.78, 127.77, 127.74, 96.00, 78.18, 76.24, 74.63, 65.36, 62.70, 58.10, 35.34, 30.64, 26.77, 25.23,19.28, 18.67. TOF MS ES+ (C29H38BrNO4SiNa): Calc. [M + Na]+: 594.1651, Found [M + Na]+: 594.1675. 355 OTHP O H H CN Br TBDPSO IV-87 desired diastereomer Data for the more polar diastereomer (desired diastereomer IV-87): 1H NMR (500 MHz, CDCl3) δ 7.69 – 7.62 (m, 4H), 7.47 – 7.33 (m, 6H), 4.56 (t, J = 3.7 Hz, 1H), 4.49 (q, J = 6.1 Hz, 1H), 4.26 (ddd, J = 8.5, 6.9, 4.6 Hz, 1H), 4.19 (q, J = 6.1 Hz, 1H), 4.05 (dt, J = 6.6, 5.1 Hz, 1H), 3.99 (dd, J = 10.9, 6.6 Hz, 1H), 3.88 (dd, J = 10.9, 5.4 Hz, 1H), 3.82 (ddd, J = 11.5, 7.9, 3.4 Hz, 1H), 3.49 (dt, J = 10.8, 4.7 Hz, 1H), 2.75 (dd, J = 16.7, 6.2 Hz, 1H), 2.63 (dd, J = 16.6, 6.4 Hz, 1H), 2.22 (dt, J = 13.3, 6.7 Hz, 1H), 1.97 (ddd, J = 13.6, 8.5, 5.7 Hz, 1H), 1.76 (td, J = 13.1, 11.1, 5.1 Hz, 1H), 1.64 (td, J = 9.1, 2.9 Hz, 1H), 1.59 – 1.43 (m, 4H), 1.04 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.60, 135.59, 135.53, 135.52, 133.02, 132.79, 129.87, 127.79, 127.77, 96.70, 77.12, 76.56, 74.09, 65.28, 62.81, 56.40, 34.42, 30.44, 26.78, 25.21, 19.36, 19.28. TOF MS ES+ (C29H38BrNO4SiNa): Calc. [M + Na]+: 594.1651, Found [M + Na]+: 594.1674. 356 Synthesis of di-TMS-protected diol IV-95: TBDPS O O KCN MeOH TBDPS O OH CN HO TMSCl Et3N DCM 75% IV-89 TMS O TBDPS O CN IV-81 OH Epoxy alcohol IV-81 (100 mg, 0.26 mmol) was reacted with potassium cyanide according to the procedure described above for the synthesis of IV-86. The crude alcohol was (two steps) IV-95 O TMS reacted with TMSCl (3.0 equiv, 0.78 mmol, 104 µL) and triethyl amine (3.0 equiv, 0.78 mmol, 121 µL) in the presence of DMAP (5 mg) in DCM (5 mL) at 0 °C. After completion of the reaction (20 min), the crude product was purified with column chromatography (silica basified with Et3N, ethyl acetate/hexanes 5%). IV-95 was purified as a clear oil (108 mg, 75% yield). 1H NMR (500 MHz, CDCl3) δ 7.77 – 7.60 (m, 4H), 7.48 – 7.34 (m, 6H), 5.72 (dtt, J = 11.1, 6.2, 1.5 Hz, 1H), 5.49 – 5.37 (m, 1H), 4.30 – 4.19 (m, 2H), 3.81 (ddd, J = 8.8, 4.4, 3.4 Hz, 1H), 3.52 (ddd, J = 8.6, 4.4, 3.5 Hz, 1H), 2.56 (dd, J = 16.6, 3.4 Hz, 1H), 2.28 – 2.20 (m, 1H), 2.20 – 2.10 (m, 1H), 1.90 – 1.77 (m, 1H), 1.05 (s, 10H), 0.13 (s, 9H), 0.07 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.57, 135.56, 133.70, 133.69, 131.60, 129.62, 127.67, 127.66, 126.71, 118.91, 74.01, 71.60, 60.26, 28.73, 26.79, 20.71, 19.15, 0.29, 0.15. TOF MS ES+ (C30H48NO3Si3): Calc. [M + H]+: 554.2942, Found [M + H]+: 554.2940. TBDPS O OH CN IV-89 HO 357 This diol was purified form a reaction of epoxide IV-81 and potassium cyanide as specified above. The crude diol was purified with column chromatography (silica, ethyl acetate/hexan = 30 – 40%) leading to pure diol IV-89. 1H NMR (500 MHz, CDCl3) δ 7.74 – 7.65 (m, 4H), 7.46 – 7.33 (m, 65H), 4.33 (t, J = 2.2 Hz, 2H), 3.81 (qd, J = 6.3, 2.7 Hz, 1H), 3.58 (qd, J = 6.1, 2.8 Hz, 1H), 2.61 – 2.48 (m, 2H), 2.48 – 2.36 (m, 2H), 2.14 (d, J = 6.2 Hz, 1H), 1.03 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 135.61, 135.59, 133.17, 133.12, 129.90, 129.88, 127.75, 127.74, 117.27, 82.03, 80.42, 70.52, 68.75, 52.72, 26.64, 24.29, 22.79, 19.13. TBDPS O THP O CN IV-92 O TMS Li THF, –78 °C 30% O TBDPS TMS O O O THP IV-94 Synthesis of ketone IV-94: To a flame dried flask containing diethyl ether (4.0 mL) under argon at –100 °C (liquid nitrogen in methanol bath) was added tBuLi (1.7 M in pentane, 0.42 mL, 0.7 mmol, 7.0 equiv) dropwise. To this solution, alkyl iodide (63 mg, 0.3 mmol, 3.0 equiv) in diethyl ether (2.0 mL) was added dropwise at the same temperature. The reaction was stirred at this temperature for 30 min. To this solution at –78 °C was added a solution of the nitrile IV-92 (56 mg, 0.1 mmol) in diethyl ether (2 mL)., and the reaction was stirred at for 3 h. Saturated sodium bicarbonate (10 mL) was added to this solution at –78 °C to quench the reaction while allowing it to warm up to room temperature. The aqueous layer was extracted with ethyl acetate (3 x 5 mL), and the combined organic 358 extracts were washed with brine (20 mL), dried over sodium sulfate and concentrated under reduced pressure. The crude ketone was purified via column chromatography (silica, ethyl acetate/hexanes 5 – 10%) to afford the ketone IV-94 as a clear oil (18.0 mg, 30%). 1H NMR (500 MHz, CDCl3) δ 7.72 – 7.60 (m, 4H), 7.44 – 7.32 (m, 6H), 5.73 – 5.62 (m, 1H), 5.56 (dtt, J = 10.6, 7.2, 1.6 Hz, 1H), 5.44 (dtt, J = 15.1, 6.1, 1.3 Hz, 1H), 5.30 – 5.38 (m, 1H), 4.63 – 4.59 (m, 1H), 4.28 (dd, J = 6.7, 5.2 Hz, 2H), 3.83 (ddd, J = 11.1, 8.2, 2.9 Hz, 1H), 3.55 (ddd, J = 8.4, 4.5, 3.4 Hz, 1H), 3.41 – 3.34 (m, 1H), 2.44 – 2.38 (m, 4H), 2.30 – 2.14 (m, 3H), 2.02 – 1.89 (m, 3H), 1.68 (dq, J = 8.5, 5.4, 4.9 Hz, 1H), 1.60 (td, J = 9.3, 3.1 Hz, 1H), 1.51 – 1.40 (m, 4H), 1.03 (s, 9H), 0.01 (s, 9H). 13C NMR (125 MHz, CDCl3) δ 209.27, 135.58, 133.87, 133.86, 132.95, 130.68, 129.54, 129.53, 127.82, 127.62, 127.31, 97.25, 77.75, 68.19, 62.42, 60.57, 44.49, 44.35, 30.83, 27.44, 26.80, 26.41, 25.52, 25.36, 19.43, 19.17, 13.79, 0.15. TBDPS O TMS O CN IV-95 O TMS DIBAL-H (3.0 equiv) DCM, -78 °C 71% TBDPS O TMS O IV-96 O TMS O Synthesis of aldehyde IV-96: To the nitrile IV-95 (55.0 mg, 0.1 mmol) in dry DCM (3 mL) was added DIBAL-H (1.0 M in hexane, 0.3 mL, 0.3 mmol, 3.0 equiv) at –78 °C. TLC indicated completion of the reaction within 30 min. Ethyl acetate (0.3 mL) was added to 359 the reaction followed by addition of saturated potassium sodium tartrate solution (3.0 mL). The reaction was stirred vigorously at room temperature for 2 h. The aqueous layer was extracted with DCM (3 x 5 mL), and the combined organic extracts were washed with brine, dried over sodium sulfate and concentrated under reduced pressure. The crude aldehyde was purified via column chromatography (silica, ethyl acetate/hexanes 3%) yielding the pure aldehyde (40.0 mg, 71%) as a clear oil. 1H NMR (500 MHz, CDCl3) δ 9.70 (q, J = 2.2 Hz, 1H), 7.74 – 7.60 (m, 4H), 7.44 – 7.29 (m, 6H), 5.74 – 5.60 (m, 1H), 5.49 – 5.31 (m, 1H), 4.23 (d, J = 6.1 Hz, 2H), 4.09 (dtd, J = 8.5, 4.3, 2.0 Hz, 1H), 3.49 (dq, J = 5.7, 3.1, 2.4 Hz, 1H), 2.57 (ddt, J = 16.0, 4.4, 2.0 Hz, 1H), 2.36 (ddt, J = 16.1, 8.2, 2.4 Hz, 1H), 2.20 (t, J = 10.9 Hz, 1H), 1.85 (dt, J = 15.6, 8.2 Hz, 1H), 1.02 (s, 9H), 0.03 (s, 18H). 13C NMR (125 MHz, CDCl3) δ 201.49, 135.56, 133.78, 131.26, 129.57, 127.63, 127.33, 74.61, 70.17, 60.37, 45.78, 29.02, 26.80, 19.16, 0.30, 0.23. TBDPS O TMS O CN IV-95 O TMS TBCO (1.2 equiv) MeOH (10 equiv) DCM then HCl (cat.), MeOH 87%, >95:5 dr Br TBDPSO OH H O H IV-90 CN Synthesis of IV-99: To the nitrile IV-95 (1.2 g, 2.1 mmol) in dry DCM (30 mL) were added TBCO (969 mg, 2.37 mmol, 1.1 equiv) and methanol (0.86 mL, 10.0 equiv) at room temperature. After stirring at this temperature for 2 h, TLC indicated completion of the reaction. The volatiles were removed under reduced pressure. The residue was dissolved 360 in methanol (50 mL). To this solution at room temperature was added HCl (conc. 2 drops, ~ 50 µL). After 5 min, solvent was removed under reduced pressure and the crude was purified via column chromatography (silica, ethyl acetate/hexanes 20%) to yield the pure product IV-90 (900 mg, 88%) as a clear oil. 1H NMR (500 MHz, CDCl3) δ 7.64 (tdd, J = 7.8, 2.3, 1.1 Hz, 4H), 7.46 – 7.33 (m, 6H), 4.35 (ddd, J = 8.9, 5.9, 2.8 Hz, 1H), 4.29 (td, J = 7.6, 7.2, 3.3 Hz, 1H), 4.11 – 4.04 (m, 1H), 4.03 – 3.95 (m, 2H), 3.92 (tdd, J = 6.6, 3.5, 0.8 Hz, 1H), 2.74 – 2.63 (m, 2H), 2.47 (ddd, J = 15.0, 9.0, 6.5 Hz, 1H), 2.36 (d, J = 9.8 Hz, 1H), 1.97 (ddd, J = 14.5, 6.0, 1.8 Hz, 1H), 1.05 (d, J = 0.8 Hz, 9H). 13C NMR (125 MHz, CDCl3) δ 135.60, 135.52, 132.89, 132.68, 129.96, 127.84, 117.57, 78.52, 76.05, 71.81, 65.04, 57.77, 38.72, 26.81, 19.29, 18.04. TOF MS ES+ (C24H31NO3SiBr): Calc. [M + H]+: 488.1257, Found [M + H]+: 488.1247. Br TBDPSO OH H O H IV-90 CN DHP (1.2 equiv) PPTS (cat.) 91%, 1:1 dr DCM Br TBDPSO OTHP O H H IV-100 CN Synthesis of IV-109: To a solution of alcohol IV-90 (100 mg, 0.2 mmol) in dry DCM (10 mL) at 0 °C were added 3,4-dihydro-2H-pyran (DHP, 1.1 equiv, 4.4 mmol, 21 µL) and PPTS (5 mg). The reaction was stirred for 12 h while allowing it to warm up to room temperature. Solvent was removed under reduced pressure, and the crude product was purified via column chromatography (silica, ethyl acetate/ hexanes = 10%) to yield to the 361 inseparable diastereomeric mixture of IV-100 (104 mg, 85% combined yield of two diastereomers). The following NMR data are for both diastereomers: 1H NMR (500 MHz, CDCl3) δ 7.64 (tt, J = 8.3, 1.6 Hz, 8H), 7.49 – 7.33 (m, 12H), 4.65 (dd, J = 4.3, 2.7 Hz, 1H), 4.56 (t, J = 3.6 Hz, 1H), 4.49 (q, J = 6.1 Hz, 1H), 4.41 (q, J = 6.5 Hz, 1H), 4.29 – 4.16 (m, 4H), 4.05 (p, J = 5.2 Hz, 2H), 3.98 (ddd, J = 11.0, 8.8, 6.5 Hz, 2H), 3.90 (ddd, J = 19.2, 10.9, 5.4 Hz, 2H), 3.80 (dddd, J = 19.3, 11.4, 7.9, 2.9 Hz, 2H), 3.49 (dt, J = 11.0, 5.5 Hz, 2H), 2.75 (dd, J = 16.6, 6.2 Hz, 1H), 2.68 – 2.55 (m, 3H), 2.34 (dt, J = 13.3, 6.7 Hz, 1H), 2.22 (dt, J = 13.3, 6.7 Hz, 1H), 2.10 (ddd, J = 13.2, 9.0, 6.5 Hz, 1H), 1.97 (ddd, J = 13.7, 8.5, 5.7 Hz, 1H), 1.80 – 1.65 (m, 4H), 1.65 – 1.43 (m, 10H), 1.04 (s, 18H). 13C NMR (125 MHz, CDCl3) δ 135.62, 135.61, 135.54, 133.02, 132.83, 132.79, 129.88, 129.86, 127.79, 127.78, 127.75, 118.14, 117.69, 100.57, 96.69, 77.87, 77.12, 76.56, 75.97, 65.36, 65.28, 62.81, 56.68, 56.41, 36.66, 34.42, 30.44, 30.39, 26.78, 25.26, 25.21, 19.51, 19.37, 19.33, 19.29 362 REFERENCES 363 REFERENCES 1. 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