"Hundreds of thousands of specialized metabolites are estimated to be produced and accumulated within the plant kingdom, many of which are considered potentially valuable natural compounds that can be used by humans. One of the greatest challenges for specialized metabolism analysis is the tremendous chemical diversity of such phytochemicals observed within and across plant species. To better address the diversification of plant specialized metabolites produced by glandular trichomes in the family Solanaceae, as well as understand the genetic and biochemical mechanisms underlying it, several mass spectrometry-based analytical platforms coupled with chromatographic separations, NMR, and stable isotope labeling have been employed in this study. In chapter 2, the diversity of acylsugar metabolites identified from three petunia species was explored as a complementary research to the previous analysis of acylsugar within Solanaceae family using LC-MS based strategies. 28 novel acylsucorses, including a class of malonyl ester acylsucroses, have been successfully purified through solid phase extraction followed by HPLC separation, and structures of them are proposed according to UHPLC-MS and NMR analysis. Interestingly, all acylsucroses characterized from petunia species have not previously been detected from studies of other Solanum species including petunia hybrids. This work extended our knowledge of the acylsugar diversity within Solanaceae family, and can be served as references in studying the bioactive of genes and enzymes involved in their biosynthesis. To fulfill the need for rapid characterization of plant chemical diversity and improve the efficacy of metabolite identification, a UHPLC-MS/MS coupled with stable isotopic labeling strategy was established and summarized in chapter 3. [13C]-Branched chain amino acids (BCAAs) were introduced as isotopic tracers to label and distinguish isomeric plant metabolites with diverse structures, particularly in respect to the lengths and branching type of aliphatic chains derived from metabolism of BCAAs. Metabolites tagged with stable isotopes were annotated easily according to their corresponded mass shift from unlabeled species using UHPLC-MS with both multiplexed CID and MS/MS analysis. This study provides a fast and simplified method to investigate the structural diversity of plant specialized metabolites without purification and NMR. Another question addressed in the study is how do plants make the huge diversity of acylsugar metabolites. Chapter 4 described the in vivo investigation of acylsugar related acyl chain elongation mechanism using stable isotope labeling coupled with UHPLC-MS/MS methodology. Fragments of acyl chain generated from acylsugars with and without stable isotope labels derived from [ 13C]-BCAA feeding of plants were analyzed using LC-MS/MS, and related aliphatic chain elongation pathways were proposed based on the resulting information. Evidence was observed to support that BCAA derived one-carbon elongation pathway existed in both wild tomato LA2560 and cultivated tomato, while two-carbon elongation was identified in S. pennellii LA0716 and LA2560."--Pages ii-iii.
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