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Micronutrient Testing of Greenhouse Media By Robert D. Berghage A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Horticulture 1986 ABSTRACT Micronutrient Testing of Greenhouse Media By Robert D. Berghage Accurate fertility control is essential for plant growth in soil-less plant growth substrates. A modified saturated media testing procedure was developed to permit testing for N03 - N, P, K, Mg, Ca, Fe, B, Mn, Cu, Zn, pH, and soluble salts with a single extraction. The procedure was based on adding a fixed equivalent weight of DTPA per unit volume of media sample prior to saturation with deionized water (30 ml of 0.005M DTPA per 1000c) . The use of DTPA greatly enhanced the extraction of Zn, Fe, Mn, and Cu, produced minor changes in Ca, Mg, P, and pH, and did not cause significant changes in N03 — N, K, B, or soluble salts. Competitive ligand - nutrient ion equilibria within the media extractant system will be discussed. Media test results were correlated with plant tissue tests of Chrysanthemum, marigold, and poinsettia in 3 media. Recom- mended media test micronutrient levels will be presented for each crop. The test procedure provides a quick, easy, and reliable method of testing both the macronutrient and micronutrient status of a wide range of growth media. ACKNOWLEDGEMENTS I would like to thank all the individuals who helped me in the preparation of this thesis, particularly my wife Barbara without whose help and patient understanding com— pleting this project would not have been possible. I would also like to thank the Gloeckner Foundation and W. R. Grace Co. for their financial support and Paul Ecke Poinsettias and Yoder Brothers for donating plant material. ii TABLE OF CONTENTS LIST OF TABLES ..................................... vi LIST OF FIGURES .................................... ix LITERATURE REVIEW Introduction ....................................... 1 Growth Media ..................................... 1 Basic plant nutrition ............................ 6 Nitrogen ....................................... 7 Phosphorus ..................................... 7 Potassium ...................................... 8 Sulfur ......................................... 8 Calcium ........................................ 8 Magnesium ...................................... 8 Iron ........................................... 9 Manganese ...................................... 10 Zinc ........................................... 10 Boron .......................................... 11 Copper ......................................... 11 Molybdenum ..................................... ll Chlorine & Nickel .............................. 11 Nutritional testing .............................. 12 Media/soil Testing ................................. 13 Principles ....................................... 13 pH and Soluble Salts testing ..................... 14 Macronutrient Testing ............................ 17 Micronutrient Testing ............................ 25 Plant Tissue Analysis .............................. 29 Principles...... ............. . ................... 29 Tissue analysis standards ........................ 34 Poinsettia .................................... 34 Chrysanthemum ................................. 34 Marigold ...................................... 35 DTPA Chelate Chemistry ............................. 37 Plasma Emission Spectroscopy ....................... 38 References ................ . ....................... 41 SECTION I Micronutrient Testing of Plant Growth Media: Extractant Evaluation Abstract ........................................... 48 Introduction ....................................... 49 iii Materials and Methods .............................. 51 Extractants............ ....... . .................. 51 Media preparation ................................ 51 Experimental design and statistical analysis ..... 53 Extractant evaluation .......... . ................. 53 Results and Discussion ............................. 54 References ......................................... 57 SECTION II Micronutrient Testing in Greenhouse Media: Extractant Characterization Abstract ........................................... 82 Introduction ....................................... 83 Materials and Methods .............................. 84 Results and Discussion ............................. 88 Summary ............................................ 96 References ......................................... 98 SECTION III Micronutrient Testing of Plant Growth Media: DTPA Modified Saturated Media Extraction Procedure Calibration and Correlation with Plant Growth and Plant Nutrient Uptake Abstract ........................................... 141 Introduction ....................................... 142 Materials and Methods .............................. 142 Media ............................................ 142 Media extractions ................................ 143 Tissue analysis. ................................ . 143 Experiment 1. Experiment 2. Correlation between media levels of Zn, Fe, Mn, B and Chrysanthemum morifoljum cv. Bright Golden Anne nutrient uptake and growth... 144 Correlation and calibration of media levels of Zn, Fe, Mn, B and Marigold (Tagetes patula cvs. Yellow boy and Inca Yellow) nutrient uptake and growth ........ 146 iv Experiment 3. Correlation and calibration of media levels of Zn, Fe, Mn, and B and Poinsettia (Euphorbia pulcherima Willd. Cvs. Annete Hegg Dark Red and Eckespoint C-l) nutrient uptake and growth ................... ..... 148 Regression analysis .............................. 150 Results and Discussion.... ............... . ......... 150 Summary ............................................ 158 References cited ................................... 159 Table LIST OF TABLES LITERATURE REVIEW Commercially available media and their components. Soluble salt guidelines for greenhouse growth media using various media to water ratios ......... Leaf analysis values for Poinsettia (Mastalerz 1977)............ ......... .... ......... Nutrient ranges for Chrysanthemum leaf tissue (Criley and Carlson 1970)................ ......... Nutrient tissue levels of marigold grown in sand culture (from Johnson 1973) .................. Standard operating conditions of the MSU plasma emission spectrometer (information provided by Beckman instrument company) .......... SECTION I Micronutrient concentration in media amendment solutions...... ..... , ........... . .......... . ....... Coefficients of variation of saturated media test results for a 1:1 peat to vermiculite media ammended with 1.2 Kg superphosphate, and 2.97 Kg of gypsum and dolomitic lime per cubic yard of media, drenched with 24 ppm Fe, 34 ppm Mn, 30 ppm Zn and 4.5 ppm B, extracted using 15 extractants.. SECTION II Variables examined using center composite experimental design and the levels used ........... Media amendment rates used in center composite experiments (ppm in drench)..... .......... . ....... Nutrient amendment rates used to examine the relationships between DTPA extractions and stand— ard water extractions (ppm of ion in drench)...... vi Page 16 35 36 36 40 52 59 85 85 87 'Table 4. Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concentration, and media moisture level, with a Cornell mix medium. Values presented are the significance levels of the (t) tests on the various least squares regression coefficients ..... Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concentration, and media moisture level, with a pine bark medium. Values presented are the sig- nificance levels of the (t) tests on the various least squares regression coefficients ............ Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concen- tration, and media moisture level, with a soil based medium. Values presented are the sig- nificance levels of the (t) tests on the various least squares regression coefficients ............. Coefficients of determination for regression equations using mg of DTPA/100cc media rather than DTPA concentration and initial media mois- ture level with data obtained from center com— posite experiments examining the effects of media amendment level, DTPA extractant concentration, and media moisture level........... ......... . ..... Conversion equations to convert from standard saturation extract test results to modified DTPA saturation extract test results ................... SECTION III Experiment 1. Continuous liquid feed rates ........ Experiment 1. "STEM" Application rates. ........... Experiment 2. Media Drench Application Rates ...... Experiment 3. Media Drench Application Rates ...... vii Page 100 102 104 145 145 147 149 Table 5. 10. 11. 12. Analysis of variance for Chrysanthemum tissue / DTPA media test correlation experiments. Trial 1 Cornell mix with CLF micronutrients, trial 2 pine/peat media CLF micronutrient applications and trial 3 pine/peat with Peters "STEM" micro- nutrient applications....... ...................... Correlation equations between Chrysanthemum tissue tests and DTPA saturated media extracts. Trial 1 Cornell mix with CLF micronutrients, trial 2 pine/peat media CLF micronutrient ap— plications and trial 3 pine/peat with Peters "STEM" micronutrient applications. ......... . ..... Correlation equations for relationships between marigold tissue tests, DTPA saturation extracts, and micronutrient drench rates for 3 media and 2 cultivars with parallel regression intercept ad— justments when needed for media or cultivar ....... Analysis of variance of split split plot experiment examining the relationships between marigold tissue tests and DTPA saturated media extracts of 3 media ............................... Correlations between poinsettia tissue samples and DTPA saturated extracts in 3 media with 2 cultivars. Intercept adjustments provided for parallel regressions where needed ................. Correlations between poinsettia tissue samples and DTPA saturated extracts in 3 media with 2 cultivars. Intercept adjustments provided for parallel regressions where needed ................. Analysis of variance of plant tissue and DTPA saturated media extract test results from a split plot experiment with 3 media (M), 2 poinsettia cultivars (C), 2 sampling times (T), and 13 micronutrient treatments (TR) ............. . ....... Recommendations for B, Zn, Fe, and Mn DTPA saturated media extract test values for Chrysanthemum, marigold and poinsettia ............ viii Page 161 162 163 164 165 166 167 LIST OF FIGURES Figure Page LITERATURE REVIEW 1. Schematic of the effects of nutrient supply on plant nutrient content and crop yield (Brown 1970) ........................................... 33 2. Schematic describing the relationship between nutrient concentration and crop yield (Ulrich & Hills, 1967)..... ............ ..... ..... .... ..... 33 3. Schematic representing the relationship between crop yield and nutrient concentration (Chapman 1967) ........................................... 33 SECTION I 1. Phosphorus extracted from a 1:1 peat to ver- miculite media using saturated media extraction techniques with 15 different extractants ........ 61 2. Potassium extracted from a 1:1 peat to vermicu- lite media using saturated media extraction techniques with 15 different extractants... ..... 63 3. Calcium extracted from a 1:1 peat to vermiculite media using saturated media extraction tech- niques with 15 different extractants ............ 65 4. Magnesium extracted from a 1:1 peat to vermicu— lite media using saturated media extraction techniques with 15 different extractants... ..... 67 5. pH of saturated media extracts of a 1:1 peat to vermiculite media extracted with 15 different extractants .......... . .......................... 69 6. Soluble salts of saturated media extracts of a 1:1 peat to vermiculite media extracted with 15 different extractants... ..... .............. ..... 71 7. Iron extracted from a 1:1 peat to vermiculite media amended with (1) Oppm Fe, (2) 24ppm Fe, and (3) 96ppm Fe extracted using saturated media methods with 15 different extractants........... 73 8. Manganese extracted from a 1:1 peat to vermicu— lite media amended with (1) Oppm Mn, (2) 34ppm Mn, and (3) 136ppm Mn extracted using saturated media methods with 15 different extractants..... 75 ix .Figure Page 9. Zinc extracted from a 1:1 peat to vermiculite media amended with (l) Oppm Zn, (2) 30ppm Zn, and (3) 120ppm Zn extracted using saturated media methods with 15 different extractants ..... 77 10. Boron extracted from a 1:1 peat to vermiculite media amended with (l) Oppm B, (2) 4.5ppm B, and (3) 18ppm B extracted using saturated media methods with 15 different extractants ........... 79 11. Copper extracted from a 1:1 peat to vermiculite media using saturated media extraction tech- niques with 15 different extractants ............ 81 SECTION II 1. Correlation between pH of standard water satur- ated extracts and DTPA modified extractions in 3 media with pH adjusted by addition of either 15 ml 0.2 N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction ...................................... 106 2. Calcium extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 m1 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction.. 108 3. Boron extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 ml 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction ...................................... 110 4. Copper extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 m1 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction.. 112 5. Zinc extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 ml 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction.. 114 Figure Page 6. Iron extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 m1 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction.. 116 7. Manganese extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 ml 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction.. 118 8. Correlation between soluble salts of water saturated extracts and modified DTPA extracts of 3 media drenched with increasing rates of nutrients prior to extraction. r2: .974 ......... 120 9. Correlation between nitrate concentration in water saturated extracts and modified DTPA ex- tracts of 3 media drenched with increasing rates of nutrients prior to extraction. r2: .964 ...... 122 10. Correlation between Ca concentration in water saturated extracts and modified DTPA extracts of 3 media drenched with increasing rates of nutrients prior to extraction. r2: .913 ......... 124 ll. Correlation between Mg concentration in water saturated extracts and modified DTPA extracts of 3 media drenched with increasing rates of nutrients prior to extraction. r2: .956 ......... 126 12. Correlation between K concentration in water saturated extracts and modified DTPA extracts of 3 media drenched with increasing rates of nutrients prior to extraction. r2: .984 ......... 128 13. Correlation between P concentration in water saturated extracts and modified DTPA extracts of 3 media drenched with increasing rates of nutrients prior to extraction ................... 130 14. Dose response curves for B in DTPA modified saturated extracts of 3 media drenched with in— creasing rates of B prior to extraction ......... 132 15. Dose response curves for Zn in DTPA modified saturated extracts of 3 media drenched with in— creasing rates of Zn prior to extraction ........ 134 xi Figure 16. 17. 18. Dose response curves for Cu in DTPA modified saturated extracts of 3 media drenched with in- creasing rates of Cu prior to extraction........ Dose response curves for Fe in DTPA modified saturated extracts of 3 media drenched with in— creasing rates of Fe prior to extraction.. ...... Dose response curves for Mn in standard water saturated extracts and DTPA modified extracts of 3 media drenched with increasing rates of Mn prior to extraction ............................. SECTION III Iron calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight ................... Zinc calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight ........ . .......... Boron calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight ................... Iron calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight .................... Zinc calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight ............... ..... Manganese calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight .................... xii Page 136 138 140 169 171 173 175 177 179 Figure Boron calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight .................... xiii Page 181 L ITERATURE REVIEW LITERATURE REVIEW Introduction Growth Media Plants have very few absolute requirements for their growth: light, water, appropriate temperatures and a supply of mineral and gaseous nutrients are essential. The producer of plants in a greenhouse is in a unique position to control these factors to produce high quality plants. However in order to accomplish this routinely the grower must be able to provide a uniform substrate with reproducible physical and chemical characteristics. Physically a growth media should provide adequate sup— port for the "above ground" portion of the plant, and yet be fairly light weight to allow for ease in handling. It should be porous enough to permit gas exchange, and yet provide an adequate supply of water. -It should be rela- tively stable over time (Spomer 1979). Mastalerz (1977) lists the following as objectives in preparing a growing medium. 1. porous and well drained, yet retentive of suf- . ficient moisture to meet the water requirements of plants between irrigations; 2. relatively low in soluble salts, but with an adequate exchange capacity to retain and supply the elements necessary for plant growth; 3. standardized and uniform with each batch to permit the use of standardized fertilization and irrigation programs for each successive crop; 4. free from harmful soil pests-pathogenic organisms, soil insects, nematodes, and weed seeds; 5. biologically and chemically stable following pasturization; primarily free from organic matter 1 2 that releases ammonia when it is subject to heat or chemical treatment. The first growth media was probably field soil. Be cause of the physical constraints of growing plants in con- tainers (primarily, small volume, and hence poor root water and air relations (Spomer 1979)) the soil was amended with a variety of "composts"; including peat, leafmould and pine needles (Bunt 1976). The first attempt to standardize grow- ing media was the development of the John Innes Composts (Lawrence & Newell 1939). They developed two composts a "seed compost" and a "potting compost". The composts were steam sterilized to eliminate disease problems, provided fertilizer recommendations based on a weight of fertilizer per volume of media and utilized an organic "compost" that was relatively clean (free from pathogens) and standardized (peat or leaf-mould). These potting media were found to be suitable for use with a wide range of plant species. The primary component of the Innes media was a "loam", "by loam we mean a soil in which the sand, silt, and clay are nicely balanced with some humus as well."(Lawrence & Newell 1939). It has become increasingly difficult and costly to obtain sufficient quantities of soil for use in greenhouse media. Many growers have changed from using soil based media to using "artificial" media or "loamless composts". Bunt (1976) provides the following comparison of loam-based and soilless media. 3 Loam Composts Advantages . l. The principal advantage is ease of plant nutrition. Composts made from a good loam have more plant nutrients and the nutrition, especially with respect to nitrogen and phosphorus, is easier. 2. Minor element deficiencies are not common. Disadvantages 1. Difficulty in obtaining suitable loam that does not give toxicities when steam sterilized. 2. Continuity of supply and control. 3. The loam must be stored dry and steam steril— ized before it is used. 4. The composts are heavy and difficult to handle. 5. They are more expensive to prepare if done properly. Loamless Composts Advantages 1. A greater degree of standardization of materials, less variability between successive batches of compost. 2. Do not require steam sterilization. 3. Cheaper to prepare. 4. Lighter to handle. 5. The lower nutrient content of the materials can be used to give more controlled growth. Disadvantages 1. Control of the supply of nitrogen, phosphorus, and minor elements such as boron and copper is more critical. 2. Greater dependence on liquid feeding. 3. Lack of general "buffer" capacity, i.e. they are more likely to show rapid changes in general nutrient levels. A large variety of materials have been used or tried in the production of soil-less growth media. One the most common components is peat (Sheard 1975). There are 4 basic classifications of peat, Sphagnum moss peat, Sphagnum moss peat is the most popular for naceous moss, reed and sedge peats, and humus peat or muck. greenhouse media (Mastalerz 1977). The U.C. (University of use in preparing 4 California) system uses mixtures of peat and sand (Baker 1957). There are 5 basic U.C. mixes which vary in their relative percentages of peat moss and sand. The U.C. mix C for example contains 50% sand and 50% peat. Cornell mixes (peat-lite) are composed of sphagnum peat and vermiculite or perlite (Boodley & Sheldrake 1963). The basic Cornell mix is composed of 50% peat and 50% vermiculite or perlite. The Cornell epiphyte mix is 1/3 by volume fir bark in addi- tion to 1/3 sphagnum peat and 1/3 perlite. In the Nether- lands the RHPA (Regeling Handelspotgrand Proefstation Aalsmeer) recommends the use of a standardized potting mix containing 10 parts sphagnum peat, 10 parts by volume frosted decomposed black sphagnum peat, and 1 part by volume river sand (Bunt 1976). Composted hardwood and softwood bark have gained wide acceptance as an organic component for use in growth media (Verdonck et a1 1983). It is often used as a substitute for part or all of the peat in a growth media. Plant roots penetrate into the internal pore space of pine bark particles, thus water and nutrients contained within these pores are available for plant use (Pokorny et a1 1984). Bark must be composted before use to allow the biological breakdown of toxic compounds and to help limit N depletion (Bunt 1976). It can be composted with a number of organic waste products, such as chicken manure and sewage sludge, which in and of themselves could not be used because of toxic accumulations of metals and soluble salts. Bark 5 sludge mixtures resulted in better plant growth than pure bark in a tomato yield trial (Verdonck et a1 1983). Coal Cinders have also been tested as an amendment to bark media. Their high micronutrient content makes their usefulness questionable (Neal & Wagner 1983). Sawdust is also widely used as a media component. Depletion of available nitrogen resulting from microbial activity is a major problem. Nitrogen must be added or the celulose content in the sawdust reduced to compensate for this nitrogen depletion. Plant growth in media amended with sawdust that has had the celulose content reduced by the Fersolin process was "consistently excellent" in Chrysan— themum trials (Mastalerz 1977). Other organic materials used as media components in- clude cofuna, sewage waste, corn cobs, bagasse, peanut hulls, rice hulls, spent mushroom composts, and straw. (Bunt 1976, Mastalerz 1977). Inorganic components of media include sand, perlite, vermiculite, calcined clays, Cinders, scoria and pumice, polystyrene, and urea formal- dehyde foams (Mastalerz 1977). Commercially available media contain a wide variety of components. A list of commercial media and their components is presented in table 1 (adapted from Sanderson 1983). Table l Commercially available media and their components Baccto - Horticultural grade vermiculite, granular Michigan peat Ball Growing Mix I - Composted pine bark,horticultura1 grade vermiculite, perlite, liquid nutrient starter solution (macro and micronutrients, dolomitic limestone, and a wetting agent. Ball Growing Mix II - Composted pine bark, horticultural grade vermiculite, sphagnum peat moss, perlite, liquid nutrient starter solu- tion (macro and micronutrients), dolomitic limestone, and a wetting agent. Jiffy Mix - Sphagnum peat moss, horticultural grade vermiculite, nutrient starter (macro and micronutrients), and a wetting agent. Metro-Mix 200 - Sphagnum peat moss, horticultural grade vermiculite, perlite, granite sand. Metro-mix 300 - Sphagnum peat moss, horticultural grade vermiculite, perlite, granite sand, composted pine bark. Metro-Mix 500 - Sphagnum peat moss, horticultural grade vermiculite, granite sand, processed bark, composted pine bark. Nature-Life Potting Soil — Peat moss, perlite, composted cow manure, earthworm castings. Wonder Gro - Pine bark, Sphagnum and sedge peat moss, sand, perlite, dolomitic lime. Basic plant nutrition There are 17 elements that have been established as es- sential for plant growth and reproduction. These elements can be divided into two groups: macronutrients and micronutrients, based on the amount of the nutrient that is required by the plant. The macronutrients include C, H, O, N, P, K, Ca, Mg, and S. C, H, and O are provided by air and water, and N, P, K, Ca, Mg, K, S and the micronutrients Zn, Fe, Mn, B, Cu, Mo, and Ni generally must be absorbed from the media by plant roots. Nitrogen There is a vast store of nitrogen in nature, unfortunately only a relatively small amount is available for use by higher plants. Higher plants absorb N as N03 (the primary source) or NH4. Both forms of N are required (Joiner 1983). In greenhouse plant production most N is provided in the form of inorganic chemicals; potassium nitrate, ammonium nitrate and calcium nitrate are examples. The negatively charged nitrate ion is not held by cation ex- change complex in the media, thus nitrate is subject to rapid leaching. While ammonium may be held by the cation exchange sites of the media, high concentrations of am- monium ion are toxic to most plant species (Mengel & Kirkby 1982). Phosphorus. The reactions which govern P solubility in soils are very complex. Phosphorus can combine with a num— ber of ions i.e. Ca, which if present in sufficient quan- tities can control the solubility of P (Lindsay 1979). As a result P is rapidly tied up in a soil, and is relatively immobile and resistant to leaching. This is true also for a soil based media. Artificial media are considerably less resistant to leaching of P, decreased exchange capacity results in a lower P buffering capacity as demonstrated by P adsorption isotherms (Marconi & Nelson 1983). Phosphorus is usually supplied as single or triple 8 superphosphates incorporated into the media. Potassium. Potassium fertilization in growth media is usually achieved with potassium chloride, potassium sulfate or potassium nitrate. These inorganic chemical sources are very soluble, and thus can be leached from the media. Ver- miculite contains an appreciable amount of potassium and can act to some extent as a potassium source when it is used in a growth media (Sanderson 1983). Sulfur. Sulfur is seldom deficient in greenhouse media because it is present as a carrier or contaminant in many of the fertilizers used, for example magnesium sulfate and superphosphate (Robertson et al 1976). Calcium. Calcium is usually supplied in a growth medium as lime (dolomitic or calcitic), and other compounds added to help control pH. Superphosphate can supply a large amount of Ca when it is used as a P source (Robertson et al 1976). Gypsum (calcium sulfate) is used when an increase in pH is not desirable (Warncke & Krauskopf 1983). Most of the "liming" materials dissolve slowly and are only slightly soluble. These materials must be incorporated into the growth medium. Magnesium. Deficiencies of Mg are common in floricul— tural production where dolomitic lime is not used in the pH control program. Magnesium is not usually supplied in the fertilization program and is not present in appreciable quantities in most components used to make greenhouse growth media. Vermiculite contains Mg and may provide 9 appreciable amounts when used in a medium (Warncke & Kraus- kopf 1983). Most fertilizer sources of Mg are soluble and readily leached from the media. Dolomitic lime is relatively insoluble and can thus act as a source of Mg in a growth medium for an extended period. lggg. In well oxidized soils iron solubility is con- trolled by Fe (III) oxides. Solubility of inorganic iron is a function of pH, solubility is highest at low pH and decreases as pH increases. At higher pH levels Fe 3+ in solution decreases 1000 times for each unit increase in pH (Lindsay 1979). Plants absorb iron as Fe 2+ or as chelates. The amount of inorganic iron the plant root can absorb may be a function of the plants ability to lower pH (to increase solubility) and reduce Fe 3+ to Fe 2+ (Chen & Barak 1982). The availability of iron to the plant from chelates is less dependent on the pH of the media. The chelates themselves are soluble, and pH affects are a function of the relative stability of the ligand metal complex. Very little of the chelate is actually absorbed by the plant root. Chelate splitting (reduction of Fe (III) to Fe (II) and separation of the Fe from the chelate) must occur for appreciable quantities of chelate Fe to be absorbed (Mengel & Kirkby 1982). Deficiencies of iron can occur as a result of a physi- cal lack of sufficient iron in the media or may be induced with apparently sufficient levels of iron when pH is elevated or where there are high levels of heavy metals in 10 the media such as Mn. The absolute level of Fe found in plant tissues does not correspond well to the presence or absence of Fe deficiency symptoms. It is thought that the Fe in the plant can be present in available and unavailable forms (Chen & Barak 1982). Mn induced iron deficiency is thought to be due to Mn interfering with the enzymatic ac- tivity of Fe possibly by competing for binding sites within the plant (Mengel & Kirkby 1982). The most effective sources of fertilizer iron are iron chelates. Of these EDDHA is most effective. Manganese. Manganese exists in a 3 oxidation states, in soils. Plants are only able to absorb the Mn2+ form. The availability of this form is pH dependent, Mn2+ solubility decreases 100x for each unit rise in pH (Lindsey 1979). Manganese is relatively immobile and non-leachable in soils and growth media. It forms complexes with organic matter that can reduce its availability at higher pH. Steam sterilization can increase Mn solubility by killing off Mn oxidizing microorganisms resulting in toxic levels of Mn in the medium (Mastalerz 1977). The most effective form of Mn fertilizer is manganese sulfate. Zipg. Zinc readily forms complexes with organic matter, it is thus relatively immobile and non-leachable in growth media. The availability of Zn in soils is influenced by pH; availability is greatest in acid soils and is reduced as pH increases. Zn forms both soluble and insoluble organic complexes. The soluble complexes are associated with amino 11 and fulvic acids. The non-soluble forms are derived from humic acids (Mengel & Kirkby 1982). Thus as organic material in a media decomposes and undergoes humification Zn may become less available. Zn deficiencies are common on many Michigan muck soils (Vitosh et a1 1981). The most com— mon fertilizer form of zinc is zinc sulfate. Boron. Boron exists in media and soils as the undis- sociated acid (H3B03). It is thought that this is the form that is absorbed by plants. Boron is adsorbed to organic matter through ligand exchange, where OH“ on the surface of the media particle is replaced by B(OH4‘ ). The carboxylic acid of humic acid may condense with B to provide another source of B exchange (Mengel & Kirkby 1982). Boron adsorp- tion to media is pH dependent, as the pH increases so does B adsorption. Boron availability to the plant is thus greater at lower pH. Boron is mobile in media and may be leached. Copper. Copper is bound very strongly to organic matter and other exchange sites in media. It is thus immobile and is non-leachable. Cu2+ is the predominant form of copper in the soil, it is also the form that plants can absorb (Mengel & Kirkby 1982). Molybdenum. Molybdenum is required by plants in minute amounts. It is present in the soil as the M0024+ ion. Mo availability is dependent on pH; as pH increases so does Mo availability (Joiner 1983). Chlorine & Nickel. These elements are required in very small amounts. Natural chlorine deficiencies are not known 12 to occur because chlorine is ubiquitous in nature. Nickel requirement was established in 1984 by Eskew and Cary. Nutritional Testing The nutritional status of a plant is dependent on a large number of factors including not only the nutrients in the growing media but also the plant’s environment, both past and present; the plant’s age, both chronological and physiological; and the plant’s genetic makeup. The com- plexity of these relationships make the development of nutritional testing programs difficult (Munson & Nelson 1973). There are two basic approaches to plant nutritional testing. They are plant tissue analysis, and substrate (soil or media) analysis. Each offers certain advantages. Tissue analysis reflects the plants actual nutrient status at a point in time, deficiencies or toxicities may be determined. It does not provide any information on whether the cause of the problem is physiological (in the plant), pH related, or caused by an absolute deficiency or excess of the element in the substrate. Substrate testing on the other hand provides information on the absolute status of the substrate but no information on the plant’s response. The best testing program would thus be the use of a com- bination of these methods. In the real world however time and expense are often overriding factors which preclude the routine use of both testing methods. 13 Tissue testing is expensive and time consuming. The average turn around time for most laboratories is measured in weeks rather than days. Media samples turn around time can be measured in days and the cost less. Many greenhouse growers thus use media testing routinely and tissue testing only as a diagnostic tool, when problems develop. Media/soil Testing Principles. Of the factors affecting the results of any media or soil test procedure the quality of the sample is probably the most important and the least considered. If the sample is not representative of the medium in question any test results are of very limited value. There are many sampling plans that have been proposed and tested for use in the farm field (Peck & Melsted 1973). In bulk growth media that has been well mixed obtain- ing a representative sample is not usually a problem, however care must be taken to where settling of the media may have occurred. Once the media has been placed in con— tainers and subjected to cropping, defining a good sample becomes a problem. Each pot is, essentially, different. There is no guarantee that the pots have been or even can be treated exactly alike. Multiple sampling should be encouraged. In the development of a rapid chemical analysis l4 procedure for testing soils or media it is important to be aware of the various forms of a nutrient that may be present. A testing procedure should determine the levels of plant available nutrients, unavailable forms should not be included. The available nutrients may be either in the bulk solution (intensity factor) or in some form that supplies the bulk solution (capacity factor). In a growth media the capacity factor is predominantly the cation exchange of the media. In any case the level of a nutrient measured in a test should correlate to plant growth. Experiments should be conducted with any -potential testing procedure to establish the correlation between the media test results and the growth and nutrient uptake of plants. Once this relationship has been established it is necessary to calibrate the procedure. Calibration experi- ments provide the basis for recommendations based on test results. The ideal calibration experiments would provide measurements of all the factors that might affect plant yield. This would allow the development of multiple regres— sion equations and the optimization of the inputs that can be controlled (Hanaway 1973). pH and Soluble Salts testing Soluble salts and pH testing should be a routine proce- dure in any greenhouse operation. Routine pH testing in greenhouse media is done using a slurry of media and deionized water, usually on a 1:1 volume basis. Since there 15 is essentially no buffering capacity in deionized water the ratio of water to media has only limited effects- on the results of a pH test. Dilute solutions of calcium chloride or potassium chloride are sometimes used in soil pH tests. The pH measured in these ionic salt solutions can be easily converted to the pH value obtained with a water test (Van Lierop 1981). It is desirable to use a deionized water procedure for pH testing in a growth medium because soluble salts can be tested using the same slurry. Since the soluble salts test is a measure of the total electrical conductivity of a solution, which is dependent on ionic concentration, test results are highly dependent on the amount of water added to the slurry. Common media to water ratios are. 1' part media to 2 parts water (by volume), 1 part media to 5 parts water, and a saturation extract. The saturation extract is obtained by gradually adding distilled or deionized water to 3 approximately 400 cc of medium until the sample is just saturated (Warncke 1980). The following chart (table 2) allows comparison of the 3 methods (taken from Warncke & Krauskopf 1983). Solu-Bridge readings are given in millisiemens. The unit used to express soluble salts in the past has been the millimho. 1 millisiemen = l millimho. To obtain an ap— proximate conversion of this value to parts per million (ppm) multiply the solu—bridge reading by 700. 16 Table 2 Soluble salt guidelines for greenhouse growth media using various media to water ratios. Solu—bridge Reading Saturation 1 part media 1 part media to to Extract 2 parts water 5 parts water Comments mS 0-.74 0-.25 0-.12 Very low salt levels. Indi— cates very low nutrient status .75—l.99 .25—.75 .12-.35 Suitable range for seedling and salt sensitive plants 2.00—3.49.75-l.25 .35-.65 Desirable range for most es- tablished plants. Upper range may reduce growth of some sen- sitive plants. 3.50-5.001.25-1.75 .65—.90 Slightly higher than desirable. Loss of vigor in upper range. OK for high nutrient requiring plants. 5.00-6.001.75-2.25 .90-1.10 Reduced growth and vigor. Wilting and marginal leaf burn. 6.00+ 2.25+ 1.10+ Severe salt symptoms crop failure likely. l7 Macronutrient Testing. Due to the inherent differences, physical and chemical, between greenhouse growth media and field soils quick chemical test methods developed for field soils have proven unreliable when used on growth media. Perhaps the largest problem in using soil test procedures for media analysis is the very low bulk density of these substances. The major volume components of soil-less media are generally very low bulk density materials. In addition these components may have very high water holding capacities. In order to produce analyzable solutions the ratio of solution to medium must be increased over ratios used in soils. This, however, results in a reduction in the reproducibility of the results (White & Thomas 1975). Grinding and sieving the sample, a common practice in soil testing, causes dramatic changes in the chemical and physical properties of the growth media components and thus should not be performed. C. H. Spurway (1949) proposed using a dilute acetic acid extractant to test greenhouse soils for ammonia, nitrate, magnesium, iron, aluminum, manganese, sulfate, chloride and sodium. The basic Spurway testing procedure is as follows; 1 teaspoon of sample is placed in a test tube with 13 ml of deionized water. One drop of 25% by volume acetic acid is added and the tube is shaken for about 1 minute. The extract is filtered and chemical analysis is 18 done on the filtrate (Spurway 1949). Because of the small sample size the procedure is best suited for use with samples that contain a significant proportion by volume of soil. With artificial media such small samples tend to produce highly variable test results (White & Thomas 1975). Many modifications on the basic Spurway procedure have been tried. Extractant, sample to solution ratio and shak- ing time may be varied. Markus and Steckel (1980) used 100ml of 0.018 N acetic acid with 20ml of soil mix. Wilker- son and O’Rourke (1983) used 0.025 N acetic acid with a 1:4 media to extractant ratio for times ranging from 1 to 24 hours. Prasad et al (1983) and Holcomb and White (1979) and (1980) also utilized modified Spurway extractions. Since there are no comparative data available, evaluating the results of these studies is an exceedingly frustrating task. Many other extractants have been tested. Markus and Steckel (1980) used 6 testing methods in addition to the Spurway extractant in a’ study of testing methods for monitoring tomato nutrition in a peat—vermiculite substrate. The extractants used were the North Carolina double acid (0.05 N HCl and 0.025 N H2504), 0.5N NaCl, 1 N NH40AC, 1.4 N NaOAc + 1.0 N CH2COOH, Bray P1 (0.03 N NH4F + 0.025 N HCl), and a water saturated paste (wsp). All the extractants were compared to the North Carolina double acid technique. The double acid, NaCl, NHqOAc and NaOAc were in relatively close agreement for extraction of K, Ca, Mg, and NOs-N. The 19 double acid removed much more P than the other extractants. NaCl, NHqOAc, NaOAc and Spurway extractions for P were in close agreement with the Bray P1 extraction. The high levels of nutrients extracted by the strong extractants used in this study indicates that they strip nutrients from the media exchange sites and probably solubilize solid forms of the nutrients that may be present. This is especially true of the strong acid extractants which would be expected to dissolve any solid forms of Ca, Mg and P. Since these nutrients may be only slowly available to the plant as in the case of lime (CaC03) the acid extractants probably over- estimate their availability to the plant. Tissue levels of the nutrients in corresponding plant samples were not statistically correlated to media levels. However since nutrients were present in excess in all the samples this is not surprising. Short term (3—6 days) media storage was found to have no effect on, the N status of the media. Using similar extractants (H3303 was also included), Markus et a1 (1983), examined the recovery rate of slow release fertilizers in container grown azaleas. The highest recovery of N was obtained using the Spurway extractant. The highest recovery of P was obtained with Bray P1 extraction and the highest recovery of K was ob— tained with H3803. It is interesting to note that the ex- tractants that had the "highest recovery rate" were extract— ing large quantities of the nutrient 10 months after application. In most cases wsp samples contained elevated 20 nutrient levels. Since the wsp may be considered to be a measure of the bulk soil solution this indicates at least some portion of the nutrients present were in a plant avail- able form. There is however no evidence to indicate whether the remaining nutrients extracted with the various Spurway modifications are present in a readily available form, a slowly available form or in a plant unavailable form. Extraction times have a dramatic effect on the amount of nutrient extracted from a pinebark medium with double acid (0.05 N HCl + 0.0025 N H2804), NH40Ac, Bray P1, and .l N CH2COOH (Wilkerson & O’Rourke 1983). The relationship be- tween extraction time and the amount of nutrient extracted is second order quadratic. The amount of N, P, K, Ca, and Mg extracted by modified Spurway (.1 N CH2COOH), Ammonium acetate and am- monium floride/hydrochloric acid, and Intensity balance (saturation extract) are significantly correlated to plant uptake in Chrysanthemum (Holcomb & White 1980). Correla- tions for N, P, and K were roughly equivalent. The satura- tion extract provided substantially better correlations for Mg and Ca indicating that the intensity factor in the media was more closely related to plant uptake than the capacity factor for these nutrients. Optimum values for media test levels based on plant tissue test levels for Chrysanthemum were determined for these nutrients and extractants (Holcomb & White 1979). The optimum test levels determined correspond fairly well to 21 other published standards, except for the N level obtained with the intensity balance method (1667 ppm). The major difference between the intensity balance method as reported in this study and the saturated media extract, recommended by Warncke (1980), is that the intensity balance method uses an air dried sample on a weight basis, where as the saturation extract does not call for drying the media , and uses a volume measurement. This difference does not seem to provide an adequate explanation for the difference in recommended levels, 1667 vs. 100 - 199. There are a number of other media test procedures that use water as an extractant. The Leavington method (Johnson 1980) uses distilled water in a 1 part media to 6 parts water extraction. This method was compared to two other methods: the ADAS method where a 20ml sample of dried, ground and sieved medium is extracted with 50 ml of the ex- tractant normally used in field soils (i.e. calcium sulfate), and a "Dutch" method where 60 m1 samples of media moistened to pF 1.5 are extracted with 90 ml of distilled water (Johnson, 1980). All three methods had relatively low errors of determination. The Leavington method was chosen since it does not require that the media be dried before testing. This was considered to be a great labor savings where large numbers of samples were to be analyzed. The "Dutch" method (Sonneveld et al 1974) is often referred to as a 1 : 1.5 volume extract. This test method was found to be superior to a 1 part media to 25 parts 22 water by weight test on a wide variety of potting media (Sonneveld et al 1974). Nutrients extracted by the 1 : 1.5 extraction technique are highly correlated to the media solution obtained from a saturated media sample with a hydraulic press. The relationship between phosphate measured by the l : 1.5 extract and the media solution is best described by two separate regressions one for unfertilized peats and "clayey" soil based media, and another for ar— tificial media fertilized with soluble phosphate sources. This is probably due to dilution of soluble phosphates in the artificial media, and in the case of soil based media, replacement of ions in the solution by phosphate dissolved from solid phase soluble sources or removed from the ex— change capacity of the soil. Phosphate levels in the soil based media were much lower than in the artificial media (Sonneveld et a1 1974). The l : 1.5 media testing procedure has been corre- lated and calibrated for N, P, and K (Prasad et a1 1981). Correlation coefficients between N uptake and N media test levels varied with the different media tested, ranging from .908 for peat to .616 for bark. The combined regression had a determination coefficient of .546, in a trial using tomato as the test plant. Correlations were similar or better with Chrysanthemum or verbena as the test crop. Desirable values for N in the media were determined by regressing dry weight against soil test values. A quadratic function was used to determine the desirable values for different cropping 23 periods. Determination coefficients ranged from .848 to .458. Similar results were reported for both P and K. Initial desirable values for N, P, and K for poinsettia using the l : 1.5 media extraction method have also been determined (Prasad et a1 1983). Initial media test values vs. mid-term uptake and mid-term media test values vs. final uptake were highly correlated. Initial desirable values for media N were between 120 ppm and 200 in a peat medium and 180-225 for a bark medium. Desirable mid-term media test values were approximately 150 to 210 ppm and 175 - 225 ppm respectively. Desirable P levels in bark declined from initial samples to mid-term samples (9-12 to 3-4 ppm). In peat 16—20ppm of P was reported as desirable. No values were reported for K. Correlation between cyclamen nutrient uptake and media test values for the l : 1.5 media to water test, a modified Spurway and the media saturated extract method for N, P, and K are very similar (Prasad et a1 1983). The saturation extract method gives values that are roughly three times those of the 1 : 1.5 extract method for these three nutrients while the modified Spurway results were between 6 and 7 times less than the saturated extract. Since the three methods gave comparable plant uptake correlations selection of a routine testing method can be based on other criteria (i.e. ease). The saturated media extract method (Warncke 1980) in— volves taking a media sample (approximately 400cc) and 24 gradually adding distilled water until the sample is just saturated. The sample is allowed to equilibrate for 1 hour after which the sample is checked and media or water added to re-attain saturation. After an additional half hour equi- libration the pH is measured on the saturated medium which is then vacuum filtered to obtain the extract. Subsequent tests are done on the extract; i.e. soluble salts, nitrate, P, K, Ca and Mg. The saturated media extract method offers several advantages as a medium test method. Relatively large samples are used resulting in a low sampling error. Samples do not need to be dried or processed reducing analysis time. Differences in the bulk density of the media are automatically compensated for since bulk density is re- lated to the water holding capacity of the media and there- fore does not affect the interpretation of the results. Nutrient balance can be easily calculated, and a single ex- traction allows testing for pH, soluble salts, nitrate-N, P, K, Ca, and Mg (Warncke 1980). Wilkerson and O’Rourke (1983) stated "this method has not proven satisfactory for media prepared from southern pine bark. Due to the hydrophobic nature of the bark particles, a relatively large quantity of extractant is required for "wetting"...". Data obtained by Prasad et a1 (1983) (see previous discussion) does not agree with this statement. The saturated media ex— tract method has been used in the Michigan State University soil test program for a number of years with very good results and is the recommended procedure for use in the 25 North Central Region of the United States (Warncke 1980). Published standards for interpretation of test results in greenhouse media are readily available (Warncke 1980). Standards have also been developed for use in the produc- tion of containerized nursery stock (Timmer & Parton 1984). The evidence available regarding the testing of green- house growth media for macronutrient availability indicates that with few exceptions all the test procedures in use today work about equally well. Selection of a procedure may thus be based on secondary considerations such as ease of analysis, cost, amount of handling, preparation, time required for analysis etc. The saturation media extract seems to be the method of choice. Micronutrient Testing Micronutrient testing is done relatively routinely on field soil samples. A number of testing procedures have been correlated and calibrated for use in testing micronutrient content in soils. These testing procedures are generally un— satisfactory for use in greenhouse growth media. As with the macronutrient testing techniques soil sample handling and preparation methods for micronutrient analysis are inap- propriate for use with greenhouse growth media (see previous section). Most extractants used for testing micronutrient status in soils can be broadly classified into three groups; 1) weak acids, 2) strong acids, and 3) organic chelating 26 agents. The first extractants used were acids. Since the solid state forms of these micronutrient cations which con- trol the solubility of the nutrient in the soil dissolve in acid there is a good chemical basis for the use of these extractants. In many cases however the absolute presence or absence of the nutrient is not of as great a concern as the form of the nutrient present. Strong acid extractants may dissolve nutrients that would otherwise not go into solution and would thus be unavailable to the plant. Weak acid ex— tractants may not dissolve as much of the nutrient and thus may be a better indicator of plant available nutrients. There are several acid extractants commonly used. These include the double acid extractant (0.05 N HCL + 0.025 N H2804), which is used by North Carolina and a number of other southern states, and HCl usually at 0.1N. Results from each of these extractants have been correlated to plant Zn uptake (Tucker & Kurtz 1955, Wear & Evans 1968). 0.1N H2PO4 is used to test for Mn in Ohio and Wisconsin (Whitney 1980). Acetic acid and ammonium acetate have also been tried as extractants for Zn (Wear & Sommer 1948, Shaw & Dean 1952). Recently tests using organic chelating agents have been developed. EDTA and DTPA are the most common organic chelat- ing agents used in soil micronutrient testing. EDTA (ethylenediaminetetraacetic acid) has been used as an ex— tractant by itself and in conjunction with ammonium acetate and ammonium carbonate. Viro (1955) proposed the use of 27 .05M EDTA for use as an extractant for Zn, Cu and Mo. Tucker and Kurtz (1955) used .007M EDTA in ammonium acetate to test for Zn. EDTA (0.01M) in ammonium carbonate was tested by Trierweiler and Lindsay (1969). Little work has been done on the extraction of trace elements from growth media. Peat is a natural complexing agent for trace elements (Verloo 1980) and can therefore retain applied micronutrients. Coosemans and Uttebroeck (1983) examined the correlations between extracted levels of Fe, Cu, Zn, Mn, Cd, and Pb and tomato tissue levels of these elements (using 5 extractants). Of the 5 extractants~ (ammonium lactate + acetic acid, ammonium acetate, H2804 + HCl, HN03 and HN03 + HCl), extracted metals in nitric acid were best correlated to plant tissue levels overall. Tissue Zn was correlated with ammonium acetate extractable levels of Zn and tissue Mn was correlated with ammonium lactate extractable levels of Mn. In recent years the use of a universal micronutrient extractant has been promoted (Lindsey & Norvell 1978). This extractant consists of 0.005M DTPA (diethylenetriamine- pentaacetic acid), 1M TEA (triethanolamine), and .OlM CaClz. Of the chelating agents that could be used in a soil test DTPA is the best choice as a universal extractant. The stability constants of DTPA for complexes formed with these ions allow simultaneous complexing and thus extraction under soil conditions. Calcium chloride and TEA are added to the extracting solution to buffer the system in the slightly 28 alkaline pH range (pH 7.3) to prevent the dissolution of CaCOa and occluded micronutrients in calcareous soils. At equilibrium the micronutrient cation DTPA complexes (Fe, Mn, Zn, and Cu) are highly favored in competition with Ca and Mg ions particularly at soil pH levels below 7 (Lindsey & Nor- vell 1978). The test results are affected by extractant pH, extractant concentration, temperature, and shaking time. The test has been used successfully to test soils for Zn, Fe, Cu, and Mn (Lindsey & Norvell 1978). The DTPA test procedure developed by Lindsey and Norvell is a recommended test procedure in the North Central Region and is used by Iowa, Kansas, Minnesota, North Dakota, and South Dakota. Due to the sensitivity of the procedure to analytical conditions, care must be taken to maintain uniform testing methods and conditions (Whitney 1980). A modification of this method using a 1:4 sample to DTPA extractant ratio and a 120 minute shaking time has been developed for use in testing the Mn and Zn status of peat/vermiculite media in greenhouse tomato production (Markus et a1 1981). The DTPA extraction method is better correlated to tomato yield than the double acid method. The rates of application of Mn and Zn used in this study were very high, ranging from 200— 1800 ppm for Mn and 75- 675 ppm for Zn. Toxicity symptoms were reported for the highest treatment levels. Yield prediction curves were not presented so it is impossible to determine the relationship between test levels and yield. It is obvious, given the test levels 29 used, that the deficiency range for these nutrients was not examined therefore the usefulness of this test in evaluat- ing micronutrient deficiencies in a growth medium has not been established. Plant Tissue Analysis Principles Plant analysis methods can be broadly classified in two categories; 1) Total quantitative analysis and 2) semiquan— titative analysis (i.e. sap tests) (Aldrich 1973). Total or semiquantitative analysis can be used to relate soil or media test values to nutrients absorbed by the plant. A number of problems with the use of plant analysis can be identified; There are a number of techniques used in obtaining and processing samples. It must be recognized that the levels of nutrients in a plant tissue vary with the type of tissue, position on the plant, chronological and physiological age of the plant and the tissue, environmental conditions sur- rounding the plant, the species, and in some cases even the variety. A good sampling area for a mobile nutrient like N is not likely to be as good a sample site for Fe. Analysis standards must thus be based on closely defined reproducible sampling criteria to have interpretive meaning for unknown samples (Munson & Nelson 1973). Washing tissue samples can leach nutrients, par— ticularly if the sample is not fresh. However not washing 30 samples can lead to misleading or unusable results if con- tamination (i. e. soil particles , dust, spray residue) is present on the leaves (Jones & Steyn 1973). Plant tissue samples should be dried as quickly as pos- sible to prevent the loss of weight. If tissue samples are not to be dried immediately they must be stored in a way that minimizes cell respiration, decomposition, and bac- terial and fungal growth (Aldrich 1973). Differentiation of the various active and inactive fractions of an element in the plant is not possible using the basic methodologies. Physiological disorders such as lime induced Fe deficiency may not be detectable (Mengel & Kirkby 1982). Ground tissues are easy to handle, and it is very easy to obtain a uniform subsample for analysis but grinding tissues in a commercial grinder may be a substantial source of contamination (Jones & Steyn 1973). Care must be taken to limit, as much as possible this source of contamination. Plant tissue samples must be digested prior to analysis. There are two general methods of digestion, wet ashing and dry ashing. Wet ashing is done with concentrated acid i.e. nitric, or sulfuric. The procedures recommended for dry ashing plant tissues generally call for ashing the tissue at 500 C for 2 to 8 hours. As long as high walled crucibles are used for dry ashing there is little loss of nutrients (Jones & Steyn 1973). With the exception of cer- tain analytical procedures (i.e. N analysis) there is 31 little reason to favor wet ashing over dry ashing. Since dry ashing is considerably easier it may be the method of choice for most routine analytical work (Jones & Steyn 1973) The interpretation of plant analysis results depends on developing an understanding of the relationships between yield/growth, nutrient supply, and nutrient concentration in the plant. Ideal experiments to determine these relation- ships would provide for variation of all the essential nutrients, and other environmental and chemical conditions that affect growth and yield simultaneously. This ideal can not be met due to the enormous size and complexity of such an experiment. Usually only one, two or sometimes three nutrients will be varied, with the others held constant (theoretically in the optimum range) (Munson & Nelson 1973). In interpreting this type of experiment and the relationships drawn from the results it is important to keep in mind the dynamic relationships between the plant and its environment. The law of the minimum and balanced nutrition states that when one factor that limits growth becomes optimum one or several others become limiting (Prevot 1961). The critical levels for a nutrient determined under one set of experimental conditions may not hold true under different conditions. With knowledge of their limitations curves representing these relationships can be very useful. The general relationship between nutrient supply, nutrient concentration, and growth or yield is represented in Figure 32 1 (Brown 1970). The yield response to increasing nutrient supply is sigmoidal i.e. yield increases with added nutrient supplied until the optimum nutrient level is reached. After the optimum nutrient level is reached addi- tional nutrient is detrimental to plant growth or yield. The nutrient concentration in the plant is considered to have a biphasic curve increasing slowly from the point of minimum supply through the zone of poverty adjustment to the optimum. After the optimum is reached the nutrient con- centration in the plant increases at a much greater rate as nutrient supply is increased. This zone of the curve is known as the zone of luxury consumption. The slopes of these curves and their relative dimensions may be different with different nutrients, however the general relationships remain the same. The relationship between nutrient concentration and crop yield has been described by Ulrich and Hills (Figure 2) (1967). This type of curve is useful in developing sam- pling and interpretation guidelines for tissue analysis. The schematic does not address the problem of toxic affects of excess nutrient supply and concentration in the plant. Chapman (1967) provides a schematic graph (Figure 3) that deals with the full range of yield response with increasing nutrient concentration. It is worth noting that in each of these representations the yield drops off rapidly in response to small decreases in the nutrient concentration in the deficient or severely deficient ranges. Because of CROP YIELD —-——" CROP YIELD —-> 33 OPTIMUM YIELD --------- NUTRIENT CONCENTRATION ——> I I I I I I I I ZONE OF LUXURY r" CONSUMPTION '“ I I ' ZONE OF POVERTY ' «\Q‘)‘ I‘— ADJUSTMENT ’I 4%?” ' I nyu'. I $9” I I09" . MINIMUM NIHL'S-CRITICAL LEVEL I PERCENTAGE Rum?" | OR I.’ ,"uuv"" I OPflMUM I’nsu~"" (PHMWI ........... {-- SUPPLY . I I i NUTRIENT SUPPLY —————> ANNA?! ZONE .00 t£;.ért;6‘.. .- 2“ "TIM“. YIELD 9° 5 it 9% acoucnou INYICLC I 80 E a- g N) : I: I I; 60 I g I so 3 E ocncntut . a 40 m I 3 I . LE 30 I" E O'TIICUM zor I ' .o . ..::::;::I..;.. c 11 1 E i I l l L l l NUTRIENT CONCENTRATION m. ppm or Inca/moo) OPERATING RANGE OPTIMUM YIELD YIELO Loss RANGE—-I I I I I I I OPHMUM CONCENTRATION SEVER LOW 1 1 _ DEFICIENCY RANGE SUFFICIENCY RANGE EXCESS RANGE NUTRIENT CONCENTRATION —> Figure 1 Schematic of the ef— fects of nutrient supply on plant nutrient content and crop yield (Brown 1970). Figure 2 Schematic describing the relationship be— tween nutrient con- centration and crop yield (Ulrich & Hills, 1967). Figure 3 Schematic repre~ senting the relation— ship between crop yield and nutrient concentration (Chapman 1967). 34 genetic variability, testing errors, and environmental effects, it is, in reality, not possible to identify an Op- timum point on these curves. The optimum is better repre— sented by a range. By dividing the curve into ranges i.e. deficiency, sufficiency, and excess, information can be provided, based on test results that can allow a grower to achieve optimum yields. In most cases standard calibration curves can not realistically be divided into more than 3 - 5 response categories or ranges. Often standards are presented as a optimum range and a "critical value" below which deficiency may occur. Tissue analysis standards Poinsettia The suggested sampling site for tissue analysis for poinsettia is the most recently expanded mature leaves (Criley & Carlson 1970). Jones (1972) recommends sampling 15 - 20 plants at or prior to flowering. Mastalerz (1977) provides leaf analysis values given in table 3. Values presented by Criley and Carlson (1970) are in relatively close agreement. Chrysanthemum The suggested tissue sample area for overall nutrient analysis in Chrysanthemum is the upper leaves on young flowering stems prior to or at flowering. Twenty to 30 plants should be sampled (Jones & Steyn 1973). Detailed 35 sampling suggestions for specific nutrients have been worked out for the Chrysanthemum. Table 4 contains suggested values and sampling areas for Chrysanthemum taken from Mas— talerz (1977). The boron level reported for Chrysanthemum in table 4 may be too high under some conditions. Smilde (1975) reported boron toxicity symptoms in Chrysanthemum with leaf boron levels of 115 ppm and 145 ppm at media pH levels of 4.8 and 6.1 respectively. Sanderson (1975) noted that only a small range exists between B toxicity and deficiency in Chrysanthemum. Marigold Tissue samples of most bedding plants usually include both leaf and stem tissue, due to the relatively small size of these plants. Table 5 contains nutrient tissue levels of marigold grown in sand cultures from Johnson (1973) Table 3 Leaf analysis values for Poinsettia (Mastalerz 1977) Element Symbol Deficiency range Normal range Excess range Nitrogen (N) Less than 3 0 % 4.0 - 6.0 7.3 Phosphorus (P) Less than 0.2 % 0.3 - 0.7 0.7 Potassium (K) Less than 1.0 % 1.5 — 3.5 4.0 Calcium (Ca) Less than 0 5 % 0.7 - 2 0 Magnesium (Mg) Less than 0.2 % 0.4 — 1.0 Manganese (Mn) Less than 30ppm 100 - 200 250 + Iron (Fe) Less than 50ppm 100 - 500 Copper (Cu) Less than 5 ppm 6.0 — l5 Boron (B) Less than 20ppm 30 - 100 200 + Zinc (Zn) Less than 15ppm 25 - 60 36 Table 4 Nutrient ranges for Chrysanthemum leaf tissue (Criley and Carlson 1970) Element Adequate range Critical level Sample area N % 4.5 — 6.0 4.0 - 4.5 Upper leaves P % 0.26?—1.15 0.17?—0.26? Upper or lower leaves K % 3.5 - 10 2.15 — 2.75 Lower leaves Ca % .50 - 4.6 0.40 - 0.46 Upper leaves Mg % 0.14 -l.50 0.11? Lower leaves Fe ppm 750 < 125 Upper leaves Mn ppm 195 - 260 Upper or lower leaves B ppm 25 - 200 20.0 Upper leaves Cu ppm 10 1.7 -4.70 Middle leaves Zn ppm 7 — 26? 4.3 — 6.8 Lower leaves Table 5 Nutrient tissue levels of marigold grown in sand culture (from Johnson 1973) Nutrient solutions Nutrient - Nutrient Complete Nitrogen .40 % 5.20% Phosphorus 0.11 % .67 % Potassium .99 % 4.11 % Calcium .15 % 1.90 % Magnesium .04 % .75 % Iron 40 ppm 170 ppm Boron .68 ppm 103 ppm Manganese 49 ppm 193 ppm 37 DTPA Chelate Chemistry DTPA is diethylenetriaminepentaacetic acid (C14H23010N3). There are 5 acidic H bonds which can be dis- sociated as well as 3 sets Of electron pairs on N atoms available for ion complex formation. DTPA is thus a power- ful cation extractant. The chemistry of chelate (ligand) soil interactions has been worked out by Lindsey (1969) (1979). The ability of the chelate to extract an ion is de- pendent on the stability constants, or the relative binding power of the ligand ion complex. The relative amount of an ion extracted by a chelate is thus a result of the competi— tion of the potential forms of a given ion that may be present in the soil, and the concentrations and affinities of other ions that can form complexes with the ligand. The total ligand concentration is the sum of all the ligand ion complexes formed L(t) = L + HL + FeXL + ZnXL + MnXL + CuXL + CaXL + MgXL + MXL. The stability constants for the various potential ligand ion complexes are available in tabulated form in the literature (i.e. Lindsey 1979). With this information and knowledge of the forms of a cation present in a soil it is possible to develop stability diagrams for the chelate soil system, and to predict the relative concentrations of the various chelate ion com- plexes which will be present under equilibrium conditions. This then provides the theoretical basis for the use of a 38 DTPA soil test. Little is known about the conditions and forms of cat- ions which may be present in greenhouse growth media. It is established that greenhouse media possess only limited nutrient holding capacity. It is likely that the nutrient levels in solution in most growth media are a result of the fertilizer program and source rather than a solid state equilibrium, however the micronutrient concentration in a growth medium may be fairly constant over a relatively long period (Broschat et a1 1985). This indicates that the solution level of the micronutrients may be controlled by an exchange equilibria either with a solid state, or in ex— change complexes with the components of the growth medium. It should thus be possible to develop stability diagrams for greenhouse media similar to those developed for soils Plasma Emission Spectroscopy Plasma emission spectroscopy is a method of elemental analysis based on the measurement of the emission of energy from the molecules of a sample that have passed through an inductively-coupled argon plasma excitation source. Rapid multi-element analysis of plant tissues and soil extracts are possible using this method. The primary advantage Of plasma emission over other methods of multi-elemental analysis is the reduction of matrix problems. The effects of the sample matrix are reduced or eliminated through the 39 use of the high temperature plasma (Spiers et a1 1983). Soil extracts can be analyzed with little or no preparation, and plant digests can be prepared in similar fashion to that required for other methods of liquid analysis (Jones 1977). Few spectral interferences have been identified. Spiers (1983) reported interferences between major elements A1, Ca, Fe, K, and Ti, occurring in soil ex- tracts in excess of 1000 - 2000 ppm, and the minor elements As, Pb, Co, Mo, B and Cd. Several ions are known to self adsorb at higher con— centrations resulting in nonlinear concentration curves. This effect can be minimized and the effective linear range increased through the addition of Li to the sample. Boron exhibits a memory effect with plasma analysis (Jones 1977) (Spiers et a1 1983). The concentration of measured B will increase if the sample is analyzed repeatedly, presumably due to B adsorption to the fused quartz torch. A longer wash period between samples mini- mizes this affect. The standard Operating conditions of the Michigan State University soil testing laboratory Beckman spectrametrics III plasma emission spectrometer are listed in table 6. 40 Table 6 Standard operating conditions of the MSU plasma emission spectrometer (information provided by Beckman instrument company). Element Wavelength Linear Dynamic Range A1 308.215 0.8-1000 Zn 202.548 0.06-600 Cd 214.438 - 0.05—300 Fe 258.204 0.2-1000 B 249.678 0.08-1000 P 214.914 2-1000 Cu 324.754 0.02—10 Mg 280.270 0.006—100 Mn 257. 610 ' 0. 03—100 Mg 293.6 -1000 Ca 559.0 -3000 Mo 379.825 0.04—100 Ni 341.746 0.02—100 K 404.414 9-100 Ca 445.473 0.9-100 K 769.896 0.2-60 Pb 768.348 0.1—600 Cr 425.435 0.02—100 Na 568.820 2—1000 41 References Aldrich, S. R. 1973. Plant Analysis: Problems and Opportunities. p. 213-222. in; L. M. Walsh, and J. D. Beaton (ed). Soil Testing and Plant Analysis. Soil Sci. Soc. Amer. Inc. Madison, Wisconsin. Baker, K. F. (ed). 1957. The U. C. System for Producing Healthy Container-grown Plants. Calif. Agr. Exp. Sta. Ext. Serv. Manual 23. Berkley, Calif. Boodley, J. W., and R. Sheldrake Jr. 1963. Artificial Soils for Commercial Plant Growing. Cornell Exten- sion Bulletin No. 1104. Bunt, A. C. 1974. Some Physical and Chemical Characteris— ‘ tics of Loamless Pot-Plant Substrates and their Rela— tion to Plant Growth. Acta Hort. 37:1954-1965. Bunt, A. C. 1976. Modern Potting Composts. George Allen and Unwin LTD. London. Broschat, T. K., and H. M. Donselman. 1985. Extractable Mg, Fe, Mn, Zn, and Cu from a peat-based Container Medium Amended with Various Micronutrient Fertilizers. J Am Soc Hrt Sci. 110(2):196-200. Brown, J. R. 1971. Plant Analysis. Missouri Agr. Exp. Sta. Bull. SB881. Chapman, H. D. 1967. Plant Analysis Values Suggestive of Nutritional Status of Selected Crops. p. 77-92. in; Soil Testing and Plant Analysis. Part II. SSSA Spe- cial Publ. Series No. 2. Soil Sci. Soc. Amer. Madison, Wis. Chen, Y. and P. Barak. 1982. Iron Nutrition of Plants in Calcareous Soils. Advances in Agronomy. 35:217-240. Carlson, W. H., and E. M. Rowley. 1980. Bedding Plants. p. 479- 521. in; R. A. Larson. (ed). Introduction to Floriculture. Academic Press. New York. Coosemans, J., and P. Uyttebroeck. 1983. Comparisons of Different Extracting Methods for Heavy Metals From "Compost" Substrates and from Tomato Leaves and Fruits Grown on These Substrates. Acta Hort. 133:165—171. Criley. R. A., and W. H. Carlson. 1970. Tissue Analysis Standards for Various Floricultural Crops. Flor. Rev. 146(3771):19-20,70—73. 42 Dight, R. J. W. 1977. Nutritional Requirements of Bedding Plants. Exp. Hort. 29:63—71. Hanaway, J. J. 1973. Experimental Methods for Correlating and Calibrating Soil Tests. P. 55-66. in; L. M. Walsh, and J. D. Beaton (ed). Soil Testing and Plant Analysis. Soil Sci. Soc. Amer. Inc. Madison, Wisconsin. Holcomb, E. J., and J. W. White. 1979. Using Plant Uptake to Determine Optimum Values for Soil Tests. J. Amer. Soc. 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Madison, Wisconsin. Jones, L. H. P. 1957. the Relative Content of Manganese in Plants. Plant and Soil. 4:328-335 Krauskopf, D. M. 1982. Personal Communication Lamaire, F., A. Dartigues,and L. M. Riviere. 1980. Properties of Substrates With Ground Pine Bark. Acta Hort. 99:67-75. Lawrence, W. J. C., and J. Newell. 1939. Seed and Potting Composts. George Allen And Unwin LTD. London. 43 Lindsay, W. L. 1979. Chemical Equilibria in Soils. John Wiley and Sons. New York. Lindsay, W. L., and W. A. Norvell. 1969. Equilibrium Relationships of ZnZt, Fe3+, Ca2+, and H+ with EDTA and DTPA in Soils. Soil Sci. Soc. Amer. Proc. 33:62—68. Lindsay, W. L., and W. A. Norvell. 1978. Development of a DTPA Soil Test for Zinc, Iron, Manganese, and Copper. Soil Sci. Soc.Am. J. 42:421—428. Marconi, D. J., and P. V. Nelson. 1984. Leaching Of Ap- plied Phosphorous in Container Media. Scientia Hort. 22:275—285. Markus, D. K., and R. L. Flannery. 1983. Macronutrient Status in Potting Mix Substrates and in Tissue of Azaleas From the Effects of Slow-Release Fertilizer. Acta Hort. 133:179—190. Markus, D. K., and J. E. Steckel. 1980. Periodical Analysis of Artificial Rooting Media and Tomato Leaf Analysis From New Jersey Greenhouses. Acta Hort. 99:205—217. Markus, D. K., J. E. Steckel, and J. R. Trout. 1981. Micronutrient Testing in Artificial Mix Substrates. Acta Hort. 1262219-235. Mastalerz, J. W. 1977. The Greenhouse Environment. John Wiley and Sons. New York. Melsted, S. W., and T. R. Peck. 1973. The Principles of Soil testing. p. 13-22. in; L. M. Walsh, and J. D. Beaton (ed). Soil Testing and Plant Analysis. Soil Sci. Soc. Amer. Inc. Madison, Wisconsin. Mengel, K., and E. A. Kirkby. 1982. Principles Of Plant Nutrition. International Potash Institute. Bern, Switzerland. Munson, R. D., and W. L. Nelson. 1973. Principles and Practices in Plant Analysis. p. 223-248. in; L. M. Walsh, and J. D. Beaton (ed). Soil Testing and Plant Analysis. Soil Sci. Soc. Amer. Inc. Madison, Wisconsin. Nash, V. E., and A. J. Laiche Jr. 1981. Changes in the Characteristics of Potting Media with Time. Commun. in Soil Sci. Plant Anal. 12(10):lOll-1020. 44 Neal, J. C., D. F. Wagner. 1983. Physical and Chemical Properties of Coal Cinders as a Container Media Component. HortScience. 18(5):693-695. Peck, T. R., And 8. W. Melsted. 1973. Field Sampling for Soil Testing. p. 67-76. in; L. M. Walsh, and J. D. Beaton (ed). Soil Testing and Plant Analysis. Soil Sci. Soc. Amer. Inc. Madison, Wisconsin. Pokorny, F. A., and H. Y. Wetzstein. 1984. Internal Porosity, Water Availability and Root Penetration Of Pine Bark Particles. HortSci. 19:447—449. Prasad, M., T. M. Spiers, and I. C. Ravenwood. 1981. Soil Testing of Horticultural Substrates (1) Evaluation Of 1:1.5 Water Extract for Nitrogen. Commun. in Soil Sci. Plant Anal. 12(9):811-823. Prasad, M., T. M. Spiers, I. C. Ravenwood and R. W. Johnston. 1981. 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Soil Testing of Horticultural Substrates for Cyclamen and Poinsettia. Commun. in Soil Sci. Plant Anal. 14(7) 553-573. Robertson, L. 8., D. R. Christenson, and D. D. Warncke. 1967. Essential Secondary Elements: Calcium Mich. State Univ. Ext. Bull. E—996 45 Robertson, L. 8., M. L. Vitosh, and D. D. Warncke. 1976. Essential Secondary Elements: Sulfur. Mich. State Univ. Ext. Bull. E-997. Sanderson, K. C. 1983. Growing with Artificial Media: The Advantages, Disadvantages. Southern Flor. & Nurseryman. July 29:13-14. Shaw, E., and L. A. Dean. 1952. Use of Dithizone as an Ex- tractant to Estimate the Zinc Nutrient Status of Soils. Soil Sci. 73:341-347. Sheard, G. F. 1975. Loamless Substrates for use in Con- tainers and as Unit Products. p. 119 - 132. In D. W. Robinson and J. G. D. Lamb (ed). Peat in Horticulture. Academic Press. London. Smilde, K. W., 1975. Micronutrient Requirements of Chrysanthemums Grown on Peat Substrates. Acta Hort. 50:101-113. Sonneveld, G., J. van den Ende, and P. A. van Dijk. 1974. Analysis of Growing Media By Means of a l : 1.5 Volume Extract. Commun. in Soil Sci. Plant Anal. 5(3):183-202. Spiers, G. A., M. J. Dudas, and L. W. Hodgins. 1983. In— strumental Conditions and Procedure for Multielement Analysis of Soils and Plant Tissue by ICP—AES. Commun. in Soil Sci. Plant Anal. 14(7):629-644. Spomer, A. L. 1979. Physical Properties of a Good con- tainer Soil Amendment. Univ. Ill. Exten. Bull. FL-5- 79. Spurway, C. H. 1943. Soil Fertility Control For Greenhouses. Mich. State. Coll. Agr. Expt. Sta. Spe— cial Bull. No.325. Timmer, V. R., and W. J. Parton. 1984. Optimum Nutrient Levels in a Container Growing Medium Determined by a Saturated Aqueous Extract. Commun. In Soil Sci. Plant Anal. 15(6):607-618. Trierweiler, J. F., and W. L. Lindsay. 1969. EDTA-Ammonium Carbonate Soil Test for Zinc. Soil Sci. Soc. Amer. Proc. 33:49—53 Tucker, T. G., and L. T. Kurtz. 1955. A Comparison of Several Chemical Methods With the Bio-assay Procedure for Extracting Zinc from Soils. Soil Sci. Soc. Amer. Proc. 19:477-481. 46 Ulrich, A., and J. F. Hills. 1967. Principles and Prac— tices of Plant Analysis. p. 11-24. in; Soil Testing and Plant Analysis. Part II. SSSA Special Publ. Series No. 2. Soil Sci Soc. Amer. Madison, Wis. Van Lierop, W. 1981. Conversion of Organic Soil pH Values Measured in Water, 0.01M CaC12 or 1N KCl. Can. Jour. Soil Sci. 61:577—579. Verdonck, 0., D. De Vleeschauwer, and R. Penninck. 1983. Barkcompost, a New Accepted Growing Medium For Plants. Acta Hort. 133:221-226. Verloo, M. G. 1980. Peat as a Natural Complexing Agent for Trace Elements. Acta Hort. 99:51—56. Viro, P. J. 1955. Use of Ethylenediaminetetraacetic acid in Soil Analysis: I Experimental. Soil Sci. 75:459- 465. Vitosh, M. L., D. D. Warncke, B. D. Knezek, and R. E. Lucas. 1981. Secondary and Micronutrients for Vegetables and Field Crops. Mich. State Univ. Ext. Bull. E-486. Warncke, D. D. 1980. Recommended Test Procedures For Greenhouse Growth Media. p. 31-33. In; W. C. Dahnke (ed). Recommended Chemical Soil Test Proce- dures For the North Central Region. Bulletin NO. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. Warncke, D. D., D. M. Krauskopf. 1983. Greenhouse Growth Media Testing and Nutrition Guidelines. Mi. State Univ. Exten. Bull. E-1736. Watson, M. E. 1980. Recommended Soil Boron Tests. p. 22- 24 In; W. C. Dahnke (ed). Recommended Chemical Soil Test Procedures For the North Central Region. Bul— letin No. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. Wear, J. I., and C. E. Evans. 1968. Relationship of Zinc Uptake by Corn and Sorghum to Soil Zinc Measured by Three Extractants. Soil Sci. Soc. Amer. Proc. 32:543-546. Wear, J. I., and A. L. Sommer. 1948. Acid—extractable Zinc of Soils in Relation to the Occurrence of Zinc Deficiency Symptoms of Corn: A Method of Analysis. Soil Sci. Soc. Amer. Proc. 12:143-144. White, J. W., and R. J. Thomas. 1975. Analytical Methods for Peat Substrates. Acta Hort. 50:157-161. 47 Whitney, D. A. 1980. Micronutrient Soil Tests-Zinc, Iron, Manganese, and Copper. p. 18—21 In; W. C. Dahnke (ed). Recommended Chemical Soil Test Procedures For the North Central Region. Bulletin NO. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. and E. N. O’Rourke. 1983. Three Analytical Systems on Nutrient "Availability" HortScience. Wilkerson, D. C., The Effects of the Interpretation of in Pine Bark Growing Media. l8(3):301-302. SECTION I Micronutrient Testing of Plant Growth Media: Extractant Evaluation Micronutrient Testing of Plant Growth Media: Extractant Evaluation Key Words Greenhouse media, Micronutrients, Saturation extract, Media testing, Media analysis, DTPA. R. D. Berghage, D. M. Krauskopf, D. D. Warncke, and I. Widders. Department of Horticulture, Michigan State University, Department Of Crop and Soil Science, Michigan State University, East Lansing, Michigan Abstract Fifteen extractants were evaluated using saturation extract procedures for their ability to remove B, Fc, Mn, and Zn from a l : l by volume, peat and vermiculite media amended with three levels of micronutrients. Five millimolar (0.005M) DTPA (diethylenetriamine pentaacetic acid) was found to be the best over all extractant of those tested. 48 49 Introduction The physical and chemical properties of greenhouse plant growth media, particularly those formulated without soil are very different than those of soils. As a result of these differences nutritional testing procedures developed for use with soils do not work well in greenhouse media. The saturated media extract method (3) offers a number of ad- vantages over other media testing procedures (4,10): l). Relatively large samples are used resulting in a low sampling error; 2) Samples do not need to be dried and processed thus reducing analysis time; 3) Differences in the bulk density of the media are compensated for since bulk density is related to the water holding capacity of the media and hence does not affect the interpretation of the results; 4) Nutrient balance can be easily calculated; 5) A single extraction allows testing for pH soluble salts N,P,K,Ca, and Mg (10). One major limitation of the saturated extract method is that it is not used to test media for micronutrient content since these nutrients are present at very low concentrations in the DI water extract. Zinc, for example, is present in saturated media extracts at concentrations of 0.1 to 1.2 ppm (9) which are too low to allow reliable measurement and interpretation. A solution to this problem would be to modify the saturated media 50 extract method to allow simultaneous testing Of both macro— nutrients and micronutrients. Any modification should have extract test values that are correlated to plant uptake and result in a wide range of test values, so that deficient, adequate, and toxic levels of the nutrient can be separated. It would be time consuming to develop new macronutrient standards for a different extractant, so any new extractant should produce macronutrient test levels that are sig- nificantly correlated to those obtained with DI water. The objective of this study was to evaluate a variety of extrac- tants using saturated extract methodology. The extractants were chosen on the basis Of their previous use in soils. A number of strong and weak acids have been used as ex- tractants for micronutrients in soils (7, 12, 13). Ammonium acetate is a common soil extractant (7,5). The chelates DTPA (1,2), and EDTA (8) have also been used successfully in testing soils for micronutrient status. DTPA is usually used with calcium chloride and TEA (12). Calcium chloride is used to establish an equilibrium with calcium carbonate in the soil and thus prevent the dissolution of occluded nutrients. Since the objective of this study was to find an extractant that could be used for both micronutrients and macronutrients, calcium could not be included in the ex— tracting solution. EDTA by itself (8) or in combination with ammonium acetate (7) or ammonium carbonate (6) have been used to test for soil Zn. Soil boron is commonly 51 extracted with boiling water (11) so several of the extrac— tants were heated to boiling prior to their use. Materials and Methods Extractants The following extraction treatments were tested: D.I. water Acetic acid 0.1N (0.1N HOAc) Acetic acid 1.0N (1.0N HOAc) Hydrochloric acid ( 0.1N HCl) 0.05N HCl + 0.025N H2804 (double acid) Ammonium acetate (1.0N NH4OAc) DTPA 0.005M (Diethylenetriaminepentaacetic acid) DTPA 0.005M + TEA 0.1M at pH 7.3 (TEA=triethanolamine) .OlM (Ethylenediaminetetraacetic acid) .01M in NH4COs at pH 8.6 .007M in NH40Ac at pH 7.0 12. DTPA .005M boiling 13. EDTA .005M boiling 14. Acetic acid 0.1N boiling 15. D.I. water boiling mqmmawmn—a 9. EDTA 10. EDTA 11. EDTA OOOOO Nutrient test levels obtained with each of the extrac- tants were evaluated in comparison with the test levels of the deionized water extractions. Media preparation A Cornell mix (1 part course sphagnum peat to 1 part course vermiculite, by volume) was amended with 2.97Kg dolomitic lime, 1.2Kg superphosphate, and 2.97Kg gypsum per cubic meter of medium. The medium was charged with Fe, Mn, Zn, and B by saturating it with a solution of these 52 micronutrients in deionized water (table 1) and allowing this mixture to equilibrate for 48 hours. The medium was then purged with deionized water to remove any excess micronutrients remaining in the bulk solution. Ap- proximately 1 liter of deionized water purging solution was used for every liter of medium. The purge volume used was based on an elution gradient series done under identical conditions. Table l Micronutrient concentration in media amendment solutions Micronutrient source level 1 level 2 1eVel 3 Iron Fe chelate Oppm 24ppm 96ppm (Sequestrene 330) Manganese MnSO H O Oppm 34ppm 136ppm Zinc ZnSO 7H 0 Oppm 30ppm 120ppm Boron Boric acid Oppm 4.5ppm 18ppm Extractions were done using saturated media extract procedures (10). pH was measured on the saturated media while soluble salts and elemental concentrations were measured on the extract. A Beckman spectrametrics IIIA plasma emission spectrometer was used for the elemental analysis. 53 Experimgptal Design and Statistical Analysis The experiment was conducted as a split plot with amendment rate as the whole plot. There were no replica- tions at the whole plot level. The 15 extractant treatments in the split plots were done in 3 randomized complete blocks. Extractions were done on 3 days, media amended with the level 1' micronutrient charge was extracted on 10/13/83, level 2 on 10/15/83, and level 3 on 10/16/83. Extractant Evaluation Extractants were evaluated on the basis of the follow— ing criteria: A) Low standard error between replications (low coeffi— cient of variation). B) Minimal differences between macronutrient values Ob— tained with the extractant and those obtained by extraction with D.I. water. C) Minimal differences between the pH and soluble salts obtained with the extractant and those obtained with D.I. water. D) Large measured differences between media equi~ librated with low and high levels of micronutrients. 54 Results and Discussion The coefficients Of variation for extractions of media amended with micronutrients at level 2 are given in Table 2. These coefficients can be used to compare the variability of each extraction procedure. With few excep- tions the variability is less than 10%. One notable excep- tion is Mn in the DI water extract which had 9 % variability. This extraction procedure appears to be un- suitable for testing media for Mn. There were large differences in the efficacy of the extractants. The concentration Of macronutrients in extract solutions for the various extractants and the least sig- nificant differences (lsd .01) for each are shown in figures 1-4. All extractants except EDTA in ammonium car— bonate removed significantly more calcium from the media than D.I. water or boiling water extracts. The strong acid extractants and those containing ammonium carbonate removed 4 to 5 times as much Ca, K and Mg as the D.I. water extracts. It is likely that these extractants are dissolving relatively unavailable, (to the plant), forms of these nutrients such as calcium and magnesium carbonate and K bound within the layers of vermiculite. The concentration of K extracted by .005M DTPA, .005M EDTA and .005M DTPA +.1M TEA was not significantly different than the level of K obtained with the D.I. water extract. Magnesium extracted 55 with each of the extractants was significantly greater than was extracted with D.I. water, however differences were small for the DTPA extractants, 0.005M EDTA and the Boiling DI. water extractions. The amount of P extracted by the various extractants appears to be partially a result of the pH of the extracting solution. The solutions with higher pH removed less P than those with low pH. For example EDTA in ammonium carbonate had the highest pH and removed the least P, while HCl had the lowest pH and extracted the most P. Differences in P extracted by .005M DTPA, .005M EDTA, am+ monium acetate extractions and D.I. water were small but significant. The EDTA extracts produced pH values that were closest to those obtained with deionized water (figure 5). DTPA ex— tracts had lower pH test results than EDTA but were higher than the strong acid extracts. The buffered extracts had pH readings that were higher than DI water extracts. Three of the extracting solutions, NH4OAc, EDTA with NH4CO, and EDTA with NH4OAc produce soluble salts test results much too high for use. DTPA and EDTA extractions resulted in soluble salts test levels that were not sig— nificantly different from those Obtained with D.I. water (Figure 6). Boron, Mn, Fe, Zn, and Cu concentrations obtained from the 15 extractants are given in figures 7 — 11. None of the extractants was very effective in extracting B. One normal 56 (1.0N) acetic acid extractions resulted in the highest B test level. Heating extractants produced very little in- crease in the concentration of B extracted. The differences in extracted boron between amendment levels were small but significant and could potentially be used in a testing program. All of the extractants removed more Mn and Fe than D.I. water. This was true for Zn as well with the exception of Zn extracted with 0.1N acetic acid which was not sig— nificantly different than Zn extracted by water. The acids were, in general, less effective in extracting Mn, and Zn than the chelating agents. The strong acids 1.0N HCl and 1.0 N acetic, were more effective than the weaker acids, removing as much Fe as the buffered DTPA and EDTA solutions. EDTA, and DTPA and any of the mixtures containing them.were effective extractants for Mn, Zn, and Fe. Boiling DTPA and EDTA extractants removed large amounts of iron some of which is probably bound to the organic matter of the media and not available for plant uptake. The extractants containing EDTA were very difficult to analyze due to difficulties in preparing standards for use with the plasma emission spectrometer. Standards crystalized if allowed to stand. According to our criteria the best overall extractant was .005M DTPA. It was easy to handle and extracted large amounts of ZN, Mn, and Fe. The DTPA extraction resulted in a 57 relatively small but significant change in pH, Mg, Ca, and P tests as compared to DI water. Potassium extracted with 0.005M DTPA was not significantly different than K in water extracts. Substitution of 0.005M DTPA for water in the saturated media extract method will provide significant en- hancement of micronutrient concentrations in the extracts while producing minimal alterations in macronutrient concentrations. References 1. Lindsay, W. L. and W. A. Norvell. 1969. Equilibrium relationships of Zn ,Fe ,Ca ,and H with EDTA and DTPA in soils. Soil Sci. Soc. Amer. Proc. 33:62-68. 2. Lindsay, W. L. and W. A. Norvell. 1978. Development of a DTPA soil test for zinc, iron, manganese,and copper. Soil Sci. Soc. AM. J. 42:421-428 3. Lucas, R. E., P.E. Ricke and E.C. Doll. 1972. Soil saturation extract method for determining plant nutrient levels in peats and other soil mixes. 4th International Peat Congress. 3:221-230 4. Markus, D.K., J.E. Steckel and J.R. Trout. 1981. Micronutrient testing in artificial mix substrates. Acta Hort. 126:219-235. 5. Shaw, E. and L.A. Dean. 1952. Use of dithizone as an extractant to estimate the zinc nutrient status of soils. Soil Sci. 73:341—347. 6. Trierweiler, J.F. and W.L. Lindsay. 1969. EDTA- ammonium carbonate soil test for zinc. Soil Sci. Soc. Amer. Proc. 33:49-53. 7. Tucker, T.C. and L.T. Kurtz. 1955. A comparison of several chemical methods with the bio-assay procedure for extracting zinc from soils. Soil Sci. Soc. Proc. 19:477-481. 10. ll. 12. 13. 58 Viro, P.J. 1955. Use of ethylenediaminetetraacetic acid in soil analysis: I experimental. Soil Sci. 75:459-465. Warncke, D.D. 1979. Testing greenhouse growing mediazupdate and research. Proc. 7th Annual Soil- Plant Analysts Workshop, NCR—13 Committee, Bridgetown, Mo., Nov. 6, 1970. Warncke, D.D. 1980. Recommended test procedures for greenhouse growth media. In: Recommended Chemical soil Test Procedures for the North Central Region. North Dakota State University,~ Bulletin No. 999(revised). p 31-33 Watson, M.E. 1980. Recommended soil boron tests. In: Recommended Chemical soil Test Procedures for the North Central Region. North Dakota State University, Bulletin No. 999(revised). p 22-24. Whitney, D.A. 1980. Micronutrient soil testsflzinc, iron, manganese, and copper. In: Recommended Chemi- cal soil Test Procedures for the North Central Region. North Dakota State University, Bulletin No. 999(revised). p 18-21. Wear, J.I. and A.L. Sommer. 1948. Acid-extractable zinc of soils in relation to the occurrence of zinc deficiency symptoms of corn: A method of analysis. Soil Sci. Soc. Amer. Proc. 12:143—144 Wear, J.I. and C.E. Evans. 1968. Relationship of zinc uptake by corn and sorghum to soil zinc measured by three extractants. Soil Sci. Soc. Amer. Proc. 32:543—546. drenched with 24 ppm Fe, 34 ppm Mn, 30 ppm Zn and 4.5 ppm B, Extractant D.I. water D. I. water (boiling) EDTA .005M EDTA .005M (boiling) HCl 0.1N NH4OAC 1N HOAc 1.0N HOAc 0.1N HOAc 0.1N (boiling) DTPA .005M (boiling) DTPA .005M HCL + H2804 EDTA in NH4OAc EDTA in NH4C03 DTPA + TEA * Below detection limits 59 Table 2 Coefficients of variation Of saturated media test results for a 1:1 peat to vermiculite media ammended with 1.2 Kg superphosphate, and 2.97 Kg of gypsum and dolomitic lime per cubic yard of media, extracted using 15 extractants. Mg CO C) oak-1.500 NC!) COCO COP-t NK‘I 000qu I—JCOI-amoo‘ CDCO-bO‘IUI Ca C0 00 I—II-hoolm 0CD .e-q UINO‘ICDC) OULONQCO (DONWCO K 01~J taco 12 4.4 N—DQNO) \ILDOCOUI 1 I—‘I—J N I-‘I--‘ unfit-WI) 11 P A (.0 NCO OI—I moor-acne.) NWO mNI—‘NI-J 01-4ka Zn c» .s-qcnco.> (Did ~Jeao>~3eo LO OICDU'ICDCD \INIbNOO (ON Fe .507 I—IUI 11 6.0 OOCDOOLO CDIDNWCD 2.5 11 8.8 15 5.5 010.) 001 O) Nat—HAG! co N qqmmou £00103 NCOO‘I Mn O (.0 I043 \100 rah-0.50) NOI‘OON IOWIQ-DN COLONION 95 27 Cu 60 Figure 1. Phosphorus extracted from a 1:1 peat to ver- media using saturated media extraction techniques with 15 different extractants. miculite (DCDQCDOIQOONI—J D.I. water 0.1N HOAc 1.0N HOAc 0.1N HCl 0.05N HCl + 0.025N H2804 1.0N NH4OAc DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH40Ac at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D.I. water boiling 61 EBoobxm m—ermrmerrormmhmmwmm—o o om d w. o... m 1. I D 3 1| 8 \2 d d W cm 5. Um. H 09. 62 Figure 2. Potassium extracted from a 1:1 peat to ver- media using saturated media extraction with 15 different extractants. miculite techniques COWQO‘IOI-DOONI—I I—‘I—‘r—‘I—‘H ANNE-'0' 15. D.I. water 0.1N HOAc 1.0N HOAc 0.1N HCl 0.05N HCl + 0.025N H2804 1.0N NH4OAc DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH4OAc at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D .I. water boiling 63 m— .1 «.4 NF : o— EBoobxm mmhomemme 5. nm_ H o (Ludd) ioohxe U1 )1 64 Figure 3. Calcium extracted from a 1:1 peat to vermiculite media using saturated media extraction techniques with 15 different extractants. COW'QQUI-Dwml—l D.I. water 0.1N HOAc 1.0N HOAc 0.1N HCl 0.05N HCl + 0.025N H2804 1.0N NH4OAc DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH4OAc at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D. I. water boiling 65 m? 4— HF NF FF or Eoyoobxm m m h m m e m a F Fo.cw_H o 0 com no 0 w. 094 e X \IT J O .3. com \I/ d d 3... com 000? Figure 4. Magnesium LOWQCDOIIDCJIOI—J 66 extracted from a 1:1 peat to ver- miculite media using saturated media extraction techniques with 15 different extractants. D.I. water 0.1N HOAc 1.0N HOAc 0.1N HCl 0.05N HCl + 0.025N H2804 1.0N NH4OAc DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH4OAc at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D.I. water boiling 67 m? 3 Mr NF : or EBoobxm mmhmmwmm F o oo— O O (\I oom (Ludd) ioonxe u! 6w Figure 5. pH of LOCDQCDOIubwmI—J 68 saturated media extracts of a 1:1 peat to vermiculite media extracted with 15 different extractants. D.I. water 0.1N HOAc 1.0N HOAc 0.1N HCl 0.05N HCl + 0.025N H2504 1.0N NH40Ac DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH40Ac at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D.I. water boiling 69 m— I: n? NF C or Egoobxm mwhomemm _. o iooiixe 3Io Hd 70 Figure 6. Soluble salts of saturated media extracts of a 1:1 peat to vermiculite media extracted with 15 different extractants. 1. D.I. water 2. 0.1N HOAc 3. 1.0N HOAc 4. 0.1N HCl 5. 0.05N HCl + 0.025N H2304 6. 1.0N NH40AC 7. DTPA 0.005M 8. DTPA 0.005M + TEA 0.1M at pH 7.3 9. EDTA 0.01M 10. EDTA 0.01M in NH4COs at pH 8.6 11. EDTA 0.007M in NH4OAc at pH 7.0 12. DTPA 0.005M boiling 13. EDTA 0.005M boiling l4. Acetic acid 0.1N boiling 15. D.I. water boiling 7 rr/ gags \\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\‘ I\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\I 7 é I\\\\\\I 7’ é I\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\I I\\\\\\\\‘ \\\\\\\\\\\I N\\\\‘ I\\\\‘ _ I led .01 g r r ’l’ r 101112131415 r rrl 89 r 7 L-CO LL!) em LN _F 0 fi" 304 (\I (84!) IODJIXEI U! SIIDS qunIos T 1 O O O Extractant Figure 7. 72 Iron extracted from a 1:1 peat to vermiculite media amended with (l) Oppm Fe, (2) 24ppm Fe, and (3) 96ppm Fe extracted using saturated media methods with 15 different extractants. 1. D.I. water 2. 0.1N HOAC 3. 1.0N HOAC 4. 0.1N HCl 5. 0.05N HCl + 0.025N H2804 6. 1.0N NH40Ac 7. DTPA 0.005M 8. DTPA 0.005M + TEA 0.1M at pH 7.3 9. EDTA 0.01M 10. EDTA 0.01M in NH4C03 at pH 8.6 11. EDTA 0.007M in NH4OAc at pH 7.0 12. DTPA 0.005M boiling 13. EDTA 0.005M boiling 14. Acetic acid 0.1N boiling 15. D.I. water boiling 73 m_. .1 Mr N— : or EBoobxm m _o>o._ S. cm_ H N _o>oI_ F0. om_ H F _o>om Po. Um_ H n _o>oI_ U N _o>om m P _m>om @ (wdd) ioohxe UI e_.I Figure 8. 74 Manganese extracted from a 1:1 peat to ver- miculite media amended with (1) Oppm Mn, (2) 34ppm Mn, and (3) 136ppm Mn extracted using saturated media methods with 15 different extractants. 1. D.I. water 2. 0.1N HOAc 3. 1.0N HOAC 4. 0.1N HCl 5. 0.05N HCl + 0.025N H2804 6. 1.0N NH4OAc 7. DTPA 0.005M 8. DTPA 0.005M + TEA 0.1M at pH 7.3 9. EDTA 0.01M 10. EDTA 0.01M in NH4C03 at pH 8.6 11. EDTA 0.007M in NH4OAc at pH 7.0 12. DTPA 0.005M boiling 13. EDTA 0.005M boiling 14. Acetic acid 0.1N boiling 15. D.I. water boiling 75 m— I: Mr N— : or Egoobxm m m _o>ol_ Po. Um_ H N _o>o4 Po. be H F _o>o._ F0. “32 H n as: .U N as: m a 6%.. @ (wdd) ioonxe uI UW Figure 9. 76 Zinc extracted from a 1:1 peat to vermiculite media amended with (1) Oppm Zn, (2) 30ppm Zn, and (3) 120ppm Zn extracted using saturated media methods With 15 different extractants. 1. D.I. water 2. 0.1N HOAC 3. 1.0N HOAc 4. 0.1N HCl 5. 0.05N HCl + 0.025N H2804 6. 1.0N NH4OAc 7. DTPA 0.005M 8. DTPA 0.005M + TEA 0.1M at pH 7.3 9. EDTA 0.01M 10. EDTA 0.01M in NH4CO3 at pH 8.6 11. EDTA 0.007M in NH40AC at pH 7.0 12. DTPA 0.005M boiling 13. EDTA 0.005M boiling 14. Acetic acid 0.1N boiling 15. D.I. water boiling 77 mr .1 MP N_. Z or €9.00be m 5,3 5. n2 _ 64,3 5. as n 34,3 U N at: m _ as: Q m. _o>o4 Po. Um_ H H H (wdd) ioonxe uI UZ Figure 10. 78 Boron extracted from a 1:1 peat to vermiculite media amended with (l) Oppm B, (2) 4.5ppm B, and (3) 18ppm B extracted using saturated media methods with 15 different extractants. 1. D.I. water 2. 0.1N HOAC 3. 1.0N HOAC 4. 0.1N HCl 5. 0.05N HCl + 0.025N H2804 6. 1.0N NH4OAC 7. DTPA 0.005M 8. DTPA 0.005M + TEA 0.1M at pH 7.3 9. EDTA 0.01M 10. EDTA 0.01M in NH4C03 at pH 8.5 11. EDTA 0.007M in NH4OAc at pH 7.0 12. DTPA 0.005M boiling l3. EDTA 0.005M boiling 14. Acetic acid 0.1N boiling l5. D.I. water boiling 79 Egoobxm. m—GFMFNFF—orm v 0.0 To m6 N4 o>o P. m m_ m o U _H mé n _o>o._ D N _o>mj 5. U2 H N _o>om N ®>® .m 233% I neon; ON 8 (Ludd) ioohxe uI 80 Figure 11. Copper extracted from a 1:1 peat to vermiculite media using saturated media extraction techniques with 15 different extractants. LOWQCDUIIDOJI‘Or—I D.I. water 0.1N HOAC 1.0N HOAC 0.1N HCl 0.05N HCl + 0.025N H2804 1.0N NH4OAC DTPA 0.005M DTPA 0.005M + TEA 0.1M at pH 7.3 EDTA 0.01M EDTA 0.01M in NH4C03 at pH 8.6 EDTA 0.007M in NH4OAc at pH 7.0 DTPA 0.005M boiling EDTA 0.005M boiling Acetic acid 0.1N boiling D.I. water boiling 81 F j l\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\I I\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\I I\\\\\\\\\\\\\\\\\\\\\\\\\\\‘ .\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\‘ II? E i **l* *I’ 101112131415 T I I\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\‘ CD I\\\\\\\\\\\\\\\\\\\\ \\I L- co I\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\\V #- I\ ’ 3 o ' —-N 8 _ __ I 4 I i j 1 1 1 1 fl I j I 1 O ‘9 L0. 81 ‘2 9! T: O, O O O o o o o (LUdd) ioonxe UI no Extractant SECTION II Micronutrient Testing in Greenhouse Media: Extractant Characterization Micronutrient Testing in Greenhouse Media: Extractant Characterization Key Words Greenhouse media, Micronutrients, Saturation extract, Media testing, DTPA. R. D. Berghage, D. M. Krauskopf, D. D. Warncke, and I. Widders Department of Horticulture, Department of Crop and Soil Science, Michigan State University, East Lansing, Michigan Abstract Nutrient extraction by saturated media methods using a DTPA solution was found to be dependent on the ultimate DTPA concentration in the media - extractant system. A test procedure was developed in which 30 ml of .005M DTPA was added to the extractant system per unit volume (100 cc) of media extracted. This method provides enhanced resolution of micronutrient cation levels in media with low to moderately high supplies of micronutrients. 82 83 Introduction The use of soil-less greenhouse growth media has been increasing steadily in the last two decades. Nutritional testing procedures developed for field soils are unsatis- factory for use with these media so testing procedures have been modified and new procedures developed to more adequately reflect media nutrient status. The Spurway or modified Spurway technique (10), the 1 :1.5 water extrac- tion (9) and saturated media extract (11) are three of the most widely used techniques of analyzing growth media for macronutrients. Little work has been done on testing the micronutrient status of artificial substrates. Coosmans (3) examined cor- relations between extracted levels of Fe, Cu, Zn, Mn, Cd, Pb and tomato tissue levels of these elements. Markus (7) used a modification of the DTPA test procedure of Lindsay (6) to test Mn, and Zn status of media for greenhouse tomato production. In an earlier paper (1) the authors evaluated the use of 15 extractants with saturated media methodology; DTPA was found to most closely match the evaluation criteria. The DTPA soil test procedure is based on the chemical properties of the soils to be tested (5,6). At equilibrium the micronutrient cation — DTPA complexes (Fe, Mn, Zn, and Cu) are highly favored in competition with Ca and Mg ions, 84 particularly below pH 7. The extractant system contains a buffer, triethanolamine and calcium chloride, to prevent the dissolution of calcium carbonate occluded micronutrients. Test results are affected by extractant pH, extractant concentration, temperature, and shaking time (13). A series of experiments were conducted to determine the effects of DTPA concentration, media moisture level, nutrient amendment level, and pH in three media. Materials and Methods Three media were used in these experiments: 1) Cornell mix; 50% Canadian sphagnum peat moss and 50% course ver- miculite (by volume): 2) Pine/peat; 50% pine bark and 50% Canadian sphagnum peat moss (by volume): and 3) Soil based; 33% sandy loam soil 33% Canadian sphagnum peat and 33% course vermiculite (by volume). Each was amended with 2.97Kg dolomitic lime, 1.2Kg superphosphate, and 2.97Kg gyp— sum per cubic meter of media. A center composite rotatable design with 3 variables (2) was used to examine the relationships between micronutrient amendment level, media moisture level, and DTPA concentration. Four replications of the center point were used for a total of 15 treatment combinations. This ex- periment was repeated with each of the three media. Tables 1 and 2 show the treatment combinations used. 85 Table 1 Variables examined using center composite experimental design and the levels used Coded level (Cochran & Cox 1957) Variable -2 -1 0 1 2 Rate Media amendments 0 1 2 3 4 Extractant cone. 0 .001 .0025 .004 .005 (M DTPA) Media moisture 10 22 40 58 70 (% sat) Micronutrients were added as a drench. Solutions con- taining the desired rate were applied to the media in a closed (non draining) system, until saturation was reached. The media was allowed to equilibrate with the micronutrient solution for . 24 hours, then was drained and purged with 1000 ml of deionized water per liter of media. Media moisture levels (percent saturation basis) were determined by saturating an oven dried sample with deionized water. Treatment rates for each media were determined on a volume media to weight water added basis. The charged medium used in the experiment was air dried to 10% moisture and deionized water was added as required. Media for each treatment was allowed to equilibrate with the added water for 1 hour prior to extraction. Extractions were done using standard saturated media procedures (11) except that DTPA of various molar concentrations was substituted for water as 86 the extractant. pH and soluble salts were determined for each sample. Macronutrient and micronutrient concentrations (P, K, CA, Mg, Zn, Fe, Mn, B, and Cu) were determined with a Beckman spectrametrics IIIA plasma emission spectrophoto- meter. 1000 ppm lithium chloride (lml of 10,000ppm LiCl/9m1 of sample) was added to the samples before analysis with the plasma to provide a more stable background matrix. Table 2 Media amendment rates used in center composite experiments (ppm in drench) Source level 0 1 2 3 4 Boric acid 0 3.6 9 14.4 18 Zn sulfate 0 24 60 96 120 Mn Sulfate 0 27.2 68 108 136 Fe sequestrene 330 0 19.2 48 76.8 96 pH effects were examined using media prepared for the center point of the center composite experiments (amendment rate 2). pH was altered by adding hydrochloric acid or sodium hydroxide to the media prior to extraction. Five treatments were applied: 1) 15ml 0.2N HCl / 100cc medium; 2) no treatment (control); 3) 5m1 0.2N NaOH / 100cc medium; 4) 7.5m1 0.2N NaOH / 100cc medium; and 5) 15ml 0.2N NaOH / 100cc medium. Samples from each of the 3 media were tested using standard saturation extract or a modified procedure. In the modified procedure 30ml of 0.005 M DTPA / 100cc of 87 media was added to the sample. Deionized water was used to complete the saturation of the media sample immediately after the DTPA solution was added. pH, soluble salts, P, K, CA, Mg, Zn, Fe, Mn, B, and Cu were analyzed as in the pre- vious experiments. Table 3 Nutrient amendment rates used to examine the relationships between DTPA extractions and standard water extractions (ppm of ion in drench) Treatment Nutrient Source 1 2 3 4 5 6 50 100 200 400 800 10 20 40 80 50 100 200 400 800 N calcium nitrate P potassium phosphate K potassium phosphate potassium chloride 000 01 Ca calcium nitrate 0 63 125 250 501 1001 Mg magnesium sulfate 0 25 50 100 200 400 Fe Iron sequestrene 330 0 6 12 24 48 96 Mn Manganese sulfate 0 8.5 17 34 68 136 B boric acid 0 1.1 2.25 4.5 9 18 Cu copper sulfate 0 5 10 20 40 80 Zn zinc sulfate 0 7.5 15 30 60 120 The relationship between the macronutrient content of the modified saturation extract and the standard procedure was examined. Media was saturated with solutions contain- ing increasing rates of nutrients (table 3). Excess solu— tion was allowed to drain from the bottom of the containers. The media was air dried to approximately 15% moisture, by weight, required for saturation. Extractions were done 88 using the procedures described in the pH experiment. Nitrate analysis was done on the extracts using an Orion specific ion electrode. Results and Discussion A key advantage of the saturation extract method is that media initial moisture levels do not significantly ef— fect test results. However when DTPA was used as the extractant, initial moisture content of the media was found to have significant effects on the extracted levels of many of the nutrients analyzed (tables 4—6). Regression models shown were those with the minimum residual sums of squares. No significant regression could be generated based on the soluble salts data in the Cornell mix. The calcium regres— sion is omitted in this media because the standards used in this analysis were found to be incorrect. With all three media there was interaction between ini— tial moisture level and DTPA concentration. The primary af- fect of initial media moisture level is to alter DTPA con— centration in the media slurry. This can be demonstrated by substitution of a combined term (DTPA added per 100cc of media) for the moisture level and DTPA concentration terms. Differences in the coefficients of determination (table 7) for these equations and those obtained using the separate model terms (tables 4—6) demonstrate that nearly all the variability accounted for in the complete 3 term model can 89 be accounted for using just 2 terms. Greater differences in coefficients of determination for the soil based mix (tables 6 and 7) may indicate that some portion of the variability in test levels in this medium is due to either initial moisture level or the experimental procedure used to produce this initial moisture level. Possibly the soil based medium did not reach equilibrium with the extractant system as quickly as the other media. The water added to the soil based media to increase the initial moisture level would, in this case be acting as an extractant for the hour of moisture level adjustment equilibration time, resulting in increased variability in the combined moisture level DTPA concentration term. The quadratic nature of the relationships between in- creasing DTPA concentration and increasing nutrient extrac- tion (tables 4—6) indicate that if sufficient DTPA is added to the system a non competitive equilibrium will be achieved and DTPA will remove all the extractable ions. However there are some practical limitations to the useful range of DTPA concentrations: 1) DTPA is only slightly soluble in water, 0.005 M was found to be a good working concentration; 2) The amount of liquid added to achieve saturation is limited; and 3) Macronutrient extraction is also affected by DTPA concentration. Slowly soluble forms of calcium, for example, can be rapidly dissolved by the DTPA solution causing inflated estimates of availability. 590 However published standards for macronutrient content of saturation extracts would still be valid with low DTPA concentrations. The effect of pH on the DTPA extractant / media system is critical, since unlike the test developed by Lindsay and Norvel (6), no buffer was used. The pH of 0.005M DTPA in deionized water ranged from 2.5 to 2.7. Therefore DTPA causes a consistent reduction in the pH test results com~ pared to those obtained with deionized water (S.E. + .ll)(figure 1). Since the relationship is linear it is pos~ sible to easily interpret DTPA extraction pH test results on the basis of the standards developed for the deionized water extraction (table 8). Increasing the media pH resulted in reduced macro- nutrient extraction with both DI water and DTPA extractants. Calcium extracted dropped from 700~800 ppm below pH 4 to under 300 ppm at pH 5.5 in all 3 media (figure 2). This trend was repeated with Mg and to a lesser extent P and K. In each case the main effects of pH, media, and extractant, and the (pH * media) interaction were sig- nificant at the 1% level. The lack of a significant (pH * extractant) interaction term indicates that media pH is not a concern in interpreting DTPA macronutrient test results with DI water standards. Micronutrient extraction in response to pH was not as straight forward as the macronutrients. Boron in the ex— 91 tracts declined linearly as pH increased (figure 3). The general trend was the same in all media, however more B was extracted from the Cornell mix than from the other media. There were no significant changes in the amount of Cu or Zn extracted as pH varied over the range tested (Figures 4,5). Solubility of these ions in the medium over the pH range examined can not be controlled by the same solid state and exchange equilibria that exist in soils since in soils these equilibria are pH sensitive. pH had no affect on the iron level in water extracts. DTPA extracts, on the other hand, contained. a quadrati— cally decreasing Fe level as pH increased (figure 6). Above a pH of 5.5 the rate of change of iron concentration in the DTPA extract as pH increased leveled off. Recommended pH ranges for soilless media are between pH 5.3 — 6.5 (11) so in most commercial media tests pH levels would be in the asymptotic region of the curve and thus would not cause substantial changes in the iron test results. A quadratic decline of manganese in water extracts oc— curred as pH increased, while manganese levels in the DTPA extracts increased as pH increased (figure 7). Increasing Mn in DTPA extracts appears to be due to competitive equi- libria within the DTPA extractant—media system. When high concentrations of potentially DTPA complexable ions (Zn=Cu>Fe>Mn>Ca>Mg) are present and available (low pH) the proportion of DTPA complexed with Mn is suppressed. Mn ex— tracted by DTPA chelation is thus reduced and Mn concentra~ tion in the extract is largely controlled by the Mn ex- tracted by water. As pH increases fewer competitive ions are present and a greater proportion of the DTPA in the system can be complexed with Mn. Manganese in the DTPA extracts, then increases greatly over the level of Mn in the water extracts. At low pH and high fertility this relationship would have to be considered in interpreting DTPA saturated media extracts. There is a high correlation between the soluble salts in the standard saturated media extract procedure and those in the DTPA modification (figure 8). Although the regres- sion equation is significant (table 8) the difference be— tween soluble salts obtained with these two procedures is so small that no correction is needed to use water stand— ards for interpretation of DTPA test results. High correlations were also found between macro— nutrients removed with DI water and those obtained with DTPA extractions. Linear regression equations (table 8) al- low the conversion of DTPA test results to their water ex- tract equivalents and thus the use of published standards. DTPA in the extractant system has no significant effect on the extraction of nitrate (figure 9). The correlation between calcium in DTPA extracts and standard water extracts is shown in figure 10. Despite the fact that calcium test levels were different for each of 93 the three media the correlation between DTPA extracts and water extracts was very good (r2 .914). Calcium levels in each of the 3 media are in different portions of the same curve. It appears that as the calcium levels increase Ca ions in solution become so common that due to mass action they occupy an increasing percentage of the DTPA chelating sites, hence the slope greater than 1. The concentration of other ions competing for DTPA exchange sites are likely to be very low compared to Ca concentrations so Ca test levels may be considered to be independent of competitive DTPA chelation affects. Magnesium levels likewise increase when DTPA is added to the extracting solution (figure 11). However there was no increase in the difference between DTPA and water ex— tractable Mg as Mg concentration in the media increased. It is possible that the DTPA in the extracting solution in- creases the extracted level of Mg simply by dissolution of dolomitic lime caused by the reduced pH of the DTPA extractant. This reduction in pH might also account for a portion of the enhanced CA extraction with the DTPA procedure. Potassium levels in the DTPA extracts were slightly higher than in the standard water extractions (figure 12). Although the regression equation is significant (table 8) differences are so small that no correction is likely to be needed to interpret DTPA test results using water 94 standards. The relationship between phosphorus in the DTPA ex- tracts and P in the standard saturated media extracts could not be adequately described by a single regression line. Separate correlations for each media were determined (figure 13). The amount of B extracted by each procedure was nearly the same (r2 = .990, slope = 1.09). The slope was very slightly (significant at the 5% level) greater than 1, however the difference is so small that it is not likely to be of concern in a routine testing program. Boron in DTPA extracts increased linearly with increasing B in the amend— ing drench (figure 14). The dose response for Zn and Cu were relatively linear up to ~60 ppm of applied micronutrient (figures 15 and 16). The linear range of these curves is 2 - 3 times the rate of nutrients normally applied (12). In the linear portion of the curve the DTPA extractable levels of these nutrients were very similar for each of the 3 media tested. For routine test interpretation a single standard curve would be adequate. The soil based medium dose response curve for iron was significantly different (1%) than that of the Cornell mix and the pine/peat media (figure 17). The strong quadratic response in the soil based medium may be the result of a strong affinity of this media to iron. When levels of ions 95 competing for DTPA binding sites are low and hence relative DTPA activity is high Fe-DTPA complexes are favored in relation to soil-Fe exchange. However when levels of com— peting ions are higher (relative DTPA activity lower) soil exchange may be competing more effectively for Fe than the exchange in the soiless media. The nonlinear portions of each of these response curves can be better understood by a close examination of the dose response curves for manganese (figure 18). In general Mn is the least competitive micronutrient cation for DTPA ex- change sites and as such is presumably the first of these nutrients displaced when DTPA exchange saturation is reached. Manganese test levels in standard water saturation extracts increased linearly as amendment level increased while DTPA extraction test levels of Mn were quadratic in nature (figure 18). At low to moderate amendment rates Mn concentration in the DTPA extractions increases rapidly in relation to the corresponding water extracts. Water extrac— tion of Mn continues to increase with little or no change in the corresponding DTPA extracts after the DTPA exchange is saturated. The relative contribution of DTPA-Mn chela— tion to the total Mn in the DTPA extract is thus reduced. For example, when 17 ppm of Mn was added to the Cornell mix in the amendment drench, 7% of the total Mn in the DTPA ex- tract can be accounted for on the basis of the water extraction. At the 136 ppm drench rate that percentage has 96 climbed to 46%. Based on the nonlinear regression curves generated for the data of this experiment, with 272 ppm in the drench Mn in the water extraction would equal the Mn in the DTPA extraction. Further increases in Mn in the media amendment drench would result in equivalent increases in' the DTPA extracts and the water extracts since water soluble levels of Mn would be the factor controlling ex- tracted Mn concentration. Summary The saturated media extract method (11) of testing greenhouse media is an attractive choice for a routine testing program, primarily because it gives reproducible results, and is simple and fast. The modified procedure developed to allow testing of micronutrient levels requires only 2 additional steps, adding the DTPA, and lithium chloride (if plasma emission spectroscopy is used for elemental analysis). The use of a single extraction pro» cedure is a great labor savings over the use of multiple independent extractions. Macronutrient content, pH and soluble salts test results in the DTPA modified extraction can be related to the test levels obtained with the standard procedure by simple equations. The differences for P, Mg, and K are small enough that conversion is probably not required and published standards for the saturation extract can be used. 97 Micronutrient cation extraction is greatly enhanced at low to moderate amendment rates by using DTPA in the ex- tracting system. This enhancement allows differentiation of media with low micronutrient content which was not possible with the standard procedure. At higher micronutrient levels competitive displacement occurs in the various DTPA-cation complexes and the extraction curves become nonlinear. As micronutrient cation concentrations in the media increase within this competitive range the portion of each of the extracted micronutrients that is attributable to DTPA chelation decreases. The water soluble and hence water ex— tractable portion of these ions then controls the extracted levels, and the extraction curves will reflect additional amended micronutrients. Since the nonlinear portions of the extraction curves occur at 2 or more times the usual recom- mended rate of amendment, the vast majority of media tested should fall within the linear range. Toxic levels of the micronutrient cations should be differentiable since they should fall within the range of the curve controlled by the water soluble levels of these ions. It is interesting to note that the results of competitive displacement of a micronutrient cation by high media concentration of another micronutrient cation may in some cases mimic the competi~ tive affects that occur in plants. For example plant uptake of Mn can be depressed in many crops by high levels of Zn, Fe, or Cu in the soil (8). This potential relationship be— 98 tween plant uptake and media test levels remains to be investigated. Based on the results of these studies the DTPA modified saturated media extract procedure described in this paper provides a method for a single extraction analysis of greenhouse media for pH, soluble salts, nitrate, P, K, Ca, Mg, Zn, Fe, B, Mn, and Cu when the micronutrient cations are present in low to moderately high amounts (less than 60 - 80 ppm in the amending drench if all 4 cations are balanced). If any micronutrient cation is present in very high concentrations in the medium or if all cations are present in excess of the equivalent of 60 - 80 ppm in the amending drench interpretation of the test results must be done guardedly since interactions could be expected to occur. References 1. Berghage, R. D., D. M. Krauskopf, D. D. Warncke, and I. Widders. 1985. Micronutrient Testing of Plant Growth Media: Extractant Evaluation. in press. 2. Cochran, W. G., and G. M. Cox. 1957. Experimental Designs. 2nd edition pp 335-375. 3. Coosemans, J., and P. Uyttebroeck. 1983. Comparisons of Different Extracting Methods for Heavy Metals From "Compost" Substrates and from Tomato Leaves and Fruits Grown on these Substrates. Acta Hort. 1332165—171. 4. Lindsay, W. L. 1979. Chemical equilibria in Soils. John Wiley and Sons. New York. 01 10. ll. 99 Lindsay, W. L., and W. A. Norvell. 1969. Equilibrium Relationships of Zn2+, Fe3+, Ca2+, and H+ with EDTA and DTPA in Soils. Soil Sci. Soc. Amer. Proc. 33:62- 68. Lindsay, W. L., and W. A. Norvell. 1969. Development of a DTPA Soil Test for Zinc, Iron, Manganese and Copper. Soil Sci. Soc. Am. J. 42:421-428. Markus, D. K., J. E. Steckel, and J. R. Trout. 1981. Micronutrient Testing in Artificial Mix Substrates. Acta Hort. 126:219-235 Mengel, K., and E. A. Kirkby. 1982. Principles of Plant Nutrition. International Potash Institute. Bern, Switzerland. Sonneveld, G., J. van den Ende, and P. A. Dijk. 1974. Analysis of Growing Media by Means of a l : 1.5 Volume Extract. Commun. in Soil Sci and Plant Anal. 5(3):183—202. Spurway, C. H. 1943. Soil Fertility Control For Greenhouses. Mich. State Coll. Agr. Expt. Sta. Spe- cial Bulletin No. 325. Warncke, D. D. 1980 Recommended Test Procedures for Greenhouse Growth Media. p. 31-35. In; W. C. Dahncke (ed). Recommended Chemical Soil Test Proce— dures for the North Central Region. Bulletin No. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. Warncke, D. D., and D. M. Krauskopf. 1983. Greenhouse Growth Media Testing and Nutrition Guidelines. Mi. State Univ. Exten. Bull. E—1736. Whitney, D. A. 1980. Micronutrient Soil Tests—Zinc, Iron, Manganese, and Copper. pp 18—21 in; W. C. Dahnke (ed). Recommended Chemical Soil Test Proce- dures for the North Central Region. Bull. No. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. 100 Table 4 Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concentration, and media moisture level, with a Cornell mix medium. Values presented are the significance levels of the (t) tests on the various least squares regression coefficients Variable pH salts P K Ca Mg Zn Fe B Mn Cu 1. Amendment * ** .3 .l * ** * level 2. DTPA ** ** * 2 .3 ** concentration 3. Moisture .2 .2 .2 * level 1 squared .1 ** .4 .1 * ** 2 squared ** ** ** 2 .3 2 * ** 3 squared 2 .2 2 1 X 2 *1: *2: 1 X 3 3 3 2 X 3 ** 3 2 l X 2 X 3 .1 .3 coefficient .917 ———— .766 .752 ———— .543 .896 .987 .912 .983 .878 of determination 101 Table 5 Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concentration, and media moisture level, with a pine bark medium. Values presented are the significance levels of the (t) tests on the various least squares regression coefficients Variable pH salts P K Ca Mg Zn Fe B Mn Cu 1 . Amendment ** ** * * ** . 2 ** * ** level 2. DTPA * .2 1k .2 ** ** .2 .4 * concentration 3. Moisture** ** .3 level 1 squared * ** .2 .2 .1 ** ** ** 2 squared .3 ** ** .1 ** .3 * * 3 squared * * x .1 l X 2 ** l ** l 2 * ** 3 l X 3 ** 2 .4 3 * * ** 2 X 3 ** * l l X 2 X 3 * .l .3 .l ** * coefficient .939 .889 .959 .822 .959 .862 .965 .899 .995 .982 .905 of determination medium. Table 6 Summary of regression analysis of center composite experiment examining the effects of media amendment level, DTPA extractant concentration, and media moisture level, with a soil based on the various least squares regression coefficients. Values presented are the significance levels of the (t) tests Variable pH Salts P K Ca Mg Zn Fe B Mn Cu 1. Amendment * * ** .2 * .1 ** * * level 2. DTPA ** .1 ** .2 ** ** concentration 3. Moisture * * * .l .3 * level 1 squared .2 ** .2 .3 * .4 .3 .4 .1 2 squared .3 1 ** ** ** .l .l .2 ** ** 3 squared .2 .2 .l .3 .4 .3 * ** l X 2 .4 .2 * l x 3 .2 1k * .3 at an: 2 X 3 1k ** .3 .2 ** l X 2 X 3 .2 ** coefficient .985 .848 .887 .759 .899 .865 .950 .951 .953 .997 .948 of determination 103 Table 7 Coefficients of determination for regression equations using mg of DTPA/100cc media rather than DTPA concentration and initial media moisture level with data obtained from center composite experiments examining the effects of media amendment level, DTPA extractant concentration, and media moisture level. Test Media Cornell mix Pine/peat Soil based pH .919 .479 .737 Soluble salts ns .769 .723 P .752 .807 .610 K .673 .765 .441 Ca —-— .841 .583 Mg .547 .851 .633 Zn .898 .957 .835 Fe .986 .826 .656 B .895 .959 .940 Mn .976 .958 .720 Cu .749 .964 .741 104 Table 8 Conversion equations to convert from standard saturation extract test results to modified DTPA saturation extract test results. Test Equation Water extracts DTPA extracts pH pH = .142 + .930 X pH Salts salts (m8) = -.150 + 1.02 X salts Nitrate Nitrate (ppm) : .794 + 1.00 X nitrate Water extracts (ppm) DTPA extracts (ppm) Phosphorus (Cornell) P = ~29.7 + 1.3 X P (Pine peat) P = 8.85 + .763 X P (Soil base) P = 1.25 + .300 X P Potassium K = —.498 + .980 X K Calcium Ca = -19.1 + .877 X Ca Magnesium Mg : —l6.5 + .980 X Mg 105 Figure 1 Correlation between pH of standard water saturated extracts and DTPA modified extractions in 3 media with pH adjusted by addition of either 15 ml 0.2 N HC1,no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 ccmedia sample prior to extraction. 106 mcosoobxo $53 eo Ia . _ . _ . _ . _ . a) N l. 1m 1w Ammmfno 0c: coimwocaoe I: IN. vomon __om < Hooa\ocE n x_E =mEoo x suogioonxe VCLLQ Dto Hd it 107 Figure 2 Calcium extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 m1 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction. 108 282 83838 .6 Ia —co If) —~.+ :8 (Eb zom c803 2.3 EB 05a 2863 .6500 Stb =oEoo e303 I ”I I III ICON [00¢ loom oom ioolixe u! (wdd) 03 109 Figure 3 Boron extracted in standard water saturated extracts or modified DTPA extracts of 3 media with pH adjusted by addition of either 15 ml 0.2N HCl, no treatment, 5, 7.5, or 15 ml 0.2 N NaOH / 100 cc media sample prior to extraction. 110 —I\ 282 aflosam so Ia :8 152 rates. Significance appears to be primarily caused by tissue levels in plants grown at the highest CLF rate. These tissues were visibly necrotic so it is not surprising that significant effects on tissue composition should occur. Increased rates of "STEM" application (trial 3) caused significant increases in plant tissue and media extract levels of Zn, Mn, Cu, and B. There were significant treat- ment effects on Fe in media extracts but no significant ef- fect on plant tissue levels. Effects on media Zn and Mn were reflected in plant tissue levels of these nutrients (r2=.528, and r2=.538 respectively). Copper uptake by plants did not correlate with media extract levels. In— creases in plant Cu uptake may in some way be actively limited, occurring only at low rates of added Cu. The maxi— mum uptake in this experiment was observed to be at about 10 ppm of Cu in the media drench. Copper in the media ex- tracts increased with increasing application rates, however plant tissue levels did not reflect this increase. Boron levels in plant tissues correlate to media levels in the first sampling (r2=.585), but there were no sig— nificant treatment effects on tissue B levels in the second sampling. The initial correlation may reflect a high ini— tial uptake of B when the drench was applied. Dilution and or leaching of B in the media through subsequent waterings may have reduced the B content of the media. Since sampling date did not have a significant effect on B in the media 153 extracts it must be assumed that this dilution or leaching occurred prior to the first sampling. There were no deficiency or toxicity symptoms observed on any plants in this trial and there were no significant differences in dry weights attributable to "STEM" applic— ations. Marigold plant tissue micronutrient levels (experiment 2) also correlate well with DTPA micronutrient extractions (Table 7). Correlations between B, Fe, Zn, and Mn in ini— tial media extracts and in plant tissue at the end of the experiment were, r2=.846, r2=.650, r2=.663, and r2=.863 respectively. Final media micronutrient extract level and plant tissue test level correlations were very similar, r2=.837, r2=.539, r2=.7l5, and r2=.902. Parallel regressions with separate intercepts for each media, and in the case of Zn and Fe each cultivar as well, were used to fit these relationships. The media used significantly affected media extract results in all cases but B extraction in the final media analysis. With the exception of Ca, media effects~ were also highly significant in the results of plant tissue analysis. Cultivar differences in plant tissue levels of Zn, Fe, B, Ca, and K were significant. Zinc and Fe tissue levels in the cultivar "Yellow boy" were on average 2 to 3 times less than in the cultivar "Apollo". The grand mean levels were 134 ppm vs 337 ppm for Zn and 165 ppm vs 383 ppm for Fe. Differences in B, Ca, and K tissue levels between 154 cultivars were not quite as dramatic. Mean nutrient levels for "Yellow boy" and "Apollo” were respectively; B: 63 ppm and 58 ppm, Ca: 2.0% and 2.3%, and K: 8.4% and 9.1%. Media sampled from cells in which "Apollo" was grown contained significantly less K than media from cells used to grow "Yellow boy" (means; 60 ppm vs 66 ppm). Media supplies of other nutrients were not significantly effected by dif— ferential uptake. In general media nutrient test levels were lower in the samples taken at the end of the experiment than those at initial sampling. The main effect of time (analyzed as a split split plot) was significant for Fe, B, Mn and Cu (Table 8). In each case there was a general overall reduc- tion in test levels from the initial sampling to the final sampling. Coefficient of determination for parallel regres— sions of Fe, Zn, Mn, and B were r2=.888, r2=.922, r2=.935, and r2=.529. The lack of significance of the main effect of time for Zn and the highly significant regression r2=.922 can be explained by examining the time x micronutrient treatment x media interaction. There were essentially no differences in initial test levels and final test levels of Zn for the soil base mix or the pine peat mix, while the Zn levels in the Cornell mix decreased with time for all treatments. For Fe, Mn and B this interaction was due to the relatively more rapid decline in the test levels of the highest amendment rates in comparison with the lower amend- 155 ment rates. Parallel regressions were developed to relate the amendment rate of Fe, Zn, B, and Mn to initial media extract levels of these nutrients. The coefficients of determination for these curves were r2=.872, r2=.860, r2=.788 and r2=.861 respectively (Table 7). The plant growth response as measured by differences in accumulated dry weight was very small and erratic. Significant regressions (based on F test statistics) could be generated for B, Fe, and Zn (Figures 1 — 3). No significant regression could be generated for Mn. The coefficients of determination for these regressions were r2=.376, r2=.478 and r2=.275. In each case the trend is toward decreasing dry weight with increasing media amendment rates of these nutrients. The small growth response and high variability make calibration of the test procedure diffi- cult. The lack of a significant decrease in growth and ab- sence of deficiency symptoms in marigolds when no B, Fe, Mn, or Zn was added to the media indicate that the deficiency threshold (critical level) lies below the levels naturally present in the media tested, and thus below 1 ppm of Zn, 13 ppm of Fe, .5 ppm of B and 4 ppm of Mn in the media extracts. In most of the media cultivar combinations there appears to be a small decrease in growth (dry weight) at the lowest amendment rates however this is not significant. The affects of low micronutrient amendment levels may be masked by genetic and environmental variation. For this 156 reason it is recommended that test levels of B, Zn, Fe, and Mn should be higher than the lowest levels obtained in this experiment. The small general decline in plant dry weight accumulation with increasing amendment rate shows that the B, Zn, and Fe, at the highest levels used were detrimental to plant growth. Recommended maximum DTPA extraction test levels for Zn, Fe, and B in marigold are given in Table 12. Plant tissue levels of Fe, Zn, B, and Mn in poinsettia were correlated with the media test results of the DTPA extractions. Mid crop (ll/2/84) plant tissue and media test level correlations for B and Mn were as good as those in the final sampling, r2=.896, .809 and r2=.925, and .729 respectively. Mid crop Fe and Zn media and plant test cor- relations were lower than correlations at the final sampling, r2=.268, .681, and r2=.556 and .816. It is likely that plant tissue levels of Fe and Zn in the mid crop sam— pling were effected by initial plant status (prior to treat— ment applications) of these nutrients. With increased treat— ment exposure the plant tissue levels of these nutrients reflect treatment effects to a greater degree and initial nutrient status to a lesser degree. The early high correla— tion between B and Mn in plant and media tests might indi- cate a more rapid uptake of these nutrients, possibly the result of a lower relative initial nutrient status of these nutrients, or a lower initial variability. Correlations between mid crop media samples and final 157 plant tissue nutrient levels were similar to those for final media and final plant test correlations (Tables 9 and 10). This can in part be explained by the lack of any significant simple effect of sampling time (Table 11), for Zn, Fe, B, or Mn in media extracts. This is in contrast to the sig— nificance of sampling time in experiment 2. The results of these 2 experiments seem to indicate an initial, possibly rapid, decline in media nutrient levels after which the levels remain relatively constant over extended periods. A further observation is that the amount and rate of decline in the nutrient media test levels is related to the initial media amendment or test levels. This relationship is demonstrated by the quadratic correlations between mid crop sampling time and final sampling time in experiment 3 (Table 10). And the significant time X treatment interactions in both experiments 2 and 3. There were significant media effects on both media ex- tractions and plant tissue tests for Zn, Fe, B, and Mn. Cultivar effects were significant for Zn, and B in the plant tissue. The correlation coefficients for regressions between plant dry weight and nutrient amendment rate were r2=.615 for Fe, r2:.400 for Zn, r2=.638 for B and r2=.318 for Mn. The quadratic nature of these curves permits the calibration of the media test to plant growth (dry weight). For example if the optimum range is defined to be approximately + or — 158 5 N of the predicted maximum dry weight, the optimum dry weight range will fall between approximately 5 and 20 ppm of B in the amendment drench. This level corresponds to a mid crop media test value of .65 to 2.2 ppm and a plant tissue test of 69 to 228 ppm. Tissue test levels for poinsettia given by Mastalerz (4) list 30 to 100 ppm as the "normal range" and 200+ as "excess range". These relationships for Zn Fe, and Mn, are presented in Tables 9 and 10, and Figures 4—7. Summary The modified DTPA saturated extract media test proce— dure results are highly correlated with tissue test results in poinsettia, marigold, and Chrysanthemum. The calibration curves presented for poinsettia test levels provide a basis for the prediction of optimum DTPA media test levels. Op- timum media test levels can also be predicted based on plant tissue test levels of the nutrient through the use of DTPA media test and plant tissue test correlation regression equations. The lack of a quadratic growth response in the marigold experiment means that no low end to the optimum range can be experimentally defined. However media test and plant tissue test correlations can be used with published tissue test standards to calculate a theoretical low end to the optimum range. This method can also be used to develop standards for the Chrysanthemum. Recommended media test 159 levels for Zn, B, Fe, and Mn in poinsettia, marigold and Chrysanthemum are presented in Table 12. Based on the evidence presented in this and previous studies a non equilibrium DTPA saturated media extract pro— cedure is the media test of choice for the analysis of growth media. It is simple, quick, and allows simultaneous testing for pH, salts, N03, Fe, Zn, Mn, B, P, Cu, K, Ca, and Mg. The macronutrient test levels can be easily compared to normal saturated extract test levels, and micronutrient test levels of Fe, Zn, Mn, and B are highly correlated to plant tissue test levels. References Cited 1. Coosemans, J., and P. Uyttebroeck. 1983. Comparisons ' of Different Extracting Methods for Heavy Metals From "Compost" Substrates and from Tomato Leaves and Fruits Grown on these Substrates. Acta Hort. 1332165—171. 2. Ecke, P. Jr., and 0. A. Matkin. 1976. The Poinsettia Manual. Paul Ecke Poinsettias, Encinitas, Ca. 3. Markus, D. K., J. E. Steckel, and J. R. Trout. 1981. Micronutrient Testing in Artificial Mix Substrates. Acta Hort. 126:219—235. 4. Mastalerz, J. W. 1977. The Greenhouse Environment. John Wiley and Sons. New York. 5. Warncke, D. D. 1980. Recommended Test Procedures for Greenhouse Growth Media. p. 31-33 in; W. C. Dahncke (ed). Recommended Chemical Soil Test Procedures for The North Central Region. Bulletin No. 499 (revised) North Dakota Agr. Exp. Sta. Fargo, North Dakota. 160 Table 5 Analysis of variance for Chrysanthemum tissue / DTPA media test correlation experiments. Trial 1 Cornell mix with CLF micronutrients, trial 2 pine/peat media CLF micronutrient applications and trial 3 pine/peat with Peters "STEM" micronutrient applications. Variate Trial 1 Trial 2 Trial 3 Source of variation Time Interaction Time Interaction Treatment Treatment Treatment Zinc Plant ** NS NS NS NS ** NS Media NS ** NS NS ** ** NS Iron Plant ** ** NS NS NS NS NS Media ** ** NS ** NS NS NS Manganese Plant ** ** ** ** * ** NS Media NS ** NS ** * ** NS Boron Plant ** ** ** ** ** ** ** Media NS *¥ NS ** NS ** NS Copper Plant NS X NS NS ** ** NS Media NS NS NS NS NS ** NS Phosphorus Plant ** ** X ** ** NS NS Media ** ** ** NS ** NS NS Potassium Plant ** ** * NS ** * NS Media ** NS NS NS ** NS NS Magnesium Plant ** NS NS * ** NS NS Media * NS NS NS ** NS NS Calcium Plant ** ** NS NS ** NS NS Media ** NS NS NS it NS NS Plant total dry weight ** ** NS ** ** NS NS Media pH u M NS an: n NS NS Media soluble salts NS NS NS ** NS NS 161 Table 6 Correlation equations between Chrysanthemum tissue tests and DTPA saturated media extracts. Trial 1 Cornell mix with CLF micronutrients, trial 2 pine/peat media CLF micronutrient applications and trial 3 pine/peat with Peters "STEM" r2 .700 .452 .753 .582 .935 .971 .538 .528 .445 micronutrient applications. Trial 1 Correlation equations Plant (Fe) (B) (Fe) (Mn) (8) (Mn) (Zn) (E) II II II Media 146 - .687(Fe) + .02(FE)2 121 + 13.1(Mn) - .19(Mn)2 ~104 + 211.9(3) - 9.7(B)2 Trial 2 217 + 3.81(Fe) 64 + 3.0(Mn) -35.8 + 305.7(3) Trial 3 130 + 21.3(Mn) 81.9 + 2.4(Zn) 31.6 + 24.3(B) Intercept adj. time 2 (15.3) time 3 (123.9) time 2 (—41) time 3 (33) time 2 (64) time 3 (211) time 2 (-77) time 2 (-22.5) time 2 (—l3.4) 162 Table 7 Correlation equations for relationships between marigold tissue tests, DTPA saturation extracts, and micronutrient drench rates for 3 media and 2 cultivars with parallel regression intercept adjustments when needed for media or cultivar r2 Correlation equations Intercept adj. Final media - final plant sample correlations Plant Media .539 (Fe) = —372 + 30(Fe) — .212(Fe)2 media 2 (114) media 3 (-260) cv 2 (258) .715 (Zn) = —161.8 + ll(Zn) media 2 (173) media 3 (—209) cv 2 (469) .902 (Mn) 2 137 + 17.9(Mn) — .075(Mn)2 media 2 (~71) media 3 (—616) .837 (B) = —4.98 + 161(8) — 33.4(B)2 media 3 (-28.4) cv 2 (9.1) Initial media — final plant sample correlations .650 (Fe) 2 -184 + 18.8(Fe) — .074(Fe)2 media 3 (—413) cv 2 (234) .663 (Zn) = -222 + 8.2(Zn) media 2 (388) cv 2 (470) .863 (Mn) = 147.3 + 6.65(Mn) media 2 (-74) media 3 (~300) .846 (B) = 15.2 + 64.4(B) - 5.38(B)2 media 3 (18.4) Initial media —final media sample correlations final initial .888 (Fe) = —6.8 + .99(Fe) — .0032(Fe)2 media 3 (6.37) .922 (Zn) = -8.1 + 1.7(Zn) - .0064(Zn)2 media 2 (7.5) .935 (Mn) = .037 + .583(Mn) media 3 (23.9) .529 (B) = .136 + .326(B) media 3 (.739) Initial media sample — micronutrient treatment rate correlations media ose .872 (Fe) = 27.3 + .150(Fe) — .00038(Fe)2 media 2 (-16) media 3 (-6.1) cv 2 (~4.3) .860 (Zn) = 12.7 + .68(Zn) — .001(Zn)2 media 2 (—11) media 3 (~21) .861 (Mn) = 12.8 + .49(Mn) — .00062(Mn)2 media 3 (26) .788 (B) = .945 + .05(B) media 2 (-.71) media 3 (-1.32) Plant dry weight - micronutrient treatment rate correlations .478 dry weight = .69 — .00059(Fe) cv 2 (.15) .275 dry weight = .73 - .0003(Zn) cv 2 (.12) .376 dry weight = .72 — .0008(B) media 2 (*.13) cv 2 (.19) 163 Table 8 Analysis of variance of split split plot experiment examining the relationships between marigold tissue tests and DTPA saturated media extracts of 3 media. Variate Source of variation Initial media samples (24 micronutrient treatment rates) Treatment Media Interaction Zn *** *** ** Fe *** *** ** B *** *** *** Mn xxx xxx NS Cu xx xxx NS P xx xxx NS Mg xx xxx NS Ca * *** NS K NS *** NS Plant dry weights (24 treatments T, 3 media M, and 2 cultivars C) T M C (T x M) (T x C) (M x C) (T x M x C) dry weight * *t ** NS NS NS NS Final media samples, and final plant tissue samples (9 micronutrient treatments T, 3 media M, and 2 cultivars C) T M C (T x M) (T x C) (M x C) (T x M x C) Zn (plant) *** *** *** *** *** *** *** Fe (plant) *** *** *** ** ** *** * B (plant) *** *** ** NS NS NS NS Mn (plant) *** ** NS *** * NS * Cu (plant) xxx x NS x NS NS NS P (plant) NS xxx x NS NS xx NS Mg (plant) ** ** NS NS NS ** NS Ca (plant) xxx NS xxx NS xx NS NS K (plant) ** *** *** NS NS *** NS Zn (media) *** *** NS NS * * NS Fe (media) *** *** NS NS NS NS NS B (media) * NS NS NS NS NS NS Mn (media) *** *** NS *** * NS NS Cu (media) *** *** NS *** NS NS NS P (media) NS *** NS NS ** NS NS Mg (media) NS ** NS NS ** NS NS Ca (media) NS *** NS NS ** NS NS K (media) * *** *** ** NS ** NS Initial and final media with time as a split split plot. (9 micronutrient treatments TR, 3 media M, 2 cultivars C, and 2 times T) T TR TxTR M C TxM TRXM TXC TRxC MxC TxTRxM Zn ‘ NS *** NS *** NS *** *** NS NS NS *** Fe * *** NS *** NS *** *** NS NS NS *** B * *** NS NS NS NS NS NS NS NS NS Mn *** *** *** *** NS *** *** NS NS NS ** Cu *** *** ** *** NS *** ** NS NS NS *** P NS NS NS *** NS *** ** NS NS NS ** Mg NS NS NS *** NS *** NS NS NS ** NS Ca NS NS NS *** NS *** NS NS NS NS NS K NS *** *** *** * *** *** *** NS *** *** 1134 Table 9 Correlations between poinsettia tissue samples and DTPA saturated extracts in 3 media with 2 cultivars. Intercept adjustments provided for parallel regressions where needed. mid crop tissue tests and mid crop extracts r2 plant tissue (ppm) DTPA extracts (ppm) .263 Fe : 39.3 + 1.06(Fe) .631 Zn : 17.6 + 3(Zn) - .022(2n)2 .896 0 : -31.9 + 196(8) - 15.501)2 .309 Hn : 46.3 + 7.13(nn) plant tissue (ppm) amendment drench (ppm) .230 Fe : 96.5 + .116(Fe) .773 Zn : 24.9 + .27(Zn) .905 3 : —5.52 + 12.0(0) .329 Mn : 50.32 + 1.4(nn) DTPA extracts (ppm) amendment drench (ppm) .896 Fe = 6.0 + .ll4(Fe) .905 Zn = *2.97 + .29(Zn) .882 B = .10 + .11(B) .852 Hn = 2.02 t .18(Hn) plant dry weight (9) amendment drench (ppm) .255 weight : 4.69 + .02(Fe) + .0007(Fe)2 NS Zn ------------------------ .459 weight : 5.1 + .09(B) -.003(0)2 NS Mn ------------------------ end crop plant tissue and end crop DTPA extracts plant tissues (ppm) DTPA extracts (ppm) .556 Fe ' 66 + 2.39(Fe) .816 Zn 25.6 + 2.07(Zn) ' .012(Zn)2 .925 0 —13.3 + 151.4(3) .792 Hn 7.54 + 12.52(Hn) - .1(Hn)2 plant tissue (ppm amendment drench (ppm) .590 Fe 87 + .l7(Fe) .316 Zn 30.7 + .275(2n) .905 3 16.4 + 10.610) .745 Hn 43.1 + 1.04(nn) .866 Fe 8.96 + .069(Fe) .971 Zn -.86 + .277(Zn) .931 3 .252 + .079(8) .882 Hn 4.08 + .158(Hn) ) DTPA extract (ppm) amendment drench (ppm) plant dry weight (g) amendment drench (ppm) .610 weight 7.74 + .015(Fe) - .00008(Fe)2 .403 weight 6.85 + .02(Zn) - .00007(Zn)2 .6” wiflt 766+ JINB)-.0M(M2 .277 weight 7.13 + .018(Hn) - .00005(Hn)2 intercept adj. media 2 (9.9) media 3 ('14.3) media 2 (9.36) cv 2 (14.5) CV 2 (59.9 media 2 (61.1) media 3 (“47.2) media 2 (10.5) media 2 (16.7) cv 2 (11.6) cv 2 (50.8) media 2 (63.3) media 3 (3.31) media 2 (7.33) media 3 (11.1) media 2 (-.30) media 3 (10 9) cv 2 (-4 04) media 2 (".607) media 2 (14.3) cv 2 (21) media 2 (33.2) media 3 (~57.5) cv 2 (27) media 2 (8.9) media 3 (-9.5) cv 2 (9.4) media 2 ('38) cv 2 (21) media 2 (32) media 2 (-2.6) media 3 (3.1) media 2 (2.4) cv 2 (2.3) media 2 (-.419) media 3 (6.4) media 3 (.85) cv 2 (-.S72) media 2 (-.52) media 3 (1.1) cv 2 (.66) media 3 (.715) media 3 (1.12) 1635 Table 10 Correlations between poinsettia tissue samples and DTPA saturated extracts in 3 media with 2 cultivars. Intercept adjustments provided for parallel regressions where needed. mid crop DTPA extracts and end crop plant tissue r2 plant tissue (ppm) DTPA extracts (ppm) Fe : 65.0 + 3.36(Fe) - 0.33(Fe)2 media 3 (- 8. 26) .372 Zn : 17.3 + 3.89(Zn) + .03(2n)2 media 3 (- 66 6) cv 2 (12.5) .911 3 = —7 34 + 174(3) - 13.7(3)2 cv 2 (23.1) .734 Hn : 13.7 + 10.5(Hn) - .07(xn)2 media 3 (~55. 9) cv 2 (40.11) mid crop DTPA extracts and end crop DTPA extracts end crop extracts mid crop extracts .360 pH : .566 + .91(pH) .293 salts : .57 + .40(salts) .365 Fe (ppm) : 1.14 + 1.22(Fe) — .012(Fe)2 media 2 (-3 3) .397 Zn (ppm) : -l.26 + 1 74(2n) - .0036(2n)2 media 2 (4 7) cv 2 (5. 0) .947 8 (ppm) : .08 + 1.22(B) - .089(B)2 media 2 (-. 26) media 3 (-.19) .903 Hn (ppm) : 1.52 + 1.11(Hn) — .0039(nn)2 media 2 (-3 .1) mid crop plant tissue and end crop plant tissues end crop tissue (ppm) mid crop tissue (ppm) .686 Fe 2 22.4 + .744(Fe) media 3 (6.89) .826 Zn : -2.36 + 1.45(Zn) - .003(Zn)2 media 2 (-6.7) media 3 (-9 9 .927 3 = 9.03 + 1.3(3) - .00069(3)2 media 2 (~35.6) .832 Hn : -18.3 + 1.2(Hn) - .00075(Hn)2 media 3 (-30) cv 2 (27.6 166 Table 11 Analysis of variance of plant tissue and DTPA saturated media extract test results from a split plot experiment with 3 media (H), 2 poinsettia cultivars (C), 2 sampling times (T), and 13 micronutrient treatments (TR). Variate source of variation - T TR H c TxTR TxH TRxH Txc TRxC ch TxTRxH TxTRxH TxTRxC TxHxTRxC media tests » pH 1” 1‘" *1” NS NS m NS NS NS * NS NS NS NS salts 1‘" NS “1* * ’1 *H ’4 NS NS 4‘ NS NS NS NS Zn NS *1” “1' NS NS NS *1” NS NS * * NS NS NS Fe NS 1‘“ “6* NS “4 NS * NS NS NS NS NS NS NS B NS “1* *1” NS “1* NS “1* NS NS NS NS NS NS NS Hn NS "4' fit NS NS 4 Hi NS * Ht NS NS N3 in P NS “1* *1“ NS NS 1” NS NS NS 1" NS NS NS NS 119 * NS *1“ NS NS *4 NS NS NS NS NS NS NS NS Ca N tit in NS Ht Ht 1“ NS NS NS *4 NS NS NS K *1" NS *1” NS 1‘“ NS "1* NS "1* NS NS NS NS NS plant tissues dry weight *1” *1” *1” NS 1‘ NS *1” NS *** NS NS NS NS NS Zn NS *1” "i "1* NS NS "1* NS NS ”1* NS NS NS NS Fe NS "1* *1” NS NS NS *1” NS NS ”’4 NS NS NS * 8 NS t“ 3 “4 “8 NS Ht NS 1“ Hi NS H NS ”8 Mn * *1” NS NS *1“ NS 1‘" NS NS *1” NS NS , NS NS P *1” *1” ”1‘ NS NS * NS *1” NS NS NS NS NS NS 119 NS NS *** *1) NS NS NS NS NS NS NS NS NS NS [:a it fit NS “‘8 8H 8 NS “8 NS Ht 8 NS H N3 |( 1’“ ““1 1*“ *** NS NS NS NS NS NS NS NS NS NS 167 Table 12 Recommendations for B, Zn, Fe, and Mn DTPA saturated media extract test values for Chrysanthemum, marigold and poinsettia. Chrysanthemum Ranges based on correlation regressions between media test levels and plant tissue test levels and plant tissue test standards from Mastalerz (7). Nutrient Adequate range B .2 ppm - .7 ppm Mn 43 ppm — 65 ppm Fe ———————— Zn -------- Marigold Maximum values based on a 5% reduction in predicted maximum plant dry weight. Nutrient Optimum range Fe - 80 ppm Zn — 120 ppm B - 4.5 ppm Mn - Poinsettia Optimum range defined as a 5% reduction of predicted maximum plant dry weight. Nutrient Optimum range Fe 10 ppm - 24 ppm Zn 16 ppm - 58 ppm B 0.7 ppm — 2.2 ppm Mn 16 ppm 52 ppm 168 Figure 1 Iron calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight. 169 (61.1.1) 3111619791 [Up Wold AEQQV 50:96 ycoEvcmEo E mm 00... com com 03 0 CO. _ . _ . _ . q _ . O 9.? mom ” aw woe ON.QJ . lv/ ” at .. .O/w 1.00 0001 .v . as... mom 9...? av _ . omdm . a H iomr . m .7 u . cm 01 Av [at 393., b not: . . . b wc a on 01. “ 05 100— cm. u of (wdd) sioonxe oipew U1 9;; 170 Figure 2 Zinc calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight. 171 (6111) 31115191171 Mp Wold AEaav 50:96 EoEccmEo E cN 00¢ com com 00— o OO. _ . _ . . _ L _ . o 8.01 low 8.01 u e? .. ” oz. 10... on? 6663 .. ” 1V0! low 0*.3 u .\\¢ 1 u 099 ROG 0001 n .. 0903 m WOOF 8.3 ”m8.” m mom? ” ~ 399: ED “CUE .. om. . o: (wdd) siooiixe oipew ui UZ 172 Figure 3 Boron calibration curves for DTPA modified saturated media extracts and marigold plant growth. Maximum recommended test level defined as the level which corresponds to a 5% reduction in predicted plant dry weight. 173 (6111) 8111519713 Alp 11101:) cog“ om AEaav 50:96 59565950 :_ m 00 06 ON 5 - — b — b o 201 8.? one. 3.3 8.01 8.0.. OBAYJ P- - - - - QNVJNNW‘ w£@_@3 an wCO—Q 0mg“ (wdd) siooiixa [)1pr 111 a 174 Figure 4 Iron calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight. 175 (6111) 11151913 10p iuold AEaav 50:96 50:66:50 :_ 0..._ com com L GDP 5 0&T oz..- OAT OAT 05.065 296; be age F""" 0A; (wdd) siooJixe ogpaw u! 9;] mom mom 100? nop— now? nonp 10¢F (wdd) senssp, iuold u! 9:] - 176 Figure 5 Zinc calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight. 177 (611.1) 111619791 Mp iuold AEaav 50:96 #:9565950 :_ :N can owm cm? 0 . H 69” o J n 000 . . O/O . o.Ni H 3+0 “ . oov . 0.”! ” «wax u u as . Devi ” n60 ” u o 8. _ O.WJ . IVO/ . . 0.0/v.0. ” 0.0. . 6% . n 3...... n 0 . 0.? u 0’ ” QT . 6 “8.4.1.3. 296; be anm a 1 7 (wdd) siooJixa oipaw U1 uz (wdd) sansspr iuold 111 uz C 178 Figure 6 Manganese calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight. 179 (6111) 1116197111 Kip lUDId 0 c5 0. O. 0. 0. ‘— J ? J (lo J j. l (1’ 4 y\ [\ (‘1 11 N (I 4.: .C.’ .9 0 3 b T, 4.! C .2 CL. 1 1 j j C O O (.0 l0 ‘1' (wdd) siomixe o1paw u1 uw 1 ' l ' l f l ' fl 0 O O O O O O O 0 fi' I") N '— (wdd) senssp, lupld u1 uw Mn in amendment drench (ppm) 180 Figure 7 Boron calibration curves for DTPA modified saturated media extracts and poinsettia plant growth and tissue nutrient levels. Optimum range defined as the range which is within 5% of the predicted maximum dry weight. 181 (611.1) 11161913 Kip 41101:) 50:96 05086:ch :_ AEQaV m om 0+ on ON 0— o h . _ b _ . _ .0 4 _ . . _\. _ .7 x .3 _ o/we .. _ l . _ on... _ . m1 .aLQ a0). _ 1N .Qv n%@ _ 5 6 7w _ . \6 or \ 5.064 _ an 1 _ _ . . _ hm _ _ 16 1 _ _ . _ . mi _ _ 1m . 86.365. 3,596; e to; .. m _ _ c m siopiixe o1pe1.u 111 (LUdd) 8 no loop ICON 7000 100+ room 100w senssp, lUDld (wdd) g ““1111111‘1111111111111111111118)“