Identification of Gibberellins in Spinach and Effects of Light and Darkness on their Levels

The endogenous gibberellin (GA) content of spinach (Spinacia oleracea) was reinvestigated by combined gas chromatography-mass spectrometry analysis. The 13-hydroxy GAs: GA₅₃, GA₄₄, GA₁₉, GA₁₇, GA₂₀, GA₅, GA₁, GA₂₉, and GA₈; the non-3, 13-hydroxy GAs: GA₁₂, GA₁₅, GA₉, and GA₅₁; and the 3β-hydroxy GAs: GA₄, GA₇, and GA₃₄, were identified in spinach extracts by comparing full-scan mass spectra and Kovats retention indices with those of reference GAs. In addition, spinach plants contained GA₇-isolactone, 16,17-dihydro-17-hydroxy-GA₅₃, GA₂₉-catabolite, 3-epi-GA₁, and 10 uncharacterized GAs with mass spectra indicative of mono- and dihydroxy-GA₁₂, monohydroxy-GA₂₅, dihydroxy-GA₂₄, and dihydroxy-GA_g. The effect of light-dark conditions on the GA levels of the 13-hydroxylation pathway was studied by using labeled internal standards in selected ion monitoring mode. In short day, the GA levels were higher at the end of the light period than at the end of the dark period. Levels of GAs at the end of each short day were relatively constant. During the first supplementary light period of long day treatment, GA₅₃ and GA₁₉ declined dramatically, GA₄₄ and GA₁ decreased slightly, and GA₂₀ increased. During the subsequent high-intensity light period, the GA₂₀ level decreased and the levels of GA₅₃, GA₄₄, GA₁₉, and GA₁ increased slightly. Within 7 days after the beginning of long day treatment, similar patterns for GA₅₃ and GA₁₉ occurred. Furthermore, when these plants were transferred to darkness, an increase in the levels of GA₅₃ and GA₁₉ was observed. These results are compatible with the idea that in spinach, the flow through the GA biosynthetic pathway is much enhanced during the high-intensity light period, although GA turnover occurs also during the supplementary period of long day, both effects being responsible for the increase of GA₂₀ and GA₁ in long day.     

Qualitative and Quantitative Analysis of Endogenous Gibberellins in Onion Plants and Their Effects on Bulb Development

Evidence has been reported that bulb development in onion plants (Allium cepa L.) is controlled by endogenous bulbing and anti-bulbing hormones, and that gibberellin (GA) is a candidate for anti-bulbing hormone (ABH). In this study, we identified a series of C-13-H GAs (GA₁₂, GA₁₅, GA₂₄, GA₉, GA₄, GA₃₄, and 3-epi-GA₄) and a series of C-13-OH GAs (GA₄₄, GA₂₀, GA₁ and GA₈) from the leaf sheaths including the lower part of leaf blades of onion plants (cv. Senshu-Chuko). These results suggested that two independent GA biosynthetic pathways, the early-non-hydroxylation pathway to GA₄ (active GA) and early-13-hydroxylation pathway to GA₁ (active GA), exist in onion plants. It was also suggested that GA₄ and GA₁ have almost the same ability to inhibit bulb development in onion plants induced by treatment with an inhibitor of GA biosynthesis, uniconazole-P. The endogenous levels of GA₁ and GA₄, and their direct precursors, GA₂₀ and GA₉, in leaf blades, leaf sheaths, and roots of 4-week-old bulbing and non-bulbing onion plants were measured by gas chromatography/selected ion monitoring with the corresponding [²H]labeled GAs as internal standards. In most cases, the GA levels in long-day (LD)-grown bulbing onion plants were higher than those of short-day (SD)-grown non-bulbing onion plants, but the GA₁ level in leaf blades of SD-grown onion plants was rather higher than that of LD-grown onion plants. Relationship between the endogenous GAs and bulb development in onion plants is discussed.     

Identification of Endogenous Gibberellins in Dormant Bulbils of Chinese Yam, Dioscorea opposita

It is known that dormancy of the genus Dioscorea is induced by application of gibberellin (GA) A₃. To understand the role of GAs in dormancy induction, endogenous GAs have been identified by Kovats retention indices and full mass spectra from capillary gas chromatography-mass spectrometry analysis of purified extract from dormant bulbils of Dioscorea opposita Thunb. These include GA₄, GA₉, GA₁₂, GA₁₉, GA₂₀, GA₂₄, GA₃₆, and GA₅₃; their presence suggests the occurrence of two biosynthetic pathways in D. opposita bulbils, the early 13-hydroxylation pathway and the non-13-hydroxylation pathway.     

The endogenous gibberellins of dwarf mutants of lettuce

The gibberellin (GA) content of E-1, a tall genotype of early flowering lettuce (Lactuca sativa L.), and of three selected GA-responsive dwarfs, dwf1, dwf2, and dwf2¹, has been determined using ¹³C-labeled internal standards and gas chromatographymass spectrometry (GC-MS). In the shoots of the E-1 parent, GA₁, 3-epi-GA₁, GA₃, GA₅, GA₈, GA₁₉, GA₂₀, GA₂₉, and GA₅₃ were identified by full scan GC-MS and Kovats retention indices. Purification by immunoaffinity chromatography selective for 13-hydroxy GAs, was necessary for GA identification. Relative to the parent E-1, the concentrations of GA₁, GA₈, GA₂₀, and GA₂₉ in the shoots of dwf2 plants were reduced to about 10% and in shoots of dwf2¹ plants to less than 50%. In dwf1 the levels of GA₁, GA₈, and GA₂₉ were also reduced to less than 50% of the parent E-1, but the level of GA₂₀ was fivefold higher than in E-1. Plant height was correlated with the endogenous levels of GA₁ and GA₈.     

Quantification of GA1, GA3, GA4, GA7, GA8, GA9, GA19 and GA20; and GA20 metabolism in dormant and non-dormant beechnuts

The endogenous levels of GA₁, GA₃, GA₄, GA₇, GA₈, GA₉, GA₁₉ and GA₂₀ were determined in beech seeds (Fagus sylvatica L.) treated with different dormancy breaking treatments. Gibberellins were analysed separately in cotyledons and embryo axes. After purification of the extracts, GAs were quantified by GC-MS-selected ion monitoring (GC-MS-SIM) with deuterated GAs as internal standards. The results showed that GAs corresponding to the 13-OH pathway seemed to be involved in dormancy breaking. Strong differences in GA₁, GA₃, GA₈, GA₁₉ and GA₂₀ levels between embryo axes and cotyledons of dormant and non-dormant beechnuts were detected with less pronounced differences for GA₄, GA₇ and GA₉ levels. Both the quantitative differences between dormant and non-dormant seeds in the analysed GAs corresponding to the 13-OH pathway, and the capacity of non-dormant seeds to carry out metabolic conversions when labelled GA₂₀ was injected into the seeds, reveal a dynamic role of GAs in dormancy release.     

Light effects on endogenous levels of gibberellins in photoblastic lettuce seeds

Gibberellin A₁ (GA₁), 3-epi-GA₁ GA₁₇, GA₁₉, GA₂₀, and GA₇₇ were identified by Kovats retention indices and full-scan mass spectra from gas chromatography-mass spectrometry analysis of a purified extract of mature seeds of photoblastic lettuce (Lactuca sativa L. cv. Grand Rapids). Non-13-hydroxylated GAs such as GA₄ and GA₉ were not detected even by highly sensitive radioimmunoassay. These results show that the major biosynthetic pathway of GAs in lettuce seeds is the early-13-hydroxylation pathway leading to GA₁, which is suggested to be physiologically active in lettuce seed germination. Quantification of endogenous GAs in the lettuce seeds by gas chromatography-selected ion monitoring using deuterated GAs as internal standards indicated that the endogenous level of GA₁ increased to a level about three times that of dark control 6 h after a brief red light irradiation, and that far-red light given after red light suppressed the effect of red light. The contents of GA₂₀ and GA₁₉ were not affected by the red light irradiation. Evidence is also presented that 3-epi-GA₁ is a native GA in the lettuce seeds.     

Quantification of endogenous gibberellins in leaves and tubers of Chinese yam, Dioscorea opposita Thunb. cv. Tsukune during tuber enlargement

Elevengibberellins (GAs) were identified and quantified in extracts of leaves andtubers of the Chinese yam, Dioscorea opposita Thunb. cv.Tsukune by GC-MS-SIM and Kovats retention indices. Five of these gibberellinsare members of the early-13-hydroxylation pathway (GA₅₃,GA₄₄, GA₁₉, GA₂₀ and GA₁), and sixare members of the non-13-hydroxylation pathway (GA₁₂,GA₁₅, GA₂₄, GA₉, GA₃₆ andGA₄). Of these eleven, GA₄₄, GA₁₅ andGA₁ were detected for the first time in Dioscoreaopposita leaf tissues. The major biosynthetic GA pathway in leavesofChinese yam was non C-13 hydroxylation (NCH). In addition, the activeGA₄ content for all harvest dates was greater than that ofGA₁ in the leaves and tubers during tuber development. It issuggested that the higher level of GA₄ in the leaves and tubers maybe closely related to tuber enlargement.     

Genetic Regulation of Development in Sorghum bicolor: V. The ma(3) Allele Results in Gibberellin Enrichment

Sorghum bicolor genotypes, near isogenic with different alleles at the third maturity locus, were compared for development, for responsiveness to GA₃ and a GA synthesis inhibitor, and occurrence and concentrations of endogenous GAs, IAA, and ABA. At 14 days the genotype 58M (ma₃^Rma₃^R) exhibited 2.5-fold greater culm height, 1.75-fold greater total height, and 1.38-fold greater dry weight than 90M (ma₃ma₃) or 100M (Ma₃Ma₃). All three genotypes exhibited similar shoot elongation in response to GA₃, and 58M showed GA₃-mediated hastening of floral initiation when harvested at day 18 or 21. Both 90M and 100M had exhibited hastening of floral initiation by GA₃ previously, at later application dates. Tetcyclacis reduced height, promoted tillering, and delayed flowering of 58M resulting in plants which were near phenocopies of 90M and 100M. Based on bioassay activity, HPLC retention times, cochromatography with ²H₂-labeled standards on capillary column GC and matching mass spectrometer fragmentation patterns (ions [m/z] and relative abundances), GA₁, GA₁₉, GA₂₀, GA₅₃, and GA₃ were identified in extracts of all three genotypes. In addition, based on published Kovats retention index values and correspondence in ion masses and relative abundances, GA₄₄ and GA₁₇ were detected. Quantitation was based on recovery of coinjected, ²H₂-labeled standards. In 14 day-old-plants, total GA-like bioactivity and GA₁ concentrations (nanograms GA/gram dry weight) were two- to six-fold higher in 58M than 90M and 100M in leaf blades, apex samples, and whole plants while concentrations in culms were similar. Similar trends occurred if data were expressed on a per plant basis. GA₁ concentrations for whole plants were about two-fold higher in 58M than 90M and 100M from day 7 to day 14. Concentrations of ABA and IAA did not vary between the genotypes. The results indicate the mutant allele ma₃^R causes a two- to six-fold increase in GA₁ concentrations, does not result in a GA-receptor or transduction mutation and is associated with phenotypic characteristics that can be enhanced by GA₃ and reduced by GA synthesis inhibitor. These observations support the hypothesis that the allele ma₃^R causes an overproduction of GAs which results in altered leaf morphology, reduced tillering, earlier flowering, and other phenotypic differences between 58M and 90M or 100M.     

Quantification of GA1, GA3, GA4, GA7, GA9, and GA20 in vegetative and male cone buds from juvenile and mature trees of Pinus radiata

The gibberellins GA₁, GA₃, GA₄, GA₇, GA₉ and GA₂₀ were quantified in vegetative and pollen cone buds of juvenile and mature trees of Pinus radiata by combined gas chromatography-mass spectrometry and selected ion monitoring (GC-MS-SIM) using deuterated GAs as internal standards. Higher levels of GA₇ and GA₉ and lower levels of GA₄ were detected in juvenile vegetative buds compared to mature buds, and there were no differences in relation to age for GA₁, GA₃ and GA₂₀. Conversely, when differences between vegetative and pollen cone buds from a mature tree were studied, the highest levels of GA₁ and GA₄ were found in pollen cone buds, similar levels of GA₃, GA₇ and GA₉ were observed in both, and ten fold lower levels of GA₂₀ were found in pollen cone buds as compared with vegetative buds. These results indicate a difference in GA metabolism in relation to both the tree age as well as the physiological status of buds: vegetative or reproductive in this conifer.     

Distribution of endogenous gibberellins in vegetative and reproductive organs of Brassica

Recognizing the physiological diversity of different plant organs, studies were conducted to investigate the distribution of endogenous gibberellins (GAs) in Brassica (canola or oilseed rape). GA₁ and its biosynthetic precursors, GA₂₀ and GA₁₉, were extracted, chromatographically purified, and quantified by gas-chromatography-selected ion monitoring (GC-SIM), using [²H₂]GAs as internal standards. In young (vegetative) B. napus cv. Westar plants, GA concentrations were lowest in the roots, increased acropetally along the shoot axis, and were highest in the shoot tips. GA concentrations were high but variable in leaves. GA₁ concentrations also increased acropetally along the plant axis in reproductive plants. During early silique filling, GA₁ concentrations were highest in siliques and progressively lower in flowers, inflorescence stalks (peduncles plus pedicels), stem, leaves, and roots. Concentrations of GA₁₉ and GA₂₀ showed similar patterns of distribution except in leaves, in which concentrations were higher, but variable. Immature siliques were qualitatively rich in endogenous GAs and GA₁, GA₃, GA₄, GA₈, GA₉, GA₁₇, GA₁₉, GA₂₀, GA₂₄, GA₂₉, GA₃₄, GA₅₁, and GA₅₃ were identified by GC-SIM. In whole siliques, GA₁₉, GA₂₀, GA₁, and GA₈ concentrations declined during maturation due to declining levels in the maturing seeds; their concentrations in the silique coats remained relatively constant and low. These studies demonstrate that GAs are differentially distributed in _Brassica with a general pattern of acropetally increasing concentration in shoots and high concentration in actively growing and developing organs.     

Accumulation of C19-gibberellins in the gibberellin-insensitive dwarf mutantgai ofArabidopsis thaliana (L.) Heynh

The endogenous gibberellins (GAs) from shoots of the GA-insensitive mutant,gai, ofArabidopsis thaliana were analyzed and compared with the GAs from the Landsberg erecta (Ler) line. Twenty GAs were identified in Ler plants by full-scan gas chromatography-mass spectrometry (GC-MS) and Kovats retention indices (KRI's). These GAs are members of the early-13-hydroxylation pathway (GA₅₃, GA₄₄, GA₁₉, GA₁₇, GA₂₀, GA₁, GA₂₉, and GA₈), the non-3,13-hydroxylation pathway (GA₁₂, GA₁₅, GA₂₄, GA₂₅, GA₉, and GA₅₁), and the early-3-hydroxylation pathway (GA₃₇, GA₂₇, GA₃₆, GA₁₃, GA₄, and GA₃₄). The same GAs, except GA₅₃, GA₄₄, GA₃₇, and GA₂₉ were detected in thegai mutant by the same methods. In addition, extracts fromgai plants contained GA₄₁ and GA₇₁. Both lines also contained several unknown GAs. In Ler plants these were mainly hydroxy-GA₁₂ derivatives, whereas in thegai mutant hydroxy-GA₂₄, hydroxy-GA₂₅, and hydroxy-GA₉ compounds were detected. Quantification of seven GAs by GC-selected ion monitoring (SIM), using internal standards, and comparisons of the ion intensities in the SIM chromatograms of the other thirteen GAs, demonstrated that thegai mutant had reduced levels of all C₂₀-dicarboxylic acids (GA₅₃, GA₄₄, GA₁₉, GA₁₂, GA₁₅, GA₂₄, GA₃₇, GA₂₇, and GA₃₆). In contrast,gai plants had increased levels of C₂₀-tricarboxylic acid GAs (GA₁₇, GA₂₅, and GA₄₁) and of all C₁₉-GAs (GA₂₀, GA₁, GA₈, GA₉, GA₅₁, GA₄, GA₃₄, and GA₇₁) except GA₂₉. The 3β-hydroxylated GAs, GA₁ and GA₄, and their respective 2β-hydroxylated derivatives, GA₈ and GA₃₄, were the most abundant GAs found in shoots of thegai mutant. Thus, thegai mutation inArabidopsis results in a phenotype that resembles GA-deficient mutants, is insensitive to both applied and endogenous GAs, and contains low levels of C₂₀-dicarboxylic acid GAs and high levels of C₁₉-GAs. This indicates that theGAI gene controls a step beyond the synthesis of an active GA. Thegai mutant is presumably a GA-receptor mutant or a mutant with a block in the transduction pathway between the receptor and stem elongation. We thank Dr. L.N. Mander, Australian National University, Canberra, for providing [²H]gibberellins, Dr. B.O. Phinney, University of California, Los Angeles, USA for [¹³C]GA₈, and Dr. D.A. Gage, MSU-NIH Mass Spectrometry Facility (grant No. DRR00480), for advice with mass spectrometry. This work was supported by a fellowship from the Spanish Ministry of Agriculture (I.N.I.A.) to M.T., by the U.S. Department of Energy under Contract DE-ACO2-76ERO-1338, and by U.S. Department of Agriculture grant No. 88-37261-3434 to J.A.D.Z.     

Endogenous Gibberellins in Aralia cordata

Eight gibberellins (GAs) were identified in extracts of buds of Aralia cordata by full scan GC/MS and by Kovats retention indices. These GAs comprised five GAs on the early-13-hydroxylation pathway [GA₁, GA₁₉, GA₂₀, GA₄₄, and GA₅₃] and three other GAs [GA₄, GA₁₅, and GA₃₇]. The major GAs were GA₁₉ and GA₄₄.     

Gibberellins and Stem Growth as Related to Photoperiod in Silene armeria L

Stem growth and flowering in the long-day plant Silene armeria L. are induced by exposure to a minimum of 3 to 6 long days (LD). Stem growth continues in subsequent short days (SD), albeit at a reduced rate. The growth retardant tetcyclacis inhibited stem elongation induced by LD, but had no effect on flowering. This indicates that photoperiodic control of stem growth in Silene is mediated by gibberellins (GA). The objective of this study was to analyze the effects of photoperiod on the levels and distribution of endogenous GAs in Silene and to determine the nature of the photoperiodic after-effect on stem growth in this plant. The GAs identified in extracts from Silene by full-scan combined gas chromatography-mass spectrometry (GC-MS), GA₁₂, GA₅₃, GA₄₄, GA₁₇, GA₁₉, GA₂₀, GA₁, GA₂₉, and GA₈, are members of the early 13-hydroxylation pathway. All of these GAs were present in plants under SD as well as under LD conditions. The GA₅₃ level was highest in plants in SD, and decreased in plants transferred to LD conditions. By contrast, GA₁₉, GA₂₀, and GA₁ initially increased in plants transferred to LD, and then declined. Likewise, when Silene plants were returned from LD to SD, there was an increase in GA₅₃, and a decrease in GA₁₉, GA₂₀, and GA₁ which ultimately reached levels similar to those found in plants kept in SD. Thus, measurements of GA levels in whole shoots of Silene as well as in individual parts of the plant suggest that the photoperiod modulates GA metabolism mainly through the rate of conversion of GA₅₃. As a result of LD induction, GA₁ accumulates at its highest level in shoot tips which, in turn, results in stem elongation. In addition, LD also appear to increase the sensitivity of the tissue to GA, and this effect is presumably responsible for the photoperiodic after-effect on stem elongation in Silene.     

Assessing Gibberellins Oxidase Activity by Anion Exchange/Hydrophobic Polymer Monolithic Capillary Liquid Chromatography-Mass Spectrometry

Bioactive gibberellins (GAs) play a key regulatory role in plant growth and development. In the biosynthesis of GAs, GA3-oxidase catalyzes the final step to produce bioactive GAs. Thus, the evaluation of GA3-oxidase activity is critical for elucidating the regulation mechanism of plant growth controlled by GAs. However, assessing catalytic activity of endogenous GA3-oxidase remains challenging. In the current study, we developed a capillary liquid chromatography – mass spectrometry (cLC-MS) method for the sensitive assay of in-vitro recombinant or endogenous GA3-oxidase by analyzing the catalytic substrates and products of GA3-oxidase (GA₁, GA₄, GA₉, GA₂₀). An anion exchange/hydrophobic poly([2-(methacryloyloxy)ethyl]trimethylammonium-co-divinylbenzene-co-ethylene glycol dimethacrylate)(META-co-DVB-co-EDMA) monolithic column was successfully prepared for the separation of all target GAs. The limits of detection (LODs, Signal/Noise = 3) of GAs were in the range of 0.62–0.90 fmol. We determined the kinetic parameters (K _m) of recombinant GA3-oxidase in Escherichia coli (E. coli) cell lysates, which is consistent with previous reports. Furthermore, by using isotope labeled substrates, we successfully evaluated the activity of endogenous GA3-oxidase that converts GA₉ to GA₄ in four types of plant samples, which is, to the best of our knowledge, the first report for the quantification of the activity of endogenous GA3-oxidase in plant. Taken together, the method developed here provides a good solution for the evaluation of endogenous GA3-oxidase activity in plant, which may promote the in-depth study of the growth regulation mechanism governed by GAs in plant physiology.     

Gibberellins and the Legume-Rhizobium Symbiosis : III. Quantification of Gibberellins from Stems and Nodules of Lima Bean and Cowpea

Lima bean (Phaseolus lunatus L.) plants inoculated with Bradyrhizobium sp. strain 127E14 displayed a period of marked internode elongation that was not observed in plants inoculated with other compatible bradyrhizobia, including strain 127E15. When strain 127E14 nodulated an alternate host, cowpea (Vigna unguiculata L. Walp), a similar, although less dramatic growth response induced by the bacteria was observed. It has been speculated that the elongative growth promotion brought about by inoculation with strain 127E14 is mediated by gibberellins (GAs). Using deuterated internal standards and gas chromatography-mass spectroscopy analysis, we have quantified the levels of GA₁, GA₂₀, GA₁₉, and GA₄₄ in nodules and stems of two varieties of lima bean (bush and pole) and one variety of cowpea that were inoculated with either strain 127E14 or 127E15. In nodules formed by strain 127E14 on lima bean, endogenous levels of GA₂₀ and GA₁₉ were 10 to 40 times higher (35-88 ng/g dry weight) than amounts found in nodules formed by strain 127E15 (2.2-3.9 ng/g dry weight). Relative amounts of GA₄₄ were also higher (4- to 11-fold) in 127E14 nodules, but this increase was less pronounced. The rhizobial-induced increase of these GAs in the nodule occurred in both pole and bush varieties and seemed to be independent of host morphology. Regardless of rhizobial inoculum, levels of the “bioactive” GA₁ in the nodule (0.3-1.1 ng/g dry weight) were similar. In cowpea nodules, a similar, although smaller, difference in GA content due to rhizobial strain was observed. The concentration of GA₁ in lima bean stems was generally higher than that observed in the nodule, whereas concentrations of the other GAs measured were lower. In contrast with the nodule, GA concentrations in lima bean stems were not greater in plants inoculated with strain 127E14, and in some cases the slower growing plants inoculated with strain 127E15 actually had higher levels of GA₂₀, GA₁₉, and GA₄₄. Thus, there were major differences in concentrations of the precursors to GA₁ in nodules formed by the two bacterial strains, which were positively correlated with the observed elongation growth. These results support the hypothesis that the rhizobial strain modifies the endogenous GA status of the symbiotic system. This alteration in GA balance within the plant, presumably, underlies the observed growth response.     

Identification of gibberellins in leaf tissues of day-neutral strawberry (Fragaria × ananassa Duch.) cultivars

The endogenous gibberellins (GAs) in leaf tissues of two day-neutral cultivars (Rapella and Selva) of strawberry (Fragaria × ananassa Duch.) were analysed using combined gas chromatography -- mass spectrometry (GC-MS). Seven of the later members of the 13-hydroxylation GA biosynthetic pathway were identified, by comparison of Kovats retention indices and mass spectral data obtained for methyl ester trimethylsilyl ether derivatives, either with data obtained from authentic compounds or literature values. GA₁, GA₃, GA₈, GA₁₇, GA₁₉, GA₂₀ and GA₂₉ were detected in extracts of both cultivars.     

Internode length in pisum. Mutants lk,lka, and lkb do not accumulate gibberellins

The levels of gibberellin A₁ (GA₁), GA₈, GA₁₉, GA₂₀, GA₂₉, and GA₄₄ in the short Pisum sativum L. mutants lk, lka, and lkb, and comparable wild-type plants, were determined by gas chromatography-selected ion monitoring (GC-SIM) using ²H or ¹³C internal standards. The mutants possessed similar GA₁ levels to wild-type plants, consistent with their classification as GA-sensitivity rather than GA-synthesis mutants. However, these mutants differ from certain sensitivity mutants in other species, in which substantial accumulation of GA₁ occurs. The results suggest that if the proposed feedback model for the regulation of GA synthesis occurs in peas it is not the reduced growth per se that is the trigger for elevated levels of C₁₉ GAs. The results are also consistent with the hypothesis that in those GA-sensitivity mutants which do not accumulate C₁₉ GAs, the biochemical lesion may be well down the transduction pathway which leads from GA₁ reception to stem elongation.     

Identification of endogenous gibberellins from sorghum

Gibberellins (GA) A₁, A₁₉, and A₂₀ were identified in shoot cylinders containing the apical meristems from sorghum (Sorghum bicolor L.). Extracts were purified by sequential SiO₂ partition chromatography and reversed-phase C₁₈ high performance liquid chromatography and biologically active (dwarf rice cv Tan-ginbozu microdrop assay) fractions were subjected to gas chromatography-selected ion monitoring. Based on the use of [³H]GA and [²H](d₂)GA internal standards, amounts of GA₁, GA₁₉, and GA₂₀ were estimated to be 0.7, 8.8, and 1.5 namograms per gram dry weight of tissue, respectively.     

Identification of endogenous gibberellins in navel orange shoots

Eight gibberellins (GAs) were identified from vegetative shoots of navel orange trees (Citrus sinensis L. Osbeck cv Washington) after sequential purification by reverse-phase C₁₈ high performance liquid chromatography, Nucleosil 5N(CH₃)₂ high performance liquid chromatography, and capillary gas chromatography-mass spectrometry. GA₁, GA₁₇, GA₁₉, GA₂₀, GA₂₉, and iso-GA₃ were identified based on the full scan mass spectra and Kovats retention indices. GA₈ was tentatively identified based on the comparison of the full scan mass spectra with the published spectra. GA₄₄ was tentatively identified from the characteristic masses at the correct Kovats retention index.     

Comparison of Biological Activities of Gibberellins and Gibberellin-Precursors Native to Thlaspi arvense L

Field pennycress (Thlaspi arvense L.) is a winter annual weed with a cold requirement for stem elongation and flowering. The relative abilities of several native gibberellins (GAs) and GA-precursors to elicit stem growth were compared. Of the eight compounds tested, gibberellin A₁, (GA₁), GA₉, and GA₂₀ caused stem growth in noninduced (no cold treatment) plants. No stem growth was observed in plants treated with ent-kaurene, ent-kaurenol, ent-kaurenoic acid, GA₅₃, or GA₈. Moreover, of the biologically active compounds, GA₉ was the most active followed closely by GA₁. In thermoinduced plants (4-week cold treatment at 6°C) that were continuously treated with 2-chlorocholine chloride to reduce endogenous GA production, GA₉ was the most biologically active compound. However, the three kaurenoid GA precursors also promoted stem growth in thermoinduced plants, and were almost as active as GA₂₀. No such increase in activity was observed for either GA_[unk] or GA₅₃. The results are discussed in relation to thermoinductive regulation of GA metabolism and its significance to the initiation of stem growth in field pennycress. It is proposed that thermoinduction results in increased conversion of ent-kaurenoic acid to GAs through the C-13 desoxy pathway and that GA₉ is the endogenous mediator of thermoinduced stem growth in field pennycress.