Identification of 3-Hydroxy-2-indolone-3-acetylaspartic Acid as a New Indole-3-acetic Acid Metabolite in Vicia Roots

A new indole-3-acetic acid (IAA) metabolite in the root of Viciafaba L. cv. Chukyo was identified as 3-hydroxy-2-indolone-3-acetylasparticacid, with the simpler name of dioxindole-3-acetylaspartic acid,by comparison with the authentic sample. Formation of dioxindole-3-aceticacid conjugates seems to be a major route of IAA metabolismin Vicia roots. (Received October 22, 1985; Accepted January 7, 1986)     

The biosynthesis and conjugation of indole-3-acetic acid in germinating seed and seedlings ofDalbergia dolichopetala

Germinating seed ofDalbergia dolichopetala converted both [²H₅]l-tryptophan and [²H₅]indole-3-ethanol to [²H₅]indole-3-acetic acid (IAA). Metabolism of [2-¹⁴C]IAA resulted in the production of indole-3-acetylaspartic acid (IAAsp), as well as several unidentified components, referred to as metabolites I, II, IV and V. Re-application of [¹⁴C]IAAsp to the germinating seed led to the accumulation of the polar, water-soluble compound, metabolite V, as the major metabolite, together with a small amount of IAA. Metabolites I, II and IV were not detected, nor were these compounds associated with the metabolism of [2-¹⁴C]IAA by shoots and excised cotyledons and roots from 26-d-oldD. dolichopetala seedlings. Both shoots and cotyledons converted IAA to IAAsp and metabolite V, while IAAsp was the only metabolite detected in extracts from excised roots. The available evidence indicates that inDalbergia, and other species, IAAsp may not act as a storage product that can be hydrolysed to provide the plant with a ready supply of IAA.Abbreviations HPLC-RC high-performance liquid chromatography-radiocounting - IAA indole-3-acetic acid - IAAsp indole-3-acetylaspartic acid - IAlnos 2-O-indole-3-acetyl-myo-inositol - IEt indole-3-ethanol     

Ethylene-enhanced catabolism of [C]indole-3-acetic Acid to indole-3-carboxylic Acid in citrus leaf tissues

Exogenous [¹⁴C]indole-3-acetic acid (IAA) is conjugated in citrus (Citrus sinensis) leaf tissues to one major substance which has been identified as indole-3-acetylaspartic acid (IAAsp). Ethylene pretreatment enhanced the catabolism of [¹⁴C]IAA to indole-3-carboxylic acid (ICA), which accumulated as glucose esters (ICGIu). Increased formation of ICGIu by ethylene was accompanied by a concomitant decrease in IAAsp formation. IAAsp and ICGIu were identified by combined gas chromatography-mass spectrometry. Formation of ICGIu was dependent on the concentration of ethylene and the duration of the ethylene pretreatment. It is suggested that the catabolism of IAA to ICA may be one of the mechanisms by which ethylene reduces endogenous IAA levels.     

Metabolism of [14C]Indole-3-Acetic Acid by Coleoptiles of Zea mays L.

Reverse-phase high-performance liquid chromatography was usedto analyse [¹⁴C]-labelled metabolites of indole-3-acetic acid(IAA) in coleoptile segments of Zeo mays seedlings. After incubationfor 2 h in 10^–2 mol m^–3 [2-¹⁴C]IAA, methanolic extractsof coleoptiles contained between six and ten radioactive compounds,one of which co-chromatographed with IAA. The metabolic productsin coleoptile extracts appeared to be similar to those in rootextracts, with an oxindole-3-acetic-acid-like component as theprincipal metabolite, but the rate of metabolism was slowerin coleoptile than in root segments. Decarboxylation did notappear to play a major role in the metabolism of exogenous IAAduring the short incubation periods. Moreover, external IAAconcentration had little effect on the pattern of metabolism.Coleoptile segments were also supplied with [¹⁴C]IAA from agardonor blocks placed at the apical ends, and agar receiver blockswere placed at the basal ends. After incubation for 4 h, theidentity of the single radioactive compound in the receiverblocks was shown to be IAA by both reverse-phase high-performanceliquid chromatography and gas chromatography-mass spectrometrytechniques. Key words: Zea mays, Coleoptile, High-performance liquid chromatography, Indole-3-acetic acid     

Metabolism of Auxin in Tomato Fruit Tissue: Formation of High Molecular Weight Conjugates of Oxindole-3-Acetic Acid via the Oxidation of Indole-3-Acetylaspartic Acid

High performance liquid chromatography of extracts of tomato (Lycopersicon esculentum Mill.) incubated with a relatively low concentration (4 μm) of [1-¹⁴C]indole-3-acetic acid (IAA) revealed the presence of two major polar metabolites. Hydrolysis of the two metabolites with 7 n NaOH yielded the same compound, which had a retention time similar to that of ring-expanded oxindole-3-acetic acid (OxIAA) on high performance liquid chromatography. The identity of the indolic moiety of these conjugates as OxIAA was further confirmed by gas chromatography-mass spectrometry. Chromatography of the two OxIAA conjugates on a calibrated Bio-Gel P-2 column indicated that their molecular weights are about 1200 and 1000. Aspartic acid and glutamic acid were the major amino acids detected in acid hydrolysates of the two conjugates. Increasing the concentration of IAA in the incubation medium resulted in an increase in the formation of indole-3-acetylaspartic acid (IAAsp) with a concomitant decrease in the formation of the two OxIAA conjugates. Feeding experiments with labeled IAAsp and OxIAA showed that IAAsp and not OxIAA is the precursor of these conjugates. The data obtained indicate that exogenous IAA is converted in tomato pericarp tissue to high molecular weight conjugates, presumably peptides, of OxIAA via the oxidation of IAAsp. The oxidation of IAAsp seems to be a rate-limiting step in the formation of these conjugates from exogenous IAA.     

Metabolism of Indole-3-Acetic Acid by Pericarp Discs from Immature and Mature Tomato (Lycopersicon esculentum Mill)

[1′-¹⁴C, ¹³C₆]Indole-3-acetic acid was infiltrated into immature pericarp discs from fruits of tomato (Lycopersicon esculentum Mill., cv Moneymaker). After a 24-h incubation period the discs were extracted with methanol and the partially purified extract was analyzed by reversed-phase high-performance liquid chromatography-radiocounting. Five metabolite peaks (1-5) were detected and subsequently analyzed by combined high-performance liquid chromatography-frit-fast atom bombardment-mass spectrometry. The metabolite 4 fraction was found to contain [¹³C₆]-indole-3-acetylaspartic acid, and analysis of metabolite 5 identified [¹³C₆]indole-3-acetyl-β-d-glucose. The other metabolites could not be identified, but alkaline hydrolysis studies and gel permeation chromatography indicated that metabolites 1 and 3 were both amide conjugates with a molecular weight of approximately 600. Studies with radiolabeled indole-3-acetic acid, indole-3-acetylaspartic acid, and indole-3-acetyl-β-d-glucose demonstrated that in immature pericarp indole-3-acetic acid is deactivated primarily via metabolism to indole-3-acetylaspartic acid, which is further converted to metabolites 1, 2, and 3. In mature, pink pericarp discs, indole-3-acetic acid is converted more extensively to its glucosyl conjugate. Conjugation of indole-3-acetic acid to indole-3-acetylaspartic acid appears to be dependent upon protein synthesis because it is inhibited by cycloheximide. In contrast, cycloheximide has little effect on the further conversion of indole-3-acetylaspartic acid to metabolites 1, 2, and 3.     

Transport of shoot- and cotyledon-applied indole-3-acetic acid to Vicia faba root

To clarify the participation of indole-3-acetic acid (IAA) originatingfrom the shoot in root growth regulation and the mechanism ofIAA translocation from shoot to root, the movement of ¹⁴C-IAAwhich was applied to the epicotyl or the cotyledon of Viciafaba seedlings was investigated. The radioactivity of IAA appliedto the cotyledon moved faster to the root tip than that appliedto the epicotyl. On the basis of the effect of 2,3,5-triiodobenzoic acid on IAAmovement, a comparison with ¹⁴C-glucose movement and autoradiographicexamination, the nature of IAA movement was concluded to bepolar transport from the epicotyl to the basal part of the roots,while IAA movement from the epicotyl to the cotyledon, fromthe basal part of roots to the apical part, and from the cotyledonto the epicotyl and to the root took place in the phloem. Theradioactivity from ¹⁴C-IAA applied to the cotyledon accumulatedin lateral root primordia and vascular bundles. These factssuggest that IAA produced in cotyledons may participate in theregulation of Vicia root development. (Received December 21, 1979; )     

Measurement of the Rates of Oxindole-3-Acetic Acid Turnover, and Indole-3-Acetic Acid Oxidation in Zea mays Seedlings

Nonhcbcl, H. M. 1986. Measurement of the rates of oxindole-3-aceticacid turnover and indole-3-acetic acid oxidation in Zea maysseedlings.—J. exp. Bat. 37: 1691–1697. Oxindole-3-acetic acid is the pnncipal catabolite of indole-3-aceticacid in Zea mays seedlings. In this paper measurements of theturnover of oxindole-3-acetic acid are presented and used tocalculate the rate of indole-3-acetic acid oxidation. [³H]Oxindolc-3-acetic acid was applied to the endosperm of Zeamays seedlings and allowed to equilibrate for 24 h before thestart of the experiment. The subsequent decrease in its specificactivity was used to calculate the turnover rate. The averagehalf-life of oxindole-3-acetic acid in the shoots was foundto be 30 h while that in the kernels had an average half-lifeof 35 h. Using previously published values of the pool sizesof oxindole-3-acetic acid in shoots and kernels from seedlingsof the same age and variety, and grown under the same conditions,the rate of indole-3-acetic acid oxidation was calculated tobe I-I pmol plant^–1 h^–1 in the shoots and 7·1pmol plant^–1 h^–1 in the kernels. Key words: Oxindole-3-acetic acid, indole-3-acetic acid, turnover, Zea mays     

Analysis of Indole-3-Acetic Acid Metabolites from Dalbergia dolichopetala by High Performance Liquid Chromatography-Mass Spectrometry

A mixture of [2-¹⁴C₁] and [¹³C₆]indole-3-acetic acid was applied to the cotyledons of 6-day-germinated seeds of “jacarandá do cerrado” (Dalbergia dolichopetala) and after 8 hours the seeds were extracted. Analysis of the fractionated extract by reversed-phase high performance liquid chromatography-radiocounting revealed the presence of five radiolabeled metabolite peaks (I-V). After further purification, the individual peaks of radioactivity were analyzed by combined high performance liquid chromatography-steel filter-fast atom bombardment-mass spectrometry. The metabolite fraction V was found to contain [¹⁴C₁, ¹³C₆]indole-3-acetylas-partic acid and unlabeled indole-3-acetylglutamic acid. Analysis of the metabolite fraction II revealed the presence of dioxindole-3-acetylaspartic acid and putative dioxindole-3-acetylglutamic acid as well as putative benzene ring-hydroxylated derivatives of oxindole-3-acetylaspartic acid and oxindole-3-acetylglutamic acid. There was no evidence of significant incorporation of label from [2′-¹⁴C₁] or [¹³C₆]indole-3-acetic acid into any of these conjugated indoles.     

Indole-3-acetic acid homeostasis in transgenic tobacco plants expressing the Agrobacterium rhizogenes rolB gene

The rolB gene of the plant pathogen Agrobacterium rhizogenes has an important role in the establishment of hairy root disease in infected plant tissues. When expressed as a single gene in transgenic plants the RolB protein gives rise to effects indicative of increased auxin activity. It has been reported that the RolB product is a β-glucosidase and proposed that the physiological and developmental alterations in transgenic plants expressing the rolB gene are the result of this enzyme hydrolysing bound auxins, in particular (indole-3-acetyl)-β-D-glucoside (IAGluc), and thereby bringing about an increase in the intracellular concentration of indole-3-acetic acid (IAA). Using tobacco plants as a test system, this proposal has been investigated in detail. Comparisons have been made between the RolB phenotype and that of IaaM/iaaH transformed plants overproducing IAA. In addition, the levels of IAA and IAA amide and IAA ester conjugates were determined in wild-type and transgenic 35S-rolB tobacco plants and metabolic studies were carried out with [¹³C₆]IAA [2′-¹⁴C]IAA, [¹⁴C]IAGluc, [5-³H]-2-o-(indole-3-acetyl)-myo-inositol and [¹⁴C]indole-3-acetylaspartic acid. The data obtained demonstrate that expression of the rolB encoded protein in transgenic tobacco does not produce a phenotype that resembles that of IAA over producing plants, does not alter the size of the free IAA pool, has no significant effect on the rate of IAA metabolism, and, by implication, appears not to influence the overall rate of IAA biosynthesis. Furthermore, the in vivo hydrolysis of IAGluc, and that of the other IAA conjugates that were tested, is not affected. On the basis of these findings, it is concluded that the RolB phenotype is not the consequence of an increase in the size of the free IAA pool mediated by an enhanced rate of hydrolysis of IAA conjugates.     

Auxin Concentrations in Nodes and Internodes ofImpatiens sultani

Greater concentrations of auxin at nodes than in internodes,resulting from some nodal barrier to basipetal transport, havelong been postulated as the cause of early differentiation ofinitially isolated xylem and cambium at the nodes. However,this study, using [¹⁴C] indole-3-acetic acid (IAA) applied apicallyand gas chromatography-mass spectrometry, found that in stemsofImpatiens sultanithe IAA concentrations (per unit f. wt) atnodes were similar to those in adjacent internodes, though alittle greater at nodes if expressed per unit length of stemand a little less per unit d. wt. By contrast, in decapitatedshoots and in stem explants of dicotyledons, loss of the apicalsource of basipetally flowing auxin can result in auxin drainagewith some auxin retention in the uppermost remaining nodes.When [¹⁴C]IAA was applied apically to shoots for 4 h and stemexplants were excised, the explants had no nodal accumulationinitially whereas comparable explants incubated for 20 h revealedsignificant nodal accumulation. If decapitation leads both tonodal auxin accumulation and to adventitious abscission justabove the node, this fits the hypothesis that abscission sitesare positioned where auxin concentration decreases locally inthe apical direction. Difficulties in quantifying nodal auxindynamics are discussed, and some crude estimates of metabolicrates and locations of the auxin are presented.Copyright 1999Annals of Botany Company Abscission, auxin,Impatiens sultani, indole-3-acetic acid, node.     

Translocation of Radiolabeled Indole-3-Acetic Acid and Indole-3-Acetyl-myo-Inositol from Kernel to Shoot of Zea mays L.

Either 5-[³H]indole-3-acetic acid (IAA) or 5-[³H]indole-3-acetyl-myo-inositol was applied to the endosperm of kernels of dark-grown Zea mays seedlings. The distribution of total radioactivity, radiolabeled indole-3-acetic acid, and radiolabeled ester conjugated indole-3-acetic acid, in the shoots was then determined. Differences were found in the distribution and chemical form of the radiolabeled indole-3-acetic acid in the shoot depending upon whether 5-[³H]indole-3-acetic acid or 5-[³H]indole-3-acetyl-myo-inositol was applied to the endosperm. We demonstrated that indole-3-acetyl-myo-inositol applied to the endosperm provides both free and ester conjugated indole-3-acetic acid to the mesocotyl and coleoptile. Free indole-3-acetic acid applied to the endosperm supplies some of the indole-3-acetic acid in the mesocotyl but essentially no indole-3-acetic acid to the coleoptile or primary leaves. It is concluded that free IAA from the endosperm is not a source of IAA for the coleoptile. Neither radioactive indole-3-acetyl-myo-inositol nor IAA accumulates in the tip of the coleoptile or the mesocotyl node and thus these studies do not explain how the coleoptile tip controls the amount of IAA in the shoot.     

Conjugation of Indole-3-Acetic Acid (IAA) in Wild-Type and IAA-Overprodcing Transgenic Tobacco Plants,and Identification of the Main Conjugates by Frit-Fast Atom Bombardment Liquid Chromatography-Mass Spectrometry

Transgenic plants overproducing indole-3-acetic acid (IAA) from expression of the Agrobacterium tumefaciens T-DNA IAA biosynthesis genes were used to study the conjugation of IAA. At the 11-node stage, free IAA, as well as ester- and amide-conjugated IAA, was analyzed in wild-type tobacco SR1 and in transgenic plants denoted 35S-iaaM/iaaH (line C) and 35S-iaaM x 35S-iaaH (line X). The transgenic plants contained increased levels of both free and conjugated IAA, and the main increase in IAA conjugates occurred in amide conjugates. Two amide conjugates were identified by fritfast atom bombardment liquid chromatography-mass spectrometry as indole-3-acetylaspartic acid (IAAsp) and indole-3-acetylglutamic acid (IAGlu), and one ester conjugate was identified as indole-3-acetylglucose. IAAsp and IAGlu were also identified as endogenous substances in wild-type plants. In wild-type plants, the percent of total IAA in the free form was significantly higher in young leaves (73 [plus or minus] 7%, SD) than in old leaves (36 [plus or minus] 8%), whereas there was no difference between young (73 [plus or minus] 8%) and old internodes (70 [plus or minus] 9%). In IAA-overproducing transformants, both free and conjugated IAA levels were increased, but the percent free IAA was maintained constant (57 [plus or minus] 10%) for both leaves and internodes, independent of the total IAA level or tissue age. These results suggest that synthesis or transport of IAA conjugates is regulated in the vegetative wild-type plant, and that different organs possess a unique balance between free and conjugated IAA. The IAA-overproducing plant, however, acquires a lower proportion of free IAA in the stem and younger leaves, presumably determined by a higher conjugation in those tissues compared with wild type.     

Distribution of Radioactivity from Exogenously Supplied [1-14C]Indol-3-yl-acetic Acid and [3, 4-3H (N)] Gibberellin A, in Geotropically-stimulated Picea abies (L.) Karst. Roots

The distribution of exogenously-supplied radioactive labelledindol-3-yl-acetic acid (IAA) and gibberellin A₁ (GA₁) in geotropicallystimulated roots of Norway spruce (Picea abies (L.) Karst.)has been demonstrated. Seedlings were positioned with theirroot tips in 2.1 x 10^–6 M [¹⁴C]IAA or 1.3 x 10^–8m ³H-GA₁ for 4 and 20 h, respectively. After geotropic stimulationfor 90 min in the horizontal position the root tips were cutlongitudinally in 50 µm thick sections, using a freeze-microtome.The radioactivity in the ¹⁴C-IAA treated roots occurred in higherconcentration in the lower than in the upper halves (ratio 1.25:1). A similar trend was observed in the [³H]GA₁-treated rootswhere the ratio lower: upper halves was 2.04: 1. The ratio ofradioactivity in right and left halves of vertical roots wasapproximately the same in roots supplied with [¹⁴C]IAA and [³H]GA₁(1.09: 1). The supplied radioactive compounds were analysed chromatographicallyafter extraction in methanol of 6 mm apical root segments. Onlya small fraction (7–8 per cent) of the supplied [¹⁴C]IAAwas revealed unchanged in the segments. The major part of thechromatographed, labelled compound has not been identified,but on basis of its R_F value it is suggested that it may beindol-3-acetyl-aspartic acid (IAAasp). The chromatographic analysis of the [³H]GA,-treated segmentsshowed that only small fractions of this gibberellin has beenconverted to other compounds. These results have been discussed and correlated with knowledgeof plant growth regulators and their participation in root geotropism. Picea abies, spruce, geotropism, gibberellin A₁, indol-3-yl-acetic acid, growth regulators, redistribution in roots     

Auxin-induced ethylene production as related to auxin metabolism in leaf discs of tobacco and sugar beet

Exogenously supplied indole-3-acetic acid (IAA) stimulated ethylene production in tobacco (Nicotiana glauca) leaf discs but not in those of sugar beet (Beta vulgaris L.). The stimulatory effect of IAA in tobacco was relatively small during the first 24 hours of incubation but became greater during the next 24 hours. It was found that leaf discs of these two species metabolized [1-¹⁴C]IAA quite differently. The rate of decarboxylation in sugar beet discs was much higher than in tobacco. The latter contained much less free IAA but a markedly higher level of IAA conjugates. The major conjugate in the sugar beet extracts was indole-3-acetylaspartic acid, whereas tobacco extracts contained mainly three polar IAA conjugates which were not found in the sugar beet extracts. The accumulation of the unidentified conjugates corresponded with the rise of ethylene production in the tobacco leaf discs. Reapplication of all the extracted IAA conjugates resulted in a great stimulation of ethylene production by tobacco leaf discs which was accompanied by decarboxylation of the IAA conjugates. The results suggest that in tobacco IAA-treated leaf discs the IAA conjugates could stimulate ethylene production by a slow release of free IAA. The inability of the exogenously supplied IAA to stimulate ethylene production in the sugar beet leaf discs was not due to a deficiency of free IAA within the tissue but rather to the lack of responsiveness of this tissue to IAA, probably because of an autoinhibitory mechanism existing in the sugar beet leaf discs.     

Mechanism of Indole-3-acetic Acid Conjugation: No Induction by Ethylene

Formation of indole-3-acetic acid-aspartate in detached primary leaves of cowpea (Vigna sinensis Endl.) floating on (14)C-indole-3-acetic acid (3 muc; 3.15 mum, phosphate-citrate buffer, pH 4.75), almost doubled when leaves were pretreated with 31.5 mum(12)C-indole-3-acetic acid for 17 hr and then transferred to (14)C-indole-3-acetic acid for 4 hours as compared with leaves preincubated in buffer only. When leaves were preincubated with ethylene (11.0 and 104 mul/l) instead of (12)C-indole-3-acetic acid, no induction of indole-3-acetylaspartic acid formation was observed, and the rate of indole-3-acetylaspartic acid formation decreased as compared with control leaves. Rhizobitoxine (1.87 mum) inhibited indole-3-acetic acid-induced ethylene production but did not prevent the formation of indole-3-acetylaspartic acid. In view of the similarity of these results and those previously obtained with alpha-naphthaleneacetic acid, it is concluded that ethylene has no role in the auxin-induced indole-3-acetylaspartic acid formation in cowpea leaves.     

Biosynthesis and metabolism of indole-3-ethanol and indole-3-acetic acid by Pinus sylvestris L. needles

Combined gas chromatography-mass spectrometry has been used to identify indole-3-ethanol (IEt) in a purified extract from needles of Pinus sylvestris L. Quantitative estimates obtained by high-performance liquid chromatography with fluorescence detection, corrected for samples losses occurring during purification, indicate that Pinus needles contain 46±4 ng g^-1 IEt. This compares with 24.5±6.5 ng g^-1 indole-3-acetic acid (IAA) and 2.3±0.4 ng g^-1 indole-3-carboxylic acid (ICA) (Sandberg et al. 1984, Phytochemistry, 23, 99–102). Metabolism studies with needles incubated in a culture medium in darkness revealed that both [3-¹⁴C]-tryptophan and [2-¹⁴C]tryptamine mine are converted to [¹⁴C]IEt. It was also shown that [3-¹⁴C]IEt acted as a precursor of [¹⁴C]IAA. The observed metabolism appears to be enzymic in nature. The [2-¹⁴C]IAA was not catabolised to [¹⁴C]ICA in detectable quantities implying that, at best, only a minor portion of the endogenous ICA pool in the Pinus needles originates from IAA.Abbreviations DEAE diethylaminoethyl - GC-MS gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - IAA indole-3-acetic acid - ICA indole-3-carboxylic acid - IEt indole-3-ethanol - PVP polyvinylpyrrolidone     

Analysis of Indole-3-Acetic Acid and Related Indoles in Culture Medium from Azospirillum lipoferum and Azospirillum brasilense

Analysis of neutral and acidic ethyl acetate extracts from culture medium of Azospirillum brasilense 703Ebc by high-performance liquid chromatography (HPLC) and combined gas chromatography-mass spectrometry demonstrated the presence of indole-3-acetic acid (IAA), indole-3-ethanol, indole-3-methanol, and indole-3-lactic acid. IAA in media of 20 strains of A. brasilense and Azospirillum lipoferum was analyzed quantitatively by both the colorimetric Salkowski assay and HPLC-based isotopic dilution procedures. There was little correlation between the estimates obtained with the two procedures. For instance, the Salkowski assay suggested that the culture medium from A. brasilense 703Ebc contained 26.1 μg of IAA ml⁻¹, whereas HPLC revealed the presence of only 0.5 μg of IAA ml⁻¹. Equivalent estimates with A. brasilense 204Ed were 10.5 and 0.01 μg of IAA ml⁻¹, respectively. The data demonstrate that the Salkowski assay is not a reliable method for measuring the IAA content of Azospirillum culture medium and that estimates in excess of 10 μg of IAA ml⁻¹ should be viewed with particular caution. Metabolism of [2′-¹⁴C]IAA by A. brasilense 703Ebc yielded radiolabeled indole-3-methanol, whereas roots of maize (Zea mays L.) seedlings gave rise to [¹⁴C]oxindole-3-acetic acid and an array of polar metabolites. Metabolism of [2′-¹⁴C]IAA by maize roots inoculated with A. brasilense 703Ebc produced a metabolic profile characteristic of maize rather than Azospirillum species.     

Metabolism of [14C]indole-3-acetic acid by the cortical and stelar tissues of Zea mays L. roots

Reverse-phase high-performance liquid chromatography was used to analyse ¹⁴C-labelled metabolites of indole-3-acetic acid (IAA) formed in the cortical and stelar tissues of Zea mays roots. After a 2-h incubation in [¹⁴C]IAA, stelar segments had metabolised between 1–6% of the methanol-extractable radioactivity compared with 91–92% by the cortical segments. The pattern of metabolites produced by cortical segments was similar to that produced by intact segments bathed in aqueous solutions of [¹⁴C]IAA. In contrast, when IAA was supplied in agar blocks to stelar tissue protruding from the basal ends of segments, negligible metabolism was evident. On the basis of its retention characteristics both before and after methylation, the major metabolite of [¹⁴C]IAA in Zea mays root segments was tentatively identified by high-performance liquid chromatography as oxindole-3-acetic acid.Abbreviations HPLC High-performance liquid chromatography - IAA Indole-3-acetic acid     

Changes in the Level of [C]Indole-3-Acetic Acid and [C]Indoleacetylaspartic Acid during Root Formation in Mung Bean Cuttings

Changes in the levels of [¹⁴C]indole-3-acetic acid (IAA) and [¹⁴C]indole-acetylaspartic acid (IAAsp) were examined during adventitious root formation in mung bean (Vigna radiata [L.] R. Wilcz. `Berken') stem cuttings. IAAsp was identified by GC-MS as the primary conjugate in IAA-treated cuttings. During root formation in IAA-treated cuttings, the level of [¹⁴C]IAAsp increased rapidly the first day and then declined; [¹⁴C]IAA was rapidly metabolized and not detected after 12 hours.