Indole Auxins Research Articles

Optimization of a high speed countercurrent chromatograph for analytical separations

AbstractThe analytical capability of the high speed countercurrent chromatograph model 4000 (HSCCC‐4000) has been improved by optimizing the dimensions of the multilayer coiled column. Using a two‐phase solvent system of n‐hexane – ethyl acetate – methanol – water (1:1:1:1) and a set of indole auxins as test samples, a series of studies was conducted to evaluate performance of coiled columns with i.d.s ranging from 0.1 to 0.55 mm. The studies on the stationary phase retention indicated that multilayer coils of 0.55 mm and 0.30 mm i.d. provide satisfactory retention of the stationary phase while the 0.10 mm i.d. column fails to yield reproducible retention. The best partition efficiencies were obtained from the 0.30 mm i.d. multilayer coil with a 6 ml capacity which produced theoretical plate numbers ranging from 5500 to 10500 with a resolution factor of 2.30. The feasibility of interfacing the HSCCC‐4000 with a mass spectrometer is briefly discussed.

Dark and photoinduced interactions between riboflavin and indole auxins

Interactions between riboflavin (RF) and indole auxins in the darkness and under conditions of irradiation with visible light were carried out. Dark complexation takes place in buffered (pH 7) aqueous solution. The association constants determined by static fluorescence quenching range from 70 to 150 mol-1 dm3. The complexation is significant at relatively high indole auxin concentrations (above 10-3 mol dm-3). It is governed by an additive effect of charge transfer, known for flavine-indole systems, and of the hydrophobic bond formation. The latter appears in the case of auxins as the driving force of the interaction. No quenching of singlet-excited RF was detected up to indole auxin concentrations of 5 . 10-3 mol dm-3. Under irradiation with visible light a complex mechanism of competitive reactions is operative: both RF and especially the auxins are decomposed by a combination of type I and II photooxidations at indole auxins concentration of 10-4 mol dm-3. The rate constants of the order of 109 mol-1 dm3 s-1 were estimated for the quenching of triplet excited RF by the indoles. For comparative purposes Rose Bengal sensitized photooxidation of a series of indolecarboxylic acids of indole-3-acetic, -propionic and –butyric acid was examined. The rate constants of the photooxidative process, mediated by singelt molecular oxygen, were of the order of 107 mol-1 dm3 s-1 for the auxins. As a result of the interaction, RF is protected against photobleaching and the auxins decompose via a different mechanism, depending on the relative flavin-auxin concentrations.

Systematic search for suitable two-phase solvent systems for high-speed counter-current chromatography

We have introduced two series of two-phase solvent systems which facilitate the systematic search for the solvent systems suitable for high-speed counter-current chromatography. The n-hexane—ethyl acetate— n-butanol—methanol—water systems provide a broad range of hydrophobicity, while the chloroform—methanol—water systems are extremely useful for separations of various natural products with moderate hydrophobicity. The practical use of these solvent series was demonstrated with several test samples which include dinitrophenyl amino acids, S-triazine herbicides, indole auxin, and non-ionic organic solvents.

Studies on Chlorinated Indole Auxins and Antiauxin

Studies on Chlorinated Indole Auxins and Antiauxin

Improved High-Speed Countercurrent Chromatograph with Three Multilayer Coils Connected in Series. III. Evaluation of Semianalytical Column with Various Biological Samples and Inorganic Elements

Abstract The partition efficiency of the counter-current chromatographic centrifuge was improved by mounting a set of three multilayer coils prepared from a 1.07 mm I.D. and ca 300 m length of PTFE (polytetrafluoroethylene) tube with a total capacity of 270 ml. The high performance of the present apparatus has been successfully demonstrated in separations of various testing samples which include DNP (dinitrophenyl) amino acids, indole auxins, tetracycline derivatives, flavonoids, bacitracin, triterpenoic acids, and lanthanoid chlorides.

Evaluation of Analytical Countercurrent Chromatographs: High-Speed Countercurrent Chromatograph-4000 Vs. Analytical Toroidal Coil Centrifuge

Abstract Performance of two countercurrent chromatographic models, high speed countercurrent chromatograph (HSCCC-4000) and analytical toroidal coil centrifuge (TCC), is evaluated in terms of theoretical plate number, resolution factor and separation time to assess their analytical capability. A series of experiments was conducted to investigate the effects of internal diameter and length of the coiled column, and flow rate of the mobile phase on the separation of indole auxins in two-phase solvent systems composed of n-hexane-ethyl acetate-methanol-water at different volume ratios. The three components of indole auxins were completely resolved in 16 min with a HSCCC system equipped with a multilayer coil of a 0.55 mm I.D. PTFE tube with theoretical plates ranging from 1290 to 829. Similar separation was achieved in 24 min with a TCC systems equipped with a 0.3 mm I.D. PTFE tube with theoretical plates ranging from 1811 to 969. It is concluded that both systems have comparable analytical capability at the...

Improved high-speed counter-current chromatography with three multilayer coils connected in series : II. Separation of various biological samples with a semi-preparative column

The semipreparative capability of the newly developed high-speed counter-current chromatograph equipped with a set of three multilayer coils has been demonstrated in separations of a variety of biological samples including triterpenoic acids, indole auxins, bacitracin, flavonoids and tetracycline derivatives, each with a suitable two-phase solvent system. The sample quantities ranging from 50 to 500 mg were efficiently separated within a few hours. The separation of tetracycline derivatives was remarkably improved by adding ammonium acetate to the solvent system.

Influences of the new auxins 4-carbomethoxyvinylenephenyl indole-3-butyrate and 2, 4-dichlorophenyl indole-3-butyrate on adventitious root formation

Tests were conducted with cuttings of bean (Phaseolus vulgaris L. cv. Top Crop) and jack pine (Pinus banksiana Lamb.) seedlings to determine the influences of the new auxins 4-carbomethoxyvinylenephenyl indole-3-butyrate (CMVP-IBA) and 2,4-dichlorophenyl indole-3-butyrate (DCP-IBA) on adventitious rooting. For comparison, bean cuttings were used to determine the effects on adventitious rooting of the unsubstituted phenyl ester of IBA, phenyl indole-3-butyrate (P-IBA). Results indicated that, compared with IBA, treatment with CMVP-IBA resulted in less adventitious rooting of jack pine and bean cuttings, treatment with DCP-IBA resulted in equal adventitious rooting of jack pine and bean cuttings, and treatment with P-IBA resulted in greater rooting of bean cuttings (jack pine was not tested). Based on results of the present and previous research, the phenyl esters of indole auxins have equal or greater root-inducing activity, compared with the parent auxin, only if their phenolic moieties do not contain a substituent (e.g., P-IBA) or contain only a simple substituent such as -OH (e.g., 3-hydroxyphenyl indole-3-acetate) or -Cl (e.g., DCP-IBA), rather than a more complex substituent like -CH=CH—COOCH3 (e.g., CMVP-IBA). However, further research is needed to test the accuracy of these generalizations.

Improved cross-axis synchronous flow-through coil planet centrifuge for performing counter-current chromatography : II. Studies on retention of stationary phase in short coils and preparative separations in multilayer coils

Performance of the apparatus was evaluated in terms of stationary phase retention, partition efficiency and sample loading capacity. Preliminary studies with short coils revealed high retention of the stationary phase under a proper combination of the head—tail elution and planetary motion. Preparative capability of the apparatus was successfully demonstrated on efficient multigram separations of 2,4-dinitrophenyl amino acids, indole auxins, and bacitracin in a pair of large multilayer coils with a total capacity of 1.5 1.

The green algal pigment caulerpin as a plant growth regulator

Caulerpin, a green algal pigment possessing a unique bis-indole structure, has been shown to be a plant growth regulator. Root growth assays were conducted with caulerpin and its hydrolysis product, and the results were compared with data obtained with indole-3-acetic acid (IAA), indole-3-pyruvic acid (IPA) and indole-3-acrylic acid (IAcA). This study has indicated that caulerpin, in essence a dimer of indole-3-acrylic acid, behaves much like the indole auxins.

Plant Hormone Analysis by Countercurrent Chromatography

Abstract Countercurrent chromatography (CCC) has been successfully applied for the separation of plant hormones; namely, indole auxins, gibberellins, cytokinins and abscisic acid. In our present study three different types of CCC devices were evaluated for their performance in separation of plant hormones with a special emphasis on analysis and purification of abscisic acid (ABA). A large-scale preparative CCC apparatus consisting of a slowly rotating coil assembly was used for preliminary separations of ABA from a large volume of crude plant extracts. The toroidal coil planet centrifuge (CPC) for analytical-scale separations was subsequently applied for purification of ABA, the final confirmation being obtained by HPLC and combined gas chromatographic-mass spectrometric method. This two-step procedure utilizing preparative CCC and toroidal CPC was successfully applied for determination of ABA content in several plant tissues. A recently introduced high-speed CCC apparatus was tested for semipreparative s...

Production of plant growth regulators by some Fusarium species.

About 10 species of Fusarium were screened and tested for their growth regulatory activity on oat coleoptile straight growth test. Culture filtrates of four species contained growth-stimulating factors while others showed growth-inhibition responses on oat sections. On the whole, F. moniliforme was found to be produce higher stimulatory effects. Chromatographic analysis revealed the presence of several indole auxin and gibberellin-like plant-growth regulators in their culture filtrates. The quantity and biological activity of each indole spot was also measured.

Separation of plant hormones by counter-current chromatography

Counter-current chromatography (CCC) using the toroidal coil planet centrifuge was applied to the separation of plant hormones. Indole auxins were separated in either hexane-ethyl acetate-methanol-water (0.6:1.4:1.0:1.0) or chloroform-acetic acid-water (2:2:1). The latter solvent system was especially useful for the separation of abscisic acid from indole-3-acetic acid. Gibberellins (GA 3, GA 4 and GA 7) were separated from each other in ether-methanol-phosphate buffer (pH 7) (3:1:2). The CCC method was suitable for the separation of four cytokinins in ethyl acetate-methanol-phosphate buffer (pH 7) (3:1:3). Abscisic acid from Zoysia grass seed was successfully analysed by CCC. The potential practical application of CCC to plant hormone analysis is discussed.

A method for the quantitation of indole auxins in the picogram range by high performance gas chromatography of their N-heptafluorobutyryl methyl esters

A method is described for the extraction and HPGC-ECD quantitation of indolic acids in the picogram range. It includes an improved GLC technique and a new derivatization reagent; it is applied to the estimation of IAA in Bryonia leaves.

Metabolism of Ingested Auxins in the Bug Lygus disponsi : Indole Compounds Appearing in Excreta of Bugs Fed with Host Plants and the Effect of Indole-3-Acetic Acid on the Feeding

Several indole compounds were detected in the excreta of bugs, Lygus disponsi LINNAVUORI, fed with rape flower bud. The Rf values on thin layer chromatogram were 0.02, 0.13, 0.18, 0.25 and 0.30 in the solvent system butanol-ethanol-water (76 : 19 : 5). These were in general accord with those of the indole-3-acetic acid (IAA) metabolites in excreta of bugs fed with IAA. Significantly high auxin activiry was detected in the crude excreta of the bugs fed with various host plants, but not in the fractions of the excreta fractionated by thin layer chromatography. IAA in high concentration inhibited the feeding of the bugs, whereas IAA in suitable concentration had a tendency to simulate it in the presence of sucrose. The significance of indole auxin metabolism in the nature is discussed.

A specific radioimmunoassay for nanogram quantities of the auxin, indole-3-acetic acid

We have developed a specific radioimmunoassay [RIA] for indole-3-acetic acid (IAA) in the 0.2 ng to 12 ng range which, in principle, can be extended to other indole auxins as well. Methods are presented for obtaining suitable antibody, for the RIA procedure, and for measuring IAA in methanolic extracts of plant tissues. Antibody specific for IAA was obtained from rabbits immunized with IAA bound to bovine serum albumin by formaldehyde treatment. In assays with this antibody, 2,4-dichlorophenoxyacetic acid and indoles structurally related to IAA reacted from 300- to 3000-fold less than did IAA itself. However, α-and β-naphthaleneacetic acid reacted significantly and hence interfered with the assay. Extracts of tobacco (Nicotiana tabacum L.) tissue were immunoassayed after partial purification by buffer-ether partition. Crown-gall tumor tissue, which is auxin-autotrophic, and pith tissue depleted of auxin by the diffusion method contained, respectively, 26.7 ng and <0.5 ng extractable IAA per gram fresh weight.

Phenol auxins and erlich reactors in soils

An attempt was made to isolate indole auxins from four different soil types (cultivated, coniferous woodland, deciduous woodland, grassland), by the use of a variety of extracting agents. Despite the fact that over 100 soil extracts were examined, we were unable to demonstrate the presence of indole auxins in these soils. Erlich reacting compounds were however, extracted. On further analysis these compounds proved to be phenolic acids, two of which possessed auxin activity. These findings are discussed in relation to the current evidence regarding the distribution and characteristics of soil auxins.

Indole auxins in spinach plants grown in long and short days

The endogenous auxin-like substances were analyzed in the shoot extracts of young spinach seedlings, exposed to photoperiodic induction. At least eight indole auxins were found. One of them was identified as tryptophan, the other one is most probably IAA. The plants grown in long days had a higher level of ether soluble auxins than the controls in short days. Separate extractions of plants after each of the eight inductive days showed that the auxin content was not constant, but subjected to irregular oscillations. However, parallel oscillations were also found in control plants grown in short days. Staminate plants were found to contain more endogenous auxins than the pistillate ones. It is concluded that the quantitative changes in auxins during the photoperiodic induction are probably not related to flowering, but to some other growth process, common to all plants in that phase of growth. The higher level of auxins in staminate plants may be the cause of their faster elongation before the onset of flowering.

Cyclic AMP Promotion and Abscisic Acid Inhibition of α-Amylase Activity in the Seeds of Rice (Oryza sativaL.)

Cyclic AMP Promotion and Abscisic Acid Inhibition of α-Amylase Activity in the Seeds of Rice (<i>Oryza sativa</i>L.)

Occurrence and distribution of acidic indole auxins in lucerne at various stages

AbstractThe variation of concentration of ‘free’ acidic indole auxins has been studied in different lines of lucerne at various stages of growth. Acidic indole auxins were separated by gas chromatography from the plant ethanolic extract.A large decrease in indole‐3‐acetic acid (IAA) and indole‐3‐carboxylic acid (ICA) was recorded as the plants aged. The trend during the life‐cycle is the same either in open pollinated progenies or in clonal lines. The IAA and ICA contents in various lines, determined in plants at the same stage, differ considerably from each other; this difference may have a genetic basis. The distribution in different parts of a single plant is a falling gradient in auxin concentration from the young to the mature tissues.By comparing root formation with ‘free’ auxin concentration of stem cuttings, evidence is collected which suggests that IAA may have primary action in the regulation of root formation.