Modulation of plant morphology, root architecture, and cell structure by low vitamin C in Arabidopsis thaliana.

Ascorbic acid (AA) fulfils many essential functions in plants. It is a key antioxidant and an important reducing substrate for a number of enzymes. The effects of low AA on plant architecture and leaf ultrastructure were studied in Arabidopsis thaliana mutants, which have constitutively moderately low (vtc1) or very low (vtc2) leaf AA contents compared with the wild type. Shoot development was comparable in all accessions over the first 14 d of growth. The production of primary roots was slightly different in vtc1, vtc2, and wild-type plants. However, the most notable difference was that a high proportion of the primary roots of the vtc2 plants grown on soil had lost the wild-type responses to gravity. The vtc mutants showed the antagonistic interaction between nitrate and sugar in the regulation of lateral root (LR) development that was observed in the wild type. However, the vtc2 mutants produced greater numbers of longer LRs than wild-type or vtc1 plants at all levels of nitrate. At later stages of development, the vtc rosettes were smaller than those of the wild type and the leaves showed intracellular structural changes that are consistent with programmed cell death (PCD). PCD symptoms such as nuclear chromatin condensation, the presence of multivesicular bodies, and extensive degradation and disorganization of the grana stacks were observed in 8-week-old vtc2 leaves and in 10-week-old vtc1 leaves. The data presented here illustrate the importance of tissue AA contents in regulating whole plant morphology, cell structure, and development.

High-performance liquid chromatographic separation of natural and synthetic desulphoglucosinolates and their chemical validation by UV, NMR and chemical ionisation-MS methods.

Methods are described for the optimised extraction, desulphation and HPLC separation of desulphoglucosinolates. These methods provide rapid separation, identification and quantitative measurements of glucosinolates extracted from Brassica napus L and related crops, of unusual glucosinolates found in crucifer weed species, and also of synthetic alkylglucosinolates. The desulphoglucosinolates used in these studies were either chemically synthesised (at least one example from each major structural class), or purified from various plant sources. Validation of the identities of the desulphoglucosinolates was by comparison of retention times with standards, and by UV, 1H- and 13C-NMR and chemical ionisation MS analysis. A list of useful species, and the specific tissues, from which high concentrations of standards can be extracted is included.

Differences in glucosinolate patterns and arbuscular mycorrhizal status of glucosinolate-containing plant species.

Under defined laboratory conditions it was shown that two glucosinolate-containing plant species, Tropaeolum majus and Carica papaya, were colonized by arbuscular mycorrhizal (AM) fungi, whereas it was not possible to detect AM fungal structures in other glucosinolate-containing plants (including several Brassicaceae). Benzylglucosinolate was present in all of the T. majus cultivars and in C. papaya it was the major glucosinolate. 2-Phenylethylglucosinolate was found in most of the non-host plants tested. Its absence in the AM host plants indicates a possible role for the isothiocyanate produced from its myrosinase-catalysed hydrolysis as a general AM inhibitory factor in non-host plants. The results suggest that some of the indole glucosinolates might also be involved in preventing AM formation in some of the species. In all plants tested, both AM hosts and non-hosts, the glucosinolate pattern was altered after inoculation with one of three different AM fungi (Glomus mosseae, Glomus intraradices and Gigaspora rosea), indicating signals between AM fungi and plants even before root colonization. The glucosinolate induction was not specifically dependent on the AM fungus. A time-course study in T. majus showed that glucosinolate induction was present during all stages of mycorrhizal colonization.

Indole glucosinolate and auxin biosynthesis in Arabidopsis thaliana (L.) Heynh. glucosinolate mutants and the development of clubroot disease.

Mutants and wild type plants of Arabidopsis thaliana were analysed for differences in glucosinolate accumulation patterns, indole-3-acetic acid (IAA) biosynthesis and phenotype. A previously identified series of mutants, termed TU, with altered glucosinolate patterns was used in this study. Only the line TU8 was affected in shoot phenotype (shorter stems, altered branching pattern). Synthesis of IAA and metabolism were not much affected in the TU8 mutant during seedling development, although the content of free IAA peaked earlier in TU8 during plant development than in the wild type. Indole glucosinolates and IAA may, however, be involved in the development of clubroot disease caused by the obligate biotrophic fungus Plasmodiophora brassicae since the TU3 line had a lower infection rate than the wild type, and lines TU3 and TU8 showed decreased symptom development. The decline in clubroot formation was accompanied by a reduced number of fungal structures within the root cortex and slower development of the fungus. Indole glucosinolates were lower in infected roots of TU3 and TU8 than in control roots of these lines, whereas in wild-type plants the differences were not as prominent. Free IAA and indole-3-acetonitrile (IAN) were increased in infected roots of the wild type and mutants with normal clubroot symptoms, whereas they were reduced in infected roots of mutants TU3 and TU8. These results indicate a role for indole glucosinolates and IAN/IAA in relation to symptom development in clubroot disease.

Residual dipolar couplings as new conformational restraints in isotropically 13C-enriched oligosaccharides.

We report the measurement of residual dipolar couplings for l3C-enriched NeuNAcalpha2-3Galbeta1-4Glc in a dilute liquid-crystalline medium. These couplings provide long-range conformational restraints that hitherto have not been available for oligosaccharides. We utilise these restraints in dynamical simulated annealing calculations, which support current models of the solution behaviour of the trisaccharide.

Involvement of Cytochrome P450 in Glucosinolate Biosynthesis in White Mustard (A Biochemical Anomaly).

One of the first steps in glucosinolate biosynthesis is the conversion of amino acids to their aldoximes. The biochemistry of this process is controversial, and several very different enzyme systems have been described. The major glucosinolate in white mustard (Sinapis alba) is sinalbin, which is derived from tyrosine via its aldoxime, and this conversion is catalyzed by a cytochrome P450 (Cyt P450) monooxygenase. Phenylethyl- and alkenylglucosinolates are also present in white mustard leaves, as are the enzymes catalyzing the relevant aldoxime formation from homophenylalanine and methionine homologs, respectively. These enzymes are similar to those found in Brassica sp. and are distinct from the tyrosine-dependent enzyme in that they contain no heme and are unaffected by Cyt P450 inhibitors. They are instead inhibited by the flavoprotein inhibitor diphenylene iodonium and by Cu2+. In both white mustard and oilseed rape (Brassica napus) methyl jasmonate specifically stimulates indolylglucosinolate biosynthesis and yet has no effect on sinalbin accumulation in either cotyledons or leaves of white mustard. White mustard appears to be unique among crucifers in having a Cyt P450 aldoxime-forming enzyme for biosynthesis of one glucosinolate, although it also contains all of the non-Cyt P450 enzyme systems found in other members of the family. Sinalbin biosynthesis in white mustard is therefore an inappropriate model system for the synthesis of other glucosinolates in crucifers, including canola and oilseed rape.

Sustained release metoprolol: a comparison with conventional formulation in the treatment of hypertension.

Thirty patients with essential hypertension (WHO Stage I or II) completed a double blind crossover trial in which they received metoprolol as monotherapy as either a slow release formulation of 200 mg once daily or a conventional formulation of 100 mg bd. Duration of treatment was six weeks on each preparation, given in randomised order. Mean values of resting blood pressure and heart rate, measured 22 to 24 hours after the daily dose of the SR formulation or 12-14 hours after the second dose of the conventional form were significantly lower than pretreatment levels. The two treatments were equally effective and equally well tolerated.