The influence of the heavy metal ions Hg++, Cu++, Cd++ and Zn++ and of arsenite on growth, amino acid uptake, protein- and heat shock protein synthesis was investigated in cell cultures of a heavy metal tolerant Silene vulgaris and the sensitive Lycopersicon peruvianum.A distinct tolerance of Silene cell growth in comparison to tomato cells against Cu, Cd and Zn was observed. Synthesis of the small heat stress proteins was induced in both species, however, with quantitative differences depending on species and metal ion. While in tomato cells all metals induce HSP synthesis, in Silene the induction by Cu and Cd was found to be significantly lower in comparison to Hg and arsenite, and Zn did not induce HSP. Therefore, generally a lower tolerance of the cells against a metal is connected with a higher HSP synthesis. From comparison of cell growth and HSP accumulation in the presence of metal ions it was further concluded that HSP synthesis is a part of HM stress response of tolerant and non tolerant cells as under heat shock, but HSPs are not responsible for the heritable metal tolerance of Silene.In contrast to heat shock, metal stress does not inhibit the cell protein synthesis directly. In cultured tomato and Silene cells the inhibition of protein synthesis under metal stress was found to be a consequence of the inhibition of amino acid uptake. Zn has no effect on amino acid uptake of Silene cells. It is concluded that only Zn tolerance of Silene seems to be related with membrane stability.

Both the vicinal side chain hydroxyl groups and the 6-oxo function of brassinosteroids were modified by fluorescence tagging. Dansylaminophenylboronic acid was used as a derivatizing agent to form fluorescent esters of brassinosteroids containing a side-chain cis-diol structure. 6-Oxo type brassinosteroids were derivatized by means of dansylhydrazine. The modified brassinosteroids, as far as possible derivatized both at the diol and the oxo group, were separated by HPLC and the optimal emission wavelength was determined. By this approach almost all brassinosteroids, including biosynthetic precursors, were susceptible to highly sensitive analysis in the fmol range. This method has been verified as an analytical tool to determine brassinosteroids in cell culture extracts and to monitor brassinosteroid biosynthetic enzymes. 24-Epibrassinolide has been detected in tomato cell suspension cultures. Several steps of brassinosteroid biosynthesis, including the Baeyer–Villiger oxidation of 24-epicastasterone to give 24-epibrassinolide, were monitored in vitro with protein preparations of the same cell culture line.

Synthetic transformations utilizing chromium(II) or chromium(III) reagents, mainly C-C-coupling reactions, are discussed. Chromium reagents find increasing application in complex total syntheses where other organometallics are difficult to apply. They are easy to prepare and exhibit extraordinary chemoselectivity and high diastereoselectivity. In this article emphasis is laid on recent results in synthetic procedures, on little known general aspects of (organo)chromium chemistry, and on areas not reviewed before. The most important recent advances include reactions catalytic in chromium ions, anti-selective Reformatsky-type aldol reactions with excellent asymmetric induction, domino radical/carbanion reactions, asymmetric chromium(III) catalyzed epoxide openings and homogeneous alkene polymerization catalysts. Some progress is also made in ligand controlled enantioselective reactions with Cr(II) reagents, but satisfying solutions remain a major challenge in the field.

Tissue-specific accumulation of phenylpropanoids was studied in mycorrhizas of the conifers, silver fir (Abies alba Mill.), Norway spruce [Picea abies (L.) Karst.], white pine (Pinus strobus L.), Scots pine (Pinus silvestris L.), and Douglas fir [Pseudotsuga menziesii (Mirbel) Franco], using high-performance liquid chromatography and histochemical methods. The compounds identified were soluble flavanols (catechin and epicatechin), proanthocyanidins (mainly dimeric catechins and/or epicatechins), stilbene glucosides (astringin and isorhapontin), one dihydroflavonol glucoside (taxifolin 3′-O-glucopyranoside), and a hydroxycinnamate derivative (unknown ferulate conjugate). In addition, a cell wall-bound hydroxycinnamate (ferulate) and a hydroxybenzaldehyde (vanillin) were analysed. Colonisation of the root by the fungal symbiont correlated with the distribution pattern of the above phenylpropanoids in mycorrhizas suggesting that these compounds play an essential role in restricting fungal growth. The levels of flavanols and cell wall-bound ferulate within the cortex were high in the apical part and decreased to the proximal side of the mycorrhizas. In both Douglas fir and silver fir, which allowed separation of inner and outer parts of the cortical tissues, a characteristic transversal distribution of these compounds was found: high levels in the inner non-colonised part of the cortex and low levels in the outer part where the Hartig net is formed. Restriction of fungal growth to the outer cortex may also be achieved by characteristic cell wall thickening of the inner cortex which exhibited flavanolic wall infusions in Douglas fir mycorrhizas. Long and short roots of conifers from natural stands showed similar distribution patterns of phenylpropanoids and cell wall thickening compared to the respective mycorrhizas. These results are discussed with respect to co-evolutionary adaptation of both symbiotic partners regarding root structure (anatomy) and root chemistry.

Treatment of the halophyte Mesembryanthemum crystallinum L. (ice plant) (Aizoaceae) with high intensities of white light resulted in a rapid cell-specific accumulation of betacyanins and flavonoids with 6-methoxyisorhamnetin 3-O-{[(2‴-E-feruloyl)-3‴-O-(β-d-glucopyranosyl)](2″-O-β-d-xylopyranosyl)}-β-d-glucopyranoside (mesembryanthin) as the predominant component, within bladder cells of the leaf epidermis. Induced accumulation of these metabolites was first detected 18 h after the initiation of light treatment in bladder cells located at the tip of young leaves followed by the bladder cells located on the epidermis of fully expanded leaves. UV-A light apparently is sufficient to induce accumulation of betacyanins and flavonoids. Application of 2-aminoindan 2-phosphonic acid, a specific inhibitor of phenylalanine ammonia-lyase (PAL; EC 4.3.1.5), not only inhibited the accumulation of flavonoids but also reduced betacyanin formation. Based on these observations we suggest these bladder cells as a model system to study regulation of betacyanin and flavonoid biosyntheses.

Based on protein sequence data and RT–PCR, a full length cDNA encoding betanidin 5‐O‐glucosyltransferase (5‐GT) was obtained from a cDNA library of Dorotheanthus bellidiformis (Burm.f.) N.E.Br. (Aizoaceae). 5‐GT catalyses the transfer of glucose from UDP‐glucose to the 5‐hydroxyl group of the chromogenic betanidin. Betanidin and its conjugates, referred to as betacyanins, are characteristic fruit and flower pigments in most members of the Caryophyllales, which fail to synthesise anthocyanins. The 5‐GT cDNA displayed homology to previously published glucosyltransferase sequences and exhibited high identity to sequences of several inducible glucosyltransferases of tobacco and tomato (Solanaceae). The open reading frame encodes a polypeptide of 489 amino acids with a calculated molecular mass of 55.24 kDa. The corresponding cDNA was expressed in Escherichia coli . The recombinant protein displayed identical substrate specificity compared to the native enzyme purified from D. bellidiformis cell suspension cultures. In addition to the natural substrate betanidin, ortho‐dihydroxylated flavonols and flavones were glycosylated preferentially at the B‐ring 4′‐hydroxyl group. 5‐GT is the first enzyme of betalain biosynthesis in plants, of which the corresponding cDNA has been cloned and expressed. The results are discussed in relation to molecular evolution of plant glucosyl‐ transferases.

The narcotic analgesic morphine is the major alkaloid of the opium poppy Papaver somniferum . Its biosynthetic precursor codeine is currently the most widely used and effective antitussive agent. Along the morphine biosynthetic pathway in opium poppy, codeinone reductase catalyzes the NADPH‐dependent reduction of codeinone to codeine. In this study, we have isolated and characterized four cDNAs encoding codeinone reductase isoforms and have functionally expressed them in Escherichia coli . Heterologously expressed codeinone reductase‐calmodulin‐binding peptide fusion protein was purified from E. coli using calmodulin affinity column chromatography in a yield of 10 mg enzyme l ‐1 . These four isoforms demonstrated very similar physical properties and substrate specificity. As least six alleles appear to be present in the poppy genome. A comparison of the translations of the nucleotide sequences indicate that the codeinone reductase isoforms are 53% identical to 6′‐deoxychalcone synthase from soybean suggesting an evolutionary although not a functional link between enzymes of phenylpropanoid and alkaloid biosynthesis. By sequence comparison, both codeinone reductase and 6′‐deoxy‐ chalcone synthase belong to the aldo/keto reductase family, a group of structurally and functionally related NADPH‐dependent oxidoreductases, and thereby possibly arise from primary metabolism.

In addition to a known alkaloid, some limonoids and coumarins, the new coumarins excavatins A–M have been isolated from Clausena excavata. Their structures have been assigned by NMR and CD investigations.

In addition to (−)-culantraramine and (−)-culantraraminol the bishordeninyl terpene alkaloids, (−)-culantraramine N-oxide, (−)-culantraraminol N-oxide and avicennamine, have been isolated from the leaves of Zanthoxylum avicennae. Their structures have been assigned by MS and especially by NMR investigations.

A tyrosine-hydroxylating enzyme was partially purified from betacyanin-producing callus cultures of Portulaca grandiflora Hook. by using hydroxyapatite chromatography and gel filtration. It was characterized as a tyrosinase (EC 1.14.18.1 and EC 1.10.3.1) by inhibition experiments with copper-chelating agents and detection of concomitant o-diphenol oxidase activity. The tyrosinase catalysed both the formation of L-(3,4-dihydroxyphenyl)-alanine (Dopa) and cyclo-Dopa which are the pivotal precursors in betalain biosynthesis. The hydroxylating activity with a pH optimum of 5.7 was specific for L-tyrosine and exhibited reaction velocities with L-tyrosine and D-tyrosine in a ratio of 1:0.2. Other monophenolic substrates tested were not accepted. The enzyme appeared to be a monomer with an apparent molecular mass of ca. 53 kDa as estimated by gel filtration and SDS-PAGE. Some other betalain-producing plants and cell cultures were screened for tyrosinase activity; however, activities could only be detected in red callus cultures and plants of P. grandiflora as well as in plants, hairy roots and cell cultures of Beta vulgaris L. subsp. vulgaris (Garden Beet Group), showing a clear correlation between enzyme activity and betacyanin content in young B. vulgaris plants. We propose that this tyrosinase is specifically involved in the betalain biosynthesis of higher plants.