Volume 47, Issue 3-4

The 5,6-dimethylbenzimidazole unit of vitamin B 12 is formed from riboflavin via FMN in aerobic and some aerotolerant bacteria. Thereby C-1′ of the ribityl side chain gets C-2 of 5,6-dimethylbenzimidazole. Experiments with homogenates of Propionibacterium shermanii on the fate of C-atoms 2′ to 5′ of the ribityl side chain of riboflavin in this transformation are reported. It was found that [5′- 3 H]riboflavin leads to radioactive sugar phosphates. These were isolated, dephosphorylated, and separated. Thus 3 H-D-fructose and 3 H-D-glucose were detected. The degradation of 3 H-D-glucose revealed that 14 per cent of the of the total radioactivity was located in C-1, and 86 per cent in C-6. This indicates that during 5,6-dimethylbenzimidazole biosynthesis a three carbon unit is formed from the ribityl side chain of riboflavin.

Two new isoflavone dimers, kudzuisoflavone A and B have been isolated from yeast extracttreated cell suspension cultures of Pueraria lobata (Leguminosae). Their structures were characterized by spectral data, and they were considered to be formed by phenol oxidative coupling of two isoflavone moieties through C -C ortho-ortho and ortho-para couplings, respectively.

The relationships between the chemical structures and the odor characteristics were examined on a series of highly purified n-hexenols and n-hexenals (14 isomers). The C 6 -enals and the corresponding alcohols resembled in odor characteristics each other. However, the odor strength of the aldehydes was 100 - 1000 times higher than those of the alcohols. In the principal components analysis, the position of carbon-carbon double-bond in the C 6 -compounds was closely related to the scores of the first principal component, whereas the functional groups, hydroxy and formyl groups, were related to those of the second principal component except for (Z)-3-hexenol.

The surface structures of beech ( Fagus sylvatica) leaf waxes were studied by SEM and correlated with chemical compositions of the extracted wax lipids during one vegetation period. The very young leaflets just unfolding from buds contained already a wax layer without any wax sculptures or crystalloids. This wax layer is quite different in yield and composition to that of mature leaves. With the unfolding of beech leaves, a dynamic biosynthesis of several wax lipids was started, but the biosynthesis of wax esters was not continued further. Ten days after leaf unfolding the de novo biosynthesis of aldehydes could be detected for the first time. Aldehyde amount increased rapidly to about 13% of the wax. The predominant individual wax lipids synthesized were aldehydes, alcohols and fatty acids with C 28 and hydrocarbons with C 27 chain lengths, respectively. The biosynthesis of wax lipids in beech leaves was completed at the end of May and remained nearly constant in quantity and com position during the remaining season. At the same time when aldehydes were found for the first time, wax sculptures were observed on beech leaf waxes coming out of the continuous wax layer, exclusively on the upper leaf side. These wax sculptures increased in size and quantity in the following time and were present on the upper leaf side all over the season, only some wax sculptures show a trend to crystalline forms.

Bud exudates from 9 poplars showing a range of resistance to bacterial canker and poplar rust were analyzed by gas chromatography-mass spectrometry and the components of the bud exudate identified. Those compounds previously reported in the literature to have notable antimicrobial activity were listed and the quantities present in the bud exudates of the specimens analyzed were compared. No correlation was observed between the amounts of these com pounds and the known resistance of the poplars to bacterial canker or rusts.

We present results on the compartmentation of carbohydrates such as sucrose, glucose, fructose, and mannose in different parts of an ectomycorrhiza established between Picea abies and Amanita muscaria and compare it with non-mycorrhized fine roots. Lyophilized mycorrhizas and fine roots (< 2 mm length) were dissected into about 0.5 mm thick slices which represent 4 zones of different physiological functions. The total amount of the analyzed carbohydrates was about 30% higher in non-mycorrhized (n-myc) compared to mycorrhized (myc) fine roots, with sucrose being the dominating sugar in both root types. A longitudinal distinction of sucrose pools showed lowest levels in the middle parts of a mycorrhiza, which represent areas of most intense symbiotic interaction. Fine roots without fungal infection did not show longitudinal variations in sugar content.

Glucose was found to be a potential stimulator of carthamin formation in triturated florets of C. tinctorius. At a 10 mM glucose level, carthamin-producing activity was raised from 3.2- fold (upper) to 2.0-fold (lower) level compared with the control having no sugar. Glucose oxidase (β-D-glucose: oxygen 1-oxidoreductase, EC 1.1.3.4) from Aspergillus niger could catalyze the conversion of precarthamin to carthamin, indicative of flower colour modification in C. tinctorius. For optimum reaction, precarthamin, β-D-glucose, and oxygen were required. No manganese enhanced the catalytic enzyme process.

We have observed that high SOD-like function (decomposition of superoxide anion) was observed for several iron(III) com pounds with tripodal ligands and several oxovanadium(IV) compounds, and also that these compounds exhibit high catalytic activity for oxidative cleavage of 3,5-di-tert-butylcatechol in non-donating solvents such as dichloromethane or nitromethane. These are suggesting that the same reaction intermediate exists in reaction mixtures of both the SOD-like and catecholase-like functions of these compounds. Based on these facts, we have proposed a new reaction mechanism for the oxidative cleavage of catechol catalyzed by the native non-heme iron dioxygenases; this includes formation of an iron(III)-peroxide adduct as a reaction intermediate, which exhibits electrophilic nature.

Keywords A study of the subcellular localization of peroxidase isoenzymes capable of oxidizing 4-hydroxystilbenes to viniferin-type compounds has been carried out in cultured cells derived from Vitis vinifera cv. Gamay berries. The study revealed that these isoenzymes are apparently located, in soluble form, in the vacuolar sap and are strongly inhibited by vacuolar anthocyani( di)ns in the concentration range found in planta. These results suggest that the inhibition of the 4-HS peroxidase system in vacuoles, as a consequence of anthocyani(di)n accumulation, could be responsible for the loss of viniferin production potential which accompanies grape berry veraison.

Hairy root cultures of Peganum harmala established by genetic transformation with Agrobacterium rhizogenes wild type strains A4, 15834 or carrying the binary vector pLTCgus 1, respectively, were analyzed with respect to their tryptophan-derived metabolites. Beside the previously detected β-carboline alkaloids harmine, harmol, harmaline and harmalol all hairy root cultures were found to contain the β-carboline glucoside ruine and serotonin. The appearance of the root cultures and the levels of β-carboline alkaloids were greatly affected by the nitrogen source of the medium. In the presence of higher levels of ammonium ions (as in MS medium) the roots were short and thick, while in B50 medium (low ammonium) they were thin and long. Ammonium ions had a negative effect on the levels of β-carbolines. The best root cultures accumulated on B50 1.5-2% β-carbolines on a dry mass basis and in the presence of 6% sucrose ca. 10 mg β-carbolines/L x d were formed. In the presence of phytohorm ones root cultures were converted into low producing suspension cultures, from which the highly productive roots were reinduced on phytohormone-free medium.

Pheophorbide a was prepared from a mixture of chlorophylls a and b by differential extraction with HCl and saponification. The insertion of the following metal ions was investigated: Mg, Zn, Co, Cu, Ni. In the enzyme test with chlorophyll synthetase, the metallopheophorbides fall into two categories: the Mg- and Zn-complexes are good substrates, the Co-, Cu- and Ni-complexes are neither substrates nor competitive inhibitors for the enzyme reaction. This corresponds to two categories of complex structures: Mg- and Zn-porphyrins prefer pentacoordinate square-pyramidal structures, Co-, Cu- and Ni-porphyrins prefer tetracoordinate square-planar structures. A model for substrate binding to chlorophyll synthetase is proposed.

The proton-translocating ATPase from chloroplasts, CF 0 F 1 , was isolated, purified and reconstituted into asolectin liposomes. The effect of the energy transfer inhibitor, tentoxin, on different functions of the enzyme was investigated. Tentoxin does not inhibit the nucleotide release during energization by a pH /ΔΨ jump, i.e. the activation of the enzyme is not influenced. ATP synthesis driven by a pH /ΔΨ jump and multi-site ATP hydrolysis are completely inhibited by tentoxin, whereas uni-site ATP hydrolysis is not influenced.

A herbicide resistant Euglena mutant (MSI) has been obtained by adapting wild type cells to increasing concentrations of DCMU (3-(3′,4′-dichlorophenyl)-1,1-dimethylurea). Lower resistance levels towards DCMU and metribuzin were observed in MSI when compared with Euglena or Chlamydomonas mutants with Ser 264 to Ala substitutions. RNA-sequence analysis identified a Ser to Thr change at position 265 (equivalent to position 264 in other organisms), thus making it possible to compare the influence of amino acids Ser, Ala and Thr at identical positions on the inhibitory effect of structurally different herbicides in the same species.

Three hours after X-irradiation in vivo with 8 Gy the in vitro incorporation of [ 3 H]uridine into total RNA of liver(L)- and brain(B)-cells of the chick embryo was reduced to 77% and 90%, respectively; the m RNA fraction was strongest inhibited. Under the same conditions, protein synthesis of L-cells declined to 62%, while protein synthesis of B-cells was not influenced. RNA and protein metabolism was not altered following X-irradiation in vitro (1.75 -56 Gy). - Compared to thymic - and splenic cells of the rat, chicken embryo cells exhibited higher constitutive poly(adenosine diphosphate-ribose)polymerase activity and lower X-irradiation-induced DNA damage. - Whereas the slight inhibition of RNA and protein synthesis by X-irradiation in ovo may be an abscopal and/or secondary phenomenon reflecting DNA and/or cellular damage, the present investigations comprising various cell types argue for an efficient DNA repair in chicken embryo cells caused, at least partly, by a high constitutive activity of DNA repair proteins.

Bifilarly BU-substituted ColE 1 plasmid and monofilarly BU-substituted M 13 phage DNA were irradiated with UV light of 313 nm. Using agarose gel electrophoresis and “reversed phase” HPLC technique ssb, dsb induction and uracil formation, respectively, could be detected in the irradiated DNA in dependence on the UV fluence. The analysis of the strandbreaks in bifilar Col E1 DNA shows a significant part of directly induced dsb. Cross sections of ssb induction from 4.1 m 2 /J x 10 7 in 28%, 3.9 m 2 /J x 10 7 in 55% and 3.1 m 2 /J x 10 7 in 8 5 -9 0% BU-substituted DNA were calculated. The cross section for dsb induction was found to be 0.04 m 2 /J x 10 7 , estimated from the linear part of the fluence effect curve. In monofilar M13 DNA a linear fluence effect curve for dsb induction was obtained. Excluding other than the direct production of dsb by using an in vitro approach for M13 DNA , the results strongly support the hypothesis that dsb can be induced by one photochem ical absorption event. The cross section for ssb was 3.8 m2/J x 107 and for dsb 0.05 m2/J x 107 in 41.5% monofilarly BU-substituted M13 DNA . The comparison of ssb, dsb, and uracil production in bifilar and monofilar DNA with similar BU substitution showed no significant difference between the two DNA systems (ColE 1, M 13), indicating that the location of BU molecules in one or in both DNA strands will not lead to a different number of lesions after UV 313 exposure.

2′,3′-O-(2,4,6-trinitrophenyl)ADP (= TNP-ADP) is a competitive inhibitor of photophosphorylation of A DP. Here, the interaction of TNP-ADP with the H + -translocating chloroplast ATPase and other thylakoid membrane proteins was studied more in detail and compared with the interactions of the parental compounds ADP and 2,4,6-trinitrophenol (= picric acid). The following results were obtained: (1) Preilluminated thylakoids incorporate about 1 nmol [ 14 C]ADP per mg chlorophyll or the same amount of T N P-[14C]ADP in the following dark. If T NP-[I4C]ADP is present in the light, however, up to 4 nmol label is bound per mg chlorophyll whereas the amount of incorporated [ 14 C]ADP is unchanged. [ 14 C]ADP is exclusively bound to CF 1 , yielding a molar ratio of 1 nucleotide per enzyme. Likewise 1 mol of 14 C label per mol enzyme is found in CF, when thylakoids are illuminated in the presence of TNP-[ 14 C]ADP; the residual bound radioactivity is related with other components of the thylakoid membrane. (2) TNP-ADP as well as picrate are electron acceptors of the photosynthetic electron transport chain. Picramate is one main product of photosynthetic reduction of picrate. Hence at least one of the three nitro groups of the trinitrophenyl moiety is reduced to an amino group. However, additional non-identified, reactive intermediates are formed, which can bind to membrane components. About 20% of the bound label is covalently linked to membrane proteins. (3) The patterns of covalently labeled polypeptides are identical whether [ 14 C]picrate or TNP- [ 14 C]ADP is employed, except for α and β subunits of CF 1 which are specifically labeled in the presence of TNP-[ 14 C]ADP. This reaction is protected by (A DP + P i ) > ATP > ADP > P i . Most likely radicalic intermediates of picrate or TNP-ADP photoreduction are responsible for covalent binding. In case of CF, the radicalic intermediate of TNP-ADP reduction may be accomodated by its ADP moiety into a nucleotide binding site. Subsequent covalent linkage via the reactive phenyl ring substituent seems to occur in both, the nucleotide and the P i binding domain.

The protective effects of a series of phenolic compounds, phenolic acids and flavonoids on the cytotoxicity of CCl 4 in rat hepatocytes were studied. A number of flavones, 7,8-dihydroxyflavone, luteolin and hypolaetin-8-glucoside, flavonols, morin, quercetin, robinetin and gossypin, phenolic acids, gallic, caffeic and chlorogenic acids, as well as the flavane ( + )-catechin significantly inhibited alanine amine transferase (ALT) release. Catechol groups are determinant for the protective activity of flavonoids and cinnamic acid derivatives, as well as the resorcinol or pyrogallol moieties in the B ring of flavonoids. In benzoic acid derivatives a pyrogallol group is required. This feature is associated with the inhibition of ALT spontaneous release.

The two brassinosteroids, 22S′,23S′-homobrassinolide and 22S′,23S′-homocastasterone are weak competitors of the binding of [ 3 H]ponasterone A to the intracellular ecdysteroid receptor from the epithelial cell line from Chironom us tentans. The relative affinities to the ecdysteroid receptor are 0.001 for both brassinosteroids as compared to 20-OH-ecdysone and 0.1 in comparison to ecdysone. Both substances exert morphological effects similar to those observed with 20-OH-ecdysone. Like moulting hormones both brassinosteroids inhibit chitin synthesis. However, these effects were observed only at rather high concentrations (10 -5 to 10 -4 m) which were cytotoxic for 22 S′,23 S′-homobrassinolide.

Dufour’s gland secretion of several species of the desert ant Cataglyphis from different geographical localities was analyzed. The secretions constituted mostly of alkanes ranging from undecane to nonadecane. Species specificity is expressed as variations in the major component as well as the relative intensities of the additional constituents. Phylogenetically related species that are allopatric exhibited similar secretory composition whereas their sympatric counterparts had disparate composition, suggesting that character displacement occurred. Analyses of colonies of C. cursor from different localities also showed divergence in their glandular composition.

Within the beetle family Anthicidae or ant-like flower beetles the canthariphilous species Notoxus monoceros shows a pair of notch-like structures on the apices of male elytra. A reservoir- like swelling at the apex of each elytron contains a bilobed gland. Its glandular tubules run into the sclerotized notch. Gaschromatographic investigations of elytral notches and the testes of cantharidin-fed and unfed male Notoxus monoceros revealed huge quantities of cantharidin stored in these organs in fed specimens. Other body parts, such as legs, elytral parts or flight muscles, had low cantharidin-titres, both in cantharidin-fed and unfed beetles. The cantharidin- titres of unfed individuals were very low compared with cantharidin-fed specimens, although cantharidin was present, mostly in notches. During sexual display the females bite several times into these notches and easily copulate, if the males had access to offered cantharidin crystals. Thus, the elytral notches of male Notoxus monoceros seem to function as „testorgans” for females which are able to select males with high cantharidin loads prior to copulation. Even small quantities of cantharidin stored in notches of field caught specimens seem of advantage for males in competing for a female partner.

CCAP (Crustacean Cardioactive Peptide), Proctolin, FMRFamide, Met- and Leu-enkephalin. Substance P, RPCH (red pigment concentrating hormone) and PDH (pigment dispersing hormone) were applied to the isolated retina of the crayfish Orconectes limosus. Changes in light sensitivity, measured as changes of the amplitude of the electroretinogram (ERG) were observed after application of RPCH, PDH and CCAP. RPCH caused an increase of the ERG amplitude to 133% of its reference value whereas PDH and CCAP decreased the amplitude to 78 % and 30% respectively. A dose-response curve half-maximal effect.

The sesquiterpenoic alcohol nerolidol was separated into its 4 stereoisomers by MPLC of the diastereomeric (1 S, 4 R)-camphanoates. An analytical GC method was found by which both the enantiomeric pairs of (Z)- and (E)-nerolidol are resolved on a chiral cyclodextrin stationary phase. The olfactoric properties of the nerolidol stereoisomers were investigated.

Analysis of GC-MS data revealed 58 com ponents of bud exudate from P. cathayana (Section Tacamahaca) and 40 components of bud exudate from P. szechuanica (Section Tacamahaca) of which 47 and 35 respectively were phenolics. The bud exudates were very similar in composition, containing flavanones and chalcones (61% and 51% respectively), flavones (15% and 12%) and pentenyl caffeates (11% and 6%). The major single component of the exudate of both poplars was pinobanksin- 3-acetate (26% and 33%). The bud exudates of P. cathayana and P. szechuanica are similar to those of European and American Section Aigeiros poplars and unlike those of North American Section Tacamahaca poplars.

Analysis of GC-MS data revealed 76 components of bud exudates from Populus koreana, P. maximowiczii and P. suaveolens (Section Tacamahaca), of which 49 were phenolics. The bulk of the exudates were composed of phenones, pinobanksin-3-acetate and aliphatic esters of caffeic acid. Bud exudates of P. koreana and P. m aximowiczii were virtually identical in composition and closely resembled those of P. suaveolens. The three poplars were alike in having as major com ponents of their bud exudate pentanophenones, which do not occur in the bud exudates of most other poplars. These three Asiatic poplars appear more closely related chemotaxonomically to Section Aigeiros poplars than to those of Section Tacamahaca.

Periodate-induced degradation of cyclobrassinin monosulphoxide gives brassilexin in 40% yield. The sequence: cyclobrassinin to brassilexin through cyclobrassinin sulphoxide is now demonstrated for the group of phytoalexins found in the Cruciferae.

Apparent Michaelis constants for nucleotides in transcription of yeast tRN Agene by hom ologous RNA polymerase III with auxiliary protein factors, were found to be remarkably higher in initiation than in elongation of RNA chain. This supports presumptions regarding topological similarities between catalytic centers of bacterial and eukaryotic RNA polymerases.