Volume 54, Issue 12

The liverwort Riella helicophylla was cultivated in vitro under aseptic conditions. The lipophilic extract of the plant material yielded seventeen monoterpenes and eleven diterpenes. Seven monoterpenes were hydroperoxides. From the diterpenes six belonged to the labdane type skeleton and one to the kaurane type, the other diterpenes were phytane derivatives.

Three new dammarane-type triterpenes, named arcapitin A, B and C, were isolated from the roots of Argyreia capitata (Convolvulaceae). They turned out to be 11α-acetoxydammar-24-ene-20(S)-ol-3-one, damm ar-24-ene-11a,20(S)-diol-3-one and 11a-acetoxydammar-24- ene-3β,20(S)-diol. While the same compounds were also detected in the aerial vegetative parts of this species, their occurrence in a number of other convolvulaceous species could not be confirmed.

Aerial parts of A chillea clusiana include two new chrysanthemol derivatives and three new guaianolides in addition to six known sesquiterpenoids. The structures of the new compounds were elucidated by spectroscopic methods.

Several species of freshwater green algae belonging to the order Zygnematales (Spirogyra crassa (Ktz.) Czurda, S. condensata (Vauch.) Czurda, S. longata (Vauch.) Ktz., S. juergensii Ktz., S. olivascens Rabenh. and Mougeotia viridis (Ktz.) Wittr.) were shown to have specific sterol content and characteristic monosaccharide composition of the biomass. Polysaccharide fractions were obtained by stepwise extraction of S. condensata and characterised by a determination of monosaccharides liberated after acid hydrolysis. Action of amyloglucosidase was used to prove the presence of starch in algal biomass. The main polysaccharide seems to be a complex mucilage composed of rhamnose, arabinose, xylose, galactose, and uronic acid, and the structure of this mucilage needs further investigation.

For a pyoverdin isolated from Pseudomonas putida it could be shown that Lys is incorporated into the peptide chain by its ε-amine group in contrast to the normally observed connection by the α-amino group. The structure elucidation of the pyoverdin by chemical degradation and spectroscopic methods is reported and the criteria for the distinction between α-and ε-connection in the case of Lys are discussed.

Neodictyoprolenol [(-)-(S)-(1,5Z,8Z)-undecatrien-3-ol], dictyoprolenol [(-)-(S )-(1,5Z)- undecadien-3-ol] and their acetates neodictyoprolene [(+)-(5)-3-acetoxy-(1,5Z,8Z)-undecatriene] and dictyoprolene [(+)-(S)-3-acetoxy-(1,5Z)-undecadiene], which are interesting as possible biosynthetic intermediates of the sex pheromones (dictyopterene B, C′ and D′) of brown algae, were synthesized by chemo-enzymatic methods through optical resolution of racemic neodictyoprolenol and dictyoprolenol using two lipases; Amano PS (Pseudom onas sp.) and Novozym 435 ® (Candida sp.). A combination of acylation of the alcohols and hydrolysis of the acetates by Novozym 435 ® produced neodictyoprolenol, neodictyoprolene, dictyoprolenol and dictyoprolene with high optical purities over 99% enantiomeric excess (e.e.). This snythetic methods will make it easier to search these compounds in marine algae and to study their biosynthesis.

Glutathione conjugation of 1-chloro-2,4-dinitrobenzene, 1,2-dichloro-4-nitrobenzene, NBD-Cl, monobromobimane and monochlorobimane was found to occur in epidermal tissue of onion bulbs (Allium cepa L.). Conjugation required the presence of glutathione S-transferases (GST). In order to follow glutathione conjugation microscopically, bimanes were utilized. Monochlorobimane was converted to a brightly fluorescing conjugate that was shown to be transported to the nucleus before sequestration in the vacuole occurred. GST activity was stimulated as well as induced by several electrophilic xenobiotics, by cycloheximide and by several glutathione conjugates. γ-Glutamylcysteine conjugates that are formed during enzymatic cleavage of glutathione conjugates in plants, were not active as inducers of enzyme activity. In the light of the stimulating effects of xenobiotic glutathione conjugates on GST activity, it is concluded that glutathione conjugates may act as signal molecules.

Based on structure-activity relationship studies, auxin analogs that can be covalently bound to a polymeric support are proposed. Molecular modeling studies were carried out by comparing different parameters of substituted phenoxyacetic acids with their auxin activity. A good correlation of the activity with the size and shape of the HOMO orbital of the acids was found. Accordingly, analogs with a substituent in the 5 position of the aromatic ring, capable to be bound to a polymeric matrix were synthesized and their auxin activity was evaluated with the wheat coleoptile elongation test. Compounds with a hydroxymethyl- and with a carboxymethyloxy- substituent were active in this test. Their use as probes for the 2,4-D receptor is proposed.

Cultures of the fungal strain Humicola lutea 110 were grown in a 3-l bioreactor. Effects of dissolved oxygen concentration (DO) on cell growth, intracellular protein content and antioxidant enzyme activities (SOD and catalase) were investigated. Controlling DO from 20 to 60% lead to: (I). The lethal phase of growth was reached faster; (ii) strong reduction of the intracellular protein content, and (iii) increase of antioxidant enzyme activities. The most efficient SOD biosynthesis was achieved at the 1st maximum of activity in the culture grown under DO uncontrolled conditions.

The phase transfer method was applied to perform the nucleophilic substitution of 2,6- dichloropurines by modified arylalkyl alcohol or phenols. Since under these conditions only the 6-halogen is exchanged, this method gives 2-chloro-6-aryloxy- and 2-chloro-6-arylalkoxypurines. 2-Chloro-6-benzylthiopurine was synthesized by alkylation of 2-chloro-6-thiopurine with benzyl bromide. The stereoisomers of 2-chloro-6-(1-phenyl-1-ethoxy)purine were obtained from R- and S-enantiomers of sec.-phenylethylalcohol and 2,6-dichloropurine. All derivatives were tested for inhibition with purified hexameric E. coli purine nucleoside phosphorylase (PNP). For analogues showing IC 50 < 10 μm, the type of inhibition and inhibition constants were determined. In all cases the experimental data were best described by the mixed-type inhibition model and the uncompetitive inhibition constant, K iu , was found to be several-fold lower than the competitive inhibition constant, K ic . This effect seems to be due to the 6-aryloxy- or 6-arylalkoxy substituent, because a natural PNP substrate adenine, as well as 2-chloroadenine, show mixed type inhibition with almost the same inhibition constants K iu and K iC . The most potent inhibition was observed for 6-benzylthio-2-chloro-, 6-benzyloxy-2-chloro-, 2-chloro-6-(2-phenyl-l-ethoxy), 2-chloro-6-(3-phenyl-l-propoxy)- and 2-chloro-6-ethoxypurines (K iu = 0.4, 0.6, 1.4, 1.4 and 2.2 μm, respectively). The R-stereoisomer of 2-chloro-6-(1pheny-1-ethoxy)purine has K iu = 2.0 μm, whereas inhibition of its S counterpart is rather weak (IC 50 > 12 μm). More rigid (e.g. phenoxy-), non-planar (cyclohexyloxy-), or more bulky (2,4,6-trimethylphenoxy-) substituents at position 6 of the purine base gave less potent inhibitors (IC 50 = 26, 56 and >100 μm, respectively). The derivatives are selective inhibitors of hexameric “high-molecular mass” PNPs because no inhibitory activity vs. trimeric Cellulomonas sp. PNP was detected. By establishing the ligand-dependent stabilization pattern of the E. coli PNP it was shown that the new derivatives, similarly as the natural purine bases, are able to form a dead-end ternary complex with the enzyme and orthophosphate. It was also shown that the derivatives are substrates in the reverse synthetic direction catalyzed by E. coli PNP

Following in vitro- and in ovo-exposure of chicken embryo cells, the level of bleomycin (BM) - induced damage was evaluated by using DNA synthesis, nucleoid sedimentation (SED), and viscometry of alkaline cell lysates (VISC). This damage was compared to Xirradiation, using 5.9-378 nM BM in vitro, 1.5-116 μg BM/egg in ovo, and 2-32 Gy, respectively, in vitro as well as in ovo. With respect to BM. the most notable result is the increase in DNA synthesis and VISC at the lowest concentrations of the drug. A decrease in both parameters was observed at high BM concentrations and following exposure to X-rays, concomitantly with an increase in SED. Regarding the radiomimetic drug BM and X-rays, different modes of DNA damage and DNA repair are suggested by previous investigations and the present results. Therefore, further evidence is presented, that the chicken embryo can act as a simple, rapid and inexpensive test system to characterize the biological effects of many nucleo- and/or cytotoxic agents.

Thirteen naturally occurring 1H-cyclopenta[b]benzofuran lignans of the rocaglamide type as well as one naturally occurring aglain congener all of them isolated from three Aglaia species (Aglaia duperreana, A. oligophylla and A. spectabilis) collected in Vietnam were studied for their antiproliferative effects using the human monocytic leukemia cell lines MONO-MAC-1 and MONO-MAC-6. Only rocaglamide type compounds showed significant inhibition of [ 3 H-]thymidine incorporation and the most active compound didesmethylrocaglamide inhibited cell growth in a similar concentration range as the well-known anticancer drug vinblastine sulfate. Detailed structure-activity analysis indicated that the OH-group at C-8b which is a common structural feature of most naturally occurring rocaglamide compounds is essential for the described antiproliferative activity since replacement of this group by methylation led to a complete loss of the inhibitory activity for the resulting derivative. Rocaglamide derivatives rapidly inhibited DNA as well as protein biosynthesis of MONOMAC- 6 cells at concentrations well below those of actinomycin D or cycloheximide which were used as positive controls in the respective experiments. Didesmethylrocaglamide was furthermore able to induce growth arrest of MONO-MAC-1 cells in the G2/M and probably G0/Gl-phase of the cell cycle with no morphological indication of cellular damage. Our data suggests that 1H-cyclopenta[b]benzofuran lignans of the rocaglamide type act primarily by a cytostatic mechanism.

Protective capabilities were studied of carboxymethylated (1→3)-β-ᴅ-glucan from Saccharomyces cerevisiae cell wall against lipid peroxidation in phosphatidylcholine liposomes in duced by OH· radicals produced with Fenton’s reagent (H 2 O 2 /Fe 2+ ) and also by microwave radiation using absorption UV-VIS spectrophotometry. A significant decrease in the conjugated diene production, quantified as Klein oxidation index, was observed in the presence of a moderate amount of added glucan. Increase of the oxidation index was accompanied with enhanced carboxyfluorescein leakage as a result of liposome membrane destabilization. This process was markedly suppressed with glucan present in the liposome suspension. Therefore, glucan may be considered as a potent protector against microwave radiation-induced cell damage.

The plasma membrane has gained increasing attention as a possible target of antitumor drugs. It has been reported that they act as growth factor antagonists, growth factor receptor blockers, interfere with mitogenic signal transduction or exert direct cytotoxic effects. Chlorambucil (4-[p-(bis[2-chloroethyl]amino)phenyl]butyric acid) is an alkylating agent widely used in the treatment of chronic lymphocytic leukaemia. Contradictory reports have been published concerning its interaction with cell membranes. Whereas a decrease in the fluidity of Ehrlich ascite tumor cells has been adduced, no evidences were found that chlorambucil changes membrane lipid fluidity and alkylating agents had effects in these systems even at highly toxic concentrations. Our results showed that chlorambucil at a dose equivalent to its therapeutical concentration in the plasma (3.6 μm) caused the human erythrocyte membrane to develop cup-shaped forms (stomatocytes). Accordingly to the bilayer couple hypothesis, this means that the drug is inserted into the inner monolayer of the erythrocyte membrane, a conclusion supported by X-ray diffraction performed on multilayers of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylethanolamine (DMPE), representative of phospholipid classes located in the outer and inner monolayers of the erythrocyte membrane, respectively. It is concluded that the cytotoxic effect of chlorambucil might be due to alteration of the structure and therefore of the physiological properties of cell membranes such as fluidity, permeability, receptor and channel functions.

The platelet-derived endothelial cell growth factor (PD-ECGF) is one of the potent angiogenic factors. Recently, its homology with thymidine phosphorylase (dThdPase), an enzyme involved in pyrimidine nucleoside metabolism, has been shown. In the present study, dThdPase activity was evaluated spectrophotometrically in 43 breast carcinomas and in 19 cases of non-neoplastic breast tissues. The mean dThdPase activity in breast cancer was almost six fold higher than in normal, non-neoplastic breast tissues (1.92 and 0.29 μmol thymine (T) x mg prot. -1 x h -1 respectively). The enzyme activity significantly correlated with axillary lymph node status (p = 0.0076) and with tumor size (p = 0.0099). Besides, the intratumoral microvessel density (MD) was evaluated using the CD 31 mouse anti-human monoclonal antibody, and there was no correlation between the level of enzymatic activity and a number of microvessels. The positive significant correlation of thymidine phosphorylase activity with prognostic factors in breast cancer patients with no relation to the number of microvessels needs further examination to confirm the prognostic significance of the level of dThdPase.

The activities of mitochondrial malate dehydrogenase (mMDH ) and the total mitochondrial proteins increase as a function of body mass in the freshwater catfish, Clarias batrachus. It clearly indicates an increase in energy production in larger- sized individuals for various purposes including prey-predator interactions. The higher activity of lactate dehydrogenase (LDH) in larger fish may indicate more production of lactate for gluconeogenesis in the liver to meet emergency requirements of increased energy demand. However, the activity of cytoplasmic malate dehydrogenase (cMDH) decreases with the increasing body mass of the fish which reflects reduction in NADPH production and, in turn, reduced lipogenesis in liver of larger individuals. Thus, the present observations suggest an adaptive mechanism dealing with the higher energy budget, and reduced synthetic activities (lipogenesis) in the liver of larger-sized freshwater catfish. This type of biochemical scaling might be also supporting other metabolic pathways in order to adjust some physiological functions for survival in the aquatic environment.

Many hypotheses have been developed to explain aging and age-related neurodegenerative disorders; one of the most compelling is the role of oxidative stress to induce changes in protease activity in brains of patients of Alzheimer’s disease and prion disease. At the moment however, there is no clear answer how protein degradation may be achieved in the brain. We have observed that several metal compounds can degrade proteins in the presence of hydrogen peroxide, and elucidated the reaction scheme based on the new theoretical point for the reactivity of a metal-peroxide adduct with η 1 -coordination mode. In this article we would like to point out the importance of a copper(II)-peroxide adduct to promote neurodegenerative diseases such as prion disease and amyotrophic lateral sclerosis through its oxidative protease function.

A series of high molecular mass pentacyclic terpenoid wax esters (TTFAE) were characterized for the first time in hexane extracts of Brazilian propolis (Carangola, Minas Gerais State). Identification was by HT-HRGC (High Temperature High Resolution Gas Chromatography) and HT-HRGC coupled to mass spectrometry (HT-HRGC-MS).

Males of several palaearctic Syntomis/Amata species (Lepidoptera: Arctiidae) possess androconial hairbrushes in connection with the foreleg coxa. The cuticular structure of these potentially behaviour-related and pheromone dissipating brushes is described. Such malespecific organs and signals play a crucial role in the female choice procedure. The presence of hairbrushes was found in 17 out of 28 inspected species of the tribe Syntomini. All members of the Syntomis phegea group (Europe to Central Asia, as well as Caspian, Caucasian and near-middle East species) have these structures, and only three oriental and south Asian, but none of three African species, carry this trait. The common genetic base of this morphological character is supported by an analysis of mitochondrial 16S rRNA from 19 representative taxa; species with hairbrushes form a monophyletic clade and the brushes are a synapomorphic character. This genetic finding corroborates the ethological significance of these organs. Phylogenetic data show a substantial genetic divergence between the tribe Ctenuchini (New World species) and the Old World Syntomini. Furthermore, DNA sequence data suggest a split of the genus Amata (sensu Obraztsov, 1966) in two distinct genera, Amata (without hairbrushes) and Syntomis (with hairbrushes).

The flavonoids 5-hydroxy-7-methoxyflavanone, 6,8 -dimethyl- 5,7-dihydroxyflavanone and 2′,4′-dihydroxy-6′- methoxy-3′,5′-dimethylchalcone, a mixture of alkyl esters of p-coumaric acid, the triterpenoids oleanolic acid and maslinic acid, the monoterpenoid 1α,2β,4β-trihydroxy- p-menthane, the sesquiterpenoid clovandiol and β-sitosterol were isolated from the aerial parts of Luma gayana (Barn.) Burret. This is the first report on the chemistry of this species.

Sex pheromones, released with the coelomic fluid by male Platynereis dumerilii initiate egg release in swarming females. The egg release pheromone, isolated from the coelomic fluid of sexually mature males, was identified as ʟ-Ovothiol A, which was found in male marine invertebrates for the first time. Isolation was obtained by reversed-phase high-performance liquid chromatography in the biologically inactive disulfide form.