Band 65, Heft 5-6

The N-(benzoylphenyl)-1H-indole-2-carboxamide derivatives 1 - 6 were synthesized, and the lipid-lowering effects of two of these novel compounds were studied using hyperlipidemic rats as an experimental model. Treatment of ethyl-1H-indole-2-carboxylate with aminobenzophenones in the presence of sodium ethoxide and DMF, followed by purifi cation using column chromatography, gave the target compounds in good yields. The tested animals were divided into control, hyperlipidemic, compounds 2-, 3- and bezafibrate-treated groups. At a dose of 15 mg/kg body weight, compounds 2, 3 and bezafibrate significantly reduced the elevated plasma triglyceride levels after 7 and 24 h. Furthermore, the high-density lipoprotein- cholesterol levels were remarkably increased in all treated groups after 7 and 24 h compared to the hyperlipidemic control group. However, only compounds 2- and 3-treated groups obviously showed a significant reduction in plasma total cholesterol levels after 24 h. It is therefore reasonable to assume that 2 and 3 may have a promising potential in the treatment of hyperlipidemia and coronary heart diseases.

The chemical structure of a polysaccharide from the broth of cultured Streptomyces virginia H03 was investigated. There might be 1→2 and 1→4 and no 1→6 glycosidic linkages in the polysaccharide according to periodate oxidation and Smith degradation. Four fragments including 2,3,4,6-Me 4 -Man, 2,3,6-Me 3 -Gal, 2,4,6-Me 3 -Glc, and 3,6-Me 2 -Man were found in the methylated polysaccharide. Furthermore, the polysaccharide has a β-Glc(1→4)- α-Man(1→4)-α-Gal(1→3)-linked backbone and a branch at the C-2 position of (1→2)-linked mannose residues as determined by the method of nuclear magnetic resonance (NMR) spectroscopy. The assumed structure of the polysaccharide is [→3)-β-Glc(1→4)-α-Man (1→4)-α-Gal(1→] n . 2 ↑ α-Man(1 Additionally, the polysaccharide has a wide antibiogram and was found to be most effective against Bacillus subtilis, Staphylococcus aureus, Listeria monocytogenes, and Escherichia coli.

A new trimeric proanthocyanidin, epigallocatechin-3-O-gallat-(4β→8)-epigallocatechin- (4β→8)-catechin (1), was isolated together with three known flavan-3-ols, catechin (2), epicatechin (3), and epigallocatechin (4), and three dimeric proanthocyanidins, 5 - 7, from the air-dried leaves of Mangifera indica. Their chemical structures were determined on the basis of 1D- and 2D-NMR spectra (HSQC, HMBC) of their peracetylated derivatives, MALDITOF- mass spectra, and by acid-catalyzed degradation with phloroglucinol. The isolated compounds 1 - 7 were in vitro tested for their inhibitory activities against COX-1 and COX-2. Compound 1 was found to have a potent inhibitory effect on COX-2, while compounds 1 and 5 - 7 exhibited moderate inhibition against COX-1.

The compound crotocaudin extracted from the stem bark of the medicinal plant Croton tiglium Linn. was administered for 24 h or 96 h to the freshwater vector snail Lymnaea (Radix) acuminata Lamarck in order to test its toxicity. L. acuminata is the intermediate host of Fasciola hepatica and Fasciola gigantica which cause immense harm to man and his domestic animals. It was observed that the molluscicidal activity of crotocaudin against L. acuminata is time- as well as dose-dependent. There was a significant negative correlation among LC 50 values and exposure periods, i.e. increasing the exposure time, the LC50 value of crotocaudin decreased from 5.37 μM (24 h) > 2.08 μM (48 h) > 1.36 μM (72 h) to 1.01 μM (96 h), respectively, against L. acuminata. The toxicological experiments to proof for environmental toxicity, if any, have also been carried out on the non-target freshwater fish Channa punctatus (Bloch) [Channidae (Ophicephalidae)], which shares the habitat with L. acuminata. The sublethal doses of crotocaudin (40% and 80% of LC50) administered over 24 h caused significant changes in the carbohydrate and nitrogenous metabolisms in nervous, hepatopancreas, and ovotestis tissues of Lymnaea acuminata. Channa punctatus was also exposed to sublethal doses of crotocaudin (40% and 80% of 24-h LC 50 of L. acuminata) for 96 h which showed significant alterations in the metabolism in muscle, liver, and gonad tissues. After withdrawal of crotocaudin the snail tissues recovered in part after 7 days and the fish tissues completely.

Silymarin, a purified extract of seeds of Silybum marianum L. and well known for its hepatoprotective abilities, has been evaluated for inherent utility as a radioprotective agent. A fraction (INM-7035) was authenticated by characterizing the percentage composition of silybin A and B (39.9% and 57.4%). Free radical scavenging activities of INM-7035 against superoxide radicals (>68%), hydroxyl radicals (>33.75%), DPPH (67.2%), and ABTS (32.4%) were also evaluated. The fraction chelated (>30%) ferrous ions, thereby able to restrict amplification. INM-7035 exhibited >50% peroxyl radical scavenging activity in the lipid phase along with dose-dependent (R 2 = 0.990) reducing power in the aqueous phase. Radiation-induced free radical flux can lead to disruption of biomolecules like membrane lipids. INM-7035 completely inhibited lipid peroxidative stress in case of membranes against supralethal radiation stress in the liposomal system. The ability of INM-7035 to modulate the levels of NF-κB, indicated its inherent potential as a radioprotective bioactive constituent.

Current evidences demonstrated that the activity of protein kinases can be controlled through oxidative stress induced by reactive oxygen species (ROS) and normalized by antioxidants. Recent studies with ROS, generated by mitochondria, suggested the potential signalling role of these species, where ROS, especially hydrogen peroxide, were proposed as membrane-related signalling components. The protein regulation by cellular redox states has shown that protein tyrosine kinase members, such as Src kinase and some of the members of the Src family kinases (SFKs), are proteins regulated by the cellular oxidation and reduction status. In this context, the oxidant or antioxidant potential of the synthetic Src kinase inhibitors previously synthesized and studied by our research group, such as N-substituted indole-3-imine and -amine derivatives, were investigated employing various acellular in vitro methods including microsomal NADPH-dependent inhibition of lipid peroxidation (LP), interaction of 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical and scavenging of superoxide anion radicals. Here, we report that some of the synthetic inhibitors designed for Src kinase target have both antioxidant and kinase inhibition properties.

To date, two types of ribosome-inactivating proteins (RIPs) have been found in Jatropha curcas. One is curcin, which has been isolated from the endosperm, and the other is curcin-L, which is expressed in leaves upon stress treatment. Phylogenetic analysis of the predicted amino acid sequences of the RIPs in plants revealed that these belong to a major subfamily and are close to trichosanthin (TCS). Studies on the mRNA and protein levels showed that both curcin and curcin-L have an organ-specific expression pattern. Curcin is only expressed and accumulated in the endosperm; its expression begins in the globular embryo period and peaks during the mature embryo period. In contrast, curcin-L is only expressed in the leaves, but its expression is induced by certain conditions such as treatment with phytohormones or polyethylene glycol, exposure to high and low temperatures, and fungal infection. Analysis of the 5’ flanking regions of curcin and curcin-L revealed that the 5’ flanking region of curcin-L has three major inserted fragments, which are not present in the corresponding region of curcin. Comparison of characteristic cis-elements suggests the presence of several motifs that are involved in the endosperm-specific expression in the 5’ flanking region of curcin, while in curcin-L some stress- and defense-responsive motifs are found to be mainly located in the three inserted fragments. Comparison of the antifungal activity of the two RIPs showed that the one of curcin-L is higher than that of curcin. Differences in the expression and activity of curcin and curcin-L suggest that these two RIPs have different functions.

The biosynthetic origin of isopentenyl diphosphate in the polyisoprenoid biosynthesis of the rubber-producing plant Eucommia ulmoides Oliver was elucidated for the fi rst time by feeding experiments using 13C-labeled isotopomers of (RS)-mevalonate, 1-deoxy- D -xylulose- 3,4,5-triacetate, 2C-methyl- D -erythritol-1,2,3,4-tetraacetate, and pyruvate. After 13 C-labeled isotopomers were fed to the young seedlings, the polyisoprenoid fractions were prepared and analyzed by 13 C NMR. The NMR data showed that the isoprene units of polyisoprenoid derived from isopentenyl diphosphate, which was biosynthesized using both mevalonate and 1-deoxy- D -xylulose-5-phosphate in E. ulmoides. It is assumed that the cross-talk of isopentenyl diphosphate, derived from both pathways, occurs during the biosynthesis of polyisoprenoid; therefore, it was observed in the formation of low-molecular weight isoprenoids.

Plants are a potential source of a large number of valuable secondary metabolites. In vitro cultures are being considered as an alternative to agricultural processes for studying valuable secondary metabolites. In this way, nutritive factors are important parameters influencing the production of these compounds in plants. Effects of nitrate concentrations (KNO3) on the production of two tropane alkaloids, hyoscyamine and scopolamine, and the growth of aerial parts and roots of two in vitro propagated accessions of Atropa belladonna and hairy roots were investigated. As hairy roots cultures are able to keep a stable production of alkaloids over long periods of subculturing, they are considered as an interesting option for the study of alkaloid biosynthesis. A hairy roots culture of Atropa belladonna was established by transformation with Agrobacterium rhizogenes strain AR15834. The results of our study showed that a rise in KNO 3 concentration caused a decline in hairy roots growth, and had a remarkable effect on the alkaloid content. The alkaloid concentrations obtained in the hairy roots were 3 - 20 times higher than that in the plants at 35 mM of KNO 3 . Increasing the nitrate concentration in the medium of hairy roots also improved the hyoscyamine/scopolamine ratio, while it increased the scopolamine/hyoscyamine ratio in the studied plants.

An electric field may have different effects on plant metabolism depending upon its application style and density, and environmental conditions. The effects of an electric field, low temperature, and their combinations on tissue vitality and some physiological variables regarding antioxidant responses of “bean” (Phaseolus vulgaris L. cv. Gina) and “cole” (Brassica oleracea L. cv. Acephale) leaves were studied. Fifteen-day-old seedlings were exposed to an electric field (100 kV m -1 ) for 10 or 40 min prior to cold treatment. In both plant leaves, cold application caused statistically significant increments in total soluble protein levels and selected antioxidant enzyme activities such as catalase, peroxidase and superoxide dismutase activities. However, tissue vitality and H 2 O 2 levels did not change in “cole”, while tissue vitality decreased and H 2 O 2 levels increased in “bean”. Electric field application itself did not cause any significant changes in “bean” and “cole” leaves. On the other hand, 40 min electric field application increased the deteriorative effect of cold in both plant species, while 10 min electric field augmented the chilling resistance by increasing the tissue vitality and antioxidant enzyme activities resulting in decreased H 2 O 2 levels.

Extraction of mannan oligosaccharides from the yeast cell wall and methods of deproteinization were studied. We extracted crude mannan oligosaccharides by the dilute alkali- Sevage method. The percentages of deproteinization and mannan oligosaccharide loss were compared as indexes using the Sevage method, trichloroacetic acid method, and hydrochloric acid method. The results showed that the hydrochloric acid method exhibited the highest percentage of deproteinization, but only a little higher percentage of mannan oligosaccharide loss than the other two methods.

A method to determine the content of total glutathione (GSHt) was introduced based on high-performance liquid chromatography (HPLC) with dansylation. The minimum detection concentration of GSHt was 0.5 μg/mL and the measurable range 1.0 - 300 μg/mL. GSHt in yeasts was obtained by hot-water extraction, GSH complete autoxidation to oxidized glutathione (GSSG) in alkaline solution and purification by thin-layer chromatography (TLC). The quantitative determination of GSSG was derived by dansyl chloride at pH 9.5, 60 ºC for 60 min and assayed by HPLC. GSHt in Saccharomyces cerevisiae is higher than in Candida rugosa and Candida utilis. S. cerevisiae can be chosen as the better target for mutagenesis and industrial scale.

Polima and Shaan 2A are the two most widely used forms of cytoplasmic male sterility (CMS) in the utilization of heterosis of rapeseed (Brassica napus) in China. A previous study indicated that the mitochondrial gene, orf224, was the only gene with a differential expression pattern among the normal, sterile and fertility-restored lines in rapeseed. DNA sequences of orf224, including coding sequences from Shaan 2A and Polima CMS, were then amplified and analyzed. DNA sequence alignment indicated both the coding sequences were 675 bp in length and had 99.9 and 99% homology in nucleotides and amino acids, respectively, and shared certain similarity to homologues from other Brassica spp. and Arabidopsis thaliana. The probable promoter regions of orf224 were conserved between B. napus and A. thaliana, but the upstream regions of probable promoter regions were completely divergent from each other. Additionally, analysis of the primary and secondary structure of the proteins encoded by orf224 from the two lines predicted that the proteins contain a α-helix, extended strand, and random coil. After cloning a in vitro experiment showed that these two proteins could be expressed in Escherichia coli BL21.

Extracts of female sex pheromone glands of the sand Salix carpenterworm moth, Holcocerus arenicola, a serious pest of desert thicket, were analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). Based on comparison of retention times and mass spectra of synthetic standards, four compounds were identifi ed as cis-7-tetradecen-1-ol (Z7 - 14:OH), cis-5-tetradecen-1-yl acetate (Z5 - 14:OAc), cis-7-tetradecen- 1-yl acetate (Z7 - 14:OAc), and cis-9-hexadecen-1-yl acetate (Z9 - 16:OAc) with the ratio of 24:39:100:43. Electroantennographic (EAG) analyses of these standard chemicals and their analogues showed that Z7 - 14:OAc elicited the largest male EAG response, followed by Z5 - 14:OAc and Z9 - 16:OAc. In fi eld trials, traps baited with either Z7 - 14:OAc or Z5 - 14:OAc captured males while Z7 - 14:OH-, Z9 - 16:OAc- or solvent-baited traps caught no males. Z7 - 14:OAc as a single component was signifi cantly more attractive than Z5 - 14:OAc alone. The combination of Z7 - 14:OAc and Z5 - 14:OAc showed an evidently synergistic effect and attracted much more males than the individual compounds in the field. Addition of Z7 - 14:OH to the blend of Z7 - 14:OAc and Z5 - 14:OAc enhanced slightly the trap catches. We conclude that the major components of the sex pheromone of H. arenicola are Z7 - 14:OAc and Z5 - 14:OAc. Currently, a triangle trap baited with the synthetic compounds Z7 - 14:OAc, Z5 - 14:OAc, and Z7 - 14:OH in a 1:0.4:0.25 ratio at 825 μg/trap dosage can be effectively used to monitor the H. arenicola population level and catch the males within the desert regions in China.

The control of Lepidoptera pests should be carried out before hatching of their caterpillars to avoid damage to the crops. The aim of this work was to assess the activity of neem (trade name: Natuneem®, producer: Base Fértil, Chapadão do Sul, Brazil) and pyroligneous extracts (trade name: Biopirol 7M®, producer: Biocarbo, Itabirito, Brazil) at 10 mL/L (1%) and 20 mL/L (2%) contents on egg masses of different ages of Spodoptera frugiperda Smith (Lepidoptera: Noctuidae) and of Diatraea saccharalis F. (Lepidoptera: Pyralidae) at Embrapa Corn and Sorghum in Sete Lagoas, Minas Gerais State, Brazil. The tests took place in an unbiased casualized design with 12 treatments and four replications. The insecticides were diluted in water, and 0.04 mL of the solution was applied to recently laid and one- and two-day-old eggs of S. frugiperda and D. saccharalis. Caterpillars hatching from recently laid egg masses of S. frugiperda was lower with 2% pyroligneous extract [(0.02 ± 0.00)%]. Recently laid eggs and one- or two-day-old eggs of D. saccharalis presented lower caterpillar hatching with 1% neem extract [(0.00 ± 0.00)%, (0.00 ± 0.00)%, and (1.00 ± 0.01)%] and 2% neem extract [(0.00 ± 0.00)%], compared to 1% pyroligneous extract [(27.30 ± 3.22)%, (28.40 ± 3.32)%, and (37.80 ± 4.14)%] and 2% pyroligneous extract [(42.20 ± 4.49)%, (48.70 ± 4.97)%, and (56.60 ± 5.52)%], respectively. Neem and pyroligneous extracts had impact on hatching of S. frugiperda and D. saccharalis caterpillars.

The combined protective effect of a polyphenol-rich extract, isolated from Geranium sanguineum L. (PC), and a novel naturally glycosylated Cu/Zn-containing superoxide dismutase, produced from the fungal strain Humicula lutea 103 (HL-SOD), in the experimental influenza A virus infection (EIVI) in mice, induced with the virus A/Aichi/2/68 (H3N2), was investigated. The combined application of HL-SOD and PC in doses, which by themselves do not defend significantly mice in EIVI, resulted in a synergistically increased protection, determined on the basis of protective indices and amelioration of lung injury. Lung weights and consolidation as well as infectious lung virus titers were all decreased significantly parallel to the reduction of the mortality rates; lung indices were raised. The excessive production of reactive oxygen species (ROS) by alveolar macrophages (aMØ) as well as the elevated levels of the lung antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), induced by EIVI, were brought to normal. For comparative reasons the combined protective effect of PC and vitamin C was investigated. The obtained results support the combined use of antioxidants for the treatment of influenza virus infection and in general indicate the beneficial protective role of combinations of viral inhibitors of natural origin with diverse modes of action.