Evaluation of in vitro antioxidant, anti-inflammatory, anticoagulant and antiplatelet potential of Rhus mysorensis

Since ancient times human beings are using plant-based medicines for the treatment of various ailments, especially in the rural areas, due to their availability and affordability. Rhus mysorensis (RM) is widely used as a traditional medicine to treat various ailments. Owing to its potential medicinal value, the present study was designed to explore the in vitro antioxidant, anti-inflammatory, anticoagulant and antiplatelet properties of purified column fraction of RM. The methanol extract of stem bark powder was sequentially fractioned by solvent partitioning. The liquid methanol fraction was further fractionated by column chromatography using gradient elution. Eluted fractions were analyzed using HPLC for percentage purity and yield. The fraction with higher percentage of purity and yield was assessed for in vitro antioxidant activity by measuring SOD and GPx activities, anti-inflammatory activity by the inhibition of nitric oxide (NO) production in LPS induced RAW264.7 cells, anticoagulation by plasma recalcification time and antiplatelet activity by agonists induced platelet aggregation respectively. The antioxidant potency of column fraction (B8) revealed that, highest enzyme activities were recorded at a concentration of 320µg/ml. The enzyme activity was found to be 2.45 U/ml for SOD and 135.75 U/L for GPx respectively. Purified column fraction (B8) of RM significantly reduces the production of NO in LPS stimulated RAW 264.7 cell lines at 320????g/ml concentration with 31.90% of inhibition. The anticoagulant activity of purified fraction was determined in terms of plasma recalcification time. Interestingly, the fraction showed the most potent anticoagulant activity both in PRP and PPP as it prolonged the clotting time. The findings indicate that the stem bark of RM possesses potent antioxidant, anti-inflammatory, anticoagulant and antiplatelet activities, supporting the use of this species for treating oxidative stress-induced inflammatory diseases. Further, bioactivity guided fractionation studies to characterize and identify specific phytochemicals responsible for these biological activities are needed.

ISOLATION AND CHARACTERISATION OF (17E)-CYCLOART-17, 26-DIEN-3β-OL AND CYCLOART-3β, 25-DIOL FROM THE PEEL OF ARTOCARPUS HETEROPHYLLUS L.

Two triterpenoids namely (17E)-cycloart-17,26-dien-3β-ol and cycloart-3β,25-diol, were isolated as a mixture from the column fraction by elution with n-hexane/30% ethyl acetate of methanol extract of ripe jackfruit (Artocarpus heterophylus) peel. This is the first report of their isolation from this plant. Their structures were elucidated by comparing the nuclear magnetic resonance (NMR) data with those published for these compounds. The antioxidant activity of these isolated compounds was determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and showed a low scavenging attitude compared with standard phenolic compounds (2-tert- butyl-4-methoxyphenol (BHA), Trolox, L(+)-ascorbic acid and gallic acid). Extracts and isolated compounds did not exhibit any antibacterial activity against two Gram-positive and two Gram-negative bacteria.

In silico, in vitro: antioxidant and antihepatotoxic activity of gnetol from Gnetum ula Brongn

Introduction: Gnetum ula is a notable medicinal plant used to cure various ailments. The stem part of the plant is used traditionally to treat jaundice and other disorders. The present work is to investigate the in vitro hepatoprotective and antioxidant activity of ethanol extract of stem of G. ula (GUE) and its isolated compound gnetol. Methods: Column chromatography was carried out for GUE and various column fractions were obtained. DPPH and reducing power assays were performed for GUE and column fractions. The potent fraction was characterized, interpreted and tested for in vitro hepatoprotective activity on the BRL3A cell line. In silico docking studies of gnetol compound on the protein TGF-β (transforming growth factor – β) and Peroxisome proliferator-activated receptor α (PPARα) was carried out. Results: DPPH scavenging and reducing power assay showed that the fourth column fraction has antioxidant potential than other fractions. The fourth column fraction was characterized to obtain gnetol compound. BRL3A cell line was used for the toxicity study of GUE and gnetol. Both, the extract and the isolated compound were found to be nontoxic with CTC50 value more than 1000 µg/mL. At the concentration of 200 µg/mL, GUE and gnetol offered cell protection of 50.2% and 54.3%, however, silymarin showed 77.15% protection at 200 µg/mL concentration against CCl4 treated BRL3A cell line. The docking results of the ligand molecule TGF-β showed that gnetol has the binding affinity of -7.0 and standard silymarin being -6.8. TGF-β showed good hydrophobic interactions and formed two hydrogen bonds with the amino acids. For PPARα protein, gnetol showed the binding affinity of -8.4 and silymarin with -6.5. Hydrogen bonding and good hydrophobic interactions against the amino acid molecules in relation to the PPARα protein are shown. Conclusion: Gnetum ula stem extract and its isolated compound are safe and offered significant hepatoprotection against CCl4 induced toxicity. Isolated compound gnetol exhibited a potent antioxidant activity offering protection to liver damage. However, in vivo studies need to be carried out to validate the traditional use of G. ula.

Evaluation of antimicrobial activity and phytochemical composition of some fractions of methanol stem bark extract of Diospyros mespiliformis

The aim of this study was to determine the active phytochemical(s) most probably responsible for microbial inhibitions, following a bioassay guided protocol. Column chromatographic fractions (AG) obtained from n-Butanol partitioned portion of stem bark extract of Diospyros mespiliformis were analyzed for phytochemical composition. These were subjected to antimicrobial activity tests on clinical isolates of Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa, Klebsiella pneumoniae and Candida albicans. Phytochemical screening conducted on the column fraction E revealed the presence of tannins, free anthraquinones, cardiac glycosides, saponins, terpenoids, and flavonoids. The anti-microbial test results from fractions A, B, F and G, obtained showed no inhibition against most of the micro-organisms tested, while fractions C, D and E showed significant (P<0.05) activities with diameters of inhibition zone of inhibition ranging from 15.00±1.00 mm to 13.00±6.67 mm against Streptococcus pyogenes at all the concentrations tested (50, 12.5, 6.25 mg/ml equivalent to 3, 1.5 and 0.75 mg/disc). The column fraction E showed the highest inhibition zones with broader concentration dependent pattern with MIC at 12.5 mg/ml. The findings from this study showed the presence of bio-active components against Streptococcus pyogenes with relative higher potency in fraction E. Based on this findings, it can be surmised that Fraction E with significant dose-dependent activity is expected to revealed a potent broad-spectrum antimicrobial agent and thus recommended for further purification stages towards compound(s) isolation of a novel antimicrobial agent.Keywords: Antimicrobial, Diospyros mespiliformis, Phytochemical, Potency, Stem Bark

3β-Hydroxy-Δ5-steroidal congeners from a column fraction of Dendronephthya puetteri attenuate LPS-induced inflammatory responses in RAW 264.7 macrophages and zebrafish embryo model

Bioactive compounds from marine organisms and their action mechanisms have provided new insights into medicinal and natural product research.

Toxicity of Vernonia anthelmintica Linn. (Asteracea) seeds against mosquitoes vectors

The Toxicological activity (larvicidal, adulticidal and repellent toxicity) of Vernonia anthelmintica seeds fraction was tested against different species of mosquito vectors viz, malaria (Anopheles culicifacies and Anopheles stephensi), filaria (Culex quinquefasciatus) and dengue (Aedes aegypti). The larvicidal toxicity of Vernonia anthelmintica seeds fraction was evaluated against the early 4th instars larvae of different mosquitoes species. Mean LC50 value of the column fraction KAL-4 from seeds of V. anthelmintica against the larvae of An. culicifacies, An. stephensi, Culex quinquifaciatus and Aedes aegpyti were found to be 64 ppm, 70 ppm, 143 ppm and 166 ppm respectively. The larvicidal toxicity was more against An. culicifacies, An. stephensi than Culex quinquifaciatus and Aedes aegypti. The seed extracts did not show any adulticidal toxicity and repellent toxicity even at 10% concentrated impregnated paper and 5% on human hand, respectively.

Bioactivity of the red algae Asparagopsis taxiformis collected from the Southwestern coast of India

Among the diverse variety of red algae, Asparagopsis taxiformis constitutes one of the abundant biomass in the Kollam coast (Southwest coast of India). Therefore, in the present study, A. taxiformis was collected, extracted and fractionated using column chromatography. The individual fractions were evaluated in vitro for their antifouling, anticyanobacterial, piscicidal and crustaceans toxicity assays. The fraction eluted with 2:8, petroleum ether and ethyl acetate exhibited strong and broad spectrum of bioactivity. In antifouling assay against Limnea truncatula, the active algal fraction produced 80% of foot repellency at 150 mg/L whereas in anticyanobacterial assay, the active fraction inhibited 100% growth of Trichodesmium sp. at 320 mg/L. The algal fraction showed higher piscicidal effect at the level of 60 mg/L. The crustacean toxicity of the active fraction was also evaluated to find compounds without toxicity in non target organisms, Penaeus monodon and Macrobrachium rosenbergii. It was found that column fraction showed less toxicity against the non target organisms. The chemical constituents of the active fraction were identified by means of chromatographic systems such as TLC, reverse phase HPLC and GC-MS. The overall activity profile envisages that the active column fraction of A. taxiformis might contain synergistic bioactive metabolites that could be utilized for the control of fouling organisms, algal bloom and herbivorous/predaceous fishes in aquaculture ponds.

Role of factors downstream of caspases in nuclear disassembly during apoptotic execution

We used cytoplasmic extracts from chicken DU249 cells at various stages along the apoptotic pathway to analyse the events of apoptotic exe–cution. So–called S/M extracts from morphologically normal ‘committed–stage’ cells induce apoptotic morphology and DNA cleavage in substrate nuclei. These apoptotic changes appear to require the function of multiple caspases (cysteine aspar–tases, a specialized class of proteases) acting in parallel. Extracts from ‘execution–stage’ apoptotic cells induce apoptotic events in added nuclei in a caspase–independent manner. Biochemical frac–tionation of these extracts reveals that a column fraction enriched in endogenous active caspases is un–able to induce DNA fragmentation or chromatin condensation in substrate nuclei, whereas a caspase–depleted fraction induces both changes. ‘Execution–stage’ extracts contain an ICAD/DFF45–inhibitable nuclease resembling CAD, plus another activity that is required for the apoptotic chromatin condensation. ‘Committed–stage’ S/M extracts lack these downstream activities. These observations reveal that caspases act in an executive fashion, serving to activate downstream factors that disassemble the nucleus rather than disassembling it themselves. They also suggest that activation of the downstream factors (rather than the caspases) is the critical event that occurs at the transition from the latent to the execution phase of apoptosis.