Mahendra Pandurang Darokar

Synergistic effect of silymarin and standardized extract of Phyllanthus amarus against CCl4-induced hepatotoxicity in Rattus norvegicus

In search of the effective and standardized hepatoprotective combination therapy, silymarin and standardized extract of Phyllanthus amarus has been evaluated against CCl(4)-induced hepatotoxicity in rats. Eight groups of rats were used. The animals of group A served as normal and were given only vehicle. The animals of group B served as toxin control and were administered with CCl(4) (50% solution of CCl(4) in liquid paraffin, 2 ml/kg b.w., intraperitoneally). The animals of groups C-H received silymarin (100 mg/kg b.w.), Phyllanthus amarus aqueous extract (100 mg/kg b.w.), Phyllanthus amarus ethanolic extract (100 mg/kg b.w.), silymarin (50 mg/kg b.w.)+P. amarus aq. ext. (50 mg/kg b.w.), silymarin (50 mg/kg b.w.)+P. amarus eth. ext. (50 mg/kg b.w.) and marketed formulation (M.F.) 5 ml/kg b.w. for 6 days orally as well as CCl(4) (2 ml/kg b.w.) on 4th day intraperitoneally. The test materials were found effective as hepatoprotective as evidenced by plasma and liver biochemical parameters. The combination of silymarin and Phyllanthus amarus showed synergistic effect for hepatoprotection and silymarin with ethanolic extract of P. amarus showed better activity due to the higher concentration of phyllanthin in ethanolic extract in comparison to aqueous extract of P. amarus as estimated by HPLC.

Antiproliferative and antioxidant activities of Juglans regia fruit extracts

Context: Cancer chemopreventive action of walnut [Juglans regia L. (Juglandaceae)] has been explored. Objective: This study evaluated antiproliferative and antioxidant activities of walnut. Materials and methods: Various fractions of walnut extract have been screened for antiproliferative activity against human cancer cell lines using the MTT assay. All these fractions have also been evaluated for total phenolic content, antioxidant activity, and reducing power capacity. Results and discussion: Chloroform and ethyl acetate fractions exhibited a high level of antiproliferation against HepG-2, liver cancer cell line (IC50 = 9 and 15 µg/mL, respectively). Conclusion: Exhibiting high phenolic content, antioxidant activity, and potent antiproliferative activity, walnut may act as a cancer chemopreventive agent.

A promising anticancer and antimalarial component from the leaves of Bidens pilosa.

Bidens pilosa is used in folk medicine for various applications due to the presence of polyacetylenes, flavonoids, terpenoids, phenylpropanoids and others. Bioactivity-guided fractionation of different extracts of B. pilosa leaf showed potential in vitro anticancer and antimalarial activity and led to the identification of a potential marker compound, phenyl-1,3,5-heptatriyne. Erythrocyte osmotic fragility experiments revealed the various extracts as well as the marker components toxicity profiles on normal blood cells.

Glabridin induces oxidative stress mediated apoptosis like cell death of malaria parasite Plasmodium falciparum.

Plants are known as the source of novel agents for developing new antimalarial drugs. Glabridin is a polyphenolic flavonoid, a main constituent in the roots of Glycyrrhiza glabra possesses various biological activities. However, its anti-plasmodial activity is unexplored. In the present work, it is for the first time demonstrated that glabridin inhibits Plasmodium falciparum growth in vitro with an IC50 23.9±0.43μM. Glabridin showed poor cytotoxicity in vitro with an IC50 246.6±0.88μM against Vero cell line and good selectivity index (9.6). In erythrocytic cycle, trophozoite stage was found to be most sensitive to glabridin. In silico study showed that glabridin inhibits Pf LDH enzyme activity by acting on NADH binding site. Glabridin induced oxidative stress by the generation of reactive oxygen and nitrogen species. Glabridin could induce apoptosis in parasite as evidenced by the depolarization of mitochondrial membrane potential (Δψm), activation of caspase like proteases and DNA fragmentation. These results indicate that glabridin exhibits antiplasmodial activity and is suitable for developing antimalarial agent from a cheap and sustainable source.

Low levels of genetic diversity detected by RAPD analysis in geographically distinct accessions of Bacopa monnieri

Brahmi (Bacopa monnieri) is atraditional Indian medicinal plant known for its natural nootropicaction of saponins present in large amount in its shoots. Acollection of 24 B.monnieri accessions from differentagro-climatic zones of India and an introduction from Malaysiamaintained in the field genebank at CIMAP was analysed for RAPDvariation. Among the 40 random primers tested, 29 primers generatedone or more polymorphic bands. The number of polymorphic bandsgenerated was primer dependent, ranging from 2 to maximum of 8.Similarity matrices were generated from the RAPD data on the basis ofNeis estimates of similarity indices and dendrograms wereconstructed based on UPGMA clustering. All the accessions were foundto be in the range of 0.8–1.0 of similarity, which isindicative of a narrow genetic base among the various accessions witha medium level of polymorphism. It was possible to differentiateindividual accessions, showing differences in morphological andgrowth properties at DNA level. The observed low levels of geneticvariation were attributed to interplay of sexual and vegetative modesof reproduction and similarity of local environments in habitats ofB. monnieri.

Molecular Profiling for Genetic Variability in Capsicum Species Based on ISSR and RAPD Markers

The taxonomic identity of Capsicum species is found to be difficult as it displays variations at morpho-chemical characters. Twenty-two accessions of six Capsicum species, namely, C. annuum, C. baccatum, C. chinense, C. eximium, C. frutescens, and C. luteum were investigated for phenotypic diversity based on flower color and for genetic differences by molecular makers. The genetic cluster analyses of 27 RAPD and eight ISSR primers, respectively, revealed genetic similarities in the ranges of 23–88% and 11–96%. Principal component analysis of the pooled RAPD and ISSR data further supports the genetic similarity and groupings. Different species showed variations in relation to corolla shade of flower. C. annuum accessions formed a single cluster in the molecular analysis as maintaining their flower characteristic. C. chinense accession shared flower features with the accessions of C. frutescens and were found to be closer at genotypic level. C. luteum was found to be rather closer to C. baccatum complex, both phenotypically and genetically. The only accession of C. eximium presenting purple flowers falls apart from the groupings. The floral characteristics and the molecular markers are found to be useful toward the delineation of the species specificity in Capsicum collection and identification of genetic stock.

Enhancement of artemisinin content through four cycles of recurrent selection with relation to heritability, correlation and molecular marker in Artemisia annua L.

Due to the high demand and low yield of the anti-malarial drug artemisinin in natural populations of Artemisia annua (Quinghao), an attempt has been made to enhance the artemisinin content through 4 cycles of recurrent selection (C(0)-C(3)) using selected genotypic and phenotypic traits. Based on their phenotypic and genotypic characteristics, the top 5% plants of each cycle were selected, and their seedlings were planted in poly-cross block to produce seeds for the subsequent generation. A significant increase in the artemisinin content (0.15% in C (0) to 1.16% in C (3), i.e., about 40% genetic gain over the generation) was observed. This enhancement was directly correlated with the plant height and branching intensity in all four cycles. Similarly, the PCV (phenotypic coefficient of variation) and GCV (genotypic coefficient of variation) have been observed to have a higher value for artemisinin content. The DNA marker (MAP 12) with relation to artemisinin was also identified for high yielding genotypes in all four cycles of selection. Over the four cycles of recurrent selection, the plant developed an oval appearance (Variety: CIM-Arogya) and a high artemisinin content (1.16%).

Antimycobacterial Activity of Lichens

Abstract Ethanol extracts of nine lichen species, namely Everniastrum cirrhatum. (Fr.) Hale ex Sipman (Parmeliaceae), Flavoparmelia caperata. (L) Hale (Parmeliaceae), Heterodermia leucomela. (L) Poelt (Physciaceae), Lecanora flavidorufa. Hue (Lecanoraceae), Leptogium pedicellatum. P.M. Jorg (Collemataceae), Lobaria isidiosa. (Bory) Trevisan (Stictaceae), Rimelia reticulata. (Taylor) Hale and Fletcher (Parmeliaceae), Phaeophyscia hispidula. (Ach.) Essl (Physciaceae), and Stereocaulon foliolosum. Nyl. (Stereocaulaceae), were evaluated for antimycobacteral properties against Mycobacterium tuberculosis. H37Rv and H37Ra strains using the radiometric BACTEC method. Among the tested lichens, the virulent strain of M. tuberculosis. H37Rv was found more susceptible to ethanol extract of F. caperata. and H. leucomela. (MIC 250 µg/mL). E. cirrhatum., R. reticulata., and S. foliolosum. were found active at the concentration of 500 µg/mL. L. isidiosa., L. pedicellatum., P. hispidula., and L. flavidorufa. did not exhibit activity at the maximum tested concentration of 1000 µg/mL.

Drug Resistance Reversal Potential of Ursolic Acid Derivatives against Nalidixic Acid- and Multidrug-resistant Escherichia coli.

As a part of our drug discovery program, ursolic acid was chemically transformed into six semi‐synthetic derivatives, which were evaluated for their antibacterial and drug resistance reversal potential in combination with conventional antibiotic nalidixic acid against the nalidixic acid‐sensitive and nalidixic acid‐resistant strains of Escherichia coli. Although ursolic acid and its all semi‐synthetic derivatives did not show antibacterial activity of their own, but in combination, they significantly reduced the minimum inhibitory concentration of nalidixic acid up to eightfold. The 3‐O‐acetyl‐urs‐12‐en‐28‐isopropyl ester (UA‐4) and 3‐O‐acetyl‐urs‐12‐en‐28‐n‐butyl ester (UA‐5) derivatives of ursolic acid reduced the minimum inhibitory concentration of nalidixic acid by eightfold against nalidixic acid‐resistant and four and eightfold against nalidixic acid‐sensitive, respectively. The UA‐4 and UA‐5 were further evaluated for their synergy potential with another antibiotic tetracycline against the multidrug‐resistant clinical isolate of Escherichia coli‐KG4. The results showed that both these derivatives in combination with tetracycline reduced the cell viability in concentration‐dependent manner by significantly inhibiting efflux pump. This was further supported by the in silico binding affinity of UA‐4 and UA‐5 with efflux pump proteins. These ursolic acid derivatives may find their potential use as synergistic agents in the treatment of multidrug‐resistant Gram‐negative infections.

Epoxide group relationship with cytotoxicity in withanolide derivatives from Withania somnifera.

Withania somnifera is one of the highly reputed medicinal plants of India. Its steroidal constituents exist in the form of two major substitution patterns, viz. withaferin A (1) and withanone (5). Withaferin A with oxidation at carbons 4, 5, and 6 is considered as an active type, especially as anticancer, whereas the withanones with oxidation at carbons 5, 6, and 7 rarely show any activity. We prepared a series of derivatives with modifications at carbons 5, 6, and 7 in ring B of these withanolides to study the role of the epoxide group towards the cytotoxic property of these bioactive steroids. We have converted withanolides into the respective thiiranes, amino alcohols and alcohols by selective reactions at the epoxide ring and were evaluated for in vitro anticancer activity against four cancer cell lines to study the structure activity relationships. The transformations of the epoxide group in withanolides of the withaferin A type showed moderate reduction in their cytotoxicity whereas the almost inactive withanones have shown some improvements in their alcohol derivatives.