Bluemomycin, a new naphthoquinone derivative from Streptomyces sp. with antimicrobial and cytotoxic properties.

Streptomyces is one of the most prolific producers of economically important bioactive compounds used against several illnesses; it has also been found to produce industrially useful enzymes. In this study, Streptomyces sp. (ERINLG-201) was isolated from the soil sample of Kodanad forest (Southern Western Ghats), The Nilgiris, Tamil Nadu, India. ERINLG-201 isolate showed promising antibacterial activity against tested Gram-positive and Gram-negative bacteria which was confirmed by perpendicular 'T' streak method. Secondary metabolites of ERINLG-201 isolate exhibited promising antibacterial activity against tested Gram-positive and Gram-negative bacteria which was confirmed by disc diffusion method using the ethylacetate extract. Further, the ethylacetate extract of ERINLG-201 (15 g) was packed in column chromatography over silica gel and eluted; it resulted in isolation of a new naphthoquinone derivative named bluemomycin from the active fraction. Bluemomycin showed promising antibacterial activity against Gram-negative bacteria and clinical isolates at least concentration (6.25 µg/mL). Cytotoxic studies of bluemomycin showed promising activity against A549, Skvo-3 and HepG2 cell lines with IC50 values of 5.9, 24.2 and 11 µM, respectively.

Enhanced production of fibrinolytic enzyme by a new Xanthomonas oryzae IND3 using low-cost culture medium by response surface methodology.

Cardiovascular diseases (CVDs) cause high mortality throughout the world. Existing fibrinolytic agents are highly expensive and have many side effects. Microbial fibrinolytic enzymes are very much considered as novel therapeutic candidate for the treatment of CVDs. Reports on fibrinolytic enzyme from Xanthomonas sp. is lacking. This study reports fibrinolytic enzymes from Xanthomonas oryzae IND3 as it shows hyperactivity on fibrin-agarose plates. This organism utilized various agro-industrial wastes for enzymes production. Among all, cow dung enhanced more enzyme production, hence it was used as the low-cost substrate for statistical optimization of fibrinolytic protease in Solid state fermentation. Response surface methodology was employed to optimize the factors and enhanced yield by 4-fold. The interactions among the variables, viz, sucrose, yeast extract, and pH of the medium were investigated using Central Composite Design (CCD). The predicted fibrinolytic enzyme activity was 2340 U/g, and the observed fibrinolytic enzyme activity was 2294 ±â€¯12.8 U/g. The fibrinolytic enzyme degraded blood clot in vitro completely. This study is the first report on statistical optimization of fibrinolytic enzyme production in SSF from Xanthomonas sp. The crude extract has immense activity on proteinaceous wastes. The production of fibrinolytic protease using the low-cost substrate could reduce the production cost of enzyme.

Optimizing the fermentation conditions and enhanced production of keratinase from Bacillus cereus isolated from halophilic environment.

Keratinase degrading Bacillus cereus was isolated from the halophilic environment in Tamilnadu, India and keratinase production was optimized using wheat bran substrate. Of the screened bacterial isolates, four were found to have the ability to produce keratinolytic enzyme. The process parameters were optimized using one-variable-at-a-time approach and response surface methodology. Supplementation of 1% lactose supported more keratinase production (120 U/g). Among the selected nitrogen sources, addition of casein significantly enhanced maximum keratinase production (132.5 U/g). Among the ions, manganese chloride significantly enhanced keratinsase production (102.6 U/g), however addition of zinc sulphate and copper sulphate decreased keratinase production. The maximum keratinase production was obtained in the wheat bran medium containing 1% lactose, 0.5% manganese with 80% moisture (292 U/g). Statistics based contour plots were generated to explore the variations in the response surface and to find the relationship between the keratinase yield and the bioprocess conditions.

Hepatoprotective effect of bisbenzylisoquinoline alkaloid tiliamosine from Tiliacora racemosa in high-fat diet/diethylnitrosamine-induced non-alcoholic steatohepatitis.

Non-alcoholic steatohepatitis (NASH) is one of the aggressive forms of non-alcoholic fatty liver disease (NAFLD) and is a potential risk factor of HCC. This study reports the curative effect of tiliamosine on NASH. Tiliamosine was isolated from Tiliacora racemosa Colebr. (Menispermaceae) and its structure was confirmed by studying the physical and spectroscopic data. The effects of tiliamsoine on lipid accumulation and lipotoxicity were evaluated using palmitate-oleate induced steatosis in HepG2 cells. The in vivo efficacy of tiliamosine was evaluated using HFD fed, DEN induced non-alcoholic steatohepatitis Wistar rats. In HepG2 cells, tiliamosine did not affect the cell viability up to 100 µM concentration and showed GI25 value of 264.28 µM. The treatment with tiliamsoine significantly lowered the ORO concentration by 44.17% and triglyceride accumulation by 69.32% at 50 µM concentration (P < 0.005). It also reduced the leakage of LDH and transaminases in PO-BSA induced HepG2 cells. The treatment with tiliamsoine significantly decreased the plasma levels of transaminases, phosphatase and LDH (P < 0.05) in HFD-DEN induced steatohepatitis. The histology and the immunohistochemistry of the hepatic sections were in accordance with the biochemical findings. Preliminary molecular analysis indicated that the hepatic FXR expression was upregulated and TNFα expression was downregulated by the treatment with tiliamsoine. This study provided preliminary evidence on the use of tiliamosine for the treatment of NASH.

Curative effect of arjunolic acid from Terminalia arjuna in non-alcoholic fatty liver disease models.

The prevalence of Non Alcoholic Fatty Liver Disease (NAFLD) is increasing globally. Terminalia arjuna W. & Arn. (Combretaceae) is an endemic tree found in India and Sri Lanka and used traditionally for its cardioprotective and hepatoprotective effects. Arjunolic acid (AA) is an oleanane triterpenoid found mainly in the heartwood of T. arjuna. This study was aimed to evaluate the hepatoprotective effect of AA using cellular and rodent models of NAFLD. AA was isolated from the ethyl acetate extract of the heartwood of T. arjuna. The structure of AA was confirmed by physical and spectroscopic data. Steatosis was induced in HepG2 cells using palmitate-oleate mixture and the effects of AA on triglyceride accumulation and lipotoxicity were assessed. In vivo effect of AA on NAFLD was assessed using HFD fed rats. The treatment with AA did not affect the cell viability upto 100 µM and showed GI25 value of 379.9 µM in HepG2 cells. The treatment with AA significantly lowered the ORO concentration by 35.98% and triglyceride accumulation by 66.36% at 50 µM concentration (P < 0.005) compared to the vehicle treated group. The treatment with AA also reduced the leakage of ALT and AST by 61.11 and 48.29% in a significant manner (P < 0.005). The in vivo findings clearly demonstrated that the animals treated with AA at 25 and 50 mg/kg concentrations showed a significant decrease in the levels of transaminases, phosphatase and GGT (P < 0.005). In the liver, the expression of PPARα and FXRα expressions were upregulated, while PPARγ expression was downregulated by the treatment with AA. The liver histology of the animals showed reduction in steatosis and MNC infiltration. These preliminary evidences suggested that AA might be a promising lead to treat NAFLD. Future robust scientific studies on AA will lead to tailoring it for the treatment of NAFLD.

A study on food-medicine continuum among the non-institutionally trained siddha practitioners of Tiruvallur district, Tamil Nadu, India.

BACKGROUND: Medicinal properties of the food species are one of the poorly documented and important areas of ethnopharmacology. The present survey quantitatively documented the medicinal foods prescribed by the non-institutionally trained siddha practitioners of Tiruvallur district of Tamil Nadu. METHODS: Field work was carried out between December 2014 and April 2017 using a questionnaire. The illnesses mentioned by the informants were grouped as illness categories on the basis of emic perceptions. Sufficiency of sampling of this survey was assessed by plotting the cumulative number of UR and Shannon-Wiener's index. The indices such as informant consensus factor (FIC), Index of Agreement on Remedies (IAR), and Cultural Food Significance Index (CFSI) were calculated. RESULTS: This study documented 165 medicinal foods used by 82 non-institutionally trained siddha practitioners of Tiruvallur district, and 73.93% of these preparations were plant based. Among the animal taxa, 82.05% were represented by fish taxa. The illness category gastrointestinal ailments is the majorly cited illness category treated with plant-based formulations. The illness categories viz., gastrointestinal ailments, hemorrhoids, and neural ailments had high consensus under the group of plant-based medicinal foods. In animal-based medicinal foods, kapha ailments had gained 23.07% of UR. The illness categories such as bone fractures, male reproductive ailments, blood ailments, and anabolic had high FIC values. CONCLUSIONS: Deeper studies on different dietary cultures of India may help to derive better interpretations on food-medicine continuum. This study identified some important claims such as the use of citron, pomegranate and Solanum americanum (gastrointestinal ailments), Abutilon indicum, onions and elephant foot yam (hemorrhoids), Boerhavia diffusa (urinary ailments), Moringa oleifera (anemia), Aloe vera (gynecological ailments), Eclipta prostrata (liver ailments), ivy gourd (diabetes), citron (hypertension), Centella asiatica (psychological ailments), spade nose shark (lactogogue), reticulate whipray (wheezing and bronchitis), Katelysia opima (impotence), Indian squid (anemia), and Indian oil sardine (anabolic). More studies on these claims will help identify novel functional foods to add to the field of medical nutrition therapy, with traditional brand identity. Robust studies on the documentation of the traditional knowledge on marine resources will yield a good database for various stakeholders and policy makers.

Hepatoprotective effect of lawsone on rifampicin-isoniazid induced hepatotoxicity in in vitro and in vivo models.

The Drug-induced liver injury is one of the common unfavourable impacts, which seriously affects any drug therapy. This study documented the hepatoprotective efficacy of lawsone, the major bioactive naphthoquinone present in Lawsonia inermis L. (Lythraceae) using in vitro and in vivo models. Lawsone was isolated from the leaves of L. inermis and its structure was confirmed using spectroscopic data. In-vitro antioxidant effect of lawsone was evaluated using ABTS assay. Hepatoprotective effect of lawsone was determined with RIF-INH treated HepG2 cells and Wistar rats. Administration of RIF-INH reduced the viability of the HepG2 cells and the treatment with lawsone significantly restored the viability of the cells even at lower concentration (7.5 µM). The other parameters such as the leakage of transaminases and MDA levels were also significantly reduced by the treatment with lawsone. Oral administration of lawsone to the animals did not show any toxicity up to 2 g/kg b.w. concentration. Treatment with lawsone to the RIF-INH administered animals significantly lowered the serum transaminases levels. The ratio of albumin to globulin was improved and the level of bilirubin was lowered. This study indicated the hepatoprotective effect of lawsone; detailed investigations will give deeper understanding of the application of lawsone for hepatoprotection.

Effect of two andrographolide derivatives on cellular and rodent models of non-alcoholic fatty liver disease.

The prevalence of Non-Alcoholic Fatty Liver Disease (NAFLD) is increasing and there is an increasing interest in natural products to treat NAFLD. This study aimed to evaluate the hepatoprotective effect of andrographolide and two of its derivatives; in one the OH group at C-14 was removed and in the other OH groups at C-3 and C-19 were protected. Andrographolide (AN) was isolated from the aerial parts of Andrographis paniculata Wall. Isoandrographolide (IAN) and 3,19-acetonylidene andrographolide (ANA) were derivatized from AN. Drug likeness of the compounds was studied using DataWarrior. The effect of the compounds in ameliorating hepatic steatosis and lipotoxicity was assessed using palmitate-oleate induced steatotic HepG2 cell lines. In vivo efficacy of the compounds was assessed by using HFD fed rats. IAN showed comparatively high drug score and low irritability than AN. MTT assay indicated that the treatment with IAN had comparatively less toxicity than AN and ANA to HepG2 cells. The treatment with IAN significantly reduced the lipid accumulation and the leakage of LDH and transaminases, while the treatments with AN and ANA did not prohibit the leakage. In the in vivo experiment, the treatment with IAN showed comparatively better hepatoprotection by reducing the serum lipid, transaminases and ALP levels than with AN and ANA. Our results showed that IAN could be a promising lead to treat NAFLD with comparatively low toxicity and improved efficacy.

Identification of putative QTLs for seedling stage phosphorus starvation response in finger millet (Eleusine coracana L. Gaertn.) by association mapping and cross species synteny analysis.

A germplasm assembly of 128 finger millet genotypes from 18 countries was evaluated for seedling-stage phosphorus (P) responses by growing them in P sufficient (Psuf) and P deficient (Pdef) treatments. Majority of the genotypes showed adaptive responses to low P condition. Based on phenotype behaviour using the best linear unbiased predictors for each trait, genotypes were classified into, P responsive, low P tolerant and P non-responsive types. Based on the overall phenotype performance under Pdef, 10 genotypes were identified as low P tolerants. The low P tolerant genotypes were characterised by increased shoot and root length and increased root hair induction with longer root hairs under Pdef, than under Psuf. Association mapping of P response traits using mixed linear models revealed four quantitative trait loci (QTLs). Two QTLs (qLRDW.1 and qLRDW.2) for low P response affecting root dry weight explained over 10% phenotypic variation. In silico synteny analysis across grass genomes for these QTLs identified putative candidate genes such as Ser-Thr kinase and transcription factors such as WRKY and basic helix-loop-helix (bHLH). The QTLs for response under Psuf were mapped for traits such as shoot dry weight (qHSDW.1) and root length (qHRL.1). Putative associations of these QTLs over the syntenous regions on the grass genomes revealed proximity to cytochrome P450, phosphate transporter and pectin methylesterase inhibitor (PMEI) genes. This is the first report of the extent of phenotypic variability for P response in finger millet genotypes during seedling-stage, along with the QTLs and putative candidate genes associated with P starvation tolerance.

Antihyperlipidemic and hepatoprotective effects of Gardenin A in cellular and high fat diet fed rodent models.

The gum of Gardenia resinifera Roth., is one of the important drugs used in the Indian system of medicine and a source of unique polymethoxylated flavones. This study was aimed to evaluate the antihyperlipidemic and anti-NAFLD effects of Gardenin A (Gar-A) from G. resinifera gum using in vitro and in vivo models. Gar-A was isolated from G. resinifera gum and was identified on the basis of the physical and spectral data. Toxicity of Gar-A to HepG2 cells was evaluated using MTT assay. The ability of Gar-A to reduce steatosis was assessed using oleate-palmitate induced HepG2 cell lines by estimating the lipid levels by ORO staining and by estimating the intracellular triglyceride content. Effect of Gar-A on amelioration of lipotoxicity was measured by estimating the LDH levels. The doses for in vivo experiments were fixed by Irwin test, between 50 and 100 mg/kg concentrations, through oral route. The acute antihyperlipidemic effect of Gar-A was assessed in Triton WR-1339 induced hyperlipidemic animals. The chronic antihyperlipidemic and anti-NAFLD effects of Gar-A were evaluated in HFD fed rats. In vitro experiments with HepG2 cell line indicated that the cells treated with Gar-A did not show any significant reduction in the viability up to 70 µg/mL concentration. Steatotic HepG2 cells treated with Gar-A showed a significant reduction in lipid accumulation at 2.5-10 µg/mL concentrations. In triton induced hyperlipidemic rats, the treatment significantly reduced the lipid levels at the synthesis phase. The treatment with Gar-A to the HFD fed animals significantly lowered the steatosis and transaminase levels. The other biochemical parameters such as TC, TG, LDL-c, ALP and ACP were also decreased significantly. Treatment with Gar-A significantly lowered the hyperlipidemia and fat accumulation in the liver; detailed molecular investigations are necessary to establish the antihyperlipidemic and hepatoprotective potentials of Gar-A.

Tracing QTLs for Leaf Blast Resistance and Agronomic Performance of Finger Millet (Eleusine coracana (L.) Gaertn.) Genotypes through Association Mapping and in silico Comparative Genomics Analyses.

Finger millet is one of the small millets with high nutritive value. This crop is vulnerable to blast disease caused by Pyricularia grisea, which occurs annually during rainy and winter seasons. Leaf blast occurs at early crop stage and is highly damaging. Mapping of resistance genes and other quantitative trait loci (QTLs) for agronomic performance can be of great use for improving finger millet genotypes. Evaluation of one hundred and twenty-eight finger millet genotypes in natural field conditions revealed that leaf blast caused severe setback on agronomic performance for susceptible genotypes, most significant traits being plant height and root length. Plant height was reduced under disease severity while root length was increased. Among the genotypes, IE4795 showed superior response in terms of both disease resistance and better agronomic performance. A total of seven unambiguous QTLs were found to be associated with various agronomic traits including leaf blast resistance by association mapping analysis. The markers, UGEP101 and UGEP95, were strongly associated with blast resistance. UGEP98 was associated with tiller number and UGEP9 was associated with root length and seed yield. Cross species validation of markers revealed that 12 candidate genes were associated with 8 QTLs in the genomes of grass species such as rice, foxtail millet, maize, Brachypodium stacei, B. distachyon, Panicum hallii and switchgrass. Several candidate genes were found proximal to orthologous sequences of the identified QTLs such as 1,4-ß-glucanase for leaf blast resistance, cytokinin dehydrogenase (CKX) for tiller production, calmodulin (CaM) binding protein for seed yield and pectin methylesterase inhibitor (PMEI) for root growth and development. Most of these QTLs and their putatively associated candidate genes are reported for first time in finger millet. On validation, these novel QTLs may be utilized in future for marker assisted breeding for the development of fungal resistant and high yielding varieties of finger millet.

Larvicidal and Pupicidal Activities of Alizarin Isolated from Roots of Rubia cordifolia Against Culex quinquefasciatus Say and Aedes aegypti (L.) (Diptera: Culicidae).

The mosquitocidal activities of different fractions and a compound alizarin from the methanol extract of Rubia cordifolia roots were evaluated on larvae and pupae of Culex quinquefasciatus Say and Aedes aegypti (L.) (Diptera: Culicidae). Larvae and pupae were exposed to concentrations of 2.5, 5.0, 7.5 and 10 ppm for fractions and 0.5, 1.0, 1.5 and 2.0 ppm for compound. After 24 h, the mortality was assessed and the LC50 and LC90 values were estimated for larvae and pupae. Among the 23 fractions screened, fraction 2 from the methanol extract of R. cordifolia showed good mosquitocidal activity against C. quinquefasciatus and A. aegypti. LC50 and LC90 values of fraction 2 were 3.53 and 7.26 ppm for C. quinquefasciatus and 3.86 and 8.28 ppm for A. aegypti larvae, and 3.76 and 7.50 ppm for C. quinquefasciatus and 3.92 and 8.05 ppm for A. aegypti pupae, respectively. Further, the isolated compound alizarin presented good larvicidal and pupicidal activities. LC50 and LC90 values of alizarin for larvae were 0.81 and 3.86 ppm against C. quinquefasciatus and 1.31 and 6.04 ppm for A. aegypti larvae, respectively. Similarly, the LC50 and LC90 values of alizarin for pupae were 1.97 and 4.79 ppm for C. quinquefasciatus and 2.05 and 5.59 ppm for A. aegypti pupae, respectively. The structure of the isolated compound was identified on the basis of spectroscopic analysis and compared with reported spectral data. The results indicated that alizarin could be used as a potential larvicide and pupicide.

In vitro antiproliferative activity of 2,3-dihydroxy-9,10-anthraquinone induced apoptosis against COLO320 cells through cytochrome c release caspase mediated pathway with PI3K/AKT and COX-2 inhibition.

The present study investigated the anticancer activity of 2,3-dihydroxy-9,10-anthraquinone against different cancer cells such as MCF-7, COLO320, HepG-2, Skov-3, MOLM-14, NB-4, CEM, K562, Jurkat, HL-60, U937, IM-9 and Vero. 2,3-dihydroxy-9,10-anthraquinone showed good antiproliferative activity against COLO320 cells when compared to other tested cells. The cytotoxicity results showed 79.8% activity at the dose of 2.07 µM with IC50 value of 0.13 µM at 24 h in COLO320 cells. So we chose COLO320 cells for further anticancer studies. mRNA expression was confirmed by qPCR analysis using SYBR green method. Treatment with 2,3-dihydroxy-9,10-anthraquinone was found to trigger intrinsic apoptotic pathway as indicated by down regulation of Bcl-2, Bcl-xl; up regulation of Bim, Bax, Bad; release of cytochrome c and pro-caspases cleaving to caspases. Furthermore, 2,3-dihydroxy-9,10-anthraquinone stopped at G0/G1 phase with modulation in protein levels of cyclins. On the other hand PI3K/AKT signaling plays an important role in cell metabolism. We found that 2,3-dihydroxy-9,10-anthraquinone inhibits PI3K/AKT activity after treatment. Also, COX-2 enzyme plays a major role in colorectal cancer. Our results showed that the treatment significantly reduced COX-2 enzyme in COLO320 cells. These results indicated antiproliferative activity of 2,3-dihydroxy-9,10-anthraquinone involving apoptotic pathways, mitochondrial functions, cell cycle checkpoint and controlling the over expression genes during the colorectal cancer. Molecular docking studies showed that the compound bound stably to the active sites of Bcl-2, COX-2, PI3K and AKT. This is the first report of anticancer mechanism involving 2,3-dihydroxy-9,10-anthraquinone in COLO320 cells. The present results might provide helpful suggestions for the design of antitumor drugs toward colorectal cancer treatment.

Assessment of genetic diversity, population structure and relationships in Indian and non-Indian genotypes of finger millet (Eleusine coracana (L.) Gaertn) using genomic SSR markers.

We evaluated the genetic variation and population structure in Indian and non-Indian genotypes of finger millet using 87 genomic SSR primers. The 128 finger millet genotypes were collected and genomic DNA was isolated. Eighty-seven genomic SSR primers with 60-70 % GC contents were used for PCR analysis of 128 finger millet genotypes. The PCR products were separated and visualized on a 6 % polyacrylamide gel followed by silver staining. The data were used to estimate major allele frequency using Power Marker v3.0. Dendrograms were constructed based on the Jaccard's similarity coefficient. Statistical fitness and population structure analyses were performed to find the genetic diversity. The mean major allele frequency was 0.92; the means of polymorphic alleles were 2.13 per primer and 1.45 per genotype; the average polymorphism was 59.94 % per primer and average PIC value was 0.44 per primer. Indian genotypes produced an additional 0.21 allele than non-Indian genotypes. Gene diversity was in the range from 0.02 to 0.35. The average heterozygosity was 0.11, close to 100 % homozygosity. The highest inbreeding coefficient was observed with SSR marker UGEP67. The Jaccard's similarity coefficient value ranged from 0.011 to 0.836. The highest similarity value was 0.836 between genotypes DPI009-04 and GPU-45. Indian genotypes were placed in Eleusine coracana major cluster (EcMC) 1 along with 6 non-Indian genotypes. AMOVA showed that molecular variance in genotypes from various geographical regions was 4 %; among populations it was 3 % and within populations it was 93 %. PCA scatter plot analysis showed that GPU-28, GPU-45 and DPI009-04 were closely dispersed in first component axis. In structural analysis, the genotypes were divided into three subpopulations (SP1, SP2 and SP3). All the three subpopulations had an admixture of alleles and no pure line was observed. These analyses confirmed that all the genotypes were genetically diverse and had been grouped based on their geographic regions.

In vitro anticancer activity of methyl caffeate isolated from Solanum torvum Swartz. fruit.

The present study was undertaken to investigate the anticancer activity of methyl caffeate isolated from Solanum torvum Swartz. fruit and to explore the molecular mechanisms of action in MCF-7 cells. Cytotoxic properties of hexane, ethyl acetate and methanol extracts were carried out against MCF-7 cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Ethyl acetate extract showed good cytototoxic activities compared to hexane and methanol extracts. Methyl caffeate was isolated from the ethyl acetate extract using column chromatography. Cytotoxic properties of methyl caffeate was investigated against MCF-7, A549, COLO320, HepG-2 and Vero cells. The compound showed potent cytotoxic properties against MCF-7 cells compared to A549, COLO320 and HepG-2 cells. Methyl caffeate significantly reduced cell proliferation and increased formation of fragmented DNA and apoptotic body in MCF-7 cells. Bcl-2, Bax, Bid, p53, caspase-3, PARP and cytochrome c release were detected by western blot analysis. The activities of caspases-3 and PARP gradually increased after the addition of isolated compound. Bcl-2 protein was down regulated; Bid and Bax were up regulated after the treatment with methyl caffeate. Molecular docking studies showed that the compound bound stably to the active sites of poly (ADP-ribose) polymerase-1 (PARP1), B cell CLL/lymphoma-2 (BCL-2), E3 ubiquitin-protein ligase (MDM2) and tubulin. The results strongly suggested that methyl caffeate induced apoptosis in MCF-7 cells via caspase activation through cytochrome c release from mitochondria.

Antimicrobial, antioxidant, and cytotoxic properties of vasicine acetate synthesized from vasicine isolated from Adhatoda vasica L.

Adhatoda vasica (L.) (Acanthaceae) is used in the indigenous system of medicine in India. The alkaloid Vasicine was isolated from ethanolic extract of the leaves of A. vasica using column chromatography. Vasicine acetate was obtained by acetylation of Vasicine. Vasicine acetate exhibited good zone of inhibition against bacteria: 10 mm against E. aerogenes, 10 mm against S. epidermidis, and 10 mm against P. aeruginosa. Vasicine acetate showed minimum inhibitory concentration values against bacteria: M. luteus (125 µg/mL), E. aerogenes (125 µg/mL), S. epidermidis (125 µg/mL), and P. aeruginosa (125 µg/mL). The radical scavenging activity of Vasicine acetate was the maximum at 1000 µg/mL (66.15%). The compound showed prominent cytotoxic activity in vitro against A549 lung adenocarcinoma cancer cell line. Quantification of Vasicine and Vasicine acetate by HPLC-DAD analysis showed their contents to be 0.2293% and 0.0156%, respectively, on dry weight basis of the leaves. Vasicine acetate could be probed further in drug discovery programme.

A flavonoid isolated from Streptomyces sp. (ERINLG-4) induces apoptosis in human lung cancer A549 cells through p53 and cytochrome c release caspase dependant pathway.

The aim of this study was to investigate the anticancer activity of a flavonoid type of compound isolated from soil derived filamentous bacterium Streptomyces sp. (ERINLG-4) and to explore the molecular mechanisms of action. Cytotoxic properties of ethyl acetate extract was carried out against A549 lung cancer cell line using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cytotoxic properties of isolated compound were investigated in A549 lung cancer cell line, COLO320DM cancer cell line and Vero cells. The compound showed potent cytotoxic properties against A549 lung cancer cell line and moderate cytotoxic properties against COLO320DM cancer cell line. Isolated compound showed no toxicity up to 2000 µg/mL in Vero cells. So we have chosen the A549 lung cancer cell line for further anticancer studies. Intracellular visualization was done by using a laser scanning confocal microscope. Apoptosis was measured using DNA fragmentation technique. Treatment of the A549 cancer cells with isolated compound significantly reduced cell proliferation, increased formation of fragmented DNA and apoptotic body. Activation of caspase-9 and caspase-3 indicated that compound may be inducing intrinsic and extrinsic apoptosis pathways. Bcl-2, p53, pro-caspases, caspase-3, caspase-9 and cytochrome c release were detected by western blotting analysis after compound treatment (123 and 164 µM). The activities of pro-caspases-3, caspase-9 cleaved to caspase-3 and caspase-9 gradually increased after the addition of isolated compound. But Bcl-2 protein was down regulated after treatment with isolated compound. Molecular docking studies showed that the compound bound stably to the active sites of caspase-3 and caspase-9. These results strongly suggest that the isolated compound induces apoptosis in A549 cancer cells via caspase activation through cytochrome c release from mitochondria. The present results might provide helpful suggestions for the design of antitumor drugs toward lung cancer treatment.

Novel 1,5,7-trihydroxy-3-hydroxy methyl anthraquinone isolated from terrestrial Streptomyces sp. (eri-26) with antimicrobial and molecular docking studies.

Streptomyces sp. isolate ERI-26 was obtained from the Nilgiris forest soil of Western Ghats, Tamil Nadu, India. Novel anthraquinone compound was isolated from the active fraction 5; it was identified by spectroscopical data using UV, IR, NMR and MASS. The isolated compound 1,5,7-trihydroxy-3-hydroxy methyl anthraquinone was tested against bacteria and fungi at minimum inhibitory concentration level. The compound showed significant antimicrobial activity against bacteria, Staphylococcus aureus at 125 µg/ml, Staphylococcus epidermidis at 62.5 µg/m, Bacillus subtilis at 31.25 µg/ml, fungi; Epidermophyton floccosum at 62.5 µg/ml, Aspergillus niger at 31.25 µg/ml, Aspergiller flavus at 31.25 µg/ml, Trichophyton rubrum at 62.5 µg/ml and Botrytis cinerea at 62.5 µg/ml. The isolated compound was subjected to molecular docking studies for the inhibition of TtgR, topoisomerase IV and AmpC ß-lactamase enzymes which are targets for antimicrobials. Docking studies of the compound showed low docking energy indicating its usefulness as antimicrobial agent. 1,5,7-Trihydroxy-3-hydroxy methyl anthraquinone is new, and its antimicrobial and molecular docking properties are reported for the first time.

Documentation and quantitative analysis of local ethnozoological knowledge among traditional healers of Theni district, Tamil Nadu, India.

ETHNOPHARMACOLOGICAL RELEVANCE: This study investigated the use of animals among the traditional healers in Theni district of Tamil Nadu, India. The data regarding the medicinal animals/animal products were documented and their usages were analyzed quantitatively. MATERIALS AND METHODS: Based on free list interviews with the traditional healers, we documented the medicinal usage of animals/animal products and calculated the indices such as informant consensus factor (Fic) to determine the consensus over the species for an illness category, as well as the Index Agreement on Remedies (IAR) to determine the extent of potential utilization of each species. RESULTS: In this study, 69 medicinal animals/animal products were documented with the help of standardized questionnaires among the local healers. The results were tabulated and Fic value for each illness category was calculated. Three illness categories viz., jaundice (milk of Capra aegagrus hircus), orthopedics (egg white and meat of Gallus gallus domesticus) and pediatrics (milk of Equus africanus asinus) had got high Fic values. Fifteen illness categories had moderate Fic values. Highly cited animals in these illness categories were: Rusa unicolor (antiemetic), Reticulitermes spp. (diabetes), flesh of Varanus benghalensis (oral ailments), milk (eye ailments, fever) and urine (antidote) of Homo sepians, meat of Trachypithecus johnii (respiratory ailments), various parts of C. aegagrus hircus (blood ailments, coolants, diarrhea, pulmonary and urinary ailments), flesh of Chamaeleon zeyalnica (neural ailments), meat of Passer domesticus (aphrodisiac), curd and dung of Bos primigenius taurus (dermatological ailments), meat of G. domesticus (musculo-skeletal disorders, analgesic), meat of Lissemys punctata (hemorrhoids), and Pherthima posthuma (psychological ailments). Six illness categories had low Fic values. CONCLUSION: This study indicated that the animals are still being used by the local healers of Theni district, to treat various illnesses. Cross-disciplinary approaches to explore the full potential of animal-derived medicines will help to improve the health of local people.

Antimicrobial and cytotoxicity properties of 2,3-dihydroxy-9,10-anthraquinone isolated from Streptomyces galbus (ERINLG-127).

Streptomyces galbus ERINLG-127 was isolated from the soil samples of the Marapalam forest, Nilgiris, India. The ethyl acetate extract was subjected to activity-guided fractionation by column chromatography over silica gel. This led to the isolation of 2,3-dihydroxy-9,10-anthraquinone as the active principle. The compound showed good antimicrobial activity against tested bacteria and fungi. The minimum inhibitory concentration values of isolated compound were 12.5 µg/mL against Pseudomonas aeruginosa, Klebsiella pneumoniae, Salmonella typhimurium, K. pneumoniae (ESBL-3971), K. pneumoniae (ESBL-3894) and Staphylococcus aureus (MRSA). The compound showed prominent cytotoxic activity in vitro against A549 lung adenocarcinoma cancer cell line. It showed 75.1 % activity at the dose of 100 µg/mL with IC50 value of 60 µg/mL. The isolated compound was subjected to molecular docking studies for the inhibition of TtgR and Topoisomerase IV enzymes which are targets for antimicrobials. Docking studies of the compound showed low docking energy indicating its usefulness as antimicrobial agent.