Unraveling anticancer potential of a novel serine protease inhibitor from marine yeast Candida parapsilosis ABS1 against colorectal and breast cancer cells.

The study was planned to isolate a serine protease inhibitor compound with anticancer potential against colorectal and breast cancer cells from marine yeast. Protease enzymes play a crucial role in the mechanism of life-threatening diseases like cancer, malaria and AIDS. Hence, blocking these enzymes with potential inhibitors can be an efficient approach in drug therapy for these diseases. A total of 12 marine yeast isolates, recovered from mangrove swamps of Sundarbans, India, showed inhibition activity against trypsin. The yeast isolate ABS1 showed highest inhibition activity (89%). The optimum conditions for protease inhibitor production were found to be glucose, ammonium phosphate, pH 7.0, 30 °C and 2 M NaCl. The PI protein from yeast isolate ABS1 was purified using ethyl acetate extraction and anion exchange chromatography. The purified protein was characterized using denaturing SDS-PAGE, Liquid Chromatography Electrospray Ionization Mass Spectrometry (LC-ESI-MS), Reverse Phase High Pressure Liquid Chromatography (RP-HPLC) and Fourier Transform Infra-red Spectroscopy (FTIR) analysis. The intact molecular weight of the PI protein was determined to be 25.584 kDa. The PI protein was further studied for in vitro anticancer activities. The IC50 value for MTT cell proliferation assay was found to be 43 µg/ml against colorectal cancer HCT15 cells and 48 µg/ml against breast cancer MCF7 cells. Hoechst staining, DAPI staining and DNA fragmentation assay were performed to check the apoptotic cells. The marine yeast was identified as Candida parapsilosis ABS1 (Accession No. MH782231) using 18s rRNA sequencing.

Biodecolorization and Biodegradation of Azo Dye Reactive Orange-16 by Marine Nocardiopsis sp.

BACKGROUND: Azo dyes are xenobiotic compounds that have bioaccumulated in the environment due to escalated industrial development. These are hazardous in nature, possessing carcinogenic and mutagenic effects on human beings. OBJECTIVES: The perspective of the present study was to isolate and to determine azo dye (Reactive Orange-16) degrading potential of marine actinobacteria isolated from sediment samples of Port Blair, India. MATERIAL AND METHODS: Actinobacteria with dye decolorization potential were isolated from sea sediment samples. The actinobacterial isolate with the highest dye decolorizing percentage was identified with the help of phenotypic, biochemical and molecular studies. The different physico-chemical parameters for dye decolorization were also optimized. The nature of decolorization by the potent isolate was determined with the help of High Performance Liquid chromatography (HPLC) and Fourier Transformed Infrared spectroscopy (FTIR) techniques. Further the toxicity of RO-16 decolorized products was investigated with the help of phytotoxcity assay. RESULTS: Out of six actinobacterial isolates, VITVAMB 1 possessed the most efficient RO-16 decolorization property. It decolorized 85.6% of RO-16 (250 mg L-1) within 24hrs. Isolate VITVAMB 1 was identified to be Nocardiopsis sp. Maximum dye decolorization occurred at pH 8, temperature 35°C, 3% salt concentration and a dye concentration of 50 mg L-1. CONCLUSIONS: The nature of decolorization by Nocardiopsis sp. was biodegradation. Additionally, the degraded dye metabolites were found to be less toxic than pure dye. The high decolorization potential of VITVAMB 1 and the low toxicity of its degradation products make it a prospective dye removal system. The marine origin of VITVAMB 1 also makes it an attractive source for novel azo dye reducing enzymes.

Characterisation and identification of antibacterial compound from marine actinobacteria: In vitro and in silico analysis.

OBJECTIVE: The present study was focused on the characterization and in silico analysis of antibacterial compound derived from marine actinobacteria isolated from the sediments of salterns of Ongole, Andhra Pradesh, India. METHODS: The sediment sample was serially diluted and marine actinobacteria were isolated in actinomycetes isolation agar. A total of 9 colonies were recovered and among them, 5 morphologically distinct isolates were selected for further processing. The antibacterial activity of these five isolates was tested against 4 clinical isolates collected from Narayani Hospital, Vellore, Tamil Nadu. RESULTS: The isolate SJP4 showed inhibitory activity against all the test pathogens viz., Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Bacillus cereus. The structure of the compounds extracted from SJP4 was identified as 8-diaza-2,9-dibenzoyl-5,6-diphenyl-2,8-decadienedioic acid diethyl ester and [1,2,4]triazol-1-ylethanone through GCMS analysis. Molecular docking studies was done using Autodock software. These two compounds were docked into the binding site of DNA gyrase and found to have binding energy of -6.55(Kcal/mol) and -4.86(Kcal/mol) respectively. The potential actinobacteria isolate was identified as Nocardiopsis dassonvillei SJPB4 strain (Accession no. MG434671) using 16s rRNA sequencing. CONCLUSION: We are concluding that as the compounds were successfully docked on to the active site of DNA gyrase.

Biofilm inhibition formation of clinical strains of Pseudomonas aeruginosa mutans, photocatalytic activity of azo dye and GC-MS analysis of leaves of Lagerstroemia speciosa.

The investigation was conducted to analyse the bioactive compounds from the leaf extracts of L. speciosa by GC-MS. The extracts were screened for antibacterial and antibiofilm activities against potential clinical strains. The bioactive compounds from the leaves of L. speciosa were extracted by soxhlet continuous extraction method and their chemical composition was analysed by Gas Chromatography-Mass Spectroscopy (GC-MS). The antibacterial activity was evaluated against clinical strain like Staphylococcus aureus, Escherichia coli, P. aeruginosa and Salmonella typhi by well diffusion technique. We also screened for antibacterial property against common food borne pathogens namely Listeria monocytogenes and Bacillus cereus at varied concentration 250µml-1 to 1000µml-1. Thereafter antibiofilm assay was carried out at from 250 to 1000µg/ml against P. aeruginosa (high biofilm forming pathogen) clinical strain by cover slip technique and the morphology of the pathogen was observed using Scanning Electron Microscopy-(SEM). It was observed that diverse class of secondary metabolites were found by GC-MS analysis for all the extracts upon the continuous extraction. It was found that only minimum inhibition was seen in alcoholic extract for antibacterial activity, whereas all other extracts showed negligible activity. P. aeruginosa biofilm inhibited to 93.0±2% and 91±2% at higher concentration (1000µg/ml) for methanolic and ethanolic extract respectively. Absence of extracellular matrix structure and the surface cracking of biofilm were viewed by SEM, which confirmed the antibiofilm activity. Hence this study reveals that L. speciosa showed significant antibiofilm activity against P. aeruginosa due to the phytoconstituents present in the leaf extracts which was well documented in the alcoholic extracts by GC-MS analysis. The methanolic and ethanolic extract showed good photocatalytic activity of 77.44% and 96.66% against azo dye degradation respectively. Further, isolating the novel phyto-compounds would yield better promising biological activities.

Efficacy of protease inhibitor from marine Streptomyces sp. VITBVK2 against Leishmania donovani - An in vitro study.

In the present study the leishmanicidal effect of potential protease inhibitor producing marine actinobacterial isolate has been investigated against Leishmania donovani, the causative agent of visceral leishmaniasis. Among 89 marine actinobacteria isolated from a salt pan in Kanyakumari, only one isolate (BVK2) showed 97% of protease inhibition activity against trypsin. Moderate to high protease inhibitor activity was shown by isolate BVK2 on proteinase (30%) and chymotrypsin (85%). In optimization study for protease inhibitor production glucose as carbon source and casein as nitrogen source showed the best activity. In the in-vitro Fluorescence-activated cell sorting (FACS) assay, 100 µg/ml of BVK2 extract was active against amastigotes in infected J774A.1 macrophages and showed 87% of parasitic inhibition. The isolate BVK2 showed significant anti-parasitic activity with an IC50 of 27.1 µg/ml after double doses were administered. The potential isolate was identified by molecular 16S rRNA gene sequencing as Streptomyces sp. VITBVK2. The results obtained suggest that the marine actinobacterial extract which have novel metabolites can be considered as a potential source for the development of drugs.

Postprandial Antihyperglycemic And Antioxidant Activities of Acalypha indica Linn Stem Extract: An In-vivo Study.

BACKGROUND: α-glucosidase inhibitors controls postprandial hyperglycemia (PPHG) by lowering sharp rise in blood glucose levels after ingestion of carbohydrate rich meal in type 2 diabetic (T2D) individuals. Acalypha indica commonly known as Indian copper leaf is used in traditional medicinal system to treat various diseases. In our previous in-vitro investigation, methanolic extract of A. indica stems (AIS) proved to be an effective a-glucosidase inhibitor, antioxidant, and well tolerated in acute and subchronic toxicity studies in albino wistar rats. OBJECTIVE: In this perspective, this study was designed to evaluate postprandial antihyperglycemic potential of AIS in maltose, sucrose, and glucose loaded streptozotocin (STZ)-induced normal and diabetic rats. As, the acute hyperglycemia at postprandial period has more triggering effect on oxidative stress, study was also aimed to evaluate the antioxidant potential of AIS on STZ-induced Albino-Wistar rats. MATERIALS AND METHODS: Rats were treated with AIS (300-600 mg/kg b.w.) to investigate effect of AIS in controling PPHG after carbohydrate loading. Hepatoprotective activity of AIS is evaluated in diabetic rats by treating them at the dosages 300-600 mg/kg b.w. RESULTS: Studies revealed 69.10 and 80.35% blood glucose-lowering effect of AIS in maltose and sucrose loaded diabetic rats in comparison with the diabetic control group. AIS recovered the liver damage caused by streptozotocin. CONCLUSION: The present study confirmed high potential of AIS in controling PPHG by inhibiting a-glucosidase enzyme in maltose and sucrose loaded diabetic rats. AIS also exhibited hepatoprotective activity in STZ-induced diabetic rats. Thus, AIS could be used as a nutraceutical supplement to treat T2D effectively. SUMMARY: AIS extract is effective in suppressing maltose and sucrose-induced postprandial hyperglycemic spikes in ratsAIS treat ment showed a 69.10 and80.35% blood glucose-lowering effect in maltose and sucrose loaded diabetic rats in comparison with the diabetic control group.AIS also improved the antioxidant status in diabetic rats and also has recovered the liver damage caused by streptozotocin.The α-glucosidase inhibitor isolated from AIS is a good supplement to control postprandial blood glucose level in the management of type 2 diabetes. Abbreviations used: AIS: Acalypha indica Stems, ALP: Alkaline Phosphatase, b/w: Body Weight, PPHG: Postprandial hyperglycemia, SE: Standard Error, SGOT: Serum glutamate oxaloacetate transaminase, SGPT: Serum glutamate pyruvate transaminase, SOD: Superoxide dismutase, STZ: Streptozotocin, TB: Total Bilirubin, T2D: Type 2 diabetes.

Evaluating the effectiveness of marine actinobacterial extract and its mediated titanium dioxide nanoparticles in the degradation of azo dyes.

Aim of the present study was to synthesize titanium dioxide nanoparticles (TiO2 NPs) from marine actinobacteria and to develop an eco-friendly azo-dye degradation method. A total of five actinobacterial isolates were isolated from Chennai marine sediments, Tamilnadu, India and analyzed for the synthesis of TiO2 NPs using titanium hydroxide. Among these, the isolate PSV 3 showed positive results for the synthesis of TiO2 NPs, which was confirmed by UV analysis. Further characterization of the synthesized TiO2 NPs was done using XRD, AFM and FT-IR analysis. Actinobacterial crude extract and synthesized TiO2 NPs was found efficient in degrading azo dye such as Acid Red 79 (AR-79) and Acid Red 80 (AR-80). Degradation percentage was found to be 81% for AR-79, 83% for AR-80 using actinobacterial crude extract and 84% for AR-79, 85% for AR-80 using TiO2 NPs. Immobilized actinobacterial cells showed 88% for AR-79 and 81% for AR-80, dye degrading capacity. Degraded components were characterized by FT-IR and GC-MS analysis. The phytotoxicity test with 500 µg/mL of untreated dye showed remarkable phenotypic as well as cellular damage to Tagetes erecta plant. Comparatively no such damage was observed on plants by degraded dye components. In biotoxicity assay, treated dyes showed less toxic effect as compared to the untreated dyes.

Protease inhibitors from marine actinobacteria as a potential source for antimalarial compound.

The study was planned to screen the marine actinobacterial extract for the protease inhibitor activity and its anti- Pf activity under in vitro and in vivo conditions. Out of 100 isolates, only 3 isolates exhibited moderate to high protease inhibitor activities on trypsin, chymotrypsin and proteinase K. Based on protease inhibitor activity 3 isolates were chosen for further studies. The potential isolate was characterized by polyphasic approach and identified as Streptomyces sp LK3 (JF710608). The lead compound was identified as peptide from Streptomyces sp LK3. The double-reciprocal plot displayed inhibition mode is non-competitive and it confirms the irreversible nature of protease inhibitor. The peptide from Streptomyces sp LK3 extract showed significant anti plasmodial activity (IC50: 25.78 µg/ml). In in vivo model, the highest level of parasitemia suppression (≈ 45%) was observed in 600 mg/kg of the peptide. These analyses revealed no significant changes were observed in the spleen and liver tissue during 8 dpi. The results confirmed up-regulation of TGF-ß and down regulation of TNF-α in tissue and serum level in PbA infected peptide treated mice compared to PbA infection. The results obtained infer that the peptide possesses anti- Pf activity activity. It suggests that the extracts have novel metabolites and could be considered as a potential source for drug development.

Streptomyces sp. LK3 mediated synthesis of silver nanoparticles and its biomedical application.

In the present study, the marine actinobacteria mediated biosynthesis of silver nanoparticles (AgNps) was achieved using Streptomyces sp LK3. The synthesized AgNps showed the characteristic absorption spectra in UV-vis at 420 nm, which confirmed the presence of nanoparticles. XRD analysis showed intense peaks at 2θ values of 27.51°, 31.87°, 45.57°, 56.56°, 66.26°, and 75.25° corresponding to (210), (113), (124), (240), (226), and (300) Bragg's reflection based on the fcc structure of AgNps. The FTIR spectra exhibited prominent peaks at 3,417 cm(-1) (OH stretching due to alcoholic group) and 1,578 cm(-1) (C=C ring stretching). TEM micrograph showed that the synthesized AgNps were spherical in shape with an average size of 5 nm. Surface morphology and topographical structure of the synthesized AgNps were dignified by AFM. The synthesized AgNps showed significant acaricidal activity against Rhipicephalus microplus and Haemaphysalis bispinosa with LC50 values of 16.10 and 16.45 mg/L, respectively. Our results clearly indicate that AgNps could provide a safer alternative to conventional acaricidal agents in the form of a topical antiparasitic formulation. The present study aimed to develop a novel, cost-effective, eco-friendly actinobacteria mediated synthesis of AgNps and its antiparasitic activity.

Biotransformation of Direct Blue 1 by a moderately halophilic bacterium Marinobacter sp. strain HBRA and toxicity assessment of degraded metabolites.

The ability of halophiles to survive in the extreme salt concentrations has gained them the importance of being used in the treatment of industrial waste waters. A moderately halophilic bacterial strain with the ability to degrade the complex azo dye Direct Blue-1 (DB-1) was isolated from sea water and identified as Marinobacter sp. strain HBRA. Complete decolorization of DB-1 (100 mg L(-1)) was achieved in 6h at 37 °C, pH 8 and with 70 g L(-1) NaCl. Decolorization was analyzed by UV-vis spectrophotometer. The FT-IR spectrum revealed that Marinobacter sp. strain HBRA specifically targeted azo bond (NN) at 1631 cm(-1) to break down Direct Blue-1. Formation of metabolites at different retention times in HPLC indicated degradation. Biotransformation pathway for DB-1 was proposed based on LC-MS. Phytotoxicity study revealed the less toxic nature of the metabolites compared to the dye. Genotoxicity with Allium cepa confirmed the cytotoxic nature of DB-1 by inducing several chromosomal abnormalities compared to the negligible effects of degraded metabolites. The current study is the first report on the detoxification of DB-1 by Marinobacter sp. strain HBRA.