Development of thermostable amylase enzyme from Bacillus cereus for potential antibiofilm activity.

The marine bacterial strain Bacillus cereus was used to produce amylase enzyme and has excellent alkali-stable and thermostable enzymatic activity. The combined effects of pH, temperature and incubation time on amylase activity were studied using response surface methodology. The amylase enzyme activity was also determined in the presence of various metal ions, chelating agents, detergents and the results showed that the maximum enzyme activity was observed in the presence of calcium chloride (96.1%), EDTA (63.4%) and surf excel (90.6%). The amylase enzyme exhibited excellent antibiofilm activity against marine derived biofilm forming bacteria Pseudomonas aeruginosa and Staphylococcus aureus in microtiter plate assay and congo red assay. Light and confocal laser scanning microscopic (CLSM) analysis were also used to confirm the potential biofilm activity of amylase enzyme. The CLSM analysis showed the inhibition of complete biofilm formation on amylase enzyme treated glass surface. Further in vivo toxicity analysis of amylase enzyme was determined against marine organisms Dioithona rigida and Artemia salina. The results showed that there is no morphological changes were observed due to the minimal toxicity of amylase enzyme. Overall these findings suggested that marine bacterial derived amylase enzyme could be developed as potential antibiofilm agent.

Structural characterization and anticancer activity (MCF7 and MDA-MB-231) of polysaccharides fractionated from brown seaweed Sargassum wightii.

The purpose of this study was to investigate the anticancer activity of polysaccharides from brown seaweed Sargassum wightii (SWP) on human breast cancer cells. Initially, two polysaccharide fractions (SWP1 and SWP2) were isolated and purified from the crude polysaccharides using DEAE-52 cellulose and Sephadex G-100 column chromatography. As a result, SWP1 was obtained with the yield of 21.48% was characterized using chemical analysis, GC-MS, 1H NMR and 13C NMR. The chemical composition of the extracted polysaccharide contains a neutral polysaccharide with a high total sugar content and low protein, phenol and flavonoid content. GC-MS analysis revealed the presence of galactofuranose and arabinose and NMR spectra shows the presence of ß-galactose signals. Anticancer activity shows that the polysaccharides significantly reduce the proliferation of breast cancer cells (MCF7 and MDA-MB-231) in a dose-dependent manner. Further, polysaccharides induced the apoptosis in the breast cancer cells by increasing ROS generation, cleaving mitochondrial membrane and nuclei damage. Finally, polysaccharides increased the activity of caspase 3/9, thus leads to apoptosis of breast cancer. Together, polysaccharides from S. wightii could be a new source of natural anticancer agent against breast cancer with potential value in the manufacturing supplements and drugs.

Marine steroid derived from Acropora formosa enhances mitochondrial-mediated apoptosis in non-small cell lung cancer cells.

p53 pathway has been revealed to mediate cellular stress responses and trigger DNA repair, cell cycle arrest, senescence, and apoptosis. We isolated 2-ethoxycarbonyl-2-ß-hydroxy-A-nor-cholest-5-ene-4one (ECHC) from butanol extracts of scleractinian coral Acropora formosa and reported its potential antioxidant and antimicrobial activity as well as less toxicity against zebrafish Danio rerio. In the present study, we intend to explore p53-mediated apoptosis pathway enhanced by ECHC in A549 human non-small cell lung cancer cell lines. This report shows that ECHC increases ROS generation and sensitizes mitochondrial membrane that leads to the release of cytochrome C (Cyto C) into cytosol. Further, ECHC decreases the expression of antiapoptotic genes such as TNF-α, IL-8, Bcl2, MMP2, and MMP9 which are actively involved in cancer cell proliferation, invasion, and metastasis etc. It also increases the expression of apoptotic genes Cyto C, Bax, and p21, which are responsible for cell cycle arrest and cell death. The tumor suppressor p53 was also observed to be upregulated during ECHC treatment in untransformed cells and was more likely to result in cell cycle arrest, senescence, and apoptosis. Finally, ECHC also down regulates the expression of caspase-9 and caspase-3 which are the death stage of intrinsic apoptosis. Our findings suggested that ECHC enhances ROS generation and mitochondrial sensitization determines the threshold for irreversible p53-mediated intrinsic apoptosis pathway.