Marwa Hamdi

Surfactant- and oxidant-stable alkaline proteases from Bacillus invictae: Characterization and potential applications in chitin extraction and as a detergent additive

A newly alkaline proteases producing strain was isolated from sea water. The strain was identified as Bacillus invictae on the basis of biochemical characteristics and 16S rRNA sequence analysis. The crude protease activity showed an optimal activity at approximately 60°C and in wide pH interval ranging from 9.0 to 11.0. At least six clear caseinolytic protease bands were observed in a zymogram. Phenylmethylsulfonyl fluoride (PMSF), a serine-protease inhibitor, was found to inhibit completely the protease activity. The crude alkaline proteases showed high stability toward solid and liquid detergents. Furthermore, wash performance analysis revealed that the crude enzyme could effectively remove blood stain when added to commercial detergent. In addition, the crude proteases were found to be effective in the deproteinization of shrimp shell waste. The percent of protein removal after 3h of hydrolysis at 50°C with an E/S ratio of 10U/mg of protein or after fermentation by the strain were about 76% and 82%, respectively. Thus, the results of the present study showed that the crude proteases of B. invectae could be effectively used in several industrial applications, as an eco-friendly agent.

Enhancement of extracellular polymeric substances (EPS) production in Spirulina (Arthrospira sp.) by two-step cultivation process and partial characterization of their polysaccharidic moiety

The interactive effects of light intensity and NaCl concentration were investigated for Spirulina two-step cultivation process using Full Factorial Design. In the experiment interval, light intensity had no effect while the NaCl concentration had significant effect on the enhancement of extracellular polymeric substances (EPS) production. Interestingly, results revealed a significant negative interaction between light and NaCl concentration indicating that high NaCl concentration (40gL-1) and low light intensity (10μmol photons m-2s-1) enhanced the EPS production. Under these conditions, EPS production reached a maximum of 1.02gg-1 of biomass (dry weight), which is 1.67-folds greater than EPS content under optimal growth conditions (10μmol photons m-2s-1, 1gL-1, 30°C). Desalting and deproteinezation steps of EPS were efficient to obtain polysaccharides (PS) with high carbohydrate (67.3±1.1%), low soluble proteins (5.14±0.32%), ash (5.85±0.71%) and sulfate (2.42±0.12%) contents. Rheological studies of PS at different concentrations (1%, 2.5% and 5%) revealed that the viscosity of the solution increased with the increase of PS concentration. In addition, PS exhibited a non Newtonian shear-thinning nature, a predominant gel-like behavior and a good resistance to consecutive heating-cooling cycles. The adopted process could be, then, a promising and economic strategy to enhance EPS production and extract polysaccharides with interesting rheological properties.

Chitin extraction from blue crab (Portunus segnis) and shrimp (Penaeus kerathurus) shells using digestive alkaline proteases from P. segnis viscera

Since chitin is closely associated with proteins, deproteinization is a crucial step in the process of extracting chitin. Thus, this research aimed to extract chitin from Portunus segnis and Penaeus kerathurus shells by means of crude digestive alkaline proteases from the viscera of P. segnis, regarding deproteinization step, as an alternative to chemical treatment. Casein zymography revealed that five caseinolytic proteases bands exist, suggesting the presence of at least five different major proteases. The optimum pH and temperature for protease activity were pH 8.0 and 60°C, respectively, using casein as a substrate. The crude enzymes extract was highly stable at low temperatures and over a wide range of pH from 6.0 to 12.0. The crude alkaline protease extract was found to be effective in the deproteinization of blue crab and shrimp shells, to produce chitin. The best efficiency in deproteinization (84.69±0.65% for blue crab shells and 91.06±1.40% for shrimp shells) was achieved with an E/S ratio of 5U/mg of proteins after 3h incubation at 50°C. These results suggest that enzymatic deproteinization of crab and shrimp wastes by fish endogenous alkaline proteases could be a potential alternative in the chitin production process.

Biomedical potential of chitosan-silver nanoparticles with special reference to antioxidant, antibacterial, hemolytic and in vivo cutaneous wound healing effects

The aim of the present study was to prepare chitosan-PVA-silver nanoparticles (CS-AgNPs) through green method. Chitosan and PVA polymers acted as stabilizing agents. DLS and TEM analyses showed that CS-AgNPs were homogeneously dispersed in matrix with an average size of 190-200u202fnm. The CS-AgNPs were tested for their antioxidant and antibacterial properties and the results revealed that they exhibited higher antioxidant activity than CS powder. Moreover, CS-AgNPs were characterized by a low cytotoxicity effect at 5-200u202fμg/ml against Chinese Hamster Ovary (CHO-K1) cells. In addition, the prepared CS-Ag NPs were found to promote significantly the wound healing, as determined by the wound contraction ratio and histological examination. A significant improvement in wound healing progression and in oxidative stress damage were observed for CS, CS-PVA and CS-AgNPs-treated wound tissues, when compared to control and CICAFLORA®-treated groups. The wound healing effect could be attributed to the antibacterial and antioxidant synergy of AgNPs and CS. Results strongly support the possibility of using CS-AgNPs for wound care applications.

Antioxidant, antibacterial and in vivo wound healing properties of laminaran purified from Cystoseira barbata seaweed

Laminaran, a polysaccharide extracted from marine algae, exhibits attractive properties being non-toxic, hydrophilic and biodegradable. The aim of this study was to investigate the effectiveness of a cream based on the brown seaweed Cystoseira barbata laminaran (CBL) for healing full thickness wounds induced on rats. The antibacterial activity of CBL was evaluated against Gram positive and Gram negative bacteria and the antioxidant properties were assessed using five different assays. To highlight the healing effectiveness of CBL-based cream, the response to treatment was assessed by macroscopic, histologic and biochemical parameters and was compared to controls. CBL, recovered (7.27%) by ultrafiltration (1-10u202fkDa) and predominantly consisted of (1u202f→u202f3)-linked β-d-glucopyranose residues with a small level of (1u202f→u202f6)-glycosidic bonds, showed noticeable antioxidant and antibacterial properties. The cream containing CBL as an active ingredient exerted a promoting healing effect. The wound contraction reached 98.57u202f±u202f1.31% after thirteen days of treatment. The derma in CBL treated group is properly arranged and revealed an improved collagen deposition and an increased fibroblast and vascular densities compared with the control groups. Overall, these results established, for the first time, a scientific in vivo evidence of the efficiency of CBL as a wound healing agent of interest in modern medicine.