Chitinases biosynthesis by immobilized Aeromonas hydrophila SBK1 by prawn shells valorization and application of enzyme cocktail for fungal protoplast preparation

• Biodegradation of prawn shell by immobilized Aeromonas hydrophila SBK1 in Ca-alginate.
• Process optimization and scale-up of chitinases production and reuse of biocatalyst.
• Evaluation of fermentation and thermodynamic viability of cell immobilization.
• Quantification of chitosaccharides and its free radical scavenging activity.
• Protoplast formation of Aspergillus niger by crude chitinases treatment and its regeneration.

Production and optimization of β-N-acetyl glucosaminidase and chitinase by Ca-alginate immobilized Aeromonas hydrophila SBK1 was carried out using prawn shell as cost-effective substrate. Beads prepared with 5.0% Na-alginate (containing 2.0% colloidal chitin) and 1.0 M CaCl2 showed considerable beads integrity and supported maximum production of chitinolytic enzymes. Bead diameter, 3 mm; temperature, 35°C; pH 7.0; agitation, 90 rpm were found ideal for the maximum production of the enzymes. The fermentation and thermodynamic indices revealed the feasibility of immobilized cells over free cells for enzymes production. Reasonable amount of chitosaccharides (degree of polymerization; 1–6) accumulated in the production media which have paramount antioxidant activity. Scale up experiment was successfully carried out in 5 L fermentor. In immobilized state, the chitosaccharides yield and antioxidant activity increased about 44.76% and 22.22%, whereas specific productivity of β-N-acetyl glucosaminidase and chitinase increased by 22.86% and 33.37% over free state. The cell entrapped beads can be reused upto ten cycles without marked loss of its biocatalytic efficiency. High level of protoplast of Aspergillus niger was generated by treating mycelia with 10 U/ml of crude chitinase after 4 h at pH 7.0 and in the temperature 35–40°C, and 67% of the protoplasts were found to be regenerated.