Журнал микробных и биохимических технологий

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Hyper-production of Alkaline Protease by Mutagenic Treatment of Bacillus subtilis M-9 using Agroindustrial Wastes in Submerged Fermentation

Sadia Javed, Munazzah Meraj, Shazia Anwer Bukhari, Rao Irfan and Saqib Mahmood

Green chemistry technologies are the powerful tool for the management of environmental wastes challenges. Agro industrial residues are composed of complex polysaccharides that support the microbial growth for the production of useful products (enzymes, organic acids, drugs, etc.). Disposal and environment friendly management of these wastes has become a global priority. The aim of present investigation was to improve the alkaline protease yield by treating the parent Bacillus subtilis M-9 strain with different mutagens UV-irradiations, N-methyl-N-nitro- N-nitrosoguinidine (NTG), Ethidium bromide (EB), using agroindustrial wastes (banana stalk and corn stover) in submerged fermentation. Fifteen positive mutants were selected on skim milk agar plates for shake flask experiments. BSU-5 mutant strain showed 81.21± 3.24 PU/mL alkaline protease activity higher than parent strain (23.57 ± 1.19 PU/mL) in optimized fermentation medium. The fermentation profile like pH (9), temperature (45°C), inoculum size (2 mL), incubation time (24 hrs, and kinetic parameters such as u (h-1), Yp/s, Yp/x, Yx/s, qs, Qs, qp also confirmed the hyper proteolytic activity of alkaline protease produced from BSU-5 mutant strain over parent strain and other mutants. Finally, the BSU-5 mutant strain was immobilized by entrapping it in calcium alginate beads and agar. Alkaline protease production and stability of biocatalyst were investigated in both free and immobilized cells. It was concluded from the study, immobilized cells were more efficient for enzyme production then free cells when used repeatedly.