Thermokinetic studies of Aspergillus tamarii Growth and alkaline protease production in a real time bio reaction calorimeter

Abstract

Protease, also known as peptidyl-peptide hydrolase, an important industrial enzyme, is responsible for approximately 60% of all enzyme sales, projected as 1.8 billion USD in the year 2015. Proteases are utilized extensively in leather industry for de-hairing. The focus of this thesis is on alkaline protease production by Aspergillus tamarii in a Real Time Biological Reaction Calorimeter (BioRTCal) for bio thermokinetic data generation. This study revealed that both growth and non-growth related reactions involved in this cell culture metabolism could be monitored efficiently by calorimeter and the heat yield values used for better design of fermentor and scale up. Optimization studies for growth and enhanced production of alkaline protease by Aspergillus tamarii were carried out in a batch stirred tank bioreactor. Using Hess s law the relationship between the online metabolic heat variable and the offline bioprocess variables, i.e. biomass generation, substrate consumption were explored and validated. Production of alkaline protease by A. tamarii was accomplished using agricultural by-products such as cotton seed meal and skimmed milk within a lower incubation period compared with what was reportedly obtained with wheat bran and soya flour. Macromorphology and broth rheology studies coupled with BioRTCal studies revealed that pellet formation resulted in low broth viscosity and high enzyme production. Alternative methods of oxygen supply are important, especially in viscous fermentations and shear-sensitive fermentations. Liquid phase (needbased) oxygen supply through H2O2 for alkaline protease production in BioRTCal by A. tamarii fungal cells was successfully attempted. This methodology helped to eliminate the gas liquid transport resistance. In BioRTcal studies, the optimal H2O2 concentration was 0.5 mM, and was added in pulses to maintain the required DO level. Higher protease yield was obtained in H2O2 based cultivation in comparison with conventional aeration method. newline newline newline