Strain Development of Aspergillus brasiliensis Using Physical and Chemicals Mutagenesis for Possible Overproduction of Xylanase, Amylase, Protease and Cellulase under Submerged Fermentation (SmF)

Aims: Xylanase is commonly involved in the complete hydrolysis of xylan which consisted of hemicelluloses and celluloses as important components of plant cell wall. This enzyme is essential in wide range of applications in various industries along with other hydrolytic enzymes such as amylase, protease and cellulase. Microbial enzymes including xylanase possess several advantages over the use of conventional chemical catalysts due to greater catalytic activity, larger amount of production, higher degree of substrate specificity, higher economical viability and better biodegradable ability without destructive threat to the environment. Global enzymes market is expected to rise to USD millions by 2020. Due to the potential and bright future prospects of these microbial enzymes in biotechnological applications, random mutagenesis that is one of the potential approaches to produce enzymes with positive motivating characteristics is proposed and implemented. Therefore, the present study was aimed to create mutants of Aspergillus brasiliensis using physical and chemical random mutagenesis and to elucidate possible overproduction of xylanase, amylase, protease and cellulase by mutants of A. brasiliensis under submerged fermentation (SmF).

Methodology: In this study, A. brasiliensis that has been classified as GRAS (generally regarded as safe) was subjected to physical mutagenesis of ultraviolet (UV) irradiation and chemical mutagenesis of nitrous acid (NA) and ethyl methane sulfonate (EMS) before screened for the possible overproduction of xylanase, amylase, protease and cellulase in SmF using wheat bran (agro-industrial residual) and yeast extract as the carbon and nitrogen source at the initial medium pH of 6.50 under the optimised growth conditions at 150 rpm at 30°C for prolonged 168 h of fermentation.

Results: Based on the results, xylanase possessed as the highest maximum production of enzyme was notably observed at the optimum fermentation period of 72 h produced by EMS-mutated mutant of A. brasiliensis. Interestingly, 2 mg/mL of EMS was exerted as the optimum chemical mutagen followed by UV irradiation of 254 nm from a distance of 20 cm and lastly 0.17 mg/mL of NA. In fact, the maximum xylanase activity of 11.325±0.881 U/mL was attained after A. brasiliensis exposed to EMS for 45 minutes. On the other hand, the UV treated mutant of 30 minutes produced 10.052±0.987 U/mL of xylanase activity. Lastly, 9.127±0.410 U/mL of xylanase activity was obtained from NA treated mutant for 15 minutes compared to the wild type with 7.821±1.460 U/mL. The xylanase activity in mutants exposed to EMS, UV and NA was notably increased by 45%, 29% and 17% compared to the wild type. On the other hand, the enhancement in amylase activity was also observed in two mutants after UV exposure for 25 minutes and EMS treatment for 60 minutes with the production of 0.481±0.007 U/mL and 0.304±0.040 U/mL compared to wild type with 0.232±0.021 U/mL, respectively. Notably, there were increment of 107% and 31% in activity in mutants compared to wild type. Nevertheless, no possible overproduction of protease and cellulase was detected after the physical and chemical mutagenesis in this study.

Conclusion: In a nutshell, the mutants of A. brasiliensis with enhanced production of xylanase and amylase were anticipated to fulfill the industrial demand in more economical approach using agro-residual waste of wheat bran under SmF.