Purification and Evaluation of Antimicrobial Activity and Biofilm Formation of Bacillus thuringiensis aizawai Strain HD283

Abstract

Objective: The study aimed to evaluate antimicrobial and antibiofilm activities, and purify the alkaline protease enzyme produced by Bacillus thuringiensis aizawai HD283. Methods: The experimental methodology encompassed the following steps: preparation of the inoculum, evaluate antimicrobial and antibiofilm activities, formulation of production media, enzyme activity assays, protein quantification, ammonium sulfate precipitation, acetone fractionation, and final purification using Sephadex Gel filtration chromatography. Results: After cultivating Bacillus thuringiensis aizawai HD283 under optimal environmental conditions and extracting the crude enzyme, as described in our previous research, the enzyme was subjected to the following purification steps: The purification pattern was as follow, the most active ammonium sulfate fractionation portion was at 60-75 % ammonium sulfate saturation, with 2.16 up to 2.41 % yield recovery, while the most active fraction by acetone precipitation, was wide range from 60-90 % acetone with yield recovery ranged between 13.4 to 12.0 %. Whereas, G-100 Sephadex column chromatography (final purification step) has 1.44 purification factor and 4.5 % final yield recovery of extracted enzyme.