Abstract
This study presents an approach that utilizes low-value agro-industrial by-products as culture media for producing high-value proteolytic enzymes. The objective was to assess the impact of six agro-industrial by-products as culture media on the production of proteolytic enzymes. Bacillus subtilis strains, confirmed through comprehensive biochemical, morphological, and molecular analyses, were isolated and identified. Enzymatic activity was evaluated using azocasein and casein substrates, and the molecular sizes of the purified extract components were determined. The results demonstrated that the isolated bacteria exhibited higher metabolic and enzymatic activity when cultured in media containing 1 % soybean oil cake or feather meal. Furthermore, higher concentrations of the culture media were found to hinder the production of protease. Optimal protease synthesis on soybean oil cake and feather meal media was achieved after 4 days, using both the azocasein and casein methods. Semi-purification of the enzymatic extract obtained from Bacillus subtilis in feather meal and soybean oil cake resulted in a significant increase in azocaseinolytic and caseinolytic activities. Gel electrophoresis analysis revealed multiple bands in the fractions with the highest enzymatic activity in soybean oil cake, indicating the presence of various enzymes with varying molecular sizes. These findings highlight the potential of utilizing low-value agro-industrial by-products as efficient culture media for the sustainable and economically viable production of proteolytic enzymes with promising applications in various industries.