Valorization of Paper Pulp Mill Sludge for Protease Production by Indigenous Bacillus tropicus P4

This study explores the potential of using paper pulp mill sludge (PPMS) as an economical substrate for producing high-value protease enzymes with an indigenous bacterial strain, Bacillus tropicus P4. Isolated directly from PPMS, B. tropicus P4 showed high protease-producing ability, approximately 134 U/mL after 48 h—more than three times the yield of the benchmark strain ( B. megaterium ). Among various additives tested to boost enzyme production, Tween 80 emerged as the most effective, increasing enzyme activity by more than threefold compared to the control. Scale-up experiments in bioreactors of 5 L and 150 L confirmed that B. tropicus P4 maintains high protease yields under typical cultivation conditions with minimal modifications, specifically the addition of Tween 80 (1%) and increased total solids concentration (25 g/L). In the 5 L bioreactor, enzyme production peaked at approximately 755 U/mL within 24 h, while the 150 L bioreactor consistently achieved high enzyme activity (~848 U/mL). These results support the feasibility of a simple and scalable approach for converting industrial sludge into high-value protease enzymes, contributing to resource recovery and circular bioeconomy strategies.