Statistical optimization and kinetic modeling simulation for fibrinolytic enzyme production by Bacillus tropicus from mahewu, a traditional South African fermented food

Abstract

Cardiovascular diseases are a leading cause of global mortality, with existing thrombolytic drugs having side effects, short half-life, and high costs, warranting alternatives. Since the discovery of nattokinase from Japanese natto, fibrinolytic enzymes from fermented foods have been explored as promising thrombolytic agents. However, most research has focused on Asian fermented foods, with limited studies on African sources. In this study, Bacillus tropicus DUT M1, an extracellular fibrinolytic enzyme producer, was isolated for the first time from mahewu, an indigenous fermented food. Optimization was performed using a combination of OFAT and RSM. Glucose and casein were the optimal carbon and nitrogen sources identified through OFAT, while casein, meat extract, and pH were significant in the Plackett-Burman design. CCD optimized the medium to 28.23 g/L casein, 6.41 g/L meat extract, and pH 7.90, increasing enzyme production 13.9-fold, 12 U/mL to 167 U/mL. Kinetic modeling validated S-Gompertz as the best fit (R² = 0.97) compared to Logistic-1, Richards, and S- Weibull- 2 models. Fibrinolytic activity was confirmed via quantitative tube assay, fibrin zymography, and blood clot lysis experiments. These findings establish mahewu as a novel source of fibrinolytic enzyme producers, highlighting the therapeutic and industrial potential of African fermented foods.