Aspergillus niger fermentation of cocoa pod husk for increased total crude protein utilizing response surface methodology

Cocoa pod husk (CPH) waste from chocolate and cocoa powder production is repurposed as raw materials for animal feed using response surface methodology (RSM) with a faced central composite design (FCCD). The investigated variables included fermentation time (ranging from 7 to 21 days), the percentage of molasses (ranging from 0 to 10%), and the liquid-to-solid (L/S) ratio ranging from 0 to 15 (mg/l). Seventeen experimental runs were executed, and the accuracy of the model was verified through ANOVA (R² = 0.9878, R²_adj = 0.9804, p < 0.0001). The optimal conditions for achieving the highest predicted total crude protein content (TCPC) were determined to be as follows: 7.93% TCPC, a fermentation period of 21 days, 10% molasses, and solid-state fermentation (L/S = 0). Aspergillus niger were used in each run to degrade CPH, boosting TCPC compared to unfermented CPH. SEM analysis confirmed CPH degradation by the microorganism. Notably, the sample collected from these optimal conditions exhibited a reduced theobromine content of 0.58 ± 0.02 mg/100 g dry mass, along with heightened antioxidant properties. This result indicates a greater-than-expected increase in protein content, highlighting fermentation's potential to enhance the value of CPH for animal feed.