Optimizing PHB and Protease Production by Box Behnken Design

Abstract

Mixed culture is more suitable to adapt more flexible fermentation process and produce different product simultaneously. In this study a mixed Bacillus culture was investigated for their ability to produce the bioplastic "Polyhydroxybutyrate" and both of the mesophilic and the thermophilic proteases in one flask. Box-Behnken experimental design was used. The produced amount of PHB has been increased significantly. Meanwhile there is a competition between PHB and proteases. The maximum produced amount of PHB using Box-Behnken design was 2.82 g/l/48 h with protease activity equal to 41.9 Units/ml/48 h for thermophilic proteases and 99.65 Units/ml/48 h for mesophilic proteases. Excel solver was used for extra-optimization for the optimum conditions obtained from Box-Behnken experiments and its model. The maximum PHB obtained after using Excel solver was 2.88 g/l/48 h. The maximum mesophilic and thermophilic activities obtained at the same PHB production conditions were 175.68 and 243.38 Units/ml respectively. The model accuracy as obtained from Excel solver was 118.8%, which prove the power of the experimental design in optimizing such complicated process. The strategies used in this study are recommended for the production of PHB and different proteases simultaneously using Bacillus mixed culture.

ABSTRAK: Kultur campuran adalah lebih sesuai bagi proses penapaian yang fleksibel dan ia boleh menghasilkan produk yang berbeza secara serentak. Dalam kajian ini keupayaan  menghasilkan "Polyhydroxybutyrate" bioplastik serta mesofilik dan termofilik protease dalam satu flask oleh  kultur Bacillus campuran telah disiasat. Eksperimen rekabentuk Box-Behnken telah digunakan. Jumlah PHB yang dikeluarkan meningkat dengan ketara dan terdapat persaingan antara PHB dan protease. Jumlah keluaran PHB maksima menggunakan rekabentuk Box-Behnken adalah 2.82 g/l/48 jam dengan aktiviti protease sama dengan 41.9 Unit/ml/48 jam untuk protease termofilik dan 99.65 Unit/ml/48 h untuk protease mesofilik. Solver Excel telah digunakan untuk memperolehi kadar optimum tambahan dari keadaan optimum yang diperolehi dari experimen dan model Box-Behnken. PHB maksimum diperolehi setelah menggunakan solver Excel adalah 2.88 g/l/48 jam. Aktiviti mesofilik dan termofilik maksima diperolehi daripada keadaan pengeluaran PHB yang sama adalah 175.68 dan 243.38 Unit / ml. Ketepatan model diperolehi daripada solver Excel adalah 118.8%, membuktikan kekuatan eksperimen tersebut bagi mengoptimumkan proses yang rumit. Strategi yang digunakan dalam kajian ini adalah disyorkan bagi pengeluaran PHB dan proteas berbeza secara serentak menggunakan kultur Bacillus campuran.

KEYWORDS: PHB; proteases; mixed culture; Box-Behnken design.