Enhanced cellulase recovery in SSF from Rhizopus oryzae SN5 and immobilization for multi-batch saccharification of carboxymethylcellulose

Abstract

In this study enhanced the cellulase recovery was achieved from wheat bran solid state fermentate of a locally isolated fungus, Rhizopus oryzae SN5 by using “one factor at a time” technique. Interestingly, 18.9% enhanced cellulase production (452.16 ± 2.38 U/gds) was observed with extraction optima of Na-citrate buffer (50 mM, pH 4.8), 1:10 solid: solvent ratio, 90 min soaking time at 40 °C. Further, cellulase was partially purified, where in 5.73-fold enhanced specific activity was achieved with product recovery of 7.1%. Moreover, the molecular weight of the purified cellulase was determined as~80 kDa using SDS-PAGE and zymogram analysis. Furthermore, Ca-alginate immobilized cellulase (IMC) exhibited increased Km (2.2 ± 0.07 to 2.78 ± 0.09 mg/ml) and decreased Vmax (31.36 ± 1.37 to 27.12 ± 1.21 μmol/min/ml) than free enzyme. In addition, the turn over number (Kcat) was found to be increased (542.4 ± 14.38 to 625.2 ± 12.11 Min−1), whereas catalytic efficiency (Kcat/Km) was decreased (246.54 ± 8.34 to 224.89 ± 9.07 Min−1.mg−1.ml−1) for IMC. The pH and thermal stability of IMC was found to be 30–40% higher in comparison to the free cellulase. Notably, the IMC displayed, easy recovery and high reusability for Carboxymethylcellulose (CMC) hydrolysis, with 70.6% cellulase activity retention after five successive cycles of their reuse. Taken together, the optimized extraction process and developed IMC could be used in cellulose based industries to cut down the saccharification cost.