Process optimization, purification, and characterization of cellulase from Aspergillus uvarum and its industrial application in saccharification

Abstract

The breakdown of cellulose, the most prevalent carbon resource on Earth, by cellulase is very important for acquiring soluble sugars. Solid-state fermentation (SSF) stands out as a proficient approach for generating economically valuable compounds, facilitating cost reduction in production. The fermentation factors were optimized to enhance cellulolytic enzyme production. Of the various inexpensive and readily accessible lignocellulosic residues, sorghum straw emerged as the utmost appropriate substrate. The maximal cellulase productivity of 14.12 ± 0.06 U/g DS was obtained after 72 h of fermentation using sorghum straw with 10% v/v moisture content at pH 7, 37 °C, and an inoculum volume of 1.5% v/v. The crude cellulase was purified using various methods. The employment of the aqueous two-phase system (ATPS) utilizing a polyethylene glycol 8000/MnSO₄ combination demonstrated optimal purification, resulting in a 26.02-fold enhancement in activity, a yield of 48.7%, and a partition coefficient of 1.27. The molecular size of cellulase was approximated to be 84 kDa using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The optimum pH and temperature for enzyme activity were identified as 7.0 and 50 °C, respectively. The cellulolytic activity exhibited the highest stimulation in the presence of Mn²⁺. Upon enzymatic saccharification of alkali-treated sorghum feedstock, the highest reducing sugar (30.51 ± 0.13 mg/mL) was obtained after an incubation period of 72 h, with a substrate loading of 4% w/v, enzyme concentration 30 U/g DS, pH 5, and the presence of Tween-80 as a surfactant. These findings may pave the way for a cost-effective and competent process within the framework of the biorefinery concept.

Graphical abstract