Screening For High Cellulase-producing Strains And Optimization Of Enzyme Complexes For Hydrolysis Of Lignocellulosic Biomass

To find a new enzyme resource for hydrolysis of lignocellulosic biomass, we screened the cellulase-producing strains from soil samples. By developing a high-throughput microplate-based technique for microbial cultivation and cellulase assay, and combining with traditional strain-screening procedure, we successfully obtained a high cellulase-producing strain Aspergillus fumigatus ECU0811. After simple optimization of cultivation conditions for the strain, the cellulase activity was improved from0.23FPU/ml to0.41FPU/ml which was close to that of Trichoderma reesei. The optimized culture condition was:corn cob powders3.0%, peptone0.3%, yeast extract0.05%,(NH4)2SO40.15%, KH2PO40.2%, Mg2SO40.015%, CaCl20.03%, Tween-800.2%, Mandel’s trace (MnSO4;1.6g/1; ZnSO4;1,4g/1; FeSO4,5g/1; CoCl22g/1)0.1%, pH3.0. Next, we prepared a crude cellulase (powder) of Aspergillus fumigatus through liquid fermentation, centrifugation, ultrafiltration and lyophilization. It was combined with the other two individual enzymes (crude almond β-glucosidase and xylanase) as supplementary enzymes for enzymatic hydrolysis of steam explosion-pretreated corn stover (CS). A high-throughput method was again used in the biohydrolysis of CS as well as in the assay of glucose released. Using a response surface methodology, we attempted to compose an optimally supplemented multiple-enzyme mixture on the basis of commercially available cellulose (AccelleraseTM1000, from Genencor Co.). The enzyme mixture contained crude almond β-glucosidase1.63mg protein/g glucan, Aspergillus fumigatus crude cellulase2.52mg protein/g glucan, xylanase7.85mg/g glucan and AccelleraseTM100029mg protein/g glucan could improve the glucan-to-glucose conversion to99.6%, with lower protein load as compared to neat AccelleraseTM1000.