Journal of Pure and Applied MicrobiologyVol. 11 No. 4

Partial Purification and Characterization of Cellulolytic Enzymes Extracted from Trichoderma reesei Inoculated Digested Biogas Slurry

Ajit Kaur1*, Surekha Bhatia2 and Urmila Gupta Phutela3

1Department of Biofuels, Centre of Excellence in Farm Machinery, CSIR-CMERI Gill Road-141006, Ludhiana, India. 2Department of Processing and Food Engineering, Punjab Agricultural University-141004, Ludhiana, India. 3School of Renewable Energy Engineering, Punjab Agricultural University, Ludhiana-141004, Punjab, India.

Received on 10 November 2017 and accepted on 16 December 2017

 

ABSTRACT

Alternative sources of energy are the demand of the modern world. For generating different alternate fuels, utilization of lignocellulolytic biomass is on top of the priority list, for which cellulase enzymes are required specifically. In this context, the aim of the presented paper was the partial purification and characterization of cellulolytic enzymes from Trichoderma reesei inoculated digested biogas slurry. The enzyme was produced at previously standardized conditions (Incubation period: 15 day, Spore concentration: 108 spores/ml, Slurry concentration: 25%). The cellulolytic enzymes viz. CMCase, Cellobiase and FilterPaperase produced were then partially purified by ammonium sulphate precipitation (0-30 and 30-80%) and dialysis followed by ion exchange chromatography using DEAE-cellulose column. Twelve fold purification was achieved for cellobiase. Specific activity of 20.18 U/mg was measured. Two isoforms of Cellobaise (C-I and C-II) were found with 21 and 32 fold purification, respectively. Upon characterization, the optimal pH and optimal temperature values for cellobiase came out to be 7.5 and 55C for C-I and 25C for C-II. Carboxymethyl cellulase was partially purified up to 10.4 fold with specific activity of 1.87 U/mg of protein and Fpase was purified to 11.3 fold with 1.47 U/mg of protein specific activity. Partially purified enzyme activities were compared with that of commercial enzymes. This is a novel work where cellulases were extracted and partially purified from digested slurry from biogas plant, which is very significant with reference to not only disposal of digested biogas slurry but also its value addition for industrial applications.

Keywords : Digested Biogas Slurry, Enzymes, Ion Exchange Chromatography, Partial Purification, Trichoderma reesei.