European Journal of Experimental Biology Open Access

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Lab- scale production of extracellular pectinase by soil bacteria using fruit waste as substrate

Rachana Oak-Dhamapurkar, Sayali Naphade-Mahajan, Meeta Bhot and Jossy Varghese

Pectinases are pectin-degrading enzymes predominantly used as biocatalyst in various industries for wine extraction, fruit juice extraction, making of paper pulp etc. The large-scale production of pectinase using biological systems (bacteria, fungi, plants etc.) is one of the common methods used in industry. In the current study, bacterial isolates obtained from various soil samples were used for pectinase production. An enrichment culture technique was used to isolate 04 bacterial strains named as P1, P2, P3 and P4 which were subsequently characterized by 16S rRNA gene sequencing and biochemical tests. Lab-scale submerged fermentation was carried out using these isolates in synthetic media containing pectin as substrate (extracted from peels of Lemon, Sweet lime, Orange) & Lab pectin. Fermentation conditions were optimized. Supernatant containing the extra-cellular crude pectinase enzyme was collected after fermentation. The maximum pectinase production was seen after 7 days of fermentation at R.T. (28±20C) and pH 7.0 in shaker conditions. Effect of environmental parameters (pH, temperature, enzyme and substrate concentration) was studied on the activity of crude pectinase enzyme. The maximum enzyme activity of crude pectinase enzyme was found to be at R.T. (28±20C) and 37°C in the pH range of 6-8. Partial purification of pectinase was done using ammonium sulfate precipitation method followed by dialysis. The dialyzed enzymes obtained were subjected to DNSA assay (for enzyme activity) and Folin-Lowry assay (for protein estimation) for determining maximum pectinase activity and the amount of pectinase present in the supernatant respectively. By using SDS- PAGE, molecular weight of pectinase was determined. Thus, the current study may help in providing alternate and eco-friendly source of pectin that can be degraded by bacterial pectinase and used in food industry.