Pectinase Activity

Pectinase assay was carried out using the DNS method (Miller 1959). The reaction mixture (1.0 mL) containing equal amounts of the substrate (0.5%) prepared in citrate buffer (0.05 M pH 4.4) and the suitably diluted enzyme was incubated at 50°C for 30 min in a water bath. After incubation, 3 mL DNS solution was added to stop the reaction and tubes were kept in boiling water for 10 min. On cooling, the developed colour was read at 575 nm using a UV-visible spectrophotometer. The amount of released reducing sugar was quantified using D-galacturonic acid as a standard. The enzyme activity was calculated as the amount of enzyme required to release one micromole equivalent of D-galacturonic acid per minute under assay condition.