Time and Temperature Stability of Collagenase Produced by Bacillus licheniformis

The aim of the present study was to evaluate the stability during storage period of collagenase obtained when culturing Bacillus licheniformis ATCC 14580 and Bacillus amyloliquefaciens isolated from soil, in a basal salt medium supplemented with 0.3% sucrose and 0.6% sheep fur meal. The obtained enzyme was lyophilized and used for further trials. Enzyme solutions (0.5 g/ml) were prepared using lyophilized product stored in different conditions, at room temperature (20-22 °C), refrigeration (4-6 °C) and freezing (-15 to -18 °C) and tested for collagenase activity immediately after preparation, after 24 hours, 1, 7, 14, 21 and 30 days. The influence of enzyme concentration, when keeping the product in buffer solution at refrigerator, was also tested and 0.2, 0.5 and 0.75 g/ml solutions were analyzed for their proteolytic activity during 30 days of maintaining at 4-8 °C. Resistance to repeated frosting and defrosting was assayed using 0.5 g/ml enzyme solution, up to five cycles, with 24 hours freezing period. The enzymatic activity of collagenase was determined by spectrophotometric method based on ninhydrin reaction and protein content was assessed according to Lowry method.