Abstract
Colorimetric enzyme assays are suitable for evaluating enzyme activity in nutrient solutions. In this study, variations of the standard methods were tested in order to simplify handling, adapt set-ups to processing large numbers of samples and increase sensitivity of the assays. Hide Powder Azure (HPA) was identified as the most suitable substrate for proteases in nutrient solutions. With this substrate significant higher protease activity was found compared to Gelatin Remazol Brilliant Blue (Gel-RBB), whereas Casein Remazol Brilliant Blue (Cas-RBB) did not show any proteolytic activity. Large number of samples might be treated in a miniaturized set-up in 96-well-microplates. Suitability of assays for chitinolytic, cellulolytic and β-1,3-glucanolytic enzymes was proven in routine tests during 4 weeks. Lyophilization of nutrient solution enhanced the sensitivity of the protease and chitinase assay using Gel-RBB and Chitin Remazol Brilliant Violet, respectively. Compared to the standard assay, concentrated variants showed up to five times higher chitinase and a sixfold increase in protease activity. Due to inconsistent results, no statement concerning cellulase and β-1,3-glucanase is possible. Dissolving the lyophilized residue in water prior to mixing with the enzyme substrate solution is unfavorable for detection of enzyme activity. Dialysis of nutrient solution prior to lyophilization is not essential.
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