Evaluation of a chromogenic chito-oligosaccharide analogue, p-nitrophenyl-beta-D-N,N'-diacetylchitobiose, for the measurement of the chitinolytic activity of bacteria.

Three methods of quantifying chitinase activity were compared. The activities of crude chitinases of 10 bacterial isolates from different environments were estimated in terms of (1) the release of p-nitrophenol from the chromogenic chito-oligosaccharide analogues, p-nitrophenyl-beta-D-N,N'-diacetylchitobiose, p-nitrophenyl-N-acetyl-beta-D-glucosamine and p-nitrophenyl-beta-D-N,N',N"-triacetylchitotriose, (2) the release of reducing sugars from chitin and (3) the formation of clearing zones on chitin agar. When crude chitinase from Bacillus pabuli was used the hydrolysis of p-nitrophenyl-beta-D-N,N'-diacetylchitobiose correlated well with the release of reducing sugars from chitin and the formation of clearing zones on chitin agar. However, when the activity of crude chitinases from the different bacterial isolates were compared no agreement was found between the hydrolysis of p-nitrophenyl-beta-D-N,N'-diacetylchitobiose and the release of reducing sugars from chitin or the formation of clearing zones on chitin agar. It was concluded that the assay with chromogenic p-nitrophenyl chito-oligosaccharide analogues is not well suited for studies that compare the chitinase activity of different bacteria.