A dye-labelled soluble substrate for the assay of endo-chitosanase activity

Coupling Remazol Brilliant Blue R (RBB) to chitosan that has been subjected to partial enzymatic hydrolysis gave rise to a new dyed derivative of chitosan which is highly soluble in slightly acidic buffers or microbiological media. The RBB:chitosan ratio has been carefully optimized, as well as the reaction conditions, to obtain a quasi-linear, sensitive assay for chitosanase. The substrate (sRBB-C) is useful for microtitre plate-based screening and for assay of chitosanase activity in the presence of high backgrounds of reducing sugars. Due to its solubility, sRBB-C is particularly suitable for the detection of chitosanase-positive organisms on agar media.