CAPILLARY ZONE ELECTROPHORESIS FOR THE QUANTITATION OF OLIGOSACCHARIDES FORMED THROUGH THE ACTION OF CHITINASE

Capillary zone electrophoresis with fluorescence detection was used to analyze the products formed by chitinase acting on N-acetylchitooligosaccharide-fluorescent conjugates. Six oligosaccharides of the structure [N-acetylglucosamine(1 --> 4)]n (where n = 1-6) were conjugated to 7-amino-1,3-naphthalene disulfonic acid by reductive amination. Each oligosaccharide-fluorescent conjugate was purified by preparative gradient polyacrylamide gel electrophoresis, semi-dry electrotransfer to a positively-charged nylon membrane and recovered by washing the membrane with salt solution. The products formed by treating each oligosaccharide-fluorescent conjugate with chitinase were analyzed by capillary zone electrophoresis. The chitinase treatment hexasaccharide-fluorescent conjugate was also examined kinetically to study the action pattern of this enzyme.