Syntheses of modified 2-chloro-4-nitrophenyl β-maltopentaosides as useful substrates for assay of human alpha amylase

Abstract Twenty-three novel 2-chloro-4-nitrophenyl β- d -maltopentaosides modified at the 6 5 and/or 4 5 position were synthesized as substrates for human alpha amylase. Two human alpha amylases hydrolyzed 6 5 -deoxy-6 5 -, 6 5 - O -, and 4 5 ,6 5 -di- O -substituted derivatives at essentially a single d -glucosidic linkage, but 4 5 ,6 5 - O -bridged and 4 5 - O -substituted derivatives were hydrolyzed at two or more linkages. The amylases displayed smaller K m values for the compounds having hydrophobic modifications. In these derivatives, 2-chloro-4-nitrophenyl O -(6-bromo-6-deoxy-α- d -glucopyranosyl)-(1 → 4)-tris[ O -α- d -glucopyranosy-(1 → 4)]-β- d -glucopyranoside ( 10 ), 2-chloro-4-nitrophenyl O -(6-azido-6-deoxy-α- d -glucopyranosyl)-(1 → 4)-tris[ O -α- d -glucopyranosyl-(1 → 4)]-β- d -glucopyranoside ( 19 ), and 2-chloro-4-nitrophenyl O -[6- O -( N -isopropyl)carbamoyl-α- d -glucopyranosyl]-(1 → 4)-tris[ O -α- d -glucopyranosyl-(1 → 4)]-β- d -glucopyranoside ( 30 ), which were rapidly hydrolyzed by the two amylases at a limited position at an approximately equal rate, were shown to be very useful blocked-type substrates for assay of human alpha amylase.