Azo mediators and methods of use thereof

A detection reagent for determining presence or concentration of glucose, the detection reagent comprising:

• (a) a flavine-, nicotinamide- or pyrroloquinoline quinone-dependent oxidoreductase;
• (b) a reducible flavine, nicotinamide or pyrroloquinoline quinone coenzyme;
• (c) an azo compound of the general formula (I):
• wherein R1 denotes a residue selected from the group consisting of OH, OR7, SR7, NHR7 and NR7R8;
• wherein R2, R3, R4 and R5 independently of one another denote a residue selected from the group consisting of H, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl, heteroaryl, CN, COO−, halogen, O(alkyl), O(aryl), NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NO2 and SO3−, and wherein the alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl, heteroaryl, O(alkyl), O(aryl), NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl) or N(aryl)2 can contain one or more substituents selected from the group consisting of CN, COO−, halogen, NH2, NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NH—C(═NH2)—NH2+, NO2, OH, O—(CH2CH2)n—OH, O—(CH2CH2)n—O(alkyl), O(alkyl), O(aralkyl), O(aryl), OPO32−, PO32− and SO3−;
• wherein R6 denotes a residue selected from the group consisting of CN, C(═X)NH2, C(═X)NHR9, C(═X)NR9R10, C(═X)NHOH and C(═X)R9;
• wherein R7 and R8 independently of one another denote a residue selected from the group consisting of alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl and heteroaryl, and wherein the alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl or heteroaryl can contain one or more substituents selected from the group consisting of CN, COO−, halogen, NH2, NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NH—C(═NH2)—NH2+, NO2, OH, O—(CH2CH2)n—OH, O—(CH2CH2)n—O(alkyl), O(alkyl), O(aralkyl), O(aryl), OPO32−, PO32− and SO3−;
• alternatively wherein R7 and R8 together with the N atom to which they are bound form a 5-membered to 7-membered heterocycle, and wherein the heterocycle can contain a further heteroatom selected from the group consisting of N, O and S and/or one or more substituents selected from the group consisting of CN, COO−, halogen, NH2, NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NH—C(═NH2)—NH2+, NO2, OH, O—(CH2CH2)n—OH, O—(CH2CH2)n—O(alkyl), O(alkyl), O(aralkyl), O(aryl), OPO32−, PO32− and SO3−;
• wherein R9 and R10 independently of one another denote a residue selected from the group consisting of alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl and heteroaryl, and wherein the alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heterocycloalkyl or heteroaryl can contain one or more substituents selected from the group consisting of CN, COO−, halogen, NH2, NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NH—C(═NH2)—NH2+, NO2, OH, O—(CH2CH2)n—OH, O—(CH2CH2)n—O(alkyl), O(alkyl), O(aralkyl), O(aryl), OPO32−, PO32− and SO3−;
• alternatively wherein R9 and R10 together with the N atom to which they are bound form a 5-membered to 7-membered heterocycle, and wherein the heterocycle can contain a further heteroatom selected from the group consisting of N, O and S and/or one or more substituents selected from the group consisting of CN, COO−, halogen, NH2, NH(alkyl), N(alkyl)2, N(alkyl)3+, NH(aryl), N(aryl)2, NH—C(═NH2)—NH2+, NO2, OH, O—(CH2CH2)n—OH, O—(CH2CH2)n—O(alkyl), O(alkyl), O(aralkyl), O(aryl), OPO32−, PO32− and SO3−;
• wherein X denotes a heteroatom selected from the group consisting of O and S, and wherein n denotes an integer from 1 to 6;
• and
• (d) optionally a reducible optical indicator.