Isolation by solid-phase extraction and liquid chromatographic determination of mutagenic amines in beef extracts

A solid-phase extraction method was successfully optimized for the isolation and preconcentration of five mutagenic amines, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole, 3-amino-1-methyl-5H-pyrido[4,3-b]indole, 2-amino-9H-pyrido[2,3-b]indole, 2-amino-3-methyl-9H-pyrido[2,3-b]indole and 2-amino-1-methyl-6-phenyl-imidazo[4,5-b]pyridine, and two co-mutagens, 1-methyl-9H-pyrido[4,3-b]indole and 9H-pyrido[4,3-b]indole. Coupling of diatomaceous earth, propylsulphonyl silica gel, and octadecylsilane cartridges was used to separate selectively the imidazopyridine and indolpyridine derivatives from those of quinoxaline and quinoline. A method based on this sample preparation was applied to the determination of twelve heterocyclic amines and related substances in a commercial beef extract using HPLC with electrochemical and fluorescence detection. Good recovery values were obtained, ranging between 55 and 99%. The co-mutagens 1-methyl-9H-pyrido[4,3-b]indole (harman) and 9H-pyrido[4,3-b]indole (norharman) were found in the beef extract at levels of 110 and 53 ng g-1, respectively, and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) and 2-amino-9H-pyrido[2,3-b]indole (A alpha C) were tentatively identified.