ESTABLISHMENT OF A SALMONELLA TESTER STRAIN HIGHLY SENSITIVE TO MUTAGENIC HETEROCYCLIC AMINES

Heterocyclic amines (HCAs) that are present in cooked foods require me tabolic activation to exert their genotoxicity. They undergo activatio n via N-hydroxylation by cytochrome P450 1A2 (CYP1A2), followed by O-e sterification by O-acetyltransferase (OAT). To develop a Salmonella te ster strain that is highly sensitive to mutagenic HCAs, we introduced a coexpression plasmid (p1A2OR) carrying human CYP1A2 and NADPH-CYP re ductase cDNAs and an expression plasmid (pOAT) carrying Salmonella OAT to Salmonella typhimurium TA1538 to yield a TA1538/ARO strain. The TA 1538/ARO strain was proven to express the enzymes, as indicated by hig h activities of 7-ethoxyresorufin O-deethylase and isoniazid N-acetyla se. The TA1538/ARO strain exhibited very high sensitivity to mutagenic HCAs 2-amino-3,4-dimethylimidazo[4,5-f]quinoline, 2-amino-3-methylimi dazo[3,5-f]quinoline (IQ), and 2-amino-3,8-dimethylimidazo[4,5-f]quino xaline and a somewhat higher sensitivity to 2-amino-1-methyl-6-phenyli midazo[4,5-b]pyridine compared with the parent Ames tester strain TA15 38. The minimum concentrations of 2-amino-3,3-dimethylimidazo[4,5-f]qu inoline, IQ, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, and 2-amin o-1-methyl-6-phenylimidazo[4,5-b]pyridine giving positive results were defined by evidence that the number of colonies increased in a dose-d ependent manner and reached a number two times higher than that obtain ed by vehicle alone as a control in the TA1538/ARO strain at concentra tions of 0.3, 3, 30, and 1000 pM, respectively. When the membrane and cytosol fractions prepared from TA1358/ARO were added to a mixture con taining the parental TA1538, the sensitivity of TA1538 to IQ was much lower than that seen with TA1538/ARO. These results indicate that the intracellular expression of drug-metabolizing enzymes makes the establ ished strain of Salmonella highly sensitive to mutagenic HCAs.