Metabolism of the carcinogen N-hydroxy- N-2-fluorenylacetamide in germ-free rats

Abstract

The question whether the carcinogen N-hydroxy- N-2-fluorenylacetamide (N-OH-FAA) was affected by the microbial flora in the intestinal tract was investigated by comparing the metabolism of this compound in germ-free and conventional Fischer strain rats, employing isotopic techniques. Although the amounts of total urinary glucosiduronic acids and sulfuric acid esters were similar in axenic and control rats after an i.p. dose of N-OH-FAA, there were important differences among the individual glucosiduronic acids. Germ-free rats excreted considerably larger amounts of the glucuronide of N-OH-FAA and appreciably less of the conjugates from the ring-hydroxylated metabolites. Furthermore, the cecal and fecal metabolites in the conventional rats were mostly free, unconjugated materials, wherease in axenic rats the major fraction was conjugated with sulfuric and glucuronic acid. Injection of N-OH-FAA into the cecum, or intraluminal administration of glucosiduronic acids of N-OH-FAA or of phenolphthalein showed that these materials could be absorbed readily from the gastrointestinal tract. In the cecum of germ-free rats, β-glucuronidase activity was low and had an optimum at a pH characteristic of mammalian enzyme, but in conventional rats it was higher and with a pH optimum more like that of the bacterial enzyme. The data indicate that N-OH-FAA is metabolized differently in germ-free rats: (1) because they lack the bacterial flora of the intestinal tract in conventional rats, which can hydrolyze glucosiduronic acids excreted into the gut by bile; and (2) because N-OH-FAA is a substrate for a bacterial N-dehydroxylase, also absent in axenic rats. In part, the enterohepatic cycle undergone by N-OH-FAA may be related to the liberation in the gut of readily absorbed metabolites, which are then further modified and excreted in urine or in bile.