Metabolism of N '-nitrosonornicotine enantiomers by cultured rat esophagusand in vivo in rats

People who use tobacco products are exposed to considerable amounts of N'-n itrosonornicotine (NNN), a well-established esophageal carcinogen in rats. NNN is believed to play a significant role as a cause of esophageal and ora l cavity cancer in smokers and snuff dippers, The carcinogenicity ofNNN is dependent on its metabolic activation. However, virtually all studies carri ed out to date on NNN metabolism have used racemic material. In this study, we examined the metabolism of [5-H-3]-(S)-NNN and [5-H-3]-(R)-NNN in cultu red rat esophagus and in vivo in rats. Cultured rat esophagus metabolized ( S)-NNN (1 mu M) predominantly to products of 2'-hydroxylation, 4-oxo-4-(3-p yridyl)butanoic acid (keto acid) and 4-hydroxy-1-(3-pyridyl)-1-butanone (ke to alcohol). In contrast, the major metabolite of (R)-NNN under these condi tions was 4-hydroxy-4-(3-pyridyl)butanoic acid (hydroxy acid), a product; o f NNN 5'-hydroxylation. The 2'-hydroxylation:5'-hydroxylation metabolite ra tio ranged from 6.22 to 8.06 at various time intervals in the incubations w ith (S)-NNN, while the corresponding ratios were 1.12-1.33 in the experimen ts with (R)-NNN. These differences were statistically significant (P < 0.00 1). Since 2'-hydroxylation is believed to be the major metabolic activation pathway of NNN in the rat esophagus, the results demonstrate that (S)-NNN is metabolically activated more extensively than (R)-NNN in this tissue, an d therefore may be more carcinogenic. Rats were treated with 0.3 mg/kg of [ 5-H-3]-(R)-NNN, [5-H-3]-(S)-NNN, or racemic [5-H-3]NNN by gavage, and the u rinary metabolites were analyzed. The major metabolites were hydroxy acid a nd keto acid. As in the in vitro studies, products of 2'-hydroxylation pred ominated in the urine of the rats treated with (S)-NNN while products of 5' -hydroxylation were more prevalent in the rats treated with (R)-NNN. 2'-Hyd roxylation:5'-hydroxylation metabolite ratios ranged from 1.66 to 2.04 in t he urine at various times after treatment with (S)-NNN, while the ratios we re 0.398-0.450 for the rats treated with (R)-NNN (P < 0.001). The results o f this study provide new insights into NNN metabolism in rats and suggest t hat the carcinogenicity of (S)-NNN, the predominant enantiomer in tobacco p roducts, may be greater than that of (R)-NNN or racemic NNN.