Cyclic 1,N2-propanodeoxyguanosine adducts are formed in vitro in DNA treated with alpha-acetoxy-N-nitrosopyrrolidine or its metabolite, crotonaldehyde. However, the in vivo formation of these cyclic adducts in DNA has not been demonstrated due to the lack of a sensitive detection method. In this study, a 32P-postlabeling method specific for the detection of 1,N2-propanodeoxyguanosine adducts was developed by using the corresponding 3'-monophosphates as standards. This method was validated by using DNA modified in vitro. It was then applied for the in vivo experiments in which hepatic DNA of rats treated with N-nitosopyrrolidine (NPYR) (total dose, 1.0 mmol) in drinking water or skin DNA of Sencar mice treated topically with crotonaldehyde (1.4 mmol) was isolated and subjected to 32P-postlabeling analysis. 1,N2-Propanodeoxyguanosine adducts were detected in these DNA samples. The minimal levels of adducts from liver DNA and skin DNA detected were estimated to be approximately 0.06 and approximately 0.24 mumol/mol guanine respectively. Interestingly, a background adduct spot chromatographically indistinguishable from the 1,N2-cyclic adducts was observed in the liver DNA of untreated rats. However, no such background adduct was detected in skin DNA of mice. This method demonstrated for the first time the in vivo formation of the cyclic 1,N2-propanodeoxyguanosine adducts.

We compared the metabolism in the F-344 rat of the moderately potent esophageal carcinogen N'-nitrosonornicotine (NNN, 2'-(3-pyridyl)-N-nitrosopyrrolidine) and its weakly active homologue N'-nitrosoanabasine (NAB, 2'-(3-pyridyl)-N-nitrosopiperidine). Urine was the major pathway of excretion for both nitrosamines. The major urinary metabolites of dl-NNN resulted from 2'-hydroxylation (8.1% of the dose), 5'-hydroxylation (37.6%), and pyridine N-oxidation (10.8%). The percentages of the dose of the corresponding metabolites of dl-NAB were: 2'-hydroxylation (not detected), 6'-hydroxylation (9.8%), pyridine-N-oxidation (30.0%). Similar results were obtained when the urinary metabolites of l-NNN and l-NAB were compared. In 48 h cultures of rat esophagus, the major metabolites of [2'-14C]dl-NNN resulted from 2'-hydroxylation (47%) and to a lesser extent from 5'-hydroxylation (15%). In contrast the major metabolite of [2'-14C]dl-NAB resulted from 6'-hydroxylation (35%) with lesser amounts from 2'-hydroxylation (8%). 6'-Hydroxylation of [2'-14C]dl-NAB also exceeded 2'-hydroxylation in cultures of 3, 6, 12 or 24 h duration. Pyridine-N-oxidation was not observed in the esophagus for either nitrosamine. These results demonstrate a high degree of regiospecificity in the metabolism of these structurally related nitrosamines. Among the identified urinary metabolites the ratio of alpha-hydroxylation to N-oxidation was 4.2 for NNN and 0.3 for NAB. Among the 48 h esophageal metabolites the ratio of 2'-hydroxylation to 5'- or 6'-hydroxylation was 3.1 for NNN and 0.2 for NAB. The results also suggest a basis for the weak carcinogenicity of NAB: facile excretion as its pyridine-N-oxide and detoxification in the esophagus by 6'-hydroxylation.

Maximum plasma levels of diphenhydramine in normal sub;ects ranged from 81 to 159 ng/ml in 2 to 4 hr after single 100 mg oral doses, with an estimated half-life of 7 hr. Multiple oral doses of 50 mg four times daily for 3 days produced mean peak levels of approximately 110 ng/ml by the second day, with an estimated half-life of 6 ~r. Total amine levels were 50% to 100% higher, due to the presence of N-dealkylated metabolites. Single 100 mg oral doses of diphenhydramine also produced a progressive increase in the plasma levels of acidic metabolites over a 24 hr period, reaching values 10 times those of the maximum diphenhydramine levels. Urinary excretion of total diphenhydramine metabolites represented about 64% of the dose in single dose studies, and 49% after multiple doses. These observations are consistent with earlier studies on the metabolic disposition of labeled diphenhydramine in laboratory animals.