ANALYSIS OF HUMAN URINE FOR PYRIDINE-N-OXIDE METABOLITES OF 4-(METHYLNITROSAMINO)-1-(3-PYRIDYL)-1-BUTANONE, A TOBACCO-SPECIFIC LUNG CARCINOGEN

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a potent pulmo nary carcinogen in rodents and is believed to be a causative factor fo r lung cancer in smokers, NNK also may be involved in oral cancer etio logy in users of smokeless tobacco products, Pyridine-N-oxidation of N NK and its major metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-but anol (NNAL), produces NNK-N-oxide and NNAL-N-oxide, respectively, whic h are detoxification products of NNK metabolism and are excreted in th e urine of rodents and primates, Our goal is to develop a panel of uri nary biomarkers to assess the metabolic activation and detoxification of NNK in humans, In this study, we developed methodology to analyze h uman urine for NNK-N-oxide and NNAL-N-oxide, The key step in the metho d was conversion of the N-oxides to NNK and NNAL by treatment with Pro teus mirabilis, The resulting samples were then analyzed essentially b y methods that we have described previously, Methylnitrosamino)-4-(3-p yridyl-N-oxide)-1-butanol (iso-NNAL-N-oxide) was used as internal stan dard, Levels of NNAL-N-oxide in smokers' urine ranged from 0.06 to 1.4 pmol/mg creatinine, mean +/- SD 0.53 +/- 0.36 pmol/mg creatinine, Its presence was confirmed by high performance liquid chromatography-elec trospray ionization-tandem mass spectrometry, NNK-N-oxide was not dete cted in smokers' urine, Levels of NNAL-N-oxide in the urine of smokele ss tobacco users ranged from 0.02 to 1.2 pmol/mg creatinine, mean +/- SD 0.41 +/- 0.35 pmol/mg creatinine. The amounts of NNAL-N-oxide in ur ine were less than 20% of those of no)-1-(3-pyridyl)but-1-yl]-beta-O-D -glucosiduronic acid (NNAL-Gluc) and were approximately 50% as great a s those of free NNAL, These results demonstrate that pyridine-N-oxidat ion is a relatively minor detoxification pathway of NNK and NNAL in hu mans, The method was applied to analysis of urine from 11 smokers who consumed a diet containing watercress, In an earlier study (S. S. Hech t et al., Cancer Epidemiol., Biomarkers & Prev., 4: 877-884, 1995), we showed that consumption of watercress, a source of phenethyl isothioc yanate (PEITC), caused an increase in urinary excretion of NNAL plus N NAL-Gluc, This was attributed to inhibition of alpha-hydroxylation of NNK by PEITC, as seen in rodents in which PEITC also inhibits the pulm onary carcinogenicity of NNK, However, PEITC also could have inhibited pyridine-N-oxidation of NNK and NNAL, The urine of these smokers was analyzed for NNAL-N-oxide, The results demonstrated that watercress co nsumption had no effect on levels of NNAL-N-oxide in urine, supporting the conclusion that PEITC does inhibit the metabolic activation of NN K in humans.