A method for measurement of nanogram quantities of 3-methylcholanthrene in stool samples

The carcinogen 3-methylcholanthrene can be produced from deoxycholic acid and is postulated by some investigators to play a role in the pathogenesis of colon carcinoma. The small quantities of this compound which could be carcinogenic have been difficult to measure in feces because of many potentially interfering compounds. Using 3-[6-14C]methylcholanthrene as an internal standard, petroleum ether extraction, C-18 SepPak separation, preparative high performance liquid chromatography, and gas-liquid chromatography-mass spectrometry with selected ion monitoring, we developed an assay capable of detecting less than 35 ng of 3-methylcholanthrene per gram of stool. Application of this technique to stools of five patients with colon carcinoma and two normal controls revealed no detectable 3-methylcholanthrene in any stool sample. This negative result was confirmed by incubating radiolabeled cholic acid in fecal homogenates. Although greater than 90% of this radiolabeled bile acid was converted to deoxycholic acid, none of the radioactivity was found in the thin-layer chromatography fraction corresponding to 3-methylcholanthrene. These observations provide evidence against a role for 3-methylcholanthrene in pathogenesis of human colon carcinoma. Similar assays could be used for analysis of other carcinogens in stool samples.