Using a combination of gas chromatography, mass spectrometry, and selected ion recording techniques, we have identified nicotine and its major metabolite, continine, in the breast fluid of nonlactating women smokers. As little as 25 picograms could be measured by using the deuterated variants, [5',5'-2H]nicotine and [3,3-2H]cotinine, both as internal standards and as carriers in an inverse isotope dilution method.