A gas chromatographic/mass spectrometric method for analysis of nicotine in scalp hair and measurements of nicotine in hair from 20 smokers and non-smokers with a one year interval are presented. The gas chromatographic/mass spectrometric method, based on selected ion monitoring of the ion m/z 162, with ions m/z 133 and 84 as qualifier ions, allows a highly selective and sensitive detection of nicotine in hair samples avoiding interference from other chemicals present in hair. Nicotine is extracted after alkaline dissolution i 5M NaOH. The sampling procedure takes into account that significant amounts of nicotine in hair is probably adsorbed and deposited from atmospheric air. In consequence, the hair samples must be taken from a site with sufficient contact with surrounding air and the measured concentration of nicotine must be related to the length of the hair segment and its distance from the scalp. Our study demonstrated very clear and statistically significant differences in the concentrations of nicotine in hair from smokers and non-smokers, a high reproducibility of hair nicotine measurements over time, and that the presented method is sensitive enough to detect individual changes not only in smoking habits, but also in environmental nicotine exposure.