Other relevant data Causal associations have been clearly established between active smoking and adverse reproductive outcomes and numerous non-neoplastic diseases, including chronic obstructive pulmonary disease and cardiovascular diseases. Tobacco smoking is addictive, and nicotine has been established as the major addictive constituent of tobacco products. Measurement of the nicotine metabolite, cotinine, in human blood, urine or saliva provides a specific and sensitive test for exposure to tobacco smoke and can be used to distinguish active and passive smokers from nonsmokers. Active smoking raises the concentrations of carbon monoxide, benzene and volatile organic compounds in exhaled air. The concentrations of urinary metabolites of some important tobacco smoke carcinogens and related compounds are consistently higher in smokers than in nonsmokers. These include metabolites of benzene, a known carcinogen in humans, as well as metabolites of several carcinogens that cause lung tumours in rodents. Covalent binding to blood proteins by carcinogens present in tobacco smoke has been demonstrated to occur at significantly higher levels in smokers than in nonsmokers. The adducts are derived from various compounds including aromatic amines (e.g. 4-aminobiphenyl), polycyclic aromatic hydrocarbons (e.g. benzo[a]pyrene), tobacco-specific nitrosamines (e.g. 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone), benzene, acrylamide and acrylonitrile. Smoking-related DNA adducts have been detected by a variety of analytical methods in the respiratory tract, urinary bladder, cervix and other tissues. In many studies the levels of carcinogen-DNA adducts have been shown to be higher in tissues of smokers than in tissues of nonsmokers. Some but not all studies have demonstrated elevated levels of these adducts in the peripheral blood and in full-term placenta. Smoking-related adducts have also been detected in cardiovascular tissues. Collectively, the available biomarker data provide convincing evidence that carcinogen uptake, activation and binding to cellular macromolecules, including DNA, are higher in smokers than in nonsmokers. The exposure of experimental animals, primarily rodents, to mainstream tobacco smoke results in a number of biological effects that include (i) increases or decreases in the activities of phase I and phase II enzymes involved in carcinogen metabolism, (ii) increases in the activation of antioxidant enzymes, (iii) increased expression of nitric oxide synthase and of various protein kinases and collagenase, (iv) the formation of tobacco smoke-related DNA adducts in several tissues and (v) reduced clearance of particulate material from the lung. Smoking is known to have inhibitory or inducing effects on the activities of many enzymes in human tissues. These include xenobiotic metabolizing enzymes, which affect drug and carcinogen metabolism. Numerous studies have reported effects on enzymes in cells treated in culture with tobacco smoke or tobacco smoke condensates. In humans, smoking produces gene mutations and chromosomal abnormalities. Urine from smokers is mutagenic. Relative to nonsmokers, lung tumours of smokers contain higher frequencies of TP53 and KRAS mutations, and the spectrum of mutations has unique features. Most of the genetic effects seen in smokers are also observed in cultured cells or in experimental animals exposed to tobacco smoke or smoke condensate. Tobacco smoke is genotoxic in humans and in experimental animals.