The ratio of cotinine to 3′-trans-hydroxycotinine is a validated probe of CYP2A6 activity and nicotine clearance [14]. CYP2A6 is responsible for nearly all in vivo cotinine hydroxylation[39], while other enzymes contribute modestly to the conversion of nicotine to cotinine [40, 41]. The far longer half-lives of cotinine and 3′-trans-hydroxycotinine compared to nicotine also make their ratio a stable measure amenable to interpretation during ad libitum smoking[13]. However, neither the 3′-trans-hydroxycotinine/cotinine ratio following oral administration [14], nor the ad libitum 3′-trans-hydroxycotinine/(cotinine + nicotine) ratio [13] were previously reported to be strong enough predictors of CYP2A6 genotype to use in developing a predictive model of CYP2A6 activity. Furthermore, it is the metabolism of nicotine that is most relevant to models of smoking phenotypes. An important disadvantage of an oral nicotine administration experiment is that this is not a smoker’s typical route of absorption. However, oral nicotine metabolism has been demonstrated to be a worthwhile probe of CYP2A6 activity [14, 42] and it is not unreasonable to assert that the relative impact of different CYP2A6 alleles on first-pass metabolism is likely to translate
