Int. J. Environ. Res. Public Health 2010, 7 1078 2. Historical Debate on the Identification of Enzyme(s) in the So-Called ‘Non-ADH’ Pathway* of Alcohol Metabolism Animal experiments using pyrazoles, which are specific inhibitors of ADH, have left no doubt that ADH1 is the main enzyme responsible for alcohol metabolism in the body [1]. However, alcohol continues to be metabolized to some extent even after the administration of pyrazoles; this is called the non-ADH pathway [2,3]. This pyrazole-insensitive pathway has a greater metabolic role when the level of blood alcohol is high or when the intake of alcohol is chronic [2-7]. Recently, we used ADH1-null mice to show that ADH1 accounts for about 70% of systemic alcohol metabolism [25,26], which means that the non-ADH1 pathway accounts for the remaining 30%. Regarding the identity of the main enzyme responsible for this pathway, a heated scientific debate has continued for three decades over whether it is catalase [10-12] or MEOS, which is mainly composed of CYP2E1 [2-9]. Indeed, both enzymes seem to be responsible for the non-ADH1 pathway, because both exhibit ethanol-oxidizing activity with a Km for ethanol (around 10 mM [7,10]) that is higher than that for ADH1, and both are insensitive to pyrazoles in vitro [2-6]. Furthermore, the induction of MEOS activity due to chronic alcohol consumption seems to explain the accelerated rate of alcohol metabolism observed in chronic drinkers [3,5-7]. However, it has recently been demonstrated that there is no difference in the elimination rate for blood ethanol between CYP2E1-null mice and wild-type mice, even after the chronic administration of ethanol [27]. The role of catalase in systemic alcohol metabolism has not been clarified in vivo either, even though mice genetically lack this enzyme [28]. Since there is no in-vivo evidence for either MEOS or catalase, the debate still has not reached a conclusion. Furthermore, the first-order kinetics of the elimination of blood alcohol has been observed at very high concentrations [29-31], which cannot be explained by the Km of either MEOS [7] or catalase [10]. These data suggest the involvement of a very high-Km enzyme in the non-ADH pathway. Many investigators have reported that the total ADH activity of liver correlates with the rate of systemic alcohol metabolism [1]. Mammalian livers are known to contain two ADH isozymes other than ADH1, namely, ADH2 (Class II) and ADH3 (Class III), both of which have a higher Km for ethanol than ADH1 [32]. These data suggest a possible role for these high-Km ADHs in the non-ADH1 pathway of alcohol metabolism. 3. Discovery of ADH3 (Class III ADH) In 1981, X. Pares and B. L. Vallee reported a new form of human liver ADH with unique kinetic characteristics [33] and classified it as Class III ADH (ADH3) because it was the third isozyme of mammalian ADH to be discovered [15]. However, two years earlier in 1979, at a meeting on alcohol medicine in Japan [13], we reported that we had found a new type of ADH with an acidic isoelectric point in mouse liver that had a very high Km for ethanol and was insensitive to pyrozoles. The new mouse ADH corresponded to an electrophoretic ADH band previously designated as ADHB2 on a gel [14,34] and was later also classified as Class III ADH (ADH3) [14,33]. Around that time, many investigators considered ADH3 to have a negligible role in alcohol metabolism because it exhibits only very low activity towards ethanol due to its very high Km when measured in vitro by the standard ADH