The metabolism of trans-4-hydroxy-2-hexenal (t-4HH) was compared with that of propionaldehyde (PAL) and trans-2-hexenal (t-2H). Kinetic experiments with hepatic cytosol as a source for aldehyde dehydrogenase(s) (ALDH) (E.C. 1.2.1.3) resulted in linear Lineweaver-Burk plots for both t-4HH and t-2H, while PAL produced biphasic plots. Values of 43 and 3 microM were obtained for the apparent Km values for t-4HH and t-2H, respectively, with the two apparent Kms for PAL being 5 microM and 0.8 mM. DEAE-cellulose chromatography of cytosol isolated two peaks capable of oxidizing PAL, but only one peak was able to metabolize t-4HH. Unlike cytosol, mitochondria produced biphasic Lineweaver-Burk plots for t-4HH with two estimated Kms of 52 microM and 0.2 mM. The majority of activity for the oxidation of t-4HH, like PAL, was localized in the cytosolic fraction. This study indicates that ALDH(s) may play an important role in the detoxification of the pyrrolizidine alkaloid metabolite t-4HH, as well as other 4-hydroxyalkenals that arise from membrane lipid peroxidation.