1. Isoprene is metabolised in vitro by oxygenation of either double bond to 2-ethenyl-2-methyloxirane (epoxide A) and 2-(1'-methylethenyl)oxirane (epoxide B). The reactivity in vitro and formation in vivo of the monoepoxides of isoprene were studied by the formation of adducts to N-terminal valines in haemoglobin (Hb). These adducts were analysed by mass spectrometry after cleavage and derivatization by a modified Edman degradation method. 2. When red blood cells were incubated with commercial isoprene oxide (about 95% epoxide A, < or = 5% epoxide B) adducts from both epoxides were formed. 3. It is confirmed that epoxide A is hydrolysed much faster than epoxide B. The rates are enhanced by phosphate buffer (epoxide A), probably through acid catalysis, and by the presence of red blood cells (both epoxides), due to enzymatic detoxification. 4. Comparison of total valine adduct levels in Hb from isoprene and isoprene oxide injected i.p. led to the conclusion that 23 and 1% of injected isoprene was metabolized to the epoxides in mouse and rat, respectively.