AbstractMale C57BL/6 mice were exposed to 1% (w/w) (+)‐ or (−)‐2‐ethylhexanoic acid or an equimolar mixture of these enantiomers in their diet for 4 or 10 days. A significant increase in liver weight and a 2‐ to 3‐fold increase in the protein content of the mitochondrial fraction were seen in all cases. Peroxisomal palmitoyl‐CoA oxidation was increased 2‐ to 3.5‐fold after 4 days of treatment and 4‐ to 5‐fold after 10 days, while the corresponding increases in peroxisomal lauroyl‐CoA oxidase activity were 2‐ to 3‐fold and 9‐ to 12‐fold, respectively. Peroxisomal catalase activity was unchanged, whereas the microsomal and cytosolic activities were increased 2‐ to 3‐fold and 6‐ to 16‐fold, respectively. These treatments also induced microsomal ω‐hydroxylation of lauric acid 7‐fold and soluble epoxide hydrolase activity in the mitochondrial and cytosolic fractions, as well as microsomal epoxide hydrolase activity about 50–100%. The only significant differences observed between the effects of (+)‐2‐ethylhexanoic acid and its (−)‐enantiomer were on peroxisomal palmitoyl‐CoA oxidation and lauroyl‐CoA oxidase activity after 4 days of treatment. In both these cases the (+)‐enantiomer resulted in increases which were 50–75% gre