A study of the substrate specificity and transfer of electrons for the conversion of very long fatty acyl-CoA to hydrocarbon and CO 2 by a cytochrome P450 (P450hyd) type enzyme in microsomal preparations from abdominal epidermal tissue of the house fly, Musca domestica, showed that the enzyme which converts aldehyde to hydrocarbon contributes little to the chain length of the hydrocarbon products observed in vivo. NADPH (and not NADH) was required for the reduction of the fatty acyl-CoA to the corresponding aldehyde, and when both cofactors were added together, the amount of hydrocarbon formed from the acyl-CoA derivative was about the same as with NADPH alone. Both NADPH and NADH (less effectively) supported the conversion of the aldehyde to hydrocarbon and CO 2 by P450hyd. When both pyridine nucleotides were added together with the aldehyde substrate, conversion to hydrocarbon was intermediate between the levels formed with either cofactor alone, suggesting that the same electron carrier was transferring electrons from both NADH and NADPH and that cytochrome b 5 does not participate in the reaction. Antibody to house fly cytochrome P450 reductase inhibited the reaction when either NADPH or NADH was used. In addition, both NADP + and 2-AMP stimulated, rather than inhibited, hydrocarbon production in microsomes from female houseflies, presumably by inhibiting competing P450 enzymes which are supplied with electrons by the NADPH-cytochrome P450 reductase. These data suggest a novel mode of electron transfer to the P450hyd.