Enzymic catalysis of the accumulation of acetaldehyde from ethanol in human prenatal cephalic tissues: Evaluation of the relative contributions of CYP2E1, alcohol dehydrogenase, and catalase/peroxidases

Background: The human prenatal brain is very sensitive to the toxic effects of ethanol, but very little information is available concerning the conver sion of ethanol to the highly cytotoxic metabolite, acetaldehyde, in that o rgan. Thus, experiments were designed to investigate rates of accumulation of acetaldehyde from ethanol in the prenatal human brain. Methods: Prenatal human cephalic tissue homogenates were used as enzyme sou rces and were compared with analogous preparations of adult rat livers. Gen erated acetaldehyde was derivatized with cyclohexane-1,3-dione to yield flu orescent decahydroacrizine-1,8-dione, which was readily separated, detected , and quantitated with HPLC. Results: Detected rates of accumulation were unexpectedly high, even in the absence of added NADPH, NAD(+), or H2O2, which are cofactors/cosubstrates for cytochrome P-450-, alcohol dehydrogenase- and catalase/peroxidase-catal yzed reactions, respectively. Without added cofactors/cosubstrates or other components and under linear reaction conditions, rates in human prenatal c ephalic preparations were approximately 20% of those observed with analogou s preparations of adult rat livers. Cofactor/cosubstrate-independent reacti ons were localized in the cytosolic (soluble) fraction and were strongly de pendent on molecular oxygen (O-2). They were not inhibited substantially by carbon monoxide (CO:O-2 = 80:20 vs N-2:O-2 = 80:20) or by pyrazole in conc entrations up to 10 mM and were only weakly inhibited by azide. Preincubati ons with excess catalase did not result in decreased activity. Reactions ex hibited substrate saturation and heat inactivation indicating enzymic catal ysis. Conclusions: Experiments indicated a relatively rapid accumulation of aceta ldehyde from ethanol in human prenatal brain tissues and suggested that the observed cofactor/cosubstrate-independent reactions were largely independe nt of P-450 cytochromes, alcohol dehydrogenases, or catalase/peroxidases. R esults were consistent with catalysis by an as yet unidentified cytosolic o xidase(s).