Plasma free acetaldehyde levels were measured by an improved method in baboons fed ethanol chronically and in pair-fed controls during an intravenous infusion of ethanol. After an appropriate loading dose, ethanol was infused at the rate of its elimination to achieve a steady state at one of three different blood ethanol levels (50 ± 10 mM, 10 ± 2 mM, or 5 ± 1 mM). The rate of production of acetaldehyde was calculated from the rate of ethanol elimination by subtracting losses into urine and expired air. Liver mitochondrial aldehyde dehydrogenase (AlDH) activity was measured in surgical biopsy samples with 50 μM acetaldehyde as substrate. Chronic ethanol administration resulted in both higher plasma free acetaldehyde levels and faster acetaldehyde production at each level of blood ethanol. When the blood level of ethanol was increased from 5 to 50 mM, the level of plasma free acetaldehyde also rose in both groups of animals. The rate of acetaldehyde production, however, increased only in alcohol-fed baboons. Plasma free acetaldehyde had a significant positive correlation with production rate of acetaldehyde ( r = 0.69) and a significant negative correlation with liver mitochondrial AIDH specific activity ( r = −0.59). When these two parameters were combined (acetaldehyde production rate/AIDH activity), a correlation coefficient of 0.84 resulted, suggesting that, in addition to increased production, decreased catabolism may contribute to the higher acetaldehyde levels seen after chronic consumption.