Abstract A systematic study of the bioreduction of ethyl 3-oxobutyrate by Saccharomyces cerevisiae under aerobic and anaerobic conditions was performed by using flow isothermal microcalorimetry. In association with this technique, other properties directly related to the metabolism of S. cerevisiae such as consumption of O 2, glucose and pH were also studied. The metabolic aspects of the bioreduction, such as compartmentalization and preferential use of the cofactors (NADH or NADPH), are discussed based on heat and ethanol production, and oxygen and glucose consumption data obtained during a period of 20 h. These data allows us to suggest the compartment in the microorganism cell where the bioreduction can occur for each experimental condition. The obtained results indicate that under aerobic conditions, the process occurs preferentially in the mitochondrial matrix and is associated with the consumption of the cofactor NADH which is regenerated by the respiratory pathway. However, under anaerobic conditions, the bioreduction occurs in the cytosol and is associated with the consumption of the cofactor NADPH, which is regenerated by the pentose phosphate pathway.