The heterofermentative lactic acid bacterium Oenococcus oeni requires pantothenic acid for growth. In the presence of sufficient pantothenic acid, glucose was converted by heterolactic fermentation stoichiometrically to lactate, ethanol and CO2. Under pantothenic acid limitation, substantial amounts of erythritol, acetate and glycerol were produced by growing and resting bacteria. Production of erythritol and glycerol was required to compensate for the decreasing ethanol production and to enable the synthesis of acetate. In ribose fermentation, there were no shifts in the fermentation pattern in response to pantothenate supply. In the presence of pantothenate, growing O. oeni contained at least 10.2 microM HSCoA, whereas the HSCoA content was tenfold lower after growth in pantothenate-depleted media. HSCoA and acetyl-CoA are cosubstrates of phosphotransacetylase and acetaldehyde dehydrogenase from the ethanol pathway. Both enzymes were found with activities commensurate with their function in ethanol production during heterolactic fermentation. From the kinetic data of the enzymes and the HSCoA and acetyl-CoA contents, it can be calculated that, under pantothenate limitation, phosphotransacetylase, and in particular acetaldehyde dehydrogenase activities become limiting due to low levels of the cosubstrates. Thus HSCoA deficiency represents the major limiting factor in heterolactic fermentation of glucose under pantothenate deficiency and the reason for the shift to erythritol, acetate, and glycerol fermentation.