An automatic spectrofluorimetric method for the simultaneous determination of glycerol and ethanol is presented. The method is based on the use of a flow injection (FI) manifold and dehydrogenase enzyme immobilized on controlled-pore glass. The proposed method allows for the simultaneous determination of glycerol (linear range between 1 and 10 mu g ml(-1), r = 0.9989 and s(r) <1%) and ethanol (linear range between 10 and 100 mu g ml(-1), r = 0.9998 and s(r) <0.9%). It was applied to the determination of both analytes in different wines with recoveries between 97 and 105%. The sampling frequency was 60 samples h(-1). The results obtained on eight wines were compared with those provided by conventional methods and the agreement between them corroborated the usefulness of the proposed method.1201179182