Attenuated total reflectance infrared spectroscopy (ATR-FTIR) combined with soft independent modeling of class analogy (SIMCA) was used to study the physiological state (exponential and stationary) of three commercial strains of Saccharomyces cerevisiae (ES454, E491, and ES181) fermented in Grenache blanc and Chardonnay musts. Microvinifications were performed with 150 mL of cleaned must at 17 °C. Fermented juices at exponential and stationary phase were centrifuged, and the pellets obtained were placed onto ZnSe crystal. Spectra were collected in the attenuated total reflectance (ATR) mode in the mid-infrared region (4000–700/cm) and were analyzed by a multivariate analysis technique, SIMCA. The chemical differences detected between S. cerevisiae strains at exponential and stationary phases were mainly related to differences in their cell wall composition. ATR-FTIR combined with multivariate technique was a rapid and simple method to study psychological states of S. cerevisiae during wine fermentation.