Four different yeast species (Metschnikowia pulcherrima, Saccharomycopsis vini, Kluyveromyces marxianus, Cryptococcus albidus), isolated from surface of grapes, were evaluated for biocontrol potential against Penicillium glabrum. In order to investigate the influence of temperature, pH, water activity and yeast cell concentration on Penicillium glabrum inhibition, the individual effects and the interaction of these factors were analyzed by means of a Central Composite Design (CCD). All yeast species tested showed antagonistic effects which were more pronounced at high cell concentrations. The other variables affected the antagonistic effect differentially depending on the yeast species. Results of the experimental design showed that the selective success of a competitive microflora is under environmental control; moreover, when microbial cells are subjected to multiple factors, the effects and the reciprocal interactions of the individual variables cannot be independently evaluated.