The work consists of an experimental analysis to evaluate the effects of the variables temperature (T), pH, agitation rate (K) and initial lactose concentration (L) on the batch fermentation process of Ricotta Cheese Whey (RCW) into bio-ethanol by using the yeast Kluyveromyces marxianus. A central composite design, constituted by 26 runs, has been carried out, and the effects of the parameters have been evaluated. Eventually, once eliminated the negligible effects, Response Surface Methodology (RSM) has been applied to optimize the four parameters values in RCW fermentation process. After a preliminary experimental analysis, the chosen factors values were 32° and 40°C for T, 4 and 6 for pH, 100 and 300 rpm for K, 40 and 80 g L-1; for L. Best operating conditions resulted to be T = 36°C, pH = 5.0, K = 200 rpm and L = 60 g L-1. Copyright © 2010 AIDIC Servizi S.r.l.