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Cheese whey, a by-product of dairy industry that usually is disposed without any treatment, represents a valuable source of carbohydrate for fermentation. Its utilization along with rice bran, a by-product derived from the rice milling process, rich in nutrients, represents an alternative for the reduction of industrial cost in bio-ethanol production. Thus, the valorisation of agro-industrial by-products in bioprocess is of large interest for economic and environmental reasons. In the present work, five Kluyveromyces yeast strains were evaluated on their capability to produce bio-ethanol in a medium containing permeated whey at an initial lactose concentration of 100 g/L. This medium was supplemented with rice bran, inoculated to achieve an initial cell concentration of 0.5 g/L and incubated under micro-aerobic conditions at 30 oC for 72h. The maximum yields achieved were 0.40 g/g and 0.30 g/g for Kluyveromyces marxianus UFV-3 and Kluyveromyces marxianus CCT 4086, respectively. Since K. marxianus UFV-3 presented a promising potential for bio-ethanol production, a 2³ full-factorial composite design and response surface methodology was applied to determine the effect of initial pH (4.0 - 7.0), lactose concentration (50 - 150 g/L), and rice bran concentration (10 - 100 g/L) in the production of ethanol. Therefore, the maximum ethanol concentration (61 g/L), yield (0.50 g/g) and productivity (1.27 g/L/h) were attained after 48 h of fermentation, in the medium with an initial lactose concentration of 150 g/L, at pH 7.0 supplemented with 100 g/L of rice bran. A positive correlation was observed when cheese whey was supplemented with rice bran and used as medium for bio-ethanol.