Wheat bran (WB), Soybean meal (SM) and Cottonseed meal (CSM) were applied together as substrates in solid fermentation for lipase production by a new Penicillium sp. A simplex centroid mixture design (SCMD) defined the best formulation for the simultaneous optimization of highest lipase activity with the lowest cost using the variables as units of lipase activity per milliliter of enzymatic extract (U/mL) and units of lipase activity per brazilian monetary unit (U/R$). SM has the highest cost, followed by CSM and WB. The experimental conditions were: 10g raw material: 10g water, 30ºC, 1 mL inoculum and 96 hours of incubation. The analysis of variance (ANOVA) validated the experimental data, indicating that the SCMD is a reliable method for determining the optimum mixture. It was not observed a synergist effect in the substrate mixture for lipase activity. The optimization by the desirability function showed that cotton seed meal is the most promise substrate, although the second highest price cost, it obtained the best results (6.7 U/mL and 111216.19 U/R$). The experimental validation using 100% of cotton seed meal achieved 9.9 U/mL and 162295.08 U/R$. The results also indicated a direct correlation between lipase activity and lipid concentration of the mixture (r = 0.84).