Energy demand currently is mostly satisfied by the use of fossil fuels that present not only problems for the environment, but also are not renewable. Dark fermentation (DF) is a biological process that can provide an alternative to meet energy needs. The use of industrial effluents from carbon-rich waters in the production of hydrogen and volatile fatty acids (VFAs) through the DF is a promising strategy. However, determining the optimal operating conditions to increase the production of the sub-products has been not being thoroughly studied. This study aims at determining the optimal condition of hydraulic retention time (HRT) and pH to improve hydrogen production in an internal recirculation (IC) reactor treating beverage wastewater using Response Surface Methodology and interaction between factors. The study also assessed VFAs concentrations when operating at optimal conditions. The results showed that the combination of an 8.0 h HRT and 5.5 produced 30% of hydrogen and the interaction between factors established that the pH was the main factor influencing the results. Also, it was observed that the concentration of lactic acid did not inhibit the production of H2 for the optimal value of pH. However, it is evident that more studies using HRT of less than 8 h and of VFAs distribution are needed. Future optimization technique considerations are necessary to reduce uncertainties for biological hydrogen production purposes.