In this work, the Taguchi method was used to optimize the dark fermentative H2 production from the organic fraction of municipal solid waste (OFMSW). The experiments were planned using the L16 orthogonal array design with each trial conducted at different levels of substrate concentration, inoculum-to-substrate ratio (ISR), and temperature. Based on the results, the optimal setting of the process parameters was the substrate concentration of 6 g-VS/L, ISR 0.5, and temperature of 55 °C. Furthermore, substrate concentration was the most important parameter affecting bio-H2 production among the three process parameters considered. Finally, a confirmation experiment under optimal conditions yielded 62.5 mL H2/g-VSadded, which was higher than all the bio-H2 yield values obtained in the other conditions tested in this study. The measured and predicted bio-H2 yields in the verification test were also very close to each other, confirming the reliability of the Taguchi method in optimizing the bio-H2 production process.