Biohydrogen production from renewable resources using dark fermentation has become an increasingly attractive solution in sustainable global energy supply. So far, there has been no report on the controllability analysis of biohydrogen production using dark fermentation. Process controllability is a crucial factor determining process feasibility. This paper presents a new criterion for assessing biohydrogen process controllability based on PI control. It proposes the critical loop gain derived via Routh stability analysis as a measure of process controllability. Results show that the dark fermentation using the bacteria from anaerobic dairy sludge and substrate source from sugarcane vinasse can lead to a highly controllable process with a critical loop gain value of 4.3. For the two other cases, an increase of substrate concentration from 10 g/L to 40 g/L substantially reduces the controllability. The proposed controllability criterion is easily adopted to assess the process feasibilty based on experimental data.