A study of bio‐hydrogen production was performed in a continuous flow anaerobic fermentation reactor (with an available volume of 5.4 L). The continuous stirred tank reactor (CSTR) for bio‐hydrogen production was operated under the organic loading rates (OLR) of 8–32 kg COD/m3 reactor/d (COD: chemical oxygen demand) with molasses as the substrate. The maximum hydrogen production yield of 8.19 L/d was obtained in the reactor with the OLR increased from 8 kg COD/m3 reactor/d to 24 kg COD/m3 d. However, the hydrogen production and volatile fatty acids (VFAs) drastically decreased at an OLR of 32 kg COD/m3 reactor/d. Ethanoi, acetic, butyric and propionic were the main liquid fermentation products with the percentages of 31%, 24%, 20% and 18%, which formed the mixed‐type fermentation.A study of bio‐hydrogen production was performed in a continuous flow anaerobic fermentation reactor (with an available volume of 5.4 L). The continuous stirred tank reactor (CSTR) for bio‐hydrogen production was operated under the organic loading rates (OLR) of 8–32 kg COD/m3 reactor/d (COD: chemical oxygen demand) with molasses as the substrate. The maximum hydrogen production yield of 8.19 L/d was obtained in the reactor with the OLR increased from 8 kg COD/m3 reactor/d to 24 kg COD/m3 d. However, the hydrogen production and volatile fatty acids (VFAs) drastically decreased at an OLR of 32 kg COD/m3 reactor/d. Ethanoi, acetic, butyric and propionic were the main liquid fermentation products with the percentages of 31%, 24%, 20% and 18%, which formed the mixed‐type fermentation.