This study investigated the roles of calcium lignosulfonate (CL) and its derived biochar (BC) in biohydrogen production. CL was pyrolyzed at 250 degrees C to produce BC, and the main characteristics of the two materials were compared. A series of anaerobic processes were carried out at 54 +/- 1 degrees C, and the results suggested that CL remarkably lowered the H-2 yield compared with that obtained with CL-derived BC. The highest H-2 yield (262 mL/g glucose) was obtained at 20 g/L, corresponding to an increase of 50.9%, whereas the lowest H-2 yield (110 mL/g glucose) occurred at 20 g/L and was 36.4% lower than that in the control group (173 mL/g glucose, without CL and BC). The CL-derived BC exhibited more pore structures than CL, providing more attachment sites for anaerobes. In addition, the high potassium, calcium, and magnesium contents and low sulfur dosage enhanced the microbial activity and process stability.