Co-substrate utilization of various wastes with complementary characteristics can provide a complete medium for higher hydrogen production. This study evaluated potential of apple pomace hydrolysate (APH) co-fermented with crude glycerol (CG) for increased H-2 production and decreased by-products formation. The central composite design (CCD) along with response surface methodology (RSM) was used as tool for optimization and 15 g/L of CG, 5 g/L of APH and 15% (v/v) inoculum were found to be optimum to produce as high as 26.07 +/- 1.57 mmol H-2/L of medium. The p-value of 0.0017 indicated that APH at lower concentration had a significant effect on H-2 production. By using CG as sole carbon source, reductive pathway of glycerol metabolism was favored with 19.46 mmol H-2/L. However, with APH, oxidative pathway was favored with higher H-2 production (26.07 +/- 1.57 mmol/L) and decrease in reduced by-products (1,3-propanediol and ethanol) formation. APH inclusion enhanced H-2 production, and decreased substrate inhibition. (C) 2015 Elsevier Ltd. All rights reserved.