In the present study, the effect of amino acids (viz. methionine, alanine, histidine, cysteine, and lysine) on hydrogen production from organic fraction of municipal solid waste was evaluated using co-culture of E. colt and E. aerogenes. The amino acids were applied in the concentration range of 1.0, 2.5, 5.0, 7.5, and 10.0 g/L. Modified Gompertz model was used to analyze cumulative hydrogen production (P), maximum hydrogen production rate (Rmax) and lag phases (lambda). The results exhibited that the hydrogen production was positively affected by each amino acid at every concentration applied. Application of alanine resulted in the highest cumulative hydrogen production and volumetric yield of 685.4 +/- 10.1 mL and 1.9561 L-H2/L-substrate, respectively. Amino acid supplementation resulted in the increase of hydrogen yield by 87.7%, 126.6%, 75.5%, 108.7% and 101.7%, respectively by addition of methionine, alanine, histidine, cysteine, and lysine as compared to control. COD removal and VFA generation were positively affected by amino acid addition.