Process parameters on enzymatic hydrolysis and molecular weight (MW) distribution of collagen hydrolysates from Gadus morrhua skin were investigated. The optimal process parameters were obtained by the single-factor and orthogonal experiments. The molecular weight distribution of hydrolysates was determined using both Sephadex G25 partition and high speed liquid chromatography electricity spray mass spectrum (HPLC-ESI-MS). Collagen hydrolysates were first gained by an alkaline protease “alcalase” for 3 h at temperature (50°C), pH (10.0), substrate concentration (75 g L-1), and E/S (3%). The molecular weight distribution of collagen hydrolysates ranged from 300 to 1 500 Da, and most of peptides were under 1 200 Da. Sephadex G25 partition and HPLC-ESI-MS should be successfully employed to determine the molecular weight distribution of collagen hydrolysates. The optimal process parameters were obtained by the single-factor and orthogonal experiments. The molecular weight distribution of hydrolysates was determined using both Sephadex G25 partition and high speed liquid chromatography electricity spray mass spectrum (HPLC-ESI-MS). Collagen hydrolysates were first gained by an alkaline protease “alcalase” for 3 h at 50 ° C, pH (10.0) 75 g L-1), and E / S (3%). The molecular weight distribution of collagen hydrolysates ranged from 300 to 1 500 Da and most of the peptides were under 1 200 Da. Sephadex G25 partition and HPLC-ESI-MS should be successfully employed to determine the molecular weight distribution of collagen hydrolysates.