Fish protein hydrolysates (FPH), which comprise a combination of small protein or peptide and free amino acids, may be suitable for use in a variety of applications. For this purpose, the hydrolysis process was carried out for the by-products of trout, whiting, and anchovy with alkaline protease, protamex, and flavourzyme, and the quality of the protein hydrolysate generated under optimized conditions was determined by proximate compositions, amino acid contents, heavy metal, and trace element levels and peptide molecular weight sizes. The time, temperature, and enzyme proportions were determined according to the hydrolysis degree (DH%) under optimum conditions identified by response surface methodology (RSM). The crude protein, lipid, and ash contents of trout, whiting, and anchovy by-products hydrolysates ranged from 77.03% to 88.54%, 3.28% to 6.12%, and 8.18% to 16.8% for dry matter, respectively.Considering the total amino acid contents, the highest number of amino acids was aspartic and glutamic acid in all fish and enzyme groups. In terms of essential amino acids, the highest concentrations of cystine, histidine, lysine, and methionine were found in anchovy by-product hydrolysates produced using the flavourzyme. Using FAO/WHO reference protein as a base, lysine was found to have the highest chemical score for trout by-products hydrolysates using protamex (85.14%), whiting byproducts hydrolysates using alkaline protease (83.66%), and anchovies by-products hydrolysates using flavourzyme (88.45%). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed that the electrophoresis pattern of the hydrolysates of trout, whiting, and anchovy by-products optimized with the RSM using alkaline protease, protamex, and flavourzyme enzymes showed variations depending on the enzyme type and fish by-products.
