Abstract

Fish processing waste is a secondary raw material that can be used as part of value-added product systems. Fish protein hydrolysates contain essential amino acids and biologically active peptides that can be included in food formulations. This article introduces an improved enzymatic hydrolysis technology for obtaining protein hydrolysates from secondary fish raw materials.
The study featured Central-East Atlantic cod (Gadus morhua) and its processing waste, which included protein-containing backbones, muscle tissue, and fins. Five different enzymes (Pancreatin, Collagenase, Protozyme B, Alcalase 2.4 L FG, and Enzy-Mix U) were applied in a wide range of concentrations. The new technology involved one additional step to improve the sensory properties: protein hydrolysates were treated with an acetic acid solution. The alkaline dissolution stage was also optimized to increase the degree of protein solubility. The resulting hydrolysates underwent a chemical analysis and high-performance liquid chromatography. Other tests made it possible to reveal their amino acid composition and molecular weight distribution.
The fish protein hydrolysates demonstrated a high protein content of 85–90% and a high amino acid ratio (110–190%) for virtually all essential amino acids. The product yield ranged from 22 to 55%. As the concentration increased from 1 to 8 g per 1 kg of raw material, the yield of protein hydrolysate and the degree of protein hydrolysis also increased. However, the amount of lowmolecular-weight peptides also grew, which spoilt the sensory profile (bitter taste, strong fishy smell).
Only Protozyme proved suitable for the food industry: its concentration could be varied whereas the enzymolysis time could be shortened, if necessary. Pancreatin and Alcalase 2.4 L FG provided high-yield protein hydrolysates for microbiological use.

Keywords

Fish protein hydrolysate, Gadus morhua, fish processing waste, enzymatic hydrolysis, food technology

References

1. Yugay AV. Relevance of the use of fish protein hydrolysates in food technology. Health, Food & Biotechnology. 2023;5(2):51– 66. (In Russ.) https://doi.org/10.36107/hfb.2023.i2.s173
2. Lu X, Qian S, Wu X, Lan T, Zhang H, et al. Research progress of protein complex systems and their application in food: A review. International Journal of Biological Macromolecules. 2024;265(Part 2):130987. https://doi.org/10.1016/j.ijbiomac.2024.130987
3. Ramakrishnan SR, Jeong C-R, Park J-W, Cho S-S, Kim S-J. A review on the processing of functional proteins or peptides derived from fish by-products and their industrial applications. Heliyon. 2023;9(3):e14188. https://doi.org/10.1016/j.heliyon.2023.e14188
4. Honrado A, Miguel M, Ardila P, Beltrán JA, Calanche JB. From waste to value: Fish protein hydrolysates as a technological and functional ingredient in human nutrition. Foods. 2024;13(19):3120. https://doi.org/10.3390/foods13193120
5. Sokolov AV. Current state and development trends of the Russian fisheries sector. Technologies for the food and processing industry of aic – Healthy food. 2019;(4):36–48. (In Russ.) https://doi.org /10.24411/2311-6447-2019-10021
6. Zhao H, Wang M, Peng X, Zhong L, Liu X, et al. Fish consumption in multiple health outcomes: An umbrella review of meta-analyses of observational and clinical studies. Annals of Translational Medicine. 2023;11(3):152. https://dx.doi.org/10.21037/atm-22-6515
7. Zamora-Sillero J, Gharsallaoui A, Prentice C. Peptides from fish by-product protein hydrolysates and its functional properties: An overview. Marine Biotechnology. 2018;20(2):118–130. https://doi.org/10.1007/s10126-018-9799-3
8. Rafieezadeh D, Esfandyari G. Marine bioactive peptides with anticancer potential, a narrative review. International Journal of Biochemistry and Molecular Biology. 2024;15(4):118–126. https://doi.org/10.62347/TUVQ7468
9. Ishak NH, Sarbon NM. A review of protein hydrolysates and bioactive peptides deriving from wastes generated by fish processing. Food and Bioprocess Technology. 2018;11(11):2–16. https://doi.org/10.1007/s11947-017-1940-1
10. Idowu AT, Igiehon OO, Idowu S, Olatunde OO, Benjakul S. Bioactivity potentials and general applications of fish protein hydrolysates. International Journal of Peptide Research and Therapeutics. 2021;27:109–118. https://doi.org/10.1007/s10989-020-10071-1
11. Nemati M, Shahosseini SR, Ariaii P. Review of fish protein hydrolysates: Production methods, antioxidant and antimicrobial activity and nanoencapsulation. Food Science and Biotechnology. 2024;33:1789–1803. https://doi.org/10.1007/s10068-024-01554-8
12. Yu Y, Fan F, Wu D, Yu C, Wang Z, et al. Antioxidant and ACE inhibitory activity of enzymatic hydrolysates from Ruditapes philippinarum. Molecules. 2018;23(5):1189. http://dx.doi.org/10.3390/molecules23051189
13. Honrado A, Rubio S, Beltrán JA, Calanche J. Fish by-product valorization as source of bioactive compounds for food enrichment: Characterization, suitability and shelf life. Foods. 2022;11(22):3656. https://doi.org/10.3390/foods11223656
14. Petrova I, Tolstorebrov I, Zhivlyantseva J, Eikevik TM. Utilization of fish protein hydrolysates as peptones for microbiological culture medias. Food Bioscience. 2021;42(5):101063. http://dx.doi.org/10.1016/j.fbio.2021.101063
15. Broli G, Nygaard H, Sletta H, Sandnes K, Aasen IM. Farmed salmon rest raw materials as a source of peptones for industrial fermentation media. Process Biochemistry. 2021;102:157–164. https://doi.org/10.1016/j.procbio.2020.12.004
16. Horn SJ, Aspmo SI, Eijsink VGH. Growth of Lactobacillus plantarum in media containing hydrolysates of fish viscera. Journal of Applied Microbiology. 2005;99(5):1082–1089. http://dx.doi.org/10.1111/j.1365-2672.2005.02702.x
17. Hyun J, Rho Y, Nagahawatta DP, Lee G, Lee S, et al. Upcycling fish byproducts for skin health: An enzymatic approach to sustainable nutricosmetics. Food Bioscience. 2025;66(11):106205. http://dx.doi.org/10.1016/j.fbio.2025.106205
18. Antipova LV, Storublevtsev SA, Bolgova SB. Creation of collagen products fish raw material. Proceedings of the Voronezh state university of engineering technologies. 2015;1(63):130–133. (In Russ.) https://elibrary.ru/TTUFGP
19. Ribeiro TB, Maia MRG, Fonseca AJM, Marques B, Caleja C, et al. A comprehensive review of fish protein hydrolysates targeting pet food formulations. Food Reviews International. 2024;41(5):1321–1359. https://doi.org/10.1080/87559129.2024.2430660
20. Gao R, Yu Q, Shen Y, Chu Q, Chen G, et al. Production, bioactive properties, and potential applications of fish protein hydrolysates: Developments and challenges. Trends in Food Science & Technology. 2021;110:687–699. https://doi.org/10.1016/j.tifs.2021.02.031
21. Shahidi F, Varatharajan V, Peng H, Senadheera R. Utilization of marine by-products for the recovery of value-added products. Journal of Food Bioactives. 2019;6:10–61. https://doi.org/10.31665/JFB.2019.6184
22. Ozogul F, Cagalj M, Simat V, Ozogul Y, Tkaczewska J, et al. Recent developments in valorisation of bioactive ingredients in discard/seafood processing by-products. Trends in Food Science & Technology. 2021;116:559–582. https://doi.org/10.1016/j.tifs.2021.08.007
23. Kaushik N, Falch E, Slizyte R, Kumari A, Khushboo, et al. Valorization of fish processing by-products for protein hydrolysate recovery: Opportunities, challenges and regulatory issues. Food Chemistry. 2024;459:140244. https://doi.org/10.1016/j.foodchem.2024.140244
24. Hoeling A, Grimm T, Volkov VV, Mezenova NYu. Hydrolysis process of fish cannery by-products. Journal of International Academy of Refrigeration. 2016;(1):3–8. (In Russ. ) https://elibrary.ru/VVRJSX
25. Mezenova OYa, Volkov VV, Merzel T, Grimm T, Kyun S, et al. Hydrolysis methods for collagen-containing fish raw materials for obtaining peptides and their amino acid profile. Proceedings of Universities. Applied Chemistry and Biotechnology. 2018;8(4):83–94. (In Russ.) https://doi.org/10.21285/2227-2925-2018-8-4-83-94
26. Yuan Z, Ye X, Hou Z, Chen S. Sustainable utilization of proteins from fish processing by-products: Extraction, biological activities and applications. Trends in Food Science & Technology. 2023;143:104276. https://doi.org/10.1016/j.tifs.2023.104276
27. Lebedeva SN, Bolkhonov BA, Zhamsaranova SD, Bazhenova BA, Leskova SYu. Functional profile of enzymatic hydrolysates in food proteins. Food Processing: Techniques and Technology. 2024;54(2):412–422. (In Russ.) https://doi.org/10.21603/2074-9414-2024-2-2515
28. Mazorra-Manzano MA, Ramírez-Suárez JC. Proteolytic enzymes for production of functional protein hydrolysates and bioactive peptides. In: Yada RY, Dee DR, editors. Improving and Tailoring Enzymes for Food Quality and Functionality, 2th ed. UK: Woodhead Publishing; 2024. pp. 325–354. https://doi.org/10.1016/B978-0-443-15437-9.00013-6
29. Abdullah FI, Hamid NH, Abd Karim MM, Ismail MF, Sin NLWW, et al. Fish protein hydrolysate for fish health. Biocatalysis and Agricultural Biotechnology. 2024;60:103292. https://doi.org/10.1016/j.bcab.2024.103292
30. Derkach SR, Kolotova DS, Kuchina YuA, Shumskaya NV. Characterization of fish gelatin obtained from Atlantic cod skin using enzymatic treatment. Polymers. 2022;14(4):751. https://doi.org/10.3390/polym14040751
31. Alp-Erbay E, Yeşilsu AF. Fish protein and its derivatives: Functionality, biotechnology and health effects. Aquatic Food Studies. 2021;1(1):AFS13. https://doi.org/10.4194/AFS13
32. Bredikhina OV, Zarubin NYu. Development of complex technology of processing of organic waste of fish processing enterprises on collagencontaining hydrolysates for food purposes. Trudy VNIRO. Aquatic bioresources processing technologies. 2019;176:109–121. (In Russ.) https://elibrary.ru/QADCUA
33. Association of Official Analytical Chemist AOAC. Official Methods of Analysis, 20th ed. NY: AOAC International; 2016.
34. Dinakarkumar Yu, Krishnamoorthy S, Margavelu G, Ramakrishnan G, Chandran M. Production and characterization of fish protein hydrolysate: Effective utilization of trawl by-catch. Food Chemistry Advances. 2022;1:100138. https://doi.org/10.1016/j.focha.2022.100138
35. Coles GD, Wratten SD, Porter JR. Food and nutritional security require adequate protein as well as energy, delivered from whole-year crop production. Peer J. 2016;4(6245):e2100. http://dx.doi.org/10.7717/peerj.2100
36. Verdú S, Fuentes C, Fuentes A, Pérez AJ, Barat JM, et al. Predicting fish by-product proteolysis status by RGB laser-scattering imaging combined with machine learning procedures. Journal of Food Engineering. 2023;358:111660. https://doi.org/10.1016/j.jfoodeng.2023.111660
37. Moreira TFM, Pessoa LGA, Seixas FAV, Ineu RP, Gonçalves OH, et al. Chemometric evaluation of enzymatic hydrolysis in the production of fish protein hydrolysates with acetylcholinesterase inhibitory activity. Food Chemistry. 2022;367:130728. https://doi.org/10.1016/j.foodchem.2021.130728
38. Tadesse SA, Emire SA, Barea P, Illera AE, Melgosa R, et al. Valorisation of low-valued ray-finned fish (Labeobarbus nedgia) by enzymatic hydrolysis to obtain fish-discarded protein hydrolysates as functional foods. Food and Bioproducts Processing. 2023;141:167–184. https://doi.org/10.1016/j.fbp.2023.08.003
39. Khan MU, Hamid K, Tolstorebrov I, Eikevik TM. A comprehensive investigation of the use of freeze concentration approaches for the concentration of fish protein hydrolysates. Food Chemistry. 2024;452:139559. https://doi.org/10.1016/j.foodchem.2024.139559
40. Chorhirankul N, Janssen AEM, Boom RM. UF fractionation of fish protein hydrolysate. Separation and Purification Technology. 2024;330(Part A):125232. https://doi.org/10.1016/j.seppur.2023.125232