Purification of Trypsin from Sardine (<i>Sardina pilchardus</i>) Viscera and Its Application in Preparation of Antioxidative Fish Protein Hydrolysates: http://www.doi.org/10.26538/tjnpr/v8i1.25

Laila Manni; Nouhaila Zouine; Ibtissam Ouahidi; Meryem Bouraqqadi; Samir Ananou

Authors

Laila Manni Laboratory of Microbial Biotechnology and Bioactive Molecules, Sciences and Technologies Faculty, Sidi Mohamed Ben Abdellah University, P. O. Box 2202, Imouzzer Road Fez, Morocco
Nouhaila Zouine Laboratory of Microbial Biotechnology and Bioactive Molecules, Sciences and Technologies Faculty, Sidi Mohamed Ben Abdellah University, P. O. Box 2202, Imouzzer Road Fez, Morocco
Ibtissam Ouahidi High Institute of Nursing and Technical Health (ISPITS). Ministry of Health, Fez, Morocco laila.manni@usmba.ac.ma
Meryem Bouraqqadi Laboratory of Microbial Biotechnology and Bioactive Molecules, Sciences and Technologies Faculty, Sidi Mohamed Ben Abdellah University, P. O. Box 2202, Imouzzer Road Fez, Morocco
Samir Ananou Laboratory of Microbial Biotechnology and Bioactive Molecules, Sciences and Technologies Faculty, Sidi Mohamed Ben Abdellah University, P. O. Box 2202, Imouzzer Road Fez, Morocco

Keywords:

Antioxidant activity, Hydrolysates, Purification, Alkaline trypsin, Sardine by-products

Abstract

A substantial quantity of solid fish by-products is generated during the fish processing. These by-products offer an intriguing opportunity as a source of high added-value compounds, including digestive proteases. Indeed, fish trypsin is one of the most beneficial and useful biomolecules that can be recovered from fish wastes. Additionally, fish protein hydrolysates have emerged as a valuable and abundant source of high-quality bioactive molecules, which can be efficiently recovered through enzymatic hydrolysis. This study aims to provide valuable insights into the extraction and utilization of fish trypsin, as well as protein hydrolysates derived from fish by-products, specifically those from sardine (Sardina pilchardus). Firstly, trypsin from the viscera of sardine was purified with an approximate fourteen-fold increase in specific activity using ammonium sulfate precipitation, followed by DEAE-cellulose chromatography. SDS–PAGE analysis revealed a single band of approximately 27 kDa. Interestingly, the purified enzyme exhibited significant features, including an optimum temperature of 60 °C, and high stability at low temperatures, retaining respectively 100% and 64% of its initial activity at 20 °C and 50 °C. The enzyme also demonstrated excellent stability at pH range 5–10 with an optimum at pH 8. Furthermore, it maintained 40% of its enzymatic activity at the pH 11. The purified enzyme was inhibited by benzamidine and PMSF and partially inhibited by EDTA.
Subsequently, four protein hydrolysates (H1, H2, H3, and H4) were prepared from sardine, using purified trypsin and other bacterial proteases. All hydrolysates exhibited noteworthy antioxidant properties in vitro, indicating their promising potential for application in functional foods as natural preservatives.

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