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SUMMARY   Research background  Cardiovascular diseases and diabetes are the biggest causes of death globally. Bioactive peptides derived from many food proteins using enzymatic proteolysis and food processing have a positive impact on the prevention of these diseases. The bioactivity of Chinese pond turtle muscle proteins and their enzymatic hydrolysates has not received much attention, thus this study aims to investigate their antioxidant, antidiabetic and cytotoxic activities.    Experimental approach  Chinese pond turtle muscles were hydrolysed using four proteolytic enzymes (Alcalase, Flavourzyme, trypsin and bromelain) and the degrees of hydrolysis were measured. High-performance liquid chromatography (HPLC) was conducted to explore the amino acid profiles and molecular mass distribution of the hydrolysates. The antioxidant activities were evaluated using various  in vitro  tests, including 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), hydroxyl radical scavenging activity, reducing capacity, chelating Fe 2+  and lipid peroxide inhibition activity. Antidiabetic activity was evaluated using α-amylase inhibition and α-glucosidase inhibition assays. Besides, cytotoxic effect of hydrolysates on human colon cancer (HT-29) cells was assessed.    Results and conclusions  The amino acid composition of the hydrolysates revealed higher mass fractions of glutamic, aspartic, lysine, hydroxyproline and hydrophobic amino acids. Significantly highest inhibition of lipid peroxidation was achieved when hydrolysate obtained with Alcalase was used. Protein hydrolysate produced with Flavourzyme had the highest radical scavenging activity measured by DPPH (68.32%), ABTS (74.12%) and FRAP ( A  700 nm =0.300) assays, α-glucosidase (61.80%) inhibition and cytotoxic effect (82.26%) on HT-29 cell line at 550 µg/mL. Hydrolysates obtained with trypsin and bromelain had significantly highest (p<0.05) hydroxyl radical scavenging (92.70%) and Fe 2+  metal chelating (63.29%) activities, respectively. The highest α-amylase (76.89%) inhibition was recorded when using hydrolysates obtained with bromelain and Flavourzyme.    Novelty and scientific contribution  Enzymatic hydrolysates of Chinese pond turtle muscle protein had high antioxidant, cytotoxic and antidiabetic activities. The findings of this study indicated that the bioactive hydrolysates or peptides from Chinese pond turtle muscle protein can be potential ingredients in pharmaceuticals and functional food formulations.

In vitro Antioxidant, Cytotoxic and Antidiabetic Activity of Protein Hydrolysates Prepared from Chinese Pond Turtle (Chinemys reevesii)