Enhancing Digestibility and Intestinal Peptide Release of Pleurotus eryngii Protein: An Enzymatic Approach

<i>Pleurotus eryngii</i> is a tasty and low-calorie mushroom containing abundant high-quality protein. This study aims to improve the digestibility of <i>P. eryngii</i> protein (PEP) and hence to facilitate its development as a healthy alternative protein. The extracted PEP was pretreated with 1000-5000 U of papain, neutral protease and alkaline protease. The Chyme collected from in vitro simulated gastrointestinal digestion was analyzed by fluorescence microscopy and protein particle analyzer, and the endpoint profiles of peptides and amino acids were determined by UHPLC-MS/MS and NanoLC-MS/MS. The particle size curve and fluorescence microscopy images jointly supported that protease hydrolysis improved decomposition and dispersion of PEP during digestion, particularly in the gastric phase. The impact on Zeta potential was minimal. Proteases effectively increased the abundance of amino acids after digestion, particularly L-isomer Lys and Arg Maximum release was achieved when pretreated with 5000 U of alkaline protease, reaching 7.54 times that of control. Pretreatments by proteases also notably increased digestive yields of 16,736-19,870 peptides, with the maximum reaching 1.70 times that of the control, which mainly consisted of small peptides composed of 7-15 amino acids with molecular weight below 800 Da. The findings indicated that protease hydrolysis, especially pretreatment with 5000 U of alkaline protease, effectively enhanced the digestibility of PEP, which shed light on providing enzymatic approaches for improving bioavailability and developing healthy fungal proteins.