In this paper,a method for the identification of functional peptides in donkey-hide gelatin by liquid chromatography-high resolution mass spectrometry was established.In addition,the in vitro activity verification was carried out.After stewing,trypsin hydrolysis and purification with C₁₈ SPE column,donkey-hide gelatin was detected by UPLC-Q-Exactive quadrupole-electrostatic field orbital trap high resolution mass spectrometer,and analyzed by MaxQuant software.The peptide segments were identified by BLAST sequence comparison on UniProt.The results showed that among 63 consensus peptides obtained from three repeated analyses,there were 19 peptides with clear protein and species origin.According to the structure-activity relationship of peptide activity,nine peptides that may be active were selected for antioxidant activity and anticoagulant activity determination.The results showed that nine peptides all had different degrees of antioxidant activity,of which five of them had anticoagulant activity.The method established in this paper is rapid and effective,and provides technical support for revealing the relationship between the structure and activity of donkey-hide gelatin polypeptide.