Species-specific identification of Colla corii asini and its non-donkey adulterating ingredients based on liquid chromatography–tandem mass spectrometry proteomics

Colla corii asini (CCA), also called Ejiao, is a protein-rich, healthy food prepared from the dried or fresh skins of donkeys with high nutritional and medicinal value. Therefore, identifying CCA and its non-donkey adulterating ingredients is of great significance. In this study, liquid chromatography–tandem mass spectrometry proteomics technology combined with bioinformatics was used to discover the specific peptide biomarkers in CCA and its non-donkey adulterating ingredients (sheep, horse, pig, camel, and cattle). A total of nine specific peptide biomarkers (one from CCA; one each from horse and pig skin gelatin; and two each from sheep, camel, and cow skin gelatin) with good signal responses were screened. After synthesizing these nine specific peptide biomarkers, an ultrahigh-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) multireaction monitoring (MRM) analysis method was established and the limits of specific peptide biomarkers of non-donkey adulterating ingredients content in CCA were proposed. This study established a rapid, simple, highly sensitive, and specific method for the authenticity certification and quality assessment of CCA, ensuring product quality and safety. The method enables identification of specific peptide biomarkers in ass hide glue while simultaneously detecting five adulterated ingredients: sheep, horse, pig, camel, and cattle skins. • Identified unique peptide biomarkers for CCA & five adulterants • Developed UPLC-MS/MS MRM method for CCA authentication & adulterant detection. • Overcomes Pharmacopoeia method limits, especially for horse skin adulteration.