Species-specific identification of donkey-hide gelatin and its adulterants using marker peptides

Abstract

Donkey-hide gelatin is an important traditional Chinese medicine made from donkey skin. Despite decades of effort, identifying the animal materials (donkeys, horses, cattle and pigs) in donkey-hide gelatin remains challenging. In our study, we aimed to identify marker peptides of donkey-hide gelatin and its adulterants and develop a liquid chromatography–tandem mass spectrometry method to identify them. Theoretical marker peptides of four animals (donkeys, horses, cattle and pigs) were predicted and verified by proteomic experiments, and 12 species-specific marker peptides from donkey-hide gelatin and its adulterants were identified. One marker peptide for each gelatin was selected to develop the liquid chromatography–tandem mass spectrometry method. The applicability of the method was evaluated by investigating homemade mixed gelatin samples and commercial donkey-hide gelatin products. Using the liquid chromatography–tandem mass spectrometry method, the addition of cattle-hide gelatin and pig-hide gelatin to donkey-hide gelatin could be detected at a level of 0.1%. Horse-hide gelatin was detected when added at a level of 0.5%. Among 18 batches of donkey-hide gelatin products, nine were identified as authentic, and eight of the remaining samples were suspected to be adulterated with horse materials. These results provide both a practical method to control the quality of donkey-hide gelatin and a good reference for quality evaluations of other medicinal materials and foods containing protein components.