Abstract: OBJECTIVE Enrichment， identification， and characterization of thiol-containing peptides in water buffalo horn by using core-shell Fe₃O₄@PDA@Au nano-material. METHODS The core-shell Fe₃O₄@PDA@Au magnetic nano-materials were prepared by seed growth method， and the material was applied to the enrichment of thiol-containing peptides in water buffalo horn extract. The contents of -SH and peptide in water buffalo horn samples were determined by Ellman method and peptide assay kit， respectively. And then the peptides in water buffalo horn extract and enriched fraction were identified and analyzed by nano LC-MS/MS system. RESULTS The enrichment factor of Fe₃O₄@PDA@Au nano-materials in the enrichment of thiol-containing peptides in water buffalo horn extract was calculated as 23.41. A total of 4114 peptides were identified in the water buffalo horn extract， including 11 peptides containing Cys residue; a total of 2254 peptides were identified in the enriched fraction， including 709 peptides containing Cys residue. After enrichment， the ratio of the number of thiol-containing peptide to the total peptide in water buffalo horn extract increased from 0.003 to 0.314. The results indicated that the nano-material had a high efficiency on the enrichment of thiol-containing peptides in water buffalo horn extract. CONCLUSION Fe₃O₄@PDA@Au nano-materials can be used for the enrichment of thiol-containing peptides in water buffalo horn， which is of great significance for the basic research of water buffalo horn.