During the processing of dry-cured ham many biochemical changes occur, including the degradation of muscle proteins. These changes are due to the intense action of endogenous proteolytic enzymes. In the present study, the isolation and identification of a large number of peptides derived from creatine kinase has been achieved for the first time in dry-cured ham. A total of 58 peptides coming from different regions of the protein have been identified by mass spectrometry. This study provides evidence for the extensive degradation of creatine kinase during the processing of dry-cured ham as well as the role played by endo- and exopeptidases in the generation of small peptides and free amino acids from polypeptides. These peptides are important in the development of characteristic sensory properties associated with dry-cured ham.