The susceptibility to in-source decay (ISD) in myoglobin, using matrix-assisted laser desorption/ionization (MALDI) coupled with a delayed extraction reflectron time-of-flight (TOF) mass spectrometer, has been described from the standpoint of secondary structures. The ISD data of three different myoglobins showed a common intense product of c35 ion originated from the cleavage of amine bond N-C_α at Gly35-His36 or Ser35-His36 residues. It is suggested that the relatively abundant formation of c35 ion in Gly35-His36 or Ser35-His36 residues is not due to the presence of His36 residue, but due to a turn region between helices in secondary structure of myoglobins.