Quantitative determination of peptides and proteins by MALDI MS

A modified method of isotope dilution was applied: to the quantitative dete rmination of peptides and proteins by MALDI MS at subpicomolar level. The e ssence of the method consists in the quantitative analysis of the enzymic h ydrolysis products rather than the starting compounds. This allows the meas urements to be performed at a higher resolution and makes the method indepe ndent of the molecular mass of oligopeptides and proteins examined. Fragmen ts obtained by hydrolysis of the Same oligopeptide or protein in a known co ncentration by the same enzyme and labeled with the stable O-18 isotope are used as internal standards. The label is introduced by carrying out the hy drolysis in H-2 O-18, and the oligopeptide concentration is calculated from the isotope distribution between the labeled and unlabeled hydrolysis prod ucts in the mass spectrum. This method was tested in the determination of c oncentrations of the angiotensinogen (1-14) fragment (oligopeptide), extrac ellular RNAase from Bacillus amyloliquefaciens (protein) and its protein in hibitor, barstar M. Usefulness of this method in kinetic studies was also d emonstrated.