Determination of cysteine plus half-cystine in protein and peptide hydrolysates: use of dithiodiglycolic acid and phenylisothiocyanate derivatization.
Cysteine and half-cystine in proteins and peptides can be determined by acid hydrolysis in the presence of dithiodiglycolic acid, derivatization with phenyl isothiocyanate, and HPLC separation of the phenylthiocarbamyl amino acids. The determination requires no additional sample handling, oxidation, reduction, alkylation, or desalting. Samples containing cystine are treated in the same manner as those containing cysteine. Derivatization of cysteine or half-cystine by dithiodiglycolic acid has no effect on any other amino acids. Recovery of the PTC-cysteine derivative is greater than 90% over the linear range of 5 pmol to 1 nmol. Precision of the cysteine plus half-cystine determination is equivalent to that observed for all other amino acids. Structure work on the derivative indicates that it is a mixed disulfide containing cysteine and thioglycolic acid.
Hoogerheide
J G
JG
Fine Chemical Analytical Methods and Services-Analytical Biochemistry, Upjohn Company, Kalamazoo, Michigan 49001.
Campbell
C M
CM
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine / Q000032:analysis*; D006016:Glycolates; D006801:Humans; D006868:Hydrolysis; D017879:Isothiocyanates; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D015203:Reproducibility of Results; D013861:Thiocyanates; D013864:Thioglycolates
D000596:Amino Acids; D006016:Glycolates; D017879:Isothiocyanates; D010446:Peptide Fragments; D011506:Proteins; D013861:Thiocyanates; D013864:Thioglycolates; C005441:phenylisothiocyanate; C017487:2-mercaptoacetate; D003545:Cysteine