Application of amino acid analysis using hydrophilic interaction liquid chromatography coupled with isotope dilution mass spectrometry for peptide and protein quantification.
Amino acid analysis that is based on the use of hydrophilic interaction liquid chromatography (HILIC) coupled with isotope dilution mass spectrometry (IDMS) has been developed for the accurate quantification of underivatized amino acids from hydrolyzed protein/peptide. Sufficient separation of amino acids on a zwitterion chromatography (ZIC)-HILIC column was achieved after removal of chloride ions in the hydrolyzate. The detection limits and quantification limits as concentration of the four amino acids ranged from 0.003 to 0.04pmol microL(-1) and from 0.01 to 0.1pmol microL(-1), respectively. The analytical results for the certified reference materials, angiotensin I and bovine serum albumin (BSA), were satisfactory. Furthermore, the quantitative results by this method were compared with those by the commercially available precolumn method, derivatizd with aminoquinolylhydroxysuccinimidyl carbamate (AQC method), and better recovery and more precise data were obtained with this method.
Kato
Megumi
M
Bio-Medical Standards Section, Organic Analytical Chemistry Division, National Metrology Institute of Japan, National Institute of Advanced Industrial Science and Technology, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8563, Japan. katou-megu@aist.go.jp
Kato
Hisashi
H
Eyama
Sakae
S
Takatsu
Akiko
A
D000596:Amino Acids / Q000032:analysis*; D000803:Angiotensin I / Q000032:analysis; D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D007553:Isotope Labeling / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D012680:Sensitivity and Specificity; D012710:Serum Albumin, Bovine / Q000032:analysis
D000596:Amino Acids; D010455:Peptides; D011506:Proteins; D012710:Serum Albumin, Bovine; D000803:Angiotensin I