title: Rapid analysis of phosphoamino acids from phosvitin by near-infrared cyanine 1-(ε-succinimydyl-hexanoate)-1'-methyl-3,3,3',3'-tetramethyl-indocarbocyanine-5,5'-disulfonate potassium derivatization and polyacrylamide-coated CE with LIF detection.
abstract: A polyacrylamide-coated CE method with LIF detection was developed for analyzing three phosphoamino acids including phosphotyrosine (p-Tyr), phosphothreonine (p-Thr), and phosphoserine (p-Ser). A near-infrared dye, 1-(ε-succinimydyl-hexanoate)-1'-methyl-3,3,3',3'-tetramethyl-indocarbocyanine-5,5'-disulfonate potassium (MeCy5-OSu) was employed for derivatization of these phosphoamino acids. Results indicated that the complete baseline resolution of each phosphoamino acid was obtained within 6.1 min, using 10 mmol/L phosphate buffer (pH 4.0) containing 60 mmol/L SDS as running buffer. The highest derivatization efficiency was achieved in 0.2 mol/L borate buffer (pH 8.8) for 30 min at 30 °C. Linearity of response was found in the range of 0.05-1 μmol/L. The correlation coefficients for these phosphoamino acids were from 0.9940 to 0.9976. The LODs for phosphotyrosine, phosphothreonine, and phosphoserine were about 6, 8, and 8 nmol/L, respectively. The proposed method has been successfully applied to the determination of phosphoamino acids in the hydrolysis sample of a phosphorylated phosvitin. Average recoveries for phosvitin sample were in the range of 94.0-98.0% and coefficients of variation ranged from 2.7 to 4.8%.
authors.affiliation: Department of Chemistry, Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Wuhan University, Wuhan, P. R. China.
authors.forename: Ni-Na
authors.initials: NN
authors.lastname: Fu
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Hua-Shan
authors.initials: HS
authors.lastname: Zhang
mesh_terms: D000180:Acrylic Resins / Q000737:chemistry*; D002232:Carbocyanines / Q000737:chemistry*; D053841:Electroosmosis; D019075:Electrophoresis, Capillary / Q000379:methods*; D005452:Fluoresceins / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D019606:Phosphoamino Acids / Q000032:analysis*; D010774:Phosvitin / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013696:Temperature; D000099166:Polyacrylamides
chemical_list: D000180:Acrylic Resins; D002232:Carbocyanines; D005452:Fluoresceins; D019606:Phosphoamino Acids; D010774:Phosvitin; D000099166:Polyacrylamides; C519607:cyanine-1-(epsilon-succinimidyl-hexanoate)-1'-methyl-3,3,3',3'-tetramethyl-indocarbocyanine-5,5'-disulfonate potassium