Capillary electrophoresis with electrochemiluminescence detection for measurement of aspartate aminotransferase and alanine aminotransferase activities in biofluids.
A new sensitive assay for aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities in biofluids was developed, based on the separation and detection of alanine, glutamate, and aspartate using capillary electrophoresis (CE) with electrochemiluminescence (ECL) detection. The three amino acids were separated in 5 mM phosphate of pH 2.1 as background electrolyte, and detected on a 500 microm platinum disk electrode at 1.2V (versus Ag/AgCl) in the presence of 10 mM tris(2,2'-bipyridyl)ruthenium(II) dissolved in 80 mM phosphate of pH 10.5. A mass detection limit of 37.3 fmol (or 81.5 fmol) for glutamate, corresponding to the product in the enzyme reaction catalyzed by 1.24 x 10(-9)U AST (or 2.72 x 10(-9)U ALT) in a 30 min reaction period, was achieved. This assay was applied to investigate the cytotoxicity effect of ethanol on HepG2 cells and differentiating nonalcoholic steatohepatitis (NASH) from alcoholic liver disease, indicating that the technique is promising for the application in the cell biological and clinical fields.
Li
Tao
T
State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin 130022, China.
Yuan
Jipei
J
Yin
Jianyuan
J
Zhang
Zhiwei
Z
Wang
Erkang
E
D000409:Alanine / Q000302:isolation & purification; D000410:Alanine Transaminase / Q000097:blood* / Q000378:metabolism*; D001219:Aspartate Aminotransferases / Q000097:blood* / Q000378:metabolism*; D001224:Aspartic Acid / Q000302:isolation & purification; D001826:Body Fluids / Q000201:enzymology*; D003937:Diagnosis, Differential; D019075:Electrophoresis, Capillary; D000431:Ethanol; D005260:Female; D018698:Glutamic Acid / Q000032:analysis / Q000302:isolation & purification; D006505:Hepatitis / Q000097:blood / Q000175:diagnosis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008108:Liver Diseases, Alcoholic / Q000097:blood / Q000175:diagnosis; D008163:Luminescent Measurements / Q000379:methods*; D008297:Male; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D001224:Aspartic Acid; D000431:Ethanol; D018698:Glutamic Acid; D001219:Aspartate Aminotransferases; D000410:Alanine Transaminase; D000409:Alanine