On-capillary derivatization and analysis of amino acids in human plasma by capillary electrophoresis with laser-induced fluorescence detection: application to diagnosis of aminoacidopathies.
A capillary electrophoresis with laser-induced fluorescence detection method for the analysis of free amino acids (AA) in human plasma was developed. A mixture of 16 AA was on-capillary derivatized with 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ) and separated inside the capillary in less than 30 min using 70 mM borax-3.5 mM SDS pH 9.3 as running buffer. Four plasma samples from a healthy donor and patients suffering from phenylketonuria, propionic acidemia, and tyrosinemia type II were studied. Repeatabilities calculated as intra-day RSD (n = 3) values for the AA involved in these aminoacidopathies (glycine, phenylalanine, and tyrosine) were in the range of 0.3 to 1.2% for migration time and 3.7 to 8.2% for peak height. Reproducibilities calculated as inter-day RSD (n = 4) values for the same AA were between 0.7 and 1.4% for migration time and 4.7 and 9.1% for peak height. A fast qualitative analysis allowed the identification of the corresponding disease by comparing the electrophoretic profiles from the patient and the healthy donor and noting the increased level of the specific AA accompanying each individual disease. The results of the quantitative analysis for glycine, phenylalanine, and tyrosine in the plasma samples studied using the developed method showed a good agreement with those provided by the Center of Diagnosis of Molecular Diseases using a standard method for AA analysis.
Veledo
Maria Teresa
MT
Institute of Organic Chemistry (C.S.I.C.), Juan de la Cierva 3, E-28006 Madrid, Spain.
de Frutos
Mercedes
M
Diez-Masa
Jose Carlos
JC
D000592:Amino Acid Metabolism, Inborn Errors / Q000097:blood / Q000175:diagnosis*; D000596:Amino Acids / Q000097:blood* / Q000302:isolation & purification; D016022:Case-Control Studies; D019075:Electrophoresis, Capillary / Q000379:methods*; D007834:Lasers; D012015:Reference Standards; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence / Q000379:methods*
D000596:Amino Acids