RESUMO
BACKGROUND: The Beckman 6300/7300 analyzer, which was widely used for amino acid (AA) analysis, is no longer commercially available. METHODS: To set up an affordable AA analysis program, a Beckman system gold HPLC 126AA analyzer and Pickering Laboratories reagents were used. Two quantitative AA analysis programs were developed. One was an 18-min short program quantitating seven AAs from plasma and dried blood spots (DBS) specimens using Lithium eluents Li-365 and Li-375 at 70 degrees C column temperature. The short program could be used for diagnosis and follow-up dietary management for phenylketonuria (PKU), maple syrup urine disease (MSUD), tyrosinemia and homocystinuria patients. The second program was a 118-min long AA screening panel quantitating 40 AAs using Lithium eluents Li-275, Li-365 and Li-375 at 32, 48 and 72 degrees C column temperatures from plasma and urine specimens. RESULTS: The values obtained from DBS specimens were in good agreement with certified results from the Centers for Disease Control and Prevention. The values obtained from plasma and urine samples were in good correlation with those obtained from Beckman 6300 analyzer (0.9076 < or = r < or = 0.999). CONCLUSIONS: Amino acid quantitation from physiological samples using a Beckman 126AA Analyzer and Pickering Laboratories reagents was useful for clinical diagnosis and monitoring of aminoacidopathies.
Assuntos
Aminoácidos/sangue , Aminoácidos/urina , Cromatografia Líquida de Alta Pressão/métodos , Software , Coleta de Amostras Sanguíneas/métodos , Homocistinúria/diagnóstico , Homocistinúria/metabolismo , Humanos , Processamento de Imagem Assistida por Computador , Doença da Urina de Xarope de Bordo/diagnóstico , Doença da Urina de Xarope de Bordo/metabolismo , Fenilcetonúrias/diagnóstico , Fenilcetonúrias/metabolismo , Tirosinemias/diagnóstico , Tirosinemias/metabolismoRESUMO
Isothermal and isocratic HPLC chromatography for rapid quantitation of isoleucine and leucine (merged peak), tyrosine and phenylalanine from a single dried blood filter paper specimen (1/8" dot blot) is described. The net chromatography time of the relevant peaks is less than 3 min. The results can be used to screen for PKU, MSUD, and tyrosinemia in one single assay. The advantage for early determination of PKU by simultaneous quantitation of both phenylalanine and tyrosine with the determination of their ratio is discussed. Using a dual column system this procedure might be suitable for large-scale newborn screening.
Assuntos
Autoanálise/métodos , Isoleucina/sangue , Leucina/sangue , Triagem Neonatal , Papel , Fenilalanina/sangue , Tirosina/sangue , Erros Inatos do Metabolismo dos Aminoácidos/sangue , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Cromatografia Líquida de Alta Pressão , Humanos , Recém-Nascido , Doença da Urina de Xarope de Bordo/sangue , Doença da Urina de Xarope de Bordo/diagnóstico , Fenilcetonúrias/sangue , Fenilcetonúrias/diagnósticoRESUMO
In this paper we report a high-resolution method and special buffers for the analysis for 45 common amino acids and related compounds and seven less-common components occasionally found in physiological fluids. Careful optimization of the chromatographic protocol resolves cysteine-penicillamine mixed disulfide, penicillamine disulfide, and dihydroxyphenylalanine from cystine and methionine; cysteine-homocysteine mixed disulfide is eluted between tyrosine and phenylalanine. Argininosuccinic acid is eluted before ethanolamine; glycerophosphoethanolamine is separated from taurine; glucosaminic acid is eluted between urea and aspartic acid. A Beckman Instruments System 6300 Amino Acid Analyzer was used. System performance is examined in terms of coefficient of variation (CV) over the concentration range of 1.0-5.0 nmol/50 microL; average CV for all components is better than 1.5%. Method performance is assessed through the analysis of normal and abnormal urine and plasma specimens.