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L-Phenylalanine concentration in blood of phenylketonuria patients: a modified enzyme colorimetric assay compared with amino acid analysis, tandem mass spectrometry, and HPLC methods
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Veronica De Silva
Published/Copyright:
July 7, 2010
Abstract
Background: Phenylketonuria (PKU) is an autosomal recessive disorder caused by an impaired conversion of L-phenylalanine (Phe) to L-tyrosine, typically resulting from a deficiency in activity of a hepatic and renal enzyme L-phenylalanine hydroxylase. The disease is characterized by an increased concentration of Phe and its metabolites in body fluids.
Methods: A modified assay based on an enzymatic-colorimetric methodology was developed for measuring blood Phe levels in PKU patients; this method is designed for use with undeproteinized samples and avoids the use of solvents or amphiphilic agents. Thus, the method could be suitable for incorporation into a simple home-monitoring device.
Results: We report here on a comparison of blood Phe concentrations in PKU patients measured in undeproteinized plasma using this enzyme colorimetric assay (ECA), with values determined by amino acid analysis (AAA) of deproteinized samples, and HPLC and tandem mass spectrometry (MS/MS) analyses of dried blood spot (DBS) eluates. Pearson correlation coefficients of 0.951, 0.976 and 0.988 were obtained when AAA-measured Phe concentrations were compared with the ECA-, HPLC- or MS/MS-measured values, respectively. A Bland-Altman analysis revealed that mean Phe concentrations determined using AAA were on average 65 μmol/L lower than values measured by our ECA. These results may be the result of minimizing the manipulations performed on the patient sample compared with AAA, HPLC, and MS/MS methods, which involve plasma deproteinization or DBS elution and derivatization.
Conclusions: The results reported here confirm that Phe concentrations determined by our ECA method are comparable to those determined by other widely used methods for a broad range of plasma Phe concentrations.
Clin Chem Lab Med 2010;48:1271–9.
Keywords: enzyme colorimetric analysis; hyperphenylalaninemia; phenylalanine; phenylalanine dehydrogenase
Received: 2010-1-14
Accepted: 2010-4-7
Published Online: 2010-07-07
Published in Print: 2010-09-01
©2010 by Walter de Gruyter Berlin New York
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Keywords for this article
enzyme colorimetric analysis;
hyperphenylalaninemia;
phenylalanine;
phenylalanine dehydrogenase
Articles in the same Issue
- Editorial
- Lost in translation…wallowing in transformation?
- Reviews
- Coeliac disease – a diagnostic and therapeutic challenge
- Advancements in anemias related to chronic conditions
- Opinion Paper
- Translations of the “Evidence-Based Medicine” concept in different languages: is it time for international standardisation?
- Genetics and Molecular Diagnostics
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- Retrospective analysis of 88,429 serum and urine glucose EQA results obtained from professional laboratories and medical offices participating in surveys organized by three European EQA centers between 1996 and 2007
- Liquid phase immunoassays utilizing magnetic markers and SQUID magnetometer
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- Amniotic fluid α-fetoprotein microheterogeneity in the prenatal diagnosis of congenital disorders of glycosylation type Ia
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