C V Mihali1, M C Petrescu1, I Mândruţiu2, D Bechet2, T V Nistor3, V Turcuş1, A Ardelean1, Gh Benga1,2,4,5,6,7. 1. "Vasile Goldiş" Western University of Arad - Institute of Life Sciences, Arad, Romania. 2. Cluj County Clinical Emergency Hospital - Laboratory of Genetic Explorations, Cluj-Napoca, Romania. 3. "Iuliu Haţieganu" University of Medicine and Pharmacy of Cluj-Napoca - Discipline of Biochemistry, Cluj-Napoca, Romania. 4. Romanian Academy - Cluj-Napoca Branch, Cluj-Napoca, Romania. 5. Academy of Medical Sciences of Romania - Cluj-Napoca Branch, Cluj-Napoca, Romania. 6. The Gheorghe Benga Foundation, Cluj-Napoca, Romania. 7. The OUTNOBEL Foundation, Cluj-Napoca, Romania.
Abstract
OBJECTIVE: To compare two chromatographic methodologies for determination of plasma phenylalanine (Phe) and their usefulness for diagnosing hyperphenylalaninemia (HPA) and phenylketonuria (PKU). METHODS: The plasma amino acids were isolated and concentrated from blood collected from infants with HPA detected by newborn screening. The plasma Phe was determined in parallel by HPLC and by image-densitometry of 2D-TLC plates. RESULTS: Typical examples of 2D-TLC plates and HPLC chromatograms from infants with HPA and PKU are presented and evaluated. The Phe spot was visible on 2D - TLC plates at Phe concentrations higher than 300 µmol/L. The standard calibration curve traced after image-densitometry of the Phe spot presented high dispersion of values at each concentration of Phe, high SD values, the equation of the curve having a low R-squared value (0.862). In contrast, the standard calibration curve obtained by HPLC shows linearity on the range of concentrations from 100 - 16,000 µmol/L, extremely small SD values, the equation of the curve has a very high R-squared value (0.999). CONCLUSIONS: The HPLC methodology is appropriate to confirm HPA detected by newborn or selective screening of PKU. The 2D - TLC methodology is adequate to detect patients with severe PKU.
OBJECTIVE: To compare two chromatographic methodologies for determination of plasma phenylalanine (Phe) and their usefulness for diagnosing hyperphenylalaninemia (HPA) and phenylketonuria (PKU). METHODS: The plasma amino acids were isolated and concentrated from blood collected from infants with HPA detected by newborn screening. The plasma Phe was determined in parallel by HPLC and by image-densitometry of 2D-TLC plates. RESULTS: Typical examples of 2D-TLC plates and HPLC chromatograms from infants with HPA and PKU are presented and evaluated. The Phe spot was visible on 2D - TLC plates at Phe concentrations higher than 300 µmol/L. The standard calibration curve traced after image-densitometry of the Phe spot presented high dispersion of values at each concentration of Phe, high SD values, the equation of the curve having a low R-squared value (0.862). In contrast, the standard calibration curve obtained by HPLC shows linearity on the range of concentrations from 100 - 16,000 µmol/L, extremely small SD values, the equation of the curve has a very high R-squared value (0.999). CONCLUSIONS: The HPLC methodology is appropriate to confirm HPA detected by newborn or selective screening of PKU. The 2D - TLC methodology is adequate to detect patients with severe PKU.
Authors: Kleopatra H Schulpis; Ioannis Papassotiriou; Maria Vounatsou; George A Karikas; Stylianos Tsakiris; George P Chrousos Journal: J Clin Endocrinol Metab Date: 2004-08 Impact factor: 5.958
Authors: Jerry Vockley; Hans C Andersson; Kevin M Antshel; Nancy E Braverman; Barbara K Burton; Dianne M Frazier; John Mitchell; Wendy E Smith; Barry H Thompson; Susan A Berry Journal: Genet Med Date: 2013-10-10 Impact factor: 8.822