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Determination of marker pteridines in urine by HPLC with fluorimetric detection and second-order multivariate calibration using MCR-ALS

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Abstract

A liquid chromatographic method has been developed, in combination with the multivariate curve resolution-alternating least squares algorithm (MCR-ALS), for the simultaneous determination of marker pteridines in urine samples. A central composite design has been applied to optimize the factors influencing the separation (buffer concentration, buffer pH, flow rate, oven temperature, mobile-phase composition). A set of 15 calibration samples were randomly prepared, in a concentration range of 0.5–10.5 ng mL−1 for neopterin, biopterin, and pterin; 4.0–8.0 ng mL−1 for xanthopterin; and 0.5–4.5 ng mL−1 for isoxanthopterin. The validation was carried out with fortified urine samples from healthy adults. The optimized conditions were a mobile-phase composition of 10 mM citric buffer at pH 5.44 and acetonitrile (94.5/5.5, v/v), a flow rate of 1.0 mL min−1, and an oven temperature of 25 °C. The detection system consisted of a fast-scanning spectrofluorimeter, which allows obtaining of second-order data matrices containing the fluorescence intensity as a function of retention time and emission wavelength. In this work, MCR-ALS was used to cope with coeluting interferences, on account of the second-order advantage inherent to this algorithm which, in addition, is able to handle data sets deviating from trilinearity, like the high-performance liquid chromatography data analyzed in the present report. The developed approach enabled us to determine five pteridines, some of them with overlapped profiles, reducing the experimental time and reagent consumption. Ratio values for pteridines/creatinine in urine, for infected children with different pathologies, are reported in this work.

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Acknowledgements

The authors are grateful to the Ministerio de Ciencia e Innovación of Spain (Project CTQ2008-06657-C02-01/BQU), cofinanced by the European FEDER funds, CONICET (PIP 112-200801-02988), and Universidad Nacional del Litoral (CAI + D 12-65). Funding from the Junta de Extremadura and European Social Funds (Consolidation Project of Research Group FQM003) is also acknowledged. A. Mancha de Llanos is grateful to the Consejería de Educación y Tecnología de la Junta de Extremadura for a fellowship (DOE 29/06/2006). M.J.C. and H.C.G. are members of the research career from CONICET. The volunteers who provided the urine samples are acknowledged.

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Authors and Affiliations

  1. Department of Analytical Chemistry, University of Extremadura, 06006, Badajoz, Spain

    A. Mancha de Llanos, A. Espinosa-Mansilla & A. Muñoz de la Peña

  2. Laboratory of Analytical Development and Chemometrics (LADAQ), Cathedra of Analytical Chemistry I, Faculty of Biochemistry and Biological Science, University National of Litoral, University City, Santa Fe, S3000ZAA, Argentina

    M. M. De Zan, M. J. Culzoni & H. C. Goicoechea

  3. Department of Sciences and Mathematics Education, University of Extremadura, 06006, Badajoz, Spain

    F. Cañada-Cañada

Authors
  1. A. Mancha de Llanos
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  2. M. M. De Zan
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  3. M. J. Culzoni
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  4. A. Espinosa-Mansilla
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  5. F. Cañada-Cañada
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  6. A. Muñoz de la Peña
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  7. H. C. Goicoechea
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Corresponding authors

Correspondence to A. Muñoz de la Peña or H. C. Goicoechea.

Additional information

Published in the special issue Chemometrics (VII Colloquium Chemiometricum Mediterraneum) with guest editors Marcelo Blanco, Juan M. Bosque-Sendra, and Luis Cuadros-Rodríguez

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de Llanos, A.M., De Zan, M.M., Culzoni, M.J. et al. Determination of marker pteridines in urine by HPLC with fluorimetric detection and second-order multivariate calibration using MCR-ALS. Anal Bioanal Chem 399, 2123–2135 (2011). https://doi.org/10.1007/s00216-010-4071-3

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  • DOI: https://doi.org/10.1007/s00216-010-4071-3

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