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Speciation analysis of calcium, iron, and zinc in casein phosphopeptide fractions from toddler milk-based formula by anion exchange and reversed-phase high-performance liquid chromatography–mass spectrometry/flame atomic-absorption spectroscopy

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Abstract

Casein phosphopeptides (CPP) are phosphorylated casein-derived peptides that can be released by in-vitro or in-vivo enzymatic hydrolysis of αs1-casein, αs2-casein, and β-casein (CN). Many of these peptides contain a highly polar acidic sequence of three phosphoseryl groups followed by two glutamic acid residues. These domains are binding sites for minerals such as calcium, iron, and zinc and play an important role in mineral bioavailability. The aim of this study was speciation analysis of calcium, iron, and zinc in CPP fractions from the soluble fraction of a toddler milk-based formula. Methods for CPP separation by anion-exchange high-performance liquid chromatography (AE-HPLC) were combined with CPP identification by reversed-phase high performance liquid chromatography–electrospray ionization mass spectrometry and determination of the calcium, iron, zinc, and phosphorus content of the fractions obtained by AE-HPLC. Calcium and phosphorus were detected in all the analyzed AE-HPLC fractions. Calcium and zinc could be bound to CPP derived from αs1-CN and αs2-CN in fraction 3. Iron could be bound to CPP in fraction 4 in which β-CN(15-34)4P was present with the cluster sequence S(P)S(P)S(P)EE. The results obtained prove the different distribution of calcium, iron, and zinc in heterogeneous CPP fractions.

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Acknowledgements

This study is part of the project AGL2000-1483 financially supported by the FEDER/CICYT (Spain). We thank Hero S.A. (Murcia, Spain) for providing the samples and for financial support of this study. Thanks also to Generalitat Valenciana for the financial support given to the Bionutest group. E. Miquel is the holder of a grant from the Ministerio de Ciencia y Tecnología, Spain.

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Authors and Affiliations

  1. Nutrition and Food Chemistry, Faculty of Pharmacy, University of Valencia, Av. Vicent Andrés Estellés s/n, 46100, Burjassot, Valencia, Spain

    Esther Miquel, Amparo Alegría, Reyes Barberá & Rosaura Farré

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  1. Esther Miquel
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  2. Amparo Alegría
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  3. Reyes Barberá
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  4. Rosaura Farré
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Correspondence to Rosaura Farré.

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Miquel, E., Alegría, A., Barberá, R. et al. Speciation analysis of calcium, iron, and zinc in casein phosphopeptide fractions from toddler milk-based formula by anion exchange and reversed-phase high-performance liquid chromatography–mass spectrometry/flame atomic-absorption spectroscopy. Anal Bioanal Chem 381, 1082–1088 (2005). https://doi.org/10.1007/s00216-004-3002-6

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  • DOI: https://doi.org/10.1007/s00216-004-3002-6

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