Quantification of N-acetyl- and N-glycolylneuraminic acids by a stable isotope dilution assay using high-performance liquid chromatography-tandem mass spectrometry |
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Authors: | Allevi Pietro Femia Eti Alessandra Costa Maria Letizia Cazzola Roberta Anastasia Mario |
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Institution: | Dipartimento di Chimica, Biochimica e Biotecnologie per la Medicina, Università di Milano, via Saldini 50, I-20133 Milan, Italy. pietro.allevi@unimi.it |
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Abstract: | The present report describes a method for the quantification of N-acetyl- and N-glycolylneuraminic acids without any derivatization, using their (13)C(3)-isotopologues as internal standards and a C(18) reversed-phase column modified by decylboronic acid which allows for the first time a complete chromatographic separation between the two analytes. The method is based on high-performance liquid chromatographic coupled with electrospray ion-trap mass spectrometry. The limit of quantification of the method is 0.1mg/L (2.0ng on column) for both analytes. The calibration curves are linear for both sialic acids over the range of 0.1-80mg/L (2.0-1600ng on column) with a correlation coefficient greater than 0.997. The proposed method was applied to the quantitative determination of sialic acids released from fetuin as a model of glycoproteins. |
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