Journal article


Relative Quantification of N-(Carboxymethyl)lysine, Imidazolone A, and the Amadori Product in Glycated Lysozyme be MALDI-TOF Mass Spectrometry


Publication Details
Author(s): Kislinger T, Humeny A, Peich C, Zhang X, Niwa T, Pischetsrieder M, Becker CM
Publisher: American Chemical Society
Publication year: 2003
Journal issue: 51
Pages range: 51-57
ISSN: 0021-8561

Abstract

The nonenzymatic glycation of proteins by reducing sugars, also known as the Maillard reaction, has received increasing recognition from nutritional science and medical research. In this study, we applied matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) to perform relative and simultaneous quantification of the Amadori product, which is an early glycation product, and of Nε-(carboxymethyl)lysine and imidazolone A, two important advanced glycation end products. Therefore, native lysozyme was incubated with D-glucose for increasing periods of time (1, 4, 8, and 16 weeks) in phosphate-buffered saline pH 7.8 at 50 °C. After enzymatic digestion with endoproteinase Glu-C, the N-terminal peptide fragment (m/z 838; amino acid sequence KVFGRCE) and the C-terminal peptide fragment (m/z 1202; amino acid sequence VQAWIRGCRL) were used for relative quantification of the three Maillard products. Amadori product, Nε-(carboxymethyl)lysine, and imidazolone A were the main glycation products formed under these conditions. Their formation was dependent on glucose concentration and reaction time. The kinetics were similar to those obtained by competitive ELISA, an established method for quantification of Nε-(carboxymethyl)lysine and imidazolone A. Inhibition experiments showed that coincubation with Nα-acetylargine suppressed formation of imidazolone A but not of the Amadori product or Nε-(carboxymethyl)lysine. The presence of Nα-acetyllysine resulted in the inhibition of lysine modifications but in higher concentrations of imidazolone A. o-Phenylenediamine decreased the yield of the Amadori product and completely inhibited the formation of Nε-(carboxymethyl)lysine and imidazolone A. MALDI-TOF-MS proved to be a new analytical tool for the simultaneous, relative quantification of specific products of the Maillard reaction. For the first time, kinetic data of defined products on specific sites of glycated protein could be measured. This characterizes MALDI-TOF-MS as a valuable method for monitoring the Maillard reaction in the course of food processing.



How to cite
APA: Kislinger, T., Humeny, A., Peich, C., Zhang, X., Niwa, T., Pischetsrieder, M., & Becker, C.-M. (2003). Relative Quantification of N-(Carboxymethyl)lysine, Imidazolone A, and the Amadori Product in Glycated Lysozyme be MALDI-TOF Mass Spectrometry. Journal of Agricultural and Food Chemistry, 51, 51-57. https://dx.doi.org/10.1021/jf020768y

MLA: Kislinger, Thomas, et al. "Relative Quantification of N-(Carboxymethyl)lysine, Imidazolone A, and the Amadori Product in Glycated Lysozyme be MALDI-TOF Mass Spectrometry." Journal of Agricultural and Food Chemistry 51 (2003): 51-57.

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