Rapid assessment of early glycation products by mass spectrometry |
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Authors: | M Prabhakaram JB Smith BJ Ortwerth |
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Affiliation: | Mason Institute of Ophthalmology, University of Missouri, Columbia 65212, USA. ophthmp@showme.missouri.edu |
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Abstract: | In order to detect the early glycation products, we have reacted a model peptide (t-boc-lys-ala-ala) with L-threose (a degradation product of ascorbic acid) and analyzed the reaction products by a combination of HPLC and mass spectrometry. Amino group modification, as observed by a fluorescamine assay, indicated complete modification after 3 days of incubation with a 10-fold excess of threose. As much as 60% of the adducts were acid labile and only 4% of the adducts could be observed by amino acid analysis. However, Fast atom bombardment mass spectrometry (FABMS) of the samples incubated for 6 hr showed relative molecular masses consistent with the formation of adducts corresponding to the addition of one and two molecules of L-threose to the peptide. Likewise, samples incubated for 12 hr showed peptide adducts with two and three L-threoses. The number of threose molecules added to the peptide was also confirmed from the FABMS analysis by using 1-13C]-threose as the glycating agent. |
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