Development of a stable isotope dilution assay for an accurate quantification of protein-bound N-epsilon-(1-deoxy-D-fructos-1-yl)-L-lysine using a C-13-labeled internal standard

Citation
F. Vinale et al., Development of a stable isotope dilution assay for an accurate quantification of protein-bound N-epsilon-(1-deoxy-D-fructos-1-yl)-L-lysine using a C-13-labeled internal standard, J AGR FOOD, 47(12), 1999, pp. 5084-5092
Citations number
21
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Agricultural Chemistry","Chemistry & Analysis
Journal title
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
ISSN journal
0021-8561 → ACNP
Volume
47
Issue
12
Year of publication
1999
Pages
5084 - 5092
Database
ISI
SICI code
0021-8561(199912)47:12<5084:DOASID>2.0.ZU;2-4
Abstract
Syntheses of the labeled Amadori compound [C-13(6)]-N-epsilon-(1-deoxy-D-fr uctos-1-yl)-L-lysine ([C-13(6)]-DFLys) and the labeled glycated tetrapeptid e Ala-[C-13(6)]-DFLys-Leu-Gly are presented. The compounds were used in the development of stable isotope dilution assays for the quantification of th e degree of glycosylation of bovine serum albumin treated for 20 min at 95 degrees C in the presence of glucose. The experiments revealed that the use of the labeled standards in combination with LC/MS allowed the exact quant ification of protein-bound DFLys with the high recovery rate of 95% (at a s pike level of 150 nmol/mg of protein) and a low detection limit of 5 nmol/m g of protein. The data revealed, however, that DFLys is significantly degra ded during the enzymic hydrolysis of the protein backbone generally needed in the quantification procedure and, furthermore, incomplete digestion of t he protein was observed. Both sources of errors were clearly overcome by us ing in particular the labeled peptide as the internal standard.