Advanced glycation end products: a highly complex set of biologically relevant compounds detected by mass spectrometry

Abstract

Structural information on 'AGE-peptides,' a class of substances belonging to advanced glycation end products (AGE) and originating by proteolysis of glycated proteins, was gained through various analytical approaches on the mixture produced by proteinase K digestion of in vitro glycated bovine serum albumin. Both matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS) were employed, and the results were compared with those from conventional spectroscopic methods (UV, fluorescence, gel permeation). The data acquired by the various techniques all depict the digestion mixtures as highly complex, with components exhibiting molecular mass in the range 300-3500 Da. In the analysis of HPLC/ESI-MS data, identification of AGE-peptides was facilitated by 3D mapping. Structural information was gained by means of multiple mass spectrometric experiments.

Mass spectrometric characterization of glycated β-lactoglobulin peptides derived from galacto-oligosaccharides surviving the in vitro gastrointestinal digestion

“…Due to the complexity of the peptides profile and to the incomplete chromatographic separation between nonglycated and glycated peptides, analogous digestion patterns were apparently obtained for unglycated and glycated ␤-LG [13,19]. A similar behavior was observed for other proteins such as bovine serum albumin (BSA) [34] or HSA [14,38]. In good agreement with this, 33 of the 47 unglycated peptides identified in the digest of native ␤-LG were also detected in that of glycated ␤-LG (Table 2).…”

“…Although a large number of ions remained present in both samples, most of those present in the glycated ␤-LG were much less abundant. This fact could be attributed to a sum of multiple facts such as the lower ionization power of glycated peptides, the lower susceptibility to digestion for glycated proteins [14,34], or the partial glycation of Lys-containing and/or N-terminal peptides. Overall, 58 and 23 peptides covering 97% and 63% of the mature ␤-LG sequence could be identified by MALDI-MS in the digests of native and glycated samples, respectively ( Table 1).…”

Analysis of carbohydrates and glycoconjugates by matrix‐assisted laser desorption/ionization mass spectrometry: An update covering the period 2001–2002

“…MALDI analysis of such products from bovine serum albumin following proteinase K digestion has revealed a highly complex mixture with molecular weights in the 300-3,500 Da range (Lapolla et al, 2001a). The presence of AGE-modified proteins has been shown to affect the peptide mixtures produced by proteinase K digestion (Lapolla et al, 2002a).…”

Mass spectrometric study of in vivo production of advanced glycation end‐products/peptides

“…However, published data reveal the enormous complexity of low molecular mass protein fractions present in plasma and serum samples. 13 For these reasons, identification of AGE/peptides directly in the sera of healthy diabetic and nephropathic subjects was believed to be a very difficult task and, consequently, it was thought of interest to undertake a preliminary investigation. This was based on the results described and discussed by Gugliucci and co-workers, 8,9 suggesting that products analogous to AGE/peptides present in plasma samples may also be produced by the enzymatic digestion of in vitro glycated human serum albumin.…”