Characterization by Ionization Mass Spectrometry of Lactosyl β-Lactoglobulin Conjugates Formed During Heat Treatment of Milk and Whey and Identification of One Lactose-Binding Site

Abstract

The extent of the early stage of the Maillard-type reaction that impaired functional properties of whey proteins was evaluated by electrospray ionization mass spectrometry. Under conditions of mild heat treatment (63 degrees C for 20 s) applied to milk before whey separation at room temperature 23 degrees C), a modification of the relative molecular mass of beta-lactoglobulin (beta-LG) was observed that differed from that of the native form by 324. This specific modification of beta-LG occurred in acidified whey as well as in sweet whey and increased with the extent of the heat treatment. Incubation of purified beta-LG dissolved in milk ultrafiltration permeate or in lactose solution at 50 to 80 degrees C demonstrated the presence of a lactosyl residue that was covalently bound to beta-LG; beta-casein, used as a control, showed no mass modification. Studies of kinetics showed that a maximum of 35% of the beta-LG was lactosyl-beta-LG conjugate after heat treatment at 70 degrees C for 1 h. This study provides the first direct evidence of specific lactosylation of beta-LG during the initial stage of the Maillard reaction. One of the first lactose-binding sites was identified as a Lys47 by protease mapping and analysis by means of on-line liquid chromatography combined with mass spectrometry. In addition, collision-activated dissociation performed on the lactosylated peptide beta-LG (f 46-51) showed the rearrangement reactions occurring during the fragmentation process by electrospray. A mechanism is proposed.

Protein composition and polymorphism in the milk of Skopelos goats

“…The mass 18 197 ± 1 detected in the β-Lg peaks of the milks of the present study corresponds to the theoretical mass of this goat protein (18 191.3). In some samples a mass of 18 523 ± 1 was detected in the front part of the β-Lg peak that is consistent with a covalent linkage of lactosyl residue to the protein [24,48]. No protein variants were observed, in accordance with the previous works that report only silent alleles for goat α-La [11] and β-Lg loci [2,19,36,49].…”

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

“…Figure 3 illustrates the resulting total ion current (TIC) chromatogram for glycated and unglycated digested proteins. 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].…”

Functional Improvements in Food Proteins in Multiple Aspects by Conjugation with Saccharides: Case Studies of .BETA.-Lactoglobulin-Acidic Polysaccharides Conjugates.

“…In Fig. 10 Leonil et al, 1997;Fogliano et al, 1998;Hattori et al, 2000a;Siciliano et al, 2000). The response to the epitope regions in the vicinity of these Lys residues in each conjugate prepared in our study was lowered in the three strains of mice.…”