The non-enzymatic reaction between reducing sugars and proteins, known as glycation, has received increased attention from nutritional and medical research. In addition, there is a large interest in obtaining glycoconjugates of pure well-characterized oligosaccharides for biological research. In this study, glycation of bovine serum albumin (BSA) by d-glucose, d-galactose and d-lactose under dry-heat at 60 degrees C for 30, 60, 120, 180 or 240 min was assessed and the glycated products studied in order to establish their biological recognition by lectins. BSA glycation was monitored using gel electrophoresis, determination of available amino groups and lectin binding assays. The BSA molecular mass increase and glycation sites were investigated by mass spectrometry and through digestion with trypsin and chymotrypsin. Depending on time and type of sugar, differences in BSA conjugation were achieved. Modified BSA revealed reduction of amino groups' availability and slower migration through SDS/PAGE. d-galactose was more reactive than d-glucose or d-lactose, leading to the coupling of 10, 3 and 1 sugar residues, respectively, after 120 minutes of reaction. BSA lysines (K) were the preferred modified amino acids; both K256 and K420 appeared the most available for conjugation. Only BSA-lactose showed biological recognition by specific lectins.

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Chitin neoglycoconjugates (BSA-CO) were obtained by the conjugation of bovine serum albumin (BSA) with chitin oligosaccharides (CO) through the Maillard reaction (nonenzymatic glycation). CO produced by acid hydrolysis of chitin were fractionated using an ultrafiltration membrane system (1-3 kDa cutoff). The Maillard reaction was carried out by heating a freeze-dried mixture containing BSA and CO at 60 °C (under 43% relative humidity for 6 and 12 h). BSA-CO were characterized by available amino groups content, intrinsic tryptophan emission spectra, gel electrophoresis, and mass spectrometry. Biological assays included interaction with wheat germ agglutinin (WGA) and with bacterial adhesins of Escherichia coli K88+ and Salmonella choleraesuis. Glycation of BSA was revealed by reduction of available amino groups and fluorescence intensity and also retarded migration through SDS-PAGE. Conjugation of BSA with chitin oligomers appeared to be time dependent and was confirmed by mass spectrometry, by which molecular mass increase for monomers and dimers was observed. Monomers were estimated to contain either one or two glycation sites (at 6 and 12 h of treatment, respectively), with one or two tetrasaccharide units attached. Consequently, dimers showed two or four glycation sites. BSA-CO presented biological recognition by WGA and E. coli K88+ and S. cholerasuis adhesins. The strategy used in this work represents a simple method to obtain glycoconjugates to study applications involving protein-carbohydrate recognition.

Red cell concentrate, serum, albumin, and immunoglobulin fractions were separated from porcine blood collected at a HACCP-implemented (Hazard Analysis and Critical Control Points system-implemented) slaughterhouse in Hermosillo, México. Functional and microbiological properties were examined. Red cell concentrate and serum isolates were high in protein (92 and 85%, respectively) but limited in methionine and isoleucine. Porcine serum and albumin fractions showed emulsifying properties comparable to bovine sources. Porcine immunoglobulins were 96% soluble. All fractions were of excellent microbiological quality. These results indicate the optimal sanitary conditions during slaughter and processing procedures, as well as the potential functionality of porcine protein fractions.

Recently, glyco-therapy is proposed to prevent the interaction of bacterial lectins with host ligands (glycoconjugates). This interaction represents the first step in infection. Neoglycans referred to as PSA-Lac (PSA-Glu (β1-4) Gal) were obtained by conjugation of porcine serum albumin (PSA) with lactose at 80 °C, 100 °C and 120 ºC. Characterization studies of the products showed that PSA could contain 1, 38 or 41 added lactoses, depending on the reaction temperature. These neoglycans were approximately 10 times more glycated than PSA-Lac obtained in previous work. Lactose conjugation occurred only at lysines and PSA-Lac contained terminal galactoses as confirmed by Ricinus communis lectin recognition. Furthermore, Escherichia coli K88+, K88ab, K88ac and K88ad adhesins showed affinity toward all PSA-Lac neoglycans, and the most effective was the PSA-Lac obtained after 100 ºC treatment. In vitro, this neoglycan partially inhibited the adhesion of E. coli K88+ to piglet mucin (its natural ligand). These results provide support for the hypothesis that glycated proteins can be used as an alternative for bioactive compounds for disease prevention.

Glycoconjugates consist of glycans attached to proteins or lipids. Glycans are involved in important biological functions such as trafficking of glycoconjugates, mediation, and modulation of cell adhesion and signaling. This study was conducted to obtain neoglycoconjugates containing a large number of carbohydrates, added through the condensation of reducing sugars with protein amino groups, whose structures were recognized by lectins. Neoglycoconjugates (BSA-Lac) of bovine serum albumin (BSA) with d-lactose were obtained using two sets of glycation conditions, each previously selected by its ability to glycate proteins extensively. The conditions included dry heat at 60 degrees C (for 7, 14, 21, and 28 days) and wet heat in 43% relative humidity (RH) at 50 degrees C (for 5, 10, 15, and 20 h). Products were characterized by gel electrophoresis, tryptophan fluorescence emission spectra, mass spectrometry, free amino group analysis, and their biological recognition established by a galactose-specific lectin and Escherichia coli K88 adhesins. BSA-Lac when compared to BSA revealed an increase in monomer mass due to addition of either 13 (dry heat) or 14 (wet heat) lactoses and formation of polymers (in wet conditions). All BSA-Lac products showed reduced intensity of intrinsic fluorescence, decreased amino groups' availability, and were recognized by Ricinus communis I lectin (RCAI) and by E. coli K88 adhesins. Overall, glycation using both conditions was time-dependent, but greater biorecognition was observed with wet-heat products, due to a higher global glycation and/or to the carbohydrate accessibility. The strategy used in this work represents a simple procedure to obtain glycoconjugates that could be used for recognition studies in biological systems.

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