β-Lactoglobulin (β-LG) was modified and conjugated to xylobiose using the Maillard reaction. The antioxidant activity of the Maillard reaction product, β-LG-xylobiose, was measured in vitro and compared to that of conjugated β-LG-lactose. The reaction for 7 days led to conjugated β-LG-xylobiose with a relative molecular mass ranging between 19 and 22 kDa based on SDS-PAGE analysis. It is confirmed that xylobiose bound to β-LG by gas-liquid chromatography. One milligram of conjugated β-LG-xylobiose contains 108 μg of xylobiose, while the available ε-amino group content decreased to 40% after the Maillard reaction. Conjugated β-LG-xylobiose had a higher radical scavenging activity than free β-LG. As the modification by xylobiose had a higher efficiency than the modification by lactose, it is found that xylobiose is a useful oligosaccharide for protein modification. Furthermore, the radical scavenging activity of β-LG was improved by modification with xylobiose.
Lactosucrose (4G-β-D-galactosylsucrose, LS) is suggested to be an oligosaccharide required for the proliferation of Bifidobacteria in the intestine. We have examined the dietary effects of LS on the intestinal immune function of mice. BALB/c mice were fed with 2 and 5% LS for 4 weeks, and the intestinal mucosal immune responses were determined. In the 2 and 5% LS fed groups, the amounts of IgA in feces and in cecum contents were significantly increased. In addition, IgA, transforming growth factor-β1 (TGF-β1) and interleukin-6 (IL-6) secretion by Peyer’s patch (PP) cells were enhanced in LS fed mice. In LS fed mice, pH in the cecum was decreased. LS, in addition, suppressed serum IgG1. These results suggest that LS supplementation changes the intestinal environment of microflora, and indirectly enhances the immune function in the gut, and suppresses the systemic immune response to the dominant type 2 helper T (Th2).
Acid-treated potato starch (ATS), ε-poly(L-lysine) (PL) and glucose/fructose stearic acid monoester (GE/FE) were conjugated by the Maillard reaction to prepare ATS-PL-GE/FE and PL-GE/FE conjugates having molar ratios of ATS:PL:GE/FE=2.2:1.0:1.2 and 1.9:1.0:0.86, and PL:GE/FE=1.0:3.3. These conjugates exhibited an excellent emulsifying ability tolerant of acidic pH and NaCl, and had antibacterial activity comparable to that of free PL against Staphylococcus aureus and Candida utilis, although somewhat less activity against Escherichia coli than that of free PL. The conjugates contributed to a specific control of the gelatinization and retrogradation of potato starch, appearing to markedly reduce the viscosity of the paste, and to augment the absorbance of the retrograded paste. The conjugates resulted in little change in the degree of gelatinization during retrogradation, and the ATS-PL-GE/FE conjugate increased the reconstructed crystalline content, while the PL-GE/FE conjugate reduced it.
Polymerization processes of 4´-hydroxyphenyl α-glucoside (α-Arb) and 4´-hydroxyphenyl β-glucoside (arbutin, β-Arb) catalyzed by horseradish peroxidase (HRP) were analyzed in detail. A calculated model based on the coupling of the free radicals formed from the substrates with every degree of polymerization reproduced successfully most of the experimental results: An increase of the polymer with a decrease of the monomer and a temporal accumulation of the dimer responding to the addition of H2O2. Reaction controlled at neutral pH improved the polymer yields by preventing partial hydrolysis of the glucose residues. A sequential addition of divided portions of H2O2 also increased the yields considerably compared to the single addition, although the yields with too low enzyme activities could not be restored. The main chain structure of the α-Arb polymer was the same as that of the β-Arb polymer which had linkages at the 3´- and 5´-positions of hydroquinone moieties. The polymers showed molecular weight distribution at 1-25 kDa in gel permeation chromatography, when maltooligosaccharides and pullulan were used as standards. Mass spectrometric analysis of deglycosylated polymers suggested that the α-Arb polymer contained smaller components more than the β-Arb polymer. Peroxidases from horseradish, soybean and Arthromyces polymerized β-Arb faster than α-Arb. HRP polymerized the two glucosides faster than the other two peroxidases. Oxidation potentials were determined to be 821 and 835 mV for α-Arb and β-Arb, respectively, by cyclic voltammetry, suggesting almost the same electrochemical reactivity. A higher reactivity of β-Arb in the enzymatic polymerization was thus attributed not to electrochemical reactivity of the substrates, but to stereoselectivity of the enzymes.
Fresh spears of asparagus were stored in the dark at 4, 10 or 20°C for 2 weeks. During storage contents of glucose, fructose, sucrose, 1-kestose, neokestose and nystose, and activities of invertase, 1-kestose hydrolyzing enzyme (1-KHE), sucrose: sucrose 1-fructosyltransferase (1-SST), fructan: fructan 1-fructosyltransferase (1-FFT) and fructan: fructan 6G-fructosyltransferase (6G-FFT) were determined in the top, middle and bottom portions of the spears. A gradient was observed, from the bottom to the top, for glucose, fructose and sucrose which constitute the major proportion of carbohydrates, while fructooligosaccharides, neokestose and nystose, exhibited low levels. Glucose and fructose varied significantly during storage, while sucrose was stable. The average variations were from 7.8 to 12.21 mg/g FW in the middle portion and 7.88 to 13.52 mg/g FW in the bottom portion for glucose and fructose, respectively. 1-Kestose and nystose increased at the end of the storage period and this increase was more apparent at 20°C. Invertase activity showed similar variation at 4 and 10°C but increased sharply after 2 days, before decreasing abruptly after 1 week of storage, while 1-kestose hydrolyzing activity showed a similar pattern to that of invertase activity. 1-SST did not vary in the bottom portion but initially increased in the middle and top portions. 1-FFT was high in the top portion and decreased during storage, while in the middle and bottom portions its activity varied slightly. The variation of 6G-FFT activity was similar to that of 1-FFT, however, the level of 6G-FFT was higher, and the 6G-FFT to 1-FFT activity ratio was temperature independent. These results suggest that short fructooligosaccharides and their metabolizing enzymes could play a role of balance between the hydrolysis and synthesis activities of carbohydrates. The high content of sugars may also extend the rapid decline of sugars in the top portion of the spears.
Novel tri- and tetra-saccharides were synthesized by glucosyltransfer from β-D-glucose 1-phosphate (β-D-G1P) to palatinose using Thermoanaerobacter brockii kojibiose phosphorylase. There saccharides were isolated using carbon-Celite column chromatography and preparative high performance liquid chromatography. Gas liquid chromatography analysis of methyl derivatives, MALDI-TOF MS and NMR measurements were used for structural confirmation of the saccharides. The 1H and 13C NMR signals of the saccharides were assigned using 2D-NMR including COSY, HSQC, HSQC-TOCSY and HMBC. These oligosaccharides were identified as 2G-α-D-glucopyranosyl-palatinose; O-α-D-glucopyranosyl-(1→2)-O-α-D-glucopyranosyl-(1→6)-D-fructofuranose and 2G(2-α-D-glucopyranosyl)2-palatinose; O-α-D-glucopyranosyl-(1→2)-O-α-D-glucopyranosyl-(1→2)-O-α-D-glucopyranosyl-(1→6)-D-fructofuranose.