Effect of high-intensity ultrasonic time on structural, mechanical, and physicochemical properties of ß-conglycinin (7S)- Transglutaminase (TGase) composite edible films.

The ß-conglycinin (7S) was pre-treated with high-intensity ultrasonic (HIU) and subsequently formed into composite edible films with the transglutaminase (TGase) method. Effects of HIU pretreatment time (0, 5, 10, 15, and 20 min) on the conformation of 7S and structural and application properties of 7S-TGase films were evaluated. The analysis of 7S conformation results revealed that HIU pretreatment for 0-10 min significantly dissociated the 7S, exposed internal hydrophobic groups of protein, increased its intermolecular hydrogen bonds, and altered the protein secondary and tertiary structure. The structural properties of films were evaluated by SEM, XRD, and ATR-FTIR. SEM showed that HIU reduced film wrinkles and cracks and improved unevenness. XRD and ATR-FTIR indicated that the film obtained an enlarged crystallinity, and the amide I and amide II regions of films were peak-shifted which is usually associated with the formation of covalent bonds. Notably, analysis of intermolecular force showed that HIU facilitated the formation of hydrogen bonds, hydrophobic interactions, and ε-(γ-glutamyl) lysine bonds in 7S-TGase films. The above structural changes in 7S and films were beneficial for the application properties of films. Results indicated that 10 min HIU pretreatment effectively improved the mechanical properties and water resistance, reduced water vapor permeability and oxygen permeability, and decreased the opacity of 7S-TGase films. However, the color of the film was not affected by the HIU, with an overall bright and yellowish color.

Effect of High-Intensity Ultrasound Pretreatment on the Properties of the Transglutaminase (TGase)-Induced ß-Conglycinin (7S) Gel.

In this study, we investigated the effects of different high-intensity ultrasound (HIU) pretreatment times (0-60 min) on the structure of ß-conglycinin (7S) and the structural and functional properties of 7S gels induced by transglutaminase (TGase). Analysis of 7S conformation revealed that 30 min HIU pretreatment significantly induced the unfolding of the 7S structure, with the smallest particle size (97.59 nm), the highest surface hydrophobicity (51.42), and the lowering and raising of the content of the α-helix and ß-sheet, respectively. Gel solubility showed that HIU facilitated the formation of ε-(γ-glutamyl)lysine isopeptide bonds, which maintain the stability and integrity of the gel network. The SEM revealed that the three-dimensional network structure of the gel at 30 min exhibited filamentous and homogeneous properties. Among them, the gel strength and water-holding capacity were approximately 1.54 and 1.23 times higher than those of the untreated 7S gels, respectively. The 7S gel obtained the highest thermal denaturation temperature (89.39 °C), G', and G″, and the lowest tan δ. Correlation analysis demonstrated that the gel functional properties were negatively correlated with particle size and the α-helix, while positively with Ho and ß-sheet. By contrast, gels without sonication or with excessive pretreatment showed a large pore size and inhomogeneous gel network, and poor properties. These results will provide a theoretical basis for the optimization of HIU pretreatment conditions during TGase-induced 7S gel formation, to improve gelling properties.

Effects of different dry heating temperatures on the spatial structure and amino acid residue side-chain oxidative modification of soybean isolated proteins.

Spatial structure and amino acid residue side-chain oxidative modification of soybean isolated protein (SPI) at different dry heating temperatures (70, 100, 130, 160 and 190 °C) were investigated, respectively in this study. The results showed that the dry heating promoted the formation of disulfide bonds and oxidative modification of SPI, such as carboxylation and hydroxylation under the below 160 °C. With increasing temperature, ß-sheet and α-helix shifted to random coil and ß-turn. The conformation of SPI changed, the solubility decreased and the particle size became smaller resulting from the combination of protein oxidation and chemical bond redistribution, but the structural integrity of SPI was better ensured below 130 °C. SPI was severely hydrolyzed at 190 °C. These results provide a theoretical basis for the study of protein modification by dry heating, which is a guideline for controlling the degree of protein denaturation in the food industry.

Advanced Glycation End Products, Bone Health, and Diabetes Mellitus.

Advanced glycation end products (AGEs), the compounds resulting from the non-enzymatic glycosylation between reducing sugars and proteins, are derived from food or produced de novo. Over time, more and more endogenous and exogenous AGEs accumulate in various organs such as the liver, kidneys, muscle, and bone, threatening human health. Among these organs, bone is most widely reported. AGEs accumulating in bone reduce bone strength by participating in bone structure formation and breaking bone homeostasis by binding their receptors to alter the proliferation, differentiation, and apoptosis of cells involved in bone remodeling. In this review, we summarize the research about the effects of AGEs on bone health and highlight their associations with bone health in diabetes patients to provide some clues toward the discovery of new treatment and prevention strategies for bone-related diseases caused by AGEs.

Effect of Baking Temperature and Time on Advanced Glycation End Products and Polycyclic Aromatic Hydrocarbons in Beef.

ABSTRACT: Beef is an important red meat that contains essential nutrients for human growth and development. Baking is a popular beef cooking method. Temperature and time play key roles in the final quality of beef. How temperature and time affect the changes of nutrients and the formation of harmful products in beef is not clear. The purpose of this study was to measure the content of water, fat, protein, ash, nitrite, total volatile base nitrogen, advanced glycation end products (AGEs) and their precursors, and polycyclic aromatic hydrocarbons (PAHs) at different temperatures (150, 190, 230, 270, and 310°C) for 20 min and at 190°C for different times (10, 20, and 30 min), so as to discuss the effect of different temperatures and times on beef nutrients and harmful products. The results showed that the moisture content of beef decreased with increased baking temperature and time, resulting in the increase of the relative content of fat, protein, and ash. The content of total volatile base nitrogen increased continuously. Compared with the control group, the content of glyoxal in beef decreased, whereas the content of methylglyoxal, pentosidine, and fluorescent AGEs increased, indicating the continuous accumulation of AGEs in beef. A total of 13 PAHs were identified by gas chromatography-mass spectrometry. The concentrations of 13 PAHs in beef increased with increases in baking temperature and time. The concentrations of BkP and BaP, which are the most carcinogenic to humans, were 0.36 and 0.35 µg/kg in raw meat, respectively; these were increased by high temperature and long baking times. After beef was baked at 270 and 310°C for 20 min, the concentration of BkP increased to 9.49 and 5.66 µg/kg, respectively, and the concentration of BaP increased to 5.45 and 4.42 µg/kg, respectively. After baking at 190°C for 30 and 40 min, the concentration of BkP increased to 4.81 and 24.20 µg/kg, respectively, and the concentration of BaP increased to 3.85 and 17.79 µg/kg, respectively.

Effect of roasting temperature on lipid and protein oxidation and amino acid residue side chain modification of beef patties.

Beef is rich in nutrients and is one of the most important ingredients in the world. But in the process of cooking and heating, the nutrients of beef will change to varying degrees. How temperature affects the oxidation of lipids and proteins in beef, and the modification of amino acid residues is unclear. This study intended to heat beef at different roasting temperatures (150 °C, 190 °C, 230 °C, 270 °C, 310 °C), measure parameter including colour, peroxide value (PV), thiobarbituric acid-reactive substances (TBARS), thiol and carbonyl content, protein solubility, tryptophan and Schiff base content, protein molecular weight distribution and modification of amino acid residues to discussed the effects of different temperatures on the lipid and protein oxidation of beef patties, as well as the modification of amino acid residues. The results showed that the values of L* and b* increased with the temperature increased, and the values of a* decreased. With the increase of temperature, the lipid oxidation indexes PV and TBARS, Schiff base and carbonyl content also increased, and the thiol content and protein solubility decreased significantly (p < 0.001). SDS-PAGE showed that the band of myosin heavy chain (MHC, 220 kDa) was significantly degraded, while the band of actin (42 kDa) was still clearly visible. The analysis of UPLC-MS/MS results found that the aromatic amino acid residues in all samples were oxidized to a certain extent, especially tryptophan. Other oxidative modifications, including α-amiooadipic acid (AAA), hydroxyethyl lysine (CEL) and malondialdehyde (MDA), were only present in roasted samples and not in raw meat. The results suggested that lipid oxidation and protein oxidation were closely related to colour parameters. The oxidation of proteins and lipids was aggravated at higher temperature. Amino acid side chains were also modified at high temperature, and this change was particularly evident in aromatic amino acids. These results provided new insights for the oxidation of proteins and lipids of beef and the modification level of amino acid residues under high temperature conditions, which will help us to improve the cooking quality of meat foods.

Immunization with recombinant Salmonella expressing SspH2-EscI protects mice against wild type Salmonella infection.

BACKGROUND: Enhancing caspase-1 activation in macrophages is helpful for the clearance of intracellular bacteria in mice. Our previous studies have shown that EscI, an inner rod protein of type III system in E. coli can enhance caspase-1 activation. The purpose of this study was to further analyze the prospect of EscI in the vaccine design. RESULTS: A recombinant Salmonella expressing SspH2-EscI fusion protein using the promotor of Salmonella effector SspH2, X4550(pYA3334-P-SspH2-EscI), was constructed. A control recombinant Salmonella expressing SspH2 only X4550(pYA3334-P-SspH2) was also constructed. In the early stage of in vitro infection of mouse peritoneal macrophages, X4550(pYA3334-P-SspH2-EscI) could significantly (P < 0.05) enhance intracellular caspase-1 activation and pyroptotic cell death of macrophages, when compared with X4550(pYA3334-P-SspH2). Except for the intracellular pH value, the levels of reactive oxygen species, intracellular concentration of calcium ions, nitric oxide and mitochondrial membrane potential in macrophages were not significantly different between the cells infected with X4550(pYA3334-P-SspH2-EscI) and those infected with X4550(pYA3334-P-SspH2). Besides, only lower inflammatory cytokines secretion was induced by X4550(pYA3334-P-SspH2-EscI) than X4550(pYA3334-P-SspH2). After intravenous immunization of mice (1 × 106 cfu/mouse), the colonization of X4550(pYA3334-P-SspH2-EscI) in mice was significantly limited at one week post immunization (wpi), when compared with X4550(pYA3334-P-SspH2) (P < 0.05). The population of activated CD8+T lymphocytes in mouse spleens induced by X4550(pYA3334-P-SspH2-EscI) was lower than that induced by X4550(pYA3334-P-SspH2) at 2-3 wpi, and the ratio of CD4+T cells to CD8+T cells decreased. The blood coagulation assay indicated that no significant difference was found between X4550(pYA3334-P-SspH2-EscI) and uninfected control, while X4550(pYA3334-P-SspH2) could induce the quick coagulation. Notably, immunization of X4550(pYA3334-P-SspH2-EscI) could limit the colonization of challenged Salmonella strains in the early stage of infection and provide more effective protection. CONCLUSION: The activation of caspase-1 in macrophages by EscI can be used in the design of live attenuated Salmonella vaccine candidate.

Recombinant Salmonella expressing SspH2-EscI fusion protein limits its colonization in mice.

BACKGROUND: Activation of inflammasome contributes to the clearance of intracellular bacteria. C-terminus of E. coli EscI protein can activate NLRC4 (NLR family, CARD domain containing-4) inflammasome in macrophages. The purpose of this study was to determine if activation of NLRC4 inflammasome by EscI can reduce the colonization of Salmonella in mice. RESULTS: A recombinant S. typhimurium strain expressing fusion protein of the N-terminal SspH2 (a Salmonella type III secretion system 2 effector) and C-terminal EscI was constructed and designated as X4550(pYA3334-SspH2-EscI). In vitro assay showed that X4550(pYA3334-SspH2-EscI) significantly enhanced IL-1ß and IL-18 secretion (P < 0.05) and pyroptotic cell death of mouse peritoneal macrophages, compared with those infected with control strain, X4550(pYA3334-SspH2). In vivo studies showed that colonization of X4550(pYA3334-SspH2-EscI) in both spleen and liver were significantly lower than that of X4550(pYA3334-SspH2) (P < 0.05). The bacterial counts of X4550(pYA3334-SspH2-EscI) in mice decreased, while those of X4550(pYA3334-SspH2) increased over the time after infection. Additionally, X4550(pYA3334-SspH2-EscI) induced a less pathological alteration in spleen and liver than X4550(pYA3334-SspH2). CONCLUSION: Fusion protein SspH2-EscI may be translocated into macrophages and activate NLRC4 inflammasome, which limits Salmonella colonization in spleen and liver of mice.

[Relationship between interferon-α conformation and its anti-viral activity determined by circular dichroism and flow cytometry].

The relationship between the conformation of interferon-α (IFN-α) and its anti-viral activity were analyzed by circular dichroism (CD) and flow cytometry (FCM) techniques. The recombinant human IFN-α (rIFN-α2b and rIFN-α2a) were used. CD spectra from 190 nm to 240 nm indicated that two the IFN-α showed stable secondary structure at 65 degrees C, but unstable when the temperature was above 65 degrees C, and the change was irreversible. FCM data of the anti-viral activity of IFN-α indicated that the change of its secondary structures partly weakened its anti-viral activity. The rIFN-α2b and rIFN-α2a showed the same phenomenon. These data indicated that the conformation of IFN-α is one of the factors to influence its anti-viral activity and the combination of CD and FCM is a good method to analyze the relationship between the conformation of protein drugs and their biological activities in single cell level.