Effects of Dityrosine on Lactic Acid Metabolism in Mice Gastrocnemius Muscle During Endurance Exercise via the Oxidative Stress-Induced Mitochondria Damage.

Dityrosine (Dityr) has been detected in commercial food as a product of protein oxidation and has been shown to pose a threat to human health. This study aims to investigate whether Dityr causes a decrease in lactic acid metabolism in the gastrocnemius muscle during endurance exercise. C57BL/6 mice were administered Dityr or saline by gavage for 13 weeks and underwent an endurance exercise test on a treadmill. Dityr caused a severe reduction in motion displacement and endurance time, along with a significant increase in lactic acid accumulation in the blood and gastrocnemius muscle in mice after exercise. Dityr induced significant mitochondrial defects in the gastrocnemius muscle of mice. Additionally, Dityr induced serious oxidative stress in the gastrocnemius muscle, accompanied by inflammation, which might be one of the causes of mitochondrial dysfunction. Moreover, significant apoptosis in the gastrocnemius muscle increased after exposure to Dityr. This study confirmed that Dityr induced oxidative stress in the gastrocnemius muscle, which further caused significant mitochondrial damage in the gastrocnemius muscle cell, resulting in decreased capacity of lactic acid metabolism and finally affected performance in endurance exercise. This may be one of the possible mechanisms by which highly oxidized foods cause a decreased muscle energy metabolism.

Soybean-derived antihypertensive hydrolysates attenuate Ang II-induced renal damage by modulating MAPK and NF-κB signaling pathways.

Hypertension-induced kidney injury is considered a vital consequence of long-term and uncontrolled hypertension, which is commonly associated with an excessive accumulation of angiotensin II (Ang II) from hyperactivated RAS. Antihypertensive peptides have a significant effect on blood pressure regulation, but few studies have focused on the ameliorative function of antihypertensive peptides on renal injury. This study explored the effects of soybean protein-derived hydrolysate (SPH) on SHR and Ang II-induced HK-2 cells. SPH significantly attenuated blood pressure and alleviated renal pathological injury in SHRs after oral gavage administration. According to the pathological results, the kidneys of SHRs showed inflammation and SPH attenuated inflammatory cell infiltration in the kidneys of SHRs. Immunohistochemical analysis further revealed that SPH inhibited MCP-1 expression and increased Nrf2 expression in the kidneys. An in vitro HK-2 cell model demonstrated that SPH exhibited optimal activity for reducing Ang II-induced inflammatory cytokines and ROS overproduction. Mechanistically, SPH was observed to regulate MAPK/JNK and NF-κB signaling pathways. These findings indicate that potent antihypertensive SPH significantly ameliorates hypertension-induced kidney damage.

Modulating in vitro digestion of whey protein cold-set emulsion gels via gel properties modification with gallic acid and EGCG.

Gallic acid (GA) and epigallocatechin gallate (EGCG), cooperated at varied ratios (1:0, 3:1, 1:1, 1:3, and 0:1), were employed to modify gel properties of calcium induced-whey protein emulsion gel. The effects of GA/EGCG on emulsion morphology, as well as gel properties and in vitro digestive behavior of the emulsion gels were investigated. Compared with emulsions without phenolics, GA/EGCG induced slightly smaller particle size and stronger electrostatic repulsion between emulsion droplets. Moreover, GA/EGCG, notably at a ratio of 3:1, promoted electrostatic and hydrophobic interactions between protein molecules and the formation of a compact and filamentous gel microstructure, resulting in a remarkable increment in the gel strength (up to 106 %). Furthermore, in vitro oral digestion, dynamic gastric digestion (using an artificial gastric digestive system, AGDS), and intestinal digestion of the emulsion gels were simulated. Particle size and protein hydrolysis results revealed that GA/EGCG was prone to weaken the physical disintegration of gels, reduce protein hydrolysis, and enhance the stability of emulsified oil droplets during dynamic gastric digestion. As a consequence, delayed release of oil droplets was observed in the gels and more free fatty acids were released in the intestinal digestion, particularly in the gel with GA/EGCG (3:1). These findings would provide novel strategies for application of phenolic compounds in developing protein gel-based delivery systems.

Dityrosine Aggravates Hepatic Insulin Resistance in Obese Mice by Altering Gut Microbiota and the LPS/TLR4/NF-κB Inflammatory Pathway.

SCOPE: Dityrosine is the main product of protein oxidation, which has been proved to be a threat to human health. This study aims to investigate whether dityrosine exacerbates insulin resistance by inducing gut flora disturbance and associated inflammatory responses. METHODS AND RESULTS: Mice fed with normal diet or high-fat diet (HFD) received daily gavage of dityrosine (320 µg kg-1 BW) or saline for consecutive 13 weeks. The effects of dityrosine on gut microbiota are verified by in vitro fermentation using fecal microbiota from db/m mice and db/db mice. As a result, dityrosine causes the insulin resistance in mice fed normal diet, and aggravates the effects of HFD on insulin sensitivity. Dityrosine increases the levels of lipopolysaccharide (LPS), lipopolysaccharide-binding protein (LBP), toll-like receptor 4 (TLR4), nuclear factor kappa-B (NF-κB), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-8 (IL-8) but decreases levels of interleukin-10 (IL-10) in the plasma of CON and HFD-fed mice. The changes of gut flora composition caused by dityrosine are significantly correlated with the changes of inflammatory biomarkers. CONCLUSION: The effects of dityrosine on insulin resistance may be attributed to the reshaping of the gut microbiota composition and promoting the activity of the LPS/TLR4/NF-κB inflammatory pathway in HFD-induced obese individuals.

Orally administered octacosanol improves liver insulin resistance in high-fat diet-fed mice through the reconstruction of the gut microbiota structure and inhibition of the TLR4/NF-κB inflammatory pathway.

1-Octacosanol (Octa) is reported to possess many physiological properties. However, its relative mechanism has not been illustrated yet. Herein, we aimed to investigate the effect of Octa on insulin resistance in mice fed with a high fat diet (HFD) and used an in vitro simulated gastrointestinal tract to analyze its digestive behavior. The effects of Octa on the gut microbiota were verified by in vitro fermentation using the mouse fecal microbiota. As a result, the Octa monomer was digested into shortened saturated and unsaturated fatty acids (C10-C24) in the simulated gastrointestinal tract. Octa improved the fasting blood glucose (FBG), insulin resistance (IR), plasma lipids, and inflammatory response in HFD-fed mice in a dose-dependent manner. This study also suggested that a high-dose of Octa effectively decreased the levels of toll-like receptor 4 (TLR4), nuclear factor kappa-B (NF-κB), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) in the plasma of HFD-fed mice. Octa improved the oxidative stress induced by a HFD and increased the expression of the Nrf2/ARE signaling pathway. Importantly, Octa reshaped gut microbiota through decreasing Firmicutes content and increasing Bacteroidota and Verrucomicrobiota contents at the phylum level, and the changes of intestinal flora structure caused by Octa were significantly correlated with the changes of inflammatory biomarkers. In conclusion, the effects of Octa on insulin resistance might be attributed to the reconstruction of the gut microbiota structure and inhibition of the TLR4/NF-κB inflammatory pathway in HFD-induced obese individuals.

Dityrosine in food: A review of its occurrence, health effects, detection methods, and mitigation strategies.

Protein and amino acid oxidation in food products produce many new compounds, of which the reactive and toxic compound dityrosine, derived from oxidized tyrosine, is the most widely studied. The high reactivity of dityrosine enables this compound to induce oxidative stress and disrupt thyroid hormone function, contributing to the pathological processes of several diseases, such as obesity, diabetes, cognitive dysfunction, aging, and age-related diseases. From the perspective of food safety and human health, protein-oxidation products in food are the main concern of consumers, health management departments, and the food industry. This review highlights the latest research on the formation pathways, toxicity, detection methods, occurrence in food, and mitigation strategies for dityrosine. Furthermore, the control of dityrosine in family cooking and food-processing industry has been discussed. Food-derived dityrosine primarily originates from high-protein foods, such as meat and dairy products. Considering its toxicity, combining rapid high sensitivity dityrosine detection techniques with feasible control methods could be an effective strategy to ensure food safety and maintain human health. However, the current dityrosine detection and mitigation strategies exhibit some inherent characteristics and limitations. Therefore, developing technologies for rapid and effective dityrosine detection and control at the industrial level is necessary.

Real-Time Profiling and Distinction of Lipids from Different Mammalian Milks Using Rapid Evaporative Ionization Mass Spectrometry Combined with Chemometric Analysis.

The price of mammalian milk from different animal species varies greatly due to differences in their yield and nutritional value. Therefore, the authenticity of dairy products has become a hotspot issue in the market due to the replacement or partial admixture of high-cost milk with its low-cost analog. Herein, four common commercial varieties of milk, including goat milk, buffalo milk, Holstein cow milk, and Jersey cow milk, were successfully profiled and differentiated from each other by rapid evaporative ionization mass spectrometry (REIMS) combined with chemometric analysis. This method was developed as a real-time lipid fingerprinting technique. Moreover, the established chemometric algorithms based on multivariate statistical methods mainly involved principal component analysis, orthogonal partial least squares-discriminant analysis, and linear discriminant analysis as the screening and verifying tools to provide insights into the distinctive molecules constituting the four varieties of milk. The ions with m/z 229.1800, 243.1976, 257.2112, 285.2443, 299.2596, 313.2746, 341.3057, 355.2863, 383.3174, 411.3488, 439.3822, 551.5051, 577.5200, 628.5547, 656.5884, 661.5455, 682.6015, and 684.6146 were selected as potential classified markers. The results of the present work suggest that the proposed method could serve as a reference for recognizing dairy fraudulence related to animal species and expand the application field of REIMS technology.

Real-time authentication of minced shrimp by rapid evaporative ionization mass spectrometry.

Minced shrimp is popular seafood due to its delicious flavor and nutritional value. However, the biological species of raw material of minced shrimp are not distinguished by naked eyes after processing. Thus, an in situ and real-time minced shrimp authentication method was established using iKnife rapid evaporative ionization mass spectrometry (REIMS) based lipidomics. The samples were analyzed under ambient ionization without any tedious preparation step. Seven economic shrimp samples were tested, whose phenotypes were used to develop a real-time recognition model. A total of 19 fatty acids and 45 phospholipid molecular species were efficiently identified and statistically analyzed by multivariate statistical analysis. The results showed that the seven shrimp species were well distinguished, and the most contributing ions at m/z 255.2, 279.2, 301.2, 327.2, 699.5, 742.5, etc., were revealed by variable importance in projection. The proposed iKnife REIMS showed excellent performance in minced shrimp authentication.

Preparation of Gum Arabic-Maltose-Pea Protein Isolate Complexes for 1-Octacosanol Microcapsule: Improved Storage Stability, Sustained Release in the Gastrointestinal Tract, and Its Effect on the Lipid Metabolism of High-Fat-Diet-Induced Obesity Mice.

1-Octacosanol (Octa) is a natural compound with several beneficial properties. However, its poor water solubility and metabolism in the digestive tract reduce its efficacy. The Octa-GA-Malt-PPI microcapsule was prepared as follows: gum Arabic (GA):maltose (Malt):pea protein isolate (PPI) = 2:1:2; core:shell = 1:7.5; emulsification temperature 70 °C; pH 9.0. An in vitro simulated gastrointestinal tract was used to analyze the digestion behavior. C57BL/6 mice were selected to establish an obesity model induced by a high-fat diet (HFD) to evaluate the effect of Octa monomer and the microcapsule. The diffusivity in water and storage stability of Octa improved after encapsulation. The microcapsule was ascribed to electrostatic interactions, hydrogen bonding, and hydrophobic interactions. The sustained release of Octa from the microcapsule was observed in a simulated gastrointestinal tract. Compared with Octa monomer, the microcapsule was more effective in alleviating the symptoms of weight gain, hypertension, and hyperlipidemia induced by HFD in mice. In conclusion, the construction of microcapsule structure can improve the dispersibility and stability of Octa in water, achieve sustained release of Octa in the gastrointestinal tract, and improve its efficiency in alleviating the effects of HFD on the body.

The relationship between the chemical composition and the quality of the soybean film during peeling process.

Soybean film is a traditional nonfermented soy product in China. It is a film formed on the surface of the soymilk during heating process. The nutrient components of soymilk will affect the quality of the soybean film. The results of this study showed that during the peeling process, the proportion of protein and carbohydrate in soymilk decreased, and the proportion of lipid increased. The mechanical properties (fracture extension and tensile strength) of the soybean film decreased during the peeling process. During the heating treatment, the Maillard reaction occurred and its intermediate products accumulated, which caused the change in soybean film color. White globular protein granules (<100 nm) existed on the surface of the soybean film. The lipid that was not wrapped by the protein network structure was exposed, and the evaporation of water led to the formation of black and gray holes on the skin (<500 nm). In addition, the results of correlation analysis showed that the changes in color, taste, and odor, as well as the mechanical properties of the skin, were all related to the changes in nutrients in the soybean film during peeling. This research provided a deeper understanding of the quality change in the soybean milk and the soybean film during the heating process.

Dityrosine suppresses the cytoprotective action of thyroid hormone T3 via inhibiting thyroid hormone receptor-mediated transcriptional activation.

Dityrosine (Dityr) is the most common oxidized form of tyrosine. In the previous studies of mice treated with dityrosine, cell death in the pancreas, kidneys, and liver was detected in the presence of enhanced plasma triiodothyronine (T3) content. Due to its structural similarity with the thyroid hormone T3, we hypothesized that dityrosine might disrupt T3-dependent endocrine signaling. The cytotoxic effect of dityrosine was studied in C57BL/6 mice by gavage with a dityrosine dose of 320 µg per kg per day for 10 weeks. Cell death in the liver was detected in the presence of enhanced plasma thyroid hormone content in mice treated with dityrosine. The antagonistic effect of dityrosine on T3 biofunction was studied using HepG2 cells. Dityrosine incubation reduced T3 transport ability and attenuated the T3-mediated cell survival via regulation of the PI3k/Akt/MAPK pathway. Furthermore, dityrosine inhibited T3 binding to thyroid hormone receptors (TRs) and suppressed the TR-mediated transcription. Dityrosine also downregulated the expressions of T3 action-related factors. Taken together, this study demonstrates that dityrosine inhibits T3-dependent cytoprotection by competitive inhibition, resulting in downstream gene suppression. Our findings offer insights into how dityrosine acts as an antagonist of T3. These findings shed new light on cellular processes underlying the energy metabolism disorder caused by dietary oxidized protein, thus contributing to a better understanding of the diet-health axis at a cellular level.

Effect of the heating process on the physicochemical characteristics and nutritional properties of whole cotyledon soymilk and tofu.

This study focused on the effect of the heating process on the whole cotyledon soymilk and tofu. Whole cotyledon soymilk was made from soybean cotyledon and processed by enzymatic hydrolysis using cellulase and high-pressure homogeneity. In this study, a one-step heating method was selected for the cooking process of whole cotyledon soybean milk, and the whole cotyledon soybean milk was heated to 90 °C and held for 4 min. Results showed that the protein, total saccharides and dietary fiber content of the whole cotyledon soymilk were higher than those of the tradition soymilk due to the existence of bean dregs (okara). Both protein aggregation and protein-polysaccharide interaction were observed during the heating process. We also found a change in soymilk physicochemical characteristics such as particle size distribution, viscosity, surface hydrophobicity and soluble protein during the heating process. The results in this study showed that compared with traditional tofu, the phytic acid and trypsin inhibitor content in whole cotyledon tofu was lower, so its protein had higher digestibility in vitro. In conclusion, whole cotyledon tofu had better health properties and application prospects.

Dityrosine administration induces dysfunction of insulin secretion accompanied by diminished thyroid hormones T3 function in pancreas of mice.

Oxidized tyrosine products are commonly found in food with high protein content and have been demonstrated to cause damage of liver and kidney in our previous studies. Dityrosine (Dityr) is a typical oxidized tyrosine product. Due to its structural homology with thyroid hormones T3, we assumed that one of the endocrine systems most likely considered in connection with its disruption by Dityr may be the T3 action. T3 plays important roles in insulin synthesis, and thyroid hormone resistance (RTH) is associated with the impairment of glucose metabolism. Therefore, this study determined whether Dityr exposure impaired T3 function in pancreas leading to glucose metabolism disruption. After 10-week gavage with Dityr, mice exhibited impaired glucose tolerance and disturbed energy metabolism. The elevated free THs content in plasma, the up-regulation of THs synthesis-specific genes expressions in thyroid glands, and the increased thyroid follicles histology shapes and areas indicated that Dityr enhanced the THs synthesis in thyroid glands. In addition, Dityr-induced RTH, which reflected as elevated plasma free THs in the presence of unsuppressed thyroid stimulating hormone. The mRNA downregulation of membrane transporter of T3 (MCT8) and co-activator factors (RXRα, Src-1), together with the decreased protein level of thyroid hormone receptor ß1 (TRß1) in pancreas illustrated that the activation ability of T3 to downstream gene involved in insulin synthesis was suppressed by Dityr. In MIN-6 cell experiment, T3 improved glucose-stimulated insulin secretion by upregulating mRNA levels of insulin synthesis-related genes (Ins2, MafA, Pdx1) and T3 action-related genes, as well as increasing protein level of TRß1. These data suggest that Dityr suppress T3-regulated insulin synthesis stimulated by glucose via an indirect way of decreasing sensibility to T3 in pancreas. All these findings indicate that Dityr can disrupt THs function in pancreas leading to glucose metabolism disorder.

Dietary oxidized tyrosine (O-Tyr) stimulates TGF-ß1-induced extracellular matrix production via the JNK/p38 signaling pathway in rat kidneys.

Oxidized tyrosine (O-Tyr) products have been detected in commercial food and have been demonstrated to induce liver injury in our previous study, but the precise mechanisms of the impact induced by dietary O-Tyr are still unclear. Kidney plays an important role in the metabolism of protein. Accumulation of O-Tyr products, especially the dityrosine (Dityr) and advanced oxidation protein products (AOPPs), in vivo was shown to be associated with many kidney diseases. Therefore, this study determined whether chronic exposure to dietary O-Tyr impaired renal function in rats. After O-Tyr treatment for 24 weeks, rats exhibited oxidative stress and protein oxidation in the kidneys, accompanied with inflammatory reaction and renal dysfunction. Elevated extracellular matrix (ECM) contents and the histological examination (HE and Masson stain) results indicated renal fibrosis. The Real-time PCR and Western blotting assay showed that O-Tyr activated phosphorylation of JNK/p38 and up-regulated the expression of transforming growth factor-ß1 (TGF-ß1) and Smad 2/3. These results suggest that dietary O-Tyr could induce oxidative stress, inflammation and renal fibrosis through JNK/p38/TGF-ß1 signaling pathway. Dityr (accounting for 22 % of the total O-Tyr material) may be responsible for the O-Tyr-induced injury. This study also provides a modified procedure for separation and purification of Dityr, the main oxidized product in O-Tyr.

Dityrosine administration induces novel object recognition deficits in young adulthood mice.

Dietary modifications have been shown to contribute to the physical and mental diseases. Oxidative modifications of protein can be easily found in protein-rich food such as meat and milk products. Previous studies mainly focus on the consequences of lipid oxidation products intake in vivo, but the effects of protein oxidation products consumption have been largely neglected. Oxidants have been shown to play an important role in aging and neurodegenerative diseases. Dityrosine is the oxidated product of tyrosine residues in protein which is considered as a biomarker for oxidative stress, but the potential deleterious effects of dityrosine are unknown. In the present study, we explored the effects of dityrosine administration on the behavioral aspect. We found that dityrosine-ingested mice displayed impaired memory during novel object recognition test, but no influence to the spatial memory in Morris water maze compared with the saline group. Other aspects of neurobehavioral function such as locomotor activity, anxiety and social behavior were not affected by dityrosine ingestion. Furthermore, we found that dityrosine-ingested mice showed decreased expression level of NMDA receptor subunits Nr1, Nr2a, Nr2b as well as Bdnf, Trkb. Our study suggests that dityrosine exposure impairs hippocampus-dependent nonspatial memory accompanied by modulation of NMDA receptor subunits and Bdnf expression.

24-Week Exposure to Oxidized Tyrosine Induces Hepatic Fibrosis Involving Activation of the MAPK/TGF-ß1 Signaling Pathway in Sprague-Dawley Rats Model.

SCOPE: Oxidized tyrosine (O-Tyr) has been widely detected in many consumer protein products. O-Tyr products such as dityrosine (Dityr) and 3-nitrotyrosine (3-NT) are universal biomarkers of protein oxidation and have been demonstrated to be associated with metabolic disorders in biological system. Evaluation of potential intracorporal effects of dietary O-Tyr is important since the mechanism of biological impacts induced by oral oxidized protein products (OPPs) is still limited although we have proved that some dietary OPPs would induce oxidative injury to liver and kidney. METHODS AND RESULTS: The present study aimed to investigate the dose-dependent hepatic injury caused by oral O-Tyr in rats. 24-week feeding of O-Tyr enhanced aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities, increased total bilirubin (TBiL) content, and led to oxidative damage in rats liver. Besides, O-Tyr distinctly increased the phosphorylation of p38 and ERK2 MAPKs and enhanced fibrosis-related TGF-ß1 and Smad2/3 levels. Higher extracellular matrix (ECM) indexes (ICTP, PIIINP) and histological examination (HE and Masson staining) also supported dose-dependent hepatic fibrosis caused by O-Tyr. CONCLUSION: These findings reveal that O-Tyr may induce oxidative damage and hepatic fibrosis via MAPK/TGF-ß1 signaling pathway, in which ROS together with malondialdehyde (MDA) and OPPs act as the pivotal mediators.

Alterations of the gut microbiota in high-fat diet mice is strongly linked to oxidative stress.

Alterations of the gut microbiota induced by diet exert a strong influence on the development of metabolic syndrome. In this study, we prove the hypothesis that the long-term high-fat diet (HFD) may influence gut microbiota directly and/or indirectly by changing the redox state. Lipoic acid (LA), as a universal antioxidant, was used to improve the redox state. Reactive oxygen species (ROS), total antioxidant capacity (T-AOC), and malondialdehyde (MDA) were analyzed to profile oxidative stress states. PCR-denaturing gradient gel electrophoresis (DGGE) was used to describe gut flora structures, while plate count was employed for the quantitative analysis of Escherichia coli, lactobacilli, and enterococcus. The influence of redox state on the vitality of gut-derived bacteria was measured in vitro. ROS and MDA, which significantly decreased in LA mice compared with HFD mice, showed a strong positive association with E. coli and enterococcus (P < 0.05) and a negative association with lactobacilli (P < 0.05). Increased T-AOC in LA mice showed a high positive association with lactobacilli (P < 0.05) and a negative correlation with E. coli and enterococcus. These correlations implied that the dietary effects on the gut microbiota were conferred, at least in part, through an effect on oxidative stress. This study provides evidence that modulation of the redox state by an antioxidant has the potential to improve gut microbiota, which has relevance for metabolic health.