Interaction mechanism between ZnO nanoparticles-whey protein and its effect on toxicity in GES-1 cells.

The interaction between zinc oxide nanoparticles (ZnO NPs) and whey protein (WP) was studied. The gastric epithelial cell line (GES-1) was used to evaluate the toxicity intensity of ZnO NPs. The interaction mechanism of ZnO NPs and WP was studied by spectroscopic techniques. The results showed that the inhibitory effect of ZnO NPs on cells activity could be reduced when added to ZnO NPs at a concentration of 50 µg/ml. The fluorescence quenching mechanism of ZnO NPs on WP is a combination of dynamic and static quenching. The interaction force between ZnO NPs and WP can be considered as H-bond and VdW force, and they have two binding sites. The interaction between WP and ZnO NPs leads to the loosening of the structural skeleton of WP and the extension of peptide chain, which exposes the tyrosine (Tyr) and tryptophan (Trp) hydrophobic groups in the hydrophobic region of protein molecules and reduces the hydrophobicity of the microenvironment. The ZnO NPs might form a complex with WP through H-bond, hydrophobic interactions, and so on, leading to peptide chain rearrangement, and finally causing changes in the secondary structure of α-helix. Practical Application This study provides a theoretical basis for future research on the interaction between food ingredients and nanomaterials, the evaluation of toxicity of nanomaterials and the application scope of nanomaterials in food field.

The hydrogel of whey protein isolate coated by lotus root amylopectin enhance the stability and bioavailability of quercetin.

In this study, whey protein isolate (WPI)-quercetin (Que)-lotus root amylopectin (LRA) hydrogels (WPI-QUE-LRA) was developed to improve the solubility, stability and bioavailability of quercetin. Results showed that the favorable WPI-QUE-LRA was formed using WPI and LRA in the ratio of 1:2 at pH 7.0. Under this condition, the average size, polydispersity index, zeta potential of the WPI-QUE-LRA was 179.5 nm, 0.271, -18.6 mV, respectively. The analysis of transmission electron microscopy, fourier transform infrared spectroscopy and X-ray diffractometer revealed that the quercetin was successfully encapsulated in WPI-LRA, giving a high encapsulation efficiency of 92.4 %. Moreover, the WPI-LRA could significantly improve the storage stability and photochemical stability of quercetin. The in vitro and in vivo experiments showed that LRA-coated WPI hydrogel can enable quercetin to be stable in stomach and be effectively released in small intestine, leading to the enhancement of the bioavailability of quercetin.

Toxicity study and blood pressure-lowering efficacy of whey protein concentrate hydrolysate in rat models, plus peptide characterization.

We evaluated the acute (single-dose) and subacute (repeated-dose) oral toxicity of alcalase-hydrolyzed whey protein concentrate. Our acute study revealed no death or treatment-related complications, and the median lethal dose of whey protein concentrate hydrolysate was >2,500 mg/kg. In the subacute study, when the hydrolysate was fed at 3 different concentrations (200, 400, and 800 mg/kg), no groups showed toxicity changes compared with controls. Then, whey protein concentrate hydrolysate was orally administered to spontaneously hypertensive rats. Results revealed significant reductions in blood pressure in a dose-dependent manner, and dosing at 400 mg/kg led to significant blood pressure reduction (-47.8 mm Hg) compared with controls (blood pressure maintained) and the findings of previous work (-21 mm Hg). Eight peptides-RHPEYAVSVLLR, GGAPPAGRL, GPPLPRL, ELKPTPEGDL, VLSELPEP, DAQSAPLRVY, RDMPIQAF, and LEQVLPRD-were sequentially identified and characterized. Of the peptides, VLSELPEP and LEQVLPRD showed the most prominent in vitro angiotensin-I converting enzyme inhibition with half-maximal inhibitory concentrations of 0.049 and 0.043 mM, respectively. These findings establish strong evidence for the in vitro and in vivo potential of whey protein concentrate hydrolysate to act as a safe, natural functional food ingredient that exerts antihypertensive activity.

Effects of whey protein and conjugated linoleic acid on acrolein-induced cardiac oxidative stress, mitochondrial dysfunction and dyslipidemia in rats.

Acrolein is a ubiquitous environmental pollutant. Whey protein and conjugated linoleic acid are widely used weight-loss supplements. We aimed to evaluate blood lipid profiles, oxidative stress and mitochondrial bioenergetics function in hearts of rats treated with acrolein and/or the weight-loss supplements. The animals were orally gavaged with acrolein, whey protein, conjugated linoleic acid, acrolein + whey protein or acrolein + conjugated linoleic acid for six days per week during 30 days. Acrolein caused dyslipidemia and oxidative stress in red blood cells and haert mitochondria. Moreover, it caused dysfunction in mitochondrial bioenergetics by decreasing levels of oxidative phosphorylation enzymes, tricarboxylic acid cycle enzymes and ATP. Co-treatment with acrolein + whey protein and acrolein + conjugated linoleic acid ameliorated acrolein-induced oxidative stress and dysfunction in mitochondrial bioenergetics. This amelioration effect was more prominent in acrolein + conjugated linoleic acid group. Interestingly, co-treatment with acrolein + whey protein negatively affected some markers of cardiac injury such as creatinine kinase-MB, lactate dehydrogenase and homocysteine. Conjugated linoleic acid may also cause dyslipidemia because it increased the levels of triacylglycerol, low density lipoproteins and very low density lipoproteins. In conclusion, using some weight loss supplements such as whey protein may adversely affect the biochemical parameters related to cardiovascular system.

Whey Protein Concentrate WPC-80 Intensifies Glycoconjugate Catabolism and Induces Oxidative Stress in the Liver of Rats.

The aim of this study was to evaluate the effect of whey protein concentrate (WPC-80) on glycoconjugate catabolism, selected markers of oxidative stress and liver inflammation. The experiment was conducted on male Wistar rats (n = 63). The animals from the study group were administered WPC-80 at a dose of 0.3 or 0.5 g/kg body weight for 7, 14 or 21 days, while rats from the control group received only 0.9% NaCl. In liver homogenates, we assayed the activity of N-acetyl-ß-D-hexosaminidase (HEX), ß-glucuronidase (GLU), ß-galactosidase (GAL), α-mannosidase (MAN), α-fucosidase (FUC), as well as the level of reduced glutathione (GSH), malondialdehyde (MDA), interleukin-1ß (IL-1ß) and transforming growth factor-ß1 (TGF-ß1). A significantly higher activity of HEX, GLU, MAN and FUC were found in the livers of rats receiving WPC-80 compared to controls. Serum ALT and AST were significantly higher in the animals supplemented with WPC-80 at a dose of 0.5 g/kg body weight for 21 days. In the same group of animals, enhanced level of GSH, MDA, IL-1ß and TGF-ß1 were also observed. WPC-80 is responsible for intensive remodelling of liver tissue and induction of oxidative stress especially at a dose of 0.5 g/kg body weight.

Oral sensitization to whey proteins induces age- and sex-dependent behavioral abnormality and neuroinflammatory responses in a mouse model of food allergy: a potential role of mast cells.

BACKGROUND: Growing evidence has strengthened the association of food allergy with neuropsychiatric symptoms such as depression, anxiety, and autism. However, underlying mechanisms by which peripheral allergic responses lead to behavioral dysfunction are yet to be determined. Allergen-activated mast cells may serve as mediators by releasing histamine and other inflammatory factors that could adversely affect brain function. We hypothesized that eliciting food allergy in experimental animals would result in behavioral changes accompanied by mast cell accumulation in the brain. Our hypothesis was tested in a mouse model of milk allergy using bovine milk whey proteins (WP) as the allergen. METHODS: Male and female C57BL/6 mice at 4 weeks (young) and 10 months (old) of age underwent 5-week WP sensitization with weekly intragastric administration of 20 mg WP and 10 µg cholera toxin as an adjuvant. Age-matched sham animals were given the vehicle containing only the adjuvant. All animals were orally challenged with 50 mg WP in week 6 and their intrinsic digging behavior was assessed the next day. Animals were sacrificed 3 days after the challenge, and WP-specific serum IgE, intestinal and brain mast cells, glial activation, and epigenetic DNA modification in the brain were examined. RESULTS: WP-sensitized males showed significantly less digging activity than the sham males in both age groups while no apparent difference was observed in females. Mast cells and their activities were evident in the intestines in an age- and sex-dependent manner. Brain mast cells were predominantly located in the region between the lateral midbrain and medial hippocampus, and their number increased in the WP-sensitized young, but not old, male brains. Noticeable differences in for 5-hydroxymethylcytosine immunoreactivity were observed in WP mice of both age groups in the amygdala, suggesting epigenetic regulation. Increased microglial Iba1 immunoreactivity and perivascular astrocytes hypertrophy were also observed in the WP-sensitized old male mice. CONCLUSIONS: Our results demonstrated that food allergy induced behavioral abnormality, increases in the number of mast cells, epigenetic DNA modification in the brain, microgliosis, and astrocyte hypertrophy in a sex- and age-dependent manner, providing a potential mechanism by which peripheral allergic responses evoke behavioral dysfunction.

Usage of whey protein may cause liver damage via inflammatory and apoptotic responses.

The purpose of this study was to investigate the long- and short-term inflammatory and apoptotic effects of whey protein on the livers of non-exercising rats. Thirty rats were divided into three groups namely (1) control group, (2) short-term whey (WS) protein diet (252 g/kg for 5 days), and (3) long-term whey (WL) protein diet (252 g/kg for 4 weeks). Interleukin 1ß (IL-1ß), IL-6, tumor necrosis factor α (TNF-α), and cytokeratin 18 (CK-18-M30) were assessed using enzyme-linked immunosorbent assay and immunohistochemical methods. Apoptosis was evaluated using the terminal transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) method. Hepatotoxicity was evaluated by quantitation of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT). Based on the biochemical levels and immunohistochemical results, the highest level of IL-1ß was identified in the WL group (p < 0.01). The IL-6 and TNF-α results were slightly lower in the WS group than in the control group and were highest in the WL group (p < 0.01). The CK-18-M30 and TUNEL results were highest in the WS group and exhibited medium intensity in the WL group (p < 0.01). AST results were statistically significant for all groups, while our ALT groups were particularly significant between the WL and control groups (p < 0.01). The results showed that when whey protein is used in an uninformed manner and without exercising, adverse effects on the liver may occur by increasing the apoptotic signal in the short term and increasing inflammatory markers and hepatotoxicity in the long term.