Studies On Caco-2Cell Protective Effects Of Myricetin After Acrylamide Induced

Acrylamide is an unsaturated amide, studies show that starchy foods at high temperatures (>120℃) under cooking prone to generate acrylamide, such as french fries, potato chips etc. The content in such food exceeded the maximum limit of allowed over500times in drinking water. Acrylamide has acute toxicity, neurotoxicity and reproductive toxicity, genotoxicity and carcinogenicity. IARC identified acrylamide as category2A carcinogen. Acrylamide in our food processing is very easily generated, so understanding acrylamide, to explore the mechanism of toxicity of acrylamide and the prevention of the natural substances is particularly important.In this paper, we use vitro antioxidant experiments and cell models to study myricetin antioxidant activity (oxygen radical scavenging capacity) and studies on Caco-2cell protective effects of myricetin after acrylamide induced and delve into its protective mechanism.Acrylamide which produced in food processing is a huge threat to human health. Functional factors, extracted from natural plants safe, has become more and more hot research to explore the protective effect of acrylamide-induced cell damage in recent years on functional food and natural medicine field. Myricetin has anti-inflammatory, analgesic, anti-cancer, lowering blood glucose, protecting liver and other effects, but about its acrylamide-induced intestinal damage cell protective effect has not been reported. In this paper, using acrylamide as harmful substances, combined with in vitro antioxidant experiments and Caco-2cell model, intended to in-vitro antioxidant capacity, cell viability, intracellular antioxidant, enzyme activity and gene expression, with the use of molecular probe needles, fluorescent labeling technology to explore myricetin on acrylamide Caco-2cell damage protective effect and its mechanism of action. The main results are as follows:(1) In vitro antioxidant activity of myricetinUsing oxygen radical scavenging ABTS method, DPPH assay method and the hydroxyl radical method, myricetin measured in vitro antioxidant activity. The results showed that:myricetin has strong antioxidant capacity in vitro, and showed a dose-dependent, if set free radical scavenging rate at50%myricetin concentration by IC50, IC50for the ABTS is10.41μg/mL; DPPH IC50is5.66μg/mL; hydroxyl radical IC50is30.18μg/mL.(2) Caco-2Cell Protective Effects of Myricetin after Acrylamide inducedUsed by Caco-2cell model, the measurement of cell viability was observed that low concentrations of myricetin components are not toxic to cell, acrylamide can cause liver oxidative damage to Caco-2cells (cell viability46.91%(5mM), and40.54%(10mM)). And myricetin pretreated cells in the experimental group, the ratio of acrylamide added separately in the experimental group significantly increased cell viability, increased by33.42%and31.68%, so myricetin could significantly inhibit acrylamide-induced cytotoxicity. Use modern molecular biology techniques such as dynamic fluorescence visualization technology, molecular probes to study the mechanism of cell protective effect. The results showed that myricetin can effectively suppress the generation of acrylamide stimulate intracellular reactive oxygen such as superoxide anion levels, reduce mitochondrial membrane lipid peroxidation, restore mitochondrial membrane potential and cellular glutathione levels, to avoid damage to the nucleus, so as to achieve the purpose of protective cells from oxidative damage.(3) Study of mechanism of myricetin protective acrylamide-induced oxidative damage in cellsBy cell enzyme method, detect cell antioxidant enzymes SOD and CAT activities; using qPCR technology, test antioxidant genes Nrf2, HO-1, NQO1, GCS, GSTP, GPX, SOD and CAT expression at the transcriptional level.The results showed that:myricetin sample (5μg/mL and10μg/mL) can suppress and restore acrylamide caused by SOD, CAT activity decreased, myricetin can increase intracellular free radical scavenging activity of antioxidant enzymes, so that the antioxidant capacity enhancement and thus protection of acrylamide induced cell damage. At the same role under AA cells produce oxidative stress, increased NRF2, HO-1, NQO1mRNA expression; while under the effect of myricetin sample, cell will increase the expression of antioxidant genes, and enhance the antioxidant capacity of cells. This study showed that myricetin on Caco-2cells strongly induced two-phase detoxification enzymes HO-1, NQO1and y-GCS, and by restoring the antioxidant enzymes SOD, CAT enzyme activity to achieve protective effect on cellular oxidative damage induced by acrylamide.In summary, the results of this study provide that how to prevent acrylamide toxicity of a new way of thinking and methods, as well as a scientific basis for the development of natural, safe and effective myricetin functional products.