The Inhibition Of Aronia Melanocarpa Anthocyanin Extracts On Macrophages Inflammation Induced By Lipopolysaccharide

Black chokeberry（Aronia melanocarpa）is a distinctive berry with a high content of polyphenol compounds.There is ample evidence for chokeberry health benefits including antidiabetic,cardioprotective,hepatoprotective,anticarcinogenic,and anti-inflammatory activities.In this experiment,ultrasonic-assisted extracts of anthocyanin from the Aronia melanocarpa was optimized,and the antioxidant capacity of the anthocyanin extracts was determined by comparison with vitamin C.the composition of the anthocyanin extracts from the Aronia melanocarpa were analyzed by high performance liquid chromatography--tandem mass spectrometry（HPLC-MS/MS）,and Aronia melanocarpa anthocyanin extracts Aronia melanocarpa performs the possible inhibition mechanisms on LPS-induced macrophages RAW264.7 inflammation from the oxidative stress response,expression of inflammatory mediators and apotosis.The main results are as follows:Using ultrasonic-assisted extraction to select single factors that have more significant effects,the extraction conditions of anthocyanins were optimized by response surface methodology.The results showed that the optimum extraction conditions were as follows:extraction temperature,60℃,ultrasonic power,100 W,solid-to-liquid,1:30（g/mL）,ultrasonic time,31 min,ethanol concentration,64%and pH,2.0.The analysis and quantification of the anthocyanins and phenolic acids in the Aronia melanocarpa were carried out by high performance liquid chromatography-tandem mass spectrometry and high performance liquid chromatography.The corresponding standards were used as references.There are 6 kinds of anthocyanins and 9 kinds of phenolic acids in the anthocyanin extract.Theeffectsof1,1-Diphenyl-2-picrylhydrazylradical2,2-Diphenyl-1-（2,4,6-trinitrophenyl）hydrazyl radical scavenging capacity（DPPH）,2,2’-Azinobis-（3-ethylbenzthiazoline-6-sulphonate）radical scavenging capacity（ABTS+·）and Fe³⁺reducing ability of plasma（FRAP）were used to evaluate their bioactivity,he vitro antioxidant experiments showed that the antioxidant activity of anthocyanins was significantly higher than that of VC under the same mass concentration,about 2-4 times.The present study investigated the anti-inflammatory effect of anthocyanin-rich phenolic extracts of Aronia melanocarpa（Ame）in lipopolysaccharide（LPS）-stimulated RAW 264.7murine macrophage cells by measuring the main oxidative,inflammatory and apoptosis biomarkers.The results showed that Ame pre-treatment at 50μg/mL significanly ameliorate oxidative stress and inflammatory mediators activities mediated inflammatory responses,as evidenced by the reduction in the production of reactive oxygen species and malondialdehyde.In particular,ROS was decreased by 36%compared with LPS exposure group.as well as the suppression of levels of nitric oxide,inducible nitric oxide synthase,cyclooxygenase-2 and prostaglandin E2 mRNA.The most significant of these is a 60%reduction in NO production;remarkably elevated LPS-induced anti-inflammatory cytokine interleukins-10 and reduced pro-inflammatory cytokines IL-1β,IL-6 and tumour necrosis factor-α,which are well-known key factors of inflammation.Comparing with cells treated with LPS,mRNA expression of IL-1β,IL-6,and TNF-αin Ame/LPS was decreased by approximately 66%,16%,and 21%,respectively.;additionally,observably attenuated the LPS-induced cells apoptosis,which were verified by the cell cycle distribution and apoptosis levels measured by flow cytometry and the mRNA expression of apotosis factors such as caspase-3 and caspase-9 are lower than that of control groups.Aronia melanocarpa extracts may be developed as a potential novel ingredient in anti-inflammatory health products or a candidate drug for the prevention of inflammation.