The Mechanism Of The Interaction Of Anthocyanins(Aronia Melanocarpa Elliot)binding In Protein

The fruit of Aornia mealnocarpa Elliot is rich in anthocyanins and its content is up to 1%.The fruit extracts of Ralstonia variegata has strong anti-oxidation and free radical(ROS)scavenging capacity,it has certain preventive and therapeutic effects in improving immunity,anti-cancer,anti-aging,and anti-cardiovascular diseases.Our group's previous experiments optimized the extraction and purification of anthocyanin in Aornia mealnocarpa Elliot(AMA),and identified its components and biological activities,it was found that anthocyanin can inhibit the occurrence of diabetes and obesity,and it also can regulate the metabolic balance of the body and keep the redox system stablely.However,because anthocyanins are susceptible to the surrounding environment,such as temperature,p H,light intensity,and metal ions,which greatly reduce its biological activity and its nutritional value.Anthocyanins can exist in a stable form of astragalus salt cations under acidic conditions,While with the increase of p H gradually,the anthocyanins form become unstable chalcone.Therefore,improving the stability of anthocyanins is a current technical problem that needs to be solved.Based on the important influence of the low stability of anthocyanins on daily use,this study intends to use AMA as the research object and ?-casein as the conjugate,spectroscopic techniques are used to analyze the combination mode,the binding distance and the structural changes;The stability changes in different environments after their binding were studied to determine the degradation mode and kinetic parameters.This study will explain the influence mechanism of AMA and ?-casein on the stability of anthocyanin after binding,and provide a scientific theoretical basis for the rational and effective development of the anthocyanins resources and its application in the food industry.Methods and results:1)Extraction of AMAUsing the principle of similar dissolution,AMA crude extraction was extracted by vacuum distillation and purified by AB-8 macroporous resin.The extraction rate under different conditions was analyzed.Finally,under the static purification conditions,when the adsorption temperature is 20 ?,AMA crude extract 8 g/L,desorption temperature is 50 ?,using 60 % ethanol with p H 3.0 as the desorption liquid has the highest extraction efficiency.Under dynamic adsorption conditions,8 g/L AMA crude extract was passed twice at a flow rate of 0.6 m L/min,and at p H 3.0 using 60 % ethanol at a resolution of 0.6 m L/min.The main components of AMA were obtained by HPLC were cyaniding-arabinosin,cyaniding-galactoside,cyaniding-glucoside and cyaniding-xyloside,cyaniding-arabinosin and cyaniding-galactoside in them were more abundant.2)The interaction of AMA and proteinsAccording to the previous study on the separation and purification of AMA components,the fluorescence quenching technique was used to analyze the quenching type of the two components.It was found that the two species reacted and the quenching type was static quenching.At the same time,data analysis showed that the binding site between the two was 1,there was a binding site.By calculating the thermodynamic parameters,the binding forces between the two was mainly hydrophobic forces.By infrared spectroscopy and circular dichroism analysis,the secondary structure of the protein after the combination of the two,mainly the increase in the content of ?-sheet,random coiling and reduction in the amount of rotation,indicating that the two formed a more stable after binding.The structure makes it more stable.Finally,using molecular modeling techniques to analyze the binding mode and binding sites,to further confirm the above conclusions.3)Stability changes after binding of AMA to proteinsAfter confirming the stable combination of the two,the AMA residual rate was measured under conditions of high temperature,high p H,and continuous light.With increasing p H and temperature and the addition of H2O2 oxidant,the residual rate was 80%,demonstrating that the conjugate can be maintained higher stability.However,with the addition of continuous light and reducing agent,the residual rate was 82%,indicating that the stability of the conjugate was not improved significantly.This may be due to the lack of light intensity and the protective effect of the reducing agent on anthocyanins.By calculating the stability response of this segment,the first-order kinetic equation is satisfied.