Pilot Study On The Mechanism Of Main Active Components In Berberis Heteropoda Schrenk Against A?-induced Alzheimer's Disease

Objective:To optimize the ultrasonic extraction of anthocyanins from Berberis heteropoda Schrenk,and determine the main active components in Berberis heteropoda Schrenk;To study the mechanism of the major active components of Berberis heteropoda Schrenk on the protective effect of AP25-35 induced PC12 cells.Methods:1 pH differential method for determination of anthocyanins content as evaluation index,using single factor rotation method and the orthogonal experiment method to ascertain the extraction time,extraction times,extraction solvent and other parameters;Using thin layer chromatography and high performance liquid chromatography to analyze the main active components of anthocyanins from Berberis heteropoda Schrenk.2 The cytotoxicity of different concentrations of cyanidin-3-O-glucoside to PC 12 cells,the concentration of AP25-35 as a model agent,and the effects of different concentrations of cyanidin-3-O-glucoside to protect A?25-35 injured PC 12 cells was detected by MTT assay;After the drug effect the release of LDH,SOD,MDA,ROS,GSH-PX of PC12 cells was detected by ELISA assay;Using Laser scanning confocal microscope to detecte apoptosis of PC 12 cells;After the drug effect the expression of Bax,Bcl-2,Akt,p-Akt,Caspase-3,Cleaved Caspase-3,Caspase-9 and Cleaved Caspase-9 was detected by WB.Results:1 Results showed that the extraction conditions were determined as follows:ethanol concentration 70%,hydrochloric acid concentration 0.8%,solid-to-liquid ratio 1:25(g/mL),extraction time 20 min,extraction times 4 times,the anthocyanin content in extracts is higher than the anthocyanin content in extracts;The main active components in Berberis heteropoda Schrenk is cyanidin-3-O-glucoside.2 Cyanidin-3-O-glucoside is not cytotoxic at concentrations of 0-500 ?g/mL,the concentration of A?25-35 as a model agent was 40 ?mol/L,and the different concentrations of cyanidin-3-O-glucoside to protect A?2S-35 injured PC 12 cells,the low dose group was 3.90625 ?g/mL,the middle dose group was 15.625 ?g/mL and the high dose group was 62.5 ?g/mL;After the drug effect cyanidin-3-O-glucoside can inhibit the release of LDH,MDA and ROS,and promote the release of SOD and GSH-PX of PC12 cells;After the drug effect cyanidin-3-O-glucoside can inhibit the apoptosis of PC 12 cells;After the drug effect cyanidin-3-O-glucoside can upregulate the expression of Bcl-2?Akt?p-Akt protein and down-regulate the expression of Bax?Caspase-3?Cleaved Caspase-3?Gaspase-9?Cleaved Caspase-9 protein in PC12cells.Conclusion:1 Optimized extraction method for the extraction of anthocyanins from Berberis heteropoda Schrenk is stable and feasible;Cyanidin-3-O-glucoside is the main active ingredient.2 Cyanidin-3-O-glucoside can protect A325-35 injured PC 12 cells,its protective effect may be through the inhibition of cell oxidation,or the activation of PI3K/Akt pathway inhibiting apoptosis,Cyanidin-3-O-glucoside a potential candidate for AD.