Studies Of Extraction Process Of Total Flavones From Chinese Olive And Its Protective Effects Against Cerebral Ischemia/reperfudion Injury In Rats

OBJECTIVE To determine of total flavone ( rutin as standard ) of the extraction is controlled by visible spectrophotometry and to establish the optimal condition for ultrasonic extraction of total flavone from Chinese olive and investigate appropriate macroporous resins ;The optimal technological parameters of the purification process of TFCO are found ; To study the protective effects and possible mechanisms of TFCO on focal cerebral ischemia-reperfusion injury (CIRI) in rats by reversible middle cerebral artery occlusion (MCAO) model.METHODS The ultrasonic extraction and orthogonal test were used to choose optimum extractive method which was as follows: optimum extractive solvent's concentration; solvent volume; extractive time .The purification process of total flavone with macroreticular resin was studied and the type of macroporous resin, review the absorption ratio; desorption ration; liquor's pH; ethanol's concentration, volume, velocity were established.MCAO model was produced by intraluminal suture technique, and reperfusion 24h was begun after 2h of occlusion. TFCO was intragastrically administrated at 1d,2d,3d and 4d of the reperfusion. After reperfusion 24h, the extent of neurological deficits was evaluated by Longer's method; the infarct size of the brain and the degree of cerebral edema were measured by Auto CAD image analysis soft ware; the morphological changes of the brain tissue were observed; the expressions of iNOS and TNF-αproteins were measured;T-AOC,MPO and GSH level in the brain tissue were detected.RESULTS (1) The optimum extraction procedures was as follows:4times of 95% ethanol ;extractive time :40 minutes .the best technics of purification is as follows: the type of macroporous resin was D101,the sample fluid(pH4.5),the velocity is 2BV/h,the water is 3BV with velocity of 2BV/h,60% ethanol is 3BV with velocity of 1BV/h.(2)After 2h of occlusion and 24h of reperfusion, the neurological deficits were evidently observed;the brain infarct size and edema degree were increased; the changes of the brain histology were appeared and the expressions of iNOS and TNF-αproteins were increased;the activity of MPO were increased, the activity of GSH was decreased, the contents of T-AOC were increased.(3)Both 30 and 60 mg.kg^-1 doses of TFCO can dramatically reduce neurological score, the infarct size and degree of cerebral edema comparing with ischemia reperfusion group. Both 30 and 60 mg.kg^-1 doses of TFCO can increase the activity of GSH,decrease the activity of MPO and the contents of T-AOC,inhibit the expression of iNOS protein and TNF-αprotein.CONCLUSION (1) The total flavones's purity could reach 90.1%.(2) TFCO 30,60 mg.kg^-1 can play a protective role in the brain injury. TFCO could protect the focal CIRI by depressing permeability of the blood brain barrier, inhibiting the expressions of iNOS and TNF-αproteins, depressing inflammation reaction, decreasing lipid peroxidation.