| Objective: ① to study the extraction and purification process of total balsamic acids from Benzoinum andcerebral protection effect of total balsamic acids.Methods:(① establishment of evaluation methods for total balsam acids gravi-metric method as a reference, establishing UV method to determinae total Balsam benzoin acid; investigating HPLC method to determinate benzoic acid in total balsam acid.② the extraction process of total balsam acids the transfer rates of total Balsam acid and benzoic acid as indices, to study the effects by solvent extraction with different solvents and extraction method. Based on these, the orthogonal test method was used to arrange experiments to test the effect of three main factors(the concentration of sodium hydroxide, the amount of solvent, extraction time) to determine extraction process parameters of total balsam acids.Based on the extraction proces, study the purification process by anion exchange resin.③ the purification process of total balsam acids adsorption and elution rate of total Balsam acid、benzoic acid as indices, visiting the kind of resin, pH、 concentration、volume、flow rate of sample and the amount、concentration、flow rate of adsorbent to determine purification technology parameters④ the pharmacodynamic study on cerebral protection selecte the modeles of acute cerebral ischemia and anoxia in mice, such as saturated magnesium chloride in mice induced by acute cerebral ischemia and anoxia in mice experiments to the survival time of mice and verified the effect of "adjuvant and messenger" of borneol by compatibilityResults:① establishment of evaluation methods for total balsam acids the detecter wavelength of UV method was set at 223 nm and the regression equation was A=66.750C+0.0377(r=0.9991); the regression equation of HPLC method was Y=5839.9X-259.06 (r=0.9994). Compared the content difference between UV and gravimetric method for determination of total balsamic.from Benzoinum, P=0.437 (>0.05), it suggests no substantial difference between two methods. UV method is fitting for the determination of total balsamic acids.② the extractionn process the optimal extraction process was refluxed with 15 times of 0.75mol/1NaOH, extracting 1h 1 time;③ the purificatio process the optimal technological parameters are as follows:the concentration of the sample solution is 0.02g/mlwith pH 12.0, the sample volume is 8BV, the flow rate is 2BV/h; impurities are removed by 3BV of deionized water,1BV of dilute HCl is used to desorpt,5BV of 95% ethanol is used to elute with the flow rate of 1BV/h. After purification, the average purity of total balsamic acids and benzonic acid are 73.49% and 63.97%, the purity time of total balsamic acids and benzonic acid are 2.78 and 2.95.④ The pharmacodynamic study on cerebral protection Compared to blank group, the effect of all the drug group, total balsamic acids group and Benzoinum group, were no significantly. But, to some extent, Benzoinum group and total balsamic acids group prolonged the survival time of mice. Total balsamic acids group compared to Benzoinum group, the compatibility group of total balsamic acids with borneol compared to the compatibility group of Benzoinum with borneol, there were no obvious difference; but total balsamic acids group and the compatibility group of total balsamic acids with borneol prolonged the survival time of mice to some degree.Conclusion:① UV method is sensitive and simple, and the determination result is accurate and reliable.It is fitting for the determination of total balsamic acids, for There is on significant difference between UV methods and gravimetric method for determination of total balsam acid. The steps of gravimetric method are too complexed to make mistake; the steps of UV method were reduced and turnround were shorten. UV method is accurate and reliable and can be used for determination of total Balsam benzoin acid.②The extraction process is rational、stable and feasible with high extraction; with the highly alkaline anion-exchange resin of 717, the efficiency of purification for total balsamic acids from Benzoinum is good, the cost and cycle is less than before. Spectrophotometry. These process laid the foundation for pharmacodynamic study on cerebral protection.③ Compared to Benzoinum group, the effect of total balsamic acids group was no no remarkable difference, but total balsamic acids delayed the death time of mice to a certain extent. It shows total balsamic acids may be a part of phamocodynamic substance of Benzoinum. Moreover, it futher verified the effect of "adjuvant and messenger’of borneol and laid the foundation for study on Benzoinum. |
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