| The leaves of Toxicodendron vernicifera Stokes. which belong to Toxicodendron in Anacardiaceae can be used as medcine and was seen in the Cuba "Qiyeqingzhansan" first.The book"Compendium of Materia Medica" recorded that the leaves of Toxicodendron vernicifera Stokes. was a spicy,warm and small poison medcine that maily used in the erea of dead diseases and insecticidal. The one was a little using medcine in traditional Chinese medcine,but it has good activity and often used in the manufacturing erea of laquer tree and the Folk.Modern scientific research shows that the effect of the leaves is very significant. Recently,there are little study about the leaves of Toxicodendron vernicifera Stokes. and only few literature about Botany and Pharmacognosy, but maily concentrated on planting of the lacquer tree, producing of raw lacquer and the applications and laccase and urushiol.In this study, by selecting the leaves of Toxicodendron vernicifera Stokes. as the object and from the point of researching for pharmaceutical chemistry,we constuct the evaluating method for high performance liquid chromatography (HPLC) firstly, combine with Liquid Chromatography-Mass Spectrometry(HPLC-MS) technology to analysis the urushiol qualitatively,then determine the content of main urushiol in different breeds in the leaves of lacquer tree. Based on these, we establish the HPLC fingerprints of fresh and dried leaves in different breeds of the lacquer tree and by the comparison of them to analysis the changes of main chemical compounds between fresh with dried leaves and different breeds.At the same time, with discussing the mainly active ingredients,we should offer the scientific basis for further exploring and passing on our medicine of ancient Cuba,developing and utilization of this medical resources of the leaves, conducting the quality control and comprehensive evaluation system for it.1 Using physical and chemical identification and thin layer chromatography (TLC) to predict the composition of leaves in Toxicodendron vernicifera Stokes. and define that mainly contain urushiol compounds at first.By comparison different solvent extraction, we found that Chloroform can be a good solvent for extracting urushiol.The result of TLC indicated that the agent of Petroleum ether-ethyl acetate and coloring of FeCl3 can get much more spots and better seperation. Furthermore, this text showed that the result of physical and chemical identification and TLC between fresh and dried leaves were also the same and urushiols are the main components.2 This research combined with other stduies both home and broad to conduct a new method for HPLC analysis of urushiol compounds in the leaves of the lacquer tree by screening and optimization the chromatographic conditions as extraction mobile phase selection, detection wavelength,flow rate and column temperature. The Optimum chromatographic conditions are as follows:using a elution with a acetonitrile-water(91:9) at 380nm,0.6mL·min-1,35℃and this method is very Simple, rapid, sensitive and repro-ducible.Then we rebuild the analysis way for the acetonitrile extraction of urushiol qualitatively by Liquid Chromatography-Mass Spectrometry technology. The rusult show that We have separated 10 peaks by LC, and identified 9 peaks were the saturated, single, diene, and triene of C15 or C17 different urushiol compounds according to the MS and existing literature.3 As saturated urushiol for reference substance, we determined each mark with HPLC quantitatively. The results show that there are diversities in different varieties of content of the total urushiol, four categories of substance urushiol and 9 urushiol compounds. The total urushiol has the higher content in the leaves of Gaobachi Pingli and Dahongpao Pingli, but lower in Qieketou Shangnan and Huoyanzi Shangnan. For the content of four categories of substance urushiol, triene urushiol is highest, saturated and diene urushiol are the second, the single one is lowest.In different varieties,the highest content of four categories of substance urushiol is from the leaves of Dahongpao Pingli, then is Dahongpao and Qieketou, Huoyanzi is the lowest.4 We have conducted the HPLC fingerprints for fresh leaves of five lacquer trees which come from different two areas. By examining the coefficient of variation for the peaks of retention time and peak area value,the results showed that the basical chemical constituents of different varieties are very similar, but the contents are quiet different, with the similarity is between 0.963-0.994 and the average similarity is 0.985, also we see that the established HPLC fingerprint analysis method is reproducible, practical and simple. Compared the HPLC fingerprints of dried leaves with the fresh one, there has a similar in general marks, but less consistent,with the similarity is between 0.928-0.993 and the average similarity is 0.970.The HPLC fingerprints of fresh and dried leaves showed that the area value of general marks account for 90% in that total peaks,and the 9 marks which are urushiol compounds in the 10 general marks can be as characteristic fingerprint peaks.5 By comparied the components and contents of urushiol and the HPLC fingerprints between the fresh and dried leaves of lacquer tree,the results consistently reviewed that the components of urushiol are almostly consistent,but the contents of the total urushiol and four categories of substance urushiol in the fresh leaves are all obviously higher than the dried one and the extent of variation due to different species. |
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