| Objective:This paper is part of the National Natural Science Foundation project titled "the correlation of functional changes before and after processed and the changes of the material basis in Polygonum orientale L.(Project No:30873437)".Polygonum orientale is the dried ripe fruit of Polygonum orientale L. of Polygonum plant. It is cold and salty. It is in the liver and stomach channels; It has the functions of Swelling with casual blood, elimination pain of accumulation, indicate Cyst, Cancer Pain, Indigestion, stomach pain. It shows that it has anti-tumor, antibacterial and diuretic effect in modern pharmacological studies. This research mainly includes:the study of the chemical constituents in Polygonum orientale in butanol layer, the analysis of chemical composition in Polygonum orientale by LC/MS, the analysis of the related factors of the quality of Polygonum orientale, the study of the development of other medicinal parts of Polygonum orientale and the research of the formulations and technology of Polygonum orientale.The research of the chemical constituents in Polygonum orientale is further improved through the study of the chemical constituents in n-butanol layer.The new analysis method of chemical constituents in Polygonum orientale is established by the analysis of LC/MS method.The processing principle of Polygonum orientale is explored to further extent and the Chinese medicine theory is verified through the analysis of the chemical constituents changes in Polygonum orientale before and after processed by HPLC method. The determination method of the active ingredient paprazine is established and it supplies a new assessment reference of the quality control of vitro pharmaCod-ynamics by examining the affect of different storage time on Polygonum orientale.It offers the scientific reference for the development of the resources of Polygonum orientale throuth the chemical constituents comparison of different medical parts and the experimental research of vitro pharmacodynamics. The former working basis of the new drug development for Polygonum orientale is supplied through the examinating the formulation of the process of Polygonum orientaleMaterials and methods:Material: the herbs used in content determination are procured from herbal markets, and other herbs are collected in Dalian Nanguanling identified by Professor Yanjun Zhai in Liao Ning University of TCM as the dried ripe fruit of Polygonum orientale L. of Polygonaceae plant. Reference substances used in experiments are made in the laboratory.Method:1. The research of chemical components of n-butanol layer:the separation of chemical constituents in n-butanol layer of Polygonum orientale is completed by column chromatography, recrystallization, thin layer chromatography and liquid preparation chromatography methods and the compound structures are identified with the spectral analysis.2. The chemical composition contained in Polygonum orientale L. is determined by the analysis of comparison of the LC\MS spectral data and the data collected from literature, and the compound data isolated from Polygonum orientale L. is collected through checking plenty of related literature.3. The analysis of related factors to quality of Polygonum orientale:to analyze the chemical components changes before and after processed in Polygonum orientale, the storage rhythm and the content determination of paprazine.4. The research and development of other pharmaceutical parts in Polygonum orientale:the pharmacological analysis of anticancer activity of root, stem and leaf is done by MTT reduction, and the content determination of the target components is done by HPLC method.5. The formulation and technology research of Polygonum orientale:to filter the rational and effective drugs in traditional medicine theory from literature collected, and combine them into new anti-cancer compounds. The best extraction technology and formulations are investigated and established By orthogonal design.Results:2. The chemical components research of n-butanol layer in Polygonum orientale:8 compounds are isolated from n-butanol layer, respe:β-sitosterol, palmitic acid, 3,3-di-o-methylellagicacid, quercetin,daucosterol,p-hydroxyphenylethanol-p-coum-asric, taxifolin, paprazine.3. The chemical composition analysis of Polygonum orientale by LC/MS:8 compounds are determined by LC/MS and literature comparison. They are:taxifolin,paprazine,quercetin,palmitic acid,p-hydroxyphenylethanol-p-coumari,orientin,isoorientin。3,5,7-trihydrochromone,4. The analysis of related factors of quality of Polygonum orientale:the quercetin content in Polygonum orientale is significantly increased after processed, quercetin is one of the by-products from the heating experiments in the same processing conditions as taxifolin and the content is more than non-processed.5. It is indicated that by the study of the rhythm of storage time:the active component paprazine is slowly increased with the storage time increasing, and there are unknown components increasing slowly as well, while the contents of taxifolin and quercetin are not significantly changed. So the storage time of Polygonum orientale has no correlation to its quality in 3 years. It shows through the content determination that the content differences of N-p-coumaroyl tyramine acid paprazine in the 10 commercial drugs of Polygonum orientale is obvious, of which the hightest one is 0.032 percents, the lowest is 0.005 percent, and the average is 0.020 percents. The contents of 5 commercial drugs are above 0.020 percents and only one is 0.005 percents.6. The development and study of other medical parts of Polygonum orientale:there is taxifolin only in the leaves of Polygonum orientale by the comparison chemical compents in taxifolin and paprazine in 3 different parts-roots, stems and leaves. It shows by the pharmacological research that the extracts of roots, stems and leaves have the function of inhibition to cell increasing of HeLa,MCF-7,A375-S2 and HepG2 to different extent, the order from high to low for IC50 of HepG2 is roots>stems>leaves, for A375-S2 is roots> stems>leaves, for MCF-7 is roots>stems>leaves and for HeLa is stems>roots> leaves.7. The study of formulation Technology of Polygonum orientale:to filter the rational and effective drugs in traditional medicine theory from literature collected, and combine them into new anti-cancer compounds. The best extraction technology of compound is determined as boiled in water for 3 times by the orthogonal experiment, added respectively 12,10,10 times amounts of water, boiled for 1 hour. The compound is made into tablets and capsules according to the actual medication for taken facilitately.Conclusion:1. It is indicated that the experimental methods are effective and feasible through the following tests:8 compounds are isolated from the n-butanol layer of Polygonum orientale by column chromatography and liquid chromatography and their structures are determined by liquid chromatography, thin layer chromatography and spectral analysis. To filter the activity and determine the contents using the compounds isolated.2.8 compounds are islolated from Polygonum orientale by analyzing chemical components using LC/MS and literature comparison for the first time. It is indicated that the experimental method is effective and feasible. Further study will be going on since the current LC/MS database is not so perfect.3. It is found that the content of quercetin in Polygonum orientale after processed increased significantly for the first time by checking the changes of chemical components before and after processed, which supplies certain scientific references for indicating the processed mechanisms of Polygonum orientale. It is indicated that the the storage time of Polygonum orientale in 3 years has nothing to do with the quality through checking the storage rhythm, which supplies scientific references for safely clinical medication. It supplies specific references for the storage time of Polygonum orientale for the first time.4. It is found that the leaves have obvious anti-cancer effects on cervical cancer cells for the first time with the results of HPLC and pharmacological activity experiments, and the anti-cancer effects of leaves are stronger than that of fruits. It may have something to do with the taxifolin and it supplies scientific references for the development of leaves for further study.5. It supplies an initial basis for new drug development of Polygonum orientale by checking the best extraction technology and taken fomulations, combing anti-cancer componds by filtering the best compatibility with modern pharmacological studies and the analysis of traditional Chinese medicine. |
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