| Objective:Euphorbia Kansui has many bioactive effects such as purgative actibity, anti-virus, anti-leukemia, anti-tumor, anti-allergy,and antinematodal activity. But its toxi-city and skin irritation have seriously restricted its clini-cal application. This study aim to explore the mechanism of dis-toxicity of Euphorbia Kansui by vinegar-preparing and screen anti-tumor fractions from Euphorbia Kansui.Method:In this experiment, Euphorbia Kansui, two kinds of vinegar-preparing products and their extracts of different polarity solvents as the test samples were studied using different methods.Euphorbia Kansui were baked with high and low temperature differently after soaked enough in vinegar. The products were named vinegar-preparing kansui A and vinegar-preparing kansui B respectively. With Kou's method modified, the acute toxicity of LD50 of Kansui and the other two vinegar-preparing products were determined on mice. Moreover, skin irritation induced by kansui and the other two was investigated in mice. The measure-ment data were analyzed statistically. The high, medium and low doses were designed based on one-thirtieth,one-fiftieth, one-seventieth of LD50 determined above. The mice were administered different doses a week, then blood biochemical parameters of mice were measured, tissue slicing and HE staining were made to observe the morphology changes of liver and kidney tissue. Furthermore, the mice were in metabolic cage and their urine was collected, the diuretic effects of kansui and the two kinds of vinegar-preparing products on mice were observed.The high, medium and low doses were designed based on one-tenth, one-twentieth, one-thirtieth of LD50 determined above.The mice in different groups were administered different doses a week once a day. the survival curves of mice were analyzed statistically. The differents effects of the two vinegar-preparing methods on kansui toxicity were compared.The growth-moduating effects of the different polar parts extracts from Kansui and vinegar-preparing products on the proliferation of MCF-7, A549, HepG2 cells in vitro were observed with MTT colorimetric assay. There were ethanol ex-tract, acetate extract, cyclohexane extract and water extract from Kansui and vinegar-preparing products as samples, they were prepared for different concentration gradient for the test solution. The tests respectively had effects on MCF-7 cells, A549 cells, HepG2 cells in vitro after 48h, the proliferation effects on these tumor cells were observed by OD values detected.The data were analysed statistically.The HPLC-ESI-MS method has been established for analysis the changes of components of kansui before and after vinegar-preparing. Referred to the mass behavior and literature, the chemical components changed significantly have been identified preliminarily.Results:The LD50 values of Kansui and the other two vinegar-preparing products were determined at 20±6mg/g, 92±10mg/g and 105±11mg/g. The acute toxicity and skin irritation tests all indicates vinegar-preparing can reduce the toxicity of Euphorbia Kansui significantly. Moreover, the two processed methods applied for Euphorbia Kansui baking were different obviously. The acute toxicity of kansui after vinegar-preparing reduced significantly according for the LD50 values, and skin irritation decreased greatly. the LD50 values of the two vinegar-preparing products were obvious in difference. (P＜0.05). The liver and kidney toxicity of kansui by vinegar-preparing reduced significantly. The liver and kidney toxicity of vinegar-preparing Kansui B reduced more than which of vinegar preparing kansui A. Above all tests indicate the toxicity of Euphorbia Kansui can be reduced by vinegar preparing, and vinegar-preparing Kansui B with high temperatue has less toxicity, but keep the diuretic effect.The extract of Kansui could obviously reduce the figure of OD of MCF-7, and the inhibiting effect might be in a dose-dependent manner from 200μg/mL to 800μg/mL. Whereas, there were no effects on A549, HepG2 cells. Acetate extract and cyclohexane extract from Kansui could reduce the figure of OD of MCF-7, and IC50 values were 552μg/mL and 378μg/mL. Water extract from Kansui cann't inhibit the proliferation of MCF-7 cell, but it can stimulate the proliferation of A549 and HepG2 cell in high concentration. Acetate extract from vinegar-preparing Kansui kept inhibiting effects on the proliferation of MCF-7, A549 and HepG2 cell in different degree. But water extract of which had neither inhibiting effects nor stimulating effects on proliferation of the tumor cells in vitro cultured.The changes of components of kansui before and after vinegar-preparing have been analyzed with the HPLC-ESI-MS method. Referred to the mass behavior and literature, the 21 chemical components were identified preliminarily. including 19 of these in common; there are 2 chemical components increased and 14 chemical components significantly reduced in kansui by vinegar-baking, and the vinegar-preparing kansui B was lower than the vinegar-preparing kansui A in the rate. By which a component disappeared after vinegar-baking; while four kinds of components in vinegar-preparing kansui B disappeared. In addition, a new chemical composition produced after vinegar-baking, and its peak area of vinegar-preparing kansui B was bigger than vinegar-preparing kansui A.conclusion:It can be inferred that vinegar-preparing kansui B has the least toxicity, according to the acute toxicity test, investigation of liver and kidney toxicity and the diuretic effect, analysis of the survival curves. The method of vinegar-preparing kansui B is better, just baking kansui in 10 minutes at 280℃before soaked enough in vinegar. Baking time and temperature had some effect on the toxicity of Kansui. So the request on baking time and temperature would be necessary in the processing technology of vinegar-preparing Kansui. The results provide a new theoretical basis for the control of baking time and temperature in vinegar-processing and clinical application safely. The study can lay a foundation for explaining the mechanism of detoxification of Euphorbia Kansui through processing.MTT test results show that acetate extracts from kansui and vinegar-preparing Kansui all have cytotoxicity on proli-feration of the tumor cells in vitro cultured in some degree. But water extract in some concentration from kansui has sti-mulating effects on proliferation of the tumor cells in vitro cultured. The water extract from vinegar-preparing kansui has no stimulating effects. All these point out the cancer-inhibitting compotent of Kansui might exist in the acetate extract and the cancer-stimulating component in the water extract from Kansui. So acetate extracts can be regarded as the anti-tumor components of kansui.This study provided a preli-minary basis for screening anti-tumor components of kansui, and a new theoretical basis for the mechanism of detoxification of Euphorbia Kansui through processing from the cellular level.The changes of components of kansui before and after vinegar-preparing have been analyzed with the HPLC-ESI-MS method. Referred to the mass behavior and literature, the 21 chemical components were identified preliminarily. Seen from overall, the changes of components of kansui after processing embody mainly that the contents of majority decreased significantly, minority disappear and new produced. So the mechanism of detoxification of Euphorbia Kansui through processing is to reduce the content of some substances, which of vinegar-preparing kansui B less than vinegar-preparing kansui A. All this revealed the least toxicity of vinegar euphorbia B and the mechanism of detoxification vinegar-preparing from the material basis. Because there is different significantly in the mass behavior of kansui before and after processing,the method resemble fingerprinting mass spectrometry is to standardize TIC of kansui before and after processing. Then it can be used for qualitative or semi-quantitative quality control standards including distinguishing scientifically the vinegar-preparing degree. This study shows that the liquid chromatography-mass spectrometry technique can be used for quality control of vinegar-preparing Euphorbia Kansui.In this study, the mechanism of detoxification of Euphorbia Kansui through processing and anti-tumor fractions from Euphorbia Kansui have been explored, which based on animal experiments, cell level and the material basis of kansui before and after vinegar-processing. On the whole, All the results were significant help in revealing the mechanism of detoxification of Euphorbia Kansui through processing and facilitating understanding of how to promote safer and better use of the herb. The proposed method provides a scientific and technical platform which Euphorbia Kansui could be used as a basis for quality control and safely assurance of herbal preparations...
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