Study On Toxicity Decrease & Effect Improvement Of Dioscorea Bulbifera L. And Its Combination With Angelica Based On Serum Pharmacology Of TCM

Dioscorea bulbifera L. has been widely used for the treatment of goitre and various cancers in recent years, particularly for the treatment of cancers. However, it is discovered that Dioscorea bulbifera L. has a certain toxic reaction in clinical application, which can damage liver and kidney. It is reported that the extent of the damage is relevant to the doses and time. The damage to liver can be demonstrated in a short period, while the damage to kidneys appear after a long time. It was reported in the literature, there are 50 cases of drug-induced hepatitis for the use of Dioscorea bulbifera L., and three persons died. But research shows that the combination of Dioscorea bulbifera L. with angelica could more significantly reduce the damage to liver cells than by simply using Dioscorea bulbifera L. , which can also release the damage to the kidneys. Therefore, this study will further explore the toxic effect of Dioscorea bulbifera L . and the attenuation & synergies mechanisms by combining Dioscorea bulbifera L. with angelica applying to serum pharmacology of traditional Chinese medicine. It has great effects on how to use drug reasonably and safely in clinic.Serum pharmacology of traditional Chinese medicine is a new methodology to develop pharmacology experiments in vitro which suits for the characteristic of TCM. It not only rules out the interference of chemical properties of traditional Chinese medicine itself in the experiment results during pharmacology experiments in vitro , but also reflects transformation process and its results of the drug in the body. Based on the former results, this research will further explore the anti-cancer effect and hepatotoxicity of the Dioscorea bulbifera L. and the attenuation by combination with angelica based on Serum pharmacology of traditional Chinese medicine. Meanwhile the toxicity-effect of Dioscorea bulbifera L. , the impact of angelica on these toxicities and its mechanisms of reducing toxicities were explored through HPLC, MTT test, P-glycoprotein, cell toxicity testing and biochemical indicators etc. The results are as follows:1. Through HPLC analysis, the best time for collecting the serum contained with Dioscorea bulbifera L. and angelica was determined. The results showed that the content of Dioscorea bulbifera L. and angelica in the serum reached a peak after 30 min, but the metabolism was fast. 60～180 minutes later, it was stable. There are many components of Dioscorea bulbifera L. and angelica in blood and the peak was high during this period. So the result was that 60 minutes after administration was the best time for haemospasia.2. To study the inhibition of the serum with Dioscorea bulbifera L. and angelica on the proliferation of carneus cells (S₁₈₀) and liver cancer cells (H₂₂) through MTT test. The results showed that the best time for anti-tumor effect of the serum with Dioscorea bulbifera L. and angelica was at 48h; its inhibition rates were respetively 37.18%, 11.90%, 45.56% and 21.54% .It demonstrated that the pattern by combining Dioscorea bulbifera L. with angelica had a synergistic effect. Angelica may enhance the anti-tumor effect of Dioscorea bulbifera L. through some processes.3. To observe the influence of the serum with Dioscorea bulbifera L. and Chinese Angelica combined on the expression of P-glycoprote in H22 and S₁₈₀. The results showed that the fluorescence intensity of Rho-123 accumulated in cells obtained the highest speed from 0.5h to 1h. It remained steady after reaching the absorption maximum at 1h in the control group, In the Dioscorea bulbifera L . group the fluorescence intensity was significantly lower than the control group after 2h. The fluorescence intensity of compatibility group was significantly higher than Dioscorea bulbifera L. group after reaching absorption maximum at 1h which reduced the expression of P-gp caused by Dioscorea bulbifera L. Therefore it was clarified that the antitumor efficacy of Dioscorea bulbifera L. was strengthened after it combined with Chinese Angelica in compatible proportion via lowering the expression of P-glycoprotein, which is one of the mechanisms for antagonizing the toxicity of Dioscorea bulbifera L.4. To observe and analyze the influence of the serum with Dioscorea bulbifera L. and Chinese Angelica combined on the cell-survival rate of hepatocytes in vitro and the activity of enzymes including GPT, GOT and LDH. The results demonstrated that Dioscorea bulbifera L. decreased cell-survival rate of hepatocytes of normal rats compared with control group. The difference was especially significant(P<0.05). whereas the cell-survival rate in compatibility group significantly increased compared with control group. It had no significant difference between compatibility group and control group (P>0.05). Dioscorea bulbifera L. may cause noticeable damage to the hepatocytes and lead to the leaks of GOT, GPT, LDH of these cells, the activity of GPT, GOT and LDH in supernatant fluid (Cs) obviously increased in comparison with the control group (P<0.01). GOT and GPT were the characteristic zymogram of hepatocyte damage, and the increase of LDH activity indicated that the integrity of Cell membranes were damaged. Therefore the above showed that Dioscorea bulbifera L. has the potential toxicity to liver. The activity of LDH, GOT and GPT in compatibility group was inhibited and had no significant difference compared with the control group (P>0.05). So It indicated that Chinese Angelica had an certain protective effect on hepatocyte.