Study On The Anti-foot-and-mouth Disease Experimention By Hypericin New Preparation And The Chemosynthesis Of Hypericin

As the tide of "back to the nature"surges day by day, natural Pharmaceuticals become more and more popular in China as to the world for their low side and toxic effects, environmental protection, safety, validity and so on. Being one of the major active ingredients in Hypericum perforakiitum L., Hypencin is reported to have remarkable effects in anti-tumor, anti-virus and anti-depression treatments, but little has been reported about its application in the prevention of acute infectious diseases of animals. The experiment uses the FMDV ( short for foot-and-mouth disease virus, and FMD is one of the OIE list A diseases )as the model to study the Hypericin New preparation effects of anti-RNA virus. Through in-vitro experiments, animal experiment and lymphocyte proliferation of little mice assay to explore the effect for the application of Hypericin New preparation in prevention and control of some epidemic diseases. In addition, this work studies the chemo-synthesis of Hypericin.The details are as follows: (1) Study the anti-FMDV effects of the bio-experiment on Hypericin New preparation1. In-vitro anti-FMDV experimentsThe experiment comparisons among the anti-FMDV effects obtained using Hypericin New preparation together with different density levels of Ribavirin or Zedoray Turmeric Oil in BHK-21 cells and 3—4 -day mice and makes sure the effect of Hypericin New preparation. The result indicates that Hypericin New preparation has obvious effect on anti-virus. In the model of BHK21 cell,Hypericin New preparation can reduce TCID50 from 6.22 to 3.22;In the model of mice, Hypericin New preparation can make two types serum of FMDV reduce 3⁴ LD50 on the concentration of 25mg.mL^-1 and differentia prominence comparison with virus group (P<0.05).2. Animal anti-FMDV experimentsIn this experiment four health cattle free of infection and immunization were divided into two groups at random, 3 for experimental group and 1 for control group. Each cattle was challenged through the tongue at 10000ID₅₀ and meanwhile coarse Hypericin New preparation was administered through drink at 15g.Kg^-1 once a day for the following 7 days. Clinical symptoms showed that in the experimental group one cattle caught light disease, whereas in the control group the cattle caught serious disease. Antibody titers were determined by LB-ELISA and the results showed that the production of antibody in the experimental group was somewhat later than in the control group. Besides, the antibody levels in the former group was lower, which illustrated that Hypericin may have influence on the absorption of virus by the host cells, virus replication or the body's immune conditions.3. lymphocyte proliferation assayThis experiment studied the influence of Hypericin New preparation on the proliferation of mice spleno-lymphocytes. The results showed that all the six concentration of Hypericin New preparation from 50ug.mr' to 800ug.mr' could enable mice sple no-lymphocytes to proliferate in a dose-dependent way(P < 0.01), from which can be inferred that Hypericin New preparation can improve the body's immunity to some extent. (2) Chemo-synthesis of HypericinBased on foreign and domestic literatures, this work explores into the synthesis process of Hypericin chemical reference substances and makes some necessary improvements in the procedure. At 96 °C,under alkaline situation and through co-action with catalysts hydroquinone and caustic alkali, l,6,8-trihydroxy-3-methyl-amthraquinone is transformed into Hypericin by TLC track, pH adjustment, extraction and final preparation. The whole process is under photo-condition for two weeks.The structure of the target product is identified through TLC, Ms ,'HNMR, !3CNMR and DEPT and its content reaches 85%, which is determined by HPLC.