Studies On Injections Of Anti-HIV Components Extrated From Arnebia Euchroma

Ever since the discovery of caffeic acid tetramer（CAT）as an active compound for the cure of HIV,our laboratory made great endeavor to extract and purified anti-HIV components from Arnebia euchroma,with which we can explore promising anti-HIV drugs for clinic use and offer an option in the limited number of traditional anti-HIV Chinese medication.The content of the subject presented here includes:The physical chemical properties of CAT;the extracting and purifying technique;the formulation and preparation of the extract of Arnebia euchroma and the extract of Arnebia euchroma for injection;the ordinary quality control and stability of the preparations,the fingerprint of Arnebia euchroma,semi-product and the two injections;safety test;pharmacokinetic test.The study of the physical chemical properties of CAT revealed that CAT is insoluble in non-polar organic solvents.Its solubility in water soared as the pH increases:5 times more at 6 than 2.CAT is easy to chelate with Cu²⁺,Fe³⁺, structurally changes beyond pH 7,and keeps stable beneath the temperature of 50℃. The light and salt also plays some role on the stability of CAT.Various conditions of were compared to find the optimum extract technology, with the objective to enhance the content of CAT and reduce the unwanted materials in the extract.In the following purifying process,several kinds of macroporous resins were offered and the most proper one was chosen to participate the next exploration of purifying technology.The technique established finally could be described like this: the dried root of Arnebia euchroma was grinded and screened to the diameter under 0.05 mm,then 10 times of 20%ethanol（pH 3.0）was added to get the extract solution after 3 hours at the temperature of 50℃.After repeated the extract process 3 times, the solution was blended and filtered with the 0.45μm microporous film.After that, the solution obtained above was purified by the macroporous resin.With the flow rate of 3 BV·h^-1,the solution went down the resin LK001 column before the eluting process.The eluting course was applying super-purified water and then 20%ethanol condition until no sugar was tested in the eluting liquid.Then 45%ethanol was used to wash down the objective product.The ethanol was removed and the water was evaporated under 50℃.The final product was light brown with more than 83 percent of CAT content in it.The formulations of the injection of the extract of Arnebia euchroma and the extract of Arnebia euchroma for injection were optimized.Adjust the pH to 4.2,then 0.2%Na₂S₂O₅,0.02%EDTA and N₂ were applied to prepare the injection with the concentration of 10 mg·mL^-1.The freeze-drying powder for injection was prepared by adjusting the pH to 4.2 and adding 10%mannitol fallowing with the filtering and freeze-drying process.Muscle,blood vessel,eye and skin irritation tests as well as hemolyty test were carried out for the exploration on the drug safety.The injection of the extract of Arnebia euchroma and the extract of Arnebia euchroma for injection did not show any tendency to causing hemolyty,but coagulation occurred when the volume percentage of the drug solution climbed above 8.Under 6%,the coagulation condition were ameliorated or even disappeared.Negative results were observed in the Muscle,blood vessel,eye and skin irritation tests,demonstrating the security of those preparations.The qualities and accelerating stability of the injection of the extract of Arnebia euchroma and the extract of Arnebia euchroma for injection were studies.Under the accelerating condition,the later were more stable that the former.And they were both stable under the long-term conditions.The HPLC conditions and methods for the construction of chromatographic fingerprints of Arnebia euchroma and the two injections and semi-product were established.Peaks in the chromatogram separated well.The accuracy,reproducibility and stability complied with related requirements. Primary chromatographic fingerprints were constructed.The relationship between them was studied.The pharmacokinetic characteristics of low（12 mg/kg）and high（37 mg/kg）dose of CAT and injection of the extract of Arnebia euchroma and the extract of Arnebia euchroma for injection were studied.The results revealed that they share the same one one compartment model and a relatively fast clearance.Moreover,their biological half lifes(t_1/2)were all shorter than one hour.