Studies On The Chemical Constituents And Bioavailability Of Saponins From The Leaves And Stems Of Panax Quinquefolium L.

Panax quinquefolium L. as a plant of Araliaceae Panax perennial herbs, is worldwidely known for the precious Medicinal Plants.They are cold-natured, mildly bitter flavor. They also can conduce to lung yin, get rid of deficiency heat, and promote the production of body fluid to quench thirst. Panax quinquefolium L.is originally distributed in virgin forest of the North America. Our country began to cultivate the Panax quinquefolium L. in 1980's. According to the modern science, the main chemical constituents are ginsenosides which are glycosides that contain an aglycone with dammarane skeleton in the Panax quinquefolium L., the leaves and stems of P.ginseng contains the same ginsenosides with the roots, and the content was higher than the roots. In order to making multiple use of resources of Panax quinquefolium L., many scholars from different countries began to study the main active constituents of the aboveground parts in Panax quinquefolium L.(leaves, stems)-Panax quinquefolium L.saponins from leaves and stems.There are many pharmacologically active actions in Panax quinquefolium L.saponins from leaves and stems, such as stay caducity, immunological enhancement, anti-myocardial ischemia and myocardial protection, anti-arrhythmia, anti-hyperhy pertension, anti-cancer and anti-tumor and so on. Therefore, they are of great value in medicinal and have a prospect of a wide range of drug development and researching.As yet, there have been 33 saponins in the leaves and stems of P.ginseng. Panax quinquefolium L. as follow, ginsenoside Rb₂, Rd, Re ,Rg₁, Rb₃, PF₁₁, RT₅, F₂, Rh₁, 20(S)-Rh₂, 20(R)-Rh₂, Rb₁, Rc, Rg₂, M6a, majoroside-F1, Gy-â…¨, Gy-â…©â…¦, quinquenoside-L1, L2, L3, Rh3, 20(S)-Rg₃, 20(R)-Rg₃, 20(S)-Rh₁, 20(R)-Rh₁, 20(S)-Rg₂, 20(R)-Rg₂, Majoroside F₄, F₁, Rg₆, Notoginsenoside Fe, Dammar-23-ene- 3β,6α,12β,20S,25- hydroxy-3- O-β-xylopyraose (1-6)-β- glucopyranosyl-6-O-D-glucosyl-20-O-β-D- glucopyranoside.In this thesis, there is an overview of comparison in the leaves and stems of Panax quinquefolium L.and leaves and stems of ginseng. In the chemical constituents of saponins, main active ingredients in Panax quinquefolium L. and ginseng are triterpenoid saponins, which contain dammarane type, Oleanic acid type and Ocotillol type. Panax quinquefolium L. contain PF11, RT5 and other Ocotillol saponins for identifying characteristics that distinguish it from ginseng. Panax quinquefolium saponins from leaves and stems were major in protopanoxadiol type saponins, but saponins of Ginseng from leaves and stems were major in panaxatriol type saponins.The physiological activity of panaxatriol was higher than panaxadiol, but compared with the panaxatriol, the biological activity of the panaxadiol were more moderate, which may affect the drug-nature differences in Panax quinquefolium L.and ginseng. Consistent with the thesis of traditional Chinese medicine, Panax quinquefolium L. was cool-natured, but ginseng was warm-natured. Therefore, those who can not use ginseng, Panax quinquefolium L. can be used.In order to further study on the chemical constituents of saponins from the leaves and stems of Panax quiquefolium L., this thesis investigate on the saponin composition of stems and leaves of Panax quiquefolium L.cultivated in Fusong county of Jilin Province. 10 kinds of saponin composition were extracted, separated and purified by macroporous, resin column chromatography of silica gel, reversed phase silica column, semi-preparative high performance liquid chromatography and recrystallization methods. Then they were identified according to their physico-chemical properties and spectrum analysis. The structures of them were identified as: RT4, RT5, 24(R)-Ocotillol, 20(S)-Rh1, 20(S)-Rg1, 20(S)-Rg2, 20(S)-Rh2, 20(R)-Rh2, 20(S)-Rg3 and PF11. Compounds RT4 and 24(R)-Ocotillol were extracted and separated from the leaves and stems of Panax quiquefolium L. for the first time.In this thesis, complete ascription in their C, H signal were carried out by 2D-NMR (HMBC, HMQC) for this 10 kinds of triterpenoid saponins.Bioavailability refers to drugs of preparation are absorbed into the rate and extent of the systemic circulation. Absolute bioavailability refers to the ratio of drugs absorbed into the systemic circulation of the quantity and dosage .In the actual work, the ratio of the drug-time area under the curve (AUC) preparation of extravascular administration and intravenous injections was calculated the absolute bioavailability ratio of extravascular administration preparation.In the process of bolting drug, bioavailability is an important parameter and also is an important index in the pharmacokinetic test, currently the domestic require pre-clinical pharmacokinetic trials when reforming dosage forms and submission new drugs. In the the leaves and stems of Panax quiquefolium L., Ginsenoside Rg₃ is one of the main active ingredient, which has anti-tumor, anti-cancer and other biological activitives. It can promote tumor cell apoptosis, inhibit tumor cell proliferation, inhibit tumor cell invasion and metastasis, inhibit tumor angiogenesis, reversing the role of multidrug resistance, affecting tumor cell signal transduction-related gene expression, enhancing tumor patients'immunity and so on. The preliminary biological activity in the cancer study found that ginsenoside Rg₃ can inhibit B16-BL6 lung metastasis. In order to ginsenoside Rg₃ developed into anti-tumor and anti-cancer agents, we must conduct bioavailability studies, and lay the foundation for Clinical Research and development of new drugs.The blood plasma samples were determined by High performance liquid - evaporative light scattering detection (HPLC/ELSD), and carry out the the verification methodology Investigation by specific experiments, the standard curve (calibration curve) of preparation, precision, stability and recovery experiments. The program of"3P87"was applied to process the data, the methods were quick and easy, which show that the approach is consistent with technical requirements of pharmacokinetic test.Adopting the analysis method in this thesis, it is determined of blood concentration -time curve of the five health subject dogs after they were intramuscular and intravenous injection given 20(S)-ginsenoside Rg₃. By"3P87"Pharmacokinetics procedures fitting, pharmacokinetic characteristic was summarized as follow: Dogs were intramuscular given 0.5mg·kg^-1 20(S)-ginsenoside Rg₃, the plasma concentration-time curve assay the main data by"3P87"Pharmacokinetics procedures fitting, the results showed that the pharmacokinetic model were one-compartment model. Pharmacokinetic parameters: T_1/2 (ke) was 0.455h, T (peak) was 0.779h, C (max) was 519.1ng·ml^-1, AUC is 1594.2h·ng·ml^-1; Dog were intravenous injection given 0.5mg·kg^-1 20(S)-ginsenoside Rg₃, the plasma concentration-time curve assay the main data by"3P87"Pharmacokinetics procedures fitting, the results showed that the pharmacokinetic model were one-compartment model. Pharmacokinetic parameters: T_1/2(ke) was 1.557h, V(c)was 0.00136L·kg^-1 , CL was 0.000607L/(kg·h) , AUC was 1648.1h·ng·mL^-1. The bioavailability of Intramuscular 20(S)-ginsenoside Rg₃ in dogs was 96.7%. The results showed that the bioavailability of Intramuscular 20(S)-ginsenoside Rg3 was higher, and intramuscular injection is the appropriate route of administration.In summary, this thesis lucubrated on the chemical constituents and bioavailability of saponins from the leaves and stems of Panax quinquefolium L., 10 saponins was separated and identified, among them pseudo-ginsenoside RT₄ and 24(R)-Ocotillol were found from the leaves and stems of Panax quinquefolium L for the first time.It is significance to the studies on Panax quinquefolium L saponins from leaves and stems and its biological activity. In this thesis, to determine the bioavailability of intramuscular 20(S)-ginsenoside Rg₃ provided the preclinical pharmacokinetics basis for the new drugs'research and development of 20(S)-ginsenoside Rg₃ injection.