KR19990015090A - Antagonist for platelet activating factor receptor binding, containing (20S) -ginsenoside Rg₃ and / or ginsenoside Rg₃ - Google Patents

Abstract

The present invention relates to antagonists for Platelet-Activating Factor (PAF), which contain (20S) -ginsenoside Rg ₃ and / or ginsenoside Rg ₅ .

Description

Antagonist for platelet activating factor receptor binding, containing (20S) -ginsenoside Rg₃ and / or ginsenoside Rg5

The present invention relates to antagonists for platelet activating factor receptor binding, containing (20S) -ginsenoside Rg ₃ and / or ginsenoside Rg _5, and more particularly, to (20S) -ginsenoside Rg ₃ and Newly discovered that ginsenoside Rg ₅ has antagonistic activity against platelet activator receptor binding and utilizing these antagonistic activities of (20S) -ginsenoside Rg ₃ and ginsenoside Rg ₅ in relation to platelet activating factor An antagonist to platelet activating factor receptor binding that can be usefully used in the treatment of inflammatory diseases and the like.

Platelet-Activating Factor (PAF) is a factor that activates platelets by stimulating basophils of IgE-sensitized rabbits with antigens and activating platelets (Benveniste, J., PM Henson and CG Cochrane (1972) Leucocyte dependent histamine release from rabbit platelet, The role of Ig E, basophils and a platelet activating factor.J.Exp. Med. 1356-1377.), multinuclear leukocytes (neutrophils, eosinophils, basophils), monocytes, macrophages, mast cells, etc. Produced in immune cells, vascular endothelial cells and platelets (Benveniste, J. and B. Arnoux (1983) Platelet-activating factor and structurally related ether-lipids, Elsevier, Amsterdam., Benveniste, J. and BB Vargaftig (1983) Platelet -activating factor, An ether lipid with biological activity in ether lipids in 'Biochemical and Biomedical Aspects' HK Magnold and F. Paltauf.ed.Academic Press, New York, 355-476., TC Lee and F. Snyder. (1984) 'Function, metabolism, and regulation of platelet-a ctivating factor and related ether lipids, in phospholipids and cellular regulation. 'JF Kuo ed.CRC Press, Boca Raton, F1, 1-39). The physiological function of platelet activators is 1 / 1,000 to 1 / 10,000 lower concentrations (10 ^-9 to 10 ^-10 M) than inflammatory mediators such as histamine, leukotrienes and prostaglandins. Is an initiator of an acute inflammatory response that activates inflammatory cells to induce the synthesis and release of other inflammatory mediators (Braquet, P., T. Touqui, TY Shen and BB Vergaftig (1987) Perspective in platelet-activating factor research, Pharmacol Rev. 39 , 97-145). When the inflammatory response of platelet activator is activated, it causes various inflammatory diseases such as bronchial contraction, asthma, tissue transplant rejection, cardiac anaphylaxy, endotoxin shock, gastric ulcer, acute allergic reaction, and irritable dermatitis (Gonzalez-Crussi). , F. and W. Hsueh (1983) Experimental model of ischemic bowl necrosis.The role of platelet-activating factor and endotoxin.Am J. Pathol. 112 , 127-135., Katoh, N., M. Itoh, Y Yokoyama and T. Joh (1994) Evaluation of the mechanism of endothelin-induced rat gastric mucosal lesion formation.Nippon Shokakibyo Gakkai Zasshi, 91 , 27-35., Frerichs, KU, PJ Lindsberg, JM Hallenbeck and GZ Feuerstein (1990). Platelet-activating factor and progressive brain damage following focal brain injury.J. Neurosurg. 73 , 223-233., Tokutomi, T., M. Sigemori, T. Kikuchi and M. Hirohata (1994) Effect of platelet-activating factor antagonist on brain injury in rats. Acta. Neurochir. suppl. Wien. 60, 508-510., Makowk a, L., FA Chapman, DV Cramer, SG Qian, H. Sun and TE Starzl (1990) Platelet-activating factor antagonists, SRI 63-441, on rejection of xenografts and allografts in sensitized hosts. Transplantation 50 , 359-365.).

However, if the platelet activator does not bind to the receptor, it is inactivated by the hydrolytic enzyme acetylhydrolase in plasma or cells to lyso-PAF (Braquet, P., T. Touqui). , TY Shen and BB Vergaftig (1987) Perspective in platelet-activating factor research, Pharmacol. Rev. 39 , 97-145), antagonizing the binding of platelet activating factors to platelet membrane receptors for the purpose of improving inflammatory diseases. The search for interfering substances has been made. Platelet activator-like structural antagonists include CV 3988 (Terashita, ZI, S. Tsushima, Y. Yoshioka, H. Nomura, Y. Inada and K. Nishikawa (1983) CV-3988, A specific antagonist of platelet activating factor (PAF) Life Sci. 32. 1975-1982), CV 6209 (Terashita, ZI, Y. Imura, M. Takatani, S. Tsushima and K. Nishikawa (1986) CV-6209, A highly potent platelet activating factor (PAF) antagonist. (abstract) .Gatlingurg, Tennessee, October, p. 29), ONO 6240 (Miyamoto, T., H. Ohno, T. Yano, T. Okada, N. Hamanaka and A. Kawasaki (1983) Ono-6240 , A new potent antagonist of platelet activating factor, Third International Congress of Inflammation, Paris, Sept, Abstr. 513-519, SRI series (Lee, ML, CM Winslow, C. Jaeggi, F. D'Aries, G. Frisch) , C. Farley, MK Melden, DA Handly and RN Saunders (1985) Inhibition of platelet activating factor, Synthesis and biological activity of SRI 63-073, a new phospholipid PAF-acether antagonist in, Is there a Case for PAF-acether Antagonists ?, Paris, June, Abstr. A4) Natural product-derived platelet activator antagonists are Ginkolide B (Braquet, P (1985) BM52021 and related compounds, a new series of highly specific PAF-acether receptor antagonists. Prostaglandins 30 , 689), Kadsurenone (Shen, TY, SB Hwang, MN Chang, TW Doebber, MH Lam, MS Wu, X. Wang, GQ Han and RZ Li (1985) Characterization of a platelet -activating factor receptor antagonist isolated from haifenteng (Piper futokadsura) : Specific inhibition of in vitro and in vivo platelet-activating factor-induced effects.Proc. Natl. Acad. Sci. USA 82 , 672-676. Coumarin (Takeuchi, N., T. Kasama, K.Mayuzumi, K. Tamura, H. Fukaya, T. Ohono and S. Tobinaga (1988) The antagonist effects of compounds derived from Khellacton on platelet-activating factor. Chem. Pharm. Bull. 36 , 4221-4224), Magnolin (Pan, JX, OD Hensens, DL Wink, MN Chang and SB Hwang (1987) Lignans with platelet activating factor antagonist activity from Magnolia biondii. Phytochemistry 26, 1377-1379).

Meanwhile, Panax Ginseng is a perennial herb belonging to the Araliaceae family, and its root is called Ginseng Radix, and the tonic, Gangjeong, hematopoiesis, and warmth ), Health, fatigue, mental stability, sedative effects, etc. (Korea Ginseng Research Institute, Ginseng Research Institute, Daesung Print Co., p. 143 (1985)., Korean ginseng, Korea Ginseng and Tobacco Research Institute, Cheonil Print Co., p. 89 (1994).) Unlike other plants, ginseng saponin is not toxic and does not show hemolytic activity below 0.001%. (田中 人, 藥用 人蔘 共 立 出版, p. 43 (1981)., Nak-Doo Kim (1978), Korean Ginseng, Korea Ginseng Research Institute, p. 118.) Central nervous inhibitory activity (Tanaki, K., Saito, H. and Nabata, H. (1972) J. Pharmacol . 33, 245., Katu, Y. , Miyata, T., Urano, T., Sato, I. and Kinoshita, A. (1975) Arzneim-Forsch (Drug Res.) (1975), 25, 4.) , Protein synthesis (Yokozawa, T. and Oura, H. (1990), J. Nat. Prod. , 53 , 1514., Yamamoto, M., Takeuchi, N., Yamada, K., Hayashi, Y., Hirai, A and Kumakai, K. (1978) Arzneim-Forsch. 27 , 1169., Yamamoto, M., Masaki, M., Yamada, K., Hayashi, Y., Hirai, A. and Kumakai, K. (1978) Arzneim-Forsch. 28 , 2238.), corticosteroid secretion promoting activity (Pearec, PT, Zois, I., Wynne, KN and Funder, JW (1982) Endocrinol. 29 , 567., Han Byung-hoon, Jang Il-mu (1977), Korean Journal of Ginseng 2 , 17), Platelet Aggregation Inhibition (田村 泰 (1992), The Ginseng Review , 13 , 198., Nakanishi and Onishi (1992), Journal of Medicinal Ginseng Research Abstracts Collection (9), Medicinal Ginseng Research Society, p . 12.) Promoting Absorption of Neutral Lipids (Cho Chung-no, Im-Soon Choi, Lee Sang-jik, Sung-Hee Cho, Myung-Hee Son (1973), Korean Journal of Biochemistry 6 , 185., Joo, CN, Cho, YD, Koo, JH, Kim, CW and Lee, SJ ( 1977) Korean Biochem.J. 13. 1.), Immune Enhancement Activity (Won Young Lee, Woo Nam Kim, Joo Deok Kim, Eun Hee Ko, Byung Soo Kim (1988) Korean Cancer Society 20 , 126., Kim, YS, Kang, KS and Kim, SI (1991) Korean J. Ginseng Sci. 15 , 13., Kim Mi-jung, Jung Nopal (1987) Korean Ginseng Society 11 , 119)

Red ginseng is a ginseng product that has been processed for 6 years to produce 12.5-13.5% water content through red ginseng, drying, etc., and the browning reaction is promoted by non-enzymatic browning during the manufacturing process. It is a ginseng product processed into a very hard form and maintained in its original form (Korea Ginseng & Tobacco Research Institute (1993), Korea Ginseng, Cheonil Printed Firm, pp. 44).

Therefore, the present inventors studied the antagonistic activity of platelet activating factor in a substance contained in red ginseng, indicating that (20S) -ginsenoside Rg ₃ and ginsenoside Rg ₅ have antagonistic activity against platelet activating factor. The present invention has been completed.

An object of the present invention is to provide an antagonist for platelet activator receptor binding, which contains (20S) -ginsenoside Rg ₃ and / or ginsenoside Rg ₅ .

The present invention provides an antagonist for platelet activating factors, containing (20S) -ginsenoside Rg ₃ represented by formula (1) and / or ginsenoside Rg ₅ represented by formula (2).

Antagonists against platelet activators of the invention are used to modulate PAF activity in vivo or ex vivo. Antagonists against platelet activators of the present invention can be used as an inflammatory therapy or an inflammatory therapy adjuvant. That is, the antagonist against platelet activating factor of the present invention can be usefully used for inflammatory diseases associated with platelet activating factor and pregnancy and egg implantation control. Inflammatory diseases that can be treated by antagonists against platelet activating factors of the present invention include asthma, allergies, arthritis, rheumatoid, thrombosis, acute inflammation, systemic hypersensitivity, retinitis, hepatitis, enteritis, pancreatitis, nephritis, rhinitis, atopic dermatitis, Cardiac hypersensitivity, endotoxin shock, central nervous system damage, or organ (tissue) transplant rejection.

Antagonists for platelet activating agents of the invention may be prepared in unit dose form by formulating with (20S) -ginsenoside Rg ₃ and / or ginsenoside Rg _{5 of the invention} and a pharmaceutically acceptable carrier, excipient or Or by incorporating into a multi-dose container.

Formulation forms may be in the form of solutions, suspensions or emulsions in oils or aqueous media or in the form of extracts, powders, granules, tablets, capsules, and may contain dispersants, suspensions or stabilizers.

(20S) -ginsenoside Rg ₃ and / or ginsenoside Rg ₅ having antagonistic activity against platelet activating factor receptor binding of the present invention is typically isolated, prepared from extraction from red ginseng or diol-based saponins of ginseng It can be prepared in high yield by treating with acetic acid.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, the following examples are only illustrative of the present invention and the present invention should not be considered as limited by the following examples.

Example 1 (20S) -ginsenoside Rg from Red Ginseng ₃ And ginsenoside Rg ₅ Manufacture

3 kg of red ginseng was ground and extracted three times for 2 days using 10 L of 95% methanol at 5 ° C. The filtrates were collected and concentrated under reduced pressure to obtain 540 g of extract. The extract was dissolved in 5 L of distilled water, washed three times sequentially with 2 L of petroleum ether, and the aqueous layer was extracted three times with 2 L of n-butanol to obtain 320 g of butanol fraction.

The butanol fraction was dissolved in 1 L of methanol, and then cooled to remove the precipitate, followed by silica gel column chromatography (CHCl ₃ / CH ₃ OH / H ₂ O, 9: 3: 1, v / v) to obtain pure ginsenoside Rg _5. 480 mg and 1.5 g of (20R) -ginsenoside Rg ₃ and (20S) -ginsenoside Rg ₃ racemic mixture were obtained. 1.5 g of the (20R) -ginsenoside Rg ₃ and (20S) -ginsenoside Rg ₃ racemic mixture were suspended in 30 ml of methanol by heating and sonication, cooled to remove the precipitate, and the filtrate was concentrated under reduced pressure. The concentrate under reduced pressure (720 mg) was dissolved in pyridine (20 mL), acetic anhydride (20 mL) was added under ice cooling, followed by stirring at room temperature for 10 hours. The reaction product was partitioned into ethyl acetate (100 mL), extracted three times, washed three times with 5% HCl (150 mL), three times with saturated aqueous NaHCO ₃ (150 mL), and three times with distilled water, and then ethyl acetate. Fractions were concentrated under reduced pressure. Concentration under reduced pressure (940 mg) was carried out by silica gel column chromatography (CH ₂ Cl ₂ / ethyl acetate, 8: 1, v / v) to give (20S) -ginsenoside Rg ₃ peracetate (620 mg). It was dissolved in 5% HCl / CH ₃ OH (50 mL) and stirred at room temperature for 5 hours. Distilled water (100 mL) was added to the reaction solution, and the mixture was concentrated under reduced pressure at 30 ° C. to remove methanol, and then partitioned and extracted three times with butanol (50 mL). The butanol fraction was concentrated under reduced pressure and purified by silica gel chromatography (CHCl ₃ / CH ₃ OH / H ₂ O, 8: 3: 1, v / v) to obtain pure (20S) -ginsenoside Rg ₃ (390 mg).

Example 2 (20S) -ginsenoside Rg from diol-based saponins ₃ And ginsenoside de Rg ₅ Manufacture

Ginsenosides Rb ₁ , Rb ₂ and Rc mixture (5 g) were dissolved in 50% acetic acid (500 mL) and sonicated at 70 ° C. for 3 hours. The reaction solution was concentrated under reduced pressure at 70 ° C. to about 1/10 volume (50 mL), and the precipitate was left to stand in a 4 ° C. refrigerator overnight to remove the precipitate by filtration. The filtrate was diluted with water (500 mL) and extracted three times with n-butanol (250 mL). The butanol extract was washed with 2 포화 saturated aqueous NHCO ₃ solution and concentrated under reduced pressure, and the concentrate was purified by silica gel column chromatography (CHCl ₃ / CH ₃ OH / H ₂ O, 9: 3: 1, v / v) to give (20S)- Cenoside Rg ₃ (1.5 g) and ginsenoside Rg ₅ (1.2 g) were obtained.

(20S) -Ginsenoside Rg ₃ carbon nuclear magnetic resonance spectrum (pyridine-d ₅ , δ)-15.7, 16.3, 16.3, 16.4, 17.4, 18.3, 22.4, 25.2, 26.1, 26.2, 27.5, 30.7, 31.4, 35.3, 36.3, 37.3, 38.5, 39.1, 39.4, 47.9, 49.8, 51.1, 54.2, 55.7, 62.0, 62.2, 70.4, 70.0, 72.3, 72.4, 76.5, 77.3, 77.5, 77.8, 78.2, 82.8, 88.3, 104.5, 105.4, 126.9, 130.1

And carbon nuclear magnetic resonance spectra of ginsenoside Rg ₅ (pyridine-d ₅ , δ)-13.1, 15.7, 16.3, 16.5, 16.9, 17.7, 18.3, 25.6, 26.6, 27.4, 28.0, 28.7, 32.1, 32.5, 35.2 , 36.9, 39.2, 39.6, 40.1, 50.3, 50.7, 50.8, 50.9, 56.3, 62.6, 62.7, 71.5, 71.5, 72.5, 77.0, 77.8, 78.0, 78.1, 78.2, 83.3, 88.8, 105.0, 123.2, 123.5, 131.1 , 140.1

Experimental Example (20S) -ginsenoside Rg ₃ And ginsenoside Rg ₅ Platelet Activation Factor Antagonism Test

Antagonistic tests for platelet activator receptor binding of (20S) -ginsenoside Rg ₃ and ginsenoside Rg ₅ were carried out by methods of Valone, FH, E. Coles, VR Reinhold. and EJ Goetzl (1982) Specific binding of phospholipid platelet-activating factor by human platelets.J. Immunol. 129. 1637-1641) and the method of Yang, HO et al (Yang, HO, DY Suh and BH Han (1995) Isolation and characterization of platelet-activating factor receptor binding antagonists from Biota orientalis.Planta Med. 61 , 37-40), and the addition of ginseng saponins from alcohol extracts of ginseng, The phenomenon of inhibiting specific binding was tested. The results are shown in Table 1.

Antagonistic Activity of Platelet Activator Factor Binding of (20S) -ginsenoside Rg ₃ and Ginsenoside Rg ₅ (20S) -ginsenoside Rg ₃ Ginsenoside Rg ₅ Concentration (micromolar) PAF receptor binding inhibition rate (%) Concentration (micromolar) PAF receptor binding inhibition rate (%) 200 84.4 160 73.0 100 56.3 80 48.2 25 40.3 40 29.3 12.5 20.0 20 24.4 IC ₅₀ = 4.9 x 10 ^-5 M IC ₅₀ = 9.2 x 10 ^-5 M

The present invention provides an antagonist to platelet activating factor, which contains (20S) -ginsenoside Rg ₃ represented by Formula 1 and / or ginsenoside Rg ₅ represented by Formula 2, wherein the platelet activation of the present invention is provided. It can be seen that the antagonistic activity against the factor is excellent as shown in Table 1 above. Therefore, the antagonist of the platelet activating factor of the present invention can be used for the regulation of platelet activating factor in vivo or ex vivo, and also inflammatory diseases associated with platelet activating factor, for example, asthma, allergy, arthritis, rheumatoid, thrombosis, Inflammatory diseases such as acute inflammation, systemic hypersensitivity, retinitis, hepatitis, enteritis, pancreatitis, nephritis, rhinitis, atopic dermatitis, heart hypersensitivity, endotoxin shock, central nervous system damage, or organ (tissue) rejection It can be usefully used.

Claims (4)

An antagonist to platelet activator receptor binding, which contains at least one compound selected from the group consisting of (20S) -ginsenoside Rg ₃ and ginsenoside Rg ₅ . The antagonist of platelet activator receptor binding according to claim 1, wherein the antagonist is used as an inflammatory agent or an inflammatory agent. 3. The antagonist of platelet activator receptor binding according to claim 2, wherein the inflammatory agent or an inflammatory therapeutic agent is useful for inflammatory diseases associated with platelet activators and for controlling pregnancy and egg implantation. The method of claim 3, wherein the inflammatory disease associated with platelet activator is asthma, allergy, arthritis, rheumatism, thrombosis, acute inflammation, systemic hypersensitivity, retinitis, hepatitis, enteritis, pancreatitis, nephritis, rhinitis, atopic dermatitis, heart hypersensitivity, internal An antagonist to platelet activator receptor binding, characterized in that the toxin shock, central nervous system damage or organ (tissue) transplant rejection.