JPH04173733A - Platelet aggregation inhibiting agent - Google Patents

Abstract

PURPOSE:To provide the subject inhibiting agent containing a specific polyacetylene compound or sesquiterpene contained in a galenical, Aracylodes lancea L, as an active ingredient and useful as a medicine for thrombosis and circulatory diseases. CONSTITUTION:A platelet aggregation-inhibiting agent contains a compound of formula I, II, III or IV as an active ingredient. The compound is suitably administered in a dose of 10mg to 7g in an oral administration agent or in a dose of 0.5mg to 1.5g in a parenteral administration agent per adult a day. The compounds are obtained as follows; extracting Aracylodes lancea L with a solvent selected from hexane, diethyl ether, petroleum ether, chloroform, acetone, methanol, etc., removing the solvent from the extraction solution, subjecting the obtained residue to a column chromatography or high performance liquid chromatography using a solvent selected from the above-mentioned solvents, acetonitrile, water, etc., once to several times, and further subjecting the obtained fraction to a thin layer chromatography or high performance liquid chromatography.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Field of Application] The present invention relates to polyacetylene compounds and sesquiterpenes that have platelet aggregation inhibiting activity.

[Prior Art and Problems] In recent years, with changes in dietary habits and aging of the population in Japan, the rapid increase in thrombotic diseases such as myocardial infarction and cerebral thrombosis has become a major social problem.

Therefore, therapeutic drugs for this thrombotic disease have been investigated and developed from all aspects of their pharmacological action.

[Means for Solving the Problems] The present inventors have conducted extensive research in search of a compound that has a platelet aggregation inhibitory effect that is effective in treating thrombotic diseases, and as a result, has found a compound that has been widely used clinically. 7ft of herbal medicine (
The present invention was completed based on the discovery that several compounds contained in Atracylodes 1ancea L) or other plants of the same genus have an inhibitory effect on platelet aggregation.

That is, the present invention is a platelet aggregation inhibitor containing a compound represented by the above formulas 1, II, III, and (2) (hereinafter referred to as a compound of the formula) as an active ingredient.

For example, the following methods can be used to obtain the compound of the formula.

That is, blue light is extracted with at least one solvent selected from hexane, diethyl ether, petroleum ether, ethyl acetate, chloroform, acetone, methanol, and ethanol, and the solvent is removed from the resulting extract, and the resulting residue is extracted with hexane. Porous polymers such as Diaion HP-20, MCI Gel CHP20P, Sepha Column chromatography or high-performance liquid chromatography using Sephadex such as Dex LH-20, reversed-phase silica gel, silicage polyamide or cellulose as a carrier once or several times, and then thin-layer chromatography or high-performance liquid chromatography. A polyacetylene compound and a sesquiterpene compound can be obtained by fractionating while confirming the target component by graphing.

Next, an example of manufacturing a compound of the formula is shown below.

Production Example 1 616 g of pulverized blue tuna was subjected to reflux extraction three times with 31 hexane. The solvent of this extract was distilled off under reduced pressure, and extract 45
I got g. This extract was subjected to silica gel column chromatography, and eluted with n-hexane and successively increasing amounts of ethyl acetate. n-hexane: ethyl acetate (4
The fraction eluted in step 1) was subjected to column chromatography using silica gel containing 10% silver nitrate, and n-
9.80 g of the fraction eluted with hexane:ethyl acetate (17:3) (hereinafter referred to as fraction A), then the fraction eluted with n-hexane:ethyl acetate (7:3) (hereinafter referred to as fraction B)2. 30g,
Furthermore, 2.22 g of a fraction eluted with ethyl acetate (hereinafter referred to as fraction C) was obtained.

Fraction B was subjected to silica gel column chromatography, eluted with n-hexane:acetone (75:4), and the first eluted fraction (hereinafter referred to as fraction D)
) and 0.20 g of the next eluted fraction (hereinafter referred to as fraction E). The obtained fraction E was subjected to silica gel column chromatography, and the solvent of the fraction eluted with n-hexane:ethyl acetate (4:1) was distilled off under reduced pressure to obtain 130 mg of a pale yellow eraser.

This pale yellow oily substance exhibits the following physical and chemical properties, and has the physical and chemical properties described in the literature (Washino Inui et al., Journal of the Japanese Society of Agricultural Chemistry, Vol. 60, No. 5, pp. 377-383 (1986)). 4E,6E,12E-tetradecatriene-8,10-diyne-1,3-diyl diacetate (4E,6E,12E-tetrad
ecatrien-8,10-diyn-1,3-di
yldiacetate).

Specific optical rotation: [α]■+15.3° (cm1.05.Chloroform) Mass spectrum 018H2004EI-MS
m/z: 300 [M]+, 240.198.197.180.
179.165.155.153゜152, 141.1
15.93.7'l, 55.43 Infrared absorption spectrum, CC】a cm": 3028, 2960.292
8.2852.2196.1744.1444.137
0°1228, 1126.1042.1020.982
．． 844 ultraviolet absorption spectrum λ nm (log ε):
337 (4,29), 315 (4,42), 296
(4,28), 280 (4,06).

266 (4, 31), 249 (4, 36), 212 (4
, 21) Proton nuclear magnetic resonance spectrum (δppm in CDCl5): 1.82 (3H, d, d, J = 6.9Hz, J
= 1.9Hz).

1.97 (2H, d, t, J = 13.7Hz, J
=6.3Hz), 2.04(3H,s).

2.06 (3H, s), 4.09 (IH, t, J
=6.3Hz), 4.12(LH,m).

5.40 (LH, d, ii, J = 13.7Hz,
J = 7.1Hz).

5.58 (LH, d, J=15.8Hz), 5.7
0 (IH, d, J = 15.5 Hz).

5.74 (LH, d, d, J=15.5Hz, J=
7.1Hz).

6.30 (LH, d, d, J = 15.5Hz, J
= 11.0Hz).

6.33 (IH, d, q, J=15.8Hz, J=
6.9Hz).

6.63 (LH, d, d, J = 15.5Hz,
J = 11.0Hz) 13C-nuclear magnetic resonance spectrum (δppm in CDCl5): 18.93(q), 20.88(q), 21.10
(q), 33.34(t), 60.30(t).

70.81(d), 70.81(d), 72.40
(s), 77.43(s), 79.72(s).

82.12(s), 109.94(d), 111.
80(d), 131.62(d).

133.96(d), 142.91(d), 143
．． 83(d), 170.06(s).

170.89 (s) Production Example 2 Fraction A obtained in Production Example 1 was purified by sublimation under reduced pressure to obtain 2.0 g of a colorless needle-like substance.

This colorless acicular crystal substance exhibits the physical and chemical properties shown below, and has the physical and chemical properties described in the literature (Hiroshi Nishikawa et al., Journal of Pharmaceutical Sciences, Vol. 29, No. 2, pp. 139-146 (1975)). Hinesol (hinesol)
) was determined to be.

Melting point 59.5-60.0°C Specific optical rotation: [α] -43.3° (c = 0.19.chloroform) Mass spectrum EI-MS m/z 222 [M]", 204.189.162 .161.
147.135.133.121゜119, 107.1
05.93.91.81.79.67, 59.55.4
3.41 Infrared absorption spectrum CCI, crn4
.

3620, 3504 (br), 2924.2960.
2924.1660.1470°1454, 1380.
1368.1218.1124.1078.936 Proton nuclear magnetic resonance spectrum (δppm in CDC13): 0.92 (3H, d, J=7.8Hz), 1.20
(3H,s), 1.21(3H,s).

1.68 (3H, d, d, J = 3.4Hz, J
=2.0Hz).

5.31 (LH, t, q, J = 3.7Hz, J
=1.4Hz) 13C-nuclear magnetic resonance spectrum (δppm in CDC13): 16.17(q), 19. Feather (q), 24.18 (
t), 27.69(t), 27.95(t).

28.01(q), 28.42(q), 33.27
(t), 35.74(t), 36°71(d).

48.78(s), 51.43(d), 71.97
(s), 121.69(d), 140.03(s)
Production Example 3 Fraction D obtained in Production Example 1 was purified by sublimation under reduced pressure to obtain 0.4 g of a colorless needle-like substance.

This colorless needle-like substance exhibits the physical and chemical properties shown below, and has the physical and chemical properties described in the literature (Hiroshi Nishikawa et al., Journal of Pharmaceutical Sciences, Vol. 29, No. 2, pp. 139-146 (1975)). β-eudesmol (β-eu
desmol).

Melting point 79.0-81.0°C Specific optical rotation: [α] +63.9' (cm0.39.chloroform) Mass spectrum EI-MS m/z; 222 [M]'', 204.・189.164. 161
．． 149.133.121.109°108, 107.
105.95.93.91.81.79.67, 59 Infrared absorption spectrum 'mad ""'3620, 3
488(br), 3080.2972.2936.2
868.2844°1644, 1468.1454.1
440.1380.1216.1190.1150゜1
124, 1090.986.958.932.888.
856 proton nuclear magnetic resonance spectrum (δppm in CDC1a): 0.70 (3H, s), 1.20 (6H, s).

4.45 (IH, d, d, J = 3.5Hz, J =
1.8Hz).

4.71 (IH, d, d, J=3.5Hz, J=
1.7Hz) 13C-nuclear magnetic resonance spectrum (δppm in CDCl3): 16.33(q
), 22.43(t), 23.54(t), 25
．． 08(t), 27.17(q).

27.22(q), 35.92(s), 36.95
(t), 41.21(t), 41.92(t).

49.52(d), 49.87(d), 72.87
(s), 105.36(t), 151.13(s)
Production Example 4 Fraction C obtained in Production Example 1 was subjected to silica gel column chromatography, and benzene:ethyl acetate (1
The fraction eluted at 0:1) was purified by sublimation under reduced pressure to obtain 0.5 g of a colorless needle-like substance.

This colorless acicular crystal substance exhibits the physical and chemical properties shown below, and is described in the literature (Hiroshi Saiyo et al., Journal of Pharmaceutical Sciences, Vol. 29, No. 2, pp. 139-146 (1975)). Elemol (elemol)
It was decided that

Melting point 51.5-53.0°C Specific optical rotation: [α12'+4.3° (c = 0.19, chloroform) Mass spectrum EI-MS m/z: 222 [M]'', 204.189.161. 147.
135.133.121.119゜107, 105.9
3.91.81.79.68.67, 55.43.41
Infrared absorption spectrum, CCl4c, 4.

3620, 3480 (br), 3084.2972.
2936.2860.1744゜1640, 1468.
1442.1414.1378.1162.1110.
1060°1004, 932.910.892.846
Proton nuclear magnetic resonance spectrum (δppm in CDC13): 0.99 (3H, s), 1.20 (6H, s).

1.71 (3H, d, d, J=2.3Hz, J=0
．． 9Hz).

1.96 (LH, d, d, J=12.4Hz, J
= 3.3Hz).

4.59 (IH, d, q, J=1.8Hz, J=1
．． 5Hz).

4.82 (IH, d, q, J=1.8Hz, J=1
．． 5Hz).

4.89 (IH, d, d, J = 10.7Hz,
J = 1.3Hz).

4.90 (IH, d, d, J = 17.7Hz, J
= 1.3Hz).

5.81 (IH, d, d, J = 17.7Hz, J
= 10.7Hz) 13C-nuclear magnetic resonance spectrum (δppm in CDC1a): 16.63 (q), 22.58 (t), 24.78
(q), 27.17(q), 27.21(q).

28.52(t), 39.71(s), 39.90
(t), 49.37(d), 52.76(d).

72.72(s), 109.90(t), 112.
07(t), 147.90(s), 150.25(
d) Next, the fact that the compound of the formula has an excellent platelet aggregation inhibitory effect and is useful as a drug for thrombotic diseases and the cardiovascular system will be explained using experimental examples.

Experimental Example 1 (Platelet aggregation inhibitory effect) Preparation of platelet-rich plasma Nine volumes of arterial blood were collected from the lower limb femoral artery of a rabbit into a polypropylene syringe containing one volume of 3.8% sodium citrate. The collected blood was heated to 800°C at room temperature.
Centrifugation at rpm for 10 minutes, and the supernatant was diluted with platelet-rich plasma (p
latelet-rich-plasma (PRP). This residue was centrifuged for 15 minutes at OOO rpm and the supernatant was collected as platelet-poor plasma.
-plasma, PPP). Dilute PRP using this PPP, and the platelet count will be 2.5 to 3.2.
The density was adjusted to 105 pieces/mm'.

(2) Measurement of platelet aggregation ability with arachidonic acid To 4 ml of PRPo obtained as above, 50. ．． 1 and incubate for 2 minutes at 37°C, then add arachidonic acid solution (final concentration 8
0 to 100 □g/ml) to induce aggregation, and the agglutination ability was measured. As a control, 2% ethanol-saline was used instead of adding the compound of formula. The measurement is performed using PAYTONAGGREGATION MODULE (
MODEL600B, PAYTON ASSOCIA
The transmittance at the time of maximum aggregation was measured by nephelometric method using TES), and the aggregation inhibition rate was determined when the control was set as 100%. The results are shown in Table 1.

Table 1 [Experiment Example 2] ■Measurement of aggregation ability with collagen Table 2 shows the results of measurements carried out in the same manner as above, except that a collagen solution (final concentration 10-20□g/ml) was added instead of the arachidonic acid solution. Shown below.

Table 2 Next, an acute toxicity test of the compound obtained in the production example was conducted using ICR male mice, and there was no death after oral administration of Ig/kg.

Next, formulation of the compound of the formula will be explained.

The compounds of the formula can be administered to animals and humans neat or with conventional pharmaceutical carriers. The dosage form is not particularly limited and can be selected and used as necessary, and includes oral preparations such as tablets, capsules, granules, fine granules, and powders, and parenteral preparations such as injections and half-open forms. .

In order to exert the desired effect as an oral agent, the weight of the compound of the formula for an adult is usually 10 rng to 7. It seems appropriate to take the drug in divided doses several times a day.

Oral agents include, for example, starch, lactose, sucrose, mannitol,
It is manufactured using conventional methods using carboxymethyl cellulose, corn starch, inorganic salts, etc.

In addition to the excipients mentioned above, binders, disintegrants, surfactants, lubricants, fluidity promoters, flavoring agents, coloring agents, fragrances, and the like can be used in this type of preparation as appropriate.

Specific examples of each are shown below.

[Bound Saige starch, dextrin, gum arabic powder, gelatin,
Hydroxypropyl starch, methylcellulose, sodium carboxymethylcellulose, hydroxypropylcellulose, crystalline cellulose, ethylcellulose, polyvinylpyrrolidone, macrogol.

[Disintegrant] Starch, hydroxypropyl starch, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, carboxymethyl cellulose, low substituted hydroxypropyl cellulose.

[Surfactant 1 Sodium lauryl sulfate, soybean lecithin, sucrose fatty acid ester, polysorbate 80° [Nasawa Saizuki talc, waxes, hydrogenated vegetable oil, sucrose fatty acid ester, magnesium stearate, calcium stearate, aluminum stearate, Polyethylene glycol.

[Fluidity promoter] Light anhydrous silicic acid, dry water 1 sod aluminum gel, synthetic aluminum silicate, magnesium silicate.

The compound of the formula can also be administered as a suspension, emulsion, syrup, or elixir, and in order to achieve the desired effect as a parenteral agent, the age, weight, and disease of the patient are important. Although it varies depending on the severity, the usual adult dose of the compound of the formula is 0.5 mg to 1.5 mg per day. Intravenous injection, intravenous drip, subcutaneous injection, and intramuscular injection are considered appropriate. These various dosage forms may contain flavoring agents and coloring agents.

Parenteral preparations are prepared according to conventional methods, and diluents generally include vegetable oil for injection, sesame oil, peanut oil, and diced oil. Yu, corn? Therefore, propylene glycol, polyethylene glycol, etc. can be used.

Furthermore, a bactericide, a preservative, and a stabilizer may be added as necessary. In addition, from the viewpoint of stability, this parenteral preparation may be filled into a vial or the like, then frozen, water removed by ordinary freeze-drying techniques, and a liquid preparation prepared from the freeze-dried product immediately before use.

Furthermore, isotonizing agents, stabilizers, preservatives, soothing agents, etc. may be added as appropriate.

Other parenteral preparations include liquid preparations for external use, liniments such as ointments, half-open preparations for intrarectal administration, etc., and are manufactured according to conventional methods.

[Example 1] ■Corn starch 44g■Crystalline cellulose 40g■Carboxymethyl cellulose calcium 5g ■Light silicic anhydride 0.5g■Magnesium stearate 0.5g6 Compound obtained in Production Example 1 10 Total 100g According to the above recipe ■～■ The mixture was mixed uniformly and compressed using a tablet machine to obtain 200 mg tablets.

This tablet contains 20 mg of the compound obtained in Production Example 1.
It contains 3 to 10 tablets per day for adults, divided into several doses.

[Example 2] ■Crystalline cellulose 84.5g■Magnesium stearate 0.5g■Carboxymethyl cellulose calcium 5g 1 to 100 g A portion of the mixture was uniformly mixed, compression molded, pulverized, and the remaining amounts of ■ and ■ were added and mixed, and compressed using a tablet machine to obtain a portion of 200 mg tablets.

This tablet contains 20 mg of the compound obtained in Production Example 2.
It contains 3 to 10 tablets per day for adults, divided into several doses.

[Example 3] ■ Crystalline cellulose 34.5 g ■ 10% hydroxypropyl cellulose ethanol solution 50 g ■ Carboxymethylcellulose calcium 5 g ■ Magnesium stearate 0.5 g Compound obtained in Production Example 3 10 Total 100 g According to the above recipe ■, ■ and ■ are homogeneously mixed, and then the mixture is made into a net by a conventional method, granulated using an extrusion granulator, dried and crushed. 200 mg tablets were obtained.

This tablet contains 20 mg of the compound obtained in Production Example 3.
It contains 3 to 10 tablets per day for adults, divided into several doses.

[Example 4] ■Corn starch 84g ■Magnesium stearate 0.5g ■Carboxymethylcellulose calcium 5g ■Light silicic anhydride 0.5g Compound 10 obtained in Production Example 4 Total 100g Mix ■~■ uniformly according to the above recipe After compression molding using a compression molding machine, the mixture was crushed using a crusher and sieved to obtain granules.

1 g of this granule contains 100% of the compound obtained in Production Example 4.
It contains 0.6 to 2 g per day for adults, divided into several doses.

[Example 5] (1) Crystalline cellulose 55 g (2) 10% hydroxypropyl cellulose ethanol solution 35 g (3) Compound obtained in Production Example 1 (10) Total 100 g (1) to (2) were uniformly mixed according to the above recipe and then stirred. After granulation using an extrusion granulator, the mixture was dried and sieved to obtain granules.

1 g of this granule contains 100% of the compound obtained in Production Example 1.
It contains 0.66 to 2 g per day for adults, divided into several doses.

[Example 6] ■ Corn starch 89.5 g ■ Light anhydrous silicic acid 0.5 g 3 Compound obtained in Production Example 2 10 Total 100 g Mix ■ to ■ uniformly according to the above recipe, and give 200 mg
was filled into a No. 2 capsule.

One capsule of this preparation contains 20 mg of the compound obtained in Production Example 2, and adults should take 3 to 1 capsules in several doses per day.

[Example 7] ■ Distilled water for injection 89.5g ■ Dog bean oil
5g■ dog bean phospholipid
2.5g ■ Glycerin 2
g Compound obtained in Production Example 3 1 Total amount 100g Dissolve ■ in ■ and ■ according to the above recipe, and add ■
The solutions of (1) and (2) were added and emulsified to obtain an injection.

Claims (4)

[Claims] (1) Formula I ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ I A platelet aggregation inhibitor containing the compound represented by as an active ingredient. (2) Formula II ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ II A platelet aggregation inhibitor containing the compound represented by as an active ingredient. (3) Formula III ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ III A platelet aggregation inhibitor containing the compound represented by as an active ingredient. (4) Formula IV ▲Contains mathematical formulas, chemical formulas, tables, etc.▼ IV A platelet aggregation inhibitor containing the compound represented by as an active ingredient.