JPH0625001A - Vascular endothelium growth promoter - Google Patents

Abstract

PURPOSE:To provide a vascular endothelium growth promoter useful for preventing and improving arterial sclerosis caused by thrombus, etc., as a malignant factor, arterial thromboembolism, etc. CONSTITUTION:This invented vascular endothelium growth promoter contains a polyvalent alcoholic substance having about 8000 molecular weight obtained by ultrafiltering or dialyzing an extract of a gardenia and carrying out gel- filtration or ion-exchange treatment of the resultant low-molecular fraction and glycerol or D-mannitol as the active components.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a vascular endothelial cell growth promoter useful for the prevention and treatment of arteriosclerosis and arterial thromboembolism, etc., in which vascular endothelial cells are proliferated and adverse factors include thrombus. Is.

[0002]

2. Description of the Related Art Thrombi are formed by the interaction of vascular endothelium, fluid blood coagulation and platelets in blood vessels and excessive formation of fimbrin. At that time, in the way of forming an arterial thrombus, no thrombus is formed without damage to the vascular endothelium. Moreover, it is said that it is not necessary to be a significant obstacle, but rather a slight obstacle is more likely to be formed.

Therefore, if the proliferation of vascular endothelial cells can be promoted, it is useful for the prevention and treatment of arteriosclerosis and arterial thromboembolism, etc., which are adverse factors such as thrombus.

Up to now, several proteins called growth factors have been known to promote the growth of vascular endothelial cells, but all of them are difficult to isolate and purify and are unstable, and they are useful as drugs. Is hard to say. Heparin is also
It is one of the substances that enhances the growth of vascular endothelial cells stimulated by growth factors, but does not itself have the action of promoting the growth of vascular endothelial cells.

Therefore, it has been desired to develop a substance having a vascular endothelial cell growth promoting action.

[0006]

Means for Solving the Problems The inventors of the present invention have conducted extensive studies in order to find a substance having a vascular endothelial cell growth-promoting effect, and as a result, a crude drug which is a ripe fruit of gardenia of Rubiaaceae, That is, Yamakako (Ga
The present inventors have found that the polyvalent ruler, glycerol and D-mannitol contained in R. denierae Fructus) have a vascular endothelial cell growth promoting action, and have completed the present invention.

That is, the present invention is as follows.

(1) A polyhydric alcohol having a molecular weight of about 8,000, which is obtained by ultrafiltration or dialysis of an extract of Yamaboshi, and gel filtration and ion exchange of the resulting low molecular weight fraction.

(2) A polyhydric alcohol compound having a molecular weight of about 8,000 (hereinafter referred to as polyhydric alcohol) obtained by ultrafiltration or dialysis of the extract of Yamaboshi and gel filtration and ion exchange of the resulting low molecular weight fraction. A vascular endothelial cell growth-promoting agent containing as an active ingredient.

(3) A vascular endothelial cell growth promoting agent containing glycerol as an active ingredient.

(4) A vascular endothelial cell growth promoting agent containing D-mannitol as an active ingredient. It is already known that Yamajuku or its extract has a vascular endothelial cell growth promoting action, etc., but it is not known at all that the polyhydric alcohol derivative of the present invention, glycerol and D-mannitol are active ingredients. .

The polyhydric alcohol of the present invention is obtained by ultrafiltration or dialysis of an extract of Yamaboshi as shown below,
The low molecular weight fraction obtained is obtained by gel filtration and ion exchange.

Glycerol and D-mannitol are
Similar to the polyhydric alcohol compound of the present invention, the specific example 1 from Yamakako
Alternatively, a commercially available product (Tokyo Kasei etc.) may be used.

Concrete Example 1 1. Yamajiko (20 g) was cut into small pieces, put in a clay bottle, added with distilled water (1.2 l), heated with a 600 W electric heater, and boiled until almost half the amount (about 1 hour). The extract was filtered, concentrated under reduced pressure and freeze-dried to obtain a brown powdery extract (5.3 g).

2. Next, the extracted extract (5.3 g) was dissolved in distilled water and the centrifugal separation ultrafilter (CENTRICCU) was used.
TU-10) was used to fractionate into a high molecular weight fraction of 10,000 or more and a low molecular weight fraction of 10,000 or less. The fraction liquid was concentrated under reduced pressure and then freeze-dried to obtain a brown powdery low molecular weight fraction (3.85).
g) was obtained.

3. The low molecular weight fraction (3.85 g) was dissolved in a small amount of distilled water and the activated carbon column (15 g, 2.5 cm x 1) was used.
(4.0 cm). Elution was repeated with distilled water, methanol, and then chloroform, and the eluate was fractionated into a water eluate and a methanol and chloroform eluate. The fraction liquid was concentrated under reduced pressure, freeze-dried, and the water elution part (1.16 g)
And methanol and chloroform elution part (1.11g)
Got

4. The water elution part (0.1 g) was dissolved in a small amount of distilled water, and TOYOPEARL HW-40F was used as a packing material for fractionation by a gel filtration column (1.3 cm × 60.0 cm). It was eluted with distilled water and a chromatogram was prepared using the absorbance at 220 nm as an index and fractionated into fraction 1 and fraction 2. This operation was performed for 1.8 g of water eluate, and fraction 1
(1.47 g) and Fraction 2 (0.011 g) were obtained.

5. Dissolve Fraction 1 (1.17 g) in a small amount of distilled water and use DEAE TOYOPEARL.
It was applied to a 650M column (2.5 cm x 60 cm).
As eluent, distilled water, then H ₂ O / 0.5M Na
A Cl gradient solvent was used to collect 6 ml.
A chromatogram was prepared using the absorbance at 220 nm as an index, and No. 28 to No. The 60 fractions were collected and concentrated under reduced pressure to obtain a yellowish brown viscous substance (1.03 g).

6. A yellowish brown viscous substance (0.98 g) was dissolved in a small amount of distilled water, and then a gel filtration column (2.6 cm × 70.0 cm) using Bio Gel P-4 as a packing material.
Was repeated, the polyhydric alcohol compound of the present invention (0.035 g) was obtained in the fraction 3.

SPECIFIC EXAMPLE 2 In the operation 2 of SPECIFIC EXAMPLE 1, dialysis was carried out using a cellophane tube instead of the centrifugal separation type ultrafiltration device, and SPECIFIC EXAMPLE 1
The polyhydric alcohol compound of the present invention (0.035 g) was obtained in the same manner as in.

Next, the action of the polyhydric alcohol, glycerol and D-mannitol of the present invention will be described with reference to experimental examples.

Experimental Example The vascular endothelial cells obtained from the intimal surface of bovine aorta were treated with 5.0
The cells were transferred to a 35 mm culture plate at 10 ⁴ cells / plate and cultured in 10% FBS-DME medium for 24 hours. Furthermore, ASF3 obtained by removing cultured cells from serum
After washing twice with 01 medium, it was cultured for 72 hours in ASF301 medium (Ajinomoto Co., Inc.) without serum containing the polyhydric alcohol of the present invention, glycerol or D-mannitol at a concentration of 0.1 μg / ml. After that, [ ³
[H] thymidine and [ ¹⁴ C] leucine were added, and the mixture was further cultured for 3 hours, and [ ³ H] thymidine was used as an indicator of DNA synthesis, [ ¹⁴ H] thymidine.
C] Leucine was used as an index for protein synthesis. After culturing, the cells were scraped from the culture plate in phosphate-buffered saline without calcium and magnesium using a rubber policeman, and the radiation of [ ³ H] thymidine and [ ¹⁴ C] leucine incorporated into the cells per plate was radiated. The quantity was measured. The results are shown in Table 1.

Table 1

From these results, it was confirmed that the polyhydric alcohol compound of the present invention, glycerol and D-mannitol have excellent vascular endothelial cell growth promoting action.

Next, the dose and formulation of the polyhydric alcohol, glycerol and D-mannitol of the present invention will be described.

The polyhydric alcohol, glycerol and D-mannitol of the present invention can be administered to animals and humans as they are or together with a conventional pharmaceutical carrier. The dosage form is not particularly limited and may be appropriately selected and used as needed, and examples thereof include oral agents such as tablets, capsules, granules, fine granules and powders, parenteral agents such as injections and suppositories. To be

In order to exert a desired effect as an oral preparation, it varies depending on the age, body weight and degree of disease of a patient, but is usually the weight of the polyhydric alcohol compound of the present invention, glycerol and D-mannitol in an adult. 5g ~ 12.5g 1
It seems appropriate to take it several times a day.

Oral preparations include, for example, starch, lactose, sucrose,
Mannitol, carboxymethyl cellulose, corn starch, inorganic salts and the like are used in a conventional manner. In addition to the above-mentioned excipients, a binder, a disintegrant, a surfactant, a lubricant, a fluidity promoter, a flavoring agent, a coloring agent, a fragrance and the like can be appropriately used in this type of formulation. Specific examples of each are as follows.

[Binder] Starch, dextrin, gum arabic powder, gelatin, hydroxypropyl starch,
Methyl cellulose, sodium carboxymethyl cellulose, hydroxypropyl cellulose, crystalline cellulose, ethyl cellulose, polyvinylpyrrolidone, macrogol.

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

[Surfactant] sodium lauryl sulfate,
Soy lecithin, sucrose fatty acid ester, polysorbate 80.

[Lubricant] Talc, wax, hydrogenated vegetable oil, sucrose fatty acid ester, magnesium stearate, calcium stearate, aluminum stearate, polyethylene glycol.

[Fluidity promoter] Light anhydrous silicic acid, dried aluminum hydroxide gel, synthetic aluminum silicate, magnesium silicate.

The polyhydric alcohol, glycerol and D-mannitol of the present invention can also be administered as suspensions, emulsions, syrups, and elixirs. Agents and colorants may be contained.

In order to exert a desired effect as a parenteral agent, it depends on the age, body weight and degree of disease of the patient.
5 to 200 mg per day as the weight of the polyhydric alcohol compound, glycerol or D-mannitol of the present invention in an adult.
Intravenous injection, intravenous drip infusion, subcutaneous injection, and intramuscular injection are considered appropriate.

This parenteral preparation is manufactured according to a conventional method and is generally used as a diluent in distilled water for injection, physiological saline, glucose solution, vegetable oil for injection, sesame oil, peanut oil, soybean oil, corn oil, propylene glycol, polyethylene glycol. Etc. can be used. Further, if necessary, a bactericide, a preservative, and a stabilizer may be added. Further, from the viewpoint of stability, this parenteral preparation may be filled in a vial or the like and then frozen, the water content may be removed by a usual freeze-drying technique, and a liquid preparation may be re-prepared from the freeze-dried product immediately before use. Further, an isotonicity agent, a stabilizer, a preservative, a soothing agent and the like may be added as needed.

Other parenteral agents include external preparations, coating agents such as ointments, and suppositories for rectal administration.
It is manufactured according to a conventional method.

Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited thereto.

Example 1 Corn starch 44.0 parts by weight Crystalline cellulose 40.0 parts by weight Carboxymethyl cellulose calcium 5.0 parts by weight Light anhydrous silicic acid 0.5 parts by weight Magnesium stearate 0.5 parts by weight Glycerol 10.0 parts by weight Department

According to the above formulation, the ingredients (1) to (2) were uniformly mixed and compression-molded with a tableting machine to give tablets (2 g each).

Each tablet has 200 m of glycerol.
g is contained, and 25 to 50 tablets for adults are taken in several divided doses per day.

Example 2 Crystalline cellulose 84.5 parts by weight Magnesium stearate 0.5 parts by weight Carboxymethyl cellulose calcium 5.0 parts by weight D-mannitol 10.0 parts by weight

According to the above formulation, a part of and was uniformly mixed, compression-molded, crushed, and the remaining amount of and was added and mixed, and compression-molded by a tableting machine to give one tablet.
g tablets were obtained.

D-mannitol 20 was added to one tablet.
It contains 0 mg, and 25 to 50 tablets for adults are taken in several divided doses a day.

Example 3 Crystalline cellulose 49.5 parts by weight 10% Hydroxypropyl cellulose ethanol solution 35.0 parts by weight Carboxymethyl cellulose calcium 5.0 parts by weight Magnesium stearate 0.5 parts by weight Polyhydric alcohol of the present invention 10 0.0 parts by weight

According to the above formulation, and were uniformly mixed, the mixture was kneaded by a conventional method, granulated by an extrusion granulator, dried and crushed, and then mixed, and compressed by a tableting machine. It was molded to obtain a tablet weighing 2 g.

Each tablet contains 200 mg of the polyhydric alcohol of the present invention, and is used for adults in an amount of 25 to 50 per day.
Take the tablet in several doses.

Example 4 Corn starch 84.0 parts by weight Magnesium stearate 0.5 parts by weight Carboxymethyl cellulose calcium 5.0 parts by weight Light anhydrous silicic acid 0.5 parts by weight Glycerol 10.0 parts by weight

According to the above formulation, the components (1) to (4) are uniformly mixed, compression-molded by a compression molding machine, and then crushed by a crusher,
Sieve to obtain granules.

1 g of this granule contains 100 g of glycerol.
Containing mg, 50 to 100 g of an adult is taken in several divided doses.

Example 5 Crystalline cellulose 55.0 parts by weight 10% Hydroxypropyl cellulose ethanol solution 35.0 parts by weight D-mannitol 10.0 parts by weight

According to the above-mentioned recipe, the ingredients (1) to (4) were uniformly mixed and kneaded. After granulating with an extrusion granulator, it was dried and sieved to obtain granules.

1 g of this granule contains 1 part of D-mannitol.
It contains 00 mg, and 50 to 100 g for an adult is divided into several doses and taken.

Example 6 Corn starch 89.5 parts by weight Light anhydrous silicic acid 0.5 parts by weight Polyhydric alcohol compound of the present invention 10.0 parts by weight

According to the above recipe, was mixed uniformly and 2 g was filled into capsules.

One capsule of this capsule contains 200 mg of the polyhydric alcohol of the present invention.
Take 25 to 50 capsules several times daily.

Example 7 Distilled water for injection 89.5 parts by weight Soybean oil 5.0 parts by weight Soybean phospholipid 2.5 parts by weight Glycerin 2.0 parts by weight Glycerol 1.0 part by weight

According to the above formulation, and were dissolved in, and a solution of and was added to this and emulsified to obtain an injection.

Claims (4)

[Claims] 1. A polyhydric alcohol having a molecular weight of about 8,000, which is obtained by ultrafiltration or dialysis of an extract of Yamaboshi, and gel filtration and ion exchange of the resulting low molecular weight fraction. 2. A blood vessel containing, as an active ingredient, a polyhydric alcohol having a molecular weight of about 8,000, which is obtained by ultrafiltration or dialysis of an extract of Yamaboshi extract, and gel filtration and ion exchange of the resulting low molecular weight fraction. Endothelial cell growth promoter. 3. A vascular endothelial cell growth promoting agent containing glycerol as an active ingredient. 4. A vascular endothelial cell growth promoter containing D-mannitol as an active ingredient.