JP2613714B2 - Human platelet aggregation inhibitor - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001] The present invention relates to a human platelet aggregation inhibitor, and more particularly, to a highly non-toxic, highly safe human platelet aggregation comprising, as an active ingredient, a squeezed component of a green leaf of a gramineous plant, particularly a wheat plant. Related to inhibitors.

The present inventors have paid attention to the physiological action of squeezed juice or its dry powder on young leaves of a grass plant, especially a barley plant before maturity, and have been studying it for many years. Prevention or cure effect of hardening (JP-B-46-385)
No. 48), an anticancer effect (JP-B-59-36890).
), Blood pressure lowering action (JP-B-59-36888).
JP-A No. 59-39410 and JP-B-59-39410), blood glucose lowering action (see JP-A-62-226927), antiviral action (see JP-A-3-169823),
The antioxidant activity (see Japanese Patent Application No. 2-220398) and the like have been found, and some of them have been successfully isolated and confirmed as the active substance.

[0003] In the course of the above series of research, the present inventors have
This time, it was found that the squeezed component of green leaves of grasses, particularly barleys, has an inhibitory effect on human platelet aggregation, and the present invention has been completed.

[0004] Thus, according to the present invention, there is provided a human platelet aggregation inhibitor comprising as an active ingredient a squeezed component of a green leaf of a gramineous plant.

Hereinafter, the human platelet aggregation inhibitor of the present invention will be described in more detail.

[0006] As a grass plant which is a raw material of the active ingredient of the inhibitor of the present invention, a barley plant is preferable. Examples of the barley plant include barley, wheat, naked barley, oats, oats,
Corn, millet, Italian diegrass, and the like, among which barley, wheat, and naked barley are particularly suitable.

According to the present invention, among these wheat plants, fresh stems and / or leaves of young plants harvested before the maturity stage (the stems and / or leaves are collectively referred to herein as "green leaves") Is particularly suitable.

[0008] First, the green leaves of grasses, especially wheat,
The juice is squeezed by a mechanical crushing means such as a mixer or a dicer under conditions that do not cause undue heat denaturation, and the squeezed liquid (hereinafter, referred to as “the squeezed liquid” is removed by removing coarse solids by means such as sieving or filtration. Green soup). The green juice thus obtained can be used as it is as the juice component of the present invention. In general, it is preferable that the green leaf be sterilized with a bactericide such as sodium hypochlorite before the above-mentioned juice and then subjected to the juice processing.

[0009] The green juice obtained as described above is preferably further centrifuged to collect a supernatant, and if necessary, sterilized and filtered with a sterile filter or the like. Can also be used as the juice component of the present invention.

Further, the green juice or supernatant component obtained as described above is neutralized to a pH of about 5 to 9 in accordance with, for example, the method described in JP-B-46-38548, and substantially subjected to spray drying, freeze drying and the like. The powder may be dried and powdered by an appropriate means that does not cause thermal denaturation to obtain a green juice powder or a supernatant powder. These green juice powders or supernatant powders can also be used as the juice component of the present invention.

Further, a liquid obtained by re-dissolving the green juice powder or the supernatant powder in water or a liquid obtained by removing insoluble components from the re-dissolved aqueous solution can be used as the juice component of the present invention. A liquid obtained by extracting a green juice powder or a supernatant powder with an aqueous alcohol, for example, 50% aqueous methanol, or a powdered product obtained by removing the aqueous alcohol therefrom can also be used as the juice component of the present invention.

The liquid from which the solid content has been removed from the green juice or the liquid from which the green juice powder has been redissolved in water to remove the solid content, for example, the above-mentioned supernatant component may be subjected to reverse-phase chromatography, ion-exchange chromatography, A fraction having human platelet aggregation inhibitory activity can also be collected by subjecting it to gel filtration or the like. Reversed phase chromatography is, for example, Delta-Pak3
Using a 00Å 15 μm C ₁₈ column (Waters),
The active fraction can be eluted using acetonitrile containing 0.01 to 0.5% trifluoroacetic acid and 0.01 to 0.5% trifluoroacetic acid as an eluent. Examples of ion exchange chromatography include, for example, Protein-Pak DEAE-5P
W (Waters) containing 1-100 mM Tris-HCl buffer pH 8.7 and 0.1-1 M NaCl 2
An active fraction can be obtained by performing chromatography with a linear gradient of 0 to 100% with 0 mM Tris HCl buffer. Further, gel filtration is performed, for example, using Protein-
This can be performed using Pak 300SW (manufactured by Waters). The green juice or supernatant component obtained as described above can be orally administered as it is or by mixing it with, for example, milk, skim milk, other colloidal protein-containing substances, sweeteners, etc. It is preferable to use the green juice powder or the supernatant powder, which has better stability, or a re-dissolved product or extract thereof.

The "squeezed component of green leaves of grasses" used as an active ingredient in the present invention includes not only green juice obtained as described above from green leaves of grasses, especially barley plants, but also green juice. As described above, the term is used in such a manner that all treatment components or fractions having a human platelet aggregation inhibitory effect obtained by further treatment are included.
The human platelet aggregation inhibitor of the present invention can be substantially composed of only the squeezed components described above, but is generally suitable for administration by blending various pharmaceutically acceptable additives. It is preferable to formulate the composition in a dosage form.

Examples of the additives that can be used include additives for freeze-drying or spray-drying, as well as any dispensing additives usually used for formulating a desired dosage form. Specifically, for example, ascorbic acid, biotin, calcium pantothenate, carotene, choline chloride, magnesium oxide, niacin, pyridoxine chloride, lipoflavin, sodium pantothenate, thiamine hydrochloride, tocopherol, vitamin A,
Nutrients such as vitamin B ₁₂ and vitamin D ₂ ; sodium metaphosphate, sodium phosphate (first, second, third)
Salts), sodium pyrophosphate, sodium tripolynate, etc .; preservatives, such as calcium sorbate, benzoic acid, methyl parahydroxybenzoate, sodium benzoate; arabic gum, tragacanth, sodium alginate, methylcellulose, carboxymethylcellulose ,
Carriers or diluents such as calcium alginate, aluminum silicate, calcium silicate, mannite, sorbitol, lactose, fructose, soluble starch, amino acids, glucose, sugar, honey, sucrose and fatty acid esters can be mentioned.

The human platelet aggregation inhibitor of the present invention can be administered orally or parenterally, and can be formulated into any dosage form suitable for each administration route.
Dosage forms suitable for oral administration such as granules, pellets or tablets, coated tablets, capsules, troches, solutions, syrups, etc .; non-injectable forms such as injections, drops, suppositories, eye drops, nasal drops, sprays, etc. The dosage form can be made suitable for oral administration.

Further, external preparations such as ointments, creams, tinctures, and patches can be prepared.

The dose of the human platelet aggregation inhibitor of the present invention depends on the weight, gender, age, weight,
Although it can be changed in a wide range according to the judgment of a physician or the like, as a rough guide, it is generally about 30 to about 1000 mg / day, preferably about 100 mg / day as an active ingredient per adult.
A range of about to about 500 mg / day can be exemplified. The above dose can be administered once or several times a day. However, when administered orally, the human platelet aggregation inhibitor of the present invention is substantially non-toxic and has no side effects as described below, so that it can be administered in large amounts beyond the above range. The acute toxicity LD ₅₀ of the juice component of green leaves of grasses, which is an active ingredient of the human platelet aggregation inhibitor of the present invention, for example, green juice powder from young leaves of barley before maturation is 1
It is practically non-toxic at 2,000 mg / kg (oral, mouse), and substantially no toxicity and side effects are observed from the results of the subacute toxicity test of 1000 mg / kg continuous administration (oral, mouse).

Therefore, in view of its great pharmacological effect and no toxicity and side effects, the squeezed component of the green leaf of the gramineous plant of the present invention is practically non-toxic to human platelet aggregation. It was found to be a very unique human platelet aggregation inhibitor which has both the ability to be administered in large amounts and the ability to administer it.

Hereinafter, the present invention will be described more specifically with reference to examples.

[0020]

EXAMPLES Example 1: Preparation of Barley Juvenile Green Juice Powder After washing 1 kg of barley young leaves before maturation, they were ground with a crusher and squeezed using a juicing machine to extract barley young leaves green juice. 1
1 was obtained. Add calcium hydroxide to this green juice and adjust the pH
Was adjusted to 7.0 and freeze-dried by a conventional method to obtain 40 g of green juice powder (hereinafter referred to as BLE powder) of young barley leaves.

Example 2: Measurement of human platelet aggregation inhibitory activity Preparation of sample solution : 5 g of BLE powder prepared in Example 1 was dissolved in 10 ml of distilled water, and sterilized with a 0.22 μm sterile filter (Wa).
ters) to obtain a sample solution.

Measurement method : 9 ml of blood of a healthy person was collected into a polypropylene tube containing 1 ml of a 3.13% aqueous solution of sodium citrate, centrifuged at 2300 rpm for 4 minutes, and the cloudy supernatant was separated. Platelet-rich plasma (hereinafter,
PRP), while buffer was poured into the remaining lower layer to make the total volume 10 ml, and then 1800 at 2800 rpm.
After centrifugation for 0 minutes, the supernatant was separated to obtain platelet poor plasma (PPP).

The Paybon dual-channel aggregometer is defined as P
The RP was adjusted so as to have an optical density of 0% by RP and 100% by PPP, and the cell for measurement contained P1 containing 1 mM CaCl _2.
After adding 40 μl of RP and allowing to stand for 1 minute, the sample solution in the amount shown in Table 1 below was added, and after culturing at 37 ° C. for 2 minutes, a platelet aggregation promoting agent was added at the concentration shown in Table 1, and the optical density was read. This optical density corresponds to the human platelet aggregation inhibition rate.
Table 1 shows the results.

[0024]

[Table 1] As shown in Table 1 above, BLE powder shows a remarkable inhibitory effect on human platelet aggregation.

Next, a formulation example using BLE powder produced in the same manner as in Example 1 will be described.

Example A: Tablets BLE powder 40.0 mg Lactose 82.4 mg Crystalline cellulose 70.0 mg Talc 5.0 mg Binder (carboxymethylcellulose) 2.0 mg Magnesium stearate 0.6 mg 20.0 mg BLE powder, lactose, After the crystalline cellulose, talc and binder are uniformly mixed and granulated, magnesium stearate is added to form a tablet of 200 mg per tablet.

Example B: Enteric coated tablet The tablet obtained in Example A was subjected to an enteric coating having the following formulation to produce one 430 mg enteric coated tablet.

Methacrylic acid / ethyl acrylate copolymer 10.8% Polyethylene glycol 6000 1.6% Surfactant (Tween 80) 1.1% Talc 7.2% Purified water 79.3% 100% Example C: Granules BLE powder 1.0 mg Lactose 700.0 mg Starch 289.0 mg Gelatin 10.0 mg 100.0 mg BLE powder, lactose and starch are uniformly mixed, a small amount of water is added, and the mixture is further kneaded and granulated. And dried (columnar granules 0.8 mm in particle size). Example D: Capsules Granules are prepared according to the following formulation, coated with an enteric coating, and filled into capsules.

Granules (columnar granules with a diameter of 0.8 mm) BLE powder 1.4 mg Lactose 140.0 mg Starch 56.6 mg Gelatin 2.0 mg 20.0 mg Enteric coating (coating amount 430 mg / g)
Granules) The formulation is as in Example B above.

Capsule filling Gelatin capsule No. 2 is filled with 200 mg.

Example E: Lozenge BLE powder 1.0 mg White sugar 920.0 mg Arabic gum 79.0 mg Purified water appropriate amount 100.0 mg BLE powder and white sugar are uniformly mixed, and Arabic gum is dissolved in a small amount of purified water. In addition, the mixture was kneaded, granulated, dried, and tableted to give a troche.

Example F: Suppository BLE powder 1.0 g Polyoxyethylene lauryl ether (21E.O.) 30.0 g Polyoxyethylene sorbitan monostearate (6E.O.) 100.0 g Lipophilic glyceryl monostearate 16 6.0 g polyethylene glycol 400 6.0 g polyethylene glycol 4000 appropriate amount Polyoxyethylene lauryl ether, polyoxyethylene sorbitan monostearate, lipophilic glyceryl monostearate, polyethylene glycol 400 and polyethylene glycol 4000 were heated to 60 ° C. and dissolved. Thereafter, the mixture was cooled to 45 ° C., BLE powder was added thereto, and the mixture was uniformly mixed, and then molded into a suppository of 2 g with a suppository molding machine.

Example G: Microcapsule Lactose 740.0 g BLE powder 20.0 g Eudragit RS 50.0 g Magnesium stearate 10.0 g 100.0 g After lactose is put into a vacuum mixing and drying coating device, it is rotated and mixed. After heating to about 50 ° C., the inside of the tank was evacuated by a vacuum pump, and coated with an aqueous solution of BLE powder. After coating, Eudragit RS
Was coated under a vacuum in the same manner using a methylene chloride solution (containing 1% of magnesium stearate based on the total weight of the product) to obtain microcapsules of BLE powder.

 ──────────────────────────────────────────────────続 き Continuation of the front page (56) References JP-A-62-226927 (JP, A) JP-A-61-194017 (JP, A) JP-A-3-27246 (JP, A) 38548 (JP, B2) JP-B 46-39548 (JP, B2)

Claims (2)

(57) [Claims] 1. A barley plant selected from barley, wheat and naked barley
A human platelet aggregation inhibitor comprising, as an active ingredient, a green leaf squeezed component before the maturation period of a product. 2. The human platelet aggregation inhibitor according to claim 1, wherein the barley plant is barley .