JPS61140525A - Antistress agent - Google Patents

Abstract

PURPOSE:An antistress agent that contains garlic extract as an active ingredient. CONSTITUTION:Garlics are extracted with water or an aqueous lower alcohol of 1-3 carbon atoms at room temperature, preferably lower than 40 deg.C for 10-20 hours. The resultant extract is subjected to at least one selected from dialysis, gel filtration and ultrafiltration to remove compunds of low molecular weights such as organic and inorganic compounds of less than 10,000 molecular weight. The product is subjected to anion-exchange chromatography to obtain the fraction having antistress activity. The dose is 100mg to 10g of the fraction/ day and given once or in portions. The fraction is prepared in powder, granules, tablets or capsules for oral purpose and into an injection solution for parenteral purpose.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to an anti-stress agent containing a garlic extract fraction as an active ingredient.

Garlic is a perennial herb cultivated in China, Korea, Japan, and other countries, and is generally known as a tonic and tonic, and has been used since ancient times as a stomachic, diuretic, expectorant, intestinal regulation, bactericidal, and anthelmintic. It is used.

On the other hand, it is no exaggeration to say that the word stress represents a modern disease. Stress can be said to be a burden that requires special functional tension for a given individual. Broadly speaking, the burden can be divided into (1) physical factors (overwork, lack of sleep, trauma, poisoning, hunger, etc.), (2) external factors (natural environment, social, cultural situation, family, school, workplace, etc.). (3) internal factors (conflicts in childhood, mental trauma, personality, psychological conflicts in reality, etc.). In modern society, it is impossible not to experience stress. Moreover, these stresses not only occur independently, but also occur additively or synergistically, further complicating stress symptoms.

However, there are no pharmaceuticals that are useful for the various disorders caused by these stresses (neurological diseases, digestive, circulatory, respiratory, endocrine, metabolic, pediatric, gynecological, dermatological, etc.) has not been developed. Therefore, there is currently a need for effective anti-stress agents.

As a result of repeated research on garlic components, the present inventor found that
The present invention was completed by discovering that garlic extract fraction has an anti-stress effect.

The present invention aims to provide a novel anti-stress agent, and is based on the unexpected discovery even for those skilled in the art that a garlic extract fraction has this effect.

Therefore, the anti-stress agent according to the present invention is characterized in that both components obtained by the following steps are used as active ingredients.

(b) Extracting garlic with water or a hydrous lower alcohol having 1 to 3 carbon atoms.

(0) Separating and removing low-molecular compounds by subjecting the obtained extract to at least one means selected from dialysis, gel i-filtration, and ultrafiltration.

(c) Obtaining an adsorbed fraction by subjecting the material treated by the above means to anion exchange chromatography 1--1 ・This specific extracted fraction of garlic has such specific physiological activity. This was unexpected, and needless to say, the provision of an anti-stress agent according to the present invention makes a significant contribution to modern disease control.

Garlic as used in the present invention includes members of the Liliaceae family (L 1liaceae), Allium (A 1liu
i) Allium sativum Linnaeus (A l
llium sativum L, ), for example, A. lium sativum L.
, forma pekinese Mahtno) corresponds to this.

The part that becomes the material for obtaining the target fraction is particularly preferably the bulb part (which usually contains 5 to 20 small spherical bulbs that have been split internally), which can be dried or It can be used for extraction as is.

In addition, these plants were subjected to tissue culture using conventional methods,
The culture can also be used.

Extraction of 1-1 garlic and acquisition of the desired fraction can basically be performed by any means commonly used for extraction of herbal medicines.

First, the extraction measure can be any part of the garlic plant, but the bulb part is most preferred.

The acquisition method according to the present invention consists of the following unit steps.

(b) Extraction Extraction is performed with water or a water-containing lower alcohol.

The lower alcohol to be used as an extractant has 1 to 3 carbon atoms.
(usually 1!ii alcohol), particularly preferred is ethanol.

Extraction can be carried out either under heating or at room temperature, but extraction time at room temperature is usually long, requiring several hours to several days. Preferably, the extraction is carried out at 40° C. or below for 10 to 20 hours.

Furthermore, in order to increase the extraction efficiency, it goes without saying that the target plant is preferably crushed.

(b) Separation and removal of low-molecular substances The above extract is subjected to any polymer production means such as dialysis, gel filtration, ultrafiltration or reverse osmosis after the oil alcohol is distilled off. Impurities such as low molecular weight substances and ions can be separated and removed. Preferably, it is subjected to at least one means selected from dialysis, gel filtration and ultrafiltration.

Note that the low molecular weight compound as used in the present invention refers to organic and inorganic substances having a molecular weight of 10,000 or less.

1) Dialysis Dialysis is a method of separating polymer components that utilizes the molecular sieving effect of semipermeable membranes, and animal membranes, cellophane membranes, Comidion membranes, gelatin membranes, etc. are commonly used as semipermeable membranes. ing.

Dialysis in the present invention can also be performed using any known means.

2) Gel filtration Gel filtration is a method of separating water-soluble polymer substances of different molecular weights using a polymer gel filtration agent having a three-dimensional network structure with a nearly uniform pore size. Gel filtration agents include dextran gels (e.g. Sephadex
, (P harg+acia F 1ne(
:, chemicals)), polyacrylamide gels (e.g. Bio-Gel (Bio-Ra
Any method such as dL laboratories) or agarose gel can be used.

3) Ultrafiltration Ultrafiltration is an operation in which polymer components are removed from a dispersion medium by applying a pressure difference by pressurization or suction to both sides of a membrane having pores of a certain size.

As the ultrafiltration membrane, a Comidione membrane, gelatin membrane, cellophane membrane, etc. can be used as in the case of dialysis.

(c) Purification by anion-exchange zero adsorption The above-mentioned treated solution can be concentrated under reduced pressure at low temperatures or freeze-dried, and then subjected to anion-exchange chromatography to obtain the target fraction as an adsorbed fraction. can.

Anion exchange chromatography in the present invention refers to resins that take in anions in a liquid phase and release their own anions, such as polystyrene divinylbenzene copolymers, polyamines, hydrated zirconium oxide, water This refers to a method of separating a cationic fraction and anionic fraction in a solution using any anion exchanger such as titanium oxide or cellulose, and can be carried out using conventional known means.

In addition, the adsorbed fraction obtained by subjecting to the above chromatography may contain inorganic strong acid-strong base salts, such as Na C1, K
By changing the ionic strength by treating with CI, Na2804, K2SO3, etc., it can be easily eluted and recovered.

The eluate can be concentrated under reduced pressure at a low temperature or freeze-dried to obtain the desired extracted fraction.

The anti-stress effect can be determined, for example, from an immunological perspective, and specifically, it can be evaluated using, for example, antibody production ability as an index.

When the anti-stress agent of the present invention is administered, the decrease in antibody-producing cells due to stress load is suppressed.

1λΣdan11 The anti-stress agent of the present invention is composed of the garlic extract fraction itself or mixed with appropriate excipients, binders, and diluents for preparation, and can be prepared as powder, granules, tablets, capsules, or syrup. It can be administered orally or parenterally in the form of a drug or injection. Further, other drugs may be mixed as necessary.

The dosage may be adjusted depending on age, body weight, and symptoms, but for oral administration, it is usually 100q to 1 day as an extracted fraction for adults.
It is about 0g, more preferably about 500q to 5g. A preferred embodiment of the present invention consists of a garlic extract fraction and a pharmaceutical auxiliary ingredient. Another preferred embodiment of the present invention is a unit dosage form in which the above-mentioned daily dose is divided into one or several doses.

In addition, the toxicity of the garlic extract fraction in the present invention is, for example, a dilute ethanol extract of garlic (extract content 14.5%).
, alcohol number 1.18), the LD5Q value was 30 dlυ for all oral, intraperitoneal and subcutaneous administration routes.
The JOtlrnal or To
xicolooical 3sciences, ,
9゜57 (1984)) and because garlic is commonly used as a food, it generally has low toxicity.

Song-1Jt 200g of garlic bulbs crushed into appropriate size
400% of 20% ethanol was added to the mixture, and extraction was performed at 4°C for 15 hours.

This garlic extract is heated to 40 degrees Celsius or below, and M pressure reaches approximately 80 alt! After curling, tc, MWcut off 1, 0
Transfer to 00 dialysis tube (manufactured by SPECTRUM),
Dialysis was performed at 4° C. for 15 hours against 10 liters of water. The dialyzed fluid was further dialyzed eight times in the same manner against 10 liters of water, and then freeze-dried (5.47 g in total).

4g of the obtained freeze-dried product was added to 0.05M Tris-
DEAE-TOYOP equilibrated with hydrochloric acid buffer pH 8)
EARL 650M (manufactured by Toyo Soda) 2.5φX 2
0 am) and the flow-through fraction was separated and removed using the buffer. The adsorbed fraction is 0,0 with 2M NaCj added.
Elute with 500ml of 5M Tris-HCl buffer and freeze-dry to obtain the desired fraction (hereinafter referred to as fraction A).
(Yield: 1270#1ff).

The fraction was a white powder, odorless and tasteless, easily soluble in water and almost insoluble in organic solvents.

Experiments were conducted from an immunological perspective to examine the anti-stress effect of the garlic extract fraction in the present invention. -The results are shown below along with the test method.

1) Anti-stress effect using antibody production ability as an indicator (1)
Animal Experiment After raising BALB/c male mice for a certain period of time, 5-week-old mice that appeared to be healthy were used in this experiment as a group of 8 mice.

(2) Experimental method The test solution at the following concentration was orally administered to the above mice every day (
The experiment was continued for a total of 11 days until the end of the experiment), and 4 days after the initial administration, the animals were placed in a stress cage and subjected to restraint stress (12 hours, 71 days) for 4 days. After that, 2.5
X10 /d of sheep red blood cells (SRBC) 0.27 were administered through the tail vein, and after administration of 5RBG, the mice at the injection port 4 were bled to death, the spleen was removed, and the PFC (Plat
quel”orming Ce1l) assay method (3
C1enCe.

140.405 (1963)), the number of antibody-producing cells was measured.

(Dosage of test solution) (a) Physiological saline (control) 10ae/Nf (b)
Fraction A20IIIy/! g→2jv/M
1→10Id/Kg (c) Fraction A 50j13F/Ks
-+5QF/d→10d/N9 The statistical processing of the results was performed using Stenipunt (S tu
dent's) was performed using the -test.

(3) Experimental Results The increase in the number of antibody-producing cells due to administration of Ryobu A was as shown in the figure.

It was revealed that the decrease in antibody-producing cells due to stress load was controlled by administration of fraction A.

In addition, in the process of obtaining both fractions A, similar pharmacological experiments were conducted on the fraction that was not adsorbed to anion exchange chromatography (pass-through fraction), but no anti-stress effect was observed.

Fraction 1050g, lactose 14009 and starch 450
g and hydroxypropyl cellulose (
Wet granulation was performed using an aqueous solution (containing 100 g) according to a conventional method. After drying, the particles were sized to make sachets of 2 g each.

11■-1 Fraction 10507, lactose 1200g, crystalline cellulose 57
0 g and 15 g of magnesium stearate were uniformly mixed and made into slap tablets using a slap tablet machine. This was sized, and 150 g of starch and 159 g of magnesium stearate were mixed therewith and tableted at 500 q per tablet.

Example 3 In a 2000 m beaker, 500 d of purified fishing rod was poured, and 238 g of A was dissolved in both parts. After adding 1000ml of simple syrup to this and mixing, 17ml of an ethanol solution in which 0.24g of butylparaben was dissolved! was added and further mixed. Purified water was added to this to bring the total volume to 1700 d, and then filtered with a membrane filter. Add this furnace liquid to 1
After filling each bottle for 30 minutes, the mixture was heat sterilized to obtain a syrup.

Example 4 Take 1000 d of purified fishing rod in a 2000 m beaker, dissolve and mix both A1055F, and then add 200 d of simple syrup.
added. After dissolving 0.21 g of butylparaben in this, purified water was added to make the total amount 1500++d. After filtering this with a membrane filter, according to the usual method,
Each bottle was filled with 30 m of water and heat sterilized to make a so-called drink.

[Brief explanation of the drawing]

The figure is a graph showing the effect of Ryobu A on increasing the number of antibody-producing cells. Applicant's representative Kiyoka Inomata:

Claims (1)

[Scope of Claims] 1. An anti-stress agent characterized by containing a fraction obtained by the following process as an active ingredient. (b) Extracting garlic with water or a hydrous lower alcohol having 1 to 3 carbon atoms. (b) Subjecting the obtained extract to at least one means selected from dialysis, gel filtration, and ultrafiltration to separate and remove low-molecular compounds. (c) Obtaining an adsorbed fraction by subjecting the product treated by the above means to anion exchange chromatography.