JP2822286B2 - Leukocyte activity modifier - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to a leukocyte activity modifier. The leukocyte activity modifier of the present invention is useful as an agent for preventing or treating allergies, autoimmune diseases, and other diseases affected by leukocyte activity.

[0002]

2. Description of the Related Art The activity of leukocytes ranges from adhesion to blood vessel walls, migration, release of various active substances, and phagocytosis. When the activity of leukocytes is weak or strong, a disease may occur, and certain diseases that increase or control the activity have been used for prevention or treatment of the disease. For example, in the case of allergic hypersensitivity, Behcet's disease, strong inflammation, autoimmune diseases such as collagen disease, pregnancy toxemia, rejection after organ or tissue transplantation, it is necessary to suppress the activity, In the case of prophylaxis or treatment of malignant tumors, it is necessary to enhance the activity, and for this purpose, various immunosuppressants or immunostimulants have been used.

For example, to increase the activity of leukocytes,
Organic germanium, bacteria or extracts thereof (eg, OK
432, etc.) or interferon or the like has been administered, but its effect is not sufficient, and there is a possibility that strong side effects may occur when administered in large amounts over a long period of time. In addition, steroid preparations have been administered to suppress the activity of leukocytes. However, steroid therapy is
The optimal dose varies from individual to individual and differs from patient to patient, so it is difficult to determine the dose. Further, if the dose is incorrect, the adverse effect of increasing the activity of leukocytes may occur.
For this reason, there has been a strong demand for the emergence of a drug that enhances or suppresses the activity of sharp white blood cells without side effects even when administered in large amounts for a long period of time.

The present inventors have studied a method for quantitatively measuring the activity of leukocytes which has a great effect on the prevention or treatment of such various diseases. Nucleopore filter method or a more accurate silicon channel method that measures activity based on the degree of occlusion of stomata of leukocytes was developed.
upplement 171-175 (1990)]. Using this method, the degree of activation based on the degree of porosity occlusion of leukocytes was measured for various materials conventionally used as health food materials. Was found to affect the activity of leukocytes, and the present invention was completed.

[0005]

DISCLOSURE OF THE INVENTION The present invention has been made as described above, and an object of the present invention is to enhance or suppress leukocyte activity without side effects even when administered in large quantities for a long period of time. It is to provide a leukocyte activity modifier which can be used.

[0006]

That is, the present invention relates to a method for extracting a fraction obtained by water-extracting a kurozu, a concentrate thereof, a kurozu dry powder or a kurozu moromi, and dialyzing the extract to obtain an active ingredient. A leukocyte activity modifier.

[0007] The kurozu in the present invention is a food conventionally known as a health food, and is generally manufactured by the following method.

Seed Koji ↓ Rice → Washing → Soaking → Steaming → Cooling → Koji making → Koji ↓ Rice → Washing → Soaking → Steaming → Cooling → Mixing → Fermentation → Filtration → Sterilization → Bottle → Black Zu (rice vinegar)

Usually, the mixing is carried out twice a year in spring and autumn in the brewing kettle, and the fermentation and aging take a long time in the open air in the turtle, and the sugar action by koji, alcohol fermentation, acetic acid fermentation It is said that various useful components are produced.

[0010] The mash of kurozu and its juice lees includes blood sugar,
It has been reported that it has an effect of lowering blood lipid and serum cholesterol contents [Health Science 10, 85-89 (198
8), pp. 12 , 139-141 (1990)], and dried kurozu or its squeezed cake is used as a health food.

The present inventors have clarified the aqueous extract of kurozu, kurozu concentrate, kurozu dry powder and moromi (squeezed lees) with an aqueous solution containing cellulase in a fraction not exceeding 10,000 in dialysis. And more than 10,000 fractions, it was found that both fractions had an effect of changing the degree of leukocyte activity.

The kurozu moromi of the present invention can be prepared by various methods known in the art, for example, the method described in JP-A-61-58577. The fractionation is not particularly limited as long as the fractionation is performed by dialysis. An example is as follows. Kurozu moromi, especially obtained by drying it, is suspended in water, left at low temperature for several days, and centrifuged to obtain a supernatant. The amount of water used for the Sakuro moromi mash is preferably 2 to 4 volumes of water per 1 part by weight of Kurozu moromi, and the water is extracted at a low temperature of about 4 ° C. for about 1 to 4 days. The supernatant is concentrated, and the concentrated solution is subjected to dialysis fractionation at a low temperature for several days against distilled water (external solution) using a dialysis membrane capable of dialyzing the concentrated solution into a fraction having a molecular weight of 10,000 or more and a molecular weight not exceeding 10,000. This external solution is exchanged several times during dialysis, and the collected external solution is concentrated to obtain a dialysate external solution. The dialysate solution is centrifuged with a centrifuge to obtain a supernatant. As such a dialysis membrane, a seamless cellulose tube (trade name) or the like is used.

The inner or outer dialysis solution is used as it is, or further concentrated, or dried and powdered by freeze drying, spray drying, or other means. In the present invention, the dried powder of black and white can also be treated in the same manner and used as a leukocyte activity modifier.

[0014] Further, the dried black powder is obtained by freeze-drying ordinary black mash obtained by squeezing moromi, or spray-drying by adding 1 to 10 times the amount of cyclodextrin to black mash or its concentrate. Obtainable. In the present invention, the black powder is fractionated in the same manner as described above.

Further, in the present invention, when enzymatic decomposition and water extraction are carried out using an aqueous cellulase solution instead of water extraction, the nitrogen content in the extract is significantly increased as compared with the case of water extraction,
The yield of the water-extracted fraction can be improved.

[0016] These may be tablets, granules, capsules and the like by further adding excipients, lubricants and other auxiliaries which are commonly used in formulation, or may be drinks. Administration is usually oral, and the dosage varies depending on age, sex, symptoms, etc., but for adult males, 1 mg to 10 mg (0.3 g to 3 g as raw moromi powder) per day is divided into 1 to several divided doses Good to do. The active ingredient of the present invention is an ingredient which has been conventionally used for eating and drinking as a kurozu moromi, and has at least no acute toxicity.

Next, the pharmacological action of the present invention will be described. The dialysis solution obtained by the method of Example 1 described below is
Diluted 10-fold. On the other hand, saline was added to the external dialysis solution to make it equal to the salt concentration of physiological saline. Heparin was collected from 4 healthy individuals, and whole blood was diluted with blood plasma to obtain white blood cell counts.
Samples adjusted to 1200-1300 were loaded with 2 nM of the potent leukocyte chemoattractant FMLP (formyl-methionyl-leucyl-
Phenylalanine), and to 1 ml of the blood, 10 μl of the above inner or outer dialysis solution was added.
Volume 18 Suppl., Pages 171 to 175 (1990). A constant pressure was applied to a silicon filter having a pore size of 5 μm to flow the same, and the blood passage time was measured.

As a result, the obtained flow rate curve is shown in FIG.

C in the figure shows a control without the addition of FMLP, and V shows the addition of only the internal solution of dialysis. F is FM
LP indicates the addition, and V + F indicates the addition of the dialysis solution and FMLP. V + F (I), V + F (II), V + F
(III) and V + F (IV) indicate the individual receiving treatment, respectively. In the figure, an increase in the slope of the curve from F indicates that blood passes faster and the response to FMLP is suppressed, and a decrease in the slope indicates that the response to FMLP is enhanced.

As shown in the figure, when only the inner dialysis solution was added, the passage time of blood was slightly shortened as compared with the control without addition. This indicates that leukocyte activity was suppressed. The slight change is due to the fact that leukocytes are not normally activated in healthy subjects. But,
Leukocytes were activated by adding FMLP.
The reaction was suppressed or promoted by adding the dialysis solution V of the present invention to this. This indicates that the addition of Kurozu dialysis solution modifies the activation of leukocytes.

Further, FIG. 2 shows the results obtained by adding the kurozu obtained in Example 1 and the Kurozu moromi dialysis solution to the blood of a patient with toxemia of pregnancy.

As can be seen from the figure, in a pregnancy addict,
The control state without the addition of MLP shows a marked prolongation of the transit time of the blood, indicating that the leukocytes are already activated. In this bed example, when the Kurozu dialysis solution is administered, the activity of leukocytes can be suppressed,
It is found to be effective in treating toxemia of pregnancy. 1 and 2
Showed the effect of the dialysis solution, but the dialysis solution also has the same effect. FIG. 3 shows the results of administration of Kuromizu and Kurumimoromi dialysis external solution to the blood of a Behcet patient. As shown in the figure, the administration of Kurozu dialysate prevents the stomata of leukocytes, suppresses leukocyte activity, and is similar to cepharanthin (trade name) conventionally used for the treatment of Behcet's disease. Was found to be effective.

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

Example 1 After washing the glutinous rice well for 7 minutes, immersing it in water overnight, steaming and cooling it, adding yellow koji (commercially available) to it, and in a koji making room at a temperature of 32 ° C. for 70 hours. Koji making was performed. 2 to 2.5 parts by weight of the above koji, 5 to 5.5 parts by weight of steamed rice obtained in the same manner as above (as raw rice) in a brewing pot,
And 16 to 16.5 parts by weight of groundwater for brewing.
0.3 to 0.5 parts by weight were sprinkled as shaking koji, and the pots were arranged outdoors and left for 6 months for fermentation and ripening. After fermentation and aging, this was sterilized and pressed at about 80 ° C. for about 30 minutes to filter the filtrate, and the filter residue was filtered to obtain moromi. This kurozu moromi was dried to obtain kurozu moromi powder.

[0024] 2 kg of the obtained Kurozu moromi powder was suspended in 6 liters of distilled water and allowed to stand at 4 ° C for 2 days, after which the suspension was centrifuged at 7,500 rpm for 25 minutes at 4 ° C. After further filtering the centrifuged supernatant, the obtained supernatant was concentrated to about 100 ml with an evaporator. The concentrate was placed in a dialysis membrane tube (size: 20/32) and dialyzed against 5 liters of distilled water (external solution) at 4 ° C. for 2 days. This external solution is 5
The external solution was exchanged about once, and these external solutions were collected and concentrated by an evaporator to obtain a dialysate external solution. The yield of the concentrated external solution was 36.3 g. The dialysate was centrifuged at 16,000 rpm for 30 minutes, and the obtained supernatant was lyophilized. The yield was 11.0 g.
The residue (moromi lees) was 1.89 g.

These were used as they were as leukocyte activity modifiers.

[0026]

EXAMPLE 2 29 parts of a mixed powder of an adsorbent such as silicic acid anhydride, Avicel (trade name) and a usual excipient (starch, lactose, etc.) per 1 part of the dialysis solution obtained in Example 1 And compression-molded to give a tablet containing 1 mg of the outer dialysate.

[0027]

Example 3 To 1 part of the freeze-dried dialysis solution obtained in Example 1, 99 parts of an excipient was added and granulated to give granules.

[0028]

Example 4 The crocodile obtained in Example 1 was freeze-dried,
This was fractionated in the same manner as in Example 1 to obtain a leukocyte activity modifier.

[0029]

Example 5 To 5 g of Kurozu moromi powder obtained in Example 1, 10 mg of various cellulases shown in Table 1 and 150 mg of water were added.
The enzyme treatment and water extraction were performed while shaking at 50 ° C. for 5 hours. As a result, the components of the obtained extract are shown in Table 1. Using this extract, a leukocyte activity modifier was obtained in the same manner as in Example 1.

[0030]

[Table 1]

[0031]

EFFECTS OF THE INVENTION The leukocyte activity modifier obtained by the present invention suppresses or activates the activity of leukocytes when administered orally, so that diseases affecting leukocyte activity are affected.
For example, it is effective for treating toxemia of pregnancy, Pechet disease and the like. In particular, Kurozu has been used as a health food,
The component has no side effects and can be administered in large amounts over a long period of time.

[Brief description of the drawings]

FIG. 1 shows the effect of Kurozu dialysis fraction on leukocyte activity.

FIG. 2 shows the effect of Kuromizu and Kuromizuromori dialysis solution on pregnancy toxemia patients.

FIG. 3 shows the effect of Kurozu and Kurozu-Mori dialysis external solution on Behcet patients.

──────────────────────────────────────────────────続 き Continuation of the front page (56) References JP-A-62-120320 (JP, A) JP-A-3-4755 (JP, A) JP-A-58-105923 (JP, A) (58) Field (Int.Cl. ⁶ , DB name) A61K 35/66-35/78 A23L 1/29-1/308

Claims (7)

(57) [Claims] 1. A leukocyte activity-modifying agent comprising extracting a kurozu moromi with water, dialyzing the extract, and using the obtained fraction as an active ingredient. 2. A leukocyte activity-modifying agent comprising the steps of subjecting Kurozu moromi to an enzyme treatment with an aqueous cellulase solution and extracting with water, dialyzing the extract, and using the obtained fraction as an active ingredient. 3. The leukocyte activity modifier according to claim 1, wherein the fraction is a fraction not exceeding 10,000 in molecular weight. 4. The leukocyte activity modifier according to claim 1, wherein the fraction is a fraction having a molecular weight of 10,000 or more. 5. A leukocyte activity-modifying agent comprising extracting a dried powder of Kurozu with water, dialyzing the obtained fraction, and using the obtained fraction as an active ingredient. 6. A leukocyte activity-modifying agent comprising treating a dried powder of black with enzymatic treatment with an aqueous cellulase solution and extracting with water, dialyzing the extract, and using the obtained fraction as an active ingredient. 7. A leukocyte activity modifier comprising a black fraction or a concentrate thereof extracted with water and dialyzed, and the resulting fraction as an active ingredient.