JPH0146490B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Application Field] The present invention is directed to Coix lacryma-jobi L.
var.ma-yuen (Roman) Staph) seed-derived ovulation-inducing agent. [Prior art and problems to be solved by the present invention] Representative ovulation-inducing agents currently administered to humans include clomiphen and cyclohexyl, and their pharmacological effects are clinically satisfactory to some extent. It is known that However, these drugs are not necessarily sensitive and can cause abnormalities in the estrous cycle and various disorders caused by them (e.g. multiple pregnancies,
Pregnancy failure) and other side effects are known. Although these drugs have been used for more than 20 years, there are no known substitutes for them. Research on new ovulation-inducing agents is also being conducted, and it is known that there are substances in the leaves of corn, rye, wheat, etc. that induce ovulation in domestic rabbits (Masasu Suzuki Niigata Medical Society Journal, Vol. 78, p. 305, (Showa 39). On the other hand, some pharmacological effects of Coix seed extract or extract of Coix seed from which the skin and pericarp have been removed are already known, and according to Inagaki et al. (Herbal Pharmacology, p. 162, Nankodo, 1975). It is as follows. 1. Due to its diuretic effect, it is used to treat edema, beriberi, kidney and bladder stones, neuralgia, and cough. 2. It has analgesic and antispasmodic effects, so it is used for muscle spasms. 3 Used for warts, rough skin, etc. Furthermore, in Japan, it has been used as an emulsifying agent in folk medicine for a long time, but it has been revealed that protein extracted from unthreshed adlay powder promotes milk secretion (Masahiko Shigemitsu: Newsletter of the Kumamoto Regional Division of the Japanese Gynecological Society, 3). Vol., p. 191, 1944), and a substance with anticancer activity has also been isolated from Yokuinin. [Chemical and Pharmaceutical Bulletin Japan]
Bulletin, Japan) vol. 9, p. 43, (1961)]. However, nothing is known about the ovulation-inducing effect of Coix barley or Coix barley extract. The present inventors have conducted repeated research on substances that have an ovulation-inducing effect, and in the research, they have found that coix has an ovulation-inducing effect, and that the ingredients in coix that have an ovulation-inducing effect are oil from coix seeds or coix bran. It was found that it is contained in large amounts in the soluble fraction, and based on these findings, the present invention was achieved. The present invention solves the above-mentioned problems of typical conventional ovulation-inducing agents, and provides a novel ovulation-inducing agent that promotes natural ovulation without side effects and has the effect of maintaining lutein formation and increasing the number of corpus luteum. The purpose is to provide a drug. [Means for solving the problems] The present invention provides whole or part of n-
This is an ovulation-inducing agent whose active ingredient is a hexane extracted fraction. In the present invention, the whole grain of Coix seed can be used as a powder, or it can be threshed and purified by a conventional method and separated into yokuinin, bran, and hull. Coix seed is roughly divided into kernel, pericarp, and pericarp from the outside, but the bran in the present invention is
It is produced when threshed pearl barley is milled in the process of obtaining refined pearl barley by a conventional method, and refers to the part mainly composed of the pericarp.
It does not contain husk or grain. However, among the latter three types, bran has the highest extraction amount of active ingredients per unit weight;
From the viewpoint of increasing the yield, it is preferable to use bran or the whole shell containing bran. The active ingredient of the ovulation-inducing agent of the present invention is extracted with n-hexane from whole grains or a part of adlay seeds, and the active ingredient of the ovulation-inducing agent of the present invention has an LD ₅₀ value of It has low toxicity, about 0.6ml per animal. Furthermore, the configuration of the present invention will be explained in detail by the records of experiments. (Experiment 1) In Experiment 1, the water-soluble fraction and oil-soluble fraction of whole pearl barley seeds were separated, and for each fraction,
Ovulation-inducing activity was tested. First, 500 g of powdered whole pearl barley seeds was extracted with 1.5 g of n-hexane at a temperature of 15 to 20°C, and the solvent was distilled off under reduced pressure at a temperature below 40°C. Approximately 45 g of a yellow oily substance was obtained (extracted (approximately 5% by weight). This extract was designated as fraction A. Furthermore, the A fraction residue was extracted with ethanol 1.5, but the extract was
The amount was extremely small. Next, this very small amount of extraction residue was cold soaked in a 50% ethanol aqueous solution, and the soaking liquid was soaked.
When concentrated under reduced pressure at 40℃ or below, the precipitate and extract are reduced to about 2
I got g. This extract was designated as the B fraction. Furthermore, when the B fraction residue was further extracted with 1.5 ml of water, a small amount of extract was obtained, which was designated as the C fraction. The above extraction procedure is shown in FIG. In addition, when the components of each fraction A, B, and C were analyzed by a conventional method, A contained glycerides, fatty acids, and other esters, B contained polyamide, and C contained amino acids and peptides. It was something to do. Then, each fraction was tested for ovulation-inducing activity as follows. Seventy golden hamsters aged 5 to 8 weeks were equally divided into 7 groups, and 1% by weight of each of fractions A, B, and C was added to the feed and orally administered to these groups. Since the feed intake of Daleden hamsters is 19g/day per animal, the amount added was 190mg/day in both cases. and,
The animals were sacrificed after 3 weeks of administration. In addition, after administering the same fractions A, B, and C for 3 weeks to groups , , and , the mice were switched to basal feed to which these fractions were not added, and after being reared for an additional 5 weeks, they were sacrificed. The other group was sacrificed after being fed only basal feed for 3 weeks from the beginning. For groups ~, the estrous cycle was observed before sacrifice, the number of spontaneous ovulation was measured, and the ovary weight after sacrifice was measured and ovary specimens fixed with ovaries were observed. The experimental results of ovulation-inducing activity are summarized in Table 1.

【table】

[Table] From the results in Table 1, it was found that the n-hexane extract (A fraction) of whole shells of pearl barley seeds significantly promoted natural ovulation without disturbing the estrous cycle in any way. It is clear that it is useful as an active ingredient for inducing ovulation without accompanying it. In addition, organ anatomical findings indicate that this active ingredient acts centrally and has a favorable effect on lutein formation. On the other hand, it is shown that this active ingredient is hardly contained in the ethanol aqueous solution and water extract. That is, it was shown that the above-mentioned active ingredient having ovulation-inducing activity was present only in the oil-soluble fraction of the whole grain of Coix seed. (Experiment 2) In Experiment 2, the effective dose of the active ingredient in this A fraction was tested. First, the A fraction obtained in Experiment 1 was purified by the following method. That is, the A fraction was cooled stepwise with ethanol, a 50% aqueous ethanol solution, and water, the leachate fraction was removed, and further subjected to column chromatography using n-hexane. As a result, a purified n-hexane extract was obtained with a yield of 90% of the A fraction. This purified n-hexane extract was administered to 3 groups of 10 golden hamsters at a dose of 95 mg for each group.
It was dissolved in 0.2 ml of soybean oil in 3 steps in the range of ~380 mg/day, and administered by force orally once a day using a syringe. The number of ovulations and ovary weight were measured, and the correlation with the dose per body weight of one golden hamster was investigated. The results are shown in Figure 2. In FIG. 2, the solid line indicates the number of ovulations, and the broken line indicates ovarian weight. From the results shown in FIG. 2, it can be seen that the effective dose ranges from 0.76 mg/day/g body weight to 1.4 mg/day/g body weight per golden hamster. These results show that when administering to humans, it is desirable to use approximately 4.6 to 8.4 g of purified n-hexane extract per day, assuming an adult body weight of 60 kg. Furthermore, the administration period may be increased or decreased as appropriate depending on the symptoms, but one reel is 3 to 4 weeks. (Experiment 3) In Experiment 3, whole pearl barley seed shells, Yokuinin,
The amount of active ingredients extracted from each of the bran and the hull was measured, and the extraction rate from each was tested. Experiment 1: Prepare 500 g of powder of coix seed, bran, and rind, and use the same method as Experiment 1.
An extract corresponding to the A fraction was obtained. The results are shown in Table 2. Note that for whole grains, the results of Experiment 1 have been transcribed. From Table 2, it can be seen that bran has the highest extraction rate, and for efficient extraction, it is better to use bran or whole grains containing bran.

[Table] (Experiment 4) In Experiment 4, the acute toxicity of the n-hexane extracted fraction, which is the active ingredient of the ovulation-inducing agent of the present invention, was tested. The n-hexane extracted fraction prepared in the same manner as in Example 1 was administered to 4 ICR male mice (7 weeks old) per group.
The mice were orally administered using a gastric tube, and the mice were observed 1, 24 and 48 hours later, and the weight of surviving mice was measured one week later. The results are shown in Table 5, and from Table 5
The LD ₅₀ value can be seen as 0.6 ml per mouse. The average body weight of mice in this group at the time of administration was 32.3
g, the daily dose per mouse of this weight is approximately 0.6 g. This amount is about 15 to 30 times excessive compared to the effective dose of Experiment 2, which is about 23 to 42 mg/day, which is calculated based on the average mouse body weight (32.3 g). The active ingredient n
-The hexane extraction fraction was found to be extremely safe.

[Table] The ovulation-inducing agent of the present invention contains the oil-soluble fraction from adlay seeds obtained as described above or its purified product as an active ingredient, and optionally contains auxiliary ingredients in the formulation.
For example, it consists of excipients, compounding agents, diluents, and other substances, and depending on the type of adjuvant, it can be administered orally in the form of powder, fine granules, tablets, capsules, syrups, injections, etc. It can be administered locally or parenterally. Example 1 5 kg of powdered whole grains of adlay seeds was
-extracted with hexane 15 at a temperature of 20°C,
When the solvent was distilled off under reduced pressure at a temperature below 40°C, about 450 g of a yellow oily substance was obtained (extraction rate about 9% by weight). Furthermore,
This fraction was subjected to silica gel chromatography using n-hexane, and this purified n-hexane extract was administered to 10 golden hamsters aged 5 to 8 weeks in a daily feed intake of 19 g per hamster. The dose per animal was 171 mg/day (i.e. 1.3 mg/day).
During administration for 3 weeks, the animals were sacrificed and the number of natural ovulations and condition of the ovaries were examined. In addition, a group of 10 animals given only the same amount of basal feed to this administration group was used as a control group. The results are shown in Table 3, indicating that the oil-soluble fraction of whole grains of Coix seed significantly promotes natural ovulation without disturbing the estrous cycle, and also has a favorable effect on luteinization. Ta.

[Table] Example 2 Whole grains of adlay seeds were used in Example 1, but in this experimental example, 500 g of bran was used instead and extracted with 1.5 n-hexane at a temperature of 20°C. Approximately 80g of oily substance (extraction rate approximately 16)
%), which was further subjected to silica gel chromatography using n-hexane, and the solvent was distilled off under reduced pressure at 40°C to obtain about 72 g of a purified oil and fat fraction. This extract 1.5
The ovulation-inducing effect was observed by administering the drug at the rate of mg/day/g body weight and examining the number of natural ovulations in the sexual cycle and ovarian condition using golden hamsters in the same manner as in Example 1. The results are shown in Table 4. The oil-soluble fraction of the bran of adlay seeds significantly promoted natural ovulation without disturbing the estrous cycle, and also had a favorable effect on lutein formation.

〔Effect of the invention〕

The ovulation-inducing agent according to the present invention does not disturb the estrous cycle and significantly promotes natural ovulation compared to conventional ovulation-inducing agents, and has an active ingredient that induces ovulation without any hindrance. It is useful as In addition, organ anatomical findings indicate that the active ingredient acts centrally and has a favorable effect on lutein formation. Additionally, this suggests a new treatment for ovulation disorders caused by hypogonadotropic hypogonadism, and furthermore, it may be useful for the treatment of pathological conditions caused by katsu body dysfunction, for which no treatment has yet been established. It opens up a new path.

[Brief explanation of drawings]

Figure 1 shows n-hexane of whole grains of adlay seeds,
FIG. 2 is a process diagram showing an extraction procedure using ethanol, a 50% ethanol aqueous solution, and water; FIG.
- Dosage of hexane extract and number of natural ovulations (solid line)
FIG.

Claims (1)

[Scope of Claims] 1. An ovulation-inducing agent characterized in that the active ingredient is whole grains of adlay seeds or a fraction extracted with n-hexane. 2. The ovulation-inducing agent according to claim 1, wherein a part of the Coix seed is bran.