WO2011093599A2

WO2011093599A2 - Yeast hydrolysate for alleviating female menopausal symptoms and food containing same

Info

Publication number: WO2011093599A2
Application number: PCT/KR2010/009631
Authority: WO
Inventors: 배송환; 서형주; 정은영
Original assignee: (주)새롬바이오
Priority date: 2010-02-01
Filing date: 2010-12-31
Publication date: 2011-08-04
Also published as: KR100989174B1; WO2011093599A3

Abstract

The present invention relates to compositions containing yeast hydrolysate for prevention and improvement of disorders associated with menopause or post menopause, and a method for manufacturing the compositions. The compositions of the present invention normalize blood hormones, improve osseous tissue and the like, and are effective for various symptoms and diseases caused by menopause such as osteoporosis, depression and the like, reduction of sex hormones and ovarian malfunction.

Description

Yeast hydrolyzate having effect on improving female menopausal symptoms and food containing same

The present invention relates to the improvement or prevention of menopausal or menopausal disorders using yeast hydrolysates.

Recent advances in medicine and economic growth have led to an increase in the average life expectancy of humans, leading to an increase in the global elderly population. In particular, the average life expectancy of Koreans in 2001 was 72.8 years for males and 80 years for females. However, in 2006, the average lifespan was rapidly increasing to 75.7 years for males and 82.4 years for females. Entered into. Therefore, research is required to improve the health and quality of life of the elderly in preparation for the growing old age unprecedentedly.

In particular, menopause, which is known to affect about 20% of all women, is a aging phenomenon that begins slowly after age 40, when the function of the ovary gradually decreases. Account for more than a third of life. The dysfunction of the gonadotrophy leading to the hypothalamus-pituitary-ovary caused by a decrease in ovarian function results in physical and mental changes such as sex hormones, lipid and cardiovascular metabolism, bone metabolism and memory. Menopause begins with menopause and postmenopausal women are at risk for many diseases due to hormonal imbalances, calcium deficiency and increased oxidative stress in the body. In other words, the incidence of diseases such as coronary artery disease, osteoporosis, Alzheimer's disease increases rapidly due to estrogen change in menopause, and in particular, the decrease in estrogen after menopause causes rapid bone loss.

Hormone therapy for postmenopausal women uses Hormone Replacement Therapy, which artificially administers estrogen, which is usually administered alone or in combination with progesterone. However, estrogen administration improves menopausal symptoms but increases the risk of breast and uterine cancer, and long-term administration increases the risk of breast cancer, venous thrombosis, high density cholesterol, elevated blood pressure, heart disease and gallbladder disease.

Therefore, while the present inventors are researching to develop a safer and more effective estrogen substitute, yeast hydrolyzate (Bioindustry, 14, 53, 1997) used as a raw material for microbial fermentation medium and health food, etc. The present invention was completed by confirming that it is effective in increasing blood estradiol and testosterone levels and improving bone metabolism and bone tissue.

It is an object of the present invention to provide a composition for the prevention and improvement of safe and effective menopausal or menopausal disorders.

It is also an object of the present invention to provide a method for preparing a composition for the prevention and improvement of safe and effective menopausal or menopausal disorders.

It is also an object of the present invention to provide a method for preventing and improving menopausal or menopausal disorders.

In order to achieve the above object, the present invention provides a composition for preventing and improving menopausal or menopausal disorders comprising yeast hydrolyzate as an active ingredient.

In addition, the present invention provides a method for producing a composition for preventing and improving menopausal or menopausal disorders comprising the step of hydrolyzing the yeast.

The present invention also provides a method for preventing, treating and ameliorating menopausal or menopausal disorders comprising administering a yeast hydrolyzate to a subject.

Yeast hydrolyzate of the present invention is effective in preventing or ameliorating menopause or menopausal symptoms caused by menopause, decreased ovarian function, and reduced sex hormones.

1 shows the effect of dosing of yeast hydrolyzate on the body weight of ovarian resected SD rats.

2 shows the effect of dosing of yeast hydrolyzate on dietary intake of ovarian resected SD rats.

3 shows the effect of dosing of yeast hydrolyzate on organ weight of ovarian resected SD rats.

4 shows the effect of dosing of yeast hydrolyzate on the hormone of ovarian ablation SD rats.

5 shows the effect of dosing of yeast hydrolyzate on bone metabolism of ovarian resected SD rats.

6 shows the effect of dosing of yeast hydrolyzate on bone tissue morphology of ovarian resected SD rats.

7 shows the effect of dosing of yeast hydrolyzate on brain tissue of ovarian resected SD rats.

The present invention provides a composition for preventing and improving menopausal or menopausal disorders comprising yeast hydrolyzate as an active ingredient.

The present invention also provides a method for preparing a composition for preventing and improving menopausal or menopausal disorders comprising the step of hydrolyzing the yeast.

The present invention also provides a method for preventing, treating and / or ameliorating menopausal or menopausal disorders comprising administering a yeast hydrolyzate to a subject (patient).

Hereinafter, the present invention will be described.

The yeast of this invention should just be a yeast used as a foodstuff, The kind is not specifically limited. For example, yeasts of the present invention include Saccharomyces cerevisiae, Saccharomyces carlsbergensis, Saccharomyces fermentati, Saccharomyces bayanus, Saccharomyces sake, Saccharomyces mandshuricus, Saccharomyces anamensis, Saccharomyces formosensis,. Saccharomyces ellipsoideus, Saccharomyces coreanus and the like can be used, preferably Saccharomyces cerevisiae .

The yeast hydrolyzate of the present invention is hydrolyzed by adding an enzyme to the yeast. In addition, the method for producing a composition for preventing and improving menopausal or menopausal disorders of the present invention includes the step of hydrolyzing by adding an enzyme to the yeast. At this time, the enzyme is preferably a protease. The protease is bromelain (Bromelain), Ficin, Papain (Papain), Flavozyme (Flavourzyme), Protamax (Neutrase) and Protease-A (Protease-A It is preferably one or more selected from the group consisting of Normalization Effect of Blood Estradiol, Testosterone and Growth Hormone in Ovarian SD Rats Treated with Yeast Bromeline Hydrolyzate When Dosing Yeast Hydrolyzate Hydrolyzed with Picin, Papain, Flavozyme, or Protamax (Recovery to SD rat levels without ovarian ablation), the enzyme is more preferably bromelain. However, a feature of the present invention is that the yeast hydrolyzate hydrolyzed by proteolytic enzymes to buffer the physiological changes caused by ovarian ablation, even if the protease is different, relieves menopausal, menopausal symptoms with yeast hydrolysates If, treatment, prevention will be within the scope of the present invention.

The menopausal or menopausal disorder refers to a disorder caused by menopause or a rapid decrease in female hormones, and the symptoms include depression, tachycardia, anxiety, hot flashes, palpitations, incontinence, difficulty urinating, urinary incontinence, recurrent urinary system inflammation, and Sweating, etc. However, if the symptoms are caused by a sharp drop in female hormones are not menopausal or menopausal disorders and are not limited to the above symptoms.

In addition, the yeast hydrolyzate of the present invention may be administered after the symptoms of menopausal or menopausal disorders, but is preferably taken in advance when a decrease in female hormone is expected or diagnosed. This is because the yeast hydrolyzate of the present invention can counteract the decrease in sex hormone which is a cause and phenomenon of menopause or menopause by promoting the secretion of sex hormones and growth hormones.

Comprising the hydrolyzate as an active ingredient, the composition for preventing and improving menopausal or menopausal disorders may be a food composition or a pharmaceutical composition.

A pharmaceutical composition for preventing and improving menopausal or menopausal disorders, including the yeast hydrolyzate of the present invention as an active ingredient, is effective in preventing and improving menopausal or menopausal disorders. In addition, the pharmaceutical composition of the present invention is effective in the prevention and improvement of symptoms caused by the rapid reduction of menopause or female hormones, such symptoms are depression, tachycardia, anxiety, hot flashes, palpitations, urinary incontinence, difficulty urinating, urinary fetus , Recurrent urinary system inflammation or sweating.

A pharmaceutical composition for preventing and improving menopausal or menopausal disorders, including the yeast hydrolyzate of the present invention as an active ingredient, can be administered orally or parenterally and can be used in the form of a general pharmaceutical preparation. Preferred pharmaceutical preparations include oral preparations such as tablets, hard or soft capsules, solutions, suspensions and the like, which can be used in the form of excipients in conventional pharmaceutically acceptable carriers such as oral preparations, Binders, disintegrants, lubricants, solubilizers, suspending agents, preservatives or extenders can be used.

The dosage of the pharmaceutical composition for preventing and improving menopausal or menopausal disorders including the yeast hydrolyzate of the present invention as an active ingredient may be determined by a specialist according to various factors such as the condition, age, sex, and complications of the patient. Generally, it can be administered at a dose of 0.1 mg to 10 g, preferably 10 mg to 1 g per kg of adult. In addition, it is intended to contain a daily dose of the pharmaceutical composition or a dose of 1/2, 1/3 or 1/4 thereof per unit dosage form, and may be administered 1 to 6 times a day. However, in the case of long-term intake for health and hygiene or health control, the amount may be below the above range, and the active ingredient may be used in an amount above the above range because there is no problem in terms of safety.

The present invention also provides a food composition for preventing and improving menopausal or menopausal disorders comprising yeast hydrolyzate as an active ingredient. The food is not limited to, but not limited to, health supplements, health functional foods, functional foods, and the like, but also includes the addition of the yeast hydrolyzate of the present invention to natural foods, processed foods, and general food materials.

Food composition for the prevention and improvement of menopausal or menopausal disorders comprising the yeast hydrolyzate of the present invention as an active ingredient, may be added as it is or used in combination with other food or food compositions, and may be appropriately used according to conventional methods Can be. The blending amount of the active ingredient can be suitably determined according to the purpose of its use (prevention, health or therapeutic treatment). In general, the health functional food of the present invention may be added in the amount of 40 to 70% by weight, preferably 50 to 60% by weight based on the raw materials in the manufacture of food or beverage. The effective dose of the yeast hydrolyzate of the food composition may be used in accordance with the effective dose of the pharmaceutical composition, but may be below the above range for long term intake for health and hygiene purposes or for health control purposes. However, since the active ingredient has no problem in terms of safety, it may be used in an amount above the above range.

There is no particular limitation on the kind of food. Food compositions comprising the yeast hydrolyzate as an active ingredient may be used in the form of oral preparations, such as tablets, hard or soft capsules, solutions, suspensions, and the like, these preparations are acceptable conventional carriers, for example In the case of oral preparations, excipients, binders, disintegrants, lubricants, solubilizers, suspending agents, preservatives or extenders may be used.

Examples of foods to which the yeast hydrolyzate can be added include meat, sausage, bread, chocolate, candy, snacks, confectionary, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, drinks, tea, Drink, alcoholic beverages and vitamin complexes, but are not limited to these types of food.

The present invention also provides a method for preventing, treating and / or ameliorating menopausal or menopausal disorders comprising administering a yeast hydrolyzate of the invention to a subject (patient). The subject may be a human or a mammal except a human.

The present invention also provides for the use of the yeast hydrolyzate of the invention for the prevention, treatment and / or amelioration of menopausal or menopausal disorders.

Hereinafter, the present invention will be described in more detail by the following examples and experimental examples. However, the following Examples and Experimental Examples are only illustrative of the contents of the present invention and the scope of the invention is not limited by the Examples and Experimental Examples.

Example 1 Materials and Methods

Preparation of Yeast Bromelain Hydrolysates

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of bromelain were added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. Thereafter, the supernatant was recovered, and peptides having a molecular weight of 10 k to 30 kDa were isolated and dried using Hydrosart membrane 10 k and 30 k (Sartorius AG, Goettingen, Germany).

Laboratory animals

Experimental animals were Nara biotech. A 10-week-old female Sprague-Dawley (SD) series rat (250 ± 5 g) supplied from (Seoul, Korea) was used. The experimental animals were kept at a laboratory temperature of 22-24 ° C and a humidity of 60 ± 5% using a breeding cage (42 × 28 cm), and the day and night cycle (12 hours light / 12 hours dark) was controlled by an automatic controller. After 1 week of prenatal breeding, ovarian resection was performed for 8 weeks.

Group settings

Ovarian excision is performed under general anesthesia with Ketamine (Keara 100 mg / kg) and 2% Xylazine (Rumpun 0.15 ml / kg), followed by conventional hair removal and preoperative sterile treatment (10% povine-iodine scrub followed by 70% alcohol wipe). Was implemented. A 1 cm incision was made in the ventral center of the experimental animal, the ovaries were ligated with a suture thread, and ovarian resection was performed on both sides (ovariectomy). After ovarian resection, each organ was relocated into the abdominal cavity, and each layer was sutured with a suture thread, and antibiotics (cafazolin 50 mg / kg) were injected into the muscle to prevent postoperative infection. One week after the procedure, only mice that recovered normally after the recovery process were selected.

The study group was divided into four groups, seven per group by random sampling (Table 1). In order to give the same stress as the experimental group, the experimental control group was the Sham control group (Sham), which performed laparotomy without ovarian ablation, the untreated negative control group (OVX), and the complex estrogen (Premina, Dalim biotech) , Seoul, Korea) was treated with a positive control (OVX + E). In the experimental group, ovarian resection was performed and the yeast hydrolyzate (Ystrogen) was mixed with 0.5% drinking water (OVX + Yst), and the positive control group (OVX + E) was mixed with 0.005% drinking water. All control and experimental groups participated in the experiment using the AIN-93 basic diet (Samyang Co., Seoul, Korea) and drinking water and diet was freely ingested.

Table 1

		Treatment	Aid
Control	Sham: sham-control	Mock-operation	No treatment
Experimental group	OVX: negative control	ovariotomy	No treatment
	OVX + E: positive control	ovariotomy	Complex Estrogen (Premina) 0.0005%
	OVX + Yst	ovariotomy	Yeast Hydrolyzate (Ystrogen) 0.5%

Statistical analysis

The experimental results were statistically analyzed using SPSS 12.0 (SPSS Inc., IL, USA), and the mean and standard error of the mean (SEM) for all measurement items were calculated. Significance between the experimental groups was confirmed by Duncan's multiple range test at p <0.05 level after ANOVA test.

Experimental Example 1 Effect of Yeast Hydrolyzate on Body Weight and Dietary Intake

<1-1> Effect of Yeast Hydrolyzate on Body Weight

Body weight gain in OVX group (2 weeks; 58.4 g, 4 weeks; 100.1 g) was significantly higher than Sham group (2 weeks; 31.1 g, 4 weeks; 56.8 g) until 4 weeks after ovarian resection (p <0.05) Weight gain was accelerated early in ovarian resection. After 4 weeks, weight gain for ovarian ablation was not statistically significant.

Combined estrogen treatment (OVX + E) significantly inhibited weight gain due to ovarian ablation throughout the 8-week observation period from the beginning of treatment (p <0.05). Could see. However, the yeast hydrolyzate (Ystrogen) 0.5% intake group was very high weight gain (p <0.05) it was confirmed that the yeast hydrolyzate has no inhibitory effect of weight gain after ovarian ablation (Fig. 1).

<1-2> Effect of Yeast Hydrolyzate on Dietary Intake

The OVX group, which had undergone ovarian resection for 4 weeks after ovarian resection, had higher dietary intake than the Sham group (p <0.05), but after 4 weeks, the intake between the two groups was statistically similar. The differences in dietary intake between the Sham and OVX groups early after ovarian resection seemed to affect the weight gain of both controls. The combined estrogen treatment group (OVX + E), which showed the most significant weight gain, was consistently consumed at the lowest dose, especially up to 5 weeks compared with the OVX group (p). <0.05). On the other hand, the yeast hydrolyzate (Ystrogen) group showed a very high dietary intake, so that the yeast hydrolyzate (Ystrogen) promoted the diet, which seems to affect the weight gain of the yeast hydrolyzate group. However, after 4 weeks, the difference between the dietary intakes of the control and experimental groups began to decrease, and there was no significant difference between the dietary intakes at week 8 (FIG. 2).

Experimental Example 2 Effect of Yeast Hydrolyzate on Organ Weight

At the end of the experiment, the animals fasted for 12 hours were sacrificed with ethyl ether, sacrificed, and the thoracic cavity was opened to collect blood from the aorta. After blood collection, 0.1 mg of pentobarbital per gram of body weight was anesthetized by intraperitoneal injection into experimental animals, and perfusion fixation was performed through the heart. First, 150 ml of saline mixed with 10 u / ml of heparin was perfused to remove blood, and immediately fixed solution was perfused. 4% paraformaldehyde-lysine-periodate was used as a fixative solution. After about 500 ml of perfusion, liver, kidney, spleen, uterus and brain were extracted.

The harvested organs were weighed and the measured organ weights were expressed relative to 100 g of body weight. The liver, kidney, and spleen weights were not statistically significant among the experimental groups. Therefore, ovarian resection did not affect the organs such as liver, kidney, spleen, and the difference according to the treatment of complex estrogen and yeast hydrolyzate (Ystrogen). Although not shown, it is determined that the treatment does not cause a change in organ weight. However, the weight of the uterus was significantly reduced due to ovarian resection (Sham; 0.27 g / 100g BW vs OVX; 0.12 g / 100g BW, p <0.05), and each uterus after complex estrogen and yeast hydrolysate (Ystrogen) treatment. The weight was slightly increased but not statistically significant (FIG. 3).

Experimental Example 3 Effect of Yeast Hydrolyzate on Hormone

At the end of the experiment, the animals fasted for 12 hours were sacrificed by anesthetizing with ethyl ether, and the thoracic cavity was opened to collect blood from the aorta. The collected blood was immediately placed in a test tube treated with heparin, and centrifuged at 4 ° C. and 3,000 × g for 10 minutes to obtain a supernatant plasma and stored at −70 ° C. until analysis.

Plasma sex hormones (estradiol and testosterone) and growth hormone levels were measured using the ELISA method, respectively, in rat estradiol Elisa kit (Uscn Life Science & Technology Co., Beijing, China), rat testosterone Elisa kit (R & D system Inc , Minneapolis, MN, USA) and rat growth hormone Elisa kit (Uscn Life Science & Technology Co.).

Estradiol

Plasma estradiol levels decreased after ovarian resection (Sham; 92.7 ng / ml vs OVX; 88.5 ng / ml), but estradiol levels were higher than in the Sham group after treatment with complex estrogen and yeast hydrolysates (97.0 ng / each, respectively). ml and 97.9 ng / ml). The yeast hydrolyzate therefore appears to promote estradiol secretion (upper left corner of FIG. 4).

Testosterone

Plasma testosterone levels decreased after ovarian resection but were higher than the Sham group when treated with complex estrogens and yeast hydrolysates, and yeast hydrolysates seem to promote testosterone secretion (upper right in FIG. 4). It has been reported that testosterone reduction is inhibited by administration of a substance having estrogen activity. As shown in FIG. 4, soldier OVX-E administered complex estrogen was found to inhibit testosterone reduction and was significantly different from that of the Sham group. There was no difference. Testosterone was significantly increased in the yeast hydrolyzate group.

Growth hormone

Plasma growth hormone levels also decreased after ovarian ablation as with sex hormones, but recovered Sham group levels after complex estrogen and yeast hydrolyzate (Ystrogen) treatment (bottom left of FIG. 4).

Experimental Example 4 Effect of Yeast Hydrolyzate on Bone Metabolism

<4-1> Bone metabolism indicator measurement

In the present invention, plasma alkaline phosphatase (ALP), calcium and osteocalcin were measured as bone formation index, and cross-linked telopeptide of type 1 collagen (CTx) was measured as an index of bone resorption. Plasma alkaline phosphatase (ALP) and calcium were measured by FUJI DRI-CHEM 3500 and osteocalcin was measured by rat osteocalcin Elisa kit (Uscn Life Science & Technology Co.). In addition, the cross-linked telopeptide of type 1 collagen (CTx), which is an index of bone resorption, was measured by ELISA using a rat cross-linked C-teminal telopeptide of type 1 collagen Elisa kit (Uscn Life Science & Technology Co.).

ALP

Bone metabolism index is a generic term for assessing bone formation ability or bone resorption capacity, which is the basis of bone metabolism dynamics, using blood or urine. ALP is an enzyme secreted by osteoblasts and is one of the most commonly used bone formation indicators in the clinic, and is known to increase in bone metabolic disorders such as osteoporosis and osteomalacia.

In the present invention, it was confirmed that ALP levels were increased due to ovarian ablation (OVX; 301.7 U / l) and that ALP levels were decreased upon administration of complex estrogen and yeast hydrolysates (280.3 U / L and 273.1 U, respectively). / L) (Figure 5 upper left).

calcium

After ovarian ablation, blood calcium decreased and calcium levels increased with the administration of complex estrogens and yeast hydrolysates. However, the amount of calcium in the blood itself was not a statistically significant level (upper right of Figure 5).

Osteocalcin

Osteocalcin is the second most abundant protein in the bone after collagen, and has a vitamin K-dependent γ-carboxyglutamic acid (Gla) residue, which is used as a calcium-binding site. Osteocalcin is formed in the osteoblasts and then deposited in the bone matrix, and a part of the newly formed osteocalcin is released into the blood, so the degree of bone formation can be determined by measuring blood concentration. As with ALP, osteocalcin is increased.

In this study, ovarian resection or complex estrogen treatment had little effect on blood osteocalcin levels. The yeast hydrolyzate treatment group had decreased blood osteocalcin compared with the ovarian resection group (OVX) and the combined estrogen treatment group, but it was not significant (OVX; 83.0 ng / ml, OVX + E; 84.1 ng / ml, OVX + Yst; 77.5 ng / ml) (Figure 5 bottom left).

CTx

CTx, one of the substances produced by the breakdown of collagen molecules by osteoclasts in bone resorption, is known as an indicator of high specificity and sensitivity to bone resorption. Ctx was increased upon ovarian ablation (Sham; 30.0 ng / ml vs. OVX; 33.2 ng / ml). Upon administration of the combined estrogen and yeast hydrolyzate, it was found to decrease to the level before ovarian ablation (OVX + E; 29.6 ng / ml, OVX + Yst; 29.5 ng / ml) (bottom right of FIG. 5).

<4-2> Bone tissue morphology measurement

Ward's triangles of the femurs of the sacrificed mice were photographed with micro-CT (Skyscan1072, SKYSCAN, Antwerpen, Belgium). The imaging tube voltage was 80 kVp, the tube current was 100 μA, and 1 mm aluminum filtration was used. The shooting time was 3,400 ms and the pixel size was 11.51 μm at 23.75 magnification. The volume of interest (VOI) was set using an acrylic tap attached to the cut surface, and the size was 3 × 3 × 5 mm3. The threshold value was set to 150 to reconstruct the bone marrow and the bone marrow from the individual images, and three-dimensional images were reconstructed and bone microstructures were obtained from the images taken with the Micro-CT using the SkyscanTM CT-analyzer software. Indicators were analyzed. Indicators analyzed for use in the present invention were bone volume (BV / TV, cancellous bone volume), trabecular number (Tb.N) and trabecular separation (Tb.Sp).

Bone shochu gap

The trabecular separation (Tb.Sp), in micrometers, representing the average distance between the bone shovels, is morphologically better at lower values. Osteoclast gap increased (OVX; 82.3 μm) during ovarian ablation, but the osteoblast gap was reduced after treatment with complex estrogen and yeast hydrolysates (77.4 μm and 77.0 μm, respectively). (Upper left corner of FIG. 6).

Trabecular bone volume fraction

The ratio of bone surface area (BNC / TV) to bone volume is the percentage of spongy bone in the total bone marrow area, including bone ossein, slightly decreased due to ovarian resection (Sham; 57.4 μm vs OVX; 54.0 μm). However, there was a tendency to increase slightly due to complex estrogen treatment, but there was no change in the treatment of yeast hydrolyzate. However, this BV / TV did not change to a statistically significant level by the treatment of ovarian ablation and complex estrogen (upper right of Figure 6).

Average number of bone weeks

The trabecular number (Tb.N) was decreased by ovarian resection. The average number of osteoblasts increased slightly after treatment with complex estrogens, but it was not significant. However, when the yeast hydrolyzate was treated, it was confirmed that the average number of osteoblasts significantly increased compared to the OVX group, which is an ovarian ablation group (p <0.05) (lower left of FIG. 6).

Taken together, the yeast hydrolyzate (Ystrogen) showed a tendency to increase the average number of bone shochu strains, reduce the gap of bone shochu strains, and improve bone tissue.

Experimental Example 5 Brain Tissue Analysis

The brains of the isolated SD rats were placed in 4% paraformaldehyde-lysine-periodate fixative and post-fixed at 4 ° C. for 1 hour. Thereafter, the resultant was washed with 0.1 M sodium phosphate buffer (PB) at 4 ° C. for 1 hour and then immersed in 20% phosphate buffered sucrose solution for 12 to 48 hours for cryoprotection. Cryoprotected brain stem tissues were made by using a cryoablation section. Coronary sections of about 40 μm in thickness were made and subjected to immunohistochemical staining using hematoxylin & eosin staining (H & E) or free floating methods. Donkey anti-rat IgG (Jackson Immunoresearch Lab Inc., PA, USA) diluted 1: 700 was added to 0.1 M PB for the purpose of improving the specificity of the secondary antibody to the primary antibody by masking IgG in the tissue. The solution (PBGT) mixed with% normal goat serum (Vectastatin) + 0.3% Triton X-100 (Sigma from Sigma Chemical Co. MO, USA) was treated for 1 hour at room temperature and washed twice with 0.1 M PB for 15 minutes. The primary antibody, ChAT (monoclonal rat anti-ChAT, Boehringer Mannheim Biochemica, Mannheim, Germany) was diluted to 2 mg in 0.1 M PB and then again diluted 1:10 in PBGT. After soaking the tissue for 12-24 hours at 4 ℃ and washed with 0.1M PB in the same manner at room temperature, and then in the biotinylated donkeyanti-rat IgG diluted 1: 100 in the secondary antibody for about an hour and a half at room temperature Reacted. After washing with the same method again, the solution was immersed in an ABC solution labeled with peroxidase for about 1 hour at room temperature. After washing again with 0.1M PB, 50 mg of 3-3'-diaminobenzidine and 250 mg of nickel chloride were reacted in a substrate solution dissolved in 50 ml of 0.1M PB for 5 minutes, and 20 μl of H ₂ O ₂ was added to give a dark blue color. After staining for about 5 minutes and washed several times with 0.1M PB, the tissue sections were put on gelatin-coated slides and dried at 4 ℃ for more than 48 hours. After dehydration and clarification of ethanol and xylene according to a conventional method, the cover glass was encapsulated and observed with an optical microscope.

The result is shown in FIG. Pathological observation of brain tissue (FIG. 7), histological cystic lesions due to ovarian resection, complex estrogen administration and yeast hydrolyzate administration were not observed (A: Sham group, B: OVX group, C: OVX group in FIG. 7). + E group, D: OVX + Yst group).

Example 2 Preparation of Yeast Hydrolyzate by Hydrolase

Example 2-1 Preparation of Yeast Ficin Hydrolyzate

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0) and 1,000 units of Ficin were added and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. After that, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kDa is isolated and dried using Hydrosart membrane 10k and 30k (Sartorius AG, Goettingen, Germany).

Example 2-2 Preparation of Yeast Papain Hydrolysate

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of papain was added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. After that, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kDa is isolated and dried using Hydrosart membrane 10k and 30k (Sartorius AG, Goettingen, Germany).

Example 2-3 Preparation of Yeast Protease-A Hydrolyzate

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of protease-A were added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. After that, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kDa is isolated and dried using Hydrosart membrane 10k and 30k (Sartorius AG, Goettingen, Germany).

Example 2-4 Preparation of Yeast Flavozyme Hydrolyzate

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of Flavozym were added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. After that, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kDa is isolated and dried using Hydrosart membrane 10k and 30k (Sartorius AG, Goettingen, Germany).

Example 2-5 Preparation of Yeast Protamax Hydrolyzate

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of Protamax were added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. After that, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kDa is isolated and dried using Hydrosart membrane 10k and 30k (Sartorius AG, Goettingen, Germany).

Example 2-6 Preparation of Yeast Nutracere Hydrolysates

Yeast ( Saccharomyces cerevisiae IFO 2346) contains 2% molasses, (NH ₄ ) ₂ SO ₄ 0.6%, MgSO ₄ 7H ₂ O 0.1%, KH ₂ PO ₄ 0.2%, K ₂ HPO ₄ 0.03%, NaCl 0.1% Incubated for 3 days at 30 ℃ in one medium. The cultured cells were suspended in 20 mM phosphate buffer (pH 7.0), and 1,000 units of Neutrase were added, and the yeast hydrolyzate was centrifuged at 30 ° C. for 4 hours. Then, the supernatant is recovered and the peptide having a molecular weight of 10 k to 30 kKDa is isolated and dried using Hydrosart membrane 10 k and 30 k (Sartorius AG, Goettingen, Germany).

The composition comprising the yeast hydrolyzate of the present invention is effective in preventing or improving menopausal or menopausal disorders caused by menopause, decreased ovarian function, decreased sex hormones, and is useful for the prevention and treatment of menopausal disorders, which are harmless to the human body. Will be used.

Claims

Yeast ( Saccharomyces cerevisiae ) A composition for preventing and improving menopausal or menopausal disorders comprising a hydrolyzate as an active ingredient.
The method of claim 1,

The composition is a food composition or a composition for preventing and improving menopausal or menopausal disorders, characterized in that the pharmaceutical composition.
The method of claim 1,

The yeast hydrolyzate is a composition for preventing and menopausal or menopause, characterized in that the hydrolysis by adding proteolytic enzymes to the yeast.
The method of claim 3, wherein

The protease is bromelain (Bromelain), Ficin, Papain (Papain), Flavozyme (Flavourzyme), Protamax (Neutrase) or Protease-A (Protease-A) Composition for the prevention and improvement of menopausal or menopausal disorders, characterized in that.
The method of claim 1,

Prevention of menopausal or menopausal disorders, characterized in that any one selected from the group consisting of hot flashes, palpitations, urinary incontinence, difficulty urination, urinary incontinence, recurrent urinary system inflammation, osteoporosis and sweating And a composition for improvement.
Method of producing a composition for the prevention and improvement of menopausal or menopausal disorders comprising the step of hydrolyzing the yeast ( Saccharomyces cerevisiae ).
The method of claim 6,

The composition is a food composition or a method for producing a composition for preventing and improving menopause or menopausal disorders, characterized in that the pharmaceutical composition.
The method of claim 6,

Method for producing a composition for preventing and improving menopausal or menopausal disorders, characterized in that the yeast is hydrolyzed with proteolytic enzymes.
The method of claim 8,

The protease is bromelain (Bromelain), Ficin, Papain (Papain), Flavozyme (Flavourzyme), Protamax (Neutrase) or Protease-A (Protease-A) Method for producing a composition for preventing and improving menopausal or menopausal disorders, characterized in that.
The method according to claim 6, wherein the menopausal or menopausal disorder is any one selected from the group consisting of hot flashes, palpitations, urinary incontinence, difficulty urinating, urinary abortion, recurrent urinary system inflammation, osteoporosis and sweating Method for producing a composition for preventing and improving menopausal or menopausal disorders.
A method for preventing and ameliorating menopausal or menopausal disorders comprising administering a yeast hydrolyzate to a subject.