Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K38/00—Medicinal preparations containing peptides
  • A61K38/01—Hydrolysed proteins; Derivatives thereof
  • A61K38/012—Hydrolysed proteins; Derivatives thereof from animals
  • A61K38/018—Hydrolysed proteins; Derivatives thereof from animals from milk
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
  • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
  • A61K35/20—Milk; Whey; Colostrum
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P19/00—Drugs for skeletal disorders
  • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P19/00—Drugs for skeletal disorders
  • A61P19/08—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease
  • A61P19/10—Drugs for skeletal disorders for bone diseases, e.g. rachitism, Paget's disease for osteoporosis
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00

Definitions

• the present invention has a basic protein fraction and / or a basic protein fraction degradation product and a whey protein hydrolyzate as active ingredients, is excellent in bone strengthening action, proliferates osteoblasts, and
• the present invention relates to a bone strengthening agent that has an action of suppressing differentiation and bone resorption by the cells, is effective in the prevention and treatment of various bone diseases such as osteoporosis, fracture treatment, rheumatism, and arthritis, and is excellent in stability and safety.
• the present invention further relates to a bone-strengthening food / beverage, a bone-strengthening nutrition composition, a bone-strengthening feed, or a bone-strengthening pharmaceutical, which further contains the bone-strengthening agent.
• Bone quality refers to bone microstructure, turnover, micro fracture, and calcification.
• osteoclasts As a method for preventing various bone diseases such as osteoporosis, fractures, and back pain, it is conceivable to suppress bone resorption by osteoclasts. Bone repetitively undergoes balanced resorption and formation (remodeling), but due to post-menopausal hormone balance changes, etc., bone resorption exceeds bone formation, which is a variety of bone diseases such as osteoporosis, fractures, and back pain Cause. Therefore, it is possible to strengthen the bone as a result by suppressing bone resorption by osteoclasts and keeping the bone strength constant.
• calcium salts such as calcium carbonate, calcium phosphate, and calcium lactate
• natural calcium preparations such as whey calcium, beef bone meal, eggshell, etc.
• substances having a calcium absorption promoting effect such as casein phosphopeptides and oligosaccharides to pharmaceuticals, foods and drinks, and feeds.
• the absorption rate of calcium is 50% or less, and more than half of the calcium is not absorbed and discharged outside the body.
• calcium absorbed in the body also does not necessarily show an improvement in bone metabolism or a bone strengthening action because its affinity to bone varies depending on its form and the types of other nutritional components taken at the same time.
• milk-derived basic protein fraction examples include a milk-derived basic protein fraction (hereinafter referred to as “milk-derived basic protein fraction”) as described in Patent Document 1, and a milk-derived basic protein.
• a basic protein fraction degradation product (hereinafter referred to as “milk-derived basic protein fraction degradation product”) obtained by degrading a protein fraction with a proteolytic enzyme has been reported.
• An object of the present invention is to provide a bone strengthening agent that is highly safe, can strengthen bones, and is useful for the prevention and treatment of various bone diseases such as osteoporosis, fractures, rheumatism, and arthritis. It is another object of the present invention to provide a bone-strengthening food / beverage, a bone-strengthening nutrition composition, a bone-strengthening feed, or a bone-strengthening pharmaceutical that contains a bone-strengthening agent.
• a bone strengthening agent comprising a milk-derived basic protein fraction and / or a milk-derived basic protein fraction degradation product and a whey protein hydrolyzate as active ingredients.
• the milk-derived basic protein fraction degradation product decomposes the milk-derived basic protein fraction using at least one proteolytic enzyme selected from the group consisting of pepsin, trypsin, chymotrypsin and pancreatin.
• the bone strengthening agent according to (1) (3) The bone strengthening agent according to (1) or (2), wherein the milk-derived basic protein fraction is a fraction containing 15% by weight or more of basic amino acids in its amino acid composition.
• the milk-derived basic protein fraction is prepared by bringing milk or a milk-derived material into contact with a cation exchange resin to adsorb basic protein, and the fraction adsorbed on the resin has a salt concentration of 0.1 M to 1 M
• the bone strengthening agent according to (1) or (2) which is a fraction obtained by elution with a 0 M eluate.
• the bone strengthening agent according to (1), wherein the whey protein hydrolyzate has a degradation rate of 25% or more.
• the bone strengthening agent according to (1), wherein the whey protein hydrolyzate has the following characteristics.
• (A) The molecular weight is 10 kDa or less, and the main peak is 200 Da to 3 kDa.
• APL average peptide chain length
• C The free amino acid content is 20% or less.
• D Antigenicity is 1 / 10,000 or less of that of ⁇ -lactoglobulin.
• the whey protein hydrolyzate decomposes the whey protein by heat denaturation using a heat-resistant protein hydrolase at pH 6 to 10 and 50 to 70 ° C., and heats to inactivate the enzyme.
• the bone strengthening agent according to (1) which is obtained.
• the whey protein hydrolyzate hydrolyzes whey protein using a protein hydrolase at pH 6 to 10 and 20 to 55 ° C., and raises the temperature to 50 to 70 ° C.
• a bone strengthening agent as described in.
• a bone-strengthening food / beverage, a bone-strengthening nutrition composition, a bone-strengthening feed or a bone-strengthening pharmaceutical comprising the bone-strengthening agent according to any one of (1) to (8).
• a bone strengthening method by simultaneously ingesting a milk-derived basic protein fraction and / or a milk-derived basic protein fraction degradation product and a whey protein hydrolyzate.
• the bone strengthening agent of the present invention is useful for the prevention and treatment of various bone diseases such as osteoporosis, fractures, rheumatism and arthritis.
• the feature of the present invention is that a milk-derived basic protein fraction and / or a milk-derived basic protein fraction degradation product and a whey protein hydrolyzate are used as active ingredients.
• the milk-derived basic protein fraction used in the present invention is obtained from mammalian milk such as cow's milk, human milk, goat milk, sheep milk, and the milk-derived basic protein fraction degradation product used in the present invention is It can be obtained by reacting a milk-derived basic protein fraction with a proteolytic enzyme.
• This milk-derived basic protein fraction has the following properties. 1) According to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), it consists of several kinds of proteins having a molecular weight ranging from 3,000 to 80,000. 2) 95% by weight or more is protein and contains a small amount of other fat and ash. 3) Protein mainly consists of lactoferrin and lactoperoxidase. 4) The amino acid composition of the protein contains 15% by weight or more of basic amino acids such as lysine, histidine and arginine.
• SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis
• Such a milk-derived basic protein fraction is obtained by, for example, bringing a milk raw material such as skim milk or whey into contact with a cation exchange resin to adsorb the basic protein, and the basic protein fraction adsorbed on the resin is obtained. Elution is carried out with an eluate having a salt concentration of 0.1 M to 1 M, and this elution fraction is collected, desalted and concentrated by a reverse osmosis (RO) membrane or electrodialysis (ED) method, and dried if necessary. Can be obtained.
• RO reverse osmosis
• ED electrodialysis
• milk or a milk-derived raw material is contacted with a cation exchanger to adsorb a basic protein, and then adsorbed to the cation exchanger.
• a method of obtaining a basic protein fraction by elution with an eluent exceeding pH 5 and having an ionic strength of 0.5 Japanese Patent Laid-Open No. 5-202098
• a method using alginate gel Japanese Patent Laid-Open No. 61-246198.
• No. 1 a method of obtaining from whey using inorganic porous particles (Japanese Patent Laid-Open No.
• a milk-derived basic protein fraction obtained by such a method can be used in the present invention.
• the milk-derived basic protein fraction degradation product has the same amino acid composition as the milk-derived basic protein fraction.
• pepsin is added to the milk-derived basic protein fraction obtained by the above method.
• a peptide composition having an average molecular weight of 4,000 or less can be obtained by allowing a proteolytic enzyme such as trypsin or chymotrypsin to act, and further allowing a proteolytic enzyme such as pancreatin to act as necessary.
• the whey protein hydrolyzate used in the present invention can be obtained, for example, by the method described in JP-A-4-112753.
• the whey protein is adjusted to pH 6-10, 50-70 ° C., heat-stable protein hydrolase is added thereto, the enzyme is decomposed while thermally denatured, and this is heated to inactivate the enzyme. It is done.
• the whey protein Prior to the above enzymatic degradation, the whey protein is enzymatically degraded with a protein hydrolase at pH 6 to 10 and 20 to 55 ° C., and immediately after the enzymatic degradation under the above conditions without cooling, the yield is further increased. Can be increased.
• the whey protein hydrolyzate prepared as described above is subjected to ultrafiltration (UF) membrane with a molecular weight cut off of 1 kDa to 20 kDa, preferably 2 kDa to 10 kDa and / or a molecular weight cut off of 100 Da to 500 Da, preferably 150 Da to It is also possible to concentrate by a method selected from a 300 Da microfiltration (MF) membrane.
• UF ultrafiltration
• MF microfiltration
• the above-mentioned solution is adjusted to pH 6 to 10.
• the special pH should be adjusted.
• the pH is adjusted to 6 to 10 using an acid solution such as hydrochloric acid, citric acid and lactic acid or an alkaline solution such as caustic soda, calcium hydroxide and sodium phosphate. Heating is performed at 50 to 70 ° C., but it is preferable from the viewpoint of yield that the heat-resistant protein hydrolase is added before heating rather than being added at this temperature.
• thermostable protein hydrolase has a residual activity of about 10% or more when heated at 80 ° C. for 30 minutes.
• the reaction is preferably performed for about 30 minutes to 10 hours.
• the reaction solution is heated to deactivate the enzyme.
• the enzyme can be deactivated by heating the reaction solution at 100 ° C. or higher for 10 seconds or longer.
• the reaction solution is centrifuged to collect the supernatant, and the supernatant is dried to obtain a powder product.
• the precipitate generated upon centrifugation is less allergen-reduced than the supernatant, so it is preferable to remove it.
• the reaction solution may be dried and used as it is.
• the APL (average peptide chain length) of the obtained whey protein hydrolyzate can be measured by a method such as the TNBS (2, 4, 6-trinitrobenzenesulfonic acid) method.
• the molecular weight distribution of the whey protein hydrolyzate can be measured by a method such as the Highperformance sizeexclusion chromatography (HPSEC) method, and the free amino acid content is extracted with 75% ethanol etc. Etc. can be measured.
• the degradation rate of the whey protein hydrolyzate can be measured by an orthophthalaldehyde (OPA) method in which a free amino group is modified and measured.
• the whey protein in the present invention refers to whey prepared from mammalian milk such as cow's milk, human milk, goat milk, and sheep milk, its aggregate, powder, or purified protein. When this is enzymatically reacted, it is in the form of an aqueous solution. Used in.
• the milk-derived basic protein fraction and / or milk-derived basic protein fraction hydrolyzate and whey protein hydrolyzate of the present invention may be used together as a bone strengthening agent when ingested together, Accordingly, it can be formulated into powders, granules, tablets, capsules, drinks and the like according to conventional methods.
• milk-derived basic protein fraction and / or the milk-derived basic protein fraction degradation product and the whey protein hydrolyzate there is no particular limitation on the method of blending the milk-derived basic protein fraction and / or the milk-derived basic protein fraction degradation product and the whey protein hydrolyzate.
• milk-derived basic protein fraction and / or milk-derived basic protein fraction degradation product and whey protein hydrolyzate may be mixed uniformly, and ultradisperser, TK homomixer, etc. are used. It is also possible to stir and mix.
• the solution of the said composition can be used by concentrating with an RO membrane etc.
• the sterilization process normally used for manufacture of a pharmaceutical, food-drinks, and feed can be performed, and if it is a powder form, dry heat sterilization is also possible.
• various forms of pharmaceuticals such as liquids, gels, powders, granules and the like containing the milk-derived basic protein fraction and / or milk-derived basic protein fraction degradation product and whey protein hydrolyzate of the present invention, It is possible to produce foods and drinks and feeds. Furthermore, after formulating them, it is also possible to blend them into foods and drinks such as nutrients, yogurt, milk drinks, wafers, nutritional compositions, feeds and pharmaceuticals. .
• the bone-strengthening food and drink, the bone-strengthening nutrition composition, the bone-strengthening feed and the bone-strengthening pharmaceutical of the present invention are the milk-derived basic protein fraction and / or the milk-derived basic protein fraction degradation product and whey. Besides containing only protein hydrolyzate, it can contain stabilizers, saccharides, lipids, flavors, vitamins, minerals, flavonoids, polyphenols, and other raw materials usually contained in foods, feeds and medicines. .
• a bone-strengthening food or drink a bone-strengthening nutrition composition, a bone-strengthening feed or a bone-strengthening pharmaceutical can be used as a raw material, and the raw materials usually contained in other food and drink can be prepared. Is possible.
• the amount is not particularly limited, but it is preferable to take 1 mg or more of a milk-derived basic protein fraction and / or a milk-derived basic protein fraction degradation product and 2 mg or more of a whey protein hydrolyzate per day for each adult.
• whey protein hydrolyzate Is contained in an amount of 0.001 to 10% (w / w), preferably 0.1 to 5% (w / w) based on the total mass.
• the bone strengthening agent of the present invention can be formulated into an arbitrary form by adding an appropriate auxiliary agent to the above active ingredient to obtain a bone strengthening composition that can be administered orally.
• diluents or excipients such as fillers, fillers, binders, disintegrants, surfactants, lubricants and the like that are usually used can be used.
• the excipient include sucrose, lactose, starch, crystalline cellulose, mannitol, light anhydrous silicic acid, magnesium aluminate, synthetic aluminum silicate, magnesium magnesium metasilicate, calcium carbonate, sodium hydrogen carbonate, calcium hydrogen phosphate
• carboxymethylcellulose calcium and the like can be added in combination.
• a column (5 cm in diameter ⁇ 30 cm in height) packed with 400 g of a cation exchange resin sulfonated chitopearl (Fujibo Co., Ltd.) was thoroughly washed with deionized water, and then 40 liters of unsterilized skim milk (pH 6. 7) was passed at a flow rate of 25 ml / min. After passing through the column, the column was thoroughly washed with deionized water, and the basic protein fraction adsorbed on the resin was eluted with 0.02 M carbonate buffer (pH 7.0) containing 0.98 M sodium chloride.
• 0.02 M carbonate buffer pH 7.0
• Example Product 1 The eluate was desalted with a reverse osmosis (RO) membrane, concentrated, and then freeze-dried to obtain 21 g of a powdered milk-derived basic protein fraction (Example Product 1).
• the obtained milk-derived basic protein fraction was measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and the molecular weight was distributed in the range of 3,000-80,000.
• the composition was as shown in Table 1.
• Table 2 shows the results of analyzing the amino acid composition with an amino acid analyzer (L-8500, manufactured by Hitachi, Ltd.) after hydrolysis with 6N hydrochloric acid at 110 ° C. for 24 hours.
• amino acid analyzer L-8500, manufactured by Hitachi, Ltd.
• Example product 3 After dissolving 50 g of the milk-derived basic protein fraction obtained in Example 1 in 10 liters of distilled water, 1% pancreatin (manufactured by Sigma) was added and reacted at 37 ° C. for 2 hours. After the reaction, the enzyme was inactivated by heat treatment at 80 ° C. for 10 minutes to obtain 48.3 g of a milk-derived basic protein fraction degradation product (Example product 3).
• Papain 50 U / g whey protein and Proleather (Amano Enzyme) 150 U / g whey protein are added to 1 L of 10% aqueous whey protein solution, adjusted to pH 8 and enzyme denatured at 55 ° C. for 6 hours. Decomposition was performed. The reaction solution was heated at 100 ° C. for 15 seconds or more to inactivate the enzyme, centrifuged to collect the supernatant, and dried to obtain a whey protein hydrolyzate (Example product 4). The molecular weight distribution of the obtained whey protein hydrolyzate was 10 kDa or less, the main peak was 1.3 kDa, APL was 7.2, and the free amino acid content for all components was 18.9%.
• Papain 50 U / g whey protein and Pro Leather (Amano Enzyme) 150 U / g whey protein were added to 1 L of a 10% whey protein aqueous solution and subjected to enzymatic degradation at pH 8 and 50 ° C. for 3 hours. The temperature was raised to 55 ° C. and maintained at this temperature for 3 hours to denature the protein and enzymatically decompose the protein, and heated at 100 ° C. for 15 seconds or more to deactivate the enzyme.
• This reaction solution is treated with a UF membrane (manufactured by STC) having a fractional molecular weight of 10 kDa and an MF membrane (manufactured by STC) having a fractional molecular weight of 300 Da to collect a concentrated liquid fraction, which is dried and dried with whey protein hydrolyzate.
• a decomposition product (Example product 5) was obtained.
• the molecular weight distribution of the obtained whey protein hydrolyzate was 10 kDa or less, the main peak was 500 Da, APL was 3.0, and the free amino acid content for all components was 15.2%.
• the decrease in antigenicity against ⁇ -lactoglobulin was measured by the Inhibition ELISA method, it was 1 / 10,000 or less, the degradation rate was 32%, the yield was 65.4%, and the bitterness was 2.
• a hydrolyzate of whey protein was prepared by the method reported in JP-A-4-69315. 120 g of whey protein was dissolved in 1,800 ml of purified water, and the pH was adjusted to 7.0 with 1M caustic soda solution. Next, the mixture was sterilized by heating at 60 ° C. for 10 minutes, and maintained at 45 ° C., 20 g of Amano A (Amano Enzyme) was added, and reacted for 2 hours. The enzyme was inactivated by heating at 80 ° C. for 10 minutes and freeze-dried to obtain a whey protein hydrolyzate (Example product 6).
• the molecular weight distribution of the obtained whey protein hydrolyzate was 14 kDa or less, the main peak was 3.1 kDa, APL was 17.2, and the free amino acid content for all components was 13.2%.
• the decrease in antigenicity with respect to ⁇ -lactoglobulin was measured by Inhibition ELISA and found to be 1 / 5,000 or less, the degradation rate was 18%, the yield was 80.6%, and the bitterness was 2.
• a hydrolyzate of whey protein was prepared by the method reported in JP-A-4-69315. 120 g of whey protein was dissolved in 1,800 ml of purified water, and the pH was adjusted to 7.0 with 1M caustic soda solution. Subsequently, the mixture was sterilized by heating at 60 ° C. for 10 minutes, and maintained at 45 ° C., 20 g of Amano A (manufactured by Amano Enzyme) was added, and the mixture was reacted for 8 hours. The enzyme was inactivated by heating at 80 ° C. for 10 minutes and freeze-dried to obtain a whey protein hydrolyzate (Example product 7).
• the molecular weight distribution of the obtained whey protein hydrolyzate was 10 kDa or less, the main peak was 1.8 kDa, APL was 10.0, and the free amino acid content for all components was 19.3%.
• the decrease in antigenicity against ⁇ -lactoglobulin was measured by the Inhibition ELISA method, it was 1 / 10,000 or less, the degradation rate was 25%, the yield was 80.6%, and the bitterness was 2.
• mice Group in which physiological saline is administered to mice (Group A), group in which 1 mg of the milk-derived basic protein fraction of Example Product 1 is administered per kg of mouse body weight (Group B), and whey protein hydrolysis of Example Product 4 Group (group C) administered 2 mg / kg body weight of mouse, group (group D) administered 3 mg / kg of mouse-derived basic protein fraction of example product 1 per kg body weight, whey protein hydrolysis of example product 4 Group (E group) in which 3 mg / kg of mouse body weight is administered, milk-derived basic protein fraction of Example product 1 and whey protein hydrolyzate of Example product 4 are simultaneously administered at 1 mg and 2 mg / kg of mouse weight, respectively.
• the test group (Group F) was divided into 6 test groups (10 animals per group).
• Example products 1 and 4 and a mixture of Example products 1 and 4 were each suspended in physiological saline and orally administered to groups BF.
• the bone density of the tibia of the right foot of the mouse was measured using 3D micro X-ray CT (Rigaku Corporation). The results are shown in Table 4.
• the bone density of the tibia after the administration for 2 weeks was as follows: the group obtained by administering 1 mg or 3 mg of the milk-derived basic protein fraction of Example Product 1 per 1 kg of the mouse body weight, and the whey protein hydrolyzate of Example Product 4 In the group administered 2 mg or 3 mg per kg body weight, the group derived from the milk-derived basic protein fraction of Example product 1 and the whey protein hydrolyzate of Example product 4 at 1 mg and 2 mg per kg body weight of the mouse, respectively, the control Compared to the group, bone density was significantly increased.
• Example Product 5 are administered orally once a day with a sonde to give 1 mg and 2 mg (Group C), respectively, per 1 kg body weight of the rat, or physiological saline that is a solvent that does not contain any Example product Was administered orally once daily with a sonde (control group) and reared for 16 weeks.
• the rats subjected to sham operation were orally administered with saline only once a day (sham operation group) as in the control group.
• the bone strength of the right femur of the rat was measured with a bone strength measuring device (RX-1600, iTechno). The results are shown in Table 5.
• Example 3 The effect of osteoblast proliferation on the milk-derived basic protein fraction or the milk-derived basic protein fraction degradation product of Example products 2 and 3 and the whey protein hydrolyzate of Example products 6 and 7 were examined.
• the established osteoblast (MC3T3-E1) was seeded in a 96-well flat cell culture plate and cultured in ⁇ -MEM medium containing 10% fetal bovine serum for 24 hours. After removing all the medium, 90 ⁇ l of ⁇ -MEM medium not containing fetal calf serum was added, and Example Products 2, 3, 6, 7 and a mixture of Example Products 2 and 6, Example Products 3 and 7 The mixture was added in an amount of 10 ⁇ l, and the culture was continued for another 24 hours.
• Bromodeoxyuridine (BrdU) attached to Cell Proliferation kit (manufactured by GE Healthcare) was added and incubated for 2 hours, followed by reaction with peroxidase-labeled anti-BrdU antibody, and the substrate 3,3 ′, 5,5′-tetra
• the amount of BrdU incorporated into the cells was measured to determine the osteoblast proliferation activity. The results are shown in Table 6.
• the mixture of No. 6 and the mixture of Example product 3 and Example product 7 were added, the proliferation of osteoblasts was significantly promoted compared to the case where physiological saline was added to the medium.
• milk-derived basic protein fraction and / or milk-derived basic protein fraction degradation product and whey protein hydrolyzate are added to the medium at the same time, osteoblasts are significantly higher than when they are added alone. It showed proliferative activity.
• the milk-derived basic protein fraction and / or the mixture of the milk-derived basic protein fraction hydrolyzate and the whey protein hydrolyzate of the present invention are compared with each other in synergistic osteoblastic cells. It was found to have a growth promoting effect.
• Example 1 and 3 milk-derived basic protein fraction or milk-derived basic protein fraction degradation product and Example products 4 and 5 whey protein hydrolyzate were examined for the effect of inhibiting bone resorption by osteoclasts. It was. 5 days old rabbit tibia and femur were removed and soft tissue was removed, and then total bone marrow cells including osteoclasts mechanically minced in DMEM / F12 medium containing 5% FBS were obtained at 1,000,000 cells / The cells were plated on wells of a crystalline calcium phosphate plate (Corrig) so as to be well and cultured.
• Corrig crystalline calcium phosphate plate
• Example Product 4 5-Whey Protein Hydrolyzate, Example Product 1 and Example Product 5 were added to the culture medium.
• the area of the pits was significantly reduced as compared with the case where physiological saline was added to the medium.
• the milk-derived basic protein fraction and / or the milk-derived basic protein fraction degradation product and the whey protein hydrolyzate are added to the medium at the same time, the pit area is significantly reduced compared to the case where each is added alone. did.
• the milk-derived basic protein fraction and / or the mixture of the milk-derived basic protein fraction hydrolyzate and the whey protein hydrolyzate according to the present invention are based on synergistic osteoclasts in comparison with each other. It was found that there is an action to suppress bone resorption.
• the bone strengthening tablet of the present invention was produced by molding and tableting into 1 g by a conventional method. In 1 g of this tablet, 25 mg of the milk-derived basic protein fraction of Example product 1 and 50 mg of the whey protein hydrolyzate of Example product 4 were contained.
• the beverage for bone strengthening contained 50 mg of the milk-derived basic protein fraction of Example Product 2 and 100 mg of the whey protein hydrolyzate of Example Product 5 per 100 ml.

Landscapes

• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• General Health & Medical Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Physical Education & Sports Medicine (AREA)
• Veterinary Medicine (AREA)
• Medicinal Chemistry (AREA)
• Engineering & Computer Science (AREA)
• Public Health (AREA)
• Pharmacology & Pharmacy (AREA)
• Animal Behavior & Ethology (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Organic Chemistry (AREA)
• General Chemical & Material Sciences (AREA)
• Orthopedic Medicine & Surgery (AREA)
• Chemical Kinetics & Catalysis (AREA)
• Rheumatology (AREA)
• Epidemiology (AREA)
• Immunology (AREA)
• Zoology (AREA)
• Proteomics, Peptides & Aminoacids (AREA)
• Biomedical Technology (AREA)
• Biotechnology (AREA)
• Cell Biology (AREA)
• Developmental Biology & Embryology (AREA)
• Virology (AREA)
• Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
• Coloring Foods And Improving Nutritive Qualities (AREA)
• Fodder In General (AREA)
• Non-Alcoholic Beverages (AREA)

Applications Claiming Priority (2)

Publications (1)

Family

ID=49116799

Family Applications (1)

Country Status (3)

Cited By (2)

Citations (6)

• 2012
  • 2012-03-09 JP JP2012053825A patent/JP5969225B2/ja active Active
• 2013
  • 2013-03-06 WO PCT/JP2013/056157 patent/WO2013133326A1/fr not_active Ceased
  • 2013-03-08 TW TW102108276A patent/TWI607709B/zh active

Patent Citations (6)

Non-Patent Citations (4)

Also Published As

Similar Documents

Legal Events