Novel protein material

a protein and protein technology, applied in the field of new protein materials, can solve the problems of drug side effects, affecting the quality of protein, and becoming a serious social problem, and achieve the effects of suppressing osteoclast differentiation and osteoclastic bone resorption, promoting osteoblast proliferation, and promoting osteoblast proliferation

Inactive Publication Date: 2015-10-22
SNOW BRAND MILK PROD CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0019]The protein material of the invention exhibits a remarkable bone-strengthening effect through the function of promoting osteoblast proliferation, and suppressing osteoclast differentiation and osteoclastic bone resorption. The drug, food, drink, or feed of the invention strengthens bones, and is useful for prevention and treatment of various bone diseases, such as osteoporosis, fracture, rheumatism, and arthritis.EMBODIMENTS FOR CARRYING OUT THE INVENTION
[0020]A protein material of the invention is characterized in that the protein material includes angiogenin and / or angiogenin hydrolysate in a specific amount, and further includes lactoperoxidase and / or lactoperoxidase hydrolysate in a specific mass ratio with respect to angiogenin and / or angiogenin hydrolysate.
[0021]The protein material of the invention may be a mixture obtained by mixing a fraction containing angiogenin and / or angiogenin hydrolysate and a fraction containing lactoperoxidase and / or lactoperoxidase hydrolysate in a specific mass ratio, a material prepared by directly extracting a fraction containing angiogenin and / or angiogenin hydrolysate and lactoperoxidase and / or lactoperoxidase hydrolysate in a specific mass ratio from milk or a material derived from milk, such as skim milk or whey, or the like. The protein material of the invention may also include a material prepared by enzymatically degrading angiogenin and / or lactoperoxidase.
[0022]When preparing the protein material of the invention by mixing a fraction containing angiogenin and / or angiogenin hydrolysate and a fraction containing lactoperoxidase and / or lactoperoxidase hydrolysate, a fraction prepared from milk of a mammal, such as human, cow, buffalo, goat, or sheep, a fraction produced by genetic engineering, a fraction purified from blood or an internal organ, or the like may be used as the fraction containing angiogenin and / or angiogenin hydrolysate and the fraction containing lactoperoxidase and / or lactoperoxidase hydrolysate. A commercially available purified angiogenin or lactoperoxidase reagent may also be used. In this case, the protein material of the invention may be prepared by adjusting the mass ratio of lactoperoxidase and / or lactoperoxidase hydrolysate to angiogenin and / or angiogenin hydrolysate.
[0023]A product obtained by enzymatically degrading the above fraction containing angiogenin, the angiogenin reagent, the fraction containing lactoperoxidase, the lactoperoxidase reagent, or the like using one or more proteases may be used as angiogenin hydrolysate or lactoperoxidase hydrolysate.
[0024]When preparing the protein material of the invention by directly extracting a fraction which contains angiogenin and / or angiogenin hydrolysate and lactoperoxidase and / or lactoperoxidase hydrolysate in a specific mass ratio from milk or a material derived from milk, such as skim milk or whey, for example, milk or a material derived from milk may be brought into contact with a cation-exchange resin, and then milk-derived proteins adsorbed on the resin may be eluted at a salt concentration of 0.1 to 2.0 M, desalted and concentrated using a reverse osmosis membrane, an electrodialysis membrane, an ultrafiltration membrane, a microfiltration membrane, or the like, after that optionally subjected to limited degradation to a molecular weight of 8000 or less using a protease, such as trypsin, pancreatin, chymotrypsin, pepsin, papain, kallikrein, cathepsin, thermolysin, or V8 protease, When subjecting to limited degradation using a protease, it is preferable that the lower limit of the molecular weight is 500 or more. The protein material thus obtained may be dried by freeze-drying, spray drying, or the like.

Problems solved by technology

In recent years, various bone diseases, such as osteoporosis, fracture, and backache have increased on a global basis along with aging of society and the like, and have become a serious social problem.
However, these drugs may have side effects such as buzzing in the ear, a headache, or loss of appetite.
Moreover, the above substances are in a situation that they cannot be added to a food or drink at present from the viewpoint of safety, cost, and the like.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

reference example 1

Preparation (1) of Angiogenin Fraction

[0036]A column filled with 30 kg of cation-exchange resin (Sulfonated Chitopearl; manufactured by Fuji Spinning Co., Ltd.) was thoroughly washed with deionized water, and 1000 liters of unpasteurized skim milk (pH 6.7) was then applied to the column. After thoroughly washing the column with deionized water, the absorbed protein was eluted with a linear gradient of 0.1 to 2.0 M sodium chloride. The elution fraction containing angiogenin was fractionated using an S-Sepharose cation-exchange chromatography (manufactured by Amersham Bioscientific), and the resulted angiogenin-containing fraction was heat-treated at 90° C. for 10 minutes, and centrifuged to remove a precipitate. The angiogenin-containing fraction was further subjected to gel filtration chromatography (column: Superose 12). The eluate obtained was desalted using a reverse osmosis membrane, and the desalted eluate was freeze-dried to obtain 16.5 g of an angiogenin fraction having an an...

reference example 2

Preparation (2) of Angiogenin Fraction

[0037]A column filled with 10 kg of Heparin Sepharose (manufactured by GE Healthcare) was thoroughly washed with deionized water, and 1000 liters of unpasteurized skim milk (pH 6.7) was then applied to the column. After thoroughly washing the column with a 0.6 M sodium chloride solution, the absorbed protein was eluted with a 1.5 M sodium chloride solution. The eluate was desalted using a reverse osmosis membrane, and the desalted eluate was freeze-dried to obtain 32 g of an angiogenin fraction having an angiogenin purity of 2%. The above successive operations were repeated 50 times.

reference example 3

Preparation of Lactoperoxidase Fraction

[0038]A column (diameter: 5 cm, height: 30 cm) filled with 600 g of cation-exchange resin (Sulfonated Chitopearl; manufactured by Fuji Spinning Co., Ltd.) was thoroughly washed with deionized water, and 360 liters of unpasteurized skim milk (pH 6.7) was applied to the column at a flow rate of 25 ml / min. After thoroughly washing the column with deionized water, the absorbed protein was eluted with a 0.02 M carbonate buffer (pH 7.0) containing 2.0 M sodium chloride. The eluted fraction, containing lactoperoxidase was adsorbed on an S-Sepharose FF column (manufactured by Amersham Bioscientific), and the column was thoroughly washed with deionized water. After equilibration with a 10 mM phosphate buffer (pH 7.0), the adsorbed fraction was eluted with a linear gradient of 0 to 2.0 M sodium chloride to collect a fraction containing lactoperoxidase. The fraction was subjected to gel filtration chromatography using a HiLoad 16 / 60 Superdex 75pg (manufac...

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Abstract

The invention relates to a protein material includes angiogenin and / or angiogenin hydrolysate in an amount of 2 to 15 mg / 100 mg, and lactoperoxidase and / or lactoperoxidase hydrolysate, in the mass ratio to angiogenin and / or angiogenin hydrolysate of 0.3 to 20.

Description

TECHNICAL FIELD[0001]This invention relates to a novel protein material, and a drug, food, drink, or feed that includes the protein material and is useful for prevention and treatment of bone diseases. The protein material has functions of promoting osteoblast proliferation, and suppressing osteoclast differentiation and osteoclastic bone resorption. Therefore, the protein material is useful for prevention and treatment of various bone diseases, such as osteoporosis, fracture, rheumatism, and arthritis.BACKGROUND ART[0002]In recent years, various bone diseases, such as osteoporosis, fracture, and backache have increased on a global basis along with aging of society and the like, and have become a serious social problem. These diseases are caused by insufficient calcium intake, depression of calcium absorption ability, hormone imbalance after menopause, and the like. It is considered that increase the body bone mass as much as possible by activating the osteoblast and bone formation ...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K38/54A23K1/165A23L1/305A23L2/66A61K38/44A61K38/46
CPCA61K38/54A61K38/44A61K38/465A23L1/305A23V2002/00A23L2/66A23K1/1653C12Y111/01007C12Y301/27A23L1/3053A61K38/18A23V2200/306A61K2300/00A61K38/1891A23V2200/00A23K20/147A23K20/142A23K20/189A23K50/40A23L33/17A23L33/18A61P19/00A61P19/08A61P19/10A61P43/00
Inventor OHMACHI, AIKOMATSUYAMA, HIROAKIMORITA, YOSHIKAZUISHIDA, YUKONARA, TAKAYUKIKATO, KENSERIZAWA, ATSUSHI
Owner SNOW BRAND MILK PROD CO LTD
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