CN114042146A - Bovine bone peptide composition and application thereof in preparing medicines for regulating intestinal flora and preventing and treating osteoporosis - Google Patents
Bovine bone peptide composition and application thereof in preparing medicines for regulating intestinal flora and preventing and treating osteoporosis Download PDFInfo
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Abstract
The invention relates to the technical field of bioactive peptides, in particular to a bovine bone peptide composition and application thereof in preparing medicines for regulating intestinal flora and preventing and treating osteoporosis. The bovine bone peptide composition provided by the invention can be used for preventing and treating osteoporosis, promoting the growth and development of thighbone, regulating calcium and phosphorus metabolism, promoting bone formation and reducing bone absorption. Through biological evaluation in animal bodies, the bovine bone peptide composition provided by the invention can change the diversity and species community composition of mouse intestinal flora, so that the disordered intestinal flora tends to be normal, and the bone metabolism is regulated, thereby improving osteoporosis.
Description
Technical Field
The invention relates to the technical field of bioactive peptides, in particular to a bovine bone peptide composition and application thereof in preparing medicines for regulating intestinal flora and preventing and treating osteoporosis.
Background
In addition to the cells of the human body, there are also millions of microorganisms, of which about 90% are present in the human intestinal tract, called the intestinal flora. With the development of the technology, researchers find that the intestinal flora is closely related to the physiological states of human characters, aging and the like, and also related to the occurrence and development of various diseases of human bodies, such as autoimmune diseases including inflammatory bowel diseases, asthma and the like, osteoporosis, even brain diseases including parkinson and the like. It has also been suggested by researchers that modulating the intestinal flora may be an effective way to treat diseases, such as diabetes, parkinson's disease, alcoholic fatty liver, etc. by using the intestinal flora.
Osteoporosis is a common disorder of bone metabolism, and the main clinical manifestations are decreased bone density, damaged bone microstructure, bone pain, susceptibility to fracture, etc. The pathogenic factors of osteoporosis are complex and mainly involve inflammation, aging, nutritional disorders, oxidative stress, hormonal metabolic disorders, and the like. At present, the common osteoporosis treatment drugs are mainly chemical drugs, such as estrogen regulators, bisphosphonates, selective cathepsin K inhibitors and the like. The use of the medicines improves the bone health to a certain extent, improves the survival rate of patients, and increases the risk of breast cancer and venous vascular embolism to different extents. In recent years, more and more studies have shown that osteoporosis is closely related to the intestinal flora. The intestinal flora can influence the bone metabolism through the intestinal tract-bone axis and is an important cause of osteoporosis. Further research shows that intestinal flora can directly or indirectly influence the absorption and metabolism of medicine and nutrient substances by organisms, thereby influencing the treatment effect of medicine and the like on osteoporosis. Therefore, the prevention and treatment of osteoporosis by taking the intestinal flora as a target point is a treatment strategy with great application prospect.
The bone collagen peptide, called bone peptide for short, is a bioactive polypeptide prepared by enzymolysis of animal bones and is widely used in foods and medicines for improving bone health. Bovine bone peptide is the most common type of bone peptide, and mainly comprises bovine bone peptide, yak bone peptide and the like. Research shows that the bovine bone peptide can promote bone formation, enhance osteoblast activity, inhibit apoptosis and weaken bone loss in vivo by activating Wnt/beta-catenin, EGFR and other signal channels, and has obvious anti-osteoporosis activity in vitro cell level and in vivo animal level. Although bovine bone peptide is an important source of drugs for preventing and treating osteoporosis, the clinical therapeutic effect of bovine bone peptide alone is not significant.
The traditional Chinese medicine has the characteristics of obvious curative effect, multiple action targets, small toxic and side effects, suitability for long-term administration and the like, and has an important effect in preventing and treating osteoporosis. Eucommia bark and achyranthes bidentata are common Chinese medicinal materials used as both medicine and food, are listed as top grade in Shen nong's herbal Jing, and have the effects of nourishing liver and kidney, strengthening tendons and bones and the like. Based on the research of modern pharmacology, eucommia ulmoides and achyranthes bidentata can inhibit skeletal inflammation, promote osteoblast proliferation and inhibit osteoclast differentiation by regulating signal transduction pathways such as RANKL/RANK/OPG, Wnt/beta-catenin and the like. At present, no published report on the application of eucommia ulmoides and radix achyranthis bidentatae in combination with bovine bone peptide in intestinal flora remodeling and osteoporosis prevention is found.
Disclosure of Invention
The invention aims to provide a bovine bone peptide composition which can be used for preparing medicines for regulating intestinal flora and preventing and treating osteoporosis. The bovine bone peptide composition provided by the invention can prevent and treat osteoporosis by remodeling intestinal flora.
In a first aspect, the invention provides a bovine bone peptide composition, which is prepared from the following raw materials in parts by weight: 8-50 parts of bovine bone peptide, 10-30 parts of eucommia ulmoides and 5-20 parts of radix achyranthis bidentatae; the bovine bone peptide is the bovine bone collagen peptide obtained by digesting fresh yellow bovine bone or yak bone with neutral protease and compound protease.
Preferably, the bovine bone peptide composition provided by the invention is prepared from the following raw materials in parts by weight: 30-50 parts of bovine bone peptide, 10-20 parts of eucommia ulmoides and 5-10 parts of radix achyranthis bidentatae.
The Eucommiae cortex is dry bark of eucommia ulmoides Oliver belonging to family Eucommiaceae; the Achyranthis radix is derived from dried root of Achyranthis radix of Amaranthaceae.
Preferably, the bovine bone peptide composition provided by the invention is prepared from the following raw materials in parts by weight: 8-10 parts of bovine bone peptide, 20-30 parts of eucommia ulmoides and 10-15 parts of radix achyranthis bidentatae.
In a second aspect, the invention provides a preparation method of the bovine bone peptide composition, which comprises the steps of mixing eucommia ulmoides and radix achyranthis bidentatae with water, heating and decocting for 0.5-1h, adding bovine bone peptide, continuing to decoct for 15-30min, filtering, and removing medicine residues, thus obtaining the bovine bone peptide composition.
In order to obtain the bovine bone peptide composition in an oral liquid state, the preparation method of the bovine bone peptide composition comprises the following steps:
soaking Eucommiae cortex and Achyranthis radix in water for 0.5-2 hr, boiling, heating, decocting until the water content is 20-35%, removing residue, standing the decoction at room temperature, and adding bovine bone peptide to dissolve completely.
In a third aspect, in the preparation method provided by the present invention, in order to better combine with functional food or pharmaceutically acceptable excipients to perform tabletting to prepare solid preparations such as powder, tablet and the like, when the eucommia ulmoides extract and the achyranthes bidentata extract are used for preparing the bovine bone peptide composition, the eucommia ulmoides extract and the achyranthes bidentata extract can be obtained by the following methods:
firstly, the eucommia ulmoides and the radix achyranthis bidentatae are respectively added with water and boiled to prepare the eucommia ulmoides or the radix achyranthis bidentatae extract. Then, mixing the eucommia ulmoides extract, the achyranthes bidentata extract and the bovine bone peptide in a weight ratio of (2-1.5): (1-2): (6-8) mixing to obtain the bovine bone peptide composition.
Preferably, decocting Eucommiae cortex or Achyranthis radix with water respectively until the volume of the mixture is 45-55% of the initial volume, removing residue, heating to viscous, cooling, solidifying, and grinding into powder to obtain Eucommiae cortex extract or Achyranthis radix extract;
the invention also claims application of the bovine bone peptide composition or the bovine bone peptide composition prepared by the preparation method in preparing powder, oral liquid, granules, tablets and/or capsules by combining with pharmaceutically or functionally food acceptable auxiliary materials.
And the application of the bovine bone peptide composition or the preparation method in preparing the medicine for preventing and treating osteoporosis.
The invention has the beneficial effects that:
(1) the bovine bone peptide composition provided by the invention can obviously improve basic growth and development indexes such as weight, body length and the like of an osteoporosis rat, and promote the growth of size and weight of thighbone; effectively improve negative calcium balance and bone metabolism disorder caused by calcium deficiency, and regulate balance between bone absorption and bone formation.
(2) The bovine bone peptide composition provided by the invention can obviously change the intestinal flora structure of an osteoporosis rat, obviously increase the abundance of beneficial pileus family, reduce the abundance of harmful erysipelothrix family, improve the intestinal metabolic function and the intestinal health by influencing the distribution of the intestinal flora, and maintain the intestinal immune steady state.
(3) The bovine bone peptide composition provided by the invention is simple in component, can be used for preparing medicines for regulating intestinal flora and preventing and treating osteoporosis, can be added into functional foods for preventing and treating osteoporosis and regulating intestinal flora, develops a new medicinal approach of the bovine bone peptide composition, and has practical application and popularization values.
Drawings
FIG. 1 is a graph showing the effect of different substances on the diversity of intestinal flora in osteoporotic rats in accordance with the present invention; wherein, M, osteoporosis model group; CBP, buffalo bone peptide group; YBP, yak bone peptide group; TCM, eucommia ulmoides/achyranthes bidentata group; CBP _ TCM, buffalo bone peptide combination group; BP _ TCM, yak bone peptide combination group.
Detailed Description
The present invention will be described in detail with reference to examples. In the embodiment of the invention, the bovine bone peptide is prepared by performing two-step enzymolysis on fresh yellow bovine bones or fresh yak bones by neutral protease and compound protease. The Eucommiae cortex is derived from dried bark of eucommia ulmoides Oliver belonging to family Eucommiaceae. Said Achyranthis radix is derived from dried root of Achyranthis radix of Amaranthaceae.
Example 1 bovine bone peptide composition prepared from bovine bone peptide, eucommia ulmoides and achyranthes bidentata
This example provides a bovine bone peptide composition prepared from bovine bone peptide, eucommia ulmoides and achyranthes bidentata. The method comprises the following specific steps:
mixing 30g of eucommia bark and 15g of achyranthes bidentata with 500mL of water, heating and decocting for 1h, adding 10g of yak bone peptide or cattle bone peptide, continuing to decoct for 15min, filtering, and removing residues to obtain the traditional Chinese medicine.
Example 2 oral liquid prepared from bovine bone peptide, eucommia ulmoides and achyranthes bidentata
This example provides an oral liquid prepared from bovine bone peptide, eucommia ulmoides and achyranthes bidentata. The method comprises the following specific steps:
soaking 50g of eucommia ulmoides and 50g of achyranthes bidentata decoction pieces in 600mL of water, soaking the medicinal materials for 1h, continuously heating for 1h after boiling, heating and boiling to 120mL, removing decoction dregs, standing the decoction at room temperature, adding 20g of bovine bone peptide to fully dissolve the bovine bone peptide, adding a proper amount of pharmaceutically acceptable auxiliary materials, and preparing the oral liquid.
Example 3 preparation of a bovine bone peptide composition from eucommia ulmoides extract, achyranthes bidentata extract and bovine bone peptide
This example provides a bovine bone peptide composition prepared from eucommia ulmoides extract, achyranthes bidentata extract, and bovine bone peptide. The method comprises the following specific steps:
respectively mixing 48g of eucommia ulmoides, 24g of radix achyranthis bidentatae and 800mL of water, heating and decocting to 300mL, removing medicine residues, respectively freeze-drying to obtain eucommia ulmoides extract powder and radix achyranthis bidentatae extract powder, and mixing the eucommia ulmoides extract, the radix achyranthis bidentatae extract and yak bone peptide according to a weight ratio of 10: 5: 30, and mixing uniformly to obtain the product.
Example 4 other formulations prepared from eucommia ulmoides extract, achyranthes bidentata extract and bovine bone peptide
This example provides additional formulations of the bovine ossotide compositions. The method comprises the following specific steps:
decocting Eucommiae cortex and Achyranthis radix in water respectively until the volume is reduced by half, removing residues, heating to viscous solution, cooling, solidifying, grinding into powder, mixing Eucommiae cortex extract, Achyranthis radix extract and beef bone peptide at weight ratio of 15:8:40, adding appropriate functional food or pharmaceutically acceptable adjuvants, and tabletting to obtain solid preparation such as powder and tablet.
The bovine bone peptide composition prepared by the invention has good technical effect after repeated experiments, and the experimental data are as follows:
experiment one: effect of bovine bone peptide composition on rat body weight and femur
Establishing an osteoporosis rat model: selecting SPF (specific pathogen free) grade SD rats (70 +/-10 g) with the age of 4 weeks, carrying out normal adaptive feeding for one week, and then feeding low-calcium feed to construct an osteoporosis rat model. The experiment is divided into six groups, namely an osteoporosis model group, a yellow cattle bone peptide group, a yak bone peptide group, an eucommia/achyranthes bidentata group, a yellow cattle bone peptide combination group and a yak bone peptide combination group. Gavage was administered 1 time daily for 6 weeks. Body weights of the rats in each group were measured and recorded during feeding. After the experiment, all groups of rats are sacrificed, femurs on both sides of all groups of rats are stripped, muscles and connective tissues on the surfaces of bones are removed, and the wet weight of the femurs and the diameter of the midpoint of the femurs of all groups of rats are measured. The experimental results (table 1) showed that the weight, the wet weight of femur, and the diameter of femur of rats were increased to different degrees not only in the osteoporosis model group but also in the cattle bone peptide combination group and yak bone peptide combination group fed alone and in the eucommia/achyranthes bidentata group.
TABLE 1 Effect of bovine ossotide compositions on rat body weight and femur
| Grouping | Average body weight/g | FemurWet weight/g | Diameter/cm of femur |
| Osteoporosis model set | 291.83±25.06 | 0.60±0.09 | 4.05±0.19 |
| Cattle bone peptide group | 311.06±38.46 | 0.65±0.08 | 4.09±0.30 |
| Yak bone peptide group | 297.22±18.45 | 0.67±0.02 | 4.12±0.11 |
| Eucommia/achyranthes bidentata group | 298.30±18.74 | 0.67±0.04 | 4.17±0.15 |
| Cattle bone peptide combination group | 324.72±16.21 | 0.68±0.08 | 4.25±0.35 |
| Yak bone peptide combination group | 302.86±24.80 | 0.72±0.09 | 4.24±0.24 |
Experiment two: influence of bovine bone peptide composition on biochemical indexes of osteoporosis rat serum
The rats in each group were sacrificed after the abdominal aorta was bled, the bled blood was allowed to stand at 37 ℃ for 2h overnight at 4 ℃ and the supernatant serum was frozen at-80 ℃ for further use. The serum calcium content, the content of bone morphogenetic protein-2 (BMP-2) and the content of inflammatory factor interleukin-1 beta (IL-1 beta) in the serum of each group of rats are detected strictly according to the instruction of a corresponding kit. The experimental results show (table 2) that compared with the control group, the bovine bone peptide composition can significantly improve the blood calcium content, promote the expression of a bone formation marker BMP-2 and inhibit the secretion of a bone resorption marker IL-1 beta. The experimental result shows that the bovine bone peptide composition corrects negative calcium balance to a certain extent, promotes bone formation and inhibits bone resorption.
TABLE 2 Effect of bovine ossotide compositions on the Biochemical indicators of serum from osteoporotic rats
Experiment three: influence of bovine bone peptide composition on diversity of intestinal flora of osteoporosis rats
The feces of each group of rats were collected under sterile conditions and stored at-80 ℃ for future use. Separating microorganisms in excrement, extracting genome DNA, and performing microbial diversity analysis on bacterial 16SrDNA by using an Illumina Miseq platform. NMDS analysis was performed at OTU level based on unweighted-unifrac distance algorithm. The experimental results show (fig. 1) that the distribution of each set of samples is relatively discrete. The results show that the intestinal flora structure of the rats is changed after the treatment of the bovine bone peptide composition group, and the strain is obviously different among the groups (0.159).
Experiment four: effect of bovine ossotide composition on the composition of intestinal bacterial community in osteoporotic rats
Based on statistics of species abundance of intestinal flora of each group of rats at the family level, the results show (table 3) that the bovine bone peptide composition can increase the abundance of beneficial bacteria lactobacillus (lactobacillus) in intestinal tract of the osteoporotic rats and simultaneously decrease the abundance of harmful bacteria bacteroidetes (bacteroideteae). The Lactobacillus can be used as beneficial bacteria for maintaining intestinal flora steady state, and has effects of increasing calcium bioavailability, increasing bone mass, and protecting bone health. Bacteroides family is a lethal bacterium of intestinal tract condition, and the increase of the abundance of the bacteroides family is closely related to the immune function disorder of the body and the imbalance of the intestinal flora. The results show that the bovine bone peptide composition can regulate intestinal flora of osteoporosis rats and achieves the effect of regulating osteoporosis by regulating the intestinal flora steady state.
TABLE 3 relative abundance of intestinal flora at the family level in each group of rats
| Grouping | Lactobacillaceae family | Bacteroides family |
| Osteoporosis model set | 4.39±1.96 | 11.97±9.88 |
| Cattle bone peptide group | 6.87±2.08 | 7.31±0.42 |
| Yak bone peptide group | 5.20±1.01 | 2.10±1.43 |
| Eucommia/achyranthes bidentata group | 6.58±2.16 | 7.93±5.34 |
| Cattle bone peptide combination group | 9.76±8.52 | 1.75±0.71 |
| Yak bone peptide combination group | 6.15±1.37 | 3.43±0.51 |
The above experiments respectively performed on the bovine bone peptide compositions prepared in different embodiments of the present invention can achieve the same or similar effects, so only summarized descriptions are given, and detailed descriptions are omitted.
In conclusion, the bovine bone peptide composition provided by the invention can obviously improve basic growth and development indexes such as weight, body length and the like of an osteoporosis rat, and promote growth of size and weight of thighbone; effectively improve negative calcium balance and bone metabolism disorder caused by calcium deficiency, and regulate balance between bone absorption and bone formation. Meanwhile, the composition can remarkably change the intestinal flora structure of an osteoporosis rat, remarkably increase the abundance of beneficial pileus family, reduce the abundance of harmful erysipelothrix family, improve the intestinal metabolic function and intestinal health and maintain the intestinal immune steady state by influencing the distribution of the intestinal flora, can be effectively used for preparing medicines for regulating the intestinal flora and preventing and treating osteoporosis, can also be used for preventing and treating osteoporosis and regulating the intestinal flora in functional food, develops a new medicinal way of the bovine bone peptide composition, and has practical application and popularization values.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (9)
1. The bovine bone peptide composition is characterized by being prepared from the following raw materials in parts by weight: 8-50 parts of bovine bone peptide, 10-30 parts of eucommia ulmoides and 5-20 parts of radix achyranthis bidentatae; the bovine bone peptide is a bovine bone small molecular bioactive peptide obtained by performing two-step enzymolysis treatment on fresh yellow bovine bones or fresh yak bones by neutral protease and compound protease.
2. The bovine ossotide composition according to claim 1, wherein the bovine ossotide composition is prepared from the following raw materials in parts by weight: 30-50 parts of bovine bone peptide, 10-20 parts of eucommia ulmoides and 5-10 parts of radix achyranthis bidentatae;
the Eucommiae cortex is derived from dried bark of eucommia ulmoides Oliver belonging to family Eucommiaceae; said Achyranthis radix is derived from dried root of Achyranthis radix of Amaranthaceae.
3. The bovine ossotide composition according to claim 1, wherein the bovine ossotide composition is prepared from the following raw materials in parts by weight: 8-10 parts of bovine bone peptide, 20-30 parts of eucommia ulmoides and 10-15 parts of radix achyranthis bidentatae.
4. The method for preparing the bovine bone peptide composition as claimed in any one of claims 1 to 3, wherein the bovine bone peptide composition is prepared by mixing eucommia ulmoides and achyranthes bidentata with water, heating and decocting for 0.5-1h, adding bovine bone peptide, continuing to decoct for 15-30min, filtering, and removing residues.
5. The method for preparing the bovine bone peptide composition as claimed in any one of claims 1 to 3, wherein the bovine bone peptide composition is obtained by immersing eucommia ulmoides and achyranthes bidentata in water for 0.5 to 2 hours, boiling, then continuously heating and boiling until the immersion liquid is 20 to 35% of the initial water content, filtering to remove the residues, standing the decoction liquid at room temperature, and then adding the bovine bone peptide to fully dissolve the bovine bone peptide composition.
6. A preparation method of a bovine bone peptide composition is characterized in that eucommia ulmoides and achyranthes bidentata are respectively decocted with water to prepare traditional Chinese medicine extracts; then, mixing the eucommia ulmoides extract, the achyranthes bidentata extract and the bovine bone peptide in a weight ratio of (2-1.5): (1-2): (6-8) mixing.
7. The Chinese medicinal extract as claimed in claim 6, wherein the eucommia ulmoides or achyranthes bidentata extract is obtained by decocting eucommia ulmoides or achyranthes bidentata in water until the volume of the mixed solution is 45-55% of the initial water content, removing the residue, continuously heating until the mixed solution is viscous, cooling and solidifying, and grinding into powder.
8. Use of the bovine bone peptide composition according to any one of claims 1 to 3 or the bovine bone peptide composition prepared by the preparation method according to any one of claims 4 to 7 in combination with pharmaceutically or functionally food acceptable excipients to prepare powders, oral liquids, granules, tablets and/or capsules.
9. Use of the bovine ossotide composition according to any one of claims 1 to 3 or the bovine ossotide composition obtained by the preparation method according to any one of claims 4 to 7 in the preparation of a medicament for preventing and treating osteoporosis.
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