A composition for promoting joint health, and methods of making and using the same

Abstract

The application provides a composition for promoting joint health and a preparation method and application thereof, and the composition comprises the following components in parts by weight: 2-20 parts of bovine bone collagen peptide powder, 2-15 parts of N-acetyl glucosamine, 2-15 parts of a calcium compound, 0.1-3 parts of avocado powder and 0.1-3 parts of polygonatum sibiricum redoute powder. The composition can play multiple roles such as promoting cartilage repair and anti-inflammation, achieves the purpose of promoting joint health, and has a good application prospect.

Description

Technical Field

[0001] This invention relates to a composition for promoting joint health, its preparation method, and its application. Background Technology

[0002] Joints are an important component of the human musculoskeletal system, and their health directly affects an individual's quality of life. Articular cartilage is a thin layer of transparent cartilage covering the articular surfaces of bones. It is mainly composed of type II collagen, proteoglycans, chondrocytes, and water, possessing a degree of elasticity and a smooth surface. Articular cartilage plays a vital role in cushioning mechanical stress, resisting pressure, reducing friction, and supporting joint movement. Because articular cartilage lacks a lymphatic and blood supply, its metabolic nutrients and waste products are absorbed and excreted through diffusion from surrounding tissues, resulting in extremely limited self-repair capabilities. Therefore, once articular cartilage is damaged, repair is slow, leading to a series of joint dysfunctions.

[0003] Articular cartilage damage can be caused by a variety of factors, including acute trauma, long-term excessive load, obesity, and aging. In the early stages, it often manifests as joint pain, swelling, limited range of motion, and a grinding sensation. As the disease progresses, the cartilage gradually degenerates and thins, eventually exposing the subchondral bone, leading to osteoarthritis (OA). Osteoarthritis is a common joint disease characterized by degenerative changes in articular cartilage, abnormal subchondral bone remodeling, and low-grade chronic inflammation. Patients often experience symptoms such as pain, stiffness, and functional impairment; in severe cases, it can lead to joint deformities and loss of function.

[0004] Currently, interventions for joint health problems mainly include drug therapy, surgical treatment, and nutritional intervention. Nonsteroidal anti-inflammatory drugs (NSAIDs) and intra-articular injections of hyaluronic acid are commonly used clinically to relieve pain and improve joint lubrication. However, long-term use may cause side effects such as gastrointestinal discomfort and cartilage metabolic disorders, and is not suitable for daily health maintenance in early-stage or sub-healthy individuals. Surgical treatments such as arthroscopic debridement, cartilage transplantation, or joint replacement can restore joint function to some extent, but they are highly invasive, expensive, and cannot reverse the natural degeneration process of cartilage. Although some products targeting joint health exist, they generally suffer from the following shortcomings: the bioavailability of single active ingredients is limited, making it difficult to achieve synergistic effects; most products only focus on a single aspect of cartilage repair or inflammation relief, lacking comprehensive regulation of joint health; and some raw materials pose safety risks, such as allergenicity.

[0005] Therefore, developing a product that can synergistically exert multiple effects such as anti-inflammation and promoting cartilage repair is of great significance for delaying joint degeneration, improving patients' quality of life, and reducing the medical burden. Summary of the Invention

[0006] In view of the shortcomings of the prior art described above, the purpose of this invention is to provide a composition for promoting joint health, its preparation method and application, to solve the problem that the products in the prior art have single efficacy and cannot synergistically exert multiple effects such as anti-inflammation and promoting cartilage repair.

[0007] To achieve the above and other related objectives, the present invention provides a composition for promoting joint health, a method for preparing the composition, and its application.

[0008] The first aspect of the present invention provides a composition for promoting joint health, wherein the composition comprises, by weight, the following components: 2-20 parts bovine bone collagen peptide powder, 2-15 parts N-acetylglucosamine, 2-15 parts calcium compound, 0.1-3 parts avocado powder, and 0.1-3 parts Polygonatum sibiricum powder.

[0009] Specifically, the weight parts of bovine bone collagen peptide powder can be 2 parts, 5 parts, 8 parts, 10 parts, 12 parts, 15 parts, 18 parts, or 20 parts.

[0010] Preferably, the bovine bone collagen peptide powder is in the form of 5-20 parts by weight.

[0011] More preferably, the bovine bone collagen peptide powder is in the form of 5-15 parts by weight.

[0012] Specifically, the weight portions of N-acetylglucosamine can be 2, 5, 8, 10, 12, or 15 parts.

[0013] Preferably, the N-acetylglucosamine is present in 2-10 parts by weight.

[0014] Specifically, the calcium compound can be in parts by weight of 2, 5, 8, 10, 12, or 15.

[0015] Preferably, the calcium compound is present in parts by weight of 2-10 parts.

[0016] Specifically, the weight parts of avocado powder can be 0.1 parts, 0.2 parts, 0.5 parts, 0.8 parts, 1 part, 1.2 parts, 1.5 parts, 1.8 parts, 2 parts, 2.2 parts, 2.5 parts, 2.8 parts, or 3 parts.

[0017] Preferably, the avocado powder is 0.5-3 parts by weight.

[0018] More preferably, the avocado powder is 0.5-2 parts by weight.

[0019] Specifically, the weight parts of Polygonatum powder can be 0.1 parts, 0.2 parts, 0.5 parts, 0.8 parts, 1 part, 1.2 parts, 1.5 parts, 1.8 parts, 2 parts, 2.2 parts, 2.5 parts, 2.8 parts, or 3 parts.

[0020] Preferably, the amount of Polygonatum powder is 0.5-3 parts by weight.

[0021] More preferably, the amount of Polygonatum powder is 0.5-2 parts by weight.

[0022] Preferably, the mass ratio of avocado powder to polygonatum powder is 0.3-3:1; for example, it can be 3:1, 2.5:1, 2:1, 1.5:1, 1:1, 0.5:1, or 0.3:1.

[0023] More preferably, the mass ratio of the avocado powder to the polygonatum powder is 0.5-2:1.

[0024] Preferably, the calcium compound is selected from any one or two of calcium aspartate and calcium citrate.

[0025] More preferably, the calcium compound is calcium aspartate.

[0026] Preferably, the method for preparing the avocado powder includes: peeling and pitting the avocado, slicing the pulp to obtain fresh avocado slices; pre-freezing and vacuum freeze-drying the fresh avocado slices to obtain freeze-dried avocado slices; and pulverizing and grinding the freeze-dried avocado slices to obtain avocado powder.

[0027] More preferably, the temperature during the preparation of the fresh avocado slices is ≤10℃.

[0028] More preferably, the temperature during the preparation of the fresh avocado slices is 2-10℃. Low-temperature operation can reduce the oxidation of unsaturated fatty acids in avocados, prevent the degradation and inactivation of enzymes, vitamins and other components, and ensure their anti-inflammatory activity.

[0029] Preferably, the thickness of the fresh avocado slices is ≤1cm.

[0030] More preferably, the thickness of the fresh avocado slices is 0.1-1 cm. Slicing the avocado pulp makes it easier to dry, ensuring uniform drying, which is beneficial for the preservation of active ingredients. Furthermore, the dry, loose, and porous slices are more suitable for subsequent pulverization.

[0031] Preferably, the pre-freezing includes freezing fresh avocado slices at -60℃ to -80℃ for 3-8 hours. Pre-freezing ensures that the avocado slices are fully frozen before vacuum freeze-drying, which is more conducive to subsequent freeze-drying.

[0032] Preferably, the sublimation drying stage of the vacuum freeze-drying includes: maintaining a vacuum of 5-20 Pa, a temperature of -5 to 5°C, and a time of 15-25 h.

[0033] Specifically, the vacuum level during the sublimation drying stage can be 5 Pa, 10 Pa, 15 Pa, or 20 Pa.

[0034] Specifically, the temperature during the sublimation drying stage can be -5℃, -2℃, 0℃, 2℃, or 5℃.

[0035] Specifically, the time for the sublimation drying stage can be 15h, 18h, 20h, 22h, or 25h.

[0036] Preferably, the desorption drying stage of the vacuum freeze-drying includes: maintaining a vacuum degree of 1-10 Pa, a temperature of 15-25°C, and a time of 5-15 h;

[0037] Most of the water is removed through a sublimation drying stage, followed by further increasing the temperature and vacuum level. The remaining water is then evaporated in a desorption drying stage. Compared to conventional high-temperature drying, vacuum freeze-drying effectively protects the anti-inflammatory components in avocados, such as phytosterols, unsaturated fatty acids, and phenolic compounds, from destruction.

[0038] Preferably, the preparation method further includes cooling the freeze-dried avocado slices in liquid nitrogen before pulverizing.

[0039] By cooling avocado slices in liquid nitrogen before pulverizing, the avocado slices become crisper, making them easier to pulverize, reducing pulverizing speed, and saving pulverizing time.

[0040] Preferably, the avocado powder has a mesh size of 100-200; for example, it can be 100 mesh, 120 mesh, 150 mesh, or 200 mesh.

[0041] More preferably, the avocado powder has a mesh size of 100-150.

[0042] If the powder has a small mesh size, such as 50 mesh, the particles are too coarse and not conducive to brewing. If the powder has a large mesh size, such as 300 mesh, the powder is too fine, difficult to dissolve, and prone to clumping and excessive stickiness.

[0043] Preferably, the preparation method of the Polygonatum powder includes: taking slices of fresh Polygonatum rhizome to obtain fresh Polygonatum slices; steaming, simmering and drying the fresh Polygonatum slices to obtain dried Polygonatum slices; and pulverizing the dried Polygonatum slices to obtain Polygonatum powder.

[0044] More preferably, the thickness of the fresh Polygonatum slices is ≤1cm.

[0045] More preferably, the thickness of the fresh Polygonatum slices is 0.2-1cm.

[0046] Steaming sliced ​​Polygonatum can shorten the steaming time, increase the heating area, and the steam can quickly deactivate enzymes, preventing them from decomposing the effective anti-inflammatory components such as polysaccharides and saponins in Polygonatum. Slicing is also more conducive to removing the numbing sensation of raw Polygonatum in a short time, and it is also beneficial for subsequent drying and pulverization.

[0047] Preferably, the steaming time is 2-5 hours; for example, it can be 2 hours, 2.5 hours, 3 hours, 3.5 hours, 4 hours, 4.5 hours, or 5 hours.

[0048] More preferably, the steaming time is 3-5 hours.

[0049] The specific steaming time can be adjusted according to the thickness of the slices, steaming until soft and tender; for example, when the slices are 0.3cm, the steaming time can be shortened to 2-3 hours, and when the slices are 0.8cm, the steaming time can be 3-4 hours.

[0050] Preferably, the simmering time is 20-100 minutes; for example, it can be 20 minutes, 30 minutes, 40 minutes, 50 minutes, 60 minutes, 70 minutes, 80 minutes, 90 minutes, or 100 minutes.

[0051] More preferably, the simmering time is 20-40 minutes.

[0052] Simmering allows the residual heat to cook the Polygonatum slices more thoroughly and evenly, while also allowing them to cool down slowly, preventing damage from thermal expansion and contraction.

[0053] Preferably, the drying temperature is 50-65℃; for example, it can be 50℃, 55℃, 60℃, or 65℃.

[0054] More preferably, the drying temperature is 50-60°C.

[0055] Preferably, the drying time is 8-15 hours; for example, it can be 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, or 15 hours.

[0056] More preferably, the drying time is 8-12 hours.

[0057] If the drying temperature is too low, such as 40℃, the moisture evaporates slowly and the drying time is too long, making the Polygonatum slices prone to spoilage. If the drying temperature is too high, such as 80℃, it will lead to the degradation of the anti-inflammatory active ingredient Polygonatum polysaccharide, and will also cause uneven drying, with the surface becoming dry and the inside remaining moist.

[0058] Preferably, the particle size of the Polygonatum powder is 100-200 mesh; for example, it can be 100 mesh, 120 mesh, 150 mesh, or 200 mesh.

[0059] More preferably, the particle size of the Polygonatum powder is 100-150 mesh.

[0060] If the powder has a small mesh size, such as 50 mesh, the particles are too coarse and not conducive to brewing. If the powder has a large mesh size, such as 300 mesh, the powder is too fine, difficult to dissolve, and prone to clumping and excessive stickiness.

[0061] Compared to the traditional nine-steaming and nine-drying process for processing Polygonatum, this invention uses slicing, steaming, and simmering processes, which greatly shortens the processing time of Polygonatum and preserves the activity of anti-inflammatory substances to the greatest extent.

[0062] A second aspect of the present invention provides a method for preparing the above-mentioned composition for promoting joint health, the method comprising: mixing bovine bone collagen peptide powder, N-acetylglucosamine, calcium compound, avocado powder and polygonatum powder evenly to obtain the composition.

[0063] Other ingredients may be added to the composition of this invention, such as anti-inflammatory ingredients like turmeric and sodium hyaluronate, and / or sugar alcohols like erythritol and maltitol, and / or flavoring ingredients like pineapple juice and apple juice, all of which fall within the scope of protection of this invention.

[0064] A third aspect of the present invention provides the use of the above-described composition for promoting joint health in the preparation of products for promoting joint health.

[0065] Preferably, the joint health promotion is selected from one or more of the following: repairing cartilage damage and anti-inflammatory.

[0066] A fourth aspect of the present invention provides a product having a joint health promoting effect, the product comprising the aforementioned joint health promoting composition and food-grade acceptable excipients.

[0067] Preferably, the excipients are selected from any one or more of fillers, flavoring agents, coloring agents, antioxidants, thickeners, stabilizers, emulsifiers, dispersants, anti-caking agents, and lubricants.

[0068] As described above, the composition for promoting joint health, its preparation method, and its application according to the present invention have the following beneficial effects:

[0069] This invention is the first to scientifically combine bovine bone collagen peptide powder, N-acetylglucosamine, calcium compounds, avocado powder, and polygonatum powder to prepare a composition that can promote joint health. This composition has low cytotoxicity, is safe, and can also play a variety of roles such as promoting cartilage repair and anti-inflammation.

[0070] In this invention, bovine bone collagen peptide powder and N-acetylglucosamine promote the synthesis of glycosaminoglycans by chondrocytes, which helps repair damaged or worn cartilage tissue. Avocado powder and Polygonatum powder synergistically inhibit pro-inflammatory cytokines, thereby reducing the stimulation of chondrocytes by inflammation, reducing cartilage matrix degradation, reshaping the anti-inflammatory microenvironment, and promoting tissue repair and cartilage regeneration. The synergistic effect of repair and anti-inflammation enhances the cartilage repair effect. Calcium strengthens the joint bone, increases bone density, reduces additional wear caused by uneven joint stress, provides a stable mechanical support environment for cartilage repair, and maintains joint health.

[0071] N-acetylglucosamine is an important raw material for chondrocytes to synthesize proteoglycans and collagen fibers. It can stimulate the proliferation and differentiation of chondrocytes, promote the production of proteoglycans with multimeric structures, increase the repair capacity of chondrocytes, and help repair damaged or worn cartilage, maintaining the normal structure and function of cartilage. The fat-soluble components in avocados, such as phytosterols, vitamins, carotenoids, and unsaturated fatty acids, as well as the water-soluble phenolic components, have certain anti-inflammatory effects. The polysaccharides, steroidal saponins, and flavonoids in Polygonatum also have certain anti-inflammatory effects. Calcium compounds are an important component of bones and can provide raw materials for bone renewal and repair. Bovine bone collagen peptide powder can stimulate the activity of osteoblasts, promote the differentiation of bone marrow mesenchymal stem cells into osteoblasts, and combine with calcium ions to form a soluble complex, preventing calcium precipitation in the alkaline environment of the intestine, thereby improving calcium absorption and allowing calcium compounds to be better deposited in bones, thus increasing bone density. The composition of the above raw materials works synergistically to promote joint health. Attached Figure Description

[0072] Figure 1 The diagram shows the EGFP fluorescence intensity of cartilage in each group of zebrafish in this invention. Detailed Implementation

[0073] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0074] Furthermore, it should be understood that the existence of other method steps before or after the combined steps, or the insertion of other method steps between these explicitly mentioned steps, does not preclude the presence of other method steps before or after the combined steps, unless otherwise stated. It should also be understood that the combined connection relationship between one or more devices / apparatus mentioned in this invention does not preclude the existence of other devices / apparatus before or after the combined devices / apparatus, or the insertion of other devices / apparatus between these explicitly mentioned devices / apparatus. Moreover, unless otherwise stated, the numbering of each method step is merely a convenient tool for identifying each method step, and not for limiting the order of the method steps or defining the scope of the invention. Changes or adjustments to their relative relationships, without substantially altering the technical content, should also be considered within the scope of the invention.

[0075] When numerical ranges are given in the embodiments, it should be understood that, unless otherwise stated in the present invention, both endpoints of each numerical range and any value between the two endpoints may be selected. Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art. In addition to the specific methods, apparatus, and materials used in the embodiments, based on the knowledge of the prior art possessed by one of ordinary skill in the art and the description of this invention, any prior art methods, apparatus, and materials similar to or equivalent to those described, apparatus, and materials in the embodiments of this invention may be used to implement the present invention.

[0076] Bovine bone collagen peptide powder is a functional protein peptide powder made primarily from bovine bones through biological extraction and enzymatic hydrolysis. Rich in various amino acids, it promotes the proliferation of osteoblasts (HOBs), providing the cellular basis for bone growth and repair. Furthermore, by stimulating chondrocytes to synthesize proteoglycans and type II collagen, it helps repair articular cartilage, slows down degenerative joint changes, and improves osteoarthritis. It can also chelate with calcium ions, effectively preventing calcium deposition in the intestines, allowing calcium to be deposited in bones and increasing bone density.

[0077] N-acetylglucosamine (NAG) is a functional monosaccharide found in human articular cartilage and connective tissue. It exerts cartilage-protective and anti-inflammatory effects through a multi-target mechanism. On one hand, as a key precursor for chondrocytes in synthesizing proteoglycans and collagen fibers, it directly stimulates chondrocyte proliferation and differentiation, promoting the production of proteoglycans with normal structure, thereby enhancing the repair capacity of chondrocytes and helping to repair damaged or worn cartilage tissue, maintaining the normal structure and mechanical function of articular cartilage. On the other hand, it also exhibits significant anti-inflammatory activity, inhibiting the synthesis and release of intra-articular inflammatory mediators. For example, by reducing the activity of nitric oxide synthase, it lowers the production of prostaglandin E2 and other inflammatory factors, thereby reducing the inflammatory response of the synovial membrane and alleviating joint discomfort and stiffness caused by inflammation.

[0078] Calcium aspartate is an organic calcium compound formed by the combination of aspartic acid and calcium ions. It is a form of calcium supplement with high bioavailability. Aspartic acid in its molecular structure acts as an active carrier, effectively chelating calcium ions and promoting the transport and absorption of calcium in the body. Calcium aspartate provides a high-quality calcium source for bone metabolism, ensuring bone strength and mineral reserves.

[0079] Avocado (scientific name: *Vitellaria paradoxa* CFGaertn.) is a deciduous tree belonging to the Sapotaceae family and the *Vitellaria* genus. Its flesh is not only rich in fat, but also contains abundant dietary fiber, potassium, vitamin E, and B vitamins. It is also rich in carotenoids, phytosterols, and polyphenolic compounds. These abundant nutrients endow avocados with antioxidant and anti-inflammatory potential. By directly scavenging free radicals, protecting cells from oxidative stress damage, and combining its ability to regulate inflammatory mediators, the multi-dimensional active ingredients from the flesh to the seed work together to exert a comprehensive antioxidant and anti-inflammatory effect.

[0080] Polygonatum sibiricum Delar. ex Redoute is a perennial herb belonging to the order Asparagales, family Liliaceae, and genus Polygonatum. Its active ingredients include polysaccharides, flavonoids, saponins, alkaloids, amino acids, and trace elements. Its rhizome is used medicinally; it is sweet in taste, neutral in nature, and enters the spleen, lung, and kidney meridians. It has the effects of nourishing yin and moistening the lungs, tonifying the spleen and replenishing qi, and nourishing the kidneys and replenishing essence. Modern research shows that Polygonatum extract has significant antioxidant effects, can alleviate oxidative damage, and enhance the activity of antioxidant enzymes such as superoxide dismutase (SOD). It also possesses various health benefits such as antibacterial, immune-regulating, blood sugar-lowering, anti-fatigue, and anti-aging properties.

[0081] Unless otherwise specified, the raw materials used in the following embodiments and comparative examples of this application are all conventional commercially available raw materials and are all in powder form.

[0082] Bovine bone collagen peptide powder was purchased from Hubei Ruibang Biotechnology Co., Ltd.

[0083] N-acetylglucosamine was purchased from Shandong Runde Biotechnology Co., Ltd.

[0084] Calcium aspartate was purchased from Zhengzhou Ruipu Bioengineering Co., Ltd.

[0085] The avocado powder was purchased from Topcon Powder Technology (Kunshan) Co., Ltd.

[0086] Polygonatum powder was purchased from Nanjing Zelang Biotechnology Co., Ltd.

[0087] Calcium citrate was purchased from Rizhao Jinhe Boyuan Biochemical Co., Ltd.

[0088] Example 1

[0089] Example 1 provides a composition for promoting joint health and a method for preparing the same, the composition comprising the following components in parts by weight:

[0090] 5.3 parts bovine bone collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium aspartate, 1 part avocado powder, and 1 part polygonatum powder.

[0091] The preparation method of the composition includes: mixing the above components evenly to obtain the composition.

[0092] The preparation of avocado powder includes the following steps:

[0093] S1. Take a Has avocado, peel and pit it in an environment below 10℃, take out the flesh and cut it into thin slices with a thickness of <1cm. Place the avocado slices flat in a stainless steel tray and place it in an ultra-low temperature freezer at -80℃ for 5 hours.

[0094] S2. Pre-cool the avocado slices in the vacuum freeze dryer to -60℃, then quickly transfer them to the vacuum freeze dryer. Start the vacuum pump to rapidly reduce the vacuum level to below 20Pa, maintaining the cold trap temperature ≤60℃. Perform sublimation drying: raise the heating plate temperature to 0℃ at a rate of 1℃ / min, maintaining a vacuum level of 5-20Pa, and dry for 20 hours. Separation drying stage: raise the heating plate temperature to 20℃ at a rate of 1℃ / min, maintaining a vacuum level of 1-10Pa, and dry for 10 hours to obtain freeze-dried avocado slices.

[0095] S3. In an environment with a temperature ≤20℃ and humidity <30℃, freeze-dried avocado slices are rapidly cooled with liquid nitrogen to make them brittle. They are then transferred to a high-speed grinder and pulverized at 8000 rpm with intermittent cycles of 3 times (20s each time, 30s interval) to coarsely grind them to 10-20 mesh. After that, they are ground in a ball mill at 200 rpm with intermittent cycles of 5 times (5min each time, 5min interval). The powder is then passed through a 120-mesh sieve to obtain avocado powder.

[0096] The preparation of Polygonatum sibiricum powder includes the following steps:

[0097] S1. Take fresh rhizomes of Polygonatum, remove the rootlets, wash and cut into slices with a thickness of <1cm to obtain sliced ​​Polygonatum.

[0098] S2. Add enough water to the steamer, place the steamer basket on top, spread the sliced ​​Solomon's Seal evenly in the steamer basket, keeping the thickness of the spread <3cm, cover and steam. After the water boils and steam rises, turn to medium-low heat (800W), keep the steam in the pot stable, and continue steaming for 3 hours. After turning off the heat, let it sit for 30 minutes before opening the lid to obtain the steamed Solomon's Seal slices.

[0099] S3. Place the steamed Polygonatum slices in an oven and dry at 55°C for 10 hours to obtain dried Polygonatum slices.

[0100] S4. Place the dried Polygonatum slices into a grinder and grind them repeatedly. Pass the powder through a 120-mesh sieve to obtain Polygonatum powder.

[0101] Example 2

[0102] Example 2 provides a composition for promoting joint health and a method for preparing the same, the composition comprising the following components in parts by weight:

[0103] 9 parts bovine bone collagen peptide powder, 6 parts N-acetylglucosamine, 7 parts calcium aspartate, 2 parts avocado powder, and 1.5 parts polygonatum powder.

[0104] The preparation method of the composition includes: mixing the above components evenly to obtain the composition.

[0105] The preparation methods of avocado powder and polygonatum powder in this Example 2 are the same as those in Example 1.

[0106] Example 3

[0107] Example 3 provides a composition for promoting joint health and a method for preparing the same, the composition comprising the following components in parts by weight:

[0108] 12 parts bovine bone collagen peptide powder, 8 parts N-acetylglucosamine, 10 parts calcium aspartate, 2 parts avocado powder, and 3 parts polygonatum powder.

[0109] The preparation method of the composition includes: mixing the above components evenly to obtain the composition.

[0110] The preparation methods of avocado powder and polygonatum powder in this Example 3 are the same as those in Example 1.

[0111] Example 4

[0112] Example 4 provides a composition for promoting joint health and a method for preparing the same, the composition comprising the following components in parts by weight:

[0113] 20 parts bovine bone collagen peptide powder, 15 parts N-acetylglucosamine, 15 parts calcium aspartate, 3 parts avocado powder, and 3 parts polygonatum powder.

[0114] The preparation method of the composition includes: mixing the above components evenly to obtain the composition.

[0115] The preparation methods of avocado powder and polygonatum powder in Example 4 are the same as those in Example 1.

[0116] Example 5

[0117] Example 5 provides a composition for promoting joint health and its preparation method, which differs from Example 1 in that it uses commercially available avocado powder.

[0118] The preparation method of Polygonatum powder in Example 5 is the same as that in Example 1.

[0119] Example 6

[0120] Example 6 provides a composition for promoting joint health and its preparation method, which differs from Example 1 in that it uses commercially available Polygonatum sibiricum powder.

[0121] The preparation method of avocado powder in Example 6 is the same as that in Example 1.

[0122] Example 7

[0123] Example 7 provides a composition for promoting joint health and its preparation method. The difference between this composition and Example 1 is that calcium aspartate is replaced with calcium citrate. Specifically, it includes the following components by weight: 5.3 parts bovine bone collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium citrate, 1 part avocado powder, and 1 part Polygonatum sibiricum powder.

[0124] The preparation methods of avocado powder and polygonatum powder in Example 7 are the same as those in Example 1.

[0125] Example 8

[0126] Example 8 provides a composition for promoting joint health and its preparation method. The difference between this composition and Example 1 is that the amount of avocado powder and polygonatum powder added is reduced. Specifically, it includes the following components by weight: 5.3 parts bovine bone collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium citrate, 0.5 parts avocado powder, and 0.5 parts polygonatum powder.

[0127] The preparation methods of avocado powder and polygonatum powder in Example 8 are the same as those in Example 1.

[0128] Example 9

[0129] Example 9 provides a composition for promoting joint health and its preparation method. The difference between Example 1 and Example 9 is that the addition ratio of avocado powder and polygonatum powder is changed, while the total addition amount remains the same. Specifically, it includes the following components by weight: 5.3 parts of fish collagen peptide powder, 2.7 parts of N-acetylglucosamine, 4.9 parts of calcium aspartate, 1.7 parts of avocado powder, and 0.3 parts of polygonatum powder.

[0130] The preparation methods of avocado powder and polygonatum powder in Example 9 are the same as those in Example 1.

[0131] Example 10

[0132] This embodiment 10 provides a composition for promoting joint health and its preparation method. The difference between this composition and that of embodiment 1 is that the addition ratio of avocado powder and polygonatum powder is changed, while the total addition amount remains the same. Specifically, it includes the following components by weight: 5.3 parts fish collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium aspartate, 0.3 parts avocado powder, and 1.7 parts polygonatum powder.

[0133] The preparation methods of avocado powder and polygonatum powder in this Example 10 are the same as those in Example 1.

[0134] Comparative Example 1

[0135] Comparative Example 1 provides a composition for promoting joint health and its preparation method. The difference between this composition and Example 1 is that avocado powder is not added. Specifically, it includes the following components by weight: 5.3 parts bovine bone collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium aspartate, and 1 part Polygonatum sibiricum powder.

[0136] The preparation method of Polygonatum powder in Comparative Example 1 is the same as that in Example 1.

[0137] Comparative Example 2

[0138] Comparative Example 2 provides a composition for promoting joint health and its preparation method. The difference between this composition and Example 1 is that no Polygonatum sibiricum powder is added. Specifically, it includes the following components by weight: 5.3 parts bovine bone collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium aspartate, and 1 part avocado powder.

[0139] The preparation method of avocado powder in Comparative Example 2 is the same as that in Example 1.

[0140] Comparative Example 3

[0141] Comparative Example 3 provides a composition for promoting joint health and its preparation method. The difference between this composition and Example 1 is that bovine bone collagen peptide powder is replaced with fish collagen peptide powder. Specifically, it includes the following components by weight: 5.3 parts fish collagen peptide powder, 2.7 parts N-acetylglucosamine, 4.9 parts calcium aspartate, 1 part avocado powder, and 1 part Polygonatum sibiricum powder.

[0142] The preparation methods of avocado powder and polygonatum powder in Comparative Example 3 are the same as those in Example 1.

[0143] Comparative Example 4

[0144] Comparative Example 4 provides a composition for promoting joint health and its preparation method, which differs from Example 4 in that: the amount of bovine bone collagen peptide powder added is increased. The specific composition includes the following components in parts by weight:

[0145] 40 parts bovine bone collagen peptide powder, 15 parts N-acetylglucosamine, 15 parts calcium aspartate, 3 parts avocado powder, and 3 parts polygonatum powder.

[0146] The preparation method of the composition includes: mixing the above components evenly to obtain the composition.

[0147] The preparation methods of avocado powder and polygonatum powder in Comparative Example 4 are the same as those in Example 1.

[0148] The formulations of the compositions in Examples 1-10 and Comparative Examples 1-4 are shown in Table 1.

[0149] Table 1. Composition formulations in Examples 1-10 and Comparative Examples 1-4

[0150]

[0151] Experimental Section

[0152] Experiment 1: The effect of the composition on cartilage repair

[0153] 1. Reagents, consumables and instruments

[0154] Reagents and consumables: Transgenic cartilage green fluorescent zebrafish Tg (Co12ala:eGFP) (purchased from Huante Biotechnology), methylcellulose (sigma, CAS: 497-76-7), chondroitin sulfate A sodium salt (C758506-5g, Aladdin), dexamethasone (source leaf BR, 99%, S17003-5g).

[0155] Instruments: Biochemical incubator (HT-250H-T, Huante Biotechnology, China), six-well culture plate (corning, USA), stereomicroscope (MZ62, Mshot, China), microscope imaging system (BX53, OLYMPUS, Japan).

[0156] 2. Experimental Procedure

[0157] Normally developing 2-3 dpf Tg (col2a1a:EGFP) transgenic cartilaginous fluorescent zebrafish embryos were randomly selected and placed in six-well plates, 30 embryos per well. Without harming the embryos, the standard dilution water was removed from the six-well plate, and 3 mL of the corresponding concentration of the test substance dilution was quickly added to each well. Except for the blank control group, all other groups were treated with 25 μM (9.81 μg / mL, prepared as 9.81 mg / mL stock solution dissolved in DMSO) of dexamethasone to establish a zebrafish cartilage injury model. The positive control was 1 mg / mL chondroitin sulfate. The culture plate was covered and wrapped with aluminum foil, and incubated in a (28.5±1)℃ biochemical incubator in the dark for 3 days. After 3 days, the MTC of the samples on zebrafish was measured. When the concentration of each composition was 0.3 mg / mL, no obvious abnormalities or deaths were observed in the zebrafish. When the concentration of each composition was 0.5 mg / mL, a small number (3%-10%) of zebrafish died in each group. Therefore, the MTC of this composition is 0.3 mg / mL.

[0158] 2.1 Experimental Grouping

[0159] Blank control group: containing 2-3 dpf zebrafish embryos and 3 mL of standard dilution water;

[0160] Model control group: containing 2-3 dpf zebrafish embryos and 25 μM dexamethasone prepared with standard dilution water;

[0161] Positive control group: containing 2-3 dpf zebrafish embryos, 25 μM dexamethasone prepared with standard dilution water, and 1 mg / mL chondroitin sulfate;

[0162] Composition test groups 1-15: corresponding to Examples 1-10 and Comparative Examples 1-4 respectively, containing 2-3 dpf zebrafish embryos, 25 μM dexamethasone prepared with standard dilution water, and 0.3 mg / mL of the composition prepared in each example or comparative example;

[0163] 2.2 Observation, photography, and data analysis

[0164] After treatment with the test substance at 28℃ for 3 days, zebrafish from each experimental group were observed and photographed under a fluorescence microscope. At least 10 zebrafish were randomly selected from each experimental group, fixed with 3% methylcellulose, and their craniofacial cartilage was photographed and its green fluorescence was collected. The fluorescence intensity (A) of the zebrafish craniofacial cartilage was quantitatively counted using ImageJ software. The statistical analysis results of this index were used to evaluate the repair capacity of the samples against dexamethasone-induced cartilage damage. Statistical results are expressed as mean ± SE.

[0165] Formula for calculating cartilage repair:

[0166] Cartilage regeneration rate (repair rate) C = (A1 - A0) / A0 × 100%,

[0167] In the formula, C represents the cartilage regeneration rate (repair rate)%;

[0168] A0—Fluorescence intensity of zebrafish craniofacial cartilage in the model control group (n=10).

[0169] A1—Fluorescence intensity of zebrafish craniofacial cartilage in each test group (n=10).

[0170] 3. Experimental Results

[0171] The EGFP fluorescence intensity and cartilage regeneration rate of each group of zebrafish are shown in Table 2 below:

[0172] Table 2. EGFP fluorescence intensity and cartilage regeneration rate of zebrafish in each group.

[0173]

[0174] As shown in Table 2, compared with the model control group, the positive control drug and the compositions prepared in Examples 1-10 of this application all achieved a cartilage regeneration rate of over 25% in the zebrafish cartilage injury model. In particular, the compositions prepared in Examples 1-4 and Example 7 of this application showed better cartilage damage resistance than the positive control drug, chondroitin A sodium salt, indicating that the compositions prepared in this application have a better cartilage regeneration promoting effect. The effects of Examples 1 and 5-6 show that the avocado powder and polygonatum powder prepared using the method of this application have a better cartilage repair effect. Changing the addition ratio of avocado powder and polygonatum powder in Examples 9 and 10 led to a decrease in the cartilage regeneration rate, indicating that a suitable ratio is more conducive to improving the cartilage repair effect.

[0175] Compared to Example 1, Comparative Example 1 did not add avocado powder, and Comparative Example 2 did not add polygonatum powder, so the synergistic anti-inflammatory effect of avocado powder and polygonatum powder could not be achieved, thus reducing the cartilage repair effect of the composition. In Comparative Example 3, replacing bovine bone collagen peptide powder with fish collagen peptide powder also led to a decrease in cartilage regeneration rate, indicating that the addition of bovine bone collagen peptide powder can make the cartilage repair effect of the composition better than that of fish collagen peptide powder. In Comparative Example 4, adding excessive bovine bone collagen peptide powder actually reduced the cartilage regeneration rate of the composition, indicating that an appropriate amount of bovine bone collagen peptide powder is more conducive to promoting cartilage repair.

[0176] Specifically, Figure 1 The images show the EGFP fluorescence intensity of zebrafish cartilage in the blank control group, model control group, positive control group, Example 1 group, Comparative Example 3 group, and Comparative Example 4 group.

[0177] Experiment 2: Anti-inflammatory effect of the composition

[0178] 1. Reagents, consumables and instruments

[0179] Reagents and consumables: RAW 264.7 macrophages (mouse mononuclear macrophage leukemia cells, purchased from Wuhan Pronosei Biotechnology Co., Ltd.), lipopolysaccharide and nitric oxide (NO) kits were purchased from Shanghai Beyotime Biotechnology Co., Ltd., and dexamethasone (Source Leaf BR, 99%, S17003-5g).

[0180] Instruments: Carbon dioxide incubator (Shanghai Zhichu Instrument Co., Ltd.), microplate reader (Molecular Device).

[0181] 2. Experimental Procedure

[0182] 2.1 Cell seeding: Macrophages in the logarithmic growth phase were gently scraped off with a spatula, mixed evenly, and the cell concentration per well was 1×10⁻⁶. 5 Inoculate 100 µL per well into a 96-well plate at a density of 1 / mL and incubate at 37°C in a 5% CO2 incubator for 24 h to allow it to adhere to the plate and grow.

[0183] 2.2 Experimental grouping: A certain mass of lipopolysaccharide (LPS) was dissolved in complete culture medium to make a final mass concentration of 1 mg / L. Then, different masses of the test samples were dissolved in complete culture medium containing LPS to prepare the required mass concentrations (0.05 mg / mL, 0.125 mg / mL, 0.25 mg / mL, 0.3 mg / mL). Five replicates were set for each mass concentration.

[0184] Blank control group: Complete culture medium without sample and LPS;

[0185] Model control group: Complete culture medium with only LPS added and no sample added;

[0186] Positive control group: Complete culture medium containing LPS and 10 μM dexamethasone;

[0187] Composition test groups 1-15: corresponding to Examples 1-10 and Comparative Examples 1-4, respectively, containing LPS and complete culture media of compositions prepared in each example or comparative example at different mass concentrations (200 µL of complete culture medium per well added to a 96-well plate).

[0188] 2.3 NO detection

[0189] After 24 h of treatment, 100 µL of cell supernatant from each well was transferred to another 96-well plate. The NO reagent kit was prepared according to the instructions: 50 µL of Griess A reagent was added, and the reaction was allowed to proceed for 10 min. Then, 50 µL of Griess B reagent was added, and the plate was incubated at room temperature for 10 min before measuring the absorbance at 540 nm. The mass of NO2 in each well was calculated based on the standard curve, which represents the mass of NO in the supernatant. The NO mass in the model control group was considered 100%, and all other groups were compared to this value to determine the amount of NO generated in the supernatant. The inhibition rate of each sample against NO was calculated using the formula: Inhibition rate (%) = (NO concentration in the model control group - NO concentration in the sample group) / NO concentration in the model control group × 100%.

[0190] The results of NO generation detection showed that when the composition concentration was in the range of 0.05 mg / mL to 0.3 mg / mL, the NO generation gradually decreased with the increase of composition concentration.

[0191] 2.4 Cell viability test

[0192] Macrophages in the logarithmic growth phase were gently scraped off with a spatula, mixed evenly, and the cell concentration per well was 1×10⁻⁶. 5 Cells were seeded at a rate of 100 µL per well in 96-well plates and incubated at 37°C with 5% CO2 for 24 h to allow for adherent growth. After 24 h, the culture medium was aspirated, the cells were washed once with 1×PBS, and samples were added. Specifically, 200 µL of different concentrations of the combined solution and dexamethasone were added according to the above-mentioned addition amounts. After 24 h of treatment, the culture medium was removed, and 100 μL of medium containing 10% CCK-8 solution was added. The plates were incubated for 1 h, and the absorbance at 450 nm was measured using a microplate reader. Simultaneously, the solvent without the sample was used as a control group. Cell viability was calculated using the following formula.

[0193] Cell viability (%) = (absorbance of sample group / absorbance of control group) × 100%.

[0194] Calculations showed that after adding different concentrations of the composition set in this experiment (0.125 mg / mL, 0.25 mg / mL, 0.5 mg / mL, 1 mg / mL, 2 mg / mL), the cell survival rate could reach over 95%, proving that the reduction in NO caused by the positive drug and the composition prepared in this application is due to the anti-inflammatory effect of the sample, rather than cytotoxicity.

[0195] Table 3 shows the NO generation and NO inhibition rate when the added composition concentration is 0.25 mg / mL and the positive control concentration is 10 μM.

[0196] Table 3 NO production and NO inhibition rate for each group

[0197]

[0198] As shown in Table 3, compared with the model control group, the positive control drug and the compositions prepared in Examples 1-10 of this application all achieved an inhibition rate of over 60% in the NO release of the LPS-induced macrophage inflammation model at a concentration of 0.25 mg / mL, indicating that the compositions prepared in this application have good anti-inflammatory effects. The effects of Examples 1 and 5-6 show that the avocado powder and polygonatum powder prepared using the method of this application have better anti-inflammatory effects. Changing the addition ratio of avocado powder and polygonatum powder in Examples 9-10 reduced the anti-inflammatory effect, indicating that a suitable ratio is more conducive to synergistic anti-inflammatory effects.

[0199] Compared with Example 1, the absence of avocado powder in Comparative Example 1 and the absence of Polygonatum powder in Comparative Example 2 both resulted in a significant reduction in the anti-inflammatory effect of the composition, indicating that avocado powder and Polygonatum powder have a synergistic anti-inflammatory effect. In Comparative Example 4, the addition of excessive bovine bone collagen peptide powder actually reduced the cartilage regeneration rate of the composition, indicating that adding an appropriate amount of bovine bone collagen peptide powder to the composition is more beneficial to improving the anti-inflammatory effect, while excessive amounts will promote the occurrence of inflammation and lead to a decrease in the anti-inflammatory effect.

[0200] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any form or substance. It should be noted that those skilled in the art can make various improvements and additions without departing from the method of the present invention, and these improvements and additions should also be considered within the scope of protection of the present invention. Any modifications, alterations, and equivalent changes made by those skilled in the art based on the above-disclosed technical content without departing from the spirit and scope of the present invention are equivalent embodiments of the present invention. Furthermore, any modifications, alterations, and evolutions made to the above embodiments based on the essential technology of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. A composition for promoting joint health, characterized in that, The composition comprises, by weight, the following components: 2-20 parts bovine bone collagen peptide powder, 2-15 parts N-acetylglucosamine, 2-15 parts calcium compound, 0.1-3 parts avocado powder, and 0.1-3 parts polygonatum powder; the mass ratio of avocado powder to polygonatum powder is 0.3-3:

1.

2. The composition according to claim 1, characterized in that, The calcium compound is selected from any one or two of calcium aspartate and calcium citrate.

3. The composition according to claim 1, characterized in that, The method for preparing avocado powder includes: peeling and pitting the avocado, slicing the pulp to obtain fresh avocado slices; pre-freezing and vacuum freeze-drying the fresh avocado slices to obtain freeze-dried avocado slices; and pulverizing and grinding the freeze-dried avocado slices to obtain avocado powder.

4. The composition according to claim 3, characterized in that, The temperature during the preparation of the fresh avocado slices is ≤10℃; and / or the thickness of the fresh avocado slices is ≤1cm; And / or, the pre-freezing includes freezing fresh avocado slices at -60°C to 80°C for 3-8 hours; And / or, the sublimation drying stage of the vacuum freeze-drying includes: maintaining a vacuum of 5-20 Pa, a temperature of -5 to 5°C, and a time of 15-25 h; And / or, the analytical drying stage of the vacuum freeze-drying includes: maintaining a vacuum of 1-10 Pa, a temperature of 15-25 °C, and a time of 5-15 h; And / or, the preparation method further includes cooling the freeze-dried avocado slices in liquid nitrogen before pulverizing; And / or, the avocado powder has a mesh size of 100-200 mesh.

5. The composition according to claim 1, characterized in that, The preparation method of the Polygonatum powder includes: taking fresh Polygonatum rhizomes and slicing them to obtain fresh Polygonatum slices; steaming, simmering and drying the fresh Polygonatum slices to obtain dried Polygonatum slices; and pulverizing the dried Polygonatum slices to obtain Polygonatum powder.

6. The composition according to claim 5, characterized in that, The thickness of the fresh Polygonatum slices is ≤1cm; And / or, the steaming time is 2-5 hours; And / or, the simmering time is 20-100 min; And / or, the drying temperature is 50-65°C; And / or, the drying time is 8-15 hours; And / or, the mesh size of the Polygonatum powder is 100-200 mesh.

7. A method for preparing a composition for promoting joint health as described in any one of claims 1 to 6, characterized in that, The method includes: mixing bovine bone collagen peptide powder, N-acetylglucosamine, calcium compounds, avocado powder and polygonatum powder evenly to obtain the composition.

8. The use of a joint health promoting composition as described in any one of claims 1 to 6 in the preparation of a product for promoting joint health.

9. A product that promotes joint health, characterized in that, The product includes the joint health-promoting composition as described in any one of claims 1 to 6 and food-grade excipients.

10. The product according to claim 9, characterized in that, The excipients are selected from any one or more of the following: fillers, flavoring agents, coloring agents, antioxidants, thickeners, stabilizers, emulsifiers, dispersants, anti-caking agents, and lubricants.

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