Preparation method and application of peptide calcium chelate

Abstract

The invention discloses a preparation method and application of a peptide calcium chelate, and belongs to the field of functional food or biological medicine. According to the invention, livestock and poultry processing by-products eggshells and shell membranes or animal cartilages are respectively used as a peptide source and a calcium source, release of polypeptides, calcium ions, phosphate radicals and amino chondroitin in raw materials is completed at the same time, phosphorylation of protein peptides and chelation of the protein peptides and calcium are synchronously realized, a one-pot method for preparing the peptide-calcium chelate is established, the chelation rate reaches 150mg / g (calcium / peptide), and the chelation rate reaches 150mg / g (calcium / peptide). Application of the composition in preparation of products beneficial to skin and hair growth; the method has the advantages of simple process, no use of strong acid, strong alkali and organic reagents, high safety, no use of hydrolase preparations, effective reduction of the product cost, simple production steps, short preparation period and important application potential. The method solves the problems of poor safety, complicated operation, high cost, long period and low flavor acceptability caused by the processes of preparing peptide by protease enzymolysis and chelating peptide calcium by using chemical reagents in the prior art.

Description

Technical Field

[0001] The invention belongs to the field of functional food or biomedicine, and specifically relates to a preparation method of a peptide calcium chelate and its use in improving skin and hair health. Background Art

[0002] Calcium is an important component of animal bones. It is a mineral element necessary for maintaining normal physiological activities such as metabolism and regulating endocrine. It is also the most obvious nutrient lacking in residents' diet. The national average daily calcium intake per person is 405 mg, which only reaches 49% of the RDA. It can be seen that calcium supplementation has become an urgent problem to be solved in my country's diet, and it is necessary to develop various calcium supplements. Therefore, effective calcium preparations are urgently needed to supplement and meet the additional calcium supplement needs. Compared with previous generations of calcium supplement products such as calcium carbonate and organic acid calcium, polypeptide chelated calcium is a cyclic complex formed by the combination of polypeptides and calcium ions in the form of coordination bonds or ionic bonds. Therefore, it has the advantages of stable structure, high bioavailability, and fast absorption rate, and can achieve the simultaneous supplementation of peptides and calcium, which is of great significance to human health.

[0003] Eggshell and eggshell membrane are also typical byproducts of egg product processing. Among them, eggshell accounts for about 11-13% of the total egg mass, contains mineral components such as calcium carbonate and calcium phosphate, and is a good source of calcium. Eggshell membrane is a thin and highly cross-linked fibrin membrane located between the egg white and the inner surface of the eggshell. It is mainly composed of keratin, collagen, glucuronic acid, hyaluronic acid and chondroitin sulfate, and is a good source of protein peptides. Animal bones are the waste of livestock and poultry food processing, but they also contain rich protein and hydroxyapatite, which are natural sources of calcium supplements and high-quality protein. For example, more than 200g of natural composite calcium can be extracted from 1kg of cattle bones. However, due to the special hydroxyapatite or fibrin structure of animal bones and eggshell membranes, the nutrients contained therein are not easy to be fully digested and absorbed by the human body. If these active ingredients in livestock and poultry processing byproducts such as discarded bones and eggshell membranes can be protected and utilized to prepare peptide calcium products that are easy for the human body to absorb and utilize, adding them as additives to food or medical health products will produce very good economic and social benefits.

[0004] At present, the preparation method of peptide chelated calcium mainly uses proteolytic enzyme to prepare peptide first, and then adds ionic calcium for chemical chelation. In order to improve the chelation rate of peptide calcium, it is usually necessary to phosphorylate the peptide before the chelation reaction to enhance its binding ability with calcium ions. For the dense structure of raw materials such as animal bones and eggshell membranes, the process of preparing hydrolyzed peptides by enzymatic reaction requires strict control of reaction conditions and pretreatment, and requires the combination of multiple enzymes to achieve effective hydrolysis of elastin and collagen, which undoubtedly increases production costs and complex operation processes. At the same time, the enzymatic process generally has problems such as the formation of bad product flavors and low sensory acceptance of consumers. In addition, bones and eggshells usually need to release calcium ions under strongly acidic conditions, while phosphorylation modification and peptide calcium chelation reactions need to be carried out under neutral conditions. The existing method adjusts the pH by NaOH, etc., which not only introduces additional sodium salts, but also causes hydroxide ions to combine with calcium ions to form precipitation, reducing the efficiency of the chelation reaction.

[0005] In summary, the current conventional preparation process of peptide chelated calcium mainly involves first using composite protein hydrolysis to extract and prepare the peptide, then using chemical reagents to phosphorylate the peptide to enhance its chelating ability, and finally adding an additional calcium source for chelation reaction. This results in the existing preparation process of peptide chelated calcium being relatively complex and time-consuming, requiring the use of strong acids and proteases, increasing costs and requiring the treatment of acid waste liquid. Therefore, the development of a new preparation technology for peptide calcium chelates with simple, efficient and low-cost processes is of great significance to the development of peptide calcium chelated calcium products and the promotion of human health. Summary of the invention

[0006] In order to solve the above technical problems, the present invention provides a preparation method and application of a peptide calcium chelate.

[0007] The present invention uses eggshells and shell membranes or animal cartilage, which are byproducts of livestock and poultry processing, as peptide sources and calcium sources, respectively, and simultaneously completes the release of polypeptides, calcium ions, phosphates and aminochondroitin in the raw materials, and synchronously realizes the phosphorylation of protein peptides and their chelation with calcium, thereby establishing a one-pot method for preparing peptide calcium chelates; and its application in the preparation of products that are beneficial to skin and hair growth. The present invention has a simple process, does not use strong acids, strong bases and organic reagents, has high safety, does not use hydrolytic enzyme preparations, can effectively reduce product costs, has simple production steps, and a short preparation cycle, and has important application potential.

[0008] A method for preparing a peptide calcium chelate is specifically completed by the following steps:

[0009] 1. Wash, dry and crush the animal bones of livestock and poultry or the egg shells with inner membranes in turn, and then pass through a 40-200 mesh sieve to obtain a powdered raw material;

[0010] 2. Mixing the powdered raw material and the organic acid solution, stirring the reaction for a period of time or stirring the reaction until no bubbles are generated, and then performing a first subcritical water treatment reaction, and then filtering the solution after the first subcritical water treatment reaction, collecting the filtrate and the filter residue to obtain a filtrate A and a filter residue A;

[0011] 3. Wash the filter residue A with water until it is neutral, then add distilled water, and then perform a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B;

[0012] 4. Combining filtrate A and filtrate B to obtain an aqueous solution of polypeptide chelated calcium; concentrating or drying the aqueous solution of polypeptide chelated calcium under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

[0013] The peptide calcium chelate described in step 4 contains free calcium, peptide chelated calcium, amino acid chelated calcium, glucosamine chondroitin, hyaluronic acid, desmosin and collagen peptide components.

[0014] The peptide calcium chelate is used in medicine, food or skin care products.

[0015] The peptide calcium chelate is used in the preparation of medicines and nutritionally fortified foods for supplementing calcium, lubricating and repairing cartilage damage, and promoting bone and hair growth.

[0016] The food is food for humans, food or feed for pets or livestock.

[0017] The present invention has the following beneficial effects:

[0018] (1) Based on the characteristic that water has an extremely low dielectric constant in a subcritical state, the present invention has the catalytic ability to replace the traditional acid hydrolysis or enzymatic hydrolysis method, and can hydrolyze collagen and elastin without adding additional catalysts, and release bone calcium and eggshell calcium; by controlling the temperature and time of the subcritical water reaction, the release and chelation of polypeptides, calcium ions, phosphates and aminochondroitin in animal cartilage and eggshell membranes are synchronously achieved, making full use of livestock and poultry processing by-products, increasing added value and reducing environmental pollution;

[0019] (2) The process of the present invention is simple and does not use hydrolase preparations, which can effectively reduce product costs; the entire method process only takes 5 to 6 hours to complete, which greatly shortens the time, improves efficiency, and reduces labor and time costs compared to the existing method that takes more than 5 days;

[0020] (3) The preparation process of the present invention does not use strong alkali and chemical reagents for treatment, which effectively avoids the formation of toxic products and bad flavors. The prepared polypeptide chelated calcium product has high safety and high consumer acceptance, and is suitable as a functional ingredient for food and medicine;

[0021] (4) The peptide calcium chelate product of the present invention is rich in chondroitin sulfate, glucosamine, hyaluronic acid, peptide chelated calcium, and amino acid chelated calcium, which plays an important role in regulating cartilage damage and improving calcium absorption, can better maintain skin health, and is beneficial to hair growth. DETAILED DESCRIPTION

[0022] Specific implementation method 1: This implementation method is a preparation method of a peptide calcium chelate, which is specifically completed according to the following steps:

[0023] 1. Wash, dry and crush the animal bones of livestock and poultry or the egg shells with inner membranes in turn, and then pass through a 40-200 mesh sieve to obtain a powdered raw material;

[0024] 2. Mixing the powdered raw material and the organic acid solution, stirring the reaction for a period of time or stirring the reaction until no bubbles are generated, and then performing a first subcritical water treatment reaction, and then filtering the solution after the first subcritical water treatment reaction, collecting the filtrate and the filter residue to obtain a filtrate A and a filter residue A;

[0025] 3. Wash the filter residue A with water until it is neutral, then add distilled water, and then perform a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B;

[0026] 4. Combining filtrate A and filtrate B to obtain an aqueous solution of polypeptide chelated calcium; concentrating or drying the aqueous solution of polypeptide chelated calcium under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

[0027] The chelation rate of the peptide calcium chelate prepared in this embodiment reaches 150 mg / g (calcium / peptide); the peptide calcium chelate solution (or powder) prepared by the present invention also contains glycosaminoglycans, hyaluronic acid, desmosin and other ingredients, has good water solubility, is easy to be absorbed by the human body, not only supplements the calcium element required by the human body, but also supplements collagen peptides and glycosaminoglycans that promote calcium absorption, and has the effects of repairing osteoarthritis, promoting the production of collagen and GAGs in the dermis of the skin, maintaining skin elasticity, improving bone and hair health, etc., and can be used to prepare drugs and / or food compositions, skin care product raw materials, food / feed (supplements) or food / feed additives, etc. that maintain and / or improve an organism to a good state.

[0028] Specific implementation method 2: This implementation method is different from specific implementation method 1 in that the livestock and poultry described in step 1 are pigs, cattle, sheep, chickens, ducks or geese. The other steps are the same as those in specific implementation method 1.

[0029] Specific implementation method 3: This implementation method is different from specific implementation method 1 or 2 in that the poultry eggs described in step 1 are chicken eggs, duck eggs or goose eggs. The other steps are the same as those of specific implementation method 1 or 2.

[0030] Specific embodiment 4: This embodiment differs from specific embodiments 1 to 3 in that: in step 1, animal bones of livestock and poultry or eggshell membranes of poultry eggs are washed with water for 3 to 5 times; the drying temperature in step 1 is 60°C to 80°C, and the drying time is 1h to 2h. The other steps are the same as those in specific embodiments 1 to 3.

[0031] Specific implementation method 5: This implementation method is different from specific implementation methods 1 to 4 in that the organic acid solution in step 2 is acetic acid solution or acetic acid solution; the mass fraction of the organic acid solution is 8% to 15%. The other steps are the same as specific implementation methods 1 to 4.

[0032] Specific embodiment 6: This embodiment differs from specific embodiments 1 to 5 in that the volume ratio of the powdered raw material to the organic acid solution in step 2 is 1:(1 to 20); the temperature of the first subcritical water treatment reaction in step 2 is 100°C to 180°C, and the reaction time is 10min to 60min. The other steps are the same as those in specific embodiments 1 to 5.

[0033] Specific embodiment 7: This embodiment differs from specific embodiments 1 to 6 in that the volume ratio of the filter residue A described in step 3 to the added distilled water is 1:(1 to 20); the temperature of the second subcritical water treatment reaction described in step 3 is 140°C to 240°C, and the reaction time is 10min to 60min. The other steps are the same as specific embodiments 1 to 6.

[0034] Specific embodiment 8: This embodiment differs from specific embodiments 1 to 7 in that the peptide calcium chelate described in step 4 contains free calcium, peptide chelated calcium, amino acid chelated calcium, glucosamine chondroitin, hyaluronic acid, desmosin and collagen peptide components. The other steps are the same as those of specific embodiments 1 to 7.

[0035] Specific implementation method 9: This implementation method is the application of the peptide calcium chelate in medicine, food or skin care products.

[0036] Specific embodiment 10: This embodiment differs from specific embodiments 1 to 9 in that the food is human food, food or feed for pets or livestock. The other steps are the same as those of specific embodiments 1 to 9.

[0037] The following examples are used to verify the beneficial effects of the present invention:

[0038] Example 1: A method for preparing a peptide calcium chelate, characterized in that the preparation method is specifically completed according to the following steps:

[0039] 1. Collect the discarded egg shells with inner membranes produced by liquid egg processing, wash them with clean water for 3 times, then dry them at 60°C, and then crush them with a grinder, and then pass them through a 100-mesh sieve to obtain a powdered raw material;

[0040] 2. Mixing the powdered raw material and the organic acid solution, stirring the mixture to react until no bubbles are generated, and then placing the mixture in a high-pressure reactor, introducing nitrogen to remove the air in the high-pressure reactor, and then heating the mixture to perform a first subcritical water treatment reaction, and then filtering the solution after the first subcritical water treatment reaction, collecting the filtrate and the filter residue to obtain a filtrate A and a filter residue A;

[0041] The organic acid solution in step 2 is an acetic acid solution with a mass fraction of 10%;

[0042] The volume ratio of the powdered raw material to the organic acid solution in step 2 is 1:10;

[0043] The temperature of the first subcritical water treatment reaction in step 2 is 150° C. and the reaction time is 30 min.

[0044] 3. Wash the filter residue A with water until it is neutral, then add distilled water, put it into a high-pressure reactor, introduce nitrogen to remove the air in the high-pressure reactor, heat it to carry out a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B;

[0045] The volume ratio of the filter residue A described in step 3 to the added distilled water is 1:10;

[0046] The temperature of the second subcritical water treatment reaction in step 3 is 180° C. and the reaction time is 30 min.

[0047] 4. Combine filtrate A and filtrate B to obtain an aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition; concentrate or dry the aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

[0048] Embodiment 2: The difference between this embodiment and embodiment 1 is that: the temperature of the first subcritical water treatment reaction described in step 2 is 180 ℃, and the reaction time is 20min; The temperature of the second subcritical water treatment reaction described in step 3 is 210 ℃, and the reaction time is 40min. Other steps and parameters are the same as embodiment 1.

[0049] Example 3: A method for preparing a peptide calcium chelate, characterized in that the preparation method is specifically completed according to the following steps:

[0050] 1. Collect the discarded duck egg shells with inner membranes produced by salted duck egg processing, wash them with clean water for 3 times, then dry them at 60°C, and then crush them with a crusher, and then pass them through a 40-200 mesh sieve to obtain a powdered raw material;

[0051] 2. Mixing the powdered raw material and the organic acid solution, stirring the mixture to react until no bubbles are generated, and then placing the mixture in a high-pressure reactor, introducing nitrogen to remove the air in the high-pressure reactor, and then heating the mixture to perform a first subcritical water treatment reaction, and then filtering the solution after the first subcritical water treatment reaction, collecting the filtrate and the filter residue to obtain a filtrate A and a filter residue A;

[0052] The organic acid solution in step 2 is an acetic acid solution with a mass fraction of 10%;

[0053] The volume ratio of the powdered raw material to the organic acid solution in step 2 is 1:20;

[0054] The temperature of the first subcritical water treatment reaction in step 2 is 150° C. and the reaction time is 30 min.

[0055] 3. Wash the filter residue A with water until it is neutral, then add distilled water, put it into a high-pressure reactor, introduce nitrogen to remove the air in the high-pressure reactor, heat it to carry out a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B;

[0056] The volume ratio of the filter residue A described in step 3 to the added distilled water is 1:10;

[0057] The temperature of the second subcritical water treatment reaction in step 3 is 180° C. and the reaction time is 30 min.

[0058] 4. Combine filtrate A and filtrate B to obtain an aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition; concentrate or dry the aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

[0059] Example 4: A method for preparing a peptide calcium chelate, characterized in that the preparation method is specifically completed according to the following steps:

[0060] 1. Collect the long bones of beef produced by beef processing, wash them with clean water for 3 times, then dry them at 60°C, and then crush them with a grinder, and then pass them through a 40-200 mesh sieve to obtain a powdered raw material;

[0061] 2. Mix the powdered raw material and the organic acid solution, stir and react for 3 hours, then put them into a high-pressure reactor, introduce nitrogen to remove the air in the high-pressure reactor, and then heat them to perform a first subcritical water treatment reaction, and then filter the solution after the first subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain filtrate A and filter residue A;

[0062] The organic acid solution in step 2 is an acetic acid solution with a mass fraction of 10%;

[0063] The volume ratio of the powdered raw material to the organic acid solution in step 2 is 1:15;

[0064] The temperature of the first subcritical water treatment reaction in step 2 is 150° C. and the reaction time is 40 min.

[0065] 3. Wash the filter residue A with water until it is neutral, then add distilled water, put it into a high-pressure reactor, introduce nitrogen to remove the air in the high-pressure reactor, heat it to carry out a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B;

[0066] The volume ratio of the filter residue A described in step 3 to the added distilled water is 1:10;

[0067] The temperature of the second subcritical water treatment reaction described in step 3 is 200° C. and the reaction time is 30 min;

[0068] 4. Combine filtrate A and filtrate B to obtain an aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition; concentrate or dry the aqueous solution of glucosamine chondroitin-polypeptide chelate calcium composition under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

[0069] Composition analysis:

[0070] The peptide calcium chelate powder prepared in Examples 1 to 4 was tested for component content. The total protein content in the product was determined by colorimetry, the calcium content was determined by atomic absorption, and the hydroxyproline content was determined by spectrophotometry described in GBT 9695.23-2003. The hydroxyproline content was multiplied by a conversion factor of 13% to obtain the collagen content. The hyaluronic acid content was determined by the Bitter-Muir carbazole method using glucuronic acid as a standard. The test results are shown in Table 1.

[0071] Table 1 Content of ingredients of peptide calcium chelates prepared in Examples 1 to 4

[0072]

[0073]

[0074] Completely different from the traditional single peptide calcium chelate preparation method, the peptide calcium chelate composition product prepared by the present invention has a chelation rate of more than 1:5 (i.e. 200 mg calcium ion / g protein peptide), and is naturally rich in glycosaminoglycan components such as hyaluronic acid and chondroitin sulfate, as well as collagen and elastin peptides, the structures of which are similar to the dermis of the skin.

[0075] Effect of promoting calcium absorption:

[0076] 70 healthy male Kunming mice, weighing 10g±2g, were purchased from the Experimental Animal Center of Xi'an Jiaotong University. After three days of adaptation to low-calcium feed, they were randomly divided into seven experimental groups, namely, blank control group I (distilled water), positive control group (commercially available calcium gluconate oral solution), low-dose group of Example 1, medium-dose group of Example 1, high-dose group of Example 1, Example 3, and Example 4. The recommended daily dose of commercially available calcium gluconate oral solution for humans is 30mL / d, which is expanded 10 times as the oral gavage dose for the test mice, and the calcium content of the samples of mice gavaged every day is calculated and prepared. Among them, the test mice in the blank control group I were orally gavaged with 0.2mL distilled water every day. The amount of calcium gavage in each group was blank control group (0 μg / 0.2 mL·d), positive control group (787.5 μg / 0.2 mL·d), low-dose group of Example 1 (393.75 μg / 0.2 mL·d), medium-dose group of Example 1 (787.5 μg / 0.2 mL·d), high-dose group of Example 1 (1575 μg / 0.2 mL·d), Example 3 (787.5 μg / 0.2 mL·d), and Example 4 (787.5 μg / 0.2 mL·d), with 10 mice in each group. Body weight statistical analysis was strictly performed, and formal experiments were performed after no significant difference was found by t test.

[0077] Each group of animals was housed in separate cages at room temperature of 25°C. Animals were fed with 6 g of low-calcium feed every day and deionized water. They were gavaged every morning, and their body weight and feed weight were weighed. Their body weight and feed intake were recorded, and the bedding was changed. The animals were raised for 6 weeks. Every week, mouse feces within 3 days were collected, labeled, dried and weighed in a 70°C oven, ground into powder for digestion, and the calcium content was determined. At the end of the 6th week, all mice were killed, blood was taken from the eyeballs, and serum was obtained by centrifugation. After the mice were killed by neck fracture, the left femur was taken, the meat attached to the bone was removed, and the mice were classified and marked by group. The femur of the mice was completely removed, the attached soft tissue and cartilage were removed, and the femur length was measured with a double-ring brand vernier caliper. According to the principle of buoyancy, the femur volume was measured and the bone density was calculated using a one-hundred-thousandth balance. The results are shown in Table 2. It can be clearly seen that the peptide calcium chelate has a significant effect on the growth of the femur of mice, especially the eggshell-derived peptide calcium chelate of Example 1 shows superiority over the blank control group and the positive control group under low-dose oral administration, suggesting that it has a certain effect on delaying bone aging.

[0078] Table 2 Changes in femur of mice after six weeks of testing

[0079]

[0080]

[0081] At the end of the experiment, the eyeballs were removed and blood was drawn. The subjects were fasted for half an hour before blood was drawn. The collected blood was centrifuged (3000r / min, 15min) to separate the serum, which was stored at -70℃ for the determination of various serum indicators. Serum indicators such as serum calcium, serum phosphorus, alkaline phosphatase and tartrate-resistant acid phosphatase were determined by conventional biochemical methods, and the kits were provided by Nanjing Jiancheng Bioengineering Institute.

[0082] Table 3 Effects on serum calcium, serum phosphorus, alkaline phosphatase and tartrate-resistant acid phosphatase in mice

[0083]

[0084]

[0085] In summary, from the changes in the weight, femur growth and various indicators in the serum of mice, we can see that the peptide calcium chelate composition provided by the present method has a certain positive effect on the weight, apparent absorption rate of calcium and various indicators in the serum of mice, which is consistent with the positive control calcium gluconate of this test, indicating that the peptide calcium chelate composition provided by the present method shows good absorption and utilization at low doses.

[0086] Evaluation of the effect of endogenous improvement of skin elasticity:

[0087] GAGs (glycosaminoglycans) and collagen and elastin are the three major components of the dermis. GAGs are filled in the mesh structure formed by the interweaving of collagen fibers and elastic fibers, and play an important role in maintaining moisture in the skin and regulating the assembly of collagen fibers. In order to verify the effect of the peptide calcium chelate composition provided by the present method on the production of glycosaminoglycan GAGs in the dermis, fibroblast growth factor was used as a positive control group, and the peptide calcium chelate composition provided by the present method prepared in Examples 1 to 4 was used for cell culture for 3-12 days. Method: Fibroblasts were cultured in a complete culture medium (DMEM / F12 containing 15% FBS by volume and 5% double antibiotic-penicillin + streptomycin mixture by volume) at 37°C and 5% CO2 incubator. When the cell content reached 80% to 90%, trypsin-EDTA enzyme digestion solution was used for digestion and passage. Take 1×10 fibroblasts in the logarithmic growth period 4 ~5×10 4, 50 μL / well was inoculated into a 96-well culture plate, 50 μL of peptide calcium chelates of different mass concentrations were added to each well, and the metabolic synthesis of glycosaminoglycans was observed.

[0088] Table 4 Effect of the peptide calcium chelate powder prepared in Examples 1 to 4 on the formation of glycosaminoglycans GAGs in the dermis

[0089]

[0090]

[0091] The results showed that the experimental group had a significant promoting effect on the metabolic synthesis of glycosaminoglycan GAGs, and the synthesis of GAGs increased by 43% after 3 days of intervention. It can be seen that the peptide calcium chelate composition provided by the method can endogenously promote the secretion of glycosaminoglycan GAGs by fibroblasts, thereby increasing the key anti-aging components of the dermis of the skin.

[0092] Pet Clinical Hair and Nail Test:

[0093] The experiment collected 60 cat subjects, and gave them peptide calcium chelate intervention at 500 mg / kg body weight per day. Questionnaires were conducted after 30 days and 30 days of discontinuation. After statistical analysis of the survey results, it was found that the peptide calcium chelate provided by the present invention has many positive effects on the glossiness, dryness, brittleness and other aspects of pet cat hair, and can also reduce hair loss and strengthen nails. And there was very good feedback after 30 days of consumption, and 86% of pet owners believed that the peptide calcium chelate provided by the present invention has a positive effect on the hair and nails of pet cats.

[0094] Table 5 Survey results on the effects of peptide calcium chelate powder prepared in Examples 1 to 4 on the hair health of pet cats

[0095]

[0096]

[0097] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for preparing a peptide calcium chelate, characterized in that The preparation method is specifically completed according to the following steps:

1. Wash, dry and crush the animal bones of livestock and poultry or the egg shells with inner membranes in turn, and then pass through a 40-200 mesh sieve to obtain a powdered raw material; 2. Mixing the powdered raw material and the organic acid solution, stirring the reaction for a period of time or stirring the reaction until no bubbles are generated, and then performing a first subcritical water treatment reaction, and then filtering the solution after the first subcritical water treatment reaction, collecting the filtrate and the filter residue to obtain a filtrate A and a filter residue A; 3. Wash the filter residue A with water until it is neutral, then add distilled water, and then perform a second subcritical water treatment reaction, filter the solution after the second subcritical water treatment reaction, collect the filtrate and the filter residue, and obtain a filtrate B and a filter residue B; 4. Combining filtrate A and filtrate B to obtain an aqueous solution of polypeptide chelated calcium; concentrating or drying the aqueous solution of polypeptide chelated calcium under reduced pressure to obtain a concentrated solution containing peptide calcium chelate or peptide calcium chelate powder.

2. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The livestock and poultry described in step 1 are pigs, cattle, sheep, chickens, ducks or geese.

3. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The poultry eggs described in step 1 are chicken eggs, duck eggs or goose eggs.

4. The method for preparing a peptide calcium chelate according to claim 1, characterized in that In step 1, animal bones of livestock and poultry or eggshell membranes of eggs are cleaned 3 to 5 times with water; the drying temperature in step 1 is 60° C. to 80° C., and the drying time is 1 to 2 hours.

5. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The organic acid solution described in step 2 is acetic acid solution or acetic acid solution; the mass fraction of the organic acid solution is 8% to 15%.

6. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The volume ratio of the powdered raw material to the organic acid solution in step 2 is 1:(1-20); the temperature of the first subcritical water treatment reaction in step 2 is 100° C.-180° C., and the reaction time is 10 min-60 min.

7. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The volume ratio of the filter residue A described in step 3 to the added distilled water is 1:(1-20); the temperature of the second subcritical water treatment reaction described in step 3 is 140° C.-240° C., and the reaction time is 10 min-60 min.

8. The method for preparing a peptide calcium chelate according to claim 1, characterized in that The peptide calcium chelate described in step 4 contains free calcium, peptide chelated calcium, amino acid chelated calcium, glucosamine chondroitin, hyaluronic acid, desmosin and collagen peptide components.

9. Use of the peptide calcium chelate prepared by the preparation method according to claim 1, characterized in that The peptide calcium chelate is used in medicine, food or skin care products.

10. The use of the peptide calcium chelate according to claim 9, characterized in that The food is food for humans, food or feed for pets or livestock.

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