Lactobacillus rhamnosus Lr-Ca011 capable of promoting calcium absorption, metagen, calcium product and nano-emulsification homogenizing preparation method of lactobacillus rhamnosus Lr-Ca011

Abstract

The invention provides a lactobacillus rhamnosus Lr-Ca011 capable of promoting calcium absorption, a metagen, a calcium product and a nano emulsification and homogenization preparation method of the lactobacillus rhamnosus Lr-Ca011, and belongs to the technical field of probiotics. The lactobacillus rhamnosus Lr-Ca011 is preserved in the China General Microbiological Culture Collection Center (CGMCC), the preservation number is CGMCC No.32037, and the lactobacillus rhamnosus Lr-Ca011 can promote the absorption of calcium. According to the invention, the lactobacillus rhamnosus Lr-Ca011 is used for fermentation to prepare the metagen, and the metagen also has the effect of promoting calcium absorption. In addition, a water phase and an oil phase containing the metabiotics are mixed and dispersed to obtain a crude emulsion, and then a calcium product is obtained by adopting a nano-emulsification homogenization method. According to the nano emulsifying and homogenizing method, the crude emulsion forms more stable nano particles, release of functional components can be effectively controlled, a calcium product has the characteristic of high absorptivity, and the bioavailability of calcium is improved.

Description

Technical Field

[0001] The invention relates to the technical field of probiotics, and in particular to a Lactobacillus rhamnosus Lr-Ca011, a postbiotic, a calcium product and a nano-emulsification homogenization preparation method thereof for promoting calcium absorption. Background Art

[0002] Calcium is an important nutrient in the human body and the most abundant inorganic element in the human body, with a total content of more than 1 kilogram. Most of the calcium (about 99%) is deposited in the bones in a solid state, and is constantly metabolized throughout our lives, dynamically changing the shape and hardness of the bones; a small amount of calcium (1%) exists in the blood and soft tissue cells in a free state, regulating various functions that maintain life activities, such as maintaining brain development, heart beating, muscle function, etc. Calcium deficiency can lead to a series of functional disorders in the body. Every year, various diseases caused by "calcium deficiency" will bring heavy economic pressure and mental burden to the patient's family and society. Therefore, how to improve the body's absorption of calcium is not only related to personal health, but also to the major issue of national health.

[0003] At present, the common methods to improve human calcium absorption are: (1) eating products rich in calcium, such as black sesame, seaweed, black beans, black fungus, etc.; (2) eating foods rich in vitamin D and lysine, such as egg yolks, pork liver, beans, etc.; (3) sunbathing to make human skin produce vitamin D, which is also called "sunshine vitamin". As long as the skin on the face and hands is exposed to sunlight for 15 minutes a day, the vitamin D produced can play a role in promoting human calcium absorption; (4) exercise can increase the body's blood circulation speed, speed up the blood to transport calcium into the bones, promote bone growth, increase bone mass, and stimulate the body's demand for calcium. With the development of science and technology, many studies have confirmed that supplementing probiotics can also play a role in promoting calcium absorption, but the effects of different probiotic strains on calcium absorption vary greatly, and even if the same strain is used, the raw materials or end products made using different processes will also lead to significant differences in their promotion effects.

[0004] Nanoparticles are particles with a diameter of less than 1 μm. Functional factors can be embedded in the interior of nanoparticles or adsorbed on the surface and delivered to the body. Nanoparticles have the advantages of small particle size and strong adhesion. They can not only enter and exit cells through a variety of different pathways (such as: (1) endocytosis, enhancing the ability of functional factors to pass through intestinal epithelial cells; (2) microfold cells (M cells) in Peyers patches in the ileum to take up particles; (3) tight junctions (TJs) between epithelial cells also form a natural epithelial barrier, affecting the permeability of functional factors; (4) transported into and out of cells through transport proteins, affecting the accumulation of functional factors in cells), but also increase the contact time between functional factors and intestinal mucosa, thereby increasing the absorption of functional factors and improving the bioavailability of oral preparations. Of course, nanoparticles are not the smaller the better. Excessively small particles are easily absorbed by cells and sometimes cause toxic effects on cells. If the particles are not small enough, the ability to promote the absorption of functional factors is not strong enough.

[0005] Therefore, providing a Lactobacillus rhamnosus that promotes calcium absorption and preparing postbiotics and calcium products through suitable nanoemulsification homogenization technology poses a challenge to current research. Summary of the invention

[0006] In view of the above problems, the present invention provides a Lactobacillus rhamnosus Lr-Ca011, a postbiotic, a calcium product and a nano-emulsification homogenization preparation method thereof for promoting calcium absorption.

[0007] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions: The present invention provides a method for promoting calcium absorption of Lactobacillus rhamnosus Lr-Ca011, which is classified and named as Lactobacillus rhamnosus Lacticaseibacillus rhamnosus , deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit date is September 23, 2024, and the deposit number is CGMCC No.32037.

[0008] The present invention also provides the use of the Lactobacillus rhamnosus Lr-Ca011 in preparing food or health products that promote calcium absorption.

[0009] Preferably, the food is a postbiotic and the health product is a calcium product.

[0010] The present invention also provides a postbiotic prepared by fermenting the Lactobacillus rhamnosus Lr-Ca011.

[0011] Preferably, the postbiotics include the following raw materials: Lactobacillus rhamnosus Lr-Ca011, quinoa, potato, pumpkin seed kernel, wolfberry, glucose monohydrate and purified water.

[0012] The present invention also provides a method for preparing the above-mentioned postbiotics, comprising: mixing quinoa, potato, pumpkin seed kernel, wolfberry, monohydrate glucose and purified water in a weight ratio of (15-43):(5-35):(10-30):(8-12):(10-30):(200-250), inoculating Lactobacillus rhamnosus Lr-Ca011 for fermentation, high-pressure homogenization, drying and crushing to obtain the postbiotics.

[0013] Preferably, the high pressure homogenization is performed at a pressure of 1000-1500 bar, for a time of 5-15 min, and for 1-4 times.

[0014] The present invention also provides a calcium product prepared according to the above-mentioned postbiotics, comprising the following raw materials in parts by weight: 40-50 parts of oil, 25-35 parts of calcium citrate, 3-7 parts of stabilizer, 2-5 parts of postbiotics, 1-3 parts of emulsifier and 10-20 parts of purified water.

[0015] Preferably, the oil is one or more of conjugated linoleic acid glyceride, medium chain triglyceride, caprylic decanoic acid glyceride, cocoa butter, anhydrous butter, walnut oil, and linseed oil; the emulsifier is one or more of whey protein, casein, soy protein isolate, pasteurized egg yolk liquid, and lecithin; the stabilizer is one or more of pectin, L-arabic gum, carrageenan, xanthan gum, gelatin, and konjac flour.

[0016] The present invention also provides a nano-emulsification homogenization preparation method for the above-mentioned calcium product, comprising: mixing oil, emulsifier, postbiotics and calcium citrate to obtain an oil phase; mixing a stabilizer with purified water to obtain an aqueous phase; mixing and dispersing the oil phase and the aqueous phase, and then high-pressure homogenizing to obtain the calcium product; the pressure of the high-pressure homogenization is 100-160MPa, the time is 10-20min, and the number of times is 1-4 times.

[0017] Compared with the prior art, the present invention has the following beneficial effects: The invention provides a Lactobacillus rhamnosus Lr-Ca011, which is preserved in the General Microbiological Center of China Microbiological Culture Collection Administration Committee with a preservation number of CGMCC No.32037 and can promote calcium absorption.

[0018] The present invention also provides a postbiotic obtained by fermenting Lactobacillus rhamnosus Lr-Ca011, which has a promoting effect on calcium absorption. ® 6162、CICC ® 6158、CICC ® 22173、CICC ® 6139、CICC ® 6241、CICC ®6135, etc.) fermentation, the postbiotics of the present invention have a better effect of promoting calcium absorption.

[0019] The present invention adopts nano-emulsification homogenization technology to prepare calcium products, after the water phase and the oil phase containing the postbiotics of Lactobacillus rhamnosus Lr-Ca011 are evenly mixed, a crude emulsion is obtained by dispersion, and then the droplets in the crude emulsion are fully broken into more stable nanoparticles again by high pressure to form a nano-emulsion, which can greatly reduce the use of surfactants, effectively reduce the particle size of the emulsion, enhance the stability of the emulsion system, and produce a pleasant mouthfeel. Nano-emulsion can effectively prevent the separation of emulsions due to its smaller particle size, has the function of quickly transporting bioactive ingredients to the cell membrane, can well overcome the shortcomings of poor water solubility, low stability and easy oxidation, effectively control the release of functional ingredients, make calcium products have the characteristics of high absorption rate, and improve the bioavailability of calcium. The test results show that the calcium absorption enhancement effect of the calcium product of the present invention is 3.18 times that of the calcium carbonate tablets with the same calcium content.

[0020] The Lactobacillus rhamnosus Lr-Ca011 of the present invention is classified and named as Lactobacillus rhamnosus Lacticaseibacillus rhamnosus , deposited in the General Microbiological Culture Collection Center of China Microorganism Culture Collection Administration, with the deposit number CGMCC No.CGMCC No.32037, the deposit date is September 23, 2024, and the deposit address is the Institute of Microbiology, Chinese Academy of Sciences. DETAILED DESCRIPTION

[0021] The present invention provides a method for promoting calcium absorption of Lactobacillus rhamnosus Lr-Ca011, which is classified and named as Lactobacillus rhamnosus Lacticaseibacillus rhamnosus , deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit date is September 23, 2024, and the deposit number is CGMCC No.32037.

[0022] The Lactobacillus rhamnosus Lr-Ca011 of the present invention is derived from the intestine of a healthy adult.

[0023] The Lactobacillus rhamnosus Lr-Ca011 of the present invention can effectively improve the absorption rate and retention rate of calcium by the human body, and has a very significant effect on promoting human bone health.

[0024] The present invention also provides the use of the Lactobacillus rhamnosus Lr-Ca011 in preparing food or health products that promote calcium absorption.

[0025] The food of the present invention is preferably a postbiotic, and the health product is preferably a calcium product.

[0026] The present invention also provides a postbiotic containing the Lactobacillus rhamnosus Lr-Ca011.

[0027] The postbiotics of the present invention include the following raw materials: Lactobacillus rhamnosus Lr-Ca011, quinoa, potato, pumpkin seed kernel, wolfberry, glucose monohydrate and purified water.

[0028] The present invention also provides a method for preparing the above-mentioned postbiotics, comprising: mixing quinoa, potato, pumpkin seed kernel, wolfberry, monohydrate glucose and purified water in a weight ratio of (15-43):(5-35):(10-30):(8-12):(10-30):(200-250), inoculating Lactobacillus rhamnosus Lr-Ca011 for fermentation, high-pressure homogenization, drying and crushing to obtain the postbiotics.

[0029] The pressure of the high-pressure homogenization of the present invention is preferably 1000-1500 bar, more preferably 1200 bar, the time is preferably 5-15 min, more preferably 10 min, the number of times is preferably 1-4 times, more preferably 2 times.

[0030] The quinoa protein of the present invention is rich in content and balanced in proportion, contains all natural amino acids, and is rich in minerals, vitamins and various functional ingredients (such as flavonoids, polyphenols, quercetin, rutin, etc.). It is a complete nutritional food that can meet the basic nutritional needs of humans and is known as "nutritional gold". Among them, the high content of lysine can promote the absorption of calcium. Potatoes are rich in nutrients such as B vitamins, a large amount of trace elements, high-quality protein, amino acids, fats and high-quality starch. Among them, the protein composition is closest to animal protein, and the protein and vitamin C content are both 10 times that of apples. It has strong antioxidant properties and can not only provide nutrition for the growth of probiotics and promote the proliferation of probiotics, but also protect the strains during freeze-drying and improve the survival rate of the strains. Pumpkin seed kernel is a plant seed kernel mainly composed of fat and protein, which contains a variety of nutrients. It not only contains 17 kinds of amino acids, but also contains zinc, selenium, manganese, copper, iron, calcium, phosphorus, magnesium, potassium, sodium and other minerals. Among these elements, the higher content is Zn, Fe, K, Ca, which can promote the growth, development and intelligence of the human body and enhance the immune function. Lycium barbarum polysaccharide in wolfberry is one of the main active ingredients, which has a prebiotic-like effect, can quickly promote the proliferation of probiotics, and induce the production of short-chain fatty acids. The monosaccharides that constitute Lycium barbarum polysaccharide are mainly glucose, rhamnose, galactose, arabinose, mannose, fucose, and xylose. Among them, the content of glucose, galactose, and arabinose is relatively high. In addition, there are sucrose, raffinose, ethyl-β-D-pyranoglucoside, 6-phosphogluconate, etc.

[0031] The Lactobacillus rhamnosus Lr-Ca011 of the present invention is fully fermented using a novel culture medium when preparing postbiotics. The culture medium not only contains a variety of nutrients such as rich high-quality proteins, amino acids, vitamins, minerals, lipids, carbohydrates, etc., which can provide sufficient nutrition for the growth and metabolism of Lr-Ca011, but also contains natural ingredients like prebiotics, which can promote the large-scale proliferation of Lr-Ca011. In addition, the culture medium also has a variety of functional sugars, and no additional freeze-drying protective agent is needed to reduce the irreversible damage caused by ice crystals to the bacterial body during freeze-drying, so that the strain is safely in a dormant state and protects the activity of the strain. The obtained bacterial powder is rich in active Lr-Ca011 and its metabolites, which is conducive to promoting calcium absorption after human consumption.

[0032] The present invention also provides a calcium product containing the above-mentioned postbiotics, comprising the following raw materials in parts by weight: 40-50 parts of oil, 25-35 parts of calcium citrate, 3-7 parts of stabilizer, 2-5 parts of postbiotics, 1-3 parts of emulsifier and 10-20 parts of purified water.

[0033] The oil of the present invention is preferably one or more of conjugated linoleic acid glyceride, medium-chain triglyceride, caprylic decanoic acid glyceride, cocoa butter, anhydrous butter, walnut oil, and linseed oil, and is more preferably medium-chain triglyceride; the emulsifier is one or more of whey protein, casein, soy protein isolate, pasteurized egg yolk liquid, and lecithin, and is more preferably whey protein; the stabilizer is preferably one or more of pectin, L-arabic gum, carrageenan, xanthan gum, gelatin, and konjac flour, and is more preferably xanthan gum.

[0034] The present invention also provides a nanoemulsified homogenized preparation method for the above-mentioned calcium product, comprising: mixing oil, emulsifier, postbiotics and calcium citrate to obtain an oil phase; mixing a stabilizer with purified water to obtain an aqueous phase; mixing and dispersing the oil phase and the aqueous phase, and high-pressure homogenizing to obtain the calcium product; the pressure of the high-pressure homogenization is 100-160MPa, the time is 10-20min, and the number of times is 1-4 times. The high-pressure homogenization pressure is preferably 130MPa, the time is preferably 13min, and the number of times is preferably 3 times.

[0035] The nano-emulsification homogenization technology of the present invention first prepares the water phase and the oil phase according to a certain ratio and mixes them evenly, performs preliminary dispersion by a high-speed disperser to obtain a crude emulsion, and then uses high pressure to fully break the droplets in the crude emulsion into more stable nanoparticles again to form a nano-emulsion, which can greatly reduce the use of surfactants, effectively reduce the particle size of the emulsion, enhance the stability of the emulsion system, and produce a pleasant taste. Nano-emulsion can effectively prevent the separation of the emulsion due to its smaller particle size, has the function of quickly transporting bioactive ingredients to the cell membrane, can well overcome the shortcomings of poor water solubility, low stability and easy oxidation, effectively control the release of functional ingredients, make the calcium product have the characteristics of high absorption rate, and improve the bioavailability of calcium.

[0036] In the present invention, unless otherwise specified, all raw material components are commercially available products well known to those skilled in the art.

[0037] The technical solutions in the present invention will be described clearly and completely below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0038] In the following embodiments, unless otherwise specified, all of them are conventional methods.

[0039] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0040] Examples 1-6 Preparation of Lactobacillus rhamnosus Lr-Ca011 bacterial powder 1. Choose dry, mildew-free, and insect-free quinoa and pumpkin seeds, wash and drain them; choose fresh, unsprouted potatoes, peel them, and cut them into pieces; choose wolfberries and glucose monohydrate of qualified quality.

[0041] 2. Liquid 1: Weigh several parts of quinoa, potato, pumpkin seed kernel, wolfberry, and glucose monohydrate as shown in Table 1, mix well, put into a wall breaking machine, add 235 parts by weight of purified water, break the wall at 35000r / min for 3min, filter with a 80-mesh filter, take the filtrate, sterilize at 121℃ for 20min, and cool to 35℃.

[0042] 3. Bacterial liquid: Take out the freeze-dried storage tube of Lactobacillus rhamnosus Lr-Ca011, let it stand at room temperature for 1 hour, draw 0.6mL of physiological saline, add it to the freeze-dried storage tube, shake until the contents are dissolved, draw 10μL, spread it on MRS solid medium, culture at 35℃ for 36h, pick 2 loops of colonies with an inoculation loop, inoculate them on 10mL MRS liquid medium, culture at 35℃ for 36h, and adjust the bacterial liquid concentration to 5×10 9 CFU / mL is obtained.

[0043] 4. Add 10% of the bacterial solution (volume percentage) into liquid 1, mix well, culture at 35°C for 72 hours, and then freeze-dry using a vacuum freeze dryer. Grind the resulting product and pass it through an 80-mesh sieve. The collected powder is the bacterial powder.

[0044] Table 1 Raw material configuration of Example 1-6 Feed solution 1 (parts by weight)

[0045] Test Example 1 Calcium absorption test 1. Experimental animals: SPF healthy male ICR mice, weighing 40g-50g.

[0046] 2. The mice were randomly divided into 13 groups, with 10 mice in each group, marked as Group A-Group M. Group B-Group M were all given intraperitoneal injection of 120 mg / (kg d) of D-galactose for 56 consecutive days to establish an aging osteoporosis mouse model. Group A was given intraperitoneal injection of the same dose of normal saline.

[0047] As people age, the intestinal absorption rate of calcium usually shows a gradual decline, leading to the appearance of osteoporosis symptoms. D-galactose-induced osteoporosis in male mice is a more practical animal model for studying aging-related osteoporosis. After continuous injection of D-galactose, excessive oxygen free radicals are produced in the male mice, destroying collagen, leading to oxidative damage to various cells, affecting the normal metabolism of bone cells, causing mineral loss, and accelerating the aging of male mice.

[0048] 3. After modeling, mice in each group were gavaged with a volume of 1 mL / 100 g bw for 28 consecutive days. The dosing conditions are shown in Table 2. All gavage samples were dissolved with sterile saline, and the inorganic calcium source for gavage was CaCl2. During the dosing period, mice were fed standard feed and drinking water was deionized water.

[0049] Table 2 Grouping and dosing of mice

[0050] 4. On the 15th day of administration, all mice were transferred to a special metabolic cage. On the 26th, 27th and 28th days, the feces and urine of each test mouse were collected. The calcium content in the ingested feed and the collected feces and urine was determined by flame atomic absorption spectrometry in accordance with GB 5009.92-2016 National Food Safety Standard Determination of Calcium in Foods. The average values ​​of calcium absorption rate (%) and retention rate (%) were calculated according to the following formula. The results are shown in Table 3.

[0051] (1) Calcium intake = calcium in feed + calcium administered intragastrically; (2) Calcium absorption = calcium intake – fecal calcium; (3) Calcium absorption rate = absorbed calcium / intake calcium × 100%; (4) Calcium retention = calcium intake – calcium in feces – calcium in urine; (5) Retention rate = stored calcium / calcium intake.

[0052] Table 3 Calcium absorption and retention in mice

[0053] As shown in Table 3, compared with Group A, the calcium absorption rate and retention rate of mice in Group B were significantly reduced, indicating that the aging mouse model was successfully established and the calcium absorption capacity of the intestine was significantly reduced. Compared with Group B, the calcium absorption rate and retention rate of mice in Groups C-H increased to varying degrees, indicating that the Lactobacillus rhamnosus Lr-Ca011 powder promoted calcium absorption. Due to the difference in the culture medium formula, the powder had different effects on promoting calcium absorption. Among them, the calcium absorption rate and retention rate of mice in Group H were the highest, indicating that the powder of Example 6 promoted calcium absorption best. Therefore, Example 6 was selected as the best formula of the culture medium, and the obtained Lactobacillus rhamnosus Lr-Ca011 powder could promote calcium absorption.

[0054] Embodiment 7-11 Preparation of postbiotics 1. Choose dry, mildew-free, and insect-free quinoa and pumpkin seeds, wash and drain them; choose fresh, unsprouted potatoes, peel them, and cut them into pieces; choose wolfberries and glucose monohydrate of qualified quality.

[0055] 2. Liquid 1: Weigh 33 parts by weight of quinoa, 20 parts by weight of potatoes, 17 parts by weight of pumpkin seed kernels, 10 parts by weight of wolfberry, and 20 parts by weight of glucose monohydrate, mix well, place in a wall breaking machine, add 235 parts by weight of purified water, break the wall at 35000r / min for 3min, filter with a 80-mesh filter, take the filtrate, sterilize at 121℃ for 20min, and cool to 35℃.

[0056] 3. Bacterial liquid: Take out the freeze-dried storage tube of Lactobacillus rhamnosus Lr-Ca011, let it stand at room temperature for 1 hour, draw 0.6mL of physiological saline, add it to the freeze-dried storage tube, shake until the contents are dissolved, draw 10μL, spread it on MRS solid medium, culture at 35℃ for 36h, pick 2 loops of colonies with an inoculation loop, inoculate them on 10mL MRS liquid medium, culture at 35℃ for 36h, and adjust the bacterial liquid concentration to 5×10 9 CFU / mL is obtained.

[0057] 4. Add 10% of the bacterial liquid (volume percentage) to the feed liquid 1, mix well, and culture at 35°C for 72 hours. The obtained product is homogenized by a high-pressure homogenizer. The homogenization condition is 1200 bar and the time is 10 minutes. The homogenization is repeated. The number of homogenizations is shown in Table 4. Then, the product is freeze-dried by a vacuum freeze dryer. The obtained product is pulverized by a 80-mesh sieve, and the collected powder is the postbiotic.

[0058] Table 4 Homogenization times of postbiotics in different embodiments

[0059] Test Example 2 1. The mice modeled in the experiment 1 (SPF healthy male ICR mice, weighing 40g-50g) were divided into 5 groups, 10 mice in each group, and the mice in each group were gavaged with a volume of 1mL / 100g bw for 28 consecutive days. The dosing conditions are shown in Table 5. All gavage samples were dissolved with sterile saline, and the inorganic calcium source for gavage was CaCl2. During the dosing period, the mice were fed standard feed and the drinking water was deionized water.

[0060] Table 5 Grouping and dosing of mice

[0061] 2. On the 15th day after administration, all mice were transferred to a special metabolic cage. On the 26th, 27th and 28th days, the feces and urine of each test mouse were collected. The calcium content in the ingested feed and the collected feces and urine was determined by flame atomic absorption spectrometry in accordance with GB 5009.92-2016 National Food Safety Standard Determination of Calcium in Food. The average values ​​of calcium absorption rate (%) and retention rate (%) were calculated. The results are shown in Table 6.

[0062] Table 6 Calcium absorption and retention in mice

[0063] From the data in Table 3 and Table 6, it can be seen that compared with Group A, the calcium absorption rate and retention rate of mice in Groups I-L increased to varying degrees, indicating that postbiotics have a promoting effect on calcium absorption. The calcium absorption rate and retention rate of mice in Groups I-K continued to increase, indicating that when the number of homogenizations is ≤2 times, the more times the homogenizations are performed, the better the effect of promoting calcium absorption. The calcium absorption rate and retention rate of mice in Groups K-M are relatively close, indicating that when the number of homogenizations is greater than 2 times, the effect of promoting calcium absorption is also significantly increased when the number of homogenizations increases. That is, when homogenized twice, the effect of postbiotics on promoting calcium absorption is basically saturated, so 2 times of homogenization is selected as the optimal number of postbiotic homogenizations.

[0064] Comparative Examples 1-6 Six different strains of Lactobacillus rhamnosus were used to prepare postbiotics according to the method of Example 9 (compared with Example 9, only the strains of Lactobacillus rhamnosus were different). The six different strains of Lactobacillus rhamnosus are shown in Table 7.

[0065] Table 7 Rhamnosus lactobacillus strains of comparative examples 1-6

[0066] The Lactobacillus rhamnosus strains used in Comparative Examples 1-6 were all from the China Industrial Microbiological Culture Collection Administration Center.

[0067] Test Example 3 1. Model mice (SPF healthy male ICR mice, weighing 40g-50g) were made using the method of Experiment 1 and divided into 7 groups, 10 mice in each group. Each group of mice was given drugs by gavage, with a gavage volume of 1mL / 100g bw for 28 consecutive days. The drug administration is shown in Table 8. All gavage samples were dissolved with sterile saline, and the inorganic calcium source for gavage was CaCl2. During the drug administration period, the mice were fed standard feed and the drinking water was deionized water.

[0068] Table 8 Grouping and dosing of mice

[0069] 2. On the 15th day after administration, all mice were transferred to a special metabolic cage. On the 26th, 27th and 28th days, the feces and urine of each test mouse were collected. The calcium content in the ingested feed and the collected feces and urine was determined by flame atomic absorption spectrometry in accordance with GB 5009.92-2016 National Food Safety Standard Determination of Calcium in Foods. The average values ​​of calcium absorption rate (%) and retention rate (%) were calculated. The results are shown in Table 9.

[0070] Table 9 Calcium absorption and retention in mice

[0071] It can be seen from the data in Table 9 that the postbiotics prepared by using Lactobacillus rhamnosus from different sources have different effects on calcium absorption. Among them, the postbiotics obtained by the preparation method of Example 6 of the present invention can significantly improve the calcium absorption rate and retention rate of mice. Obviously, the Lactobacillus rhamnosus Lr-Ca011 of the present invention has a better effect of promoting calcium absorption than other Lactobacillus rhamnosus of the same category.

[0072] Examples 12-16 Preparation of calcium products 1. Select soybean oil, medium-chain triglycerides, xanthan gum, postbiotics, and whey protein of qualified quality; select calcium citrate of qualified quality.

[0073] 2. Coarse emulsion: Weigh 45.5 parts by weight of medium-chain triglycerides, add 1.5 parts by weight of whey protein while stirring, continue stirring for 10 minutes, then add 3 parts by weight of the postbiotics of Example 9, continue stirring for 10 minutes, then add 30 parts by weight of calcium citrate, continue stirring for 3 hours, filter through a 100-mesh filter, collect the filtrate, and the obtained filtrate is the oil phase; add 5 parts by weight of xanthan gum to 100°C and 15 parts by weight of purified water, stir and dissolve, and the obtained solution is the water phase. Mix the oil phase and the water phase evenly, shear at 8000 rpm for 3 minutes with a high-speed disperser, and the obtained product is the coarse emulsion.

[0074] 3. Use a high-pressure homogenizer to homogenize the crude emulsion at a temperature of 25°C. The homogenization pressure is as shown in Table 10. Homogenize 3 times, each time for 13 minutes. The obtained product is a calcium product.

[0075] Table 10 Homogenization pressure of calcium products in different embodiments

[0076] Test Example 4 The particle size of the calcium products of Examples 12 to 16 was measured using a nanoparticle size and potential analyzer. Each product was measured 3 times and the average value was calculated. The results are shown in Table 11.

[0077] Table 11 Effect of pressure on nanoparticle size

[0078] It can be seen from Table 11 that pressure has a significant effect on the particle size of the calcium product, but it does not decrease as the pressure increases. The particle size of the nanoemulsion shows a trend of first decreasing and then increasing as the pressure continues to increase. The particle size of the calcium product in Example 15 is the smallest, that is, when the pressure is 130 MPa, the calcium product reaches the minimum particle size.

[0079] Test Example 5 Crowd testing 1. Volunteers aged 12-17 were recruited as subjects to participate in the trial and randomly divided into 5 groups, denoted as Group A-Group E, with 8 people in each group. Calcium was supplemented in the form of calcium products. The groups and consumption conditions are shown in Table 12. All subjects met the following conditions: voluntary participation, good health, and no liver, kidney disease or skeletal system disease that affects bone metabolism.

[0080] Table 12 Subject groups and sample consumption

[0081] 2. Consume 3 times a day, 1 hour after meals, for 10 days. The subjects are provided with centralized accommodation and cannot consume any food or drink other than the food provided by the experiment. The subjects are first given 3 days to adapt to the experimental diet, and then the experimental diet is consumed for 7 days with a 3-day diet cycle. The diet of the subjects for 10 days (including meals, snacks and drinking water) is collected.

[0082] 3. Take 2 capsules of Kahong capsules (200 mg) before breakfast on the 4th and 11th days to mark the stool. Collect the stool of each subject from the time the stool turns red until the red color appears again on the 11th day. The stool is collected once every 24 hours.

[0083] 4. Collect the subject's urine for 24 hours every day, add 1% hydrochloric acid solution, and mix well.

[0084] 5. According to the flame atomic absorption spectrometry method in "GB 5009.92-2016 National Food Safety Standard Determination of Calcium in Food", the calcium content in the diet and collected feces and urine was determined, and the average values ​​of calcium absorption rate (%) and retention rate (%) were calculated. The results are shown in Table 13.

[0085] Table 13 Calcium absorption and retention of subjects

[0086] As shown in Table 13, among the groups A to E, the calcium absorption rate and retention rate of the subjects in group C are the highest, indicating that the calcium product of Example 14 has the best effect in promoting calcium absorption. Combined with the experimental results of Table 11, the particle size of group C is larger than that of group D, indicating that the smaller the particle size, the better the calcium absorption effect. The calcium product with the best particle size has the best effect in promoting calcium absorption. Therefore, Example 14 is selected as the optimal pressure for homogenizing the calcium product, and the obtained calcium product has a high absorption rate for calcium element.

[0087] Test Example 6 Calcium absorption rate test in rats 1. Sample preparation: 70 parts by weight of sorbitol and 30 parts by weight of calcium carbonate were mixed and made into tablet candy by direct compression. The tablet candy was dissolved in purified water to make a solution with a concentration of 50%, which was recorded as solution A.

[0088] 2. Experimental animals: SD rats, female, 10-12 weeks old.

[0089] 3. After 7 days of adaptive feeding, the 18 rats were divided into three experimental groups, named model control group, experimental group 1 and experimental group 2, with 6 rats in each group.

[0090] 4. Osteoporosis modeling of rats: anesthesia, ventral skin preparation, hair removal, disinfection, use sterile surgical scissors to make an incision of about 2 cm in length in the midline of the abdomen, separate and expose the ovaries, use sutures to ligate and remove both ovaries. Put the uterus back into the abdominal cavity, suture the muscles and skin layer by layer, disinfect the wound with iodine, place it on a heating pad and wait for the animal to wake up and feed normally.

[0091] 5. After the modeling surgery, medication was started every other day. Experimental group 1 was gavaged with 1 mL of the calcium product of Example 14 every day, experimental group 2 was gavaged with 2 mL of solution A every day, and the model control group was gavaged with 1 mL of normal saline every day for 4 consecutive weeks.

[0092] 6. After the last administration, rats in each group were euthanized, femurs were removed, muscles and connective tissues around the femurs were removed, carbonized on the furnace, and then placed in a muffle furnace at 800°C for 2 hours. After cooling to room temperature, the ashes were weighed with an analytical balance after being placed in a desiccator. 0.1g of bone ash was weighed for each rat, 10mL of 5% nitric acid solution was added and transferred to a 25mL colorimetric tube. The solution was placed at room temperature for 48 hours to digest, and the solution was colorless and transparent. The digestion solution was removed, the absorbance was measured, and a reagent blank was made at the same time. The calcium content in the sample was calculated from the standard curve, and the average value of each group was calculated. Two decimal places were retained and rounded off. The results are shown in Table 14.

[0093] Table 14 Bone calcium absorption of rats in each group

[0094] It can be seen from Table 14 that the ratio of the improvement rate between experimental group 1 and experimental group 2 is about 3.18, indicating that the calcium product of Example 14 has the effect of significantly promoting bone calcium absorption, and its improvement effect is 3.18 times that of calcium carbonate tablets with the same calcium content.

[0095] 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 Lactobacillus rhamnosus Lr-Ca011 for promoting calcium absorption, characterized in that: The classification of the Lactobacillus rhamnosus Lr-Ca011 is named as Lactobacillus rhamnosus Lacticaseibacillus rhamnosus , deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit date is September 23, 2024, and the deposit number is CGMCCNo.32037.

2. Use of the Lactobacillus rhamnosus Lr-Ca011 according to claim 1 in preparing foods that promote calcium absorption.

3. The use according to claim 2, characterized in that: The food products include postbiotics and calcium products.

4. A postbiotic prepared by fermenting Lactobacillus rhamnosus Lr-Ca011 according to claim 1.

5. The postbiotic according to claim 4, characterized in that The invention comprises the following raw materials: Lactobacillus rhamnosus Lr-Ca011, quinoa, potato, pumpkin seed kernel, wolfberry, dextrose monohydrate and purified water.

6. A method for preparing the postbiotics according to claim 4 or 5, characterized in that: include: Quinoa, potato, pumpkin seed kernel, wolfberry, glucose monohydrate and purified water are mixed in a weight ratio of (15-43):(5-35):(10-30):(8-12):(10-30):(200-250), inoculated with Lactobacillus rhamnosus Lr-Ca011 for fermentation, homogenized under high pressure, dried and crushed to obtain the postbiotics.

7. The preparation method according to claim 6, characterized in that: The high pressure homogenization is performed at a pressure of 1000-1500 bar, for a time of 5-15 minutes, and for 1-4 times.

8. A calcium product prepared from the postbiotics according to claim 4 or 5, characterized in that: The invention comprises the following raw materials in parts by weight: 40-50 parts of oil, 25-35 parts of calcium citrate, 3-7 parts of stabilizer, 2-5 parts of postbiotics, 1-3 parts of emulsifier and 10-20 parts of purified water.

9. The calcium product according to claim 8, characterized in that The oil is one or more of conjugated linoleic acid glyceride, medium chain triglyceride, caprylic decanoic acid glyceride, cocoa butter, anhydrous butter, walnut oil, and linseed oil; the emulsifier is one or more of whey protein, casein, soy protein isolate, pasteurized egg yolk liquid, and lecithin; the stabilizer is one or more of pectin, L-arabic gum, carrageenan, xanthan gum, gelatin, and konjac flour.

10. The method for preparing the calcium product by nanoemulsification and homogenization according to claim 8 or 9, characterized in that: include: The oil, emulsifier, postbiotics and calcium citrate are mixed to obtain an oil phase; the stabilizer is mixed with purified water to obtain an aqueous phase; the oil phase and the aqueous phase are mixed and dispersed, and then high-pressure homogenized to obtain the calcium product; the pressure of the high-pressure homogenization is 100-160MPa, the time is 10-20min, and the number of times is 1-4 times.

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