Composition for promoting skeletal development and application thereof

Abstract

The invention belongs to the technical field of microbial engineering, and discloses a composition for promoting skeletal development and application thereof, and the composition contains bifidobacterium longum subsp. Infantis i18, 2 '-fucosyllactose and lactose-N-neotetraose. All the components in the composition have a synergistic effect, the content of calcium ions in bone cells can be remarkably increased, the bone density is improved, and the purpose of promoting skeletal development is achieved. The preparation method is suitable for preparing formula milk powder capable of promoting skeletal development or health-care food beneficial to improving bone mineral density.

Description

Technical Field

[0001] This invention belongs to the field of microbial engineering technology, specifically relating to a composition that promotes bone development and its application. Background Technology

[0002] Bone growth and development not only affect human morphology but also directly impact the development of internal organs and motor skills. Furthermore, the bone marrow within the bones plays a crucial role in hematopoiesis. Skull development during infancy influences brain development, while childhood and adolescence are critical periods for peak bone mass accumulation. The level of peak bone mass directly affects an individual's bone health and risk of osteoporosis in middle and old age. Therefore, bone development determines long-term health, and key factors influencing bone development include calcium absorption, calcium ion accumulation in bone cells, and the regulation of hormones related to bone development.

[0003] In recent years, the regulatory role of gut microbiota in various bodily functions has been frequently reported. Studies have shown that there is a clear link between the composition of gut microbiota and bone growth. Some probiotics are beneficial to bone growth and development, and further affect immune cells in bone marrow, thereby affecting the body's health.

[0004] Currently, there are numerous known types and strains of probiotics, with varying mechanisms of action and effects. Some strains, such as Lactobacillus reuteri, Lactobacillus rhamnosus, and Lactobacillus casei, have been reported to have certain effects on bone health and can improve osteoporosis-related symptoms. While Bifidobacterium longum and its subspecies have shown outstanding effects in treating or alleviating gastrointestinal diseases in infants and young children—for example, Bifidobacterium longum 35624 can prevent T-cell metastatic colitis—there is little public information on its use in promoting bone health.

[0005] Furthermore, even within the same species and genus, different strains exhibit functional differences due to phenotypic variations. For instance, the ATCC 15697 strain of Bifidobacterium longum subsp. infantis can successfully ferment and metabolize HMOs (human milk oligosaccharides), while another strain, Bifidobacterium infantis R0033, utilizes HMOs with extremely low efficiency.

[0006] Therefore, exploring new uses for known probiotic strains or discovering new strains that promote bone development are key and challenging issues in addressing bone health problems from the perspective of probiotics. Summary of the Invention

[0007] To address the aforementioned problems in existing technologies, this invention screens strains of Bifidobacterium longum that exhibit outstanding effects in promoting bone development from dozens of strains. In particular, it was discovered that a new strain, Bifidobacterium longum subsp. infanti 18, has significant efficacy in promoting bone development. Furthermore, it was unexpectedly found that when Bifidobacterium longum subsp. infanti 18 is used in combination with a specific ratio of 2'-fucosylated lactose and lactose-N-neotetrasaccharide, it can significantly increase the calcium ion content in bone cells, thereby promoting bone development.

[0008] The present invention also provides the use of a composition containing Bifidobacterium longum infantis subsp. i18 that promotes bone development, the composition being able to achieve the purpose of promoting bone development.

[0009] The present invention also provides the use of compositions that promote bone development in the preparation of infant formula, children's formula or health foods that promote bone development or help improve bone density.

[0010] The present invention also provides formula milk powder containing a composition that promotes bone development.

[0011] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0012] A composition for promoting bone development, comprising *Bifidobacterium longum subsp. infantis* i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide, wherein the ratio of *Bifidobacterium longum subsp. infantis* i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide is 1 × 10⁻⁶. 6 ~1×10 10 CFU: 560~4000mg: 160~1200mg.

[0013] Among them, Bifidobacterium longum subsp. infanti (i18) was deposited on July 25, 2024, at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 31411.

[0014] Bifidobacterium longum infant subspecies i18 promotes bone development by increasing bone density and / or intracellular calcium ion concentration.

[0015] As a limitation of the present invention, the composition further contains colostrum basic protein (CBP), casein phosphopeptide (CPP), or 1,3-dioleoyl-2-palmitoylglycerol triglyceride (OPO).

[0016] As a further limitation of the present invention, the ratio of Bifidobacterium longum subsp. infantis i18 to colostrum basic protein in the composition is 1×10⁻⁶. 6 ~1×10 10 CFU: 50-120mg.

[0017] As a further limitation of the present invention, the ratio of Bifidobacterium longum subsp. infantis i18 to casein phosphopeptide in the composition is 1×10⁻⁶. 6 ~1×10 10 CFU: 50-150mg.

[0018] As a further limitation of the present invention, the ratio of Bifidobacterium longum subsp. infantis i18 to 1,3-dioleoyl-2-palmitoylglycerol triglyceride in the composition is 1×10⁻⁶. 6 ~1×10 10 CFU: 3000~8000mg.

[0019] As a further limitation of the present invention, the ratio of Bifidobacterium longum subsp. infantis i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide is 1×10⁻⁶. 6 ~1×10 10 CFU: 560~1920mg: 160~480mg.

[0020] The preferred ratio of Bifidobacterium longum subsp. infantis i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide is 1×10⁻⁶. 9 CFU: 920mg: 160mg.

[0021] The present invention also provides the application of the above-mentioned composition for promoting bone development, specifically for the preparation of infant formula, children's formula or health food that helps improve bone density.

[0022] As a limitation of the present invention, the bone development promoting composition is added in infant formula, toddler formula, fermented milk, or health foods that help improve bone density at a weight percentage of 0.6% to 4.5%, wherein Bifidobacterium longum subsp. infantis i18 is added in the form of bacterial powder, and the viable count of the bacterial powder is ≥4×10⁻⁶. 10 CFU / g.

[0023] As a further limitation of the present invention, the composition for promoting bone development is added in infant formula, toddler formula, or health foods that help improve bone density at a weight percentage of 0.6% to 2.2%, wherein Bifidobacterium longum subsp. infantis i18 is added in the form of bacterial powder, and the viable count of the bacterial powder is ≥4×10⁻⁶. 10 CFU / g.

[0024] The present invention also provides a formula milk powder that promotes bone development, comprising the composition for promoting bone development as described in any of the above technical solutions.

[0025] The present invention also provides a health food that promotes bone development, comprising the composition for promoting bone development described in any one of the above technical solutions.

[0026] By adopting the above-described technical solution, the beneficial effects achieved by this invention compared to the prior art are as follows:

[0027] (1) The present invention discovered that Bifidobacterium longum subsp. infanti18 has the effect of promoting bone development, and developed a composition to promote bone development, effectively utilizing the mechanism of action of Bifidobacterium longum subsp. infanti18 to promote bone development by increasing bone density and / or calcium ion concentration in bone cells.

[0028] (2) The components of the composition for promoting bone development of the present invention have a synergistic effect, so that 2'-fucosylated lactose (2'-FL) and lactose-N-neotetrasaccharide (LNnT), which have no effect on improving calcium ion concentration and promoting bone development when used alone at a certain concentration, are combined with Bifidobacterium longum subsp. infanti18 at a specific concentration and used in the form of a composition, which has a very significant effect on increasing calcium ion concentration and promoting bone development.

[0029] (3) The addition of specific amounts of colostrum basic protein or casein phosphopeptide to the composition for promoting bone development of the present invention significantly increases calcium ion concentration and helps improve bone density, thereby promoting bone development.

[0030] (4) The composition for promoting bone development in this invention can be applied to products such as infant formula, children's formula, fermented milk, and health foods that help improve bone density.

[0031] This invention is applicable to the preparation of compositions that promote bone development, and is used to prepare infant formula, children's formula, or health foods that help improve bone density. Attached Figure Description

[0032] The present invention will now be described in further detail with reference to the accompanying drawings and specific embodiments.

[0033] Figure 1 This is a typical result diagram of fluorescence intensity in each group during the verification experiment of promoting bone development according to the present invention;

[0034] Figure 2 This is a graph showing the results of fluorescence intensity data analysis after sample treatment in the verification experiment of promoting bone development in this invention. Detailed Implementation

[0035] The present invention will be further described in detail below with reference to specific embodiments and accompanying drawings. It should be understood that the described embodiments are only used to explain the present invention and do not limit the present invention.

[0036] Unless otherwise specified, all materials and reagents used in the embodiments of this invention are commercially available. Experimental methods not specifically described in the embodiments are generally performed under standard conditions or as recommended by the manufacturer.

[0037] In the verification experiments of this invention, the reagent dosages were all verified through minimum toxic concentration detection experiments. The reagents were prepared as follows: CPP (casein phosphopeptide) (hereinafter referred to as CPP), prepared as a 20.0 mg / mL stock solution with standard dilution water, to be used immediately. LNnT (lactose-N-neotetrasaccharide) (hereinafter referred to as LNnT), prepared as a 20.0 mg / mL stock solution with standard dilution water, to be used immediately. 2'-FL (2'-fucosylated lactose) (hereinafter referred to as 2'-FL), prepared as a 20.0 mg / mL stock solution with standard dilution water, to be used immediately. CBP (colostrum basic protein) (hereinafter referred to as CBP), prepared as a 20.0 mg / mL stock solution with standard dilution water, to be used immediately. OPO (1,3-dioleoyl-2-palmitoylglycerol) (hereinafter referred to as OPO), prepared as a 20.0 mg / mL stock solution with standard dilution water, to be used immediately. Lutein, prepared as a 20.0 mg / mL stock solution with standard dilution water, should be prepared and used immediately. Calcium carbonate (hereinafter referred to as CaCO3), prepared as a 20.0 mg / mL stock solution with standard dilution water, should be prepared and used immediately. Bifidobacterium infantis i18 (hereinafter referred to as i18), prepared as a 20.0 mg / mL stock solution with standard dilution water, should be prepared and used immediately. Commercial Bifidobacterium infantis A (total viable count consistent with i18), prepared as a 2.00 mg / mL stock solution with standard dilution water, should be prepared and used immediately. Swisse "Calcium & Vitamin D", yellow capsules, batch number 47295, Swisse, store in a cool, dry, and light-protected place. Prepare as a 20.0 mg / mL stock solution with standard dilution water, prepare and use immediately.

[0038] In the verification experiments of this invention, zebrafish were raised in aquaculture water at 28℃ (water quality: 200mg of readily soluble sea salt added per 1L of reverse osmosis water; conductivity 450-550μS / cm; pH 6.5-8.5; hardness 50-100mg / L CaCO3). Wild-type AB strain zebrafish were bred through natural pair mating. Zebrafish samples aged 3 days past their first month (dpf) were used to verify the efficacy in promoting bone development.

[0039] Example 1: A composition for promoting bone development

[0040] The composition for promoting bone development in this embodiment contains 1 × 10⁶ live bacteria.11 100 mg of Bifidobacterium longum subsp. infantis i18 (CFU / g) (i.e., the total viable count of Bifidobacterium longum subsp. infantis i18 is 1×10⁻⁶) 10 920 mg of CFU, 2'-fucosylated lactose, and 160 mg of lactose-N-neotetrasaccharide were mixed to obtain a composition that promotes bone development by increasing bone density and / or intracellular calcium ion concentration.

[0041] Example 2: A bone development promoting composition containing CBP

[0042] The composition of this embodiment contains 1×10⁶ viable bacteria. 11 100 mg of Bifidobacterium longum subsp. infantis i18 (CFU / g) (i.e., the total viable count of Bifidobacterium longum subsp. infantis i18 is 1×10⁻⁶) 10 CFU), 920 mg of 2'-fucosylated lactose, 160 mg of lactose-N-neotetrasaccharide, and 50 mg of colostrum basic protein were mixed to obtain a bone development-promoting composition containing CBP.

[0043] Example 3: A CPP-containing composition for promoting bone development

[0044] The composition of this embodiment contains 1×10⁶ viable bacteria. 11 100 mg of Bifidobacterium longum subsp. infantis i18 (CFU / g) (i.e., the total viable count of Bifidobacterium longum subsp. infantis i18 is 1×10⁻⁶) 10 The following ingredients were mixed: CFU, 2'-fucosylated lactose 920 mg, lactose-N-neotetrasaccharide 160 mg, and casein phosphopeptide 50 mg to obtain a bone development promoting composition containing CPP.

[0045] (I) Verification test of efficacy in promoting bone development

[0046] (1.1) Group settings:

[0047] 510 zebrafish with a 3dpf fin were randomly selected and divided into 17 groups of 30 each in beakers.

[0048] The treatments for the 17 experimental groups were as follows:

[0049] The normal control group received no special treatment.

[0050] The positive control group was treated with Swisse "Calcium & Vitamin D" 250 μg / mL.

[0051] Groups 1 through 5 were administered equal amounts of the separately applied samples according to Table 1, with Group 1 containing 4 × 10⁻⁶ viable *Bifidobacterium longum* subsp. infantis i18. 10CFU / g, the number of live commercial Bifidobacterium infantis A added in group 5 was also 4×10⁻⁶. 10 CFU / g.

[0052] Groups 6 to 15 were each given an equal amount of the sample in the form of a composition, according to Table 1.

[0053] Table 1. Types and dosages of samples used in each group

[0054]

[0055]

[0056] Note: ① Groups 9, 11, and 13 are the zebrafish dosages converted from the daily human doses of Examples 1-3. The dosages for other groups were adaptively adjusted based on the dosages in Group 9 combined with the safe zebrafish dose. Specifically, the OPO dosages for Groups 14 and 15 were converted from an infant intake of 4.83 g / d to a zebrafish dosage of 805 μg / mL. ② Commercial Bifidobacterium infantis A.

[0057] (1.2) Test methods:

[0058] After 96 hours of treatment at 28℃, samples from each experimental group were collected. Alizarin red staining of the vertebrae was used to characterize the calcium ion content in bone cells. Ten zebrafish were randomly selected from each experimental group and photographed under a fluorescence microscope to obtain fluorescence intensity images of the zebrafish vertebrae from each group. Data were analyzed and collected using NIS-ElementsD3.20 advanced image processing software. The fluorescence intensity of the zebrafish vertebrae was analyzed, and the statistical analysis results of this index were used to evaluate the bone development-promoting efficacy of each applied sample. Statistical results are expressed as mean ± SE. Statistical analysis was performed using SPSS 26.0 software. * indicates p < 0.05, indicating a statistically significant difference; ** indicates p < 0.01, indicating an extremely significant difference.

[0059] (1.3) Experimental Results:

[0060] Typical images were selected from the fluorescence intensity photographs of zebrafish vertebrae, and the results are as follows: Figure 1 As shown;

[0061] The statistical results of fluorescence intensity in zebrafish vertebrae are shown in Table 2 and Figure 2 As shown.

[0062] Table 2. Statistical results of fluorescence intensity of zebrafish vertebrae in each group.

[0063]

[0064]

[0065] (1.4) Results Analysis:

[0066] From Table 2, Figure 1 and Figure 2 It can be seen that, under the experimental conditions, compared with the normal control group, group 2 (using commercial Bifidobacterium infantis alone), group 3 (using low concentration 2'FL alone), group 4 (using medium concentration 2'FL alone), and group 5 (using LNnT alone) had no significant effect on promoting bone development; while group 1 (using i18 alone) had a significant effect on promoting bone development (<0.05). This indicates that the present invention has found that Bifidobacterium longum subsp. infanti 18 has the effect of promoting bone development. Since the fluorescence intensity of vertebral bone directly reflects the calcium ion concentration, it indicates that Bifidobacterium longum subsp. infanti 18 promotes bone development through bone density and / or intracellular calcium ion concentration.

[0067] Compared with the normal control group, groups 8 (low concentration of i18+2'-FL+LNnT) and 9 (medium concentration of i18+2'-FL+LNnT) showed significant and more significant effects on promoting bone development, respectively (<0.01), indicating that the components in the composition of the present invention have a synergistic effect and significantly promote bone development compared with the use of the corresponding concentrations of 2'FL and LNnT alone. The significantly high fluorescence intensity of groups 10-15 indicates that "Bifidobacterium longum infantis i18++2'FL+LNnT+CaCO3" can synergistically enhance the effect of promoting bone development when used in conjunction with CPP, CBP or OPO.

[0068] Example 4: A composition for promoting bone development

[0069] The composition of this embodiment contains 1×10⁶ viable bacteria. 11 100 mg of Bifidobacterium longum subsp. infantis i18 (CFU / g) (i.e., the total viable count of Bifidobacterium longum subsp. infantis i18 is 1×10⁻⁶) 10 CFU), 2'-fucosyl lactose 560mg, lactose-N-neotetrasaccharide 480mg and 1,3-dioleoyl-2-palmitoylglycerol triglyceride 4830mg.

[0070] Examples 5-8: Compositions for Promoting Bone Development

[0071] Examples 5-8 are compositions for promoting bone development, and their specific components and dosages are shown in Table 5.

[0072] Table 5 shows the specific components and dosages of each composition in Examples 5-8.

[0073]

[0074] Experimental results have shown that the above-mentioned compositions all have outstanding effects in promoting bone development.

[0075] Examples 9-12 are infant formula milk powders containing a composition that promotes bone development.

[0076] The infant formula in Example 9 contains 0.6% by weight of the bone-promoting composition from Example 1, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0077] The infant formula in Example 10 contains 4.0% by weight of the bone-promoting composition from Example 2, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0078] The infant formula in Example 11 contains 4.5% by weight of the bone-promoting composition from Example 3, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0079] The infant formula in Example 12 contains 2.4% by weight of the bone-promoting composition from Example 4, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0080] Examples 13-24 are formula milk powders containing compositions that promote bone development.

[0081] The infant formula in Example 13 contains 0.7% by weight of the bone-promoting composition from Example 1, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0082] The infant formula in Example 14 contains 1.6% by weight of the bone-promoting composition from Example 2, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0083] The infant formula in Example 15 contains 2.0% by weight of the bone-promoting composition from Example 3, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0084] The infant formula in Example 16 contains 2.2% by weight of the bone-promoting composition from Example 4, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0085] The infant formula in Example 17 contains 2.4% by weight of the bone-promoting composition from Example 5, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0086] The formula milk powder in Example 18 contains 0.8% by weight of the bone development promoting composition from Example 6, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4×10⁻⁶. 10 CFU / g.

[0087] The formula milk powder in Example 19 contains 1.7% by weight of the bone development promoting composition from Example 1, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4×10⁻⁶. 10 CFU / g.

[0088] The formula milk powder in Example 20 contains 2.3% by weight of the bone development promoting composition from Example 2, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4×10⁻⁶. 10 CFU / g.

[0089] The formula milk powder in Example 21 contains 2.2% by weight of the bone development promoting composition from Example 3, wherein Bifidobacterium longum subsp. infantis i18 is added in the form of a bacterial powder with a viable count ≥4×10⁻⁶. 10 CFU / g.

[0090] The formula milk powder in Example 22 contains 2.9% by weight of the bone development promoting composition from Example 4, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4×10⁻⁶. 10 CFU / g.

[0091] The formula milk powder in Example 23 contains 4% by weight of the bone development promoting composition from Example 3, wherein Bifidobacterium longum subsp. infantis i18 is added in the form of a bacterial powder with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0092] The formula milk powder in Example 24 contains 3.5% by weight of the bone development promoting composition from Example 4, wherein Bifidobacterium longum subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g.

[0093] In other embodiments, the bone-developing composition is added to infant formula, toddler formula, or health food at a weight percentage of 0.6% to 4.5%, wherein *Bifidobacterium longum* subsp. infantis i18 is added in powder form with a viable count ≥4 × 10⁻⁶. 10 CFU / g. All products obtained have the effect of promoting bone development.

[0094] In other embodiments, the ratio of Bifidobacterium longum subsp. infantis i18 to 1,3-dioleoyl-2-palmitoylglycerol triglyceride (opo) in the composition promoting bone development is 1 × 10⁻⁶. 6 ~1×10 10 CFU: 3000–8000 mg. For example, the total viable count of *Bifidobacterium longum* subsp. infantis i18 is 1 × 10⁻⁶. 6 CFU, 1×10 8 CFU, 2×10 9 CFU or 1×10 10 CFU, corresponding to the dosage of 1,3-dioleoyl-2-palmitoylglycerol triglyceride, is 3000 mg, 6500 mg, 7500 mg, or 8000 mg.

[0095] It should be noted that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art can still modify the technical solutions described in the above embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the claims of the present invention.

Claims

1. A composition for promoting bone development, characterized in that, The composition contains *Bifidobacterium longum* subsp. infantis i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide, wherein the ratio of *Bifidobacterium longum* subsp. infantis i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide is 1 × 10⁻⁶. 6 ~1×10 10 CFU: 560~4000mg; 160~1200mg.

2. The composition for promoting bone development according to claim 1, characterized in that, The composition also contains colostrum basic protein, casein phosphopeptide, or 1,3-dioleoyl-2-palmitoylglycerol.

3. The composition for promoting bone development according to claim 2, characterized in that, The ratio of Bifidobacterium longum infantis subsp. i18 to colostrum basic protein in the composition is 1×10⁻⁶. 6 ~1×10 10 CFU: 50~120mg.

4. The composition for promoting bone development according to claim 2, characterized in that, The ratio of Bifidobacterium longum subsp. infantis i18 to casein phosphopeptide in the composition is 1×10⁻⁶. 6 ~1×10 10 CFU: 50~150mg.

5. The composition for promoting bone development according to claim 2, characterized in that, The composition contains Bifidobacterium longum subsp. infantis i18 in a ratio of 1×10⁻⁶ to 1,3-dioleoyl-2-palmitoylglycerol triglyceride. 6 ~1×10 10 CFU: 3000~8000mg.

6. The composition for promoting bone development according to claim 3 or 4, characterized in that, The ratio of Bifidobacterium longum subsp. infantis i18, 2'-fucosylated lactose, and lactose-N-neotetrasaccharide is 1×10⁻⁶. 6 ~1×10 10 CFU: 560~1920mg; 160~480mg.

7. The application of the composition for promoting bone development according to any one of claims 1 to 6, characterized in that, The composition for promoting bone development is used in the preparation of infant formula, children's formula, or health foods that help improve bone density.

8. The application of the composition for promoting bone development according to claim 7, characterized in that, The composition promoting bone development is added at a weight percentage of 0.6% to 4.5% in infant formula, toddler formula, or health foods that help improve bone density, wherein Bifidobacterium longum subsp. infantis i18 is added in the form of bacterial powder with a viable count ≥4×10⁻⁶. 10 CFU / g.

9. A formula milk powder that promotes bone development, characterized in that, A composition comprising any one of claims 1 to 6 that promotes bone development.

10. A health food product that helps improve bone density, characterized in that, A composition comprising any one of claims 1 to 6 that promotes bone development.

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