A postbiotic composition for protecting lung health and methods of making and using the same

Abstract

This invention discloses a post-biotic composition for protecting lung health, its preparation method, and its application. The active ingredient is derived from Bifidobacterium infantis B1398. This post-biotic is prepared by co-fermentation, inactivation, and drying of lily bulb, dried tangerine peel, yam, and licorice root as the fermentation medium. Animal experiments show that this composition can significantly regulate the immune function of rats with lipopolysaccharide-induced lung inflammation, reducing serum IgM and increasing IgG and IgA; it regulates inflammation-related indicators, reducing pro-inflammatory factors such as TNF-α, IL-6, and IL-8 in bronchoalveolar lavage fluid; and it can reduce the lung wet / dry weight ratio, with effects superior to post-biotics prepared from known subspecies strains. This invention provides a new drug option for daily lung health protection and auxiliary regulation of lung function.

Description

Technical Field

[0001] This invention relates to a post-biotic composition for protecting lung health, its preparation method, and its application, belonging to the field of microbial technology. Background Technology

[0002] With increasing air pollution, the prevalence of respiratory pathogens, and the widespread presence of risk factors such as smoking, the incidence of lung diseases continues to rise, posing a serious challenge to global public health. Metabiotics refer to non-viable microorganisms or components obtained through fermentation that are beneficial to host health, including microbial components and metabolites (such as short-chain fatty acids, bacteriocins, and extracellular polysaccharides). Compared with traditional probiotics, metabiotics have advantages such as high stability, good safety, well-defined targets, and ease of standardized production. Studies have shown that certain metabiotics can positively impact distal lung health through "gut-lung axis" mechanisms such as regulating gut microbiota, enhancing mucosal immunity, and modulating excessive inflammatory responses. Currently, research has explored the potential immunomodulatory roles of metabiotics from some Bifidobacterium strains.

[0003] However, existing technologies still have significant limitations: First, while numerous Bifidobacterium strains with potential health benefits have been reported, the biological effects of different strains, and even different strains of the same species, vary significantly. Their protective effects on specific organs (such as the lungs) cannot be simply extrapolated, making it crucial to screen for strains with specific and superior effects on lung health. Second, conventional postbiotic preparations often use single-component culture media, resulting in relatively singular product functions. Furthermore, simply physically mixing prebiotics or plant extracts with postbiotics has limited synergistic effects and fails to achieve deep integration and enhancement between components.

[0004] Based on this, the present invention aims to overcome the shortcomings of the prior art and provide a postbiotic composition prepared by a specific high-efficiency strain and integrated with an innovative fermentation process, in order to achieve better and unexpected technical effects in protecting lung health. Summary of the Invention

[0005] To address the problems of unclear strain effects, limited functionality, and lack of deep synergy in existing technologies described in the background section, this invention provides a metabiotic composition with outstanding protective effects on lung health, prepared from specific high-efficiency strains using an innovative process, as well as its preparation method and application.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a metabiotic composition for protecting lung health. The active ingredient of the composition comprises a metabiotic obtained from Bifidobacterium infantis B1398 (accession number CGMCC No. 22241) through fermentation and inactivation. Compared with known strains of the same subspecies in the prior art, the present invention provides a new pharmaceutical option for protecting lung health and assisting in the regulation of lung function.

[0007] Preferably, to further enhance efficacy and achieve deep integration of components, the fermentation is carried out in a culture medium containing specific raw materials. The raw materials are selected from at least one of lily bulb, dried tangerine peel, yam, and licorice.

[0008] More preferably, the postbiotic composition further comprises prebiotics (such as fructooligosaccharides, inulin) and / or other natural ingredients (such as curcumin).

[0009] Secondly, the present invention provides a method for preparing the above-mentioned post-biotic composition. The key to this method lies in the composition of the fermentation medium and the fermentation process, which includes the following steps: 1) Prepare a fermentation medium containing a carbon and nitrogen source and at least one of lily, dried tangerine peel, yam, and licorice; (2) Inoculate Bifidobacterium infantis B1398 into the culture medium at a specific inoculation amount and ferment it under temperature-controlled anaerobic conditions until the predetermined fermentation endpoint (such as pH value) is reached. (3) The fermentation broth is inactivated (e.g., pasteurized) and dried (e.g., spray dried) to obtain postbiotic powder; and optionally (4) the obtained postbiotic powder is mixed with prebiotics and other excipients.

[0010] This invention provides the application of the aforementioned post-biotic composition in the preparation of a medicament for protecting lung health and assisting in the regulation of lung function; the medicament is used to regulate lipopolysaccharide-induced lung-related indicators or to regulate the lung wet / dry weight ratio. The medicament may be in the form of a special medical purpose formulation, etc.

[0011] Compared with the prior art, the present invention has the following beneficial effects: High strain specificity and significant effect: Comparative experiments have confirmed that the B1398 strain selected in this invention has significantly better effects on key indicators than other known strains of the same subspecies (such as NKU FB 3-14 and M-63), solving the problems of uncertain strain effects and weak specificity in the prior art.

[0012] Technological innovation and deep integration of ingredients: Creatively using specific plant raw materials (such as lily, dried tangerine peel, yam, and licorice) as fermentation substrates, and through the co-fermentation process of Bifidobacterium infantis B1398, a deep integration of microbial metabolism and plant component transformation is achieved. This may not only generate new active substances, but also endow the product with more synergistic and diverse lung health functions.

[0013] Scientific formulation and synergistic effect: By combining the post-fermentation matrix obtained by co-fermentation with specific prebiotics, a synergistic effect of "1+1>2" is generated, especially in maintaining the integrity of the lung epithelial barrier, providing a better solution than single components or simple mixtures.

[0014] High safety and broad application prospects: The product ingredients are derived from natural strains and plant materials, ensuring high safety and suitability for long-term conditioning. Its diverse product forms and usage methods provide a brand-new option for daily protection and auxiliary regulation of lung health.

[0015] Preservation of biological materials A strain of Bifidobacterium infantis B1398, taxonomically named Bifidobacterium infantis It was deposited on April 27, 2021, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 22241. The deposit address is Institute of Microbiology, Chinese Academy of Sciences, No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. Attached Figure Description

[0016] Figure 1 Effects of post-biotics on serum IgG levels in rats; Figure 2 Effects of post-biotics on serum IgM levels in rats; Figure 3 Effects of post-biotics on serum IgA levels in rats; Figure 4 Effects of post-biotics on the levels of TNF-α, IL-6, IL-8, and IL-10 in rat BALF; Figure 5 Effect of post-biotics on the ratio of wet to dry lung weight in rats. Detailed Implementation

[0017] The following embodiments are provided to better understand the present invention and are not limited to the preferred embodiments described. They do not constitute a limitation on the content and scope of protection of the present invention. Any product that is the same as or similar to the present invention, derived by any person under the guidance of the present invention or by combining the features of the present invention with other prior art, falls within the protection scope of the present invention.

[0018] For any experimental steps or conditions not specified in the following examples, the procedures or conditions described in the literature in this field can be followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional reagent products.

[0019] Example 1: Preparation of Bifidobacterium infantis B1398 postbiotic and its composition This embodiment creatively uses raw materials with lung-health benefits as an important component of the fermentation culture medium. Through the fermentation transformation of a specific strain B1398, a more complex and synergistic probiotic and its composition are prepared. The preparation method of the probiotic probiotic is as follows: 1. Preparation of fermentation medium By weight percentage, the fermentation medium contains the following components: Carbon and nitrogen base: 6% skim milk powder, 1.5% soy peptone.

[0020] Lily powder: 2% Tangerine peel extract: 1.5% Yam powder: 2% Licorice extract: 0.5% Remaining amount: purified water.

[0021] Mix all ingredients and stir at 60℃ and 150 rpm for 30 minutes to ensure complete dissolution and homogenization. Then homogenize at 58℃ using a first-stage pressure of 15 MPa and a second-stage pressure of 5.0 MPa. The homogenized solution is then sterilized at 95℃ for 20 minutes and cooled to 36±1℃ for later use.

[0022] 2. Co-fermentation and metabiotic preparation of strain B1398 Inoculation and fermentation: After activating Bifidobacterium infantis B1398, it was inoculated into the above culture medium at an inoculation rate of not less than 6×10^6 CFU / mL.

[0023] Fermentation control: Fermentation was carried out at 36±1℃ under anaerobic conditions. pH was monitored, and fermentation was terminated when the pH of the fermentation broth dropped to 4.60. At this point, the cells proliferated significantly and completed the initial transformation of the culture medium components.

[0024] Post-processing: The fermentation broth was pasteurized (70℃, 30 minutes) to inactivate the bacteria. Then, it was spray-dried (inlet air temperature 170℃, outlet air temperature 70℃) to obtain powdered metabiotic.

[0025] 3. Preparation of the compound composition To further enhance the effect, the above-mentioned co-fermented biogener can be compounded with functional excipients.

[0026] The composition formulation is as follows, based on parts by weight: B1398 Postbiotic Powder: 10 parts Prebiotics (fructooligosaccharides and inulin in a 1:1 ratio): 3 servings Natural anti-inflammatory adjuvant (curcumin microcapsule powder): 1 part All ingredients are physically mixed evenly in a three-dimensional mixer to obtain the final "post-lung health protective composition".

[0027] Comparative Example 1 Comparative Example 1 was prepared by replacing the strain B1398 described in this invention with Bifidobacterium infantis NKU FB 3-14 (CGMCC No. 25762) and following the same postbiotic preparation method as in Example 1.

[0028] Comparative Example 2 Two comparative groups were prepared by replacing the strain B1398 described in this invention with Bifidobacterium infantis M-63 (purchased from Shandong Xinxiong Biotechnology Co., Ltd.) and following the same postbiotic preparation method as in Example 1.

[0029] Example 2: Effects on lipopolysaccharide-induced pulmonary immune inflammatory response in rats 1. Laboratory animals Fifty SPF-grade male SD rats, weighing 200–220 g, were purchased from SPF (Beijing) Biotechnology Co., Ltd. The rats were housed separately in cages at a temperature of 22±2℃, humidity of 55%–75%, with a 12-hour light / dark cycle, and free access to water and food. All rats underwent acclimatization for one week.

[0030] 2. Experimental Methods Fifty rats were randomly divided into five groups using a random number table: normal group, model group, metageneric group, control group 1, and control group 2, with ten rats in each group. Except for the normal group, all other groups received lipopolysaccharide (LPS, 1 mg / kg) via intratracheal instillation. - ¹, a lung inflammation model was established by dissolving the compound in sterile saline; the normal group was infused with an equal volume of sterile saline. The lung inflammation model rats were randomly divided into a model group, a metagenic group, a control group 1, and a control group 2, with 10 rats in each group. The metagenic group and the control group were given 5.0 × 10⁻⁶ mmol / L of sterile saline solution. 8 Cells / d / animal, where Cells is the unit of metabiotic cells, representing the number of cells, and Cells / d / animal is the daily dose unit. The normal group and the model group were given an equal volume of physiological saline by gavage once daily for 14 consecutive days.

[0031] 2. Observation indicators (1) Determination of IgG, IgM, and IgA levels Rats in each group were anesthetized by intraperitoneal injection of 3% sodium pentobarbital. 1.5 mL of blood was collected from the abdominal aorta of each group. The blood was allowed to stand at room temperature for 8 min, then subjected to a pulse at 3500 r·min. -1 Centrifuge for 12 min (centrifugation radius 8 cm, the same below) to prepare the serum of each group of rats. Place the serum on an ice box and use an ELISA kit to detect the levels of IgG, IgM and IgA in the serum of each group of rats according to the kit instructions.

[0032] (2)TNF-α, IL-6, IL-8, IL-10 content After blood collection, the thoracic cavity was opened to expose the lung tissue. The lung tissue was separated, and the trachea was fully exposed. Endotracheal intubation was performed using an indwelling needle. 5 mL of pre-cooled phosphate-buffered saline (PBS) was drawn and the alveoli were lavaged. This lavage was repeated three times, and the lavage fluid was collected in centrifuge tubes. The recovery rate of bronchoalveolar lavage fluid (BALF) was above 85%. The BALF was centrifuged at 4 ℃ and 2000 r·min⁻¹ for 10 min. The supernatant was aliquoted into centrifuge tubes and stored at -80 ℃ for later use. The levels of TNF-α, IL-6, IL-8, and IL-10 in the BALF of each group of rats were detected using an ELISA kit according to the kit instructions.

[0033] (3) Immune organ mass and lung wet / dry weight (W / D) The thoracic and abdominal cavities were cut open, and the thymus and lungs were removed, weighed, and recorded. The upper lobe of the left lung of the rat was removed, washed, dried, weighed, and the wet weight (W) was recorded. The lung tissue was placed in a 70 ℃ constant temperature incubator for continuous drying for more than 48 h until the weight no longer changed. The weight was recorded again and the dry weight (D) was recorded. The lung W / D ratio was calculated.

[0034] 4. Results (1) Effects of post-biotics on the levels of IgG, IgM, and IgA in rat serum The concentrations of the three immunoglobulins (IgG, IgA, IgM) are as follows: Figures 1-3As shown in the figure, the model group showed significant differences in all three indicators. Compared with the normal group, the model group had significantly lower IgG and IgA concentrations and significantly higher IgM concentrations. This suggests that the model group may be in a state of immune dysfunction, manifested by abnormalities in immunoglobulin synthesis and secretion. The post-biotic group performed better than the model group in all three indicators. Specifically, the post-biotic group significantly increased IgG and IgA concentrations and effectively reduced IgM concentrations compared to the model group. These results indicate that post-biotics may have a positive effect on regulating normal immune function. The control group had IgG and IgA concentrations between the normal and model groups, failing to significantly regulate the low IgG and IgA state of the model group. However, regarding IgM, the control group showed a lower concentration than the model group, but still higher than the normal group. This suggests that the control group may have partially alleviated the immune abnormalities of the model group, but its effect was not as significant as that of the post-biotic group. In summary, the experimental results show that the model group exhibited significant immune dysfunction, while the post-biotic group demonstrated a significant effect in regulating immunoglobulin concentration, contributing to the restoration of a normal immune state. In contrast, although the control group also affected immunoglobulin concentration to some extent, its effect was weaker. Therefore, post-biotics may be one of the effective ways to regulate immune function, and their potential application value warrants further research and exploration.

[0035] (2) Effects of post-biotics on the levels of TNF-α, IL-6, IL-8 and IL-10 in rat BALF Figure 4 The concentrations of four cytokines (TNF-α, IL-6, IL-8, and IL-10) were presented in five different groups (normal group, model group, post-biotic group, and control groups 1 and 2). The results showed that, compared with the model group, the concentrations of TNF-α, IL-6, and IL-8 in the post-biotic group were decreased, but still higher than those in the normal group; the concentration of IL-10 was slightly increased, but still lower than that in the normal group, indicating that the post-biotic group could regulate the inflammatory response to some extent.

[0036] The model group exhibited a significant inflammatory response, characterized by elevated levels of various pro-inflammatory cytokines (TNF-α, IL-6, IL-8) and decreased levels of the anti-inflammatory cytokine (IL-10). Both the post-genetic group and the control group were able to regulate the levels of these cytokines to varying degrees, with the post-genetic group showing a more significant effect, demonstrating its potential in regulating inflammation-related indicators.

[0037] (3) Effect of post-biotic on the wet weight to dry weight ratio (W / D) of rat lungs like Figure 5As shown, the wet-to-dry weight ratio of the lungs in the model group was significantly higher than that in the normal group, which may indicate a significant increase in this ratio in the model group, leading to increased lung tissue water content. Compared with the model group, the wet-to-dry weight ratio of the lungs in the post-genetic group decreased, but was still higher than that in the normal group. This indicates that the post-genetic group can reduce this ratio to some extent. Compared with the model group, the wet-to-dry weight ratio of the lungs in the control group did not change significantly, indicating that the control group failed to significantly alter this ratio.

[0038] These results indicate a significant increase in the lung wet / dry weight ratio in the model group. The post-genetic group was able to reduce this ratio to some extent, demonstrating its potential in regulating the lung wet / dry weight ratio. In contrast, the two control groups failed to significantly reduce the lung wet / dry weight ratio, and their effect on the control was less pronounced than that of the post-genetic group.

[0039] 5. Conclusion Experimental results show that the postbiotic prepared from Bifidobacterium infantis B1398 provided by this invention has a comprehensive and significant effect on protecting lung health in a lipopolysaccharide (LPS)-induced rat lung inflammation model. Its overall effect is significantly better than that of the known subspecies strains NKU FB 3-14 and M-63.

[0040] Specifically, the B1398 postbiotic of the present invention exhibits unexpected technical advancements in the following key aspects: the B1398 postbiotic of the present invention can significantly reduce the lung W / D ratio in model rats ( P <0.05), while the postbiotics prepared from known strains NKU FB 3-14 and M-63 (comparative group) did not show statistically significant changes in this indicator. B1398 postbiotic more effectively increased the levels of immunoprotective IgG and IgA, while more significantly reducing abnormally elevated IgM levels, bringing all indicators closer to the normal group. Its regulatory effect on the immune system was significantly stronger than the two comparative groups in both magnitude and comprehensiveness. Regarding the inflammatory cytokine profile in bronchoalveolar lavage fluid (BALF), B1398 postbiotic was superior to the comparative group in reducing pro-inflammatory factors (TNF-α, IL-6, IL-8) and increasing anti-inflammatory factors (IL-10), indicating that it has greater potential for regulating lung inflammation-related indicators.

[0041] In summary, although the NKU FB 3-14 and M-63 strains used in the comparative examples belong to the same genus as the Bifidobacterium infantis as the strain of this invention, and existing technologies have suggested that this genus / species of microorganism may have general immunomodulatory potential, the specific strain B1398 of this invention achieves significantly superior technical effects in the core function of "regulating the lung wet / dry weight ratio" and in the "systematic and balanced regulation" of lung immune-related indicators—effects that cannot be foreseen or simply deduced by existing technologies. Obviously, the above embodiments are merely examples for clear illustration and are not intended to limit the implementation. Those skilled in the art can make other variations or modifications based on the above description. It is neither necessary nor possible to exhaustively list all embodiments here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. An epigenetic composition for protecting lung health, characterized in that, Its active ingredients include Bifidobacterium infantis (Bifidobacterium infantis) Bifidobacterium infantis The metabiotic obtained by fermentation and inactivation of Bifidobacterium infantis B1398; the preservation number of Bifidobacterium infantis B1398 is CGMCC No. 22241; the fermentation is carried out in a culture medium containing at least one of lily powder, tangerine peel extract, yam powder and licorice extract.

2. The post-genetic composition according to claim 1, characterized in that, The culture medium contains, by weight percentage, 5-8% skim milk powder, 1-2% soybean peptone, 1-3% lily powder, 1-2% dried tangerine peel extract, 1-3% yam powder, and 0.3-1.0% licorice extract.

3. The epigenetic composition according to any one of claims 1-2, characterized in that, It also contains prebiotics and / or natural anti-inflammatory ingredients; the prebiotics are selected from at least one of fructooligosaccharides and inulin.

4. The epigenetic composition according to claim 3, characterized in that, By weight, it comprises: 8-12 parts of postbiotics obtained by fermentation and inactivation of the aforementioned Bifidobacterium infantis B1398, and 2-5 parts of prebiotics.

5. A method for preparing the post-genetic composition of claim 1, characterized in that, Includes the following steps: (1) Preparation of fermentation medium; (2) Inoculate Bifidobacterium infantis B1398 into the culture medium for fermentation culture to obtain fermentation broth; (3) The fermentation broth is inactivated and dried to obtain the post-biotic composition.

6. The method according to claim 5, characterized in that, In step (2), the inoculation amount of the fermentation culture is not less than 6 x 10 6 CFU / mL, the culture temperature is 35-37℃, and the fermentation end-point pH is 4.50-4.70; in step (3), the inactivation is pasteurization, and the drying is spray drying or freeze drying.

7. The use of the post-biotic composition according to claim 1 in the preparation of a medicine for protecting lung health and assisting in the regulation of lung function.

8. The application according to claim 7, characterized in that, The physiological states that assist in regulating lung function include at least one of the following: regulating changes in lung indicators induced by lipopolysaccharide, regulating the wet / dry weight ratio of lung tissue, or regulating changes in lung function parameters caused by external stimuli.

9. The application according to claim 8, characterized in that, The dosage form of the drug is an oral preparation or an inhaled preparation; the oral preparation includes powder, granules, tablets or capsules.

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