Selenium-dot chimeric probiotic and preparation method therefor and use thereof

Abstract

Provided are a selenium-dot chimeric probiotic and a preparation method therefor and the use thereof, wherein the selenium-dot chimeric probiotic comprises a probiotic and elemental selenium grown in-situ on the surface of the probiotic. Using a probiotic as the main structure, a probiotic with selenium dots grown in-situ on the surface is prepared by means of in-situ reduction. The preparation method is simple, and the process is simple to operate and easy to implement. In addition, the selenium-dot chimeric probiotic not only retains the biological activity of the probiotic itself, but also has the function of nano selenium, and shows better therapeutic effects on inflammation-related diseases. Furthermore, the provided preparation method enriches the preparation of engineered probiotics, and further expands the application of probiotics in the fields of nanomedicine and animal feeding.

Description

A chimeric selenium-dot probiotic and its preparation method and application

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to application number CN202311254049.5, filed on September 27, 2023, entitled “A Chimeric Selenium-Dot Probiotic, Preparation Method and Application Thereof,” the contents of which are incorporated herein by reference in their entirety. Technical Field

[0003] The present invention relates to the technical field of functional materials, and in particular to a chimeric selenium-dot probiotic and a preparation method and application thereof. Background Art

[0004] The intestinal microbiome is composed of trillions of bacteria, fungi, viruses and other microorganisms, and is involved in the host's immune regulation process. Studies have shown that the intestinal microbiome is one of the driving factors of inflammatory bowel disease and other related diseases. The occurrence of inflammatory bowel disease is usually accompanied by an imbalance of the intestinal microbiome. In recent years, researchers have used probiotics to intervene and regulate the intestinal microbiome, increase the colonization of beneficial microbial communities in the host, and thus regulate the imbalanced intestinal microbial homeostasis under inflammatory conditions. However, the treatment of inflammatory bowel disease with probiotics alone has limited effect, because the inflammatory response will produce excessive reactive oxygen free radicals, causing oxidative stress damage to the body's tissues, exacerbating and intensifying the inflammatory response. Therefore, while using probiotics to treat inflammatory-related diseases, it is crucial to eliminate reactive oxygen free radicals in the lesion site.

[0005] Selenium is an essential trace element for the human body, present in the antioxidant activity center of glutathione peroxidase. It plays a key role in regulating immune function and mitigating free radical oxidative damage. Therefore, moderate selenium supplementation can help treat inflammatory diseases. Furthermore, due to its enhanced biosafety and antioxidant activity, nano-selenium is considered a new generation of selenium supplements.

[0006] Although nano-selenium has antioxidant function, it has poor adhesion in the intestine and is easily metabolized and excreted from the body. Therefore, improving the enrichment efficiency of nano-selenium in the intestine and combining probiotics with nano-selenium to synergistically treat inflammatory-related diseases are issues that need to be urgently addressed.

[0007] Summary of the Invention

[0008] To address at least some of the technical problems in the prior art, the present invention provides a chimeric selenium-dot probiotic, its preparation method, and its use. This chimeric selenium-dot probiotic not only retains the biological activity of the probiotic itself but also, due to the presence of in situ-grown selenium dots, possesses the functions of nano-selenium, such as antioxidant activity. Therefore, it has excellent therapeutic effects on inflammatory diseases. Specifically, the present invention encompasses the following.

[0009] A first aspect of the present invention provides a chimeric selenium-dot probiotic comprising a probiotic and a selenium element growing in situ on the surface of the probiotic.

[0010] In certain preferred embodiments, according to the chimeric selenium-dot probiotics of the present invention, the probiotics include intestinal probiotics.

[0011] In certain preferred embodiments, according to the chimeric selenium-dot probiotics of the present invention, the intestinal probiotics include at least one of the genera Lactobacillus, Bifidobacterium, Streptococcus, Bacillus, Clostridium, Bacteroides, Lactococcus, Leuconostoc, Propionibacterium, Pediococcus and Saccharomyces.

[0012] In certain preferred embodiments, according to the chimeric selenium-dot probiotics of the present invention, the selenium element has a particle size of 1-100 nm.

[0013] In certain preferred embodiments, the chimeric selenium-dot probiotics according to the present invention are obtained by in situ adsorption of a selenium precursor on the probiotics, followed by reduction with a reducing agent, to obtain probiotics with selenium dots grown in situ on the surface.

[0014] The second aspect of the present invention provides a method for preparing a chimeric selenium-dot probiotic, comprising:

[0015] (1) culturing probiotics in a culture medium, and then co-incubating the cultured probiotics in a solvent containing a selenium precursor to obtain probiotics adsorbed with the selenium precursor;

[0016] (2) subjecting the probiotics adsorbed with selenium precursor to a reduction reaction with a reducing agent to obtain the chimeric selenium-dot probiotics.

[0017] In certain embodiments, according to the method for preparing the chimeric selenium dot probiotics of the present invention, the ratio of the probiotics to the selenium precursor is 10:1. 7-10 CFU:(0.002-10)mmol.

[0018] In certain embodiments, according to the method for preparing the chimeric selenium-dot probiotics of the present invention, the selenium precursor comprises organic selenium and / or inorganic selenium;

[0019] Preferably, the selenium precursor is selected from any one or combination of sodium selenite, potassium selenite, sodium selenate, potassium selenate, selenium dioxide, selenium oxide, selenomethionine, hydroxyselenomethionine, selenocysteine, selenocystine, selenomethylselenocysteine, selenomethylselenomethionine or seaweed selenium polysaccharide, preferably sodium sulfite.

[0020] In certain embodiments, according to the method for preparing chimeric selenium-dot probiotics of the present invention, the reducing agent includes any one or a combination of ascorbic acid, glutathione, sodium borohydride, hydrazine hydrate and polyphenols (such as catechin, hydroquinone, gallic acid, quercetin, resveratrol, etc.).

[0021] In certain embodiments, according to the method for preparing chimeric selenium-dot probiotics of the present invention, the solvent is selected from at least one of physiological saline, phosphate buffer solution, M17 medium, MRS medium, LB medium, BH medium, Bifidobacterium BS medium and Roche broth medium.

[0022] The third aspect of the present invention provides a use of a chimeric selenium-dot probiotic in the preparation of a medicine or dietary supplement for preventing, improving or treating inflammatory diseases.

[0023] The fourth aspect of the present invention provides a pharmaceutical composition or dietary supplement containing the chimeric selenium-dot probiotics of the present invention.

[0024] The fifth aspect of the present invention provides a feed additive containing the chimeric selenium-dot probiotics of the present invention.

[0025] The technical solution provided by the present invention has the following advantages compared with the prior art:

[0026] (1) The present invention uses probiotics as the main structure, in situ adsorbs selenium precursors, and then uses a reducing agent to reduce them to obtain probiotics with selenium spots growing in situ on the surface. The preparation method is simple and the process operation is simple and easy;

[0027] (2) The chimeric selenium dot probiotics provided by the present invention not only retain the biological activity of the probiotics themselves but also have the function of nano-selenium. Compared with the aqueous-phase synthesized nano-selenium (100-400 nm) reported in traditional literature, they have more outstanding functionality. For example, in terms of antioxidant activity, the chimeric selenium dot probiotics provided by the present invention can show a better therapeutic effect on inflammatory-related diseases.

[0028] (3) The preparation method provided by the present invention not only enriches the preparation methods of engineered probiotics, but also provides a theoretical basis and technical guidance for the application of probiotics in the fields of nanomedicine and animal husbandry. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] FIG1 shows a transmission electron micrograph of a chimeric selenium-dot Lactobacillus casei.

[0030] FIG2 shows the elemental analysis spectrum of the chimeric selenium-dot Lactobacillus casei.

[0031] FIG3 shows transmission electron micrographs of chimeric selenoid-containing Streptococcus thermophilus and Pediococcus acidilactici.

[0032] FIG4 shows a transmission electron micrograph of a chimeric selenium-dot Lactobacillus casei prepared using selenomethionine.

[0033] FIG5 shows the growth curve of chimeric selenium-dot Lactobacillus casei.

[0034] FIG6 shows the survival quantification of chimeric selenium-dot Lactobacillus casei and Lactobacillus casei in simulated gastric fluid.

[0035] FIG7 shows the fluorescence imaging images and quantitative curves of the chimeric selenium-dot Lactobacillus casei and Lactobacillus casei.

[0036] Figure 8 shows the body weight, colon length and HE section score data of ulcerative colitis mice after treatment, where the columns from left to right are the healthy group, model group, Lactobacillus casei group, Lactobacillus casei + nano-selenium mixed group, and chimeric selenium-dot Lactobacillus casei group.

[0037] FIG9 shows data on intestinal flora diversity, genus-level composition, and relative abundance of beneficial and harmful bacterial genera in mice after treatment of ulcerative colitis.

[0038] FIG10 shows the data on the scavenging of reactive oxygen species free radicals in vivo after treatment of ulcerative colitis mice.

[0039] FIG11 shows flow cytometry data of colon immune cells in mice after treatment with ulcerative colitis.

[0040] FIG12 shows the quantitative data of cytokines in the colon of mice with ulcerative colitis after treatment.

[0041] FIG13 shows the colon length and HE section score data after delayed treatment of ulcerative colitis mice with chimeric selenium-containing Lactobacillus casei.

[0042] FIG14 shows the colon length and HE section score data after chimeric selenium-containing Lactobacillus casei treated TNBS-induced ulcerative colitis mice.

[0043] FIG15 shows the colon length and HE section score data after treating chronic ulcerative colitis mice with chimeric selenium-containing Lactobacillus casei.

[0044] Among them, in Figures 13-15, the columns from left to right are the healthy group, the model group, and the chimeric selenium-dot Lactobacillus casei group.

[0045] FIG16 shows the evaluation of the biosafety of short-term oral administration of chimeric selenium-dot Lactobacillus casei in healthy mice.

[0046] FIG17 shows the evaluation of the biosafety of long-term oral administration of chimeric selenium-dot Lactobacillus casei in healthy mice.

[0047] FIG18 shows the content of reactive oxygen free radicals and the composition of intestinal flora genus levels after the chimeric selenium-dot Lactobacillus casei was used to treat ulcerative colitis cynomolgus monkeys.

[0048] FIG19 shows the colonoscopy results after treatment of ulcerative colitis in cynomolgus monkeys with chimeric selenium-containing Lactobacillus casei.

[0049] FIG20 shows the evaluation of the biosafety of chimeric selenium-dot Lactobacillus casei in cynomolgus monkeys (Note: ND means not detected). DETAILED DESCRIPTION

[0050] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0051] It should be understood that the terms described in the present invention are only for describing particular embodiments and are not intended to limit the present invention. In addition, for numerical ranges in the present invention, it should be understood that the upper and lower limits of the ranges and each intermediate value therebetween are specifically disclosed. Each smaller range between any stated value or intermediate value within a stated range and any other stated value or intermediate value within the stated range is also included in the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0052] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention belongs. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In the event of any conflict with any incorporated document, the content of this specification shall prevail.

[0053] Chimeric Selenium Dot Probiotics

[0054] The first aspect of the present invention provides an engineered probiotic. The probiotic can be any microorganism naturally present in the intestine that can adhere to and colonize the intestine. Research has shown that combining probiotics with nano-selenium not only addresses the poor targeting of nano-selenium but also enhances the antioxidant capacity of the probiotic, achieving a synergistic therapeutic effect of probiotics and nano-selenium for inflammatory diseases.

[0055] The chimeric selenium dot probiotics of the present invention are based on probiotics as the main structure, and utilize their surface to achieve in-situ adsorption of selenium precursors, which are then reduced using a reducing agent to obtain probiotics with selenium dots grown in-situ only on the surface.

[0056] The selenium elemental particle size (average particle size) on the surface of the chimeric selenium dot probiotics of the present invention is 1-100nm, preferably 2-80nm, also preferably 2-50nm, preferably 2-20nm, preferably 2-10nm such as 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50nm etc. The inventors have found that compared to large particle size (for example, greater than 100, 200, 300 or more particle size), nano-selenium within the particle size range of the present invention has more excellent functions, such as antioxidant effect, and the chimeric selenium dot probiotics containing the nano-selenium can show more excellent therapeutic effect for inflammation-related diseases. In the present invention, selenium elemental substance can also be referred to as selenium dot elemental substance.

[0057] In the present invention, the type of probiotic is not particularly limited. In a preferred embodiment, the probiotic is an intestinal probiotic, examples of which include, but are not limited to, at least one or a combination of the following: Lactobacillus, Bifidobacterium, Streptococcus, Bacillus, Clostridium, Bacteroides, Lactococcus, Leuconostoc, Propionibacterium, Pediococcus, and Saccharomyces. Among them, the probiotic of the genus Lactobacillus is preferably selected from at least one of Lactobacillus casei, Lactobacillus bulgaricus, and Lactobacillus lactis. The probiotic of the genus Bifidobacterium is preferably selected from at least one of Bifidobacterium adolescentis, Bifidobacterium longum, and Bifidobacterium infantis. The probiotic of the genus Streptococcus is preferably Streptococcus thermophilus. The probiotic of the genus Bacillus is preferably at least one of Bacillus subtilis and Bacillus licheniformis. The probiotic of the genus Pediococcus is preferably Pediococcus acidilactici. The probiotic of the genus Clostridium is preferably Clostridium butyricum. The probiotic of the genus Bacteroides is preferably Bacteroides fragilis. The probiotic of the genus Lactococcus is preferably Lactococcus lactis. The genus Leuconostoc is preferably Leuconostoc mesenteroides. The genus Propionibacterium is preferably Propionibacterium freudenreichii.

[0058] In a preferred embodiment, the probiotics used in the present invention are Lactobacillus casei, and more preferably Lactobacillus casei with a preservation number of CICC20280, Streptococcus thermophilus with a preservation number of CICC6222, and Pediococcus acidilactici with a preservation number of JCM8797.

[0059] In a preferred embodiment of the present invention, the selenium element located on the surface of the probiotics is obtained by an in situ reduction method, which is described in detail below.

[0060] Preparation method

[0061] The preparation method of the chimeric selenium-dot probiotics of the present invention comprises: (1) co-incubating the cultured probiotics in a solvent containing a selenium precursor to obtain probiotics adsorbed with the selenium precursor; and (2) subjecting the probiotics adsorbed with the selenium precursor to a reduction reaction with a reducing agent to obtain the chimeric selenium-dot probiotics.

[0062] Step (1)

[0063] In step (1) of the present invention, the probiotics are first cultured in a culture medium. The culture medium preferably contains factors that promote polysaccharide production, and then the probiotics are allowed to adsorb the selenium precursor in situ under suitable co-incubation conditions. The appropriate culture medium and culture conditions can be selected according to the specific type of probiotics to be cultured, and are not particularly limited to this, as long as the desired number of probiotics can be achieved.

[0064] In a preferred embodiment, the culture medium used in the present invention is Msgg culture medium. Also preferably, the culture medium contains 0.2-1.5 g / L K2HPO4, 10-40 g / L MOPS, 1-10 g / L glycerol, 1-10 g / L amino acids (glutamic acid, phenylalanine and tryptophan), 0.1-1.5 mg / L B1, and 5-50 mg / L metal salts. Further preferably, the culture medium contains 0.5-1.0 g / L K2HPO4, 15-25 g / L MOPS, 2-8 g / L glycerol, 3-8 g / L amino acids, 0.4-0.6 mg / L B1, and 10-30 mg / L metal salts. The metal salt is selected from at least one or a combination of MgCl2, CaCl2, FeCl3·6H2O, MnCl2·4H2O, and ZnCl2. The amino acids are selected from at least one or a combination of glutamic acid, phenylalanine, and tryptophan.

[0065] In a preferred embodiment of the present invention, the ratio of probiotics to the selenium precursor is 10 7-10 CFU: (0.002-100) mmol, preferably 10 7-10 CFU: (0.002-10) mmol, wherein, preferably, the number of probiotics is preferably 10 8-10 CFU, preferably 10 9 CFU, the selenium precursor is preferably 0.002-10mmol, and more preferably 0.006-10mmol. Furthermore, the ratio of probiotics to the selenium precursor is preferably 10 7-10 CFU: (0.006-1) mmol, more preferably 10 7-10 CFU: (0.006-0.2) mmol, for example, 0.006, 0.008, 0.010, 0.012, 0.02, 0.1, 0.15, 0.2 mmol, etc.

[0066] In a preferred embodiment of the present invention, any high-valent selenium precursor compound that can obtain nano-selenium element through reduction reaction can be used in the present invention. The high-valent selenium precursor can be a divalent, tetravalent, or hexavalent selenium precursor compound, and can be organic selenium and / or inorganic selenium. Examples include but are not limited to: sodium selenite, potassium selenite, sodium selenate, potassium selenate, selenium dioxide, selenium oxide, selenomethionine, hydroxyselenomethionine, selenocysteine, selenocystine, selenomethylselenocysteine, selenomethylselenomethionine, or any one or a combination of at least two of seaweed selenium polysaccharides.

[0067] In a preferred embodiment of the present invention, the type of solvent is not particularly limited, and examples include, but are not limited to, any one or a combination of at least two of physiological saline, buffer (e.g., phosphate buffer solution), M17 medium, MRS medium, LB medium, BH medium, Bifidobacterium BS medium and Roche broth medium.

[0068] In a preferred embodiment of the present invention, the co-incubation temperature is 4-45°C, preferably 4-40°C, and further preferably 36-38°C, for example 36°C, 36.5°C, 37°C, 37.5°C, 38°C, etc. The co-incubation temperature should not be too high or too low. The inventors have found that co-incubation at the above temperature, especially 36-38°C, can enable the selenium precursor to be adsorbed on the surface of the probiotics as quickly as possible. If the co-incubation temperature is high, it is not conducive to the survival of the probiotics. If the co-incubation temperature is low, the therapeutic effect (including antioxidant effect, etc.) of the chimeric selenium dot probiotics is poor.

[0069] In the present invention, the co-incubation time is not particularly limited and can be adjusted as needed, but is preferably 1-12 h, preferably 1-8 h, for example, 1 h, 2 h, 3 h, 4 h, 5 h, 6 h, 7 h, 8 h, etc.

[0070] In the present invention, the pH of the co-incubation is not particularly limited and can be adjusted as needed, but is preferably 3-11, preferably 4-8, for example, 4, 5, 6, 7, 8, etc.

[0071] In addition, the probiotics co-cultured with the selenium precursor are preferably probiotics that do not contain free polysaccharides. Therefore, the method of the present invention further includes a step of pre-treating the cultured probiotics to remove free exopolysaccharides. The pre-treatment method is not particularly limited, and for example, free exopolysaccharides can be removed by low-speed centrifugation, wherein low speed means that the centrifugal force is not higher than 6000g, preferably not higher than 3000g.

[0072] In a preferred embodiment of the present invention, the order of adding the solvent, the probiotics and the selenium precursor is not particularly limited. Preferably, the probiotics can be mixed with the solvent first and then with the selenium precursor.

[0073] Step (2)

[0074] Step (2) of the present invention is a step of obtaining the chimeric selenium dot probiotics by surface in situ reduction using a reducing agent. The type of reducing agent is not particularly limited, and examples thereof include but are not limited to any one or a combination of ascorbic acid, glutathione, sodium borohydride, and polyphenols.

[0075] In a preferred embodiment of the present invention, in order to obtain selenium dots or nano-selenium with a fixed particle size, the molar ratio of the reducing agent to the selenium atoms in the selenium precursor is preferably (1-320):1, further preferably (1-100):1, further preferably (1-80):1, and more preferably (1-60):1, for example 1:1, 4:1, 8:1, 10:1, 20:1, 40:1, 60:1, etc.

[0076] In a preferred embodiment of the present invention, the temperature of the reduction reaction is 4-40°C, preferably 5-35°C, for example 5°C, 10°C, 15°C, 20°C, 25°C, 30°C, 35°C, etc.

[0077] In a preferred embodiment of the present invention, the reduction reaction time is 0.5-24 h, preferably 1-10 h, more preferably 2-10 h, for example 2 h, 3 h, 4 h, 5 h, 6 h, 7 h, 8 h, 9 h, 10 h, etc.

[0078] In a preferred embodiment of the present invention, the pH of the reduction reaction is 3-11, preferably 4-8, and more preferably 5-8, such as 5, 6, 7, 8, etc.

[0079] In addition, the present invention also uses infrared spectroscopy and nuclear magnetic resonance hydrogen spectrum to characterize the interaction between hydroxyl groups and selenium points in polysaccharides on the surface of Lactobacillus casei, confirming that hydroxyl groups are ligands of nano-selenium and proving that selenium can disrupt the hydrogen bonding system of sugars.

[0080] use

[0081] One aspect of the present invention provides a pharmaceutical composition comprising the chimeric selenium-dot probiotics of the present invention.

[0082] In one aspect of the present invention, a dietary supplement is provided, comprising the chimeric selenium-dot probiotics of the present invention.

[0083] One aspect of the present invention provides a feed additive containing the chimeric selenium-dot probiotics of the present invention.

[0084] The present invention also provides a use of the chimeric selenium-dot probiotic in the preparation of the following medicines, dietary supplements or feed additives, wherein the medicines, dietary supplements or feed additives can be:

[0085] (1) Prevent, improve, alleviate or treat intestinal inflammatory diseases;

[0086] (2) Drugs for preventing, improving, alleviating or treating nasal inflammatory diseases;

[0087] (3) Increase or improve the efficiency of scavenging reactive oxygen free radicals;

[0088] (4) increasing the diversity of intestinal flora or improving the abundance of probiotics;

[0089] (5) regulating immune cells associated with intestinal inflammation, preferably restoring the immune cells associated with intestinal inflammation to normal levels;

[0090] (6) Regulating intestinal inflammation-related cytokines, preferably promoting the expression or amount of anti-inflammatory cytokines, and / or inhibiting the expression or amount of pro-inflammatory cytokines.

[0091] Those skilled in the art will appreciate that the chimeric selenium dot probiotics of the present invention can be used for diagnostic or therapeutic purposes, or for non-diagnostic or therapeutic purposes. Based on this, the present invention also provides a method for increasing or improving the efficiency of scavenging reactive oxygen free radicals in vitro, or regulating inflammation-related cytokines in vitro, comprising the step of contacting the chimeric selenium dot probiotics of the present invention with cells.

[0092] As used herein, the terms "prevent, ameliorate, alleviate, or treat" refer to improving a condition before or after the onset of a disease or disorder. This reduction or prevention is achieved by at least 5%, 10%, 20%, 40%, 50%, 60%, 80%, 90%, 95%, or 100% as measured by any standard technique compared to an untreated control group under the same conditions. As used herein, the term "treat" refers to both therapeutic treatment and prophylactic or preventative measures aimed at preventing or slowing down (reducing) undesirable physiological changes or disorders, such as reducing the risk of developing an inflammatory disease. Beneficial or desired clinical outcomes include, but are not limited to, alleviation of symptoms, reduction in disease severity, stabilization of the disease state (i.e., non-worsening), delay or slowing of disease progression, amelioration or alleviation of the disease state, and alleviation (whether partial or complete) of the disease state, whether detectable or undetectable. Those in need of treatment include those already suffering from the disease or disorder, as well as those susceptible to developing the disease or disorder, or those in need of prevention of the disease or disorder.

[0093] Example 1

[0094] The following is an exemplary description using Lactobacillus casei as a probiotic.

[0095] 1. Preparation of Chimeric Selenium-dot Lactobacillus casei

[0096] (1) Lactobacillus casei (CICC20280) cultured overnight in Msgg medium was centrifuged at low speed to remove polysaccharides in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of Lactobacillus casei to selenium precursor was 10:1. 9 CFU: 0.012 mmol sodium selenite was added and incubated at 37°C for 3 h to obtain Lactobacillus casei adsorbed with selenium precursors;

[0097] (2) Ascorbic acid was added according to the molar ratio of the reducing agent to the selenium atoms in the selenium precursor being 40:1, and the reduction reaction was carried out at 37°C for 2 hours to obtain the chimeric selenium-dot Lactobacillus casei.

[0098] 2. Performance test of chimeric selenium-dot Lactobacillus casei

[0099] The micromorphology of the chimeric selenium-dot Lactobacillus casei was observed using a transmission electron microscope. Fig. 1 is a transmission electron microscope image of the chimeric selenium-dot Lactobacillus casei obtained in Example 1, wherein the embedded image is a selenium-dot lattice, and Fig. 2 is an elemental analysis spectrum of the chimeric selenium-dot Lactobacillus casei (the arrows indicate the energy spectrum position of the selenium element). As can be seen from Figs. 1 and 2, the preparation method provided by the present invention can successfully grow selenium dots on the surface of Lactobacillus casei in situ, i.e., obtain the product chimeric selenium-dot Lactobacillus casei. Simultaneously, as can be seen from the figure, the average particle size of the selenium dots is about 5 nm.

[0100] Example 2

[0101] Preparation of Chimeric Selenium-dot Lactobacillus casei:

[0102] (1) Lactobacillus casei (CICC20280) cultured overnight in Msgg medium was centrifuged at low speed to remove polysaccharides in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of Lactobacillus casei to selenium precursor was 10:1. 9 CFU: 0.006 mmol sodium selenite was added and incubated at 37°C for 3 h to obtain Lactobacillus casei adsorbed with selenium precursors;

[0103] (2) Ascorbic acid was added according to the molar ratio of the reducing agent to the selenium atoms in the selenium precursor being 40:1, and the reduction reaction was carried out at 37°C for 2 hours to obtain the chimeric selenium-dot Lactobacillus casei.

[0104] Example 3

[0105] Preparation of Chimeric Selenium-dot Lactobacillus casei:

[0106] (1) Lactobacillus casei (CICC20280) cultured overnight in Msgg medium was centrifuged at low speed to remove polysaccharides in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of Lactobacillus casei to selenium precursor was 10:1. 9 CFU: 0.02 mmol sodium selenite was added and incubated at 37°C for 3 h to obtain Lactobacillus casei adsorbed with selenium precursors;

[0107] (2) Ascorbic acid was added according to the molar ratio of the reducing agent to the selenium atoms in the selenium precursor being 40:1, and the reduction reaction was carried out at 37°C for 2 hours to obtain the chimeric selenium-dot Lactobacillus casei.

[0108] Example 4

[0109] Preparation of Chimeric Selenium-dot Lactobacillus casei:

[0110] (1) Lactobacillus casei (CICC20280) cultured overnight in Msgg medium was centrifuged at low speed to remove polysaccharides in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of Lactobacillus casei to selenium precursor was 10:1. 9 CFU: 0.012 mmol sodium selenite was added and incubated at 40°C for 3 h to obtain Lactobacillus casei adsorbed with selenium precursors;

[0111] (2) Ascorbic acid was added according to the molar ratio of the reducing agent to the selenium atoms in the selenium precursor being 40:1, and the reduction reaction was carried out at 37°C for 2 hours to obtain the chimeric selenium-dot Lactobacillus casei.

[0112] Example 5

[0113] Preparation of chimeric selenium-dot Streptococcus thermophilus and chimeric selenium-dot Pediococcus acidilactici:

[0114] (1) The thermophilic Streptococcus (CICC6222) or the acidococcus (JCM8797) cultured in Msgg medium overnight were centrifuged at low speed to remove the polysaccharide in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of thermophilic Streptococcus or acidococcus to selenium precursor was 10:1. 9 CFU: 0.012 mmol sodium selenite was added and incubated at 37°C for 3 h to obtain Streptococcus thermophilus or Pediococcus acidilactici that adsorbed selenium precursors;

[0115] (2) Ascorbic acid was added at a molar ratio of 40:1 between the reducing agent and the selenium atoms in the selenium precursor, and the reduction reaction was carried out at 37° C. for 2 h to obtain chimeric selenium-dot Streptococcus thermophilus and chimeric selenium-dot Pediococcus acidilactici. The microscopic morphology of the chimeric selenium-dot Lactobacillus casei was observed using a transmission electron microscope. FIG3 is a transmission electron microscope image of the chimeric selenium-dot Streptococcus thermophilus and chimeric selenium-dot Pediococcus acidilactici obtained in Example 5. The preparation method provided by the present invention can successfully grow selenium dots in situ on the bacterial surface, i.e., obtain the chimeric selenium-dot Streptococcus thermophilus or chimeric selenium-dot Pediococcus acidilactici as products.

[0116] Example 6

[0117] Preparation of chimeric selenium-containing Lactobacillus casei using selenomethionine as a selenium precursor:

[0118] (1) Lactobacillus casei (CICC20280) cultured overnight in Msgg medium was centrifuged at low speed to remove polysaccharides in the solution. After centrifugation at 3000×g for 5 min, the supernatant was discarded and the cells were resuspended in physiological saline solution. The ratio of Lactobacillus casei to selenium precursor was 10:1. 9 CFU: 0.1 mmol of selenomethionine was added, and the pH of the solution was then adjusted to 7.0. After incubation at 37°C for 3 h, Lactobacillus casei adsorbed with selenium precursors was obtained;

[0119] (2) Ascorbic acid was added according to a molar ratio of the reducing agent to the selenium atoms in the selenium precursor of 60:1, and then the pH of the solution was adjusted to 6.0, and a reduction reaction was carried out at 37°C for 9 hours to obtain a chimeric selenium-dot Lactobacillus casei. The microscopic morphology of the chimeric selenium-dot Lactobacillus casei was observed using a transmission electron microscope. Figure 4 is a transmission electron microscope image of the chimeric selenium-dot Lactobacillus casei obtained in Example 6. The preparation method provided by the present invention can use selenomethionine as a precursor to successfully grow selenium dots on the bacterial surface in situ, that is, to obtain the product chimeric selenium-dot Lactobacillus casei.

[0120] Comparative Example 1

[0121] Preparation of nano-selenium:

[0122] (1) Sodium selenite (the same mass as in Example 1) was added to a phosphate buffer solution and placed at 37°C for 3 h. Ascorbic acid was then added at a molar ratio of the reducing agent to the selenium atoms in the selenium precursor of 40:1, and the reduction reaction was carried out at room temperature for 12 h to obtain nano-selenium.

[0123] Comparative Example 2

[0124] Preparation of the mixed solution of nano-selenium and Lactobacillus casei:

[0125] The Lactobacillus casei provided in Example 1 was mixed with the nano-selenium provided in Comparative Example 1 to obtain a mixed solution of nano-selenium and Lactobacillus casei provided in this comparative example (the amount of the solvent used was the same as that in Example 1).

[0126] Comparative Example 3

[0127] This comparative example provides a kind of isolated lactobacillus casei secretion exopolysaccharide, and is used for the synthesis of nano-selenium.Described lactobacillus casei secretion exopolysaccharide concrete extraction method: with lactobacillus casei (CICC20280) culture fluid boiling water bath inactivation (30min), be cooled to room temperature, centrifugal (12000 × g, 30min) remove impurities such as thalline and enzymolysis protein.Get supernatant subsequently, adding 80.0% trichloroacetic acid solution while stirring to final content is 4.0%, 4 ℃ of standing overnight back centrifugal (12000 × g, 30min) remove remaining denatured protein.Get supernatant while stirring vigorously while adding 3 times of volume 95% ice ethanol, 4 ℃ of standing 24h, once more centrifugal (12000 × g, 30min) get precipitation, with suitable deionized water, it is redissolved, solution dialyzed 48h (dialysis bag with 8-14kDa molecular weight cut-off) every 8h change 1 water, dialyzate is following used polysaccharide solution. Sodium selenite (the same mass as in Example 1) was added to the polysaccharide solution and placed at 37° C. for 2 h. Ascorbic acid was then added at a molar ratio of 40:1 between the reducing agent and the selenium atoms in the selenium precursor, and the reduction reaction was carried out at room temperature for 2 h to obtain a connection product of extracellular polysaccharide and nano-selenium.

[0128] Test Case

[0129] The performance tests of the chimeric selenium-dot Lactobacillus casei prepared in Examples 1-4 are shown below.

[0130] (1) Antioxidant activity test of chimeric selenium-dot Lactobacillus casei

[0131] The ability to scavenge the classic free radical DPPH (1,1-diphenyl-2-trinitrophenylhydrazine) was used as a representative test: 50 μg / mL of the chimeric selenium-dot Lactobacillus casei was reacted with 0.5 mmol / L of a purple free radical DPPH solution. The absorbance change at 526 nm after the entire reaction system reacted for 30 minutes was measured, and the scavenging rate of the DPPH free radical was calculated. The results are shown in Table 1:

[0132] Table 1

[0133] Note: Control group 1 is the untreated control group (containing only Lactobacillus casei).

[0134] As can be seen from Table 1, the chimeric selenium-dot Lactobacillus casei provided by the present invention has excellent antioxidant activity. As can be seen from the comparison of Examples 1-3 and Comparative Example 1, the ratio of Lactobacillus casei to selenium precursor in the present invention has a better effect within a certain range; as can be seen from the comparison of Examples 1 and 4, the present invention is co-incubated at 36-40°C, and the chimeric selenium dots on the surface of Lactobacillus casei have a good antioxidant effect. As can be seen from the comparison of Example 1 and Comparative Examples 1 and 2, the preparation method provided by the present invention can obtain a chimeric selenium-dot Lactobacillus casei with excellent antioxidant effect.

[0135] (2) In vitro growth ability of the chimeric selenium-dot Lactobacillus casei prepared in Example 1

[0136] 1×10 9 CFU of chimeric selenium-containing Lactobacillus casei with 1×10 9 CFU of Lactobacillus casei were added to a culture tube containing MRS liquid medium, and the absorbance at 600 nm of the culture solution at different incubation times was measured, and a growth curve was plotted based on this. As shown in Figure 5, the growth ability of Lactobacillus casei was not affected after the chimeric selenium dots were added.

[0137] (3) Gastric acid environment tolerance of the chimeric selenium-dot Lactobacillus casei prepared in Example 1

[0138] A certain amount of chimeric selenium-dot Lactobacillus casei and Lactobacillus casei were added to a simulated gastric acid solution and co-incubated. Appropriate amounts of the bacterial suspension were diluted at different time points and then spread onto pre-prepared plates (MRS solid medium). The plates were then inverted and incubated for 24 hours at 37°C for counting. As shown in Figure 6, 1% of the chimeric selenium-dot Lactobacillus casei bacteria survived the 4-hour co-incubation, while the Lactobacillus casei itself had been inactivated.

[0139] (4) Intestinal adhesion ability of the chimeric selenium-dot Lactobacillus casei prepared in Example 1

[0140] Female BALB / c mice aged 5-6 weeks were used. The abdominal hair of the mice was removed with depilatory cream and 1×10 8 Fluorescence imaging of CFU of chimeric selenium-dot Lactobacillus casei was performed at different time points. As shown in Figure 7, the fluorescence intensity in the abdomen of mice in the chimeric selenium-dot Lactobacillus casei group was significantly higher than that in the Lactobacillus casei group within 4-6 hours, demonstrating that the chimeric selenium-dot Lactobacillus casei adhered well to and remained in the intestines, which helped alleviate intestinal inflammation.

[0141] (5) Therapeutic effect of chimeric selenium-dot Lactobacillus casei in ulcerative colitis model mice

[0142] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days, and drinking water was replaced with an aqueous solution (w / v) of 3% dextran sulfate sodium (DSS); on the 1st to 7th day of modeling, mice were given different treatments (respectively PBS, Lactobacillus casei, a mixed solution of nano-selenium and Lactobacillus casei prepared in the above-mentioned comparative example 2, and the chimeric selenium-dot Lactobacillus casei prepared in the above-mentioned embodiment 1); within the 9-day period of modeling, the weight of the mice was recorded every day. On the 9th day of modeling, the mice were killed, and the colon tissue of the mice was taken out and the length was measured for analysis. As shown in Figure 8, the results show that the chimeric selenium-dot Lactobacillus casei provided by the present invention is used for treatment, which is conducive to alleviating inflammation in ulcerative colitis mice and alleviating intestinal damage, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a good therapeutic effect in, for example, inflammatory disease models.

[0143] (6) Effect of chimeric selenium-dot Lactobacillus casei on the recovery of intestinal flora in ulcerative colitis model mice

[0144] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days, and drinking water was replaced with an aqueous solution of 3% DSS (w / v); on the 1st to 7th day of modeling, mice were given different treatments (respectively, PBS, Lactobacillus casei, a mixed solution of nano-selenium and Lactobacillus casei prepared in the above-mentioned comparative example 2, and a chimeric selenium-dot Lactobacillus casei prepared in the above-mentioned embodiment 1); on the 9th day of modeling, mouse feces were collected for 16SrRNA sequencing analysis. As shown in Figure 9, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, which was beneficial to alleviating the imbalanced intestinal flora in ulcerative colitis mice, increasing the diversity of intestinal flora, and improving the abundance of probiotics, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a good recovery effect on the imbalanced intestinal flora.

[0145] (7) Effect of chimeric selenium-dot Lactobacillus casei on scavenging reactive oxygen free radicals in ulcerative colitis model mice

[0146] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days, and drinking water was replaced with an aqueous solution of 3% DSS (w / v); on the 1st to 7th day of modeling, mice were given different treatments (respectively PBS, Lactobacillus casei, a mixed solution of nano-selenium and Lactobacillus casei prepared in the above-mentioned comparative example 2, and a chimeric selenium-dot Lactobacillus casei prepared in the above-mentioned embodiment 1); on the 9th day of modeling, the mouse intestine was subjected to vital staining for 30 minutes using Dihydroethidium dye, and then the colon of the mouse was removed and observed and photographed under a two-photon microscope. As shown in Figure 10, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, which was conducive to removing the reactive oxygen free radicals excessively produced in the ulcerative colitis mice, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention had an excellent scavenging effect on reactive oxygen free radicals in vivo.

[0147] (8) Effect of Chimeric Selenium-dot Lactobacillus casei on Immune Cell Regulation in Ulcerative Colitis Model Mice

[0148] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days, and drinking water was replaced with an aqueous solution of 3% DSS (w / v); on the 1st to 7th day of modeling, mice were given different treatments (respectively, PBS, Lactobacillus casei, a mixture of nano-selenium and Lactobacillus casei prepared in the above-mentioned comparative example 2, and a chimeric selenium-dot Lactobacillus casei prepared in the above-mentioned embodiment 1); on the 9th day of modeling, the mice were dissected to remove the colon and separate the lamina propria cells for flow cytometry staining. As shown in Figure 11, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, and the proportion of anti-inflammatory Treg cells increased, while the proportion of pro-inflammatory Th17 and Macrophage cells decreased, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a regulatory effect on intestinal inflammation-related immune cells.

[0149] (9) Effect of Chimeric Selenium-dot Lactobacillus casei on Cytokine Regulation in Ulcerative Colitis Model Mice

[0150] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days, and drinking water was replaced with an aqueous solution of 3% DSS (w / v); on the 1st to 7th day of modeling, mice were given different treatments (respectively, PBS, Lactobacillus casei, a mixture of nano-selenium and Lactobacillus casei prepared in the above-mentioned comparative example 2, and a chimeric selenium-dot Lactobacillus casei prepared in the above-mentioned embodiment 1); on the 9th day of modeling, the mice were dissected to remove the colon and homogenized to extract protein, and cytokine ELISA was performed. As shown in Figure 12, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, and the anti-inflammatory cytokines IL-10 and TGF-β increased, and the pro-inflammatory TNF-α, IL-6, and IL-1β ratios decreased, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a regulating effect on intestinal inflammation-related cytokines.

[0151] (10) Effect of Chimeric Selenium-Dot Lactobacillus Casei in Delayed Treatment of Ulcerative Colitis Model

[0152] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days. On days 1-7 of modeling, their drinking water was replaced with a 3% DSS aqueous solution (w / v). On days 8-11, the mice were given different treatments (PBS and the chimeric selenium-dot Lactobacillus casei prepared in Example 1). On day 12, the mice were killed, and their colon tissue was removed and measured for length analysis. The results are shown in Figure 13. Treatment with the chimeric selenium-dot Lactobacillus casei provided by the present invention is beneficial for alleviating inflammation in mice with ulcerative colitis and reducing intestinal damage, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a good therapeutic effect in, for example, inflammatory disease models.

[0153] (11) Therapeutic effect of chimeric selenium-dot Lactobacillus casei in TNBS-induced ulcerative colitis model

[0154] Female c57BL / 6 mice aged 5-6 weeks were adaptively raised for 7 days. After anesthesia on the 0th day of modeling, the mice were haired and subjected to percutaneous absorption of TNBS solution. On the 7th day, TNBS solution was administered rectally. From the 8th to the 11th day, mice were given different treatments (PBS and the chimeric selenium-dot Lactobacillus casei prepared in Example 1, respectively). On the 12th day, mice were killed, and the colon tissue of the mice was taken out and the length was measured for analysis. As shown in Figure 14, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, which was conducive to alleviating inflammation in ulcerative colitis mice and alleviating intestinal damage, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a good therapeutic effect in, for example, inflammatory disease models.

[0155] (12) Therapeutic effect of chimeric selenium-dot Lactobacillus casei in a chronic ulcerative colitis model

[0156] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days. During the first, third, and fifth weeks of modeling, their drinking water was replaced with a 3% DSS aqueous solution (w / v). During the second, fourth, and sixth weeks, the mice were given drinking water and different treatments (PBS and the chimeric selenium-dot Lactobacillus casei prepared in Example 1, respectively). On the 43rd day, the mice were sacrificed, and their colon tissue was removed and measured for length analysis. The results, as shown in Figure 15, show that treatment with the chimeric selenium-dot Lactobacillus casei provided by the present invention is beneficial for alleviating inflammation in mice with ulcerative colitis and reducing intestinal damage, indicating that the chimeric selenium-dot Lactobacillus casei provided by the present invention has a good therapeutic effect in, for example, inflammatory disease models.

[0157] (13) Evaluation of the biosafety of chimeric selenium-containing Lactobacillus casei in mice after short-term / long-term oral administration

[0158] Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days and given drinking water and the chimeric selenium-dot Lactobacillus casei prepared in Example 1 for one week; on the 8th day, the mice were killed, and the whole blood, heart, liver, spleen, lung, and kidney of the mice were taken and physiological and pathological tests were performed. Female BALB / c mice aged 5-6 weeks were adaptively raised for 7 days and given drinking water and the chimeric selenium-dot Lactobacillus casei prepared in Example 1 for four weeks; on the 29th day, the mice were killed, and the whole blood, heart, liver, spleen, lung, and kidney of the mice were taken and physiological and pathological tests were performed. The results are shown in Figures 16 and 17. When the chimeric selenium-dot Lactobacillus casei provided by the present invention was orally administered, all the indicators of the mice were within the normal range.

[0159] (14) Effect of chimeric selenium-dot Lactobacillus casei on scavenging reactive oxygen free radicals in cynomolgus monkeys with ulcerative colitis

[0160] Male cynomolgus monkeys aged 4 to 4.5 years old and weighing 4.5 ± 0.5 kg were kept in cages alone with free access to food and water. On the 1st to 10th day of modeling, each cynomolgus monkey was gavaged with 50 mL of an aqueous solution of 2.5% DSS (w / v) every day. On the 2nd to 10th day of modeling, different treatments (PBS and the chimeric selenium-dot Lactobacillus casei prepared in Example 1) were given to the cynomolgus monkeys. On the 11th day after the treatment, the feces of the cynomolgus monkeys were analyzed for the detection of the content of reactive oxygen species and 16S rRNA sequencing. As shown in Figure 18, the chimeric selenium-dot Lactobacillus casei provided by the present invention was used for treatment, which was conducive to removing the reactive oxygen species excessively produced in ulcerative colitis cynomolgus monkeys, and was conducive to alleviating the imbalanced intestinal flora in ulcerative colitis mice, increasing intestinal flora diversity, and improving the abundance of probiotics. This showed that the chimeric selenium-dot Lactobacillus casei provided by the present invention had an excellent scavenging effect on reactive oxygen species in vivo, and had a good recovery effect on the imbalanced intestinal flora.

[0161] (15) Effect of chimeric selenium-dot Lactobacillus casei on improving intestinal ulcers in cynomolgus monkeys with ulcerative colitis

[0162] Male cynomolgus monkeys aged 4 to 4.5 years and weighing 4.5 ± 0.5 kg were individually caged with free access to food and water. On days 1 to 10 of modeling, each cynomolgus monkey was gavaged with 50 mL of a 2.5% DSS aqueous solution (w / v) daily. On days 2 to 10 of modeling, the monkeys were given different treatments (PBS and the chimeric selenium-doped Lactobacillus casei prepared in Example 1, respectively). On the 11th day after the completion of treatment, the monkeys were subjected to a colonoscopy. As shown in Figure 19, treatment with the chimeric selenium-doped Lactobacillus casei provided by the present invention was beneficial to improving symptoms such as bleeding and ulcers in the intestinal cavity of the cynomolgus monkeys.

[0163] (16) Safety evaluation of chimeric selenium-containing Lactobacillus casei in the treatment of ulcerative colitis in cynomolgus monkeys

[0164] Male cynomolgus monkeys aged 4 to 4.5 years old and weighing 4.5 ± 0.5 kilograms were kept in cages alone with free access to food and water. On the 1st to 10th day of modeling, each cynomolgus monkey was gavaged with an aqueous solution (w / v) of 50mL 2.5% DSS every day. On the 2nd to 10th day of modeling, different treatments (PBS and the chimeric selenium-dot Lactobacillus casei prepared by the above-described embodiment 1) were given to the cynomolgus monkeys. The situation of the cynomolgus monkeys after treatment was observed every day. On the 11th day after treatment, the blood of the cynomolgus monkeys was taken for physiological testing. As shown in Figure 20, the results showed that the chimeric selenium-dot Lactobacillus casei provided by the present invention was taken orally, and the cynomolgus monkeys had no adverse oral reactions, and the blood parameters tested were all within the normal range.

[0165] Although the present invention has been described with reference to exemplary embodiments, it should be understood that the invention is not limited to the disclosed exemplary embodiments. Various modifications and variations may be made to the exemplary embodiments of the present specification without departing from the scope or spirit of the present invention. The scope of the claims is to be given the broadest interpretation so as to encompass all such modifications and equivalent structures and functions.

Claims

1. A chimeric selenium-dot probiotic, characterized in that: The invention comprises probiotics and selenium element which grows in situ on the surface of the probiotics.

2. The chimeric selenium-dot probiotic according to claim 1, characterized in that The probiotics include intestinal probiotics.

3. The chimeric selenium-dot probiotic according to claim 2, characterized in that The intestinal probiotics include at least one of Lactobacillus, Bifidobacterium, Streptococcus, Bacillus, Clostridium, Bacteroides, Lactococcus, Leuconostoc, Propionibacterium, Pediococcus and Saccharomyces; Preferably, the intestinal probiotics are Lactobacillus casei, Streptococcus thermophilus or Pediococcus acidilactici.

4. The chimeric selenium-dot probiotic according to claim 3, characterized in that: The selenium element has a particle size of 1-100 nm.

5. A method for preparing a chimeric selenium-dot probiotic, characterized in that: include: (1) culturing probiotics in a culture medium, and then co-incubating the cultured probiotics in a solvent containing a selenium precursor to obtain probiotics adsorbed with the selenium precursor; (2) subjecting the probiotics adsorbed with selenium precursor to a reduction reaction with a reducing agent to obtain the chimeric selenium-dot probiotics.

6. The method for preparing the chimeric selenium-dot probiotics according to claim 5, characterized in that: The co-incubation temperature is 4-45°C.

7. The method for preparing the chimeric selenium-dot probiotics according to claim 6, characterized in that: The ratio of the probiotics to the selenium precursor is 10 7-10 CFU: (0.002-100) mmol, preferably 10 7-10 CFU: (0.002-10)mmol.

8. The method for preparing the chimeric selenium-dot probiotics according to claim 7, characterized in that: The selenium precursor includes organic selenium and / or inorganic selenium; Preferably, the selenium precursor is selected from any one or a combination of sodium selenite, potassium selenite, sodium selenate, potassium selenate, selenium dioxide, selenium oxide, selenomethionine, hydroxyselenomethionine, selenocysteine, selenocysteine, selenomethylselenocysteine, selenomethylselenomethionine or seaweed selenium polysaccharide; Preferably, the solvent is selected from at least one of physiological saline, phosphate buffer solution, M17 medium, MRS medium, LB medium, BH medium, Bifidobacterium BS medium and Roche broth medium.

9. The method for preparing the chimeric selenium-dot probiotics according to claim 8, characterized in that: The reducing agent includes any one or a combination of ascorbic acid, glutathione, sodium borohydride, hydrazine hydrate, catechin, hydroquinone, gallic acid, quercetin and resveratrol; Preferably, the temperature of the reduction reaction is 4-40°C; Preferably, the reduction reaction time is 0.5-24h; Preferably, the pH of the reduction reaction is 3-11.

10. Use of the chimeric selenium-dot probiotic according to any one of claims 1 to 9 in the preparation of a medicine, dietary supplement or feed additive for preventing, improving or treating inflammatory diseases.

11. A pharmaceutical composition or dietary supplement comprising the chimeric selenium-dot probiotic according to any one of claims 1 to 10.

12. A feed additive comprising the chimeric selenium-dot probiotic according to any one of claims 1 to 10.

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