Bifidobacterium longum extracellular vesicles and preparation method and application thereof

Abstract

The present invention discloses extracellular vesicles of Bifidobacterium longum, as well as a preparation method and application thereof. The extracellular vesicles of Bifidobacterium longum are extracted from the culture medium of Bifidobacterium longum XZY01. The extracellular vesicles of Bifidobacterium longum prepared by the present invention can effectively inhibit the proliferation of human colorectal cancer cells HT29 and SW480 and the murine colon cancer cell line MC38, while inducing the production of reactive oxygen species in tumor cells, while having no effect on normal cells. It is expected that the extracellular vesicles of Bifidobacterium longum according to the present invention will be widely used as a drug with low side effects for alleviating, preventing, or treating cancer.

Description

Technical Field

[0001] The present invention belongs to the field of biotechnology, and in particular relates to an extracellular vesicle of Bifidobacterium longum and a preparation method and application thereof. Background Art

[0002] Colorectal cancer (CRC) is a major disease that seriously threatens human life and health. In the early stages of colon cancer, there may be no symptoms. In the middle and late stages, the disease may manifest as abdominal distension and indigestion, followed by changes in bowel habits, abdominal pain or blood in the stool. The treatment of colorectal cancer is currently mainly based on surgery, with radiotherapy, chemotherapy and immunotherapy as postoperative adjuvants. However, the prognosis of patients is poor, and it has serious side effects and toxicity, including nausea, vomiting, oral ulcers, diarrhea, hepatotoxicity, bone marrow suppression, immunosuppression, etc., which seriously affect the quality of life of patients. Therefore, the development of new tumor treatment methods is of great significance to the treatment of cancer patients.

[0003] The intestinal flora refers to the community of microorganisms that reside in the human intestine. It participates in important physiological processes of the host, such as metabolism and immunity. A large number of studies have found that the intestinal flora is involved in the progression of colon cancer. Intestinal microorganisms play a key role in regulating the body's immune response. Disturbances in the intestinal microbiome often lead to a variety of diseases, including cancer. Probiotics are a type of active microorganism that is beneficial to the host by colonizing in the human body and changing the composition of the microbiome in a certain part of the host. It is well known that probiotics can change the composition of the intestinal microbiome and trigger a strong immune response in the host, thereby more effectively fighting cancer.

[0004] Bacterial extracellular vesicles (EVs) are nanoscale phospholipid bilayer particles released by bacteria that encapsulate nucleic acids, proteins, lipids, and other molecules. They are a key product of bacterial life. As research into the functions of the human microbiome in health and disease has shown, related studies have found that bacterial EVs play an important role in anti-tumor therapy. Therefore, exploring the role of bacterial EVs, especially probiotic EVs, in anti-tumor therapy has important clinical significance and translational value. Summary of the Invention

[0005] The present invention aims to solve at least one of the technical problems existing in the above-mentioned prior art. To this end, the present invention provides an extracellular vesicle of Bifidobacterium longum.

[0006] The present invention also provides a method for preparing the extracellular vesicles of Bifidobacterium longum.

[0007] The present invention also provides an application of the extracellular vesicles of Bifidobacterium longum.

[0008] According to one aspect of the present invention, an extracellular vesicle of Bifidobacterium longum is provided. The extracellular vesicle of Bifidobacterium longum is extracted from the culture solution of Bifidobacterium longum XZY01.

[0009] In some embodiments of the present invention, the diameter of the vesicle is 20-500 nm.

[0010] In some embodiments of the invention, the diameter of the vesicle is 80-90 nm.

[0011] According to a second aspect of the present invention, a method for preparing extracellular vesicles of Bifidobacterium longum is proposed. The method comprises the following steps: ultracentrifuging a culture solution of Bifidobacterium longum XZY01.

[0012] In some embodiments of the present invention, obtaining the culture fluid comprises adding Bifidobacterium longum to a fermentation medium, culturing for 24 to 60 hours, separating the solid and the liquid, and collecting the culture fluid.

[0013] In some embodiments of the present invention, the fermentation medium comprises MRS liquid medium.

[0014] In some embodiments of the present invention, the solid-liquid separation method includes centrifugation.

[0015] In some embodiments of the present invention, the centrifugation conditions are 3000-6000 g, 2-6° C., and 40-60 min.

[0016] In some embodiments of the present invention, the culture solution further comprises a filtration step before ultracentrifugation.

[0017] In some embodiments of the present invention, the filtration is performed using a 0.2-0.6 μm filter membrane.

[0018] In some embodiments of the present invention, the ultracentrifugation conditions are 100,000-1,200,000 g, 2-6° C., and centrifugation for 60-80 min.

[0019] According to a third aspect of the present invention, a use of the extracellular vesicles of Bifidobacterium longum is proposed, wherein the use is in preparing a product for treating tumors.

[0020] In some embodiments of the present invention, the tumor includes at least one of colorectal cancer, testicular cancer, colorectal cancer, small intestine cancer, large intestine cancer, gastric cancer, esophageal cancer, hypopharyngeal cancer, laryngeal cancer, oral cancer, nasal cancer, pancreatic cancer, liver cancer, lung cancer, breast cancer, ovarian cancer, cervical cancer, endometrial cancer, vaginal cancer, fallopian tube cancer, kidney cancer, melanoma, brain tumor, thyroid cancer, parathyroid cancer, leukemia, lymphoma, myeloma, sarcoma, prostate cancer, bladder cancer, bile duct cancer, and gallbladder cancer.

[0021] In some embodiments of the present invention, the application is application in preparing a ROS promoter for tumors.

[0022] In a fourth aspect of the present invention, a product for preventing or treating tumors is provided, wherein the product comprises the extracellular vesicles of Bifidobacterium longum.

[0023] In some embodiments of the present invention, the product is a medicine or a food.

[0024] In some embodiments of the present invention, the drug further comprises a drug carrier and / or a pharmaceutical excipient.

[0025] In some embodiments of the present invention, the pharmaceutical carrier includes at least one of a diluent, an excipient, a filler, a binder, a disintegrant, an absorption enhancer, a surfactant, an adsorption carrier, a lubricant, a sweetener, and a flavoring agent.

[0026] In some embodiments of the invention, the excipient comprises water.

[0027] In some embodiments of the present invention, the filler comprises at least one of starch and sucrose.

[0028] In some embodiments of the present invention, the binder comprises at least one of a cellulose derivative, alginate, gelatin, and polyvinyl pyrrolidone.

[0029] In some embodiments of the invention, the humectant comprises glycerin.

[0030] In some embodiments of the present invention, the disintegrant comprises at least one of agar, calcium carbonate and sodium bicarbonate.

[0031] In some embodiments of the present invention, the absorption enhancer comprises a quaternary ammonium compound.

[0032] In some embodiments of the invention, the surfactant comprises cetyl alcohol.

[0033] In some embodiments of the present invention, the adsorption carrier includes at least one of kaolin and bentonite.

[0034] In some embodiments of the present invention, the lubricant includes at least one of talc, calcium stearate, magnesium stearate and polyethylene glycol.

[0035] In some embodiments of the present invention, the dosage form of the drug is powder, ointment, drops, gel, lozenge, granules, suspension, syrup, patch, capsule, spray, tablet, pill, injection, gel or oral solution.

[0036] In some embodiments of the present invention, the powder includes a powder and a lyophilized powder.

[0037] In some embodiments of the present invention, the cream comprises an emulsion or a cream.

[0038] According to some embodiments of the present invention, at least the following beneficial effects are achieved: the extracellular vesicles of Bifidobacterium longum prepared by the present invention can effectively inhibit the proliferation of human colorectal cancer cells HT29, SW480 and mouse colon cancer cell line MC38, and can induce the production of reactive oxygen species in tumor cells without affecting normal cells. It is expected that the extracellular vesicles of Bifidobacterium longum according to the present invention can be widely used as drugs with low side effects for alleviating, preventing or treating cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] The present invention will be further described below with reference to the accompanying drawings and embodiments, in which:

[0040] Figure 1 This is a flow chart of the extraction of extracellular vesicles of Bifidobacterium longum in Example 1 of the present invention;

[0041] Figure 2 This is a morphological result of the extracellular vesicles of Bifidobacterium longum XZY01 in the test example of the present invention under a 100,000X electron microscope;

[0042] Figure 3 This is a morphological result of the extracellular vesicles of Bifidobacterium longum XZY01 in the test example of the present invention under a 150,000X electron microscope;

[0043] Figure 4 This is a particle size distribution diagram of the extracellular vesicles of Bifidobacterium longum XZY01 in the test example of the present invention;

[0044] Figure 5Figure 1 is a graph showing the effects of different concentrations of extracellular vesicle solutions of Bifidobacterium longum XZY01 and Lactobacillus plantarum DSM10667 on the proliferation of colorectal cancer cell lines SW480, HT-29, MC-38 and normal intestinal epithelial cells NCM-460 in the test examples of the present invention, wherein A is a graph showing the effects of different concentrations of extracellular vesicle solutions of Bifidobacterium longum XZY01 on the proliferation of colorectal cancer cell lines SW480, HT-29, MC-38 and normal intestinal epithelial cells NCM-460 when co-cultured for 24 hours; B is a graph showing the effects of different concentrations of extracellular vesicle solutions of Bifidobacterium longum XZY01 on the proliferation of colorectal cancer cell lines SW480, HT-29, MC-38 and normal intestinal epithelial cells when co-cultured for 48 hours. A is the result of the effect of different concentrations of extracellular vesicle solution of Lactobacillus plantarum DSM10667 on the proliferation of colorectal cancer cell lines SW480, HT-29, MC-38 and normal intestinal epithelial cells NCM-460 after co-culture for 24 hours; D is the result of the effect of different concentrations of extracellular vesicle solution of Lactobacillus plantarum DSM10667 on the proliferation of colorectal cancer cell lines SW480, HT-29, MC-38 and normal intestinal epithelial cells NCM-460 after co-culture for 48 hours; "*" is p < 0.05; "**" is p < 0.01; "***" is p < 0.005; "****" is p < 0.001;

[0045] Figure 6 This is a graph showing the reactive oxygen species detection results of the colorectal cancer cell line SW480 in the non-drug-added group in the test example of the present invention;

[0046] Figure 7 This is a graph showing the results of reactive oxygen species detection in normal intestinal epithelial cells NCM-460 using a 25 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in a test example of the present invention;

[0047] Figure 8 This is a graph showing the results of reactive oxygen species detection in normal intestinal epithelial cells NCM-460 using a 50 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in a test example of the present invention;

[0048] Figure 9 This is a graph showing the results of reactive oxygen species detection in normal intestinal epithelial cells NCM-460 using a 100 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in a test example of the present invention;

[0049] Figure 10 This is a graph showing the results of reactive oxygen species detection in the colorectal cancer cell line SW480 using a 25 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in a test example of the present invention;

[0050] Figure 11This is a graph showing the results of reactive oxygen species detection in the colorectal cancer cell line SW480 using a 50 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in a test example of the present invention;

[0051] Figure 12 This is a graph showing the results of reactive oxygen species detection in the colorectal cancer cell line SW480 using a 100 μg concentration of extracellular vesicle solution of Bifidobacterium longum XZY01 in the test example of the present invention. DETAILED DESCRIPTION

[0052] The following will clearly and completely describe the concept and technical effects of the present invention in conjunction with the embodiments to fully understand the purpose, features and effects of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are all within the scope of protection of the present invention.

[0053] Experimental Materials:

[0054] MRS culture medium: Qingdao Haibo Biotechnology Co., Ltd. (catalog number: HB0384-5);

[0055] Bacterial genomic DNA extraction kit: Tiangen Biochemical Technology (Beijing) Co., Ltd., (catalog number: DP302-02);

[0056] Taq PCR Mix premix: Sangon Biotech (Shanghai) Co., Ltd. (catalog number: B639295-0001);

[0057] Phosphotungstic acid: China Mirror Instruments (Beijing) Co., Ltd. (catalog number: GA040116);

[0058] 300-mesh circular hole copper mesh: Zhongjing Scientific Instrument (Beijing) Co., Ltd. (Art. No.: AGH300);

[0059] PBS: purchased from Guangzhou Yizibang Biological Instrument Co., Ltd., brand: GIBCO, (catalog number: C10010500BT); BCA quantitative kit: Shanghai Biyuntian Biotechnology Co., Ltd., (catalog number: P0011);

[0060] Reactive oxygen species detection kit: Shanghai Biyuntian Biotechnology Co., Ltd. (Cat. No.: S0033S).

[0061] Example 1

[0062] In this example, extracellular vesicles of Bifidobacterium longum were prepared. The specific process was as follows:

[0063] 1. Recovery culture of Bifidobacterium longum XZY01

[0064] In a clean bench, wipe a cryovial of the XZY01 strain (disclosed in patent CN202110592894.8 and deposited with the Guangdong Provincial Microbial Culture Collection under accession number GDMCC No. 61618) with cotton wool soaked in 75% alcohol. After drying the alcohol, unwrap the sealed film and heat the tube opening with an alcohol lamp. Open the cryovial and add 500 μL of sterile water to the lyophilized powder of the strain. After dissolving and mixing, use a sterile inoculating loop to streak the bacterial solution onto an MRS plate (containing 2% agar) and incubate at 37°C under anaerobism for 48 hours. After 48 hours, pick a single colony from the streaked plate and culture it on a new blood plate and incubate it at 37°C under anaerobism for 48 hours.

[0065] 2. Identification of Bifidobacterium longum XZY01

[0066] Bacterial genomic DNA extraction: Genomic DNA from strain XZ01 was extracted using a bacterial genomic DNA extraction kit (refer to the instructions for specific procedures). After obtaining XZ01 DNA, its concentration and purity were tested. An OD260 / OD280 ratio in the range of 1.7-1.9 was considered acceptable.

[0067] PCR amplification: Thaw upstream and downstream primers, Taq PCR Master Mix, and sterile water on ice. Mix each reagent thoroughly with a pipette before use. The amplification system is shown in Table 1, and the primer sequences are shown in Table 2. After preparation, briefly centrifuge the PCR tube to remove any reaction solution adhering to the wall. Then, perform the PCR amplification procedure. The amplification conditions are shown in Table 3.

[0068] Table 1 PCR amplification system

[0069]

[0070] Table 2 16S rRNA amplification primer sequences

[0071]

[0072] Table 3 PCR amplification conditions

[0073]

[0074] The PCR products were commissioned to Sangon Biotech (Shanghai) Co., Ltd. for 16S rRNA sequencing.

[0075] 3. Obtaining extracellular vesicles of Bifidobacterium longum

[0076] The specific extraction process is as follows Figure 1The procedure is as follows: The correctly sequenced XZY01 strain stored at -80°C was removed from the culture medium and cultured at 37°C for 24 hours. After two activation passages, the strain was inoculated into MRS liquid medium at a 2% (v / v) inoculum for expansion at 37°C for 48 hours. After 48 hours of culture, the fermentation broth was centrifuged (4500 rpm, 45 minutes, 4°C) to remove bacterial cells. The sterile supernatant was collected, filtered through a 0.45 μm filter, transferred to an ultracentrifuge tube, and ultracentrifuged at 120,000 g for 70 minutes. After centrifugation, the pellet was resuspended in pre-chilled PBS. Each batch of pellets was diluted 20-30 times with PBS and ultracentrifuged again to remove residual culture medium. The supernatant was discarded, the pellet was resuspended in PBS, and BCA quantification was performed. The pellet was stored in a -80°C ultra-low temperature freezer.

[0077] Comparative Example 1

[0078] This comparative example provides an extracellular vesicle of Lactobacillus plantarum DSM10667 (purchased from MINZHOUBIO), and the preparation method is the same as the preparation method of the extracellular vesicle of Bifidobacterium longum XZY01 in Example 1.

[0079] Test example

[0080] This test example tested the performance of the extracellular vesicles of Bifidobacterium longum prepared in Example 1.

[0081] 1. Identification of extracellular vesicles of Bifidobacterium longum

[0082] The morphology and particle size distribution of the extracellular vesicles of Bifidobacterium longum XZY01 prepared in Example 1 were detected using a transmission electron microscope and a particle size analyzer.

[0083] Transmission electron microscopy morphology detection: Take 20 μL of extracellular vesicles of Bifidobacterium longum XZY01 and place them on a 300-mesh copper grid. After evaporation, add 20 μL of phosphotungstic acid for negative staining. After evaporation, detect the surface of the grid by electron microscopy.

[0084] Particle size distribution: 700 μL of 20 μg / mL extracellular vesicles of Bifidobacterium longum XZY01 was added to a particle size test dish and diluted gradually until the particle size remained constant. The final result was repeated five times.

[0085] Test results such as Figure 2-4 As shown, from Figure 2-3 As can be seen from the figure, the extracellular vesicles of Bifidobacterium longum XZY01 are spherical, have a clear bilayer structure, and are about 85 nm in diameter. Figure 4 It can be seen that the obtained extracellular vesicles of Bifidobacterium longum XZY01 have a uniform particle size distribution and an average particle size of 87 nm.

[0086] 2. Application of Bifidobacterium longum extracellular vesicles in the preparation of inhibitory agents for colorectal cancer cells

[0087] (1) Preparation of Bifidobacterium longum XZY01 extracellular vesicles: 100 μL of 2 mg / mL extracellular vesicles of Bifidobacterium longum XZY01 prepared in Example 1 was added to 1640 complete medium to prepare a 200 μg / mL stock solution. The solution was filtered through a 0.22 μm microporous filter membrane and then stored in a refrigerator at 4°C until use. The solutions were diluted to 150 μg / mL and 50 μg / mL, respectively, before administration.

[0088] Preparation of extracellular vesicles of Lactobacillus plantarum DSM10667: Dissolve 100 μL of 2 mg / mL extracellular vesicles of Lactobacillus plantarum DSM10667 prepared in the comparative example in 1640 complete medium to create a 200 μg / mL stock solution. Filter through a 0.22 μm microporous filter membrane, aliquot, and store in a refrigerator at 4°C until use. Prior to administration, dilute to 150 μg / mL and 50 μg / mL solutions, respectively.

[0089] (2) Test method: After the colorectal cancer cell lines SW480, HT29, MC38 and the normal intestinal epithelial cell line NCM460 were revived and passaged for more than three generations, they were seeded in 96-well plates, with approximately 5,000 cells per well. After 24 hours of adherent culture, 200 μL of 200 μg / mL, 150 μg / mL, and 50 μg / mL extracellular vesicle solutions of Bifidobacterium longum XZY01 or 200 μL of extracellular vesicle solutions of Lactobacillus plantarum DSM10667 were added to each well, with three replicate wells for each dosing concentration. After culturing at 37°C for 24 hours and 48 hours, 10% CCK-8 reagent was added to the cell system. After incubation at 37°C in the dark for 30 minutes, the OD value at CD450 was measured using a microplate reader. The cell survival rate was calculated using the following formula:

[0090] Cell viability (%) = [A(drug added) - A(blank)] / [A(0 drug added) - A(blank)] × 100

[0091] A (drug addition): OD value of the wells with cells, CCK-8 solution, and drug solution;

[0092] A (0 drug addition): OD value of the well with cells and CCK-8 solution but no drug solution;

[0093] A (blank): OD value of a well without cells.

[0094] The above experiment was repeated three times. The CCK-8 cell proliferation toxicity assay was used to examine the effects of the extracellular vesicle solution of Bifidobacterium longum XZY01 on the proliferation of human colorectal cancer cell lines SW480 and HT-29, mouse colon adenocarcinoma cell line MC-38, and human normal intestinal epithelial cell line NCM-460.

[0095] The results are as follows Figure 5 As shown, from Figure 5 It can be seen that compared with the normal group cells (Control) without drug administration, 200μg / mL Bifidobacterium longum XZY01 extracellular vesicle solution can significantly inhibit the proliferation of SW480, HT-29, and MC-38 cells, and show time and concentration dependence. When incubated with cells for 24 hours, 150μg / ml Bifidobacterium longum XZY01 extracellular vesicle solution can have a significant inhibitory effect on SW480 cells, but has no inhibitory effect on the proliferation of normal human intestinal epithelial cells NCM-460. 200μg / ml Bifidobacterium longum XZY01 extracellular vesicle solution can have a significant inhibitory effect on SW480, HT29 and MC38 cells, but has no inhibitory effect on the proliferation of normal human intestinal epithelial cells NCM-460 ( Figure 5 A). When incubated with cells for 48 hours, 150 μg / ml of extracellular vesicle solution of Bifidobacterium longum XZY01 can significantly inhibit the proliferation of SW480 and MC38 cells, but not the normal human intestinal epithelial cells NCM-460. 200 μg / ml of extracellular vesicle solution of Bifidobacterium longum XZY01 can significantly inhibit the proliferation of SW480, HT29 and MC38 cells, but not the normal human intestinal epithelial cells NCM-460 ( Figure 5 B). After 24 hours of intervention with Lactobacillus plantarum DSM10667 extracellular vesicle solution, we found that compared with the normal control group cells (Control), 200 μg / mL Lactobacillus plantarum DSM10667 extracellular vesicle solution can significantly stimulate the proliferation of human colon cancer cell line HT-29 and normal intestinal epithelial cells NCM460 in a time- and concentration-dependent manner ( Figure 5 C). After 48 hours of intervention with Lactobacillus plantarum DSM10667 extracellular vesicle solution, 200 μg / mL Lactobacillus plantarum DSM10667 extracellular vesicle solution can significantly inhibit the proliferation of human colon cancer cell line SW480, while stimulating the proliferation of HT29, MC38 and NCM460 ( Figure 5 D) These results indicate that the extracellular vesicle solution of Bifidobacterium longum XZY01 can significantly inhibit the proliferation of colorectal cancer cells in a dose- and time-dependent manner, but has no effect on normal NCM-460 cells. In contrast, the extracellular vesicle solution of Lactobacillus plantarum DSM10667 can affect the proliferation of colorectal cancer cells in a dose- and time-dependent manner, while stimulating the proliferation of normal NCM-460 cells.

[0096] 3. Reactive oxygen detection

[0097] Reactive oxygen species (ROS) are the main molecules produced by the body during oxidative stress and have long been considered a key factor in tumor development, progression, and recurrence. Therefore, a reactive oxygen species detection kit was used to detect extracellular vesicles of Bifidobacterium longum XZY01 as follows:

[0098] Colorectal cancer cell line SW480 and normal intestinal epithelial cell line NCM460 were revived and passaged for at least three passages before being seeded in 24-well plates at approximately 10,000 cells per well. After 24 hours of adherence, 1000 μL of a solution of extracellular vesicles of Bifidobacterium longum XZY01 at 25 μg / mL, 50 μg / mL, or 100 μg / mL was added to each well. Three replicate wells were prepared for each dose concentration and incubated at 37°C for 24 hours. Following the protocol for the Reactive Oxygen Species Assay Kit, DCFH-DA was diluted 1:1000 in serum-free culture medium to a final concentration of 10 μmol / L. The cell culture medium was removed, and 1 mL of the diluted DCFH-DA was added. The cells were incubated at 37°C in a cell culture incubator for 20 minutes. The cells were washed three times with serum-free culture medium to fully remove the DCFH-DA that had not entered the cells. The cells were then harvested by trypsinization and analyzed.

[0099] The control group (NC) was the colorectal cancer cell line SW480 without the addition of extracellular vesicles of Bifidobacterium longum XZY01.

[0100] The results of the test on the effect of extracellular vesicles of Bifidobacterium longum XZY01 on the reactive oxygen species of tumor cell line SW480 and normal intestinal epithelial cell NCM460 after 24 hours of intervention are as follows Figure 6-12 As shown, from Figure 6-12 As can be seen, 25μg, 50μg, and 100μg of extracellular vesicles of Bifidobacterium longum XZY01 can significantly increase the level of reactive oxygen species in SW480 cells, but have no effect on NCM460 cells. This suggests that extracellular vesicles of Bifidobacterium longum XZY01 can inhibit the proliferation of tumor cells by stimulating the production of ROS in tumor cells.

[0101] While the embodiments of the present invention have been described in detail above with reference to the accompanying drawings, the present invention is not limited to the embodiments described above. Various modifications may be made within the scope of knowledge possessed by a person skilled in the art without departing from the spirit of the present invention. Furthermore, the embodiments of the present invention and the features thereof may be combined with one another unless there is a conflict.

Claims

1. A use of extracellular vesicles of Bifidobacterium longum in the preparation of a medicine for treating colorectal cancer, characterized in that: The extracellular vesicles of Bifidobacterium longum are extracted from the culture solution of Bifidobacterium longum XZY01; the Bifidobacterium longum XZY01 is deposited in Guangdong Provincial Microbiological Culture Collection Center with a deposit number of GDMCC No: 61618.

2. The use according to claim 1, characterized in that The diameter of the vesicles is 20-500 nm.

3. The use according to claim 2, characterized in that The diameter of the vesicles is 80-90 nm.

4. The use according to claim 1, characterized in that The Bifidobacterium longum extracellular vesicles are used as ROS promoters for tumors and are used to treat colorectal cancer.

5. The use according to claim 1, characterized in that The medicine also includes a drug carrier and / or pharmaceutical excipients.

6. The use according to claim 5, characterized in that The pharmaceutical carrier includes at least one of a diluent, an excipient, a filler, a binder, a disintegrant, an absorption promoter, a surfactant, an adsorption carrier, a lubricant, a sweetener and a flavoring agent.

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