CN112210511A - Bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof - Google Patents

Bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof Download PDF

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CN112210511A
CN112210511A CN202011079328.9A CN202011079328A CN112210511A CN 112210511 A CN112210511 A CN 112210511A CN 202011079328 A CN202011079328 A CN 202011079328A CN 112210511 A CN112210511 A CN 112210511A
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bifidobacterium longum
colon cancer
polysaccharide
protein
inhibiting
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高杰
王潇潇
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Inner Mongolia Puze Biologics Co ltd
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Inner Mongolia Puze Biologics Co ltd
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Abstract

The invention relates to Bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life, wherein the strain number of the Bifidobacterium longum is BB20071001, the Bifidobacterium longum is classified and named as Bifidobacterium longum, and the deposit number is: CGMCC No 4470, preservation date of 2010, 12 months and 14 days, preservation unit: china general microbiological culture Collection center. The bifidobacterium longum BB20071001 is separated from intestinal flora of healthy infants, has better capability of tolerating simulated gastrointestinal fluid, has no toxic or side effect on a human body, has the functions of inhibiting HT-29 cell proliferation and benefiting life, can relieve colitis or colon cancer, and has certain advantages compared with the traditional medicines for treating colitis or colon cancer and wide market prospect.

Description

Bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof
Technical Field
The invention relates to bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof, belonging to the field of application of microbial technology.
Background
The human intestinal flora is one of the most complex micro-ecosystem, and a large number of microorganisms are planted in the human intestinal tract, are influenced and evolved together with a host, and are closely related to the health of a human body. Numerous studies have shown that gut microorganisms affect the proliferation of host gut cells and the development of the immune system, which are associated with many human diseases, such as obesity, Inflammatory Bowel Disease (IBD), infection with pathogenic gut bacteria, cirrhosis of the liver, diabetes, etc. The bifidobacterium longum is used as a dominant bacterium in human intestinal flora, has a symbiotic relationship with a host, and has an important function on maintaining normal intestinal flora and human health. Therefore, some specific bifidobacterium longum strains can be used as probiotics for the intervention and treatment of certain intestinal diseases. The microbial Exopolysaccharide (EPS) is a macromolecular polymer synthesized by some microorganisms and secreted to the outside of cells, and can improve the tolerance of the cells to the gastrointestinal tract environment and protect the cells from the influence of a host immune system.
The extracellular polysaccharide produced by some bifidobacterium longum can regulate the immune response of a host, reduce the inflammation level of the host and play a role in preventing and treating colitis. The drugs currently used for treating inflammatory bowel diseases mainly include anti-inflammatory drugs, immunosuppressants, antibiotics and the like, and these drugs usually only relieve symptoms of diseases and cannot achieve the purpose of treatment, and may cause side effects such as allergy and liver diseases after long-term use. The bifidobacterium longum is used as symbiotic bacteria in intestinal tracts of healthy people, has a certain immunoregulation effect on a host through interaction with an intestinal immune system, and has no toxic or side effect on a human body. The extracellular polysaccharide is an important mechanism for bifidobacterium longum to generate an immunoregulation effect on a host, and the extracellular polysaccharide generated by different microorganisms has different functions due to different monosaccharide compositions and molecular structures. Studies have shown that viscous exopolysaccharides generally have a relatively large molecular weight and have an effect of suppressing an immune response, whereas low molecular weight exopolysaccharides generally promote an immune response. Therefore, the screening of the bifidobacterium longum which can synthesize viscous exopolysaccharides has important significance and broad prospect for developing medicaments for treating colitis or particularly induced colon cancer.
The colon adenocarcinoma is a common digestive tract malignant tumor derived from colon gland epithelium, belongs to one of a plurality of pathological types of colon cancer, the etiology is not clear, but the occurrence of the colon adenocarcinoma is closely related to the occurrence of adenomatous polyp, schistosoma coli, nonspecific ulcerative colitis, colon adenocarcinoma bacillary dysentery, amoebic bowel disease and the like, and the occurrence of the colon adenocarcinoma is related to the diet with high fat and little fiber. About 40% of colon cancer is distributed in the rectum and rectum sigmoid flexure, and the rest is distributed in the sigmoid, cecum, ascending colon, descending colon, transverse colon, and hepatic and splenic flexure. Colon adenocarcinoma is the most common type of colon cancer, and the rest are classified into mucinous adenocarcinoma and undifferentiated carcinoma, and the gross morphology can be polypoid, ulcer type, and the like. The clinical manifestations are that colon cancer is hidden, only fecal occult blood positive is seen at the early stage, bloody stool and dysentery-like purulent bloody stool are gradually generated, tenesmus are generated, intractable constipation sometimes occurs, the stool shape becomes thin, or pasty stool is generated, or diarrhea and constipation alternate, and the changes become prominent manifestations of colon cancer. Patients often have abdominal pain with different degrees, often have erosion, necrosis and secondary infection, and if the abdominal pain occurs on the right side, dull pain of the right abdomen can be generated, and the abdominal pain after meal can be generated sometimes. Left colon cancer often accompanies intestinal obstruction, sometimes abdominal cramps, accompanied by abdominal distension, hyperactive bowel sounds, etc. Abdominal masses are mostly seen in the right abdomen and are one of the manifestations of right colon cancer, suggesting that when the tumor reaches the middle and late stages, the surface of the mass may have a nodular sensation and can be generally pushed, but when the tumor reaches the late stage, the mass is fixed and may have tenderness when the infection is combined. Patients with colon cancer may develop progressive anemia, low grade fever, progressive wasting, cachexia, hepatomegaly, edema, colon adenocarcinoma jaundice, ascites, and the like.
Therefore, the search for new therapeutic drugs that effectively control the development of colitis and prevent the carcinogenesis of colitis has been a focus of attention of researchers.
Disclosure of Invention
The technical problem to be solved by the invention is to provide bifidobacterium longum with the functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof, and the bifidobacterium longum can be used for preparing food or medicine for relieving colitis or colon cancer and solving the technical problem.
The technical scheme for solving the technical problems is as follows: bifidobacterium longum having HT-29 cell proliferation inhibiting and probiotic functions, wherein the strain number of Bifidobacterium longum is BB20071001, which is classified and named as Bifidobacterium longum, and the deposit number is: CGMCC No 4470, preservation date of 2010, 12 months and 14 days, preservation unit: china general microbiological culture Collection center, the preservation unit address: beijing, Chaoyang, Beijing, Xilu No. 1 Yuan No. 3.
The bifidobacterium longum has the following properties:
(1) has acid resistance, good growth under the environment condition of pH3.0-9.0, good survival under the environment of pH2.5;
(2) has adhesion and inhibition, can better adhere to the colon cancer cell HT-29 and inhibit the proliferation of the colon cancer cell HT-29.
The invention has the beneficial effects that: the bifidobacterium longum BB20071001 is separated from intestinal flora of healthy infants, has better capability of tolerating simulated gastrointestinal fluid, has no toxic or side effect on a human body, has the functions of inhibiting HT-29 cell proliferation and benefiting life, can relieve colitis or colon cancer, and has certain advantages compared with the traditional medicines for treating colitis or colon cancer and wide market prospect.
The invention also relates to application of the bifidobacterium longum with the functions of inhibiting HT-29 cell proliferation and benefiting life in preparing food or medicines for relieving colitis or colon cancer.
On the basis of the technical scheme, the invention can be further improved as follows.
Further, the food or the medicine for relieving colitis or colon cancer is fermented milk or microcapsules.
Further, the preparation method of the fermented milk comprises the following steps:
(1) preparation of fermented milk raw materials: mixing milk powder, cane sugar and yeast powder with water according to the proportion of 120-140 g/L, 60-70 g/L and 1-2 g/L respectively, and then sterilizing at 65-75 ℃ for 20-40 min;
(2) adding a leavening agent into the fermented milk raw material by 0.1-1 per mill of mass fraction, and fermenting for 4-8 hours at 37-39 ℃ to obtain a fermented milk product; the leaven consists of the bifidobacterium longum BB20071001, lactobacillus bulgaricus and streptococcus thermophilus.
Further, the bifidobacterium longum BB20071001, the lactobacillus bulgaricus and the streptococcus thermophilus in the leavening agent are in a mass ratio of 1-8:1: 1.
Further, the wall material of the microcapsule comprises denatured protein and polysaccharide, and the core material is bifidobacterium longum BB 20071001; wherein the mass ratio of the denatured protein to the polysaccharide is 1-5:1, and the bacterial content of the microcapsule is 108-1011CFU/g。
Further, the wall material also comprises a freeze-drying protective agent, and the mass ratio of the freeze-drying protective agent to the denatured protein to the polysaccharide is 12.5-25:1-5: 1.
Further, the denatured protein is denatured whey protein or denatured gelatin; the polysaccharide is sodium alginate, chitosan or Arabic gum; the freeze-drying protective agent is glycerol, trehalose or skimmed milk powder.
Further, the preparation method of the microcapsule comprises the following steps:
(1) culturing activated bifidobacterium longum BB20071001, centrifuging the fermentation liquor, and washing the thallus by using normal saline to obtain bifidobacterium longum wet thallus;
(2) adjusting pH of the protein solution to 7.0-8.5, heating to fully denature, mixing with the polysaccharide solution, and sterilizing to obtain protein-polysaccharide mixed solution; adjusting pH of the calcium chloride solution to 6.0-8.5, and sterilizing;
(3) under the aseptic condition, adding the wet thalli obtained in the step (1) into the protein-polysaccharide mixed solution obtained in the step (2), uniformly mixing, then dropwise adding the mixture into a calcium chloride solution for solidification, and carrying out aseptic filtration and washing to obtain wet microcapsules;
(4) and (4) freeze-drying the wet microcapsules obtained in the step (3) to obtain the bifidobacterium longum microcapsules.
Detailed Description
The principles and features of this invention are described below in conjunction with examples which are set forth to illustrate, but are not to be construed to limit the scope of the invention.
Example 1 isolation and screening of Bifidobacterium longum BB20071001
The bifidobacterium longum with the functions of inhibiting HT-29 cell proliferation and benefiting life is the optimal strain screened from a plurality of infant feces through a large number of experiments, and the specific screening method is as follows:
separation and screening of lactic acid bacteria
(1) Collecting a plurality of infant feces of one week of birth, and enriching feces samples in an MRS + 0.5-1 ‰ (mass percentage) cysteine culture medium containing fructo-oligosaccharide for 12 h;
(2) after being diluted in a gradient way, the fecal sample is coated on a solid plate added with 0.02 percent of bromocresol purple by mass percent and MRS + 0.5-1% of cysteine, and cultured for 24-48 h;
(3) selecting single bacterial colony with obvious color changing ring and according with the basic shape of lactic acid bacteria (milky white, semitransparent, round, moist bacterial colony surface, regular edge, etc.), carrying out plate scribing purification, screening and separating out lactic acid bacteria;
(4) and (3) culturing the single colony in liquid MRS + 0.5-1 ‰ (mass percent) cysteine for 24h, performing gram staining, and selecting gram-positive bacteria for subsequent test.
(II) screening strains with HT-29 cell proliferation inhibition and probiotic functions
(1) Respectively inoculating the obtained gram-positive bacteria into 10ml of MRS liquid culture medium in a clean bench, culturing for 12h at 36.5-37.5 ℃ in a constant temperature and humidity chamber by using an anaerobic tube, and repeatedly carrying out passage for 2-3 times to obtain bacterial liquids of different strains.
(2) The human colon cancer cell strain HT-29 is grown in DMEM medium containing 10% fetal calf serum at 37 deg.C and 5% CO2Culturing in a cell culture box with saturated humidity, replacing culture solution every 2-3 days, and taking cells in logarithmic phase for experiment.
(3) Collecting logarithmic phase human colon cancer cells, adjusting cell suspension concentration to 5 × 104Adding 100 mul of the mixture into each hole per ml, and plating;
(4)5％CO2incubating at 37 ℃ until the cell monolayer is paved on the bottom of a well (96-well flat bottom plate), adding bacterial solutions of different strains, and setting a blank control group (without adding the bacterial solution);
(5)5％CO2incubating at 37 ℃, continuously culturing for 3h, 6h, 12h, 24h, 48h and 72h, adding 20ul of MTT solution (5mg/ml, namely 0.5% MTT) into each hole, and continuously culturing for 4 h;
(6) terminating the culture, and carefully sucking out the culture solution in the holes;
(7) adding 150 μ l dimethyl sulfoxide into each well, shaking on a shaker at low speed for 10min to dissolve the crystal completely, and measuring absorbance (A value) of each well at OD490nm of ELISA detector;
(8) the experiment was repeated 3 times;
(9) and calculating the inhibition rate.
Determining one strain with the highest HT-29 cell inhibition rate by an MTT colorimetric method and repeated measurement variance analysis, thereby obtaining the strain with the functions of inhibiting HT-29 cell proliferation and benefiting life, identifying the strain as Bifidobacterium longum by utilizing the microbiological characteristics such as morphological characteristics, culture traits, physiological and biochemical characteristics and the like and molecular biological means such as 16SrDNA and the like, preserving the strain with the strain number of BB20071001 and the classification name of Bifidobacterium longum, wherein the preservation number is as follows: CGMCC No 4470, preservation date: 12/14/2010, depository: china Committee for culture Collection of microorganisms general culture Collection.
The biological properties of bifidobacterium longum BB20071001 are as follows:
colony characteristics: the bacterial colony grows well on a culture medium of MRS +0.5 per mill (mass percent) of cysteine, forms milky, semitransparent and convex round bacterial colony with the diameter of about 3mm, and has smooth and moist surface and regular edges. The cells are long and curved, have rod-shaped, and are partially dumbbell-shaped, without motility and spore production.
Growth characteristics: the lowest growth temperature of the strain is 20 ℃, the highest growth temperature is 45 ℃, the growth temperature of the strain is optimal at 37 ℃, the highest and lowest initial growth pH values are 9.0 and 3.0, and the optimal initial growth pH value is 6.8.
Example 2: fermented milk prepared from Bifidobacterium longum BB20071001
Preparing raw materials, namely, adopting conventional products sold in the market, purchasing a yoghurt starter sold in the market (the mass ratio of lactobacillus bulgaricus to streptococcus thermophilus is 1:1), continuously activating bifidobacterium longum BB20071001 strain for 3 generations, inoculating the strain into an MRS culture medium added with 0.05% cysteine hydrochloride according to the inoculation amount of 2%, anaerobically culturing at 37 ℃ for 24 hours, centrifuging for 30 minutes at 8000g, collecting bacterial sludge, cleaning for 2 times by using physiological saline, adding freeze-drying protective agents (120g/L skim milk powder, 100g/L trehalose, 150g/L sucrose and the balance water) according to 3 times of the weight of the bacterial sludge, uniformly mixing, carrying out vacuum freeze drying, and finally carrying out vacuum packaging on the bacterial powder obtained by freeze drying. The viable bacteria concentration of the obtained bacterial powder can reach 1 × 1010CFU/g, can be used as a starter for preparing fermented milk.
Scheme 1: (1) preparation of fermented milk raw materials: mixing milk powder, sucrose and yeast powder with water at ratio of 120g/L, 70g/L and 1g/L, respectively, and sterilizing at 65 deg.C for 40 min;
(2) adding 0.1 per mill of leaven into the raw material of the fermented milk by mass fraction, fermenting for 8h at 37 ℃ to prepare the fermented milk product; the leaven consists of the bifidobacterium longum BB20071001, the lactobacillus bulgaricus and the streptococcus thermophilus according to the mass ratio of 1:1: 1.
Scheme 2: (1) preparation of fermented milk raw materials: mixing milk powder, sucrose and yeast powder with water at a ratio of 140g/L, 60g/L and 2g/L, respectively, and sterilizing at 75 deg.C for 20 min;
(2) adding a leaven into the raw material of the fermented milk by 1 per mill of mass fraction, and fermenting for 4 hours at 39 ℃ to prepare the fermented milk product; the leaven consists of the bifidobacterium longum BB20071001, the lactobacillus bulgaricus and the streptococcus thermophilus according to the mass ratio of 8:1: 1.
Scheme 3: (1) preparation of fermented milk raw materials: mixing milk powder, sucrose and yeast powder with water at the ratio of 130g/L, 65g/L and 1.2g/L, respectively, and sterilizing at 70 deg.C for 30 min;
(2) adding 0.5 per mill of leaven into the raw material of the fermented milk by mass fraction, fermenting for 6h at 38 ℃ to prepare the fermented milk product; the leaven consists of the bifidobacterium longum BB20071001, the lactobacillus bulgaricus and the streptococcus thermophilus according to the mass ratio of 5:1: 1.
The obtained fermented milk has good organoleptic and physicochemical properties, is fine and smooth in taste, has no bad taste, and has no obvious difference in flavor from the traditional fermented milk, and the effect of relieving colitis or colon cancer can be realized due to the bifidobacterium longum BB20071001 contained in the fermented milk.
Example 3: preparation of microcapsules by using bifidobacterium longum BB20071001
Scheme 1: bifidobacterium longum BB20071001 is used as a core material, and sodium alginate and lactalbumin are used as wall materials.
(1) Preparation of the strain: inoculating 0.1ml of frozen bifidobacterium longum BB20071001 bacterial liquid into 10ml of MRS liquid culture medium in a clean bench, culturing for 12h at 36.5-37.5 ℃ in a constant temperature and humidity chamber by using an anaerobic tube, and repeatedly carrying out passage for 2-3 times to obtain the activated bifidobacterium longum.
Inoculating activated Bifidobacterium longum into 250ml MRS liquid culture medium at volume ratio of 0.1%, placing in constant temperature and humidity box, culturing at 36.5-37.5 deg.C for 15h, centrifuging at 4500rpm for 15min, and collecting thallus. Then, the cells were washed with 0.9% physiological saline, centrifuged at 4500rpm for 15min, and washed repeatedly for 2 times to obtain wet cells of Bifidobacterium longum in the form of a paste.
(2) Adding whey protein into deionized water, stirring for 2h at room temperature (about 25 ℃) by using a magnetic stirrer, standing overnight in an environment at 4 ℃ to ensure complete dissolution and hydration of the whey protein, balancing to room temperature the next day, adjusting the pH to 8.0 by using 1mol/LNaOH to obtain 8 wt% whey protein solution, and heating in a water bath at 80 ℃ for 30min to fully denature the whey protein.
Adding sodium alginate into deionized water, stirring at room temperature until the sodium alginate is uniformly dispersed, standing overnight in an environment at 4 ℃, and balancing to room temperature the next day to obtain a 4 wt% sodium alginate solution.
Mixing the denatured whey protein solution and the polysaccharide solution, wherein the mass ratio of the whey protein to the sodium alginate is 1:1, uniformly mixing the whey protein solution and the sodium alginate by magnetic stirring, and sterilizing the mixture in a boiling water bath at 100 ℃ for 30min to serve as a wall material for later use.
(3) Preparing 3 wt% calcium chloride solution, adjusting pH value to 6.0 with 1mol/LHCl, and sterilizing at 121 deg.C for 20 min.
(4) And (3) adding the thalli obtained by centrifugation into the wall material in a clean bench, and carefully stirring to uniformly mix the thalli and the wall material to avoid bubbles. And (3) under the action of a magnetic stirrer, sucking the mixed solution by using a 2ml sterile syringe, and vertically dropping the mixed solution into a calcium chloride coagulation bath to form microcapsule particles. The wet capsule is solidified in calcium chloride solution for 30min, filtered by sterile sieve, and washed with 0.9% sterile normal saline for 2-3 times to obtain wet microcapsule.
(5) Freeze drying the wet microcapsule in vacuum freeze dryer for 24-48 hr to obtain freeze dried Bifidobacterium longum microcapsule with bacteria content of about 1010CFU/g。
Scheme 2: bifidobacterium longum BBMN68 is used as core material, and sodium alginate, lactalbumin and glycerol are used as wall material.
The polysaccharide solution is prepared by dissolving polysaccharide in 50 wt% glycerol water solution, the mass ratio of glycerol, sodium alginate and whey protein in the final product is 12.5:1:1, and other specific preparation methods are the same as the scheme 1.
Scheme 3: the specific preparation method is the same as scheme 2, but sodium alginate is dissolved in a 50 wt% glycerol aqueous solution, wherein the mass ratio of glycerol to sodium alginate is 25:1, the sodium alginate is mixed with 8 wt% whey protein, and the mass ratio of glycerol, whey protein and chitosan in the final product is 25:5: 1.
Scheme 4: the preparation method is the same as scheme 1, but the pH of 8 wt% whey protein solution is adjusted to 7.0, and the whey protein solution and the chitosan solution are mixed to obtain protein-polysaccharide mixed solution, wherein the mass ratio of whey protein to chitosan is 5:1, and the bacteria content of the microcapsule is about 108cfu/g。
Scheme 5 the specific preparation method is the same as scheme 1, but the pH of a 4 wt% gelatin solution is adjusted to 8.5, and the gelatin and gum arabic solution are mixed to obtain a protein-polysaccharide mixed solution, wherein the mass ratio of the gelatin to the gum arabic is 2:1, and the bacteria content of the microcapsule is about 1011cfu/g。
In addition, in other specific embodiments, the wall material may also be a combination of other polysaccharides (such as chitosan and acacia), other proteins (such as gelatin) and other freeze-drying protective agents (such as trehalose, skim milk powder, etc.), which is not limited to the above-mentioned embodiments.
The microcapsule obtained by the scheme has better gastric acid resistance and enteric solubility, can enter an intestinal tract through a stomach, can be quickly disintegrated in the intestinal tract to release bifidobacterium longum, can be used for preparing a medicament for treating colitis or colon cancer, can also be added into foods such as yoghourt and the like, and plays roles in relieving colitis and preventing canceration of colitis.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.

Claims (10)

1. Bifidobacterium longum having HT-29 cell proliferation inhibiting and probiotic functions, wherein the strain number of Bifidobacterium longum is BB20071001, which is classified and named as Bifidobacterium longum, and the deposit number is: CGMCC No 4470, preservation date of 2010, 12 months and 14 days, preservation unit: china general microbiological culture Collection center.
2. Bifidobacterium longum having HT-29 cell proliferation inhibiting and probiotic properties according to claim 1, characterized in that: the bifidobacterium longum has the following properties:
(1) has acid resistance, good growth under the environment condition of pH3.0-9.0, good survival under the environment of pH2.5;
(2) has adhesion and inhibition, can better adhere to the colon cancer cell HT-29 and inhibit the proliferation of the colon cancer cell HT-29.
3. Use of the bifidobacterium longum according to claim 1 or 2 for inhibiting the proliferation of HT-29 cells and for preparing a food or a medicament for alleviating colitis or colon cancer.
4. The use according to claim 3, wherein the food or medicament for alleviating colitis or colon cancer is fermented milk or microcapsules.
5. Use according to claim 4, characterized in that the method for the preparation of fermented milk comprises the following steps:
(1) preparation of fermented milk raw materials: mixing milk powder, cane sugar and yeast powder with water according to the proportion of 120-140 g/L, 60-70 g/L and 1-2 g/L respectively, and then sterilizing at 65-75 ℃ for 20-40 min;
(2) adding a leavening agent into the fermented milk raw material by 0.1-1 per mill of mass fraction, and fermenting for 4-8 hours at 37-39 ℃ to obtain a fermented milk product; the leaven consists of the bifidobacterium longum BB20071001, lactobacillus bulgaricus and streptococcus thermophilus.
6. The use according to claim 5, wherein Bifidobacterium longum BB20071001, Lactobacillus bulgaricus and Streptococcus thermophilus are present in the starter at a mass ratio of 1-8:1: 1.
7. The use according to claim 4, wherein the wall material of the microcapsules comprises denatured proteins and polysaccharides, and the core material is Bifidobacterium longum BB 20071001; wherein the mass ratio of the denatured protein to the polysaccharide is 1-5:1, and the bacterial content of the microcapsule is 108-1011CFU/g。
8. The use of claim 7, wherein the wall material further comprises a lyoprotectant, and the mass ratio of the lyoprotectant to the denatured protein to the polysaccharide is 12.5-25:1-5: 1.
9. Use according to claim 8, wherein the denatured protein is denatured whey protein or denatured gelatin; the polysaccharide is sodium alginate, chitosan or Arabic gum; the freeze-drying protective agent is glycerol, trehalose or skimmed milk powder.
10. Use according to claim 7, characterized in that the process for the preparation of microcapsules comprises the following steps:
(1) culturing activated bifidobacterium longum BB20071001, centrifuging the fermentation liquor, and washing the thallus by using normal saline to obtain bifidobacterium longum wet thallus;
(2) adjusting pH of the protein solution to 7.0-8.5, heating to fully denature, mixing with the polysaccharide solution, and sterilizing to obtain protein-polysaccharide mixed solution; adjusting pH of the calcium chloride solution to 6.0-8.5, and sterilizing;
(3) under the aseptic condition, adding the wet thalli obtained in the step (1) into the protein-polysaccharide mixed solution obtained in the step (2), uniformly mixing, then dropwise adding the mixture into a calcium chloride solution for solidification, and carrying out aseptic filtration and washing to obtain wet microcapsules;
(4) and (4) freeze-drying the wet microcapsules obtained in the step (3) to obtain the bifidobacterium longum microcapsules.
CN202011079328.9A 2020-10-10 2020-10-10 Bifidobacterium longum with functions of inhibiting HT-29 cell proliferation and benefiting life and application thereof Pending CN112210511A (en)

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CN114574390A (en) * 2022-03-09 2022-06-03 江南大学 Bifidobacterium longum subspecies of infant for relieving colitis and application
CN114847483A (en) * 2022-04-26 2022-08-05 微康益生菌(苏州)股份有限公司 Application of bifidobacterium longum BL21 and microbial inoculum containing same in preparation of product for preventing, relieving or treating colorectal cancer

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CN103932186A (en) * 2014-04-22 2014-07-23 中国农业大学 Bifidobacterium longum microcapsule and preparation method thereof
CN105112333A (en) * 2015-08-31 2015-12-02 江南大学 Bifidobacterium longum with good intestinal tract colonizing ability and screening method and application of bifidobacterium longum
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CN102204679A (en) * 2011-06-16 2011-10-05 高杰 Quadruple bacterial tablets and preparation method thereof
CN103932186A (en) * 2014-04-22 2014-07-23 中国农业大学 Bifidobacterium longum microcapsule and preparation method thereof
CN105112333A (en) * 2015-08-31 2015-12-02 江南大学 Bifidobacterium longum with good intestinal tract colonizing ability and screening method and application of bifidobacterium longum
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Publication number Priority date Publication date Assignee Title
CN114574390A (en) * 2022-03-09 2022-06-03 江南大学 Bifidobacterium longum subspecies of infant for relieving colitis and application
CN114574390B (en) * 2022-03-09 2024-03-01 江南大学 Bifidobacterium longum subspecies infantis for relieving colonitis and application thereof
CN114847483A (en) * 2022-04-26 2022-08-05 微康益生菌(苏州)股份有限公司 Application of bifidobacterium longum BL21 and microbial inoculum containing same in preparation of product for preventing, relieving or treating colorectal cancer
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