Propionibacterium fermentation product, composition and application of propionibacterium fermentation product and composition in preparation of product with anti-enteritis effect

Abstract

The invention relates to the technical field of biological medicines, and particularly discloses a propionibacterium fermentation product, a composition and application of the propionibacterium fermentation product to preparation of a product with an anti-enteritis effect. The preparation method of the propionibacterium fermentation product comprises the following steps: adding propionibacterium into a fermentation culture medium, and fermenting at 30-40 DEG C for 3-5 days; after fermentation is finished, separating fermentation liquor, and concentrating and drying the fermentation liquor to obtain the propionibacterium fermentation product. The fermentation culture medium comprises the following components in parts by weight: 5-15 parts of peptone; 5-15 parts of yeast extract powder; 2-4 parts of glucose; 0.5 to 2 parts of monopotassium phosphate; 0.1-1 part of magnesium sulfate; and 1000 parts of water. Researches show that the propionibacterium fermentation product obtained by adopting the method disclosed by the invention has an anti-colitis effect; therefore, the propionibacterium fermentation product serving as an active ingredient has important application value in preparation of drugs or health care products with the effect of resisting enteritis, especially colitis.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine, and in particular to a propionibacterium fermentation product, a composition and application thereof in preparing a product with anti-enteritis effect. Background Art

[0002] Enteritis is a common digestive tract disease. Ulcerative colitis is a type of enteritis. It is a chronic nonspecific inflammatory intestinal disease. Clinical symptoms include diarrhea, abdominal pain, mucus, pus and blood in the stool, etc. Severe cases may be complicated by intestinal perforation, massive bleeding, cancer, etc. The main treatment for ulcerative colitis is drug therapy. At present, the drugs used to treat ulcerative colitis are mainly Western medicines, such as aminosalicylic acid preparations (such as mesalazine), corticosteroids (such as prednisone), and immunomodulators (such as thiopurine, 6-thiopurine purine nucleotides). However, since ulcerative colitis is a chronic disease, it is difficult to cure completely and often recurs; long-term use of artificially synthesized Western medicines has the problem of large side effects and drug resistance. It is well known that active products of natural origin have the advantage of small side effects; therefore, developing more active products of natural origin for the treatment of ulcerative colitis has important application value for solving the side effects or drug resistance in the treatment of ulcerative colitis. Summary of the invention

[0003] In order to overcome at least one technical problem existing in the prior art, the present invention provides a propionibacterium fermentation product, a composition and an application thereof in preparing a product with anti-enteritis effect.

[0004] The present invention first provides a method for preparing a propionibacterium fermentation product, which comprises the following steps: adding propionibacterium to a fermentation medium, and fermenting at 30-40°C for 3-5 days; separating the fermentation liquid after the fermentation is completed, and concentrating and drying the fermentation liquid to obtain the propionibacterium fermentation product;

[0005] The fermentation medium comprises the following components in parts by weight: 5-15 parts of peptone; 5-15 parts of yeast extract powder; 2-4 parts of glucose; 0.5-2 parts of potassium dihydrogen phosphate; 0.1-1 parts of magnesium sulfate; and 1000 parts of water.

[0006] The inventors found in their research that the Propionibacterium fermentation product obtained by the method of the present invention has an anti-colitis effect.

[0007] Preferably, the weight of the added propionibacterium accounts for 1-5% of the weight of the fermentation medium;

[0008] Most preferably, the added weight of the Propionibacterium accounts for 3% of the weight of the fermentation medium.

[0009] Preferably, the Propionibacterium is Propionibacterium freudenreichii.

[0010] Preferably, the fermentation medium comprises the following components in parts by weight: 10 parts of peptone; 10 parts of yeast extract powder; 3 parts of glucose; 1 part of potassium dihydrogen phosphate; 0.5 parts of magnesium sulfate; and 1000 parts of water.

[0011] Preferably, the fermentation medium further comprises pine pollen.

[0012] Preferably, the fermentation medium comprises the following components in parts by weight: 5-15 parts of peptone; 5-15 parts of yeast extract powder; 2-4 parts of glucose; 3-6 parts of pine pollen; 0.5-2 parts of potassium dihydrogen phosphate; 0.1-1 part of magnesium sulfate; and 1000 parts of water.

[0013] The inventors surprisingly found in further studies that the anti-colitis effect of the Propionibacterium fermentation product obtained by adding pine pollen to the fermentation medium was further improved compared with the Propionibacterium fermentation product obtained by using the fermentation medium without adding pine pollen.

[0014] The present invention also provides a method for preparing a propionibacterium fermentation product.

[0015] The present invention also provides a composition comprising the propionibacterium fermentation product according to claim 6 and a natural plant extract;

[0016] Wherein, the weight ratio of the propionibacterium fermentation product and the natural plant extract described in claim 6 is 3 to 5:1.

[0017] Most preferably, the weight ratio of the propionibacterium fermentation product to the natural plant extract according to claim 6 is 4:1.

[0018] Preferably, the natural plant extract is prepared by the following method:

[0019] Dandelion and purslane are mixed and extracted with a solvent. After the extraction is completed, the extract is separated, and the extract is concentrated and dried to obtain a dandelion-purslane solvent extract. The dandelion-purslane solvent extract is the natural plant extract.

[0020] The inventors found in further studies that the anti-colitis effect of a composition formed by adding a dandelion-purslane solvent extract prepared using dandelion and purslane as raw materials to the Propionibacterium fermentation product was further significantly improved compared to the Propionibacterium fermentation product.

[0021] Further studies have shown that the selection of natural plants in the present invention is very critical. Only when the composition formed by adding the dandelion-purslane solvent extract prepared with both dandelion and purslane as raw materials to the propionibacterium fermentation product, the anti-colitis effect is further significantly improved compared with the propionibacterium fermentation product; however, the anti-colitis effect of the composition formed by only adding the solvent extract prepared with dandelion or purslane as raw materials cannot be further significantly improved.

[0022] Preferably, the solvent is an ethanol aqueous solution with a volume fraction of 50 to 95%.

[0023] Preferably, the ratio of the total weight of dandelion and purslane to the solvent is 1 kg:5-10 L.

[0024] Most preferably, the ratio of the total weight of dandelion and purslane to the solvent is 1 kg:8 L.

[0025] Preferably, the natural plant extract further comprises the following steps: suspending the dandelion-purslane solvent extract in water, and then extracting with an organic solvent; taking the organic solvent extract after extraction, concentrating and drying to obtain an organic solvent extract; taking the organic solvent extract to obtain the natural prepared extract.

[0026] Preferably, the organic solvent consists of n-butanol and cyclopentane.

[0027] More preferably, the volume ratio of n-butanol to cyclopentane is 1 to 3:1.

[0028] Most preferably, the volume ratio of n-butanol to cyclopentane is 2:1.

[0029] The inventors found in further research that the anti-colitis effect of a composition formed by adding an organic solvent extract obtained by further extracting the dandelion-purslane solvent extract with an organic solvent to the Propionibacterium fermentation product was further significantly improved compared to a composition obtained by adding an unextracted dandelion-purslane solvent extract.

[0030] Further studies have shown that the selection of the organic solvent of the present invention is very critical. In the Propionibacterium fermentation product, only by adding a composition formed by an organic solvent extract obtained by further extraction with an organic solvent consisting of n-butanol and cyclopentane, can the anti-colitis effect be further greatly improved; however, only by adding a composition formed by further extraction with n-butanol or cyclopentane, the anti-colitis effect cannot be further greatly improved. It can be seen that in the Propionibacterium fermentation product, the addition of an organic solvent extract obtained by extraction with an organic solvent consisting of n-butanol and cyclopentane can synergistically improve the anti-colitis effect of the composition compared to the addition of an organic solvent extract obtained by extraction with a single n-butanol or a single cyclopentane.

[0031] Preferably, the usage ratio of dandelion-purslane solvent extract to water and mixed organic solvent is 1kg:10-30L:10-30L.

[0032] Most preferably, the usage ratio of dandelion-purslane solvent extract to water and mixed organic solvent is 1kg:20L:20L.

[0033] The present invention also provides an application of the propionibacterium fermentation product and the composition in preparing a product with anti-enteritis effect.

[0034] Preferably, the enteritis is colitis.

[0035] More preferably, the enteritis is ulcerative colitis.

[0036] Preferably, the product is a medicine or a health product.

[0037] Beneficial effects: The present invention provides a propionibacterium fermentation product prepared by a new method; studies have shown that the propionibacterium fermentation product obtained by the method of the present invention has an anti-colitis effect; therefore, using the propionibacterium fermentation product of the present invention as an active ingredient to prepare drugs or health products with anti-enteritis, especially anti-colitis effects has important application value. DETAILED DESCRIPTION

[0038] The present invention is further explained below in conjunction with specific examples, but the examples do not limit the present invention in any form.

[0039] Example 1 Preparation method of propionibacterium fermentation product

[0040] Propionibacterium freudenreichii powder (with a Propionibacterium freudenreichii content of 10 billion cfu / g) is added to a fermentation medium and fermented at 37° C. for 4 days; after the fermentation is completed, the fermentation liquid is filtered and separated, and the fermentation liquid is concentrated and freeze-dried to obtain the propionibacterium fermentation product;

[0041] The weight of the added Propionibacterium freudenreichii powder accounts for 3% of the weight of the fermentation medium;

[0042] The fermentation medium is prepared by mixing the following components in parts by weight: 10 parts of peptone; 10 parts of yeast extract powder; 3 parts of glucose; 1 part of potassium dihydrogen phosphate; 0.5 parts of magnesium sulfate; and 1000 parts of water.

[0043] Example 2 Preparation method of propionibacterium fermentation product

[0044] Propionibacterium freudenreichii powder (with a Propionibacterium freudenreichii content of 10 billion cfu / g) is added to a fermentation medium and fermented at 37° C. for 4 days; after the fermentation is completed, the fermentation liquid is filtered and separated, and the fermentation liquid is concentrated and freeze-dried to obtain the propionibacterium fermentation product;

[0045] The weight of the added Propionibacterium freudenreichii powder accounts for 3% of the weight of the fermentation medium;

[0046] The fermentation medium is prepared by mixing the following components in parts by weight: 10 parts of peptone, 10 parts of yeast extract powder, 3 parts of glucose, 5 parts of pine pollen, 1 part of potassium dihydrogen phosphate, 0.5 parts of magnesium sulfate, and 1000 parts of water.

[0047] Example 3 Preparation of the composition

[0048] The composition is obtained by uniformly mixing the propionibacterium fermentation product and the natural plant extract in a weight ratio of 4:1.

[0049] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0050] The natural plant extract is prepared by the following method: mixing dandelion and purslane, heating and refluxing with a solvent at 75°C for 2h, separating the extract after the extraction, concentrating and drying the extract to obtain a dandelion-purslane solvent extract, and taking the dandelion-purslane solvent extract to obtain the natural plant extract;

[0051] The solvent is an ethanol aqueous solution with a volume fraction of 95%; the total weight of dandelion and purslane to the solvent is 1kg:8L.

[0052] Example 4 Preparation of the composition

[0053] The composition is obtained by uniformly mixing the fermentation product of Propionibacterium and the natural plant extract in a weight ratio of 4:1.

[0054] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0055] The natural plant extract is prepared by the following method:

[0056] (1) Mix dandelion and purslane, and perform extraction by heating and refluxing with a solvent at 75° C. for 2 h. After the extraction, separate the extract, and concentrate and dry the extract to obtain a dandelion-purslane solvent extract; wherein the solvent is an ethanol aqueous solution with a volume fraction of 95%; and the total weight of dandelion and purslane to the solvent is 1 kg:8 L;

[0057] (2) suspending the dandelion-purslane solvent extract in water, and then extracting it with an organic solvent; taking the organic solvent extract after extraction, concentrating and drying to obtain an organic solvent extract; taking the organic solvent extract to obtain the naturally prepared extract; wherein the organic solvent is composed of n-butanol and cyclopentane in a volume ratio of 2:1; the dosage ratio of the dandelion-purslane solvent extract to water and the mixed organic solvent is 1kg:20L:20L.

[0058] Comparative Example 1 Preparation of the composition

[0059] The composition is obtained by uniformly mixing the propionibacterium fermentation product and the natural plant extract in a weight ratio of 4:1.

[0060] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0061] The natural plant extract is prepared by the following method: taking dandelion, heating and refluxing it with a solvent at 75°C for 2h, separating the extract after the extraction, concentrating and drying the extract to obtain a solvent extract, and taking the solvent extract to obtain the natural plant extract;

[0062] The solvent is an ethanol aqueous solution with a volume fraction of 95%; the weight of dandelion and the amount of the solvent are in the ratio of 1kg:8L.

[0063] Comparative Example 2 Preparation of the composition

[0064] The composition is obtained by uniformly mixing the propionibacterium fermentation product and the natural plant extract in a weight ratio of 4:1.

[0065] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0066] The natural plant extract is prepared by the following method: taking purslane, heating and refluxing the extract with a solvent at 75°C for 2h, separating the extract after the extraction, concentrating and drying the extract to obtain a solvent extract, and taking the solvent extract to obtain the natural plant extract;

[0067] The solvent is an ethanol aqueous solution with a volume fraction of 95%; the weight ratio of purslane to the solvent is 1kg:8L.

[0068] Comparative Example 3 Preparation of Composition

[0069] The composition is obtained by uniformly mixing the propionibacterium fermentation product and the natural plant extract in a weight ratio of 4:1.

[0070] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0071] The natural plant extract is prepared by the following method:

[0072] (1) Mix dandelion and purslane, and perform extraction by heating and refluxing with a solvent at 75° C. for 2 h. After the extraction, separate the extract, and concentrate and dry the extract to obtain a dandelion-purslane solvent extract; wherein the solvent is an ethanol aqueous solution with a volume fraction of 95%; and the total weight of dandelion and purslane to the solvent is 1 kg:8 L;

[0073] (2) suspending the dandelion-purslane solvent extract in water, and then extracting it with an organic solvent; taking the organic solvent extract after extraction, concentrating and drying to obtain an organic solvent extract; taking the organic solvent extract to obtain the naturally prepared extract; wherein the organic solvent is n-butanol.

[0074] Comparative Example 4 Preparation of Composition

[0075] The composition is obtained by uniformly mixing the propionibacterium fermentation product and the natural plant extract in a weight ratio of 4:1.

[0076] The Propionibacterium fermentation product was prepared according to the method described in Example 2.

[0077] The natural plant extract is prepared by the following method:

[0078] (1) Mix dandelion and purslane, and perform extraction by heating and refluxing with a solvent at 75° C. for 2 h. After the extraction, separate the extract, and concentrate and dry the extract to obtain a dandelion-purslane solvent extract; wherein the solvent is an ethanol aqueous solution with a volume fraction of 95%; and the total weight of dandelion and purslane to the solvent is 1 kg:8 L;

[0079] (2) suspending the dandelion-purslane solvent extract in water, and then extracting it with an organic solvent; taking the organic solvent extract after extraction, concentrating and drying to obtain an organic solvent extract; taking the organic solvent extract to obtain the naturally prepared extract; wherein the organic solvent is cyclopentane.

[0080] Experimental Example 1

[0081] The colitis model rats were constructed by conventional DSS method, and the specific method was as follows: 108 male SD rats weighing 300±30g were taken, and 50mL of 4% DSS (dextran sulfate sodium salt) aqueous solution was gavaged daily for 7 consecutive days to obtain the colitis model rats.

[0082] The colitis model rats were randomly divided into 9 groups, with 12 rats in each group; they were experimental groups 1 to 8 and a model control group; 12 normal SD rats were selected as the normal control group; the rats in the experimental group were gavaged at a dose of 100 mg / kg per day; the model control group and the normal control group were gavaged with an equal amount of normal saline; the rats in each group drank water and ate normally, and the rats were killed after continuous gavage for 7 days, and blood was collected from the eye sockets; the detection kit was used to detect the content of IL-1β in the serum of the rats in each group, and the test results are shown in Table 1.

[0083] Among them, experimental groups 1 to 7 were gavaged with the Propionibacterium fermentation product prepared in Examples 1 to 2, the composition prepared in Examples 3 to 4, and the composition prepared in Comparative Examples 1 to 2, respectively; the Propionibacterium fermentation product and the composition were suspended in physiological saline before gavage.

[0084] Table 1. Anti-colitis experimental results of the Propionibacterium fermentation products and compositions of the present invention

[0085]

[0086] It can be seen from the experimental results in Table 1 that the content of inflammatory factor IL-1β in the serum of rats in the experimental group 1 is significantly lower than that in the model control group; this indicates that the Propionibacterium fermentation product obtained by the method of the present invention has an anti-colitis effect.

[0087] From the experimental results in Table 1, it can be seen that the content of inflammatory factor IL-1β in the serum of rats in Experimental Group 2 is further lower than that in Experimental Group 1; this indicates that the anti-colitis effect of the Propionibacterium fermentation product obtained by adding pine pollen to the fermentation medium is further improved compared with the Propionibacterium fermentation product obtained by fermentation in the fermentation medium without adding pine pollen.

[0088] It can be seen from the experimental results in Table 1 that the content of inflammatory factor IL-1β in the serum of rats in experimental group 3 is further significantly lower than that in experimental group 2; this indicates that the anti-colitis effect of the composition formed by adding the dandelion-purslane solvent extract prepared with dandelion and purslane as raw materials to the Propionibacterium fermentation product has been further significantly improved compared with the Propionibacterium fermentation product.

[0089] It can be seen from the experimental results in Table 1 that the inflammatory factor IL-1β in the serum of rats in experimental groups 5 and 6 was not significantly reduced compared with that in experimental group 2; this shows that: the selection of natural plants in the present invention is very critical. In the said propionibacterium fermentation product, only the composition formed by adding the dandelion-purslane solvent extract prepared by using dandelion and purslane as raw materials, compared with the propionibacterium fermentation product, its anti-colitis effect is further significantly improved; however, the anti-colitis effect of the composition formed by only adding the solvent extract prepared by using dandelion or purslane as raw materials cannot be further significantly improved.

[0090] It can be seen from the experimental results in Table 1 that the content of the inflammatory factor IL-1β in the serum of rats in the experimental group 4 is further significantly lower than that in the experimental group 3; it is close to that in the normal control group; this shows that: in the said Propionibacterium fermentation product, the composition formed by adding the organic solvent extract obtained by further extracting the dandelion-purslane solvent extract with an organic solvent, compared with the composition obtained by adding the unextracted dandelion-purslane solvent extract, its anti-colitis effect has been further significantly improved; so that the prepared composition has a very excellent anti-colitis effect.

[0091] From the experimental results in Table 1, it can be seen that the content of inflammatory factor IL-1β in the serum of rats in experimental groups 7 and 8 is not much reduced compared with that in experimental group 3, and the reduction is much smaller than that in experimental group 4; this shows that: the selection of the organic solvent of the present invention is very critical. In the fermentation product of Propionibacterium, only by adding a composition formed by further extracting with an organic solvent composed of n-butanol and cyclopentane, its anti-colitis effect can be further greatly improved; however, only by adding a composition formed by further extracting with n-butanol or cyclopentane, its anti-colitis effect cannot be further greatly improved. It can be seen that in the fermentation product of Propionibacterium, adding an organic solvent extract obtained by extracting with an organic solvent composed of n-butanol and cyclopentane at the same time can synergistically improve the anti-colitis effect of the composition compared with adding an organic solvent extract obtained by extracting with a single n-butanol or a single cyclopentane.

Claims

1. A method for preparing a propionibacterium fermentation product, characterized in that: The method comprises the following steps: adding propionibacterium to a fermentation medium, and fermenting at 30-40° C. for 3-5 days; separating the fermentation liquid after the fermentation is completed, and concentrating and drying the fermentation liquid to obtain the propionibacterium fermentation product; The fermentation medium comprises the following components in parts by weight: 5-15 parts of peptone; 5-15 parts of yeast extract powder; 2-4 parts of glucose; 0.5-2 parts of potassium dihydrogen phosphate; 0.1-1 parts of magnesium sulfate; and 1000 parts of water.

2. The method for preparing a propionibacterium fermentation product according to claim 1, characterized in that: The weight of the added propionibacterium accounts for 1 to 5% of the weight of the fermentation medium; Most preferably, the added weight of the Propionibacterium accounts for 3% of the weight of the fermentation medium.

3. The method for preparing a propionibacterium fermentation product according to claim 1, characterized in that: The fermentation medium comprises the following components in parts by weight: 10 parts of peptone; 10 parts of yeast extract powder; 3 parts of glucose; 1 part of potassium dihydrogen phosphate; 0.5 parts of magnesium sulfate; and 1000 parts of water.

4. The method for preparing a propionibacterium fermentation product according to claim 1, characterized in that: The fermentation medium also contains pine pollen.

5. The method for preparing a propionibacterium fermentation product according to claim 4, characterized in that: The fermentation medium comprises the following components in parts by weight: 5-15 parts of peptone; 5-15 parts of yeast extract powder; 2-4 parts of glucose; 3-6 parts of pine pollen; 0.5-2 parts of potassium dihydrogen phosphate; 0.1-1 part of magnesium sulfate; and 1000 parts of water.

6. A Propionibacterium fermentation product prepared by the preparation method according to any one of claims 1 to 5.

7. A composition, characterized in that Comprising the Propionibacterium fermentation product according to claim 6 and a natural plant extract; Wherein, the weight ratio of the propionibacterium fermentation product and the natural plant extract described in claim 6 is 3 to 5:

1.

8. The composition according to claim 7, characterized in that The natural plant extract is prepared by the following method: Dandelion and purslane are mixed and extracted with a solvent. After the extraction is completed, the extract is separated, and the extract is concentrated and dried to obtain a dandelion-purslane solvent extract. The dandelion-purslane solvent extract and the natural prepared extract are obtained.

9. The composition according to claim 8, characterized in that The solvent is an ethanol aqueous solution with a volume fraction of 50-95%.

10. Use of the propionibacterium fermentation product of claim 6 and the composition of any one of claims 7 to 9 in the preparation of a product having an anti-enteritis effect.

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