Bifidobacterium symbiotic system as well as culture method and application thereof

Abstract

The invention provides a bifidobacterium symbiotic system and a culture method and application thereof, and belongs to the technical field of microorganisms. The preparation method comprises the following steps: firstly, culturing streptococcus thermophilus, then inoculating bifidobacterium for culturing to obtain a mixed culture solution of the two bacteria, and then co-culturing the mixed culture solution and a chlorella culture solution to obtain the bifidobacterium symbiotic system. The bifidobacterium symbiotic system obtained by the method maintains the activity and performance of bifidobacteria, also improves the immunity of the organism and promotes the growth of the organism.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and in particular relates to a bifidobacterium symbiotic system and a culture method and application thereof. Background Art

[0002] Bifidobacterium is a type of Gram-positive, non-motile, non-spore-forming, obligately anaerobic bacteria. It is an important component of the intestinal microbial community of humans and animals. Bifidobacterium has a variety of physiological functions that are beneficial to the host. It has excellent performance in maintaining the balance of intestinal microecology, enhancing intestinal barrier function, participating in nutrient metabolism, and regulating the immune system. However, Bifidobacterium is an anaerobic bacterium that is extremely sensitive to environmental conditions, and its activity is easily lost under the influence of external environmental factors. In an aerobic environment, Bifidobacterium lacks an effective antioxidant defense mechanism. Oxygen causes a large amount of reactive oxygen species (ROS) to be produced in its cells. These ROS can damage cell membranes, proteins, and DNA, thereby affecting the survival and normal function of Bifidobacterium. In addition, changes in environmental conditions such as temperature, humidity, and pH value can also have a significant effect on the activity of Bifidobacterium. For example, high temperature may cause protein denaturation and damage to cell membrane structure, and an overly acidic or alkaline environment may affect the activity of its enzymes and the balance of biochemical reactions in cells.

[0003] At present, the common preservation methods of bifidobacteria are freeze-drying and low-temperature preservation. Freeze-drying can maintain the activity of bifidobacteria to a certain extent, but during the freezing and drying process, the formation of ice crystals and dehydration stress may damage the cell structure, resulting in the death of some bacteria or reduced activity; and during the storage process after freeze-drying, if the seal is not appropriate, bifidobacteria are prone to moisture absorption, and re-absorption of water may further damage its cell structure. Storing bifidobacteria in a low-temperature environment (such as liquid nitrogen or a low-temperature refrigerator) can reduce its metabolic rate, thereby extending its active preservation time; but this method requires continuous low-temperature conditions and corresponding equipment support, which is costly, and temperature fluctuations during storage and use may still affect the activity of bifidobacteria; in addition, long-term low-temperature storage may cause some adaptive changes, so that bifidobacteria cannot quickly restore their optimal functional state when they return to normal physiological temperature. In addition, bifidobacteria are used to maintain their activity by symbiosis with other microorganisms, but different microorganisms have different absorption and utilization efficiencies of nutrients, which may cause some bifidobacteria to be unable to obtain enough nutrients to maintain their activity, and the growth and metabolism of bifidobacteria are inhibited. It is necessary to select appropriate microbial species for symbiosis with different types of bifidobacteria in order to effectively maintain their activity. Summary of the invention

[0004] In view of this, the object of the present invention is to provide a bifidobacterium symbiotic system and a culture method and application thereof, wherein the bifidobacterium symbiotic system maintains the activity and performance of bifidobacteria, and effectively improves the body's immunity and promotes the body's growth.

[0005] In order to solve the above technical problems, the present invention provides the following technical solutions:

[0006] The present invention provides a bifidobacterium symbiotic system, which comprises bifidobacterium, thermophilic streptococcus and chlorella.

[0007] Preferably, the bifidobacterium is Bifidobacterium animalis, strain number CICC 6165; the thermophilic Streptococcus is strain number CICC 22563.

[0008] The present invention also provides a method for culturing a bifidobacterium symbiotic system, comprising the following steps: inoculating thermophilic streptococci onto an MRS culture medium for culturing, adding supplementary nutrients and inoculating bifidobacteria, and anaerobic culturing to obtain a mixed culture solution of thermophilic streptococci and bifidobacteria; inoculating Chlorella species onto a photoautotrophic culture medium for culturing to obtain a Chlorella culture solution; and co-culturing the mixed culture solution of thermophilic streptococci and bifidobacteria with the Chlorella culture solution to obtain a bifidobacterium symbiotic system.

[0009] Preferably, the thermophilic streptococcus is cultured on the MRS medium at a temperature of 36-38° C., a time of 10-14 h, and a rotation speed of 100-150 r / min.

[0010] Preferably, the temperature of the anaerobic culture of the bifidobacteria is 36-38°C, the time is 46-50h, and the rotation speed is 100-150r / min.

[0011] Preferably, the formula of the MRS medium includes: 8-12g of peptone, 3-7g of beef extract powder, 18-22g of glucose, 1.5-2.5g of dipotassium hydrogen phosphate heptahydrate, 1.5-2.5g of triammonium citrate, 3-7g of ammonium acetate trihydrate, 0.1-0.3g of magnesium sulfate heptahydrate, and 0.02-0.04g of manganese sulfate tetrahydrate. Add water to 1L and adjust the pH to 6.8-7.0.

[0012] Preferably, the nutritional supplement formula comprises: 0.5-2.5 g of yeast extract and 0.05-1 g of bile acid, 0.5 L of water is added, and the pH is adjusted to 6.8-7.0.

[0013] Preferably, the photoautotrophic culture medium formula includes: 2-4g glucose, 1-3g sodium chloride, 0.120-0.130g magnesium sulfate, 1.0-1.5g potassium dihydrogen phosphate, 1.0-1.5g citric acid, 1.2-1.6g ammonium ferric citrate, add water to 1L, and adjust the pH to 7.0-7.2.

[0014] Preferably, the temperature for culturing the Chlorella species inoculated into the photoautotrophic culture medium is 27-32° C., the time is 93-100 h, and the rotation speed is 50-60 r / min.

[0015] Preferably, the co-culture temperature is 32-35°C, the time is 22-26 hours, and the rotation speed is 70-90 r / min.

[0016] The present invention provides the use of the bifidobacterium symbiotic system or the bifidobacterium symbiotic system obtained by the culturing method in preparing an immunity enhancing product.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] The present invention prepares a symbiotic system for the first time by combining Bifidobacterium animalis (strain number CICC 6165), Streptococcus thermophilus (strain number CICC 22563), and Chlorella to maintain the activity and performance of Bifidobacterium. The bacterial activity in the liquid Bifidobacterium symbiotic system can be maintained for more than two weeks at room temperature. At the same time, after consuming the Bifidobacterium symbiotic system, the immunity of the body can be improved and the growth of the body can be promoted. DETAILED DESCRIPTION

[0019] The present invention provides a bifidobacterium symbiotic system, the bifidobacterium symbiotic system comprises bifidobacterium, thermophilic streptococcus and chlorella. The bifidobacterium described in the present invention is Bifidobacterium animalis, strain number CICC 6165, purchased from China Industrial Microorganism Culture Collection Administration Center; the thermophilic streptococcus is Streptococcus thermophilus, strain number CICC 22563, purchased from China Industrial Microorganism Culture Collection Administration Center; the chlorella is food grade, there is no special limitation on its source, and it can be purchased through commercial channels.

[0020] The present invention also provides a method for cultivating the bifidobacterium symbiotic system, comprising the following steps: after thermophilic streptococcus is inoculated on MRS culture medium and cultured, supplementary nutrients are added and bifidobacterium is inoculated, anaerobic culture is performed, and a mixed culture of thermophilic streptococcus and bifidobacterium is obtained; Chlorella species are inoculated on a photoautotrophic culture medium to obtain a chlorella culture solution; thermophilic streptococcus and bifidobacterium mixed culture solution are co-cultured with chlorella culture solution to obtain a bifidobacterium symbiotic system. The present invention first cultures thermophilic streptococcus, consumes oxygen in the culture medium, and simultaneously produces numerous primary metabolites, providing an anaerobic environment and sufficient effective active ingredients for bifidobacterium; the supplementary nutrients added before inoculating bifidobacterium provide nutrition for thermophilic streptococcus and bifidobacterium again, maintaining strain activity; Chlorella is rich in nutrients such as protein, chlorophyll, carotenoids, polysaccharides, and in the co-culture process, when the chlorella cells rupture or die, these nutrients are released into the culture medium, and thermophilic streptococcus and bifidobacterium can use the components therein as a nutrient source.

[0021] In the present invention, the thermophilic streptococcus is cultured on the MRS medium at a temperature of 36-38°C, a time of 10-14h, and a rotation speed of 100-150r / min; preferably, the temperature is 37°C, the time is 12h, and the rotation speed is 120r / min. The inoculation amount of the thermophilic streptococcus in the present invention is 1-5% (V / V), preferably 3%. The thermophilic streptococcus purchased by the present invention is activated by conventional methods to obtain OD 600 = 0.2-0.4 of thermophilic Streptococcus liquid for inoculation and culture.

[0022] In the present invention, the temperature of the anaerobic culture of the bifidobacteria is 36-38°C, the time is 46-50h, and the speed is 100-150r / min; preferably, the temperature is 37°C, the time is 48h, and the speed is 120r / min. The inoculation amount of the bifidobacteria of the present invention is 1-5% (V / V), preferably 3%. The bifidobacteria purchased by the present invention are activated by conventional methods to obtain OD 600 = 0.15-0.35 of Bifidobacterium liquid for inoculation and culture.

[0023] In the present invention, the formula of the MRS culture medium includes: 8-12g of peptone, 3-7g of beef extract powder, 18-22g of glucose, 1.5-2.5g of dipotassium hydrogen phosphate heptahydrate, 1.5-2.5g of ammonium citrate trihydrate, 3-7g of ammonium acetate trihydrate, 0.1-0.3g of magnesium sulfate heptahydrate, 0.02-0.04g of manganese sulfate tetrahydrate, add water to 1L, and adjust the pH to 6.8-7.0; preferably, 10g of peptone, 5g of beef extract powder, 20g of glucose, 2g of dipotassium hydrogen phosphate heptahydrate, 1.5g of triammonium citrate, 4g of ammonium acetate trihydrate, 0.15g of magnesium sulfate heptahydrate, 0.025g of manganese sulfate tetrahydrate, add water to 1L, and adjust the pH to 6.9. The raw materials used in the MRS culture medium of the present invention can be purchased through commercial channels.

[0024] In the present invention, the supplementary nutrient formula includes: 0.5-2.5g yeast extract and 0.05-1g bile acid, add 0.5L water, and adjust the pH to 6.8-7.0; preferably 1.57g yeast extract and 0.089g bile acid, add 0.5L water, and adjust the pH to 6.8. The raw materials used for the supplementary nutrient of the present invention can be purchased through commercial channels. The supplementary nutrient of the present invention provides nutrients for bifidobacteria, promotes their growth and reproduction, and creates a living environment for bifidobacteria during dormancy.

[0025] In the present invention, the photoautotrophic culture medium formula includes: 2-4g glucose, 1-3g sodium chloride, 0.120-0.130g magnesium sulfate, 1.0-1.5g potassium dihydrogen phosphate, 1.0-1.5g citric acid, 1.2-1.6g ammonium ferric citrate, add water to 1L, adjust the pH to 7.0-7.2; preferably 2.5g glucose, 1.8g sodium chloride, 0.125g magnesium sulfate, 1.3g potassium dihydrogen phosphate, 1.25g citric acid, 1.45g ammonium ferric citrate, add water to 1L, adjust the pH to 7.1. The raw materials used for the nutritional supplements of the present invention can be purchased through commercial channels.

[0026] In the present invention, the temperature of the chlorella species inoculated into the photoautotrophic medium is 27-32°C, the time is 93-100h, and the rotation speed is 50-60r / min; preferably, the temperature is 28°C, the time is 95h, and the rotation speed is 55r / min. In the present invention, 2g of the purchased chlorella species is inoculated into 100mL of the photoautotrophic medium.

[0027] In the present invention, the mixed culture solution of thermophilic streptococcus and bifidobacterium and the culture solution of chlorella are mixed in a ratio of 2-8:1, preferably 4:1. The temperature of the mixed culture solution of thermophilic streptococcus and bifidobacterium and the culture solution of chlorella is 34°C, the time is 24h, and the speed is 80r / min. The bifidobacterium symbiotic system after co-cultivation of the present invention can be directly bottled and vacuumed, or the bifidobacterium symbiotic system culture solution can be prepared into freeze-dried powder by conventional freeze-drying method.

[0028] The present invention also provides the use of the bifidobacterium symbiotic system or the bifidobacterium symbiotic system obtained by the culturing method in the preparation of an immunity enhancing product. The bifidobacterium symbiotic system of the present invention is suitable for the general public, especially children or elderly people with low immunity.

[0029] In the present invention, unless otherwise specified, all components, reagents or culture media are commercially available products well known to those skilled in the art.

[0030] The technical solutions in the present invention will be described clearly and completely below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0031] Example 1

[0032] 1. Culture medium preparation

[0033] The formula of MRS medium is: 10 g peptone, 5 g beef extract powder, 20 g glucose, 2 g dipotassium hydrogen phosphate heptahydrate, 1.5 g triammonium citrate, 4 g ammonium acetate trihydrate, 0.15 g magnesium sulfate heptahydrate, 0.025 g manganese sulfate tetrahydrate, add water to 1 L and adjust the pH to 6.9.

[0034] The nutritional supplement formula is: 1.57 g yeast extract and 0.089 g bile acid, add 0.5 L water, and adjust the pH to 6.8.

[0035] The formula of the photoautotrophic culture medium is: 2.5 g glucose, 1.8 g sodium chloride, 0.125 g magnesium sulfate, 1.3 g potassium dihydrogen phosphate, 1.25 g citric acid, 1.45 g ammonium ferric citrate, add water to 1 L, and adjust the pH to 7.1.

[0036] The preparation method of the above culture medium is unified as follows: weigh each culture medium component, add it into the weighed distilled water, heat and boil it to make it completely dissolved, calibrate the pH value (25°C), and after packaging, sterilize it at 121°C for 15 minutes and set aside.

[0037] 2. Preparation of Bifidobacterium Symbiotic System

[0038] (1) Preparation of bacterial solution: Inoculate the purchased Streptococcus thermophilus (strain number CICC 22563) onto MRS medium and culture until OD 600 =0.3, and obtain thermophilic Streptococcus bacterial solution; inoculate the purchased animal Bifidobacterium (strain number CICC 6165) into MRS medium and culture until OD 600 =0.25, and a bifidobacterium culture solution was obtained. (2) The thermophilic Streptococcus culture solution was inoculated into 250 mL of MRS culture medium at an inoculum volume of 3% (V / V), and cultured at a rotation speed of 120 r / min and 37° C. for 12 h.

[0039] (3) Then, 50 mL of supplementary nutrients were added, and a 3% (V / V) inoculated bifidobacterium culture solution was inoculated. The mixture was cultured anaerobically at a speed of 120 r / min and 37° C. for 48 h to obtain a mixed culture solution of thermophilic streptococci and bifidobacteria.

[0040] (4) 2 g of Chlorella was inoculated into 100 mL of photoautotrophic culture medium, and the culture was carried out at a rotation speed of 55 r / min and 28° C. for 96 h to obtain Chlorella culture solution.

[0041] (5) The mixed culture solution of thermophilic Streptococcus and Bifidobacterium was mixed with the culture solution of Chlorella at a mixing ratio of 4:1, and co-cultured at a speed of 80 r / min and 34° C. for 24 h to obtain a bifidobacterium symbiotic system.

[0042] Example 2

[0043] The culture medium preparation was the same as that in Example 1.

[0044] Preparation of Bifidobacterium Symbiotic System

[0045] (1) Preparation of bacterial solution: Inoculate the purchased Streptococcus thermophilus (strain number CICC 22563) onto MRS medium and culture until OD 600 =0.5, and obtain thermophilic Streptococcus bacterial liquid; the purchased animal Bifidobacterium (strain number CICC 6165) was inoculated into MRS medium and cultured until OD 600 =0.4, and obtain bifidobacterium liquid.

[0046] (2) The thermophilic Streptococcus liquid was inoculated into 250 mL of MRS culture medium at an inoculum size of 5% (V / V), and cultured at a rotation speed of 150 r / min and 32° C. for 18 h.

[0047] (3) Then, 40 mL of supplementary nutrients were added, and a 5% (V / V) inoculation amount of bifidobacterium culture solution was inoculated. The mixture was anaerobically cultured at a speed of 150 r / min and 32° C. for 50 h to obtain a mixed culture solution of thermophilic streptococci and bifidobacteria.

[0048] (4) 4 g of the purchased Chlorella species was inoculated into 100 mL of photoautotrophic culture medium, and cultured at a rotation speed of 50 r / min and 30° C. for 98 h to obtain a Chlorella culture solution.

[0049] (5) The mixed culture solution of thermophilic Streptococcus and Bifidobacterium was mixed with the culture solution of Chlorella at a mixing ratio of 6:1, and co-cultured at a speed of 75 r / min and 35° C. for 28 h to obtain a bifidobacterium symbiotic system.

[0050] Comparative Example 1

[0051] The thermophilic Streptococcus (strain number CICC 22563) was replaced by thermophilic Streptococcus (CICC 22565, purchased from China Industrial Microbiological Culture Collection Center), and the remaining steps were the same as in Example 1.

[0052] Comparative Example 2

[0053] The difference from Example 1 is that the thermophilic Streptococcus and related steps are discarded, and the remaining steps are the same as Example 1.

[0054] Comparative Example 3

[0055] The animal Bifidobacterium (strain number CICC 6165) was replaced with Bifidobacterium longum (Bifidobacterium longum, CICC 6207, purchased from China Industrial Microbiological Culture Collection Center), and the remaining steps were the same as in Example 1.

[0056] Comparative Example 4

[0057] The beef extract in the MRS medium was replaced with yeast extract, and the remaining steps were the same as in Example 1.

[0058] Comparative Example 5

[0059] The yeast extract in the supplementary nutrient was replaced with corn syrup, and the remaining steps were the same as in Example 1.

[0060] Comparative Example 6

[0061] The difference from Example 1 is that the steps of step (2) and step (3) in Example 1 are changed to: the thermophilic streptococcus liquid and the bifidobacterium liquid are respectively inoculated into 250mL MRS medium according to the inoculation amount of 3% (V / V), and the thermophilic streptococcus and bifidobacterium mixed culture liquid are cultured at a speed of 120r / min and 37°C for 12h to obtain a thermophilic streptococcus and bifidobacterium mixed culture liquid; step (5) is changed to add 50mL of supplementary nutrients, and then the thermophilic streptococcus and bifidobacterium mixed culture liquid is mixed with the chlorella culture liquid at a mixing ratio of 4:1, and the speed is 80r / min and co-cultured at 34°C for 24h to obtain a bifidobacterium symbiotic system.

[0062] Test Example 1

[0063] The bifidobacterium symbiotic system obtained from Example 1-2 and Comparative Example 1-6 was placed at room temperature for 4 weeks, the existence status of the symbiotic system was observed, and samples were taken at 1 week, 2 weeks, 3 weeks, and 4 weeks to detect the number of live bifidobacteria in the symbiotic system.

[0064] The detection method is: add 1 mL of sample to 9 mL of sterile diluent (normal saline) and dilute it in the same way. -5 The dilution was spread on MRS medium plates (prepared by adding 2 g agar to the above MRS), and three plates were spread for each sample. The plates were then placed in an anaerobic incubator and cultured at 37°C for 48 hours. The typical bifidobacterium colonies (round, neatly edged, milky white, etc.) on the plates were counted.

[0065] Table 1 Results of stability study on the bifidobacterium symbiotic system in each group

[0066]

[0067] As shown in Table 1, the bifidobacterium symbiotic system prepared by combining thermophilic Streptococcus (strain number CICC 22563) and animal Bifidobacterium (strain number CICC6165) and adopting specific nutrients and culture methods has high stability, and can still maintain high bifidobacterium activity after being placed at room temperature for four weeks; while abandoning thermophilic Streptococcus and replacing other bifidobacteria cannot achieve a complete symbiotic effect because the required nutrients and culture conditions are different.

[0068] Test Example 2

[0069] Healthy young SD mice, weighing 10-20 g, male, were selected from Jiangsu Huachuang Xinnuo Pharmaceutical Technology Co., Ltd. Rats were raised in an animal room at a temperature of (22±2)°C and a humidity of (50±10)%, with free access to water and food, and the experiment began after one week of adaptive feeding.

[0070] Enzyme-linked immunosorbent assay (ELISA) kits were used to determine serum immunoglobulin IgM, IgG, IgA, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). The activities of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase were determined on an Rx Daytona automatic biochemical analyzer; serum growth hormone (GH) was determined using a mouse growth hormone (GH) ELISA kit (Wuhan Elaruite Biotechnology Co., Ltd.) The results are shown in Table 2.

[0071] The bifidobacterium symbiotic system obtained in Examples 1-2 and Comparative Examples 1-6, as well as the thermophilic streptococcus liquid and bifidobacterium liquid obtained in step (1) of Example 1 were fed to rats, and the control group was fed with the same amount of normal saline. The feeding was continued for 25 days. After the feeding, blood was collected from the jugular vein of all rats, serum was separated, and serum biochemical indices, hormone levels, and serum immune factors were measured. The results are shown in Table 3.

[0072] Table 2 Effects of Bifidobacterium symbiotic system on blood biochemical indexes and hormone levels in rats

[0073]

[0074] Table 3 Effects of Bifidobacterium symbiotic system on immune indexes of rats

[0075]

[0076]

[0077] The results of Table 2-3 show that the IgM, IgG, IgA, IL-6, and TNF-α contents in the rat serum of the bifidobacterium symbiotic system prepared by feeding Example 1-2 are significantly increased relative to the comparative example group, the control group, the bifidobacterium liquid group, and the thermophilic streptococcus liquid group, indicating that the bifidobacterium symbiotic system prepared by the present invention has the effect of significantly enhancing the immunity of rats; the aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and serum growth hormone contents in the rat serum of the bifidobacterium symbiotic system prepared by feeding Example 1-2 are significantly increased relative to the comparative example group, the control group, the bifidobacterium liquid group, and the thermophilic streptococcus liquid group, indicating that the bifidobacterium symbiotic system prepared by the present invention has the effect of significantly improving the growth performance of rats. At the same time, the replacement of each component or the change of the culture part in Comparative Example 1-6 has an effect on the above-mentioned indicators, and compared with the results of Example 1-2, each indicator is significantly reduced. This shows that the present invention scientifically combines thermophilic Streptococcus (strain number CICC 22563), animal Bifidobacterium (strain number CICC 6165) and Chlorella, and adopts a specific culture medium formula and culture method to obtain a bifidobacterium symbiont with high stability, and can enhance the immunity of the organism and promote the growth of the organism.

[0078] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A bifidobacterium symbiotic system, characterized in that: The bifidobacterium symbiotic system comprises bifidobacterium, thermophilic streptococcus and chlorella; the bifidobacterium is animal bifidobacterium, strain number CICC 6165; the thermophilic streptococcus strain number CICC 22563.

2. A method for cultivating a bifidobacterium symbiotic system, characterized in that: The steps include: After thermophilic streptococci are inoculated on MRS medium for cultivation, supplementary nutrients are added and bifidobacteria are inoculated, and anaerobically cultured to obtain a mixed culture solution of thermophilic streptococci and bifidobacteria; Inoculating the Chlorella species into a photoautotrophic culture medium for cultivation to obtain a Chlorella culture solution; The mixed culture solution of thermophilic streptococcus and bifidobacterium is co-cultured with the culture solution of chlorella to obtain a bifidobacterium symbiotic system.

3. The culture method according to claim 2, characterized in that The thermophilic streptococcus is cultured on the MRS culture medium at a temperature of 36-38° C., a time of 10-14 hours, and a rotation speed of 100-150 r / min.

4. The culture method according to claim 2, characterized in that The temperature of the bifidobacterium anaerobic culture is 36-38°C, the time is 46-50h, and the rotation speed is 100-150r / min.

5. The culture method according to claim 2, characterized in that The formula of the MRS culture medium includes: 8-12g of peptone, 3-7g of beef extract powder, 18-22g of glucose, 1.5-2.5g of dipotassium hydrogen phosphate heptahydrate, 1.5-2.5g of triammonium citrate, 3-7g of ammonium acetate trihydrate, 0.1-0.3g of magnesium sulfate heptahydrate, and 0.02-0.04g of manganese sulfate tetrahydrate. Water is added to 1L, and the pH is adjusted to 6.8-7.

0.

6. The culture method according to claim 2, characterized in that: The formula of the supplementary nutrient comprises: 0.5-2.5 g of yeast extract and 0.05-1 g of bile acid, 0.5 L of water is added, and the pH is adjusted to 6.8-7.

0.

7. The culture method according to claim 2, characterized in that The photoautotrophic culture medium formula includes: 2-4g of glucose, 1-3g of sodium chloride, 0.120-0.130g of magnesium sulfate, 1.0-1.5g of potassium dihydrogen phosphate, 1.0-1.5g of citric acid, 1.2-1.6g of ammonium ferric citrate, add water to 1L, and adjust the pH to 7.0-7.

2.

8. The culture method according to claim 2, characterized in that The temperature of the chlorella inoculated into the photoautotrophic culture medium is 27-32° C., the time is 93-100 hours, and the rotation speed is 50-60 r / min.

9. The culture method according to claim 2, characterized in that: The co-culture temperature is 32-35° C., the time is 22-26 hours, and the rotation speed is 70-90 r / min.

10. Use of the bifidobacterium symbiotic system according to claim 1 or 2 or the bifidobacterium symbiotic system obtained by the culturing method according to claims 3 to 9 in the preparation of an immunity enhancing product.

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