WO2020098097A1 - 一株发酵植物乳杆菌及其用途 - Google Patents
一株发酵植物乳杆菌及其用途 Download PDFInfo
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- WO2020098097A1 WO2020098097A1 PCT/CN2018/124327 CN2018124327W WO2020098097A1 WO 2020098097 A1 WO2020098097 A1 WO 2020098097A1 CN 2018124327 W CN2018124327 W CN 2018124327W WO 2020098097 A1 WO2020098097 A1 WO 2020098097A1
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
- C12R2001/25—Lactobacillus plantarum
Definitions
- the invention relates to the technical field of microbial fermentation, in particular discloses a strain of Lactobacillus plantarum, a direct-injection starter containing the strain of Lactobacillus plantarum, and a preparation method thereof, as well as their preparation of fermented dairy products, fermented cereal drinks, and fermented soybean products ,
- the use in the field of fermented fruit and vegetable products, fermented feed and also discloses the use of the Lactobacillus plantarum in the preparation of live bacterial preparations and the prepared live bacterial preparations.
- Lactic acid bacteria (lactic acid bacteria, LAB) is a generic term for a class of bacteria that can use fermentable carbohydrates to produce large amounts of lactic acid. This is a group of quite complex bacteria, currently divided into at least 18 genera, a total of more than 200 species, with rich species diversity.
- Traditional fermented dairy products in China's Qinghai, Cambodia, Inner Mongolia, Xinjiang and other places contain rich resources of lactic acid bacteria, which is a rich treasure trove of probiotic bacteria.
- Lactobacillus plantarum strains in lactic acid bacteria are widely used in food production, especially fermented foods.
- Lactic acid bacteria as potential probiotics need to have the following properties: have outstanding probiotic functions, and need to tolerate the acidic environment of the gastrointestinal tract and bile salt environment, and have good cell adhesion.
- lactic acid bacteria The main role of lactic acid bacteria in fermentation products is to produce acid and produce flavor substances.
- the acid-producing characteristics of lactic acid bacteria are important characteristics for screening starters for fermentation products.
- a lactic acid bacterium with a strong acid-producing ability has a strong tendency to acidify thereafter.
- Lactic acid bacteria with fast acid production and large acid production can bring about lower final product fermentation acidity, which is an effective measure to ensure low post-acidification (such as dairy products).
- gastric juice and intestinal juice constitute the human body's "biological barrier". Lactic acid bacteria must pass through the gastrointestinal tract and be attached to the intestinal epithelial cells to achieve colonization in order to exert beneficial effects on the human body or animal host.
- Exopolysaccharides are produced by lactic acid bacteria during growth and metabolism and secreted to the outside of the cell, released into the growth matrix or capsular polysaccharide tightly bound to the cell surface. Mainly have the following functions: improve product quality, make the product delicate and even, increase the smooth taste; increase water retention, improve product viscosity, help to increase the non-specific adhesion of lactic acid bacteria to the intestinal surface, and increase the colonization of lactic acid bacteria Opportunities, and has a certain role in promoting the regulation of intestinal microflora composition.
- the present invention screened a plant Lactobacillus plantarum with fast acid production, stable acid production and curd genetic properties, and a unique flavor of the fermented product, and used it in fermented dairy products, cereal products, beans In products, fruit and vegetable products and feed products, improve the quality of fermented products.
- the Lactobacillus plantarum can also tolerate the acidic environment of the gastrointestinal tract, can produce high extracellular polysaccharides, improve the colonization of cells, and is used to prepare probiotic live bacterial preparations, which helps to regulate the composition of intestinal microflora.
- the present invention provides a Lactobacillus plantarum strain.
- the classification of the Lactobacillus plantarum strain is named Lactobacillus plantarum. .
- the present invention provides the use of Lactobacillus plantarum as described above in the field of preparing fermented dairy products, fermented cereal drinks, fermented soy products, fermented fruit cooked products, and fermented feed.
- the present invention provides the use of Lactobacillus plantarum as described above in the preparation of a direct-feed starter.
- the present invention provides a direct-throwing starter, which is prepared by fermenting the Lactobacillus plantarum as described above in a fermentation medium.
- the present invention provides a method of direct-injection starter as described above, the method comprising:
- Lactobacillus plantarum is fermented and cultured in a fermentation medium, and the number of viable bacteria is 10 8 cfu / mL or more;
- step (2) Centrifuge the fermentation broth obtained in step (1), then rinse with buffer, add lyoprotectant, and adjust the concentration of viable bacteria to 10 10 cfu / mL or more, mix evenly, and then freeze-dry in vacuum , To get the direct-injection starter.
- the present invention provides the direct-injection starter as described above and / or the direct-injection starter prepared by the method as described above. Uses in the field of fermented feed.
- the present invention provides the use of Lactobacillus plantarum as described above in the preparation of a live bacterial preparation.
- the present invention provides a live bacterial preparation, which is prepared by fermenting Lactobacillus plantarum as described above in a fermentation medium.
- the Lactobacillus plantarum CGMCC No. 16441 of the present invention is a safe strain that can be used for food selected from the milk cake made by the traditional method in the herdsman's house in Inner Mongolia.
- the method of the present invention has no chemical addition and is green Natural, nutritious and healthy;
- the Lactobacillus plantarum CGMCC No. 16441 of the present invention can effectively improve the fermentation quality of fermented dairy products, cereal products, soy products, fruit and vegetable products, feed products, and play the probiotic function of probiotics; the present invention uses Lactobacillus plantarum CGMCC No. As a starter, 16441 can effectively increase the acidity of the product, increase the content of polysaccharides, improve the nutritional value and flavor characteristics of the product, and have a certain improvement on the sensory quality characteristics of the product. Effectively improve the colonization of cells;
- the Lactobacillus plantarum CGMCC No. 16441 of the present invention is easy to culture and prepare as a fermentation agent and a probiotic viable bacteria preparation, and has a high concentration of live bacteria in the fermentation agent of Lactobacillus plantarum.
- the Lactobacillus plantarum provided by the present invention was deposited in the General Microbiology Center of the China Microbial Culture Collection Management Committee on September 7, 2018. Its deposit number is CGMCC No. 16441, and the deposit address is Beichenxi, Chaoyang District, Beijing No. 3, No. 1 Road, Institute of Microbiology, Chinese Academy of Sciences (abbreviated as CGMCC).
- Figure 1 is the colony morphology of Lactobacillus plantarum of the present invention on the culture medium;
- Figure 2 is the micromorphology of Lactobacillus plantarum of the present invention.
- 3 is a graph showing the dynamic change of the pH value of acid production by different strains of the present invention.
- FIG. 4 is the analysis and detection result of the fermentation flavor substance of Lactobacillus plantarum of the present invention.
- the present invention provides a Lactobacillus plantarum strain.
- the classification of the Lactobacillus plantarum strain is named Lactobacillus plantarum, which is deposited in the General Microbiology Center of the China Microbial Culture Collection Management Committee.
- the Lactobacillus plantarum CGMCC No. 16441 has the following properties:
- Morphological characteristics the colonies on MRS medium are milky white, round, smooth, moist, and opaque. The morphology of the cells is rod-shaped under the microscope, and the Gram stain is positive;
- the fermented product also contains ingredients such as 3-hydroxy-2-butanone, 2-heptanone, 2-nonanone, and ethyl caproate to give the product a fresh flavor;
- Adhesion reach 1000 bacteria / 100 cells (HT-29).
- the Lactobacillus plantarum CGMCC No. 16441 described in the present invention is selected from the milk cake made by the traditional method in the herdsmen's home in Inner Mongolia.
- the Lactobacillus plantarum provided by the present invention can produce a large number of viable Lactobacillus plantarum through liquid culture, and the culture method is not particularly required, as long as it can proliferate the Lactobacillus plantarum, for example, according to 10 6- 8 CFU / mL inoculation volume Inoculate the live bacterial cells of Lactobacillus plantarum in the Lactobacillus culture medium, and under anaerobic or aerobic conditions, cultivate at a temperature of 15-40 ° C for 8-72 hours to obtain Culture medium.
- the medium for Lactobacillus can be any medium suitable for the cultivation of Lactobacillus known in the art, for example, it can be milk and / or "lactic acid bacteria-biological basis and application" (Yang Jiebin, Light Industry Press, 1996) ) Described in the lactic acid bacteria (MRS) medium.
- MFS lactic acid bacteria
- the present invention can further isolate the live bacterial cells of Lactobacillus plantarum in the above-mentioned culture liquid.
- the method for the separation is not particularly limited, as long as the cells can be enriched from the culture liquid, for example, by centrifugation and / or filtration Method, the conditions of the centrifugation and the filtration may be well-known conditions, and the present invention will not repeat them here.
- the present invention provides the application of the Lactobacillus plantarum CGMCC No. 16441.
- Lactobacillus plantarum CGMCC No. 16441 in one embodiment of the present invention, refers to the application in fermented food.
- the fermented products include dairy products, cereal products, soy products, fruit and vegetable products, and feed products.
- the dairy product may include yogurt, active lactic acid bacteria beverage, and the like.
- the cereal product may include oat fermented beverage, black rice fermented beverage, buckwheat fermented beverage, and the like.
- the soybean product may include fermented soybean milk and the like.
- the fruit and vegetable product may include fermented fruit and vegetable juice.
- the feed product may include solid anaerobic biological feed.
- Lactobacillus plantarum CGMCC No. 16441 in the present invention is in the conventional production process of producing fermented dairy products, cereal products, soy products, fruit and vegetable products, feed products Inoculate Lactobacillus plantarum CGMCC No. 16441 into the raw material to be treated according to conventional use, and ferment or survive under the temperature and pressure that can propagate the plant Bacillus plantarum CGMCC No. 16441.
- CGMCC No. 16441 By adding CGMCC No. 16441 to the fermentation substrate, its metabolic products make the fermented products have certain advantages such as acidity and aroma, while prolonging the preservation time of the products and improving the nutritional value and flavor characteristics of the products.
- the present invention provides the use of the Lactobacillus plantarum CGMCC No. 16441 in the preparation of a direct-feeding starter.
- the present invention provides a direct-throwing starter, which is prepared by fermenting Lactobacillus plantarum CGMCC No. 16441 in a fermentation medium.
- the fermentation medium may be various conventionally used for fermenting lactic acid bacteria in the art, for example, may be the MRS medium as described above, or may be various suitable for optimization based on the MRS medium.
- Culture medium for fermenting lactic acid bacteria may be various conventionally used for fermenting lactic acid bacteria in the art, for example, may be the MRS medium as described above, or may be various suitable for optimization based on the MRS medium.
- the fermentation conditions may be conventional conditions known in the art for lactic acid bacteria fermentation culture, for example, the fermentation culture temperature may be 30-40 ° C.
- the preparation method of the direct-injection starter can be prepared according to a conventional method in the art.
- it can be prepared according to the following method: Lactobacillus plantarum CGMCC No. 16441 Until the number of viable bacteria reaches more than 10 8 cfu / mL, it is prepared by vacuum freeze drying after mixing evenly.
- the direct-injection starter mentioned above in one embodiment of the present invention, is a bacterial powder prepared by vacuum freeze-drying Lactobacillus plantarum CGMCC No. 16441 fermentation broth, which contains more than 1 billion / g of active Lactobacillus plantarum CGMCC No. 16441.
- the present invention provides a method for preparing the direct-injection starter as described above, the method comprising:
- Lactobacillus plantarum CGMCC No. 16441 is fermented and cultured in a fermentation medium, and the number of viable bacteria reaches 10 8 cfu / mL or more;
- step (2) Centrifuge the fermentation broth obtained in step (1), then rinse with buffer, add lyoprotectant, and adjust the concentration of viable bacteria to 10 10 cfu / mL or more, mix evenly, and then freeze-dry in vacuum , To get the direct-injection starter.
- the method of centrifuging the fermentation bacterial liquid can be performed according to a conventional method in the art.
- a bacterial pellet can be obtained by centrifuging at a speed of 5000-12000 rpm for 5-20 minutes in a refrigerated centrifuge.
- the lyoprotectant may be various cryoprotectants conventional in the art, for example, it may be at least one of skim milk powder, maltodextrin, trehalose, dextran, and glycerin.
- the buffer solution may be a buffer solution conventionally used for washing bacterial cells, for example, it may be physiological saline or PBS buffer solution.
- the present invention provides the direct-injection starter as described above and / or the direct-injection starter prepared by the method described above in the preparation of fermented dairy products, fermented cereal beverages, fermented soy products, fermented fruit cooked products 1. Use in the field of fermented feed.
- the present invention provides the use of Lactobacillus plantarum CGMCC No. 16441 in the preparation of a live bacterial preparation.
- the live bacterial preparation may be in various known forms, such as but not limited to capsules, tablets, oral liquids, and powders.
- the present invention provides a live bacterial preparation, which is prepared by fermenting the Lactobacillus plantarum as described above in a fermentation medium.
- the method for preparing the live bacterial preparation can be prepared by referring to the direct-feeding starter as described above. In order to avoid unnecessary repetition, it will not be repeated here.
- MRS medium Man Rogosa Sharpe Broth: 1 wt% peptone, beef powder 0.5 wt%, 0.4 wt% yeast extract, 2% glucose, 0.1 Tween 80 wt%, K 2 HPO 4 .7H 2 O 0.2 wt% , 0.5% by weight of sodium acetate ⁇ 3H 2 O, 0.2% by weight of triammonium citrate, 0.02% by weight of MgSO 4 ⁇ 7H 2 O, 0.005% by weight of MnSO 4 ⁇ 4H 2 O, 1.5% by weight of agar powder, dissolved by heating, and adjusted to pH 6.2 ⁇ 0.2, autoclaved at 121 °C for 15-20min.
- MRS medium Man Rogosa Sharpe Broth
- MC medium soybean peptone 0.5% by weight, beef powder 0.3% by weight, yeast powder 0.3% by weight, glucose 2% by weight, lactose 2% by weight, calcium carbonate 1% by weight, agar 1.5% by weight, 1% neutral red solution 0.5 weight%.
- Lactobacillus plantarum CGMCC No. 16441 was isolated from Inner Mongolian traditional milk cakes and was preserved by the Nutrition and Healthy Strains Collection Center of COFCO Nutrition and Health Research Institute.
- the inventors collected samples of traditional puff pastry, milk shreds, milk bars, milk tofu and other samples made by traditional methods from herdsmen's homes in Hulunbeier, Xilinguole League, Inner Mongolia, etc., and diluted them with sterile physiological saline gradient to 10 -6 .
- MRS plates and MC plates were incubated at 36 ⁇ 1 °C for 72h. Inoculate needles to select colonies with different colony morphology to the MRS plate and MC plate to streak until single colonies are uniform in size and consistent in shape.
- the strain with Gram staining purple, catalase negative, and no spore formation was selected.
- the above-mentioned strains tentatively identified as lactic acid bacteria were activated in MRS liquid medium for 3 generations for physiological and biochemical identification and molecular biological identification, and the lactic acid bacteria's acid production and curd genetic stability, fermentation performance, sensory evaluation, fermentation Flavor substances, polysaccharide-producing ability, gastrointestinal tract acidic environment and gastrointestinal tract bile salt environment, etc. were studied. After several rounds of research and demonstration, a strain of Lactobacillus plantarum CGMCC was finally screened from many wild lactic acid bacteria .16441.
- the selected Lactobacillus plantarum CGMCC No. 16441 was cultured at 36 ⁇ 1 ° C for 72 hours. As shown in Figure 1, the colonies were milky white, round, smooth, moist and opaque on the MRS medium. As shown in Figure 2, the cell morphology of Lactobacillus plantarum CGMCC No. 16441 was rod-shaped under the microscope, and was Gram-positive.
- Physiological and biochemical identification of the screening strain CGMCC No. 16441 was carried out using the French Merrill API identification system. The identification results are shown in Table 1 below. After physiological and biochemical identification, the isolated strain was Lactobacillus plantarum.
- the 16s rDNA of the isolated strain was cloned and sequenced.
- the nucleotide sequence of the 16s rDNA gene is shown in SEQ ID No. 1.
- the 16s rDNA sequence of the strain of the present invention was compared with the sequence of NCBI Lactobacillus plantarum The 16s rDNA sequence and Lactobacillus plantarum sequence similarity reached 99%.
- this strain was identified as Lactobacillus plantarum.
- the isolated strain was identified as Lactobacillus plantarum, named Lactobacillus plantarum LCZ , and has been deposited in the General Microbiology Center of the China Microbial Culture Collection Management Committee. Deposit address: Chinese Academy of Sciences, No. 3, Beichen West Road, Chaoyang District, Beijing The Institute of Microbiology has the deposit number CGMCC No. 16441 and the deposit date is September 7, 2018.
- test strains were connected to liquid MRS medium and cultured at 37 °C for 24-48h. Inoculate into the subculture medium (whole milk powder is formulated into 12% by weight reconstituted milk solution, add 6% by weight white sugar and mix well, boil for 10 min), and ferment at 37 °C After the milk solidifies, record the fermentation time, and measure the acidity and texture.
- the experimental results are shown in Table 2 and Table 3 after 10 consecutive passages.
- This example is used to illustrate the determination of the acid production capacity of lactic acid bacteria
- the isolated and purified lactic acid bacteria were inoculated into an airtight container filled with MC liquid medium after microscopic examination, cultured at 37 ° C under constant temperature, 15mI of culture liquid was taken out every 2h, and the pH value was measured with a precision acidity meter for a total of 24h. Taking time as the abscissa and the pH value of the fermentation broth as the ordinate, the dynamic change curve of the pH value of acid production of each strain is shown in Figure 3.
- strain CGMCC NO.16441 can reach a pH of 3.76 after 24 hours of fermentation, and its acid production capacity is higher than that of commercial strains (commercial strain No. 1: purchased from Jiangsu Weikang Biotechnology Co., Ltd., lactic acid bacteria strain, strain type: 1807B, Bulgarian milk Bacillus and Streptococcus thermophilus; commercial strain No. 1: purchased from Hebei Yiran Biotechnology Co., Ltd., lactic acid bacteria strain, strain model: Yo-C698-1F, Lactobacillus delbrueckii subsp. Bulgaricus and Streptococcus thermophilus).
- Phenol sulfuric acid method was used for the total sugar: after 2.0mL of the test sample was added to the test tube, 1mL of 6% phenol and 5mL of concentrated sulfuric acid were added in sequence, and then left to stand for 10min, shaken, and the absorbance was detected at a wavelength of 490nm after 20min at room temperature.
- polysaccharide total sugar-reducing sugar
- the screened strain CGMCC NO.16441 has an extracellular polysaccharide yield of up to 803.0 mg / L, which has a high polysaccharide production capacity. It is much higher than 150-600mg / L of similar strains after optimized culture conditions.
- the preparation of simulated intestinal juice 0.1g trypsin, 1.8g bile salt dissolved in a sterile solvent containing 1.1g NaHCO3, 0.2g NaCl and 100mL distilled water, adjust the pH to 8.0 with 0.5M NaOH.
- the solution was sterilized through a 0.45 ⁇ m filter membrane.
- the bacterial liquid that has been activated three times is inoculated into the simulated gastric juice (pH 3.0) according to the inoculation amount of 10% by volume, mixed and incubated at 37 ° C anaerobicly, and the plates are counted at 0 and 3h respectively.
- N 1 represents the number of lactic acid bacteria after simulated gastrointestinal tract culture
- N 0 represents the number of lactic acid bacteria before simulated gastrointestinal tract culture.
- This example is used to illustrate the intestinal adhesion of Lactobacillus plantarum CGMCC No. 16441
- the HT-29 cells cultured good digestion made cell suspension (approximately 10 5 cells / mL), added to 24-well plates of 2mL coverslips have been placed in, 37 °C, 5% CO 2 , 95% Cultivate in the air.
- the culture medium was 1640 culture solution and 10% by weight of inactivated calf serum and L-glutamine. After the cells on the coverslip are basically covered, aspirate the old cell culture solution, rinse with sterile PBS 3 times, and add a mixture of 1 mL bacterial solution (containing 10 8 cfu / mL of bacterial cells) and 1 mL 1640 cell culture solution to each well. Incubate at 37 ° C, 5% CO 2 , 95% air for 2h.
- the adhesion of CGMCC NO.16441 to HT-29 cells is 1000 bacteria / 100 cells, which is higher than 200 bacteria / 100 cells of the same strain with the optimized culture conditions.
- the main role of lactic acid bacteria in dairy fermentation is to produce acid and produce flavor substances.
- the acid-generating properties of lactic acid bacteria are important characteristics for screening starters for fermented dairy products.
- a lactic acid bacterium with a strong acid-producing ability has a strong tendency to acidify thereafter.
- Lactic acid bacteria that produce acid quickly and produce a large amount of acid can bring about a low final product fermentation acidity, which is an effective measure to ensure low post-acidification.
- LLD is a variant of Lactococcus lactis subsp. Butadione; LLL is a subspecies of Lactococcus lactis subsp .; LLC is a subspecies of Lactococcus lactis emulsifiable concentrate;
- Lactobacillus plantarum CGMCC No. 16441 fermented milk flavor substances by taking 20ml coagulated yogurt added 5mL saturated NaCl solution to oscillate and mix, transfer 5mL solution to the headspace bottle to be tested, GC-MS analysis of flavor substances.
- test results of fermented flavor substances are shown in Figure 4.
- a total of 25 main flavor substances were identified in fermented milk, including ketones, alcohols, esters, ethers, etc.
- the ordinary yogurt and CGMCC No. strain fermented yogurt contain the characteristic flavor substances, and also contain 3-hydroxy-2-butanone, 2-heptanone, 2-nonanone and ethyl caproate and other flavor components, which give the yogurt a fragrance Flavor.
- LLD is a variant of Lactococcus lactis subsp. Butadione; LLL is a subspecies of Lactococcus lactis subsp .; LLC is a subspecies of Lactococcus lactis emulsifiable concentrate;
- CGMCC No. 16441 fermented lactic acid bacteria beverage has a fermented lactic acid aroma and a unique taste. The color is pure white and uniform.
- the slurry enters the colloid mill cycle for 2-4 minutes. Add citric acid to adjust the pH of the slurry to 4.0-5.0, add 1-3% saccharification enzyme of the composite slurry, and saccharify at a constant temperature of 40-60 ° C for 4-8h.
- LLD is a variant of Lactococcus lactis subsp. Butadione; LLL is a subspecies of Lactococcus lactis subsp .; LLC is a subspecies of Lactococcus lactis emulsifiable concentrate;
- CGMCC No. 16441 fermented grain beverages have a compound aroma and pleasure
- the fermented flavor has a unique taste, light yellow color and uniformity.
- the fermented soymilk described in this example is prepared by the following method: according to the volume ratio of soft water to soybean 2: 4, soak the soybean in soft water at a temperature of 75-85 ° C for 1-2 hours, remove the soybean hull, remove the soaking water, and then follow the soybean and
- the weight ratio of boiling water is 1: 6-10 plus boiling water grinding slurry, the slurry is kept at a temperature of 80-85 ° C for 10-15min, filtered through a 150 mesh screen to obtain soy milk; add 5-10% by weight of soy milk Sucrose, homogenized again, with a homogenization pressure of 15-25Mpa, followed by a sterilization treatment at a temperature of 95 ° C for 5-10min, after its temperature was reduced to 28-38 ° C, inoculated with 0.04-0.05% (by weight of soybean milk) of commercial Using Lactobacillus bulgaricus, Streptococcus thermophilus and CGMCC NO. 16441 starter, the ratio of bacteria is 1: 1:
- the fermented soybean milk prepared by using only commercial Lactobacillus bulgaricus and Streptococcus thermophilus (the ratio of the number of bacteria is 1: 1) without the addition of Lactobacillus plantarum CGMCC NO.16441 starter was compared.
- the results show that the acidity of fermented soybean milk added with L. plantarum CGMCC NO. 16441 is 2.3 times higher than that without fermented soybean milk. Therefore, the use of L. plantarum CGMCC NO. 16441 in combination with commercial strains can shorten the fermentation time 1-1.5 hours.
- the acid value of the fermented fruit and vegetable juices of Lactobacillus plantarum during refrigeration is smaller, the post-acidification is smaller, and the fermentation stability is good.
- the crop straw is one or more selected from rape stalks, peanut seedlings, corn stalks and the like.
- the herbaceous plant is a mixture of one or more selected from the common herbs of Astragalus.
- the lactic acid content in the feed fermented by Lactobacillus plantarum CGMCC No. 16441 is 21.28g / kg, which is 4.4 times the content of lactic acid in the fermented feed for commercial compound probiotic bacteria.
- Lactobacillus plantarum selected above is used as a live bacterial preparation.
- Lactobacillus plantarum CGMCC NO. 16441 is inoculated into MRS liquid medium according to the inoculation amount of 2-4% (v / v), and cultivated at 37-42 ° C for 20-24h to make the number of viable bacteria of L. plantarum CGMCC NO. Above 10 8 cfu / mL, centrifuge (4000rpm, 10min), rinse the pellet twice with PBS buffer pH6.5-7.5, add skim milk and trehalose as lyoprotectant, adjust the cell concentration to 10 9 cfu / mL, after mixing, vacuum freeze-drying to obtain a live bacterial preparation.
- the number of viable bacteria is 10 8 -10 10 cfu / mL.
- the fungus can be directly added to the fermentation product for fermentation, or it can be added to the food industry or clinically acceptable by conventional methods.
- the dosage form includes various dosage forms such as capsules, microcapsules, tablets and powders.
- the live bacterial preparation made from the above-mentioned screened Lactobacillus plantarum also includes products that maintain the activity of strains through technical means such as domestication.
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Abstract
Description
项目 | 结果 | 项目 | 结果 | 项目 | 结果 |
对照 | - | 肌醇 | - | D-松三糖 | + |
甘露醇 | - | 甘露醇 | + | D-棉子糖 | - |
赤藻糖醇 | - | 山梨醇 | - | 淀粉 | - |
D-***糖 | - | 甲基-αD-吡喃甘露糖苷 | - | 糖原 | - |
L-***糖 | + | 甲基-αD-吡喃葡萄糖苷 | - | 木糖醇 | - |
D-核糖 | + | N-乙酰葡萄糖胺 | + | D-龙胆二糖 | + |
D-木糖 | - | 苦杏仁苷 | + | D-土伦糖 | + |
L-木糖 | - | ARBULIN | + | D-来苏糖 | - |
D-侧金盏花醇1 | - | 七叶灵柠檬酸铁 | + | D-塔格糖 | - |
甲基-βD吡喃木糖苷 | - | 水杨苷 | + | D-岩藻糖 | - |
D-半乳糖 | + | D-纤维二糖 | + | L-岩藻糖 | - |
D-葡萄糖 | + | D-麦芽糖 | + | D-***糖 | - |
D-果糖 | + | D-乳糖 | + | L-***醇 | - |
D-甘露糖 | + | D-蜜二糖 | - | 葡萄糖酸钾 | + |
L-山梨醇 | - | D-蔗糖 | + | 2酮基葡萄糖酸钾 | - |
L-鼠李糖 | - | D-海藻糖 | + | 5酮基葡萄糖酸钾 | - |
卫矛醇 | - | 菊粉 | - |
编号 | 发酵时间/h | 发酵终点pH | 后熟滴定酸度 |
10代 | 15 | 4.53±0.5 | 89.74±0.32 |
1代 | 10 | 4.32±0.67 | 95.93±0.78 |
编号 | 持水力/% | 稠度/g | 粘度 |
10代 | 48.15±0.48 | 4721.86±15.80 | 2388±10.55 |
1代 | 42.90±0.37 | 3886.11±12.47 | 2852±12.36 |
CGMCC NO. | 浓度(mg/L) |
总糖 | 803.0±7.5 |
还原糖 | 0 |
多糖 | 803.0 |
菌株 | LLD | LLL | LLC | CGMCC NO.16441 |
酸度/°T | 72.7±2.4 | 64.4±2.9 | 71.65±3.4 | 95.93±0.78 |
pH值 | 4.39±0.13 | 4.77±0.08 | 4.45±0.14 | 4.32±0.67 |
发酵时间/h | 6 | 6.5 | 6 | 5 |
发酵菌种 | 色泽 | 滋、气味 | 组织状态 | 总分 |
LLD | 10 | 20 | 12 | 42 |
LLL | 15 | 33 | 20 | 68 |
LLC | 14 | 38 | 20 | 72 |
CGMCC No.16441 | 18 | 44 | 22 | 84 |
发酵菌种 | 色泽 | 滋、气味 | 外观 | 总分 |
LLD | 16 | 20 | 11 | 47 |
LLL | 20 | 33 | 13 | 66 |
LLC | 18 | 27 | 12 | 57 |
CGMCC | 26 | 41 | 15 | 82 |
Claims (10)
- 一株植物乳杆菌,其特征在于,该植物乳杆菌的分类命名为Lactobacillus plantarum,保藏于中国微生物菌种保藏管理委员会普通微生物中心,其保藏编号为CGMCC No.16441。
- 权利要求1所述的植物乳杆菌在制备发酵乳制品、发酵谷物饮料、发酵豆制品、发酵果熟制品、发酵饲料领域中的用途。
- 根据权利要求2所述的用途,其中,所述用途包括将权利要求1所述的植物乳杆菌与适配的发酵底物相接触的步骤。
- 权利要求1所述的植物乳杆菌在制备直投式发酵剂中的用途。
- 一种直投式发酵剂,其特征在于,该直投式发酵剂由权利要求1所述的植物乳杆菌在发酵培养基中进行发酵培养后制得。
- 一种制备权利要求5所述的直投式发酵剂的方法,其特征在于,该方法包括:(1)将权利要求1所述的植物乳杆菌在发酵培养基中进行发酵培养,并使活菌数达到10 8cfu/mL以上;(2)将步骤(1)得到的发酵菌液进行离心处理,之后用缓冲液进行冲洗,加入冻干保护剂,并调整活菌浓度至10 10cfu/mL以上,混合均匀后进行真空冷冻干燥,即得所述直投式发酵剂。
- 权利要求5所述的直投式发酵剂和/或权利要求6所述的方法制备的直投式发酵剂在制备发酵乳制品、发酵谷物饮料、发酵豆制品、发酵果熟制品、发酵饲料领域中的用途。
- 权利要求1所述的植物乳杆菌在制备活菌制剂中的用途。
- 根据权利要求8所述的用途,其中,所述活菌制剂包括胶囊、片剂、口服液和粉剂。
- 一种活菌制剂,其特征在于,该活菌制剂由权利要求1所述的植物乳杆菌在发酵培养基中进行发酵培养后制得。
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