Lactobacillus kefiranofaciens subsp. Kefiranofaciens W182 and application thereof

Abstract

The invention relates to the technical field of microorganisms, in particular to a lactobacillus kefir-producing subsp. Kefir W182 and application thereof. The preservation number of the lactobacillus kefir-producing subspecies is CGMCC No.27058, the preservation time is April 11, 2023, and the lactobacillus kefir-producing subspecies can survive for a long time in intestinal tracts, has a strong adhesion characteristic, can produce exopolysaccharides at high yield, can be used for fermenting dairy products, fruit juice, fruit puree, soybean milk and the like, and can also be used for preparing leavening agents and natural thickening agents. Especially when applied to fermented milk products, the fermented milk products can have rich flavor and very good viscosity without adding any existing thickening agent and emulsifying agent, can remarkably reduce separation of whey, can be stored for a long time, and have wide application in the field of food.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, in particular to kfu-producing Lactobacillus kfu-producing subspecies W182 and applications thereof. Background Art

[0002] Lactic acid bacteria (LAB) are Gram-positive bacteria that can produce large amounts of lactic acid by fermenting carbohydrates. They are widely distributed and are usually found in fermented foods such as dairy products and kimchi.

[0003] The application history of lactic acid bacteria is long. In ancient times, the storage and production of a large amount of human food were related to the fermentation of lactic acid bacteria, which not only extended the shelf life of food, but also added special flavors and gave the product probiotic properties. In recent years, more and more lactic acid bacteria fermented products have appeared in people's vision, such as fermented dairy products yogurt, cheese, kefir, kumis, enzymes, and fermented fruit juices, sauerkraut, kimchi and other fermented fruits and vegetables; studies have shown that lactic acid bacteria give fermented foods a good taste, unique flavor and healthy probiotic functions, so lactic acid bacteria and their fermented products have attracted more and more attention from microbiologists, food scientists and nutritionists. Lactic acid bacteria are generally considered safe (GRAS), and in recent years, it has been found that lactic acid bacteria are also widely present in specific environments such as the mouth, intestines, and skin of animals and humans. Lactic acid bacteria are the most common probiotics.

[0004] Lactobacillus has a long history of consumption as a probiotic and is the most widely used microorganism as a starter or auxiliary culture in the fermented food industry. Commonly used species are mainly Lactobacillus plantarum (Lactobacillus casei), Lactobacillus paracasei, Lactobacillus bulgaricus, Streptococcus thermophilus, Lactobacillus acidophilus, Bifidobacterium, etc. In December 2020, Lactobacillus kefiranofaciens (Lactobacillus kefiranofaciens subspecies) belonging to the Lactobacillaceae family and the Lactobacillus genus was added as a new food ingredient. Studies have shown that Lactobacillus kefiranofaciens subspecies ( Lactobacillus kefiranofaciens subsp.kefiranofaciens) does not have any toxicity or pathogenicity and is a safe strain. Wang Yanping and others isolated a strain of Lactobacillus kumiss ZW3 from Tibetan kefir grains. It has good food processing characteristics and probiotic functions, and has the functions of regulating intestinal flora and improving mental health. It has great application potential in the field of green food and functional food. However, there are only a few reports on Lactobacillus kumiss, and its application is even more blank. At present, there are still no products fermented with Lactobacillus kumiss on the market. Given that the relevant research time is short and the results are few, there are many potential strains and related functional characteristics that have not yet been explored, and the market is blank. Therefore, the development, identification and application of Lactobacillus kumiss strain resources with different functions and biological characteristics are of great significance. Summary of the invention

[0005] In view of the above technical status, the present invention provides a strain of Lactobacillus kumiss-producing subspecies kumiss-producing, which has the characteristics of long survival time in the intestine and high production of extracellular polysaccharides.

[0006] The first aspect of the present invention is to provide a strain of Lactobacillus kumiss subspecies, which is Lactobacillus kumiss subspecies W182, classified and named Lactobacillus kefiranofaciens subsp. Kefiranofaciens , was deposited in the General Microbiology Center of China Microbiological Culture Collection Administration on April 11, 2023, with the deposit number CGMCC No. 27058. The discovery of the kefir-producing Lactobacillus W182 fills the relative gap in the existing technology of isolating kefir-producing Lactobacillus from Tibetan kefir grains. The isolated W182 strain has a long survival time in the intestine and has the function of high production of extracellular polysaccharides, and can be used as a potential probiotic in the field of food fermentation.

[0007] The second aspect of the present invention is to provide a fermentation agent, the fermentation agent is prepared from Lactobacillus kumiss subspecies W182, and the number of viable bacteria in the fermentation agent is 10 8 The W182 starter is prepared from kefir-producing Lactobacillus W182, is easy to store and transport, and can be used in the field of food fermentation to reduce the requirements for fermentation conditions, relax the range of fermentation conditions, and improve fermentation efficiency to a certain extent, which helps to save the production cost of industrialized food fermentation products.

[0008] The third aspect of the present invention is to provide an application of a starter, wherein the starter is used in dairy product fermentation, fruit juice fermentation, fruit puree fermentation, and soy milk fermentation.

[0009] Further, the amount of the starter added is such that the final concentration of the kumiss-producing Lactobacillus subspecies W182 in the fermented product reaches 10 6 CFU / g-10 7 CFU / g.

[0010] The fourth aspect of the present invention is to provide a fermented dairy product, including Kfusogenic Lactobacillus Kfusogenic subspecies W182 and / or a starter. Based on the high production of exopolysaccharides by Kfusogenic Lactobacillus W182, and the good emulsifying and thickening properties of the produced exopolysaccharides, the corresponding fermented dairy product produced has good viscosity and taste, and can be stored for a longer time, further filling the gap in the relevant market.

[0011] Further, the dairy product is cow's milk, the fermented dairy product is a fermented cow's milk product, and the final concentration of the kumiss-producing Lactobacillus subspecies W182 in the fermented cow's milk product is 10 6 CFU / mL-10 7 CFU / mL, extracellular polysaccharide content greater than or equal to 1.8g / L, and viscosity greater than or equal to 200mPa·s.

[0012] The fifth aspect of the present invention is to provide a natural thickener, including Kfusogenic Lactobacillus Kfusogenic subspecies W182 and / or dairy fermentation products, which are applied to dairy fermentation products, fruit puree fermentation products, and soy milk fermentation products. Dairy fermentation products can be used alone or in combination with W182 strain and its derivative products, and further applied to other fermentation products as natural thickeners. On the one hand, it can relax the range of fermentation conditions, improve fermentation efficiency, save fermentation production costs, etc. On the other hand, it can reduce the use of chemical additives in other fermentation products, so that other fermentation products can achieve a certain viscosity and taste without adding any existing thickeners and emulsifiers, and can further improve the food safety of other fermentation products.

[0013] A sixth aspect of the present invention is to provide a fermented fruit juice beverage, comprising Lactobacillus kumissogenes subsp. kumissogenes W182 and / or a starter.

[0014] A seventh aspect of the present invention is to provide a fruit puree fermented milk beverage, comprising Kfusogenic Lactobacillus subsp. Kfusogenic W182 and / or a starter.

[0015] An eighth aspect of the present invention is to provide a fermented soymilk product, comprising Lactobacillus kumissogenicus subsp. kumissogenicus W182 and / or a starter.

[0016] Compared with the prior art, the present invention has the following advantages: (1) The present invention provides a kumiss-producing Lactobacillus kumiss subspecies W182, which is isolated and screened from Tibetan kefir grains. The strain is a probiotic that can survive in the intestine for a long time and has strong adhesion properties. Therefore, it has a wide range of applications in the food field and can be used for fermented dairy products, fruit juices, fruit purees, soy milk, etc.

[0017] (2) The kefir-producing Lactobacillus kefir subspecies W182 provided by the present invention has the function of high production of extracellular polysaccharides, and the extracellular polysaccharides have good emulsifying and thickening properties. Based on this function, the kefir-producing Lactobacillus kefir subspecies W182 can be used to ferment dairy products. The obtained fermented dairy products have rich flavor and very good viscosity without adding any existing thickeners and emulsifiers, and can significantly reduce the precipitation of whey. After being stored at 4°C for 30 days, there is still no whey precipitation, and the storage time is relatively long.

[0018] (3) The kumissogenic Lactobacillus kumissogenic subspecies W182 provided by the present invention can ferment fruit juices such as crabapple juice, apple juice, hawthorn juice, pear juice, etc., giving the product a soft sweet and sour taste and a unique flavor.

[0019] (4) The kfusogenic Lactobacillus kufusogenic subspecies W182 provided by the present invention can be used to prepare starter cultures, natural thickeners and other products, and can be further used in food fermentation products. On the one hand, it can save industrial production costs, and on the other hand, it can improve the taste of fermented products and reduce the use of chemical additives. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 This is a colony morphology diagram of the W182 strain in Example 1 of the present application on MRS medium; Figure 2 This is a cell morphology diagram of the W182 strain in Example 1 of the present application; Figure 3 This is a schematic diagram of the comparison of the 16S rDNA sequencing results of the W182 strain in Example 1 of the present application; Figure 4 The survival of the W182 strain in Example 2 of the present application in the mouse intestine; Figure 5 This is the sugar production curve of the extracellular polysaccharide of the W182 strain in Example 2 of the present application; Figure 6 This is a state diagram of fermentation of yogurt by strain W182 of Example 4 of the present application; Figure 7 This is a diagram showing the state of soy milk fermented by the W182 strain in Example 6 of the present application.

[0021] The kefrucigenous Lactobacillus kefrucigenus subspecies of the present invention is: kefrucigenous Lactobacillus kefrucigenus subspecies W182, the preservation date is April 11, 2023, the preservation unit is the General Microbiology Center of the China Microbiological Culture Collection Administration, and the classification name is Lactobacillus kefrucigenus subspecies ( Lactobacillus kefiranofaciens subsp. kefiranofaciens ), the deposit number is CGMCC No.27058. DETAILED DESCRIPTION

[0022] The present invention will be further described below in conjunction with specific embodiments. The following embodiments are only used to more clearly illustrate the technical solution of the present invention, and cannot be used to limit the scope of protection of the present invention. The MRS solid culture medium, MRS liquid culture medium, skim milk culture medium, etc. that are not described in detail in the present patent application can be understood as common knowledge in the art.

[0023] Example 1 Isolation, identification and preservation of W182 strain 1. Isolation of W182 strain A small amount of Tibetan kefir grains was thoroughly ground with 2 mL of saline in a sterile mortar to form a uniform liquid. -3 , 10 -4 , 10 -5 The three gradient homogenates were spread on MRS solid medium and cultured in a 37°C incubator for 1-3 days. Single colonies with obvious characteristics were picked out and purified by streaking until colonies with consistent morphology and size were isolated, numbered, and the colony morphology and cell morphology were observed. They were inoculated into MRS liquid medium with an inoculation loop, cultured at 37°C for 48 hours, and stored in glycerol.

[0024] Colony morphology Figure 1 As shown in Figure 1, W182 colonies on MRS solid medium are white, round or irregular in shape, with a convex center and neat edges, and a diameter of 0.5mm-3mm. Figure 2 As shown, it can be clearly seen that W182 is a bacillus.

[0025] 2. Physiological and biochemical tests of W182 strain The W182 strain is G + Bacillus, a catalase-negative bacteria, was subjected to physiological and biochemical tests using the Merri API 50 CH identification system to study the decomposition of 48 different fermentable carbohydrates. The changes in the organic acids and pH produced caused the indicator to change color, and different fermentation effects were observed. The experimental results are shown in Table 1. The carbon sources that the W182 strain can utilize are D-galactose, D-glucose, D-fructose, D-mannose, esculin, maltose, lactose, and sucrose, and the carbon sources that cannot be utilized are salicin, trehalose, etc.; based on cell morphology and physiological and biochemical characteristics, with reference to the Bergey's Manual of Bacterial Identification and the Classification and Identification of Lactic Acid Bacteria and Experimental Methods, the lactic acid bacteria W182 strain was preliminarily determined to be a kefir-producing Lactobacillus ( Lactobacillus kefiranofaciens ).

[0026] Table 1 Carbohydrate fermentation test results of strain W182

[0027] In Table 1, +: positive; -: negative 3. Molecular Biological Identification of W182 Strain The genomic DNA of strain W182 was extracted, and the genomic extraction kit was purchased from Sangon Biotech (Shanghai) Co., Ltd. The 16SrDNA gene fragment of strain W182 was amplified by PCR using the extracted genomic DNA as a template using universal primers 27F and 1492R (Table 2), and the PCR amplification product was shown in SEQ ID No. 1. The PCR amplification product was sent to Tianjin Jinweizhi Biotechnology Co., Ltd. for sequencing, and the sequencing results were compared with the Genbank database of NCBI. The comparison results are shown in Figure 3 As shown, the strain with the highest homology to strain W182 was found to be Lactobacillus kumissogenes subsp. Lactobacillus kefiranofaciens subsp.kefiranofaciens Therefore, strain W182 was identified as Lactobacillus kumissogenes subsp. Lactobacillus kefiranofaciens subsp.kefiranofaciens .

[0028] Table 2 PCR amplification primers for strain W182

[0029] 4. Deposit of W182 strain The identified W182 strain was preserved and classified as: Lactobacillus kumiss subsp. kumiss, and classified as Lactobacillus kefiranofaciens subsp.kefiranofaciens The strain number is W182. It was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on April 11, 2023. The deposit number is CGMCCNo.27058. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing.

[0030] The subsequent examples are performance studies on Lactobacillus kefir subsp. kefir W182 and application studies in the food field. The W182 strain and Lactobacillus kefir W182 mentioned throughout the article refer to this bacterium, and W182 exopolysaccharide refers to the exopolysaccharide secreted by Lactobacillus kefir subsp. W182.

[0031] Example 2 Performance measurement of lactobacillus W182 1. Survival of Lactobacillus W182 in the intestine Prepare 0.5 mg / mL FITC solution with PBS solution. Take 1 mL and culture until the mid-logarithmic phase (OD 600 = 1.5) of W182 bacterial solution was mixed with 1 mL of FITC solution and incubated at 37°C for 45 min. The cells were collected after centrifugation and washed with PBS solution until the supernatant was colorless. The cells were then resuspended in PBS solution to obtain a concentration of 108 CFU / mL of W182 bacterial suspension. W182 bacterial suspension was used for single or continuous gavage of mice, with a daily gavage dose of 0.3 mL (3.0×10 7 CFU / mouse), and the high-sensitivity multimodal molecular imaging workstation was used to perform fluorescent imaging of the mouse intestine to observe the survival of the W182 strain in the mouse intestine on days 1, 3, 5, 7, 14, and 21.

[0032] The results are as follows Figure 4 As shown, the first group received a single intragastric gavage, and the second group received continuous intragastric gavage. Figure 4 It can be observed that: 1 day after gavage, the W182 strain was clearly present in the mouse intestines in both groups; the fluorescence intensity of the single gavage group was significantly weakened 3 days after gavage, indicating that the number of bacteria was significantly reduced compared with 1 day after gavage; the fluorescence intensity of the continuous gavage group did not have a significant difference within 2 weeks of continuous gavage (1 day-14 days), but the fluorescence intensity was significantly weakened 21 days after gavage (i.e. 7 days after gavage was stopped). In summary, the W182 strain can survive in the body for at least 21 days.

[0033] 2. Exopolysaccharide production ability of Lactobacillus W182 The fully activated W182 strain was inoculated into MRS medium at 4% inoculation rate and cultured at 37°C. The total sugar and reducing sugar content in the fermentation broth were measured every 4 hours. Figure 5 It can be seen that when fermented for 60h, the yield of extracellular polysaccharide EPS can reach 1.954mg / mL. The extracellular polysaccharide has good emulsifying and thickening properties.

[0034] The thermodynamic energy level of exopolysaccharides was measured in the range of 25℃-350℃ by differential scanning calorimeter, and xanthan gum and guar gum were used as standard products for comparison. W182 exopolysaccharides, xanthan gum and guar gum melted at 93.4℃, 153.4℃ and 490.1℃, respectively. The enthalpy change (ΔH) of the endothermic reaction was measured to melt 1g of W182 exopolysaccharides, xanthan gum and guar gum at 249.7℃, 93.2℃ and 192.9℃, respectively. This shows that W182 exopolysaccharides have different thermodynamic properties from xanthan gum and guar gum.

[0035] The emulsifying ability of the extracellular polysaccharide was determined by measuring the ability of EPS to maintain emulsification in water. The experiment compared the emulsifying properties of W182 extracellular polysaccharide with commercial polysaccharides xanthan gum and guar gum. The results showed that the emulsifying activity of W182 extracellular polysaccharide with a purity of 95% was 92.35% at 30 minutes; the emulsifying activity of guar gum was 72.67%, and that of xanthan gum was 92.66%. It can be seen that W182 extracellular polysaccharide shows higher or similar emulsifying activity compared with other common commercial polysaccharides; the emulsifying ability of W182 crude polysaccharide is 89.19%, which has similar emulsifying activity to 95% pure W182 extracellular polysaccharide, so it is more conducive to use as an emulsifier in the future.

[0036] Example 3 W182 starter culture The present embodiment provides an application of kefir-producing Lactobacillus W182, specifically a starter prepared from the W182 strain. A single colony of the purified W182 strain is selected and subcultured in an MRS liquid culture medium, and the bacterial sludge is obtained by centrifugation at 8000rpm. The bacterial sludge is mixed with 12% skim milk at a ratio of 1:3 (M / V), and pre-frozen in a -80°C environment for 4 hours, and vacuum freeze-dried for 18 hours to 24 hours to finally obtain W182 bacterial powder, which is vacuum packaged and refrigerated at low temperature to obtain the W182 starter. The moisture content of the W182 starter is 3%, and the number of viable bacteria in the W182 starter is greater than or equal to 10 8 CFU / g.

[0037] The W182 starter prepared in this embodiment has a wide range of applications and can be used for dairy fermentation, fruit juice fermentation, fruit puree fermentation, soy milk fermentation, etc. In general, the fermentation conditions of the W182 strain are to continue fermenting at 30°C-37°C for 24h-72h, and then post-ripen at 2°C-6°C for 8h-16h to obtain a fermented product. The method of using the W182 starter is to add the W182 starter to the food to be fermented, and the amount of the W182 starter added is such that the final concentration of the kefir-producing Lactobacillus W182 in the fermented product reaches 10 6 CFU / g-10 7 The corresponding fermentation products can be obtained by sealing and fermenting at a temperature range of 20℃-30℃ for 1 day to 60 days. Compared with directly fermenting with W182 strain, using W182 starter can obtain fermentation products at a lower temperature.

[0038] Example 4 Fermented dairy products This embodiment provides an application of kefir-producing Lactobacillus W182 in dairy fermentation, and also provides a fermented dairy product. In other embodiments, dairy products include but are not limited to single-ingredient cow's milk, goat's milk, and mixed dairy products; in other embodiments, the W182 strain can be directly applied to dairy fermentation, or the W182 starter prepared in Example 3 can be used in the fermentation of dairy products. The fermented dairy product obtained by fermentation with the W182 strain or the W182 starter can achieve very good viscosity and taste without adding any existing thickeners and emulsifiers. This embodiment is to directly use the W182 strain to ferment fresh cow's milk.

[0039] The W182 strain was inoculated into a 10% milk culture medium and activated at 37°C. It was inoculated into sterilized milk at an inoculum of 2%, 3%, and 4%, respectively. It was fermented at 37°C for about 48 hours. The fermentation end point was reached when the pH value was measured to be 4.5. It was then refrigerated at 4°C for 12 hours to obtain a milk fermentation product such as Figure 6 shown.

[0040] The final concentration of W182 strain in the fermented milk product was 10 6 CFU / mL-10 7 CFU / mL; the extracellular polysaccharide content of the fermented milk products with different inoculation amounts in the W182 group was measured, and the extracellular polysaccharide content of the W182 strain added at 2% was the highest, reaching 1.95mg / mL; the viscosity of the fermented milk products with different inoculation amounts in the W182 group was measured, and the control group selected yogurt prepared with commercially available yogurt starter, and the results are shown in Table 3-1. As can be seen from the table, the viscosity of the fermented milk products inoculated with the W182 strain is higher than that of the yogurt in the control group, among which the viscosity of the W182 strain added at 2% is the highest, reaching 211.6mPa·s, which may be due to the fact that the W182 strain and its metabolic pathway are different from the existing common fermentation strains.

[0041] Table 3-1 Viscosity test results of yogurt fermented by each group of strains

[0042] Gas chromatography-mass spectrometry (GC-MS) was used to study the flavor substances of fermented milk products. A total of 70 flavor substances were detected, mainly including acids, esters, ketones, alcohols, aldehydes, furan compounds, phenols, hydrocarbons and other substances. The relative contents of acids, esters and ketones were 42.95%, 32.86% and 4.77%, respectively. Among acids, in addition to common organic acids such as lactic acid and acetic acid, caproic acid with sweet cheese aroma and butyric acid with cream flavor were also detected. The relative contents of caproic acid and butyric acid were 9.04% and 0.15%, respectively. Among esters, 3-nonanoic acid methyl ester and propyl caproate with wine aroma and coconut aroma were detected, and the relative contents of the two were 0.29% and 1.54%, respectively. A total of 11 ketones were detected, including acetoin with a relative content of 0.55%. A total of 10 alcohols were detected, including 1-octanol with a lemony smell. A total of 8 aldehydes were detected, accounting for 12.85% in total, mainly including hexanal, nonanal, furfural with a flower aroma, and lauric aldehyde; among them, the relative contents of hexanal, nonanal, and furfural were 13.35%, 0.18%, and 0.2%, respectively. As one of the important flavors of yogurt, three furan compounds were detected, with a relative content of 2.26%. In summary, the high viscosity tissue state and unique flavor of the fermented milk product may be based on the unique metabolic pathway of the W182 strain, which produces a large amount of extracellular polysaccharides and a variety of flavor substances. These extracellular polysaccharides and a variety of flavor substances work together to give the fermented milk product a pleasant taste.

[0043] The sensory evaluation of the fermented milk products was conducted, and the evaluation scores were between 85 and 94 points. The control group still selected yogurt prepared with commercially available yogurt starter. The evaluation results are shown in Table 3-2. As can be seen from the table, the sensory scores of the fermented milk products inoculated with the W182 strain were higher than those of the control group. The fermented milk products containing the W182 strain were whiter in color, had higher tissue morphology and viscosity, were more refreshing, had richer flavor, and delayed the dehydration condensation of whey. After being stored at 4°C for 30 days, there was still no whey precipitation.

[0044] Table 3-2 Sensory evaluation results of fermented milk products

[0045] Example 5 Natural thickener This embodiment provides an application of a fermented dairy product and a natural thickener. The natural thickener includes kefir-producing Lactobacillus W182 and / or a fermented dairy product, and may also include a W182 starter; the natural thickener can be applied to fermented dairy products, fruit puree fermented products, and soy milk fermented products, and the fermented product added with the natural thickener can achieve very good viscosity and taste without adding any existing thickeners and emulsifiers.

[0046] Example 6 Application of strain W182 in fruit juice fermentation and fruit puree fermentation 1. Application of Kweichow Lactobacillus W182 strain in fruit juice fermentation This embodiment provides an application of the Kruskal Lactobacillus W182 strain in fruit juice fermentation, and also provides a fruit juice fermented beverage. In other embodiments, the fruit juice includes but is not limited to apple juice, pear juice, and mixed juice of a single raw material; in other embodiments, the W182 strain can be directly applied to the fruit juice fermentation, or the W182 starter prepared in Example 3 can be used in the fermentation of the fruit juice. In this embodiment, the W182 strain is directly used to ferment crabapple juice.

[0047] Wash and juice the crabapple fruits, filter out the crabapple juice, mix the crabapple juice with water in a ratio of 1:3, add 3% white sugar, sterilize at 75°C for 20 minutes, inoculate with W182 strain at a rate of 2%-4%, ferment at 37°C for 24 hours, sterilize, and fill to prepare a fermented crabapple juice beverage.

[0048] The pH of the fermented crabapple juice beverage was 4.0, the acidity was 9.5 g / KG, and the taste was soft, sweet and sour. The sensory evaluation was conducted and the evaluation score was 95 points. The control group selected a fermented juice beverage prepared with a commercially available starter. The evaluation results are shown in Table 4.

[0049] Table 4 Sensory evaluation results of crabapple juice fermented beverage

[0050] Analysis of organic acids and short-chain fatty acids in crabapple juice fermented beverages revealed that the contents of malic acid, tartaric acid, lactic acid, citric acid, succinic acid and fumaric acid in crabapple juice fermented beverages increased significantly, reaching 0.78ug / mL, 2.65ug / mL, 3.98ug / mL, 0.76ug / mL, 0.02ug / mL and 0.05ug / mL, respectively; short-chain fatty acids included formic acid, acetic acid, propionic acid and butyric acid, with contents reaching 7.01mg / mL, 4.74mg / mL, 4.95mg / mL and 0.48mg / mL, respectively; in addition, there were alcohols and 20 kinds of esters.

[0051] 2. Application of Kweichow Lactobacillus W182 in Fruit Puree Fermentation This embodiment provides an application of the Kruskal Lactobacillus W182 strain in the fermentation of fruit puree, and also provides a fruit puree fermented milk beverage. In other embodiments, the fruit puree includes but is not limited to single-ingredient blueberry puree, raspberry puree, and mixed fresh fruit puree; in other embodiments, the W182 strain, the W182 starter prepared in Example 3, the dairy fermentation product, the cow's milk fermentation product prepared in Example 4, and the natural thickener provided in Example 5 can all be used alone or in combination in the fermentation of fruit puree. In this embodiment, the cow's milk fermentation product prepared in Example 4 is directly used to ferment strawberry puree.

[0052] The fermented milk product prepared in Example 4 was demulsified, mixed with milk and strawberry puree in a ratio of 1:5:0.1, and homogenized, ultra-high temperature sterilized, and filled in sequence to prepare a strawberry puree fermented milk beverage. The beverage retains some organic acids, extracellular polysaccharides and other nutrients of the original fermented milk, giving the fermented milk beverage a soft sweet and sour taste and strawberry flavor. No stratification was observed after storage for a week, and a safe and refreshing all-natural lactic acid bacteria fermented beverage was provided to the market without the presence of any emulsifier or other food additives.

[0053] Example 7 Application of W182 strain in soymilk fermentation This embodiment provides an application of the lactobacillus W182 strain in soy milk fermentation, and also provides a soy milk fermentation product. In other embodiments, soy milk includes but is not limited to soy milk, black bean milk, and mixed soy milk made of a single raw material; in other embodiments, the W182 strain, the W182 starter prepared in Example 3, the dairy fermentation product, the cow's milk fermentation product prepared in Example 4, and the natural thickener provided in Example 5 can all be used alone or in combination in the fermentation of soy milk. This embodiment specifically provides a soybean soy milk fermentation product.

[0054] Soybeans were soaked, ground, filtered and deslagging to obtain soy milk, homogenized at 85°C and 20 MPa for 5 min to obtain a uniform liquid, pasteurized, and then cooled to below 40°C. The W182 strain or the starter prepared in Example 3 was inoculated at an inoculum of 2%-5%, stirred evenly, fermented at 37°C for 36 hours, and after curdling, post-ripened at 4°C for 12 hours to obtain a fermented soy milk product such as Figure 7 As shown. The pH of soy milk fermentation product is 4.52, and the number of viable bacteria is 10 6 cfu / mL or more, and a viscosity of 180mPa·s-200mPa·s. The fermented soymilk product does not require the addition of additional food additives such as milk powder, thickeners, stabilizers and flavors, and can achieve the taste of high-quality fermented sour soymilk without beany smell.

[0055] The above is based on the ideal embodiment of the present invention. Through the above description, relevant personnel can make various changes and modifications without departing from the technical concept of the present invention. The technical scope of the present invention is not limited to the content in the specification, and the technical scope must be determined according to the scope of the claims.

Claims

1. A strain of Lactobacillus kumissogenes subsp. kumissogenes, characterized in that: The kefir-producing Lactobacillus kuefuriensis subspecies is the kefir-producing Lactobacillus kuefuriensis subspecies W182, which is classified and named Lactobacillus kefiranofaciens subsp. kefiranofaciens , and was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on April 11, 2023, with the deposit number CGMCC No.27058.

2. A leavening agent, characterized in that The fermentation agent is prepared from the kefir-producing Lactobacillus subspecies W182 of claim 1, and the number of viable bacteria in the fermentation agent is 10 8 CFU / g and above.

3. An application of a leavening agent, characterized in that: The starter culture according to claim 2 is used in dairy product fermentation, fruit juice fermentation, fruit puree fermentation and soy milk fermentation.

4. The use of the leavening agent according to claim 3, characterized in that: The amount of the fermentation agent added is such that the final concentration of Lactobacillus kumiss-producing subspecies W182 in the fermented product reaches 10 6 CFU / g-10 7 CFU / g.

5. A fermented dairy product, characterized in that: It comprises the kfusogenic Lactobacillus subspecies W182 described in claim 1 and / or the starter described in claim 2.

6. The fermented dairy product according to claim 5, characterized in that: The dairy product is cow's milk, the dairy fermented product is a cow's milk fermented product, and the final concentration of Kfusogenic Lactobacillus subspecies W182 in the cow's milk fermented product is 10 6 CFU / mL-10 7 CFU / mL, extracellular polysaccharide content greater than or equal to 1.8g / L, and viscosity greater than or equal to 200mPa·s.

7. A natural thickener, characterized in that: The invention comprises the kfusogenic Lactobacillus subspecies W182 described in claim 1 and / or the fermented dairy product described in any one of claims 5 or 6, and is applied to fermented dairy products, fruit puree fermented products and soy milk fermented products.

8. A fermented fruit juice beverage, characterized in that: It comprises the kfusogenic Lactobacillus subspecies W182 described in claim 1 and / or the starter described in claim 2.

9. A fruit puree fermented milk beverage, characterized in that: It comprises the kfusogenic Lactobacillus subspecies W182 described in claim 1 and / or the starter described in claim 2.

10. A soymilk fermented product, characterized in that: It comprises the kfusogenic Lactobacillus subspecies W182 described in claim 1 and / or the starter described in claim 2.

Images