Preparation and quality evaluation of fermented milk with long bifidobacterium microcapsule and black fungus

Abstract

The application relates to a preparation method of black fungus fermented milk, and relates to the food field, and aims to solve the problems of intolerance to gastric acid and bile salts after long bifidobacterium is made into fermented milk, poor taste and improvement of the added value of black fungus. The application mixes bacteria mud obtained by centrifugation of long bifidobacterium culture solution with pectin, and cross-linking is carried out to obtain long bifidobacterium microcapsules. Long bifidobacterium microcapsules, black fungus liquid and whole milk are used as main raw materials, skimmed milk powder, sodium carboxymethyl cellulose and white granulated sugar are used as auxiliary materials, the raw materials are uniformly mixed, and then black fungus fermented milk is obtained through fermentation. The black fungus fermented milk prepared by the application has a fine organization state and does not have the phenomenon of whey separation. The application is applied to the field of fermented milk.

Description

Technical Field

[0001] The invention relates to the field of food, and in particular to a method for preparing black fungus fermented milk. Background Art

[0002] Black fungus (Auricularia auricula), a precious colloid fungus used in both medicine and food in my country, is also recognized worldwide as a health food. Its delicious taste and rich nutrition have earned it the reputation of "the best of the vegetarians," "the king of the vegetarians," and "the treasure of the fungi." Both medicinal and edible, black fungus boasts a rich nutritional profile, containing not only polysaccharides, melanin, collagen, polyphenols, and flavonoids, but also trace elements such as iron, zinc, calcium, and manganese. Its high iron content makes it an excellent tonic for those with anemia. The calcium content in black fungus is dozens of times higher than in meat and four to ten times higher than in vegetables. The high amount of dietary fiber in black fungus promotes gastrointestinal motility. The phospholipids in black fungus nourish brain cells and nerve cells, making it a practical and affordable brain tonic for adolescents and those engaging in mental work. Therefore, black fungus has multiple biological activities such as antioxidant, anti-tumor, immune regulation, hyperlipidemia, anti-diabetes, anti-coagulation, and liver protection. In-depth exploration of the functions of black fungus and product development have important market value.

[0003] Currently, my country boasts advantages such as a large-scale industry, mature cultivation techniques, a relatively stable market, low investment, and significant returns. my country's black fungus production accounts for over 90% of the world's total. However, a weakness in deep processing has become a bottleneck restricting the industry's development. This is primarily manifested in the following aspects: First, the processed products are mostly crude, resulting in low added value, weak market competitiveness, and a relatively limited consumption pattern; second, innovation is insufficient, and resource utilization needs to be optimized; and finally, production technology is relatively low-level, energy-intensive, and environmentally unfriendly.

[0004] Currently, processed black fungus products are primarily used in food, health supplements, pharmaceuticals, and cosmetics. In the food sector, black fungus products primarily include black fungus polysaccharide steamed buns, black fungus cakes, black fungus biscuits, and ultrafine black fungus powder. In the health supplement sector, they are primarily used in oral liquids, chewable tablets, and enzymes. Miaoji Pills, a traditional Chinese medicine with black fungus as its primary ingredient, is used to treat liver and kidney deficiency, nourishing the liver and kidneys and removing dampness and unblocking meridians. Furthermore, black fungus plays a crucial role as a prescription ingredient in medications such as Strong Bone and Blood-Growing Oral Liquid. In the cosmetics sector, the focus is on the development of facial masks. As a non-essential industry, deep processing of black fungus is an inevitable trend. Developing distinctive, high-value-added products can boost farmers' incomes and achieve a win-win situation for all parties.

[0005] A large number of research results show that fermented milk contains relatively high levels of vitamins and proteins, which are easily absorbed by the intestines. Fermented milk can enhance digestion, promote appetite, adjust the body's intestinal flora, and prevent intestinal diseases. In addition, it also has important physiological activities such as lowering cholesterol and anti-tumor. Therefore, fermented milk is highly favored by people and has become one of the most popular dairy products on the Chinese market.

[0006] Common probiotics include Bifidobacterium, Lactobacillus plantarum, Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus bulgaricus, and other strains. These probiotics can effectively colonize the intestines, inhibiting the invasion and growth of harmful intestinal bacteria, strengthening the intestinal barrier, and helping the body fully absorb and utilize nutrients while also stimulating immune cells, enhancing the ability of macrophages to phagocytose pathogens, and improving the body's immune system.

[0007] Bifidobacterium longum is a type of probiotic. As a beneficial intestinal microorganism, Bifidobacterium longum plays an important role in maintaining the health of the body. It has physiological functions such as preventing and treating constipation, inhibiting intestinal pathogens, regulating intestinal balance, lowering cholesterol, promoting digestion and absorption of nutrients, delaying aging, and enhancing the body's immune activity. As research on the health benefits of Bifidobacterium longum continues to deepen, various active drugs and foods using Bifidobacterium longum have been launched and are becoming increasingly popular. Among them, fermented milk is a product in which Bifidobacterium longum is relatively mature and popular with consumers. In order to achieve these effects, Bifidobacterium longum must remain active and metabolically stable in both the product and the host, and reach a certain number (usually considered to be no less than 10 in the product). 7 cfu / g or 10 7 cfu / mL) is particularly important. However, during the processing, transportation, storage and digestion process, it is often affected by adverse factors such as processing conditions, storage temperature, host digestive system (gastric acid, bile salts, enzymes), etc., which leads to a decrease in the number and activity of bacteria, or the number of live bacteria that eventually colonize the intestine is far lower than the minimum threshold that can theoretically exert its probiotic effect. Microencapsulation refers to the process of encapsulating tiny particles with natural or synthetic polymer materials to form semi-permeable particles with a particle size of several millimeters or even several microns. Therefore, the technology of microencapsulation can effectively protect Bifidobacterium longum from adverse environments. We have previously used pectin as a carrier to develop composite probiotic microcapsules (patent number: CN107125767A).

[0008] The preparation method of probiotic microcapsules and the related technical processes used have been gradually applied to related fields such as biotechnology, chemical engineering, and food industry. In the production process of probiotic products, probiotics are embedded by microencapsulation technology. On the one hand, the survival rate of probiotics in the gastrointestinal tract can be improved due to the protective effect of the microcapsule wall material, thereby improving its probiotic effect. On the other hand, the special flavor (especially bifidobacteria) produced by bacterial metabolism can be masked to a certain extent, thereby reducing the impact on product quality. Therefore, the black fungus fermented milk containing long bifidobacterium microcapsules obtained by the present invention has a better body regulation effect than traditional fermented milk or traditional bifidobacterium food. Summary of the Invention

[0009] The first purpose of the present invention is to solve the problem of low viable bacteria count and poor taste of Bifidobacterium longum after fermented milk is made from Bifidobacterium longum, by improving the activity of Bifidobacterium longum through microencapsulation and shielding the problem of poor flavor caused by Bifidobacterium longum participating in fermentation; the second purpose is to use fermented milk as a carrier to achieve full utilization of black fungus and improve its added value.

[0010] Based on the above purpose, the present invention adopts probiotics embedding technology to prepare Bifidobacterium longum into microcapsules, and then enzymatically hydrolyzes black fungus, and then prepares black fungus fermented milk together with the microencapsulated Bifidobacterium longum.

[0011] The black fungus fermented milk developed by the present invention is suitable for consumption by a wide range of people. It has a sweet and sour flavor, a delicate taste, and promotes gastrointestinal digestion. The microcapsules protect Bifidobacterium longum from being damaged by gastric acid and bile salts, allowing it to reach the intestines and exert its beneficial effects. This product also provides new ideas for the deep processing of black fungus products in my country and the development of the probiotic food and health food markets.

[0012] The plate count method can be used to determine the number of viable bacteria in black fungus fermented milk of this product is 10.99±0.06lgcfu / g. According to the "Regulations on the Application and Review of Probiotic Health Foods (Trial)", the number of viable bacteria per gram of probiotic health food shall not be less than 10 during its shelf life. 6 Therefore, the product of the present invention complies with relevant national requirements. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 This is a photo of Bifidobacterium longum microcapsules;

[0014] Figure 2 This is a photo of black fungus fermented milk;

[0015] Figure 3 This is the graph showing the change in viable bacterial count (cfu / g) of black fungus fermented milk during the simulated digestive tract period;

[0016] Figure 4This is the graph showing the change in viable bacterial count (cfu / g) during the 21-day storage period of black fungus fermented milk;

[0017] Figure 5 This is the pH change diagram of black fungus fermented milk during 21 days of storage;

[0018] Figure 6 This is a graph showing the acidity changes of black fungus fermented milk during 21 days of storage;

[0019] Figure 7 This is a graph showing the changes in water holding capacity of black fungus fermented milk during 21 days of storage.

[0020] Explanation of the attached table

[0021] Table 1 Orthogonal experimental scheme and results of black fungus fermented milk

[0022] Table 2 Variance analysis table of black fungus fermented milk

[0023] Table 3 Total bacterial count lg (cfu / g) after fermented milk simulated gastrointestinal tract treatment

[0024] Table 4 Viable bacteria count lg (cfu / g) during storage of black fungus fermented milk

[0025] Table 5 pH changes of black fungus fermented milk during storage

[0026] Table 6 Changes in acidity of black fungus fermented milk during storage

[0027] Table 7 Changes in water holding capacity of black fungus fermented milk during storage period. DETAILED DESCRIPTION

[0028] The exemplary embodiments of the present invention and the description thereof are used to explain the present invention but are not intended to limit the present invention.

[0029] The content of the present invention is not limited to the content of the above-mentioned embodiments. The combination of one or more specific embodiments can also achieve the purpose of the invention.

[0030] Example

[0031] The preparation method of a black fungus fermented milk of the present embodiment is achieved by the following steps:

[0032] 1. Inoculate activated Bifidobacterium longum into MRS+L-cysteine ​​hydrochloride liquid culture medium and culture at 37°C for 24 hours to obtain Bifidobacterium longum culture solution; then centrifuge and discard the supernatant to obtain a Bifidobacterium longum bacterial pellet; 2. Wash the Bifidobacterium longum bacterial pellet with sterile water, centrifuge and discard the supernatant, then dilute to volume with physiological saline at a bacterial sludge: physiological saline mass volume ratio of 1g:5mL to obtain a bacterial concentrate;

[0033] 3. Prepare a 2% pectin solution by weight, add an equal volume of the pectin solution to the bacterial concentrate prepared in step 2, and shake until uniformly mixed to obtain a mixed solution; dropwise add the mixed solution into a sterile CaCl2 solution having a concentration of 100 to 500 mmol / L to obtain wet capsules;

[0034] Fourth, the wet microcapsules were placed at 4°C for 60 minutes to solidify, and then filtered, and the capsule surface was rinsed with sterile water to obtain rinsed microcapsules;

[0035] 5. Soak the black fungus at room temperature for 2 hours, then crush it in a wall-breaking machine according to the mass volume ratio of black fungus to distilled water of 1g:60mL, and then make it into fungus pulp in a colloid mill (the black fungus should turn white in color).

[0036] 6. Weigh cellulase and pectinase and dilute them with 10 times the volume of water to dissolve them, add them to the black fungus pulp, place them in a water bath, 50°C, enzymolysis time 3.5h, take them out after the time is up, and sterilize them under high pressure at 121°C for 20min;

[0037] 7. Mix skim milk powder and sodium carboxymethyl cellulose evenly, add to whole milk, stir evenly, add white sugar and black fungus liquid, mix evenly, and sterilize;

[0038] 8. Fermenting the Bifidobacterium longum microcapsules and milk together at 42° C. for 6 hours to obtain black fungus fermented milk.

[0039] The specific operation of activating Bifidobacterium longum in step 1 is as follows: the Bifidobacterium longum strain preserved in glycerol at -20°C is thawed at room temperature for a while, inoculated into 5 mL of liquid TPY medium, cultured in a constant temperature incubator at 37°C for 24 hours, and then the revived strain is streaked into solid TPY medium at 37°C and anaerobically cultured for 48 hours. This step is repeated until the bacteria are fully revived; finally, the fermentation liquid with the best bacterial activity is inoculated into sterilized MRS+L-cysteine ​​hydrochloride liquid medium at an inoculum size of 5%, and expanded culture is carried out at 37°C for 24 hours.

[0040] In step 1 of this example, the inoculation amount of activated Bifidobacterium longum is 5%.

[0041] In this example, the Bifidobacterium longum strain, numbered CCFM 760, was purchased from the Culture Collection Center of the Institute of Microbiology, Chinese Academy of Sciences.

[0042] The MRS+L-cysteine ​​hydrochloride liquid culture medium in this embodiment was prepared as follows: 48.3 g of finished MRS culture medium, 0.5 g of L-cysteine ​​hydrochloride, 1 L of distilled water, heated and boiled to dissolve, packaged, and sterilized by high-pressure steam at 121° C. for 15 min.

[0043] The TPY liquid culture medium in this embodiment was prepared as follows: 38 g of finished MRS culture medium, 0.5 g of L-cysteine ​​hydrochloride, 1 L of distilled water, heated and boiled to dissolve, divided and packaged, and sterilized by high-pressure steam at 121° C. for 15 min.

[0044] The height at which the mixed solution in this example is added to the sterile CaCl2 solution and the rotation speed of the vortex oscillator determine the size of the microcapsules, while the amounts of skim milk powder, white sugar, black fungus liquid, and sodium carboxymethyl cellulose added determine the quality of the black fungus fermented milk product. The fermented milk prepared in this embodiment has a milky white appearance, a granular texture, a smooth texture, a uniform and stable texture, and no whey precipitation.

[0045] The black fungus fermented milk of this embodiment was subjected to the following experimental verification:

[0046] 1. Single-factor experiment: The amount of black fungus liquid, skim milk powder, and sodium carboxymethyl cellulose added were used as single factors, and fuzzy mathematical sensory scoring was used as the evaluation indicator to determine the addition amount of each single factor. The experimental conclusion was that the sensory scores were the highest when the black fungus liquid was added at 10%, the skim milk powder was added at 4%, and the sodium carboxymethyl cellulose was added at 1.75%, which was the highest value in the single-factor experiment.

[0047] 2. Orthogonal experiment: According to the design principle of orthogonal experiment, a three-factor three-level orthogonal experiment was designed. The experimental design and experimental results are shown in Table 1.

[0048] The variance analysis of the orthogonal experiment is shown in Table 2.

[0049] The influence of various factors on the formula of black fungus fermented milk: the addition amount of black fungus liquid > the addition amount of skimmed milk powder > the addition amount of sodium carboxymethyl cellulose.

[0050] Through single factor experiment and orthogonal experiment, the optimal formula of black fungus fermented milk was obtained as follows: the addition amount of black fungus liquid was 10%, the addition amount of skim milk powder was 4%, and the addition amount of sodium carboxymethyl cellulose was 1.75%.

[0051] 3. Evaluate the quality of the best formula of this embodiment.

[0052] For fermented milk, pH, acidity and viable bacteria count are relatively important factors, especially for fermented milk with added Bifidobacterium longum microcapsules. Its viable bacteria count is of utmost importance and is the key to ensuring its probiotic function. The specific steps are: a: Use a pH meter to measure the pH of the fermented milk, and measure three parallels for each sample. b: Determine the acidity of the fermented milk by acid-base titration: For acid-base titration, weigh 10g of fermented milk sample in a 150mL conical flask, mix it evenly with 20mL of distilled water containing 2-3 drops of phenolphthalein, and titrate it with 0.1mol / L NaOH standard solution until the solution just turns slightly red and does not fade within 30s. The titration is complete. Record the volume V of the consumed solution, set up three parallels for each sample, and calculate using the formula: acidity (°T) = V × 10. c) Take an appropriate amount of fermented milk sample from each group, add EDTA to break the microcapsules, centrifuge at 1699 × g for 10 minutes, wash with sterile water until the supernatant is clear and transparent, dilute with physiological saline in a gradient, and spread the appropriate gradient on a plate, aspirating 100 μL for each gradient. Incubate at 37°C for 48 hours and count the cells. Set up three gradients for each sample.

[0053] The pH of the fermented milk was measured to be around 4.4, with an acidity of 103.3°T, which is greater than the 70°T stipulated in the national standard. Therefore, it complies with the GB19302-2010 National Food Safety Standard for Fermented Milk. The number of viable bacteria in the fermented milk was 10.99±0.06lgcfu / mL. According to the "Regulations on the Application and Review of Probiotic Health Foods (Trial)", the number of viable bacteria per gram of probiotic health foods should not be less than 10 during their shelf life. 6 CFU. Therefore, this product complies with relevant national requirements.

[0054] [Protein] According to the national standard GB5009.3, the Kjeldahl method was used for testing, and the average protein content was 3.38% ± 0.1, which meets the requirements of the national standard for fermented milk.

[0055]

Fat

[0056] [Acidity] The acid-base titration method was used, and the measured acidity was 103.3°T±2.31, which is greater than the national standard of 70°T and meets the national standard for fermented milk.

[0057] [Ash content] According to the national standard GB5009.4, the ash is directly weighed after burning and the calculated value is 1.02%±0.14.

[0058] 4. The fermented milk was subjected to a simulated gastric juice experiment, and the live bacteria count was performed to detect its tolerance and gastric juice release.

[0059] Specific steps are: respectively to the fermented milk, add 5ml artificial gastric juice, placed in 37℃ constant temperature shaking bed, 180 r / min vibration 2h; 5000r / min centrifugal 20min, discard supernatant, collect the bottom precipitate, to the fermented milk precipitate add 5mL EDTA solution completely broken microcapsule after centrifugal 20min, wash 2-3 times, until the supernatant is no longer turbid, discard supernatant after adding 4 times the volume of physiological saline, mix evenly after gradient dilution, take 100 μL diluent coating plate, 37℃ culture 48h, then carry out colony count; compare and analyze the viable count after artificial gastric juice treatment.

[0060] Five, the prepared fermented milk simulates bile in vitro experiment, the prepared Bifidobacterium longum microcapsule tablet is counted, and the tolerance and bile release are detected.

[0061] Specific steps are: respectively to the artificial gastric juice treated fermented milk, add 5ml artificial bile, placed in 37℃ constant temperature shaking bed, 180r / min vibration 20min; 5000r / min centrifugal 20min, discard supernatant, collect the bottom precipitate, to the fermented milk precipitate add 5mL EDTA solution completely broken microcapsule after centrifugal 20min, wash 2-3 times, until the supernatant is no longer turbid, discard supernatant after adding 4 times the volume of physiological saline, mix evenly after gradient dilution, take 100 μL diluent coating plate, 37℃ culture 48h, then carry out colony count; compare and analyze the viable count after artificial bile treatment.

[0062] Six, the prepared fermented milk simulates intestinal fluid in vitro experiment, the prepared fermented milk is counted, and the tolerance and intestinal fluid release are detected.

[0063] Specific steps are: respectively to the artificial gastric juice treated fermented milk, add 5ml artificial intestinal fluid, placed in 37℃ constant temperature shaking bed, 180r / min vibration 20min; 5000r / min centrifugal 20min, discard supernatant, collect the bottom precipitate, to the fermented milk precipitate add 5mL EDTA solution completely broken microcapsule after centrifugal 20min, wash 2-3 times, until the supernatant is no longer turbid, discard supernatant after adding 4 times the volume of physiological saline, mix evenly after gradient dilution, take 100 μL diluent coating plate, 37℃ culture 48h, then carry out colony count; compare and analyze the viable count after artificial intestinal fluid treatment.

[0064] The fermented milk obtained in this example was subjected to experiments simulating the human digestive tract, as shown in Table 3. After treatment with gastrointestinal fluid, the fermented milk containing the black fungus enzymatic hydrolyzate had a survival rate of 79.79%, demonstrating the excellent protective effect of the black fungus enzymatic hydrolyzate. These experimental results demonstrate that the black fungus enzymatic hydrolyzate effectively protects the microcapsules, increasing the survival rate of Bifidobacterium longum microcapsules and enabling them to exert a more effective probiotic effect in the intestine.

[0065] VII. Storage Period Experiment: Fermented milk was stored in a refrigerator at 4°C for 21 days. Samples were collected and the viable bacterial count in the fermented milk was determined. 10 mL of EDTA solution (pH 8.0) was added to each batch of fermented milk. The mixture was shaken at 37°C and 180 rpm / min for 20 minutes to completely disrupt the microcapsules. Appropriate dilutions were then plated and counted for viable bacterial counts. Microencapsulated milk containing Bifidobacterium longum and fermented milk containing black fungus pulp served as controls, treated under the same conditions as the microencapsulated milk.

[0066] The stability test of black fungus fermented milk during storage (as shown in Table 4) shows that black fungus fermented milk has good stability. After 21 days of storage, the number of live bacteria in the microcapsule fermented milk containing Bifidobacterium longum decreased by 1.28lg cfu / g, while the number of live bacteria in the microcapsule fermented milk containing Bifidobacterium longum added with black fungus pulp decreased by 1.39lg cfu / g; the number of live bacteria in our black fungus fermented milk decreased by 1.19lg cfu / g. The number of live bacteria in the black fungus fermented milk during storage was 9.83lg cfu / g, far exceeding the national standard of 6lg cfu / g (10 6 cfu / g), and the color and flavor of black fungus fermented milk are relatively stable, which meets the standards of probiotic live bacteria health food.

[0067] pH is an important indicator for evaluating the quality of fermented milk. Changes in pH during storage can, to a certain extent, reflect the degree of post-acidification. Table 4 shows the pH of the fermented milks in each group at 4°C over the 21-day storage period. As can be seen, the pH of the fermented milks in all three groups decreased overall. The acidity of the milk fermented with Bifidobacterium longum microencapsulated microencapsulated milk changed significantly, decreasing from 4.47 to 3.94. The pH of the milk fermented with Bifidobacterium longum microencapsulated ... milk was consistently lower than that of the other two groups.

[0068] pH represents the free H in fermented milk. +The lactic acid bacteria will continue to grow and reproduce, consume the remaining sugar to produce lactic acid, and the acidity will continue to rise, and the pH value will also continue to decline. The pH values of the three groups of fermented milk showed a downward trend, and the change of the B. longum microcapsule fermented milk group was very large. B. longum has strong acid-producing capacity, although the embedding technology can well protect B. longum and avoid the participation of the fermentation process, reduce its negative impact on yogurt quality and flavor taste, etc. However, the addition of black fungus liquid can slow down the post-acidification of B. longum microcapsule fermented milk. The reason may be that some active substances in black fungus can prevent the continuous fermentation of lactic acid bacteria.

[0069] The titration acidity represents the ability of water and strong alkali, and in the neutralization process, the fermented milk can react with the number of H+ of strong alkali, and the titration acidity of fermented milk is simply called acidity. As shown in Table 6, the acidity of fermented milk during storage. The titration acidity can also reflect the degree of post-acidification to a certain extent, which shows the same trend as the pH value. From the table we can see that on the 0th day, the initial acidity of the three groups of fermented milk is 103.3 °T, 105.3 °T, 103.3 °T, respectively. In the 0-7th day, each group of fermented milk showed an increasing trend in acidity, and the group added with black fungus liquid and black fungus enzyme liquid reached the maximum value of 137.5 °T and 123 °T on the 7th day. The B. longum microcapsule fermented milk group still showed an increasing trend from the 7th day to the 14th day, reaching a maximum of 134.67 °T, while the fermented milk added with black fungus liquid and black fungus enzyme liquid showed a downward trend, and by the 21st day, the acidity of each group of fermented milk reached 134 °T, 147 °T, 121 °T. Too high acidity is not a very good thing for fermented milk, too high acidity will cause the fermented milk to have too much sour taste, whey precipitation, sensory quality decline, and the taste and flavor of yogurt will change greatly. This shows that the addition of black fungus enzyme liquid can alleviate the post-acidification of fermented milk. This can keep the quality of fermented milk within a good range.

[0070] Water holding capacity influences the taste and texture of yogurt and is a key factor influencing the quality of fermented milk. It is generally related to the total solids content and protein content of the fermented milk. Higher water holding capacity indicates that the macromolecules in the gel can bind more water molecules through stronger forces, resulting in greater stability. Table 7 shows the water holding capacity of each fermented milk group over a 21-day storage period. As can be seen, the water holding capacity of each group decreased over time. Starting from day 0, the water holding capacity of the milk fermented with Bifidobacterium longum microencapsulated was significantly lower than that of the groups supplemented with black fungus liquid and the group supplemented with black fungus enzymatic hydrolyzate. The initial water holding capacities of the three groups were 84.53%, 96.55%, and 96.92%, respectively. At the end of storage, the water holding capacities of the two groups supplemented with black fungus liquid remained higher than those of the milk fermented with Bifidobacterium longum microencapsulated, indicating that the addition of black fungus liquid enhances the water retention of the fermented milk. This may be because the polysaccharide components in black fungus and the addition of polysaccharides interact with proteins in the gel structure of yogurt in the form of chemical bonds during the fermentation process, thereby enhancing the gel structure, retaining moisture more effectively, preventing excessive precipitation of whey, and improving its water holding capacity.

[0071] Serial number Addition amount of fungus Amount of milk powder added CMC addition amount blank Fraction 1 1(8％) 1(3％) 1(1.5％) 1 6.75±0.34 2 1 2(4％) 2(1.75％) 2 7.23±0.66 3 1 3(5％) 3(2％) 3 6.87±0.84 4 2(10％) 1 2 3 7.52±0.55 5 2 2 3 1 7.69±0.19 6 2 3 1 2 7.46±0.28 7 3(12％) 1 3 2 6.11±0.31 8 3 2 1 3 6.3±0.48 9 3 3 2 1 6.36±0.24 K1 6.950 6.793 6.837 6.933 K2 7.557 7.073 7.037 6.933 K3 6.257 6.890 6.890 6.897 Range R 1.300 0.280 0.200 0.036

[0072] factor Sum of Squared Deviations degrees of freedom F ratio F critical value Significance fungus 2.539 2 3.726 3.110 * milk powder 0.120 2 0.176 3.110 CMC 0.064 2 0.094 3.110 error 2.73 8

[0073] Table 3

[0074]

[0075] Table 4

[0076]

[0077] Table 5

[0078]

[0079] Table 6

[0080]

[0081] Table 7

[0082]

Claims

1. A method for preparing black fungus fermented milk containing microcapsules of Bifidobacterium longum, characterized in that It is implemented as follows:

1. Inoculate the activated Bifidobacterium longum into MRS+L-cysteine ​​hydrochloride liquid culture medium and culture at 37°C for 24 hours to obtain a Bifidobacterium longum culture solution; then centrifuge the solution and remove the supernatant to obtain a Bifidobacterium longum bacterial pellet; 2. Wash the Bifidobacterium longum bacterial precipitate with sterile water, and then mix it with normal saline at a mass volume ratio of 1 g of bacterial sludge to 5 mL of normal saline to obtain a bacterial concentrate; 3. Prepare a low-methoxyl pectin solution with a mass percentage of 0.5% to 4%, mix the pectin solution with a mass concentration of 2% and the bacterial concentrate in step 2 in equal volumes, and drop the mixture into a sterile CaCl2 solution with a concentration of 100 to 500 mmol / L to obtain wet capsules; Fourth, the wet microcapsules were placed at 4°C for static crosslinking for 20-120 minutes, then filtered, and the capsule surface was rinsed with sterile water to obtain rinsed microcapsules; 5. After soaking the black fungus at room temperature for 2 hours, crush it in a wall-breaking machine according to the mass volume ratio of black fungus to distilled water of 1g:60mL, and then make it into black fungus pulp in a colloid mill; 6. Weigh cellulase and pectinase and dilute and dissolve them with 10 times the volume of water, add them to the black fungus pulp, place them in a water bath, 50℃, and enzymolysis time 3.5h. After the time is up, take them out and sterilize them by high pressure at 121℃ for 20min to inactivate the enzymes; the mass ratio of cellulase to pectinase is 3:2; 7. Mix skim milk powder and sodium carboxymethyl cellulose evenly, add to whole milk, stir evenly, add white sugar and black fungus liquid, mix evenly, and sterilize; 8. Fermenting the Bifidobacterium longum microcapsules and milk together at 42° C. for 6 hours to obtain black fungus fermented milk.

2. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that The method for activating Bifidobacterium longum is as follows: after taking out Bifidobacterium longum strains stored in glycerol at -20°C, inoculating them into 5 mL of liquid TPY culture medium, culturing them in a constant temperature incubator at 37°C for 24 hours, then streaking the revived strains into solid TPY culture medium at 37°C for anaerobically culturing for 48 hours, and repeating this step until the bacteria are completely revived; finally, inoculating the fermentation liquid into sterilized MRS+L-cysteine ​​hydrochloride liquid culture medium at a 5% inoculum volume, and expanding the culture at 37°C for 24 hours.

3. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that The MRS+L-cysteine ​​hydrochloride liquid culture medium is obtained by adding 1 L of distilled water to a mixture of 48.3 g of MRS culture medium and 0.5 g of L-cysteine ​​hydrochloride, heating and boiling to dissolve, packaging, and autoclaving at 121° C. for 15 minutes.

4. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that The second step is to obtain a concentrated solution of bacteria by washing the Bifidobacterium longum sludge with sterilized saline, centrifuging, discarding the supernatant, and diluting the solution with saline at a mass volume ratio of 1 g:5 mL to obtain a concentrated solution of naked Bifidobacterium longum.

5. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that The mass ratio of cellulase and pectinase is 3:

2. Taking 50g of black fungus pulp as an example, there are 0.96g of cellulase and 0.64g of pectinase.

6. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that In step six, the amount of skimmed milk powder added is 4%, the amount of sodium carboxymethyl cellulose added is 1.75%, the amount of white sugar added is 7%, and the amount of black fungus liquid added is 10%.

7. The method for preparing the Bifidobacterium longum-loaded microcapsule fungus fermented milk according to claim 1, characterized in that The amount of microcapsules added in step seven is 1 g of Bifidobacterium longum microcapsules per cup of fermented milk.

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