Postbiotic, preparation method thereof and application of postbiotic in yoghurt

Abstract

The invention discloses a postbiotic and a preparation method and application thereof in yoghourt, the postbiotic takes bacillus subtilis GDAAS-A32 or / and metabolite of the bacillus subtilis GDAAS-A32 as an active ingredient, and the preservation number of the bacillus subtilis GDAAS-A32 is GDMCC No.62816. The invention further discloses a preparation method of the postbiotic and application of the postbiotic in yoghourt. The bacillus subtilis GDAAS-A32 is inoculated into a culture medium to be cultured, and a cultured substance is the metagen. The bacillus subtilis GDAAS-A32 is firstly inoculated into an LB culture medium to be cultured and then inoculated into an M9 culture medium to be cultured. The culture temperature in the culture medium is 30-50 DEG C, and the rotating speed is 120-250 rpm. The metabiotics are used for preparing yoghourt. According to the method, the quality of the yoghourt is improved, the viable count of probiotics in the yoghourt is increased by 10 times, and good application potential is shown.

Description

Technical Field

[0001] The present invention relates to the field of food technology, and more specifically, to a postbiotic, a preparation method thereof, and an application thereof in yogurt. Background Art

[0002] Yogurt is fermented by lactic acid bacteria and is rich in protein, polysaccharides and other nutrients. During the fermentation process, the protein in yogurt is degraded into amino acids and polysaccharides are converted into monosaccharides, which facilitate digestion and absorption. The widespread popularity and high consumption of yogurt make it an attractive choice for adding various value-added ingredients such as probiotics, prebiotics, plant fibers and extracts.

[0003] In the production and preservation of yogurt, microbial activity plays a vital role in the quality of yogurt. First of all, microbial activity is the core of the yogurt fermentation process. Lactic acid bacteria such as thermophilic Streptococcus and Bulgarian Lactobacillus convert lactose in milk into lactic acid through metabolism, causing the pH value of milk to drop and coagulate into yogurt. This fermentation process not only gives yogurt its unique sour taste and texture, but also promotes the release and absorption of nutrients in yogurt. Secondly, microbial activity has an important influence on the taste and flavor of yogurt. Lactic acid and other metabolites produced by lactic acid bacteria during the fermentation process, such as acetaldehyde and pyruvic acid, together constitute the unique flavor of yogurt. At the same time, the metabolic activity of lactic acid bacteria can also produce volatile compounds such as ethyl acetate, which also makes an important contribution to the aroma of yogurt. Therefore, the level of microbial activity directly affects the taste and flavor quality of yogurt. In addition, microbial activity is also closely related to the shelf life and nutritional value of yogurt. During the storage of yogurt, the activity of lactic acid bacteria will gradually decrease, but their presence still plays an important role in maintaining the quality of yogurt. my country stipulates that the activity of probiotics in yogurt during the storage period shall not be less than 10 6 CFU / mL is one of the core indicators of yogurt quality.

[0004] Probiotics produce water-soluble bioactive compounds called "postbiotics", which include various metabolites such as bioactive lipids, antimicrobial peptides and exopolysaccharides. These compounds have various biological activities such as anti-inflammatory, antibacterial, anti-diabetic, anti-cancer, immunomodulatory, anti-atherosclerotic and anti-obesity. However, there is no report that postbiotics can effectively enhance the activity of probiotics and thus improve the quality of yogurt. Therefore, the present invention provides a postbiotic that can effectively improve the quality of yogurt. Summary of the invention

[0005] An object of the present invention is to solve at least the above problems and to provide at least the advantages which will be described hereinafter.

[0006] Another object of the present invention is to provide a postbiotic, which improves the quality of yogurt and increases the number of live probiotics in yogurt by 10 times, showing good application potential.

[0007] In order to achieve these purposes and other advantages according to the present invention, a postbiotic is provided, wherein the postbiotic uses Bacillus subtilis GDAAS-A32 or / and its metabolites as active ingredients, wherein the deposit number of Bacillus subtilis GDAAS-A32 is GDMCC No.62816.

[0008] The present invention further provides a method for preparing a postbiotic, wherein the Bacillus subtilis GDAAS-A32 is inoculated into a culture medium for culture, and the cultured substance is the postbiotic.

[0009] Preferably, the Bacillus subtilis GDAAS-A32 is inoculated in LB or M9 medium for culture, and then inoculated in M9 medium for culture.

[0010] Preferably, the culture temperature in the culture medium is 30-50°C and the rotation speed is 120-250 rpm.

[0011] The present invention further provides an application of a postbiotic in yogurt, wherein the postbiotic is used for preparing yogurt.

[0012] Preferably, the preparation method of yogurt is: first mixing the milk base with the postbiotics, then adding the starter to obtain a fermentation liquid, and fermenting the fermentation liquid at 35-45°C for 5-8h to obtain yogurt, wherein the content of the postbiotics in the fermentation liquid is 10-20% (v / v).

[0013] Preferably, the milk base is fresh milk or milk powder.

[0014] Preferably, the fermentation agent comprises thermophilic streptococcus and bulgaricus lactobacillus, and the usage ratio of bulgaricus lactobacillus to thermophilic streptococcus is 1:5-30.

[0015] The present invention has at least the following beneficial effects:

[0016] First, the new postbiotics prepared by the present invention can significantly improve the activity of lactic acid bacteria, and the effect is stronger than that reported previously.

[0017] Second, the novel postbiotic prepared by the present invention can increase the rennet activity in yogurt by more than 87%.

[0018] Third, the new postbiotics prepared by the present invention improve the quality of yogurt and increase the number of live probiotics in yogurt by 10 times, showing good application potential.

[0019] Fourthly, the novel postbiotics prepared by the present invention can improve the viscosity and water holding capacity of yogurt and slow down the rate at which yogurt goes sour.

[0020] Other advantages, objectives and features of the present invention will be embodied in part through the following description, and in part will be understood by those skilled in the art through study and practice of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 The pH value during the yogurt fermentation process according to one of the technical solutions of the present invention;

[0022] Figure 2 The measurement result of the changes of Lactobacillus bulgaricus during the yogurt fermentation process described in one of the technical solutions of the present invention;

[0023] Figure 3 The measurement result of the changes of Streptococcus thermophilus during the yogurt fermentation process described in one of the technical solutions of the present invention;

[0024] Figure 4 This is a photograph of the yogurt fermentation at the end of one of the technical solutions of the present invention;

[0025] Figure 5 The sensory evaluation result at the end of yogurt fermentation according to one of the technical solutions of the present invention;

[0026] Figure 6 A comparison chart of the overall preference values ​​at the end of yogurt fermentation according to one of the technical solutions of the present invention;

[0027] Figure 7 The viscosity measurement result of yogurt at the end of fermentation according to one of the technical solutions of the present invention;

[0028] Figure 8 The measurement result of the water holding capacity of yogurt at the end of fermentation according to one of the technical solutions of the present invention;

[0029] Fig. 9 The result of measuring rennet activity at the end of yogurt fermentation according to one of the technical solutions of the present invention;

[0030] Fig.10 The measurement result of pH change of yogurt during storage according to one of the technical solutions of the present invention;

[0031] Fig.11 The measurement result of the change of the acidity of the yogurt during storage according to one of the technical solutions of the present invention;

[0032] Fig.12The results of the determination of Lactobacillus bulgaricus during the storage of yogurt according to one of the technical solutions of the present invention;

[0033] Fig.13 This is the measurement result of thermophilic Streptococcus during the storage of yogurt according to one of the technical solutions of the present invention. DETAILED DESCRIPTION

[0034] The present invention is further described in detail below in conjunction with embodiments so that those skilled in the art can implement the invention with reference to the description.

[0035] <Example 1>

[0036] A postbiotic, wherein the postbiotic uses Bacillus subtilis GDAAS-A32 or / and its metabolites as active ingredients, wherein the deposit number of Bacillus subtilis GDAAS-A32 is GDMCC No.62816.

[0037] The Bacillus subtilis GDAAS-A32 is inoculated into a culture medium for cultivation, and the cultured substance is the postbiotic. The specific cultivation method is: streak the Bacillus subtilis GDAAS-A32 on an LB plate culture medium, place it in a 37°C incubator and invert it for 12 hours, pick a single colony into an M9 liquid culture medium, place it in a 37°C, 120rpm shaker and culture it until the OD 600 The centrifugal force is 6000 (×g) and the centrifugal force is 0.5-0.7 (centrifugation is performed after the shaking table culture is completed). The supernatant obtained is the postbiotics.

[0038] The preparation method of yogurt is as follows: a milk base is first mixed with a postbiotic, and then a starter is added to obtain a fermented milk, and the fermented milk is fermented at 35 to 45° C. for 5 hours to obtain yogurt, wherein the amount of the postbiotic in the fermented milk is 10% (v / v), and the amount of the starter is 10% (v / v).

[0039] The milk base material is milk powder (the milk powder is dissolved in water, the milk powder content is 20%, and the mixture is placed at 75° C. for pasteurization for 30 minutes and cooled).

[0040] Preparation method of starter culture: single colonies of thermophilic Streptococcus and Bulgarian Lactobacillus were selected and cultured in MRS liquid culture medium at 30°C until OD 600 The mixture is evenly mixed at a ratio of 1.0 to 1.5 to obtain a mixed bacterial solution, which is centrifuged at a centrifugal force of 6000 (×g). The precipitate is taken and washed 2-3 times with sterile water to obtain a fermentation agent.

[0041] The fermentation agent includes thermophilic streptococcus and bulgaricus lactobacillus, and the dosage ratio of bulgaricus lactobacillus to thermophilic streptococcus is 1:20.

[0042] <Example 2>

[0043] The postbiotics and yogurt were prepared by the method of Example 1, wherein the difference was that the amount of the postbiotics in the fermented milk was 20% (v / v).

[0044] <Example 3>

[0045] The method of Example 1 was used to prepare postbiotics and yogurt, except that the milk base was fresh milk.

[0046] <Example 4>

[0047] The postbiotics and yogurt were prepared by the method of Example 1, wherein the difference was that the milk base was fresh milk, and the amount of the postbiotics in the fermented milk was 20% (v / v).

[0048] <Comparative Example 1>

[0049] Yogurt was prepared by the method of Example 1, wherein no postbiotics were added during the preparation of the yogurt.

[0050] <Experiment>

[0051] In the experimental characterization, Example 1 corresponds to Group B, Example 2 corresponds to Group C, and Comparative Example 1 corresponds to Control Group A;

[0052] 1. Effect of adding different concentrations of postbiotics on pH during yogurt fermentation

[0053] A pH meter was used to detect the pH during the yogurt fermentation process. Figure 1 It can be seen that the pH continued to decrease during the fermentation process. There was no significant difference between the experimental group with 10% postbiotics added and the control group (p>0.05), while there was a significant difference between the experimental group with 20% postbiotics added and the control group (p<0.05), that is, the addition of 20% postbiotics accelerated the acid production ability of lactic acid bacteria.

[0054] 2. Effects of adding different concentrations of postbiotics on Lactobacillus bulgaricus during yogurt fermentation

[0055] The detection of lactic acid bacteria during yogurt fermentation was performed by Q-PCR. Figure 2 It can be seen that with the growth of time, Lactobacillus bulgaricus continued to proliferate, and the addition of different concentrations of postbiotics had a certain growth-promoting effect on Lactobacillus bulgaricus. When the concentration of postbiotics was 20%, Lactobacillus bulgaricus grew best, and the number of viable bacteria reached 1.53×10 at the end of fermentation. 8 CFU / g, higher than the control group of 1.71×10 8 CFU / g. The viable count reached 6.27×10 8 CFU / g, which is 3.23×10 8CFU / g is 1.94 times.

[0056] 3. Effects of adding different concentrations of postbiotics on Streptococcus thermophilus during yogurt fermentation

[0057] The detection of lactic acid bacteria during yogurt fermentation was carried out by Q-PCR. Figure 3 It can be seen that thermophilic Streptococcus proliferates continuously with time. Adding different concentrations of postbiotics has different degrees of enhancement effect on the growth of thermophilic Streptococcus. When the concentration of postbiotics is 20%, the thermophilic Streptococcus grows best, and the number of viable bacteria reaches 6.71×10 at the end of fermentation. 8 CFU / g, which was higher than that of the control group (3.53×10 8 CFU / g. The viable count reached 2.29×10 9 CFU / g, which is 2.94×10 8 CFU / g is 7.8 times.

[0058] 4. Effect of adding different concentrations of postbiotics on sensory evaluation at the end of yogurt fermentation

[0059] Sensory analysis results of different groups of yogurt Figure 5 As shown, the trend of smell and texture results is consistent, and there is a significant difference between the experimental group (10% postbiotics in Example 1 and 20% postbiotics in Example 2) and the control group (p<0.05), and there is no significant difference between the experimental groups (p>0.05); the trend of taste and mouthfeel results is consistent, and there is a significant difference between the experimental group and the control group, and there is a significant difference between the experimental groups. It can be seen from the results of room temperature placement (see Figure 4 ): The control group was yellowish in color, with a small amount of whey separation. After being inverted at room temperature for 1 day, the position of the yogurt changed; the experimental group with 10% postbiotics was whiter in color, with no or a small amount of whey separation. After being inverted at room temperature for 1 day, the position of the yogurt remained unchanged, but a large amount of whey precipitated; the experimental group with 20% postbiotics was milky white in color, with no or a small amount of whey separation. After being inverted at room temperature for 1 day, the position of the yogurt remained unchanged, and a small amount of whey precipitated; the sensory evaluation score results showed (see Figure 5 ): The yogurt in the experimental group with 20% postbiotics was milky white in appearance, with a uniform and delicate texture, no whey separation; it tasted slightly sour and delicious, delicate and tender; it contained a strong fermented milk aroma, no bad smell; and it had a mellow taste. The control group was yellowish in color, more uniform in texture, with a small amount of whey separation; it tasted sour and delicate; it contained a lighter fermented milk aroma, and no bad smell. The sensory evaluation results were basically consistent with the results of the room temperature treatment.

[0060] 5. Effect of adding different concentrations of postbiotics on the overall liking of yogurt

[0061] The 9-point Chinese preference scale was used to measure the overall preference differences of different types of yogurt products. Figure 6 It can be seen that the overall preference value distribution range of different types of yogurt is 4.1 to 6.5 points. In the present invention, except for the control group, the preference values ​​of the yogurt prepared by the experimental group with 10% postbiotics and the experimental group with 20% postbiotics are all above 5 points, indicating that consumers tend to like the dairy products prepared in Example 1 and Example 2. There is a significant difference in the preference value between the experimental group with 10% postbiotics and the control group (p<0.05), and the experimental group with 20% postbiotics has the highest significance compared with the control group (p<0.001). There is no significant difference in the overall preference value between the experimental group with 10% postbiotics and the experimental group with 20% postbiotics (p>0.05). Compared with the control group, the experimental group with 10% postbiotics and the experimental group with 20% postbiotics have significantly improved their preference after adding postbiotics, indicating that the addition of postbiotics has a significant effect on improving overall preference.

[0062] 6. Effects of adding different concentrations of postbiotics on yogurt viscosity and water holding capacity

[0063] Depend on Figure 7 , 8 It can be seen that compared with the control group, the viscosity (μ) and water holding capacity (WHC) of the yogurt samples in the experimental group were significantly improved, and the overall viscosity and water holding capacity of the fermented yogurt with 20% postbiotics added were higher, which was consistent with the trend of the results of room temperature storage, indicating that the addition of postbiotics has a significant effect on improving the viscosity and water holding capacity of yogurt.

[0064] 7. Effect of adding different concentrations of postbiotics on yogurt rennet activity

[0065] Curd formation is crucial in the production of fermented dairy products such as yogurt, in which rennet-induced gelation plays a key role. Fig. 9 It can be seen that there are significant differences between the experimental group and the control group. When the fermentation end point is reached, the rennet activity of the control group is 142.51SU / mL, and the rennet activity of yogurt with 10% postbiotics is 178.48SU / mL, which is 25.24% higher than that of the control group; the rennet activity of yogurt with 20% postbiotics is 267.27SU / mL, which is 87.54% higher than that of the control group.

[0066] 8. Effect of adding different concentrations of postbiotics on the pH of yogurt during storage

[0067] A pH meter was used to detect the pH of yogurt during storage. Fig.10It can be seen that the pH continued to decrease during storage. On the first day of storage, the pH of the experimental group with 20% postbiotics was the lowest, and the pH of the control group was the highest. As the storage time increased, the pH of the control group decreased the fastest and had the lowest pH, while the pH of the experimental group with 20% postbiotics decreased the slowest and had the highest pH, that is, the addition of postbiotics slowed down the rate of sourness of yogurt during storage.

[0068] 9. Effect of adding different concentrations of postbiotics on the acidity of yogurt during storage

[0069] Changes in acidity (TA) during yogurt storage Fig.11 As shown in the figure, as the storage days increased, the TA of all yogurts continued to rise, and the TA remained below 70°T during the whole process. At the end of storage, the control group reached the highest TA of 63.25°T, followed by the experimental group with 10% postbiotics, with a TA of 58.5°T, and the experimental group with 20% postbiotics had the lowest TA of 55°T, that is, the addition of postbiotics can slow down the rate of yogurt spoilage.

[0070] 10. Effect of adding different concentrations of postbiotics on microbial activity during yogurt storage

[0071] The number of viable Streptococcus thermophilus and Lactobacillus bulgaricus during yogurt storage Fig.12 , 13 As can be seen from the figure, the trends of the number of live thermophilic Streptococcus and Bulgarian Lactobacillus in the fermented yogurt samples of different groups are consistent, showing a gradual downward trend. The number of live bacteria during storage is higher in the experimental group with 20% postbiotics added> the experimental group with 10% postbiotics added> the control group. After 7 days, the number of live thermophilic Streptococcus in the yogurt of the experimental group with 20% postbiotics added is still 6.18×10 8 CFU / mL, Lactobacillus bulgaricus 1.03×10 8 CFU / mL, which were higher than those in the control group (5.49×10 7 and 1.11×10 7 CFU / mL) is 10 times higher, and the number of live bacteria is higher than that of the National Yogurt Association (NYA) in the United States, which stipulates that biological yogurt products must contain 10 8 CFU / g lactic acid bacteria (LAB) are produced using the “live and active culture” logo.

[0072] 11. Characterization of the effect of adding new postbiotics on yogurt prepared from fresh milk

[0073] Postbiotic yogurt was prepared by the method of Example 3 and Example 4. By comparing the water holding capacity, rennet activity and the number of viable probiotics, it was found that with the increase in the amount of the new postbiotics added, the water holding capacity, rennet activity and other capabilities were significantly improved, especially the number of viable bacteria increased by 10 times, and it had higher nutritional value and application prospects.

[0074] Although the embodiments of the present invention have been disclosed as above, they are not limited to the applications listed in the specification and the implementation modes, and they can be fully applied to various fields suitable for the present invention. For those familiar with the art, additional modifications can be easily implemented. Therefore, without departing from the general concept defined by the claims and the scope of equivalents, the present invention is not limited to the specific details and the illustrations shown and described herein.

Claims

1. Postbiotics, characterized in that The postbiotics use Bacillus subtilis GDAAS-A32 or / and its metabolites as active ingredients, wherein the deposit number of Bacillus subtilis GDAAS-A32 is GDMCC No.62816.

2. The method for preparing postbiotics according to claim 1, characterized in that: The Bacillus subtilis GDAAS-A32 is inoculated into a culture medium for culture, and the cultured substance is the postbiotic.

3. The method for preparing postbiotics according to claim 2, characterized in that: The Bacillus subtilis GDAAS-A32 was grown in LB or M9 medium.

4. The method for preparing postbiotics according to claim 2, characterized in that: The culture temperature is 30-50°C and the rotation speed is 120-250rpm.

5. The use of postbiotics in yogurt according to claim 1, characterized in that: The postbiotics are used in the preparation of yogurt.

6. The use of postbiotics in yogurt according to claim 5, characterized in that: The preparation method of yogurt is as follows: a milk base is first mixed with postbiotics, and then a starter is added to obtain fermented milk, and the fermented milk is fermented at 35-45° C. for 5-8 hours to obtain yogurt, wherein the content of postbiotics in the fermentation liquid is 10-20% (v / v).

7. The use of postbiotics in yogurt according to claim 6, characterized in that: The milk-based material is fresh milk or milk powder.

8. The use of postbiotics in yogurt according to claim 6, characterized in that: The fermentation agent includes thermophilic streptococcus and bulgaricus lactobacillus, and the dosage ratio of bulgaricus lactobacillus to thermophilic streptococcus is 1:5-30.

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