A composite prebiotic-embedded probiotic microcapsule and its preparation method and application

Abstract

The present invention relates to the field of food technology, and specifically discloses a composite prebiotic-embedded probiotic microcapsule, a preparation method and an application thereof. The present invention uses green tea leaves as a main raw material, performs water extraction on the microcapsule, and simultaneously adopts ultrasound-assisted treatment, thereby more effectively releasing active ingredients in the green tea leaves, shortening the extraction time, and improving the extraction rate of the active ingredients in the green tea leaves; subsequently, the extract is fermented by Lactobacillus acidophilus, so that the green tea fermentation liquid contains more antioxidant active ingredients, and during the fermentation process, the bound polyphenols in the extract can release small molecular polyphenols, thereby increasing the content of flavonoids, polyphenols and other substances in the green tea fermentation liquid, thereby further improving the antioxidant activity; and then, modified dietary fiber is used as the main wall material of the microcapsule to embed the green tea fermentation liquid and probiotics, thereby improving the stability of the active substances and probiotics in the green tea fermentation liquid, and enabling them to play a protective and sustained-release role in a simulated digestion process.

Description

Technical Field

[0001] The present invention relates to the field of food technology, and in particular to a composite prebiotic-embedded probiotic microcapsule and a preparation method and application thereof. Background Art

[0002] Green tea is a vital component of traditional Chinese tea culture. As one of the leading teas, its unique production process and rich nutritional profile have earned it a global reputation. The polyphenols and catechins found in green tea have been extensively studied, demonstrating their remarkable antioxidant, anti-aging, antibacterial, lipid-lowering, and metabolic benefits. As potent antioxidants, polyphenols can effectively neutralize free radicals, slowing the aging process and thus helping to boost the body's immune system.

[0003] With increasing emphasis on healthy eating, the application of green tea extract has gradually expanded to various fields, including food, health products, pharmaceuticals, and daily chemicals. In particular, green tea extract is widely used in the development of functional foods due to its rich bioactive components. However, because these active ingredients are extremely sensitive to environmental factors such as temperature, light, pH, and oxygen, their practical application faces many challenges. For example, green tea extract may produce an unpleasant taste during certain food processing processes, affecting the taste and quality of the final product.

[0004] In order to overcome these problems, researchers are constantly exploring new technical means, among which microencapsulation technology has become an effective solution. Microencapsulation technology can effectively protect active ingredients from the influence of the external environment by encapsulating them in tiny capsules, thereby maintaining their stability and biological activity. This technology can not only prevent the degradation of active ingredients, but also control their release in food, thereby improving the overall quality of the product. In the microencapsulation process, the selection of encapsulation materials is crucial. Commonly used encapsulation materials include natural polymers such as gelatin, gum arabic, and starch. They not only have good biocompatibility, but also can provide sufficient protection. In addition, the optimization of encapsulation technology, such as adjusting the encapsulation ratio, temperature, and time, will also directly affect the stability of the microcapsules and the release efficiency of the active ingredients.

[0005] Chinese patent document CN202310845151.6 discloses a microcapsule containing green tea extract, its preparation method and application. The whitening composition is prepared by compounding green tea extract with other plant extracts, and then the whitening composition is prepared into microcapsules. The green tea extract and the like are embedded by spray drying encapsulation technology, which can effectively control the release of its active substances, improve its bioavailability and stability, and ensure product quality. However, the stability of the microcapsules still needs to be improved. Summary of the Invention

[0006] In view of the shortcomings of the prior art, the present invention aims to provide a composite prebiotic-embedded probiotic microcapsule and a preparation method and application thereof.

[0007] In order to achieve the above object, the present invention adopts the following technical solutions:

[0008] In a first aspect, the present invention provides a method for preparing composite prebiotic-embedded probiotic microcapsules, comprising the following steps:

[0009] S1. Grind and sieve the green tea leaves, add them to deionized water, then add an amino acid surfactant, perform ultrasonic-assisted extraction, and filter after the extraction to obtain an extract and a filter residue;

[0010] S2, adding the extract to sterilized coconut water, then inoculating Lactobacillus acidophilus, fermenting and culturing, and after the fermentation is completed, filtering and collecting the filtrate to obtain green tea fermentation broth;

[0011] S3, adding the filter residue to a hydrogen peroxide solution for treatment, and after the treatment is completed, filtering, washing, drying, grinding and screening to obtain modified dietary fiber;

[0012] S4. Add inulin, sodium alginate and modified dietary fiber to deionized water and stir evenly. Then add green tea fermentation broth and Lactobacillus rhamnosus powder and mix evenly. Then add zinc lactate solution and continue stirring to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0013] Preferably, in step S1, the mass ratio of green tea leaves, deionized water and amino acid surfactant is 15-25:300-400:0.5-1.

[0014] Preferably, in step S1, the ultrasonic power is 300-600w, the extraction temperature is 70-80°C, and the extraction time is 10-20min.

[0015] Preferably, in step S2, the amount of the extract added is 20-30% by volume of the sterilized coconut water, and the inoculation amount of Lactobacillus acidophilus is 1-3%.

[0016] Preferably, in step S2, during the fermentation culture process, the fermentation temperature is 28-32°C, the fermentation time is 48-72h, and the stirring speed is 100-200r / min.

[0017] Preferably, in step S3, the mass ratio of the filter residue to the hydrogen peroxide solution is 10-20:100, and the mass fraction of the hydrogen peroxide solution is 10-20%.

[0018] Preferably, in step S4, the mass ratio of inulin, sodium alginate, modified dietary fiber, deionized water, green tea fermentation broth, Lactobacillus rhamnosus powder and zinc lactate solution is 0.4-0.8:1-1.5:2-3:10-20:10-15:0.3-0.6:10-20.

[0019] Preferably, in step S4, the mass fraction of the zinc lactate solution is 3-6%.

[0020] In a second aspect, the present invention provides composite prebiotic-embedded probiotic microcapsules prepared by the above-mentioned preparation method.

[0021] In a third aspect, the present invention further provides the use of the above-mentioned composite prebiotic-embedded probiotic microcapsules in the preparation of solid beverage products.

[0022] Compared with the prior art, the present invention has the following beneficial effects:

[0023] (1) The present invention uses green tea leaves as the main raw material and performs water extraction on the green tea leaves. During the water extraction process, an amino acid surfactant is added, which can effectively improve the permeability of the cell wall and effectively dissolve the flavonoids and polyphenols in the cells. At the same time, ultrasound is used as an auxiliary treatment. Ultrasonic waves can generate tiny bubbles and rupture the cell walls through the cavitation effect of ultrasound, thereby more effectively releasing the active ingredients in the green tea leaves, shortening the extraction time, and improving the extraction rate of the active ingredients in the green tea leaves.

[0024] (2) The present invention uses Lactobacillus acidophilus to ferment the extract, and at the same time uses coconut water as a fermentation matrix. Coconut water contains a large amount of nutrients, which reduces the utilization of effective substances in the extract by Lactobacillus acidophilus, so that the green tea fermentation liquid contains more antioxidant active ingredients. In addition, during the fermentation process, the bound polyphenols in the extract can release small molecular polyphenols, thereby increasing the content of flavonoids, polyphenols and other substances in the green tea fermentation liquid, thereby further improving the antioxidant activity; at the same time, Lactobacillus acidophilus can ferment to produce some biologically active peptide substances, which work together with prebiotics such as inulin and sodium alginate to promote the growth and reproduction of beneficial bacteria such as lactic acid bacteria, bifidobacteria and acetic acid bacteria in the intestine. Lactobacillus acidophilus can also be used as a probiotic species to regulate gastrointestinal health.

[0025] (3) The present invention uses modified dietary fiber as the main wall material of the microcapsule, and first uses hydrogen peroxide to treat the surface of the filter residue, with the aim of increasing the activity of the surface groups of the filter residue, increasing the specific surface area of ​​the filter residue, and improving the binding force between the filter residue and inulin and sodium alginate. Subsequently, zinc lactate solution is added, and zinc ions form a gel through chelation with the hydroxyl groups in inulin, sodium alginate and modified dietary fiber, which encapsulates the green tea fermentation broth and rhamnosus lactobacillus, thereby improving the stability of the active substances in the green tea fermentation broth and rhamnosus lactobacillus, and enabling them to play a protective and sustained-release role in the simulated digestion process; at the same time, zinc also has certain antioxidant properties, which can protect intestinal cells from oxidative damage, and work together with the active ingredients in the green tea fermentation broth to further improve the antioxidant effect of the composite prebiotic-encapsulated probiotic microcapsules.

[0026] (4) The composite prebiotic-encapsulated probiotic microcapsules provided by the present invention form a protective barrier on the outer layer of the probiotics, helping the probiotics to resist the damage of digestive juices such as gastric acid and bile, ensuring that more live bacteria reach the intestines; at the same time, the prebiotics provide nutrition for the probiotics after fermentation in the intestines, further enhancing their colonization and reproduction capabilities. This synergistic effect not only improves the survival rate of probiotics, but also more effectively regulates the balance of intestinal flora, improves digestive function, enhances immunity, and alleviates problems such as intestinal inflammation. The composite prebiotic-encapsulated probiotic microcapsules provided by the present invention have broad application prospects in functional foods and health products. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 This is a comparison chart of the in vitro DPPH free radical scavenging rates of different groups of microcapsules;

[0028] Figure 2 This is a comparison chart of the storage stability of microcapsules in different groups;

[0029] Figure 3 This is a comparison chart of the polyphenol extraction rate and flavonoid extraction rate of different groups. DETAILED DESCRIPTION

[0030] The present invention is further described in detail below through specific preferred embodiments, but the present invention is not limited to the following embodiments.

[0031] It should be noted that, unless otherwise specified, all chemical reagents involved in the present invention were purchased through commercial channels.

[0032] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0033] S1. Preparation of extract and filter residue

[0034] The green tea leaves are crushed and sieved and then added into deionized water, and then an amino acid surfactant is added, and ultrasonic-assisted extraction is performed. After the extraction is completed, the extraction is filtered to obtain an extract and a filter residue.

[0035] In this step, the green tea leaves are first crushed and passed through a 40-80 mesh sieve for processing.

[0036] In this step, the mass ratio of green tea leaves, deionized water and amino acid surfactant is 15-25:300-400:0.5-1. In some embodiments of the present invention, for example, 15:300:0.5, 15:400:1, 20:300:0.5, 20:300:1, 25:300:0.5, and 25:400:1 can be selected, but the values ​​are not limited to the listed values. Other values ​​not listed within the numerical range are also applicable.

[0037] Among them, the amino acid surfactant can be selected from proline or glycine.

[0038] In this step, the ultrasonic power is 300-600W, for example, 300W, 350W, 400W, 450W, 500W, 550W, and 600W can be selected; the extraction temperature is 70-80°C, for example, 70°C, 72°C, 74°C, 75°C, 76°C, 78°C, and 80°C can be selected; the extraction time is 10-20min, for example, 10min, 12min, 14min, 15min, 16min, 18min, and 20min can be selected; but the values ​​are not limited to the listed values, and other values ​​not listed within the numerical range are also applicable.

[0039] In this step, green tea leaves are used as the main raw material and are subjected to water extraction. Amino acid surfactants are added during the water extraction process, which can effectively improve the permeability of the cell walls and effectively dissolve the flavonoids and polyphenols in the cells. At the same time, ultrasound-assisted treatment is used. Ultrasonic waves can generate tiny bubbles and rupture the cell walls through the cavitation effect of ultrasound, thereby more effectively releasing the active ingredients in the green tea leaves, shortening the extraction time, and improving the extraction rate of the active ingredients in the green tea leaves.

[0040] S2. Preparation of green tea fermentation liquid

[0041] The extract is added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus for fermentation. After the fermentation is completed, the filtrate is filtered and collected to obtain green tea fermentation liquid.

[0042] In this step, the amount of the extract added is 20-30% (v / v) based on the volume of sterilized coconut water, and the inoculation amount of Lactobacillus acidophilus is 1-3% (v / v), wherein the number of viable Lactobacillus acidophilus is ≥1.0×10 11 cfu / mL.

[0043] In this step, the fermentation temperature is 28-32°C, for example, 28°C, 30°C, and 32°C can be selected; the fermentation time is 48-72h, for example, 48h, 54h, 60h, 66h, and 72h can be selected; the stirring speed is 100-200r / min, for example, 100r / min, 150r / min, and 200r / min can be selected; but it is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable.

[0044] In this step, the extract is fermented by Lactobacillus acidophilus, and coconut water is used as a fermentation matrix. Coconut water contains a large amount of nutrients, which reduces the utilization of effective substances in the extract by Lactobacillus acidophilus, so that the green tea fermented liquid contains more antioxidant active ingredients. In addition, during the fermentation process, the bound polyphenols in the extract can release small molecular polyphenols, thereby increasing the content of flavonoids, polyphenols and other substances in the green tea fermented liquid, thereby further improving the antioxidant activity. At the same time, Lactobacillus acidophilus can ferment to produce some biologically active peptide substances, which work together with prebiotics such as inulin and sodium alginate to promote the growth and reproduction of beneficial bacteria such as lactic acid bacteria, bifidobacteria and acetic acid bacteria in the intestine. Lactobacillus acidophilus can also be used as a probiotic species to regulate gastrointestinal health.

[0045] S3. Preparation of modified dietary fiber

[0046] The filter residue is added into a hydrogen peroxide solution for treatment, and after the treatment is completed, it is filtered, washed, dried, ground and sieved to obtain modified dietary fiber.

[0047] In this step, the mass ratio of the filter residue to the hydrogen peroxide solution is 10-20:100, for example, 10:100, 12:100, 15:100, 18:100, and 20:100 can be selected; wherein the mass fraction of the hydrogen peroxide solution is 10-20%, for example, 10%, 12%, 15%, 18%, and 20% can be selected; but the present invention is not limited to the listed values, and other values ​​not listed within the numerical range are also applicable.

[0048] In this step, the filter residue is surface treated by using hydrogen peroxide in order to increase the activity of the surface groups of the filter residue and increase the specific surface area of ​​the filter residue.

[0049] S4. Preparation of composite prebiotic-embedded probiotic microcapsules

[0050] Inulin, sodium alginate and modified dietary fiber are added to deionized water and stirred evenly. Green tea fermentation broth and Lactobacillus rhamnosus powder are then added and mixed evenly. Zinc lactate solution is then added and stirred continuously to obtain a colloidal solution. The solution is freeze-dried and then ground into powder to obtain composite prebiotic-encapsulated probiotic microcapsules.

[0051] In this step, the mass ratio of inulin, sodium alginate, modified dietary fiber, deionized water, green tea fermentation broth, Lactobacillus rhamnosus powder and zinc lactate solution is 0.4-0.8:1-1.5:2-3:10-20:10-15:0.3-0.6:10-20, wherein the mass fraction of the zinc lactate solution is 3-6%, for example, 3%, 4%, 5% and 6% can be selected.

[0052] In this step, zinc ions in the zinc lactate solution form a gel with hydroxyl groups in inulin, sodium alginate and modified dietary fiber through chelation, which encapsulates the green tea fermentation broth, thereby improving the stability of the active substances and Lactobacillus acidophilus in the green tea fermentation broth, enabling them to play a protective and sustained-release role during the simulated digestion process; at the same time, zinc also has certain antioxidant properties, which can protect intestinal cells from oxidative damage, and works together with the active ingredients in the green tea fermentation broth to further enhance the antioxidant effect of the composite prebiotic-encapsulated probiotic microcapsules.

[0053] The composite prebiotic-embedded probiotic microcapsules provided by the present invention form a protective barrier on the outer layer of the probiotics, helping the probiotics resist damage from digestive juices such as gastric acid and bile, ensuring that more live bacteria reach the intestines; at the same time, the prebiotics provide nutrition for the probiotics after fermentation in the intestines, further enhancing their colonization and reproduction capabilities. This synergistic effect not only improves the survival rate of the probiotics, but also more effectively regulates the balance of intestinal flora, improves digestive function, enhances immunity, and relieves problems such as intestinal inflammation.

[0054] The present invention is further described below by means of specific examples. The green tea leaves provided by the present invention are produced in Enshi City, Hubei Province; Lactobacillus acidophilus, CAS No. 308084-36-8, was purchased from Shanghai Ruiyingying Biotechnology Co., Ltd., with a viable count of ≥1.0×10 11 cfu / mL; Lactobacillus rhamnosus LGG bacterial powder was purchased from Shanghai Ruiyingying Biotechnology Co., Ltd., with a viable cell count of ≥1.0×10 10 cfu / g.

[0055] Example 1

[0056] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0057] S1. Grind 15 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 300 g of deionized water. Then, add 0.5 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 300 W, an extraction temperature of 80° C., and an extraction time of 10 min. After the extraction, filter to obtain an extract and a filter residue.

[0058] S2, the extract was added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus. The amount of the extract added was 20% by volume of the sterilized coconut water, and the inoculum size of Lactobacillus acidophilus was 1%. The fermentation was carried out at 30°C for 60h with a stirring speed of 150r / min. After the fermentation was completed, the filtrate was filtered and collected to obtain a green tea fermentation broth.

[0059] S3, 10g of the filter residue was added to 100g, 10wt% hydrogen peroxide solution and treated for 3h. After the treatment, it was filtered, washed, dried, and ground through a 400 sieve to obtain modified dietary fiber;

[0060] S4. Add 0.4 g of inulin, 1 g of sodium alginate and 2 g of modified dietary fiber to 10 g of deionized water and stir evenly. Then add 10 g of green tea fermentation broth and 0.5 g of Lactobacillus rhamnosus powder and mix evenly. Then add 10 g of 3 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0061] Example 2

[0062] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0063] S1. Grind 25 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 400 g of deionized water. Then, add 1 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 400 W, an extraction temperature of 70° C., and an extraction time of 20 min. After the extraction, filter to obtain an extract and a filter residue.

[0064] S2, the extract was added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus. The amount of the extract added was 30% by volume of the sterilized coconut water, and the inoculum size of Lactobacillus acidophilus was 3%. The fermentation was carried out at 30°C for 60h with a stirring speed of 150r / min. After the fermentation was completed, the filtrate was filtered and collected to obtain a green tea fermentation broth.

[0065] S3, adding 20g of the filter residue to 100g, 20wt% hydrogen peroxide solution and treating for 3h. After the treatment is completed, filtering, washing, drying, and grinding through a 400 sieve to obtain modified dietary fiber;

[0066] S4. Add 0.8 g of inulin, 1.5 g of sodium alginate and 3 g of modified dietary fiber to 20 g of deionized water and stir evenly. Then add 15 g of green tea fermentation broth and 0.6 g of Lactobacillus rhamnosus powder and mix evenly. Then add 10 g of 5 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0067] Example 3

[0068] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0069] S1. Grind 20 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 400 g of deionized water. Then, add 0.8 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 600 W, an extraction temperature of 70° C., and an extraction time of 15 min. After the extraction, filter to obtain an extract and a filter residue.

[0070] S2, the extract was added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus. The amount of the extract added was 25% by volume of the sterilized coconut water, and the inoculum size of Lactobacillus acidophilus was 2%. The fermentation was carried out at 30°C for 48h with a stirring speed of 150r / min. After the fermentation was completed, the filtrate was filtered and collected to obtain a green tea fermentation broth.

[0071] S3, 15g of the filter residue was added to 100g, 20wt% hydrogen peroxide solution and treated for 3h. After the treatment, it was filtered, washed, dried, and ground through a 400 sieve to obtain modified dietary fiber;

[0072] S4. Add 0.5 g of inulin, 1.2 g of sodium alginate and 2.5 g of modified dietary fiber to 15 g of deionized water and stir evenly. Then add 15 g of green tea fermentation broth and 0.3 g of Lactobacillus rhamnosus powder and mix evenly. Then add 20 g of 3 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0073] Comparative Example 1

[0074] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0075] S1. Grind 15 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 300 g of deionized water. Then, add 0.5 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 300 W, an extraction temperature of 80° C., and an extraction time of 10 min. After the extraction, filter to obtain an extract and a filter residue.

[0076] S2, 10g of the filter residue was added to 100g, 10wt% hydrogen peroxide solution and treated for 3h. After the treatment was completed, it was filtered, washed, dried, and ground through a 400 sieve to obtain modified dietary fiber;

[0077] S3. Add 0.4 g of inulin, 1 g of sodium alginate and 2 g of modified dietary fiber to 10 g of deionized water and stir evenly. Then add 10 g of the extract and 0.5 g of Lactobacillus rhamnosus powder and mix evenly. Then add 10 g of 3 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. The solution is freeze-dried and then ground into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0078] Compared with Example 1, in Comparative Example 1, the extract was not subjected to fermentation treatment.

[0079] Comparative Example 2

[0080] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0081] S1. Grind 15 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 300 g of deionized water. Then, add 0.5 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 300 W, an extraction temperature of 80° C., and an extraction time of 10 min. After the extraction, filter to obtain an extract and a filter residue.

[0082] S2, the extract was added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus. The amount of the extract added was 20% by volume of the sterilized coconut water, and the inoculum size of Lactobacillus acidophilus was 1%. The fermentation was carried out at 30°C for 60h with a stirring speed of 150r / min. After the fermentation was completed, the filtrate was filtered and collected to obtain a green tea fermentation broth.

[0083] S3. Add 0.4 g of inulin and 1 g of sodium alginate to 10 g of deionized water and stir evenly. Then add 10 g of green tea fermentation broth and 0.5 g of Lactobacillus rhamnosus powder and mix evenly. Then add 10 g of 3 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-embedded probiotic microcapsules.

[0084] Compared with Example 1, in Comparative Example 2, no filter residue was added to the capsule wall material.

[0085] Comparative Example 3

[0086] A method for preparing composite prebiotic-embedded probiotic microcapsules comprises the following steps:

[0087] S1. Grind 15 g of green tea leaves, pass them through an 80-mesh sieve, and add them to 300 g of deionized water. Then, add 0.5 g of glycine and perform ultrasonic-assisted extraction at an ultrasonic power of 300 W, an extraction temperature of 80° C., and an extraction time of 10 min. After the extraction, filter to obtain an extract and a filter residue.

[0088] S2, the extract was added to sterilized coconut water, and then inoculated with Lactobacillus acidophilus. The amount of the extract added was 20% by volume of the sterilized coconut water, and the inoculum size of Lactobacillus acidophilus was 1%. The fermentation was carried out at 30°C for 60h with a stirring speed of 150r / min. After the fermentation was completed, the filtrate was filtered and collected to obtain a green tea fermentation broth.

[0089] S3. Add 0.4 g of inulin, 1 g of sodium alginate and 2 g of filter residue (400 mesh) to 10 g of deionized water and stir evenly. Then add 10 g of green tea fermentation broth and 0.5 g of Lactobacillus rhamnosus powder and mix evenly. Then add 10 g of 3 wt % zinc lactate solution and continue stirring for 2 h to obtain a colloidal solution. After freeze-drying, grind into powder to obtain composite prebiotic-encapsulated probiotic microcapsules.

[0090] Compared with Example 1, in Comparative Example 3, no modification treatment was performed on the filter residue.

[0091] The composite prebiotic-embedded probiotic microcapsules prepared in Example 1 and Comparative Examples 1-3 were subjected to an in vitro DPPH free radical scavenging test, and the specific steps were as follows:

[0092] 40% ethanol-physiological saline release media with different pH values ​​(3, 7.4, and 8.5) were prepared respectively, and 5 mL was measured and placed in a 10 mL centrifuge tube. Then, 10 mg of the composite prebiotic-encapsulated probiotic microcapsules prepared in Example 1 and Comparative Examples 1-3 were added, respectively. The mixture was shaken in a constant temperature shaker (180 r / min) at 37°C in the dark for 3 h, and then centrifuged at 2000 × g for 5 min. The supernatant obtained was the released active substance solution. 100 μL of the supernatant was added to 300 μL of DPPH ethanol solution (0.04 mg / mL), shaken well, incubated in the dark for 30 min, and centrifuged at 4000 r / min for 10 min. The supernatant was taken and its absorbance was measured at 517 nm.

[0093] The DPPH free radical scavenging rate was calculated according to the formula:

[0094] DPPH free radical scavenging rate / %=[1-(A-A0) / A1]×100;

[0095] Wherein, A refers to the absorbance value after adding the active substance solution, A0 refers to the background absorbance value of the active substance solution, and A1 refers to the absorbance value of the blank control group.

[0096] All experiments were repeated three times independently, and the test results are shown in Figure 2. Figure 1 As shown, from Figure 1It can be seen that the scavenging rate of the composite prebiotic-embedded probiotic microcapsules prepared in Example 1 and Comparative Examples 1-3 for DPPH free radicals gradually increases with increasing pH. The reason is that with increasing pH, the amount of active substances released from the composite prebiotic-embedded probiotic microcapsules gradually increases, and therefore the scavenging rate of DPPH free radicals gradually increases. From the data in Example 1, it can be seen that at a pH of 3, the scavenging rate of the composite prebiotic-embedded probiotic microcapsules for DPPH free radicals is about 43%, while when the pH is 8.5, the scavenging rate of DPPH free radicals can reach 87%. The reason may be that when the pH is 8.5, the hydrogen bonding effect between the various substances of the capsule wall material is weakened, the structure becomes loose, and the embedded active substances are released in large quantities. This shows that the release rate of the active substances in the composite prebiotic-embedded probiotic microcapsules prepared by the present invention is low in the gastric stage, but can be released in large quantities in the intestinal stage, and can achieve intestinal sustained-release function. In Comparative Example 1, the extract was not fermented, and its DPPH radical scavenging rate was lower than that in Example 1, indicating that fermentation of the extract can increase the content of antioxidant active ingredients. In Comparative Example 2, no filter residue was added to the capsule wall material, and in Comparative Example 3, the filter residue was not modified. The DPPH radical scavenging rates of the composite prebiotic-encapsulated probiotic microcapsules prepared in Comparative Examples 2 and 3 were both higher than that in Example 1. This may be because by adding modified dietary fiber to the capsule wall material, the active substances in the green tea fermentation broth are embedded in the lamellar structure and pores of the modified dietary fiber, thereby enhancing the encapsulation effect of the active substances and delaying the release of the active substances, thereby achieving the purpose of sustained release.

[0097] The storage stability test of the composite prebiotic-embedded probiotic microcapsules prepared in Example 1 and Comparative Examples 2-3 was performed. The specific experimental steps were as follows: the composite prebiotic-embedded probiotic microcapsules were stored at room temperature for 4 weeks, and the viable bacteria counts at the beginning of storage (recorded as week 0) and after 4 weeks of storage were tested. When measuring the viable bacteria count, 0.1 g of microcapsules were placed in 10 mL of sterile PBS each time, and the cells were shaken at a constant temperature of 37°C until completely dissolved. The cells were then plated and counted using the dilution gradient method. All experiments were repeated three times independently. The test results are shown in the figure. Figure 2 As shown, in Comparative Example 2, no filter residue was added to the capsule wall material, and the number of viable bacteria decreased by about 2.8 (1 g (CFU / g)), in Comparative Example 3, no filter residue was modified, and the number of viable bacteria decreased by about 1.5 (1 g (CFU / g)), while the number of viable bacteria in Example 1 of the present invention decreased by about 0.7 (1 g (CFU / g)). It can be seen that the composite prebiotic-embedded probiotic microcapsules prepared by the present invention can maintain good stability when stored at room temperature.

[0098] Polyphenol extraction rate test: The extract obtained in step S1 of Example 1-3 was lyophilized to obtain a lyophilized powder, which was then prepared into a 2 mg / ml test solution. A control group was also set up. The experimental steps of the control group were as follows: 15 g of green tea leaves were crushed, passed through an 80-mesh sieve, and added to 300 g of deionized water. Ultrasonic-assisted extraction was performed at an ultrasonic power of 300 W, an extraction temperature of 80° C., and an extraction time of 10 min. After the extraction, the extract and filter residue were filtered. The extract was lyophilized to obtain a lyophilized powder, which was then prepared into a 2 mg / ml test solution for later use.

[0099] 1 mL of each test solution from Examples 1-3 and the control group was placed in a test tube. 5.00 mL of 10% Folin phenol reagent was added to the test tube. The mixture was allowed to stand for 5 minutes, and then 4 mL of 7.5% sodium carbonate solution was added. The mixture was shaken and allowed to react at room temperature for 60 minutes. The absorbance at 765 nm was measured using a UV-visible spectrophotometer. The absorbance was compared with the absorbance at 765 nm of a 2 mg / mL aqueous solution of a gallic acid reference substance treated in the above steps. The polyphenol extraction rate was calculated as follows: polyphenol extraction rate (%) = mass of polyphenols in the freeze-dried powder / mass of green tea leaves × 100%. The test was repeated three times, and the results were averaged.

[0100] Flavonoid extraction rate test: 1 mL of the extract obtained in Examples 1-3 and the control group was respectively placed in a 10 mL volumetric flask, 0.3 mL of 5% NaNO2 solution was added, and the mixture was shaken and allowed to stand for 6 minutes. Then, 0.3 mL of 10% aluminum nitrate solution was added, and the mixture was shaken and allowed to stand for 6 minutes. 4.0 mL of 4% NaOH solution was added, and the volume was made up to the mark with 75% ethanol. The mixture was shaken and allowed to stand for 12 minutes. The absorbance was measured at 510 nm, and the mass of flavonoids in the extract was calculated. Flavonoid extraction rate (%) = mass of flavonoids in the extract / mass of green tea leaves × 100%. The test was repeated three times, and the results were averaged.

[0101] The test results are as follows Figure 3 As shown, from Figure 3 It can be seen that the control group did not add glycine during the extraction process. Compared with the examples of the present invention, the polyphenol extraction rate and flavonoid extraction rate were significantly reduced. It can be seen that the addition of amino acid surfactants during the water extraction process can effectively dissolve flavonoids and polyphenols in cells.

[0102] Finally, it should be noted that the above embodiments do not limit the present invention in any form. Those skilled in the art will appreciate that modifications and improvements can be made based on the present invention. Therefore, any modifications or improvements made without departing from the spirit of the present invention are intended to fall within the scope of protection claimed in the present invention.

Claims

1. A method for preparing composite prebiotic-embedded probiotic microcapsules, characterized in that: The steps include: S1. Grind and sieve green tea leaves, add them to deionized water, then add glycine, perform ultrasonic-assisted extraction, and filter after the extraction to obtain an extract and a filter residue; S2, adding the extract to sterilized coconut water, then inoculating Lactobacillus acidophilus, fermenting and culturing, and after the fermentation is completed, filtering and collecting the filtrate to obtain green tea fermentation broth; S3, adding the filter residue to a hydrogen peroxide solution for treatment, and after the treatment is completed, filtering, washing, drying, grinding and screening to obtain modified dietary fiber; S4, adding inulin, sodium alginate and modified dietary fiber to deionized water, stirring evenly, then adding green tea fermentation broth and Lactobacillus rhamnosus powder thereto, mixing evenly, then adding zinc lactate solution, continuing stirring to obtain a colloidal solution, and then freeze-drying and grinding into powder to obtain composite prebiotic-embedded probiotic microcapsules; In step S2, during the fermentation culture process, the fermentation temperature is 28-32°C, the fermentation time is 48-72 hours, and the stirring speed is 100-200 r / min.

2. The preparation method according to claim 1, characterized in that In step S1, the mass ratio of green tea leaves, deionized water and glycine is 15-25:300-400:0.5-1.

3. The preparation method according to claim 1, characterized in that In step S1, the ultrasonic power is 300-600w, the extraction temperature is 70-80°C, and the extraction time is 10-20min.

4. The preparation method according to claim 1, characterized in that In step S2, based on the volume of the sterilized coconut water, the amount of the extract added is 20-30%, and the inoculum amount of Lactobacillus acidophilus is 1-3%.

5. The preparation method according to claim 1, characterized in that In step S3, the mass ratio of the filter residue to the hydrogen peroxide solution is 10-20:100, and the mass fraction of the hydrogen peroxide solution is 10-20%.

6. The preparation method according to claim 1, characterized in that In step S4, the mass ratio of inulin, sodium alginate, modified dietary fiber, deionized water, green tea fermentation broth, Lactobacillus rhamnosus powder and zinc lactate solution is 0.4-0.8:1-1.5:2-3:10-20:10-15:0.3-0.6:10-20.

7. The preparation method according to claim 1, characterized in that In step S4, the mass fraction of the zinc lactate solution is 3-6%.

8. Composite prebiotic-embedded probiotic microcapsules prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the composite prebiotic-embedded probiotic microcapsules according to claim 8 in the preparation of solid beverage products.

Images