WO2019200499A1 - Probiotic microcapsule for maintaining strain activity, and preparation method thereof - Google Patents

Abstract

Provided are a probiotic microcapsule for maintaining strain activity and preparation method thereof; the probiotic microcapsule uses a composite wall structure constituted by a first wall material and a second wall material; the first wall material is a protein plant polysaccharide composite gel made of casein, malt extract, and xylooligosaccharide which can prevent damage to a core material caused by light, heat, oxygen, metal ions, and the like, and positively promote the stability of an embedded material; the second wall material is a plant polysaccharide compound made of Tremella fuciformis polysaccharide, Bletilla striata polysaccharide, Atractylodes macrocephala polysaccharide, guar gum, and pectin which can reduce the impact of vacuum freeze-drying processing technology on the biological activity of core probiotic bacteria; at the same time, when the outside of the first wall material is covered with the second wall material, the first wall material, which contains a large amount of protein, increases the film-forming effectiveness and embedding effect of the second wall material, thereby improving the performance of the composite wall material as a whole such that the composite wall material better protects the core material from biological activity and improves the stability of the core material.

Description

Probiotic microcapsule for maintaining strain activity and preparation method thereof Technical field

The invention relates to the field of biotechnology, in particular to a probiotic microcapsule for maintaining the activity of a strain and a preparation method thereof.

Background technique

The intestine is the largest digestive organ and detoxification organ in the human body. The state of the intestine determines the appearance and beauty of the human body. The digestive tract of the human body is as large as a tennis court. The small intestine is about 4-6 meters long and the large intestine is about 1.5 meters long. The intestine is also the largest immune organ of the human body. It has the largest micro-ecological environment of the human body and produces about 80% of the resistance. The normal flora is about 1.5kg, the number is 100 trillion, and the flora is 500-1000. The human intestine is actually an organic component of the symbiosis between the self organ and the resident microbial group. Among them, the beneficial bacteria and harmful bacteria in the intestine maintain a dynamic balance, which can help digest food, produce immune components, and enhance the body's own immunity. Force, promote the synthesis of certain vitamins, improve the absorption of lactose, protect the human body from pathogens, promote bowel movements, and inhibit intestinal corruption. The intestinal barrier is an important biological barrier of the human body. The imbalance of the intestinal flora causes the intestinal barrier to be destroyed, and harmful bacteria or endotoxins enter the body, leading to inflammation and various discomforts. Modern medical evidence tells us that "aging begins with the intestines" and "important adult diseases such as bowel cancer, breast cancer, heart disease, high blood pressure, and senile dementia are closely related to intestinal health."

With the development of society, people's lifestyle and diet structure are undergoing tremendous changes. Long-term use of computers, mobile phones and other electronic devices, work pressure, staying up late to work overtime, rarely participating in sports or exercise, often eating raw and cold Irritating foods such as spicy foods, irregular eating and drinking, or excessive drinking, can lead to insufficient blood supply to the gastrointestinal tract, decreased immunity, and reduced beneficial intestinal flora, resulting in a sub-health state.

Probiotics are biologically active beneficial microorganisms that, when taken in sufficient amounts, improve the micro-ecological balance of the host's gut and thus have a beneficial effect on the health of the host. In recent years, the probiotics industry has developed rapidly and is now used in many fields such as medicine, food, and health care products. In the field of medicine, probiotics are mainly used to treat diseases such as diarrhea, constipation and vaginitis; in the food industry, probiotics are widely used in fermented flavor foods, such as traditional yogurt, lactic acid bacteria beverages, kimchi, vinegar, liquor, etc.; In the health food industry, probiotics are made into tablets, capsules, granules, powders, etc. in order to achieve a variety of health functions.

For probiotics to exert their health effects and produce probiotic functions, they must maintain sufficient biological activity when ingested by the human body. The number of probiotics ingested must be 106 cfu/g, and the official standard proposed by the World Food Organization is 106. ~107 cfu/g. At present, liquid probiotic live bacteria can be stored for 3 to 14 days below 4 ° C. The method of using traditional process skim milk powder to embed probiotics has a shelf life of 42 days at 4 ° C, which is difficult to improve significantly. The most widely used probiotics are Lactobacilli (such as Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus bulgaricus, etc.), Streptococcus (such as Streptococcus thermophilus, etc.) and Bifidobacteria (such as bifidus). Bacillus, Bifidobacterium infantis, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium adolescentis, etc., which are extremely susceptible to the influence of the external environment during storage, how to maintain the activity of the strain and prolong the shelf life, become this Recognized technical problems in products.

Summary of the invention

The technical problem to be solved by the present invention is to provide a probiotic microcapsule which can reduce the adverse effects of environmental factors on the activity of probiotics, and maintain the activity of the strain to improve the quality of probiotic products, in view of the current state of the art.

Another technical problem to be solved by the present invention is to provide a method for preparing the above probiotic microcapsules according to the state of the art, which can reduce the adverse effects of environmental factors on the activity of probiotics and maintain the activity of the strain for a long time. Probiotic product quality.

The technical solution adopted by the present invention to solve the above technical problems is: a probiotic microcapsule that maintains the activity of a strain, comprising a core material and a wall material coated on the outside of the core material, wherein the core material is a probiotic bacteria, and is characterized in that The wall material includes a first wall material coated on the outside of the core material and a second wall material wrapped on the outside of the first wall material.

The first wall material comprises the following components: casein, malt extract and xylooligosaccharide;

The second wall material comprises the following components: Tremella polysaccharide, white and polysaccharide, atractylodes polysaccharide, guar gum and pectin.

The first and second wall materials are each diluted with a mixture of 3 to 5 times the mass of the wall material as a solvent to form a mixed solution, and then embedded.

In the above aspect, the mass ratio of the probiotic to the first wall material is 1: (0.5 to 2), and the casein is 5 to 15 parts by weight, and the malt extract is 1 by weight. ~6 parts, oligosaccharide sugar is 0.5 to 3 parts. The mass ratio of the probiotic to the second wall material is 1: (0.5-1.5), and the second wall material is 2-8 parts of the white fungus polysaccharide, and the white and polysaccharide is 2-5 parts, atractylodes The polysaccharide is 3-6 parts, the guar gum is 0.2-0.6 parts, and the pectin is 0.6-1.2 parts. The invention optimizes and adjusts the mass ratio of the probiotic bacteria to the first wall material and the second wall material and the ratio of the raw materials in the wall materials, so that the prepared microcapsules have good biological activity of the strain, so that the wall material It can completely envelop probiotics without causing the low number or waste of live bacteria. At the same time, it improves the stability of microcapsules in freeze-drying, high-temperature storage, production and processing, improves the survival rate of the strains, and improves the strains. Activity, reducing waste and the production of defective products. The probiotic microcapsules prepared by the above preparation method have a particle size of 75-150 μm, uniform particle size distribution and controllable quality, and the microcapsule embedding rate is greater than 96%.

Preferably, the probiotic is one or more of Lactobacillus, Bifidobacterium, Streptococcus, wherein the Lactobacillus may be a strain of Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus bulgaricus, Streptococcus For strains such as Streptococcus thermophilus, the bifidobacteria may be Bifidobacterium bifidum, Bifidobacterium infantis, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium adolescentis and the like.

A method for preparing a probiotic microcapsule for maintaining the activity of the above-mentioned bacteria, characterized in that the method comprises the following steps

(1) Centrifugally treating the probiotic culture medium in the activated proliferation culture, and removing the supernatant after the treatment to obtain the probiotic bacteria;

(2) adding casein, malt extract and xylooligosaccharide to water to obtain a mixed solution of the first wall material, adding the probiotic bacteria to the mixed solution of the first wall material, and stirring to obtain a bacterial suspension The bacterial suspension is slowly sprayed into a gallic acid solution having a pH of 3.0 to 5.0 by a sprayer, allowed to stand for 5 to 20 minutes, and a small particle precipitate is collected, and the collected small particles are washed in a sterile water having a pH of 3.8 to 4.2;

(3) adding Tremella polysaccharide, white and polysaccharide, Atractylodes polysaccharide, guar gum and pectin to water to obtain a mixed solution of the second wall material, and the small particles obtained in the step (2) are put into the mixed solution of the second wall material. The bacterial suspension is prepared by stirring, and the bacterial suspension is slowly sprayed into a 0.05-0.5 mol/L calcium lactate solution by a sprayer to be solidified, washed, and filtered to obtain microcapsules;

(4) The above microcapsules are obtained by vacuum freeze-drying to obtain a probiotic microcapsule dry powder.

The medium used for the probiotic culture solution is MRS liquid medium, the culture temperature is 36-40 ° C, the culture time is 24-36 h, the centrifugation temperature is 0-10 ° C, the centrifugal speed is 4000-6000 r/min, and the centrifugation time is 5 to 12 minutes.

The solvent used in the above mixed solution and washing water is sterile water, and the above MRS medium, wall material mixed solution, gallic acid solution and calcium lactate solution are subjected to high temperature moist heat sterilization, and the sterilization temperature is 115 to 125 ° C, sterilization. The time is 15 to 30 minutes.

The probiotic microcapsules prepared by the above preparation method have a particle size of 75-150 μm, uniform particle size distribution and controllable quality, and the microcapsule embedding rate is greater than 92%, so that the obtained microcapsules have good strain activity. It can adapt to changes in light, heat, oxygen, metal ions, pH and other environmental factors, and has good stability.

Preferably, the centrifugation speed of the centrifugation in the step (1) is 4000 to 6000 r/min, and the centrifugation time is 5 to 12 min.

Preferably, the pre-freezing initial temperature of the vacuum freeze-drying in the step (4) is -30 to -40 ° C, the pre-freezing rate is 0.4-0.8 ° C/min, the pre-freezing end temperature is -60 ° C, and the drying chamber pressure is 40-100 Pa. The heating plate temperature is 30 to 40 ° C, and the drying time is 12 to 20 hours.

Preferably, the malt extract is prepared as follows

(a) pulverization: pulverizing barley malt or barley into ultrafine barley malt powder or barley flour of 90-110 μm by ultrafine pulverization technology;

(b) pulping; the barley malt powder or barley flour is prepared into a protein dispersion with pure water, and the pH of the dispersion is adjusted to between 6 and 7;

(c) Enzymatic hydrolysis: heating and maintaining the temperature of the dispersion between 40 and 50 ° C, then adding α-amylase, β-amylase and neutral protease to the dispersion, stirring uniformly and then enzymolysis for 2 h~ 4h, wherein the amount of α-amylase added is 0.3% to 0.5% of the substrate content, the amount of β-amylase added is 0.1% to 0.3% of the substrate content, and the amount of neutral protease added is 0.2% of the substrate content. ~0.4%, after the end of enzymatic hydrolysis, the enzyme is subjected to high temperature;

(d) Centrifugal filtration: Centrifugation of the enzymatic hydrolysate after inactivation of the enzyme, maintaining a centrifugal speed of 12000 to 16000 r/min, feeding speed of the peristaltic pump of 1.5 to 3.5 L/min, collecting the supernatant, and starting the membrane filtration device. The supernatant is filtered through a microporous membrane, the membrane thickness is 90-150 μm, the filtered particle size is 4-8 μm, and the operating pressure is 0.01-0.2 MPa, ensuring that the filtrate is clear and transparent, and the filter residue is removed;

(e) vacuum concentration: the filtrate is concentrated in a vacuum, concentrated at 65-85 ° C, vacuum -0.07 ~ -0.09Mpa, to obtain a concentrated liquid, the relative density is controlled between 1.04 ~ 1.08;

(f) Spray drying: The above concentrated liquid was spray-dried, and the process parameters were a nozzle temperature of 175 to 190 ° C, an outlet temperature of 80 to 95 ° C, and a feeding speed of 1.5 to 3.5 L/min to obtain the malt extract.

The above-mentioned malt extract is selected from high-quality barley malt/barley as raw material, which is pulverized, pulped and enzymatically decomposed into wort, and then subjected to centrifugal filtration, vacuum concentration, spray drying and other production processes to obtain high-quality malt extract. The malt extract has an enzymatic hydrolysis rate of more than 90%, and the energy efficiency improves the absorption and utilization rate of the above-mentioned extracts, and the effect can be absorbed without digesting or slightly digesting.

Preferably, the white and polysaccharide/Atractylodes polysaccharide are obtained by the following steps

(a) Dynamic countercurrent extraction; take pulverized white and / atractylodes, add 8-10 times the weight of the material, and dynamically countercurrently extract at 90 ~ 100 ° C to obtain the extract, the number of extraction times is 1 to 3 times, each time The extraction time is 1 to 3 hours;

(b) centrifugation: the extract is centrifuged to obtain a clear liquid;

(c) Concentrated alcohol precipitation: The clear liquid is concentrated in a vacuum, the relative density is controlled between 1.10 and 1.20, and the edible liquid is added to the concentrated liquid to make the alcohol content in the mixed solution reach 40-80%, fully stirred and allowed to stand. Overnight; the supernatant recovers the solvent and leaves the precipitate for use;

(d) ion exchange: the alcohol precipitation polysaccharide is dissolved in 4-8 times purified water, stirred well, dissolved, and filtered, and the filtered filtrate is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(e) vacuum concentration under vacuum: the ion exchanged solution is concentrated in a vacuum, concentrated temperature 60-80 ° C, vacuum -0.07 ~ -0.09Mpa, to obtain a polysaccharide concentrate, the relative density is controlled between 1.03 ~ 1.06;

(f) spray drying: the polysaccharide concentrate is sterilized and filtered, and then spray-dried, the process parameters are nozzle temperature 170-185 ° C, outlet temperature 75-95 ° C, feeding speed 2.0-4.0 L / min, to obtain the white And polysaccharide / Atractylodes polysaccharide.

The polysaccharide extraction rates of white and polysaccharide/Atractylodes polysaccharides were 35% and 8%, respectively. White and polysaccharide/Atractylodes polysaccharides are prepared by pulverizing, dynamic countercurrent extraction, centrifugation, concentrated alcohol precipitation, ion exchange, vacuum concentration and spray drying. Dynamic countercurrent extraction can make full use of the concentration gradient of solid-liquid two-phase, and diffuse the active ingredient to the stripping solution with lower initial concentration step by step, so that the extract of the discharged material reaches a higher equilibrium concentration, which can ensure the extraction. The extraction rate of the material can save energy, shorten the extraction time, and greatly reduce the workload and energy consumption of the subsequent concentration process, and comprehensively solve the problems existing in the intermittent leaching of the single can currently widely used.

Preferably, the tremella polysaccharide is obtained by the following steps

(a) Enzymatic hydrolysis; the white fungus is pulverized into ultrafine powder of 90-110 μm by ultrafine pulverization technology, 30~50 times of water and 0.5~2% pectinase are added, and the temperature is 40-50 °C. 30-90min, warming up to 95 ° C or more to kill the enzyme:

(b) Extraction: immersion in 95-100 ° C temperature, 500-mesh sieve to obtain Tremella polysaccharide extract, the number of extraction times is 1-2 times, each extraction time is 1-3 hours;

(c) ion exchange: the filtered Tremella polysaccharide extract is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(d) Vacuum concentration: the ion exchanged solution is concentrated in a vacuum, the concentration temperature is 60-80 ° C, the degree of vacuum is -0.07 - -0.09 Mpa, and the tremella polysaccharide concentrate is obtained, and the relative density is controlled between 1.02 and 1.04;

(e) Vacuum freeze-drying: Tremella polysaccharide powder is prepared by vacuum freeze-drying technique, wherein the initial temperature of pre-freezing is -35 to -45 ° C, the pre-freezing rate is 0.2-0.5 ° C / min, and the pre-freezing end temperature - 65 ° C, drying chamber pressure 50 ~ 120pa, heating plate temperature 30 ~ 40 ° C, drying time is 16 ~ 24h.

The above-mentioned Tremella polysaccharides are prepared by a production process such as pulverization, enzymatic hydrolysis, extraction, ion exchange, vacuum concentration, and vacuum freeze-drying. The polysaccharide extraction rate of Tremella polysaccharides is as high as 36%.

Compared with the prior art, the invention has the advantages that the probiotic microcapsules for maintaining the activity of the invention adopt the composite wall structure composed of the first wall material and the second wall material, and the first wall material is composed of casein. The protein plant polysaccharide composite gel made of malt extract and xylooligosaccharide can well prevent the damage of the core material such as light, heat, oxygen and metal ions, and positively affect the stability of the embedded material. Promoting effect; the second wall material is a plant polysaccharide complex made of Tremella polysaccharide, white and polysaccharide, Atractylodes polysaccharide, guar gum and pectin, which can greatly reduce the vacuum freeze-drying process to the core probiotic bacteria The effect of biological activity; meanwhile, when the first wall material is coated with the second wall material, the first wall material containing a large amount of protein can increase the film forming property and embedding effect of the second wall material, thereby improving the whole The performance of the composite wall material enables the composite wall material to better protect the core material activity and significantly improve the stability of the core material, so that the prepared microcapsule has strong acid resistance and can withstand the test of stomach acid in the intestine. Fixed point release To improve the bioavailability of probiotics.

The preparation method of the invention adopts dynamic countercurrent extraction, vacuum concentration, enzymolysis separation and purification, ion exchange, microcapsule embedding, vacuum freeze-drying to prepare probiotic microcapsules, and the natural raw materials together with the probiotics are made into microcapsules, Maximize the biological activity of probiotics, improve the performance of probiotics, and extend shelf life. Traditional probiotic microencapsulation methods mostly use gum arabic, polyamide, polyvinyl alcohol, polyethylene glycol, sodium alginate and sodium carboxymethyl cellulose as capsule materials. These embedding agents have strong mechanical strength and mass transfer. The performance is poor, and some of the raw materials have certain toxicity, and there is a certain food safety risk, and the present invention uses natural raw materials to improve the storage conditions of the probiotics, thereby achieving the purpose of maintaining the activity of the strain and improving the function of the gastrointestinal tract.

Specifically, white and dry tubers are rich in sticky polysaccharides, which are bitter, sweet, glutinous, slightly cold, and belong to the lungs, liver and stomach. The effect is astringent hemostasis, swelling and muscle growth, for internal and external bleeding, sore swollen, skin chapped. It has a good therapeutic effect on gastrointestinal damage and gastric ulcer, and can protect and repair the gastric mucosa. Atractylodes macrocephala is the dry rhizome of plants, rich in polysaccharides, bitter, sweet, warm, spleen, stomach. The effect is to spleen and qi, dampness and water, antiperspirant and fetus. For spleen deficiency, less bloating, diarrhea, sputum, edema, edema, spontaneous sweating, fetal movement, spleen and stomach. Tremella is a fruiting body of the fungus Tremella fuciformis, rich in polysaccharides, sweet, light, and non-toxic. It not only has the effect of strengthening the kidney, invigorating the stomach, but also benefiting the blood and blood. It can nourish the lungs, improve the body's immunity, and enhance the tolerance of cancer patients to radiotherapy and chemotherapy. Casein is a phosphorus-calcium-binding protein widely found in mammals including cows, sheep and human milk. Casein is both a source of amino acids and a source of calcium and phosphorus. Casein forms a curd in the stomach. digestion. Casein can treat dental caries, prevent and treat osteoporosis and rickets, regulate blood pressure, treat iron deficiency anemia, magnesium deficiency neuritis and other physiological effects, and promote the efficient absorption of minerals and trace elements. Malt extract is a natural food derived from selected high-quality barley malt (barley), which is completely derived from cereals. It contains vitamins, minerals and soluble dietary fiber. It is rich in glucose, maltose, oligosaccharides, small molecular peptides and more. It is an essential nutrient for human essential and non-essential amino acids and contains active polysaccharide β-glucan. Xylooligosaccharide is a prebiotic, a functional polymeric sugar composed of 2 to 7 xylose molecules combined with β-1,4 glycosidic bonds. It is difficult for xylooligosaccharides to be decomposed by human digestive enzymes and has reduced toxicity. Fermentation products and harmful bacterial enzymes, inhibit pathogens and diarrhea, protect the liver, lower serum cholesterol, lower blood pressure, enhance immunity, stimulate intestinal peristalsis to prevent constipation, promote the reproduction of beneficial bacteria in the digestive tract of animals, and improve the balance of microbial flora The effect. Guar gum is a highly purified natural polysaccharide extracted from the widely grown leguminous guar. Guar gum is a macromolecular natural hydrophilic colloid, belonging to natural galactomannan, a natural thickener. Pectin is widely found in the fruits and roots of plants and is a component of the cell wall. When the pectin is dissolved in water, it forms a milky white viscous colloidal solution, which is weakly acidic and has strong heat resistance, and can form a gel with elasticity.

The invention adopts traditional Chinese medicines such as white fungus, white and atractylodes, such as nourishing yin and moistening the lungs, moistening the intestines and tonifying the stomach, and combining with casein, malt extract, xylooligosaccharide, pectin, guar gum and other natural sources of raw materials, can form a good The microencapsulation of microcapsules greatly enhances the activity of probiotics, prolongs the shelf life, increases the ability of probiotics to resist stomach acid, and can smoothly transfer to the intestines and release them. While improving the balance of intestinal stomach and ecology, The protein and active polysaccharide in the wall material can provide sufficient nutrients for the probiotics, and can also play the role of moistening the intestines and benefiting the stomach.

The invention subverts the traditional single-layer embedding process of probiotic microcapsules, and adopts a double-layer microcapsule embedding process of natural raw materials. First, protein, xylooligosaccharide, malt extract, etc. are wrapped on the probiotic bacteria to form stable condensed tuberculosis, and then natural plant polysaccharides and natural polymer compounds are used to continue embedding on the above condensed nodules. The curing process forms a stable microcapsule shell outside the probiotic. The probiotic microcapsules prepared by the above double-layer microcapsule embedding process have extremely stable biological activity, can resist the corrosion of gastric acid, and dissolve the capsule shell fully and rapidly after reaching the pH value of the intestinal tract. A large number of probiotics are released, which have a release rate of more than 86% in intestinal fluid and provide sufficient nutrition for the intestines. The microencapsulated probiotics have stable biological activity and durability, and have superior performance in high temperature resistance, freezing resistance, light resistance and high oxygen activity. Compared with the non-microencapsulated probiotics, the shelf life is prolonged. 2.5 times or more.

The capsule material of the invention adopts white and polysaccharide, tremella polysaccharide, atractylodes polysaccharide and xylooligosaccharide as a polysaccharide composition to introduce a wall material system, wherein the white and viscous polysaccharide can converge to stop bleeding, reduce swelling and build muscle, protect and repair gastrointestinal mucosa, Tremella polysaccharides benefit Qi and blood, strong invigorous kidney, Runchangyiwei, Atractylodes polysaccharide can strengthen the spleen and stomach, this combination of Chinese herbal medicine extract and strong benefit of oligo-oligosaccharide, while maintaining the activity of probiotic microcapsules It can also protect the gastrointestinal tract, promote the rapid proliferation of probiotics, and greatly improve the actual effect and bioavailability of the above microcapsules.

The preparation method of the probiotic microcapsule of the invention has universal applicability, and is suitable for the production and processing of microcapsules of one or several probiotics of Lactobacillus, Bifidobacterium and Streptococcus, and the operation is simple and easy, and the process is simple and stable. The quality is controllable, the production efficiency is high, and the economic added value is high, which is suitable for large-scale industrialized production.

DRAWINGS

1 is a graph showing a change trend of the amount of probiotic bacteria in the yogurt during the shelf life in Example 6 of the present invention;

2 is a graph showing changes in the viable environment viability of probiotic bacteria and microcapsule powder of the present invention;

Fig. 3 is a graph showing the results of an accelerated test of the activity of probiotic bacteria and microcapsule powders of the present invention.

detailed description

The invention will be further described in detail below with reference to the embodiments of the drawings.

Example 1;

The probiotic microcapsules for maintaining the activity of the strain in the embodiment include a core material and a wall material coated on the core material, the core material is a probiotic bacteria, and the wall material comprises a first wall material coated on the outside of the core material and coated. a second wall material outside the first wall material, the first wall material comprises the following components: casein, malt extract and xylooligosaccharide; the second wall material comprises the following components; Tremella polysaccharide, white and polysaccharide, atractylodes The polysaccharide, guar gum and pectin, the probiotic bacteria are Lactobacillus acidophilus, and the first and second wall materials are diluted into a mixed solution by using 4 times sterile water as a solvent, and then embedded. The mass ratio of the Lactobacillus acidophilus cells to the first wall material was 1:1, the casein in the first wall material was 12 parts, the malt extract was 6 parts, and the xylooligosaccharide was 0.5 parts. The mass ratio of Lactobacillus acidophilus cells to the second wall material is 1:1.5, the amount of Tremella polysaccharides in the second wall material is 3 parts, the white and polysaccharides are 5 parts, the atractylodes polysaccharide is 3 parts, and the guar gum is 0.2 parts. The pectin is 1 part.

In the present embodiment, the malt extract is obtained by pulverizing, slurrying, enzymatic hydrolysis, centrifugal filtration, vacuum concentration, and spray drying, and the specific preparation steps are as follows:

(a) pulverization: pulverizing barley malt or barley into ultrafine powder of about 100 μm by ultrafine pulverization technology;

(b) pulping: the barley malt powder or barley flour is prepared into a protein dispersion with pure water, and the pH of the dispersion is adjusted to 6.5;

(c) Enzymatic hydrolysis: heating and maintaining the temperature at 45 ° C, then adding α-amylase, β-amylase and neutral protease respectively in the dispersion, stirring and homogenizing for 2 h, wherein α-amylase The amount added is 0.3% of the substrate content, the amount of β-amylase added is 0.3% of the substrate content, and the amount of neutral protease added is 0.2% of the substrate content, and the enzyme is subjected to high temperature inactivation after the end of enzymatic hydrolysis;

(d) Centrifugal filtration: the enzymatic hydrolysate after the enzyme is centrifuged to remove impurities, maintain the centrifugal speed of 14000r/min, the peristaltic pump feed rate is 2.5L/min, collect the supernatant, start the membrane filtration equipment, and the supernatant After filtration through a microporous membrane, the membrane thickness is 90 μm, the filtered particle size is 8 μm, and the operating pressure is 0.1 MPa, ensuring that the filtrate is clear and transparent, and the filter residue is removed;

(e) vacuum concentration: the filtrate is concentrated in vacuo, concentration temperature 75 ° C, vacuum -0.08Mpa, a concentrated solution, the relative density is controlled at 1.04;

(f) Spray drying: The above concentrated liquid was spray-dried, and the process parameters were a nozzle temperature of 175 ° C, an outlet temperature of 90 ° C, and a feeding speed of 2.5 L/min, to obtain a malt extract required to meet the quality requirements.

The above malt extract has an enzymatic hydrolysis rate of more than 90%, and the energy efficiency improves the absorption and utilization rate of the above-mentioned extract by the body, and the effect can be absorbed without digesting or slightly digesting.

In the present embodiment, the white and polysaccharides and Atractylodes polysaccharides are prepared by the same preparation method, which is prepared by pulverizing, dynamic countercurrent extraction, centrifugation, concentrated alcohol precipitation, ion exchange, vacuum concentration and spray drying. . The specific preparation process is as follows:

(a) Dynamic countercurrent extraction: taking pulverized white and or atractylodes, adding 10 times the weight of the material, and dynamically countercurrently extracting at 95 ° C to obtain an extract, the extraction times are 2 times, each extraction time is 2 hours;

(b) centrifugation: the extract is centrifuged to obtain a clear liquid;

(c) Concentrated alcohol precipitation: The clear liquid was concentrated in a vacuum, the relative density was controlled at 1.12, and the edible liquid was added to the concentrated liquid to make the alcohol content in the mixed solution reach 50%, stirred well, and allowed to stand overnight. The supernatant is recovered from the solvent, and the precipitate is reserved for use;

(d) ion exchange: the alcohol precipitation polysaccharide is dissolved in 6 times purified water, fully stirred and dissolved, and then filtered, and the filtered filtrate is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(e) Vacuum concentration: The ion exchanged solution is concentrated in a vacuum, the concentration is 60 ° C, the degree of vacuum is -0.09 MPa, and the polysaccharide concentrate is obtained, and the relative density is controlled between 1.04:

(f) Spray drying: The polysaccharide concentrate was sterilized and filtered, and then spray-dried. The process parameters were a nozzle temperature of 180 ° C, an outlet temperature of 85 ° C, and a feed rate of 2.0 L/min to obtain a white polysaccharide or an Atractylodes polysaccharide.

The polysaccharide extraction rates of white and polysaccharide and Atractylodes polysaccharide were 35.2% and 8.28%, respectively.

In this embodiment, the Tremella fuciformis polysaccharide is prepared by the production process of the Tremella by pulverization, enzymatic hydrolysis, extraction, ion exchange, vacuum concentration and vacuum freeze drying. The specific preparation process is as follows;

(a) Enzymatic hydrolysis: taking the pulverized Tremella ultrafine powder, adding 50 times of the weight of the material and 0.5% pectinase, thermolyzing at 50 ° C for 60 min, and heating to 95 ° C or above;

(b) Extraction: immersion in 95 ° C temperature, 500 mesh screen filtration to obtain Tremella polysaccharide extract, the number of extraction times is 2, each extraction time is 2 hours;

(c) ion exchange: the filtered Tremella polysaccharide extract is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(d) vacuum concentration: the ion exchanged solution is concentrated in a vacuum, the concentration of 65 ° C, the degree of vacuum -0.09Mpa, the Tremella polysaccharide concentrate, the relative density is controlled at 1.02;

(e) Vacuum freeze-drying: Tremella polysaccharide powder is prepared by vacuum freeze-drying technique, wherein pre-freezing initial temperature is -45 ° C, pre-freezing rate is 0.2 ° C / min, pre-freezing end temperature is -65 ° C, drying chamber The pressure was 50 Pa, the heating plate temperature was 30 ° C, and the drying time was 24 h.

The polysaccharide extraction rate of Tremella polysaccharides was as high as 37.4%.

The preparation method of the probiotic microcapsules for maintaining the activity of the strain in the present embodiment is as follows:

(1) Culture and isolation of the strain: Lactobacillus acidophilus was planted in MRS liquid medium for activation and proliferation culture at a culture temperature of 37 ° C and a culture time of 24 h. The medium is subjected to low temperature centrifugation, the supernatant is removed, and the obtained Lactobacillus acidophilus cells are collected, wherein the centrifugation temperature is 4 ° C, the centrifugal speed is 5000 r / min, and the centrifugation time is 10 min;

(2) coating the first wall material: adding the above-mentioned Lactobacillus acidophilus cells to the mixed solution of the first wall material, fully stirring to obtain a bacterial suspension, stirring time is 8 min, and then centrifuging the above-mentioned bacterial suspension The sprayer was slowly sprayed into the gallic acid solution of pH=4.0, allowed to stand for 15 min, and the small particle precipitate was collected and washed several times with sterile water adjusted to pH 4.0;

(3) coating the second wall material; putting the small particles into the mixed solution of the wall material, uniformly stirring for 5 minutes to prepare the bacterial suspension, and then slowly spraying the above-mentioned bacterial suspension into the 0.2 mol/L by a centrifugal atomizer. Calcium lactate solution, curing for 30 min, washing and filtering to obtain microcapsules;

(4) Vacuum freeze-drying: the above microcapsules were obtained by vacuum freeze-drying to obtain a dry powder of Lactobacillus acidophilus microcapsules, wherein the initial temperature of pre-freezing was -35 ° C, the pre-freezing rate was 0.6 ° C / min, and the pre-freezing end temperature was -60 ° C. The drying chamber pressure was 60 Pa, the heating plate temperature was 35 ° C, and the drying time was 18 h.

The solvent used in the above mixed solution and washing water is sterile water, and the above MRS medium, wall material mixed solution, gallic acid solution and calcium lactate solution are subjected to high temperature moist heat sterilization, and the sterilization temperature is 121 ° C, and the sterilization time is 20min. The dry powder of Lactobacillus acidophilus microcapsules prepared by the above preparation method has a viable cell count of 1.92×1011 CFU/mL, a particle size of 100 μm, uniform particle size distribution and controllable quality, and the microcapsule embedding rate is greater than 92%. The obtained microcapsules have good bacterial activity and can adapt to changes in various environmental factors such as light, heat, oxygen, metal ions and pH, and have good stability.

Example 2: The preparation method of the probiotic microcapsules for maintaining the activity of the strain in the present embodiment is the same as that of the first embodiment, except that the probiotics in the present embodiment are Bifidobacterium longum, and the first and second wall materials are both The mixture was diluted with a sterile water having a wall material of 5 times as a solvent to form a mixed solution, and then embedded. The mass ratio of the probiotic to the first wall material was 1:0.75, the casein in the first wall material was 15 parts, the malt extract was 6 parts, and the oligomeric sugar was 3 parts. The mass ratio of probiotics to the second wall material is 1:1.2, the white wall polysaccharides are 8 parts, the white and polysaccharides are 5 parts, the atractylodes polysaccharide is 6 parts, the guar gum is 0.6 parts, and the pectin is 1.2. Share.

In the present embodiment, the malt extract is obtained by pulverizing, slurrying, enzymatic hydrolysis, centrifugal filtration, vacuum concentration, and spray drying, and the specific preparation steps are as follows:

(a) pulverization; pulverizing barley malt or barley into ultrafine powder of about 110 μm by ultrafine pulverization;

(b) pulping; the barley malt powder or barley flour is prepared into a protein dispersion with pure water, and the pH of the dispersion is adjusted to 6.0;

(c) Enzymatic hydrolysis: heating and maintaining the temperature at 40 ° C, then adding α-amylase, β-amylase and neutral protease to the dispersion, stirring and homogenizing for 2 h, wherein α-amylase The amount added is 0.4% of the substrate content, the amount of β-amylase added is 0.2% of the substrate content, and the amount of neutral protease added is 0.3% of the substrate content, and the enzyme is subjected to high temperature inactivation after the end of enzymatic hydrolysis;

(d) Centrifugal filtration: Centrifugation of the enzymatic hydrolysate after inactivation of the enzyme, maintaining a centrifugal speed of 14000 r/min, feeding speed of the peristaltic pump 3.0 L/min, collecting the supernatant, starting the membrane filtration device, and the supernatant After filtration through a microporous membrane, the membrane thickness is 90 μm, the filtered particle size is 6 μm, and the operating pressure is 0.15 MPa, ensuring that the filtrate is clear and transparent, and the filter residue is removed;

(e) vacuum concentration: the filtrate is concentrated in vacuo, concentration temperature 80 ° C, vacuum -0.08Mpa, to obtain a concentrated liquid, the relative density is controlled at 1.05;

(f) Spray drying: The above concentrated liquid was spray-dried, and the process parameters were a nozzle temperature of 180 ° C, an outlet temperature of 85 ° C, and a feeding speed of 2.0 L/min, to obtain a malt extract required to meet the quality requirements.

The above malt extract has an enzymatic hydrolysis rate of more than 90%, and the energy efficiency improves the absorption and utilization rate of the above-mentioned extract by the body, and the effect can be absorbed without digesting or slightly digesting.

In the present embodiment, the white and polysaccharides and Atractylodes polysaccharides are prepared by the same preparation method, which is prepared by pulverizing, dynamic countercurrent extraction, centrifugation, concentrated alcohol precipitation, ion exchange, vacuum concentration and spray drying. . The specific preparation process is as follows:

(a) Dynamic countercurrent extraction: taking pulverized white or atractylodes, adding 9 times the weight of the material, and extracting it dynamically at 95 ° C to obtain the extract, the extraction times are 2 times, each extraction time is 1.5 hours;

(b) centrifugation: the extract is centrifuged to obtain a clear liquid;

(c) Concentrated alcohol precipitation; the clear liquid was concentrated in a vacuum, the relative density was controlled at 1.10, and the edible liquid was added to the concentrated liquid to make the alcohol content in the mixed solution reach 60%, stirred well, and allowed to stand overnight. The supernatant is recovered from the solvent, and the precipitate is reserved for use;

(d) ion exchange: the alcohol precipitation polysaccharide is dissolved in 6 times purified water, fully stirred and dissolved, and then filtered, and the filtered filtrate is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(e) vacuum concentration: the ion exchanged solution is concentrated in a vacuum, the concentration temperature is 70 ° C, the degree of vacuum -0.09 Mpa, the polysaccharide concentrate is obtained, the relative density is controlled between 1.03;

(f) Spray drying: The polysaccharide concentrate was sterilized and filtered, and then spray-dried. The process parameters were a nozzle temperature of 180 ° C, an outlet temperature of 85 ° C, and a feed rate of 2.0 L/min to obtain a white polysaccharide or an Atractylodes polysaccharide.

The polysaccharide extraction rates of white and polysaccharides and Atractylodes polysaccharides were 35.4% and 8.24%, respectively.

In this embodiment, the Tremella fuciformis polysaccharide is prepared by the production process of the Tremella by pulverization, enzymatic hydrolysis, extraction, ion exchange, vacuum concentration and vacuum freeze drying. The specific preparation process is as follows:

(a) Enzymatic hydrolysis: taking the pulverized white fungus ultrafine powder, adding 50 times the weight of the material and 1% pectinase, and digesting at 50 °C for 80 min, and heating to 95 ° C or above;

(b) Extraction: immersion in 95 ° C temperature, 500 mesh screen filtration to obtain Tremella polysaccharide extract, the number of extraction times is 2, each extraction time is 2 hours;

(c) ion exchange: the filtered Tremella polysaccharide extract is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

(d) vacuum concentration: the ion exchanged solution is concentrated in a vacuum, the concentration of 65 ° C, the degree of vacuum -0.09Mpa, the Tremella polysaccharide concentrate, the relative density is controlled at 1.02;

(e) Vacuum freeze-drying: Tremella polysaccharide powder is prepared by vacuum freeze-drying technique, wherein pre-freezing initial temperature is -40 ° C, pre-freezing rate is 0.3 ° C / min, pre-freezing end temperature is -65 ° C, drying chamber The pressure was 50 Pa, the heating plate temperature was 35 ° C, and the drying time was 24 h.

The polysaccharide extraction rate of Tremella polysaccharides was as high as 36.9%.

The preparation method of the probiotic microcapsules for maintaining the activity of the strain in the present embodiment is as follows:

(1) Culture and isolation: Bifidobacterium longum was planted into MRS liquid medium for activation and proliferation culture at a culture temperature of 36 ° C and a culture time of 36 h. The medium is subjected to low temperature centrifugation, the supernatant is removed, and the obtained Bifidobacterium longum cells are collected, wherein the centrifugation temperature is 10 ° C, the centrifugal speed is 4000 r / min, and the centrifugation time is 8 min;

(2) coating the first wall material: adding the above Bifidobacterium longum cells to the first wall material mixing solution, stirring time is 10 min, and then slowly spraying the above-mentioned bacterial suspension into the PH=3.6 with a centrifugal sprayer. In the gallic acid solution, it was allowed to stand for 10 min, and the small particle precipitate was collected and washed several times with sterile water adjusted to pH 4.0;

(3) coating the second wall material: the small particles are put into the second wall material mixed solution, uniformly stirred for 8 minutes to prepare a bacterial suspension, and the above suspension is slowly sprayed into the 0.15 mol/L by a centrifugal atomizer. Calcium lactate solution, curing for 50 min, washing and filtering to obtain microcapsules;

(4) Vacuum freeze-drying: the above microcapsules are obtained by vacuum freeze-drying to obtain a dry powder of Bifidobacterium longum microcapsules, wherein the initial temperature of pre-freezing is -30 ° C, the pre-freezing rate is 0.8 ° C / min, and the pre-freezing end temperature is -60 ° C. The drying chamber pressure was 80 Pa, the heating plate temperature was 30 ° C, and the drying time was 20 h.

The solvent used in the above mixed solution and washing water is sterile water, and the above MRS medium, wall material mixed solution, gallic acid solution and calcium lactate solution are subjected to high temperature moist heat sterilization, and the sterilization temperature is 125 ° C, and the sterilization time is 15min. The number of live bacteria of the Bifidobacterium longum microcapsules prepared by the above preparation method is 1.39×1011 CFU/mL, the particle size is 80 μm, the particle size distribution is uniform and the quality is controllable, and the microcapsule embedding rate is greater than 92%, The obtained microcapsules have good strain activity and can adapt to various environmental factors such as light, heat, oxygen, metal ions and pH, and have good stability.

Example 3;

The preparation method of the probiotic microcapsule tablet in this embodiment is:

The Lactobacillus acidophilus microcapsule powder and the Bifidobacterium longum microcapsule powder prepared in the above Examples 1 and 2 were mixed in a ratio of 2:1, and 10% of the total weight of the probiotic microcapsule powder was weighed. A 1% magnesium stearate was put into the above mixture and stirred for 5 minutes, and the pellet was tableted with 0.45 g/tablet to prepare a probiotic microcapsule tablet. After the 2-year shelf life, the above-mentioned tablets had a total viable cell count of 31% at the time of production, and the strain activity was good and the survival rate was high.

Example 4:

The preparation method of the probiotic microcapsule capsule in the embodiment is:

The Lactobacillus acidophilus microcapsule powder and the Bifidobacterium longum microcapsule powder prepared in the above Examples 1 and 2 were mixed in a ratio of 1:1, and 15% of the total weight of the probiotic microcapsule powder was weighed. 1.5% magnesium stearate was put into the above mixture, and stirred for 5 minutes, that is, the granules were filled with 0# capsules, 0.35 g/granules to prepare probiotic microcapsule capsules. After the 2-year shelf life, the capsule has a total viable cell count of 34% at the time of production, and the activity of the strain is good and the survival rate is high.

Example 5:

The preparation method of the probiotic microcapsule granule in the embodiment is:

The Lactobacillus acidophilus microcapsule powder and the Bifidobacterium longum microcapsule powder prepared in the above Examples 1 and 2 were mixed in a ratio of 3:1, and the oligomeric wood of 1% of the total weight of the probiotic microcapsule powder was weighed. Sugar, 25% maltodextrin, 1% silica was put into the above mixture, and stirred for 5 minutes, that is, the granules were filled with an aluminum foil strip packaging bag, 5 g/bag, to prepare probiotic microcapsule granules. After the 2-year shelf life, the granules have a total viable cell count of 29% at the time of production, and the strain activity is good and the survival rate is high.

Example 6

The preparation method of the probiotic microcapsule yogurt in the embodiment is:

In the prepared yoghurt, the Lactobacillus acidophilus microcapsule powder and the Lactobacillus casei microcapsule powder prepared in the above Examples 1 and 2 were added, and the amount of each added was 1% by weight of the yoghurt, and the mixture was uniformly stirred to obtain a probiotic microbe. Capsule yoghurt, by sampling the yoghurt of different storage dates, the viable count of the two probiotics is measured, reflecting the change of the number of live probiotics within the shelf life of the yoghurt added with the probiotic microcapsules, as shown in Figure 1 below. The trend of the amount of live bacteria of Lactobacillus acidophilus and Lactobacillus casei over time.

Yogurt has a low PH value and is not suitable for the survival of most probiotics. Due to the special microcapsule embedding layer of the present invention, the probiotic microcapsules form agglomerates in the yogurt, fully protecting the internal probiotics until after drinking. Into the small intestine, the pH is significantly improved, the aggregation phenomenon gradually disappears, the capsule shell dissolves into nutrients such as protein and polysaccharide, and the probiotics are fully released to produce the proper effect in the intestine.

Lactobacillus acidophilus decreased from the initial 1.86 x 109 CFU/mL viable count to 1.45 x 109 CFU/mL, a decrease of only 22.1%. Lactobacillus casei decreased from the initial 1.48 x 109 CFU/mL viable count to 1.1 x 109 CFU/mL, a decrease of only 25.7%. From the above data, it can be seen that the survival rate of probiotics in the shelf life of yogurt is high, and the amount of live bacteria is not in the order of magnitude.

The invention also tests the acid stability, durable storage property and thermal stability of the probiotic microcapsules prepared in Examples 1 and 2, specifically:

1.1 Probiotic microcapsule stability test

The microcapsule shell of the probiotic microcapsule of the invention is mainly composed of protein and compound plant polysaccharide, and the aggregate structure formed by them is not dissolved in an acidic environment, and the core probiotic can be protected from being safely passed through the gastric juice into the intestinal tract for dissolution and colonization. The released nutrients in the capsule shell can quickly provide nutrients to the probiotic bacteria, and the probiotics can rapidly multiply. Therefore, the significance of microcapsule embedding is to isolate the direct contact between the probiotics and the external harsh environment, control the place where the probiotics are released, and provide a nutrient source for the probiotics, and maintain the stability of the strain activity to a large extent. The bioavailability of probiotics.

The acid stability test was carried out with Lactobacillus acidophilus, Bifidobacterium longum, Lactobacillus casei unembedded cells and microcapsules as test materials. The material test environment was artificial gastric acid solution (pH=2.1), focusing on the investigation. The effect of microencapsulation on the survival rate of the acid environment of the strain was as shown in Fig. 2 below. It can be seen from Fig. 2 that the probiotic bacteria which have not been encapsulated by microcapsules rapidly die in a strong acidic environment, and the survival rate is extremely low, and the microcapsule-embedded probiotics have excellent acid resistance and high survival rate. It can be seen from the above that the microcapsule embedding method of the present invention can significantly improve the acid resistance of the strain and improve the survival rate and the activity of the strain.

1.2 Probiotic microcapsule durability test

The durability test was carried out by using Lactobacillus acidophilus, Bifidobacterium longum, Lactobacillus casei unembedded cells and microencapsulated dry powder as experimental materials, and focusing on microcapsule embedding for accelerated test environment (40 ° C, The effect of survival rate within 35 days under humidity 75%) is shown in Figure 3 below. It can be seen from Fig. 3 that after three weeks of accelerated test environment, the three probiotic microcapsule powders reduced the number of live bacteria of the strain by only one log, the survival rate of the strain was higher, and the three probiotics were not embedded. At least 5 to 6 logs were lost to the cells, and the survival rate of the strains was low. It can be seen from the above that the microcapsule embedding method of the present invention can significantly increase the survival rate of the strain and maintain the activity of the strain.

1.3 Probiotic microcapsule thermal stability test

The invention not only has excellent acid resistance and durability, but also has excellent high temperature resistance and good thermal stability. Lactobacillus acidophilus microcapsule powder, Bifidobacterium longum microcapsule powder and Lactobacillus casei microcapsule powder were used as test materials to investigate the changes of their species survival rate under different temperature and time conditions. The specific data are shown in Table 1 below. Show.

Table 1 Results of heat stability test of probiotic microcapsules

It can be seen from the data in Table 1 that the probiotics embedded in the microcapsules have excellent high temperature resistance characteristics, can withstand the test of severe processing techniques, still have a large survival rate, excellent strain activity and good thermal stability. It can have a broader industrial application prospect.

The above description is only a preferred embodiment of the present invention, and is not intended to limit the scope of the invention, and the equivalents of the embodiments of the present invention may be made without departing from the principles of the invention. A number of improvements and refinements, either directly or indirectly in other related technical fields, are included within the scope of the patent protection of the present invention.

Claims (10)

1. A probiotic microcapsule for maintaining the activity of a strain, comprising a core material and a wall material coated on the outside of the core material, the core material being a probiotic, characterized in that the wall material comprises a coating outside the core material a first wall material and a second wall material wrapped around the first wall material,

   The first wall material comprises the following components: casein, malt extract and xylooligosaccharide;

   The second wall material comprises the following components: Tremella polysaccharide, white and polysaccharide, atractylodes polysaccharide, guar gum and pectin.
2. The probiotic microcapsule for maintaining the activity of the strain according to claim 1, wherein the first and second wall materials are each diluted with a sterile water as a solvent to form a mixed solution, and then embedded. The quality of the sterile water used when the second wall material is diluted into a mixed solution is 3 to 5 times the mass of the wall material.
3. The probiotic microcapsule for maintaining bacterial activity according to claim 1, wherein the mass ratio of the probiotic to the first wall material is 1: (0.5 to 2), the first by weight 5 to 15 parts of casein in the wall material, 1 to 6 parts of the malt extract, and 0.5 to 3 parts of the xylooligosaccharide; the mass ratio of the probiotic to the second wall material is 1: (0.5 to 1.5) According to the weight, the second wall material has 2 to 8 parts of Tremella polysaccharide, 2 to 5 parts of white and polysaccharide, 3 to 6 parts of atractylodes polysaccharide, 0.2 to 0.6 parts of guar gum, and 0.6 to pectin. ~ 1.2 parts.
4. The probiotic microcapsule for maintaining the activity of a strain according to claim 1 or 2 or 3, wherein the probiotic is one or more of Lactobacillus, Bifidobacterium, and Streptococcus.
5. A method for preparing a probiotic microcapsule for maintaining strain activity according to any one of claims 1 to 4, characterized in that it comprises the following steps

   (1) Centrifugally treating the probiotic culture medium in the activated proliferation culture, and removing the supernatant after the treatment to obtain the probiotic bacteria;

   (2) adding casein, malt extract and xylooligosaccharide to water to obtain a mixed solution of the first wall material, adding the probiotic bacteria to the mixed solution of the first wall material, and stirring to obtain a bacterial suspension The bacterial suspension is slowly sprayed into a gallic acid solution having a pH of 3.0 to 5.0 by a sprayer, allowed to stand for 5 to 20 minutes, and a small particle precipitate is collected, and the collected small particles are washed in a sterile water having a pH of 3.8 to 4.2;

   (3) adding Tremella polysaccharide, white and polysaccharide, Atractylodes polysaccharide, guar gum and pectin to water to obtain a mixed solution of the second wall material, and the small particles obtained in the step (2) are put into the mixed solution of the second wall material. The bacterial suspension is prepared by stirring, and the bacterial suspension is slowly sprayed into a 0.05-0.5 mol/L calcium lactate solution by a sprayer to be solidified, washed, and filtered to obtain microcapsules;

   (4) The above microcapsules are obtained by vacuum freeze-drying to obtain a probiotic microcapsule dry powder.
6. The method for preparing a probiotic microcapsule for maintaining the activity of a strain according to claim 5, wherein the centrifugation speed of the centrifugation in the step (1) is 4000 to 6000 r/min, and the centrifugation time is 5 to 12 min.
7. The method for preparing a probiotic microcapsule for maintaining bacterial activity according to claim 5, wherein the vacuum freeze-drying pre-freezing initial temperature in the step (4) is -30 to -40 ° C, and the pre-freezing rate is 0.4. ~0.8 °C / min, pre-freezing end temperature -60 ° C, drying chamber pressure 40 ~ 100pa, heating plate temperature 30 ~ 40 ° C, drying time is 12 ~ 20h.
8. The method for preparing a probiotic microcapsule for maintaining strain activity according to claim 5, wherein the malt extract is prepared as follows

   (a) pulverization: pulverizing barley malt or barley into ultrafine barley malt powder or barley flour of 90-110 μm by ultrafine pulverization technology;

   (b) pulping: the barley malt powder or barley flour is prepared into a protein dispersion with purified water, and the pH of the dispersion is adjusted to between 6 and 7;

   (c) Enzymatic hydrolysis: heating and maintaining the temperature of the dispersion between 40 and 50 ° C, then adding α-amylase, β-amylase and neutral protease to the dispersion, stirring uniformly and then enzymolysis for 2 h~ 4h, wherein the amount of α-amylase added is 0.3% to 0.5% of the substrate content, the amount of β-amylase added is 0.1% to 0.3% of the substrate content, and the amount of neutral protease added is 0.2% of the substrate content. ~0.4%, after the end of enzymatic hydrolysis, the enzyme is subjected to high temperature;

   (d) Centrifugal filtration: Centrifugation of the enzymatic hydrolysate after inactivation of the enzyme, maintaining a centrifugal speed of 12000 to 16000 r/min, feeding speed of the peristaltic pump of 1.5 to 3.5 L/min, collecting the supernatant, and starting the membrane filtration device. The supernatant is filtered through a microporous membrane, the membrane thickness is 90-150 μm, the filtered particle size is 4-8 μm, and the operating pressure is 0.01-0.2 MPa, ensuring that the filtrate is clear and transparent, and the filter residue is removed;

   (e) vacuum concentration: the filtrate is concentrated in a vacuum, concentrated at 65-85 ° C, vacuum -0.07 ~ -0.09Mpa, to obtain a concentrated liquid, the relative density is controlled between 1.04 ~ 1.08;

   (f) Spray drying: The above concentrated liquid was spray-dried, and the process parameters were a nozzle temperature of 175 to 190 ° C, an outlet temperature of 80 to 95 ° C, and a feeding speed of 1.5 to 3.5 L/min to obtain the malt extract.
9. The method for preparing a probiotic microcapsule for maintaining strain activity according to claim 5, wherein the white and polysaccharide/Atractylodes polysaccharide are obtained by the following steps

   (a) Dynamic countercurrent extraction: take pulverized white and / white atractylodes, add 8 to 10 times the weight of the material, and dynamically countercurrently extract at 90 ~ 100 ° C to obtain the extract, the number of extraction times is 1 to 3 times, each time The extraction time is 1 to 3 hours;

   (b) centrifugation: the extract is centrifuged to obtain a clear liquid;

   (c) Concentrated alcohol precipitation: The clear liquid is concentrated in a vacuum, the relative density is controlled between 1.10 and 1.20, and the edible liquid is added to the concentrated liquid to make the alcohol content in the mixed solution reach 40-80%, fully stirred and allowed to stand. Overnight; the supernatant recovers the solvent and leaves the precipitate for use;

   (d) ion exchange: the alcohol precipitation polysaccharide is dissolved in 4-8 times purified water, stirred well, dissolved, and filtered, and the filtered filtrate is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

   (e) vacuum concentration under vacuum: the ion exchanged solution is concentrated in a vacuum, concentrated temperature 60-80 ° C, vacuum -0.07 ~ -0.09Mpa, to obtain a polysaccharide concentrate, the relative density is controlled between 1.03 ~ 1.06;

   (f) spray drying: the polysaccharide concentrate is sterilized and filtered, and then spray-dried, the process parameters are nozzle temperature 170-185 ° C, outlet temperature 75-95 ° C, feeding speed 2.0-4.0 L / min, to obtain the white And polysaccharide / Atractylodes polysaccharide.
10. The method for preparing a probiotic microcapsule for maintaining bacterial activity according to claim 5, wherein the tremella polysaccharide is obtained by the following steps

    (a) Enzymatic hydrolysis: The white fungus is pulverized into ultrafine powder of 90-110 μm by ultrafine pulverization technology, 30~50 times of water and 0.5~2% pectinase are added, and the temperature is 40-50 °C. 30-90min, the temperature is raised to above 95 °C to kill the enzyme;

    (b) Extraction: immersion in 95-100 ° C temperature, 500-mesh sieve to obtain Tremella polysaccharide extract, the number of extraction times is 1-2 times, each extraction time is 1-3 hours;

    (c) ion exchange: the filtered Tremella polysaccharide extract is subjected to deionization and decolorization treatment through an ion exchange column and an activated carbon column;

    (d) Vacuum concentration: the ion exchanged solution is concentrated in a vacuum, the concentration temperature is 60-80 ° C, the degree of vacuum is -0.07-0.09 Mpa, and the white fungus polysaccharide concentrate is obtained, and the relative density is controlled between 1.02 and 1.04;

    (e) Vacuum freeze-drying: Tremella polysaccharide powder is prepared by vacuum freeze-drying technique, wherein the initial temperature of pre-freezing is -35 to -45 ° C, the pre-freezing rate is 0.2-0.5 ° C / min, and the pre-freezing end temperature - 65 ° C, drying chamber pressure 50 ~ 120pa, heating plate temperature 30 ~ 40 ° C, drying time is 16 ~ 24h.

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