CN101856604A - Method for preparing probiotic microcapsules by using electrostatic spraying - Google Patents
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- CN101856604A CN101856604A CN201010193794A CN201010193794A CN101856604A CN 101856604 A CN101856604 A CN 101856604A CN 201010193794 A CN201010193794 A CN 201010193794A CN 201010193794 A CN201010193794 A CN 201010193794A CN 101856604 A CN101856604 A CN 101856604A
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Abstract
The invention discloses a method for preparing probiotic microcapsules by using electrostatic spraying. Sodium alga acid and gelatin are taken as wall materials, probiotic suspension or microporous starch-adsorbed probiotics are taken as core materials, and calcium chloride is taken as solidifying solution. The method comprises the following steps of: a) mixing 1.5 percent solution of sodium alga acid and 1 percent gelatin in the same volume, and uniformly mixing with the probiotic suspension or microporous starch-adsorbed probiotics to form mixed solution; b) putting the mixed solution into a container, and injecting the mixed solution into 2 to 3 percent continuously stirred solution of calcium sodium chloride in the electrostatic spraying mode by using a microcapsule molding device to form microcapsules; c) standing and solidifying the encapsulated probiotic microcapsules for 20 minutes in the solution of calcium chloride, and filtering and cleaning the solution by using physiological saline; and d) saving the probiotic microcapsules in the physiological saline. The prepared microcapsules have the advantages of uniform grain diameter, good encapsulation performance, simple process, readily available wall materials, and low cost, and can be widely applied to the fields of food processing, biomedicines, chemical industry, and the like.
Description
Technical field
The present invention relates to a kind ofly prepare the method for probiotic microcapsule, keep the activity of probio, be applied to technical fields such as health food, biological feedstuff with electrostatic spray.
Background technology
Present confirmed probio has Pseudomonas kinds such as Bacillus acidi lactici, Bifidobacterium, streptococcus, bacillus and yeast.A large amount of research reports and clinical testing confirm, probio as: lactobacillus acidophilus is as the normal flora in the human body intestinal canal, have the lactose intolerance of alleviation, the formation of inhibition tumour cell, reduce cholesterol levels, alleviate physiological functions such as diarrhoea and enhancing immunity of organisms, it is the important microbe in the human body intestinal canal, healthy closely bound up with human body, be the probio of very paying attention to research and development in the present lactic acid bacteria family, be regarded as third generation yogurt starter culture.In recent years, along with deepening continuously that probio is studied, probio has formed powerful industry in the application aspect food, the health products.As sour milk, cheese, sour cream and the ice-cream product etc. that add the baby of leben, liquid milk, interpolation Bifidobacterium of lactobacillus and Bifidobacterium and person in middle and old age's formula milk, the bread that adds Bifidobacterium and biscuits and add probio; Probio also is used as microbial forage additive and is applied to aquatic products animal husbandry breed field.
But the most key problem that exists in the lactobacillus product is the product viable count to descend comparatively fast.According to Kurmann, in lactobacteria-containing food compositions, have only when the viable lactic acid bacteria number to reach 10
6More than the cfu/ml, the normal physiological hygiene function of competence exertion.Probio is used for the positive-effect of fermented food or food additives is generally acknowledged, but many urgency problem to be solved is also arranged:
(1) probio such as lactobacillus and Bifidobacterium mostly is anaerobic bacteria or oxygen resistence anaerobic bacteria, in growth course, does not form gemma, resistance is relatively poor, most acid resistance is relatively poor, very responsive to oxygen, the active difficulty that keeps, easy inactivation in processing, transportation, storage process;
(2) probio will produce beneficial effect to human body, and at first it must also be grown on intestinal mucosa with a large amount of survival bacterium arrival enteron aisles surely by gastric environment.But in the environment of hydrochloric acid in gastric juice and cholate, lactobacillus acidophilus and Bifidobacterium bacterium number can very fast declines in the dairy products, and making does not finally have enough probios to play a role in human body.
In order to improve the activity of probio, people study and have adopted several different methods, for example: utilize genetic engineering that probio is transformed, foreign DNA is incorporated on the chromosome of object bacteria, and utilize regulating system to modify.By this method; can obtain under adverse environments such as food processing process, keeping great-hearted probio; improve the research of condition of culture raising activity of probiotic and carrying out always, for example: condition of storage, the interpolation of protectiveness compound and the optimization of freezing and drying condition etc. of probiotic active in the research process.
In addition, utilize microcapsule embedded technology, the probio embedding is got up, the survival of probio under gastroenteric environment made moderate progress, and their protectiveness shell can make their better survivals in food system.Simultaneously, embedding techniques can improve the survival rate of thalline in dry run and storage process, and utilizing microcapsule embedded technical protection probio is the focus of studying both at home and abroad at present, is considered to have a kind of method of application prospect.Yet, there is the microcapsule method of bibliographical information that emulsion process and extrusion are arranged at present, the envelop rate of emulsion process is low, only has an appointment 60%, and the microencapsulation probio particle diameter of extrusion preparation is more greatly at 2～3mm, uses to be restricted in food.
Summary of the invention
The invention discloses and a kind ofly prepare the method for probiotic microcapsule, application invention patent with electrostatic spray: " microcapsules shaped device ", (ZL00218100.2) probio is carried out microcapsule embedded, to strengthen its resistance to adverse circumstance.Lactic acid bacteria is carried out microencapsulation not only can make the thalline and the poor environment in the external world separate; Odour masking; Formation is similar to spherical solid microparticle, is convenient to add in the various food such as liquid, jelly and powder, and helps storing and transportation; Adopt enteric solubility wall material, can also prevent the destruction of gastric juice, thereby make thalline as much as possible arrive enteron aisle, play wholesome effect.By microencapsulation, be expected to improve the stability of probio in production, storage and process of consumption, prepare anti-storage, high temperature resistant, high pressure resistant, acid proof probiotics bacterial probiotics.
1, a kind of method for preparing probiotic microcapsule with electrostatic spray, with sodium alginate and gelatin as the wall material, adsorb probio as core with probio suspension or micropore starch, calcium chloride is consolidation liquid, sodium alginate and gelatin, interpolation probio suspension or micropore starch absorption probio is mixed, under electrostatic interaction, be sprayed in the calcium chloride solution, form the calcium alginate film, in calcium chloride solution, solidify behind the encystation, store with physiological saline washing back; It is characterized in that step is as follows:
A) be 1.5% sodium alginate soln with concentration with concentration be that 1% gelatin mixes by equal-volume, form mixed liquor with probiotics bacterial suspension or micropore starch absorption probio mixing again;
B) mixed liquor being placed container, is in the calcium chloride acid sodium solution of 2%-3% by the nozzle of microcapsules shaped device by the concentration that the electrostatic spray mode sprays into continuous stirring, forms microcapsules;
C) with the probiotic microcapsule behind the encystation, in calcium chloride solution, leave standstill and solidify 20min, filter and clean with physiological saline;
D) place physiological saline to preserve probiotic microcapsule then.
Described core is 1 with the quality of wall material or the ratio of volume: 2-3.
The addition of described micropore starch is 6% of a probiotics bacterial suspension quality.
Described its electric-field intensity of electrostatic spray mode is 4～6kV, and the distance between syringe needle and the calcium chloride liquid level is 20～30mm, and the fltting speed of syringe is 50～70mm/h.
Described electrostatic spray mode is the electrostatic field that adds high pressure between nozzle and calcium chloride solution, and mixed liquor is dispersed into even, tiny drop with spray liquid under the progradation of electric field force and nozzle.Adjust electric-field intensity, sodium alginate, gelatin and calcium chloride concentration, the distance between nozzle and the calcium chloride liquid level and the fltting speed of syringe can be controlled the probiotic microcapsule particle diameter.
The present invention can be used for the slow controlled release of microencapsulation probio, in cell cultivation and transplanting and the biochemical industry.
The microcapsules particle diameter that adopts electrostatic spray method of the present invention to make is even, the encystation performance is good and technology is simple; The wall material is easy to get, cheap; Can be widely used in fields such as food processing, biological medicine, chemical industry.
Description of drawings
Below in conjunction with drawings and Examples the present invention is described in detail.
Fig. 1 is in the lactic acid bacteria colonial morphology photo of exponential phase through cultivation;
Fig. 2 lactobacillus micro-capsule microphoto;
Fig. 3 bacillus subtilis colonial morphology photo;
Fig. 4 bacillus subtilis bacteria microcapsule microphoto;
The microcapsules shaped device of Fig. 5 high-voltage electrostatic field.
The specific embodiment
Below in conjunction with accompanying drawing and fact Example the present invention is described in detail.
With sodium alginate, gelatin is the wall material, and calcium chloride is consolidation liquid, selects for use to be in the exponential phase physiological property to reach stable probiotics bacterial suspension be core.The microcapsules shaped device (ZL00218100.2) of high-voltage electrostatic field comprises as shown in Figure 5: container 1, nozzle 2, syringe 3, propeller 4, support 5, anodal wiring 6, negative pole wiring 7, high-pressure electrostatic field device 8, nozzle adopt smooth flat syringe needle.
Embodiment 1:
With concentration is that 1.5% sodium alginate soln and concentration are that 1% gelatin solution is by the equal-volume mixing, with miillpore filter (aperture 0.22um) filtration sterilization, promptly get wall material solution, choose then and be in exponential phase, the lactic acid bacteria bacteria suspension of the about 1.1 * 109cfu/ml of viable count (the lactic acid bacteria colonial morphology as shown in Figure 1) and above-mentioned wall material are inhalation syringe behind 1: 2 mixing by mass ratio, utilize the microcapsules shaped device of high-voltage electrostatic field then, choosing electric-field intensity is 3.7 * 10kV, distance between syringe needle and the calcium chloride liquid level is 2.5cm, and the fltting speed of syringe is 50mm/h.The concentration that mixed liquor is sprayed to continuous stirring is 3% to have sterilized in the calcium chloride solution Ca
2+Form microcapsules with the sodium alginate cross-linking reaction, and in calcium chloride solution, leave standstill curing 20min.Filtration makes the Bacillus acidi lactici microcapsules with physiological saline washing 2-3 time.
Also can adopt identical wall material, core adopts has good adsorption properties, the micropore starch that can be used as functional materials, after being 6% micropore starch absorption lactic acid bacteria with content, as core, be to carry out microencapsulation at 1: 3 by core wall material ratio, can find out by the photo in kind of Fig. 2 microencapsulation micropore starch absorption probio, micro-capsule is complete, microcapsule membrane has certain thickness, and embedding is good.
Adopt the lactobacillus micro-capsule of the inventive method preparation, can reach 96.3% the embedding rate of lactic acid bacteria; To micropore starch absorption lactic acid bacteria embedding rate is 85.88%; The lactic acid bacteria of microencapsulation is after handling 2h through simulated gastric fluid, viable count is than high two orders of magnitude of control group without microencapsulation, and microcapsules almost completely discharge after simulated intestinal fluid is handled 40min, sodium alginate of selecting for use and gelatin mixing wall material and lactic acid bacteria have better biocompatibility, and certain acid resistance and enteric solubility are arranged.
The microencapsulation sour milk beverage of preparation, its storage stability with contrast without the lactic acid bacteria and the market milk acid bacteria beverage of embedding.The result shows, without the lactic acid bacteria of embedding and market milk acid bacteria beverage about 2 orders of magnitude that descended in 4 weeks; And the lactobacillus micro-capsule beverage is just by original 4.35 * 10
9Drop to 2.14 * 10
9, fall is very little.As seen microcapsules milk has better storage stability than stoste lactic acid bacteria milk, market milk acid bacteria beverage.
Embodiment 2:
To grow to logarithmic phase bacillus subtilis bacterium liquid (viable count is 1.23 * 109cfu/ml, and the bacillus subtilis colonial morphology as shown in Figure 3),, remove supernatant, collect bacterium mud at 4 ℃ of centrifugal 20min of J-20XP high speed freezing centrifuge (7000rpm).In bacterium mud, add and nutrient solution equal-volume physiological saline, standby after mixing; Sodium alginate soln with 1.5% and 1% gelatin solution mixing, miillpore filter (aperture 0.22 μ m) filtration sterilization is as the wall material; Then, with inhalation syringe behind bacillus subtilis bacteria suspension and the wall material mixing; Utilize the microcapsules shaped device (ZL00218100.2) of high-voltage electrostatic field, choosing electric-field intensity is 4～6kV, distance between syringe needle and the calcium chloride liquid level is 20～30mm for distance, the fltting speed of syringe is 50～70mm/h, and it is in 2% the calcium chloride solution that core wall material mixed liquor is sprayed to the sterilized concentration of continuous stirring.Ca2+ and sodium alginate cross-linking reaction form microcapsules, leave standstill and solidify 20min, filtration is washed 2-3 time with physiological saline, obtain the bacillus subtilis bacteria microcapsule, adopt the bacillus subtilis bacteria microcapsule of the inventive method preparation, observe its profile rounding, smooth surface with inverted microscope, average diameter is 130 μ m, and microcapsules can reach 90.7% to the embedding rate of bacillus subtilis.Fig. 4 is for using the bacillus subtilis bacteria microcapsule microphoto of the inventive method preparation, micro-capsule rounding as can be seen, and bacillus subtilis is wrapped in the micro-capsule, is evenly distributed.
By simulating the experiment of artificial stomach environment and acid resistance; the result shows that sodium alginate, gelatin-compounded wall material have certain protective role to the bacillus subtilis thalline; through the storage endurance experimental verification, adopt the probiotic microcapsule of the inventive method preparation can improve the storage stability of probio.
Claims (4)
1. method for preparing probiotic microcapsule with electrostatic spray, with sodium alginate and gelatin as the wall material, adsorb probio as core with probio suspension or micropore starch, calcium chloride is consolidation liquid, sodium alginate and gelatin, interpolation probio suspension or micropore starch absorption probio is mixed, under electrostatic interaction, be sprayed in the calcium chloride solution, form the calcium alginate film, in calcium chloride solution, solidify behind the encystation, store with physiological saline washing back; It is characterized in that step is as follows:
A) be 1.5% sodium alginate soln with concentration with concentration be that 1% gelatin mixes by equal-volume, form mixed liquor with probiotics bacterial suspension or micropore starch absorption probio mixing again;
B) mixed liquor being placed container, is in the calcium chloride acid sodium solution of 2%-3% by the nozzle of microcapsules shaped device by the concentration that the electrostatic spray mode sprays into continuous stirring, forms microcapsules;
C) with the probiotic microcapsule behind the encystation, in calcium chloride solution, leave standstill and solidify 20min, filter and clean with physiological saline;
D) place physiological saline to preserve probiotic microcapsule then.
2. according to claim 1ly a kind ofly prepare the method for probiotic microcapsule with electrostatic spray, it is characterized in that: described core is 1 with the quality of wall material or the ratio of volume: 2-3.
3. according to claim 1ly a kind ofly prepare the method for probiotic microcapsule with electrostatic spray, it is characterized in that: the addition of described micropore starch is 6% of a probio suspension quality.
4. a kind of method for preparing probiotic microcapsule with electrostatic spray according to claim 1, it is characterized in that: described its electric-field intensity of electrostatic spray mode is 4～6kV, distance between syringe needle and the calcium chloride liquid level is 20～30mm, and the fltting speed of syringe is 50～70mm/h.
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