CN105105146A - Probiotics activity retention method and application thereof to solid-state fatty food - Google Patents
Probiotics activity retention method and application thereof to solid-state fatty food Download PDFInfo
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- CN105105146A CN105105146A CN201510605065.3A CN201510605065A CN105105146A CN 105105146 A CN105105146 A CN 105105146A CN 201510605065 A CN201510605065 A CN 201510605065A CN 105105146 A CN105105146 A CN 105105146A
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- lactobacillus
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23G—COCOA; COCOA PRODUCTS, e.g. CHOCOLATE; SUBSTITUTES FOR COCOA OR COCOA PRODUCTS; CONFECTIONERY; CHEWING GUM; ICE-CREAM; PREPARATION THEREOF
- A23G1/00—Cocoa; Cocoa products, e.g. chocolate; Substitutes therefor
- A23G1/30—Cocoa products, e.g. chocolate; Substitutes therefor
- A23G1/32—Cocoa products, e.g. chocolate; Substitutes therefor characterised by the composition containing organic or inorganic compounds
- A23G1/42—Cocoa products, e.g. chocolate; Substitutes therefor characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins
- A23G1/423—Cocoa products, e.g. chocolate; Substitutes therefor characterised by the composition containing organic or inorganic compounds containing microorganisms or enzymes; containing paramedical or dietetical agents, e.g. vitamins containing microorganisms, enzymes
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23G—COCOA; COCOA PRODUCTS, e.g. CHOCOLATE; SUBSTITUTES FOR COCOA OR COCOA PRODUCTS; CONFECTIONERY; CHEWING GUM; ICE-CREAM; PREPARATION THEREOF
- A23G1/00—Cocoa; Cocoa products, e.g. chocolate; Substitutes therefor
- A23G1/30—Cocoa products, e.g. chocolate; Substitutes therefor
- A23G1/32—Cocoa products, e.g. chocolate; Substitutes therefor characterised by the composition containing organic or inorganic compounds
- A23G1/44—Cocoa products, e.g. chocolate; Substitutes therefor characterised by the composition containing organic or inorganic compounds containing peptides or proteins
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Abstract
The invention discloses a probiotics activity retention method and application thereof to solid-state fatty food, and belongs to the technical field of food processing. According to the probiotics activity retention method, high-concentration glycerol is utilized for dehydration pretreatment of active probiotics by the osmotic equilibrium principle to achieve the purpose of reducing remaining moisture in probiotics. The obtained active bacterial sludge is added in the chocolate temperature-regulating technological process to form a favorable crystal structure together with chocolate fat, and time for probiotics to be active in chocolate is effectively prolonged, so that probiotic chocolate is obtained. The activity retention effect of probiotics in prepared chocolate is approximate to that of a product obtained through processing of freeze-dried bacterial powder. Through adoption of the probiotics activity retention method, a high cost caused by preparation of freeze-dried powder is avoided, energy consumption and equipment investment are low, and the processing cost can be greatly reduced.
Description
Technical field
The present invention relates to the activity retention method of a kind of probio and the application in Solid lipid food thereof, belong to food processing technology field.
Background technology
Probio is the active microorganism that a class is useful to host, can be colonizated in human body intestinal canal, reproductive system, and produces definite health efficacy, and then improves the active beneficial microorganism general name of host's microecological balance, performance beneficial effect.In recent years, along with people are to the rising increasingly of healthy attention rate, probio is subject to the extensive concern of numerous domestic and international scientific researcher, and it has to comprise and improves digestive system function, strengthens immunity, reduces blood fat and blood pressure, suppresses harmful microorganism, recovers many beneficial function such as liver function, even anti-malignant tumor.But the performance of probio beneficial function need be based upon the absorption of human body to a certain amount of active somatic cell number, and active bacteria can complete in the gastrointestinal tract and surely grows.Therefore, for the maintenance of probio growth activity in food processing process and storage thereof, become the great difficult problem that food industry and even sphere of learning face.
The maintenance of bacterial activity is the physicochemical property according to microorganism, the means such as physics, biology are used to create suitable condition artificially: to reduce the nutritional labeling in the water content of matrix, minimizing culture medium, reduce partial pressure of oxygen or adopt low temperature, suppress the respiration of microorganism, make its metabolism be in the state least active, growth and breeding is suppressed or is in geo-stationary.Namely the essential condition of microbial metabolism effectively can be reduced by low temperature, drying, anoxic three, make microorganism temporarily be in the state of semidormancy or the state of complete rest dormancy, thus make bacterial classification through original biological characteristics and good hereditary capacity still can be kept after preserving for a long time.
Ministry of Health of China was issued and was defended method prison and send out (2001) No. 84 prebiotic mushroom health foods evaluation regulation Article 10 and explicitly point out March calendar year 2001: do not advocate and produce prebiotic mushroom health food live products in liquid form.This is because liquid form product moisture activity is higher, and dissolved oxygen amount is relatively high, and for most of microbe, the optimum moisture activity of its growth is > 0.99.Growth, metabolism and breeding, finally cause the quick death of bacterial classification fast.For this reason, the producer adopts the dry means such as spraying dry, freeze drying usually, in conjunction with means such as embedding techniques, vacuum packaging and cryopreservations, with the growth activity keeping bacterial classification good.On the one hand, spraying dry and Freeze Drying Technique can effectively reduce residual moisture content in bacterial classification; On the other hand, high/low temperature and drying and dehydrating can trigger cell damages.Embedding techniques combines by albumen/polysaccharide or the mode such as outer layer covers, effectively reduces the external environment infringement suffered by probio, can effectively completely cut off bacterial classification with the direct of oxygen and contact, effective its growth activity of reduction in conjunction with vacuum packaging.But spray-dired processing temperature is higher, there is the drawbacks such as survival rate is lower in obtained probiotics bacterial powder; And although the probiotics bacterial powder adopting freeze drying to obtain possesses higher quality, due to the processing cost of its costliness, seriously hinder the Popularization And Development of probio.In addition, because dry processing technology limit, commercially available probio series products occurs there is product form more single, and generally have medicine external form, easily cause consumer to deposit the drawbacks such as mistaken ideas on concept mainly with forms such as dry powder state, Tablet and Capsulas on the one hand.On the other hand, research shows that the maintenance effect of existing bacterium powder dry technology to thalline vigor is poor, and usual cryopreservation is after 6 months, and bacterium powder vigor loss late is comparatively large, has a strong impact on the quality of product and prebiotic function.
In addition, the performance of probio functional activity not only needs the absorption of a certain amount of active thalline, also needs bacterial classification through digestions such as stomach and intestine simultaneously and finally realizes the long-term field planting in small intestine.Therefore, it is possible to resisting stronger sour environment and possessing good resistance to bile salt ability is the prerequisite that probio can survive and play a role in enteron aisle.But hydrochloric acid in gastric juice and bile salt all have stronger biocidal property, in vitro digestion result of the test shows, and probio is through acid and bile salt tolerance experimental test, and thalline survival rate is generally only about 40%, even lower.At present, the main embedding techniques that adopts keeps effect with the vigor improving probio, but the method is limited to embedding coating material and application vector thereof, make complicated, and effect is not remarkable.Therefore, the trend that solid-state probiotic products becomes new is developed.
Summary of the invention
The present invention provide firstly a kind of method keeping probiotic active, and by the active probiotic bacterium mud that described method prepares.
Described method, be by mass fraction be 70% ~ 90% glycerite and probiotics bacterial mud in mass ratio (5-20): 1 mixes, after pressure balanced to be infiltrated, collected by centrifugation active probiotic bacterium mud.
Described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium etc.
In one embodiment of the invention, be by 1 ~ 2 weight portion concentration 2 ~ 8 × 10
10the glycerine water solution that active bacteria mud and 10 ~ 20 weight portion mass fractions of cfu/g are 70% ~ 90% mixes, and leaves standstill 18 ~ 24h, then centrifugal dehydration, make gained active probiotic bacterium mud water content be 35 ~ 40% under low temperature.
In one embodiment of the invention, the peptide matters also containing 1 ~ 3% in described glycerine water solution.
In one embodiment of the invention, described method comprises the steps:
A, preparation activation probiotics bacterial liquid;
Described probiotics bacterial liquid can be formed by one or more probio mixing manufactures.
The step of described preparation activation probiotics bacterial liquid, comprise and draw probiotics bacterial liquid in glycerine guarantor tube, be inoculated in 800 ~ 1200mLMRS broth bouillon, then under temperature 36 ~ 38 DEG C, anaerobic condition, activation culture 16 ~ 18h is carried out, repeat above-mentioned activation act 2 ~ 4 times, obtain activating probiotics bacterial liquid.
Described MRS broth bouillon proportion of composing is as follows: 500mL distilled water, 5g peptone, 5g beef extract, 2.5g yeast extract, 2.5g sodium acetate, 10g glucose, 0.05gMgSO
4.7H
20,0.025gMnSO
4.H
20,1g dibasic ammonium citrate, 1gK
2hPO
4with 0.5ml tween, pH6.2 ~ 6.4;
B, the activation probiotics bacterial liquid high speed centrifugation, the separation under the condition of 6000 ~ 8000g that steps A are obtained, be separated the thalline sterile phosphate buffer obtained and wash 2 ~ 4 times, obtain concentration 2 ~ 8 × 10
10the active bacteria mud of cfu/g.
Described high speed centrifugation uses high-speed centrifuge to carry out centrifugation 10 ~ 15min.
The proportion of composing of described phosphate buffer is as follows: 0.2gKCl, 0.27gKH
2pO
4, 8.5gNaCl, 2.85gNa
2hPO
412H
2o and 1000mL distilled water, pH to 7.2.
The meaning of described cfu/g refers to the bacterial community sum contained in every gram of sample, and the mensuration of its bacterial community sum adopts conventional method well known to those skilled in the art of the present technique.
C, take 70 ~ 90 weight portion food grade glycerin, add in 10 ~ 30 parts by weight of deionized water, vibration mixing, obtained glycerine mixed solution;
Gained active bacteria mud in described step B is added in above-mentioned glycerine mixed solution (as above), and be placed in 4 DEG C of insulating box inner equilibrium 18 ~ 24h, ready to balance is complete, take out, by the glycerine mixed solution containing active bacteria mud high speed centrifugation, separation under the condition of 6000 ~ 8000g, obtain the active probiotic bacterium mud with low Residual water.
The active probiotic bacterium mud water content of described low Residual water is 35 ~ 40%.
Described Solid lipid system, primarily of the lipid composition deriving from animal or plant, lipid content >=70%, fusing point >=30 DEG C, under 35 DEG C ~ 45 DEG C conditions, have good rheological properties, when storing under 4 DEG C ~ 15 DEG C conditions, are generally solid-state.
The present invention also provides the application of described active probiotic bacterium mud in the prebiotic Solid lipid food of preparation, and such as chocolate, shortening, cheese etc., be the later stage at Solid lipid Product processing, add active probiotic bacterium mud and stir and evenly mix.
Understand the application of the method for the invention in Solid lipid food to be more convenient for, the present invention also provides described active probiotic bacterium mud preparing the application process in chocolate.
Described application process is in the process of processing dissimilar chocolate product, and described active probiotic bacterium mud is added in the production processing later stage and stirred and evenly mixed.With decline or the forfeiture of avoiding active probiotic can cause vigor because of production process, active probiotic can be protected further at follow-up storage.
In one embodiment of the invention, the temperature in described production processing later stage should be in 35 ~ 45 DEG C, product good fluidity or in viscous liquid.
In one embodiment of the invention, described application process comprises the following steps:
(1) take 70% ~ 75% cocoa liquor after high temperature melting, add 35% ~ 38% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 45 ~ 60 DEG C, refining 36 ~ 72h;
(3) the chocolate homogenate that completes of refining, to be cooled to 40 ~ 45 DEG C, described active probiotic bacterium mud is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases.The first stage of temperature adjustment, material is cooled to 29 DEG C from 40 DEG C; Second stage, is cooled to 27 DEG C from 29 DEG C; Phase III, go up to 29 ~ 30 DEG C from 27 DEG C
(4) moulding by casting: cool 25 ~ 30min, the demoulding under being placed in 8 ~ 10 DEG C of environment, packaging.
The present invention has following advantage:
1. preparation method is easy, and after long-time storage, activity of probiotic keeps excellent:
The present invention utilizes high concentration glycerine to carry out dehydration pretreatment by osmotic balance principle to active probiotic, to reach the object reducing Residual water contained by probio inside.Somatic cells is after the pretreatment of high concentration glycerine, and ICW viscosity significantly promotes, and water activity declines, and phage surface can be wrapped to form good " embedding " drop by glycerine, forms good air barrier, effectively avoids thalline because of oxidative death.Active bacteria mud is after treatment applied in Solid lipid food, and add in Solid lipid food processing later stage tempering process process, itself and lipid can be made to form good crystal structure, the vigor of effective prolongation probio in Solid lipid, and vigor keeps effect to be better than commercially available freeze-dried vaccine powder class probiotic products, avoids the great number cost prepared freeze-dried powder and produce simultaneously.In addition, glycerine, as a kind of emulsifying agent, contributes to the Homogeneous phase mixing of lipid and thalline drop.
2. in vitro digestion test result is more excellent, contributes to the field planting of probio enteron aisle and the performance of functional characteristic
Adopt the method for the invention to obtain product, find through in vitro digestion test, itself and existing probio series products process technology (dry bacterium powder, embedding techniques etc.) are compared, and active thalline survival rate obviously promotes.This is mainly due in Solid lipid system of the present invention, probio thalline exists mainly with " oily bag bacterium " form, lipid layer is thicker, and acid and bile salt tolerance ability is strong, therefore, active thalline still has higher survival rate after enteron aisle digestion process, contributes to the field planting of probio enteron aisle and the performance of functional characteristic.
3. materials safety is reliable, low processing cost, and equipment investment is little:
The present invention adopts glycerine as osmotic pressure water balance raw material, and it is put in the food additives list that can use in right amount by need of production in varieties of food items in China GB2760-2014 food additives use standard, therefore, does not have food security hidden danger.Bacterium mud preprocess method is easy, and energy consumption is low, and equipment investment is little, greatly can save processing cost.
In sum, the present invention effectively can keep probiotic active, reduce the processing and fabricating cost of probio series products, and method is easy, and after long-time storage, in product, contained probio still has good growth activity, nutrient health.
Accompanying drawing explanation
Fig. 1. the maintenance of the chocolate thalline vigor of probio is with the change of storage time
Detailed description of the invention
In order to better set forth the advantage that the method for the invention is described, the present invention has also carried out related property test to active probiotic and the chocolate containing active probiotic, specific as follows:
According to GB/T5497-1985 method of testing, carry out measuring comparing to moisture in probiotics bacterial mud;
According to SN/T0168-2015 method of testing, carry out measuring comparing to probiotics viable bacteria sum contained in the probio chocolate of long-time storage;
Activity of probiotic keeps storage experiment: store 6 months under obtained probio chocolate being placed in 4 DEG C of environment, and regularly carries out test and comparison to probio content contained in product.
In vitro digestion is tested: the testing sample solid getting suitable quality, add simulated gastric fluid or the simulated intestinal fluid of certain volume, control group is set simultaneously, isopyknic PBS is added in control group, 2h is cultivated in 37 DEG C of water-baths, put upside down every 30min and shake up 1 time, after cultivating 2h, adopt gradient dilution method to carry out plate count.
Embodiment 1 is clay that probio is chocolate without glycerine pretreated probiotic active bacterium
A, to protect in tube from glycerine and draw 200 μ L probiotics bacterial liquids, be inoculated in 1000mLMRS broth bouillon, then under temperature 37 DEG C, anaerobic condition, carry out activation culture 17h, repeat above-mentioned activation act 2 times, obtain a kind of activation probiotics bacterial liquid;
Described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
Described probiotics bacterial liquid can be formed by one or more probio mixing manufactures above-mentioned.
Described MRS broth bouillon is composed as follows: 500mL distilled water, 5g peptone, 5g beef extract, 2.5g yeast extract, 2.5g sodium acetate, 10g glucose, 0.05gMgSO
47H
20,0.025gMnSO
4h
20,1g dibasic ammonium citrate, 1gK
2hPO
4with 0.5ml tween, pH6.2;
B, the activation bacterium liquid high speed centrifugation, the separation under the condition of 6500g that steps A are obtained, be separated the thalline sterile phosphate buffer obtained and wash 3 times, so obtain concentration 7.3 × 10
10cfu/g active bacteria mud.
In this step B, described activation bacterium liquid uses high-speed centrifuge to carry out centrifugation 10min.
Described phosphate buffer composed as follows: 0.2gKCl, 0.27gKH
2pO
4, 8.5gNaCl, 2.85gNa
2hPO
412H
2o and 1000mL distilled water, pH to 7.2.
The meaning of described cfu/g refers to the bacterial community sum contained in every gram of sample, and in employing the art of its bacterial community sum, conventional method known by the technical staff measures.
Described active probiotic bacterium mud water content is 80.92%.
C, preparation chocolate
(1) take 70% cocoa liquor after high temperature melting, add 36% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 50 DEG C, refining 40h;
(3) the chocolate homogenate that completes of refining, to be cooled to 40 DEG C, active probiotic bacterium mud in the method for the invention step B is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases.The first stage of temperature adjustment, material is cooled to 29 DEG C from 40 DEG C; Second stage, is cooled to 27 DEG C from 29 DEG C; Phase III, go up to 29 DEG C from 27 DEG C;
(4) moulding by casting, cools 25min, the demoulding under being placed in 8 DEG C of environment, packaging.
Embodiment 2 freeze-dried vaccine powder obtains probio chocolate
A, to protect in tube from glycerine and draw 200 μ L probiotics bacterial liquids, be inoculated in 1000mLMRS broth bouillon, then under temperature 37 DEG C, anaerobic condition, carry out activation culture 17h, repeat above-mentioned activation act 2 times, obtain a kind of activation probiotics bacterial liquid;
Described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
Described probiotics bacterial liquid can be formed by one or more probio mixing manufactures above-mentioned.
Described MRS broth bouillon is composed as follows: 500mL distilled water, 5g peptone, 5g beef extract, 2.5g yeast extract, 2.5g sodium acetate, 10g glucose, 0.05gMgSO
47H
20,0.025gMnSO
4h
20,1g dibasic ammonium citrate, 1gK
2hPO
4with 0.5ml tween, pH6.2;
B, the activation bacterium liquid high speed centrifugation, the separation under the condition of 6500g that steps A are obtained, be separated the thalline sterile phosphate buffer obtained and wash 3 times, so obtain concentration 7.3 × 10
10cfu/g active bacteria mud.
In this step B, described activation bacterium liquid uses high-speed centrifuge to carry out centrifugation 10min.
Described phosphate buffer composed as follows: 0.2gKCl, 0.27gKH
2pO
4, 8.5gNaCl, 2.85gNa
2hPO
412H
2o and 1000mL distilled water, pH to 7.2.
The meaning of described cfu/g refers to the bacterial community sum contained in every gram of sample, and in employing the art of its bacterial community sum, conventional method known by the technical staff measures.
The making of C, probio freeze-dried vaccine powder
By the mud of active bacteria described in step B, add 30 weight portion skimmed milk solution, mix, then through freeze drying, obtain high-activity probiotics freeze-dried vaccine powder, for subsequent use, simultaneously for storing the mensuration of experiment viable count.
Described probio freeze-dried vaccine powder total plate count is 8.9 × 10
9cfu/g;
Described probio freeze-dried vaccine powder moisture is 8.21%.
D, preparation chocolate
(1) take 70% cocoa liquor after high temperature melting, add 36% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 50 DEG C, refining 40h;
(3) the chocolate homogenate that completes of refining, to be cooled to 40 DEG C, active probiotic freeze-dried vaccine powder in the method for the invention step C is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases.The first stage of temperature adjustment, material is cooled to 29 DEG C from 40 DEG C; Second stage, is cooled to 27 DEG C from 29 DEG C; Phase III, go up to 29 DEG C from 27 DEG C;
(4) moulding by casting, cools 25min, the demoulding under being placed in 8 DEG C of environment, packaging.
The active probiotic that embodiment 3 the inventive method obtains is chocolate for the preparation of probio
A, to protect in tube from glycerine and draw 200 μ L probiotics bacterial liquids, be inoculated in 1000mLMRS broth bouillon, then under temperature 37 DEG C, anaerobic condition, carry out activation culture 17h, repeat above-mentioned activation act 2 times, obtain a kind of activation probiotics bacterial liquid;
Described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
Described probiotics bacterial liquid can be formed by one or more probio mixing manufactures above-mentioned.
Described MRS broth bouillon is composed as follows: 500mL distilled water, 5g peptone, 5g beef extract, 2.5g yeast extract, 2.5g sodium acetate, 10g glucose, 0.05gMgSO
47H
20,0.025gMnSO
4h
20,1g dibasic ammonium citrate, 1gK
2hPO
4with 0.5ml tween, pH6.2;
B, the activation bacterium liquid high speed centrifugation, the separation under the condition of 6500g that steps A are obtained, be separated the thalline sterile phosphate buffer obtained and wash 3 times, so obtain concentration 7.3 × 10
10cfu/g active bacteria mud.
In this step B, described activation bacterium liquid uses high-speed centrifuge to carry out centrifugation 10min.
Described phosphate buffer composed as follows: 0.2gKCl, 0.27gKH
2pO
4, 8.5gNaCl, 2.85gNa
2hPO
412H
2o and 1000mL distilled water, pH to 7.2.
The meaning of described cfu/g refers to the bacterial community sum contained in every gram of sample, and in employing the art of its bacterial community sum, conventional method known by the technical staff measures.
C, take 90 weight portion food grade glycerin, add in 10 parts by weight of deionized water, vibration mixing, obtained glycerine mixed solution;
Gained active bacteria mud in described step B is added in above-mentioned glycerine mixed solution, and be placed in 4 DEG C of insulating box inner equilibrium 20h, ready to balance is complete, take out, by the glycerine mixed solution containing active bacteria mud high speed centrifugation, separation under the condition of 6500g, obtain the active probiotic bacterium mud with low Residual water, for the processed and applied in chocolate;
The active probiotic bacterium mud water content of described low Residual water is 35.67%.
D, preparation chocolate
(1) take 70% cocoa liquor after high temperature melting, add 36% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 50 DEG C, refining 40h;
(3) the chocolate homogenate that completes of refining, to be cooled to 40 DEG C, the active probiotic bacterium mud of Residual water low in the method for the invention step C is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases.The first stage of temperature adjustment, material is cooled to 29 DEG C from 40 DEG C; Second stage, is cooled to 27 DEG C from 29 DEG C; Phase III, go up to 29 DEG C from 27 DEG C;
(4) moulding by casting, cools 25min, the demoulding under being placed in 8 DEG C of environment, packaging.
The active probiotic that embodiment 4 the inventive method obtains is chocolate for the preparation of probio
A, to protect in tube from glycerine and draw 200 μ L probiotics bacterial liquids, be inoculated in 1200mLMRS broth bouillon, then under temperature 38 DEG C, anaerobic condition, carry out activation culture 18h, repeat above-mentioned activation act 2 times, obtain a kind of activation probiotics bacterial liquid;
Described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
Described probiotics bacterial liquid can be formed by one or more probio mixing manufactures above-mentioned.
Described MRS broth bouillon is composed as follows: 500mL distilled water, 5g peptone, 5g beef extract, 2.5g yeast extract, 2.5g sodium acetate, 10g glucose, 0.05gMgSO
47H
20,0.025gMnSO
4h
20,1g dibasic ammonium citrate, 1gK
2hPO
4with 0.5ml tween, pH6.3;
B, the activation bacterium liquid high speed centrifugation, the separation under the condition of 8000g that steps A are obtained, be separated the thalline sterile phosphate buffer obtained and wash 4 times, so obtain concentration 6.3 × 10
10cfu/g active bacteria mud.
In this step B, described activation bacterium liquid uses high-speed centrifuge to carry out centrifugation 15min.
Described phosphate buffer composed as follows: 0.2gKCl, 0.27gKH
2pO
4, 8.5gNaCl, 2.85gNa
2hPO
412H
2o and 1000mL distilled water, pH to 7.2.
The meaning of described cfu/g refers to the bacterial community sum contained in every gram of sample, and in employing the art of its bacterial community sum, conventional method known by the technical staff measures.
C, take 75 weight portion food grade glycerin and 1.6 weight portion peptide matters, add in 25 parts by weight of deionized water, vibration mixing, obtained glycerine mixed solution;
Gained active bacteria mud in described step B is added in above-mentioned glycerine mixed solution, and be placed in 4 DEG C of insulating box inner equilibrium 24h, ready to balance is complete, take out, by the glycerine mixed solution containing active bacteria mud high speed centrifugation, separation under the condition of 8000g, obtain the active probiotic bacterium mud with low Residual water, for the processed and applied in chocolate;
The active probiotic bacterium mud water content of described low Residual water is 36.06%.
D, preparation chocolate
(1) take 75% cocoa liquor after high temperature melting, add 38% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 45 DEG C, refining 60h;
(3) the chocolate homogenate that completes of refining, to be cooled to 45 DEG C, the active probiotic bacterium mud of Residual water low in the method for the invention step C is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases.The first stage of temperature adjustment, material is cooled to 29 DEG C from 40 DEG C; Second stage, is cooled to 27 DEG C from 29 DEG C; Phase III, go up to 30 DEG C from 27 DEG C;
(4) moulding by casting, cools 30min, the demoulding under being placed in 10 DEG C of environment, packaging.
The comparison of probio moisture under table 1. different disposal condition
The pretreatment of table 2. glycerine balance is on the impact of growth of probiotics activity
The external tolerance experimental results of table 3 in simulated gastric fluid and simulated intestinal fluid
As can be seen from Figure 1, the chocolate utilizing probio obtained is after long-time storage is placed, adopt fresh bacterium clay chocolate in total amount of probiotics with the prolongation of storage time, decline more obvious, and adopt the chocolate that the inventive method and freeze-dried vaccine powder obtain, in it, contained total amount of probiotics does not decline along with the prolongation of storage time significantly, and total plate count tends towards stability after 30 days.In addition, embodiment 3,4 gained chocolate flavouring is not subject to the impact of glycerine.
Although the present invention with preferred embodiment openly as above; but it is also not used to limit the present invention, any person skilled in the art, without departing from the spirit and scope of the present invention; all can do various changes and modification, what therefore protection scope of the present invention should define with claims is as the criterion.
Claims (10)
1. preparing a method for prebiotic Solid lipid product, it is characterized in that, is the later stage at Solid lipid Product processing, adds active probiotic bacterium mud and stirs and evenly mixs; Described active probiotic bacterium mud is glycerite by being 70% ~ 90% and concentration by mass fraction is 2 ~ 8 × 10
10the probiotics bacterial mud of cfu/g is in mass ratio (5-20): 1 mixing, after pressure balanced to be infiltrated, and the bacterium mud that collected by centrifugation obtains.
2. method according to claim 1, is characterized in that, the temperature in the later stage of described Solid lipid Product processing is in 35 ~ 45 DEG C, product good fluidity or in viscous liquid.
3. method according to claim 1, is characterized in that, described Solid lipid product is chocolate, shortening or cheese.
4. method according to claim 1, it is characterized in that, described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
5., according to the arbitrary described method of claim 1-4, it is characterized in that, described active probiotic bacterium mud is by 1 ~ 2 weight portion concentration 2 ~ 8 × 10
10the glycerine water solution that active bacteria mud and 10 ~ 20 weight portion mass fractions of cfu/g are 70% ~ 90% mixes, under low temperature, leave standstill 18 ~ 24h, and then bacterium mud is collected in centrifugal dehydration; Described active probiotic bacterium mud water content is 35 ~ 40%.
6. method according to claim 3, is characterized in that, comprises the following steps:
(1) take 70% ~ 75% cocoa liquor after high temperature melting, add 35% ~ 38% white sugar powder, mix, obtained chocolate homogenate, for subsequent use;
(2) by above-mentioned chocolate homogenate, under being placed in the temperature environment of 45 ~ 60 DEG C, refining 36 ~ 72h;
(3) the chocolate homogenate that completes of refining, to be cooled to 40 ~ 45 DEG C, described active probiotic bacterium mud is added in chocolate homogenate, and completes the temperature adjustment flow process of chocolate three phases; Described temperature adjustment flow process comprises: the first stage of temperature adjustment, and material is cooled to 29 DEG C from 40 DEG C, second stage, is cooled to 27 DEG C, the phase III from 29 DEG C, gos up to 29 ~ 30 DEG C from 27 DEG C;
(4) moulding by casting: cool 25 ~ 30min, the demoulding under being placed in 8 ~ 10 DEG C of environment, packaging, obtains prebiotic chocolate.
7. method according to claim 1, is characterized in that, the peptide matters also containing 1 ~ 3% in described glycerite.
8. the prebiotic Solid lipid product that method prepares according to claim 1 or 2 or 3 or 4 or 6 or 7.
9. keep a method for probiotic active, it is characterized in that, be by mass fraction be 70% ~ 90% glycerite and concentration be 2 ~ 8 × 10
10the probiotics bacterial mud of cfu/g is in mass ratio (5-20): 1 mixing, leaves standstill 18 ~ 24h, pressure balanced to be infiltrated, collected by centrifugation active probiotic bacterium mud under low temperature, described active probiotic bacterium mud water content is 35 ~ 40%, described probio comprises lactobacillus fermenti, lactobacillus cellobiosas, lactobacillus curvatus, lactobacillus lactis, lactobacillus plantarum, lactobacillus reuteri, lactobacillus acidophilus, Lactobacillus brevis, lactobacillus bulgaricus, Lactobacillus casei, the bright beading coccus of mesenterium, pediococcus cerevisiae, Pediococcus pentosaceus, lactic acid bacteria tablet coccus, streptococcus lactis, streptococcus lactis, streptococcus thermophilus, bifidobacterium adolescentis, animal bifidobacteria, bifidobacterium infantis, bifidobacterium longum, thermophilic Bifidobacterium.
10. the active probiotic bacterium mud for preparing of method according to claim 9.
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