CN110122576A - It is a kind of the method for fermented soybean milk to be prepared by lactobacillus fermenti and the fermented soybean milk prepared and application - Google Patents
It is a kind of the method for fermented soybean milk to be prepared by lactobacillus fermenti and the fermented soybean milk prepared and application Download PDFInfo
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- CN110122576A CN110122576A CN201910435065.1A CN201910435065A CN110122576A CN 110122576 A CN110122576 A CN 110122576A CN 201910435065 A CN201910435065 A CN 201910435065A CN 110122576 A CN110122576 A CN 110122576A
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- 239000001963 growth medium Substances 0.000 abstract description 6
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Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C11/00—Milk substitutes, e.g. coffee whitener compositions
- A23C11/02—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins
- A23C11/10—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins containing or not lactose but no other milk components as source of fats, carbohydrates or proteins
- A23C11/103—Milk substitutes, e.g. coffee whitener compositions containing at least one non-milk component as source of fats or proteins containing or not lactose but no other milk components as source of fats, carbohydrates or proteins containing only proteins from pulses, oilseeds or nuts, e.g. nut milk
- A23C11/106—Addition of, or treatment with, microorganisms
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/143—Fermentum
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/169—Plantarum
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Microbiology (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Beans For Foods Or Fodder (AREA)
- Dairy Products (AREA)
Abstract
The invention particularly discloses a kind of by lactobacillus fermenti prepares the method for fermented soybean milk and the fermented soybean milk prepared and application, carries out fermentating metabolism method includes the following steps: lactobacillus fermenti CGMCC No.17321 is inoculated in soya-bean milk, obtains fermented soybean milk；Present invention has the advantage that by lactobacillus fermenti CGMCC No.17321/ lactobacillus fermenti CGMCC No.17321 and lactobacillus plantarum (L.plantarum) ATCC 14917, (purchase is fermented from ATCC) using soya-bean milk as culture medium, it is prepared with streptococcus mutans biofilm inhibitor activity fermented soybean milk, it can be compared with the bacterial strain and other fermentation mediums of fermented soybean milk with other, gained fermented soybean milk is more significant to the inhibitory effect of streptococcus mutans biofilm formation, and good stability.
Description
Technical field
The invention belongs to field of biotechnology, and in particular to a kind of preparation method of fermented soybean milk.In addition, the present invention also relates to
And a kind of fermented soybean milk and application.
Background technique
Saprodontia is commonly called as decayed tooth, decayed tooth, is a kind of bacteriosis, can be with secondary pulpitis and periapical inflammation, or even can draw
Play alveolar bone and inflammation of jaws.Such as it is not treated in time, lesion continues to develop, and cavity is formed, destroys disappearance completely to corona eventually,
The final result of development is loss of tooth.Saprodontia feature is disease incidence height, and distribution is wide.The disease is the main common disease in oral cavity,
It is one of most common disease of the mankind, it is listed as the big emphasis of the mankind three with tumour and cardiovascular disease by the World Health Organization
Disease preventing and treating.
The factor for causing saprodontia is numerous, such as bacterium, oral environment (food, saliva), host, wherein bacterium is saprodontia
Necessary condition occurs, it is considered that there are two types of types for cariogenic bacteria, and one is acid-producing bacteria categories, wherein predominantly streptococcus mutans
(Streptococcus mutans, abbreviation S.mutans), actinomyces and lactobacillus can make carbohydrate breakdown produce acid,
Lead to tooth inanimate matter demineralization；Another kind is gram-positive cocci, can destruction of organic material, tooth can be made to be formed by long term
Cavity.In above-mentioned cariogenic bacteria, streptococcus mutans are known as being the main and most important disease for causing saprodontia by domestic and foreign scholars
Opportunistic pathogen.However these above-mentioned pathogens be not directly be colonized in dental surface, but by with caused by streptococcus mutans
After biomembrane (insoluble polysaccharide) combination forms plaque, it is adhered to dental surface indirectly, as cariogenic bacteria is constantly in biology
The degree of accumulation on film, saprodontia runs down.Therefore, the yield for reducing its biomembrane also can reach the purpose of prevention and treatment saprodontia.
Currently, overwhelming majority report concentrates on control or reduces streptococcus mutans in oral cavity in the research in relation to dental caries therapeutic
Quantity, however, the oral environment of health is typically at harmful bacteria and beneficial bacterium reaches the state of balance, harmful bacteria is excessive
Reduction can bring oral cavity flora unbalance, and then cause other adverse reactions of human body.In addition, what is clinically used is some for change
The streptococcic germ killing drugs of shape, such as ammoniacal silver nitrate compound, side effect is strong, and long-time service can enable dental stain, therefore not applicable
It is treated in labial teeth.And saprodontia risk can be equally reduced by reducing streptococcus mutans biomembrane yield, and do not destroy mouth
The inherent balance of chamber flora generates side effect, but the report of this respect is relatively fewer, especially has with microbial fermentation product
The research of pass is extremely limited, has and inhibits the fermented product of streptococcus mutans biofilm formation quite deficient.
Therefore it provides the fermented product with inhibition streptococcus mutans biofilm formation is the task of top priority.This is this field
Technical staff's urgent problem to be solved.
Summary of the invention
Based on above-mentioned technical problem, the present invention provides a kind of preparation methods of fermented soybean milk, can be made using this method
There must be the fermented soybean milk for inhibiting streptococcus mutans biofilm formation.
Specifically, on the one hand, provide a kind of preparation method of fermented soybean milk by lactobacillus fermenti preparation, including following
Step: lactobacillus fermenti CGMCC No.17321 is inoculated in soya-bean milk as fermenting agent and carries out fermentating metabolism, is fermented
Soya-bean milk.
Through the above technical solutions, the application uses lactobacillus fermenti (Lactobacillus fermentum) for the first time
CGMCC No.17321, ferments using soya-bean milk as culture medium, is prepared with streptococcus mutans biofilm inhibitor activity
Fermented soybean milk, that is, the fermented soybean milk for disclosing lactobacillus fermenti (L.fermentum) CGMCC No.17321 fermentating metabolism have
The new application of anti-caries tooth.
In addition, above-mentioned lactobacillus fermenti (L.fermentum) CGMCC No.17321 bacterial strain is protected on March 8th, 2019
Ensconce China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC), it is proposed that classification naming: acidified milk bar
Bacterium Lactobacillus fermentum, preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, postcode: 100101.
Preferably, the inoculum concentration of lactobacillus fermenti CGMCC No.17321 is 2x107~1x108cfu/mL。
Through the above technical solutions, the inoculum concentration as lactobacillus fermenti (L.fermentum) CGMCC No.17321 is
2x107~1x108When cfu/mL, can be had by fermented soybean milk prepared by lactobacillus fermenti (L.fermentum) CGMCC No.17321
Effect inhibits the forming amount of streptococcus mutans biomembrane；And when inoculum concentration is 4x107~8x107Cfu/mL, inhibitory effect achieved
Preferably；When inoculum concentration is 6x107Cfu/mL, inhibitory effect achieved are best.
Further, the fermenting agent be inoculative proportion 1~9:1~9 lactobacillus fermenti CGMCC No.17321 and
Lactobacillus plantarum (L.plantarum) ATCC 14917 (buys the composite fermentation microbial inoculum from ATCC).
And the application is for the first time by lactobacillus fermenti (L.fermentum) CGMCC No.17321 and lactobacillus plantarum
(L.plantarum) (purchase is inoculated in soya-bean milk ATCC 14917 from ATCC), is prepared with streptococcus mutans biomembrane
Inhibitory activity fermented soybean milk is disclosed by lactobacillus fermenti (L.fermentum) CGMCC No.17321 and lactobacillus plantarum
(L.plantarum) ATCC 14917 (buys the new application for having anti-caries tooth from the fermented soybean milk of ATCC) fermentating metabolism.
Further, the soya-bean milk be soybean, red bean, mung bean, semen viciae fabae, big white kidney bean, small white kidney bean it is one or more.
Through the above technical solutions, the effect of soya-bean milk is made in soybean, red bean, mung bean, semen viciae fabae, big white kidney bean, small white kidney bean
During fruit compares, wherein the soya-bean milk effect of soybean preparation is best.
Further, the solid quality degree in the soya-bean milk is 1%~9%.
Through the above technical solutions, when solid quality degree 1%~9% in soya-bean milk, gained fermented soybean milk
The inhibitory effect having had to streptococcus mutans biofilm formation；It is right when solid quality degree is 3%~7%
Streptococcus mutans biofilm formation has preferable inhibitory effect, when solid quality degree is 5%, to deformation chain
Coccus biofilm formation has better inhibitory effect；Wherein principle is the biomembrane of solid content and final products in soya-bean milk
Inhibitory effect has direct relation, when solid content is excessively high, thallus habitat osmotic pressure can be caused to increase, influence the growth of thallus
And reduce suppression membrane substance synthetic quantity, in addition production cost can also rise, and solid content is too low then can be because of nutritional ingredient not
Foot causes viable bacteria cultivation effect poor, equally will affect the generation of suppression membrane substance.
Further, the temperature of the fermentation is 15 DEG C~45 DEG C, and the time of fermentation is 2~48h.
Through the above technical solutions, when the time of fermentation is 2~48h, being inoculated with when the temperature of fermentation is 15 DEG C~45 DEG C
In the case that amount, soymilk concentration, cultivation temperature and fermentation time influence each other, gained fermented soybean milk is to streptococcus mutans biomembrane
Form the inhibitory effect having had；And the temperature preferably fermented is 25 DEG C~40 DEG C, the temperature more preferably fermented is 37 DEG C；Compared with
Good ground fermentation time is 12~36h；More preferably fermentation time is for 24 hours.
Further, the fermentation is stationary culture.
Through the above technical solutions, lactobacillus fermenti CGMCC No.17321 belongs to facultative anaerobic bacteria, stationary culture has
The fast feature of reproduction speed.
Second aspect additionally provides a kind of fermented soybean milk, is made by any of the above-described kind of preparation method.
Further, inhibiting rate >=41% of the above-mentioned fermented soybean milk to streptococcus mutans biofilm formation.
The third aspect additionally provides fermented soybean milk in the application of preparation anti-caries tooth food.
Compared with prior art, the application uses lactobacillus fermenti (L.fermentum) CGMCC No.17321/ to send out for the first time
(purchase is certainly by kefir milk bacillus (L.fermentum) CGMCC No.17321 and lactobacillus plantarum (L.plantarum) ATCC 14917
ATCC), ferment, be prepared with streptococcus mutans biofilm inhibitor activity fermented soybean milk using soya-bean milk as culture medium,
Disclose lactobacillus fermenti (L.fermentum) CGMCC No.17321/ lactobacillus fermenti (L.fermentum) CGMCC
No.17321 and lactobacillus plantarum (L.plantarum) ATCC 14917 (buy the metabolite from ATCC) gained fermented soybean milk
New application with anti-caries tooth；Can be compared with the bacterial strain and other fermentation mediums of fermented soybean milk with other, lactobacillus fermenti
(L.fermentum) CGMCC No.17321/ lactobacillus fermenti (L.fermentum) CGMCC No.17321 and lactobacillus plantarum
(L.plantarum) ATCC 14917 (buys from ATCC) gained fermented soybean milk and imitates to the inhibition of streptococcus mutans biofilm formation
Fruit is more significant, and good stability.
Specific embodiment
Technical solution of the present invention is illustrated to be clearer, below in conjunction with specific embodiment to skill of the invention
Art scheme is further elaborated:
In a specific embodiment, a kind of preparation method of fermented soybean milk is provided, comprising the following steps: will send out
Kefir milk bacillus CGMCC No.17321, which is inoculated in soya-bean milk, carries out fermentating metabolism, obtains fermented soybean milk.
Preparation method in above-mentioned technical proposal, the application use lactobacillus fermenti (L.fermentum) CGMCC for the first time
No.17321 ferments using soya-bean milk as culture medium, is prepared with streptococcus mutans biofilm inhibitor activity fermentation beans
Slurry discloses the new application that lactobacillus fermenti (L.fermentum) CGMCC No.17321 fermented soybean milk has anti-caries tooth.
In addition, bacterial strain used by preparation method in above-mentioned technical proposal is edible natural bacterial strain, the fermentation of use
Culture medium (all kinds of soya-bean milk) from a wealth of sources, low in cost, natural safety improves food while reducing Material Cost
Safety.
Further, in above-mentioned steps, the inoculum concentration of lactobacillus fermenti (L.fermentum) CGMCC No.17321 is
2x107~1x108cfu/mL；Preferably 4x107~8x107Cfu/mL is more preferably 6x107cfu/mL。
Further, in above-mentioned steps, preferred soya-bean milk is soybean, red bean, mung bean, semen viciae fabae, big white kidney bean, little Bai rue
One or more, the more preferably soybean milk of beans.
Further, in above-mentioned steps, preferred soya-bean milk solid quality degree is 1%~9%；Preferably
It is 3%~7%；It is more preferably 5%.
Further, in above-mentioned steps, preferred fermentation temperature is 15 DEG C~45 DEG C；Preferably 25 DEG C~40 DEG C；
It is more preferably 37 DEG C.
Further, in above-mentioned steps, preferred fermentation time is 2~48h；Preferably 12~36h；More preferably it is
24h。
Further, in above-mentioned steps, preferred fermentation method is stationary culture.
In another particular embodiment of the invention, a kind of fermented soybean milk is provided, which is sent out by any of the above-described kind
The preparation method of ferment soya-bean milk is made.
Since the preparation method of fermented soybean milk has above-mentioned beneficial effect, the fermented soybean milk as made from the preparation method
With corresponding beneficial effect, details are not described herein again.
Further, inhibiting rate >=41% of the above-mentioned fermented soybean milk to streptococcus mutans biofilm formation.In conjunction with the embodiments
Also it is found that lactobacillus fermenti (L.fermentum) CGMCC No.17321 fermented soybean milk is to streptococcus mutans biofilm formation
Inhibitory effect be significantly higher than other can fermented soybean milk bacterial strain, lactobacillus fermenti (L.fermentum) CGMCC No.17321 and
(purchase is from ATCC) fermented soybean milk to streptococcus mutans biofilm formation by lactobacillus plantarum (L.plantarum) ATCC 14917
Inhibitory effect is apparently higher than lactobacillus fermenti (L.fermentum) CGMCC No.17321 fermented soybean milk.
Above-mentioned specific embodiment is further illustrated below by embodiment, but is not therefore limited the present invention to described
Scope of embodiments among.In the following examples, the experimental methods for specific conditions are not specified, according to conventional methods and conditions, or presses
It is selected according to product manual.
In following embodiments, all raw materials are commercially available, and meet relevant national standard.
Embodiment 1
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe): by lactobacillus fermenti (Lactobacillus
Fermentum) freeze-dried powder of CGMCC No.17321 is dissolved with a small amount of sterile distilled water, takes a ring to line MRS with oese
On solid medium (Merck Co. Germany), 37 DEG C of Anaerobic culturels take out for 24 hours, are put into 1mL MRS with oese picking single colonie
Bacterium colony, is dispersed in fluid nutrient medium, 37 DEG C of Anaerobic culturels by liquid (Merck Co. Germany) with vortex oscillator
It takes out for 24 hours, 50mL MRS liquid is inoculated in 2% (v/v) inoculum concentration, after 37 DEG C of cultures for 24 hours, culture 9,000rpm centrifugation 10
Minute, it discards supernatant, after thallus washs 2 times with sterile distilled water, is suspended, fermented with the sterile distilled water of former volume of culture
Seed, the bacteria concentration of seed liquor are 2x109cfu/mL。
The preparation of streptococcus mutans bacteria suspension: by streptococcus mutans CGMCC No.1.2499's (being purchased from CGMCC, China)
Freeze-dried powder is dissolved with a small amount of sterile distilled water, with oese take a ring line BHI solid medium (purchase from OXOID Co.,
Britain), 37 DEG C of Anaerobic culturels take out for 24 hours, and being put into 10mL BHI fluid nutrient medium with oese picking single colonie, (purchase is certainly
OXOID Co., Britain), bacterium colony is dispersed in fluid nutrient medium with turbula shaker, 37 DEG C of Anaerobic culturels take for 24 hours
Out, BHI fluid nutrient medium is inoculated in 2% (v/v) inoculum concentration, 37 DEG C of Anaerobic culturels for 24 hours after, culture 15000rpm centrifugation
10min is abandoned supernatant, after thallus washs 2 times with sterile distilled water, is resuspended with BHI fluid nutrient medium to 106CFU/mL.
(b) preparation of soya-bean milk: after the weighing of big dried bean curd beans, the water that five times of quality are added is placed in 37 DEG C of immersion 8h, discards water,
A certain proportion of wet beans and water are added in soy bean milk making machine, the soya-bean milk of solid content needed for being made is obtained through 121 DEG C of sterilizing 20min
To the sterile soybean milk of specified solid content.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation:
The pretreatment of fermented soybean milk: fermented soybean milk pH is adjusted to 6.80 to get sample to be tested.
The measurement of inhibitory activity: after sterile magnetic stir bar A (5 × 10mm) is immersed in 0.22 μm of filter membrane aseptic filtration
Healthy population saliva in 37 DEG C of incubation 4h, pick up magnetic stir bar A with sterile magnetic stir bar B (10 × 50mm), with
The PBS of 10mM, pH7.0 are eluted, and are removed magnetic stir bar A, are placed in the culture hole of 24 orifice plates.1.6mL is added into culture hole
BHI fluid nutrient medium, 200 μ L streptococcus mutans bacteria suspensions (106CFU/mL) and 400 μ L testing sample solutions or blank control it is molten
Liquid (unfermentable soya-bean milk), 37 DEG C of anaerobism are incubated for (purchase is German from Greiner company) for 24 hours again.It is inhaled with magnetic stir bar B
Magnetizing force stirrer A after being eluted with PBS, is removed magnetic stir bar A, is placed in the culture hole of 24 orifice plates, natural air drying.Xiang Pei
The crystal violet solution dyeing 15min that 1.6mL, 0.1% (w/v) are added in hole is supported, picks up magnetic stir bar A with magnetic stir bar B,
Extra dyeing liquor is removed with PBS elution, magnetic stir bar A is removed, is placed in the culture hole of 24 orifice plates, natural air drying.Xiang Pei
It supports and 300 μ L dehydrated alcohols decoloration 15min is added in hole, 200 μ L is taken to be transferred to 96 orifice plates (purchase is German from Greiner company)
It is interior, OD600nm, as quantized data corresponding to streptococcus mutans biofilm formation amount are measured with microplate reader.In the above way
The OD600nm of blank control and sample to be tested is measured, and calculates the inhibiting rate of bioflm inhibiting agents.Biomembrane inhibiting rate (I):
I=(OD600nm (blank control)-OD600nm (sample to be tested))/OD600nm (blank control) × 100%
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with inoculum concentration 3% (v/v, kind
Sub- liquid accounts for the percent by volume of fermentation liquid, similarly hereinafter) in solid content 5%, (w/w, soybean account for the quality percentage of water to aseptic inoculation
Than similarly hereinafter) soybean milk in, 37 DEG C of stationary cultures for 24 hours fermented soybean milk A.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk A is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=45% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 2
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of soya-bean milk: after the dry beans weighing of red bean, the water that five times of quality are added is placed in 37 DEG C of immersion 8h, discards water,
A certain proportion of wet beans and water are added in soy bean milk making machine, the soya-bean milk of solid content needed for being made is obtained through 121 DEG C of sterilizing 20min
To the sterile small red bean milk of specified solid content.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with inoculum concentration 1% (v/v) nothing
Bacterium is inoculated in the small red bean milk of solid content 9% (w/w), and 15 DEG C of stationary culture 48h obtain fermented soybean milk B.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk B is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=41% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 3
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of soya-bean milk: after the dry beans weighing of mung bean, the water that five times of quality are added is placed in 37 DEG C of immersion 8h, discards water,
A certain proportion of wet beans and water are added in soy bean milk making machine, the soya-bean milk of solid content needed for being made is obtained through 121 DEG C of sterilizing 20min
To the sterile mung bean milk of specified solid content.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with inoculum concentration 5% (v/v) nothing
Bacterium is inoculated in the mung bean milk of solid content 1% (w/w), and 45 DEG C of stationary culture 2h obtain fermented soybean milk C.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk C is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=42% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 4
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of soya-bean milk: after the dry beans weighing of semen viciae fabae, the water that five times of quality are added is placed in 37 DEG C of immersion 8h, discards water,
A certain proportion of wet beans and water are added in soy bean milk making machine, the soya-bean milk of solid content needed for being made is obtained through 121 DEG C of sterilizing 20min
To the sterile semen viciae fabae soya-bean milk of specified solid content.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with inoculum concentration 2% (v/v) nothing
Bacterium is inoculated in the semen viciae fabae soya-bean milk of solid content 7% (w/w), and 25 DEG C of stationary culture 36h obtain fermented soybean milk D.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk D is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=44% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 5
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) after big white kidney bean and the dry beans of small white kidney bean are mixed with mass ratio 1:1 weighing, five times the preparation of soya-bean milk: are added
The water of quality is placed in 37 DEG C of immersion 8h, discards water, a certain proportion of wet beans and water are added in soy bean milk making machine, solid content needed for being made
The soya-bean milk of content obtains the sterile navy bean soya-bean milk of specified solid content through 121 DEG C of sterilizing 20min.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with inoculum concentration 4% (v/v) nothing
Bacterium is inoculated in the navy bean soya-bean milk of solid content 3% (w/w), and 40 DEG C of stationary culture 12h obtain fermented soybean milk E.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk E is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=43% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 6
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of lactobacillus plantarum (fermenting microbe): the freeze-dried powder of lactobacillus plantarum ATCC14917 is used on a small quantity respectively
Sterile distilled water dissolution takes a ring to line on MRS solid medium (purchase is German from Merck Co.) with oese, and 37 DEG C
Anaerobic culturel takes out for 24 hours, 1mL MRS liquid (purchase is German from Merck Co.) is put into oese picking single colonie, with whirlpool
Bacterium colony is dispersed in fluid nutrient medium by rotation oscillator, and 37 DEG C of Anaerobic culturels take out for 24 hours, with the inoculation of 2% (v/v) inoculum concentration
After 50mL MRS liquid, 37 DEG C of cultures for 24 hours, culture 9,000rpm is centrifuged 10 minutes, is discarded supernatant, the sterile distillation of thallus
After water washing 2 times, is suspended with the sterile distilled water of former volume of culture, obtain the seed of corresponding fermentation, the bacterium of seed liquor is dense
Degree is 2x109cfu/mL。
(c) preparation of soybean milk: with embodiment 1.
(d) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 and lactobacillus plantarum
(L.plantarum) (purchase is from ATCC) respectively with inoculum concentration 1.5% (v/v) aseptic inoculation in solid content by ATCC 14917
In the soybean milk of 5% (w/w), 37 DEG C of stationary cultures obtain fermented soybean milk F for 24 hours.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk F is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=60% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 7
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of lactobacillus plantarum (fermenting microbe): with embodiment 6.
(c) preparation of soybean milk: with embodiment 1.
(d) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with 2.7% (v/ of inoculum concentration
V) and lactobacillus plantarum (L.plantarum) ATCC 14917 (is bought from ATCC) with inoculum concentration 0.3% (v/v) aseptic inoculation
In the soybean milk of solid content 5% (w/w), 37 DEG C of stationary cultures obtain fermented soybean milk G for 24 hours.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk G is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=55% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 8
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of lactobacillus plantarum (fermenting microbe): with embodiment 6.
(c) preparation of soybean milk: with embodiment 1.
(d) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with 0.3% (v/ of inoculum concentration
V) and lactobacillus plantarum (L.plantarum) ATCC 14917 (is bought from ATCC) with inoculum concentration 2.7% (v/v) aseptic inoculation
In the soybean milk of solid content 5% (w/w), 37 DEG C of stationary cultures obtain fermented soybean milk H for 24 hours.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk H is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=51% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 9
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of lactobacillus plantarum (fermenting microbe): with embodiment 6.
(c) preparation of soybean milk: with embodiment 1.
(d) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with 1.5% (v/ of inoculum concentration
V) and lactobacillus plantarum (L.plantarum) ATCC 14917 (is bought from ATCC) with inoculum concentration 1.5% (v/v) aseptic inoculation
In the soybean milk of solid content 3% (w/w), 37 DEG C of stationary cultures obtain fermented soybean milk I for 24 hours.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk I is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=49% of the fermented soybean milk to streptococcus mutans biofilm formation.
Embodiment 10
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of lactobacillus plantarum (fermenting microbe): with embodiment 6.
(c) preparation of soybean milk: with embodiment 1.
(d) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with 1.5% (v/ of inoculum concentration
V) and lactobacillus plantarum (L.plantarum) ATCC 14917 (is bought from ATCC) with inoculum concentration 1.5% (v/v) aseptic inoculation
In the soybean milk of solid content 5% (w/w), 40 DEG C of stationary culture 12h obtain fermented soybean milk J.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
Fermented soybean milk J is in the above way prepared as to measure its suppression to streptococcus mutans biofilm formation after sample to be tested
System activity, the results showed that inhibiting rate I=47% of the fermented soybean milk to streptococcus mutans biofilm formation.
Effect example 1Product taste and fancy grade are tested
Using fermented soybean milk product A, B, C, D, E, F, G, H, I, J prepared by above-described embodiment 1-10 as experimental subjects, into
The taste tests of row product.50 people of test number.Trial test mode: it is tasted by the way of blank marking；Respectively to upper
Color, flavor, mouthfeel, the nutrition item for stating fermented soybean milk product A, B, C, D, E, F, G, H, I, J are individually given a mark, and each single item is full
Dividing is 25 points, calculates average mark and its total score, statistical result are recorded in table 1.Meanwhile it being given according to the whole fancy grade to product
Opinion out, counts the hobby number to each single-item, and statistical result is recorded in table 2.
1 product taste tests result data statistical form of table
2 product fancy grade test result data statistical form of table
It can be seen that in general from product taste tests and fancy grade statistical result, through the invention technical solution
In method obtained by by lactobacillus fermenti, lactobacillus fermenti and lactobacillus plantarum preparation have streptococcus mutans biomembrane
The fermented soybean milk of inhibitory activity can be received in terms of product special flavour, mouthfeel, nutrition by most of consumer.
Effect example 2Stability of the fermented soybean milk to streptococcus mutans biofilm inhibitor activity under refrigerated condition
By fermented soybean milk A, B, C, D, E, F, G, H, I, J prepared by embodiment 1-10 be placed in (4 DEG C) of refrigerated condition save 0,
10, it is taken out after 20 and 30 days, measures each sample respectively to streptococcus mutans biofilm formation inhibiting rate, the results are shown in Table 3.
Stability of the fermented soybean milk to streptococcus mutans biofilm formation inhibitory effect under 3 refrigerated condition of table
As shown in Table 3, the fermented soybean milk of all tests is raw to streptococcus mutans after (4 DEG C) of refrigerated condition save 30 days
Object film forms inhibiting rate and is stably held in same level, and stability is preferable.
Comparative example 1
By the inoculum concentration in embodiment 1, soymilk concentration, cultivation temperature and fermentation time are adjusted one by one, are obtained
The fermented soybean milk of following set of distinct methods preparation, fermented soybean milk obtained by each group to the inhibitory effect of streptococcus mutans biomembrane such as
Shown in table 4.
Inhibitory effect of 4 distinct methods of the table preparation gained fermented soybean milk to streptococcus mutans biofilm formation
From result shown in table 4 it can be concluded that, by inoculum concentration in the preparation method of the fermented soybean milk, soymilk concentration,
When cultivation temperature and fermentation time are adjusted to except preferred scope, by comparing embodiment 1 and comparative example 1 it is found that by
The fermented soybean milk of lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 preparation can still inhibit to become
The forming amount of shape streptococcus organism film, but its inhibitory effect is decreased obviously.
Therefore, within preferred scope, inoculum concentration, soymilk concentration, cultivation temperature and fermentation time influence each other, so that by
The fermented soybean milk of lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 preparation, which has, more preferably to be deformed
The inhibitory effect that streptococcus organism film is formed.
Comparative example 2
By the inoculum concentration in embodiment 6, soymilk concentration, cultivation temperature and fermentation time are adjusted one by one, are obtained
The fermented soybean milk of following set of distinct methods preparation, fermented soybean milk obtained by each group to the inhibitory effect of streptococcus mutans biomembrane such as
Shown in table 5.
Inhibitory effect of 5 distinct methods of the table preparation gained fermented soybean milk to streptococcus mutans biofilm formation
From result shown in table 5 it can be concluded that, by inoculum concentration in the preparation method of the fermented soybean milk, soymilk concentration,
When cultivation temperature and fermentation time are adjusted to except preferred scope, by comparing embodiment 6 and comparative example 2 it is found that by
Lactobacillus fermenti (L.fermentum) CGMCC No.17321 and the (purchase of lactobacillus plantarum (L.plantarum) ATCC 14917
The fermented soybean milk prepared from ATCC) can still inhibit the forming amount of streptococcus mutans biomembrane, but its inhibitory effect is obvious
Decline.
Therefore, within preferred scope, inoculum concentration, soymilk concentration, cultivation temperature and fermentation time influence each other, so that by
Lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 and lactobacillus plantarum (L.plantarum)
ATCC 14917 (buys the inhibitory effect that the fermented soybean milk prepared from ATCC) has more preferably streptococcus mutans biofilm formation.
Comparative example 3
1 the method for reference implementation example compares by lactobacillus fermenti (Lactobacillus fermentum) CGMCC
No.17321, lactobacillus plantarum (L.plantarum) ATCC 14917 (are bought from ATCC), streptococcus thermophilus
(S.thermophilus) ST-BODY-3 (being provided by Hansen Corp., section), (purchase of Lactobacillus casei (L.casei) ATCC 393
For the fermented soybean milk prepared from ATCC) to the inhibitory effect of streptococcus mutans biofilm formation, concrete operations are as follows:
1, materials and methods
(a) preparation of seed (fermenting microbe):
The preparation of lactobacillus fermenti seed (fermenting microbe): with embodiment 1.
The preparation of Lactobacillus casei and lactobacillus plantarum seed: by Lactobacillus casei ATCC 393 and lactobacillus plantarum
The freeze-dried powder of ATCC14917 is dissolved with a small amount of sterile distilled water respectively, and each personal oese takes a ring to line MRS solid culture
On base (purchase is German from Merck Co.), 37 DEG C of Anaerobic culturels take out for 24 hours, are put into 1mL MRS liquid with oese picking single colonie
Bacterium colony, is dispersed in fluid nutrient medium by body (purchase is German from Merck Co.) with vortex oscillator, 37 DEG C of anaerobism trainings
It supports and takes out for 24 hours, 50mL MRS liquid is inoculated in 2% (v/v) inoculum concentration, after 37 DEG C of cultures for 24 hours, culture 9,000rpm centrifugation
It 10 minutes, discards supernatant, after thallus washs 2 times with sterile distilled water, is suspended with the sterile distilled water of former volume of culture, obtain phase
The seed for the fermentation answered.
The preparation of streptococcus thermophilus seed: the freeze-dried powder of streptococcus thermophilus ST-BODY-3 is molten with a small amount of sterile distilled water
Solution takes a ring to line on M17 solid medium (purchase is German from Merck Co.) with oese, and 40 DEG C of Anaerobic culturels take for 24 hours
Out, it is put into 1mL M17 liquid (purchase from Merck Co. Germany) with oese picking single colonie, with vortex oscillator by bacterium
It falls and is dispersed in fluid nutrient medium, 40 DEG C of Anaerobic culturels take out for 24 hours, are inoculated in 50mL M17 liquid with 2% (v/v) inoculum concentration
Body, after 40 DEG C of cultures for 24 hours, culture 9,000rpm is centrifuged 10 minutes, is discarded supernatant, after thallus washs 2 times with sterile distilled water,
It is suspended with the sterile distilled water of former volume of culture, obtains the seed of fermentation.
The preparation of streptococcus mutans bacteria suspension: with embodiment 1.
(b) preparation of soybean milk: with embodiment 1.
(c) fermented soybean milk measures the inhibiting rate of streptococcus mutans biofilm formation: with embodiment 1.
2, the preparation of fermented soybean milk
By each bacterial strain with 3% (v/v) inoculum concentration aseptic inoculation in the soybean milk of solid content 5% (w/w), respectively
It cultivates (37 DEG C of lactobacillus fermenti, lactobacillus plantarum and Lactobacillus casei Anaerobic culturels, 40 DEG C of Anaerobic culturels of streptococcus thermophilus)
For 24 hours, corresponding fermented soybean milk is obtained.
3, measurement of the fermented soybean milk to streptococcus mutans biofilm formation inhibiting rate
The fermented soybean milk of above-mentioned different strains preparation is as shown in table 6 to the inhibitory effect of streptococcus mutans biofilm formation:
Inhibitory effect of the fermented soybean milk of 6 different strains of table preparation to streptococcus mutans biofilm formation
As shown in Table 6, the hair prepared by lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321
Ferment soya-bean milk inhibiting rate is 45%, and the fermented soybean milk of other bacterial strains preparation does not have streptococcus mutans biofilm inhibitor activity.
By embodiment 6 it is found that lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 and plant
(buy the fermented soybean milk inhibiting rate prepared jointly from ATCC) is 60% to object lactobacillus (L.plantarum) ATCC 14917, will
The fermented soybean milk of itself and lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 preparation compares, and sends out
Kefir milk bacillus (Lactobacillus fermentum) CGMCC No.17321 and lactobacillus plantarum (L.plantarum) ATCC
14917 (it is significant to the inhibitory activity of streptococcus mutans biomembrane to buy the fermented soybean milk prepared jointly from ATCC).
Comparative example 4
1, materials and methods
(a) preparation of lactobacillus fermenti seed (fermenting microbe) and streptococcus mutans bacteria suspension: with embodiment 1.Plant cream
The preparation of bacillus (fermenting microbe): with embodiment 6.
(b) fermentation liquid measures the inhibiting rate of streptococcus mutans biofilm formation:
The pretreatment of fermentation liquid: fermentation liquid pH is adjusted to 6.80 to get sample to be tested.
The measurement of inhibitory activity: it referring to 1 the method for embodiment, is surveyed using unfermentable culture medium as blank control
It is fixed.
2, the preparation of lactobacillus fermenti single bacterium fermentation liquid, lactobacillus fermenti and lactobacillus plantarum mixed fermentation liquid
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 with 3% (v/v) inoculum concentration without
Bacterium is inoculated in MRS fluid nutrient medium (Merck Co. Germany), and 37 DEG C of Anaerobic culturels for 24 hours, obtain the fermentation of lactobacillus fermenti single bacterium
Liquid；
By lactobacillus fermenti (Lactobacillus fermentum) CGMCC No.17321 and lactobacillus plantarum
(L.plantarum) ATCC 14917 (from ATCC) respectively with 1.5% (v/v) inoculum concentration, train in MRS liquid for purchase by aseptic inoculation
Base is supported, 37 DEG C of Anaerobic culturels for 24 hours, obtain lactobacillus fermenti and lactobacillus plantarum mixed fermentation liquid.
3, measurement of the fermentation liquid to streptococcus mutans biofilm formation inhibiting rate
Its inhibition to streptococcus mutans biofilm formation is measured after fermentation liquid to be in the above way prepared as to sample to be tested
Activity, the results showed that lactobacillus fermenti single bacterium fermentation liquid, lactobacillus fermenti and lactobacillus plantarum mixed fermentation liquid are to deformation hammer
The inhibiting rate of bacterium biofilm formation is respectively 12%, 13%.
Comparative example 5
The pH of fermented soybean milk A, B, C, D, E, F, G, H, I and J prepared by embodiment 1-10 are adjusted to 6.80, then with 4 DEG C,
10,000rpm centrifugation 10min, supernatant fraction is sterile fermented soybean milk (sample to be tested).According to method described in embodiment 1
It measures above-mentioned sample to carry out to streptococcus mutans biofilm formation inhibiting rate, the results are shown in Table 7.
The sterile fermented soybean milk of table 7 is to streptococcus mutans biofilm formation inhibitory effect
As shown in Table 7, sterile fermented soybean milk still has stronger inhibition to streptococcus mutans biofilm formation inhibiting rate
Activity illustrates product of the inhibiting effect after bacterial strain metabolism soya-bean milk.But the biomembrane inhibitory effect of the sterile fermented soybean milk of each group
Sample sets before being below aseptic process, this shows that the effect of the inhibition biomembrane mycoderm of the fermented soybean milk before aseptic process comes from
Viable bacteria and metabolite collective effect.
The preparation method to fermented soybean milk provided by the present invention, fermented soybean milk and application have carried out detailed Jie above
It continues.Used herein a specific example illustrates the principle and implementation of the invention, and the explanation of above embodiments is only
It is to be used to help understand method and its core concept of the invention.It should be pointed out that for those skilled in the art
For, it without departing from the principle of the present invention, can be with several improvements and modifications are made to the present invention, these improve and repair
Decorations are also fallen within the protection scope of the claims of the present invention.
Claims (10)
1. a kind of preparation method of the fermented soybean milk by lactobacillus fermenti preparation, which comprises the following steps: will ferment
Lactobacillus CGMCC No.17321 is inoculated in soya-bean milk as fermenting agent and carries out fermentating metabolism, obtains fermented soybean milk.
2. the preparation method of fermented soybean milk according to claim 1, which is characterized in that lactobacillus fermenti CGMCC
The inoculum concentration of No.17321 is 2x107~1x108cfu/mL。
3. the preparation method of fermented soybean milk according to claim 2, which is characterized in that the fermenting agent is inoculative proportion
(purchase is certainly by lactobacillus fermenti CGMCC No.17321 and lactobacillus plantarum (L.plantarum) ATCC 14917 of 1~9:1~9
ATCC composite fermentation microbial inoculum).
4. the preparation method of fermented soybean milk according to claim 1, which is characterized in that the soya-bean milk is soybean, red bean, green
Beans, semen viciae fabae, big white kidney bean, small white kidney bean it is one or more.
5. the preparation method of fermented soybean milk according to claim 1, which is characterized in that the solid quality in the soya-bean milk
Degree is 1%~9%.
6. the preparation method of fermented soybean milk according to claim 1, which is characterized in that the temperature of the fermentation be 15 DEG C~
45 DEG C, the time of fermentation is 2~48h.
7. the preparation method of fermented soybean milk according to claim 1, which is characterized in that the fermentation is stationary culture.
8. a kind of fermented soybean milk prepared by lactobacillus fermenti, which is characterized in that by any preparation method of claim 1~7
It is made.
9. fermented soybean milk according to claim 8, which is characterized in that the fermented soybean milk is to streptococcus mutans biomembrane
Inhibiting rate >=41% of formation.
10. a kind of fermented soybean milk as claimed in claim 8 is in the application of preparation anti-caries tooth food.
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| CN111134199A (en) * | 2020-02-20 | 2020-05-12 | 杭州九阳豆业有限公司 | Preparation method of beany flavor-free fermented soybean milk and prepared beany flavor-free fermented soybean milk |
| CN115011528A (en) * | 2022-07-13 | 2022-09-06 | 光明乳业股份有限公司 | Application of lactobacillus fermentum in preparation of normal-temperature dairy product |
| CN115152842A (en) * | 2022-07-13 | 2022-10-11 | 光明乳业股份有限公司 | Lactobacillus fermentum normal-temperature milk beverage and preparation method thereof |
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| CN115152842A (en) * | 2022-07-13 | 2022-10-11 | 光明乳业股份有限公司 | Lactobacillus fermentum normal-temperature milk beverage and preparation method thereof |
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