CN109402005A - A kind of probiotics leaven and its has the function of the application in ethanol degradation acidified milk in fermentation preparation - Google Patents
A kind of probiotics leaven and its has the function of the application in ethanol degradation acidified milk in fermentation preparation Download PDFInfo
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- CN109402005A CN109402005A CN201811319810.8A CN201811319810A CN109402005A CN 109402005 A CN109402005 A CN 109402005A CN 201811319810 A CN201811319810 A CN 201811319810A CN 109402005 A CN109402005 A CN 109402005A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/12—Fermented milk preparations; Treatment using microorganisms or enzymes
- A23C9/123—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt
- A23C9/1234—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt characterised by using a Lactobacillus sp. other than Lactobacillus Bulgaricus, including Bificlobacterium sp.
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- A23C9/1238—Fermented milk preparations; Treatment using microorganisms or enzymes using only microorganisms of the genus lactobacteriaceae; Yoghurt using specific L. bulgaricus or S. thermophilus microorganisms; using entrapped or encapsulated yoghurt bacteria; Physical or chemical treatment of L. bulgaricus or S. thermophilus cultures; Fermentation only with L. bulgaricus or only with S. thermophilus
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- A23C9/1307—Milk products or derivatives; Fruit or vegetable juices; Sugars, sugar alcohols, sweeteners; Oligosaccharides; Organic acids or salts thereof or acidifying agents; Flavours, dyes or pigments; Inert or aerosol gases; Carbonation methods
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Abstract
The present invention provides a kind of probiotics leaven and its applications in alcohol prepared by fermenting degradation function acidified milk, belong to fermented food technical field, lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus viable count ratio are (1.5~2.5): (0.8~1.2): (0.8~1.2) in the probiotics leaven.Strain in the probiotics leaven is resistant to ethyl alcohol; and has the function of ethanol degradation; it is worked well in terms of reducing internal ethanol content, removing the free radical that organism metabolism generates using the composite fermentation cream with ethanol degradation function of the probiotics leaven fermenting and producing; stomach lining can be protected and absorption of the ethyl alcohol in stomach and intestine is effectively reduced, product nutritive value is high, with rich flavor, effect is obvious.
Description
Technical field
The invention belongs to fermented food technical field more particularly to a kind of probiotics leaven and its in alcohol prepared by fermenting
Application in degradation function acidified milk.
Background technique
After people drinks, ethyl alcohol is metabolized the deposition that can be generated many harmful substances and a large amount of free radicals occur in vivo, gives people
Body causes different degrees of injury.Currently, it is commercially available relieve the effect of alcohol, product of sobering up play its effect mostly be based on such product in human body
Alcohol metabolism can be influenced after taking or ethyl alcohol is inhibited to absorb, but the internal ethyl alcohol that cannot directly degrade.Many hangover medicines are not
Substantive sobering-up functions only play the effects of comfort is calm, alleviation is had a headache, though some hangover medicines relieve the effect of alcohol, wake up with certain
Wine effect, but can be also harmful to human health along with different side effects.The people of especially different constitutions, for different
The reaction of hangover medicine is not identical, therefore it is not recommended that drunk person is sobered up using hangover medicine.
Microorganism growth process needs carbohydrate, if it can use ethyl alcohol as one of carbon source, can play drop
Solve the effect of ethyl alcohol.But the research of this respect is rarely reported, at present still without it is a kind of health, be effectively applied to it is awake
The food of wine.
Summary of the invention
In view of this, the purpose of the present invention is to provide a kind of probiotics leaven and its in alcohol prepared by fermenting degradation function
Application in energy acidified milk.The acidified milk of the probiotics leaven fermentation preparation, not only nutritive value is high, with rich flavor, also
Free radical and protection gastric mucosa for being generated with the internal ethanol content of reduction, removing organism metabolism and other effects.
In order to achieve the above-mentioned object of the invention, the present invention provides following technical schemes:
A kind of probiotics leaven, including following components: lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus,
The lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus viable count ratio are (1.5~2.5): (0.8~1.2): (0.8
~1.2)；Living bacteria count >=10 in the probiotics leaven10CFU/g。
Preferably, the probiotics leaven further includes protective agent；The lactobacillus acidophilus, streptococcus thermophilus and Bao Jiali
Sub- lactobacillus gross mass and protectant mass ratio are 1:(2.5~5).
The present invention also provides the preparation methods of the probiotics leaven, comprising the following steps:
1) by lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus it is activated, domestication proliferation after be inoculated into respectively
Seed liquor, the seed liquor of streptococcus thermophilus and Bao Jia that seed liquor culture obtains lactobacillus acidophilus are carried out in MRS fluid nutrient medium
The seed liquor of Leah lactobacillus；
2) by the seed liquor of the lactobacillus acidophilus, the seed liquor of the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus
Combined inoculation fermented and cultured into MRS fluid nutrient medium obtains mixed fermentation liquid；
3) centrifugation step 2) described in mixed fermentation liquid, collection solid phase components be probiotics leaven.
It preferably, further include that the solid phase components are mixed with protective agent, freezes and does after collecting solid phase components in step 3)
Dry acquisition probiotics leaven freeze-dried powder.
Preferably, the culture medium that proliferation is tamed described in step 1) is degreasing milk medium, the temperature of the domestication proliferation
It is 39~41 DEG C, the time of the domestication proliferation is 6~10h.
Preferably, the time of seed liquor culture described in step 1) is 22~26h, and the temperature of the seed liquor culture is 36
~38 DEG C.
The present invention provides application of the probiotics leaven in alcohol prepared by fermenting degradation function acidified milk.
The present invention also provides a kind of preparation methods of acidified milk with ethanol degradation function, comprising the following steps:
A) cow's milk, white granulated sugar and whipping cream are mixed and obtain mixture, two sections of homogeneous are carried out to the mixture, sterilize,
It is cooling, obtain mixture after cooling；
B probiotics leaven of any of claims 1 or 2) is inoculated with into the mixture after cooling, 28~32 DEG C quiet
It sets 28~32h of fermentation and obtains acidified milk.
The preparation method of the present invention also provides a kind of composite fermentation cream with ethanol degradation function, including following step
It is rapid:
A) pueraria lobata is mixed with beating with water, after sterilizing, cooling down, is inoculated with probiotics leaven of any of claims 1 or 2,
40~44 DEG C of 12~18h of fermentation obtain pueraria lobata fermentation liquid；
B) pueraria lobata fermentation liquid described in the acidified milk and step a) is mixed and obtains composite fermentation cream.
Preferably, the quality of pueraria lobata fermentation liquid described in step b) is the 3.5~4.5% of acidified milk quality.
Beneficial effects of the present invention: probiotics leaven provided by the invention include lactobacillus acidophilus, streptococcus thermophilus and
Lactobacillus bulgaricus is resistant to ethyl alcohol, and has the function of ethanol degradation, by the lactobacillus acidophilus, streptococcus thermophilus
Can be applied to exploitation with the probiotics leaven of lactobacillus bulgaricus compounding preparation, there is the functional of ethanol degradation effect to produce
The viable bacteria content of product, the probiotics leaven is high, reaches 1010~1012CFU/g。
Further, the composite fermentation cream pH with ethanol degradation function of the probiotics leaven fermenting and producing is utilized
For 3.57~3.69, acidity be 86.30~94.13 ° of T, viscosity is 382.5~508.3MPa.S, retention ability be 55.01%~
56.92%, viable count reaches 5.42 × 109~7.15 × 109CFU/mL, 82.1~83.3 points of sensory evaluation scores, manual simulation's stomach,
It is respectively 397.59 ± 4.12mg GSH that ethanol degradation rate, which reaches 67%, ABTS and DPPH free radical scavenging ability, under the conditions of intestinal juice
Equivalents/mL and 224.34 ± 21.37mg GSH equivalents/mL and 4.96 ± 0.12mg GSH
Equivalents/mL and 3.46 ± 0.10mg GSH equivalents/mL, alcohol dehydrogenase (ADH) activity ratio 40.14%,
Mouse blood alcohol concentration is remarkably decreased from filling wine 1h to acid filling milk 3h, and mouse liver function index measures during relieving the effect of alcohol
It reduces the raising of AST and ALT, hepar damnification be greatly reduced and protection liver, mouse sobers up the time than sobering up after normally drinking
Time shortens 29.12%.It can be seen that the composite fermentation cream is reducing internal ethanol content, is removing what organism metabolism generated
It works well in terms of free radical, pueraria lobata fermentation liquid therein, which contains Puerarin, can protect stomach lining and ethyl alcohol is effectively reduced in stomach and intestine
In absorption, product nutritive value is high, with rich flavor, effect is obvious.
Detailed description of the invention
Fig. 1 is complex function fermentation cream to ethanol degradation effect picture in Mice Body；
Fig. 2 is complex function fermentation cream to ALT changes of contents influence diagram in mouse liver；
Fig. 3 is complex function fermentation cream to AST changes of contents influence diagram in mouse liver；
Fig. 4 is complex function fermentation cream to SGOT changes of contents influence diagram in mouse blood；
Fig. 5 is complex function fermentation cream to SGPT changes of contents influence diagram in mouse blood.
Specific embodiment
The present invention provides a kind of probiotics leavens, including following components: lactobacillus acidophilus, streptococcus thermophilus and Bao Jia
Leah lactobacillus, the lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus viable count ratio are (1.5~2.5): (0.8
~1.2): (0.8~1.2)；Living bacteria count >=10 in the probiotics leaven10CFU/g。
Heretofore described probiotics leaven includes lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus；This
Lactobacillus acidophilus described in invention (CICC6091), streptococcus thermophilus (CICC6063) and lactobacillus bulgaricus (CICC6064)
Preferably it is purchased from Chinese industrial Microbiological Culture Collection administrative center (CICC).Heretofore described lactobacillus acidophilus, thermophilic chain
Coccus and lactobacillus bulgaricus are resistant to ethyl alcohol, and have good ethanol degradation ability.It is heretofore described thermophilic
Lactobacillus lactis, streptococcus thermophilus and lactobacillus bulgaricus viable count ratio are (1.5~2.5): (0.8~1.2): (0.8~
1.2), preferably 2:1:1.Living bacteria count >=10 in heretofore described probiotics leaven10CFU/g, preferably 1010
~1012CFU/g.Heretofore described probiotics leaven preferably further includes protective agent, the lactobacillus acidophilus, thermophilus
The gross mass of bacterium and lactobacillus bulgaricus and protectant mass ratio are 1:(2.5~5), preferably 1:3.The present invention
Described in protective agent preferably include glycerol, glucose and ascorbic acid, the mass ratio of the glycerol, glucose and Vitamin C bolt
Preferably 2:1:2；In the present invention, the lactobacillus acidophilus, streptococcus thermophilus and Bulgaria are protected in protectant effect
Lactobacillus is not influenced by external conditions such as low gently drieds.
The present invention also provides the preparation methods of the probiotics leaven, comprising the following steps:
1) by lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus it is activated, domestication proliferation after be inoculated into respectively
Seed liquor, the seed liquor of streptococcus thermophilus and Bao Jia that seed liquor culture obtains lactobacillus acidophilus are carried out in MRS fluid nutrient medium
The seed liquor of Leah lactobacillus；
2) by the seed liquor of the lactobacillus acidophilus, the seed liquor of the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus
Combined inoculation fermented and cultured into MRS fluid nutrient medium obtains mixed fermentation liquid；
3) centrifugation step 2) described in mixed fermentation liquid, collection solid phase components be probiotics leaven.
In the present invention, the lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus are activated first；The present invention
The method of the activation is not particularly limited, using this field conventional bacterial classification activation method, is embodied in the present invention
In the process, the activation preferably distinguishes the freeze-dried powder of the lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus
It is inoculated in MRS broth bouillon, 37 DEG C of constant temperature incubations are for 24 hours.The present invention is after the activation, by the acidophilus cream bar after activation
Bacterium, streptococcus thermophilus and lactobacillus bulgaricus carry out domestication proliferation, in the present invention, the inoculation strain of the domestication breeding
Volume be preferably the 2~6% of culture volume, more preferably 3~5%；The culture medium of the domestication proliferation is skimmed milk training
Base is supported, the temperature of the domestication proliferation is preferably 39~41 DEG C, and more preferably 40 DEG C, the time of the domestication proliferation is preferably 6
~10h, more preferably 7~9h.The effect of heretofore described domestication proliferation is to improve bacterial activity and its fermenting property.
The present invention is inoculated into MRS fluid nutrient medium after domestication proliferation, by the strain after the domestication proliferation respectively
Middle seed liquor, the kind of the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus for carrying out seed liquor culture and obtaining lactobacillus acidophilus
Sub- liquid.In the present invention, the time of the seed liquor culture is preferably 22~26h, more preferably 23~25h, most preferably for 24 hours；
The temperature of the seed liquor culture is preferably 36~38 DEG C, and more preferably 37 DEG C.In heretofore described seed liquor incubation,
The volume for being inoculated with strain is preferably the 2~6% of culture volume, more preferably 3~5%.In the present invention, the lactobacillus acidophilus
Seed liquor, independent preferably 2 ╳ of viable count in the seed liquor of the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus
108CFU/mL。
The present invention is by the seed liquor of the lactobacillus acidophilus of acquisition, the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus
Seed liquor combined inoculation fermented and cultured into MRS fluid nutrient medium obtains mixed fermentation liquid.In the present invention, the acidophilus cream bar
The mixed volume ratio of the seed liquor of the seed liquor of bacterium, the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus be preferably (1.5~
2.5): (0.8~1.2): (0.8~1.2), more preferably 2:1:1.In the present invention, the volume of the combined inoculation inoculation strain
Preferably the 3~7% of MRS fluid nutrient medium volume, more preferably 4~6%；The time of the mixed fermentation is preferably 22~
26h, more preferably 23~25h, most preferably for 24 hours；The temperature of the mixed fermentation is preferably 36~38 DEG C, more preferably 37
℃。
The present invention obtains mixed fermentation liquid after the mixed fermentation, is centrifuged the mixed fermentation liquid, collects solid phase
Group is divided into probiotics leaven.In the present invention, the temperature of the centrifugation is preferably 3~5 DEG C, and more preferably 4 DEG C, the centrifugation
Revolving speed be preferably 6000~8000rpm, more preferably 6500~7500rpm, the time of the centrifugation is preferably 15~
25min, more preferably 20min.For the present invention after the centrifugation, collecting solid phase components is probiotics leaven.
The present invention preferably further includes mixing the solid phase components with protective agent, being cold after the collection solid phase components
Dry acquisition probiotics leaven freeze-dried powder is lyophilized.The gross mass of the solid phase components and protectant quality in the present invention
Ratio is 1:(2.5~5), preferably 1:3.Heretofore described protective agent preferably includes glycerol, glucose and ascorbic acid,
The mass ratio of the glycerol, glucose and Vitamin C bolt is preferably 2:1:2.Heretofore described freeze-drying preferably includes will
The mixed material after -80 DEG C of pre-cooling 12h, with vacuum and low temperature freeze drier in -40 DEG C be freeze-dried 20~for 24 hours.
The present invention provides application of the probiotics leaven in alcohol prepared by fermenting degradation function acidified milk.This hair
In bright, the probiotics leaven fermentation cow's milk obtains the acidified milk with ethanol degradation function；It is described that there is ethanol degradation function
The acidified milk of energy further includes composite fermentation cream, i.e., compounds other function product with acidified milk and prepare composite fermentation cream.
The present invention also provides a kind of preparation methods of acidified milk with ethanol degradation function, comprising the following steps: A)
Cow's milk, white granulated sugar and whipping cream are mixed and obtain mixture, two sections of homogeneous are carried out to the mixture, sterilization, cooling obtain cold
But the mixture after；B the probiotics leaven, 28~32 DEG C of standing for fermentation) are inoculated with into the mixture after cooling
28~32h obtains acidified milk.
In the present invention, cow's milk, white granulated sugar and whipping cream are mixed and obtains mixture.In the present invention, the cow's milk, white sand
The mass ratio of sugar and whipping cream is preferably (90~110): (10~20): (30~50), more preferably (95~105): (12~
18): (35~45), most preferably 100:15:40.In the present invention, the cow's milk is preferably fresh milk；The present invention is to described white
The source of granulated sugar and whipping cream is not particularly limited, using this field conventional commercial product.The present invention is obtaining described mix
After closing material, two sections of homogeneous are carried out to the mixture；Two sections of homogeneous include the first homogeneous and the second homogeneous, and described first
The pressure of matter is preferably 7.5~8.5MPa, more preferably 8MPa, and the time of first homogeneous is preferably 8~12min, more excellent
It is selected as 10min；The pressure of second homogeneous is preferably 9.5~10.5MPa, more preferably 10MPa, first homogeneous when
Between preferably 15~25min, more preferably 20min.The present invention carries out the material after homogeneous after two sections of homogeneous
Sterilization, cooling, obtain mixture after cooling.In the present invention, the method for the sterilization is preferably pasteurize, the Pasteur
The temperature of sterilization is preferably 80~90 DEG C, and more preferably 82~87 DEG C, most preferably 85 DEG C, the time of the pasteurize is preferred
For 12~18min, more preferably 15min.In the present invention, the cooling is preferably circulating water, mixture after the cooling
Temperature be preferably 42~47 DEG C, more preferably 45 DEG C.
The present invention is inoculated with the probiotics leaven, 28~32 DEG C of standings into the mixture after cooling of the acquisition
28~the 32h that ferments obtains acidified milk.In the present invention, the inoculum concentration of the probiotics leaven is preferably 1~3wt%, more excellent
It is selected as 1.5~2.5wt%.The present invention preferably further includes continuously stirring to make described mix after being inoculated with the probiotics leaven
It closes material to be uniformly mixed with the probiotics leaven, the revolving speed continuously stirred is preferably 30~50rpm, described to continuously stir
Time be preferably 20~30min.The present invention is left to ferment after described continuously stir；The temperature of the standing for fermentation
Preferably 28~32 DEG C, more preferably 30 DEG C, the time of the standing for fermentation is preferably 28~32h, more preferably 30h.This hair
The bright container to the standing for fermentation is not particularly limited, using this field normal sterile round.The present invention is in institute
It preferably further include that the acidified milk is placed in 3~5 DEG C of progress after-ripening, the time of the after-ripening is excellent after stating standing for fermentation
It is selected as 8~12h, more preferably 10h.
The present invention obtains the acidified milk with ethanol degradation function after the standing for fermentation, after-ripening.In the present invention
The acidified milk with ethanol degradation function individually eat with reduce internal ethanol content, remove that organism metabolism generates from
The effect of by base, can be used in sobering up.The heretofore described acidified milk with ethanol degradation function can also be with other functionality
Product mix prepares composite fermentation cream.
The preparation method of the present invention also provides a kind of composite fermentation cream with ethanol degradation function, including following step
It is rapid: pueraria lobata is mixed with beating with water a), after sterilizing, cooling down, is inoculated with the probiotics leaven, 40~44 DEG C of fermentations 12~
18h obtains pueraria lobata fermentation liquid；B) acidified milk is obtained with the mixing of pueraria lobata fermentation liquid described in step a) compound
Acidified milk.
In the present invention, pueraria lobata and water are mixed with beating.In the present invention, the pueraria lobata is preferably the fresh pueraria lobata removed the peel
Piece, preferred be soaked in colour protecting liquid carries out color protection before the pueraria lobata is water mixing；The colour protecting liquid is preferably bicarbonate two
The mass concentration of sodium water solution, the disodium bicarbonate aqueous solution is preferably 0.3%, and the disodium bicarbonate is preferably food-grade
Disodium bicarbonate；The present invention does not have particular/special requirement to the volume of the colour protecting liquid, can submerge the pueraria lobata, in the present invention
The time of the color protection is preferably 8~12min, more preferably 10min.The present invention is after the color protection, by pueraria lobata and water
It is mixed with beating, the mass ratio of the pueraria lobata and water is preferably (0.8~1.2): (0.8~1.2), more preferably 1:1；The mashing
It is preferably carried out using high-speed homogenization machine, the parameter 10000rpm of the high-speed homogenization machine.The present invention after mashing,
It is sterilized, the sterilization is preferably high pressure moist heat sterilization, and the temperature of the sterilization is preferably 98~102 DEG C, more preferably 100
DEG C, the time of the sterilization is preferably 3~8min, more preferably 5min.The present invention cools down after the sterilization, institute
Stating cooling is preferably natural cooling, and the temperature after cooling is preferably 41~43 DEG C, and more preferably 42 DEG C.The present invention is to described
It is inoculated with the probiotics leaven in material after cooling and carries out fermentation acquisition pueraria lobata fermentation liquid；The temperature of the fermentation is 40
~44 DEG C, preferably 42 DEG C, the time of the fermentation are 12~18h, preferably 13~17h, more preferably 15h.
The acidified milk and the pueraria lobata fermentation liquid are mixed after obtaining the pueraria lobata fermentation liquid by the present invention
Obtain composite fermentation cream.In the present invention, the quality of the pueraria lobata fermentation liquid is preferably the 3.5~4.5% of acidified milk quality, more excellent
It is selected as 3.8~4.2%.The revolving speed of heretofore described mixing is preferably 20~30rpm, and the time of the mixing is excellent
It is selected as 10~15min.The present invention preferably further includes packing and cryopreservation step after the mixing；The present invention is to described
The specification and packaging material of packing are not particularly limited, using the specification and packaging material of this field normal fermentation cream；This
The temperature of refrigeration described in invention is preferably 3~5 DEG C, and more preferably 4 DEG C.
Technical solution provided by the invention is described in detail below with reference to embodiment, but they cannot be understood
For limiting the scope of the present invention.
Embodiment 1
The preparation of probiotics leaven
Steps are as follows:
1) 3 kinds of lactobacillus acidophilus, streptococcus thermophilus, lactobacillus bulgaricus bacterial strain freeze-dryings the domestication proliferation of probiotics: are taken
Powder is inoculated in respectively in 10mL sterilizing MRS broth bouillon, and vortex is mixed in 37 DEG C of constant temperature incubation activation for 24 hours, then will activation
Bacterial strain afterwards is inoculated into degreasing milk medium for 2% inoculum concentration by volume respectively and carries out domestication proliferation, 40 DEG C of constant temperature trainings
10h is supported to curdled milk；
2) preparation of probiotics leaven: the inoculation that 3 kinds of strain cultured solutions by domestication proliferation are respectively 2% with volume ratio
Amount access MRS Liquid Culture is based on 37 DEG C of constant temperature incubations and obtains seed liquor for 24 hours, and seed liquor is pressed lactobacillus acidophilus: thermophilus
Bacterium: the bacterial strain ratio that lactobacillus bulgaricus volume ratio is 2:1:1 mixes, and then accesses MRS with the inoculum concentration that volume ratio is 7%
In fluid nutrient medium for 24 hours in 37 DEG C of constant temperature incubations, the hybrid bacterial strain culture solution obtained is obtained in 4 DEG C, 6000r/min centrifugation 15min
Bacterium mud sediment is obtained, according to bacterium mud sediment: in -80 DEG C of pre-cooling 12h after the ratio mixing that protective agent is 1:3, using vacuum and low temperature
Freeze drier freeze-drying 20h obtains probiotics leaven freeze-dried powder, and detection viable bacteria content reaches 1010CFU/g。
The preparation of composite fermentation cream with ethanol degradation function:
Ferment-fermented cow's milk: the white granulated sugar and 40% for being 15% by milk quality percentage is added into Fresh Milk
Whipping cream is heated to 65 DEG C, is stirred continuously to whole dissolutions, then carries out two sections of homogeneous, first segment homogenization pressure is adjusted to 8MPa
10min is kept, second segment homogenization pressure rises to 10MPa and keeps 20min, cow's milk is heated to 85 DEG C after homogeneous and is kept the temperature
15min is sterilized, and is then rapidly cooled to 45 DEG C with cool circulation water, the probiotics that inoculation milk quality percentage is 1% sends out
Ferment agent (embodiment 1 prepares), tank enters in sterilized stainless steel metal container after being continuously stirring to uniformly, then is put
The standing for fermentation for entering to carry out 32h in the Yogurt making machine through ultraviolet sterilization and temperature adjustment to 30 DEG C, is placed in after fermentation in 4 DEG C of refrigerator
Refrigeration 10h obtains acidified milk；
The preparation of pueraria lobata fermentation liquid: it is fresh pueraria lobata is peeled and washed, thinly slice, it is soaked in 0.3% bicarbonate two
Color protection 10min in sodium water solution guarantees that solution can be totally submerged pueraria lobata surface, is cleaned after color protection with clear water, then will
Pueraria lobata: tap water is to be beaten in high-speed homogenization machine after 1:1 is mixed according to mass ratio, and slurries are packed into 500mL triangular flask and exist
100 DEG C of sterilization 5min are carried out in high-pressure sterilizing pot, taking-up, which is put, after sterilizing is cooled to 42 DEG C into gnotobasis, is inoculated with pueraria lobata
The probiotics leaven that mass percent is 2% obtains pueraria lobata fermentation liquid after the 15h that ferments in 42 DEG C of constant temperature fermentation boxes；
The compounding of complex function fermentation cream: the acidified milk 1kg using probiotics leaven production is weighed in the stainless of sterilizing
In steel metal container, it is that pueraria lobata fermentation liquid is added in 3.5% additive amount by acidified milk mass percent, is stirred continuously to uniform, point
It is stored refrigerated in 4 DEG C in Sour milk bottle loaded on 200mL, it can be prepared by function fermented dairy product；
The Product Quality Evaluation of complex function fermentation cream: product pH is 3.57, acidity is that 94.13 ° of T, viscosity are
508.3MPa.S, retention ability are that 56.92% and viable count reach 7.15 × 109CFU/mL, color, smell, flavour, structural state
4 index sensory evaluation criterias are shown in Table 1, and the sensory evaluation scores of the complex function fermentation cream are 82.6 points.
Embodiment 2
The preparation of probiotics leaven
1) 3 kinds of lactobacillus acidophilus, streptococcus thermophilus, lactobacillus bulgaricus bacterial strain freeze-dryings the domestication proliferation of probiotics: are taken
Powder is inoculated in respectively in 10mL sterilizing MRS broth bouillon, and vortex is mixed in 37 DEG C of constant temperature incubation activation for 24 hours, by 3 after activation
Kind of bacterial strain is inoculated into degreasing milk medium for 4% inoculum concentration by volume respectively and carries out domestication proliferation, 40 DEG C of constant temperature incubations
8h is to curdled milk；
2) preparation of probiotics leaven: the inoculation that 3 kinds of strain cultured solutions by domestication proliferation are respectively 2% with volume ratio
Amount access MRS Liquid Culture is based on 37 DEG C of constant temperature incubations and obtains seed liquor for 24 hours, and seed liquor is pressed lactobacillus acidophilus: thermophilus
Bacterium: the bacterial strain ratio that lactobacillus bulgaricus volume ratio is 2:1:1 mixes, and then accesses MRS with the inoculum concentration that volume ratio is 7%
In fluid nutrient medium for 24 hours in 37 DEG C of constant temperature incubations, the hybrid bacterial strain culture solution obtained, 4 DEG C, 7000r/min centrifugation hybrid bacterial strain
Culture solution 20min obtains bacterium mud sediment, according to bacterium mud sediment: being pre-chilled after the ratio mixing that protective agent is 1:3 at -80 DEG C
12h obtains probiotics leaven freeze-dried powder with vacuum and low temperature freeze drier freeze-drying 22h, and viable bacteria content reaches
1011CFU/g。
The preparation of composite fermentation cream with ethanol degradation function:
Ferment-fermented cow's milk: the white granulated sugar and 40% for being 15% by milk quality percentage is added into Fresh Milk
Whipping cream is heated to 65 DEG C, is stirred continuously to whole dissolutions, then carries out two sections of homogeneous, first segment homogenization pressure is adjusted to 8MPa
10min is kept, second segment homogenization pressure rises to 10MPa and keeps 20min, cow's milk is heated to 85 DEG C after homogeneous and is kept the temperature
15min is sterilized, and is then rapidly cooled to 45 DEG C with cool circulation water, and the cow's milk for being cooled to 45 DEG C to sterilization is inoculated with milk quality
Percentage be 1% probiotics leaven, be continuously stirring to uniformly after tank enter in sterilized stainless steel metal container, then by its
It is put into the standing for fermentation that 30h is carried out in the Yogurt making machine through ultraviolet sterilization and temperature adjustment to 30 DEG C；
The preparation of pueraria lobata fermentation liquid: it is fresh pueraria lobata is peeled and washed, thinly slice, it is soaked in 0.3% bicarbonate two
Color protection 10min in sodium water solution guarantees that solution can be totally submerged pueraria lobata surface, is cleaned after color protection with clear water, then will
Pueraria lobata: tap water is to be beaten in high-speed homogenization machine after 1:1 is mixed according to mass ratio, and slurries are packed into 500mL triangular flask and exist
100 DEG C of sterilization 5min are carried out in high-pressure sterilizing pot, taking-up, which is put, after sterilizing is cooled to 42 DEG C into gnotobasis, is inoculated with pueraria lobata
The probiotics leaven that mass percent is 2% obtains pueraria lobata fermentation liquid after the 15h that ferments in 42 DEG C of constant temperature fermentation boxes；
The compounding of function acidified milk: the acidified milk 5kg using probiotics leaven production is weighed in the stainless steel gold of sterilizing
Belong in container, is that pueraria lobata fermentation liquid is added in 4.0% additive amount by acidified milk mass percent；
Function fermented dairy product quality evaluation: product pH is 3.61, acidity is 91.43 ° of T, viscosity 458.1MPa.S, holds
Waterpower is that 56.12% and viable count reach 6.45 × 109CFU/mL, 4 color, smell, flavour, structural state index sense organs are commented
Divide and reaches 83.3 points.
Embodiment 3
The production of probiotics leaven and ethanol degradation function acidified milk
Implementation process is substantially with embodiment 1, and only the following is different:
1) 3 kinds of lactobacillus acidophilus, streptococcus thermophilus, lactobacillus bulgaricus bacterial strain freeze-dryings the domestication proliferation of probiotics: are taken
Powder is inoculated in respectively in 10mL sterilizing MRS broth bouillon, and vortex is mixed in 37 DEG C of constant temperature incubation activation for 24 hours, by 3 after activation
Kind of bacterial strain is inoculated into degreasing milk medium for 6% inoculum concentration by volume respectively and carries out domestication proliferation, 40 DEG C of constant temperature incubations
6h is to curdled milk；
2) preparation of probiotics leaven: the inoculation that 3 kinds of strain cultured solutions by domestication proliferation are respectively 2% with volume ratio
Amount access MRS Liquid Culture is based on 37 DEG C of constant temperature incubations and obtains seed liquor for 24 hours, and seed liquor is pressed lactobacillus acidophilus: thermophilus
Bacterium: the bacterial strain ratio that lactobacillus bulgaricus volume ratio is 2:1:1 mixes, and then accesses MRS with the inoculum concentration that volume ratio is 7%
In fluid nutrient medium for 24 hours in 37 DEG C of constant temperature incubations, the hybrid bacterial strain culture solution obtained.It is obtained in 4 DEG C, 8000r/min centrifugation 25min
Bacterium mud sediment is obtained, according to bacterium mud sediment: in -80 DEG C of pre-cooling 12h after the ratio mixing that protective agent is 1:3, using vacuum and low temperature
Freeze drier freeze-drying obtains probiotics leaven freeze-dried powder for 24 hours, and viable bacteria content reaches 1012CFU/g；
3) white granulated sugar and 40% for being 15% by milk quality percentage ferment-fermented cow's milk: is added into Fresh Milk
Whipping cream, be heated to 65 DEG C, be stirred continuously to whole dissolutions, then carry out two sections of homogeneous, first segment homogenization pressure is adjusted to
8MPa keeps 10min, and second segment homogenization pressure rises to 10MPa and keeps 20min, cow's milk is heated to 85 DEG C after homogeneous and is kept the temperature
15min is sterilized, and is then rapidly cooled to 45 DEG C with cool circulation water, and the cow's milk for being cooled to 45 DEG C to sterilization is inoculated with milk quality
Percentage be 2% probiotics leaven, be continuously stirring to uniformly after tank enter in sterilized stainless steel metal container, then by its
It is put into the standing for fermentation that 28h is carried out in the Yogurt making machine through ultraviolet sterilization and temperature adjustment to 30 DEG C；
4) preparation of pueraria lobata fermentation liquid: it is fresh pueraria lobata is peeled and washed, thinly slice, it is soaked in 0.3% bicarbonate
Color protection 10min in two sodium water solutions guarantees that solution can be totally submerged pueraria lobata surface, is cleaned after color protection with clear water, then
By pueraria lobata: tap water is to be beaten in high-speed homogenization machine after 1:1 is mixed according to mass ratio, and slurries are packed into 500mL triangular flask
100 DEG C of sterilization 5min are carried out in high-pressure sterilizing pot, taking-up, which is put, after sterilizing is cooled to 42 DEG C into gnotobasis, is inoculated with Pueraria lobota
The probiotics leaven that root mass percent is 2% obtains pueraria lobata fermentation liquid after the 15h that ferments in 42 DEG C of constant temperature fermentation boxes；
5) compounding of function acidified milk: the acidified milk 10kg using probiotics leaven production is weighed in the stainless steel of sterilizing
It is that pueraria lobata fermentation liquid is added in 4.5% additive amount by acidified milk mass percent in canister；
6) function fermented dairy product quality evaluation: product pH is 3.69, acidity is 86.30 ° of T, viscosity 382.5MPa.S,
Retention ability is that 55.01% and viable count reach 5.42 × 109CFU/mL, 4 color, smell, flavour, structural state index sense organs
Scoring reaches 82.1 points.
Complex function fermentation milk product sensory evaluation scores table is shown in Table 1 in above-described embodiment 1~3
1 complex function fermentation milk product sensory evaluation scores table of table
Embodiment 4
Probiotics leaven alcohol dehydrogenase (ADH) activity ratio, free radical scavenging ability and has the function of ethanol degradation
Fermented dairy product animal sober up experimental result
External functional activity measurement:
(1) alcohol dehydrogenase activity measures
Using watt Le-Huo He (Valle&Hoch) method, study sobering-up fermented milk in the gastrointestinal tract to alcohol dehydrogenase
(ADH) activation.Sodium pyrophosphate buffer solution 1.5mL, NAD+1.0mL, ethanol solution 0.5mL, sample 0.1mL are drawn, is mixed
After conjunction at 25 DEG C, warm bath 5min is covered, alcohol dehydrogenase enzyme solutions 0.1mL is added immediately, uses spectrophotometer after shaking up immediately
It measures absorbance (A340 value), later primary every 10s reading, METHOD FOR CONTINUOUS DETERMINATION 5min.It is returned to zero with control tube, the anti-of drug is not added
Ying Zuwei blank.
(2) alcohol dehydrogenase activity calculating method
It is plotted against time with A, negates and answer initial stage part, calculated the value added of A 340/10s, existed according to NADH
Molar extinction coefficient at 340nm is 6.2, calculates enzyme activity.The nanomole number table that the activity of ADH is generated with NADH per minute
Show.
(activation) rate (%)=(enzymatic activity is without medicine control group-enzymatic activity dosing measurement group)/enzymatic activity is inhibited to compare without medicine
Group × 100%
(3) solution is prepared
The preparation of artificial simulation intestinal juice:
4 6.85g of KH 2PO is weighed, distilled water 500m L is added to make it dissolve, with the N aO H solution tune of 0.4g/100m L
PH value is saved to 6.8, is diluted with water, is then that 1g/100m L is added according to trypsase additive amount, water is added to dissolve it sufficiently,
It is settled to 1L, is mixed spare.
The preparation of artificial simulation gastric juices: 2 are adjusted to 100mL, PH with the concentrated hydrochloric acid of distilled water dilution 23.4mL, is therefrom taken dilute
Then hydrochloric acid 16.4mL is added pepsin according to the additive amount of 1g/100mL, adds water to dissolve it sufficiently, be settled to 1L, mix
It is even spare.
(4) alcohol dehydrogenase (ADH) activity ratio measures
It takes complex function fermentation cream 10mL, 4 DEG C of 4000r/min to be centrifuged 20min, takes supernatant measurement simulated gastrointestinal tract front and back
ADH activity ratio, PH are adjusted to 2.0.It takes 1mL sobering-up fermented milk in the test tube containing 9mL simulate the gastric juice, is vortexed and mixes, 37 DEG C of guarantors
After warm 2h, boiling water bath heats 10min to terminate reaction.Extract reaction solution the ADH activity ratio of measurement digestion front and back.Its pH is adjusted to again
6.8, it is added with the volume ratio of 1:9 into simulated intestinal fluid, simulation digestion 2h is protected from light in 37 DEG C of waters bath with thermostatic control, it is cooling rapidly, so
Sample liquid is centrifuged 15min in 4 DEG C, 6000r/min again afterwards, takes the ADH activity ratio of supernatant measurement digestion front and back.Equivalent is added to steam
Distilled water is as a control group.
(5) free radical scavenging ability measures
It takes complex function fermentation cream 10mL, 4 DEG C of 4000r/min to be centrifuged 20min, takes supernatant measurement simulated gastrointestinal tract front and back
Antioxidant activity, pH value are adjusted to 2.0.It takes 1mL complex function fermentation cream in the test tube containing 9mL simulate the gastric juice, is vortexed and mixes,
After 37 DEG C of heat preservation 2h, boiling water bath heats 10min to terminate reaction.Extract reaction solution the antioxidant activity of measurement digestion front and back.Again will
Its pH is adjusted to 6.8, is added with the volume ratio of 1:9 into simulated intestinal fluid, simulation digestion 2h is protected from light in 37 DEG C of waters bath with thermostatic control, rapidly
It is cooling, sample liquid is then centrifuged 15min in 4 DEG C, 6000r/min again, supernatant is taken to measure its antioxidant activity.Equivalent is added to steam
Distilled water is as a control group.
Animal in vivo functionality determination of activity:
(1) zoopery
KM mouse 40, four groups are randomly divided into, is divided into control group, low dose group, middle dose group, high dose group, every group 10
Only.Every group of mouse carries out stomach-filling with 16mL/kg dosage white wine after each group mouse fasting 16h.With righting reflex loss for drunk finger
Mark.Reach the composite fermentation cream stomach-filling carried out after state of intoxication in embodiment 1, blank group stomach-filling physiological saline；Low middle high dose
The acidified milk of group difference stomach-filling 1.5mL/kg, 3mL/kg, 6mL/kg, take whole blood to carry out ethyl alcohol after 60,120,180min
Measurement of concetration.
(2) whole blood ethanol content measures
Using ethanol content in 1300 gas chromatograph of TRACE detection whole blood, referring to Pharmacopoeia of the People's Republic of China second
Alcohol content measuring method carries out alcohol levels in blood samples concentration mensuration using external standard method.
Chromatographic condition: vaporizing chamber: 230 DEG C, sensing chamber: 230 DEG C, chromatographic column temperature: 70 DEG C, nitrogen 20mL/min, hydrogen
30mL/min, air 100mL/min.PEG-20M chromatographic column is selected, the alcohols application in the chromatographic column headspace analysis blood is more wide
It is general.
Head space condition: sample room temperature 70 C, 90 DEG C of line temperature, 90 DEG C of valve road temperature, carrier gas (N2) 120.0kPa, it blows
Sweep pressure 220.0kPa.
Standard curve preparation: ethyl alcohol standard reserving solution (200mg/100mL) 0,0.1,0.2,0.4,0.8mL is taken to push up in 10mL
In empty bottle plus blank group blood is to 1mL, then mass concentration is 0,20,40,80,160mg/100mL, mixes, is placed in Head-space equipment
In 70 DEG C of constant temperature 30min, automatic sampling top tank air 1mL, it is qualitative with retention time, with peak area quantification, with concentration of alcohol
It (mg/100mL) is abscissa, peak area (mV) is that ordinate draws standard curve.
(3) mouse liver functional testing
Tissue homogenate preparation: tissue takes 0.5g, cleans in the physiological saline of pre-cooling, removes blood.With 9 times of tissue weight
Pre- cold saline impregnate after.It is put into the container for being loaded with ice cube.It is homogenized, the homogenate of electric homogenizer ice bath, with
3000rpm4 DEG C of centrifugation 10min, takes supernatant, and -20 DEG C of freezen protectives are to be measured.Serum preparation: it is centrifuged, takes at once after blood sampling
Clear liquid is put into -80 DEG C of refrigerators and saves.Sample detection builds up the provided kit of Bioengineering Research Institute using Nanjing, exists respectively
It is detected under 510nm wavelength using microplate reader, tissue sample carries out absorbance detection after being diluted.It is calculated by light absorption value
Enzyme activity.Calculation formula: standard curve/tissue homogenate protein concentration (unit U/ obtained by tissue enzyme activity=standard items
Gprot) seroenzyme activity: protein concentration (unit U/L) in standard curve/serum obtained by seroenzyme activity=standard items
ALT (glutamic-pyruvic transaminase) is primarily present in various cells, especially most with liver cell content.Under normal condition, only
It to be released into blood on a small quantity, the activity of its enzyme can be significantly raised in serum.Therefore transaminase (especially ALT) is that Acute Hepatic is thin
The sensitive labels of born of the same parents' damage.Drinking may be such that transaminase increases.Illustrate wine when AST (glutamic-oxalacetic transaminease) and ALT are generally increased
Essence generates damage to liver, and complex function fermentation cream can reduce the raising of AST and ALT during relieving the effect of alcohol, can be significantly
Hepar damnification is reduced, liver is protected.When liver damages, SGOT (content in glutamic-oxalacetic transaminease blood) and SGPT (paddy third
Content in transaminase blood) blood can be entered by liver, so this two indexs be also detect hepar damnification degree index it
One.
(4) sober up effect is tested
Mouse (male and female is fifty-fifty) 20 is taken, is randomly divided into 2 groups, every group 5, respectively blank control group and product is tested
Group.After fasting 12h, 2 groups of white wine for pouring into 0.2mL/10g respectively record drunk (righting reflex loss) of each group and sober up and (turn over
Positive reflection restores) time, specifically start timing after drinking to fill.
Simulated gastrointestinal tract alcohol dehydrogenase (ADH) activity ratio measurement result is shown in Table 2
The measurement of 2 simulated gastrointestinal tract alcohol dehydrogenase (ADH) activity ratio of table
Note: sample 1 is the Yoghourt using streptococcus thermophilus, bulgaria lactobacillus fermentation；Sample 2 is to utilize thermophilus
The Yoghourt that bacterium, lactobacillus bulgaricus, lactobacillus acidophilus are fermented；Sample 3 be using streptococcus thermophilus, lactobacillus bulgaricus,
The acidified milk of lactobacillus acidophilus fermentation addition pueraria lobata.
Simulated gastrointestinal tract free radical scavenging ability measurement result hands over table 3.
The measurement of 3 simulated gastrointestinal tract free radical scavenging ability of table
Note: sample 1 is the Yoghourt using streptococcus thermophilus, bulgaria lactobacillus fermentation；Sample 2 is in embodiment 1
Acidified milk；Sample 3 is the composite fermentation cream in embodiment 1.
Mouse liver functional testing the results are shown in Table 4,5,6,7.
ALT assay in 4 mouse liver of table
AST assay in 5 mouse liver of table
SGOT assay in 6 mouse blood of table
SGPT assay in 7 mouse blood of table
Ethanol degradation function fermented dairy product results of animal is shown in Table 8.
There is table 8 acidified milk of ethanol degradation function to sober up experimental result
As can be seen from the above embodiments, 3 kinds of streptococcus thermophilus, lactobacillus bulgaricus, lactobacillus acidophilus bacterial strain interworking preparations
What probiotics leaven was produced has the function of that ethanol degradation acidified milk is reducing internal ethanol content, removing organism metabolism generation
Free radical in terms of work well, the pueraria lobata fermentation liquid in product, which contains Puerarin, can protect stomach lining and ethyl alcohol is effectively reduced exist
Absorption in stomach and intestine, product nutritive value is high, with rich flavor, effect is obvious.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (10)
1. a kind of probiotics leaven, which is characterized in that including following components: lactobacillus acidophilus, streptococcus thermophilus and Bao Jiali
Sub- lactobacillus, the lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus viable count ratio are (1.5~2.5): (0.8~
1.2): (0.8~1.2)；Living bacteria count >=10 in the probiotics leaven10CFU/g。
2. probiotics leaven according to claim 1, which is characterized in that the probiotics leaven further includes protection
Agent；The lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus gross mass and protectant mass ratio are 1:
(2.5~5).
3. the preparation method of probiotics leaven of any of claims 1 or 2, comprising the following steps:
1) by lactobacillus acidophilus, streptococcus thermophilus and lactobacillus bulgaricus it is activated, domestication proliferation after be inoculated into MRS liquid respectively
Seed liquor, the seed liquor of streptococcus thermophilus and bulgarian milk that seed liquor culture obtains lactobacillus acidophilus are carried out in body culture medium
The seed liquor of bacillus；
2) seed liquor of the seed liquor of the lactobacillus acidophilus, the seed liquor of streptococcus thermophilus and lactobacillus bulgaricus is mixed
It is seeded to fermented and cultured in MRS fluid nutrient medium and obtains mixed fermentation liquid；
3) centrifugation step 2) described in mixed fermentation liquid, collection solid phase components be probiotics leaven.
4. the preparation method of probiotics leaven according to claim 3, which is characterized in that collect solid phase group in step 3)
It further include that the solid phase components are mixed with protective agent, are freeze-dried acquisition probiotics leaven freeze-dried powder after point.
5. the preparation method of probiotics leaven according to claim 3, which is characterized in that tame and increase described in step 1)
The culture medium grown be degreasing milk medium, it is described domestication proliferation temperature be 39~41 DEG C, it is described domestication proliferation time be 6~
10h。
6. the preparation method of probiotics leaven according to claim 3, which is characterized in that seed liquor described in step 1)
The time of culture is 22~26h, and the temperature of the seed liquor culture is 36~38 DEG C.
7. prepared by preparation method described in probiotics leaven of any of claims 1 or 2 or claim 3~6 any one
Application of the probiotics leaven of acquisition in alcohol prepared by fermenting degradation function acidified milk.
8. a kind of preparation method of the acidified milk with ethanol degradation function, comprising the following steps:
A) cow's milk, white granulated sugar and whipping cream are mixed and obtain mixture, two sections of homogeneous are carried out to the mixture, sterilizes, is cooling,
Obtain mixture after cooling；
B probiotics leaven of any of claims 1 or 2,28~32 DEG C of standing hairs) are inoculated with into the mixture after cooling
28~32h of ferment obtains acidified milk.
9. a kind of preparation method of the composite fermentation cream with ethanol degradation function, comprising the following steps:
A) pueraria lobata is mixed with beating with water, after sterilizing, cooling down, is inoculated with probiotics leaven of any of claims 1 or 2,40~
44 DEG C of 12~18h of fermentation obtain pueraria lobata fermentation liquid；
B) pueraria lobata fermentation liquid described in acidified milk described in claim 8 and step a) is mixed and obtains composite fermentation
Cream.
10. preparation method according to claim 9, which is characterized in that the quality of pueraria lobata fermentation liquid described in step b) is
The 3.5~4.5% of acidified milk quality.
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