CN106434473B - The whey pear wine of lactobacillus plantarum WZMX-2 and application and its preparation - Google Patents
The whey pear wine of lactobacillus plantarum WZMX-2 and application and its preparation Download PDFInfo
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- CN106434473B CN106434473B CN201610916194.9A CN201610916194A CN106434473B CN 106434473 B CN106434473 B CN 106434473B CN 201610916194 A CN201610916194 A CN 201610916194A CN 106434473 B CN106434473 B CN 106434473B
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- lactobacillus plantarum
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- fermentation
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Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
- C12R2001/25—Lactobacillus plantarum
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
- C07K14/335—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Lactobacillus (G)
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12G—WINE; PREPARATION THEREOF; ALCOHOLIC BEVERAGES; PREPARATION OF ALCOHOLIC BEVERAGES NOT PROVIDED FOR IN SUBCLASSES C12C OR C12H
- C12G3/00—Preparation of other alcoholic beverages
- C12G3/02—Preparation of other alcoholic beverages by fermentation
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- Biophysics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Tropical Medicine & Parasitology (AREA)
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- Biomedical Technology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Dairy Products (AREA)
Abstract
The present invention discloses the whey pear wine of lactobacillus plantarum WZMX-2 and application and its preparation, 4:4:2 is uniformly mixed the whey fermentation liquid formed by lactobacillus plantarum WZMX-2, Kluyveromyces marxianus bacterium WWMJ-1 and S. cervisiae with whey bergamot pear juice mixed liquor by volume, -72h for 24 hours is cultivated at 25 DEG C -37 DEG C, alcoholic strength reaches 7.8%vol, total acid reaches 60.84 ° of T, total sugar content reaches 42.34g/L for measurement, bitter taste peptide content reaches 0.15g/100mL fermentation and obtains without bitter taste whey pear wine；The whey pear wine of preparation in pistac, clarification it is bright, with whey and bergamot pear intrinsic color and fragrance, without bitterness sense, the time of mice burden swimming can be extended through functional trial confirmation, the generation of serum urea when reducing movement, increase the deposit of hepatic glycogen, the weight and blood lactase acid of mouse are had no significant effect, there is function of relieving physical fatigue effect.
Description
Technical field
The invention belongs to the technical fields of drinks processing technology, and more specifically, it is mixed that the present invention relates to a kind of microorganism fungus kinds
Close the technical field of fermentation preparation whey pear wine method.
Background technique
Whey is the byproduct caused by during cow's milk production cheese, casein, is in yellow green, translucent.Cream
Clear nutritive value is extremely abundant, and up to 55% or so nutriment has stayed in whey in raw milk, mainly includes thick egg
White about 1%, fat 0.3~0.4%, lactose 3%~5%, lactalbumin 20%, also containing minerals, vitamin etc., wherein
Lactalbumin is divided into beta lactoglobulin, α-lactoprotein, and wherein beta lactoglobulin can combine calcium and some hydrophobic molecules such as
Retinol, vitamin E play the role of blood pressure lowering, resisting stress, the disappearance for promoting fear memory, reduce cholesterol levels；α-cream
Albumen can promote galactosis, it may have anti-hypertension adjusts cell growth, immunological regulation, the effect relieved stress, and raising is recognized
Intellectual can and improve sleep；Lactose can provide energy, 1g lactose about production capacity 16.75kJ for body.Lactose is as unique in milk
Carbohydrate, be one of the energy that is obtained at first from milk after infant's birth.Lactose can promote calcium in enteron aisle, phosphorus and
The absorption of the minerals such as manganese inhibits the breeding of other harmful bacterias, adjusts colony balance.In addition, whey mineral is in good
Equilibrium state, there is complete nutritive effect, required minerals can be supplemented for the mankind.Therefore, whey is considered as one
There is the byproduct of application value in a pole, can be applied to the fields such as food, medicine, feed, chemical industry.
In China, the development and utilization level of whey is very low at present, although containing multiple nutritional components in whey, it is total
Soma content of material is low, and the separation of various composition is extracted, and especially technology needed for the separation and Extraction of mineral composition is complicated, sets
Standby expensive, relative cost height, and for whey drink, the milk beverage of various brands has formd certain pin on the market
Scale is sold, but whey class drink is extremely rare.For intuitive, lactose, lactalbumin, minerals and vitamins are contained in whey
Strong Deng valuable nutritional ingredient, newborn taste, whey source is sufficient, and it is more can to develop whey beverage, milone etc. using whey
Kind has the whey product compared with high added value, and the selling market of whey product should be not less than other milk beverages；But in fact, mesh
Preceding whey drink rare in the market is constantly in trial sale state and fails industrialization production listing.To find out its cause, being due to whey
In contain bitter taste, make converted products with this " rear bitter taste " phenomenon of obvious bitter taste.And current debitterness technology do not obtain also it is prominent
Broken, also not up to production functionization.De- suffering topic in whey has become the technical bottleneck for restricting whey processing and utilization, related
Technical research, accumulation, breakthrough will start the industrialized development of whey processing industry, there is highly important economic value and society
It can meaning.
Summary of the invention
State of the art deficiency and there are aiming at the problem that, to solve the above-mentioned problems, the present invention provides plants
The preparation method of lactobacillus WZMX-2 and its whey pear wine of application, compared with prior art, plant cream bar provided by the invention
It is the preparation method high production efficiency of bacterium WZMX-2 and its whey pear wine of application, no added, acidified milk pure mellow wine bitter taste is effectively removed,
Overcome the technical bottleneck for having apparent " rear bitter taste " using whey converted products in the prior art, the whey pear wine prepared
Uniform color, in pistac, clarification it is bright, with whey and bergamot pear intrinsic color and fragrance, no bitterness sense, mouthfeel is inclined
Sweet tea, mild acidity, harmony is moderate, and mellowness is simple and elegant, and mouthfeel is plentiful, has good harmony.Orally administration mouse is born
Weight swimming test, serum urea measurement test, hepatic glycogen measurement test and blood lactase acid measurement test, utilize plant cream as the result is shown
The whey pear wine of bacillus WZMX-2 preparation can be obviously prolonged the time of mice burden swimming, the life of serum urea when reducing movement
At increasing the deposit of hepatic glycogen, had no significant effect to the weight and blood lactase acid of mouse, shown using lactobacillus plantarum WZMX-2 system
Standby whey pear wine has the significant technical effect for alleviating physical fatigue.
The present invention specifically adopts the following technical scheme that:
The lactobacillus plantarum (Lactobacillus plantarum) that the present invention selects is the geographical environment according to Xinjiang
Particularity filters out the well-grown micro- life of a batch by sampling from self-control cheese that Xinjiang South Mountain acquires, Kuerle delicious pear
Object bacterial strain carries out culture, separation and the screening of microorganism fungus kind from cheese and bergamot pear, therefrom preferably goes out one plant of number and is
The lactobacillus plantarum of WZMX-2, the bacterial strain were preserved in budapest treaty microorganism International Depository Authority before the applying date: in
State's Microbiological Culture Collection administration committee common micro-organisms center (CGMCC).Preservation date is on 08 08th, 2016, preservation
It number is CGMCC No.12832.The 16S rRNA homogeneous assays of the bacterial strain show itself and Lactobacillus plantarum parent
Source relationship is nearest, and homologous similitude is 99%, and Phylogenetic Analysis result all shows that deposit number is CGMCC No.12832 plant
Lactobacillus (Lactobacillus plantarum) WZMX-2 bacterial strain is one plant of novel bacterial, utilizes the novel bacterial of screening and outer
Purchase S. cervisiae and existing preservation of bacteria strain number are WWMJ-1, and Marx's Crewe that deposit number is CGMCC No.8432 ties up ferment
Female bacterium (Kluyveromyces marxianus) compatibility fermentation bergamot pear juice and whey preparation are without bitter taste whey pear wine.
The present invention specifically provides lactobacillus plantarum (Lactobacillus plantarum) CGMCC of preparation whey pear wine
No.12832.The bacterium numbering used is the lactobacillus plantarum (Lactobacillus plantarum) of WZMX-2, bacterium warp
MRS culture medium is separately cultured, then can be generated molten calcium circle in improved MRS culture medium and be carried out preliminary screening, which cultivates in MRS
The flat lining out culture of base for 24 hours, can form that colony edge is neat, surface is smooth, milky, the circle of protrusion, diameter 0.3mm-
The bacterium colony of 5mm；The strain culturing condition is 37 DEG C of cultivation temperature, and preferred growth is in MRS media surface, the group of MRS culture medium
It is divided into beef extract 10.0g/L, yeast extract 5.0g/L, glucose 20.0g/L, sodium acetate 5.0g/L, citric acid hydrogen diamine 2.0g/L,
Tween 80 1.0ml/L, dipotassium hydrogen phosphate 2.0g/L, epsom salt 0.2g/L, seven water manganese sulfate 0.05g/L, distilled water 1.0L,
Solid medium adds agar 20.0g/L；Bacterial strain colony diameter size on solid medium is 0.3mm-5mm, and front is rounded,
Lateral projections；Neat in edge, surface are smooth；Bacterium colony is opaque, is creamy white；With certain toughness, in Physiology and biochemistry and bacterium
In terms of kind of characteristic, number be with the bacterium numbering that uses for the lactobacillus plantarum (Lactobacillus of WZMX-2
Plantarum) there are visibly different strain properties with common lactobacillus plantarum (Lactobacillus plantarum).
Lactobacillus plantarum (Lactobacillus plantarum) WZMX-2 was preserved in Budapest before the applying date
Treaty microorganism International Depository Authority: China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC), ground
Location: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica, postcode: 100101.Preservation date is
On 08 08th, 2016, deposit number was CGMCC No.12832.Lactobacillus plantarum (Lactobacillus plantarum)
The gene order of WZMX-2 such as SEQUENCE LISTING.Through the 16S rRNA homogeneous assays bacterium and Lactobacillus
Plantarum parent source relationship is nearest, and homologous similitude is 99%, and Phylogenetic Analysis result all shows that deposit number is CGMCC
No.12832 bacterial strain WZMX-2 is the novel bacterial in lactobacillus plantarum (Lactobacillus plantarum), be with it is common often
The lactobacillus plantarum (Lactobacillus plantarum) seen has the novel bacterial of significant difference feature.
Meanwhile the present invention specifically provides a kind of no bitter taste whey pear wine, prepares especially by the preparation method provided as follows
It obtains:
(1) activation of strain: by the lactobacillus plantarum saved at 4 DEG C (Lactobacillus plantarum) WZMX-
2CGMCC No.12832 is inoculated in MRS culture medium in an aseptic environment, is placed in 37 DEG C of cultures for 24 hours；The mark that will be saved at 4 DEG C
Think Kluyveromyces sp (Kluyveromyces marxianus) WWMJ-1CGMCC No.8432 and common S. cervisiae
It is inoculated in YPD culture medium and activates respectively in an aseptic environment, be placed in 28 DEG C of cultures for 24 hours, constant-temperature preserving is spare.
(2) bergamot pear pre-processes: selecting fresh Kuerle delicious pear that handle is gone to clean, filters after being squeezed the juice with automatic juice extractor, obtains fragrant
Pear juice is stored spare at room temperature.
(3) whey pre-processes: being added after selecting fresh whey to remove impurity and foreign matter in whey with four layers of filtered through gauze
The amount of 6%-14% v sucrose, mixing be preheating to 85 DEG C, homogeneous 5min under the conditions of 20MPa；By whey liquid after homogeneous
Pasteurize is carried out, is cooled to room temperature, sterile preservation is spare.
(4) preparation of whey bergamot pear juice mixed liquor: the bergamot pear juice and 90% debulking step (3) that step (2) are obtained obtain
Whey by bergamot pear juice account for mixeding liquid volume 40%-60% be uniformly mixed, save backup at room temperature.
(5) preparation of female fermentation liquid: volume percentage is pressed, the whey that (3) the step of 10% obtain is sub-packed in hair
In ferment container, aseptically, the lactobacillus plantarum (Lactobacillus plantarum) that will be activated by step (1)
WZMX-2CGMCC No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1CGMCC
No.8432 and common S. cervisiae are respectively connected in round by 3% inoculative proportion, and condition of culture successively distinguishes 37 DEG C
Culture for 24 hours, 28 DEG C culture for 24 hours, 28 DEG C culture for 24 hours, constant-temperature preserving is spare.
(6) it ferments: lactobacillus plantarum (Lactobacillus plantarum) WZMX-2CGMCC prepared by step (5)
No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1 and saccharomyces cerevisiae fermented liquid
After mixing according to volume ratio 4:4:2 meter, it is added in the whey bergamot pear juice mixed liquor of the residue 90% of step (4) preparation,
- 72h for 24 hours, fermentation termination setting are cultivated at 25 DEG C -37 DEG C are as follows: measurement alcoholic strength reaches 7.8%vol, and measurement total acid reaches
60.84 ° of T, measurement total sugar content reach 42.34g/L, and measurement bitter taste peptide content reaches 0.15g/100mL.
(7) post-process: after product to fermentation termination, flash distillation sterilization carries out sterile filling after cooling and prepares no hardship
Taste whey pear wine.
The present invention is selected from sieve strain: Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1,
The bacterial strain was preserved in budapest treaty microorganism International Depository Authority: Chinese microorganism strain preservation management before the applying date
Committee's common micro-organisms center (CGMCC), address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, the micro- life of the Chinese Academy of Sciences
Object research institute, postcode: 100101.Preservation date is on November 05th, 2013, and deposit number is CGMCC No.8432, using the bacterium
The condition of culture of kind is 28 DEG C of cultivation temperature, and preferred growth uses yeast extract in YPD media surface, YPD nutrient media components
20g/L agar powder is added in 10g/L, peptone 20g/L, glucose 20g/L, solid medium.Meanwhile the present invention provides bacterial strain horse
Gram think Kluyveromyces sp (Kluyveromyces marxianus) WWMJ-1 zymotechnique: aseptically, picking should
Bacterial strain is cultivated for 24 hours at 28 DEG C.
S. cervisiae selects the common saccharomyces cerevisiae of outsourcing, selects outsourcing S. cervisiae condition of culture and fermentation condition all
Using common knowledge.
In the present invention, alcoholic strength measurement is measured after obtaining alcohol water blend using distillation with alcohol meter；Acidity assaying uses
Determination of acid-basetitration；Pol measurement is measured using hand-held saccharometer: bitter peptides measurement is measured using chloroform-methanol extraction.
The present invention prepares a kind of whey pear wine through the above technical solution, and product quality characteristic is as follows:
(1) organoleptic indicator: color is in pistac, micro- yellow color for having whey intrinsic；Fragrance: frankincense taste is along with light
Light fruit aroma, flavour: wine body gracefulness, sour-sweet tasty and refreshing, free from extraneous odour；Posture: a little precipitating of the bright permission of clarification.
(2) physical and chemical index: total acid: 60.84 ° of T, total sugar content: 42.34g/L.
(3) microbiological indicator: total plate count (cfu/mL)≤100；Coliform (MPN/100mL)≤3；Pathogenic bacteria: not
Detection.
Using technical solution provided by the invention can achieve it is following the utility model has the advantages that
(1) technical solution of the present invention is used, acidified milk pure mellow wine bitter taste is effectively removed, overcomes and utilizes whey in the prior art
Converted products has a technical bottleneck of apparent " rear bitter taste ", the whey pear wine uniform color of preparation, saturating in pistac, clarification
It is bright, with whey and bergamot pear intrinsic color and fragrance, no bitterness sense, the inclined sweet tea of mouthfeel, mild acidity, harmony is moderate, mellowness
Simple and elegant, mouthfeel is plentiful, has good harmony.
(2) whey pear wine orally administration mouse prepared by the present invention carries out Loaned swimming test, and serum urea measurement is tested,
Hepatic glycogen measurement test and blood lactase acid measurement test, whey pear wine can be obviously prolonged the time of mice burden swimming as the result is shown,
The generation of serum urea, increases the deposit of hepatic glycogen, has no significant effect, show to the weight and blood lactase acid of mouse when reducing movement
The novel bacterial screened using the present invention and outsourcing S. cervisiae and existing preservation of bacteria strain Kluyveromyces marxianus bacterium
(Kluyveromyces marxianus) WWMJ-1CGMCC No.8432 compatibility fermentation bergamot pear juice and whey, utilize the complete of offer
It is whole to remove bitter taste and there is alleviation physical fatigue without bitter taste whey pear wine using prepared by whey, bergamot pear juice mixed fermentation technology
Function affect obtains significant technical effect.
Detailed description of the invention:
Fig. 1 is shown as lactobacillus plantarum provided by the invention (Lactobacillus plantarum) WZMX-2CGMCC
The colonial morphology figure of No.12832.
Fig. 2 is shown as lactobacillus plantarum provided by the invention (Lactobacillus plantarum) WZMX-2CGMCC
The phyletic evolution of No.12832 develops tree graph.
Fig. 3 is shown as the process flow chart of whey pear wine preparation method provided by the invention.
Fig. 4 is shown as the influence that different vaccination ratio provided by the invention ferments to whey pear wine, and in figure, inoculative proportion is
Refer to lactobacillus plantarum (Lactobacillus plantarum) WZMX-2CGMCC No.12832, Kluyveromyces marxianus bacterium
The volume ratio of (Kluyveromyces marxianus) WWMJ-1CGMCC No.8432 and S. cervisiae.
Fig. 5 is shown as the influence that different fermentations temperature provided by the invention ferments to whey pear wine.
It is the influence fermented to whey pear wine that Fig. 6, which is shown as the different fermentations time provided by the invention,.
Fig. 7 is shown as the influence that different sucrose additions provided by the invention ferment to whey pear wine.
The response surface that Fig. 8 is shown as inoculative proportion provided by the invention and fermentation time interaction influences alcoholic fermentation
Figure.
The contour that Fig. 9 is shown as inoculative proportion provided by the invention and fermentation time interaction influences alcoholic fermentation
Figure.
The response that Figure 10 is shown as inoculative proportion provided by the invention and fermentation temperature interaction influences alcoholic fermentation
Face figure.
Figure 11 be shown as inoculative proportion provided by the invention and fermentation temperature interaction alcoholic fermentation is influenced it is contour
Line chart.
The response that Figure 12 is shown as fermentation temperature provided by the invention and fermentation time interaction influences alcoholic fermentation
Face figure.
Figure 13 be shown as fermentation temperature provided by the invention and fermentation time interaction alcoholic fermentation is influenced it is contour
Line chart.
Figure 14 is shown as the surface chart of inoculative proportion provided by the invention Yu fermentation time reciprocal effect bitter peptides.
Figure 15 is shown as the contour map of inoculative proportion provided by the invention Yu fermentation time reciprocal effect bitter peptides.
Figure 16 is shown as the surface chart of fermentation temperature and inoculative proportion reciprocal effect bitter peptides provided by the invention.
Figure 17 is shown as the contour map of fermentation temperature and inoculative proportion reciprocal effect bitter peptides provided by the invention.
Figure 18 is shown as the surface chart of fermentation time and fermentation temperature reciprocal effect bitter peptides provided by the invention.
Figure 19 is shown as the high line chart of fermentation time and fermentation temperature reciprocal effect bitter peptides provided by the invention.
Specific embodiment
1-19 and embodiment with reference to the accompanying drawing, specific embodiments of the present invention will be described in further detail, but this
Invention is not limited to following embodiments.
Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1 CGMCC that the present invention uses
No.8432, S. cervisiae can be bought by public channel；The main instrument and equipment used: LD2X-50KB vertical electric
Pressure steam sterilizer (Shanghai Shen An Medical treatment device factory)；HR40- II A2 Biohazard Safety Equipment (the limited public affairs of Qingdao Haier special type electric appliance
Department)；DHP-781 electro-heating standing-temperature cultivator (Hebei province Huangshi medical apparatus and instruments factory)；DZKW-172 electric-heated thermostatic water bath (Beijing
Light Medical Instruments factory, city)；CH1776J microcomputer electromagnetic oven (Glanz living electric apparatus Manufacturing Co., Ltd, Zhongshan city)；SXKW
Digital display control-temperature electric heating set (Beijing forever bright Medical Instruments factory)；EYELA water-bath (Shanghai Ai Lang Instrument Ltd)；Rotation
Evaporimeter (German IKA, 8V-C).
All raw and auxiliary materials that the present invention selects, equipment, and bacterium culture medium, the cultural method selected all are this field
Known selection, but implementation of the invention is not limited, other some material and facilities well known in the art are applied both to this hair
The implementation of bright following implementation.The % being related in the present invention is weight percentage, unless otherwise indicated except.
Embodiment one: the sieve of lactobacillus plantarum (Lactobacillus plantarum) WZMX-2CGMCC No.12832
It selects, isolate and purify and identifies
(1) it screens: according to the particularity of the geographical environment in Xinjiang, passing through the self-control cheese that acquires from Xinjiang South Mountain, Kuerle
It is sampled in bergamot pear, filters out the well-grown microbial strains of a batch, take cheese and each 25g of bergamot pear, be put into 225mL salt water,
It is sufficiently mixed on vortex mixer uniformly, 1mL is taken to be put into the test tube equipped with 9mL sterile saline.It is carried out with 10 times of grades
Dilution, dilution 10-2、10-3、10-4、10-5、10-6.Above-mentioned each 1mL of dilution sample liquid is drawn with pipettor, using tilt-pour process
Modified MRS culture medium is poured into, plate is placed in 37 DEG C of incubators and cultivates for 24 hours.Observe and record colony characteristics and picking have it is transparent
The different shape single bacterium colony of circle carries out Gram's staining, and picking G+ bacteria is fallen, in microscope oil microscopic observation
Cellular morphology, and doubtful strain is numbered.
(2) it purifies: the doubtful bacterial strain for producing transparent circle being inoculated in MRS agar medium, is trained in 37 DEG C of incubator constant temperature
It supports for 24 hours, repeats 3~4 purified lactic acid bacterium.Lactic acid bacteria strains after purification are inoculated in modified MRS inclined-plane, 37 DEG C of incubator constant temperature
After culture for 24 hours, 4 DEG C are saved backup.
The strain culturing condition: 37 DEG C of cultivation temperature；Preferred growth is in MRS media surface, MRS nutrient media components beef
Cream 10.0g/L, yeast extract 5.0g/L, glucose 20.0g/L, sodium acetate 5.0g/L, citric acid hydrogen diamine 2.0g/L, tween
801.0ml/L, dipotassium hydrogen phosphate 2.0g/L, epsom salt 0.2g/L, seven water manganese sulfate 0.05g/L, distilled water 1L, solid training
Feeding base adds agar 20.0g/L.Colony morphology characteristic: bacterial strain colony diameter size on solid medium is 0.5mm-3mm, front
It is rounded, lateral projections；Neat in edge, surface are smooth；Bacterium colony is opaque, is creamy white；With certain toughness.
(3) it identifies:
Physiology and biochemistry identification: the bacterium is separately cultured through MRS culture medium, then can generate molten calcium in improved MRS culture medium
Circle carries out preliminary screening, it is tentatively defined lactic acid bacteria strains；The bacterial strain crosses culture for 24 hours on MRS culture medium flat plate, can shape
Neat at colony edge, the smooth milky protrusion in surface circle, the bacterium colony of diameter 0.5mm-3mm are shown in attached drawing 1.It is blue to carry out leather
Albert'stain Albert, dyeing qualification result are Gram-positive bacillus.The bacterium is negative in catalase test, catalase
It is negative, nitrate can not be restored, do not liquefy gelatin, edwardsiella hoshinae and H2S, it does not move, azymic rhamnose, Neng Gou
15 DEG C of growths, are accredited as lactobacillus plantarum (Lactobacillus plantarum), and the bacterium numbering used is WZMX-2's
Lactobacillus plantarum (Lactobacillus plantarum) and common lactobacillus plantarum (Lactobacillus
Plantarum) there are visibly different strain properties.
16S rDNA sequence alignment and Phylogenetic Analysis: the 16S rDNA sequence and GenBank data that sequencing is obtained
Nucleotide sequence in library carries out BLAST analysis, therefrom obtains similar 16S rDNA sequence, lactobacillus plantarum
The gene order such as SEQUENCE LISTING of (Lactobacillus plantarum) WZMX-2.With in MEGA 6.0
Maximum Likelihood method constructs systematic evolution tree, as a result referring to attached drawing 2.The bacterial strain of bacterial strain WZMX-2 is under the jurisdiction of plant cream
Bacillus (Lactobacillus plantarum), it is nearest with Lactobacillus plantarum parent source relationship, it is homologous similar
Property be 99%, determine that bacterial strain WZMX-2 is lactobacillus plantarum, molecular level Phylogenetic Analysis result all shows that deposit number is
The bacterial strain of CGMCC No.12832 is the novel bacterial in lactobacillus plantarum (Lactobacillus casei), be with it is common common
Lactobacillus plantarum (Lactobacillus casei) have significant difference feature novel bacterial.In conjunction with the strain of above-mentioned offer
Colonial morphology that number is WZMX-2, physio-biochemical characteristics and molecular level identification classification, it is comprehensive in biological classification title
It is recommended that being lactobacillus plantarum (Lactobacillus plantarum).
The bacterial strain was preserved in budapest treaty microorganism International Depository Authority: Chinese microorganism strain before the applying date
Preservation administration committee common micro-organisms center (CGMCC).Address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Chinese section
Institute of microbiology, institute, postcode: 100101.Preservation date is on 08 08th, 2016, and deposit number is CGMCC No.12832.
Embodiment two: the preparation process of whey pear wine
The present invention specifically provides a kind of no bitter taste whey pear wine, prepares especially by the preparation method provided as follows:
(1) activation of strain: by the lactobacillus plantarum saved at 4 DEG C (Lactobacillus plantarum) WZMX-
2CGMCC No.12832 is inoculated in MRS culture medium in an aseptic environment, is placed in 37 DEG C of cultures for 24 hours；The mark that will be saved at 4 DEG C
Think Kluyveromyces sp (Kluyveromyces marxianus) WWMJ-1CGMCC No.8432 and common S. cervisiae
It is inoculated in YPD culture medium and activates respectively in an aseptic environment, be placed in 28 DEG C of cultures for 24 hours, constant-temperature preserving is spare.
(2) bergamot pear pre-processes: selecting fresh Kuerle delicious pear that handle is gone to clean, filters after being squeezed the juice with automatic juice extractor, obtains fragrant
Pear juice is stored spare at room temperature.
(3) whey pre-processes: being added after selecting fresh whey to remove impurity and foreign matter in whey with four layers of filtered through gauze
The amount of 6%-14% v sucrose, mixing be preheating to 85 DEG C, homogeneous 5min under the conditions of 20MPa；By whey liquid after homogeneous
Pasteurize is carried out, is cooled to room temperature, sterile preservation is spare.
(4) preparation of whey bergamot pear juice mixed liquor: the bergamot pear juice and 90% debulking step (3) that step (2) are obtained obtain
Whey by bergamot pear juice account for mixeding liquid volume 40%-60% be uniformly mixed, save backup at room temperature.
(5) preparation of female fermentation liquid: volume percentage is pressed, the whey that (3) the step of 10% obtain is sub-packed in hair
In ferment container, aseptically, the lactobacillus plantarum (Lactobacillus plantarum) that will be activated by step (1)
WZMX-2CGMCC No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1CGMCC
No.8432 and common S. cervisiae are respectively connected in round by 3% inoculative proportion, and condition of culture successively distinguishes 37 DEG C
Culture for 24 hours, 28 DEG C culture for 24 hours, 28 DEG C culture for 24 hours, constant-temperature preserving is spare.
(6) it ferments: lactobacillus plantarum (Lactobacillus plantarum) WZMX-2CGMCC prepared by step (5)
No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1 and saccharomyces cerevisiae fermented liquid
After mixing according to volume ratio 4:4:2 meter, it is added in the whey bergamot pear juice mixed liquor of the residue 90% of step (4) preparation,
- 72h for 24 hours, fermentation termination setting are cultivated at 25 DEG C -37 DEG C are as follows: measurement alcoholic strength reaches 7.8%vol, and measurement total acid reaches
60.84 ° of T, measurement total sugar content reach 42.34g/L, and measurement bitter taste peptide content reaches 0.15g/100mL.
(7) post-process: after product to fermentation termination, flash distillation sterilization carries out sterile filling after cooling and prepares no hardship
Taste whey pear wine.
The present invention is selected from sieve strain: Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-
1CGMCC No.8432, the bacterial strain were preserved in budapest treaty microorganism International Depository Authority before the applying date: China is micro-
Biological inoculum preservation administration committee common micro-organisms center (CGMCC).Address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3
Number, Institute of Microorganism, Academia Sinica, postcode: 100101.Preservation date is on November 05th, 2013, and deposit number is CGMCC
No.8432 uses the condition of culture of the strain for 28 DEG C of cultivation temperature, and preferred growth is in YPD media surface, YPD culture medium
Component uses yeast extract 10g/L, peptone 20g/L, glucose 20g/L, and 20g/L agar powder is added in solid medium.Meanwhile this
Invention provides bacterial strain Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1CGMCC No.8432's
Zymotechnique: aseptically, the picking bacterial strain is cultivated for 24 hours at 28 DEG C.
Outsourcing S. cervisiae of the present invention selects outsourcing S. cervisiae condition of culture and fermentation condition all to use known neck
Domain provides, and bacterial strain selects activity strong, and acid resistance is strong, and production alcohol ability is strong, and fermentation period is short.
In the present invention, alcoholic strength measurement is measured after obtaining alcohol water blend using distillation with alcohol meter；Acidity assaying uses
Determination of acid-basetitration；Pol measurement is measured using hand-held saccharometer: bitter peptides measurement is measured using chloroform-methanol extraction.
The process flow chart that composite bacteria mixed fermentation prepares whey fragrant pear wine is as shown in Fig. 3.
Embodiment three: fermentation condition single factor experiment in the zymotechnique of whey pear wine
Inoculative proportion (lactobacillus plantarum WZMX-2/%: Kluyveromyces marxianus bacterium/%: saccharomyces cerevisiae is chosen respectively
Bacterium/%) be 6:6:1,5:5:1.5,4:4:2,3:3:2.5,2:2:3, cultivation and fermentation temperature be 25 DEG C, 28 DEG C, 31 DEG C, 34 DEG C,
37 DEG C, fermentation time be for 24 hours, 48h, 72h, 96h, 120h, 144h, sugared additive amount be 6%, 8%, 10%, 12%, 14%；Into
Row single factor test fermentation test carries out single factor exploration by measuring alcoholic strength, bitter peptides, acidity, pol.
(1) influence of the different vaccination ratio to fermentation
As attached drawing 4 it is found that the alcohol content of fermentation liquid is higher when the ratio shared by the barms is more.When inoculation ratio
Example (lactobacillus plantarum WZMX-2/%: Kluyveromyces marxianus bacterium/%: S. cervisiae/%) ratio is 4:4:2, fermentation liquid
Alcohol content be up to 7.5%vol, pol is lower with acidity, illustrates that a large amount of sugar and acid are all converted into alcohol, and bitter
Gustin content 0.12g/mL.Comprehensively consider, for guarantee whey pear wine mouthfeel, therefore selection connect bacterium ratio (lactobacillus plantarum
WZMX-2/%: Kluyveromyces marxianus bacterium/%: S. cervisiae/%) ratio 4:4:2 is the most appropriate.
(2) influence of the different fermentations temperature to fermentation
5 as can be seen that the total reducing sugar of fermentation liquid is gradually decreased with the raising of fermentation temperature with reference to the accompanying drawings, and acidity gradually rises
Height, when temperature is 31 DEG C, fermentation liquid alcohol content reaches up to 7.7%vol, and bitter peptides are at 28 DEG C and 31 DEG C, content
Reach lower value, but when temperature it is excessively high more than 31 DEG C when, alcoholic strength is declined, and bitter peptides gradually rise, and temperature, which is crossed, makes strain
Breeding and metabolic capability decline instead, the accumulation of alcohol gradually decreases, comprehensive ferment local-flavor and mouthfeel, thus select 31 DEG C for
Optimum fermentation temp the most.
(3) influence of the different fermentations time to fermentation
By attached drawing 6 it is found that with fermentation time extension, saccharomycete obtained sufficient growth, alcoholic strength and total acid by
Edge up height.In -72h for 24 hours, the alcohol content of fermentation liquid is gradually risen, and bitter taste peptide content is gradually decreased with total reducing sugar, fermentation time
To be inflection point when 72h, bitter taste peptide content reaches minimum 0.72g/100mL in fermentation liquid, and it is 6.5% that alcoholic strength, which reaches peak,
Vol, illustrate fermenting microbe growth it is vigorous, can make full use of carbon source, be translated into alcohol, when fermenting between in 72h-120h
When, bitter peptides, total acid gradually rise, and the alcohol accumulation of fermentation liquid is substantially reduced, and is comprehensively considered, select fermentation time for
72h is best.
(4) influence of the different sugar additive amount to fermentation
During the fermentation, sugared additive amount provides carbon source for saccharomycete and lactic acid bacteria, and sufficient carbon source enhances bacterial strain
Fermentability.By attached drawing 7 it is found that total reducing sugar and alcohol content are increased with the raising of sugared additive amount, sugared additive amount is in 8%-
When between 10%, alcoholic strength accumulation is gradually increased, and it is 0.36g/100mL that bitter taste peptide content, which reaches minimum value, but works as sugaring amount
When more than 10%, alcohol content tends to be steady, and bitter peptides are increased, and strain fermentation ability is gentle and has a declining tendency, can
It can be the sugared excessive concentration in whey pear juice fermentation liquid, it is suppressed that saccharomycete and lactic acid bacteria because addition cane sugar content is excessively high
Fermentation, so that alcoholic strength declines, the higher reason of pol.Therefore, select for the above reasons sugaring amount for 10% the most
Appropriately.
Example IV: the optimization for preparing zymotechnique of whey pear wine
On the basis of the single factor experiment result that embodiment three provides, set according to the center combination design of Box-Behnken
Principle is counted respectively to three factors for influencing fermentation bitter taste peptide content (Z) and alcoholic strength (Y): fermentation time (A), fermentation temperature
(B), inoculative proportion (lactobacillus plantarum WZMX-2/%: Kluyveromyces marxianus bacterium/%: S. cervisiae/%) (C) is utilized
8.05 software of response surface software Design-Expert designs according to Box-Behnken test combinations and carries out Three factors-levels
Experiment.Factor and level code the value such as table 1 of response surface experiments.
The factor and level code value of 1 response surface experiments of table
The response surface optimization that ferments is analyzed: comprehensive single factor experiment is as a result, using Box-Behnken Responds Surface Methodology to it
It optimizes.Response surface experiments design and response the results are shown in Table 2.
2 response surface experiments of table and response
Multiple regression equation fitting is carried out according to 2 test data of table, can be established with fermentation liquid alcoholic strength (Y) and bitter peptides
(Z) respectively to the fit equation of fermentation time (A), fermentation temperature (B), inoculative proportion (C) are as follows:
Y=7.74+0.62A+0.46B+0.47C-0.410AB-0.087AC-0.41BC-1.08A2-0.7B2-0.68C2
Z=0.14-9.732E-003A+0.06B+0.037C-0.12AB+0.043AC-0.039BC+0 .12A2+0.1B2+
0.12C2
3 alcoholic strength response surface experiments result of table and variance analysis
Note: in table 3 and table 4, " * " indicates significant (0.01 < p < 0.05)；" * * " indicates extremely significant (p < 0.01).
4 bitter peptides response surface experiments result of table and variance analysis
The conspicuousness that each variable influences index (response) in regression equation is determined, the value of probability P is got over by F inspection
Small, then the significance degree of relevant variable is higher.By table 3, table 4 it is found that when model F value is respectively F=22.94, F=32.30, P <
0.0001, illustrate that model is extremely significant.When the quasi- item of the mistake of alcoholic strength and bitter peptides is respectively F=1.63, P=0.3021 >
It is not significant that 0.05, F=3.85, P=0.0826 > 0.05 illustrates that model loses quasi- item.The coefficient of determination is respectively R2=0.9538, R2=
0.9667 correction coefficient is respectively R2 Adj=0.9122, R2 Adj=0.9368 shows have well between measured value and predicted value
Degree of fitting.It is possible thereby to which the foundation for illustrating model illustrates that the fitting degree of this model is preferable, test operation accurately may be used in conspicuousness
Letter, therefore can use this model and whey pear juice fermentation process is analyzed and predicted.
From table 3 it is observed that first order and quadratic term show extremely significant water to the Y value influence of whey pear juice fermentation
Flat, influence of interaction item AB, the BC to whey pear juice fermentation Y value shows as the level of signifiance, shadow of the interaction item AC to whey pear wine Y value
Ring the influence without the level of signifiance.The Z value of first order B, C and quadratic term to whey pear juice fermentation liquid as can be seen from Table 4
Influence shows significant and extremely significant level, and significant water is presented in the influence of interaction phase AB, AC, BC to whey pear juice fermentation liquid Z value
It is flat.
Response surface map analysis: in order to investigate influence of each interactive item to fermentation liquid alcoholic strength and bitter peptides, other because
In the case that element immobilizes, operation is carried out to regression equation using 8.05 software of Design-Expert, makes interactive item
Response surface figure.Three-dimensional response surface figure can more intuitively explain the influence between each variable and variable to response.
With the increase of inoculative proportion ratio it can be seen from attached drawing 8-13, alcoholic strength is in downward trend after first rising,
Fermentation time influences obvious significant interaction between fermentation time and inoculative proportion ratio on alcoholic strength；Alcoholic strength is with hair
The raising of ferment temperature and inoculative proportion ratio is made in downward trend after first rising, the interaction of fermentation temperature and inoculative proportion ratio
With significant；With the raising of fermentation temperature, alcoholic strength is in first to rise the trend reduced afterwards, the two significant interaction.
It can be seen that by attached drawing 14-19, with the increase of inoculative proportion ratio, bitter peptides are in the trend risen after falling before,
Contour levels off to ellipse, and inoculative proportion is affected to bitter peptides, and fermentation time influences obvious, fermentation time on bitter peptides
With the significant interaction between inoculative proportion ratio；Bitter peptides are in fall before with the raising of fermentation temperature and inoculative proportion ratio
The trend risen afterwards, curved surface radian is steeper, the significant interaction of fermentation temperature and inoculative proportion ratio；With fermentation temperature
It increases, bitter peptides are in first to reduce the trend risen afterwards, the two significant interaction.
Alcoholic fermentation alcoholic strength is maximized respectively, bitter taste peptide content is minimized, by software Design-Expert
8.05 automatically analyze optimal conditions of fermentation theoretical value can be obtained are as follows: inoculative proportion (lactobacillus plantarum WZMX-2/%: Marx gram
Shandong tie up saccharomycete/%: S. cervisiae/%) be 4:4:2, fermentation time 75.8h, fermentation temperature are 31 DEG C, surveyed alcoholic strength
For 7.8% (V/V), bitter taste peptide content is 0.14g/100mL.Consider that practical operation is convenient, selection inoculative proportion is 4:4:2, sends out
The ferment time is 76h, fermentation temperature is 31 DEG C, carries out 3 parallel tests, and alcoholic strength is 8% (V/V), 7.9% (V/V) and 7.6%
(V/V).Bitter peptides value is 0.13g/100mL, 0.15g/100mL, 0.17g/100mL, approaches with theoretical expectation values, illustrates equation
Good with actual conditions fitting, the obtained Optimized model of response surface analysis is that reliably, the mathematical model is to optimization whey pears
Wine fermentation process conditions be it is feasible, have practical value.
On the basis of single factor experiment, using response surface software, single factor experiment is carried out, and utilize response surface software point
Analysis, show that alcoholic fermentation optimal conditions of fermentation is respectively as follows: inoculative proportion (lactobacillus plantarum (Lactobacillus
Plantarum) WZMX-2CGMCC No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus)
The volume ratio of WWMJ-1CGMCC No.8432 and S. cervisiae) it is 4:4:2, fermentation temperature is 31 DEG C, fermentation time 76h,
Sugared additive amount is 10%.Alcoholic strength average out to 7.8%vol with this condition, bitter taste peptide content are 0.15g/100mL.It is verified
The measured actual acidity of test and theoretical value difference are smaller, it was demonstrated that two mathematical models are to optimization whey pear wine zymotechnique item
Part is feasible, provides theoretical foundation for comprehensive utilization whey.
The whey pear wine prepared by above-mentioned campaign effectively removes acidified milk pure mellow wine bitter taste, overcomes the prior art
The whey pear wine of the middle technical bottleneck that apparent " rear bitter taste " is had using whey converted products, preparation has typical case as product
Property is as follows:
(1) organoleptic indicator: color: pistac, micro- yellow color for having whey intrinsic；Fragrance: frankincense taste is along with light
Light fruit aroma, flavour: wine body gracefulness, sour-sweet tasty and refreshing, free from extraneous odour；Posture: the bright a little precipitating of clarification.
(2) physical and chemical index: total acid: 60.84 ° of T, total sugar content 42.34g/L.
(3) microbiological indicator: total plate count (cfu/mL)≤100；Coliform (MPN/100mL)≤3；Pathogenic bacteria: not
Detection.
Embodiment five: the typicalness subjective appreciation of the whey pear wine of preparation
1. sensory evaluation criteria
By being all from the sensory evaluation group progress data sampling for preparing different 20 people of technical staff in fruit wine field and forming,
The whey pear wine of the technical solution preparation provided using embodiment two to four is coordinated from mouthfeel, color, mellowness, sweet tea, acid, fragrance
Property aspect carry out overall merit, organoleptic indicator's evaluation criterion of whey pear wine is as shown in table 5.
5 whey pear juice fermented wine sensory evaluation scores table of table
2. Fuzzy Evaluation
Evaluation domain (U), comment domain (V), weight vectors (A) are determined according to fuzzy evaluation, and to Analyses Methods for Sensory Evaluation Results
It is handled, obtains fuzzy matrix for assessment, then by calculating Comprehensis pertaining, finally obtain whey pear wine aesthetic quality synthesis and comment
The result vector sentenced, and carry out grade classification.
2.1 determine evaluation domain
Evaluation domain refers to the set that can most reflect one group of index of the food quality, is denoted as: U={ u1, u2, u3 ...
Un }, u indicates evaluation index.Mouthfeel, color, mellowness, sweet tea, sour fragrance harmony is chosen to refer to as the sensory evaluation of whey pear wine
Mark.It selects 10 people of Majors of Food and forms subjective appreciation group.Mouthfeel can be indicated with u1, u2 indicates that color, u3 indicate mellowness, u4
Indicate that sweet tea, u5 indicate sour fragrance harmony, obtain the evaluation domain U={ u1, u2, u3, u4, u5 } about whey pear wine.
2.2 determine comment domain
According to the quality grade evaluation standard of table 5, obtain one about whey pear wine comment domain V=v1, v2, v3,
v4}。
2.3 determine weight vectors
Weight vectors relevant to whey pear wine are obtained according to the weighted value of indices in table 1 are as follows: A=(0.30,
0.10,0.10,0.10,0.10,0.30)。
2.4 determine fuzzy matrix for assessment
Evaluating member is as shown in table 6 to the evaluation result of whey pear wine.
The results of sensory evaluation of 6 whey pear wine of table
The data of table 6 are all obtained into fuzzy matrix for assessment respectively divided by panelist's total number of persons 20:
3. calculating Comprehensis pertaining
The weight vectors of known 6 evaluation indexes of whey pear wine be A=(0.30,0.10,0.10,0.10,0.10,
0.30), according to the fuzzy comprehensive evoluation of standard, replace max min algorithm with ordinary channel multiplication and according to comprehensive score formula
Calculate comprehensive score.The Comprehensis pertaining of whey pear wine is Y=A × R=(b1, b2, b3 ... bn), and whey pear wine sense organ can be obtained
The result vector that quality comprehensive is judged:
Wherein:
Y1=0.3 × 15/20+0.10 × 13/20+0.10 × 14/20+0.10 × 16/20+0.10 × 13/20+0.30 ×
15/20=0.73.
Y2=0.3 × 3/20+0.10 × 5/20+0.10 × 3/20+0.10 × 3/20+0.10 × 4/20+0.30 × 4/20
=0.18.；
Y3=0.3 × 2/20+0.10 × 2/20+0.10 × 3/20+0.10 × 1/20+0.10 × 2/20+0.30 × 1/20
=0.085.；
Y4=0.3 × 0+0.10 × 0+0.10 × 0+0.10 × 0+0.10 × 1/20+0.30 × 0=0.005.
That is: Y1=(0.73,0.18,0.085,0.005)
According to comprehensive score formula:Calculate comprehensive score:
H1=1 × 0.73+2 × 0.18+3 × 0.085+4 × 0.005=1.335；
The comprehensive score of whey pear wine close to 1, illustrates whey pear wine quality between becoming reconciled very well partially between 1 and 2
To in fine.
Fuzzy mathematics subjective appreciation is integrated it is found that whey pear wine quality is in whey pears that are fine, brewing with mixed bacteria
Wine quality characteristic are as follows: acidity is low, and total reducing sugar is within the scope of national standard, and wine degree is higher than simple commercial yeast, has whey pear juice
The characteristic feature pistac of wine sample, bright color, with whey outstanding and pear juice fruit flavor, no bitterness sense, mouthfeel is inclined
Sweet tea, mild acidity, harmony is moderate, and mellowness is simple and elegant, and mouthfeel is plentiful, has good harmony.To sum up, selected by the application
Mixed bacteria be suitable for fermenting no bitter taste whey pear wine fermentation strain, a kind of composite bacteria mixed fermentation provided by the present application
Method of the preparation without bitter taste whey pear wine effectively reduces whey pear wine bitter taste peptide content, while in the mouthfeel for promoting whey pear wine
Outstanding contributions are made that in terms of flavor.
Embodiment six: the whey pear wine functional trial of preparation
It chooses KM mouse and is randomly divided into 4 big group, be further divided into 5 groups per big group, every group 10.By basic, normal, high dosage,
I.e. each group mouse is respectively daily 8.4mL/kg.bw, 16.7mL/ using whey pear wine amount provided by the above embodiment
kg.bw,50.1mL/kg.bw.Since stomach-filling amount is larger, the stomach-filling requirement of mouse is not met, so with 20 times of concentrates of sample,
Through conversion dilution 8 times after be used as high dose stomach-filling sample liquid, with high dose group stomach-filling sample liquid handle after wine base be configured to it is low, in
Stomach-filling sample liquid used in dosage group, stomach-filling volume is based on 20mL/kg.bw.It is by the treatment that alcoholic strength is 48% (V/V)
8% (V/V), wine base control group stomach-filling isometric 8% (V/V) wine base, the isometric deionized water of blank control group stomach-filling.It is daily to fill
Stomach is primary, continuous 30d.
Mice burden swimming experiment, the experiment of mice serum urea, mouse hepatic glycogen reality are carried out to each group experiment mice respectively
It tests and is tested with blood lactase acid, measure related experiment parameter.
Table 7: influence of the whey pear wine to swimming with a load attached to the body experiment mice weight and swimming time
By table 7 as it can be seen that there was no significant difference (P > 0.05) for weight of each dosage to mouse in Loaned swimming test, and
The walking weight load of high dose group mouse extends (P < 0.05) than control group conspicuousness.
Table 8: influence of the whey pear wine to serum urea experiment mice weight and serum urea
By table 8 as it can be seen that there was no significant difference (P > 0.05) for weight of each dosage to mouse in serum urea experiment, and
The serum urea content conspicuousness of high dose group mouse is lower than control group (P < 0.05).
Table 9: influence of the whey pear wine to hepatic glycogen experiment mice weight and hepatic glycogen content
By table 9 as it can be seen that there was no significant difference (P > 0.05) for weight of each dosage to mouse in hepatic glycogen experiment, and it is high
The hepatic glycogen content conspicuousness of dosage group mouse is higher than control group (P < 0.05).
Table 10: influence of the whey pear wine to blood lactase acid experiment mice weight
By table 10 as it can be seen that in blood lactase acid experiment weight of each dosage group of whey pear wine to mouse there was no significant difference (P >
0.05)。
Table 11: the influence of three time point Serum lactic acid contents before and after whey pear wine swims to blood lactase acid experiment mice
By table 11 as it can be seen that whey pear wine various dose group mouse is before swimming in blood lactase acid examination experiment, after swimming, trip
There was no significant difference (P > 0.05) for the Serum lactic acid content of 20min after swimming.
Table 12: influence of the whey pear wine to swimming mouse blood lactase acid area under the curve
By table 12 as it can be seen that in blood lactase acid experiment, the blood lactase acid area under the curve of each dosage group swimming mouse of whey pear wine is equal
There was no significant difference (P > 0.05).
To sum up result: experiment shows the whey pear wine sample of orally administration mouse various dose, to the body weight increase of mouse
It has no adverse effects, the mice burden swimming time can be obviously prolonged, the generation of serum urea, increases the storage of hepatic glycogen when reducing movement
It is standby, the weight and blood lactase acid of mouse are had no significant effect, therefore whey pear wine has function of relieving physical fatigue effect.
The preparation method of lactobacillus plantarum WZMX-2 provided by the invention and its whey pear wine of application
It is high production efficiency, no added, be effectively improved acidified milk pure mellow wine bitter taste, the whey pear wine uniform color prepared, in pistac,
Clarify it is bright, with whey and bergamot pear intrinsic color and fragrance, no bitterness sense, the inclined sweet tea of mouthfeel, mild acidity, harmony is suitable
In, mellowness is simple and elegant, and mouthfeel is plentiful, has good harmony.Orally administration mouse carries out Loaned swimming test, and serum urea is surveyed
Fixed test, hepatic glycogen measurement test and blood lactase acid measurement test, whey pear wine can be obviously prolonged mice burden swimming as the result is shown
Time, reduce movement when serum urea generation, increase the deposit of hepatic glycogen, the weight and blood lactase acid to mouse are without obvious shadow
It rings, shows that whey pear wine has function of relieving physical fatigue effect.
As described above, the present invention can be realized preferably, the above embodiments are only to preferred implementation side of the invention
Formula is described, and is not intended to limit the scope of the present invention, and without departing from the spirit of the design of the present invention, this field is general
The various changes and improvements that logical technical staff makes technical solution of the present invention, should all fall into present invention determine that protection scope
It is interior.
SEQUENCE LISTING
<110>Xinjiang Agricultural Univ
<120>the whey pear wine of lactobacillus plantarum WZMX-2 and application and its preparation
<130> WZMX-2
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 1440
<212> DNA
<213> WZMX-2
<220>
<221> WZMX-2
<222> (1)..(1440)
<400> 1
agtcgacgag ctctggtatt gattggtgct tgcatcatga tttacatttg agtgagtggc 60
gaactggtga gtaacacgtg ggaaacctgc ccagaagcgg gggataacac ctggaaacag 120
atgctaatac cgcataacaa cttggaccgc atggtccgag tttgaaagat ggcttcggct 180
atcacttttg gatggtcccg cggcgtatta gctagatggt ggggtaacgg ctcaccatgg 240
caatgatacg tagccgacct gagagggtaa tcggccacat tgggactgag acacggccca 300
aactcctacg ggaggcagca gtagggaatc ttccacaatg gacgaaagtc tgatggagca 360
acgccgcgtg agtgaagaag ggtttcggct cgtaaaactc tgttgttaaa gaagaacata 420
tctgagagta actgttcagg tattgacggt atttaaccag aaagccacgg ctaactacgt 480
gccagcagcc gcggtaatac gtaggtggca agcgttgtcc ggatttattg ggcgtaaagc 540
gagcgcaggc ggttttttaa gtctgatgtg aaagccttcg gctcaaccga agaagtgcat 600
cggaaactgg gaaacttgag tgcagaagag gacagtggaa ctccatgtgt agcggtgaga 660
tgcgtagata tatggaagaa caccagtggc gaaggcggct gtctggtctg taactgacgc 720
tgaggctcga aagtatgggt agcaaacagg attagatacc ctggtagtcc ataccgtaaa 780
cgatgaatgc taagtgttgg agggtttccg cccttcagtg ctgcagctaa cgcattaagc 840
attccgcctg gggagtacgg ccgcaaggct gaaactcaaa ggaattgacg ggggcccgca 900
caagcggtgg agcatgtggt ttaattcgaa gctacgcgaa gaaccttacc aggtcttgac 960
atactatgca aatctaagag attagacgtt cccttcgggg acatggatac aggtggtgca 1020
tggttgtcgt cagctcgtgt cgtgagatgt tgggttaagt cccgcaacga gcgcaaccct 1080
tattatcagt tgccagcatt aagttgggca ctctggtgag actgccggtg acaaaccgga 1140
ggaaggtggg gatgacgtca aatcatcatg ccccttatga cctgggctac acacgtgcta 1200
caatggatgg tacaacgagt tgcgaactcg cgagagtaag ctaatctctt aaagccattc 1260
tcagttcgga ttgtaggctg caactcgcct acatgaagtc ggaatcgcta gtaatcgcgg 1320
atcagcatgc cgcggtgaat acgttcccgg gccttgtaca caccgcccgt cacaccatga 1380
atcagcatgc cgcggtgaat acgttcccgg gccttgtaca caccgcccgt cacaccatga 1440
Claims (4)
1. a kind of lactobacillus plantarum (Lactobacillus plantarum) WZMX-2, which is characterized in that the plant cream bar
The deposit number of bacterium (Lactobacillus plantarum) WZMX-2 is GCMCC No.12832,16S rDNA gene sequence
Column are as shown in SEQ ID NO.1.
2. lactobacillus plantarum (Lactobacillus plantarum) WZMX-2 as described in claim 1 is in whey pear wine system
Application in standby.
3. a kind of preparation method of whey pear wine, which is characterized in that adopt the following technical scheme that
(1) activation of strain: by the lactobacillus plantarum saved at 4 DEG C (Lactobacillus plantarum) WZMX-2
CGMCC No. 12832 is inoculated in MRS culture medium in an aseptic environment, is placed in 37 DEG C of cultures for 24 hours；The mark that will be saved at 4 DEG C
Think Kluyveromyces sp (Kluyveromyces marxianus) WWMJ-1 CGMCC No.8432 and common S. cervisiae
It is inoculated in YPD culture medium and activates respectively in an aseptic environment, be placed in 28 DEG C of cultures for 24 hours, constant-temperature preserving is spare；
(2) bergamot pear pre-processes: it selects fresh Kuerle delicious pear that handle is gone to clean, is filtered after being squeezed the juice with automatic juice extractor, obtain bergamot pear juice,
It stores at room temperature spare；
(3) whey pre-processes: 6%- is added after selecting fresh whey to remove impurity and foreign matter in whey with four layers of filtered through gauze
The amount of 14% v sucrose is uniformly mixed, is preheating to 85 DEG C, homogeneous 5min under the conditions of 20MPa；Whey liquid is carried out after homogeneous
Pasteurize is cooled to room temperature, and sterile preservation is spare；
(4) preparation of whey bergamot pear juice mixed liquor: the whey that the bergamot pear juice and 90% debulking step (3) that step (2) are obtained obtain
Mixeding liquid volume 40%-60% is accounted for by bergamot pear juice to be uniformly mixed, and is saved backup at room temperature；
(5) preparation of female fermentation liquid: volume percentage is pressed, the whey that (3) the step of 10% obtain is sub-packed in round
In, aseptically, lactobacillus plantarum (Lactobacillus plantarum) WZMX-2 that will be activated by step (1)
CGMCC No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1 CGMCC
No.8432 and common S. cervisiae are respectively connected in round by 3% inoculative proportion, and condition of culture successively distinguishes 37 DEG C
Culture for 24 hours, 28 DEG C culture for 24 hours, 28 DEG C culture for 24 hours, constant-temperature preserving is spare；
(6) it ferments: lactobacillus plantarum (Lactobacillus plantarum) WZMX-2 CGMCC prepared by step (5)
No.12832, Kluyveromyces marxianus bacterium (Kluyveromyces marxianus) WWMJ-1 and saccharomyces cerevisiae fermented liquid
After mixing according to volume ratio 4:4:2 meter, it is added in the whey bergamot pear juice mixed liquor of the residue 90% of step (4) preparation, 25
- 72h for 24 hours, fermentation termination setting are cultivated at DEG C -37 DEG C are as follows: measurement alcoholic strength reaches 7.8%vol, and measurement total acid reaches 60.84 °
T, measurement total sugar content reach 42.34g/L, and measurement bitter taste peptide content reaches 0.15g/100mL；
(7) post-process: after product to fermentation termination, flash distillation sterilization carries out sterile filling and prepares no bitter taste cream after cooling
Clear pear wine.
4. a kind of whey pear wine, which is characterized in that obtain whey pear wine using preparation method as claimed in claim 3.
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Effective date of registration: 20230410 Address after: 841000 South side of Anchen Avenue and east side of Jinhe Road in Korla Economic and Technological Development Zone, Bayingolin Mongolian Autonomous Prefecture, Xinjiang Uygur Autonomous Region Patentee after: XINJIANG KAIRUIKE FOOD TECHNOLOGY CO.,LTD. Address before: 830011 No.42 Nanchang Road, shayibak District, Urumqi, Xinjiang Uygur Autonomous Region Patentee before: XINJIANG AGRICULTURAL University Patentee before: Wu Yun |
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Denomination of invention: Lactobacillus plantarum WZMX-2 and its application in the preparation of whey pear wine Granted publication date: 20190716 Pledgee: China Construction Bank Corporation Bayingolin Mongolian Autonomous Prefecture Branch Pledgor: XINJIANG KAIRUIKE FOOD TECHNOLOGY CO.,LTD. Registration number: Y2024980040373 |