CN109439548A - A kind of strengthening porcelain and Preparation method and use that can generate tyrosol and hydroxytyrosol - Google Patents
A kind of strengthening porcelain and Preparation method and use that can generate tyrosol and hydroxytyrosol Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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Abstract
The present invention discloses a kind of strengthening porcelain and Preparation method and use that can generate tyrosol and hydroxytyrosol, belongs to Liquor Making Technology field, the preparation method of the yeast be include (1) screening strain, (2) cultivate strain and (3) make strengthening porcelain.The present invention screens the peculiar microorganism for generating tyrosol and hydroxytyrosol active secondary metabolites, prepares strengthening porcelain by forced fermentation method, during strengthening porcelain is applied to brewed spirit, produces the white wine rich in tyrosol and the hydroxytyrosol healthy functions factor.
Description
Technical field
The invention belongs to Liquor Making Technology fields, and in particular to a kind of strengthening porcelain that can generate tyrosol and hydroxytyrosol
And Preparation method and use.
Background technique
China white wine brewing has very long history.The White wine brewing process and distillation technique of more bacterium spontaneous fermentations have
Unique skill and superb level, be the national heritage of China's preciousness.With the raising of Chinese national economy level, consumption
Person increasingly focuses on the relationship between food and health.China white wine lacks always the health that bioactivity is significant, from the horse's mouth
The factor is difficult to incorporate international consumption market.The characteristics of China white wine is made is use nature koji-making, is mainly micro- life with yeast
The open production technology of the multi-cultur es co-fermentation in object source.Separation and directed screening metabolism have wide from brewing microorganism
Health factor is brought into Liquor Products by the secondary metabolite of general bioactivity by fermentation method, is brewed rich in health
The Liquor Products of active constituent are the quick Critical policies for pushing China white wine development.
Tyrosol and hydroxytyrosol have extensive bioactivity, and fool proof, at home and abroad food, health products trade tool
There are very high popularity and significant impact power.The bioactivity of tyrosol and hydroxytyrosol mainly has the following aspects: (1)
2016, Stiuso, Paola et al. are in international bio chemistry publication " International Journal of Molecular
Sciences " independent academic paper is delivered, determine that tyrosol is protected with significant liver protection from cell, gene, protein, animal level
Liver effect.The same year, Sarna, Lindsei K. et al. give to the world the result of one's studies on International Publication " lipids ", determine that tyrosol can
Hepatic injury caused by high-fat foods is reduced, further determines that the liver protecting effect of the compound.(2) epidemiological study table
Bright, the food for being rich in the two compounds is used for a long time in the crowd of Mediterranean Region, and the disease incidence of vascular diseases significantly reduces,
Others research also indicates that the two compounds have cardiovascular protective effect.(3) a large amount of International Academic paper determine this two
A compound all has significant antioxidant activity, can prevent the disease incidence of the various diseases of human body, delays senescence.(4) prevent
There is obvious action in terms of the generation of cancer and control cancer metastasis.(5) mitochondrial function disorder is prevented, flesh is prevented
Meat atrophy.(6) inhibit lethal nerve degeneration caused by prion, inhibit beta-amyloid protein to the toxicity of nerve cell, protect
Nerve cell is protected, senile dementia is prevented.
Currently, producing Tetramethylpyrazine functional microorganism mainly around screening, and apply it to white in brewed spirit field
In wine production, the content of Tetramethylpyrazine active constituent in white wine is improved, there is many patents and document report.But there is not yet
Breeding produces tyrosol and hydroxytyrosol function bacterium from brewing microorganism, is applied to brewed spirit, and production is rich in tyrosol and hydroxyl junket
The patent and document report of alcohol bioactive molecule white wine.
Summary of the invention
In order to solve above-mentioned prior art problem, the invention proposes a kind of reinforcing that can generate tyrosol and hydroxytyrosol is big
Bent and Preparation method and use.
To achieve the goals above, the present invention adopts the following technical scheme:
A kind of preparation method for the strengthening porcelain that can generate tyrosol and hydroxytyrosol, it is characterised in that: the following steps are included:
(1) it screens strain: common yeast culture 5-7 days separate to microorganism pure using microbe separation technology
Change, obtains single culture, the single culture is fermented, the bacterium in fermentation material containing tyrosol and hydroxytyrosol is filtered out
Kind, it is slant strains by the fungi preservation filtered out；
(2) it cultivates strain: the slant strains is inoculated into fluid nutrient medium, carry out culture 24-96 hours, obtain function
It can bacterium seed liquor；
(3) it makes strengthening porcelain: the function bacterium seed liquor being uniformly inoculated on the solid medium of sterilizing, 20-50
DEG C cultivation and fermentation 15-30 days, the raw material fermented is taken out, tyrosol can be generated and the reinforcing of hydroxytyrosol is big up to described by drying in the shade
It is bent.
Further, the function bacterium seed liquor is accounted for the 1- of solid medium weight according to the volume of function bacterium seed liquor
20% ratio uniform is inoculated on the solid medium of sterilizing.
Further, the solid medium the preparation method comprises the following steps: by base-material weight 40% be added 0.7-5.0% egg
White peptone water solution is uniformly mixed, and loaded in 500 milliliters of triangular flasks, 100 grams every bottle, 121 DEG C of sterilizing 30min, the base-material is big
One of rice, wheat and wheat bran are a variety of.
A kind of strengthening porcelain that can generate tyrosol and hydroxytyrosol, is made by above-mentioned preparation method.
A kind of above-mentioned strengthening porcelain that can generate tyrosol and hydroxytyrosol is in white wine of the preparation rich in tyrosol and hydroxytyrosol
In purposes.
A kind of preparation method of the white wine rich in tyrosol and hydroxytyrosol, it is characterised in that: the following steps are included: will be described
The strengthening porcelain that tyrosol and hydroxytyrosol can be generated is uniformly mixed with traditional yeast, then enters pond hair according to traditional wine-making technology
Ferment, distillation takes wine after fermentation, can be obtained the white wine rich in tyrosol and hydroxytyrosol.
Further, the weight ratio of the strengthening porcelain that can generate tyrosol and hydroxytyrosol and traditional yeast is 1:100-
1:1。
Compared with prior art, the beneficial effects of the present invention are:
The present invention screens the peculiar microorganism for generating tyrosol and hydroxytyrosol active secondary metabolites, passes through forced fermentation
Method prepares strengthening porcelain, during strengthening porcelain is applied to brewed spirit, produces rich in tyrosol and hydroxytyrosol health
The white wine of function factor；The structure of tyrosol and hydroxytyrosol is entirely different with Tetramethylpyrazine, and bioactivity also has many differences
It is different, meanwhile, tyrosol and hydroxytyrosol belong to phenylethanol compound, and belonging to homologue and Liquor Products with ethyl alcohol has very
Good compatibility, can be obviously improved the comfortable taste degree of Liquor Products, can generate tyrosol and hydroxyl junket using of the present invention
The white wine mouthfeel of the strengthening porcelain preparation of alcohol is sweet.
Specific embodiment
The present invention will be further described with reference to the examples below, and described embodiment is only present invention a part
Embodiment is not whole embodiment.Based on the embodiments of the present invention, those skilled in the art are not making
Other all embodiments obtained, belong to protection scope of the present invention under the premise of creative work.Used in embodiment
Test material is unless otherwise specified that routine biochemistry reagent shop is commercially available.
Embodiment one
A kind of preparation method for the strengthening porcelain that can generate tyrosol and hydroxytyrosol, comprising the following steps:
(1) culture medium is prepared:
A prepares slant medium: weighing 200g potato, is cut into small pieces, is placed in 1000 milliliters of water, boils 20-30
Minute, residue is filtered off with filter cloth.2% glucose is added in filtrate, agar 15-20g (is done broken) in advance, after heating fusing, adds water
It is settled to 1000mL, pH value is naturally, be sub-packed in test tube, 121 DEG C of sterilizing 30min.
B prepares liquid seed culture medium: weighing 200g potato, is cut into small pieces, is placed in 1000 milliliters of water, boils
20-30 minutes, residue was filtered off with filter cloth.2% glucose, 0.3%KH are added in filtrate2PO4, 0.15%MgSO4·7H2O、
PH is adjusted to neutrality with NaOH aqueous solution, is sub-packed in triangular flask by 0.15% yeast extract, 121 DEG C of sterilizing 30min.
C prepares solid medium: by the peptone water solution of 40% addition 0.7% of rice or wheat weight, mixing is equal
It is even, it is loaded in 500 milliliters of triangular flasks, 100 grams every bottle, 121 DEG C of sterilizing 30min.
(2) it screens strain: taking yeast sample 10g to be added in the triangular flask equipped with 100mL sterile water, add sterile glass
Pearl is placed on shaking table, and 28 DEG C shake 30 minutes.After standing, take bacterium solution according to 10 times of gradient dilutions to 10-6, take each dilution
1mL is put into equipped with being uniformly mixed in the culture dish for being cooled to 50 DEG C of enriched mediums after fusing, 30 DEG C of constant temperature trainings is put in after solidification
It supports case culture 5 days.The bacterium colony of picking growth, and isolate and purify to obtain pure culture by scribing line.Liquid will be inoculated by strain after purification
In body culture medium, cultivate 48 hours, after be transferred to ordinary solid culture medium, be placed in incubator, 30 DEG C cultivate 25 days.Culture is had
The ethyl acetate of 5 times of amounts is added in the ordinary solid culture medium of strain, impregnates 3d under room temperature.With filtered through gauze, 50 DEG C of decompressions are dense
Contracting obtains medicinal extract, with methanol by medicinal extract constant volume to certain volume.Using high-efficient liquid phase chromatogram technology, to the fermentation material after constant volume into
Row content analysis, liquid phase analysis condition are as follows: (microorganism is received in Suzhou by chromatographic column UniSiL 10 μm of -100-C18,4.6 × 250mm
Science and Technology Ltd.)；Detection wavelength 258nm；35 DEG C of column temperature；Flow 0.8mL/min；35% acetonitrile-water isocratic elution, detection knot
Fruit is as shown in table 1.Wherein DH-6 content is higher, further chooses slant medium prepared by the strain that bacterium numbering is DH-6
Save as slant strains.
Table 1 screens obtained strain and produces tyrosol and hydroxytyrosol result (mg/100g culture medium)
(3) strain is cultivated: in desinfection chamber, the cooling liquid of the sterilizing that the strain DH-6 for being stored in inclined-plane is inoculated into preparation
Body seed culture medium, 200r/min, 32 DEG C cultivate 48h, obtain cell concentration and reach 106-107The function bacterium seed of cfu/ml
Liquid.
(4) make strengthening porcelain: by the volume of DH-6 function bacterium seed liquor account for solid medium weight 1%, 5%,
10%, 20% inoculum concentration is respectively connected to DH-6 function bacterium seed liquor to the solid medium of preparation, after shaking up, 30 DEG C of cultures
25d takes out the raw material fermented, the DH-6 strengthening porcelain to dry in the shade up to four kinds of various concentrations, as DH-6 strengthening porcelain -1%,
DH-6 strengthening porcelain -5%, DH-6 strengthening porcelain -10%, DH-6 strengthening porcelain -20%.
Embodiment two
A kind of preparation method for the strengthening porcelain that can generate tyrosol and hydroxytyrosol, comprising the following steps:
(1) culture medium is prepared:
Prepare solid medium: by the peptone water solution of 40% addition 5.0% of rice or wheat weight, mixing is equal
It is even, it is loaded in 500 milliliters of triangular flasks, 100 grams every bottle, 121 DEG C of sterilizing 30min.
(2) it screens strain: taking yeast sample 10g to be added in the triangular flask equipped with 100mL sterile water, add sterile glass
Pearl is placed on shaking table, and 28 DEG C shake 30 minutes.After standing, take bacterium solution according to 10 times of gradient dilutions to 10-6, take each dilution
1mL is put into equipped with being uniformly mixed in the culture dish for being cooled to 50 DEG C of enriched mediums after fusing, 30 DEG C of constant temperature trainings is put in after solidification
It supports case culture 7 days.The bacterium colony of picking growth, and isolate and purify to obtain pure culture by scribing line.Liquid will be inoculated by strain after purification
In body culture medium, cultivate 48 hours, after be transferred to ordinary solid culture medium, be placed in incubator, 30 DEG C cultivate 25 days.Culture is had
The ethyl acetate of 5 times of amounts is added in the ordinary solid culture medium of strain, impregnates 3d under room temperature.With filtered through gauze, 50 DEG C of decompressions are dense
Contracting obtains medicinal extract, with methanol by medicinal extract constant volume to certain volume.Using high-efficient liquid phase chromatogram technology, to the fermentation material after constant volume into
Row content analysis, liquid phase analysis condition is as in the first embodiment, choose tyrosol and hydroxytyrosol yield highest strain embodiment one
The slant medium of preparation saves as slant strains.
(3) it cultivates strain: in desinfection chamber, it is cold that the strain for being stored in inclined-plane being inoculated into the sterilizing prepared in embodiment one
But liquid seed culture medium, 200r/min, 32 DEG C, culture for 24 hours, obtain cell concentration and reach 106-107The function bacterium of cfu/ml
Seed liquor.
(4) it makes strengthening porcelain: accounting for 10% inoculum concentration of solid medium weight to originally by the volume of function bacterium seed liquor
The solid medium access function bacterium seed liquor prepared in embodiment, after shaking up, 25 DEG C of culture 15d take out the raw material fermented,
It dries in the shade up to strengthening porcelain.
Embodiment three
A kind of preparation method for the strengthening porcelain that can generate tyrosol and hydroxytyrosol, comprising the following steps:
(1) culture medium is prepared:
Prepare solid medium: by the peptone water solution of 40% addition 2.5% of rice or wheat weight, mixing is equal
It is even, it is loaded in 500 milliliters of triangular flasks, 100 grams every bottle, 121 DEG C of sterilizing 30min.
(2) it screens strain: taking yeast sample 10g to be added in the triangular flask equipped with 100mL sterile water, add sterile glass
Pearl is placed on shaking table, and 28 DEG C shake 30 minutes.After standing, take bacterium solution according to 10 times of gradient dilutions to 10-6, take each dilution
1mL is put into equipped with being uniformly mixed in the culture dish for being cooled to 50 DEG C of enriched mediums after fusing, 30 DEG C of constant temperature trainings is put in after solidification
It supports case culture 6 days.The bacterium colony of picking growth, and isolate and purify to obtain pure culture by scribing line.Liquid will be inoculated by strain after purification
In body culture medium, cultivate 48 hours, after be transferred to ordinary solid culture medium, be placed in incubator, 30 DEG C cultivate 25 days.Culture is had
The ethyl acetate of 5 times of amounts is added in the ordinary solid culture medium of strain, impregnates 3d under room temperature.With filtered through gauze, 50 DEG C of decompressions are dense
Contracting obtains medicinal extract, with methanol by medicinal extract constant volume to certain volume.Using high-efficient liquid phase chromatogram technology, to the fermentation material after constant volume into
Row content analysis, liquid phase analysis condition is as in the first embodiment, choose tyrosol and hydroxytyrosol yield highest strain embodiment one
The slant medium of preparation saves as slant strains.
(3) strain is cultivated: cold to the sterilizing prepared in the strain inoculation embodiment one on inclined-plane by being stored in desinfection chamber
But liquid seed culture medium, 200r/min, 32 DEG C cultivate 72h, obtain cell concentration and reach 106-107The function bacterium of cfu/ml
Seed liquor.
(4) make strengthening porcelain: by the volume of function bacterium seed liquor account for 10% inoculum concentration of solid medium weight to
The solid medium access function bacterium seed liquor prepared in the present embodiment, after shaking up, 45 DEG C of culture 30d take out the original fermented
Material dries in the shade up to strengthening porcelain.
The present invention also provides the purposes of the yeast of can ferment tyrosol and the hydroxytyrosol, below with specific embodiment for
It is bright.
Example IV
DH-6 strengthening porcelain -10% prepares the purposes in Production of Luzhou-flavor Liquor
In Production of Luzhou-flavor Liquor, according to the weight ratio of 1:10, DH-6 strengthening porcelain -10% is pulverized and mixed with yeast
Uniformly, mixture starter powder is formed.Strengthen song due to having used, mixed starters dosage is suitably reduced when bent under airing, than usual yeast
When few 10% or so, then pit entry fermentation, fermentation time are identical as common process.After fermentation, routinely technique distillation is
It can get healthy functions white wine.High performance liquid chromatography (HPLC) method content measuring shows junket in giving off a strong fragrance white wine that the technique spawns
The content of pure and mild hydroxytyrosol respectively reaches 30-50 mg/litre and 15-30 mg/litre.
Embodiment five
DH-6 strengthening porcelain -5% is preparing the purposes in Maotai-flavor liquor production
In the production of Maotai-flavor liquor, DH-6 strengthening porcelain -5% and paste flavor high-temperature daqu are mixed according to the weight ratio of 1:20
Mixing starter powder is obtained after conjunction, after smashing.Traditionally, bent under starter powder airing to mix, it is close with traditional handicraft with song amount,
Pit entry fermentation, distillation can be obtained the Maotai-flavor liquor containing tyrosol and hydroxytyrosol after fermentation.HPLC method content measuring
Show in the Maotai-flavor liquor spawned using the technique the content of tyrosol and hydroxytyrosol respectively reach 15-25 mg/litre and
10-20 mg/litre.
Embodiment six
DH-6 strengthening porcelain -1% is preparing the purposes in the production of yeast fen-flavor type white spirit
In the production of yeast fen-flavor type white spirit, by DH-6 strengthening porcelain -1% with delicate fragrance type yeast according to the weight of 1:1
After mixing, mixing starter powder is obtained after smashing.Traditionally, bent under starter powder airing to mix, with song amount and traditional handicraft
It is close, enter ground cylinder fermentation, distillation can be obtained the fen-flavor type white spirit containing tyrosol and hydroxytyrosol after fermentation.HPLC method contains
Measurement take temperature it is bright, in the fen-flavor type white spirit spawned using the technique the content of tyrosol and hydroxytyrosol respectively reach 20-35 milligrams/
It rises and 10-20 mg/litre.
Embodiment seven
DH-6 strengthening porcelain -20% is preparing the purposes in distilled spirit with sesame flavour production
In the production of distilled spirit with sesame flavour, by DH-6 strengthening porcelain -20% with the total starter powder of sesame type according to the weight of 1:100
After amount is than mixing, mixing starter powder is obtained after smashing.Traditionally, bent under starter powder airing to mix, with song amount and traditional work
Skill is close, pit entry fermentation, and distillation can be obtained the distilled spirit with sesame flavour containing tyrosol and hydroxytyrosol after fermentation.HPLC method
Content measuring shows that the content of tyrosol and hydroxytyrosol respectively reaches 15-30 milli in the fen-flavor type white spirit spawned using the technique
Grams per liter and 10-25 mg/litre.
Claims (7)
1. a kind of preparation method for the strengthening porcelain that can generate tyrosol and hydroxytyrosol, it is characterised in that: the following steps are included:
(1) it screens strain: by common yeast culture 5-7 days, microorganism being isolated and purified using microbe separation technology, is obtained
Single culture is obtained, the single culture is fermented, the strain in fermentation material containing tyrosol and hydroxytyrosol is filtered out, will sieve
The fungi preservation selected is slant strains；
(2) it cultivates strain: the slant strains being inoculated into fluid nutrient medium, are cultivated to obtain function bacterium seed liquor；
(3) it makes strengthening porcelain: the function bacterium seed liquor being uniformly inoculated into after being fermented on the solid medium of sterilizing,
The raw material fermented is taken out, is dried in the shade up to the strengthening porcelain that can generate tyrosol and hydroxytyrosol.
2. the preparation method of the strengthening porcelain according to claim 1 that tyrosol and hydroxytyrosol can be generated, it is characterised in that:
The ratio uniform for the 1-20% that the function bacterium seed liquor accounts for solid medium weight according to the volume of function bacterium seed liquor is inoculated with
Onto solid medium.
3. the preparation method of the strengthening porcelain according to claim 1 that tyrosol and hydroxytyrosol can be generated, it is characterised in that:
The solid medium the preparation method comprises the following steps: the peptone water solution of 0.7-5.0% is added by the 40% of base-material weight, mixing is equal
Even, loaded in 500 milliliters of triangular flasks, 100 grams every bottle, 121 DEG C of 30 min of sterilizing, the base-material is in rice, wheat and wheat bran
It is one or more.
4. a kind of strengthening porcelain that can generate tyrosol and hydroxytyrosol, it is characterised in that: as described in any one of claims 1 to 3
Preparation method is made.
5. a kind of strengthening porcelain that can generate tyrosol and hydroxytyrosol as claimed in claim 4 is in preparation rich in tyrosol and hydroxyl
Purposes in the white wine of tyrosol.
6. a kind of preparation method of the white wine rich in tyrosol and hydroxytyrosol as claimed in claim 5, it is characterised in that: including
Following steps: the strengthening porcelain that can generate tyrosol and hydroxytyrosol is uniformly mixed with traditional yeast, then according to tradition
Wine-making technology pit entry fermentation, after fermentation distillation take wine, can be obtained the white wine rich in tyrosol and hydroxytyrosol.
7. a kind of preparation method of the white wine rich in tyrosol and hydroxytyrosol as claimed in claim 6, it is characterised in that: described
The weight ratio of strengthening porcelain and traditional yeast that tyrosol and hydroxytyrosol can be generated is 1:100-1:1.
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| CN115353946A (en) * | 2022-08-24 | 2022-11-18 | 梁山瑞升生物工程有限公司 | Directional reinforced Daqu cultivation method |
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