CN109674051A - A kind of method lactobacillus-fermented enrichment wheat polyphenol and prepare antioxidant - Google Patents
A kind of method lactobacillus-fermented enrichment wheat polyphenol and prepare antioxidant Download PDFInfo
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Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Life Sciences & Earth Sciences (AREA)
- Botany (AREA)
- Mycology (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The present invention provides a kind of enrichment method of wheat polyphenol and its applications in terms of preparing antioxidant, belong to food function development field.Using first wheat kernel as raw material, the content of wherein phenolic substances can be improved in lactobacillus-fermented, the wheat polyphenol being prepared by techniques such as extraction, concentration, extraction, concentration, freeze-dryings can effectively clear DPPH free radical activity, have higher iron ion reducing power and oxyradical absorbability.Cell experiment proves that the wheat polyphenol after fermentation is to H2O2The protective effect of the cell oxidative damage of induction is more obvious, such as can reduce ROS is horizontal, improves activities of antioxidant enzymes, improves reduced glutathione content etc..Method of the invention is environmentally protective, low-carbon energy-saving, the deep development wheat resource in China, and product form is stable, be readily transported, antioxidant activity is higher, can be used as functional components and is widely used in food service industry.
Description
One, technical field
The present invention provides a kind of methods lactobacillus-fermented enrichment wheat polyphenol and prepare antioxidant, and it is raw to belong to food
Object utilizes and functional development field.
Two, background technique
Cereal crops of the highland barley as a kind of grass family (Gramineae) Hordeum (Hordeum), have long in China
Cultivation history, due to its feature exposed with seed, also referred to as naked barley or wheat.Since 21st century, Quan Gu
The health benefits of object food are very popular, recommend ingestion standard to improve year after year, various nutrients, including egg are contained in wheat
White, amino acid, polysaccharide, starch, dietary fiber, vitamin and phenolic substances, in addition to as conventional foods such as first ale, zanba, roasted qingke barley flours
Primary raw material, be also widely used for functional food exploitation and medical science research in.
Currently, in all artificial growth cereal crops, the nutrition of wheat and use value are higher, have " high protein,
The distinguishing feature of high microsteping content, low fat, low sugar ", nutritional ingredient are higher than other industrial crops such as rice, wheat, corn.With
Research deeply, related scholar also found that the content of phenolic compounds in wheat is also higher, can effectively it is anti-oxidant, antitumor,
Anti arteriosclerosis, the prevention cardiovascular diseases such as coronary heart disease and apoplexy.Studies have shown that numerous diseases and histoorgan aging all and from
There is certain association by base, the intracorporal free radical of people can both help the energy for transmitting, sustaining life, and can also be used to kill germ
With helminth, participation toxin discharge.On the other hand, excessive free radical can cause Normal cell death and disorganization, cause
Human health injury, further causes chronic disease.Phenols can be removed as a kind of safe and effective free radical scavenger
Excessive free radical in vivo, the cellular damage induced free radical play a protective role.
Lactic acid bacteria is a kind of bacterium that a large amount of lactic acid can be generated using carbohydrate, belongs to Lactobacillaceae
(Lactobacillaceae), Eubacteriales (Eabacteriales), eubacteria guiding principle (Eubacteriac), not according to source
Together, animal sources lactic acid bacteria and Plant Lactobacilli are generally divided into.In comparison, the use of Plant Lactobacilli is more extensive, takes the photograph
Taken amount will not cause rejection to human body when larger, have stronger intestinal colonisation and efficacy exertion ability.Utilize lactic acid
Bacterium fermentation wheat, not only can be improved the content of phenolic substances in wheat, additionally it is possible to the potential function activity of wheat is given full play to,
It is a kind of not only environmental protection but also efficient food development mode, meets " energy-saving and emission-reduction, the low-carbon production " that modern industry is proposed
It is required that.
Patent (publication number CN103445068B, publication date on June 10th, 2015) discloses that " lactobacillus-fermented barley is extracted
The preparation method and its antitumor action of object ", patent (publication number CN108783220A, publication date on November 13rd, 2018) are open
" a kind of fermentation coarse cereals and preparation method thereof ", patent (publication number CN201811081573, publication date September in 2018 17 days) are public
" a kind of method of low temperature ultrasonic extraction line leaf goldspink flower polyphenol " is opened.Above-mentioned patent only ferments to cereal such as barleys,
Or only the extracting method of phenolic substances in raw material is invented, its aldehydes matter content is improved by lactobacillus-fermented wheat
And the method for antioxidant activity is not directed to patent disclosure.This patent uses biotechnological method, has selected have local characteristic
Grain variety --- wheat is studied, invented it is a kind of enrichment wheat polyphenol and prepare high activity antioxidant method.
It is demonstrated experimentally that wheat aldehydes matter content significantly improves through everfermentation, extract has stronger oxidation resistance, Ke Yizuo
For effective antioxidant.
Three, summary of the invention
Present invention solves the technical problem that: a kind of antioxidant is developed in the way of environmentally protective, being can be deep
Degree exploitation distinct Chinese characteristics cereal --- wheat has not only given full play to the potential function activity of wheat, and what is be prepared high-end spreads out
Production product --- antioxidant also can be widely applied in food processing industry, have simple to operation, strong applicability, economic effect
The advantages that benefit is obvious.
In order to solve the above technical problems, the technical method that the present invention takes: a kind of lactobacillus-fermented enrichment wheat polyphenol and
The method for preparing antioxidant, comprising the following steps:
(1) bacteria selection: selection plant lactobacillus Lactobacillus plantarum B1-6, Gen Bank sequence
Number KM200717, is preserved in food science and technology institute Microbiological Lab, Agricultural University Of Nanjing；
(2) pretreatment of raw material: first wheat kernel that selection is dried after cleaning is ground up, sieved, and certain proportion water is added, and high pressure is gone out
Bacterium is made into wheat lotion；
(3) enrichment of wheat polyphenol: member obtained in step (2) is added in activation fermentation strain used by a certain percentage
Wheat lotion seals, is placed in stationary culture in constant incubator, obtains fermentation wheat cream；
(4) prepared by antioxidant: fermentation wheat cream obtained in above-mentioned steps (3) is collected, extract, be concentrated, extracting,
Concentration, drying, are prepared fermentation wheat extract, which has more obvious anti-oxidant than wheat extract before fermenting
Ability can be used as antioxidant；
(5) with the preparation of antioxidant activity food: according to a certain percentage, will be anti-oxidant obtained in above-mentioned steps (4)
Agent is added in liquid or solid food, and the products such as functional beverage, functional bread are made.
In step (2), the first flour size being ground up, sieved is 40-80 mesh；
In step (2), the ratio that water is added is 1: 5-1: 25 (with the volume basis of wheat silty amount and water)；
In step (2), the temperature of the moist heat sterilization is 121 DEG C, time 20min.
In step (3), the first time activation is, Lactobacillus plantarum B1-6 cryopreservation tube is direct
Put into MRS fluid nutrient medium, 37 DEG C of culture 24-60h；
In step (3), second of activation is that 1-10% (in terms of fresh culture volumetric concentration) activation one is added
In fresh MRS fluid nutrient medium, 37 DEG C are continued to cultivate 12-48h secondary culture solution；
In step (3), the addition strain ratio is 1-20% (in terms of wheat lotion volumetric concentration), cultivation temperature 30-
37 DEG C, incubation time 12-72h.
In step (4), the extraction conditions are, with 1: 1-1: 10 ratios (with wheat cream and the ethanol-water solution volume of fermenting
Than meter) be added ethanol solution wheat of ferment obtained in step (3) newborn, 30-50 DEG C of water bath sonicator 1-4h, 5000-
It is centrifuged 1-5min under the conditions of 10000rpm/min, is repeated 3 times, Extraction solvent is the second of 60-100% (in terms of the volumetric concentration of water)
Alcohol-water solution；
In step (4), the first time concentration condition is, using 80-150rpm/min as revolving speed, 30-50 DEG C of water bath condition
Lower rotary evaporation, concentration said extracted liquid；
In step (4), the extraction conditions is that a small amount of saturated salt solution is added in above-mentioned concentrate, by 1: 1-1: 5 ratios
Example (by be concentrated after extracting solution volume multiple in terms of) be added ethyl acetate extracted, concussion shake up, static 5-15min, to complete
Full layering, is repeated 3 times, and collects supernatant liquid；
In step (4), second of concentration condition is, using 80-150rpm/min as revolving speed, 30-50 DEG C of water bath condition
Lower rotary evaporation, the above-mentioned extract liquor of concentration, are added 1-3mL pure water, dissolution residual substance；
In step (4), the drying condition is -20 DEG C of above-mentioned residue solution of pre-cooling 24-72h, -60 DEG C of vacuum refrigerations
Dry 12-60h.
In step (5), the antioxidant additional proportion is 1-20%, preferably 1-10%.
The above method can obtain the antioxidant of fermentation wheat polyphenol preparation.
Preferably, the phenolic substances that above-mentioned fermentation wheat contains is than improving 0.1-0.8 times (with wheat cream before fermentation before fermentation
The multiple meter of total phenol mass concentration in liquid dry matter).
The present invention uses the means of biotechnology, and the macromolecular substances in wheat is made to be converted into tool under microbial action
There is higher active small-molecule substance, the phenolic substances of a part of reference state has become free state.After fermentation, phenols in wheat
Compounds content increases 10-80%, and oxidation resistance of the antioxidant being prepared also than wheat extract before fermenting is significant；
In order to prepare antioxidant, the present invention mainly selects ethanol solution as extractant, extracts in fermentation wheat cream
Active constituent, the wheat antioxidant safety obtained in this way is higher, is easy to store, transport, usage mode is simple, can be extensive
Applied to food processing field；
By establishing different experiments model, for example, DPPH radicals scavenging test, the test of iron ion reducing power, oxidation from
By the test of base absorbability, H2O2Cellular oxidative damage protection test, it was confirmed that antioxidant prepared by the present invention is in 1-
There is apparent oxidation resistance in 10mg/L concentration range.In cell experiment, the present invention has selected cell ROS level, cell
Antioxidase enzyme activity (crucial oxidation preventive content in superoxide dismutase (Superoxide dismutase, SOD), cell
(reduced glutathione (GSH)) and characterization cell membrane integrity important enzyme burst size (lactic dehydrogenase (Lactate
Dehydrogenase, LDH)) as evaluation antioxidant capabilities index, and confirm fermentation wheat extract, i.e., it is anti-oxidant
Agent has apparent effect on a cellular level.
The advantages and positive effects of the present invention:
Wheat is as a kind of crop with Chinese characteristics, and planting range is wide, development and utilization degree is low, traditional cereal processing
Mode is in the form of a single, consumes energy higher, it is difficult to excavate to the potential value of wheat.The present invention has selected generally recognized as safe food
Grade microorganism --- lactic acid bacteria is fermented, not only high-efficiency environment friendly, is also enriched important activity ingredient --- and phenolic substances is filled
The functional activity of wheat has been waved in distribution；Herein on basis, select ethyl alcohol as extractant, safety non-pollution is prepared
Wheat antioxidant have higher anti-oxidant vigor, to radicals scavenging, iron ion reduction, oxyradical absorb and
Oxidative damage protection on cellular level has positive effect.
Four, Detailed description of the invention
The total phenol content of wheat obtained in Fig. 1 embodiment 1.
The free radical scavenging of antioxidant obtained in Fig. 2 embodiment 2.
The iron ion reducing power of antioxidant obtained in Fig. 3 embodiment 2.
The oxyradical absorbability of antioxidant obtained in Fig. 4 embodiment 2.
Influence of the antioxidant obtained in Fig. 5 embodiment 3 to human hepatoma HepG2 cell's ROS level.
Influence of the antioxidant obtained in Fig. 6 embodiment 3 to human hepatoma HepG2 cell's SOD enzyme activity.
Influence of the antioxidant obtained in Fig. 7 embodiment 3 to human hepatoma HepG2 cell's GSH content.
Influence of the antioxidant obtained in Fig. 8 embodiment 3 to human hepatoma HepG2 cell's LDH content.
Five, specific embodiment
Below with reference to the embodiment of the present invention, the technical solution in the present invention is clearly and completely described.Based on this
Embodiment in invention, all other reality obtained by those of ordinary skill in the art without making creative efforts
Example is applied, shall fall within the protection scope of the present invention.Below with reference to specific implementation case, the invention will be further described:
Case study on implementation 1:
Picking full grains clean the first wheat kernel dried, and smash it through 60 meshes, obtain wheat powder, weigh sample matter
Water is added by 1: 10 after amount, 121 DEG C of high pressure sterilization 20min obtain wheat lotion.By Lactobacillus plantarum
B1-6 cryopreservation tube direct plunges into MRS fluid nutrient medium, 37 DEG C of culture 48h, then is added in fresh MRS fluid nutrient medium with 3%
Re-activation is carried out, condition is 37 DEG C and cultivates for 24 hours.The culture of re-activation is added in above-mentioned wheat lotion with 3% inoculum concentration
Liquid, 37 DEG C of stationary culture 36h obtain fermentation wheat cream.It is molten in 1: 10 ratio, 60% alcohol-water of addition in fermentation wheat cream
Liquid mixes well, and 4h is ultrasonically treated under 40 DEG C of water bath conditions, then with 5000rpm/min pelleted by centrifugation 5min, repeats to extract 3 times,
Collect supernatant.Revolving, concentrated extracting solution, are settled to 25mL with 60% ethanol-water solution under the conditions of 40 DEG C, 120rpm/min.
Wheat lotion, wheat extracting solution before being fermented before processing is fermented in aforementioned manners.
Total phenol compounds content is measured using Folin-Ciocalteu method.Draw 200 μ L wheat extracting solutions, thereto plus
Enter 0.5mL forint phenol and 2.3mL pure water, stand 1min, 2mL 7.5% (w/v) sodium carbonate liquor is added, mix at room temperature,
Dark reaction 2h measures absorbance value of the reaction solution under 760nm wavelength later.Replace sample as blank using 60% alcohol-water
Control, indicates total phenol content with gallic acid equivalant (GAE).The result shows that the total phenol content by lactobacillus-fermented, in wheat
It is improved by 4.87 ± 0.10mg GAE/g before fermenting to 5.61 ± 0.02mg GAE/g, is dramatically increased in 0.01 level of P <, increased
Width is 15.81%(Fig. 1)。
Case study on implementation 2:
Picking full grains clean the first wheat kernel dried, and smash it through 60 meshes, obtain wheat powder, weigh sample matter
Water is added by 1: 10 after amount, 121 DEG C of high pressure sterilization 20min obtain wheat lotion.By Lactobacillus plantarum
B1-6 cryopreservation tube direct plunges into MRS fluid nutrient medium, 37 DEG C of culture 48h, then is added in fresh MRS fluid nutrient medium with 3%
Re-activation is carried out, condition is 37 DEG C and cultivates for 24 hours.The culture of re-activation is added in above-mentioned wheat lotion with 3% inoculum concentration
Liquid, 37 DEG C of stationary culture 36h obtain fermentation wheat cream.It is molten in 1: 10 ratio, 60% alcohol-water of addition in fermentation wheat cream
Liquid mixes well, and 4h is ultrasonically treated under 40 DEG C of water bath conditions, then with 5000rpm/min pelleted by centrifugation 5min, repeats to extract 3 times,
Collect supernatant.Revolving, concentrated extracting solution under the conditions of 40 DEG C, 120rpm/min.A small amount of saturated salt solution is added in concentrate,
Ethyl acetate is added in 1: 1 ratio again to be extracted, concussion shakes up, static 10min is repeated to extract 3 times, be received to be layered completely
Collect upper layer of extraction liquid.Using 120rpm/min as revolving speed, 40 DEG C of concentrated extracts.2mL pure water dissolution residual substance is added, -20 DEG C pre-
It is cold for 24 hours after, vacuum freeze drying 48h under the conditions of -60 DEG C obtains fermentation wheat extract, i.e. antioxidant.In aforementioned manners
Wheat lotion, wheat extract before being fermented before processing is fermented.
Using the DPPH free radical scavenging ability of spectrophotometry measurement extract.80% methanol-water of said extracted object
It is made into the solution that concentration is 1-10mg/L, for use.2mL extract solution and 2mL 0.4mmol/L DPPH solution are (with 80% first
Alcohol-water dissolution) it mixes, dark reaction 30min, measures absorbance value of the reaction solution under 517nm wavelength at room temperature.80% methanol-
Water replaces sample as blank control, and DPPH free radical scavenging ability is indicated with clearance rate (%).DPPH is calculated according to the following formula
Free radical scavenging activity: DPPH free radical scavenging activity (%)=(A1-A2)/A1× 100%, in formula, A1For blank control reaction solution
Absorbance value；A2For the absorbance value of example reaction liquid.The result shows that by lactobacillus-fermented, the removing of wheat extract
DPPH free radical ability is remarkably reinforced, and a certain amount effect relationship is presented in antioxidant concentration and oxidation resistance, when extract is dense
When degree is 10mg/L, the DPPH clearance rate of wheat extract is up to 91.44% after fermentation, is 80.45% before fermentation(Fig. 2)。
Using the iron ion reducing power (FRAP) of spectrophotometry measurement extract.Said extracted object is made into dense with pure water
Degree is the solution of 1-10mg/L, for use.By the acetate buffer solution of 100mL 0.3mol/L pH 3.6,10mL 10mmol/L
TPTZ solution (being prepared with 40mmol/L hydrochloric acid) and 10mL 20mmol/L ferric chloride solution mix, and prepare FRAP reagent.Take 1mL
5mL FRAP reagent is added in extract solution thereto, in 37 DEG C of water-bath 10min after mixing, measures reaction solution in 593nm
Absorbance value at wavelength.Replace sample as blank control, iron ion reducing power FeSO using pure water4Equivalent indicates.Knot
Fruit shows that, by lactobacillus-fermented, the iron ion reducing power of wheat extract is remarkably reinforced, antioxidant concentration with it is anti-oxidant
A certain amount effect relationship is presented in ability, and when extract concentrations are 10mg/L, the FRAP of wheat extract is 511.30 μ after fermentation
M FeSO4, it is 398.80 μM of FeSO before fermentation4 (Fig. 3)。
Using the oxyradical absorbability (ORAC) of spectrophotometry measurement extract.Said extracted object is matched with PBS
The solution for being 1mg/L at concentration, for use.200 μ L are added in 96 microwell plates and dilute the extract solution after 1000 times, with 12
Pipettor 2 is serially diluted again, is 10,5,2.5,1.25,0.625 μm of ol/L Trolox controls with concentration.50 μ L are added in every hole
0.2 μm of ol/L fluorescein sodium mixing, 37 DEG C of incubation 15min add 50 μ L 80mmol/L AAPH.96 orifice plates are immediately placed in
Fluorescence intensity, excitation wavelength 485nm, absorbing wavelength 535nm are measured in microplate reader, measuring temperature is 37 DEG C, minute
For 100min.Fluorescence intensity after each micropore reaction calculates area (Area under under fluorescence decline curve with integration method
Curve, AUC), ORAC value is by sample gradient concentration-anti-oxidation protection curve of areas and Trolox standard items gradient concentration-antioxygen
Change protected area slope of a curve ratio to obtain, with the expression of Trolox equivalent, i.e. mol Trolox/g.The result shows that by lactic acid
Bacterium fermentation, the oxyradical absorbability of wheat extract are remarkably reinforced, and antioxidant concentration and oxidation resistance are presented one
Fixed dose-effect relationship, the ORAC value of wheat extract is 11.60 ± 0.43mol Trolox/g after fermentation, before fermentation for 9.66 ±
It is dramatically increased in 0.05 level of 0.39mol Trolox/g, P <, improves 20.08%(Fig. 4)。
Case study on implementation 3:
Wheat extract before fermentation wheat extract, fermentation obtained in case study on implementation 2 is dissolved in sterile water, is made dense
Degree is the solution of 1-10mg/L, crosses 0.22 μm of sterilised membrane filter, for use.The human hepatoma HepG2 cell for taking out -80 DEG C of preservations freezes
Pipe, gently shakes in 37 DEG C of water-baths, and thawing is moved back into centrifuge tube, and 2mL is added and contains 100U/mL penicillin, 0.1mg/mL chain
Dulbecco ' s Modified Eagle ' the s Medium-high glucose of mycin, 1%1mol/L Heps buffer
(DMEM) culture medium is centrifuged 5min under the conditions of 1000rpm/min, gives up supernatant, and 5mL is added later and contains 20% fetal calf serum
Above-mentioned culture medium, piping and druming uniformly, metastatic cells suspension to floor space be 25-75cm2Tissue Culture Flask in, be put into 37 DEG C,
The constant incubator that gas concentration lwevel is 5% is cultivated.When cell is covered with to bottom of bottle area 70-90%, 3mL pancreas is added
Protease digests attached cell 2min, during which takes out culture bottle, the microscopically observation for being 10 times in object lens, until cell base
This becomes free shape from adherent strip or sheet, and 5mL is added, and serum-containing media does not terminate reaction, 3000rpm/min from
After heart 5min, appropriate culture medium is added into cell precipitation, at single cell suspension, concentration is 1 × 10 for piping and druming5-1×106A/mL.
Every hole is inoculated with 100 μ L cell suspensions in 96 orifice plates, and outmost turns are substituted with 100 μ L sterile distilled waters and are added, and is placed in 37 DEG C, dioxy
Change the constant incubator culture that concentration of carbon is 5% for 24 hours, covers with after cell to 96 orifice plate bottoms, be grouped: (1) control group
It is protected to be not only not exposed to wheat extract, but also not by H2O2The cell of stimulation；(2) oxidative damage group is only by H2O2Stimulation
Cell；(3) experimental group is first to be exposed to 1,2,5,10 μ g/mL wheat extract solutions, then by 2400 μm of ol/L H2O2Thorn
Sharp cell.The guard time of wheat extract is 6h, H2O2The oxidative damage time is 2h.
The cell supernatant and cell precipitation of above-mentioned different groups are collected respectively,Human liver cancer is measured using RNA isolation kit respectively
HROS level, superoxide dismutase (SOD) enzyme activity, reduced glutathione (GSH) content, lactic dehydrogenase in epG2 cell Enzyme (LDH) burst size.The result shows that wheat extract has bright in the oxidation resistance of cellular level by lactobacillus-fermented
It is aobvious to improve.After fermentation wheat extract can in significantly more efficient reduction oxidative damage cell ROS it is horizontal(Fig. 5), more efficient
Raising cell SOD enzyme activity(Fig. 6), significantly more efficient increase GSH content(Fig. 7), significantly more efficient protection cell membrane stability
Property, it shows as more effectively reducing LDH burst size(Fig. 8).When extract concentrations are within the scope of 1-10mg/L, the change of different indexs
Change trend has apparent dose-effect relationship(Fig. 5-8)。
Claims (9)
1. a kind of method lactobacillus-fermented enrichment wheat polyphenol and prepare antioxidant, which is characterized in that be with fresh wheat
Raw material selects lactobacillus plantarum to carry out liquid state fermentation, improves the content of phenolic substances in wheat.Herein on basis, system is extracted
Standby wheat polyphenol have higher antioxidant activity, can more efficient removings DPPH free radical, oxyradical, reduction oxygen
It is horizontal to change ROS in damaging cells, improves antioxidase activity etc. in cell.It is characterized in that, following the steps below:
(1) bacteria selection: selection plant lactobacillus Lactobacillus plantarum B1-6, Gen Bank sequence number
KM200717 is preserved in food science and technology institute Microbiological Lab, Agricultural University Of Nanjing；
(2) pretreatment of raw material: first wheat kernel that selection is dried after cleaning is ground up, sieved, and certain proportion water is added, and high pressure sterilization is matched
At wheat lotion；
(3) enrichment of wheat polyphenol: the cream of wheat obtained in step (2) is added in activation fermentation strain used by a certain percentage
Liquid seals, is placed in stationary culture in constant incubator, obtains fermentation wheat cream；
(4) prepared by antioxidant: collecting fermentation wheat cream obtained in above-mentioned steps (3), extracts, is concentrated, extracting, is dense
Contracting, drying, are prepared fermentation wheat extract, which has more obviously anti-oxidant energy than wheat extract before fermenting
Power can be used as antioxidant；
(5) with the preparation of antioxidant activity food: according to a certain percentage, antioxidant obtained in above-mentioned steps (4) being added
Enter in liquid or solid food, the products such as functional beverage, functional bread are made.
2. the method according to claim 1, wherein first wheat kernel is ground up, sieved to obtain in step (2)
Size is first flour of 40-80 mesh, in proportion 1: 5-1: 25 (with the volume basis of wheat silty amount and water) plus water, and 121 DEG C wet
Wheat lotion is obtained after heat sterilization.
3. the method according to claim 1, wherein the actication of culture carries out in two times in step (3),
Lactobacillus plantarum B1-6 cryopreservation tube is once direct plungeed into MRS fluid nutrient medium, 37 DEG C of culture 24-60h,
It takes out, is added in fresh MRS fluid nutrient medium by 1-10% (in terms of fresh culture volumetric concentration) and carries out re-activation, 37 DEG C
Continue to cultivate 12-48h, take out, is added in above-mentioned wheat lotion by inoculum concentration 1-20% (in terms of wheat lotion volumetric concentration),
30-37 DEG C of stationary culture 12-72h, the fermentation wheat cream after obtaining polyphenol enrichment.
4. the fermentation wheat cream that any the method for claim 1-3 obtains.
5. fermentation wheat cream as claimed in claim 4, it is characterised in that total phenol content is than improving 0.1- before fermentation in dry matter
0.8 times (before fermenting in wheat lotion dry matter in terms of the multiple of total phenol mass concentration).
6. the method according to claim 1, wherein joined 1: 1-1 in fermentation wheat cream in step (4):
10 (with wheat cream and the ethanol-water solution volume basis of fermenting) concentration are 60-100% ethanol-water solution (with the volumetric concentration of water
Meter), it is centrifuged 1-5min under the conditions of 30-50 DEG C of water bath sonicator 1-4h, 5000-10000rpm/min, repeats to extract 3 times, collection liquid
Body；Again using 80-150rpm/min as revolving speed, rotary evaporation, concentration said extracted liquid under 30-50 DEG C of water bath condition；Above-mentioned dense
A small amount of saturated salt solution is added in contracting liquid, acetic acid second is added by 1: 1-1: 5 ratios (after being concentrated in terms of the volume multiple of extracting solution)
Ester is extracted, and concussion shakes up, static 5-15min repeats to extract 3 times, collect supernatant liquid to be layered completely；80-
Rotary evaporation, the above-mentioned extract liquor of concentration under the conditions of 150rpm/min, 30-50 DEG C；1-3mL pure water, dissolution residual substance is added；-20
DEG C pre-cooling the above-mentioned residue solution of 24-72h, -60 DEG C of vacuum freeze drying 12-60h.
7. the method according to claim 1, wherein in step (5), by wheat extract of the invention with certain
Ratio is directly added into liquid or solid food, and the products such as beverage, the bread with anti-oxidation function can be made, it is generally preferable to
1-20%, preferably 1-10%.
8. the fermentation wheat extract that any the method for claim 1-7 obtains, i.e., antioxidant of the invention.
9. antioxidant according to any one of claims 8, which is characterized in that in studying and test in vitro, if antioxidant concentration exists
Between 1-10mg/L (in terms of the mass concentration of the be dissolved in solvent of extract), a variety of oxidation resistances can be provided simultaneously with, including
More significant DPPH free radical scavenging ability, iron ion reducing power, oxyradical absorbability, and for cell oxygen
Change and damage significantly more efficient protective effect, cell ROS is horizontal, improve cell SOD enzyme activity, increases GSH in cell contains as reduced
Amount reduces cell LDH burst size.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110432423A (en) * | 2019-07-24 | 2019-11-12 | 芜湖职业技术学院 | A kind of antioxidant of microbial food and preparation method thereof |
| CN111357928A (en) * | 2020-02-24 | 2020-07-03 | 江苏省农业科学院 | Method for processing dough sheet by using kefir modifier and application of method |
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2018
- 2018-12-07 CN CN201811500270.3A patent/CN109674051A/en active Pending
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| CN110432423A (en) * | 2019-07-24 | 2019-11-12 | 芜湖职业技术学院 | A kind of antioxidant of microbial food and preparation method thereof |
| CN111357928A (en) * | 2020-02-24 | 2020-07-03 | 江苏省农业科学院 | Method for processing dough sheet by using kefir modifier and application of method |
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