CN107656081B - 70 to 80 degree wine alcoholic strength simplicity rapid assay methods - Google Patents
70 to 80 degree wine alcoholic strength simplicity rapid assay methods Download PDFInfo
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- CN107656081B CN107656081B CN201710925076.9A CN201710925076A CN107656081B CN 107656081 B CN107656081 B CN 107656081B CN 201710925076 A CN201710925076 A CN 201710925076A CN 107656081 B CN107656081 B CN 107656081B
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- 230000001476 alcoholic effect Effects 0.000 title claims abstract description 78
- 235000014101 wine Nutrition 0.000 title claims abstract description 66
- 238000003556 assay Methods 0.000 title claims description 9
- 238000000034 method Methods 0.000 claims abstract description 52
- 238000002835 absorbance Methods 0.000 claims abstract description 29
- 239000012634 fragment Substances 0.000 claims abstract description 19
- 108020004414 DNA Proteins 0.000 claims description 39
- ZHNUHDYFZUAESO-UHFFFAOYSA-N Formamide Chemical compound NC=O ZHNUHDYFZUAESO-UHFFFAOYSA-N 0.000 claims description 18
- 230000036425 denaturation Effects 0.000 claims description 15
- 238000004925 denaturation Methods 0.000 claims description 15
- 239000003795 chemical substances by application Substances 0.000 claims description 12
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 9
- 239000004202 carbamide Substances 0.000 claims description 9
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 238000002965 ELISA Methods 0.000 claims description 7
- 239000012153 distilled water Substances 0.000 claims description 6
- 238000002156 mixing Methods 0.000 claims description 6
- 239000007864 aqueous solution Substances 0.000 claims description 4
- 102000053602 DNA Human genes 0.000 claims description 2
- 230000000295 complement effect Effects 0.000 claims description 2
- 102000004190 Enzymes Human genes 0.000 claims 1
- 108090000790 Enzymes Proteins 0.000 claims 1
- 238000005259 measurement Methods 0.000 abstract description 29
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 abstract description 25
- 238000006062 fragmentation reaction Methods 0.000 abstract description 6
- 238000013467 fragmentation Methods 0.000 abstract description 4
- 238000011088 calibration curve Methods 0.000 abstract 1
- 235000019441 ethanol Nutrition 0.000 description 12
- 235000020097 white wine Nutrition 0.000 description 10
- 240000006394 Sorghum bicolor Species 0.000 description 7
- 235000011684 Sorghum saccharatum Nutrition 0.000 description 7
- 241001247821 Ziziphus Species 0.000 description 7
- 229960004756 ethanol Drugs 0.000 description 7
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 238000001514 detection method Methods 0.000 description 6
- 239000002023 wood Substances 0.000 description 5
- 239000007788 liquid Substances 0.000 description 3
- 238000002844 melting Methods 0.000 description 3
- 230000008018 melting Effects 0.000 description 3
- 238000004497 NIR spectroscopy Methods 0.000 description 2
- 239000003398 denaturant Substances 0.000 description 2
- 238000000186 gas chromatography-infrared spectroscopy Methods 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- WPPDXAHGCGPUPK-UHFFFAOYSA-N red 2 Chemical compound C1=CC=CC=C1C(C1=CC=CC=C11)=C(C=2C=3C4=CC=C5C6=CC=C7C8=C(C=9C=CC=CC=9)C9=CC=CC=C9C(C=9C=CC=CC=9)=C8C8=CC=C(C6=C87)C(C=35)=CC=2)C4=C1C1=CC=CC=C1 WPPDXAHGCGPUPK-UHFFFAOYSA-N 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000009825 accumulation Methods 0.000 description 1
- 235000013334 alcoholic beverage Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 229960000935 dehydrated alcohol Drugs 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 238000012203 high throughput assay Methods 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229940046166 oligodeoxynucleotide Drugs 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000002798 spectrophotometry method Methods 0.000 description 1
Classifications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/98—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving alcohol, e.g. ethanol in breath
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
- G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
- G01N21/31—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry
- G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
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- Engineering & Computer Science (AREA)
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- Pathology (AREA)
- Hematology (AREA)
- General Physics & Mathematics (AREA)
- Biochemistry (AREA)
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- Spectroscopy & Molecular Physics (AREA)
- Biomedical Technology (AREA)
- Urology & Nephrology (AREA)
- Molecular Biology (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Biotechnology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Distillation Of Fermentation Liquor, Processing Of Alcohols, Vinegar And Beer (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
Alcoholic strength is that alcohol product must survey index, lacks have both accurate and reliable, easy quick and economical and practical alcoholic strength measuring method at present.In containing ethanol system unwinding, which can occur, for the present invention using DNA fragmentation causes its absorbance under 260nm wavelength to change, to establish a kind of new alcoholic strength measuring method.The present invention screens suitable short double chain DNA fragment mixed system as shown in SEQ ID NO:1-2 for 70 to 80 degree wine, and the calibration curve equation for establishing its absorbance x and wine sample alcoholic strength y is y=8.6163x+68.592(70≤y≤80).Alcoholic strength measuring method of the invention is suitable for the measurement of 70 to 80 degree wine, has good accuracy and reliability, safer without distilling, it is easy to operate rapid, required wine sample amount is seldom and cheap, is suitable for brewery 70 to 80 and spends the measurement practice of wine alcoholic strength.
Description
Technical field
The present invention relates to ethyl alcohol detection methods, in particular to ethanol content, that is, alcoholic strength simplicity of 70 to 80 degree wine of one kind
Rapid assay methods.
Background technique
Ethyl alcohol is the main component of wine, and content is one of the important indicator of wine quality, is referred to as alcoholic strength in national standard, is led to
Often it refers at 20 DEG C, the ethyl alcohol ml contained in every 100 ml wine liquid.Alcoholic strength is that must surveying for alcohol product refers to
Mark, measuring method has much at present, and common includes alcoholic strength meter method, density bottle method, gas chromatography and near infrared spectroscopy
Deng.Both are the methods based on modern analytical technique for gas chromatography and near infrared spectroscopy, have that detection sensitivity is high, knot
Fruit is accurate, can be with high throughput assay the advantages that, but there is also equipment it is expensive, complicated for operation and testing cost is high the deficiencies of, because
It is practical that this is not suitable for brewery's detection.The most commonly used is alcoholic strength meter methods and density bottle method in brewery, however both methods detects
Before be both needed to distill alcoholic drink in advance, after removing impurity, then alcohol water blend is measured.Time-consuming for entire detection process,
Step is more, stability is poor, is not able to satisfy the demand of quickly detection development.
In summary, it needs to establish accurate and reliable, easy quick and economical and practical alcoholic strength measuring method at present, for
The present invention provides be based on specific ultrashort DNA fragmentation (i.e. oligodeoxynucleotide chain) in different alcoholic strength wine samples for 70 to 80 degree wine
Middle denaturation degrees difference leads under 260nm wavelength absorbance difference, and establishes a kind of new alcoholic strength measuring method i.e.
Meet this demand.DNA double chain is that the hydrogen bond and accumulation force between the base maintain, however some external factors such as high temperature,
In the presence of extreme pH, organic reagent such as urea, formamide, methanol and ethyl alcohol, these maintenance energies can be destroyed, so that
DNA double chain unwinding becomes single-stranded, causes its absorbance at 260nm that (also known as hyperchromicity) is significantly increased.Utilize this spy
Property, suitable DNA system is constructed, wine sample is added thereto processing, using the DNA system absorbance between alcoholic strength in wine sample
Association, to realize the measurement of alcoholic strength.
Summary of the invention
The purpose of the present invention is to provide the easy rapid assay methods of 70 to 80 degree wine alcoholic strengths.
The technical solution adopted by the present invention to solve the technical problems is the easy quickly measurement side of 70 to 80 degree wine alcoholic strengths
The key step of method is as follows:
(1) pipette 400ul room temperature 70 to 80 degree wine wine sample to be measured into empty ELISA Plate, be added the agent of 50ul denaturation and
The short double chain DNA fragment mixed system of 50ul simultaneously stands 5min after mixing well, while it is that 400ul wine sample to be measured adds that control wells, which are arranged,
50ul denaturation agent adds 50ul distilled water, and wine sample to be measured and control make even row survey three times, by ELISA Plate after sample process is good
It is placed in microplate reader.
(2) it sets microplate reader program and measures absorbance in sample well and control wells under 260nm wavelength, then sample
Mean light absorbency subtract control mean light absorbency be short double chain DNA fragment mixed system in sample practical absorbance.It will
In sample DNA system practical absorbance x substitute into relational expression y=8.6163x+68.592(70≤y≤80) in converse wine sample
Alcoholic strength y.
Wherein, 70 to 80 degree wine as described in step (1) refers to that alcoholic strength is more than or equal to 1 and is less than or equal to 20 liquor
Product；The wine sample temperature is normal room temperature, it is preferable that control is between 30.0 ± 5.0 DEG C；The denaturation agent
It is the urea of 18% mass volume fraction and the formamide mixed aqueous solution of 20% volume volume fraction, i.e., every 100ml mixing is water-soluble
Contain 18g urea and 20ml formamide in liquid；The short double chain DNA fragment mixed system includes 2 short double chain DNA fragments
1 and 2, for sequence respectively as shown in SEQ ID NO:1-2, each short double chain DNA fragment concentration of item is 10 μm of ol/l.
The present invention screens suitable short double chain DNA fragment mixed system, and discloses its extinction in 70 to 80 degree wine
Relationship between degree and alcoholic strength constructs the easy rapid assay methods of 70 to 80 degree wine alcoholic strengths on this basis.This hair
Bright alcoholic strength measuring method is suitable for the measurement of 70 to 80 degree wine, has good accuracy and reliability, without distilling ratio
Safer, easy to operate required wine sample amount is seldom and cheap rapidly, is suitable for brewery 70 to 80 and spends the measurement of wine alcoholic strength
Practice.
Detailed description of the invention
Fig. 1 is absorbance-alcoholic strength standard curve that the present invention 70 to 80 spends wine alcoholic strength measurement new method.Abscissa is
Absorbance of the artificial wine sample for adding short double chain DNA fragment mixed system and denaturation agent to handle at 260nm, ordinate is it
Alcoholic strength.Y=8.6163x+68.592 in figure is the relational expression of the absorbance x and alcoholic strength y of 23.0 software of SPSS fitting, R2
=0.9988 indicate be matched curve the coefficient of determination.
Specific embodiment
Below in conjunction with specific embodiment, the present invention is further explained.
The present invention is additionally added urea and the formamide mixing of optimized certain concentration in wine sample to be measured and DNA system
Liquid, primarily to being in DNA fragmentation in the environment that one is easy to be denaturalized, so that alcohol determining method is with higher
Sensitivity.Short double-stranded DNA system provided by the invention contains 2 DNA fragmentations, by SEQ ID NO:1-2 be incremented by sequence its
Melting temperature is to rise, therefore increasing with alcoholic strength, the type and quantity of the interior denaturation double-strand of system are consequently increased, because
It is positively related between this absorbance and alcoholic strength.In addition, the denaturation of DNA is to realize in a narrow range, therefore need
Screening and condition optimizing are carried out, so that boundary is that have under the specified conditions for the DNA fragmentations of different melting temperatures in DNA system
It is superimposed continuity.In ethyl alcohol, the narrow mobility scale (within 10ml) and the ultrashort segment of DNA of this mild denaturant are (insufficient
30bp), it has been investigated that the melting extent of reaction of Denaturant for ethanol concentration and specific DNA system may be implemented in a certain range
Linear fit.Since second alcohol and water accounts for main body in wine, solid content is very low therefore its absorbance on system in 260nm influences
Very little.Only add the wine sample of denaturation agent as control in addition, being provided with, influence of the background to absorbance can be eliminated as much as.
System of the present invention and under the conditions of, 70 degree of alcoholic strengths below are still insufficient to allow any short double chain DNA fragment that partial denaturization occurs,
And 80 degree or more of alcoholic strength has been enough to be denaturalized all short double chain DNA fragments completely, therefore alcoholic strength provided by the invention
Standard curve is only applicable to the alcoholic strength measurement of 70 to 80 degree wine.In the detection of drinks product quality, spectrophotometry is common
Means, can measure including coloration, methanol and specific amino acids etc., therefore spectrophotometer is conventional equipment.
Embodiment 1
Embodiment 1 is that the present invention 70 to 80 spends alcoholic strength measurement standard curve in wine alcoholic strength simplicity rapid assay methods
Production.
(1) artificial 70 to 80 degree wine sample is prepared
70.0,72.0,74.0,76.0,78.0 and 80.0ml dehydrated alcohol is taken to hold in 100ml respectively with corresponding pipette
In measuring bottle, it is continuously added distilled water and slightly shakes and stand repeatedly, final constant volume to 100ml is stored at room temperature 10min to get arriving
Artificial 70 to the 80 degree wine sample of different alcoholic strengths.
(2) artificial 70 to 80 degree wine sample pre-treatment
Pipette 400ul 70 to the 80 artificial wine sample (30.0 DEG C) of degree to be measured into empty common ELISA Plate with liquid-transfering gun, first plus
Enter the urea of 18% mass volume fraction and the formamide mixed aqueous solution 50ul of 20% volume volume fraction as denaturation agent,
Add the short double chain DNA fragment mixed system as shown in SEQ ID NO:1-2 (concentration of every bar segment is 10 μm of ol/l)
50ul, then total system is 500ul, stands 5min after mixing well.It should be noted that the urea of 18% mass volume fraction and
The formamide of 20% volume volume fraction refers to the formamide of urea and 20ml in 100ml distilled water dissolved with 18g, short double-strand
DNA fragmentation is that the complementary single strand mixed in equal amounts synthesized by Shanghai Sheng Gong bioengineering Co., Ltd forms.
It is that artificial 70 to the 80 degree wine sample of 400ul adds 50ul denaturation agent to add 50ul distilled water that control wells are arranged simultaneously, i.e.,
To replace short double chain DNA fragment mixed system with distilled water, each artificial 70 to 80 degree wine sample and corresponding control are made even for control
ELISA Plate three times, is placed in microplate reader (the Multiskan GO of Thermo company of the U.S.) by row after sample preparation is good.
(3) it is fitted alcoholic strength measuring method standard curve
It sets microplate reader program and measures absorbance in sample well and control wells under 260nm wavelength, then sample is flat
It is DNA system in sample that equal absorbance (three are averaged in parallel), which subtracts the mean light absorbency (three are averaged in parallel) of control,
Practical absorbance, as shown in table 1.
The scatterplot of absorbance and corresponding alcoholic strength is observed, finds 70.0,72.0,74.0,76.0,78.0 and
Linear trend is presented in the corresponding point of 80.0 this 6 alcoholic strengths, is fitted to obtain the relational expression between alcoholic strength y and absorbance x
For y=8.6163x+68.592(70≤y≤80), and R2=0.9988 very close 1 shows that linear relationship is significant, then this i.e. alcohol
Spend measuring method standard curve.
1 alcoholic strength standard curve data of table
| Mean light absorbency x | Alcoholic strength y |
| 0.177 | 70.0 |
| 0.381 | 72.0 |
| 0.611 | 74.0 |
| 0.877 | 76.0 |
| 1.102 | 78.0 |
| 1.314 | 80.0 |
Embodiment 2
Embodiment 2 is the practical measurement using new method of the present invention to certain 70 to 80 degree wine alcoholic strength.
(1) wine sample pre-treatment to be measured
Commercially available Mount Taishan original degree white wine (production of Mount Taishan the wine industry Group Plc) is selected, two bottles of (former degree white wine 1 and originals are taken
Spend white wine 2) carry out alcoholic strength measurement.Pipette the 400ul wine sample (30.0 DEG C) into empty common ELISA Plate with liquid-transfering gun, first plus
Enter the urea of 18% mass volume fraction and the formamide mixed aqueous solution 50ul of 20% volume volume fraction as denaturation agent,
Add the short double chain DNA fragment mixed system as shown in SEQ ID NO:1-2 (concentration of every bar segment is 10 μm of ol/l)
50ul, then total system is 500ul, stands 5min after mixing well.
It is that 400ul wine sample to be measured adds 50ul denaturation agent that 50ul is added to distill aqueous systems (total volume that control wells are arranged simultaneously
Also it is 500ul), each wine sample to be measured and corresponding control make even row three times, and ELISA Plate is placed in microplate reader after sample preparation is good
In (the Multiskan GO of Thermo company of the U.S.).
(2) wine sample alcoholic strength measurement to be measured
It sets microplate reader program and measures absorbance in sample well and control wells under 260nm wavelength, then sample is flat
It is DNA system in sample that equal absorbance (three are averaged in parallel), which subtracts the mean light absorbency (three are averaged in parallel) of control,
Practical absorbance, as shown in table 2.The relational expression substituted between alcoholic strength y and absorbance x is y=8.6163x+68.592(70
≤ y≤80), it calculates alcoholic strength result (table 2).In addition, the alcoholic strength of the wine sample is measured using density bottle method simultaneously, as a result such as table
Shown in 2.
The alcoholic strength numerical value of 2 the method for the present invention of table and the measurement of density bottle method
| White wine | Absorbance values | The alcoholic strength that the method for the present invention is surveyed | The alcoholic strength that density bottle method is surveyed | Deviation % |
| Former degree white wine 1 | 0.336 | 71.49 | 71.23 | 0.37 |
| Former degree white wine 1 | 0.331 | 71.44 | 71.18 | 0.37 |
In table 2, deviation refers on the basis of the alcoholic strength that density bottle method measures, alcoholic strength that the method for the present invention measures and its
Difference accounts for the percentage of alcoholic strength meter method result.As shown in Table 2, the two bottles of Mount Taishan original degree white wine wine measured using the method for the present invention
Precision and density bottle method measurement result deviation are all 0.37%, and the accurate of the alcoholic strength measurement of national regulations is met within 1%
Degree.
Embodiment 3
Embodiment 3 is using new method of the present invention to red jujube magma white wine (the Shijiazhuang City Fuxi platform the wine industry of commercially available jujube wood
Co., Ltd produces) measurement of the alcoholic strength of this 70 to 80 degree wine, two bottles of (jujube wood red 1 and red 2) of jujube wood are taken altogether.Referring to implementation
Example 2 operates, and the results are shown in Table 3.In addition, carrying out the measurement of this wine alcoholic strength using density bottle method simultaneously.
The alcoholic strength numerical value of 3 the method for the present invention of table and the measurement of density bottle method
| White wine | Absorbance values | The alcoholic strength that the method for the present invention is surveyed | The alcoholic strength that density bottle method is surveyed | Deviation % |
| Jujube wood red 1 | 0.778 | 75.30 | 74.93 | 0.49 |
| Jujube wood red 2 | 0.770 | 75.23 | 75.06 | 0.23 |
As shown in Table 3, using the red white wine alcoholic strength of two bottles of jujubes wood and density bottle method measurement result of the method for the present invention measurement
Deviation is respectively 0.49% and 0.23%, and the accuracy of the alcoholic strength measurement of national regulations is met within 1%.
Embodiment 4
Embodiment 4 is that commercially available imperial ease liquor made from sorghum, (it is limited that the wine industry is contained by five town Ke Shu China of Yushu City using new method of the present invention
Company produce) this 70 to 80 degree wine alcoholic strength measurement, take two bottles (liquor made from sorghum 1 and liquor made from sorghums 2) altogether.Referring to embodiment 2
Operation, the results are shown in Table 4.In addition, carrying out the measurement of this wine alcoholic strength using density bottle method simultaneously.
The alcoholic strength numerical value of 4 the method for the present invention of table and the measurement of density bottle method
| Liquor made from sorghum | Absorbance values | The alcoholic strength that the method for the present invention is surveyed | The alcoholic strength that density bottle method is surveyed | Deviation % |
| Liquor made from sorghum 1 | 1.229 | 79.18 | 79.83 | 0.81 |
| Liquor made from sorghum 2 | 1.243 | 79.30 | 79.91 | 0.76 |
As shown in Table 4, using the two bottles of imperial ease liquor made from sorghum alcoholic strengths and density bottle method measurement result of the method for the present invention measurement
Deviation is respectively 0.81% and 0.76%, and the accuracy of the alcoholic strength measurement of national regulations is met within 1%.
In summary, alcoholic strength measuring method of the invention is suitable for the measurement of 70 to 80 degree wine, and it is good accurate to have
Property and reliability；It is safer without distilling compared with alcoholic strength meter method or density bottle method, operate 10min and simple, convenient and rapid；Separately
Wine sample amount needed for outer is seldom, and testing cost is very cheap less than 1 yuan/sample, is suitable for the such wine alcoholic strength measurement practice of brewery.
Sequence table
<110>yellow race mountain
<120>70 to 80 degree wine alcoholic strength simplicity rapid assay methods
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
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<213>artificial sequence (Artificial Sequence)
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atatatatat actgactgac tgact 25
Claims (1)
1.70 to 80 degree wine alcoholic strength simplicity rapid assay methods, it is characterised in that alcoholic strength simplicity rapid assay methods it is main
Step are as follows:
(1) 400ul room temperature 70 to 80 degree wine wine sample to be measured is pipetted into empty ELISA Plate, and 50ul denaturation is added in sample well
Agent and the short double chain DNA fragment mixed system of 50ul simultaneously stand 5min after mixing well, while it is 400ul wine to be measured that control wells, which are arranged,
Sample adds 50ul denaturation agent to add 50ul distilled water, and sample well and control wells make even row survey three times, by enzyme after sample process is good
Target is placed in microplate reader；The denaturation agent being wherein added is the urea and 20% volume integral of 18% mass volume fraction
Several formamide mixed aqueous solutions；The short double chain DNA fragment mixed system being wherein added includes 2 specific short double-stranded DNAs
Segment, short double chain DNA fragment are formed by complementary single strand mixed in equal amounts, the single-stranded sequence of two specific short double chain DNA fragments
For column respectively as shown in SEQ ID NO:1 and SEQ ID NO:2, each short double chain DNA fragment concentration of item is 10 μm of ol/l；
(2) it sets microplate reader program and measures absorbance in sample well and control wells under 260nm wavelength, then sample well is flat
The mean light absorbency that equal absorbance subtracts control wells is the practical absorbance of short double chain DNA fragment mixed system in sample；It will
This practical absorbance x substitutes into the alcoholic strength model of y, 70≤y≤80 that wine sample to be measured is conversed in relational expression y=8.6163x+68.592
It encloses interior effective.
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2017
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| Title |
|---|
| A Study of DNA Melting in Concentrated Water-Alcohol Solutions;I. A. Mikhailenko & L. S. Shlyakhtenko;《Journal of Biomolecular Structure and Dynamics》;20120521;第1卷(第6期);1501-1510 |
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| CN107656081A (en) | 2018-02-02 |
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