CN102147397A - Method for detecting taurine in functional beer by adopting high performance liquid chromatography (HPLC) - Google Patents
Method for detecting taurine in functional beer by adopting high performance liquid chromatography (HPLC) Download PDFInfo
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- CN102147397A CN102147397A CN 201010301398 CN201010301398A CN102147397A CN 102147397 A CN102147397 A CN 102147397A CN 201010301398 CN201010301398 CN 201010301398 CN 201010301398 A CN201010301398 A CN 201010301398A CN 102147397 A CN102147397 A CN 102147397A
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- taurine
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- XOAAWQZATWQOTB-UHFFFAOYSA-N taurine Chemical compound NCCS(O)(=O)=O XOAAWQZATWQOTB-UHFFFAOYSA-N 0.000 title claims abstract description 131
- 229960003080 taurine Drugs 0.000 title claims abstract description 64
- 235000013405 beer Nutrition 0.000 title claims abstract description 50
- 238000000034 method Methods 0.000 title claims abstract description 33
- 238000004128 high performance liquid chromatography Methods 0.000 title claims abstract description 19
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims abstract description 18
- 238000004458 analytical method Methods 0.000 claims abstract description 16
- 150000001413 amino acids Chemical class 0.000 claims abstract description 15
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims abstract description 10
- 239000003643 water by type Substances 0.000 claims abstract description 9
- 238000009795 derivation Methods 0.000 claims abstract description 7
- 230000002452 interceptive effect Effects 0.000 claims abstract description 7
- BDKLKNJTMLIAFE-UHFFFAOYSA-N 2-(3-fluorophenyl)-1,3-oxazole-4-carbaldehyde Chemical compound FC1=CC=CC(C=2OC=C(C=O)N=2)=C1 BDKLKNJTMLIAFE-UHFFFAOYSA-N 0.000 claims abstract description 6
- 235000017281 sodium acetate Nutrition 0.000 claims abstract description 6
- 229940087562 sodium acetate trihydrate Drugs 0.000 claims abstract description 6
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims abstract description 5
- 230000014759 maintenance of location Effects 0.000 claims abstract description 5
- 239000000126 substance Substances 0.000 claims abstract description 5
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 111
- 239000012071 phase Substances 0.000 claims description 36
- 239000000872 buffer Substances 0.000 claims description 32
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 16
- 238000001514 detection method Methods 0.000 claims description 7
- 239000000463 material Substances 0.000 claims description 6
- 239000012074 organic phase Substances 0.000 claims description 6
- 230000009466 transformation Effects 0.000 claims description 6
- 238000011084 recovery Methods 0.000 claims description 5
- 238000013016 damping Methods 0.000 claims description 4
- 238000010790 dilution Methods 0.000 claims description 4
- 239000012895 dilution Substances 0.000 claims description 4
- 230000009977 dual effect Effects 0.000 claims description 4
- 239000012530 fluid Substances 0.000 claims description 4
- RJSRSRITMWVIQT-UHFFFAOYSA-N quinolin-6-amine Chemical compound N1=CC=CC2=CC(N)=CC=C21 RJSRSRITMWVIQT-UHFFFAOYSA-N 0.000 claims description 4
- 238000012360 testing method Methods 0.000 claims description 4
- 238000011156 evaluation Methods 0.000 claims description 3
- 238000010829 isocratic elution Methods 0.000 claims description 3
- 238000011002 quantification Methods 0.000 claims description 3
- 230000001105 regulatory effect Effects 0.000 abstract description 5
- 238000010828 elution Methods 0.000 abstract description 4
- 239000007853 buffer solution Substances 0.000 abstract description 2
- 230000035945 sensitivity Effects 0.000 abstract description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-M Formate Chemical compound [O-]C=O BDAGIHXWWSANSR-UHFFFAOYSA-M 0.000 abstract 1
- 235000001014 amino acid Nutrition 0.000 description 12
- 239000003795 chemical substances by application Substances 0.000 description 5
- 238000002360 preparation method Methods 0.000 description 5
- 239000003153 chemical reaction reagent Substances 0.000 description 4
- 238000010438 heat treatment Methods 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 239000000843 powder Substances 0.000 description 4
- ZWLUXSQADUDCSB-UHFFFAOYSA-N phthalaldehyde Chemical compound O=CC1=CC=CC=C1C=O ZWLUXSQADUDCSB-UHFFFAOYSA-N 0.000 description 3
- 239000011347 resin Substances 0.000 description 3
- 229920005989 resin Polymers 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 description 2
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N Sodium azide Chemical compound [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000001212 derivatisation Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000003480 eluent Substances 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000005374 membrane filtration Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 239000012086 standard solution Substances 0.000 description 2
- 239000012749 thinning agent Substances 0.000 description 2
- 229910021642 ultra pure water Inorganic materials 0.000 description 2
- 239000012498 ultrapure water Substances 0.000 description 2
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical group [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 238000002479 acid--base titration Methods 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 210000000941 bile Anatomy 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 239000000337 buffer salt Substances 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- 150000001768 cations Chemical class 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000007872 degassing Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000003797 essential amino acid Substances 0.000 description 1
- 235000020776 essential amino acid Nutrition 0.000 description 1
- 230000005284 excitation Effects 0.000 description 1
- 238000001506 fluorescence spectroscopy Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 229940054441 o-phthalaldehyde Drugs 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 238000001303 quality assessment method Methods 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000011496 sports drink Nutrition 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 229940086542 triethylamine Drugs 0.000 description 1
- 238000000825 ultraviolet detection Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a method for detecting taurine in functional beer by adopting a high performance liquid chromatography (HPLC), comprising the steps: (1) a derivation pretreatment procedure: carrying out pre-column derivation on 6-aminoquinoline-N-hydroxysuccinimido formate; (2) a gradient elution procedure: regulating the pH value of a mobile phase A to 4.98 by using 50% phosphoric acid, and optimizing the gradient elution procedure aiming at a standard sample and a beer sample respectively so as to completely separate the taurine from other interfering substances in the beer, wherein the mobile phase A is a buffer solution including 139.92mmol/L of sodium acetate trihydrate and 16.39mmol/L of triethylamine, a chromatographic column used for sample analysis is an AccQ-Tag amino acid analysis column C18 of the American Waters company, and the chromatographic column is in the specification of 3.9mm*50mm and 4mum; and (3) a data treatment procedure: comparing beer to be detected with a standard taurine sample spectrogram so as to realize nature determination in a retention time and quantity determination in a chromatograph peak area. By means of the method disclosed by the invention, the taurine in the beer is completely separated from other amino acids and can be accurately determined on the nature and quantity. The method has the advantages of simplicity for operation, accuracy, reliability and higher sensitivity.
Description
Technical field
The invention belongs to beer quality assessment technique field, in more detail, relate to the high performance liquid chromatography that is used for measuring ability beer taurine.
Background technology
Is taurine (Taurine) a kind of sulfur-bearing that extensively is present in the humans and animals body? amino acid was just found from the bile of ox and was separated as far back as 1827, and therefore gained the name.Taurine has biologic activity widely as a kind of conditionality essential amino acid in the human body, is the important substance of regulating the body normal physiological function.Studies show that, taurine because of its to the Radical Metabolism in the motion body, Ca
2+Transhipment etc. has good regulating action, thereby shows good anti-sports fatigue effect.At present, release the drink with function of multiple interpolation taurine on the market, as contained sports beer, sports drink of taurine etc.
For these functional beers are carried out strict quality, need set up accurately taurine assay method in the beer.Content of taurine method commonly used has acid base titration, amino acid autoanalyzer method, thin-layered chromatography and high performance liquid chromatography etc. in the detection by quantitative food at present; and the fluorophotometric method that mostly is that detects at beer specially; other method is not applied to beer and detects; because the otherness of the whole bag of tricks and sample is all different at aspects such as sample pre-treatments, analytical procedure and employed reagent.
The fluorophotometric method adopts strong acidic ion resin to isolate taurine in the beer, and carries out fluorescence spectrometry behind o-phthalaldehyde(OPA) (OPA)/2 mercaptoethanol derivatization.Because taurine is sulfonated aminoacid (NH
2-CH
2-CH
2-SO
3H), contain sulfonate radical (SO
3H), least tight with resin-bonded, when at first being eluted through the cationic resin column time, may exist in the sample with more weak other amino acid of resin-bonded and flow out altogether, thereby cause measured value higher.In addition, it is comparatively loaded down with trivial details that resin cation is handled operation, is unfavorable for conventional sense.And owing to there are tens kinds of different types of free amino acids in the beer, easily the detection of taurine produced and disturb, only depend on single fluorophotometric method to be difficult to taurine is separated from other amino acid fully, it is accurately quantitative thereby influence.
Summary of the invention
The objective of the invention is to set up and a kind ofly can carry out accurately qualitative and quantitative high performance liquid chromatography, be used for the type quality of beer and control taurine in the functional beer.The present invention adopts high performance liquid chromatography (English abbreviation: HPLC); utilize 6-aminoquinoline base-N-hydroxyl succinimide carbamate (english abbreviation AQC) column front derivation; diode array or UV-detector and fluorescence detector series connection detect; by regulating buffer concentration, pH and solvent gradient elution; realize in the beer that taurine and other are amino acid whose and separate fully, thereby it is carried out accurately qualitative and quantitative.
A kind of method that adopts taurine in the high performance liquid chromatography measuring ability beer of the present invention comprises the steps: (one) pre-treatment program of deriving: 6-aminoquinoline base-N-hydroxyl succinimide carbamate (AQC) column front derivation;
The present invention also comprises the steps:
(2) gradient elution program: mobile phase A is the damping fluid of 139.92mmol/L sodium acetate trihydrate+16.39mmol/L triethylamine, regulate pH to 4.98 with 50% phosphoric acid, Mobile phase B is a pure water, moving phase C is an acetonitrile, optimize the gradient elution program at standard specimen and beer sample respectively, make that other interfering material separates fully in taurine and the beer;
(3) stratographic analysis and data processor:
The employed chromatographic column of sample analysis is the AccQ-Tag amino acid analysis C of U.S. Waters company
18Post, 3.9mm*50mm, 4um, beer to be measured and the contrast of taurine standard specimen spectrogram is qualitative with retention time, the chromatographic peak peak area quantification, computing formula is as follows:
In the formula: C
SampleThe content of taurine in-----beer to be measured, mg/L;
C
Mark----content of taurine in the-standard specimen, mg/L;
A
SampleThe peak area of taurine in-----beer to be measured;
A
Mark----peak area of taurine in the-standard specimen;
The dilution gfactor of f-----beer to be measured.
The concrete elution program of step (two) is: in (1) 0 ~ 0.5 minute, moving phase is 100% buffer A, carries out isocratic elution; In the time of (2) 0.5 minutes, moving phase moment switches to 99% buffer A+1% acetonitrile; (3) 0.5 ~ 27.0 minutes, moving phase was 95% buffer A+5% acetonitrile by 99% buffer A+1% acetonitrile linear transformation, increased organic phase (acetonitrile) ratio gradually to reduce moving phase polarity; (4) 27.0 ~ 33.0 minutes, follow the non-linear commutation mode that is of curve 8 representatives, moving phase is transformed to 91% buffer A+9% acetonitrile gradually by 95% buffer A+5% acetonitrile, further reduces moving phase polarity, makes that the stronger component of reservation elutes on chromatographic column; (5) 33.0 ~ 33.5 minutes, moving phase was 60% pure water+40% acetonitrile by 91% buffer A+9% acetonitrile linear transformation, begins to wash chromatographic column; (6) 33.5 ~ 34.0 minutes, moving phase became 40% pure water+60% acetonitrile from 60% pure water A+40% acetonitrile, further strengthened the organic phase ratio to strengthen backwash rate; (7) 34.0 ~ 37.0 minutes, washed chromatographic column 3 minutes with 40% pure water+60% acetonitrile; (8) 37.1 minutes, moving phase was transformed to 100% buffer A, beginning balance chromatographic column; (9) 37.1 ~ 45.0 minutes, to original state, ready for analyzing next sample with 100% buffer A balance chromatographic column.
This method is in preceding ultraviolet and the fluorescence dual-detector series connection trace routine of preferably including of step (two); after soon diode array or UV-detector and fluorescence detector will be connected on chromatographic column; taurine derivative products in the sample is realized separating on chromatographic column with other interfering material; and successively by diode array or UV-detector and fluorescence detector; utilize the ultraviolet and the fluorescent characteristic of test substance to detect respectively, thereby realize the dual qualitative and quantitative of taurine.
Can also comprise proving program after step (three): the method evaluation index comprises that recovery of standard addition, reappearance and minimum detection limit etc. reach the analysis requirement of beer sample.
The present invention adopts high performance liquid chromatography; utilize the AQC column front derivation; diode array (or ultraviolet) and the series connection of fluorescence dual-detector detect; dual qualitative and quantitative; at this complex sample that is rich in multiple free amino acid of beer; by regulating buffer concentration, pH and solvent gradient elution, having realized in the beer that taurine and other are amino acid whose separates fully, thereby it is carried out accurately qualitative and quantitative.Method is simple to operate, accurately and reliably, has higher sensitivity.The recovery of standard addition 97.1%-102.0% of method, reappearance RSD (n=6)≤2.2%, minimum detection limit 0.5mg/L.
Description of drawings
Fig. 1 is high performance liquid chromatography ultraviolet (HPLC-UV) the contrast spectrogram of taurine standard specimen and functional beer sample.
Fig. 2 is high performance liquid chromatography fluorescence (HPLC-FLD) the contrast spectrogram of taurine standard specimen and functional beer sample.
Embodiment
Embodiment 1.
A kind of method that adopts taurine in the high performance liquid chromatography measuring ability beer of the present invention comprises the steps:
(1) pre-treatment program of deriving: 6-aminoquinoline base-N-hydroxyl succinimide carbamate (AQC) column front derivation:
The preparation of liquid 1.1 derive
Heating arrangement is preheated to 55 degrees centigrade, before opening the 2A bottle, knock gently, guarantee that all reagent powder all drop on bottle at the bottom of, by drawing the 1ml thinning agent in the 2B bottle and bleeding off, to clean micropipet, draw the 1ml thinning agent and put into the 2A bottle that the derivating agent powder is housed, seal, vortex shook for 10 seconds, 55 degrees centigrade of heating 2A bottles all dissolve until the derivating agent powder in baking oven.Be no more than 10 minutes heat time heating time.
1.2 sample preparation:
Use the 0.45um membrane filtration after the beer exhaust, can suitably dilute according to content of taurine in the testing sample.
1.3 derivatization reaction:
The beer sample that pipettes 10ul taurine standard operation liquid or handle well adds 70ul borate buffer solution (Buffer 1), vortex mixed in the pipe of deriving.Pipette 20 Ma derivating agents, vortex limit, limit adds in the pipe of deriving, and continues for 10 seconds.The pipe of will deriving places 55 degrees centigrade of baking ovens heating 10 minutes, then with sample transfer to the micro-sampling bottle.
(2) ultraviolet and fluorescence dual-detector series connection trace routine
After diode array or UV-detector and fluorescence detector be connected on chromatographic column; taurine derivative products in the sample is realized separating on chromatographic column with other interfering material; and successively by diode array or UV-detector and fluorescence detector; utilize the ultraviolet and the fluorescent characteristic of test substance to detect respectively, thereby realize the dual qualitative and quantitative of taurine.
(3) gradient elution program:
3.1 taurine standard solution preparation
Earlier taurine (Tau) standard reserving solution of preparation 2.5mmol/L pipettes Tau standard reserving solution 100ul with liquid-transfering gun then, adds the 900ul ultrapure water, and the vortex mixing obtains the taurine standard operation liquid of 0.25mmol/L.
3.2 the preparation of mobile phase A:
Take by weighing sodium acetate trihydrate 19.04g,, add the 2.37ml triethylamine, be settled to 1L with the ultrapure water dissolving.Regulate pH to 4.98 with 50% phosphoric acid then, the 0.45um water-based membrane filtration and the degassing.
That is, mobile phase A is the damping fluid of 139.92mmol/L sodium acetate trihydrate+16.39mmol/L triethylamine, with 50% phosphoric acid (H
3PO
4) regulate pH to 4.98, optimize the gradient elution program at standard specimen and beer sample respectively, make that other interfering material separates fully in taurine and the beer.
The eluent gradient table:
Annotate:
1.A be the buffer salt solution of pH4.98; B is a pure water; C is an acetonitrile.
2. the linear commutation of curve 6 representatives, curve 11 is represented the moment commutation, and the non-linear commutation of certain rule is followed in curve 8 representatives, is finished automatically according to setting value by instrument.Curve number is that U.S. Waters company liquid chromatograph carries function, in order to adjust proportion of mobile phase.
3. the eluent gradient tabulation is released: in (1) 0 ~ 0.5 minute, moving phase is 100% buffer A, carries out isocratic elution; In the time of (2) 0.5 minutes, moving phase moment switches to 99% buffer A+1% acetonitrile; (3) 0.5 ~ 27.0 minutes, moving phase was 95% buffer A+5% acetonitrile by 99% buffer A+1% acetonitrile linear transformation, increased organic phase (acetonitrile) ratio gradually to reduce moving phase polarity; (4) 27.0 ~ 33.0 minutes, follow the non-linear commutation mode that is of curve 8 representatives, moving phase is transformed to 91% buffer A+9% acetonitrile gradually by 95% buffer A+5% acetonitrile, further reduces moving phase polarity, makes that the stronger component of reservation elutes on chromatographic column; (5) 33.0 ~ 33.5 minutes, moving phase was 60% pure water+40% acetonitrile by 91% buffer A+9% acetonitrile linear transformation, begins to wash chromatographic column; (6) 33.5 ~ 34.0 minutes, moving phase became 40% pure water+60% acetonitrile from 60% pure water A+40% acetonitrile, further strengthened the organic phase ratio to strengthen backwash rate; (7) 34.0 ~ 37.0 minutes, washed chromatographic column 3 minutes with 40% pure water+60% acetonitrile; (8) 37.1 minutes, moving phase was transformed to 100% buffer A, beginning balance chromatographic column; (9) 37.1 ~ 45.0 minutes, to original state, ready for analyzing next sample with 100% buffer A balance chromatographic column.
(4) stratographic analysis and data processor
Chromatographic column: AccQ-Tag amino acid analysis C18 post (3.9mm*50mm, 4um), Waters company;
Column temperature: 37 degrees centigrade;
Sample size: 10ul;
Ultraviolet detection wavelength: 248nm;
Fluoroscopic examination: excitation wavelength 250nm, emission wavelength 395nm.
See figures.1.and.2, beer to be measured and the contrast of taurine standard specimen spectrogram is qualitative with retention time, the chromatographic peak peak area quantification.Computing formula is as follows:
In the formula: C
SampleThe content of taurine in-----beer to be measured, mg/L;
C
Mark----content of taurine in the-standard specimen, mg/L;
A
SampleThe peak area of taurine in-----beer to be measured;
A
Mark----peak area of taurine in the-standard specimen;
The dilution gfactor of f-----beer to be measured.
(5) proving program: the method evaluation index comprises that recovery of standard addition, reappearance and minimum detection limit etc. reach the analysis requirement of beer sample.
Add the taurine standard solution of variable concentrations in functional beer, addition is respectively 1.0mg/L, 2.5mg/L and 5.0mg/L, records recovery of standard addition between 97.1%～102.0%.
Same beer sample is handled by preceding method, parallel doing 6 times, the RSD of taurine retention time≤0.09%, the RSD of peak area≤2.2%, reappearance is better.
With the required sample size of the response of 3 times of noises is detection limit, and detecting of taurine is limited to 0.5mg/L, satisfies beer and detects needs.
The apparatus and the reagent that adopt in the said method are as follows:
Apparatus: Waters Alliance system: 2695 separative elements, 2996 diode array detector, 2475 fluorescence detectors, Empower chromatogram management system, Waters company.The Delta320pH meter, MettlerToledo company.Chromatographic column: AccQ-Tag amino acid analysis C
18Post (3.9mm*50mm, 4um), Waters company.
Reagent: kit: borate buffer (Buffer 1), derivating agent powder 2A, derivating agent dilution 2B, Waters company; Taurine, sodium acetate trihydrate, triethylamine, Fluka company, wherein taurine is a chromatographically pure, it is pure that other medicine is analysis; Acetonitrile, UV level, Burdick﹠amp; Jackson company; Sodium azide is analyzed pure.
Because beer matrix more complicated, must operate in strict accordance with this chromatographic condition, the particularly concentration of damping fluid and pH, and can the gradient program be the key point that carry out accurate separation and quantitative to the taurine in the beer.
The present invention describes with reference to drawings and Examples, but protection domain is not limited thereto, and has those of ordinary skill and can pass through simple conversion in the technology of the present invention scope, and obtain the same technique effect of the present invention, equally in protection scope of the present invention.
Claims (6)
1. method that adopts taurine in the high performance liquid chromatography measuring ability beer is characterized in that comprising the steps: (one) pre-treatment program of deriving: 6-aminoquinoline base-N-hydroxyl succinimide carbamate column front derivation.
2. a kind of method that adopts taurine in the high performance liquid chromatography measuring ability beer as claimed in claim 1, it is characterized in that also comprising (two) gradient elution program: mobile phase A is the damping fluid of 139.92mmol/L sodium acetate trihydrate+16.39mmol/L triethylamine, regulate pH to 4.98 with 50% phosphoric acid, Mobile phase B is a pure water, moving phase C is an acetonitrile, optimize the gradient elution program at standard specimen and beer sample respectively, make that other interfering material separates fully in taurine and the beer;
(3) stratographic analysis and data processor:
The employed chromatographic column of sample analysis is the AccQ-Tag amino acid analysis C18 post of U.S. Waters company, 3.9mm*50mm, and 4um, beer to be measured and the contrast of taurine standard specimen spectrogram is qualitative with retention time, the chromatographic peak peak area quantification.
3. a kind of method that adopts taurine in the high performance liquid chromatography measuring ability beer as claimed in claim 2, it is characterized in that described step (two) gradient elution program is: in (1) 0 ~ 0.5 minute, moving phase is 100% buffer A, carries out isocratic elution; In the time of (2) 0.5 minutes, moving phase moment switches to 99% buffer A+1% acetonitrile; (3) 0.5 ~ 27.0 minutes, moving phase was 95% buffer A+5% acetonitrile by 99% buffer A+1% acetonitrile linear transformation, increased organic phase (acetonitrile) ratio gradually to reduce moving phase polarity; (4) 27.0 ~ 33.0 minutes, follow the non-linear commutation mode that is of curve 8 representatives, moving phase is transformed to 91% buffer A+9% acetonitrile gradually by 95% buffer A+5% acetonitrile, further reduces moving phase polarity, makes that the stronger component of reservation elutes on chromatographic column; (5) 33.0 ~ 33.5 minutes, moving phase was 60% pure water+40% acetonitrile by 91% buffer A+9% acetonitrile linear transformation, begins to wash chromatographic column; (6) 33.5 ~ 34.0 minutes, moving phase became 40% pure water+60% acetonitrile from 60% pure water A+40% acetonitrile, further strengthened the organic phase ratio to strengthen backwash rate; (7) 34.0 ~ 37.0 minutes, washed chromatographic column 3 minutes with 40% pure water+60% acetonitrile; (8) 37.1 minutes, moving phase was transformed to 100% buffer A, beginning balance chromatographic column; (9) 37.1 ~ 45.0 minutes, to original state, ready for analyzing next sample with 100% buffer A balance chromatographic column.
4. as claim 2 or 3 described a kind of methods that adopt taurine in the high performance liquid chromatography measuring ability beer; it is characterized in that preceding diode array or ultraviolet and the fluorescence dual-detector series connection trace routine of comprising of step (two); after soon diode array or UV-detector and fluorescence detector will be connected on chromatographic column; taurine derivative products in the sample is realized separating on chromatographic column with other interfering material; and successively by diode array or UV-detector and fluorescence detector; utilize the ultraviolet and the fluorescent characteristic of test substance to detect respectively, thereby realize the dual qualitative and quantitative of taurine.
5. as claim 2 or 3 described a kind of methods that adopt taurine in the high performance liquid chromatography measuring ability beer, it is characterized in that step (three) computing formula is as follows:
In the formula: C
SampleThe content of taurine in-----beer to be measured, mg/L;
C
Mark----content of taurine in the standard specimen, mg/L;
A
Sample--the peak area of taurine in----beer to be measured;
A
Mark------peak area of taurine in the standard specimen;
The dilution gfactor of f-----beer to be measured.
6. as claim 2 or 3 described a kind of methods that adopt taurine in the high performance liquid chromatography measuring ability beer; it is characterized in that comprising proving program after the step (three), the method evaluation index comprises that recovery of standard addition, reappearance and minimum detection limit etc. reach the analysis requirement of beer sample.
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| CN103454358A (en) * | 2013-09-13 | 2013-12-18 | 厦门大学 | Method for detecting taurine |
| CN103822894A (en) * | 2014-02-11 | 2014-05-28 | 广东恒兴饲料实业股份有限公司 | Method for detecting sulfonic acid content of fishmeal based on near infrared spectroscopy method |
| CN104330377A (en) * | 2014-10-28 | 2015-02-04 | 南京白云化工环境监测有限公司 | Method for measuring 16 polycyclic aromatic hydrocarbons by using serial ultraviolet fluorescence detector |
| CN105044019A (en) * | 2015-06-01 | 2015-11-11 | 浙江万里学院 | Rapid detection method for taurine content in marine product processing process |
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