CN108414658A - A kind of method that UPLC-Q-TOF-MS-MS combinations measure eight kinds of sweeteners in white wine - Google Patents
A kind of method that UPLC-Q-TOF-MS-MS combinations measure eight kinds of sweeteners in white wine Download PDFInfo
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Abstract
The invention discloses a kind of UPLC Q TOF MS MS to be combined the method for measuring eight kinds of sweeteners in white wine, and wine sample to be measured first is evaporated by boiling water bath except alcohol utilizes C then using UPLC as piece-rate system18Chromatographic column carries out gradient elution separation to wine sample to be measured, segment data acquisition is carried out using MS MS patterns during gradient elution, it is monitoring ion with the characteristic molecular ion of eight kinds of sweeteners and fragment ion, the sample to be tested spectrogram of acquisition and standard items spectrogram are compared, identical retention time is tie substance, is then quantified to each sweetener by equation of linear regression.For the method for the present invention detection limit down to 10 μ g/L, linearly dependent coefficient is more than 0.99, and for the rate of recovery between 80% 120%, associated standard deviation illustrates that eight kinds of sweeteners can be detected with this method in white wine within 10%.
Description
Technical field
The present invention relates to a kind of UPLC-Q-TOF-MS-MS to be combined the method for measuring eight kinds of sweeteners in white wine, belongs to fermentation
Chemical analysis in industry and instrument analysis technology field.
Background technology
Caused by the pure and sweet sense of traditional high quality liquor is the sweet substances such as polyalcohol or ketone by itself containing, such as 2,3- fourths
Glycol, glycerine, biacetyl etc..In order to adapt to new consumption demand, increases the sweet taste of white wine, returns sweet tea sense, and can effectively improve
Production efficiency becomes the project that numerous Liquor-making Enterprises ＆s are competitively studied.Many enterprises are in order to improve mouthfeel now, to the white wine of production
The middle various sweeteners of addition, and do not allow addition non-fermented generation in national standard clear stipulaties white spirit by solid state method is in fragrant flavor
Substance.Only allow in solid-liquid and white wine of liquid with fragrant unstrained spirits flavour passage or use food additives seasoning and adjusting incense, without allowing addition times
What sweetener.
It is found according to investigation, the sweetener that may be now detected from the white wine spot-check in market has following 7 kinds：Saccharin sodium, peace
Match honey, honey element, Aspartame, neotame, Sucralose and steviol glycoside etc..Saccharin sodium, also known as soluble saccharin are a kind of white
The sweetener of color crystalline powder, sugariness are 300-500 times of sucrose, and non-energy sweetener is not involved in the metabolic adsorption of human body,
But large amounts of food can cause damages to liver and nervous system.Acesulfame potassium also known as acesulfame potassium, 3,4-Dihydro-6-methyl-1,2,3-oxathiazin-4-one 2,2-dioxide potassium salt are white crystalline
Powder is highly soluble in water, non-hygroscopic, and sugariness is 200 times of sucrose, with no nutrition, empty calory, is not metabolized in human body, no
The characteristics of absorption, but it can result in harm to liver and nervous system.Honey element is white needles, flaky crystal or crystalloid
Powder also known as cyclohexylamine sulfonic acid potassium, be sulfamic acid and cyclohexylamine and sodium hydroxide reaction and made of non-nutritive synthesize
Sweetener, sugariness are 30 times of sucrose, and good water solubility, absolute acid stability is strong, and cost is relatively low, but studies have shown that edible honey element
Carcinoma of urinary bladder can be caused with the mixture of saccharin.Aspartame also known as sweetener, protein sugar, aspartame, radix asparagi sweet extract, day benzene sugar
Deng, be by L-phenylalanine first with methanol esterification after again with amino acid L-Aspartic acid condensation amidation generate it is a kind of through methyl
Dipeptide compounds after esterification, sugariness are 200 times of sucrose, account for the 62% of world's sugar substitute market share, due to its height
It is sugariness, low in calories, non-hazardous to human body, it is suitable for all other men group except phenyl ketonuria patient, is widely used in food
In.Neotame is a kind of white crystalline powder, and noenergy belongs to dipeptide intense sweetener, and sugariness is 7000-13000 times of sucrose,
40 times of Aspartame, sweet taste is pure, no bitter taste and metallic taste, because its stability is good, sweet taste coordinate, without saprodontia, be applicable in
Crowd is wide, safe, is applied in various food by vast enterprise.Sucralose, a kind of white powder substance are easily molten
Yu Shui, it is 600-800 times of sucrose to have preferable absolute acid stability, photostability and thermal stability, sugariness, and noenergy, sweet taste is pure,
Zero calory will not cause saprodontia, be suitble to any crowd, it is considered to be current safest sweetener, by including China
More than 30 a state approvals can be used as sweetener use.Steviol glycoside sterling is white crystalline powder, soluble easily in water, methanol, second
Alcohol, it is 200-300 times of sucrose to have preferable heat resistance, salt tolerance and stability, sugariness, and calorific value is the 1/300 of sucrose, to people
Body is without side-effects, is the natural low caloric value sweetener closest to sucrose taste for ratifying to use through the Ministry of Public Health of China, Ministry of Light Industry,
It is the 3rd kind of natural sucrose substitute for having Development volue and health to praise highly except sugarcane, beet sugar, is described as in the world
" third place in the world sucrose ", therefore many food processing enterprises are as the substitute of sucrose.
Currently, the detection method to sweetener in white wine mainly has：High performance liquid chromatography, the chromatography of ions, efficient liquid
One mass spectrometric hyphenated technique of phase chromatography, one mass spectrometric hyphenated technique of ultra-performance liquid chromatography, gas chromatography, thin layer chromatography, ratio
Color method etc..Thin layer chromatography and colorimetric method are mainly used in the detection of Determination of Saccharin in Food, and because detection limit is relatively high, operation is numerous
Trivial, poor reproducibility has been rarely used in the detection of sweetener in food, and after the detection national standard revision of Determination of Saccharin in Food
This two methods is deleted.
Invention content
The present invention is intended to provide a kind of UPLC-Q-TOF-MS-MS (ultra performance liquid chromatography and flight time mass spectrum) combination
Measure white wine in eight kinds of sweeteners method, can in white wine Simultaneous Quantitative Analysis acesulfame potassium, saccharin sodium, honey element, A Siba
Eight kinds of sweet tea, alitame, neotame, Sucralose, steviol glycoside sweeteners.The pre-treatment of detection method is simple, is boiling water
Bath evaporation removes alcohol, and for quantitative approach detection limit down to 10 μ g/L, linearly dependent coefficient is more than 0.99, the rate of recovery 80-120% it
Between, associated standard deviation illustrates that above eight kinds of sweeteners in white wine can carry out standard by the method for the invention within 10%
Really detection, and method is simple, efficient, high sensitivity.
The method that UPLC-Q-TOF-MS-MS combinations of the present invention measure eight kinds of sweeteners in white wine, wine sample to be measured first pass through
Boiling water bath evaporation utilizes C except alcohol then using UPLC as piece-rate system18Chromatographic column carries out gradient elution separation to wine sample to be measured,
Segment data acquisition is carried out using MS-MS patterns during gradient elution, with the characteristic molecular ion of eight kinds of sweeteners and from
Sub- fragment is monitoring ion, and the sample to be tested spectrogram of acquisition and standard items spectrogram are compared, and identical retention time is to correspond to
Then substance quantifies each sweetener by equation of linear regression.
The method that UPLC-Q-TOF-MS-MS of the present invention measures eight kinds of sweeteners in white wine, includes the following steps：
Step 1：Pre-treatment
The ethyl alcohol in wine sample to be measured is removed using boiling water bath evaporation, the ethyl alcohol contained in white wine is more, if in preceding processing
In do not remove ethyl alcohol, it will generate matrix effect, influence the quantitative analysis of sweetener in white wine.
Step 2：Chromatographic test strip part
Chromatographic column：BEH C18(2.1mm×50mm 1.7μm)；
Mobile phase：Mobile phase A：0.1% acetic acid-water, Mobile phase B：0.1% acetic acidacetonitrile；
The process for preparation of mobile phase A is as follows：1mL acetic acid is measured in 1L volumetric flasks, 1L is settled to ultra-pure water, 0.22 μm
Water phase filter membrane is for use after filtering.
The process for preparation of Mobile phase B is as follows：0.5mL acetic acid is measured in 500mL volumetric flasks, with acetonitrile (chromatographically pure) constant volume
To 500mL.
Eluent gradient elutes ratio：
| Time (min) | Flow velocity (mL/min) | A (%) | B (%) |
| 0.0 | 0.2 | 90.0 | 10.0 |
| 4.0 | 0.2 | 80.0 | 20.0 |
| 6.0 | 0.2 | 10.0 | 90.0 |
| 8.0 | 0.2 | 10.0 | 90.0 |
| 8.1 | 0.2 | 90.0 | 10.0 |
| 10.0 | 0.2 | 90.0 | 10.0 |
Flow velocity：0.2mL/min；
Pressure limit：0-18000psi；
Column temperature：30℃；
Sampling volume：5μL.
Step 3：Mass Spectrometer Method condition
MS-MS patterns；
ESI anionic textiles patterns；
Orifice potential：20-40V；
Ion source temperature：120℃；
Desolvation temperature：500℃；
Desolventizing gas flow：800L/h；
Taper hole throughput：50L/h；
Mass range：50-1200Da.
Step 4：The preparation of standard items
Prepare respectively the acesulfame potassium of 1g/L, saccharin sodium, honey element, Sucralose, Aspartame, alitame, steviol glycoside,
The standard solution of eight kinds of sweeteners is mixed and is diluted step by step, obtains the hybrid standard product of various concentration by the standard solution of neotame
Solution；
In step 4, dilution step by step obtains the hybrid standard product solution of at least five various concentrations, in hybrid standard product solution
The μ g/L of the concentration of each sweetener >=50.
Step 5：The drafting of standard spectrogram and standard curve
The hybrid standard product solution sample presentation for the various concentration that step 4 is prepared carries out chromatography eluant separation, according to eight kinds of sweet teas
Taste agent retention time (acesulfame potassium 1.07min, saccharin sodium 1.41min, honey element 1.60min, Sucralose 2.81min, A Siba
Sweet tea 3.29min, alitame 4.30min, steviol glycoside 5.52min, neotame 5.75min) the different mistakes in chromatography eluant separation
Piecewise acquisition is carried out using MS-MS patterns in journey, the standard spectrogram of eight kinds of sweetener standard items is obtained, with the dense of each sweetener
Degree maps to its retention time, obtains its corresponding standard curve；Equation of linear regression and detection limit see the table below：
| Component to be measured | The range of linearity | Equation of linear regression | Linearly dependent coefficient | Detection limit (mg/L) |
| Acesulfame potassium | 0.05-0.2 | 5.56575*X-70.8607 | 0.999 | 0.01 |
| Saccharin sodium | 0.05-0.2 | 1.02524*X-15.9781 | 0.998 | 0.01 |
| Honey element | 0.05-0.2 | 20.642*X-333.272 | 0.998 | 0.01 |
| Sucralose | 0.05-0.2 | 21.1612*X-615.415 | 0.997 | 0.05 |
| Aspartame | 0.05-0.2 | 7.11332*X-150.8 | 0.999 | 0.01 |
| Neotame | 0.05-0.2 | 47.6501*X-1410.49 | 0.998 | 0.01 |
| Alitame | 0.05-0.2 | 44.6377*X-887.097 | 0.999 | 0.01 |
| Steviol glycoside | 0.1-0.5 | 52.634*X-3347.87 | 0.999 | 0.01 |
Step 6：The detection of sample to be tested
Wine sample sample introduction to be measured after step 1 pre-treatment is subjected to chromatography eluant separation, in the process of chromatography eluant separation
The middle MS-MS pattern piecewise acquisition data using UPLC-Q-TOF-MS-MS, by the sample to be tested spectrogram and standard spectrogram of acquisition
Comparison, identical retention time is tie substance, is quantified to each sweetener by standard curve equation of linear regression.For
The accuracy for ensuring sample, uses the daughter ion of each component to monitor ion, in sample with and standard items in monitoring ion phase
Together, can further determine that in sample in standard items be same substance.
According to the retention time of sweetener, the MS-MS pattern piecewise acquisition data of UPLC-Q-TOF-MS-MS, An Sai are utilized
The acquisition time of honey is 0-1.3min, and the acquisition time of saccharin sodium is 1.3-1.5min, and the acquisition time of honey element is 1.5-
The acquisition time of 1.7min, Sucralose are 1.7-3.0min, and the acquisition time of Aspartame is 3.0-3.5min, alitame
Acquisition time is 3.5-4.5min, and the acquisition time of steviol glycoside is 4.5-5.65, and the acquisition time of button sweet tea is 5.65-
10.0min；It is monitored under ESI negative ion modes, it is 161.9861 to select acesulfame potassium qualitative, quantitative ion pair>82.0289 honey element
Qualitative, quantitative ion pair is 178.0538>79.9559, alitame qualitative, quantitative ion pair is 330.1488>312.1366, trichlorine
Sucrose qualitative, quantitative ion pair is 395.0067>395.0067, button sweet tea qualitative, quantitative ion pair is 377.2076>200.0711
Aspartame qualitative, quantitative ion pair is 293.1137>200.0711 saccharin sodium qualitative, quantitative ion pair is 181.9912>
105.9600 steviol glycoside 803.3701>640.1577 detection time 10min.
The pre-treatment of the method for the present invention is simple, and ultra high efficiency liquid phase systems can carry out eight kinds of sweeteners within 10min fast
Speed separation, and qualitative, quantitative is carried out using eight kinds of sweeteners of the MS-MS piecewise acquisitions pattern pair of UPLC-Q-TOF-MS-MS, not only
It can ensure that the qualitative accuracy of each substance, while piecewise acquisition can make the collection of illustrative plates of each substance collect more data points, with
Ensure quantitative accuracy.The method of the present invention is easy, and rapidly and efficiently, the quick detection of eight kinds of sweeteners suitable for white wine is
The detection of eight kinds of sweeteners provides new detection means in white wine.
Description of the drawings
Fig. 1 is acesulfame potassium daughter ion ion flow graph.
Fig. 2 is saccharin sodium daughter ion ion flow graph.
Fig. 3 is honey element daughter ion ion flow graph.
Fig. 4 is Sucralose daughter ion ion flow graph.
Fig. 5 is Aspartame daughter ion ion flow graph.
Fig. 6 is alitame daughter ion ion flow graph.
Fig. 7 is steviol glycoside daughter ion ion flow graph.
Fig. 8 is neotame daughter ion ion flow graph.
Fig. 9 is acesulfame potassium standard curve.
Figure 10 is saccharin sodium standard curve.
Figure 11 is sodium cyclamate standard curve.
Figure 12 is Sucralose standard curve.
Figure 13 is Aspartame standard curve.
Figure 14 is alitame standard curve.
Figure 15 is steviol glycoside standard curve.
Figure 16 is neotame standard curve.
Figure 17 is mark-on wine sample ion flow graph.
Specific implementation mode
Explanation is further analyzed to technical solution of the present invention below by specific embodiment.
1 material
1.1 instrument
Ultra performance liquid chromatography system (ACQUITY Ultra Performance LC Waters companies) is equipped with mass spectrum
(G2-Q Tof time of-flight mass spectrometer Waters companies), electric furnace (Beijing forever bright Medical Devices Co., Ltd.), ACQUITY
UPLC Ben C18 Column (1.7 μm, 2.1 × 50mm) chromatographic column.
1.2 reagent
Eight kinds of sweetener standard items (Dr.Ehrenstorfer)；Acetonitrile (chromatographically pure)；Formic acid (chromatographically pure)；Use for laboratory
Water is Milli-Q ultra-pure waters (Millipore companies of the U.S.).
1.3 wine samples
Wine Sample to be checked be 8 kinds of sweeteners mark-on positive wine sample (8 kinds of sweeteners of spiked levels are 100 μ g/L) and
Blank control wine sample.
1.4 standard reserving solution
50mg acesulfame potassiums, saccharin sodium, honey element, Sucralose, Aspartame, alitame, steviol glycoside, knob are weighed respectively
The standard items of sweet tea are settled to 50mL with 50% methanol solution, the single sweetener standard items of sweetener of 1g/L are obtained, in 4 DEG C of ice
It is preserved in case.
2, pre-treatment
The ethyl alcohol in wine sample to be measured is removed using boiling water bath evaporation, the ethyl alcohol contained in white wine is more, if in preceding processing
In do not remove ethyl alcohol, it will generate matrix effect, influence the quantitative analysis of sweetener in white wine.
3, chromatographic test strip part
Chromatographic column：BEH C18(2.1mm×50mm 1.7μm)；
Mobile phase：Mobile phase A：0.1% acetic acid-water, Mobile phase B：0.1% acetic acidacetonitrile；
The process for preparation of mobile phase A is as follows：1mL acetic acid is measured in 1L volumetric flasks, 1L is settled to ultra-pure water, 0.22 μm
Water phase filter membrane is for use after filtering.
The process for preparation of Mobile phase B is as follows：0.5mL acetic acid is measured in 500mL volumetric flasks, with acetonitrile (chromatographically pure) constant volume
To 500mL.
Eluent gradient elutes ratio：
| Time (min) | Flow velocity (mL/min) | A (%) | B (%) |
| 0.0 | 0.2 | 90.0 | 10.0 |
| 4.0 | 0.2 | 80.0 | 20.0 |
| 6.0 | 0.2 | 10.0 | 90.0 |
| 8.0 | 0.2 | 10.0 | 90.0 |
| 8.1 | 0.2 | 90.0 | 10.0 |
| 10.0 | 0.2 | 90.0 | 10.0 |
Flow velocity：0.2mL/min；
Pressure limit：0-18000psi；
Column temperature：30℃；
Sampling volume：5μL.
4, Mass Spectrometer Method condition
MS-MS patterns；
ESI anionic textiles patterns；
Orifice potential：20-40V；
Ion source temperature：120℃；
Desolvation temperature：500℃；
Desolventizing gas flow：800L/h；
Taper hole throughput：50L/h；
Mass range：50-1200Da.
5, the preparation of standard items
Prepare respectively the acesulfame potassium of 1g/L, saccharin sodium, honey element, Sucralose, Aspartame, alitame, steviol glycoside,
The standard solution of eight kinds of sweeteners is mixed and is diluted step by step, obtains the hybrid standard product of various concentration by the standard solution of neotame
Solution, concentration are respectively 50 μ g/L, 80 μ g/L, 100 μ g/L, 150 μ g/L, 200 μ g/L.
6, the drafting of standard spectrogram and standard curve
The hybrid standard product solution sample presentation for the various concentration that step 4 is prepared carries out chromatography eluant separation, according to eight kinds of sweet teas
Taste agent retention time (acesulfame potassium 1.07min, saccharin sodium 1.41min, honey element 1.60min, Sucralose 2.81min, A Siba
Sweet tea 3.29min, alitame 4.30min, steviol glycoside 5.52min, neotame 5.75min) the different mistakes in chromatography eluant separation
Piecewise acquisition is carried out using MS-MS patterns in journey, the standard spectrogram of eight kinds of sweetener standard items is obtained, with the dense of each sweetener
Degree maps to its retention time, obtains its corresponding standard curve；Equation of linear regression and detection limit see the table below：
| Component to be measured | The range of linearity | Equation of linear regression | Linearly dependent coefficient | Detection limit (mg/L) |
| Acesulfame potassium | 0.05-0.2 | 5.56575*X-70.8607 | 0.999 | 0.01 |
| Saccharin sodium | 0.05-0.2 | 1.02524*X-15.9781 | 0.998 | 0.01 |
| Honey element | 0.05-0.2 | 20.642*X-333.272 | 0.998 | 0.01 |
| Sucralose | 0.05-0.2 | 21.1612*X-615.415 | 0.997 | 0.05 |
| Aspartame | 0.05-0.2 | 7.11332*X-150.8 | 0.999 | 0.01 |
| Neotame | 0.05-0.2 | 47.6501*X-1410.49 | 0.998 | 0.01 |
| Alitame | 0.05-0.2 | 44.6377*X-887.097 | 0.999 | 0.01 |
| Steviol glycoside | 0.1-0.5 | 52.634*X-3347.87 | 0.999 | 0.01 |
7, the detection of sample to be tested
Wine sample sample introduction to be measured after step 2 pre-treatment is subjected to chromatography eluant separation, in the process of chromatography eluant separation
The middle MS-MS pattern piecewise acquisition data using UPLC-Q-TOF-MS-MS, by the sample to be tested spectrogram and standard spectrogram of acquisition
Comparison, identical when reservation is tie substance, is quantified to each sweetener by standard curve equation of linear regression.In order to
The accuracy for ensuring sample uses the daughter ion of each component to monitor ion, in sample with and standard items in monitor ion identical,
Can further determine that in sample in standard items be same substance.
Using UPLC-Q-TOF-MS-MS MS-MS pattern piecewise acquisition data when, the acquisition time of acesulfame potassium is 0-
The acquisition time of 1.3min, saccharin sodium are 1.3-1.5min, and the acquisition time of honey element is 1.5-1.7min, and Sucralose is adopted
Integrate the time as 1.7-3.0min, the acquisition time of Aspartame is 3.0-3.5min, and the acquisition time of alitame is 3.5-
The acquisition time of 4.5min, steviol glycoside are 4.5-5.65, and the acquisition time of button sweet tea is 5.65-10.0min；ESI anion moulds
It is monitored under formula, it is 161.9861 to select acesulfame potassium qualitative, quantitative ion pair>82.0289 honey element qualitative, quantitative ion pair is
178.0538>79.9559, alitame qualitative, quantitative ion pair is 330.1488>312.1366 Sucralose qualitative, quantitative ion
To being 395.0067>395.0067, button sweet tea qualitative, quantitative ion pair is 377.2076>200.0711 Aspartame qualitative, quantitative
Ion pair is 293.1137>200.0711 saccharin sodium qualitative, quantitative ion pair is 181.9912>105.9600, steviol glycoside
803.3701>640.1577 detection time 10min.
8, interpretation of result
The detection spectrogram of mark-on positive wine sample is shown in Figure 17.By to 8 kinds of sweetener mark-on positive wine samples (100 μ g/L of mark-on)
And the detection and analysis of blank wine sample, the retention time and quota ion pair of sweetener and sweetener standard items in wine sample are compared, is determined
Sweetener type in wine sample, and by equation of linear regression, obtain detecting without sweetener in blank wine sample, it is each in mark-on wine sample
The content of sweetener, acesulfame potassium, saccharin sodium, honey element, Sucralose, Aspartame, alitame, steviol glycoside, neotame content
Respectively：96.5μg/L、89.6μg/L、98.2μg/L、85.3μg/L、88.7μg/L、92.8μg/L、91.7μg/L、90.4μg/
L illustrates the recovery of standard addition of the method between 85.3%-98.2%, is suitble to the analysis detection of 8 kinds of sweeteners in white wine.
Claims (7)
1. a kind of method that UPLC-Q-TOF-MS-MS combinations measure eight kinds of sweeteners in white wine, it is characterised in that：It is to be measured first
Wine sample is evaporated by boiling water bath except alcohol utilizes C then using UPLC as piece-rate system18Chromatographic column carries out gradient to wine sample to be measured and washes
De- separation, carries out segment data acquisition, with the characteristic molecular of eight kinds of sweeteners during gradient elution using MS-MS patterns
Ion and fragment ion are monitoring ion, and the sample to be tested spectrogram of acquisition and standard items spectrogram are compared, and retention time is identical
Then as tie substance quantifies each sweetener by equation of linear regression.
2. according to the method described in claim 1, it is characterized by comprising following steps：
Step 1：Pre-treatment
The ethyl alcohol in wine sample to be measured is removed using boiling water bath evaporation；
Step 2：Chromatographic test strip part
Chromatographic column：BEH C18(2.1mm×50mm 1.7μm)；
Mobile phase：Mobile phase A：0.1% acetic acid-water, Mobile phase B：0.1% acetic acidacetonitrile；
Type of elution：Gradient elution；
Flow velocity：0.2mL/min；
Pressure limit：0-18000psi；
Column temperature：30℃；
Sampling volume：5μL；
Step 3：Mass Spectrometer Method condition
MS-MS patterns；
ESI anionic textiles patterns；
Orifice potential：20-40V；
Ion source temperature：120℃；
Desolvation temperature：500℃；
Desolventizing gas flow：800L/h；
Taper hole throughput：50L/h；
Mass range：50-1200Da.
Step 4：The preparation of standard items
The acesulfame potassium of preparation 1g/L, saccharin sodium, honey element, Sucralose, Aspartame, alitame, steviol glycoside, neotame respectively
Standard solution, the standard solution of eight kinds of sweeteners is mixed and is diluted step by step, the hybrid standard product solution of various concentration is obtained；
Step 5：The drafting of standard spectrogram and standard curve
The hybrid standard product solution sample presentation for the various concentration that step 4 is prepared carries out chromatography eluant separation, according to eight kinds of sweeteners
The difference of retention time carries out piecewise acquisition during chromatography eluant detaches using MS-MS patterns, obtains eight kinds of sweeteners
The standard spectrogram of standard items maps to its retention time with the concentration of each sweetener, obtains its corresponding standard curve；
Step 6：The detection of sample to be tested
Wine sample sample introduction to be measured after step 1 pre-treatment is subjected to chromatography eluant separation, it is sharp during chromatography eluant detaches
With the MS-MS pattern piecewise acquisition data of UPLC-Q-TOF-MS-MS, the sample to be tested spectrogram of acquisition and standard spectrogram are compared,
Identical retention time is tie substance, is quantified to each sweetener by standard curve equation of linear regression.
3. according to the method described in claim 2, it is characterized in that：
In step 2, it is as follows that eluent gradient elutes ratio：
4. according to the method described in claim 2, it is characterized in that：
It in step 3, is monitored under ESI negative ion modes, it is 161.9861 to select acesulfame potassium qualitative, quantitative ion pair>82.0289 sweet tea
Sweet element qualitative, quantitative ion pair is 178.0538>79.9559, alitame qualitative, quantitative ion pair is 330.1488>312.1366
Sucralose qualitative, quantitative ion pair is 395.0067>395.0067, button sweet tea qualitative, quantitative ion pair is 377.2076>
200.0711 Aspartame qualitative, quantitative ion pair is 293.1137>200.0711 saccharin sodium qualitative, quantitative ion pair is
181.9912>105.9600 steviol glycoside 803.3701>640.1577 detection time 10min.
5. according to the method described in claim 2, it is characterized in that：
In step 4, dilution step by step obtains the hybrid standard product solution of at least five various concentrations, each sweet tea in hybrid standard product solution
The μ g/L of the concentration of taste agent >=50.
6. according to the method described in claim 2, it is characterized in that：
In step 5, when carrying out piecewise acquisition using MS-MS patterns, the acquisition time of acesulfame potassium is 0-1.3min, and saccharin sodium is adopted
Integrate the time as 1.3-1.5min, the acquisition time of honey element is 1.5-1.7min, and the acquisition time of Sucralose is 1.7-
The acquisition time of 3.0min, Aspartame are 3.0-3.5min, and the acquisition time of alitame is 3.5-4.5min, steviol glycoside
Acquisition time is 4.5-5.65, and the acquisition time of button sweet tea is 5.65-10.0min.
7. according to the method described in claim 2, it is characterized in that：
In step 6, in order to ensure the accuracy of sample, use the daughter ion of each component to monitor ion, in sample with and standard
In product monitor ion it is identical, can further determine that in sample in standard items be same substance.
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| CN109490452A (en) * | 2018-10-31 | 2019-03-19 | 中国农业科学院茶叶研究所 | Method that is a kind of while detecting 6 kinds of synthetic sweeteners in tealeaves |
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