CN114236032A - Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry - Google Patents
Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry Download PDFInfo
- Publication number
- CN114236032A CN114236032A CN202111489161.8A CN202111489161A CN114236032A CN 114236032 A CN114236032 A CN 114236032A CN 202111489161 A CN202111489161 A CN 202111489161A CN 114236032 A CN114236032 A CN 114236032A
- Authority
- CN
- China
- Prior art keywords
- glyphosate
- water
- sample
- solution
- ion
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/88—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86
- G01N2030/8809—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample
- G01N2030/884—Integrated analysis systems specially adapted therefor, not covered by a single one of the groups G01N30/04 - G01N30/86 analysis specially adapted for the sample organic compounds
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The invention provides a method for determining glyphosate in water body by combining a double ionization scanning mode and liquid chromatography-tandem mass spectrometry, which comprises the steps of carrying out chelate fracture and derivatization on a pretreated water sample, firstly adding 1,2-13C into the pretreated water sample215N-tracing glyphosate aqueous solution to be used as an internal standard for derivatization, adding EDTA-Na2 aqueous solution, shaking up and keeping for a period of time, adding sodium borate buffer solution to adjust the pH value of the system, adding FMOC-Cl acetonitrile solution, performing water bath ultrasound, adding phosphoric acid to neutralize the solution, and finally filtering the obtained solution to obtain a sample to be analyzed; and analyzing and testing the sample to be analyzed by adopting a liquid chromatography-tandem mass spectrometer in a double-ionization scanning mode so as to quantify and qualitatively identify the glyphosate and the main metabolite aminomethyl phosphonic acid thereof. The method can eliminate matrix interference and enhance qualitative accuracy, and the detection limit can reach 0.03 mu g/L.
Description
Technical Field
The invention relates to a method for measuring glyphosate in water by combining a double ionization scanning mode with liquid chromatography-tandem mass spectrometry, belonging to the technical field related to pesticide detection.
Background
The water body is an important component of natural ecological environment, is a complex multi-medium carrier and is also an important attribution of herbicide. The chemical name of the glyphosate is N-phosphine hydroxymethyl glycine, and the glyphosate is one of the widely used herbicides due to the advantages of high efficiency, low toxicity and the like, and is widely used for preventing and removing various weeds and treating non-farmland weeds since the glyphosate is marketed; the glyphosate is continuously enriched in the environment and organisms and enters the organisms through food and drinking water, thus causing harm to human bodies. Therefore, the detection and management of the glyphosate residue in the water body or other media are extremely important.
The limit of glyphosate is specified in the ground water quality standard (GB/T14848-2017), and the limit of class I is 0.1 mu g/L, which is used as a pollution index in ground water for control. At present, standard detection methods of glyphosate are post-column derivatization-high performance liquid chromatography (GB/T5750.9-200618) and ion chromatography (CJ/T141-20187.14), but the detection limits of the two methods are high, wherein the detection limit of the post-column derivatization-high performance liquid chromatography is 25 mug/L, the detection limit of the ion chromatography is 44 mug/L, the requirements of 'underground water quality standard' evaluation cannot be met, and the chromatography method is difficult to determine.
Therefore, the method for determining glyphosate in water by combining the double ionization scanning mode with the liquid chromatography-tandem mass spectrometry is an urgent technical problem in the field.
Disclosure of Invention
In order to solve the defects and shortcomings, the invention aims to provide a method for measuring glyphosate in water by combining a double ionization scanning mode with liquid chromatography-tandem mass spectrometry.
In order to achieve the above object, the present invention provides a method for measuring glyphosate in a water body by a double ionization scanning mode-liquid chromatography tandem mass spectrometry, wherein the method comprises:
(1) and carrying out chelating fracture and derivatization on the pretreated water sample.
Firstly, adding 1,2-13C into the pretreated water sample215N-tracing glyphosate aqueous solution to be used as an internal standard for derivatization, adding EDTA-Na2 aqueous solution, shaking up and keeping for a period of time, adding sodium borate buffer solution to adjust the pH value of the system, adding FMOC-Cl acetonitrile solution, performing water bath ultrasound, adding phosphoric acid to neutralize the solution, and finally filtering the obtained solution to obtain a sample to be analyzed;
(2) and analyzing and testing the sample to be analyzed by adopting a liquid chromatography-tandem mass spectrometer in a double-ionization scanning mode so as to quantify and qualitatively identify the glyphosate and the main metabolite aminomethyl phosphonic acid thereof.
As a specific embodiment of the above method of the present invention, wherein, in the step (1), the pretreatment comprises:
washing a filtering membrane by using ultrapure water, filtering a water sample by using the filtering membrane, discarding an initial part of the water sample, collecting filtrate, and storing the filtrate in an environment with the temperature of 4 +/-3 ℃, wherein the storage time is not more than one week.
As a specific embodiment of the above method according to the present invention, in step (1), if the water sample contains free chlorine, the pretreatment further comprises adding a chlorine removal reagent to the water sample. For water samples containing free chlorine, such as those after disinfection, the presence of free chlorine can oxidize glyphosate, thereby causing its loss. Therefore, a chlorine removal reagent needs to be added to the water sample to remove free chlorine in the water sample.
In a specific embodiment of the above method of the present invention, the dechlorination reagent is sodium thiosulfate.
As a specific embodiment of the above method of the present invention, 2 to 4mg of sodium thiosulfate is added per 100mL of the water sample.
As a specific embodiment of the above method of the present invention, in step (1), if the mass spectrum used does not have sufficient detection capability, the method further comprises pre-concentrating the filtered water sample to help achieve the expected quantitative performance limit. Wherein, the skilled person can routinely determine whether the mass spectrum used has sufficient detection capability. In some embodiments, the pre-concentration may be achieved by solid phase extraction.
As a specific embodiment of the above method of the present invention, wherein, in the step (1), the chelate cleavage and derivatization of the pretreated water sample specifically include:
10mL of the pretreated water sample was first added to a test tube with a lid, and then 50. mu.L of 1,2-13C with a concentration of 20. mu.g/L was added to the test tube215N-tracer glyphosate aqueous solution to be used as an internal standard for derivatization, adding 100 mu L of EDTA-Na2 aqueous solution with the concentration of 0.1mol/L into the test tube, shaking up and keeping for 10min, adding 0.5mL of sodium borate buffer solution with the concentration of 0.05mol/L into the test tube to adjust the pH value of the system to 9.0-9.5, adding 500 mu L of FMOC-Cl acetonitrile solution with the concentration of 0.5mg/mL into the test tube, performing water bath ultrasound at 50-55 ℃ for 50-60min or performing ultrasound in normal temperature water bath for 4-12h (overnight), adding 100 mu L of phosphoric acid with the mass concentration of 30% into the test tube to neutralize the solution in the test tube, and filtering the obtained solution to obtain a sample to be analyzed.
As a specific embodiment of the above-mentioned method of the present invention, wherein, in the step (1), said 1,2-13C2The 15N-tracer glyphosate aqueous solution and the EDTA-Na2 aqueous solution are both prepared by adopting ultrapure water.
As a specific embodiment of the above method of the present invention, in the step (2), the analysis and test conditions of the liquid chromatogram in the liquid chromatogram tandem mass spectrometer include: the column used was a reverse phase column, BIO C182.1X 150mm, 2.0 μm, mobile phase A: acetonitrile, mobile phase B: triethylamine buffer with volume fraction of 0.1%, pH adjusted to 9-9.5 with chromatographically pure acetic acid, flow: 0.3mL/min, column temperature: 40 ℃, injection volume: 50 mu L of the solution; the gradient elution conditions were: and (3) 0-1.5min, wherein the initial mobile phase contains 8% acetonitrile by volume fraction for 1.5-3min, the acetonitrile volume fraction is increased to 95% and is 3-4.5min, the acetonitrile volume fraction is kept at 95% and is 4.5-6min, the acetonitrile volume fraction is reduced to 8% and is 6-11min, the acetonitrile volume fraction is continuously stabilized at 8% and is 12min, and the process is stopped.
In a specific embodiment of the above method of the present invention, the mobile phase B is triethylamine acetate buffer with a pH of 9.5.
As a specific embodiment of the above method of the present invention, in the step (2), the analysis and test conditions of mass spectrum in the lc-ms include: the ion source is an electrospray ion source, and the monitoring mode is as follows: a multiple reaction monitoring mode; scanning mode: a positive ion scanning mode and a negative ion mode;
flow rate of atomizing gas: 3L/min; heating air flow: 10L/min; DL temperature: 250 ℃; temperature of the heating block: 400 ℃, dry air flow: 10L/min.
As a specific embodiment of the above method of the present invention, wherein, in the step (2), the glyphosate and its main metabolite aminomethyl phosphonic acid are characterized, comprising:
comparing the retention time of the target compound with the retention time of a corresponding standard substance, comparing the positive and negative ionization mode mass spectrometry result of the MRM parameter of the target compound with the result obtained by performing multi-reaction monitoring (MRM) positive and negative ionization mode mass spectrometry on the standard substance, and determining whether the standard substance exists in an analysis sample according to the characteristic ion pair and the retention time to realize the identification of the target compound;
the quantification of glyphosate and its major metabolite aminomethylphosphonic acid comprises:
and drawing a standard working curve according to the standard concentration and the peak surface, substituting the peak area of the target compound in the sample to be detected into the standard working curve, and calculating to obtain the corresponding concentration of the target compound, thereby realizing quantification.
As a specific embodiment of the above method of the present invention, wherein, in step (2), 332.1 and 110 are selected as quantitative ion pairs of aminomethyl phosphate-anions, and 332.6 and 135.9 are selected as qualitative ion pairs of aminomethyl phosphate-anions; 390.1 and 168.2, 402.9 and 180.2 are selected as quantitative ion pairs of glyphosate-negative ions, and 390.1 and 150.1, 402.9 and 205.9 are selected as qualitative ion pairs of glyphosate-negative ions; 334 and 179.1 were chosen as the quantitative ion pair for the aminomethyl phosphate-positive ion, and 334 and 112.1 were chosen as the qualitative ion pair for the aminomethyl phosphate-positive ion; 392 and 88 were selected as the quantitative ion pair of glyphosate-positive ions, and 392 and 214 were selected as the qualitative ion pair of glyphosate-positive ions.
As a specific embodiment of the above method of the present invention, the water body is a hard water body in which the total amount of calcium and magnesium is 2 to 3 mmol/L.
Wherein when the calcium and magnesium content in the water body is higher, EDTA-Na needs to be properly increased in the derivatization stage2The concentration of the aqueous solution. For certain types of water where the total amount of calcium and magnesium is less than 2-3mmol/L, it may also be necessary to acidify them prior to derivatization.
In some embodiments of the invention, the acidification is preferably carried out according to the following specific steps:
firstly, 350 mu L of hydrochloric acid is added into 5mL of pretreated water sample to adjust the pH value of the sample to 1, the mixed solution is fully shaken and then stands for 1min, and then potassium hydroxide solution is added to adjust the pH value of the sample to 6-7.
In some embodiments of the present invention, the acidification may be further performed according to the following specific steps:
firstly, 50 mu L of oxalic acid dihydrate is added into 5mL of pretreated water sample to adjust the pH value of the sample to 2-3, the mixed solution is fully shaken and then stands for 1h, and then the potassium hydroxide solution is used for neutralizing the sample until the pH value is 7-8.
In one embodiment of the above method of the present invention, the water body comprises drinking water, ground water or surface water.
In the process of the present invention, glyphosate and AMPA (soluble fraction after filtration) are derivatized with 9-fluorenylmethylcarbamate (FMOC-Cl), which can reduce their polarity and increase their retention time in a reverse phase column (e.g., C18) separation; and analyzing the sample to be analyzed after derivatization by adopting high performance liquid chromatography-tandem mass spectrometry and matrix matching correction, wherein a double ionization scanning mode, namely a positive/negative ion simultaneous scanning mode is adopted for qualitative analysis in the analysis process, mutual evidence can be obtained, and matrix interference is eliminated, so that the accuracy of qualitative analysis and quantitative analysis can be enhanced.
The concentration measuring range of the method provided by the invention is 0.03 mu g/L-1.5 mu g/L, namely the detection limit of the method can reach 0.03 mu g/L.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed to be used in the description of the embodiments will be briefly introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and it is obvious for those skilled in the art that other drawings can be obtained according to these drawings without creative efforts.
FIG. 1 shows a mass spectrum obtained in example 1 of the present invention.
FIG. 2 is a mass spectrum obtained in example 2 of the present invention.
FIG. 3 is a mass spectrum obtained in example 3 of the present invention.
FIG. 4 is a mass spectrum obtained in example 4 of the present invention.
FIG. 5 is a mass spectrum obtained in example 5 of the present invention.
FIG. 6 is a mass spectrum obtained from a blank control according to the present invention.
FIG. 7 is a chromatogram of the separation of the target compound obtained in the calibration experiment of the present invention.
FIG. 8 is a graph showing the operation of the aminomethyl phosphoric acid anion scan mode in the calibration experiment according to the present invention.
FIG. 9 is a graph showing the operation of glyphosate anion scan in the calibration experiment of the present invention.
FIG. 10 is a graph showing the operation of the scanning mode of aminomethylphosphonic acid positive ion in the calibration experiment of the present invention.
FIG. 11 is a graph showing the operation of glyphosate positive ion scan mode in the calibration experiment of the present invention.
FIG. 12 shows the results of the calibration experiment of the present invention, 1,2-13C2，15N-tracing glyphosate anion scanning working curve.
FIG. 13 shows the results of the calibration experiment of the present invention, 1,2-13C2，15N-tracing glyphosate positive ion scanning working curve.
Detailed Description
The "ranges" disclosed herein are given as lower and upper limits. There may be one or more lower limits, and one or more upper limits, respectively. The given range is defined by the selection of a lower limit and an upper limit. The selected lower and upper limits define the boundaries of the particular range. All ranges defined in this manner are combinable, i.e., any lower limit can be combined with any upper limit to form a range. For example, ranges of 60-120 and 80-110 are listed for particular parameters, with the understanding that ranges of 60-110 and 80-120 are also contemplated. Further, if the minimum range values listed are 1 and 2 and the maximum range values listed are 3, 4, and 5, then the following ranges are all contemplated: 1-3, 1-4, 1-5, 2-3, 2-4 and 2-5.
In the present invention, unless otherwise stated, the numerical range "a-b" represents a shorthand representation of any combination of real numbers between a and b, where a and b are both real numbers. For example, a numerical range of "0 to 5" indicates that all real numbers between "0 to 5" have been listed throughout this disclosure, and "0 to 5" is only a shorthand representation of the combination of these numbers.
In the present invention, all the embodiments and preferred embodiments mentioned in the present invention may be combined with each other to form a new technical solution, if not specifically stated.
In the present invention, all the technical features mentioned in the present invention and preferred features may be combined with each other to form a new technical solution, if not specifically stated.
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings, the attached tables and the embodiments. The following described embodiments are some, but not all embodiments of the present invention, and are merely illustrative of the present invention and should not be construed as limiting the scope of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.
The reagents and instruments used in the examples of the invention are as follows:
all reagents and solvents should be of sufficient purity to be useful for trace analysis, unless otherwise specified.
Reagent
1 deionized water.
2 ultrapure water, according to ISO 3696 grade requirements.
3 nitrogen, N2The purity is more than or equal to 99.996 percent (volume fraction).
4 laboratory cleaner, alkaline cleaner.
6 acetonitrile, C2H3And N, chromatographic grade.
7 methanol, CH4O, chromatographic grade.
8 ethanol, C2H6O, 95%, chromatographic grade and mass fraction.
9 ethyl acetate, C4H8O2。
10 ammonium acetate, C2H7O2N。
11 Triethylamine, C6H15N。
12 ammonia, NH4OH, 28% mass fraction.
13 formic acid, CH2O2。
14 hydrochloric acid, HCl, concentration 300 g/L.
15 glacial acetic acid, C2H4O2。
16 disodium ethylenediaminetetraacetic acid (EDTA) dihydrate, C10H14N2O8Na2·2H2O (marked as EDTA-Na2) with the minimum purity of 99 percent by mass.
179-fluorenylmethyl chloroformate, (FMOC-Cl), C15H11ClO2The minimum purity is 97 percent by mass.
FMOC-Cl was used to prepare the derivatizing reagent, FMOC-Cl acetonitrile (6) solution, 50 mg/mL. The solution was stored at-18 ℃. + -. 3 ℃ for 6 months.
The derivatization was carried out using FMOC-Cl in acetonitrile, 0.5 mg/mL.
18 reference material
Glyphosate, N- (phosphonomethyl) glycine, C3H8NO5P, purity>98 percent, mass fraction, molecular weight of 169.1g/mol, and chemical Abstract service number, namely CAS-RN, of 1071-83-6.
AMPA, aminomethylphosphoric acid, CH6NO3P, purity>98 percent, mass fraction, molecular weight of 111.0g/mol, and chemical abstract service number, namely CAS-RN, of 1066-51-9.
1,2-13C215N-tracing glyphosate, and the mass concentration of the internal standard stock solution is 100 mg/L.
19 calibration solution
100mg/L of glyphosate stock solution and 100mg/L of AMPA stock solution are respectively prepared by ultrapure water. These solutions were stored at 4 ℃. + -. 3 ℃ for 1 month.
Preparation of 1,2-13C2，15The N-trace glyphosate aqueous solution is 100 mg/L. The solution was stored at 4 ℃. + -. 3 ℃ for 1 month.
Mixed solution of substitutes: preparation of 1,2-13C with ultrapure Water215N-trace glyphosate aqueous solution, 20 mug/L. The solution was stored at 4 ℃. + -. 3 ℃ for 1 month.
Note that: if there is sufficient assurance in terms of stability, the stock and calibration solutions can be stored for longer periods of time.
Sodium tetraborate 21 decahydrate, Na2B4O7·10H2O。
22 sodium tetraborate buffer solution, 0.05 mol/L; pH 9.2.
It is prepared by dissolving 19 + -0.1 g sodium tetraborate decahydrate (21) in 1L water (1). This solution can be stored at 4 ℃. + -. 3 ℃ for about 1 month.
23 mineral water containing divalent cation (Mg) less than 3.2mmol/L2+And Ca2+Total amount) for preparing a matrix matching calibration.
Second, instrument
The material or any parts that may come into contact with the sample should be free of any residue that may cause unacceptable interference with the blank.
Glass and plastic containers can be used for all steps prior to sampling and derivatization. Glass vials and glass test tubes were used after derivatization.
1 glassware or apparatus commonly used in laboratories, particularly the following.
2 glass, Polyethylene (PE) or polypropylene (PP) bottles, at least 50mL, for sampling.
3 glass, Polyethylene (PE) or polypropylene (PP) syringe, 50mL, for sample filtration.
4 Disposable syringe for filtration, diameter 25mm, hydrophilic membrane, 0.45 μm, for example from regenerated cellulose.
5 glass, or disposable PE or PP conical tube, about 50mL, was used for derivatization.
6 micropipettes, adjustable from 100. mu.L to 500. mu.L.
And 7, a pH meter.
8 glass bottles, suitable for autosampler, with Polytetrafluoroethylene (PTFE) or silica gel spacers.
9 glass test tube, 15mL or less.
10 liquid chromatography mass spectrometer, LCMS-8050.
11 reversed phase column, BIO C182.1X 150mm, 2.0 μm.
Example 1
The embodiment provides a method for measuring glyphosate in underground water (wherein the underground water is underground water collected from a place of a Shijiazhu and recorded as MS-1) by using a double ionization scanning mode-liquid chromatography-tandem mass spectrometry, and the method comprises the following specific steps of:
1) sampling:
if the sample vial is not disposable, the sample vial is first rinsed with deionized water, then with a laboratory cleaner, then with water, then with ultra-pure water, and finally with 95% ethanol.
Samples were taken according to ISO 5667-3 (about 50 mL).
If the sample is suspected of containing free chlorine after sampling, about 2mg of sodium thiosulfate or any other dechlorination reagent can be added to each 100mL of sample to remove the free chlorine.
2) Pretreatment (suspended particulate matter):
the filter membrane was placed on a syringe and rinsed with 5mL of ultra pure water.
The sample (approximately 50mL) was filtered, the initial 5mL discarded, and the filtrate collected in a conical tube.
The filtered sample was stored in a temperature environment of 4 ℃ ± 3 ℃ for up to one week, followed by the derivatization step.
3) Chelate cleavage and derivatization:
firstly, 10mL of water sample (different amounts of samples need to be proportionally adjusted by the amount of the following reagents) after pretreatment is added into a 15mL test tube with a cover, and then 50 muL of 1,2-13C with the concentration of 20 mug/L is added into the test tube215N-tracing glyphosate aqueous solution (prepared by ultrapure water) to be used as an internal standard for derivatization, adding 100 mu L of EDTA-Na2 aqueous solution (prepared by ultrapure water) with the concentration of 0.1mol/L into the test tube, shaking uniformly and keeping for 10min, adding 0.5mL of sodium borate buffer solution with the concentration of 0.05mol/L into the test tube to adjust the pH value of the system to 9.0-9.5, adding 500 mu L of FMOC-Cl acetonitrile solution with the concentration of 0.5mg/mL into the test tube, carrying out water bath ultrasound for 60min, adding 30% phosphoric acid into the test tube to neutralize the solution in the test tube, and filtering the obtained solution to obtain a sample to be analyzed.
If a mass spectrometer with enough detection capability is used in the subsequent analysis process, preconcentration is not needed, and the derivatized sample is directly injected into an instrument for analysis;
otherwise, the pre-concentration step may be performed on the derivatized sample, specifically including: the SPE tubes were rinsed with 1mL of 0.1% by volume aqueous formic acid followed by 2X 500. mu.L of ultra-pure water. The column was dried under vacuum or a gentle stream of nitrogen for about 1min to remove most of the water. Preparing an eluent: methanol/ammonia (ultrapure water, 20g/L) volume fraction of 70:30, and 3X 700. mu.L of the prepared eluent was taken to elute the SPE tube. The eluate was collected in a glass test tube and purged with nitrogen until the methanol was removed. The final volume obtained after concentration was about 0.7 mL. Note that: methanol remaining under chromatographic conditions causes degradation, resulting in peak splitting. The resulting 0.7mL of concentrate was adjusted to 1mL with ultrapure water. The purified extract, due to the stability of the FMOC derivative in water prior to analysis, can be stored at 4 ℃ ± 3 ℃ for 48h, a step which can help to reach the expected limit of quantitative performance (LOQ).
4) Liquid quality analysis:
the model of the liquid chromatography-mass spectrometer used in this embodiment is LCMS-8050, and in the test process, the apparatus is used according to the instructions provided by the manufacturer, and the system stability needs to be checked periodically, and the apparatus parameter settings are adjusted and optimized according to the instructions of the manufacturer.
Since a buffered mobile phase is used in this example, it is recommended that the analytical column and the chromatographic system should be cleaned periodically according to the manufacturer's instructions.
Wherein the liquid chromatography conditions are:
and (3) separating glyphosate and the derivative AMPA by adopting a reverse phase column and proper chromatographic conditions. The pH of the mobile phase should be basic (pH should be between 9 and 9.5) in order to obtain better chromatographic performance (peak symmetry close to 1 and narrow peak), which means that a specific column (mixed stationary phase) is used.
Chromatographic conditions are as follows:
the column used was a reverse phase column, BIO C182.1X 150mm, 2.0 μm, mobile phase A: acetonitrile, mobile phase B: triethylamine buffer with volume fraction of 0.1%, pH adjusted to 9.5 with chromatographically pure acetic acid, flow: 0.3mL/min, column temperature: 40 ℃, injection volume: 50 mu L of the solution; the gradient elution conditions were: and (3) 0-1.5min, wherein the initial mobile phase contains 8% acetonitrile by volume fraction for 1.5-3min, the acetonitrile volume fraction is increased to 95% and is 3-4.5min, the acetonitrile volume fraction is kept at 95% and is 4.5-6min, the acetonitrile volume fraction is reduced to 8% and is 6-11min, the acetonitrile volume fraction is continuously stabilized at 8% and is 12min, and the process is stopped.
The mass spectrum conditions are as follows:
analyzing by using a liquid chromatography-mass spectrometer with the model number of LC-MS 8050;
the ion source is an electrospray ion source, and the monitoring mode is as follows: a multiple reaction monitoring mode; scanning mode: a positive ion scanning mode and a negative ion mode;
flow rate of atomizing gas: 3L/min; heating air flow: 10L/min; DL temperature: 250 ℃; temperature of the heating block: 400 ℃, dry air flow: 10L/min.
Wherein the identification and confirmation of the analyte comprises:
comparing the retention time of the target compound with the retention time of a corresponding standard substance, comparing the positive and negative ionization mode mass spectrometry result of the MRM parameter of the target compound with the result obtained by carrying out multi-reaction monitoring positive and negative ionization mode mass spectrometry on the standard substance, and determining whether the standard substance exists in an analysis sample according to the characteristic ion pair and the retention time to realize the identification of the target compound;
the MRM parameters are shown in table 1 below.
TABLE 1 MRM reference parameter Table
The ratio between the quantitative and the qualitative ions depends on the MRM conditions (i.e. the collision energy and the gas pressure in the collision cell). These ratios should be determined using standard materials and using the same equipment optimization, check whether the sample is within ± 20% of the observed value for the reference material.
The data of the experimental results obtained in this example are shown in table 2 below and fig. 1.
TABLE 2
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 4.015 | 29691 | 2.480 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.977 | 350133 | 1.972 | μg/ | Target | |
| 3 | Glyphosate- |
3.978 | 354397 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.979 | 258986 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 4.044 | 490584 | 2.240 | μg/ | Target | |
| 6 | Glyphosate-positive ion | 3.978 | 243709 | 2.032 | μg/L | Target |
Example 2
The embodiment provides a method for measuring glyphosate in underground water (wherein the underground water is collected from a place of a Shijiazhu and is recorded as MS-2) by using a double ionization scanning mode-liquid chromatography-tandem mass spectrometry, and the flow, the used substances, the used process parameters and the like of the method are the same as those in the embodiment 1.
The data of the experimental results obtained in this example are shown in table 3 and fig. 2 below.
TABLE 3
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 4.013 | 28781 | 2.461 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.977 | 347561 | 2.004 | μg/ | Target | |
| 3 | Glyphosate- |
3.978 | 346182 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.979 | 253440 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 4.047 | 431970 | 2.016 | μg/ | Target | |
| 6 | Glyphosate-positive ion | 3.979 | 231918 | 1.976 | μg/L | Target |
Example 3
The embodiment provides a method for measuring glyphosate in underground water (wherein the underground water is collected from a place of a Shijiazhu and is recorded as MS-3) by using a double ionization scanning mode-liquid chromatography-tandem mass spectrometry, and the flow, the used substances, the used process parameters and the like of the method are the same as those in the embodiment 1.
The data of the experimental results obtained in this example are shown in table 4 and fig. 3 below.
TABLE 4
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 4.011 | 26571 | 2.171 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.964 | 326350 | 1.798 | μg/ | Target | |
| 3 | Glyphosate- |
3.964 | 362308 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.963 | 271643 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 4.028 | 406301 | 1.769 | μg/ | Target | |
| 6 | Glyphosate-positive ions | 3.963 | 235495 | 1.872 | μg/L | Target |
Example 4
The embodiment provides a method for preparing glyphosate in a quality control sample (containing glyphosate-13C 2 and 15N with the mass concentration of 2 mug/L and marked as CC-1, namely CC-1 contains glyphosate and glyphosate internal standard substance with fixed concentration) with the concentration of 5 mug/L by adding glyphosate stock solution into ultrapure water through double ionization scanning mode-liquid chromatography-tandem mass spectrometry, wherein the flow, the used substances, the used process parameters and the like of the method are the same as those in the embodiment 1.
The data of the experimental results obtained in this example are shown in table 5 below and fig. 4.
TABLE 5
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 4.014 | 75969 | 5.238 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.981 | 986362 | 4.586 | μg/ | Target | |
| 3 | Glyphosate- |
3.983 | 429369 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.983 | 285450 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 4.052 | 1195868 | 4.954 | μg/ | Target | |
| 6 | Glyphosate-positive ion | 3.983 | 638220 | 4.827 | μg/L | Target |
Example 5
This example provides a method for measuring glyphosate in ultrapure water (containing glyphosate-13C 2,15N at a mass concentration of 2 μ g/L) by using a double ionization scanning mode-liquid chromatography tandem mass spectrometry, wherein the procedures, materials and process parameters are the same as those in example 1.
The data of the experimental results obtained in this example are shown in table 6 and fig. 5 below.
TABLE 6
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 3.970 | 1639 | 0.125 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.956 | 4347 | 0.022 | μg/ | Target | |
| 3 | Glyphosate- |
3.956 | 387546 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.958 | 263681 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 4.025 | 17594 | 0.079 | μg/ | Target | |
| 6 | Glyphosate-positive ion | 3.954 | 1958 | 0.016 | μg/L | Target |
Blank control example
The detection object used in the blank control example is only added with 1,2-13C2，15Mineral water/mineral water (as shown above 23) of N-tracer glyphosate (as shown above in 18).
The blank control sample and the water sample to be tested should be processed and analyzed simultaneously, and the flow, the used substances, the used process parameters and the like of the analysis are the same as those in the example 1. Both glyphosate and AMPA residual levels should be below 1/3 of limit of quantitation (LOQs). The data of the experimental results obtained in this blank control example are shown in table 7 below and fig. 6.
TABLE 7
| ID# | Name | Ret.Time | Area | Conc. | | Type | |
| 1 | Aminomethylphosphonic acid anion | 3.921 | 390 | 0.020 | μg/ | Target | |
| 2 | Glyphosate-anion | 3.889 | 2527 | 0.007 | μg/ | Target | |
| 3 | Glyphosate- |
3.891 | 633369 | 2.000 | μg/ | I.STD | |
| 4 | Glyphosate- |
3.889 | 357129 | 2.000 | μg/ | I.STD | |
| 5 | Aminomethylphosphonic acid-anions | 3.944 | 4784 | 0.015 | μg/ | Target | |
| 6 | Glyphosate-positive ion | 3.888 | 1292 | 0.008 | μg/L | Target |
Combining the data in tables 2-5 above, excluding data from blank and blank control samples (both not detected), comparing data from aminomethylphosphonic acid and glyphosate in negative ion mode and positive ion mode, and performing one-way analysis of variance (α ═ 0.05) on aminomethylphosphonic acid and glyphosate, respectively, the results are shown in tables 8-10 below.
TABLE 8 data of significant difference analysis of aminomethylphosphonic acid and glyphosate
Table 9 results data for one-way anova (α ═ 0.05)
TABLE 10 Glyphosate one-way ANOVA (α ═ 0.05) results data
As can be seen from tables 8-10, there is no significant difference in the positive and negative ion mode data for aminomethylphosphonic acid and glyphosate, which can be considered to be identical. Therefore, in actual sampling, the data of the negative ion mode and the positive ion mode can be comprehensively analyzed and averaged to seek the accuracy of the data, and the effect of enhancing the qualitative and quantitative effects of the double ionization mode analysis is achieved.
Calibration experiment
Matrix matching calibration was performed according to ISO8466-1 for glyphosate and AMPA (both at 0.05. mu.g/L, 0.2. mu.g/L, 0.5. mu.g/L, 2.0. mu.g/L, 5.0. mu.g/L, 10.0. mu.g/L and 20.0. mu.g/L), and 1,2-13C2, 15N-labeled glyphosate (constant concentration of 2.00. mu.g/L), wherein the matrix matching calibration was performed using a 7-point calibration using the same liquid chromatography and mass spectrometry conditions as in example 1.
The target compound separation chromatogram obtained in the calibration experiment is shown in FIG. 7, the working curve of aminomethyl phosphoric acid negative ion scanning mode, the working curve of glyphosate negative ion scanning mode, the working curve of aminomethyl phosphonic acid positive ion scanning mode, the working curve of glyphosate positive ion scanning mode, 1,2-13C2，15Working curve of N-tracing glyphosate anion (internal standard, fixed concentration, all 2 mug/L) scanning and 1,2-13C2，15The working curves of N-trace glyphosate positive ion (internal standard, fixed concentration, all 2 mug/L) scans are respectively shown in figures 8-138-13, it can be seen that the glyphosate, aminomethylphosphonic acid positive and negative ion scan pattern exhibits a good linear relationship over the concentration range of 0.05-20.0. mu.g/L.
The above description is only exemplary of the invention and should not be taken as limiting the scope of the invention, so that the invention is intended to cover all modifications and equivalents of the embodiments described herein. In addition, the technical features and the technical inventions of the present invention, the technical features and the technical inventions, and the technical inventions can be freely combined and used.
Claims (10)
1. A method for measuring glyphosate in water by double ionization scanning mode-liquid chromatography-tandem mass spectrometry is characterized by comprising the following steps:
(1) chelating and breaking and derivatizing the pretreated water sample
Firstly, adding 1,2-13C into the pretreated water sample215N-tracing glyphosate aqueous solution to be used as an internal standard for derivatization, adding EDTA-Na2 aqueous solution, shaking up and keeping for a period of time, adding sodium borate buffer solution to adjust the pH value of the system, adding FMOC-Cl acetonitrile solution, performing water bath ultrasound, adding phosphoric acid to neutralize the solution, and finally filtering the obtained solution to obtain a sample to be analyzed;
(2) and analyzing and testing the sample to be analyzed by adopting a liquid chromatography-tandem mass spectrometer in a double-ionization scanning mode so as to quantify and qualitatively identify the glyphosate and the main metabolite aminomethyl phosphonic acid thereof.
2. The method according to claim 1, wherein in step (1), the pre-processing comprises:
washing a filtering membrane by using ultrapure water, filtering a water sample by using the filtering membrane, discarding an initial part of the water sample, collecting filtrate, and storing the filtrate in an environment with the temperature of 4 +/-3 ℃, wherein the storage time is not more than one week.
3. The method according to claim 2, wherein in step (1), if the water sample contains free chlorine, the pre-treating further comprises adding a chlorine removal reagent to the water sample;
preferably, the dechlorination reagent is sodium thiosulfate, and more preferably, 2-4mg of sodium thiosulfate is added to every 100mL of water sample.
4. The method of claim 1, wherein in step (1), if the mass spectrum used does not have sufficient detection capability, the method further comprises pre-concentrating the filtered water sample to help achieve the desired quantitative performance limit.
5. The method according to claim 1, wherein in the step (1), the chelate cleavage and derivatization are performed on the pretreated water sample, and specifically comprise:
10mL of the pretreated water sample was first added to a test tube with a lid, and then 50. mu.L of 1,2-13C with a concentration of 20. mu.g/L was added to the test tube215N-tracer glyphosate aqueous solution to be used as an internal standard for derivatization, adding 100 mu L of EDTA-Na2 aqueous solution with the concentration of 0.1mol/L into the test tube, shaking up and keeping for 10min, adding 0.5mL of sodium borate buffer solution with the concentration of 0.05mol/L into the test tube to adjust the pH value of the system to 9.0-9.5, adding 500 mu L of FMOC-Cl acetonitrile solution with the concentration of 0.5mg/mL into the test tube, performing water bath ultrasound at 50-55 ℃ for 50-60min or performing ultrasound in normal temperature water bath for 4-12h (overnight), adding 100 mu L of phosphoric acid with the mass concentration of 30% into the test tube to neutralize the solution in the test tube, and filtering the obtained solution to obtain a sample to be analyzed.
6. The method according to claim 1, wherein in the step (2), the analysis test conditions of the liquid chromatogram in the liquid chromatogram tandem mass spectrometer comprise: the column used was a reverse phase column, BIO C182.1X 150mm, 2.0 μm, mobile phase A: acetonitrile, mobile phase B: triethylamine buffer with volume fraction of 0.1%, pH adjusted to 9-9.5 with chromatographically pure acetic acid, flow: 0.3mL/min, column temperature: 40 ℃, injection volume: 50 mu L of the solution; the gradient elution conditions were: 0-1.5min, the initial mobile phase contains 8% acetonitrile by volume fraction, 1.5-3min, the acetonitrile volume fraction is increased to 95%, 3-4.5min, the acetonitrile volume fraction is kept at 95%, 4.5-6min, the acetonitrile volume fraction is decreased to 8%, 6-11min, the acetonitrile volume fraction is continuously stabilized at 8%, 12min, and the process is stopped;
preferably, the mobile phase B is triethylamine acetate buffer solution with the pH value of 9.5.
7. The method of claim 1, wherein in step (2), the analytical test conditions of mass spectrometry in the LC-MS comprise: the ion source is an electrospray ion source, and the monitoring mode is as follows: a multiple reaction monitoring mode; scanning mode: a positive ion scanning mode and a negative ion mode;
flow rate of atomizing gas: 3L/min; heating air flow: 10L/min; DL temperature: 250 ℃; temperature of the heating block: 400 ℃, dry air flow: 10L/min.
8. The method of claim 1, wherein in step (2), the characterization of glyphosate and its major metabolite aminomethylphosphonic acid comprises:
comparing the retention time of the target compound with the retention time of a corresponding standard substance, comparing the positive and negative ionization mode mass spectrometry result of the MRM parameter of the target compound with the result obtained by carrying out multi-reaction monitoring positive and negative ionization mode mass spectrometry on the standard substance, and determining whether the standard substance exists in an analysis sample according to the characteristic ion pair and the retention time to realize the identification of the target compound;
the quantification of glyphosate and its major metabolite aminomethylphosphonic acid comprises:
and drawing a standard working curve according to the standard concentration and the peak surface, substituting the peak area of the target compound in the sample to be detected into the standard working curve, and calculating to obtain the corresponding concentration of the target compound, thereby realizing quantification.
9. The method according to claim 8, wherein in step (2), 332.1 and 110 are selected as quantitative ion pairs of aminomethyl phosphate-anions, and 332.6 and 135.9 are selected as qualitative ion pairs of aminomethyl phosphate-anions; 390.1 and 168.2, 402.9 and 180.2 are selected as quantitative ion pairs of glyphosate-negative ions, and 390.1 and 150.1, 402.9 and 205.9 are selected as qualitative ion pairs of glyphosate-negative ions; 334 and 179.1 were chosen as the quantitative ion pair for the aminomethyl phosphate-positive ion, and 334 and 112.1 were chosen as the qualitative ion pair for the aminomethyl phosphate-positive ion; 392 and 88 were selected as the quantitative ion pair of glyphosate-positive ions, and 392 and 214 were selected as the qualitative ion pair of glyphosate-positive ions.
10. The method according to any one of claims 1 to 9, wherein the water body is a hard water body with a total amount of calcium and magnesium of 2 to 3 mmol/L;
preferably, the body of water comprises drinking water, ground water or surface water.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111489161.8A CN114236032A (en) | 2021-12-07 | 2021-12-07 | Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202111489161.8A CN114236032A (en) | 2021-12-07 | 2021-12-07 | Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN114236032A true CN114236032A (en) | 2022-03-25 |
Family
ID=80753849
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202111489161.8A Pending CN114236032A (en) | 2021-12-07 | 2021-12-07 | Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114236032A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114910594A (en) * | 2022-05-31 | 2022-08-16 | 湖北省地质实验测试中心(国土资源部武汉矿产资源监督检测中心) | Method for rapidly measuring glyphosate by synchronously extracting and deriving |
| CN115406981A (en) * | 2022-08-04 | 2022-11-29 | 青岛理工大学 | Detection method based on PPCPs mass balance accounting in MBBR sewage treatment system |
-
2021
- 2021-12-07 CN CN202111489161.8A patent/CN114236032A/en active Pending
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114910594A (en) * | 2022-05-31 | 2022-08-16 | 湖北省地质实验测试中心(国土资源部武汉矿产资源监督检测中心) | Method for rapidly measuring glyphosate by synchronously extracting and deriving |
| CN114910594B (en) * | 2022-05-31 | 2024-05-14 | 湖北省地质实验测试中心(国土资源部武汉矿产资源监督检测中心) | Method for rapidly determining glyphosate by synchronously extracting and deriving |
| CN115406981A (en) * | 2022-08-04 | 2022-11-29 | 青岛理工大学 | Detection method based on PPCPs mass balance accounting in MBBR sewage treatment system |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN114236032A (en) | Method for measuring glyphosate in water body by double ionization scanning mode-liquid chromatography-tandem mass spectrometry | |
| CN114270165A (en) | Triggered sampling system and method | |
| CN114894913B (en) | Method for synchronously and rapidly determining total arsenic and arsenic metabolite content in urine | |
| EP3875944A1 (en) | Atomic fluorescence spectrometry method and device employing water as carrier fluid | |
| CN112326812A (en) | Method for simultaneously detecting five pesticides in underground water by isotope dilution-ONLINESPE-HRMS | |
| CN112858503A (en) | Method for measuring residual quantity of 10 aminoglycoside antibiotics in eggs | |
| JP2009139177A (en) | Aminoglycoside antibiotic analyzing method | |
| CN111220722B (en) | Method for simultaneously determining 8 p-hydroxybenzoate compounds in soil | |
| CN109900777B (en) | Device for rapidly analyzing gas components of combustion products of materials on line and application | |
| CN113820421A (en) | Method for measuring 6 phenolic compounds in water | |
| CN111474279B (en) | Method and kit for detecting macrolide antibiotic compounds | |
| CN115326994A (en) | Method and system for simultaneously analyzing multi-class smoke exposure biomarkers and using method | |
| Xu et al. | Simultaneous measurement of multiple organic tracers in fine aerosols from biomass burning and fungal spores by HPLC-MS/MS | |
| Chen et al. | Performance Liquid Chromatography/Tandem Mass Spectrometry | |
| CN114544281A (en) | Method for measuring cesium and compounds thereof in air of workplace | |
| CN113391014A (en) | Method for measuring 18 polycyclic aromatic hydrocarbons in air filter membrane | |
| Mukhtar et al. | On-line determination of water-soluble zinc in airborne particulate matter using a dynamic extraction procedure coupled to flame atomic absorption spectrometry | |
| CN110749691A (en) | HPLC-MS/MS method for determining aflatoxin and homologue thereof in infant auxiliary food | |
| CN111208231B (en) | Method for simultaneously detecting six illegally added compounds in plant beverage | |
| CN117649897B (en) | Method for predicting gas production of soy sauce in shelf life | |
| CN220893913U (en) | Sample injection atomization treatment device and system thereof | |
| CN112255340B (en) | Method for determining PPO (polyphenylene oxide) and PPA (polyphthalamide) in glufosinate ammonium product by liquid chromatography-mass spectrometry | |
| CN115343394B (en) | Method for determining two tridecyl silanes in spray product and application thereof | |
| US20030194816A1 (en) | Method of collecting chemically contaminating impurity constituents contained in air | |
| CN114720570B (en) | Method for detecting 8 estrogens in fish meat |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |