CN115032292A - Detection and analysis method for organic phosphate in water sample - Google Patents
Detection and analysis method for organic phosphate in water sample Download PDFInfo
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- CN115032292A CN115032292A CN202210497084.9A CN202210497084A CN115032292A CN 115032292 A CN115032292 A CN 115032292A CN 202210497084 A CN202210497084 A CN 202210497084A CN 115032292 A CN115032292 A CN 115032292A
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- 229910019142 PO4 Inorganic materials 0.000 title claims abstract description 65
- 239000010452 phosphate Substances 0.000 title claims abstract description 46
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 title claims abstract description 44
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 40
- 238000001514 detection method Methods 0.000 title claims description 13
- 238000004458 analytical method Methods 0.000 title description 7
- 239000012086 standard solution Substances 0.000 claims abstract description 32
- 238000000034 method Methods 0.000 claims abstract description 31
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 30
- 239000000126 substance Substances 0.000 claims abstract description 20
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims abstract description 12
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims abstract description 12
- 239000012074 organic phase Substances 0.000 claims abstract description 10
- 238000001914 filtration Methods 0.000 claims abstract description 7
- 238000010813 internal standard method Methods 0.000 claims abstract description 6
- 229910052757 nitrogen Inorganic materials 0.000 claims abstract description 6
- 239000012071 phase Substances 0.000 claims abstract description 6
- 238000004445 quantitative analysis Methods 0.000 claims abstract description 6
- 238000007865 diluting Methods 0.000 claims abstract description 4
- 238000003260 vortexing Methods 0.000 claims abstract description 4
- 238000007664 blowing Methods 0.000 claims abstract description 3
- 235000021317 phosphate Nutrition 0.000 claims description 60
- 238000011084 recovery Methods 0.000 claims description 14
- 150000002500 ions Chemical class 0.000 claims description 12
- 150000003013 phosphoric acid derivatives Chemical class 0.000 claims description 12
- 238000002156 mixing Methods 0.000 claims description 10
- 238000000589 high-performance liquid chromatography-mass spectrometry Methods 0.000 claims description 9
- 239000013535 sea water Substances 0.000 claims description 9
- GTVWRXDRKAHEAD-UHFFFAOYSA-N Tris(2-ethylhexyl) phosphate Chemical compound CCCCC(CC)COP(=O)(OCC(CC)CCCC)OCC(CC)CCCC GTVWRXDRKAHEAD-UHFFFAOYSA-N 0.000 claims description 8
- 239000008399 tap water Substances 0.000 claims description 8
- 235000020679 tap water Nutrition 0.000 claims description 8
- 238000012360 testing method Methods 0.000 claims description 8
- STCOOQWBFONSKY-UHFFFAOYSA-N tributyl phosphate Chemical compound CCCCOP(=O)(OCCCC)OCCCC STCOOQWBFONSKY-UHFFFAOYSA-N 0.000 claims description 8
- XZZNDPSIHUTMOC-UHFFFAOYSA-N triphenyl phosphate Chemical compound C=1C=CC=CC=1OP(OC=1C=CC=CC=1)(=O)OC1=CC=CC=C1 XZZNDPSIHUTMOC-UHFFFAOYSA-N 0.000 claims description 8
- 238000002347 injection Methods 0.000 claims description 7
- 239000007924 injection Substances 0.000 claims description 7
- YSMRWXYRXBRSND-UHFFFAOYSA-N TOTP Chemical compound CC1=CC=CC=C1OP(=O)(OC=1C(=CC=CC=1)C)OC1=CC=CC=C1C YSMRWXYRXBRSND-UHFFFAOYSA-N 0.000 claims description 6
- 125000004965 chloroalkyl group Chemical group 0.000 claims description 6
- DQWPFSLDHJDLRL-UHFFFAOYSA-N triethyl phosphate Chemical compound CCOP(=O)(OCC)OCC DQWPFSLDHJDLRL-UHFFFAOYSA-N 0.000 claims description 6
- QJAVUVZBMMXBRO-UHFFFAOYSA-N tripentyl phosphate Chemical compound CCCCCOP(=O)(OCCCCC)OCCCCC QJAVUVZBMMXBRO-UHFFFAOYSA-N 0.000 claims description 6
- ASLWPAWFJZFCKF-UHFFFAOYSA-N tris(1,3-dichloropropan-2-yl) phosphate Chemical compound ClCC(CCl)OP(=O)(OC(CCl)CCl)OC(CCl)CCl ASLWPAWFJZFCKF-UHFFFAOYSA-N 0.000 claims description 6
- BGNTUSKZDOUZCZ-UHFFFAOYSA-N tris(1-butoxyethyl) phosphate Chemical compound CCCCOC(C)OP(=O)(OC(C)OCCCC)OC(C)OCCCC BGNTUSKZDOUZCZ-UHFFFAOYSA-N 0.000 claims description 6
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 6
- 239000012498 ultrapure water Substances 0.000 claims description 6
- 125000000217 alkyl group Chemical group 0.000 claims description 5
- 125000003118 aryl group Chemical group 0.000 claims description 5
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 5
- 239000012528 membrane Substances 0.000 claims description 5
- 239000000203 mixture Substances 0.000 claims description 4
- 238000002360 preparation method Methods 0.000 claims description 4
- 238000004885 tandem mass spectrometry Methods 0.000 claims description 4
- HQUQLFOMPYWACS-UHFFFAOYSA-N tris(2-chloroethyl) phosphate Chemical compound ClCCOP(=O)(OCCCl)OCCCl HQUQLFOMPYWACS-UHFFFAOYSA-N 0.000 claims description 4
- CGSLYBDCEGBZCG-UHFFFAOYSA-N Octicizer Chemical compound C=1C=CC=CC=1OP(=O)(OCC(CC)CCCC)OC1=CC=CC=C1 CGSLYBDCEGBZCG-UHFFFAOYSA-N 0.000 claims description 3
- 239000007983 Tris buffer Substances 0.000 claims description 3
- 239000008346 aqueous phase Substances 0.000 claims description 3
- 150000002148 esters Chemical class 0.000 claims description 3
- 239000011521 glass Substances 0.000 claims description 3
- 230000004044 response Effects 0.000 claims description 3
- RXPQRKFMDQNODS-UHFFFAOYSA-N tripropyl phosphate Chemical compound CCCOP(=O)(OCCC)OCCC RXPQRKFMDQNODS-UHFFFAOYSA-N 0.000 claims description 3
- GTRSAMFYSUBAGN-UHFFFAOYSA-N tris(2-chloropropyl) phosphate Chemical compound CC(Cl)COP(=O)(OCC(C)Cl)OCC(C)Cl GTRSAMFYSUBAGN-UHFFFAOYSA-N 0.000 claims description 3
- 238000001294 liquid chromatography-tandem mass spectrometry Methods 0.000 abstract 1
- 150000003014 phosphoric acid esters Chemical class 0.000 description 14
- -1 electronics Substances 0.000 description 10
- 230000007613 environmental effect Effects 0.000 description 4
- 238000000622 liquid--liquid extraction Methods 0.000 description 4
- 239000000178 monomer Substances 0.000 description 4
- 238000000638 solvent extraction Methods 0.000 description 4
- 239000007789 gas Substances 0.000 description 3
- 238000002372 labelling Methods 0.000 description 3
- 239000004033 plastic Substances 0.000 description 3
- 229920003023 plastic Polymers 0.000 description 3
- 230000000711 cancerogenic effect Effects 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000010828 elution Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000003063 flame retardant Substances 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- 239000004014 plasticizer Substances 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 125000001424 substituent group Chemical group 0.000 description 2
- 239000004753 textile Substances 0.000 description 2
- WVLBCYQITXONBZ-UHFFFAOYSA-N trimethyl phosphate Chemical compound COP(=O)(OC)OC WVLBCYQITXONBZ-UHFFFAOYSA-N 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 238000005299 abrasion Methods 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 239000012491 analyte Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 231100000357 carcinogen Toxicity 0.000 description 1
- 231100000315 carcinogenic Toxicity 0.000 description 1
- 239000003183 carcinogenic agent Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000037406 food intake Effects 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 231100000024 genotoxic Toxicity 0.000 description 1
- 230000001738 genotoxic effect Effects 0.000 description 1
- 231100000003 human carcinogen Toxicity 0.000 description 1
- 239000010720 hydraulic oil Substances 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000002887 neurotoxic effect Effects 0.000 description 1
- 239000003973 paint Substances 0.000 description 1
- 238000004451 qualitative analysis Methods 0.000 description 1
- 238000012502 risk assessment Methods 0.000 description 1
- 238000011896 sensitive detection Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- WTLBZVNBAKMVDP-UHFFFAOYSA-N tris(2-butoxyethyl) phosphate Chemical compound CCCCOCCOP(=O)(OCCOCCCC)OCCOCCCC WTLBZVNBAKMVDP-UHFFFAOYSA-N 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
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- 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
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- 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
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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- 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
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/28—Control of physical parameters of the fluid carrier
- G01N30/30—Control of physical parameters of the fluid carrier of temperature
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- 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
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/28—Control of physical parameters of the fluid carrier
- G01N30/32—Control of physical parameters of the fluid carrier of pressure or speed
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- 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
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
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- G01N30/34—Control of physical parameters of the fluid carrier of fluid composition, e.g. gradient
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G01N30/02—Column chromatography
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/28—Control of physical parameters of the fluid carrier
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Abstract
The invention discloses a method for detecting and analyzing organic phosphate in a water sample, which comprises the following steps: diluting the mixed standard solution of the organic phosphate to obtain gradient mixed standard solutions with different concentrations; after filtering the sample, adding an internal standard substance, and vortexing for 30 s; add 1mL dichloromethane, vortex for 1 min; centrifuging at 3000rpm for 2min, removing upper water phase, blowing organic phase nitrogen to near dryness, diluting with methanol to constant volume, and filtering to obtain pretreated sample; and (3) determining the standard solution and the sample by using a high performance liquid chromatography-tandem mass spectrometry method, and carrying out quantitative analysis by using an internal standard method to obtain the content of the organic phosphate in the sample.
Description
Technical Field
The invention relates to the field of environmental detection and analysis methods, in particular to a trace rapid detection and analysis method for organic phosphate in a water sample.
Background
Organophosphates (ops) are widely used in textile, electronics, plastics and other industries as flame retardants and plasticizers. OPEs are a group of compounds with the same phosphate basic unit-the phosphate center and different substituents. OPEs are classified into halogenated alkyl phosphates, non-halogenated alkyl phosphates and aryl phosphates according to the type of the substituent. In use, chloroalkyl and aryl phosphates are mainly used as flame retardants for plastics, textiles, electronics and furniture and construction to improve fire resistance. Non-chlorinated alkyl phosphates are mainly used as plasticizers, also as defoamers, paint additives, hydraulic oils or floor polishes.
Chloroalkyl phosphates are listed in the European Union (EU) priority list 2 (1995) and 4 (2000) for risk assessment, and tris (2-chloroethyl) phosphate (TCEP) has been listed as a "potential human carcinogen" (carcinogen class 3) by the european union in 2008. Non-chlorinated organophosphates, trimethyl phosphate (TMP), are considered genotoxic, tributyl phosphate (TnBP) and triphenyl phosphate (TPHP) also have neurotoxic effects, and trioxyethyl phosphate (TBEP) is also a suspected carcinogenic compound.
Because the OPEs used are incorporated into the material in a physical blend rather than a chemical bond, they are readily released from the plastic and enter the environment by volatilization, abrasion and dissolution, and are transported remotely by further atmospheric air and recycled into different environmental media. Humans may be exposed to OPEs by various routes, such as inhalation, ingestion, and skin contact. It has been reported that OPEs and their metabolites have been detected in animal tissues and human blood and urine. Given the continuing reports of adverse effects of OPEs, the presence, migration, and transformation of such chemicals in the environment has attracted considerable attention. In order to accurately evaluate the environmental risk of the OPEs, a convenient, accurate and sensitive detection method is urgently needed to be established, in particular to a detection method of the OPEs in a water sample.
Disclosure of Invention
The invention aims to overcome the defects of the prior art and provides a method for detecting and analyzing organic phosphate in a water sample. Aiming at the problems of low concentration of OPEs in an environmental medium, large sample volume, large organic solvent consumption, complex operation and the like in the traditional liquid-liquid extraction process, the method adopts a trace liquid-liquid extraction method to extract the organic phosphate in a water sample, and has the advantages of simple operation and short time consumption; and the enriched organic phosphate is quantitatively analyzed by using HPLC-MS, so that the organic phosphate can be rapidly and quantitatively determined.
A method for detecting and analyzing organic phosphate in a water sample comprises the following steps:
preparing a standard solution: for 1. mu.g.mL -1 The organic phosphate mixed standard solution was diluted and added at a concentration of 200. mu.g.L -1 Obtaining the concentration of the internal standard substance of 20. mu.g.L -1 The concentrations of the 12 organic phosphates were 0.2, 1, 5, 10, 25, 50, 100, 200 and 500. mu.g.L, respectively -1 The prepared gradient mixed standard solution is prepared; the internal standard substances in the standard solution preparation stage are TNBP-d27 and TPHP-d 15;
sample pretreatment: filtering the collected water sample by using a 0.45-micron microporous filter membrane, adding 10mL of the filtered water sample into a 10mL glass centrifuge tube, adding a certain amount of internal standard substance, performing vortex for 30s, and fully and uniformly mixing; adding 1mL of dichloromethane, vortexing for 1min, and fully and uniformly mixing; centrifuging at 3000rpm for 2min to obtain water phase and organic phase with obvious layering; removing the upper aqueous phase, sucking the lower organic phase into a sample injection vial, blowing the organic phase nitrogen to be nearly dry, fixing the volume to 200 mu L with methanol, uniformly mixing the organic phase nitrogen and the methanol in a vortex manner for 30s, and filtering the mixture through a 0.22 mu m filter membrane to obtain a pretreated sample; storing the pretreated sample in a refrigerator at the temperature of-20 ℃; wherein the internal standard substances in the sample pretreatment stage refer to TNBP-d27 and TPHP-d15 with the mass of 4 ng;
determining the gradient mixed standard solution and the pretreated sample by using a high performance liquid chromatography tandem mass spectrometry method to obtain HPLC-MS parameters of each organic phosphate monomer;
and (3) carrying out quantitative analysis by adopting an internal standard method, drawing a standard curve of the organic phosphate by using detection data of the gradient mixed standard solution, and obtaining the content of the organic phosphate in the sample according to the corresponding relation between response and concentration.
Further, the mixed standard solution of organic phosphates comprises six alkyl organic phosphates, three chloroalkyl organic phosphates, and three aryl organic phosphates, wherein the alkyl organic phosphates comprise: triethyl phosphate (TEP), tripropyl phosphate (TPrP), tri-n-butyl phosphate (TnBP), tri-n-pentyl phosphate (TPeP), tri (2-ethylhexyl) phosphate (TEHP), tri (butoxyethyl) phosphate (TBOEP); the chloroalkyl organophosphate esters include: tris (2-chloroethyl) phosphate (TCEP), tris (2-chloropropyl) phosphate (TCIPP), tris (1, 3-dichloroisopropyl) phosphate (TDCIPP); the aryl organic phosphate ester: triphenyl phosphate (TPHP), tricresyl phosphate (TCrP), tris (2-ethylhexyl diphenyl) phosphate (EHDPP).
Further, the drawing of the standard curve comprises the following steps: concentration of 20. mu.g.L for internal standard -1 And the concentrations of the 12 organic phosphates are 0.2, 1, 5, 10, 25, 50, 100, 200 and 500 mug. multidot.L -1 The mixed standard solution is subjected to HPLC-MS test to obtain the peak area of the quantitative ion integral, and a relation curve of the concentration ratio and the area ratio of the component to be tested and the internal standard substance is made, namely the standard curve.
Further, after the content of the organic phosphate is obtained, a blank labeling or matrix labeling method is adopted for verification, and the method specifically comprises the following steps:
adding 1 mu g/mL into ultrapure water, seawater, tap water and lake water samples respectively -1 Mixing 10 mu L of standard solution, repeatedly measuring each water sample for five times, and testing the standard adding recovery rate of the sample to be tested so as to judge the accuracy of the content of the organic phosphate.
Compared with the prior art, the technical scheme of the invention has the following beneficial effects:
according to the invention, the trace liquid-liquid extraction method is adopted to extract the organic phosphate in the water sample, so that the problems of large volume demand, large organic solvent consumption, complex operation and the like of the traditional liquid-liquid extraction sample can be avoided, and meanwhile, the used extraction solvent has a wide polarity selection range, can complete the rapid extraction of a plurality of organic phosphates, and improves the detection efficiency.
The method adopts high performance liquid chromatography tandem mass spectrometry to detect the organic phosphate in the enriched sample, greatly improves the sensitivity and selectivity of the analysis method in a quantitative reaction monitoring mode, reduces the detection limit, and is suitable for quantitative detection of trace organic phosphate in various water environments.
The invention can effectively separate, accurately determine the nature and the quantity of 12 organic phosphate esters, and the linear range of the 12 organic phosphate esters is 0.2-500 mu g.L -1 Correlation coefficient (R) of standard curve 2 ) All are more than 0.99, the blank standard addition recovery rate of the target object is 86-109%, and the detection limit is 2.19-125.26 ng.L -1 。
Drawings
FIG. 1 is a schematic flow chart of the method for detecting and analyzing organic phosphate in a water sample according to the present invention;
FIG. 2 is a chromatogram of a total ion flow chart of a ultrapure water spiked sample before and after treatment in example 1;
FIG. 3 is a chromatogram of a total ion flow chart of a seawater spiked sample before and after treatment in example 2;
FIG. 4 is a chromatogram of a total ion flow chart of a tap water standard sample before and after treatment in example 2;
FIG. 5 is a chromatogram of a total ion flow chart of a lake water spiked sample before and after treatment in example 2.
Detailed Description
The technical solutions of the present invention are further described in detail with reference to the drawings and specific embodiments, which are only illustrative and not intended to limit the present invention.
Example 1: blank value and blank standard addition recovery rate experiment.
A method for detecting and analyzing organic phosphate in a water sample comprises the following steps:
the method comprises the following steps: preparing a standard solution: diluting the mixed standard solution of organic phosphate to obtain the concentrations of 0.2, 1, 5, 10, 20, 50, 200 and 500. mu.g.L -1 The gradient of (2) mixes the standard solution. The organophosphates analyzed included six alkyl organophosphates: triethyl phosphate (TEP), tripropyl phosphate (TPrP), tri-n-butyl phosphate (TnBP), tri-n-pentyl phosphate (TPeP), tri (2-ethylhexyl) phosphate (TEHP), tri (butoxyethyl) phosphate (TBOEP); three chloroalkyl organophosphates: tris (2-chloroethyl) phosphate (TCEP), tris (2-chloropropyl) phosphate(TCIPP), tris (1, 3-dichloroisopropyl) phosphate (TDCIPP); three aryl organophosphates: triphenyl phosphate (TPHP), tricresyl phosphate (TCrP), tris (2-ethylhexyl diphenyl) phosphate (EHDPP).
The preparation of the standard solution in the step one is specifically to prepare internal standard substances (TNBP-d27 and TPHP-d15) with the concentration of 20 mu g.L -1 The concentrations of the 12 organic phosphates were 0.2, 1, 5, 10, 25, 50, 100, 200, 500. mu.g.L -1 The standard solutions were mixed in a gradient and the formulation details are shown in table 1.
TABLE 1 preparation of the Standard solutions
Wherein in Table 1, (. sup.X) is 1. mu.g/mL -1 The mixed internal standard A refers to the internal standard substance added in the step one and is 200 mug.L -1 TNBP-d27 and TPHP-d 15.
② taking a sample injection bottle Y, adding 100 mu L X (1 mu g. mL) -1 ) 900 μ L of methanol, totaling 1000 μ L, denoted as Y (100 μ g. L) -1 )。
③ taking a sample injection bottle Z, adding 100 mu L Y (100 mu g. L) -1 ) 900 μ L of methanol, totaling 1000 μ L, and is reported as Z (10 μ g. L) -1 )。
Fourthly, taking a sample injection bottle W, adding 100 mu L Z (10 mu g.L) -1 ) 900 μ L of methanol, 1000 μ L in total, and is denoted as W (1 μ g. L) -1 )。
Step two: sample pretreatment: filtering the collected water sample by using a 0.45-micron microporous filter membrane, adding 10mL of the filtered water sample into a 10mL glass centrifuge tube, continuously adding 4ng of internal standard substances TNBP-d27 and TPHP-d15, swirling for 30s, and fully and uniformly mixing; adding 1mL of dichloromethane, vortexing for 1min, and fully and uniformly mixing; centrifuging at 3000rpm for 2min, and separating two phases; the upper aqueous phase was removed with a dropper and the lower organic phase was transferred to a 2mL injection vial with a syringe; nitrogen was blown to near dryness and the volume was made up to 200 μ L with methanol, vortexed for 30s and filtered through a 0.22 μm filter. And storing the pretreated sample in a refrigerator at the temperature of 20 ℃ below zero for HPLC-MS detection.
Step three: the standard solution and the sample were determined by high performance liquid chromatography tandem mass spectrometry.
The conditions of the high performance liquid chromatography-mass spectrometry of the step three are as follows:
liquid chromatography conditions:
an Agilent Infinity 1260 high performance liquid chromatography system; a chromatographic column: zorbax Eclipse Plus C 18 A column (2.1 mm. times.150 mm; 3.5 μm); the column temperature was 40 ℃; mobile phase: a: 0.1% aqueous formic acid solution, B: pure methanol; the flow rate of the mobile phase is 0.4 mL/min -1 (ii) a The sample injection volume is 10 mu L; the gradient elution conditions are shown in Table 2
TABLE 2 gradient elution conditions
Mass spectrum conditions: an Agilent 6460 triple quadrupole mass spectrometer detector; an ion source: ESI, ion source temperature: 300 ℃, ionization mode: a positive ion mode; flow rate of drying gas: 7L min -1 (ii) a Atomizer pressure: 45 psi; temperature of sheath gas: 350 ℃, sheath gas flow rate: 11 L.min -1 (ii) a Collision voltage: 3.5 kV; nozzle voltage: 0 kV; the collection mode is as follows: quantitative reaction monitoring mode (MRM). The HPLC-MS parameters for each organophosphate monomer are shown in Table 3:
TABLE 3 HPLC-MS parameters
Step four: quantitative analysis was performed by the internal standard method, and a standard curve of organophosphate was plotted using the detection data of the gradient mixed standard solution, with the results shown in table 4. And obtaining the content of the organic phosphate in the sample according to the corresponding relation between the response and the concentration. The internal standard method is characterized in that a certain amount of internal standard substance is added into a sample so as to correct the system error of the whole experimental process, and the operation condition is slightly changed or the error of an instrument does not have great influence on the result. Plotting the organophosphate Standard curves specifically for the concentrations of the internal standards (TNBP-d27 and TPHP-d15)20 μ g.L -1 The concentrations of the 12 organic phosphates were 0.2, 1, 5, 10, 25, 50, 100, 200, 500. mu.g.L -1 The gradient mixed standard solution is subjected to HPLC-MS test to obtain the peak area of the quantitative ion integral, and a relation curve of the concentration ratio (the measured component and the internal standard substance) and the area ratio (the measured component and the internal standard substance) is made, and the curve is a standard curve.
Table 4 calibration curve:
to the ultrapure water sample was added 10. mu.L of a mixed standard solution (1. mu.g. mL) -1 ) The recovery of the analyte was tested and the assay was repeated five times for each sample as a parallel control. FIG. 2 is a chromatogram of a total ion flow chart of a sample of ultrapure water spiked before and after the treatment in example 1; by using the method to extract and measure the content of each organic phosphate monomer in the ultrapure water, the blank value and the blank standard addition recovery rate of the method can be calculated (see table 5).
Table 5 methods blank value and blank normalized recovery (n ═ 5)
ND: not detected, Not detected
Example 2: the method is applied to the analysis of organic phosphate in Bohai Bay near-shore seawater, tap water and landscape lakes.
The specific implementation steps of example 2 are similar to the steps one to four of example 1, and only the differences are described below.
In the fourth step, an internal standard method is adopted for quantitative analysis, and 12 organic phosphate esters in seawater, tap water and landscape lake water are respectively extracted and measured, wherein each organic phosphate ester monomer is not detected (see table 6).
To test the utility of the proposed method, the recovery of the method in water samples from different sources was tested by matrix labeling experiments.
In the seawater,Adding 1 mu g/mL into tap water and lake water samples -1 10 μ L of standard solution was mixed and set in five replicates. The method for extracting and measuring the organic phosphate in the seawater, tap water and lake water standard adding samples can obtain the standard adding recovery rates of different matrixes, can examine the recovery effect of the organic phosphate in actual water samples, and confirms the effectiveness of the method, and the test results are shown in Table 6.
FIGS. 3 to 5 show the total ion flow chromatogram of the seawater, tap water and lake water spiked samples before and after treatment in example 2, respectively.
TABLE 6 contents of 12 organophosphates in three water samples and standard recovery (n ═ 5)
In the analysis of actual water samples, the recovery rate of the added standard is between 90% and 125% (the recovery rate of TEHP in seawater is 140%, the recovery rate of TEHP in lake water is 145%), and the RSD is lower than 20%. Compared with other methods, the method has the advantages of simplicity, convenience, rapidness, reagent saving and the like. Fig. 3-5 show that the method can effectively realize the enrichment of the OPEs in different water bodies and is suitable for the rapid qualitative and quantitative analysis of the OPEs in the highly polluted water body.
Although the preferred embodiments of the present invention have been described with reference to the accompanying drawings, the present invention is not limited to the above-described embodiments, which are only illustrative and not restrictive, and those skilled in the art can make many modifications without departing from the spirit and scope of the invention as claimed.
Claims (4)
1. A method for detecting and analyzing organic phosphate in a water sample is characterized by comprising the following steps:
preparing a standard solution: for 1. mu.g.mL -1 Diluting the mixed standard solution of the organic phosphate, and adding a certain amount of internal standard substance to obtain the internal standard substance with the concentration of 20 mu g.L -1 The concentrations of the 12 organic phosphates were 0.2, 1, 5, 10, 25, 50, 100, 200 and 500. mu.g.L, respectively -1 The prepared gradient mixed standard solution is prepared; the internal standard substances in the standard solution preparation stage are TNBP-d27 and TPHP-d 15;
sample pretreatment: filtering the collected water sample by using a 0.45-micron microporous filter membrane, adding 10mL of the filtered water sample into a 10mL glass centrifuge tube, adding a certain amount of internal standard substance, performing vortex for 30s, and fully and uniformly mixing; adding 1mL of dichloromethane, vortexing for 1min, and fully and uniformly mixing; centrifuging at 3000rpm for 2min to obtain water phase and organic phase with obvious layering; removing the upper aqueous phase, sucking the lower organic phase into a sample injection vial, blowing the organic phase nitrogen to be nearly dry, fixing the volume to 200 mu L with methanol, uniformly mixing the organic phase nitrogen and the methanol in a vortex manner for 30s, and filtering the mixture through a 0.22 mu m filter membrane to obtain a pretreated sample; storing the pretreated sample in a refrigerator at the temperature of-20 ℃; wherein the internal standard substances in the sample pretreatment stage refer to TNBP-d27 and TPHP-d15 with the mass of 4 ng;
and (3) determining the gradient mixed standard solution and the sample by using a high performance liquid chromatography tandem mass spectrometry, performing quantitative analysis by using an internal standard method, drawing a standard curve of the organic phosphate by using the detection data of the standard solution, and obtaining the content of the organic phosphate in the sample according to the corresponding relation of response and concentration.
2. The method for detecting and analyzing organic phosphate in water samples according to claim 1, wherein the mixed standard solution of organic phosphate comprises six kinds of alkyl organic phosphate, three kinds of chloroalkyl organic phosphate and three kinds of aryl organic phosphate, wherein the alkyl organic phosphate comprises: triethyl phosphate (TEP), tripropyl phosphate (TPrP), tri-n-butyl phosphate (TnBP), tri-n-pentyl phosphate (TPeP), tri (2-ethylhexyl) phosphate (TEHP), tri (butoxyethyl) phosphate (TBOEP); the chloroalkyl organophosphate esters include: tris (2-chloroethyl) phosphate (TCEP), tris (2-chloropropyl) phosphate (TCIPP), tris (1, 3-dichloroisopropyl) phosphate (TDCIPP); the aryl organophosphate ester: triphenyl phosphate (TPHP), tricresyl phosphate (TCrP), tris (2-ethylhexyl diphenyl) phosphate (EHDPP).
3. The method for detecting and analyzing organic phosphate in water samples according to claim 1, wherein the standard curve is drawn by the following steps: concentration of 20. mu.g.L for internal standard -1 The concentrations of the 12 organic phosphates were 0.2, 1, 5, 10, 25, 50, 100, 200, 500. mu.g.L -1 The gradient mixed standard solution is subjected to HPLC-MS test to obtain the peak area of the quantitative ion integral, and a relation curve of the concentration ratio and the area ratio of the component to be tested and the internal standard substance is made, namely the standard curve.
4. The method for detecting and analyzing organic phosphate in water samples according to claim 1, further comprising the following steps after obtaining the content of the organic phosphate:
adding 1 mu g/mL into ultrapure water, seawater, tap water and lake water samples respectively -1 Mixing 10 μ L of standard solution, repeating the determination for five times for each sample, and testing the recovery rate of the sample to be tested by adding standard, thereby testing the accuracy of the obtained content of the organic phosphate.
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| CN111044643A (en) * | 2019-12-31 | 2020-04-21 | 中国环境科学研究院 | Detection method of organic phosphate |
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