Non-targeted identification method for phthalate pollutants in site soil

Abstract

The invention discloses a non-targeted recognition method for phthalate pollutants in site soil. The non-targeted recognition method comprises the following steps: collecting a soil sample from soil possibly polluted by phthalate; extracting organic pollutants in the soil sample by using an ultrasonic-assisted solid-liquid extraction method; and performing phthalate analysis and detection on a to-be-detected sample solution obtained by extracting the organic pollutants from the soil sample by using high performance liquid chromatography-tandem mass spectrometry. Based on the solid phase extraction and liquid chromatography-tandem mass spectrometry technology, cross contamination in the pretreatment and detection process can be reduced, meanwhile, compared with traditional targeted screening for detecting limited pollutants, interference is smaller, more geological spectrum peaks can be recognized, and wide and comprehensive recognition can be conducted.

Description

Technical Field

[0001] The invention relates to the technical field of environmental detection, and in particular to a non-targeted identification method for phthalate pollutants in site soil. Background Art

[0002] Phthalates (PAEs), also known as phthalates, are common plasticizers that are widely used in daily life and production. They are one of the most produced and widely used synthetic organic compounds in the world. With the extensive use of PAEs, they are also commonly found in the atmosphere, water bodies, soil, etc. PAEs can interfere with the endocrine system of humans and animals at extremely low concentrations, and some of them have carcinogenic, teratogenic, and mutagenic effects. They are a typical endocrine disruptor. However, the detection and management of PAEs in soil have not yet been fully established. Therefore, detecting its residual level in soil is of great significance to controlling soil pollution and protecting human health.

[0003] According to literature research, there are currently few methods for detecting phthalates in soil. On the one hand, ultrasonic extraction pretreatment technology coupled with fluorescence method is used to detect PAEs in soil. On the other hand, C 18 The pretreatment method of solid phase membrane extraction is coupled with gas chromatography-mass spectrometry to detect PAEs in soil. In addition, the method of using QuEChREs method to pretreat samples and couple them with gas chromatography-mass spectrometry to determine PAEs in soil is also mentioned. It is worth noting that these traceability methods are still obviously insufficient. This is mainly reflected in the current field of analytical chemistry. The content of PAEs that can be qualitatively and quantitatively analyzed only constitutes a very limited fragment of the amount of PAEs present. In view of the diversity and complexity of chemical substances in the environment, traditional targeted analysis methods are more difficult to fully depict the composition map of environmental organic pollutants due to problems such as low accuracy and large background interference. Summary of the invention

[0004] The purpose of the present invention is to provide a non-targeted identification method for phthalate ester pollutants in site soil in view of the limitations of the current qualitative and quantitative analysis methods of phthalate esters in the field of analytical chemistry, the problems of low spectral accuracy and large background interference.

[0005] The present invention realizes the high-throughput screening of the whole process for new pollutants of phthalic acid esters in soil through soil sample collection, preservation, transportation, pretreatment, on-machine detection, and analysis and identification; based on solid-phase extraction (SPE) and liquid chromatography-tandem mass spectrometry (LC-MS) technologies, cross-contamination during the pretreatment and detection processes is reduced, and compared with the traditional targeted screening for detecting a limited number of pollutants, it has less interference, can identify more geological spectral peaks, and can conduct extensive and comprehensive identification.

[0006] To achieve the above object, the technical solution provided by the present invention is:

[0007] A non-targeted identification method for phthalic acid ester pollutants in site soil, comprising the following steps:

[0008] Collect soil samples from the soil that may be contaminated with phthalic acid esters;

[0009] Use ultrasonic-assisted solid-liquid extraction method to extract organic pollutants in the soil samples;

[0010] Use high-performance liquid chromatography-tandem mass spectrometry to analyze and detect phthalic acid esters in the test sample solution obtained by extracting organic pollutants from the soil samples;

[0011] The conditions of high-performance liquid chromatography-tandem mass spectrometry are as follows:

[0012] High-performance liquid chromatography determination:

[0013] Use a C18 ultra-high performance liquid chromatography column, column temperature: 35 - 45 °C;

[0014] Use an aqueous methanol solution containing 1 - 3 mM ammonium acetate and 3 - 7% methanol as mobile phase A, and methanol as mobile phase B;

[0015] Mobile phase flow rate: 0.2 - 0.4 L / min; elution detection time 45 - 65 min;

[0016] Mass spectrometry determination:

[0017] Use a quadrupole tandem high-resolution orbitrap mass spectrometer, and use an electrospray ionization ion source;

[0018] Spray voltage: -2500 V / +3500 V; capillary temperature 300 - 340 °C; the first-stage mass spectrometry scanning range is m / z 80 to 1000, and the resolution is 70000; the mass spectrometry collects fragment ions with m / z greater than 50, and three second-stage fragment ions of the three most abundant first-stage parent ions are collected for each first-stage scanning window;

[0019] The method for the analysis and detection of phthalates is as follows: By retrieving the substance information of PAEs in the MSDIAL software of the mass spectrometry, a PAEs database is constructed, and the sample data is compared based on the PAEs database.

[0020] To optimize the above technical solution, the specific measures taken also include:

[0021] When collecting soil samples, manual sampling of shallow soil is carried out using a Luoyang shovel, and sampling of deep soil is carried out using a DP-50 diesel core drill. After collecting the soil samples, ensure the integrity of the samples and store them at a low temperature of 4 °C during the process of transferring and transporting the soil samples to the determination laboratory.

[0022] The ultrasonic-assisted solid-liquid extraction method is used to extract organic pollutants in soil samples. Specifically: Weigh the soil samples and grind them evenly. Use anhydrous sodium sulfate to remove water from the soil samples, put them into centrifuge tubes, add internal standards and age for 15 - 20 h. Use methanol to ultrasonically extract the samples 3 times at 38 - 42 °C in a water bath using different extraction solvents respectively, and combine the extraction solutions.

[0023] Further, the different extraction solvents used 3 times are dichloromethane - n-hexane at 1:1 v / v, n-hexane - acetone at 1:1 v / v, and acetone respectively.

[0024] After extraction by the ultrasonic-assisted solid-liquid extraction method, concentration, volume fixation, and filtration are carried out to obtain the sample solution to be measured.

[0025] Further, in the high-performance liquid chromatography determination conditions, the column temperature is 40 °C; a 5% methanol aqueous solution containing 2 mM ammonium acetate is used as mobile phase A, and methanol is used as mobile phase B.

[0026] Further, in the high-performance liquid chromatography determination conditions, the flow rate of the mobile phase is 0.3 L / min; the elution detection time is 55 min.

[0027] Further, in high-performance liquid chromatography - tandem mass spectrometry, the injection volume is 4 - 20 μm.

[0028] Further, in the mass spectrometry determination conditions, the capillary temperature is 320 °C; the sheath gas flow rate is 48 L / h, the auxiliary gas flow rate is 11 L / h, and the standby gas flow rate is 2 L / h.

[0029] Further, in the analysis and detection of phthalates, only substances within a mass error range of 5 mDa and matching 2 or more secondary mass spectrometry fragments in the PAEs database are retained. If the secondary mass spectrometry information of a substance in the PAEs database contains only 1 secondary mass spectrometry fragment, then the substance is excluded.

[0030] Compared with the prior art, the beneficial effects of the present invention are:

[0031] The present invention proposes a full - process identification scheme from sampling to analytical detection and a pretreatment technology represented by solid - phase extraction. While reducing cross - contamination, it can accurately identify PAEs in soil. The detection of PAEs in soil by the present invention uses liquid chromatography - high - resolution mass spectrometry technology, which can significantly reduce interference during the analytical detection process and perform high - throughput screening of PAEs in soil. The present invention forms a database based on the PAEs data in MSDIAL to compare with the sample data in MSDIAL, which is more convenient and has higher accuracy compared to traditional detection and analysis methods.

[0032] The present invention fills the gap in the lack of a full - process identification method for PAEs in soil from sample collection to analytical detection. It conducts full - process analytical detection of new PAEs pollutants in soil and simplifies the pretreatment process using a pretreatment technology represented by solid - phase extraction, preventing the problem of cross - contamination easily caused by the original pretreatment technology, providing technical support for the investigation and control of PAEs in the soil environment; the present invention is the first solution proposed for phthalate pollutants in the environmental medium of site soil, and more suitable extraction solvents and extraction methods are selected according to the different medium characteristics in site soil; the present invention uses liquid chromatography - high - resolution mass spectrometry technology to widely and comprehensively identify new phthalic acid pollutants in soil, significantly reducing interference during the analytical detection process, and proposes a screening method of database screening to perform high - throughput screening of PAEs in soil, enabling comprehensive analysis of a large number of compounds. Description of the Drawings

[0033] Figure 1 : Schematic flow chart of the non - targeted identification method for phthalate pollutants in the site soil of the present invention. Detailed Embodiments

[0034] The above - mentioned content of the present invention will be further described in detail below in the form of specific embodiments, but it should not be understood that the scope of the above - mentioned subject matter of the present invention is limited to the following embodiments. All technologies implemented based on the above - mentioned content of the present invention belong to the scope of the present invention.

[0035] The experimental methods used in the following embodiments are all conventional methods unless otherwise specified. The reagents, methods, and equipment used are all conventional reagents, methods, and equipment in the technical field unless otherwise specified.

[0036] For the sake of brevity, only some numerical values and optional ranges are specifically disclosed herein. However, any lower limit can be combined with any upper limit to form a range not explicitly recorded; and any lower limit can be combined with other lower limits to form a range not explicitly recorded. Similarly, any upper limit can be combined with any other upper limit to form a range not explicitly recorded; the optional items in the optional range can also be combined arbitrarily.

[0037] The present invention provides a non-targeted identification method for phthalate pollutants in site soil, comprising the following steps:

[0038] Collect soil samples from soil where phthalate pollution may exist;

[0039] Use ultrasonic-assisted solid-liquid extraction to extract organic pollutants from the soil samples;

[0040] Use high performance liquid chromatography-tandem mass spectrometry to analyze and detect phthalates in the test sample solution obtained by extracting organic pollutants from the soil samples;

[0041] The conditions for high performance liquid chromatography-tandem mass spectrometry are as follows:

[0042] High performance liquid chromatography determination:

[0043] Use a C18 ultra-high performance liquid chromatography column, column temperature: 35 - 45 °C;

[0044] Use an aqueous methanol solution containing 1 - 3 mM ammonium acetate and 3 - 7% methanol as mobile phase A, and methanol as mobile phase B;

[0045] Mobile phase flow rate: 0.2 - 0.4 L / min; elution detection time 45 - 65 min;

[0046] Mass spectrometry determination:

[0047] Use a quadrupole tandem high-resolution orbitrap mass spectrometer, and use an electrospray ionization ion source;

[0048] Spray voltage: -2500 V / +3500 V; capillary temperature 300 - 340 °C; the first-stage mass spectrometry scanning range is m / z 80 to 1000, and the resolution is 70000; the mass spectrometry collects fragment ions with m / z greater than 50, and for each first-stage scanning window, the second-stage fragment ions of the 3 most abundant first-stage parent ions are collected;

[0049] The method for analyzing and detecting phthalates is: by retrieving the substance information of PAEs in the MSDIAL software of the mass spectrometry, constructing a PAEs database, and comparing it with the sample data based on the PAEs database.

[0050] When collecting soil samples in this application, manual sampling of shallow soil is carried out using a Luoyang shovel, and deep soil is sampled using a DP-50 diesel core drill; after collecting the soil samples, during the process of transferring and transporting the soil samples to the determination laboratory, ensure that the samples are intact and stored at a low temperature of 4 °C.

[0051] Use ultrasonic-assisted solid-liquid extraction method to extract organic pollutants from soil samples. Specifically, weigh the soil samples and grind them evenly. Use anhydrous sodium sulfate to remove water from the soil samples, put them into centrifuge tubes, add internal standards and age for 15 - 20 h. Use methanol to ultrasonically extract the samples 3 times at 38 - 42 °C in a water bath with different extraction solvents respectively, and combine the extraction solutions.

[0052] In some embodiments, the different extraction solvents used 3 times are dichloromethane - n - hexane at 1:1 v / v, n - hexane - acetone at 1:1 v / v, and acetone respectively.

[0053] After extraction by ultrasonic-assisted solid-liquid extraction method, concentrate, make up the volume and filter to obtain the sample solution to be measured.

[0054] In some embodiments, in the high performance liquid chromatography determination conditions, column temperature: 40 °C; use 5% methanol aqueous solution containing 2 mM ammonium acetate as mobile phase A, and methanol as mobile phase B.

[0055] In some embodiments, in the high performance liquid chromatography determination conditions, mobile phase flow rate: 0.3 L / min; elution detection time 55 min.

[0056] In some embodiments, in the mass spectrometry determination conditions, capillary temperature 320 °C; sheath gas flow rate: 48 L / h, auxiliary gas flow rate: 11 L / h, standby gas flow rate: 2 L / h.

[0057] Construct a PAEs database by retrieving the substance information of PAEs in MSDIAL of the mass spectrometry, and compare the sample data with the PAEs database formed based on the PAEs data in MSDIAL.

[0058] In some embodiments, in the analysis and detection of phthalic acid esters, only retain the substances within a mass error range of 5 mDa and matching 2 or more secondary mass spectrometry fragments with the PAEs database. If the secondary mass spectrometry information of the substance in the PAEs database only contains 1 secondary mass spectrometry fragment, then exclude the substance.

[0059] The technical solution of the present invention will be further described in detail with specific embodiments as follows:

[0060] Example

[0061] A non-targeted identification survey method for new phthalic acid ester pollutants in site soil, comprising the following steps:

[0062] (1) Sample collection, preservation, and transportation: Based on the original land use function and pollution characteristics, several working units with relatively heavy potential PAEs pollution were selected as the working units for pollutant identification. Manual sampling of shallow soil was carried out using a Luoyang shovel, and deep soil sampling was performed using a DP-50 diesel core drill.

[0063] Before sampling, the sampling instruments must be thoroughly cleaned and disinfected to avoid cross-contamination. Before sample shipment, fill out the "Sample Transfer Form" and firmly attach the label outside the tube, indicating the project name, borehole number, sampling number, depth, date, tube number, and soil type. After sample collection, the samples were sent to the designated laboratory within the specified time according to the sampling plan. The soil sample transfer and transportation ensured the integrity of the samples and were stored at a low temperature of 4°C.

[0064] (2) Laboratory extraction: Weigh 10.0 g of soil samples and grind them evenly. Use anhydrous sodium sulfate to remove water from the soil samples, put them into centrifuge tubes, add internal standards and age for 16 h. Use methanol to extract the samples three times at 40°C in a water bath using dichloromethane-n-hexane (1:1, v / v), n-hexane-acetone (1:1, v / v), and acetone by ultrasonic extraction. The ultrasonic time for each time is 20 min, and the ultrasonic temperature is 40°C. A control experiment of the whole procedure blank was carried out for the extraction of soil samples.

[0065] (3) Concentration and volume fixation: After ultrasonic extraction of soil samples, the ultrasonic extraction supernatants were combined, nitrogen blown to dryness, fixed to 1 mL with n-hexane, passed through a 0.45 μm PTFE filter membrane, and stored at -20°C in a chromatographic injection vial.

[0066] (4) On-machine detection: High-performance liquid chromatography-tandem mass spectrometry was used to analyze and detect phthalic acid esters in the samples. The liquid chromatography system was the UltiMate 3000 Series, and the high-resolution mass spectrometry system was a quadrupole tandem high-resolution orbitrap mass spectrometer. The liquid chromatography column was a C18 ultra-high performance liquid chromatography column, and the column temperature was 40°C. Phase A was a 5% methanol aqueous solution containing 2 mM ammonium acetate as a buffer salt, and Phase B was methanol. The mobile phase flow rate was 0.3 L / min, and gradient elution was carried out for 55 min using a two-phase mobile phase for gradient elution to achieve better separation effects. The mass spectrometry ionization source was an electrospray ionization (ESI) ion source, the spray voltage was -2500 V / +3500 V, the capillary temperature was 320°C, the sheath gas flow rate was 48 L / min, the auxiliary gas flow rate was 11 L / min, and the standby gas flow rate was 2 L / min.

[0067] The first-level mass spectrometry scanning range was m / z 80 to 1000, and the resolution was 70000. The second-level mass spectrometry collected fragment ions with m / z greater than 50, and the resolution was 17500. The second-level fragment ions of the three most abundant first-level parent ions were collected in each first-level scanning window.

[0068] (5) Analysis and identification: Import the detection results into MSDIAL. MSDIAL only retains substances that match two or more secondary mass spectrometry fragments in the database within a mass error range of 5 mDa. If the secondary mass spectrometry information of a substance in the database contains only one secondary mass spectrometry fragment, the substance is not retained. At the same time, establish a PAEs database based on the substance information of PAEs in MSDIAL, and compare the data detected by MSDIAL with the database, so as to identify phthalic acid substances in the soil.

[0069] The PAEs database is as follows:

[0070] Table 1: List of screening databases in positive ion mode

[0071]

[0072] Table 2: List of screening databases in negative ion mode

[0073]

[0074]

[0075] The above are only the preferred embodiments of the present invention, and do not impose any form of limitation on the present invention. Any person skilled in the art, without departing from the scope of the technical solution of the present invention, based on the technical essence of the present invention, any simple modification, equivalent replacement, and improvement made to the above embodiments still fall within the protection scope of the technical solution of the present invention.

Claims

1. A non-targeted identification method for phthalate pollutants in site soil, characterized in that: The following steps are involved: Collect soil samples from soils that may be contaminated with phthalates; Extraction of organic pollutants from soil samples using ultrasound-assisted solid-liquid extraction; The sample solution obtained by extracting organic pollutants from soil samples was analyzed and tested for phthalates using high performance liquid chromatography-tandem mass spectrometry; The conditions for HPLC-MS / MS were: High performance liquid chromatography determination: Use C18 ultra-high performance liquid chromatography column, column temperature: 35-45°C; A 3-7% methanol aqueous solution containing 1-3 mmM ammonium acetate was used as mobile phase A, and methanol was used as mobile phase B; Mobile phase flow rate: 0.2-0.4 L / min; elution detection time 45-65 min; Mass spectrometry: A quadrupole-tandem high-resolution orbital ion trap mass spectrometer was used with an electrospray ionization ion source; Spray voltage: -2500V / +3500V; capillary temperature 300~340℃; primary mass spectrometry scanning range is m / z 80 to 1000, resolution is 70000; mass spectrometry collects fragment ions with m / z greater than 50, and each primary scanning window collects secondary fragment ions of the three most abundant primary parent ions; The method for analyzing and detecting phthalates is as follows: by retrieving the material information about PAEs in the MSDIAL software of the mass spectrometer, a PAEs database is constructed, and the sample data is compared based on the PAEs database.

2. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: When collecting soil samples, shallow soil was sampled manually using a Luoyang shovel, and deep soil was sampled using a DP-50 diesel core drill. After collecting soil samples, they were transported to the testing laboratory to ensure that the samples were intact and stored at 4°C.

3. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: Ultrasonic-assisted solid-liquid extraction was used to extract organic pollutants from soil samples. Specifically, the soil samples were weighed and ground evenly, the soil samples were dehydrated with anhydrous sodium sulfate, placed in a centrifuge tube, internal standards were added and aged for 15 to 20 hours, and the samples were ultrasonically extracted with methanol in a water bath at 38 to 42°C for three times using different extraction solvents, and the extracts were combined.

4. The non-targeted identification method of phthalate esters in site soil according to claim 3, characterized in that: The different extraction solvents used in three times were 1:1 v / v dichloromethane-n-hexane, 1:1 v / v n-hexane-acetone, and acetone.

5. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: After extraction by ultrasonic-assisted solid-liquid extraction, the sample solution to be tested is obtained by concentration, volume adjustment and filtration.

6. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: The HPLC determination conditions were as follows: column temperature: 40°C; 5% methanol aqueous solution containing 2 mmM ammonium acetate as mobile phase A, and methanol as mobile phase B.

7. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: The HPLC determination conditions were as follows: mobile phase flow rate: 0.3 L / min; elution detection time: 55 min.

8. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: In HPLC-MS / MS, the injection volume is 4 to 20 μm.

9. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: The mass spectrometry conditions were as follows: capillary temperature: 320°C; sheath gas flow rate: 48 L / h; auxiliary gas flow rate: 11 L / h; and standby gas flow rate: 2 L / h.

10. The non-targeted identification method of phthalate esters in site soil according to claim 1, characterized in that: In the analysis and detection of phthalates, only substances within the 5mDa mass error range and matching 2 or more secondary mass spectrometry fragments with the PAEs database are retained. If the secondary mass spectrometry information of a substance in the PAEs database contains only 1 secondary mass spectrometry fragment, the substance is excluded.

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