Quantitative determination method for tris (2, 4-di-tert-butylphenyl) phosphate

Abstract

The invention relates to a quantitative determination method of tris (2, 4-di-tert-butylphenyl) phosphate, which comprises the following steps: S1, preparing a plurality of samples to be determined; s2, preparing a standard stock solution from tris (2, 4-di-tert-butylphenyl) phosphate; s3, preparing a standard working solution by diluting the standard stock solution; s4, respectively loading samples to be detected into centrifugal tubes, adding a small amount of ethyl acetate for multiple times, carrying out vortex extraction and centrifugation, and taking supernate; s5, nitrogen-blowing the supernatant obtained in the step S4 to be nearly dry, adding ethyl acetate for redissolving, uniformly mixing, and passing through a membrane to obtain a to-be-analyzed solution; s6, placing the standard working solution and the to-be-analyzed solution in a gas chromatograph-mass spectrometer for chromatography and mass spectrometry detection analysis, and screening out a to-be-determined sample containing a target object pair; s7, taking a to-be-determined sample containing the target object pair, and repeating the steps S4-S5 to obtain a to-be-determined liquid; s8, preparing a series of working solutions by diluting the standard stock solution; and S9, putting the series of working solutions and the to-be-determined solution into a gas chromatograph-mass spectrometer for chromatography and mass spectrometry detection.

Description

Technical Field

[0001] This invention relates to the field of food and pharmaceutical component detection technology, and to a method for the quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate. Background Technology

[0002] Tris(2,4-di-tert-butylphenyl) phosphate, as a class of organophosphate compounds with significant activity, may exert anti-inflammatory effects by inhibiting secretory phospholipase A2 (sPLA2), a key mediator of inflammatory responses. Therefore, the precise detection of tris(2,4-di-tert-butylphenyl) phosphate shows potential application value in food science, medicinal chemistry, and other fields, and is of significant research importance.

[0003] However, existing detection technologies have significant gaps and limitations, making it difficult to meet the current demand for high-precision and interference-resistant analysis. Technical challenges exist, such as insufficient research on sample pretreatment methods. Relying on traditional adsorption and filtration methods fails to effectively purify the matrix, interfering with the detection of target analytes. Therefore, developing an analytical method that can overcome matrix effects, possess both high sensitivity and high selectivity, and accurately quantify tris(2,4-di-tert-butylphenyl) phosphate has become a crucial technological gap that urgently needs to be filled in this field. Summary of the Invention

[0004] To address the above technical problems, this invention proposes a quantitative determination method for tris(2,4-di-tert-butylphenyl) phosphate. The method employs gas chromatography-mass spectrometry (GC-MS) to rapidly screen the sample for identification and quantitative analysis of the target analyte. By comparing the GC-MS detection results of the target sample with those of a series of working solutions, the content of the target analyte in the target sample is determined.

[0005] A method for the quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate, including preliminary screening and quantitative analysis, specifically includes the following steps: Initial screening: S1: Prepare several samples to be tested; S2: Prepare a standard stock solution from tris(2,4-di-tert-butylphenyl) phosphate; S3: Prepare standard working solution by diluting standard stock solution; S4: Take the samples to be tested and put them into centrifuge tubes, add ethyl acetate, vortex extract, centrifuge, and take the supernatant; S5: Blow the supernatant from S4 to near dryness with nitrogen, add ethyl acetate to redissolve, mix well and pass through a membrane to obtain the solution to be analyzed; S6: Place the standard working solution and the solution to be analyzed in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric analysis to screen out the solution to be analyzed containing tris(2,4-di-tert-butylphenyl) phosphate, and the corresponding sample to be tested is the target sample; Quantitative analysis: S7: Take the target sample and follow steps S4~S5 to obtain the solution to be tested; S8: A series of working solutions were prepared by diluting the standard stock solution. The tris(2,4-di-tert-butylphenyl) phosphate concentrations of the series of working solutions were 0.2, 0.5, 1, 2 and 5 mg / L, respectively. S9: Place the series of working solutions and the solution to be tested in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric detection, and compare the detection results of the series of working solutions and the solution to be tested to quantitatively analyze the content of tris(2,4-di-tert-butylphenyl) phosphate in the target sample.

[0006] Furthermore, in step S2, the standard stock solution is an ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate at a concentration of 100 mg / L, and in step S3, the standard working solution is an ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate at a concentration of 5 mg / L.

[0007] Furthermore, the chromatographic conditions for the gas chromatography-mass spectrometry (GC-MS) in step S6 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: Electron impact ion source (EI); Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: Full scan 15 m / z - 550 m / z.

[0008] Furthermore, the chromatographic conditions for the gas chromatography-mass spectrometry (GC-MS) in step S9 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: electron impact ion source; Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: ion scan; Quantitative ion: 316; Qualitative ions: 207, 57, 191.

[0009] Furthermore, step S4 specifically involves: Take the sample to be tested and put it into a centrifuge tube. Add ethyl acetate, vortex extract for 3 min, centrifuge at 8500 r / min for 3 min, take the supernatant, repeat the extraction twice for the same sample to be tested and combine the supernatants.

[0010] Furthermore, in step S4, the volume ratio of the sample to be tested to ethyl acetate is 3:1.

[0011] Compared with the prior art, the beneficial effects achieved by the present invention are as follows: 1. This invention employs gas chromatography-mass spectrometry (GC-MS) for rapid screening of tris(2,4-di-tert-butylphenyl) phosphate, and simultaneously uses GC-MS for quantitative analysis.

[0012] 2. This invention employs gas chromatography-mass spectrometry (GC-MS) to determine tris(2,4-di-tert-butylphenyl) phosphate, achieving good peak shape and high resolution. Under the chromatographic conditions, the concentration and peak area exhibit a good linear relationship, with recoveries all exceeding 80%. Attached Figure Description

[0013] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0014] Figure 1 This is a full scan matching diagram of tris(2,4-di-tert-butylphenyl) phosphate in this invention.

[0015] Figure 2 This is a selected ion chromatogram of tris(2,4-di-tert-butylphenyl) phosphate in this invention. Detailed Implementation

[0016] To further illustrate the technical means and effects of the present invention in achieving its intended purpose, the following detailed description of the specific implementation methods, structures, features, and effects of the present invention, in conjunction with the accompanying drawings and preferred embodiments, is provided.

[0017] Please see Figures 1-2A method for the quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate includes two main parts: preliminary screening of target samples and quantitative analysis of the target analyte (tris(2,4-di-tert-butylphenyl) phosphate). The initial screening includes the following steps: S1: Prepare several samples to be tested; S2: Prepare a 100 mg / L ethyl acetate solution from tris(2,4-di-tert-butylphenyl) phosphate as a standard stock solution; S3: Prepare a 5 mg / L ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate by diluting the standard stock solution to serve as the standard working solution; S4: Take 20-25 ml of the sample to be tested and put them into centrifuge tubes. Add ethyl acetate in small amounts several times, for a total of 8 ml. Vortex extract for 3 min, centrifuge at 8500 r / min for 3 min, take the supernatant, and repeat the extraction once more. Combine the two supernatants. S5: Blow the supernatant from S4 to near dryness with nitrogen, add 2 ml of ethyl acetate to redissolve, mix well and pass through a membrane to obtain the solution to be analyzed; S6: Place the standard working solution and the solution to be analyzed in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric analysis to screen out the solution to be analyzed containing tris(2,4-di-tert-butylphenyl) phosphate. The corresponding sample to be measured is the target sample.

[0018] The chromatographic conditions for gas chromatography-mass spectrometry in step S6 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: Electron impact ion source (EI); Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: Full scan 15 m / z - 550 m / z.

[0019] By utilizing the target analyte database in the NIST standard library of the software, and simultaneously matching the data of the screened samples with the target analytes in the database, it is possible to quickly determine whether a sample contains the target analyte. Figure 1 The full-scan ion map can obtain the main characteristic ion fragment information of the target object, and quickly screen to determine whether the target object is present.

[0020] Quantitative analysis includes the following steps: S7: Take the target sample and follow steps S4~S5 to obtain the solution to be tested; S8: A series of working solutions were prepared by diluting the standard stock solution. The tris(2,4-di-tert-butylphenyl) phosphate concentrations of the series of working solutions were 0.2, 0.5, 1, 2 and 5 mg / L, respectively. S9: Place the series of working solutions and the solution to be tested in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric detection, and compare the detection results of the series of working solutions and the solution to be tested to quantitatively analyze the content of tris(2,4-di-tert-butylphenyl) phosphate in the target sample.

[0021] The chromatographic conditions for the gas chromatography-mass spectrometry (GC-MS) instrument in step S9 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: Electron impact ion source (EI); Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: Ion scan; Quantitative ion: 316; Qualitative ions: 207, 57, 191.

[0022] The target sample was quantitatively analyzed using the above chromatographic-mass spectrometric conditions, and the selected ion chromatogram of tris(2,4-di-tert-butylphenyl) phosphate was obtained as follows: Figure 2 As shown, the horizontal axis represents retention time (min), and the vertical axis represents ion abundance. Figure 2 Selective ion chromatograms can accurately determine the retention time of target compounds, facilitating quantitative analysis.

[0023] This invention provides a method for the quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate. Gas chromatography-mass spectrometry (GC-MS) is used to rapidly screen the sample to identify the target sample and perform quantitative analysis of the target analyte. By comparing the GC-MS detection results of the target sample with those of a series of working solutions, the content of the target analyte in the target sample is determined. The feasibility of this method will be further verified below: (1) Linear equation, correlation coefficient and limit of quantitation Prepare a series of working solutions according to step S8, and inject and determine them according to chromatographic and mass spectrometric conditions. The linear range was investigated, and a standard curve was plotted with the injection amount (X, μg) as the abscissa and the corresponding peak area (Y) as the ordinate. The linear equation, linear range, correlation coefficient, limit of detection, and limit of quantitation were calculated. The limit of detection (RSN=3) and limit of quantitation (RSN=10) of the target analyte were determined according to the signal-to-noise ratio. The results are shown in Table 2.

[0024] (2) Method repeatability Six blank samples (20g each) were taken and mixed with 0.09ml of a 100mg / L ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate. Extraction and analysis were performed using the extraction method, chromatographic conditions, and mass spectrometry conditions described in this invention. The average content of tris(2,4-di-tert-butylphenyl) phosphate was determined to be 0.44mg / kg. The RSD was 7.67%, indicating that the test method of this invention has good repeatability.

[0025] (3) Instrument stability A 1 mg / L ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate was repeatedly detected 6 times using gas chromatography-mass spectrometry. The changes in the peak area of ​​the target analyte were observed and are shown in Table 3. The relative standard deviation (RSD) of the peak area of ​​the target analyte was 1.8%.

[0026] (4) Recovery and precision of the method Spiked recovery experiments were conducted on low-background samples in S8. Six samples were independently weighed and spiked for determination. The recoveries and relative standard deviations were then calculated, and the results are shown in Table 4. The recoveries ranged from 81.51% to 102.88%, and the RSDs ranged from 1.50% to 3.44%.

[0027] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or alterations to the above-disclosed technical content to create equivalent embodiments without departing from the scope of the present invention. Any simple modifications, equivalent changes and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the scope of the present invention.

Claims

1. A method for the quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate, characterized in that, This includes preliminary screening and quantitative analysis, specifically the following steps: Initial screening: S1: Prepare several samples to be tested; S2: Prepare a standard stock solution from tris(2,4-di-tert-butylphenyl) phosphate; S3: Prepare standard working solution by diluting standard stock solution; S4: Take the samples to be tested and put them into centrifuge tubes, add ethyl acetate, vortex extract, centrifuge, and take the supernatant; S5: Blow the supernatant from S4 to near dryness with nitrogen, add ethyl acetate to redissolve, mix well and pass through a membrane to obtain the solution to be analyzed; S6: Place the standard working solution and the solution to be analyzed in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric analysis to screen out the solution to be analyzed containing tris(2,4-di-tert-butylphenyl) phosphate, and the corresponding sample to be tested is the target sample; Quantitative analysis: S7: Take the target sample and follow steps S4~S5 to obtain the solution to be tested; S8: A series of working solutions were prepared by diluting the standard stock solution. The tris(2,4-di-tert-butylphenyl) phosphate concentrations of the series of working solutions were 0.2, 0.5, 1, 2 and 5 mg / L, respectively. S9: Place the series of working solutions and the solution to be tested in a gas chromatograph-mass spectrometer for chromatographic and mass spectrometric detection, and compare the detection results of the series of working solutions and the solution to be tested to quantitatively analyze the content of tris(2,4-di-tert-butylphenyl) phosphate in the target sample.

2. The method for quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate according to claim 1, characterized in that, In step S2, the standard stock solution is a 100 mg / L ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate, and in step S3, the standard working solution is a 5 mg / L ethyl acetate solution of tris(2,4-di-tert-butylphenyl) phosphate.

3. The method for quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate according to claim 1, characterized in that, The chromatographic conditions for gas chromatography-mass spectrometry in step S6 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: Electron impact ion source (EI); Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: Full scan 15 m / z - 550 m / z.

4. The method for quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate according to claim 1, characterized in that, The chromatographic conditions for the gas chromatography-mass spectrometry (GC-MS) instrument in step S9 are as follows: Column: DB-5, 30 m * 0.25 mm * 0.25 μm; Injector temperature: 300℃; Split mode: Splitless; Column flow rate: 1 ml / min; Temperature program: Initial temperature: 150℃, hold for 5 min, increase to 270℃ at a rate of 20℃ / min, hold for 20 min; Mass spectrometry conditions are: Solvent delay: 1 min; Ion source: Electron impact ion source; Electron energy: 70 eV; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Mass spectrometry interface temperature: 280℃; Scan mode: Ion scan; The quantitative ion was 316, and the qualitative ions were 207, 57, and 191.

5. The method for quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate according to any one of claims 1 to 4, characterized in that, Step S4 is as follows: Take the sample to be tested and put it into a centrifuge tube. Add ethyl acetate, vortex extract for 3 min, centrifuge at 8500 r / min for 3 min, take the supernatant, repeat the extraction twice for the same sample to be tested and combine the supernatants.

6. The method for quantitative determination of tris(2,4-di-tert-butylphenyl) phosphate according to any one of claims 1 to 4, characterized in that, In step S4, the volume ratio of the sample to be tested to ethyl acetate is 3:1.

Images