A method for detecting 2,4-dimethylphenol and 2,6-dimethylphenol by high-resolution mass spectrometry in chromatographic co-elution and application

Abstract

The application discloses a high-resolution mass spectrometry detection method for chromatographic co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol and application, and relates to the technical field of environmental detection. 1 ) and secondary mass spectrum (MS 2 ) data of standard sequence samples, conversion factor calibration samples and samples are detected and collected by using LC-HRMS under an acetonitrile-5.0-8.0 mmol / L ammonia mobile phase system; according to MS 1 data, the standardized concentrations of co-elution components 2,4-dimethylphenol and 2,6-dimethylphenol and the concentrations of other 11 kinds of phenols and alkyl-substituted phenols are calculated; according to MS 2 data, the concentration of 2,4-dimethylphenol is calculated; according to MS 1 data, the conversion factor of the co-elution components is calculated, and the concentration of 2,6-dimethylphenol is further calculated. The detection limit of the method is lower than 1.0 ug / L, and the sensitivity is 1-2 orders of magnitude higher than that of the standard HJ744-2015 (derivation-GC-MS method) test phenolic compound method.

Description

Technical Field

[0001] This invention relates to the field of environmental monitoring technology, and in particular to a high-resolution mass spectrometry method and its application for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol by chromatography. Background Technology

[0002] Stripping water discharged from sulfur- and saline wastewater stripping units is oily wastewater and is the most significant process wastewater in oil refineries, with a unit discharge volume reaching 0.2 m³. 3 / t crude oil, with a volatile phenol content as high as 0.2~0.5kg / m 3 (Yan Song, Li Lingbo, Han Congbi, Zhou Yanhong. Study on organic components of high-sulfur and high-acid crude oil wastewater [J]. Modern Chemical Industry, 2015, 44(09): 2081-2083). Coal gasification is an important process in modern coal chemical projects. Lurgi pressurized gasification is a relatively mature gasification process. The gas washing wastewater produced by gasification contains high concentrations of recalcitrant organic pollutants such as phenols and ammonia (Li Xueyuan, Wei Danqing. Detection of phenolic compounds in coal gasification wastewater by gas chromatography-mass spectrometry [J]. Modern Chemical Industry, 2015, 35(09): 187-188). The high content of volatile phenolic substances in stripping water and coal gasification wastewater mainly includes phenol, methylphenol (3 isomers) and dimethyl / ethylphenol (9 isomers), which have recycling and reuse value. Clarifying the composition of volatile phenolic substances is the basis for carrying out the recycling and comprehensive utilization of volatile phenolic substances.

[0003] Volatile phenolic components contain numerous isomers. When using chromatography-mass spectrometry (GC-MS) for quantification, chromatographic separation of isomers is necessary, which has always been a technical challenge in this field. Liu Yulong et al. established a method for calculating the content of single-component isomers of dimethylphenol in the same reverse-phase C18 liquid chromatography column under two different mobile phase systems, demonstrating changes in the elution order and co-elution of dimethylphenol isomers. This method enabled the quantitative characterization of the aforementioned 13 alkyl-substituted phenol components in oily wastewater (Liu Yulong, Wang Zixian, Zhang Xiaofei, Shao Zhiguo, Tong Kun. A characterization method and application of C0-C2-alkyl-substituted phenols. Chinese Patent: 202211482228.X; Liu Yulong, Shao Zhiguo, Tong Kun, Zhang Xiaofei, Wang Zixian. A detection method and application of C1-C2-alkyl-substituted phenol isomers. Chinese Patent: 202211482206.3). In existing technologies, whether gas chromatography or liquid chromatography is used, it is rare to find a solution that can accurately quantify single components of chromatographic co-eluent isomers using only one mobile phase system. Summary of the Invention

[0004] The purpose of this invention is to provide a high-resolution chromatographic co-eluent of 2,4-dimethylphenol and 2,6-dimethylphenol.

[0005] The detection method and application employed liquid chromatography-high resolution mass spectrometry (LC-HRMS) in an acetonitrile-ammonia mobile phase system, simultaneously acquiring full scan mass spectrometry (MS1) and selected ion mass spectrometry (MS2). 2 This invention enables the quantitative detection of phenol, three methylphenol isomers, and nine dimethyl / ethylphenol isomers in oily wastewater. Furthermore, it is applicable to the quantitative detection of phenol, three methylphenol isomers, and nine dimethyl / ethylphenol isomers in drinking water, surface water, groundwater, coal gasification wastewater, and oily solid waste.

[0006] To achieve the above objectives, the present invention provides the following technical solution:

[0007] On one hand, the present invention provides a high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol by chromatography, the detection method comprising the following steps:

[0008] Based on the range of petroleum content in the sample, select the appropriate sample pretreatment procedure to prepare the sample;

[0009] In an acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system, standard sequence samples, conversion factor calibration samples, and the aforementioned sample were detected using LC-HRMS. A full-scan mass spectrometry (MS) scan was performed. 1 ) and selected ion secondary mass spectrometry (MS) 2 )data;

[0010] According to MS 1 The data were used to calculate the standardized concentrations of the co-eluting components 2,4-dimethylphenol and 2,6-dimethylphenol, as well as the concentrations of phenol, 2-methylphenol, 3-methylphenol, 4-methylphenol, 2,3-dimethylphenol, 2,5-dimethylphenol, 3,4-dimethylphenol, 3,5-dimethylphenol, 2-ethylphenol, 3-ethylphenol, and 4-ethylphenol using the internal standard calibration method.

[0011] According to MS 2 The concentration of 2,4-dimethylphenol was calculated using 2,4-dimethylphenol-3,5,6-D3 as an internal standard.

[0012] According to MS 1 The data is used to calculate the conversion factor of the co-efferent component, and based on the conversion factor of the co-efferent component, the standardized concentration of the co-efferent component, and the concentration of 2,4-dimethylphenol, the concentration of 2,6-dimethylphenol is calculated.

[0013] As a preferred embodiment, the step of selecting an appropriate sample pretreatment process to prepare the sample based on the petroleum content range in the sample specifically includes:

[0014] When the petroleum content in the sample does not exceed 50 mg / L, the sample shall be filtered through a 0.22 μm filter before being injected for testing.

[0015] When the petroleum content in the sample exceeds 50 mg / L, the sample is alkalized with NaOH and the organic phase is extracted with dichloromethane and n-hexane and then separated by centrifugation. The aqueous phase is then filtered through a 0.22 μm filter membrane, neutralized with formic acid, and then injected for detection.

[0016] As a preferred embodiment, the elution conditions of the acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system include:

[0017] The column flow rate was 0.20–0.25 mL / min;

[0018] Gradient elution is performed as follows: the initial volume ratio of acetonitrile is 20% to 30%, and after maintaining zero gradient elution for 1 min, the elution gradient of acetonitrile is maintained at an isogradient range of 0.43% to 1.56% / min until all components elute;

[0019] The column temperature of the chromatographic column is 35–40℃.

[0020] As a preferred embodiment, the elution conditions of the acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system include:

[0021] The column flow rate was 0.25 mL / min;

[0022] The gradient elution was as follows: 0–1 min, 30% acetonitrile; 1–9 min, 30%–42.5% acetonitrile; 9–9.1 min, 42.5%–90% acetonitrile; 9.1–13 min, 90% acetonitrile; 13–13.1 min, 90%–30% acetonitrile; 13.1–18 min, 30% acetonitrile.

[0023] The column temperature was 40℃.

[0024] As a preferred embodiment, the selected ion secondary mass spectrometry (MS) scan... 2 In the sample, the precursor ion of 2,4-dimethylphenol-3,5,6-D3 is 124.08472 m / z, and the quantitative product ion is 94.07415 m / z; the precursor ion of 2,4-dimethylphenol is 121.06589 m / z, and the quantitative product ion is 91.05532 m / z.

[0025] In a preferred embodiment, the conversion factor for the co-eluent component is calculated using the following formula:

[0026]

[0027] In the formula, k is the conversion factor for the co-efferent component, and A 2,4 / 2,6-DMP To determine the peak areas of the quantitative precursor ions of 2,4-dimethylphenol and 2,6-dimethylphenol, which co-elute at calibration points in the calibration series of samples, A 2,4-DMP To calibrate the peak area of ​​the quantitative precursor ion of 2,4-dimethylphenol in the sample using the corresponding conversion factor, A IS，2,4 / 2,6-DMP and A IS，2,4-DMP The peak area of ​​the corresponding internal standard quantitative precursor ion in the calibration sample is determined by the calibration point and conversion factor of the above calibration sample series.

[0028] In a preferred embodiment, the concentration of 2,6-dimethylphenol is calculated using the following formula:

[0029]

[0030] In the formula, c 2,6-DMP The concentration of 2,6-dimethylphenol is represented by k, which represents the conversion factor for the co-eluting components, and c is the concentration of 2,6-dimethylphenol. STD c represents the normalized concentration of co-efferentiated 2,4-dimethylphenol and 2,6-dimethylphenol. 2,4-DMP This indicates the concentration of 2,4-dimethylphenol.

[0031] On the other hand, the present invention also provides an application of a high-resolution mass spectrometry detection method for chromatographic co-eluent 2,4-dimethylphenol and 2,6-dimethylphenol. The detection method is applied to the quantitative detection of single components of phenol, three methylphenols and nine dimethylphenol isomers in oily wastewater, coal gasification wastewater, oily solid waste, drinking water, surface water and groundwater.

[0032] The technical effects and advantages of this invention are as follows:

[0033] (1) A single injection was used to simultaneously acquire a full scan of a primary mass spectrometer (MS). 1 ) and selected ion secondary mass spectrometry (MS) 2 )

[0034] The data enabled accurate quantification of single components of the chromatographic co-eluting 2,4-dimethylphenol and 2,6-dimethylphenol isomers.

[0035] (2) It was discovered that at a collision energy NCE≥80, only the 2,4-dimethylphenol parent ion has a m / z ratio of 121.06589 ([C8H9O)). - After fragmentation, it produces high-intensity daughter ions (91.05532 m / z [C7H7]). -This daughter ion can be used for the quantification of 2,4-dimethylphenol.

[0036] (3) The sample is not enriched or derivatized. The detection limit of the single component method for phenol, three methylphenols and nine dimethylphenol isomers is less than 1.0 μg / L. Compared with the standard HJ744-2015 (derivative-GC-MS method) for testing phenolic compounds, the sensitivity of this method is 1 to 2 orders of magnitude higher.

[0037] Other features and advantages of the invention will be set forth in the following description, and will be apparent in part from the description, or may be learned by practicing the invention. The objects and other advantages of the invention may be realized and obtained by means of the structures pointed out in the description and the drawings. Attached Figure Description

[0038] Figure 1 This is a flowchart of a high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to the present invention.

[0039] Figure 2 This invention relates to nine dimethyl / ethylphenol [MH] samples eluted under different conditions in an acetonitrile-6.0 mmol / L ammonia mobile phase system, as part of an embodiment of the invention. - Extraction ion map;

[0040] Figure 3 This embodiment of the invention describes the separation of [MH] from nine dimethyl / ethylphenols using an acetonitrile-4.0–10.0 mmol / ammonia mobile phase system. - Extraction ion map. Detailed Implementation

[0041] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0042] To address the shortcomings of existing technologies, this invention discloses a high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol by chromatography. Figure 1 This is a flowchart of a high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to the present invention. Figure 1 As shown, the method includes the following steps:

[0043] Step S1: Based on the range of petroleum content in the sample, select the appropriate sample pretreatment process to prepare the sample;

[0044] Step S2: In an acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system, the standard sequence sample, conversion factor calibration sample, and the sample were detected using LC-HRMS. A full-scan mass spectrometry (MS) scan was performed. 1 ) and selected ion secondary mass

[0045] Spectral scanning (MS) 2 )data;

[0046] Step S3, based on MS 1 The data were used to calculate the standardized concentrations of the co-eluting components 2,4-dimethylphenol and 2,6-dimethylphenol, as well as the concentrations of phenol, 2-methylphenol, 3-methylphenol, 4-methylphenol, 2,3-dimethylphenol, 2,5-dimethylphenol, 3,4-dimethylphenol, 3,5-dimethylphenol, 2-ethylphenol, 3-ethylphenol, and 4-ethylphenol using the internal standard calibration method.

[0047] Step S4, based on MS 2 The concentration of 2,4-dimethylphenol was calculated using 2,4-dimethylphenol-3,5,6-D3 as an internal standard.

[0048] Step S5, based on MS 1 The data is used to calculate the conversion factor of the co-efferent component, and based on the conversion factor of the co-efferent component, the standardized concentration of the co-efferent component, and the concentration of 2,4-dimethylphenol, the concentration of 2,6-dimethylphenol is calculated.

[0049] In step S1 of the present invention, the step of selecting the appropriate sample pretreatment process to prepare the sample according to the range of petroleum content in the sample specifically includes:

[0050] When the petroleum content in the sample does not exceed 50 mg / L, the sample shall be filtered through a 0.22 μm filter before being injected for testing.

[0051] When the petroleum content in the sample exceeds 50 mg / L, the sample is alkalized with NaOH and the organic phase is extracted with dichloromethane and n-hexane and then separated by centrifugation. The aqueous phase is then filtered through a 0.22 μm filter membrane, neutralized with formic acid, and then injected for detection.

[0052] In step S2 of the present invention, the elution conditions of the acetonitrile-5.0-8.0 mmol / L ammonia mobile phase system include:

[0053] The column flow rate was 0.20–0.25 mL / min;

[0054] Gradient elution is performed as follows: the initial volume ratio of acetonitrile is 20% to 30%, and after maintaining zero gradient elution for 1 min, the elution gradient of acetonitrile is maintained at an isogradient range of 0.43% to 1.56% / min until all components elute;

[0055] The column temperature of the chromatographic column is 35–40℃.

[0056] Preferably, the elution conditions of the acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system include:

[0057] The column flow rate was 0.25 mL / min;

[0058] The gradient elution was as follows: 0–1 min, 30% acetonitrile; 1–9 min, 30%–42.5% acetonitrile; 9–9.1 min, 42.5%–90% acetonitrile; 9.1–13 min, 90% acetonitrile; 13–13.1 min, 90%–30% acetonitrile; 13.1–18 min, 30% acetonitrile.

[0059] The column temperature was 40℃.

[0060] In step S4 of the present invention, the selected ion secondary mass spectrometry scan (MS) 2 In the sample, the precursor ion of 2,4-dimethylphenol-3,5,6-D3 is 124.08472 m / z, and the quantitative product ion is 94.07415 m / z; the precursor ion of 2,4-dimethylphenol is 121.06589 m / z, and the quantitative product ion is 91.05532 m / z.

[0061] In step S5 of the present invention, the formula for calculating the conversion factor of the co-eluent component is as follows:

[0062]

[0063] In the formula, k is the conversion factor for the co-efferent component, and A 2,4 / 2,6-DMP To determine the peak areas of the quantitative precursor ions of 2,4-dimethylphenol and 2,6-dimethylphenol, which co-elute at calibration points in the calibration series of samples, A 2,4-DMP To calibrate the peak area of ​​the quantitative precursor ion of 2,4-dimethylphenol in the sample using the corresponding conversion factor, A IS，2,4 / 2,6-DMP and A IS，2,4-DMP The peak area of ​​the corresponding internal standard quantitative precursor ion in the calibration sample is determined by the calibration point and conversion factor of the above calibration sample series.

[0064] The formula for calculating the concentration of 2,6-dimethylphenol is as follows:

[0065]

[0066] In the formula, c2,6-DMP The concentration of 2,6-dimethylphenol is represented by k, which represents the conversion factor for the co-eluting components, and c is the concentration of 2,6-dimethylphenol. STD c represents the normalized concentration of co-efferentiated 2,4-dimethylphenol and 2,6-dimethylphenol. 2,4-DMP This indicates the concentration of 2,4-dimethylphenol.

[0067] Example 1

[0068] This embodiment provides two sample pretreatment methods for LC-HRMS injection testing.

[0069] 1. Stripping water or coal gasification wastewater with a petroleum content ≤50mg / L;

[0070] The water sample was filtered through a 0.22 μm filter membrane. Depending on the volatile phenol content, 10–900 μL of the filtrate was used to prepare the sample.

[0071] This process is also applicable to the preparation of samples of relatively clean water such as drinking water, groundwater and surface water; or the preparation of samples of oil-water mixtures such as oily wastewater.

[0072] 2. Stripping water or coal gasification wastewater with petroleum content > 50 mg / L;

[0073] Take 10.00 mL of water sample into a 20 mL round-bottom screw-top sample bottle (with a PTFE-lined sample cap), add 100 μL of 2.5 mol / L NaOH solution to adjust the pH to ≥12, add 1–3 mL of dichloromethane-n-hexane (2+1) mixed solvent, and tighten the cap. Shake for 30 min and centrifuge. Neutralize the aqueous phase with formic acid, filter the aqueous phase through a 0.22 μm filter membrane, and prepare the sample by taking 10–900 μL of the filtrate, depending on the volatile phenol content.

[0074] Example 2

[0075] In this embodiment, the chromatographic elution conditions for nine types of dimethyl / ethylphenol were optimized in an acetonitrile-ammonia mobile phase system.

[0076] When the chromatographic mobile phase is an acetonitrile-ammonia system, the three methylphenol isomers can be well separated by chromatography. Therefore, the effect of mobile phase elution conditions on the chromatographic separation of dimethyl / ethylphenol was investigated.

[0077] Optimization experiments were conducted by varying parameters such as column flow rate, initial acetonitrile volume ratio, and elution gradient under conditions of column temperature 40℃ and ammonia concentration 6 mmol / L. Gradient elution was initiated after 1 min of initial zero-gradient elution in all optimization experiments. Figure 2 This invention provides examples of the [MH] elution of nine dimethyl / ethylphenols under different elution conditions in an acetonitrile-6.0 mmol / L ammonia mobile phase system. -Extract ion maps, where, Figure 2 Elution conditions for A are: column flow rate 0.20 mL / min; 0–1 min, 20% acetonitrile; 1–24 min, 20%–30% acetonitrile. Elution conditions for B are: column flow rate 0.25 mL / min; 0–1 min, 25% acetonitrile; 1–24 min, 25%–35% acetonitrile. Elution conditions for C are: column flow rate 0.25 mL / min; 0–1 min, 25% acetonitrile; 1–17 min, 25%–50% acetonitrile. The elution conditions for % acetonitrile, D, were: column flow rate 0.25 mL / min; 0–1 min, 30% acetonitrile; 1–17 min, 30%–55% acetonitrile; the corresponding chromatographic peaks were: 1, 3,4-dimethylphenol; 2, 3-ethylphenol; 3, 3,5-dimethylphenol; 4, 2,6-dimethylphenol; 5, 2,3-dimethylphenol; 6, 4-ethylphenol; 7, 2,5-dimethylphenol; 8, 2-ethylphenol; 9, 2,4-dimethylphenol.

[0078] The results showed that, under the conditions of a column flow rate of 0.20 mL / min and an initial volume ratio of 20% acetonitrile, when the elution gradient (defined as the percentage increase rate of acetonitrile in the mobile phase) was 0.43% / min, [the following conditions were met]. Figure 2 As shown in Figure A, three groups of components co-elute: 3,5-dimethylphenol and 3-ethylphenol, 4-ethylphenol and 2,3-dimethylphenol, and 2,5-dimethylphenol and 2,6-dimethylphenol.

[0079] Maintaining the elution gradient at 0.43% / min, the column flow rate was increased to 0.25 mL / min, and the initial acetonitrile volume ratio was increased to 25%. Figure 2 As shown in Figure B, two groups of components co-elute: 4-ethylphenol and 2,3-dimethylphenol, and 2,4-dimethylphenol and 2,6-dimethylphenol.

[0080] Then increase the elution gradient from 0.43% / min to 1.56% / min, such as... Figure 2 As shown in Figure C, only 2,4-dimethylphenol and 2,6-dimethylphenol co-elute.

[0081] Maintain the elution gradient at 1.56% / min, then increase the initial volume percentage of acetonitrile to 30%. Figure 2 As shown in Figure D, the overall chromatographic separation of the nine dimethyl / ethylphenols is better and can be used for quantitative analysis; under these conditions, the chromatographic peak of 2,6-dimethylphenol co-elutes with 2,4-dimethylphenol in the form of a tailing peak.

[0082] Under optimized elution conditions, the effects of column temperatures of 30℃, 35℃, 40℃, and 45℃ on the chromatographic separation of nine dimethyl / ethylphenols were further compared. The results showed that the chromatographic separation effect was better when the column temperature was between 35℃ and 40℃, therefore, the preferred column temperature was 40℃.

[0083] In summary, preferably, the chromatographic elution conditions include: column flow rate of 0.25 mL / min and column temperature of 40 °C; elution sequence: 0–1 min, 30% acetonitrile; 1–9 min, 30%–42.5% acetonitrile; 9–9.1 min, 42.5%–90% acetonitrile; 9.1–13 min, 90% acetonitrile; 13–13.1 min, 90%–30% acetonitrile; 13.1–18 min, 30% acetonitrile.

[0084] Example 3

[0085] In this embodiment, the effect of ammonia concentration in the range of 4.0–10.0 mmol / L on the chromatographic separation of nine dimethyl / ethylphenols was investigated in an acetonitrile-ammonia mobile phase system.

[0086] Figure 3 This embodiment of the invention describes the chromatographic separation of nine dimethyl / ethylphenols under an acetonitrile-4.0–10.0 mmol / ammonia mobile phase system. - The extracted ion chromatograms show that A corresponds to an ammonia concentration of 4.0 mmol / L; B to 5.0 mmol / L; C to 6.0 mmol / L; D to 8.0 mmol / L; and E to 10.0 mmol / L. The corresponding chromatographic peaks are: 1, 3,4-dimethylphenol; 2, 3-ethylphenol; 3, 3,5-dimethylphenol; 4, 2,6-dimethylphenol; 5, 2,3-dimethylphenol; 6, 4-ethylphenol; 7, 2,5-dimethylphenol; 8, 2-ethylphenol; and 9, 2,4-dimethylphenol.

[0087] like Figure 3 As shown in Figure A, the nine dimethyl / ethylphenols separated into seven peaks at an ammonia concentration of 4.0 mmol / L. Among these, two groups of components, 4-ethylphenol and 2,5-dimethylphenol, and 2,4-dimethylphenol and 2,6-dimethylphenol, co-eluted. Figure 3 As shown in Figures B to D, eight peaks were observed when the ammonia concentration was between 5.0 and 8.0 mmol / L. Among these, 2,4-dimethylphenol and 2,6-dimethylphenol co-eluted. Figure 3 As shown in Figure E, when the ammonia concentration is 10.0 mmol / L, only 7 peaks are separated. Among them, 3,5-dimethylphenol and 3-ethylphenol, and 2,4-dimethylphenol and 2,6-dimethylphenol are two groups of components that elute together.

[0088] In summary, the chromatographic separation of the nine dimethyl / ethylphenols was relatively superior when the ammonia concentration was 5.0–8.0 mmol / L, with the best result achieved when the ammonia concentration was 6.0 mmol / L.

[0089] Example 4

[0090] This embodiment provides a method for detecting 13 single phenolic components, including phenol, methylphenol, and dimethyl / ethylphenol, in stripped water using ultra-high performance liquid chromatography-electrostatic field orbital trap high resolution mass spectrometry (UPLC-Orbitrap HRMS).

[0091] 1. Instrument conditions

[0092] UPLC-Orbitrap HRMS: Vanquish C-H10 ultra-high performance liquid chromatograph and Q-Exactive Plus electrostatic field orbit trap high resolution mass spectrometer (ThermoFisher, USA).

[0093] (1) Liquid chromatography conditions

[0094] Mobile phase: Acetonitrile—6.0 mmol / L ammonia. Column temperature: 40℃; Column flow rate: 0.25 mL / min; Sample chamber temperature: 6℃; Injection volume: 2 μL. Gradient elution: 0–1 min, 30% acetonitrile; 1–13 min, 30%–48.75% acetonitrile; 13–13.1 min, 48.75%–30% acetonitrile; 13.1–17 min, 30% acetonitrile.

[0095] (2) Orbitrap HRMS conditions

[0096] Heated electrospray ion source: negative ion mode; sheath gas flow rate: 35 Arb; auxiliary gas flow rate: 10 Arb; spray voltage: 2.6kV; ion transmission tube temperature: 320℃; S-lens RF voltage: 50V; auxiliary gas heater temperature: 380℃.

[0097] Full scan of mass spectrometry (MS) 1 Acquisition parameters: Resolution: 70000; Maximum C-trap ion number capacity (AGCTarget): 1×10⁻⁶ 6 C-trap maximum injection time (Maximum IT): 200ms; Quality range: m / z 90~155;

[0098] Selected ion secondary mass spectrometry (MS) scan 2Acquisition parameters: Selected ion frequencies of 121.06589 m / z and 124.08472 m / z; Resolution: 17500; Maximum C-trap ion count (AGC Target): 1 × 10⁻⁶ 5 C-trap maximum injection time (Maximum IT): 100ms; Isolation window: 1m / z; Normalized collision energy (steppedNCE): 80.

[0099] 2. Preparation of standard sequence samples and conversion factor calibration samples

[0100] Thirteen phenols (phenol, methylphenol, and dimethyl / ethylphenol) with a purity greater than 98% and an internal standard (phenol-) were used. 13 C6 and 2,4-dimethylphenol-3,5,6-D3) standard samples were prepared into single-standard stock solutions. The 13 phenolic mixed working solutions, 2,4-dimethylphenol working solutions, and mixed internal standard working solutions were prepared by stepwise dilution with methanol. The standard sequence samples and conversion factor calibration samples were then prepared using the working solutions.

[0101] Standard series samples: Add 900 μL of pure water to each of six 2 mL sample vials. Using a microsyringe, sequentially add the mixed working solution of 13 phenols and the mixed internal standard working solution. Add methanol to bring the total methanol content in the calibration standard solution to 100 μL. Shake well and store at 4–6 °C until analysis. The concentrations of each component of the 13 phenols in the standard series are 10, 100, 500, 1000, 2500, and 5000 μg / L, respectively, and the concentration of each component of the internal standard is 200 μg / L.

[0102] Conversion factor calibration sample: Add 900 μL of pure water to a 2 mL sample vial. Using a microsyringe, add the 2,4-dimethylphenol working solution and the mixed internal standard working solution sequentially. Add methanol to bring the total methanol content in the calibration standard solution to 100 μL. Shake well and store at 4–6 °C until analysis. The concentration of 2,4-dimethylphenol in the conversion factor calibration sample is 1000 μg / L, and the concentration of each component of the internal standard is 200 μg / L.

[0103] 3. Sample collection and specimen preparation

[0104] Stripping water or coal gasification wastewater samples were collected in 40 mL VOA sample bottles lined with polytetrafluoroethylene (PTFE). The sample bottles were required to be completely filled without overflowing, and no preservatives were added. Samples were refrigerated at 4–6°C, and sample preparation was completed within 48 hours.

[0105] Remove the refrigerated water sample and allow it to return to room temperature before filtering it through a 0.22 μm nylon filter membrane. Add 90 μL of methanol to a 2 mL sample vial beforehand. Depending on the volatile phenol content, add 10–900 μL of the filtrate to the sample vial, then add pure water to bring the total sample volume to 990 μL. Add 10 μL of the mixed internal standard working solution, shake well, and store at 4–6 °C until analysis.

[0106] 4. Data Collection and Result Calculation

[0107] Data was acquired using Xcalibur 4.4 software and analyzed using TraceFinder 5.1 software. The target compound in the sample was qualitatively identified based on retention time matching and the parent ion [MH]. - The concentration of the target compound can be calculated from the peak area of ​​the ordinal ion.

[0108] Table 1 shows the retention times, quantitative parent ions, and quantitative daughter ions for the mixed internal standard and 13 phenolic components. The effects of different collision energies on the secondary mass spectrometry fingerprint characteristics of dimethylphenol and 2,6-dimethylphenol were investigated in an acetonitrile-6.0 mmol / L ammonia mobile phase system. The study found that at lower collision energies (NCE ≤ 50), 2,4-dimethylphenol...

[0109] Both methylphenol and 2,6-dimethylphenol produce the same characteristic daughter ion 93.03459 m / z ([C6H5O]) upon fragmentation. - Other daughter ions have very low relative strengths to the parent ion and can be ignored; increasing the collision energy (NCE≥80) is crucial, especially for the 2,4-dimethylphenol parent ion with a m / z of 121.06589 ([C8H9O)). - After fragmentation, it produces 91.05532 m / z ([C7H7]). - The daughter ions of 2,4-dimethylphenol have relatively high intensities and can be used for the quantitative determination of 2,4-dimethylphenol. Interfering ions at 91.01899 m / z and 90.98577 m / z were also found. For low-resolution mass spectrometry (such as triple quadrupole mass spectrometers), the 91 m / z interference cannot be distinguished, while high-resolution mass spectrometry (HRMS) has a relative error better than 5 × 10⁻⁶ m / z. -6 This can effectively eliminate such interference.

[0110] Table 1. Retention time, quantitative parent ion, and quantitative daughter ion of the mixed internal standard and 13 phenolic components.

[0111]

[0112] At each calibration point, with equal concentrations of 2,4-dimethylphenol (2,4-DMP) and 2,6-dimethylphenol (2,6-DMP), a calibration curve was plotted with the sum of the concentrations of 2,4-dimethylphenol and 2,6-dimethylphenol as the independent variable and the sum of their responses (e.g., peak areas of the quantitative ion) as the dependent variable.

[0113] The concentration values ​​of the co-eluting components calculated from the calibration curve are defined as the standardized concentrations of co-eluting 2,4-dimethylphenol and 2,6-dimethylphenol, denoted by c. STD express.

[0114] The conversion factor for 2,6-dimethylphenol to 2,4-dimethylphenol is represented by k and calculated according to formula (1):

[0115]

[0116] In the formula, A 2,4 / 2,6-DMP To determine the peak areas of the quantitative precursor ions of 2,4-dimethylphenol and 2,6-dimethylphenol, which are co-eluted components, at calibration points (e.g., 1000 μg / L) in a calibration series of samples, A 2,4-DMP To calibrate the peak area of ​​the quantitative precursor ion of 2,4-dimethylphenol in the corresponding conversion factor sample (e.g., 1000 μg / L), A IS，2,4 / 2,6-DMP and A IS，2,4-DMP The peak area of ​​the corresponding internal standard quantitative precursor ion in the calibration sample for the calibration point and conversion factor of the above calibration sample series; wherein, the relevant calibration samples of the conversion factor can be repeatedly measured, and the average value is taken as k.

[0117] For MS 1 The data was processed, and the peak area of ​​the parent ion was used to calculate the standardized concentrations (c) of the co-eluting 2,4-dimethylphenol and 2,6-dimethylphenol using the internal standard method. STD The concentrations of phenol, 2-methylphenol, 3-methylphenol, 4-methylphenol, 2,3-dimethylphenol, 2,5-dimethylphenol, 3,4-dimethylphenol, 3,5-dimethylphenol, 2-ethylphenol, 3-ethylphenol and 4-ethylphenol;

[0118] For MS 2 The data was processed, and the peak area of ​​the quantitative ion was used to calculate the concentration of 2,4-dimethylphenol in the sample using the internal standard method, denoted as c. 2,4-DMP .

[0119] The concentration of 2,6-dimethylphenol in the sample, c 2,6-DMP Calculate according to formula (2):

[0120]

[0121] 5. Method performance indicators

[0122] (1) Calibration range and detection limit

[0123] The first regression correlation coefficient (R) of 13 phenolic components in the range of 10–5000 μg / L 2 All values ​​were greater than 0.997. The detection limits for all 13 phenolic components were ≤1.0 μg / L. Compared with the standard HJ744-2015 (derivative-GC-MS method) for testing phenolic compounds, this method has a sensitivity that is 1 to 2 orders of magnitude higher.

[0124] (2) Sample matrix spike recovery and precision

[0125] In the spiking of stripped water sample matrix, the concentrations of 13 phenolic compounds were 10, 500, and 2000 μg / L. The recoveries of the target analytes ranged from 77% to 117.8%, with RSDs ranging from 0.4% to 10.2%. This indicates that the method is accurate and reproducible.

[0126] It should be noted that this embodiment is merely a preferred example. This invention does not limit the type and specifications of the chromatographic column or the type of LC-HRMS instrument. Other similar reverse-phase chromatographic columns can be used to implement this invention based on the technical solutions disclosed herein.

[0127] Finally, it should be noted that the above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol by chromatographic extraction, characterized in that, The detection method includes the following steps: preparing the sample by selecting the appropriate sample pretreatment process according to the range of petroleum content in the sample; In an acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system, standard sequence samples, conversion factor calibration samples, and the aforementioned sample were detected using LC-HRMS, with full-scan MS data acquired via primary mass spectrometry. 1 Selected ion secondary mass spectrometry scanning MS 2 data; According to MS 1 The data were used to calculate the standardized concentrations of the co-eluting components 2,4-dimethylphenol and 2,6-dimethylphenol, as well as the concentrations of phenol, 2-methylphenol, 3-methylphenol, 4-methylphenol, 2,3-dimethylphenol, 2,5-dimethylphenol, 3,4-dimethylphenol, 3,5-dimethylphenol, 2-ethylphenol, 3-ethylphenol, and 4-ethylphenol using the internal standard calibration method. According to MS 2 The concentration of 2,4-dimethylphenol was calculated using 2,4-dimethylphenol-3,5,6-D3 as an internal standard. According to MS 1 The data is used to calculate the conversion factor of the co-efferent component, and the concentration of 2,6-dimethylphenol is calculated based on the conversion factor of the co-efferent component, the standardized concentration of the co-efferent component, and the concentration of 2,4-dimethylphenol. The formula for calculating the conversion factor of the co-eluent component is as follows: In the formula, k is the conversion factor for the co-efferent component, and A 2,4 / 2,6-DMP To determine the peak areas of the quantitative precursor ions of 2,4-dimethylphenol and 2,6-dimethylphenol, which co-elute at calibration points in the calibration series of samples, A 2,4-DMP To calibrate the peak area of ​​the quantitative precursor ion of 2,4-dimethylphenol in the sample using the corresponding conversion factor, A IS，2,4 / 2,6-DMP and A IS，2,4-DMP The peak area of ​​the corresponding internal standard quantitative precursor ion in the calibration sample is determined by the calibration points and conversion factors of the above calibration sample series. The formula for calculating the concentration of 2,6-dimethylphenol is as follows: In the formula, c 2,6-DMP The concentration of 2,6-dimethylphenol is represented by k, which represents the conversion factor for the co-eluting components, and c is the concentration of 2,6-dimethylphenol. STD c represents the normalized concentration of co-efferentiated 2,4-dimethylphenol and 2,6-dimethylphenol. 2,4-DMP This indicates the concentration of 2,4-dimethylphenol.

2. The high-resolution mass spectrometry detection method for chromatographic co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to claim 1, characterized in that, The preparation of samples by selecting the appropriate sample pretreatment process based on the petroleum content range in the sample specifically includes: When the petroleum content in the sample does not exceed 50 mg / L, the sample shall be filtered through a 0.22 μm filter before being injected for testing. When the petroleum content in the sample exceeds 50 mg / L, the sample is alkalized with NaOH and the organic phase is extracted with dichloromethane and n-hexane and then separated by centrifugation. The aqueous phase is then filtered through a 0.22 μm filter membrane, neutralized with formic acid, and then injected for detection.

3. A high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to claim 1 or 2, characterized in that, The elution conditions for the acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system include: The column flow rate was 0.20–0.25 mL / min; Gradient elution was performed as follows: the initial volume ratio of acetonitrile was 20%–30%, and after maintaining zero gradient elution for 1 min, the elution gradient of acetonitrile was maintained at an isogradient range of 0.43–1.56% / min until all components eluted; the column temperature was 35–40℃.

4. The high-resolution mass spectrometry detection method for chromatographic co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to claim 1, characterized in that, The elution conditions for the acetonitrile-5.0–8.0 mmol / L ammonia mobile phase system include: The column flow rate was 0.25 mL / min; The gradient elution was as follows: 0–1 min, 30% acetonitrile; 1–9 min, 30%–42.5% acetonitrile; 9–9.1 min, 42.5%–90% acetonitrile; 9.1–13 min, 90% acetonitrile; 13–13.1 min, 90%–30% acetonitrile; 13.1–18 min, 30% acetonitrile. The column temperature was 40℃.

5. The high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol according to claim 1, characterized in that, Selected ion secondary mass spectrometry scanning MS 2 In the study, the precursor ion of 2,4-dimethylphenol-3,5,6-D3 was 124.08472 m / z, and the quantitative product ion was 94.07415 m / z; the precursor ion of 2,4-dimethylphenol was 121.06589 m / z. The quantitative ion was 91.05532 m / z.

6. The application of a high-resolution mass spectrometry method for the co-elution of 2,4-dimethylphenol and 2,6-dimethylphenol as described in any one of claims 1-5, characterized in that, The detection method is applied to the quantitative detection of single components of phenol, three methylphenols and nine dimethylphenol isomers in oily wastewater, coal gasification wastewater, oily solid waste, drinking water, surface water and groundwater.

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