A high-throughput method for the identification of carbonyl compounds in environmental matrices

Abstract

This invention discloses a high-throughput identification method for carbonyl compounds in environmental matrices, comprising: extracting carbonyl compounds from a sample of the environmental matrix to be tested, removing the solvent from the extract to obtain a concentrated solution of the carbonyl compounds; adding a stable isotope-labeled reagent to the concentrated solution of the carbonyl compounds to perform a derivatization reaction, terminating the reaction, drying, and reconstituted to obtain a derivatized sample; performing ultra-high resolution mass spectrometry analysis on the derivatized sample to obtain raw ultra-high resolution mass spectrometry data; screening peak pair information of products containing stable isotope-labeled products from the raw ultra-high resolution mass spectrometry data, obtaining the elemental molecular composition information of the carbonyl compounds labeled with stable isotope-labeled products based on the peak pair information through elemental molecular formula matching, and performing molecular-level characterization of the corresponding carbonyl compounds in the environmental matrix. The method of this invention can achieve high-throughput identification of unknown carbonyl compounds in environmental matrices.

Description

Technical Field

[0001] This invention relates to the field of environmental technology, and in particular to a high-throughput method for identifying carbonyl compounds in environmental matrices. Background Technology

[0002] Carbonyl compounds are a class of compounds containing a carbonyl group in their molecules, including aldehydes and ketones, and are ubiquitous in the atmospheric environment. The main sources of carbonyl compounds in the environment are thermal processes such as the incomplete combustion of fossil fuels or biomass fuels. They can also be generated through oxidation or photochemical reactions from volatile or semi-volatile precursors of alkanes, alkenes (such as terpenes), and polycyclic aromatic hydrocarbons from biological or anthropogenic sources. Carbonyl compounds are also present in the atmosphere (e.g., PM2.5). 2.5 It is widely distributed and accounts for a significant proportion in environmental media such as automobile exhaust, industrial smoke, and indoor dust.

[0003] Carbonyl compounds can participate in atmospheric photochemical reactions, promoting the formation of fine particulate matter and leading to regional or localized air pollution. Typically, carbonyl compounds exhibit biotoxicity and negative health effects, such as genotoxicity through covalent binding to DNA nucleophilic sites, inducing cellular oxidative stress leading to cell and tissue inflammation, and increasing the burden of respiratory and cardiovascular diseases, thus harming both biological and human health. Therefore, identifying carbonyl compounds in the environmental matrix is ​​crucial for understanding the reaction mechanisms of carbonyl compound formation, potential sources, environmental health risk assessment, and governance and control.

[0004] Currently, most sampling and analysis of carbonyl compounds relies on manual sampling and offline analysis, such as liquid chromatography and gas chromatography-mass spectrometry (HPLC / GC-MS). For example, Chinese patent document CN104807899B discloses an online analysis device for volatile carbonyl compounds. This device collects and thermally desorbs volatile carbonyl compound samples. The thermally desorbed sample is then fed into a gas chromatograph / mass spectrometer through a fused silica tube for the separation and quantification of carbonyl compounds.

[0005] Conventional mass spectrometry techniques (such as HPLC / GC-MS) are widely used for the qualitative identification and quantitative detection of carbonyl compounds. However, large-scale environmental analysis of unknown carbonyl compounds using reference standards requires significant manpower, resources, and time for mass spectrometry analysis and massive peak labeling. Furthermore, carbonyl compounds exist only at trace levels in complex environmental matrices such as the atmosphere, which further complicates their accurate identification and analysis.

[0006] High-resolution mass spectrometry techniques (such as orbital trap mass spectrometry and Fourier transform ion cyclotron resonance mass spectrometry) have received increasing attention and application due to their higher mass resolution, accuracy, and sensitivity. They have already successfully achieved the measurement of PM2.5.2.5 Non-targeted identification of trace organic pollutants in the middle.

[0007] It is worth noting that carbonyl compounds have low ionization efficiency and poor stability, resulting in insufficient sensitivity and low response efficiency in high-resolution mass spectrometry detection, which is not conducive to the high-precision and high-efficiency identification of unknown carbonyl compounds in complex environmental matrices. Summary of the Invention

[0008] This invention provides a high-throughput identification method for carbonyl compounds in an environmental matrix, enabling high-throughput identification of unknown carbonyl compounds in an environmental matrix.

[0009] The technical solution of the present invention is as follows:

[0010] A high-throughput identification method for carbonyl compounds in an environmental matrix includes the following steps:

[0011] (1) Extract carbonyl compounds from the environmental matrix sample to be tested, and remove the solvent from the extract to obtain a concentrated carbonyl compound solution;

[0012] (2) Add stable isotope labeling reagents to the concentrated carbonyl compound solution to carry out derivatization reaction, terminate the reaction, dry and reconstitute to obtain derivatized sample; the stable isotope labeling reagents are Girard reagent P (d0-GRP) and deuterated Girard reagent P (d5-GRP).

[0013] (3) Perform ultra-high resolution mass spectrometry analysis on the derivatized samples to obtain the original ultra-high resolution mass spectrometry data;

[0014] (4) Screen out the peak pair information of the products labeled with stable isotope pairs from the original ultra-high resolution mass spectrometry data, obtain the elemental molecular composition information of carbonyl compounds labeled with stable isotope pairs by matching the elemental molecular formula based on the peak pair information, and characterize the carbonyl compounds in the corresponding environmental matrix at the molecular level.

[0015] Characteristic peaks of carbonyl compounds are screened from ultra-high resolution samples based on the precise mass difference between isotope pairs of carbonyl compounds and the abundance ratio of mass spectrometry response. Non-target analysis methods are then used to identify carbonyl compounds.

[0016] The high-throughput identification method of this invention can be applied to complex environmental samples such as atmospheric fine particulate matter and indoor dust to achieve high-throughput identification of unknown carbonyl compounds, providing technical support for risk assessment and priority control of pollutants containing carbonyl functional groups.

[0017] Preferably, in step (1), when extracting carbonyl compounds from the environmental matrix sample to be tested, the extraction solvent used is acetonitrile or a mixed solution of acetonitrile and water.

[0018] The abbreviation for Girard's reagent P is d0-GRP, CAS: 1126-58-5; the abbreviation for deuterated Girard's reagent P is d5-GRP, CAS: 1505505-87-2.

[0019] Preferably, in the stable isotope pair labeling reagent, the molar ratio of Girard reagent P to deuterated Girard reagent P is 1:1.

[0020] Preferably, in step (2), the concentration of the stable isotope labeling reagent in the derivatization reaction system is 0.01-10 mmol / L; the molar ratio of the stable isotope labeling reagent to the carbonyl compound is (10-1000):1.

[0021] Preferably, in step (2), hydrochloric acid is used to adjust the derivatization reaction system to acidity, and the concentration of HCl in the derivatization reaction system is 0.01-10 mmol / L.

[0022] Preferably, in step (2), the derivatization reaction is carried out at room temperature in the dark for 5-60 minutes.

[0023] More preferably, in step (2), the concentration of the stable isotope labeling reagent in the derivatization reaction system is 0.01-10 mmol / L; the concentration of HCl is 0.01-10 mmol / L; and the derivatization reaction time is 5-60 min.

[0024] Preferably, in step (2), the reaction is terminated at -20°C.

[0025] Preferably, in step (2), the solvent used for resolution is acetonitrile or a mixture of acetonitrile and water.

[0026] More preferably, in the mixed solution of acetonitrile and water, the volume ratio of acetonitrile to water is (8-9.5):1.

[0027] Preferably, in step (3), a Fourier transform ion cyclotron resonance mass spectrometer is used to perform ultra-high resolution mass spectrometry analysis on the derivatized sample. The scanning mode is full ion scan in electrospray ionization positive ion mode, and the full scan mass range is 120-900 m / z to obtain ultra-high resolution mass spectrometry data.

[0028] Preferably, step (3) further includes conducting field blank, procedural blank and solvent blank tests, and using the carbonyl compounds detected in the field blank, procedural blank and solvent blank as background carbonyl compounds.

[0029] Field blank refers to: taking particulate matter sampling as an example, placing the sampling membrane for 22-24 hours when the sampler is not working to monitor background interference during the sampling process; procedural blank refers to: performing a complete pretreatment process on the blank sampling membrane to monitor background interference during the pretreatment process; solvent blank refers to: injecting pure solvent for analysis to monitor background interference caused by instrument residue.

[0030] Preferably, step (4) includes:

[0031] (4-1) Perform mass spectrometry peak detection and identification on the raw ultra-high resolution mass spectrometry data (peak response IR>1×10). 6 (S / N≥3), remove isotope peaks and adduct ion peaks, perform internal standard correction, and obtain the peak list;

[0032] (4-2) Select target peak pairs from the peak list to generate a list of potential carbonyl compound light and heavy isotope labeled derivative pairs (M p1 M p2 M p1 <M p2 M p1 M p2 These represent the mass-to-charge ratio of the target peak pair and the characteristic mass distance M of the target peak pair, respectively. p2 -M p1 The target peak-to-response ratio (IR) is 5.03138 ± 0.00015 Da. P1 / IR P2 Between 0.33 and 3, IR P1 IR P2 These represent the response intensities of the target peak pairs;

[0033] (4-3) For M p1 The exact mass number is used to match the elemental molecular formula, and the number of elements in the molecular formula is limited to 1. 12 C is 1~100, 1 H is 1~200, 16 O is 0~50, 15 N is 3~6 32 S is 0~1, and the absolute value of molecular formula matching error is <1ppm;

[0034] After deducting the C7H8N3 portion of the derivatizing reagent, a list of molecular formulas for potential carbonyl candidates was obtained;

[0035] (4-4) Screening out the molecular formulas of reliable carbonyl candidates from the list of molecular formulas of potential carbonyl candidates through the rule constraints on the number of atoms of elements in the molecular formula; the rule constraints are 0.3 ≤ H / C ≤ 2.25, 0 < O / C ≤ 1.2, 0 ≤ N ≤ 3, and 0 < DBE / C ≤ 1, where H, C, O, and N are the number of atoms of elements hydrogen, carbon, oxygen, and nitrogen in the molecular formula, and DBE is the number of unsaturated double bonds.

[0036] The calculation formula for the number of unsaturated double bonds is DBE = 1 + C - 0.5H + 0.5N.

[0037] Further preferably, in step (4-1), the hydrazone derivatives of background carbonyl compounds are used for in-spectrum internal standard correction.

[0038] Further preferably, step (4-1) also includes background subtraction from the peak list to remove interfering mass spectrometry peaks that coincide with background carbonyl compounds (IR sample / IR blank ratio < 3, IR sample is the peak response value in the peak list, and IR blank is the peak response value of the background carbonyl compound).

[0039] Further preferably, in step (4-2), the characteristic mass distance M p2 –M p1 of the target peak pair is 5.03138 ± 0.00015 Da; the response ratio IR P1 / IR P2 of the target peak pair is between 0.67 and 1.5.

[0040] Further preferably, in step (4-3), for peaks with multiple molecular formula candidates, principles such as considering the molecular formula matching score (score), isotope peak shape matching degree (msigma value), matching mass error (mass errors), and the presence of homologues are comprehensively considered to exclude interfering options and obtain the molecular formula of the carbonyl derivative; subtracting part of C7H8N3 of the derivatization reagent to obtain the list of molecular formulas of potential carbonyl candidates.

[0041] After the present invention screens out the accurate mass and instrument response information of the d0 / d5-GRP labeled product through ultra-high resolution mass spectrometry, it matches and deduces the elemental composition molecular formula of the carbonyl compound based on the accurate mass and derivatization reaction mechanism, and conducts molecular-level characterization of the carbonyl compound composition.

[0042] The selected carbonyl compounds are aldehydes, ketones, and quinone compounds, which are aliphatic or aromatic aldehydes and ketones with a carbonyl functional group (R1-(C=O)-R2) in the molecular structure.

[0043] Preferably, the carbonyl compound is at least one selected from aliphatic aldehydes (decanal), aliphatic ketones (undecane-2-one), aromatic aldehydes (4-phenylbenzaldehyde), and aromatic ketones (fluorene-9-one).

[0044] The environmental matrix is ​​volatile gases and / or fine particulate matter; the fine particulate matter originates from at least one of the following: atmosphere, indoor dust, vehicle exhaust, and industrial smoke.

[0045] Preferably, the environmental substrate is derived from the atmosphere during typical haze weather in winter and summer; each atmospheric sample is collected continuously for 22-24 hours using a medium-sized sampler.

[0046] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0047] (1) This invention introduces characteristic mass tags through isotope labeling technology, which not only endows the test compound with characteristic information such as isotope mass difference and abundance ratio, but also makes the labeled product easier to ionize and be captured by mass spectrometer (improving mass spectrometry detection sensitivity), effectively eliminating false positive interference and achieving high-precision identification of unknown carbonyl compounds;

[0048] (2) The method of the present invention only requires a very small amount of labeling reagent to achieve efficient labeling of carbonyl functional groups, avoiding the complex and cumbersome carbonyl group derivation / or pretreatment process of other technologies;

[0049] (3) The method of the present invention can identify thousands of carbonyl compounds at the same time, making up for the shortcomings of the existing technology in identifying unknown carbonyl compounds, and realizing high-throughput identification of trace carbonyl compounds in complex environmental matrices. Attached Figure Description

[0050] Figure 1 This is a schematic diagram of the isotope labeling reaction process and recognition mechanism in Example 1;

[0051] Figure 2 The mass distance and intensity ratio of the peak pairs after d0 / d5-GRP labeling of the decanal standard identified in Example 1 are I. d0 / I d5 ;

[0052] Figure 3 The mass distance and intensity ratio of the peak pairs after d0 / d5-GRP labeling of the undecane-2-one standard identified in Example 1 are I. d0 / I d5 ;

[0053] Figure 4 The mass distance and intensity ratio of the peak pairs after d0 / d5-GRP labeling of the 4-phenylbenzaldehyde standard identified in Example 1 are I. d0 / I d5;

[0054] Figure 5 The mass distance and intensity ratio of the peak pairs after d0 / d5-GRP labeling of the fluorene-9-one standard identified in Example 1 are I. d0 / I d5 ;

[0055] Figure 6 This is a statistical graph of peak pair mass distance and peak pair intensity ratio in the unknown carbonyl compound identified in the particulate matter standard material in Example 2;

[0056] Figure 7 This is a statistical chart showing the quantity and abundance of carbonyl compounds in atmospheric particulate matter samples from summer and winter in Example 3. Detailed Implementation

[0057] A high-throughput identification method for carbonyl compounds in atmospheric particulate matter includes the following steps:

[0058] (1) Carbonyl compounds are labeled with chemical isotopes;

[0059] (2) High-throughput identification of carbonyl compounds labeled with isotopes.

[0060] In some specific embodiments, the reagents used for chemical isotope labeling are Girard reagents and natural deuterated Girard reagent P (abbreviated as d0 / d5-GRP), with a concentration of 0.01-10 mmol / L.

[0061] In some specific embodiments, the carbonyl compound is an aliphatic or aromatic aldehyde or ketone compound, with a concentration of 1-10 μmol / L.

[0062] In some specific embodiments, the labeling reagent and the carbonyl compound are reacted at room temperature in the dark for 5-60 minutes.

[0063] In some specific embodiments, the reaction between the labeling reagent and the carbonyl compound needs to be carried out under acidic conditions of hydrochloric acid, with a concentration of 0.01-10 mmol / L.

[0064] In some specific embodiments, high-throughput identification of the characteristics of isotopically labeled carbonyl compounds includes: 1) screening by mass spectrometry for mass-to-charge ratios of 120 to 900 (… m / z Within the quality range of d0 ( 1 H) and d5 ( 2 H) Using the peak pairs of the labeled products as signals, combined with a custom progressive filtering algorithm, the mass distance of the extracted peak pairs is determined to be 5.03138±0.00015 Da; 2) The intensity ratio of the peak pairs is 0.67-1.5; 3) The molecular formula matching error is within 1 ppm.

[0065] In some specific embodiments, after screening the peak pair information of the products containing d0 / d5-GRP labeling by high-resolution mass spectrometry, the elemental molecular composition information of the carbonyl compound labeled by d0 / d5-GRP is obtained by elemental molecular formula matching, and the carbonyl compound in the environmental matrix is ​​characterized at the molecular level.

[0066] In some specific embodiments, the sample after the chemical isotope-labeled carbonyl compound reaction is immediately blown to a near-dry state, centrifuged at high speed, and then diluted with an equal volume of a mixed solution of acetonitrile and water. Subsequently, high-resolution mass spectrometry analysis is performed. The inter-ion mass difference of each pair of peaks in the full scan mass spectrum is calculated, and unknown carbonyl compounds are efficiently identified by the isotopic characteristics of the d0 and d5 labeled carbonyl compounds.

[0067] In some specific embodiments, the carbonyl compound precursor is selected as an aldehyde or ketone carbonyl compound with an aliphatic or aromatic structure.

[0068] In some specific embodiments, the carbonyl compound is selected as one or any combination of two or more of the following: aliphatic aldehyde (decanal), aliphatic ketone (undecane-2-one), aromatic aldehyde (4-phenylbenzaldehyde), and aromatic ketone (fluorene-9-one).

[0069] In some specific embodiments, high-resolution mass spectrometry analysis of the system after adjusting the volume of the acetonitrile and water mixture involves using a Fourier transform ion cyclotron resonance mass spectrometer to analyze the precise mass of the carbonyl compound before and after isotope labeling. The scanning mode is a full ion scan in electrospray ionization positive ion mode, with a mass-to-charge ratio range of 120 to 900. A custom progressive filtering algorithm is used to accurately extract peak pairs within a fixed mass distance and a specific peak intensity ratio range, achieving high-throughput identification of carbonyl compounds.

[0070] In some specific embodiments, the environmental sample matrix in which the carbonyl compound is located is selected from the atmosphere during typical haze weather in winter and summer; each atmospheric sample is collected continuously for 22-24 hours using a medium-sized sampler.

[0071] The present invention will now be described through specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Variations and advantages that can be conceived by those skilled in the art without departing from the spirit and scope of the inventive concept are included in the present invention, and the scope of protection of the present invention is defined by the appended claims and any equivalents thereof.

[0072] The reagents and analytical equipment used in the following examples are described below:

[0073] Reagents: Girard reagent P and natural deuterated Girard reagent P (d0 / d5-GRP), hydrochloric acid solution, standard substances such as decanal, undecane-2-one, 4-phenylbenzaldehyde, fluorene-9-one, and atmospheric particulate matter standard substances (NIST SRM 1649b, provided by the National Institute of Standards and Technology, USA).

[0074] Mass spectrometer: Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS, Bruker Daltonik, Germany).

[0075] Example 1

[0076] The following steps are followed to perform the reaction of chemically isotopically labeled carbonyl compounds, with a focus on high-precision identification of the labeled carbonyl compounds. Specific steps include:

[0077] S1: Prepare acetonitrile solutions of standard substances such as decanal, undecane-2-one, 4-phenylbenzaldehyde, and fluorene-9-one at a concentration of 1 μmol / L;

[0078] S2: Dissolve d0 / d5-GRP (d0-GRP and d5-GRP in a 1:1 ratio) in a mixed solution of acetonitrile / water (90:10 by volume) to obtain a labeled reagent solution with a concentration of 0.01 mmol / L.

[0079] S3: Prepare hydrochloric acid solution by diluting commercial concentrated hydrochloric acid with acetonitrile, with the final concentration of hydrochloric acid being 0.01 mmol / L;

[0080] S4: Mix 200 µL of standard solution with 300 µL of labeled reagent solution, add 400 µL of hydrochloric acid solution, react at room temperature (e.g., 22-30℃) for 5 min, and terminate the reaction at -20℃. After centrifuging the reaction mixture at 12000 rpm for 10 min, blow it to near dryness with pure nitrogen, and then reconstitute it with 400 µL of a acetonitrile / water (volume ratio 1:1) mixture.

[0081] S5: Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) Analysis: Before each analysis, the instrument was calibrated using a 10 mmol / L sodium formate solution. The reconstituted reaction mixture was injected directly into the mass spectrometer at a rate of 2 µL / min. The flow rates of the ion source drying gas (heated nitrogen) and the nebulizer gas (nitrogen) were 4 L / min and 1 bar, respectively. The spray voltage was 3.8 kV, the desiccator temperature was 180 °C, and the ion saturation time in the ion cyclotron resonance mass spectrometer was 0.2 s, with a flight time of 0.6 s. To avoid cross-contamination of chemical substances, the sample introduction system was flushed sequentially with a mixture of acetonitrile and water in the same volume ratio between each sample analysis interval.

[0082] In addition, field blank, procedural blank, and solvent blank tests were conducted simultaneously to control potential contamination and interference that may be introduced during sampling, pretreatment, and instrument analysis. Field blank: Taking particulate matter sampling as an example, the sampling membrane was placed for 22-24 hours without the sampler in operation to monitor background interference during the sampling process. Procedural blank: A complete pretreatment process was performed on the blank membrane to monitor background interference during the pretreatment process. Solvent blank: Pure solvent was injected for analysis to monitor background interference caused by instrument residue. Setting blanks (field blank, procedural blank, solvent blank) is a quality control method to eliminate background contamination interference. A list of commonly detected carbonyl compounds in the blank samples (field blank, procedural blank, solvent blank) was constructed within the laboratory as background carbonyl compounds.

[0083] Ultra-high resolution spectroscopy was employed, with data acquisition performed in a broadband scanning mode within the mass range of 120-900 m / z. For each injection, 200 acquisition data points (transient size 4 mega-nodes, 32-bit data format) were superimposed to improve the signal-to-noise ratio of the spectra. Linear internal standard correction was performed on the acquired spectra using background carbonyl compounds to ensure that the mass error remained below 0.2 ppm throughout the entire acquisition mass range.

[0084] S6: By screening peak pairs with characteristic d0 / d5-GRP labels in the spectrum within the mass range of 120-900 m / z, carbonyl compounds can be distinguished from other ion signals. A custom progressive filtering algorithm developed using Python software is used, combined with the following three screening rules: (1) The mass distance between peak pairs is 5.03138 ± 0.00015 Da (theoretical mass distance ± error threshold), where theoretical mass distance = d5-GRP monoisotope mass - d0 monoisotope mass, and error = 0.00015 Da. The principle is explained in the figure. Figure 1(2) The intensity ratio of the peak pairs is 0.67~1.5:1 (i.e., the ratio of the peak intensities of d0-GRP and d5-GRP is between 0.67 and 1.5); (3) The molecular formula matching error is within 1 ppm. Finally, the above four carbonyl compounds were accurately identified ( Figures 2-5 ).

[0085] The specific steps are as follows:

[0086] (1) For the raw data of FT-ICR MS analysis, mass spectrometry peak detection and peak identification were first performed using DataAnalysis (version 4.0, Bruker Daltonics GmbH, Bremen, Germany) (peak response IR>1×10). 6 (S / N≥3), isotope peaks and adduct ion peaks were removed. Internal standard correction (Cal 2 linear mode) was performed using background carbonyl hydrazone derivatives to further reduce the FT-ICR MS mass axis error (<0.2ppm). A peak list was obtained.

[0087] (2) A data processing package PMF (Paired Mass Filtering) was developed based on Python for the obtained peak list. First, background subtraction was performed on the sample peak list to remove interfering mass spectrometry peaks (IR peaks) that overlapped with those in the solvent blank sample and the experimental blank sample. sample / IR blank A ratio <3 ensures the reliability of mass spectrometry data.

[0088] (3) Based on peak pairs (M) p1 M p2 M p1 <M p2 Characteristic mass distance M p2 -M p1 =5.03138±0.00015Da, a progressive filtering algorithm based on PMF packets is used to select target peak pairs, and the peak pair response ratio IR P1 / IR P2 A list of potential carbonyl compound light and heavy isotope labeled derivatives is generated between 0.67 and 1.5.

[0089] (4) For M p1 The exact mass numbers are used to match the elemental molecular formulas, with the number of elements constituting the formula limited to a certain limit. 12 C (1–100) 1 H(1–200), 16 O (0–50) 15 N(3–6) and 32S (0–1), with the absolute value of the molecular formula matching error < 1 ppm. For mass peaks with multiple molecular formula candidates, principles such as the molecular formula matching score (score), isotope peak shape matching degree (msigma value), matching mass error (mass errors), and the presence of homologues are comprehensively considered to exclude interfering options, and finally the molecular formula of the carbonyl derivative is determined. Subtracting a part of C7H8N3 (corresponding to the d0-GRP group) of the derivatization reagent gives a list of molecular formulas of potential carbonyl candidates, and the reliability of the molecular formulas of the selected carbonyl candidates is ensured by rules such as 0.3 ≤ H / C ≤ 2.25, 0 < O / C ≤ 1.2, 0 ≤ N ≤ 3, and 0 < DBE / C ≤ 1.

[0090] Example 2

[0091] Compared with Example 1, the high-precision identification of unknown carbonyl compounds in complex matrices was carried out using atmospheric particulate matter reference material (NIST SRM 1649b). The specific steps include:

[0092] S1: Dissolve 2.5 mg of NIST SRM 1649b atmospheric particulate matter reference material in 10 mL of acetonitrile and perform ultrasonic extraction. Then centrifuge at 12000 rpm / min for 10 min and collect the supernatant. Combine the supernatants obtained from three consecutive extractions and concentrate to 1 mL under the conditions of 40 °C and 1200 rpm / min. Then, blow the concentrated solution to a nearly dry state under a pure nitrogen atmosphere and redissolve it with 400 μL of acetonitrile to prepare the carbonyl compound precursor NIST SRM 1649b solution;

[0093] S2: Dissolve d0 / d5-GRP in a mixed solution of acetonitrile / water (volume ratio 90:10) to obtain a labeling reagent solution with a concentration of 1 mmol / L;

[0094] S3: Prepare a hydrochloric acid solution by diluting commercial concentrated hydrochloric acid with acetonitrile, and the final concentration of the prepared hydrochloric acid is 0.5 mmol / L;

[0095] S4: Take 200 µL of the NIST SRM 1649b solution and mix it with 300 µL of the labeling reagent solution. Add 400 µL of the hydrochloric acid solution and react at room temperature (such as 22 - 30 °C) for 10 min and terminate the reaction under the condition of -20 °C. After the reaction mixture is centrifuged at 12000 rmp / min at high speed for 10 min, blow it to a nearly dry state with pure nitrogen, and then redissolve it with 400 µL of the same volume ratio of acetonitrile / water mixed solution;

[0096] S5: Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) Analysis: Before each analysis, the instrument was calibrated using a 10 mmol / L sodium formate solution. The reconstituted reaction mixture was injected directly into the mass spectrometer at a rate of 2 µL / min. The ion source drying gas flow rate and nebulizer gas flow rate were 4 L / min and 1 bar, respectively. The spray voltage was 3.8 kV, the desiccator temperature was 180 °C, and the ion saturation time in the ion cyclotron resonance mass spectrometer was 0.2 s, with a flight time of 0.6 s. To avoid cross-contamination of chemical substances, the sample introduction system was flushed sequentially with a mixture of acetonitrile and water in the same volume ratio between each sample analysis interval. In addition, field blank, program blank, and solvent blank tests were conducted simultaneously to control potential contamination and interference that may be introduced during sampling, pretreatment, and instrument analysis. Ultra-high resolution spectroscopy was used, and data was acquired in broadband scanning mode within the mass range of 120–900 m / z. For each injection, 200 acquisition data points (transient size 4 mega-nodes, 32-bit data format) were superimposed to improve the mass spectrometry signal-to-noise ratio. Background carbonyl compounds were used to perform linear internal standard correction on the acquired spectra, ensuring that the mass error was below 0.2 ppm throughout the entire acquisition mass range.

[0097] S6: By screening peak pairs with characteristic d0 / d5-GRP labels in the spectrum within the mass range of 120-900 m / z, carbonyl compounds can be distinguished from other ion signals. A custom progressive filtering algorithm developed using Python software is used, combined with the following three screening rules: (1) The mass distance between peak pairs is 5.03138 ± 0.00015 Da (theoretical mass distance ± error threshold), where theoretical mass distance = d5-GRP monoisotope mass - d0 monoisotope mass, and error = 0.00015 Da. The principle is explained in the figure. Figure 1 (2) The intensity ratio of peak pairs is 0.67-1.5; (3) The molecular formula matching error is within 1 ppm.

[0098] S7: Similar to the process in Example 1, the reaction conditions for isotope-labeled carbonyl compounds were changed to verify the effectiveness and accuracy of the method under complex matrix conditions and different reaction conditions. Specifically, the dosage of the labeling reagent, the dosage of hydrochloric acid, and the reaction time were examined.

[0099] With the same volume of reactants added, the recognition ability was verified under different concentrations of d0 / d5-GRP in S2 (0.5, 1, 2, 5, and 10 mmol / L), different concentrations of hydrochloric acid solution in S3 (0.5, 1, 2, 5, and 10 mmol / L), and different reaction times in S4 (5, 10, 20, 30, and 60 min). Using NIST SRM 1640b standard material, thousands of carbonyl compounds labeled with d0 / d5-GRP were accurately screened (see Table 1 below). The results showed that the highest number of carbonyl compounds were identified under the conditions of 1 mM d0 / d5-GRP, 0.5 mM HCl, and 10 min of reaction. The results of the three parallel experiments were almost identical (n=1233±11), indicating that this method can efficiently identify unknown carbonyl compounds under the conditions of 1 mmol / L d0 / d5-GRP, 0.5 mmol / L HCl, and 10 min of reaction.

[0100] Statistical results show that CHO is the most common chemical component, accounting for as high as 93±1%. Analysis of all detected carbonyl compounds reveals ( Figure 6 The method showed that 85% of the peak-to-mass distance errors were within ±0.10 mDa, and 93% of the peak-to-intensity ratios were within the range of 0.67 to 1.5, indicating the stability and reliability of the method and its potential for application in non-targeted analysis of complex environmental matrices.

[0101] Table 1

[0102]

[0103] Example 3

[0104] The purpose of this embodiment is to further verify the ability of the method to identify unknown carbonyl compounds in real environments under hazy weather conditions in summer and winter. It is worth noting that carbonyl compounds in atmospheric hazy samples are mostly at trace levels.

[0105] Except for the carbonyl compounds being actual atmospheric particulate matter samples, the other steps were basically similar to those in Example 2. Ultimately, over 2000 unknown carbonyl compounds with molecular formulas ranging from 317 to 1122 were identified from 38 samples. Statistical analysis revealed that CHO compounds overwhelmingly accounted for 74±3% and 84±4% of the samples from summer and winter, respectively, with individual samples contributing even more, at 85±2% and 92±2%, respectively. Figure 7 This is a statistical chart showing the number and proportion of identifiable carbonyl compounds in samples from summer and winter.

[0106] The embodiments described above provide a detailed explanation of the technical solutions and beneficial effects of the present invention. It should be understood that the above descriptions are merely specific embodiments of the present invention and are not intended to limit the present invention. Any modifications, additions, and equivalent substitutions made within the scope of the principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A high-throughput identification method for carbonyl compounds in an environmental matrix, characterized in that, It includes the following steps: (1) Extract carbonyl compounds from the environmental matrix sample to be measured. After removing the solvent in the extract, a concentrated solution of carbonyl compounds is obtained. (2) Add a stable isotope pair labeling reagent to the concentrated solution of carbonyl compounds for derivatization reaction. After terminating the reaction, perform drying treatment and re-dissolve to obtain a derivatized sample. The stable isotope pair labeling reagent is Girard's reagent P and deuterated Girard's reagent P. (3) Perform ultra-high resolution mass spectrometry analysis on the derivatized sample to obtain original ultra-high resolution mass spectrometry data. Conduct field blanks, procedural blanks and solvent blanks tests. The carbonyl compounds detected in the field blanks, procedural blanks and solvent blanks are used as background carbonyl compounds. Use a Fourier transform ion cyclotron resonance mass spectrometer to perform ultra-high resolution mass spectrometry analysis on the derivatized sample. The scanning mode is full ion scan in the positive ion mode of electrospray ionization. The full scan mass range is 120 - 900 m / z to obtain ultra-high resolution mass spectrometry data. (4) Screen the peak pair information of the product after being labeled with a stable isotope pair from the original ultra-high resolution mass spectrometry data. Based on the peak pair information, obtain the elemental molecular composition information of the carbonyl compounds labeled with a stable isotope pair through elemental molecular formula matching, and conduct molecular-level characterization of the carbonyl compounds in the corresponding environmental matrix, including: (4-1) Perform mass spectrometry peak detection and peak identification on the original ultra-high resolution mass spectrometry data,剔除 isotope peaks and adduct ion peaks, perform internal standard correction to obtain a peak list. Perform background subtraction on the peak list and剔除 interference mass spectrometry peaks that coincide with the background carbonyl compounds. (4-2) Select target peak pairs from the peak list to generate a list of potential carbonyl compound light and heavy isotope labeled derivative pairs (M p1 M p2 M p1 < M p2 M p1 M p2 These represent the mass-to-charge ratio of the target peak pair and the characteristic mass distance M of the target peak pair, respectively. p2 -M p1 The peak-to-response ratio (IR) is 5.03138 ± 0.00015 Da. P1 / IR P2 Between 0.33 and 3, IR P1 IR P2 These represent the response intensities of the target peak pairs; (4-3) For M p1 The exact mass number is used to match the elemental molecular formula, and the number of elements in the molecular formula is limited to 1. 12 C is 1~100, 1 H is 1~200, 16 O is 0~50, 15 N is 3~6 32 S is 0~1, and the absolute value of molecular formula matching error is <1ppm; 扣除部分 of the derivatizing reagent C7H8N3 to obtain a list of molecular formulas of potential carbonyl candidates. (4-4) Screen out the molecular formulas of reliable carbonyl candidates from the list of molecular formulas of potential carbonyl candidates through the rule constraints of the atomic numbers of elements in the molecular formula. The rule constraints are 0.3 ≤ H / C ≤ 2.25, 0 < O / C ≤ 1.2, 0 ≤ N ≤ 3, and 0 < DBE / C ≤ 1, where H, C, O, and N are the atomic numbers of elements hydrogen, carbon, oxygen, and nitrogen in the molecular formula respectively, and DBE is the number of unsaturated double bonds.

2. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 1, characterized in that, In step (1), when extracting carbonyl compounds from the environmental matrix sample to be measured, the extraction solvent used is acetonitrile or a mixed solution of acetonitrile and water.

3. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 2, characterized in that, In the mixed solution of acetonitrile and water, the volume ratio of acetonitrile to water is (8 - 9.5):

1.

4. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 1, characterized in that, In the stable isotope pair labeling reagent, the molar ratio of Girard's reagent P to deuterated Girard's reagent P is 1:

1.

5. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 1, characterized in that, In step (2), the derivatization reaction is carried out at room temperature in the dark for 5-60 min.

6. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 1, characterized in that, In step (4-3), for peaks with multiple molecular formula candidates, comprehensively consider the molecular formula matching score, isotope peak shape matching degree, matching mass error, and the presence of homologues to排除 interference options and obtain the molecular formula of the carbonyl derivative.扣除部分 of the derivatizing reagent C7H8N3 to obtain a list of molecular formulas of potential carbonyl candidates.

7. The high-throughput identification method for carbonyl compounds in an environmental matrix according to claim 1, characterized in that, The environmental matrix is volatile gas and / or fine particulate matter; the fine particulate matter comes from at least one of atmosphere, indoor dust, vehicle exhaust, and industrial soot.

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