A method for analyzing molecular structure information of dissolved organic matter based on high-resolution tandem mass spectrometry

Abstract

The present application relates to the technical field of dissolved organic matter, and particularly relates to a method for analyzing molecular structure information of dissolved organic matter based on high-resolution tandem mass spectrometry. The method for analyzing molecular structure information of dissolved organic matter based on high-resolution tandem mass spectrometry comprises the following steps: obtaining tandem mass spectrometry data; tracking fragmentation path and identifying functional groups, identifying possible functional group combinations based on neutral loss mass difference between starting point molecules and fragment ions, and tracking fragmentation path; identifying and quantifying isomers, identifying and quantifying isomers of target molecules based on fragmentation path; algorithm evaluation and parameter optimization; structure evolution and statistical analysis. The method for analyzing molecular structure information of dissolved organic matter based on high-resolution tandem mass spectrometry establishes an analysis process from parent ions to structural isomers by constructing fragmentation path, identifying neutral loss combinations and extracting isomer characteristics, and significantly improves the depth and accuracy of molecular structure analysis of dissolved organic matter.

Description

Technical Field

[0001] This invention relates to the field of soluble organic matter technology, and in particular to a method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry. Background Technology

[0002] Dissolved organic matter (DOM), as one of the largest pools of active organic carbon in the ocean, plays a crucial role in regulating the global carbon cycle. The structure and composition of DOM molecules determine their reactivity, bioavailability, and fate in water, thereby influencing the cycling of marine biogenic elements and environmental ecological processes.

[0003] Currently, the direct introduction of ultra-high resolution mass spectrometry (DI-UHR MS) techniques, such as Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS), can achieve molecular formula allocation of DOM molecules, significantly improving our understanding of DOM chemical composition. However, since the obtained results are essentially average signals of multiple isomers, they lack structural sensitivity and cannot resolve the large number of isomers and their functional group information present in DOM.

[0004] Therefore, tandem mass spectrometry (MS / MS) fragmentation analysis has been used as a supplementary method to provide structural clues for DOM molecules. MS / MS fragmentation analysis, by controlling collision energies, induces the neutral loss of parent ions to generate fragment ions, thereby inferring the functional group types and molecular skeleton characteristics of the original molecule. However, current strategies for interpreting fragmentation data are still immature, lacking standardized algorithms, which poses a significant limitation to the interpretation of the results.

[0005] The existing technology has the following main problems: the current fragmentation path identification is basically in a black box state, and there is no clear description and evaluation of the structural path analysis method between the parent ion and all its fragments. Summary of the Invention

[0006] To address the problems existing in the prior art, this invention provides a method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry, comprising:

[0007] Tandem mass spectrometry data were obtained, and high-quality molecular formulas were obtained through sample injection analysis. Typical mass-to-charge ratio windows were selected for tandem mass spectrometry fragmentation analysis. Fragmentation parameters were dynamically adjusted to ensure that the main peak intensity was significantly reduced and there were obvious fragmentation peak signals in the spectrum.

[0008] Fragmentation path tracing and functional group identification: Based on the neutral loss mass difference between the starting molecule and fragment ions, possible functional group combinations are identified and their fragmentation paths are traced.

[0009] Isomer identification and quantification: Identifying and quantifying isomers of target molecules based on fragmentation pathways;

[0010] Algorithm evaluation and parameter optimization: Core indicators are used to evaluate the explanatory effect, thereby selecting the optimal explanatory strategy;

[0011] Structural evolution and statistical analysis were used to calculate the Bray-Curtis distance based on relative fragmentation intensity, and to compare the differences in DOM structure among samples from different environments. The Spearman rank correlation coefficient between functional group loss and elemental composition was analyzed to verify and explore the chemical driving factors and laws behind these structural differences from a mechanistic perspective.

[0012] For substances like DOM, which are primarily detected using electrospray ionization (ESI), in most cases, the detected ions are single-charged ions ([M+H)). + Or [MH] - Therefore, the mass-to-charge ratio (m / z) is numerically approximately equal to the molecular mass (M) of its neutral molecule.

[0013] Neutral loss refers to the phenomenon in mass spectrometry where the parent ion loses an uncharged neutral molecule or radical fragment during fragmentation, generating a daughter ion. The mass-to-charge ratio (m / z) of the daughter ion minus the mass-to-charge ratio (m / z) of the parent ion equals the mass of the lost neutral fragment (in Da or u).

[0014] Traditional tandem mass spectrometry (MS / MS) analysis often focuses on single fragmentation events or functional group markers, failing to distinguish between different structures with the same molecular formula. This invention, by constructing a neutral loss path and based on differences in fragmentation paths, achieves isomer identification for the first time in non-targeted DOM samples, significantly expanding the analytical capabilities of DOM molecular structures.

[0015] The fragmentation path tracing algorithm designed in this invention can continuously identify multiple rounds of neutral loss events and automatically construct the spectral relationship between the parent ion, fragments, and functional groups. This far surpasses the traditional method that only analyzes first- and second-order fragment pairs, thus improving analytical coverage and structure reduction capabilities.

[0016] Based on the relative fragmentation intensity (i.e., the proportion of fragment intensity to the total intensity of parent and daughter molecules) of target molecules in each sample, the structural similarity matrix (Bray-Curtis) between samples is calculated. This invention applies the aforementioned structural analysis algorithm to natural estuarine samples, achieving for the first time a dynamic characterization of DOM structural diversity under salinity gradients. It has been successfully applied to DOM samples along the salinity gradient in the Yangtze River estuary, revealing the trend of increased DOM structural isomer numbers and changing functional group loss patterns during the transition from freshwater to seawater. This represents the first quantitative study of DOM structural evolution based on isomer levels.

[0017] The algorithm proposed in this invention can be used in conjunction with other experimental methods such as LC-UHR-MS (liquid chromatography-ultra-high resolution mass spectrometry), stable isotope labeling, and photodegradation experiments. It supports the construction of various ecological function simulation systems such as DOM structure stability models and reactivity prediction models, and has great potential for expansion.

[0018] Furthermore, the dynamic adjustment of the fragmentation parameters includes: adjusting the impact voltage to between 10eV and 20eV, and adjusting the cumulative time to between 0.2s and 1s.

[0019] Typically, the collision voltage is adjusted to around 10eV-20eV, and the cumulative time is adjusted to between 0.2s and 1s to ensure that the intensity of the main peak in the spectrum is significantly reduced and there is a clear fragmentation peak signal.

[0020] Furthermore, the identification of possible functional group combinations and tracing of their fragmentation paths based on the neutral loss mass difference between the starting point molecule and fragment ions includes:

[0021] Set parameters and define the functional group library. Setting parameters includes setting the signal-to-noise ratio threshold and setting the quality error.

[0022] The path tracing strategy employs a recursive path tracing algorithm, starting from the initial peak of each precursor ion and gradually matching neutral losses. Each neutral loss is treated as a fragmentation step, and the new ion is used as the next precursor ion to continue tracing.

[0023] The functional group matching process involves calculating the mass difference between each parent ion and all daughter ions; matching this mass difference with the theoretical mass of each functional group in the functional group library; allowing a maximum of one type of functional group for each mass difference; recording the mass-to-charge ratio of the parent ion, the mass-to-charge ratio of the fragment ions, the mass error, the matched functional groups, and the number of functional groups at each step; checking whether there are ions corresponding to intermediate fragments in each complete path; and retaining reasonable path combinations.

[0024] Record the parent ion, terminal fragments, and neutral loss combination for each effective fragmentation path, and output the results.

[0025] Furthermore, the signal-to-noise ratio threshold is set to 4, 6, and 10 to filter valid peaks; the quality error is the allowable matching quality error range, set to 0.1 ppm, 0.2 ppm, and 0.5 ppm.

[0026] Furthermore, define the functional group library, including:

[0027] The functional group library includes a comprehensive functional group library or a simplified functional group library; the comprehensive functional group library includes: CO2, H2O, CO, SO2, SO3, C2H4O, CH3OH; the simplified functional group library includes: CO2, H2O, CO, CH3OH.

[0028] This invention establishes a joint optimization framework based on recovery rate and isomer number by systematically testing different signal-to-noise ratios, mass errors, and functional group combinations. This ensures that the interpretation results are stable in complex environmental samples and are applicable to different mass spectrometry platforms and sample types.

[0029] Furthermore, the output results include:

[0030] Output all matching results for each step of neutral loss; the maximum number and intensity of functional group combinations for each parent ion; and a statistical table of functional group types.

[0031] Furthermore, the isomer identification and quantification includes:

[0032] Identify isomers; those with the same molecular formula but different combinations of neutral loss or terminal fragment ions are considered as different structural isomers.

[0033] For each parent ion, count the number of identifiable isomers;

[0034] Calculate the relative fragmentation strength of each isomer It is used to reflect structural stability and diversity.

[0035] Furthermore, the core metrics include:

[0036] Molecular formula number recovery rate, which is the proportion of the number of identified fragments to the total number of fragments; intensity recovery rate, which is the proportion of the intensity of the resolved fragments to the total intensity.

[0037] A dual-index optimization mechanism based on molecular formula recovery rate and isomer number is introduced to improve the stability and generalization of the algorithm. Through systematic testing under different signal-to-noise ratios, mass errors, and functional group combinations, this invention establishes a joint optimization framework based on molecular formula recovery rate and isomer number, ensuring stable performance of the interpretation results in complex environmental samples and making it applicable to different mass spectrometry platforms and sample types.

[0038] Furthermore, the Bray-Curtis distance is calculated using the following formula (1).

[0039]

[0040] Where C ij x is the Bray-Curtis distance between sample i and sample j; ikx represents the k-th relative fragmentation strength in sample i; jk Let x be the k-th relative fragmentation strength in sample j; min(x) ik ,x jk ) represents the minimum value of the k-th feature in the two samples; the denominator is the sum of all features in the two samples.

[0041] This invention applies the aforementioned structural analysis algorithm to natural estuarine samples, achieving for the first time a dynamic characterization of DOM structural diversity under salinity gradients. Through inter-sample structural similarity calculation and functional group loss mode analysis, the evolutionary trend of DOM structures from land to sea is revealed.

[0042] Furthermore, the Spearman rank correlation coefficient is calculated as shown in equation (2).

[0043]

[0044] in d is the Spearman rank correlation coefficient, ranging from [-1, 1], representing the degree of monotonic correlation between variables; i Let be the rank difference of the i-th pair of data; n is the number of paired samples.

[0045] Spearman rank correlation coefficient analysis is performed on the average loss number of each functional group in each isomer of the molecule (such as the average CO2 and H2O loss number) and the elemental composition parameters of the molecule (such as O / C and NOSC) to determine the degree of correlation. This determines the degree of association between the molecular composition and structural features of the DOM, and indirectly judges the effectiveness of the analysis. A higher degree of correlation indicates that the analysis effect may be better.

[0046] In summary, the advantages and beneficial effects of the present invention are as follows:

[0047] This invention provides a method for analyzing DOM molecular structure information based on high-resolution tandem mass spectrometry. By systematically constructing fragmentation paths, identifying neutral loss combinations, and extracting isomer features, an analytical process from parent ion to structural isomers is established, significantly improving the depth and accuracy of DOM molecular structure analysis. This fragmentation data interpretation process, isomer identification mechanism, and structural evolution modeling method constitute an integrated system.

[0048] This invention innovatively designs a recursive fragmentation path tracing algorithm based on the loss of neutral functional groups (such as CO2, H2O, SO3, etc.), which can automatically identify the structural paths between the parent ion and its multiple fragments. This mechanism not only realizes the automatic interpretation of the fragmentation process, but also supports multi-round matching and path termination condition control, constructing a complete structure transformation map.

[0049] This invention proposes a structure-sensitive isomer discrimination method: parent ions with the same molecular formula but different fragmentation combinations (neutral loss paths) are considered as different structural isomers, and the isomer distribution and structural diversity are quantified by combining their corresponding fragment intensities. This strategy breaks through the traditional assumption of "molecular formula uniqueness" and achieves isomer identification in non-targeted environmental samples for the first time. Attached Figure Description

[0050] Figure 1 This is a flowchart of a method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry, as disclosed in this invention. Detailed Implementation

[0051] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below in conjunction with specific embodiments and corresponding drawings. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0052] Embodiments of the present invention are described in detail below. Examples of these embodiments are shown in the accompanying drawings, wherein the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0053] This invention provides a method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry, such as... Figure 1 As shown, it includes:

[0054] S10: Obtain tandem mass spectrometry data, inject samples for analysis to obtain high-quality molecular formulas; select a typical mass-to-charge ratio window for tandem mass spectrometry fragmentation analysis; dynamically adjust fragmentation parameters to ensure that the main peak intensity on the spectrum is significantly reduced and there is a clear fragmentation peak signal.

[0055] In step S10, the fragmentation parameters are dynamically adjusted, including adjusting the impact voltage to between 10eV and 20eV and the cumulative time to between 0.2s and 1s.

[0056] For example, three representative DOM samples (Y1-freshwater, Y2-medium saline, Y3-seawater) were collected from the Yangtze River estuary, filtered using a 0.2 μm pore size filter membrane, and then solid-phase extracted using PPL (styrene-divinylbenzene polymer solid-phase extraction column packing) to concentrate to 100 mg / L.

[0057] Tandem mass spectrometry detection: Direct injection analysis was performed using an FT-ICR MS system, yielding 4398 high-quality molecular formulas after formula assignment. MS / MS fragmentation experiments were conducted within five typical m / z windows (335, 350, 355, 415, 453). Fragmentation parameters (collision voltage and accumulation time) were dynamically adjusted, typically with the collision voltage set to approximately 10-20 eV and the accumulation time between 0.2 s and 1 s, ensuring a significant decrease in the main peak intensity and the presence of clear fragmentation peak signals in the spectrum.

[0058] S20: Fragmentation path tracking and functional group identification. Based on the neutral loss mass difference between the starting molecule and fragment ions, it identifies possible functional group combinations and tracks their fragmentation paths.

[0059] S30: Isomer identification and quantification, based on fragmentation pathways to identify and quantify isomers of target molecules.

[0060] S40: Algorithm evaluation and parameter optimization, using core indicators to evaluate the explanatory effect, thereby selecting the optimal explanatory strategy.

[0061] S50: Structural evolution and statistical analysis, calculating the Bray-Curtis distance based on relative fragmentation intensity, comparing the differences in DOM structure among samples from different environments as a whole; analyzing the Spearman rank correlation coefficient between functional group loss and elemental composition, verifying and exploring the chemical driving factors and laws behind these structural differences from a mechanistic perspective.

[0062] In one embodiment, step S20 involves identifying possible functional group combinations based on the neutral loss mass difference between the starting point molecule and the fragmentation ions, and tracing their fragmentation paths, including the following steps:

[0063] S201: Set parameters and define functional group library. Setting parameters includes setting the signal-to-noise ratio threshold and setting the quality error.

[0064] Specifically, the signal-to-noise ratio (S / N) threshold is set to 4, 6, and 10 to filter valid peaks; the quality error (in ppm) is the allowable range of matching weight error, set to 0.1 ppm, 0.2 ppm, and 0.5 ppm.

[0065] Define a library of functional groups, including:

[0066] The functional group library includes a comprehensive functional group library or a simplified functional group library; the comprehensive functional group library includes: CO2, H2O, CO, SO2, SO3, C2H4O, CH3OH; the simplified functional group library includes: CO2, H2O, CO, CH3OH.

[0067] Depending on the actual situation, you can choose to use a comprehensive functional group library or a simplified functional group library that only includes common functional groups.

[0068] S202: Path tracking strategy, adopting a recursive path tracking algorithm, starting from each parent ion's initial peak, gradually matching neutral losses, treating each neutral loss as a fragmentation step, and using the new ion as the next parent ion to continue tracking.

[0069] For example, a recursive path tracing algorithm is used, starting from each precursor ion's initial peak and progressively matching neutral losses (such as CO2, H2O, CH3OH, SO3, etc.). Each neutral loss is considered a fragmentation step, and the new ion is used as the next precursor ion to continue tracing. Each round involves a maximum of 30 steps, forming a complete neutral loss path.

[0070] A complete fragmentation path tracing mechanism is constructed, supporting multi-round iterations and functional group combination identification. The fragmentation path tracing algorithm designed in this invention can continuously identify multiple rounds of neutral loss events and automatically construct the spectral relationship between the parent ion, fragments, and functional groups, far exceeding traditional methods that only analyze primary / secondary fragment pairs, thus improving analytical coverage and structure reduction capabilities.

[0071] S203: Functional group matching process: For each parent ion, calculate its mass difference (Δm / z) with all daughter ions; match this mass difference with the theoretical mass of each functional group in the functional group library (within ppm); for each mass difference, a maximum of one functional group is allowed; record the mass-to-charge ratio (m / z) of the parent ion, the mass-to-charge ratio (m / z) of the fragment ions, the mass error, the matched functional group, and the number of functional groups for each step; check whether there is an ion corresponding to the intermediate fragment in each complete path; retain reasonable path combinations.

[0072] S204: Record the parent ion, terminal fragments, and neutral loss combination for each effective fragmentation path, and output the results.

[0073] Specifically, the output results include:

[0074] Output all matching results for neutral loss at each step; the maximum number and intensity of functional group combinations for each parent ion; and a statistical table of functional group types (combination frequency, total intensity, average intensity, etc.).

[0075] In one embodiment, step S30, isomer identification and quantification, includes:

[0076] S301: Identify isomers. Paths with the same molecular formula but different neutral loss combinations or terminal fragment ions are considered as different structural isomers.

[0077] S302: For each parent ion, count the number of identifiable isomers.

[0078] S303: Calculate the relative fragmentation strength of each isomer It is used to reflect structural stability and diversity.

[0079] This invention utilizes the unique "fragmentation path" in tandem mass spectrometry as a fingerprint to identify and quantify different structural isomers with the same molecular formula in DOM (Dissociative Oxidant). Step S301 defines isomer discrimination criteria based on fragmentation patterns. Step S302 counts the number of isomers corresponding to each molecular formula. Step S303 calculates the relative abundance of each isomer. This method deepens DOM composition analysis from "what molecular formula it is" to "what different structures these molecular formulas may exist in, and what proportion of each exists," thereby revealing deeper structural stability, diversity, and transformation information of DOM at the sub-molecular formula level.

[0080] In one embodiment, in step S40, the core indicators include:

[0081] Molecular formula number recovery rate, the proportion of identified fragments to the total number of fragments;

[0082] Strength recovery rate, the proportion of the strength of the analyzed fragments to the total strength.

[0083] The higher these two indicators are, the stronger the algorithm's ability to cover the parent ions and fragments detected in the spectrum, thus enabling the selection of the optimal interpretation strategy.

[0084] A dual-index optimization mechanism based on molecular formula number, recovery rate, and isomer number is introduced to improve the stability and generalization of the algorithm. Through systematic testing under different signal-to-noise ratios, mass errors, and functional group combinations, this invention establishes a joint optimization framework based on recovery rate and isomer number, ensuring stable performance of the interpretation results in complex environmental samples and making it applicable to different mass spectrometry platforms and sample types.

[0085] This invention introduces several key parameters (signal-to-noise ratio threshold, quality error tolerance, functional group combination) into the algorithm implementation, and selects the optimal interpretation strategy by jointly evaluating the recovery rate index (molecule number recovery rate, intensity recovery rate) and the number of isomers.

[0086] In one embodiment, in step S50, the Bray-Curtis distance is calculated using the following formula (1).

[0087]

[0088] Where C ij x is the Bray-Curtis distance between sample i and sample j; ik x represents the k-th relative fragmentation strength in sample i;jk Let x be the k-th relative fragmentation strength in sample j; min(x) ik ,x jk ) represents the minimum value of the k-th feature in the two samples; the denominator is the sum of all features in the two samples.

[0089] The Bray-Curtis dissimilarity index was used to systematically calculate the degree of structural difference among all sample pairs and organize them into a matrix. The purpose of this step was to elevate the analytical perspective from local features to a global model; to quantify the magnitude of DOM structure changes along the environmental gradient (salinity); and to provide a core data foundation for subsequent statistical visualizations (such as clustering and ordination plots) and hypothesis testing.

[0090] Specifically, the Spearman rank correlation coefficient is calculated as shown in equation (2).

[0091]

[0092] in d is the Spearman rank correlation coefficient, ranging from [-1, 1], representing the degree of monotonic correlation between variables; i Let be the rank difference of the i-th pair of data; n is the number of paired samples (i.e., the number of sample pairs).

[0093] Spearman rank correlation coefficient analysis was performed on each functional group loss type (such as average CO2 and H2O loss number) and elemental composition parameters (such as O / C and NOSC) to determine the degree of correlation, thereby judging the degree of association between the molecular composition and structural characteristics of DOM, and indirectly judging the effectiveness of the analysis. The higher the degree of correlation, the better the analysis effect may be.

[0094] The above-mentioned method for analyzing dissolved organic matter based on high-resolution tandem mass spectrometry can be used to analyze the loss of neutral functional groups in DOM at the structural level, thereby indirectly revealing the structural characteristics of DOM and providing new evidence for understanding the compositional characteristics and potential degradation mechanisms of DOM.

[0095] Finally, it should be noted that any modification or equivalent substitution of some or all of the technical features made based on the technical solutions of the present invention and the embodiments thereof, without departing from the corresponding technical solutions of the present invention, shall fall within the patent scope of the present invention and the embodiments thereof.

Claims

1. A method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry, characterized in that, include: Obtain tandem mass spectrometry data, and perform sample injection analysis to obtain high-quality molecular formulas; Typical mass-to-charge ratio windows were selected for tandem mass spectrometry fragmentation analysis; fragmentation parameters were dynamically adjusted to ensure that the main peak intensity was significantly reduced and there were obvious fragmentation peak signals in the spectrum. Fragmentation path tracing and functional group identification: Based on the neutral loss mass difference between the starting molecule and fragment ions, possible functional group combinations are identified and their fragmentation paths are traced. Isomer identification and quantification: Identifying and quantifying isomers of target molecules based on fragmentation pathways; Algorithm evaluation and parameter optimization: Core indicators are used to evaluate the explanatory effect, thereby selecting the optimal explanatory strategy; Structural evolution and statistical analysis were conducted, and the Bray-Curtis distance was calculated based on relative fragmentation intensity to compare the differences in DOM structure among samples from different environments. The Spearman rank correlation coefficient between functional group loss and elemental composition was analyzed to verify and explore the chemical driving factors and patterns behind these structural differences from a mechanistic perspective. The method of identifying possible functional group combinations and tracing their fragmentation paths based on the neutral loss mass difference between the starting point molecule and fragment ions includes: Set parameters and define the functional group library. Setting parameters includes setting the signal-to-noise ratio threshold and setting the quality error. The path tracing strategy employs a recursive path tracing algorithm, starting from the initial peak of each precursor ion and gradually matching neutral losses. Each neutral loss is treated as a fragmentation step, and the new ion is used as the next precursor ion to continue tracing. The functional group matching process involves calculating the mass difference between each parent ion and all daughter ions; matching this mass difference with the theoretical mass of each functional group in the functional group library; allowing a maximum of one type of functional group for each mass difference; recording the mass-to-charge ratio of the parent ion, the mass-to-charge ratio of the fragment ions, the mass error, the matched functional groups, and the number of functional groups at each step; checking whether there are ions corresponding to intermediate fragments in each complete path; and retaining reasonable path combinations. Record the parent ion, terminal fragments, and neutral loss combination for each effective fragmentation path, and output the results; The isomer identification and quantification include: Identify isomers; those with the same molecular formula but different combinations of neutral loss or terminal fragment ions are considered as different structural isomers. For each parent ion, count the number of identifiable isomers; Calculate the relative fragmentation strength of each isomer It is used to reflect structural stability and diversity.

2. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, The dynamic adjustment of fragmentation parameters includes: adjusting the impact voltage to between 10eV and 20eV, and adjusting the cumulative time to between 0.2s and 1s.

3. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, The signal-to-noise ratio threshold is set to 4, 6, and 10 to filter valid peaks; the quality error is the allowable matching quality error range, set to 0.1 ppm, 0.2 ppm, and 0.5 ppm.

4. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, Define a library of functional groups, including: The functional group library includes a comprehensive functional group library or a simplified functional group library; the comprehensive functional group library includes: CO2, H2O, CO, SO2, SO3, C2H4O, CH3OH; the simplified functional group library includes: CO2, H2O, CO, CH3OH.

5. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, The output results include: Output all matching results for each step of neutral loss; the maximum number and intensity of functional group combinations for each parent ion; and a statistical table of functional group types.

6. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, The core metrics include: Molecular formula number recovery rate, which is the proportion of the number of identified fragments to the total number of fragments; intensity recovery rate, which is the proportion of the intensity of the resolved fragments to the total intensity.

7. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 1, characterized in that, The Bray-Curtis distance is calculated using the following formula (1). Where C ij x is the Bray-Curtis distance between sample i and sample j; ik x represents the k-th relative fragmentation strength in sample i; jk Let x be the k-th relative fragmentation strength in sample j; min(x) ik ,x jk ) represents the minimum value of the k-th feature in the two samples; the denominator is the sum of all features in the two samples.

8. The method for analyzing the molecular structure information of soluble organic matter based on high-resolution tandem mass spectrometry according to claim 7, characterized in that, The Spearman rank correlation coefficient is calculated as shown in equation (2). in d is the Spearman rank correlation coefficient, ranging from [-1, 1], representing the degree of monotonic correlation between variables; i Let be the rank difference of the i-th pair of data; n is the number of paired samples.

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