Multi-target tandem mass spectrometry analysis method, device, electronic equipment and storage medium

Abstract

The application relates to a multi-target tandem mass spectrometry method and device, electronic equipment and a storage medium. The method comprises the following steps: acquiring ion mobility-one mass spectrum of sample ions after ion mobility separation based on an ion mobility-tandem mass spectrometer; acquiring ion mobility-two mass spectrum of sample ions after ion mobility separation and wide window fragmentation based on the ion mobility-tandem mass spectrometer; performing peak detection and extraction on the ion mobility-one mass spectrum and the ion mobility-two mass spectrum to obtain an intensity matrix of parent ions and fragment ions in the mass-to-charge ratio and drift time dimensions; and performing deconvolution calculation on the intensity matrix of the parent ions and the fragment ions to obtain two mass spectra of multiple parent ions. Thus, the problems that related technologies cannot realize single-ion sampling multi-target tandem mass spectrometry, the sample utilization rate of tandem mass spectrometry is low, and the analysis coverage and structure analysis capability of mass spectrometry for molecules in trace samples are weak are solved.

Description

Technical Field

[0001] This application relates to the field of tandem mass spectrometry analysis technology, and in particular to a multi-target tandem mass spectrometry analysis method, apparatus, electronic device and storage medium. Background Technology

[0002] Mass spectrometry is a label-free, highly sensitive detection technique that can simultaneously detect a large number of compound molecules in a sample. High-resolution mass analyzers, such as Fourier transform ion cyclotron resonance mass analyzers and orbital trap mass analyzers, can obtain the precise molecular mass of a compound, providing information on its elemental composition. However, they cannot distinguish or identify isomers.

[0003] The combination of mass spectrometry and ion mobility separation techniques can distinguish ions with the same mass-to-charge ratio but different structures. However, accurate identification of ion structures depends on the establishment of an ion collision cross-sectional area database. Tandem mass spectrometry can achieve precise molecular structure identification; however, in conventional tandem mass spectrometry analysis, only one precursor ion can be analyzed in a single tandem mass spectrometry scan, leading to reduced sample utilization and prolonged analysis time. Targeted (data-dependent) tandem mass spectrometry sequentially isolates and analyzes different precursor ions within an analysis cycle; however, this analytical mode significantly increases sample consumption and prolongs analysis time, especially in the analysis of trace samples. For example, in mass spectrometry imaging and single-cell analysis, the number of times a single pixel (or single cell) can be sampled is limited, thus limiting the number of target precursor ions that can be analyzed in this mode.

[0004] Currently, relevant technologies allow data-independent tandem mass spectrometry (TAMS) based on liquid chromatography to be applied to proteomics and metabolomics, significantly improving the number of identified molecular structures and the stability of quantification. Applying similar data-independent tandem mass spectrometry methods in ion mobility mass spectrometry helps improve ion utilization per injection and resolve more molecular structures. Because ion mobility is a post-ionization separation technique with a separation timescale in the millisecond to second range, it is suitable for analyzing samples with trace amounts and short signal durations, especially in mass spectrometry imaging and single-cell analysis.

[0005] However, no non-target or multi-target tandem mass spectrometry analysis method for trace samples based on ion mobility-tandem mass spectrometry has been proposed, and related technologies cannot achieve tandem mass spectrometry analysis of multiple targets in a single ion sampling. The sample utilization rate of tandem mass spectrometry analysis is low, and the analytical coverage and structural resolution of molecules in trace samples by mass spectrometry are weak, which urgently need to be solved. Summary of the Invention

[0006] This application provides a multi-target tandem mass spectrometry analysis method, apparatus, electronic device, and storage medium to solve the problems of related technologies that cannot achieve tandem mass spectrometry analysis of multiple targets in a single ion sampling, low sample utilization rate of tandem mass spectrometry analysis, and weak analytical coverage and structural resolution capability of mass spectrometry for molecules in trace samples.

[0007] The first aspect of this application provides a multi-target tandem mass spectrometry analysis method, comprising the following steps: acquiring an ion mobility-first-order mass spectrum of sample ions after ion mobility separation based on an ion mobility-tandem mass spectrometer; acquiring an ion mobility-second-order mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation based on the ion mobility-tandem mass spectrometer; performing peak detection and extraction on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum to obtain the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and performing deconvolution calculation on the intensity matrices of the parent ion and fragment ions to obtain secondary mass spectra of multiple parent ions.

[0008] Optionally, in one embodiment of this application, the peak detection and extraction of the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum includes: performing two-dimensional feature detection on the ion mobility-first-order mass spectrum data in the time dimension and the mass-to-charge ratio dimension to obtain the features of multiple precursor ions; extracting the first column vector of the intensity change of the features of the multiple precursor ions over time to generate the intensity matrix of the precursor ions.

[0009] Optionally, in one embodiment of this application, the peak detection and extraction of the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum includes: performing one-dimensional feature detection on the ion mobility-second-order mass spectrum data in the mass-to-charge ratio dimension to obtain the features of multiple fragment ions; extracting the second column vector of the intensity of the features of the multiple fragment ions as a function of time to generate the intensity matrix of the fragment ions.

[0010] Optionally, in one embodiment of this application, the deconvolution calculation of the intensity matrices of the parent ion and fragment ions includes: solving a preset optimization problem based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, and calculating the total intensity of the parent ion to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

[0011] A second aspect of this application provides a multi-target tandem mass spectrometry analysis device, comprising: a first acquisition module for acquiring an ion mobility-first-order mass spectrum of sample ions after ion mobility separation based on an ion mobility-tandem mass spectrometer; a second acquisition module for acquiring an ion mobility-second-order mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation based on the ion mobility-tandem mass spectrometer; a third acquisition module for performing peak detection and extraction on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum to acquire the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and a fourth acquisition module for performing deconvolution calculation on the intensity matrices of the parent ion and fragment ions to acquire secondary mass spectra of multiple parent ions.

[0012] Optionally, in one embodiment of this application, the third acquisition module includes: a first detection unit, used to perform two-dimensional feature detection on ion mobility-first-order mass spectrometry data in the time dimension and mass-to-charge ratio dimension to obtain features of multiple precursor ions; and a first generation unit, used to extract a first column vector of the intensity of the features of the multiple precursor ions changing over time to generate an intensity matrix of the precursor ions.

[0013] Optionally, in one embodiment of this application, the third acquisition module further includes: a second detection unit, used to perform one-dimensional feature detection on the ion mobility-secondary mass spectrometry data in the mass-to-charge ratio dimension to obtain the features of multiple fragment ions; and a second generation unit, used to extract a second column vector of the intensity of the features of the multiple fragment ions as a function of time to generate the intensity matrix of the fragment ions.

[0014] Optionally, in one embodiment of this application, the fourth acquisition module includes: a calculation unit, used to solve a preset optimization problem based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, and calculate the total intensity of the parent ion, so as to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

[0015] A third aspect of this application provides an electronic device, including: a memory, a processor, and a computer program stored in the memory and executable on the processor, wherein the processor executes the program to implement the multi-target cascade mass spectrometry analysis method as described in the above embodiments.

[0016] A fourth aspect of this application provides a computer-readable storage medium storing a computer program that, when executed by a processor, implements the above-described multi-target cascade mass spectrometry analysis method.

[0017] Therefore, the embodiments of this application have the following beneficial effects:

[0018] The embodiments of this application can obtain an ion mobility-first-order mass spectrum of sample ions after ion mobility separation; obtain an ion mobility-second-order mass spectrum of sample ions after ion mobility separation and wide-window fragmentation; perform peak detection and extraction on the obtained first-order mass spectrum and ion mobility-second-order mass spectrum to obtain the intensity matrix of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and perform deconvolution calculation on the intensity matrix of the parent ion and fragment ions to obtain the second-order mass spectrum of multiple parent ions. Thus, this application improves the sample utilization rate of tandem mass spectrometry analysis, enables tandem mass spectrometry analysis of multiple targets in a single ion sampling, and significantly improves the analytical coverage, structural characterization ability and analysis speed of biomolecules in trace samples. In addition, this application is particularly suitable for tandem mass spectrometry analysis of non-target or multi-target substances in high-resolution mass spectrometry imaging and single-cell analysis to quickly obtain the structure and content of a large number of molecules. This solves the problems of related technologies being unable to achieve tandem mass spectrometry analysis of multiple targets in a single ion sampling, low sample utilization rate in tandem mass spectrometry analysis, and weak analytical coverage and structural resolution of molecules in trace samples.

[0019] Additional aspects and advantages of this application will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of this application. Attached Figure Description

[0020] The above and / or additional aspects and advantages of this application will become apparent and readily understood from the following description of the embodiments taken in conjunction with the accompanying drawings, wherein:

[0021] Figure 1 This is a flowchart of a multi-target cascade mass spectrometry analysis method provided according to an embodiment of this application;

[0022] Figure 2 A schematic diagram illustrating the principle of mass spectrometry imaging data acquisition and deconvolution is provided as an embodiment of this application;

[0023] Figure 3 A schematic diagram of a data acquisition system provided for one embodiment of this application;

[0024] Figure 4 A schematic diagram of a peak detection and spectral deconvolution process is provided as an embodiment of this application;

[0025] Figure 5 A schematic diagram of the result of spectral deconvolution is provided as an embodiment of this application;

[0026] Figure 6 A schematic diagram of a high-throughput cascade mass spectrometry imaging result provided for one embodiment of this application;

[0027] Figure 7This is an example diagram of a multi-target cascade mass spectrometry analysis apparatus according to an embodiment of this application;

[0028] Figure 8 This is a schematic diagram of the structure of an electronic device provided in an embodiment of this application.

[0029] Explanation of reference numerals in the attached figures: Multi-target cascade mass spectrometry analysis device-10; First acquisition module-100, Second acquisition module-200, Third acquisition module-300, Fourth acquisition module-400; Memory-801, Processor-802, Communication interface-803. Detailed Implementation

[0030] The embodiments of this application 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 intended to explain this application, and should not be construed as limiting this application.

[0031] The following description, with reference to the accompanying drawings, outlines a multi-target tandem mass spectrometry analysis method, apparatus, electronic device, and storage medium according to embodiments of this application. Addressing the problems mentioned in the background section, this application provides a multi-target tandem mass spectrometry analysis method. This method involves acquiring an ion mobility-first-order mass spectrum of sample ions after ion mobility separation; acquiring an ion mobility-second-order mass spectrum of sample ions after ion mobility separation and wide-window fragmentation; performing peak detection and extraction on the acquired first-order and second-order mass spectra to obtain the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and performing deconvolution calculations on the intensity matrices of the parent ion and fragment ions to obtain the second-order mass spectra of multiple parent ions. This application improves the sample utilization rate of tandem mass spectrometry analysis, enabling tandem mass spectrometry analysis of multiple targets in a single ion sampling. It significantly enhances the analytical coverage, structural characterization ability, and analytical speed for biomolecules in trace samples. Furthermore, this application is particularly suitable for tandem mass spectrometry analysis of non-targeted or multi-target substances in high-resolution mass spectrometry imaging and single-cell analysis, enabling rapid acquisition of the structure and content of a large number of molecules. This solves the problems of related technologies being unable to achieve tandem mass spectrometry analysis of multiple targets in a single ion sampling, low sample utilization rate in tandem mass spectrometry analysis, and weak analytical coverage and structural resolution of molecules in trace samples.

[0032] Specifically, Figure 1 This is a flowchart of a multi-target cascade mass spectrometry analysis method provided in an embodiment of this application.

[0033] like Figure 1 As shown, this multi-target cascade mass spectrometry analysis method includes the following steps:

[0034] In step S101, based on the ion mobility tandem mass spectrometer, the ion mobility-first-order mass spectrum of the sample ions after ion mobility separation is obtained.

[0035] The embodiments of this application can utilize an ion mobility-tandem mass spectrometer to obtain an ion mobility-first-order mass spectrum of sample ions after ion mobility separation.

[0036] It should be noted that any soft ionization source can be used for sample ionization. In the embodiments of this application, the DESI XS desorption electrospray ionization source from Waters can be used for sample ionization.

[0037] Furthermore, the above-mentioned ion mobility separation can be performed using an ion mobility spectrometer with time separation effect, such as a drift tube ion mobility spectrometer, a traveling wave ion mobility spectrometer, a captured ion mobility spectrometer, etc. In the embodiments of this application, a ring traveling wave ion mobility spectrometer and a time-of-flight mass spectrometer can be used to perform ion mobility separation and mass spectrometry analysis, respectively.

[0038] The ion mobility spectrometer and mass analyzer are arranged sequentially in space. Additionally, a quadrupole filter can be placed before the ion mobility spectrometer to isolate a range of ions by mass for subsequent analysis.

[0039] Specifically, in the embodiments of this application, frozen sections of mouse brain tissue can be used as samples, and the sample sections can be subjected to mass spectrometry imaging analysis using the aforementioned desorption electrospray ionization source.

[0040] like Figure 2 As shown, in the mass spectrometry imaging analysis of this embodiment, a spatial grid region is defined, where each small rectangular grid represents a pixel, and each pixel is further divided into two sub-pixels. The two sub-pixels are used to acquire ion mobility-primary mass spectrometry data and ion mobility-secondary mass spectrometry data, respectively. The size of the rectangular pixel is 50 μm × 100 μm, the size of the sub-pixels is 50 μm × 50 μm, and the spatial resolution is 50 μm × 100 μm.

[0041] It should be noted that the mass spectrometry imaging analysis method described above is also applicable to high-throughput single-cell analysis.

[0042] Understandably, since the ion mobility separation time is only a few hundred milliseconds, it does not significantly prolong the data acquisition time. Therefore, the embodiments of this application have the characteristics of high throughput and high sensitivity, and are particularly suitable for non-target or multi-target tandem mass spectrometry analysis of samples with trace amounts and short signal durations, such as mass spectrometry imaging and single-cell analysis.

[0043] In step S102, based on the ion mobility tandem mass spectrometer, the ion mobility-secondary mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation is obtained.

[0044] After obtaining the ion mobility-first-order mass spectrum of the sample ions after ion mobility separation, the embodiments of this application can further obtain the ion mobility-second-order mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation using an ion mobility-tandem mass spectrometer.

[0045] It should be noted that the embodiments of this application may employ collision-induced dissociation technology, using a fixed excitation energy, to achieve wide-window ion fragmentation, i.e., ion excitation, between the ion migration analyzer and the mass analyzer.

[0046] In the specific implementation process, those skilled in the art can also achieve ion excitation through any technology with ion fragmentation function, such as ultraviolet light dissociation, surface-induced dissociation, electron capture-induced dissociation, or electron transfer-induced dissociation, without making any specific restrictions.

[0047] Furthermore, the data acquisition system used in the embodiments of this application is a Waters Cyclic IMS mass spectrometer, such as... Figure 3 As shown. The main components of this system include an ion source (desorption electrospray ionization source (DESI)), a quadrupole mass filter, a ring ion mobility spectrometer, a collision cell, and a time-of-flight mass analyzer.

[0048] The DESI ion source is used to ionize compounds on the sample surface, with a spatial resolution greater than 30 μm, preferably 50 μm; the quadrupole mass filter is used to screen ions within a specific mass-to-charge ratio range; the ring ion mobility spectrometer is used to separate the ion mobility of analytes in the sample, with a maximum mobility resolution of approximately 750, preferably 50–100; the collision cell is used for collision-induced dissociation of ions; and the time-of-flight mass analyzer is used to detect the ion mass-to-charge ratio, with a maximum mass resolution of approximately 100,000, preferably 50,000.

[0049] Therefore, the embodiments of this application use an ion mobility-tandem mass spectrometer to obtain ion mobility-secondary mass spectra of sample ions after ion mobility separation and wide-window fragmentation, providing a basis for subsequent deconvolution calculations and effectively ensuring the performance of multi-target tandem mass spectrometry analysis.

[0050] In step S103, peak detection and extraction are performed on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum to obtain the intensity matrix of the parent ion and fragment ions in the dimensions of mass-to-charge ratio and drift time.

[0051] After obtaining the ion mobility-primary mass spectrum and the ion mobility-secondary mass spectrum, since the data formats output by the mass spectrometry instrument software from different manufacturers are different, the raw data file can be converted into a .mzML format data file using the open-source software MSConvert. In the embodiments of this application, the .raw format data from Waters can be converted into a .mzML format data file.

[0052] It should be noted that the analysis method in this application embodiment is also applicable to data collected from instruments manufactured by other instrument manufacturers.

[0053] Furthermore, embodiments of this application can use the open-source Python library pymzml to process .mzML format data files, converting them into .mat format data files for subsequent processing using MATLAB scripts, such as... Figure 4 As shown. The .mat data file contains ion mobility-primary mass spectrometry data and ion mobility-secondary mass spectrometry data.

[0054] Therefore, the above data preprocessing operations provide a reliable data basis for peak detection and extraction in subsequent ion mobility-first-order mass spectra and ion mobility-second-order mass spectra.

[0055] Optionally, in one embodiment of this application, peak detection and extraction are performed on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum, including: performing two-dimensional feature detection on the ion mobility-first-order mass spectrum data in the time dimension and mass-to-charge ratio dimension to obtain the features of multiple precursor ions; extracting the first column vector of the intensity change of the features of multiple precursor ions over time to generate the intensity matrix of the precursor ions.

[0056] It should be noted that the embodiments of this application can perform two-dimensional feature detection on the ion mobility-first-order mass spectrometry data obtained above in the time dimension and mass-to-charge ratio dimension to obtain the features of N parent ions, and extract the column vector p of the intensity change of the N features over time. i (i = 1, 2, ..., N), N column vectors arranged into a matrix P T×N , where T represents the number of time points for ion mobility sampling.

[0057] Specifically, in the embodiments of this application, the process of performing two-dimensional feature detection on ion mobility-first-order mass spectrometry data is as follows:

[0058] 1. Detect the top N data points in terms of intensity in both the mass-to-charge ratio and drift time dimensions;

[0059] 2. For each detected data point, extract the ion migration spectrum at ±0.01Th and ±10ms.

[0060] 3. Arrange all migration spectra to obtain a data matrix P of N parent ions. T×N .

[0061] This enables two-dimensional feature detection of ion mobility-first-order mass spectrometry data, ensuring the reliability of second-order mass spectra of multiple precursor ions.

[0062] Optionally, in one embodiment of this application, peak detection and extraction are performed on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum, including: performing one-dimensional feature detection on the ion mobility-second-order mass spectrum data in the mass-to-charge ratio dimension to obtain the features of multiple fragment ions; extracting the second column vector of the intensity of the features of multiple fragment ions as a function of time to generate the intensity matrix of fragment ions.

[0063] Furthermore, embodiments of this application can perform one-dimensional feature detection on the ion mobility-secondary mass spectrometry data obtained above in the mass-to-charge ratio dimension to obtain the features of M fragment ions, and extract the column vector f of the intensity change of the M features over time. j (j = 1, 2, ..., M), M column vectors arranged into matrix F T×M .

[0064] Based on the matrix P obtained above T×N sum matrix F T×M We can obtain the following formula:

[0065] F T×M =P T×N ·C N×M ,

[0066] Among them, C N×M The element in the i-th row and j-th column represents the proportion of the mass spectrometry signal intensity of the j-th fragment ion generated by the fragmentation of the i-th parent ion under a specific excitation energy.

[0067] As one possible approach, the process of performing one-dimensional feature detection on ion mobility-secondary mass spectrometry data according to embodiments of this application is described below:

[0068] 1. Detect the data points of intensity M in the mass-to-charge ratio dimension;

[0069] 2. For each detected data point, extract the migration spectra of ions within a 0.01Th range to its left and right, and arrange them into a data matrix F of M fragment ions. T×M .

[0070] Therefore, the embodiments of this application effectively ensure the rapid determination and structural characterization of a large number of molecules in the sample by detecting and extracting spectral peaks from ion mobility-first-order mass spectra and ion mobility-second-order mass spectra.

[0071] In step S104, the intensity matrices of the parent ion and fragment ions are deconvolved to obtain secondary mass spectra of multiple parent ions.

[0072] After obtaining the intensity matrices of the parent ion and fragment ions in terms of mass-to-charge ratio and drift time, embodiments of this application can further perform deconvolution calculations on the intensity matrices of the parent ion and fragment ions to obtain secondary mass spectra of multiple parent ions.

[0073] It is understood that the embodiments of this application achieve non-targeted tandem mass spectrometry data acquisition by utilizing ion mobility separation and wide-window ion fragmentation. Based on the mathematical relationship between the intensity of the parent ion and fragment ions, the secondary spectrum of the parent ion is reconstructed, thereby achieving nearly 100% ion utilization and effectively improving the detection sensitivity of tandem mass spectrometry analysis.

[0074] Optionally, in one embodiment of this application, the deconvolution calculation of the intensity matrices of the parent ion and fragment ions includes: solving a preset optimization problem based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, and calculating the total intensity of the parent ion to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

[0075] It should be noted that the embodiments of this application can utilize the non-negative least squares method, as shown in the following equation, to solve the optimization problem:

[0076] argmin C ||FP·C||,subject to C≥0

[0077]

[0078] Thus, an approximate solution to matrix C is obtained. To obtain the coefficient matrix C for the generation of fragment ions from the parent ion. N×M ,in, Each row represents a normalized second-order mass spectrum of different parent ions.

[0079] It should be noted that, in the specific implementation process, those skilled in the art can also solve the optimization problem through methods such as nonnegative matrix factorization, and no specific restrictions are imposed here.

[0080] Furthermore, based on vector p i The intensity of the parent ion can be obtained by solving the peak area using the following formula:

[0081]

[0082] And calculate the absolute intensity of the i-th parent ion producing the j-th fragment:

[0083] IF j=IP i ·c ij

[0084] Furthermore, embodiments of this application can perform pairwise cross-correlation analysis on the migration spectra of all parent ions to obtain the distance d between migration peaks and the peak shape similarity s. If d and s satisfy the relationship with the set thresholds d0 and s0: d < d0 and s > s0, then the migration peaks are merged to obtain a feature, and the absolute intensities of fragment ions are merged to obtain a new coefficient matrix. The merger process is as follows:

[0085] 1. The set of features to be merged includes those numbered i1, i2, ..., i l The parent ions have absolute mass spectrometry signal intensities of [missing information]. Then the i1, i2, ..., i-th elements of the fragmentation coefficient matrix l The rows are weighted averages based on the absolute intensity of the parent ion signal and merged into one row:

[0086]

[0087] The absolute intensity of the i-th parent ion producing the j-th fragment ion is I. i ·c ij , where I i This represents the absolute intensity of the i-th parent ion;

[0088] 2. Normalize the total ion intensity of the pixel to obtain the normalized intensity of the ion. Use the normalized intensity or the square root or logarithm of the normalized intensity as the pixel value to plot the mass spectrometry imaging results of the parent ion and fragment ions.

[0089] In summary, the embodiments of this application calculate the cross-correlation of the intensity change vectors of all precursor ion features over time. The offset of the maximum cross-correlation value represents the distance between migration peaks, and the maximum cross-correlation value represents the similarity of the migration peak shapes. If the distance between migration peaks is less than a set threshold, and the peak similarity is greater than a set threshold, then the two features and their corresponding fragment ions are merged into one feature, and the intensities of the corresponding fragment ions are combined.

[0090] Therefore, the embodiments of this application can increase the ion utilization rate in traditional tandem mass spectrometry analysis from 1 / N (where N is the number of precursor ions to be measured) to nearly 100%, thereby enabling the analysis of the structural information of multiple precursor ions in a single injection, effectively improving the analytical coverage, structural characterization ability, and analysis speed of biomolecules in trace samples.

[0091] The data deconvolution method proposed in the embodiments of this application does not rely on a database and has the characteristics of simplicity, speed, accuracy, and automation. It can also be applied to the deconvolution of data based on data acquisition methods such as liquid chromatography-tandem mass spectrometry, gas chromatography-electron ionization mass spectrometry, and capillary electrophoresis-tandem mass spectrometry, and has strong versatility.

[0092] The multi-target cascade mass spectrometry analysis method of this application will be described below with reference to the accompanying drawings and specific embodiments.

[0093] Specifically, in this embodiment, mass spectrometry analysis of frozen sections of mouse brain tissue in negative mode is performed using a desorption electrospray ionization source. Secondary mass spectrometry is deconvoluted at individual pixels, resulting in secondary mass spectra of four different parent ions obtained in a single mass spectrometry analysis, such as... Figure 5 As shown, the second-order mass spectrum obtained by deconvolution using the non-targeted cascade mass spectrometry method is highly similar to the second-order mass spectrum obtained by targeted cascade mass spectrometry.

[0094] Furthermore, embodiments of this application also use DESI to perform mass spectrometry imaging analysis on frozen sections of mouse brain tissue in positive mode. The defined mass spectrometry imaging region is 4.5 mm × 9 mm, with a pixel size of 50 μm × 100 μm. Each pixel is further divided into two sub-pixels, each 50 μm × 50 μm in size. The left sub-pixel is used to acquire ion mobility-mass spectrometry data, and the right sub-pixel is used to acquire ion mobility-secondary mass spectrometry data. The mass range of the quadrupole filter is 730–880 Th. A single mass spectrometry imaging experiment yielded secondary mass spectra of 21 different parent ions after deconvolution. Some representative secondary mass spectrometry imaging results are shown below. Figure 6 As shown. Figure 6 The first column shows the mass spectrometry imaging results of the parent ion directly reconstructed from ion mobility-first-order mass spectrometry data; the second column shows the mass spectrometry imaging results of the parent ion directly reconstructed from ion mobility-second-order mass spectrometry data; and the third (fourth) column shows the mass spectrometry imaging results of the corresponding fragment ions after deconvolution. Fragment ions can be used for the structural identification of the parent ion. Simultaneously, this invention can also resolve the structure of biomolecular isomers or the spatial distribution of isomers of the same weight. For example, for an ion with a mass-to-charge ratio of 782Th, its second-order mass spectrometry fragment ions have two spatial distributions, corresponding to [PC 34: 1+Na] respectively. + (782.5670Th) and [PC 36:4+H+(782.5694Th) are objects of the same weight. The mass-to-charge ratio separation requires a resolution of over 300,000, which cannot be distinguished by time-of-flight mass spectrometry alone. However, this application can simultaneously achieve structural identification and spatial distribution characterization.

[0095] The multi-target tandem mass spectrometry analysis method proposed in this application involves obtaining an ion mobility-first-level mass spectrum of sample ions after ion mobility separation; obtaining an ion mobility-second-level mass spectrum of sample ions after ion mobility separation and wide-window fragmentation; performing peak detection and extraction on the obtained first-level and second-level mass spectra to obtain the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and performing deconvolution calculation on the intensity matrices of the parent ion and fragment ions to obtain the second-level mass spectra of multiple parent ions. This method improves the sample utilization rate of tandem mass spectrometry analysis, enabling tandem mass spectrometry analysis of multiple targets in a single ion sampling, significantly improving the analytical coverage, structural characterization ability, and analysis speed of biomolecules in trace samples. Furthermore, this method is particularly suitable for tandem mass spectrometry analysis of non-target or multi-target substances in high-resolution mass spectrometry imaging and single-cell analysis, enabling rapid acquisition of the structure and content of a large number of molecules.

[0096] Next, referring to the accompanying drawings, a multi-target cascade mass spectrometry analysis apparatus according to an embodiment of this application is described.

[0097] Figure 7 This is a block diagram of a multi-target cascade mass spectrometry analysis device according to an embodiment of this application.

[0098] like Figure 7 As shown, the multi-target cascade mass spectrometry analysis device 10 includes: a first acquisition module 100, a second acquisition module 200, a third acquisition module 300, and a fourth acquisition module 400.

[0099] The first acquisition module is used to acquire the ion mobility-first-order mass spectrum of sample ions after ion mobility separation based on the ion mobility-tandem mass spectrometer.

[0100] The second acquisition module is used to acquire the ion mobility-secondary mass spectrum of sample ions after ion mobility separation and wide-window fragmentation based on the ion mobility-tandem mass spectrometer.

[0101] The third acquisition module is used to detect and extract peaks from the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum, and to obtain the intensity matrix of the parent ion and fragment ions in the dimensions of mass-to-charge ratio and drift time.

[0102] The fourth acquisition module is used to perform deconvolution calculations on the intensity matrices of the parent ion and fragment ions to obtain secondary mass spectra of multiple parent ions.

[0103] Optionally, in one embodiment of this application, the third acquisition module 300 includes: a first detection unit and a first generation unit.

[0104] The first detection unit is used to perform two-dimensional feature detection on the ion mobility-first-order mass spectrometry data in the time dimension and mass-to-charge ratio dimension to obtain the features of multiple parent ions.

[0105] The first generation unit is used to extract the first column vector of the intensity changes of multiple precursor ions over time, and generate the intensity matrix of the precursor ions.

[0106] Optionally, in one embodiment of this application, the third acquisition module 300 further includes: a second detection unit and a second generation unit.

[0107] The second detection unit is used to perform one-dimensional feature detection on the ion mobility-secondary mass spectrometry data in the mass-to-charge ratio dimension to obtain the features of multiple fragment ions.

[0108] The second generation unit is used to extract the second column vector of the intensity of multiple fragment ions as a function of time, and generate the intensity matrix of the fragment ions.

[0109] Optionally, in one embodiment of this application, the fourth acquisition module 400 includes: a calculation unit, used to solve a preset optimization problem based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, and to calculate the total intensity of the parent ion, so as to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

[0110] It should be noted that the foregoing explanation of the multi-target cascade mass spectrometry analysis method embodiment also applies to the multi-target cascade mass spectrometry analysis device of this embodiment, and will not be repeated here.

[0111] According to the multi-target tandem mass spectrometry analysis device proposed in the embodiments of this application, the following steps are taken: First, an ion mobility-first-order mass spectrum of sample ions after ion mobility separation is obtained; second, an ion mobility-second-order mass spectrum of sample ions after ion mobility separation and wide-window fragmentation is obtained; peak detection and extraction are performed on the obtained first-order mass spectrum and ion mobility-second-order mass spectrum to obtain the intensity matrix of the parent ion and fragment ions in the dimensions of mass-to-charge ratio and drift time; third, deconvolution calculation is performed on the intensity matrix of the parent ion and fragment ions to obtain the second-order mass spectrum of multiple parent ions. Thus, this application improves the sample utilization rate of tandem mass spectrometry analysis, enabling tandem mass spectrometry analysis of multiple targets in a single ion sampling, significantly improving the analytical coverage, structural characterization ability, and analysis speed of biomolecules in trace samples. Furthermore, this application is particularly suitable for tandem mass spectrometry analysis of non-target or multi-target substances in high-resolution mass spectrometry imaging and single-cell analysis, to rapidly obtain the structure and content of a large number of molecules.

[0112] Figure 8 A schematic diagram of the structure of an electronic device provided in an embodiment of this application. The electronic device may include:

[0113] The memory 801, the processor 802, and the computer program stored on the memory 801 and capable of running on the processor 802.

[0114] When the processor 802 executes the program, it implements the multi-target cascade mass spectrometry analysis method provided in the above embodiments.

[0115] Furthermore, electronic devices also include:

[0116] Communication interface 803 is used for communication between memory 801 and processor 802.

[0117] The memory 801 is used to store computer programs that can run on the processor 802.

[0118] The memory 801 may include high-speed RAM memory, and may also include non-volatile memory, such as at least one disk storage device.

[0119] If the memory 801, processor 802, and communication interface 803 are implemented independently, then the communication interface 803, memory 801, and processor 802 can be interconnected via a bus to complete communication between them. The bus can be an Industry Standard Architecture (ISA) bus, a Peripheral Component Interconnect (PCI) bus, or an Extended Industry Standard Architecture (EISA) bus, etc. Buses can be divided into address buses, data buses, control buses, etc. For ease of representation, Figure 8 The bus is represented by a single thick line, but this does not mean that there is only one bus or one type of bus.

[0120] Optionally, in a specific implementation, if the memory 801, processor 802, and communication interface 803 are integrated on a single chip, then the memory 801, processor 802, and communication interface 803 can communicate with each other through an internal interface.

[0121] The processor 802 may be a central processing unit (CPU), an application specific integrated circuit (ASIC), or one or more integrated circuits configured to implement the embodiments of this application.

[0122] This application also provides a computer-readable storage medium storing a computer program thereon, which, when executed by a processor, implements the above-described multi-target cascade mass spectrometry analysis method.

[0123] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of this application. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of different embodiments or examples.

[0124] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this application, "N" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0125] Any process or method described in the flowchart or otherwise herein can be understood as representing a module, segment, or portion of code comprising one or N executable instructions for implementing custom logic functions or processes, and the scope of the preferred embodiments of this application includes additional implementations in which functions may be performed not in the order shown or discussed, including substantially simultaneously or in reverse order depending on the functions involved, as should be understood by those skilled in the art to which embodiments of this application pertain.

[0126] The logic and / or steps represented in the flowchart or otherwise described herein, for example, can be considered as a sequenced list of executable instructions for implementing logical functions, and can be embodied in any computer-readable medium for use by, or in conjunction with, an instruction execution system, apparatus, or device (such as a computer-based system, a processor-included system, or other system that can fetch and execute instructions from, an instruction execution system, apparatus, or device). For the purposes of this specification, "computer-readable medium" can be any means that can contain, store, communicate, propagate, or transmit programs for use by, or in conjunction with, an instruction execution system, apparatus, or device. More specific examples (a non-exhaustive list) of computer-readable media include: an electrical connection having one or more wires (electronic device), a portable computer disk drive (magnetic device), random access memory (RAM), read-only memory (ROM), erasable and editable read-only memory (EPROM or flash memory), fiber optic devices, and portable optical disc read-only memory (CDROM). Alternatively, the computer-readable medium may be paper or other suitable media on which the program can be printed, since the program can be obtained electronically by optically scanning the paper or other medium, followed by editing, interpreting, or otherwise processing as necessary, and then stored in a computer memory.

[0127] It should be understood that the various parts of this application can be implemented using hardware, software, firmware, or a combination thereof. In the above embodiments, the N steps or methods can be implemented using software or firmware stored in memory and executed by a suitable instruction execution system. If implemented in hardware, as in another embodiment, it can be implemented using any one or a combination of the following techniques known in the art: discrete logic circuits having logic gates for implementing logical functions on data signals, application-specific integrated circuits (ASICs) having suitable combinational logic gates, programmable gate arrays (PGAs), field-programmable gate arrays (FPGAs), etc.

[0128] Those skilled in the art will understand that all or part of the steps of the methods in the above embodiments can be implemented by a program instructing related hardware. The program can be stored in a computer-readable storage medium, and when executed, the program includes one or a combination of the steps of the method embodiments.

[0129] Furthermore, the functional units in the various embodiments of this application can be integrated into a processing module, or each unit can exist physically separately, or two or more units can be integrated into a module. The integrated module can be implemented in hardware or as a software functional module. If the integrated module is implemented as a software functional module and sold or used as an independent product, it can also be stored in a computer-readable storage medium.

[0130] The storage medium mentioned above can be a read-only memory, a disk, or an optical disk, etc. Although embodiments of this application have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting this application. Those skilled in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of this application.

Claims

1. A multi-target cascade mass spectrometry analysis method, characterized in that, Includes the following steps: Based on ion mobility-tandem mass spectrometry, the ion mobility-first-order mass spectrum of sample ions after ion mobility separation is obtained. Based on the ion mobility tandem mass spectrometer, the ion mobility-secondary mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation was obtained. Peak detection and extraction were performed on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum to obtain the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and The intensity matrices of the parent ion and fragment ions are deconvolved to obtain secondary mass spectra of multiple parent ions.

2. The method according to claim 1, characterized in that, The peak detection and extraction of the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum includes: Two-dimensional feature detection was performed on the ion mobility-first-order mass spectrometry data in the time and mass-to-charge ratio dimensions to obtain the characteristics of multiple parent ions. Extract the first column vector of the intensity variation of the features of the multiple precursor ions over time to generate the intensity matrix of the precursor ions.

3. The method according to claim 2, characterized in that, The peak detection and extraction of the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum includes: One-dimensional feature detection was performed on the ion mobility-secondary mass spectrometry data in the mass-to-charge ratio dimension to obtain the characteristics of multiple fragment ions. The intensity matrix of the fragment ions is generated by extracting the second column vector of the intensity variation over time of the features of the multiple fragment ions.

4. The method according to claim 3, characterized in that, The deconvolution calculation of the intensity matrices of the parent ion and fragment ions includes: Based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, a preset optimization problem is solved, and the total intensity of the parent ion is calculated to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

5. A multi-target cascade mass spectrometry analysis device, characterized in that, include: The first acquisition module is used to acquire the ion mobility-first-order mass spectrum of sample ions after ion mobility separation based on the ion mobility-tandem mass spectrometer. The second acquisition module is used to acquire the ion mobility-secondary mass spectrum of the sample ions after ion mobility separation and wide-window fragmentation based on the ion mobility-tandem mass spectrometer. The third acquisition module is used to perform peak detection and extraction on the ion mobility-first-order mass spectrum and the ion mobility-second-order mass spectrum to obtain the intensity matrices of the parent ion and fragment ions in the mass-to-charge ratio and drift time dimensions; and The fourth acquisition module is used to perform deconvolution calculations on the intensity matrices of the parent ion and fragment ions to obtain secondary mass spectra of multiple parent ions.

6. The apparatus according to claim 5, characterized in that, The third acquisition module includes: The first detection unit is used to perform two-dimensional feature detection on ion mobility-first-level mass spectrometry data in the time dimension and mass-to-charge ratio dimension to obtain the characteristics of multiple parent ions. The generation unit is used to extract the first column vector of the intensity change of the features of the multiple parent ions over time, and generate the intensity matrix of the parent ions.

7. The apparatus according to claim 6, characterized in that, The third acquisition module further includes: The second detection unit is used to perform one-dimensional feature detection on the ion mobility-secondary mass spectrometry data in the mass-to-charge ratio dimension to obtain the characteristics of multiple fragment ions. The second generation unit is used to extract the second column vector of the intensity of the features of the multiple fragment ions as a function of time, and generate the intensity matrix of the fragment ions.

8. The apparatus according to claim 7, characterized in that, The fourth acquisition module includes: The calculation unit is used to solve a preset optimization problem based on the intensity matrix of the parent ion and the intensity matrix of the fragment ions, and to calculate the total intensity of the parent ion, so as to calculate the distance and peak shape similarity of all parent ion characteristic migration peaks.

9. An electronic device, characterized in that, include: A memory, a processor, and a computer program stored in the memory and executable on the processor, the processor executing the program to implement the multi-target cascade mass spectrometry analysis method as described in any one of claims 1-4.

10. A computer-readable storage medium having a computer program stored thereon, characterized in that, The program is executed by the processor to implement the multi-target cascade mass spectrometry analysis method as described in any one of claims 1-4.

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