Method and device for performing energy resolution tandem mass spectrometry on sample based on ion trap

Abstract

The invention discloses a method and a device for performing energy resolution cascade mass spectrometry on a sample based on an ion trap, and the method comprises the following steps: performing ionization treatment on a to-be-detected sample to obtain to-be-detected ions; enabling the to-be-detected ions to enter a linear ion trap, and screening out target ions; amplitude scanning processing is carried out on the target ions, and the amplitude scanning processing comprises the steps that first alternating voltage is applied to the x direction of the linear ion trap so that the target ions can be cracked into daughter ions, and meanwhile second alternating voltage is applied to the y direction of the linear ion trap so that the daughter ions can be separated from the linear ion trap; enabling a detector to obtain the daughter ions so as to obtain a detection result of the to-be-detected sample. According to the method disclosed by the invention, continuous energy scanning and fragmentation of parent ions can be realized through the first alternating-current voltage amplitude which is increased along with time by utilizing a single ion trap and only through single sample introduction, and daughter ions are synchronously emitted by utilizing the constant second alternating-current voltage amplitude in the orthogonal direction for detection; and the efficiency and the accuracy of energy resolution cascade mass spectrometry are improved.

Description

Technical Field

[0001] The present application relates to the field of analysis. Specifically, the present application relates to a method and apparatus for energy-resolved tandem mass spectrometry analysis of a sample based on an ion trap. Background Art

[0002] Energy-resolved tandem mass spectrometry technology (Energy-resolved MS / MS, ER-MS / MS) refers to obtaining energy-related fragment ion information by precisely controlling the energy parameters during the ion activation process, so as to more comprehensively analyze the structure of compounds. In a traditional triple quadrupole mass spectrometer, energy control is mainly achieved by adjusting the collision energy (CE). Generally speaking, energy-resolved tandem mass spectrometry technology mainly establishes the fragmentation curve of precursor ions - product ions (i.e., the correlation curve between relative ion abundance and collision energy) through multiple injections, respectively fits the fragmentation curves of precursor ions and product ions using Sigmoid curves and Gaussian curves, and further extracts the fragmentation energy information of ions through the collision energy at half response of precursor ions (CE 50 ).) or the optimal collision energy (OCE) of product ions.

[0003] When a traditional triple quadrupole mass spectrometer performs energy-resolved tandem mass spectrometry analysis, it is necessary to perform multiple injections to establish the fragmentation curve of precursor ions - product ions, and extract the fragmentation energy information by plotting the product ion intensity - energy curve point by point. This is not only cumbersome in operation, but also has fitting errors. Multiple injections result in a long measurement time, reducing the analysis efficiency, and at the same time, the utilization rate of the sample is insufficient, causing waste of the sample.

[0004] Therefore, the current methods for energy-resolved tandem mass spectrometry still need to be improved. Summary of the Invention

[0005] The present application aims to solve at least one of the technical problems existing in the prior art to some extent. For this purpose, the present application provides a method and apparatus for energy-resolved tandem mass spectrometry of a sample based on an ion trap. The method of the present application uses a single ion trap, and only requires a single injection to achieve continuous energy scanning and fragmentation of precursor ions through an AC voltage amplitude that increases with time, and simultaneously uses a constant AC voltage in the orthogonal direction to eject product ions for detection, improving the efficiency and accuracy of mass spectrometry analysis.

[0006] In the first aspect of the present application, a method for energy-resolved tandem mass spectrometry of a sample based on an ion trap is proposed, including: ionizing a sample to be measured to obtain ions to be measured; introducing the ions to be measured into a linear ion trap and screening out target ions; performing amplitude scanning on the target ions, and the amplitude scanning includes: applying a first alternating voltage in the x direction of the linear ion trap to cause the target ions to fragment into daughter ions, and at the same time applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; enabling a detector to acquire the daughter ions to obtain a detection result of the sample to be measured. Thus, by increasing the amplitude of the first alternating voltage with time in a single ion trap, continuous energy scanning and fragmentation of the parent ions can be achieved with a single injection, and at the same time, the constant second alternating voltage is used to excite the daughter ions for detection, enabling the fragmentation of the parent ions and the detection of the daughter ions to be synchronized, improving the efficiency and accuracy of mass spectrometry analysis.

[0007] According to an embodiment of the present application, the first alternating voltage is a sine voltage.

[0008] According to an embodiment of the present application, the second alternating voltage is a sine voltage.

[0009] According to an embodiment of the present application, the frequency of the first alternating voltage is the same as the long-term frequency when the target ions are isolated in the linear ion trap.

[0010] According to an embodiment of the present application, the frequency of the second alternating voltage is the same as the long-term frequency when the daughter ions are isolated in the linear ion trap.

[0011] According to an embodiment of the present application, the x direction and the y direction are orthogonal to each other.

[0012] According to an embodiment of the present application, the amplitude of the first alternating voltage increases with the time of the amplitude scanning process.

[0013] According to an embodiment of the present application, the amplitude of the first alternating voltage increases from 0V to 0.5 - 1V.

[0014] According to an embodiment of the present application, the amplitude of the second alternating voltage remains constant.

[0015] According to an embodiment of the present application, the amplitude of the second alternating voltage is 0.5 - 1.5V.

[0016] According to an embodiment of the present application, before performing the amplitude scanning process, a collision gas is introduced into the linear ion trap.

[0017] According to an embodiment of the present application, the time of the amplitude scanning process is 400 - 600ms.

[0018] In a second aspect of the present application, there is provided an apparatus for energy-resolved tandem mass spectrometry of a sample based on an ion trap, comprising: an injection module for ionizing a sample to be measured to obtain ions to be measured; a screening module for allowing the ions to be measured to enter a linear ion trap and screening out target ions; a fragmentation module for applying a first alternating voltage in the x direction of the linear ion trap to fragment the target ions into daughter ions; an ejection module for simultaneously applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; and a detection module for enabling a detector to acquire the daughter ions to obtain a detection result of the sample to be measured.

[0019] Additional aspects and advantages of the present application will be given in part in the following description, become apparent in part from the following description, or be learned through the practice of the present application. BRIEF DESCRIPTION OF THE DRAWINGS

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

[0021] Figure 1 is a schematic flowchart of a method for energy-resolved tandem mass spectrometry according to an embodiment of the present application;

[0022] Figure 2 is a schematic diagram of the principle of energy-resolved tandem mass spectrometry scanning according to an embodiment of the present application;

[0023] Figure 3 is the structure of the thermometer ion used in Embodiment 1 of the present application;

[0024] Figure 4 is a schematic diagram of the control device of the mass spectrometer for non-continuous injection in Embodiment 1 of the present application;

[0025] Figure 5 is a graph of the energy-resolved tandem mass spectrometry results in Embodiment 1 of the present application;

[0026] Figure 6 is a graph of the signal verification results in Comparative Example 1 of the present application; where Figure 6 (A) is a waveform diagram of ACx; Figure 6 (B) is the detection signal when ACy is turned off; Figure 6 (C) is the detection signal when ACy is turned on. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0027] Embodiments of the present application will be described in detail below. The following described embodiments are exemplary and are only used to explain the present application, and should not be construed as a limitation to the present application.

[0028] It should be noted that the terms "first" and "second" are only used for descriptive purposes and should not be construed as indicating or implying relative importance or implicitly specifying the quantity of the indicated technical features. Thus, features defined with "first" and "second" may explicitly or implicitly include one or more of such features. Further, in the description of the present application, unless otherwise specified, the meaning of "a plurality" is two or more.

[0029] The endpoints and any values in the ranges disclosed herein are not limited to the exact ranges or values, and these ranges or values should be understood to include values close to these ranges or values. For numerical ranges, between the endpoint values of each range, between the endpoint values of each range and individual point values, and between individual point values, they can be combined with each other to obtain one or more new numerical ranges, and these numerical ranges should be regarded as specifically disclosed herein.

[0030] In this document, the term "comprising" or "including" is an open-ended expression, that is, it includes the content specified in the present application, but does not exclude other aspects of the content.

[0031] In this document, the terms "optionally", "optional" or "option" generally mean that the subsequent events or conditions may but do not necessarily occur, and this description includes the cases where such events or conditions occur, as well as the cases where such events or conditions do not occur.

[0032] In the first aspect of the present application, the present application provides a method for energy-resolved tandem mass spectrometry of a sample based on an ion trap, including: ionizing a sample to be measured to obtain ions to be measured; making the ions to be measured enter a linear ion trap and screening out target ions; performing amplitude scanning processing on the target ions, and the amplitude scanning processing includes: applying a first alternating voltage in the x direction of the linear ion trap to cause the target ions to fragment into daughter ions, and at the same time applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; making a detector acquire the daughter ions to obtain a detection result of the sample to be measured. Thus, by the increasing amplitude of the first alternating voltage with time of a single ion trap, continuous energy scanning and fragmentation of the parent ions can be achieved with a single injection, and at the same time, the constant second alternating voltage is used to excite the daughter ions for detection, enabling the fragmentation of the parent ions and the detection of the daughter ions to be synchronized, improving the efficiency and accuracy of mass spectrometry analysis.

[0033] Specifically, referring to Figure 1 , the method for energy resolution of a sample based on an ion trap includes:

[0034] Sample introduction

[0035] According to some specific embodiments of the present application, in this step, the sample to be tested is ionized to obtain the ions to be tested. The method of ionization is not particularly limited, as long as it can ionize the molecules to be tested, and those skilled in the art can select it according to actual circumstances. For example, electrospray ionization, atmospheric pressure chemical ionization, electron bombardment ionization, etc. can be used.

[0036] Ion trapping

[0037] According to some specific embodiments of the present application, in this step, the ions to be detected are caused to enter a linear ion trap. Specifically, the ions to be detected include impurity ions in addition to the target ions.

[0038] Ion Isolation

[0039] According to some specific embodiments of the present application, in this step, an AC voltage with the same frequency as that of the impurity ions is applied to the linear ion trap, so that the impurity ions are excited to fly out of the ion trap, leaving only the target ions trapped in the linear ion trap.

[0040] Adjust air pressure

[0041] According to some specific embodiments of the present application, a collision gas is introduced into the linear ion trap in this step. The collision gas is not particularly limited, as long as it is used for subsequent collision-induced fragmentation. For example, air, nitrogen, helium, argon, etc. can be used.

[0042] Amplitude-scanned collisional fragmentation

[0043] According to some specific embodiments of the present application, in this step, a first AC voltage is applied in the x direction of the linear ion trap to cause the target ions to be fragmented into daughter ions. Figure 2 In a linear ion trap, a pair of electrodes applies a first AC voltage in the x-direction, and the amplitude of the first AC voltage increases with time. The ions are subjected to resonance excitation that gradually increases with time, and the collision energy also increases with time. This is accomplished using a single ion trap, and a single injection can capture the fragmentation information of parent ions in different energy states.

[0044] Simultaneous detection of specific product ions

[0045] According to some specific embodiments of the present application, in this step, a second AC voltage is simultaneously applied in the y direction of the linear ion trap to allow the daughter ions to escape from the linear ion trap. Figure 2, an alternating voltage in the y direction is applied to another pair of electrodes in the linear ion trap, and the amplitude of the second alternating voltage remains constant. When the target ions undergo fragmentation to generate daughter ions, the daughter ions are simultaneously excited by the second alternating voltage in the y direction, exit the linear ion trap, and enter the detector where the daughter ion signals are detected. The detector can be installed at the axial end of the quadrupole for axial ion emission detection; or it can be installed in the radial y direction for radial daughter ion emission detection.

[0046] Drawing of energy-resolved tandem spectra

[0047] According to some specific embodiments of the present application, in this step, the detector is used to obtain the daughter ions to obtain the detection result of the sample to be measured. In the present application, the fragmentation of the target ions, the emission of the daughter ions, and the detection of the daughter ion signals are carried out synchronously. By recording the relationship between the signal intensity of the daughter ions and the amplitude of the corresponding first alternating voltage, an energy-resolved tandem mass spectrum is formed.

[0048] According to some specific embodiments of the present application, the ion trapping method refers to the conventional trapping method of a linear ion trap in the art. Its key parameters are the Mathieu parameters a and q of the parent ions during trapping, which can be calculated by the following formulas 1 and 2. The q value of the parent ions during trapping needs to satisfy less than 0.908.

[0049]

[0050] Where, e is the electron charge (1.6021766208×10-19 C), z is the number of charges carried by the corresponding ion, V_RF is the voltage amplitude of the radio frequency trapping field of the ion trap, U is the DC voltage amplitude of the ion trap (set to 0 V in this example), m is the mass of the corresponding ion, Ω is the frequency of the radio frequency trapping field of the ion trap, and r_0 is the radius of the ion trap.

[0051] According to some specific embodiments of the present application, the frequency of the alternating voltage is calculated by the following formula 3.

[0052]

[0053] Where, Ω is the frequency of the radio frequency trapping field of the ion trap (typical value is about 1 MHz), n is a natural number 0≤n≤∞ (typical value is 0), β is the trapping parameter of the corresponding ion (parent ion or daughter ion), which is related to the mass-to-charge ratio of the corresponding ion, and can be specifically obtained by the continued fraction of formula 4:

[0054]

[0055] Where, both a and q are Mathieu parameters, obtained from formulas 1 and 2.

[0056] According to an embodiment of the present application, the first alternating current voltage is a sinusoidal voltage. Thus, by utilizing the characteristic of a single sine wave frequency, the target ions can smoothly obtain gradually increasing energy during a single injection process, achieving continuous energy scanning and fragmentation from low to high.

[0057] According to an embodiment of the present application, the second alternating current voltage is a sinusoidal voltage. Thus, by utilizing the characteristic of a single sine wave frequency, the daughter ions can be stably and continuously excited in the y direction, thereby ensuring that the daughter ions can efficiently and stably escape from the linear ion trap and be captured by the detector.

[0058] According to an embodiment of the present application, the frequency of the first alternating current voltage is the same as the long-term frequency when the target ions are isolated in the linear ion trap. Thus, the target ions are resonantly excited in the x direction, thereby obtaining energy and achieving continuous energy scanning and fragmentation from low to high.

[0059] According to an embodiment of the present application, the frequency of the second alternating current voltage is the same as the long-term frequency when the daughter ions are isolated in the linear ion trap. Thus, when the daughter ions generated by the fragmentation of the target ions are formed, the alternating current voltage in the y direction excites these daughter ions, causing them to leave the ion trap and enter the detector.

[0060] According to an embodiment of the present application, the x direction and the y direction are orthogonal to each other. In the present application, there are no special restrictions on the x direction and the y direction in the linear ion trap, as long as the x direction and the y direction are orthogonal to each other, enabling the fragmentation of the target ions and the ejection of the daughter ions to be relatively independent and accurate.

[0061] According to an embodiment of the present application, the amplitude of the first alternating current voltage increases with the time of the amplitude scanning process. Thus, the ions are resonantly excited by a gradually increasing energy over time, and the collision energy also increases with time. By using a single ion trap, the fragmentation information of the parent ions in different energy states can be captured in a single injection.

[0062] According to an embodiment of the present application, the amplitude of the first alternating current voltage increases from 0V to 0.5 - 1V. For example, it can increase from 0V to 0.5V, 0.6V, 0.7V, 0.8V, 0.9V, 1V, etc. Thus, sufficient energy is provided to fragment the target ions, and at the same time, the problem of excessive amplitude scanning range reducing the energy resolution accuracy is avoided, ensuring the stable generation and effective detection of the daughter ions.

[0063] According to an embodiment of the present application, the amplitude of the second alternating current voltage remains constant. Thus, when the target ions fragment to generate daughter ions, the daughter ions are simultaneously excited by the second alternating current voltage in the y direction, exit the linear ion trap, and enter the detector to detect the daughter ion signal.

[0064] According to an embodiment of the present application, the amplitude of the second alternating voltage is 0.5 to 1.5 V. For example, it can be 0.5 V, 0.6 V, 0.7 V, 0.8 V, 0.9 V, 1 V, 1.1 V, 1.2 V, 1.3 V, 1.4 V, 1.5 V, etc. Thereby, the daughter ions can obtain sufficient energy to more quickly escape from the linear ion trap and be effectively captured by the detector, while avoiding excessive fragmentation of the daughter ions or reduction of the emission efficiency to the detector due to too high a voltage.

[0065] According to an embodiment of the present application, before performing the amplitude scanning process, collision gas is introduced into the linear ion trap. Thereby, a necessary collision environment is provided for the subsequent fragmentation process of the target ions.

[0066] According to an embodiment of the present application, the time of the amplitude scanning process is 400 to 600 ms. For example, it can be 400 ms, 500 ms, 600 ms, etc. Thereby, this time setting helps to obtain complete energy resolution information in a single injection, while maintaining the high efficiency and accuracy of mass spectrometry analysis.

[0067] In a second aspect of the present application, the present application proposes an apparatus for energy resolution tandem mass spectrometry of a sample based on an ion trap, including: an injection module for ionizing a sample to be measured to obtain ions to be measured; a screening module for allowing the ions to be measured to enter a linear ion trap and screening out target ions; a fragmentation module for applying a first alternating voltage in the x direction of the linear ion trap to cause the target ions to fragment into daughter ions; an emission module for simultaneously applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; and a detection module for allowing a detector to acquire the daughter ions to obtain a detection result of the sample to be measured.

[0068] Those skilled in the art can understand that the features and advantages described above for the method of energy resolution tandem mass spectrometry of a sample based on an ion trap also apply to this apparatus, and will not be elaborated here.

[0069] Hereinafter, the solution of the present application will be explained in conjunction with embodiments. Those skilled in the art will understand that the following embodiments are only used to illustrate the present application and should not be regarded as limiting the scope of the present application. For those not specified in the embodiments regarding specific technologies or conditions, they shall be carried out according to the technologies or conditions described in the literature in the art or according to the product specifications. For reagents or instruments not specified by the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0070] Example 1

[0071] Sample introduction: After ionizing one of the samples 4-tert-butylbenzylamine, 4-chlorobenzylamine, or 4-nitrobenzylamine into the target gas-phase ions by the ion source respectively, they enter the linear ion trap through the discontinuous atmospheric pressure interface; the structures of the three ions are as Figure 3 shown, and the dotted line is the dissociation bond position; the control device of the discontinuous sampling mass spectrometer is as Figure 4 shown.

[0072] Ion trapping: The ions of a single injection are trapped in the linear ion trap, including the target ions and impurity ions.

[0073] Ion isolation: The impurity ions are excited and fly out of the ion trap, leaving only the target ions trapped in the linear ion trap to complete the selective isolation of the target ions.

[0074] Adjusting the air pressure: Open the air replenishing solenoid valve for 15 ms to introduce air into the linear ion trap for subsequent collision-induced dissociation.

[0075] Collision dissociation with amplitude scanning: The linear ion trap contains a pair of electrodes in each of the x and y directions. Apply an alternating voltage ACx to the electrodes in the x direction, and the amplitude of the alternating voltage increases from 0 V to 0.5 V over time. The frequency of the alternating voltage is kept consistent with the long-term frequency of the target ion trapping. The scanning time is 500 ms.

[0076] Product ion detection: While applying the alternating voltage ACx in the x direction, apply another alternating voltage ACy in the y direction, whose voltage amplitude remains constant at 1 V and the frequency is kept consistent with the long-term frequency of the product ions of the target ion. The detector is installed at the axial end of the quadrupole for axial emission detection after the product ions are excited.

[0077] Drawing the energy-resolved tandem mass spectrum: Taking the amplitude of ACx as the horizontal axis x and the signal of the product ions as the vertical axis y, draw the energy-resolved tandem mass spectrum.

[0078] The results are as Figure 5 shown. The signal acquisitions of the above steps are respectively performed on the three selected samples to obtain the energy-resolved tandem mass spectra. Their peak positions represent the dissociation energy magnitudes. The magnitude of the horizontal axis Vacx corresponding to the peak position represents the AC voltage value required for fragmentation. The peak signal with a left-leaning peak position represents a smaller Vacx required for fragmentation, indicating a smaller ion dissociation energy (4-tert-butylbenzylamine ion, the bond dissociation energy is 1.35 eV). The peak signal with a right-leaning peak position represents a larger Vacx required for fragmentation, indicating a larger ion dissociation energy (4-nitrobenzylamine ion, the bond dissociation energy is 1.96 eV). Through this embodiment, it is proved that the linear ion trap can achieve energy-resolved tandem mass spectrometry analysis under single injection, using a linearly scanned AC voltage for dissociation and a constant AC voltage for product ion excitation.

[0079] Comparative Example

[0080] In this comparative example, the difference from the embodiment lies in that in the sub-ion detection, ACy is respectively turned off and ACy is turned on to detect 4-chlorobenzylamine.

[0081] The detector signal results are as Figure 6 shown, where Figure 6 (A) is a waveform schematic diagram of ACx; Figure 6 (B) is the detection signal when ACy is turned off; Figure 6 (C) is the detection signal when ACy is turned on. This proves that the signal of the detector comes from the sub-ion, and the target ion has no influence on the detector signal.

[0082] In the description of this specification, the description with reference to terms such as "one embodiment", "some embodiments", "examples", "specific examples", or "some examples" means that the specific features, structures, materials, or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of this application. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials, or characteristics described can be combined in a suitable manner in any one or more embodiments or examples. In addition, without contradiction, those skilled in the art can combine and combine the different embodiments or examples described in this specification and the features of different embodiments or examples.

[0083] Although the embodiments of this application have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting this application. Those of ordinary skill in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of this application.

Claims

1. A method for energy-resolved tandem mass spectrometry of a sample based on an ion trap, characterized in that, Comprising: Ionizing a sample to be measured to obtain ions to be measured; Introducing the ions to be measured into a linear ion trap to screen out target ions; Performing amplitude scanning on the target ions, the amplitude scanning comprising: Applying a first alternating voltage in the x direction of the linear ion trap to cause the target ions to fragment into daughter ions, and simultaneously applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; Enabling a detector to acquire the daughter ions to obtain a detection result of the sample to be measured.

2. The method according to claim 1, wherein The first alternating voltage is a sinusoidal voltage; Optionally, the second alternating voltage is a sinusoidal voltage.

3. The method according to claim 1, wherein The frequency of the first alternating voltage is the same as the long-term frequency when the target ions are isolated in the linear ion trap; Optionally, the frequency of the second alternating voltage is the same as the long-term frequency when the daughter ions are trapped in the linear ion trap.

4. The method according to claim 1, wherein The x direction and the y direction are orthogonal to each other.

5. The method according to claim 1, characterized in that, The amplitude of the first alternating voltage increases with the time of the amplitude scanning process.

6. The method according to claim 1, characterized in that The amplitude of the first alternating voltage increases from 0 V to 0.5 - 1 V.

7. The method according to claim 1, characterized in that, The amplitude of the second alternating voltage remains constant.

8. The method according to claim 1, characterized in that, The amplitude of the second alternating voltage is 0.5 - 1.5 V.

9. The method according to claim 1, characterized in that, Before performing the amplitude scanning process, introducing a collision gas into the linear ion trap; Optionally, the time of the amplitude scanning process is 400 - 600 ms.

10. An apparatus for energy-resolved tandem mass spectrometry of a sample based on an ion trap, characterized in that, Comprising: An injection module for ionizing a sample to be measured to obtain ions to be measured; A screening module for introducing the ions to be measured into a linear ion trap to screen out target ions; A fragmentation module for applying a first alternating voltage in the x direction of the linear ion trap to cause the target ions to fragment into daughter ions; An ejection module for simultaneously applying a second alternating voltage in the y direction of the linear ion trap to cause the daughter ions to escape from the linear ion trap; A detection module for enabling a detector to acquire the daughter ions to obtain a detection result of the sample to be measured.

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