High resolution ion isolation method and system for an ion trap mass spectrometer

Abstract

The application provides a high-resolution ion isolation method and system of an ion trap mass spectrometer, and relates to the field of mass spectrometry analysis. According to the mass-to-charge ratio of target ions and interfering ions, the frequency of an auxiliary alternating current signal is determined. The amplitude scanning range of the auxiliary alternating current signal is set. The auxiliary alternating current signal with a fixed frequency and a continuously or stepwise changing amplitude in the amplitude scanning range is applied to the ion trap electrode. During the amplitude scanning process, the interfering ions are expelled from the ion trap, and the target ions are retained in the ion trap. The application can realize high-resolution isobaric isolation of 0.1 Th order of magnitude of a mass interval, and significantly improves the isolation resolution of the ion trap mass spectrometer.

Description

Technical Field

[0001] This application relates to the field of mass spectrometry analysis, and in particular to a high-resolution ion isolation method and system for an ion trap mass spectrometer. Background Technology

[0002] Mass spectrometry is a method for separating and detecting substances based on the different motion states of ions with different mass-to-charge ratios (m / z) in an electric or magnetic field. It is characterized by high sensitivity and fast response. Ion trap mass spectrometers, due to their tandem mass spectrometry (MS / MS) capabilities... n Its analytical capabilities are widely used in fields such as food and drug analysis, environmental monitoring, metabolite identification, and biomolecule analysis.

[0003] Ion isolation technology is one of the key techniques in ion trap tandem mass spectrometry. By isolating precursor ions with a specific mass-to-charge ratio in an ion trap, exciting and excluding other ions, the space charge effect can be reduced, the spectral resolution and signal-to-noise ratio can be improved, and pure precursor ions can be provided for subsequent collision-induced dissociation (CID).

[0004] The most widely used ion isolation technique currently is the Stored Waveform Inverse Fourier Transform (SWIFT) technique. SWIFT technology designs a rectangular broadband amplitude spectrum starting from the frequency domain, sets up notch bands within a specific frequency range to retain target ions, and assigns larger amplitudes to frequency components outside the notch bands to excite non-target ions. Then, a time-domain isolation waveform is obtained through inverse Fourier transform and applied to the end cap electrode or radial electrode of the ion trap to achieve selective retention of ions with specific mass-to-charge ratios.

[0005] However, SWIFT technology faces significant challenges when the mass-to-charge ratio difference between the target ion and the interfering ion is extremely small (e.g., less than 1 Th). The secular frequency of an ion in the ion trap is related to its mass-to-charge ratio, but the motion frequencies of ions with adjacent mass-to-charge ratios are very close. In the presence of space charge effects, the ion motion frequency shifts, and the excitation frequency bandwidth broadens, causing overlap in the excitation frequency bands of adjacent ions. Simply reducing the width of the SWIFT notch band or increasing the amplitude of the notch band edge frequency components will lead to mis-excitation of the target ion, resulting in intensity loss.

[0006] To address this, researchers have proposed several improvement schemes. The SWIFTSIN method (see Chinese patent application CN202211120157) achieves fine excitation by superimposing a sinusoidal waveform of a specific frequency onto a SWIFT waveform, achieving isolation resolution at the level of 1 Th mass number. However, this method relies on the selective excitation of the sinusoidal wave in the frequency domain, and it is still difficult to achieve effective separation for ions with a mass difference of less than 0.5 Th.

[0007] The separation of isobaric substances has been a long-standing challenge in mass spectrometry. Ion pairs with a mass-to-charge ratio difference of only 0.11Th (such as isotope peaks of certain drugs and adjacent metabolite peaks) are almost indistinguishable under conventional mass spectrometry conditions, which seriously affects the accurate qualitative and quantitative analysis of complex systems.

[0008] The existing technology has the following problems: The SWIFT method has limited isolation resolution: due to the small secular frequency interval between neighboring mass-to-charge ratio ions in the ion trap, the frequency shift and bandwidth broadening caused by the space charge effect, the traditional SWIFT method can usually only achieve an isolation resolution of 1.0Th, and cannot separate ions with a mass difference of less than 0.5Th.

[0009] Simple methods to improve resolution can lead to the loss of target ions: compressing the notch band width or enhancing the edge frequency components can cause the target ion's motion frequency to shift due to space charge effects or processing errors, resulting in the target ions falling into the excitation band and being lost.

[0010] There is a lack of effective separation methods for isotopic interference: When dealing with isotopic substances with a mass difference of 0.1Th, existing technologies lack effective single-stage isolation and separation methods. High-resolution mass analyzers (such as FT-ICR and Orbitrap) are usually required to distinguish them on the first-stage mass spectrometer, which is costly and still cannot obtain a pure second-stage spectrum. Summary of the Invention

[0011] This application aims to at least partially address one of the technical problems in the related art.

[0012] To address the technical problem of insufficient resolution and inability to separate isotopic substances with a mass interval of 0.1Th in existing ion isolation methods, this application provides a high-resolution ion isolation method for ion trap mass spectrometers based on amplitude scanning AC. By continuously scanning the amplitude of the auxiliary alternating current (AC) signal from low to high, and utilizing the characteristic that ions with different mass-to-charge ratios have different resonance excitation efficiencies under different AC amplitude conditions, interfering ions are expelled while target ions are retained, thereby achieving high-resolution isolation on the order of 0.1Th.

[0013] Another aspect of this application proposes a high-resolution ion isolation system for an ion trap mass spectrometer.

[0014] To achieve the above objectives, the first aspect of this application proposes a high-resolution ion isolation method for an ion trap mass spectrometer, comprising: The frequency of the auxiliary AC signal is determined based on the mass-to-charge ratio of the target ion and the interfering ion. Set the amplitude scanning range of the auxiliary AC signal; An auxiliary AC signal with a fixed frequency and continuously or stepwise varying amplitude within the amplitude scanning range is applied to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

[0015] In one embodiment of this application, determining the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion includes: Based on the mass-to-charge ratio of the target ion and the interfering ion, as well as the operating conditions of the ion trap, the secular frequencies of the target ion and the interfering ion in the ion trap are calculated. The fixed frequency of the auxiliary AC signal is set to the secular frequency of the target ion plus an increment or minus a decrement, so that the fixed frequency is closer to the secular frequency of the interfering ion.

[0016] In one embodiment of this application, the frequency of the auxiliary AC signal is determined based on the mass-to-charge ratio of the target ion and the interfering ion, such that the interfering ion reaches the resonance excitation condition before the target ion during the amplitude scanning process; the operating conditions of the ion trap include the radio frequency, trap structure parameters, type of buffer gas, and pressure.

[0017] In one embodiment of this application, the amplitude scanning range of the auxiliary AC signal is set such that the range covers the resonance excitation threshold of the interfering ion and is lower than the resonance excitation threshold of the target ion; the setting of the amplitude scanning range of the auxiliary AC signal includes: setting the initial amplitude to 0V, and determining the termination amplitude by scanning an emission spectrum, wherein the termination amplitude makes the efficiency of expelling the interfering ion greater than 95% and the target ion not expelled.

[0018] In one embodiment of this application, the setting of the amplitude scanning range of the auxiliary AC signal further includes: setting a scanning rate such that the scanning rate decreases accordingly as the mass-to-charge ratio difference between the target ion and the interfering ion decreases, so as to ensure that each ion obtains sufficient interaction time under the resonance amplitude condition, thereby achieving selective excitation.

[0019] In one embodiment of this application, applying the auxiliary AC signal with a fixed frequency and continuously or stepwise varying amplitude within the scanning range to the ion trap electrode includes: An auxiliary AC signal is coupled to the electrodes of the ion trap via a transformer.

[0020] In one embodiment of this application, the amplitude changing continuously or in steps within the scanning range includes: One of the following methods is employed: linear continuous scanning, exponentially increasing scanning, or segmented constant amplitude step scanning. The duration and amplitude increment of each segment in the segmented constant amplitude step scanning are optimized according to the mass interval between the target ion and interfering ions at different distances.

[0021] In one embodiment of this application, the auxiliary AC signal contains two frequency components, which correspond to different interfering ions distributed on both sides of the target ion. The amplitude scanning range and scanning rate of each frequency component are independently set according to the mass-to-charge ratio of their respective interfering ions.

[0022] In one embodiment of this application, before applying the auxiliary AC signal, the method further includes: first performing wide notch band coarse isolation using the stored waveform inverse Fourier transform waveform to exclude interfering ions far from the target ions, wherein the width of the wide notch band is 1 to 5Th. The auxiliary AC signal is applied to the end cap electrode of the three-dimensional ion trap to achieve axial excitation, or simultaneously applied to electrodes in multiple directions of the linear ion trap to enhance excitation efficiency.

[0023] Furthermore, to achieve the above embodiments, this application proposes a high-resolution ion isolation system for an ion trap mass spectrometer, comprising: The frequency determination module is used to determine the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion; The scanning range setting module is used to set the amplitude scanning range of the auxiliary AC signal; The ion separation module is used to apply an auxiliary AC signal with a fixed frequency and a continuously or stepwise varying amplitude within the amplitude scanning range to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

[0024] The methods and systems of this application can achieve high-resolution isolation of isotopic materials with a mass interval of only 0.1Th, breaking through the resolution limit of traditional methods in the frequency domain and significantly improving the isolation accuracy and complex sample analysis capabilities of ion trap mass spectrometers.

[0025] 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

[0026] 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: Figure 1 A flowchart illustrating a high-resolution ion isolation method for an ion trap mass spectrometer provided in this application embodiment; Figure 2 This is a schematic diagram illustrating the principle of this application; Figure 3 This is a graph showing the experimental data of the isolation effect of this application; Figure 4This is a result diagram of Embodiment 2 of this application; Figure 5 This is a result diagram of Embodiment 3 of this application; Figure 6 This is a structural diagram of a high-resolution ion isolation system for an ion trap mass spectrometer provided in an embodiment of this application. Detailed Implementation

[0027] It should be noted that, unless otherwise specified, the embodiments and features described in this application can be combined with each other. This application will now be described in detail with reference to the accompanying drawings and embodiments.

[0028] To enable those skilled in the art to better understand the present application, the technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present application, and not all embodiments. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative effort should fall within the scope of protection of the present application.

[0029] The following description, with reference to the accompanying drawings, describes a high-resolution ion isolation method and system for an ion trap mass spectrometer according to embodiments of this application.

[0030] Example 1 like Figure 1 As shown, this high-resolution ion isolation method for an ion trap mass spectrometer specifically includes the following steps: S1. Determine the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion.

[0031] Specifically, in the high-resolution ion isolation method of ion trap mass spectrometry, the frequency parameters of the auxiliary AC signal applied to the ion trap electrodes must first be determined. This frequency setting is based on the motion characteristics of the target ion and interfering ions within the ion trap. Specifically, there is a definite correspondence between the secular frequency of an ion and its mass-to-charge ratio in a quadrupole field. This relationship is determined by the ion trap's RF operating parameters, trap structure parameters, and buffer gas conditions. By calculating or experimentally determining the secular frequencies of the target ion and interfering ions, the frequency of the auxiliary AC signal is configured to be closer to the secular frequency of the interfering ion, while maintaining a certain offset from the secular frequency of the target ion. This ensures that, during subsequent amplitude scanning, the interfering ion can reach the resonance excitation condition before the target ion. This frequency selection strategy utilizes the characteristic that ions with different mass-to-charge ratios have different resonance excitation efficiencies under the same frequency of AC electric field. That is, the closer the AC signal frequency is to the secular frequency of a certain ion, the more resonance energy that ion can obtain at a lower amplitude and be expelled from the ion trap. Therefore, by setting the frequency of the auxiliary AC signal appropriately, it can be ensured that interfering ions are preferentially excited and eliminated during the scanning process from low to high amplitude, while target ions require a higher amplitude to be excited due to the large difference between their duration frequency and the AC signal frequency, thus being retained throughout the scanning process.

[0032] As one implementation method, the frequency of the auxiliary AC signal can be set to the target ion's duration frequency plus or minus a preset frequency increment or decrement, based on the target ion's duration frequency under a given radio frequency voltage condition. The magnitude of this increment or decrement is optimized based on the mass-to-charge ratio difference between the target ion and the interfering ion, as well as the actual operating conditions of the ion trap. For example, it can be set to an offset of approximately 0.5 kHz to make the duration frequency of the interfering ion closer to the AC signal frequency.

[0033] This step establishes the sequential relationship of ion expulsion during amplitude scanning by precisely selecting the frequency of the auxiliary AC signal, laying the foundation for subsequent high-resolution isolation. Its advantage lies in the fact that it overcomes the limitation of frequency resolution on isolation capability in traditional methods by utilizing the difference in resonance excitation thresholds of different ions during amplitude scanning, without relying on narrowband filtering or high-precision frequency selection in the frequency domain. This enables effective separation of isotopic ions with extremely small mass-to-charge ratio differences (e.g., on the order of 0.1Th), significantly improving the isolation resolution of the ion trap mass spectrometer.

[0034] S2 sets the amplitude scanning range of the auxiliary AC signal.

[0035] Specifically, when setting the amplitude scanning range of the auxiliary AC signal, it is necessary to determine an amplitude range that can selectively expel interfering ions while retaining target ions, based on the difference in amplitude required for resonant excitation of target ions and interfering ions in the ion trap. The lower limit of this amplitude scanning range should be lower than or equal to the initial amplitude at which interfering ions begin to undergo significant resonant excitation, ensuring that interfering ions can be effectively excited and expelled during the scanning process; the upper limit of this amplitude scanning range should be lower than the critical amplitude at which the target ions reach the resonant excitation threshold and begin to be expelled, thereby avoiding loss of target ions during the scanning process. The specific value of the amplitude scanning range can be obtained through experimental determination or calculation based on ion motion theory. For example, by performing amplitude scanning emission spectrum analysis on a sample containing target ions and interfering ions in advance, the amplitude at which interfering ions are completely expelled and the amplitude at which target ions begin to be lost can be determined based on the emission peak positions corresponding to each ion, and then the scanning termination amplitude can be set between the two. The scanning start amplitude can be set to a low value below the excitation threshold of interfering ions, including an initial state of zero amplitude or near-zero amplitude, to ensure that the scanning process starts from a state where interfering ions have not yet been excited. The scanning method can be a continuous linear change, a piecewise constant amplitude step change, or a non-linear incremental method, as long as the amplitude increases monotonically with time and covers the range.

[0036] As one implementation method, the initial amplitude can be set to 0V, and the termination amplitude can be determined by scanning the emission spectrum so that the range just covers the resonance excitation threshold of the interfering ion but is lower than the resonance excitation threshold of the target ion. The scanning rate can be optimized according to the excitation amplitude window interval between the target ion and the interfering ion, for example, using a linear scanning rate of 1.2V / s.

[0037] By precisely setting the amplitude scanning range, the amplitude change of the auxiliary AC signal only covers the resonant excitation region of interfering ions and avoids the excitation region of target ions. This achieves selective expulsion of interfering ions and complete retention of target ions during the amplitude scanning process, significantly improving the selectivity and resolution of ion isolation. It is especially suitable for the separation of isotopic substances with extremely small mass-to-charge ratio differences (such as on the order of 0.1Th). At the same time, it avoids the loss of target ions due to excessive amplitude, ensuring the reliability of the isolation operation and the ion strength.

[0038] S3, apply an auxiliary AC signal with a fixed frequency and an amplitude that changes continuously or stepwise within the amplitude scanning range to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

[0039] Specifically, after ion introduction and cooling in the ion trap, an auxiliary AC signal with a fixed frequency and continuously or incrementally varying amplitude within a preset scanning range is applied to at least one pair of electrodes in the ion trap during the mass isolation stage. The frequency of this auxiliary AC signal is predetermined based on the mass-to-charge ratio of the target ion and interfering ions, ensuring that the interfering ions reach the resonant excitation emission condition before the target ion during the amplitude scanning process. The amplitude scanning range is set to cover the resonant excitation threshold of the interfering ions but is lower than the resonant excitation threshold of the target ion. When this signal is applied, the auxiliary AC signal is coupled to the ion trap electrodes, establishing an alternating electric field coupled to the ion motion within the ion trap. As the amplitude gradually increases from the initial value to the final value, ions with different mass-to-charge ratios within the trap sequentially experience the resonant excitation condition: ions with a mass-to-charge ratio corresponding to the set frequency of the auxiliary AC signal first gain sufficient energy and are expelled from the trap; subsequently, ions with adjacent mass-to-charge ratios reach their respective excitation thresholds and are expelled as the amplitude continues to increase. By precisely controlling the rate and range of the amplitude scanning, the target ion never reaches its resonant excitation threshold throughout the entire scanning process, thus remaining within the trap. In one implementation, the amplitude of the auxiliary AC signal can be linearly and continuously scanned, starting from a zero-volt or near-zero-volt initial amplitude and linearly increasing at a constant rate to the final amplitude. Alternatively, a segmented constant amplitude step-scanning method can be used, maintaining a constant amplitude for a certain time within different amplitude segments before stepping to the next amplitude segment. The duration and amplitude increment of each segment can be optimized according to the mass interval between the target ion and interfering ions at different distances. Furthermore, the auxiliary AC signal can be applied to the end cap electrodes of a three-dimensional ion trap to achieve axial excitation, or to the radial electrode pair of a linear ion trap, or simultaneously applied in multiple directions. This step, through scanning operations in the amplitude dimension, utilizes the characteristic that different ions have different resonant excitation amplitude thresholds at the same frequency due to differences in mass-to-charge ratio, achieving selective ion expulsion and retention.

[0040] This step replaces the traditional fixed amplitude excitation method with an auxiliary AC signal that scans the amplitude, breaking through the physical limitations of frequency domain resolution. This allows for selective isolation of adjacent ions with extremely small mass gaps (such as on the order of 0.1Th) on conventional ion traps, while avoiding the loss of target ions due to mis-excitation caused by frequency shift, thus significantly improving the resolution and selectivity of ion isolation.

[0041] Furthermore, during the amplitude scanning process, an auxiliary AC signal with a fixed frequency and continuously or incrementally varying amplitude within a preset range is applied to the ion trap electrode. This utilizes the characteristic that ions with different mass-to-charge ratios have different resonant excitation efficiencies under the same fixed-frequency auxiliary AC signal, achieving selective separation of interfering ions and target ions. Specifically, as the amplitude of the auxiliary AC signal gradually increases from a lower value, interfering ions, whose resonant frequency is closer to that of the auxiliary AC signal, will reach the resonant excitation condition first, thus gaining sufficient energy to be expelled from the ion trap. However, because the target ion's resonant frequency deviates from the auxiliary AC signal frequency, its resonant excitation efficiency is lower under the same amplitude conditions. Therefore, before the amplitude of the auxiliary AC signal reaches its resonant excitation threshold, the target ion cannot obtain enough energy to be expelled and remains within the ion trap. By precisely controlling the start and end values ​​of the amplitude scan and the scan rate, interfering ions are completely expelled throughout the scanning process, while the target ion remains in a stable trapped state, ultimately retaining only the target ion within the ion trap. As one implementation method, during the amplitude scanning process, a wider amplitude scanning range can be used to perform full-spectrum analysis to determine the resonance excitation threshold of interfering ions. Then, based on this threshold, the termination value of subsequent fine isolation amplitude scanning can be set, thereby ensuring that interfering ions are completely eliminated while avoiding the loss of target ions. In addition, for multiple interfering ions distributed on both sides of the target ion, auxiliary AC signals of different frequencies can be applied sequentially to perform multiple rounds of amplitude scanning, or auxiliary AC signals containing multiple frequency components can be applied simultaneously, with each frequency component corresponding to a different interfering ion, so as to achieve the sequential or synchronous expulsion of multiple interfering ions.

[0042] This technique overcomes the resolution limitations of traditional ion isolation methods in the frequency domain by scanning in the amplitude dimension rather than selecting in the frequency dimension. It enables the effective separation of interfering ions with extremely small mass-to-charge ratio differences (such as on the order of 0.1Th) from target ions on conventional ion traps, while avoiding the loss of target ions due to false excitation caused by frequency shift, thus significantly improving the selectivity and fidelity of ion isolation.

[0043] Example 2 Figure 2 This is a schematic diagram of this application; see [link / reference]. Figure 2 (a) Figure 2 (b) and Figure 2 (c) The core principle of this application is as follows: A fixed-frequency auxiliary alternating current (AC) resonance excitation signal is applied to the electrodes of an ion trap mass analyzer, and the amplitude of this AC signal is gradually increased starting from a low value. During resonance excitation, ions with ion motion frequencies closer to fac have a larger resonance amplitude ( Figure 2 Ions whose ion motion frequency is far from that of the fac have a smaller resonance amplitude. Figure 2 During the scanning process from low to high AC amplitude, impurity ions with frequencies close to fac are removed first, leaving target ions far from fac. By rationally designing the scanning frequency fac, initial amplitude, and final amplitude, successive excitation and ejection of ions with different mass-to-charge ratios can be achieved, realizing high-resolution separation. Specifically, this technical solution includes the following steps: Step S101: Determine the frequency parameters of the AC signal: Target ions (ions to be retained) and interfering ions (ions to be excluded) are trapped in an ion trap. Based on the mass-to-charge ratio of the target and interfering ions, and the operating conditions of the ion trap (RF frequency Ω, trap structure parameters, type and pressure of buffer gas, etc.), the secular frequency of each ion in the trap is calculated. The fixed frequency of the AC signal is set to the secular frequency of the target ion plus an appropriate increment or decrement, such as ±0.5 kHz. The relationship between the secular frequency ω of the ion in the quadrupole field and the mass-to-charge ratio m is:

[0044] in Let ω be the angular frequency of the radio frequency signal, and β be a dimensionless parameter related to the Mathieu equation's steady-state parameter q.

[0045] Where e is the charge, V is the amplitude of the radio frequency voltage, and r0 is the field radius of the ion trap. From the above relationship, it can be seen that ions with different mass-to-charge ratios have different secular frequencies, and the motion frequency of a specific ion under a given q value can be changed by adjusting the radio frequency voltage V.

[0046] Step S102: Set the amplitude scanning parameters of the AC signal: The amplitude of the AC signal is scanned from the starting amplitude V_start to the ending amplitude V_end.

[0047] The principle for setting the initial amplitude V_start is that V_start is generally set to 0V, that is, the amplitude is gradually increased from 0V during the scan.

[0048] The principle for setting the termination amplitude V_end: Setting V_end too high will cause the target ion to be ejected by resonant excitation, making it impossible to retain the target ion. Setting V_end too low will result in incomplete removal of impurity ions. The specific V_end value is determined by scanning an emission spectrum. For example, in... Figure 3 In the middle, an output spectrum is scanned by AC amplitude ( Figure 3 (a) in the formula determines the V_end threshold for each substance peak.

[0049] Scan rate setting principle: The scan rate should match the excitation amplitude window interval of the target ion and interfering ions. The slower the scan rate, the longer the interaction time of each ion under its respective resonance amplitude condition, resulting in higher excitation efficiency and better selectivity. In this embodiment, a scan rate of 1.2 V / s is used for high-resolution isolation.

[0050] Step S103: Apply an amplitude scanning AC signal to perform isolation operation: After ion introduction and cooling are completed in the ion trap, an AC signal with a fixed frequency and linear amplitude is applied to the electrodes of the ion trap (such as ring electrodes or rod electrodes) during the mass isolation phase. The AC signal is coupled to the ion trap electrodes via a transformer.

[0051] During the AC amplitude scan from low to high, ions with different mass-to-charge ratios successively experience the resonance condition. Ions with a mass-to-charge ratio corresponding to the AC set frequency first gain sufficient energy and are expelled from the trap; then, ions with adjacent mass-to-charge ratios successively reach the excitation threshold and are expelled as the amplitude continues to increase. Figure 3 (b) By precisely controlling the scan rate and amplitude range, the target ion is kept within the trap without ever reaching its excitation threshold throughout the scan. This ultimately achieves a high-resolution isolation effect where the target ion is retained while interfering ions with a mass gap of only 0.1Th are expelled.

[0052] It is understood that in step S102 above, the amplitude scanning method is not limited to linear scanning, but can also employ exponentially increasing scanning, segmented constant amplitude step scanning, etc. The duration and amplitude increment of each segment of the segmented step scanning can be optimized according to the mass interval between the target ion and interfering ions at different distances.

[0053] It is understood that in step S101 above, the AC signal is not limited to a single frequency and can simultaneously contain multiple frequency components to simultaneously exclude different interfering ions distributed on both sides of the target ion. The amplitude scanning range and rate corresponding to each frequency component can be independently set according to the mass-to-charge ratio of their respective interfering ions.

[0054] It is understood that in step S103, the AC signal can be applied to the end cap electrode of the three-dimensional ion trap (axial excitation) to replace the application method of the x / y electrode pair of the linear ion trap, or applied in multiple directions at the same time to enhance the excitation efficiency.

[0055] Example 3 This embodiment demonstrates the isolation effect of this application on the multicharge isotope peak of bradykinin with a mass difference of 0.5Th.

[0056] This embodiment was performed using an ion trap mass spectrometer. The sample used was a solution of bradykinin (average molecular weight approximately 1060 Da), and multiply charged ions were generated using an electrospray ionization (ESI) source. The [M+2H]² fraction of bradykinin was selected for the experiment. + The ion being studied has a theoretical monoisotope mass-to-charge ratio of approximately 530.8 Th, with a mass interval of 0.5 Th (i.e., 1 Da / 2 charge) between adjacent isotope peaks. In this embodiment, the target ion is bradykinin [M+2H]². + The ion trap shows an A1 isotope peak (m / z 531.3), with interfering ions being the adjacent A0 isotope peak (m / z 530.8) and the adjacent A2 isotope peak (m / z 531.8). The mass-to-charge ratio difference between the three is 0.5 Th. The ion trap radio frequency is Ω / 2π = 1.030 MHz, and the buffer gas is air at a pressure of approximately 1 × 10⁻⁶. -5 Torr.

[0057] First, the secular frequency of the target ion (m / z 531.3) is calculated based on its secular frequency under a given RF voltage condition. Two AC signals of different frequencies are used to eliminate interfering ions distributed on both sides of the target ion. AC1 is fixed at 150.92 kHz, and AC2 is fixed at 150.45 kHz. The AC1 amplitude scan uses a linear scan method, starting from V_start = 0V and linearly increasing to V_end = 1.2V within 1 second. The initial scan range is large to ensure that the interfering ion m / z 531.8 is completely expelled from the ion trap and detected. Figure 4 (a) Based on the full spectrum analysis above, it can be determined that in order to retain the target ion m / z 531.3 and remove the impurity ion m / z 530.8, the V_end of AC1 only needs to be scanned to 0.24V. The V_end confirmation method for AC2 is the same; in order to retain the target ion m / z 531.3 and remove the impurity ion m / z 531.8, the V_end of AC2 only needs to be scanned to 0.24V. First, AC1 is used to remove impurity ions with m / z less than the target ion, and then AC2 is used to remove impurity ions with m / z greater than the target ion. Figure 4 (b) in the middle, finally leaving the target ion ( Figure 4 c). The mass spectra before and after the removal demonstrate that this method can achieve high-resolution ion isolation with a 0.5Th gap (c). Figure 4 (d in the text)

[0058] Example 4 This embodiment demonstrates the isolation effect of this application on a mixture of fendiline (m / z 316.0926) and chlorprothixene (m / z 316.2065) with a mass difference of 0.11Th.

[0059] This embodiment was performed using a linear ion trap mass spectrometer. The sample used in the experiment was fendiline ([M+H)). + The theoretical mass-to-charge ratio (m / z 316.2065) and chlorprothixene ([M+H)) + An equimolar concentration mixed solution with a theoretical mass-to-charge ratio (m / z 316.0926). The mass-to-charge ratio difference is Δ = 0.1139 Th (approximately 0.11 Th), which is a typical isobaric ion pair. Protonated molecular ions [M+H] were generated using an electrospray ionization (ESI) source. + The ion trap radio frequency is Ω / 2π = 1.0 MHz, the buffer gas is air, and the pressure is approximately 1 × 10⁻⁶. -5 Torr.

[0060] In this embodiment, the target ion is fentiline (m / z 316.2065), and the interfering ion is chlorprothiaton (m / z 316.0926), with a mass difference of only about 0.11 Th. Due to their extremely close mass-to-charge ratios, traditional SWIFT or SWIFTSIN methods cannot effectively separate them while maintaining the target ion strength.

[0061] The secular frequency of the target fenidrine ion at q=0.4 was calculated under the given ion trap operating conditions. The AC signal frequency was fixed at 150.87 kHz. The AC amplitude scan started at V_start = 0.00 V and linearly increased to V_end = 0.25 V, with a total scan time of approximately 1 s. The AC signal frequency is greater than the secular frequency of the target fenidrine ion and closer to the secular frequency of the impurity chlorothiaton ion; therefore, the impurity chlorothiaton ion will be removed first during the amplitude scan.

[0062] Before amplitude scanning and AC isolation, a full-spectrum MS1 analysis is performed. Figure 5 (a) is used in the figure, followed by coarse isolation using a wide-band SWIFT waveform (5(b)) to shorten the scan time of the fine isolation stage and eliminate interference from distant ions. After coarse isolation, only the target ion and a small number of ions with a mass-to-charge ratio in the ±0.5 Th range are retained in the ion trap. Figure 5 (c) in the middle), then the first amplitude scan AC is applied to perform a full spectrum scan ( Figure 5In (d) of the results, a double peak appears, indicating a mixture of target and impurity ions. It can also be determined that chlorothiaton ions can be completely removed when scanning to 0.12V; therefore, V_end = 0.12V is determined to be used for the removal of chlorothiaton ions.

[0063] To verify the selectivity of isolation, CID secondary mass spectra were also collected before and after isolation. The CID spectrum before isolation contained characteristic fragment peaks of both fendiline and chlorprothiazide, resulting in a more complex spectrum. Figure 5 (e)); while in the CID spectrum after amplitude scanning and AC isolation, only the characteristic fragment peak of findilin (m / z 212) appears ( Figure 5 (f) further confirms that the isolation method effectively separates two isotopic substances with a mass difference of 0.11Th.

[0064] In summary, compared with existing technologies, this solution has the following technical advantages: 1. Achieve high-resolution ion isolation at the 0.1Th level: Through amplitude scanning AC technology, it is possible to separate isotopic substances with a mass interval of only 0.11Th, which is a resolution level that is difficult to achieve by existing methods such as SWIFT and SWIFTSIN. Figure 4 The experimental results clearly demonstrate the effective separation of 0.11Th mass difference ions.

[0065] 2. Breaking the physical limit of sine wave frequency selectivity: The amplitude scanning AC method does not rely on the frequency selectivity of a single sine wave at a fixed amplitude. Instead, it excites different ions by scanning in the amplitude dimension, thereby breaking the physical limitation of frequency domain resolution and making it possible to achieve ultra-high resolution isolation on conventional ion traps.

[0066] 3. No need to change the instrument hardware structure: This method only requires adding the AC amplitude scanning control function to the instrument's measurement and control software, without changing the hardware structure of the ion trap. It is applicable to various commercially available ion trap mass spectrometers and has broad compatibility and feasibility.

[0067] 4. Enhanced Qualitative and Quantitative Capabilities in Mass Spectrometry: High-resolution isolation capabilities enable ion trap mass spectrometers to distinguish isotopic interferences that were previously indistinguishable. This significantly reduces false positives and quantitative biases in the analysis of complex matrix samples, and is of great significance for applications such as drug metabolite identification, endogenous substance analysis, and environmental pollutant detection. In secondary mass spectrometry (MS / MS) experiments, selective isolation of precursor ions with a mass difference of 0.1Th allows fragment ions to be clearly associated with specific precursor ions, aiding in the accurate derivation of fragmentation pathways.

[0068] Example 5 In yet another embodiment, such as Figure 6As shown, this application provides a high-resolution ion isolation system 10 for an ion trap mass spectrometer, comprising: The frequency determination module 100 is used to determine the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion. The scanning range setting module 200 is used to set the amplitude scanning range of the auxiliary AC signal; The ion separation module 300 is used to apply an auxiliary AC signal with a fixed frequency and a continuously or stepwise variable amplitude within the amplitude scanning range to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

[0069] This application discloses a high-resolution ion isolation system for an ion trap mass spectrometer. By continuously scanning the amplitude of the auxiliary alternating current (AC) signal from low to high, and utilizing the characteristic that ions with different mass-to-charge ratios have different resonance excitation efficiencies under different AC amplitude conditions, interfering ions are expelled while target ions are retained, thereby achieving high-resolution isolation on the order of 0.1Th.

[0070] 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.

[0071] 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, "multiple" means at least two, such as two, three, etc., unless otherwise explicitly specified.

Claims

1. A high-resolution ion isolation method for an ion trap mass spectrometer, characterized in that, include: The frequency of the auxiliary AC signal is determined based on the mass-to-charge ratio of the target ion and the interfering ion. Set the amplitude scanning range of the auxiliary AC signal; An auxiliary AC signal with a fixed frequency and continuously or stepwise varying amplitude within the amplitude scanning range is applied to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

2. The method as described in claim 1, characterized in that, The determination of the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion includes: Based on the mass-to-charge ratio of the target ion and the interfering ion, as well as the operating conditions of the ion trap, the secular frequencies of the target ion and the interfering ion in the ion trap are calculated. The fixed frequency of the auxiliary AC signal is set to the secular frequency of the target ion plus an increment or minus a decrement, so that the fixed frequency is closer to the secular frequency of the interfering ion.

3. The method as described in claim 2, characterized in that, Based on the mass-to-charge ratio of the target ion and the interfering ion, the frequency of the auxiliary AC signal is determined so that the interfering ion reaches the resonant excitation emission condition before the target ion during the amplitude scanning process; the operating conditions of the ion trap include the radio frequency, trap structure parameters, type of buffer gas, and pressure.

4. The method as described in claim 1, characterized in that, Set the amplitude scanning range of the auxiliary AC signal so that the range covers the resonance excitation threshold of the interfering ion and is lower than the resonance excitation threshold of the target ion; The amplitude scanning range of the auxiliary AC signal is set as follows: the initial amplitude is set to 0V, and the termination amplitude is determined by scanning an emission spectrum. The termination amplitude ensures that the efficiency of expelling interfering ions is greater than 95% and that the target ion is not expelled.

5. The method as described in claim 1, characterized in that, The amplitude scanning range of the set auxiliary AC signal also includes: The scanning rate is set to decrease accordingly as the mass-to-charge ratio difference between the target ion and the interfering ion decreases, so as to ensure that each ion obtains sufficient interaction time under the resonance amplitude condition, thereby achieving selective excitation.

6. The method as described in claim 1, characterized in that, The auxiliary AC signal applied to the ion trap electrode, which has a fixed frequency and whose amplitude varies continuously or in steps within the scanning range, includes: An auxiliary AC signal is coupled to the electrodes of the ion trap via a transformer.

7. The method as described in claim 1, characterized in that, The amplitude changing continuously or in steps within the scanning range includes: One of the following methods is employed: linear continuous scanning, exponentially increasing scanning, or segmented constant amplitude step scanning. The duration and amplitude increment of each segment in the segmented constant amplitude step scanning are optimized according to the mass interval between the target ion and interfering ions at different distances.

8. The method as described in claim 1, characterized in that, The auxiliary AC signal contains two frequency components, which correspond to different interfering ions distributed on both sides of the target ion. The amplitude scanning range and scanning rate of each frequency component are set independently according to the mass-to-charge ratio of their respective interfering ions.

9. The method as described in claim 1, characterized in that, Before applying the auxiliary AC signal, the method further includes: first using the stored waveform inverse Fourier transform waveform to perform wide notch band coarse isolation to eliminate interfering ions far from the target ion, wherein the width of the wide notch band is 1 to 5Th. The auxiliary AC signal is applied to the end cap electrode of the three-dimensional ion trap to achieve axial excitation, or simultaneously applied to electrodes in multiple directions of the linear ion trap to enhance excitation efficiency.

10. A high-resolution ion isolation system for an ion trap mass spectrometer, characterized in that, include: The frequency determination module is used to determine the frequency of the auxiliary AC signal based on the mass-to-charge ratio of the target ion and the interfering ion; The scanning range setting module is used to set the amplitude scanning range of the auxiliary AC signal; The ion separation module is used to apply an auxiliary AC signal with a fixed frequency and a continuously or stepwise varying amplitude within the amplitude scanning range to the ion trap electrode. During the amplitude scanning process, interfering ions are expelled from the ion trap, while the target ions are retained within the ion trap.

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