Rapid mass spectrometric detection method and device for multi-component compounds in body fluid
By introducing ion migration pathways and segmented electric fields into mass spectrometry detection, combined with stable isotope internal standards and eluent membrane technology, the problems of sequential ion release and quantitative accuracy of compounds were solved, enabling rapid and accurate detection of multi-component compounds.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- ZHONGYAN TESTING CO LTD
- Filing Date
- 2026-03-16
- Publication Date
- 2026-05-12
Smart Images
Figure CN122016990A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of mass spectrometry detection technology, and in particular to a rapid mass spectrometry detection method and apparatus for multi-component compounds in body fluids. Background Technology
[0002] In the field of mass spectrometry analysis, the detection technology for multi-component compounds in body fluids has evolved from traditional separation coupling to environmental ionization and high-resolution instruments. The emergence of soft ionization techniques such as electrospray ionization and matrix-assisted laser desorption / ionization has enabled the gentle ionization of macromolecules and polar compounds, driving breakthroughs in biological sample analysis. Subsequently, liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS) became mainstream for the separation and qualitative and quantitative analysis of drug metabolites, narcotic residues, and endogenous metabolites in body fluids (such as blood, urine, and saliva). The development of high-resolution mass spectrometers has further improved detection accuracy and multiplexing capabilities, supporting the simultaneous monitoring of hundreds of compounds and significantly advancing metabolomics and toxicology research.
[0003] However, existing mass spectrometry detection techniques still have room for improvement. First, they lack an inherent ion sequential release mechanism, making it impossible to naturally distinguish ions of compounds with different physicochemical properties over time. This causes the resolution of isomers or compounds of similar mass to rely on external chromatography or additional post-processing, further increasing operational complexity and instrument dependence. Second, although the integration of stable isotope internal standards in existing methods can correct some matrix effects, the physicochemical environment of the internal standard and the target compound is often not completely consistent, affecting the reliability of the quasi-quantitative results. Summary of the Invention
[0004] In view of the aforementioned existing problems, the present invention is proposed.
[0005] Therefore, this invention provides a rapid mass spectrometry detection method for multi-component compounds in body fluids, which solves the problems of lacking an intrinsic ion release mechanism and the fact that the physicochemical environment of the internal standard and the target compound is often not completely consistent.
[0006] To solve the above-mentioned technical problems, the present invention provides the following technical solution: In a first aspect, the present invention provides a rapid mass spectrometry detection method for multi-component compounds in body fluids, comprising, Body fluid samples were collected from the subjects and transferred to the functionalized treatment interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized treatment interface adjacent to the ion source inlet. The sample surface area is positioned inside the ion source. The sampling needle releases the desorption solvent into the sample surface area from inside the ion source, and the tip of the sampling needle contacts the sample surface area. The desorption solvent forms a desorption liquid film on the sample surface area that carries the target compound and stable isotope internal standard. The eluent film is retained inside the ion source. Under the conditions of electric field, thermal assistance and airflow assistance of the ion source, the solvent in the eluent film is gradually evaporated, the target compound in the eluent film is ionized to form target compound ions, and the target compound ions are released in the order of physical migration and energy response differences and guided to the mass analysis region to form an ion signal time series. Mass spectrometry was used to detect the time series of ion signals. Characteristic ion signals of target compound ions and stable isotope internal standard ions were collected through multiple reaction monitoring mode. Qualitative identification was completed, and the concentration range of the target compound was determined based on the signal intensity ratio, generating structured detection results.
[0007] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, the body fluid sample of the subject is obtained by directly dipping a sampling needle into the body fluid on the surface of the subject.
[0008] In a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, the formation of the sample surface region specifically refers to: The body fluid sample collected by the sampling needle is introduced into the functionalized processing interface of the pre-set stable isotope internal standard, and the body fluid sample droplets are formed on the functionalized processing interface and defined as the sample surface area. Exposing the body fluid sample in the sample surface area to the acidic chemical environment of the functionalized treatment interface causes the macromolecular components in the body fluid sample to undergo conformational changes and aggregate and become inactive. By bringing the body fluid sample in the sample surface area into contact with the functional material of the functionalized treatment interface, the non-target matrix components in the body fluid sample are retained at the functionalized treatment interface and the interference is weakened. The target compound and stable isotope internal standard in the sample surface region are enriched at the functionalization interface and maintained in the sample surface region.
[0009] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, wherein: the formation of the eluent membrane carrying the target compound and a stable isotope internal standard specifically comprises: The desorption solvent is introduced into the sample surface area from inside the ion source by a sampling needle, and the tip of the sampling needle touches the sample surface area to form a local solvent coating layer in the sample surface area. The target compound and stable isotope internal standard in the sample surface area are desorbed by a local solvent coating layer, and the liquid film morphology of the local solvent coating layer is maintained inside the ion source to form an eluent film carrying the target compound and stable isotope internal standard.
[0010] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, wherein: the formation of target compound ions specifically includes: The eluent film carrying the target compound and stable isotope internal standard is kept in the eluent position inside the ion source, and an electric field is established inside the ion source to act on the eluent film. Heat assistance and airflow assistance are applied to the desorption location inside the ion source to drive the desorption solvent in the desorption liquid film to gradually evaporate and maintain the desorption liquid film in a volatile state; During the gradual evaporation of the eluent membrane, the electric field inside the ion source is maintained, causing the target compound in the eluent membrane to ionize and form target compound ions.
[0011] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, wherein: the formation of the ion signal time series specifically includes: An ion migration path is set up inside the ion source and a segmented electric field is applied to the ion migration path to enable the target compound ions to migrate in a controlled manner along the ion migration path. Simultaneous application of thermal and gas flow assistance along the ion migration path alters the time distribution of target compound ions released from the eluent film into the gas phase, causing target compound ions with different physicochemical properties to be released sequentially according to differences in physical migration and energy response. By setting a guiding electrode at the end of the ion migration path and applying a guiding electric field, the target compound ions released sequentially are guided to the mass analysis region, forming an ion signal time series in the time dimension.
[0012] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, wherein: the acquisition of characteristic ion signals of target compound ions and stable isotope internal standard ions through multiple reaction monitoring mode specifically includes: During mass spectrometry detection, a multi-reaction monitoring mode is set and the parent ion-daughter ion conversion relationship corresponding to the target compound ion is preset; In multi-reaction monitoring mode, the parent ion signal and daughter ion signal of the target compound are acquired simultaneously and the signal intensity is recorded. In the multi-reaction monitoring mode, the parent ion signal and daughter ion signal of stable isotope internal standard ions are acquired simultaneously and the signal intensity is recorded.
[0013] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, wherein: determining the concentration range of the target compound specifically involves: The characteristic ion signal intensity of the target compound ion is extracted from the parent ion signal and daughter ion signal of the target compound ion; The characteristic ion signal intensity of stable isotope internal standard ions is extracted from the parent ion signal and daughter ion signal of stable isotope internal standard ions. Calculate the ratio of the characteristic ion signal intensity of the target compound ion to the characteristic ion signal intensity of the stable isotope internal standard ion, and determine the concentration range of the target compound based on the preset concentration range correspondence.
[0014] As a preferred embodiment of the rapid mass spectrometry detection method for multi-component compounds in body fluids according to the present invention, the generation of structured detection results specifically includes: Extract the time series signals corresponding to the target compound ions from the ion signal time series and establish the target compound ion time series record; The target compound ion time series records are correlated with the target compound concentration range to form a target compound detection record; The detection records of the target compound are uniformly packaged and structured detection results are generated.
[0015] Secondly, the present invention provides a rapid mass spectrometry detection device for multi-component compounds in body fluids, comprising, The acquisition module collects body fluid samples from the subject and transfers the body fluid samples to the functionalized processing interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized processing interface adjacent to the ion source inlet. The analysis module positions the sample surface area inside the ion source, releases the analysis solvent into the sample surface area, and makes the tip of the sampling needle contact the sample surface area. The analysis solvent forms an analysis liquid film on the sample surface area that carries the target compound and stable isotope internal standard. The detection module retains the eluent membrane inside the ion source. Under the conditions of electric field, heat assistance, and airflow assistance of the ion source, the solvent in the eluent membrane gradually evaporates, causing the target compound in the eluent membrane to ionize and form target compound ions. The target compound ions are released in sequence according to the differences in physical migration and energy response and are directed to the mass analysis area, forming an ion signal time series. The output module performs mass spectrometry detection on the time series of ion signals. It acquires characteristic ion signals of target compound ions and stable isotope internal standard ions through multi-reaction monitoring mode, completes qualitative identification, determines the concentration range of the target compound based on the signal intensity ratio, and generates structured detection results.
[0016] The beneficial effects of this invention are as follows: By setting up ion migration paths and applying segmented electric fields inside the ion source, and applying a guiding electric field at the end of the path to guide the formation of ion signal time series in the mass analysis region, in-situ time separation of ions based on intrinsic differences is achieved. This effectively distinguishes isomers or similar mass compounds without external chromatography or additional post-processing, significantly improving multi-component resolution while reducing operational complexity and instrument dependence. Furthermore, by forming an eluent film to support both components, and combining this with subsequent extraction of characteristic intensities based on parent and daughter ion signals and calculation of signal intensity ratios to determine concentration ranges, the physicochemical conditions are unified to the greatest extent, effectively correcting matrix effects and instrument variability. This improves the accuracy and repeatability of concentration estimation, overcoming the problem of insufficient quasi-quantitative reliability caused by inconsistencies between the internal standard and the target environment in existing methods. Attached Figure Description
[0017] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings used in the following description of the embodiments will be briefly introduced. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.
[0018] Figure 1 This is a flowchart of a rapid mass spectrometry method for detecting multi-component compounds in body fluids.
[0019] Figure 2 A block diagram of a rapid mass spectrometry detection device for multi-component compounds in body fluids.
[0020] Figure 3 A flowchart for forming the target compound ions.
[0021] Figure 4 A flowchart for generating a time series of ion signals. Detailed Implementation
[0022] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
[0023] Many specific details are set forth in the following description in order to provide a full understanding of the invention. However, the invention may also be practiced in other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of the invention. Therefore, the invention is not limited to the specific embodiments disclosed below.
[0024] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The phrase "in one embodiment" appearing in different places in this specification does not necessarily refer to the same embodiment, nor is it a single or selective embodiment that is mutually exclusive with other embodiments.
[0025] Reference Figures 1-4 As one embodiment of the present invention, this embodiment provides a rapid mass spectrometry detection method for multi-component compounds in body fluids, comprising the following steps: S1: Collect body fluid samples from the subject and transfer the body fluid samples to the functionalized treatment interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized treatment interface adjacent to the ion source inlet.
[0026] The sampling needle tip comes into contact with the body fluid on the surface of the subject. The surface tension of the needle tip adsorbs the body fluid, forming a sample. The sampling needle carries the sample to the functionalized interface, where it contacts a pre-positioned stable isotope internal standard. The sample is released onto the interface, forming a droplet. This droplet maintains a fixed contact position on the interface, forming a defined region, which serves as the sample surface area. The functionalized interface is a solid surface structure located adjacent to the ion source inlet. This solid surface pre-fixes an acidic chemical environment, functional materials, and a stable isotope internal standard to support the sample and achieve matrix inhibition and target compound enrichment.
[0027] To further explain, the solid surface can be a metal substrate, glass substrate, or silicon substrate. An acidic chemical environment, functional materials, and a stable isotope internal standard are pre-fixed on the solid surface through surface modification. The acidic chemical environment can be achieved by covalently fixing acidic groups (such as sulfonic acid groups or carboxylic acid groups) on the solid surface or by coating and drying the solid surface to form a stable acidic buffer layer, so that a stable acidic environment can be formed when the body fluid sample comes into contact with the solid surface. The functional materials can be fixed to the solid surface by physical coating, sol-gel fixation, or polymer adhesion. For example, C18 modified silica gel particles or polymer microbeads are dispersed in an adhesive solution, uniformly coated on the solid surface, and dried and cured, so that the functional materials are stably attached to the solid surface. The stable isotope internal standard can be deposited on the solid surface by micro-dropping a standard solution containing a stable isotope label and then dried and fixed, so that the stable isotope internal standard can quickly dissolve into the body fluid sample when it comes into contact with the body fluid sample, and thus serve as a quantitative calibration reference in the subsequent mass spectrometry detection process.
[0028] The body fluid sample in the sample surface area is in continuous contact with the acidic chemical environment provided by the functionalized treatment interface. The hydrogen ions in the acidic chemical environment act on the macromolecular components in the body fluid sample, changing the spatial structure of the macromolecular components in the body fluid sample, causing the macromolecular components in the body fluid sample to undergo conformational changes and form an aggregated state. The macromolecular components remain in an inactive state in the aggregated state and remain inside the sample surface area.
[0029] To further explain, an acidic chemical environment can be an acidic solution with a pH of about 2 to 4. Acidic hydrogen ions neutralize the surface charge of proteins and destroy the three-dimensional structure of proteins and other macromolecules, causing proteins and other macromolecules to expose hydrophobic cores and lose their stable dispersion state, thereby increasing intermolecular collisions and adhesion, and ultimately forming aggregates or precipitates; macromolecules include serum proteins, enzymes and peptides, etc.
[0030] After aggregation, the body fluid sample in the sample area simultaneously comes into contact with the surface of the functional material at the functionalized interface. The surface of the functional material is a polymer or silica-based material fixed on a solid carrier, such as C18 modified silica gel or polymer microbeads. Non-target matrix components in the body fluid sample are adsorbed through hydrophobic interactions, electrostatic interactions, or hydrogen bonding. Non-target matrix components include salts, small protein fragments, sugars, and other soluble macromolecules that can interfere with mass spectrometry detection. The target compounds are small molecule drugs and metabolites to be detected in blood, urine, or saliva. Adsorption causes the non-target matrix components to form a retention state on the surface of the functional material. Retention means that the non-target matrix components do not migrate with the electric field in the sample surface area and under the applied ion source electric field, thereby reducing interference with the detection of the target compounds.
[0031] After the non-target matrix components are retained, the target compound and the stable isotope internal standard in the sample surface area undergo an enrichment process at the functionalized treatment interface. The functionalized treatment interface maintains a stable distribution of the target compound and the stable isotope internal standard within the sample surface area, thus ensuring that the target compound and the stable isotope internal standard remain enriched and maintained in the sample surface area.
[0032] It should also be noted that the stable isotope internal standard is a compound with the same structure as the target compound but containing a stable isotope label, such as ^2H or ^13C label, used to correct for sample matrix effects and instrument response fluctuations, thereby ensuring the accuracy of quantitative detection of the target compound. In addition, the acidic chemical environment on the functionalized interface is achieved by covalently immobilizing acidic groups on the surface of the solid support or coating with an acidic buffer layer, so that the acidic environment maintains a stable pH value during body fluid contact and subsequent analysis.
[0033] To further clarify, this method is applicable to a variety of small molecule target compounds with different polarities, volatility, and thermal stability, and is suitable for both human body fluid samples and other mammalian body fluid samples.
[0034] S2: Position the sample surface area inside the ion source, and release the desorption solvent into the sample surface area from inside the ion source by the sampling needle, and make the tip of the sampling needle contact the sample surface area. The desorption solvent forms a desorption liquid film carrying the target compound and stable isotope internal standard on the sample surface area.
[0035] The sampling needle carries the desorption solvent into the ion source, causing the tip of the sampling needle to move to a position above the sample surface area. The tip of the sampling needle gradually approaches the sample surface area until it comes into contact with the sample surface area. The tip of the sampling needle releases the desorption solvent, which spreads on the sample surface area to form a liquid coverage area. The desorption solvent forms a continuous liquid layer on the sample surface area, which covers the sample surface area and forms a local solvent coverage layer.
[0036] To further explain, the eluent is a low-volatility polar or medium-polar organic solvent that can be used alone or mixed with water, buffer solutions, such as methanol, water, acetonitrile, or a mixed solution, to dissolve the target compound and stable isotope internal standard in the sample and form a stable liquid film to achieve the eluent effect.
[0037] The local solvent coating layer comes into contact with the target compound and stable isotope internal standard in the sample surface area. The eluent molecules in the local solvent coating layer undergo solvation with the target compound and stable isotope internal standard, causing the target compound and stable isotope internal standard in the sample surface area to enter the eluent from the functionalized interface and form a dissolved state. The target compound and stable isotope internal standard remain uniformly distributed inside the local solvent coating layer.
[0038] The internal environment of the ion source maintains a stable liquid state for the local solvent coating layer. This local solvent coating layer maintains a continuous liquid film structure on the sample surface. Inside the ion source, it remains in a liquid film state and continuously carries the target compound and stable isotope internal standard. The local solvent coating layer forms an eluent film carrying the target compound and stable isotope internal standard within the ion source. The eluent film refers to the continuous micro-liquid film structure formed by the eluent on the sample surface. The target compound enters the solvent phase through solvent dissolution and interfacial desorption, a mechanism consistent with solvent phase extraction in liquid-liquid extraction or thin-film microextraction. The eluent film maintains a stable liquid film morphology by selecting the aforementioned eluent combination and controlling the internal temperature and airflow conditions of the ion source, ensuring a controlled evaporation rate of the eluent in the ion source environment, thereby maintaining the continuous existence of the eluent film during the ionization of the target compound.
[0039] To further explain, the thickness of the eluent film is typically 50 to 200 micrometers. Under the conditions of thermal and gas flow assistance from the ion source, the film can maintain a stable structure and complete controlled solvent evaporation within a few seconds to tens of seconds, ensuring that the target compound can migrate smoothly to the surface of the film and ionize during the continuous existence of the film.
[0040] S3: The eluent film is retained inside the ion source. Under the conditions of electric field, thermal assistance and airflow assistance of the ion source, the solvent in the eluent film is gradually evaporated, causing the target compound in the eluent film to ionize and form target compound ions. The target compound ions are released in the order of physical migration and energy response differences and guided to the mass analysis region to form an ion signal time series.
[0041] S3.1: The eluent film carrying the target compound and the stable isotope internal standard is kept in the eluent position inside the ion source. The eluent position inside the ion source is formed by arranging adjustable electrodes above the eluent region to create a segmented electric field. The electrodes are arranged at intervals along the surface of the eluent film. An adjustable voltage is applied between the electrodes to form a stable potential gradient above the eluent film, thereby applying the electric field inside the ion source to the eluent film and forming a continuous ionization interface. The eluent film carrying the target compound and the stable isotope internal standard is within the coverage of the electric field inside the ion source, so that the target compound in the eluent film is continuously in the electric field environment.
[0042] The resolving position inside the ion source is simultaneously assisted by both electric heating and airflow. Electric heating continuously provides heat by placing electric heating elements (such as resistance heating plates or heating rings) near the resolving position inside the ion source, creating a stable temperature environment in the area where the resolving liquid film is located, thereby increasing the evaporation rate of the resolving solvent in the resolving liquid film. Airflow creates a stable gas flow channel along the surface direction of the resolving position inside the ion source, carrying away the solvent vapor formed by evaporation on the surface of the resolving liquid film, gradually reducing the amount of resolving solvent in the resolving liquid film and keeping the resolving liquid film in a continuous evaporation state. During the gradual evaporation process, the resolving liquid film maintains its liquid film shape without overall rupture, thus maintaining a stable liquid film existence time at the resolving position inside the ion source.
[0043] To further explain, the electric heating aid generates heat by energizing the electric heating element and transfers the heat to the area where the eluent film is located through heat conduction and heat convection, thereby creating a stable temperature environment around the eluent film; the airflow aid continuously inputs inert gas through a gas channel set inside the ion source to form a directional gas flow, thereby creating a stable gas flow path on the surface of the eluent film and carrying away the volatile solvent vapor.
[0044] During the gradual evaporation of the solvent in the eluent film, the electric field inside the ion source continuously acts on the surface of the eluent film, forming a charged interface region on the surface of the eluent film. The target compound in the eluent film undergoes ionization in the charged interface region and gradually transforms into target compound ions. Under the action of the electric field inside the ion source, the target compound ions are released from the surface of the eluent film into the gas phase space and maintain a stable charged state.
[0045] The process of target compound ionization in the eluent membrane to form target compound ions occurs simultaneously with the gradual evaporation of the eluent membrane. The continuous evaporation of the eluent membrane maintains the migration of the target compound to the membrane surface, while the continuous electric field inside the ion source maintains the ionization process of the target compound. The eluent membrane forms a continuous ionization interface at the eluent position inside the ion source, enabling the target compound to be continuously converted into target compound ions at the eluent position inside the ion source. Compared with the traditional body fluid mass spectrometry detection process that relies on complex sample pretreatment or liquid phase separation steps, the eluent membrane carrying the target compound and stable isotope internal standard directly completes the ionization of the target compound to form target compound ions at the eluent position inside the ion source, realizing the direct conversion of the target compound to target compound ions in the body fluid sample.
[0046] It should also be noted that the segmented electric field can be achieved in the existing mass spectrometer ion source through electrode arrangement and voltage control, without the need to build a new dedicated device.
[0047] S3.2: An ion migration path is established inside the ion source. This path is formed by arranging multiple spaced electrodes along the mass analysis region inside the ion source. These electrodes are arranged sequentially along the ion migration direction, forming a continuous electric field channel. The space between the electrodes constitutes the ion migration path. After the target compound ions are released from the eluent film, they enter the space between the electrodes and move along the electric field direction. The ion migration path extends along the direction in which the target compound ions leave the eluent film and connects to the mass analysis region. Multiple electric field interaction regions are formed sequentially along the ion migration path. Each region forms an electric field gradient through the potential difference between adjacent electrodes. For example, the initial electrode potential of the ion migration path is 0V, the second electrode potential is 30V, the third electrode potential is 60V, and the fourth electrode potential is 90V. A progressively increasing potential gradient is formed between adjacent electrodes. The multiple electric field interaction regions are arranged sequentially along the ion migration path. The potential gradient in the ion migration path propels the target compound ions forward along the path, maintaining a continuous migration state and achieving a controlled migration process.
[0048] To further clarify, the electrode spacing in the ion migration path should be between 0.5 mm and 5 mm. It is recommended that the electrode spacing near the eluent film region be set to 0.5 mm to 2 mm to avoid segmented electric field interference with the ionization of the target compound on the eluent film surface. The electrode spacing in the middle section of the ion migration path can be appropriately increased to 5 mm to form a stable electric field channel and ensure the continuous and controlled movement of the target compound ions during migration. The electric field gradient should be between 5 V / mm and 30 V / mm. The electrode potentials are adjustable parameters. The migration speed of the target compound ions is controlled by adjusting the electrode potential difference, while avoiding excessively high electric field strength that could lead to ion instability or interference with the ionization of the liquid film surface.
[0049] A continuous thermal and gas flow assist is applied to the space containing the ion migration path. The thermal assist is an electric heating assist, which uses a resistance heating element (e.g., heating plate, heating ring, or heating wire) placed near the desorption location inside the ion source or outside the ion migration path to continuously transfer heat to the space containing the ion migration path, creating a stable temperature environment. The adjustable range of the thermal assist temperature is 80°C to 180°C. The specific temperature is adjusted according to the thermal stability of the target compound and the thickness of the desorption liquid film to ensure that the liquid film completes the evaporation of the desorption solvent and the ionization of the target compound in a very short time (e.g., several seconds to tens of seconds). For thermosensitive target compounds, a lower thermal assist temperature can be selected and the gas flow rate can be appropriately increased to maintain the evaporation rate of the desorption solvent, thereby avoiding thermal decomposition or structural changes of the target compound. To maintain the continuous release of target compound ions from the surface of the eluent film, a stable gas flow channel is formed along the ion migration path with the assistance of airflow. Target compound ions are released from the surface of the eluent film into the gas phase space. The gas flow channel propels the target compound ions in the gas phase space into the ion migration path. Due to differences in molecular weight, polarity, and energy response characteristics, different target compound ions migrate at different speeds in the ion migration path. Different target compound ions gradually form a spatial separation state in the ion migration path. The spatial separation state corresponds to the release order of different target compound ions in the ion migration path, thereby enabling target compound ions with different physicochemical properties to be released in order according to differences in physical migration and energy response.
[0050] To further explain, a heat-assisted temperature range of 80°C to 180°C can promote stable evaporation of the eluent solvent and maintain the continuous desorption process of the eluent film, while avoiding thermal decomposition of the target compound due to excessively high temperatures. The airflow assistance uses high-purity nitrogen (>99.999%) or helium to form a directional flow, pushing the target compound ions into the ion migration path along the ion migration path, while carrying away the evaporating solvent. The airflow velocity is in the range of 1 to 3 L / min and can be adjusted according to the migration characteristics of the target compound to ensure stable ion migration and the formation of a time-resolved signal.
[0051] The sequentially released target compound ions continue to move along the ion migration path to the end of the ion migration path. At the end of the ion migration path, a guiding electric field generated by a guiding electrode is formed. The guiding electric field forms a potential gradient in a single direction at the end of the ion migration path. Under the action of the guiding electric field, the sequentially released target compound ions enter the mass analysis region along a fixed direction. Different target compound ions enter the mass analysis region in the order of their arrival time, forming an ion signal time series in the time dimension.
[0052] It should also be noted that the mass analysis region refers to a dedicated area inside the mass spectrometer used for ion mass selection and detection, such as a quadrupole, time-of-flight tube, or ion trap region. This region is used to guide the release of target compound ions in sequence according to different migration velocities for mass spectrometry detection, thereby achieving mass resolution and signal acquisition. This method can be implemented on existing triple quadrupole (QqQ) mass spectrometer platforms, such as AB-Sciex-5500-QTRAP, Thermo-TSQ, or Waters-Xevo-TQ-S, without requiring additional instrument modifications.
[0053] Preferably, the present invention forms a sequential release process of target compound ions through the combined action of ion migration paths and segmented electric fields. The ion signal time series is generated by the migration time difference of target compound ions in the ion migration path. Compared with the traditional method of relying on liquid chromatography separation to form time signals in the body fluid mass spectrometry detection process, the ion migration path and segmented electric field directly form the ion signal time series inside the ion source, so that the target compound ions are released sequentially inside the ion source and enter the mass analysis region, realizing a rapid sequential detection process of multi-component compounds in body fluids.
[0054] S4: Mass spectrometry detection of ion signal time series is performed. Characteristic ion signals of target compound ions and stable isotope internal standard ions are collected through multiple reaction monitoring mode to complete qualitative identification and determine the concentration range of target compound based on the signal intensity ratio, generating structured detection results.
[0055] S4.1: Characteristic ion signals of target compound ions and stable isotope internal standard ions are acquired through multiple reaction monitoring (MRM) mode, specifically: After the ion signal time series is guided from inside the ion source to the mass analysis region, it is directly entered into the mass spectrometry detection. The mass analysis region performs a mass scan on the ion signal time series and starts the multi-reaction monitoring mode.
[0056] To further explain, multiple reaction monitoring (MRM) is a commonly used scanning mode in existing mass spectrometry techniques. It is used to simultaneously select the parent ion of a target compound and induce its fragmentation to generate daughter ions, subsequently detecting the mass signal of the daughter ions to achieve highly selective and sensitive qualitative and quasi-quantitative analysis. It is widely used in triple quadrupole mass spectrometer platforms.
[0057] Before starting the multi-reaction monitoring mode, the mass spectrometer predefines the conversion relationship between the parent ion mass and the daughter ion mass of the target compound. The conversion relationship between the parent ion mass and the daughter ion mass is determined based on the molecular structure and fragmentation mode of the target compound. For example, when the target compound is caffeine (as an example only), the parent ion mass is 194.1 Da, and the daughter ion mass is 138.1 Da and 110.1 Da, respectively. The conversion relationship between the parent ion mass and the daughter ion mass is stored as a fixed parameter.
[0058] After the multiple reaction monitoring mode is activated, the mass analysis region performs a mass scan on the ion signal time series and captures the parent ion signal of the target compound ion. The parent ion signal corresponds to the initial charged molecular mass signal of the target compound ion when it enters the mass spectrometry detection process. The peak intensity of the parent ion signal is recorded as the signal intensity value during the mass spectrometry detection process. At the same time, the mass spectrometry detection process induces the fragmentation of the parent ion and captures the daughter ion signal. The daughter ion signal corresponds to the mass signal of the ion determined according to the mass conversion relationship between the parent ion mass and the daughter ion mass after the parent ion fragmentation. The peak intensity of the daughter ion signal is recorded as the signal intensity value during the mass spectrometry detection process.
[0059] In the multiple reaction monitoring (MRM) mode, the mass analysis region simultaneously scans the parent ion signal of the stable isotope internal standard ion in the ion signal time series. The parent ion signal of the stable isotope internal standard ion corresponds to the initial charged molecular mass signal when the stable isotope internal standard ion enters the mass analysis region. The mass analysis region records the peak intensity of the parent ion signal of the stable isotope internal standard ion as the signal intensity value and induces the fragmentation of the parent ion of the stable isotope internal standard ion to capture the daughter ion signal. The daughter ion signal of the stable isotope internal standard ion corresponds to the mass signal of a specific fragment ion after the stable isotope internal standard ion fragmentation. The mass analysis region records the peak intensity of the daughter ion signal of the stable isotope internal standard ion as the signal intensity value.
[0060] S4.2: Determine the concentration range of the target compound, specifically: The acquired precursor ion signal and daughter ion signal of the target compound ion enter the extraction process. The mass analysis region selects the part with the highest peak intensity from the precursor ion signal of the target compound ion as the characteristic ion signal intensity of the target compound ion, and selects the part with the highest peak intensity from the daughter ion signal of the target compound ion as the characteristic ion signal intensity of the target compound ion. The peak intensities of the precursor ion signal and daughter ion signal of the target compound ion are combined to obtain the characteristic ion signal intensity value of the target compound ion.
[0061] The parent ion signal and daughter ion signal of the stable isotope internal standard ion enter the extraction process. The mass analysis region selects the part with the highest peak intensity from the parent ion signal of the stable isotope internal standard ion as the characteristic ion signal intensity of the stable isotope internal standard ion, and at the same time selects the part with the highest peak intensity from the daughter ion signal of the stable isotope internal standard ion as the characteristic ion signal intensity of the stable isotope internal standard ion. The peak intensities of the parent ion signal and daughter ion signal of the stable isotope internal standard ion are used as the characteristic ion signal intensity values of the stable isotope internal standard ion.
[0062] The signal intensity ratio is calculated by comparing the characteristic ion signal intensity values of the target compound ion with those of the stable isotope internal standard ion. This signal intensity ratio reflects the relative abundance of the target compound in the body fluid sample, and is expressed as follows: ; in, Indicates the ratio of signal strength. The numerical value representing the characteristic ion signal intensity of the target compound ion. The numerical value representing the characteristic ion signal intensity of stable isotope internal standard ions.
[0063] The signal intensity ratio is matched with a preset concentration range. The preset concentration range is established based on a standard curve. For example, the concentration range corresponding to a signal intensity ratio in the range of 0.1 to 1.0 is 0.1 ng / mL to 1.0 ng / mL. The concentration range of the target compound is determined according to the preset concentration range in which the signal intensity ratio falls.
[0064] It should also be noted that the standard curve is obtained by collecting the characteristic ion signal intensity ratio of the target compound ion and the stable isotope internal standard ion under known concentration gradient conditions; the standard curve is used to establish the correspondence between the signal intensity ratio and the concentration range of the target compound. This method is mainly used for rapid screening and quasi-quantitative detection of multi-component compounds in body fluids.
[0065] S4.3: Generate structured detection results, specifically: The quality analysis region identifies the specific time window signal corresponding to the target compound ion from the ion signal time series. The specific time window signal is defined based on the migration time difference in the ion migration path, and the specific time window signal is converted into a digital record to establish the target compound ion time series record.
[0066] By binding the signal characteristics of the target compound ion time series record with the numerical value of the target compound concentration range, a target compound detection record containing qualitative signals and quasi-quantitative ranges is formed.
[0067] Multiple target compound detection records are organized into a fixed-format data package, which includes the target compound name, qualitative confirmation results, and concentration range values. The fixed-format data package serves as a structured detection result.
[0068] It should also be noted that this invention is applicable to various body fluid types, including human blood, plasma, serum, urine, and saliva samples. Different body fluid samples are detected through the same ionization process after forming a sample surface area at the functionalized treatment interface. For different body fluid matrices, parameters can be optimized by adjusting the acidity of the functionalized treatment interface, the type of functional material, and the composition of the eluent. It is also applicable to target compounds with different physicochemical properties, including small molecule compounds with large polarity differences, drugs and metabolites with different volatility, and organic compounds with different thermal stability, such as amphetamine compounds, opioids, sedative-hypnotic drugs, and related metabolites. For different types of target compounds, detection conditions can be optimized by adjusting the type of eluent, the ion source temperature, and the electric field parameters of the ion migration path.
[0069] This embodiment also provides a rapid mass spectrometry detection device for multi-component compounds in body fluids, including: The acquisition module collects body fluid samples from the subject and transfers the body fluid samples to the functionalized processing interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized processing interface adjacent to the ion source inlet. The analysis module positions the sample surface area inside the ion source, releases the analysis solvent into the sample surface area, and makes the tip of the sampling needle contact the sample surface area. The analysis solvent forms an analysis liquid film on the sample surface area that carries the target compound and stable isotope internal standard. The detection module retains the eluent membrane inside the ion source. Under the conditions of electric field, heat assistance, and airflow assistance of the ion source, the solvent in the eluent membrane gradually evaporates, causing the target compound in the eluent membrane to ionize and form target compound ions. The target compound ions are released in sequence according to the differences in physical migration and energy response and are directed to the mass analysis area, forming an ion signal time series. The output module performs mass spectrometry detection on the time series of ion signals. It acquires characteristic ion signals of target compound ions and stable isotope internal standard ions through multi-reaction monitoring mode, completes qualitative identification, determines the concentration range of the target compound based on the signal intensity ratio, and generates structured detection results.
[0070] This embodiment also provides a computer device suitable for a rapid mass spectrometry detection method for multi-component compounds in body fluids, comprising: a memory and a processor; the memory is used to store computer-executable instructions, and the processor is used to execute the computer-executable instructions to realize the rapid mass spectrometry detection method for multi-component compounds in body fluids as proposed in the above embodiment.
[0071] The computer device can be a terminal, comprising a processor, memory, communication interface, display screen, and input devices connected via a system bus. The processor provides computing and control capabilities. The memory includes non-volatile storage media and internal memory. The non-volatile storage media stores the operating system and computer programs. The internal memory provides an environment for the operation of the operating system and computer programs stored in the non-volatile storage media. The communication interface is used for wired or wireless communication with external terminals; wireless communication can be achieved through Wi-Fi, carrier networks, NFC (Near Field Communication), or other technologies. The display screen can be an LCD screen or an e-ink screen. The input devices can be a touch layer covering the display screen, buttons, a trackball, or a touchpad on the computer device's casing, or an external keyboard, touchpad, or mouse.
[0072] This embodiment also provides a storage medium storing a computer program that, when executed by a processor, implements the rapid mass spectrometry detection method for multi-component compounds in body fluids as proposed in the above embodiments. The storage medium can be implemented by any type of volatile or non-volatile storage device or a combination thereof, such as Static Random Access Memory (SRAM), Electrically Erasable Programmable Read-Only Memory (EEPROM), Erasable Programmable Read Only Memory (EPROM), Programmable Red-Only Memory (PROM), Read-Only Memory (ROM), magnetic storage, flash memory, magnetic disk, or optical disk.
[0073] In summary, this invention achieves in-situ time-separation of ions based on intrinsic differences by setting up ion migration paths within the ion source and applying segmented electric fields, and by applying guiding electric fields at the end of the paths to guide the formation of ion signal time series in the mass analysis region. This effectively distinguishes isomers or similarly massed compounds without external chromatography or additional post-processing, significantly improving multi-component resolution while reducing operational complexity and instrument dependence. Furthermore, by forming an eluent film to support both components, and combining this with subsequent extraction of characteristic intensities based on parent and daughter ion signals and calculation of signal intensity ratios to determine concentration ranges, the invention maximizes the uniformity of physicochemical conditions, effectively corrects matrix effects and instrument variability, and improves the accuracy and repeatability of concentration estimation. This overcomes the problem of insufficient quasi-quantitative reliability caused by inconsistencies between the internal standard and the target environment in existing methods.
[0074] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all such modifications or substitutions should be covered within the scope of the claims of the present invention.
Claims
1. A rapid mass spectrometry method for detecting multi-component compounds in body fluids, characterized in that: include, Body fluid samples were collected from the subjects and transferred to the functionalized treatment interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized treatment interface adjacent to the ion source inlet. The sample surface area is positioned inside the ion source. The sampling needle releases the desorption solvent into the sample surface area from inside the ion source, and the tip of the sampling needle contacts the sample surface area. The desorption solvent forms a desorption liquid film on the sample surface area that carries the target compound and stable isotope internal standard. The eluent film is retained inside the ion source. Under the conditions of electric field, thermal assistance and airflow assistance of the ion source, the solvent in the eluent film is gradually evaporated, the target compound in the eluent film is ionized to form target compound ions, and the target compound ions are released in the order of physical migration and energy response differences and guided to the mass analysis region to form an ion signal time series. Mass spectrometry was used to detect the time series of ion signals. Characteristic ion signals of target compound ions and stable isotope internal standard ions were collected through multiple reaction monitoring mode. Qualitative identification was completed, and the concentration range of the target compound was determined based on the signal intensity ratio, generating structured detection results.
2. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The body fluid samples of the tested subjects were obtained by directly dipping a sampling needle into the body fluid on the surface of the tested subjects.
3. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The formation of the sample surface region specifically refers to: The body fluid sample collected by the sampling needle is introduced into the functionalized processing interface of the pre-set stable isotope internal standard, and the body fluid sample droplets are formed on the functionalized processing interface and defined as the sample surface area. Exposing the body fluid sample in the sample surface area to the acidic chemical environment of the functionalized treatment interface causes the macromolecular components in the body fluid sample to undergo conformational changes and aggregate and become inactive. By bringing the body fluid sample in the sample surface area into contact with the functional material of the functionalized treatment interface, the non-target matrix components in the body fluid sample are retained at the functionalized treatment interface and the interference is weakened. The target compound and stable isotope internal standard in the sample surface region are enriched at the functionalization interface and maintained in the sample surface region.
4. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The formation of the eluent film carrying the target compound and the stable isotope internal standard specifically involves: The desorption solvent is introduced into the sample surface area from inside the ion source by a sampling needle, and the tip of the sampling needle touches the sample surface area to form a local solvent coating layer in the sample surface area. The target compound and stable isotope internal standard in the sample surface area are desorbed by a local solvent coating layer, and the liquid film morphology of the local solvent coating layer is maintained inside the ion source to form an eluent film carrying the target compound and stable isotope internal standard.
5. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The formation of the target compound ion specifically involves: The eluent film carrying the target compound and stable isotope internal standard is kept in the eluent position inside the ion source, and an electric field is established inside the ion source to act on the eluent film. Heat assistance and airflow assistance are applied to the desorption location inside the ion source to drive the desorption solvent in the desorption liquid film to gradually evaporate and maintain the desorption liquid film in a volatile state; During the gradual evaporation of the eluent membrane, the electric field inside the ion source is maintained, causing the target compound in the eluent membrane to ionize and form target compound ions.
6. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The formation of the ion signal time series is specifically as follows: An ion migration path is set up inside the ion source and a segmented electric field is applied to the ion migration path to enable the target compound ions to migrate in a controlled manner along the ion migration path. Simultaneous application of thermal and gas flow assistance along the ion migration path alters the time distribution of target compound ions released from the eluent film into the gas phase, causing target compound ions with different physicochemical properties to be released sequentially according to differences in physical migration and energy response. By setting a guiding electrode at the end of the ion migration path and applying a guiding electric field, the target compound ions released sequentially are guided to the mass analysis region, forming an ion signal time series in the time dimension.
7. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The characteristic ion signals of the target compound ions and stable isotope internal standard ions acquired through the multi-reaction monitoring mode are specifically as follows: During mass spectrometry detection, a multi-reaction monitoring mode is set and the parent ion-daughter ion conversion relationship corresponding to the target compound ion is preset; In multi-reaction monitoring mode, the parent ion signal and daughter ion signal of the target compound are acquired simultaneously and the signal intensity is recorded. In the multi-reaction monitoring mode, the parent ion signal and daughter ion signal of stable isotope internal standard ions are acquired simultaneously and the signal intensity is recorded.
8. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The determination of the target compound concentration range specifically involves: The characteristic ion signal intensity of the target compound ion is extracted from the parent ion signal and daughter ion signal of the target compound ion; The characteristic ion signal intensity of stable isotope internal standard ions is extracted from the parent ion signal and daughter ion signal of stable isotope internal standard ions. Calculate the ratio of the characteristic ion signal intensity of the target compound ion to the characteristic ion signal intensity of the stable isotope internal standard ion, and determine the concentration range of the target compound based on the preset concentration range correspondence.
9. The rapid mass spectrometry detection method for multi-component compounds in body fluids as described in claim 1, characterized in that: The generation of structured detection results specifically includes: Extract the time series signals corresponding to the target compound ions from the ion signal time series and establish the target compound ion time series record; The target compound ion time series records are correlated with the target compound concentration range to form a target compound detection record; The detection records of the target compound are uniformly packaged and structured detection results are generated.
10. A rapid mass spectrometry detection device for multi-component compounds in body fluids, based on the rapid mass spectrometry detection method for multi-component compounds in body fluids according to any one of claims 1 to 9, characterized in that: include, The acquisition module collects body fluid samples from the subject and transfers the body fluid samples to the functionalized processing interface of the pre-set stable isotope internal standard, forming a sample surface area on the functionalized processing interface adjacent to the ion source inlet. The analysis module positions the sample surface area inside the ion source, releases the analysis solvent into the sample surface area, and makes the tip of the sampling needle contact the sample surface area. The analysis solvent forms an analysis liquid film on the sample surface area that carries the target compound and stable isotope internal standard. The detection module retains the eluent membrane inside the ion source. Under the conditions of electric field, heat assistance, and airflow assistance of the ion source, the solvent in the eluent membrane gradually evaporates, causing the target compound in the eluent membrane to ionize and form target compound ions. The target compound ions are released in sequence according to the differences in physical migration and energy response and are directed to the mass analysis area, forming an ion signal time series. The output module performs mass spectrometry detection on the time series of ion signals. It acquires characteristic ion signals of target compound ions and stable isotope internal standard ions through multi-reaction monitoring mode, completes qualitative identification, determines the concentration range of the target compound based on the signal intensity ratio, and generates structured detection results.