阅读说明：本技术 一种直接采样电离分析系统及方法、应用 (Direct sampling ionization analysis system and method and application ) 是由 欧阳证 张文鹏 焦斌 于 2021-09-02 设计创作，主要内容包括：本发明公开了一种直接采样电离分析系统,包括：采样模块,用于接触采集待测样品,采集待测成分；试剂盒模块,用于固定采样模块,洗脱待测成分,获得洗脱溶液；反应模块,用于对洗脱溶液进行高通量的化学反应；进样模块,用于将试剂盒模块固定在质谱仪的进样端,使得进样模块和质谱仪配合进样；采样模块为一端固定有多孔膜的金属探针,多孔膜用于接触采集待测样品。本发明涉及的直接采样电离分析系统,通过用一端固定有多孔膜的金属探针进行采样,操作便捷,且能迅速得到测试结果,适用于现场快速检测、临床快检等应用,满足了快速取样分析的需求,大幅简化质谱分析的样品前处理过程。本发明还提供了一种使用上述系统的一种直接采样电离分析方法。(The invention discloses a direct sampling ionization analysis system, comprising: the sampling module is used for collecting a sample to be detected in a contact manner and collecting components to be detected; the reagent box module is used for fixing the sampling module, eluting the component to be detected and obtaining an elution solution; the reaction module is used for carrying out high-flux chemical reaction on the elution solution; the sample injection module is used for fixing the reagent box module at a sample injection end of the mass spectrometer, so that the sample injection module and the mass spectrometer are matched for sample injection; the sampling module is a metal probe with one end fixed with a porous membrane, and the porous membrane is used for collecting a sample to be detected in a contact manner. The direct sampling ionization analysis system provided by the invention is convenient to operate and can quickly obtain a test result by sampling with the metal probe with one end fixed with the porous membrane, is suitable for the applications of on-site quick detection, clinical quick detection and the like, meets the requirements of quick sampling analysis, and greatly simplifies the sample pretreatment process of mass spectrometry. The invention also provides a direct sampling ionization analysis method using the system.)

1. A direct sampling ionization analysis system, comprising:

the sampling module is used for contacting a sample to be detected and collecting components to be detected;

the reagent box module is used for fixing the sampling module and eluting the component to be detected to obtain an elution solution;

a reaction module for performing a high-throughput chemical reaction on the elution solution;

the sample injection module is used for fixing the kit module at a sample injection end of a mass spectrometer, so that the sample injection module and the mass spectrometer are matched for sample injection;

the sampling module is a metal probe with one end fixed with a porous membrane, and the porous membrane is used for collecting the sample to be detected in a contact manner.

2. The direct sampling ionization analysis system of claim 1, wherein the porous membrane further comprises a chemical reagent modified thereon for improving contact collection conditions.

3. The direct sampling ionization analysis system of claim 1 or 2, wherein the porous membrane has a pore size of 0.1 to 10 μ ι η.

4. The direct sampling ionization analysis system of claim 3, wherein a nano-spray glass tube is fixed on the reagent box module, an elution solvent for eluting the component to be detected is contained in the nano-spray glass tube, and the metal probe is inserted into the nano-spray glass tube after collection; and after voltage is applied to the metal probe, electrospray is formed at the tip of the nano-spray glass tube.

5. The direct sampling ionization analysis system of claim 4, wherein the metal probes are used to immobilize the porous membrane, and the outer diameter of the metal probes is 0.2 mm to 1 mm.

6. The direct sampling ionization analysis system of claim 5, wherein the sample module comprises a single mass spectrometry sample injector and a sequential mass spectrometry sample injector.

7. A direct sampling ionization analysis method based on the direct sampling ionization analysis system according to any one of claims 1, 2, 4, 5 and 6, characterized by comprising the steps of:

(1) contacting a porous membrane of a metal probe with a sample to be detected, and absorbing components to be detected;

(2) placing the metal probe in a cleaning solution to remove adhered matrix components;

(3) inserting the metal probe into a nano-spray glass tube filled with an elution solvent, and standing or shaking for elution to obtain an elution solution;

(4) in the reaction module, performing a high-throughput chemical reaction on the elution solution;

(5) and carrying out mass spectrometry on the eluted solution after the reaction to obtain chemical molecular information in the sample to be detected.

8. The direct sampling ionization analysis method of claim 7 wherein, when the sample to be tested is a solid, the porous membrane is in contact with the sample to be tested in a manner that: inserting the porous membrane into the sample to be detected or rolling the porous membrane on the surface of the sample to be detected; when the sample to be detected is liquid, the porous membrane is contacted with the sample to be detected in a manner that the porous membrane is inserted into the sample to be detected.

9. The direct sampling ionization analysis method of claim 8 wherein the means for sampling using the metal probe comprises sampling directly with the metal probe and sampling with a kit.

10. Use of the direct sampling ionization analysis method of claim 8 comprising qualitative and quantitative analyses in which the direct sampling ionization analysis method is used to determine elemental composition, sn isomers of phospholipids and carbon-carbon double bond positional isomers; in the quantitative analysis aspect, the direct sampling ionization analysis method is used for realizing the quantitative detection of the component to be detected by adding an internal standard substance on the porous membrane or in the elution solution.

Technical Field

The invention belongs to the technical field of mass spectrometry, and particularly relates to a direct sampling ionization analysis system, a method and application.

Background

In recent years, the characteristics of mass spectrometers, such as high sensitivity, high accuracy and high throughput, have made them play an important role in the analysis of complex mixtures, such as biological tissues, body fluids and environmental samples. The mass spectrometer can be used for detecting most compounds in a sample to be detected, so that the structure identification of target compounds, the quantitative analysis of low-content medicines and biomarkers in vivo and the like are realized. In addition to on-site rapid detection of drugs, prohibited drugs and the like, rapid detection is also in great demand in the medical field, and disease markers detected by using a mass spectrometer play more and more roles in clinical diagnosis. Related studies have shown that the ratio of carbon-carbon double bond isomers of unsaturated phospholipids is greatly different in patients with diabetes, breast cancer, and the like, compared to normal persons.

However, the conventional sample pretreatment method consumes a long time, is not matched with the rapid detection capability of a mass spectrometer, is not beneficial to improving the detection efficiency, and cannot give an analysis result in time. In recent years, the quantity of adsorbed substances to be detected is limited by using modes such as direct sampling by a metal probe and the like, so that low-content target substances are difficult to detect, and simultaneously, solid analysis substances carried by the substances can block a sample introduction passage, thereby causing troubles for analysis; when the solid-phase microextraction mode is used for sampling, the sampling process is long in time consumption, the extraction recovery rate is low, selectivity exists, and some key chemical information can be lost. Meanwhile, the above method cannot realize high-throughput sampling, reaction and analysis of biological samples, and cannot accurately and efficiently obtain the required analysis result. Therefore, there is a need to develop a novel sampling technique that can be used with a mass spectrometer to achieve micro and quantitative sampling and simultaneously preserve the complete chemical information of the sample, so as to meet the requirements of rapid sampling and analysis of the mass spectrometer.

Disclosure of Invention

Aiming at the technical problems of long time consumption, chemical information loss and the like in the sample sampling process in the prior art, the invention aims to provide a direct sampling ionization analysis system, a method and application.

To achieve the above object, the present invention provides a direct sampling ionization analysis system, comprising: the sampling module is used for contacting a sample to be detected and collecting components to be detected; the reagent box module is used for fixing the sampling module, eluting the component to be detected and obtaining an elution solution; the reaction module is used for carrying out high-flux chemical reaction on the elution solution; the sample injection module is used for fixing the reagent box module at a sample injection end of the mass spectrometer, so that the sample injection module and the mass spectrometer are matched for sample injection; the sampling module is a metal probe with one end fixed with a porous membrane, and the porous membrane is used for collecting a sample to be detected in a contact manner.

The reaction module of the invention can carry out high-flux chemical reaction on the elution solution, thereby being convenient for subsequent monitoring. The advance kind module is for can fixing the device at the advancing kind end of mass spectrograph with the reagent box module for advance kind with the mass spectrograph cooperation, in order to promote analysis efficiency.

Further, the porous membrane may also include a chemical agent modified thereon for improving contact collection conditions.

The porous membrane can collect substances to be detected in samples to be detected, such as solid tissues or biological fluids, and the like, and is of a porous structure, the porous structure can collect more substances to be detected, and eluent obtained through subsequent elution can be used for mass spectrometry, so that corresponding substances to be detected can be detected, and corresponding analysis is completed. According to the analysis requirement, the porous structure of the porous membrane is chemically modified to improve the sampling condition and better sample the object to be detected, and the chemical modification is to add other chemical reagents on the porous structure of the porous membrane.

Further, the pore diameter of the porous membrane is 0.1 to 10 μm.

Furthermore, a nano-spray glass tube is fixed on the reagent box module, an elution solvent for eluting the component to be detected is contained in the nano-spray glass tube, and the metal probe is inserted into the nano-spray glass tube after collection; after applying a voltage to the metal probe, an electrospray was formed at the tip of the nano-spray glass tube.

Further, a metal probe is used for immobilizing the porous membrane, and the outer diameter of the metal probe is 0.2 to 1 mm.

Further, the sample injection module comprises a single mass spectrometry sample injection device and a sequence mass spectrometry sample injection device.

The invention also provides a direct sampling ionization analysis method, which comprises the following steps:

(1) contacting a porous membrane of a metal probe with a sample to be detected, and absorbing components to be detected;

(2) placing the metal probe in a cleaning solution to remove the adhered matrix component;

(3) inserting the metal probe into a nano-spray glass tube filled with an elution solvent, standing or shaking for elution to obtain an elution solution;

(4) in the reaction module, carrying out high-flux chemical reaction on the elution solution;

(5) and carrying out mass spectrometry on the reacted elution solution to obtain chemical molecular information in the sample to be detected.

Further, when the sample to be detected is a solid, the contact mode of the porous membrane and the sample to be detected is as follows: inserting the porous membrane into the sample to be detected or rolling the porous membrane on the surface of the sample to be detected; when the sample to be detected is liquid, the porous membrane is contacted with the sample to be detected in a manner that the porous membrane is inserted into the sample to be detected.

Further, the sampling mode using the metal probe includes directly sampling with the metal probe and matching the metal probe with the kit.

The specific using method for matching the metal probe and the kit for sampling comprises the following steps: firstly, taking out a half of the kit fixed with the metal probe, then contacting the porous membrane of the metal probe with a sample to be detected for a period of time, then pushing the metal probe to a proper position to facilitate subsequent operations such as elution and mass spectrometry, and finally splicing the two parts of the kit to form the complete kit.

Further, the application of the direct sampling ionization analysis method comprises qualitative analysis and quantitative analysis, and in the aspect of qualitative analysis, the direct sampling ionization analysis method is used for determining the composition of elements, sn isomers of phospholipid and carbon-carbon double bond position isomers; in the aspect of quantitative analysis, the direct sampling ionization analysis method is used for realizing the quantitative detection of the components to be detected by adding an internal standard substance on a porous membrane or in an elution solution.

Compared with the prior art, the invention has the technical effects that: the direct sampling ionization analysis system provided by the invention has the advantages that the metal probe with one end fixed with the porous membrane is used for sampling, the operation is convenient, the test result can be quickly obtained, the direct sampling ionization analysis system is suitable for the applications of on-site quick detection, clinical quick detection and the like, the requirement of quick sampling analysis is met, and the sample pretreatment process of mass spectrometry is greatly simplified; in addition, the high-throughput reaction and analysis method improves the overall analysis efficiency. The invention also relates to a direct sampling ionization analysis method using the system and application of the direct sampling ionization analysis method.

Drawings

The foregoing and/or additional aspects and advantages of the present invention will become apparent and readily appreciated from the following description of the embodiments, taken in conjunction with the accompanying drawings of which:

FIG. 1 is a schematic diagram of the overall structure of a direct sampling ionization analysis system of the present invention;

FIG. 2 is a schematic diagram of the configuration of a direct sampling ionization analysis system of the present invention in combination with a mass spectrometer;

FIG. 3 is a schematic structural diagram of a metal probe according to the present invention;

FIG. 4 is an electron micrograph of a porous membrane of the present invention before and after modification;

FIG. 5 is a flow chart of the operation of the direct sampling ionization analysis method of the present invention;

FIG. 6 is a schematic diagram of a sampling mode using a metal probe;

FIG. 7 is a positive ion mode mass spectrum of murine brain tissue;

FIG. 8 is a high-quality terminal anion mode mass spectrum of rat brain tissue;

FIG. 9 is a mass spectrum of a low mass end anion mode of rat brain tissue;

FIG. 10 is a comparison graph of lipid signals of the same rat brain tissue extracted using the extraction method of the present invention and the conventional extraction method;

FIG. 11 is a mass spectrum of isomers of fatty acid carbon-carbon double bonds in murine brain tissue;

FIG. 12 is the mass spectrum of the sn isomer and the carbon-carbon double bond isomer of the lipid of the rat brain tissue.

Description of reference numerals:

the device comprises a sampling module 1, a kit module 2, a reaction module 3, a sample introduction module 4, a mass spectrometer 5, a metal probe 6, a porous membrane 7, a sample to be detected 8 and a kit 9.

Detailed Description

Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to the same or similar elements or elements having the same or similar function throughout. The embodiments described below with reference to the drawings are illustrative and intended to be illustrative of the invention and are not to be construed as limiting the invention.

A direct sampling ionization analysis system and method according to an embodiment of the present invention will be described with reference to the accompanying drawings.

As shown in fig. 1-4, the direct sampling ionization analysis system includes: the sampling module 1 is used for contacting a sample 8 to be detected and collecting components to be detected; the kit module 2 is used for fixing the sampling module 1 and eluting the component to be detected to obtain an elution solution; the reaction module 3 is used for carrying out high-flux chemical reaction on the elution solution; the sample injection module 4 is used for fixing the reagent box module 2 at a sample injection end of the mass spectrometer 5, so that the sample injection module 4 and the mass spectrometer 5 are matched for sample injection; the sampling module 1 is a metal probe 6 with one end fixed with a porous membrane 7, and the porous membrane 7 is used for collecting a sample 8 to be detected in a contact manner.

The module is a metal probe 6 with a porous membrane 7 fixed at one end and is used for directly contacting a sample 8 to be detected and collecting components to be detected. The metal probe 6 is used for fixing the porous membrane 7, the outer diameter of the metal probe 6 is 0.2-1mm, and the metal probe 6 is a metal wire such as a platinum wire or a stainless steel wire. The porous membrane 7 is an integrated membrane material fixed on the metal probe 6, the aperture of the porous membrane 7 is 0.1-10 μm, and the porous membrane 7 is made of nylon, polyether sulfone, polytetrafluoroethylene, mixed cellulose or polypropylene and is used for adsorbing a sample 8 to be detected. The porous membrane 7 can collect substances to be detected in a sample 8 to be detected, such as solid tissues or biological fluids, and the like, the porous membrane 7 is of a porous structure, the porous structure can collect more substances to be detected, and eluent obtained through subsequent elution can be used for mass spectrometry, so that corresponding substances to be detected can be detected, and corresponding analysis is completed. According to the analysis requirement, the porous structure of the porous membrane 7 is chemically modified to improve the sampling condition and better sample the object to be detected, and the chemical modification is to add other chemical structures on the porous structure of the porous membrane 7. Taking a polypropylene film as an example, Polydopamine (PDA) and cysteine (Cys) are modified on a porous structure of the polypropylene film, as shown in fig. 4, fig. 4(a), fig. 4(c), and fig. 4(e) are electron micrographs of 5000X, 500X, and 50X magnification of the modified polypropylene film, respectively, fig. 4(b), fig. 4(d), and fig. 4(f) are electron micrographs of 5000X, 500X, and 50X magnification of the polypropylene film before modification, respectively, and it can be seen from comparing fig. 4(a) and fig. 4(b) that polydopamine and cysteine have been modified on the polypropylene film, as shown in a part outlined by a dotted line in fig. 4 (a). In addition, electron micrographs of the polypropylene film with different magnifications before and after modification show that the polypropylene film has uniform aperture and good shape and is suitable for the invention.

The kit module 2 is adapted to the sampling module 1 and used for fixing the sampling module 1 and eluting the component to be detected to obtain an elution solution. A nano-spray glass tube is fixed on the reagent box module 2, an elution solvent is contained in the nano-spray glass tube, the metal probe 6 is inserted into the nano-spray glass tube after collection, the elution solvent is used for eluting the component to be detected on the porous membrane 7 to obtain an elution solution, and the elution solution is contained in the nano-spray glass tube. The mass spectrometer 5 provides a voltage that is applied to the metal probe 6 such that electrospray forms at the nanotube tip. The reaction module 3 is used for performing a high-throughput chemical reaction on the elution solution. The reaction module 3 may have a multi-chamber structure, and one reagent cartridge 9 may be disposed in one chamber to perform a chemical reaction on the elution solutions in a plurality of reagent cartridges 9 at the same time. The chemical reactions in the reaction module 3 include photochemical reactions, peroxidation, and epoxidation reactions. The chemical reaction in the reaction module 3 is not limited to the above reaction types, and in practical applications, whether the chemical reaction is performed or not can be selected according to the properties and requirements of the sample 8 to be tested. The sample injection module 4 is used for fixing the reagent box module 2 at a sample injection end of the mass spectrometer 5, so that the sample injection module 4 and the mass spectrometer 5 are matched for sample injection, and the sample injection module 4 comprises a single mass spectrometry sample injection device and a sequence mass spectrometry sample injection device.

The invention also provides a direct sampling ionization analysis method, the operation flow chart of which is shown in figure 5, and the method comprises the following steps:

(1) contacting the porous membrane 7 of the metal probe 6 with a sample 8 to be detected, and absorbing components to be detected;

(2) placing the metal probe 6 in a cleaning solution to remove the adhered matrix component;

(3) inserting the metal probe 6 into a nano-spray glass tube filled with an elution solvent, standing or shaking for elution to obtain an elution solution;

(4) in the reaction module 3, a high-flux chemical reaction is performed on the elution solution;

(5) and carrying out mass spectrometry on the elution solution after the reaction to obtain chemical molecular information in the sample 8 to be detected.

Wherein, when the sample 8 to be measured is a solid, the contact mode of the porous membrane 7 and the sample 8 to be measured is as follows: inserting the porous membrane 7 into the sample 8 to be detected or rolling the porous membrane 7 on the surface of the sample 8 to be detected; when the sample 8 to be measured is a liquid, the porous membrane 7 is in contact with the sample 8 to be measured in such a manner that the porous membrane 7 is inserted into the sample 8 to be measured. As shown in fig. 6, the sampling using the metal probe 6 includes directly sampling with the metal probe 6 and the kit 9. The specific using method for sampling by matching the metal probe 6 and the kit 9 is as follows: firstly, taking out a half of the kit 9 fixed with the metal probe 6, then contacting the porous membrane 7 of the metal probe 6 with a sample 8 to be detected for a period of time, then pushing the metal probe 6 to a proper position to facilitate subsequent operations such as elution and mass spectrometry, and finally splicing the two parts of the kit 9 to form the complete kit 9.

In addition, the application of the direct sampling ionization analysis method comprises qualitative analysis and quantitative analysis, and in the aspect of qualitative analysis, the direct sampling ionization analysis method is used for determining the composition of elements, sn isomers of phospholipid and carbon-carbon double bond position isomers; in the aspect of quantitative analysis, the direct sampling ionization analysis method is used for realizing the quantitative detection of the components to be detected by adding an internal standard substance on a porous membrane or in an elution solution. In the aspect of qualitative detection, the element composition can be determined, and the structures such as sn isomers, carbon-carbon double bond position isomers and the like of phospholipid can be determined by means of photochemical derivatization reaction, epoxidation reaction and the like in combination with cascade mass spectrometry and the like; in the quantitative detection aspect, absolute quantitative analysis of the substance to be detected can be achieved by adding an internal standard substance on the porous membrane or in the elution solution.

The method of the invention is used for monitoring the sample rat brain tissue, and fig. 7, 8 and 9 are respectively a positive ion mode mass spectrogram of the rat brain tissue, a high-quality terminal negative ion mode mass spectrogram of the rat brain tissue and a low-quality terminal negative ion mode mass spectrogram of the rat brain tissue. The positive ion mode spectrum peak of the murine brain tissue lipid can be obtained from fig. 7, the negative ion mode spectrum peak of the murine brain tissue lipid can be obtained from fig. 8, and the spectrum peak of the murine brain tissue fatty acid can be obtained from fig. 9. The method can be proved to be capable of effectively obtaining the lipid and fatty acid information in the rat brain tissue, and the sample pretreatment system can effectively extract the lipid, thereby facilitating the subsequent identification and analysis. Respectively counting lipid signals in a homogenized rat brain tissue extracted by using the metal probe 6 sampling method and a traditional lipid extraction method, wherein a contrast graph of the rat brain signals is shown in figure 10, wherein PC is phosphatidylcholine; PE is phosphatidylethanolamine; PS is phosphatidylserine; PG is phosphatidyl glycerol; PI is phosphatidylinositol; TAG is triglyceride; from fig. 10, it can be seen that the lipid signal ratio of the two methods is around 1, the lipid species and abundance extracted by the two methods are not obviously different, and the stability is good, thus proving that the method has universal applicability.

Fatty acid FA 18:1 in rat brain tissue is selected respectively for identifying carbon-carbon double bond isomers, lipid PC 34:1 is selected for identifying sn isomers and carbon-carbon double bond isomers, and identification spectrograms are shown in figures 11 and 12 respectively. In the reaction module 3, the lipid and the fatty acid are subjected to photochemical derivatization reaction, and the position information of the carbon-carbon double bond of the lipid is obtained through tandem mass spectrometry, so that the lipid is subjected to fine structure analysis, and as can be seen from fig. 11 and 12, FA 18:1 comprises three carbon-carbon double bond isomers of delta 8(m/z 246.2, m/z 248.2), delta 9(m/z 232.2, m/z 262.2) and delta 11(m/z 204.2 and m/z 290.2); PC 34:1 has two sn isomers (m/z 380.4, m/z 396.1 and m/z 466.2) and two carbon-carbon double bond isomers (m/z 489.2, m/z 517.2, m/z 578.5 and m/z 606.5), wherein m/z is a mass-to-charge ratio, and thus, the method can prove that the method can carry out fine structure analysis on the lipid.

In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.

Furthermore, the terms "first", "second" and "first" are used for descriptive purposes only and are not to be construed as indicating or implying relative importance or implicitly indicating the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one such feature. In the description of the present invention, "a plurality" means at least two, e.g., two, three, etc., unless specifically limited otherwise.

While embodiments of the invention have been shown and described, it will be understood by those of ordinary skill in the art that: various changes, modifications, substitutions and alterations can be made to the embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the claims and their equivalents.