A system for in-situ preparation of polymer-coated capillaries, a probe and uses thereof

Abstract

The application discloses a system for preparing a polymer-coated capillary in situ by rotation, a probe and application thereof, and belongs to the technical field of rapid sample extraction analysis. The system comprises: a surface activation treatment module for forming a pre-modification layer on the inner surface of a capillary base through a bifunctional grafting reagent; a local etching module for performing patterned treatment on the pre-modification layer through electric heating or a laser mask technology; a rotary polymerization module for driving the capillary to rotate around an axis and exciting in-situ polymerization of a monomer solution; and a control module for controlling polymerization conditions to generate a polymer in-situ growth layer with adjustable porosity. The probe is composed of the capillary base and the polymer in-situ growth layer, the inner surface of the probe has axial uniformity and multi-segment bit patterning characteristics, and the probe is suitable for in-situ extraction and detection of small-volume biological samples such as whole blood, plasma and urine. The application realizes parameterized control of the coating thickness and porosity by rotation, is compatible with various chemical derivatization methods, and significantly improves the detection sensitivity.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of solid phase extraction analysis technology, and in particular to a system for preparing a rotating in-situ polymer-coated capillary, and a method for rapid extraction, in-situ ionization and photoderivatization reaction using the prepared probe. BACKGROUND

[0002] When analyzing organic components in complex samples such as biological tissues, environmental samples, etc., due to the complexity of the sample composition, sufficient pretreatment of the obtained sample is an essential step; liquid-liquid extraction is first performed on the sample, then liquid chromatography is used for separation, and finally mass spectrometry analysis has become the industry standard. The current commonly used liquid-liquid extraction method has a complex operation process, takes a long time, and introduces chloroform and other reagents that are highly polluting and toxic. There is also a non-negligible loss of sample during repeated transfer. Liquid chromatography also has the disadvantages of high running cost and long separation time. Rapid sampling extraction and in-situ ionization technology has significant advantages in this field. Such technology not only saves time and human resources, improves the convenience and timeliness of detection, but also reduces the dependence on a large amount of organic solvent, thereby reducing the environmental pollution burden.

[0003] In view of the above problems, researchers have developed many solid phase extraction-based methods to speed up the sample extraction and purification separation steps. Such methods mainly use functional particles adhered to the outside of a solid rod, and only the rapid pretreatment of the sample is simplified and improved. The sample still needs to be transferred after extraction before ionization and mass spectrometry analysis can be achieved, and a complete analysis method independent of liquid chromatography-mass spectrometry technology has not been achieved. The method of adhering extraction materials to the outside of a solid rod requires a sample volume that is much larger than the actual volume that is effectively extracted, resulting in waste of valuable samples, and has great limitations when faced with micro-sample analysis scenarios.

[0004] The extraction method in the capillary can significantly save the sample volume, and macroporous materials such as porous polymers have high specific surface area, providing sufficient adsorption sites for analyte adsorption, which can improve the extraction rate and material recovery rate in the limited space of the capillary. The capillary has a mature interface device for mass spectrometry analysis, which can directly complete electrospray ionization in-situ to achieve rapid analysis. Porous polymers can be adjusted in framework and pore size according to the structure and properties of the target analyte to enhance their adsorption selectivity, and can be functionally modified to improve the interaction between the target analyte and the porous polymer, eliminate the interference of complex sample matrix, and improve the selectivity of analysis.

[0005] However, due to the extremely limited space of the inner wall of the capillary, it cannot directly contact the inner wall surface, so it is impossible to transfer the process of making a solid-phase microextraction fiber to the scene of in-tube modification. The free radical reaction used for modifying the polymer has a fast reaction speed, and it is difficult to accurately control the initiation and termination of the reaction, and the polymerization in the capillary is easy to block the hollow structure. Therefore, the process technology of modifying a high-quality polymer thin layer on a small-diameter capillary still needs to be further developed.

[0006] Therefore, in view of the rapid analysis requirements of specific substances in biological samples, it is necessary to develop a device and application technology for rapid detection of biological fluid samples highly coupled with mass spectrometry to meet the requirements of micro-sample extraction and in-situ rapid ionization. SUMMARY

[0007] The present application aims to at least solve one of the technical problems existing in the prior art. To this end, one object of the present application is to propose a system for preparing a polymer-coated capillary in situ by rotation, and a method for rapid extraction, in-situ ionization and photo-derivation reaction using a probe prepared thereby.

[0008] The first aspect of the present application proposes a system for preparing a polymer-coated capillary in situ by rotation, for preparing a polymer-coated capillary probe, wherein the polymer in-situ growth layer in the polymer-coated capillary probe is porous, has a uniform and stable thickness, and does not change the hollow structure of the capillary base.

[0009] The system for preparing a polymer-coated capillary in situ by rotation according to the first aspect of the present application comprises:

[0010] A surface activation treatment module is configured to treat the inner surface of the capillary base with a surface activation solution to form a pre-modification layer.

[0011] A local etching module is configured to perform a patterning treatment on the pre-modification layer, wherein the patterning treatment is performed by electric heating decomposition or laser mask irradiation.

[0012] A rotating polymerization module comprises a motor, a rotating shaft, a clamping device and a reaction initiation device, wherein the clamping device clamps the capillary base and makes the capillary base coaxial with the rotating shaft, and the motor drives the rotating and initiates the in-situ polymerization of the monomer solution in the capillary base to generate a polymer in-situ growth layer.

[0013] A control module is electrically connected with the motor and the reaction initiation device, and controls the porosity and thickness of the polymer in-situ growth layer by adjusting the rotating speed and reaction time.

[0014] In summary, the rotating in-situ polymerization coating capillary system of the first aspect of the present application can prepare a polymer coating capillary probe, the polymer in-situ growth layer on the inner wall of the capillary substrate is uniform and stable in thickness, and does not change the hollow structure of the capillary substrate, so that the polymer coating capillary probe has an axial channel coaxial with the capillary substrate; the polymer in-situ growth layer has the characteristics of porosity and large specific surface area.

[0015] In some embodiments, the surface activation treatment module uses a surface activation solution to sufficiently rinse the inner surface of the capillary substrate, the surface activation solution comprising a bifunctional grafting agent, the first functional group of the bifunctional grafting agent being covalently bonded to the surface of the capillary substrate, and the second functional group of the bifunctional grafting agent being covalently bonded to the polymer monomer, so that the inner surface of the capillary substrate obtains a grafting active group.

[0016] In some embodiments, the local etching module includes an electric heating element arranged in a desired shape on the outer wall of the capillary substrate or a light mask device arranged on the outer wall of the capillary substrate. The local grafting active group on the inner surface of the capillary substrate is removed by pyrolysis; or a light mask is arranged on the outer wall of the capillary, and the active group in the unshielded area is decomposed by laser irradiation.

[0017] In some embodiments, the plurality of reactants in the monomer solution includes a free radical initiator, a polymerization monomer, and an extraction modifier. The free radical initiator, the polymerization monomer, and the extraction modifier can undergo a polymerization reaction to generate a polymer. Among them, the free radical initiator includes but is not limited to a free radical initiator excited by ultraviolet light and a free radical initiator excited by heating. The extraction modifier includes but is not limited to a hydrophilic group modifier, a hydrophobic group modifier, and a specific antibody. By changing the extraction modifier, the polymer in-situ growth layer can be modified to achieve high selective extraction.

[0018] In some embodiments, the rotating polymerization module injects a monomer solution into the inner surface of the capillary substrate, contacts and polymerizes the reactants by rotation, and forms the polymer in-situ growth layer after washing and drying.

[0019] In some embodiments, the rotating polymerization module comprises a motor, a rotating shaft, a clamping device and a reaction initiation device, the rotating motor is connected to the rotating shaft, the clamping device is arranged on the rotating shaft, the clamping device clamps the capillary substrate and makes the capillary substrate coaxial with the rotating shaft, and the reaction initiation device is arranged near one side of the capillary substrate. A proper amount of monomer solution is injected into the inner surface of the capillary substrate, the monomer solution is fully contacted with the grafted active groups on the inner surface of the capillary substrate, then the capillary substrate is rotated around its own axis and the polymerization reaction of the plurality of reactants in the monomer solution is initiated by the reaction initiation device, so that the generated polymer is directly grown and fixed on the patterned pre-modified layer on the inner surface of the capillary substrate, thereby obtaining the patterned polymer in-situ growth layer.

[0020] In some embodiments, the control module and the reaction initiation module are electrically connected to control the operation of the motor and the reaction initiation module. The porosity of the polymer in-situ growth layer is controlled by adjusting the rotating speed of the motor by the control module; and the thickness of the polymer in-situ growth layer is controlled by adjusting the initiation time of the polymerization reaction by the control module.

[0021] The second aspect of the present application provides a rotating in-situ prepared polymer coated capillary probe.

[0022] The system for rotating in-situ preparation of polymer coated capillary according to the first aspect of the present application can obtain the polymer coated capillary probe according to the second aspect of the present application, the thickness and position of the polymer in-situ growth layer of the probe are controllable, the polymer in-situ growth layer is firmly modified, and no shedding phenomenon occurs in use.

[0023] The preparation method of the polymer coated capillary probe according to the second aspect of the present application comprises the following steps:

[0024] S1: treating the inner surface of the capillary substrate by the surface activation treatment module to form a pre-modified layer;

[0025] S2: performing local etching treatment on the pre-modified layer on the inner surface of the capillary substrate to form a patterned pre-modified layer on the remaining active groups on the inner surface of the capillary substrate;

[0026] S3: injecting a monomer solution into the capillary substrate, clamping the capillary on the rotating polymerization module, driving the capillary to rotate around its own axis, initiating a controllable in-situ polymerization reaction, and growing a patterned polymer in-situ growth layer at the patterned pre-modified layer on the inner surface of the capillary substrate.

[0027] The polymer-coated capillary probe of the second aspect of the present application has a polymer in-situ growth layer coaxially in-situ grown on the inner surface of the capillary substrate, does not block the central passage of the capillary substrate, has a central passage coaxial with the capillary substrate, and can suck in a trace amount of solution, which can flow freely in the polymer-coated capillary probe.

[0028] By sucking in a trace amount of the solution to be measured, the polymer in-situ growth layer in the polymer-coated capillary probe of the second aspect of the present application can adsorb or enrich the target analyte in the solution to be measured. By sucking in a trace amount of the washing solution, the polymer in-situ growth layer in the polymer-coated capillary probe of the second aspect of the present application can purify the target analyte on the polymer in-situ growth layer and remove impurities in the target analyte. By sucking in a trace amount of the eluent, the polymer in-situ growth layer in the polymer-coated capillary probe of the second aspect of the present application can elute or release the target analyte on the polymer in-situ growth layer into the eluent in the polymer-coated capillary probe. By applying high pressure to the polymer-coated capillary probe, the eluent containing the target analyte in the polymer-coated capillary probe of the second aspect of the present application can be in-situ ionized and form an electrospray at the tip.

[0029] When the eluent contains a photo-derivatization reagent, the polymer-coated capillary probe of the second aspect of the present application can also simultaneously perform in-situ ionization and photo-derivatization, thereby realizing the coupling with mass spectrometry, being compatible with various chemical derivatization analysis methods, and being suitable for high-throughput mass spectrometry analysis and in-situ on-site detection.

[0030] The polymer-coated capillary probe of the second aspect of the present application has a simple operation process, is convenient and fast, has good extraction effect, and has high detection sensitivity.

[0031] In some embodiments, the thickness of the polymer in-situ growth layer is uniform in different directions of any one transverse section perpendicular to the axis of the capillary substrate; and the thickness of the polymer in-situ growth layer is uniform on any multiple transverse sections with equal inner diameters perpendicular to the axis of the capillary substrate.

[0032] In some embodiments, the polymer in-situ growth layer is directly grown on the inner surface of the capillary substrate by in-situ reaction of a monomer solution, and the polymer in-situ growth layer has a porous structure with adjustable porosity.

[0033] In some embodiments, the polymer in-situ grown layer is a continuous polymer in-situ grown layer along the inner surface of the capillary substrate or a patterned polymer in-situ grown layer discontinuously distributed along the inner surface of the capillary substrate or a plurality of different modified polymer in-situ grown layers patterned at different sites of the inner surface of the capillary substrate.

[0034] In some embodiments, the material of the polymer in-situ grown layer includes but is not limited to organic polymer, silica microsphere cross-linked polymer, graphene, metal / covalent organic framework; and the modification method of the polymer in-situ grown layer includes but is not limited to hydrophilic group modification, hydrophobic group modification, and addition of specific antibody or aptamer.

[0035] The third aspect of the present application provides an application of the polymer-coated capillary probe, which is used for in-situ extraction detection of target objects in whole blood, plasma or urine samples, and can be used in combination with chemical derivatization method.

[0036] Additional aspects and advantages of the present application will be partially given in the following description, partially will become obvious from the following description, or will be understood by the practice of the present application. BRIEF DESCRIPTION OF DRAWINGS

[0037] Figure 1 Structure schematic diagram of a system for preparing a polymer-coated capillary by rotation in-situ according to the present application;

[0038] Figure 2 Structure schematic diagram of a local etching module of the system according to the present application;

[0039] Figure 3 Structure schematic diagram of a rotation polymerization module of the system according to the present application;

[0040] Figure 4 Working process schematic diagram of the rotation polymerization module of the system according to the present application;

[0041] Figure 5 Cross-sectional scanning electron microscope photo of the polymer-coated capillary prepared by the system according to the present application;

[0042] Figure 6a Curve of the relationship between the thickness of the polymer in-situ grown layer and the polymerization reaction time of the polymer-coated capillary prepared by the system according to the present application;

[0043] Figure 6b Scanning electron microscope photos of the thickness of the polymer in-situ grown layer of the polymer-coated capillary prepared by the system according to the present application at different polymerization reaction time lengths;

[0044] Figure 7Structure diagram of polymer coated capillary probe prepared for the system of the present application;

[0045] Figure 8 Physical photograph and micro-imaging diagram of polymer coated capillary probe and capillary substrate of the present application;

[0046] Figure 9 Extraction effect diagram of polymer coated capillary probe of the present application and comparison diagram of number of lipid species extracted by liquid-liquid extraction method;

[0047] Figure 10a Mass spectrum diagram of positive ion mode of plasma sample extracted by polymer coated capillary probe of the present application;

[0048] Figure 10b Mass spectrum diagram of negative ion mode of plasma sample extracted by polymer coated capillary probe of the present application;

[0049] Figure 11a Extraction effect diagram of phosphatidylcholine by polymer coated capillary probe of the present application;

[0050] Figure 11b Extraction effect diagram of phosphatidylethanolamine by polymer coated capillary probe of the present application;

[0051] Figure 11c Extraction effect diagram of triglyceride by polymer coated capillary probe of the present application;

[0052] Figure 12 Analysis result diagram of unsaturated lipid structure identification of human plasma sample by polymer coated capillary probe of the present application.

[0053] Reference signs:

[0054] System 1000 for preparing polymer coated capillary in situ by rotation; surface activation treatment module 1; local etching module 2; electric heating element 201; photo mask device 202; rotation polymerization module 3; motor 301, rotation shaft 302, clamping device 303; reaction excitation device 304; control module 4; polymer coated capillary probe 2000; main tube segment 501; closed tube segment 502; needle tip tube segment 503; capillary substrate 5; main tube segment 501, closed tube segment 502; needle tip tube segment 503; pre-modification layer 6; patterned pre-modification layer 6'; polymer in-situ growth layer 7. DETAILED DESCRIPTION

[0055] Embodiments of the present application are described in detail below, examples of which are shown in the accompanying drawings, wherein the same or similar reference signs represent the same or similar elements or elements having the same or similar functions throughout. The embodiments described below by reference to the drawings are exemplary and are only used to explain the present application, and cannot be understood as a limitation of the present application.

[0056] The following is combined with Figures 1 to 12 The present invention describes a system 1000 for preparing polymer-coated capillaries by rotation in situ, a method for preparing polymer-coated capillary probes 2000 using the present system, and a method for extraction and photoderivation reaction within the polymer-coated capillary probes 2000.

[0057] like Figure 1 As shown, the first aspect of the present invention provides a system 1000 for the in-situ preparation of polymer-coated capillaries by rotation.

[0058] A system 1000 for preparing polymer-coated capillaries by rotation in situ according to a first aspect of the present invention includes the following components: a surface activation treatment module 1, a local etching module 2, a rotation polymerization module 3, and a control module 4. The surface activation module 1 is used to treat the inner surface of the capillary substrate with a surface activation solution to form a pre-modified layer; the local etching module 2 is configured to pattern the pre-modified layer by electrothermal decomposition or laser mask irradiation; the rotary polymerization module 3 includes a motor 301, a rotary shaft 302, a clamping device 303 and a reaction excitation device 304. The clamping device clamps the capillary substrate 5 and makes the capillary substrate 5 coaxial with the rotary shaft 302. The motor 301 drives the rotation and excites the monomer solution to polymerize in situ, generating a polymer in situ growth layer 7; the control module 4 is electrically connected to the motor 301 and the reaction excitation device 304, and controls the porosity and thickness of the coating (i.e., the polymer in situ growth layer 7) by adjusting the rotation speed (2000~12000rpm) and the reaction time (30~500s).

[0059] In some embodiments, the surface activation solution used in the surface activation treatment module 1 contains a bifunctional grafting reagent. The first functional group of the bifunctional grafting reagent is covalently bonded to the surface of the capillary substrate 5, and the second functional group of the bifunctional grafting reagent is covalently bonded to the polymer monomer. This facilitates a firm connection between the pre-modified layer 6 and the capillary substrate 5 and the polymer in-situ growth layer 7.

[0060] In some embodiments, the local etch module 2 includes an electrically heated element 201 of a desired shape disposed on the outer wall of the capillary substrate 5, or a photomask device 202 disposed on the outer wall of the capillary substrate 5. Specifically, such as Figure 2 As shown, the local etch-removal module 2 specifically involves an electric heating element 201 heating a local area of ​​the capillary substrate 5, removing locally grafted active groups on the inner surface of the capillary substrate 5 through pyrolysis, thereby obtaining a patterned pre-modified layer 6'; or a photomask device 202 is set on the outer wall of the capillary substrate 5, and a laser is used to irradiate and decompose the active groups in the unmasked area, thereby obtaining a patterned pre-modified layer 6'. Subsequently, a corresponding patterned polymer in-situ growth layer 7 can be modified within the capillary substrate 5.

[0061] In some embodiments, the monomer solution comprises a radical initiator, a polymerizable monomer, and an extraction modifier. The radical initiator includes, but is not limited to, a radical initiator excited by ultraviolet light and a radical initiator excited by heating. The extraction modifier includes, but is not limited to, a hydrophilic group modifier, a hydrophobic group modifier, and a specific antibody. By changing the extraction modifier, the polymer in-situ growth layer 7 can be modified to achieve high selective extraction.

[0062] In some embodiments, the rotating polymerization module 3 injects the monomer solution into the inner surface of the capillary substrate 5, and the reactants are contacted and polymerized by rotation, and the polymer in-situ growth layer 7 is formed after washing and drying. Specifically, as shown in Figure 3 The rotating polymerization module 3 comprises a motor 301, a rotating shaft 302, a clamping device 303, and a reaction excitation device 304. The control module 4 is connected to the motor 301 and the reaction excitation device 304. The motor 301 is connected to the rotating shaft 302 for driving the rotating shaft 302 to rotate around its own axis. The clamping device 303 is arranged on the rotating shaft 302 and clamps the capillary substrate 5 so that the capillary substrate 5 is coaxial with the rotating shaft 302. In this way, the capillary substrate 5 rotates synchronously around its own axis with the rotating shaft 302. The reaction excitation device 304 is arranged near one side of the capillary substrate 5 to excite the polymerization reaction of the various reactants in the monomer solution in the capillary substrate 5. The generated polymer particles are settled on the inner surface of the capillary substrate 5 under the action of centrifugal force and further polymerize and consolidate on the inner surface of the capillary substrate 5. The coating adheres to the capillary substrate 5 with sufficient strength while maintaining the porous morphology. The polymer-coated capillary probe 2000 has an axial channel, which can normally absorb and release solution, so that a polymer-coated capillary probe 2000 with a smaller spray opening can be prepared for the detection of a smaller amount of sample.

[0063] The control module 4 is electrically connected to the motor 301 and the reaction excitation device 304 to control the operation of the motor 301 and the reaction excitation device 304. By adjusting the rotating speed of the motor 301, the size of the centrifugal force can be controlled, and further the porosity of the polymer in-situ growth layer 7 can be controlled. By adjusting the polymerization reaction time, the thickness of the polymer in-situ growth layer 7 can be controlled. Using this device to prepare a thin-layer polymer material micro-extraction probe (polymer-coated capillary probe 2000), a probe with a highly uniform polymer thickness and a controllable modification thickness can be obtained without changing the hollow nature of the polymer-coated capillary probe 2000, ensuring that the polymer-coated capillary probe 2000 has an axial channel.

[0064] In some embodiments, the reaction excitation device 304 includes, but is not limited to, an ultraviolet light source module and a heating source module, which can be selected according to actual needs. As shown in Figure 3 andFigure 4 As shown, a specific example of a method for preparing a polymer-coated capillary probe is given. After the inner surface of the capillary substrate 5 is fully cleaned and activated surface groups, a pre-modification layer 6 of patterned grafting active groups is prepared using a grafting reagent. The inner surface of the capillary substrate 5 is filled with a monomer solution containing polymerization monomers, free radical initiators and other components, and the monomer solution is brought into full contact with the surface to be modified. The capillary substrate 5 is fastened on the clamping device 303 with its axis collinear with the rotation axis 302, and rotation is started and polymerization is initiated. Under the condition of rotation around the axis, the polymerization generates polymer deposition and interconnection on the inner surface of the capillary substrate 5 under the action of centrifugal force, forming a uniform polymer in-situ growth layer 7. Due to the symmetry of rotation, the thickness of the polymer grown on each part of the capillary substrate 5 is uniform and stable, and the polymer is only distributed on the inner surface without changing the hollow structure of the capillary substrate 5, so that the polymer-coated capillary probe 2000 has an axial central channel coaxial with the capillary substrate 5.

[0065] Figure 5 (a) in FIG. 7 is a scanning electron microscope image of the polymer in-situ growth layer 7, which can be seen to have the characteristics of porosity and large specific surface area; Figure 5 (b) and (c) in FIG. 7 are cross-sectional scanning electron microscope images of the capillary substrate 5, which can be seen to have a uniform polymer in-situ growth layer 7 attached to the inner wall of the capillary substrate 5, and at the tip of one end of the polymer-coated capillary probe 1000, there is still a layer of uniform growth on the inner surface of the capillary substrate 5, indicating that the tip of the capillary substrate 5 also has extraction function. By adjusting the length of time of the polymerization reaction, the thickness of the polymer in-situ growth layer 7 can be controlled, as shown in Figure 6a and Figure 6b

[0066] As shown in FIG. 6, the porosity of the polymer in-situ growth layer 7 can be controlled by adjusting the motor speed through the control module 4, and the thickness of the polymer in-situ growth layer 7 can be controlled by adjusting the initiation time of the polymerization reaction through the control module 4, and the initiation time of the polymerization reaction is 30-500 s. In this way, the polymer-coated capillary probe 2000 can be quickly prepared, and the thickness of the polymer-coated capillary probe 2000 is highly related to the polymerization reaction time.

[0067] In summary, the rotating in-situ preparation of the polymer-coated capillary system 1000 of the first aspect of the present application can prepare the polymer-coated capillary probe 2000, the thickness of the polymer in-situ growth layer 7 on the inner wall of the capillary substrate 5 is uniform and stable, and the hollow structure of the capillary substrate 5 is not changed, so that the polymer-coated capillary probe 2000 has an axial central channel coaxial with the capillary substrate 5; the polymer in-situ growth layer 7 has the characteristics of porosity and large specific surface area. ​

[0068] The second aspect of the present application provides a polymer-coated capillary probe 2000 prepared by rotation in situ.

[0069] As shown in FIG. 2, the polymer-coated capillary probe 2000 of the second aspect of the present application is prepared by the system 1000 of the first aspect of the present application. Figure 7

[0070] Figure 8 The photos and optical microscope images of the polymer-coated capillary probe 2000 prepared by in-situ controlled uniform growth of thin-layer polymer in the capillary substrate 5 show that the inner surface of the polymer-coated capillary probe 2000 after modification is covered with an opaque polymer in-situ growth layer 7. The appearance of the capillary substrate 5 is not damaged before and after modification, and the hollow characteristics of the capillary substrate 5 are also retained.

[0071] In some embodiments, the capillary substrate 5 in the capillary probe 2000 includes a main body tube segment 501, a tapered tube segment 502, and a needle tip tube segment 503 connected in sequence coaxially. The needle tip tube segment 503 can serve as a sampling tube segment, i.e., by inserting the needle tip tube segment 503 into a solution, so that the polymer-coated capillary probe 2000 can absorb the solution; the tapered tube segment 502 can serve as an ionization spray tube segment, i.e., by applying high pressure to the polymer-coated capillary probe 2000, so that the solution in the capillary substrate 5 can generate an electrospray at the tapered tip of the tapered tube segment 502. The material of the capillary substrate 5 includes but is not limited to glass, quartz, polypropylene, polyether ether ketone, other high molecular materials, etc.

[0072] ​The polymer in-situ growth layer 7 is tubular and coaxially in-situ grown on the inner surface of the needle tip tube segment 503, the inner surface of the closed tube segment 502 or / and the inner surface of the main tube segment 501, so that the polymer coated capillary probe 2000 has an axial channel. That is, the polymer in-situ growth layer 7 is directly in-situ deposited and fixed on the inner surface of the capillary substrate 5 by polymerization of multiple reactants in the monomer solution in the capillary substrate 5, and the polymer in-situ growth layer 7 is tubular and coaxially fixed on the inner surface of the capillary substrate 5 without blocking the central channel of the capillary substrate 5, so that the polymer coated capillary probe 2000 has an axial channel coaxial with the capillary substrate 5, ensuring that the polymer coated capillary probe 2000 can suck in the solution, and the solution can freely flow in the axial channel of the polymer coated capillary probe 2000 without obstruction; the position of the polymer in-situ growth layer 7 in-situ grown in the capillary substrate 5 is controllable, and it can be fixed on the inner surface of the needle tip tube segment 503, the inner surface of the closed tube segment 502 or / and the inner surface of the main tube segment 501 according to actual needs. The polymer in-situ growth layer 7 has a high specific surface area and can quickly adsorb or rapidly enrich target analytes in the solution to be measured.

[0073] In some embodiments, the preparation method of the polymer coated capillary probe 2000 includes the following steps, as shown in Figure 4

[0074] S1: treating the inner surface of the capillary substrate 5 with a surface activation solution to form a pre-modification layer 6.

[0075] S2: performing local etching treatment on the pre-modification layer 6 on the inner surface of the capillary substrate 5, so that the remaining active groups on the inner surface of the capillary substrate 5 form a patterned pre-modification layer 6'.

[0076] ​S3: injecting the monomer solution into the capillary substrate 5, clamping the capillary substrate 5 on the rotating polymerization module 3 to drive the capillary substrate 5 to rotate around its own axis, and growing the patterned polymer in-situ growth layer 7 in-situ at the patterned pre-modification layer 6' on the inner surface of the capillary substrate 5 through controllable in-situ polymerization. That is, the polymer in-situ growth layer 7 is directly deposited and fixed in-situ on the inner surface of the capillary substrate 5 by the polymer formed by the polymerization of the various reactants in the monomer solution, and the polymer in-situ growth layer 7 is coaxially fixed on the inner surface of the capillary substrate 5 in a tubular shape without blocking the central passage of the capillary substrate 5, so that the polymer-coated capillary probe 2000 has a central passage coaxial with the capillary substrate 5, ensuring that the polymer-coated capillary probe 2000 can suck in the solution, and the solution can flow freely in the central passage of the polymer-coated capillary probe 2000 without obstruction. The position of the polymer in-situ growth layer 7 grown in-situ in the capillary substrate 5 is controllable, and the polymer in-situ growth layer 7 can be fixed on the inner surface of the converging tube section 502, on the inner surface of the main tube section 501, or on the inner surfaces of both the converging tube section 502 and the main tube section 501 according to actual needs. The polymer in-situ growth layer 7 has a high specific surface area and can quickly adsorb or enrich the target analyte in the solution to be measured.

[0077] Through step S3, the polymer in-situ growth layer 7 with extremely uniform thickness along the inner surface of the capillary substrate 5 can be prepared, and the polymer in-situ growth layer 7 can be modified without causing blockage at the spray end opening even if the spray end opening is extremely small. The polymer particles generated in the monomer solution are subjected to centrifugal action and settle on the inner surface of the capillary substrate 5, and are further polymerized and fixed on the inner surface of the capillary substrate 5, ensuring the adhesion strength of the polymer in-situ growth layer 7 and ensuring that the polymer-coated capillary probe 2000 can normally suck and release the solution, so that a probe with a smaller spray opening can be prepared for the detection of a smaller amount of sample. Because most of the generated polymer particles eventually settle on the inner surface of the capillary substrate 5 and are fixed, the thickness of the polymer in-situ growth layer 7 is highly related to the polymerization time, and the thickness of the polymer in-situ growth layer 7 inside the capillary substrate 5 can be controlled by controlling the polymerization time. The polymer-coated capillary probes 2000 prepared in different batches have stable quality, and the polymer-coated capillary probes 2000 can be easily prepared in large quantities for high-throughput analysis.

[0078] In some embodiments, the thickness of the polymer in-situ growth layer 7 is uniform in different directions of any one of the transverse sections perpendicular to the axis of the capillary base 5; and the thickness of the polymer in-situ growth layer 7 is uniform in any one of the transverse sections with equal inner diameters perpendicular to the axis of the capillary base 5. This is conducive to not changing the central channel characteristics of the capillary base 5, and makes the polymer-coated capillary probe 2000 have an axial central channel coaxial with the capillary base 5.

[0079] In some embodiments, the material of the polymer in-situ growth layer 7 includes but is not limited to organic polymers, silica microsphere cross-linked polymers, graphene, metal / covalent organic frameworks; and the modification method of the polymer in-situ growth layer 7 includes hydrophilic / hydrophobic group modification or antibody / aptamer addition, which can achieve high selective extraction.

[0080] In some embodiments, as shown in FIG. 6, the polymer in-situ growth layer 7 is a porous structure with adjustable porosity, which is directly generated by in-situ polymerization reaction. That is, the porosity of the polymer in-situ growth layer 7 is adjustable during the in-situ growth of the polymer in-situ growth layer 7, so as to realize the enrichment of the polymer in-situ growth layer 7 to the target analyte.

[0081] In some embodiments, the polymer in-situ growth layer 7 is in a continuous distribution, a discontinuous patterned distribution, or a multi-modification section distribution, which can be directly generated by in-situ polymerization reaction according to actual needs. In some embodiments, obviously, the polymer in-situ growth layer 7 can be arranged on the inner surface of the capillary base 5 with very small inner diameter without changing the hollow characteristics of the capillary base 5, and the polymer-coated capillary probe 2000 has an axial central channel for liquid flow, which is conducive to saving the sample to be tested and realizing rapid enrichment, and has high detection sensitivity.

[0082] The third aspect of the embodiments of the present application provides an application of a polymer-coated capillary probe. The polymer-coated capillary probe 2000 is used for in-situ extraction detection of target objects in whole blood, plasma or urine samples, and the in-situ extraction detection can be combined with a chemical derivatization method.

[0083] For example, the extraction method in the polymer-coated capillary probe of the embodiments of the present application is used for detecting a plasma sample. Figure 9 、 Figures 10a-10b 、 Figures 11a-11cThe statistical comparison chart of the identified lipid species in the extract of the polymer coating capillary probe 2000 according to the third aspect of the present application and the traditional liquid-liquid extraction, the mass spectrum chart collected in positive and negative ion modes, and the comparison chart of the mass spectrum of the detected phosphatidylcholine, phosphatidylethanolamine and triglyceride species are shown respectively. It can be seen that the results obtained by the rapid extraction method according to the third aspect of the present application are basically consistent with the results obtained by the current industry gold standard, most of the lipids can be extracted, and the time is greatly shortened.

[0084] The extraction and photoderivatization reaction method of the polymer coating capillary probe according to the embodiments of the present application are compatible, the same method is used to elute by preparing an elution solution containing photoderivatization reaction reagents. When the electrospray is ionized in situ, a light source with a characteristic wavelength is placed on the side of the polymer coating capillary probe 2000 where the electrospray is generated, so that the photoderivatization reaction can be carried out online during the electrospray, and a more accurate judgment can be made on the molecular structure of the analyte. Figure 12 The results of identifying the position of the carbon-carbon double bond of unsaturated phosphatidylcholine and phosphatidylethanolamine in human plasma by adding acetone as a photoderivatization reagent in the elution solvent are shown. The position information of the carbon-carbon double bond of the lipid is obtained by analyzing the lipid by tandem mass spectrometry, so as to analyze the fine structure of the lipid. It can be seen that the phosphatidylcholine PC 34:1 has two double bond position isomers of Δ9 and Δ11, reflecting that the method can simultaneously perform in situ photoderivatization reaction for fine structure analysis.

[0085] In the description of the present specification, the description of the terms "one embodiment", "some embodiments", "exemplary embodiment", "example", "specific example", or "some examples" means that the specific features, structures, or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present application. In the present specification, the exemplary description of the above terms does not necessarily mean the same embodiment or example. Moreover, the specific features, structures, or characteristics described can be combined in any one or more embodiments or examples in a suitable manner.

[0086] Although the embodiments of the present application have been shown and described, those skilled in the art can understand that various changes, modifications, replacements and variations can be made to the embodiments without departing from the principles and purposes of the present application, and the scope of the present application is defined by the claims and their equivalents.

Claims

1. A method of fabricating a polymer-coated capillary probe using a system for in-situ fabrication of polymer-coated capillaries by rotation, characterized in that, The system for preparing a polymer-coated capillary in situ by rotation comprises: a surface activation treatment module for treating the inner surface of a capillary substrate with a surface activation solution to form a pre-modification layer; a local etching module configured to perform a patterned treatment on the pre-modification layer by means of electric heating decomposition or laser mask irradiation; the local etching module comprises an electric heating element of a desired shape arranged on the outer wall of the capillary substrate or a light mask device arranged on the outer wall of the capillary substrate; a rotation polymerization module comprising a motor, a rotating shaft, a clamping device, and a reaction initiation device; the clamping device clamps the capillary substrate and makes the capillary substrate coaxial with the rotating shaft; the motor drives rotation and initiates in-situ polymerization of a monomer solution in the capillary substrate to generate a polymer in-situ growth layer; a control module electrically connected to the motor and the reaction initiation device to control the porosity and thickness of the polymer in-situ growth layer by adjusting the rotation speed and reaction time; the method for preparing a polymer-coated capillary probe comprises the following steps: S1: treating the inner surface of a capillary substrate with a surface activation solution by the surface activation treatment module to form a pre-modification layer; S2: performing a local etching treatment on the pre-modification layer on the inner surface of the capillary substrate to form a patterned pre-modification layer with active groups remaining on the inner surface of the capillary substrate; S3: injecting a monomer solution into the capillary substrate, clamping the capillary substrate on the rotation polymerization module, driving the capillary substrate to rotate around its own axis, and initiating a controllable in-situ polymerization reaction to grow a patterned polymer in-situ growth layer at the patterned pre-modification layer on the inner surface of the capillary substrate; the polymer in-situ growth layer is discontinuously patterned or distributed in multiple modified segments.

2. A method of fabricating a polymer-coated capillary probe using the system for fabricating polymer-coated capillary tubes in situ by rotation according to claim 1, characterized by, The surface activation solution used by the surface activation treatment module comprises a bifunctional grafting agent, the first functional group of which covalently binds to the surface of the capillary substrate, and the second functional group of which covalently binds to a polymer monomer.

3. The method of claim 1, wherein the polymer-coated capillary probe is prepared using the system for preparing polymer-coated capillary tubes in rotation in situ, characterized in that, The monomer solution comprises a free radical initiator, a polymerization monomer, and an extraction modifier.

4. The method of claim 1, wherein the polymer-coated capillary probe is prepared using the system for preparing polymer-coated capillary in rotation in situ, characterized in that, The rotation polymerization module injects a monomer solution into the inner surface of the capillary substrate, contacts and polymerizes the reactants by rotation, and forms the polymer in-situ growth layer after washing and drying.

5. The method of claim 1, wherein the polymer-coated capillary probe is prepared using the system for preparing polymer-coated capillary in rotation in situ, characterized in that, The polymer in-situ growth layer is a porous structure with adjustable porosity, directly generated by in-situ polymerization.

6. The method of claim 1, wherein the polymer-coated capillary probe is prepared using the system for preparing polymer-coated capillary in rotation in situ, and The material of the polymer in-situ growth layer includes organic polymers, silica microsphere cross-linked polymers, graphene, and metal / covalent organic frameworks; the modification method of the polymer in-situ growth layer includes hydrophilic / hydrophobic group modification or antibody / aptamer addition.

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