Method for preparing gun tip type solid phase extraction column and gun tip type solid phase extraction column filling device

Abstract

The present invention discloses a method for preparing a gun-tip solid phase extraction column and a gun-tip solid phase extraction column filling device, the method comprising: (1) mixing solid phase extraction filler and methanol to obtain a filler suspension, and oscillating with an oscillator; (2) using a pipetting device to absorb the filler suspension, and accurately pipetting it into a sieve plate pipetting gun tip; (3) centrifuging the sieve plate pipetting gun tip or a sample plate equipped with a plurality of sieve plate pipetting gun tips to volatilize the solvent methanol, and obtaining a gun-tip solid phase extraction column, collecting liquid methanol in the sample plate, and collecting volatilized gaseous methanol, condensing it and combining it with liquid methanol, condensing it for purification, and recycling it. The present invention utilizes a column filling device to uniformly suspend the filler in a methanol solution and activate it, pipetting the filler suspension into a sieve plate pipetting gun tip, centrifuging it for precipitation, removing the solvent, and realizing the production of high-throughput SPE tips, and the obtained gun-tip solid phase extraction column can be directly loaded with samples for SPE experiments without activation.

Description

Technical Field

[0001] The invention relates to a method for preparing a gun-tip type solid phase extraction column and a gun-tip type solid phase extraction column filling device. Background Art

[0002] Solid Phase Extraction (SPE) is a technique used to separate and purify target compounds in liquid samples. SPE has high separation and concentration capabilities, and can process complex matrix samples using a small amount of solvent carrier, significantly improving sample analysis efficiency, recovery rate and detection sensitivity. SPE can be used for environmental analysis (water quality testing, soil and sediment analysis), food safety testing (pesticide residues, food additives and pollutants), biomedical analysis (drugs, clinical samples), pharmaceutical industry research (drug purification, drug metabolism), forensic testing (poisons, trace substances), cosmetic ingredient testing (active ingredients, preservatives and pollutants), etc.

[0003] SPE mainly uses SPE devices for separation and extraction, and its components mainly include containers, solid phase adsorbents (fillers), eluents, sieve plates (filters) and connectors at various parts.

[0004] SPE containers mainly include small columns, tips, plates, tubes, etc., among which SPE small columns are suitable for most samples; SPE tips are used with automatic pipetting workstations to achieve automated operation and accurately control the volume of sample box solvents, and are often used in high-throughput screening; SPE plates belong to the multi-well plate format and can process multiple samples at the same time, and are also suitable for high-throughput screening and pharmaceutical research; SPE tubes are currently the largest containers in solid phase extraction, suitable for processing large-volume samples and for environmental analysis, etc.

[0005] The solid phase adsorbent used in SPE is the core component of SPE. Choosing the right filler is the key to effective extraction and separation. Currently, the commonly used fillers are C18 (octadecyl bonded silica gel), C8 (octyl bonded silica gel), HLB (lipophilic-hydrophilic polymer mixed material), silica gel, ion exchange resin, polymer adsorbent, etc.

[0006] SPE eluent is used to desorb the target substance in the filler and transfer it to the solution phase for subsequent analysis. It has the characteristics of good solubility, polarity matching with the target substance and adsorbent, strong selectivity, and compatibility with methods and instruments. Common eluents are divided into organic solvents (methanol, acetonitrile, ethanol, isopropanol, etc.), water-organic solvent mixtures (water-methanol, water-acetonitrile), acidic solutions (0.1% formic acid aqueous solution, 0.1% acetic acid aqueous solution), alkaline solutions (0.1% ammonium hydroxide aqueous solution) and salt solutions (phosphate buffer), etc. according to their properties.

[0007] The sieve plate is usually located at the top and bottom of the container or the upper and lower ends of the adsorbent to fix the adsorbent and filter the sample. The sieve plate is usually made of polyethylene (PE) or polypropylene (PP) with a pore size between 20 and 60 μm.

[0008] Currently, SPE devices are mainly produced manually or semi-automatically, which is prone to problems such as long filling time, large quality errors, and unevenness, and cannot meet the homogenization and higher-level SOP production requirements.

[0009] The DPX gun-tip dispersed solid phase microextraction column is a new type of solid phase extraction device. The solid phase extraction filler is loaded in the pipette tip and the pipette is used for blowing and suction operations. This can reduce the amount of filler used and is suitable for loading small volume samples. It effectively reduces the amount of solvent used and greatly improves the solid phase extraction rate. It is suitable for high-throughput multi-sample screening.

[0010] At present, the SPE devices on the market are mainly produced by individual components and then assembled. This method is highly flexible and a specific device combination can be selected according to the needs of subsequent experiments. However, in terms of operation, whether it is manual or semi-automatic, it is time-consuming and labor-intensive, with large errors, and cannot effectively achieve high throughput, and the cost is high. The specifications of commercially available gun-tip solid phase extraction columns are limited, and they need to be used with specific gun tips, which also leads to high costs, and commercially available SPE tips cannot meet the needs of different types or volumes of fillers for different tests. In addition, the production process and the use process of the SPE device are separated and have a lag. The filler of the commercially available SPE tips needs to be activated before it can be used. If an automated SPE device is used, an activation step needs to be added. If you choose to make your own SPE tips, due to the small volume of the filler, the error of self-filling is large, which affects the accuracy and consistency of the results.

[0011] It is necessary to develop an automated self-filling column device and column filling method that can meet the needs of high-throughput sample SPE processing, can perform precise equal-amount filling, and can simultaneously activate the filling stage to reduce subsequent detection steps. Summary of the invention

[0012] In order to meet the production requirements of SPE adsorption columns such as complicated processes and high precision, and to solve the problems of low manual production efficiency, uncontrollable quality, limited specifications of commercially available SPE tips, and the need for activation of fillers, the purpose of the present invention is to provide a low-cost, mechanized and large-scale new automated production method and column filling device that can meet the needs of high-throughput sample SPE processing, which can quickly, accurately and efficiently produce gun-tip solid phase extraction columns (SPE tips), and the activation of fillers is completed while producing SPE tips, so that they can be directly used in subsequent SPE experiments.

[0013] The technical solution adopted by the present invention is:

[0014] A method for preparing a gun-tip solid phase extraction column, the method comprising the following steps:

[0015] (1) mixing the solid phase extraction filler and methanol to obtain a filler suspension, placing the filler suspension into a filler mixing container, covering the container with a sealing cover, and oscillating the filler mixing container with an oscillator to keep the filler suspension uniformly suspended;

[0016] (2) placing a plurality of sieve plate pipette tips on the sample plate, wherein a filter sieve plate is provided inside the pointed outlet at the lower end of the sieve plate pipette tip; after opening the cover of the filler mixing container, using a pipetting device to absorb the filler suspension and accurately pipetting it into the sieve plate pipette tip on the sample plate;

[0017] (3) placing a sieve plate pipette tip or a sample plate equipped with multiple sieve plate pipette tips into a centrifuge; using the centrifuge to perform centrifugation to separate the filler suspension in the sieve plate pipette tip into solid and liquid to obtain a filler precipitate and a solvent; volatilizing the solvent methanol to prepare the tip-type solid phase extraction column, collecting liquid methanol in the sample plate, extracting and collecting the volatilized gaseous methanol, condensing it and combining it with the liquid methanol, purifying it and recycling it in step (1).

[0018] In the step (1), the volume amount of methanol is generally 3-500 μL / mg based on the mass of the filler.

[0019] The solid phase extraction filler can be a common SPE filler on the market or a self-made filler. Commonly used SPE fillers include C18 (octadecyl bonded silica gel), C8 (octyl bonded silica gel), HLB (lipophilic-hydrophilic polymer mixed material), silica gel, ion exchange resin, polymer adsorbent, etc. The present invention does not limit the type of filler.

[0020] In the step (1), the oscillation is generally vertical oscillation with a frequency of 100 to 500 times per minute, so that the filler is fully suspended in the methanol.

[0021] In the step (2), the filter sieve plate in the sieve plate pipette tip has holes or pores that allow liquid solvent to pass through, but cannot allow solid phase extraction filler particles to pass through.

[0022] The shape of the screen plate pipette tip is generally a conical tube, with an upper opening for connecting a pipette or a pipette interface and a pointed outlet at the lower end.

[0023] In the step (2), the single pipetting volume of the filler suspension can be 5 to 200 μL, or the same screening pipette tip can be pipetted multiple times, the number of times being 1 to 10 times.

[0024] In step (2), the sample plate is used to place a plurality of sieve plate pipette tips. The sample plate comprises a porous plate, and the porous plate is provided with a plurality of openings for placing a plurality of sieve plate pipette tips to collect the liquid solvent flowing out of the outlet at the lower end of the sieve plate pipette tip.

[0025] Furthermore, the sample plate is a hollow container with a porous plate on the top, and the interior of the container is hollow, which can collect and hold the liquid solvent flowing out of the pointed outlet at the lower end of the sieve plate pipette head.

[0026] In a preferred embodiment, the sample plate is a rectangular hollow container, such as a rectangular hollow box, the top surface of the rectangle is a porous plate with multiple openings for placing multiple sieve plate pipette tips, and the container can collect and hold liquid solvents.

[0027] In the step (2), after the transfer of the filler suspension is completed, the sealing cover of the filler mixing container is preferably covered to keep the filler mixing container sealed to prevent methanol from volatilizing.

[0028] In the step (3), the centrifugal speed can be 1000-8000 rpm, and the centrifugal time is 1-5 minutes.

[0029] In the step (3), a blower may be used to volatilize the solvent methanol.

[0030] In the step (3), the volatilized gaseous methanol can be collected by a vacuum tube, condensed by a condensing device, and combined with the liquid methanol collected by the sample plate, and recycled for use in the step (1) after purification.

[0031] In the method of the present invention, the filler and methanol are mixed and wet-filled into a column, and the carrier solvent methanol also plays the role of activating the filler, that is, the filler is activated by methanol when filling the column. After removing the methanol solvent, the obtained gun-tip solid phase extraction column can directly perform the SPE detection step without activation and can be directly loaded with samples.

[0032] The present invention also provides a column filling device for preparing a gun tip type solid phase extraction column, the device comprises an automated liquid transfer device, a sieve plate liquid transfer gun tip, a working liquid transfer gun tip, a filler mixing container, an oscillator and a centrifuge; a filtering sieve plate is provided inside the tip outlet at the lower end of the sieve plate liquid transfer gun tip; the automated liquid transfer device comprises a liquid transfer gun, a mechanical arm and a moving device for controlling the movement of the mechanical arm; the liquid transfer gun is arranged at one end of the mechanical arm, and the liquid transfer gun and the sieve plate liquid transfer gun tip, and the liquid transfer gun and the working liquid transfer gun tip are detachably assembled.

[0033] Furthermore, the pipette is provided with a gun head connector, which is matched with the upper end opening of the sieve plate pipette head or the working pipette head for detachable assembly.

[0034] The working pipette tip is used to absorb the filler suspension for wet column packing, and no filtering sieve plate is arranged inside the working pipette tip.

[0035] The filter screen in the screen pipette tip has holes or pore structures that allow liquid solvent to pass through.

[0036] The oscillator is provided with a clamp to detachably fix the filler mixing container. The filler mixing container is used to fill the filler suspension. The oscillator is used to oscillate the filler mixing container to keep the filler suspension in the filler mixing container uniformly suspended.

[0037] The upper opening of the filler mixing container is provided with a seal cover that can be opened and closed to prevent the solvent of the filler suspension from volatilizing. In a preferred embodiment, the filler mixing container can be a centrifuge tube with a cover.

[0038] The automated liquid transfer device may further include a cover opening robot arm, which is used to open and close the sealing cover of the filler mixing container.

[0039] The pipette gun is provided with a pipetting control device, which can accurately control the suction and discharge amounts of the liquid.

[0040] Preferably, the pipette gun is connected to an air pump, and the pipette control device controls the air suction or exhaust of the air pump, so that the working pipette gun head mounted on the pipette gun can aspirate or discharge liquid, and the amount of liquid aspirated and discharged can be accurately controlled.

[0041] The moving device for controlling the movement of the robotic arm can be connected to a motion control device to set and control the start and stop and movement path of the robotic arm.

[0042] One or more mechanical arms can be provided, and one or more working pipette tips can also be provided, and parallel processing can be performed according to the automation program. For example, a first mechanical arm can be provided, assembled with a first working pipette tip, sucking up the filler suspension, and pipetting it into the first sieve plate pipette tip; at this time, a second mechanical arm and a second working pipette tip are assembled, sucking up the filler suspension, and pipetting it into the second sieve plate pipette tip, and multiple mechanical arms can operate in parallel. Multiple filler mixing containers can also be provided, and multiple filler mixing containers can share the same oscillator or use different oscillators, and multiple filler mixing containers can be filled with the same filler suspension or different filler suspensions.

[0043] That is, the same or different filler suspensions can be pipetted to multiple screen plate pipette tips at the same time.

[0044] The automated liquid transfer device may also be provided with a transfer robot arm for transferring the sieve plate liquid transfer gun tip containing the filler suspension to the centrifuge.

[0045] Furthermore, the column filling device includes a sample plate for placing a plurality of sieve plate pipette tips and collecting the liquid solvent flowing out of the outlet at the lower end of the sieve plate pipette tip.

[0046] Specifically, the sample plate comprises a porous plate, and the porous plate is provided with a plurality of openings for accommodating a plurality of sieve plate pipette tips.

[0047] Furthermore, the sample plate is a hollow container with a porous plate on the top, and the interior of the container is hollow, which can collect and hold the solvent liquid flowing out of the pointed outlet at the lower end of the sieve plate pipette head.

[0048] In a preferred embodiment, the sample plate is a rectangular hollow container, such as a rectangular hollow box, the top surface of the rectangle is a porous plate with multiple openings for placing multiple sieve plate pipette tips, and the interior of the container can collect and hold solvent liquid.

[0049] The transfer robot can also be used to transfer the sample plate to the centrifuge, or a transfer robot can be provided for transferring the sample plate.

[0050] The centrifuge can centrifuge a sieve plate pipette tip containing a filler suspension or a sample plate containing multiple sieve plate pipette tips, so that the filler in the sieve plate pipette tip is precipitated and separated from the solvent to obtain a filler precipitate and a solvent.

[0051] The device of the present invention may further include a blower, which is used to volatilize the solvent in the screen plate pipette tip after centrifugation.

[0052] The device of the present invention may also include an air extraction pipe and a condensation device, wherein the air extraction pipe is used to extract the volatilized gaseous solvent, and the condensation device is used to condense the gaseous solvent. The solvent recovered by condensation is combined with the liquid solvent collected by the sample plate and can be recycled.

[0053] The device of the present invention may also include a control unit for receiving user instructions and transmitting the instructions to the motion control device and the pipetting control device.

[0054] The present invention designs an automated self-filling column device, uniformly suspends the filler in a methanol carrier solution and activates it, uses a working pipette tip to transfer the filler-methanol suspension to a sieve plate pipette tip, centrifuges the filler, removes the solvent, and realizes the production of high-throughput SPE tips without the support of additional tips and other devices. Thanks to the advantages of the automated small volume precision pipetting of the filler self-filling and the filler suspension, the program can be set to accurately control the column loading amount of the filler, and the carrier solution also plays the role of activating the filler, reducing the use of the processing solvent, and efficiently realizing the production of the SPE tips on the basis of ensuring the accuracy of the column loading amount of the filler. In addition, the SPE tip is small because the injection volume is small, and it is limited by the sample volume. It can process multiple samples simultaneously in combination with the automated SPE device, so the SPE tip is very suitable for high-throughput SPE experiments, and it is placed in a 96-well sample plate for production, and 8 channels or even 96 channels of SPE experiments can be directly performed later, meeting the needs of various high-throughput technologies, and there is a very large practical value.

[0055] The beneficial effects of the present invention are:

[0056] 1. Use a fully automatic mechanized method to solve the time-consuming problem of SPE column loading in the market.

[0057] 2. The carrier solution is used to suspend the filler particles and oscillate them, so that the carrier particles will not precipitate and stratify, and the density of each part of the suspension is kept consistent. The filler quality in the obtained SPE pipette tip is consistent and the error is small, which solves the problem of large error in the quality of the filler in the SPE pipette tip on the market.

[0058] 3. The method of the present invention can produce SPE tip groups of 96 in a unit, which can subsequently provide 8-channel high-throughput SPE experiments simultaneously. In addition, for the demand for particularly large sample volumes, the pipetting system can also be converted to a maximum of 96 channels to achieve high-throughput sample detection, solving the time-consuming problem of SPE experiments on the market.

[0059] 4. The SPE pipette tip is obtained by a self-filling column device, and the quality of the filler is highly reproducible and controllable. The amount of filler is calculated in advance according to the batch sample, which avoids the problem of limited specifications of SPE pipette tips on the market and waste of filler when processing trace samples. For example, an SPE pipette tip with a 5mg filler specification may be used to process a 10 ng-300 μg sample, which will lead to a waste of filler. The present invention can reduce the amount of filler and solvent used, and the filler loading amount can be as low as the microgram level, such as 5 μg. For high-throughput testing, the amount of expensive filler can be greatly reduced, thereby reducing operating costs and solving the problem of high SPE cost on the market.

[0060] 5. Solve the problem of separation between the production and use of SPE devices in the market. If you want to connect the production, filler activation and use processes of SPE tips, you can only combine the three steps into two, otherwise the time cost will increase, which is not conducive to the efficient processing of samples. The present invention adopts a self-filling column workstation to produce SPE tips. The filler is also activated during filling. Finally, the activation solvent, that is, the carrier solution, is evaporated and removed. The tip assembly is completed. At the same time, the filler activation is completed, and the sample can be directly loaded for SPE experiment to obtain the target compound, which reduces the subsequent SPE experimental steps, simplifies the experimental process, and improves the experimental efficiency.

[0061] 6. The solvent methanol not only acts as a carrier solution to suspend the filler particles, but also activates the filler. After evaporation, the methanol gas is collected through the exhaust pipe and condensed for recovery. The liquid methanol is recovered through the sample plate. Therefore, the methanol solvent can be recycled, further reducing the production cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0062] Figure 1 This is a schematic diagram of the structure of a column filling device for preparing a gun-tip solid phase extraction column. DETAILED DESCRIPTION

[0063] The technical solution of the present invention is further described below with reference to specific embodiments, but the protection scope of the present invention is not limited thereto.

[0064] Example 1

[0065] The present invention provides a column filling device for preparing a gun-tip type solid phase extraction column, the structural schematic diagram of which is shown in FIG. Figure 1 As shown, Figure 1 In the figure, 1 is a working pipette tip, 2 is a sieve plate pipette tip, 3 is a sieve plate pipette tip filled with a filler suspension, 4 is a pipette, 5 is a robot arm, 6 is a cover opening robot arm, 7 is a filler mixing container, 8 is an oscillator, 9 is a transfer robot arm, 10 is a sample plate, 11 is a centrifuge, 12 is a blower, and 13 is an exhaust pipe.

[0066] The column filling device for preparing a gun tip type solid phase extraction column comprises an automated liquid transfer device, a sieve plate liquid transfer gun tip 2, a working liquid transfer gun tip 1, a filler mixing container 7, an oscillator 8 and a centrifuge 11; a filtering sieve plate is provided inside the tip outlet at the lower end of the sieve plate liquid transfer gun tip 2; the automated liquid transfer device comprises a liquid transfer gun 4, a mechanical arm 5 and a moving device (not shown) for controlling the movement of the mechanical arm; the liquid transfer gun 4 is arranged at one end of the mechanical arm 5, and the liquid transfer gun 4 and the sieve plate liquid transfer gun tip 2, and the liquid transfer gun 4 and the working liquid transfer gun tip 1 are all detachably assembled.

[0067] Furthermore, the pipette gun 4 is provided with a gun head connector, which is adapted to the upper end opening of the sieve plate pipette gun head 2 or the working pipette gun head 1 for detachable assembly. The detachable assembly between the gun head connector of the pipette gun and the gun head is well known to those skilled in the art. For example, the gun head connector is a gun head slot, and the upper end opening of the sieve plate pipette gun head 2 or the working pipette gun head 1 is pushed in to install and fix it. Alternatively, it can be a snap-on method, etc.

[0068] The working pipette tip 1 is used to absorb the filler suspension for wet column loading, and no filter screen is provided inside the working pipette tip 1 .

[0069] The filter screen in the screen pipette tip 2 has holes or pores that allow liquid solvent to pass through.

[0070] The oscillator 8 is provided with a clamp to detachably fix the filler mixing container 7. The filler mixing container 7 is used to fill the filler suspension. The oscillator 8 is used to oscillate the filler mixing container 7 to keep the filler suspension in the filler mixing container uniformly suspended.

[0071] A seal cover that can be opened and closed is provided at the upper opening of the filler mixing container 7. The filler mixing container 7 can be a centrifuge tube with a cover or the like.

[0072] The automated liquid transfer device may further include a cover opening robot arm 6 , and the cover opening robot arm 6 is used to open and close the sealing cover of the filling mixing container 7 .

[0073] The pipette gun 4 is provided with a pipetting control device, which can accurately control the amount of liquid sucked and discharged.

[0074] Preferably, the pipette 4 is connected to an air pump, and the pipette control device controls the suction or exhaust of the air pump, so that the working pipette head assembled on the pipette can aspirate or discharge liquid, and the aspiration and discharge amounts of liquid can be accurately controlled.

[0075] The moving device for controlling the movement of the robotic arm can be connected to a motion control device to set and control the start and stop and movement path of the robotic arm.

[0076] One or more mechanical arms can be provided, and one or more working pipette tips can also be provided, and parallel processing can be performed according to the automation program. For example, a first mechanical arm can be provided, assembled with a first working pipette tip, sucking up the filler suspension, and pipetting it into the first sieve plate pipette tip; at this time, a second mechanical arm and a second working pipette tip are assembled, sucking up the filler suspension, and pipetting it into the second sieve plate pipette tip, and multiple mechanical arms can operate in parallel. Multiple filler mixing containers can also be provided, and multiple filler mixing containers can share the same oscillator or use different oscillators, and multiple filler mixing containers can be filled with the same filler suspension or different filler suspensions.

[0077] That is, the same or different filler suspensions can be pipetted to multiple screen plate pipette tips at the same time.

[0078] The automated liquid transfer device may also be provided with a transfer robot 9 for transferring the sieve plate pipette tip containing the filler suspension to the centrifuge 11 .

[0079] Furthermore, the column filling device includes a sample plate 10 for placing a plurality of sieve plate pipette tips 2 and collecting the liquid solvent flowing out of the outlet at the lower end of the sieve plate pipette tips 2 .

[0080] The sample plate 10 comprises a porous plate, and the porous plate is provided with a plurality of openings for accommodating a plurality of screen plate pipette tips 2 .

[0081] Furthermore, the sample plate 10 is a hollow container with a porous plate on the top, and the interior of the container is hollow, which can collect and hold the solvent liquid flowing out of the outlet of the tip at the lower end of the sieve plate pipette head.

[0082] In a preferred embodiment, Figure 1 As shown, the sample plate 10 is a rectangular box with a hollow interior. The top surface of the rectangular box is a porous plate with 96 openings for placing multiple sieve plate pipette tips. The interior of the box can collect and hold solvent liquid.

[0083] A valve for liquid outflow may be provided at the bottom or side of the rectangular hollow container, so as to transfer and recycle the liquid methanol collected in the container after the column filling is completed.

[0084] When the filler suspension is wet packed into the column, the solvent will flow out from the pointed outlet at the lower end of the frit pipette tip due to gravity and can be collected at the bottom of the container on the sample plate.

[0085] When the filler suspension is centrifuged for solid-liquid separation, part of the solvent flows out from the pointed outlet at the lower end of the sieve plate pipette tip and is collected by the hollow container of the sample plate, while part of the solvent evaporates from the opening at the upper end of the sieve plate pipette tip.

[0086] The transfer robot 9 can also be used to transfer the sample plate 10 .

[0087] When the transfer robot arm 9 is used to transfer the sieve plate pipette tip 2, the transfer can be performed by arranging a pipette on the transfer robot arm and detachably assembling the pipette tip 2 with the sieve plate pipette tip.

[0088] When the transfer robot arm 9 is used to transfer the sample plate 10, a gripper may be provided to transfer the sample plate.

[0089] The centrifuge 11 can centrifuge a sieve plate pipette tip containing a filler suspension or a sample plate 10 containing multiple sieve plate pipette tips containing a filler suspension, so as to separate the filler suspension in the sieve plate pipette tip from solid to liquid to obtain a precipitate and a solvent.

[0090] The device of the present invention may also include a blower 12 and an exhaust pipe 13. The blower 12 blows air to volatilize the solvent in the centrifuged sieve plate pipette tip, and the exhaust pipe 13 is used to collect the volatilized gaseous solvent. The exhaust pipe 13 can be connected to a condensing device to condense the gaseous solvent alcohol, which can be combined with the liquid solvent collected at the bottom of the sample plate 10 to obtain a recovered solvent. The concentration of the solvent is tested. If it meets the use standard, it can be directly recycled. If it does not meet the standard, it can be further purified and recycled. The method for purifying and recovering methanol can be distillation, organic membrane purification, etc.

[0091] The device of the present invention may also include a control unit for receiving user instructions and transmitting the instructions to the motion control device and the pipetting control device.

[0092] In a preferred embodiment, the sample plate adopts a rectangular hollow box with 96 holes on the top, and 96 screen plate pipette tips are used as a unit. The filling material is mainly 5 mg, 10 mg, 30 mg, 60 mg, etc. according to the needs. There are also lower requirements for the weight of the filler during the refining process, such as 5 μg, 10 μg, 30 μg, etc. The bottom or side of the rectangular hollow box can be provided with a valve for liquid outflow, which is used to recover liquid methanol after the column filling is completed.

[0093] The volume of the screen pipette tip can be 200 μL (suspension concentration is 2 mg ml -1 , 10 mg ml -1 , 25mg·ml -1 , 50 mg ml -1 , 150 mg ml -1 、300 mg·ml -1 ), which can be used in single-channel, eight-channel and even 96-channel automatic pipetting systems, laying the foundation for subsequent high-throughput purification and recovery.

[0094] The centrifuge can be a 96-well plate centrifuge, which can process two 96-well sample plates at the same time. The 96-well plate centrifuge is designed with a special rotor adapted to the 96-well sample plate to ensure that each well can be evenly stressed. Existing commercially available devices can be used.

[0095] The present invention also provides a method for automatically preparing a gun-tip type solid phase extraction column using a column filling device, which specifically comprises the following steps:

[0096] (1) Mix 12 g of HLB filler and 40 mL of methanol to prepare a filler suspension. Pour the filler suspension into a 50 mL centrifuge tube with a lid and cover it with a sealing cap. Oscillate the centrifuge tube with an oscillator, oscillate vertically at a frequency of 200 times / min to keep the filler suspension evenly suspended. Keep oscillating and keep the centrifuge tube sealed when not pipetting to prevent methanol from volatilizing.

[0097] (2) The cover-opening robot opens the sealing cover, and the pipette gun and the working pipette gun head are assembled and connected. The working pipette gun head can be used to transfer the liquid volume from 50 μL to 200 μL. The motion control system controls the movement of the robot gun to transfer the liquid to the sieve plate pipette gun head placed on the 96-well sample plate. The transfer volume can be selected from 5 μL to 200 μL, and the number of transfers can be adjusted. Multiple pipette guns and robot arms can be used to perform parallel pipetting operations until all 96 sieve plate pipette gun heads contain the same amount of filler suspension. Different amounts or types of filler suspensions can also be loaded. In this way, the sieve plate pipette gun heads on two 96-well sample plates are filled with filler suspensions, and the liquid methanol flowing out of the lower end outlet of the sieve plate pipette gun head is collected in the hollow container at the bottom of the sample plate.

[0098] (3) The transfer robot (with a gripper) transfers the 96-well sample plate equipped with 96 sieve plate pipette tips to a 96-well plate centrifuge; two 96-well sample plates are placed symmetrically and centrifuged at 3800 rpm for 2 minutes to precipitate HLB to obtain HLB filler precipitate and solvent; during centrifugation, part of the solvent flows out from the tip outlet at the lower end of the sieve plate pipette tip and is collected in a container at the bottom of the sample plate, and part of the solvent evaporates from the opening at the upper end of the sieve plate pipette tip; the transfer robot transfers the 96-well sample plate to the blower outlet, starts the blower to blow air to evaporate the solvent methanol, and obtains a tip-type solid phase extraction column. The exhaust pipe continuously exhausts air at negative pressure to collect the volatilized methanol solvent, which is condensed by a condensation device and combined with the collected liquid methanol. After purification, it can be recycled in step (1).

[0099] The present invention adopts a mechanical automation device to produce SPE tips, and simultaneously performs the production of tips and the activation steps of fillers, so as to ensure the quality requirements of tip production and the sufficiency of filler activation. Under the premise of ensuring that mass spectrometry can be directly loaded, 96 SPE tips can be produced as a unit, meeting the requirements of high-throughput sample processing. From the cost point of view, the automated process ensures that the entire process only needs to transfer a fixed amount of filler-carrier solution gun tips and containers for suspending filler-carrier solution, without the need for additional clean containers and gun tips, and when centrifuging to evaporate the solvent, the 96-well plates are paired in pairs, the production efficiency is doubled, the production time is greatly reduced, the cost is greatly reduced, and basically no manual operation is required.

Claims

1. A method for preparing a gun-tip solid phase extraction column that does not require activation, characterized in that The method comprises the following steps: (1) The solid phase extraction filler and methanol are mixed to obtain a filler suspension, the filler suspension is placed in a filler mixing container, and the container is sealed with a sealing cover. The filler mixing container is shaken by an oscillator to keep the filler suspension uniformly suspended; the volume of methanol used is 3-500 μL / mg based on the mass of the filler; (2) placing a plurality of sieve plate pipette tips on the sample plate, wherein a filter sieve plate is provided inside the pointed outlet at the lower end of the sieve plate pipette tip; after opening the cover of the filler mixing container, using a pipetting device to absorb the filler suspension and accurately pipetting it into the sieve plate pipette tip on the sample plate; (3) placing a sieve plate pipette tip or a sample plate equipped with multiple sieve plate pipette tips into a centrifuge; using the centrifuge to perform centrifugation to separate the filler suspension in the sieve plate pipette tip into solid and liquid to obtain filler precipitate and solvent; volatilizing the solvent methanol to prepare the tip-type solid phase extraction column that does not require activation, collecting liquid methanol in the sample plate, extracting and collecting the volatilized gaseous methanol, condensing it and combining it with the liquid methanol, purifying it and recycling it in step (1).

2. The method according to claim 1, characterized in that In the step (2), the filter sieve plate in the sieve plate pipette tip has holes or pores that allow liquid solvent to pass through, and solid phase extraction filler particles cannot pass through the holes or pores.

3. A column filling device for preparing a gun-tip solid phase extraction column that does not require activation, characterized in that The device comprises an automated liquid transfer device, a screen plate liquid transfer gun head, a working liquid transfer gun head, a filler mixing container, an oscillator and a centrifuge; a filter screen plate is provided inside the tip outlet at the lower end of the screen plate liquid transfer gun head; the automated liquid transfer device comprises a liquid transfer gun, a mechanical arm and a moving device for controlling the movement of the mechanical arm; the liquid transfer gun is arranged at one end of the mechanical arm, and the liquid transfer gun and the screen plate liquid transfer gun head, and the liquid transfer gun and the working liquid transfer gun head are detachably assembled; The column filling device comprises a sample plate, and the sample plate is used to place a plurality of sieve plate liquid transfer gun tips and collect the liquid solvent flowing out of the outlet at the lower end of the sieve plate liquid transfer gun tip; The filler mixing container is used to fill the filler suspension; The oscillator is provided with a clamp to detachably fix the filler mixing container; The oscillator is used to oscillate the filler mixing container to keep the filler suspension in the filler mixing container uniformly suspended; The centrifuge is used to centrifuge the sieve plate pipette tip containing the filler suspension, so that the filler in the sieve plate pipette tip is precipitated and separated from the solvent solid and liquid; the device also includes a blower, which is used to volatilize the solvent in the sieve plate pipette tip after centrifugation; The device also includes an air extraction pipe and a condensation device. The air extraction pipe is used to extract the volatilized gaseous solvent, and the condensation device is used to condense the gaseous solvent.

4. The column filling device for preparing a gun-tip type solid phase extraction column that does not require activation as claimed in claim 3, characterized in that The working pipette tip is used to absorb the filler suspension for wet column packing, and no filtering sieve plate is arranged inside the working pipette tip.

5. The column filling device for preparing a gun-tip type solid phase extraction column that does not require activation as claimed in claim 3, characterized in that The upper opening of the filler mixing container is provided with an openable and closable sealing cover to prevent the solvent of the filler suspension from volatilizing.

6. The column filling device for preparing a gun-tip type solid phase extraction column that does not require activation as claimed in claim 5, characterized in that The automated liquid transfer device further comprises a cover-opening mechanical arm, which is used for opening and closing the sealing cover of the filling mixing container.

7. The column filling device for preparing a gun-tip type solid phase extraction column that does not require activation as claimed in claim 3, characterized in that The automated liquid transfer device also includes a transfer robot arm for transferring a sample plate or a transfer screen plate liquid transfer gun head.

8. The column filling device for preparing a gun-tip type solid phase extraction column that does not require activation as claimed in claim 3, characterized in that The centrifuge is used to centrifuge a sample plate equipped with a plurality of sieve plate pipette tips, so as to separate the solid and liquid of the filler suspension in the sieve plate pipette tips to obtain filler precipitation and solvent.

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