CN116338074A - Sample application method and device - Google Patents

Abstract

The automatic sample application method includes the steps of cleaning the inner wall of the sample application needle, cleaning the outer wall of the sample application needle, sucking gas or immiscible liquid, sucking liquid to be tested, eliminating liquid drop and applying sample, and the six steps constitute one operation unit. The device and the method of the invention can automatically sample the sample and automatically collect the fraction. The automatic sample application and automatic fraction collection are fast, the sample application amount and the fraction collection amount are accurate, the fraction collection and detection analysis of chemical reaction can be accelerated, the detection cost is saved, the time cost is shortened, and the research and development process is accelerated.

Description

Sample application method and device

Technical Field

The invention relates to the field of organic synthetic chemistry, in particular to a sample application method and a sample application device.

Background

Thin layer chromatography is a chromatographic separation technique that uses a support coated on a support plate as a stationary phase and a suitable solvent as a mobile phase to separate, identify and quantify a mixed sample. The adsorption capacity of each component to the same adsorbent is different, so that adsorption, desorption, re-adsorption and re-desorption are continuously generated in the process of flowing a mobile phase (solvent) through a stationary phase (adsorbent), and the purpose of mutually separating each component is achieved. Thin layer chromatography is an important experimental technique for rapid separation and qualitative analysis of small amounts of substances, and is commonly used in organic synthetic chemistry to track the progress of chemical reactions. The thin layer chromatography has the advantages of convenient operation, simple equipment, easy color development and high development rate, and is the most common method for detecting and analyzing the components of the reaction liquid and tracking the progress of chemical reaction in the reaction process.

Half of the thin layer chromatography process involves:

1) Sample application, in which the sample is made into 1% -5% solution, and the sample is applied by using capillary tube whose internal diameter is less than 1 mm. A line is gently drawn by a pencil at a distance of 1-1.5cm from the lower end of the thin layer plate, which is called an initial line, and the sample is spotted for the second time after the solvent volatilizes (generally repeated for 3-5 times). Of course, we can spot multiple samples on one chromatographic plate, typically 1-1.5cm apart.

2) And (3) spreading, namely obliquely placing the sampled thin layer plate in a spreading cylinder, and taking out the thin layer plate and drawing a front line by using a pencil when the front edge of the spreading agent moves upwards to be 1-1.5cm away from the upper edge of the thin layer plate.

3) Color development, if the sample is colored after being unfolded, the sample can be directly observed. If there is no color, the color is developed by spraying a color developing agent or by iodine fumigation. If the product we want to do is fluorescent, it can also be observed under UV light.

However, thin layer chromatography, which is common in laboratories at present, is manually operated for sample application, and has high labor and time costs; operators are easy to be poisoned by toxic reagents, and the safety is low; in addition, in manual operation, the sample application amount of the reaction liquid is difficult to accurately control, accurate quantification is difficult to carry out, and the requirement of an experimental process cannot be met.

Disclosure of Invention

According to a first aspect, in an embodiment, there is provided a spotting method comprising the steps of, in order:

a step of cleaning the inner wall of the sample application needle, wherein a control mechanism controls the sample application needle to move to the upper part of the waste liquid bottle, and a pump connected with the sample application needle pumps cleaning solvent into the sample application needle to clean the inner wall of the sample application needle;

a step of cleaning the outer wall of the sample application needle, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle filled with the cleaning solvent and controls the sample application needle to extend into the reagent bottle filled with the cleaning solvent, the sample application needle at least partially submerges into the cleaning solvent in the reagent bottle, and the cleaning solvent in the reagent bottle is used for cleaning the outer wall of the sample application needle; and/or the control mechanism controls the other flow path to pump a cleaning solvent to the outer wall of the sample application needle so as to clean the outer wall of the sample application needle;

a step of sucking gas or immiscible liquid, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle containing air, and the pump controls the sample application needle to suck air as a gas interval or suck liquid which is immiscible with the solution to be measured as a liquid interval;

a liquid sucking step, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle containing the sample liquid, controls the sample application needle to extend into the reagent bottle containing the sample liquid, and controls the sample application needle to suck the sample liquid through the pump;

A liquid drop removing step, wherein the control mechanism controls the sample application needle to contact with the liquid suction ball, so that the liquid suction ball wipes off liquid drops of the needle head of the sample application needle and enters the sample application step;

and a sample application step, wherein the control mechanism controls the sample application needle to move to the upper part of the sample application plate, controls the sample application needle to contact with the sample application position on the sample application plate, injects a set amount of solution to be tested into the sample application needle through a pump, returns to the inner wall cleaning step after the sample application is finished, and circulates in this way to finish sample application of the sample application position on the sample application plate.

According to a second aspect, in an embodiment, there is provided a method of collecting fractions comprising performing fraction collection using the spotting device of the first aspect, the control mechanism controlling the spotting needles to inject the fractions to be collected into the respective empty reagent bottles. The fraction to be collected may be a fraction obtained after a chemical reaction, and the liquid collected to the reagent bottle may be used for spot plate, detection, separation, or the like.

According to a third aspect, in an embodiment, a spotting device is provided comprising a spotting needle for collecting, sucking and spotting a sample;

a control mechanism for controlling the movement of the sample application needle;

a plurality of reagent bottles which are respectively used for containing solution to be tested, cleaning solvent, air and waste liquid;

The sample plate is used for sample application of sample application needles;

a liquid suction ball for removing liquid drops of the needle head of the sample application needle;

the control mechanism controls the sample application needle to finish liquid injection, sample suction, sample application and cleaning;

or, the reagent bottles are empty bottles for containing liquid to be collected, the control mechanism controls the sample application needle to inject the liquid to be collected into each empty reagent bottle, the liquid to be collected can be fractions obtained after chemical reaction, and the liquid collected into the reagent bottles can be used for spot plates, detection, separation and the like.

According to the sample application method and the sample application device, the sample application method has the advantages of high sample application speed and accurate sample application amount, and can accelerate fraction collection and/or detection analysis of chemical reaction, save detection cost, shorten detection time and accelerate research and development processes.

In one implementation, the device of the present invention may be automated for both spotting and collecting fractions.

Drawings

FIG. 1 is a schematic perspective view of an automatic spotting device according to one embodiment;

FIG. 2 is a schematic diagram showing an exploded structure of the automatic spotting device according to one embodiment;

FIG. 3 is a side view of an automated spotting device of one embodiment;

FIG. 4 is an enlarged schematic view of portion A of FIG. 1;

FIG. 5 is a schematic view of the structure of the print needle in FIG. 1;

FIG. 6 is a schematic perspective view of an automatic spotting device with a wash line;

FIG. 7 is an enlarged schematic view of portion B of FIG. 6;

FIG. 8 is a schematic diagram of an automatic spotting process according to one embodiment;

FIG. 9 is a photograph of a spot plate of one embodiment;

fig. 10 is a standard graph of an embodiment.

Description of the reference numerals: 1. a base; 2. a console; 3. an X axis; 4. a Y axis; 5. a Z axis; 500. a connecting piece; 51. a clamping table; 52. a locking piece; 521. a locking hole; 522. a locking member; 53. a cutting sleeve; 54. a groove; 6. a sample application needle; 7. a sample cartridge; 71. a reagent bottle; 8. a sample application box; 81. a spot plate; 82. a liquid suction ball; 83. a clamping groove; 9. cleaning a pipeline; 91. and a liquid outlet.

Detailed Description

The invention will be described in further detail below with reference to the drawings by means of specific embodiments. Wherein like elements in different embodiments are numbered alike in association. In the following embodiments, numerous specific details are set forth in order to provide a better understanding of the present application. However, one skilled in the art will readily recognize that some of the features may be omitted, or replaced by other elements, materials, or methods in different situations. In some instances, some operations associated with the present application have not been shown or described in the specification to avoid obscuring the core portions of the present application, and may not be necessary for a person skilled in the art to describe in detail the relevant operations based on the description herein and the general knowledge of one skilled in the art.

Furthermore, the described features, operations, or characteristics of the description may be combined in any suitable manner in various embodiments. Also, various steps or acts in the method descriptions may be interchanged or modified in a manner apparent to those of ordinary skill in the art. Thus, the various orders in the description and drawings are for clarity of description of only certain embodiments, and are not meant to be required orders unless otherwise indicated.

The numbering of the components itself, e.g. "first", "second", etc., is used herein merely to distinguish between the described objects and does not have any sequential or technical meaning. The terms "connected" and "coupled," as used herein, are intended to encompass both direct and indirect connections (couplings), unless specifically indicated otherwise.

Thin layer chromatography (thin-layer chromatography, TLC): the chromatographic separation technology is to separate, identify and quantify mixed sample with the support coated on the support board as stationary phase and proper solvent as mobile phase. The adsorption capacity of each component to the same adsorbent is different, so that adsorption, desorption, re-adsorption and re-desorption are continuously generated in the process of flowing a mobile phase (solvent) through a stationary phase (adsorbent), and the purpose of mutually separating each component is achieved. Thin layer chromatography is an important experimental technique for rapid separation and qualitative analysis of small amounts of substances, and is commonly used in organic synthetic chemistry to track the progress of chemical reactions.

Organic synthetic chemistry (organic synthesis chemistry): refers to a process of synthesizing an organic substance from a simpler compound or simple substance through a chemical reaction. Sometimes also including the degradation from complex starting materials to simpler compounds.

Flow chemistry (Flow chemistry), also known as continuous Flow chemistry or microchannel chemistry, refers to the process of adding, mixing, reacting, separating and purifying chemical reactants in a continuous Flow system, wherein the process of adding, mixing, reacting, separating and purifying reactants is performed continuously. Is a concept for distinguishing batch chemistry, flow chemistry typically uses microchannel chips, coils, packed beds, etc. as reactors.

The invention belongs to the field of organic synthesis, and relates to a process for synthesizing an organic substance from a simpler compound or simple substance through chemical reaction. Sometimes also including the degradation from complex starting materials to simpler compounds. Organic synthesis is the main means and tool for creating new molecules as the basis of organic chemistry. Over 100 years, organic chemists synthesize thousands of medicines, pesticides, fragrances, dyes, other organic functional molecules and the like, so that the organic synthesis industry rises rapidly, and the rapid development of a plurality of industries closely related to national economy and daily life of people is promoted. Organic synthetic chemistry presents no more attractive prospect to people and becomes the most active and most vitalizing branch of organic chemistry.

One core of chemical synthesis is the synthesis of a wide variety of compound molecules, such as natural product molecules, drug molecules, functional molecules, material molecules, and the like. During synthesis, a large amount of chemical reactions are often required, glass flasks or reaction kettles are mostly adopted as reactors in traditional laboratories, and some laboratories adopt flow chemistry to perform chemical synthesis experiments, and microchannel reactors or coils and the like are adopted as reactors. The general procedure for the experimental operation is: firstly, preparing a reaction reagent, a solvent, a flow chemical device and the like, and preparing the reaction reagent solvent into a solution for later use; then the reaction liquid is input into a reactor through a transfusion pump for mixed reaction, and the reactor is placed under the required reaction condition; detecting and analyzing the components of the reaction liquid in real time in the reaction process, and tracking the progress of chemical reaction; finally, the required reaction solution is collected and subjected to post-treatment.

Among them, detection and analysis of the components of the reaction solution and tracking of the progress of the chemical reaction are very important works, which determine the success or failure and quality of the chemical reaction. In the experimental process, liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS), ultraviolet visible spectrometry and thin layer chromatography are generally adopted to detect and analyze the components of the reaction solution, and the progress of chemical reaction is tracked. The thin layer chromatography is free from expensive detection instruments, short in detection and analysis time, low in cost, simple and convenient to use, and is the most widely used method for tracking the progress of chemical reaction.

According to a first aspect, in an embodiment, a printing method is provided, and a printing device used may refer to fig. 1 to 5, and the method sequentially includes the following steps:

a step of cleaning the inner wall of the sample application needle, wherein the control mechanism controls the sample application needle 6 to move to the upper part of the waste liquid bottle, and a pump connected with the sample application needle 6 pumps cleaning solvent into the sample application needle 6 to clean the inner wall of the sample application needle 6;

and a step of cleaning the outer wall of the sample application needle, wherein the control mechanism controls the sample application needle 6 to move to the position above the reagent bottle filled with the cleaning solvent, and controls the sample application needle 6 to extend into the reagent bottle filled with the cleaning solvent, and at least part of the sample application needle 6 is immersed into the cleaning solvent in the reagent bottle, and the cleaning solvent in the reagent bottle is used for cleaning the outer wall of the sample application needle 6.

A step of sucking gas or immiscible liquid, wherein the control mechanism controls the sample application needle 6 to move to the position above the reagent bottle containing air, and the pump controls the sample application needle 6 to suck air as a gas interval or suck liquid which is immiscible with the solution to be tested as a liquid interval; the gas interval or the liquid interval effectively reduces the cross contamination of the solution, so that the waste liquid is easier to remove after the sample is printed;

a liquid sucking step, wherein the control mechanism controls the sample application needle 6 to move to the position above the reagent bottle containing the sample liquid, controls the sample application needle 6 to extend into the reagent bottle containing the sample liquid, and controls the sample application needle 6 to suck the sample liquid through the pump;

A drop removing step, wherein the control mechanism controls the sample application needle 6 to contact the liquid suction ball 82, so that the liquid suction ball 82 wipes off drops of the needle head of the sample application needle 6 and then enters a sample application step; the step effectively avoids inaccurate sample application results caused by different amounts of liquid remained by the needle points of the sample application needle 6 during sample application, and the amounts of the liquid at the needle points are almost equal after the sample application needle points are wiped by the liquid suction ball before sample application, so that the accuracy of detection results is improved;

and in the sample application step, the control mechanism controls the sample application needle 6 to move to the upper part of the sample application plate 81, controls the sample application needle 6 to contact the sample application position on the sample application plate 81, injects a set amount of solution to be tested into the sample application needle 6 through a pump, returns to the inner wall cleaning step after sample application is finished, and circulates in this way, so as to finish sample application of the sample application position on the sample application plate 81.

In one embodiment, the above six basic steps constitute one operation unit, and the spotting method is a cyclic operation process consisting of a custom number of operation units.

In another embodiment the control means control a further flow path to pump a cleaning solvent to the outer wall of the print needle 6, cleaning the outer wall of the print needle 6. Specifically, as shown in fig. 6 and 7, a cleaning pipeline 9 is disposed on the Z-axis 5, a liquid outlet of the cleaning pipeline 9 is close to the outer wall of the sample needle 6, and the control mechanism pumps the cleaning solvent into the cleaning pipeline 9 by controlling the pump to flow out from the liquid outlet 91 so as to clean the outer wall of the sample needle 6. The cleaning pipeline 9 can be arranged on the clamping table 51, so that the stability of the structure is improved, and shaking is avoided.

The sample application needle 6 can only wash the outer wall through the cleaning solvent in the reagent bottle, can also only wash the outer wall through the cleaning pipeline 9, also can combine reagent bottle cleaning and pipeline cleaning, realize the more thorough washing to sample application needle 6 outer wall.

In one embodiment, after each sample site on the sample plate 81 is completely sampled, the sample plate 81 is put into the developing agent to climb the plate, and after the climbing plate is completed, the developing agent on the sample plate 81 is dried and removed.

In one embodiment, the dried silica gel plate is placed under an ultraviolet lamp, or iodized or other color development modes, and then photographed to obtain a photograph of the silica gel plate, and the concentration of the product in the solution to be measured is obtained according to the analytical calculation of the photograph.

In one embodiment, the dried silica gel plate is placed in a gel imaging analyzer or other photographing device to take a photograph of the silica gel plate.

In one embodiment, after the pipetting step and before the de-dripping step, the method further comprises an evacuating step, wherein the control mechanism controls the sample needle 6 to move above a reagent bottle for receiving waste liquid on the sample box 7, and the sample needle 6 is evacuated by a pump; the pump has a gear clearance when switching from suction to injection mode, and the evacuation is to remove the clearance when switching modes; if there is no play in the gears of the pump, no evacuation step is required.

In one embodiment, the control mechanism controls the sample application needle 6 to move to the position above the reagent bottle on the sample application box 8 and controls the sample application needle 6 to contact with the liquid suction ball 82 at the bottle mouth of the reagent bottle, and the sample application needle 6 is injected with the solution to be tested through the pump while the sample application needle 6 moves, and the liquid suction ball 82 is used for entering the sample application step after wiping off the liquid drops of the needle head of the sample application needle 6.

In one embodiment, as shown in fig. 1, 2 and 3, the spotting device used in the spotting method includes a spotting needle 6 for collecting a sample, sucking the sample, and spotting;

a control mechanism for controlling the movement of the sample application needle 6;

a plurality of reagent bottles which are respectively used for containing solution to be tested, cleaning solvent, air and waste liquid;

a sample plate 81 for sample application by the sample application needle 6;

a liquid suction ball 82 for removing the liquid drop of the needle head of the sample application needle 6 before sample application;

the control mechanism controls the sample application needle 6 and the pump to finish liquid injection, sample suction, sample application and cleaning;

or, the reagent bottles are empty bottles for containing the liquid to be collected, the control mechanism controls the sample application needle 6 to inject the liquid to be collected into each empty reagent bottle, the liquid to be collected can be fractions obtained after chemical reaction, and the liquid collected into the reagent bottles can be used for spot plates, detection, separation and the like. The printing device herein is only exemplary and other devices capable of performing the printing method of the present invention are within the scope of the present invention.

In one embodiment, as shown in fig. 1, the control mechanism includes a console 2, and an X axis, a Y axis, and a Z axis disposed on the console 2, where the X axis, the Y axis, and the Z axis are perpendicular to each other, the sample application needle 6 is fixed on the Z axis, and the control mechanism controls the sample application needle 6 to perform translational motion along the X axis and the Y axis, and perform linear lifting motion along the Z axis, so as to accomplish liquid injection, sample suction, sample application, and cleaning. The action of the sample application needle 6 completed under the control of the control mechanism comprises suspending above the waste liquid bottle to clean the inner wall of the sample application needle, injecting liquid into the reagent bottle for containing the solution to be tested, sucking sample from the reagent bottle containing the solution to be tested, applying sample at the contact point template 81, entering the reagent bottle containing the cleaning solvent to clean the outer wall of the sample application needle, and contacting the liquid suction ball to remove the liquid drop of the needle head.

In an embodiment the printing device further comprises a fluid circuit system in communication with the print needle 6, the fluid circuit system comprising a pump and a pipeline for injecting a liquid into the print needle 6 or controlling the suction of the print needle 6. The pump is connected to the sample application needle 6 by a tubing.

In an embodiment, the control mechanism is electrically connected to the pump, and the control mechanism controls the X-axis, Y-axis, Z-axis movements such that the sample application needle 6 completes the injection of the liquid to be collected into the empty reagent bottle.

In an embodiment, the control mechanism further comprises a hand touch screen 21 arranged on the console 2, and a worker can set a required program through the hand touch screen 21, so that the subsequent sample application needle 6 performs corresponding actions according to the set program.

In one embodiment, the control mechanism comprises a programmable controller or a single chip microcomputer embedded system for controlling the X-axis, Y-axis and Z-axis movements and the liquid injection and washing of the sample application needle 6.

In an embodiment, as shown in fig. 1, the deposition device further includes a deposition box 8, a deposition plate 81 is disposed on the deposition box 8, and the control mechanism controls the deposition needle 6 to contact the deposition plate 81 to complete deposition.

In one embodiment, as shown in fig. 1, a reagent bottle (i.e. a waste liquid bottle) for receiving waste liquid is arranged on the sample application box 8, a liquid suction ball 82 is arranged on the bottle mouth of the reagent bottle, a part of solution to be tested is injected into the sample application needle 6 through a pump while the sample application needle 6 moves, a control mechanism controls the sample application needle 6 to contact the liquid suction ball 82, and liquid drops on the needle head of the sample application needle 6 are removed before sample application. The pipette 82 is used to remove needle droplets. The liquid absorbing ball 82 may be a cotton ball, or may be made of other materials having liquid absorbing property.

In one embodiment, as shown in fig. 1, the sample application device further comprises a base 1, the base 1 is provided with a sample box 7 and a sample application box 8, the sample box 7 is provided with a plurality of holes for placing reagent bottles, and each hole is provided with one reagent bottle; the control console 2 and the sample application box 8 are oppositely arranged at two sides of the sample box 7; the sample plate 81 is obliquely arranged on the sample box 8; the side of the spot plate 81 close to the liquid suction ball 82 is lower than the side far from the liquid suction ball 82; the reagent bottle with the liquid suction ball 82 at the bottle mouth is close to the sample box 7; from the direction of the control console 2 approaching the sample application box 8 to the direction of the control console far from the sample application box 8, the sample box 7 is sequentially provided with a reagent bottle for containing sample liquid, a reagent bottle for containing air (namely an empty reagent bottle), a reagent bottle for containing cleaning solvent and a reagent bottle for receiving waste liquid; the cleaning solvent in the reagent bottle is used for cleaning the outer wall of the sample application needle 6; the arrangement structure is beneficial to minimizing the stroke of the sample application needle 6, reducing the design requirement on a control program, and rapidly completing the steps of liquid injection, sample application, cleaning and the like.

The sample application needle 6 is connected with a pump for sucking and injecting liquid; the spot plate 81 is a silicone plate. In an embodiment, as shown in fig. 4 and 5, the connector 500 further comprises a connector 500, the connector 500 comprising a clamping sleeve 53 connected to the sample application needle 6 and a clamping table 51 arranged on the Z-axis, the clamping table 51 having a recess for clamping the clamping sleeve 53. One side of the lowest level of the sample plate 81 is slightly higher than the bottle mouth of each reagent bottle on the sample box 7, which is helpful for reducing the stroke of the sample application needle 6 and improving the sample application efficiency.

In one embodiment, as shown in fig. 3, the sample box 8 is provided with a slot 83 for placing the sample plate 81, so as to prevent the sample plate 81 from sliding downward.

In one embodiment, the clamping table 51 is hinged with a clamping piece 52 for pressing the clamping sleeve 53 to the groove of the clamping table 51.

In an embodiment, the connecting member 500 further includes a locking member 522, the locking member 52 is provided with a locking hole 521, the clamping table 51 is provided with a concave hole (not shown) matching the locking hole 521, and the locking member 522 penetrates the locking hole 521 and extends into the concave hole of the clamping table 51 to lock the locking member 52. The locking member 522 may be a bolt, the locking hole 521 and a corresponding concave hole on the clamping table 51 are screw holes matched with the bolt, and the locking member 522 is in threaded connection with the locking hole 521 and the corresponding concave hole on the clamping table 51 to lock the locking member 52 to the clamping table 51. This structure can realize locking the stopper 52 to the chuck 51 fast, effectively avoid sample application needle 6 to rock in the removal in-process simultaneously.

In another embodiment, the locking member 52 is a magnetic member, and is magnetically attached to the clamping table 51, so that the locking member 52 can be quickly locked to the clamping table 51. In another embodiment, the latch 52 may also be bonded to the catch 51 by an adhesive. This structure can lock the stopper 52 to the chuck 51 as well.

The control mechanism controls the movement of the sample application needle 6. The control mechanism can comprise a programmable controller (Programmable Logic Co ntroller, PLC), an embedded system of a singlechip.

In one embodiment, the sample application needle 6 is suspended vertically above each reagent bottle and the sample application plate 81, and enters the corresponding reagent bottle or the sample application plate 81 under the control of the control mechanism if necessary, so as to finish sample application.

In an embodiment the sample application needle 6 has a hollow channel which communicates to the liquid supply system, which hollow channel may be arranged vertically for easy pipetting and pipetting. The sample application needle 6 can be connected to the liquid supply system through the hollow channel of the connector 500, and liquid suction, liquid injection and the like can be performed when needed.

In one embodiment, the sample application needle 6 is suspended vertically above each reagent bottle and sample application plate 81 to facilitate liquid injection and sample application.

According to a second aspect, in an embodiment, there is provided a method of collecting fractions comprising liquid collection using the spotting device of the first aspect, the control mechanism controlling the spotting needles to inject the fractions to be collected into the respective empty reagent bottles. The fraction to be collected may be a fraction obtained after chemical reaction, and the fraction collected to the reagent bottle may be used for spot plate, detection, separation, or the like.

In one embodiment, if the collected liquid is used for spotting, a portion of the reagent bottles may be removed, and empty reagent bottles required for the spotting process, reagent bottles for containing the cleaning solvent, may be placed.

According to a third aspect, in an embodiment, as shown in fig. 1, 2, 3, a spotting device is provided comprising a spotting needle 6 for collecting a sample, sucking the sample and spotting;

a control mechanism for controlling the movement of the sample application needle 6;

a plurality of reagent bottles which are respectively used for containing solution to be tested, cleaning solvent, air and waste liquid;

a sample plate 81 for sample application by the sample application needle 6;

a liquid suction ball 82 for removing the liquid drop from the needle head of the sample application needle 6;

the control mechanism controls the sample application needle 6 to finish liquid injection, sample suction, sample application and cleaning;

or, the reagent bottles are empty bottles for containing the liquid to be collected, the control mechanism controls the sample application needle 6 to inject the liquid to be collected into each empty reagent bottle, the liquid to be collected can be fractions obtained after chemical reaction, and the liquid collected into the reagent bottles can be used for spot plates, detection, separation and the like.

In one embodiment, the automatic sample application method provided by the invention has the advantages of high sample application speed and accurate sample application amount, and can accelerate the detection and analysis of chemical reaction, save time and cost and accelerate the development process.

In one embodiment, the invention designs an instrument device integrating the functions of automatically collecting reaction liquid and automatically spotting, and develops an automatic spotting method and a thin layer chromatography method based on the automatic spotting.

In one embodiment, the device of the invention can automatically collect the reaction liquid and automatically sample the sample, can liberate both hands to save labor, avoid artificial errors and protect operators from toxic reagents.

In one embodiment, the thin layer chromatography method based on automatic sample application provided by the invention can be used for quantifying a required product, and is helpful for accurately tracking the progress of chemical reaction.

Example 1

The automatic reaction liquid collecting and automatic spotting device of this embodiment, as shown in fig. 1 to 5, comprises a three-axis structure (X, Y, Z axis), a programmable controller (PLC), a spotting needle 6, a sample cartridge 7, and a spotting cartridge 8. The material of the sample application needle 6 may be metal.

The programmable controller is used for controlling the motion of the triaxial. The programmable controller is electrically connected with a hand touch screen 21, and the operation of the motor can be controlled by clicking the hand touch screen 21 so as to control the movement of the three shafts; the three-axis motion can be controlled by computer software through the USB line connection computer.

The sample application needle 6 is fixed on the connector 500, and the connector 500 is fixed on the Z-axis clamping groove, so that the sample application needle can move along the Z-axis in the directions X, Y, Z. In addition, the sample application needle 6 and the connection member 500 have a hole in the center, the connection member 500 connects the sample application needle 6 and a pipeline at the end of the flow chemical system (i.e., the liquid path system), and the liquid of the flow chemical system flows through the pipeline, the connection member 500 and the sample application needle 6 and flows out from the needle tip.

Referring to fig. 2, the sample cartridges 7 and the sample application cartridges 8 are placed in the clamping grooves of the instrument bottom plate, two sample cartridges 7 are provided, 48 (8 rows, 6 columns) holes are provided on each sample cartridge 7, and 48 sample bottles can be placed. In this embodiment, the sample bottle is 48, or other amounts, such as 4*6, 8×12, etc., can be made as needed, and the sample bottle of this embodiment has a capacity of 2mL. The sample application box 8 is provided with a row of 8 holes which are arranged in a straight line and used for placing sample bottles (also called reagent bottles), a certain interval is reserved between the adjacent sample bottles, a small group of cotton for wiping a needle is placed at the bottle mouth of each sample bottle, and the sample application box 8 is also provided with a clamping groove 83 for placing a silica gel plate.

The centers of 48 sample bottles in 48 holes on the sample box correspond to 48 two-dimensional coordinates (X and Y), a certain coordinate is clicked on a PLC screen or control software, the sample application needle 6 can move to the upper part of the corresponding position along with the Z axis, and the sample application needle can extend into a certain depth in the bottle mouth according to the requirement.

Referring to fig. 2 and 3, the centers of 8 sample bottles in 8 holes on the sample application box correspond to 8 two-dimensional coordinates (X and Y), a coordinate is clicked on a PLC screen or control software, and the sample application needle can move to the upper side of the corresponding position along with the Z axis and move down the Z axis to make the sample application needle contact the cotton mass. The sample application box 8 is provided with a row of sample application positions C31, the distance between the C31 and the lower edge of the silica gel plate is 1-1.5 cm, the distance between the C31 and the silica gel plate is adjustable, a row of sample application positions are provided with 8 positions and corresponding X, Y coordinates, a certain coordinate is clicked on a PLC screen or control software, a sample application needle can move to the upper part of the corresponding position along with a Z axis, the Z axis is downwards moved to enable the sample application needle to contact the silica gel plate to finish the sample application process, the time of the sample application needle contacting the silica gel plate can be flexibly adjusted, and the quantity of sample application can be flexibly adjusted.

As shown in fig. 4 and 5, the connector 500 is further included, and the connector 500 includes a card sleeve 53 connected to the sample application needle 6 and a card table 51 disposed on the Z axis, and the card table 51 has a groove for clamping the card sleeve 53.

The clamping table 51 is hinged with a clamping piece 52 for pressing the clamping sleeve 53 to the groove of the clamping table 51.

The connecting piece 500 further includes a locking piece 522, the locking piece 52 is provided with a locking hole 521, the clamping table 51 is provided with a concave hole (not shown) matched with the locking hole 521, and the locking piece 522 penetrates the locking hole 521 and extends into the concave hole of the clamping table 51 to lock the locking piece 52. The locking member 522 may be a bolt, the locking hole 521 and a corresponding concave hole on the clamping table 51 are screw holes matched with the bolt, and the locking member 522 is in threaded connection with the locking hole 521 and the corresponding concave hole on the clamping table 51 to lock the locking member 52 to the clamping table 51. This structure effectively prevents the locking member 52 from loosening and affecting the operation of the print needle 6.

The automatic reaction liquid collecting and automatic sample application device of the embodiment has two functions, namely an automatic reaction liquid collecting function and an automatic sample application function.

Referring to fig. 1, 2 and 3, when the automatic reaction solution collecting function is used, the sample needle 6 is connected to the outlet of the flow chemistry system through the connector 500, and an empty reagent bottle for receiving the reaction solution is placed in the sample box 7. The liquid of the flow chemical system flows through the pipeline, the connecting piece and the sample application needle 6, flows out from the needle point to an empty reagent bottle at a set position, collects the reaction liquid within a time set by software, and moves to the next position along with the triaxial structure after the set time is over. The two sample boxes 7 can be provided with 96 empty reagent bottles, namely 96 reaction liquid samples can be collected at one time. In the process of moving the sample application needle 6 from one reagent bottle to the next, the XYZ three-axis movement mode is as follows: the Z axis moves upwards to enable the sample application needle 6 to be out of the bottle mouth, the XY axis moves to enable the sample application needle 6 to reach the position right above the next reagent bottle, and the Z axis moves downwards to set depth to enable the sample application needle 6 to be in the bottle mouth. If the liquid collected in each reagent bottle is used for subsequent sample application, the liquid to be collected can be injected into the reagent bottles of the first to three rows (A1-A3), or after all the reagent bottles finish liquid collection, the reagent bottles of the fourth row (A4), the fifth row (A5) and the sixth row (A6) are removed, the reagent bottles are replaced, then the cleaning solvent is injected into the empty reagent bottles of the fifth row (A5) for cleaning the outer wall of the sample application needle during subsequent sample application, and the fourth and the sixth rows of reagent holes are kept in an empty state and are respectively used for sucking air and receiving waste liquid by the subsequent sample application needle 6.

Referring to fig. 1, 2 and 3, when the automatic spotting function is used, the spotting needle 6 is connected to a pump having an injection and suction function through a connector 500 and a pipeline, and the pump is filled with acetonitrile solvent for cleaning the pipeline and the inner wall of the spotting needle. The first to three rows (A1 to A3) of reagent bottles of the sample box 7 are filled with the solution to be spotted, the fourth row (A4) is used for sucking air by placing empty reagent bottles, the fifth row (A5) of reagent bottles is filled with acetonitrile for cleaning the outer wall of the needle head, and the sixth row (A6) of reagent bottles is used for receiving waste liquid. The cotton ball is placed to the reagent bottleneck of sample application box 8 first row (C1) and is used for wiping off syringe needle liquid drop, places the silica gel board that needs the sample application in the draw-in groove of sample application box 8, and the sample application position sets up in position C3 (down along 1cm from the silica gel board), and the sample application position is sharp and arranges, has 8 sample application positions on every silica gel board, and the direction of arranging of sample application position is parallel with the reagent bottle direction of arranging on the sample application box 8. The sample application is only exemplified here, and more sample application boxes and corresponding silica gel plates can be arranged, so that sample application of more samples can be finished at one time, and the sample application efficiency is improved.

The spotting process will be described in detail below by taking a sample of the reaction solution at the A31 position in the first column as an example. Referring to FIGS. 3 and 8, in the first step, the sample needle 6 is moved to the reagent bottle A61 by a triaxial structure, and 50. Mu.L acetonitrile (100. Mu.L/min) is pumped to wash the inner wall of the sample needle; secondly, the sample application needle 6 moves to a reagent bottle A51 through a triaxial structure, and moves downwards by 1.2-1.8 cm to enable the sample application needle 6 to be immersed in acetonitrile solvent to clean the outer wall of the sample application needle; third, the sample application needle 6 is moved to the reagent bottle A41 through the three-axis structure, and 20. Mu.L of air (100. Mu.L/min) is pumped up as a gas interval; fourth, the sample application needle 6 moves to the reagent bottle A31 through a triaxial structure, and moves downwards by 1.2-1.8 cm to enable the sample application needle part to be immersed into the solution to be applied (namely the solution to be tested) so as to absorb the solution, and a pump absorbs 25 mu L of the solution for applying the sample; fifthly, the sample application needle 6 moves to the reagent bottle A61 through a triaxial structure, a pump injects 20 mu L of clearance for emptying an injection-to-suction mode, the pump has gear clearance when the suction is converted into the injection mode, and the emptying is to remove the clearance when the mode is switched; the clearance of the pump of the embodiment is about 15 mu L, and the 20 mu L is emptied to ensure that the liquid to be detected is conveyed to the needle head, and the liquid can be normally discharged when the injection mode is started again; sixth, the sample application needle 6 is moved to C11 through the three-axis structure, the sample application needle is moved (from the reagent bottle a61 to the reagent bottle C11) while the pump injects the solution to be tested (the sample application liquid is not dropped out from the needle port) at a speed of 40 μl/min, so that the action of the pump and the action of the three-axis are sometimes crossed for saving time, that is, the pump may be inhaling or injecting during the movement of the three-axis, here, injecting, and the sample application needle 6 is moved to the next position after contacting the cotton mass 3s (such that the needle head liquid drop is removed); seventh, the sample application needle 6 moves to a sample application position C31 through a triaxial structure, the sample application needle 6 moves while the pump injects the solution to be tested at a speed of 40 mu L/min (the sample application liquid does not drop out from a needle opening), and the sample application needle 6 contacts the silica gel plate 7s to finish sample application and then moves to the next position; eighth, the sample needle 6 was moved to the reagent bottle A61 by a triaxial structure, and the inner wall of the sample needle was washed by pumping 50. Mu.L of acetonitrile (100. Mu.L/min). Thus, the sample application of the reagent bottle A31 sample was completed by the above eight steps, which takes 120 seconds. The sample application of other samples to be tested is the same as that of the sample application box 8 in the embodiment, and 1 sample application plate 8 can be placed on each sample application box, and the sample application plate 8 is specifically a silica gel plate. 2 silica gel plates can be placed on the two sample application boxes, and 8 sample application positions are arranged on each silica gel plate, so that 16 samples can be applied at a time; a new sample application can be started by replacing a new silica gel plate; in addition, the whole spotting process is automatically controlled by control software. And, the whole sample application process does not need to replace the sample application needle 6, thereby obviously improving the sample application efficiency.

After the sample application of the silica gel plate is completed, the silica gel plate is put into a developing agent (set according to the property of the product) with specific polarity to climb the plate, taken out when the front edge of the developing agent is 0.5 cm to 1cm away from the upper edge of the silica gel plate, and baked in a baking oven at 100 ℃ for 4 minutes to 6 minutes to remove the developing agent. The silica gel plate is then placed in a gel imaging analyzer to take a photograph of the silica gel plate (ultraviolet light), or after developing color by other means such as iodine fumigation. Finally, the photo is processed by software, and the concentration of the product to be detected can be calculated by comparing the gray value of the product with a standard curve, so that the product can be quantified.

The standard curve was established as follows:

the sample solution used in this example was p-cyanobenzamide. The chemical formula of the p-cyanobenzamide is as follows:

the specific method comprises the following steps: 6 solutions of paracyanobenzamide with different concentrations were prepared, the concentrations being respectively: 0.005, 0.01, 0.02, 0.03, 0.04, 0.05mmol/mL, then spotted using the apparatus and method described above, six solutions were spotted onto the same silica gel plate, and then the silica gel plate was placed in ethyl acetate: n-hexane=5: 2 (volume ratio) in a developing agent, baked at 100 ℃ for 5 minutes, and then placed in a gel imaging analyzer to take a photograph of a silica gel plate (254 nm ultraviolet light), the photograph of the spot plate being shown in fig. 9.

The photograph shown in fig. 9 is processed by using the picture processing software to obtain the gray value of each point, a compound concentration-sample application gray relation curve is established by combining the corresponding concentration of each point, and the specific view is shown in fig. 10, and as can be seen from the figure, the p-cyanobenzamide solution-sample application gray is in good linear relation, R ² 0.9763. Thus, the concentration and thus the yield can be calculated from the gray scale of the corresponding spotting of paracyanobenzamide in solution.

The calculated yields were tested as follows:

the photocatalytic reaction formula for synthesizing the p-cyanobenzamide from the terephthalonitrile is as follows:

synthesizing p-cyanobenzamide under the condition of visible light catalysis by utilizing terephthalonitrile, synthesizing by utilizing a flow chemical system according to the conditions shown in a reaction formula, automatically collecting reaction liquid after reaction by utilizing the equipment, and then automatically spotting. Finally, the yield of the reaction was calculated to be 84% by comparing the compound concentration-spotting gray scale relationship curve.

The sample application experiment was repeated as follows:

the sample solution used in this example was p-cyanobenzamide at a concentration of 0.02mmol/mL.

The specific method comprises the following steps: preparing 0.02mmol/mL of paracyanobenzamide solution, then spotting by the equipment and the method of the example 1, repeating the spotting for six times, spotting the six times on the same silica gel plate, and marking as a liquid suction ball group; the non-wicking ball set removed the wicking balls during spotting, with the other conditions being the same as the set with wicking balls. After spotting was completed, the silica gel plate was placed in ethyl acetate: n-hexane=5: 2 (volume ratio) in developing agent, baking at 100deg.C for 5 min, placing in gel imaging analyzer, taking photo of silica gel plate (254 nm ultraviolet light), processing photo with picture processing software to obtain gray value of each point, and experimental results are shown in Table 1.

TABLE 1 Effect of pipette on repeated spotting experiments

As can be seen from table 1, the standard deviation of the six spotting results is 9.73 without the pipette, and 1.20 with the pipette, indicating that the presence of the pipette significantly reduces the standard deviation of the repeated experiments, improving the reproducibility and accuracy of the spotting experiments. In an embodiment, through the cooperation of control mechanism, triaxial structure (X, Y, Z axle), sample application needle, sample box and sample application box, can realize the function of automatic collection reaction liquid and automatic sample application, avoid artificial error, save time cost protects operating personnel from toxic reagent's poisoning, and sample application is fast simultaneously, and sample application volume is accurate, is favorable to the detection analysis of chemical reaction.

In one embodiment, the desired product can be quantified by an automated spotting method developed, an automated spotting-based thin layer chromatography method, to facilitate accurate tracking of chemical reaction progress.

Those skilled in the art will appreciate that all or part of the functions of the various methods in the above embodiments may be implemented by hardware, or may be implemented by a computer program. When all or part of the functions in the above embodiments are implemented by means of a computer program, the program may be stored in a computer readable storage medium, and the storage medium may include: read-only memory, random access memory, magnetic disk, optical disk, hard disk, etc., and the program is executed by a computer to realize the above-mentioned functions. For example, the program is stored in the memory of the device, and when the program in the memory is executed by the processor, all or part of the functions described above can be realized. In addition, when all or part of the functions in the above embodiments are implemented by means of a computer program, the program may be stored in a storage medium such as a server, another computer, a magnetic disk, an optical disk, a flash disk, or a removable hard disk, and the program in the above embodiments may be implemented by downloading or copying the program into a memory of a local device or updating a version of a system of the local device, and when the program in the memory is executed by a processor.

The foregoing description of the invention has been presented for purposes of illustration and description, and is not intended to be limiting. Several simple deductions, modifications or substitutions may also be made by a person skilled in the art to which the invention pertains, based on the idea of the invention.

Claims (10)

1. A sample application method, which is characterized by comprising the following steps in sequence:

a step of cleaning the inner wall of the sample application needle, wherein a control mechanism controls the sample application needle to move to the upper part of the waste liquid bottle, and a pump connected with the sample application needle pumps cleaning solvent into the sample application needle to clean the inner wall of the sample application needle;

a step of cleaning the outer wall of the sample application needle, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle filled with the cleaning solvent and controls the sample application needle to extend into the reagent bottle filled with the cleaning solvent, the sample application needle at least partially submerges into the cleaning solvent in the reagent bottle, and the cleaning solvent in the reagent bottle is used for cleaning the outer wall of the sample application needle; and/or the control mechanism controls the other flow path to pump a cleaning solvent to the outer wall of the sample application needle so as to clean the outer wall of the sample application needle;

a step of sucking gas or immiscible liquid, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle containing air or immiscible liquid, and the pump controls the sample application needle to suck air as a gas interval or suck liquid immiscible with the solution to be detected as a liquid interval;

A liquid sucking step, wherein the control mechanism controls the sample application needle to move to the position above the reagent bottle containing the sample liquid, controls the sample application needle to extend into the reagent bottle containing the sample liquid, and controls the sample application needle to suck the sample liquid through the pump;

a liquid drop removing step, wherein the control mechanism controls the sample application needle to contact with the liquid suction ball, so that the liquid suction ball wipes off liquid drops of the needle head of the sample application needle and enters the sample application step;

a sample application step, wherein a control mechanism controls a sample application needle to move to the upper part of a sample application plate, controls the sample application needle to contact a sample application position on the sample application plate, and injects a set amount of solution to be tested into the sample application needle through a pump, and returns to the inner wall cleaning step after sample application is finished;

the sample application method is circulated by the six steps, and the sample application of each sample application position on the sample application plate is completed.

2. The spotting method of claim 1, further comprising a step of evacuating the spotting needle by a pump after the pipetting step and before the de-pipetting step, the control mechanism controlling the spotting needle to move over a reagent bottle for receiving waste liquid on the sample cartridge;

after the sample application of each sample application position on the sample application plate is finished, putting the sample application plate into a developing agent to climb the plate, and drying to remove the developing agent on the sample application plate after the climbing plate is finished;

Placing the dried silica gel plate under an ultraviolet lamp or performing iodine fumigation, and then photographing to obtain a silica gel plate photo, and analyzing and calculating according to the photo to obtain the concentration of a product in the solution to be detected;

in the drop removing step, a control mechanism controls a sample application needle to move to the position above a reagent bottle on a sample application box, and controls the sample application needle to contact with a liquid suction ball at the bottle opening of the reagent bottle, when the sample application needle moves, a solution to be detected is injected into the sample application needle through a pump, and after the liquid suction ball wipes off the drop of the sample application needle head, the sample application step is carried out.

3. A spotting method according to claim 1, characterized in that the spotting device used comprises a spotting needle (6) for collecting, sucking and spotting the sample;

a control mechanism for controlling the movement of the sample application needle (6);

the reagent bottles are respectively used for containing a solution to be detected, a cleaning solvent, air or immiscible liquid and waste liquid;

-a spotting plate (81) for spotting by said spotting needle (6);

a pipette (82) for removing the droplets from the needle head of the sample application needle (6) before the sample application;

the control mechanism controls the sample application needle (6) and the pump to finish liquid injection, sample suction, sample application and cleaning;

or the reagent bottles are empty bottles and are used for containing fractions to be collected, and the control mechanism controls the sample application needle (6) to inject the fractions to be collected into each empty reagent bottle.

4. A spotting method according to claim 3, characterized in that the control mechanism comprises a console (2) and an X-axis, a Y-axis and a Z-axis which are arranged on the console (2), wherein the X-axis, the Y-axis and the Z-axis are mutually perpendicular, the spotting needle (6) is fixed on the Z-axis, and the control mechanism controls the spotting needle (6) to do translational movement along the X-axis and the Y-axis and to do linear lifting movement along the Z-axis so as to complete the injection, the suction, the spotting and the cleaning;

the sample application device further comprises a liquid path system communicated with the sample application needle (6), wherein the liquid path system comprises a pump and a pipeline, the pump is used for injecting liquid into the sample application needle (6) or controlling the sample application needle (6) to absorb the liquid, and the pump is communicated to the sample application needle (6) through the pipeline;

the control mechanism is electrically connected to the pump and controls the X-axis, the Y-axis and the Z-axis to move so that the sample application needle (6) can finish liquid injection, sample suction, sample application and cleaning;

or the control mechanism controls the X-axis, Y-axis and Z-axis to move, so that the sample application needle (6) finishes injecting the liquid to be collected into the empty reagent bottle.

5. A deposition method according to claim 3, characterized in that the deposition device further comprises a deposition cartridge (8), the deposition plate (81) being arranged in the deposition cartridge (8), the control means controlling the deposition needle (6) to contact the deposition plate (81) for deposition.

6. A spotting method according to claim 5, characterized in that the spotting box (8) is provided with a reagent bottle for receiving waste liquid, the pipette ball (82) is arranged on the bottle mouth of the reagent bottle, the spotting needle (6) is moved while the solution to be tested is injected into the spotting needle (6) by a pump, and the control mechanism controls the spotting needle (6) to contact with the pipette ball (82) to remove the liquid drop of the needle of the spotting needle (6) before spotting.

7. A spotting method according to claim 3, characterized in that the spotting device further comprises a base (1), the base (1) is provided with a sample box (7) and a spotting box (8), and the sample box (7) is provided with a plurality of holes for placing the reagent bottles;

the control console (2) and the sample application box (8) are oppositely arranged at two sides of the sample box (7);

the sample plate (81) is obliquely arranged on the sample box (8);

the side of the spot plate (81) close to the liquid suction ball (82) is lower than the side far away from the liquid suction ball (82);

the reagent bottle with the liquid suction ball (82) at the bottle mouth is close to the sample box (7);

from the direction of the control console (2) approaching the sample application box (8) to the direction of the control console far away from the sample application box (8), a reagent bottle for containing sample liquid, a reagent bottle for containing air, a reagent bottle for containing cleaning solvent and a reagent bottle for receiving waste liquid are sequentially arranged on the sample box (7);

The cleaning solvent in the reagent bottle is used for cleaning the outer wall of the sample application needle (6);

the sample application needle (6) is connected with a pump for sucking and injecting liquid;

the spot plate (81) is a silica gel plate;

the sample application box (8) is provided with a clamping groove (83) for placing the sample application plate (81).

8. A spotting method according to claim 3, characterized in that the spotting device further comprises a connector (500), the connector (500) comprising a ferrule (53) connected to the spotting needle (6) and a clamping table (51) arranged in the Z-axis, the clamping table (51) having a recess for clamping the ferrule (53);

the clamping table (51) is hinged with a clamping piece (52) for pressing the clamping sleeve (53) to a groove of the clamping table (51);

the clamping piece (52) is a magnetic piece and is tightly attached to the clamping table (51) through magnetism;

the sample application needle (6) is provided with a hollow channel which is communicated with a liquid supply system;

the sample application needle (6) is vertically suspended above each reagent bottle and the sample application plate (81).

9. A method of collecting fractions, characterized in that it comprises the use of the spotting device according to any one of claims 3-8 for fraction collection, the control means controlling the spotting needle (6) to inject the fractions to be collected into the respective empty reagent bottles.

10. A sample application device, characterized by comprising a sample application needle (6) for collecting a sample, sucking the sample and applying the sample;

a control mechanism for controlling the movement of the sample application needle (6);

the reagent bottles are respectively used for containing a solution to be tested, a cleaning solvent, air and waste liquid;

-a spotting plate (81) for spotting by said spotting needle (6);

a pipette (82) for removing the droplets from the needle head of the sample application needle (6) before the sample application;

the control mechanism controls the sample application needle (6) to finish liquid injection, sample suction, sample application and cleaning;

or the reagent bottles are empty bottles and are used for containing fractions to be collected, and the control mechanism controls the sample application needle (6) to inject the fractions to be collected into each empty reagent bottle.

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