A microfluidic liquid chromatography system manufactured by using a full light curing 3D printing technology and a method and application thereof

Abstract

The application discloses a kind of microfluidic liquid chromatography systems manufactured by using full light curing 3D printing technology and method and application thereof, it is related to the full light curing 3D printing technology field of microfluidic liquid chromatography system, the main structure of this microfluidic liquid chromatography system is completely manufactured by light curing 3D printing technology, the mechanical structure and flow path system of microfluidic liquid chromatography system are manufactured using high-resolution full light curing 3D printing technology, wherein the mechanical structure and flow path system include fluid pumping system, sample injection system, flow path management system, gradient system and liquid chromatography module including controllable chromatographic column of microcolumn bed structure, using 3D printing manufacturing instrument parts or even manufacturing instrument itself can effectively reduce instrument system building cost, improve the user customization capability of instrument system.

Description

Technical Field

[0001] This invention relates to the field of photopolymerization 3D printing technology for microfluidic liquid chromatography, and particularly to a method and application of manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology, which can be applied to fields such as analytical chemistry, biological analysis, and portable rapid testing. Background Technology

[0002] 3D printing, also known as additive manufacturing, is a manufacturing technology that constructs solid objects from the bottom up through the interaction of material units, including stacking, assembling, and polymerizing. 3D printing excels at creating objects with complex geometries, offering advantages such as low waste and rapid prototyping. In industrial production, 3D printing is widely used in aerospace, construction, and biomedicine, playing a crucial role in rapid prototyping and cost reduction. Currently, 3D printing is also widely applied in analytical chemistry, providing analytical chemists with solutions for various non-standard parts, assisting in experiments and the operation of experimental instruments. Using 3D printing to manufacture instrument parts, or even the instruments themselves, can effectively reduce the cost of instrument system construction and enhance the user customization capabilities of instrument systems.

[0003] Liquid chromatography (LC), as a separation and analysis technique, achieves the separation of analytes through physicochemical processes such as distribution, diffusion, and adsorption between a solid support containing a stationary phase and a liquid mobile phase. It then performs qualitative analysis based on time-scale resolution. LC can also be coupled with numerous other analytical techniques to achieve powerful and diverse analytical capabilities. LC has become an important analytical technique with extremely wide applications.

[0004] Current liquid chromatography (LC) instruments are primarily modular laboratory instruments, typically comprising a liquid pumping system, injection system, chromatographic column, and detection system. These LC system modules require sophisticated manufacturing processes and the use of multiple materials in their composite production. They also necessitate the integration of numerous flow path management and sensor modules, resulting in high manufacturing costs. Furthermore, users require comprehensive training to effectively operate and maintain these instruments. The large size of LC instruments makes them difficult to deploy outside of fixed locations such as laboratories and hospitals. Their primary operating mode is to be placed in a fixed location to receive samples for testing, lacking portability and on-site detection capabilities. Summary of the Invention

[0005] The main objective of this invention is to provide a method and application for manufacturing a microfluidic liquid chromatography system using all-photocurable 3D printing technology. By manufacturing most of the mechanical structure and flow path system of the microfluidic liquid chromatography system through photocurable 3D printing, the manufacturing cost of the liquid chromatography system is effectively reduced, and control over the structure of the chromatographic column bed is achieved.

[0006] To achieve the above objectives, this invention utilizes high-resolution photopolymerization 3D printing technology to manufacture key liquid chromatography modules, including a fluid pumping system, an injection system, a flow path management system, a gradient system, and a chromatographic column, thereby realizing the full 3D photopolymerization printing manufacturing of the liquid chromatography system. The specific technical solution adopted is as follows:

[0007] This invention provides a method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology, comprising the following steps:

[0008] Step S1: Using Lambert-Beer's law, determine the optimal molding conditions for 3D printing through a normalized light absorption equivalent model;

[0009] Step S2: Draw the model of the 3D printed liquid chromatography system component. Based on the optimal molding conditions optimized in step S1, generate the 3D printed model slice file and import the slice file into the photopolymerization 3D printer for printing.

[0010] Step S3: After cleaning and drying the 3D printed liquid chromatography system component model with pure ethanol, it is then cured by ultraviolet exposure and assembled to form a microfluidic liquid chromatography system.

[0011] As a further optimization of the present invention, the method for determining the optimal forming conditions for 3D printing by adopting the Lambert-Beer law and using a normalized light absorption equivalent model includes:

[0012] Step S101: According to Beer-Lambert law, convert absorbance A into the ratio of light intensity I to penetration depth z:

[0013] ；

[0014] In the formula: I0 is the initial light intensity, and I(z) is the light intensity at the penetration depth z;

[0015] Step S102: Define the light absorption equivalent D required for polymerization to occur under light irradiation as being related to light intensity I and irradiation time t, and obtain the light absorption equivalent D:

[0016] ;

[0017] In the formula: t is the illumination time, h α The feature is the penetration depth;

[0018] Step 103: Normalize the light absorption equivalent with the critical equivalent for the polymerization reaction to obtain the normalized light absorption equivalent Ω model for the photopolymerization 3D printing process:

[0019] ;

[0020] Where: h α With T C h represents the model features. α For the characteristic penetration depth, T C This is the critical polymerization time.

[0021] As a further optimization of the present invention, the 3D printed liquid chromatography system component module includes a liquid chromatography module comprising a fluid pumping system, an injection system, a mobile phase gradient generation system, and a chromatographic column.

[0022] As a further optimization of the present invention, the fluid pumping system is a two-phase peristaltic pump system, and the construction method includes:

[0023] Step A1: The 3D-printed peristaltic pump rotor compresses the silicone tubes placed symmetrically on both sides to achieve phase-complementary peristaltic liquid pumping.

[0024] Step A2: The two phases of liquid flow are converged into one through a 3D-printed three-way pipe to achieve low-pulse peristaltic liquid pumping.

[0025] As a further optimization of the present invention, the chromatographic column is a liquid chromatography column with a controllable microscopic geometry containing a plate array. Different geometries can enable the chromatographic column to provide different chromatographic kinetic performance.

[0026] As a further optimization of the present invention, the injection system includes a face-sealed six-way injection valve, the valve body of which is composed of a stator and a rotor, and the valve body of the six-way injection valve is connected to other structures of the liquid chromatography system through a standard chromatographic interface.

[0027] The specific construction method of the six-way injection valve body is as follows:

[0028] Step B1: Flow channels are manufactured on the rotor and stator respectively using 3D printing, with a large-volume flow channel, called a metering ring, being manufactured on the stator.

[0029] Step B2: The stator and rotor are aligned and pressed together by bearings to form the valve body. The flow channel is sealed by the flat surface formed by 3D printing.

[0030] As a further optimization of the present invention, the mobile phase gradient generation system is a flow path module containing an array of arc-shaped flow channels of decreasing length, used for generating the mobile phase gradient. When the flow channel array is filled with mobile phase A, mobile phase B is injected through the module's inlet. At this time, mobile phase A in the shorter arc-shaped pipes inside the module is discharged and mixes with a small amount of phase B to form the mobile phase entering the chromatographic column. As phase A is gradually discharged from the module, the proportion of mobile phase B gradually increases until it is completely converted into phase B. This fluid exchange process enables automatic mobile phase gradient generation without the need for mechanical structures.

[0031] The present invention also provides a microfluidic liquid chromatography system manufactured using the method described above, including a mobile phase storage bottle, a 3D-printed biphase peristaltic pump, a 3D-printed three-way connector, a 3D-printed surface-sealed six-way valve, a 3D-printed microstructure chromatography column, an Arduino microcontroller, and a DC power supply.

[0032] The 3D-printed biphase peristaltic pump's silicone tube inlet end is inserted into the storage bottle through the cap hole. The 3D-printed biphase peristaltic pump's outlet end is connected to the 3D-printed surface-sealed six-way valve pump interface through a 3D-printed three-way connector. The 3D-printed surface-sealed six-way valve column interface is connected to the 3D-printed microstructure chromatographic column. The chromatographic column outlet end is connected to the detector through a capillary tube. The flow rate of the 3D-printed biphase peristaltic pump is controlled by an Arduino microcontroller with an added CNC expansion board. The 3D-printed biphase peristaltic pump and the Arduino microcontroller are powered by a 12V DC power supply. During gradient separation, a gradient generation module filled with mobile phase can be connected between the six-way valve and the chromatographic column.

[0033] The present invention also provides an application of the microfluidic liquid chromatography system described above in the separation of nanoparticles, using pure water containing 0.05 wt% SDS as the mobile phase, a peristaltic pump flow rate of 30 μL / min, isocratic separation, and a 3D-printed chromatographic column bed structure of orthogonal plate structure with column bed dimensions of 1.0*1.0*150 mm.

[0034] Compared with the prior art, the present invention has the following beneficial effects:

[0035] In this invention, a liquid chromatography module including a fluid pumping system, an injection system, a flow path management system, a gradient system, and a microscopic column bed structure controllable is manufactured by using high-resolution photopolymerization 3D printing technology. This enables rapid prototyping of a microfluidic liquid chromatography system. Using 3D printing to manufacture instrument parts or even the instrument itself can effectively reduce the cost of instrument system construction and improve the user customization capability of the instrument system. Furthermore, different chromatographic separation effects can be obtained by using 3D printed chromatographic columns with different geometries. Attached Figure Description

[0036] Figure 1This is a schematic diagram of the peristaltic pump structure of the present invention;

[0037] Figure 2 This is a schematic diagram of the 3D printed three-way and six-way valve structure of the present invention;

[0038] Figure 3 This is a schematic diagram of the 3D-printed microfluidic liquid chromatography system according to an embodiment of the present invention;

[0039] Figure 4 This is a schematic diagram of the internal structure of the gradient generation module;

[0040] Figure 5 Chromatograms of nanoparticle separation and electron micrographs of cross-sections of 3D-printed chromatographic column beds;

[0041] Figure 6 This is a schematic diagram of a simulation calculation for photopolymerization 3D printing.

[0042] In the diagram: 1. Liquid storage tank; 2. 3D-printed biphase peristaltic pump; 3. 3D-printed tee connector; 4. 3D-printed surface-sealed six-way valve; 5. 3D-printed microstructure chromatographic column; 6. Arduino microcontroller; 7. 12V DC power supply; 8. Gradient generation module. Detailed Implementation

[0043] The present application will now be described in further detail with reference to the accompanying drawings. It should be noted that the following specific embodiments are only used to further illustrate the present application and should not be construed as limiting the scope of protection of the present application. Those skilled in the art can make some non-essential improvements and adjustments to the present application based on the above application content.

[0044] Example:

[0045] Construction of a fully 3D printed microfluidic liquid chromatography system

[0046] (1) Optimization of 3D printing condition modeling ;

[0047] According to Beer-Lambert Law:

[0048] ;

[0049] The relationship between the ratio of absorbance A to light intensity I and the penetration depth z is as follows:

[0050] The light absorption equivalent D required for polymerization to occur under light irradiation is defined as being related to the light intensity I and the irradiation time t, from which the absorption can be obtained.

[0051] ;

[0052] The absorbance equivalent D can be obtained:

[0053] By normalizing the light absorption equivalent with the critical equivalent for the polymerization reaction, we can obtain the normalized light absorption equivalent Ω model for the photopolymerization 3D printing process: ;

[0054] Based on the above model, after measuring the characteristic curve between the film thickness z of the photocurable resin and the exposure time t, the model characteristic quantity h can be determined. α With T C Furthermore, the model calculates whether, under specific 3D printing conditions (e.g., model slice thickness, exposure time per slice, solid layer interval), the printed entity can be generated normally (normalized equivalent Ω greater than 1), and whether the model cavity can be preserved (normalized equivalent Ω less than 1). Figure 6 As shown:

[0055] This model system can be used to optimize 3D printing conditions. For example, using AnyCubic Basic Clear photopolymer 3D printing resin, calculations show that when the model slice thickness is 30 μm and the exposure time per layer is 1 s, the solid part of the printed model can be successfully formed, and the cavity structure larger than 70 μm can be preserved.

[0056] (2) Photopolymerization 3D printing of components for liquid chromatography system

[0057] Complete the modeling of all components of the 3D printed liquid chromatography system using SolidWorks software (as shown below). Figure 1 , 2 4) After converting to STL format, according to the optimal printing conditions optimized in (1), the 3D printing model slice file is generated in Photon Workshop software. The slice file is then imported into AnyCubic MONO 4K LCD photopolymer 3D printer for printing. AnyCubic Basic Clear photopolymer 3D printing resin is used for printing, and AnyCubic Basic Black resin is used for 3D printing chromatographic columns.

[0058] (3) Post-processing and assembly of components of liquid chromatography system

[0059] After the printed liquid chromatography system components are shaken and cleaned in pure ethanol solution for 10-30 minutes, the surface of the parts is cleaned with ethanol spray. The parts are then placed in a UV exposure chamber and cured with 405 nm UV light for 7 minutes. Finally, the components are rinsed with pure water and air-dried. Individual modular parts can be fixed together with screws (commercial metal screws or 3D-printed screws can be used) to form functional independent modules (e.g., a peristaltic pump module is formed by...). Figure 1The components shown are assembled with the 42 stepper motor using screws. Connections between individual modules can be made via standard liquid chromatography threaded interfaces (flat head 1 / 16 thread) (e.g., Figure 4 The diagram shows a 3D-printed three-way interface and a 3D-printed surface-sealed six-way valve 4 (with an interface on top). The chromatographic column is connected to the detector via a 250 μm ID capillary tube. The 42 stepper motors driving the peristaltic pump are connected to an Arduino microcontroller 6 equipped with a CNC expansion board via stepper motor cables. The peristaltic pump is controlled by the Arduino microcontroller 6 connected to a 12 V DC power supply 7.

[0060] like Figure 3 As shown, the inlet end of the biphase peristaltic pump 1 is inserted into the storage bottle 1 through the cap hole. The outlet end of the biphase peristaltic pump 1 merges the two flow paths into one through a 3D-printed T-connector 3 to reduce flow pulses in the peristaltic pump. The outlet end of the T-connector is connected to the pump interface of a 3D-printed surface-sealed six-way valve 4. The column interface of the six-way valve is connected to a 3D-printed microstructure chromatographic column 5. The outlet end of the chromatographic column is connected to the detector through a capillary tube. The flow rate of the peristaltic pump 2 is controlled by an Arduino microcontroller with an added CNC expansion board. The peristaltic pump and the microcontroller are powered by a 12V DC power supply 7. When gradient separation is required, a gradient generation module 8 filled with mobile phase can be connected between the six-way valve 4 and the chromatographic column 5.

[0061] Application of nanoparticle separation in fully 3D printed microfluidic liquid chromatography systems

[0062] Polystyrene-acrylic acid nanoparticles with particle sizes of 50 nm and 200 nm were dispersed in pure methanol to prepare dispersions with a concentration of 20 mg / mL. These dispersions were mixed at a volume ratio of 1:1 to prepare the test sample. The nanoparticle dispersion sample was drawn into a syringe, which was then connected to the sample end of a six-way valve. The sample was injected into the six-way valve injection position to fill 10 μL of the quantitative structure. The six-way valve was then switched to the separation position to begin chromatographic separation. Chromatographic separation conditions: pure water containing 0.05 wt% SDS was used as the mobile phase; the peristaltic pump flow rate was 30 μL / min; isocratic separation was performed; and the 3D-printed chromatographic column bed structure was an orthogonal plate structure (as shown below). Figure 5 (As shown in the electron microscope image), the column bed dimensions are 1.0*1.0*150 mm. The separation results are as follows. Figure 5 As shown, the present invention can separate two types of nanoparticles with different particle sizes within 6 minutes by utilizing the difference in migration speed of nanoparticles with different particle sizes, and different chromatographic separation effects can be obtained by using 3D printed chromatographic columns with different geometries.

[0063] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of this invention is defined by the appended claims and their equivalents.

Claims

1. A method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology, characterized in that: Includes the following steps: Step S1: Using Lambert-Beer's law, determine the optimal molding conditions for 3D printing through a normalized light absorption equivalent model; Step S2: Draw the model of the 3D printed liquid chromatography system component. Based on the optimal molding conditions optimized in step S1, generate the 3D printed model slice file and import the slice file into the photopolymerization 3D printer for printing. Step S3: After cleaning and drying the 3D printed liquid chromatography system component model with pure ethanol, it is then cured by ultraviolet exposure and assembled to form a microfluidic liquid chromatography system. Methods for determining the optimal forming conditions for 3D printing using the Lambert-Beer law and a normalized light absorption equivalent model include: Step S101: According to Beer-Lambert law, convert absorbance A into the ratio of light intensity I to penetration depth z: ； In the formula: I0 is the initial light intensity, and I(z) is the light intensity at the penetration depth z; Step S102: Define the light absorption equivalent D required for polymerization to occur under light irradiation as being related to light intensity I and irradiation time t, and obtain the light absorption equivalent D: ； In the formula: t is the illumination time. The feature is the penetration depth; Step 103: Normalize the light absorption equivalent with the critical equivalent for the polymerization reaction to obtain the normalized light absorption equivalent Ω model for the photopolymerization 3D printing process: ； In the formula: With T C For model features, For the characteristic penetration depth, T C This refers to the critical polymerization time. Based on the normalized absorbance equivalent Ω model, after measuring the characteristic curve between the film thickness z of the photocurable resin and the exposure time t, the model characteristic quantities are determined. With T C The normalized absorbance equivalent Ω model was used to calculate whether the printed entity could be generated normally and whether the model cavity could be retained under specific 3D printing conditions. The specific 3D printing conditions include the model slice thickness, the exposure time of each slice, and the interval between the solid layers.

2. The method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology according to claim 1, characterized in that: The 3D-printed liquid chromatography system component modules include a liquid chromatography module comprising a fluid pumping system, an injection system, a mobile phase gradient generation system, and a chromatographic column.

3. The method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology according to claim 2, characterized in that: The fluid pumping system is a two-phase peristaltic pump system, and its construction method includes: Step A1: The 3D-printed peristaltic pump rotor compresses the silicone tubes placed symmetrically on both sides to achieve phase-complementary peristaltic liquid pumping. Step A2: The two phases of liquid flow are converged into one through a 3D-printed three-way pipe to achieve low-pulse peristaltic liquid pumping.

4. The method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology according to claim 2, characterized in that: The chromatographic column is a liquid chromatography column with a controllable microscopic geometry containing a plate array.

5. A method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology according to claim 2, characterized in that: The injection system includes a face-sealed six-way injection valve. The valve body of the six-way injection valve is composed of a stator and a rotor. The valve body of the six-way injection valve is connected to other structures of the liquid chromatography system through a standard chromatographic interface. The specific construction method of the six-way injection valve body is as follows: Step B1: Flow channels are manufactured on the rotor and stator respectively using 3D printing, with a large-volume flow channel, called a metering ring, being manufactured on the stator. Step B2: The stator and rotor are aligned and pressed together by bearings to form the valve body. The flow channel is sealed by the flat surface formed by 3D printing.

6. A method for manufacturing a microfluidic liquid chromatography system using all-photopolymerization 3D printing technology according to claim 2, characterized in that: The mobile phase gradient generation system is a flow path module containing an array of arc-shaped flow channels with decreasing lengths.

7. A microfluidic liquid chromatography system manufactured using any one of claims 1-6, characterized in that, The components include a mobile phase storage bottle, a 3D-printed biphase peristaltic pump, a 3D-printed three-way connector, a 3D-printed surface-sealed six-way valve, a 3D-printed microstructure chromatographic column, an Arduino microcontroller, and a DC power supply. The 3D-printed biphase peristaltic pump's silicone tube inlet end is inserted into the storage bottle through the cap hole. The 3D-printed biphase peristaltic pump's outlet end is connected to the 3D-printed surface-sealed six-way valve pump interface through a 3D-printed three-way connector. The 3D-printed surface-sealed six-way valve column interface is connected to the 3D-printed microstructure chromatographic column. The chromatographic column outlet end is connected to the detector through a capillary tube. The flow rate of the 3D-printed biphase peristaltic pump is controlled by an Arduino microcontroller with an added CNC expansion board. The 3D-printed biphase peristaltic pump and the Arduino microcontroller are powered by a 12V DC power supply. During gradient separation, a gradient generation module filled with mobile phase is connected between the six-way valve and the chromatographic column.

8. An application of the microfluidic liquid chromatography system as described in claim 7 in the separation of nanoparticles, characterized in that: Using pure water containing 0.05 wt% SDS as the mobile phase, a peristaltic pump flow rate of 30 μL / min was employed for isocratic separation. The 3D-printed chromatographic column bed structure was an orthogonal plate structure. The column bed specifications were as follows. .

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