CN104096526B - A kind of perfluoropolymer microreactor and its application - Google Patents

Abstract

The invention provides a kind of (per) fluoropolymer microreactor and application thereof, this microreactor has the composite construction that perfluor cover plate, perfluorinated elastomer film and perfluor substrate stack gradually bonding, and described composite construction has micro-air valve.(per) fluoropolymer microreactor provided by the invention can adapt to number of chemical reaction condition, and be suitable for the solid-state chemical reaction method reaction of multicomponent circulation, reaction efficiency is high, and reagent dosage is few, and cost of manufacture is low, and can realize the control of automation course of reaction.

Description

A kind of perfluoropolymer microreactor and its application

technical field

The invention relates to the technical field of microfluidic chips and microreactors, in particular to a perfluoropolymer microreactor and its application in solid-phase organic synthesis.

Background technique

Microreactor refers to a micro chemical reaction system manufactured by micromachining or precision machining technology, and its reaction interface scale is on the order of submicron to submillimeter. Microreactor has the characteristics of large specific surface area, fast mass and heat transfer rate, low reagent consumption, easy large-scale integration and high-throughput reaction. A model of success in the field of chemistry.

Since chemical reactions often require the use of strong acids and bases, various organic solvents, etc., and may also require reaction environments such as heating and pressure, the material selection and processing technology of the microreactor are subject to many restrictions. Based on specific chemical reaction conditions, suitable materials and processing methods can be selected to design specific microreactors, thereby reducing raw material or energy consumption and improving production efficiency. For example, a fast constant temperature microreactor disclosed in CN101696929A can provide a constant temperature microreaction environment. However, the series of microreactors disclosed in CN102240535A, CN102247787A and the like with different structures can effectively mix the reaction raw materials. At present, microfluidic chip reactors for solid-phase peptide synthesis based on hard materials such as glass have also been reported (eg: Wanget.al, LabChip, 2011, 11, 929-935, CN102527306A).

However, the above studies are only optimized for a specific environment, and their microreactors can only adapt to a specific reaction environment. At present, the research on general-purpose microreactors that can adapt to various reaction environments is still a difficult problem. Because it requires the material of the microreactor to have almost harsh chemical resistance, ordinary polymer materials and metal materials are not competent, and inert metals such as gold and platinum are well tolerated, but they are expensive and difficult to process into microreactors. device. At present, the polymer material with the strongest chemical resistance is recognized as a perfluoropolymer material whose side chain is completely replaced by fluorine atoms, and the trade name is "Teflon". Microreactors based on perfluorinated or slightly less resistant fluorinated polymer materials have also been reported, but their performance or application range is still limited. Studies have reported a microfluidic chip made of perfluoropolyether (PFPE), which can work well in a variety of organic solvent atmospheres, and the chip can perform organic synthesis of DNA (Huanget.al , LabChip, 2007, 7, 24-26). But the main problem with this material is that it is too expensive, and its synthesis requires complex equipment and processes. Although the price of commercial perfluoropolymer materials such as polytetrafluoroethylene is low, due to technical and other reasons, except for a small amount of research exploration (Renet.al, Proc.Natl.Acad.Sci.U.S.A., 2011, 108, 8162-8166), there are no reports of mature perfluoropolymer microreactors.

Contents of the invention

The purpose of the present invention is to overcome the defect of prior art, provide a kind of perfluoropolymer microreactor and application thereof, described microreactor can work under the environment of various complex chemical reaction, can but not limited to be used in Multi-step cyclic solid-phase chemical synthesis reactions, such as peptide synthesis reactions.

To achieve the aforementioned object, the present invention provides a perfluoropolymer microreactor, wherein the microreactor comprises: a perfluoro cover sheet, a perfluoroelastic film and a perfluorosubstrate;

Wherein, the perfluorinated cover sheet includes: a main liquid flow pipeline with a reaction chamber, a main sample inlet and a product or waste liquid outlet located at both ends of the main liquid flow pipeline, and liquid flow branch pipes located on both sides of the main liquid flow pipeline Road, the reaction chamber has a carrier binding structure, one end of the liquid flow branch pipeline communicates with the liquid flow main pipeline, and the liquid flow branch pipeline has a liquid branch inlet, and in the liquid flow The branch pipeline is provided with a flow channel disconnection in the valve area, and the carrier restraint structure is used to control the solid carrier in the reaction chamber not to flow out of the reaction chamber, but to allow the liquid in the reaction chamber to flow out of the reaction chamber;

The perfluorinated substrate includes: a gas flow branch pipeline, the gas flow branch pipeline has a gas branch inlet and a valve area, and the number of the gas flow branch pipeline is the same as that of the liquid flow branch pipe in the perfluorinated cover sheet The number of roads is the same;

The perfluoroelastic film has the function of controlling the on-off of the liquid flow branch pipeline according to the air pressure;

Among them, the perfluorinated substrate, the perfluoroelastic film and the perfluorinated cover are laminated and bonded in sequence, and after bonding, the valve area of the perfluorinated cover is disconnected, the valve area of the perfluorinated substrate, and The perfluoroelastic film between them forms a micro-air valve, and the air flow branch pipeline corresponds to the liquid flow branch pipeline one by one.

For the present invention, it is preferred that the materials of the perfluoro cover sheet, the perfluoroelastic film and the perfluoro substrate are each selected from polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and one or more of polyfluoroethylene propylene (FEP), preferably tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and/or polyfluoroethylene propylene (FEP), but not limited to here.

In the present invention, the polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and polyperfluoroethylene propylene (FEP) can all be commercially available materials.

For the present invention, it is preferred that the melting point of tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) is 302-310°C, for example, it can be the DuPont

For the present invention, it is preferred that the melting point of polyperfluoroethylene propylene (FEP) is 255-265 ° C, for example, it can be the

In the present invention, the liquid flow branch pipelines located on both sides of the main liquid flow pipeline form a comb-like structure on the perfluorinated cover sheet. And the main liquid flow pipeline includes an upstream main pipeline (located between the reaction chamber and the main sample inlet) and a downstream main pipeline (located between the reaction chamber and the product or waste liquid outlet).

In the present invention, the micro-air valve is a three-layer air valve structure obtained by aligning, laminating and bonding the perfluorinated substrate, perfluorinated elastic film and perfluorinated cover sheet (which can be called stacked bonding). The number is consistent with the number of liquid flow branch pipelines. Since the micro-air valve is formed by the disconnection of the flow channel of the valve area of the perfluorinated cover sheet, the valve area of the perfluorinated substrate and the perfluoroelastic film between them, the micro-air valve includes the valve area of the perfluorinated cover sheet The opening of the flow channel, the valve area of the perfluorinated substrate, and the perfluoroelastic film located between the opening of the flow channel of the valve area and the valve area.

In the present invention, the liquid flow branch pipeline is generally used to feed the reaction raw materials, and because the reactor is generally used for multiple rounds of cyclic reactions, and the reaction raw materials in each round may be different, so generally multiple liquid flow branch pipelines are required.

In the present invention, the air flow branch pipeline is used to control the on-off of the liquid flow branch pipeline, wherein each air flow branch pipeline can separately control the on-off of a liquid flow branch pipeline, and the principle of its control is to pass from Introduce gas into or cut off the gas in the gas flow branch pipeline to act on the micro-air valve to control the on-off of the liquid flow branch pipeline. When the liquid flow branch pipeline is connected, it can be used to feed the reaction raw materials. When the reaction raw material is not needed, the perfluoroelastic film at the micro-gas valve can be relaxed as long as the gas supply is removed, so that the liquid flow branch pipeline disconnect.

In the present invention, preferably, the valve area can completely cover the disconnection of the flow channel of the valve area, that is, the valve area matches the disconnection of the flow channel of the valve area.

It is further preferred that the valve area has a circular structure, and the width of the disconnected part of the flow channel of the valve area (the distance between the two ends of the disconnected liquid flow branch pipeline) is 0.01-5 mm, and the diameter of the valve area is 0.01-5mm, more preferably 0.1-2mm.

In the present invention, there are many types of carrier binding structures, and any carrier binding structure that can meet the aforementioned requirements of the present invention can be used in the present invention. For the present invention, in order to facilitate processing, it is preferable that the carrier binding structure is composed of 1-50 groups , preferably composed of 3-6 sets of dams, and each set of dams is formed on the bottom of the perfluorinated substrate, and there is a gap between each set of dams and at least one side wall of the reaction chamber. For the present invention, it is further preferred that two adjacent groups of dams are arranged in a staggered manner. Among them, it is further preferred that the width of each group of dams is 0.01-1 mm, more preferably 0.05-0.2 mm; the spacing of each group of dams is preferably 0.1-8 mm, more preferably 0.5-5 mm, and adjacent dams are arranged in a staggered manner.

In the present invention, since the reaction chamber has a carrier binding structure, the solid carrier in the reaction chamber cannot flow out of the reaction chamber, but the liquid can flow out of the reaction chamber.

In the present invention, preferably, the number of liquid branch pipelines of the perfluorinated cover sheet is 2-60, preferably 6-16, and the number is equally distributed on both sides of the main liquid flow pipeline.

In the present invention, the thickness of the perfluoroelastic film is preferably 0.01-0.1 mm, more preferably 0.020-0.05 mm.

In the present invention, the main sample inlet, the liquid branch sample inlet and the gas branch inlet can all transport samples through air pressure.

In the present invention, the micro-air valve can be switched on and off through software control.

The microstructure on the perfluorinated cover sheet and perfluorinated substrate of the microreactor of the present invention can adopt methods such as mechanical milling method, chemical etching method, plasma etching method, hot embossing method, laser ablation method, etc. to make.

As mentioned above, in the present invention, the perfluorinated cover sheet and perfluorinated substrate having the aforementioned microstructure can be formed by the method of the prior art, specifically, for example, it can be formed according to the following steps:

Obtain the required stainless steel male pattern through outsourcing processing (choose according to the required pattern); place a perfluorinated plate (polytetrafluoroethylene, tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer) on the stainless steel male pattern One or more of polyfluoroethylene propylene), hot embossing is carried out under the condition of heating and pressing, the hot embossing temperature is 240-300°C, preferably the hot embossing temperature is 260-290°C, and the pressure is applied It is 0.02-0.4Mpa, preferably the applied pressure is 0.1-0.2Mpa. The pressurization time is 1-40 minutes, preferably the pressurization time is 10-20 minutes; after the pressure is removed, the pattern negative mold is obtained on the perfluorinated plate, and the perfluorinated substrate and the perfluorinated cover sheet of the required structure of the present invention are obtained. . But the present invention is not limited thereto.

In the present invention, the perfluorinated cover sheet, the perfluoroelastic film and the perfluorinated substrate are bonded by thermal bonding or a special adhesive. The fluorine cover sheet, the perfluoroelastic film and the perfluorinated substrate are sequentially stacked and bonded.

In the present invention, as mentioned above, the method of the prior art can be used to bond the perfluoro cover sheet, the perfluoroelastic film and the perfluoro substrate, specifically when the method of the prior art is used to form the perfluoro substrate , After the perfluorinated cover sheet, it can be stacked and bonded according to the following steps:

The perfluorinated substrate, perfluorinated cover slip and perfluoroelastic film were carefully cleaned, carefully aligned with the aid of a micro-alignment device, and fixed with stainless steel clamps; the perfluorinated substrate, perfluoro cover slip and perfluoroelastic film The fluoroelastic film and stainless steel fixture are placed in a chip bonding instrument for bonding. The bonding temperature is 240-300°C, the preferred bonding temperature is 240-270°C; the bonding time is 0.5-6 hours, and the preferred bonding time is 2-4 hours. But the present invention is not limited thereto.

In the present invention, the manufacturing method of the microreactor is not particularly limited, for example, it can be manufactured by mechanical processing template and hot embossing method, or it can be made by chemical wet etching template and hot embossing method.

In the present invention, the perfluoropolymer microreactor can be connected with the accessory device through the following method: each liquid branch inlet is connected to the raw material storage device through an infusion pipeline, and each liquid branch inlet can be connected through a sealant or Elastic and corrosion-resistant rubber hose is sealed, and the raw material is fed through a syringe pump or pneumatically driven. Each gas branch inlet is connected to a gas source through a connecting pipeline, and the sealing method is the same as that of each liquid branch inlet. Wherein, the gas source can be switched and controlled by a solenoid valve. The material of the infusion pipeline and gas connection pipeline can be polytetrafluoroethylene.

The present invention also provides the application of the above-mentioned perfluoropolymer microreactor and its accessory devices in the solid-phase synthesis of polypeptides.

The method that the microreactor of the present invention is used for polypeptide solid-phase synthesis is as follows: the solid-phase carrier is passed into the microreactor through the main inlet, and is blocked by the carrier binding structure and stays in the reaction chamber, and the synthesis process is started after cleaning with the reaction solvent . Each round requires successive passes through deprotection reagents and amino acid/coupling reagents. The solvents and deprotection reagents used in the synthesis enter the reaction chamber from the main inlet, and the amino acids/coupling reagents used in each round of synthesis are passed into the synthesis chamber from different liquid branch inlets. Each liquid branch pipeline uses Automatically controlled perfluorinated pneumatic film micro-air valve control to prevent liquid mixing between pipelines to prevent cross-contamination. After the synthesized peptide is cleaved, it is received from the product collection port, verified by mass spectrometry, and its purity is identified by reversed-phase liquid chromatography.

The perfluoropolymer microreactor provided by the present invention can solve the problems of high price and difficult processing of special perfluoropolymers, and at the same time has the excellent chemical resistance of perfluoropolymer itself, and can adapt to various harsh chemical reaction environments, such as Strong acids and bases, organic solvents, high temperatures, etc., are suitable for but not limited to multi-step cyclic solid-phase chemical synthesis reactions. The micro-reactor integrates multiple perfluorinated pneumatic thin-film micro-gas valve structures, which can be easily automated and integrated control . The microreactor of the invention has the advantages of miniaturization, integration, high efficiency, low cost and environmental friendliness, and has wide application value in the fields of fine chemical industry and biomedicine.

Beneficial effect

1. The present invention uses perfluoropolymer as the microreactor base material, because perfluoropolymer has excellent chemical inertness, it can withstand almost all chemical reagents except molten alkali metal, and can work for a long time at minus 200 Therefore, in addition to the characteristics of large specific surface area, fast mass and heat transfer rate, low reagent consumption, large-scale modular integration and high-throughput reaction, the microreactor produced by using it also has extremely Good universality.

2. The perfluoropolymer microreactor integrates a plurality of perfluoropneumatic film micro-valve structures, which can automatically adjust the opening and closing of the micro-valve according to needs, so as to achieve flexible control of the reaction liquid, integration and flexibility High performance, very suitable for automated operation.

3. The raw materials of the microreactor used are all commercial materials, the manufacturing method is flexible, the manufacturing effect is stable, and the manufacturing cost is greatly reduced.

4. Compared with the prior art, the present invention has the advantages of outstanding performance, good universality, high degree of integration, convenient processing, low cost, etc., and is easy to be automatically controlled, and is very suitable for industrial applications.

Description of drawings

The accompanying drawings are used to provide a further understanding of the present invention, and constitute a part of the description, together with the following specific embodiments, are used to explain the present invention, but do not constitute a limitation to the present invention. In the attached picture:

Fig. 1 is the schematic diagram of the manufacturing process of the perfluoropolymer microreactor with ten liquid flow branch pipelines and ten air flow branch pipelines;

Figure 2 is a schematic diagram of the design specification (top view) of the perfluoropolymer substrate of the perfluoropolymer microreactor with ten gas flow branch pipelines;

Figure 3 is a schematic diagram of the design specification (bottom view) of the perfluorinated cover sheet of the perfluoropolymer microreactor with ten liquid flow branch pipelines;

4 is a schematic diagram of the microstructure of the reaction chamber;

5 is a schematic diagram of the alignment of each layer of the perfluoropolymer microreactor with ten liquid flow branch pipelines and ten gas flow branch pipelines;

Fig. 6 is the complete structural representation of the perfluoropolymer microreactor with ten liquid flow branch pipelines and ten air flow branch pipelines;

Fig. 7 is a schematic diagram of the detailed structure of the micro-air valve.

reference sign

1 Air flow branch pipeline 2 Gas branch inlet

3 valve zones 4 main inlets

5 upstream main line 6 liquid branch inlet

7 Liquid flow branch pipeline 8 Disconnection of flow channel in valve area

9 Reaction chamber 10 Carrier binding structure

11 Product or waste liquid outlet 12 Downstream main pipeline

13 Perfluorinated cover sheet 14 Perfluoroelastic film

15 perfluorinated substrate

detailed description

The specific embodiments described here are only used to illustrate and explain the present invention, not to limit the present invention.

The experimental methods used in the following examples are conventional methods unless otherwise specified.

The materials and reagents used in the following examples can be obtained from commercial sources unless otherwise specified.

Example 1

A perfluoropolymer microreactor with ten liquid flow branch lines and ten gas flow branch lines was fabricated by chemical wet etching template and hot embossing method.

The manufacture process of described perfluoropolymer microreactor is as shown in accompanying drawing 1:

Wherein, step A represents spin-coating photoresist;

Step B represents UV exposure;

Step C represents development, transferring the pattern on the mask to the photoresist;

Step D represents wet etching of stainless steel, thereby obtaining a pattern positive mold on stainless steel;

Step E represents removing the residual photoresist on the stainless steel;

Step F represents hot embossing, transfers the pattern on the stainless steel positive mold to the perfluoropolymer sheet, and then obtains the perfluoro substrate and the perfluoro cover sheet with the microstructure required by the present invention;

Step G represents stacking and bonding, aligning, stacking, and bonding three layers of perfluorinated materials to form a complete microreactor.

The main implementation process is as follows: according to the microreactor of the present invention, the designed pattern is made into a film mask; the photoresist is spin-coated on a polished stainless steel (model: 304) plate, and after covering the film mask, ultraviolet light is carried out. Exposure; the exposed stainless steel (model: 304) plate is developed in a photoresist (photoresist) developing solution to obtain a photoresist micropattern; the stainless steel plate is placed in a stainless steel etching solution for wet etching to obtain a pattern anode Mold, the line depth is 0.05-0.4mm; use photoresist remover to remove residual photoresist and clean; place perfluorinated substrate or perfluorinated cover sheet on the stainless steel male mold (the materials are all tetrafluoroethylene-full Fluoroalkyl vinyl ether copolymer, that is, PFA), hot embossing is carried out under the condition of heating and pressing, the hot embossing temperature is 260-290°C, the applied pressure is 0.1-0.2Mpa, and the pressing time is 15-20 minute. After the pressure is removed, a negative mold with a corresponding pattern is obtained on the perfluorinated substrate or the perfluorinated cover sheet, thereby promptly obtaining the perfluorinated substrate and the perfluorinated cover sheet with the required microstructure of the present invention; Perfluorinated substrate, perfluorinated cover and perfluoroelastic film (perfluorinated substrate and perfluorinated cover are made of DuPont The melting point range is 302-310°C, and the material of the perfluoroelastic film is DuPont Company of the United States. Melting point range 302-310°C) After careful cleaning, carefully align with the aid of a micro-alignment device, and fix with stainless steel fixtures; place the perfluorinated substrate, perfluorocover, perfluoroelastic film and stainless steel fixtures on the chip The bonding is carried out in a sealing instrument, the bonding temperature is 240-270°C; the bonding time is 2-4 hours.

Figure 2 and Figure 3 show the structure and micropipe size design of the perfluorinated cover sheet and perfluorinated substrate of the fabricated perfluoropolymer microreactor. For the convenience of description, the top view and the bottom view of the microreactor are specially designated here, and each view is in accordance with this designation: the complete perfluoropolymer microreactor is normally placed on a certain plane, and the observer is from top to bottom The view seen by the observer is the top view, and the view viewed from the bottom up by the observer is the bottom view. Figure 2 is a schematic diagram of the structure of a perfluorinated substrate. The reference signs in Fig. 2 are as follows: 1 gas flow branch pipeline, 2 gas branch inlet, 3 valve area. Fig. 2 is a top view, the microstructure of the perfluoropolymer substrate of the normally placed perfluoropolymer microreactor faces upward. Fig. 3 is a schematic diagram of the structure of the perfluorinated cover sheet, which is a bottom view, and the microstructure of the perfluorinated cover sheet of the normally placed perfluoropolymer microreactor faces downward. Among them, the reference signs in Fig. 3 are as follows: 4 main inlet, 5 upstream main pipeline, located upstream of the carrier restraint structure, 6 liquid branch inlet, 7 liquid flow branch pipeline, 8 valve area flow channel disconnection, 9 reaction cavity, 10 carrier binding structure, 11 product or waste liquid outlet. Wherein, the fine structure of the reaction chamber 9 is shown in FIG. 4 , and the downstream main pipeline of the reaction chamber 12 . Perfluorinated cover sheet 13, perfluoroelastic film 14, and perfluorinated substrate 15 are bonded after being fully laminated according to the order shown in Figure 5 to obtain an overall structure, that is, the upper layer of the microreactor is a perfluorinated cover sheet 13, and the microreactor is a perfluorinated cover sheet 13. The structure faces downward, the middle layer is a perfluoroelastic film 14, the lower layer is a perfluoro substrate 15, and its microstructure faces upward. The structure of the microreactor obtained by laminating and bonding the perfluorinated cover sheet 13, the perfluoroelastic film 14 and the perfluoroelastic substrate 15 is shown in FIG. 6 . The micro-valve structure formed by bonding is shown in Figure 7.

The specifications of the perfluoropolymer microreactor and its accessories produced in this example (the accessories refer to: when the microreactor is used for synthesis reactions, the parts that need to be used include connecting pipelines, sealing rings, joints, etc.) are as follows: The size of the fluorine cover is 4cm (length) × 2cm (width) × 2mm (thick); the length of the main liquid flow pipeline is 2.4cm, the width is 400μm, and the depth is 150μm; the length of the liquid flow branch pipeline is 4mm, the width The length of the reaction chamber is 150 μm, the depth is 100 μm; the length of the reaction chamber is 8 mm, the width is 800 μm, and the depth is 150 μm; the single-layer width of the dam part of the reaction chamber is 200 μm, the width of the liquid-passing part is 50 μm, and the distance between the dams is 300 μm , the number of dams is 4 groups; the size of the perfluorinated substrate is 4cm (length) × 2cm (width) × 2mm (thick); the width of the gas flow branch pipeline is 150μm, and the depth is 150μm; The inner diameter of the product or waste liquid outlet and the gas branch inlet is 1mm; all the gas and liquid inlets and outlets are connected to stainless steel capillaries with an outer diameter of 0.9mm folded at a 45-degree angle. The thickness of the film is 20 μm, and the main inlet, liquid branch inlet and product or waste liquid outlet contacting the reaction liquid are also covered with a fluorine rubber O-ring at the front end of the silicone tube; the other end of the stainless steel capillary is connected to the infusion or gas pipeline Connection, wherein the material of the infusion line is polytetrafluoroethylene.

When the above-mentioned device is used, the main inlet 4 is connected to the common reagent bottle used in the reaction process, and the liquid branch inlet 6 can be connected to different changeable raw material pools respectively, and nitrogen gas is used as the driving force to drive the amino acid from the raw material bottle The raw material enters the microreactor, the gas branch inlet 2 is respectively connected with a plurality of ventilation solenoid valves, the opening and closing of each solenoid valve is controlled by the circuit board and software, and the product or waste liquid port 11 is connected with the receiving bottle.

Example 2

A perfluoropolymer microreactor with ten liquid flow branch pipelines and ten gas flow branch pipelines was fabricated by mechanical processing template and hot embossing method.

The manufacturing method of the perfluoropolymer microreactor of embodiment 2 is as follows: the stainless steel plate is processed by lathe and milling machine cutting mode, and the stainless steel positive mold pattern shown in Fig. 2 and Fig. 3 is obtained, and the hot embossing method is adopted , transfer the pattern on the stainless steel male mold to the perfluoropolymer sheet (including the perfluorinated cover sheet and the perfluorinated base sheet) to obtain the perfluorinated base sheet and the perfluorinated cover sheet with the structure required by the present invention; Through bonding, the three-layer perfluorinated material of perfluorinated substrate, perfluorinated elastic film and perfluorinated cover is bonded into a complete microreactor. In this embodiment, the materials of the perfluorinated substrate and the perfluorinated cover sheet are from DuPont Company of the United States. The melting point range is 302-310°C, and the material of the perfluoroelastic film is DuPont Company of the United States. The melting point range is 302-310°C.

The main implementation process is as follows: with reference to the perfluoropolymer microreactor structure of Example 1, the required stainless steel male mold pattern is obtained through outsourcing processing; a PFA plate is placed on the stainless steel male mold, and heat is carried out under conditions of heating and pressurization. Embossing, the hot embossing temperature is 260-290°C, the applied pressure is 0.1-0.2Mpa, and the pressurization time is 15-20 minutes; after the pressure is removed, the pattern negative mold is obtained on the PFA plate, and the structure required by the present invention is obtained The perfluorinated substrate and perfluorinated cover slip; after carefully cleaning the perfluorinated substrate, perfluorinated cover slip and perfluoroelastic film, carefully align them with the aid of a micro-alignment device, and fix them with stainless steel clamps; Place the perfluorinated substrate, perfluorinated cover sheet, perfluoroelastic film and stainless steel fixture in a chip bonding instrument for bonding, the bonding temperature is 240-270°C; the bonding time is 2-4 hours.

The structure of the perfluoropolymer cover sheet and the perfluorosubstrate of the fabricated perfluoropolymer microreactor is similar to the corresponding structure of the perfluoropolymer microreactor produced in Example 1.

The specifications of the perfluoropolymer microreactor produced in this example are different from those in Example 1. The differences are as follows: the size of the perfluoropolymer cover sheet is 6cm (length) × 3cm (width) × 2mm (thick); The length of the main flow pipeline is 3.6cm, the width is 1mm, and the depth is 200μm; the length of the liquid flow branch pipeline is 6mm, the width is 600μm, and the depth is 100μm; the length of the reaction chamber is 12mm, the width is 1.5mm, and the depth is 200μm; the single-layer width of the dam part of the reaction chamber is 600μm, the width of the liquid-passing part is 50μm, the dam spacing is 1mm, and the number of dams is 2 groups; the size of the perfluorinated substrate is 6cm (length) × 3cm (width) × 2mm (thickness); the width of the gas flow branch pipeline is 600 μm, and the depth is 200 μm; the inner diameters of all liquid branch inlets, product or waste liquid outlets, and gas branch inlets are 1 mm; the thickness of the perfluoroelastic film is 20 μm. All the other specifications are the same as those in Embodiment 1. The attachments used are the same as those in Example 1.

Example 3

A perfluoropolymer microreactor with ten branched pipelines for liquid flow and ten branched pipelines for gas flow was fabricated by mechanical processing.

The manufacturing method of the perfluoropolymer microreactor of Example 3 is as follows: the perfluoro plate is processed by lathe and milling machine cutting, and the perfluoro substrate and the perfluoro cover sheet having the structure shown in Fig. 2 and Fig. 3 are obtained ; By bonding, the perfluorinated substrate, the perfluoroelastic film and the perfluorinated cover sheet (in this embodiment, the perfluorinated substrate and the perfluorinated cover sheet are made of U.S. DuPont The melting point range is 302-310°C, and the material of the perfluoroelastic film is DuPont Company of the United States. Melting point range 255-265°C) Three layers of perfluorinated materials are laminated and bonded into a complete microreactor.

The main implementation process is as follows: with reference to the perfluoropolymer microreactor structure of Example 1, the perfluorinated substrate and the perfluorinated cover sheet with the structure required by the present invention are obtained through outsourcing processing; the perfluorinated substrate is , perfluorinated cover slip and perfluoroelastic film are carefully cleaned, carefully aligned with the aid of a micro-alignment device, and fixed with stainless steel fixtures; Put them in a chip bonding machine and perform bonding after stacking. The bonding temperature is 240-270° C., and the bonding time is 2-4 hours.

The structure of the perfluorinated cover sheet and the perfluorinated base sheet of the perfluoropolymer microreactor made is the same as that of the perfluoropolymer microreactor made in embodiment 1 and embodiment 2. structure is similar. The specification of the perfluoropolymer microreactor produced in this embodiment is slightly different from the specification of the perfluoropolymer microreactor produced in Example 2, that is, the single-layer width of the dam part of the reaction chamber is 1mm, and the dam spacing is 600 μm. All the other specifications and accessories thereof are the same as in Embodiment 2.

Example 4

The perfluoropolymer microreactor device produced in Example 1 was used for microreactor solid-phase peptide synthesis.

The solid phase carrier is commercial Wang resin (carrier particle size: 100 μm), and the amino acid at the C-terminal of the polypeptide is connected to the resin in advance according to the sequence of the polypeptide to be synthesized. In this example, Fmoc-Leu modified Wang resin was used as a solid phase carrier for polypeptide synthesis, and leucine enkephalin was synthesized using the perfluoropolymer microreactor device produced in Example 1. According to the method reported in the literature (Int.J.Pept.ProteinRes., 1990, 35, 161-214.), the polypeptide leucine enkephalin was synthesized, and its sequence was Tyr-Gly-Gly-Phe-Leu.

Concrete synthetic steps are as follows:

a. Use a syringe to pour Fmoc-Leu modified Wang resin dispersed in N'N-dimethylformamide (DMF) into the reaction chamber 9 from the main injection port 4 (as shown in Figure 6). The structure 10 exists, the carrier is bound in the reaction chamber 9, and the DMF solution is discharged from the product or waste liquid port 11.

b. The amino acid reagents Fmoc-Phe, Fmoc-Gly, Fmoc-Gly, and Fmoc-Tyr at the second to fifth positions of the C-terminal were respectively mixed with benzotriazole-N, N, N', N'-tetramethylurea hexa Fluorophosphate (HBTU) was mixed equimolarly, and an activation reagent (0.4mol/L N-methylmorpholine/DMF) was added to make an amino acid coupling reagent, which was added to the corresponding raw material pool for later use. Assemble the entire synthesis apparatus.

c. Deprotection: the deprotection reagent (DMF solution dissolved in hexahydropyridine) was injected into the microreactor through the main injection port 4, the injection flow rate was 2 μL/min, and the deprotection time was 5 minutes.

d. Cleaning the resin: Inject DMF solution from the main injection port 4 to wash the resin and remove residual deprotection reagents and impurities after the reaction. The injection flow rate is 100 μL/min, and the washing time is 6 minutes.

e. Amino acid coupling: open the micro-air valve of the second amino acid coupling reagent at the C-terminal corresponding to the liquid flow branch pipeline, and simultaneously close the micro-air valves of the other branch liquid flow branch pipelines. The method is: close the gas flow path corresponding to the micro-gas valve by software, remove the pressure corresponding to the valve area 3, and relax the perfluoroelastic film 14 at the micro-gas valve, so that the reaction liquid can be disconnected from the flow channel of the valve area 8 and the loose perfluoroelastic film 14 through the gap, thereby opening the micro-air valve; while maintaining the pressure of other valve areas 3, the perfluoroelastic film 14 at these micro-air valves is tense, making these micro-air valves The gap between the flow channel disconnection 8 of the valve area at the valve and the perfluoroelastic film 14 is closed, thereby closing these micro-air valves. Turn on the syringe pump, introduce the above-mentioned amino acid coupling reagent into the reaction chamber 9, and start the continuous flow reaction, the injection flow rate is 2 μL/min, and the coupling time is 20 minutes. Since the micro-air valves of other liquid flow branch lines are closed, the amino acid coupling reagent will not flow back into other liquid flow branch lines, thereby avoiding cross-contamination.

f. Wash the resin once according to step d.

g. Repeat steps c, d, e and f 3 times in sequence, wherein the amino acid reagents in step e are amino acid reagents Fmoc-Tyr, Fmoc-Gly and Fmoc-Gly in sequence, to finally obtain leuenkephalin.

h. After the synthesis of leupenkephalin is completed, a deprotection reagent is injected to remove the protective group of the N-terminal amino acid, and DMF is injected to wash the resin.

i. Solvent replacement: inject dichloromethane and methanol successively to replace the DMF solvent; the liquid pumping flow rate is 25 μL/min, and the replacement time is 3 minutes respectively.

j. In-situ cracking: Inject cracking reagent (aqueous solution of trifluoroacetic acid (TFA) with a mass percentage of 97.5%), the injection flow rate is 2 μL/min, and the cracking time is 30 minutes; the cracking product is collected at the outlet of the chip. Peptide Leucine Enkephalin.

Example 5

This example is used to illustrate the solid-phase decapeptide synthesis in a microreactor based on a perfluoropolymer microreactor using a rigid porous solid support.

The solid phase carrier is selected from self-made rigid, porous, high-load polyhydroxymethylmethoxymethylstyrene (HMP) resin, and the amino acid at the C-terminal of the polypeptide is connected to the resin in advance according to the sequence of the polypeptide to be synthesized. In this example, Fmoc-Lys modified rigid porous is used as a solid phase carrier for polypeptide synthesis, and the synthesis sequence is GGDYKDDDDK using the perfluoropolymer microreactor device prepared in Example 1.

Concrete synthetic steps are as follows:

a. Use a syringe to pour Fmoc-Lys modified HMP resin (carrier particle size: 10 μm) dispersed in N'N-dimethylformamide (DMF) into the reaction chamber from the main injection port 4 (as shown in Figure 6) In the body 9, due to the existence of the carrier binding structure 10, the carrier is bound in the reaction chamber 9, and the DMF solution is discharged from the product or waste liquid port 11.

b, the second to fifth amino acid reagents Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Lys, Fmoc-Tyr, Fmoc-Asp, Fmoc-Gly, Fmoc-Gly respectively with benzene Mix triazole-N,N,N',N'-tetramethyluronium hexafluorophosphate (HBTU) equimolarly, add activating reagent (0.4mol/LN-methylmorpholine/DMF) to form amino acid couple Add reagents to the corresponding raw material pool for later use. Assemble the entire synthesis apparatus.

c. Deprotection: the deprotection reagent (DMF solution dissolved in hexahydropyridine) was injected into the microreactor through the main injection port 4, the injection flow rate was 0.5 μL/min, and the deprotection time was 5 minutes.

d. Cleaning the resin: Inject DMF solution from the main injection port 4 to wash the resin and remove residual deprotection reagents and impurities after the reaction. The injection flow rate is 10 μL/min, and the washing time is 6 minutes.

e. Amino acid coupling: open the micro-air valve of the second amino acid coupling reagent at the C-terminal corresponding to the liquid flow branch pipeline, and close the micro-air valves of other liquid flow branch pipelines at the same time. The method is: close the gas flow path corresponding to the micro-gas valve through the control software, remove the pressure corresponding to the valve area 3, and relax the perfluoroelastic film 14 at the micro-gas valve, so that the reaction liquid can be cut off from the flow channel of the valve area. The slit between the opening 8 and the loose perfluoroelastic film 14 passes through, thereby opening the micro-air valve; while maintaining the pressure of other valve areas 3, the perfluoroelastic film 14 at these micro-air valves is tense, making these micro-air valves The gap between the flow channel disconnection 8 of the valve area at the air valve and the perfluoroelastic film 14 is closed, thereby closing these micro-air valves. Turn on the syringe pump, introduce the above-mentioned amino acid coupling reagent into the reaction chamber 9, and start the continuous flow reaction, the injection flow rate is 2 μL/min, and the coupling time is 20 minutes. Since the micro-air valves of other liquid flow branch lines are closed, the amino acid coupling reagent will not flow back into other liquid flow branch lines, thereby avoiding cross-contamination.

f. Wash the resin once according to step d.

g, repeat steps c, d, e and f8 times in sequence, wherein the amino acid reagents in step e are amino acid reagents Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Lys, Fmoc-Tyr, Fmoc-Asp, Fmoc-Gly, Fmoc-Gly, and finally a decapeptide final product.

h. After the synthesis is completed, inject a deprotection reagent to remove the protective group of the N-terminal amino acid, and inject DMF to wash the resin.

i. Solvent replacement: inject dichloromethane and methanol successively to replace the DMF solvent; the liquid pumping flow rate is 10 μL/min, and the replacement time is 3 minutes respectively.

j. In-situ cracking: Inject cracking reagent (aqueous solution of trifluoroacetic acid (TFA) with a mass percentage of 97.5%), the injection flow rate is 0.5 μL/min, and the cracking time is 30 minutes; the cracking product is collected at the outlet of the chip. target peptide.

The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention. These simple modifications All belong to the protection scope of the present invention.

In addition, it should be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable way if there is no contradiction. The combination method will not be described separately.

In addition, various combinations of different embodiments of the present invention can also be combined arbitrarily, as long as they do not violate the idea of the present invention, they should also be regarded as the disclosed content of the present invention.

Claims (9)

1. a (per) fluoropolymer microreactor, is characterized in that, described microreactor comprises: perfluor cover plate, perfluorinated elastomer film and perfluor substrate;

Wherein, described perfluor cover plate comprises: the liquid stream main line with reaction cavity, be positioned at the main injection port at liquid stream main line two ends and product or waste liquid outlet, be positioned at the liquid flow branching pipeline of liquid stream main line both sides, described reaction cavity is with support bound structure, one end of described liquid flow branching pipeline is communicated with liquid stream main line, and described liquid flow branching pipeline has liquid branch injection port, and valve district runner gap is set on described liquid flow branching pipeline, described support bound structure does not flow out reaction cavity for the solid-state carrier controlled in reaction cavity, and allow the liquid in reaction cavity to flow out reaction cavity,

Described perfluor substrate comprises: air flows pipeline, and described air flows pipeline has gas branch entrance and valve district, and the number of described air flows pipeline is identical with the number of the described liquid flow branching pipeline in perfluor cover plate;

The with good grounds gas pressure of described perfluorinated elastomer thin film controls the function of liquid flow branching pipeline break-make;

Wherein, perfluor substrate, perfluorinated elastomer film and perfluor cover plate stack gradually and bonding, and make after bonding the valve district runner gap of perfluor cover plate, the valve district of perfluor substrate and between perfluorinated elastomer film form micro-air valve, and air flows pipeline and liquid flow branching pipeline one_to_one corresponding.

2. microreactor according to claim 1, wherein, the material of described perfluor cover plate, perfluorinated elastomer film and perfluor substrate be selected from polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer and perfluoroethylene-propylene separately one or more.

3. microreactor according to claim 1 and 2, wherein, described valve district all can cover described valve district runner gap.

4. microreactor according to claim 3, wherein, described valve district is circular configuration, and the width of described valve district runner gap is 0.01-5mm, and the diameter in described valve district is 0.01-5mm.

5. microreactor according to claim 1 and 2, wherein, described support bound structure is made up of 1-50 group ponding, and often organizes the bottom that described ponding is formed in perfluor substrate, there is interval between at least one sidewall simultaneously often organizing described ponding and reaction cavity.

6. microreactor according to claim 5, wherein, two adjacent groups ponding is staggered.

7. microreactor according to claim 1 and 2, wherein, the number of described liquid flow branching pipeline is 2-60, and in distributed number such as liquid stream main line both sides.

8. microreactor according to claim 1 and 2, wherein, the thickness of described perfluorinated elastomer film is 0.01-0.1mm.

9. the application of the microreactor in claim 1-8 described in any one in Solid-phase organic synthesis.