CN104096526A - Perfluoropolymer micro-reactor and use thereof - Google Patents
Perfluoropolymer micro-reactor and use thereof Download PDFInfo
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- CN104096526A CN104096526A CN201310114549.9A CN201310114549A CN104096526A CN 104096526 A CN104096526 A CN 104096526A CN 201310114549 A CN201310114549 A CN 201310114549A CN 104096526 A CN104096526 A CN 104096526A
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Abstract
The invention provides a perfluoropolymer micro-reactor and a use thereof. The perfluoropolymer micro-reactor has a composite structure obtained by orderly laminating and bonding a perfluorinated cover plate, a perfluorinated elastic film and a perfluorinated base plate. The composite structure is provided with a miniature air valve. The perfluoropolymer micro-reactor is suitable for multiple chemical reaction conditions, is suitable for multicomponent-cycle solid phase chemical synthesis reaction, has high reaction efficiency, a less reagent use amount and a low making cost and can realize automatic reaction process control.
Description
Technical field
The present invention relates to micro-fluidic chip and microreactor technical field, be specifically related to a kind of (per) fluoropolymer microreactor and the application in solid phase organic synthesis thereof.
Background technology
Microreactor refers to the very small chemical reaction system of manufacturing by micro-processing or accurate machining technique, its reaction interface yardstick in sub-micron to submillimeter magnitude.It is large that microreactor has specific area, and mass-and heat-transfer speed is fast, and reagent consumes little, easily scale integrated with the feature such as high flux reacts, in fine chemistry industry and the good prospect of biomedicine field, be also the successful model that micro-fluidic chip technology is applied to chemical field.
Because chemical reaction usually needs to use strong acid-base, multiple organic solvent etc., and may need the reaction environments such as heating, pressurization, the aspects such as the therefore material of microreactor selection, processing technology are subject to a lot of restrictions.Based on specific chemical reaction condition, can select applicable material and processing method, design specific microreactor, thereby reduce raw material or energy resource consumption, enhance productivity.The disclosed a kind of fast constant temperature microreactor of for example CN101696929A, can provide micro-reaction environment of constant temperature.And the serial microreactor of the disclosed different structures such as CN102240535A, CN102247787A can effectively mix reaction raw materials.At present, also there is report (as: Wang et.al, Lab Chip, 2011,11,929-935, CN102527306A) based on hard materials such as glass for the synthetic micro-fluidic chip reactor of solid-phase polypeptide.
But above-mentioned research is only optimized for specific a certain environment, its microreactor can only adapt to specific reaction environment.The research that at present, can adapt to the universal microreactor of multiple reaction environment remains a difficult problem.Because it requires the material of microreactor to have the harsh chemical resistance of being close to, common macromolecular material and metal material all cannot be competent at, and the inert metal tolerances such as gold, platinum are good, but expensive, and are difficult to be processed into microreactor.At present, the generally acknowledged the strongest macromolecular material of chemical resistance is the perfluoro-polymer material that side chain is replaced by fluorine atom completely, and trade name is " Teflon ".Also report to some extent of microreactor based on perfluor or the slightly poor inclined to one side fluorine macromolecular material of tolerance, but its performance or range of application still have limitation.Studies have reported that a kind of micro-fluidic chip taking PFPE (PFPE) as material, this chip can be good at being operated in multiple organic solvent atmosphere, and this chip can carry out organic synthesis (the Huang et.al of DNA, Lab Chip, 2007,7,24-26).But it is too expensive that the subject matter of this material is price, and the equipment and process of synthetic this material require complexity.Although commercialization (per) fluoropolymer materials is as cheap in polytetrafluoroethylene (PTFE) etc., but due to reasons such as technology, (Ren et.al except the exploration of a small amount of research, Proc.Natl.Acad.Sci.U.S.A., 2011,108,8162-8166), have not yet to see the report of ripe (per) fluoropolymer microreactor.
Summary of the invention
The object of the invention is the defect for overcoming prior art, a kind of (per) fluoropolymer microreactor and application thereof are provided, described microreactor can be worked under the chemical reaction environment of Various Complex, can and be not limited to be applied in the circulating solid state chemistry synthetic reaction of multistep, in polypeptide synthetic reaction.
For realizing aforementioned object, the invention provides a kind of (per) fluoropolymer microreactor, wherein, described microreactor comprises: perfluor cover plate, perfluor elastic film and perfluor substrate;
Wherein, described perfluor cover plate comprises: the liquid stream main line with reaction cavity, be positioned at main injection port and product or the waste liquid outlet at liquid stream main line two ends, be positioned at the liquid flow branching pipeline of liquid stream main line both sides, described reaction cavity fetters structure with carrier, 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 on described liquid flow branching pipeline, valve district runner gap is set, described carrier constraint structure does not flow out reaction cavity for the solid-state carrier of controlling in reaction cavity, and liquid in permission reaction cavity flows out reaction cavity,
Described perfluor substrate comprises: air flows pipeline, and described air flows pipeline has gas branch entrance He Fa 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;
Described perfluor elastic film has according to the function of gas pressure control liquid flow branching pipeline break-make;
Wherein, perfluor substrate, perfluor elastic film and perfluor cover plate stack gradually and bonding, and after bonding, make perfluor cover plate Fa district runner gap, perfluor substrate Fa district and between perfluor elastic film form micro-air valve, and air flows pipeline is corresponding one by one with liquid flow branching pipeline.
For the present invention, the material of preferred described perfluor cover plate, perfluor elastic film and perfluor substrate is selected from one or more in polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and perfluoroethylene-propylene (FEP) separately, be preferably tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and/or perfluoroethylene-propylene (FEP), but be not limited to this.
In the present invention, described polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and perfluoroethylene-propylene (FEP) can be all the commercially available material that is purchased.
For the present invention, preferably the fusing point of tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) is 302-310 DEG C, for example, can be du pont company
du pont company
For the present invention, preferably the fusing point of perfluoroethylene-propylene (FEP) is 255-265 DEG C, for example, can be du pont company
In the present invention, the liquid flow branching pipeline that is positioned at liquid stream main line both sides forms pectinate texture on perfluor cover plate.And liquid stream main line comprises upstream main line (between reaction cavity and main injection port) and downstream main line (between reaction cavity and product or waste liquid outlet).
In the present invention, the three layer air valve structure of described micro-air valve for being obtained after align stacked and bonding (can be described as stack bonding) by perfluor substrate, perfluor elastic film, perfluor cover plate, its number is consistent with the number of liquid flow branching pipeline.Due to micro-air valve by perfluor cover plate Fa district runner gap, perfluor substrate Fa district and between perfluor elastic film form, therefore described micro-air valve comprises perfluor cover plate Fa district runner gap, perfluor substrate Fa district and the perfluor elastic film between described valve district runner gap and described valve district.
In the present invention, described liquid flow branching pipeline is generally used for into reaction raw materials, and because reactor is generally used for many wheel circular responses, and every reaction raw materials of taking turns may be different, so generally need multiple liquid flow branching pipelines.
In the present invention, described air flows pipeline is for controlling the break-make of liquid flow branching pipeline, wherein, each air flows pipeline can be controlled respectively the break-make of a liquid flow branching pipeline, thereby the principle of its control is exactly the break-make that acts on described micro-air valve control liquid flow branching pipeline by passing into gas or cut off gas from air flows pipeline, for example, in the time passing into gas, the perfluor elastic film at micro-air valve place is tight, now liquid flow branching pipeline connection, can be for entering reaction raw materials, and in the time not needing into reaction raw materials, as long as it is lax to remove the perfluor elastic film that the supply of gas can make micro-air valve place, thereby liquid flow branching pipeline is disconnected.
In the present invention, preferred described valve district can all cover described valve district runner gap, and Yu Fa district of Ji Fa district runner gap matches.
Further preferred described valve district is circular configuration, and the width of described valve district runner gap (distances between the liquid flow branching pipeline two ends of disconnection) is 0.01-5mm, and the diameter in described valve district is 0.01-5mm, more preferably 0.1-2mm.
In the present invention, described carrier fetters the of a great variety of structure, every carrier constraint structure that can meet the aforementioned requirement of the present invention all can be used for the present invention, for the present invention, for the ease of processing, preferred described carrier constraint structure is by 1-50 group, and preferably 3-6 group ponding forms, and dam, hurdle is formed on the bottom of perfluor substrate described in every group, described in every group, between ponding and at least one sidewall of reaction cavity, there is interval simultaneously.For the present invention, further preferably two adjacent groups ponding is staggered.Wherein, further preferred every group of ponding width is 0.01-1mm, more preferably 0.05-0.2mm; Preferred every group of ponding spacing is 0.1-8mm, more preferably 0.5-5mm, and adjacent ponding is staggered.
In the present invention, because described reaction cavity has carrier constraint structure, make the solid-state carrier in reaction cavity can not flow out reaction cavity, and liquid can flow out reaction cavity.
In the present invention, the liquid branch pipeline of preferred described perfluor cover plate is 2-60, is preferably 6-16, and in distributed number such as liquid stream main line both sides.
In the present invention, preferred described perfluor elastic film thickness is 0.01-0.1mm, more preferably 0.020-0.05mm.
In the present invention, described main injection port, liquid branch injection port and gas branch entrance can be carried sample by air pressure.
In the present invention, described micro-air valve can be by software control break-make.
The perfluor cover plate of described microreactor of the present invention and the on-chip micro-structural of perfluor, can adopt the methods such as mechanical milling method, chemical etching method, plasma etching method, hot padding method, laser ablation method to make.
As previously mentioned, in the present invention, have the described perfluor cover plate of aforementioned micro-structural, method that perfluor substrate can adopt prior art forms and obtains, concrete example is as formed as follows:
Obtain required stainless steel formpiston figure (selecting according to required figure) through cooperation processing; On stainless steel formpiston, place perfluor plate (for one or more in polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer and perfluoroethylene-propylene), under the condition of heating pressurization, carry out hot padding, hot padding temperature is 240-300 DEG C, preferably hot padding temperature is 260-290 DEG C, exert pressure as 0.02-0.4Mpa, preferably exert pressure as 0.1-0.2Mpa.Be 1-40 minute pressing time, and be preferably 10-20 minute pressing time; After removing pressure, on perfluor plate, obtain pattern former, obtain perfluor substrate and the perfluor cover plate of structure of the presently claimed invention.But the present invention is not limited to this.
In the present invention, between described perfluor cover plate, perfluor elastic film and perfluor substrate, carry out bonding by thermal bonding or adhesive specially, can, by methods such as micromanipulation, scale are auxiliary, perfluor cover plate, perfluor elastic film and perfluor substrate be stacked gradually and bonding particularly.
In the present invention, as previously mentioned, can adopt the method for prior art that perfluor cover plate, perfluor elastic film and perfluor substrate are carried out to bonding, when the method that adopts prior art has formed after perfluor substrate, perfluor cover plate, can carry out as follows stacked and bonding particularly:
Through carefully cleaning, in the auxiliary lower careful alignment of micro-aligning apparatus, and use stainless steel fixture to fix described perfluor substrate, perfluor cover plate and perfluor elastic film; Perfluor substrate, perfluor cover plate and perfluor elastic film and stainless steel fixture are placed in to chip involution instrument and carry out bonding, bonding temperature is 240-300 DEG C, and preferably bonding temperature is 240-270 DEG C; Bonding time is 0.5-6 hour, and preferably bonding time is 2-4 hour.But the present invention is not limited to this.
In the present invention, the preparation method of described microreactor without particular restriction, for example, can adopt machining template and hot padding method to make, and also can adopt chemical wet etching template and hot padding method to make.
In the present invention, described (per) fluoropolymer microreactor can be connected with auxiliary equipment by the following method: each liquid branch injection port is connected with raw material device for storing liquid by transfer line, and each liquid branch injection port can be by fluid sealant or the corrosion-resistant sebific duct sealing of elasticity, and raw material drives charging by syringe pump or air pressure.Each gas branch entrance is connected with gas source by connecting line, and encapsulating method is with the encapsulating method of each liquid branch injection port.Wherein, gas source can be carried out switch control by magnetic valve.The material of described transfer line and gas connecting line can be polytetrafluoroethylene (PTFE).
The present invention also provides above-mentioned (per) fluoropolymer microreactor and the application of auxiliary equipment in Solid-phase Polypeptide is synthetic thereof simultaneously.
Microreactor of the present invention is as follows for the synthetic method of Solid-phase Polypeptide: solid phase carrier passes into microreactor through main injection port, and is stopped and stay in reaction cavity by carrier constraint structure, after cleaning with reaction dissolvent, starts building-up process.Each is taken turns and need pass through successively deprotecting regent and amino acid/coupling reagent.Synthetic solvent and deprotecting regent used enters reaction cavity by main injection port; often take turns synthetic amino acid/coupling reagent used and all pass into synthetic cavity from different liquid branch injection ports; each liquid branch pipeline uses the micro-Air Valve Control of perfluor air bladder of automatically controlling to prevent liquid mixed flow between pipeline, to stop cross pollution.After the synthetic polypeptide cracking obtaining, receive from product-collecting mouth, verify by mass spectrography, and identify purity with reversed phase liquid chromatography.
(per) fluoropolymer microreactor provided by the invention can solve that extraordinary perfluorinated material price is high, the problem of processing difficulties, possesses the good chemical resistance of (per) fluoropolymer itself simultaneously, can adapt to the chemical reaction environment of multiple harshness, as strong acid-base, organic solvent, high temperature etc., be suitable for and be not limited to the circulating solid state chemistry synthetic reaction of multistep, the micro-air valve structure of perfluor air bladder that is integrated with multichannel on microreactor, can easily carry out the integrated control of automation.Microreactor of the present invention has microminiaturization, integrated, high efficiency, low cost and eco-friendly advantage, in fine chemistry industry and the biomedicine field value that is widely used.
Beneficial effect
1. the present invention adopts (per) fluoropolymer as microreactor base material; because (per) fluoropolymer has fabulous chemical inertness; can tolerate almost all chemical reagent except the alkali metal of melting; and can long-term work in the environment of subzero 200 DEG C to 260 DEG C; therefore the microreactor that uses its making to obtain is large except having specific area, and mass-and heat-transfer speed is fast, and reagent consumes little; module scale is integrated with outside the feature such as high flux reacts, and also has fabulous universality.
2. integrated micro-air valve structure of multiple perfluor air bladders on (per) fluoropolymer microreactor, automation regulates micro-air valve to open and close as required, thereby realize the flexible manipulation to reaction liquid, degree of integration and flexibility are high, be applicable to very much automation mechanized operation.
3. microreactor raw material used is commercialization material, and manufacture means are flexible, makes effect stability, greatly reduces manufacturing cost.
4. compared with prior art, the present invention has the advantages such as performance is outstanding, universality is good, degree of integration is high, easy to process, with low cost, and is easy to automation control, is applicable to very much industrial applications.
Brief description of the drawings
Accompanying drawing is to be used to provide a further understanding of the present invention, and forms a part for description, is used from explanation the present invention, but is not construed as limiting the invention with detailed description of the invention one below.In the accompanying drawings:
Fig. 1 is the manufacturing process schematic diagram with the (per) fluoropolymer microreactor of ten liquid flow branching pipelines and ten air flows pipelines;
Fig. 2 is the design specification schematic diagram (top view) with the perfluor substrate of the (per) fluoropolymer microreactor of ten air flows pipelines;
Fig. 3 is the design specification schematic diagram (lower view) with the perfluor cover plate of the (per) fluoropolymer microreactor of ten liquid flow branching pipelines;
Fig. 4 is the micro-structural schematic diagram of reaction cavity;
Fig. 5 is the each layer of alignment thereof schematic diagram of (per) fluoropolymer microreactor with ten liquid flow branching pipelines and ten air flows pipelines;
Fig. 6 is the complete structure schematic diagram with the (per) fluoropolymer microreactor of ten liquid flow branching pipelines and ten air flows pipelines;
Fig. 7 is the thin portion structural representation of micro-air valve.
Reference numeral
1 air flows pipeline 2 gas branch entrances
The 3 main injection ports in valve district 4
5 upstream main line 6 liquid branch injection ports
7 liquid flow branching pipeline 8 valve district runner gaps
9 reaction cavity 10 carrier constraint structures
11 products or waste liquid outlet 12 downstream main lines
13 perfluor cover plate 14 perfluor elastic films
15 perfluor substrates
Detailed description of the invention
Detailed description of the invention described herein only, for description and interpretation the present invention, is not limited to the present invention.
The experimental technique using in following embodiment if no special instructions, is conventional method.
Material, reagent etc. used in following embodiment, if no special instructions, all can obtain from commercial channels.
Embodiment 1
Adopt chemical wet etching template and hot padding method to there is the making of the (per) fluoropolymer microreactor of ten liquid flow branching pipelines and ten air flows pipelines.
The manufacturing process of described (per) fluoropolymer microreactor is as shown in Figure 1:
Wherein, steps A represents spin coating photoresist;
Step B represents uv-exposure;
Step C represents to develop, and the figure on mask is transferred on photoresist;
Step D represents stainless steel wet etching, thereby on stainless steel, obtains pattern formpiston;
Step e represents to remove the residual photoresist on stainless steel;
Step F represents hot padding, and the figure on stainless steel formpiston is transferred on (per) fluoropolymer sheet material, then obtains perfluor substrate and the perfluor cover plate of the micro-structural with requirement of the present invention;
Step G represents stacked and bonding, is bonded to complete microreactor stacked the alignment of three layers of perfluorinated material.
Main implementation process is as follows: according to microreactor of the present invention, the pattern designing is made to film mask; Photoresist is spin-coated on to the stainless steel (model: 304) on plate, carry out uv-exposure after overlap film mask of surface finish; Stainless steel (model: 304) plate develops in photoresist (optical cement) developer solution, obtains the micro-pattern of photoresist after exposure; Corrosion resistant plate is put into stainless steel etching solution and carry out wet etching, obtain pattern formpiston, its lines degree of depth is 0.05-0.4mm; The liquid that makes to remove photoresist is with photoresist removed residual photoresist and is cleaned; (material is tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer on stainless steel formpiston, to place perfluor substrate or perfluor cover plate, be PFA), under the condition of heating pressurization, carry out hot padding, hot padding temperature is 260-290 DEG C, exert pressure as 0.1-0.2Mpa, be 15-20 minute pressing time.After removing pressure, on perfluor substrate or perfluor cover plate, obtain the former with corresponding pattern, obtain thus described perfluor substrate and the perfluor cover plate with desired micro-structural of the present invention; By described perfluor substrate, perfluor cover plate and perfluor elastic film, (perfluor substrate and perfluor cover plate material are du pont company
melting range 302-310 DEG C, perfluor elastic film material is du pont company
melting range 302-310 DEG C) after carefully cleaning, in the auxiliary lower careful alignment of micro-aligning apparatus, and use stainless steel fixture to fix; Perfluor substrate, perfluor cover plate and perfluor elastic film and stainless steel fixture are placed in to chip involution instrument and carry out bonding, bonding temperature is 240-270 DEG C; Bonding time is 2-4 hour.
The perfluor cover plate of the (per) fluoropolymer microreactor of made and the structure of perfluor substrate and microtubule road size design are as shown in Figures 2 and 3.For convenience of describing, here specify top view and the lower view of microreactor, described each view all meets this appointment: complete (per) fluoropolymer microreactor is normally placed in a certain plane, the view that observer sees is from top to bottom top view, and the view that observer sees is from the bottom up lower view.Fig. 2 is the structural representation of perfluor substrate.In Fig. 2, each Reference numeral is as follows: 1 air flows pipeline, 2 gas branch entrances, 3 valve districts.Fig. 2 is top view, and the micro-structural of the perfluor substrate of the normal (per) fluoropolymer microreactor of placing upward.Fig. 3 is the structural representation of perfluor cover plate, is lower view, and the perfluor cover plate micro-structural of the normal (per) fluoropolymer microreactor of placing down.Wherein, in Fig. 3, Reference numeral is as follows: 4 main injection ports, 5 upstream main lines, be positioned at carrier constraint structure upstream, 6 liquid branch injection ports, 7 liquid flow branching pipelines, 8 valve district runner gaps, 9 reaction cavities, 10 carrier constraint structure, 11 products or waste liquid outlets.Wherein, the fine structure of reaction cavity 9 as shown in Figure 4,12 downstream main lines.Perfluor cover plate 13, perfluor elastic film 14, perfluor substrate 15 close according to the complete stacked laggard line unit of the order shown in Fig. 5, obtain overall structure, the upper strata that is microreactor is perfluor cover plate 13, its micro-structural down, middle level is perfluor elastic film 14, lower floor is perfluor substrate 15, and its micro-structural upward.The microreactor structure obtaining after the stacked and key of perfluor cover plate 13, perfluor elastic film 14, perfluor substrate 15 as shown in Figure 6.Micro-air valve structure that bonding forms as shown in Figure 7.
The specification of the (per) fluoropolymer microreactor that the present embodiment is made and auxiliary equipment thereof (auxiliary equipment refers to: microreactor, for the synthesis of when reaction, needs the parts that use to comprise connecting line, sealing ring, joint etc.) is as follows: it is long that perfluor cover plate is of a size of 4cm() × 2cm(is wide) × 2mm(is thick); The length of liquid stream main line is 2.4cm, and width is 400 μ m, and the degree of depth is 150 μ m; The length of liquid flow branching pipeline is 4mm, and width is 150 μ m, and the degree of depth is 100 μ m; The length of reaction cavity is 8mm, and width is 800 μ m, and the degree of depth is 150 μ m; The ponding part individual layer width of reaction cavity is 200 μ m, and liquid can be 50 μ m by the width of part, and ponding spacing is 300 μ m, and ponding number is 4 groups; It is long that perfluor substrate is of a size of 4cm() × 2cm(is wide) × 2mm(is thick); The width of air flows pipeline is 150 μ m, and the degree of depth is 150 μ m; The internal diameter of all liq branch injection port, product or waste liquid outlet, gas branch entrance is 1mm; All gas and liquid entrance are all connected the stainless steel capillary of the external diameter 0.9mm that is converted into miter angle, stainless steel capillary overcoat has the applicable silicone tube of yardstick, the thickness of perfluor elastic film is 20 μ m, wherein, the main injection port of haptoreaction liquid, liquid branch injection port and product or waste liquid outlet also overlap and have fluororubber O-type ring at silicone tube front end; The stainless steel capillary other end is connected with transfusion or gas line, and wherein transfer line material is polytetrafluoroethylene (PTFE).
While using said apparatus, the public reagent bottle that main injection port 4 need use with course of reaction is connected, liquid branch injection port 6 can connect from different variable feed reservoir respectively, use nitrogen as driving force, drive amino acid starting material to enter microreactor from stock bottle, gas branch entrance 2 is connected with multiple AVSV Air Vent Solenoid Valves respectively, and by circuit board and the each magnetic valve folding of software control, product or waste liquid mouth 11 are connected with receiving bottle.
Embodiment 2
Adopt machining template and hot padding method to there is the making of the (per) fluoropolymer microreactor of ten liquid flow branching pipelines and ten air flows pipelines.
The preparation method of the (per) fluoropolymer microreactor of embodiment 2 is as follows: adopt lathe, milling machine cutting way to process stainless steel materials, obtain with respect to thering is the stainless steel formpiston figure shown in Fig. 2 and Fig. 3, adopt hot padding method, figure on stainless steel formpiston is transferred to (per) fluoropolymer sheet material (comprising perfluor cover plate and perfluor substrate) upper, obtain having perfluor substrate and the perfluor cover plate of structure of the presently claimed invention; By bonding, perfluor substrate, perfluor elastic film and three layers of perfluorinated material of perfluor cover plate are bonded to complete microreactor.In the present embodiment, perfluor substrate and perfluor patch material are du pont company
melting range 302-310 DEG C, perfluor elastic film material is du pont company
melting range 302-310 DEG C.
Main implementation process is as follows: with reference to the (per) fluoropolymer microreactor structure of embodiment 1, obtain required stainless steel formpiston figure through cooperation processing; On stainless steel formpiston, place PFA plate, under the condition of heating pressurization, carry out hot padding, hot padding temperature is 260-290 DEG C, exerts pressure as 0.1-0.2Mpa, and be 15-20 minute pressing time; After removing pressure, on PFA plate, obtain pattern former, obtain perfluor substrate and the perfluor cover plate of structure of the presently claimed invention; After carefully cleaning, in the auxiliary lower careful alignment of micro-aligning apparatus, and use stainless steel fixture to fix described perfluor substrate, perfluor cover plate and perfluor elastic film; Perfluor substrate, perfluor cover plate and perfluor elastic film and stainless steel fixture are placed in to chip involution instrument and carry out bonding, bonding temperature is 240-270 DEG C; Bonding time is 2-4 hour.
The perfluor cover plate of the (per) fluoropolymer microreactor of made is similar to the counter structure of the (per) fluoropolymer microreactor of embodiment 1 made with the structure of perfluor substrate.
Specification and the embodiment 1 of (per) fluoropolymer microreactor that the present embodiment is made is different, and difference is as follows: perfluor cover plate is of a size of 6cm(and grows) × 3cm(is wide) × 2mm(is thick); The length of liquid stream main line is 3.6cm, and width is 1mm, and the degree of depth is 200 μ m; The length of liquid flow branching pipeline is 6mm, and width is 600 μ m, and the degree of depth is 100 μ m; The length of reaction cavity is 12mm, and width is 1.5mm, and the degree of depth is 200 μ m; The ponding part individual layer width of reaction cavity is 600 μ m, and liquid can be 50 μ m by the width of part, and ponding spacing is 1mm, and ponding number is 2 groups; It is long that perfluor substrate is of a size of 6cm() × 3cm(is wide) × 2mm(is thick); The width of air flows pipeline is 600 μ m, and the degree of depth is 200 μ m; The internal diameter of all liq branch injection port, product or waste liquid outlet, gas branch entrance is 1mm; The thickness of perfluor elastic film is 20 μ m.All the other specifications are identical with the corresponding part specification in embodiment 1.Identical with embodiment 1 of the auxiliary equipment adopting.
Embodiment 3
Adopt mechanical processing method to there is the making of the (per) fluoropolymer microreactor of ten liquid flow branching pipelines and ten air flows pipelines.
The preparation method of the (per) fluoropolymer microreactor of embodiment 3 is as follows: adopt lathe, milling machine cutting way to process perfluor sheet material, obtain and have perfluor substrate and the perfluor cover plate of structure as shown in Figures 2 and 3; By bonding, perfluor substrate, perfluor elastic film and perfluor cover plate, (in the present embodiment, perfluor substrate and perfluor cover plate material are du pont company
melting range 302-310 DEG C, perfluor elastic film material is du pont company
melting range 255-265 DEG C) three layers of layer of perfluorinated material poststack be bonded to complete microreactor.
Main implementation process is as follows: with reference to the (per) fluoropolymer microreactor structure of embodiment 1, obtain described perfluor substrate and the perfluor cover plate with structure of the presently claimed invention through cooperation processing; Described perfluor substrate, perfluor cover plate and perfluor elastic film, after carefully cleaning, in the auxiliary lower careful alignment of micro-aligning apparatus, and use stainless steel fixture to fix; Perfluor substrate, perfluor cover plate and perfluor elastic film and stainless steel fixture are placed in to chip involution instrument middle level poststack and carry out bonding, bonding temperature is 240-270 DEG C, and bonding time is 2-4 hour.
The perfluor cover plate of (per) fluoropolymer microreactor of the perfluor cover plate of the (per) fluoropolymer microreactor of made and the structure of perfluor substrate and embodiment 1 and embodiment 2 mades and the structural similarity of perfluor substrate.The (per) fluoropolymer microreactor specification that the specification of the (per) fluoropolymer microreactor that the present embodiment is made is made from embodiment 2 is slightly different, and the ponding part individual layer width of reaction cavity is 1mm, and ponding spacing is 600 μ m.All the other specifications and auxiliary equipment thereof are identical with embodiment 2.
Embodiment 4
Utilize (per) fluoropolymer microreactor device that embodiment 1 makes to carry out microreactor solid-phase polypeptide synthetic.
Solid phase carrier is chosen commercial Wang resin (diameter of carrier: 100 μ m), according to peptide sequence that will be synthetic, connect the amino acid of peptide C end in advance on resin.The Wang resin that the present embodiment is modified using Fmoc-Leu, as the synthetic solid phase carrier of polypeptide, utilizes the (per) fluoropolymer microreactor device that embodiment 1 makes to synthesize LEK.The synthetic polypeptide LEK of method of reporting according to document (Int.J.Pept.Protein Res., 1990,35,161-214.), its sequence is Tyr-Gly-Gly-Phe-Leu.
Concrete synthesis step is as follows:
A, use syringe from main injection port 4(as Fig. 6) pour into Wang resin that the Fmoc-Leu that is scattered in N ' dinethylformamide (DMF) modifies to reaction cavity 9, because carrier constraint structure 10 exists, carrier is bound in reaction cavity 9, and DMF solution is discharged from product or waste liquid mouth 11.
B, by C end the second to five amino acids reagent Fmoc-Phe, Fmoc-Gly, Fmoc-Gly, Fmoc-Tyr respectively with BTA-N, N, N ', mole mixing such as N '-tetramethylurea hexafluorophosphate (HBTU), add activating reagent (0.4mol/LN-methylmorpholine/DMF) to be made into amino acid coupling reagent, add corresponding feed reservoir for subsequent use.Assemble whole synthesizer.
C, deprotection: deprotecting regent (being dissolved with the DMF solution of hexahydropyridine) injects microreactor by main injection port 4, injection flow velocity is 2 μ L/min, the deprotection time is 5 minutes.
D, cleaning resin: inject DMF solution from main injection port 4, in order to washing resin, remove remaining deprotecting regent and reaction rear impurity, injection flow velocity is 100 μ L/min, and wash time is 6 minutes.
E, amino acid coupling: open micro-air valve of the corresponding liquid flow branching of C end second amino acid coupling reagent pipeline, close micro-air valve of other branch's liquid flow branching pipelines simultaneously.Method is: close by software the gas flow path that this micro-air valve is corresponding, shed the pressure in corresponding valve district 3, make the perfluor elastic film 14 at this micro-air valve place lax, make reaction liquid can Cong Fa district runner gap 8 and lax perfluor elastic film 14 between gap pass through, thereby open this micro-air valve; Keep the pressure in other valve districts 3 simultaneously, make this perfluor elastic film 14 anxieties at air valve place slightly, this is closed in the gap between air valve Chu Fa district runner gap 8 and perfluor elastic film 14 slightly, thereby close this air valve slightly.Open syringe pump, above-mentioned amino acid coupling reagent is incorporated in reaction cavity 9, start continuous-flow reaction, injecting flow velocity is 2 μ L/min, and coupling time is 20 minutes.Because micro-air valve of other liquid flow branching pipelines is closed, amino acid coupling reagent can not blow back into other liquid flow branching pipelines, thereby avoids cross pollution.
F, according to d step washing resin once.
G, repeating step c, d, e and f3 time successively, wherein, the amino-acid reagent in step e is followed successively by amino-acid reagent Fmoc-Tyr, Fmoc-Gly and Fmoc-Gly, finally obtains leu-enkaphalin.
After h, leu-enkaphalin end of synthesis, inject deprotecting regent, the blocking group of N terminal amino acid is removed, and inject DMF washing resin.
I, solvent exchange: successively inject carrene, methyl alcohol displacement DMF solvent; It is 25 μ L/min that liquor pump enters flow velocity, and time swap is respectively 3 minutes.
J, lysisin situ: inject lytic reagent (aqueous solution of the trifluoroacetic acid (TFA) that quality percentage composition is 97.5%), injection flow velocity is 2 μ L/min, and pyrolysis time is 30 minutes; Collect pyrolysis product in chip exit and obtain peptide LEK.
Embodiment 5
The present embodiment for illustrate utilize rigidity porous solid phase carrier to carry out microreactor solid phase decapeptide based on (per) fluoropolymer microreactor synthetic.
Solid phase carrier is chosen self-control volume rigidity, porous, poly-methylol methoxyl methyl styrene (HMP) resin of high capacity amount, according to peptide sequence that will be synthetic, connects in advance the amino acid of peptide C end on resin.The rigidity porous that the present embodiment is modified using Fmoc-Lys is as the synthetic solid phase carrier of polypeptide, and utilizing the (per) fluoropolymer microreactor device composition sequence that embodiment 1 makes is GGDYKDDDDK.
Concrete synthesis step is as follows:
A, use syringe from main injection port 4(as Fig. 6) pour into the HMP resin (diameter of carrier: 10 μ are m) to reaction cavity 9 that the Fmoc-Lys that is scattered in N ' dinethylformamide (DMF) modifies, because carrier constraint structure 10 exists, carrier is bound in reaction cavity 9, and DMF solution is discharged from product or waste liquid mouth 11.
B, by C end the second to five amino acids reagent Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Lys, Fmoc-Tyr, Fmoc-Asp, Fmoc-Gly, Fmoc-Gly respectively with BTA-N, N, N ', mole mixing such as N '-tetramethylurea hexafluorophosphate (HBTU), add activating reagent (0.4mol/LN-methylmorpholine/DMF) to be made into amino acid coupling reagent, add corresponding feed reservoir for subsequent use.Assemble whole synthesizer.
C, deprotection: deprotecting regent (being dissolved with the DMF solution of hexahydropyridine) injects microreactor by main injection port 4, injection flow velocity is 0.5 μ L/min, the deprotection time is 5 minutes.
D, cleaning resin: inject DMF solution from main injection port 4, in order to washing resin, remove remaining deprotecting regent and reaction rear impurity, injection flow velocity is 10 μ L/min, and wash time is 6 minutes.
E, amino acid coupling: open micro-air valve of the corresponding liquid flow branching of C end second amino acid coupling reagent pipeline, close micro-air valve of other liquid flow branching pipelines simultaneously.Method is: close by controlling software the gas flow path that this micro-air valve is corresponding, shed the pressure in corresponding valve district 3, make the perfluor elastic film 14 at this micro-air valve place lax, make reaction liquid can Cong Fa district runner gap 8 and lax perfluor elastic film 14 between gap pass through, thereby open this micro-air valve; Keep the pressure in other valve districts 3 simultaneously, make this perfluor elastic film 14 anxieties at air valve place slightly, this is closed in the gap between air valve Chu Fa district runner gap 8 and perfluor elastic film 14 slightly, thereby close this air valve slightly.Open syringe pump, above-mentioned amino acid coupling reagent is incorporated in reaction cavity 9, start continuous-flow reaction, injecting flow velocity is 2 μ L/min, and coupling time is 20 minutes.Because micro-air valve of other liquid flow branching pipelines is closed, amino acid coupling reagent can not blow back into other liquid flow branching pipelines, thereby avoids cross pollution.
F, according to d step washing resin once.
G, repeating step c, d, e and f8 time successively, wherein, amino-acid reagent in step e is followed successively by amino-acid reagent Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Asp, Fmoc-Lys, Fmoc-Tyr, Fmoc-Asp, Fmoc-Gly, Fmoc-Gly, finally obtains decapeptide end-product.
After h, end of synthesis, inject deprotecting regent, the blocking group of N terminal amino acid is removed, and inject DMF washing resin.
I, solvent exchange: successively inject carrene, methyl alcohol displacement DMF solvent; It is 10 μ L/min that liquor pump enters flow velocity, and time swap is respectively 3 minutes.
J, lysisin situ: inject lytic reagent (aqueous solution of the trifluoroacetic acid (TFA) that quality percentage composition is 97.5%), injection flow velocity is 0.5 μ L/min, and pyrolysis time is 30 minutes; Collecting pyrolysis product in chip exit is target polypeptides.
More than describe the preferred embodiment of the present invention in detail; but the present invention is not limited to the detail in above-mentioned embodiment, within the scope of technical conceive of the present invention; can carry out multiple simple variant to technical scheme of the present invention, these simple variant all belong to protection scope of the present invention.
It should be noted that in addition, each concrete technical characterictic described in above-mentioned detailed description of the invention, in reconcilable situation, can combine by any suitable mode, for fear of unnecessary repetition, the present invention is to the explanation no longer separately of various possible combinations.
In addition, between various embodiment of the present invention, also can be combined, as long as it is without prejudice to thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (9)
1. a (per) fluoropolymer microreactor, is characterized in that, described microreactor comprises: perfluor cover plate, perfluor elastic film and perfluor substrate;
Wherein, described perfluor cover plate comprises: the liquid stream main line with reaction cavity, be positioned at main injection port and product or the waste liquid outlet at liquid stream main line two ends, be positioned at the liquid flow branching pipeline of liquid stream main line both sides, described reaction cavity fetters structure with carrier, 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 on described liquid flow branching pipeline, valve district runner gap is set, described carrier constraint structure does not flow out reaction cavity for the solid-state carrier of controlling in reaction cavity, and liquid in permission reaction cavity flows out reaction cavity,
Described perfluor substrate comprises: air flows pipeline, and described air flows pipeline has gas branch entrance He Fa 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;
Described perfluor elastic film has according to the function of gas pressure control liquid flow branching pipeline break-make;
Wherein, perfluor substrate, perfluor elastic film and perfluor cover plate stack gradually and bonding, and after bonding, make perfluor cover plate Fa district runner gap, perfluor substrate Fa district and between perfluor elastic film form micro-air valve, and air flows pipeline is corresponding one by one with liquid flow branching pipeline.
2. microreactor according to claim 1, wherein, the material of described perfluor cover plate, perfluor elastic film and perfluor substrate is selected from one or more in polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer and perfluoroethylene-propylene separately.
3. microreactor according to claim 1 and 2, wherein, described valve district can all 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, the diameter in described valve district is 0.01-5mm.
5. microreactor according to claim 1 and 2, wherein, described carrier constraint structure is made up of 1-50 group ponding, and described in every group, dam, hurdle is formed on the bottom of perfluor substrate, has interval described in every group between ponding and at least one sidewall of reaction cavity simultaneously.
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 perfluor elastic film is 0.01-0.1mm.
9. the application of the microreactor described in any one in solid phase organic synthesis in claim 1-8.
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| US6632655B1 (en) * | 1999-02-23 | 2003-10-14 | Caliper Technologies Corp. | Manipulation of microparticles in microfluidic systems |
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| US6632655B1 (en) * | 1999-02-23 | 2003-10-14 | Caliper Technologies Corp. | Manipulation of microparticles in microfluidic systems |
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