CN104096526B - A kind of (per) fluoropolymer microreactor and application thereof - Google Patents
A kind of (per) fluoropolymer microreactor and application thereof Download PDFInfo
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- CN104096526B CN104096526B CN201310114549.9A CN201310114549A CN104096526B CN 104096526 B CN104096526 B CN 104096526B CN 201310114549 A CN201310114549 A CN 201310114549A CN 104096526 B CN104096526 B CN 104096526B
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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
Technical field
The present invention relates to micro-fluidic chip and microreaction technology 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 manufactured by micro Process 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, easy scale integrated with high flux reaction etc. feature, in fine chemistry industry and the good prospect of biomedicine field, be also the successful model that microfluidic 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, therefore the aspect such as Material selec-tion, processing technology of microreactor is subject to a lot of restriction.Based on specific chemical reaction condition, applicable material and processing method can be selected, design specific microreactor, thus reduce raw material or energy resource consumption, enhance productivity.Such as a kind of fast constant temperature microreactor disclosed in CN101696929A, can provide micro-reaction environment of constant temperature.And the serial microreactor of the disclosed different structure such as CN102240535A, CN102247787A, effectively can mix reaction raw materials.At present, the micro-fluidic chip reactor for Solid phase peptide synthssis based on hard materials such as glass also has report (as: Wanget.al, LabChip, 2011,11,929-935, CN102527306A).
But above-mentioned research is only optimized for specific a certain environment, and its microreactor can only adapt to specific reaction environment.At present, the research that can adapt to the universal microreactor of multiple reaction environment remains a difficult problem.Because it requires that the material of microreactor has and is close to harsh chemical resistance, 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 macromolecular material that the chemical resistance of generally acknowledging is the strongest is the perfluoro-polymer material that side chain is replaced by fluorine atoms completely, and trade name is " Teflon ".Microreactor based on perfluor or the slightly poor inclined fluorine macromolecular material of tolerance is also reported to some extent, but its performance or range of application still have limitation.Studies have reported that the micro-fluidic chip that one is material with PFPE (PFPE), this chip can be good at being operated in multiple organic solvent atmosphere, and this chip can carry out the organic synthesis (Huanget.al of DNA, LabChip, 2007,7,24-26).But it is too expensive that the subject matter of this material is price, and synthesize the equipment and process of this material require complexity.Although commercialization (per) fluoropolymer materials is as cheap in polytetrafluoroethylene (PTFE) etc., but due to reasons such as technology, (Renet.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 the (per) fluoropolymer microreactor of maturation.
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 work under the chemical reaction environment of Various Complex, can and be not limited to be applied in the circulating solid-state chemical reaction method reaction of multistep, as in Peptide systhesis reaction.
For realizing aforementioned object, the invention provides a kind of (per) fluoropolymer microreactor, wherein, 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.
For the present invention, the material of preferred described perfluor cover plate, perfluorinated elastomer film and perfluor substrate be selected from polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) and perfluoroethylene-propylene (FEP) separately one or more, 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 commercially available commercial material.
For the present invention, the fusing point of preferred tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer (PFA) is 302-310 DEG C, such as, can be du pont company
du pont company
For the present invention, the fusing point of preferred perfluoroethylene-propylene (FEP) is 255-265 DEG C, such as, can be du pont company
In the present invention, the liquid flow branching pipeline being 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, described micro-air valve is that its number is consistent with the number of liquid flow branching pipeline by perfluor substrate, perfluorinated elastomer film, perfluor cover plate through the three layers of air valve structure that are stacked and that obtain after bonding (can be described as superposition bonding) that align.Due to micro-air valve by the valve district runner gap of perfluor cover plate, the valve district of perfluor substrate and between perfluorinated elastomer film formed, therefore described micro-air valve comprises the valve district runner gap of perfluor cover plate, the valve district of perfluor substrate and the perfluorinated elastomer 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 and takes turns circular response more, and the reaction raw materials of often taking turns may be different, need multiple liquid flow branching pipeline so general.
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 control the break-make of a liquid flow branching pipeline respectively, its principle controlled is exactly act on described micro-air valve thus the break-make controlling liquid flow branching pipeline by passing into gas or cut off gas from air flows pipeline, such as when passing into gas, the perfluorinated elastomer film at micro-air valve place is tight, now liquid flow branching pipeline connection, may be used for into reaction raw materials, and when not needing into reaction raw materials, as long as the supply of removing gas can make the perfluorinated elastomer film at micro-air valve place relax, thus liquid flow branching pipeline is disconnected.
In the present invention, preferred described valve district all can cover described valve district runner gap, and Ji Fa district and valve district runner gap match.
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, is more preferably 0.1-2mm.
In the present invention, described support bound structure of a great variety, every support bound structure that can meet aforementioned claim of the present invention all can be used for the present invention, for the present invention, for the ease of processing, preferred described support bound structure is by 1-50 group, and preferred 3-6 group ponding is formed, and often organize the bottom that dam, described hurdle is formed in perfluor substrate, there is interval between at least one sidewall simultaneously often organizing described ponding and reaction cavity.For the present invention, preferably two adjacent groups ponding is staggered further.Wherein, preferably often organizing ponding width is further 0.01-1mm, is more preferably 0.05-0.2mm; Preferably often organizing ponding spacing is 0.1-8mm, be more preferably 0.5-5mm, and adjacent ponding is staggered.
In the present invention, because described reaction cavity has support bound structure, make the solid-state carrier in reaction cavity 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 perfluorinated elastomer film thickness is 0.01-0.1mm, is more preferably 0.020-0.05mm.
In the present invention, described main injection port, liquid branch injection port and gas branch inlet can carry sample by air pressure.
In the present invention, described micro-air valve can pass through 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 stamping, laser ablation method to make.
As previously mentioned, in the present invention, there is the described perfluor cover plate of aforementioned micro-structural, perfluor substrate can adopt the method for prior art formed obtain, concrete example is as formed as follows:
Stainless steel formpiston figure (selecting according to required figure) needed for obtaining through cooperation processing; Stainless steel formpiston is placed perfluor plate (for one or more in polytetrafluoroethylene (PTFE), tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer and perfluoroethylene-propylene), hot padding is carried out under the condition of heating pressurization, hot padding temperature is 240-300 DEG C, preferred hot padding temperature is 260-290 DEG C, applying pressure is 0.02-0.4Mpa, and preferably applying pressure is 0.1-0.2Mpa.Pressing time is 1-40 minute, and preferred pressing time is 10-20 minute; On perfluor plate, obtain pattern former after removing pressure, 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, described perfluor cover plate, between perfluorinated elastomer film and perfluor substrate, carry out bonding by thermal bonding or adhesive specially, particularly by methods such as micromanipulation, scale are auxiliary, perfluor cover plate, perfluorinated elastomer film and perfluor substrate are stacked gradually and bonding.
In the present invention, as previously mentioned, can adopt the method for prior art that perfluor cover plate, perfluorinated elastomer film and perfluor substrate are carried out bonding, particularly after adopting the method for prior art to define perfluor substrate, perfluor cover plate, stacked and bonding can be carried out as follows:
By described perfluor substrate, perfluor cover plate and perfluorinated elastomer film through carefully cleaning, at the auxiliary lower careful alignment of microscopic alignment device, and stainless steel fixture is used to fix; Perfluor substrate, perfluor cover plate and perfluorinated elastomer film and stainless steel fixture are placed in chip involution instrument and carry out bonding, bonding temperature is 240-300 DEG C, and preferred bonding temperature is 240-270 DEG C; Bonding time is 0.5-6 hour, and preferred 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 special restriction, such as, can adopt machining template and hot stamping to make, and chemical wet etching template and hot stamping also can be adopted to make.
In the present invention, described (per) fluoropolymer microreactor is connected with auxiliary equipment by 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 is by fluid sealant or the corrosion-resistant sebific duct sealing of elasticity, 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 carries out switch control rule 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 synthesis peptides thereof simultaneously.
The method that microreactor of the present invention is used for Solid-phase synthesis peptides is as follows: solid phase carrier passes into microreactor through main injection port, and loaded body constraint structure stops and stays in reaction cavity, after with reaction dissolvent cleaning, start building-up process.Each is taken turns need successively by deprotecting regent and amino acid/coupling reagent.Synthesize solvent used and deprotecting regent enters reaction cavity by main injection port; often wheel synthesis amino acid/coupling reagent used all passes into synthetic cavity from different liquid branch injection ports; each liquid branch pipeline uses the micro-Air Valve Control of perfluor air bladder automatically controlled to prevent liquid mixed flow between pipeline, to stop cross pollution.After synthesizing the polypeptide cleavage obtained, 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 chemical resistance that (per) fluoropolymer itself is excellent simultaneously, the chemical reaction environment of multiple harshness can be adapted to, as strong acid-base, organic solvent, high temperature etc., be suitable for and be not limited to the circulating solid-state chemical reaction method reaction of multistep, microreactor is integrated with the micro-air valve structure of perfluor air bladder of multichannel, can easily automation integrating control be carried out.Microreactor of the present invention has microminiaturization, integrated, high efficiency, low cost and eco-friendly advantage, to be widely used value in fine chemistry industry and biomedicine field.
Beneficial effect
1. the present invention adopts (per) fluoropolymer as microreactor base material; because (per) fluoropolymer has fabulous chemical inertness; almost all chemical reagent except the alkali metal of melting can be tolerated; and can long-term work in the environment of subzero 200 DEG C to 260 DEG C; therefore the microreactor using 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 features such as high flux reaction, also has fabulous universality.
2. on (per) fluoropolymer microreactor, be integrated with micro-air valve structure of multiple perfluor air bladder, can automation micro-air valve be regulated to open and close as required, thus realize flexible manipulation to reaction liquid, degree of integration and flexibility high, be very applicable to automation mechanized operation.
3. microreactor raw material used is commercial materials, manufactures means 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 Automated condtrol, is very applicable to industrial applications.
Accompanying drawing explanation
Accompanying drawing is 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 of the (per) fluoropolymer microreactor with ten liquid flow branching pipelines and ten air flows pipelines;
Fig. 2 is the design specification schematic diagram (top view) of the perfluor substrate of the (per) fluoropolymer microreactor with ten air flows pipelines;
Fig. 3 is the design specification schematic diagram (lower view) of the perfluor cover plate of the (per) fluoropolymer microreactor with ten liquid flow branching pipelines;
Fig. 4 is the micro-structural schematic diagram of reaction cavity;
Fig. 5 is (per) fluoropolymer microreactor each layer alignment thereof schematic diagram with ten liquid flow branching pipelines and ten air flows pipelines;
Fig. 6 is the complete structure schematic diagram of the (per) fluoropolymer microreactor with 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 entrance
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 support bound structures
11 products or waste liquid outlet 12 downstream main line
13 perfluor cover plate 14 perfluorinated elastomer films
15 perfluor substrates
Detailed description of the invention
Detailed description of the invention described herein, only for instruction and explanation of the present invention, is not limited to the present invention.
The experimental technique used in following embodiment if no special instructions, is conventional method.
Material used in following embodiment, reagent etc., if no special instructions, all can obtain from commercial channels.
Embodiment 1
Chemical wet etching template and hot stamping is adopted to carry out the making of the (per) fluoropolymer microreactor with 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 development, the Graphic transitions on mask on photoresist;
Step D represents stainless steel wet etching, thus obtains pattern formpiston on stainless steel;
Step e represents the residual photoresist removed on stainless steel;
Step F represents hot padding, the Graphic transitions on stainless steel formpiston on perfluoropolymer sheets, then obtains perfluor substrate and the perfluor cover plate of the micro-structural with application claims;
Step G represents stacked and bonding, is bonded to complete microreactor stacked for the alignment of three layers of perfluorinated material.
Main implementation process is as follows: according to microreactor of the present invention, and the pattern designed is made film mask; (model: 304) on plate, carries out uv-exposure after overlap film mask photoresist to be spin-coated on the stainless steel of surface finish; (model: 304) plate develops in photoresist (optical cement) developer solution, obtains the micro-pattern of photoresist to stainless steel after exposure; Corrosion resistant plate is put into stainless steel etching solution and carry out wet etching, obtain pattern formpiston, its line depth is 0.05-0.4mm; Make to remove photoresist with photoresist liquid removing residual photoresist cleaning; Stainless steel formpiston is placed perfluor substrate or perfluor cover plate, and (material is tetrafluoroethylene-perfluoroalkyl vinyl ether copolymer, i.e. PFA), under the condition of heating pressurization, carry out hot padding, hot padding temperature is 260-290 DEG C, applying pressure is 0.1-0.2Mpa, and pressing time is 15-20 minute.On perfluor substrate or perfluor cover plate, obtain the former with corresponding pattern after removing pressure, namely obtain the described perfluor substrate and perfluor cover plate with required micro-structural of the present invention thus; By described perfluor substrate, perfluor cover plate and perfluorinated elastomer film, (perfluor substrate and perfluor cover plate material are du pont company
melting range 302-310 DEG C, perfluorinated elastomer film material is du pont company
melting range 302-310 DEG C) after carefully cleaning, at the auxiliary lower careful alignment of microscopic alignment device, and use stainless steel fixture to fix; Perfluor substrate, perfluor cover plate and perfluorinated elastomer film and stainless steel fixture are placed in chip involution instrument and carry out bonding, bonding temperature is 240-270 DEG C; Bonding time is 2-4 hour.
The made perfluor cover plate of (per) fluoropolymer microreactor and the structure of perfluor substrate and microtubule road size design are as shown in Figures 2 and 3.For convenience of describing, here the top view of microreactor and lower view is specified, described each view all meets this and specifies: normally placed on a certain plane by complete (per) fluoropolymer microreactor, the view that observer sees from top to bottom is top view, and the view that observer sees from the bottom up is 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 placed 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 placed down.Wherein, in Fig. 3, Reference numeral is as follows: 4 main injection ports, 5 upstream main lines, be positioned at support bound structure upstream, 6 liquid branch injection ports, 7 liquid flow branching pipelines, 8 valve district runner gaps, 9 reaction cavities, 10 support bound structures, 11 products or waste liquid outlet.Wherein, the fine structure of reaction cavity 9 as shown in Figure 4,12 downstream main lines.Perfluor cover plate 13, perfluorinated elastomer film 14, perfluor substrate 15 close according to the complete stacked laggard line unit of the order shown in Fig. 5, obtain overall structure, namely the upper strata of microreactor is perfluor cover plate 13, its micro-structural down, middle level is perfluorinated elastomer film 14, lower floor is perfluor substrate 15, and its micro-structural upward.Stacked and the microreactor structure obtained after key of perfluor cover plate 13, perfluorinated elastomer film 14, perfluor substrate 15 as shown in Figure 6.Micro-air valve structure that bonding is formed as shown in Figure 7.
The specification of the (per) fluoropolymer microreactor that the present embodiment makes and auxiliary equipment (auxiliary equipment refers to: when microreactor is for the synthesis of reaction, needs the parts used to comprise connecting line, sealing ring, joint etc.) thereof 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 μm, and the degree of depth is 150 μm; The length of liquid flow branching pipeline is 4mm, and width is 150 μm, and the degree of depth is 100 μm; The length of reaction cavity is 8mm, and width is 800 μm, and the degree of depth is 150 μm; The ponding fractional monolayer width of reaction cavity is 200 μm, and liquid is 50 μm by the width of part, and ponding spacing is 300 μm, 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 μm, and the degree of depth is 150 μm; 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 being converted into miter angle, the silicone tube that stainless steel capillary overcoat has yardstick to be applicable to, the thickness of perfluorinated elastomer film is 20 μm, wherein, the main injection port of haptoreaction liquid, liquid branch injection port and product or waste liquid outlet also overlap in silicone tube front end and have fluororubber O-type ring; The stainless steel capillary other end is connected with transfusion or gas line, and wherein transfer line material is polytetrafluoroethylene (PTFE).
When using said apparatus, the public reagent bottle that main injection port 4 and course of reaction need use is connected, liquid branch injection port 6 can connect from different modifiable feed reservoir respectively, use nitrogen as driving force, amino acid starting material is driven to enter microreactor from stock bottle, gas branch entrance 2 is connected with multiple AVSV Air Vent Solenoid Valve respectively, and by circuit board and each magnetic valve folding of software control, product or waste liquid port 11 are connected with receiving bottle.
Embodiment 2
Machining template and hot stamping is adopted to carry out the making of the (per) fluoropolymer microreactor with ten liquid flow branching pipelines and ten air flows pipelines.
The preparation method of the (per) fluoropolymer microreactor of embodiment 2 is as follows: employing lathe, milling machine cutting way are processed stainless steel materials, obtain relative to having the stainless steel formpiston figure shown in Fig. 2 and Fig. 3, adopt hot stamping, figure on stainless steel formpiston is transferred on perfluoropolymer sheets (comprising perfluor cover plate and perfluor substrate) again, obtains perfluor substrate and the perfluor cover plate with structure of the presently claimed invention; By bonding, perfluor substrate, perfluorinated elastomer film and perfluor cover plate three layers of perfluorinated material 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, perfluorinated elastomer 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, the stainless steel formpiston figure needed for obtaining through cooperation processing; Stainless steel formpiston is placed PFA plate, and under the condition of heating pressurization, carry out hot padding, hot padding temperature is 260-290 DEG C, and applying pressure is 0.1-0.2Mpa, and pressing time is 15-20 minute; On PFA plate, obtain pattern former after removing pressure, obtain perfluor substrate and the perfluor cover plate of structure of the presently claimed invention; By described perfluor substrate, perfluor cover plate and perfluorinated elastomer film after carefully cleaning, at the auxiliary lower careful alignment of microscopic alignment device, and stainless steel fixture is used to fix; Perfluor substrate, perfluor cover plate and perfluorinated elastomer film and stainless steel fixture are placed in chip involution instrument and carry out bonding, bonding temperature is 240-270 DEG C; Bonding time is 2-4 hour.
The made perfluor cover plate of (per) fluoropolymer microreactor is similar to the counter structure of the (per) fluoropolymer microreactor made by embodiment 1 with the structure of perfluor substrate.
Specification and the embodiment 1 of the (per) fluoropolymer microreactor that the present embodiment makes are 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 μm; The length of liquid flow branching pipeline is 6mm, and width is 600 μm, and the degree of depth is 100 μm; The length of reaction cavity is 12mm, and width is 1.5mm, and the degree of depth is 200 μm; The ponding fractional monolayer width of reaction cavity is 600 μm, and liquid is 50 μm 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 μm, and the degree of depth is 200 μm; The internal diameter of all liq branch injection port, product or waste liquid outlet, gas branch entrance is 1mm; The thickness of perfluorinated elastomer film is 20 μm.All the other specifications are identical with the corresponding part specification in embodiment 1.Identical with embodiment 1 of the auxiliary equipment adopted.
Embodiment 3
Mechanical processing method is adopted to carry out the making of the (per) fluoropolymer microreactor with ten liquid flow branching pipelines and ten air flows pipelines.
The preparation method of the (per) fluoropolymer microreactor of embodiment 3 is as follows: employing lathe, milling machine cutting way are processed perfluor sheet material, obtain perfluor substrate and the perfluor cover plate with structure as shown in Figures 2 and 3; By bonding, perfluor substrate, perfluorinated elastomer 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, perfluorinated elastomer 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, obtains 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 perfluorinated elastomer film, after carefully cleaning, at the auxiliary lower careful alignment of microscopic alignment device, and use stainless steel fixture to fix; Perfluor substrate, perfluor cover plate and perfluorinated elastomer film and stainless steel fixture are placed in 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 made perfluor cover plate of (per) fluoropolymer microreactor is similar with the structure of perfluor substrate to the perfluor cover plate of the (per) fluoropolymer microreactor made by embodiment 1 and embodiment 2 with the structure of perfluor substrate.The (per) fluoropolymer microreactor specification that specification and the embodiment 2 of the (per) fluoropolymer microreactor that the present embodiment makes make is slightly different, and namely the ponding fractional monolayer width of reaction cavity is 1mm, and ponding spacing is 600 μm.All the other specifications and auxiliary equipment identical with embodiment 2.
Embodiment 4
The (per) fluoropolymer microreactor device utilizing embodiment 1 to make carries out microreactor Solid phase peptide synthssis.
Solid phase carrier chooses commercial Wang resin (diameter of carrier: 100 μm), according to the peptide sequence that will synthesize, and the amino acid that connecting peptides C holds on resin in advance.The Wang resin that the present embodiment is modified using Fmoc-Leu as Peptide systhesis solid phase carrier, the (per) fluoropolymer microreactor device synthesis LEK utilizing embodiment 1 to make.According to the method improvement on synthesis LEK that document (Int.J.Pept.ProteinRes., 1990,35,161-214.) is reported, 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 and be scattered in Wang resin that Fmoc-Leu N ' dinethylformamide (DMF) modifies in reaction cavity 9, because support bound structure 10 exists, carrier is bound in reaction cavity 9, and DMF solution is then discharged from product or waste liquid port 11.
B, by C hold 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) and be made into amino acid coupling reagents, it is for subsequent use to add corresponding feed reservoir.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, and the deprotection time is 5 minutes.
D, cleaning resin: inject DMF solution from main injection port 4, in order to washing resin, the deprotecting regent that removing is remaining and reaction rear impurity, injection flow velocity is 100 μ L/min, and wash time is 6 minutes.
E, amino acid couplings: open micro-air valve that C holds second amino acid coupling reagents corresponding liquid flow branching pipeline, close micro-air valve of other branch's liquid flow branching pipelines simultaneously.Method is: close gas flow path corresponding to this micro-air valve by software, shed the pressure in corresponding valve district 3, the perfluorinated elastomer film 14 at this micro-air valve place is relaxed, reaction liquid can be passed through from the gap between valve district runner gap 8 and lax perfluorinated elastomer film 14, thus open this micro-air valve; Keep the pressure in other valve districts 3 simultaneously, make the perfluorinated elastomer film 14 at this air valve place slightly nervous, this gap slightly between air valve Chu Fa district runner gap 8 and perfluorinated elastomer film 14 is closed, thus closes this air valve slightly.Open syringe pump, be incorporated into by above-mentioned amino acid coupling reagents 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 reagents can not blow back into other liquid flow branching pipelines, thus avoids cross pollution.
F, according to Step d washing resin once.
G, repeat 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 injects DMF washing resin.
I, solvent are replaced: successively inject carrene, methanol replacement 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 (mass percentage is the aqueous solution of the trifluoroacetic acid (TFA) of 97.5%), injection flow velocity is 2 μ L/min, and pyrolysis time is 30 minutes; Collect pyrolysis product in chip exit and namely obtain peptide LEK.
Embodiment 5
The present embodiment for illustration of utilize Rigid Porous solid phase carrier based on (per) fluoropolymer microreactor carry out microreactor solid phase decapeptide synthesis.
Solid phase carrier chooses self-control volume rigidity, porous, high capacity amount gather methylol methoxyl methyl styrene (HMP) resin, according to the peptide sequence that will synthesize, and the amino acid that connecting peptides C holds on resin in advance.The Rigid Porous that the present embodiment is modified using Fmoc-Lys is as Peptide systhesis solid phase carrier, and the (per) fluoropolymer microreactor device composition sequence utilizing embodiment 1 to make is GGDYKDDDDK.
Concrete synthesis step is as follows:
A, use syringe from main injection port 4(as Fig. 6) pour into and be scattered in HMP resin (diameter of carrier: 10 μm) that Fmoc-Lys N ' dinethylformamide (DMF) modifies in reaction cavity 9, because support bound structure 10 exists, carrier is bound in reaction cavity 9, and DMF solution is then discharged from product or waste liquid port 11.
B, by C hold 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) and be made into amino acid coupling reagents, it is for subsequent use to add corresponding feed reservoir.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, and the deprotection time is 5 minutes.
D, cleaning resin: inject DMF solution from main injection port 4, in order to washing resin, the deprotecting regent that removing is remaining and reaction rear impurity, injection flow velocity is 10 μ L/min, and wash time is 6 minutes.
E, amino acid couplings: open micro-air valve that C holds second amino acid coupling reagents corresponding liquid flow branching pipeline, close micro-air valve of other liquid flow branching pipelines simultaneously.Method is: close gas flow path corresponding to this micro-air valve by control software design, shed the pressure in corresponding valve district 3, the perfluorinated elastomer film 14 at this micro-air valve place is relaxed, reaction liquid can be passed through from the gap between valve district runner gap 8 and lax perfluorinated elastomer film 14, thus open this micro-air valve; Keep the pressure in other valve districts 3 simultaneously, make the perfluorinated elastomer film 14 at this air valve place slightly nervous, this gap slightly between air valve Chu Fa district runner gap 8 and perfluorinated elastomer film 14 is closed, thus closes this air valve slightly.Open syringe pump, be incorporated into by above-mentioned amino acid coupling reagents 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 reagents can not blow back into other liquid flow branching pipelines, thus avoids cross pollution.
F, according to Step d washing resin once.
G, repeat 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 injects DMF washing resin.
I, solvent are replaced: successively inject carrene, methanol replacement 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 (mass percentage is the aqueous solution of the trifluoroacetic acid (TFA) of 97.5%), injection flow velocity is 0.5 μ L/min, and pyrolysis time is 30 minutes; Pyrolysis product and target polypeptides is collected in chip exit.
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 characteristic described in above-mentioned detailed description of the invention, in reconcilable situation, can be combined by any suitable mode, in order to avoid unnecessary repetition, the present invention illustrates no longer separately to various possible combination.
In addition, also can be combined between various different embodiment of the present invention, 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, 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.
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| US6632655B1 (en) * | 1999-02-23 | 2003-10-14 | Caliper Technologies Corp. | Manipulation of microparticles in microfluidic systems |
| CN102527306A (en) * | 2010-12-28 | 2012-07-04 | 中国科学院化学研究所 | Array type continuously-flowing microfluidic chip device and manufacture method and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US6632655B1 (en) * | 1999-02-23 | 2003-10-14 | Caliper Technologies Corp. | Manipulation of microparticles in microfluidic systems |
| CN102527306A (en) * | 2010-12-28 | 2012-07-04 | 中国科学院化学研究所 | Array type continuously-flowing microfluidic chip device and manufacture method and application thereof |
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