CN101881750A - Microchip detector of content of arteannuin in Artemisia apiacea and method thereof - Google Patents
Microchip detector of content of arteannuin in Artemisia apiacea and method thereof Download PDFInfo
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Abstract
The invention discloses a microchip detector of the content of arteannuin in Artemisia apiacea and a method thereof, belonging to the technical field of content detection of arteannuin in the Artemisia apiacea raw material. The microchip detector of the invention mainly comprises a power source, a microfluidic chip, a detection circuit, a data acquisition circuit and a computer. The method comprises the following steps: taking the Artemisia apiacea as raw material; firstly, preparing arteannuin extract; using the device of the invention to carry out conductivity detection; and working out the content of the arteannuin in the Artemisia apiacea with a standard curve method. The device of the invention has simple and microminiaturization structure, good portability and convenient utilization; and the method of the invention is simple, rapid and accurate to check, has high efficiency and small detection sample usage, and meets the requirement of quick detection. The invention can also be widely used for analysis and detection of natural product active constituent and is particularly suitable for the detection of the content of the arteannuin in the Artemisia apiacea raw material, particularly the rapid detection.
Description
One. technical field
The invention belongs to artemislnin content detection technique field in the sweet wormwood raw material, be specifically related to the microchip detector and the detection method of artemislnin content in the sweet wormwood raw material.
Two. background technology
Qinghaosu is the sesquiterpene lactone compounds that contains peroxide bridge that extracts from sweet wormwood, is described as unique in the world effective antimalarial agent by the World Health Organization (WHO).Though the chemical synthesis process of qinghaosu has just been arranged from nineteen eighty-three, sweet wormwood have a plurality of asymmetric carbon atoms, the synthesis step complexity, productive rate is low, cost is high, obtains promoting on synthetic thereby be difficult in industry.From the natural plants sweet wormwood, extract the separation qinghaosu at present and be still its main source.Seek accurate, easy, qinghaosu quantitative detecting method fast, for optimize the extraction process condition, the on-the-spot purchase of research qinghaosu pharmacological effect, particularly medicinal material is all significant.
The pick-up unit and the detection method thereof of artemislnin content in the existing sweet wormwood raw material, as the patent No. is " assay method of artemislnin content in a kind of sweet wormwood " patent of ZL 200610095017.5, disclosed is qinghaosu in a kind of incomplete extraction sweet wormwood to be measured, adopt the ultraviolet spectro-photometric analysis method to measure artemislnin content data M (mg/g), substitution by with qinghaosu is extracted fully the linear model that the corresponding relation that adopts the artemislnin content data that liquid phase chromatography measures in the back is set up, calculate the actual content X (mg/g) of the qinghaosu in the sweet wormwood of surveying.The major defect of this patent is: the ultraviolet character of utilizing qinghaosu, need with aqueous slkali the qinghaosu 30min that in 50 ℃ of water-baths, derives, generation is carried out ultraviolet detection with ultraviolet spectrophotometer at wavelength 292nm after having the compound (a, alpha, beta-unsaturated ketone hydrochlorate) of uv absorption.This method complex operation step, time-consuming is damaged through qinghaosu structure after the alkali derivatization treatment, produces content loss.Extract qinghaosu in the raw material fully with the Soxhlet extracting method, use that to extract quantity of solvent bigger, and the 30h that reaches consuming time.And the ultraviolet spectrophotometer volume is bigger.And for example roll up 109～111 pages of the 2nd phases " content that the high performance capillary electrophoresis conductance method is measured qinghaosu " literary compositions in March, 2006 " analytical test journal " the 25th, the capillary electrophoresis apparatus of employing comprises that mainly high-voltage power supply, separation length are quartz capillary column, the Capillary Electrophoresis data station of 50cm; The method that adopts is: be to extract solvent with the sherwood oil, with a certain amount of activated carbon decolorizing of sweet wormwood raw material, and use sonic oscillation 30min behind the immersion 10min, obtain the qinghaosu extract after repeating to extract 3 times.Successively with NaOH, redistilled water, the buffer solution flushing kapillary 5min of 0.1mol/L, wash kapillary 3min between per twice sample introduction before each the detection.Adopt the gravity sample introduction, sample introduction height 20cm, sample injection time 15s.With Tris-H
3BO
2Be electrophoretic medium, at the 15kV high pressure, the styletable conductance method detects artemislnin content under the alkali condition of pH9.0.(ρ mg/L) sets up linear regression side Y=202+102 ρ, thereby calculates artemislnin content with mass concentration with peak height (Y).The major defect of this capillary electrophoresis apparatus is: capillary electrophoresis apparatus is the bigger commercialization instrument of volume, and structure is comparatively complicated, be not easy to carry, and the instrument costliness.The major defect of this method is: extract the qinghaosu complicated operation, and the sample separation time is longer, detection speed is slower, uses the whole cost of the bigger capillary electrophoresis apparatus of volume higher.
Three. summary of the invention
The objective of the invention is deficiency at artemislnin content pick-up unit and detection method in the existing sweet wormwood raw material, the microchip detector and the method thereof of artemislnin content in a kind of sweet wormwood are provided, have efficiently, easy, with low cost, be applicable to characteristics such as qinghaosu express-analysis detection.
Mechanism of the present invention: electricity is led and detected is a kind of universal Electrochemical Detection mode, just can be detected as long as can change the material of electrical conductivity of solution, and applicability is wide, and sensitivity is higher, and detection limit can reach 10
-6Mol/L.The micro-fluidic chip analytic system has the advantage that the analytic sample amount is few, analysis time is short, the huge advantage aspect microminiaturized, integrated and portability.Use the micro-fluidic chip conductance method to detect qinghaosu and not only can avoid the qinghaosu derivatization process, the step that simplifies the operation can be finished the assay of qinghaosu in 0.5 hour, greatly shortened the detection time of qinghaosu.And by detection technique and analysis chip integrated make instrument microminiaturized and integrated aspect have more advantage.The present invention is the specific extraction solvent with a kind of organic solvent in chloroform, ethyl acetate, the normal hexane, in microminiature glass extraction element, to the qinghaosu in the sweet wormwood raw material carry out fast, high efficiency extraction.Then extract is led in the microchip detector of detection by miillpore filter direct injected to the integrated electricity of self manufacture, the qinghaosu extract is not had derive, online express-analysis detects.
The technical scheme that realizes the object of the invention is: the microchip detector of artemislnin content in a kind of sweet wormwood mainly comprises power supply, micro-fluidic chip, testing circuit, data acquisition circuit and computing machine.
Described power supply is that commercial model is the micro-fluidic chip small intelligent high-voltage power supply of XCDY, and input voltage is civil power 220V, and output voltage is that 0～8000KV is adjustable, and electric current is that 0～500 μ Ak is adjustable.Power supply is connected with two platinum electrode pins by concentric cable, in order to supply with the separation voltage of the qinghaosu extract in the microchannel.
Described micro-fluidic chip is to be integrated with the micro-fluidic chip that electricity is led integrated microelectrode of detection chip and criss-cross microchannel, mainly leads the integrated microelectrode of detection chip, dimethyl silicone polymer (PDMS) cover plate formation by glass substrate, electricity.Its glass substrate is the substrate of electrophoresis chip, and the length of glass substrate is that 40000～50000 μ m, width are that 20000～30000 μ m, thickness are 500～1000 μ m.In an end of glass substrate and be positioned at the terminal position of the horizontal microchannel of criss-cross microchannel, directly metallic gold (Au) or platinum (Pt) or platinum evanohm materials such as (Pt-Cr) are deposited on the glass substrate and form electricity and lead the integrated microelectrode of detection chip.Electricity is led the integrated microelectrode of detection chip and is made up of 2～4 groups of electrode pairs, and the length of every pair of electrode is that 3000～5000 μ m, width are that 20～100 μ m, thickness are 10～50 μ m, and the spacing between the every pair of electrode is 20～100 μ m.And to cover a layer thickness at the outside surface of microelectrode be 0.01～0.1 μ m silicon dioxide insulator thin layer, to guarantee the isolation between the buffer solution and detected liquid in microelectrode and the microchannel, avoids the microelectrode corrosion failure.At the two ends of microelectrode, be connected with the input end of testing circuit by pin respectively, feed back to testing circuit in order to conductance signal with qinghaosu.On glass substrate, be coated with the dimethyl silicone polymer that a layer thickness is 1000～1500 μ m (PDMS) cover plate.On dimethyl silicone polymer (PDMS) cover plate, be provided with the criss-cross microchannel that utilizes the SU-8 anode membrane to make by solid casting method, the width of microchannel is that 30～80 μ m, the degree of depth are 10～30 μ m, length along the horizontal microchannel of glass substrate length direction is 5000～15000 μ m, is 1000～3000 μ m along the length of vertical microchannel of glass substrate Width.Two ends at horizontal, vertical microchannel, the delivery hole that volume is 3～5 μ L is set (promptly in the end of criss-cross microchannel respectively, be provided with A, B, four delivery holes of C, D altogether), in order to ensure in the criss-cross microchannel of micro-fluidic chip the supply of buffer solution and detected liquid.Micro-fluidic chip is fixedly connected on the testing circuit by pin.
Described testing circuit is led testing circuit for ordering integrated electricity, mainly comprises by I/V change-over circuit, multiplying operational circuit, low-pass filter circuit, differential amplifier circuit constituting, and is integrated on the pcb board.Electricity is led testing circuit and is connected by lead with data acquisition circuit.Testing circuit provides 220V power supply by commercial power supply (model is 4NIC-X39).
Described data acquisition circuit is for ordering integrated A/D signal acquisition circuit, and described computing machine is commercial computing machine.Data acquisition circuit is connected with computing machine by serial ports, by the conductance signal capture program conductance signal is transformed into digital signal, calculate, and output and preservation data, display result.
The microchip detection method of artemislnin content in a kind of sweet wormwood raw material is a raw material with the sweet wormwood, and preparation qinghaosu extract utilizes apparatus of the present invention to carry out electricity again and leads detection earlier, calculates artemislnin content by calibration curve method then, and its concrete detection step is as follows:
(1) qinghaosu extract preparation
With the sweet wormwood is raw material, is to extract solvent with organic solvents such as chloroform or ethyl acetate or normal hexanes.Earlier with the sweet wormwood raw material pulverizing and after 40～60 mesh sieves sieve, be 1: 6～8 ratio again in the ratio of the quality (g) of sweet wormwood and volume of organic solvent (m1), sweet wormwood and organic solvent are positioned in the extraction element, after 1～3min is extracted in jolting at ambient temperature, standing demix 1～3min, collect the upper strata stillness of night with the glass injection-tube, abandon lower floor's dregs of a decoction.The supernatant liquor of collecting is the qinghaosu extract.
(2) the micro-fluidic chip electricity of qinghaosu is led detection
(1) step finish after, earlier in the criss-cross microchannel of apparatus of the present invention, pass through glass syringe, inject buffer solution (as trishydroxymethylaminomethane-boric acid or potassium dihydrogen phosphate-sodium dihydrogen phosphate or borax-boric acid), after with (1) the qinghaosu extract prepared of step, by front end the glass syringe of miillpore filter is housed, injects two delivery holes (being C, D delivery hole) at vertical microchannel two ends of criss-cross microchannel.Then by two platinum electrode pins and two concentric cable, the output terminal of power supply is connected with two delivery holes (being A, B delivery hole) at the horizontal microchannel two ends of the criss-cross microchannel of micro-fluidic chip respectively, qinghaosu extract in the criss-cross microchannel is under 300～900V voltage, carry out electrophoretic separation, and the electricity of the horizontal microchannel rear end by criss-cross microchannel is led the integrated microelectrode of detection chip, with the conductance signal of the electrophoretic separation of the qinghaosu that detects, flow to electricity by signal wire and lead testing circuit and detect.Last electricity is led testing circuit electricity is led the conductance signal of the qinghaosu that the integrated microelectrode of detection chip detects, and after data acquisition circuit converts digital conductance signal to, sends computing machine to by signal system.
(3) artemislnin content calculates
(2) step finish after, the electricity of digital conductance signal V ' (electricity that is qinghaosu in the qinghaosu extract is led response) substitution qinghaosu standard items in (2) step that computing machine is received is led among the linear standard curve equation V=aC+b of the relation foundation between response V and its concentration C, calculate the content C ' of qinghaosu in the tested sweet wormwood, and preserve, export and show.
The present invention adopts technique scheme, mainly contains following effect:
1. the present invention is integrated with electrophoresis detection technology and microfluidic analysis chip, simple in structure, and has realized that effectively integrated electricity leads the microminaturization of the micro-fluidic chip pick-up unit of detection, and good portability is easy to use, can realize fast detecting.
2. the present invention serves as to extract behind the solvent rapid extraction qinghaosu direct injected to integrated electricity to lead in the integrated micro-fluidic chip of detection chip and carry out analyzing and testing with chloroform, ethyl acetate, normal hexane organic solvent, method is simple, has avoided the derivatization process in the conventional qinghaosu detection.And the leaching process time is short, efficient is high, and the extraction solvent load of use is few, and the test sample consumption is few, detects fast and accurately.Realize the continuity of the content analysis of qinghaosu in the raw material, simplified operation steps, greatly shortened the detection time of qinghaosu, can in 0.5 hour, finish the fast detecting of the content of qinghaosu.
3. adopt the integrated electricity of chip to lead the pattern of detection,, accurately and fast obtain the content of qinghaosu in the sweet wormwood raw material by calibration curve method.
The present invention can be widely used in the analyzing and testing of other active ingredient of natural product, is specially adapted to the detection of artemislnin content in the sweet wormwood raw material, especially is convenient to fast detecting.
Four. description of drawings
Fig. 1 is the theory diagram of apparatus of the present invention;
Fig. 2 is apparatus of the present invention microfluidic chip structure synoptic diagram;
Fig. 3 is a microelectrode structure for amplifying synoptic diagram in the present embodiment 1;
Fig. 4 is the chip conductance signal response diagram of detected qinghaosu among the embodiment 1.
Among the figure: 1 power supply, 2 micro-fluidic chips, 3 testing circuits, 4 data acquisition circuits and computing machine, 5 glass substrates, 6 dimethyl silicone polymers (PDMS) cover plate, 7 horizontal microchannels, 8 vertical microchannels, 9 delivery holes, 10 microelectrodes.
Five. embodiment
Below in conjunction with embodiment, further specify the present invention.
Embodiment 1
Shown in Fig. 1～3, the microchip detector of artemislnin content in a kind of sweet wormwood mainly comprises power supply 1, micro-fluidic chip 2, testing circuit 3, data acquisition circuit and computing machine 4.
Described power supply 1 is the micro-fluidic chip small intelligent high-voltage power supply of XCDY for commercial model, and input voltage is civil power 220V, and output voltage is that 0～8000KV is adjustable, and electric current is that 0～500 μ Ak is adjustable.Power supply 1 is connected with two platinum electrode pins by concentric cable, in order to supply with the separation voltage of the qinghaosu extract in the criss-cross microchannel.
Described micro-fluidic chip 2 is mainly led the integrated microelectrode of detection chip 10, dimethyl silicone polymer (PDMS) cover plate 6 formations by glass substrate 5, electricity for being integrated with the micro-fluidic chip 2 that electricity is led detection chip integrated microelectrode 10 and criss-cross microchannel.Its glass substrate 5 is the substrate of electrophoresis chip, and the length of glass substrate 5 is that 40000 μ m, width are that 20000 μ m, thickness are 500 μ m.In an end of glass substrate 5 and be positioned at the terminal position of the horizontal microchannel 6 of criss-cross microchannel, directly gold (Au) is deposited on the glass substrate 5 and forms electricity and lead the integrated microelectrode of detection chip 10.Electricity is led the integrated microelectrode of detection chip 10 and is made up of 2 groups of electrode pairs, and the length of every pair of electrode is that 3000 μ m, width are that 20 μ m, thickness are 10 μ m, and the spacing between the every pair of electrode is 20 μ m.And to cover a layer thickness at the outside surface of microelectrode 10 be 0.01 μ m silicon dioxide insulator thin layer, to guarantee the isolation between the buffer solution and detected liquid in microelectrode 10 and the microchannel, avoids microelectrode 10 corrosion failures.At the two ends of microelectrode 10, be connected by the input end of pin respectively with testing circuit 3, feed back to testing circuit 3 in order to conductance signal with qinghaosu.On glass substrate 5, be coated with the dimethyl silicone polymer that a layer thickness is 1000 μ m (PDMS) cover plate 6.On dimethyl silicone polymer (PDMS) cover plate 6, be provided with the criss-cross microchannel that utilizes the SU-8 anode membrane to make by solid casting method, the width of microchannel is that 30 μ m, the degree of depth are 10 μ m, length along the horizontal microchannel 7 of glass substrate length direction is 5000 μ m, is 1000 μ m along the length of vertical microchannel 8 of glass substrate Width.Two ends at horizontal, vertical microchannel 7,8, the delivery hole 9 that volume is set is 3 μ L respectively is (promptly in the end of criss-cross microchannel, be provided with A, B, four delivery holes 9 of C, D altogether), in order to ensure in the criss-cross microchannel of micro-fluidic chip the supply of buffer solution and detected liquid.Micro-fluidic chip 2 is fixedly connected on the testing circuit 3 by pin.
Described testing circuit 3 is led testing circuit for ordering integrated electricity, mainly comprises by I/V change-over circuit, multiplying operational circuit, low-pass filter circuit, differential amplifier circuit constituting, and is integrated on the pcb board.Electricity is led testing circuit 3 and data acquisition circuit and computing machine 4 and is connected by lead.Testing circuit provides 220V power supply by commercial power supply (model is 4NIC-X39).
Described data acquisition circuit 4 is for ordering integrated A/D signal acquisition circuit, and described computing machine 4 is commercial computing machine.Data acquisition circuit is connected with computing machine by serial ports, by the conductance signal capture program conductance signal is transformed into digital signal, calculate, and output and preservation data, display result.
The microchip detector of artemislnin content in a kind of sweet wormwood, with embodiment 1, wherein: the length of glass substrate 5 is that 50000 μ m, width are that 30000 μ m, thickness are 1000 μ m.Metal platinum is deposited on the electric integrated microelectrode of detection chip (10) of leading of formation on the glass substrate.Electricity is led the microelectrode 10 of detection and is made up of 4 groups of electrode pairs, and the length of every pair of electrode is that 5000 μ m, width are that 100 μ m, thickness are 50 μ m, and the spacing between the every pair of electrode is 100 μ m.The silicon dioxide insulator thickness of thin layer that microelectrode 10 outside surfaces cover is 0.1 μ m.The thickness of dimethyl silicone polymer (PDMS) cover plate 6 is 1500 μ m.The width of microchannel is that 80 μ m, the degree of depth are 30 μ m, and laterally the length of microchannel 7 is 15000 μ m, and vertically the length of microchannel 8 is 3000 μ m.The volume of delivery hole 9 is 5 μ L.
Embodiment 3
The inspection microchip of artemislnin content is surveyed device in a kind of sweet wormwood, and with embodiment 1, wherein: the length of glass substrate 5 is that 45000 μ m, width are that 25000 μ m, thickness are 800 μ m.Platinum evanohm (Pt-Cr) is deposited on the electric integrated microelectrode of detection chip (10) of leading of formation on the glass substrate.Electricity is led the microelectrode 10 of detection and is made up of 3 groups of electrode pairs, and the length of every pair of electrode is that 4000 μ m, width are that 60 μ m, thickness are 30 μ m, and the spacing between the every pair of electrode is 60 μ m.The silicon dioxide insulator thickness of thin layer that microelectrode 10 outside surfaces cover is 0.05 μ m.The thickness of dimethyl silicone polymer (PDMS) cover plate 6 is 1200 μ m.The width of microchannel is 50 μ m, and the degree of depth is 20 μ m, and laterally the length of microchannel 7 is 10000 μ m, and vertically the length of microchannel 8 is 2000 μ m.The volume of delivery hole 9 is 4 μ L.
Embodiment 4
The concrete steps of the microchip detection method of artemislnin content are as follows in a kind of sweet wormwood raw material:
(1) qinghaosu extract preparation
With the sweet wormwood is raw material, is to extract solvent with the chloroform organic solvent.Earlier with the sweet wormwood raw material pulverizing and after 40 mesh sieves sieve, again in the quality (g) of sweet wormwood: the ratio of volume of organic solvent (ml) is 1: 6 a ratio, sweet wormwood and organic solvent are positioned in the extraction element, after 1min is extracted in jolting at ambient temperature, standing demix 1min, collect the upper strata stillness of night with the glass injection-tube, abandon lower floor's dregs of a decoction.The supernatant liquor of collecting is the qinghaosu extract.
(2) the micro-fluidic chip electricity of qinghaosu is led detection
(1) step finish after, earlier in the criss-cross microchannel of apparatus of the present invention, pass through glass syringe, inject trishydroxymethylaminomethane-borate buffer solution, after with (1) the qinghaosu extract prepared of step, the glass syringe that miillpore filter is housed by front end injects two delivery holes 9 (being C, D delivery hole 9) at vertical microchannel 8 two ends of criss-cross microchannel.Then by two platinum electrode pins and two concentric cable, the output terminal of power supply is connected with two delivery holes 9 (being A, B delivery hole 9) at horizontal microchannel 7 two ends of the criss-cross microchannel of micro-fluidic chip 2 respectively, qinghaosu extract in the criss-cross microchannel is under 300V voltage, carry out electrophoretic separation, and the electricity of horizontal microchannel 7 rear ends by the criss-cross microchannel microelectrode 10 of leading detection, with the conductance signal of the electrophoretic separation of the qinghaosu that detects, flow to electricity by signal wire and lead testing circuit 3 and detect.Last electricity is led testing circuit 3 electricity is led the conductance signal of the qinghaosu that the microelectrode of detection detects, and after data acquisition circuit 4 converts digital conductance signal to, sends computing machine to by signal system.
(3) artemislnin content calculates
(2) step finish after, with computing machine receive (2) step digital conductance signal V ' (electricity that is qinghaosu in the qinghaosu extract is led response) be 2.58 * 10
5The electricity of substitution qinghaosu standard items is led linear standard curve equation V=1.70 * 10 that the relation between response V and its concentration C is set up
5C+883.6, R
2In=0.9837, the content C ' that calculates qinghaosu in the tested sweet wormwood is 0.91%, and preserves, exports and show.
The concrete steps of the microchip detection method of artemislnin content in a kind of sweet wormwood raw material, with embodiment 4, wherein:
In (1) step, extracting solvent is the ethyl acetate organic solvent, and the sweet wormwood raw material sieves through 60 orders, and the quality of sweet wormwood (g) is 1: 8 with the ratio of volume of organic solvent (ml), and 3min, standing demix 3min are extracted in jolting.
In (2) step, buffer solution is potassium dihydrogen phosphate-sodium dihydrogen phosphate, carries out electrophoretic separation under 900V voltage.
In (3) step, (2) the digital conductance signal V ' that goes on foot that computing machine is received is 1.76 * 10
5The electricity of substitution qinghaosu standard items is led linear standard curve equation V=1.70 * 10 that the relation between response V and its concentration C is set up
5C+883.6, R
2In=0.9837, the content C ' that calculates qinghaosu in the tested sweet wormwood by program is 0.83%.
The concrete steps of the microchip detection method of artemislnin content in a kind of sweet wormwood raw material, with embodiment 4, wherein:
In (1) step, extracting solvent is the normal hexane organic solvent, and the sweet wormwood raw material sieves through 50 orders, and the quality of sweet wormwood (g) is 1: 7 with the ratio of volume of organic solvent (ml), and 2min, standing demix 2min are extracted in jolting.
In (2) step, buffer solution is borax-boric acid, carries out electrophoretic separation under 600V voltage.
In (3) step, (2) the digital conductance signal V ' that goes on foot that computing machine is received is 2.11 * 10
5The electricity of substitution qinghaosu standard items is led linear standard curve equation V=1.70 * 10 that the relation between response V and its concentration C is set up
5C+883.6, R
2In=0.9837, the content C ' that calculates qinghaosu in the tested sweet wormwood by program is 0.87%.
Claims (8)
1. the microchip detector of artemislnin content in the sweet wormwood, mainly comprise power supply (1), testing circuit (3), data acquisition circuit and computing machine (4), it is characterized in that micro-fluidic chip (2) in addition, described micro-fluidic chip (2): mainly by glass substrate (5), electricity is led the integrated microelectrode of detection chip (10), dimethyl silicone polymer cover plate (6) constitutes, its glass substrate (5) is the substrate of electrophoresis chip, the length of glass substrate is 40000～50000 μ m, width is 20000～30000 μ m, thickness is 500～1000 μ m, in an end of glass substrate and be positioned at the terminal position of the horizontal microchannel (6) of criss-cross microchannel, directly metallic gold or platinum or platinum chromium alloy material being deposited on glass substrate (5) goes up and forms electricity and lead the integrated microelectrode of detection chip (10), electricity is led the integrated microelectrode of detection chip (10) and is made up of 2～4 groups of electrode pairs, the length of every pair of electrode is 3000～5000 μ m, width is 20～100 μ m, thickness is 10～50 μ m, spacing between the every pair of electrode is 20～100 μ m, and cover the silicon dioxide insulator thin layer that a layer thickness is 0.01～0.1 μ m at the outside surface of microelectrode (10), two ends in microelectrode (10), be connected by the input end of pin respectively with testing circuit (3), on glass substrate (5), being coated with a layer thickness is the dimethyl silicone polymer cover plate (6) of 1000～1500 μ m, on dimethyl silicone polymer cover plate (6), be provided with the criss-cross microchannel that utilizes the SU-8 anode membrane to make by solid casting method, the width of microchannel is 30～80 μ m, the degree of depth is 10～30 μ m, length along the horizontal microchannel (7) of glass substrate length direction is 5000～15000 μ m, length along vertical microchannel (8) of glass substrate Width is 1000～3000 μ m, at horizontal stroke, vertical microchannel (7,8) two ends, the delivery hole that volume is 3～5 μ L (9) is set respectively, promptly in the end of criss-cross microchannel, be provided with A altogether, B, C, four delivery holes of D (9), micro-fluidic chip (2) is fixedly connected on (3) on the testing circuit by pin.
2. according to the microchip detector of artemislnin content in the described sweet wormwood of claim 1, the length that it is characterized in that glass substrate (5) is 40000 μ m, width is 20000 μ m, thickness is 500 μ m, metallic gold is deposited on the electric integrated microelectrode of detection chip (10) of leading of formation on the glass substrate, this microelectrode (10) is made up of 2 groups of electrode pairs, the length of every pair of electrode is 3000 μ m, width is 20 μ m, thickness is 10 μ m, spacing between the every pair of electrode is 20 μ m, the silicon dioxide insulator thickness of thin layer that microelectrode (10) outside surface covers is 0.01 μ m, the thickness of dimethyl silicone polymer cover plate (6) is 1000 μ m, the width of microchannel is 30 μ m, the degree of depth is 10 μ m, laterally the length of microchannel (7) is 5000 μ m, vertically the length of microchannel (8) is 1000 μ m, and the volume of delivery hole (9) is 3 μ L.
3. according to the microchip detector of artemislnin content in the described sweet wormwood of claim 1, the length that it is characterized in that glass substrate (5) is 50000 μ m, width is 30000 μ m, thickness is 1000 μ m, metal platinum is deposited on the electric integrated microelectrode of detection chip (10) of leading of formation on the glass substrate, this microelectrode (10) is made up of 4 groups of electrode pairs, the length of every pair of electrode is 5000 μ m, width is 100 μ m, thickness is 50 μ m, spacing between the every pair of electrode is 100 μ m, the thick 0.1 μ m of silicon dioxide insulator thin layer that microelectrode (10) outside surface covers, the thickness of dimethyl silicone polymer cover plate (6) is 1500 μ m, the width of microchannel is 80 μ m, the degree of depth is 30 μ m, laterally the length of microchannel (7) is 15000 μ m, and vertically the length of microchannel (8) is 3000 μ m, and the volume of delivery hole (9) is 5 μ L.
4. according to the microchip detector of artemislnin content in the described sweet wormwood of claim 1, the length that it is characterized in that glass substrate (5) is 45000 μ m, width is 25000 μ m, thickness is 800 μ m, the platinum evanohm is deposited on the electric integrated microelectrode of detection chip (10) of leading of formation on the glass substrate, this microelectrode (10) is made up of 3 groups of electrode pairs, the length of every pair of electrode is 4000 μ m, width is 60 μ m, thickness is 30 μ m, spacing between the every pair of electrode is 60 μ m, the thick 0.05 μ m of silicon dioxide insulator thin layer that microelectrode (10) outside surface covers, the thickness of dimethyl silicone polymer cover plate (6) is 1200 μ m, the width of microchannel is 50 μ m, the degree of depth is 20 μ m, laterally the length of microchannel (7) is 10000 μ m, vertically the length of microchannel (8) is 2000 μ m, and the volume of delivery hole (9) is 4 μ L.
5. the microchip detection method of artemislnin content in the sweet wormwood raw material is characterized in that concrete steps are as follows:
(1) qinghaosu extract preparation
With the sweet wormwood is raw material, with organic solvents such as chloroform or ethyl acetate or normal hexanes is to extract solvent, earlier with the sweet wormwood pulverizing and after 40～60 mesh sieves sieve, be 1: 6～8 ratio again in the ratio of the quality of sweet wormwood and volume of organic solvent, sweet wormwood and organic solvent are positioned in the extraction element, after 1～3min is extracted in jolting at ambient temperature, standing demix 1～3min, collect the upper strata stillness of night with the glass injection-tube, abandon lower floor's dregs of a decoction;
(2) the micro-fluidic chip electricity of qinghaosu is led detection
(1) step finish after, earlier in the criss-cross microchannel of apparatus of the present invention, pass through glass syringe, inject buffer solution, be trishydroxymethylaminomethane-boric acid or potassium dihydrogen phosphate-sodium dihydrogen phosphate or borax-boric acid, after with (1) the qinghaosu extract prepared of step, the glass syringe of miillpore filter is housed by front end, inject two delivery holes (9) at vertical microchannel two ends of criss-cross microchannel, be C, in the D delivery hole (9), then by two platinum electrode pins and two concentric cable, with the output terminal of power supply respectively with two delivery holes at horizontal microchannel (7) two ends of the criss-cross microchannel of micro-fluidic chip (2), be A, B delivery hole (9) connects, qinghaosu extract in the criss-cross microchannel is under 300～900V voltage, carry out electrophoretic separation, and the electricity of horizontal microchannel (7) rear end by criss-cross microchannel is led the integrated microelectrode of detection chip (10), conductance signal with the electrophoretic separation of the qinghaosu that detects, flowing to electricity by signal wire leads testing circuit (3) and detects, last electricity is led the conductance signal that testing circuit (3) is led electricity the qinghaosu that the microelectrode (10) of detection detects, after data acquisition circuit (4) converts digital conductance signal to, send computing machine (4) to by signal system;
(3) artemislnin content calculates
(2) step finish after, the electricity of digital conductance signal V ' substitution qinghaosu standard items in (2) step that computing machine is received is led among the linear standard curve equation V=aC+b of the relation foundation between response V and its concentration C, calculate the content C ' of qinghaosu in the tested sweet wormwood, and preserve, export and show.
6. according to the concrete steps of the microchip detection method of artemislnin content in the described sweet wormwood raw material of claim 5, it is characterized in that:
In (1) step, extracting solvent is the chloroform organic solvent, and the sweet wormwood raw material sieves through 40 orders, and the quality of sweet wormwood is 1: 6 with the ratio of volume of organic solvent, and 1min, standing demix 1min are extracted in jolting;
In (2) step, buffer solution is trishydroxymethylaminomethane-boric acid, carries out electrophoretic separation under 300V voltage.
7. according to the concrete steps of the microchip detection method of artemislnin content in the described sweet wormwood raw material of claim 5, it is characterized in that:
It is characterized in that extracting solvent is the ethyl acetate organic solvent in (1) step, the sweet wormwood raw material sieves through 60 orders, and the quality of sweet wormwood is 1: 8 with the ratio of volume of organic solvent, and 3min, standing demix 3min are extracted in jolting;
In (2) step, buffer solution is potassium dihydrogen phosphate-sodium dihydrogen phosphate, carries out electrophoretic separation under 900V voltage.
8. according to the concrete steps of the microchip detection method of artemislnin content in the described sweet wormwood raw material of claim 5, it is characterized in that:
It is characterized in that extracting solvent is the normal hexane organic solvent in (1) step, the sweet wormwood raw material sieves through 50 orders, and the quality of sweet wormwood is 1: 7 with the ratio of volume of organic solvent, and 2min, standing demix 2min are extracted in jolting;
In (2) step, buffer solution is borax-boric acid, carries out electrophoretic separation under 600V voltage.
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