CN102116758B - Tetracycline medicament selective electrode and preparation method thereof - Google Patents
Tetracycline medicament selective electrode and preparation method thereof Download PDFInfo
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- CN102116758B CN102116758B CN 200910156697 CN200910156697A CN102116758B CN 102116758 B CN102116758 B CN 102116758B CN 200910156697 CN200910156697 CN 200910156697 CN 200910156697 A CN200910156697 A CN 200910156697A CN 102116758 B CN102116758 B CN 102116758B
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Abstract
The invention discloses a tetracycline medicament selective electrode and a preparation method thereof. The electrode comprises a quartz glass tube containing a molecularly imprinted monolithic column with an electroactive substance, an internal filling liquid, an internal reference electrode, an electrode stem and epoxy resin. By increasing the thickness of a sensitive film, using a porous monolithic column material, efficiently binding tetracycline medicaments in the electroactive substance to the sensitive film and optimizing the internal filling liquid, the detection limit of the electrode is reduced to the order of magnitude of 10<-9> mol/L. Because the molecularly imprinted monolithic column has a specific recognition action on the tetracycline medicaments, the selectivity is high, and common interfering substances have no influence on the potential measurement of the electrode. By using the tetracycline medicament selective electrode provided by the invention as an indicator electrode and a saturated calomel electrode or a Pt electrode as a reference electrode, the tetracycline medicaments can be detected with high sensitivity and high selectivity by using the potential analysis method, equipment is simple, the analysis cost is low, and the residual analysis of the tetracycline medicaments can be realized.
Description
Technical field
The present invention relates to Electroanalytical Chemistry, chemical sensor field, be specifically related to a kind of preparation method of selective electrode of tetracycline drugs.
Background technology
Tetracycline medication (Tetracycline, TCs) be a class extensive pedigree antibiotic, have ten dihydro aphthacene based structures, be mainly used in clinically treating the pathogenic former infection that causes such as Gram-positive and negative bacterium, mycoplasma, Chlamydia, Richettsia and conveyor screw etc.TCs mainly comprises two large classes, be that natural Tetracyclines comprises tetracycline (TC), terramycin (OTC), aureomycin (CTC), demethylchlortetra cylinum (DMC) etc., comprise methacycline (MTC), fortimicin (DC), minocycline (MNC) etc. with semi-synthetic Tetracyclines, wherein the most frequently used TCs is TC, OTC, CTC, DC.The advantages such as it is low with price, has a broad antifungal spectrum are widely used as feed addictive in aquaculture, with prevention, control and treatment Animal diseases.The long-term TCs that takes in accumulates in human body gradually, can cause acute and chronic poisoning, finally causes the pathology of multiple organ, thus many countries to the strict routine monitoring of the residual enforcement of TCs, the maximum residue limit of TCs is 0.1mg/kg in China's regulation animal derived food.
The common method that detects at present TCs has: high performance liquid chromatography, capillary electrophoresis, euzymelinked immunosorbent assay (ELISA), microbial method etc.1. high performance liquid chromatography (HPLC): be the method for most widely used quantitative detection TCs, have the characteristic of efficient separation and high-sensitivity detection.The method weak point is that sample pre-treatments operating personnel, instrument complicated, need to be professional be expensive, analysis cost is high, analysis speed is slow, be unwell to Site Detection.2. capillary electrophoresis (CE): have the advantages that analysis speed is fast, sample size is few, but instrument is expensive, need operating personnel, stability and the precision of specialty to have much room for improvement.3. euzymelinked immunosorbent assay (ELISA) (ELISA): in rapid screening, be most widely used, sensitivity higher (about 0.001～0.005mg/kg), be applicable to on-the site analysis and industrialization and detect, it is existing at present that to detect the residual kit of TCs on sale.Main weak point is: be not suitable for coloured, muddy, viscous fluid, be prone to false positive results.4. microbial method: be the method for the primary dcreening operation TCs of widespread use, simple to operate, with low cost, be fit to the gross sample analysis, can reflect biological value, (detection limit＞3mg/kg), poor selectivity, there is cross reaction but sensitivity is low.
Potentiometry based on ion-selective electrode has many merits: equipment needed thereby is simple, is convenient to Site Detection; Analysis speed is very fast; Coloured, muddy, viscous fluid can be used for, generally Chemical Decomposition need be do not carried out, simple to operate rapid; Higher selectivity is arranged.Major defect is: detectability higher (10
-6～10
-5Mol/L), can not be used for the extremely low concentration sample analysis, greatly limit its range of application.Adopt the selective Electrodes Method For The Determination tetracycline medication, existing forefathers' report (Microchem.J., 1987,36 (1): 107, straits medical science, 1994,4 (3): 23.), but sensitivity is all lower, and detectability only is equivalent to 0.9～2.2mg/kg approximately, is difficult to reach the analysis requirement.
Molecular imprinting is that " lock and key model ", the antibody that combine the enzyme-to-substrate effect forms theory and " selectivity adsorption theory " and a novel preparation technology growing up.Molecularly imprinted polymer (the Molecular Imprinting Polymers of this technology preparation, MIPs), have that compatibility and selectivity are good, the advantages such as the anti-adverse environment ability is strong, good stability, long service life and applied range, be widely used in chromatographic resolution, mimetic enzyme catalysis, clinical medicine analysis, sensing technology, film separates and the field such as Solid-Phase Extraction.Integral post typically refers to the bar-shaped block polymer that obtains by the reaction reagent in-situ polymerization, has high-permeability, poriness and large specific surface area.The molecular imprinting of high selectivity is combined with the integral post isolation technics of high-permeability, and (Talanta 2009,79:926 for the existing relevant report of preparation tetracycline medication molecular engram integral column; Chinese patent, application number: 200810052541.3), but all as Solid-Phase Extraction material or liquid chromatography stuffing.Pretsch etc. have prepared highly sensitive metalloform-selective electrode based on integral post, and detectability reaches 10
-9Mol/L (Anal.Chem.2005,77:3966).Have not yet to see based on the preparation method of the electrodes selective of molecular engram integral column and the relevant report of application thereof.
For above-mentioned several mensuration TCs methods deficiency separately, the present invention proposes a kind of preparation method of the selective electrode of tetracycline drugs based on molecular engram integral column, based on this electrodes selective, adopt potentiometry, the in high sensitivity tetracycline medication in the test sample.The sensitive membrane of traditional selective electrode of tetracycline drugs is for being dissolved with Polyvinylchloride (PVC) film of electroactive material (tetracycline medication-sodium phosphotungstate associated matter or tetracycline medication-tetraphenylboron sodium associated matter), (general＜10 when sample concentration is extremely low
-6Mol/L), the ionic flux that tetracycline medication spreads from sensitive membrane to testing sample solution occurs, cause the ion concentration to be measured on sensitive membrane surface to be higher than bulk concentration and can't further to reduce detectability.The selective electrode of tetracycline drugs that the present invention proposes, adopt following three technological means to suppress above-mentioned diffusion ion flux: to be synthetic tetracycline medication molecular engram integral column in the quartz glass tube of 1～3mm at diameter 1., behind the wash-out template molecule of tetracycline class medicine, the PVC solution that is dissolved with electroactive material is sucked this molecular engram integral column, form sensitive material after volatilizing solvent.Because the thickness substance of sensitive membrane increases to more than 10 times of traditional sensitive membrane, and the porous structure of integral post limited the diffusive migration behavior of tetracycline medication in film, so greatly reduce ionic diffusion coefficient; 2. the specific binding site of tetracycline medication is contained in the molecular engram integral column inside behind the wash-out template molecule, can efficiently bind the tetracycline medication in the electroactive material; 3. add in the internal-filling liquid can with the Ca of tetracycline medication complexing
2+, reduced its Cf, the cross-film ionic flux that water wettability causes the common transmission of ion in establishment tetracycline medication and the internal-filling liquid.So the detectability of selective electrode of tetracycline drugs is reduced to 10
-9The mol/L order of magnitude is lower than 0.005mg/L (mg/kg), and sensitivity is higher; Because molecular engram integral column is to the specific recognition of template molecule, so selectivity is higher, the common interference material on potential measurement all without affecting; The range of linearity is wider, is 1 * 10
-8～1 * 10
-3Mol/L; But direct-detection colored solutions; Detection speed is very fast, and low concentration sample can be finished in 2 hours; Equipment needed thereby is simple, is convenient to Site Detection.The preparation method of above-mentioned selective electrode of tetracycline drugs based on molecular engram integral column is not reported in other documents or patent.
Summary of the invention
Primary technical matters to be solved by this invention is the deficiency that overcomes the several method of tetracycline medication content in the above-mentioned existing test sample, comprise: sample pre-treatments is complicated, the operating personnel that need specialty, instrument is expensive, analysis cost is high, analysis speed is slow, detection time is long, be unwell to Site Detection, stability and precision are not good, and sample itself is had relatively high expectations, be not suitable for coloured, muddy, viscous fluid, be prone to false positive, as a result sensitivity is lower, and (detection limit＞3mg/kg), specificity is not strong etc., a kind of selective electrode of tetracycline drugs based on molecular engram integral column is provided, it is highly sensitive, selectivity is high, but direct-detection has color solution, and detection speed is very fast; Preci-sion and accuracy is higher; Institute's use instrument is simple, and analysis cost is cheap.
Another technical matters to be solved by this invention provides relevant preparation method.
For achieving the above object, the present invention for the technical scheme that solves primary technical matters and propose is:
A kind of selective electrode of tetracycline drugs based on molecular engram integral column is characterized in that: this electrode comprises and includes quartz glass tube, internal-filling liquid, internal reference electrode, electrode stem, the epoxy resin that sucks the molecular engram integral column that electroactive material is arranged; The outer wall stage casing that includes the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged is bonded at the electrode stem lower end, and sealing abutting edge, add in the electrode stem internal-filling liquid is arranged, internal reference electrode electrode insertion bar also is immersed in the internal-filling liquid, with fixing internal reference electrode and the enclosed electrode bar upper end of epoxy resin; Freshly prepd electrode places 10
-6～10
-8Activate in the mol/L quadracycline class drug solution.
Described tetracycline medication comprises tetracycline (TC), terramycin (OTC), aureomycin (CTC) or fortimicin (DC).
Described internal reference electrode is Pt electrode or Ag/AgCl electrode.
The described quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged that includes prepares by the following method:
1. template molecule of tetracycline class medicine, function monomer methacrylic acid are dissolved in the mixed solution of methyl alcohol and acetone, ultrasonic dissolution adds pore-foaming agent, behind stirring 6～8h, add crosslinking chemical ethylene glycol dimethacrylate and initiating agent azoisobutyronitrile, continue to be stirred to dissolving; Wherein, when template molecule was 0.1mmol, the mixed liquor volume of methyl alcohol and acetone was 0.5～1.5mL, and the volume of the shared mixed solution of methyl alcohol is: 45～100%; The mol ratio of template molecule of tetracycline class medicine, function monomer, crosslinking chemical is 1: 2～10: 20～50; The cumulative volume of function monomer, crosslinking chemical and the volume ratio of pore-foaming agent are 1～2: 3～4; Pore-foaming agent is the mixed solution of toluene and lauryl alcohol, and wherein the shared volume of toluene is 10～20%; Initiator amount is 8～20% of function monomer quality;
2. with behind the logical nitrogen 5～10min of said mixture, inserting internal diameter is that 1～3mm, length are in the quartz glass tube of 2～5cm, under nitrogen protection, with the quartz glass channel closure, place polyase 13～8h under the uviol lamp, obtain including the quartz glass tube of tetracycline medication molecular engram integral column;
8. intercepted length is the above-mentioned quartz glass tube that includes the tetracycline medication molecular engram integral column that makes of 0.5～1cm, soxhlet extraction is removed template molecule of tetracycline class medicine, 45～60 ℃ of drying 12～36h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline medication; Wherein the used extractant of soxhlet extraction is pure methyl alcohol;
4. above-mentioned quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline medication is immersed and be dissolved with in the uniform mix of electroactive material, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged.
The described synthetic method that is dissolved with the uniform mix of electroactive material, step is as follows:
Get dry electroactive material, PVC, tetrahydrofuran and plastifier and join in the test tube, after the sealing, blend together uniform mix with the vortex oscillator; Wherein, the mass ratio of electroactive material, PVC and plastifier is 0.5～10: 30～35: 30～70; When the quality of PVC was 1mg, the volume of tetrahydrofuran was 30～80 μ L; Plastifier is a kind of of following plastifier: dibutyl phthalate (DBP), dioctyl phthalate (DOP), dinonyl phthalate (DNP), Plexol 201 (DOS), O-Nitrophenylfluorone Octyl Ether (o-NPOE).
The preparation method of described electroactive material, step is:
With 0.005～0.02mol/L quadracycline class drug solution and 0.005～0.02mol/L Salkowski's solution in molar ratio 3～4: 1 or 0.005～0.02mol/L quadracycline class drug solution with mix under 0.005～0.02mol/L sodium tetraphenylborate solution 1: 1 in molar ratio～1.5 stirs, leave standstill 10～12h, filter, after sediment concentration is the salt acid elution 5～8 times of 0.01～0.05mol/L, 45～60 ℃ of oven dry namely get dry electroactive material.
Described internal-filling liquid (2) is: contain 10
-3～10
-6The quadracycline class medicine, 10 of mol/L
-1～10
-3The NaCl of mol/L and 10
-1～10
-3The CaCl of mol/L
2Mixed solution.
The present invention is that the technical scheme that second technical matters of solution taked is: a kind of preparation method of the selective electrode of tetracycline drugs based on molecular engram integral column, it is characterized in that: the outer wall stage casing that will include the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged is bonded at the electrode stem lower end, and sealing abutting edge, add internal-filling liquid in the electrode stem, with internal reference electrode electrode insertion bar and be immersed in the internal-filling liquid, with the fixing internal reference electrode and with end closure on the electrode stem of epoxy resin, freshly prepd electrode being placed concentration at last is 10
-6～10
-8Activate 48～96h in the quadracycline class drug solution of mol/L, thereby obtain the selective electrode of tetracycline drugs based on molecular engram integral column;
The employed preparation method who includes the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged, step is as follows:
1. template molecule of tetracycline class medicine, function monomer methacrylic acid are dissolved in the mixed solution of methyl alcohol and acetone, ultrasonic dissolution adds pore-foaming agent, behind stirring 6～8h, add crosslinking chemical ethylene glycol dimethacrylate and initiating agent azoisobutyronitrile, continue to be stirred to dissolving; Wherein, when template molecule was 0.1mmol, the mixed liquor volume of methyl alcohol and acetone was 0.5～1.5mL, and the volume of the shared mixed solution of methyl alcohol is: 45～100%; The mol ratio of template molecule of tetracycline class medicine, function monomer, crosslinking chemical is 1: 2～10: 20～50; The cumulative volume of function monomer, crosslinking chemical and the volume ratio of pore-foaming agent are 1～2: 3～4; Pore-foaming agent is the mixed solution of toluene and lauryl alcohol, and wherein the shared volume of toluene is 10～20%; Initiator amount is 8～20% of function monomer quality;
2. with behind the logical nitrogen 5～10min of said mixture, inserting internal diameter is that 1～3mm, length are in the quartz glass tube of 2～5cm, under nitrogen protection, with the quartz glass channel closure, place polyase 13～8h under the uviol lamp, obtain including the quartz glass tube of tetracycline medication molecular engram integral column;
3. intercepted length is the above-mentioned quartz glass tube that includes the tetracycline medication molecular engram integral column that makes of 0.5～1cm, soxhlet extraction is removed template molecule of tetracycline class medicine, 45～60 ℃ of drying 12～36h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline medication; Wherein the used extractant of soxhlet extraction is pure methyl alcohol;
4. above-mentioned quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline medication is immersed and be dissolved with in the uniform mix of electroactive material, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged.
The described preparation method who is dissolved with the uniform mix of electroactive material, step is followed successively by:
Get dry electroactive material, PVC, tetrahydrofuran and plastifier and join in the test tube, after the sealing, blend together uniform mix with the vortex oscillator; Wherein, the mass ratio of electroactive material, PVC and plastifier is 0.5～10: 30～35: 30～70; When the quality of PVC was 1mg, the volume of tetrahydrofuran was 30～80 μ L; Plastifier is a kind of of following plastifier: dibutyl phthalate (DBP), dioctyl phthalate (DOP), dinonyl phthalate (DNP), Plexol 201 (DOS), O-Nitrophenylfluorone Octyl Ether (o-NPOE).
Described electroactive material is synthetic by following methods:
With 0.005～0.02mol/L quadracycline class drug solution and 0.005～0.02mol/L Salkowski's solution in molar ratio 3～4: 1 or 0.005～0.02mol/L quadracycline class drug solution with mix under 0.005～0.02mol/L sodium tetraphenylborate solution 1: 1 in molar ratio～1.5 stirs, leave standstill 10～12h, filter, after sediment concentration is the salt acid elution 5～8 times of 0.01～0.05mol/L, 45～60 ℃ of oven dry namely get dry electroactive material.
Described internal-filling liquid is: contain 10
-3～10
-6The quadracycline class medicine, 10 of mol/L
-1～10
-3The NaCl of mol/L and 10
-1～10
-3The CaCl of mol/L
2Mixed solution.
Compared with prior art, the invention has the advantages that to have following technique effect:
(1) selective electrode of tetracycline drugs that proposes among the present invention, thickness by increasing sensitive membrane, use the porous integral column material, efficiently bind tetracycline medication in the electroactive material in sensitive membrane, optimize internal-filling liquid and form, establishment the diffusion ion flux of tetracycline medication, reduce the electrode detection limit to 10
-9The mol/L order of magnitude is lower than 0.005mg/L (mg/kg), and sensitivity is higher.
(2) because molecular engram integral column has the specific recognition effect to template molecule of tetracycline class medicine, so selectivity is higher, the common interference material on the potential measurement of electrode all without impact.
(3) selective electrode of tetracycline drugs that proposes in the present invention is as indicator electrode, adopt the potentiometric analysis method, can high sensitivity, highly selective detects tetracycline medication, detection speed is very fast, can detect colored solutions, equipment needed thereby is simple, and analysis cost is cheap, can realize the Site Detection of tetracycline medication.
Description of drawings
Fig. 1 is the selective electrode of tetracycline drugs synoptic diagram based on molecular engram integral column;
Fig. 2 a-i is the working curve diagram of the selective electrode of tetracycline drugs of a series of different preparation methods' acquisitions.
Embodiment
The below elaborates to embodiments of the invention: present embodiment is implemented under take technical solution of the present invention as prerequisite, provided detailed embodiment and concrete operating process, but protection scope of the present invention is not limited only to following embodiment.
The first step: include the preparation of the quartz glass tube of the molecular engram integral column behind the wash-out tetracycline, step is as follows:
1. tetracycline 0.1mmol, methacrylic acid 0.6mmol are added in the mixed solution of 1.0mL methyl alcohol and acetone (1: 1), ultrasonic dissolution, add 0.16mL toluene and 1.44mL lauryl alcohol, after stirring 6h, add 4.5mmol ethylene glycol dimethacrylate and 8mg azoisobutyronitrile, continue to be stirred to dissolving, logical nitrogen 10min.
2. under nitrogen protection, it is that 2mm, length are in the quartz glass tube of 3cm that said mixture is added internal diameter, and with its sealing, places polymerization 6h under the uviol lamp, obtains including the quartz glass tube of tetracycline molecular engram integral column.
3. intercepted length is the above-mentioned quartz glass tube that includes the tetracycline molecular engram integral column that makes of 0.5cm, methyl alcohol is extractant, soxhlet extraction is removed tetracycline, and 60 ℃ of dry 24h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline.
Second step: include the preparation method of the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged, step is as follows:
1. the preparation method of electroactive material: with 65mL 1.0 * 10
-2Mol/L quadracycline solution and 20mL 1.0 * 10
-2The mol/L Salkowski's solution stirs lower the mixing, leaves standstill 12h, filters, and after sediment concentration was the salt acid elution 5～8 times of 0.01mol/L, 60 ℃ of oven dry namely got dry electroactive material.
2. electroactive material 35mg, the PVC 157.5mg, DBP 157.5mg, the tetrahydrofuran 6mL that get above-mentioned drying join in the test tube, after the sealing, blend together even mixed liquor with the vortex oscillator.
3. quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline that makes in the first step is immersed in the above-mentioned uniform mix, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged.
The 3rd step: the preparation of tetracycline electrodes selective, step is as follows:
1. the above-mentioned suction that makes there is the quartz glass tube outer wall stage casing of the molecular engram integral column of electroactive material to be bonded at the electrode stem lower end, and the sealing abutting edge.
2. the preparation of internal-filling liquid: preparation contains 10
-4The quadracycline of mol/L, 10
-2The NaCl of mol/L and 10
-2The CaCl of mol/L
2Mixed solution.
3. get internal-filling liquid 0.8mL and add in the electrode stem, with Ag/AgCl electrode electrode insertion bar, and be immersed in below the internal-filling liquid liquid level, with the fixing also sealing of epoxy resin, namely make the tetracycline electrodes selective, such as Fig. 1.
4. the above-mentioned electrode that makes is inserted 10
-8Activate 60h in the quadracycline solution of mol/L.
The 4th step: the drafting of working curve.
1. the preparation of standard solution: accurately take by weighing quadracycline 0.1205g, with a small amount of water-soluble solution, add 50.00mL Michaelis buffer solution (tartrate-sodium tartrate buffer solution, pH 2.8), deionized water is settled to the brown volumetric flask of 250mL, gets 10
-3The mol/L standard solution.Stepwise dilution becomes 10 again
-4～10
-9Mol/L quadracycline standard solution series.
2. take the above-mentioned tetracycline electrodes selective that makes as indicator electrode, saturated calomel electrode is contrast electrode, according to concentration order from low to high, measures successively the primary element electromotive force E that two electrodes form with potential difference meter.Take the logarithm value (lgc) of tetracycline concentration as horizontal ordinate, take electromotive force E as ordinate, the drawing curve is such as Fig. 2-a.
The 5th step: the mensuration of Electrode selectivity coefficient.
K
+, Na
+, NH
4 +, Ca
2+, Mn
2+, Zn
2+, Ba
2+, Mg
2+, Sr
2+, Fe
2+, Fe
3+, the common interference materials such as tobramycin, glycocoll, alanine, lysine, glucose, maltose, lactose measure without impact electrode potential.
Template molecule is terramycin, and other are with embodiment 1, such as Fig. 2-b.
Template molecule is aureomycin, and other are with embodiment 1, such as Fig. 2-c.
Template molecule is fortimicin, and other are with embodiment 1, such as Fig. 2-d.
The first step: include the preparation of the quartz glass tube of the molecular engram integral column behind the wash-out tetracycline.
1. tetracycline 0.1mmol, methacrylic acid 0.4mmol are added in the mixed solution of 1.0mL methyl alcohol and acetone (8: 2), ultrasonic dissolution, add 0.24mL toluene and 1.36mL lauryl alcohol, after stirring 6h, add 3mmol ethylene glycol dimethacrylate and 6mg azoisobutyronitrile, continue to be stirred to dissolving, logical nitrogen 10min.
2. under nitrogen protection, it is that 2mm, length are in the quartz glass tube of 3cm that said mixture is added internal diameter, and with its sealing, places polymerization 4h under the uviol lamp, obtains including the quartz glass tube of template molecule of tetracycline molecular engram integral column.
3. intercepted length is the above-mentioned quartz glass tube that includes the template molecule of tetracycline molecular engram integral column that makes of 0.5cm, methyl alcohol is extractant, soxhlet extraction is removed tetracycline, and 60 ℃ of dry 24h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline.
Other are with embodiment 1, such as Fig. 2-e.
The first step: include the preparation of the quartz glass tube of the molecular engram integral column behind the wash-out tetracycline.
1. tetracycline 0.1mmol, methacrylic acid 0.8mmol are added in the 1.0mL methyl alcohol, ultrasonic dissolution adds 0.16mL toluene and 1.44mL lauryl alcohol, after stirring 6h, add 2mmol ethylene glycol dimethacrylate and 10mg azoisobutyronitrile, continue to be stirred to dissolving, logical nitrogen 10min.
2. under nitrogen protection, it is that 2mm, length are in the quartz glass tube of 3cm that said mixture is added internal diameter, and with its sealing, places polymerization 4h under the uviol lamp, obtains including the quartz glass tube of template molecule of tetracycline molecular engram integral column.
3. intercepted length is the above-mentioned quartz glass tube that includes the template molecule of tetracycline molecular engram integral column that makes of 0.5cm, methyl alcohol is extractant, soxhlet extraction is removed tetracycline, and 60 ℃ of dry 24h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline.
Other are with embodiment 1, such as Fig. 2-f.
The preparation method of electroactive material: with 20mL 1.0 * l0
-2Mol/L quadracycline solution and 25mL 1.0 * 10
-2The mol/L sodium tetraphenylborate solution stirs lower the mixing, leaves standstill 12h, filters, and after sediment concentration was the salt acid elution 5～8 times of 0.01mol/L, 60 ℃ of oven dry namely got dry electroactive material.
Other are with embodiment 1, such as Fig. 2-g.
Include the preparation method of the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged, step is as follows:
1. the preparation method of electroactive material: with 65mL 1.0 * 10
-2Mol/L quadracycline solution and 20mL 1.0 * 10
-2The mol/L Salkowski's solution stirs lower the mixing, leaves standstill 12h, filters, and after sediment concentration was the salt acid elution 5～8 times of 0.01mol/L, 60 ℃ of oven dry namely got dry electroactive material.
2. electroactive material 0.5mg, the PVC 31.5mg, DOP 68mg, the tetrahydrofuran 2.5mL that get above-mentioned drying join in the test tube, after the sealing, blend together even mixed liquor with the vortex oscillator.
3. quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline that makes in the first step is immersed in the above-mentioned uniform mix, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube that sucks the molecular engram integral column that electroactive material is arranged.
Other are with embodiment 1, such as Fig. 2-h.
The preparation of internal-filling liquid: preparation contains 10
-5The quadracycline of mol/L, 10
-3The NaCl of mol/L and 10
-3The CaCl of mol/L
2Mixed solution.
Other are with embodiment 1, such as Fig. 2-i.
Application Example 10
The first step: sample preparation:
Accurately take by weighing eel dried product that 5.0g blends in 50ml teflon centrifuge tube, add 25mLMcllvaine ' s damping fluid, behind the vibration 10min, add the 5mL trichloroacetic acid solution, with the centrifugal 2min of the speed of 2000r/min, behind standing demix, the centrifugal 20min of 5000r/min, supernatant filters with filtering head, adds the 10mL damping fluid again, 2mL trichloroacetic acid solution, extract 10min through supersonic oscillations, the centrifugal 10min of 5000r/min filters, and repeats once, merges filtered fluid, be placed on the automatic evaporation and concentration instrument 45 ℃ of nitrogen and blow and be concentrated into 0.5mL, be settled to 10mL with the 0.01mol/L citric acid solution.
Second step: sample measurement:
Get the above-mentioned sample solution 1mL that handles well, add 2.00mL Michaelis buffer solution, deionized water is settled in the 10mL volumetric flask.Get mentioned solution and place measuring cell, the selective electrode of tetracycline drugs that makes take the present invention is as indicator electrode, and saturated calomel electrode is contrast electrode, measures the primary element electromotive force E that two electrodes form with potential difference meter.Concentration according to various tetracycline medications in the working curve derived sample.
Adopt this method and marine industry standard determination method " the mensuration High Performance Liquid Chromatography with Fluorescence Detection of terramycin, tetracycline, aureomycin, fortimicin residual quantity in the aquatic products " (DB33T 691-2008), measure simultaneously 10 parts of mark-on samples, result such as following table 1.The result shows: through t check, this method measurement result and add there was no significant difference between scalar, the standard method measurement result (P＜0.05) is so this method can be used for content 1 * 10
-8～1 * 10
-3The residual detection of tetracycline medication in the aquatic products of mol/L scope.
Table 1 this method measurement result with add scalar, standard method measurement result relatively
Claims (7)
1. selective electrode of tetracycline drugs is characterized in that: this electrode comprises and includes quartz glass tube (1), internal-filling liquid (2), internal reference electrode (3), electrode stem (4) and the epoxy resin (5) that sucks the molecular engram integral column that electroactive material is arranged; The outer wall stage casing that includes the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged is bonded at electrode stem (4) lower end, and sealing, add in the electrode stem (4) internal-filling liquid (2) is arranged, internal reference electrode (3) electrode insertion bar (4) also is immersed in the internal-filling liquid (2), with the fixing internal reference electrode (3) and electrode stem (4) upper end sealed of epoxy resin (5); The electrode of preparation places 10
-6～10
-8Activate in the quadracycline class drug solution of mol/L;
Described internal-filling liquid (2) is: contain 10
-3～10
-6The quadracycline class medicine, 10 of mol/L
-1～10
-3The NaCl of mol/L and 10
-1～10
-3The CaCl of mol/L
2Mixed solution;
Described electroactive material is obtained by following methods: with 0.005～0.02mol/L quadracycline class drug solution and 0.005～0.02mol/L Salkowski's solution in molar ratio 3～4: 1 or 0.005～0.02mol/L quadracycline class drug solution with mix under 0.005～0.02mol/L sodium tetraphenylborate solution 1: 1 in molar ratio～1.5 stirs, leave standstill 10～12h, filter, after sediment concentration is the salt acid elution 5～8 times of 0.01～0.05mol/L, 45～60 ℃ of oven dry namely get dry electroactive material.
2. selective electrode of tetracycline drugs according to claim 1, it is characterized in that: described tetracycline medication comprises tetracycline (TC), terramycin (OTC), aureomycin (CTC) or fortimicin (DC).
3. selective electrode of tetracycline drugs according to claim 1, it is characterized in that: described internal reference electrode is Pt electrode or Ag/AgCl electrode.
4. selective electrode of tetracycline drugs according to claim 1 is characterized in that: described to include the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged be to obtain by the following method:
1. template molecule of tetracycline class medicine, function monomer methacrylic acid are dissolved in the mixed solution of methyl alcohol and acetone, ultrasonic dissolution adds pore-foaming agent, behind stirring 6～8h, add crosslinking chemical ethylene glycol dimethacrylate and initiating agent azoisobutyronitrile, continue to be stirred to dissolving; Wherein, when template molecule was 0.1mmol, the mixed liquor volume of methyl alcohol and acetone was 0.5～1.5mL, and the volume of the shared mixed solution of methyl alcohol is: 45～100%; The mol ratio of template molecule of tetracycline class medicine, function monomer, crosslinking chemical is 1: 2～10: 20～50; The cumulative volume of function monomer, crosslinking chemical and the volume ratio of pore-foaming agent are 1～2: 3～4; Pore-foaming agent is the mixed solution of toluene and lauryl alcohol, and wherein the shared volume of toluene is 10～20%; Initiator amount is 8～20% of function monomer quality;
2. with behind the logical nitrogen 5～10min of said mixture, inserting internal diameter is that 1～3mm, length are in the quartz glass tube of 2～5cm, under nitrogen protection, with the quartz glass channel closure, place polyase 13～8h under the uviol lamp, obtain including the quartz glass tube of tetracycline medication molecular engram integral column;
3. intercepted length is the above-mentioned quartz glass tube that includes the tetracycline medication molecular engram integral column that makes of 0.5～1cm, soxhlet extraction is removed template molecule of tetracycline class medicine, 45～60 ℃ of drying 12～36h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline medication; Wherein the used extractant of soxhlet extraction is pure methyl alcohol;
4. above-mentioned quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline medication is immersed and be dissolved with in the uniform mix of electroactive material, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged;
The described synthesis step that is dissolved with the uniform mix of electroactive material is: get dry electroactive material, PVC, tetrahydrofuran and plastifier and join in the test tube, after the sealing, blend together uniform mix; Wherein, the mass ratio of electroactive material, PVC and plastifier is 0.5～10: 30～35: 30～70; When the quality of PVC was 1mg, the volume of tetrahydrofuran was 30～80 μ L; Plastifier is a kind of of following plastifier: dibutyl phthalate (DBP), dioctyl phthalate (DOP), dinonyl phthalate (DNP), Plexol 201 (DOS) or O-Nitrophenylfluorone Octyl Ether (o-NPOE).
5. the preparation method of a selective electrode of tetracycline drugs, it is characterized in that: the outer wall stage casing that will include the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged is bonded at electrode stem (4) lower end, and sealing abutting edge, add internal-filling liquid (2) in the electrode stem (4), with internal reference electrode (3) electrode insertion bar (4) and be immersed in the internal-filling liquid (2), with the fixing internal reference electrode (3) and with the upper end closure of electrode stem (4) of epoxy resin (5), freshly prepd electrode being placed concentration at last is 10
-6～10
-8Activate 48～96h in the quadracycline class drug solution of mol/L, thereby obtain the selective electrode of tetracycline drugs based on molecular engram integral column;
Described electroactive material is synthetic by following methods:
With 0.005～0.02mol/L quadracycline class drug solution and 0.005～0.02mol/L Salkowski's solution in molar ratio 3～4: 1 or 0.005～0.02mol/L quadracycline class drug solution with mix under 0.005～0.02mol/L sodium tetraphenylborate solution 1: 1 in molar ratio～1.5 stirs, leave standstill 10～12h, filter, after sediment concentration is the salt acid elution 5～8 times of 0.01～0.05mol/L, 45～60 ℃ of oven dry namely get dry electric active matter;
Described internal-filling liquid (2) is: contain 10
-3～10
-6The quadracycline class medicine, 10 of mol/L
-1～10
-3The NaCl of mol/L and 10
-1～10
-3The CaCl of mol/L
2Mixed solution.
6. the preparation method of selective electrode of tetracycline drugs according to claim 5 is characterized in that: the employed preparation method who includes the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged, and step is as follows:
1. template molecule of tetracycline class medicine, function monomer methacrylic acid are dissolved in the mixed solution of methyl alcohol and acetone, ultrasonic dissolution adds pore-foaming agent, behind stirring 6～8h, add crosslinking chemical ethylene glycol dimethacrylate and initiating agent azoisobutyronitrile, continue to be stirred to dissolving; Wherein, when template molecule was 0.1mmol, the mixed liquor volume of methyl alcohol and acetone was 0.5～1.5mL, and the volume of the shared mixed solution of methyl alcohol is: 45～100%; The mol ratio of template molecule of tetracycline class medicine, function monomer, crosslinking chemical is 1: 2～10: 20～50; The cumulative volume of function monomer, crosslinking chemical and the volume ratio of pore-foaming agent are 1～2: 3～4; Pore-foaming agent is the mixed solution of toluene and lauryl alcohol, and wherein the shared volume of toluene is 10～20%; Initiator amount is 8～20% of function monomer quality;
2. with behind the logical nitrogen 5～10min of said mixture, inserting internal diameter is that 1～3mm, length are in the quartz glass tube of 2～5cm, under nitrogen protection, with the quartz glass channel closure, place polyase 13～8h under the uviol lamp, obtain including the quartz glass tube of tetracycline medication molecular engram integral column;
3. intercepted length is the above-mentioned quartz glass tube that includes the tetracycline medication molecular engram integral column that makes of 0.5～1cm, soxhlet extraction is removed template molecule of tetracycline class medicine, 45～60 ℃ of drying 12～36h namely get the quartz glass tube that includes the molecular engram integral column behind the wash-out tetracycline medication; Wherein the used extractant of soxhlet extraction is pure methyl alcohol;
4. above-mentioned quartz glass tube one end that includes the molecular engram integral column behind the wash-out tetracycline medication is immersed and be dissolved with in the uniform mix of electroactive material, suck fast this uniform mix, after the tetrahydrofuran volatilization, namely form and include the quartz glass tube (1) that sucks the molecular engram integral column that electroactive material is arranged.
7. the preparation method of selective electrode of tetracycline drugs according to claim 6 is characterized in that: the described preparation method who is dissolved with the uniform mix of electroactive material, and step is followed successively by:
Get dry electroactive material, PVC, tetrahydrofuran and plastifier and join in the test tube, after the sealing, blend together uniform mix; Wherein, the mass ratio of electroactive material, PVC and plastifier is 0.5～10: 30～35: 30～70; When the quality of PVC was 1mg, the volume of tetrahydrofuran was 30～80 μ L; Plastifier is a kind of of following plastifier: dibutyl phthalate (DBP), dioctyl phthalate (DOP), dinonyl phthalate (DNP), Plexol 201 (DOS) or O-Nitrophenylfluorone Octyl Ether (o-NPOE).
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| CN101246150A (en) * | 2008-03-28 | 2008-08-20 | 南开大学 | Tsiklomitsin molecular engram integral column preparation method |
| CN101324541A (en) * | 2008-07-18 | 2008-12-17 | 大连理工大学 | Abricycline molecular imprinting polymer membrane electrode and preparation and application thereof |
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| CN101246150A (en) * | 2008-03-28 | 2008-08-20 | 南开大学 | Tsiklomitsin molecular engram integral column preparation method |
| CN101324541A (en) * | 2008-07-18 | 2008-12-17 | 大连理工大学 | Abricycline molecular imprinting polymer membrane electrode and preparation and application thereof |
Non-Patent Citations (4)
| Title |
|---|
| Ligang Chen等.Preparation of magnetic molecularly imprinted polymer for the separation of tetracycline antibiotics from egg and tissue samples.《Journal of Chromatography A》.2009,(第1216期),第3710-3719页. |
| Preparation of magnetic molecularly imprinted polymer for the separation of tetracycline antibiotics from egg and tissue samples;Ligang Chen等;《Journal of Chromatography A》;20090221(第1216期);第3710-3719页 * |
| 四环素选择性电极的研制和应用;安镜如等;《海峡药学》;19941231;第6卷(第3期);第23-25页 * |
| 安镜如等.四环素选择性电极的研制和应用.《海峡药学》.1994,第6卷(第3期),第23-25页. |
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