CN109060780A - Broad-spectrum specific molecularly imprinted polymer of sulfonamide, chemiluminescence kit, detection method and application - Google Patents
Broad-spectrum specific molecularly imprinted polymer of sulfonamide, chemiluminescence kit, detection method and application Download PDFInfo
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- CN109060780A CN109060780A CN201810911434.5A CN201810911434A CN109060780A CN 109060780 A CN109060780 A CN 109060780A CN 201810911434 A CN201810911434 A CN 201810911434A CN 109060780 A CN109060780 A CN 109060780A
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- Prior art keywords
- imprinted polymer
- molecularly imprinted
- sulfa drugs
- aperture
- titer plates
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- 229920000344 molecularly imprinted polymer Polymers 0.000 title claims abstract description 31
- 229940124530 sulfonamide Drugs 0.000 title claims abstract description 20
- 238000001514 detection method Methods 0.000 title claims abstract description 18
- 150000003456 sulfonamides Chemical class 0.000 title abstract 3
- 239000003814 drug Substances 0.000 claims abstract description 61
- 229940079593 drug Drugs 0.000 claims abstract description 59
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 23
- FDDDEECHVMSUSB-UHFFFAOYSA-N sulfanilamide Chemical compound NC1=CC=C(S(N)(=O)=O)C=C1 FDDDEECHVMSUSB-UHFFFAOYSA-N 0.000 claims abstract description 17
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims abstract description 14
- GEVPIWPYWJZSPR-UHFFFAOYSA-N tcpo Chemical compound ClC1=CC(Cl)=CC(Cl)=C1OC(=O)C(=O)OC1=C(Cl)C=C(Cl)C=C1Cl GEVPIWPYWJZSPR-UHFFFAOYSA-N 0.000 claims abstract description 7
- DBCAQXHNJOFNGC-UHFFFAOYSA-N 4-bromo-1,1,1-trifluorobutane Chemical compound FC(F)(F)CCCBr DBCAQXHNJOFNGC-UHFFFAOYSA-N 0.000 claims abstract description 6
- 239000003431 cross linking reagent Substances 0.000 claims abstract description 6
- STVZJERGLQHEKB-UHFFFAOYSA-N ethylene glycol dimethacrylate Substances CC(=C)C(=O)OCCOC(=O)C(C)=C STVZJERGLQHEKB-UHFFFAOYSA-N 0.000 claims abstract description 6
- 239000003999 initiator Substances 0.000 claims abstract description 6
- 239000000178 monomer Substances 0.000 claims abstract description 6
- 239000003054 catalyst Substances 0.000 claims abstract description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 24
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 24
- 239000000126 substance Substances 0.000 claims description 20
- 239000000243 solution Substances 0.000 claims description 17
- 239000004793 Polystyrene Substances 0.000 claims description 16
- 238000004020 luminiscence type Methods 0.000 claims description 16
- 229920002223 polystyrene Polymers 0.000 claims description 16
- 239000007787 solid Substances 0.000 claims description 15
- 239000013618 particulate matter Substances 0.000 claims description 13
- 238000000034 method Methods 0.000 claims description 12
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 10
- ZZORFUFYDOWNEF-UHFFFAOYSA-N sulfadimethoxine Chemical compound COC1=NC(OC)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 ZZORFUFYDOWNEF-UHFFFAOYSA-N 0.000 claims description 10
- 229960000973 sulfadimethoxine Drugs 0.000 claims description 10
- 238000012360 testing method Methods 0.000 claims description 9
- 239000002904 solvent Substances 0.000 claims description 7
- 229960002135 sulfadimidine Drugs 0.000 claims description 7
- ASWVTGNCAZCNNR-UHFFFAOYSA-N sulfamethazine Chemical compound CC1=CC(C)=NC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 ASWVTGNCAZCNNR-UHFFFAOYSA-N 0.000 claims description 7
- 238000006243 chemical reaction Methods 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- 150000002460 imidazoles Chemical class 0.000 claims description 6
- 238000001035 drying Methods 0.000 claims description 5
- 125000005395 methacrylic acid group Chemical class 0.000 claims description 5
- 229960004730 sulfabenzamide Drugs 0.000 claims description 5
- PBCZLFBEBARBBI-UHFFFAOYSA-N sulfabenzamide Chemical compound C1=CC(N)=CC=C1S(=O)(=O)NC(=O)C1=CC=CC=C1 PBCZLFBEBARBBI-UHFFFAOYSA-N 0.000 claims description 5
- 229960004306 sulfadiazine Drugs 0.000 claims description 5
- SEEPANYCNGTZFQ-UHFFFAOYSA-N sulfadiazine Chemical compound C1=CC(N)=CC=C1S(=O)(=O)NC1=NC=CC=N1 SEEPANYCNGTZFQ-UHFFFAOYSA-N 0.000 claims description 5
- GPTONYMQFTZPKC-UHFFFAOYSA-N sulfamethoxydiazine Chemical compound N1=CC(OC)=CN=C1NS(=O)(=O)C1=CC=C(N)C=C1 GPTONYMQFTZPKC-UHFFFAOYSA-N 0.000 claims description 5
- 229960002229 sulfametoxydiazine Drugs 0.000 claims description 5
- 229950003874 sulfamonomethoxine Drugs 0.000 claims description 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 4
- 229960005404 sulfamethoxazole Drugs 0.000 claims description 4
- VLYWMPOKSSWJAL-UHFFFAOYSA-N sulfamethoxypyridazine Chemical compound N1=NC(OC)=CC=C1NS(=O)(=O)C1=CC=C(N)C=C1 VLYWMPOKSSWJAL-UHFFFAOYSA-N 0.000 claims description 4
- 229960004936 sulfamethoxypyridazine Drugs 0.000 claims description 4
- JLKIGFTWXXRPMT-UHFFFAOYSA-N sulphamethoxazole Chemical compound O1C(C)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 JLKIGFTWXXRPMT-UHFFFAOYSA-N 0.000 claims description 4
- 239000004372 Polyvinyl alcohol Substances 0.000 claims description 3
- 239000012535 impurity Substances 0.000 claims description 3
- 239000011259 mixed solution Substances 0.000 claims description 3
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 3
- 235000019422 polyvinyl alcohol Nutrition 0.000 claims description 3
- 229940068984 polyvinyl alcohol Drugs 0.000 claims description 3
- KVNRLNFWIYMESJ-UHFFFAOYSA-N butyronitrile Chemical compound CCCC#N KVNRLNFWIYMESJ-UHFFFAOYSA-N 0.000 claims description 2
- 235000019441 ethanol Nutrition 0.000 claims description 2
- 239000000284 extract Substances 0.000 claims description 2
- 239000002994 raw material Substances 0.000 claims description 2
- SMWDFEZZVXVKRB-UHFFFAOYSA-N Quinoline Chemical compound N1=CC=CC2=CC=CC=C21 SMWDFEZZVXVKRB-UHFFFAOYSA-N 0.000 claims 4
- RZVAJINKPMORJF-UHFFFAOYSA-N Acetaminophen Chemical compound CC(=O)NC1=CC=C(O)C=C1 RZVAJINKPMORJF-UHFFFAOYSA-N 0.000 claims 1
- 150000003851 azoles Chemical class 0.000 claims 1
- OZAIFHULBGXAKX-UHFFFAOYSA-N 2-(2-cyanopropan-2-yldiazenyl)-2-methylpropanenitrile Chemical compound N#CC(C)(C)N=NC(C)(C)C#N OZAIFHULBGXAKX-UHFFFAOYSA-N 0.000 abstract description 4
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 abstract 3
- RAXXELZNTBOGNW-UHFFFAOYSA-N imidazole Natural products C1=CNC=N1 RAXXELZNTBOGNW-UHFFFAOYSA-N 0.000 abstract 3
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 abstract 1
- 150000001875 compounds Chemical class 0.000 abstract 1
- 229920000642 polymer Polymers 0.000 description 7
- 241001465754 Metazoa Species 0.000 description 6
- 238000003786 synthesis reaction Methods 0.000 description 6
- WMPXPUYPYQKQCX-UHFFFAOYSA-N Sulfamonomethoxine Chemical compound C1=NC(OC)=CC(NS(=O)(=O)C=2C=CC(N)=CC=2)=N1 WMPXPUYPYQKQCX-UHFFFAOYSA-N 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 238000003018 immunoassay Methods 0.000 description 4
- XSCHRSMBECNVNS-UHFFFAOYSA-N quinoxaline Chemical compound N1=CC=NC2=CC=CC=C21 XSCHRSMBECNVNS-UHFFFAOYSA-N 0.000 description 4
- 230000000844 anti-bacterial effect Effects 0.000 description 3
- -1 bacteresulf Chemical compound 0.000 description 3
- 239000002245 particle Substances 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 238000012216 screening Methods 0.000 description 3
- NHZLNPMOSADWGC-UHFFFAOYSA-N 4-amino-N-(2-quinoxalinyl)benzenesulfonamide Chemical compound C1=CC(N)=CC=C1S(=O)(=O)NC1=CN=C(C=CC=C2)C2=N1 NHZLNPMOSADWGC-UHFFFAOYSA-N 0.000 description 2
- 231100000703 Maximum Residue Limit Toxicity 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003640 drug residue Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- CTAPFRYPJLPFDF-UHFFFAOYSA-N isoxazole Chemical compound C=1C=NOC=1 CTAPFRYPJLPFDF-UHFFFAOYSA-N 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 238000001179 sorption measurement Methods 0.000 description 2
- 229960003097 sulfaquinoxaline Drugs 0.000 description 2
- ASFHDLDAWYTMJS-UHFFFAOYSA-N 3-methoxypyridazine Chemical compound COC1=CC=CN=N1 ASFHDLDAWYTMJS-UHFFFAOYSA-N 0.000 description 1
- 206010067484 Adverse reaction Diseases 0.000 description 1
- 208000019838 Blood disease Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 206010020751 Hypersensitivity Diseases 0.000 description 1
- ZCQWOFVYLHDMMC-UHFFFAOYSA-N Oxazole Chemical compound C1=COC=N1 ZCQWOFVYLHDMMC-UHFFFAOYSA-N 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 239000005864 Sulphur Substances 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- 230000006838 adverse reaction Effects 0.000 description 1
- 208000026935 allergic disease Diseases 0.000 description 1
- 230000007815 allergy Effects 0.000 description 1
- DKNWSYNQZKUICI-UHFFFAOYSA-N amantadine Chemical compound C1C(C2)CC3CC2CC1(N)C3 DKNWSYNQZKUICI-UHFFFAOYSA-N 0.000 description 1
- 229960003805 amantadine Drugs 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 238000003975 animal breeding Methods 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 229960005091 chloramphenicol Drugs 0.000 description 1
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000003480 eluent Substances 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 238000004401 flow injection analysis Methods 0.000 description 1
- 208000014951 hematologic disease Diseases 0.000 description 1
- 208000018706 hematopoietic system disease Diseases 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 238000010324 immunological assay Methods 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000002414 normal-phase solid-phase extraction Methods 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- LISFMEBWQUVKPJ-UHFFFAOYSA-N quinolin-2-ol Chemical compound C1=CC=C2NC(=O)C=CC2=C1 LISFMEBWQUVKPJ-UHFFFAOYSA-N 0.000 description 1
- 238000011896 sensitive detection Methods 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 239000000021 stimulant Substances 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- 238000004704 ultra performance liquid chromatography Methods 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/76—Chemiluminescence; Bioluminescence
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
Landscapes
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Plasma & Fusion (AREA)
- Engineering & Computer Science (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention discloses a broad-spectrum specific molecularly imprinted polymer of sulfonamide drugs, which is prepared by using sulfanilamide benzene as a pseudo molecular template, methacrylic acid as a functional monomer, azodiisobutyronitrile as an initiator and ethylene glycol dimethacrylate as a cross-linking agent. The molecular imprinting compound is used as a recognition reagent, imidazole is used as a catalyst, and bis (2,4, 6-trichlorophenyl) oxalate and hydrogen peroxide are used as chemiluminescent reagents to prepare a chemiluminescent kit, and a detection method using the kit is also provided. The molecularly imprinted polymer can capture 12 drugs, and the chemiluminescence kit can accurately, quickly and efficiently detect 12 sulfonamides.
Description
Technical field
The present invention relates to animal food safety fields, and in particular to a kind of molecule print that can capture 12 kinds of sulfa drugs
The detection method and application of mark polymer, the chemical luminescence reagent kit thus prepared and the kit.
Background technique
Sulfa drugs is a kind of chemical synthesis antibacterials, and such medicine antibacterial spectrum is wide, and antibacterial activity is strong, uses extensively
It is treated in the bacteriosis of animals and humans.Common kind has sulphadiazine, sulfadimidine, bacteresulf, sulfanilamide (SN)
First oxazole, sulfametoxydiazine, sulfadimethoxine, daimeton, sulfadimethoxine, sulfaquinoxaline, sulfanilamide (SN)
Methoxy pyridazine, cistosulfa, sulfabenzamide etc..With a large amount of uses of sulfa drugs during food animal-breeding,
They in animal derived food caused by residue problem attract extensive attention.The residual of sulfa drugs is likely to result in consumption
Long-term a small amount of intake of person, causes the adverse reactions such as blood disease, allergy, in some instances it may even be possible to induce the drug resistance of humanized pathogenic bacteria.
Therefore, it is necessary to pay attention to residue problem of such drug in animal derived food.USA and EU all defines such drug
Maximum residue limit, the Ministry of Agriculture have also issued the maximum residue limit of sulfa drugs.
Currently, there is many methods that can detect to the residual of sulfa drugs, wherein immunoassay is simple, square
Just, the screening that can be used for batch samples, it is widely used in testing agency of base.But core reagent-antibody of immunoassay
The immunoassay or commercialization immunologic function test reagent for needing 4-6 months production cycles, and being established using antibody as recognition component
Box is disposable.Therefore, detection method/product gesture having the advantages that immunoassay, can be recycled is developed to exist
It must go.
Molecularly imprinted polymer is a kind of chemical synthesis material, can be formed have to specific objective object in the synthesis process
There is the three-dimensional lumen of specific recognition capability, therefore, referred to as " plastics antibody ".Moreover, molecularly imprinted polymer preparation the phase it is short,
It is at low cost, repeatable to be recycled.Chemoluminescence method is a kind of easy to operate, the detection method of analysis speed fast (10-30 seconds), spirit
Sensitivity is higher than conventional immunological assays, and agents useful for same is chemical reagent, is not influenced by factors such as temperature, times.Currently,
There are a small number of documents to use the drug original shape of sulfa drugs for molecular template both at home and abroad, synthesizes corresponding molecularly imprinted polymer and make
It is recognition component for establishing chemiluminescence detecting method.But these methods are all made of double pump flow injection mode, and operation sequence is multiple
It is miscellaneous, it not can be carried out batch screening, i.e., multiple samples can only singly be detected.Secondly, reported method can only
Detect a certain sulfa drugs.In addition, the sensitivity of these methods can not detect the sulfa drug residue of trace.
Summary of the invention
It is an object of the present invention to provide a kind of molecularly imprinted polymer of the Broadspectrum specificity of sulfa drugs, this points
Sub- imprinted polymer can capture a variety of sulfa drugs, and can be directly used for the detection of chemiluminometry.
To achieve the goals above, the present invention adopts the following technical scheme that:
A kind of molecularly imprinted polymer of the Broadspectrum specificity of sulfa drugs, it is characterised in that prepare by the following method, below
Raw material presses the meter of substance:
(a) 1 part of false molecular template N'-phenylsulfanilamide, 5 ~ 7 parts of function monomer methacrylic acids, 20 ~ 30 parts of initiator azo two is different
Butyronitrile and 20 ~ 30 parts of crosslinking agent ethylene glycol dimethacrylate are placed in solvent, and 60 ~ 70 DEG C are reacted 10 ~ 12 hours, are obtained
To solid particulate matter；
(b) solid particulate matter is placed in Soxhlet extractor, with extracting solution continuous backflow 12 ~ 24 hours, by the false mould
Plate molecule N'-phenylsulfanilamide extracts；
(c) by after the solid particulate matter drying after step (b) reaction, the molecular engram for obtaining the Broadspectrum specificity of sulfa drugs is poly-
Close object.
Preferably, solvent used in the step (a) is chloroform, extracting solution used in step (b) be methanol with
The mixed solution of acetic acid, and the volume ratio of methanol and acetic acid is 9:0.5 ~ 2.
It is a further object to provide a kind of chemical illuminating reagents of the Broadspectrum specificity of sulfa drugs detection
Box, the kit can accurately detect a variety of sulfa drugs, and it is convenient to operate, as a result reliably.For this purpose, the present invention takes
Following technical solution:
A kind of chemical luminescence reagent kit of the Broadspectrum specificity detection of sulfa drugs, the chemical luminescence reagent kit is with previous molecular
Imprinted polymer is identification agent, is to change with bis- (2,4,6- trichlorophenyl) oxalates and hydrogen peroxide using imidazoles as catalyst
Luminescence reagent is learned, according to luminous intensity and testing concentration there is proportionate relationship to make detection to testing concentration.
The present invention also provides the detection methods using above-mentioned chemical luminescence reagent kit to sulfa drugs, including walk as follows
It is rapid:
(a) above-mentioned molecularly imprinted polymer is suspended in weight percent is then to be added in 0.3 ~ 2% poly-vinyl alcohol solution
Into the aperture of polystyrene titer plates, place 1 ~ 2 hour；
(b) sulfa drugs to be detected or sample extracting solution are added in the aperture of polystyrene titer plates, are placed at room temperature for 5
~ 50 minutes；
(c) aperture of the polystyrene titer plates is washed, removes impurity and without captured sulfa drugs；
(d) will be bis- (2,4,6- trichlorophenyl) oxalate, imidazoles and hydrogen peroxide be added to the small of the polystyrene titer plates
The polystyrene titer plates are placed in Chemiluminescence Apparatus or multi-function microplate reader by Kong Zhong, read the values of chemiluminescence of each aperture.
Preferably, it in the step (c), is washed with aperture of the ethyl alcohol to the polystyrene titer plates.
The present invention provides a kind of above-mentioned chemical luminescence reagent kit and detects sulphadiazine, sulfadimidine, sulfanilamide (SN) at the same time
Isoxazole, sulfamethoxazole, sulfametoxydiazine, sulfadimethoxine, daimeton, sulfadimethoxine, sulfanilamide (SN)
The application of quinoxaline, sulfamethoxypyridazine, cistosulfa and sulfabenzamide field.
The beneficial effects of the present invention are: being false molecular template with N'-phenylsulfanilamide, the Broadspectrum specificity of sulfa drugs has been synthesized
Molecularly imprinted polymer can identify 12 common kinds of such drug: sulphadiazine, sulfadimidine, sulfanilamide (SN) simultaneously
Isoxazole, sulfamethoxazole, sulfametoxydiazine, sulfadimethoxine, daimeton, sulfadimethoxine, sulfanilamide (SN)
Quinoxaline, sulfamethoxypyridazine, cistosulfa, sulfabenzamide.The change prepared using the molecularly imprinted polymer as recognition component
Luminescence reagent box is learned, 12 kinds of sulfa drugs in animal muscle can be carried out with more residuals, quick, Sensitive Detection, to realize
The field quick detection of sulfa drug residue is laid a good foundation, and can ensure animal food safety in a wider context.In addition,
Since the tolerance that molecularly imprinted polymer changes chemical/physical condition is strong, chemical illuminating reagent provided by the invention
Box may be reused, and greatly reduce testing cost, reduce resource waste, and meet current social Green Sustainable
Trend.
Detailed description of the invention
Fig. 1 be control without molecular template when the electron-microscope scanning picture of polymer that synthesizes；
Fig. 2 is the electron-microscope scanning picture of the molecularly imprinted polymer of this patent preparation；
Fig. 3 is capture effect of the molecularly imprinted polymer to 9 kinds of sulfa drugs of this patent preparation；
Fig. 4 is the concentration of sulfadimidine detection and the standard curve of OD value.
Specific embodiment
Below by specific embodiment, the present invention will be further described, presses the meter of substance below:
1 molecularly imprinted polymer synthesis example 1 of embodiment
(a) by 1 part of false molecular template N'-phenylsulfanilamide, 5 parts of function monomer methacrylic acids, 20 parts of initiator azodiisobutyronitrile and
20 parts of crosslinking agent ethylene glycol dimethacrylate is placed in chloroform solvent, and 60 DEG C are reacted 12 hours, obtains solid particle
Object；
(b) solid particulate matter is placed in Soxhlet extractor, with methanol and acetic acid (9:1, V/V) extracting solution continuous backflow 12
Hour, the false template molecule N'-phenylsulfanilamide of trace is extracted；
(c) by after the solid particulate matter drying after step (b) reaction, the molecular engram for obtaining the Broadspectrum specificity of sulfa drugs is poly-
Close object.
2 molecularly imprinted polymer synthesis example 2 of embodiment
(a) by 1 part of false molecular template N'-phenylsulfanilamide, 7 parts of function monomer methacrylic acids, 30 parts of initiator azodiisobutyronitrile and
30 parts of crosslinking agent ethylene glycol dimethacrylate is placed in chloroform solvent, and 70 DEG C are reacted 10 hours, obtains solid particle
Object；
(b) solid particulate matter is placed in Soxhlet extractor, with methanol and acetic acid (9:2, V/V) extracting solution continuous backflow 24
Hour, the false template molecule N'-phenylsulfanilamide of trace is extracted；
(c) by after the solid particulate matter drying after step (b) reaction, the molecular engram for obtaining the Broadspectrum specificity of sulfa drugs is poly-
Close object.
3 molecularly imprinted polymer synthesis example 3 of embodiment
(a) by 1 part of false molecular template N'-phenylsulfanilamide, 6 parts of function monomer methacrylic acids, 25 parts of initiator azodiisobutyronitrile and
25 parts of crosslinking agent ethylene glycol dimethacrylate is placed in chloroform solvent, and 65 DEG C are reacted 11 hours, obtains solid particulate matter；
(b) solid particulate matter is placed in Soxhlet extractor, with methanol and acetic acid (9:0.5, V/V) extracting solution continuous backflow
18 hours, the false template molecule N'-phenylsulfanilamide of trace is extracted；
(c) by after the solid particulate matter drying after step (b) reaction, the molecular engram for obtaining the Broadspectrum specificity of sulfa drugs is poly-
Close object.
The electron microscope photo scanning of molecularly imprinted polymer prepared by this example is as shown in Fig. 2, its surface is more as seen from Figure 2
Hole can identify sulfa drugs for the cavity that template molecule trace is formed；It is distinguished with this, if do not used
False molecular template is stated, the electron microscope photo scanning of the polymer synthesized is as shown in Figure 1, its surfacing, smooth, without hole,
Sulfa drugs cannot be identified.The electron microscope photo scanning of molecularly imprinted polymer prepared by embodiment 1 and embodiment 2
It is similar with Fig. 2 situation in this example.
Capture validity check of the above-mentioned molecularly imprinted polymer of embodiment 4 to nine kinds of sulfa drugs
The molecularly imprinted polymer particle that sulfa drugs is directed to prepared by embodiment 3 is placed in empty solid phase extraction as filler
It takes in column, by 9 kinds of drugs (1=sulphadiazine, 2=sulfadimethoxine, 3=bacteresulf, 4=sulfamethoxazole, 5=sulphur
Sulfamonomethoxine, 6=sulfadimidine, 7=sulfadimethoxine, 8=sulfabenzamide, 9=sulfaquinoxaline between amine) mark
Quasi- product mixed solution is added on the column, and liquid flows out naturally.Then the medicine that will be adsorbed with methanol/acetic acid solution (8:2, V/V)
Object elution, eluent are detected with ultra-performance liquid chromatography.Chromatogram shows that the polymer has the above-mentioned 9 kinds of drugs of specific adsorption
Ability.Similar experiment shows the Molecularly Imprinted Polymer to 3 kinds of sulfametoxydiazine, sulfamethoxypyridazine, cistosulfa medicines
Object also has specific adsorption ability.It attempts non-with amantadine, quinolone, beta-stimulants, tetracycline, chloramphenicol, phenthazine etc.
Sulfa drugs carries out similar experiment, it is found that this molecular imprinting polymer does not adsorb these drugs.Therefore, by the polymer
It is coated in the aperture of microwell plate as recognition component, the kit being assembled into can adsorb above-mentioned 12 kinds of sulfa drugs.
By luminesceence analysis, can quickly screen in sample to be tested whether there are also at least one of above-mentioned 12 kinds of sulfa drugs.
The composition of 5 chemical luminescence reagent kit of embodiment
Chemical luminescence reagent kit includes following composition:
Molecularly imprinted polymer prepared by embodiment 1,2 or 3 can capture sulfa drugs as identification agent, for inspection
It surveys and basis is provided；
Bis- (2,4,6- trichlorophenyl) oxalates and hydrogen peroxide, both as chemical illuminating reagent；
Imidazoles plays catalytic action to reaction, it is made quickly to shine as catalyst；
After shining, according to luminous intensity and testing concentration there is proportional relationship to do to testing concentration using Chemiluminescence Apparatus
It detects out.
Embodiment 6 utilizes the detection method of mentioned reagent box sulfa drugs
Detection method is as follows:
(a) previous molecular imprinted polymer is suspended in weight percent is to be then added in 0.3 ~ 2% poly-vinyl alcohol solution
In the aperture of polystyrene titer plates, place 1 ~ 2 hour；
(b) sulfa drugs to be detected or sample extracting solution are added in the aperture of polystyrene titer plates, are placed at room temperature for 5
~ 50 minutes；
(c) aperture of the polystyrene titer plates is washed, removes impurity and without captured sulfa drugs；
(d) will be bis- (2,4,6- trichlorophenyl) oxalate, imidazoles and hydrogen peroxide be added sequentially to the polystyrene titer plates
Aperture in, which is placed in Chemiluminescence Apparatus or multi-function microplate reader, read each aperture chemistry hair
Light value.
(e) according to whether there is luminous value, judge whether at least one containing 12 kinds of sulfa drugs used in embodiment 4
Kind.
After above-mentioned molecularly imprinted polymer-chemical luminescence reagent kit is washed with methanol/acetic acid, reusable 5 times.
This example illustrates that this kit disposably can quickly screen in sample to be tested whether contain above-mentioned sulfa drugs
It is one or more, above-mentioned 12 kinds of sulfa drugs are free of for the sample explanation of not luminescence phenomenon, for there is luminescence phenomenon
Sample can be analyzed again by specific type and content of the high performance liquid chromatography to contained sulfa drugs.Therefore, originally
Kit greatly improves the screening efficiency to sample.
The detection effect of 7 chemical luminescence reagent kit of embodiment
By taking sulfadimidine as an example, by drug standards with sample extracting solution carry out series of concentrations dilution (0.01,0.05,
0.1,0.5,1.0,5.0,10,50 ng/mL), it is then respectively added to be surrounded by the different apertures of molecularly imprinted polymer, press
The above method is tested.The result shows that being in good linear relationship between drug concentration and values of chemiluminescence, illustrate to use the examination
When agent box, the concentration of drug can be calculated by the OD value of measurement, accuracy with higher, experimental result is such as
Shown in Fig. 4.
Above-described embodiment is the explanation conceived to the utility model and realized, is not intended to limit it, in present inventive concept
Under, without the technical solution that substantially converts still in protection scope.
Claims (6)
1. a kind of molecularly imprinted polymer of the Broadspectrum specificity of sulfa drugs, it is characterised in that it prepares by the following method, with
Lower raw material presses the meter of substance:
(a) 1 part of false molecular template N'-phenylsulfanilamide, 5 ~ 7 parts of function monomer methacrylic acids, 20 ~ 30 parts of initiator azo two is different
Butyronitrile and 20 ~ 30 parts of crosslinking agent ethylene glycol dimethacrylate are placed in solvent, and 60 ~ 70 DEG C are reacted 10 ~ 12 hours, are obtained
To solid particulate matter；
(b) solid particulate matter is placed in Soxhlet extractor, with extracting solution continuous backflow 12 ~ 24 hours, by the false mould
Plate molecule N'-phenylsulfanilamide extracts；
(c) by after the solid particulate matter drying after step (b) reaction, the molecular engram for obtaining the Broadspectrum specificity of sulfa drugs is poly-
Close object.
2. molecularly imprinted polymer as described in claim 1, it is characterised in that: solvent used in the step (a) is
Chloroform, extracting solution used in step (b) are the mixed solution of methanol and acetic acid.
3. the chemical luminescence reagent kit that a kind of Broadspectrum specificity of sulfa drugs detects, it is characterised in that the chemiluminescence examination
Agent box is using molecularly imprinted polymer of any of claims 1 or 2 as identification agent, using imidazoles as catalyst, with bis- (2,4,6- tri-
Chlorphenyl) oxalate and hydrogen peroxide be chemical illuminating reagent, proportionate relationship pair is had according to luminous intensity and testing concentration
Testing concentration makes detection.
4. using chemical luminescence reagent kit as claimed in claim 3 to the detection method of sulfa drugs, it is characterised in that including
Following steps:
(a) molecularly imprinted polymer of any of claims 1 or 2 is suspended in weight percent is 0.3 ~ 2% poly-vinyl alcohol solution
In, it is then added in the aperture of polystyrene titer plates, places 1 ~ 2 hour；
(b) sulfa drugs to be detected or sample extracting solution are added in the aperture of polystyrene titer plates, are placed at room temperature for 5
~ 50 minutes；
(c) aperture of the polystyrene titer plates is washed, removes impurity and without captured sulfa drugs；
(d) will be bis- (2,4,6- trichlorophenyl) oxalate, imidazoles and hydrogen peroxide be added to the small of the polystyrene titer plates
The polystyrene titer plates are placed in Chemiluminescence Apparatus or multi-function microplate reader by Kong Zhong, read the values of chemiluminescence of each aperture.
5. method as claimed in claim 4, which is characterized in that in the step (c), titrated with ethyl alcohol to the polystyrene
The aperture of plate is washed.
6. chemical luminescence reagent kit as claimed in claim 3 detects the different evil of sulphadiazine, sulfadimidine, sulfanilamide (SN) at the same time
Azoles, sulfamethoxazole, sulfametoxydiazine, sulfadimethoxine, daimeton, sulfadimethoxine, sulfanilamide (SN) quinoline are disliked
The application of quinoline, sulfamethoxypyridazine, cistosulfa and sulfabenzamide field.
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| CN111151227A (en) * | 2019-10-21 | 2020-05-15 | 深圳市易瑞生物技术股份有限公司 | Semi-molecular imprinting material and preparation method and application thereof |
| CN111551622A (en) * | 2020-06-29 | 2020-08-18 | 盐城工学院 | Preparation method of high-sensitivity sulfadiazine molecular imprinting electrochemical sensor |
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