CN103524679B - The preparation method of the molecular imprinting polymkeric substance of anticoccidial drug diclazuril and application - Google Patents
The preparation method of the molecular imprinting polymkeric substance of anticoccidial drug diclazuril and application Download PDFInfo
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- ZSZFUDFOPOMEET-UHFFFAOYSA-N 2-(4-chlorophenyl)-2-[2,6-dichloro-4-(3,5-dioxo-1,2,4-triazin-2-yl)phenyl]acetonitrile Chemical compound C1=CC(Cl)=CC=C1C(C#N)C1=C(Cl)C=C(N2C(NC(=O)C=N2)=O)C=C1Cl ZSZFUDFOPOMEET-UHFFFAOYSA-N 0.000 title claims abstract description 61
- 229960000248 diclazuril Drugs 0.000 title claims abstract description 60
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- 239000003814 drug Substances 0.000 title claims abstract description 14
- 230000001165 anti-coccidial effect Effects 0.000 title claims abstract description 13
- 239000000126 substance Substances 0.000 title claims abstract description 13
- 229940079593 drug Drugs 0.000 title claims abstract description 12
- 239000000178 monomer Substances 0.000 claims abstract description 22
- 229920000642 polymer Polymers 0.000 claims abstract description 18
- 238000002414 normal-phase solid-phase extraction Methods 0.000 claims abstract description 17
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 claims abstract description 11
- STVZJERGLQHEKB-UHFFFAOYSA-N ethylene glycol dimethacrylate Substances CC(=C)C(=O)OCCOC(=O)C(C)=C STVZJERGLQHEKB-UHFFFAOYSA-N 0.000 claims abstract description 11
- DBCAQXHNJOFNGC-UHFFFAOYSA-N 4-bromo-1,1,1-trifluorobutane Chemical compound FC(F)(F)CCCBr DBCAQXHNJOFNGC-UHFFFAOYSA-N 0.000 claims abstract description 10
- 239000003795 chemical substances by application Substances 0.000 claims abstract description 7
- 238000001514 detection method Methods 0.000 claims abstract 3
- 238000002203 pretreatment Methods 0.000 claims abstract 2
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 57
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 26
- 229920000344 molecularly imprinted polymer Polymers 0.000 claims description 23
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 12
- 241000287828 Gallus gallus Species 0.000 claims description 8
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 claims description 8
- 238000005251 capillar electrophoresis Methods 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 6
- 239000002904 solvent Substances 0.000 claims description 6
- PBCJIPOGFJYBJE-UHFFFAOYSA-N acetonitrile;hydrate Chemical compound O.CC#N PBCJIPOGFJYBJE-UHFFFAOYSA-N 0.000 claims description 5
- 239000000203 mixture Substances 0.000 claims description 5
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 claims description 5
- 230000010355 oscillation Effects 0.000 claims description 5
- 238000004090 dissolution Methods 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- 239000011491 glass wool Substances 0.000 claims description 4
- 238000002390 rotary evaporation Methods 0.000 claims description 4
- 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 claims description 3
- OZAIFHULBGXAKX-VAWYXSNFSA-N AIBN Substances N#CC(C)(C)\N=N\C(C)(C)C#N OZAIFHULBGXAKX-VAWYXSNFSA-N 0.000 claims description 3
- 206010013786 Dry skin Diseases 0.000 claims description 3
- 239000008367 deionised water Substances 0.000 claims description 3
- 229910021641 deionized water Inorganic materials 0.000 claims description 3
- 238000001035 drying Methods 0.000 claims description 3
- 238000011068 loading method Methods 0.000 claims description 3
- VVWRJUBEIPHGQF-UHFFFAOYSA-N propan-2-yl n-propan-2-yloxycarbonyliminocarbamate Chemical compound CC(C)OC(=O)N=NC(=O)OC(C)C VVWRJUBEIPHGQF-UHFFFAOYSA-N 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 238000000605 extraction Methods 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 2
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 2
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 2
- 238000000034 method Methods 0.000 abstract description 8
- 241001465754 Metazoa Species 0.000 abstract description 5
- 235000013305 food Nutrition 0.000 abstract description 3
- 239000000945 filler Substances 0.000 abstract description 2
- 238000006116 polymerization reaction Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 16
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 12
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 8
- 230000027455 binding Effects 0.000 description 8
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 7
- 230000000694 effects Effects 0.000 description 7
- 235000013330 chicken meat Nutrition 0.000 description 6
- 239000000758 substrate Substances 0.000 description 5
- 239000007788 liquid Substances 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 4
- 238000010521 absorption reaction Methods 0.000 description 3
- 230000000274 adsorptive effect Effects 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 239000000523 sample Substances 0.000 description 3
- 238000001179 sorption measurement Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 150000001336 alkenes Chemical class 0.000 description 2
- 125000003118 aryl group Chemical group 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 239000003431 cross linking reagent Substances 0.000 description 2
- 235000010855 food raising agent Nutrition 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 238000001157 Fourier transform infrared spectrum Methods 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 238000005202 decontamination Methods 0.000 description 1
- 230000003588 decontaminative effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000002329 infrared spectrum Methods 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 239000004005 microsphere Substances 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 230000009871 nonspecific binding Effects 0.000 description 1
- 244000045947 parasite Species 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 239000002861 polymer material Substances 0.000 description 1
- 238000007639 printing Methods 0.000 description 1
- 230000000384 rearing effect Effects 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 150000003918 triazines Chemical class 0.000 description 1
- 239000000273 veterinary drug Substances 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
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Abstract
The present invention relates to the preparation method of the molecular imprinting polymkeric substance of anticoccidial drug diclazuril, the method with diclazuril (DIC) for template molecule, methacrylic acid (MAA) is function monomer, take ethylene glycol dimethacrylate as linking agent, being obtained by steps such as dissolving, polymerization, wash-outs take diclazuril as the imprinted polymer of template.This imprinted polymer is used for the filler of solid-phase extraction column, and obtained solid-phase extraction column has good identity to diclazuril medicine, can be used for the sample pre-treatments of anticoccidial drug diclazuril residue detection in animal derived food.
Description
Technical field
The present invention relates to the preparation method of molecularly imprinted polymer; The preparation method of the molecularly imprinted polymer of anticoccidial drug diclazuril in animal derived food specifically.
Background technology
Diclazuril belongs to triazines anticoccidial drug, and because having efficient, low toxicity, wide spectrum expelling parasite feature, its anticoccidial effect is better than other anticoccidial drugs and is used widely in China.As incorrect use anticoccidial drug diclazuril will cause its animal food safety problem in animal rearing.
Molecular imprinting (MolecularImprintingTechniqe, MIT) be the new technology that the preparation occurred in recent years has the polymer materials of recognition function, the molecularly imprinted polymer (molecularlyimprintedpolymers, MIPs) mated completely with certain a part on space structure and binding site can be obtained.This technology has been applied to the Solid-Phase Extraction Column preparation of many veterinary drugs, but has no report for the molecular imprinting Solid-Phase Extraction column preparation method of diclazuril.
Summary of the invention
The object of the present invention is to provide a kind of molecular imprinting Solid-Phase Extraction column preparation method of diclazuril, namely be the function monomer of molecularly imprinted polymer with diclazuril, obtained molecularly imprinted polymer, mix with different ratios raising agent (diatomite) after crushed and fill post, obtained solid-phase extraction column has good recognition performance to diclazuril.
Technical solution of the present invention is as follows:
The preparation method of the molecular imprinting polymkeric substance of anticoccidial drug diclazuril, this preparation method carries out as follows:
(1) take quality≤0.408g template molecule diclazuril (DIC) and be dissolved in 20mLN, in dinethylformamide solution, add function monomer methacrylic acid (MAA) 0.265-0.795mL, at sonic oscillation 30min under room temperature, template molecule is fully combined with function monomer; Then linking agent ethylene glycol dimethacrylate (EDMA) 1.29-6.43mL is added; Add Diisopropyl azodicarboxylate (AIBN) 0.05g again, mix, sonic oscillation 15min, at N
2protect in underlying 60 DEG C of water-baths and heat 24h, obtain solid white polymer;
(2) solid white polymer of gained is pulverized by step (1), ground 220 mesh sieves, with the template molecule diclazuril in the acetonitrile-water of volume ratio 8:2 or 7:3 or 6:4 or the methyl alcohol/acetic acid solution eluted polymer of volume ratio 8:2, use deionized water eluting solvent again, collect product 60 DEG C of dryings in baking oven, obtaining take diclazuril as the imprinted polymer of template.
As preferably, template molecule described in the inventive method: function monomer: the mol ratio of linking agent is 1:5:30.
Described template molecule quality is not more than finger and comprises but be not more than 0.408g.
A kind of solid phase extraction column stuffing, described filler is that the imprinted polymer of template and diatomite form by diclazuril according to claim 1, and described imprinted polymer and diatomaceous mass ratio are 2:1.
Compared with the prior art, beneficial effect of the present invention is embodied in:
The present invention take diclazuril as the template of molecularly imprinted polymer, and obtained molecularly imprinted polymer, mix with different ratios raising agent (diatomite) after crushed and fill post, obtained solid-phase extraction column has good recognition performance to diclazuril medicine.
Accompanying drawing explanation
Fig. 1 is the infrared spectrogram of NMIPs
Fig. 2 is the infrared spectrogram of MIPs
Fig. 3 is the scanning electron microscope of NMIPs
Fig. 4 is the scanning electron microscope sem figure of MIPs before wash-out
Fig. 5 is the scanning electron microscope sem figure of MIPs after wash-out
Fig. 6 is through the mark-on chicken meat sample electrophorogram of MISPE
Embodiment
Below by way of specific embodiment, technical solution of the present invention is described further.Testing sequence in embodiment and test product used, if special instruction is this area routine techniques means and conventional products.
Embodiment 1
(1) compare methyl alcohol, N, N-dimethylformamide, acetonitrile, tetrahydrofuran (THF) respectively as solvent, investigate the solute effect of different solvents to diclazuril (DIC) and MAA.Result shows, DIC and MAA is insoluble in methyl alcohol, and slightly soluble in acetonitrile, compares with tetrahydrofuran (THF), the most molten at N, N-dimethylformamide.Therefore, N, N-dimethylformamide is selected to be the solvent preparing diclazuril molecularly imprinted polymer.
(2) impact of function monomer MAA and the ratio of template molecule diclazuril consumption: test fixed die plate molecule consumption, change the amount of function monomer, the mol ratio investigating function monomer and template molecule is 1:1, 3:1, 5:1, 7:1, interaction situation during 9:1 between function monomer and template molecule, at N, template molecule is fully combined in N-DMF solution with function monomer after, N is measured in 220nm place with Shimadzu 2500 type ultraviolet-visible pectrophotometer, the absorbance remaining template molecule diclazuril in N-DMF solution is respectively 0.439, 0.418, 0.386, 0.381, 0.379.When increasing along with function monomer amount, absorbancy constantly declines, and pattern of descriptive parts molecule and function monomer interact and form host-guest complex.When the mol ratio of function monomer and template molecule is increased by 5:1 again, absorbancy change is no longer obvious, illustrate that now template molecule and function monomer have acted on fully, can self-association be there is in unnecessary function monomer, form nonspecific binding site, absorption resistance to mass transfer is increased, is unfavorable for preparing imprinted polymer, so the mol ratio 5:1 of function monomer and template molecule is optimal selection.
The impact of dosage of crosslinking agent:
Experiment synthesizing glycol dimethacrylate (EDMA) and diclazuril mol ratio are the polymkeric substance of 10:1,20:1,30:1,40:1,50:1, take each 20mg of molecularly imprinted polymer powder of above-mentioned 5 different ratioss respectively, add the diclazuril standardized solution 2mL of 1.0mmol/L, after vibration absorption 2h, survey the content remaining diclazuril in solution, then calculate the adsorptive capacity of molecularly imprinted polymer powder, result such as table 1 shows.
Table 1 molecularly imprinted polymer powder in proportion and the binding capacity of diclazuril
As seen from the above table, along with the increase of dosage of crosslinking agent, the binding capacity of MIPs also increases thereupon, and when n (EDMA): n (DIC)=30:1 reaches maximum value, adsorptive capacity reduces subsequently.Therefore, experimental selection n (EDMA): n (DIC)=30:1 is optimum proportion.
The template molecule of different ratio, function monomer, linking agent are on the impact of imprinted polymer adsorptivity after measured, and the ratio finally establishing the template molecule/function monomer/linking agent of 1:5:30 is optimum proportioning.
(3) methyl alcohol/acetic acid (8/2), acetonitrile/water (8/2), acetonitrile/water (7/3), acetonitrile/water (6/4) four kind of solution is contrasted to the removal effect of template.Result shows, acetonitrile/water (6/4) can not remove template molecule completely; Methyl alcohol/acetic acid (8/2), acetonitrile/water (7/3) remove template molecule completely all needs more than 80h, and acetonitrile/water (8/2) can remove template molecule completely and required time is shorter, so experimental selection acetonitrile/water (8/2) is as the solution of removing template.
(4) take the molecularly imprinted polymer that 3.00g prepares and put into apparatus,Soxhlet's, add acetonitrile/water (V:V=8:2) solution of 150mL, reflux in 95 DEG C of water-baths wash-out, within every 3 hours, get the liquid in an apparatus,Soxhlet's, then 0.22 μm of millipore filtration is crossed, detect with HPCE, the liquid detecting in apparatus,Soxhlet's is less than till diclazuril molecule, and after result shows 75h, molecularly imprinted polymer wash-out is complete.
(5) containing non-molecularly imprinted polymer NMIPs and molecularly imprinted polymer MIPs results contrast
The infrared spectra of non-molecularly imprinted polymer NMIPs and molecularly imprinted polymer MIPs is surveyed, with pure KBr for original pattern is tested, at wave-number range 4000 ~ 400cm with FTIR spectrum
-1measure.As shown in Figure 1, wave number 3456cm
-1and 2970cm
-1be respectively O-H stretching vibration and the methyl-CH of hydroxyl-OH
3c-H stretching vibration, wave number 1739cm-1,1638cm
-1, 1461cm
-1, 1261cm
-1, 1161cm
-1to be respectively in C=O, alkene C=C, C-C and C-O key stretching vibration absorption peak in C=C, aromatic nucleus.As shown in Figure 2, after wash-out, hydroxyl O-H stretching vibration peak moves to 3429cm
-1, the C-H stretching vibration peak of methyl moves to 2975cm
-1, in C=O, alkene, in C=C, aromatic nucleus, C=C, C-C and C-O key stretching vibration peak moves to 1729cm respectively
-1, 1634cm
-1, 1448cm
-1, 1256cm
-1and 1161cm
-1place.Comparison diagram 1 and Fig. 2, find the bulk migration of just stretching vibration peak value, figure moves towards consistent substantially, proves that MIP reaches whole removals of template.Meanwhile, the appearance at these peaks illustrates that MIP has the point of application forming hydrogen bond.
Before adopting S-4800 scanning electronic microscope directly to observe MIPs wash-out, after wash-out and the surperficial mode of appearance of NMIPs, to realize the visual understanding to sample.From Fig. 3,4,5, before wash-out, MIPs and NMIPs surface is more even, smooth, and quality is tight; And MIPs is comparatively coarse after wash-out, quality is loosened, coarse microsphere surface has a large amount of holes, may be template molecule by left by after wash-out, a large amount of holes, be conducive to contacting of binding site in polymkeric substance and substrate, thus make molecularly imprinted polymer have higher charge capacity and the higher identity to substrate.
Adopt the Scatchard assay binding capacity of MIPs and NMIPs to substrate molecule, result shows, MIPs is greater than the binding capacity of NMIPs to substrate molecule significantly to the binding capacity of substrate molecule.Illustrate that binding site that in printing process, template molecule stays in MIPs determines MIPs to the high affinity of template molecule and specific recognition.
Embodiment 2
Compare the MIPs of mass ratio 1:1,2:3,2:1,3:1 and diatomaceous filling effect.Comparative sample solution, by the speed of molecularly imprinted solid phase extraction column (MISPE) and the adsorption effect to template molecule, the results are shown in Table 2.
The selection of table 2 molecularly imprinted polymer solid-phase extraction column preparation condition
| Polymkeric substance: diatomite | 1：1 | 3：2 | 2：1 | 3：1 |
| Filtyration velocity (mL/min) | 2.5 | 1.8 | 1.5 | 0.5 |
| Adsorptive capacity (umol/g) | 16.5 | 20.2 | 28.5 | 30.0 |
As can be seen from table: when the adsorption effect of MIPs MISPE when diatomite ratio is 3:1 is better, loading speed is excessively slow, although when proportioning is 1:1 and 3:2 sample through-rate comparatively fast still adsorption effect is poor.Therefore optimum proportioning is selected to be 2:1.
The preparation of diclazuril solid-phase extraction column takes Solid-Phase Extraction (SPE) post that 60mgMIPs and 30mg diatomite (MIPs: diatomite=2:1) loads 3mL sky) in, compress with sieve plate, continue to rap pillar, until sieve plate be combined with polymkeric substance tight till.
Embodiment 3
(1) accurately take template molecule diclazuril (DIC) 0.408g to be dissolved in and to fill 20mLN, in the container of N-dimethylformamide, add function monomer methacrylic acid (MAA) 0.44mL, vibrate 30min under room temperature in ultrasonic apparatus, and template molecule is fully combined with function monomer; Then in container, add linking agent ethylene glycol dimethacrylate (EDMA) 3.86mL, Diisopropyl azodicarboxylate (AIBN) 0.05g, mixes, and sonic oscillation 15min is filled with N in container
2and seal, put in 60 DEG C of water-baths and heat 24h, obtain solid white polymer;
(2) solid white polymer of gained is pulverized by step (1), ground 220 orders (particle diameter 0.065mm) sieve, with the template molecule diclazuril in the acetonitrile-water of volume ratio 8:2 or 7:3 or 6:4 or the methyl alcohol/acetic acid solution eluted polymer of volume ratio 8:2, use deionized water eluting solvent again, collect product 60 DEG C of dryings in baking oven, obtaining take diclazuril as the imprinted polymer of template.
(3) in 3mL solid phase extraction column blank pipe, load glass wool from bottom to top successively and take diclazuril as the molecularly imprinted polymer of Template preparation, strike reality and make surface concordant, add one deck glass wool more above, loading height is about 0.5cm, makes diclazuril MISPE post.Diclazuril MISPE post is first with 2mL methyl alcohol, the activation of 3mL water; Get 1mL lysate, cross post with the flow velocity of 5mL/min; With the drip washing of 5mL normal hexane, abandon leacheate; Wash-out is carried out again with 5mL acetonitrile solution; Collect elutriant, dry up with Nitrogen evaporator, dissolve with 1.0mL60% acetonitrile-aqueous solution, after crossing 0.22 μm of millipore filtration, obtained upper HPCE detects liquid.
(4) accurately take 5.00g Chicken Tissues in 30mL centrifuge tube, add the standard mixing solutions 1.0mL of diclazuril anticoccidial drug, 10.0mL acetonitrile, 5.0mL normal hexane.Vortex vibrates, and the centrifugal 5min of 5000r/min, removes acetonitrile layer to another centrifuge tube, repeats extraction 1 time, merges acetonitrile layer.Add 5mL n-propyl alcohol, 50 DEG C of rotary evaporations, are concentrated into dry.With 1.0mL dissolve with methanol residue, add 4mL0.05mol/L potassium dihydrogen phosphate dissolution residual substance, cross diclazuril MISPE post, after normal hexane drip washing, acetonitrile, collect elutriant.Add 3.0mL n-propyl alcohol, 50 DEG C of rotary evaporations, are concentrated into dry, and use 1.0mL60% acetonitrile-water dissolution residual substance, cross 0.22 μm of millipore filtration, HPCE detects (see figure 5).
(5) as shown in Figure 6, compared with the chicken pre-treating process had been reported, purified by chicken extracting solution diclazuril MISPE, HPCE detects, and achieves good decontamination effect improving.
In order to verify that the accuracy that diclazuril in chicken remains pre-treating process has carried out recovery testu, respectively to adding 2.0 in blank chicken extracting solution, 5.0, the diclazuril reference liquid of a 10.0mg/kg3 mass concentration level, extract by the method for (4), purify after detect with HPCE, each Pitch-based sphere replication 3 times.Result shows, and after diclazuril MISPE process, it is 5.7% that the diclazuril rate of recovery can reach 87.6%, RSD, meets wild animal resources requirement.
Claims (1)
1. the molecular imprinting polymkeric substance of anticoccidial drug diclazuril remains the application in pre-treatment at detection chicken diclazuril, it is characterized in that comprising the steps:
One, the preparation of molecularly imprinted polymer
(1) take quality 0.408g template molecule diclazuril (DIC) and be dissolved in 20mLN, in dinethylformamide solution, add function monomer methacrylic acid (MAA) 0.265-0.795mL, sonic oscillation 30min under room temperature, makes template molecule fully be combined with function monomer; Then linking agent ethylene glycol dimethacrylate (EDMA) 1.29-6.43mL is added; Add Diisopropyl azodicarboxylate (AIBN) 0.05g again, mix, sonic oscillation 15min, at N
2protect in underlying 60 DEG C of water-baths and heat 24h, obtain solid white polymer;
(2) solid white polymer of step (1) gained is pulverized, ground 220 mesh sieves, with the template molecule diclazuril in the acetonitrile-water of volume ratio 8:2 or 7:3 or 6:4 or the methyl alcohol/acetic acid solution eluted polymer of volume ratio 8:2, use deionized water eluting solvent again, collect product 60 DEG C of dryings in baking oven, obtaining take diclazuril as the imprinted polymer of template;
Two, the preparation of solid-phase extraction column
In 3mL solid phase extraction column blank pipe, load glass wool from bottom to top successively and take diclazuril as the molecularly imprinted polymer of Template preparation, strike reality and make surface concordant, add one deck glass wool more above, loading height is 0.5cm, makes diclazuril MISPE post;
Three, diclazuril composition detection in chicken
Accurately take 5.00g Chicken Tissues in 30mL centrifuge tube, add the standard mixing solutions 1.0mL of diclazuril anticoccidial drug, 10.0mL acetonitrile, 5.0mL normal hexane; Vortex vibrates, and the centrifugal 5min of 5000r/min, removes acetonitrile layer to another centrifuge tube, repeats extraction 1 time, merges acetonitrile layer; Add 5mL n-propyl alcohol, 50 DEG C of rotary evaporations, are concentrated into dry; With 1.0mL dissolve with methanol residue, add 4mL0.05mol/L potassium dihydrogen phosphate dissolution residual substance, cross solid-phase extraction column prepared by step 2, after normal hexane drip washing, acetonitrile, collect elutriant; Add 3.0mL n-propyl alcohol, 50 DEG C of rotary evaporations, are concentrated into dry, and use 1.0mL60% acetonitrile-water dissolution residual substance, cross 0.22 μm of millipore filtration, HPCE detects.
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| CN102174148A (en) * | 2011-03-15 | 2011-09-07 | 安徽出入境检验检疫局检验检疫技术中心 | Preparation of triazine phytocide molecular imprinting solid phase extracting material |
| CN102532390A (en) * | 2011-12-05 | 2012-07-04 | 中国农业科学院农业质量标准与检测技术研究所 | Triazine weedicide, and metabolite molecular engram polymer microspheres, preparation method and application thereof |
| CN102924645A (en) * | 2012-11-21 | 2013-02-13 | 安徽农业大学 | Preparation method and application of molecularly imprinted polymer of penicillin antibiotics and intermediate of penicillin antibiotics |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102174148A (en) * | 2011-03-15 | 2011-09-07 | 安徽出入境检验检疫局检验检疫技术中心 | Preparation of triazine phytocide molecular imprinting solid phase extracting material |
| CN102532390A (en) * | 2011-12-05 | 2012-07-04 | 中国农业科学院农业质量标准与检测技术研究所 | Triazine weedicide, and metabolite molecular engram polymer microspheres, preparation method and application thereof |
| CN102924645A (en) * | 2012-11-21 | 2013-02-13 | 安徽农业大学 | Preparation method and application of molecularly imprinted polymer of penicillin antibiotics and intermediate of penicillin antibiotics |
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| Behaviour of triazine herbicides and their hydroxylated and dealkylatedmetabolites on a propazine-imprinted polymer Comparative study in organic and aqueous media;R.Carabias-Martinez, et al.;《Analytica Chimica Acta》;20060117;第559卷(第2期);第186-194页 * |
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