The Preparation Of Hydrophilic Molecularly Imprinted Polymers For Determination Of Olaquindox

Olaquindox as an antimicrobial growth accelerant, is usually added into chick feed tocontrol chick dysentery and bacterial enteritis in young chick. However, due to the possiblecarcinogenic, mutagenic, and photoallergenic effects, its use as an additive in animalfeedstuffs have been prohibited in the European Union and many other countries. In order toprotect human health, an accurate and reliable analytical method for the determination ofolaquindox in the feed samples is required. The aim of this paper is to study the determinationof preconcentration of olaquindox using molecular imprinting technology in aqueous phase.MIT is one of the most promising approaches whose principle came from “Key-Lock”. In thepresent work, we prepared four new hydrophilic functionalized materials in aqueous solutionby a series of molecular imprinting techniques in combination with high pressure liquidchromatography or enzyme-linked immunospot assay.（1） Separation and determination of olaquindox using molecularly imprinted solid-phase extraction coupled with high-performance liquid chromatographyA new and hydrophilic molecularly imprinted polymer （MIP）, selective for olaquindox,was prepared by bulk polymerization technique using olaquindox as the template, methacrylicacid as the functional monomer, and ethylene glycoldimethacrylate as the cross-linker. Thesynthesized polymer was characterized by Fourier transform infrared and static adsorptionexperiments, and the results showed that the MIP had good recognition and selective abilityfor olaquindox. A novel method of molecularly imprinted solid-phase extraction coupled withhigh-performance liquid chromatography （HPLC） was developed for separation anddetermination of trace olaquindox in feed samples. Under the selected experimental condition,the detection limit （S/N=3） was38.0ng L^-1, and the RSD for five replicate extractions of50μg L^-1olaquindox was4.9%. This method was employed for quantitative determination ofolaquindox in fish feed with recoveries ranging from89.8⁹7.4%.（2） Chitosan beads as molecularly imprinted polymer matrix for selectiveseparation of olaquindox in aqueous phaseA simple molecularly imprinted polymer （MIP） was prepared using olaquindox as theimprinted molecule and acrylamide as the functional monomer. The MIP was achieved bygrafting of the selective soft polyacrylamide gel to the chitosan beads by letting the monomers and the template molecule diffuse into the pores of the chitosan matrix before starting thepolymerization. This prepared material was evaluated by FT-IR spectra, SEM images, andstatic adsorption experiments. The adsorption test indicated that the imprinted polymerexhibited higher selectivity and adsorption capacity toward olaquindox than that ofstructurally related compounds. The adsorption capacity of the imprinted polymer were10.14mg g^-1. The MIP has much higher adsorption capacity for OLA than the non-imprintedpolymer with the same chemical composition, and also has a higher selectivity for theimprinted molecule.（3） Core-shell magnetic molecularly imprinted solid phase extraction coupled withhigh-performance liquid chromatography for recognition of olaquindox.The magnetic nanoparticles were synthesized by the chemical coprecippitation of Fe2+and Fe3+in an ammonia solution. Subsequently, silica was coated on the Fe₃O₄nanoparticlesusing a sol-gel method to obtain silica shell magnetic nanoparticles. Subsequently,acrylamide（AA） as the functional monomer, and ethylene glycoldimethacrylate（EGDMA） asthe cross-linker. The synthesized polymer was coated onto the silica-modified Fe₃O₄surfacethrough oxidation with AIBN in an aqueous solution. The polymer was characterized byFourier transform infrared spectra, static adsorption experiments and thermogtavimetricanalysis. The results showed that the MIP had good recognition and selective ability forolaquindox. A novel method of molecularly imprinted solid-phase extraction coupled withhigh-performance liquid chromatography （HPLC） was developed for separation anddetermination of trace olaquindox in feed samples. Under the selected experimental condition,the detection limit （S/N=3） was32.0ng L^-1, and the RSD for five replicate extractions of50μg L^-1olaquindox was4.7%. This method was employed for quantitative determination ofolaquindox in fish feed with recoveries ranging from90.2%⁹8.2%.（4） Development of a biomimetic Enzyme-Linked Immunosorbent Assay Methodbased on a hydrophilic molecularly imprinted polymer film for determination ofolaquindoxWe developed a fast and new competitive biomimetic enzyme-linked immunosorbentassay （BELISA） method for the determination of olaquindox in chick feed based on a novelmolecularly imprinted film as an artificial antibody. The imprinted film was directlysynthesized on the well surface of Maxisorp polystyrene96-well plate by bulk polymerizationwith molecular imprinting technique. Then the experimental conditions which areconcentration of enzyme conjugate, diluent choice and pH values were optimized through theexperiments followed by the establishment of the standard curve. Using it as the recognition element, a fast and new direct competitive biomimetic enzyme-linked immunosorbent assay（BELISA） method for the determination of olaquindox in feed was developed. This BELISAmethod had low cross-reactivities of6.2%and12%for two analogues. Under the optimalconditions, the sensitivity (IC₅₀) and the limit of detection (IC₁₅) were700±60μgL^-1and17±1.6μg L^-1, respectively. The blank chick feed samples spiked with olaquindox at threelevels were determined by this developed method with recoveries ranging from89.0%⁹6.0%...