【作者】 刘美华；

【导师】 陈曦；

【作者基本信息】 福州大学 ， 食品安全与药物化学， 2010， 博士

【摘要】 近年来，农业生产以及农副产品加工过程中滥用药物导致其在环境和食品中的残留现状已经引起人类的高度重视。关于这类药残的分离分析方法的研究显得极其重要。本论文以一些常见的药物如杀虫剂、雌激素等为研究对象，主要从两个方面进行探讨：一，新型的样品前处理方法在药残分析检测中的应用；二，分子印迹复合膜在药物分离分析中的应用。本论文研究的主要目的是建立微型化、环境友好、易操作、快速、成本低廉、抗干扰能力强、能适应复杂基质的样品前处理方法以及制备对目标物质具有特异识别能力的分子印迹复合膜，并应用于一些常见药物的分离分析中。本论文共分为三部分，共六章，分述如下：论文的第一部分（第一章）绪论部分首先介绍了常见的样品前处理方法、现状及特点、样品前处理方法中的新技术以及分子识别技术中分子印迹膜的制备与应用现状；并对采用前处理方法进行药残分析的相关文献报道进行综述，探讨其发展趋势；最后简要介绍了本论文的研究目的和研究内容。论文的第二部分（包括第二章、第三章、第四章）分别以杀虫剂和雌激素为研究对象，探讨了三种液相微萃取前处理方法，并应用于实际样品中的残留分析。第二章通过设计新的前处理装置，建立了中空纤维管保护液相微萃取结合高效液相色谱分离检测三种雌激素的新方法。实验对液相微萃取条件如：有机溶剂、给相与接受相的体积比、提取时间、搅拌速度以及给相与接受相的pH值等进行了优化。一段长45mm的中空纤维管在正辛醇中饱和后，去除内腔中的正辛醇。然后将一端封口，管内腔充入0.5mol/L NaOH溶液10μL作为接受相，浸入10mL试剂瓶中。调节溶液pH至1.5，经过40min提取达到平衡，接受相直接进入HPLC系统。在优化的条件下，三种雌激素的富集倍数均超过300倍，检测限为0.25～0.5ng/mL (信噪比S/N=3)。该方法简单、富集倍数高、费用低廉、快速、有机溶剂消耗少，检测限较低。所建立的方法可用于养殖鱼塘废水样品中己烯雌酚、双烯雌酚和己烷雌酚残留的分析检测，回收率在89%以上。论文第三章的研究工作采用混合溶剂作为萃取相，建立了一种新型的单滴悬挂式液相微萃取前处理方法，并结合GC-MS对蔬菜中9种有机氯农药进行分析。试验通过对有机溶剂、悬挂液滴体积、萃取时间、搅拌速度等进行考察，优化了单滴悬挂式液相微萃取的萃取条件。试验优化的结果为：采用丙酮与二甲苯的混合溶剂（体积比为2/8）作为萃取溶剂，微量进样器悬挂液滴体积1.00μL，样品溶液体积2mL，搅拌速度400rpm，萃取时间30min。在优化的微萃取条件下，采用该方法结合GC-MS对9种有机氯农药进行分析。9种物质的线性范围如下：α-, β-, γ-, δ-六氯苯和氯苯三氯乙醇为0.05～20ng/mL，六氯萘和DDD为0.5～20ng/mL， DDE和DDT为0.5～50ng/mL。各物质的检测限（S/N=3）：α-, β-, γ-, δ-六氯苯为0.05ng/mL，氯苯三氯乙醇、六氯萘和DDT为0.2ng/mL，回收率为63.3%～100.0%，相对标准偏差为8.74%～18.9%。试验结果表明所建立的单滴悬挂液相微萃取作为一种快速而简单的前处理方法，可用于蔬菜样品中有机氯农药的前处理中。论文第四章的研究工作将分散液相微萃取中的有机溶剂采用当前热门的离子液体代替，并借助超声辅助萃取技术与分散液相微萃取方法相结合，建立了离子液体-超声辅助分散液相微萃取技术的前处理方法，讨论并优化了萃取的各种条件，结合高效液相色谱分离检测技术实现对雌激素类物质的检测。试验采用50μL的离子液体，考察了溶液体积、溶液pH值、超声时间、静置时间、离心时间等因素对富集效果的影响。最佳的萃取条件为：溶液体积为7.5mL，甲醇体积为0.3mL，溶液pH值为2.0，超声时间为6min，静置时间为30min，离心时间为10min。在优化的萃取条件下，三种雌激素的富集倍数可达到100倍左右；方法的线性范围为1.0～100.0ng/mL；检测限为0.55～1.0ng/mL。对浓度为5.0ng/mL的三种物质测定6次的日内相对标准偏差为9.2%～10.8%。最后将所建立的前处理方法分别用于生活污水和牛奶厂废水中雌激素残留的检测。论文的第三部分（包括第五章、第六章）工作分别以商业膜材料为载体，合成了两种分子印迹复合膜材料，并用于几种雌激素及其结构类似物的前处理和分离中。论文第五章工作介绍了设计合成己烯雌酚的分子印迹-中空纤维管（MIP-HFT）复合材料，并用于对己烯雌酚及其结构类似物的识别。对于合成的复合膜的表面特性、吸附能力及选择性试验进行了研究。为考察所合成材料对目标分子的选择识别能力，选用双烯雌酚（DS）、己烷雌酚（HS）作为模板分子的结构类似物，苯酚和甲苯作为参考物质。将所合成的分子印迹复合材料通过自组装的吸附装置，对三种物质混合液进行微萃取前处理，以HPLC为分析检测手段，并应用于对牛奶中三种雌激素物质残留进行分析。三种物质的检测限为2.5～3.33ng/mL，牛奶中添加回收率范围为83.7%～90.6%。表明所合成的分子印迹中空纤维管对己烯雌酚及其结构类似物双烯雌酚,己烷雌酚具有较好的选择性识别能力，能用于复杂样品中这类物质分析的前处理技术中。该分子印迹复合膜对己烯雌酚及其结构类似物具有很好的特异选择性。论文第六章工作以商业多孔乙酸纤维膜为载体，17β-雌二醇为模板，采用紫外光引发方法制备了17β-雌二醇乙酸纤维分子印迹复合膜。试验对膜的选择吸附性和分离透过性进行了探讨。实验结果表明，所合成的乙酸纤维分子印迹复合膜对模板分子17β-雌二醇具有较好的选择吸附性，吸附量达10.2μmol／g；17β-雌二醇／17α-炔雌醇的分离因子为4.7。膜透过实验结果表明，该分子印迹复合膜能将17β-雌二醇与17α-炔雌醇分离开，表明17α-炔雌醇作为结构类似物在复合膜上的结合牢固性不如模板分子。更多还原

【Abstract】 For the past few years, drug residues in environment and food had evokedpeople’s attention because of drug abuse in agricultural production andagricultural products. In this thesis, we selected some pesticides and estrogen astarget and focused on these two problems. One was sample preparation indetermination of drug residues and the other was molecular imprinting membrane inapplication in seperation and determination of estrogen. The aim purpose of thethesis was to develop sample pretreatment with high enrichment fact,miniaturization, environment friendly, easily operation, less time, cheap andanti-interference, and molecular recognition technologies with selectivilyrecognition capability to target molecular, adapting complicated matrix, andapplication in seperation and determination of some estrogen.In general, this thesis is divided into three parts, including six chapters. Thefirst part is also the first chapter. In this chapter, we reviewed the recentdevelopment of sample preparation technology and the molecular imprintingmembrane, and we also proposed the future prospects of sample preparationtechnology. At the end of this chapter, we briefly introduced the research goals aswell as the contents and achievements of this thesis.The second part of the thesis includes three chapters, which is chapter2,chapter3and chapter4. This part mainly focuses on the sample preparation(hollow fiber-mediated liquid-phase microextraction, single-drop liquid-phasemicroextraction and dispersive liquid-phase microextraction). In chapter2, weestablished three-phase hollow fiber-mediated liquid-phase microextractionfollowed by HPLC for the determination of three synthetic estrogens, namelydiethylstilbestrol, dienestrol and hexestrol. Extraction conditions including organicsolvent, volume ratio between donor solution and acceptor phase, extraction time,stirring rate, donor phase and acceptor phase were optimized. The targetcompounds were extracted from a10mL aqueous sample at pH1.5(donorsolution) through a45mm in length hollow polypropylene fiber that wasimmersed in1-octanol in advance, and then the hollow fiber filled with10μL0.5mol/L sodium hydroxide solution (acceptor phase). After a40min extraction, the acceptor phase was directly injected into a HPLC system for detection. Under theoptimized extraction conditions, a large enrichment factor (more than300fold)was achieved for the three estrogens. The determination limit at an S/N of3ranged from0.25to0.5ng/mL for the estrogens. The recovery ratio was more than89%in the determination of these estrogens in wastewater.In chapter3, A new approach for the extraction of nine kinds oforganochlorine pesticides (OCPs) from vegetable samples coupling single-dropmicroextraction with gas chromatography–mass spectrometry was presented.Experimental parameters, such as organic solvent, exposure time, agitation andorganic drop volume were controlled and optimized. An effective extraction wasachieved by suspending a1.00μL mixed drop of acetone and p-xylene (2:8, v/v) tothe tip of a micro-syringe immersed in a2mL donor aqueous solution and stirredat400rpm. The approach was applied to the determination of OCPs in vegetablesamples with a linearity range of0.05to20ng/mL for α-, β-, γ-,δ-hexachlorobenzene (BHC) and dicofol,0.5to20ng/mL for dieldrin and2,2-bis(4-chlorophenyl)-1,1-dichloroethane (DDD) or0.5to50ng/mL for2,2-bis(4-chlorophenyl)-1,1-dichloroethylene (DDE) and2-(2-chlorophenyl)-2(4-chlorophenyl)-1,1,1-trichloroethane (p,p’-DDT). Correspondingly, thedetermination limit at an S/N of3ranged from0.05ng/mL for α-, β-, γ-, δ-BHC to0.2ng/mL for dicofol, dieldrin or p,p’-DDT. The relative recoveries were from63.3%to100.0%, with repeatability ranging from8.74%to18.9%(relativestandard deviation, RSD). The single-drop microextraction was proved to be a fastand simple approach for the pre-concentration of organochlorine pesticides invegetable samples.In chapter4, a novel method was investigated for the determination ofestrogens in wastewater by ultrasonic assisted ionic liquid (1-butyl-3-methylimidazolium hexafluorophosphate,([C4MIM][PF6])dispersive liquid-phasemicroextraction coupled with high performance liquid chromatography. Someconditions that might affect the extraction efficiency such as volume of solution,pH value of solution, ultrasonic time, hierarchical standing time and centrifugingtime, were optimized. The results indicated that the optimal conditions were7mLsolution at pH2.0for6min of ultrasonication,30min of hierarchical standing and10min of centrifugation. Under the above mentioned conditions, the extractionefficiency of ionic liquid based ultrasonic microextraction for the three estrogens were about100. Their linear ranges and detection limits were1.0～100.0ng/mLand0.55～1.0ng/mL respectively. The proposed method was applied to determinethe three estrogens in wastewater. The recoveries of the three estrogens spiked inwastewater were in the range of89.0%～96.6%.The third part of this thesis mainly focuses on the synthesis and application ofmolecular imprinting membrane. This part includes chapter5and chapter6. Inchapter5, we synthesized a novel molecularly imprinted polymer (MIP)-coatedpolypropylene hollow fiber tube (HFT) of diethylstilbestrol (DES) as a templatemolecule, α-methacrylic acid (MAA) as a functional monomer, and ethyleneglycol dimethacrylate (EGDMA) as a crosslinking agent. The characteristics andapplications of the MIP-coated hollow fiber tube (MIP-HFT) were investigated. Inorder to compare the characteristics with the non-imprinted polymer-immergedhollow fiber tube (NIP-HFT), the selectivity of the MIP-HFT was investigatedwith dienestrol and hexestrol as the structural analogues of diethylstilbestroltemplate, and phenol and phenolphthalein were taken as reference compounds.The MIP-HFT was employed in HPLC application for milk samples. The detectionlimits were found to be in the range of2.5to3.33ng/mL for DES, dienestrol andhexestrol, and their recoveries were83.7to90.6%, respectively, in spiked milksamples. The experimental results revealed that the MIP-HFT provides a goodcarrier for the selective adsorption of DES and its analogs in chemical structure,and can be used for the pre-concentration of these compounds in complicatedsamples.In chapter6, we prepared the17β-Estradiol molecularly imprintedcomposite membrane through UV initiation by using porous cellulose acetatemembrane as support. The selective binding properties and separation capacity ofthe membranes were investigated．The results showed that the17β-Estradiolmolecularly imprinted composite membrane had high binding capacity for17β-Estradiol，the binding amount Q for17β-Estradiol was10.2μmol／g andseparation factor was4.7for17β-Estradiol／17α-ethinylestradiol．Permeationexperiment results showed that the molecularly imprinted composite membranecould separate17β-Estradiol from17α-ethinylestradiol. The bonding forcebetween17α-ethinylestradiol and the membrane was weaker than the template.更多还原