【作者】 赵莉莉；

【导师】 王晓岚；

【作者基本信息】 江南大学 ， 营养与食品卫生学， 2011， 硕士

【摘要】 人工合成的新型抗菌药物恩诺沙星（enrofloxacin）是第三代喹诺酮类药物（FQNs）之一,它通过抑制细菌DNA螺旋酶而抗菌,具有抗菌谱广、杀菌力强、体内分布广泛及与其它抗生素之间无交叉耐药性等特点,在预防和治疗畜禽的细菌性感染及支原体病方面取得良好的效果,现已广泛地应用于兽医临床。但由于致病菌产生耐药性和长期应用所产生的不良反应以及潜在的致癌性,其残留问题已引起广泛关注。许多国家和组织等都将其列入限制使用的兽药范围。本研究采用时间分辨荧光免疫分析法（TRFIA）,建立了恩诺沙星的高灵敏的检测方法。方法采用稀土离子Eu³⁺标记技术建立恩诺沙星时间分辨免疫检测法并对其进行方法学的考核,其检测的灵敏度可达5 ng/L,测量范围为0.01μg/L～100μg/L:向鳗鱼、猪肉、鸡肉样品中分别添加0.1μg/kg、10μg/kg、200μg/kg三个水平的恩诺沙星标准品,蜂蜜样品中添加0.34μg/kg、34.3μg/kg、342.8μg/kg,各样品的平均回收率分别为89.1℅、88.4℅、89. 0℅、89.4℅。方法的批内变异系数小于10 %,批间变异小于15 %。特异性实验表明恩诺沙星抗体与其他喹诺酮类药物交叉反应均较低,抗体的特异性很好。同时,初步建立了恩诺沙星纳米均相发光免疫检测法,并简要的对方法的稳定性、样品添加回收率等方面进行了考核。恩诺沙星纳米均相发光免疫检测法两步共30 min的反应模式大大缩短了检测时间,同时又保证了良好的精密度,批内、批间变异率均小于5 %;方法的灵敏度为2 ng/L,线性范围为0.007¹00μg/L。研究表明,恩诺沙星-时间分辨免疫检测法是目前已报道文献中检测恩诺沙星的最灵敏的方法,具有很好的应用前景。同时,本论文还对新兴的纳米均相技术进行了较全面的探讨,并初步建立了恩诺沙星纳米均相发光免疫检测法,为我国这一领域以及相关领域的研究提供了很有价值的参考资料。更多还原

【Abstract】 Enrofloxacin, the newly artificially synthesized antimicrobial drug, is one of the third generation quinolone derivatives. It functions effectively on preventing and treating bacterial infection by inhibiting bacterial DNA gyrase. Currently, it is used widely in veterinary clinical attributing to its broad spectrum of activity, strong bactericidal,widely distributed in vivo and no cross-resistance with other antibiotics. However, it is concerned widly of the residues because of its resistance, adverse reaction and potential carcinogenic trait. Many countries and organizations have included it to the scope of restrictions on the use of veterinary drugs.The main purpose of this thesis is to develop the ultrasensitive detection for enrofloxacin by time-resolved fluoroimmunoassay（TRFIA）.The method is to establish the Enrofloxacin -TRFIA by europium (Eu³⁺) marker and to carry out the evaluation methodology. The sensitivity of detection was 5 ng/L and the practical linear response was from 0.01 to 100μg/L.The average recoveries of detection for enrofloxacin spiked in eel, pork, chicken samples were 89.1℅,88.4℅,89. 0℅ when the added enrofloxacin were 0.1μg/kg,10μg/kg and 200μg/kg respectively,while the honey sample was 89.4℅, with enrofloxacin added 0.34μg/kg,34.3μg/kg and 342.8μg/kg respectively. The intra-assay variation of the method was less than 10℅, and the inter-assay variation was less than15℅.The specificity test indicated that the antibody of Enrofloxacin is low cross reaction with other quinolones and the antibody enables high specificity.Furthermore, we also initially established a homogenously indirect competitive luminescent oxygen channeling immunoassay（LICLIA） for the detection of enrofloxacin and examined constancy and recovery of the assay briefly. It only took 30min of this immunoassay to complete the detection. The results indicated that the detection limit of the assay was 2ng/L for homogenously indirect competitive LICLIA at a practical working concentration range from 0.007 to 100μg/L.The research indicated that Enrofloxacin-TRFIA was the most sensitive method of detecting Enrofloxacin reported so far,and it has good prospects of application.Furthermore, the thesis discussed fully on the newly emerged homogeneous nanotechnology,and the enrofloxacin-LICLIA has been initially established, which would provide great valuable reference to this area.更多还原