头孢喹诺肺靶向微球在动物肺组织及巨噬细胞的选择性分布研究

The efficient, safe, effective, targeted drugs for the treatment of animal respiratory disease is an urgent issue to be addressed in breeding industry. And the study on influencing factors of the preparation of targeting microsphere and its selective distribution in lung tissues and macrophages is also very important to the development of clinical veterinary in China. However, the study on microsphere is still in primary stage in China. And the researchs on influencing factors of the preparation of animal-specific drugs targeting microsphere, its targeting mechanism and selectively distribution in lung tissues and macrophages have not been reported in literatures. In this study, poly lactic / glycolic acid copolymer ( PLGA ) is selected as carrier material, and cefquinome is selected as the representative of cephalosporins drug. The influencing factors of the preparation of cefquinome targeting microsphere with different methods will be studied. The targeting microspheres prepared will be studied comprehensively, and then preparation technology of targeting microsphere will be obtained and the influential factors of the release rule of microspheres and correlation between in vivo and in vitro, etc will be made an intensive study. Moreover, methods of 125I markers and UPLC-MS/MS are studied in biological samples, its selective distribution in lung tissues and macrophages of rat/mice , rabbit and target animal dog in vivo will be studied. Finally,WinNonlin 5.2 and DAS 1.0 software package was used for the analysis of the time and concentration data.The mechanism of lung targeting and pharmacokinetic characteristics of its distribution in tissues in vivo will be revealed, which will lay a solid foundation for the application of targeting microspheres in veterinary clinical.

高效、安全、长效、靶向的动物呼吸道病治疗药是动物养殖生产中急需解决的问题，研究靶向微球制备影响因素及探究对肺组织和巨噬细胞的选择性分布对我国兽医临床具有重要意义。而我国动物专用靶向制剂研究尚处于起步阶段，对动物专用靶向微球制备影响因素、靶向机理及对组织选择性分布的研究也未见报道。选用PLGA等载体材料，头孢喹诺为代表药物，研究微球制备影响因素，通过对靶向微球的性能表征，获得较理想的靶向微球制备工艺，对影响微球释放规律的因素及体内外相关性等做深入研究。利用125I示踪标记及建立UPLC-MS/MS生物样品检测方法，进而研究头孢喹诺靶向微球对大/小鼠、家兔肺组织和巨噬细胞的选择性分布；以靶动物犬为模型，进行肺组织和巨噬细胞的选择性分布研究，揭示其肺靶向机理及体内组织分布的动力学特征，通过WinNolin5.2及DAS1.0软件分析靶向性评价参数，为靶向制剂在兽医临床的开发应用奠定基础。

开发出高效、安全、长效、靶向的动物呼吸道病治疗药对我国兽医临床的发展具有重要意义。利用明胶、PLGA、PLA等聚合物作为载体材料，通过乳化交联法、复乳化法、喷雾干燥法制备微球，以形貌表征、粒径大小分布、载药量、包封率和药物体外释放速率为指标，优化靶向微球制备工艺，成功制得2种头孢喹诺微球制剂，总结出一套制备头孢喹诺靶向微球的影响因素规律。完成了对其各项指标评价和组织中药物含量测定及肺靶向性分析。对大鼠肺组织的选择性分布研究表明肺组织中药物浓度最高，微球主要聚集在肺部；所获得的两种微球制剂在大鼠体内具有明显的肺靶向性，建立了大鼠组织心、肝、脾、肺、肾中头孢喹诺药物的UPLC/MS/MS检测方法，该方法简易快捷、合适可靠。以普通制剂“克百特”作为阳性药物对照，微球制剂缓释效果明显，具有较强的肺靶向，达到缓释目的，增加了药物靶向性。完成了病理模型对头孢喹诺肺靶向微球在大鼠肺组织及巨噬细胞选择性分布及功能影响。成功建立了以肺炎克雷伯杆菌感染大鼠肺炎模型，研究结果表明，细菌感染严重降低大鼠的免疫功能，经过普通制剂和靶向制剂治疗后，在4、12、24、48h巨噬细胞的数目和吞噬能力显著提高，有利于肺部细菌的清除；经靶向制剂治疗后细菌数量显著减少，明显改善细菌感染有引起的大鼠肺组织损伤，与普通制剂“克百特”相比，治疗效果更好。完成了头孢喹诺靶向微球在靶动物猪体内对肺组织及巨噬细胞的选择性分布研究、药代动力学和安全性试验。研究结果表明微球在静脉注射后快速浓集于肺组织，5分钟时单位重量的肺组织中聚集的微球数量最多，制备的头孢喹诺微球具有显著的肺靶向性；药动学参数结果表明，靶向制剂的消除半衰期比普通制剂的显著延长，且AUC值也明显大于普通制剂组，靶向制剂在血浆中的缓释性能良好；靶动物安全试验结果表明，肺靶向微球的低、中、高剂量对猪的呼吸系统、心血管系统、消化系统、泌尿系统、神经传导系统以及体内电解质和微量元素均无影响。推荐剂量（2 mg/kg）一次给药治疗肺部疾病时，其使用是安全的。本研究通过揭示靶向微球药物的体内作用机制、代谢机制、促进动物临床靶向制剂药物的开发，为其他药物靶向微球制剂在兽医临床的应用提供方法和手段。