再沉淀法制备复合纳米药物及体外抗肿瘤研究

Preparation of composite nano drugs by reprecipitation and Study on its antitumor activity in vitro

摘要:

摘要：目的：为了解决药物传递系统中载体材料和药-药共价键缀合的新药带来的不确定威胁及采用更简单的药物传输系统；方法：将两种不同的抗肿瘤药物阿霉素（DOX）和苯丁酸氮芥(Cb)混合,利用再沉淀法制备一种新型的复合药物纳米粒子，并采用1H核磁（1H NMR）、DLS、SEM、AFM、TEM及MTT、Hoechst染色、Western、共聚焦实验等证明制备了复合药物纳米粒子及体外抗肿瘤活性；结果：1H NMR证明了DOX和Cb混合后通过氨基和羧基静电力相互作用形成超分子复合物，DLS、SEM、AFM、TEM表征证明了复合药物纳米粒子具有规整的形貌，较窄的尺寸分布，三种肿瘤细胞模型（MCF-7、A549、HepG2）MTT实验证明复合药物纳米粒子相比于单独的DOX、Cb、DOX/Cb混合物有着更好杀死肿瘤细胞的效果．采用Hoechst染色检测凋亡细胞成像以及Western说明复合药物纳米粒子通过诱导细胞凋亡杀伤肿瘤，共聚焦实验则验证了相比于自由药阿霉素（DOX），复合药物纳米粒子可以更容易进入肿瘤细胞；结论：通过DOX与Cb静电作用力，采用再沉淀的方法可制得复合纳米粒子，且这种策略制备的复合纳米粒子有着更好的抗肿瘤效果．

Abstract:

Abstract：Objective:To solve these uncertain threat that carrier materials and a new drug which is synthesized by drug-drug conjugate through covalent bond bring, insteading by more easy drug delivery system through non-covalent bond. Methods: A novel composite drug nanoparticles (DOX-Cb nanoparticles) have been prepared by reprecipitation after hydrophobic anticancer drug Doxorubicin (DOX) and hydrophobic anticancer drug chlorambucil (Cb) being mixed. Then using 1H nuclear magnetism(1H NMR), DLS,SEM,AFM ,TEM, Hoechst staining,western and confocal experiment confirm that DOX-Cb nanoparticles,which have antitumor activity in vitro, are prepared. Results: 1H NMR confirms that DOX and Cb compound is formed via electrostatic force between amino group and carboxyl group. DLS,SEM,AFM and TEM confirm that Composite drug nanoparticles have regular morphology and a narrow size distribution. Cytotoxicity test by using three tumor models（MCF-7，A549，HepG2） shows Composite drug nanoparticles have better effect of killing tumor cells than free DOX and Cb. Hoechst staining and western’result suggest that Composite drug nanoparticles kill tumor by inducing tumor cells apoptosis. confocal experiment verifys DOX-Cb nanoparticles are easier endocytosed by tumor cells than DOX. Conclusion: A novel composite drug nanoparticles (DOX-Cb nanoparticles) have been prepared by reprecipitation through electrostatic force between DOX and Cb, and composite drug nanoparticles were prepared by this strategy having better antitumor effect.