阿霉素修饰纳米银的制备及其体外抗肿瘤活性研究

Preparation of doxorubicin modified silver nanoparticle and its anti-tumor activity in vitro

目的设计合成一类新型的具有p H响应性的阿霉素-纳米银(DOX-Ag NPs)联合抗肿瘤药物,对其理化性质进行表征,并研究其体外响应性释药行为和抗肿瘤活性。方法通过硫辛酰肼(LA-NHNH2)连接纳米银(AgNPs)和阿霉素(DOX),得到DOX-Ag NPs。利用核磁氢谱(1H NMR)和高分辨质谱(HRMS)对硫辛酰肼-阿霉素(LA-NHN=DOX)进行结构确证;通过动态光散射(DLS)和透射电镜(TEM)分析纳米粒的粒径和形貌;通过紫外-可见吸收光谱和荧光光谱表征纳米粒的光学性质;通过透析法结合荧光光谱检测DOX-Ag NPs在不同p H下的DOX释放行为;采用噻唑蓝比色法研究DOX-Ag NPs对Hep G2肿瘤细胞的增殖抑制效果。结果 LA-NHN=DOX的1H NMR数据及HRMS检测到746.275 6处的分子离子峰均证明LA-NHN=DOX成功合成。DOX-Ag NPs为粒径(40.4±3.8)nm的球形纳米粒;在弱酸性条件下DOX-Ag NPs能够快速响应性释放DOX;DOX-Ag NPs对Hep G2肿瘤细胞增殖抑制呈现浓度依赖性,当DOX浓度为0.5~20 mg/L(Ag浓度为0.45~18 mg/L)时,DOX-Ag NPs组细胞生存率均明显低于DOX组和Ag NPs组(均P<0.05)。结论 DOX-Ag NPs是一种具有p H响应性的联合抗肿瘤纳米制剂,能在肿瘤组织快速释放DOX,并通过与Ag NPs的协同治疗,发挥良好的体外抗肿瘤作用。

Objective To design and synthesize a novel type of combined anti-tumor drug-doxorubicin modified silvernanoparticles（DOX-Ag NPs） with p H response,characterize its physical and chemical properties,and investigate its drugresponsive release and anti-tumor activity in vitro.Methods DOX-Ag NPs were prepared by coupling silver nanoparticles（Ag NPs） with doxorubicin（DOX） via a LA-NHNH2 linker.The structure of LA-NHN=DOX was confirmed by nuclearmagnetic resonance（1 H NMR） and high resolution mass spectrometry（HRMS）.The particle size and micromorphology of thenanoparticles were detected by dynamic light scattering（DLS） and transmission electron microscopy（TEM）,respectively.The optical properties of the nanoparticles were characterized by UV-vis absorption spectroscopy and fluorescencespectroscopy.The DOX release kinetics of DOX-Ag NPs under different p H conditions were examined by dialysis methodcombined with fluorescence spectroscopy.The in vitro anti-tumor effects of DOX-Ag NPs were evaluated by MTT assay.Results DOX-Ag NPs were spherical nanoparticles with a particle size of（40.4±3.8） nm.DOX-Ag NPs could rapidlyrelease DOX under weak acid condition.DOX-Ag NPs significantly inhibited the proliferation and cell viability of Hep G2 cells in concentration dependent manner.When DOX concentration was 0.5-20 mg/L（Ag concentration was 0.45-18 mg/L）,the cell survival rate was significantly lower in DOX-Ag NPs group than that of DOX group and Ag NPs group（P〈0.05）.Conclusion DOX-Ag NPs are a combined anti-tumor nano-drug with p H-responsive ability,which can release DOXrapidly in tumor tissues and play an anti-tumor effect through synergistic treatment with Ag NPs in vitro.