超小的羧甲基壳聚糖超顺磁氧化铁纳米粒制备及处方优化

The Synthesis and Characterizations of O-Carboxymethyl-Chitosan Ultra-small Superparamagnetic Iron Oxide Nanoparticles

关键词:  羧甲基壳聚糖  超小超顺磁氧化铁纳米粒  正交实验  MRI造影剂

Key words:  O-carboxymethyl-chitosan  ultra-small superparamagnetic iron oxide nanoparticle  orthogonal experimental design  MRI contrast agent

摘要:

目的 优化羧甲壳聚糖超小超顺磁氧化铁纳米粒(O-carboxymethyl-chitosan ultra-small superparamagnetic iron oxide nanoparticle，OCMCS-USPIO-NPs)处方并对其理化性质进行表征。方法 共沉淀法合成超顺磁氧化铁纳米粒核并用羧甲基壳聚糖对其表面进行共价修饰，用正交实验L₉(3^₄)对OCMCS-USPIO-NPs的处方优化和工艺进行优化筛选；用透射电镜、电子相关光谱仪(LSD)、X-Ray衍射法、傅立叶红外、磁性测定仪、MRI扫描仪和邻二氮菲法测定铁含量等对制备的OCMCS-USPIO-NPs进行表征；用普鲁士蓝染色法体外评价OCMCS-USPIO-NPs抗吞噬能力。结果 正交实验处方优化处方为：羧甲基壳聚糖的分子量为1~2万，浓度为3%，超声时间为45 min，功率为600 W；傅立叶红外和X-Rays结果证实了OCMCS-SPIO-NPs的合成，羧甲基壳聚糖的修饰显著降低超顺磁氧化铁的流体粒径及超顺磁性，修饰后饱和磁性为73.4 emu·g^-1 Fe，磁性较强；同时增加纳米粒表面的Zeta电位，普鲁士蓝染色结果表明巨噬细胞吞噬的纳米粒量依次是：uncoated SPIO-NPs＞dextran-SPIO-NPs＞OCMCS-SPIO-NPs。结论 合成的OCMCS-USPIO-NPs的流体粒径<50 nm，具有强的超顺磁性，分散性好，能逃避巨噬细胞的摄取，可用于MRI照影。

Abstract:

OBJECTIVE To develop a novel USPIO-NPs system by surface-modifying superparamagnetic iron oxide nanoparticles (USPIO-NPs) with O-carboxymethyl-chitosans (OCMCS) to improve their biocompatibility and ability to evade reticuloendothelial system(RES). METHODS The OCMCS-USPIO-NP was synthesized by two-step methods: at first, the plain SPIO-NPs was synthesize by alkaline co-precipitation, then the surface of the SPIO-NPs were modified by conjugating the OCMCS to USPIO-NPs, orthogonal experimental design of L9(34) was used to optimize the technology of preparing OCMCS-USPIO-NPs. The resulting USPIO nanoparticles were characterized by dynamic light scattering (DLS), fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscope (TEM), zeta-potential measurement and vibrating sample magnetometry (VSM) and MRI. The internalization of uncoated SPIO-NPs, dextran-SPIO-NPs and OCMCS-USPIO-NPs was visualized via prussian blue staining to evaluate the ability of the different SPIO-NPs to escape the macrophage capture. RESULTS The orthogonal experimental design demonstrated that the best technology was as follows: carboxymethyl chitosan molecular weight was 10-20 K, the concentration of OCMCS was 3%, ultrasonic time was 45 min, ultrasonic power was 600 W. FTIR and XRD results confirmed that the successful synthesis of OCMCS-SPIO-NPs. VSM results demonstrated both the modified magnetic Fe₃O₄ nanoparticles and the uncoated SPIO-NPs were superparamagnetic. The results of characterization also demonstrated that the modification with OCMCS results in a drop in hydrodynamic size, saturated hysteresis (r) while lead to a increase in zeta potential. The prussian blue staining in vitro results indicated the internalization of OCMCS-USPIO-NPs into macrophage was much lower than that of dextran-SPIO-NPs and further lower than SPIO-NPs. CONCLUSION The synthesized OCMCS-USPIO-NPs system is stable with hydrodynamic size less than 50 nm, strong electrostatic repulsion, strong super-paramagnetic and low macrophage uptake. Thus, OCMCS-USPIO-NPs system can be a desirable alternative for MRI contrast agent.