纳米羟基磷灰石的表面修饰及其与DNA结合的实验研究

Experimental Study on Surface Modification and DNA Binding of Hydroxyapatite Nanoparticles

目的探讨纳米羟基磷灰石(nHAP)的表面修饰及其与DNA结合的可行性。方法采用均相沉淀法制备nHAP;透射电镜观察纳米粒结构;经超声分散及碳酸钠预处理后,在pH值7.4的环境下应用多聚赖氨酸(PLL)修饰nHAP;对修饰前后的纳米粒行Zeta电位检测;采用离心后测上清DNA浓度检测PLL修饰后的nHAP(nHAP-PLL)结合DNA的能力;应用DNA抗核酸酶保护实验测定nHAP-PLL保护DNA的能力。结果透射电镜下nHAP呈针状颗粒,粒径较均匀,约(15～20)nm×(60～80)nm左右,分散程度良好;nHAP的Zeta电位为(-42.4±7.5)mV,nHAP-PLL的Zeta电位为(8.5±1.5)mV,两组比较差异有统计学意义(P<0.01);DNA结合及抗核酸酶保护实验显示nHAP-PLL和DNA质量比达20∶1时,nHAP-PLL可有效结合并保护相应DNA。结论nHAP经PLL表面修饰后可成为一种有效的DNA结合载体。

Objective To investigate the feasibility of surface modification and DNA binding of nano-hydroxyapatite （nHAP）. Methods The nHAP was synthesized by the homogeneous precipitation method. The structure of the nanopartieles was observed under a transmission electron microscope. The nHAP was prepared using ultrasonication and Na2 CO3 and was modified with poly L-lysine （PLL） at pH 7.4. Zeta potential of nHAP and nHAP modified with poly-L-lysine （nHAP-PLL） was detected respectively. The binding capacity of nHAP-PLL to DNA was tested by measuring DNA concentration in superna- rant after centrifugation. The capacity of protecting DNA by nHAP-PLL was assayed by the experiment of DNA degradation. Results Under the transmission electron microscopy, the synthesized product presented needle-like and well dispersed particles with evenly distributed sizes of （15-20） nm× （60-80） nm. The Zeta potential of nHAP was --42.4±7.5 mV, significantly lower than that of nHAP-PLL （8.5±1.5 mV, P〈0.01）. DNA binding test showed that nHAP-PLL could completely bind DNA when the w/w ratio of nHAP PLL and DNA was more than 20 ： 1. The nHAP-PLL/DNA mixtures with w/w ratio of 20 ： 1 upwards could effectively protect DNA against being digested by DNase Ⅰ. Conclusion The nHAP modified with PLL can effectively bind and protect DNA and may be used as a potential DNA carrier.