分子印迹空心球在蛋白质固相萃取中的应用

建立了分子印迹固相萃取-高效液相色谱相结合分离复杂样品中牛血红蛋白(Hb)的方法. 以单分散的二氧化硅球为载体, Hb为模板, 采用表面印迹法制备了分子印迹空心球(MIHS). 采用透射电子显微镜(TEM)对该印迹材料的形貌进行了表征. 结果表明, 所制备的MIHS的单分散性好、 粒径均匀, 是一种理想的固相萃取填料. 以该印迹材料作为固相萃取吸附剂, 并结合高效液相色谱(HPLC)研究了其对Hb的吸附行为, 结果表明, 制备的MIHS在磷酸盐缓冲液(pH=7.0)中对Hb的回收率高于80%, 而对核糖核酸酶A(RNase A)和细胞色素C(Cyt C)的回收率低于50%, 方法的精密度高(RSD<5%). 与Hb印迹硅球相比, MIHS具有较快的传质速率和萃取效率. 将MHIS应用于复杂样品中Hb的分离, 结果表明, MIHS可以从加标牛血清中选择性吸附Hb, 吸附率达82%.

Molecularly Imprinted Hollow Spheres for the Solid Phase Extraction of Protein†

Molecularly imprinted polymer was widely used as the stuffing of solid phase extraction(SPE) due to its high selectivity. Hb isolation from complex matrix was established based on molecularly imprinted solid phase extraction(MISPE). Mono-dispersed molecularly imprinted hollow spheres(MIHS) of hemoglobin(Hb) were synthesized using silica nanospheres as the sacrificial matrix. It is proved that the MIHS with good monodispersion and uniform particle size by TEM scanning was suitable for SPE enrichment matrix. Therefore, the extraction of Hb, bovine serum albumin(BSA) and cytochrome C(Cyt C) by MIHSs was investigated by SPE combined with liquid chromatographic method. The recovery of Hb was higher than 80% in phosphate buffer solution(pH=7) while it was lower than 50% for RNase A and Cyt C. Meanwhile, the degree of precision(RSD) was lower than 5%. Compared to the surface imprinted silica(SIS), the MIHS had good recognition and fast adsorption-desorption dynamics for Hb. Moreover, this method was also confirmed in real sample, and the adsorption of Hb was 82% in the spiked bovine serum.