沉淀聚合制备粒径可控的温度响应型蛋白纳米胶囊

以牛血清蛋白(BSA)为模型蛋白,N-异丙基丙烯酰胺(NIPAM)为单体,N,N'-亚甲基双丙烯酰胺(BIS)为交联剂,采用原位沉淀聚合法制备了单分散的温度响应型蛋白纳米胶囊(nBSA).通过调整单体与蛋白的比例制备了粒径大小不同的含有单个蛋白分子的BSA纳米胶囊.采用基质辅助激光解吸电离飞行时间质谱仪(MALDI-TOF MS)、透射电镜(TEM)和动态光散射仪(DLS)等对BSA蛋白的修饰度,nBSA的形貌和结构,以及温度响应性能进行了表征,并用HeLa细胞对nBSA的体外安全性进行了初步评价.结果表明,在一定范围内,随着单体和蛋白比例的升高,蛋白纳米胶囊的粒径也逐渐增大,且在7.4～17 nm之间可控,而nBSA的响应温度则逐渐减小在33～41℃之间可控;制备的nBSA单分散性较好;nBSA具有温度响应性能,当环境温度高于其响应温度时,nBSA的粒径可显著增大16～33倍,且这种变化随温度呈现可逆性,并通过对nBSA细胞毒性的初步考察,评价将其用于生物领域的潜力.

FABRICATION OF THERMOSENSITIVE PROTEIN NANOCAPSULES via PRECIPITATION POLYMERIZATION

Thermoresponsive protein nanocapsules were prepared via in situ precipitation polymerization using protein bovine serum album(BSA) as the model protein,N-isopropyl acrylamide(NIPAM) as the monomer,and N,N′-methylene bis-acrylamide(BIS) as the crosslinker.To synthesize such nanocapsules,BSA was first attached with polymerizable groups through reacting with N-acryloxysuccinimide(NAS),and the degree of reaction was examined using MALDI-TOF mass spectrometer.By adjusting the NIPAM/BSA ratio,BSA nanocapsules(nBSA) with different sizes(7.4 nm to 17 nm) were synthesized successfully and confirmed by dynamic light scattering(DLS) and transmission electron microscopy(TEM).Thermoresponsive behavior of nBSA was studied by monitoring the size-temperature trending test through DLS.Reversible thermoresponsive property of obtained nBSA was observed from DLS test,with responsive temperatures ranging from 33℃ to 41℃ accordingly of different sized protein nanocapsules synthesized from various NIPAM/BSA ratios.There is an obvious particle size increase of all synthesized BSA nanocapsules after their exposure to higher temperatures above their responsive ones.The observed particle sizes of nBSA at higher temperatures were increased by 16 to 33 times accordingly.By observing the HeLa cell viability after incubation with the nanocapsules at different concentrations for 24 h,the result shows the nanocapsules exhibit low cytotoxicity.This work provides a new synthesis method of thermoresponsive protein nanocapsules for various applications.