负载表没食子儿茶素没食子酸酯硅酸钙微球的制备及抗菌性能评价

Preparation of calcium silicate microspheres loaded with epigallocatechin gallate and investigation on its antibacterial performance

背景:茶多酚的主要活性成分为表没食子儿茶素没食子酸酯,其具有抗氧化、抗菌、抗凋亡、抗炎等多种生物学功能。为实现表没食子儿茶素没食子酸酯的可控释放、提高其生物利用度,开发具有生物活性的茶多酚载体是亟待解决的难题。目的:制备高效负载表没食子儿茶素没食子酸酯的微球,同时加测其抗菌性能及生物相容性。方法:采用化学沉淀法分别制备硅酸钙微球与二氧化硅微球,利用X射线光电子能谱、透射电镜、场发射扫描电子显微镜、比表面积分析仪、激光粒度分析仪对微球的性能进行表征。将两种微球分别浸泡于表没食子儿茶素没食子酸酯溶液中,制备负载表没食子儿茶素没食子酸酯的硅酸钙微球与二氧化硅微球(分别记为CSM-EGCG、SM-EGCG),检测微球的载药量及包封率,以及体外表没食子儿茶素没食子酸酯的释放情况。将未载药的硅酸钙微球、二氧化硅微球及CSM-EGCG、SM-EGCG分别与金黄色葡萄球菌、大肠杆菌共培养,检测抑菌率。将上述4种微球分别与人皮肤成纤维细胞共培养,通过CCK-8法评估细胞活性。结果与结论:①表征实验结果显示,硅酸钙微球和二氧化硅微球均为介孔微球,分散性好,硅酸钙微球的比表面积、总孔体积、平均孔径均大于二氧化硅微球;②CSM-EGCG的包封率为(72.0±0.5)%,载药量为(58.4±0.4)%;SM-EGCG的包封率为(41.6±0.7)%,载药量为(45.0±1.3)%;CSM-EGCG微球较SM-EGCG微球具有更好的药物携载能力,且体外释药时间可持续19 d以上,累计释放量达到(88.1±3.0)%;③二氧化硅微球无抗菌能力,硅酸钙微球、SM-EGCG、CSM-EGCG对金黄色葡萄球菌的抑菌率分别为(28.0±4.2)%,(63.9±1.0)%,(95.6±0.5)%,对大肠杆菌的抑菌率分别为(27.5±7.0)%,(51.9±1.4)%,(93.4±1.0)%;④CCK-8检测显示,硅酸钙微球及CSM-EGCG具有良好的细胞相容性,具有促进人皮肤成纤维细胞增殖的能力;⑤结果表明,负载表没食子儿茶素没食子酸酯的硅酸钙微球具有良好的抗菌性及细胞相容性。

BACKGROUND:The main active component of tea polyphenols is epigallocatechin gallate,which has various biological functions such as antioxidation,antibacteria,anti-apoptosis,and anti-inflammation.To realize the controllable release of tea polyphenols and improve their bioavailability,it is urgent to develop bioactive carriers for tea polyphenols.OBJECTIVE:To prepare microspheres loaded with high-efficiency epigallocatechin gallate,and measure its antibacterial performance and biocompatibility.METHODS:Calcium silicate microspheres were prepared by chemical precipitation method.X-ray photoelectron spectroscopy,transmission electron microscopy,field emission scanning electron microscopy,surface area measurement and laser particle size analyzer were used to characterize the performance of the microspheres.The two kinds of microspheres were immersed in epigallocatechin gallate solution respectively to prepare calcium silicate microspheres and silica microspheres loaded with epigallocatechin gallate(named as CSM-EGCG and SM-EGCG,respectively).The drug loading rate and encapsulation rate of the microspheres,as well as the release of epigallocatechin gallate in vitro were detected.The unloaded calcium silicate microspheres,silica microspheres,CSM-EGCG and SM-EGCG were co-cultured with Staphylococcus aureus and Escherichia coli,respectively,to detect the antibacterial capacity.The above four kinds of microspheres were co-cultured with human skin fibroblasts respectively,and the cell viability was evaluated by CCK-8 assay.RESULTS AND CONCLUSION:(1)The characterization experiment results showed that the prepared calcium silicate spheres and silica spheres were mesoporous microspheres,and the specific surface area,total pore volume and average pore size of calcium silicate spheres were larger than those of silica spheres.(2)The encapsulation rate and drug loading rate of CSM-EGCG were(72.0±0.5)%and(58.4±0.4)%,respectively.The encapsulation rate and drug loading rate of SMEGCG were(41.6±0.7)%and(45.0±1.3)%,respectively.CSM-EGCG microspheres had better drug loading capacity than SM-EGCG microspheres,and the drug release time in vitro lasted for more than 19 days,and the cumulative release reached(88.1±3.0)%.(3)Silica microspheres had no antibacterial activity.The antibacterial rates of calcium silicate microspheres,SM-EGCG,and CSM-EGCG against Staphylococcus aureus were(28.0±4.2)%,(63.9±1.0)%,and(95.6±0.5)%,respectively.The antibacterial rates against Escherichia coli were(27.5±7.0)%,(51.9±1.4)%,and(93.4±1.0)%,respectively.(4)CCK-8 assay results showed that the calcium silicate spheres and CSM-EGCG had good cell biocompatibility and showed enhanced proliferation of human skin fibroblasts.(5)These results verified that the epigallocatechin gallate-loaded calcium silicate spheres have good antibacterial activity and cytocompatibility.