丁香酚纳米脂质体的制备条件优化、表征及其体外抑菌效果  

Preparation conditions optimization and characterization of eugenol nanoliposomes and its antibacterial effect in vitro

作　　者：许书雅 詹轶雯 魏思敏 田启明 伯若楠 刘明江[1,2] 李金贵 XU Shuya;ZHAN Yiwen;WEI Simin;TIAN Qiming;BO Ruonan;LIU Mingjiang;LI Jingui(College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou 225009,China)

机构地区：[1]扬州大学兽医学院,江苏扬州225009 [2]江苏省高校动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《畜牧与兽医》2023年第7期108-117,共10页Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(32072911);江苏省高校优势学科建设工程项目。

摘　　要：为解决丁香酚易挥发、难溶于水和不稳定等问题,将其制备成丁香酚纳米脂质体(ENL),并检测其体外抑菌效果。采用薄膜水合超声法制备ENL,在单因素分析基础上以BBD响应面法优化制备工艺;对其形态、粒径、多分散指数(PDI)、体外释放等进行表征;以微量肉汤稀释法检测ENL对大肠杆菌的抑菌效果。ENL的最佳制备条件为:大豆卵磷脂与胆固醇的质量比5.4∶1、丁香酚浓度1.5%、超声时间10 min,包封率可达到(72.42±0.22)%。平均粒径大小为(203.97±8.66)nm, PDI为0.23±0.03,Zeta电位为(-2.18±0.18)mV,具有较好的分散性。透射电镜下,ENL形态近似球形;ENL中丁香酚的红外光谱特征峰消失,提示丁香酚被成功包裹到脂质体中;24 h内在胃肠模拟液中的累积释放量约为60%。30 d内在4℃和25℃两种储存情况下,ENL平均粒径大小及PDI变化不大,与丁香酚相比,ENL具有更好的稳定性。体外抑菌试验结果表明:ENL的抑菌效果比丁香酚增强2~4倍,其对大肠杆菌的最小抑菌浓度(MIC)为0.31 mg/mL,该浓度可明显抑制大肠杆菌从生长期到对数期的进程;4 MIC处理4 min可杀灭参考株ATCC25922,10 min可杀灭临床分离耐药株AHM5C23;扫描电镜发现,ENL处理引起菌体发生皱缩和细胞膜破损。优化制备的ENL具有较好的均一性和分散性,方便使用,在胃肠模拟液中缓慢释放,且增强了丁香酚的抑菌效果。In order to solve its problems of being volatile,insoluble in water and unstable,eugenol was prepared into nanoliposomes(ENL),and its antibacterial effect in vitro was tested.ENL was prepared using the thin film hydration ultrasonic method,and the prepara-tion process was optimized using the BBD response surface method based on single factor analysis.Its morphology,particle size,polydispersi-ty index(PDI),in vitro release and other characteristics were characterized;and the micro-broth dilution method was used to detect the an-tibacterial effect of ENL on Escherichia coli(E.coli).The optimal preparation conditions of ENL were as follows:The mass ratio of soybean lecithin to cholesterol was 5.4:1,the concentration of eugenol was 1.5%,and the ultrasonic time was 10 min.The encapsulation efficiency could reach(72.42±0.22)%.The average particle size was(203.97±8.66)nm,the PDI was 0.23±0.03,and the zeta potential was(-2.18±0.18)mV,which had a good dispersion.Under the transmission electron microscope,ENL was almost spherical.The characteristic peak of eugenol in infrared spectrum disappeared,indicating that eugenol was successfully encapsulated into liposomes.The cumulative re-lease in gastrointestinal fluid within 24 h was about 60%.Within 30 days,the average particle size and PDI of ENL did not change much un-der the two storage conditions of 4℃ and 25℃.Compared with eugenol,ENL had better stability.The results of in vitro antibacterial test showed that ENL enhanced the antibacterial effect of eugenol by 2-4 times and its minimum inhibitory concentration(MIC)against E.coli strains was O.31 mg/mL,which significantly inhibited the progression of E.coli from the growth phase to the log phase.4 MIC treatment for 4 min could kill the standard strain ATCC25922;and for 10 min,could kill the clinically isolated drug-resistant strain E.coli AHM5C23.Under a scanning electron microscope,ENL treatment was found to have caused shrinkage and cell membrane damage.The optimized ENL had good uniformity and dispersion and was

关 键 词：丁香酚纳米脂质体 BBD响应面法 优化 表征 体外抑菌 

分 类 号：S859[农业科学—临床兽医学]