聚(ε-己内酯)-多肽共聚物胶束增强抗生素的抗菌活性

Poly(ε-caprolactone)-Polypeptide Copolymer Micelles Enhance the Antibacterial Activities of Antibiotics

细菌感染对人类健康构成了严重威胁,而传统抗生素治疗可能导致如胃病、细菌耐药等一系列副作用。因此,亟需发展提高抗生素抗菌效率的新方法。在本文中,我们通过氨基酸环内酸酐和ε-己内酯开环聚合合成了多肽基的聚合物[PCL_(34)-b-PGA_(30)-b-P(Lys_(16)-stat-Phe_(12))]。在碱性环境中,此聚合物能够自组装成带负电荷的胶束(其zeta电位值为−26.7 mV)。通过透射电子显微镜和动态光散射可以证实组装体的胶束结构。此胶束具有药物缓释的特性,并且胶束上含有的抗菌多肽链段能够有效杀死细菌,因此该胶束能够很大程度上提高抗生素的抗菌效率。实验证实该载药胶束对革兰氏阴性菌(大肠杆菌)和革兰氏阳性菌(金黄色葡萄球菌)均具有较好的抗菌活性,最小抑菌浓度值分别为7.8和18.2μg·mL^(−1)。这种胶束包载妥布霉素的载药率和载药量分别为24.3%和5.2%。因此,胶束中实际含有的抗生素量为0.4和0.9μg·mL^(−1),小于妥布霉素的最小抑菌浓度值。总的来说,该胶束有望大幅降低抗生素的使用量,并减少临床上抗生素过量使用造成的副作用。

Bacterial infection is a major threat to human health,and can cause several diseases including gastroenteritis,influenza,tetanus,and tuberculosis.As conventional antibiotic treatment may cause various undesirable effects such as stomach disorder and bacterial resistance,it is necessary to improve the antibacterial efficiency of antibiotics.Here,we synthesized a peptide-based copolymer,poly(ε-caprolactone)-blockpoly(glutamic acid)-block-poly(lysine-stat-phenylalanine)[PCL_(34)-b-PGA_(30)-b-P(Lys_(16)-stat-Phe_(12))]by ring-opening polymerization(ROP)ofε-caprolactone and amino acid N-carboxyanhydride(NCA).Successful synthesis of the copolymer was verified by proton nuclear magnetic resonance and size exclusion chromatography.This copolymer can self-assemble into negatively charged micelles(−26.7 mV)under alkaline conditions by solvent switch method.The micelle structure was confirmed by transmission electron microscopy and dynamic light scattering,and revealed to have a diameter of~42 nm.Antibiotics were loaded into micelles during the self-assembly process,and cell viability assay was conducted to evaluate its cytotoxicity with and without tobramycin.No obvious cytotoxicity was observed for both micelles when the concentration was lower than 300μg·mL^(−1).The antibiotic-loaded micelles demonstrated very low minimum inhibitory concentrations(MICs)against both Gram-negative Escherichia coli(E.coli)(7.8μg·mL^(−1))and Gram-positive Staphylococcus aureus(S.aureus)(18.2μg·mL^(−1)),while the MICs of free tobramycin were 3.9 and 1.0μg·mL^(−1),respectively.The drug-loading content and efficiency of the micelles were 5.2%and 24.3%,respectively.Therefore,the MICs of the loaded tobramycin against E.coli and S.aureus were 0.4 and 0.9μg·mL^(−1),respectively,suggesting that the micelle could enhance the antibacterial activity of antibiotics.Tobramycin-loaded micelles demonstrated a sustained release characteristic,with 85%of the antibiotics released after 8 h.In bacteria-induced acidic microenvironment,the coil conformation of PGA blocks transforms and PGA blocks shrink toward the micelle core.Concomitantly,the carboxyl side chains are protonated in an acidic environment,increasing the hydrophobicity of this micelle.Antibiotics will be captured when reaching the outer core to slow down the releasing process.Furthermore,the poly(lysine-stat-phenylalanine)[P(Lys-stat-Phe)]coronas with broad spectrum intrinsic antibacterial activity can penetrate the bacterial cell membrane,leading to leakage of the cellular contents of the bacteria and ultimately their death.Due to the sustained release property of micelle and the intrinsic activity of the antibacterial peptide segments,this micelle can greatly enhance the antibacterial activity of antibiotics.Overall,this antibiotic-loaded micelle provides a novel approach for significantly reducing the antibiotics dosage and avoiding the associated health risks.