该实验选用大肠杆菌和李斯特菌为G-、G+菌的代表菌，探究月桂酰精氨酸乙酯（LAE）对其以细菌细胞膜为作用靶点的抑菌机理，通过测定细菌的抑菌曲线、中和内毒素的活性、细菌表面特性、构建脂质体模拟LAE与磷脂双分子层的相互作用、离子的泄漏和LAE对G-菌外膜及细胞质膜的渗透性等探究其抑菌机理。实验结果表明，LAE对G-、G+菌均有明显的杀菌活性，最小抑菌浓度均为8 μg/mL，LAE结合脂多糖对内毒素的中和率可高达96.56%，降低细胞表面Zeta电位，增强细菌表面疏水性，且对G-菌的影响更大。LAE能引起脂质体包裹的荧光素钙黄绿素的泄漏，并呈浓度依赖性，但LAE并不能使脂质体膜完全破裂。LAE能够增加G-菌外膜渗透性，使大肠杆菌对抗生素探针利福平和红霉素更敏感，同时对细胞质膜产生较大扰动，使内容物从胞内渗出，从而抑制细菌的生长。该研究结果表明LAE主要以改变细胞壁膜渗透性，导致胞内物质的泄漏，而达到抑菌作用。

In this experiment, Escherichia coli and Listeria innocua were selected as the representative G- and G+ bacteria to explore the antibacterial mechanism of LAE towards bacterial membranes. The antibacterial mechanism was examined via investigating LAE’s antibacterial curve, the ability of neutralizing endotoxin, and bacterial surface characteristics, and constructing liposomes to simulate the interaction between LAE and phospholipid bilayers and evaluate the ion leakage and the permeability of LAE through G- bacterial outer membrane and cytoplasmic membrane. The experimental results showed that LAE had significant bactericidal activities against both G- and G+ bacteria with the minimum inhibitory concentration as 8 μg/mL. LAE could neutralize up to 96.56%endotoxin through binding to lipopolysaccharide, reduce the zeta potential on the cell surface, enhance hydrophobicity of bacterial surface, and exert a greater impact on G- bacteria. LAE could cause the leakage of a liposome-encapsulated fluorescein, calcein, in a concentration-dependent manner, but LAE didn’t collapse completely the liposome membrane. LAE was able to increase the permeability of the outer membrane of G- bacteria, which made E. coli more sensitive to the antibiotic probes, rifampicin and erythromycin, while disturbed greatly the cytoplasmic membrane, causing the leakage of intracellular contents, thereby inhibiting the growth of bacteria. The results of this study indicate that LAE exerts bacteriostatic effects mainly through changing the cell wall membrane permeability and consequently leading to the leakage of intracellular substances.

中央高校基本科研业务费专项（2019NYB26）