目的 探究诃子水提物（water extract of Terminalia chebula，TRW）对球形马拉色菌的体外抗菌活性及对生物被膜的抑制作用，并初步探讨其抗菌机制。方法 采用微量稀释法测定TRW对马拉色菌的体外抗菌活性；活菌计数法测定TRW对球形马拉色菌作用的时间-杀菌曲线。通过微板法构建球形马拉色菌生物被膜体外模型，用XTT法测定生物被膜代谢活性；采用活菌计数法和扫描电子显微镜分别检测TRW对球形马拉色菌生物被膜活菌量以及微观形态的影响。采用山梨糖醇保护实验、碘化丙啶（PI）染色分别测定TRW对球形马拉色菌细胞壁和细胞膜的作用影响。结果 TRW对7株马拉色菌的最低抑菌质量浓度为0.90～4.59 mg/mL，其中对TRW敏感性最强的是球形马拉色菌MC14。时间-杀菌曲线结果显示TRW对MC14整个生长周期均有抑制作用。球形马拉色菌MC14生物被膜的初黏附期、聚集期和成熟期分别为0～24 h、24～96 h和96～168 h。TRW可降低MC14各个阶段生物被膜的活菌量，作用效果由强到弱排列为聚集期＞黏附期＞成熟期；且对MC14各个阶段的生物被膜均有清除作用，可减少MC14生物被膜中的细菌量和胞外基质，破坏生物被膜的结构。TRW可以通过改变细胞壁通透性、破坏细胞膜完整性产生抗菌作用。结论 TRW不仅对糠秕马拉色菌、球形马拉色菌和限制性马拉色菌有较好的体外抗菌效果，而且对球形马拉色菌不同阶段的生物被膜有清除作用，其抗菌机制与影响细胞膜通透性、细胞壁完整性有关，为TRW在新型抗菌药物研究和应用提供了一定的基础理论和依据。

Objective To investigate the antibacterial activity and anti-biofilm effect of water extract of Terminalia chebula (TRW) against Malassezia globosa, and preliminary explored its antibacterial mechanism. Methods Microdilution method was used to determine the antibacterial activity of TRW against Malassezia in vitro; The time-kill curve of TRW against M. globosa was determined by viable count method. An in vitro model of M. globosa biofilm was constructed by microplate method; The metabolic activity of biofilm was measured by XTT method. The effect of TRW on the number of viable bacteria in biofilm of M. globosa and the microscopic morphology of biofilm were detected by means of viable bacteria counting method and scanning electron microscopy. Sorbitol protection test and PI staining were used to determine the effect of TRW on cell wall and membrane of M. globosa. Results The minimum inhibitory concentration of TRW to seven Malassezia strains were ranged from 0.90 to 4.59 mg/mL, and the most sensitive to TRW was M. globosa MC14. The results of time-kill curve showed that TRW inhibited the entire growth cycle of MC14. The initial adhesion period, aggregation period and mature period of MC14 biofilm were 0—24 h, 24—96 h and 96—168 h, respectively. TRW could reduce the number of viable bacteria in biofilm at each stage of MC14, and the effect was as following: aggregation stage > adhesion stage > mature period. TRW could clear the biofilm at all stages of MC14, reduce the number of bacteria and extracellular matrix in MC14 biofilm, and destroy the structure of biofilm. TRW could produce antibacterial effect by changing the permeability of cell wall and destroying the integrity of cell membrane. Conclusion TRW not only has a good antibacterial effect on M. furfur, M. globosa and M. restricta in vitro, but also can remove the biofilm of M. globosa at different stages. Its antibacterial mechanism is related to the influence on cell membrane permeability and cell wall integrity, which provides a certain basic theory and basis for the research and application of TRW in new antibacterial drugs.

R285.5

国家重点研发计划“中医药现代化”重点专项项目（2018YFC1707100）