Antifungal mode of action of macrocarpal C extracted from Eucalyptus globulus Labill ( Lan An ) towards the dermatophyte Trichophyton mentagrophytes

摘要

The fresh leaves of Eucalyptus globulus Labill. (Lan An) have been used in Chinese medicine for many years to treat dermatomycosis. Macrocarpal C was isolated from this herb and identified as its major antifungal component by bioassay-guided purification. This study aims to investigate the antifungal activity of macrocarpal C against Trichophyton mentagrophytes, which can cause tinea pedis. Fresh leaves of E. globulus were extracted with 95?% ethanol, and the resulting ethanolic extracts were dried before being partitioned with n-hexane. The n-hexane layer was then subjected to chromatographic purification to give macrocarpal C. The antifungal minimum inhibitory concentration (MIC) of macrocarpal C was determined using the standard M38-A2 method described by the Clinical Laboratory Standards Institute (CLSI). The mode of action of macrocarpal C was elucidated using three in vitro assays, including (1) a fungal membrane permeability test using SYTOX? Green; (2) a reactive oxygen species (ROS) production test using 5-(and-6)-carboxy-2′,7′-dihydrodichlorofluorescein diacetate as a cell-permeable fluorogenic probe; and (3) a DNA fragmentation test based on terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) detection. Terbinafine hydrochloride and nystatin were used as positive controls. The suppression in the growth of T. mentagrophytes following its treatment with macrocarpal C was associated with an increase in the permeability of the fungal membrane (P?=?0.0043 when compared to control); an increase in the production of intracellular ROS (P?=?0.0063); and the induction of apoptosis as a consequence of DNA fragmentation (P?=?0.0007). This study demonstrated that the antifungal action of macrocarpal C was associated with increases of membrane permeability, intracellular ROS and DNA fragmentation.