Abstract

Ou-gon, an extract from Scutellaria baicalensis Georgi root, has been shown to exhibit pronounced antifungal activity. The present study aimed to identify antifungal components of Ou-gon and to determine their mechanism of action against pathogenic fungi. Antifungal activity was assessed by the microbroth dilution method using four common human pathogenic fungi, Trichophyton rubrum, Trichophyton mentagrophytes, Aspergillus fumigatus, and Candida albicans. Components of crude Ou-gon extract were separated by reversed-phase high-performance liquid chromatography. Active antifungal components were identified by liquid chromatography-electrospray ionization tandem mass spectrometry. Terminal deoxynucleotidyl transferase dUTP nick end-labelling assay, SYTOX® green uptake assay, determination of intracellular reactive oxygen species and mitochondrial membrane potential as well as microscopy (confocal laser microscopy, scanning and transmission electron microscopy) were used to probe the mode of action. Two components with potent antifungal activity, baicalein and wogonin, were identified in Ou-gon. Baicalein showed potent antifungal activity against the four fungi tested. Wogonin displayed antifungal activity against all four fungi except C. albicans. The components are considered to induce apoptosis-like programmed cell death via hyperproduction of reactive oxygen species. This study enhances our understanding of the antifungal activity of Kampo medicine, and may contribute to the development of new and safe antifungal therapeutics.

Highlights

  • Because of its most pronounced relative antifungal activity, the acetic acid extract was selected for reversed-phase high-performance liquid chromatography (RP-HPLC) analysis

  • The extract was separated by RP-HPLC on a C18 column, and the antifungal activity of the eluted fractions was tested

  • transferase dUTP nick end-labelling (TUNEL) staining of baicalein- and wogonin-treated fungal cells suggested the presence of apoptotic DNA breaks

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Summary

Introduction

Kampo is a well-known type of traditional Japanese medicine that is widely used to relieve symptoms of a variety of diseases because of the low incidence of adverse effects associated with its use. It is currently prescribed by over 80% of medical doctors in Japan[7]. We previously evaluated the antifungal activity of 61 commercially available Kampo medicines towards Trichophyton rubrum using the microbroth dilution assay[7] Seven of these medicines exhibited antifungal activity, with six containing Ou-gon, an extract from the roots of Scutellaria baicalensis Georgi. The current study aimed to identify antifungal components in Ou-gon and to determine their mechanism of action against pathogenic fungi

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