Anti-pathogenic yeast malassezia globosa activity by propolis of stingless bee geniotrigona thoracica

Abstract

Malassezia globosa is a lipophilic pathogen which relates to various chronic skin diseases. Azole drugs are main available treatments, but they have been reported with many side effects and microbial drug resistance. Therefore, the alternative medications should be developed. As the antimicrobial activity of propolis is outstanding, this study aims to investigate the potential of propolis from stingless bee, Geniotrigona thoracica against the pathogenic yeast. The extraction and characterization of the propolis components were done after the obtained extracts were evaluated for anti-M. globosa growth by using agar well diffusion assay and broth microdilution assay. Moreover, Inhibitory Concentration values at 50% (IC50) was estimated by using nonlinear regression analysis. The sample was firstly extracted with 80% methanol (MeOH), then was partitioned with different polarities of organic solvents. The first obtained crude extracts were crude 80% MeOH extract (CME) which exhibited zone of inhibition beginning at 200 mg/ml (11.83 ± 0.50 mm), and its IC50 was estimated at 2.21 mg/ml. After CME was partitioned, only crude MeOH partitioned extract (CMPE) showed the activity. The zone of inhibition from CMPE was observed beginning at 12.5 mg/ml (9.00 ± 0.00 mm) and IC50 values was estimated 1.22 mg/ml. Hence, CMPE was selected to further purification by column chromatography. The obtained CMPE 1 presented the best activity with zone of inhibition beginning at 12.5 mg/ml (11.83 ± 0.83 mm) and IC50 was predicted at 185.0 μg/ml. CMPE 1 was then identified as methyl gallate (MG) by using nuclear magnetic resonance (NMR). Accordingly, our synthetic MG and standard MG were subsequently tested, which the IC50 values were 223.7 μg/ml and 59.14 μg/ml, respectively. The Minimal Fungicidal Concentration of MG was 8 mg/ml as standard MG was used as a representative due to the limitation of the extract. Since extracellular lipase is necessary for survival and pathogenicity, the efficiency of MG toward the lipase was additionally investigated. Both colorimetric and agar-based methods were examined. The results suggested that activity of lipase and growth inhibition of MG might be not related, which percentage of lipase inhibition of standard MG (30.25 ± 1.91%) was lower than the growth inhibition (57.35 ± 0.59%) at the same concentration. Either it did not trend to increase in higher concentrations. However, it still supported the reduction of pathogenicity at low concentration. This is the first report that proves the activity against both growth and lipase of M. globosa by MG, which is explored from the natural resource. Thus, propolis from G. thoracica could be a good candidate source for searching new anti-Malassezia or other antifungal agents.