IN VITRO AND IN SITU ANTIFUNGAL ACTIVITY OF COMBINATION OF ROSEMARY LEAVES, JAVA TURMERIC RHIZOMES, AND GINGER RHIZOMES

• Main Author: Susanto, Eriwan
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/45964
• Institution: Institut Teknologi Bandung

Indonesia is a tropical region with high temperatures and humidity that will facilitate the growth of mold and yeast, so that fungus infection commonly found in this country. Dermatophytosis and candidiasis is a skin infection caused by fungus with a high prevalence among other skin diseases in Indonesia. However, effective antifungal is still very limited and not all have a good safety profile. Therefore, the search for new compounds that have antifungal activity and better safety profile is still being conducted. The aim of this research is to test the antifungal activity of some combinations from ten plants against Candida albicans, Microsporum gypseum, and Trichophyton mentagrophytes. The powdered of crude drug was extracted by reflux method using ethanol 96% as solvent. Antifungal activity of extracts was tested by microdilution and agar diffusion method against Candida albicans, Microsporum gypseum, and Trichophyton mentagrophytes to determine the Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC). The extracts which showed antifungal activity, then were examined its combination properties. Three plant extracts with the highest activity and had synergistic interaction in combination were selected. Then, the three extracts were fractionated by liquid-liquid extraction method using n-hexane and ethyl acetate. MIC and MFC values of fractions of each extracts were determined. The three selected extracts were in ointment and cream preparations to be tested against Trichophyton mentagrophytes in infected rabbit’s skin. The selected extracts and fractions were then compared with ketoconazole as reference drug. Active fractions were investigated its activity through TLC bioautography method. Three fractions with the highest activity from selected plant extracts were observed its work site by Scanning Electron Microscopy (SEM). Characterization of crude drug and extracts were conducted in plants with the highest activity, included phytochemical screening, total ash content, and loss on drying for crude drug and phytochemical screening, water content, loss on drying, water and ethanol extractable matter, density, yield of extraction, and chromatogram pattern for the extract. The ethanol extract of rosemary leaves, Java turmeric rhizome, and ginger rhizome showed the highest activity and synergistic interaction in combination against Trichophyton mentagrophytes. Cream o/w which contained combination of three plant extracts showed activity by decreasing edema and erythema total scores was significantly different (p<0,01) with positive control on day 8 after treatment. The preparation of cream o/w showed average healing effect after 16.67 days, which faster than the positive control 24 days. Java turmeric extract and ginger nhexane fraction showed the highest activity towards ketoconazole than other extracts and fractions. The equivalency of antifungal activity of Java turmeric extract to ketoconazole showed that 1 mg extract had equivalency activity to 0.00701 mg of ketoconazole, while the equivalency of antifungal activity of ginger n-hexane fraction to ketoconazole showed that 1 mg n-hexane fraction had equivalency activity to 0.01489 mg ketoconazole in testing against Trichophyton mentagrophytes. The results of TLC bioautography showed three components of ethyl acetate fraction of rosemary could inhibit the growth of Trichophyton mentagrophytes. Five components in ethyl acetate fraction of ginger had antifungal activity, while only one component in Java turmeric ethyl acetate fraction had antifungal activity. In the n-hexane fraction of Java turmeric, six components showed iv growth inhibitory activity against Trichophyton mentagrophytes, while four components in ginger n-hexane fraction had antifungal activity. SEM results showed that three fractions from selected plant extracts induced morphological change of Trichophyton mentagrophytes cells. This might be due to the inhibition of cell wall synthesis and disruption of cell membrane. The ethanol extracts of rosemary leaves, Java turmeric rhizome, and ginger rhizome showed the highest antifungal activity with synergistic interaction in combination. Cream o/w that contained combination of three plant extracts showed antifungal activity better than positive control in situ. Chromatogram of ethyl acetate fraction of rosemary, n-hexane fractions of ginger and Java turmeric showed some spots with certain Rf values that specifically showed antifungal activity. SEM results showed the occurrence of morphological changes induced by exposure to each fraction with the highest activity from selected plant extracts.