IN VITRO AND IN SITU ANTIBACTERIAL ACTIVITIES STUDY OF FINGERROOT RHIZOME (Kaempferia pandurata Roxb.) AGAINST Staphylococcus aureus

• Main Author: Kamil, Annisa
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/45957
• Institution: Institut Teknologi Bandung

Staphylococcus aureus is the most common cause of bacterial skin and soft tissue infections in the world. The prevalence of S.aureus skin infection is high in nosocomial infections. Treatment of S.aureus infections increasingly difficult due to the presence of resistant strains, known as Methicillin-resistant Staphylococcus aureus (MRSA). Previous research reported that ethanolic extract of fingerroot rhizome has antibacterial activity against S.aureus. This study will evaluate antibacterial activity of fractions from ethanolic extract of fingerroot rhizome in vitro against S.aureus and MRSA as well as in situ antibacterial activity against skin infection caused by S.aureus in rabbits, determine the compound which act as antibacterial agent in active fraction, and determining antibacterial work site of active fraction against S.aureus. The crude drug of fingerroot rhizome extracted by reflux using ethanol 96% as solvent, then was fractionated by liquid-liquid extraction using n-hexane and ethyl acetate. In vitro antibacterial activity of ethanolic extract, n-hexane, ethyl acetate, and water fraction was tested by microdilution method to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) as well as determination equivalency of antibacterial activity to standard antibiotic (Tetracycline HCl) with agar diffusion method against S.aureus and MRSA. Bioautography was done to the most active fraction to determine antibacterial compounds. Determination of antibacterial work site of active fraction observed by SEM. In situ antibacterial activity was performed by giving the ointment and cream of extract, and oxytetracycline ointment in rabbits that injected by S.aureus. The observed parameters were diameter of erythema, edema thickness, and diameter of pus. Primary and ocular irritation test were conducted to assess the safety of topical preparations. Microdilution test results showed MIC value of ethanolic extract, nhexane, and ethyl acetate fraction against S. aureus were 64, 64, and 512 ppm respectively, whereas for MRSA were 128, 128, and 1024 ppm respectively. The MBC of the ethanolic extract, n-hexane, and ethyl acetate fraction against S. aureus were 256, 64, and 1024 ppm respectively, whereas for MRSA were 8192,128, and 1024 ppm respectively. Bioautography of n-hexane fraction showed two active antibacterial spots for S.aureus. Observation by SEM exposed that cell of S.aureus which was given by n-hexane fraction showed morphological change became not flat and irregular. In situ antibacterial test showed a total score of 3% oxytetracycline ointment was lower and significantly different (p<0.05) with positive control on day 4 of treatment, while the total score of 3% extract cream was lower and significantly different (p<0.05) with positive control on day 6, and significantly different (p<0.05) with 3% oxytetracycline ointment on day 23. Ointment of extract 3% had significantly different (p<0.05) with positive control on day 25. N-hexane fraction showed the best antibacterial activity among ethanolic extract, ethyl acetate, and water fractions. Antibacterial activity equivalency of n-hexane fraction to tetracycline HCl was the highest against MRSA (1 mg fraction ~ 1.79 x 10 -2 mg tetracycline HCl). There was flavonoid iv compound besides panduratin A that have the most powerful role as an antibacterial agent in n-hexane fraction. Based on SEM result, n-hexane fraction was supposed to be worked on cell membrane or protein. Cream of extract 3% gave the best antibacterial activity than 3% oxytetracycline ointment and 3% extract ointment against skin which was infected by S.aureus. Ointment and cream of extract 3% did not irritate the skin and eyes of rabbits.