Aims and Objectives: The aim of this study was to identify the optimum method to obtain one of the chemical compounds in the water fraction and to identify the hypothesized chemical isolates in the water fraction katuk leave's ethanol extract.
Materials and Methods: The methods used in this study included the collection and determination of the katuk plant, the processing of the katuk, phytochemical filtrating, extracting with ethanol 96%, and fractionation using the liquid-liquid extraction method with n-hexane, ethyl acetate, and water solvents The water fraction of katuk leaves was analyzed by its components by thin-layer chromatography using the stationary phase of silica gel 60 F254, developer of n-butanol:acetic acid:water (4:1:5), and detection under ultraviolet (UV) light at a wavelength of 366 and 254nm, as well as with vanillin-sulfuric acid reagent. To isolate the compounds from water fraction of katuk leaves, it was then eluted with a vacuum column chromatography by eluent with a level polarity that would get 11 subfractions. Each subfraction was checked by two-dimensional thin-layer chromatography to see subfraction purity characterized by the appearance of a spot on the chromatogram plate. The isolate was analyzed using spot test, ultraviolet-visible spectrophotometer, infrared spectrophotometer, and liquid chromatography-mass spectrometry.
Results: The isolate was an alkaloid compound with a molecular mass of 406.3131 m/z with the molecular formula C21H39N6O2 as S, S-5, 5'-amino-4,4'-dihexyl-propyldihydropyrazol-3, 3-one.
Conclusion: One of the chemical compounds contained in the water fraction of the ethanol extract of the katuk leaf was an alkaloid group.
METHODOLOGY: Qualitative phytochemical analysis was firstly carried out to determine the possible active compounds in P. betle leaves methanolic extract. The antibacterial activities of major compounds from this extract against nine fish pathogenic bacteria were then assessed using TLC-bioautography agar overlay assay and their quantity were determined simultaneously by HPLC method.
RESULTS: The use of methanol has proved to be successful in extracting numerous bioactive compounds including antibacterial compounds. The TLC-bioautography assay revealed the inhibitory action of two compounds which were identified as hydroxychavicol and eugenol. The $-caryophyllene however was totally inactive against all the tested bacterial species. In this study, the concentration of hydroxychavicol in extract was found to be 374.72±2.79 mg g-1, while eugenol was 49.67±0.16 mg g-1.
CONCLUSION: Based on these findings, it could be concluded that hydroxychavicol and eugenol were the responsible compounds for the promising antibacterial activity of P. betle leaves methanolic extract. This inhibitory action has significantly correlated with the amount of the compounds in extract. Due to its potential, the extract of P. betle leaves or it compounds can be alternative source of potent natural antibacterial agents for aquaculture disease management.