SECONDARY METABOLITES OF ENDOPHYTE FUNGI Colletotrichum phyllanthi FROM THE CALYX OF Physalis peruviana, ANTIBACTERIAL ASSAY AND EFFECT OF ADDITION VALPROIC ACID ELICITOR

• Main Author: HIDAYAH - NIM : 20516020, RATIH
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/30236
• Institution: Institut Teknologi Bandung

<p align="justify">Endophytic fungi are microorganisms that live colonize in plant tissue without causing negative effect to its host. It takes symbiotic mutualism. Endophytic fungi as secondary metabolite sources that have important biological activities including anticancer, antifungal and antibacterial. Endophytic fungi can be isolated from various types of plants both high levels of plants and herbaceous plants. Physalis peruviana (Solanaceae) is one of the herbaceous plants are widely used as a traditional medicine. Phytochemically this plant produces steroid, terpenoid, alkaloid, phenyl propanoid and flavonoid compounds. Studies on secondary metabolites of endophytic fungi from P. peruviana have not been previously reported. The study of endophytic fungi of the same genus has been performed on P. alkekengi that contain Gibberellin-producing fungus Fusarium proliferatum KGL0401. In another study, from the leaf of P. alkekengi was successfully isolated endophytic fungus Thielavia sp. PA0001 (Chaetomiaceae). Isolation of the fungus produces some secondary metabolites nor-spiro-azafilon and bis-spiro-azafilon skeleton. Thus, research on the isolation of secondary metabolites of endophytic fungi from the genus Physalis is important to develop. Therefore, in this study, the isolation and characterization of secondary metabolites of endophytic fungi from the calyx of P. peruviana and the determination of their antibacterial activity. <br /> <br /> <br /> The steps of this study began with sample selection, surface sterilization to kill epiphytic microorganisms, tissue inoculation in PDA solid medium (Potato Dextrose Agar) and followed by subculture to get single isolate CCK3.2. The results of molecular identification is Colletotrichum phyllanthi. The next process is large scale cultivation in liquid PDB medium (Potato Dextrose Broth) with incubation at 28° C for 14 days. The endophytic fungi is separated between mycelia and its medium by filtration method using Buchner. The mycelia treathed by a liquid solid extraction with maceration method using methanol. While the liquid medium treated by liquid-liquid extraction using ethyl acetate. The extract methanol of mycelia and the ethyl acetate extract of medium were fractionated and purified using chromatographic techniques which include column chromatography of gravity and radial chromatography. The isolated secondary metabolite structure was determined based on 1H NMR, 13C NMR, HSQC, HMBC, TOCSY1D and NOESY spectroscopy data. Antibacterial activity was determined in vitro by microdilution assay to get Minimum Inhibitory Concentration (MIC). <br /> <br /> <br /> Based on the methodology, two steroid skeleton secondary metabolites have been successfully isolated from the methanol extract of miselia, ergosterol (1) and ergosterol peroxide (2). From the ethyl acetate extract of liquid media obtained three isolates identified as four secondary metabolites, tyrosol (3), 2,3-dihydroxybenzoic acid (4), and mixture of nectriapyrone (5) and 2-phenylethanol (6). The addition of 500 ?M valproic acid elicitor to the culture medium showed an altered TLC profile on the ethyl acetate extract of liquid media. The isolation of the extract yielded 2-methylheptan-1,5-diol (7) which was not previously obtained from the ethyl acetate extract of the medium without the addition of an elicitor. The determination of antibacterial activity was performed on two Gram-(+) bacteria, Staphylococcus aureus and Bacillus cereus, and also two Gram-(-) bacteria, Citrobacter freundii and Salmonella enterica. Results of determination of antibacterial activity obtained MIC value >500 µg/mL for extracts and MIC >100 µg/mL for secondary metabolites with amoxicillin as a positive control having MIC value 0,78 – 6,25 µg/mL. Based on these results it can be concluded that C. phyllanthi produces secondary metabolites that are inactive as antibacterial. The addition of the valproic acid elicitor affects the formation of secondary metabolites in C. phyllanthi.<p align="justify"> <br /> <br />