Antibacterial Activity, GC MS Analysis and In Silico Validation of Bioactive Compound from Endophytic Fungus Lasiodiplodia pseudotheobromae EF-9

Secondary metabolites synthesized by endophytic fungi have garnered significant interest for their broad applications in treating various ailments. In this study involving 20 plant samples, 11 endophytic fungi were isolated and cultured, and Lasiodiplodia pseudotheobromae EF-9, derived from Hibiscus rosa-sinensis, demonstrated greater antibacterial efficacy than the other isolated endophytes. Phylogenetic analyses using 18S rRNA gene confirmed the EF-9 identity as L. pseudotheobromae. Following mass production, the active compound was partially purified using column chromatography. The fraction collected at the 60^th min exhibited good antibacterial activity against Bacillus coagulans (MTCC 6735) and Shigella flexneri (ATCC 12022), with an inhibition zone of approximately 20 mm in diameter. UV spectral studies revealed a wide absorption band at 430 nm. High Performance Liquid Chromatography (HPLC) of the active fraction showed a distinct peak with a retention time of 4.216 min at 430 nm. Gas Chromatography-Mass Spectrometry (GC-MS) identified the active compound in the L. pseudotheobromae EF-9 culture broth extract as Bis(2-ethylhexyl) phthalate, which displayed a peak at 16.856 min and covered 66.69 % of the area in the spectral analysis.