Endophytic fungi allied to plants have sparked substantial promise in discovering new bioactive compounds. In this study, propagation of the endophytic fungusAlternaria alternataHE11 obtained fromColocasia esculantaleaves led to the isolation of Ergosterol (1),β-Sitosterol (2), Ergosterol peroxide (3), in addition to three dimeric naphtho-γ-pyrones, namely Fonsecinone A (4), Asperpyrone C (5), and Asperpyrone B (6), which were isolated from genusAlternariafor the first time. Structures of the isolated compounds were established on the basis of extensive 1D and 2D NMR and, MS measurements. The ethyl acetate extract, as well as compounds 1, 3, 4 and 6 were evaluated for their antimicrobial activity using agar well-diffusion and broth microdilution assays. Molecular docking study was carried out to explore the pharmacophoric moieties that governed the binding orientation of antibacterial active compounds to multidrug efflux transporter AcrB and the ATP binding site toE. coliDNA gyrase using MOE software. Results revealed that the most active antibacterial compounds 4 and 6 bind with high affinity in the phenylalanine-rich cage and are surrounded with other hydrophobic residues. The antiproliferative activity of all isolated compounds was in vitro evaluated using the human prostatic adenocarcinoma cell lines DU-145, PC-3, PC-3 M, 22Rv1 and CWR-R1ca adopting MTT assay. Compound 4 was the most active against almost all tested cell lines, with IC50values 28.6, 21.6, 17.1 and 13.3 against PC-3, PC-3 M, 22Rv1 and CWR-R1ca cell lines, respectively.Graphical AbstractSupplementary InformationThe online version contains supplementary material available at 10.1186/s13568-023-01568-1.