Metabolites of microorganisms have long been considered as potential sources for drug discovery. In this study, five new depsidone derivatives, talaronins A-E (1–5) and three new xanthone derivatives, talaronins F–H (6–8), together with 16 known compounds (9–24), were isolated from the ethyl acetate extract of the mangrove-derived fungusTalaromycesspecies WHUF0362. The structures were elucidated by analysis of spectroscopic data and chemical methods including alkaline hydrolysis and Mosher’s method. Compounds1and2each attached a dimethyl acetal group at the aromatic ring. A putative biogenetic relationship of the isolated metabolites was presented and suggested that the depsidones and the xanthones probably had the same biosynthetic precursors such as chrysophanol or rheochrysidin. The antimicrobial activity assay indicated that compounds5,9,10, and14showed potent activity againstHelicobacter pyloriwith minimum inhibitory concentration (MIC) values in the range of 2.42–36.04 μmol/L. While secalonic acid D (19) demonstrated significant antimicrobial activity against four strains ofH. pyloriwith MIC values in the range of 0.20 to 1.57 μmol/L. Furthermore, secalonic acid D (19) exhibited cytotoxicity against cancer cell lines Bel-7402 and HCT-116 with IC50values of 0.15 and 0.19 μmol/L, respectively. The structure–activity relationship of depsidone derivatives revealed that the presence of the lactone ring and the hydroxyl at C-10 was crucial to the antimicrobial activity againstH. pylori. The depsidone derivatives are promising leads to inhibitH. pyloriand provide an avenue for further development of novel antibiotics.Supplementary InformationThe online version contains supplementary material available at 10.1007/s42995-023-00170-5.