Background The oceans are teeming with a diverse range of marine organisms that offer unique health benefits, such as seagrass which is one of many key marine products that have garnered attention for their potential therapeutic properties. However, until now there have been few successful reports of seagrass’s metabolites profile and biological activity. Therefore, this work aims to profile metabolites or chemical constituents and assess the potential antioxidants and anti-obesity effects of two seagrasses, Thalassia hemprichii and Zostera marina. Methods Once authenticated, T. hemprichii and Z. marina were extracted with two different solvents, polar (ethanol) and nonpolar (hexane). Metabolite profiling was performed using untargeted metabolomic profiling via liquid chromatography coupled to high-resolution tandem mass spectrometry method analysis, and then antioxidant and anti-obesity capabilities were assessed by molecular docking and in vitro studies on selected receptors. Results A total of 9 and 11 metabolites were observed from T. hemprichii and Z. marina and continued molecular docking. Some of the observed compounds have promising potential as inhibitors of human inducible nitric oxide synthase, reactive oxygen species (ROS) 1 kinase, human pancreatic lipase, and fat mass and obesity-associated (FTO) proteins, including luteolin, 6-hydroxy compounds luteolin O-glucoside, luteolin-O-sulphate, Thalassiolin A, Thalassiolin C, kaempferol-7,4′-dimethylether-3-O-sulfate, apigenin, and diosmetin. T. hemprichii ethanol extract (THE) EC50 value shows antioxidant capabilities via ABTS radical scavenging activity of 76.00 μg/mL, a smaller value than standard antioxidant controls (Trolox, 76.54 μg/mL) and followed by EC50 of lipase inhibition activity by THE which has the same pattern (EC50 THE < EC50 Orlistat). Conclusions This concludes that the two seagrasses have promising biological activity as candidates for functional food and/or drugs in combating free radicals and obesity.