Marine tunicates of the genus Didemnum are a rich source of structurally diverse and biologically potent metabolites. As part of our ongoing search for bioactive secondary metabolites from ascidians, we investigated Didemnum sp. collected from Sulawesi Island (Indonesia) due to the potent cytotoxicity of its methanolic extract against tumor cells. Here, we report the isolation and structural elucidation of didemniserinolipids A-C (1-3), the first serinolipids from a marine organism. These compounds consist of a 2-amino-1,3-propanediol (serinol) unit linked to a hydroxylated α,β-unsaturated acid with an unusual 6,8-dioxabicyclo[3.2.1]octane framework. Isolation involved solvent extraction and chromatographic fractionation (silica gel flash chromatography, HPLC). Structural characterization using high-resolution FAB mass spectrometry (HRFABMS) and multi-dimensional NMR spectroscopy (1H/13C NMR, COSY, HMQC, HMBC, NOESY/ROESY) revealed their structures: compound 2 is the ethyl carboxylate 10-deacetyl derivative of 1, and compound 3 has a free hydroxyl group at C-10 and a nonesterified carboxylic acid at C-1. Notably, these are the first monosubstituted serinol derivatives isolated from a marine source, while the 6,8-dioxabicyclo[3.2.1]octane substructure has been found in potent toxins like palytoxins. Although the CH2Cl2 fraction of the extract showed strong cytotoxicity, compounds 1-3 exhibited no cytotoxic activity against P388, A549, and HT29 tumor cell lines (IC50 > 20 µg/mL).