Abstract: The sedimentary lipid pool comprises a myriad of components with some specific biomarkers used in paleoclimatic and geobiological reconstructions. However, a comprehensive view of the sedimentary lipidome is lacking. Here we conduct an untargeted analysis of the Black Sea sedimentary lipidome using high resolution mass spectrometry. Besides commonly reported phytoplankton-derived fossil lipids originate from oxic surface water, a diverse and abundant set of sphingolipids, accounting for ~20% of the lipidome, was discovered. These sphingolipids are produced in-situ by sedimentary anaerobic bacteria, likely in place of phospholipids due to the deficiency of phosphate in anoxic sediments. Our results suggest that while phytoplankton-derived lipids contribute 50–60% of the sedimentary lipidome, the importance of bacterial lipids, particularly in-situ produced sphingolipids, has been overlooked. Source data: Data 1. Spt_hits.and.backbone_sequences.MAFFT-L-INS-i.msa Data 2. Spt_hits.and.backbone_sequences.MAFFT-L-INS-i.msa.trimAl