The study proposes a new pathway for the formation of gammacerane from tetrahymanol, involving sulphuration and early C-S cleavage in marine sediments. Evidence for this pathway includes the sequestration of gammacerane skeletons in S-rich macromolecular aggregates formed by natural sulphurization of functionalized lipids, and the release of gammacerane from these aggregates upon mild artificial maturation using hydrous pyrolysis. The stable carbon isotopic compositions of gammacerane and lipids from primary producers and green sulphur bacteria in Miocene and Upper Jurassic formations indicate that gammacerane is derived from bacterivorous ciliates feeding on green sulphur bacteria. This suggests that gammacerane is an indicator of water column stratification, explaining its presence in both hypersaline and lacustrine deposits. The findings also address two current enigmas in gammacerane geochemistry: why gammacerane is often found in hypersaline environments but not exclusively so, and why lacustrine deposits may contain abundant gammacerane.The study proposes a new pathway for the formation of gammacerane from tetrahymanol, involving sulphuration and early C-S cleavage in marine sediments. Evidence for this pathway includes the sequestration of gammacerane skeletons in S-rich macromolecular aggregates formed by natural sulphurization of functionalized lipids, and the release of gammacerane from these aggregates upon mild artificial maturation using hydrous pyrolysis. The stable carbon isotopic compositions of gammacerane and lipids from primary producers and green sulphur bacteria in Miocene and Upper Jurassic formations indicate that gammacerane is derived from bacterivorous ciliates feeding on green sulphur bacteria. This suggests that gammacerane is an indicator of water column stratification, explaining its presence in both hypersaline and lacustrine deposits. The findings also address two current enigmas in gammacerane geochemistry: why gammacerane is often found in hypersaline environments but not exclusively so, and why lacustrine deposits may contain abundant gammacerane.