Petroleum associated with sulfide-rich sediment is present in Escanaba Trough at the southern end of the Gorda Ridge spreading axis offshore from northern California within the Exclusive Economic Zone (EEZ) of the U.S. This location and occurrence are important for evaluation of the mineral and energy resource potential of the seafloor under U.S. jurisdiction. In Escanaba Trough, petroleum is believed to be formed by hydrothermal processes acting on mainly terrigenous organic material in Quaternary, river-derived sediment. To attempt to simulate these processes in the laboratory, portions of a Pleistocene gray-green mud, obtained from ∼ 1.5 m below the seafloor at a water depth of ∼ 3250 m in Escanaba Trough, were heated in the presence of water in four hydrous-pyrolysis experiments conducted at temperatures ranging from 250 to 350°C and at a pressure of 350 bar for 1.0–4.5 days. Distributions of n-alkanes, isoprenoid hydrocarbons, triterpanes, and steranes in the heated samples were compared with those in a sample of hydrothermal petroleum from the same area. Mud samples heated for less than 4.5 days at less than 350°C show changes in some, but not all, molecular marker ratios of organic compounds that are consistent with those expected during hydrothermal petroleum formation. Our results suggest that the organic matter in this type of sediment serves as one possible source for some of the compounds found in the hydrothermal petroleum.