Hydrocarbon contaminants are ubiquitous in urban aquatic ecosystems, and the ability of some microbial strains to degrade certain polycyclic aromatic hydrocarbons (PAHs) is well established. However, detrimental effects of petroleum hydrocarbon contamination on nondegrader microbial populations and photosynthetic organisms have not often been considered. In the current study, fatty acid methyl ester (FAME) biomarkers in the sediment record were used to assess historical impacts of petroleum contamination on microbial and/or algal biomass in South San Francisco Bay, CA, USA. Profiles of saturated, branched, and monounsaturated fatty acids had similar concentrations and patterns downcore. Total PAHs in a sediment core were on average greater than 20× higher above ∼200 cm than below, which corresponds roughly to the year 1900. Isomer ratios were consistent with a predominant petroleum combustion source for PAHs. Several individual PAHs exceeded sediment quality screening values. Negative correlations between petroleum contaminants and microbial and algal biomarkers – along with high trans/cis ratios of unsaturated FA, and principle component analysis of the PAH and fatty acid records – suggest a negative impacts of petroleum contamination, appearing early in the 20th century, on microbial and/or algal ecology at the site.