For the first time, to our knowledge, anionic and nonionic surfactants were analyzed in an oil matrix by ultra-highperformance liquid chromatography hyphenated to high-resolution mass spectrometry (UHPLC-HRMS). The feasibility of this analysis was studied using synthetic mixtures of surfactants prepared in water (quality controls), binary THF/toluene 50/50 v/v (surfactant + THF/toluene), and binary THF/toluene containing 1 and 10% crude oil (Crudel% and Crude10%). These compositions were chosen in order to be as close as possible to petroleum related samples to be investigated in the future. Analyses were carried out by UHPLC methods using both reverse phase and anion-exchange mechanisms with a mixed mode column. Despite the complexity of the oil matrix and the presence of organic solvents used for dilution, the retention times of the surfactants were not affected whatever the concentration of crude oil present in the sample. Nevertheless, a significant matrix effect caused a loss of signal when the concentration of oil reached 10% in mass. For the analysis of samples with this crude oil concentration range, it would be advisable to dilute the sample.