Polybrominated diphenyl ethers (PBDEs) are flame retardants widely used in electronic and domestic goods. Thesepersistent pollutants are present in the environment and in humans, and their toxicological properties are of growingconcern. PBDEs can be metabolised into compounds suspected to be responsible for their toxicity. These metaboliteshave been characterised quite well in rodents and fish, but available information in humans remains scarce. For theiridentification, an efficient method for the simultaneous analysis of PBDEs, hydroxylated PBDEs (OH-PBDEs), andother PBDE metabolites in a single run was needed and has been developed in this work. Atmospheric pressureionisation modes were compared, and Atmospheric Pressure Photo-Ionization (APPI) was selected. After carefulsetting of APPI parameters such as dopant and operating temperature, the optimised method was based on APPIionization coupled to High-Resolution Mass Spectrometry operating in the full scan mode at a resolution of60 000. This provided excellent sensitivity and specificity, allowing the discrimination of signals which could notbe resolved on a triple quadrupole used as a reference. The full-scan high-resolution acquisition mode allowedmonitoring of both parent PBDEs and their metabolites, including hydroxylated PBDEs, with detection limitsranging from 0.1 pg to 4.5 pg injected on-column based on the investigated standard compounds. The method wasapplied to the study of BDE-47 metabolism after incubation with human primary cultures of hepatocytes, and provedto be efficient not only for monitoring the parent compound and expected hydroxylated metabolites, but also for theidentification of other non-targeted metabolites. In addition to hydroxy-BDE-47, several conjugated metabolitescould be located, and the formation of a dihydrodiol derivative was evidenced for the first time in the case of PBDEsin this work.