Purpose <p>The abuse of 2-fluoro-2-oxo-phenylcyclohexylethylamine (2F-2-oxo-PCE), a dissociative anesthetic structurally related to phencyclidine (PCP) and ketamine, has recently increased in South Korea. This study presented the first forensic toxicological detection of 2F-2-oxo-PCE in autopsy cases and described a validated method for the simultaneous quantification of 2F-2-oxo-PCE, methylenedioxymethamphetamine (MDMA), methylenedioxyamphetamine (MDA), ketamine, and norketamine in postmortem blood.</p> Method <p>2F-2-oxo-PCE and its metabolite, 2F-deschloronorketamine (2F-DCNK) were identified using gas chromatography-mass spectrometry (GC–MS). A liquid chromatography-tandem mass spectrometry (LC–MS/MS) with solid-phase extraction (SPE) was developed and validated for the simultaneous quantification of 2F-2-oxo-PCE, MDMA, MDA, ketamine and norketamine in blood.</p> Results <p>The validation parameters including linearity, accuracy, precision, matrix effect, and recovery were satisfactory. In postmortem blood samples, 2F-2-oxo-PCE concentrations ranged from 664 to 7911&#xa0;ng/mL. Concurrent detection with MDMA and ketamine suggested possible polydrug use contributing to fatal outcomes.</p> Conclusions <p>The validated LC–MS/MS method is suitable for forensic applications and may enhance the toxicological profiling of emerging dissociative substances. The results can provide critical baseline data for interpreting 2F-2-oxo-PCE-related intoxications.</p>