To further elucidate the metabolism of CUMYL-4CN-BINACA, a new synthetic cannabinoid with a cyano group, and to evaluate biomarkers, we incubated the substance in human hepatocytes and analysed 9 authentic urine specimens. We also quantified CUMYL-4CN-BINACA and cyanide in blood and provide comprehensive data on the 7 autopsy cases, 5 of them determined CUMYL-4CN-BINACA intoxications. For metabolite elucidation, CUMYL-4CN-BINACA was incubated with pooled human hepatocytes for up to 5hours, urine samples were analysed with and without enzymatic hydrolysis. Data was acquired in data-dependent mode by ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) with an Agilent 6550 QTOF. For quantitative analysis of CUMYL-4CN-BINACA, blood samples were precipitated and analysed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Cyanide was determined by gas chromatography-headspace-nitrogen phosphorus detection (GC-headspace-NPD). CUMYL-4CN-BINACA was metabolised via CYP450-mediated hydroxylation at 4-butyl position generating a cyanohydrin (M12), which releases free cyanide to form an aldehyde intermediate and eventually generates 4-hydroxybutyl CUMYL-BINACA (M11) and CUMYL-BINACA butanoic acid (M10). Other minor metabolites were produced by hydroxylation, dihydroxylation, N-dealkylation, and dihydrodiol formation; glucuronidation was observed. One urine sample showed high intensities of M10 and a wide variety of metabolites; the other samples contained fewer metabolites in low abundance and 1 sample showed no metabolites. CUMYL-4CN-BINACA blood concentrations ranged from 0.1 to 8.3ng/g showing an overlap between fatal and non-fatal concentrations. One blood sample contained 0.36g/g cyanide. Release of free cyanide during metabolism is worrying as it might induce liver toxicity. As suggested earlier, CUMYL-BINACA butanoic acid is the most abundant biomarker in urine, but monitoring of additional metabolites or, even better, analysis for the parent in blood is recommended.