Postmortem distribution of mepirapim and acetyl fentanyl in biological fluid and solid tissue specimens measured by the standard addition method

Forensic Toxicology https://doi.org/10.1007/s11419-018-0431-z ORIGINAL ARTICLE Postmortem distribution of mepirapim and acetyl fentanyl in biological fluid and solid tissue specimens measured by the standard addition method Akira Mochizuki1 · Hiroko Nakazawa1 · Noboru Adachi2 · Kenichi Takekawa1 · Hideki Shojo2 Received: 6 May 2018 / Accepted: 26 June 2018 © The Author(s) 2018 Abstract Purpose Mepirapim is a new synthetic cannabinoid. We previously reported that the concentrations of unchanged mepirapim in whole blood and urine were much higher than those of other synthetic cannabinoids. To determine the postmortem distribution of mepirapim and acetyl fentanyl in the deceased individual, we established a standard addition method for detailed analysis by liquid chromatography–mass spectrometry (LC–MS) for quantification of these drugs. Methods The LC–MS method was fully validated for linearity, extraction recovery, matrix effect and repeatability. Results Good linearities, extraction recoveries, matrix effects and repeatabilities were shown for both target compounds in all specimens. The concentrations of mepirapim and acetyl fentanyl in three body fluid specimens and 12 solid tissue specimens were measured. For mepirapim, the highest concentrations were found in the liver and kidney, and the concentrations in the blood and urine specimens were one order of magnitude lower than the high concentrations in the solid tissues except the psoas major muscle. For acetyl fentanyl, the highest concentrations were found in the myocardium, spleen and kidney, and the concentrations in the body fluid specimens were also one order of magnitude lower than the highest concentrations in the solid tissues. There were concentration differences of mepirapim and acetyl fentanyl among the regions of the brain. Conclusions The concentration of unchanged mepirapim in urine was much higher than those of other synthetic cannabinoids; the higher dosage, urinary excretion, metabolisms and/or pharmacokinetics of mepirapim may be quite different from those of other synthetic cannabinoids. Keywords Mepirapim · Synthetic cannabinoid · Acetyl fentanyl · Postmortem distribution · LC–MS · Standard addition method Introduction Illicit psychoactive substances (e.g., synthetic cannabinoids, cathinone derivatives and synthetic opioids) have become a serious threat worldwide as designer drugs of abuse [1–3]. We previously encountered a curious case in which two male subjects self-dosed mepirapim plus acetyl fentanyl by different routes of administration, that is, intravenously and * Hideki Shojo 1 Forensic Science Laboratory, Yamanashi Prefectural Police Headquarters, 312‑4 Kubonakajima, Isawa, Fuefuki, Yamanashi 406‑0036, Japan 2 Department of Legal Medicine, Graduate Faculty of Interdisciplinary Research, University of Yamanashi, 1110 Shimokato, Chuo, Yamanashi 409‑3898, Japan by inhalation [4]. Mepirapim is a new and unique synthetic cannabinoid that was first identified in herbal blends in Japan [5]. Very recently, the affinities of mepirapim toward CB1 and CB2 in term of Ki values have been reported. The Ki values were 2650 and 1850 nM for CB1 and CB2, respectively [6], but it actually functions as a CB receptor agonist. This compound differs from JWH-018, because it has a 4-methylpiperazine group in place of the naphthyl group [4]. We thus reported a detailed gas chromatography–tandem mass spectrometry with the internal standard (IS) method for quantification of mepirapim and acetyl fentanyl in whole blood and urine [4]; the concentrations of unchanged mepirapim in whole blood and urine were much higher than those of other common synthetic cannabinoids. On the other hand, acetyl fentanyl is a synthetic fentanyl analog in which the propionyl group of fentanyl is replaced by an acetyl group [7]. Recently, acetyl fentanyl has been 13 Vol.:(0123456789) Forensic Toxicology encountered in clinical and forensic case studies [8–15]. The number of available reports dealing with determination of acetyl fentanyl from multiple specimens in authentic cases has been limited [3, 10, 11, 15]. In addition, the number of specimens was few; one report included five specimens [11], and another one included four specimens [15]. The most detailed report was provided by Poklis et al. [10], including seven specimens using 14 fatal cases. In this study, we have quantified both mepirapim and acetyl fentanyl in as many as 15 specimens. To our knowledge, this is the most detailed study to date for the distribution of acetyl fentanyl, and also is the first demonstration of distribution of mepirapim in an authentic fatal poisoning case. Such detailed investigation of postmortem distribution is very useful for evaluating the cause(s) of death. In addition, we have used the standard addition method for analysis by liquid chromatography–mass spectrometry (LC–MS), which can overcome the different matrix effects and recovery rates in different specimens, and also does not need blank human matrices for validation experiments. Case history The deceased in the autopsy case used in our analysis was one of two subjects who had abused drugs together in December 2013. According to the confession statement of the surviving individual, the deceased individual (a male in his 60s) self-administered the drug (approximately 50–60 mg) via intravenous injection. Approximately 10 h after dosing, the surviving individual recognized that the other individual had died. An autopsy was performed, and postmortem biological fluid (heart whole blood, femoral vein whole blood and urine) and solid (cerebrum, cerebellum, pons, medulla oblongata, lung, myocardium, liver, pancreas, kidney, adrenal gland, spleen and psoas major muscle) tissue specimens were collected. The postmortem interval was estimated to be 64 h, but because the cadaver had been stored at 4 °C until autopsy, it was relatively fresh at autopsy. All specimens were stored at − 80 °C until analysis. The ‘Angela’ drug product found at the scene was seized and analyzed; the powder consisted of 73.2 ± 0.4% mepirapim and 18.9 ± 0.2% acetyl fentanyl (w/w) [4]. Drug analyses from the deceased were performed at our department by the request of judicial authorities. Materials and methods Materials Mepirapim hydrochloride, acetyl fentanyl, JWH-200 and acetyl fentanyl-d5 were purchased from Cayman Chemical 13 (Ann Arbor, MI, USA), and Isolute SLE + (1 mL capacity) columns from Biotage (Uppsala, Sweden). Other common chemicals used in this study were of the highest purity commercially available. Extraction procedure To 50 µL of each body fluid or 100 µL solid tissue homogenate (2 g tissue in 8 mL 0.01 N HCl solution thoroughly homogenized with a blender) were added 20 µL of methanol solution containing a known amount of mepirapim and acetyl fentanyl as standard additions, 20 µL of methanol solution containing 50 ng of JWH-200 and 10 ng of acetyl fentanyl-d5 as ISs for mepirapim and acetyl fe (...truncated)