A Metabolomics Study of Retrospective Forensic Data from Whole

Dec 25, 2015 - A Metabolomics Study of Retrospective Forensic Data from Whole Blood Samples of Humans Exposed to 3 ...
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A Metabolomics Study of Retrospective Forensic Data from Whole Blood Samples of Humans Exposed to 3,4Methylenedioxymethamphetamine: A New Approach for Identifying Drug Metabolites and Changes in Metabolism Related to Drug Consumption Kirstine L. Nielsen,* Rasmus Telving, Mette F. Andreasen, Jørgen B. Hasselstrøm, and Mogens Johannsen* Department of Forensic Medicine, Section for Forensic Chemistry, Aarhus University, Palle Juul-Jensens Boulevard 99, DK-8200 Aarhus N, Denmark S Supporting Information *

ABSTRACT: The illicit drug 3,4-methylenedioxymethamphetamine (MDMA) has profound physiological cerebral, cardiac, and hepatic effects that are reflected in the blood. Screening of blood for MDMA and other narcotics are routinely performed in forensics analysis using ultraperformance liquid chromatography with high-resolution time-of-flight mass spectrometry (UPLC−HR-TOFMS). The aim of this study was to investigate whether such UPLC−HR-TOFMS data collected over a twoyear period could be used for untargeted metabolomics to determine MDMA metabolites as well as endogenous changes related to drug response and toxicology. Whole blood samples from living Danish drivers’ positive for MDMA in different concentrations were compared to negative control samples using various statistical methods. The untargeted identification of known MDMA metabolites was used to validate the methods. The results further revealed changes of several acylcarnitines, adenosine monophosphate, adenosine, inosine, thiomorpholine 3-carboxylate, tryptophan, S-adenosyl-L-homocysteine (SAH), and lysophospatidylcholine (lysoPC) species in response to MDMA. These endogenous metabolites could be implicated in an increased energy demand and mechanisms related to the serotonergic syndrome as well as drug induced neurotoxicity. The findings showed that it was possible to extract meaningful results from retrospective UPLC−HR-TOFMS screening data for metabolic profiling in relation to drug metabolism, endogenous physiological effects, and toxicology. KEYWORDS: 3,4-methylenedioxymethamphetamine (MDMA), ecstasy, metabolism, metabolomics, tryptophan, acylcarnitine, thiomorpholine 3-carboxylate, S-adenosyl-L-homocysteine, UPLC-HR-TOFMS



INTRODUCTION

Genes, transcripts, and proteins are essential predictors for drug responses, addiction, and toxicity, but the metabolome may provide different additional insights and biomarkers of importance.9,10 Metabolic profilling studies on the actions of amphetamines are however few. A study by Perrine et al. (2009)11 investigated the acute cardiac effects of MDMA in rats showing an increase and decrease in cardiac carnitine and choline, respectively. The authors suggested this to be due to an increased fatty acid metabolism in response to an increased workload of the heart, whereas choline concentrations could indirectly impact membrane compositions and cell-signaling through altered phosphatidylcholine synthesis. A metabolic profiling study of the acute effects of methamphetamine in urine and plasma from rats also showed an impaired energy metabolism through TCA cycle intermediates, glucose, and β-

3,4-Methylenedioxymethamphetamine (MDMA, ecstasy) is an illicit drug that has stimulant and hallucinogenic properties. It affects the peripheral and central nervous system by acting as an indirect monoaminergic agonist causing an efflux of serotonin, dopamine, and norepinephrine.1,2 Consumption of MDMA is associated with various adverse acute, mid-, and long-term effects including hyperthermia, hypertension, tachycardia, rhabdomyolysis, liver damage, and neurotoxicity.2,3 Metabolism of MDMA happens primarily in the liver involving two main metabolic pathways: (1) N-demethylation and (2) Odemethylenation followed by O-methylation.4 The metabolites are found in plasma and urine mainly as glucuronide or sulfate conjugates.5 Oxidation to quinones may additionally form adducts with glutathione or other thiol-containing compounds that are distributed to other organs including the brain where they may cause neurotoxic effects.6−8 © XXXX American Chemical Society

Received: November 5, 2015

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DOI: 10.1021/acs.jproteome.5b01023 J. Proteome Res. XXXX, XXX, XXX−XXX

Article

Journal of Proteome Research

Table 1. Identified Metabolites Found to Distinguish Human Whole Blood Samples with MDMA from Control Samples Predicted by Multiple Linear Regressions, Pearson Correlations, LASSO Logistic Regressionk, and/or OPLS-DA VIP-Scoresa Pearson correlation metabolite d

MDMA

MDAd HMMA 4-sulfatee HMMA 4-glucuronidee HHMA 3-sulfatef AMPd adenosined inosined S-adenosyl-L-homocysteined hexanoylcarnitined,g 2-octenoylcarnitineh,g decanoylcarnitined 9-decenoylcarnitinei,g dodecanoylcarnitined tetradecenoylcarnitinef,g unknown 1 unknown 2 uric acidd tryptophand

thiomorpholine 3-carboxylatef,j LysoPC (16:0)d

LysoPC (17:0)d LysoPC (18:1)d glycerolf unknown 3

unknown 4 unknown 5

m/z

P

r

P

VIPc

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