Analytical and Bioanalytical Chemistry

, Volume 402, Issue 6, pp 2141–2151

Generation of metabolites by an automated online metabolism method using human liver microsomes with subsequent identification by LC-MS(n), and metabolism of 11 cathinones

Original Paper

DOI: 10.1007/s00216-011-5678-8

Cite this article as:
Mueller, D.M. & Rentsch, K.M. Anal Bioanal Chem (2012) 402: 2141. doi:10.1007/s00216-011-5678-8

Abstract

Human liver microsomes (HLMs) are used to simulate human xenobiotic metabolism in vitro. In forensic and clinical toxicology, HLMs are popularly used to study the metabolism of new designer drugs for example. In this work, we present an automated online extraction system we developed for HLM experiments, which was compared to a classical offline approach. Furthermore, we present studies on the metabolism of 11 cathinones; for eight of these, the metabolism has not previously been reported. Metabolites were identified based on MS2 and MS3 scans. Fifty-three substances encompassing various classes of drugs were employed to compare the established offline and the new online methods. The metabolism of each of the following 11 cathinones was studied using the new method: 3,4-methylenedioxy-N-benzylcathinone, benzedrone, butylone, dimethylcathinone, ethylone, flephedrone, methedrone, methylone, methylethylcathinone, naphyrone, and pentylone. The agreement between the offline and the online methods was good; a total of 158 metabolites were identified. Using only the offline method, 156 (98.7%) metabolites were identified, while 151 (95.6%) were identified using only the online method. The metabolic pathways identified for the 11 cathinones included the reduction of the keto group, desalkylation, hydroxylation, and desmethylenation in cathinones containing a methylenedioxy moiety. Our method provides a straightforward approach to identifying metabolites which can then be added to the library utilized by our clinical toxicological screening method. The performance of our method compares well with that of an established offline HLM procedure, but is as automated as possible.

Keywords

LC-MS Human liver microsomes Cathinones Online extraction Human metabolism 

Abbreviations

APCI

Atmospheric pressure chemical ionization

AU

Arbitrary units

bk

Beta-keto

CYP

Cytochrome P450

EDTA

Ethylenediaminetetraacetic acid

ESI

Electrospray ionization

GST

Glutathione-S-transferase

G6P

Glucose-6-phosphate

G6PD

Glucose-6-phosphate dehydrogenase

HPLC

High-pressure liquid chromatography

HLMs

Human liver microsomes

IU

International units

LC-MS

Liquid chromatography–mass spectrometry

MDPV

Methylenedioxypyrovalerone

NADP+

Nicotinic acid adenine dinucleotide phosphate (oxidized form)

NADPH

Nicotinic acid adenine dinucleotide phosphate (reduced form)

NAT

N-Acetyltransferase

Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  1. 1.Institute for Clinical ChemistryUniversity Hospital ZurichZurichSwitzerland

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