Development of an analytical method for simultaneous detection of psychotropic phenylalkylamines in hair by LC-MS/MS with a multi-mode reversed-phase column using pH gradient elution
Introduction
Methamphetamine (MA), which undergoes demethylation to amphetamine (AM), is a highly addictive drug with potent central nervous system stimulant properties [1] and is traditionally the most abused drug in Korea. Amphetamine-derived new designer drugs, such as 3,4-methylenedioxymethamphetamine (MDMA), have also become popular as rave drugs among young people since being introduced in the late 1990s. The use of ecstasy (MDMA) in combination with other substances, such as ketamine, has recently been reported [2]. In recent years, there has been increasing abuse of ketamine in East and Southeast Asia including Korea [3]. On the other hand, amphetamine-like anorectic drug, phentermine, has become popular among MA and MDMA abusers and is presumably considered an alternative or a synergist for amphetamines abuse [4]. In addition to phentermine, other stimulants used primarily for weight loss, such as fenfluramine, phendimetrazine, and amfepramone, are also of interest because they are occasionally detected in imported diet products from China and have a causative association with serious health problems [5], [6].
Hair is a useful specimen to prove chronic drug use and analytical results from hair analysis provide conclusive evidence of previous drug use and the severity of the drug abuse [7]. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) has recently become a more popular tool for hair analysis due to its sensitivity and simplicity in sample preparation relative to gas chromatography/mass spectrometry (GC–MS). Most LC methods for analyzing low molecular amine have employed a reversed-phase column in combination with low organic mobile phase. However, the use of high organic mobile phase, which works more effectively with multi-mode reversed-phase column than with reversed-phase column, is advantageous for highly sensitive electrospray ionization-mass (ESI-MS) detection [8]. Therefore, combination of multi-mode reversed-phase column and high organic mobile phase provides the highest sensitivity most suitable for detecting trace-level amine compounds. Furthermore, standard reversed-phase columns have some limitations to separate low molecular amines which have structural similarity and show same multiple reaction monitoring (MRM) transition such as MA and phentermine.
Therefore, in order to enhance the sensitivity and achieve the good separation, we adopted the multi-mode reversed-phase column, Scherzo SS-C18, which is composed of strong ionic ligands (quaternary ammonium and sulfonyl groups) and C18 ligands and used pH gradient elution to separate seven psychotropic phenylalkylamines and their metabolites, which covers the major amphetamines and anorectics in Korea (Table 1). In this study, a simultaneous analytical method for determining MA, AM, MDMA, 3,4-methylenedioxyamphetamine (MDA), phentermine, fenfluramine, norfenfluramine, phendimetrazine, phenmetrazine, amfepramone, diethylnorephedrin, ketamine, and norketamine levels in hair was developed and validated for application to actual hair samples collected from drug abuse suspects. The obtained quantitative results were compared with those generated using a reference GC–MS method [12].
Section snippets
Chemicals and reagents
All solvents were of high-performance liquid chromatography grade. Methanolic solutions of MA, AM, MDMA, MDA, phentermine, fenfluramine, ketamine, and norketamine for the analytical study were obtained from Cerilliant (Round Rock, TX, USA). Norfenfluramine, phendimetrazine, and diethylnorephedrin were purchased from Sigma–Aldrich (St. Louis, MO, USA). Phenmetrazine and amfepramone were obtained from U.S. Pharmacopeia (Rockville, MD, USA). Methanolic solution of the deuterated internal standard
Principle of separation
The principle of separation of amines using the multi-mode reversed-phase column, which is composed of strong ionic ligands and C18 ligands, is ionic and hydrophobic interactions [13]. Retention of an ionic compound can be affected by eluent pH. The amines were ionized in the initial acidic gradient and retained strongly on the column via ionic interactions with the anionic ligands of the column. Then, the analytes were neutralized during pH gradient elution, depending on their pKa values. The
Conclusion
In this study, a simultaneous analytical method for seven psychotropic phenylalkylamines and their metabolites in hair by LC-MS/MS with multi-mode reversed-phase column using pH gradient elution was established and validated successfully. The analytical separation using the Scherzo SS-18 column employing pH gradient showed good separation and peak shape and narrow peak width compared with those of a previous study using normal reversed-phase column. The developed method was successfully applied
Conflict of interest
There are no financial or other relations that could lead to a conflict of interest.
Acknowledgements
This study was supported by funding from the National Forensic Service of Korea (2015-02).
References (15)
Pharmacology and Toxicology of Amphetamine and Related Designer Drugs. NIDA Research Monograph 94
(1989)- et al.
Characteristics and trends of 3,4-methylenedioxymethamphetamine (MDMA) tablets found in Taiwan from 2002 to February 2005
Forensic Sci. Int.
(2006) - et al.
Recent trends in drug abuse in China
Acta Pharmacol. Sin.
(2006) - et al.
Detection of phentermine in hair samples from drug suspects
Forensic Sci. Int.
(2011) - et al.
Appetite-suppressant drugs and the risk of primary pulmonary hypertension
New Engl. J. Med.
(1996) - et al.
Hair analysis for fenfluramine and norfenfluramine as biomarkers for N-nitrosofenfluramine ingestion
Forensic Sci. Int.
(2004) - et al.
Distribution of methamphetamine and amphetamine in drug abusers’ head hair
Forensic Sci. Int.
(2009)
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