SOFT 2004 Meeting Abstracts - Society of Forensic Toxicologists

A17
HIGH.PERFORMANCE LIQUID CHROMATOGRAPHY AS AN ALTERNATIVE METHOD
FOR COMPARATIVE ANALYSIS OF SEIZED DRUG SAMPLES
2
Bogumila Byrska\ Dariusz Zuba*\ Malgorzata Swisf, Andrzej Parczewski l •
I Institute of Forensic Research, ul. Westerplatte 9,31-033 Cracow, Poland; 2 Faculty of Chemistry,
Jagiellonian University, ul. Ingardena 3, 30-060 Cracow, Poland
The comprehensive testing of seized drug samples performed in forensic laboratories allows identification
and determination of the controlled psychoactive substances, identification of additives (e.g. caffeine or
paracetamol) and diluents (e.g. palmitylic acid or glucose), as well as chemical impurities. The results of
analysis deliver information about synthesis methods applied by illegal producers. It is also possible to
establish if the seizure drug samples originate from the same production batch or the same illegal
laboratory and even to link the dealer with drug users. Hitherto, mainly gas chromatographic methods
(GClFlO, GCIMS) are used for this purpose.
The aim of the present study was to develop an analytical procedure that makes possible separation of the
substances occurring in the seized ecstasy tablets by means of the high-performance liquid chromatography
(HPLC). The studies have been carried out on the samples of 3,4-methylenedioxymethamphetamine
(MDMA, ecstasy) prepared by different routes: reductive amination, Leuckart reaction and safrole
bromonation. The reductive amination was performed using various reducing agents: sodium
cyanoborohydride, aluminium amalgam and sodium borohydride. In order to estimate the repeatability,
each synthesis route was repeated three times. HPLC analyses were carried out on LaChrom D·7000
System (Merck-Hitachi) liquid chromatograph.
The solid-phase extraction (SPE) was used as the sample preparation method. FIrst, 200 mg of ecstasy
sample was dissolved in mixture of 1.5 ml of the ammonium buffer (pH 8.5) and 0.5 ml of 30 mg/l
diphenylamine solution (internal standard). Than the suspension was vigorously shaken for 25 min at 2000
rpm. The optimisation ofSPE conditions was focused on selection of the appropriate column filling, which
enables to bind the main compound and to thicken the impurities that are important in view of comparative
analysis. Bakerbond spe Octadecyl (3 ml, 500 mg) turned out to be the most effective. SPE was
performed as follows: 1.5 ml of sample was rinsed with 6.0 ml of water and 0.3 ml of methanol and the
impurities were extracted with 0.8 ml of methanol. Than, 0.3 ml of collected fraction was diluted with the
phosphoric acid solution up to pH of 6.
Chromolith Performance RP-18 e (100 - 4.6 mm) monolithic column was used in HPLC analysis. Taking
into account the variety of physicochemical properties of the impurities, the separation was done in
gradient conditions. In order to optimise the resolution, the simplex method was applied. Four factors were
taken into account which describe gradient conditions. After eight steps of optimisation the following
conditions were selected: 0 min - 100% of phase A (water + 0.1 mIll phosphoric acid), 21.3 min - 59% of
phase A + 41 % of phase B (acetonitryle), 32.7 min 100% of phase B, 34 min 100% of phase A. The
flow was 1.0 mUmin. Diode array detector (DAD) was used.
The applied conditions of SPE extraction and HPLC analysis allow good separation of the characteristic
impurities occurring in ecstasy samples. Because most of the impurities give similar UVIVIS spectra, it is
impossible to identifY them using diode-array detector. It is highly probable that application of LCIMS
system will solve this problem. The obtained HPLC chromatograms could be treated equivalently to FlO
chromatograms where the impurities pattern is a subject to further chemometric analysis. The preliminary
results show that the worked-out method allows to distinguish the samples prepared by different synthesis
route as well as it could be useful in comparative analysis of seized drug samples.
The study was supported by the grant of the State Committee for Scientific Research, Warsaw, Poland,
number OT OOC 01024.
Keywords: Comparative analysis, Ecstasy, HPLC
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