Simultaneous Determination of Mephedrone, Methylone, MDPV, and
Amphetamines in Urine by LC/MS/MS Amanda Rigdon1*, Mike Coyer2, Jack Cochran1, Ty Kahler1, and Paul Kennedy3.
(1)Restek. Bellefonte, PA, USA. (2) Northern Tier Research, Mayfield, PA, USA. (3)Cayman Chemical, Ann Arbor, MI, USA.
Introduction
Abuse of substances marketed as ‘research chemicals’ often sold
for research purposes only or added to consumer products labeled
‘not for human consumption’ has become increasingly popular.
Cathinones, including mephedrone, methylone, and MDPV, are one
class of compounds that have appeared on the market as part of the
‘research chemical’ movement. These compounds are commonly
sold as bath salts, however drug users often snort or ingest these
compounds to induce an amphetamine-like high. (Figure 1). In
September 2011, mephedrone, methylone, and MDPV were placed
on the DEA Schedule I list on an emergency basis. Because of this
scheduling, demand for testing will increase.
Figure 1: Photos of Bath Salts (left) and ‘Research Chemicals’ (right)
Chromatographic Method Development and Validation:
Because these compounds are very similar to amphetamines, and
are used either as a substitute for or in conjunction with
amphetamines [1], a chromatographic method was developed to
detect both amphetamines and cathinones in a single run. In
addition to the compounds mentioned in the title of this poster,
several additional ‘research chemicals’ marketed to be similar to
MDPV were included in the method. The final chromatographic
method and results are detailed in Figure 2.
Figure 2: 100 ng/mL Spiked Urine Sample
Table 1: Peak List and Transitions Used
Method Performance:
A partial validation was performed on the three cathinones and
amphetamines listed in the title of this work. Preliminary data for the
remaining compounds was also collected, and is available upon
request. LOD/LOQ, linearity, precision, and accuracy data were
collected and are shown in Table 2. Note that at the time this work
was performed, only mephedrone and its deuterated analog were
available as certified reference standards. In order to evaluate the
method for the remainder of the compound list, the ‘research
chemicals’ shown in Figure 1 were used as mock reference standards
for this work. Although a thorough characterization was not performed,
all substances were analyzed for purity using LC-MS, and no
additional compounds from 100 – 1000m/z were detected in the
products. All samples were analyzed on a Shimadzu UFLCXR equipped with
an AB SCIEX API 4000 MS/MS. Rather than being extracted, samples were
prepared using a dilute-and-shoot methodology. All samples were diluted 10x
in starting mobile phase containing 30 ng/mL internal standard.
Table 2: Method Performance Summary
Discussion:
All values in the table above reflect results from the quantifier ion.
MDA, MDMA, and MDEA were not evaluated using external control
samples due to an error in writing the MS/MS method. These three
compounds were evaluated for the remaining parameters at a later
date. The calibration curve for this project was prepared from 1
ng/mL to 500 ng/mL, however, based on recent findings, urine
concentrations of MDPV can range from < 10 ng/mL to > 8000
ng/mL. When samples > 1000 ng/mL were analyzed according to
the method detailed above, both detector and column overloading
were observed. This can be remedied by using either a higher
sample dilution factor and/or a smaller injection volume, however a
range such as this may exceed the linear dynamic range of the
MS/MS detector.
Authentic Sample Analysis:
One authentic sample was
obtained for this study. This
sample was diluted as
described above and analyzed
along with a calibration curve.
MDPV was found at a level of
116 ng/mL in the sample
(Figure 3).
Figure 3: Authentic Sample
Positive for MDPV
Conclusion:
The method developed here is
suitable for the quantitative
analysis of cathinones and
amphetamines in urine at low
levels. Further work needs to be performed to determine
performance with high-level samples. Preliminary data indicates
that this method is also suitable for several of the newer designer
drugs that may become used more widely in the near future due
to the DEA scheduling of several cathinones.
References:
[1] P. Kriikku, L. Wilhelm, O. Schwarz, J. Rintatalo. Forensic Science International. 2011, 210, 1-3; 195 - 200