Forensic Toxicol (2007) 25:1–7
DOI 10.1007/s11419-006-0018-y
ORIGINAL ARTICLE
Identification and quantitative determination of 5-methoxy-N,N-di-n-
propyltryptamine in urine by isotope dilution gas chromatography-
mass spectrometry
Akihiro Nakamoto · Akira Namera · Manami Nishida
Mikio Yashiki · Takako Kuramoto · Kojiro Kimura
Received: 8 November 2006 / Accepted: 7 December 2006 / Published online: 26 January 2007
© Japanese Association of Forensic Toxicology and Springer 2007
Abstract A simple method for analysis of 5-methoxy-
N,N-di-n-propyltryptamine (5-MeO-DPT) in urine has
been developed using gas chromatography-mass
spectrometry (GC-MS) with tetradecadeuterated 5-
MeO-DPT as internal standard, which is useful for
discrimination from 5-methoxy-N,N-diisopropyltrypt-
amine (5-MeO-DIPT). These tryptamine designer drugs
were extracted from urine with Extrelut, and derivatized
with trifluoroacetic anhydride prior to GC-MS analysis.
The recovery of 5-MeO-DPT from urine was 90.7%; the
calibration curve showed linearity in the range of 0.01–
2.0 µg/ml. When urine samples containing two different
concentrations (0.1 and 1.0 µg/ml) of 5-MeO-DPT were
analyzed, the coefficients of variation for intraday and
interday testing ranged from 3.11% to 5.82%. The estab-
lished method was applied to an acute poisoning case
in which 5-MeO-DIPT abuse had been suspected. Un-
expectedly, 5-MeO-DIPT could not be detected, but 5-
MeO-DPT was found in the subject’s urine and in an
aqueous sample seized upon investigation of the poison-
ing; the electron-impact ionization and chemical ioniza-
tion mass spectra well agreed with those of authentic
5-MeO-DPT. The concentration of 5-MeO-DPT in urine
A. Nakamoto · A. Namera · M. Nishida · M. Yashiki ·
K. Kimura
Department of Legal Medicine, Graduate School of Biomedical
Sciences, Hiroshima University, Hiroshima, Japan
A. Nakamoto (*) · T. Kuramoto
Scientific Investigation Laboratory, Hiroshima Prefectural Police
Headquarters, 2-26-3 Kohnan, Naka-ku, Hiroshima 730-0825,
Japan
e-mail: kasouken@police.pref.hiroshima.jp
of this case was 0.37 µg/ml. Although several reports
have appeared describing poisoning cases of 5-MeO-
DIPT, which carries the street name “Foxy,” to our
knowledge, this is the first report of GC-MS analysis of
5-MeO-DPT and demonstration of its use in a poisoning
case.
Keywords 5-Methoxy-N,N-di-n-propyltryptamine ·
5-Methoxy-N,N-diisopropyltryptamine · Designer
drug · GC-MS · Hallucinogenic tryptamines · Foxy
Introduction
Psychoactive compounds contained in plants and fungi
have been traditionally used for religious rituals and
augury, because of their ability to cause dysesthesia and/
or mystic experiences including hallucination [1]. These
hallucinogens are sometimes abused as psychotomimetic
drugs to alter thoughts, feelings, and perceptions. In ad-
dition to these naturally occurring compounds, many
derivatives are illegally chemically synthesized as design-
er drugs. Hallucinogens are classified into lysergic acids,
phenethylamines, tryptamines, and others according to
their chemical structures. Abuse of these drugs has be-
come a serious social problem worldwide, and many
kinds of them are available on the market; the levels of
control differ in different countries. Recently, illicit
drugs have become easily obtainable all over the world
through the Internet, and acute poisoning caused by
such drugs occurs frequently. Tryptamine derivatives are
often used and some poisoning cases have been reported
[2–6]. There are also reports of problems arising from
mislabeling of ingredients in “2C-C”, “2CT-21” and
“4-acetoxy-DIPT” [7,8].
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2 Forensic Toxicol (2007) 25:1–7
Recently, abuse and poisoning using 5-methoxy-N,N-
diisopropyltryptamine (5-MeO-DIPT) have become
topics in designer drug toxicology [2–6]. Therefore, its
isomer 5-methoxy-N,N-di-n-propyltryptamine (5-MeO-
DPT) was anticipated to appear as a drug of abuse.
As described in this report, we have had some involve-
ment in the investigation of a case of poisoning by
5-MeO-DPT. This has led us to establish a detailed
procedure for analysis of 5-MeO-DPT in urine by gas
chromatography-mass spectrometry (GC-MS), the
details of which are presented in this report. Our method
enables easy discrimination between 5-MeO-DPT and
5-MeO-DIPT.
Materials and methods
Materials
5-MeO-DIPT was donated by Dr. H. Inoue of the
National Research Institute of Police Science (Kashiwa,
Japan). 5-MeO-DPT and its tetradecadeuterated com-
pound (5-MeO-DPT-d14) were synthesized in our labora-
tory using a modification of the previously published
procedure [9]. Stock solutions (1 mg/ml) were separately
prepared by dissolving 5-MeO-DIPT, 5-MeO-DPT, or
5-MeO-DPT-d14 in acetonitrile. The solutions were
stored at 4ºC. 5-MeO-DPT-d14 was stable for at least a
year. Standard solutions and spiked samples were pre-
pared by diluting the stock solutions with ethyl acetate
and drug-free urine, respectively. The drug-free urine
collected from a healthy male adult was kept frozen at
−20ºC until used. Trifluoroacetic anhydride (TFAA)
was purchased from Wako (Osaka, Japan); other re-
agents and solvents used were of the highest purity
commercially available. Extrelut granule was purchased
from Merck (Darmstadt, Germany), and was washed
with three volumes of diethyl ether and was dried at 50ºC
for 1 h before use; 2.0 g each of Extrelut granule was
packed in a small (9-ml volume) glass tube, the bottom
of which was stopped with glass wool.
GC-MS conditions
Two GC-MS instruments were used; a Shimadzu
(Kyoto, Japan) GC-MS QP-5050A and an Agilent (Palo
Alto, CA, USA) 6890GC/5973 mass selective detector
(MSD). For qualitative analysis of the seized aqueous
sample, the Shimadzu GC-MS QP-5050A, equipped
with an HP-5MS fused silica capillary column (30 m ×
0.25 mm i.d., film thickness 0.25 µm, Agilent), was used.
The oven temperature was set at 70ºC for 1 min, then
13
programmed from 70º to 300ºC at 5ºC/min and held at
300ºC for 3 min. The temperatures of the injection port
and the interface were set at 260º and 280ºC, respec-
tively. Splitless injection mode was used. Helium with an
inlet pressure of 127 kPa was used as carrier gas. The
MSD was operated at 70 eV in the electron-impact ion-
ization (EI) mode. For the chemical ionization (CI)
mode, isobutane was used as a CI reagent gas. Scan
ranges in the EI and CI modes were m/z 50 to 550 and
m/z 150 to 550, respectively.
For quantitative analysis of urine, the Agilent
6890GC/5973MSD, equipped with the same HP-5MS
fused silica capillary column, was used. The oven tem-
perature was set at 50ºC for 1 min, then programmed
from 50º to 280ºC at 15ºC/min, and held at 280ºC for
5 min. The temperature of the injection port and the de-
tector were set at 250º and 280ºC, respectively. Splitless
injection mode was used. Helium with an inlet pressure
of 127 kPa was used as carrier gas. The MSD was oper-
ated in the EI mode at 70 eV. A calibration curve was
drawn by plotting the peak area ratio of 5-MeO-DPT
(m/z 114) to 5-MeO-DPT-d14 (m/z 128) against the con-
centration of the analyte.
Procedure for the seized aqueous sample
without derivatization
A 0.1-ml volume of a seized aqueous sample was diluted
with 0.9 ml of distilled water and mixed with 28% aque-
ous ammonia to adjust the pH to 10. The analyte was
extracted with 1.0 ml of ethyl acetate. After phase sepa-
ration by centrifugation (3000 rpm, 5 min), 1 µl of the
organic layer was injected into the GC-MS instrument
without derivatization.
Established procedure for 5-MeO-DPT in urine
with derivatization
A 0.5-ml volume of urine was mixed with 1.0 ml of
Sorensen’s glycine buffer (0.1 M glycine, 0.1 M NaCl; pH
11.0, adjusted with NaOH solution) and 5 µl of internal
standard (IS) (5-MeO-DPT-d14, 0.1 mg/ml). The mixture
was applied to the Extrelut column. After 10 min at
room temperature, 5-MeO-DPT and IS were eluted with
6 ml of ethyl acetate. Then the eluate was evaporated to
dryness under a stream of nitrogen. The residue was dis-
solved in 0.1 ml of TFAA/ethyl acetate (1 : 3, v/v) and
heated at 50ºC for 10 min. After evaporation of the de-
rivatized mixture, the residue was dissolved in 0.2 ml of
ethyl acetate and 1 µl was injected into the GC-MS
instrument.
Forensic Toxicol (2007) 25:1–7 3

Fig. 1 Total ion

chromatogram of a seized
aqueous sample (5-methoxy-
N,N-di-n-propyltryptamine;
5-MeO-DPT) superimposed
on that of 5-methoxy-N,
N-diisopropyltryptamine
(5-MeO-DIPT). The
compounds were
underivatized

Fig. 2 Electron-impact
ionization (EI) and chemical
ionization (CI) mass spectra of
the seized aqueous sample
(5-MeO-DPT) and 5-MeO-
DIPT without derivatization.
A CI mass spectrum of 5-MeO-
DIPT; B CI mass spectrum of
seized aqueous sample (5-MeO-
DPT); C EI mass spectrum of
5-MeO-DIPT; D EI mass
spectrum of the seized aqueous
sample (5-MeO-DPT). Note
the characteristic peak for
5-MeO-DPT indicated by an
arrow

Results and discussion
Encounter with a seized aqueous sample containing
a “Foxy”-like substance
We became aware of a poisoning case in which a man
injected a drug solution into the anus of another man
for sexual excitement. The administered man fell into
unconsciousness and was sent to a hospital. The patient
fortunately survived after treatment. The solution
containing a high concentration of a drug was seized
and brought to our laboratories. The man who injected
the solution alleged that the solution contained
5-MeO-DIPT.

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4 Forensic Toxicol (2007) 25:1–7
Fig. 3 Probable
fragmentation pathways in
the EI mode for
underivatized 5-MeO-DIPT
(A) and 5-MeO-DPT (B and (A)
C)
(B)
(C)
We first tried to identify the compound using its au-
thentic standard after simple liquid–liquid extraction by
GC-MS without derivatization. As shown in Fig. 1, the
retention time of the compound in question was clearly
longer than that of 5-MeO-DIPT. The EI and CI mass
spectra of the compound were very similar to those of
5-MeO-DIPT (Fig. 2); the CI mass spectra of the com-
pound in question and 5-MeO-DIPT showed protonated
molecular ions at m/z 275, which suggested that the com-
pound was an isomer of 5-MeO-DIPT. The only differ-
ence in the EI mass spectra was the appearance of a small
peak at m/z 86 for the compound in the seized solution.
Therefore, the compound was presumed to be 5-MeO-
DPT, which led us to chemically synthesize 5-MeO-DPT
and its deuterated analog. The EI mass spectrum of the
synthesized compound coincided with that obtained
from the seized solution, confirming that this was a case
of 5-MeO-DPT poisoning.
13
Fragmentation pathways for peaks appearing in the EI
mass spectra
We analyzed the fragmentation pathways for the peaks
appearing in the EI mode for both underivatized 5-MeO-
DPT and 5-MeO-DIPT. As shown in Fig. 3, the m/z 114
ions were presumed to be due to N,N-di-n-propylamino-
methylene and N,N-diisopropylaminomethylene liber-
ated, respectively. The m/z 160 ion was attributed to
methoxyindolemethylene. The m/z 86 ion observed in
the EI mass spectrum of 5-MeO-DPT was thought to
have been generated via a McLafferty-type rearrange-
ment and β-cleavage as shown in Fig. 3C. Therefore, the
generation of the m/z 86 ion was possible only for 5-
MeO-DPT [10], which has a di-n-propylamino group,
but not for 5-MeO-DIPT, which has a diisopropylamino
group (Fig. 3A). Generally, when the tertiary amino
compounds are equivalent in molecular weight and
constitutional isomers, the di-n-alkylamino-substituted
compound has a larger intermolecular force than the
Forensic Toxicol (2007) 25:1–7
diisoalkylamino-substituted compound; the boiling
point of the former becomes higher than that of the lat-
ter. Therefore, the retention time of 5-MeO-DPT be-
comes longer than that of 5-MeO-DIPT.
Establishment of the procedure for GC-MS analysis of
5-MeO-DPT in urine
The established procedure for analysis of 5-MeO-DPT
in urine is described in the Materials and methods sec-
tion. The analysis of free aromatic and aliphatic amines
by GC or GC-MS is generally associated with difficulties
in sensitivity and reproducibility, because of adsorption
to and interaction with glass inserts in the GC and the
Fig. 4 CI and EI mass spectra
of the authentic 5-MeO-DPT
and internal standard (IS; 5-
MeO-DPT-d14) after
trifluoroacetyl (TFA)
derivatization. A CI mass
spectrum of 5-MeO-DPT-
TFA; B EI mass spectrum of
5-MeO-DPT-TFA; C EI mass
spectrum of IS-TFA
5
analytical column; this results in poor peak resolution
and lower sensitivity. Therefore, derivatization is usually
needed for GC analysis of these compounds. Ishida et
al. [11] have described that 5-MeO-DIPT is not deriva-
tized using acetic anhydride. However, we tried to de-
rivatize 5-MeO-DIPT with TFAA, and it was successful;
it was found very useful to improve peak resolution and
sensitivity. The CI and EI mass spectra of the authentic
5-MeO-DPT and IS are shown in Fig. 4. The TFA group
is considered to bind to the imine group of the pyrrole
ring of 5-MeO-DPT. Even after TFA derivatization, the
EI mass spectrum of the compound showed the charac-
teristic peak at m/z 86, but that of 5-MeO-DIPT did not.
In view of the retention times, it was also easy to separate
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6 Forensic Toxicol (2007) 25:1–7

the 5-MeO-DPT peak from the 5-MeO-DIPT peak after
derivatization (data not shown). The retention times of
5-MeO-DPT-TFA and 5-MeO-DPT-d14-TFA were 14.12
and 14.07 min, respectively.
Liquid–liquid extraction is usually used for extraction
of tryptamines in urine. However, this method has many
disadvantages: (1) a large sample volume is needed for
extraction; (2) a large volume of extraction solvent is
needed; (3) an emulsion is formed. To overcome these
disadvantages, Extrelut was used for extraction of the
tryptamine drugs in urine. The target compounds could
be extracted from the biological materials using Extrelut
under alkaline conditions. The recovery rate with the
Extrelut extraction was 90.7% at 1 µg/ml for a urine
sample.
Urine samples spiked with 5-MeO-DPT were pre-
pared at concentrations of 0.001, 0.005, 0.01, 0.05, 0.1,
0.5, 1.0, and 2.0 µg/ml, and then analyzed by GC-MS
using the established procedure. There was linearity
from 0.01 to 2.0 µg/ml. The limit of detection (signal/
noise = 3) in urine was 0.005 µg/ml. The correlation coef-
ficient of the calibration curve for 5-MeO-DPT was
0.999. The intraday coefficients of variation (CVs) at
concentrations of 0.1 and 1.0 µg/ml for 5-MeO-DPT in
urine with five replicates were 3.11% and 4.61%, respec-
tively. The interday CVs at concentrations of 0.1 and
1.0 µg/ml of 5-MeO-DPT in urine with measurements
over five consecutive days were 5.82% and 4.21%, respec-
tively. Considering the above data, the proposed method
was found to be fully reproducible.

Fig. 5 Mass chromatograms

of the extracts of blank urine, Blank urine
spiked urine, and urine from
the poisoning case

Spiked urine

5-MeO-DPT-d14-TFA
5-MeO-DPT-TFA

Urine of the poisoning case

5-MeO-DPT-d14-TFA
5-MeO-DPT-TFA

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Forensic Toxicol (2007) 25:1–7
Application to the urine sample obtained from
the poisoning case
The present analytical method was applied to the urine
sample obtained from the described poisoning case.
Mass chromatograms of extracts from blank urine,
spiked urine, and the seized urine sample are shown in
Fig. 5. A sharp and symmetrical peak of 5-MeO-DPT
was obtained without disturbance by endogenous impu-
rities. The concentration of 5-MeO-DPT in the urine of
the victim was 0.37 µg/ml. The toxicity of 5-MeO-DPT
cannot be compared with that of 5-MeO-DIPT, because
no data for 5-MeO-DPT poisoning have been reported
so far; however, the detected concentration may be at a
toxic level.
Conclusions
To our knowledge, this is the first report describing a
GC-MS method for 5-MeO-DPT and demonstrating its
use in an actual poisoning case. For its isomer 5-MeO-
DIPT, some reports of poisoning and analysis have ap-
peared recently [3–6,11]. The pharmacological action of
5-MeO-DPT is expected to be analogous to that of 5-
MeO-DIPT, although it has not been documented. At
present in Japan, 5-MeO-DIPT is a controlled substance,
but 5-MeO-DPT is not. The pressing need for the ability
to discriminate between the two compounds in forensic
toxicology has been addressed by the method presented
in this report.
Acknowledgments The authors thank Dr. H. Inoue of the
National Research Institute of Police Science, Japan, for donation
of the standard of 5-MeO-DIPT, and Dr. K. Somekawa of
7
Kagoshima University for advice on the fragmentation modes for
5-MeO-DPT.
References
1. Schultes RE, Hofman A (1992) Plants of the gods. Healing
Arts, Rochester, VT
2. Morano RA, Spies C, Walker FB, Plank SM (1993) Fatal in-
toxication involving etryptamine. J Forensic Sci 38:721–725
3. Meatherall R, Sharma P (2003) Foxy, a designer tryptamine
hallucinogen. J Anal Toxicol 27:313–317
4. Vorce SP, Sklerov JH (2004) A general screening and confir-
mation approach to the analysis of designer tryptamines and
phenethylamines in blood and urine using GC-EI-MS and
HPLC-electrospray-MS. J Anal Toxicol 28:407–410
5. Wilson JM, McGeorge F, Smolinske S, Meatherall R (2005)
A foxy intoxication. Forensic Sci Int 148:31–36
6. Tanaka E, Kamata T, Katagi M, Tsuchihashi H, Honda K
(2006) A fatal poisoning with 5-methoxy-N,N-diisopropyl-
tryptamine, Foxy. Forensic Sci Int 163:152–154
7. Kojima H, Miyazawa M, Ishikawa T, Takayanagi H, Asatani
H, Doi K (2004) Conclusion of an investigation into the actual
condition about “chemical drugs” (in Japanese). Jpn J Toxicol
17:71–72
8. Kojima H, Miyazawa M, Takayanagi H, Shimizu A, Doi K
(2005) Current state of so-called “chemical drug”—difference
between display name and content element (in Japanese). Jpn
J Toxicol 18:83–85
9. Shulgin A, Shulgin A (1997) TiHKAL. Transform, Berkeley,
CA, pp 527–528
10. Brandt SD, Freeman S, Fleet IA, McGagh P, Alder JF (2005)
Analytical chemistry of synthetic routes to psychoactive trypt-
amines part II. Characterization of the Speeter and Anthony
synthetic route to N,N-dialkylated tryptamines using GC-EI-
ITMS, ESI-TQ-MS-MS and NMR. Analyst 130:330–344
11. Ishida T, Kudo K, Kiyoshima A, Inoue H, Tsuji A, Ikeda N
(2005) Sensitive determination of alpha-methyltryptamine
(AMT) and 5-methoxy-N,N-diisopropyltryptamine (5MeO-
DIPT) in whole blood and urine using gas chromatography-
mass spectrometry. J Chromatogr B 823:47–52
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