New-generation azaindole-adamantyl-derived synthetic cannabinoids

Article abstract of DOI:10.1007/s11419-019-00466-1

Purpose: This work reports the synthesis and pharmacological and analytical data for a new series of recently identified azaindole-adamantyl-derived synthetic cannabinoids (SCs). Methods: Each SC was synthesised using an efficient?and divergent synthesis, and assessed by electron ionisation mass spectrometry (EIMS). The cannabimimetic activity of each compound was conducted using a fluorometric imaging plate reader (FLIPR) assay. Results: The described EIMS method and retention time by gas chromatography were able to effectively differentiate each of the analogues regardless of the bicyclic core. For the first time in these SC structures, the bicyclic ring system was shown to have an impact on the cannabimimetic activities in the fluorometric assay of membrane potential. Analogues ranged from moderately potent at both CB₁ and CB₂ (e.g., AP4AIC EC₅₀ = 160?nM and EC₅₀ = 64?nM, respectively) to not active at either cannabinoid receptor (AP4AICA, AP5AICA, and APIC). Conclusions: Further investigation into receptor selectivity surrounding these bicyclic cores could prove useful for future therapeutic applications.

Full text of DOI:10.1007/s11419-019-00466-1

Forensic Toxicology
https://doi.org/10.1007/s11419-019-00466-1
ORIGINAL ARTICLE
New‑generation azaindole‑adamantyl‑derived synthetic cannabinoids
Mitchell Longworth1 · Tristan A. Reekie1 · Karen Blakey2 · Rochelle Boyd3 · Mark Connor4 · Michael Kassiou1
Received: 5 December 2018 / Accepted: 4 February 2019
© The Author(s) 2019
Abstract
Purpose This work reports the synthesis and pharmacological and analytical data for a new series of recently identifed
azaindole-adamantyl-derived synthetic cannabinoids (SCs).
Methods Each SC was synthesised using an efcient and divergent synthesis, and assessed by electron ionisation mass
spectrometry (EIMS). The cannabimimetic activity of each compound was conducted using a fuorometric imaging plate
reader (FLIPR) assay.
Results The described EIMS method and retention time by gas chromatography were able to efectively diferentiate each
of the analogues regardless of the bicyclic core. For the frst time in these SC structures, the bicyclic ring system was shown
to have an impact on the cannabimimetic activities in the fuorometric assay of membrane potential. Analogues ranged from
moderately potent at both CB1 and CB2 (e.g., AP4AIC EC50 =160 nM and EC50 =64 nM, respectively) to not active at either
cannabinoid receptor (AP4AICA, AP5AICA, and APIC).
Conclusions Further investigation into receptor selectivity surrounding these bicyclic cores could prove useful for future
therapeutic applications.
Keywords Synthetic cannabinoid · Azaindole · Adamantyl · CB1 and CB2 · APICA · Gas chromatography–mass
spectrometry
Introduction
JWH-018 (2) was one of the frst SCs to be identifed as
a drug of abuse. The capacity for diversifcation of the
aminoalkylindole-derived SC scafold has resulted in clan-
destine chemists making modifcations to the N-pentyl and
3-acyl substituents in an attempt to avoid detection by law
enforcement agencies. As a result, SCs have become one
of the most prominent classes of novel psychoactive sub-
stances (NPS) [1]. Because of the limited understanding of
their pharmacological efects, these compounds have caused
a number of unpredictable adverse events, posing a major
risk to public health [2–9]. The rate at which new chemical
entities are appearing, along with the plethora of potential
substitution patterns, has made it virtually impossible for
law enforcement agencies to stay ahead of newly appearing
SCs (Fig. 1).
The feld of synthetic cannabinoids (SCs) has been con-
stantly changing since their emergence in the early 2000s.
These compounds are reported to elicit efects similar to
∆9-THC (1), the psychoactive component of cannabis, by
activating the two known cannabinoid receptors: cannabi-
noid type-1 (CB1) and cannabinoid type-2 receptors (CB2).
Electronic supplementary material The online version of this
article (https://doi.org/10.1007/s11419-019-00466-1) contains
supplementary material, which is available to authorized users.
* Michael Kassiou
michael.kassiou@sydney.edu.au
1
Our group has reported the pharmacological and ana-
lytical data for a variety of identifed indole- and indazole-
derived SCs containing N-substituents, including pentyl,
5-fuoropentyl, butyl, methylcyclohexyl, and p-fuorobenzyl,
and various 3-acyl-substituted skeletons containing ada-
mantyl, l-valinate, l-tert-leucinate, and cumyl functionali-
ties (examples include 4 to 6 in Fig. 1) [10–13]. Whereas
School of Chemistry, The University of Sydney, Sydney,
NSW 2006, Australia
2
Forensic Chemistry Section, Queensland Health Forensic
and Scientifc Services, Coopers Plains, QLD 4108, Australia
3
Children’s Medical Research Institute, Westmead,
NSW 2145, Australia
4
Department of Biomedical Sciences, Macquarie University,
Sydney, NSW 2109, Australia
Vol.:(0123456789)
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Forensic Toxicology
Fig. 1 Examples of known syn-
thetic cannabinoids (SCs) and a
phytocannabinoid
∆9-THC shows only partial agonist activity at CB1, these
SCs have proven to be highly potent and efcacious agonists
at both CB1 and CB2. There have also been reports of SC
CB1 activation resulting in promiscuous G-protein coupling
and of-target receptor activation, potentially contributing to
their toxicological profles [14, 15].
2015, QHFSS obtained an illicit sample of plant material
portraying an EIMS consistent with a 1-N-pentyl azaindole/
indazole and an adamantyl carboxylate substituent at the
3-position. As nothing with these chemical characteristics
has previously been reported, EIMS was unable to distin-
guish which indazole/azaindole ring system was present in
the sample. Detection of this compound was subsequently
reported in South Korea in 2016 and Russia in 2017 [26, 27].
A series of reference samples with an indazole or azain-
dole core were synthesised to assess whether the EIMS frag-
mentation patterns could diferentiate between the indazole/
azaindole isomers. In conjunction, derivatives incorporat-
ing the amide functionality (7–11), along with the indole
bicyclic ring system (4 and 12) (see structures in Fig. 2),
were also synthesised and assessed in vitro at the CB1 and
CB2 receptors to determine functional trends within this new
class of SCs. Each analogue was designated an abbreviation
following a protocol similar to lead compounds 4 and 7 (e.g.,
Adamantan-1-yl 1-Pentyl-1H-4-AzaIndole-3-Carboxylate
was given the abbreviation AP4AIC).
In 2013, a new modification to the SC scaffold was
observed, incorporating an ester linker, seen in analogues
such as PB-22 (QUPIC), 5F-PB-22, and BB-22 (QUCHIC),
identifed in Japan and Russia [16, 17]. This modifcation
was subsequently identifed in another series of other com-
pounds including FUB-PB-22, NM-2201, and 5F-NM-
2201 [18]. PB-22 was detected in illicit samples received
at Queensland Health Forensic and Scientific Services
(QHFSS) in mid-2013, along with 5F-PB-22, and became a
prominent compound in the Queensland SC market through-
out 2013 and 2014.
Additional SC modifcations incorporating an azaindole
scafold have begun to appear in recent years. CUMYL-
5F-P7AICA, the 7-azaindole isomer of CUMYL-5F-PIN-
ACA, has been reported in forensic samples from 2015
to 2017 in Switzerland, Germany, and Australia, and was
detected in illicit samples received at QHFSS in late 2015
[19–21]. Other 7-azaindole SCs have been reported in the
NPS market, including NNL-1, NNL-3, 5F-NPB-22-7N,
5F-AKB-48-7N, CUMYL-4CN-B7AICA, and AB-5F-P7A-
ICA [20, 22–24]. While the majority of azaindole SCs
reported have been the 7-azaindole, the potential for other
isomers to enter the NPS market has been demonstrated with
the reporting of 5F-PCN, the 5-azaindole isomer of 5F-MN-
18 [25]. The presence of azaindole/indazole structural iso-
mers in the NPS market can present analytical challenges
for forensic identifcation, as electron ionisation mass spec-
trometry (EIMS) data are likely to be similar and may not
provide clear discrimination between each isomer. In early
Materials and methods
General chemical synthesis details
All reactions were performed under an atmosphere of
nitrogen or argon unless otherwise specifed. Anhydrous
methanol (MeOH), acetonitrile, and dimethyl sulfoxide
(DMSO) (Sigma-Aldrich, St. Louis, MO, USA) were used
as received. Anhydrous tetrahydrofuran (THF) and toluene
were obtained from a PureSolv MD 7 solvent purifcation
system (Innovative Technology, Inc., Oldham, UK). Other
commercially available chemicals (Sigma-Aldrich) were
used as received. Analytical thin-layer chromatography
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Forensic Toxicology
Fig. 2 Adamantyl-derived SCs
synthesised in this work
was performed using Merck aluminum-backed silica gel
60 F254 (0.2 mm) plates (Merck, Darmstadt, Germany),
which were visualised using shortwave (254 nm) UV
fuorescence. Flash chromatography was performed using
Merck Kieselgel 60 (230–400 mesh) silica gel, with the
eluent mixture reported as the volume/volume ratio (%).
Melting point ranges (m.p.) were measured in open capil-
laries using a Stanford Research Systems (SRS, Sunny-
vale, CA, USA) MPA160 melting point apparatus with
a ramp rate of 0.5–2.0 °C/min. Nuclear magnetic reso-
nance (NMR) spectra were recorded at 300 K using either
a Bruker AVANCE DRX 300 (300 MHz), AVANCE
DRX400 (400.1 MHz), or AVANCE III 500 Ascend
(500.1 MHz) spectrometer (Bruker, Billerica, MA, USA).
The data are reported as chemical shift (δ ppm) relative to
the residual protonated solvent resonance, relative integral,
multiplicity (s = singlet, br s = broad singlet, d = doublet,
t = triplet, quat. = quartet, quin. = quintet, m = multiplet),
coupling constants (J Hz), and assignment. Assignment of
signals was assisted by correlation spectroscopy (COSY),
distortionless enhancement by polarisation transfer
(DEPT), heteronuclear single quantum coherence (HSQC),
and heteronuclear multiple-bond correlation (HMBC)
experiments where necessary. Low-resolution mass spectra
(LRMS) were recorded using electrospray ionisation (ESI)
on a Finnigan LCQ ion trap mass spectrometer (Thermo
Fisher Scientifc, Waltham, MA, USA). High-resolution
mass spectra (HRMS) were run on a Bruker 7T Apex-Qe
Fourier transform (FT) ion cyclotron resonance mass spec-
trometer equipped with an Apollo II ESI/APCI/MALDI
dual source (Bruker) by the Mass Spectrometry Facility
of the School of Chemistry at the University of Sydney.
Infrared (IR) absorption spectra were recorded on a Bruker
ALPHA FT-IR spectrometer (Bruker) as solid or thin flm
from ethanol, and the data are reported as vibrational fre-
quencies (cm−1).
Preparation of N‑pentyl‑3‑trifuoroacetylindole (19)
Sodium hydride (60% dispersion in mineral oil, 137 mg,
3.42 mmol) was added portion-wise to an ice-cold solution
of indole (200 mg, 1.71 mmol) in N,N-dimethylformamide
(DMF, 6 mL) and stirred for 10 min. 1-Bromopentane (225 µL,
1.80 mmol) was added, and the mixture was warmed to ambi-
ent temperature and stirred for 1 h. The mixture was re-cooled
to 0 °C, and trifuoroacetic anhydride (600 µL, 4.28 mmol)
was added dropwise. The reaction mixture was warmed to
ambient temperature and stirred for 1 h. The reaction mix-
ture was poured onto ice (75 mL) and extracted with CH2Cl2
(3×75 mL). The combined organic extracts were washed with
H2O (100 mL) and brine (100 mL), dried (MgSO4), and con-
centrated under reduced pressure, giving 19, following purif-
cation by fash chromatography [hexane/ethyl acetate (EtOAc),
94:6 v/v], as a yellow oil (450 mg, 94%).
1H NMR (300 MHz, CDCl3): δ 8.43 (1H, d, J=8.7 Hz),
7.93 (1H, s), 7.42–7.36 (3H, m), 4.21 (2H, t, J=6.9 Hz),
1.95 (2H, quin., J=7.2 Hz), 1.43–1.32 (4H, m), 0.92 (3H,
t, J=6.6 Hz); 13C NMR (75 MHz, CDCl3): δ 174.8 (q, CO,
J=34.5 Hz), 137.5 (d, CH, J=4.5 Hz), 136.8 (quat.), 127.3
(quat.), 124.6 (CH), 124.0 (CH), 122.8 (CH), 117.3 (q,
CF3, J=289.5 Hz), 110.5 (CH), 109.5 (quat.), 47.8 (CH2),
29.5 (CH2), 29.0 (CH2), 22.3 (CH2), 14.0 (CH3); 19F NMR
(282 MHz, CDCl3): δ −72.2 (3F, s); LRMS (+ESI): m/z 306
([M+Na]+, 100%); IR (diamond cell, thin flm): 3124 (w),
2959 (m), 2933 (m), 2863 (w), 1662 (s), 1527 (s), 1397 (m),
1286 (m), 1181 (s), 1132 (s), 878 (m), 751 (m).
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Forensic Toxicology
22.2 (CH2), 13.9 (CH3); LRMS (+ ESI): 189 ([M + H]+,
100%); IR (diamond cell, thin flm): 2956 (m), 2929 (m),
2860 (w), 1604 (m), 1417 (s), 1322 (m), 1290 (m), 771 (s),
723 (m).
Preparation of 1‑N‑pentyl‑3‑indazole methyl car‑
boxylate (22)
Potassium tert-butoxide (350 mg, 3.12 mmol) was added
to an ice-cold solution of methyl-3-indazole carboxylate
(500 mg, 2.84 mmol) in THF (15 mL), warmed to ambient
temperature, and stirred for 1 h. 1-Bromopentane (415 µL,
2.98 mmol) was added and stirred at ambient temperature
for 48 h and then heated to refux for 24 h. The reaction
mixture was cooled to ambient temperature, poured into
H2O (100 mL), and extracted with diethyl ether (Et2O,
3 × 100 mL). The combined organic extracts were dried
(MgSO4) and concentrated under reduced pressure, giving
22, following purifcation by fash chromatography (hexane/
EtOAc, 90:10 v/v), as a colourless oil (585 mg, 84%).
1H NMR (300 MHz, CDCl3): δ 8.24 (1H, d, J=8.0 Hz),
7.50–7.44 (2H, m), 7.35–7.27 (1H, m), 4.47 (2H, t,
J = 7.4 Hz), 4.04 (3H, s), 1.97 (2H, quin., J = 7.0 Hz),
1.32 (4H, m), 0.87 (3H, t, J=6.6 Hz); 13C NMR (75 MHz,
CDCl3): δ 163.3 (CO), 140.6 (quat.), 134.6 (quat.), 126.8
(CH), 123.9 (quat.), 123.1 (CH), 122.3 (CH), 109.7 (CH),
52.1 (CH2), 50.1 (CH3), 29.7 (CH2), 29.0 (CH2), 22.4 (CH2),
14.0 (CH3); LRMS (+ESI): m/z 269 ([M+Na]+, 60%), 515
([2M+Na]+, 100%); IR (diamond cell, thin flm): 2954 (m),
2932 (m), 2860 (w), 1709 (s), 1477 (s), 1215 (s), 1159 (s),
1117 (s), 751 (s).
Preparation of 1‑N‑pentyl‑5‑azaindole (29)
Treating 5-azaindole (250 mg, 2.12 mmol) according to gen-
eral procedure A gave 29, following purifcation by fash
chromatography (EtOAc), as a red oil (285 mg, 71%).
1H NMR (300 MHz, CDCl3): δ 8.89 (1H, s), 8.29 (1H, d,
J=5.7 Hz), 7.22 (1H, d, J=5.7 Hz), 7.10 (1H, d, J=2.4 Hz),
6.57 (1H, d, J=1.2 Hz), 4.12 (2H, t, J=6.9 Hz), 1.82 (2H,
quint., J=7.2 Hz), 1.30 (4H, m), 0..89 (3H, t, J=7.1 Hz);
13C NMR (75 MHz, CDCl3): δ 144.1 (CH), 140.6 (CH),
139.6 (quat.), 128.8 (CH), 125.5 (quat.), 104.9 (CH), 100.9
(CH), 46.4 (CH2), 30.1 (CH2), 29.1 (CH2), 22.4 (CH2), 14.3
(CH3); LRMS (+ESI): m/z 189 ([M+H]+, 100%); IR (dia-
mond cell, thin flm): 2956 (m), 2929 (m), 2859 (w), 1602
(m), 1476 (m), 1453 (m), 1319 (m), 1295 (m), 890 (m), 801
(m), 724 (s).
Preparation of 1‑N‑pentyl‑6‑azaindole (30)
Treating 6-azaindole (1.00 g, 8.46 mmol) according to gen-
eral procedure A gave 30, following purifcation by fash
chromatography (EtOAc), as a red oil (1.35 g, 85%).
1H NMR (300 MHz, CDCl3): δ 8.77 (1H, s), 8.22 (1H, d,
J=5.4 Hz), 7.50 (1H, d, J=5.4 Hz), 7.49 (1H, m), 6.47 (1H,
m), 4.19 (2H, t, J= 7.2 Hz), 1.87 (2H, quint., J= 7.2 Hz),
1.33 (4H, m), 0.89 (3H, t, J=6.8 Hz); 13C NMR (75 MHz,
CDCl3): δ 138.5 (CH), 133.3 (quat.), 133.2 (quat.), 133.0
(CH), 131.5 (CH), 115.4 (CH), 100.5 (CH), 46.9 (CH2), 30.3
(CH2), 29.2 (CH2), 22.4 (CH2), 14.0 (CH3); LRMS (+ESI):
m/z 189 ([M + H]+, 100%); IR (diamond cell, thin flm):
3091 (w), 2956 (s), 2930 (s), 2870 (m), 1682 (m), 1500 (s),
1470 (s), 1320 (s), 1030 (m), 814 (s), 733 (s).
General procedure A: 1‑N‑alkylation of azaindoles
The appropriate azaindole (1.00 g, 8.46 mmol) in DMF
(15 mL) was slowly added to an ice-cold solution of sodium
hydride (60% dispersion in mineral oil, 675 mg, 16.9 mmol)
in DMF (5 mL) and stirred for 10 min. 1-Bromopentane
(1.15 mL, 9.31 mmol) was added, and the mixture was
warmed to ambient temperature and stirred for 1 h. The
reaction mixture was slowly added to H2O (75 mL) and
extracted with EtOAc (3 ×75 mL). The combined organic
extracts were dried (MgSO4) and concentrated under
reduced pressure.
Preparation of 1‑N‑pentyl‑7‑azaindole (31)
Treating 7-azaindole (250 mg, 2.12 mmol) according to gen-
eral procedure A gave 31, following purifcation by fash
chromatography (hexane/EtOAc, 65:35 v/v), as a red oil
(217 mg, 68%).
Preparation of 1‑N‑pentyl‑4‑azaindole (28)
Treating 4-azaindole (1.00 g, 8.46 mmol) according to gen-
eral procedure A gave 28, following purifcation by fash
chromatography (hexane/EtOAc, 70:30 v/v), as an orange
oil (1.28 g, 80%).
1H NMR (300 MHz, CDCl3): δ 8.32 (1H, d, J=3.3 Hz),
7.89 (1H, d, J = 7.8 Hz), 7.22 (1H, m), 7.04 (1H, t,
J=4.8 Hz), 6.44 (1H, m), 4.29 (2H, t, J=7.2 Hz), 1.87 (2H,
1H NMR (300 MHz, CDCl3): δ 8.57 (1H, d, J=5.4 Hz),
8.18 (1H, d, J=8.4 Hz), 7.67 (1H, d, J=3.3 Hz), 7.46 (1H,
dd, J = 8.4, 5.7 Hz), 7.00 (1H, d, J = 3 Hz), 4.26 (2H, t,
J=7.2 Hz), 1.86 (2H, quin., J=7.5 Hz), 1.29 (4H, m), 0.88
(3H, t, J = 7.0 Hz); 13C NMR (75 MHz, CDCl3): δ 138.3
(quat.), 136.8 (CH), 135.3 (CH), 132.1 (quat.), 123.9 (CH),
116.1 (CH), 98.3 (CH), 47.5 (CH2), 30.1 (CH2), 28.9 (CH2),
quint., J=6.9 Hz), 1.26 (4H, m), 0.88 (3H, t, J=6.6 Hz); 13
C
NMR (75 MHz, CDCl3): δ 147.6 (quat.), 142.8 (CH), 128.8
(CH), 128.1 (CH), 120.7 (quat.), 115.6 (CH), 99.3 (CH),
44.7 (CH2), 30.3 (CH2), 29.2 (CH2), 22.5 (CH2), 14.1 (CH3);
LRMS (+ ESI): m/z 189 ([M + H]+, 100%); IR (diamond
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Forensic Toxicology
cell, thin flm): 3051 (w), 2956 (s), 2926 (s), 2858 (m), 1509
([M+Na]+, 100%); IR (diamond cell, thin flm): 2959 (m),
2933 (m), 2864 (w), 1675 (s), 1527 (s), 1394 (m), 1295 (m),
1184 (s), 1141 (s), 1112 (s), 881 (s).
(s), 1425 (s), 1272 (s), 772 (s), 715 (s).
General procedure B: 3‑trifuoroacylation of azain‑
doles
Preparation of 1‑N‑pentyl‑3‑trifuoroacyl‑6‑azaindole (34)
A suspension of aluminium chloride (4.42 g, 33.2 mmol)
and the appropriate 1-N-pentyl azaindole (1.25 g,
6.64 mmol) in DMF (25 mL) was stirred at ambient tem-
perature for 1 h. The reaction was cooled to 0 °C and tri-
fuoroacetic anhydride (1.41 mL, 9.96 mmol) was added
dropwise, warmed to ambient temperature, and stirred
for 5 h. The reaction mixture was slowly added to H2O
(75 mL) and extracted with CH2Cl2 (3 × 75 mL). The
combined organic extracts were dried (MgSO4) and con-
centrated under reduced pressure.
Treating 30 (1.25 g, 6.64 mmol) according to general pro-
cedure B gave 34, following purifcation by fash chroma-
tography (hexane/EtOAc, 55:45 v/v), as a red oil (217 mg,
95%).
m.p. 80.5–82.5 °C; 1H NMR (300 MHz, CDCl3): δ 8.89
(1H, s), 8.53 (1H, d, J=5.1 Hz), 8.25 (1H, d, J=5.4 Hz),
8.01 (1H, s), 4.31 (2H, t, J = 7.2 Hz), 1.97 (2H, quin.,
J = 6.9 Hz), 1.42-1.30 (4H, m), 0.92 (3H, t, J = 6.9 Hz);
13C NMR (75 MHz, CDCl3): δ 174.8 (q, CO, J=35.3 Hz),
143.1 (CH), 139.3 (q, CH, J =4.5 Hz), 133.7 (CH), 133.7
(quat.), 132.6 (quat.), 116.8 (q, CF3, J = 288.8 Hz), 116.7
(CH), 109.1 (quat.), 48.3 (CH2), 29.7 (CH2), 28.9 (CH2),
22.2 (CH2), 13.8 (CH3); 19F NMR (282 MHz, CDCl3): δ
−72.9 (3F, s); LRMS (+ESI): m/z 307 ([M+Na]+, 100%);
IR (diamond cell, thin flm): 3118 (w), 2960 (m), 2938 (m),
2867 (m), 1681 (s), 1606 (s), 1525 (s), 1294 (s), 1188 (s),
1147 (s), 995 (m), 829 (m).
Preparation of 1‑N‑pentyl‑3‑trifuoroacyl‑4‑azaindole (32)
Treating 28 (1.55 g, 7.17 mmol) according to general pro-
cedure B gave 32, following purifcation by fash chroma-
tography (hexane/EtOAc, 70:30 v/v), as an of-white solid
(1.48 g, 78%).
1
m.p. 101.5–103.0 °C; H NMR (300 MHz, CDCl3):
δ 8.75 (1H, d, J = 4.5 Hz), 8.10 (1H, s), 7.76 (1H, d,
J=8.4 Hz), 7.29 (1H, t, J=6.6 Hz), 4.23 (2H, t, J=7.2 Hz),
1.92 (2H, quin., J=6.9 Hz), 1.41–1.32 (4H, m), 0.90 (3H,
t, J = 6.9 Hz); 13C NMR (75 MHz, CDCl3): δ 173.8 (q,
CO, J = 35.3 Hz), 147.1 (CH), 144.8 (quat.), 139.0 (q,
CH, J = 4.5 Hz), 130.0 (quat.), 118.8 (CH), 118.1 (CH),
116.9 (q, CF3, J = 288.8 Hz), 109.4 (quat.), 48.1 (CH2),
29.5 (CH2), 28.9 (CH2), 22.2 (CH2), 13.9 (CH3); 19F NMR
(282 MHz, CDCl3): δ −72.69 (3F, s); LRMS (+ESI): m/z
591 ([2 M+Na]+, 100%); IR (diamond cell, thin flm): 3027
(w), 2956 (m), 2932 (m), 2871 (w), 1674 (s), 1525 (s), 1382
(s), 1282 (s), 1176 (s), 1135 (s), 880 (s), 785 (s), 725 (s).
Preparation of 1‑N‑pentyl‑3‑trifuoroacyl‑7‑azaindole (35)
Treating 31 (200 mg, 1.06 mmol) according to general pro-
cedure B gave 35, following purifcation by fash chroma-
tography (hexane/EtOAc, 80:20 v/v), as a red oil (235 mg,
78%).
1H NMR (300 MHz, CDCl3): δ 8.63 (1H, d, J=7.8 Hz),
8.46 (1H, d, J = 1.5 Hz), 8.07 (1H, s), 7.32 (1H, t,
J = 4.5 Hz), 4.37 (2H, t, J = 7.2 Hz), 1.95 (2H, quint.,
J=6.6 Hz), 1.37 (4H, m), 0.91 (3H, t, J=6.3 Hz); 13C NMR
(75 MHz, CDCl3): δ 148.3 (quat,), 145.7 (CH), 137.3 (q,
CH, J = 4.4 Hz), 131.1 (CH), 119.8 (CH), 119.6 (quat.),
117.0 (q, quat., J = 290.7 Hz), 108.1 (quat.), 46.1 (CH2),
29.8 (CH2), 29.0 (CH2), 22.3 (CH2), 14.0 (CH3), unresolved
CO due to F splitting; 19F NMR (282 MHz, CDCl3): δ −72.5
(3F, s); LRMS (+ESI): m/z 307 ([M+Na]+, 100%); IR (dia-
mond cell, thin flm): 3119 (w), 2958 (w), 2933 (w), 2863
(w), 1671 (s), 1527 (s), 1385 (s), 1128 (s), 1104 (s), 882 (s).
Preparation of 1‑N‑pentyl‑3‑trifuoroacyl‑5‑azaindole (33)
Treating 29 (1.25 g, 6.64 mmol) according to general proce-
dure B gave 33, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 45:55 v/v), as a red oil (1.18 g, 63%).
1H NMR (300 MHz, CDCl3): δ 9.63 (1H, s), 8.55 (1H, d,
J=4.5 Hz), 7.94 (1H, m), 7.34 (1H, d, J=5.7 Hz), 4.21 (2H,
t, J=7.2 Hz), 1.92 (2H, quin., J=7.2 Hz), 1.42–1.31 (4H,
m), 0.92 (3H, t, J=7.1 Hz); 13C NMR (75 MHz, CDCl3): δ
174.8 (q, CO, J=36.3 Hz), 145.8 (CH), 143.9 (CH), 141.0
(quat.), 137.9 (q, CH, J=4.6 Hz), 123.4 (quat.), 116.9 (q,
quat., J=290.8 Hz), 109.8 (quat.), 105.6 (CH), 47.7 (CH2),
29.6 (CH2), 28.9 (CH2), 22.3 (CH2), 13.9 (CH3); 19F NMR
(282 MHz, CDCl3): δ −72.7 (3F, s); LRMS (+ESI): m/z 307
General procedure C: hydrolysis of methyl esters,
trifuoroacetyl and benzyl sulfonyl groups
Aqueous NaOH (4 M, 3.87 mL, 15.5 mmol) was added drop-
wise to a stirred solution of the appropriately substituted
methyl ester, trifuoroacetyl compound, or benzyl sulfo-
nyl-protected indole (2.58 mmol) in MeOH (20 mL), and
allowed to stir at ambient temperature (indazole derivatives)
1 3

Forensic Toxicology
or refux (indole/azaindole derivatives) for 18 h. The reac-
tion mixture was added to H2O (75 mL) and washed with
Et2O (75 mL). The aqueous layer was acidifed to pH ~2
using 1 M aq. HCl and extracted with Et2O (3 × 75 mL). The
combined organic extracts were dried (MgSO4) and concen-
trated under reduced pressure.
22.3 (CH2), 13.9 (CH3); LRMS (+ESI): m/z 255 ([M+Na]+,
100%); IR (diamond cell, thin flm): 3358 (bs), 3120 (w),
2948 (m), 2928 (m), 2864 (m), 1666 (m), 1538 (m), 1373
(m), 1143 (m), 1032 (m), 962 (m), 782 (s), 723 (s), 491 (s).
Preparation of 1‑N‑pentyl‑5‑azaindole‑3‑carboxylic acid
(37)
Preparation of N‑pentyl‑3‑indole carboxylic acid (20)
33 (600 mg, 2.11 mmol) was treated according to general
procedure C. Once the reaction was complete, the reac-
tion mixture was concentrated under reduced pressure and
immediately subjected to fash chromatography (hexane/
EtOAc/acetic acid, 91:8:1 v/v), giving 37 as an of-white
solid (425 mg, 87%).
Treating 19 (450 mg, 1.59 mmol) according to general pro-
cedure C gave 20 as a white solid (323 mg, 88%).
m.p. 106.5–108.0 °C; 1H NMR (300 MHz, MeOD-d4): δ
8.09 (1H, dd, J=6.5, 1.9 Hz), 7.93 (1H, s), 7.42–7.28 (3H,
m), 4.17 (2H, t, J = 7.2 Hz), 1.90 (2H, quin., J = 7.0 Hz),
1.40–1.32 (4H, m), 0.90 (3H, t, J = 6.6 Hz), COOH sig-
nal not observed; 13C NMR (75 MHz, MeOD-d4): δ 168.8
(CO2H), 138.1 (quat.), 136.3 (CH), 128.3 (quat.), 123.6
(CH), 122.6 (CH), 122.4 (CH), 111.3 (CH), 107.7 (quat.),
47.7 (CH2), 30.7 (CH2), 30.0 (CH2), 23.3 (CH2), 14.2 (CH3);
LRMS (– ESI): m/z 230 ([M–H]−, 100%); IR (diamond cell,
thin flm): 3106 (w), 2925 (m), 2856 (w), 2525 (bs), 1649
(s), 1526 (s), 1461 (m), 1273 (m), 1204 (s), 1117 (m), 940
(m), 731 (s).
m.p. 185.0–188.0 °C; 1H NMR (300 MHz, MeOD-d4):
δ 9.36 (1H, s), 8.36 (1H, d, J=6.9 Hz), 8.18 (1H, s), 7.84
(1H, d, J=6.3 Hz), 4.34 (2H, t, J=7.2 Hz), 1.89 (2H, quint.,
J = 7.4 Hz), 1.39-1.28 (4H, m), 0.89 (3H, t, J = 7.1 Hz),
COOH signal not observed; 13C NMR (75 MHz, MeOD-
d4): δ 168.0 (CO2H), 143.3 (quat.), 141.2 (CH), 139.0 (CH),
136.8 (CH), 125.0 (quat.), 113.1 (quat.), 108.5 (CH), 48.1
(CH2), 30.8 (CH2), 29.9 (CH2), 23.3 (CH2), 14.2 (CH3);
LRMS (+ ESI): m/z 233 ([M + H]+, 100%); IR (diamond
cell, thin flm): 3105 (w), 2956 (m), 2859 (w), 2391 (bs),
1677 (w), 1538 (m), 1475 (m), 1186 (s), 1007 (s), 805 (s).
Preparation of 1‑N‑pentyl‑3‑indazole carboxylic acid (23)
Treating 22 (600 mg, 2.58 mmol) according to general pro-
cedure C gave 23 as a white solid (510 mg, 85%).
Preparation of 1‑N‑pentyl‑6‑azaindole‑3‑carboxylic acid
(38)
1
m.p. 76.5–78.0 °C; H NMR (300 MHz, MeOD-d4):
δ 8.26 (1H, d, J = 8.1 Hz), 7.52–7.44 (2H, m), 7.35 (1H,
t, J = 7.8 Hz), 4.48 (2H, t, J = 7.2 Hz), 1.99 (2H, quin.,
J=7.2 Hz), 1.34 (4H, m), 0.89 (3H, t, J=6.0 Hz), COOH
signal not observed; 13C NMR (75 MHz, MeOD-d4): δ 165.4
(CO2H), 142.0 (quat.), 135.9 (quat.), 127.9 (CH), 124.5
(quat.), 124.0 (CH), 122.9 (CH), 111.0 (CH), 49.7 (CH2),
30.4 (CH2), 29.8 (CH2), 23.1 (CH2), 14.1 (CH3); LRMS
(– ESI): m/z 231 ([M–H]−, 100%); IR (diamond cell, thin
flm): 3053 (bs), 2956 (m), 2931 (m), 2860 (w), 1687 (s),
1503 (s), 1218 (s), 1176 (s), 1121 (s), 752 (s).
34 (900 mg, 3.17 mmol) was treated according to general
procedure C. Once the reaction was complete, the reaction
mixture was concentrated under reduced pressure and imme-
diately subjected to fash chromatography (CH2Cl2/MeOH,
94:6 v/v), giving 38 as an of-white solid (385 mg, 52%).
m.p. 159.0–160.5 °C; 1H NMR (300 MHz, MeOD-d4):
δ 8.90 (1H, s), 8.26 (1H, d, J=5.6 Hz), 8.22 (1H, s), 8.13
(1H, d, J=5.7 Hz), 4.37 (2H, t, J=7.1 Hz), 1.92 (2H, quin.,
J = 7.1 Hz), 1.41–1.26 (4H, m), 0.89 (3H, t, J = 7.1 Hz),
COOH signal not observed; 13C NMR (75 MHz, MeOD-
d4): δ 167.9 (CO2H), 140.6 (CH), 139.3 (CH), 135.1 (quat.),
134.5 (CH), 133.5 (CH), 117.5 (quat.), 109.4 (quat.), 48.3
(CH2), 31.0 (CH2), 29.9 (CH2), 23.2 (CH2), 14.2 (CH3);
LRMS (+ ESI): m/z 233 ([M + H]+, 100%); IR (diamond
cell, thin flm): 2930 (w), 2861 (w), 1694 (s), 1479 (s), 1183
(m), 1131 (m), 1029 (s), 822 (m).
Preparation of 1‑N‑pentyl‑4‑azaindole‑3‑carboxylic acid
(36)
Treating 32 (1.48 g, 5.21 mmol) according to general proce-
dure C gave 36 as an of-white solid (1.18 g, 97%).
1
m.p. 112.0–113.5 °C; H NMR (300 MHz, MeOD-
d4): δ 8.48 (1H, d, J = 4.5 Hz), 8.06 (1H, s), 7.78 (1H, d,
J = 8.3 Hz), 7.26 (1H, dd, J = 8.6, 4.8 Hz), 4.18 (2H, t,
J=7.1 Hz), 1.87 (2H, quin., J=7.2 Hz), 1.38–1.24 (4H, m),
0.87 (3H, t, J = 7.1 Hz), COOH signal not observed; 13C
NMR (75 MHz, MeOD-d4): δ 164.4 (CO2H), 144.4 (quat.),
143.7 (CH), 136.1 (CH), 129.2 (quat.), 118.9 (CH), 117.9
(CH), 106.6 (quat.), 47.8 (CH2), 29.8 (CH2), 28.9 (CH2),
Preparation of 1‑N‑pentyl‑7‑azaindole‑3‑carboxylic acid
(39)
Treating 35 (200 mg, 0.70 mmol) according to general pro-
cedure C gave 39 as an of-white solid (135 mg, 83%).
m.p. 143.0–147.5 °C; 1H NMR (300 MHz, MeOD-d4): δ
8.43 (1H, d, J=7.2 Hz), 8.31 (1H, m), 8.23 (1H, m), 7.25
1 3

Forensic Toxicology
(1H, s), 4.33 (2H, t, J=6.0 Hz), 1.88 (2H, quin., J=6.0 Hz),
1.41–1.15 (4H, m), 0.88 (3H, m), COOH signal not
observed; 13C NMR (75 MHz, MeOD-d4): δ 167.9 (CO2H),
148.8 (quat.), 144.5 (CH), 136.5 (CH), 131.3 (CH), 121.0
(quat.), 118.9 (CH), 106.8 (quat.), 46.2 (CH2), 30.9 (CH2),
29.9 (CH2), 23.3 (CH2), 14.3 (CH3); LRMS (–ESI): m/z 231
([M–H)−, 100%); IR (diamond cell, thin flm): 2930 (w),
2856 (w), 1685 (s), 1533 (s), 1257 (s), 1138 (s), 798 (s),
755 (s).
m.p. 66.1–68.8 °C; 1H NMR (300 MHz, CDCl3): δ 8.38
(1H, d, J = 8.1 Hz), 7.38–7.34 (2H, m), 7.27–7.22 (1H,
m), 6.80 (1H, s), 4.34 (2H, t, 6.9 Hz), 2.23–2.18 (6H, m),
2.17–2.10 (3H, m), 1.93 (2H, quin., 6.9 Hz), 1.79–1.69 (6H,
m), 1.40–1.30 (4H, m), 0.89 (3H, t, J =6.3 Hz); 13C NMR
(75 MHz, CDCl3): δ 162.2 (CO), 141.0 (quat.), 138.2 (quat.),
126.6 (CH), 123.3 (CH), 123.0 (quat.), 122.4 (CH), 109.2
(CH), 52.0 (CH2), 49.5 (quat.), 42.1 (CH2), 36.6 (CH2),
29.7 (CH), 29.6 (CH2), 29.1 (CH2), 22.4 (CH2), 12.0 (CH3);
LRMS (+ESI): m/z 388 ([M+Na]+, 100%); HRMS (+ESI):
m/z calculated [M + Na]+ 388.2359, found 388.2360; IR
(diamond cell, thin flm): 3303 (w), 2908 (s), 2846 (m),
1647 (s), 1529 (s), 1491 (m), 1451 (m), 1357 (m), 1341
(m), 1289 (m), 1218 (m), 1139 (m), 1005 (m), 859 (m), 772
(m), 748 (s).
General procedure D: amidation of 1‑N‑pen‑
tyl‑3‑indole/indazole/azaindole carboxylic
acids with adamantylamine hydrochloride
via 1‑ethyl‑3‑(3‑dimethylaminopropyl)carbodiim‑
ide) hydrochloride coupling
N,N-Diisopropylethylamine (340 µL, 1.95 mmol) was added
dropwise to a stirred solution of the appropriate 1-N-pentyl-
3-indole/indazole/azaindole carboxylic acid (0.39 mmol),
adamantylamine hydrochloride (0.41 mmol), 1-ethyl-
3-(3-dimethylaminopropyl)carbodiimide hydrochloride
(EDC·HCl, 150 mg, 0.78 mmol), and 1-hydroxybenzotria-
zole (HOBt, 119 mg, 0.78 mmol) in DMSO (5 mL), and was
stirred at ambient temperature for 18 h. The reaction mixture
was added to sat. aq. NaHCO3 (75 mL) and extracted with
EtOAc (3 × 75 mL). The combined organic extracts were
dried (MgSO4) and concentrated under reduced pressure.
Preparation of N‑(adamantan‑1‑yl)‑1‑pen‑
tyl‑1H‑pyrrolo[3,2‑b]pyridine‑3‑carboxamide (AP4AICA, 8)
Treating 36 (200 mg, 0.86 mmol) according to general pro-
cedure D gave 8, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 70:30 v/v), as a white solid (210 mg,
67%).
m.p. 212.0–214.0 °C; 1H NMR (300 MHz, CDCl3): δ 8.69
(1H, br s), 8.47 (1H, s), 7.97 (1H, s), 7.65 (1H, d, J=8.1 Hz),
7.16 (1H, t, J=3.9 Hz), 4.12 (2H, t, J=6.3 Hz), 2.13 (6H,
m), 2.04 (3H, m), 1.83–1.69 (8H, m), 1.35–1.21 (4H, m),
0.88 (3H, t, J=6.9 Hz); 13C NMR (75 MHz, CDCl3): δ 163.4
(CO), 143.5 (CH), 134.8 (CH), 129.8 (quat.), 127.1 (quat.),
117.6 (CH), 116.9 (CH), 112.0 (quat.), 51.8 (CH2), 47.2
(quat.), 42.2 (CH2), 36.8 (CH2), 29.9 (CH2), 29.8 (CH2),
29.0 (CH2), 22.4 (CH2), 14.0 (CH3); LRMS (+ ESI): m/z
388 ([M + Na]+, 100%); HRMS (+ ESI): m/z calculated
[M + Na]+ 388.2359, found 388.2362; IR (diamond cell,
thin flm): 3257 (w), 3065 (w), 2902 (s), 2849 (m), 1635
(s), 1562 (s), 1432 (m), 1276 (m), 1255 (m), 782 (s).
Preparation of N‑(adamantan‑1‑yl)‑1‑pentyl‑1H‑in‑
dole‑3‑carboxamide (APICA, 4)
Treating 20 (576 mg, 2.50 mmol) according to general pro-
cedure D gave 4, following recrystallisation from isopro-
panol/H2O, as a white solid (787 mg, 86%).
m.p. 140.0–141.0 °C; 1H NMR (500 MHz, CDCl3): δ 7.87
(1H, d, J=7.5 Hz), 7.65 (1H, s), 7.36 (1H, d, J=8.0 Hz),
7.28–7.22 (2H, m), 5.71 (1H, br s, NH), 4.11 (2H, t,
J=7.2 Hz), 2.19 (6H, br s), 2.14 (3H, br s), 1.84 (2H, quin.,
J = 7.3 Hz), 1.76 (6H, m) 1.39–1.24 (4H, m), 0.88 (3H, t,
J = 7.0 Hz); 13C NMR (125 MHz, CDCl3): δ 164.0 (CO),
136.7 (quat.), 131.5 (CH), 125.3 (quat.), 122.3 (CH), 121.2
(CH), 120.0 (CH), 112.3 (quat.), 110.4 (CH), 52.2 (quat.),
46.9 (CH2), 42.4 (CH2), 36.6 (CH2), 29.8 (CH), 29.7 (CH2),
29.1 (CH2), 22.4 (CH2), 14.0 (CH3).
Preparation of N‑(adamantan‑1‑yl)‑1‑pen‑
tyl‑1H‑pyrrolo[3,2‑c]pyridine‑3‑carboxamide (AP5AICA, 9)
Treating 37 (70 mg, 0.30 mmol) according to general pro-
cedure D gave 9, following purifcation by fash chromatog-
raphy (EtOAc), as a white solid (68 mg, 62%).
1
Preparation of N‑(adamantan‑1‑yl)‑1‑pentyl‑1H‑inda‑
zole‑3‑carboxamide (APINACA, 7)
m.p. 155.5–156.5 °C; H NMR (300 MHz, CDCl3): δ
9.24 (1H, s), 8.38 (1H, d, J=5.7 Hz), 7.63 (1H, s), 7.26 (1H,
d, J=6.0 Hz), 5.70 (1H, br s), 4.10 (2H, t, J=7.2 Hz), 2.18
(6H, m), 2.14 (3H, m), 1.84 (2H, quin., J= 7.2 Hz), 1.75
(6H, m), 1.38–1.27 (4H, m), 0.88 (3H, t, J = 7.1 Hz); 13C
NMR (75 MHz, CDCl3): δ 163.4 (CO), 143.4 (CH), 141.6
(CH), 140.5 (quat.), 131.7 (CH), 122.4 (quat.), 113.0 (quat.),
105.4 (CH), 52.2 (CH2), 46.8 (quat.), 42.2 (CH2), 36.6 (CH),
Treating 23 (100 mg, 0.43 mmol) according to general pro-
cedure D gave 7, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 95:5 v/v), as a white solid (128 mg,
81%).
1 3

Forensic Toxicology
29.9 (CH2), 29.7 (CH3), 29.0 (CH2), 22.3 (CH2), 14.0 (CH3);
LRMS (+ESI): m/z 366 ([M+H]+, 100%); HRMS (+ESI):
m/z calculated [M + Na]+ 388.2359, found 388.2362; IR
(diamond cell, thin flm): 3343 (bs), 2905 (m), 2851 (m),
1619 (s), 1537 (s), 1516 (s), 1357 (m), 1196 (m), 1141 (m),
799 (m).
General procedure E: esterifcation of 1‑N‑pen‑
tyl‑3‑indole/indazole/azaindole carboxylic acids
with 1‑bromoadamantane
A suspension of the appropriate 1-N-pentyl-indole, inda-
zole, or azaindole-3-carboxylic acid (0.43 mmol), 1-bro-
moadamantane (101 mg, 0.47 mmol), and silver carbonate
(178 mg, 0.65 mmol) in N,N-dimethylacetamide (4 mL) was
stirred at refux for 24 h. The reaction was cooled to ambi-
ent temperature and concentrated under reduced pressure.
The residue was added to H2O (50 mL) and extracted with
EtOAc (3 × 50 mL), and the combined organic extracts were
dried (MgSO4) and concentrated under reduced pressure.
Preparation of N‑(adamantan‑1‑yl)‑1‑pen‑
tyl‑1H‑pyrrolo[2,3‑c]pyridine‑3‑carboxamide (AP6AICA, 10)
Treating 38 (70 mg, 0.30 mmol) according to general proce-
dure D gave 10, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 80:20 v/v), as a white solid (88 mg,
80%).
1
m.p. 154.0–155.5 °C; H NMR (500 MHz, CDCl3): δ
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑indole‑3‑car‑
boxylate (APIC, 12)
8.79 (1H, d, J = 0.9 Hz), 8.35 (1H, d, J = 5.6 Hz), 7.75-
7.73 (2H, m), 5.67 (1H, br s), 4.19 (2H, t, J = 7.2 Hz),
2.17 (9H, m), 1.88 (2H, quin., J=7.3 Hz), 1.77–1.70 (6H,
m), 1.36–1.27 (4H, m), 0.87 (3H, t, J =7.3 Hz); 13C NMR
(125 MHz, CDCl3): δ 163.6 (CO), 140.5 (CH), 134.1 (CH),
133.8 (CH), 133.7 (quat.), 130.2 (quat.), 114.5 (CH), 112.2
(quat.), 52.4 (CH2), 47.4 (quat.), 42.3 (CH2), 36.6 (CH2),
30.1 (CH2), 29.7 (CH), 29.1 (CH2), 22.4 (CH2), 14.0 (CH3);
LRMS (+ESI): m/z 366 ([M+H]+, 100%); HRMS (+ESI):
m/z calculated [M + Na]+ 388.2359, found 388.2362; IR
(diamond cell, thin flm): 3333 (bs), 2904 (s), 2851 (m),
1618 (s), 1537 (s), 1514 (s), 2180 (m), 1245 (m), 821 (m).
Treating 20 (100 mg, 0.43 mmol) according to general pro-
cedure E gave 12, following purifcation by fash chroma-
tography (hexane/EtOAc, 93:7 v/v), as an of-white solid
(68 mg, 41%).
1
m.p. 102.0–103.0 °C; H NMR (300 MHz, CDCl3): δ
8.21–8.11 (1H, m), 7.75 (1H, s), 7.38–7.19 (3H, m), 4.12
(2H, t, J=7.1 Hz), 2.40–2.31 (6H, m), 2.28–2.19 (3H, m),
1.86 (2H, quin., J=7.3 Hz), 1.80–1.65 (6H, m), 1.44–1.25
(4H, m), 0.89 (3H, t, J = 6.5 Hz); 13C NMR (75 MHz,
CDCl3): 164.7 (CO), 136.7 (quat.), 134.2 (CH), 126.9
(quat.), 122.5 (CH), 122.0 (CH), 121.6 (CH), 110.0 (CH),
108.9 (quat.), 80.1 (quat.), 47.1 (CH2), 42.1 (CH2), 36.5
(CH), 31.1 (CH2), 29.8 (CH2), 29.1 (CH2), 22.4 (CH2), 14.0
(CH3); LRMS: m/z 388 ([M+Na]+, 100%); HRMS (+ESI):
m/z calculated [M + Na]+ 388.2247, found 388.2245; IR
(diamond cell, thin flm): 3112 (w), 2904 (s), 2847 (m), 1680
(s), 1530 (s), 1265 (s), 1102 (s), 1058 (s), 734 (s).
Preparation of N‑(adamantan‑1‑yl)‑1‑pen‑
tyl‑1H‑pyrrolo[2,3‑b]pyridine‑3‑carboxamide (AP7AICA,
11)
Treating 39 (75 mg, 0.32 mmol) according to general proce-
dure D gave 11, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 80:20 v/v), as a white solid (88 mg,
75%).
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑indazole‑3‑car‑
boxylate (APINAC, 13)
1
m.p. 149.0–149.5 °C; H NMR (300 MHz, CDCl3): δ
9.24 (1H, m), 8.26 (1H, d, J = 8.1 Hz), 7.69 (1H, s), 7.17
(1H, t, J=4.5 Hz), 5.55 (1H, br s), 4.29 (2H, t, J=6.6 Hz),
2.17 (9H, m), 1.88 (2H, quin., J= 6.8 Hz), 1.74 (6H, m),
1.33 (4H, m), 0.88 (3H, t, J=6.7 Hz); 13C NMR (75 MHz,
CDCl3): δ 164.0 (CO), 147.8 (quat.), 143.7 (CH), 130.2
(CH), 128.8 (CH), 118.8 (quat.), 117.3 (CH), 110.6 (quat.),
52.3 (CH2), 45.1 (quat.), 42.3 (CH2), 36.6 (CH), 30.1 (CH2),
29.7 (CH), 29.1 (CH2), 22.4 (CH2), 14.1 (CH3); LRMS
(+ ESI): m/z 388 ([M + Na]+, 100%); HRMS (+ ESI): m/z
calculated [M + Na]+ 388.2359, found 388.2362; IR (dia-
mond cell, thin flm): 3322 (bs), 3097 (s), 2905 (s), 2850
(m), 1622 (s), 1538 (s), 1451 (m), 1187 (m).
Treating 20 (500 mg, 1.54 mmol) according to general pro-
cedure E gave 13, following purifcation by fash chroma-
tography (hexane/EtOAc, 92:8 v/v), as an of-white solid
(215 mg, 38%).
1
m.p. 109.0–110.0 °C; H NMR (300 MHz, CDCl3): δ
8.15 (1H, d, J = 8.1 Hz), 7.46–7.37 (2H, m), 7.26 (1H, t,
J = 6.9 Hz), 4.44 (2H, t, J = 7.5 Hz), 2.39–2.37 (6H, m),
2.26–2.23 (3H, m), 1.96 (2H, quin., J=7.2 Hz), 1.80–1.69
(6H, m), 1.39–1.28 (4H, m), 0.88 (3H, t, J = 6.9 Hz); 13C
NMR (300 MHz, CDCl3): δ 161.8 (CO), 140.7 (quat.), 136.3
(quat.), 126.5 (CH), 123.5 (quat.), 122.7(0) (CH), 122.6(9)
(CH), 109.7 (CH), 81.9 (quat.), 49.9 (CH2), 41.8 (CH2),
36.4 (CH), 31.1 (CH2), 29.6 (CH2), 29.1 (CH2), 22.4 (CH2),
14.0 (CH3); LRMS (+ ESI): m/z 389 ([M + Na]+, 100%),
755 ([2 M + Na]+, 95%); HRMS (+ ESI): m/z calculated
1 3

Forensic Toxicology
[M + Na]+ 389.2199, found 389.2196; IR (diamond cell,
thin flm): 2905 (s), 2849 (m), 1690 (m), 1644 (s), 1361
(m), 1305 (s), 1169 (m), 1124 (s), 866 (m), 756 (s), 544 (w).
chromatography (hexane/EtOAc, 50:50 v/v), as a white solid
(68 mg, 43%).
m.p. 108.5–110.0 °C; 1H NMR (300 MHz, CDCl3): δ 8.78
(1H, s), 8.37 (1H, d, J=5.1 Hz), 7.97 (1H, d, J=5.4 Hz),
7.83 (1H, s), 4.21 (2H, t, J = 7.2 Hz), 2.35–2.30 (6H, m),
2.26–2.21 (3H, m), 1.90 (2H, quin., J=6.9 Hz), 1.78–1.67
(6H, m), 1.37–1.30 (4H, m), 0.89 (3H, t, J = 6.3 Hz); 13C
NMR (75 MHz, CDCl3): δ 163.8 (CO), 140.9 (CH), 136.8
(CH), 133.8 (quat.), 133.4 (CH), 131.9 (CH), 116.2 (quat.),
109.0 (quat.), 80.8 (quat.), 47.5 (CH2), 42.0 (CH2), 36.5
(CH2), 31.1 (CH), 30.0 (CH2), 29.0 (CH2), 22.3 (CH2),
14.0 (CH3); LRMS (+ ESI): m/z 367 ([M + H]+, 100%);
HRMS (+ESI): m/z calculated [M+Na]+ 389.2199, found
389.2197; IR (diamond cell, thin flm): 3109 (w), 2909 (s),
2848 (m), 1682 (s), 1603 (m), 1526 (s), 1443 (m), 1249 (s),
1194 (m), 1131 (s), 1055 (s), 820 (s), 626 (m), 424 (m).
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑pyrrolo[3,2‑b]
pyridine‑3‑carboxylate (AP4AIC, 14)
Treating 36 (100 mg, 0.43 mmol) according to general pro-
cedure E gave 14, following purifcation by fash chromatog-
raphy (hexane/EtOAc, 70:30 v/v), as a white solid (55 mg,
35%).
m.p. 83.5–85.0 °C; 1H NMR (300 MHz, CDCl3): δ 8.64
(1H, d, J=4.5 Hz), 7.86 (1H, s), 7.62 (1H, d, J=8.4 Hz),
7.15 (1H, dd, J = 7.8, 4.8 Hz), 4.11 (2H, t, J = 6.9 Hz),
2.34–2.31 (6H, m), 2.22–2.19 (3H, m), 1.85 (2H, quin.,
J = 7.2 Hz), 1.74–1.67 (6H, m), 1.38–1.25 (4H, m), 0.89
(3H, t, J = 6.3 Hz); 13C NMR (75 MHz, CDCl3): δ 162.6
(CO), 145.5 (CH), 144.7 (quat.), 136.1 (CH), 129.7 (quat.),
117.2 (CH), 117.1 (CH), 109.1 (quat.), 80.2 (quat.), 47.2
(CH2), 41.8 (CH2), 36.5 (CH2), 31.1 (CH), 29.8 (CH2),
29.0 (CH2), 22.3 (CH2), 13.9 (CH3); LRMS (+ ESI): m/z
389 ([M + Na]+, 100%); HRMS (+ ESI): m/z calculated
[M + Na]+ 389.2199, found 389.2200; IR (diamond cell,
thin flm): 3107 (w), 2911 (m), 2856 (w), 1661 (s), 1640
(s), 1517 (s), 1329 (s), 1182 (m), 885 (m), 799 (m), 549 (m).
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑pyrrolo[2,3‑b]
pyridine‑3‑carboxylate (AP7AIC, 17)
Treating 39 (100 mg, 0.43 mmol) according to general pro-
cedure E gave 17, following purifcation by fash chromatog-
raphy (hexane/EtOAc 70:30 v/v), as a yellow solid (60 mg,
38%).
m.p. 99.5–101.0 °C; 1H NMR (300 MHz, CDCl3): δ 8.38-
8.34 (2H, m), 7.87 (1H, s), 7.17 (1H, dd, J =8.1, 4.8 Hz),
4.29 (2H, t, J = 7.3 Hz), 2.34–2.29 (6H, m), 2.26–2.20
(3H, m), 1.89 (2H, quin., J= 7.2 Hz), 1.78–1.68 (6H, m),
1.39–1.28 (4H, m), 0.88 (3H, t, J = 7.0 Hz); 13C NMR
(75 MHz, CDCl3): δ 164.0 (CO), 147.9 (quat.), 143.8 (CH),
133.9 (CH), 130.1 (CH), 119.3 (quat.), 117.8 (CH), 107.4
(quat.), 80.5 (quat.), 45.2 (CH2), 42.0 (CH2), 36.5 (CH2),
31.1 (CH), 30.0 (CH2), 29.0 (CH2), 22.4 (CH2), 14.0 (CH3);
LRMS (+ESI): m/z 389 ([M+Na]+, 100%); HRMS (+ESI):
m/z calculated [M + Na]+ 389.2199, found 389.2198; IR
(diamond cell, thin flm): 3105 (w), 2915 (m), 2852 (w),
1686 (s), 1528 (s), 1403 (m), 1256 (s), 1135 (s), 1117 (s),
1058 (s), 773 (m), 758 (s).
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑pyrrolo[3,2‑c]
pyridine‑3‑carboxylate (AP5AIC, 15)
Treating 37 (100 mg, 0.43 mmol) according to general pro-
cedure E gave 15, following purifcation by fash chroma-
tography (hexane/EtOAc, 60:40 v/v), as an of-white solid
(79 mg, 50%).
1
m.p. 101.5–102.5 °C; H NMR (300 MHz, CDCl3): δ
9.56 (1H, d, J=0.9 Hz), 8.52 (1H, d, J=5.8 Hz), 7.72 (1H,
s), 7.33 (1H, dd, J =5.9, 0.8 Hz), 4.19 (2H, t, J =7.1 Hz),
2.31–2.28 (6H, m), 2.11–2.06 (3H, m), 1.90 (2H, quin.,
J = 7.3 Hz), 1.72–1.61 (6H, m), 1.39–1.28 (4H, m), 0.90
(3H, t, J = 7.0 Hz); 13C NMR (75 MHz, CDCl3): δ 162.5
(CO) 145.3 (CH), 143.3 (CH), 141.6 (quat.), 137.8 (CH),
123.0 (quat.), 116.2 (quat.), 105.7 (CH), 74.2 (quat.), 47.5
(CH2), 40.1 (CH2), 36.4 (CH2), 30.1 (CH), 29.5 (CH2), 29.0
(CH2), 22.3 (CH2), 14.0 (CH3); LRMS (+ ESI): m/z 367
([M+H]+, 100%); HRMS (+ESI): m/z calculated [M+Na]+
389.2199, found 389.2199; IR (diamond cell, thin flm):
3109 (w), 2909 (m), 2852 (w), 1683 (s), 1533 (m), 1229
(m), 1132 (s), 1055 (s), 757 (m), 421 (m).
Gas chromatography–mass spectrometry
All fnal compounds were subjected to gas chromatogra-
phy–electron ionisation–mass spectrometry (GC–EI-MS)
using an Agilent 7890 gas chromatograph ftted with a
5975C inert mass selective detector (Agilent Technolo-
gies, Santa Clara, CA, USA). The column was an HP-5
capillary column (30 m × 0.25 mm, i.d., flm thickness
0.25 µm), with helium as the carrier gas at a constant fow
of 1.8 mL/min and a split ratio of 10:1 (Agilent Technolo-
gies). The oven temperature commenced at 65 °C with a
hold time of 1 min, followed by a 40 °C/min ramp rate to
300 °C with a fnal hold time of 13 min. Mass spectra were
collected over m/z 35–550 with an ionisation energy of
Preparation of adamantan‑1‑yl 1‑pentyl‑1H‑pyrrolo[2,3‑c]
pyridine‑3‑carboxylate (AP6AIC, 16)
Treating 38 (100 mg, 0.43 mmol) according to gen-
eral procedure E gave 16, following purifcation by fash
1 3

Forensic Toxicology
70 eV. Compounds were extracted in EtOAc at an approxi-
mate concentration of 1 mg/mL. The injection volume was
1 µL.
50% of that of CP 55,940 (1 µM; Cayman Chemical, Ann
Arbor, MI, USA) when tested at 10 µM. If drugs produced a
response less than 50% of CP 55,940 (1 µM) when tested at
10 µM, experiments were repeated at 10 µM only.
In vitro pharmacological assessment
Data were analysed with PRISM (GraphPad Software
Inc., San Diego, CA, USA), using four-parameter nonlinear
regression to ft concentration-response curves. In all plates,
a maximum efective concentration of CP 55,940 (1 µM) was
added to allow for a ready comparison between experiments.
Mouse AtT-20 neuroblastoma cells stably transfected
with human CB1 or human CB2 have been described pre-
viously and were cultured in Dulbecco’s modifed Eagle
medium containing 10% fetal bovine serum (FBS), 100 U
penicillin/streptomycin, and 80 μg/mL hygromycin [10].
Cells were passaged at 80% confuence as required. Cells
for assays were grown in 75 cm2 fasks and used at 90%
confuence. The day before, the assay cells were detached
from the fask with trypsin/ethylenediaminetetraacetic acid
(Sigma-Aldrich) and resuspended in 10 mL of Leibovitz’s
L-15 media supplemented with 1% FBS, 100 U penicillin/
streptomycin, and 15 mM glucose for the below membrane
potential and Ca5 calcium assays. The cells were plated in
a volume of 90 μL in black-walled, clear-bottomed 96-well
microplates (Corning Inc., Corning, NY, USA). Cells were
incubated overnight at 37 °C in ambient CO2.
Results and discussion
Preparation of APICA (4) is outlined in Fig. 3, and fol-
lowed the same methodology as that previously reported
[13]. Indole (18) was subjected to an N-pentyl alkylation
with 1-bromopentane, followed by the addition of trif-
luoroacetic anhydride, to give 19 in good yield. Hydroly-
sis of the trifuoroacetate functionality under basic condi-
tions gave carboxylic acid 20, before HOBt/EDC-mediated
amide coupling with 1-aminoadamantane hydrochloride
gave 4. Synthesis of the adamantyl ester derivative APIC
(12) has not been reported previously, and required optimi-
sation to obtain desirable amounts of material (full detail
of attempts can be found in supplementary material).
Given the bulky nature and relatively poor nucleophi-
licity of 1-hydroxyadamantane, traditional esterifcation
conditions proved to be an inefective way to access 12.
Instead, halide abstraction of 1-bromoadamantane using
silver(I) carbonate to form the tertiary carbocation in the
presence of carboxylic acid 20 gave the desired product
12 in moderate yield.
Membrane potential was measured using a fuorometric
imaging plate reader (FLIPR) membrane potential assay kit
(blue) from Molecular Devices (San Jose, CA, USA), as
described previously [28]. The dye was reconstituted with
assay buffer of the following composition (mM): NaCl
145, HEPES 22, Na2HPO4 0.338, NaHCO3 4.17, KH2PO4
0.441, MgSO4 0.407, MgCl2 0.493, CaCl2 1.26, glucose 5.56
and bovine serum albumin (BSA) (pH 7.4 and osmolarity
315 5). Prior to the assay, 90 μL/well of the dye solution
was added, giving an initial assay volume of 180 μL/well.
Drugs were dissolved at 30 mM in DMSO and aliquoted
for storage at −30 °C. On the day of experiment, an aliquot
was unfrozen, and a portion diluted to 10 mM in DMSO.
Serial dilutions were performed in HEPES-bufered saline
(HBS) containing 0.1% BSA. The frst 1:10 dilution was
made into HBS plus BSA and 1% DMSO. Drugs were added
to the cells at 10 times the fnal concentration, to give a fnal
concentration of DMSO of 0.1% in all wells.
Synthesis of APINACA (7) also followed a methodol-
ogy described previously (Fig. 4) [10]. N-Alkylation of
methyl-3-indazole carboxylate (21) using potassium tert-
butoxide gave predominantly the 1-N-alkyl regioisomer
(22). Following hydrolysis of the methyl ester to give car-
boxylic acid 23, HOBt/EDC·HCl-mediated amide coupling
with 1-aminoadamantane hydrochloride gave 7 in good
yield. Synthesis of APINAC (13) utilised the optimised
conditions employed in the preparation of 12, using 23
and 1-bromoadamantane to obtain the desired product in
a sufcient quantity.
Plates were incubated at 37 °C at ambient CO2 for
60 min. Fluorescence was measured using a FlexStation 3
(Molecular Devices) microplate reader, with cells excited at
a wavelength of 530 nm and emission measured at 565 nm.
Baseline readings were taken every 2 s for at least 2 min, at
which time either drug or vehicle was added in a volume of
20 μL. The background fuorescence of cells without dye or
dye without cells was negligible. Changes in fuorescence
were expressed as a percentage of predrug fuorescence after
subtraction of the changes produced by vehicle addition.
Drugs were initially tested over a concentration range of
1 nM to 30 µM. Full concentration response curves were
constructed for drugs which produced an efect greater than
The azaindole-derived SCs were accessed through a
synthetic approach similar to that outlined for 4 and 12.
The appropriate azaindoles (24 to 27) were 1-N-pentylated
with 1-bromopentane to give 28–31. Attempts to introduce
the trifuoromethyl ketone in a one-pot reaction with the
N-alkylation proved unsuccessful. This was attributed to
the presence of the additional nitrogen in the bicyclic ring
system, decreasing the nucleophilicity of the azaindole’s
3-position. Friedel-Crafts acylation conditions using alu-
minium chloride and trifluoroacetic anhydride proved
1 3

Forensic Toxicology
Fig. 3 Preparation of 4 and 12. Reagents and conditions: (a)(i) NaH,
1-bromopentane, N,N,-dimethylformamide (DMF), 0–25 °C, 1 h; (ii)
(CF3CO)2O, 25 oC, 1 h, 94%; (b) 4 M aq. NaOH, methanol (MeOH),
refux, 18 h, 88%; (c) 1-aminoadamantane hydrochloride, 1-hydroxy-
benzotriazole (HOBt), 1-ethyl-3-(3-dimethylaminopropyl)carbodiim-
ide hydrochloride (EDC-HCl), N,N-diisopropylethylamine (DIPEA),
dimethyl sulfoxide (DMSO), 25 °C, 18 h, 86%; (d) 1-bromoadaman-
tane, Ag2CO3, N,N-dimethylacetamide (DMA), refux, 24 h, 41%
Fig. 4 Preparation of
7
and 13. Reagents and conditions: (a)
noadamantane, HOBt, EDC·HCl, DIPEA, DMSO, 25 °C, 18 h, 81%;
1-bromopentane, potassium tert-butoxide, tetrahydrofuran, 0–25 °C,
(d) 1-bromoadamantane, Ag2CO3, DMA, refux, 24 h, 38%
48 h, 84%; (b) 4 M aq. NaOH, MeOH, refux, 18 h, 85%; (c) 1-ami-
1 3

Forensic Toxicology
Fig. 5 Preparation of 8–11 and 14–17. Reagents and conditions:
(a) 1-bromopentane, NaH, DMF, 0–25 °C, 2 h, 68–85%; (b) AlCl3,
(CF3CO)2O, DMF, 25 °C, 2 h, 63–95%; (c) 4 M aq. NaOH, MeOH,
refux, 18 h, 52–97%; (d) 1-adamantylamine HCl, EDC·HCl, HOBt,
DIPEA, DMSO, 25 °C, 18 h, 62–80%; (e) 1-bromoadamantane,
Ag2CO3, DMA, refux, 24 h, 35–50%
successful, giving desired compounds 32–35 in good to
excellent yield. Hydrolysis of the trifuoroacetate moie-
ties to give carboxylic acids 36–39 was followed by the
aforementioned amide coupling conditions to give 8–11,
or esterifcation conditions to give 14–17 (Fig. 5).
azaindole-derived carboxylates, the parent ion fragment
was one of the major peaks recorded, whereas it was only a
minor peak observed for the 13. The other major fragmenta-
tion peaks observed within this series include m/z 135/231
corresponding to cleavage of the adamantyl ring (the base
peak for 13), m/z 215 corresponding to cleavage of the C–O
ester bond (the base peak for 14), and m/z 145, presumably
corresponding to the protonated state of the N-alkyl and
C–O ester bond cleaved ion.
Representative total ion chromatograms (TICs) and EI
mass spectra for carboxamides APINACA (7) and AP7A-
ICA (11) are shown in Fig. 6, and carboxylates APINAC
(13) and AP7AIC (17) in Fig. 7 (all other chromatograms
and mass spectra can be found in the supplementary data).
The GC method used was able to reliably diferentiate each
compound within the carboxamide series based exclu-
sively on retention times, and although each derivative
showed similar EI fragmentation patterns, the intensities
of each fragment ion could be used as an additional source
of identifcation. As demonstrated in Fig. 6, each of the
carboxamides showed the parent ion (m/z 365) and a base
peak at m/z 215 corresponding to fragmentation of the
C–N amide bond. The carboxamide series also showed a
tendency to cleave along the N-pentyl chain (e.g., APIN-
ACA m/z 294 and 336; and AP7AICA m/z 308).
The in vitro activity of compounds 4 and 7–17 at CB1 and
CB2 were compared to known agonist CP 55,940 (3) using
a FLIPR assay (summary of results displayed in Table 1,
n=5 or more independent experiments) following the same
methodology as that previously reported [5–8]. Previously
identifed SCs APICA (4) and APINACA (7) showed moder-
ate potency at both hCB1 (EC50 =109 and 142 nM, respec-
tively) and hCB2 (EC50 =56 and 141 nM, respectively), with
4 showing a similar preference for activating CB2, as previ-
ously reported [29]. Unlike previous series of SCs based
on the indole scafold, a number of compounds in the cur-
rent series showed no activity at one or both cannabinoid
receptors, indicating the importance of the bicyclic ring
system to cannabimimetic activity. There was also no cor-
relation between the activity of the amide series and the
ester series. An example of this is known SC APICA (4),
which had EC50 values of 109 and 56 nM at CB1 and CB2,
respectively. Its ester counterpart APIC (12) showed no
activities at the tested dosages at either cannabinoid recep-
tor. The opposite was seen with the 4-azaindole derivatives.
Similar to the carboxamide series, each azaindole scaf-
fold in the carboxylate series was able to be diferentiated
by retention time using the GC method described. However,
APINAC (13) and AP7AIC (17) showed nearly identical
retention times (see Fig. 7). The EIMS fragmentation ratios
for the carboxylate analogues showed greater variability
than their carboxamide counterparts, providing a more
efective way to discriminate each structure. With all of the
1 3

Forensic Toxicology
Fig. 6 Gas chromatography–mass spectrometry (GC–MS) analyses showing the total ion chromatograms (TICs, time recorded in min, top) and
electron-ionisation mass spectra (EIMS, bottom) for APINACA (7, a) and AP7AICA (11, b)
1 3

Forensic Toxicology
Fig. 7 GC–MS analyses showing the TICs (time recorded in min, top) and EIMS (bottom) for APINAC (13, a) and AP7AIC (17, b)
Whereas AP4AICA (8) showed no efcacy at either CB1 or
CB2, AP4AIC (14) showed moderate activity at both CB1
and CB2 (EC50 = 160 and 64 nM, respectively). Based on
these data, it is difcult to gauge any trend in azaindole and
ester tolerability on cannabimimetic activity. Interestingly,
a general trend for CB2 selectivity was seen with this series
of compounds, indicating that a large aliphatic substituent
at the 3-position of these bicyclic ring systems has enhanced
activity at CB2. The most notable of these was AP7AICA
1 3

Forensic Toxicology
Table 1 Functional activities of CP 55,940 (3) and synthetic cannabinoids 4 and 7–17 at hCB1 and hCB2 receptors
Compound
hCB1
hCB1
hCB2
hCB2
CB1 sel.a
pEC50 SEM (EC50, nM)
Max SEM (% CP pEC50 SEM (EC50, nM)
55,940)
Max SEM (%CP
55,940)
CP 55,940 (3)
APICA (4)
7.71 0.05 (19)
6.97 0.07 (109)
6.85 0.05 (142)
–
7.54 0.05 (29)
7.25 0.10 (56)
6.85 0.13 (141)
–
1.5
0.5
1.0
–
–
0.2
–
102
106
–
–
96
–
4
3
89
87
–
6
6
APINACA (7)
AP4AICA (8)
AP5AICA (9)
AP6AICA (10)
AP7AICA (11)
APIC (12)
APINAC (13)
AP4AIC (14)
AP5AIC (15)
AP6AIC (16)
AP7AIC (17)
b
b
–
–
–
–
b
b
–
5.99 0.09 (1022)
7
6
6.80 0.25 (158)
7.70 0.10 (20)
87
94
–
80
78
–
9
4
b
–
–
b
b
–
–
–
5.83 0.19 (1486)
6.80 0.04 (160)
6.00 0.05 (998)
6.15 0.06 (707)
6.25 0.06 (562)
98
104
104
85
97
6.72 0.10 (190)
7.19 0.08 (64)
4
4
0.1
0.4
–
0.4
0.5
2
4
b
–
3
4
6.56 0.10 (258)
6.58 0.09 (262)
72
87
4
5
SEM standard error of the mean
aCB1 selectivity is expressed as the ratio of CB2 EC50 to CB1 EC50
bEC50 >10,000 nM
mmons.org/licenses/by/4.0/), which permits unrestricted use, distribu-
tion, and reproduction in any medium, provided you give appropriate
credit to the original author(s) and the source, provide a link to the
Creative Commons license, and indicate if changes were made.
(11), which showed potent activity at CB2 (EC50 =20 nM),
with no activity at CB1 in this FLIPR assay.
Conclusions
This work explored the synthesis, forensic analysis, and can-
nabimimetic tolerance of a series of adamantyl-containing
SCs, incorporating a range of bicyclic cores and ester/amide
linkages. For the frst time, the importance of the bicyclic
ring system to cannabinoid activity within the aminoalky-
lindole core structure has been illustrated. As there is no
consistent structure-activity relationship within each scaf-
fold, it is difcult to predict whether these bicyclic cores
will readily appear in future drugs of abuse. However, the
receptor selectivity for various analogues within these series
may be of interest in the development of therapeutics.
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