The Oxidation of Phytocannabinoids to Cannabinoquinoids

Article abstract of DOI:10.1021/acs.jnatprod.9b01284

Spurred by a growing interest in cannabidiolquinone (CBDQ, HU-313, 2) as a degradation marker and alledged hepatotoxic metabolite of cannabidiol (CBD, 1), we performed a systematic study on the oxidation of CBD (1) to CBDQ (2) under a variety of experimental conditions (base-catalyzed aerobic oxidation, oxidation with metals, oxidation with hypervalent iodine reagents). The best results in terms of reproducibility and scalability were obtained with λ⁵-periodinanes (Dess-Martin periodinane, 1-hydroxy-1λ⁵,2-benziodoxole-1,3-dione (IBX), and SIBX, a stabilized, nonexplosive version of IBX). With these reagents, the oxidative dimerization that plagues the reaction under basic aerobic conditions was completely suppressed. A different reaction course was observed with the copper(II) chloride-hydroxylamine complex (Takehira reagent), which afforded a mixture of the hydroxyiminodienone 11 and the halogenated resorcinol 12. The λ⁵-periodinane oxidation was general for phytocannabinoids, turning cannabigerol (CBG, 18), cannabichromene (CBC, 10), and cannabinol (CBN, 19) into their corresponding hydroxyquinones (20, 21, and 22, respectively). All cannabinoquinoids modulated to a various extent peroxisome proliferator-activated receptor gamma (PPAR-γ) activity, outperforming their parent resorcinols in terms of potency, but the iminoquinone 11, the quinone dimers 3 and 23, and the haloresorcinol 12 were inactive, suggesting a specific role for the monomeric hydroxyquinone moiety in the interaction with PPAR-γ.

Full text of DOI:10.1021/acs.jnatprod.9b01284

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The Oxidation of Phytocannabinoids to Cannabinoquinoids
Diego Caprioglio, Daiana Mattoteia, Federica Pollastro, Roberto Negri, Annalisa Lopatriello,
Giuseppina Chianese, Alberto Minassi, Juan A. Collado, Eduardo Munoz, Orazio Taglialatela-Scafati,*
and Giovanni Appendino*
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ABSTRACT: Spurred by a growing interest in cannabidiolqui-
none (CBDQ, HU-313, 2) as a degradation marker and alledged
hepatotoxic metabolite of cannabidiol (CBD, 1), we performed a
systematic study on the oxidation of CBD (1) to CBDQ (2) under
a variety of experimental conditions (base-catalyzed aerobic
oxidation, oxidation with metals, oxidation with hypervalent iodine
reagents). The best results in terms of reproducibility and
scalability were obtained with λ5-periodinanes (Dess-Martin
periodinane, 1-hydroxy-1λ5,2-benziodoxole-1,3-dione (IBX), and
SIBX, a stabilized, nonexplosive version of IBX). With these
reagents, the oxidative dimerization that plagues the reaction under
basic aerobic conditions was completely suppressed. A different
reaction course was observed with the copper(II) chloride-
hydroxylamine complex (Takehira reagent), which afforded a mixture of the hydroxyiminodienone 11 and the halogenated
resorcinol 12. The λ5-periodinane oxidation was general for phytocannabinoids, turning cannabigerol (CBG, 18), cannabichromene
(CBC, 10), and cannabinol (CBN, 19) into their corresponding hydroxyquinones (20, 21, and 22, respectively). All
cannabinoquinoids modulated to a various extent peroxisome proliferator-activated receptor gamma (PPAR-γ) activity,
outperforming their parent resorcinols in terms of potency, but the iminoquinone 11, the quinone dimers 3 and 23, and the
haloresorcinol 12 were inactive, suggesting a specific role for the monomeric hydroxyquinone moiety in the interaction with PPAR-γ.
olor development has played a significat role in the early
Cstudies on Cannabis (Cannabis sativa L.) and cannabi-
noids. Thus, the first phytocannabinoids were purified from
Cannabis red oil, a deep-red high-vacuum distillation fraction
of Cannabis extracts.1,2 A red-purple color was also observed
when fiber hemp or hashish was treated with methanolic
KOH.3 Under these conditions, the development of a color is
specific for Cannabis and Cannabis-derived products (mar-
ijuana, hashish),4 and the reaction has long been proved as an
expeditious method for their identification in a forensic context
(Beam test).4
The nature of the pigment from Cannabis red oil is still
unclear, but color formation in the Beam test is the result of
the aerobic oxidation of cannabidiol (CBD, 1) to the
hydroxyquinone 2 (cannabidiolquinone, CBDQ, HU-331),5 a
compound that has attracted considerable interest because of
its selective anticancer activity6,7 and catalytic inhibitory
properties on topoisomerase IIα.8 While development of 2 as
a drug was abandoned, possibly because of unfavorable
stability properties (vide infra) and cellular toxicity,9 distinct
lines of research rekindled interest in this compound. Thus,
microsomial formation of 2 from CBD(1) has been associated
with P450 covalent inhibition and perturbation of hepatic
xenobiotics metabolism,10 and a similar process could also
underlie the liver toxicity reported for high dosages of CBD.11
Furthermore, 2 is formed during long-term storage of CBD
under aerobic conditions,12 and its availability is therefore
important for quality control of this active pharmaceutical
ingredient (API).
Despite the convergence of interest for CBDQ (2) from
various areas of cannabinoid research, its only reported
synthesis is the one inspired by the Beam test, that is, the
aerobic oxidation of CBD in a cooled biphasic petroleum
ether/5% ethanolic KOH system.5,6 Under these conditions,
yields are erratic, scale-dependent, and modest (ca. 20% at
best),5,6 while significant amounts of the dimeric quinone 3 are
also formed by oxidative dimerization of CBDQ.5 Both
reaction products, especially 3, are unstable and rapidly turn
into a complex mixture of polar compounds.5 In our hands, the
oxidation reaction was poorly reproducible and could not be
Received: December 31, 2019
© XXXX American Chemical Society and
American Society of Pharmacognosy
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(tert-butyl hydroperoxide (TBHP), basic H2O2), with
significant amounts of the dimer 3 being always formed
under basic conditions or during the long reaction times
required to achieve a significant conversion. A surprising and
notable exception was the behavior of the Takehira complex
(CuCl2-hydroxylamine),14 which afforded a mixture of the
hydroxyiminodienone 11 and the chlororesorcinol 12. The
regioselectivity of the formation of 11 was deduced from the
diagnostic 3J heteronuclear multiple bond correlation
(HMBC) cross-peaks of H-1″ with the hydroxyiminocarbonyl
carbon.
scaled up over a few hundred milligrams of starting material,
even when air or 80% oxygen was bubbled into the biphasic
reaction system. A more reproducible behavior was observed
with KH or LiH in tetrahydrofuran (THF) or toluene under
heterogeneous conditions, but scale-up was still problematic.
While Beam-type oxidation strategies were eventually aban-
doned, their mechanism is worth mentioning. Thus, the
reaction is presumably triggered by formation of a phenolate
anion, next oxidized to an electrophilic radical (4) that adds to
dioxygen to form a hydroperoxy radical. The latter is reduced
to the corresponding anion (5) by a second phenolate ion,
and, after tautomerization to 6, the hydroperoxy anion is
trapped by the para-carbonyl group. This generates the
bridged keto-peroxyhemiacetal 7, whose α-deprotonation
triggers cleavage of the peroxidic bond, eventually affording
the hydroxylated quinone 2 via the hydrate 8 (Figure 1).
The Takehira complex was originally developed for the
oxidation of methylpolyphenols to their corresponding
hydroxyquinones,14 a reaction of relevance for the industrial
synthesis of vitamin E,14 and was later modified by
replacement of hydroxylamine with other nitrogen bases.15
In control experiments, copper(II) chloride alone gave CBDQ
(2) and the dimer 3 as the only reaction products, while the
quinone 2 did not react with hydroxylamine, suggesting a role
for hydroxylamine in the chemoselective halogenation reaction,
possibly via the generation of an N-chlorinated species, and of
copper(II) in the activation of the quinonecarbonyl carbon
toward nucleophilic attack by hydroxylamine.
Hypervalent iodine derivatives have become increasingly
popular for a wide range of oxidative reactions,16 and
bis(trifluoroacetoxy)iodobenzene (BTIB) was reported to
oxidize the mono-O-alkylated cannabinoid Δ9-THC (9),
otherwise unreactive in Beam-type oxidations,6 to its
corresponding hydroxyquinone.6 This λ3-iodane was also able
to oxidize CBD to CBDQ, but λ5 iodanes like 2-iodoxybenzoic
acid (1-hydroxy-1λ5,2-benziodoxole-1,3-dione, IBX, 13)17 and
the Dess-Martin periodinane (DMP)18 gave much better and
more reproducible yields, with a stabilized and not explosive
version of IBX (SIBX)19 emerging as the reagent of choice.
The superior behavior of SIBX compared to IBX might be
related to the acidity of the stabilizing matrix (isophthalic and
benzoic acids), which could help the hydrolytic cleavage of
iodic esters formed in the reaction.19
The oxidation is presumably initiated by the sigmatropic
rearrangement of the iodine−oxygen bond in the mixed λ5
iodane ester 14 formed by interaction of IBX and the C-1′
phenolic hydroxy group (Figure 2). The resulting C-2′ λ3-
quinol 15, after oxidation to the corresponding λ5-iodane 16, is
transformed by [3.3]-sigmatropic rearrangement of the
carbon−oxygen bond into the C-4′ λ5-iodinane 17, with β-
elimination eventually generating the hydroxyquinone 2 and a
reduced λ-iodinane. Remarkably, dimerization was completely
suppressed under iodinane oxidation, and yields in the range of
50−60% could be obtained at multigram reaction scale.
Figure 1. Possible mechanism of the base-mediated aerobic formation
of cannabidiolquinone (CBDQ (2)) from cannabidiol (CBD (1)) in
ethanolic KOH. (R = 3-p-mentha-1,8-dienyl).
This process is reminiscent of the transformation of vitamin
K hydroquinone into its epoxyquinone form,13 and the
mechanism outlined in Figure 1 could explain the sensitivity
of the reaction to radical traps like butylated hydroxytoluene
(BHT) as well as the unreactivity of monoalkylated
phytocannabinoids, like Δ9-tetrahydrocannabinol (Δ9-THC,
9) and cannabichromene (CBC, 10), where the prototropic
equilibrium required for the formation of the peroxyhemiacetal
is not possible (cf. the formation of 6 from 5 in Figure 1).
The reaction profile of the Beam test was basically
replicated, without any substantial improvement of yield, by
metal oxidants [FeCl3, K3[Fe(CN)6], MnO2, Cr6+-based
reagents, CuCl, CuCl2, Ag2O, NH4Ce(NO3)5] under both
catalytic and stoichiometric conditions, as well as by peroxides
B
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a
Table 1. PPAR-γ Modulation Activity
compound
EC50
1
>25 μM
>25 μM
15.7 μM
>25 μM
10.5 μM
14.7 μM
4.9 μM
10
18
19
2
21
20
22
23.1 μM
a
PPAR-γ modulation activity of the phytocannabinoids 1, 10, 18, and
19 and their corresponding cannabinoquinones (2, 21, 20, and 22).
Rosiglitazone (1 μM) was used as positive control for PPAR-γ
activation (50-fold induction over basal activity).
Figure 2. Possible mechanism for the SIBX-mediated formation of
cannabidiolquinone (CBDQ (2)) from cannabidiol (CBD (1)) (R =
3-p-mentha-1,8-dienyl, R′ = n-pentyl).
quinones are axially chiral, and, since enantiomeric cannabi-
noids can show markedly different profiles of bioactivity,2 the
one from CBG (CBGQ (23)) was resolved by chromatog-
raphy on a chiral-phase column packed with amylose-tris(5-
chloro-2-methylphenylcarbamate). However, both the (aR)
and the (aS) enantiomers turned out to be inactive.23
Similarly, the hydroxyiminodienone 11 and the chlorinated
resorcinol 12 were also devoid of activity.23
In conclusion, we have developed a reproducible and
scalable synthesis of cannabinoquinoids, including CBDQ (2),
significantly enhancing access to this compound24 of relevance
not only for its bioactivity profile but also for the analytics of
CBD, the study of its binding to P450 apoproteins, and its
effects on liver function.
CBDQ, an orange powder,20 is unstable in solution, rapidly
degrading in both protic (methanol) and aprotic (acetone,
CHCl3) solvents, with generation of the more polar dimer 3
next to a host of uncharacterized more polar products. On the
other hand, it could be stored for at least 10 months as a
powder at −18 °C in a sealed flask, or for additional time as a
frozen benzene or dimethyl sulfoxide (DMSO) solution at 4
°C.21
The oxidation with SIBX is general for phytocannabinoids,
and, apart from cannabigerol (18), it could also be applied to
monoetherified compounds [cannabichromene (CBC, 10),
cannabinol (19)] that are unreactive under Beam-test
conditions, to afford their corresponding hydroxyquinones
20−22.
EXPERIMENTAL SECTION
■
General Experimental Procedures. IR spectra were recorded
on an Avatar 370 FT-IR Techno-Nicolet apparatus. 1H (400 and 500
MHz) and 13C (100 and 125 MHz) NMR spectra were measured on
Varian INOVA NMR spectrometers. Chemical shifts were referenced
to the residual solvent signal (methanol-d4: δH = 3.34, δC = 49.0 or
1
CDCl3: δH = 7.21, δC = 77.0). Homonuclear H connectivities were
determined by the correlated spectroscopy (COSY) experiment. One-
1
bond heteronuclear H−13C connectivities were determined with the
heteronuclear single quantum coherence (HSQC) spectroscopy
experiment. Two- and three-bond 1H−13C connectivities were
determined by gradient two-dimensional (2D) heteronuclear multiple
bond correlation (HMBC) experiments optimized for a 2,3J = 9 Hz.
Low- and high-resolution electrospray ionization mass spectrometry
(ESI-MS) data were determined on an LTQ OrbitrapXL (Thermo
Scientific) mass spectrometer.
Reactions were monitored by thin-layer chromatography (TLC) on
Merck 60 F254 (0.25 mm) plates, visualized by staining with 5%
H2SO4 in EtOH and heating. Organic phases were dried with Na2SO4
before evaporation. Chemical reagents and solvents were purchased
form Sigma-Aldrich and were used without further purification unless
stated otherwise. Petroleum ether with boiling point of 40−60 °C was
used. Silica gel 60 (70−230 mesh) was used for gravity column
chromatography (GCC).
SIBX Oxidation of Phytocannabinoids. Reaction with CBD
(1) as Example. To a cooled (ice bath) solution of CBD (5 g, 15,6
mmol) in ethyl acetate (EtOAc, 75 mL), SIBX (21.1 g, 31.5 mmol, 2
molar equiv) was addd in six portions of ca. 5 g each. The cooling
bath was removed, and the suspension was stirred at room
temperature for 18 h and then filtered over a pad of diatomaceous
earth. The filtration cake was washed with EtOAc (50 mL), and the
pooled filtrates were washed with saturated Na2S2O3 (4 × 75 mL) and
next with brine. After the drying and evaporation, the residue was
purified by GCC on silica gel (75 g, petroleum ether−EtOAc 9:1 as
eluant) to obtain a brown oil that solidified upon storing in the
refrigerator. Washing with cold petroleum ether removed some of the
CBDQ (2) has been reported to be non-narcotic6 and lacks
significant affinity for CB1 and CB2 receptors.9 Nevertheless, it
showed powerful modulating activity on peroxisome prolifer-
ator-activated receptor gamma (PPAR-γ),9,22 and various
degrees of PPAR-γ activating activity were also shown by the
other cannabinoquinoids (Table 1). However, dimerization
was detrimental for activity, and dimeric quinones were devoid
of significant activity in PPARγ-activity assays.23 Dimeric
C
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colored impurities and afforded an orange powder (3.17 g, 61%). The
same protocol was used for the oxidation and the purification of the
other phytocannabinoids investigated (CBC, 10; CBG, 18; CBN, 19).
The scale was 100−200 mg, and the yields were 59, (CBCQ, 21), 37
(CBGQ, 20), and 58%, (CBNQ, 22).
H-5), 1.65 (3H, s, H-7), 1.63 (3H, s, H-10), 1.60 (2H, overlapped, H-
2′′), 1.34 (2H, overlapped, H-3′′), 1.33 (2H, overlapped, H-4′′), 0.91
(3H, t, J = 6.9 Hz, H-5″); 13C NMR (methanol-d4, 100 MHz) δ 176.3
(C-1′), 168.1 (C-5′), 150.2 (C-8), 148.7 (C-3′), 148.1 (C-4′), 133.6
(C-1), 125.6 (C-2), 120.6 (C-2′), 119.1 (C-6′), 110.8 (C-9), 45.6 (C-
4), 36.2 (C-3), 32.8 (C-3″), 31.6 (C-6), 31.5 (C-1′′), 30.8 (C-5),
30.5 (C-2′′), 23.6 (C-7), 23.5 (C-4′′), 19.1 (C-10), 14.3 (C-5′′); ESI-
MS m/z 344 [M + H]+; HR ESI-MS m/z 344.2210 [M + H]+ calcd
for C21H30NO3, 344.2220.
2-Chlorocannabidiol (12). Yellow oil, IR νmax (KBr disc): 3500,
3421, 2962, 2924, 2859, 1623, 1421, 1258, 1193, 1054, 888, 817, 698
cm−1; 1H NMR (methanol-d4, 400 MHz): δ 6.21 (1H, s, H-2′), 5.25
(1H, s, H-2), 4.46 (1H, s, H-9a), 4.44 (1H, s, H-9b), 3.99 (1H, m, H-
3), 2.95 (1H, m, H-4), 2.56 (2H, t, J = 7.4 Hz, H-1″), 2.20 (1H, m,
H-5a), 2.01 (1H, d, J = 17.1 Hz, H-5b), 1.76 (2H, m, H-6), 1.68 (3H,
s, H-7), 1.65 (3H, s, H-10), 1.56 (2H, m, H-2″), 1.35 (4H, m, H-3′′-
4′′), 0.91 (3H, t, J = 6.8 Hz, H-5″); 13C NMR (methanol-d4, 100
MHz): δ 156.1, 152.7, 150.2, 139.3, 134.2, 126.6, 118.2, 112.8, 110.7,
109.6, 46.2, 38.3, 34.7, 32.7, 31.7, 30.7, 30.5, 23.7, 23.5, 19.3, 14.4.
ESI-MS m/z 349, 351 [M + H]+ ratio 3:1; HR ESI-MS m/z [M +
H]+349.1919 (calcd for C21H3035ClO2, 349.1929).
PPAR-γ Activity Evaluation. Human embryonic kidney epithelial
cells 293T cells were obtained from the American Type Culture
Collection (CRL-3216) and cultured in Dulbecco’s Modified Eagle’s
Medium (DMEM) supplemented with 10% fetal calf serum (FCS)
and antibiotics. To analyze the PPAR-γ transcriptional activity, HEK-
293T cells were cultured in 24-well plates (2 × 104 cells/well) and
transiently cotransfected with GAL4-PPAR-γ (50 ng) and GAL4-luc
(firefly luciferase, 50 ng) vectors using Roti-Fect (Carl Roth). Twenty
hours after transfection the cells were stimulated with increasing
concentrations of the compounds for 6 h, and luciferase activities
were quantified using Dual-Luciferase Assay (Promega). Rosiglitazone
(1 μM, Cayman Chemical), was used as a positive control for PPAR-γ
activation (50-fold induction over basal activity). Test compounds
and controls stocks were prepared in DMSO, and the final
concentration of the solvent was always less than 0.5% v/v. The
plasmid GAL4-PPAR-γ was obtained from Prof. C. Sinal (Dalhousie
University). Half-maximal effective concentration (EC50) was
estimated using Prism software (GraphPad). All transfection experi-
ments were performed at least three times.
Cannabigeroquinone (CBGQ, 20). Red powder, IR νmax (KBr
disc): 3272, 2955, 2923, 2856, 1644, 1637, 1350, 1316, 1191, 1175,
1
580 cm−1; H NMR (CDCl3, 400 MHz) δ 6.94 (1H, s, OH), 6.45
(1H, bs, H-2′), 5.13 (1H, t, J = 7.4 Hz, H-2), 5.04 (1H, t, J = 6.7 Hz,
H-7), 3.13 (2H, d, J = 7.4 Hz, H-1), 2.41 (2H, t, J = 7.6, H-1″), 1.99−
1.90 (4H, m, H-4, H-5), 1.73 (3H, s, H-8), 1.64 (3H, s, H-9), 1.57
(3H, s, H-10), 1.50 (2H, m, H-2′′), 1.33 (4H, m, H-3′′, H-4′′), 0.89
(3H, t, J = 6.8 Hz, H-5″); 13C NMR (CDCl3, 100 MHz) δ 187.7,
184.2, 150.9, 145.1, 137.3, 134.4, 131.5, 124.3, 120.2, 119.7, 39.8,
31.5, 28.3, 27.4, 26.7, 25.8, 22.5, 22.0, 17.8, 16.3, 14.0; ESI-MS: m/z
331 [M + H]+; high-resolution (HR) ESI-MS m/z 331.2262 [M +
H]+, calcd. for C21H31O3, 331.2268.
Cannabichromenquinone (CBCQ, 21). Red oil, IR νmax (KBr
1
disc): 2957, 2926, 2852, 1648, 1580, 1324, 1078, 969, 891 cm−1; H
NMR (CDCl3, 400 MHz) 6.47 (1H, d, J = 9.9 Hz, H-1), 6.40 (1H, bs,
H-2′), 5.56 (1H, d, J = 9.9 Hz, H-2), 5.07 (1H, t, J = 6.9 Hz, H-6),
2.39 (2H, t, J = 7.6 Hz, H-1″), 2.08 (1H, m, H-5a), 1.88 (1H, m, H-
5b), 1.66 (2H, overlapped, H-4), 1.64 (3H, s, H-8), 1.55 (3H, s, H-9),
1.49 (2H, m, H-2′′), 1.46 (3H, s, H-10), 1.32 (4H, m, H-3′′, H-4′′),
0.89 (3H, t, J = 6.7 Hz, H-5″); 13C NMR (CDCl3, 100 MHz) δ 184.6,
181.9, 150.8, 147.7, 132.2, 131.4, 128.8, 123.4, 115.4, 115.0, 83.0,
41.5, 31.4, 28.7, 27.4, 27.3, 25.6, 22.6, 22.4, 17.7, 13.9; ESI-MS m/z
329 [M + H]+; HR ESI-MS m/z 329.2107 [M + H]+, calcd for
C21H29O3, 329.2111.
Cannabinolquinone (CBNQ, 22). Red oil, IR νmax (KBr disc):
2955, 2924, 2855, 1649, 1382, 1145, 1110, 811 cm−1 1H NMR
;
(CDCl3, 400 MHz) δ 8.30 (1H, s, H-2), 7.09 (1H, d, J = 7.9 Hz, H-
6), 7.02 (1H, d, J = 7.9 Hz, H-5), 6.63 (1H, t, J = 1.4 Hz, H-2′), 2.40
(2H, t, J = 7.7 Hz, H-1″), 2.36 (3H, s, H-7), 1.69 (6H, s, H-9, H-10),
1.56 (2H, m, H-2″), 1.32 (4H, m, H-3′′, H-4′′), 0.90 (3H, t, J = 6.8
Hz, H-5″); 13C NMR (CDCl3, 100 MHz) δ 180.2, 175.3, 163.3,
144.7, 138.1, 133.8, 131.8, 128.9, 125.7, 122.3, 111.0, 82.7, 53.6, 31.6,
29.8, 29.0, 28.3, 27.4, 22.4, 21.4, 13.9; ESI-MS m/z 325 [M + H]+;
HR ESI-MS m/z 325.1791 [M + H]+, calcd. for C21H25O3, 325.1798.
Dimeric Cannabigeroquinone (23) and Chiral-Phase Chro-
matography. Red powder, IR νmax (KBr disc): 3280, 2955, 1350,
1
1188, cm−1; H NMR (CDCl3, 400 MHz) δ 6.98 (1H, s, OH), 5.17
ASSOCIATED CONTENT
■
(1H, t, J = 7.4 Hz, H-2), 5.06 (1H, t, J = 6.7 Hz, H-7), 3.16 (2H, d, J
= 7.4 Hz, H-1), 2.32 (2H, t, J = 7.6, H-1″), 2.05−1.90 (4H, m, H-4,
H-5), 1.73 (3H, s, H-8), 1.64 (3H, s, H-9), 1.57 (3H, s, H-10), 1.49
(2H, m, H-2′′), 1.32 (4H, m, H-3′′, H-4′′), 0.89 (3H, t, J = 6.8 Hz,
H-5″). ESI-MS: m/z 645 [M + H]+; HR ESI-MS m/z 645.4159 [M +
H]+, calcd. for C41H57O6, 645.4155.
A sample of compound 23 (2.0 mg) was separated on a chiral-
phase Lux 5 μ Amylose-2 250 × 4.60 mm column, Phenomenex,
eluent n-hexane/isopropyl alcohol 9:1 (0.2% trifluoroacetic acid
(TFA)) with a flow of 0.7 mL/min, and two peaks were obtained with
Rt = 8 min (0.9 mg) and Rt = 13 min (0.7 mg).
Oxidation of Cannabidiol (CBD, 1) with the Takehira
Reagent. To a stirred solution of CBD (1, 200 mg, 0,64 mmol) in
toluene−tert-butanol (3:1, 20 mL), copper(II) chloride (43 mg, 0.32
mmol, 0.5 molar equiv) and hydroxylamine hydrochloride (22 mg,
0.32 mmol, 0,.5 molar equiv) were added. The solution turned from
yellow to brown and was stirred for 2 h at rt, worked up by dilution
with 2N H2SO4 and extraction with EtOAc. The organic phase was
washed with brine, dried with Na2SO4, filtered, and evaporated. The
residue was purified by GCC (5 g silica gel, petroleum ether−EtOAc
gradient, from to petroleum ether to 95:5 petroleum ether−EtOAc as
eluent) to give 12 (135 mg, 34%) and 11 (20%).
sı
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acs.jnatprod.9b01284.
1H and 13C NMR spectra for CBDQ (2) and the other
cannabinoquinoids mentioned in this study (PDF)
AUTHOR INFORMATION
■
Corresponding Authors
Orazio Taglialatela-Scafati − Dipartimento di Farmacia,
̀
Universita di Napoli Federico II, 80131 Napoli, Italy;
orcid.org/0000-0001-8010-0180; Phone: +39-081-
678509; Email: scatagli@unina.it; Fax: +39-081678552
Giovanni Appendino − Dipartimento di Scienze del Farmaco,
̀
Universita del Piemonte Orientale, 28100 Novara, Italy;
orcid.org/0000-0002-4170-9919; Phone: +39-0321-
375744; Email: giovanni.appendino@uniupo.it; Fax: +39-
0321375744
Hydroxyiminocannabiquinone (11). Brownish oil, IR νmax (KBr
disc): 2960, 2924, 2856, 1617, 1420, 1420, 1260, 1092, 1016, 797
Authors
1
cm−1; H NMR (methanol-d4, 400 MHz) δ 6.25 (1H, s, H-2′), 5.12
Diego Caprioglio − Dipartimento di Scienze del Farmaco,
Universita del Piemonte Orientale, 28100 Novara, Italy
Daiana Mattoteia − Dipartimento di Scienze del Farmaco,
Universita del Piemonte Orientale, 28100 Novara, Italy
̀
(1H, s, H-2), 4.50 (1H, s, H-9a), 4.49 (1H, s, H-9b), 3.82 (1H, m, H-
3), 2.92 (1H, td, J = 11.7, 3.2 Hz, H-4), 2.70 (2H, t, J = 7.5 Hz, H-
1″), 2.18 (1H, m, H-6a), 1.99 (1H, m, H-6b), 1.73 (2H, overlapped,
̀
D
https://dx.doi.org/10.1021/acs.jnatprod.9b01284
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
pubs.acs.org/jnp
Note
(14) Takehira, K.; Shimizu, M.; Watanabe, Y.; Orita, H.; Hayakawa,
T. J. Chem. Soc., Chem. Commun. 1989, 1705−1706.
Federica Pollastro − Dipartimento di Scienze del Farmaco,
̀
Universita del Piemonte Orientale, 28100 Novara, Italy
(15) Takehira, K.; Shimizu, M.; Watanabe, Y.; Orita, H.; Hayakawa,
T. Tetrahedron Lett. 1989, 30, 6691−6692.
Roberto Negri − Dipartimento di Scienze del Farmaco,
̀
Universita del Piemonte Orientale, 28100 Novara, Italy
(16) Nicolaou, K. C.; Montagnon, T.; Baran, P. S.; Zhong, Y.-L. J.
Am. Chem. Soc. 2002, 124, 2245−2258.
(17) Frigerio, M.; Santagostino, M.; Sputore, S. J. Org. Chem. 1999,
64, 4537−4538.
̀
Annalisa Lopatriello − Dipartimento di Farmacia, Universita di
Napoli Federico II, 80131 Napoli, Italy
̀
Giuseppina Chianese − Dipartimento di Farmacia, Universita
di Napoli Federico II, 80131 Napoli, Italy
(18) Dess, D. B.; Martin, J. C. J. Org. Chem. 1983, 48, 4155−4156.
(19) Ozanne, A.; Pouysegu, L.; Depernet, D.; Francois, B.; Quideau,
S. Org. Lett. 2003, 5, 2903−2906 We previously disclosed the use of
SIBX for the telescoped synthesis of aminoderivatives of cannabino-
quinoids in the proprietary literature:. Appendino, G.; Cabello de
Alba, M. L. B.; Munoz, E. WO158381, 2015.
(20) The product resulting from the Beam-type oxidation was
reported as a brown powder.6 We confirmed this observation,
presumably related to the formation of highly colored impurities
under the basic conditions of the Beam oxidation that could not be
removed by hexane washing or recrystallization.
Alberto Minassi − Dipartimento di Scienze del Farmaco,
̀
Universita del Piemonte Orientale, 28100 Novara, Italy
Juan A. Collado − Maimonides Biomedical Research Institute of
́
Cordoba; Department of Cellular Biology, Physiology and
́
Immunology, University of Cordoba; University Hospital Reina
́
Sofia, 14004 Cordoba, Spain
Eduardo Munoz − Maimonides Biomedical Research Institute of
́
Cordoba; Department of Cellular Biology, Physiology and
́
Immunology, University of Cordoba; University Hospital Reina
́
Sofia, 14004 Cordoba, Spain
(21) Appendino, G.; Pollastro, F.; Verotta, L.; Ballero, M.; Romano,
A.; Wyrembek, P.; Szczuraszek, K.; Mozrzymas, J. W.; Taglialatela-
Scafati, O. J. Nat. Prod. 2009, 72, 962−965.
Complete contact information is available at:
https://pubs.acs.org/10.1021/acs.jnatprod.9b01284
́
(22) Granja, A. G.; Carrillo-Salinas, F.; Pagani, A.; Gomez-Canas,
̃
M.; Negri, R.; Navarrete, C.; Mecha, M.; Mestre, L.; Fiebich, B. L.;
Cantarero, I.; Calzado, M. A.; Bellido, M. L.; Fernandez-Ruiz, J.;
Notes
The authors declare no competing financial interest.
Appendino, G.; Guaza, C.; Mun
2012, 7, 1002−1016.
̃
oz, E. J. Neuroimmune Pharmacol.
(23) Compounds were considered inactive if unable to induce 5-fold
PPARC-induction at 50 M concentration.
ACKNOWLEDGMENTS
■
We are grateful to Dr. G. Grassi (CREA, Rovigo) for
generously supplying the Carmagnola strain of Cannabis
used for the isolation of cannabidiol. We thank MIUR for
financial support to the groups in Novara and Naples
(PRIN2017, Project No. 2017WN73PL, Bioactivity-directed
exploration of the phytocannabinoid chemical space).
̌
(24) Hanus, L. O.; Tchilibon, S.; Ponde, D. E.; Breuer, A.; Fride, E.;
Mechoulam, R. Org. Biomol. Chem. 2005, 3, 1116−1123.
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https://dx.doi.org/10.1021/acs.jnatprod.9b01284
J. Nat. Prod. XXXX, XXX, XXX−XXX