The anandamide membrane transporter. Structurea€"activity relationships of anandamide and oleoylethanolamine analogs with phenyl rings in the polar head group region

Article abstract of DOI:10.1016/j.bmc.2004.07.026

A new series of arachidonic and oleic acids derivatives, most of which with aromatic moieties in the head group region, has been synthesized and evaluated as inhibitors of anandamide uptake. A new series of anandamide and N-oleoylethanolamine analogs, most of which with aromatic moieties in the head group region, has been synthesized and evaluated as inhibitors of anandamide uptake. Some of them efficaciously inhibit the uptake process with K_i values in the low micromolar range (2.4-21.2 μM). Strict structural requisites are needed to observe a significant inhibition and in no case inhibition of fatty acid amidohydrolase overlaps with inhibition of anandamide uptake.

Full text of DOI:10.1016/j.bmc.2004.07.026

Bioorganic & Medicinal Chemistry 12 (2004) 5161–5169
The anandamide membrane transporter. Structure–activity
relationships of anandamide and oleoylethanolamine analogs
with phenyl rings in the polar head group region
Vincenzo Di Marzo,a Alessia Ligresti,a Enrico Morera,b
Marianna Nallib and Giorgio Ortarb,
*
aEndocannabinoid Research Group, Institute of Biomolecular Chemistry, Consiglio Nazionale delle Ricerche,
Via Campi Flegrei 34, 80078 Pozzuoli, Napoli, Italy
b
`
Dipartimento di Studi Farmaceutici, Universita di Roma ÔLa SapienzaÕ, Piazzale A. Moro 5, 00185 Roma, Italy
Received 19 March 2004; accepted 13 July 2004
Available online 7 August 2004
Abstract—A new series of anandamide and N-oleoylethanolamine analogs, most of which with aromatic moieties in the head group
region, has been synthesized and evaluated as inhibitors of anandamide uptake. Some of them efficaciously inhibit the uptake
process with Ki values in the low micromolar range (2.4–21.2lM). Strict structural requisites are needed to observe a significant
inhibition and in no case inhibition of fatty acid amidohydrolase overlaps with inhibition of anandamide uptake.
Ó 2004 Elsevier Ltd. All rights reserved.
1. Introduction
potential therapeutic applications of compounds acting
on the proteins that constitute the ECS.23–30 In particu-
lar, AEA transport inhibitors could offer a therapeutic
approach to a variety of diseases including pain,31 multi-
ple sclerosis,32 HuntingtonÕs chorea33 in which elevation
of AEA levels would seem to afford more favorable
responses and fewer undesirable side-effects than direct
activation of CB1 receptors by agonists.
The endogenous cannabinoid system (ECS)1 comprises
two G-protein coupled receptors, CB1 and CB2,3 their
2
endogenous ligands [anandamide (AEA),4 2-arachido-
noylglycerol (2-AG),5 2-arachidonyl glyceryl ether (2-
AGE, noladin ether)6 and O-arachidonoylethanolamine
(virodhamine)]7 and an endocannabinoid inactivation
system. AEA and 2-AG are inactivated via a cellular up-
take, facilitated by one or more transporters,8 followed
by enzymatic hydrolysis, which is catalyzed by a fatty
acid amide hydrolase (FAAH)9 and, in the case of 2-
AG, also by monoacylglycerol lipases.10 The putative
AEA transporter (AMT) has not yet been cloned and
its nature is still controversial,11,12 although consider-
able evidence, including selective inhibition by various
structural analogs of AEA, points to its existence.13–22
The well established involvement of ECS in the regula-
tion of a wide series of both central and peripheral pro-
cesses has stimulated an enormous interest in the
Studies of structure–activity relationships concerned
with the AMT and its substrates/inhibitors represent
therefore an area of considerable interest. A large num-
ber of modifications on the three potential pharmaco-
phore moieties of AEA, the hydrophobic cis-tetraene
side chain, the carboxamido group and the polar head
group, has been already carried out34 and resulted in
the identification of a number of potent inhibitors such
as AM404,35 some N-acylvanillylamides (N-AVAMs),36
VDM11,37 UCM707,15 3-pyridinylarachidonylamide38
and SKM4451.39 However, AM404 also activates vanil-
loid (TRPV1) receptors, inhibits FAAH37,40 and is not
very stable to enzymatic hydrolysis; N-AVAMs are also
agonists at TRPV1 and CB1 receptors, while VDM11,
although almost inactive at these receptors and at
FAAH, is hydrolytically instable.37 Finally, 3-pyridinyl-
arachidonylamide also has very high affinity for
FAAH.38 The identification of new potent, selective
Keywords: Endocannabinoids; Anandamide transporter; Anandamide
transporter inhibitors; Structure–activity relationships.
*
Corresponding author. Tel.: +39-649913612; fax: +39-6491491;
e-mail: giorgio.ortar@uniroma1.it
0968-0896/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.07.026

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V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
Table 1. Effect of compounds 1–15 on [14C]AEA uptake by RBL-2H3
cellsa
and metabolically stable AMT inhibitors appears thus
still highly desirable.
O
The most potent AMT inhibitors so far developed have
been obtained by introducing aromatic moieties in the
polar head group region. The stabilization of the bind-
ing has been tentatively attributed to aromatic stacking
interactions between inhibitors and the transporter.38 In
this line and taking also into account changes previously
shown to diminish the affinity of aromatic long chain
fatty acid amides for TRPV1 receptors and/or FAAH,
we have recently designed and synthesized a series of
10-(4-hydroxybenzyl) analogues of N-oleoylethanol-
amine (OEA) and AEA (OMDM1-4, compounds 1, 2
of Table 1).14 OMDM1-4 inhibited to a varied extent
the AMT, the oleoyl analogues being 6- to 7-fold more
potent than the arachidonoyl counterparts. All com-
pounds exhibited poor affinity for both CB1 and CB2
receptors and almost no activity at TRPV1 receptors
and at FAAH.
R
a: oleoyl series
O
R
b: arachidonoyl series
Compd
R
Ki (lM)
OH
OH
1a
1b
2.4 0.7b
16.6 0.9b
OH
OH
N
H
3.0 0.6b
17.7 1.5b
In the present study we have conducted a systematic
investigation on the structure–activity relationships of
AMT inhibitors related to OMDM1-4 in order to gain
further insight into the molecular requisites for the
inhibitory action. To this end, a representative series
of derivatives of arachidonic and oleic acids with general
structure I (Fig. 1) has been designed. In these deriva-
tives the influence of factors such as the position of
the aromatic ring, its distance from the ethanolamine
portion, the presence or not of phenolic or benzylic hy-
droxy groups and the stereochemistry of the substitution
has been analyzed. Furthermore, the oleoyl analogue
(II, R = oleoyl) of AM404 (II, R = arachidonoyl), two
homologs of AM404 (III) and amides derived from (S)
and (R)-prolinol (IV) have been prepared and
evaluated.
2a
2b
N
H
3a
3b
>25
>25
OH
OH
N
H
4a
4b
11.1 2.0
>25
N
H
OH
2. Results and discussion
2.1. Chemistry
5a
5b
>25
17.5 2.3
OH
N
H
The synthesis of amides listed in Table 1 has been
accomplished by direct condensation between arachi-
donic or oleic acid and the appropriate amine using
1-hydroxybenzotriazole (HOBt)/N-ethyl-N0-(3-dimeth-
ylaminopropyl)carbodiimide hydrochloride (EDC) as
the carboxylate activator in 64–87% yields (Fig. 2).
The synthesis of the noncommercial amines (R) and
(S)-4-hydroxyphenylglycinol was carried out by reduc-
tion of (R) and (S)-N-Boc-4-hydroxyphenylglycine
methyl esters with LiAlH4, followed by deprotection
with SOCl2/MeOH.
OH
6a
6b
>25
21.2 4.1
OH
N
H
7a
7b
12.9 3.1
19.4 1.2
2.2. Biological data
OH
OH
N
H
The effect of compounds 1–15 on the uptake of
[14C]AEA by intact RBL-2H3 cells, where the AMT
has been well characterized,41 is shown in Table 1. For
comparative purposes, we have also included the results
previously obtained with OMDM1-414 and AM404
(compound 12b).37
8a
8b
16.6 2.1
>25
N
H

V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
5163
Table 1 (continued)
Indeed, some of the compounds inhibited efficaciously
the uptake process with Ki values in the low micromolar
range (2.4–21.2lM), while others were inactive at the
highest concentration tested (25lM). None of our com-
pounds inhibited significantly FAAH from RBL-2H3
cells up to a concentration of 50lM. Several conclusions
on the transporter affinity can be drawn from data of
Table 1. Removal of the hydroxy group in para on the
phenyl ring of compounds 1, 2 results in a decrease of
the affinity for AMT (compounds 3 and 4), which is par-
ticularly evident in the case of (S)-isomer of oleic acid
series (3a). An analogous effect, concerned with the
introduction of an hydroxy group in para of N-phenyl-
arachidonylamide has been previously described by
Piomelli et al.42 and Hillard and co-workers38 and was
attributed to a stabilization of binding. An opposite
trend is, however, observed here for the series of a-phen-
ylethanolamine analogues (compounds 5–8), where the
presence of the para-OH group proves to be of little ef-
fect or even detrimental for the inhibitory ability. The
decrease in the distance of the aromatic ring on the eth-
anolamine moiety by elimination of the methylene
spacer produces variable effects on the inhibitory po-
tency, depending on the presence of hydroxy groups,
the stereochemistry and the position of the ring (com-
pounds 1, 2 and 3, 4 compared with 5, 6 and 7, 8, 9,
respectively). The introduction of an hydroxy group
on the benzylic carbon atom affords appreciably active
compounds only in the case of (S,S)-diastereomer of
the oleic acid series (compound 11a) and of (R,R)-dia-
stereomer of arachidonic acid series (compound 10b),
a result difficult to interpret. The inactivity of the benzyl
analogues 13 of AM404 seems to indicate that the stabil-
ization of the binding to the carrier by a hydrogen bond-
ing acceptor/donor group near to the head region
binding site is very sensitive to the position of this
group. Compound 13b has been described by Makriyan-
nis and co-workers as a poor FAAH substrate as well.43
To study the effects of head group conformational
preferences, a set of AEA analogues in which the hy-
droxyalkyl group was partially restricted by its incorpo-
ration into five- or six-membered rings has been
previously synthesized.42 The resulting 3-hydroxypyrrol-
idinyl- and 3- and 4-hydroxypiperidinylamides exhibited
affinity for the transporter similar to that of AEA.
In contrast, the conformationally restricted analogues
of AEA and OEA 14 and 15 are, rather surprisingly,
completely inactive, suggesting that the hydrogen of
the hydroxyl group may be involved in an intramole-
cular hydrogen bond. The arachidonoyl 2-pyrrolidine-
methanol derivatives 14b, 15b have been previously
investigated for their effects on the intraocular pressure
(IOP) in normotensive rats, causing a statistically signif-
icant reduction of IOP.44 Regarding the stereochemical
factor, the stereoselectivity of the transporter, already
evidenced in previous studies, is further confirmed by
the differences in the inhibitory potencies of various cou-
ples of stereoisomers such as 3a–4a, 7b–8b, 10a–11a and
10b–11b. Regarding the nature of the hydrophobic tail,
compounds of the oleic acid series are able, generally, to
compete with [14C]AEA more efficaciously than arachi-
donoyl analogues or, at least, in a comparable fashion.
The presence of only one cis double bond located in
Compd
R
Ki (lM)
OH
N
H
9a
9b
>25
>25
HO
10a
10b
>25
15.7 3.7
OH
OH
N
H
OH
11a
11b
6.4 0.5
>25
N
H
OH
12a
12b
>25
8.1 2.6c
N
H
OH
13a
13b
>25
>25
N
H
OH
OH
14a
14b
>25
>25
N
15a
15b
>25
>25
N
a The inhibitory effect is expressed as the Ki calculated using the
Cheng–Prusoff equation from the concentrations exerting 50% inhi-
bition of
experiments.
[
14C]AEA uptake. Data are means SEM of n = 3
b Data from Ref. 14.
c Value from Ref. 37.
R2
OH
R1
R
N
H
R
n
N
H
OH
I
II
OH
OH
R
R
N
H
N
III
IV
R = arachidonoyl, oleoyl; R1, R2 = H, OH; n = 0, 1
Figure 1. General structures of the compounds synthesized.
Although it is clear, according to previous studies,8 that
the head group region in the AEA transporter can toler-
ate bulk additions, strict structural requisites appear to
be necessary in order to observe a significant inhibition.

5164
V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
R2
R1
n
R
OH
N
H
1-11
1) EDC/HOBt
or
O
R2
OH
OH
O
R1
2)
R
n
N
OH
H
H2N
12a
or
or
OH
OH
or
H2N
OH
or
H2N
OH
R
N
H
13
OH
or
HN
or
OH
R
N
14,15
Figure 2. Synthesis of compounds 1–15.
the middle of the fatty acid chain has been shown by
Piomelli et al.42 to represent a sufficient requisite for
AMT recognition and, among the ethanolamides of
fatty acids other than arachidonic acid, N-oleoyletha-
nolamine proved to be the best inhibitor.8 In this re-
spect, the inactivity of the oleoyl analogue 12a of
AM404 (12b) is quite unexpected and deserves further
investigation. In conclusion, it is evident that little struc-
tural variations in the aromatic moiety of the polar head
region produce more or less profound differences in the
affinity for AMT. Most importantly, the molecular req-
uisites of the transport process differ significantly from
those of the interaction with FAAH, despite the fact
that the head group binding region in FAAH can also
tolerate large head groups. The reactivity of the sub-
strate43,45 does not seem to be a major factor in deter-
mining the affinity for FAAH.
most no activity at TRPV1 receptors, while some nota-
ble affinity for CB1 receptors was observed only for 4a.
Compounds 7a and, particularly, 11a can be viewed,
therefore, as relatively selective inhibitors of the AMT.
3. Experimental
3.1. Chemistry
Melting points were determined with a Kofler hot stage
apparatus and are uncorrected. Optical rotations were
taken at 20°C with a Schmidt–Haensch polartronic D
polarimeter in a 1dm cell. Infrared spectra were
recorded with a Perkin–Elmer Spectrum 1000 FT-IR
spectrophotometer. 1H and 13C NMR spectra were mea-
sured with a Varian Mercury 300 spectrometer using
tetramethylsilane as internal standard. Column chroma-
tographies were carried out using Merck silica gel 60
(230–400mesh).
The three most potent new inhibitors (compounds 4a, 7a
and 11a) were also assessed for their activity on cannab-
inoid CB1, CB2 and TRPV1 receptors (Table 2). The
three compounds exhibited weak affinity for CB2 and al-
3.1.1. (S)-N-Boc-4-hydroxyphenylglycine methyl ester.
To a stirred suspension of 4-hydroxyphenylglycine
(1.672g, 10mmol) in 20mL of dry MeOH, thionyl chlo-
ride (0.87mL, 12mmol) was added dropwise at 0°C and
the resulting mixture was stirred 15min at 0°C and 16h
at room temperature. Evaporation under reduced pres-
sure of the solution afforded a residue, which was dis-
solved in dry CH2Cl2 (25mL) and additioned of TEA
(1.4mL, 10mmol), NaHCO3 (1.26g, 15mmol) and
(Boc)2O (2.40g, 11mmol). The reaction mixture was
stirred at room temperature for 3h, then concentrated
under vacuum, acidified with 0.5N HCl and extracted
with AcOEt. The organic phase was washed with brine,
10% aqueous solution of NaHCO3, brine, dried over
Na2SO4 and evaporated to give crude (S)-N-Boc-4-hy-
Table 2. Effects of compounds 4a, 7a, 11a on CB1, CB2 and TRPV1
receptors
Compd hCB1 (Ki lM)a hCB2 (Ki lM)a hTRPV1 (efficacy)b
4a
7a
0.5 0.01
2.7 0.01
>10
2.5 0.02
2.0 0.1
3.2 0.2
9.3 1.6
10.5 2.0
10.5 2.1
11a
a The affinity for CB1 and CB2 receptors was measured in displacement
assays carried out using transfected human COS cells and
[3H]CP55,940.
b Efficacy at human TRPV1 receptors transfected in HEK cells was
measured by expressing the effect of a 10lM concentration of each
compound as percent of the effect of 4lM ionomycin. Data are
means SEM of n = 3 experiments.

V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
5165
droxyphenylglycine methyl ester (2.614g), which was
further purified by crystallization from CH2Cl2/hexane
3.1.7. (Z)-N-{(1S)-2-Hydroxy-1-[(4-hydroxyphenyl)meth-
yl]ethyl}-9-octadecenamide (1a). Yield 82%; mp 88–
(2.30g. 82%): mp 138–139°C; ½aꢀ þ110 (c 1.0, CHCl3);
89°C; ½aꢀ ꢁ13 (c 1.0, CHCl3); IR (KBr) 3432, 3325,
D
IR (KBr) 3431, 3368, 2987, 1733, 1673, 1614, 1506,
D
2928, 2856, 1651, 1515, 1234, 1201cmꢁ1
;
1H NMR
1441, 1320, 1213, 1164, 1061cmꢁ1
;
1H NMR
(300MHz, CDCl3) d 0.87 (3H, t, J = 6.5Hz), 1.26
(20H, m), 1.52–1.56 (2H, m), 1.96–2.00 (4H, m), 2.12
(2H, t, J = 7.6Hz), 2.65–2.78 (2H, m), 3.12 (1H, br s),
3.47 (1H, dd, J = 11.1, 5.1Hz), 3.56 (1H, dd, J = 11.1,
4.0Hz), 4.07 (1H, m), 5.26–5.37 (2H, m), 6.38 (1H, d,
J = 8.4Hz), 6.70 (2H, d, J = 8.4Hz), 6.97 (2H, d,
J = 8.4Hz), 8.05 (1H, br s); 13C NMR (75MHz, CDCl3)
d 14.09, 22.66, 25.72, 27.17, 27.21, 29.15, 29.30, 29.50,
29.74, 31.87, 36.15, 36.77, 52.73, 52.82, 63.63, 115.34,
128.46, 129.55, 129.81, 129.97, 155.05, 174.19. Anal.
Calcd for C27H45NO3 (431.7): C, 75.13; H, 10.51; N,
3.24. Found: C, 75.28; H, 10.44; N, 3.18.
(300MHz, CDCl3) d 1.43 (9H, s), 3.70 (3H, s), 5.21
(1H, d, J = 7.2Hz), 5.63 (1H, d, J = 7.2Hz), 6.72 (2H,
d, J = 8.7Hz), 6.83 (1H, br s), 7.15 (2H, d, J = 8.4Hz);
13C NMR (75MHz, CDCl3) d 28.27, 52.64, 57.00,
80.43, 115.64, 127.85, 128.19, 154.86, 156.14, 171.75.
3.1.2. (S)-N-Boc-4-hydroxyphenylglycinol. To a stirred
solution of (S)-N-Boc-4-hydroxyphenylglycine methyl
ester (2.117g, 7.53mmol) in dry THF (100mL), LiAlH4
(0.569g, 15.0mmol) was added portionwise in 30min.
The mixture was stirred at room temperature for 16h
and then Et2O saturated with water was cautiously
added. The suspension was poured into a cool 5%
tartaric acid solution and extracted with AcOEt. The or-
ganic phase was washed twice with brine, filtered, dried
over Na2SO4 and evaporated. The residue (2.291g) was
chromatographed on silica gel (70g) using hexane/
AcOEt = 1:1 as eluent to give pure (S)-N-Boc-4-hy-
droxyphenylglycinol (1.321g, 69%): mp 176–178°C
3.1.8. (all-Z)-N-{(1S)-2-Hydroxy-1-[(4-hydroxyphenyl)-
methyl]ethyl}-5,8,11,14-eicosatetraenamide (1b). Yield
74%; wax; ½aꢀ ꢁ17 (c 1.0, CHCl3); IR (CHCl3) 3433,
D
3331, 2930, 2870, 1656, 1615, 1515, 1456, 1261cmꢁ1
;
1H NMR (300MHz, CDCl3) d 0.88 (3H, t, J = 6.9Hz),
1.25–1.37 (6H, m), 1.64 (2H, quintet, J = 7.5Hz), 2.04
(4H, q, J = 6.9Hz), 2.15 (2H, t, J = 7.6Hz), 2.71–2.83
(8H, m), 3.49 (1H, m), 3.52 (1H, dd, J = 11.1, 4.9Hz),
3.61 (1H, dd, J = 11.1, 3.3Hz), 4.12 (1H, m), 5.26–5.41
(8H, m), 5.98 (1H, d, J = 7.8Hz), 6.71 (2H, d,
J = 8.4Hz), 6.97 (2H, d, J = 8.4Hz), 7.65 (1H, br s);
13C NMR (75MHz, CDCl3) d 14.07, 22.54, 25.47,
25.60, 26.53, 27.18, 29.27, 31.46, 36.12, 36.21, 52.96,
63.76, 115.44, 127.30, 127.62, 127.91, 128.07, 128.42,
128.49, 128.67, 129.94, 130.32, 154.91, 173.90. Anal.
Calcd for C29H43NO3 (453.7): C, 76.78; H, 9.55; N,
3.09. Found: C, 77.02; H, 9.42; N, 3.00.
(from acetone); ½aꢀ þ69 (c 1.0, MeOH); IR (KBr)
D
3401, 3250, 1644, 1560, 1444, 1315, 1169, 1057cmꢁ1
;
1H NMR (300MHz, CDCl3/CD3OD = 9:1) 1.42 (9H,
s), 3.62–3.76 (H, m), 4.59 (1H, m), 5.77 (1H, m), 6.79
(2H, d, J = 8.7Hz), 7.12 (2H, d, J = 8.4Hz); 13C NMR
(75MHz, CDCl3/CD3OD = 9:1) d 28.31, 56.31, 65.93,
79.85, 115.30, 127.63, 156.01, 156.40.
3.1.3. (S)-4-Hydroxyphenylglycinol hydrochloride. To a
solution of (S)-N-Boc-4-hydroxyphenylglycinol (0.253g,
1.0mmol) in dry MeOH (7.0mL) cooled at 0°C, thionyl
chloride (0.073mL, 1.0mmol) was added and the mix-
ture was stirred at 40°C for 2h. Evaporation under
reduced pressure of the solution afforded (S)-4-hydroxy-
phenylglycinol hydrochloride (0.189g), which was used
in the next steps without further purification.
3.1.9. (Z)-N-{(1R)-2-Hydroxy-1-[(4-hydroxyphenyl)meth-
yl]ethyl}-9-octadecenamide (2a). Yield 83%; mp 88–
89°C; ½aꢀ þ13 (c 1.0, CHCl3). Anal. Calcd for
D
C27H45NO3 (431.7): C, 75.13; H, 10.51, N, 3.24. Found:
C, 75.30; H, 10.42; N, 3.19.
3.1.4. (R)-N-Boc-4-hydroxyphenylglycinol. Prepared fol-
lowing the same procedure used for the synthesis of
(S) enantiomer. Yield 65.0%; mp 176–178°C (acetone);
3.1.10. (all-Z)-N-{(1R)-2-Hydroxy-1-[(4-hydroxyphen-
yl)methyl]ethyl}-5,8,11,14-eicosatetraenamide (2b). Yield
82%; ½aꢀ þ18 (c 1.0, CHCl3). Anal. Calcd for
½aꢀ ꢁ69 (c 1.0, MeOH).
D
D
C29H43NO3 (453.7): C, 76.78; H, 9.55, N, 3.09. Found:
C, 77.09; H, 9.40; N, 2.98.
3.1.5. (R)-4-Hydroxyphenylglycinol hydrochloride. Pre-
pared following the same procedure used for the synthe-
sis of (S) enantiomer.
3.1.11. (Z)-N-[(1S)-2-Hydroxy-1-(phenylmethyl)ethyl]-9-
octadecenamide (3a). Yield 66%; wax; ½aꢀ ꢁ12 (c 2.0,
D
CHCl3); IR (CHCl3) 3434, 2928, 2856, 1654, 1510,
3.1.6. General procedure for the synthesis of compounds
1–15. To a solution of fatty acid in DMF at 0°C, HOBt
(1.05equiv) and EDC (1.05equiv) were added under stir-
ring. After 15min at 0°C and 0.5h at room temperature,
the appropriate amine hydrochloride (1equiv) and TEA
(1equiv) were added and the mixture was stirred at
room temperature overnight. The mixture was diluted
with brine and extracted with AcOEt. The organic phase
was washed with 10% citric acid solution, saturated
NaHCO3 and brine, dried over Na2SO4 and evaporated
under reduced pressure. The residue was purified by col-
umn chromatography on silica gel using hexane/AcOEt
or CH2Cl2/AcOEt mixtures as eluents.
1
1465, 1238, 1088cmꢁ1; H NMR (300MHz, CDCl3) d
0.88 (3H, t, J = 6.6Hz), 1.27 (20H, m), 1.51–1.58 (2H,
m), 1.99–2.02 (4H, m), 2.11 (2H, t, J = 7.6Hz), 2.79–
2.93 (2H, m), 3.55 (1H, dd, J = 11.0, 5.1Hz), 3.63 (1H,
dd, J = 11.0, 3.9Hz), 3.78 (1H, br s), 4.16 (1H, m),
5.28–5.40 (2H, m), 6.12 (1H, d, J = 7.5Hz), 7.17–7.30
(5H, m); 13C NMR (75MHz, CDCl3) d 14.09, 22.65,
25.72, 27.16, 27.19, 29.13, 29.28, 29.49, 29.73, 31.86,
36.77, 36.95, 52.61, 63.72, 126.34, 128.31, 129.00,
129.47, 129.77, 137.58, 173.64. Anal. Calcd for
C27H45NO2 (415.7): C, 78.02; H, 10.91; N, 3.37. Found:
C, 78.23; H, 10.79; N, 3.29.

5166
V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
3.1.12. (all-Z)-N-[(1S)-2-Hydroxy-1-(phenylmethyl)-
ethyl]-5,8,11,14-eicosatetraenamide (3b). Yield 79%;
C26H43NO3 (417.6): C, 74.78; H, 10.38; N, 3.35. Found:
C, 74.89; H, 10.47; N, 3.27.
wax; ½aꢀ ꢁ17 (c 1.0, CHCl3); IR (CHCl3) 3434, 2929,
D
2858, 1657, 1509, 1455, 1234, 1039cmꢁ1
;
1H NMR
3.1.18. (all-Z)-N-[(1R)-2-Hydroxy-1-(4-hydroxyphenyl)-
ethyl]-5,8,11,14-eicosatetraenamide (6b). Yield 74%;
(300MHz, CDCl3) d 0.89 (3H, t, J = 6.9Hz), 1.29–1.38
(6H, m), 1.64 (2H, quintet, J = 7.2Hz), 2.01–2.08 (4H,
m), 2.15 (2H, t, J = 7.5Hz), 2.76–2.90 (8H, m), 3.17
(1H, br s), 3.57 (1H, dd, J = 11.1, 5.1Hz), 3.66 (1H,
dd, J = 11.1, 3.6Hz), 4.17 (1H, m), 5.29–5.44 (8H, m),
5.83 (1H, d, J = 7.2Hz), 7.20–7.33 (5H, m); 13C NMR
(75MHz, CDCl3) d 14.06, 22.54, 25.45, 25.59, 26.52,
27.18, 29.27, 31.46, 36.08, 36.92, 52.68, 64.03, 126.44,
127.28, 127.61, 127.91, 128.02, 128.39, 128.56, 128.77,
128.95, 130.29, 137.39, 173.27. Anal. Calcd for
C29H43NO2 (437.7): C, 79.59; H, 9.90; N, 3.20. Found:
C, 79.85; H, 10.02; N, 3.11.
wax; ½aꢀ ꢁ50 (c 1.0, EtOH). Anal. Calcd for
D
C28H41NO3 (439.6): C, 76.50; H, 9.40; N, 3.19. Found:
C, 76.77; H, 9.29; N, 3.09.
3.1.19. (Z)-N-[(1S)-2-Hydroxy-1-phenylethyl]-9-octade-
cenamide (7a). Yield 80%; mp 53–54°C; ½aꢀ þ31 (c
D
1.0, CHCl3); IR (CHCl3) 3436, 2928, 2856, 1662, 1505,
1
1465, 1234, 1069cmꢁ1; H NMR (300MHz, CDCl3) d
0.88 (3H, t, J = 6.3Hz), 1.28 (20H, m), 1.62–1.67 (2H,
m), 1.97–2.02 (4H, m), 2.24 (2H, t, J = 7.2Hz), 2.83
(1H, br s), 3.88 (2H, m), 5.07 (1H, m), 5.31–5.36 (2H,
m), 6.15 (1H, d, J = 6.3Hz), 7.26–7.39 (5H, m); 13C
NMR (75MHz, CDCl3) d 14.09, 22.66, 25.69, 27.16,
27.21, 29.11, 29.22, 29.50, 29.67, 29.75, 31.87, 36.74,
55.94, 66.77, 126.49, 127.74, 128.73, 129.51, 129.80,
138.78, 173.52. Anal. Calcd for C26H43NO3 (401.6): C,
75.13; H, 10.51; N, 3.24. Found: C, 75.29; H, 10.43;
N, 3.15.
3.1.13. (Z)-N-[(1R)-2-Hydroxy-1-(phenylmethyl)ethyl]-9-
octadecenamide (4a). Yield 80%; wax; ½aꢀ þ12 (c 2.0,
D
CHCl3). Anal. Calcd for C27H45NO2 (415.7): C, 78.02;
H, 10.91; N, 3.37. Found: C, 78.25; H, 10.79; N, 3.29.
3.1.14. (all-Z)-N-[(1R)-2-Hydroxy-1-(phenylmethyl)eth-
yl]-5,8,11,14-eicosatetraenamide (4b). Yield 76%; wax;
½aꢀ þ18 (c 1.0, CHCl3). Anal. Calcd for C29H43NO2
3.1.20. (all-Z)-N-[(1S)-2-Hydroxy-1-phenylethyl]-5,8,11,
14-eicosatetraenamide (7b). Yield 70%; wax; ½aꢀ þ26 (c
D
(437.7): C, 79.59; H, 9.90; N, 3.20. Found: C, 79.88;
H, 10.03; N, 3.11.
D
1.0, CHCl3); IR (CHCl3) 3434, 2930, 1662, 1505, 1455,
1
1234, 1070cmꢁ1; H NMR (300MHz, CDCl3) d 0.89
3.1.15. (Z)-N-[(1S)-2-Hydroxy-1-(4-hydroxyphenyl)-
ethyl]-9-octadecenamide (5a). Yield 73%; mp 134–
(3H, t, J = 6.9Hz), 1.29–1.31 (6H, m), 1.71 (2H, quintet,
J = 7.5Hz), 2.02–2.11 (4H, m), 2.23 (2H, t, J = 7.5Hz),
2.77–2.84 (6H, m), 3.13 (1H, br s), 3.82 (2H, d,
J = 4.8Hz), 5.03 (1H, m), 5.29–5.44 (8H, m), 6.32 (1H,
d, J = 7.2Hz), 7.25–7.37 (5H, m); 13C NMR (75MHz,
CDCl3) d 14.05, 22.54, 25.44, 25.60, 26.62, 27.18,
29.27, 31.47, 35.98, 55.79, 66.40, 126.47, 127.31,
127.62, 127.91, 128.04, 128.40, 128.49, 128.63, 128.78,
130.29, 138.84, 173.22. Anal. Calcd for C28H41NO2
(423.6): C, 79.39; H, 9.75; N, 3.31. Found: C, 79.16;
H, 9.88; N, 3.20.
135°C; ½aꢀ þ54 (c 1.0, EtOH); IR (CHCl3) 3630,
D
3423, 3013, 2943, 2837, 1651, 1516, 1463, 1334, 1238,
1
1012cmꢁ1; H NMR (300MHz, CDCl3/CD3OD = 9:1)
d 0.88 (3H, t, J = 6.6Hz), 1.28 (20H, m), 1.62 (2H, m),
2.00 (4H, m), 2.22 (2H, t, J = 7.5Hz), 3.76 (2H, m),
4.90 (1H, m), 5.34 (2H, m), 6.76 (2H, d, J = 8.9Hz),
7.90 (2H, d, J = 8.9Hz); 13C NMR (75MHz, CDCl3/
CD3OD = 9:1) d 14.10, 22.69, 25.77, 27.19, 27.22,
29.17, 29.27, 29.31, 29.52, 29.73, 31.90, 36.64, 55.12,
55.21, 65.91, 115.45, 127.73, 129.56, 129.83, 129.91,
156.09, 174.13. Anal. Calcd for C26H43NO3 (417.6): C,
74.78; H, 10.38; N, 3.35. Found: C, 74.90; H, 10.45;
N, 3.29.
3.1.21. (Z)-N-[(1R)-2-Hydroxy-1-phenylethyl]-9-octade-
cenamide (8a). Yield 76%; mp 55–56°C; ½aꢀ ꢁ31 (c
D
1.0, CHCl3). Anal. Calcd for C26H43NO3 (401.6): C,
75.13; H, 10.51; N, 3.24. Found: C, 75.31; H, 10.44;
N, 3.19.
3.1.16. (all-Z)-N-[(1S)-2-Hydroxy-1-(4-hydroxyphenyl)-
ethyl]-5,8,11,14-eicosatetraenamide (5b). Yield 66%;
wax; ½aꢀ þ50 (c 1.0, EtOH); IR (CHCl3) 3594, 3432,
3.1.22. (all-Z)-N-[(1R)-2-Hydroxy-1-phenylethyl]-5,8,11,
D
3009, 2930, 2858, 1656, 1515, 1458, 1234, 1039cmꢁ1
;
14-eicosatetraenamide (8b). Yield 71%; wax; ½aꢀ ꢁ26 (c
D
1H NMR (300MHz, CDCl3) d 0.88 (3H, t, J = 6.6Hz),
1.26–1.37 (6H, m), 1.69 (2H, quintet, J = 7.5Hz), 2.02–
2.13 (4H, m), 2.23 (2H, t, J = 7.2Hz), 2.77–2.84 (6H,
m), 3.73 (2H, m), 4.87 (1H, m), 5.29–5.42 (8H, m),
6.68 (2H, d, J = 8.5Hz), 7.16 (2H, d, J = 8.5Hz); 13C
NMR (75MHz, CDCl3) d 14.06, 22.55, 25.59, 26.64,
27.19, 29.28, 31.48, 35.94, 35.98, 55.14, 55.23, 65.81,
115.46, 127.31, 127.66, 127.91, 128.08, 128.42, 128.64,
128.75, 129.70, 130.32, 155.97, 173.92. Anal. Calcd for
C28H41NO3 (439.6): C, 76.50; H, 9.40; N, 3.19. Found:
C, 76.78; H, 9.28; N, 3.10.
1.0, CHCl3). Anal. Calcd for C28H41NO2 (423.6): C,
79.39; H, 9.75; N, 3.31. Found: C, 79.14; H, 9.87; N,
3.20.
3.1.23. (Z)-N-[(R,S)-2-Hydroxy-2-phenylethyl]-9-octade-
cenamide (9a). Yield 85%; wax; IR (CHCl3) 3451, 2928,
2856, 1656, 1518, 1455, 1234, 1065cmꢁ1 1H NMR
;
(300MHz, CDCl3) d 0.91 (3H, t, J = 6.6Hz), 1.30
(20H, m), 1.57–1.61 (2H, m), 2.01–2.05 (4H, m), 2.15
(2H, t, J = 7.5Hz), 3.27–3.36 (1H, m), 3.66 (1H, ddd,
J = 13.8, 6.6, 3.3Hz), 4.14 (1H, br s), 4.81 (1H, dd,
J = 7.5, 3.3Hz), 5.33–5.43 (2H, m), 6.21 (1H, m), 7.27–
7.36 (5H, m); 13C NMR (75MHz, CDCl3) d 14.09,
22.65, 25.69, 27.16, 27.20, 29.13, 29.19, 29.23, 29.28,
29.49, 29.70, 29.73, 31.86, 36.51, 47.39, 73.44, 125.60,
3.1.17. (Z)-N-[(1R)-2-Hydroxy-1-(4-hydroxyphenyl)-
ethyl]-9-octadecenamide (6a). Yield 72%; mp 134–
135°C; ½aꢀ ꢁ54 (c 1.0, EtOH). Anal. Calcd for
D

V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
5167
127.51, 128.22, 129.50, 129.79, 141.66, 174.47. Anal.
Calcd for C26H43NO3 (401.6): C, 75.13; H, 10.51; N,
3.24. Found: C, 75.38; H, 10.42; N, 3.16.
½aꢀ þ31 (c 1.0, CHCl3). Anal. Calcd for C29H43NO3
D
(453.7): C, 76.78; H, 9.55; N, 3.09. Found: C, 77.09;
H, 9.67; N, 2.98.
3.1.29. (Z)-N-4-Hydroxyphenyl-9-octadecenamide (12a).
Yield 64%; mp 104–105°C; IR (KBr) 3278, 2917, 2849,
3.1.24. (all-Z)-N-[(R,S)-2-Hydroxy-2-phenylethyl]-
5,8,11,14-eicosatetraenamide (9b). Yield 76%; wax; IR
(CHCl3) 3450, 2929, 2858, 1659, 1518, 1455, 1234,
1647, 1615, 1559, 1537, 1518, 1466, 1256, 1097cmꢁ1
;
1H NMR (300MHz, CDCl3) d 0.88 (3H, t, J = 6.6Hz),
1.27–1.31 (20H, m), 1.64–1.71 (2H, m), 1.97–2.02 (4H,
m), 2.30 (2H, t, J = 7.6Hz), 5.32–5.36 (2H, m), 6.72
(2H, d, J = 8.7Hz), 7.22 (2H, dd, J = 8.7, 1.8Hz), 7.65
(1H, br s); 13C NMR (75MHz, CDCl3) d 14.07, 22.65,
25.75, 27.16, 27.20, 29.13, 29.27, 29.49, 29.74, 31.86,
37.38, 115.47, 122.46, 122.58, 129.53, 129.79, 153.42,
172.51. Anal. Calcd for C24H39NO2 (373.6): C, 77.16;
H, 10.52; N, 3.75. Found: C, 77.41; H, 10.67; N, 3.67.
1
1206, 1065cmꢁ1; H NMR (300MHz, CDCl3) d 0.89
(3H, t, J = 6.6Hz), 1.26–1.38 (6H, m), 1.68 (2H, quintet,
J = 7.5Hz), 2.02–2.12 (4H, m), 2.17 (2H, t, J = 7.5Hz),
2.79–2.85 (6H, m), 3.25–3.30 (1H, m), 3.67 (1H, ddd,
J = 13.8, 6.9, 3.3Hz), 3.77 (1H, br s), 4.80 (1H, d,
J = 5.7Hz), 5.28–5.43 (8H, m), 6.02 (1H, d,
J = 7.2Hz), 7.26–7.34 (5H, m); 13C NMR (75MHz,
CDCl3) d 14.06, 22.54, 25.44, 25.61, 26.59, 27.19,
29.28, 31.47, 35.84, 47.40, 73.57, 125.60, 127.31,
127.63, 127.94, 128.05, 128.27, 128.41, 128.62, 128.79,
130.31, 141.60, 174.09. Anal. Calcd for C28H41NO2
(423.6): C, 79.39; H, 9.75; N, 3.31. Found: C, 79.14;
H, 9.88; N, 3.23.
3.1.30. (Z)-N-[(4-Hydroxymethyl)phenyl]-9-octadecen-
amide (13a). Yield 75%; wax; IR (CHCl3) 3285, 2917,
2849, 1654, 1605, 1542, 1466, 1412, 1260, 1015cmꢁ1
;
1H NMR (300MHz, CDCl3) d 0.88 (3H, t, J = 6.6Hz),
1.32 (20H, m), 1.70 (2H, m), 1.96–2.02 (4H, m), 2.32
(2H, t, J = 7.6Hz), 2.65–2.78 (2H, m), 4.58 (2H, s),
5.32–5.36 (2H, m), 7.23 (2H, d, J = 8.4Hz), 7.43 (2H,
d, J = 8.4Hz), 7.65 (1H, br s); 13C NMR (75MHz,
CDCl3) d 13.74, 15.30, 23.07, 26.01, 27.55, 27.60,
28.66, 29.53, 29.69, 29.90, 30.13, 30.91, 32.27, 38.01,
65.03, 120.34, 127.89, 129.89, 1130.20, 136.95, 137.45,
171.98. Anal. Calcd for C25H41NO2 (387.6): C, 77.47;
H, 10.66; N, 3.61. Found: C, 77.72; H, 10.77; N, 3.53.
3.1.25. (Z)-N-[(1R,2R)-Phenyl-1,3-propanodiol-2-yl]-9-
octadecenamide (10a). Yield 82%; wax; ½aꢀ ꢁ28 (c
D
1.0, CHCl3); IR (CHCl3) 3433, 3025, 2928, 2856, 1655,
1509, 1465, 1199, 1062cmꢁ1 1H NMR (300MHz,
;
CDCl3) d 0.88 (3H, t, J = 6.6Hz), 1.11–1.34 (20H, m),
1.39–1.48 (2H, m), 1.99–2.09 (6H, m), 3.66–3.77 (2H,
m), 3.90 (1H, br s), 4.04–4.11 (1H, m); 4.35 (1H, br s),
5.18 (1H, d, J = 3.6Hz), 5.29–5.40 (2H, m), 6.31 (1H,
d, J = 7.8Hz), 7.20–7.33 (5H, m); 13C NMR (75MHz,
CDCl3) d 14.09, 22.65, 25.64, 27.17, 27.21, 29.02,
29.09, 29.22, 29.30, 29.49, 29.71, 29.74, 31.86, 36.64,
56.39, 63.12, 72.93, 125.58, 127.43, 128.14, 129.50,
129.79, 141.72, 174.31. Anal. Calcd for C27H45NO3
(431.7): C, 75.13; H, 10.51; N, 3.24. Found: C, 75.34;
H, 10.40; N, 3.16.
3.1.31. (all-Z)-N-[(4-Hydroxymethyl)phenyl]-5,8,11,14-
eicosatetraenamide (13b). Yield 77%; wax; IR (CHCl3)
3605, 3433, 2929, 2858, 1686, 1596, 1518, 1411, 1306,
1
1224, 1004cmꢁ1; H NMR (300MHz, CDCl3) d 0.88
(3H, t, J = 6.9Hz), 1.25–1.37 (7H, m), 1.80 (2H, quintet,
J = 7.2Hz), 2.05 (2H, q, J = 6.3Hz), 2.16 (2H, q,
J = 6.3Hz), 2.34 (2H, t, J = 6.9Hz), 2.78–2.83 (4H, m),
4.60 (2H, s), 5.34–5.41 (8H, m), 7.25 (2H, d,
J = 8.1Hz), 7.44 (2H, d, J = 8.1Hz), 7.55 (1H, br s);
13C NMR (75MHz, CDCl3) d 21.80, 23.66, 24.01,
24.52, 24.84, 25.79, 26.02, 26.42, 28.53, 30.71, 36.10,
63.92, 119.12, 126.53, 126.78, 126.86, 127.14, 127.30,
127.64, 127.98, 128.04, 129.58, 135.83, 136.26, 170.37.
Anal. Calcd for C27H39NO2 (409.6): C, 79.17; H, 9.60;
N, 3.42. Found: C, 79.44; H, 9.49; N, 3.33.
3.1.26. (all-Z)-N-[(1R,2R)-Phenyl-1,3-propanodiol-2-yl]-
5,8,11,14-eicosatetraenamide (10b). Yield 86%; wax;
½aꢀ ꢁ31 (c 1.0, CHCl3); IR (CHCl3) 3432, 3009, 2958,
D
2929, 2858, 1656, 1508, 1453, 1210, 1063cmꢁ1
;
1H
NMR (300MHz, CDCl3) d 0.89 (3H, t, J = 6.9Hz),
1.26–1.38 (6H, m), 1.54 (2H, quintet, J = 7.5Hz), 1.92–
1.99 (2H, m), 2.02–2.12 (4H, m), 2.73–2.84 (6H, m),
3.73 (3H, br s), 4.06–4.10 (1H, m), 4.22 (1H, br s),
5.02 (1H, br s), 5.26–5.44 (8H, m), 6.26 (1H, d,
J = 8.4Hz), 7.24–7.32 (5H, m); 13C NMR (75MHz,
CDCl3) d 14.06, 22.54, 25.45, 25.58, 26.47, 27.18,
29.27, 31.46, 35.98, 56.36, 63.19, 73.10, 125.56, 127.29,
127.49, 127.62, 127.92, 128.03, 128.16, 128.40, 128.54,
128.71, 130.29, 140.96, 173.93. Anal. Calcd for
C29H43NO3 (453.7): C, 76.78; H, 9.55; N, 3.09. Found:
C, 77.11; H, 9.69; N, 2.97.
3.1.32. (Z)-(R)-1-(1-Oxo-9-octadecenyl)-2-pyrrolidine-
methanol (14a). Yield 78%; wax; ½aꢀ ꢁ32 (c 2.0,
D
CHCl3); IR (CHCl3) 3346, 2928, 2855, 1612, 1445,
1
1340, 1234, 1015cmꢁ1; H NMR (300MHz, CDCl3) d
0.85 (3H, t, J = 6.5Hz), 1.23–1.28 (20H, m), 1.53–1.63
(3H, m), 1.78–2.04 (8H, m), 2.26 (2H, t, J = 7.6Hz),
3.38–3.64 (4H, m), 4.14–4.22 (1H, m), 5.25–5.36 (2H,
m); 13C NMR (75MHz, CDCl3) d 14.09, 22.65, 24.38,
24.76, 27.14, 27.18, 28.26, 29.10, 29.30, 29.48, 29.73,
31.86, 35.06, 45.61, 48.04, 58.91, 61.00, 63.67, 67.37,
129.52, 129.74, 174.31. Anal. Calcd for C23H43NO2
(365.6): C, 75.56; H, 11.85; N, 3.83. Found: C, 75.78;
H, 11.98; N, 3.75.
3.1.27. (Z)-N-[(1S,2S)-Phenyl-1,3-propanodiol-2-yl]-9-
octadecenamide (11a). Yield 87%; wax; ½aꢀ þ28 (c
D
1.0, CHCl3). Anal. Calcd for C27H45NO3 (431.7): C,
75.13; H, 10.51; N, 3.24. Found: C, 75.28; H, 10.43;
N, 3.15.
3.1.28. (all-Z)-N-[(1S,2S)-Phenyl-1,3-propanodiol-2-yl]-
5,8,11,14-eicosatetraenamide (11b). Yield 87%; wax;
3.1.33. (all-Z)-(R)-1-(1-Oxo-5,8,11,14-eicosatetraenyl)-2-
pyrrolidinemethanol (14b). Yield 84%; wax; ½aꢀ ꢁ28 (c
D

5168
V. Di Marzo et al. / Bioorg. Med. Chem. 12 (2004) 5161–5169
1.0, CHCl3); IR (CHCl3) 3343, 2930, 1613, 1446, 1341,
References and notes
1
1227, 1075cmꢁ1; H NMR (300MHz, CDCl3) d 0.89
1. Martin, B. R. J. Pharmacol. Exp. Ther. 2002, 301, 790.
2. Matsuda, L. A.; Lolait, S. J.; Brownstein, M. J.; Young,
A. C.; Bonner, T. I. Nature 1990, 346, 561.
3. Munro, S.; Thomas, K. L.; Abu-Shaar, M. Nature 1993,
365, 61.
(3H, t, J = 6.9Hz), 1.26–1.38 (6H, m), 1.57–1.65 (1H,
m), 1.72–1.79 (2H, m), 1.84–2.09 (5H, m), 2.11–2.18
(2H, m), 2.31 (2H, t, J = 7.5Hz), 2.77–2.83 (6H, m),
3.43–3.62 (4H, m), 4.18–4.25 (1H, m), 5.19 (1H, br s),
5.29–5.45 (8H, m); 13C NMR (75MHz, CDCl3) d
14.05, 22.54, 24.37, 24.52, 25.62, 26.65, 27.19, 28.25,
29.29, 31.48, 34.26, 47.99, 61.02, 67.26, 127.32, 127.63,
127.99, 128.37, 128.56, 129.03, 130.26, 162.49, 173.93.
Anal. Calcd for C25H41NO2 (387.6): C, 77.47; H,
10.66; N, 3.61. Found: C, 77.79; H, 10.78; N, 3.52.
ˇ
4. Devane, W. A.; Hanus, L.; Breuer, A.; Pertwee, R. G.;
Stevenson, L. A.; Griffin, G.; Gibson, D.; Mandelbaum,
A.; Etinger, A.; Mechoulam, R. Science 1992, 258, 1946.
5. Sugiura, T.; Kondo, S.; Sukagawa, A.; Nakane, S.;
Shinoda, A.; Itoh, K.; Yamashita, A.; Waku, K. Biochem.
Biophys. Res. Commun. 1995, 215, 89.
ˇ
6. Hanus, L.; Abu-Lafi, S.; Fride, E.; Breuer, A.; Vogel, Z.;
Shalev, D. E.; Kustanovich, I.; Mechoulam, R. Proc. Natl.
Acad. Sci. U.S.A. 2001, 98, 3662.
7. Porter, A. C.; Sauer, J.-M.; Knierman, M. D.; Becker, G.
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D
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