Novel, regioselective transformation of an oxirane system. An efficient approach to the synthesis of endocannabinoid 2-arachidonoylglycerol

Article abstract of DOI:10.1016/S0040-4039(02)00116-8

A trifluoroacetic anhydride-catalysed opening of the oxirane system of glycidyl arachidonate with a simultaneous migration of the acyl group provides a new, efficient entry to 2-arachidonoylglycerol.

Full text of DOI:10.1016/S0040-4039(02)00116-8

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 43 (2002) 1759–1761
Novel, regioselective transformation of an oxirane system. An
efficient approach to the synthesis of endocannabinoid
2-arachidonoylglycerol
Stephan D. Stamatova,* and Jacek Stawinskib,c,
*
aDepartment of Chemical Technology, University of Plovdiv, 24 Tsar Assen St., Plovdiv 4000, Bulgaria
bDepartment of Organic Chemistry, Arrhenius Laboratory, Stockholm University, S-106 91 Stockholm, Sweden
cInstitute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, 61-704 Poznan, Poland
Received 7 December 2001; accepted 14 January 2002
Abstract—A trifluoroacetic anhydride-catalysed opening of the oxirane system of glycidyl arachidonate with a simultaneous
migration of the acyl group provides a new, efficient entry to 2-arachidonoylglycerol. © 2002 Published by Elsevier Science Ltd.
It has only recently been discovered1 that 2-arachi-
donoylglycerol (2-AG) might be an intrinsic, natural
ligand for central and peripheral cannabinoid receptors
(CB1 and CB2), that had previously been identified as
specific targets for a major psychoactive ingredient of
marijuana, D9-tetrahydrocannabinol. There is a grow-
ing line of evidence that 2-AG is a lipid mediator
governing a variety of protective or physiopathological
events in the central nervous,2 cardiovascular,3 and
immune systems,4 including hormone regulation,5
inflammation control,6,7 and also inhibiting prolifera-
tion of human breast and prostate cancer cells.7,8
poses severe complications in their synthesis, isolation,
storage, etc. Secondly, a problem specific to 2-AG is
that the arachidonoyl moiety exhibits pronounced sus-
ceptibility to autoxidation affecting integrity of the
native olefinic system and thus limiting the number of
available procedures for its preparation.
Two chemical methods described in the literature for
the synthesis of 2-AG are based on the same chemistry:
acylation of suitable 1,3-protected glycerol precursors
with an arachidonic acid derivative, followed by depro-
tection and separation of the isomeric arachidonoyl-
glycerols. In the original method developed by Martin12
and its two most recent modifications,9,13 1,3-benzyli-
deneglycerol is used as a substrate and, after introduc-
tion of the arachidonoyl moiety, the acetal group is
removed using boric acid derivatives. In the other
approach,14 triisopropylsilyl (TIPS) groups are used for
This diverse array of biological activities exhibited by
2-AG caused a high demand for isomerically pure
2-arachidonoylglycerol for biochemical and structure-
activity relation studies. Despite the apparent simplicity
of the chemical structure of 2-AG, access to this ligand
is still limited and in most instances the compound is
isolated from natural sources,7 as chemical9 and enzy-
matic methods10 for its preparation are rather
inefficient.
There are two problems that make synthesis of 2-AG
most difficult. Firstly, due to the presence of two adja-
cent primary hydroxyl functions, 2-acyl glycerols show
high propensity towards isomerisation11 (acid, base and
heat promoted migration of an acyl group) and this
Keywords: 2-acylglycerol; 2-arachidonoylglycerol; glycidyl arachi-
donate; acyl migration.
* Corresponding authors.
Scheme 1.
0040-4039/02/$ - see front matter © 2002 Published by Elsevier Science Ltd.
PII: S0040-4039(02)00116-8

1760
S. D. Stamato6, J. Stawinski / Tetrahedron Letters 43 (2002) 1759–1761
the protection of 1- and 3-hydroxyl functions of glyc-
erol and their removal from 1,3-bissilyl-2-arachi-
fluoroacetate) 2 in a CH2Cl2–pentane with pyridine
(10 equiv.) and methanol (15 equiv.). The reaction
was quantitative (completion within 3 h) and after
removal of volatile products via evaporation, isomeri-
cally homogenous 2-arachidonoylglycerol 318 (purity
donoyl intermediate is effected by
a prolonged
treatment with tetra-n-butylammonium fluoride
(TBAF) and acetic acid at low temperature.
1
>99%, H and 13C NMR spectroscopy) was obtained
Although useful in a general sense, these methods
provide only a partial solution to synthetic problems.
Difficulties include extended reaction time, acidic con-
ditions required for the removal of protecting groups,
necessity for workup after each synthetic step or sep-
aration of the intermediates from the accompanying
by-products, etc. These have frequently been reported
to contribute to isomerisation and oxidative or
hydrolytic side-reactions during the synthesis of 2-
acylglycerols. To lessen the problem of acyl migra-
tions, in these synthetic procedures the deprotection
steps were either not taken to completion,14 or the
produced isomeric compounds were separated by vari-
ous chromatographic techniques.9,14
in 92% overall yield (calculated on 1) with no neces-
sity for additional purification.
A typical procedure for the preparation of 2 and 3: To
a solution of glycidyl arachidonate 1 (0.180 g, 0.50
mmol), in alcohol-free dichloromethane (2.0 mL),
trifluoroacetic anhydride (TFAA, 0.278 mL, 2.00
mmol), in alcohol-free dichloromethane (2.0 mL), was
added at −20°C, and the reaction mixture was kept at
room temperature for 1 h. The solvent and unreacted
TFAA were removed under reduced pressure (bath
temp. 40°C), the residue was dissolved in toluene (5.0
mL) and passed through a silica gel pad (ꢀ5 g) pre-
pared in the same solvent. The support was washed
with toluene (100 mL) and the solvent was removed
under reduced pressure to provide 1,3-bis(trifluoro-
acetyl)-2-arachidonoylglycerol 2 as a yellowish oil.
Searching for an alternative methodology that would
circumvent these shortcomings during 2-acylglycerol
synthesis, we have developed an efficient and highly
stereoselective transformation of glycidyl arachidonate
1 into 2-arachidonoylglycerol derivative 2, promoted
by trifluoroacetic anhydride (TFAA) (Scheme 1).
Mechanistic details of this transformation remain to
be clarified, but the observed high regioselectivity sug-
gested intermediacy of the cyclic 1-trifluoroacetyl-2,3-
acyliumglycerol cation (formed via an intramolecular
attack of the adjacent carbonyl group on a trifluoro-
acetic anhydride-activated epoxide function in 1) that
in the presence of trifluoroacetate ion collapses into
1,3-bis(trifluoroacetyl)-2-arachidonoylglycerol 2. Com-
pound 2, from which 2-arachidonoylglycerol is gen-
erated under mild conditions, can be envisaged as a
convenient storage form of 2-AG (protection of 1-
and 3-hydroxyl groups as trifluoroacetyl esters should
prevent scrambling of the arachidonoyl moiety) or as
a possible prodrug for this lipid mediator.
1
Yield, 0.267 g (94%, purity >99%, H NMR spectro-
scopy).
To produce 2-arachidonoylglycerol 3, to a solution of
2 in pentane–CH2Cl2 (2:1, v/v, 5.0 mL), pyridine (0.40
mL, 5.0 mmol) and methanol (0.30 mL, 7.5 mmol) in
the same solvent (5.0 mL) were added at −20°C, and
the reaction mixture was left at room temperature for
3 h. After evaporating solvents, 3 was obtained as a
yellowish oil. Yield, 0.175 g (92%, calculated on 1;
1
purity >99%, H NMR spectroscopy).
In conclusion, we have developed an efficient synthetic
strategy based on a novel, regioselective transforma-
tion of glycidyl arachidonate 1 into 2-arachidonoyl-
1,3-bis(trifluoroacetyl)glycerol
2 intermediate, from
which 2-arachidonoylglycerol 3 can be retrieved under
mild conditions. The main features of this new syn-
thetic protocol are: (i) highly effective and practically
quantitative, one-pot synthesis of 2-arachidonoylglyc-
erol 3 under mild reaction conditions; (ii) the pro-
duced compounds 2 and 3 are of high purity, which
alleviates problems of their additional purification,
and thus the extent of acyl migration (and other side-
reactions) is minimised; (iii) glycidyl arachidonate 1
and intermediate 2 can be considered as convenient
storage forms of 2-AG; (iv) the method makes use of
commercially available reactants and it is easy to
scale-up.
Efficacy of this approach for the synthesis of 2-arachi-
donoyl glycerol 3 was assessed by subjecting ( )-gly-
cidyl arachidonate 1 (obtained in one step from
commercially available ( )-glycidol and arachidonic
acid in 95% yield15,16) to the transformations shown
in Scheme 1. To this end, compound
1
in
dichloromethane was treated with TFAA (4 equiv.) at
room temperature for 1 h. 1H and 13C NMR spec-
troscopy revealed that under these conditions the con-
version of
1
to 1,3-bis(trifluoroacetate) 217 was
quantitative and completely regioselective (>99%).
Thus, intermediate 2 can be either directly used for a
subsequent transformation, or isolated (ꢀ94% yield,
see Experimental) and stored for several months
(−20°C, under argon) without detectable alterations of
its spectral characteristics (1H and 13C NMR spec-
troscopy).
Acknowledgements
We are indebted to Professor P. J. Garegg for his
interest in this work. Financial support from the
Swedish Natural Science Research Council and the
Swedish Foundation for Strategic Research is grate-
fully acknowledged.
Since trifluoroacetate esters are known to undergo
smooth transesterification with alcohols,19 as a final
step of this synthetic protocol we treated 1,3-bis(tri-

S. D. Stamato6, J. Stawinski / Tetrahedron Letters 43 (2002) 1759–1761
1761
References
400 MHz) 0.88 (t, J=6.8 Hz, 20-CH3, 3H); 1.22–1.40 (m,
17-19-CH2, 6H); 1.71 (p, J=7.5 Hz, 3-CH2, 2H); 2.05,
2.11 (m, 16-CH2, 4-CH2, 4H); 2.36 (t, J=7.5 Hz, 2-CH2,
2H); 2.64 (dd, J=2.6, 2.6 Hz, C(O)CH2CHCHaHbO,
1H); 2.77–2.86 (m, 7, 10, 13-CH2, C(O)CH2CHCHaHbO,
7H); 3.19 (m, C(O)CH2CHCH2O, 1H); 3.91 (dd, J=6.2,
6.2 Hz, OCH2CHCHaHbOC(O), 1H); 4.40 (dd, J=3.3,
2.9 Hz, OCH2CHCHaHbOC(O), 1H); 5.28–5.44 (m,
CHꢀCH, 8H); 13C NMR lC (in ppm, CDCl3, 100 MHz)
14.28 (20-CH3); 22.78 (19-C); 24.91 (3-C); 25.83 (7, 10,
13-C); 26.73 (16-C); 27.43 (4-C); 29.54 (18-C); 31.73
(17-C); 33.62 (2-C); 127.76, 128.07, 128.35, 128.45,
128.80, 129.04, 129.19, 130.71 (5, 6, 8, 9, 11, 12, 14,
15-C); 173.47 (1-C): arachidonoyl fragment; 44.87 (1-C);
49.56 (2-C); 65.05 (3-C): oxirane-2-methyl fragment.
17. Yellowish oil; Rf (pentane:toluene:EtOAc=40:50:10, v/v/
v)=0.57; anal. calcd for C27H36O6F6 (570.57): C, 56.84;
1. (a) Sugiura, T.; Kondo, S.; Sukagawa, A.; Nakane, S.;
Shinoda, A.; Itoh, K.; Yamashita, A.; Waku, K.
Biochem. Biophys. Res. Commun. 1995, 215, 89–97; (b)
Mechoulam, R.; Ben-Shabat, S.; Hanus, L.; Ligumsky,
M.; Kaminski, N. E.; Schatz, A. R.; Gopher, A.; Almog,
S.; Martin, B. R.; et al. Biochem. Pharmacol. 1995, 50,
83–90.
2. (a) Pop, E. Curr. Opin. Chem. Biol. 1999, 3, 418–425; (b)
Sugiura, T.; Yoshinaga, N.; Waku, K. Neurosci. Lett.
2001, 297, 175–178; (c) Di Marzo, V.; Melck, D.;
Bisogno, T.; De Petrocellis, L. Trends Neurosci. 1999, 22,
80.
3. Mechoulam, R.; Fride, E.; Di Marzo, V. Eur. J. Pharma-
col. 1998, 359, 1–18.
4. Sugiura, T.; Kondo, S.; Kishimoto, S.; Miyashita, T.;
Nakane, S.; Kodaka, T.; Suhara, Y.; Takayama, H.;
Waku, K. J. Biol. Chem. 2000, 275, 605–612.
5. Murphy, L. L.; Munoz, R.; Adrian, B. A.; Villanua, M.
A. Neurobiol. Dis. 1998, 5, 432–446.
6. (a) Burnstein, S. H. Pharmacol. Ther. 1999, 82, 87–96; (b)
Sugiura, T.; Waku, K. Chem. Phys. Lipids 2000, 108,
89–106.
7. Di Marzo, V.; Melck, D.; De Petrocellis, L.; Bisogno, T.
Prostaglandins Other Lipid Mediat. 2000, 61, 43–61.
8. Melck, D.; De Petrocellis, L.; Orlando, P.; Bisogno, T.;
Laezza, C.; Bifulco, M.; Di Marzo, V. Endocrinology
2000, 141, 118–126.
9. Seltzman, H. H.; Fleming, D. N.; Hawkins, G. D.; Car-
roll, F. I. Tetrahedron Lett. 2000, 41, 3589–3592.
10. Schmid, P. C.; Schwartz, K. D.; Smith, C. N.; Krebs-
bach, R. J.; Berdyshev, E. V.; Schmid, H. H. O. Chem.
Phys. Lipids 2000, 104, 185–191.
11. Under acidic conditions (e.g. 0.03N perchloric acid) the
equilibrium between isomeric acylglycerols is rapidly
established (ca. 10 min) yielding usually a ca. 9:1 mixture
of 1-acyl- and 2-acylglycerols, see: Martin, J. J. Am.
Chem. Soc. 1953, 75, 5483–5486.
12. Martin, J. B. J. Am. Chem. Soc. 1953, 75, 5482–5483.
13. Suhara, Y.; Takayama, H.; Nakane, S.; Miyashita, T.;
Waku, K.; Sugiura, T. Chem. Pharm. Bull. 2000, 48,
903–907.
14. Han, L.; Razdan, R. K. Tetrahedron Lett. 1999, 40,
1631–1634.
15. Neises, B.; Steglich, W. Angew. Chem., Int. Ed. Engl.
1978, 17, 522–524.
1
H, 6.36. Found: C, 56.93; H, 6.30. H NMR lH (in ppm,
CDCl3, 400 MHz) 0.89 (t, J=6.9 Hz, 20-CH3, 3H);
1.22–1.40 (m, 17-19-CH2, 6H); 1.70 (p, J=7.3 Hz, 3-CH2,
2H); 2.05, 2.12 (m, 16-CH2, 4-CH2, 4H); 2.36 (t, J=7.5
Hz, 2-CH2, 2H); 2.72–2.90 (m, 7, 10, 13-CH2, 6H); 4.46
(dd, J=5.5, 5.5 Hz, C(O)OCHaHbCHCHaHbOC(O), 2H);
4.63 (dd, J=4.2, 4.4 Hz, C(O)OCHaHbCHCHaHbOC(O),
2H); 5.27–5.46 (m, CHꢀCH, CH2CHCH2, 9H); 13C NMR
lC (in ppm, CDCl3, 100 MHz) 14.26 (20-CH3); 22.78
(19-C); 24.70 (3-C); 25.81 (7, 10, 13-C); 26.57 (16-C);
27.43 (4-C); 29.53 (18-C); 31.73 (17-C); 33.41 (2-C);
127.73, 128.03, 128.22, 128.54, 128.68, 128.83, 129.44,
130.72 (5, 6, 8, 9, 11, 12, 14, 15-C); 172.51 (1-C): arachi-
donoyl fragment; 114.50 (q, J=285.3 Hz, 2-C); 157.17 (q,
J=43.5 Hz, 1-C): trifluoroacetyl fragment; 64.92 (1-C,
3-C); 67.22 (2-C): glycerol fragment.
18. Yellowish oil; Rf (pentane:toluene:EtOAc=30:20:50, v/v/
v)=0.33; anal. calcd for C23H38O4 (378.56): C, 72.98; H,
1
10.12. Found: C, 73.00; H, 10.18. H NMR lH (in ppm,
CDCl3, 400 MHz) 0.88 (t, J=6.8 Hz, 20-CH3, 3H);
1.22–1.41 (m, 17-19-CH2, 6H); 1.73 (p, J=7.3 Hz, 3-CH2,
2H); 2.05, 2.13 (m, 16-CH2, 4-CH2, 4H); 2.39 (t, J=7.6
Hz, 2-CH2, 2H); 2.73–2.89 (m, 7, 10, 13-CH2, 6H); 3.82
(d, J=4.8 Hz, OCH2CHCH2O, 4H); 4.92 (tt, J=4.6, 4.6
Hz, OCH2CH, 1H); 5.28–5.46 (m, CHꢀCH, 8H); 13C
NMR lC (in ppm, CDCl3, 100 MHz) 14.29 (20-CH3);
22.78 (19-C); 24.97 (3-C); 25.83 (7, 10, 13-C); 26.71
(16-C); 27.43 (4-C); 29.54 (18-C); 31.74 (17-C); 33.90
(2-C); 127.75, 128.06, 128.32, 128.54, 128.85, 128.99,
129.26, 130.76 (5, 6, 8, 9, 11, 12, 14, 15-C); 174.03 (1-C):
arachidonoyl fragment; 62.67 (1-C, 3-C); 75.28 (2-C):
glycerol fragment.
16. Yellowish oil; Rf (pentane:EtOAc=90:10, v/v)=0.33;
anal. calcd for C23H36O3 (360.54): C, 76.62; H, 10.06.
1
19. Lok, C. M. Chem. Phys. Lipids 1978, 22, 323–337.
Found: C, 76.70; H, 10.04. H NMR lH (in ppm, CDCl3,