An efficient synthesis of 3-OBn-6β,14-epoxy-bridged opiates from naltrexone and identification of a related dual MOR inverse agonist/KOR agonist

Article abstract of DOI:10.1016/j.bmcl.2012.06.056

In an effort to better understand the conformational preferences that inform the biological activity of naltrexone and related naltrexol derivatives, a new synthesis of the restricted analog 3-OBn-6β,14-epoxymorphinan 4 is described. 4 was synthesized starting from naltrexone in 50% overall yield, proceeding through the OBn-6α-triflate intermediate 8. Key steps to the synthesis include benzylation (96% yield), reduction (90% yield, α:β:3:2), followed by a one-pot triflation/displacement sequence (96% yield) to yield the desired bridged epoxy derivative 4. X-ray crystallographic analysis of intermediate 3-OBn-6α-naltrexol 7a supports population of the key boat conformation required for the epoxy ring closure. We also report that the 6β-mesylate 10-a high affinity opioid receptor ligand, the epimeric derivative of 11, and an analog of 12-functions as an inverse agonist at the mu opioid receptor using herkinorin pre-conditioned cells and an agonist at the kappa opioid receptor when evaluated in independent in vitro [ ³⁵S]-GTP-γ-S assays.

Full text of DOI:10.1016/j.bmcl.2012.06.056

Bioorganic & Medicinal Chemistry Letters 22 (2012) 6801–6805
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
An efficient synthesis of 3-OBn-6b,14-epoxy-bridged opiates from naltrexone
and identification of a related dual MOR inverse agonist/KOR agonist
David J. Martin a,b, Paul E. FitzMorris a, Bo Li c, Mario Ayestas d, Ellicott J. Sally d, Christina M. Dersch d,
Richard B. Rothman d, Amy M. Deveau a,
⇑
a University of New England, College of Arts and Sciences, Chemistry & Physics Department, Biddeford, ME 04005, USA
b University of New England, College of Osteopathic Medicine, Biddeford, ME 04005, USA
c Boston College, Chemistry Department, X-ray Crystallography Center, Chestnut Hill, MA 02467, USA
d National Institute on Drug Abuse/National Institutes of Health, Addiction Research Center, Baltimore, MD 21224, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Available online 23 June 2012
In an effort to better understand the conformational preferences that inform the biological activity of
naltrexone and related naltrexol derivatives, a new synthesis of the restricted analog 3-OBn-6b,14-epox-
ymorphinan 4 is described. 4 was synthesized starting from naltrexone in 50% overall yield, proceeding
Keywords:
Naltrexone
Naltrexol
Antagonist
Inverse Agonist
Opiate
6b,14-Epoxymorphinan
Conformation
Rigid analog
through the OBn-6
reduction (90% yield,
yield the desired bridged epoxy derivative 4. X-ray crystallographic analysis of intermediate 3-OBn-
-naltrexol 7a supports population of the key boat conformation required for the epoxy ring closure.
a
-triflate intermediate 8. Key steps to the synthesis include benzylation (96% yield),
a:b:3:2), followed by a one-pot triflation/displacement sequence (96% yield) to
6a
We also report that the 6b-mesylate 10-a high affinity opioid receptor ligand, the epimeric derivative
of 11, and an analog of 12-functions as an inverse agonist at the mu opioid receptor using herkinorin
pre-conditioned cells and an agonist at the kappa opioid receptor when evaluated in independent
in vitro [35S]-GTP-
c-S assays.
Ó 2012 Elsevier Ltd. All rights reserved.
The discovery of novel opioid receptor antagonists is needed to
probe biochemical mechanisms of functional antagonism1 that
underlie the therapeutic relevance of naltrexone/ol derivatives.2–5
Both naltrexone and naltrexol (Fig. 1) are nitrogen heterocycles
that are comprised of five rings–rings A, B, C, D, and E- (Fig. 2);
these two compounds differ only in their functionality at C6. In
order to unlock structure–activity trends for MOR functional
antagonism in the naltrexol/one series, we are investigating chem-
ical approaches and computational methods to understand C Ring
conformation.2,6
Development of high yielding syntheses of conformationally-
constrained naltrexol/one derivatives provides insight into confor-
mational preferences while improving access to biologically inter-
esting opioid receptor probes. Specifically, in 2008 and 2011,
Nagase published the synthesis of novel 6b,14-epoxy morphinans
that target the kappa opioid receptor (KOR).7,8 In 1990,9 the Por-
toghese research group published the synthesis of 17-(cyclopro-
tert-butoxide (Fig. 3).9 In fact, the 3,6
a-mesylate used by Portogh-
ese is closely related to 103, the dual MOR inverse antagonist/KOR
agonist that we have identified in this work (vide infra, Fig. 1 and
Table 1). The Portoghese group proposed that the presence of a
boat conformation directed the proper alignment of nucleophile
and leaving group in the ether-forming SN2 reaction. As we are
very interested in manipulating conformational preferences of
Ring C in our synthetic and structure-activity studies, we hypoth-
esized that a better leaving group, such as a triflate, along with a
different base, lithium triethylborohydride, would improve the
yield of bridged epoxy derivatives. The use of benzyl protection
at position 3 also allows for further synthetic manipulation of 4.10
In this research, we report a new synthesis of 4 (Fig. 1), a con-
strained 6b,14-epoxy derivative of naltrexone that is functional-
ized as a benzyl ether at C3 (Fig. 2).11 Compound 4 is a derivative
of the high affinity (Ki = 1.1 nM)2 MOR antagonist 17-(cyclopro-
pylmethyl)-3,14-dihydroxy-6b-((methylsulfonyl)oxy)-4,5a-epox-
pylmethyl)-4,5
a
:6b,14-diepoxy-3-hydroxymorphinan, 5, that has
ymorphinan, 10 (Fig. 1). Because the 6b-mesylate 10 demonstrated
affinity at both the MOR and KOR in previous studies (6.1 subtype
selectivity ratio; KOR:MOR),3 we also studied 10’s functional activ-
ity in MOR and KOR-based functional assays. Using an in vitro
since been shown to have affinity for the MOR.11
Portoghese utilized 12, the 3,6-dimesylate of 6a-naltrexol, to
form the 6b,14-epoxy derivative 5 upon treatment with potassium
[
35S]-GTP- -S assay with herkinorin12 conditioned cells and the
c
inverse agonist (IA) standard KC-2-009,13,22 10 was identified as
an MOR inverse agonist (Table 1). An in vitro [35S]-GTP-
-S assay
⇑
c
Corresponding author.
E-mail address: adeveau@une.edu (A.M. Deveau).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.06.056

6802
D. J. Martin et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6801–6805
HO
HO
RO
HO
HO
O
O
O
O
O
N
OH
N
N
OH
N
OH
N
OH
H
RO
H
O
H
RO
H
H
O
1, Naltrexone
2, R= -H; 6β-Naltrexol
10, R= -Ms
3, R= -H; 6α-Naltrexol
11, R= -Ms
4, R= -Bn
5, R= -H
9, Desoxonaltrexone
Figure 1. Structures of naltrexone, naltrexol, and related derivatives.
Carbon 3
HO
Table 1
Summary of [35S]-GTP-
c
-S functional assay data
35S]-GTP-
-S functional assay data
A-ring: aromatic
B-ring: cyclohexane
C-ring: cyclohexanol
D-ring: piperidine
E-ring: cyclic ether
D
A
Drug
[
c
l
Opioid receptor inverse
agonisma
j
Opioid receptor agonismb
B
O
E
N
C
H
OH
EC50
(nM SD)
Emax
ED50
Emax
(% SD)
HO
Carbon 14
(% SD)
(nM SD)
Carbon 6
Figure 2. Ring labeling and key carbons for 6
6b-Mesylate,
10
6b-Naltrexol,
2.6 0.3
1.2 0.1
60.5 0.9
89.0 1.3
26
—
3
48
—
1
a-naltrexol.
2
a
Assay procedures were previously described.22 Dose–response curves (10
with expressed KOR receptors also characterized 10 as a KOR ago-
nist (Table 1).
points per curve) of MOR antagonists acting as inverse agonists in herkinorin
treated membranes. The Emax reported is a normalized value with 100% being the
inhibition of [ c-S binding produced by 5 lM KC-2-009. The data of 3
35S]-s-
The synthesis of 4 was achieved in a four-step, three-pot
synthetic sequence in 50% overall yield (Fig. 4). To simplify purifi-
cation of intermediates and provide increased flexibility for syn-
thetic manipulation of intermediates in our ongoing research, we
first protected naltrexone’s phenol as a benzyl ether using benzyl
bromide with K2CO3 in refluxing acetone to yield 6 (96% yield).10,11
The ketone of benzyl ether 6 was subsequently reduced with
NaBH4 in MeOH by using procedures adapted from Uwai’s previous
work.4g Uwai applied a À20 °C bath to direct a diastereoselective
NaBH4 reduction of naltrexone’s C6 ketone and form exclusively
experiments were meaned and the best-fit estimates of the EC50 and Emax deter-
mined using nonlinear least squares curve fitting programs. Each value is standard
deviation (SD).
b
Experimental procedures were followed as previously described using Chinese
hamster ovary cells which stably express the human kappa opiate receptor.23 All
results represent the mean of three experiments. Each value is SD.
research were purified using silica gel chromatography and spectro-
scopically characterized using NMR spectroscopy (1H, 13C, and 2D
NMR spectroscopy when appropriate), high resolution mass spec-
trometry, and IR spectroscopy.15 The isolated yields reported in this
work are unoptimized.16
the 6
the reduction at 0 °C to purposely get both 6
mers (90% overall yield, :b:3:2) of 7a. The 6b-diastereomer, 7b
(36% yield from :b mixture), which was easily separated from
the 6 -diastereomer (54% from :b mixture) by column chroma-
a-diastereomer in 94% yield. Alternatively, we completed
a
and 6b-diastereo-
a
Our route into the epoxy morphinan series using SN2 displace-
ment chemistry with a triflate leaving group was discovered seren-
dipitously. We were attempting to synthesize 6-desoxonaltrexone
9 ( Fig. 1) that Wentland’s group has since generated by the
straightforward Wolff–Kishner reduction of 1,17 but instead we
formed 4 in 96% yield. It is well-established that the use of rigid
scaffolds, like the 6b,14-epoxy bridge found in the C-Ring of 4, have
long been a valued strategy in elucidating important structure-
activity relationships (SARs).18 Therefore, we anticipate that epoxy
derivative 4 will be of great use to evolve our understanding of
SARs in MOR functional antagonism—for example differentiating
inverse agonists from neutral antagonists. In fact, Nelson and
coworkers previously identified that the debenzylated 6b,14-epoxy
a
a
a
tography, is being employed in other areas of our research. It is also
important to note that 7a was recrystallized from hot acetonitrile
followed by slow evaporation to form large, transparent, colorless
prisms that were of sufficient quality for X-ray analysis (Fig. 5).14
Once in hand the 3-OBn-6a-naltrexol derivative 7a was triflated
at À30 °C with triflic anhydride and pyridine in dry CH2Cl2 (Fig. 4).
The triflate intermediate was brought to À10 °C prior to in situ addi-
tion of LiBEt3H and then worked up. The one pot, two-step sequence
of triflation followed by nucleophilic displacement generated 17-
(cyclopropylmethyl)-4,5a:6b,14-diepoxy-3-benzyloxymorphinan,
4, in 96% yield. All intermediate and final compounds in this
K
O
H
H
OH
H
O
OMs
O
N
N
N
H
O
OMs
O
O
OMs
OMs
OH
6α-mesylate,12
6α-mesylate
boat conformation
6β,14-epoxy analog,5
chair conformation
boat conformation
Figure 3. Portoghese’s precedent for synthesis of the 6b,14-epoxy skeleton.

D. J. Martin et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6801–6805
6803
N
N
N
OH
OH
OH
BnBr
NaBH4
MeOH
0 °C to RT, 17 h
K2CO3
acetone
reflux, 8 h
(96%)
O
O
O
BnO
O
BnO
OH
HO
O
1
6
7a : 7b : α : β: 3 : 2
90% yield overall
N
N
N
OH
OH
(F3CSO2)2O
pyridine
LiBEt3H
THF
O
CH2Cl2, -30 °C
75 min
-10 °C, room temp.
1 h
BnO
O
O
O
OH
BnO
OTf
BnO
4, 96%- one pot, two steps
7a
8
Figure 4. Synthetic approach to epoxy derivative 4.
efficacy with synergistic KOR agonist properties, like that found
for 10, may be of use therapeutically or in basic research as a bio-
chemical probe.19
The chemistry in this work delivered good to excellent yields,
despite procedures being unoptimized. Starting from 1, both the
benzyl protection (96% yield) and hydride reduction (90% yield—
a:b:3:2) reactions were straightforward to yield intermediate dia-
stereomers 7a and 7b (Fig. 4). Because 7a was highly crystalline
even in a crude state, X-ray quality crystals were achieved with
minimal effort by recrystallization. An ORTEP diagram of 7a, which
is purposely oriented to highlight the solid-state conformation of
Rings C (a boat) and D (a chair), is provided in Figure 5. The pres-
ence of the Ring C boat conformation justified our proposed mech-
anism for conversion of alpha diastereomer, 7a, to 4 via triflate
intermediate 8.20 We first assume that triflation occurs with reten-
tion of configuration at C6. We subsequently hypothesize, in line
with the Portoghese9,20 precedent (Fig. 3), that both (S) configura-
tion at C6 and the boat conformation of Ring C are needed for a con-
certed displacement reaction to occur. Additional support for this
hypothesis comes from attempting the triflation/ring closure with
7b instead of 7a; in this case, the reaction was unsuccessful and
only starting material was recovered.
Figure 5. ORTEP diagram of 7a showing Ring C conformation. Thermal ellipsoids
are drawn at a 50% probability level.
Regarding the selection of base for the reaction, lithium trieth-
ylborohydride (Superhydride) was initially chosen during our
attempts to synthesize 9 (vide infra). We desired a hydride source
that was softer than NaH (and related ionic hydrides) with greater
solubility in THF that might facilitate displacement of triflate with-
out cleaving the existing ether contained within Ring E (Fig. 2).21
Clearly, the combination of a very good leaving group (triflate)
and lithium triethylborohydride was effective as evidenced by
our 96% yield for the triflation/ring closure sequence.
When considering the efficiency of this procedure, it is impor-
tant to note that the yields of the protection and triflation/ring
closure steps were close to quantitative. We are also satisfied with
our 50% unoptimized overall yield for the three-pot sequence.
Employing a diastereoselective reduction of 6 in future research
should greatly enhance our overall economy of the synthesis.
Overall, we report a new four-step, three-pot approach (benzy-
lation, reduction, triflation/ring closure; 50% overall yield) to the
6b,14-epoxymorphinan skeleton and synthesized compound 4
(Figs. 1 and 4), an opioid receptor ligand and derivative of the dual
MOR antagonist/KOR agonist 10 (Fig. 1; Table 1).3,11 Although more
steps, our overall yield is comparable to that previously reported
derivative 5 displaces the radioligand DAMGO from the MOR.9 One
additional question of our work is whether 4 and 5 ( Fig. 1),
constrained analogs with defined boat conformations for Ring
C (Fig. 3), block basal signaling when studied in an agonist-condi-
tioned environment.9 This question is currently under investiga-
tion and results will be reported in due course.
It is well-known that G protein-coupled receptors, like the MOR,
exhibit heighted basal activity after prolonged exposure to an ago-
nist (like morphine or herkinorin).5,12 Thus, blockage of this consti-
tutive signaling can be used to differentiate inverse agonists (IA)
from neutral antagonists (NA) in functional binding assays.5,22
The role of functional antagonism in developing an ideal opioid
addiction therapy is becoming increasingly important.5,22 In our
identification of 10’s inverse agonism, the noninternalizing MOR
agonist herkinorin was used for preconditioning cells in the
MOR-based [35S]-GTP- -S assay.12 Unlike morphine, herkinorin
c
does not promote b-arrestin recruitment and mu-receptor inter-
nalization.12 Although compound 10 was found to be less effica-
cious than naltrexol (Table 1) and the benchmark KC-2-009,13,22
the value of having a high affinity MOR antagonist of reasonable

6804
D. J. Martin et al. / Bioorg. Med. Chem. Lett. 22 (2012) 6801–6805
for a two-step synthesis of similar epoxymorphinan 5 (Fig. 3),9,11
and also allows for protective group incorporation. Additionally,
we achieved a high quality X-ray structure of key benzyl ether
intermediate 7a (Fig. 5).14 This crystal structure of 7a clearly con-
firms the population of a Ring C boat conformation that was
hypothesized to exist during the concerted ring closure reaction.
These results, when taken together with the functional assay data
on 10 and other studies,20 begin to provide further insight into con-
formational preferences of Ring C. Such perspectives inform future
synthetic efforts as we strive to better understand structural pat-
terns aligned with MOR functional antagonism (IA vs NA).
7. Nemoto, T.; Fujii, H.; Narita, M.; Miyosh, K.; Nakamura, A.; Suzuki, T.; Nagase,
H. Bioorg. Med. Chem. Lett. 2008, 18, 6398.
8. Nemoto, T.; Yamamoto, N.; Watanabe, A.; Fujii, H.; Hasebe, K.; Nakajima, M.;
Mochizuki, H.; Nagase, H. Bioorg. Med. Chem. 2011, 19, 1205.
9. Maloney Huss, K. E.; Portoghese, P. S. J. Org. Chem. 1990, 55, 2957.
10. Nelson, T. D.; Davis, R. D.; Nelson, W. L. J. Med. Chem. 1994, 37, 4270.
11. Davis, R. D.; Nelson, T. D.; Nelson, W. L. J. Heterocycl. Chem. 1994, 31, 1509.
12. Xu, H.; Partilla, J. S.; Wang, X.; Rutherford, J. M.; Tidgewell, K.; Prisinzano, T. E.;
Bohn, L. E.; Rothman, R. B. Synapse 2007, 61, 166.
13. Cheng, K.; Lee, Y. S.; Rothman, R. B.; Dersch, C. M.; Bittman, R. W.; Jacobson, A.
E.; Rice, K. C. J. Med. Chem. 2011, 54, 957.
14. X-ray crystallographic analysis, 7a: Colorless prisms of 7a were recrystallized
from a minimum volume of hot acetonitrile. The X-ray intensity data for 7a
were measured on
a Bruker Kappa Apex Duo diffractometer using high
brightness copper source with multi-layer mirrors. The frames were
IlS
integrated with the Bruker SAINT software package (Bruker, 2010). Data were
corrected for absorption effects using the multi-scan method (SADABS). The
structure was solved and refined by full-matrix least squares procedures on |F2|
using the Bruker SHELXTL Software Package. Hydrogen atoms in hydroxyl
groups were located and refined independently. All other hydrogen atoms
were included in idealized positions for structure factor calculations.
Anisotropic displacement parameters were assigned to all non-hydrogen
atoms. X-ray crystal data for 7a: C27H31NO4, Mr = 433.53, colorless prism,
0.08 Â 0.14 Â 0.20 mm, monoclinic, space group P21, a = 8.7942(4) Å,
b = 18.8792(9) Å, c = 13.1369(6) Å, b = 90.3310(10)°, V = 2181.05(17) Å3, Z = 4,
Acknowledgements
This Letter is dedicated to Professor Timothy Macdonald for his
many contributions to the field of Organic and Medicinal
Chemistry. The authors are indebted to Dr. Kenner C. Rice and
Kejun Chung of the Drug Design and Synthesis Section, National
Institute on Drug Abuse, National Institutes of Health, Bethesda,
MD, for providing a sample of KC-2-009 for use in functional bind-
ing assays. The authors are also grateful to Mallinckrodt, Inc. for
generously donating naltrexone free base. We thank Dr. Rebecca
Conry (Colby College, Waterville, ME) for valuable advice and assis-
tance in preparing X-ray quality crystals of 7a. We gratefully
acknowledge the support of the University of New England (UNE)
for College of Arts & Sciences (CAS) Research Mini-Grants and the
Chemistry & Physics Department for monies to support research
& conference travel (awarded to A.M.D.). We thank the Carman
Pettapiece Student Research Fund of the College of Osteophatic
Medicine (COM) at UNE as well as the CAS and COM Dean’s Offices
and the Provost’s office for research support (awarded to D.J.M./
A.M.D.). Portions of this work were supported by the Intramural
Research Program, National Institute on Drug Abuse, National
Institutes of Health.
T = 100(2) K,
q a radiation (k = 1.54178 Å). A total of
cacld = 1.320 g cmÀ3, CuK
24083 reflections were collected to a maximum h angle of 68.05° (0.83 Å
resolution), of which 7735 were independent (Rint = 0.0191). The final
anisotropic full-matrix least-squares refinement on F2 with 589 parameters
and 5 restraints converged at R1 = 0.0251, wR2 = 0.0646 for 7731 reflections
with I > 2r(I) and for all data (7735 reflections). The goodness-of-fit was 1.038.
CCDC-826389 contains the supplementary crystallographic data for structure
7a. Copies of these data can be obtained free of charge, on application, via
www.ccdc.cam.ac.uk/conts/retreiving.html
or
from
the
Cambridge
Crystallographic Data Center, 12, Union Road, Cambridge, CB2 1EZ, UK; fax:
+44 1223 336 033; or email: deposit@ccdc.cam.ac.uk.
15. Procedure for One Pot triflation/epoxy bridge formation to yield 17-
(cyclopropylmethyl)-4,5
round bottom flask equipped with
a
:6b,14-diepoxy-3-benzyloxymorphinan (4). To a 15 mL
spin vane was added 7a (102 mg,
a
0.24 mmol), anhydrous dichloromethane (2.5 mL) and anhydrous pyridine
(0.1 mL). The stirring homogenous solution was cooled to À35 °C in an
acetonitrile/dry
ice
bath,
and
to
the
solution
was
added
trifluoromethanesulfonic anhydride (133 mg, 0.47 mmol) via syringe in
a
dropwise fashion. A vigorous reaction ensued yielding a white suspension
which over the course of one hour resolved to a clear yellow solution. The
progress of the reaction was monitored by TLC (dichloromethane/acetone:
95:5) with intermediate triflate 8 possessing a Rf = 0.5. The solution was then
concentrated at 0 °C via rotary evaporation providing a clear yellow viscous
liquid. Residual solvent was removed by vacuum pump distillation. The crude
triflate was immediately dissolved in 2.5 mL of anhydrous tetrahydrofuran and
cooled to À10 °C. Next, a 1 M lithium triethylborohydride/tetrahydrofuran
(9.5 mmol) solution was added dropwise. The homogeneous solution was
stirred for 1 h, taking on a dark violet hue, and the progress of the reaction was
monitored by TLC (eluent dichloromethane/acetone: 95:5). The reaction was
quenched of excess lithium triethylborohydride with 20 mL ice cold acetone.
The stirring solution was then allowed to warm to room temperature at which
point it was concentrated via rotary evaporation to produce a viscous red-
brown liquid. The crude product was dissolved in 30 mL dichloromethane and
washed with deionized water (3 Â 50 mL). The organic solution was dried over
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.
06.056.
References and notes
1. (a) Urban, J. D.; Clarke, W. P.; Von Zastrow, M.; Nichols, D. E.; Kobilka, B.;
Weinstein, H.; Javitch, J. A.; Roth, B. L.; Christopoulos, A.; Sexton, P. M.; Miller,
K. J.; Spedding, M.; Mailman, R. B. J. Pharmacol. Exp. Ther. 2007, 320, 1; (b)
Mailman, R. B. Trends Pharmacol. Sci. 2007, 28(8), 390; (c) Greasley, P. J.;
Clapham, J. C. Eur. J. Pharmacol. 2006, 553, 1.
2. (a) Comer, S. D.; Collins, E. D.; Kleber, H. D.; Nuwayser, E. S.; Kerrigan, J. H.;
Fischman, M. W. Psychopharmacology 2002, 159, 351; (b) LaSalle, G.; Rachelska,
B.; Nedelec, L. Burns 2008, 34(6), 797; (c) Leri, F. Expert Opin. Pharmacother.
2008, 9, 1387; (d) Van Dorp, E. L. A.; Yassen, A.; Dahan, A. Expert Opin. Drug Saf.
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107, 41.
magnesium sulfate, vacuum filtered, and again concentrated to
a brown
viscous liquid. The crude product was purified by column chromatography
(dichloromethane/acetone: 95:5) yielding compound 4 as a white solid (93 mg,
96% yield, Rf = 0.1). 1H NMR (CDCl3, 300 MHz): d 7.28 to 7.45 (m, 5H); 6.76 (d,
1H, J = 7.9 Hz); 6.53 (d, 1H, J = 8.2 Hz); 5.18 (dd, 2H, J = 29.6, 12 Hz); 4.98 (t, 1H,
J = 5.2 Hz); 4.66 (d, 1H, J = 4.8 Hz); 3.75 (d, 1H, J = 6.5 Hz); 3.38 (d, 1H,
J = 17.5 Hz); 2.86 (dd, 1H, J = 12.2, 5.6 Hz); 2.66 (td, 1H, J = 13.1, 3.8 Hz); 2.54
(dd, 1H, J = 12.4, 5.8 Hz); 2.42 (dd, 1H, J = 6.9, 3.8 Hz); 2.37 (d, 1H, J = 6.5 Hz);
2.20 (td, 1H, J = 13.1, 5.8 Hz); 1.75 (dd, 1H, J = 13.4, 3.1 Hz); 1.69 to 1.40 (m,
2H); 1.05 (dd, 1H, J = 12.7, 4.8 Hz); 1.02 to 0.82 (m, 1H); 0.59 to 0.46 (m, 2H);
0.20 to À0.06 (m, 2H); À0.10 to À0.22 (m, 1H). 13C NMR including DEPT
assignments (CDCl3, 75 MHz): d 150.76 (quarternary), 142.33 (quarternary),
137.38 (quarternary), 135.65 (quarternary), 131.28 (quarternary), 128.56 (CH),
127.98 (CH), 127.70 (CH), 122.14 (CH), 117.59 (CH), 92.36 (CH), 89.19
(quarternary), 86.71 (CH), 77.55 (CDCl3), 77.12 (CDCl3), 76.70 (CDCl3), 72.16
(CH2), 59.80 (CH2), 57.37 (CH), 54.43 (quarternary), 43.38 (CH2), 31.67 (CH2),
30.87 (CH2), 30.51 (CH2), 21.64 (CH2), 9.33 (CH), 4.43 (CH2), 3.56 (CH2). HRMS-
ESI (m/z): [M+H]+ calculated for C27H30NO3: 416.2226; found: 416.2212.
16. Compound 4 was previously reported as a reaction byproduct (approximately
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