The first biomimetic synthesis of a diterpenoid with the ent-verrucosin A/B skeleton

Article abstract of DOI:10.1016/j.tetlet.2016.03.106

The first biomimetic synthesis of methyl (8S,9R,13R,14R)-4,4,8,9,13-pentamethyl-20(10 → 9)-abeo-ent-isocopal-5(10),11(12)-dien-15-oate - a diterpenoid with the ent-verrucosin A/B skeleton has been performed by electrophilic isomerization of methyl 12α-hydroxy-ent-isocopal-13(16)-en-15-oate. The structure and stereochemistry of the synthesized compound have been established on the basis of spectroscopic data.

Full text of DOI:10.1016/j.tetlet.2016.03.106

Accepted Manuscript
The first biomimetic synthesis of a diterpenoid with the ent-Verrucosin A/B
skeleton
Marina Grinco, Vladilena Gîrbu, Elena Gorincioi, Alic Barba, Veaceslav
Kulciţki, Nicon Ungur
PII:
S0040-4039(16)30335-5
DOI:
http://dx.doi.org/10.1016/j.tetlet.2016.03.106
TETL 47494
Reference:
Tetrahedron Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
14 December 2015
21 March 2016
31 March 2016
Please cite this article as: Grinco, M., Gîrbu, V., Gorincioi, E., Barba, A., Kulciţki, V., Ungur, N., The first
biomimetic synthesis of a diterpenoid with the ent-Verrucosin A/B skeleton, Tetrahedron Letters (2016), doi: http://
dx.doi.org/10.1016/j.tetlet.2016.03.106
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The first biomimetic synthesis of a diterpenoid
with the ent-Verrucosin A/B skeleton
Leave this area blank for abstract info.
Marina Grinco, Vladilena Gîrbu, Elena Gorincioi,
Alic Barba, Veaceslav Kulciţki, Nicon Ungur*

1
Tetrahedron Letters
The first biomimetic synthesis of a diterpenoid with the ent-Verrucosin A/B
skeleton
Marina Grinco, Vladilena Gîrbu, Elena Gorincioi, Alic Barba, Veaceslav Kulciţki, Nicon Ungur
Institutul de Chimie al Academiei de Ştiinţe a Moldovei, Chişinău, MD 2028, Republic of Moldova
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
The first biomimetic synthesis of methyl (8S,9R,13R,14R)-4,4,8,9,13-pentamethyl- 20(109)-
abeo-ent-isocopal-5(10),11(12)-dien-15-oate – a diterpenoid with the ent-verrucosin A/B
skeleton has been performed by electrophilic isomerization of methyl 12-hydroxy-ent-
isocopal-13(16)-en-15-oate. The structure and stereochemistry of the synthesized compound
have been established on the basis of spectroscopic data.
Available online
2015 Elsevier Ltd. All rights reserved.
Keywords:
Biomimetic synthesis;
Verrucosin A;
Diterpenoids;
Isomerization.
———
 Corresponding author. Tel.: +373-22-739-775; fax: +373-22-739-954; e-mail: nicon.ungur@gmail.com

2
Tetrahedron Letters
Terpenoids are one of the most numerous and important
property of terpenoids is their isomerization under the influence
of acidic reagents.3 In this paper we report the isomerization of
the methyl ester of 12-hydroxy-ent-isocopal-13(16)-en-15-oic
acid (1)4 under the influence of p-toluenesulfonic acid.
classes of natural compounds, their roles in living systems being
extensively revealed over the past three decades. They are
particularly widespread, e.g. in lower and higher plants,1
microorganisms and aquatic animals.2 An important chemical
Scheme 1. Reagents and conditions: (a) KMnO4, Me2CO, r.t., 12 h, 90%; (b) I2, PhMe, reflux, 3 h, 78%; (c) (MeO)2P(O)CH2CO2Me,
PhH, MeONa, reflux, 2 h, 98%, (13E:13Z = 10:1); (d) FSO3H (5 eq), i-PrNO2, -78 °C, 15 min, then NEt3, 92%; (e) m-CPBA, CH2Cl2, 0
°C, 12 h, 97%; (f) Al(Oi-Pr)3, PhMe, reflux, 24 h, 78%; (g) p-TsOH/CHCl3, reflux, 3 h, 71%; (h) LiAlH4, THF, reflux, 4 h, 98%.
(-)-Sclareol (2) served as a starting compound, which was
converted into the methyl ester of ent-isocopalic acid (3),
according to the described procedure.5 The latter compound was
in turn transformed into the methyl ester of 12-hydroxy-ent-
isocopal-13(16)-en-15-oic acid (1), according to the reported
two-step sequence6 (Scheme 1).
13C, DEPT-135) and 2D homo- (1H/1H COSY-45, 1H/1H
1
NOESY) and heteronuclear (1H/13C HSQC and H/13C HMBC)
correlation spectra (Figure 1). Thus, the 1H NMR spectrum
displayed singlets of four tertiary methyl groups: at  0.98 (3H,
H-17) and 0.994 (3H, H-20), geminal dimethyls at
andeach 3H, H-18 and H-19), signals of one
secondary methyl group at  0.88 (3H, H-16) that appears as a
doublet of multiplets with J = 6.7 Hz due to splitting of H-13, H-
12 and H-14 nuclei, a downfield singlet of the methyl ester group
at   as well as deshielded signals of two sp2 methines:
5.38 (dd, J = 10.1; 1.2 Hz, H-12) and 5.45 (br. d, J = 10.1, H-
11). The assignment of signals that characterize methylene
protons belonging both to ring A (complex splitting patterns H2-
1-H2-2-H2-3) and B (splitting patterns H2-6-H2-7) is depicted in
Table 1, being to a certain degree impeded by couplings and
signal overlapping.
The reaction of a chloroform solution of ester (1) with p-
toluenesulfonic acid at reflux for 4 h afforded a ~5:2 mixture of
two compounds. The polar minor compound was identified by its
spectroscopic data and other properties as the known diterpenic
lactone (4).4
According to elemental analysis, IR and NMR spectra, the
major compound (5)7 was identified as a diterpenic diene ester
with the molecular formula C21H32O2. The NMR data of
compound (5) were assigned (Table 1) on the basis of its 1D (1H,
a
Table 1. 1H (400.13 MHz) and 13C NMR (100.61 MHz) data of compound 5 in CDCl3
Position
H
C
Position
H
C
1ax
1eq
2
1.90 (1H, mb)
27.1, t
9
-
-
42.7, s
133.4, s
133.5, d
129.1, d
31.5, d
51.3, d
175.9, s
19.9, q
18.7, q
27.9, q
28.8, q
22.3, q
51.0, q
2.00 (1H, m, 6.8)
10
11
12
13
14
15
16
17
18
19
20
OMe
1.58 (2H, m)
20.0, t
39.8, t
5.45 (1H, br.d, 10.1)
5.38 (1H, dd, 10.1, 1.2)
2.48 (1H, mb)
2.48 (1H, mb)
-
3ax
3eq
1.36 (1H, td, 12.3, 3.6)
1.44 (1H, dddd, 12.5, 4.9, 3.3, 1.5)
4
-
33.9, s
131.5, s
20.8, t
5
-
0.88 (3H, m, 6.7)
0.97 (3H, s)
6ax
6eq
7ax
7eq
8
2.35 (1H, m)
1.90 (1H, mb)
0.986 (3H, s)
0.988 (3H, s)
0.994 (3H, s)
3.69 (3H, s)
1.65 (1H, ddd, 14.0, 12.5, 6.3)
1.23 (1H, dd, 14.0, 6.3)
-
30.1, t
36.8, s
a chemical shifts () are in ppm, coupling constants (J) are in Hz.
b overlapping signals.

3
spectrum, while the presence of the function was
especially proven by long-range correlations between H3-16/C-
12, H3-20/C-11, H-12/C-14, H-11 and H-12/C-9 and by mutual
HMBC correlations as indicated in Figure 1.
During elucidation of the stereochemistry at the C-13
asymmetric center of compound (5) a problem emerged, caused
by the overlapped signals of methines H-13 and H-14 that
1
appeared in the H NMR spectrum as a multiplet centered at
2.48. In light of this, interpretation of its 1H-1H NOESY
spectrum only furnished inconclusive data for the identification
of the configuration at C-13. Therefore, we decided to solve this
problem by correlation of the NMR data of compound (5) with
Figure 1. Key 2D NMR correlations for compounds (5) and (6).
1
that of its reduction product, compound (6).8 In the H NMR
The 13C NMR data (Table 1) exhibited twenty one carbon
signals, which were assigned by DEPT as six methyls, five sp3
methylenes, two sp3 and two sp2 methines, three sp3 and three
sp2, including one ester, quaternary carbons. The presenceoftwo
double bonds, one of which being disubstituted and another
tetrasubstituted, was corroborated by the 13C NMR data [C 133.5
(C-11), 129.1 (C-12), 131.5 (C-5) and 133.4 (C-10)]. The
rearranged carbon framework of compound (5) becomes obvious
with detailed analysis of its HMBC spectrum. Thus, the observed
correlations from both H-6 and H-1 to two sp2 hybridized
carbons (C-5, C 131.5 and C-10, C 133.4) were indicative of
localization, which was also supported by the correlations of
H3-19/C-5 and H3-18/C-5. The migration of the H3-20 methyl
from the C-10 to C-9 position was ascertained by the H3-20/C-10,
H3-20/C-9, H3-20/C-11 and H3-20/C-8 cross-peaks in the HMBC
spectrum of compound (6) signals of the H-13 and H-14 nuclei
were well-separated:  1.39 (dt, J = 10.5, 3.6 Hz, 1H, H-14ax)
and 2.03 (m, J= 10.5, 7.0, 1.6, 1.3 Hz, 1H, H-13ax). The large
coupling constant (10.5 Hz) between the vicinal methine protons
under discussion demonstrated their trans-diaxial interaction,
which was confirmed by molecular modelling.9 Finally, the (R)
relative configuration at C-13 was deduced from the NOESY
correlation between H3-17/H-13 and the lack of H3-17/H-14
correlation. Thus, compound (5) was definitively identified as
methyl (8S,9R,13R,14R)-4,4,8,9,13-pentamethyl- 20(109)-
abeo-ent-isocopal-5(10),11(12)-dien-15-oate.
Scheme 2. Proposed reaction mechanism for the acid-promoted isomerization of 1 into 5.
Based on these data, it was possible to confirm that the acid-
promoted isomerization of the methyl ester of 12-hydroxy-ent-
isocopal-13(16)-en-15-oic (1) acid furnished a diene ester with a
rearranged framework (5). A plausible path for its formation
involving the electrophilic isomerization of compound (1) is
depicted in Scheme 2.
with total facial selectivity of the protonation at C-13 seems less
probable.
There are also similar reports on terpene synthesis which have
been shown to occur via such rearrangements. For example
Urones and collaborators postulated formation of a carbonium
ion very similar to A upon treatment of the related epoxide with a
Lewis acid. Its subsequent conversion involved a skeletal
rearrangement via ring contraction.11
Thus, after protonation of ester 1 carbocation A is formed,
which through a 1,3-hydride shift (H-11 moves to C-13)
generates carbocation B.
A second hydride shift forms
carbocation C, which after angular methyl migration and
dehydration forms carbocation D, which is stabilized by
deprotonation to afford the methyl ester of (8S,9R,13R,14R)-
4,4,8,9,13-pentamethyl-20(109)-abeo-ent-isocopal-
5(10),11(12)-dien-15-oic acid (5).
The fact that this mechanistic explanation relies upon a rare
1,3-hydride shift10 forced us to consider other alternatives. For
example, an acid induced dehydration could compete with the
formation of intermediate A and bring about formation of a diene
7. However, subsequent re-protonation and rearrangement would
lead to a trisubstituted compound 8, the isomer of 5. Its
isomerisation to a less favored disubstituted olefin proceeding
Figure 2. Natural verrucosins A and B.12,13
It is noteworthy that the carbon skeleton of diterpenoid (5) has
ent-stereochemistry relatively to that of verrucosins A (9) and B
(10) - acylglycerols isolated for the first time from Doris
verrucosa mollusks collected in the Mediterranean Sea.12,13

4
Tetrahedron Letters
Diterpenoids (9) and (10) are highly ichthyotoxic12 and have
mg, 0.84 mmol), which was heated at reflux for 4 h. Then, the
mixture was washed twice with water, dried, and concentrated.
The crude reaction product (94 mg) was purified by silica gel
column chromatography (4 g, 1.5% AcOEt in petroleum ether) to
give methyl (8S,9R,13S,14R)-4,4,8,9,13-pentamethyl-20(109)-
abeo-ent-isocopal-5(10),11(12)-dien-15β-oate (5) (59 mg, 67%):
white gum; [α]D25 = 37.7 (c 1.33, CHCl3). IR liquid film (, cm-1):
1164, 1732. Anal. Calcd. for C21H32O2: C 79.70, H 10.19; found:
C 79.81, H 10.14. 1H and 13C NMR see Table 1.
demonstrated in vivo bioactivity as morphogens in the Hydra
tentacle regeneration assay and their parallel function as
activators of rat brain protein kinase C was also described.14
It should also be noted that the synthesis of terpenoids with
the verrucosin A and B carbon skeleton has not yet been
achieved. The biogenetic pathway leading to 9 and 10 has been
previously suggested15 and the present communication helps to
confirm it with additional synthetic data.
8.
(8S,9R,13S,14R)-4,4,8,9,13-Pentamethyl-20(109)-abeo-ent-
isocopal-5(10),11(12)-dien-15β-ol (6). A solution of ester (5) (10
mg, 0.032 mmol) in anhydrous THF (5 mL) was treated with
LiAlH4 (15 mg, 0.40 mmol) under stirring. After 3 h at reflux, the
reaction was cooled and quenched with ethyl acetate (0.1 mL).
The reaction product was subsequently treated with a 10% H2SO4
solution (2 mL), extracted with diethyl ether (3x2 mL) and washed
with brine, sat. NaHCO3 solution and brine. The diethyl ether
extract was dried over Na2SO4 for 2 h and then evaporated in
vacuo. The crude product was purified by flash chromatography
(silica gel (0.2 g, 1.5% AcOEt in petroleum ether) to give alcohol
In conclusion - the first biomimetic synthesis of methyl
(8S,9R,13R,14R)-4,4, 8,9,13-pentamethyl- 20(109)-abeo-ent-
isocopal-5(10),11(12)-dien-15-oate – a diterpenoid with the ent-
verrucosin A/B skeleton has been performed by electrophilic
isomerization of methyl 12-hydroxy-ent-isocopal-13(16)-en-15-
oate. The presented method opens the possibility of the synthesis
of bioactive diterpenoids possessing this unique rearranged
framework.
25
6 (9 mg, 98%). Colorless viscous oil. [α]D = 31.5 (c 0.61,
CHCl3). IR liquid film (, cm-1): 1360, 1460, 2925. 3375. 1H
NMR (400 MHz, CDCl3, H): 0.75 (s, 3H, H-17), 0.86 (s, 3H, H-
19), 0.907 (s, 3H, H-20), 0.91 (s, 3H, H-18), 0.99 (d, J = 7.0 Hz,
3H, H-16), 1.26 (td, J = 12.5, 3.3 Hz, 1H, H-3ax), 1.36 (dddd, J =
12.5, 5.1, 3.3, 1.3 Hz, 1H, H-3eq),1.39 (dt, J = 10.5, 3.6 Hz, 1H, H-
14ax), 1.43 (br. s, OH), 1.47 (m, 2H, H-2), 1.55 (m, J = 14.0, 12.6,
5.9 Hz, 1H, H-7ax), 1.63 (ddd, J = 14.0, 5.9, 1.7 Hz, 1H, H-7eq),
1.87 (m, 4H, H-1 and H-6), 2.03 (m, J = 10.5, 7.0, 1.6, 1.3 Hz, 1H,
H-13), 3.66 (dd, J = 11.5, 3.3 Hz, 1H, H-15A), 3.82 (dd, J = 11.5,
3.8 Hz, 1H, H-15B), 5.31 (dd, J = 10.1, 1.6 Hz, 1H, H-12), 5.35
(dd, J = 10.1, 1.3 Hz, 1H, H-11). 13C NMR (100 MHz, CDCl3, C):
19.6 (q, C-17), 20.0 (t, C-2), 20.2 (q, C-16), 21.2 (t, C-6), 22.5 (q,
C-20), 27.2 (t, C-1), 27.8 (q, C-18), 28.6 (t, C-7), 28.9 (q, C-19),
31.1 (d, C-13), 33.9 (s, C-4), 36.4 (s, C-8), 39.8 (t, C-3), 43.3 (s,
C-9), 45.0 (d, C-14), 62.9 (s, C-15), 130.6 (d, C-12), 131.2 (s, C-
5), 133.3 (s, C-11), 134.6 (d, C-10).
Acknowledgement
The financial support from Supreme Council for Science and
Technology Development of the Republic of Moldova (Project
"Elaboration of methods for obtaining valuable terpenoids by
valorization of renewable resources from the Republic of
Moldova, 2015-2018", No 15.817.02.14A) is gratefully
acknowledged.
References and Notes
1.
2.
(a) Hanson, J. R. Nat. Prod. Rep. 2015, 32, 1654-1663 and
previous reviews of this series; (b) González, M. A. Nat. Prod.
Rep. 2015, 32, 684-704.
9.
The calculated value of the J13ax,14ax coupling constant was 11.9
Hz. For molecular modeling with energy minimization, PERCH
NMR TOOLS (version 2014.1) software was used.
(a) Blunt, J. W.; Copp, B. R.; Keyzers, R. A.; Munro, M. H. G.;
Prinsep, M. R. Nat. Prod. Rep. 2015, 32, 116-211 and previous
reviews of this series; (b) Xie, Y.; Wright, S.; Shen, Y.; Du, L.
Nat. Prod. Rep. 2012, 29, 1277-1287; (c) Berrue, F.; Kerr, R.G.
Nat. Prod. Rep. 2009, 26, 681-710; (d) Keyzers, R. A.; Northcote,
P. T.; Davies-Coleman, M. T. Nat. Prod. Rep. 2006, 23, 321–334.
Barkhash, V. A.; Polovinka, M. P. Russ. Chem. Rev. 1999, 68,
393-414.
10. (a) Kim, S.-H.; Heo, K.; Chang, Y.-J.; Park, S.-H.; Rhee, S.-K.;
Kim, S.-U. J. Nat. Prod. 2006, 69, 758–762; (b) For a review on
biosynthesis via carbocations see: Tantillo, D. J. Nat. Prod. Rep.
2011, 28, 1035-1053.
11. Basabe, P.; Diego, A.; Díez, D.; Marcos, I. S.; Mollinedo, F.;
Urones, J.G. Synthesis 2002, (11), 1523-1529.
12. Cimino, G.; Gavagnin, M.; Sodano, G.; Puliti, R.; Mattia, C. A.;
Mazzarella, L. Tetrahedron 1988, 44, 2301-2310.
13. Zubia, E.; Gavagnin, M.; Crispino, A.; Martinez, E.; Ortea, J.;
Cimino, G. Experientia 1993, 49, 268-271.
14. (a) De Petrocellis, L.; Di Marzo, V.; Arca, B.; Gavagnin, M.;
Minei, R.; Cimino, G. Comp. Biochem. Physiol. 1991, 100C, 603-
607; (b) De Petrocellis, L.; Orlando, P.; Gavagnin, M.; Ventriglia,
M.; Cimino, G.; Di Marzo, V. Experientia 1996, 52, 874-877.
15. Gavagnin, M.; Ungur, N.; Castelluccio, F.; Cimino, G.
Tetrahedron 1997, 53, 1491-1504.
3.
4.
5.
Imamura, P. M.; Sierra, M. G.; Ruveda, E. A. J. Chem. Soc.,
Chem. Comm. 1981, 734-735.
(a) Hua, S.-K.; Wang, J.; Chen, X.-B.; Xu, Z.-Y.; Zeng, B.-B.
Tetrahedron 2011, 67, 1142-1144; (b) Vlad, P. F.; Ungur, N.;
Nguen, V. T. Russ. Chem. Bull. 1995, 44, 2404-2411.
Mischne, M. P.; Sierra, M. G.; Ruveda, E. A. J. Org. Chem. 1984,
49, 2035–2037.
6.
7.
Methyl (8S,9R,13R,14R)-4,4,8,9,13-pentamethyl- 20(109)-
abeo-ent-isocopal-5(10),11(12)-dien-15-oate (5). To a solution of
1 (95 mg, 0.28 mmol) in CHCl3 (35 mL) was added p-TsOH (145

3
Graphical Abstract
To create your abstract, type over the instructions in the template box below.
Fonts or abstract dimensions should not be changed or altere
The first biomimetic synthesis of a diterpenoid
with the ent-Verrucosin A/B skeleton
Leave this area blank for abstract info.
Marina Grinco, Vladilena Gîrbu, Elena Gorincioi,
Alic Barba, Veaceslav Kulciţki, Nicon Ungur*

4
Tetrahedron Letters
The first biomimetic synthesis of a diterpenoid with the ent-Verrucosin A/B skeleton
Marina Grinco, Vladilena Gîrbu, Elena Gorincioi, Alic Barba, Veaceslav Kulciţki, Nicon Ungur*
Highlights



The first biomimetic synthesis of a diterpenoid with the ent-verrucosin A/B skeleton has been performed.
The method opens the possibility of synthesis of bioactive diterpenoids possessing this rearranged framework.
The structure and stereochemistry of the synthesized compound have been established on the basis of spectral
data.