Bicunningines A and B, two new dimeric diterpenes from Cunninghamia Lanceolata

Article abstract of DOI:10.1021/ol202915r

Two unprecedented dimeric diterpenoids, with a 2,3-dihydrofuran ring fusing an abietane and a 4,5-seco-abietane diterpene, were isolated from Cunninghamia lanceolata. Their structures were elucidated by spectroscopic measurements, and their absolute confi

Full text of DOI:10.1021/ol202915r

ORGANIC
LETTERS
2012
Vol. 14, No. 2
460–463
Bicunningines A and B, Two New Dimeric
Diterpenes from Cunninghamia lanceolata
†
†
‡
‡
†
ꢀ
ꢀ
Xian-Feng Hou, Sheng Yao, Attila Mandi, Tibor Kurtan, Chun-Ping Tang,
Chang-Qiang Ke,† Xi-Qiang Li,† and Yang Ye*,†
State Key Laboratory of Drug Research and SIMM-CUHK Joint Research Laboratory
of Promoting Globlization of Traditional Chinese Medicines, Shanghai Institute of
Materia Medica, Chinese Academy of Sciences, 555 Zu-Chong-Zhi Road, Zhangjiang
Hi-Tech Park, Shanghai 201203, China, and Department of Organic Chemistry,
University of Debrecen, POB 20, H-4010 Debrecen, Hungary
yye@mail.shcnc.ac.cn
Received October 31, 2011
ABSTRACT
Two unprecedented dimeric diterpenoids, with a 2,3-dihydrofuran ring fusing an abietane and a 4,5-seco-abietane diterpene, were isolated from
Cunninghamia lanceolata. Their structures were elucidated by spectroscopic measurements, and their absolute configurations were determined by
quantum chemical TDDFT ECD calculations, chemical transformations, and Mosher’s method. The Mosher method carried out with MPA and MTPA
esters of the sterically hindered sec-hydroxyl group gave contradictory results, while MPA afforded the correct absolute configuration.
The genus Cunninghamia (Taxodiaceae) consists of two
species and two variants, which are all distributed in
China.1 Cunninghamia lanceolata (Lamb.) Hook. is a
traditional Chinese medicine used for the treatment of
hernia, arthritis, and strangury. Previous investigations
resulted in the identification of bislabdane,2ꢀ4 lignans,5,6
diterpenoids,4 and bisflavonoids7 from its heartwoods,
roots, and leaves. As a continuation of our efforts to
explore new structures, we performed a study on this plant
and separated two new dimeric diterpenoids, bicunning-
ines A (1) and B (2) (Figure 1), which featured an abietane
and a 4,5-seco-abietane diterpene moiety fused by a 2,3-
dihydrofuran ring. In this paper, the isolation and struc-
tural elucidation of these two compounds are reported.
The air-dried barks of C. lanceolata (13.5 kg)8 were
ground into powder and percolated with acetone at
room temperature 3 times (7 days each). The combined
† Shanghai Institute of Materia Medica.
‡ University of Debrecen.
(1) Chen, S. K.; Chen, B. Y.; Li, H. Flora Reipublicae Popularis
Sinicae (Zhongguo Zhiwu Zhi); Science Press: Beijing, China, 1997; Vol.
43, pp 285ꢀ286.
(2) Irie, H.; Miyashita, M.; Kouno, I.; Hamanaka, N.; Sugioka, M.
Tetrahedron Lett. 1992, 33, 5671–5672.
(3) Du, J.; Chen, R. Y.; Yu, D. Q. J. Nat. Prod. 1999, 62, 1200–1201.
(4) Du, J.; Wang, M. L.; Chen, R. Y.; Yu, D. Q. Planta Med. 2001, 67,
542–547.
(5) Lee, T. H.; Yeh, M. H.; Chang, C. I.; Lee, C. K.; Shao, Y. Y.;
Kuo, Y. H. Biosci., Biotechnol., Biochem. 2007, 71, 2075–2078.
(6) Lee, C. K.; Yeh, M. H.; Lee, T. H.; Chiu, H. L.; Kuo, Y. H. Helv.
Chim. Acta 2009, 92, 1983–1989.
(7) Ansari, F. R.; Ansari, W. H.; Ragman, W. J. Indian Chem. Soc.
1985, 62, 406–408.
(8) The barks of C. lanceolata were collected in December, 2009, in
Gongcheng County of Suining, Hunan province, China, and identified
by Professor Jingui Shen from Shanghai Institute of Materia Medica. A
voucher (2001024) was deposited at the herbarium of Shanghai Institute
of Materia Medica, Chinese Academy of Sciences.
r
10.1021/ol202915r
Published on Web 12/27/2011
2011 American Chemical Society

percolates were concentrated under reduced pressure, af-
fording a brown syrup. The syrup was then suspended in
hot water and partitioned successively with petroleum
ether (PE), CH2Cl2, and EtOAc to give PE (15 g), CH2Cl2
(160 g), and EtOAc (470 g) extracts, respectively. The
CH2Cl2 extract was subjected to column chromatography
(CC) (100ꢀ200 mesh) over silica gel and eluted with a
gradient of PE/acetone to yield fractions 1ꢀ10. Fraction 3
was separatedoveranMCI gel columnelutedwithMeOH/
H2O (40%-95%), affording subfractions AꢀE. Subfrac-
tion E was subjected to CC over silica gel (200ꢀ300 mesh)
eluted gradiently with CHCl3/MeOH (from 100:1 to 20:1),
and then Sephadex LH-20 (CHCl3/MeOH 1:1), and finally
purified by preparative HPLC to give 1 (5.6 mg). In a
similar way, compound 2 (14 mg) was isolated from
fraction 10 by CC over Sephadex LH-20 using CHCl3/
MeOH (1:1) as solvent system and then preparative
HPLC.
Compound 1 was obtained as a white powder:
[R]2D3 = þ272 (c = 0.09, MeOH).9 It had a molecular
formula of C40H50O4 as determined by HREIMS at m/z
594.3732 [M]þ (calcd 594.3709), indicating 16 degrees of
unsaturation. The IR spectrum shows absorption bands of
hydroxyl (3432 cmꢀ1) and carbonyl (1709 cmꢀ1) groups.
The 13C and DEPT NMR spectra (Table 1) revealed 40
resonances ascribed to 10 methyls, 4 methylenes, 11
1
methines, and 15 quaternary carbons. The H NMR spec-
trum (Table 1) displayed signals of five aromatic protons at
δH 7.17 (d, 8.1 Hz, 1H), 7.61 (d, 8.1 Hz, 1H), 7.51 (s, 1H), 6.63
(s, 1H), 6.55 (s, 1H), a benzylic angular methyl at 2.50 (s, 3H),
and three isopropyl groups at 3.05 (sept, 6.9 Hz, 1H), 1.27 (d,
6.9 Hz, 6H); 2.52 (sept, 6.9 Hz, 1H), 1.03 (d, 6.9 Hz, 3H), 1.04
(d, 6.9 Hz, 3H); and 2.88 (sept, 6.9 Hz, 1H), 0.87 (d, 6.9 Hz,
3H), 0.69 (d, 6.9 Hz, 3H). These data, together with addi-
tional analyses of 2D NMR spectra, revealed the presence of
two abietane-type diterpenoid moieties.
In one moiety, characteristic signals of a tetra-substituted
benzyl [δH 6.63 (s, 1H), 6.55 (s, 1H), δC 109.3, 125.9, 127.6,
131.5, 151.5], an isopropyl group [δH 2.88, 0.87, 0.69, δC
26.3, 22.0, 22.1], a carbonyl (δC 215.2) and an oxygenated
methine (δH 5.29, dd, 7.4, 8.1 Hz, 1H; δC 83.1) suggested
the presence of a typical abietane-type diterpenoid skele-
ton. HMBC correlations (Figure 2) from two geminal
methyls to the carbonyl carbon indicated that the carbonyl
was assigned as C-3; correlations from the oxygenated
methine to two quaternary carbons (one aromatic at δC
127.6, one connected with two geminal methyls at δC 48.2)
revealed that the oxygenated position was at C-6.
1
Figure 2. Selected HMBC and 1Hꢀ H COSY for compound 1.
Other resonances, including a penta-substituted naph-
thyl [δH 7.17 (d, 8.1 Hz, 1H), 7.61 (d, 8.1 Hz, 1H), 7.51
(s, 1H), δC 118.0, 126.3, 126.8, 127.5, 129.9, 130.1, 132.0,
132.3, 134.1, 157.1], two isopropyl groups [δH 2.52
(sept, 6.9 Hz, 1H), 1.03(d, 6.9 Hz, 3H), 1.04(d, 6.9 Hz,
3H), δC 41.1, 18.0, 18.3 ], [δH 3.05 (sept, 6.9 Hz, 1H), 1.27
(d, 6.9 Hz), 1.27 (d, 6.9 Hz), δC 28.6, 21.4, 22.5], a carbonyl
(δC 214.1), and an aromatic methyl [δH 2.50, s, 3H; δC
20.7], constructed a 4,5-seco-abietane moiety. The side
chain ꢀCH2ꢀCH2ꢀCOꢀCH(CH3)2, assigned to C-100,
was interpreted by its COSY and HMBC correlations
(Figure 2). The aromatic methyl was attached to C-50
Figure 1. Structures of compounds 1 and 2 and the truncated
model compound of 1 used for the TDDFT ECD calculation.
(9) Bicunningdione A (1): white powder, [R]2D3 = þ272 (c = 0.09,
MeOH), CD (MeOH, λ [nm] (Δε), c = 1.34 ꢁ 10ꢀ3); 342 (ꢀ3.09), 326sh
(ꢀ1.72), 308sh (1.39), 292sh (2.58), 279sh (3.91), 248 (67.67), 240sh
(48.58), 218 (30.36), 199 (ꢀ70.26); IR (film) vmax 3432, 2871, 1709, 1622,
1504, 1464, 1414 cmꢀ1; UV (MeOH) λmax (log ε) 342 (3.31), 328 (3.26),
304 (3.50), 281 (3.63), 245 (4.49) nm; 1H NMR (400 MHz, CDCl3) and
13C NMR (100 MHz, CDCl3) data, see Table 1; EIMS m/z 594 [M]þ;
HREIMS m/z 594.3732 [M]þ (calcd for C40H50O4, 594.3709).
Org. Lett., Vol. 14, No. 2, 2012
461

according to the HMBC correlation from H-60 to C-100
and C-200.
correlations from H-150 to C-120 rather than C-110 further
supported such a proposed structure. The relative config-
uration of 1 was inferred from the ROESY experiment,
which showed effects for H-6/H3-20, H-7/H3-20, and
H-5/H3-18, suggesting that H-6, H-7, and H3-20 are or-
iented on the same face, while H-5 and H3-18 on the other
face of the molecule. Accordingly, the structure of 1 was
established as shown in Figure 1 and named bicunningine A.
Compound 2, orange powder,10 [R]D23 = þ320 (c = 0.14,
MeOH), possessed a molecular formula of C40H52O4 as
determined by HRESIMS at m/z 595.3799 [M ꢀ H]ꢀ
(calcd 595.3787), corresponding to 15 degrees of unsatura-
tion. Detailed analyses of 1D and 2D NMR data of 2
(Table 1) revealed a high similarity between the structures
of 1 and 2. In comparison with 1, 2 showed two additional
protons in the molecular formula, which implied one
degree of unsaturation less for compound 2. This corrobo-
rated the fact that, in the 1H and 13C NMR spectra of 2,
one oxygenated methine (δH 3.41, δC 78.7) instead of a
carbonyl (δC 215.2) of 1 was observed, suggesting that an
oxygenated methine took place at C-3 in the molecule of 2.
HMBC correlations from H-3 (δH 3.41) to the C-1 (δC
36.9), C-18 (δC 28.3), and C-19 (δC 16.2) confirmed the
hydoxyl located at C-3. The ROESY correlations of
H-3/H3-18 and H-3/H-5 further indicated that H-3 was
on the same face with H-5 and H3-18. Thus, compound 2
was deduced to be a 3-hydroxyl analogue of 1. This struc-
ture was confirmed by the chemical transformation of 2 to
1 by Jones reagent (see details in the Supporting Informa-
tion). Therefore, the structure of 2 was established and
named bicunningine B.
Table 1. NMR Data of Compounds 1 and 2 (δ in ppm, J in Hz)
1
2
no.
1
δH
δC
δH
δC
R, 2.06, dd (13.8, 3.4)
β, 2.51, m
R, 2.54, m
37.6
R, 1.79, m
β, 2.19, m
R, 1.93, m
β, 3.11, m
36.9
2
34.4
27.4
β, 3.03, m
3
215.2
48.2
54.5
83.1
46.3
127.6
145.7
36.7
109.3
151.5
131.5
125.9
26.3
22.0
22.1
25.6
22.6
19.9
41.1
24.1
3.41, dd (9.5, 6.0)
78.7
39.8
52.8
84.1
46.2
127.6
147.0
36.9
109.3
151.5
131.2
125.7
26.3
22.1
22.2
28.3
16.2
20.9
41.1
24.0
4
5
2.15, d (7.4)
1.76, d (8.3)
6
5.29, dd (7.4, 8.1)
4.91,d (8.1)
5.23, dd (8.3, 7.8)
4.76, d (7.8)
7
8
9
10
11
12
13
14
15
16
17
18
19
20
10
6.63, s
6.63, s
6.55, s
6.50, s
2.88, sept (6.9)
0.87, d (6.9)
0.69, d (6.9)
1.35, s
2.89, sept (6.8)
0.86, d (6.8)
0.69, d (6.8)
1.28, s
1.42, s
1.17, s
1.55, s
1.30, s
2.67, m
2.66, m
20
R, 3.12, m
β, 3.37, m
R, 3.11, m
β, 3.35, m
30
40
214.1
41.1
214.1
41.0
2.52, sept (6.9)
2.51, sept (6.9)
50
134.1
126.3
127.5
132.3
130.1
129.9
118.0
157.1
132.0
126.8
28.6
134.0
126.2
127.4
132.4
130.2
129.8
118.5
157.3
132.0
126.6
28.6
60
7.17, d (8.1)
7.61, d (8.1)
7.15, d (8.2)
7.60, d (8.2)
70
80
90
100
110
120
130
140
150
160
170
180
190
200
7.51, s
7.50, s
3.05, sept (6.9)
1.27, d (6.9)
1.27, d (6.9)
1.04, d (6.9)
1.03, d (6.9)
2.50, s
3.11, sept (6.9)
1.27, d (6.9)
1.28, d (6.9)
1.01, d (6.9)
1.03, d (6.9)
2.48, s
21.4
21.3
22.5
22.7
18.3
18.4
18.0
18.0
20.7
20.7
Figure 3. Two overlapped solution conformers of the truncated
model compound of bicunningines A (1) with Boltzmann pop-
ulations of 58.4 and 40.8% indicating that they differ only in the
orientation of the phenolic proton.
Thus, two moieties were fully established with a total of
15 degrees of unsaturation. Considering the overall for-
mulaof 1, themissing onedegreeof unsaturationshouldbe
ascribed to the linkage between these two subunits creating
an additional ring. HMBC correlations from H-7 to C-110
and C-120 confirmed a CꢀC linkage between C-7 and
C-110. The remaining oxygenated methine at C-6 and the
oxygenated aromatic quaternary carbon of C-120 sug-
gested that there must be an oxygen bridge between C-6
and C-120, which constructeda 2,3-dihydrofuran moiety as
a connection between the two diterpenoid subunits. HMBC
(10) Bicunningdione B (2): light yellow powder, [R]2D3 = þ320 (c =
0.14, MeOH), CD (MeOH, λ [nm] (Δε), c = 1.34 ꢁ 10ꢀ3); 342 (ꢀ2.67),
326sh (ꢀ1.18), 307sh (1.76), 292sh (3.44), 281 (4.77), 248 (63.18), 240sh
(44.26), 219 (28.59), 199 (ꢀ73.97); IR (film) vmax 3430, 2962, 2931, 2871,
1704, 1623, 1461 cmꢀ1; UV (MeOH) λmax (log ε) 342 (3.22), 329 (3.17),
280 (3.65), 246 (4.43) nm; 1H NMR (400 MHz, CDCl3) and 13C NMR
(100 MHz, CDCl3) data, see Table 1; ESIMS m/z 597.2 [M þ H]þ, 619.2
[M þ Na]þ, 595.3 [M ꢀ H]ꢀ; HRESIMS m/z 595.3799 [M ꢀ H]ꢀ (calcd
for C40H51O4, 595.3787).
462
Org. Lett., Vol. 14, No. 2, 2012

In order to determine the absolute configurations of
these two compounds, electronic circular dichroism (ECD)
spectra of 1 and 2 were measured, which were almost
identical.9,10 For 1, weak negative Cotton effect (CE)
was observed at 342 nm, an intense positive one at
248 nm with positive shoulders on both sides, and a negative
CE at 199 nm in methanol. For the conformational
analysis of 1, its alkyl side chains were truncated to methyls
and the model compound 3 with (5S,6R,7S,10R) absolute
configuration was analyzed. The MMFF conformational
search of 3 followed by DFT reoptimizations afforded two
slightly different conformers (Figure 3) in chloroform, the
ECD calculations of which were performed with different
functionals (B3LYP, BH&HLYP, PBE0) and TZVP basis
set. All the functionals gave consistently mirror image
ECD curves of the experimental ECD spectrum, with PBE0
showing the best agreement (Figure 4), which allowed
determining the absolute configuration as (5R,6S,7R,-
10S). Since compound 2 showed nearly the same ECD
spectrum and they share the same chromophoric system,
its absolute configuration was determined as (3S,5R,6R,-
7R,10S) based on the known relative configuration of C-3.
It has to be noted that, regarding the corresponding
chirality centers, 2 is homochiral with 1 and thus its (6R)
configuration is only due to the change in the priority
orders.
Parallel with ECD calculations, diastereomeric MTPA
(R-methoxy-R-trifluoromethylphenylacetic acid) esters of
2 were prepared to use the Mosher’s method for the
configurational assignment of C-3. Although the Mosher’s
esters showed consistently positive and negative chemical
shift difference values for the protons flanking the C-3
chirality center (see Figure S1 in the Supporting Infor-
mation), surprisingly their values suggested opposite ab-
solute configuration to that determined by ECD. It was
supposed that higher-energy conformers of MTPA esters
had a larger contribution to the chemical shift difference
due to the presence of the adjacent two methyl groups,
which ultimately led to the wrong assignment. Then
diastereomeric MPA (methoxyphenylacetic acid) esters
of 2 were also prepared,11,12 the results of which were in
accordance with the absolute configuration obtained by
ECD (Figure S2). The present case confirms that the
widely used MTPA Mosher esters may lead to wrong as-
signment on sterically crowded chirality centers and MPA
esters are superior to them as also suggested by Riguera
and co-workers.13
Figure 4. Solution ECD spectrum of bicunningine A (1) in
methanol compared with the PBE0/TZVP computed Boltzmann-
weighted ECD spectrum of the truncated model compound,
(5S,6R,7S,10R)-3. Bars represent the rotatory strength of the
low-energy conformer.
linkage between the two halves. Previous reports disclosed
the presence of bislabdanes from this plant, which repre-
sented compounds containing two labdane diterpenoids
coupled by a DielsꢀAlder reaction.2ꢀ4 Other diterpenoid
dimers were also reported from other species of Taxodia-
ceae, such as sugikurojins B and C and sugikurojin J from
Cryptomeria japonica.14,15 The recent finding reveals a new
type of linkage of two diterpenoid moieties and expands
our knowledge on the possible connection between diter-
penoids in this family.
Acknowledgment. Y.Y. thanks the financial support
of the National Science & Technology Major Project
“Key New Drug Creation and Manufacturing Program”
(No. 2009ZX09301-001). Y.Y.’s thanks are also given to
the National Natural Science Funds for Distinguished
Young Scholar (No. 30925043) and grants from the
Ministry of Science and Technology (2010DFA30980) and
the Shanghai Commission of Science and Technology
ꢀ
(10DZ1972700). T.K. thanks the TAMOP 4.2.1./B-09/1/
KONV-2010-0007 and National Information Infrastruc-
ture Development Institute (NIIFI 10038).
Supporting Information Available. Experimental sec-
tion and NMR spectra of the new compounds. This
material is available free of charge via the Internet at
http://pubs.acs.org.
To the best of our knowledge, this is the first report
of dimeric diterpenes containing a 2,3-dihydrofuranyl
~ ꢀ
(11) Seco, J. M.; Quinoa, E.; Riguera, R. Chem. Rev. 2004, 104, 17–
117.
(14) Arihara, S.; Umeyama, A.; Bando, S.; Imoto, S.; Ono, M.; Tani,
M.; Yoshikawa, K. Chem. Pharm. Bull. 2004, 52, 354–358.
(15) Yoshikawa, K.; Suzuki, K.; Umeyama, A.; Arihara, S. Chem.
Pharm. Bull. 2006, 54, 574–578.
(12) Word, D. E.; Rhee, C. K. Tetrahedron Lett. 1991, 32, 7165–7166.
~ ꢀ
(13) Latypov, S. K.; Seco, J. M.; Quinoa, E.; Rigvera, R. J. Org.
Chem. 1996, 61, 8569–8577.
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