Isocoumarin derivatives and benzofurans from a sponge-derived Penicillium sp. fungus

Article abstract of DOI:10.1021/np3007556

Ten new fungal metabolites, including three hydroisocoumarins, penicimarins A-C (1-3), three isocoumarins, penicimarins D-F (6-8), and four benzofurans, penicifurans A-D (11-14), together with four known isocoumarin derivatives (4, 5, 9, 10), were obtained from the sponge-derived fungus Penicillium sp. MWZ14-4, collected from the South China Sea. Their planar structures and relative configurations were elucidated by detailed analysis of spectroscopic data and by comparison with related known compounds. The absolute configurations of 1-4 were assigned by the modified Mosher's method and TDDFT ECD calculations together with comparison of their CD spectra. Compound 1 represents a rare naturally occurring isocoumarin derivative with 4-substitution, but no substituent at the 3-position. These compounds were evaluated for antibacterial activities and cytotoxic activities in vitro. Among them, penicifuran A (11) exhibited inhibitory activity against Staphylococcus albus with an MIC value of 3.13 μM.

Full text of DOI:10.1021/np3007556

Article
pubs.acs.org/jnp
Isocoumarin Derivatives and Benzofurans from a Sponge-Derived
Penicillium sp. Fungus
Jun Qi,† Chang-Lun Shao,† Zhi-Yong Li,‡ Li-She Gan,§ Xiu-Mei Fu,† Wen-Tao Bian,† Hong-Ying Zhao,†,‡
and Chang-Yun Wang*,†
†Key Laboratory of Marine Drugs, The Ministry of Education of China, School of Medicine and Pharmacy, Ocean University of
China, Qingdao 266003, People’s Republic of China
‡Marine Biotechnology Laboratory, State Key Laboratory of Microbial Metabolism, School of Life Sciences & Biotechnology,
Shanghai Jiao Tong University, Shanghai 200240, People’s Republic of China
§Institute of Modern Chinese Medicine, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310058, People’s
Republic of China
S
* Supporting Information
ABSTRACT: Ten new fungal metabolites, including three
hydroisocoumarins, penicimarins A−C (1−3), three isocou-
marins, penicimarins D−F (6−8), and four benzofurans,
penicifurans A−D (11−14), together with four known
isocoumarin derivatives (4, 5, 9, 10), were obtained from
the sponge-derived fungus Penicillium sp. MWZ14-4, collected
from the South China Sea. Their planar structures and relative
configurations were elucidated by detailed analysis of
spectroscopic data and by comparison with related known
compounds. The absolute configurations of 1−4 were assigned
by the modified Mosher’s method and TDDFT ECD calculations together with comparison of their CD spectra. Compound 1
represents a rare naturally occurring isocoumarin derivative with 4-substitution, but no substituent at the 3-position. These
compounds were evaluated for antibacterial activities and cytotoxic activities in vitro. Among them, penicifuran A (11) exhibited
inhibitory activity against Staphylococcus albus with an MIC value of 3.13 μM.
RESULTS AND DISCUSSION
arine microorganisms, especially marine fungi, have
■
Mproven to be a rich source of structurally novel and
Penicimarin A (1) was isolated as a colorless, amorphous
powder with the molecular formula assigned as C11H12O5 on
the basis of its HREIMS data, indicating six degrees of
biologically active secondary metabolites, which have become
significant resources for drug discovery.1,2 Marine-derived fungi
in the genus Penicillium produce various bioactive metabolites,
such as cytotoxic citrinadin A,3 antioxidant terrestrols,4
antifungal and antibacterial scalusamide A,5 and antileukemic
sorbicillactone A,6 and have drawn the attention of both
pharmaceutical and natural product chemists. During our
ongoing search for bioactive metabolites from marine fungi in
the South China Sea,7−10 a sponge-derived fungus, Penicillium
sp. MWZ14-4, attracted our attention because an EtOAc extract
of the fungal culture exhibited antibacterial activity. Bioassay-
guided fractionation of the active extract led to the isolation of
10 new compounds including three hydroisocoumarins,
penicimarins A−C (1−3), three isocoumarins, penicimarins
D−F (6−8), and four benzofurans, penicifurans A−D (11−
14), together with four known isocoumarin derivatives,
aspergillumarin B (4),11 aspergillumarin A (5),11 sescandelin
B (9),12 and 5,6,8-trihydroxy-4-(1′-hydroxyethyl)isocoumarin
(10).12 Herein we report the isolation, structure elucidation,
and biological activities of these compounds.
unsaturation. Detailed inspection of the H NMR and 13C
1
NMR data (Table 1) revealed that 1 belongs to the
isocoumarin class. Analysis of the H and 13C NMR data
1
(Table 1) indicated that 1 has a tetrasubstituted benzene ring.
In addition, the 1H NMR spectrum (Table 1) displayed signals
for one set of nonequivalent oxymethylene protons (δH 4.72,
dd, J = 10.8, 1.5 Hz, H-3a; 4.43, dd, J = 10.8, 3.0 Hz, H-3b), one
oxymethine (δH 3.70, dq, J = 7.8, 6.6 Hz, H-9), one methine
(δH 2.62, ddd, J = 7.8, 3.0, 1.5 Hz, H-4), and one methyl group
(δH 1.04, d, J = 6.6 Hz, H3-10). The 13C NMR spectrum (Table
1) showed resonances for one lactone carbonyl (δC 169.1), one
oxymethylene (δC 68.3), one oxymethine (δC 65.8), one
methine (δC 44.6), and one methyl (δC 21.4). The HMBC
correlations from the methyl protons (H3-10) to C-9 and C-4,
from the oxymethine (H-9) to C-3, C-4, and C-4a, and other
HMBC correlations (Figure 1) established a hydroisocoumarin
derivative with a 1-hydroxyethyl unit at C-4. The presence of a
Received: October 28, 2012
© XXXX American Chemical Society and
American Society of Pharmacognosy
A
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Chart 1
1
Table 1. H NMR Data (600 MHz, δ in ppm, J in Hz) and 13C NMR Data (150 MHz, δ in ppm) for 1−4
a
b
b
b
1
2
3
4
position
δC, type
δH (J in Hz)
δC, type
162.9, C
δH (J in Hz)
δC, type
162.9, C
δH (J in Hz)
δC, type
δH (J in Hz)
1
3
169.1, C
170.1, C
79.8, CH
68.3, CH2 4.72, dd (10.8, 1.5)
4.43, dd (10.8, 3.0)
77.9, CH
4.38, m
78.8, CH
4.41, m
4.58, m
4
44.6, CH
2.62, ddd (7.8, 3.0,
1.5)
34.6, CH2
2.90, dd (15.6,
11.7)
33.7, CH2
2.84, dd (16.2,
11.1)
33.0, CH2 2.95, dd (16.2,
10.2)
2.83, dd (15.6,
2.4)
2.79, dd (16.2,
3.0)
2.91, dd (16.2,
4.2)
4a
143.4, C
108.3, CH
164.4, C
101.3, CH
163.2, C
100.2, C
65.8, CH
142.1, C
132.2, C
139.5, C
5
6.29, d (2.1)
6.21, d (2.1)
119.3, CH
134.5, CH
111.0, CH
161.3, C
6.79, d (7.2)
123.1, CH
6.86, d (8.4)
7.12, d (8.4)
118.1, CH
136.3, CH
116.3, CH
162.3, C
6.68, d (7.2)
6
7.44, dd (8.4, 7.2) 120.6, CH
7.40, dd (8.4, 7.2)
6.88, d (8.4)
7
6.90, d (8.4)
149.0, C
148.6, C
117.5, C
8
8a
114.0, C
108.6, C
9
3.70, dq (7.8, 6.6)
10
21.4, CH3 1.04, d (6.6)
1′
2′
3′
4′
5′
8-OH
34.8, CH2
21.4, CH2
39.0, CH2
68.0, CH
23.7, CH3
1.86, 1.69, m
1.58, m
34.8, CH2
21.4, CH2
39.0, CH2
68.0, CH
23.7, CH3
1.85, 1.69, m
1.57, m
34.9, CH2 1.90, 1.74, m
21.4, CH2 1.61, m
38.8, CH2 1.51, m
1.49, m
1.49, m
3.81, m
3.82, m
68.0, CH
3.83, m
1.19, d (6.0)
1.19, d (6.6)
23.8, CH3 1.21, d (6.6)
11.00, s
11.08, brs
8-OCH3
56.3, OCH3 3.93, s
62.4, OCH3 3.95, s
a
b
DMSO-d6. CDCl3.
Figure 1. Key HMBC correlations for compounds 1, 2, 6 and 11.
B
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1-hydroxyethyl unit was reinforced by acetylation of 1 with
Ac2O−pyridine. The chemical shift for the triacetyl derivative
1a assigned to the oxymethine proton (δH 5.03) was shifted
significantly downfield (Δδ 1.33 ppm) relative to the original
oxymethine proton (δH 3.70) in 1. Furthermore, in the selective
1D NOE experiment of 1, the irradiation of H3-10 resulted in
the signal intensity of H-5, which further confirmed the
complete planar structure assignment for 1.
The relative and absolute configurations of 1 were
determined by quantum chemical calculation of its ECD
spectrum combined with the modified Mosher’s method. First,
conformational analyses of 4S,9R-1 and 4R,9R-1 were carried
out via Monte Carlo searching with the MMFF94 force field in
the SPARTAN 08 software package.13 The results showed eight
lowest energy conformers for 4S,9R-1 (4S9RC1−4S9RC8; see
Supporting Information Figure S93-1) and four for 4R,9R-1
(4R9RC1−4R9RC4; see Supporting Information Figure S93-
2), whose relative energies were within 2 kcal/mol.
Subsequently, the conformers were reoptimized using DFT at
the B3LYP/6-31G(d) level in the gas phase by the GAUSSIAN
09 program.14 The B3LYP/6-31G(d) harmonic vibrational
frequencies were further calculated to confirm their stability.
The energies, oscillator strengths, and rotational strengths of
the first 20 electronic excitations were calculated using TDDFT
methodology at the B3LYP/6-311++G(2d,2p) level in a
vacuum. The ECD spectra were simulated by the overlapping
Gaussian function (σ = 0.2 or 0.3 eV)15 in which the first nine
excitations for 4S,9R-1 and three excitations for 4R,9R-1 were
adopted, respectively. To get the final spectra for each
compound, the simulated spectra of the six lowest energy
conformations were averaged according to the Boltzmann
distribution theory and their relative Gibbs free energy (ΔG).
The theoretical ECD spectra for 4R,9S-1 and 4S,9S-1 were
obtained by directly reversing the spectra of 4S,9R-1 and 4R,9R-
1, respectively (Figures 2 and 3). The difference observed in
Figure 3. B3LYP/6-311++G(2d,2p)//B3LYP/6-31G(d)-calculated
ECD spectra of 4R,9R-1 (red) and 4S,9S-1 (blue) (σ = 0.2 eV).
4R,9S-1. The absolute configuration at C-9 was further
confirmed by the modified Mosher’s method.16 Compound 1
was treated with (R)- and (S)-MTPA chloride, to afford the
(S)- and (R)-MTPA esters, 1s and 1r, respectively. The
differences in chemical shift values (Δδ = δS − δR) for the
diastereomeric esters 1s and 1r were calculated in order to
assign the absolute configuration at C-9 (Figure 4).
Calculations for all of the relevant signals confirmed the S
absolute configuration at C-9 in 1.
Penicimarin B (2) was obtained as a colorless oil. The
molecular formula was determined as C15H20O4 by HRESIMS
1
with six degrees of unsaturation. In the H NMR spectrum
(Table 1), the proton signals and the coupling constants at δH
7.44 (dd, J = 8.4, 7.2 Hz), 6.90 (d, J = 8.4 Hz), and 6.79 (d, J =
7.2 Hz) indicated the presence of a 1,2,3-trisubstituted benzene
1
system. The H NMR and 13C NMR spectra included four
methylenes, two methines, one methoxy, and one methyl. The
13C NMR spectrum also displayed signals for six aromatic
carbons and a carbonyl carbon. These spectroscopic features
suggested that 2 was very similar to aspergillumarin B (4),11
with the main differences being the additional methoxy signal
(δH 3.93, s in 2) and the absence of a chelated hydroxy proton
signal (δH 11.01, s in 4) in the 1H NMR spectra. Thus, it could
be deduced that the 8-OH in 4 was replaced by a methoxy in 2.
The 8-OMe was further confirmed by an HMBC correlation
from the methoxy to C-8 and the correlation from H-5, one of
the ortho-coupled aromatic protons, to C-4 (Figure 1). On the
basis of these results, the planar structure of 2 was elucidated.
In order to determine the absolute configuration of C-4′ at
the side chain, the modified Mosher’s method was applied.16
When reacted with (R)- and (S)-MTPA chloride, 2 gave the
corresponding (S)- and (R)-MTPA esters, 2s and 2r,
respectively. The observed chemical shift differences ΔδH(S−R)
(Figure 4) clearly defined the S configuration at C-4′. The
absolute configuration of C-3 was determined by CD
spectroscopy. The negative circular dichroism at 259 nm
(Figure 5) suggested the R configuration at C-3, by comparison
with data for dihydroisocoumarins described in the literature.17
Thus, the absolute configuration of 2 was established as 3R, 4′S.
Penicimarin C (3) was also isolated as a colorless oil. Its
molecular formula of C15H20O5 was determined by HRESIMS,
thus revealing the addition of one oxygen atom compared with
Figure 2. B3LYP/6-311++G(2d,2p)//B3LYP/6-31G(d)-calculated
ECD spectra of 4S,9R-1 (red) and 4R,9S-1 (blue) and the
experimental ECD spectrum of 1 (black) (σ = 0.3 eV).
the second Cotton effect around 245 nm for 4R,9R-1 and
4S,9S-1 (Figure 3) may be attributed to overestimation of the
corresponding positive rotatory strengths in the calculations.
For compound 1 the experimental first negative (270 nm),
second small positive (244 nm), third small negative (234 nm),
and fourth positive (220 nm) Cotton effects compared well
with the calculated ECD curve for 4R,9S-1, which showed four
corresponding Cotton effects around 259, 241, 231, and 212
nm (Figure 2). Therefore, qualitative analysis of the result
allowed the assignment of the absolute configuration of 1 as
1
that of 2. The H NMR data (Table 1) resembled those of 2
except for the absence of one aromatic proton signal. The
signals at δH 7.12 (d, J = 8.4 Hz) and 6.86 (d, J = 8.4 Hz)
C
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Figure 4. Δδ (δS − δR) values (in ppm) for the MTPA esters of 1−4.
spectra revealed that a downfield aromatic quaternary carbon
(δC 149.0, C-7) in 3 replaced an aromatic methine carbon (δC
111.0, C-7) in 2. The downfield shift suggested the position of
the additional hydroxy group was at C-7 in 3. Additionally, the
HMBC spectrum showed a correlation from H-5, one of the
two ortho-coupled aromatic protons, to C-4, indicating the
substituted hydroxy and the methoxy should be located at C-7
and C-8. The HMBC correlation from the methoxy to C-8 and
the correlation from H-5 to C-7 confirmed the locations of 8-
OMe and 7-OH. The absolute configuration at C-4′, bearing a
secondary OH group, was determined as S by the modified
Mosher’s method (Figure 4).16 The absolute configuration of
C-3 was also determined to be R by CD (Figure 5).17
Penicimarin D (6) was isolated as a colorless, amorphous
powder, with the molecular formula C19H22O11. Analysis of the
1H and 13C NMR spectroscopic data (Table 2) of 6 revealed
similarities to those of 9, except for the presence of a hexose
residue, with an anomeric carbon at δH 4.78 (d, J = 3.6 Hz)/δC
Figure 5. CD spectra of compounds 2−5.
indicated the presence of a tetrasubstituted benzene system in
3, instead of the trisubstituted benzene in 2. The 13C NMR
1
Table 2. H NMR Data (600 MHz, δ in ppm, J in Hz) and 13C NMR Data (150 MHz, δ in ppm) for 6−9
a
a
a
a
6
7
8
9
position
δC, type
δH (J in Hz)
δC, type
δH (J in Hz)
δC, type
δH (J in Hz)
δC, type
δH (J in Hz)
1
164.8, C
154.3, C
109.6, C
139.5, C
101.4, CH
165.9, C
101.3, CH
162.9, C
98.0, C
164.5, C
155.1, C
108.6, C
138.9, C
100.7, CH
165.9, C
101.5, CH
163.1, C
97.9, C
165.1, C
153.9, C
114.5, C
139.4, C
102.4, CH
165.8, C
101.9, CH
163.0, C
98.7, C
165.8, C
152.5, C
113.2, C
139.7, C
101.5, CH
165.1, C
101.2, CH
163.0, C
98.2, C
3
4
4a
5
6.59, d (1.8)
6.34, d (1.8)
6.44, d (2.4)
6.35, d (2.4)
6.72, d (2.1)
6.39, d (2.1)
6.63, d (2.1)
6.33, d (2.1)
6
7
8
8a
9
16.7, CH3
61.7, CH2
2.33, s
16.8, CH3
58.7, CH2
2.35, s
5.06, s
57.5, CH2
55.2, CH2
4.38, s
4.49, s
16.6, CH3
55.9, CH2
2.30, s
4.41, s
10
4.59, d (12.3)
4.46, d (12.3)
11
12
1′
2′
3′
4′
5′
6′
170.3, C
20.6, CH3
2.02, s
97.8, CH
71.5, CH
72.9, CH
70.3, CH
70.1, CH
63.9, CH2
4.78, d (3.6)
3.24, ddd (9.6, 6.0, 3.6)
3.35, ddd (9.6, 9.6, 4.8)
3.06, ddd (9.6, 9.6, 5.4)
3.57, ddd (9.6, 6.9, 1.8)
4.26, dd (12.0, 1.8)
4.00, dd (12.0, 6.9)
7′
170.4, C
8′
20.6, CH3
2.03, s
6-OH
8-OH
9-OH
10-OH
2′-OH
3′-OH
4′-OH
10.88, brs
11.11, s
10.95, brs
11.06, brs
10.95, brs
11.19, s
10.90, brs
11.16, s
5.48, brs
5.06, brs
4.72, d (6.0)
4.90, d (4.8)
5.17, d (5.4)
a
DMSO-d6.
D
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1
Table 3. H NMR Data (600 MHz, δ in ppm, J in Hz) and 13C NMR Data (150 MHz, δ in ppm) for 11−14
7-methoxy-2,3-
dimethylbenzofuran-5-ol
a
b
a
a
c
11
12
13
14
δH (J in
δH (J in
δH (J in
δH (J in
δH (J in
position
δC, type
151.8, C
Hz)
δC, type
156.1, C
Hz)
δC, type
Hz)
δC, type
Hz)
δC, type
151.5, C
Hz)
2
167.8, C
117.5, C
126.1, C
96.4, CH
154.0, CH
122.0, C
125.8, C
97.0, CH
8.81, s
3
115.3, C
130.3, C
96.12, CH
111.9, C
131.5, C
96.8, CH
110.0, C
132.1, C
97.0, CH
3a
4
6.48, d
(3.3)
6.48, d
(2.1)
6.81, d
(2.1)
6.90, d
(2.1)
6.39, d
(1.8)
5
6
153.7, C
155.2, C
97.6, CH
155.5, C
155.4, C
137.8, C
95.7, CH
96.05, CH
6.29, d
(3.3)
6.34, d
(2.1)
100.5, CH
6.28, d
(2.1)
100.9, CH
6.32, d
(2.1)
6.30, d
(1.8)
7
144.4, C
136.4, C
11.9, CH3
53.5, CH2
146.4, C
139.0, C
12.0, CH3
57.7, CH2
142.3, C
136.6, C
186.3, CH
142.8, C
138.7, C
193.2, C
144.8, C
151.8, C
11.8, CH3
8.0, CH3
7a
8
2.35, s
2.43, s
5.14, s
10.11, s
2.33, s
2.04, s
9
4.44, d
(7.8)
12.7, CH3 2.72, s
27.9, CH3 2.47, s
10
172.8, C
11
20.8, CH3
2.03, s
5-OH
7-OH
7-OCH3
9-OH
9.04, s
9.18, s
9.18, brs
4.92, brs
10.11, s
10.15, brs
55.5, OCH3 3.83, s
56.5, OCH3 3.91, s
56.0, OCH3 3.92, s
4.84, t
(7.8)
a
b
c
1
DMSO-d6. CD3OD. CDCl3. H NMR data (500 MHz, δ in ppm, J in Hz) and 13C NMR data (125 MHz, δ in ppm) for 7-methoxy-2,3-
dimethylbenzofuran-5-ol from ref 19.
97.8. Assignment of the relative configuration of the sugar
moiety in 6 was accomplished by analyses of coupling
constants. The J values of 3.6, 9.6, 9.6, and 9.6 Hz for
Penicifuran A (11) was isolated as a pale brown, semicrystal-
line solid with the molecular formula C11H12O4 by HREIMS,
1
indicating six degrees of unsaturation. The H and 13C NMR
J
H‑1′,H‑2′, JH‑2′,H‑3′, JH‑3′,H‑4′, and JH‑4′,H‑5′ indicated equatorial,
spectra (Table 3) were similar to those of 7-methoxy-2,3-
dimethylbenzofuran-5-ol obtained from the culture of the
fungus Malbranchea cinnamomea HKI0286.19 The only differ-
ence between these two compounds was the replacement of the
methyl signal located at C-3 in 7-methoxy-2,3-dimethylbenzo-
furan-5-ol with a hydroxymethyl signal (δH 4.44, d, J = 7.8 Hz,
2H and 4.84, t, J = 7.8 Hz, 1H) in 11. The HMBC spectrum
(Figure 1) displayed the correlations from the hydroxy group
(δH 4.84) to C-3 and C-9 and from H-9 to C-2, C-3, and C-3a,
confirming the location of the hydroxymethyl at C-3. In
addition, the other hydroxy group (δH 9.04) showed
correlations with C-4, C-5, and C-6, confirming its location at
C-5. This suggested that the assignments for C-5 (δC 137.8)
and C-7a (δC 151.8) of 7-methoxy-2,3-dimethylbenzofuran-5-ol
in the literature19 should be exchanged.
axial, axial, and axial orientations of H-1′, H-2′, H-3′, and H-4′,
respectively. Comparison of the NMR data of the sugar unit in
6 with those of the isocoumarin glucoside halorosellin A18
confirmed the sugar unit is an α-glucopyranose in 6. The
additional signals of an acetyl group (δH 2.03, s; δC 170.4) in
the 1H and 13C NMR spectra, together with the HMBC
correlation (Figure 1) from H-6′ to the carbonyl carbon,
suggested that the hydroxy group at C-6′ was acetylated. The
HMBC correlation from H-10 to C-1′ established the linkage
between the α-glucopyranose and the isocoumarin unit.
Penicimarin E (7) was isolated as a colorless, amorphous
powder with a molecular formula of C13H12O6. Careful
comparison of the H and 13C NMR spectra of 7 (Table 2)
1
with those of 9 showed a close structural relationship. The
obvious differences in the H NMR spectra were the presence
1
Penicifuran B (12), with the molecular formula C13H14O5
(seven degrees of unsaturation) from HREIMS data, was also
of a singlet methyl signal at δH 2.02 in 7 and the significantly
downfield shift of H-10 (δH 5.06 in 7 vs 4.41 in 9 in DMSO-d6).
The presence of a carbonyl group (δC 170.3) in the 13C NMR
spectrum was also observed. These spectroscopic features
indicated that the hydroxy group at C-10 was acetylated in 7.
The location of the acetoxy group at C-10 was confirmed by an
HMBC correlation from H-10 to the carbonyl carbon.
Therefore, 7 was a 10-acetylated derivative of 9.
Penicimarin F (8) was isolated as a colorless, amorphous
powder with the molecular formula C11H10O6. The spectro-
scopic data (Table 2) of 8 also closely corresponded with those
of 9 except for the replacement of the methyl signal (δH 2.30, s)
located at C-3 in 9 with a hydroxymethyl signal (δH 4.38, s, 2H
and 5.48, brs, 1H) in 8. The location of the hydroxymethyl at
C-3 was confirmed by HMBC correlations from H-9 to C-3
and C-4.
obtained as a pale brown, semicrystalline solid. The 1H and 13
C
NMR spectroscopic data (Table 3) were very similar to those
of 11. The differences in the 1H and 13C NMR spectra were the
presence of an acetyl group (δH 2.03, s; δC 20.8/δC 172.8) and
the downfield shift of H-9. These spectroscopic features
indicated that the hydroxy group at C-9 was acetylated in 12.
The location of the acetoxy group at C-9 was confirmed by the
HMBC correlation from H-9 to the carbonyl carbon. Thus, 12
was a 9-acetylated derivative of 11.
Penicifuran C (13) was isolated as a pale brown, amorphous
powder with the molecular formula assigned as C10H8O4 by
HREIMS. Comparison of the 1H and 13C NMR data (Table 3)
of 13 with those of 11 revealed the replacement of a
hydroxymethyl (δH 4.44, d, J = 7.8 Hz, 2H and 4.84, t, J =
7.8 Hz, 1H; δC 53.5) and a methoxy (δH 3.83, s, 3H; δC 55.5) in
E
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Journal of Natural Products
Article
11 with an aldehyde group (δH 10.11, s, 1H; δC 186.3) and a
hydroxy (δH 10.11, s, 1H) in 13, respectively. The HMBC
spectrum displayed correlations from the aldehyde proton to C-
3 and C-3a, confirming the position of the aldehyde group at C-
3. HMBC correlations from the δH 9.18 hydroxy to C-4, C-5,
and C-6 and those from the other hydroxy (δH 10.11) to C-6,
C-7, and C-7a confirmed the positions of the two hydroxy
groups at C-5 and C-7, respectively.
cereus and V. parahemolyticus, with MIC values of 6.25 μM for
each. Compound 11 exhibited moderate inhibitory activity
against S. albus, with an MIC value of 3.13 μM, and weak
activity against B. cereus. All of the isolated compounds were
also evaluated for their cytotoxic activity against human
promyelocytic leukemia HL-60, human lung carcinoma A-
549, HeLa cell human cervical carcinoma, and chronic leukemia
K562 cell lines, but none of them exhibited cytotoxic activity.
In this paper, 10 new metabolites including three hydro-
isocoumarins, penicimarins A−C (1−3), three isocoumarins,
penicimarins D−F (6−8), and four benzofurans, penicifurans
A−D (11−14), were obtained from the sponge-derived fungus
Penicillium sp. MWZ14-4, collected from the South China Sea.
The absolute configurations of 1−4 were assigned by the
modified Mosher’s method and TDDFT ECD calculations
together with comparison of their CD spectra. Among the
naturally occurring isocoumarin derivatives, a 4-substituted
isocoumarin with no substituent at the 3-position such as 1 is
very rare.21−27 During the past decade, only three compounds
with such structural features have been reported.26,27
Interestingly, the co-occurrence of isocoumarins and benzofur-
ans in the same fungal species has also rarely been reported,
only from lichen mycobionts and a Canadian thistle endophytic
fungus.28,29 In this paper, the isocoumarin derivatives with six-
membered lactones and the benzofurans were simultaneously
isolated from a marine-derived Penicillium fungus for the first
time. All of the isolated isocoumarin derivatives and
benzofurans may share a similar biosynthetic pathway from a
common open-chain precursor.
Penicifuran D (14) was obtained as a pale brown,
semicrystalline solid. The molecular formula was determined
as C10H8O4 on the basis of HREIMS data. Comparison of the
1H NMR spectrum of 14 (Table 3) with that of 13 showed
similar structures between them. The significant differences in
1
the H NMR spectrum of 14 were the presence of an olefinic
proton signal for H-2, downfield shifted to δH 8.81, and the
absence of an exchangeable proton (δH 10.11 in 13). The
primary differences in the 13C NMR spectra were that the
methyl carbon signal downfield shifted from δC 12.7 (C-9) in
13 to δC 27.9 (C-9) in 14 and that the ketone carbon (δC
193.2) and olefinic methine carbon (δC 154.0) in 14 replaced
the aldehyde carbon and olefinic quaternary carbon in 13.
These results revealed the aldehyde group located at C-3 in 13
was replaced by an acetyl group in 14 and that the substituent
methyl at C-2 was not present. In addition, the HMBC
correlations from H3-9 to C-3 and C-8 implied that the acetyl
group was located at C-3.
The known compounds aspergillumarin B (4),11 aspergillu-
marin A (5),11 sescandelin-B (9),12 and 5,6,8-trihydroxy-4-(1′-
hydroxyethyl)isocoumarin (10)12 were determined by compar-
ison of their spectroscopic data with those in the literature. The
R configuration at C-3 in 4 was determined by comparison of
the CD spectrum (Figure 5) with those of 2, 3, and reported
compounds in the literature.17 The absolute configuration of C-
4′ in 4 was determined as S by the modified Mosher’s method
(Figure 4).16 Comparison of the CD spectrum (Figure 5) as
EXPERIMENTAL SECTION
■
General Experimental Procedures. Melting points were
determined on an X-6 micromelting point apparatus and are
uncorrected. Optical rotations were measured on a Jasco P-1020
digital polarimeter. UV spectra were recorded on a Beckman DU 640
spectrophotometer. CD spectra were recorded on a Jasco J-810
circular dichroism spectrometer. IR spectra were recorded on a
Nicolet-Nexus-470 spectrometer using KBr pellets. 1H NMR, 13C
NMR, and DEPT spectra of compounds 1−14 and 2D NMR spectra
of compounds 1−3, 8, and 12−14 were recorded on a JEOL JNM-
well as the specific rotation of 5 ([α]25 −92.1) with those
D
reported for similar compounds17,20 suggested that C-3 in 5 has
the R configuration. Additionally, the Dess−Martin oxidation of
4 led to 5, further confirming the R configuration at C-3 in 4.
The antibacterial activities of all isolated compounds were
evaluated with seven terrestrial pathogenic bacteria, Escherichia
coli, Staphylococcus aureus, S. albus, Bacillus subtilis, B. cereus,
Micrococcus tetragenus, and Kocuria rhizophila, and two marine
pathogenic bacteria, Vibrio parahemolyticus and V. anguillarum
(Table 4). Compound 10 showed moderate activity against B.
ECP NMR spectrometer (600 MHz for H and 150 MHz for 13C),
1
using TMS as an internal standard. 2D NMR spectra of compounds 6,
7, and 11 were recorded on an Ultrashied 400 MHz Plus spectrometer.
The 1H NMR spectra of compounds 1a, 1s, and 1r were recorded on
an Agilent DD2 500 MHz NMR spectrometer. ESIMS and HRESIMS
spectra were obtained from a Micromass Q-TOF spectrometer and
Thermo Scientific LTQ Orbitrap XL spectrometer. EIMS spectra were
measured on a Thermo DSQ EI-mass spectrometer, and HREIMS
spectra on a Thermo MAT95XP high-resolution mass spectrometer.
HPLC separations were performed using a Hitachi L-2000 prep-
HPLC system coupled with a Hitachi L-2455 photodiode array
detector. A Kromasil C18 preparative HPLC column (250 × 10 mm, 5
μm) was used. Silica gel (Qing Dao Hai Yang Chemical Group Co.;
200−300 mesh), Sephadex LH-20 (Amersham Biosciences), and
octadecylsilyl silica gel (Unicorn; 45−60 μm) were used for column
chromatography. Precoated silica gel plates (Yan Tai Zi Fu Chemical
Group Co.; G60, F-254) were used for thin-layer chromatography.
Fungal Material. The fungal strain Penicillium sp. (MWZ14-4) was
isolated from a piece of fresh tissue from the inner part of an
unidentified sponge (GX-WZ-20080014), collected from the Weizhou
coral reef in the South China Sea in September 2008. The fungus was
identified as a Penicillium sp. according to morphological traits and a
molecular biological protocol by amplification and sequencing of the
DNA sequences of the ITS region of the rRNA gene. The 532 base
pair ITS sequence had 100% sequence identity to that of Penicillium
sp. 3TMS-2011 (HQ631007). The sequence data have been submitted
Table 4. Antibacterial Activities of Compounds 4, 5, 8, and
a
10−14
MIC (μM)
compound
S. aureus
S. albus
B. cereus
V. parahemolyticus
4
5
8
>25.0
>25.0
12.5
12.5
12.5
>25.0
>25.0
>25.0
6.25
>25.0
>25.0
>25.0
6.25
>25.0
>25.0
3.13
10
12.5
11
25.0
12.5
25.0
12
>25.0
12.5
12.5
>25.0
12.5
>25.0
>25.0
25.0
13
25.0
14
>25.0
0.160
25.0
25.0
b
ciprofloxacin
0.312
0.625
0.160
a
b
Data are expressed in MIC values (μM). Ciprofloxacin was used as a
positive control.
F
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Journal of Natural Products
Article
to GenBank, accession number JN693500. The strain was deposited in
the Key Laboratory of Marine Drugs, the Ministry of Education of
China, School of Medicine and Pharmacy, Ocean University of China,
Qingdao, PR China.
1621, 1484, 1378, 1243, 1184, 1004, 853, 722 cm−1; 1H and 13C NMR
see Table 2; EIMS m/z 238 [M]+; HREIMS m/z 238.0476 [M]+
(calcd for C11H10O6, 238.0472).
Penicifuran A (11): pale brown, semicrystalline solid; mp 118−119
°C; UV (MeOH) λmax (log ε) 218 (3.74), 250 (3.63), 290 (3.31) nm;
IR (KBr) νmax 3398, 3182, 1632, 1601, 1438, 1386, 1276, 1216, 1152,
Fermentation, Extraction, and Isolation. Thirty-five Erlen-
meyer flasks of the fungal strain were cultivated in solid medium
(Erlenmeyer flasks each containing rice 80 g, water 120 mL, sea salt
2.0 g) at 27 °C for four weeks. The fermented solid medium was
extracted three times with EtOAc. The combined EtOAc layers were
evaporated to dryness under reduced pressure to give an EtOAc
extract. The EtOAc extract (9.0 g) was subjected to vacuum liquid
chromatography (VLC) on silica gel using step gradient elution with
EtOAc−petroleum ether (0−100%) and then with MeOH−EtOAc
(0−100%) to afford eight fractions (Fr. 1−Fr. 8). Fr. 3 was isolated by
column chromatography (CC) on silica gel eluted with petroleum
ether−EtOAc (v/v, 7:3), then subjected to Sephadex LH-20 CC with
petroleum ether−CHCl3−MeOH (v/v/v, 2:1:1), and further purified
by using semipreparative HPLC on an ODS column (Kromasil C18,
250 × 10 mm, 5 μm, 2 mL/min) eluted with 70% MeOH−H2O for 5
(10.0 mg) and 60% MeOH−H2O for 7 (20.0 mg) and 12 (2.3 mg).
Fr. 5 was first subjected to repeated silica gel CC (CHCl3−MeOH, v/
v, 15:1), then separated by Sephadex LH-20 CC (CHCl3−MeOH, v/v,
1:1), and further purified on HPLC with 70% MeOH−H2O for 4
(12.0 mg) and 20% MeOH−H2O for 13 (3.0 mg). Fr. 6 was applied to
CC on silica gel (petroleum ether−EtOAc, v/v, 3:2), then separated
by Sephadex LH-20 CC (CHCl3−MeOH, v/v, 1:1), and further
purified on HPLC with 30% MeOH−H2O to obtain 1 (4.0 mg) and
14 (2.0 mg), 65% MeOH−H2O for 3 (8.0 mg), 40% MeOH−H2O for
10 (45.0 mg), and 45% MeOH−H2O for 11 (3.0 mg). Fr. 7 was
fractionated on silica gel CC (CHCl3−MeOH, v/v, 10:1), then
purified by Sephadex LH-20 CC (MeOH), and further purified on
HPLC with 40% MeOH−H2O to afford 2 (6.0 mg) and 8 (8.0 mg)
and 60% MeOH−H2O for 6 (5.0 mg) and 9 (11.0 mg).
1
1000, 970, 812 cm−1; H and 13C NMR see Table 3; EIMS m/z 208
[M]+; HREIMS m/z 208.0726 [M]+ (calcd for C11H12O4, 208.0730).
Penicifuran B (12): pale brown, semicrystalline solid; mp 103−104
°C; UV (MeOH) λmax (log ε) 223 (3.94), 250 (3.88), 294 (3.47) nm;
IR (KBr) νmax 3321, 1710, 1603, 1436, 1384, 1237, 1151, 1022, 972,
836 cm−1; 1H and 13C NMR see Table 3; EIMS m/z 250 [M]+;
HREIMS m/z 250.0835 [M]+ (calcd for C13H14O5, 250.0836).
Penicifuran C (13): pale brown, amorphous powder; UV (MeOH)
λmax (log ε) 218 (3.41), 240 (3.28), 272 (2.97) nm; IR (KBr) νmax
1
3357, 1668, 1504, 1384, 1314, 1180, 1141, 835 cm−1; H and 13C
NMR see Table 3; EIMS m/z 192 [M]+; HREIMS m/z 192.0415
[M]+ (calcd for C10H8O4, 192.0417).
Penicifuran D (14): pale brown, semicrystalline solid; mp 190−192
°C; UV (MeOH) λmax (log ε) 217 (3.23), 236 (3.12), 264 (3.31), 305
(2.89) nm; IR (KBr) νmax 3299, 1650, 1506, 1358, 1279, 1153, 841
1
cm−1; H and 13C NMR see Table 3; EIMS m/z 192 [M]+; HREIMS
m/z 192.0414 [M]+ (calcd for C10H8O4, 192.0417).
Preparation of the (R)- and (S)-MTPA Esters of 1−4.
Compound 1 (1.0 mg) was dissolved in 500 μL of pyridine, and
dimethylaminopyridine (2.0 mg) and (R)-MTPACl (8 μL) were then
added in sequence. The reaction mixture was stirred for 14 h at room
temperature (rt), and 1 mL of H2O was then added. The solution was
extracted with 5 mL of CH2Cl2, and the organic phase was
concentrated under reduced pressure. Then the residue was purified
by semipreparative HPLC (90% MeOH−H2O) to yield the (S)-
MTPA ester 1s (0.9 mg). By the same procedure, the (R)-MTPA ester
1r (0.8 mg) was obtained from the reaction of 1 (1 mg) with (S)-
MTPACl (8 μL).
Penicimarin A (1): colorless, amorphous powder; [α]25 −36 (c
D
0.15, MeOH); UV (MeOH) λmax (log ε) 219 (3.91), 233 (3.99), 270
(3.93), 303 (3.74) nm; CD (0.92 mM, MeOH) λmax (Δε) 220
(+7.52), 234 (−3.65), 244 (+1.24), 270 (−11.61) nm; IR (KBr) νmax
3204, 1650, 1402, 1243, 1166, 1118, 850, 713 cm−1; 1H and 13C NMR
see Table 1; EIMS m/z 224 [M]+; HREIMS m/z 224.0680 [M]+
(calcd for C11H12O5, 224.0679).
By the same procedure as for the preparation of the (S)- and (R)-
MTPA esters of 1, (S)-MTPA esters 2s, 3s, 4s and (R)-MTPA esters
2r, 3r, 4r were obtained.
1
(S)-MTPA ester (1s): H NMR (CD3OD, 500 MHz) δ 7.71−6.88
(17H, m, aromatic protons), 5.51 (1H, m, H-9), 4.63 (1H, dd, J =
12.0, 3.5 Hz, H-3a), 4.59 (1H, dd, J = 12.0, 2.5 Hz, H-3b), 3.74 (3H, s,
OCH3-MTPA), 3.68 (3H, s, OCH3-MTPA), 3.44 (1H, m, H-4), 3.37
(3H, s, OCH3-MTPA), 1.35 (3H, d, J = 6.5 Hz, H-10); ESIMS m/z
Penicimarin B (2): colorless oil; [α]25 −69 (c 0.5, CHCl3); UV
D
(MeOH) λmax (log ε) 221 (3.32), 244 (2.35), 306 (3.27) nm; CD
(0.63 mM, MeOH) λmax (Δε) 239 (+2.01), 259 (−4.15), 290 (−1.42),
302 (−1.96) nm; IR (KBr) νmax 3521, 1710, 1598, 1477, 1279, 1250,
1
895.2 [M + Na]+, 911.2 [M + K]+. (R)-MTPA ester (1r): H NMR
(CD3OD, 500 MHz) δ 7.73−6.84 (17H, m, aromatic protons), 5.44
(1H, m, H-9), 4.51 (1H, dd, J = 12.0, 3.5 Hz, H-3a), 4.41 (1H, dd, J =
12.0, 2.5 Hz, H-3b), 3.71 (3H, s, OCH3-MTPA), 3.70 (3H, s, OCH3-
MTPA), 3.55 (3H, s, OCH3-MTPA), 3.35 (1H, m, H-4), 1.48 (3H, d,
J = 6.0 Hz, H-10); ESIMS m/z 895.2 [M + Na]+, 911.2 [M + K]+.
(S)-MTPA ester (2s): 1H NMR (CDCl3, 600 MHz) δ 6.92 (1H, d, J
= 8.4 Hz, H-7), 6.78 (1H, d, J = 7.8 Hz, H-5), 5.15 (1H, m, H-4′), 4.34
(1H, m, H-3), 3.95 (3H, s, 8-OCH3), 3.54 (3H, s, OCH3-MTPA), 2.89
(1H, dd, J = 16.2, 12.0 Hz, H-4a), 2.78 (1H, dd, J = 16.2, 2.4 Hz, H-
4b), 1.84 (1H, m, H-1′a) and 1.75 (1H, m, H-1′b), 1.63 (2H, m, H-
2′), 1.52 (2H, m, H-3′), 1.28 (3H, d, J = 6.0 Hz, H-5′); ESIMS m/z
481.0 [M + H]+, 502.9 [M + Na]+, 519.0 [M + K]+, 982.9 [2 M +
1
1232, 1111, 1088, 1063, 803 cm−1; H and 13C NMR see Table 1;
ESIMS m/z 265.3 [M + H]+, 287.3 [M + Na]+, 303.3 [M + K]+, 551.6
[2 M + Na]+; HRESIMS m/z 265.1433 [M + H]+ (calcd for
C15H21O4, 265.1434).
Penicimarin C (3): colorless oil; [α]25 −84 (c 0.1, CHCl3); UV
D
(MeOH) λmax (log ε) 220 (3.39), 250 (3.48), 319 (3.38) nm; CD
(0.71 mM, MeOH) λmax (Δε) 235 (−0.85), 256 (−2.88), 283
(−0.73), 317 (−1.98) nm; IR (KBr) νmax 3354, 1708, 1489, 1430,
1373, 1295, 1128, 1053, 894, 802 cm−1; 1H and 13C NMR see Table 1;
ESIMS m/z 281.3 [M + H]+, 303.3 [M + Na]+, 319.3 [M + K]+, 583.6
[2 M + Na]+; HRESIMS m/z 281.1382 [M + H]+ (calcd for
C15H21O5, 281.1384).
1
Na]+. (R)-MTPA ester (2r): H NMR (CDCl3, 600 MHz) δ 6.92
Penicimarin D (6): colorless, amorphous powder; [α]25D +69 (c 0.2,
MeOH); UV (MeOH) λmax (log ε) 248 (4.07), 279 (3.60), 327 (3.57)
nm; IR (KBr) νmax 3367, 1732, 1680, 1626, 1370, 1252, 1176, 1037,
846, 713 cm−1; 1H and 13C NMR see Table 2; ESIMS m/z 427.2 [M +
H]+, 449.2 [M + Na]+, 465.2 [M + K]+, 853.5 [2 M + H]+, 875.5 [2 M
+ Na]+; HRESIMS m/z 427.1229 [M + H]+ (calcd for C19H23O11,
427.1235).
Penicimarin E (7): colorless, amorphous powder; UV (MeOH) λmax
(log ε) 246 (4.37), 325 (3.83) nm; IR (KBr) νmax 3197, 1681, 1618,
1372, 1266, 1166, 1032, 847, 740 cm−1; 1H and 13C NMR see Table 2;
EIMS m/z 264 [M]+; HREIMS m/z 264.0625 [M]+ (calcd for
C13H12O6, 264.0628).
(1H, d, J = 9.0 Hz, H-7), 6.78 (1H, d, J = 7.2 Hz, H-7), 5.15 (1H, m,
H-4′), 4.25 (1H, m, H-3), 3.95 (3H, s, 8-OCH3), 3.57 (3H, s, OCH3-
MTPA), 2.87 (1H, dd, J = 15.6, 11.4 Hz, H-4a), 2.81 (1H, dd, J = 15.6,
3.9 Hz, H-4b), 1.79 (1H, m, H-1′a) and 1.69 (1H, m, H-1′b), 1.57
(2H, m, H-2′), 1.47 (2H, m, H-3′), 1.35 (3H, d, J = 6.0 Hz, H-5′);
ESIMS m/z 480.9 [M + H]+, 502.9 [M + Na]+, 518.9 [M + K]+, 982.9
[2 M + Na]+.
(S)-MTPA ester (3s): 1H NMR (CDCl3, 600 MHz) δ 7.20 (1H, d, J
= 7.8 Hz, H-6), 6.99 (1H, d, J = 7.8 Hz, H-5), 5.16 (1H, m, H-4′), 4.43
(1H, m, H-3), 3.81 (3H, s, 8-OCH3), 3.73 (3H, s, OCH3-MTPA), 3.54
(3H, s, OCH3-MTPA), 2.92 (1H, dd, J = 16.2, 12.0 Hz, H-4a), 2.83
(1H, dd, J = 16.2, 3.0 Hz, H-4b), 1.87 (1H, m, H-1′a) and 1.76 (1H,
m, H-1′b), 1.69 (2H, m, H-2′), 1.52 (2H, m, H-3′), 1.29 (3H, d, J =
6.0 Hz, H-5′); ESIMS m/z 712.9 [M + H]+, 734.8 [M + Na]+, 750.9
Penicimarin F (8): colorless, amorphous powder; UV (MeOH) λmax
(log ε) 249 (3.73), 329 (3.17) nm; IR (KBr) νmax 3444, 3258, 1681,
G
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Journal of Natural Products
Article
[M + K]+. (R)-MTPA ester (3r): 1H NMR (CDCl3, 600 MHz) δ 7.24
(1H, d, J = 7.8 Hz, H-6), 7.00 (1H, d, J = 7.8 Hz, H-5), 5.16 (1H, m,
H-4′), 4.31 (1H, m, H-3), 3.75 (3H, s, 8-OCH3), 3.72 (3H, s, OCH3-
MTPA), 3.57 (3H, s, OCH3-MTPA), 2.84 (1H, dd, J = 16.2, 11.1 Hz,
H-4a), 2.77 (1H, dd, J = 16.2, 3.0 Hz, H-4b), 1.80 (1H, m, H-1′a) and
1.69 (1H, m, H-1′b), 1.61 (2H, m, H-2′), 1.48 (2H, m, H-3′), 1.36
(3H, d, J = 6.0 Hz, H-5′); ESIMS m/z 712.9 [M + H]+, 734.9 [M +
Na]+.
ACKNOWLEDGMENTS
■
This work was supported by the Key Program of National
Natural Science Foundation of China (No. 41130858), the
National Natural Science Foundation of China (No.
40776073), the Program for New Century Excellent Talents
in University, Ministry of Education of China (No. NCET-11-
0472), the Research Fund for the Doctoral Program of Higher
Education, Ministry of Education of China (No.
20090132110002), and the Open Research Fund Program of
State Key Laboratory of Microbial Metabolism (Shanghai Jiao
Tong University) (MMLKF12-07).
(S)-MTPA ester (4s): 1H NMR (CDCl3, 600 MHz) δ 7.19 (1H, d, J
= 7.8 Hz, H-7), 6.97 (1H, d, J = 7.8 Hz, H-5), 5.16 (1H, m, H-4′), 4.48
(1H, m, H-3), 3.75 (3H, s, OCH3-MTPA), 3.53 (3H, s, OCH3-
MTPA), 2.96 (1H, dd, J = 16.2, 12.0 Hz, H-4a), 2.88 (1H, dd, J = 16.2,
3.0 Hz, H-4b), 1.85 (1H, m, H-1′a) and 1.75 (1H, m, H-1′b), 1.66
(2H, m, H-2′), 1.49 (2H, m, H-3′), 1.28 (3H, d, J = 6.6 Hz, H-5′);
ESIMS m/z 704.8 [M + Na]+. (R)-MTPA ester (4r): 1H NMR
(CDCl3, 600 MHz) δ 7.17 (1H, d, J = 8.1 Hz, H-7), 6.98 (1H, d, J =
8.1 Hz, H-5), 5.15 (1H, m, H-4′), 4.36 (1H, m, H-3), 3.77 (3H, s,
OCH3-MTPA), 3.57 (3H, s, OCH3-MTPA), 2.90 (1H, dd, J = 16.2,
12.0 Hz, H-4a), 2.82 (1H, dd, J = 16.2, 3.0 Hz, H-4b), 1.77 (2H, m, H-
1′), 1.58 (2H, m, H-2′), 1.45 (2H, m, H-3′), 1.35 (3H, d, J = 6.0 Hz,
H-5′); ESIMS m/z 704.8 [M + Na]+.
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ASSOCIATED CONTENT
■
S
* Supporting Information
1H, 13C, HMQC, HMBC, and MS spectra of the new
compounds 1−3, 6−8, and 11−14 and COSY spectra of 1−
1
3 and 6; H, MS spectra of the (R)- and (S)-MTPA esters of
1
1−4, H, MS spectra of the triacetyl derivative 1a of 1, and
computational calculation details and the original data for each
conformer of 1. These materials are available free of charge via
the Internet at http://pubs.acs.org.
AUTHOR INFORMATION
■
(17) Choukchou-Braham, N.; Asakawa, Y.; Lepoittevin, J. P.
Tetrahedron Lett. 1994, 35, 3949−3952.
(18) Chinworrungsee, M.; Kittakoop, P.; Isaka, M.; Chanphen, R.;
Tanticharoen, M.; Thebtaranonth, Y. J. Chem. Soc., Perkin Trans. 1
2002, 2473−2476.
Corresponding Author
*Tel/Fax: 86-532-82031536. E-mail: changyun@ouc.edu.cn.
Notes
The authors declare no competing financial interest.
H
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