A Pair of Enantiomeric Bis- seco-abietane Diterpenoids from Cryptomeria fortunei

Article abstract of DOI:10.1021/acs.jnatprod.8b00482

(±)-Cryptomeriolide, a pair of racemic bis-seco-abietane diterpenoids, were isolated from the bark of Cryptomeria fortunei. The separation of enantiomers was achieved by using chiral stationary phase HPLC. Their structures including the absolute configuration and conformations in solution and solid state were determined by extensive analysis of spectroscopic data, single-crystal X-ray diffraction, and comparison of calculated and experimental electronic circular dichroism data. A bioinspired one-pot enantiomeric synthesis of 1a and 1b was accomplished via a readily made intermediate orthoquinone from sugiol. All compounds including the synthetic intermediates were assayed for their cytotoxic activities on human cancer cell lines HL-60, A549, and SGC7901.

Full text of DOI:10.1021/acs.jnatprod.8b00482

Article
Cite This: J. Nat. Prod. XXXX, XXX, XXX−XXX
pubs.acs.org/jnp
A Pair of Enantiomeric Bis-seco-abietane Diterpenoids from
Cryptomeria fortunei
†
‡
†,§
‡
∥
†,§
Lu Feng, Attila Man
́
di, Chunping Tang, Tibor Kurtan
́
, Shuai Tang, Chang-Qiang Ke,
∥
§
,†,§
,†,§,⊥
Ning Shen, Ge Lin, Sheng Yao,* and Yang Ye*
†
∥
State Key Laboratory of Drug Research and Natural Products Chemistry Department and Division of Antitumor Pharmacology,
Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, People’s Republic of China
‡
Department of Organic Chemistry, University of Debrecen, P.O. Box 400, H-4002 Debrecen, Hungary
§
SIMM-CUHK Joint Research Laboratory for Promoting Globalization of Traditional Chinese Medicines, Shanghai 201203,
People’s Republic of China
⊥
School of Life Science and Technology, ShanghaiTech University, Shanghai 201203, People’s Republic of China
S Supporting Information
*
ABSTRACT: (±)-Cryptomeriolide, a pair of racemic bis-seco-abietane diterpenoids, were isolated from the bark of Cryptomeria
fortunei. The separation of enantiomers was achieved by using chiral stationary phase HPLC. Their structures including the
absolute configuration and conformations in solution and solid state were determined by extensive analysis of spectroscopic
data, single-crystal X-ray diffraction, and comparison of calculated and experimental electronic circular dichroism data. A
bioinspired one-pot enantiomeric synthesis of 1a and 1b was accomplished via a readily made intermediate orthoquinone from
sugiol. All compounds including the synthetic intermediates were assayed for their cytotoxic activities on human cancer cell
lines HL-60, A549, and SGC7901.
4
bietanes are a family of naturally occurring diterpenoids
discovered from a variety of terrestrial plant sources and
murine BV-2 microglial cells. To date, more than 80 naturally
3
−15
A
occurring seco-abietane diterpenoids have been reported.
have displayed a wide spectrum of interesting biological
activities, including cytotoxic, antimicrobial, antifungal, anti-
viral, antiulcer, cardiovascular, antioxidant, and anti-inflamma-
Particularly, hyptisolode A is a bis-seco-abietane, and its C-7/C-
8 bond (B-ring) and the C-11/C-12 bond (C-ring) both have
7
been cleaved by oxidation.
1
tory activity. Some drugs possessing an abietane skeleton,
The genus Cryptomeria (Cupressaceae) comprises two
such as Ecabet Sodium, have been on the market for years.
Ecabet Sodium is a 12-sulfodehydroabietic acid monosodium
salt used in the management of gastritis and gastric ulceration
due to its affinity to the gastric mucosa and to fibrinogen
species. Distributed mainly in China and Japan, Cryptomeria
16−20
is known for its abundance in abietane diterpenoids.
Chinese cedar (Cryptomeria fortunei Hooibrenk), also known
as Liu-Shan in Chinese, is an indigenous evergreen tree
growing in the south of the Yangtze River region. As a
traditional folk medicine, its root bark has been widely used as
2
located on the gastric ulcer base.
In recent decades, the abietanes have attracted broad
attention from natural product chemists and pharmacological
communities in their search for new lead compounds. Of the
known abietane diterpenoids, seco-abietanes constitute a small
but interesting group that originates from the ring cleavage of
21
a detoxification and an insecticidal agent. The essential oils of
its wood and leaf were reported to show antibacterial and
22,23
antitermitic activity.
Our research group reported a series of abietane
19
3
diterpenoids from C. fortunei previously. In the course of
our continuing effort to explore unique and bioactive
the core skeleton of the abietanes at unspecific positions.
They also showed remarkable bioactivity. For example,
chlorabietins B and C, two 13,14-seco-abietanes, showed
antineuroinflammatory effects by inhibiting the nitric oxide
Received: June 15, 2018
(
NO) production in lipopolysaccharide (LPS)-activated
©
XXXX American Chemical Society and
American Society of Pharmacognosy
A
DOI: 10.1021/acs.jnatprod.8b00482
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Journal of Natural Products
Article
Table 1. H (500 MHz) and 13C (125 MHz) NMR Data for
1
structures, this plant was investigated in depth. As a result, a
pair of racemic bis-seco-abietane diterpenoids was isolated, and
the two enantiomers (1a and 1b) were obtained by chiral-
phase HPLC separation. Their structures incorporate a rare
architecture presumably derived from a common abietane
diterpenoid skeleton. Their structures were characterized on
the basis of spectroscopic data analysis, a single-crystal X-ray
diffraction experiment, and comparison of experimental and
computed electronic circular dichroism (ECD) data. A concise
enantiomeric synthesis of 1a and 1b was accomplished
following a plausible biosynthetic pathway. Herein, we present
the isolation and structure elucidation of 1a and 1b,
particularly the absolute configuration determination. The
bioinspired one-pot method to construct 1a and 1b via a
readily made intermediate orthoquinone from sugiol is also
described.
Compound 1 in CDCl (δ in ppm, J in Hz)
3
position
δC
δH, mult (J in Hz)
1
2
3
4
25.0, CH2
28.1, CH2
83.7, CH
72.2, C
3.06, m; 2.55, ddd (13.8, 13.7, 7.0)
2.14, m; 1.79, m
3.88, dd (12.2, 4.5)
5
6
7
8
135.3, C
133.3, CH
127.5, CH
134.3, C
130.4, C
135.0, C
171.1, C
170.6, C
142.7, C
130.2, CH
32.1, CH
21.2, CH3
21.7, CH3
24.0, CH3
26.1, CH3
19.5, CH3
7.25, d (8.0)
7.03, d (8.0)
9
10
11
12
1
1
1
1
1
1
1
2
3
4
5
6
7
8
9
0
6.78, s
2.80, sept (6.5)
1.16, d (6.5)
1.17, d (6.5)
1.24, s
1.16, s
2.35, s
RESULTS AND DISCUSSION
■
The acetone extract of the bark of C. fortunei was dissolved in
water and partitioned with petroleum ether (PE), dichloro-
methane (CH Cl ), and ethyl acetate (EtOAc), successively.
2
2
After evaporation to dryness under vacuum of the solvent, the
PE fraction dissolved in 80% aqueous MeOH. The CH Cl
2
2
fraction and the MeOH layer were combined together based
on the TLC profiles and separated by repeated column
chromatography and preparative HPLC to afford compound 1
as a white powder. Its molecular formula of C H O was
1
1
Figure 1. Key H− H COSY (red bold line) and HMBC correlations
(
H → C) of 1.
20
26
5
deduced from the sodium adduct ion at m/z 369.1662 [M +
Na] in the HRESIMS, corresponding to eight indices of
+
aromatic C-9 and C-5, and from H-2 to the aromatic C-10
facilitated the construction of a seven-membered ring. HMBC
correlations from the olefinic H-14 to C-15 and C-12, and
from H-15 to C-12 and C-14 established an isopentyl unit with
a carboxylic group connecting to the olefinic C-13. HMBC
correlations from H-14 to the aromatic carbons C-7 and C-9
confirmed the connection of C-14 to C-8. The key HMBC
hydrogen deficiency. The IR spectrum showed absorption
−
1
−1
bands for hydroxy (3446 cm ) and carbonyl (1713 cm )
groups. A total of 20 carbon resonances were well resolved in
1
3
the C NMR and DEPT spectra, ascribed to five methyls, two
methylenes, five methines, five quaternary carbons, an
oxygenated tertiary carbon, and two carbonyls (Table 1).
The H NMR data (Table 1), combined with the C NMR
data, displayed diagnostic signals for a 1,2,3,4-tetrasubstituted
1
13
correlations from H-2 and H-3 to C-4, and from CH -20 to C-
3
6 suggested that the oxygenated isopropyl and the methyl
group were located at C-3 and C-5, respectively. The NOESY
interaction between H-14 and H-15 indicated the Z
benzene [δ 7.25 (d, J = 8.0 Hz), 7.03 (d, J = 8.0 Hz); δC
H
1
35.3, 135.0, 134.3, 133.3, 130.4, 127.5], a trisubstituted
1
3(14)
double bond [δH 6.78 (s); δC 142.7, 130.2], an aromatic
methyl [δ 2.35 (s, 3H); δ 19.5], an oxygenated isopropyl
configuration of the Δ
double bond. Therefore, the 2D
structure of 1 was established and the compound was named
cryptomeriolide.
H
C
group [δ 1.24 (s, 3H), 1.16 (s, 3H); δ 72.2, 26.1, 24.0], an
H
C
isopropyl [δ 2.80 (sept, J = 6.5 Hz, 1H), 1.17 (d, J = 6.5 Hz,
The structure of cryptomeriolide (1) has one stereogenic
H
1
13
3
H), 1.16 (d, J = 6.5 Hz, 3H); δ 32.1, 21.7, 21.2], and two
center at C-3. Although only one set of H and C NMR data
was observed, the specific rotation value (−3) of 1 suggested
that it could be racemic. Compound 1 was then analyzed on a
chiral-phase HPLC column, and two peaks with an
approximate ratio of 1:1 were observed (Figure S1, Supporting
Information). Single crystals of the racemic cryptomeriolide
(1) were obtained (CCDC 1446792), and the X-ray diffraction
data (Figure 2) showed that 1 consisted of a racemic
C
ester carbonyl groups (δ 171.1, 170.6). These observations
C
accounted for seven out of eight indices of hydrogen
deficiency, and the remaining one implied the presence of an
additional ring. The connectivity of CH (1)−CH (2)−CH(3)
2
2
could be well interpreted from correlations of H-1/H-2 and H-
1
1
2
/H-3 in the H− H COSY spectrum (Figure 1). HMBC
correlations from H-3 to the carbonyl C-11, from H-1 to the
B
DOI: 10.1021/acs.jnatprod.8b00482
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Journal of Natural Products
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Figure 2. Perspective ORTEP drawing for 1.
compound. Chiral-phase HPLC separation of rac-1 eventually
afforded the enantiomers 1a and 1b.
Compounds 1a and 1b gave the opposite specific rotations
(
+159 for 1a vs −153 for 1b) and mirror images of ECD
curves (Figure S2, Supporting Information). To assign the
absolute configuration of C-3, time-dependent density func-
tional theory (TDDFT) ECD calculations were carried out on
24,25
the arbitrarily chosen enantiomer (S).
A Merck Molecular
Force Field (MMFF) conformational search in CHCl resulted
3
in 116 conformers in a 21 kJ/mol energy window. These
conformers were reoptimized at the B3LYP/6-31G(d) level in
2
6,27
vacuo and at the B97D/TZVP
CAM-B3LYP/TZVP
PCM/MeOH and the
28,29
PCM/MeOH levels. ECD spectra
were calculated with various functionals (B3LYP, BH&HLYP,
CAM-B3LYP, and PBE0) and the TZVP basis set for
conformers above 1% Boltzmann population (Figure 3). All
Figure 4. Experimental ECD spectrum of 1a in MeOH compared
with the Boltzmann-weighted BH&HLYP/TZVP PCM/MeOH ECD
spectrum of (S)-1 computed for the CAM-B3LYP/TZVP PCM/
MeOH conformers. Bars represent the rotational strength values of
the lowest-energy conformer.
relative to the conjugating benzene ring seems to play a
decisive role in the ECD spectrum. In this case, the
comparison of the experimental ECD spectrum with the
weighted TDDFT-ECD spectrum computed for the optimized
solid-state X-ray conformers would have given an incorrect
absolute configuration, since different conformers with
opposite ECD are prevalent in solution.
Figure 3. Low-energy conformers (≥1%) of (S)-1 obtained by
optimization of the MMFF conformers at the CAM-B3LYP/TZVP
level with PCM for MeOH. Group A contains conformers A−D, F, G,
I, J, and L−O; group B contains conformers E, H, K, P, and Q.
The ECD results were also verified by OR calculations
performed on the CAM-B3LYP/TZVP PCM/MeOH con-
formers, resulting in large positive rotation values in the range
applied combinations of levels gave moderate to good
agreement with the experimental ECD spectrum recorded in
MeOH. In most cases the BH&HLYP and the CAM-B3LYP
functionals gave better agreement than the B3LYP and the
PBE0 ones. It is also noteworthy that ECD spectra computed
for the two optimized solid-state X-ray conformers with
different conformations exhibit a curve that is nearly a mirror-
image of that of the solution conformers (Figure S3,
Supporting Information and Figure 4). In solution, the
hydrogen of the carboxyl group forms a hydrogen bond with
the oxygen of the carbonyl or OH groups, while in the solid
state intermolecular hydrogen bonds are produced, which give
rise to different orientations of the α,β-unsaturated carboxyl
group. The orientation of the α,β-unsaturated carboxyl group
31,32
of +226 to +263.
Furthermore all low-energy conformers
at all the applied combinations of levels had OR values in the
range of +182 to +308, allowing the unambiguous elucidation
of the absolute configuration (see Supporting Information).
Thus, the absolute configuration and preferred conforma-
tions of 1a and 1b were determined in solution and solid
states. Compound 1a, with a positive optical rotation and S
absolute configuration, was named (+)-(S)-cryptomeriolide,
while compound 1b was named (−)-(R)-cryptomeriolide.
Cryptomeriolide (1) is an example of a 20(1→5)-abeo-
4,5;11,12-bis-seco-abietane diterpenoid isolated from natural
sources. The biosynthetic precursors of cryptomeriolide (1)
30
C
DOI: 10.1021/acs.jnatprod.8b00482
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Journal of Natural Products
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Scheme 1. Synthetic Pathway from Sugiol to 1a and 1b
could presumably be traced back to sugiol, which is widely
distributed in the family Cupressaceae and also isolated
Table 2. Cytotoxic Activities of Compounds 1−7 against the
HL-60, A549, and SGC7901 Cell Lines
3
3
previously from C. fortunei. A concise enantioselective
synthesis of 1a and 1b was effectively accomplished by using
sugiol as a starting material (Scheme 1). First, sugiol was
methylated in the presence of Me SO in acetone and then
IC50 (μM)
a
b
b
compound
1a
HL-60
>100
A549
>100
SGC7901
>100
2
4
oxidized with SeO2 in acidic medium to form the α,β-
unsaturated ketone (2). Demethylation and acetylation
afforded the acetate (3). The acetate was reduced with
NaBH4 in the presence of cerium(III) chloride and
subsequently treated with p-toluenesulfonic acid to yield a
seco ring A precursor (4). Subsequently, the exocyclic double
bond of 4 was oxidized to form an α- or β-orientated
dihydroxy group by AD-mix-α or AD-mix-β according to the
1b
2
>100
>100
>100
>100
>100
>100
3
4
5a
5
6
6
7
7
25.7 ± 0.7
20.8 ± 1.6
21.7 ± 1.1
41.2 ± 2.5
16.0 ± 4.0
17.2 ± 5.7
45.5 ± 1.4
49.4 ± 10.6
0.024 ± 0.01
26.9 ± 0.6
25.5 ± 1.8
50.5 ± 0.3
48.6 ± 0.9
68.5 ± 0.2
28.9 ± 0.7
26.2 ± 0.7
23.5 ± 0.6
22.9 ± 0.5
34.4 ± 0.9
45.8 ± 1.0
47.5 ± 1.6
88.9 ± 10.5
36.7 ± 1.9
31.7 ± 0.4
33.1 ± 1.4
b
a
b
a
b
3
4
method of Sharpless. The diol intermediate (5) was
protected with 2,2-dimethoxypropane to afford compound 6,
which was hydrolyzed to remove the acetyl group. The
resulting product (7) was oxidized to orthoquinone (8) by
c
mitoxantrone
0.036 ± 0.004
b
0.293 ± 0.005
a
Determined by the CCK8 assay. Determined by the SRB assay.
3
5
c
Value given as nM.
Fremy’s salt. The final product 1a or 1b with good
enantiomeric excess was generated by treating 8 with m-
CPBA and then hydrolyzed under acid conditions. The NMR
data and specific rotation values of synthetic 1a and 1b were
both in good agreement with those of the corresponding
natural products, which further confirmed the structural
assignments of naturally occurring compounds 1a and 1b.
The natural and synthetic bis-seco-abietane diterpenoids 1a
and 1b, together with the intermediates from the synthetic
procedures, were evaluated for cytotoxicity against three
human cancer cell lines: HL-60, A549, and SGC7901. The
results (Table 2) indicated that all intermediates (3, 4, 5a/b,
resulting in a small group of seco-abietanes, this type of
36
skeleton was in fact only described in a recent patent. This is
the first time that such a 20(10→5)-abeo-4,5;11,12-bis-seco-
abietane skeleton was unambiguously defined by both single-
crystal X-ray crystallography and synthesis.
EXPERIMENTAL SECTION
■
General Experimental Procedures. Optical rotations were
measured on a Perkin−Elmer 341 polarimeter. IR spectra were
recorded on a Nicolet 6700 spectrometer in KBr pellets. ECD spectra
were measured on a BRIGHTTIME Chirascan. NMR spectra were
recorded on a Varian Mercury-500 NMR spectrometer. The chemical
shift (δ) values are given in ppm with tetramethylsilane as internal
standard, and coupling constants (J) are in Hz. ESIMS and HRESIMS
data were recorded on Waters 2695−3100 LC-MS and Waters Xevo
TOF mass spectrometers. Single-crystal X-ray diffraction measure-
ments were conducted on a Bruker Smart Apex II diffractometer with
a graphite monochromator. Column chromatography (CC) was
performed with silica gel (100−200, 200−300, and 300−400 mesh,
Qingdao Marine Chemical Industrials, Qingdao, People’s Republic of
6
a/b, 7a/b) showed moderate cytotoxic activities with IC50
values ranging from 16.0 to 88.9 μM except for compounds 1a,
1
b, and 2.
In summary, a pair of enantiomeric bis-seco-abietane
diterpenoids from C. fortunei was isolated and fully
characterized. They represent a rare carbon skeleton formed
by migration of an angular methyl group and bond-breaking of
rings A and C. Although different degrees of oxidation of rings
A, B, or C of abietane diterpenoids have been reported,
D
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Journal of Natural Products
Article
China) and MCI gel CHP20P (75−150 μm, Mitsubishi Chemical
Industries, Tokyo, Japan). TLC was carried out on precoated silica gel
GF254 plates (Yantai Chemical Industrials, Yantai, People’s Republic
of China), and the TLC spots were viewed at 254 nm and visualized
with 5% H SO in EtOH containing 10 mg/mL vanillin followed by
Cytotoxicity Bioassays. The antiproliferative activities of
compounds 1−7 against HL-60 (human leukemia), A-549 (human
non-small-cell lung adenocarcinoma), and SGC7901 (human gastric
adenocarcinoma) tumor cell lines were evaluated.
Cell Culture and Reagents. Human acute myeloid leukemia cell
line HL-60 was purchased from the Institute of Biochemistry and Cell
Biology, Chinese Academy of Sciences, and cultured in media
according to recommendations with 10% fetal bovine serum (FBS)
(Gibco, New Zealand). The human non-small-cell lung cancer cell
line A549 was purchased from the American Type Culture Collection
(ATCC) and cultured in media according to recommendations with
10% FBS (Gibco, New Zealand). The human gastric cancer cell line
SGC-7901 was purchased from Shanghai Sixth People’s Hospital and
cultured in RPMI-1640 medium with 10% FBS. The cells were
2
4
heating. Analytical HPLC was performed on a Waters 2695
instrument with an Acquity ELSD detector. Preparative HPLC was
performed on a Varian PrepStar system with an Alltech 3000 ELSD.
Chromatographic separations were carried out on a Waters Sunfire
RP C , 5 μm, 30 mm × 150 mm column, using a gradient solvent
18
system composed of H O and CH CN, with a flow rate of 25.0 mL/
2
3
min. Chiral-phase separation was performed by K-prep LAB100S
YMC, Japan) on a chiral-phase column Daicel Chiralpak AY-H, 10
(
μm, 5.0 cm i.d. × 25.0 cm L column (Daicel Chemical Industries,
Ltd., Japan), using a gradient solvent system composed of hexane/
EtOH/HAc (90:10:0.1). Analytical chiral-phase HPLC was per-
formed by a Shimadzu LC 20A QA&QC-HPLC-02 on a chiral-phase
Daicel Chiralpak AY-H, 5 μm, 0.46 cm i.d. × 15.0 cm L column
cultured at 37 °C and 5% CO . Mitoxantrone, used as the positive
2
control, was purchased from Selleck (TX, USA), and the purity was
99.36%.
Cell Viability Assay. The antiproliferative activity of the
(
Daicel Chemical Industries, Ltd., Japan). All solvents used for CC
compounds against HL-60 cells was determined using the CCK8
37
and HPLC were of analytical grade (Shanghai Chemical Reagents Co.
Ltd.) and gradient grade (Merck KGaA), respectively.
assay (Dojindo, Japan). Briefly, cells were seeded in 96-well plates
3
(8 × 10 cells/well) overnight and then exposed to various
Plant Material. The bark of C. fortunei was collected in She
County, Anhui Province, China, in October 2013, and identified by
Professor Jin-Gui Shen of the Shanghai Institute of Materia Medica. A
voucher specimen (no. 20130506) was deposited at the Herbarium of
the Shanghai Institute of Materia Medica, Chinese Academy of
Sciences.
concentrations of compounds in triplicates for 72 h. Subsequently,
10 μL of CCK8 reagent was added per well and incubated at 37 °C
for 2 h. Cell proliferation was evaluated by optical density, which was
measured using a SpectraMax 190 multiwell spectrophotometer
(Molecular Devices, CA, USA) at 450 nm. The inhibitory rates (%) of
compounds were calculated with the following formula: [1 −
(A450 treated/A450 control)] × 100%. Cell sensitivity was expressed as
IC50 values, which were calculated by the Logit method. The
experiment was repeated independently and shown as the average of
two independent experiments.
Extraction and Isolation. The air-dried and powdered bark of C.
fortunei (110 kg) was extracted with acetone (3 × 330L) at room
temperature. After evaporation of the solvent, part of the residue (2
kg) was dissolved in water and partitioned with PE, CH Cl , and
2
2
EtOAc, successively. The PE fraction was concentrated and dissolved
in 80% aqueous MeOH. The CH Cl fraction and the MeOH layer
The antiproliferative activity of the compounds against A549 and
SGC7901 cells was evaluated using the sulforhodamine B (SRB)
2
2
38
3
3
were combined together based on the TLC profiles and subjected to
assay. Cells (A549: 2 × 10 cells/well and SGC7901: 3 × 10 cells/
well) were seeded into 96-well plates and grown for 24 h. Cells were
then treated with increasing concentrations of compounds and grown
for 72 h. DMSO was used as the vehicle control. At the end of the
exposure time, 100 μL of ice-cold 10% trichloroacetic acid (TCA) was
added to each well, left at 4 °C for 1 h, and washed five times with
distilled water. The TCA-fixed cells were stained for 15 min with 100
CC over MCI gel (EtOH/H O, 60% to 95%) to yield three fractions
2
(
(
A−C). Fraction A (75 g) was subjected to CC over MCI gel
EtOH/H O, 30% to 70%) to give 13 fractions (A1−A13). Fraction
2
A10 (10.8 g) was subjected to CC over silica gel eluting with PE/
acetone (10:1 to 1:2) in a stepwise manner to afford 16 fractions
(
A10A−A10P). Subsequently, fraction A10J (753 mg) was selected
−
1
for separation by preparative HPLC (CH CN/H O, 35% to 45%)
μL of 4 mg mL SRB in 1% HOAc. The plates were washed five
times with 1% HOAc and air-dried overnight. After air-drying,
protein-bound dye was dissolved in 150 μL of 10 mM Tris base for 5
min and was measured at 560 nm using a multiwall spectropho-
tometer (SpectraMax, Molecular Devices, USA). The inhibition rate
was calculated as (1 − A560 treated/A560 control) × 100%. The
experiment was repeated independently, and the results are shown
with the average value of two independent experiments.
3
2
and further purified by CC over silica gel eluting with CH Cl /
2
2
acetone (10:1) to yield cryptomeriolide (1) (44 mg).
2
0
Cryptomeriolide (1): white powder; [α] +159 (c 0.3, MeOH)
D
(
2
(
1a), [α]20 −153 (c 0.3, MeOH) (1b); ECD (MeOH) λ (Δε)
D
max
48 (+30.35), 233 (0), 222 (−25.75), 212 (0), 196 (+42.76) nm
1a), ECD (MeOH) λ (Δε) 248 (−33.42), 233 (0), 222 (+28.43),
max
2
2
12 (0), 197 (−47.03) nm (1b); IR (KBr) ν 3446, 2965, 2928,
max
−1 1
871, 1713, 1466, 1384, 1261, 1182, 1090, 1053, 802 cm ; H and
C NMR, see Table 1; ESIMS m/z 347.3 [M + H] , 345.2 [M −
H] , 691.4 [2M − 1] ; ESIHRMS m/z 369.1662 [M + Na] (calcd
13
+
ASSOCIATED CONTENT
Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acs.jnat-
prod.8b00482.
■
−
−
+
*
S
for C H O Na, 369.1672).
20
26
5
X-ray Crystallographic Analysis of Compound 1. From a
mixture of CH Cl /n-hexane solution, white crystals were obtained,
2
2
and a needle-like crystal was supplied for the X-ray crystallographic
analysis. The structure was solved and refined using the Bruker
SHELXTL Software Package. A total of 35 529 measurements yielded
1D and 2D NMR, IR, and ECD spectra, HRESIMS data
of new compounds, chiral-phase HPLC separation,
enantioselective synthesis, and the X-ray data for
compound 1 (PDF)
9
0
837 unique reflections (Rint = 0.0406), the final R was determined as
.0592, and wR2 was 0.1687 (I > 2σ(I)). Crystal data for 1:
1
C H O , M = 692.82 g/mol, triclinic, space group P-1, a =
40
52 10
1
6
2
7
2.8671(3) Å, b = 14.1050(3) Å, c = 15.2457(3) Å; α =
4.1960(10)°, β = 89.7430(10)°, γ = 63.3900(10)°, V =
AUTHOR INFORMATION
Corresponding Authors
*E-mail: yaosheng@simm.ac.cn (S. Yao).
*Tel: +86 21 50806600. Fax: +86 21 50807088. E-mail: yye@
mail.shcnc.ac.cn (Y. Ye).
■
3
−1
163.93(8) Å , Z = 2, T = 170 K, μ(Mo Kα) = 0.076 mm , F =
3
44, Dcalc = 1.063 g/cm . Crystallographic data for 1 have been
deposited at the Cambridge Crystallographic Data Centre as deposit
no. CCDC 1446792. Copies of these data can be obtained free of
charge via the Internet at www.ccdc.cam.ac.uk/conts/retrieving.html
or on application to CCDC, 12 Union Road, Cambridge CB2 1EZ,
UK [tel: (+44) 1223-336-408; fax: (+44) 1223-336-033; e-mail:
deposit@ccdc.cam.ac.uk].
ORCID
Attila Man
Yang Ye: 0000-0003-1316-5915
́
di: 0000-0002-7867-7084
E
DOI: 10.1021/acs.jnatprod.8b00482
J. Nat. Prod. XXXX, XXX, XXX−XXX

Journal of Natural Products
Article
Notes
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The authors declare no competing financial interest.
(
ACKNOWLEDGMENTS
■
4603−4625.
We are thankful for the financial support of the National
Science & Technology Major Project of China “Key New Drug
Creation and Manufacturing Program” (2015ZX09103002).
Our thanks are also given to the National Natural Science
Foundation of China (81473112, 81573305) and the Interna-
tional Partnership Program of the Chinese Academy of
Sciences (153631KYSB20160004). T.K. and A.M. thank the
National Research, Development and Innovation Office (NKFI
K120181 and PD121020) for financial support and the
Governmental Information-Technology Development Agency
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393, 51−57.
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DOI: 10.1021/acs.jnatprod.8b00482
J. Nat. Prod. XXXX, XXX, XXX−XXX