Briaroxalides : Novel diepoxybriarane diterpenes from an okinawan gorgonian briareum SP.

Article abstract of DOI:10.3987/COM-11-12383

Seven new 8,17- and 11,12-diepoxybriarane diterpenoids, briaroxalides A-G (1-7), were isolated from an Okinawan gorgonian Briareum sp. The structures of the diterpenoids were determined on the basis of spectroscopic analysis, chemical conversions and X-ra

Full text of DOI:10.3987/COM-11-12383

HETEROCYCLES, Vol. 85, No. 1, 2012
135
HETEROCYCLES, Vol. 85, No. 1, 2012, pp. 135 - 145. © 2012 The Japan Institute of Heterocyclic Chemistry
Received, 31st October, 2011, Accepted, 30th November, 2011, Published online, 6th December, 2011
DOI: 10.3987/COM-11-12383
BRIAROXALIDES : NOVEL DIEPOXYBRIARANE DITERPENES FROM
AN OKINAWAN GORGONIAN BRIAREUM SP.
Koichiro Ota, Naoko Okamoto, Hidemichi Mitome, and Hiroaki Miyaoka*
School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, 1432-1
Horinouchi, Hachioji, Tokyo 192-0392, Japan; E-mail: miyaokah@toyaku.ac.jp
Abstract – Seven new 8,17- and 11,12-diepoxybriarane diterpenoids,
briaroxalides A-G (1-7), were isolated from an Okinawan gorgonian Briareum sp.
The structures of the diterpenoids were determined on the basis of spectroscopic
analysis, chemical conversions and X-ray diffraction analysis.
INTRODUCTION
Briarane-type diterpenes have received a great deal of interest due to their structural features and
biological activities. These diterpenoids are characterized by highly oxygenated
a
bicyclo[8.4.0]tetradecane skeleton, and most also contain a -lactone moiety. The range of activities
reported for some briarane-type diterpenes include cytotoxic,1-3 antiviral,4,5 anti-inflamatory,6,7
insecticidal,8,9 immunomodulation,10 and reversal of multidrug resistance.11 We examined the chemical
constituents of Briareum sp., collected at Ishigaki Island in the region of Okinawa Prefecture in Japan.12
During the course of our investigations, seven new 8,17-, and 11,12-diepoxybriarane diterpenoids 1-7,
designated as briaroxalides A-G,13 have been isolated. Herein we report on the isolation and structural
elucidation of these new briarane diterpenoids, including the determination of absolute configuration
based on chemical conversions and X-ray diffraction analysis.
RESULTS AND DISCUSSION
Gorgonian specimens of Briareum sp. (177 g), collected from the coral reef of Ishigaki Island, Okinawa,
Japan in 2004, were immersed in MeOH. Repeated chromatographic separation of the combined extracts
led to the purification and subsequent characterization of seven new 8,17- and 11,12-diepoxybriaranes,
briaroxalides A-G (1-7) (Figure 1), along with known diterpenoids brianthein A,11 violide G14 and
briarlide R.15

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HETEROCYCLES, Vol. 85, No. 1, 2012
OR2
R1O
R4O15
3
16
4
2
9
5
briaroxalide A (1) : R11 = H , R22 = Ac, R33 = H , R44 = Ac
14
1
13
12
briaroxalide B (2)
: R1 = H , R2 = H , R3 = Ac, R4 = Ac
10
6
11
briaroxalide C 3
( ) : R = H , R = Ac, R = Ac, R = Ac
7
8
H
O
briaroxalide D (4) : R11 = H , R22 = H , R33 = Ac, R44 = H
O
H
briaroxalide E (5)
: R1 = H , R2 = Ac, R3 = Ac, R4 = H
R3O
20
O
briaroxalide F 6
( ) : R = Ac, R = Ac, R = Ac, R = H
19
17
briaroxalide G (7) : R1 = Ac, R2 = H , R3 = Ac, R4 = Ac
18
O
Structures of briaroxalides A-G ( - )
1 7
Figure 1.
Briaroxalide A (1) was obtained as colorless needles and the molecular formula was found to be
C24H32O10 as determined from high-resolution ESIMS. The IR spectrum of 1 suggested the presence of a
-lactone (1771 cm-1), ester (1740 cm-1) and hydroxy group (3537, 3449 cm-1). The 13C, 1H NMR (Table 1
and 2) and DEPT spectra showed signals indicating six methyl, two sp3 methylene, one sp3 methine, six
sp3 oxymethine, one sp3 quaternary carbon, three sp3 quaternary carbons bearing an oxygen functional
group, two sp2 carbons, and three carbonyl carbons. These spectral data, coupled with the degree of
unsaturation (9), suggested that compound 1 is a tricyclic diterpenoid possessing a -lactone [C 171.8
(C)], two epoxide [C 71.1 (C), C 64.6 (C), C 60.4 (C), C 59.9 (CH)], two acetoxyl [H 2.10 (3H, s), H
2.03 (3H, s), C 171.3 (C), C 170.8 (C)], and a trisubstituted olefin [H 5.27 (1H, d, J = 9.1 Hz), C 140.4
(C), C 120.3 (CH)] moiety. COSY cross-peaks indicated sequences of C-3 to C-4, C-6 to C-7, C-9 to
C-10, and C-12 to C-14. The planar structure of 1 was determined on the basis of the following
correlations (Figure 2) in the HMBC spectrum: H-2/C-1, C-4, C-10, C-15; H-3/C-3-Ac (C=O); H-4 (H
2.99)/C-2, C-5, C-6; H-6/C-4, C-16; H-7/C-5, C-19; H-9/C-7, C-8, C-11, C-17; H-10/C-1, C-8, C-11,
C-15; H-13 (H 2.24)/C-11; H-14/C-1, C-2, C-10, C-14-Ac (C=O); H-15/C-1, C-2, C-10, C-14; H-16/C-4,
C-5, C-6; H-18/C-8, C-17, C-19; H-20/ C-10, C-11, C-12; H-3-Ac (Me)/C-3-Ac (C=O); H-14-Ac
(Me)/C-14-Ac (C=O).
H3C
O
H3C
O
HO
O
O
CH3
CH3
O
O
CH3
O
HO
: 1H-1H COSY
: HMBC
H3C
O
1
Figure 2. Selective 1H H COSY and HMBC correlations of 1
-

HETEROCYCLES, Vol. 85, No. 1, 2012
137
Table 1. 13C-NMRa spectral data (150 MHz, CDCl3) for compounds 1-7
no
5
7
1
2
3
4
6
1
2
3
4
5
6
7
8
45.9 (C)
69.0 (CH)
73.0 (CH)
34.4 (CH2)
140.4 (C)
120.3 (CH)
75.5 (CH)
71.1 (C)
46.1 (C)
68.6 (CH)
68.4 (CH)
39.4 (CH2)
141.3 (C)
119.3 (CH)
74.7 (CH)
70.3 (C)
45.7 (C)
69.5 (CH)
72.0 (CH)
34.4 (CH2)
139.7 (C)
120.9 (CH)
74.8 (CH)
70.3 (C)
45.5 (C)
72.5 (CH)
68.3 (CH)
39.6 (CH2)
142.3 (C)
118.5 (CH)
74.6 (CH)
71.1 (C)
46.0 (C)
72.6 (CH)
72.2 (CH)
34.4 (CH2)
141.2 (C)
119.8 (CH)
74.8 (CH)
71.1 (C)
45.7 (C)
73.8 (CH)
71.3 (CH)
34.9 (CH2)
141.5 (C)
120.2 (CH)
74.5 (CH)
70.8 (C)
44.5 (C)
73.3 (CH)
68.9 (CH)
39.4 (CH2)
141.9 (C)
119.0 (CH)
74.6 (CH)
70.3 (C)
9
69.2 (CH)
44.5 (CH)
60.4 (C)
59.9 (CH)
26.7 (CH2)
73.3 (CH)
13.3 (CH3)
26.5 (CH3)
64.6 (C)
68.8 (CH)
43.4 (CH)
59.7 (C)
59.3 (CH)
26.4 (CH2)
73.7 (CH)
13.8 (CH3)
27.2 (CH3)
65.1 (C)
69.2 (CH)
44.0 (CH)
59.5 (C)
59.3 (CH)
26.1 (CH2)
73.0 (CH)
13.4 (CH3)
26.9 (CH3)
65.8 (C)
67.6 (CH)
43.7 (CH)
61.6 (C)
62.4 (CH)
26.0 (CH2)
72.9 (CH)
15.0 (CH3)
27.9 (CH3)
63.8 (C)
68.0 (CH)
43.8 (CH)
61.5 (C)
62.3 (CH)
26.1 (CH2)
72.2 (CH)
14.4 (CH3)
27.5 (CH3)
64.3 (C)
67.5 (CH)
42.1 (CH)
61.2 (C)
62.0 (CH)
25.8 (CH2)
72.3 (CH)
15.8 (CH3)
26.8 (CH3)
63.7 (C)
9.6 (CH3)
170.7 (C)
26.3 (CH3)
169.9 (C)
20.7 (CH3)
170.4 (C)
21.2 (CH3)
169.8 (C)
21.1 (CH3)
68.0 (CH)
43.0 (CH)
58.9 (C)
58.8 (CH)
24.9 (CH2)
72.1 (CH)
15.1 (CH3)
26.6 (CH3)
64.1 (C)
9.9 (CH3)
170.5 (C)
25.9 (CH3)
169.6 (C)
20.8 (CH3)
10
11
12
13
14
15
16
17
18
19
20
2-Ac
10.5 (CH3)
171.8 (C)
22.9 (CH3)
10.6 (CH3)
170.4 (C)
24.7 (CH3)
11.2 (CH3)
170.1 (C)
23.7 (CH3)
9.6 (CH3)
170.6 (C)
26.0 (CH3)
10.0 (CH3)
170.5 (C)
25.9 (CH3)
3-Ac
9-Ac
170.8 (C)
21.4 (CH3)
169.4 (C)
21.2 (CH3)
169.5 (C)
20.9 (CH3)
170.6 (C)
21.1 (CH3)
170.1 (C)
21.3 (CH3)
169.7 (C)
21.0 (CH3)
169.1 (C)
21.1 (CH3)
172.7 (C)
21.2 (CH3)
169.7 (C)
21.4 (CH3)
169.4 (C)
21.1 (CH3)
170.9 (C)
21.4 (CH3)
14-Ac
171.3 (C)
21.1 (CH3
)
a Carbon multiplicities were determined by DEPT experiments.

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HETEROCYCLES, Vol. 85, No. 1, 2012
Furthermore, the p-bromobenzoate derivative (8), which would be amenable to X-ray diffraction analysis
for determination of the relative and absolute configuration, was prepared by treatment of 1 with
p-bromobenzoic acid in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride
(EDC·HCl) and DMAP (Figure 3). Results showed that the absolute configuration of p-bromobenzoate 8
was 1S, 2R, 3R, 7S, 8R, 9S, 10S, 11S, 12R, 14S, 17R on the basis of the Flack parameter [0.003 (3)] in the
X-ray analysis (Figure 4). This indicated that briaroxalide A (1) possesses the same 1S, 2R, 3R, 7S, 8R, 9S,
10S, 11S, 12R, 14S, 17R configuration.
Br
O
OAc
O
AcO
H
O
H
HO
O
O
O
8
Figure 4. ORTEP drawing of p -bromobenzoate 8
Figure 3. Structure of p-bromobenzoate 8
Briaroxalide B (2), obtained as a white amorphous compound with absorption bands at 1777 (-lactone),
1730 (ester) and 3388 cm-1 (OH) in its IR spectrum. The molecular formula of 2 was determined as
13
1
C24H32O10 from high-resolution ESIMS. Comparison of the C and H NMR data of 2 with those of 1
(Tables 1 and 2) showed the presence of a 20-carbon skeleton of a briarane diterpenoid having two
hydroxy and two acetoxy groups. Furthermore, the HMBC correlation between H-9 (H 5.72) and the
carbonyl carbon (C 169.1) of the acetyl group, and between H-14 (H 5.16) and the carbonyl carbon (C
172.7) of the acetyl group revealed the position of two acetates attached to C-9 and C-14.
Briaroxalide C (3), obtained as colorless needles, was found to have the molecular formula C26H34O11 as
determined from high-resolution ESIMS. The IR spectrum of 3 suggested the presence of a -lactone
(1784 cm-1), ester (1723 cm-1) and hydroxy group (3513 cm-1). Comparison of the 13C and 1H NMR data
of 3 with those of 1 (Tables 1 and 2) showed the presence of a briarane diterpenoid having a hydroxy and

HETEROCYCLES, Vol. 85, No. 1, 2012
139
three acetoxy groups. Furthermore, the HMBC correlation between H-3 (H 5.69) and the carbonyl carbon
(C 169.4) of the acetyl group, H-9 (H 5.64) and the carbonyl carbon (C 169.5) of the acetyl group, and
between H-14 (H 5.12) and the carbonyl carbon (C 170.6) of the acetyl group revealed the position of
three acetates, one each attached to C-3, C-9 and C-14. Thus, the structure of 3 represents the
diastereomer at C-11, 12 of the known briarane-type diterpene, brianthein C.11
Briaroxalide D (4), obtained as a colorless oil with absorption bands at 1782 (-lactone), 1758 (ester) and
3457 cm-1 (OH) in its IR spectrum. The molecular formula of 4 was determined as C22H30O9 from
13
1
high-resolution ESIMS. Comparison of the C and H NMR data of 4 with those of 1 (Tables 1 and 2)
showed the presence of a briarane diterpenoid having three hydroxy and an acetoxy groups. Furthermore,
the HMBC correlation between H-9 (H 5.78) and the carbonyl carbon (C 169.7) of the acetyl group
revealed the position of one acetate attached to C-9.
Briaroxalide E (5) was obtained as a colorless oil with molecular formula C24H32O10 as determined from
high-resolution ESIMS. The IR spectrum of 5 suggested the presence of a -lactone (1783 cm-1), ester
(1732 cm-1) and hydroxy group (3478 cm-1). Comparison of the 13C and 1H NMR data of 5 with those of
1 (Tables 1 and 2) showed the presence of a briarane diterpenoid having two hydroxy and two acetoxy
groups. Furthermore, the HMBC correlation between H-3 (H 5.53) and the carbonyl carbon (C 170.1) of
the acetyl group, and between H-9 (H 5.82) and the carbonyl carbon (C 169.7) of the acetyl group
revealed the position of two acetates, one each attached to C-3 and C-9.
Briaroxalide F (6), a white amorphous compound, and briaroxalide G (7), a colorless oil were assigned as
13
1
C26H34O11 on the basis of high-resolution ESIMS. Comparison of the C and H NMR data of 6 and 7
with those of 1 (Tables 1 and 2) showed the presence of a briarane diterpenoid having three hydroxy and
an acetoxy groups. Moreover, the decision of the position of three acetates was similar to that of other
briaroxalides.
The relative and absolute configurations of briaroxalides B-G (2-7) were determined by chemical
conversion and comparison with the spectral data for tetraacetate 9 derived from briaroxalide A (1), the
absolute configuration of which had been determined. Briaroxalides B-G (2-7) and briaroxalide A (1)
were treated with acetic anhydride, pyridine and a catalytic amount of DMAP (except for 2 and 3) to
1
afford tetraacetate 9 (Figure 5). The H and 13C NMR spectra, []D value, and melting point of
tetraacetate 9 were identical with those of 9. These results clearly indicated that each of the briaroxalides
A-G (1-7) possess the same absolute configuration. The biological activity of the seven new briarane
diterpenoids briaroxalides A-G (1-7) reported here is currently under investigation.

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OAc
AcO
AcO
H
O
H
AcO
O
O
O
9
Figure 5.
9
Structure of tetraacetate
EXPERIMENTAL
General Experimental Procedures. Optical rotations were measured with a JASCO P-1030 polarimeter.
1
13
IR spectra were recorded with a JASCO FT-IR/620 spectrometer. H and C NMR spectra were taken
with Bruker Biospin AV-600 spectrometer. Chemical shifts were expressed on a  (ppm) scale with
tetramethylsilane (TMS) as the internal standard (s, singlet; d, doublet; t, triplet; m, multiplet; br, broad).
High resolution ESIMS (HRESIMS) spectra were obtained using a Micromass LCT spectrometer. Single
crystal X-ray diffraction was recorded using a Mac Science Co., Ltd DIP 2020 Image Plate. Flash column
chromatography was carried out on Kanto Chemical silica gel 60N (spherical, neutral) 40-50 m.
Animal Material. The gorgonian specimens of Briareum sp. were obtained from the coral reef of
Ishigaki Island, Okinawa, Japan, in 2004. A voucher specimen has been deposited at Tokyo University of
Pharmacy and Life Sciences (SC-04-5).
Extraction and Isolation. Wet specimens (177 g) were cut into small pieces and extracted with MeOH
(900 mL×3). The combined extracts were concentrated to give a green residue (13.4 g). The part of
MeOH-extracted portion (1.01 g) was chromatographed on silica gel using an AcOEt-MeOH (2 : 1, 1 : 1,
1 : 2) gradient and MeOH as eluent to produce fraction 1 (373 mg), and 2 (136 mg). Fraction 1 was
subjected to flash silica gel column chromatography using a hexane-AcOEt (2 : 1, 1 : 1, 1 : 5, 0 : 1)
gradient and MeOH to give fractions 1-1 (23.0 mg), 1-2 (18.4 mg), 1-3 (91.9 mg), 1-4 (152 mg), 1-5
(33.1 mg), and 1-6 (29.2 mg). Fraction 1-3 was subjected to flash silica gel column chromatography
(elution with hexane-AcOEt (1 : 1, 1 : 5, 0 : 1)) to give briaroxalide A (1) (5.4 mg) and violide G (5.9 mg).
Fraction 1-4 was subjected to flash silica gel column chromatography (elution with CHCl3-AcOEt (2 : 1))
to give briaroxalide A (1) (21.6 mg), briaroxalide B (2) (6.8 mg), and briaroxalide C (3) (36.4 mg).
Fraction 1-5 was subjected to flash silica gel column chromatography (elution with hexane-AcOEt (1 : 5))
to give briaroxalide D (4) (14.1 mg). Fraction 1-6 was subjected to flash silica gel column
chromatography (elution with CHCl3-MeOH (30 : 1)) to give briaroxalide E (5) (39.1 mg). And then the
rest of MeOH-extracted portion (12.4 g) was chromatographed on silica gel using an AcOEt-MeOH (2 : 1,

HETEROCYCLES, Vol. 85, No. 1, 2012
141
1 : 1, 1 : 2) gradient and MeOH as eluent to produce fraction 3 (4.50 g), and 4 (804 mg). Fraction 3 was
subjected to flash silica gel column chromatography using a hexane-AcOEt (2 : 1, 1 : 2, 0 : 1) gradient
and MeOH to give fractions 3-1 (359 mg), 3-2 (1.83 g), 3-3 (2.02 g), and 3-4 (282 mg). Fraction 3-1 was
subjected to flash silica gel column chromatography (elution with CHCl3-AcOEt (7 : 1)) to give
brianthein A (26.5 mg). Fraction 3-2 was subjected to flash silica gel column chromatography using a
hexane-AcOEt (2 : 1, 1 : 2, 0 : 1) gradient and MeOH to give fractions 3-2-1 (86.9 mg), 3-2-2 (215 mg),
and 3-2-3 (1.47 g). Fraction 3-2-2 was subjected to flash silica gel column chromatography (elution with
CHCl3-AcOEt (5 : 1)) to give violide G (88.2 mg), and briarlide R (10.3 mg). Fraction 3-2-3 was
subjected to flash silica gel column chromatography using a CHCl3-AcOEt (5 : 1) gradient and MeOH to
give fractions 3-2-3-1 (462 mg), 3-2-3-2 (338 mg), 3-2-3-3 (89.5 mg), 3-2-3-4 (447 mg), and 3-2-3-5
(234 mg). Fraction 3-2-3-1 was subjected to flash silica gel column chromatography (elution with
CHCl3-AcOEt (10 : 1)) to give briaroxalide C (3) (238 mg), briaroxalide F (6) (34.0 mg), and briarlide R
(38.8 mg). Fraction 3-2-3-2 was subjected to flash silica gel column chromatography (elution with
hexane-AcOEt (1 : 1)) to give briaroxalide C (3) (322 mg). Fraction 3-2-3-4 was subjected to flash silica
gel column chromatography (elution with CHCl3-MeOH (40 : 1)) to give briaroxalide A (1) (222 mg), and
briaroxalide G (7) (18.9 mg).
Briaroxalide A (1): colorless needles (Et2O); mp 220-222 ˚C; []25 +132.3 (c 0.27, MeOH); IR (KBr)
D
max: 3537, 3449, 1771, 1740, 1248 cm-1; 13C-NMR and 1H-NMR, see Table 1 and 2; COSY correlarions
(H/H) H-3/H-4 (H 2.03); H-3/H-4 (H 2.99); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.24); H-14/H-13 (H
2.24); HMBC correlations (H/C) H-2/C-1, C-4, C-10, C-15; H-3/C-3-Ac (C=O); H-4 (H 2.99)/C-2, C-3,
C-5, C-6; H-6/C-4, C-16; H-7/C-5, C-6, C-19; H-9/C-7, C-8, C-11, C-17; H-10/C-1, C-3, C-8, C-9, C-11,
C-15; H-12/C-13, C-14; H-13 (H 2.24)/C-11, C-12, C-14; H-14/C-1, C-2, C-10, C-12, C-13, C-14-Ac
(C=O); H-15/C-1, C-2, C-10, C-14; H-16/C-4, C-5, C-6; H-18/C-8, C-17, C-19; H-20/C-9, C-10, C-11,
C-12; H-3-Ac (Me)/C-3-Ac (C=O); H-14-Ac (Me)/C-14-Ac (C=O); ESIMS m/z 503 [M++Na] (100);
HRESIMS m/z 503.1892 (calcd for C24H32O10Na: M++Na, 503.1893).
Briaroxalide B (2): white amorphous; mp 208-211 ˚C; []25 +32.7 (c 0.34, CHCl3); IR (KBr) max:
D
1
3388, 2993, 1777, 1730 cm-1; 13C-NMR and H-NMR, see Table 1 and 2; COSY correlarions (H/H)
H-3/H-4 (H 2.18); H-3/H-4 (H 2.74); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.13), H-13 (H 2.18);
H-14/H-13 (H 2.13), H-13 (H 2.18); HMBC correlations (H/C) H-2/C-1, C-2, C-4, C-10, C-14, C-15;
H-3/C-1, C-3, C-4; H-4 (H 2.18)/C-3, C-5, C-6, C-16; H-4 (H 2.74)/C-2, C-3, C-5, C-16; H-6/C-4, C-16;
H-7/C-5, C-6, C-19; H-9/C-1, C-7, C-8, C-10, C-11, C-17, C-9-Ac (C=O); H-10/C-1, C-5, C-11;
H-12/C-13, C-20; H-13 (H 2.13)/C-1, C-11, C-12, C-14; H-14/C-10, C-12, C-13, C-14-Ac (C=O);
H-15/C-1, C-2, C-10, C-14; H-16/C-4, C-5, C-6; H-18/C-8, C-17, C-19; H-20/C-9, C-10, C-11, C-12;

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H-9-Ac (Me)/C-9-Ac (C=O); H-14-Ac (Me)/C-14-Ac (C=O); ESIMS m/z 481 [M++H] (100); HRESIMS
m/z 481.2103 (calcd for C24H33O10: M++H, 481.2074).
Briaroxalide C (3): colorless needles (Et2O); mp 198-201 ˚C; []25 +130.0 (c 0.20, EtOH); IR (KBr)
D
1
max: 3513, 1784, 1723, 1249 cm-1; 13C-NMR and H-NMR, see Table 1 and 2; COSY correlarions
(H/H) H-3/H-4 (H 2.03); H-3/H-4 (H 3.03); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.23), H-13 (H 2.16);
H-14/H-13 (H 2.23), H-13 (H 2.16); HMBC correlations (H/C) H-2/C-1, C-15; H-3/C-1, C-4, C-3-Ac
(C=O); H-4 (H 2.03)/C-3, C-5, C-6, C-16; H-4 (H 3.03)/C-2, C-3, C-5, C-6, C-16; H-6/C-4, C-16;
H-7/C-5, C-6, C-19; H-9/C-1, C-7, C-8, C-10, C-11, C-9-Ac (C=O); H-10/C-1, C-2, C-11, C-14, C-15;
H-12/C-13, C-14; H-13 (H 2.16)/C-1, C-12, C-14; H-14/C-1, C-2, C-10, C-12, C-13, C-14-Ac (C=O);
H-15/C-1, C-2, C-10, C-14; H-16/C-4, C-5, C-6; H-18/C-7, C-17, C-19; H-20/C-9, C-10, C-11, C-12;
H-3-Ac (Me)/C-3-Ac (C=O); H-9-Ac (Me)/C-9-Ac (C=O); C-14-Ac (Me)/C-14-Ac (C=O); ESIMS m/z
545 [M++Na] (100); HRESIMS m/z 545.2015 (calcd for C26H34O11Na: M++Na, 545.1999).
Briaroxalide D (4): colorless oil; []25D –39.7 (c 0.19, CHCl3); IR (neat) max: 3457, 2917, 1782, 1758
13
1
cm-1; C-NMR and H-NMR, see Table 1 and 2; COSY correlarions (H/H) H-3/H-4 (H 2.04); H-3/H-4
(H 2.83); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.16), H-13 (H 2.23); H-14/H-13 (H 2.16), H-13 (H
2.23); HMBC correlations (H/C) H-2/C-1, C-4, C-10, C-15; H-3/C-1, C-4; H-4 (H 2.04)/C-3, C-5, C-6,
C-16; H-4 (H 2.83)/C-2, C-3, C-5, C-6; H-6/C-4, C-16; H-7/C-5, C-6, C-19; H-9/C-7, C-8, C-10, C-11,
C-9-Ac (C=O); H-10/C-1, C-2, C-8, C-9, C-11, C-15; H-12/C-14, C-20; H-14/C-10, C-12; H-15/C-10,
C-14; H-16/C-4, C-5, C-6; H-18/C-8, C-17, C-19; H-20/C-9, C-11, C-12; H-9-Ac (Me)/C-9-Ac (C=O);
ESIMS m/z 439 [M++H] (100); HRESIMS m/z 439.2042 (calcd for C22H31O9: M++H, 439.1968).
Briaroxalide E (5): colorless oil; []25D +44.1 (c 0.37, CHCl3); IR (neat) max: 3478, 2977, 1783, 1732
13
1
cm-1; C-NMR and H-NMR, see Table 1 and 2; COSY correlarions (H/H) H-3/H-4 (H 2.13); H-3/H-4
(H 2.99); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.13), H-13 (H 2.24); H-14/H-13 (H 2.13), H-13 (H
2.24); HMBC correlations (H/C) H-2/C-1, C-2, C-4, C-10, C-15; H-3/C-1, C-4, C-3-Ac (C=O); H-4 (H
2.13)/C-3, C-5, C-6, C-14, C-16; H-4 (H 2.99)/C-5, C-16; H-6/C-4, C-16; H-7/C-5, C-6, C-19; H-9/C-7,
C-8, C-10, C-11, C-9-Ac (C=O); H-10/C-1, C-9, C-11, C-15; H-12/C-13, C-14; H-13 (H 2.24)/C-1, C-3,
C-11, C-14; H-14/C-10, C-12, C-13; H-15/C-1, C-2, C-3, C-10, C-14; H-16/C-4, C-5, C-6; H-18/C-8,
C-17, C-19; H-20/C-9, C-10, C-11; H-3-Ac (Me)/C-3-Ac (C=O); H-9-Ac (Me)/C-9-Ac (C=O); ESIMS
m/z 503 [M++Na] (100); HRESIMS m/z 503.1916 (calcd for C24H32O10Na: M++Na, 503.1916).
Briaroxalide F (6): white amorphous; mp 187-190 ˚C; []25 +70.6 (c 0.32, CHCl3); IR (KBr) max:
D
1
3490, 3001, 1792, 1747 cm-1; 13C-NMR and H-NMR, see Table 1 and 2; COSY correlarions (H/H)
H-2/H-3, H-3/H-4 (H 2.24); H-3/H-4 (H 2.90); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.11), H-13 (H

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2.24); H-14/H-13 (H 2.11), H-13 (H 2.24); HMBC correlations (H/C) H-2/C-1, C-4, C-10, C-14, C-15,
C-2-Ac (C=O); H-3/C-1, C-4, C-3-Ac (C=O); H-4 (H 2.24)/C-3, C-5, C-16; H-4 (H 2.90)/C-2, C-3, C-5;
H-6/C-4; H-7/C-6, C-19; H-9/C-7, C-8, C-10, C-11, C-9-Ac (C=O); H-10/C-1, C-2, C-5, C-8, C-9;
H-12/C-13, C-14; H-14/C-10, C-12, C-13; H-15/C-1, C-2, C-10, C-14; H-16/C-4; H-18/C-7, C-8, C-17;
H-20/C-9, C-10, C-11; ESIMS m/z 523 [M++H] (100); HRESIMS m/z 523.2171 (calcd for C26H35O11:
M++H, 523.2179).
Briaroxalide G (7): colorless oil; []25D +47.8 (c 0.095, CHCl3); IR (neat) max: 3524, 2979, 1782, 1748
13
1
cm-1; C-NMR and H-NMR, see Table 1 and 2; COSY correlarions (H/H) H-3/H-4 (H 1.97); H-3/H-4
(H 2.64); H-6/H-7; H-9/H-10; H-12/H-13 (H 2.09), H-13 (H 2.22); H-14/H-13 (H 2.09), H-13 (H
2.22); HMBC correlations (H/C) H-2/C-1, C-4, C-14, C-15, C-2-Ac (C=O); H-3/C-1, C-4; H-4 (H
1.97)/C-3, C-5, C-6, C-16; H-4 (H 2.64)/C-2, C-3, C-5, C-6; H-6/C-4, C-16; H-7/C-5, C-6, C-19;
H-9/C-7, C-8, C-11, C-17, C-9-Ac (C=O); H-10/C-1, C-2, C-8, C-9, C-11, C-12, C-15; H-12/C-13, C-14;
H-13 (H 2.22)/C-1, C-12, C-14; H-14/C-2, C-10, C-12, C-13, C-14-Ac (C=O); H-15/C-1, C-2, C-10,
C-14; H-16/C-3, C-4, C-5, C-6; H-18/C-8, C-17, C-19; H-20/C-9, C-10, C-11, C-12; H-9-Ac
(Me)/C-9-Ac (C=O), H-14-Ac (Me)/C-14-Ac (C=O); ESIMS m/z 523 [M++H] (100), 463 (M+–Ac);
HRESIMS m/z 523.2180 (calcd for C26H35O11: M++H, 523.2179).
Synthesis of p-bromobenzoate 8 from briaroxalide A (1). To a solution of briaroxalide A (1) (11.5 mg,
23.9 mol) in CH2Cl2 (400 L) were added p-bromobenzoic acid (96.2 mg, 479 mol), EDC·HCl (91.8
mg, 479 mol), and DMAP (0.1 mg, 0.820 mol). The mixture was stirred at room temperature for 24 h.
The reaction mixture was diluted with Et2O, washed with H2O and saturated aqueous NaCl, dried over
Na2SO4, and concentrated under reduced pressure. The residue was purified by silica gel column
chromatography (elusion with hexane-AcOEt (1 : 1)) to give p-bromobenzoate 8 (10.3 mg, 65% yield):
colorless scales (toluene); mp 168-170 ˚C; []25 +145.9 (c 0.39, CHCl3); IR (KBr) max: 3450, 2934,
D
1782, 1728 cm-1; 1H-NMR (600 MHz, CDCl3)  ppm: 7.87 (2H, m), 7.60 (2H, m), 6.11 (1H, dd, J = 5.9,
12.3 Hz), 5.74 (1H, d, J = 8.8 Hz), 5.52 (1H, s), 5.36 (1H, d, J = 8.8 Hz), 4.68 (1H, s), 4.59 (1H, d, J = 3.8
Hz), 3.93 (1H, s), 2.93 (1H, d, J = 2.8 Hz), 2.84 (1H, s), 2.68 (1H, s), 2.17 (2H, s), 2.12 (3H, s), 2.01 (3H,
13
s), 1.93 (3H, s), 1.90 (1H, m), 1.69 (3H, s), 1.53 (3H, s), 1.26 (3H, s); C-NMR (150 MHz, CDCl3) 
ppm: 171.6, 171.5, 170.9, 164.6, 139.9, 131.9×2, 131.2×2, 128.8, 128.4, 120.4, 75.6, 72.8, 72.0, 71.9,
71.1, 69.1, 64.3, 59.7, 59.6, 45.7, 43.6, 34.6, 30.9, 26.4, 26.0, 24.2, 21.5, 15.1, 10.3; ESIMS m/z 685
[M++Na] (100); HRESIMS m/z 685.1260 (calcd for C31H35O11BrNa: M++Na, 685.1260).
Synthesis of tetraacetate 9 from briaroxalides A-G (1-7). To a solution of briaroxalides A-G (1-7)
(3.30-11.0 mol) in pyridine (200 L) were added acetic anhydride (100 L) and catalytic amount of
DMAP (except for 2 and 3). The mixture was stirred at room temperature for 3-96 h. The reaction mixture

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was concentrated under reduced pressure. The residue was purified by silica gel column chromatography
(elusion with hexane-AcOEt (1 : 1)) to give tetraacetate 9 (44-89% yield) as a white amorphous.
Tetraacetate 9 from briaroxalide A (1): mp 183-184 ˚C; []25D +101.1 (c 0.15, CHCl3); IR (KBr) max:
1
2924, 1779, 1730 cm-1; H-NMR (600 MHz, CDCl3)  ppm: 5.86 (1H, d, J = 2.2 Hz), 5.62 (1H, dd, J =
6.0, 11.6 Hz), 5.55 (1H, d, J = 8.9 Hz), 5.37 (1H, d, J = 9.0 Hz), 5.19 (1H, s), 4.71 (1H, d, J = 3.5 Hz),
2.93 (1H, s), 2.85 (1H, m), 2.80 (1H, d, J = 2.3 Hz), 2.21 (1H, m), 2.14 (3H, s), 2.11 (3H, s), 2.08 (3H, s),
13
2.04 (1H, m), 1.98 (3H, s), 1.95 (3H, s), 1.68 (3H, s), 1.49 (3H, s), 1.03 (3H, s); C-NMR (150 MHz,
CDCl3)  ppm: 170.8, 170.5, 170.3, 170.1, 169.7, 140.4, 120.5, 74.8, 72.1, 71.8, 71.4, 70.2, 68.4, 64.6,
59.0, 58.8, 44.7, 42.8, 35.0, 26.6, 25.3, 25.2, 21.3, 21.3, 21.0, 20.7, 15.0, 10.3; ESIMS m/z 587 [M++Na]
(100); HRESIMS m/z 587.2108 (calcd for C28H36O12Na: M++Na, 587.2104).
Tetraacetate 9 from briaroxalide B (2): mp 177-179 ˚C; []25D +141.0 (c 0.085, CHCl3).
Tetraacetate 9 from briaroxalide C (3): mp 184-185 ˚C; []25D +130.6 (c 0.16, CHCl3).
Tetraacetate 9 from briaroxalide D (4): mp 184-185 ˚C; []25D +106.6 (c 0.060, CHCl3).
Tetraacetate 9 from briaroxalide E (5): mp 182-184 ˚C; []25D +109.9 (c 0.16, CHCl3).
Tetraacetate 9 from briaroxalide F (6): mp 180-181 ˚C; []25D +111.5 (c 0.085, CHCl3).
Tetraacetate 9 from briaroxalide G (7): mp 185-186 ˚C; []25D +149.2 (c 0.085, CHCl3).
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