Anti-Pseudomonas aeruginosa xanthones from the resin and green fruits of Cratoxylum cochinchinense

Article abstract of DOI:10.1016/j.tet.2009.01.083

Cochinchinones I-L (1-3 and 13) along with 11 known xanthones (4-12, 14, and 15) were isolated from the resin and green fruits of Cratoxylum cochinchinense. In addition, four new acetylated compounds (16-19) were derivatized from 7-geranyloxy-1,3-dihydrox

Full text of DOI:10.1016/j.tet.2009.01.083

Tetrahedron 65 (2009) 3003–3013
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Anti-Pseudomonas aeruginosa xanthones from the resin and green fruits
of Cratoxylum cochinchinense
,
,
b
,
a
b
Nawong Boonnak a b, Chatchanok Karalai a , Suchada Chantrapromma , Chanita Ponglimanont ,
*
Hoong-Kun Fun c, Akkharawit Kanjana-Opas d, Kan Chantrapromma e, Shigeru Kato f
a Crystal Materials Research Unit, Department of Chemistry, Faculty of Science, Prince of Songkla University, Hat-Yai, Songkhla 90112, Thailand
b Department of Chemistry and Center of Excellence for Innovation in Chemistry, Faculty of Science, Prince of Songkla University, Hat-Yai, Songkhla 90112, Thailand
c X-ray Crystallography Unit, School of Physics, Universiti Sains Malaysia, 11800 USM, Penang, Malaysia
d Department of Industrial Biotechnology, Faculty of Agro-Industry, Prince of Songkla University, Hat-Yai, Songkhla 90112, Thailand
e Institute of Research and Development, Walailak University, Thasala, Nakhon Si Thammarat, 80160, Thailand
f Department of Materials and Life Science, Seikei University, Tokyo 180 8633, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Cochinchinones I–L (1–3 and 13) along with 11 known xanthones (4–12, 14, and 15) were isolated from
the resin and green fruits of Cratoxylum cochinchinense. In addition, four new acetylated compounds (16–
19) were derivatized from 7-geranyloxy-1,3-dihydroxyxanthone (14) and 3-geranyloxy-1,7-dihydroxy-
xanthone (15). All compounds were characterized on the basis of spectroscopic analyses. The structures
of cochinchinone I (1), a monoacetate (18) and a dibrosylate (20), were also confirmed by X-ray dif-
fraction analysis. The antibacterial and antifungal activities of selected compounds were evaluated as
well.
Received 23 October 2008
Received in revised form 8 January 2009
Accepted 22 January 2009
Available online 29 January 2009
Keywords:
Cratoxylum cochinchinense
1,3,7-Trihydroxyxanthone
1,3,7-Trioxygenated xanthone
Antifungal activity
Ó 2009 Elsevier Ltd. All rights reserved.
Antibacterial activity
Pseudomonas aeruginosa
1. Introduction
investigate the bioactive compounds from this plant. The antibac-
terial and antifungal activities of isolated compounds were also
Cratoxylum cochinchinense belongs to the family Guttiferae,
which is distributed in several parts of Thailand.1 This plant is
a well-known tropical tree, and is commonly known in Thailand as
Tui Kliang. The bark, roots, and leaves of this plant have been used
in folk medicine to treat fever, coughs, diarrhea, itches, ulcers, and
abdominal complaints.2 Previous investigations revealed the major
components from the Cratoxylum genus as xanthones and anthra-
quinones.3–6 Xanthones have been reported to exhibit biological
activities such as antibacterial,3–5 antioxidant,6,7cytotoxic,3–5,8,9
and antiprotozoal activities.10 A preliminary screening of the bio-
activity of the crude extracts from the resin and green fruits of C.
cochinchinense has shown strong antibacterial activity specifically
against Pseudomonas aeruginosa. Although Pseudomonas sepsis is
a major cause of death in transplant recipients, while P. aeruginosa
is one of the most common Gram-negative pathogenic bacteria
causing life-threatening infections.11–13 This result led us to
evaluated.
2. Results and discussion
The crude CH2Cl2 extract of the resin of C. cochinchinense was
subjected to chemical investigation leading to the isolation of three
new xanthones, cochinchinones I–K (1–3), together with nine
known xanthones identified as cochinchinone A (4),6 1,3,7-tri-
hydroxy-2,4-diisoprenylxanthone (5),14 celebixanthone methyl
ether (6),15 dulxis-xanthone F (7),15 -mangostin (8),16
b a-mangostin
(9),16 macluraxanthone (10),17 pruniflorone G (11),3 and a caged-
prenylated xanthone (12).6 Their structures were elucidated by
NMR analysis and comparison of their spectroscopic data with
those reported in the literature.
Cochinchinone I (1) was obtained as a yellow powder, which
was further recrystallized from acetone to yield yellow single
crystals. The X-ray structure (Fig. 1) suggested a xanthone skeleton
with a chromene ring. A molecular formula C28H30O5 was implied
by HRMS. Its structure was supported by the 1H and 13C NMR
spectral data (Table 1). The 1H and 13C NMR spectral data of 1 were
* Corresponding author. Tel.: þ66 7428 8444; fax: þ66 7421 2918.
E-mail addresses: chatchanok.k@psu.ac.th, chatchanok_k@yahoo.com (C. Karalai).
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.01.083

3004
N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
chromene ring was connected to a xanthone skeleton in a linear
fashion whose structure was confirmed by HMBC correlations as
summarized in Table 2, which agreed well with the X-ray structure
(Fig. 1). From these data, the structure of cochinchinone I was
assigned as 1. In addition, the asymmetric unit of a cocrystal
(Fig. 1a) consisted of a disordered mixture of cochinchinone I (1)
(Fig. 1) and a new chromenoxanthone 1a (Fig. 1b). The latter was
different from 1 in which a geranyl side chain in 1 was replaced by
a (E)-300,700,700-trimethyloct-200-ene side chain in 1a. Therefore,
a trace amount of chromenoxanthone 1a in a single crystal should
be an unexpected contaminated compound from cochinchinone I
(1), which might be transformed during the resin formation.
Cochinchinone J (2) was obtained as a yellow viscous oil with
a molecular ion peak at m/z 446.2092 [M]þ in the HRMS, corre-
sponding to a molecular formula of C28H30O5. The UV spectrum
showed absorption bands of a xanthone (243, 289, 298, 320, 351, and
391 nm),14 while the IR spectrum exhibited the hydroxyl and conju-
gated carbonyl functionalities at vmax 3397 and 1649 cmꢀ1, re-
spectively. The 1H and 13C NMR data of 2 (Table 1) were similar to
thoseof 4,6 exceptfortheappearanceofthesignalsofachromenering
bearing a methyl group and six-carbon side chain of 4-methylpent-3-
enyl group, which appeared atdH 5.10 (br t, J 7.2 Hz, H-600),1.89 (m,1H-
400), 1.68 (m, 1H-400), 2.12 (m, H2-500), 1.66 (s, Me-800), and 1.57 (s,
Me-1000) (Table 1) instead of a geranyl moietyat C-4 as in 4. The loss of
4-methylpent-3-enyl moiety in EIMS, m/z 363 ([M]þ ꢀ83), also sup-
ported the proposed structure. Finally, the location of the angular
chromene ring was confirmed by HMBC correlations (Table 2) in
Figure 1. The X-ray structure of 1.
which the methine proton H-100
158.9), C-4 ( 100.2), C-4a (
150.2), and C-300 (
structure of cochinchinone J was deduced to be 2.
(d
6.88) was correlated with C-3 (
d
comparable to cochinchinone A (4), which was previously isolated
from the roots of C. cochinchinense.6 The main difference was ob-
served at C-2 and C-3, where the 1H NMR spectral data of 1 showed
d
d
d
80.6). Therefore, the
Compound 3 was isolated as an inseparable mixture with
compound 2, and the mixture was thus acetylated with Ac2O in
pyridine. The resulting product was further separated by CC eluting
with 70% CHCl3–hexane to give monoacetates 3a (18.9 mg) and 2a
the signals of a chromene ring at
d
6.74 (d, J 9.9 Hz, H-10), 5.60 (d, J
9.9 Hz, H-20), and 1.48 (s, Me-40 and Me-50) instead of a prenyl
group at C-2 and a free hydroxyl group at C-3 as in 4 (Table 1). The
Table 1
1H and 13C (300 MHz) spectral data of 1, 2, 3a, and 4 in CDCl3
Position
1
2
3a
4
1H (J in Hz)
13C (
d)
1H (J in Hz)
13C (
d
)
1H (J in Hz)
13C (
d
)
1H (J in Hz)
13C (
d)
1-OH
2
3
4
5
6
7
8
9
12.97, s
155.5
104.2
158.4
107.4
119.0
124.1
150.5
108.9
180.9
154.5
152.4
120.6
103.3
115.8
127.3
78.1
13.15, s
160.2
111.2
158.9
100.2
118.9
123.8
150.3
109.3
180.5
150.2
152.2
121.1
102.9
21.2
13.02, s
158.4
103.9
159.6
107.3
118.7
128.6
146.3
117.8
180.3
152.4
153.7
121.2
102.3
16.2
12.79, s
158.1
109.1
161.1
104.9
118.7
124.7
150.1
108.7
180.9
152.9
152.4
120.2
103.0
21.8
7.30, d, 9.0
7.23, dd, 2.4, 9.0
7.35, d, 8.7
7.25, m
7.46, d, 8.7
7.39, dd, 8.7, 2.7
7.04, d, 9.0
7.07, br d, 9.0
7.60, d, 2.4
7.60, br d, 1.8
7.92, d, 2.7
7.44, br s
4a
4b
8a
9a
10
6.74, d, 9.9
5.60, d, 9.9
3.36, d, 7.2
5.25, br t, 7.2
2.75, br t, 6.9
1.81, br t, 6.9
3.32, d, 6.9
5.19, br t, 6.9
20
122.1
131.5
25.8
31.7
76.3
26.9
26.9
121.6
134.8
25.6
30
40
1.48, s
1.48, s
28.4
28.4
1.68, s
1.81, s
1.39, s
1.39, s
1.55, s
1.75, s
50
17.9
17.9
100
200
300
400
500
600
700
800
900
1000
7-OAc
3.46, d, 7.2
5.22, br t, 7.2
21.3
6.88, d, 10.2
5.54, d, 10.2
115.9
125.4
80.6
41.8
22.8
123.8
131.9
25.6
27.1
17.6
3.48, d, 7.5
5.22, br t, 6.3
21.5
3.39, d, 6.9
5.16, br t, 6.9
21.6
122.1
135.0
39.7
122.3
134.9
39.8
121.7
137.9
39.7
1.99, ma
1.89, m,a 1.68, ma
2.12, ma
1.96, m
2.03, m
5.05, br t, 6.6
1.99, ma
2.04, ma
26.6
26.7
1.97, ma
26.4
5.04, br t, 6.6
124.2
131.3
25.6
16.3
17.6
5.10, br t, 7.2
124.4
131.3
25.6
16.3
17.7
4.95, br t, 7.2
123.9
131.8
25.8
16.2
17.6
1.59, s
1.86, s
1.53, s
1.66, s
1.44, s
1.57, s
1.60, s
1.86, s
1.55, s
2.34, s
1.67, s
1.78, s
1.48, s
20.9/169.4
a
Deduced from HMQC experiment.

N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
3005
Table 2
HMBC (300 MHz) spectral data of 1, 2, 3a, and 4 in CDCl3
Position
1
2
3a
4
1-OH
5
6
C-1, C-2, C-9a
C-1, C-2, C-9a
C-7, C8a
C-7, C-4b
C-1, C-2, C-9a
C-7, C-8a, C-9
C-5, C-7, C-4b
C-1, C-2, C-9, C-9a
C-7, C-9, C-4b, C8a
C-7, C-8
C-7, C-9, C-4b, C8a
C-5, C-7, C-8, C-4b
C-6, C-7, C-9, C-4b
C-1, C-2, C-3, C-4, C-30
C-2, C-30, C-40, C-50
C-20, C-30, C-50
8
C-8
C-6, C-7, C-4b, C-9
C-1, C-2, C-3, C-20, C-30
C-2, C-10, C-30, C-40, C-50
C-20, C-30
C-6, C-7, C-9, C-4b
10
C-1, C-2, C-20, C-30
C-20
C-1, C-2, C-3, C-20, C-30 C-40, C-50
C-2, C-10, C-50
20
40
C-20, C-30, C-50
C-20, C-30, C-40
C-3, C-4, C-4a, C-300
C-4, C-300, C-400, C-500, C-900
C-200, C-300
C-20, C-30, C-50
C-20, C-30, C-40
50
C-20, C-30, C-40
C-20, C-30
100
C-3, C-4, C-4a, C-200, C-300
C-4, C-100, C-400, C-900
C-200, C-300, C-500
C-3, C-4, C-4a, C-200, C-300
C-100, C-300, C-400, C-900
C-200, C-300, C-600
C-300, C-600, C-700
C-500, C-800, C-1000
C-600, C-700, C-1000
C-200,C-300
C-3, C-4, C-4a, C-200, C-300, C-900
C-4, C-100, C-400, C-900
C-200, C-600
200
400
500
C-300, C-400, C-600, C-700
C-400, C-500, C-800, C-1000
C-600, C-700, C-1000
C-200, C-300
C-600, C-700
C-600
C-600, C-700, C-1000
C-200, C-300, C-400
C-600, C-700, C-800
C-300, C-400, C-600, C-700
C-400, C-500, C-800
600
800
C-600, C-700, C-1000
900
C-200, C-300, C-400
1000
7-OAc
C-600, C-700, C-800
C-600, C-700, C-1000
C-7
C-600, C-700, C-800
(5.1 mg). The latter was confirmed as an acetylated derivative of 2
by comparison of its spectral data with those of compound 2.
Compound 3a is a yellow powder. The HRMS spectrum showed
a molecular ion peak at m/z 490.2355 [M]þ, corresponding to
C30H34O6. The UV spectrum showed absorption bands of a xan-
thone (243, 271, 303, 326, and 383 nm),14 while the IR spectrum
exhibited the hydroxyl and conjugated carbonyl functionalities at
vmax 3392 and 1648 cmꢀ1, respectively. The 1H and 13C NMR spec-
tral data of 3a (Table 1) were closely related to those of 1, except for
the appearance of the signals of a dimethylchromane ring and
acetoxy group revealed at dH 2.75 (br t, J 6.9 Hz, H-10), 1.81 (br t, J
6.9 Hz, H-20), 1.39 (s, Me-40 and Me-50), and 2.34 (s, 7-OAc) (Table 1)
instead of a chromene ring and a hydroxyl group in 1. The chromane
ring was fused to the xanthone nucleus in a linear fashion, which
was confirmed by HMBC correlations (Table 2), in which the
with C-1 (d 158.4), C-2 (d 103.9), and C-9a (d 102.3). Therefore, the
structure of 3a, an acetylated form of 3, was deduced. Compound 3
was named cochinchinone K.
It is interesting to note that the three newchromenoxanthones (1
and 2) and chromanoxanthone (3) could be derived from cochin-
chinone A (4). The isoprenyl or geranyl side chains of 4 were epox-
idized and further cyclized via a free hydroxyl at C-3 to produce the
chromanol ring18 as shown in Scheme 1. Further dehydration led to
the linear or angular chromene rings of 1 and 2, respectively. On the
other hand, the new chromanoxanthone (3) could be produced by
cyclization via a free hydroxyl groupto an isoprenyl side chain of 4 to
afford thechromane ring (Scheme 1). Therefore, cochinchinone A (4)
should be a precursor of cochinchinones I–K (1–3).
The investigation of the crude CH2Cl2 extract of the green fruits
(approximate 4 weeks maturity stage) led to the isolation and
identification of a new xanthone, cochinchinone L (13) along with
two major known oxygeranyl xanthones 1414 and 15.19 Compounds
14 and 15 were structural isomers of xanthones with an oxygeranyl
methylene protons H2-10 at dH 2.75 were correlated with C-1 (
158.4), C-2 ( 103.9), C-3 ( 76.3),
159.6), C-20 ( 31.7), and C-30 (
while the H-bonded hydroxyl proton 1-OH ( 13.02) was correlated
d
d
d
d
d
d
Figure 1a. The asymmetric unit of a cocrystal of 1 and 1a.
Figure 1b. The X-ray structure of 1a.

3006
N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
O
9
OH
1
O
9
OH
1
8
1'
1'
8
3'
HO
HO
2'
3'
cyclization
6
3
6
O
O
3
O
OH
1''
chromanoxanthone (3)
4
3''
5''
epoxidation and
cyclization
epoxidation and
cyclization
7''
O
8
OH
1'
OH
1
1
O
HO
9
OH
8
HO
9
6
3
3'
O
O
6
O
3 O
3''
7''
1''
5''
OH
chromanolxanthone intermediate
chromanolxanthone intermediate
dehydration
-H2O
dehydration
-H2O
OH
1
O
9
8
OH
1
O
9
HO
8
1'
HO
2'
3'
6
3
O
O
6
3''
7''
3
O
O
1''
5''
2''
chromenoxanthone (2)
chromenoxanthone (1)
Scheme 1. Plausible biosynthesis pathway of 1, 2, and 3.
side chain. The 1H and 13C NMR spectral data of both 14 and 15
6.40) and H-2 (
d
6.34). These results implied that the oxygeranyl
(Tables 3 and 4) were almost identical. However, the main differ-
side chain of 14 was connected to C-7 and that of 15 to C-3. To
confirm these structural assignments of 14 and 15, compound 14
was further reacted with p-bromobenzenesulfonyl chloride in
CH2Cl2 at room temperature to afford a crystalline dibrosylate 20
ence was observed in the NOESYexperiment in which H2-10 (
d
4.45)
of 14 showed a cross-peak with H-8 (d 7.40) and H-6 (d 7.18),
whereas the H2-10 (
d 4.63) of 15 showed a cross-peak with H-4 (d
Table 3
1H NMR (300 MHz) spectral data of 13–19 in CDCl3
Position
13
14
15
16
17
18
19
2
4
5
6
6.43, d, 2.1
6.57, d, 2.1
7.08, br d, 9.0
7.10, br d, 9.0
7.45, br s
4.42, d, 6.3
5.73, t, 6.3
1.99, m
1.97, m
4.98, t, 6.6
1.57, s
6.22, d, 1.8
6.23, d, 1.8
7.15, br d, 9.0
7.18, br d, 9.0
7.40, br s
4.45, d, 6.3
5.39, br t, 6.3
2.01, m
1.95, m
4.98, br t, 5.7
1.57, s
6.34, d, 2.4
6.40, d, 2.4
7.30, d, 9.3
7.26, d, 9.3
7.59, br s
4.63, d, 6.6
5.50, br t, 6.6
2.13, m
2.10, m
5.11, br t, 5.7
1.69, s
1.78, s
1.62, s
d
6.42, br d, 2.1
6.59, br d, 2.1
7.21, br d, 8.1
7.20, br d, 8.1
7.44, br s
4.50, d, 6.6
5.40, br t, 6.6
2.03, m
2.02, m
5.00, br t, 6.3
1.57, s
7.15, d, 2.1
6.79, d, 2.1
7.23, d, 9.3
7.28, dd, 9.3, 2.1
7.55, d, 2.1
4.56, d, 6.3
5.48, br t, 6.3
2.11, m
2.09, m
5.09, br t, 6.3
1.66, s
1.73, s
1.59, s
d
6.19, br d, 2.1
6.24, br d, 2.1
7.25, d, 9.3
7.29, br d, 9.3
7.76, br s
4.49, d, 6.3
5.37, br t, 6.3
2.03, m
1.97, m
4.99, br t, 6.9
1.57, s
6.41, d, 2.1
6.46, d, 2.1
7.25, d, 8.1
7.26, br d, 8.1
7.78, br d, 1.5
4.51, d, 6.3
5.37, br t, 6.3
2.03, m
2.01, m
4.99, br t, 6.6
1.57, s
8
10
20
40
50
60
80
90
1.60, s
1.50, s
2.38, s
d
1.64, s
1.50, s
d
1.68, s
1.51, s
d
1.66, s
1.51, s
d
1.65, s
1.51, s
d
100
1-OAc
1-OH
3-OH
7-OH
1-OAc
3-OAc
7-OAc
12.85, s
7.86, br s
d
12.72, s
12.71, s
d
d
12.55, s
d
d
d
d
d
d
7.03, br s
d
d
d
d
d
d
d
d
d
d
2.47, s
2.27, s
d
d
2.40, s
d
d
d
2.23, s
d
d
d
d
2.22, s
2.19, s

N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
3007
(Fig. 3), while compound 15 was acetylated in the usual manner
with Ac2O in pyridine to give a crystalline monoacetate 18 (Fig. 2).
Both 20 and 18 were subjected to X-ray diffraction analysis whose
results supported the structures of 14 and 15.
Cochinchinone L (13) was isolated as a yellow powder, with
a molecular ion peak at m/z 422.1718 [M]þ in the HRMS, corre-
sponding to a molecular formula of C25H26O6. The 1H and 13C NMR
spectral data of 13 (Tables 3 and 4) showed characteristic signals
similar to those of 7-geranyloxy-1,3-dihydroxyxanthone (14),14
except that the hydroxyl group at C-1 of 14 was replaced by an
acetoxy group in 13, which appeared as a proton singlet signal at
2.38. The higher field chemical shift of C-1 ( 151.2) and lower
d
d
field resonances of protons and carbons of C-2 (dH 6.43 (d, J 2.1 Hz,
H-2); dC 108.3) and C-4 (dH 6.57 (d, J 2.1 Hz, H-4); dC 101.3) as
compared to those of 14 (dH 6.22 (d, J 1.8 Hz, H-2); dC 98.5 and dH
6.23 (d, J 1.8 Hz, H-4); dC 94.4) supported the title structure (Tables
3 and 4). The 4J HMBC correlations of methyl protons of the ace-
toxy group (d 2.38) with C-1 (d 151.2) supported its location at C-1,
which was also confirmed by NOESY cross-peak between acetoxy
moiety with H-2. Cochinchinone L (13), an acetylated analogue of
5'
O
9
OH
1
OH
1
O
9
OH
1
O
9
8
1'
8
1'
8
1'
3'
9a
8a
4b
8a
4b
9a
8a
4b
9a
4a
HO
HO
O
2'
2'
R
4'
10''
2'
4'
4'
5'
6
6
6
3
3'
O 4a
1''
O
3
3'
3
O 4a
1''
O
O
O
5'
3''
7''
1''
5''
8''
9''
1
2 : R = H
2a : R = Ac
1a
3''
5''
3''
5''
9''
9''
7''
7''
8''
8''
10''
10''
11''
5'
OH
1
O
O
9
OH
O
OH
8
8
1'
1'
3'
8a
9a
4a
1
8a
9a
HO
O
HO
2'
9
R
4'
2'
4'
6
6
3
4b O
3 O
4b O 4a
1''
OH
O
OH
3'
5'
1''
3 : R = H
5
4
3''
5''
3''
5''
9''
3a : R = Ac
7''
7''
8''
8''
10''
10''
OH
O
O
OH
O
OH
O
R2
HO
O
H3CO
H3CO
O
OCH3
R1
HO
O
OH
8: R1 = OCH3
9: R1 = OH
R2 = OCH3
R2 = OCH3
6
7
OR1
1
O
O
OH
O
9
OR1
1
O
8
8
R2O
8'
7'
3'
9'
O
10'
7'
9'
3'
9
1'
R1
1'
5'
B
A
OR2
6
6
HO
O
10'
O
3
3
O
O
5'
8'
OH
13: R1 = Ac
14: R1 = H
16: R1 = H
17: R1 = Ac
15: R1 = H R2 = H
18: R1 = H R2 = Ac
19: R1 = Ac R2 = Ac
R2 = H
R2 = H
R2 = Ac
R2 = Ac
10: R1 = CH3
11: R1=
3''
Br
O
1''
O
OH
O
O
5''
S
O
9
OH
1
O
9
O
3'''
H3CO
O
Br
8
8
1
8'
HO
3'
9'
O
O
1'''
7'
10'
1'
B
5'
A
5'''
S
6
6
3 OH
O
O
3 O
O
O
20
21
12

3008
N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
Table 4
activity against all Gram-positive bacteria. Nearly all compounds,
13C NMR (75 MHz) spectroscopic data of 13–19 in CDCl3
which are active against P. aeruginosa are the 1,3,7-trihydroxy
xanthones (4 and 5) or 1,3,7-trioxygenated xanthones with an
oxygeranyl side chain either at C-3 or C-7 and dihydroxyl groups
(14–15) whose indicated structures might contribute to the strong
antibacterial activity specifically against P. aeruginosa. However,
when the free hydroxyl group of the 1,3,7-trihydroxy xanthone was
cyclized onto the isoprenyl or geranyl side chain to form a chro-
mane or chromene ring, the antibacterial activity against P. aeru-
ginosa decreased drastically as shown in compounds 1, 2, and 3a. In
other previous reports, it was suggested that hydroxy xanthones
Position
13
14
15
16
17
18
19
1
2
3
4
5
6
7
8
151.2
108.3
162.6
101.3
118.8
125.1
155.4
106.4
175.7
158.8
150.0
122.2
107.8
65.51
118.8
141.9
39.5
26.3
123.8
131.8
25.7
16.7
163.8
98.5
164.0
94.4
118.9
125.5
155.2
105.9
180.5
157.8
150.7
120.3
103.3
65.6
118.6
142.1
39.5
26.3
123.8
131.8
26.3
16.7
17.7
163.2
97.6
166.1
93.2
188.9
124.2
152.5
109.0
180.5
157.8
150.5
120.9
103.5
65.6
118.3
142.3
39.5
26.2
123.6
131.9
25.6
16.7
17.7
162.8
104.0
156.8a
100.7
119.0
126.0
155.4
106.0
181.1
156.7a
150.8
120.7
106.6
65.6
118.8
141.9
39.5
26.3
123.7
131.8
25.6
16.7
150.9
108.7
154.5
112.4
118.9
125.2
155.5
106.6
174.7
157.7
149.9
122.3
112.2
65.5
163.3
97.8
158.7
99.4
166.2
93.4
118.7
128.9
146.5
117.8
179.8
157.5
153.3
121.1
103.4
65.6
118.4
142.2
39.5
26.2
123.9
131.9
25.6
16.7
17.7
163.6
108.1
118.6
128.3
146.7
118.4
174.1
151.4
152.7
123.6
108.6
65.8
9
4a
4b
8a
9a
10
play important roles in biological activity such as antibacterial,3,5
a-
glucosidase inhibitory,20 anti-tumor,21,22 and anti-inflammatory
effects.23 Because compounds 14 and 15 are the major components
obtained from this plant, this prompts us to modify their structures
for the structure–activity relationships (SARs). To investigate
whether the free hydroxyl group was responsible for antibacterial
activity, the acetylation with acetic anhydride in pyridine was
therefore applied to 14 and 15. Four acetylated geranyloxyxanthone
derivatives: 3-acetoxy-7-geranyloxy-1-hydroxyxanthone (16) and
1,3-diacetoxy-7-geranyloxyxanthone (17) were obtained from 14,
whereas 7-acetoxy-3-geranyloxy-1-hydroxy-xanthone (18) and
1,7-diacetoxy-3-geranyloxy-xanthone (19) were obtained from 15.
The assignments of the 1H and 13C NMR spectral data of 16–19 are
summarized in Tables 3 and 4. The antibacterial activity of acety-
lated geranyloxyxanthone derivatives 16–19 and dibrosylate 20
was evaluated. All of them showed strong anti-P. aeruginosa (Table
5). This implied that the free hydroxyl groups should not be
responsible for the inhibition of P. aeruginosa. For further in-
vestigation of the role of an oxygeranyl side chain, a 1,3,7-trihy-
droxyxanthone (21) obtained from the dried fruits of C.
cochinchinense24 was tested against P. aeruginosa. It was inactive
against Gram-negative bacteria as shown in Table 5. From these
results, it can be suggested that the geranyl side chain is necessary
for anti-P. aeruginosa activity. Moreover, mixtures of compound 4
with compound 5, and compound 14 with compound 15 were
subjected to antimicrobial assay. Interestingly, the mixture of
compounds 14 and 15 significantly increased the antibacterial ac-
tivity against MRSA compared with the pure forms as indicated by
the lower MIC values shown in Table 5. The mixture of compounds
4 and 5, on the other hand, did not show any significant differences
for antibacterial activity compared to the pure forms as also shown
in Table 5. Therefore, it may be proposed that the 1,3,7-trihydroxy-
xanthone with the isoprenyl or geranyl side chain at C-2 and C-4 in
20
118.8
141.5
39.5
118.0
142.6
39.5
30
40
50
26.3
26.2
60
123.8
131.7
25.6
16.7
17.6
21.1
169.2
21.0
167.8
d
123.6
131.9
25.6
16.7
17.7
21.1
169.5
d
70
80
90
100
1-OAc
17.7
171.0
21.3
17.7
d
d
d
d
d
d
d
d
3-OAc
7-OAc
d
d
d
21.1
168.2
d
d
d
d
d
d
d
d
d
d
20.9
169.2
20.9
169.2
d
d
d
d
d
a
May be interchangeable.
14, was therefore assigned as 1-acetoxy-3-hydroxy-7-geranyl-
oxyxanthone.
The isolated compounds were evaluated for their antibacterial
activities against both Gram-positive bacteria: Bacillus subtilis,
Staphylococcus aureus, Enterococcus faecalis TISTR 459, Methicillin-
Resistant S. aureus (MRSA) ATCC 43300, Vancomycin-Resistant
E. faecalis (VRE) ATCC 51299 and Gram-negative bacteria: Salmo-
nella typhi, Shigella sonei, and P. aeruginosa. All compounds were
also subjected to antifungal assay against Candida albicans. The
results showed that most of the isolated compounds (4–6 and 13–
15) exhibited strong antibacterial activity specifically against P.
aeruginosa (Table 5). Interestingly, only compounds 9 and 10
exhibited strong activity against C. albicans. It is important to note
that compound 9 also exhibited broad spectrum antimicrobial
Figure 2. ORTEP plot of a monoacetate 18.

N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
3009
Figure 3. ORTEP plot of a dibrosylate 20.
4 and 5, respectively, and 1,3,7-trioxygenated xanthone with the
geranyl side chain at C-3 or C-7 in 13–19 are essential for their
antibacterial activity against P. aeruginosa. Therefore, 1,3,7-trihy-
droxyxanthones (4 and 5) and 1,3,7-trioxygenated xanthone with
geranyl side chain (13–19) should be considered as potent candi-
dates as anti-P. aeruginosa.
results was correlated to their strong antibacterial activity (Table 5).
Therefore, it can be suggested that compounds 4 and 13 may interact
with or damage the cell wall of P. aeruginosa as seen by the formation
of pores on the cell wall of P. aeruginosa (Figs 4 and 5).
3. Experimental
We further studied the possible mode of action of compounds 4
and 13 against P. aeruginosa by observing the bacteria cell morpho-
logy through scanning electron microscopy (SEM) at 3, 6, 9, and 15 h
after applying compounds 4 and 13. From the SEM results (see Figs 4
and 5), it was clearly indicated that the cell morphology of P. aeru-
ginosa, when treated with compounds 4 and 13, started to deform at
3 h onward, and at 15 h, most cells were completely deformed whose
3.1. General experimental procedures
Melting points were determined on a Fisher–John melting point
apparatus. Optical rotations were measured on a JASCO P-1020
digital polarimeter. UV and IR spectra were recorded on SPECORD S
100 (Analytikjena) and Perkin–Elmer FTS FT-IR spectrophotometer,
Table 5
Antimicrobial activity (MIC, mg/ml) of 1–21
No.
Antibacterial activity
Antifungal activity
b
Gram-positive bacteriaa
Gram-negative bacteria
B. subtilis
S. aureus
E. faecalis
MRSA
VRE
S. typhi
S. sonei
P. aeruginosa
C. albicansc
1
2
3a
4
5
4D5
6
7
8
9
10
11
12
13
14
15
14D15
16
17
18
19
20
21
>300
300
75
150
150
75
>150
150
300
9.37
18.75
>300
75
150
150
150
75
>300
300
>300
150
75
150
>150
300
75
9.37
37.5
300
>300
>150
>150
150
37.5
>150
>150
>150
>150
>150
75
>300
>300
300
150
150
75
>150
300
150
9.37
37.5
>300
300
150
150
150
37.5
150
75
150
>150
150
>300
300
>300
9.37
37.5
9.37
150
150
75
9.37
37.5
150
>300
300
>300
150
75
>300
>300
>300
>150
>150
>150
>150
>300
>300
>300
>300
>300
>300
>150
>150
>150
>150
>150
>150
>150
>150
>150
>300
>300
>300
>300
>150
>150
>150
>150
>300
>300
>300
>300
>300
>300
>150
>150
>150
>150
>150
>150
>150
>150
>150
>300
>300
300
>300
4.7
4.7
4.7
>300
75
300
75
37.5
75
150
300
150
2.4
150
150
150
150
9.37
37.5
150
150
150
150
150
37.5
150
150
150
150
>300
300
4.7
150
300
18.75
37.5
>300
>300
4.7
4.7
4.7
4.7
4.7
4.7
300
>300
18.75
75
37.5
37.5
150
18.75
150
150
>300
300
150
37.5
18.7
9.3
4.7
150
75
>150
>150
>150
37.5
18.75
37.5
18.75
37.5
300
150
4.7
4.7
4.7
4.7
>300
300
a
B. subtilis, S. aureus, E. faecalis TISTR 459, Methicillin-Resistant S. aureus (MRSA) ATCC 43300, Vancomycin-Resistant E. faecalis (VRE) ATCC 51299.
S. typhi, S. sonei, and P. aeruginosa.
C. albicans.
b
c

3010
N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
Figure 4. Scanning electron microscopy of cell morphology of P. aeruginosa treated with compound 4 at different time.
respectively. The 1H and 13C NMR spectra were recorded on
300 MHz Bruker FTNMR Ultra ShieldÔ spectrometer in CDCl3 with
TMS as the internal standard. Chemical shifts are reported in
subfractions (FR4A–FR4H) and 8 (150.2 mg) (Rf (15% acetone–hex-
ane (three runs)) 0.39). Subfraction FR4B was further purified by CC
and eluted with a gradient of acetone–hexane to give 7 (31.4 mg)
(Rf (15% acetone–hexane) 0.31) and 12 (56.5 mg) (Rf (15% acetone–
hexane) 0.28). Fractions FR6 and FR7 were separated by QCC and
eluted with a gradient of CH2Cl2–hexane to give six subfractions
(FR6A–FR6F). Subfraction FR6B was further separated by QCC
eluting with a gradient of acetone–hexane to give 6 (35.7 mg)
(Rf (15% acetone–hexane (three runs)) 0.41), 8 (83.2 mg) (Rf (15%
acetone–hexane (three runs)) 0.39), and 11 (1.8 mg) (Rf (15% ace-
tone–hexane (three runs)) 0.24). Subfraction FR6E was purified by
CC on reversed-phase silica gel C-18 eluting with MeOH to give 4
(849.4 mg) (Rf (MeOH) 0.54) and 5 (1.25 g) (Rf (MeOH) 0.62).
Fractions FR8 and FR9 were separated by QCC and eluted with
a gradient of CH2Cl2–hexane to afford 5 (551.3 mg) (Rf (60% CH2Cl2–
hexane (three runs)) 0.05), 7 (18.2 mg) (Rf (60% CH2Cl2–hexane
(three runs)) 0.29), 8 (116.6 mg) (Rf (60% CH2Cl2–hexane (three
runs)) 0.61), and 9 (148.7 mg) (Rf (60% CH2Cl2–hexane (three runs))
0.10). Fractions FR10 and FR11 were separated by QCC and eluted
with a gradient of acetone–hexane to give seven subfractions
(FR10A–FR10G) and 7 (25.9 mg) (Rf (15% acetone–hexane) 0.31).
Subfraction FR10B was further purified by CC on silica gel C-18 and
eluted with MeOH to furnish 1 (8.5 mg) (Rf (MeOH) 0.36). The
mixture of 1 and 1a could not be separated due to the very low
amount of 1a. The purity of 1 is much more than 90% base on
a single spot on TLC and the integral area signal in the 1H NMR
spectrum of 1. Subfraction FR10D was separated by QCC eluting
d
(ppm) and coupling constants (J) are expressed in hertz. EI and
HREI mass spectra were measured on a Kratos MS 25 RFA
spectrometer. Quick column chromatography (QCC) and column
chromatography (CC) were carried out on silica gel 60 F254 (Merck)
and silica gel 100 (Merck), respectively. All the bacteria images
were viewed with a JSM-5800LV, JEOL SEM (scanning electron
microscope).
3.2. Plant material
The resin of C. cochinchinense was collected in October 2003 at
Prince of Songkla University, Hat-Yai campus, whereas the green
fruits of C. cochinchinense were collected in October 2007 at Kaun
Kha Long district, Satun Province, Southern part of Thailand. Bo-
tanical identification was achieved through comparison with
a voucher specimen No. SL-1 (PSU) in the herbarium of Department
of Biology, Prince of Songkla University, Songkhla, Thailand.
3.3. Isolation and extraction
The resin of C. cochinchinense (87.75 g) was extracted with
CH2Cl2 (2ꢁ2.0 L, for a week) at room temperature and was evapo-
rated under reduced pressure to afford a deep green crude CH2Cl2
extract (47.04 g), which was subjected to QCC (Quick column
chromatography) on silica gel (Merck 60 F254) using hexane as
a first eluent and then increasing the polarity with acetone to give
16 fractions (FR1–FR16). Fractions FR4 and FR5 were separated by
CC eluting with a gradient of acetone–hexane to give eight
with
a gradient of acetone–hexane to give six subfractions
(FR10D1–FR10D6). Subfraction FR10D2 was further separated by CC
and eluted with a gradient of acetone–hexane to give five sub-
fractions (FR10D2A–FR10D2E), 2 (1.8 mg) (Rf (20% acetone–hexane)

N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
3011
Figure 5. Scanning electron microscopy of cell morphology of P. aeruginosa treated with compound 13 at different time.
0.36), 4 (23.1 mg) (Rf (20% acetone–hexane) 0.22), 5 (34.2 mg) (Rf
(20% acetone–hexane) 0.17), and an inseparable mixture of 2 and 3
(20.2 mg) (Rf (20% acetone–hexane) 0.35). The mixture was sepa-
rated by acetylation with Ac2O (0.1 ml) in pyridine (2.0 ml) and
stirred overnight at room temperature to give a yellow gum, which
was further purified by CC eluting with 70% CHCl3–hexane to give
acetylated derivatives 2a (5.1 mg) (Rf (70% CHCl3–hexane) 0.68) and
3a (18.9 mg) (Rf (70% CHCl3-hexane) 0.40), respectively. Subfraction
FR10D2E was purified by CC and eluted with 80% CH2Cl2–hexane to
give 8 (16.7 mg) (Rf (80% CH2Cl2–hexane) 0.91), 10 (5.0 mg) (Rf (80%
CH2Cl2–hexane) 0.85), and 11 (1.5 mg) (Rf (80% CH2Cl2–hexane)
0.38).
Air-dried green fruits of C. cochinchinense (5.5 kg) were
extracted with CH2Cl2 (2ꢁ20 L, for a week) at room temperature
and was evaporated under reduced pressure to afford a deep
green crude CH2Cl2 extract (40.04 g), which was subjected to QCC
on silica gel using hexane as a first eluent and increasing polarity
with EtOAc to give nine fractions (FS1–FS9). Fraction FS5 was
purified by CC eluting with pure CHCl3 to give 14 (1.88 g) (Rf (70%
CHCl3–hexane) (three runs) 0.46). Fraction FS6 was further sepa-
rated by CC eluting with pure CHCl3 to furnish six subfractions
(FS6A–FS6F), 14 (2.10 g) (Rf (70% CHCl3–hexane) (three runs) 0.46)
and 15 (490.2 mg) (Rf (70% CHCl3–hexane) (three runs) 0.39).
Subfraction FS6B was further purified by CC eluting with a gradi-
ent of acetone–hexane to give 13 (53.3 mg) (Rf (15% acetone–
hexane (three runs)) 0.17).
(73), 363 (76), 323 (26), 307 (15), 295 (13),137 (5), 69 (6). For 1H and
13C NMR spectroscopic data, see Table 1.
3.3.2. Cochinchinone J (2)
25
Yellow viscous oil; [
(log
a]
ꢀ69.8 (c 0.08, CHCl3); UV (CHCl3) lmax
D
3
) 243 (3.90), 289 (4.10), 298 (4.13), 320 (3.68), 351 (3.47), 391
(3.33) nm; IR (neat) nmax 3397, 1649, 1613 cmꢀ1; HRMS m/z
446.2092 for C28H30O5 (calcd 446.2093). EIMS m/z (rel int.): 446
[M]þ (11), 363 (100), 307 (18), 69 (5). For 1H and 13C NMR spec-
troscopic data, see Table 1.
3.3.3. Acetylated derivative of cochinchinone K (3a)
Yellow powder, mp 85–87 ꢂC; UV (CHCl3) lmax (log
3
) 243, 271,
303, 326, 383 nm; IR (neat) nmax 3392, 1709, 1648, 1612 cmꢀ1
;
HRMS m/z 490.2355 for C30H34O6 (calcd 490.2355). EIMS m/z (rel
int.): 490 [M]þ (53), 473 (53), 419 (44), 405 (31), 365 (100), 323 (52),
311 (41), 267 (51), 69 (24). For 1H and 13C NMR spectroscopic data,
see Table 1.
3.3.4. Cochinchinone L (13)
Yellow powder, mp 114–116 ꢂC; UV (CHCl3) lmax (log
3) 248
(4.59), 273 (4.03), 305 (4.13), 354 (3.80) nm; IR (neat) nmax 3237,
1774, 1728, 1628 cmꢀ1; HRMS m/z 422.1718 for C25H26O6 (calcd
422.1729). EIMS m/z (rel int.): 422 [M]þ (1), 286 (40), 244 (100), 187
(4), 81 (9), 69 (28). For 1H and 13C NMR spectroscopic data, see
Tables 3 and 4.
3.3.1. Cochinchinone I (1)
3.4. Acetylation of 14 and 15
Yellow needle single crystals, mp 160–162 ꢂC; UV (CHCl3) lmax
(log
3
) 261 (4.01), 297 (4.31), 342 (3.69), 393 (3.46) nm; IR (neat)
Compound 14 (82.5 mg) was treated with Ac2O (2.5 ml) in
pyridine (2.0 ml) and stirred for 6 h at room temperature. The re-
action mixture was diluted with water and extracted with CH2Cl2.
nmax 3406, 1650, 1612 cmꢀ1; HRMS m/z 446.2279 for C28H30O5
(calcd 446.2093). EIMS m/z (rel int.): 446 [M]þ (76), 431 (100), 377

3012
N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
The combined organic extract was washed with 10% HCl and then
washed with water again. After the organic solvent was removed,
the resulting residue was dried over anhydrous Na2SO4. Chroma-
tography over silica gel yielded a pale yellow powder of 16
(80.6 mg) (Rf (5% acetone–hexane (three runs)) 0.44).
Compound 14 (200.5 mg) was treated with Ac2O (6.0 ml) in
pyridine (3.0 ml) and stirred overnight at room temperature.
Chromatography over silica gel yielded a pale yellow powder of 16
(10.6 mg) (Rf (5% acetone–hexane (three runs)) 0.44) and 17
(177.8 mg) (Rf (15% acetone–hexane (three runs)) 0.46).
Compound 15 (85.5 mg) was treated with Ac2O (2.5 ml) in
pyridine (2.0 ml) and stirred for 6 h at room temperature. Chro-
matography over silica gel yielded a pale yellow powder of 18
(83.7 mg) (Rf (5% acetone–hexane (three runs)) 0.32), which was
further recrystallized from MeOH–acetone (1:99 v/v) to give yellow
single crystals.
Compound 15 (190.0 mg) was treated with Ac2O (6.0 ml) in
pyridine (3.0 ml) and stirred overnight at room temperature.
Chromatography over silica gel yielded a pale yellow powder of 18
(8.7 mg) (Rf (5% acetone–hexane (three runs)) 0.32) and 19
(170.0 mg) (Rf (15% acetone–hexane (three runs)) 0.37).
(three runs)) 0.20). Yellow needle-shaped single crystals of 20 were
obtained after recrystallization from CH3OH–CHCl3 (1:4 v/v), mp
106–108 ꢂC. dH (300 MHz, CDCl3þCD3OD) 7.82 (dd, J 9.0, 2.1 Hz,
H-200, H-600), 7.68 (d, J 2.4 Hz, H-4), 7.65 (dd, J 8.7, 2.1 Hz, H-300, H-500,
H-2000, H-6000), 7.63 (dd, J 8.7, 2.1 Hz, H-3000, H-5000), 7.50 (d, J 2.4 Hz,
H-8), 7.32 (d, J 2.4 Hz, H-2), 7.26 (d, J 9.0 Hz, H-5), 7.19 (dd, J 9.3,
2.4 Hz, H-6), 5.44 (br t, J 6.6, H-20), 5.04 (br t, J 6.3 Hz, H-60), 4.56 (d, J
6.6 Hz, H-10), 2.07 (m, 2H-50), 2.05 (m, 2H-40), 1.71 (s, 3H-90),1.60 (s,
3H-80),1.54 (s, 3H-100); dC (75 MHz, CDCl3þCD3OD) 181.6 (C-9),157.4
(C-1),155.9 (C-7),151.9 (C-3),149.7 (C-4b),148.6 (C-4a),142.1 (C-30),
134.3 (C-100), 133.4 (C-1000), 133.1 (C-300, C-500), 132.5 (C-3000, C-5000),
131.8 (C-70), 130.6 (C-400), 130.2 (C-200, C-600), 130.1 (C-4000), 129.8 (C-
2
000, C-6000),124.9 (C-6),123.7 (C-60),122.4 (C-8a),118.8 (C-5),118.6 (C-
20), 114.2 (C-9a), 112.8 (C-4), 111.1 (C-2), 106.8 (C-8), 65.6 (C-10), 39.5
(C-40), 26.2 (C-50), 25.5 (C-80), 17.5 (C-100), 16.6 (C-90). EIMS m/z (rel
int.): 816 [Mꢀ2]þ (1), 683 (7), 681 (14), 679 (7), 619 (5), 617 (10), 615
(5), 461 (55), 463 (56), 399 (41), 397 (41), 371 (19), 369 (19), 357 (34),
355 (34), 244 (27), 229 (84), 215 (100),186 (16),157 (53),155 (53),131
(11), 108 (13), 76 (16).
3.4.1. 3-Acetoxy-7-geranyloxy-1-hydroxyxanthone (16)
3.6. X-ray crystallographic studies of cocrystal of 1 and 1a,
monoacetate 18, and dibrosylate 20
Yellow powder, mp 94–95 ꢂC; UV (CHCl3) lmax (log
3) 239 (4.40),
262 (4.66), 289 (3.97), 379 (3.93) nm; IR (neat) nmax 3429, 1768,
1649, 1612 cmꢀ1
;
HRMS m/z 422.1725 for C25H26O6 (calcd
Crystallographic data were collected at 100.0 (1) K with the
Oxford Cryosystem Cobra low-temperature attachment. The data
were collected using a Bruker Apex2 CCD diffractometer with a
422.1729). EIMS m/z (rel int.): 422 [M]þ (1), 286 (43), 244 (100), 187
(4), 81 (9), 69 (24). For 1H and 13C NMR spectroscopic data, see
Tables 3 and 4.
graphite monochromated Mo Ka radiation at a detector distance of
5 cm using APEX2.25 The collected data were reduced using SAINT
program,25 and the empirical absorption corrections were per-
formed using SADABS program.25 The structures were solved by
direct methods and refined by least-squares using the SHELXTL
software package26 All non-hydrogen atoms were refined aniso-
tropically, whereas all H atoms were placed in calculated positions
with an O–H distance of 0.82 Å and C–H distances in the range
0.93–0.98 Å after checking their positions in the difference map.
The Uiso values were constrained to be 1.5Ueq of the carrier atoms
for methyl H atoms and 1.2Ueq for hydroxyl and the other H atoms.
The final refinement converged well. Materials for publication were
prepared using SHELXTL26 and PLATON.27
3.4.2. 1,3-Diacetoxy-7-geranyloxyxanthone (17)
Yellow powder, mp 96–97 ꢂC; UV (CHCl3) lmax (log
3) 253 (4.59),
300 (3.45), 361 (3.86) nm; IR (neat) nmax 3429, 1776, 1656,
1624 cmꢀ1; HRMS m/z 464.1838 for C27H28O7 (calcd 464.1835).
EIMS m/z (rel int.): 464 [M]þ (2), 328 (3), 286 (58), 244 (100), 187
(5), 81 (17), 69 (36). For 1H and 13C NMR spectroscopic data, see
Tables 3 and 4.
3.4.3. 7-Acetoxy-3-geranyloxy-1-hydroxy-xanthone (18)
Yellow powder, mp 104–106 ꢂC; UV (CHCl3) lmax (log
3) 243
(4.44), 257 (4.52), 310 (4.29), 358 (3.82) nm; IR (neat) nmax 3429,
1758, 1665, 1607 cmꢀ1; HRMS m/z 422.1726 for C25H26O6 (calcd
422.1729). EIMS m/z (rel int.): 422 [M]þ (5), 286 (16), 244 (100), 187
(2), 81 (16), 69 (56). For 1H and 13C NMR spectroscopic data, see
Tables 3 and 4.
Crystal data for cocrystal of 1 and 1a: C28.60H32.40O5, M¼465.15,
0.60ꢁ0.19ꢁ0.10 mm3, monoclinic, P21/c, a¼21.6161(6) Å, b¼
5.3826(2) Å, c¼22.8296(8) Å,
a
¼90.00ꢂ,
b
¼121.267ꢂ(2),
g
¼90.00ꢂ,
V¼2270.44(13) Å3, Z¼4, Dx¼1.334 Mg mꢀ3
,
m
(Mo K
a
)¼2.512 mmꢀ1
,
29,338 reflection measured, 6634 unique reflections, R¼0.0995,
Rw¼0.2794.
3.4.4. 1,7-Diacetoxy-3-geranyloxyxanthone (19)
Yellow powder, mp 85–87 ꢂC; UV (CHCl3) lmax (log
3) 246 (4.61),
Crystal data for 18: C25H26O6, M¼422.46, 0.58ꢁ0.27ꢁ0.06 mm3,
orthorhombic, P212121, a¼7.3280(2) Å, b¼13.0634(5) Å, c¼45.6190
275 (4.01), 302 (4.28), 334 (3.86) nm; IR (neat) nmax 3453, 1770,
1655, 1629 cmꢀ1
;
HRMS m/z 464.1834 for C27H28O7 (calcd
(18) Å,
(Mo K
a
¼
b
¼
g
¼90.00ꢂ, V¼4367.0(3) Å3, Z¼4, Dx¼1.285 Mg mꢀ3
,
464.1835). EIMS m/z (rel int.): 464 [M]þ (4), 328 (4), 286 (32), 244
(100), 187 (2), 81 (24), 69(73). For 1H and 13C NMR spectroscopic
data, see Tables 3 and 4.
m
a
)¼0.091 mmꢀ1, 81,117 reflection measured, 4877 unique
reflections, R¼0.1258, Rw¼0.3299.
Crystal
data
for
20:
C35H30Br2O9S2,
M¼818.53,
0.60ꢁ0.18ꢁ0.03 mm3, triclinic, Pꢀ1, a¼9.0779(4) Å, b¼19.3468(7)
3.5. Brosylation of 14
Å, c¼20.7939(7) Å,
a
¼108.160(2)ꢂ,
b
¼91.755(2)ꢂ,
g
¼90.046(2)ꢂ,
V¼3468.3(2) Å3,
Z¼4,
Dx¼1.568 Mg mꢀ3
,
m(Mo
K
a)¼
Compound 14 (40.0 mg, 105.14 mmol) was stirred overnight
at room temperature with p-bromobenzenesulfonyl chloride
(40.30 mg,190.2 mmol) and K2CO3 (44.1 mg, 315.4 mmol) in CH2Cl2
(3 ml). After the reaction was complete, water (10 ml) was added to
the reaction mixture. The resulting solutionwas then extracted with
CH2Cl2 (10 ml, three times). The combined organic extract was dried
over anhydrous sodium sulfate and evaporated under reduced
pressure to give a crude extract, which was further purified by col-
umn chromatography over silica gel eluting with 5% acetone–hex-
ane to yield the dibrosylate 20 (75.2 mg) (Rf (5% acetone–hexane
2.512 mmꢀ1, 41,231 reflection measured, 12,219 unique reflections,
R¼0.0817, Rw¼0.2121.
The crystallographic-information files for cocrystal of 1 and 1a,
18, and 20 have been deposited in the Cambridge Crystallographic
Data Centre as CCDC694467, CCDC689924, and CCDC689378, re-
spectively. These data can be obtained free of charge via http://
www.ccdc.cam.ac.uk/data_request/cif, or by e-mailing data_
request@ccdc.cam.ac.uk, or by contacting the Cambridge Crystal-
lographic Data Centre, 12, Union Road, Cambridge CB2 1EZ, UK; fax:
þ44 1223 336033.

N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013
3013
3.7. Antimicrobial activity
authors also would like to thank the Scientific Equipment Center,
Prince of Songkla University for the SEM analysis.
3.7.1. Antibacterial assay
The isolated compounds from the resin and green fruits of C.
cochinchinense were tested against both Gram-positive and Gram-
negative bacteria: B. subtilis, S. aureus, TISTR517, E. faecalis
TISTR459, Methicillin-Resistant S. aureus (MRSA) ATCC43300,
Vancomycin-Resistant E. faecalis (VRE) ATCC 51299, Streptococcus
faecalis, S. typhi, S. sonei and P. aeruginosa. The microrganisms were
obtained from the culture collections, Department of Industrial
Biotechnology and Department of Pharmacognosy and Botany, PSU,
except for the TISTR and ATCC strains, which were obtained from
Microbial Research Center (MIRCEN), Bangkok, Thailand. The
antibacterial assay employed was the same as described in Boonsri
et al.5 Vancomycin, which was used as a standard, showed anti-
bacterial activity against Vancomycin-Resistant E. faecalis (VRE)
References and notes
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Acknowledgements
We are greatly indebted to the Thailand Research Fund (TRF) for
research grant (RSA5280033). Financial support from the Center of
Excellence for Innovation in Chemistry (PERCH-CIC), Commission
on Higher Education, Ministry of Education is gratefully acknowl-
edged. N.B. thanks the Development and Promotion of Science and
Technology Talents Project for fellowship and Graduate School,
Prince of Songkla University for financial supports. The authors also
thank the Koshinocorporation Group, Japan and Universiti Sains
Malaysia for the Research University Golden Goose grant No. 1001/
PFIZIK/811012. N.B. thanks Asst. Prof. Dr. Surat Laphookhieo for
supplying the authentic sample of 1,3,7-trihydroxyxanthone. The