Tetrahedron 65 (2009) 3003–3013

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Tetrahedron
journal homepage: www.elsevier.com/locate/tet

Anti-Pseudomonas aeruginosa xanthones from the resin and green fruits

of Cratoxylum cochinchinense
Nawong Boonnak a, b, Chatchanok Karalai a, b, *, Suchada Chantrapromma a, Chanita Ponglimanont b,
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
Received 23 October 2008 the resin and green fruits of Cratoxylum cochinchinense. In addition, four new acetylated compounds (16–
Received in revised form 8 January 2009 19) were derivatized from 7-geranyloxy-1,3-dihydroxyxanthone (14) and 3-geranyloxy-1,7-dihydroxy-
Accepted 22 January 2009
xanthone (15). All compounds were characterized on the basis of spectroscopic analyses. The structures
Available online 29 January 2009
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
Keywords:
well.
Cratoxylum cochinchinense
1,3,7-Trihydroxyxanthone
Ó 2009 Elsevier Ltd. All rights reserved.
1,3,7-Trioxygenated xanthone
Antifungal activity
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, evaluated.
which is distributed in several parts of Thailand.1 This plant is
a well-known tropical tree, and is commonly known in Thailand as
2. Results and discussion
Tui Kliang. The bark, roots, and leaves of this plant have been used
in folk medicine to treat fever, coughs, diarrhea, itches, ulcers, and
The crude CH2Cl2 extract of the resin of C. cochinchinense was
abdominal complaints.2 Previous investigations revealed the major
subjected to chemical investigation leading to the isolation of three
components from the Cratoxylum genus as xanthones and anthra-
new xanthones, cochinchinones I–K (1–3), together with nine
quinones.3–6 Xanthones have been reported to exhibit biological
known xanthones identified as cochinchinone A (4),6 1,3,7-tri-
activities such as antibacterial,3–5 antioxidant,6,7cytotoxic,3–5,8,9
hydroxy-2,4-diisoprenylxanthone (5),14 celebixanthone methyl
and antiprotozoal activities.10 A preliminary screening of the bio-
ether (6),15 dulxis-xanthone F (7),15 b-mangostin (8),16 a-mangostin
activity of the crude extracts from the resin and green fruits of C.
(9),16 macluraxanthone (10),17 pruniflorone G (11),3 and a caged-
cochinchinense has shown strong antibacterial activity specifically
prenylated xanthone (12).6 Their structures were elucidated by
against Pseudomonas aeruginosa. Although Pseudomonas sepsis is
NMR analysis and comparison of their spectroscopic data with
a major cause of death in transplant recipients, while P. aeruginosa
those reported in the literature.
is one of the most common Gram-negative pathogenic bacteria
Cochinchinone I (1) was obtained as a yellow powder, which
causing life-threatening infections.11–13 This result led us to
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
* Corresponding author. Tel.: þ66 7428 8444; fax: þ66 7421 2918. by HRMS. Its structure was supported by the 1H and 13C NMR
E-mail addresses: [email protected], [email protected] (C. Karalai). spectral data (Table 1). The 1H and 13C NMR spectral data of 1 were

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 cm1, re-
spectively. The 1H and 13C NMR data of 2 (Table 1) were similar to
those of 4,6 except for the appearance of the signals of a chromene ring
bearing a methyl group and six-carbon side chain of 4-methylpent-3-
enyl group, which appeared at dH 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 moiety at 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
Figure 1. The X-ray structure of 1.
chromene ring was confirmed by HMBC correlations (Table 2) in
which the methine proton H-100 (d 6.88) was correlated with C-3 (d
comparable to cochinchinone A (4), which was previously isolated 158.9), C-4 (d 100.2), C-4a (d 150.2), and C-300 (d 80.6). Therefore, the
from the roots of C. cochinchinense.6 The main difference was ob- structure of cochinchinone J was deduced to be 2.
served at C-2 and C-3, where the 1H NMR spectral data of 1 showed Compound 3 was isolated as an inseparable mixture with
the signals of a chromene ring at d 6.74 (d, J 9.9 Hz, H-10 ), 5.60 (d, J compound 2, and the mixture was thus acetylated with Ac2O in
9.9 Hz, H-20 ), and 1.48 (s, Me-40 and Me-50 ) instead of a prenyl pyridine. The resulting product was further separated by CC eluting
group at C-2 and a free hydroxyl group at C-3 as in 4 (Table 1). The with 70% CHCl3–hexane to give monoacetates 3a (18.9 mg) and 2a

Table 1
1
H and 13C (300 MHz) spectral data of 1, 2, 3a, and 4 in CDCl3

Position 1 2 3a 4
1 13 1 13 1 13 1 13
H (J in Hz) C (d) H (J in Hz) C ( d) H (J in Hz) C (d) H (J in Hz) C (d)
1-OH 12.97, s 155.5 13.15, s 160.2 13.02, s 158.4 12.79, s 158.1
2 104.2 111.2 103.9 109.1
3 158.4 158.9 159.6 161.1
4 107.4 100.2 107.3 104.9
5 7.30, d, 9.0 119.0 7.35, d, 8.7 118.9 7.46, d, 8.7 118.7 7.04, d, 9.0 118.7
6 7.23, dd, 2.4, 9.0 124.1 7.25, m 123.8 7.39, dd, 8.7, 2.7 128.6 7.07, br d, 9.0 124.7
7 150.5 150.3 146.3 150.1
8 7.60, d, 2.4 108.9 7.60, br d, 1.8 109.3 7.92, d, 2.7 117.8 7.44, br s 108.7
9 180.9 180.5 180.3 180.9
4a 154.5 150.2 152.4 152.9
4b 152.4 152.2 153.7 152.4
8a 120.6 121.1 121.2 120.2
9a 103.3 102.9 102.3 103.0
10 6.74, d, 9.9 115.8 3.36, d, 7.2 21.2 2.75, br t, 6.9 16.2 3.32, d, 6.9 21.8
20 5.60, d, 9.9 127.3 5.25, br t, 7.2 122.1 1.81, br t, 6.9 31.7 5.19, br t, 6.9 121.6
30 78.1 131.5 76.3 134.8
40 1.48, s 28.4 1.68, s 25.8 1.39, s 26.9 1.55, s 25.6
50 1.48, s 28.4 1.81, s 17.9 1.39, s 26.9 1.75, s 17.9
100 3.46, d, 7.2 21.3 6.88, d, 10.2 115.9 3.48, d, 7.5 21.5 3.39, d, 6.9 21.6
200 5.22, br t, 7.2 122.1 5.54, d, 10.2 125.4 5.22, br t, 6.3 122.3 5.16, br t, 6.9 121.7
300 135.0 80.6 134.9 137.9
400 1.99, ma 39.7 1.89, m,a 1.68, ma 41.8 1.96, m 39.8 1.99, ma 39.7
500 2.04, ma 26.6 2.12, ma 22.8 2.03, m 26.7 1.97, ma 26.4
600 5.04, br t, 6.6 124.2 5.10, br t, 7.2 123.8 5.05, br t, 6.6 124.4 4.95, br t, 7.2 123.9
700 131.3 131.9 131.3 131.8
800 1.59, s 25.6 1.66, s 25.6 1.60, s 25.6 1.67, s 25.8
900 1.86, s 16.3 1.44, s 27.1 1.86, s 16.3 1.78, s 16.2
1000 1.53, s 17.6 1.57, s 17.6 1.55, s 17.7 1.48, s 17.6
7-OAc 2.34, 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 C-1, C-2, C-9a C-1, C-2, C-9a C-1, C-2, C-9a C-1, C-2, C-9, C-9a
5 C-7, C-9, C-4b, C8a C-7, C8a C-7, C-8a, C-9 C-7, C-9, C-4b, C8a
6 C-5, C-7, C-8, C-4b C-7, C-4b C-5, C-7, C-4b C-7, C-8
8 C-6, C-7, C-9, C-4b C-8 C-6, C-7, C-4b, C-9 C-6, C-7, C-9, C-4b
10 C-1, C-2, C-3, C-4, C-30 C-1, C-2, C-20 , C-30 C-1, C-2, C-3, C-20 , C-30 C-1, C-2, C-3, C-20 , C-30 C-40 , C-50
20 C-2, C-30 , C-40 , C-50 C-20 C-2, C-10, C-30 , C-40 , C-50 C-2, C-10, C-50
40 C-20 , C-30 , C-50 C-20 , C-30 , C-50 C-20 , C-30 C-20 , C-30 , C-50
50 C-20 , C-30 , C-40 C-20 , C-30 , C-40 C-20 , C-30 C-20 , C-30 , C-40
100 C-3, C-4, C-4a, C-200 , C-300 C-3, C-4, C-4a, C-300 C-3, C-4, C-4a, C-200 , C-300 C-3, C-4, C-4a, C-200 , C-300 , C-900
200 C-4, C-100, C-400 , C-900 C-4, C-300 , C-400 , C-500 , C-900 C-100, C-300 , C-400 , C-900 C-4, C-100, C-400 , C-900
400 C-200 , C-300 , C-500 C-200 , C-300 C-200 , C-300 , C-600 C-200 , C-600
500 C-300 , C-400 , C-600 , C-700 C-600 , C-700 C-300 , C-600 , C-700 C-300 , C-400 , C-600 , C-700
600 C-400 , C-500 , C-800 , C-1000 C-600 C-500 , C-800 , C-1000 C-400 , C-500 , C-800
800 C-600 , C-700, C-1000 C-600 , C-700, C-1000 C-600 , C-700, C-1000 C-600 , C-700, C-1000
900 C-200 , C-300 C-200 , C-300 , C-400 C-200 ,C-300 C-200 , C-300 , C-400
1000 C-600 , C-700, C-800 C-600 , C-700, C-800 C-600 , C-700, C-1000 C-600 , C-700, C-800
7-OAc C-7

(5.1 mg). The latter was confirmed as an acetylated derivative of 2 with C-1 (d 158.4), C-2 (d 103.9), and C-9a (d 102.3). Therefore, the
by comparison of its spectral data with those of compound 2. structure of 3a, an acetylated form of 3, was deduced. Compound 3
Compound 3a is a yellow powder. The HRMS spectrum showed was named cochinchinone K.
a molecular ion peak at m/z 490.2355 [M]þ, corresponding to It is interesting to note that the three new chromenoxanthones (1
C30H34O6. The UV spectrum showed absorption bands of a xan- and 2) and chromanoxanthone (3) could be derived from cochin-
thone (243, 271, 303, 326, and 383 nm),14 while the IR spectrum chinone A (4). The isoprenyl or geranyl side chains of 4 were epox-
exhibited the hydroxyl and conjugated carbonyl functionalities at idized and further cyclized via a free hydroxyl at C-3 to produce the
vmax 3392 and 1648 cm1, respectively. The 1H and 13C NMR spec- chromanol ring18 as shown in Scheme 1. Further dehydration led to
tral data of 3a (Table 1) were closely related to those of 1, except for the linear or angular chromene rings of 1 and 2, respectively. On the
the appearance of the signals of a dimethylchromane ring and other hand, the new chromanoxanthone (3) could be produced by
acetoxy group revealed at dH 2.75 (br t, J 6.9 Hz, H-10 ), 1.81 (br t, J cyclization via a free hydroxyl group to an isoprenyl side chain of 4 to
6.9 Hz, H-20 ), 1.39 (s, Me-40 and Me-50 ), and 2.34 (s, 7-OAc) (Table 1) afford the chromane ring (Scheme 1). Therefore, cochinchinone A (4)
instead of a chromene ring and a hydroxyl group in 1. The chromane should be a precursor of cochinchinones I–K (1–3).
ring was fused to the xanthone nucleus in a linear fashion, which The investigation of the crude CH2Cl2 extract of the green fruits
was confirmed by HMBC correlations (Table 2), in which the (approximate 4 weeks maturity stage) led to the isolation and
methylene protons H2-10 at dH 2.75 were correlated with C-1 (d identification of a new xanthone, cochinchinone L (13) along with
158.4), C-2 (d 103.9), C-3 (d 159.6), C-20 (d 31.7), and C-30 (d 76.3), two major known oxygeranyl xanthones 1414 and 15.19 Compounds
while the H-bonded hydroxyl proton 1-OH (d 13.02) was correlated 14 and 15 were structural isomers of xanthones with an oxygeranyl

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 OH O OH
8 1 1' 8 1 1'
3' HO
HO 9 2'
9
cyclization
6
6 O 3 OH O 3 O 3'

1''
4 chromanoxanthone (3)
3''

epoxidation and 5''

cyclization epoxidation and
7''
cyclization

O OH
8 1 1' O OH
HO OH 8
9 HO 1
9
6
O 3 O 3'
6 O 3 O
3'' 7''
1''
5''
OH

chromanolxanthone intermediate

chromanolxanthone intermediate dehydration

-H2O
dehydration
-H2O
O OH
O OH 8 1
8
HO 9
1 1'
HO 2'
9
6 3 O
O
6 O 3 O 3' 3'' 7''
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
ence was observed in the NOESY experiment in which H2-10 (d 4.45) confirm these structural assignments of 14 and 15, compound 14
of 14 showed a cross-peak with H-8 (d 7.40) and H-6 (d 7.18), was further reacted with p-bromobenzenesulfonyl chloride in
whereas the H2-10 (d 4.63) of 15 showed a cross-peak with H-4 (d CH2Cl2 at room temperature to afford a crystalline dibrosylate 20

Table 3
1
H NMR (300 MHz) spectral data of 13–19 in CDCl3

Position 13 14 15 16 17 18 19
2 6.43, d, 2.1 6.22, d, 1.8 6.34, d, 2.4 6.42, br d, 2.1 7.15, d, 2.1 6.19, br d, 2.1 6.41, d, 2.1
4 6.57, d, 2.1 6.23, d, 1.8 6.40, d, 2.4 6.59, br d, 2.1 6.79, d, 2.1 6.24, br d, 2.1 6.46, d, 2.1
5 7.08, br d, 9.0 7.15, br d, 9.0 7.30, d, 9.3 7.21, br d, 8.1 7.23, d, 9.3 7.25, d, 9.3 7.25, d, 8.1
6 7.10, br d, 9.0 7.18, br d, 9.0 7.26, d, 9.3 7.20, br d, 8.1 7.28, dd, 9.3, 2.1 7.29, br d, 9.3 7.26, br d, 8.1
8 7.45, br s 7.40, br s 7.59, br s 7.44, br s 7.55, d, 2.1 7.76, br s 7.78, br d, 1.5
10 4.42, d, 6.3 4.45, d, 6.3 4.63, d, 6.6 4.50, d, 6.6 4.56, d, 6.3 4.49, d, 6.3 4.51, d, 6.3
20 5.73, t, 6.3 5.39, br t, 6.3 5.50, br t, 6.6 5.40, br t, 6.6 5.48, br t, 6.3 5.37, br t, 6.3 5.37, br t, 6.3
40 1.99, m 2.01, m 2.13, m 2.03, m 2.11, m 2.03, m 2.03, m
50 1.97, m 1.95, m 2.10, m 2.02, m 2.09, m 1.97, m 2.01, m
60 4.98, t, 6.6 4.98, br t, 5.7 5.11, br t, 5.7 5.00, br t, 6.3 5.09, br t, 6.3 4.99, br t, 6.9 4.99, br t, 6.6
80 1.57, s 1.57, s 1.69, s 1.57, s 1.66, s 1.57, s 1.57, s
90 1.60, s 1.64, s 1.78, s 1.68, s 1.73, s 1.66, s 1.65, s
100 1.50, s 1.50, s 1.62, s 1.51, s 1.59, s 1.51, s 1.51, s
1-OAc 2.38, s d d d d d d
1-OH d 12.85, s 12.72, s 12.71, s d 12.55, s d
3-OH d 7.86, br s d d d d
7-OH d d 7.03, br s d d d d
1-OAc d d d d 2.47, s d 2.40, s
3-OAc d d d 2.23, s 2.27, s d d
7-OAc d 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 acetoxy group in 13, which appeared as a proton singlet signal at
with Ac2O in pyridine to give a crystalline monoacetate 18 (Fig. 2). d 2.38. The higher field chemical shift of C-1 (d 151.2) and lower
Both 20 and 18 were subjected to X-ray diffraction analysis whose field resonances of protons and carbons of C-2 (dH 6.43 (d, J 2.1 Hz,
results supported the structures of 14 and 15. H-2); dC 108.3) and C-4 (dH 6.57 (d, J 2.1 Hz, H-4); dC 101.3) as
Cochinchinone L (13) was isolated as a yellow powder, with compared to those of 14 (dH 6.22 (d, J 1.8 Hz, H-2); dC 98.5 and dH
a molecular ion peak at m/z 422.1718 [M]þ in the HRMS, corre- 6.23 (d, J 1.8 Hz, H-4); dC 94.4) supported the title structure (Tables
sponding to a molecular formula of C25H26O6. The 1H and 13C NMR 3 and 4). The 4J HMBC correlations of methyl protons of the ace-
spectral data of 13 (Tables 3 and 4) showed characteristic signals toxy group (d 2.38) with C-1 (d 151.2) supported its location at C-1,
similar to those of 7-geranyloxy-1,3-dihydroxyxanthone (14),14 which was also confirmed by NOESY cross-peak between acetoxy
except that the hydroxyl group at C-1 of 14 was replaced by an moiety with H-2. Cochinchinone L (13), an acetylated analogue of

5'
O OH O OH O OH
8 1 1' 8 1 1' 8 1 1' 3'
HO 8a 9a HO 8a 9a 8a 9a
9 2' 9 2' O
R 9
4' 2' 4'
4'
6 6 6 10''
4b O 4a 3 O 3' 5' 4b O 4a 3 O 3' 5' 4b O 4a 3 O
3'' 7''
1'' 1'' 1''
5'' 8''
1 1a 2:R=H 9''
3'' 3''
9'' 9'' 2a : R = Ac
5'' 5''
7''
7'' 8''
10'' 8'' 10''
11''

5'
O OH O OH O OH
8 1' 8 1 1' 3'
O 8a 9a 1 HO 8a 9a HO
R 9 2' 9
2' 4'
4'
6 6
4b O 4a 3 O 3' 5' 4b O 4a 3 OH O OH
1'' 1''

3:R=H 4 5
3'' 3''
9''
3a : R = Ac 5'' 5''

7'' 7''
10'' 8'' 10'' 8''

O OH O OH O OH
O R2
H3CO

H3CO O HO O OCH3 HO O R1
OH
8: R1 = OCH3 R2 = OCH3
6 7

9: R1 = OH R2 = OCH3

O OH O OR1 O OR1
8' 8 1 8 1
3' O R2O 9' 10'
9 9
7' 5' 1' B A
R1 1' 3' 7'
HO O O 10' 9' 6 O 3 OR2 6 O 3 O 5' 8'
OH
13: R1 = Ac R2 = H 15: R1 = H R2 = H
1
1
10: R = CH3 14: R = H 1
R =H2 18: R = H R2 = Ac
1 1
16: R = H R2 = Ac 19: R = Ac R2 = Ac
11: R1=
17: R1 = Ac R2 = Ac
3''
Br
O 1''
O OH 5''
S O OH
H3CO O O 3'''
8' 3' O
8 1 O Br
HO
8 1
9 O 1''' 9
7' 5' 1' B A
O O S 5'''
10' 9' 6 O 3 O 6 O 3 OH
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,

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

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 results was correlated to their strong antibacterial activity (Table 5).
geranyl side chain at C-3 or C-7 in 13–19 are essential for their Therefore, it can be suggested that compounds 4 and 13 may interact
antibacterial activity against P. aeruginosa. Therefore, 1,3,7-trihy- with or damage the cell wall of P. aeruginosa as seen by the formation
droxyxanthones (4 and 5) and 1,3,7-trioxygenated xanthone with of pores on the cell wall of P. aeruginosa (Figs 4 and 5).
geranyl side chain (13–19) should be considered as potent candi-
dates as anti-P. aeruginosa. 3. Experimental
We further studied the possible mode of action of compounds 4
and 13 against P. aeruginosa by observing the bacteria cell morpho- 3.1. General experimental procedures
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 Melting points were determined on a Fisher–John melting point
and 5), it was clearly indicated that the cell morphology of P. aeru- apparatus. Optical rotations were measured on a JASCO P-1020
ginosa, when treated with compounds 4 and 13, started to deform at digital polarimeter. UV and IR spectra were recorded on SPECORD S
3 h onward, and at 15 h, most cells were completely deformed whose 100 (Analytikjena) and Perkin–Elmer FTS FT-IR spectrophotometer,

Table 5
Antimicrobial activity (MIC, mg/ml) of 1–21

No. Antibacterial activity Antifungal activity

a b
Gram-positive bacteria Gram-negative bacteria

B. subtilis S. aureus E. faecalis MRSA VRE S. typhi S. sonei P. aeruginosa C. albicansc

1 >300 >300 >300 >300 >300 >300 >300 >300 >300
2 300 300 >300 300 300 >300 >300 300 75
3a 75 >300 300 >300 >300 >300 >300 >300 300
4 150 150 150 9.37 150 >150 >150 4.7 75
5 150 75 150 37.5 75 >150 >150 4.7 37.5
4D5 75 150 75 9.37 150 >150 >150 4.7 75
6 >150 >150 >150 150 150 >150 >150 4.7 150
7 150 300 300 150 150 >300 >300 150 300
8 300 75 150 75 150 >300 >300 300 150
9 9.37 9.37 9.37 9.37 9.37 >300 >300 18.75 2.4
10 18.75 37.5 37.5 37.5 37.5 >300 >300 37.5 4.7
11 >300 300 >300 150 150 >300 >300 >300 300
12 75 >300 300 150 150 >300 >300 >300 >300
13 150 >150 150 37.5 150 >150 >150 4.7 18.75
14 150 >150 150 18.7 150 >150 >150 4.7 75
15 150 150 150 9.3 150 >150 >150 4.7 37.5
14D15 75 37.5 37.5 4.7 37.5 >150 >150 4.7 37.5
16 150 >150 150 18.75 150 >150 >150 4.7 150
17 75 >150 75 37.5 150 >150 >150 4.7 18.75
18 >150 >150 150 18.75 150 >150 >150 4.7 150
19 >150 >150 >150 37.5 150 >150 >150 4.7 150
20 >150 >150 150 300 >300 >150 >150 4.7 >300
21 37.5 75 >300 150 300 >300 >300 300 300
a
B. subtilis, S. aureus, E. faecalis TISTR 459, Methicillin-Resistant S. aureus (MRSA) ATCC 43300, Vancomycin-Resistant E. faecalis (VRE) ATCC 51299.
b
S. typhi, S. sonei, and P. aeruginosa.
c
C. albicans.
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 subfractions (FR4A–FR4H) and 8 (150.2 mg) (Rf (15% acetone–hex-
300 MHz Bruker FTNMR Ultra ShieldÔ spectrometer in CDCl3 with ane (three runs)) 0.39). Subfraction FR4B was further purified by CC
TMS as the internal standard. Chemical shifts are reported in and eluted with a gradient of acetone–hexane to give 7 (31.4 mg)
d (ppm) and coupling constants (J) are expressed in hertz. EI and (Rf (15% acetone–hexane) 0.31) and 12 (56.5 mg) (Rf (15% acetone–
HREI mass spectra were measured on a Kratos MS 25 RFA hexane) 0.28). Fractions FR6 and FR7 were separated by QCC and
spectrometer. Quick column chromatography (QCC) and column eluted with a gradient of CH2Cl2–hexane to give six subfractions
chromatography (CC) were carried out on silica gel 60 F254 (Merck) (FR6A–FR6F). Subfraction FR6B was further separated by QCC
and silica gel 100 (Merck), respectively. All the bacteria images eluting with a gradient of acetone–hexane to give 6 (35.7 mg)
were viewed with a JSM-5800LV, JEOL SEM (scanning electron (Rf (15% acetone–hexane (three runs)) 0.41), 8 (83.2 mg) (Rf (15%
microscope). 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
3.2. Plant material 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).
The resin of C. cochinchinense was collected in October 2003 at Fractions FR8 and FR9 were separated by QCC and eluted with
Prince of Songkla University, Hat-Yai campus, whereas the green a gradient of CH2Cl2–hexane to afford 5 (551.3 mg) (Rf (60% CH2Cl2–
fruits of C. cochinchinense were collected in October 2007 at Kaun hexane (three runs)) 0.05), 7 (18.2 mg) (Rf (60% CH2Cl2–hexane
Kha Long district, Satun Province, Southern part of Thailand. Bo- (three runs)) 0.29), 8 (116.6 mg) (Rf (60% CH2Cl2–hexane (three
tanical identification was achieved through comparison with runs)) 0.61), and 9 (148.7 mg) (Rf (60% CH2Cl2–hexane (three runs))
a voucher specimen No. SL-1 (PSU) in the herbarium of Department 0.10). Fractions FR10 and FR11 were separated by QCC and eluted
of Biology, Prince of Songkla University, Songkhla, Thailand. with a gradient of acetone–hexane to give seven subfractions
(FR10A–FR10G) and 7 (25.9 mg) (Rf (15% acetone–hexane) 0.31).
3.3. Isolation and extraction 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
The resin of C. cochinchinense (87.75 g) was extracted with mixture of 1 and 1a could not be separated due to the very low
CH2Cl2 (22.0 L, for a week) at room temperature and was evapo- amount of 1a. The purity of 1 is much more than 90% base on
rated under reduced pressure to afford a deep green crude CH2Cl2 a single spot on TLC and the integral area signal in the 1H NMR
extract (47.04 g), which was subjected to QCC (Quick column spectrum of 1. Subfraction FR10D was separated by QCC eluting
chromatography) on silica gel (Merck 60 F254) using hexane as with a gradient of acetone–hexane to give six subfractions
a first eluent and then increasing the polarity with acetone to give (FR10D1–FR10D6). Subfraction FR10D2 was further separated by CC
16 fractions (FR1–FR16). Fractions FR4 and FR5 were separated by and eluted with a gradient of acetone–hexane to give five sub-
CC eluting with a gradient of acetone–hexane to give eight 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 (73), 363 (76), 323 (26), 307 (15), 295 (13), 137 (5), 69 (6). For 1H and
13
(20% acetone–hexane) 0.17), and an inseparable mixture of 2 and 3 C NMR spectroscopic data, see Table 1.
(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 3.3.2. Cochinchinone J (2)
stirred overnight at room temperature to give a yellow gum, which Yellow viscous oil; [a]25
D 69.8 (c 0.08, CHCl3); UV (CHCl3) lmax
was further purified by CC eluting with 70% CHCl3–hexane to give (log 3) 243 (3.90), 289 (4.10), 298 (4.13), 320 (3.68), 351 (3.47), 391
acetylated derivatives 2a (5.1 mg) (Rf (70% CHCl3–hexane) 0.68) and (3.33) nm; IR (neat) nmax 3397, 1649, 1613 cm1; HRMS m/z
3a (18.9 mg) (Rf (70% CHCl3-hexane) 0.40), respectively. Subfraction 446.2092 for C28H30O5 (calcd 446.2093). EIMS m/z (rel int.): 446
FR10D2E was purified by CC and eluted with 80% CH2Cl2–hexane to [M]þ (11), 363 (100), 307 (18), 69 (5). For 1H and 13C NMR spec-
give 8 (16.7 mg) (Rf (80% CH2Cl2–hexane) 0.91), 10 (5.0 mg) (Rf (80% troscopic data, see Table 1.
CH2Cl2–hexane) 0.85), and 11 (1.5 mg) (Rf (80% CH2Cl2–hexane)
0.38). 3.3.3. Acetylated derivative of cochinchinone K (3a)
Air-dried green fruits of C. cochinchinense (5.5 kg) were Yellow powder, mp 85–87  C; UV (CHCl3) lmax (log 3) 243, 271,
extracted with CH2Cl2 (220 L, for a week) at room temperature 303, 326, 383 nm; IR (neat) nmax 3392, 1709, 1648, 1612 cm1;
and was evaporated under reduced pressure to afford a deep HRMS m/z 490.2355 for C30H34O6 (calcd 490.2355). EIMS m/z (rel
green crude CH2Cl2 extract (40.04 g), which was subjected to QCC int.): 490 [M]þ (53), 473 (53), 419 (44), 405 (31), 365 (100), 323 (52),
on silica gel using hexane as a first eluent and increasing polarity 311 (41), 267 (51), 69 (24). For 1H and 13C NMR spectroscopic data,
with EtOAc to give nine fractions (FS1–FS9). Fraction FS5 was see Table 1.
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- 3.3.4. Cochinchinone L (13)
rated by CC eluting with pure CHCl3 to furnish six subfractions Yellow powder, mp 114–116  C; UV (CHCl3) lmax (log 3) 248
(FS6A–FS6F), 14 (2.10 g) (Rf (70% CHCl3–hexane) (three runs) 0.46) (4.59), 273 (4.03), 305 (4.13), 354 (3.80) nm; IR (neat) nmax 3237,
and 15 (490.2 mg) (Rf (70% CHCl3–hexane) (three runs) 0.39). 1774, 1728, 1628 cm1; HRMS m/z 422.1718 for C25H26O6 (calcd
Subfraction FS6B was further purified by CC eluting with a gradi- 422.1729). EIMS m/z (rel int.): 422 [M]þ (1), 286 (40), 244 (100), 187
ent of acetone–hexane to give 13 (53.3 mg) (Rf (15% acetone– (4), 81 (9), 69 (28). For 1H and 13C NMR spectroscopic data, see
hexane (three runs)) 0.17). Tables 3 and 4.

3.3.1. Cochinchinone I (1) 3.4. Acetylation of 14 and 15

Yellow needle single crystals, mp 160–162  C; 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
nmax 3406, 1650, 1612 cm1; HRMS m/z 446.2279 for C28H30O5 pyridine (2.0 ml) and stirred for 6 h at room temperature. The re-
(calcd 446.2093). EIMS m/z (rel int.): 446 [M]þ (76), 431 (100), 377 action mixture was diluted with water and extracted with CH2Cl2.
3012 N. Boonnak et al. / Tetrahedron 65 (2009) 3003–3013

The combined organic extract was washed with 10% HCl and then (three runs)) 0.20). Yellow needle-shaped single crystals of 20 were
washed with water again. After the organic solvent was removed, obtained after recrystallization from CH3OH–CHCl3 (1:4 v/v), mp
the resulting residue was dried over anhydrous Na2SO4. Chroma- 106–108  C. dH (300 MHz, CDCl3þCD3OD) 7.82 (dd, J 9.0, 2.1 Hz,
tography over silica gel yielded a pale yellow powder of 16 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 ,
(80.6 mg) (Rf (5% acetone–hexane (three runs)) 0.44). H-2000 , H-6000 ), 7.63 (dd, J 8.7, 2.1 Hz, H-3000 , H-5000 ), 7.50 (d, J 2.4 Hz,
Compound 14 (200.5 mg) was treated with Ac2O (6.0 ml) in 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,
pyridine (3.0 ml) and stirred overnight at room temperature. 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
Chromatography over silica gel yielded a pale yellow powder of 16 6.6 Hz, H-10 ), 2.07 (m, 2H-50 ), 2.05 (m, 2H-40 ), 1.71 (s, 3H-90 ),1.60 (s,
(10.6 mg) (Rf (5% acetone–hexane (three runs)) 0.44) and 17 3H-80 ),1.54 (s, 3H-100 ); dC (75 MHz, CDCl3þCD3OD) 181.6 (C-9),157.4
(177.8 mg) (Rf (15% acetone–hexane (three runs)) 0.46). (C-1), 155.9 (C-7), 151.9 (C-3), 149.7 (C-4b), 148.6 (C-4a), 142.1 (C-30 ),
Compound 15 (85.5 mg) was treated with Ac2O (2.5 ml) in 134.3 (C-100 ), 133.4 (C-1000 ), 133.1 (C-300 , C-500 ), 132.5 (C-3000 , C-5000 ),
pyridine (2.0 ml) and stirred for 6 h at room temperature. Chro- 131.8 (C-70 ), 130.6 (C-400 ), 130.2 (C-200 , C-600 ), 130.1 (C-4000 ), 129.8 (C-
matography over silica gel yielded a pale yellow powder of 18 2000 , C-6000 ), 124.9 (C-6), 123.7 (C-60 ), 122.4 (C-8a),118.8 (C-5), 118.6 (C-
(83.7 mg) (Rf (5% acetone–hexane (three runs)) 0.32), which was 20 ), 114.2 (C-9a), 112.8 (C-4), 111.1 (C-2), 106.8 (C-8), 65.6 (C-10 ), 39.5
further recrystallized from MeOH–acetone (1:99 v/v) to give yellow (C-40 ), 26.2 (C-50 ), 25.5 (C-80 ), 17.5 (C-100 ), 16.6 (C-90 ). EIMS m/z (rel
single crystals. int.): 816 [M2]þ (1), 683 (7), 681 (14), 679 (7), 619 (5), 617 (10), 615
Compound 15 (190.0 mg) was treated with Ac2O (6.0 ml) in (5), 461 (55), 463 (56), 399 (41), 397 (41), 371 (19), 369 (19), 357 (34),
pyridine (3.0 ml) and stirred overnight at room temperature. 355 (34), 244 (27), 229 (84), 215 (100),186 (16),157 (53),155 (53),131
Chromatography over silica gel yielded a pale yellow powder of 18 (11), 108 (13), 76 (16).
(8.7 mg) (Rf (5% acetone–hexane (three runs)) 0.32) and 19
(170.0 mg) (Rf (15% acetone–hexane (three runs)) 0.37).

3.4.1. 3-Acetoxy-7-geranyloxy-1-hydroxyxanthone (16) 3.6. X-ray crystallographic studies of cocrystal of 1 and 1a,
Yellow powder, mp 94–95  C; UV (CHCl3) lmax (log 3) 239 (4.40), monoacetate 18, and dibrosylate 20
262 (4.66), 289 (3.97), 379 (3.93) nm; IR (neat) nmax 3429, 1768,
1649, 1612 cm1; HRMS m/z 422.1725 for C25H26O6 (calcd Crystallographic data were collected at 100.0 (1) K with the
422.1729). EIMS m/z (rel int.): 422 [M]þ (1), 286 (43), 244 (100), 187 Oxford Cryosystem Cobra low-temperature attachment. The data
(4), 81 (9), 69 (24). For 1H and 13C NMR spectroscopic data, see were collected using a Bruker Apex2 CCD diffractometer with a
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
3.4.2. 1,3-Diacetoxy-7-geranyloxyxanthone (17) program,25 and the empirical absorption corrections were per-
Yellow powder, mp 96–97  C; UV (CHCl3) lmax (log 3) 253 (4.59), formed using SADABS program.25 The structures were solved by
300 (3.45), 361 (3.86) nm; IR (neat) nmax 3429, 1776, 1656, direct methods and refined by least-squares using the SHELXTL
1624 cm1; HRMS m/z 464.1838 for C27H28O7 (calcd 464.1835). software package26 All non-hydrogen atoms were refined aniso-
EIMS m/z (rel int.): 464 [M]þ (2), 328 (3), 286 (58), 244 (100), 187 tropically, whereas all H atoms were placed in calculated positions
(5), 81 (17), 69 (36). For 1H and 13C NMR spectroscopic data, see with an O–H distance of 0.82 Å and C–H distances in the range
Tables 3 and 4. 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
3.4.3. 7-Acetoxy-3-geranyloxy-1-hydroxy-xanthone (18) for methyl H atoms and 1.2Ueq for hydroxyl and the other H atoms.
Yellow powder, mp 104–106  C; UV (CHCl3) lmax (log 3) 243 The final refinement converged well. Materials for publication were
(4.44), 257 (4.52), 310 (4.29), 358 (3.82) nm; IR (neat) nmax 3429, prepared using SHELXTL26 and PLATON.27
1758, 1665, 1607 cm1; HRMS m/z 422.1726 for C25H26O6 (calcd Crystal data for cocrystal of 1 and 1a: C28.60H32.40O5, M¼465.15,
422.1729). EIMS m/z (rel int.): 422 [M]þ (5), 286 (16), 244 (100), 187 0.600.190.10 mm3, monoclinic, P21/c, a¼21.6161(6) Å, b¼
(2), 81 (16), 69 (56). For 1H and 13C NMR spectroscopic data, see 5.3826(2) Å, c¼22.8296(8) Å, a¼90.00 , b¼121.267 (2), g¼90.00 ,
Tables 3 and 4. V¼2270.44(13) Å3, Z¼4, Dx¼1.334 Mg m3, m(Mo Ka)¼2.512 mm1,
29,338 reflection measured, 6634 unique reflections, R¼0.0995,
3.4.4. 1,7-Diacetoxy-3-geranyloxyxanthone (19) Rw¼0.2794.
Yellow powder, mp 85–87  C; UV (CHCl3) lmax (log 3) 246 (4.61), Crystal data for 18: C25H26O6, M¼422.46, 0.580.270.06 mm3,
275 (4.01), 302 (4.28), 334 (3.86) nm; IR (neat) nmax 3453, 1770, orthorhombic, P212121, a¼7.3280(2) Å, b¼13.0634(5) Å, c¼45.6190
1655, 1629 cm1; HRMS m/z 464.1834 for C27H28O7 (calcd (18) Å, a¼b¼g¼90.00 , V¼4367.0(3) Å3, Z¼4, Dx¼1.285 Mg m3,
464.1835). EIMS m/z (rel int.): 464 [M]þ (4), 328 (4), 286 (32), 244 m(Mo Ka)¼0.091 mm1, 81,117 reflection measured, 4877 unique
(100), 187 (2), 81 (24), 69(73). For 1H and 13C NMR spectroscopic reflections, R¼0.1258, Rw¼0.3299.
data, see Tables 3 and 4. Crystal data for 20: C35H30Br2O9S2, M¼818.53,
0.600.180.03 mm3, triclinic, P1, a¼9.0779(4) Å, b¼19.3468(7)
3.5. Brosylation of 14 Å, c¼20.7939(7) Å, a¼108.160(2) , b¼91.755(2) , g¼90.046(2) ,
V¼3468.3(2) Å3, Z¼4, Dx¼1.568 Mg m3, m(Mo Ka)¼
Compound 14 (40.0 mg, 105.14 mmol) was stirred overnight 2.512 mm1, 41,231 reflection measured, 12,219 unique reflections,
at room temperature with p-bromobenzenesulfonyl chloride R¼0.0817, Rw¼0.2121.
(40.30 mg, 190.2 mmol) and K2CO3 (44.1 mg, 315.4 mmol) in CH2Cl2 The crystallographic-information files for cocrystal of 1 and 1a,
(3 ml). After the reaction was complete, water (10 ml) was added to 18, and 20 have been deposited in the Cambridge Crystallographic
the reaction mixture. The resulting solution was then extracted with Data Centre as CCDC694467, CCDC689924, and CCDC689378, re-
CH2Cl2 (10 ml, three times). The combined organic extract was dried spectively. These data can be obtained free of charge via http://
over anhydrous sodium sulfate and evaporated under reduced www.ccdc.cam.ac.uk/data_request/cif, or by e-mailing data_
pressure to give a crude extract, which was further purified by col- [email protected], or by contacting the Cambridge Crystal-
umn chromatography over silica gel eluting with 5% acetone–hex- lographic Data Centre, 12, Union Road, Cambridge CB2 1EZ, UK; fax:
ane to yield the dibrosylate 20 (75.2 mg) (Rf (5% acetone–hexane 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- References and notes
negative bacteria: B. subtilis, S. aureus, TISTR517, E. faecalis
1. Smitinand, T. Thai Plant Names; Prachachon: Bangkok, 2001; p 152.
TISTR459, Methicillin-Resistant S. aureus (MRSA) ATCC43300,
2. Vo, V. V. A Dictionary of Medicinal Plants in Vietnam; Y Hoc: HoChiMinh City,
Vancomycin-Resistant E. faecalis (VRE) ATCC 51299, Streptococcus 1997; Vol. 435.
faecalis, S. typhi, S. sonei and P. aeruginosa. The microrganisms were 3. Boonnak, N.; Karalai, C.; Chantrapromma, S.; Ponglimanont, C.; Hoong-Kun, F.;
obtained from the culture collections, Department of Industrial Kanjana-Opas, A.; Laphookhieo, S. Tetrahedron 2006, 62, 8850–8859.
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