Journal of Natural Products
ARTICLE
CHCl3ꢀMeOH (1:1) for elution, and finally purified by semiprepara-
tive HPLC, to yield 1 (tR 15.5 min, 5 mg). Fraction 5-8 was chromato-
graphed over silica gel, eluted by n-hexaneꢀacetone (1:1), and then
purified over a Sephadex LH-20 column, using CHCl3ꢀMeOH (1:1)
for elution, furnishing γ-mangostin (4, 2 mg). Fraction 5-9 was
chromatographed over silica gel, eluted by n-hexaneꢀacetone (1:1),
and then purified over a column containing Sephadex LH-20, using
CHCl3ꢀMeOH (1:1) for elution, affording β-sitosterol 3-O-β-D-gluco-
pyranoside (7 mg).
365 nm; UV (MeOH) λmax (log ε) 238 (4.38), 271 (4.45), 307
(4.25) nm; IR (dried film) νmax 2925, 1645, 1600, 1469, 1373, 1284,
826 cmꢀ1 1H and 13C NMR data, see Tables 3 and 4; positive
;
HRESIMS m/z 475.2052, calcd for C27H32O6Na, 475.2097.
7-O-Methylcochinchinone A (17): amorphous yellow powder
(n-hexane) showing a purple color under UV light at 365 nm; UV
(MeOH) λmax (log ε) 236 (4.75), 266 (4.79), 316 (4.44) nm; IR (dried
film) νmax 3390, 1651, 1600, 1455, 1372, 1220, 821 cmꢀ1; 1H and 13
C
NMR data, see Tables 1 and 2; positive HRESIMS m/z 485.2355, calcd
for C29H34O5Na, 485.2304.
Cochinchinoxanthone (1): amorphous, yellow powder (n-hexane)
showing a purple color under UV light at 365 nm; [R]20 þ10 (c 0.1,
3,7-Di-O-methylcochinchinone A (18): amorphous, yellow
powder (n-hexane) showing a purple color under UV light at 365 nm;
UV (MeOH) λmax (logε) 238 (4.41), 267 (4.53), 297 (3.96) nm; IR (dried
D
CH2Cl2); UV (MeOH) λmax (log ε) 226 (4.67), 327 (4.59), 346
(4.60) nm; CD (MeOH, nm) λmax (Δε) 213 (ꢀ26.96), 292 (ꢀ2.41),
303.5 (ꢀ2.64), 353 (þ1.97); IR (dried film) νmax 3230, 1744, 1644, 1597,
1461, 1333, 1274, 1165, 815 cmꢀ1; 1H and 13C NMR data, see Tables 1 and
2; positive HRESIMS m/z 419.1468, calcd for C23H24O6Na, 419.1471.
Cochinensoxanthone (2): amorphous, yellow powder (n-hexane)
1
film) νmax 2921, 1641, 1608, 1488, 1469, 1281, 821 cmꢀ1; H and 13C
NMR data, see Tables 1 and 2; positive ESIMS m/z 499.2 [M þ Na]þ;
positive HRESIMS m/z 499.2484, calcd for C30H36O5Na, 499.2460.
Benzoylation of r-Mangostin (3) and Cochinchinone A
(6). To a dried flask (50 mL) equipped with a water condenser and a
magnetic stirrer were added 12.3 mg (0.03 mmol) of R-mangostin (3),
11.0 mg of benzoic acid (0.09 mmol), 6.2 mg of DCC (0.03 mmol), and
5 mL of CH2Cl2. After the mixture was stirred at 40 ꢀC for 4 h, it was
cooled to room temperature and then washed with distilled H2O. The
organic layer was evaporated under reduced pressure, and the residue
separated by silica gel column chromatography, using n-hexane and
acetone (5:1 f 1:1) as eluents, to produce 6-O-benzoyl-R-mangostin
(12, 3.0 mg, 0.0058 mmol, 19.5%).
showing a purple color under UV light at 365 nm; [R]20 þ11 (c 0.1,
D
CH2Cl2); UV (MeOH) λmax (log ε) 242 (4.58), 308 (4.22) nm; CD
(MeOH, nm) λmax (Δε) 206.5 (þ27.43), 228 (þ2.63), 262.5 (þ3.90),
283 (ꢀ2.83); IR (dried film) νmax 3344, 1632, 1616, 1587, 1457, 1226,
1125 cmꢀ1; 1H and 13C NMR data, see Tables 1 and 2; positive HRESIMS
m/z 487.2131, calcd for C28H32O6Na, 487.2097.
Acetylation of r-Mangostin (3) and Cochinchinone A (6).
To a dried 50 mL flask equipped with water condenser and magnetic
stirrer, containing 12.3 mg (0.03 mmol) of R-mangostin (3), were added
acetic anhydride (9.2 mg, 0.09 mmol) and pyridine (5 mL). After the
mixture was stirred at 60 ꢀC for 1 h, the mixture was cooled to room
temperature. Then, 5 mL of CHCl3 was transferred into the flask, and
the solution was extracted with distilled H2O. The organic layer was
washed with distilled H2O and then evaporated at reduced pressure. The
residue was separated by silica gel column chromatography, using n-
hexane and acetone (5:1 f 1:1), to afford 3-O-acetyl-R-mangostin (7,
1.5 mg, 0.003 mmol, 11.0%) and 3,6-di-O-acetyl-R-mangostin (8, 12.0
mg, 0.024 mmol, 81.0%). Using the same procedure, reaction at 70 ꢀC
for 2 h yielded 3,6-di-O-acetyl-R-mangostin (8, 12.5 mg, 0.025 mmol,
84.3%) and 3,6,7-tri-O-acetyl-R-mangostin (9, 2.5 mg, 0.0047 mmol,
12.4%).
The same reaction as described above was performed on 13.4 mg
(0.03 mmol) of cochinchinone A (6) to afford 7-O-benzoylcochinchi-
none A (19, 2.0 mg, 0.0036 mmol, 12.1%).
7-O-Benzoylcochinchinone A (19): amorphous, yellow powder
(n-hexane) showing a purple color under UV light at 365 nm; UV
(MeOH) λmax (log ε) 239 (4.53), 257 (4.36), 313 (4.05) nm; IR (dried
film) νmax 1743, 1645, 1483, 1253, 807 cmꢀ1; 1H and 13C NMR data, see
Tables 3 and 4; positive HRESIMS m/z 575.2432, calcd for
C35H36O6Na, 575.2410.
Cyclization of r-Mangostin (3). To a dried flask (50 mL)
equipped with water condenser and magnetic stirrer, containing 12.3 mg
(0.03 mmol) of R-mangostin (3), were added 1.44 mg of formic acid
(0.09 mmol), 6.18 mg of DCC (0.03 mmol), and 5 mL of CH2Cl2. After
the mixture was stirred at 40 ꢀC for 4 h and cooled to room temperature,
it was washed with distilled H2O. The organic layer was evaporated at
reduced pressure, and the residue was separated by silica gel column
chromatography, using n-hexane and acetone (5:1 f 1:1) as eluents, to
afford 18-O-formyl-3-isomangostin hydrate (13, 1.5 mg, 0.0033 mmol,
11.0%), 3-isomangostin hydrate (14, 3.0 mg, 0.007 mmol, 23.4%), and
1-isomangostin hydrate (15, 1.5 mg, 0.0035 mmol, 11.7%).
Cytotoxicity Assay. Cytotoxicity of the samples was evaluated
against HT-29 human colon cancer cells by a previously reported pro-
cedure.32 The cells were cultured under standard conditions and trypsi-
nized. Then, the harvested cells were added to 96-well plates and treated
by the test samples dissolved in DMSO at different concentrations, the
positive control, and the negative control (DMSO). The plates were
incubated at 37 ꢀC in 5% CO2 for three days, and then the cells were
fixed, incubated at room temperature for 30 min, washed with tap water,
dried at room temperature overnight, and dyed using sulforhodamine B.
After the dyed cells were lysed in Tris-base buffer, the plates were read at
515 nm with an ELISA plate reader. Paclitaxel was used as a positive
control, and the ED50 values of the test samples in serial dilutions were
calculated using nonlinear regression analysis (Table Curve2Dv4; AISN
Software, Inc., Mapleton, OR). Measurements were performed in
triplicate and are representative of two independent experiments in
which the values generally agreed within 10%.
The same reaction as described above with 13.4 mg (0.03 mmol) of
cochinchinone A (6) at 60 ꢀC for 1 h produced 3,7-di-O-acetylcochinch-
inone A (16, 12 mg, 0.022 mmol, 75.2%).
3,7-Di-O-acetylcochinchinone A (16): amorphous, white pow-
der (n-hexane) showing a purple color under UV light at 365 nm; UV
(MeOH) λmax (log ε) 235 (4.57), 260 (4.60), 286 (4.07) nm; IR (dried
film) νmax 2968, 1767, 1635, 1616, 1587, 1484, 1463, 1372, 878 cmꢀ1
;
1H and 13C NMR data, see Tables 3 and 4; positive HRESIMS m/z
555.2416, calcd for C32H36O7Na, 555.2359.
Methylation of r-Mangostin (3) and Cochinchinone A (6).
To a dried flask (50 mL) equipped with a water condenser and a
magnetic stirrer, containing 12.3 mg (0.03 mmol) of R-mangostin (3),
were added 20.8 mg of Ag2O (0.09 mmol), 12.8 mg of MeI (0.09 mmol),
and 5 mL of CH2Cl2. After being stirred at 40 ꢀC for 4 h, the mixture was
cooled to room temperature and washed with distilled H2O. The organic
layer was evaporated at reduced pressure. The residue was separated by
silica gel column chromatography, using n-hexane and acetone (5:1 f 1:1)
for elution, to afford 4-methyl-3,6-di-O-methyl-R-mangostin (10, 2.0
mg, 0.0042 mmol, 14.7%) and 3,6-di-O-methyl-R-mangostin (11, 4.0
mg, 0.009 mmol, 30.4%).
The same reaction performed as described above with 13.4 mg (0.03
mmol) of cochinchinone A (6) and 20.8 mg (0.09 mmol) of Ag2O gave
7-O-methylcochinchinone A (17, 2.0 mg, 0.0043 mmol, 14.4%) and 3,7-
di-O-methylcochinchinone A (18, 1.5 mg, 0.0032 mmol, 10.5%).
4-Methyl-3,6-di-O-methyl-r-mangostin (10): amorphous,
yellow powder (n-hexane) showing a purple color under UV light at
Enzyme-Based ELISA NF-KB Inhibition Assay. A NF-κB
inhibition assay was carried out using a published procedure, with an
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dx.doi.org/10.1021/np200051j |J. Nat. Prod. 2011, 74, 1117–1125