960 Journal of Natural Products, 2009, Vol. 72, No. 5
Notes
1
ganodermanontriol by comparison of ESI-MS, H NMR, and 13C
Table 2. 13C NMR Data (500 MHz) of 1-3 (in CDCl3)
NMR data with those of an authentic sample.21
1
2
3
δC
Ganosinensin C (3) was obtained as a yellow, amorphous powder.
The molecular formula of 3 was determined as C51H74O8 by
HRFABMS (m/z 813.5333 [M - H]-, calcd for C51H71O9,
813.5305). The 1H NMR spectrum was similar to that of compound
2 except for some of the signals for a partial farnesyl hydroquinone
moiety (Table 1). The absence of a carbonyl signal (δC 195.5) and
a conjugated carbonyl absorption (1640 cm-1) in the 13C NMR and
IR spectra of 3, compared to those of 2, and the difference in
molecular formula suggested that a farnesyl acid is present in 3,
instead of an oxofarnesyl acid as in 2. This was further confirmed
from the HMBC spectrum, in which three olefinic protons at δH
6.59 (H-5′′), 6.62 (H-3′′), and 6.68 (H-6′′) showed long-range
correlations with C-1′′ (δC 128.0) and C-4′′ (δC 151.2), while the
signal at δH 6.62 (H-3′′) showed long-range correlations with a
carbon signal at δC 31.8 (C-1′), indicating an R-methylene hydro-
quinone moiety as a partial structure (Figure 1b).25 The configu-
ration of 3 was determined by the same method as that for 1. The
structure of the farnesyl hydroquinone moiety matched that of
ganomycin A. Accordingly, ganosinensin C (3) was determined to
be an ester of ganomycin A with ganodermanontriol.
position
δC
δC
1
2
3
4
5
6
7
8
36.7
34.9
216.9
47.5
50.7
23.7
119.4
142.7
144.3
37.2
117.2
37.7
43.8
50.3
27.8
31.5
50.9
15.8
22.1
36.5
18.7
33.5
29.8
77.2
73.9
69.6
20.3
25.4
22.5
25.5
195.3
126.6
146.3
34.6
36.6
34.8
217.0
47.5
50.8
23.6
119.4
142.8
144.4
37.2
117.3
37.7
43.7
50.3
27.8
31.5
50.8
15.7
22.0
36.4
18.5
33.1
25.8
78.8
73.6
66.5
18.1
25.3
22.5
25.4
195.5
127.8
145.2
34.5
36.6
34.8
217.0
47.5
50.8
23.6
119.4
142.7
144.4
37.1
117.2
37.7
43.7
50.2
27.8
31.4
50.7
15.7
22.0
36.5
18.5
33.1
25.7
76.8
73.5
68.7
18.3
25.3
22.4
25.4
31.8
140.6
133.3
35.9
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
1′
Since these triterpene-farnesyl hydroquinone conjugates were
detected in the methanol extract obtained under mild conditions,
they must be natural products rather than artifacts formed through
extraction processes. However, we cannot exclude a possibility of
transesterification between 1 and 2.
Experimental Section
General Experimental Procedures. Optical rotations were measured
with a JASCO DIP-360 automatic polarimeter. UV spectra were
measured with a UV-2200 UV-vis recording spectrophotometer
(Shimadzu Co., Kyoto, Japan). IR spectra were measured with a JASCO
FT/IR-230 infrared spectrometer. NMR spectra were measured with a
Varian Unity 500 (1H, 500 MHz; 13C, 125 MHz) NMR spectrometer.
TMS was used as an internal standard, and J values were reported in
hertz. HRFABMS were measured on a JEOL JMS-AX505HAD
apparatus with a resolution of 5000 and with m-nitrobenzyl alcohol as
matrix. Preparative HPLC was performed on a Tosoh CCPM- II system
(Tosoh Co., Tokyo, Japan) equipped with a UV 8020 detector and a
Cholester Waters HPLC column (20 × 250 mm). Column chromatog-
raphy was carried out on BW-820MH silica gel (Fuji Silica Chemical
Co. Aichi, Japan) or Wakogel 50C18 (38-63 µm, Wako Pure Chemical
Industries, Ltd.).
Plant Material. Ganoderma sinense Zhao, Xu et Zhang was
purchased at Mei-Lingzhi in Hehuachi crude drug market for medicinal
herbs (Chengdu, China) on August 2007 and identified by Dr. De-
Yuan Chen, Guiyang College of Traditional Chinese Medicine,
Guiyang, China. A voucher specimen (TMPW No. 25617) is deposited
at the Museum of Materia Medica, University of Toyama, Japan.
Extraction and Isolation. Dried and chopped G. sinense (2.5 kg)
was extracted with CHCl3 (each 10 L) by refluxing for 9 h (3 h × 3
times) to give 61 g of a crude extract. The crude extract (60 g) was
applied to a silica gel column (6 × 60 cm) and eluted with
hexane-acetone (9:1; Fr. 1, 18.3 g), hexane-acetone (7:3; Fr. 2,
16.7 g), and CHCl3-MeOH (1:2; Fr. 3, 16.2 g). Fr. 2 was subjected to
silica gel column chromatography with a CHCl3-MeOH gradient
solvent system (100:0 f 0:100) to yield six fractions. Fr. 2-2 was
crystallized from MeOH to afford ganoderiol A (40 mg). Fr. 2-4 was
crystallized from hexane to yield ganodermanontriol (1.2 g). Fr. 2-6
was subjected to ODS column chromatography with a H2O-MeOH
gradient solvent system to provide 1-rich (2-6-2a, 800 mg) and 2-rich
fractions (2-6-2b, 1 g). Further separation by preparative HPLC
(Cholester column; MeOH-0.1% TFA/H2O, 5 mL/min) of Fr. 2-6-2a
and Fr. 2-6-2b (H2O-MeOH, 15:85) gave compounds 1 (40 mg) and
2 (50 mg). Fr. 3 was subjected to silica gel column chromatography
with a CHCl3-MeOH gradient solvent system (100:0 f 0:100) to give
nine fractions. Then, Fr. 3-6 was combined and subjected to ODS
column chromatography with a H2O-MeOH gradient solvent system
to provide three fractions. Further separation by preparative HPLC
(cholester column; MeOH-0.1% TFA/H2O, 5 mL/min) of Fr. 3-6-2
2′
3′
4′
5′
25.5
25.6
28.5
6′
122.1
136.6
39.1
122.1
136.9
39.0
124.6
136.7
38.9
7′
8′
9′
25.5
25.6
27.3
10′
11′
12′
13′
14′
15′
1′′
2′′
3′′
4′′
5′′
6′′
125.3
134.4
68.6
13.9
16.3
168.3
157.0
114.9
117.2
148.1
126.1
119.8
125.5
134.5
68.7
13.8
16.2
169.0
157.1
112.7
117.3
148.2
126.0
119.8
126.8
135.8
69.0
13.7
16.2
172.3
128.0
149.3
114.8
151.2
116.9
117.8
(MeOH-H2O, 85:15) gave compound 3 (40 mg), and that of Fr. 3-6-1
(H2O-MeOH, 30:70) yielded ganoderiol D (30 mg).
Ganosinensin A (ganodermanontriol 26-O-{(2Z,5E,9E)-2-[2-(2,5-
dihydroxyphenyl)-2-oxo-ethylidene]-11-hydroxy-6,10-dimethylun-
deca-5,9-dienate}, 1): yellow, amorphous powder; [R]23D +19.1 (c 0.46,
CHCl3); UV (CHCl3) λmax (log ꢀ) 246 (4.3), 254 (4.2), 380 (3.4) nm;
IR (KBr) νmax 3450, 2930, 2850, 1720, 1700, 1650, 1460, 1380, 1300,
1230, 1180, 1110, 1070, and 1000 cm-1; H and 13C NMR (CDCl3)
1
data, see Tables 1 and 2; HRFABMS, m/z 827.5102 [M - H]- (calcd
for C51H71O9, 827.5104).
Ganosinensin B (ganodermanontriol 24-O-{(2Z,5E,9E)-2-[2-(2,5-
dihydroxyphenyl)-2-oxo-ethylidene]-11-hydroxy-6,10-dimethylun-
deca-5,9-dienate}, 2): yellow, amorphous powder; [R]23D +22.1 (c 0.63,
CHCl3); UV (CHCl3) λmax (log ꢀ) 246 (4.4), 254 (4.3), 377 (3.5) nm;
IR (KBr) νmax 3450, 2930, 2850, 1720, 1700, 1640, 1480, 1460, 1370,
1300, 1230, 1190, 1120, 1050, and 1000 cm-1 1H and 13C NMR
;
(CDCl3) data, see Tables 1 and 2; HRFABMS, m/z 827.5094 [M -
H]- (calcd for C51H71O9, 827.5104)
Ganosinensin C (ganodermanontriol 24-O-{(2Z,5E,9E)-2-[2-(2,5-
dihydroxyphenyl)ethylidene]-11-hydroxy-6,10-dimethylundeca-5,9-