1
072
J.-G. Xing et al.
(12 nm, S-50 um, YMC Co., Ltd., Kyoto,
Japan).
followed by 30–40% for 18 min; detection
2
1
at 254 nm; t ¼ 12 min; 2.0 ml min ) to
R
afford compound 2 (10 mg). Fraction 9
(1.6 g) was then subjected to silica gel
reverse-phase column chromatography,
3
.2 Strain isolation and cultivation
Strain L421 was isolated from the stems of
B. gymnorrhiza collected in Dongzhai,
Hainan Province of China. The strain L421
was identified as P. heterocornis by Prof.
Jing-Ze Zhang and has been deposited in
Key Laboratory of Medicinal Chemistry
and Molecular Diagnosis of Ministry of
Education of Hebei University.
eluted with H O–MeOH (100:0–0:100),
2
yielding 10 subfractions (1–10). Com-
pound 3 (15 mg) was obtained as a
colorless crystal from subfraction 2.
3.3.1 Compound 2
2
2
A yellow oil, ½aꢀ 2 64 (c ¼ 0.25,
D
The fungal strain was cultured on
slants of comprehensive potato dextrose
agar (CPDA) at 288C for 5 days. Agar
plugs were used to inoculate 1000-ml
Erlenmeyer flasks each containing 600 ml
of media (CPDA), and the final pH of the
media was adjusted to 6.5 before steriliza-
tion. Flask cultures were incubated at 288C
on a rotary shaker at 150 rpm for 5 days.
Fermentation was carried out in eighty
MeOH). UV (MeOH) l
(nm): 216,
max
21
247, and 292. IR (KBr) vmax (cm ): 3374,
13
1
1744, 1605, and 1441. For H and
C
NMR spectral data, see Table 1. HR-ESI-
þ
MS: m/z 377.1215 [M þ Na] (calculated
for C H O Na, 377.1207).
17 22 8
3
.3.2 Compound 3
A colorless crystal, MP 169–1718C.
2
2
D
5
1
00-ml Erlenmeyer flasks each containing
00 g of rice. Distilled H O (100 ml) was
½aꢀ þ 22.8 (c ¼ 0.1, MeOH). UV
2
(MeOH) l
max
(nm): 212, 249, and 291.
2
IR (KBr) vmax (cm ): 3374, 1736, 1513,
1
added to each flask, and the contents were
soaked overnight before autoclaving at 15
1 13
and 1452. For H and C NMR spectral
2
2
lb in for 30 min. After cooling to room
temperature, each flask was inoculated
with 10.0 ml of the spore inoculum and
incubated at 288C for 60 days.
data, see Table 1. HR-ESI-MS: m/z
þ
393.1156 [M þ Na] (calculated for
C H O Na, 393.1156).
17 22 9
Acknowledgements
3
.3 Extraction and isolation
This work was financially supported by
Hundred Excellent Innovation Talents from
the Universities and Colleges of Hebei
Province (SPRC008), the Key Project of
Chinese Ministry of Education and the Key
Applied Basic Research Programs of Hebei
Province (0996030917D) and National Natural
Science Foundation of China (31071701).
The fermented material was extracted with
EtOAc (8 £ 1.0 l), and the organic solvent
was evaporated to dryness under vacuum
to afford the crude extract (108.0 g). The
extract was applied on a silica gel column
chromatography, and eluted with pet-
roleum ether–EtOAc–MeOH gradient to
afford 10 fractions. Compound 1 (20 mg)
was obtained as a colorless crystal from
fraction 2 (230 mg). Fraction 6 (2.0 g) was
subjected to silica gel column chromato-
References
[1] A.A.L. Gunatilaka, J. Nat. Prod. 69, 509
(
2006).
2] G.A. Strobel, Microbes Infect. 5, 535
2003).
3] B. Schulz, C. Boyle, S. Draeger, A.K.
Rommert, and K. Krohn, Mycol. Res. 106,
996 (2002).
[
[
graphy, eluted with CHCl –MeOH
3
(
(100:0–0:100), yielding seven subfrac-
tions (1–7). Subfraction 3 was purified by
HPLC (30% CH OH in H O for 2 min,
3
2