全 文 ：· 1688 · 药学学报 Acta Pharmaceutica Sinica 2013, 48 (11): 1688−1691




Metabolites from the endophytic fungus Penicillium
sp. FJ-1 of Ceriops tagal
JIN Peng-fei1, 2, ZUO Wen-jian2, GUO Zhi-kai2, MEI Wen-li2*, DAI Hao-fu2*
(1. College of Environment and Plant Protection, Hainan University, 570228 Haikou, China;
2. Key Laboratory of Biology and Genetic Resources of Tropical Crops, Ministry of Agriculture,
Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, 571101 Haikou, China)
Abstract: To investigate the chemical constituents of the endophytic fungus Penicillium sp. FJ-1 of Ceriops
tagal, the chemical constituents were isolated by column chromatography on silica gel and Sephadex LH-20.
Their structures were elucidated on the basis of spectroscopic analysis. Their antibacterial activity was tested
by paper disco diffusion method. Two compounds were isolated and identified as 7-hydroxy-deoxytalaroflavone
(1), and deoxytalaroflavone (2). Compound 1 is a new compound, and compounds 1 and 2 showed weak
activity against Staphylococcus aureus and methicillin-resistant Staphylococcus aureus.
Key words: Ceriops tagal; endophytic fungus; Penicillium sp.; antimicrobial activity
CLC number: R284 Document code: A Article ID: 0513-4870 (2013) 11-1688-04
角果木内生青霉菌 FJ-1化学成分研究
靳鹏飞 1, 2, 左文健 2, 郭志凯 2, 梅文莉 2*, 戴好富 2*
(1. 海南大学环境与植物保护学院, 海南 海口 570228; 2. 中国热带农业科学院热带生物技术研究所,
农业部热带作物生物学与遗传资源利用重点实验室, 海南 海口 571101)

摘要: 采用硅胶、葡聚糖凝胶柱色谱等方法对角果木内生真菌 FJ-1的化学成分进行分离, 从中分离得到了 2
个化合物。通过化合物的理化性质和波谱数据鉴定其结构分别为 7-hydroxydeoxytalaroflavone (1) 和
deoxytalaroflavone (2), 其中化合物 1为新化合物。通过滤纸片扩散法测定其抗菌活性, 化合物 1、2对金葡菌和
耐甲氧西林金葡菌显示有弱抑制活性。
关键词: 角果木; 内生真菌; 青霉属; 抗菌活性

Mangrove plants are woody plants growing in
tropical and subtropical intertidal habitats[1]. Many
secondary metabolites, such as terpenoids, steroids,

Received 2013-06-13; Accepted 2013-07-31.
Project supported by the International Cooperation Projects of Hainan
Province (KJHZ2013-17); National Non-profit
Institute Research Grant of ITBB (ITBB130405);
National Support Science and Technology Subject
(2013BAI11B04); the Natural Science Foundation of
Hainan (313079).
*Corresponding author Tel / Fax: 86-898-66987529,
E-mail: meiwenli@itbb.org.cn;
Tel / Fax: 86-898-66961869,
E-mail: daihaofu@itbb.org.cn
alkaloids and polysaccharides were isolated from
mangrove plants[2−5] and showed anti-tumor, anti-HIV,
anti-bacterial, anti-proliferative, and anti-estrogenic
activities. Ceriops tagal (Rhizophoraceae) are used as
a traditional folk medicine for treatment of wound by
the people of Li folk. The chemical constituents of
Ceriops tagal have been extensively investigated[6, 7].
Previous research has revealed that diterpenoids and
triterpenoids are the mainly constituents in this plant[8−11].
In this study, a new drimane-type sesquiterpene, 7-
hydroxy-deoxytalaroflavone (1), together with a known
compound, deoxytalaroflavone (2), were isolated from
DOI:10.16438/j.0513-4870.2013.11.004
JIN Peng-fei, et al: Metabolites from the endophytic fungus Penicillium sp. FJ-1 of Ceriops tagal · 1689 ·

endophytic fungus FJ-1 and identified by extensive
spectroscopic analysis (1D and 2D NMR). Compounds
1 and 2 showed weak activity against Staphylococcus
aureus and methicillin-resistant Staphylococcus aureus.

Results and discussion
Compound 1 was obtained as brown powder. Its
molecular formula is assigned to be C14H14O5 from its
HR-ESI-MS peak at m/z 285.074 4 [M+Na]+ (calculated
for C14H14O5Na 285.073 9) and NMR data (Table 1), with
eight degrees of unsturation. The IR spectrum displayed
the presence of hydroxyls (3 418 cm−1), carbonyl (1 740
cm−1), and double bond (1 656 cm−1). The 1H NMR
spectrum of 1 (Table 1) displayed the presence of
two aromatic protons [δH 6.53 (1H, d, J = 2.3 Hz, H-4),
6.56 (1H, d, J = 2.3 Hz, H-6)], for one typical 1, 3, 4, 5-
tetrasubstituted phenyl, and a proton signals of methyl
δH 3.94 (3H, s, H3-10). The 13C NMR and DEPT
spectra of 1 (Table 1) displayed a total of 14 carbon
signals including two methyls (one oxygenated), four
methines, one methylene, one carbonyl (at δC 168.9),
and six quaternary carbons (δC 101.7, 101.9, 101.9,
121.1, 166.2 and 168.9) indicated the presence of
one phenyl group. The 1H and 13C NMR spectra of 1
(Table 1) is very similar to that of the known compound
2. The major difference in the 13C NMR spectra data
is the loss of the carbonyl signal (C-7, δC 195.5), and
appearance of an additional oxygenated methane (δC
72.9) in compound 1. The α-orientation of H-7 and
CH3-11 were deduced by the key correlation between
H-7 and CH3-11 in the NOESY spectrum. Thus the
structure of compound 1 was established as shown
in Figure 1, 1, 6-dihydroxy-8-methoxy-3-methyl-2, 3-
dihydrocyclopenta[c]isochromen-5(1H)-one, named 7-
hydroxyl-deoxytalaroflavone.


Figure 1 Key 1H-1H COSY, HMBC, and NOSEY correlations
of compound 1

The antimicrobial test demonstrated that compound
1 and 2 possessed inhibitory effect on S. aureus with the
diameter of the inhibition zone 8.34 mm and 7.06 mm
with the positive control (kanamycin sulfate) 26.05 mm.
Compound 1 possessed inhibitory effect on methicillin-
resistant S. aureus with the diameter of the inhibition
zone 6.24 mm with the positive control (kanamycin
sulfate) 20.66 mm, while compound 2 shows no inhibitory
activity on methicillin-resistant S. aureus.

Experimental
The NMR spectra were recorded on a Bruker AV-
500 spectrometer, using TMS as an internal standard.
The HR-ESI-MS spectra were measured with an API

Table 1 1H (500 MHz) and 13C (125 MHz) NMR spectral data of compound 1 (J in Hz, CD3COCD3)
Position 1H NMR 13C NMR HMBC COSY NOESY
2 168.9 (s)
2a 101.7 (s)
3 166.2 (s)
4 6.53 (d, 2.3) 101.9 (d) 2a, 3, 5, 6 10
5 168.9 (s)
6 6.56 (d, 2.3) 101.9 (d) 2a, 4, 5, 7a
6a 139.3 (s)
7 4.92 (m) 72.9 (d) 7a, 9, 9a 8 8, 9, 11
7a 122.1 (s)
8 2.70 (dt, 8.1, 13.9) 41.5 (t) 7, 7a, 9, 9a, 11 7, 9 7, 9
1.59 (dt, 2.6, 13.9)
9 3.20 (m) 34.7 (d) 7, 7a, 8, 9a, 11 8, 11 7, 8
9a 157.2 (s)
10 3.94 (s) 57.3 (q) 5 4
11 1.39 (d, 6.9) 23.1 (q) 7a, 8, 9 9 7
3-OH 11.45 (s) 2a, 3, 4

· 1690 · 药学学报 Acta Pharmaceutica Sinica 2013, 48 (11): 1688−1691

QSTAR Pulsar mass spectrometer (Bruker). The
ESI-MS spectra were measured with a VG Auto-3000
Spectrometer. Column chromatography was performed
with silica gel (60−80, 200−300 mesh, Marine Chemical
Industry Factory, Qingdao, China) and Sephadex LH-20
(Merck, Germany). TLC was preformed with silica
gel GF254 (Marine Chemical Industry Factory, Qingdao,
China). XS-212 Biological microscope (Nanjing
Jiangnan Novel Optics Co., Ltd., Nanjing, China).
The plant of Ceriops tagal was collected in Hainan
province of China, in June 2010, and identified by Prof.
WANG Zhu-nian, Tropical Crops Genetic Resources
Institute, Chinese Academy of Tropical Agricultural
Sciences. A voucher specimen (No. 20100731) was
deposited at the Institute of Tropical Bioscience and
Biotechnology, Chinese Academy of Tropical Agricultural
Sciences, Haikou, China.
Endophytic fungus FJ-1 was isolated from the
stem of Ceriops tagal tree collected in Haikou city,
Hainan province, China. The fungus was deposited at
the Institute of Tropical Bioscience and Biotechnology,
Chinese Academy of Tropical Agricultural Sciences,
and maintained on potato dextrose agar (PDA) slant at
4 ℃. The fungus was identified by analysis of the
ITS region of the rDNA. The sequence data derived
from the fungal strain have been deposited at GenBank
with the accession number DQ365947.1. The blast
result showed (Figure 2) that the sequence is the most
similar (99%) to the sequence of Penicillium strain
HS-A82 (compared with AF245261.1, AF245257.1).
The strain is preserved at Key Laboratory of Biology
and Genetic Resources of Tropical Crops, Ministry
of Agriculture, Institute of Tropical Bioscience and
Biotechnology, Chinese Academy of Tropical Agricultural
Sciences, 571101, Haikou, PR China.
1 Fermentation, extraction, and isolation[12]
The endophytic fungus FJ-1 was cultured on PDA
at room temperature for 7 days. Two pieces of mycelial
agar plugs (0.5 cm × 0.5 cm) were inoculated into 1 L
Erlenmeyer flasks containing 400 mL potato dextrose
broth (PDB). The cultivation was shaken at 120 r·min−1
at room temperature for 7 days, and then kept in still at
room temperature for 21 days. The culture broth (60 L)
was filtered to give the filtrate and mycelia. The
filtrate was evaporated in vacuo to small volume and
then partitioned in succession between H2O and
petroleum ether, EtOAc. The EtOAc solution was
evaporated under reduced pressure to give a crude
extract (8.4 g), which was separated into 10 fractions
on a silica gel column using a step gradient elution
of CHCl3/MeOH (1∶0 → 0∶1). Fr. 5 (2.4 g) was
submitted to chromatography further separated by
Sephadex LH-20 and on silica gel column with CHCl3/
MeOH (10∶1), yielding compound 1 (8.2 mg). Fr. 4
(125.0 mg) was submitted to chromatography on
silica gel column with CHCl3/MeOH (15∶1) as eluent
and further separated by column chromatography over
Sephadex LH-20 with CHCl3/MeOH (1∶1) as eluent,
yielding compound 2 (4.8 mg).
2 Structure elucidation
7-Hydroxy-deoxytalaroflavone (1) Brown powder
(MeOH); [α] 27D +3.6 (c 0.2, MeOH), UV (MeOH) λmax
(nm) (log ε): 195 (2.99); IR (KBr) νmax (cm−1): 3 418,
1 740, 1 656; 1H NMR (500 MHz, CD3COCD3) and
13C NMR (125 MHz, CD3COCD3) spectral data see
Table 1; HR-ESI-MS peak at m/z 285.074 4 [M+Na]+,
calculated for C14H14O5Na, m/z 285.073 9.
Deoxytalaroflavone (2) White powder (CHCl3)
ESI-MS m/z 261.0 [M+H]+. 1H NMR (500 MHz,
CDCl3) δ 11.32 (1H, s, 3-OH), 6.68 (2H, s, H-4, H-6),


Figure 2 Polymeric analysis of strain FJ-1 based on the sequence of ITS
JIN Peng-fei, et al: Metabolites from the endophytic fungus Penicillium sp. FJ-1 of Ceriops tagal · 1691 ·

3.93 (3H, s, H3-10), 3.43 (1H, m, J = 6.7 Hz, H-9), 2.94
(1H, dd, J = 6.5, 19.0 Hz, H-8), 2.30 (1H, dd, J = 1.2,
19.0 Hz, H-8), 1.45 (3H, d, J = 7.0 Hz, H-11); 13C NMR
(125 MHz, CDCl3) δ 195.5 (C-7), 167.1 (C-2), 165.5
(C-5), 165.2 (C-3), 148.4 (C-9a), 144.9 (C-7a), 134.8
(C-6a), 103.5 (C-6), 103.3 (C-4), 101.0 (C-2a), 56.3
(C-10), 43.1 (C-8), 28.6 (C-9), 21.2 (C-11). The data
is agreement with deoxytalaroflavone[13].
3 Antimicrobial activity[14]
Compounds 1 and 2 were tested for antibacterial
activity against S. aureus strains and methicillin-resistant
S. aureus by the filter paper disc agar diffusion method.
The strains were cultured using nutrient agar. 25 μL
(20 mg·mL−1) of the compounds were impregnated on
sterile filter paper discs (6 mm diameter), and then,
aseptically applied to the surface of the agar plates.
10 μL (0.64 mg·mL−1) of kanamycin sulfate was used as
positive control. Then the diameters of inhibition
zones were measured after 24 h.
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