Ring B aromatic norpimarane glucoside from a Xylaria sp.
ABSTRACT A novel 20-norpimarane glucoside, xylopimarane (1), together with the known sphaeropsidin C (2) and clonostachydiol (3), was isolated from the fungus Xylaria sp. BCC 4297. Compound 1 exhibited cytotoxicity to cancer cell lines KB, MCF-7, and NCI-H187 with respective IC(50) values of 1.0, 13, and 65 μM.
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ABSTRACT: The biological control of cyanobacterial harmful algal blooms (cyanoHABs) is important to promote human health, environmental protection, and economic growth. Active algicidal compounds and algicidal mechanisms should be identified and investigated to control cyanoHABs. In this study, the algicidal actinobacterium Streptomyces sp. L74 was isolated from the soil of a nearby pond which located in the center lake of Guanghzou Higher Education Mega Center. Results showed that the algicidal activities of cyanoHABs are mainly achieved via an indirect attack by producing algicidal compounds. All active algicidal compounds are hydrophilic substances that are heat and pH stable. In the present study, an active compound (B3) was isolated and purified by high-performance liquid chromatography and identified as a type of triterpenoid saponin (2-hydroxy-12-oleanene-3, 28-O-D-glucopyranosyl) with a molecular formula of C42H70O13 as determined by infrared spectrometry, electrospray ionization mass spectrometry, and nuclear magnetic resonance. Active algicidal compounds from Streptomyces sp. L74 were shown to disrupt the antioxidant systems of Microcystis aeruginosa cells.PLoS ONE 01/2013; 8(10):e76444. · 3.73 Impact Factor
Published: January 12, 2011
Copyrightr2011 American Chemical Society and
American Society of Pharmacognosy
dx.doi.org/10.1021/np100873t|J. Nat. Prod. 2011, 74, 300–302
Ring B Aromatic Norpimarane Glucoside from a Xylaria sp.
Masahiko Isaka,* Arunrat Yangchum, Patchanee Auncharoen, Kitlada Srichomthong, and
National Center for Genetic Engineering and Biotechnology (BIOTEC), 113 Thailand Science Park, Phaholyothin Road,
Klong Luang, Pathumthani 12120, Thailand
S Supporting Information
ABSTRACT: A novel 20-norpimarane glucoside, xylopimarane (1), together with the known
Compound 1 exhibited cytotoxicity to cancer cell lines KB, MCF-7, and NCI-H187 with
respective IC50values of 1.0, 13, and 65 μM.
from Xylaria sp. (No. 2508),1,2multiplolides from X. multiplex
BCC 1111,3xylactam from X. euglossa,4integric acid from a
the utilization of fungal sources in Thailand, we investigated the
Xylaria sp. strain BCC 4297, as an extract of this fungus showed
up fermentation and chemical studies resulted in the isolation of
a novel 20-norpimarane glucoside, xylopimarane (1), along with
the known sphaeropsidin C (2)8and clonostachydiol (3).9,10
ungi belonging to the genus Xylaria have been the source of
bioactive compounds such as xyloketals and xyloallenolide
The molecular formula of 1 was determined by HRESIMS as
C25H36O8. The IR spectrum exhibited absorption bands at νmax
3426 and 1636 cm-1, which suggested the presence of OH
groups and an aromatic chromophore. The1H and13C NMR,
DEPT135, and HMQC spectroscopic data demonstrated that 1
was composed of a sugar unit (C6) and aglycone (C19). The
pyranose ring (C-10-C-60) was addressed by COSY data. The
aglycone (C19) contained six aromatic quaternary carbons, a vinyl
group (δC147.6/δH6.25; δC110.2/δH5.02 and 4.96), a hydro-
4.7 Hz), two quaternary carbons, five methylenes, and three methyl
groups. The planar structure of the aglycone was deduced from
correlations to each other and also exhibited correlations to a
quaternary carbon at δC34.1 (C-4), an aromatic quaternary carbon
atδC131.7 (C-5), anda methylene carbonat δC42.0 (C-3), which
indicated the connections of these methyl groups to the same
quaternary carbon (C-4). HMBC data also demonstrated that the
(C-15/C-16), a methyl group at δC19.8 (δH0.79, CH3-17), a
hydroxylated methine (CH-14), and a methylene carbon at δC25.8
(CH2-12).Thepresenceofafullysubstitutedbenzene(ring B) was
H2-2, and HR-11 (δH2.60) to C-10 (δC132.6), from H2-1, H2-3,
H3-18, and H3-19 to C-5 (δC131.7), from H2-1, H2-11, and Hβ-12
(δC129.6), and from H-14 to C-7 (δC143.2). The quaternary
Received:December 1, 2010
dx.doi.org/10.1021/np100873t |J. Nat. Prod. 2011, 74, 300–302
Journal of Natural Products
to C-6 to constitute a benzene ring. The chemical shifts of C-6 and
Therefore, the planar structure of the aglycone was assigned as 20-
norpimara-5,7,9-triene-6,7,14-triol, wherein either 6-OH or 7-OH
should form a glucoside. HMBC correlation from the anomeric
co-occurrence of 1 with the known sphaeropsidine C (2) and the
that these compounds should have the same sense of C-13 absolute
configuration. Intense NOESY correlations of H-14 and H3-17 and
orientation of H-14. Assignment of the protons of two methylene
correlations from H3-17 to Hβ-11 (δH2.43) and Hβ-12 (δH1.50).
Finally, the D-glucose unit was confirmed by acid hydrolysis of 1.
To our knowledge, this is the first report of a natural ring B
reported the isolation of LL-S491β (4) and LL-S491γ (7β-hydroxy
analogue of 4) from the fungus Aspergillus chevalieri.11,12The paper
also describes that ethanolic hydrochloric acid treatment of 4
gave decarboxylation/aromatization products 5a and 5b.11Since
5a is identical to the aglycone of 1, we assume that the same
transformation occurred during fermentation of BCC 4297. Thus,
compound 4, produced by oxidation of sphaeropsidin C (2), is the
possible biosynthetic precursor of 1. Glucoside formation should be
the last biosynthetic step.
Xylopimarane (1) exhibited cytotoxic activity against the human
cancer cell lines KB (oral carcinoma), MCF-7 (breast cancer), and
NCI-H187 (small-cell lung cancer), with IC50values of 1.0, 13, and
65 μM, respectively. It also showed cytotoxicity (IC5041 μM) to
General Experimental Procedures. Melting points were mea-
sured with an Electrothermal IA9100 digital melting point apparatus.
Optical rotations were measured with a JASCO P-1030 digital polarimeter.
UV spectra were recorded on a GBC Cintra404 spectrophotometer. FTIR
spectra were taken on a Bruker VECTOR 22 spectrometer. NMR spectra
were recorded on Bruker DRX400 and AV500D spectrometers. ESITOF
mass spectra were measured with a Bruker micrOTOF mass spectrometer.
Fungal Material. The fungus used in this study was isolated from
an unidentified dead wood in Hala Bala Wildlife Sanctuary, Narathiwat
on February 24, 2000, as BCC 4297. On the basis of the characteristics of
fruiting bodies and morphology, this fungus was assigned to the genus
Xylaria (family Xylariaceae) by one of the authors (P.S.).
Fermentation and Isolation. The fungus BCC 4297 was main-
tained on potato dextrose agar at 25 ?C. The agar was cut into small plugs
and inoculated into 2 ? 250 mL Erlenmeyer flasks containing 25 mL of
incubation at 25 ?C for 8 days on a rotary shaker (200 rpm), each primary
same liquid medium (PDB) and incubated at 25 ?C for 8 days on a rotary
shaker (200 rpm). These secondary cultures were pooled, each 25 mL
of malt extract broth (MEB; malt extract 6.0 g/L, yeast extract 1.2 g/L,
EtOAc (3 ? 5 L), and the combined organic layer was concentrated to
obtain a brown gum (834 mg; extract A). The mycelium was macerated in
added to the filtrate, and the layers were separated. The aqueous MeOH
phase was partially evaporated, and the residue was extracted with EtOAc
(2 ? 600 mL). The combined EtOAc solution was washed with H2O
(183 mg, extract B). Extract A was subjected to Sephadex LH-20 column
chromatography (3.0 ? 50 cm, MeOH) to obtain six pooled fractions,
A1-A6. Fraction A3 (545 mg) was chromatographed on silica gel (3.0 ?
11 fractions, A3-1-A3-11. Fraction A3-4 (58 mg) was purified by pre-
parative HPLC using a reversed-phase column (SunFire Prep C18OBD,
19 ? 150 mm, 5 μm; mobile phase MeCN/H2O, gradient from 30:70 to
Table 1. NMR Spectroscopic Data for 1 in Acetone-d6
(500 MHz for1H, and 125 MHz for13C)
δH, mult. (J in Hz) HMBC
2, 3, 5, 10
1, 4, 10
1, 2, 4, 5, 18, 19
R 2.60, dd (17.3, 6.8)
β 2.43, m
R 2.21, m
β 1.50, dd (12.9, 7.0)
8, 9, 10, 12, 13
8, 9, 10, 12
11, 13, 14, 17
9, 11, 13, 14, 15, 17
4.49, d (4.7)
6.25, dd (17.7, 10.9)
5.02, dd (17.7, 1.4)
4.96, dd (10.9, 1.4)
9.47, br s
4.20, d (4.7)
5.01, d (3.8)
3.91, t (9.3)
3.45, t (9.4)
3.98, br d (11.6); 3.74, m
7, 8, 9, 12, 13, 15, 17
12, 13, 14, 17
12, 13, 14, 15
3, 4, 5, 19
3, 4, 5, 18
aThe assignment of CH3-18 and CH3-19 can be interchanged.
8, 13, 14
7, 30, 50
30, 50, 60
dx.doi.org/10.1021/np100873t |J. Nat. Prod. 2011, 74, 300–302
Journal of Natural Products
80:20,flowrate10mL/min) to furnish2 (7.7mg).FractionA3-6 (91 mg)
was also further fractionated by preparative HPLC (MeCN/H2O, gradient
from 20:80 to 40:60) to afford 3 (20.0 mg). Fraction A4 (97 mg) was sub-
jected to silica gel column chromatography (2.0 ? 15 cm, MeOH/CH2Cl2,
step gradient elution from 2:98 to 11:89) to give 1 (13.2 mg). Extract B was
chromatographed on Sephadex LH-20 (2.0 ? 60 cm, MeOH) and pre-
parative HPLC (MeCN/H2O) to furnish 3 (1.1 mg) and 1 (6.0 mg).
Xylopimarane (1): colorless solid; mp 174-175 ?C; [R]26Dþ75
(c 0.11, MeOH); UV (MeOH) λmax(log ε) 207 (4.66), 228 sh (3.95),
288 (3.44) nm; IR (KBr) νmax3420, 2928, 1636, 1429, 1029, 1008 cm-1;
1H NMR (500 MHz, acetone-d6) and13C NMR (125 MHz, acetone-d6)
data,seeTable1;HRMS(ESI-TOF) m/z487.2301[Mþ Na]þ(calcdfor
Hydrolysis of 1. Compound 1 (1.9 mg) was hydrolyzed in 5%
aqueous hydrochloric acid (0.5 mL) at 90 ?C for 2 h. The mixture was
washed with Et2O (2 ? 1 mL), and the aqueous layer was concentrated
under reduced pressure to obtain D-glucose (0.7 mg,1H NMR in D2O/
CD3OD); [R]27Dþ85 (c 0.035, H2O)).
Biological Assays. Anticancer activities against KB cells (oral
human epidermoid carcinoma), MCF-7 cells (human breast cancer),
and NCI-H187 cells (human small-cell lung cancer) were evaluated
using the resazurin microplate assay.13The IC50values of the standard
compound doxorubicin hydrochloride were 0.27 μM for KB cells,
4.9 μM for MCF-7 cells, and 0.13 μM for NCI-H187 cells. Cytotoxicity
using the green fluorescent protein microplate assay (GFPMA).14
Ellipticine was used as the standard of cytotoxicity (IC507.3 μM).
is available free of charge via the Internet at http://pubs.acs.org.
Financial support from the Bioresources Research Network,
National Center for Genetic Engineering and Biotechnology
(BIOTEC), is gratefully acknowledged.
(1) Lin, Y.; Wu, X.; Feng, S.; Jiang, G.; Luo, J.; Zhou, S.; Vrijmoed,
2001, 66, 6252–6256.
(2) Lin, Y.; Wu, X.; Feng, S.; Jiang, G.; Zhou, S.; Vrijmoed, L. L. P.;
Jones, E. B. G. Tetrahedron Lett. 2001, 42, 449–451.
(3) Boonphong, S.; Kittakoop, P.; Isaka, M.; Pittayakhajonwut, D.;
Tanticharoen, M.;Thebtaranonth, Y.J.Nat.Prod.2001,66,6252–6256.
(5) Singh, S. B.; Zink, D.; Polishook, J.; Valentino, D.; Shafiee, A.;
Silverman, K.; Felock, P.; Teran, A.; Vilella, D.; Hazuda, D. J.; Lingham,
R. B. Tetrahedron Lett. 1999, 40, 8775–8779.
(6) Smith, C. J.; Morin, N. R.; Bills, G. F.; Dombrowski, A. W.;
(7) Isaka, M.; Chinthanom, P.; Boonruangprapa, T.; Rungjindamai,
N.; Pinruan, U. J. Nat. Prod. 2010, 73, 683–687.
(8) Evidente, A.; Sparapano, L.; Fierro, O.; Bruno, G.; Giordano, F.;
Motta, A. Phytochemistry 1997, 45, 705–713.
(9) Grabley, S.; Hammann, P.; Thiericke, R.; Wink, J. J. Antibiot.
1993, 46, 343–345.
(10) Rao,A.V. R.;Murthy, V. S.; Sharma,G.V. M.Tetrahedron Lett.
1995, 36, 143–146.
(11) Ellestad, G. A.; Kunstmann, M. P.; Mirando, P.; Morton, G. O.
J. Am. Chem. Soc. 1972, 94, 6206–6208.
(12) The same compound (4) was later isolated from the fungus
Sphaeropsis sapinea f. sp. cupressi and named sphaeropsidin A: Evidente,
A.; Sparapano, L.; Motta, A.; Giordano, F.; Fierro, O.; Frisullo, F.
Phytochemistry 1996, 42, 1541–1546.
(13) O’Brien, J.; Wilson, I.; Orton, T.; Pognan, F. Eur. J. Biochem.
2000, 267, 5421–5426.
(14) Changsen, C.; Franzblau, S. G.; Palittapongarnpim, P. Anti-
microb. Agents Chemother. 2003, 47, 3682–3687.