Altenuene derivatives from an unidentified freshwater fungus in the family tubeufiaceae

Article abstract of DOI:10.1021/np0504661

Four new altenuene derivatives called dihydroaltenuenes A (1) and B (2) and dehydroaltenuenes A (3) and B (4), along with five known compounds, including isoaltenuene (5), altenuene (6), and 5′-epialtenuene (7), were isolated from cultures of an unidentif

Full text of DOI:10.1021/np0504661

612
J. Nat. Prod. 2006, 69, 612-615
Altenuene Derivatives from an Unidentified Freshwater Fungus in the Family Tubeufiaceae
Ping Jiao,† James B. Gloer,*,† Jinx Campbell,‡ and Carol A. Shearer‡
Department of Chemistry, UniVersity of Iowa, Iowa City, Iowa 52242, and Department of Plant Biology, UniVersity of Illinois,
Urbana, Illinois 61801
ReceiVed NoVember 11, 2005
Four new altenuene derivatives called dihydroaltenuenes A (1) and B (2) and dehydroaltenuenes A (3) and B (4), along
with five known compounds, including isoaltenuene (5), altenuene (6), and 5′-epialtenuene (7), were isolated from
cultures of an unidentified freshwater aquatic fungal species in the family Tubeufiaceae. The structures of 1-4 were
1
determined by analysis of NMR and MS data. The relative stereochemistry was determined on the basis of H NMR
J-values and NOE data, while the absolute configuration of a representative member of the group (5) was assigned by
CD spectral analysis of its bis-N,N-dimethylaminobenzoate derivative. Compounds 1, 3, 4, 5, and 6 showed antibiotic
activity against Gram-positive bacteria.
Fungi are well known as producers of secondary metabolites that
display a variety of biological activities. However, several ecological
groups of fungi remain underexplored as potential sources of new
bioactive natural products. One such group includes fungi that
specialize in freshwater aquatic habitats. Our research group has
undertaken investigations of freshwater aquatic fungi, and extracts
of such isolates have afforded a variety of bioactive metabolites.1-4
In the course of this work, an unusual isolate identified only as a
member of the family Tubeufiacieae was collected in the Cheoah
River in the Great Smoky Mountains, Tapoco, North Carolina.
Chemical studies of this isolate (A-00471) afforded four new
altenuene derivatives (1-4) and five known compounds. Details
of the isolation, structure elucidation, and stereochemical assignment
of compounds 1-4 are presented here.
antifungal activity against Candida albicans in disk assays and was
selected for further investigation. In addition to 1-4, studies of
this extract afforded five known compounds: isoaltenuene (5),
altenuene(6),5′-epialtenuene(7),talaroflavone,and7-hydroxy-2,5-dimeth-
ylchromone.7-10 Altenuene and isomers thereof have been previ-
ously reported from isolates of the common fungal genus Alternaria.
Altenuene, altertoxin, and alternariol are dibenzopyrone derivatives
that have been reported to show antimicrobial activity and phyto-
toxicity.5,6 The structures of the known compounds were confirmed
by comparison of their NMR and MS data with literature values.7-10
In addition, the relative stereochemical differences that distinguish
diastereomers 5-7 were independently confirmed by analysis of
1H NMR J-values and NOE data for each compound.
The molecular formula of dihydroaltenuene A (1) was determined
to be C15H18O6 (7 unsaturations) on the basis of an EIMS molecular
ion at m/z 294, together with NMR data. Analysis of the NMR
data suggested that 1 was a new altenuene analogue. One hydrogen-
bonded OH signal at δH 11.30, two meta-coupled aromatic proton
signals at δH 6.37 and 6.28, and one methoxy group (δH 3.84) are
all characteristic of the aromatic portion of the altenuenes.7,8
Comparison of the remaining signals with those of 5-7 showed
the absence of the olefinic resonance and the presence of one
additional methine proton signal at δH 3.06 and two methylene
proton signals at δH 2.30 and 1.74, suggesting saturation of the
C1′-C6′ double bond. This conclusion was supported by elucida-
tion of a spin-system corresponding to the C-3′-C6′/C-1′ unit in 1
based on analysis of 1H NMR data. HMBC correlations were
observed from the methine proton at δH 3.06 (H-1′, J ) 13 Hz) to
C-2′, C-6′, and C-1 of the aromatic ring, and the methyl singlet at
δH 1.40 displayed correlations to C-2′ and C-3′. Thus, the gross
structure was established as shown in 1.
The relative stereochemistry of 1 was assigned by analysis of
1
relevant H NMR J-values and NOE data. H-6′â appeared as a
quartet with a coupling constant of 13 Hz, consistent with germinal
and axial-axial couplings for a six-membered ring, requiring axial
orientations for H-6′â, H-1′, and H-5′, and enabling assignment of
the relative stereochemistry shown at C-1′ and C-5′. NOE correla-
tions of the methyl signal at H3-7′ to H-3′â and H-6′â suggested
trans-fusion of the lactone ring and cyclohexane rings (Figure 1)
and an axial orientation for CH3-7′ and H-1′. The equatorial
orientation proposed for H-4′ was consistent with the small vicinal
coupling constants (2.3-3.0 Hz) observed between H-4′ and both
H-3R and H-3â.
Results and Discussion
The isolate was cultured by solid-substrate fermentation on rice.
The EtOAc extract of the resulting cultures exhibited moderate
Dihydroaltenuene B (2) also displayed an EIMS molecular ion
at m/z 294, suggesting that it is an isomer of compound 1. The 1H
and 13C NMR spectra were very similar to those of 1, with the
only significant difference in the 1H NMR spectrum being a
* To whom correspondence should be addressed. Tel: 319-335-1361.
Fax: 319-335-1270.
† University of Iowa.
‡ University of Illinois.
10.1021/np0504661 CCC: $33.50
© 2006 American Chemical Society and American Society of Pharmacognosy
Published on Web 03/17/2006

Altenuene DeriVatiVes from a Freshwater Fungus
Journal of Natural Products, 2006, Vol. 69, No. 4 613
Table 1. 1H NMR Data [δH (mult, JHH)] for Compounds 1-4 (CDCl3, 300 MHz)
position
1
2
3
4
3-OH
4
6
11.30 (s)
11.36 (s)
11.38 (s)
11.32 (s)
6.37 (d, 2.3)
6.28 (d, 2.3)
3.84 (s)
6.35 (d, 2.3)
6.27 (d, 2.3)
3.84 (s)
6.64 (d, 2.3)
6.71 (d, 2.3)
3.87 (s)
6.61 (d, 2.3)
6.69 (d, 2.3)
3.89 (s)
8
1′
3.06 (br d, 13)
2.05 (dd, 14, 2.3)
2.40 (dd, 14, 3.0)
4.22 (br m)
3.57 (dd, 13, 3.4)
2.42 (dd, 13, 4.4)
2.12 (dd, 13, 2.6)
4.16 (m)
3′R
3′â
4′
2.47 (t, 13)
2.83 (dd, 14, 6.0)
2.40 (dd, 14, 7.3)
4.44 (dd, 7.3, 6.0)
2.93 (br s)
2.85 (dd, 13, 6.0)
4.30 (dd, 13, 6.0)
3.55 (br s)
4′-OHa
5′
3.95 (br m)
2.30 (dt, 13, 3.4)
1.74 (q, 13)
1.40 (s)
4.10 (m)
6′R
6′â
7′
2.25 (br d, 13)
1.95 (td, 13, 2.6)
1.39 (s)
6.53 (s)
6.43 (s)
1.73 (s)
1.72 (s)
a Broad OH signals were observed for the 4′-OH in the spectra for 3 and 4, but 4′-OH and 5′-OH signals were not discernible in the spectra of
1 and 2.
Figure 2. Key NOE correlations of dehydroaltenuenes A (3) and
B (4).
signal at δC 197.4. Assignment of the structure as shown in 3 was
enabled by analysis of HMBC data, including key correlations from
H-6 to C-2, C-5, and C-1′; from H-6′ to C-2′, C-4′, and C-1; and
from both H-3′R and H-3′â to the C-5′ carbonyl. The relative
stereochemistry of 3 was assigned by analysis of coupling constants
and NOE data. The signal for H-4′ showed one vicinal coupling
constant (J ) 14 Hz) characteristic of axial-axial-type coupling
and a smaller one (J ) 5.6 Hz) suggestive of axial-equatorial
coupling, leading to the assignment of a pseudoaxial orientation
for H-4′ as shown in Figure 2. Mutual NOE correlations observed
among CH3-7′, H-4′, and H-3′â indicated orientation of all three
of these units on the same face of the molecule.
NMR and MS data indicated that dehydroaltenuene B (4) is an
isomer of 3, and the limited NMR differences (Tables 1 and 2)
again suggested that the difference was in the relative stereochem-
istry at C-5′. The vicinal J-values observed for H-4′ (6.0 and 7.3
Hz) in comparison with those observed for dehydroaltenuene A
(3) suggested a pseudoequatorial orientation of H-4′. These coupling
constants, combined with NOE data (Figure 2), led to assignment
of the relative stereochemistry of 4 as shown.
Figure 1. Key NOE correlations of dihydroaltenuenes A (1) and
B (2).
Table 2. 13C NMR Data (δC) for Compounds 1-4 (CDCl3, 100
MHz)
position
1
2
3
4
1
2
3
4
5
6
7
8
142.4
101.6
165.2
104.1
166.5
99.0
143.6
102.4
164.9
104.2
166.4
98.9
134.6
100.5
165.1
104.3
166.6
105.3
168.0
56.3
135.8
102.6
165.2
105.9
166.9
105.9
167.8
56.3
169.1
56.0
169.1
55.9
1′
2′
3′
4′
5′
6′
7′
42.5
83.3
42.2
69.1
71.2
28.1
21.0
37.7
83.9
40.0
69.6
71.1
27.7
20.9
153.7
81.0
44.4
153.0
80.1
41.1
70.9
68.8
197.8
121.2
26.1
197.4
124.2
29.6
somewhat different splitting pattern ultimately attributed to a
different relative configuration at C-5′. Specifically, the H-5′ and
H-6′â signals were coupled with an axial-equatorial J-value rather
than the trans-diaxial coupling observed in the spectrum for 1. NOE
correlations (Figure 1) and other relevant J-values were consistent
with this being the only difference in the two structures.
To our knowledge, absolute stereochemistry has not been
assigned for altenuene or any of its previously reported stereo-
isomeric analogues. Interestingly, the original reports indicated that
altenuene and 5′-epialtenuene were obtained as racemates,9,11 and
no optical rotation values for the isomers have been reported. In
contrast to these reports, nonzero optical rotations were recorded
for all three known altenuenes 5-7 obtained in this study, as well
as the new analogues 1-4. The absolute stereochemistry of the
most abundant analogue isolated (isoaltenuene; 5) was established
by preparation of the corresponding bis-N,N-dimethylaminobenzoate
and subsequent analysis of its circular dichroism (CD) spectrum.
Initially, dibromobenzoate 5a was prepared. However, the CD
curve obtained for 5a (Figure 3) had an irregular shape and did
Dehydroaltenuene A (3) was assigned the molecular formula
C15H14O6 (9 unsaturations) on the basis of EIMS and NMR data.
1
The H and 13C NMR chemical shifts (Tables 1 and 2) matched
reasonably well with those of the corresponding signals for the
altenuenes and revealed the presence of the same structural features
found in the altenuenes,7,8 except that the oxygenated sp3 methine
carbon C-5′ was replaced by an R,â-unsaturated ketone carbonyl

614 Journal of Natural Products, 2006, Vol. 69, No. 4
Jiao et al.
Figure 3. CD and UV spectra of 5a and 5b in acetonitrile.
and CD spectra (0.1-cm cell) were collected using an Olis Cary-17
spectrometer. 1H and 13C NMR spectra were measured on Bruker DPX-
300 and DRX-400 spectrometers, respectively. HMQC and HMBC data
were measured on a Bruker AMX-600. HPLC was carried out using a
Beckman system Gold HPLC system with a model 166 detector. Other
general procedures and instrumentation have been described previ-
ously.13
Isolation, Cultivation, and Fermentation of Fungal Material.
Species A-00471 was isolated from submerged, decorticated wood
collected from the Cheoah River (fast flowing, 26 °C, pH 5), Tapoco,
North Carolina, on July 18, 2000. Procedures used in collection and
isolation have been described elsewhere.14 On the basis of the
morphological characteristics of this fungus, it belongs in the Dot-
hideomycetes, Pleosporales, and Tubeufiaceae. Due to paucity of
material from the field and absence of sexual reproduction in culture,
the fungus could not be identified further with confidence. Species
A-00471 is characterized by hyaline to tan, superficial, membranous,
broadly conical ascomata; cylindrical fissitunicate asci; hamathecium
of septate pseudoparaphyces; and 1-3 septate, narrowly fusoid, hyaline
ascospores. A voucher specimen has been deposited in the University
of Illinois Department of Plant Biology fungal collection with the
accession number A-00471.
not provide conclusive information, presumably due to interference
with the nearby aromatic ring chromophore8 already present in the
starting material. Thus, the bis-p-N,N-dimethylaminobenzoate (5b)
was prepared. The N,N-dimethylaminobenzoate chromophore is red-
shifted, with absorbance centered at approximately 314 nm,12 and
therefore circumvented the interference problem observed for the
dibromobenzoate. The CD spectrum of 5b (Figure 3) was recorded
in MeCN, and a clear negative bisignate Cotton effect was observed
centered at the dimethylaminobenzoate chromophore λmax. On the
basis of the exciton chirality rule,12 compound 5 was proposed to
have the 2′S, 4′R, and 5′R absolute stereochemistry. Altenuene (6),
5′-epialtenuene (7), and compounds 1-4 are proposed to have the
absolute stereochemistry shown based on presumption of the same
configuration at C-2′.
Although the original extract showed modest antifungal activity,
compounds 1-4 showed no activity in disk assays against
Aspergillus flaVus (NRRL 6541), Fusarium Verticillioides (NRRL
25457), or Candida albicans (ATCC 14053) at 200 µg/disk. In
antibacterial assays, compounds 1 and 4 were active against
Staphylococcus aureus (ATCC 29213), each causing a 14-mm zone
of inhibition at 100 µg/disk (standard ) gentamicin sulfate; 35-
mm zone at 25 µg/disk). Compounds 1, 3, and 4 also showed
activity against Bacillus subtilis (ATCC 6051) at the same level,
affording zones of 50, 13, and 20 mm, respectively (standard )
gentamicin sulfate; 32-mm zone at 25 µg/disk). Interestingly,
compound 2 did not show activity in these assays, despite differing
from 1 only in the relative stereochemistry at C-5′. None of the
compounds showed significant activity against Escherichia coli
(ATCC 25922) at this level. Isoaltenuene (5), altenuene (6), and
5′-epialtenuene (7) were also tested in these assays (at 100 µg/
disk). Altenuene (6) showed activity against S. aureus and B.
subtilis, causing zones of inhibition of 18 and 20 mm, respectively.
Isoaltenuene (5) was active against B. subtilis, causing an inhibition
zone of 30 mm, while 5′-epialtenuene (7) was inactive.
The fungus was subcultured onto 250 g of rice and incubated at 25
°C under 12 h light/12 h dark conditions for 5 weeks. The fermentation
mixture was broken up with a spatula and extracted twice with EtOAc
(2 × 500 mL). The combined EtOAc solution was filtered and
evaporated to afford a crude extract (539 mg).
Extraction and Isolation. The crude extract was partitioned between
hexanes and MeCN. The MeCN fraction (290 mg) was fractionated
on a Sephadex LH-20 column using a hexanes/CH2Cl2/acetone solvent
system to obtain 20 fractions. Fraction 10 (28 mg) was further separated
by reversed-phase HPLC (30% MeCN/H2O isocratic over 10 min, 30-
70% over 15 min, 70-100% over 5 min) on an Alltech HS BDS 8 µm
C18 column (10 × 250 mm) at a flow rate of 2 mL/min with UV
detection at 215 nm to afford 5′-epialtenuene (7; 6.6 mg), talaroflavone10
(3.6 mg), 1 (2.5 mg), and 3 (1.6 mg). Fraction 12 (33 mg) was separated
by gradient elution using MeCN/H2O under the same HPLC conditions
to yield 7-hydroxy-2,5-dimethylchromone10 (2.4 mg) and 4 (1.3 mg).
A subfraction (19 mg) from fraction 12 obtained as a single HPLC
peak was further purified by reversed-phase HPLC employing gradient
elution (50% MeOH/H2O isocratic over 30 min, then 50-100% over
10 min) on the same column with UV detection at 230 nm to afford
altenuene (6; 5.0 mg), isoaltenuene (5; 11 mg), and 2 (1.9 mg).
Experimental Section
General Experimental Procedures. Optical rotations were deter-
mined with a Rudolph automatic polarimeter, model AP III. UV spectra
were recorded with a Varian Cary III UV-visible spectrophotometer,

Altenuene DeriVatiVes from a Freshwater Fungus
Journal of Natural Products, 2006, Vol. 69, No. 4 615
Dihydroaltenuene A (1): colorless glass; [R]25D +20 (c 0.15, CH2-
Preparation of Bis-N,N-dimethylaminobenzoate Ester 5b. In the
same fashion, compound 5 (3.0 mg, 0.010 mmol) was treated with
p-N,N-dimethylaminobenzoyl chloride (3.9 mg, 0.021 mmol) in
CH2Cl2 (1 mL) to give the corresponding 5b (0.7 mg) as a white solid:
UV (MeCN) λmax (log ꢀ) 239 (4.1), 314 (4.4); CD (MeCN) ∆ꢀ 233
(-7.6), 256 (+3.0), 292 (17.3), 333 (-22.6); 1H NMR (300 MHz,
CDCl3) δ 11.37 (s, 3-OH), 6.55 (d, J ) 2.3 Hz, H-6), 6.48 (d, J ) 2.3
Hz, H-4), 6.26 (d, J ) 2.7 Hz, H-6′), 6.03 (dd, J ) 8.4, 2.7 Hz, H-5′),
5.55 (ddd, J ) 13, 8.4, 4.1 Hz, H-4′), 3.84 (s, H-8), 2.65 (dd, J ) 13,
4.1 Hz, H-3′â), 2.47 (t, J ) 13 Hz, H-3′R), 1.76 (s, H-7′); EIMS m/z
586 (M+; 0.1), 438 (1), 164 (100), 148 (76).
1
Cl2); UV (MeOH) λmax (log ꢀ) 305 (3.9); 270 (5.3); H and 13C NMR
data, see Tables 1 and 2; HMBC data, H-4 f C-2, 3, 5, 6; H-6 f C2,
4, 5, 1′; H-8 f C-5; OH-3 f C-2, 3; H-1′ f C-1, 2′, 6′; H-3′ f C-1′,
2′, 4′, 5′; H-4′ f C-2′; H-6′ f C-1′, 2′, 5′; H-7′ f C-2′, 3′; EIMS m/z
294 (M+; 74), 240 (100), 229 (21), 212 (50); HREIMS m/z 294.1109
(calcd for C15H18O6, 294.1103).
Dihydroaltenuene B (2): colorless glass; [R]25 +14 (c 0.085,
D
1
CH2Cl2); UV (MeOH) λmax (log ꢀ) 304 (4.0), 269 (4.3); H and 13C
NMR data, see Tables 1 and 2; EIMS m/z 294 (M+; 100), 279 (20),
240 (80); HREIMS m/z 294.1105 (calcd for C15H18O6, 294.1103).
Dehydroaltenuene A (3): colorless glass; [R]25 +12 (c 0.032,
D
1
CH2Cl2); UV (MeOH) λmax (log ꢀ) 300 (3.7), 253 (4.1); H and 13C
Acknowledgment. Financial support from the National Institutes
NMR data, see Tables 1 and 2; HMBC data, H-4 f C-2, 3, 6; H-6 f
C-2, 4, 5, 1′; H-8 f C-5; OH-3 f C-2, 3, 4; H-3′f C-2′, 4′, 5′, 7′;
H-4′ f C-3′; H-6′ f C-1, 2′, 4′; H-7′ f C-1′, 2′, 3′; EIMS m/z 290
(M+; 26), 245 (80), 217 (100), 202 (13), 174 (29); HREIMS m/z
290.0793 (calcd for C15H14O6, 290.0790).
of Health (GM 60600) is gratefully acknowledged.
Supporting Information Available: 1H NMR spectra for com-
pounds 1-4 and 13C NMR spectra for compounds 1 and 3. This material
is available free of charge via the Internet at http://pubs.acs.org.
Dehydroaltenuene B (4): colorless glass; [R]25 +65 (c 0.026,
D
1
CH2Cl2); UV (MeOH) λmax (log ꢀ) 300 (3.9), 253 (4.2); H and 13C
NMR data, see Tables 1 and 2; EIMS m/z 290 (M+; 14), 246 (25), 217
(100), 175 (55); HREIMS m/z 290.0780 (calcd for C15H14O6, 290.0790).
References and Notes
(1) Reategui, R. F.; Gloer, J. B.; Campbell, J.; Shearer, C. A. J. Nat.
Prod. 2005, 68, 701-705.
Isoaltenuene (5): [R]25 +22 (c 0.54, CH2Cl2); UV (MeOH) λmax
D
(2) Li, C.; Nitka, M. V.; Gloer, J. B. J. Nat. Prod. 2003, 66, 1302-
(log ꢀ) 243 (4.4); 281 (3.9); 325 (3.7); CD (MeCN) ∆ꢀ 220 (-1.6),
256 (-0.5). EIMS, 1H NMR, and 13C NMR data matched those
previously reported.8
1306.
(3) Oh, H.; Swenson, D. C.; Gloer, J. B.; Shearer, C. A. Tetrahedron
Lett. 2001, 42, 975-977.
(4) Oh, H.; Gloer, J. B.; Shearer, C. A. J. Nat. Prod. 1999, 62, 497-
501.
Altenuene (6): [R]25 -7.8 (c 0.205, CH2Cl2); UV (MeOH) λmax
D
(log ꢀ) 240 (4.4); 278 (3.9); 319 (3.7). EIMS, 1H NMR, and 13C NMR
data matched those previously reported.7
(5) King, A. D., Jr.; Schade, J. E. J. Food Prot. 1984, 38, 267-274.
(6) Schade, J. E.; King, A. D., Jr. J. Food Prot. 1984, 47, 978-995.
(7) Turner, C. H.; Crabb, T. A.; Bradburn, N. L.; Coker, R. D.; Blunden,
G. Phytochemistry 1994, 35, 665-669.
5′-Epialtenuene (7): [R]25 +18 (c 0.185, CH2Cl2); UV (MeOH)
D
λmax (log ꢀ) 240 (4.5); 278 (3.6); 319 (3.3). EIMS, H NMR, and 13C
1
NMR data matched those previously reported.7
(8) Visconti, A.; Botalico, A.; Solfrizzo, M.; Palmisano, F. Mycotoxin
Res. 1989, 5, 69-76.
Preparation of Dibromobenzoate Ester 5a. A solution of 5 (5.7
mg, 0.020 mmol) in CH2Cl2 (2 mL) was treated with p-bromobenzoyl
chloride (9.8 mg, 0.045 mmol) and DMAP (1 crystal). After stirring at
ambient temperature for 24 h, saturated NaHCO3 was added. The
reaction mixture was extracted with CH2Cl2. The combined organic
layer was evaporated to dryness and separated by reversed-phase HPLC
(MeCN/H2O 40-100% over 30 min) to afford 5a (3.7 mg) as a white
solid: UV (MeCN) λmax (log ꢀ) 244 (4.9); CD (MeCN) ∆ꢀ 233 (-23),
(9) McPhail, A. T.; Miller, R. W. J. Chem. Soc., Chem. Commun. 1973,
18, 682-682.
(10) Ayer, W. A.; Racok, J. S. Can. J. Chem. 1990, 68, 2085-2094.
(11) Rogers, D.; Williams, D. J. J. Chem. Soc., Chem. Commun. 1971, 8,
393-393.
(12) Cai, G.; Bozhkova, N.; Odingo, J.; Berova, N.; Nakanishi, K. J. Am.
Chem. Soc. 1993, 115, 7192-7198.
1
(13) Ho¨ller, U.; Gloer, J. B.; Wicklow, D. T. J. Nat. Prod. 2002, 65, 876-
254 (+4.5), 260 (+2.6), 282 (+8.5); H NMR (300 MHz, CDCl3) δ
882.
11.34 (s, 3-OH), 6.56 (d, J ) 2.3 Hz, H-6), 6.50 (d, J ) 2.3 Hz, H-4),
6.20 (d, J ) 2.7 Hz, H-6′), 6.09 (dd, J ) 8.4, 2.7 Hz, H-5′), 5.56 (ddd,
J ) 12, 8.4, 4.0 Hz, H-4′), 3.85 (s, H-8), 2.64 (dd, J ) 12, 4.0 Hz,
H-3′â), 2.54 (t, J ) 12 Hz, H-3′R), 1.76 (s, H-7′); EIMS m/z 660 [(M
+ 4)+; 0.02], 658 [(M + 2)+; 0.04], 656 (M+; 0.02), 458 (8), 456 (14),
182 (100).
(14) Shearer, C. A.; Langsam, D. M.; Longcore, J. E. Fungi in freshwater
habitats. In BiodiVersity of Fungi: InVentory and Monitoring
Methods; Mueller, G. M., Bills, G. F., Foster, M. S., Eds.; Elsevier
Academic Press: Burlington, MA, 2004; pp 513-531.
NP0504661