Coloured artefacts formed by oxidation of benzene-1,2,4-triol and β-dopa during the extraction of cortinarius violaceus (agaricales) with alcohols

Article abstract of DOI:10.1002/ejoc.201101215

An array of coloured artefacts was formed upon extraction of the mushroom Cortinarius violaceus with methanol. The structures of these pigments were determined by spectroscopic investigations, including NMR and mass spectrometric analyses. Cortiviolate A and dihydrocortiviol A arise by oxidative dimerization of benzene-1,2,4-triol, and cortiviolate B is formed by an unprecedented oxidative rearrangement of β-dopa. The oxidations are caused by iron(III) ions, released from the iron complex of (R)-β-dopa- the native violet pigment of the mushroom. In the course of these investigations, syntheses of the cortiviolates, dihydrocortiviol A, gomphilactone, and two related dilactones were carried out. Copyright

Full text of DOI:10.1002/ejoc.201101215

FULL PAPER
DOI: 10.1002/ejoc.201101215
Coloured Artefacts Formed by Oxidation of Benzene-1,2,4-triol and β-Dopa
During the Extraction of Cortinarius violaceus (Agaricales) with Alcohols
Franz von Nussbaum,[a] Matthias Rüth,[a] Peter Spiteller,[a] Tina Hübscher-Weissert,[a]
Florian Löbermann,[a] Kurt Polborn,[a][‡] and Wolfgang Steglich*[a]
Dedicated to Professor Gerhard Bringmann on the occasion of his 60th birthday
Keywords: Natural products / Structure determination / Dimerization / Oxidation / Quinones
An array of coloured artefacts was formed upon extraction
of the mushroom Cortinarius violaceus with methanol. The
structures of these pigments were determined by spectro-
B is formed by an unprecedented oxidative rearrangement
of β-dopa. The oxidations are caused by iron(III) ions, re-
leased from the iron complex of (R)-β-dopa – the native violet
scopic investigations, including NMR and mass spectromet- pigment of the mushroom. In the course of these investi-
ric analyses. Cortiviolate A and dihydrocortiviol A arise by
oxidative dimerization of benzene-1,2,4-triol, and cortiviolate
gations, syntheses of the cortiviolates, dihydrocortiviol A,
gomphilactone, and two related dilactones were carried out.
Introduction
Results and Discussion
Structural Elucidation
Some years ago, we identified the pigment of the dark-
violet mushroom Cortinarius violaceus (German: Violetter
Schleierling) as the iron(III) complex of (R)-β-dopa (1).[1]
The main compound, cortiviolate A, gave a brilliant-blue
solution in methanol, which exhibited a UV/Vis maximum
at 581 nm (log ε = 3.79). The energy-dispersive X-ray spec-
tra and the atomic absorption (AA) spectra indicated the
presence of potassium and minor amounts of sodium, and
the HRMS (ESI+) gave peaks that corresponded to the mo-
lecular formula C13H7KO7. The 1H NMR spectrum
(CD3OD) displayed only five signals: a methoxy singlet at
δ = 3.76 ppm, two doublets at δ = 6.17 and 7.18 (J = 12 Hz)
ppm, and two aromatic singlets at δ = 5.77 and 6.62 ppm.
In the 13C NMR spectrum, the methoxy signal was ob-
served at δ = 52.3 ppm, and, in addition, signals for four
methine carbons at δ = 103.0, 105.7, 126.8, and 138.9 ppm,
and eight quaternary carbon atoms at δ = 96.4, 153.4,
166.9, 169.3, 169.6, 178.9, 184.1, and 186.0 ppm, were ob-
served.
In the course of those investigations, we discovered that
extraction of the fresh fungi with methanol and a small
amount of acetic acid resulted in an interesting play of col-
ours: the initially brownish-grey extract soon became dark-
red and then changed to ultramarine-blue and finally to
greyish-green. Cautious concentration of the blue extract
under reduced pressure, immediately followed by rapid gel
permeation chromatography on Sephadex® LH-20 under
an argon atmosphere, gave red and blue fractions of modest
stability, which, on repeated gel permeation chromatog-
raphy, yielded three pure compounds, named cortiviolate A
and B, and dihydrocortiviol A.
According to the HMBC correlations (Figure 1), the 1H-
coupled 13C NMR spectrum, as well as selective 1H-de-
couplings, cortiviolate A possesses structure 2a, containing
a hydroxyquinone methide anion chromophore.[2] Because
extraction of the mushrooms with ethanol yielded the cor-
responding ethyl ester 2b, the blue pigment must be an arte-
fact, formed during the workup procedure.
The red component, dihydrocortiviol A, was obtained
from 2a by reduction with ascorbic acid or sulfur dioxide,
which suggested catechol structure 3, in agreement with the
spectroscopic data. The UV/Vis spectrum of 3 exhibited
characteristic maxima at 278, 366, and 467 nm and re-
[a] Department Chemie und Biochemie,
Ludwig-Maximilians-Universität München,
Butenandtstr. 5–13, 81377 München, Germany
E-mail: wos@cup.uni-muenchen.de
[‡] Crystal structure determination
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Coloured Artefacts from the Extraction of Cortinarius violaceus
(MSTFA), showed the exclusive formation of a tris(trime-
1
thylsilyl) derivative, in agreement with structure 3. The H
NMR spectrum (CD3OD) of 3 exhibited a methoxy signal
at δ = 3.91 ppm, two singlets for the aromatic protons at δ
= 6.59 and 7.37 ppm, and two doublets (J = 13.0 Hz) at δ
= 6.20 and 8.15 ppm, attributed to the protons at the Z
double bond. Reduction of cortiviolate A (2a) with sulfur
dioxide under more acidic conditions yielded dihydro deriv-
ative 3 as a 5:1 mixture of the Z and E diastereomers. Com-
pound (E)-3 exhibited a larger coupling constant of J =
16.4 Hz for the olefinic protons, thus confirming the Z con-
figuration of dihydrocortiviol A and cortiviolate A.[4]
The rather unstable, blue-violet cortiviolate B could only
be purified with difficulty and yielded no useful mass spec-
trum. The longest wavelength UV/Vis maximum at 560 nm
indicated a smaller chromophore for this pigment than that
of cortiviolate A. Extensive NMR studies including selec-
tive 1H,13C-decouplings and HMBC experiments (Figure 2)
allowed the assignment of structure 5 to cortiviolate B.
Figure 1. Prominent HMBC correlations of cortiviolate A (2a) (in
CD3OD).
sembled that of gomphilactone (4), a red pigment known
from the mushrooms Gomphidius maculatus and G. glu-
tinosus.[3]
Figure 2. Prominent HMBC correlations of cortiviolate B (5) (in
CD3OD).
The HRMS (EI) of 3 showed a molecular ion at m/z =
278.0432, which was consistent with the formula C13H10O7.
The base peak at m/z = 246 (C12H6O6) corresponds to a
ketene fragment, which is formed by elimination of meth-
Synthesis of Gomphilactone and the Cortinarius Artefacts
The structure of cortiviolate A (2a) suggested a simple
anol from the molecular ion. A GC/MS investigation of genesis from gomphilactone (4) by ring opening with meth-
the pertrimethylsilylated product, obtained from 3 by treat- anol. Because gomphilactone had not been synthesized pre-
ment with N-methyl-N-(trimethylsilyl)trifluoroacetamide viously, we developed a synthesis of this pigment from 5,6-
Scheme 1. Syntheses of gomphilactone (4).
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dimethoxybenzofuran-2(3H)-one (6)[5] and maleic anhy- weak base KOAc induces the mild air oxidation of gomphil-
dride (Scheme 1). Heating an intimate mixture of the two actone to the corresponding ortho-quinone 12, which acti-
components for several hours at 150 °C afforded dilactone vates the lactone ring for the opening with methanol
7 as a single stereoisomer in 41% yield, which, after treat- (Scheme 2).
ment with BBr3, provided gomphilactone (4). Alternatively,
addition of lactone 6 to maleic anhydride at ambient tem-
perature in the presence of triethylamine afforded the car-
boxylic acid 8 in high purity, which was directly trans-
formed into dilactone 7 by heating in toluene with a cata-
lytic amount of p-toluenesulfonic acid. The second method
proceeded with an overall yield of 57%. The olefinic pro-
tons of intermediate 8 exhibited a coupling of J = 12.8 Hz
in the 1H NMR spectrum, pointing to the Z configuration.
Scheme 2. Conversion of gomphilactone (4) into cortiviolate A
(2a).
The two-step procedure could also be carried out with
commercially available benzofuran-2(3H)-one and maleic
or phthalic anhydride. The yields of the resulting dilactones
9 and 10 were 56 and 15%, respectively. In the latter case,
the poor yield can be explained by strong steric hindrance
in the transition state. The E configuration of the central
double bond was secured by an X-ray analysis of bisdeoxyg-
omphilactone (9) (Figure 3). Because the NMR spectro-
scopic data of this compound and gomphilactone (4) are in
full agreement, the E configuration of the natural product is
firmly established. Gomphilactone and its relatives exhibit
intense orange or red colours and represent a new class of
indigo-type pigments,[6] isomeric to isoxindigo (11).[7]
The question remained as to which precursor triggers the
formation of the blue and red artefacts in the methanolic
mushroom extract. A GC/MS investigation of the extract
after concentration and trimethylsilylation revealed the
presence of β-dopa[1] and considerable quantities of ben-
zene-1,2,4-triol (13a).[3,8] Clearly, the high concentration of
iron(III) ions released from the indigenous pigment of C.
violaceus[1] induces the oxidative transformation of 13a into
the artefacts. This could be proven by addition of 6-methyl-
benzene-1,2,4-triol (13b) to the mushroom extract, which
gave rise to the 3Ј-methyl derivative 2c of cortiviolate A.
Compound 2c lacks the two doublets for the olefinic pro-
1
tons in the H NMR spectrum ([D6]DMSO) and, instead,
exhibits a narrow quartet at δ = 7.03 (J = 1.6 Hz) ppm,
which is caused by coupling with the allylic methyl group
(δ = 1.95 ppm) (Scheme 3). The chemical shift of the methyl
group is in accord with its position at C-3Ј, favored over C-
4 for steric reasons.
Figure 3. ORTEP plot derived from a single-crystal X-ray analysis
of dilactone 9 (crystallographic numbering).
Scheme 3. Oxidative dimerization of 1,2,4-trihydroxybenzenes 13
to cortiviolate A (2a) and its homologues.
The formation of cortiviolate A (2a) was also demon-
strated by stirring benzenetriol 13a with ferric chloride and
Cortiviolate A (2a) could result from ring opening of potassium acetate in methanol and a catalytic quantity of
gomphilactone (4) with methanol and oxidation of the re- acetic acid. From the resulting mixture of products, the blue
sulting catechol to the hydroxyquinone methide anion. Ini- fraction of cortiviolate A could be isolated by chromatog-
tial experiments to achieve this ring opening with meth- raphy on Sephadex® LH-20. With 6-methylbenzene-1,2,4-
anolic HCl or under base-catalyzed conditions were unsuc- triol (13b), the dimethyl derivative 2d was obtained. In both
cessful. In contrast, stirring a solution of 4 in aqueous cases, the yield was in the range of 1.5–3.5%.
methanol with potassium acetate under air yielded a deep-
A possible reaction sequence for the formation of arte-
blue solution, from which cortiviolate A (2a) could be iso- facts 2a, 2b, and 3 from 13a is depicted in Scheme 4.[9] Oxi-
lated by chromatography on Sephadex® LH-20. Clearly, the dation of the triol 13a might yield hydroxy-1,4-benzoqui-
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Coloured Artefacts from the Extraction of Cortinarius violaceus
Scheme 4. Proposal for the oxidative dimerization of 1,2,4-trihydroxybenzene (13a).
none 14, which then dimerises to give intermediate 15. Oxi- data for this compound were identical to those of the pig-
dation of the latter would yield dibenzoquinone 16, which ment from the extract of C. violaceus.
could undergo a Posternak type rearrangement[9a] after
The formation of cortiviolate B (5) cannot be explained
forming hemiacetal 17 by addition of an alcohol to the by oxidative dimerization of 1,2,4-trihydroxybenzene. How-
most reactive carbonyl group. Proton-catalyzed fragmenta- ever, it appears reasonable that aldehyde 5 originates from
tion of 17, followed by tautomerization, would yield a dihy- the (R)-β-dopa (1) present in the mushroom extract. A hy-
drocortiviol 3 and its oxidation product 2. The sequence is pothetical sequence of reactions for this transformation is
supported by the observation that dihydro compound 3 is given in Scheme 6. In the first step, oxidative cyclization of
the first product to be detected when the extraction of the (R)-β-dopa could yield spirolactone 21, which then affords
fruit bodies of C. violaceus with methanol is followed by ester 22 by a Grob fragmentation. Hydrolysis of the imin-
HPLC. Compound 3 is already present after a few minutes, ium group would yield formyl derivative 23, which, after
whereas the blue cortiviolates A (2a) and B (5) appear much oxidation to the ortho-quinone and formation of the hy-
later. This suggests that the blue quinone methide anions 2a droxymethylene enolate 24, could cyclize to yield aldehyde
and 5 are formed by oxidation of the respective catechols. 25, the direct precursor of cortiviolate B (5).
For the synthesis of cortiviolate B (5), 5,6-dimethoxyben-
To support this proposal, we studied the oxidation of
zofuran-2(3H)-one (6) was heated with N,NЈ-diphenylfor- rac-β-dopa with manganese(IV) oxide in aqueous methanol
mamidine at 145 °C (Scheme 5). The resulting (phen- in the presence of 1 equiv. of potassium acetate and a small
ylamino)methylene derivative 18 was heated to reflux with amount of acetic acid. After 1 h, the dark-blue solution was
methanolic NaOH to yield α-(hydroxymethylene)lactone carefully concentrated, purified by gel permeation
1
19, which, after deprotection with BBr3 and oxidation of chromatography and analyzed by H NMR spectroscopy.
the resulting catechol 20 with air in acetonitrile/water in the To our surprise, however, the spectrum not only showed the
presence of KOAc, afforded cortiviolate B (5); the analytical signals of cortiviolate B (5) but also those of cortiviolate
Scheme 5. Synthesis of cortiviolate B (5).
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Scheme 6. Proposal for the formation of cortiviolate B (5) from (R)-β-dopa (1).
A (2a) in a ratio of about 1:3. Clearly, the activated ester yield under similar conditions, supporting the proposed
intermediate 24 in Scheme 6 suffers partial methanolysis to fragmentation mechanism via a spirolactone intermediate
yield hydroxy-1,4-benzoquinone (14), which is then trans- (i.e., 31) (Scheme 8). Enamide 32 was obtained as a 5:2 mix-
formed into cortiviolate A according to Scheme 4.
ture of the E and Z isomers, which could be separated by
In contrast, the oxidation of rac-N-Boc-β-dopa (26) with chromatography.
PhI(OAc)2 in methanol yielded the dihydrocoumarin deriv-
The intramolecular addition of the enolate unit to the
ative 27 (Scheme 7).
ortho-quinone in Scheme 6 is supported by the formation
of a small quantity of the methyl ester analogue 36 of corti-
violate B[11] upon treatment of 3,4-dihydroxyphenyl methyl
malonate (35) with potassium carbonate in aqueous aceto-
nitrile (Scheme 9). Malonate 35 was prepared from 3,4-
bis(benzyloxy)phenol (33)[12] by esterification with methyl
3-chloro-3-oxopropanoate followed by hydrogenolytic re-
moval of the protecting groups from the resulting diester
34.
Scheme 7. Oxidative cyclization of N-Boc-β-dopa (26).
Conclusions
rac-N-Boc-3-O-methyl-β-dopa (30), which is easily pre-
pared in three steps from 4-benzyloxy-3-methoxybenzalde-
The blue and red pigments formed during the extraction
hyde (28),[10] afforded the rearranged product 32 in low of C. violaceus with alcohols are artefacts arising from the
Scheme 8. Synthesis and oxidative rearrangement of β-dopa derivative 30.
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Coloured Artefacts from the Extraction of Cortinarius violaceus
diation (λ = 0.71073 Å). Analytical TLC: silica gel 60 F254 alumin-
ium foils (Merck). Column chromatography: silica gel 60, 40–63 μm
(Merck); acetylated polyamide-6 (Polyamide SC-6AC, 50–160 μm,
Macherey–Nagel). Gel permeation chromatography: Sephadex®
LH-20 (Pharmacia).
Mushrooms: Cortinarius violaceus were collected in September
1996, 1998, 1999, and 2000 in spruce forests near Gilching (Ba-
varia).
Isolation and Structural Elucidation
Artefacts from the Methanol Extract of C. violaceus: Coarsely
chopped, deep-frozen fruit bodies of C. violaceus (305 g, 18 g dry
weight after extraction) were mechanically shaken in an open Er-
lenmeyer flask (2 L) with MeOH/AcOH (100:1, v/v, 1.5 L) at room
temperature for 6 h. The extract was initially grey-brown and
changed rapidly to red and finally to blue. It is important to start
the workup after the blue colour has fully developed, however,
longer extraction times lead to decomposition with formation of a
greenish colour. The blue extract was filtered and concentrated un-
der reduced pressure to a third of its volume, the temperature being
maintained below 22 °C. The solution was immediately applied to
a Sephadex® LH-20 column (27ϫ 7 cm) and rapidly eluted with
MeOH under an argon atmosphere. The red and blue fractions
were collected and kept at –18 °C in the freezer. The combined
red and blue fractions were then concentrated to 20 mL under the
conditions described above and re-purified by chromatography on
Sephadex® LH-20 (40ϫ 8 cm) with MeOH. Typically, four zones
developed: a weak light-blue zone, an intense red zone containing
dihydrocortiviol A (3), an intense blue zone containing cortiviolate
A (2a), and a very weak grey-blue band containing cortiviolate B
(5), which immediately followed after the blue zone. Careful con-
centration of the last three bands yielded 2a and 3 as NMR spec-
troscopically pure compounds, whereas in the case of 5, re-
chromatography on Sephadex® LH-20 was necessary.
Scheme 9. Synthesis and oxidative cyclization of malonate 35.
native metabolites benzene-1,2,4-triol (13a) and (R)-β-dopa
(1) through oxidation with FeIII ions present in the mush-
room. The cortiviolates and dihydrocortiviol A might be
considered as tertiary metabolites.[13] In the course of our
investigations, gomphilactone and a new type of oxindigo
pigment have been synthesized. The oxidative transforma-
tion of β-dopa into cortiviolate B is supported by model
experiments.
Experimental Section
General: Melting points (uncorrected): Reichert Thermovar hot-
stage apparatus. Elemental analyses: Microanalytical Laboratory,
Ludwig-Maximilians-Universität München. IR: Perkin–Elmer
FTIR. Intensity of the bands: vs (very strong), s (strong), m (me-
dium), w (weak); sh (shoulder). UV/Vis spectra: Perkin–Elmer
Lambda 16 spectrophotometer. NMR: Bruker ARX 300 and AMX
600 spectrometers, with the solvent peak as internal standard
(CDCl3: δH = 7.26 ppm, δC = 77.0 ppm; [D6]DMSO: δH = 2.49
ppm, δC = 39.5 ppm. CD3OD: δH = 3.31 ppm, δC = 49.0 ppm;
[D6]acetone: δH = 2.04 ppm, δC = 29.8 ppm). 1H-Coupled 13C
Cortiviolate A (2a): Yield 40.5 mg (0.23% of dry weight); m.p.
Ͼ380 °C; R (TLC) = 0.33 (CHCl /MeOH, 3:1). IR (KBr): ν =
˜
f
3
3435 (s, br.), 1724 (m), 1639 (s), 1590 (m), 1529 (s), 1441 (m), 1384
(w), 1232 (w), 840 (w) cm–1. UV/Vis (MeOH): λmax (logε) = 219
1
(4.43), 356 (4.21), 581 (3.79) nm. H NMR (600 MHz, CD3OD): δ
= 3.76 (s, 3 H, OCH3), 5.77 (s, 1 H, 7-H), 6.17 (d, J = 12.0 Hz, 1
H, 3Ј-H), 6.62 (s, 1 H, 4-H), 7.18 (d, J = 12.0 Hz, 1 H, 2Ј-H) ppm.
(300 MHz, [D6]DMSO): δ = 3.62 (s, 3 H, OCH3), 5.75 (s, 1 H, 7-
H), 6.19 (d, J = 11.9 Hz, 1 H, 3Ј-H), 6.36 (s, 1 H, 4-H), 7.12 (d, J
= 11.9 Hz, 1 H, 2Ј-H) ppm. 13C NMR (151 MHz, CD3OD, 1H-
coupled): δ = 52.3 (Q, J = 147.9 Hz, OCH3), 96.4 (s, C-3), 103.0
(D, J = 169.0 Hz, C-7), 105.7 (D, J = 170.5 Hz, C-4), 126.8 (D, J
= 166.0 Hz, C-2Ј), 138.9 (D, J = 163.0 Hz, C-3Ј), 153.4 (d, J =
7.1 Hz, C-3a), 166.9 (dd, J = 11.7, 4.7 Hz, C-7a), 169.3 (m, J =
9.4, 4.7 Hz, C-4Ј), 169.6 (s, C-2), 178.9 (d, J = 4.7 Hz, C-5), 184.1
(d, J = 7.0 Hz, C-6), 186.0 (dd, J = 11.7, 4.7 Hz, C-1Ј) ppm.
(151 MHz, [D6]DMSO): δ = 51.4 (OCH3), 92.4 (C-3), 101.8 (C-7),
104.7 (C-4), 126.7 (C-2Ј), 135.3 (C-3Ј), 149.9 (C-3a), 164.7 (C-7a),
167.4 (C-2), 167.6 (C-4Ј), 177.2 (C-5), 181.5 (C-6), 181.7 (C-
1Ј) ppm. The assignments were established by HMQC, HMBC, and
HETCOR experiments, and selective decouplings in the 1H-cou-
pled 13C NMR spectrum (JH4,C6 = 7.0 Hz, JH4,C7a = 11.7 Hz,
1
NMR: multiplets due to JC,H couplings are indicated by capital
3
letters, J and other couplings are in small letters. MS and HRMS
(EI): Finnigan MAT 95Q mass spectrometer (direct inlet: 20–
800 °C, 70 eV, source temperature: 200 °C, mass range: m/z = 41 to
800). GC-MS: A Finnigan MAT 95Q mass spectrometer coupled
with a Varian 3400 gas chromatograph equipped with a split/split-
less injector (split ratio, 1:30) was used. Helium was used as carrier
gas at a flow rate of 1.5 mL/min. The compounds were separated
on a fused silica DB-5ms capillary column (30 mϫ 0.25 mmϫ
0.25 μm, Macherey–Nagel). For sample separation, the following
temperature programme was used: 2 min isocratic at 50 °C, then a
linear gradient of 10 °C/min to 300 °C. Typically, 0.2 μL samples
of a 1% (m/v) solution were injected. Retention indices Ri accord-
ing to Kováts[14] were determined by co-injection of a mixture of
straight chain n-alkanes (C10–C36). MS (ESI): Finnigan TSQ 7000
mass spectrometer equipped with a Finnigan ESI ion source inter-
face [spray voltage 3 kV, mass range 50–800 mu, multiplier 1000 V
(scan modus)]. MS/MS: argon collision gas 2.0 mbar, sheath gas
(N2) 2.9 bar, multiplier 1400 V, collision energy automatically ro-
tated at –20, –30, –40 eV. HRMS (ESI): Finnigan MAT 95Q mass
spectrometer [spray voltage 3 kV, flow rate 10 μL/min, solvent
MeOH/H2O (1:1)]. X-ray diffraction: Enraf–Nonius CAC4 dif-
fractometer at 293(2) K using graphite-monochromated Mo-Kα ra-
JH7,C3a = 7.1 Hz, JH7,C5 = 4.7 Hz, JH7,C7a = 4.7 Hz, JH2Ј,C1Ј
4.7 Hz, JH2Ј,C4Ј = 9.4 Hz, JH3Ј,C1Ј = 11.7 Hz).
=
Energy Dispersive X-ray Spectroscopy (EDX) (30 kV): found C, O,
K. Atomic Absorption Spectroscopy (AAS): K ca. 81 mg/g, Na ca.
10 mg/g, Fe ca. 0.6 mg/g. MS (ESI+): m/z (%) = 353 (100)
[C13H7K2O7]+, 337 (41) [C13H7KNaO7]+, 321 (14) [C13H7Na2O7]+,
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299 (8) [C13H8NaO7]+, 276 (8) [C13H8O7]+. C13H7KO7·0.25MeOH:
calcd. C 49.38, H 2.50; found C 49.37, H 2.66.
of its volume and was then immediately applied to a Sephadex®
LH-20 column (27ϫ 7 cm). Elution with EtOH yielded, in addition
to two weak violet bands, an intense blue fraction, consisting of
the ethyl ester 2b (4.4 mg, 0.009% of fresh weight); m.p. Ͼ350 °C;
Rf (TLC) = 0.14 (CHCl3/MeOH, 5:1). 1H NMR (600 MHz,
[D6]DMSO): δ = 1.16 (t, J = 7.1 Hz, 3 H), 4.08 (q, J = 7.1 Hz, 2
H), 5.75 (s, 1 H, 7-H), 6.16 (d, J = 11.9 Hz, 1 H, 3Ј-H), 6.35 (s, 1
H, 4-H), 7.08 (d, J = 11.9 Hz, 1 H, 2Ј-H) ppm. 13C NMR
(151 MHz, [D6]DMSO): δ = 14.0 (CH3), 60.1 (OCH2), 92.5 (C-3),
101.9 (C-7), 104.7 (C-4), 126.8 (C-2Ј), 135.9 (C-3Ј), 150.0 (C-3a),
164.8 (C-7a), 167.1 (C-2 or C-4Ј), 167.5 (C-4Ј or C-2), 177.3 (C-
5), 181.6 (C-6), 182.1 (C-1Ј) ppm. MS (ESI+): m/z (%) = 367 (31)
Cortiviolate B (5): Yield 2.5 mg (0.014% of dry weight); m.p.
Ͼ340 °C (decomp.); Rf (TLC) = 0.28 (CHCl3/MeOH, 3:1). IR
(KBr): ν = 3436 (s, br.), 1771 (m), 1597 (s, br.), 1538 (s), 1442 (m),
˜
1384 (w), 1359 (w), 1320 (w), 1279 (w), 1230 (m), 836 (m) cm–1.
UV/Vis (MeOH): λmax (logε) = 214 (4.55), 295 (4.23), 345 (4.30),
560 (3.68, br.) nm. 1H NMR (600 MHz, CD3OD): δ = 5.79 (s, 1
H, 7-H), 6.40 (s, 1 H, 4-H), 9.52 (s, 1 H) ppm. 13C NMR (151 MHz,
1
CD3OD, H-coupled): δ = 96.6 (dd, J = 22.4, 1.5 Hz, C-3), 103.4
(D, J = 169.9 Hz, C-7), 105.1 (D, J = 167.8 Hz, C-4), 151.4 (t, J =
6.1 Hz, C-3a), 166.8 (dd, J = 11.5, 4.5 Hz, C-7a), 171.4 (s, C-2),
179.0 (d, J = 4.1 Hz, C-5), 183.8 (dd, J = 5.6, ca. 1.0 Hz, C-6),
184.7 (d, J = 171.4 Hz, CHO) ppm. The assignments were estab-
[C14H9K2O7]+,
351
(10)
[C14H9KNaO7]+,
335
(100)
[C14H9Na2O7]+. HRMS: Calcd. for C14H9KNaO7 [M + Na]+
350.9883; found 350.9883.
lished by HMQC, HMBC, and HETCOR experiments, and selec-
1
tive decouplings in the H-coupled 13C NMR spectrum (JH4,C5
=
Detection of Benzene-1,2,4-triol (13a) in C. violaceus: A 2-g portion
of a deep-frozen fruit body of C. violaceus was minced and ex-
tracted for 30 min with MeOH (2ϫ 50 mL). The extracts were
combined and concentrated under reduced pressure at 40 °C. The
resulting residue was treated with MSTFA (100 μL) at 40 °C for
2 h, and the reaction mixture (0.2 μL) was subjected to GC/MS
analysis. The peak with Ri = 1616 exhibited the same properties as
authentic 1,2,4-tris(trimethylsilyloxy)benzene. MS (EI): m/z (%) =
342 (100) [M+], 327 (8), 311 (1), 254 (9), 239 (80), 223 (1), 211 (6),
193 (1), 179 (4), 147 (7), 133 (5), 119 (1), 112 (4), 91 (2), 75 (8), 73
(81), 59 (2), 45 (26).
Ͼ1.0 Hz, JH4,C6 = 5.6 Hz, JH4,C7a = 11.5 Hz, JH7,C3a = 6.1 Hz,
JH7,C5 = 4.1 Hz, JH7,C7a = 4.5 Hz, JCHO,C3 = 22.4 Hz, JCHO,C3a
=
6.0 Hz). EDX (30 kV): found C, O, K.
Dihydrocortiviol A [(Z)-3]
Isolation from C. violaceus: The combined red and blue fractions
from the MeOH extract of C. violaceus (244 g) were exposed to a
stream of SO2, until the colour had completely changed to red. The
solution was concentrated to 20 mL and purified by chromatog-
raphy on a Sephadex® LH-20 column (40ϫ 8 cm). On elution with
MeOH containing 0.01% SO2, a strong red zone developed, which
was concentrated under reduced pressure to yield (Z)-3 (15 mg,
0.006% of fresh weight).
Synthetic Studies
Gomphilactone Dimethyl Ether (7)
One-Step Procedure: An intimate mixture of 5,6-dimethoxybenzo-
furan-2(3H)-one (6)[5] (200 mg, 1 mmol) and maleic anhydride
(400 mg, 4 mmol) was heated to 150 °C for 16 h. After being cooled
to room temperature, the deep-red melt was dissolved in chloro-
form (150 mL), washed with H2O (2ϫ150 mL) and brine
(150 mL), and dried (MgSO4). The solution was concentrated un-
der reduced pressure and purified by flash chromatography on sil-
ica gel (CH2Cl2/acetone, 10:1) to yield 7 (112 mg, 41%) as red crys-
tals.
Two-Step Procedure: A solution of benzofuranone 6[5] (144 mg,
0.74 mmol) and triethylamine (0.12 mL, 0.86 mmol) in anhydrous
THF (10 mL) was added dropwise to a vigorously stirred solution
of maleic anhydride (80 mg, 0.82 mmol) in anhydrous THF (8 mL).
After stirring for a further 2 h, the orange-red reaction mixture was
quenched with 1.1 m aq. KHSO4 (10 mL) and extracted with
EtOAc (3ϫ 100 mL). The combined organic phases were washed
with brine (100 mL), dried (MgSO4), and concentrated under re-
duced pressure to yield carboxylic acid 8 as red crystals (for spec-
troscopic data, see below), which were suspended in anhydrous tol-
uene (50 mL), treated with a catalytic amount of TsOH, and heated
to reflux for 3 h with the use of a Dean–Stark trap. After concen-
tration under reduced pressure, the residue was purified by flash
chromatography on silica gel (CH2Cl2/acetone, 10:1) to yield 7
(115 mg, 57%) as red crystals; m.p. Ͼ200 °C (decomp.); Rf (TLC)
From Reduction of Cortiviolate A (2a): A weak stream of SO2 was
passed for a few seconds through a solution of anion 2a (3.5 mg,
11 μmol) in MeOH (2 mL), whereupon the colour changed from
deep-blue to red. The solution was degassed with argon and con-
centrated under reduced pressure to afford a red residue, which
consisted of a ca. 5:1 mixture of (Z)-3 and (E)-3. 1H NMR
(600 MHz, [D6]DMSO): δ [(Z)-3] = 3.83 (s, 3 H, OCH3), 6.35 (d,
J = 13 Hz, 1 H, 3Ј-H), 6.59 (s, 1 H, 7-H), 7.26 (s, 1 H, 4-H), 7.91
(d, J = 13 Hz, 1 H, 2Ј-H), 12.90 (s, 1 H, OH) ppm. δ [(E)-3] = 3.74
(s, 3 H, OCH3), 6.58 (s, 1 H, 7-H), 6.77 (d, J = 16.4 Hz, 1 H, 3Ј-
H), 7.28 (s, 1 H, 4-H), 8.40 (d, J = 16.4 Hz, 1 H, 2Ј-H) ppm.
Chromatography of the mixture with MeOH on a Sephadex® LH-
20 column (25ϫ 0.4 cm) under an argon atmosphere yielded pure
(Z)-3 (3 mg, 97%) as a red solid; m.p. Ͼ140 °C (decomp.). UV/Vis
(MeOH/H2O, 1:1): λmax (qual.) = 204, ca. 230 (sh), 278, ca. 295
(sh), 366, 467 nm. 1H NMR (300 MHz, CD3OD): δ = 3.91 (s, 3 H,
OCH3), 6.20 (d, J = 13.0 Hz, 1 H, 3Ј-H), 6.59 (s, 1 H, 7-H), 7.37
(s, 1 H, 4-H), 8.15 (d, J = 13.0 Hz, 1 H, 2Ј-H) ppm. 13C NMR
(151 MHz, [D6]DMSO): δ = 53.5 (OCH3), 98.2 (CH), 106.2 (C),
110.8 (CH), 114.2 (C), 123.3 (CH), 135.2 (CH), 141.9 (C), 145.1
(C), 147.3 (C), 157.2 (C), 168.2 (C), 169.6 (C) ppm. MS (EI): m/z
(%) = 278 (6) [M+], 247 (17), 246 (100) [M – CH3OH]+, 162 (24)
[C9H6O3]+. HRMS: Calcd. for C13H10O7 [M+] 278.0427; found
278.0432.
= 0.55 (CH Cl /acetone, 6:1). IR (KBr): ν = 3435 (m, br.), 1794
˜
2
2
A sample of (Z)-3 (0.1 mg) was suspended in MSTFA (10 μL) and
shaken for 5 min at room temperature. The mixture was kept at
45 °C for 2 h and was then directly used for GC/EI-MS:
C22H34O7Si3 [M+]: calcd. 494.1612; found 494.1612.
(vs), 1767 (vs), 1649 (m), 1615 (s), 1538 (m), 1499 (s), 1477 (m),
1438 (m), 1422 (w), 1358 (w), 1338 (m), 1308 (vs), 1269 (s), 1249
(s), 1194 (s), 1156 (m), 1114 (m), 1054 (m), 1004 (s), 964 (m), 948
(m), 873 (m), 847 (m), 788 (m) cm–1. UV/Vis (MeOH): λmax (logε)
= 272 (4.08), 290 (sh, 3.99), 354 (3.90), 454 (3.89) nm. 1H NMR
(300 MHz, CDCl3): δ = 3.94 (s, 3 H, OCH3), 3.95 (s, 3 H, OCH3),
6.48 (d, J = 5.5 Hz, 1 H, 4Ј-H), 6.71 (s, 1 H, 7-H), 7.30 (s, 1 H, 4-
H), 8.47 (d, J = 5.5 Hz, 1 H, 3Ј-H) ppm. 13C NMR (75.5 MHz,
CDCl3): δ = 56.5 (OCH3), 56.8 (OCH3), 95.6 (CH), 107.1 (CH),
Isolation of the Ethyl Ester Analogue 2b of Cortiviolate A: Coarsely
chopped, deep-frozen fruit bodies of C. violaceus (51 g) were ex-
tracted with EtOH/AcOH (100:1, v/v, 300 mL) at room tempera-
ture for 2 h as described for the isolation of 2a. After filtration, the
solution was concentrated under reduced pressure (Ͻ22 °C) to half
386
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© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2012, 380–390

Coloured Artefacts from the Extraction of Cortinarius violaceus
112.7 (C), 122.1 (CH), 141.0 (CH), 147.0 (C), 149.9 (C), 152.7 (C), (CH), 154.0 (C), 155.0 (C), 166.5 (C=O), 167.3 (C=O) ppm. MS
153.5 (C), 167.2 (C=O), 167.9 (C=O), (1 Cq obscured) ppm. MS
(EI): m/z (%) = 214 (100) [M+], 186 (17) [M – CO]+, 160 (11), 145
(EI): m/z (%) = 274 (100) [M+], 259 (40) [M – CH3]+, 231 (7), 203 (15), 130 (14), 102 (17), 76 (19). C12H6O4 (214.17): calcd. C 67.29,
(7), 175 (6). C14H10O6 (274.2): calcd. C 61.32, H 3.68; found C
61.18, H 3.62.
H 2.82; found C 67.29, H 2.98.
Crystallization of 9 from CHCl3 gave crystals that were suitable for
Intermediate 8: Rf (TLC)
=
0.27 (benzene/HCO2Et/HCO2H,
X-ray structure determination.
10:5:3). 1H NMR (300 MHz, [D6]DMSO): δ = 3.74 (s, 3 H, OCH3),
3.75 (s, 3 H, OCH3), 6.11 (d, J = 12.8 Hz, 1 H), 6.84 (s, 1 H), 7.45
(s, 1 H), 7.92 (d, J = 12.8 Hz, 1 H) ppm. 13C NMR (75.5 MHz,
[D6]DMSO): δ = 56.2 (OCH3), 56.3 (OCH3), 95.6 (CH), 100.5 (C),
106.6 (CH), 118.0 (C), 130.6 (CH), 134.3 (CH), 144.0 (C), 145.1
(C), 148.0 (C), 167.9 (C), 168.7 (C), 169.2 (C) ppm. MS (EI): m/z
(%) = 292 (2) [M+], 274 (100) [M – H2O]+, 259 (27), 194 (40), 166
(33), 84 (84), 66 (93). HRMS: Calcd. for C14H12O7 [M+] 292.0583;
found 292.0554.
(E)-3-[3-Oxo-isobenzofuran-1(3H)-ylidene]benzofuran-2(3H)-one
(10): Prepared from benzofuran-2(3H)-one (100 mg, 0.74 mmol),
phthalic anhydride (120 mg, 0.81 mmol), and triethylamine
(0.12 mL, 0.86 mmol) in anhydrous THF (10 mL) according to the
two-step procedure described for the reaction of 6. The orange
crude product was purified by flash chromatography on silica gel
(hexanes/EtOAc, 1:1) to yield 10 (30 mg, 15%) as a red solid; m.p.
215 °C; Rf (TLC) = 0.83 (hexanes/EtOAc, 1:1). UV/Vis (MeOH):
λmax (logε) = 253 (3.88), 385 (3.95) nm. 1H NMR (300 MHz,
CDCl3): δ = 7.15 (d, J = 8.0 Hz, 1 H, 7-H), 7.25 (dd, J = ca. 7.7,
7.5 Hz, 1 H, 5-H), 7.40 (dd, J = 8.0, ca. 7.7 Hz, 1 H, 6-H), 7.77
(dd, J = ca. 7.7, 7.5 or 8.0 Hz, 1 H, 5Ј-H or 6Ј-H), 7.91 (dd, J ca.
7.7, 8.0 or 7.5 Hz, 1 H, 6Ј-H or 5Ј-H), 8.07 (d, J = 7.5 Hz, 1 H, 4-
H or 4Ј-H), 8.11 (d, J = 7.5 Hz, 1 H, 4Ј-H or 4-H), 9.32 (d, J =
8.0 Hz, 1 H, 7Ј-H) ppm. 13C NMR (76 MHz, CDCl3): δ = 106.4
(C), 110.5 (CH), 122.8 (C), 124.6 (CH), 125.2 (C), 125.9 (CH),
126.1 (CH), 128.1 (CH), 130.7 (CH), 133.1 (CH), 136.0 (CH), 136.3
(C), 153.0 (C), 154.4 (C), 165.0 (C=O), 167.0 (C=O) ppm. MS (EI):
m/z (%) = 264 (100) [M+], 220 (33) [M – CO2]+, 152 (13). HRMS:
Calcd. for C16H8O4 [M+] 264.0422; found 264.0409.
Gomphilactone (4): To a solution of ether 7 (100 mg, 0.36 mmol)
in anhydrous CH2Cl2 (20 mL), maintained at –78 °C, was added
dropwise a solution of BBr3 (0.16 mL, 1.17 mmol) in anhydrous
CH2Cl2 (5 mL), whereupon the solution immediately turned bor-
deaux-red. The cooling bath was removed, and the mixture was
stirred for an additional 16 h at room temperature. The solution
was poured into ice water (150 mL) and extracted with EtOAc (3ϫ
150 mL). The combined organic phases were washed with H2O
(100 mL) and brine (100 mL), dried (MgSO4), and concentrated.
The resulting red-violet powder was purified by chromatography
on acetylated polyamide (acetone) to yield 4 (54 mg, 61%); m.p.
Ͼ250 °C (decomp.); Rf (TLC) = 0.50 (benzene/HCO2Et/HCO2H,
Formation of 2a and Methyl Homologues 2c and 2d, from Benzene-
1,2,4-triol (13a) and 6-Methylbenzene-1,2,4-triol (13b), respectively
10:5:3). IR (KBr): ν = 3391 (vs, br.), 1779 (vs), 1750 (vs), 1652 (m),
˜
1612 (s), 1544 (w), 1502 (w), 1471 (s), 1382 (w), 1340 (m), 1312
(vs), 1276 (vs), 1196 (m), 1170 (m), 1114 (s), 1057 (m), 1001 (m),
948 (m), 878 (m), 815 (m) cm–1. UV/Vis (MeOH): λmax (logε) =
276 (4.07), 298 (3.91, sh), 361 (3.96), 476 (3.92) nm. 1H NMR
(600 MHz, [D6]DMSO): δ = 6.68 (s, 1 H, 7-H), 6.83 (d, J = 5.5 Hz,
1 H, 4Ј-H), 7.17 (s, 1 H, 4-H), 8.36 (d, J = 5.5 Hz, 1 H, 3Ј-H) ppm.
(300 MHz, [D6]acetone): δ = 6.70 (d, J = 5.5 Hz, 1 H, 4Ј-H), 6.72
(s, 1 H, 7-H), 7.28 (s, 1 H, 4-H), 8.42 (d, J = 5.5 Hz, 1 H, 3Ј-
H) ppm. 13C NMR (151 MHz, [D6]DMSO): δ = 98.9 (C-7), 106.2
(C-3), 110.1 (C-4), 111.4 (C-3a), 122.2 (C-4Ј), 140.6 (C-3Ј), 143.1
(C-5), 148.1 (C-7a), 150.5 (C-6), 151.4 (C-2Ј), 166.8 (C-2), 167.9
(C-5Ј) ppm. 13C NMR (75 MHz, [D6]acetone): δ = 99.6, 107.2,
111.4, 113.4, 123.0, 141.5, 143.4, 150.0, 150.9, 153.3, 167.7,
168.6 ppm. MS (EI): m/z (%) = 246 (100) [M+], 218 (5), 192 (11),
162 (33). HRMS: Calcd. for C12H6O6 [M+] 246.0164; found
246.0166.
Cortiviolate A (2a): To a solution of triol 13a (50 mg, 0.40 mmol)
in MeOH (500 mL) were added FeCl3 (10 mg) and a catalytic
amount of AcOH (0.2 mL). By using a dropping funnel, a solution
of KOAc (20 mg) in MeOH (50 mL) was added at a rate of 20
drops per min. After a while, the yellowish solution turned dirty-
brown. The stirring was continued for a further 6 h and then the
solution was concentrated under reduced pressure at 25 °C, to a
volume of 2 mL. Chromatography of the resulting solution on Se-
phadex® LH-20 (MeOH) yielded, after several reddish-brown frac-
tions, a blue zone, which was concentrated under reduced pressure
to afford the potassium salt 1a (2.2 mg, 3.5%), the spectroscopic
properties (UV/Vis, 1H NMR in CD3OD, 13C NMR) of which were
in excellent agreement with those of the compound obtained from
1
the extract of C. violaceus. H NMR (600 MHz, [D6]DMSO): δ =
3.76 (s, 3 H, OCH3), 5.77 (s, 1 H, 7-H), 6.16 (d, J = 12.0 Hz, 1 H,
3Ј-H), 6.61 (s, 1 H, 4-H), 7.18 (d, J = 12.0 Hz, 1 H, 2Ј-H) ppm.
The chemical shift of the 4-H proton (δ = 6.61 ppm) of the pure
potassium salt in [D6]DMSO differed considerably from that of the
K/Na mixture obtained from the mushroom extract (δ = 6.36 ppm).
HRMS (ESI–): calcd. for C13H7O7 [M – K]– (100%) 275.0185;
found 275.0192.
The synthetic product was identical with gomphilactone[3] by com-
parison of the IR, 1H NMR and mass spectra, as well as TLC
chromatography.
(E)-3-[5-Oxofuran-2(5H)-ylidene]benzofuran-2(3H)-one (Bisdeoxy-
gomphilactone, 9): Synthesized from benzofuran-2(3H)-one
(600 mg, 4.47 mmol), maleic anhydride (480 mg, 4.89 mmol), and
triethylamine (0.72 mL, 5.16 mmol) in anhydrous THF (100 mL)
according to the two-step procedure described for the reaction of
6. The crude product was purified by flash chromatography on sil-
ica gel (CH2Cl2/acetone, 10:1) to yield 9 (540 mg, 56%) as an
orange-red solid; m.p. 170 °C; Rf (TLC) = 0.20 (hexanes/EtOAc,
4:1). UV/Vis (MeOH): λmax (logε) = 254 (4.02), 271 (sh, 3.88), 364
(4.06), 392 (sh, 4.04) nm. 1H NMR (600 MHz, CDCl3): δ = 6.58
(d, J = 5.5 Hz, 1 H, 4Ј-H), 7.14 (d, J = 8.2 Hz, 1 H, 7-H), 7.24 (dd,
J = 7.7, ca. 7.7 Hz, 1 H, 5-H), 7.42 (dd, J = 8.2, ca. 7.7 Hz, 1 H,
6-H), 7.89 (d, J = 7.7 Hz, 1 H, 4-H), 8.53 (d, J = 5.5 Hz, 1 H, 3Ј-
H) ppm. 13C NMR (151 MHz, CDCl3): δ = 106.2 (C), 110.9 (CH),
121.4 (C), 123.7 (CH), 124.9 (CH), 125.8 (CH), 132.1 (CH), 141.2
3Ј,4-Dimethyl Derivative 2d of Cortiviolate A: Dimethyl derivative
2d was synthesized from triol 13b (50 mg, 0.36 mmol) as described
for the preparation of 2a. Yield 0.9 mg (1.5%). 1H NMR
(600 MHz, CD3OD): δ = 2.02 (s, 3 H, 4-CH3), 2.03 (d, 4J = 1.4 Hz,
4
3 H, 3Ј-CH3), 3.73 (s, 3 H, OCH3), 5.65 (s, 1 H, 7-H), 7.05 (q, J
= 1.4 Hz, 1 H, 2Ј-H) ppm. 13C NMR (151 MHz, CD3OD): δ = 13.7
(4-CH3), 20.4 (3Ј-CH3), 52.5 (OCH3), 96.7 (C-3), 101.1 (C-7), 116.0
(C-4), 133.1 (C-2Ј), 138.9 (C-3Ј), 148.6 (C-3a), 169.2 (C-7a), 170.5
(C-2), 172.7 (C-4Ј), 178.4 (C-5), 183.3 (C-6), 184.7 (C-1Ј) ppm.
HRMS: Calcd for C15H11O7 [M – K]– 303.0548; found 303.0553.
3Ј-Methyl Derivative 2c of Cortiviolate A: Either fresh or deep-fro-
zen fruit bodies (25 g) of C. violaceus were chopped and stirred
Eur. J. Org. Chem. 2012, 380–390
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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387

W. Steglich et al.
FULL PAPER
with triol 13b (20 mg, 0.14 mmol) in MeOH (200 mL) and AcOH
(0.5 mL) in an open beaker at 25 °C for 4 h. The volume of the
brownish-blue extract was reduced to 50 mL under reduced pres-
sure and the solution was purified by chromatography on Se-
phadex® LH-20 (MeOH). All coloured fractions were combined
and concentrated under reduced pressure to 2 mL. Repeated
chromatography on Sephadex® LH-20 (MeOH) yielded a deep-blue
fraction, which was concentrated under reduced pressure at 25 °C
to give potassium salt 2c (0.9 mg, 1.9%). 1H NMR (600 MHz,
(br. s, 1 H, OH) ppm. 13C NMR (76 MHz, CDCl3): δ = 56.4
(OCH3), 56.7 (OCH3), 96.5 (CH), 101.9 (CH), 103.0 (C), 112.5 (C),
145.4 (C), 146.5 (C), 149.8 (C), 156.6 (CH), 172.8 (CO) ppm. MS
(EI): m/z (%) = 222 (100) [M+], 207 (36) [M – CH3]+, 194 (28) [M –
CO]+, 193 (47) [M – CHO]+, 179 (18), 166 (24), 151 (20), 123 (17),
95 (16). HRMS: Calcd. for C11H10O5 [M+] 222.0528; found
222.0518.
5,6-Dihydroxy-3-(hydroxymethylene)benzofuran-2(3H)-one (20): To
a suspension of methyl ether 19 (450 mg, 2.02 mmol) in anhydrous
CH2Cl2 (15 mL), maintained at –78 °C, was added dropwise a 1 m
solution of BBr3 (9.56 mL, 9.56 mmol) in CH2Cl2. After removal
of the cooling bath, the mixture was stirred at room temperature
for 16 h. The reaction mixture was cooled to 0 °C, quenched by
addition of 2 n HCl (10 mL), and extracted with EtOAc (3ϫ
100 mL). The combined organic phases were washed with H2O
(100 mL) and brine (100 mL), and dried (MgSO4). Removal of the
4
[D6]DMSO): δ = 1.95 (d, J = 1.6 Hz, 3 H, 3Ј-CH3), 3.64 (s, 3 H,
4
OCH3), 5.72 (s, 1 H, 7-H), 6.35 (s, 1 H, 4-H), 7.03 (q, J = 1.6 Hz,
1 H, 2Ј-H) ppm. 13C NMR (151 MHz, [D6]DMSO): δ = 20.3 (3Ј-
CH3), 51.6 (OCH3), 92.3 (C-3), 101.6 (C-7), 104.6 (C-4), 126.7 (C-
2Ј), 139.8 (C-3Ј), 150.1 (C-3a), 164.6 (C-7a), 167.4 (C-2), 170.8 (C-
4Ј), 177.0 (C-5), 180.5 (C-1Ј), 181.4 (C-6) ppm. HRMS: Calcd. for
C14H9O7 [M – K]– 289.0348; found 289.0365.
Formation of 2a from Gomphilactone (4): To a solution of 4 (10 mg, solvent under reduced pressure gave a grey powder, which was puri-
0.04 mmol) in MeOH (10 mL) and H2O (1 mL) was added KOAc
(20 mg), and the mixture was stirred in an open beaker. After a few
seconds, the solution turned intense-blue. After 15 min of stirring,
the reaction was complete, and the solution was concentrated under
reduced pressure to 2 mL at a temperature maintained below 20 °C.
The resulting solution was immediately purified over Sephadex®
LH-20 (MeOH) to give 2a (9 mg, 72%); m.p. Ͼ300 °C. The spec-
troscopic data for the synthetic potassium salt agreed with those
of the compound obtained from benzene-1,2,4-triol.
fied by chromatography on Sephadex® LH-20 (MeOH) to yield 20
(250 mg, 64%); m.p. 222 °C; Rf (TLC) = 0.41 (toluene/HCO2Et/
HCO2H, 5:4:1). H NMR (300 MHz, CD3OD): δ = 6.57 (s, 1 H),
1
7.11 (s, 1 H), 7.79 (s, 1 H) ppm. 13C NMR (76 MHz, CD3OD): δ
= 99.4 (CH), 104.1 (C), 110.6 (CH), 115.5 (C), 143.1 (C), 146.4
(C), 146.6 (C), 156.5 (CH), 174.0 (CO) ppm. MS (EI): m/z (%) =
194 (100) [M+], 166 (41) [M – CO]+, 165 (43) [M – CHO]+, 138
(17), 137 (8), 110 (9), 69 (9), 68 (11). HRMS: Calcd. for C9H6O5
[M+] 194.0215; found 194.0215.
Synthesis of Cortiviolate B (5)
Cortiviolate B (5): To a solution of dihydro compound 20 (20 mg,
0.1 mmol) in acetonitrile (30 mL) was added KOAc (20 mg,
0.2 mmol) in H2O (2 mL). The mixture was stirred in an open
beaker at room temperature for 3 h, whereby the solution rapidly
turned blue-violet. The solution was then concentrated to 5 mL at
a temperature not exceeding 20 °C. The concentrate was lyophilized
and finally purified by chromatography on Sephadex® LH-20
(MeOH) to yield 5 (17 mg, 74%) as a greyish-blue solid; m.p.
Ͼ340 °C (decomp.); Rf (TLC) = 0.30 (CHCl3/MeOH, 3:1). UV/Vis
(MeOH): λmax (logε) = 215 (5.00), 291 (4.59), 351 (4.69), 574 (4.31)
5,6-Dimethoxy(3-phenylaminomethylene)benzofuran-2(3H)-one (18):
An intimate mixture of benzofuranone 6 (1.00 g, 5.15 mmol) and
N,NЈ-diphenylformamidine (1.01 g, 5.15 mmol) was heated in a se-
aled glass ampoule to 145 °C for 45 min. The resulting brownish
oil was treated with MeOH (20 mL), crystallized by sonication, and
cooled to 0 °C. The crystals were collected by filtration, washed
with a small amount of cold MeOH, and dried. The yellow residue
(1.20 g, 78%) was recrystallized from MeOH to yield pure 18
(0.97 g, 63%); m.p. 180 °C; Rf (TLC) = 0.82 (hexanes/EtOAc, 1:4).
UV/Vis (MeOH): λmax (logε) = 203 (4.30), 221 (sh, 4.07), 245
1
nm. H NMR (300 MHz, CD3OD): δ = 5.79 (s, 1 H, 7-H), 6.40 (s,
1 H, 4-H), 9.52 (s, 1 H, CHO) ppm. (600 MHz, [D6]DMSO): δ =
1
(3.88), 287 (3.81), 317 (3.62), 388 (4.27) nm. H NMR (300 MHz,
5.78 (s, 1 H, 7-H), 6.17 (s, 1 H, 4-H), 9.48 (s, 1 H, CHO) ppm. 13
C
CDCl3): δ = 3.87 (s, 3 H, OCH3), 3.90 (s, 3 H, OCH3), 6.75 (s, 1
H), 6.88 (s, 1 H), 7.13 (m, 3 H), 7.37 (m, 2 H), 7.88 (d, J = 13.2 Hz,
1 H), 9.89 (d, J = 13.2 Hz, 1 H, NH) ppm. 13C NMR (76 MHz,
CDCl3): δ = 56.3 (OCH3), 56.7 (OCH3), 95.4 (C), 96.3 (CH), 100.6
(CH), 115.7 (C), 116.2 (2ϫ CH), 124.2 (CH), 129.9 (2ϫ CH), 137.1
NMR (150 MHz, [D6]DMSO): δ = 92.9 (C-3), 102.4 (C-7), 103.8
(C-4), 148.1 (C-3a), 164.6 (C-7a), 169.1 (C-2), 177.1 (C-5), 181.5
(C-6), 181.5 (CHO) ppm. HRMS: Calcd. for C9H3O5 [M – K]–
190.9980; found 190.9965.
(CH), 139.3 (C), 144.5 (C), 146.0 (C), 147.8 (C), 170.9 (C=O) ppm. Oxidation of rac-β-Dopa with Manganese(IV) Oxide: rac-β-Dopa
MS (EI): m/z (%) = 297 (100) [M+], 282 (55) [M – CH3]+.
C17H15NO4 (297.31): calcd. C 68.68, H 5.08, N 4.71; found C
68.36, H 5.08, N 4.69.
(197 mg, 1 mmol) was dissolved in a mixture of MeOH/AcOH
(100:1, v/v, 100 mL) and H2O (10 mL). After the addition of KOAc
(100 mg, 1 mmol), the mixture was stirred at 20 °C for 10 min,
treated with MnO2 (2.6 g, 30 mmol), and stirred for a further 1 h.
The product was filtered through Celite® and the filter cake was
washed with MeOH. The dark-blue solution was then submitted
to chromatography on a Sephadex® LH-20 column (MeOH). The
blue fractions were collected and carefully concentrated under re-
duced pressure at a temperature not exceeding 25 °C. After drying
under reduced pressure, the residue (12 mg) was analyzed by 1H
NMR spectroscopy (600 MHz, CD3OD). The signals indicated a
mixture of the potassium salts 2a [δ = (3.72, s, 3 H), 5.76 (s, 1 H),
6.13 (d, J = 12 Hz, 1 H), 6.58 (s, 1 H), 7.13 (d, J = 12 Hz, 1 H) ppm]
and 5 [δ = 5.79 (s), 6.40 (s), 9.52 (s) ppm] in a ratio of 2.8:1. In
addition, signals of an unknown compound appeared at δ = 6.23
(s) and 7.44 (s) ppm.
3-Hydroxymethylene-5,6-dimethoxybenzofuran-2(3H)-one (19):
A
suspension of enamine 18 (100 mg, 0.34 mmol) in MeOH (6 mL)
and aq. NaOH (10%, 10 mL) was heated to reflux for 20 min,
whereupon the solution turned yellow. The mixture was cooled to
0 °C, adjusted with concd HCl to pH 2, and extracted with Et2O
(3ϫ 50 mL). The combined extracts were dried (MgSO4) and con-
centrated under reduced pressure. The crude product was recrys-
tallized from CHCl3 to yield 19 (56 mg, 74%) as yellow crystals;
m.p. 131 °C; Rf (TLC) = 0.67 (toluene/HCO2Et/HCO2H, 5:4:1). IR
(KBr): ν = 3474 (s, br.), 2961 (m), 1763 (s), 1665 (s), 1625 (m), 1602
˜
(m), 1498 (s), 1475 (m), 1440 (m), 1381 (w), 1318 (m), 1262 (m),
1216 (vs), 1118 (vs), 1093 (m), 1002 (m), 960 (w), 846 (w) cm–1.
UV/Vis (MeOH): λmax (logε) = 257 (3.95), 301 (3.81), 352 (3.80)
1
nm. H NMR (300 MHz, [D6]acetone): δ = 3.80 (s, 3 H, OCH3),
4-tert-Butoxycarbonylamino-3,4-dihydro-6,7-dihydroxy-1-benzo-
pyran-2-one (27): To a stirred solution of rac-N-Boc-β-dopa (26;
3.83 (s, 3 H, OCH3), 6.80 (s, 1 H), 7.28 (s, 1 H), 8.04 (s, 1 H), 10.90
388
www.eurjoc.org
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2012, 380–390

Coloured Artefacts from the Extraction of Cortinarius violaceus
0.50 g, 1.68 mmol) in anhydrous MeOH (15 mL) was added
PhI(OAc)2 (0.60 g, 1.86 mmol) at 0 °C. The mixture was stirred for
a further 30 min, then EtOAc (45 mL) was added and the solution
was washed with saturated aq. NaHCO3 (2ϫ 30 mL). The aqueous
phases were re-extracted with EtOAc (4ϫ 30 mL), and the com-
bined organic extracts were washed with brine (2ϫ 25 mL) and
dried (MgSO4). Concentration of the solution and chromatography
of the dark-red residue on silica gel (CHCl3/MeOH, 10:1) yielded
benzopyranone 27 (0.06 g, 12%) as an orange solid; m.p. Ͼ80 °C
with EtOAc (3ϫ 30 mL), and the combined organic phases were
washed with brine (2ϫ 30 mL) and dried (MgSO4). Concentration
under reduced pressure yielded 30 (2.57 g, 76%) as a white solid;
m.p. 154–157 °C (decomp.); Rf (TLC) = 0.42 (toluene/HCO2Et/
1
HCO2H, 10:5:3). H NMR (300 MHz, CD3CN): δ = 1.38 (s, 9 H,
2
3
2
tBu), 2.67 (dd, J = 15.6, J = 6.5 Hz, 1 H, CHAHB), 2.74 (dd, J
3
= 15.6, J = 7.7 Hz, 1 H, CHAHB), 3.84 (s, 3 H, OCH3), 4.88 (br.
dd, 1 H), 5.82 (br. s, 1 H, NH), 6.76–6.90 (m, 3 H, ArH) ppm. 13
C
NMR (76 MHz, CD3CN): δ = 28.6 [C(CH3)3], 41.6 (CH2), 52.2
(CHNH), 56.7 (OCH3), 79.6 [C(CH3)3], 111.0 (CH), 115.4 (CH),
119.8 (CH), 135.3 (C), 146.3 (C), 148.2 (C), 156.1 (CONH), 172.4
1
(slow decomp.); Rf (TLC) = 0.30 (CHCl3/MeOH, 10:1). H NMR
(300 MHz, [D6]DMSO): δ = 1.40 (s, 9 H, tBu), 2.71 (dd, J = 16.1,
7.0 Hz, 1 H, CHAHB), 2.91 (dd, J = 16.1, 5.7 Hz, 1 H, CHAHB), (CO2H) ppm. HRMS: Calcd. for C15H21NO6 [M+] 311.1369; found
4.72 (br. m, 1 H, CHNH), 6.44 (s, 1 H, ArH), 6.64 (s, 1 H, ArH),
7.41 (br. s, 1 H, NH), 8.96 (br. s, 1 H, OH), 9.23 (br. s, 1 H,
OH) ppm. 13C NMR (76 MHz, [D6]DMSO): δ = 28.4 [C(CH3)3],
36.0 (CH2), 44.7 (CHNH), 78.5 [C(CH3)3], 104.0 (CH), 113.5 (CH),
113.9 (C), 142.1 (C), 143.5 (C), 146.0 (C), 155.3 (CONH), 167.2
(lactone C=O) ppm. MS (EI): m/z (%) = 295 (2) [M+], 239 (100)
[M – C4H8]+, 222 (3) [M – C4H9O]+, 204 (2), 197 (33), 194 (2), 179
(33) [M – C5H10NO2]+, 178 (63) [M – C5H11NO2]+, 153 (46), 150
(25) [C8H6O3]+, 123 (4), 77 (2), 69 (3), 59 (7), 57 (34), 44 (9), 41
(18). HRMS: Calcd for C14H17NO6 [M+] 295.1056; found
295.1044.
3-Amino-3-(4-benzyloxy-3-methoxyphenyl)propanoic Acid (29):[15] A
solution of 4-benzyloxy-3-methoxybenzaldehyde (28)[10] (3.98 g,
16.4 mmol), malonic acid (2.22 g, 21.3 mmol), and NH4OAc
(3.22 g, 41.8 mmol) in EtOH (4.2 mL) was heated to reflux. After
1 h, a colourless solid precipitated, and heating was continued for
a further 4 h. The solid was collected by filtration, and washed with
a small volume of hot EtOH and then with Et2O to yield amino
311.1363.
4-Hydroxy-3-methoxyphenyl-(E)- and (Z)-3-(tert-Butoxycarbonyl-
amino)propenoate (32): To a stirred solution of acid 30 (0.55 g,
1.77 mmol) in absolute MeOH (18 mL) was added PhI(OAc)2
(0.67 g, 2.08 mmol) at 0 °C, and the mixture was stirred at 0 °C for
30 min. EtOAc (35 mL) was added and the solution was washed
with saturated aq. NaHCO3 (2ϫ 18 mL). The aqueous phases were
re-extracted with EtOAc (2ϫ 30 mL), and the combined organic
extracts were washed with brine (2ϫ 25 mL) and dried (MgSO4).
The yellow solution was concentrated under reduced pressure and
purified by chromatography on silica gel (hexanes/EtOAc, 2:1) to
give the Z and E diastereomers of ester 32 in yields of 10 mg (2%)
and 30 mg (5%), respectively.
(Z)-32: Brown oil; Rf (TLC) = 0.59 (hexanes/EtOAc, 2:1). 1H NMR
(300 MHz, CD3CN): δ = 1.48 (s, 9 H, tBu), 3.82 (s, 3 H, OCH3),
5.21 (d, J = 8.9 Hz, 1 H, CH=CH), 6.48 (s, 1 H, OH), 6.56 (dd, J
= 8.6, 2.6 Hz, 1 H, ArH), 6.74 (d, J = 2.6 Hz, 1 H, ArH), 6.81 (d,
acid 29 (3.66 g, 74%) as a white solid; m.p. 210–214 °C (decomp.); J = 8.6 Hz, 1 H, ArH), 7.41 (dd, J = 11.9, 8.9 Hz, 1 H, CH=CH),
Rf (TLC) = 0.56 (n-BuOH/EtOH/AcOH/H2O, 4:1:1:1). 1H NMR 9.40 (br. d, J = 11.9 Hz, 1 H, NH) ppm. 13C NMR (76 MHz,
2
3
(300 MHz, [D6]DMSO/CF3CO2D, 6:1): δ = 2.82 (dd, J = 16.6, J
= 7.3 Hz, 1 H, CHAHB), 2.96 (dd, 2J = 16.6, 3J = 7.1 Hz, 1 H,
CHAHB), 3.77 (s, 3 H, OCH3), 4.51 (br. dd, 1 H, CHNH2), 5.06 (s,
[D6]DMSO): δ = 28.2 [C(CH3)3], 56.8 (OCH3), 83.1 [C(CH3)3], 93.6
(CH=CH), 107.2 (CH), 114.7 (CH), 115.3 (CH), 143.2 (CH=CH),
144.3 (C), 144.9 (C), 148.4 (C), 152.7 (CO2NH), 168.8
3
4
2 H, OCH2), 6.94 (dd, J = 8.3, J = 1.6 Hz, 1 H, Harom), 7.00 (d, (CO2R) ppm.
3J = 8.3 Hz, 1 H, ArH), 7.14 (d, 4J = 1.6 Hz, 1 H, ArH), 7.25–7.40
(E)-32: Yellow oil; Rf (TLC) = 0.27 (hexanes/EtOAc, 2:1). 1H NMR
(m, 5 H, Ph) ppm. 13C NMR (76 MHz, [D6]DMSO/CF3CO2D,
6:1): δ = 39.4 (CH2), 51.7 (CHNH), 56.4 (OCH3), 70.9 (OCH2),
112.5 (CH), 114.4 (CH), 120.8 (CH), 128.6 (CH), 128.7 (CH), 129.3
(CH), 130.3 (C), 137.9 (C), 149.2 (C), 150.3 (C), 171.9
(CO2H) ppm. MS (ESI+): m/z (%) = 302 (2) [M + H]+, 286 (18)
[M – CH3]+, 285 (100) [M – NH2]+, 267 (1), 253 (4), 240 (3), 225
(7), 207 (6), 132 (3). HRMS: Calcd. for C17H20NO4 [M + H]+
302.1392; found 302.1388. C17H19NO4 (301.34): calcd. C 67.76, H
6.36, N 4.65; found C 67.43, H 6.31, N 4.11.
(300 MHz, CD3CN): δ = 1.48 (s, 9 H, tBu), 3.82 (s, 3 H, OCH3),
5.52 (d, J = 13.9 Hz, 1 H, CH=CH), 6.44 (s, 1 H, OH), 6.55 (dd,
J = 8.5, 2.6 Hz, 1 H, ArH), 6.72 (d, J = 2.6 Hz, 1 H, ArH), 6.80
(d, J = 8.5 Hz, 1 H, ArH), 7.81 (dd, J = 13.9, 11.8 Hz, 1 H,
CH=CH), 8.13 (br. d, J = 11.8 Hz, 1 H, NH) ppm. 13C NMR
(75.5 MHz, [D6]DMSO): δ = 28.2 [C(CH3)3], 56.8 (OCH3), 82.7
[C(CH3)3], 97.8 (CH=CH), 107.3 (CH), 114.8 (CH), 115.3 (CH),
142.7 (CH=CH), 144.6 (C), 144.9 (C), 148.4 (C), 153.0 (CONH),
167.4 (CO2R) ppm. MS (EI): m/z (%) = 309 (10) [M+], 254 (3) [M –
C4H7]+, 236 (6) [M – C4H9O]+, 170 (15) [C8H12NO3]+, 140 (100)
[C7H8O3]+, 125 (8) [C6H5O3]+, 114 (44) [C4H4NO3]+, 96 (7)
[C5H4O2]+, 85 (1), 70 (22) [C3H4NO]+, 59 (3), 57 (30), 43 (2), 41
(5). MS: Calcd. for C15H19NO6 [M+] 309.1212; found 309.1204.
3-(tert-Butoxycarbonylamino)-3-(4-hydroxy-3-methoxyphenyl)propa-
noic Acid (30): To a solution of 29 (3.40 g, 11.3 mmol) in 98%
AcOH (40 mL) was added a spatula tip of 10% Pd/C under an
argon atmosphere. The mixture was transferred to an autoclave and
hydrogenated with stirring at 20 °C and 50 bar for 18 h. The cata-
lyst was removed by filtration through a Celite® pad and the sol-
vent was removed under reduced pressure to afford the acetate of
3-amino-3-(4-hydroxy-3-methoxyphenyl)propanoic acid (4.03 g,
quant.) as a yellowish, viscous oil. The crude product (3.87 g,
10.8 mmol) was dissolved in dioxane/H2O (1:1, 50 mL) and NEt3
(4.44 g, 43.9 mmol), cooled to 0 °C, and treated with Boc2O (3.50 g,
16.0 mmol). The solution was stirred for 30 min at 0 °C and for a
further 19 h at ambient temperature. The solvent was removed un-
der reduced pressure and, after addition of H2O (40 mL), the solu-
tion was adjusted to pH 8 with 2 n NaOH. The mixture was
washed with EtOAc (2ϫ 15 mL) and the aqueous phase was ad-
justed to pH 1–2 with 2 n HCl. The Boc derivative was extracted
3,4-Bis(benzyloxy)phenyl Methyl Malonate (34): To a solution of
3,4-bis(benzyloxy)phenol (33)[12] (1.94 g, 6.34 mmol) in anhydrous
CH2Cl2 (35 mL) were added methyl 3-chloro-3-oxopropanoate
(1.06 g, 7.76 mmol) and NEt3 (0.90 g, 8.89 mmol) at 0 °C, and the
mixture was stirred at 0 °C for 30 min. The ice bath was removed
and the mixture was stirred overnight at room temperature. The
solution was concentrated under reduced pressure, H2O (30 mL)
was added, and the mixture was extracted with EtOAc (3ϫ 30 mL).
The combined organic phases were washed with saturated aq.
NaHCO3 (2ϫ 30 mL) and brine (30 mL), dried (MgSO4), and con-
centrated under reduced pressure. The yellow residue was purified
by chromatography on silica gel (CH2Cl2) to yield malonate 34
Eur. J. Org. Chem. 2012, 380–390
© 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
389

W. Steglich et al.
FULL PAPER
(2.17 g, 84%) as a waxy, yellowish solid; m.p. 56–59 °C; Rf (TLC)
Acknowledgments
1
= 0.61 (CH2Cl2). H NMR (300 MHz, CDCl3): δ = 3.58 (s, 2 H,
CH2), 3.80 (s, 3 H, OCH3), 5.13 (s, 4 H, OCH2), 6.66 (dd, 3J = 8.7,
The authors thank the Deutsche Forschungsgemeinschaft (DFG)
and the Fonds der Chemischen Industrie for financial support. We
are grateful to Ms. C. Dubler for NMR measurements, Dr. W.
Spahl and Mr. R. Seidl for the mass spectra, Mr. H. Hartl for the
AA spectra, and Prof. G. Wanner, Botanical Institute of the L. M.
University, for the EDX spectra. Furthermore, we thank Ms. C.
Gräf for technical assistance and Dr. K. Greenfield, Brisbane, Aus-
tralia, for valuable suggestions and improving the manuscript. Fi-
nally, Professor Dirk Trauner is thanked for his continuous support
and stimulating discussions.
4
4J = 2.6 Hz, 1 H, ArH), 6.76 (d, J = 2.6 Hz, 1 H, ArH), 6.92 (d,
3J = 8.7 Hz, 1 H, ArH), 7.30–7.45 (m, 10 H, Ph) ppm. 13C NMR
(76 MHz, CDCl3): δ = 41.3 (CH2), 52.7 (OCH3), 71.4 (OCH2), 71.8
(OCH2), 108.6, 113.5, 115.5 (each C-ArH), 127.3, 127.4, 127.85,
127.94, 128.48, 128.52 (each CHbenzyl), 136.7, 137.1 (each Cbenzyl),
144.5, 147.0, 149.6 (each Carom), 165.2 (CO2R), 166.6 (CO2R) ppm.
MS (EI): m/z (%) = 406 (3) [M+], 306 (10) [M – C4H4O3]+, 215 (8)
[M – C11H11O3]+, 200 (4), 181 (3), 122 (3), 107 (3), 100 (25), 91
(100), 77 (3), 69 (59), 65 (8), 51 (2), 44 (4), 41 (7). HRMS: Calcd.
for C24H22O6 [M+] 406.1416; found 406.1419. C24H22O6 (406.43):
calcd. C 70.92, H 5.46; found C 70.99, H 5.60.
[1] F. von Nussbaum, P. Spiteller, M. Rüth, W. Steglich, G.
Wanner, B. Gamblin, L. Stievano, F. E. Wagner, Angew. Chem.
1998, 110, 3483–3485; Angew. Chem. Int. Ed. 1998, 37, 3292–
3295.
[2] Blue anions of a similar type are formed by oxidation of hy-
droxypulvinic acids, causing the blueing of Boletus erythropus
and related boletes on bruising the fruit bodies. See: M. Gill,
W. Steglich, Progr. Chem. Org. Nat. Prod. 1987, 51, 36–37, and
references cited therein.
3,4-Dihydroxyphenyl Methyl Malonate (35): An autoclave was
charged with a solution of malonate 34 (0.50 g, 1.23 mmol) in
MeOH (20 mL) and, after addition of a spatula tip of 10% Pd/C,
stirred for 18 h with hydrogen at room temperature at 20 bar. The
mixture was filtered through a Celite® pad and the solvent was
removed under reduced pressure. The grey residue was purified by
chromatography on silica gel (CHCl3/MeOH, 10:1) to yield 35
(0.25 g, 90%) as a reddish, viscous oil. Rf (TLC) = 0.46 (CHCl3/
MeOH, 10:1). 1H NMR (300 MHz, CD3CN): δ = 3.59 (s, 2 H,
[3] E. Jägers, B. Steffan, R. von Ardenne, W. Steglich, Z. Natur-
forsch. C 1981, 36, 488–489.
[4] In ref.[3] the configuration of the olefinic double bond in dihy-
drocortiviol A (3) (J = 13 Hz) was wrongly assigned as E.
[5] W. C. Agosta, J. Org. Chem. 1965, 30, 2490–2491.
[6] E. Wille, W. Lüttke, Chem. Ber. 1973, 106, 3240–3257. Accord-
ing to the nomenclature of these authors, the name
"oxaphthalaurin" can be proposed for this new class of pig-
ments.
3
4
CH2), 3.74 (s, 3 H, OCH3), 6.49 (dd, J = 8.5, J = 2.7 Hz, 1 H),
6.61 (d, 4J = 2.7 Hz, 1 H), 6.82 (d, 3J = 8.5 Hz, 1 H) ppm. 13C
NMR (76 MHz, CD3CN): δ = 41.9 (CH2), 53.2 (OCH3), 110.0,
113.7, 116.1 (each CHarom), 143.6, 144.6, 145.9 (each Carom), 166.8,
168.0 (each CO2R) ppm. MS (EI): m/z (%) = 226 (10) [M+], 195
(3) [M – CH3O]+, 152 (2), 126 (100) [C6H6O3]+, 109 (2), 107 (2),
101 (5), 100 (4), 97 (3), 80 (5), 69 (11), 59 (6), 52 (3), 42 (2). HRMS:
Calcd. for C10H10O6 [M+] 226.0477; found 226.0480.
[7] a) P. Chovin, Bull. Soc. Chim. Fr. 1944, 11, 82–90, b) J. N. Chat-
terjea, J. Indian Chem. Soc. 1959, 36, 69–75, c) P. Nesvadba,
Synthesis 2000, 355–356, and references cited therein.
[8] R. von Ardenne, W. Steglich, Z. Naturforsch. C 1974, 29, 446.
[9] For similar conversions of 1,2,4-trihydroxybenzenes, hydroxy-
benzoquinones and hydroxydibenzoquinones into dilactones,
see, for example: a) T. Posternak, R. G. Huguenin, W. Alcalay,
Helv. Chim. Acta 1956, 39, 1564–1579, b) E. Jägers, W. Steg-
lich, Angew. Chem. 1981, 93, 1105, Angew. Chem. Int. Ed. Engl.
1981, 20, 1016–1017, c) F. Kiuchi, H. Takashima, Y. Tsuda,
Chem. Pharm. Bull. 1998, 46, 1229–1234, d) P. Lin, S. Li, S.
Wang, Y. Yang, J. Shi, J. Nat. Prod. 2006, 69, 1629–1632, e)
M. Lang, A. Mühlbauer, E. Jägers, W. Steglich, Eur. J. Org.
Chem. 2008, 3544–3551, and references cited therein.
[10] P. T. Anastas, R. Stevenson, J. Nat. Prod. 1991, 54, 1687–1691.
[11] Ester 36 has been obtained previously by cleavage of bisnor-
bovilactone[9b] with 4% KOH in MeOH, see: B. Sontag, Disser-
tation, Ludwig-Maximilians-Universität (LMU), München,
Germany, 1999.
Ester Analogue 36 of Cortiviolate B: To a solution of malonate 35
(30 mg, 0.13 mmol) in acetonitrile (50 mL) was added a solution of
K2CO3 (40 mg, 0.29 mmol) in H2O (2 mL). The solution rapidly
turned dark-red and was stirred in an open beaker for 3 h. The
mixture was concentrated under reduced pressure, with the tem-
perature being maintained below 20 °C. The residue was triturated
with MeOH and the solution was purified by chromatography on
a Sephadex® LH-20 column (MeOH) to afford hydroxyquinone
methide anion 36[11] (0.9 mg, 3%) as a blue solid. UV/Vis (MeOH):
λmax (logε) = 222 (4.19), 253 (sh, 3.92), 349 (3.92), 586 (3.52) nm.
1H NMR (600 MHz, CD3OD): δ = 3.76 (s, 3 H, OCH3), 5.75 (s, 1
H), 6.20 (s, 1 H) ppm.
Crystallographic Data for 9: C12H6O4, M = 214.17, crystal dimen-
sions: 0.53ϫ0.40ϫ0.40 mm, monoclinic, space group P21/n, a =
9.543(3), b = 14.844(2), c = 13.788(5) Å, α = 90°, β = 105.51(3)°, γ
= 90°, V = 1882.1(9) Å3, Z = 8, Dcalcd. = 1.512 g/cm3, F(000) =
880, μ = 0.116 mm–1, number of measured reflections: 3199, 2942
were independent (Rint = 0.0114). Absorption correction, max and
min transmissions 0.9992 and 0.9799, GOF on F2 = 1.165. Finally
refined parameters [IϾ2σ(I)]: R1 = 0.0571, wR2 = 0.1056, R indices
(all data) R1 = 0.0993, wR2 = 0.1237, max./min. residual electron
density: 0.163 and –0.214 e/Å3. Structure solution with SHELXS-
86,[16] refinement with SHELXTL PLUS. [17]
[12] a) K. Aihara, Y. Urano, T. Higuchi, M. Hirobe, J. Chem. Soc.
Perkin Trans. 2 1993, 2165–2170, b) M. Miyake, Y. Hanaoka,
Y. Fujimoto, Y. Sato, N. Takemoto, I. Yokota, Y. Yoshiyama,
Heterocycles 1996, 43, 665–674, c) C. C. Li, Z. X. Xie, Y. D.
Zhang, J. H. Chen, Z. Yang, J. Org. Chem. 2003, 68, 8500–
8504.
[13] M. Iinuma, T. Tanaka, F. Asai, Phytochemistry 1994, 36, 941–
943.
[14] E. Kováts, Helv. Chim. Acta 1958, 41, 1915–1932.
[15] W. M. Rodionow, E. A. Postovskaja, J. Am. Chem. Soc. 1929,
51, 841–847.
[16] G. M. Sheldrick, SHELXS-86, Program for the Solution of
Crystal Structures, University of Göttingen, Germany, 1990.
[17] G. M. Sheldrick, SHELXTL PLUS, rel. 4.1, Siemens Analyti-
cal X-RAY Instruments Inc., Madison, WI, 1990.
Received: August 18, 2011
CCDC-271801 (for 9) contains the supplementary crystallo-
graphic data for this paper. These data can be obtained free of
charge from The Cambridge Crystallographic Data Centre via
www.ccdc.cam.ac.uk/data_request/cif.
Published Online: November 9, 2011
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