The total synthesis of roquefortine C and a rationale for the thermodynamic stability of isoroquefortine C over roquefortine C

Article abstract of DOI:10.1021/ja800067q

The first total synthesis of roquefortine C is achieved by implementation of a novel elimination strategy to construct the thermodynamically unstable E-dehydrohistidine moiety. Molecular modeling studies are presented which explain the instability of the

Full text of DOI:10.1021/ja800067q

Published on Web 04/16/2008
The Total Synthesis of Roquefortine C and a Rationale for the
Thermodynamic Stability of Isoroquefortine C over
Roquefortine C
Ning Shangguan,§ Warren J. Hehre,† William S. Ohlinger,† Mary Pat Beavers,‡ and
Madeleine M. Joullié*,§
WaVefunction, Inc., 18401 Von Karman AVenue, Suite 370, IrVine, California 92612, Institute for
Medicine and Engineering, UniVersity of PennsylVania, 1026 Vagelos Research Labs,
Philadelphia, PennsylVania 19104-6383, and Department of Chemistry, UniVersity of
PennsylVania, Philadelphia, PennsylVania 19104-6323
Received January 4, 2008; E-mail: mjoullie@sas.upenn.edu
Abstract: The first total synthesis of roquefortine C is achieved by implementation of a novel elimination
strategy to construct the thermodynamically unstable E-dehydrohistidine moiety. Molecular modeling studies
are presented which explain the instability of the roquefortine C structure compared to that of isoroquefortine
C.
and Antartic sediments.14 A new member of this family,
Introduction
roquefortine E (3), has been isolated from an Australian soil
The roquefortines are a class of biologically active indole
alkaloids featuring a hexahydro[2,3-b]indole nucleus substituted
at the benzylic ring junction with a 1,1-dimethylallyl group. A
number of structurally similar indole alkaloids that belong to
the amauromine, ardeemin, or flustramine families have received
significant attention from synthetic chemists.1–5 The roquefortine
family of fungal metabolites was initially isolated from cultures
of Penicillium roqueforti obtained from soil samples, though
these compounds are also produced by other Penicillium
species.6–10 Roquefortine C was found in a variety of food
products, due to both natural occurrence and contamination.7,11–13
Roquefortine D (1) is the dihydro derivative of roquefortine C
(2) and the biosynthetic precursor to 2. More recently, both were
isolated from P. aurantiogriseum strains obtained from Artic
isolate of the ascomycete Gymnoascus reessii (Figure 1).15
Roquefortine E (3) is the first roquefortine to be isolated from
a fungus other than Penicillium. Roquefortine C possesses
bacteriostatic activity against Gram-positive bacteria and also
interacts with cytochrome P450 by binding to the heme. The
effect of roquefortine on the growth of gram-positive organisms
only occurs in those containing hemins.16–19 However, the
mechanisms of toxicity and metabolic pathway of roquefortine
are still unclear.12,20–22
An important structural feature of 2 is the configuration of
the attachment to the imidazole ring. The E-configuration of
the 3–17 double bond of roquefortine was established by
comparison of its spectral properties with those of its photoi-
somer, isoroquefortine C (4, Figure 2) that has the Z-configu-
ration. The unusual E-dehydrohistidine moiety typically under-
goes facile isomerization under acidic, basic, or photochemical
conditions.23–27 Isoroquefortine C (4) is not a natural product.
In contrast to roquefortine C, isoroquefortine C does not bind
to iron.
§ Department of Chemistry, University of Pennsylvania.
† Wavefunction, Inc.
‡ Institute for Medicine and Engineering, University of Pennsylvania.
(1) Depew, K. M.; Marsden, S. P.; Zatorska, D.; Zatorski, A.; Bornmann,
W. G.; Danishefsky, S. J. J. Am. Chem. Soc. 1999, 121, 11953.
(2) Lindel, T.; Braeuchle, L.; Golz, G.; Boehrer, P. Org. Lett. 2007, 9,
283.
The interest in roquefortine C (2) is due to its presence in all
blue cheeses and the health implications of its existence in
(3) Marsden, S. P.; Depew, K. M.; Danishefsky, S. J. J. Am. Chem. Soc.
1994, 116, 11143.
(4) Suarez-Castillo, O. R.; Sanchez-Zavala, M.; Melendez-Rodriguez, M.;
Aquino-Torres, E.; Morales-Rio, M. S.; Joseph-Nathan, P Heterocycles
2007, 71, 1539.
(14) Kozlovsky, A. G.; Zhelifonova, V. P.; Adanin, V. M.; Antipova, T. V.;
Overskaya, S. M.; Ivanushkina, N. E.; Grafe, U. Appl. Biochem.
Microbiol. 2003, 39, 393.
(5) Takase, S.; Itoh, Y.; Uchida, I.; Tanaka, H.; Aoki, H. Tetrahedron
Lett. 1985, 26, 847.
(15) Clark, B.; Capon, R. J.; Lacey, E.; Tennant, S.; Gill, J. H. J. Nat.
Prod. 2005, 68, 1661.
(6) Ohmomo, S.; Kitamoto, H. K.; Nakajima, T. J. Sci. Food Agric. 1994,
211.
(16) Aninat, C.; Andre, F.; Delaforge, M. Food Addit. Contam. 2005, 22,
361.
(7) Ohmomo, S.; Oguma, K.; Ohashi, T.; Abe, M. Agric. Biol. Chem.
1978, 42, 2387.
(17) Aninat, C.; Delaforge, M. AdV. Exp. Med. Biol. 2001, 500, 331.
(18) Aninat, C.; Hayashi, Y.; Andre, F.; Delaforge, M. Chem. Res. Toxicol.
2001, 14, 1259.
(8) Ohmomo, S.; Sato, T.; Utagawa, T.; Abe, M. Agric. Biol. Chem. 1975,
39, 1333.
(19) Delaforge, M.; Bouille, G.; Jaouen, M.; Jankowski, C. K.; Lamouroux,
C.; Bensoussan, C. Peptides 2001, 22, 557.
(9) Scott, P. M.; Kennedy, B. P. C. J. Agric. Food Chem. 1976, 24, 865.
(10) Scott, P. M.; Merrien, M.-A.; Polonsky, J. Experientia 1976, 32, 140.
(11) Cole, R. J.; Dorner, J. W.; Cox, R. H.; Raymond, L. W. J. Agric.
Food Chem. 1983, 31, 655.
(20) Arnold, D. L.; Scott, P. M.; MCGuire, P. F.; Hartwig, J.; Nera, E. A.
Food Cosmet. Toxicol. 1978, 16, 369.
(21) Ohmomo, S. J. Antibac. Antifung. Agents 1982, 10, 253.
(22) Wagener, R. E.; Davis, N. D.; Diener, U. L. Appl. EnViron. Microbiol.
1980, 39, 882.
(12) Häggblom, P. Appl. EnViron. Microbiol. 1990, 56, 2924.
(13) Möller, T.; Åkerstrand, K.; Massoud, T. Nat. Toxins 1997, 5, 86.
9
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A R T I C L E S
Shangguan et al.
Figure 1. The roquefortine family of fungal metabolites.
a total synthesis of this molecule, paving the way for a clear
toxicological assessment of this compound. Curious about the
reluctance of this compound to yield to synthesis, we also
present molecular modeling studies that confirm the stability
of the isoroquefortine C (4) structure compared to that of
roquefortine C (2), accounting for the difficulties encountered
in generating this compound. What is still unexplained is the
ubiquitous presence of roquefortine C in nature and the absence
of isoroquefortine C.
Figure 2. Isoroquefortine C (4).
human and animal food products. A report that intraperitoneal
injection of 50–100 mg per kg in male mice caused convulsive
seizures was followed by conflicting reports, but all these tests
were performed on isolated roquefortine obtained from a variety
of isolation procedures.10,12,20–22 The variability of responses
to different toxicity tests clearly indicates that the effects exerted
were not due to the presence of a single toxin but to several
substances present in variable quantities in the extracts of
different strains. In view of these results, 2 might be contami-
nated with other mycotoxins, and it was imperative that toxicity
tests should be carried out on a pure synthetic sample.
From a synthetic chemistry standpoint, roquefortine C (2) is
an attractive target for total synthesis because of its unique
structure. Therefore its synthesis was investigated in our
laboratory. Earlier efforts resulted only in preparations of
isoroquefortine C.28,29 Now for the first time, we have achieved
The two synthetic challenges presented by roquefortine C (2)
are the construction of the prenylated pyrroloindole tricyclic
structure and the E-dehydrohistidine moiety. As shown in the
retrosynthetic analysis, the pyrroloindole 5 will be synthesized
from L-tryptophan methyl ester 6 and the imidazole derivative
7 will be obtained from the methyl ester of urocanic acid 8.
The synthesis of acid 5 was accomplished according to the
method developed by the Danishefsky group.1,3 Starting from
L-tryptophan methyl ester hydrochloride 6 (Scheme 1), Boc-
protection of both nitrogens afforded 9, which was treated with
N-phenylselenyl phthalimide (NPSP)30 to form the selenenylated
pyrroloindole 10. The reaction of tryptophan derivatives with
NPSP was extensively investigated by Crich.31 Compound 10
was treated with methyl triflate, prenyl tributyl stannane and
Figure 3. Retrosynthetic analysis of roquefortine C (2).
Figure 4. Isoroquefortine C 4 and two tautomers of roquefortine C (2a and 2b) were the low energy tautomers selected for Hartree–Fock/6-31G* calculations
in Spartan 06.
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A R T I C L E S
Scheme 1. Construction of the Pyrroloindole Segment (5)
Scheme 2. Synthesis of the Imidazole Derivative (7)
Scheme 3. Route to Isoroquefortine C (4)
2,6-di-tert-butyl-4-methylpyridine to provide the alkylated
compound 11. Hydrolysis of the methyl ester afforded the acid
5.
The next step was the construction of the dehydrohistidine
fragment (Scheme 2) that began with the methyl ester of
urocanic acid 8.
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A R T I C L E S
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Scheme 4. Completion of the Synthesis of Roquefortine C (2)
Boc protection of the imidazole ring 12 was followed by
dihydroxylation to afford a racemic syn diol 13. Selective
nosylation of the more acidic R-hydroxyl group in 14 and
subsequent azide displacement of the nosylate afforded azide
15.32 Hydrogenation of the azide gave the amine 16.
acetonitrile/water mixture to afford the anti ꢀ-hydroxy-R-bromo
derivative 20 (Scheme 4). Azide displacement of the bromine
(21) and reduction of the azide provided amine 22. Coupling
between 5 and 22 by 2-(1H-9-azobenzotriazole-1-yl)-1,1,3,3-
tetramethylammonium hexafluorophosphate (HATU) afforded
23. Using EDC dehydration conditions provided the Z-isomer
19 and E-isomer 24 as a 1:1 mixture of isomers. Significant
amounts of the thermodynamically unstable E-isomer were
produced by syn elimination under these conditions. The E/Z
isomers were easily separable by silica gel chromatography.
Treatment of 24 with TMSI and then ammonia in methanol
completed the first total synthesis of roquefortine C.
Coupling of acid 5 and amine 16 under conventional
conditions using EDCI and HOBt afforded 17 in 93% yield
(Scheme 3). It was expected that an anti E2 elimination on
substrate 17 would give the desired E double bond. However,
treatment of 17 with mesyl chloride or thionyl chloride and
triethylamine provided oxazoline 18 as the product, and no
elimination occurred. A stereoselective elimination method to
generate dehydroamino acid derivatives was reported by Sai et
al.33 Under these conditions (EDC, copper chloride, 80 °C in
toluene), elimination provided only the Z-dehydroamino product
from substrate 17. Assuming a syn elimination transition state,
the Z-isomer 19 should be the major product.33 Global depro-
tection of Boc with TMSI and treatment with ammonia in
methanol provided isoroquefortine C.28
(27) Wild, J. Diplomarbeit. University of Stuttgart, Stuttgart, Germany,
1982.
(28) Schiavi, B.; Richard, D. J.; Joullie´, M. M . J. Org. Chem. 2002, 67,
620.
(29) Richard, D. J.; Schiavi, B.; Joullie´, M. M. Proc. Natl. Acad. Sci. U.S.A.
2004, 101, 11971.
(30) Nicolaou, K. C.; Claremon, D. A.; Barnette, W. E.; Seitz, S. P.
Tetrahedron 1985, 41, 4835.
To make the thermodynamically unstable E-isomer using the
same dehydration conditions, we needed the threo isomer. To
this end, protected ester 12 was treated with NBS in an
(31) Crich, D.; Huang, X. J. Org. Chem. 1999, 64, 7218.
(32) Nicolaou, K. C.; Jain, N. F.; Natarajan, S.; Hughes, R.; Solomon, M. E.;
Li, H.; Ramanjulu, J. M.; Takayanagi, M.; Koumbis, A. E.; Bando,
T. Angew. Chem., Int. Ed. 1998, 37, 2714.
(23) Horenstein, B. A.; Nakanishi, K. J. Am. Chem. Soc. 1989, 111, 6242.
(24) Kim, D.; Li, Y.; Horenstein, B. A. Tetrahedron Lett. 1990, 31, 7119.
(25) Poisel, H.; Schmidt, U. Chem. Ber. 1975, 108, 2547.
(26) Schmidt, U. G., H.; Leitenberger, V.; Lieberknecht, A.; Mangold, R.;
Meyer, R.; Riedl, B. Synthesis 1992, 487.
(33) Sai, H.; Ogiku, T.; Ohmizu, H. Synthesis 2003, 201.
(34) (a) Roothaan, C. C. J. ReV. Mod. Phys. 1951, 23, 69. (b) Hall, G. G.
Proc. R. Soc. 1951, A205, 541. c) Hehre, W. J.; Radom, L.; Schleyer,
P. R.; Pople, J. A. Ab Initio Molecular Orbital Theory; Wiley: New
York, 1985.
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A R T I C L E S
Figure 5. 3-Dimensional structural representations of isoroquefortine C, tautomer 1 of roquefortine C, and tautomer 2 of roquefortine C based on HF/6-
31G* geometry optimizations. Both isoroquefortine C (4) and tautomer (2) of roquefortine C (2b) have intramolecular H-bonds between the imidazole and
diketopiperazine rings. The atoms circled in red denote the centers (sp2, nitrogen, carbon, and hydrogen atoms) involved in the “rings” representing the
coplanar double bond networks.
that was developed to provide accurate heats of formation.35
The Spartan 06 program was employed.36
When comparing the lowest energy tautomer of roquefortine
C (2b) to isoroquefortine C (4), we find that roquefortine C is
Figure 6. Vinyl imidazole. The barrier to rotation of the exocyclic double
bond out of coplanarity was modeled using HF/6-31G*. If the double bond
rotates out of coplanarity with the conjugated imidazole ring, the penalty
is in the range of 17–23 kJ/mol, depending on the starting tautomer and
conformer.
nearly 17 kJ/mol higher in energy than isoroquefortine C,
according to the HF/6-31G* model and 15 kJ/mol higher
according to the T1 model (Tables 1 and 3). These energy
differences clearly demonstrate the stability of isoroquefortine
C over roquefortine C.
(35) T1 is an efficient procedure for calculating heats of formation of
uncharged, closed-shell molecules comprising H, C, N, O, F, S, Cl,
and Br. It follows the G3(MP2) recipe [Curtiss, L. A.; Redfern, P. C.;
Raghavachari, K.; Rassolov, V.; Pople, J. A. J. Chem. Phys. 1999,
110, 4703.] but substitutes a HF/6-31G* for the MP2/6-31G*
geometry, eliminates both the HF/6-31G* frequency and the QCISD(T)/
6-31G* energies and approximates the MP2/G3MP2Large energy
using dual basis set RI-MP2 techniques. Taken together, these changes
reduce computation time by 2–3 orders of magnitude. Atom counts,
Mulliken bond orders, and HF/6-31G* and RI-MP2 energies are
introduced as variables in a linear regression fit to a set of ∼1100
G3(MP2) heats of formation. The T1 procedure reproduces these
values with a mean absolute error of 1.8 kJ/mol. It reproduces
experimental heats of formation for a set of ∼1800 organic molecules
from the NIST thermochemical database with a mean absolute error
of 8.5 kJ/mol. Heats of formation of flexible molecules have been
approximated by the heats of formation of their lowest-energy
conformer. This has been identified by examining all conformers for
molecules with fewer than 100 conformers and by examining a random
sample of 100 conformers for molecules with more than 100
conformers. In terms of both overall error and errors for individual
systems, T1 provides a better account of the experimental thermo-
chemistry than any practical quantum chemical method that we have
previously examined. The T1 model will be included with the release
of the Spartan electronic structure program. A database of ∼40000
T1 calculations for both rigid and flexible organic molecules will also
be available (scheduled for late Summer 2008).
Figure 7. Cyclopentadiene analogues of isoroquefortine C and roquefortine
C (25 and 26, respectively). The isoroquefortine C analogue (25) is 14 kJ/
mol more stable than the roquefortine C analogue (26) from HF/6-31G*
and 9 kJ/mol from T1.
Computational studies on roquefortine C (2) and isoroque-
fortine C (4) were undertaken to understand why the synthesis
of roquefortine C was at first elusive, and why roquefortine C
so readily isomerized to isoroquefortine C photochemically.
Lowest energy conformers were established for the most stable
tautomers of isoroquefortine C and roquefortine C (Figure 4)
using molecular force field (MMFF) molecular mechanics.
Equilibrium geometries were then calculated using the Hartree-
–Fock method with the 6-31G* basis set (HF/6-31G*).34
Additionally, calculations were performed with the T1 model
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A R T I C L E S
Shangguan et al.
Figure 8. 3-Dimensional structural representations of cyclopentadienyl analogues of isoroquefortine C and roquefortine C (25 and 26) based on HF/6-31G*
geometry optimizations. The conjugated double bonds in 25 are nearly coplanar while those in 26 have bond angles that deviate from ideal sp2 geometries.
These double bond networks (in both 25 and 26) involve the exocyclic double bond and the double bond of the Cp ring that it is conjugated to.
Table 1. Relative Energies for Isoroquefortine C (4) and for the
two Tautomers of Roquefortine C (2a and 2b) Calculated by the
Hartree/Fock Method with the 6-31G* Basis Set
Table 3. Relative Energies for Isoroquefortine C (4) and for
Tautomer (2) of Roquefortine C (2b) Calculated by the T1
Method35
isomer
relative energies (kJ/mol)
Boltzmann distribution
isomer
relative energies (kJ/mol)
Boltzmann distribution
4
2a
2b
0.0
32
17
>99.5%
<0.5%
<0.5%
4
2b
0.0
15
>99.5%
<0.5%
Table 4. Relative Energies for the Cyclopentadiene Analogues (Cp
Replaces Imidazole) of Isoroquefortine C (25) and Roquefortine C
(26) Calculated by the T1 Method35
Table 2. Relative Energies for the Cyclopentadiene Analogues (Cp
Replaces Imidazole) of Isoroquefortine C (25) and Roquefortine C
(26) Calculated by the Hartree/Fock Method with the 6-31G* Basis
Set
isomer
relative energies (kJ/mol)
Boltzmann distribution
25
26
0.0
10
98%
2%
isomer
relative energies (kJ/mol)
Boltzmann distribution
25
26
0.0
14
>99.5%
<0.5%
Figure 5) is made up of only six atoms (four sp2 centers, a
nitrogen, and a hydrogen). Achieving ideal sp2 bond angles
(120°) will be more difficult for roquefortine C than for
isoroquefortine C and the calculated structures are in agreement:
roquefortine C has bond angles up to 134° while the largest
bond angle calculated for isoroquefortine C is 127°.
We contend that the difference in stability of roquefortine C
and isoroquefortine C is primarily due to the difference in the
energy penalty required to achieve coplanarity and not to the
difference in hydrogen-bond strengths. This view is supported
by additional calculations on analogues in which a cyclopen-
tadiene replaces imidazole (Figure 7). While there is no
possibility for intramolecular hydrogen bonding in these ana-
logues, the HF/6-31G* energy difference (14 kJ/mol favoring
the isoroquefortine C analogue) is nearly the same as in the
imidazole systems. The T1 energy difference is 10 kJ/mol. As
expected, double bonds in the diketopiperazine and cyclopen-
tadiene rings for 25 are nearly coplanar, and bond angles in the
roquefortine analogue 26 are further removed from ideal values
(Figure 8).
Examination of the equilibrium geometry structures for
roquefortine C (2b) and isoroquefortine C (4) reveals that both
compounds adopt conformations with an internal H-bond (Figure
5). In roquefortine C (2b), the hydrogen bonding is between a
carbonyl oxygen on the diketopiperazine ring and the NH on
the imidazole ring. In isoroquefortine C (4), the hydrogen bond
forms between an NH bond on the diketopiperazine ring and
the N on the imidazole ring. This observation suggests that the
large calculated energy difference between the two isomers (17
kJ/mol by the HF/6-31G* method and 15 kJ/mol by the T1
method) cannot be explained on the basis of hydrogen-bonding
stabilization alone.
Note that both roquefortine C and isoroquefortine C are planar
(or nearly so) with respect to the orientation of the diketopip-
erazine and imidazole rings (Figure 5). Coplanarity is necessary
for the exocyclic (E/Z) double bond and the ring (imidazole
and diketopiperazine) double bonds to conjugate. The preference
for planar arrangements in roquefortine C and isoroquefortine
C was modeled using vinyl imidazole37 (Figure 6), where,
according to the HF/6-31G* model, the barrier to rotation is in
the range of 17–23 kJ/mol, depending on the starting tautomer
and conformer. This is the same order of magnitude as the
difference in energy between roquefortine C (2b) and isoro-
quefortine C (4).
(36) Spartan 06 is available from Wavefunction, Inc., 18401 Von Karman
Avenue, Suite 370, Irvine, CA 92612; www.wavefun.com.
(37) Although tautomer 1 of roquefortine C 2a does not have the potential
to hydrogen bond, its coplanar double bond network (circled atoms
in 2a, Figure 5) also resembles a seven-membered “ring” involving
six sp2 centers and one hydrogen atom. Here too, the bond angles
deviate from ideal sp2 geometries. Since tautomer 2a does not gain
any stabilization from H-bonding as tautomer 2b does, it has a higher
relative energy (34 kJ/mol) than tautomer 2b (17 kJ/mol) by the HF/
6-31G* method.
The “ring” involving the H-bond in roquefortine C (Figure
5, 2b) is made up of seven atoms (five sp2 centers, a nitrogen
and the hydrogen atom), while the ring in isoroquefortine C (4,
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A R T I C L E S
The computational calculations are summarized in Tables
1-4. The computational studies support the notion that achiev-
ing coplanarity of the diketopiperazine and imidazole rings
requires greater distortion of the sp2 centers in roquefortine than
in isoroquefortine C. We believe that this is the major cause of
the thermodynamic differences in stability between the two
isomeric compounds.
Supporting Information Available: Experimental procedures,
characterization, and spectra for compounds 12–24, 2, 4. This
material is available free of charge via the Internet at http://
pubs.acs.org. (The coordinates for the 3D structures of roque-
fortine C, isoroquefortine C, and cyclopentadiene derivatives
25 and 26, resulting from HF/6-31G* geometry optimizations
can be found as pdb files).
Acknowledgment. We thank the NSF (CHEM-0130958) for
support of this work.
JA800067Q
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