Total synthesis of VM55599. Utilization of an intramolecular Diels-Alder cycloaddition of potential biogenetic relevance

Article abstract of DOI:10.1021/ja993791h

The total synthesis of VM55599, a natural metabolite of Penicillium sp. IMI332995, has been achieved via an intramolecular Diels-Alder cycloaddition of a reverse isoprene moiety across an azadiene system. The diastereoselectivity of the intramolecular Diels-Alder cycloaddition has biogenetic implications and is discussed in the context of the biogenetic relationship of VM55599 to the paraherquamides.

Full text of DOI:10.1021/ja993791h

J. Am. Chem. Soc. 2000, 122, 1675-1683
1675
Total Synthesis of VM55599. Utilization of an Intramolecular
Diels-Alder Cycloaddition of Potential Biogenetic Relevance
Emily M. Stocking,† Juan F. Sanz-Cervera,‡ and Robert M. Williams*,†
Contribution from the Department of Chemistry, Colorado State UniVersity, Fort Collins, Colorado 80523
ReceiVed October 22, 1999
Abstract: The total synthesis of VM55599, a natural metabolite of Penicillium sp. IMI332995, has been achieved
via an intramolecular Diels-Alder cycloaddition of a reverse isoprene moiety across an azadiene system. The
diastereoselectivity of the intramolecular Diels-Alder cycloaddition has biogenetic implications and is discussed
in the context of the biogenetic relationship of VM55599 to the paraherquamides.
Introduction
VM55599 is a minor secondary metabolite of Penicillium sp.
IMI332995.1 This substance was co-isolated with several known
members of the paraherquamide family including paraherqua-
mide A, VM54158 (paraherquamide G), VM54159 (paraher-
quamide E), and VM55594 (paraherquamide F) by Everett et
al.1 In addition, this Penicillium strain was found to produce
several new metabolites in the paraherquamide family including
VM55595, VM55596, and VM55597 (Figure 1).1 The relative
1
stereochemistry of VM55599 was assigned by H NMR/NOE
data but the absolute stereochemistry remains unknown; the
absolute stereostructure depicted below is a prediction based
on biogenetic considerations to be discussed below. Of particular
interest in this regard is the stereochemical disposition of the
methyl group in the â-methylproline ring which was assigned
as being syn to the bridging isoprene moiety. In all other known
members of the paraherquamide family, the methyl group in
the â-methylproline ring is disposed anti to the bridging isoprene
moiety.
VM55599,1 the paraherquamides,2 brevianamides,3 marcfor-
tines,4 and most recently, the sclerotamides,5 are indolic
secondary metabolites isolated from various fungi and have
attracted considerable attention due to their molecular complex-
ity, intriguing biogenesis,6 and some members, most notably
the paraherquamides, display potent antiparasitic activity.7 These
Figure 1.
alkaloids share the unusual bicyclo[2.2.2] ring system that has
been proposed to arise via the [4 + 2] cycloaddition of the
isoprene moiety across the R-carbons of the amino acid
units.6c,d,8,9 Previous work on the biosynthesis of these sub-
stances invoked a facial divergence in the Diels-Alder cycliza-
tion which sets the relative syn-/anti-stereochemical relationship
at this stereogenic center.10,11 Specifically, the cyclization of
the isoprenyl olefin across the azadiene ring system can proceed
via four distinct diasteromeric transition structures a, b, c, or d
(Figure 2), resulting in the four corresponding cycloadducts A,
B, C, or D. Cycloadduct B corresponds to VM55599, and
cycloadduct A is the putative structure leading to paraherqua-
† Department of Chemistry, Colorado State University.
‡ Departamento de Qu´ımica Orga´nica, Universidad de Valencia, E-46100
Burjassot, Spain.
(1) Blanchflower, S. E.; Banks, R. M.; Everett, J. R.; Reading, C. J.
Antibiot. 1993, 46, 1355.
(2) (a) Yamazaki, M.; Okuyama, E.; Kobayashi, M.; Inoue, H. Tetra-
hedron Lett. 1981, 22, 135. (b) Ondeyka, J. G.; Goegelman, R. T.; Schaeffer,
J. M.; Kelemen, L.; Zitano, L. J. Antibiot. 1990, 43, 1375. (c) Liesch, J.
M.; Wichmann, C. F. J. Antibiot. 1990, 43, 1380. (d) S. E.; Banks, R. M.;
Everett, J. R.; Manger, B. R.; Reading, C. J. Antibiot. 1991, 44, 492.
(3) (a) Birch, A. J.; Wright, J. J. J. Chem. Soc., Chem. Commun. 1969,
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A. T.; Campbell, I. M. Appl. EnViron. Microbiol. 1981, 42, 521. (e) Bird,
B. A.; Campbell, I. M. Appl. EnViron. Microbiol. 1982, 43, 345. (f) Robbers,
J. E.; Straus, J. W. Lloydia 1975, 38, 355. (g) Paterson, R. R. M.;
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J. Acta Crystallogr. 1974, B30, 2254.
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49, 8471. (e) Kuo, M. S.; Wiley: V. H.; Cialdella, J. I.; Yurek, D. A.;
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C.H.; Michael, B. F. Am. J. Vet. Res. 1992, 53, 2032. (d) Ostlind, D. A.;
Mickle, W. G.; Ewanciw, D. V.; Andriuli, F. J.; Campbell, W. C.;
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P. J.; Mrozik, H. Biochem. Pharmacol. 1992, 43, 679.
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horgne, M.; Pascard, C.; Polonsky, J. Tetrahedron Lett. 1980, 22, 1977.
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1970, 1103.
10.1021/ja993791h CCC: $19.00 © 2000 American Chemical Society
Published on Web 02/04/2000

1676 J. Am. Chem. Soc., Vol. 122, No. 8, 2000
Stocking et al.
mide; cycloadducts C and D lead to C-20-epi metabolites thus
far not detected in paraherquamide-producing fungi.
In addition, recent theoretical work on an indoxyl-based
Diels-Alder cyclization pathway supported the observed isomer
distribution of the brevianamides in Penicillium breVicompactum
which produces brevianamide A as the major metabolite and
brevianamide B as the minor metabolite, both of which possess
the anti-relationship.12 As part of a program directed primarily
at elucidating the biosynthetic mechanism of formation of the
unique bicyclo[2.2.2] ring system, particularly with respect to
the question of possible enzymatic catalysis of this reaction,
we report here the first total synthesis of VM55599 using an
intramolecular Diels-Alder cyclization reaction that may be
of biogenetic relevance.13
Results and Discussion
The synthesis of VM55599 was accomplished as shown in
Scheme 1. The benzophenone imine 1 of glycine ethyl ester
was condensed with the dimethylallylated gramine derivative
214 in the presence of tri-n-butylphosphine15 in acetonitrile to
furnish the tryptophan derivative 3 in 70% yield. Cleavage of
the benzophenone imine with hydroxylamine provided the amino
ethyl ester 4 in high yield. Subsequent t-BOC protection and
basic hydrolysis of the ethyl ester furnished the acid 5 in 78%
yield over two steps. Coupling of acid 5 with racemic â-methyl-
â-hydroxyproline ethyl ester with BOP reagent16 provided the
desired dipeptide 7 in 70-83% yield. The BOC group was
cleaved with TFA, and the resulting amino ethyl ester was
(9) For reviews on biosynthetic Diels-Alder reactions, see: (a) Tarasow,
T. M.; Eaton, B. E. CMLS Cell Mol. Life Sci. 1999, 1463 (b) Laschat, S.
Angew. Chem., Int. Ed. Engl. 1996, 35, 289. (c) Stipanovic, R. D. EnViron.
Sci. Res. 1992, 44, 319. For specific references on Diels-Alder reactions
proposed in natural products biosynthesis, see: (d) catharanthin: Scott, A.
I. Acc. Chem. Res. 1970, 3, 151. (e) Ikarugamycin: Ito, S.; Hirata, Y.
Tetrahedron Lett. 1972, 25, 2557. (f) Cytochalasans: Binder, M.; Tamm,
C. Angew. Chem., Int. Ed. Engl. 1973, 12, 370. (g) Ircinianin: Hofheinz,
W.; Schonholzer, P. HelV. Chim. Acta 1977, 60, 1367. (h) lachananthocar-
pone: Bazan, A. C.; Edwards, J. M., Weiss, U. Tetrahedron 1978, 34, 3005.
(i) Endiandric acids: Bandaranayake, W. M.; Banfield, J. E., Black, D. S.
C. J. Chem. Soc., Chem. Commun. 1980, 902. (j) Antibiotic X-14547A:
Roush, W. R.; Peseckis, S. M.; Walts, A. E. J. Org. Chem. 1984, 49, 3429.
(k) Mevinolin: Moore, R. N.; Bigam, G.; Chan, J. K.; Hogg, A. M.
Nakashima, T. T.; Vederas, J. C. J. Am. Chem. Soc. 1985, 107, 3694. Witter,
D. J.; Vederas, J. C. J. Org. Chem. 1996, 61, 2613. Kennedy, J.; Auclair,
K.; Kendrew, S. G.; Park, C.; Vederas, J. C.; Hutchinson, C. R. Science
1999, 284, 1368. (l) Bishordeninyl terpenes: Schroeder, D. R.; Stermitz, F.
R. Tetrahedron 1985, 41, 4309. (m) Coumarins: Dreyer, D. L. Recent AdV.
Phytochem. 1986, 20, 317 (n) Betaenone B: Harrison, P. H. M.; Cane, D.
E. Chemtracts: Org. Chem. 1988, 1, 369. (0) Sordaricin: Wu, X.; Kato,
N.; Takeshita, H. Tennen Yuki Kagobutsu Toronkai Koen Yoshishu 1988,
30, 424. (p) Esperamicins: Schreiber, S. L.; Kiessling, L. L. J. Am. Chem.
Soc. 1988, 110, 631. (q) Solanapyrone A: Oikawa, H.; Yokota, T.; Abe,
T.; Ichihara, A.; Sakamura, S.; Yoshizawa, Y.; Vederas, J. C. J. Chem.
Soc., Chem. Commun. 1989, 1282 and Oikawa, H.; Suzuki, Y.; Katayama,
K.; Naya, A.; Chiaki, S.; Ichihara, A. J. Chem. Soc., Perkin Trans 1 1999,
1225 and references therein. (r) Kuwanon J and chalcomoracin: Hano, Y.
Nomura, T.; Ueda, S. Chem. Pharm. Bull. 1989, 37, 554. Hano, Y.; Nomura,
T.; Ueda, S. Heterocycles 1999, 51, 231. (s) Plagiospirolides C, D and E:
Spoerle, J.; Becker, H.; Gupta, M. P.; Veith, M.; Huch, V. Tetrahedron
1989, 45, 5003. (t) Eudesmanolides: Marco, J. A.; Sanz, J. F.; Falco, E.
Tetrahedron 1990, 46, 7941. Manzamines: Baldwin, J. E.; Whitehead, R.
C. Tetrahedron Lett. 1992, 33, 2059. Baldwin, J. E. et al. Angew. Chem.,
Int. Ed. 1998, 37, 2661 and references therein. (u) Theonellastreone and
conicasterone: Kobayashi, M.; Kawazoe, K., Katori, T.; Kitagawa, I. Chem.
Pharm. Bull. 1992, 40, 1773. (v) Nargenicin: Cane, D. E.; Yang, C.-C. J.
Am. Chem. Soc. 1984, 106, 784. Cane, D. E.; Tan, W.; Ott, W. R. J. Am.
Chem. Soc. 1993, 115, 527. (w) Torreyanic acid: Lee, J. C.; Strobel, G. A.;
Lobovsky, E.; Clardy, J. J. Org. Chem. 1996, 61, 3232. (x) Cochleamy-
cins: Shindo, K.; Sakakibara, M.; Kawai, H.; Seto, H. J. Antibiot. 1996,
49, 249. (y) Wistarin: Fontana, A.; Fakhr, I.; Mollo, E.; Cimino, G.
Tetrahedron: Asymmetry 1999, 10, 3869. (z) bisorbicillinol: Nicolaou, K.
C.; Simonsen, K. B.; Vassilikogiannakis, G.; Baran, P. S.; Vidali, V. P.;
Pitsinos, E. N.; Couladouros, E. A. Angew. Chem., Int. Ed. 1999, 38, 3555
and references therein.
Figure 2.
cyclized to the corresponding piperazinedione 8 in the presence
of 2-hydroxypyridine in refluxing toluene in excellent yield.
Treatment of 8 with thionyl chloride in pyridine furnished
the unsaturated substance 9 in 75% yield. Subsequent treatment
of 9 with trimethyloxonium tetrafluoroborate in dichloromethane
provided the azadiene 10 in 72% yield. Treatment of azadiene
10 with KOH in aqueous methanol effected tautomerization to
the labile incipient azadiene 11 which spontaneously suffered
intramolecular Diels-Alder cycloaddition at room temperature
to give a mixture of all four possible racemic cycloadducts 12-
15 in 78% combined yield in a 3.7:2.6:1.6:1 ratio, respectively.
(10) The syn/anti relationship refers to the relative stereochemistry
between the C-20 stereogenic center (VM55599 numbering) and the cyclic
amino acid residue (proline, â-methylproline, or pipecolic acid):
(11) Williams, R. M.; Kwast, E.; Coffman, H,; Glinka, T. J. Am. Chem.
Soc. 1989, 111, 3064.
(12) Domingo, L. R.; Sanz-Cervera, J. F.; Williams, R. M.; Picher, M.
T.; Marco, J. A. J. Org. Chem. 1997, 62, 1662.
(13) (a) Williams, R. M.; Sanz-Cervera, J. F.; Sancenon, F., Marco, J.
A.; Halligan, K. J. Am. Chem. Soc. 1998, 120, 1090. (b) Williams, R. M.;
Sanz-Cervera, J. F.; Sancenon, F., Marco, J. A.; Halligan, K. Bioorg. Med.
Chem. 1998, 6, 1233.
(14) This substance was prepared according to ref 6d.
(15) (a) Kametani, T.; Kanaya, N.; Ihara, M. J. Chem. Soc., Perkin Trans
1 1981, 959. (b) Somei, M.; Karasawa, Y.; Kaneko, C. Heterocycles 1981,
16, 941.
(16) BOP ) (benzotriazol-1-yloxy)tris(dimethylamino)phosphonium
hexafluorophosphate (purchased from Aldrich Chemical Co.).

Total Synthesis of VM55599
J. Am. Chem. Soc., Vol. 122, No. 8, 2000 1677
Scheme 1
The Diels-Alder cycloadducts 12-15 were separable by
PTLC on silica gel, and their relative stereochemistry was
1
assigned by H NMR NOE studies.17 The syn-stereochemistry
at C-20 for 12 and 13 was assigned based on the NOE between
H-20 (using the VM55599 numbering system) and the OMe of
the lactim ether. The anti-stereochemistry assignment of C-20
for both 14 and 15 was made based on the NOE between H-23
and the OMe. The assignment of stereochemistry at C-14 for
12 and 14 was deduced from the NOE between H-17 and H-19/
19′. This NOE was also observed by Everett et al. in the original
VM55599 isolation paper.1 For 13 and 15, the stereochemical
assignment of C-14 was inferred from the NOE between H-14
and H-19/19′.
The structures of all four cycloadducts 12-15 depicting their
relative stereochemistries are shown in Figure 3. The syn/anti
relationship10 at the C-20 stereogenic center was 2.4:1 and is
consistent with results reported earlier from this laboratory on
a simpler system lacking the methyl group in the proline ring.13
Of significant interest was the unexpected observation that the
major products (12 and 14) in each diastereomeric subset
displayed the methyl group in the â-methylproline ring syn to
the bridging isoprene unit (see Figure 3). The diastereoselectivity
in this regard was 1.47:1 favoring the methyl group disposed
syn to the bridging isoprene moiety. Although it is reasonable
to expect modest diastereoselectivity for this Diels-Alder
cycloaddition, purely on the basis of the slight steric bias
expected to be exerted by the methyl group in the proline ring,
we anticipated a modest preference for cycloadducts that
displayed the methyl group anti to the bridging isoprene moiety.
Figure 3.
12 with dilute HCl effected cleavage of the lactim ether to the
corresponding secondary amide 16 in 85% yield (Scheme 2).
Selective reduction of this substance with excess DIBAH18 (20
equiv) provided VM55599 in 86% yield whose 1H and 13C NMR
spectral characteristics matched those published.1 The synthetic
material was subsequently utilized to guide reisolation of natural
VM55599 from cultures of Penicillium sp. IMI332995 (obtained
from the International Mycological Institute) grown in our
laboratory. The synthetic and natural specimens were found to
have identical 1H NMR spectra and TLC mobility thereby
confirming the assignment (see Supporting Information).
Confirmation of the structure for cycloadduct 12 was secured
through conversion into racemic VM55599. Thus, treatment of
(18) We thank Prof. Tohru Fukuyama for suggesting the use of excess
DIBAH for this transformation; see Fukuyama, T.; Liu, G. Pure Appl. Chem.
1997, 69, 501.
(17) Complete NOE data on cycloadducts 12-15 can be found in
Supporting Information.

1678 J. Am. Chem. Soc., Vol. 122, No. 8, 2000
Stocking et al.
Scheme 2
Scheme 5
Scheme 3
caused by compression between the methyl group disposed on
the â-face of the proline ring and the lactim ether methoxy group
that is relieved upon ring-opening to 17 and 22, respectively.
In substrates 12 and 14, where the methyl group in the proline
ring is on the R-face, the opportunity for A(1,3)-type strain is
obviated by the anti-relationship between the lactim ether group
and the methyl group. Subsequent DIBAH reduction of the
tertiary amides of compounds 18, 20, and 23 gave the corre-
sponding diastereomers of VM55599 (19, 21, and 24, respec-
tively).
The NMR spectra of the VM55599 diastereomers 19, 21, and
24 were fully consistent with the assigned structures, and
significantly, all were distinctly different from the spectra for
natural VM55599 (see Supporting Information).1
Scheme 4
A significant implication of these observations concerns the
biogenesis and absolute stereochemistry of VM55599. In
particular, it should first be noted that natural paraherquamide
derivatives containing a non-hydroxylated â-methylproline
residue, such as VM55594, VM55595, VM54159, SB203105,20
and SB200437,20 all display the methyl group at the â-position
of the proline residue anti to the bridging isoprene moiety. In
stark contrast, VM55599 is the only member of the paraher-
quamide family thus far isolated that displays the methyl group
at the â-position of the proline residue syn to the bridging
isoprene moiety. We previously demonstrated that the â-methyl-
â-hydroxyproline ring of paraherquamide A is biosynthetically
derived from L-isoleucine.21 Enzymatic hydroxylation of the (S)-
â-methylproline ring in paraherquamide A biosynthesis must
therefore occur with net retention of stereochemistry leaving
the methyl group anti to the bridging isoprene moiety. One
possible scenario constituting a unified biogenesis of paraher-
quamide A (and congeners) and VM55599 is depicted in
Scheme 6. Since Paraherquamide A and VM55599 both possess
the bicyclo[2.2.2] monoketopiperazine ring system and are co-
produced by the same fungi, it is tempting to speculate that
these substances arise via a related or common [4 + 2]
cycloaddition. Thus, if a similar Diels-Alder cyclization,
whether it be uncatalyzed or enzyme-catalyzed, is operating in
the biosynthetic construction of these metabolites, the isoprene
unit must approach the azadiene from the same face as the
methyl group in the proline ring for VM55599 (25b, Scheme
To further confirm this assignment, the three other cycload-
ducts 13-15 were similarly converted into the corresponding
C-14 and/or C-20 epimers of VM55599 (19, 21, and 24) as
shown in Schemes 3-5.19 It was interesting to observe that, in
the case of cycloadducts 13 and 15, cleavage of the lactim ether
with dilute HCl led to the production of the ring-opened amino
esters 17 and 22, respectively. These were readily cyclized to
the corresponding bicyclo[2.2.2]-containing secondary amides
18 and 23, respectively, by simply heating these substances in
toluene at reflux temperature overnight. In contrast, the lactim
ethers of both cycloadducts 12 and 14 could be cleaved to the
corresponding bicyclo[2.2.2]-containing substrates without at-
tendant ring-opening to the corresponding amino esters. It would
appear that there is A(1,3)-type strain in compounds 13 and 15
(20) Banks, R. M.; Blanchflower, S. E.; Everett, J. R.; Manger, B. R.;
Reading, C. J. Antibiot. 1997, 50, 840.
(19) Conditions for the conversion of compounds 13-15 to the VM55599
diasteromers 19, 21, and 24 were not optimized.
(21) Stocking, E.; Sanz-Cervera, J. F.; Williams, R. M.; Unkefer, C. J.
J. Am. Chem. Soc. 1996, 118, 7008.

Total Synthesis of VM55599
J. Am. Chem. Soc., Vol. 122, No. 8, 2000 1679
Scheme 6
6), whereas in paraherquamide A, Diels Alder cyclization must
occur from the face opposite to the methyl group (25a, Scheme
6). In both cases, the diastereofacial selectivity of the Diels-
Alder reaction must give the syn-relative stereochemistry at C-20
(VM55599 numbering). It is interesting to note that metabolites
possessing the anti-relative stereochemistry at C-20 have not
yet been isolated from paraherquamide-producing fungi. Since
VM55599 is a very minor metabolite of Penicillium sp.
IMI332995,22 it seems reasonable that a syn-selective Diels-
Alder reaction gives, via conformer 25a, cycloadduct 26 as a
major product that is then further metabolized to the paraher-
quamide family. The minor cycloaddition product (via con-
former 25b), after adjustment of the oxidation state at C-12,
would furnish VM55599 as a dead-end shunt metabolite with
the absolute stereochemistry depicted (predicted).
It is therefore quite interesting that the diastereofacial bias
of the Diels-Alder cycloaddition on synthetic azadiene 11 gives
a slight preponderance (1.47:1) of cycloaddition from the same
face as the methyl group in the â-methylproline ring and modest
selectivity (2.4:1) favoring the (C-20) syn-relative stereochem-
istry. These results indicate that the intrinsic facial bias of this
type of Diels-Alder cycloaddition is modest at best and that
the biological system may be subject to protein organization of
the precyclization conformers.9
also important to stress that the laboratory cyclization described
here occurs spontaneously at room temperature in water, which
indicates that the fundamental thermodynamics of this cycliza-
tion are amenable to cytosolic constraints. Thus, “catalysis” of
this type of cycloaddition may not be required biosynthetically,
but the predisposition of the precyclization conformers (ie., 25a/
25b) leading to the observed paraherquamide stereoisomers may
be a manifestation of incidental protein organization.9,24 Un-
certainties as to the oxidation state of the putative azadiene
moiety in the biosynthetic system still exist and are the subject
of ongoing investigations in these laboratories.
Experimental Section
N-(Diphenylmethylene)-2-(1,1-dimethyl-2-propenyl)-D,L-tryp-
tophan Ethyl Ester (3). N-(Diphenylmethylene)glycine ethyl ester (1)
(3.2 g, 12.0 mmol) and the gramine derivative 214 (3.2 g, 13.2 mmol)
were stirred in acetonitrile (110 mL) under argon until the solids
dissolved. Tri-n-butylphosphine, (1.5 mL, 6 mmol) was added, and the
mixture was brought to reflux temperature for 8 h. After being cooled
to room temperature, the solvent was concentrated under reduced
pressure, and the crude product was purified by flash silica gel column
chromatography (15% EtOAc/hex) to yield 3.78 g (70%) of 3 as sticky
1
yellow foam. H NMR (300 MHz, CDCl3): δ 1.29 (3H, t, J ) 7.0
Hz), 1.38 (3H, s), 1.41 (3H, s), 3.59 (2H, dd, J ) 1.1, 7.0 Hz), 4.23
(2H, m), 4.52 (1H, dd, J ) 6.2, 7.3 Hz), 5.04 (1H, dd, J ) 1.1, 10.6
Hz), 5.09 (1 H, dd, J ) 1.3, 17.2 Hz), 5.92 (1H dd, J ) 10.3, 17.2
Hz), 6.39 (2H, bs), 6.87 (1H, ddd, J ) 1, 8.3, 8.3 Hz), 7.06 (1H, ddd,
J ) 1.1, 7.5, 7.5 Hz), 7.09 (2H, dd, J ) 1.1, 7.5, 7.5 Hz), 7.30 (5 H,
m), 7.45 (1H, d, J ) 8.0 Hz), 7.60 (2H, m), 7.86 (1H, bs). 13C NMR
(75 MHz, CDCl3): δ 14.1, 27.50, 27.52, 28.7, 38.9, 60.8, 66.6, 107.2,
109.7, 111.5, 118.8, 119.3, 121.1, 127.5, 127.6, 127.7, 128.7, 129.9,
130.0, 133.8, 135.9, 139.2, 139.8, 146.0, 169.7, 172.4. IR (NaCl neat):
3405, 3057, 2972, 1731, 1621, 1597, 1575, 1489, 1462, 1446, 1286,
1245, 1185, 1069, 1029, 917, 781, 742, 697 cm-1. HRMS (FAB+):
Calcd for C31H33N2O2: 465.2542. Found 465.2541 (M + H).
Conclusion
This study confirms the structural and relative stereochemical
assignment made for VM555991 and further demonstrates that
the core bicyclo[2.2.2] ring system common to this family of
alkaloids very likely arises by a biosynthetic intramolecular
Diels-Alder cyclization from a preformed dioxopiperazine23
that subsequently undergoes oxidation to an azadiene species.
Finally, the C-20-epi-metabolites (with the anti-stereochemistry
corresponding to the brevianamides) have not yet been detected
from paraherquamide-producing fungi, and there have been no
reports on the isolation of similarly epimeric metabolites from
the brevianamide-producing Penicillium sp. Thus, in each
biosynthetic system, there appears to be complete facial
exclusivity in the construction of the bicyclo[2.2.2] ring nucleus
with respect to the relative stereochemistry set at C-20; such is
not the case for the laboratory cycloaddition reported here. It is
2-(1,1-Dimethyl-2-propenyl)-D,L-tryptophan Ethyl Ester (4). Com-
pound 3 (1.92 g, 4.27 mmol) was stirred with NH2OH‚HCl (2.25 g,
32.45 mmol) and anhydrous Na2CO3 (3.21 g, 30.31 mmol) in CH2Cl2
(17 mL) at room temperature under argon for 24 h. The solution was
acidified to pH 3 with 10% KHSO4 (aq), and the organic layer was
separated from the aqueous phase. The aqueous layer was extracted
three more times with EtOAc before it was made basic with 10% Na2-
CO3 (aq) and extracted three times with EtOAc. The combined organic
layers from the basic extract were dried over anhydrous Na2SO4, and
the solvent was evaporated under reduced pressure to give 1.03 g (80%)
(22) Our own work with Penicillium sp. IMI 332995 provided ca. 0.5
mg of VM55599 and ∼250 mg of paraherquamide A (∼1:500 ratio) from
12 L of surface culture media.
1
of 4 as a colorless oil. H NMR (300 MHz, CDCl3): δ 1.19 (3H, t, J
) 7 Hz), 1.52 (2H, bs), 1.58 (6H, s), 3.08 (1H, dd, J ) 14.3, 9.5 Hz),
(23) For relevant work, see (a) Dunkerton, L. V.; Chen, H.; McKillican,
B. P. Tetrahedron Lett. 1988, 29, 2539. (b) Fabre, J. L.; Farge, D.; James,
C.; Lave, D. Tetrahedron Lett. 1985, 26, 5447.
(24) For an analogous system, see Oikawa, H.; Katayama, K.; Suzuki,
Y.; Ichihara, A. J. Chem. Soc., Chem. Commun. 1995, 1321.

1680 J. Am. Chem. Soc., Vol. 122, No. 8, 2000
Stocking et al.
3.35 (1H, dd, J ) 14.7, 5.1 Hz), 3.86 (1H, dd, J ) 9.5, 5.1 Hz), 4.13
(2H, m), 5.18 (1 H, dd, J ) 0.7, 10.6 Hz), 5.20 (1 H, dd, J ) 0.7, 17.2
Hz), 6.16 (1H, dd, J ) 10.6, 17.2 Hz), 7.08 (1H, t, J ) 7 Hz), 7.14
(1H, t, J ) 7 Hz), 7.29 (1H, d, J ) 7 Hz), 7.58 (1H, d, J ) 7 Hz), 7.98
(1H, bs). 13C NMR (75 MHz, CDCl3): δ 14.0, 27.8, 27.9, 31.2, 39.1,
55.9, 60.8, 107.0, 110.3, 112.1, 118.6, 119.3, 112.5, 129.8, 134.1, 140.4,
146.0, 175.5. IR (NaCl neat) 3399, 3243, 3081, 3056, 2973, 1733, 1638,
1617, 1580, 1462, 1300, 1282, 1195, 1105, 1029, 917, 859, 743 cm
-1. HRMS (FAB+): Calcd for C18H25N2O2: 301.191603. Found
301.191898 (M + H).
by adding saturated NaCl solution (100 mL), extracting three times
with EtOAc, drying over anhydrous Na2SO4, and evaporating of the
solvent under reduced pressure. The crude product was purified by
Kugelrohr distillation; the product distilled at 102 °C at 1 mmHg
affording 15.64 g of the product as an oil (65% for the two steps). 1H
NMR (300 MHz, DMSO-d6, 120 °C): δ 1.23 (3H, ddd, J ) 1.6, 7.3,
7.3 Hz), 1.41 (9H, s), 2.10 (3H, s), 2.70 (2H, t, J ) 6.6 Hz), 3.44 (2H,
ddd, J ) 1.5, 6.6, 6.6 Hz), 3.92 (2H, s), 4.14 (2H, dddd, J ) 1.5, 7.0,
7.0, 7.0 Hz). 13C NMR (100 MHz, DMSO-d6, 120 °C): 13.3, 27.4,
27.6, 28.9, 41.6, 42.8, 49.0, 59.6, 78.8, 154.0, 169.1, 205.9. IR (NaCl,
neat): 3611, 3398, 2978, 2936, 1748, 1698, 1462, 1397, 1367, 1250,
1162, 1129, 1029, 894, 866, 778 cm-1. HRMS (FAB+) Calcd. for
C13H24NO5: 274.165448. Found 274.165921 (M + H).
3-Hydroxy-3-methyl-1,2-pyrrolidinedicarboxylic Acid 1-(1,1-
Dimethylethyl) 2-Ethyl Ester (6a). A solution of the N-Boc-protected
compound obtained above (5 g, 18.29 mmol) in toluene (100 mL) was
cooled to 0 °C. Solid potassium tert-butoxide (2.05 g, 1.0 equiv) was
added portionwise, and the solution was stirred under argon for 45
min at 0 °C. The reaction was quenched by the addition of ice cold
10% aqueous KHSO4 (pH ) 2-3). The organic layer was separated
from the aqueous layer, and the aqueous layer was extracted three times
with CH2Cl2. The combined organic fractions were washed with pH )
7 phosphate buffer and brine successively. The organic layer was dried
over anhydrous Na2SO4 and evaporated to give a yellowish oil. The
oil was resuspended in CH2Cl2 and extracted three times with pH )
10 Na2CO3 buffer. The combined aqueous extracts were extracted two
more times with CH2Cl2. The organic layers were washed with brine,
dried over anhydrous Na2SO4, and evaporated to give 1.89 g (38%) of
the product as an off-white amorphous solid. 1H NMR (300 MHz,
DMSO-d6, 120 °C): δ 1.23 (3H, ddd, J ) 1.0, 7.0, 7.0 Hz), 1.40 (12H,
s), 1.77 (1H, m), 1.98 (1H, m), 3.36 (1H, m), 3.48 (1H, m), 3.89 (1H,
d, J ) 2.2 Hz), 4.12 (2H, dddd, J ) 2.2, 7.0, 7.0, 7.0 Hz), 4.62 (1H,
bs). 13C NMR (100 MHz, DMSO-d6, 116 °C): δ 13.5, 26.6, 27.4, 37.6,
43.8, 59.1, 68.4, 78.2, 152.8, 168.9. IR (NaCl, neat‚): 3446, 3093, 2977,
2935,m 2900, 1743, 1681, 1456, 1403, 1367, 1161, 1097, 1033, 926,
860, 774, 739 cm1. HRMS (FAB+): Calcd for C13H24N1O5: 274.165448.
Found 274.166420 (M + H).
1-[N-[(1,1-Dimethylethoxy)carbonyl]-2-(1,1-dimethyl-2-propenyl)-
D,L-tryptophyl]-3-hydroxy-3-methyl-D,L-proline Ethyl Ester (7).
Compound 6a (1.21 g, 4.43 mmol) was stirred with TFA (6.8 mL) in
CH2Cl2 (7 mL) at 0 °C. The reaction was allowed to come to room
temperature and stir for an additional 3 h. A saturated solution of
NaHCO3 was added until the solution became basic, and the organic
layer was separated from the aqueous phase. The aqueous layer was
extracted three more times with EtOAc, the combined organic layers
were dried over anhydrous Na2SO4, and the solvent was removed under
reduced pressure. Compound 6b (â-hydroxy-â-methylproline ethyl
ester) was mixed with compound 5 (1.65 g, 4.43 mmol), BOP reagent
(1.96 g, 4.43 mmol), and Et3N (1.24 mL) in acetonitrile (67 mL) at
room temperature for 4 h. A saturated aqueous solution of NaCl was
added, and the reaction was extracted four times with EtOAc. The
combined organic layers were washed with 2 M HCl, water, 10%
NaHCO3 (aq), water, and brine successively. The organic phase was
dried over anhydrous Na2SO4 and evaporated to dryness under reduced
pressure. The product was partially purified by flash silica gel column
chromatography using 4% MeOH/CH2Cl2; the slightly impure mixture
of diastereomers 7 (1.88 g, 80%) were directly carried on to the next
step without further purification.
3-[[2-(1,1-Dimethyl-2-propenyl)-1H-indol-3-yl]methyl]hexahydro-
8-hydroxy-8-methylpyrrolo[1,2-a]pyrazine-1,4-dione (8). To a solu-
tion of 7 (527 mg, 1.00 mmol) in CH2Cl2 (2 mL) at 0 °C was added
TFA (1.6 mL). The ice bath was removed, and the mixture was allowed
to come to room temperature and stir for an additional 3 h. A saturated
solution of NaHCO3 was added until the solution became basic, and
the organic layer was separated from the aqueous phase. The aqueous
layer was extracted three more times with EtOAc, the combined organic
layers were dried over anhydrous Na2SO4, and the solvent was removed
under reduced pressure. The crude free amine was then dissolved in
toluene (5 mL) with 2-hydroxypyridine (19 mg), the solution was
refluxed overnight under argon, and the solvent was removed under
reduced pressure. The four diastereomers could be partially separated
N-[(1,1-Dimethylethoxy)carbonyl]-2-(1,1-dimethyl-2-propenyl)-
D,L-tryptophan Ethyl Ester. Compound 4 (1.57 g, 5.23 mmol) was
stirred with 1 equiv of 0.5 M NaOH and di-tert-butyl pyrocarbonate
(1.25 g, 5.75 mmol) in dioxane (5.23 mL) at room temperature for 3
h. The dioxane was removed under reduced pressure, and the solution
was brought to pH ) 2 with the addition of aqueous 10% KHSO4.
The aqueous layer was extracted three times with EtOAc and dried
over anhydrous Na2SO4. After removing the solvent under reduced
pressure, the product was purified by flash silica gel column chroma-
tography using 30% EtOAc/hexane to yield 1.843 g (88%) of the
1
product as an oil. H NMR (300 MHz, DMSO-d6, 120 °C): δ 1.04
(3H, t, J ) 7.0 Hz), 1.31 (9H, s), 1.55 (6H, s), 3.12 (1H, dd, J ) 7.7,
14.3 Hz), 3.30 (1H, dd, J ) 6.8, 14.8 Hz), 3.96 (2H, m), 4.30 (1H, dd,
J ) 7.7, 15.6 Hz), 5.07 (1H, dd, J ) 0, 10.3 Hz), 5.10 (1H, dd, J ) 0,
17.6 Hz), 6.22 (1H, dd, J ) 10.7, 17.6 Hz), 6.26 (1H, bs), 6.92 (1H,
t, J ) 7.3 Hz), 7.00 (1H, t, J ) 7.0 Hz), 7.31 (1H, d, J ) 8.0 Hz), 7.45
(1H, d, J ) 7.7 Hz), 10.06 (1H, bs). 13C NMR (100 MHz, DMSO-d6,
116 °C): δ 12.9, 26.8, 27.3, 27.5, 30.7, 38.3, 54.9, 59.4, 77.7, 105.0,
108.7, 110.0, 110.5, 117.4, 117.5, 119.7, 128.9, 134.4, 140.3, 145.9,
154.2, 171.5. IR (NaCl neat) 3376, 3083, 3057, 2976, 2933, 1697, 1503,
1462, 1376, 1167, 1021, 917, 861, 742 cm-1. HRMS (FAB+): Calcd
for C23H32N2O4: 400.236208. Found 400.236330 (M+).
N-[(1,1-Dimethylethoxy)carbonyl]-2-(1,1-dimethyl-2-propenyl)-
D,L-tryptophan (5). N-[(1,1-Dimethylethoxy)carbonyl]-2-(1,1-dimethyl-
2-propenyl)-D,L-tryptophan ethyl ester (1.97 g, 4.60 mmol) was stirred
with LiOH (589 mg, 25 mmol) in a THF:H2O solution (2:1) (16 mL)
overnight. The solution was acidified with 10% KHSO4(aq) and
extracted three times with EtOAc. The organic layer was dried over
anhydrous Na2SO4, and the solvent was concentrated under reduced
pressure to afford 1.63 g (89%) of 5 as an amorphous solid. The product
was deemed sufficiently pure to use directly for the next step without
1
further purification. H NMR (300 MHz, DMSO-d6, 120 °C): δ 1.28
(9H, s), 1.55 (3H, s), 1.56 (3H, s), 3.09 (1H, dd, J ) 8.1, 14.7 Hz),
3.46 (1H, dd, J ) 6.2, 14.7), 4.30 (1H, dd, J ) 8.1, 14.7 Hz), 5.06
(1H, dd, J ) 1.1, 10.6 Hz), 5.10 (1H, dd, J ) 0, 17.2 Hz), 5.98 (1H,
d, J ) 5.9 Hz), 6.22 (1H, dd, J ) 10.6, 17.2 Hz), 6.92 (1H, ddd, J )
1.1, 7.7, 7.7 Hz), 7.00 (1H, ddd, J ) 0.8, 7.7, 7.7 Hz), 7.31 (1H, d, J
) 7.7 Hz), 7.53 (1H, d, J ) 7.7 Hz), 10.02 (1H, bs), 11.51 (1H, very
bs). 13C NMR (100 MHz, DMSO-d6, 120 °C): δ 27.1, 27.3, 27.4, 38.3,
54.6, 77.6, 105.4, 110.1, 110.4, 117.5, 119.6, 129.0, 134.4, 140.3, 146.0,
154.2, 172.7. IR (NaCl neat) 3368-2563, 3368, 3087, 3053, 2974,
2926, 1712, 1502, 1460, 1394, 1367, 1245, 1164, 1054, 1010, 919,
742 cm -1. HRMS (FAB+): Calcd for C21H28N2O4: 372.2049. Found
372.2052 (M+).
Synthesis of 6a. N-[(1,1-Dimethyethoxy)carbonyl]-N-(3-oxobutyl)-
glycine Ethyl Ester. The hydrochloride salt of glycine ethyl ester was
neutralized by the addition of 1 equiv of aqueous 10% Na2CO3 and
extracting five times with CH2Cl2. After drying the organic layer over
anhydrous Na2SO4, the solvent was removed under reduced pressure,
and the crude free amine was obtained. Glycine ethyl ester (9.06 g,
87.8 mmol) was stirred with methyl vinyl ketone (7.28 mL, 1.0 equiv)
in acetonitrile (88 mL) at room temperature under argon in the absence
of light. After 3 h, the solvent was removed under reduced pressure,
and the flask was placed under vacuum for 1 h. The free amine
decomposes fairly rapidly upon standing, and it was found best to
proceed directly to the next step without further purification. To a
solution of the adduct obtained above (13.76 g, 79.4 mmol) at 0 °C in
dioxane (160 mL) were added di-tert-butyl pyrocarbonate (17.3 g, 1.0
equiv, 79.4 mmol), 1 M NaOH solution (79.4 mL), and deionized water
(79.4 mL). The reaction was allowed to stir under argon, in the absence
of light, at room-temperature overnight. The reaction was worked up

Total Synthesis of VM55599
J. Am. Chem. Soc., Vol. 122, No. 8, 2000 1681
using PTLC, but in practice the mixture of products was purified as a
mixture of diasteromers via radial silica gel chromatography using an
elutant of 2% MeOH/CH2Cl2 to afford 361 mg (95%) of 8 as a mixture
of diastereomers (solid). Data for two of the diastereomers, 8a and 8d,
are descibed below (relative stereochemistry not assigned); diastereo-
mers 8b and 8c could not be separated.
8a: 1H NMR (300 MHz, CDCl3): δ 1.57 (6H, s), 1.62 (3 H, s),
1.89 (1H, m), 2.18 (1H, ddd, J ) 7.3, 7.3, 13.5 Hz), 2.95 (1H, bs),
3.21, (1H, dd J ) 9.7, 15.4 Hz), 3.74 (3H, m), 3.91 (1H, d, J ) 1.5
Hz), 4.40 (1H, dd, J ) 2.2, 11.3 Hz), 5.19 (1H, dd, J ) 0, 17.2 Hz),
5.20 (1H, dd, J ) 0, 11.0 Hz), 5.81 (1H, bs), 6.15 (1H, dd, J ) 10.9,
16.8 Hz), 7.12 (1H, ddd, J ) 1.1, 7.3, 7.3 Hz), 7.19 (1H, ddd, J ) 1.5,
7.3, 7.3 Hz), 7.34 (1H, d, J ) 8.0 Hz), 7.50 (1H, d, J ) 7.7 Hz), 8.10
(1H, bs). 13C NMR (75 MHz, CDCl3): δ 25.9, 26.1, 27.8, 27.9, 36.9,
39.0, 43.3, 54.5, 65.8, 77.8, 104.5, 110.9, 112.9, 117.8, 120.1, 122.2,
128.0, 132.2, 141.5, 145.6, 166.0, 168.0. IR (NaCl neat) 3360, 3054,
2968, 2924, 1666, 1651, 1462, 1434, 1302, 1262, 1138, 1105, 1010,
919, 734 cm -1. HRMS (FAB+): Calcd. for C22H28N3O3: 382.213067.
Found 382.212574 (M + H). Rf 0.75 (eluted twice with 2% MeOH/
CH2Cl2).
8d: 1H NMR (300 MHz, CDCl3): δ 1.54 (3 H, s), 1.55 (3H, s),
2.04 (2H, m), 2.32 (1H, bs), 2.66 (3H, d, J ) 9.2 Hz), 3.25 (1H, dd,
J ) 9.5, 14.3), 3.46 (1H, dd, J ) 14.6, 3.6), 3.53 (2H, m), 3.74 (1H,
m), 4.25 (1H, m), 5.16 (1H, dd, J ) 1.1, 10.6 Hz), 5.19 (1H, dd, J )
0.8, 17.2 Hz), 6.12 (1H, dd, J ) 10.6, 17.6 Hz), 6.18 (1H, bs), 7.12
(2H, m), 7.28 (1H, dd, J ) 1.5, 6.6 Hz), 7.53 (1H, dd, J ) 1.5, 7.0
Hz), 8.09 (1H, bs). 13C NMR (300 MHz, CDCl3): δ 25.7, 27.7, 28.0,
36.6, 36.9, 39.1, 43.3, 58.2, 65.0, 77.9, 105.1, 110.5, 111.9, 118.4, 119.8,
121.9, 128.9, 134.2, 141.4, 146.0, 166.2, 167.6. IR (NaCl neat) 3340,
3084, 3044, 2970, 2924, 1671, 1658, 1461, 1447, 1372, 1327, 1198,
1138, 1009, 987, 732 cm -1. HRMS (FAB+): Calcd for C22H28N3O3:
382.213067. Found 382.211498 (M + H). Rf 0.43 (eluted twice with
2% MeOH/CH2Cl2).
5.18 (1H, dd J ) 1.1, 17.2 Hz), 6.15 (1H, dd, J ) 10.3, 17.2 Hz), 7.04
(1H, ddd, J ) 1.1, 8.1, 8.1 Hz), 7.15 (1H, ddd, J ) 1.1, 7.0, 7.0 Hz),
7.26 (1H, d, J ) 8.1 Hz), 7.65 (1H, d, J ) 7.7 Hz), 7.86 (1H, bs). 13
C
NMR (75 MHz, CDCl3): δ 13.7, 27.7, 27.8, 31.4, 34.0, 39.3, 42.8,
52.7, 64.0, 108.1, 109.9, 111.8, 118.6, 119.7, 121.1, 122.8, 124.5, 130.3,
134.1, 140.0, 146.2, 152.6, 166.3. IR (NaCl neat) 3345, 2962,2924,
1676, 1634, 1456, 1335, 1304, 1242, 1051, 917, 741 cm-1. HRMS
(FAB+): Calcd. for C23H28N3O2: 378.217697. Found 378.218152 (M
+ H).
2,3,11,12,12a,13-Hexahydro-14-methoxy-1,12,12-trimethyl-5H,6H-
5a,13a-(nitrilometheno)-1H-indolizino[7,6-b]carbazol-5-one (12-15).
Azadiene 10 (264 mg, 0.70 mmol) was stirred in MeOH (47 mL) and
20% KOH (aq) (12.6 mL) under an argon atmosphere at 0 °C. The
reaction mixture was allowed to come to room temperature and
continued to stir for 10 h. When the reaction was complete as indicated
by TLC analysis, phosphate buffer (pH ) 7) was added until the
solution was neutral. The aqueous phase was extracted three times with
EtOAc. The combined organic extracts were washed with brine, dried
over anhydrous Na2SO4, and concentrated under reduced pressure. The
mixture of diastereomers could be partially separated by flash silica
gel column chromatography (eluted with 2-4% MeOH/CH2Cl2);
however, cycloadducts 13 and 14 had to be separated by successive
PTLC (eluted with 2% MeOH/CH2Cl2). The order of elution was (from
fastest mobility to slowest mobility): 15, 14, 13, and finally 12.
Yield: 15, 23 mg; 14, 37 mg; 13, 62 mg; 12, 85 mg (78% combined
yield). Data for each is as follows:
(1S,5aR,12aR,13aS)-rel-2,3,11,12,12a,13-Hexahydro-14-methoxy-
1,12,12-trimethyl-5H,6H-5a,13a-(nitrilometheno)-1H-indolizino[7,6-
1
b]carbazol-5-one (12). H NMR (400 MHz, CDCl3): δ 1.08 (3H, s),
1.22 (3H, d, J ) 7.0 Hz), 1.31 (3H, s), 1.64-1.78 (3H, m), 2.16 (1H,
m), 2.28 (1H, dd, J ) 9.2, 5.5 Hz), 2.90 (1H, m), 3.12 (1H, d, J )
15.7 Hz); 3.26 (1H, m), 3.56 (1H, m), 3.81 (3H, s), 4.03 (1H, d, J )
15.7 Hz), 7.12 (2H, m), 7.27 (1H, d, J ) 7 Hz), 7.57 (1H, d, J ) 7
Hz), 7.74 (1H, bs). 13C NMR (100 MHz, CDCl3): δ 14.02, 22.1, 26.6,
27.7, 28.6, 32.4, 34.3, 35.2, 42.4, 46.4, 54.4, 65.7, 66.0, 106.8, 110.2,
118.8, 118.9, 121.2, 127.6, 136.5, 139.5, 171.1, 172.8. IR (NaCl,
neat): 3306, 2969, 1668, 1651, 1455, 1428, 1345, 1310, 1194, 995,
740, 714 cm-1. HRMS (FAB+) Calcd for C23H28N3O2: 378.2181.
Found 378.2176 (M + H). Rf 0.40 (eluted twice with 2% MeOH/CH2-
Cl2).
(3R,S)-3-[[2-(1,1-Dimethyl-2-propenyl)-1H-indol-3-yl]methyl]-
2,3,6,7-tetrahydro-8-methylpyrrolo[1,2-a]pyrazine-1,4-dione (9). Com-
pound 8 (535 mg, 1.40 mmol) was cooled to 0 °C in THF (5.6 mL)
under an argon atmosphere. Pyridine (226 µL, 2.0 equiv) was added,
and the solution was stirred for ∼15 min. SOCl2 (112 µL, 1.1 equiv)
was added, and the mixture was allowed to come to room temperature
over 3 h. Water was added to the reaction mixture which was then
extracted three times with EtOAc. The organic layer was dried over
anhydrous Na2SO4 and evaporated to dryness under reduced pressure.
The product was purified by flash silica gel column chromatography
using 2% MeOH/CH2Cl2 to afford 381 mg (75%) of compound 9 as a
glass. 1H NMR (300 MHz, CDCl3): δ 1.56 (6H, s), 2.02 (3H, s), 2.69
(2H, dd, J ) 9.2, 9.2 Hz), 3.21 (1H, dd, J ) 11.3, 14.6 Hz), 3.72 (1 H,
dd J ) 3.3, 14.7 Hz), 3.93 (2H, ddd, J ) 3.0, 11.7, 11.7 Hz), 4.47
(1H, d, J ) 10.6 Hz), 5.17 (1H, dd, J ) 0, 10.26 Hz), 5.18 (1H, dd, J
) 0, 17.2 Hz), 5.55 (1H, bs), 6.13 (1H, dd, J ) 10.6, 17.6 Hz), 7.11
(1H, ddd, J ) 1.1, 7.3, 7.3 Hz), 7.18 (1H, ddd, J ) 0.7, 7.3, 7.3 Hz),
(1R,5aR,12aR,13aS)-rel-2,3,11,12,12a,13-Hexahydro-14-methoxy-
1,12,12-trimethyl-5H,6H-5a,13a-(nitrilometheno)-1H-indolizino[7,6-
1
b]carbazol-5-one (13). H NMR (400 MHz, CDCl3): δ 1.02 (3H, s),
1.26 (3H, s), 1.43 (3H, d, J ) 7.0 Hz), 1.73 (1H, m), 1.86 (1H, dd, J
) 10.6, 12.5 Hz), 2.11(1H, dd, J ) 4.8, 12.5 Hz), 2.11 (1H, m), 2.29
(1H, dd, J ) 10.6, 4.8 Hz), 2.38 (1H, m), 3.12 (1H, d, J ) 16.1 Hz);
3.25 (1H, m), 3.62 (1H, m), 3.65 (3H, s), 4.06 (1H, d, J ) 16.1 Hz),
7.15 (2H, m), 7.31 (1H, d, J ) 7 Hz), 7.61 (1H, d, J ) 7 Hz), 7.73
(1H, bs). 13C NMR (100 MHz, CDCl3): δ 13.9, 22.9, 27.8, 28.4, 32.3,
32.9, 35.0, 41.1, 42.8, 47.5, 54.0, 65.8, 66.2, 106.9, 110.3, 118.8, 119.0,
121.9, 127.6, 136.5, 139.6, 172.9, 173.1. IR (NaCl, neat): 3306, 3047,
2951, 1167, 1633, 1462, 1435, 1372, 1311, 1185, 980, 743 cm-1. HRMS
(FAB+) Calcd for C23H28N3O2: 378.2181. Found 378.2163 (M + H).
Rf 0.49 (eluted twice with 2% MeOH/CH2Cl2).
7.32 (1H, d, J ) 7.7 Hz), 7.55 (1H, d, J ) 7.7 Hz), 8.07 (1H, bs). 13
C
NMR (75 MHz, CDCl3): δ 13.7, 26.8, 28.0, 30.8, 33.9, 39.1, 43.1,
57.0, 104.8, 110.7, 112.4, 118.3, 120.0, 122.0, 128.9, 134.30, 134.33,
141.5, 145.7, 158.1, 162.2. IR (NaCl neat) 3344, 3047, 2968, 1682,
1645, 1440, 1324, 1251, 1114, 1007, 917, 744 cm-1. HRMS (FAB+):
Calcd for C22H26N3O2: 364.2025. Found 364.2032 (M + H). Rf 0.2
(eluted with 2% MeOH/CH2Cl2).
(3R,S)-3-[[2-(1,1-Dimethyl-2-propenyl)-1H-indol-3-yl]methyl]-6,7-
dihydro-1-methoxy-8-methylpyrrolo[1,2-a]pyrazin-4(3H)-one (10).
A solution of 9 (257 mg, 0.7 mmol) was stirred with (CH3)3OBF4 (314
mg, 2.12 mmol, 3.0 equiv) and anhydrous K2CO3 (5 equiv, 489 mg,
3.54 mmol, 5.0 equiv) in CH2Cl2 (7 mL) for 7 h at ambient temperature
under an argon atmosphere. The reaction was poured into ice water
and extracted with CH2Cl2 three times. The combined organic layers
were washed with brine, dried over anhydrous Na2SO4, concentrated
under reduced pressure, and purified by silica gel column chromatog-
raphy (eluted with 50% EtOAc/hexanes) to yield 192 mg (72%) of the
azadiene 10 as a brittle foam. Rf 0.4 (eluted with 50% EtOAc/hexanes).
1H NMR (300 MHz, CDCl3): δ 1.61 (3H, s), 1.62 (3H, s), 1.99 (3H,
s), 2.55 (2H, m), 3.09 (1H, dd, J ) 9.2, 14.3 Hz), 3.66 (3H, s), 3.79
(3H, m), 4.59 (1H, d, J ) 7.0 Hz), 5.15 (1H, dd J ) 1.1, 10.3 Hz),
(1S,5aR,12aS,13aS)-rel-2,3,11,12,12a,13-Hexahydro-14-methoxy-
1,12,12-trimethyl-5H,6H-5a,13a-(nitrilometheno)-1H-indolizino[7,6-
1
b]carbazol-5-one (14). H NMR (400 MHz, CDCl3): δ 1.13 (3H, s),
1.19 (3H, d, J ) 7.0 Hz), 1.25 (3H, s), 1.68 (1H, m), 1.88 (1H, dd, J
) 9.8, 12.9 Hz), 1.97 (1H, dd, J ) 3.9, 12.9 Hz), 2.12 (1H, m), 2.23
(1H, dd, J ) 9.8, 3.9 Hz), 2.88 (1H, m), 3.22 (1H, m), 3.29 (1H, d, J
) 17.2 Hz), 3.65 (3H, s), 3.68 (1H, m), 3.92 (1H, d, J ) 17.2 Hz),
7.09 (2H, m), 7.27 (1H, d, J ) 7 Hz), 7.57 (1H, d, J ) 7 Hz), 7.94
(1H, bs). 13C NMR (100 MHz, CDCl3): δ 14.1, 25.2, 25.9, 27.9, 28.4,
32.0, 34.2, 35.0, 42.5, 45.0, 54.2, 65.8, 66.9, 106.1, 110.3, 118.8, 119.4,
121.3, 128.0, 136.4, 139.8, 170.8, 172.1. IR (NaCl, neat, cm-1): 3407,
3312, 3053, 2964, 1667, 1455, 1427, 1345, 1306, 1230, 1180, 1042,
995, 815, 738. HRMS (FAB+) Calcd for C23H28N3O2: 378.2181. Found
377.2109 (M - H). Rf 0.55 (eluted twice with 2% MeOH/CH2Cl2).

1682 J. Am. Chem. Soc., Vol. 122, No. 8, 2000
Stocking et al.
(1R,5aR,12aS,13aS)-rel-2,3,11,12,12a,13-hexahydro-14-methoxy-
1,12,12-trimethyl-5H,6H-5a,13a-(nitrilometheno)-1H-indolizino[7,6-
b]carbazol-5-one (15). H NMR (400 MHz, CDCl3): δ 1.15 (3H, s),
MHz, CDCl3 + 1 drop DMSO-d6): δ 12.9, 22.2, 24.5, 28.0, 29.4, 31.0,
34.5, 41.5, 43.0, 49.3, 59.8, 66.6, 103.9, 110.5, 118.6, 119.7, 121.0,
126.7, 136.4, 139.9, 169.9, 173.5. IR (NaCl, neat): 3664, 3326, 2960,
1677, 1453, 1258, 1092, 799, 703 cm-1. HRMS (FAB+) Calcd for
C22H26N3O2: 364.2025. Found 364.2023 (M + H). Rf 0.30 (eluted twice
with 4% MeOH/CH2Cl2).
1
1.25 (3H, s), 1.41 (3H, d, J ) 7.0 Hz), 1.71 (1H, m), 1.86 (1H, dd, J
) 9.9, 13.2 Hz), 1.97 (1H, dd, J ) 4.4, 13.2 Hz), 2.10 (1H, m), 2.33
(1H, m), 2.38 (1H, dd, J ) 9.9, 4.4 Hz), 3.27 (1H, d, J ) 17.2 Hz),
3.35 (1H, m), 3.57 (1H, m), 3.65 (3H, s), 3.88 (1H, d, J ) 17.2 Hz),
7.10 (2H, m), 7.28 (1H, d, J ) 7 Hz), 7.59 (1H, d, J ) 7 Hz), 7.70
(1H, bs). 13C NMR (100 MHz, CDCl3): δ 14.2, 25.3, 26.1, 28.4, 32.9,
33.6, 34.9, 40.7, 43.0, 45.8, 53.8, 66.2, 66.7, 106.4, 110.3, 118.9, 119.2,
121.5, 128.0, 136.4, 139.7, 171.1, 173.4. IR (NaCl, neat): 3407, 3310,
(1S,5aR,12aS,13aS)-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-tri-
methyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carbazole-
1
5,14-dione (20) from cycloadduct 14. Yield: 14.5 mg, (100%). H
NMR (400 MHz, CDCl3): δ 1.17 (3H, d, J ) 7.0 Hz), 1.26 (3H, s),
1.33 (3H, s), 1.66 (1H, m), 1.85 (1H, dd, J ) 3.9, 13.7 Hz), 2.02 (1H,
dd, J ) 10.1, 13.3 Hz), 2.14 (1H, m), 2.25 (1H, dd, J ) 3.9, 10.1),
2.89 (1H, d, J ) 17.9 Hz), 2.98 (1H, m), 3.26 (1H, m) 3.73 (1H, m),
3.90 (1H, d, J ) 17.6 Hz), 5.90 (1H, bs), 7.09 (1H, m), 7.15 (1H, m),
7.30 (1H, d, J ) 8 Hz), 7.50 (1H, d, J ) 7.8 Hz), 7.89 (1H, bs). 13C
NMR (100 MHz, CDCl3): δ 14.3, 23.8, 25.2, 27.3, 29.0, 31.9, 34.5,
34.6, 42.9, 44.9, 61.2, 68.2, 103.7, 110.7, 118.4, 119.7, 122.1, 127.2,
136.4, 139.6, 169.2, 173.1. IR (NaCl, neat): 3298, 2962, 1682, 1455,
1399, 1302, 1231, 742 cm-1. HRMS (FAB+) Calcd for C22H25N3O2:
363.1946. Found 363.1949 (M+). Rf 0.40 (eluted twice with 4% MeOH/
CH2Cl2).
2958, 2918, 1660, 1626, 1461, 1423, 1307, 1283, 1008, 741 cm-1
.
HRMS (FAB+) Calcd for C23H28N3O2: 378.2181. Found 378.2173,
(M + H). Rf 0.57 (eluted twice with 2% MeOH/CH2Cl2).
General Procedure for Lactim Ether Deprotection of Cyclo
Adducts (12-15). One equivalent of the lactim ether cycloadduct was
stirred in THF (0.025 M) at 0 °C. To this solution was added 0.1 M
HCl (3.0 equiv), and the reaction was stirred 5-15 min until starting
material was no longer detected by TLC analysis. The reaction was
netralized with pH ) 7 phosphate buffer and extracted three times with
EtOAc. The combined organic layers were dried over anhydrous Na2-
SO4, and the solvent was removed under reduced pressure. In the case
of cycloadducts 13 and 15, the ring opened products 17 and 22 were
obtained. In the case of cycloadduct 12, a small percentage of
conversion to the corresponding ring-opened amine methyl ester was
sometimes observed. These ring-opened amine methyl esters were
recyclized by refluxing in toluene (0.025 M) overnight. The corre-
sponding piperazinedione products (16, 18, 20, and 23) were purified
using flash silica gel column chromatography (eluted with 2-4%
MeOH/CH2Cl2). Data for each is as follows:
(1R,5aR,12aS,13aS)-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-tri-
methyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carbazole-
5,14-dione (23) from Cycloadduct 15 via 22. Yield: 4.7 mg, (49%).
Data for 22: 1H NMR (400 MHz, CDCl3): δ 1.04 (3H, d, J ) 7.0
Hz), 1.34 (3H, s), 1.46 (3H, s), 1.58 (2H, bs), 1.66 (1H, m), 1.90 (1H,
dd, J ) 9.8, 14.1 Hz), 2.02 (1H, m), 2.05 (1H, dd, J ) 9.4, 9.4), 2.18
(1H, m), 3.03 (1H, dd, J ) 9.0, 14.1 Hz), 3.13 (1H, d, J ) 16.4 Hz),
3.15 (1H, d, J ) 16.4 Hz), 3.64 (3H, s), 3.65 (1H, m), 3.73 (1H, m),
7.08 (1H, ddd, J ) 7.8, 7.8, 1.2), 7.14 (1H, ddd, J ) 7.8, 7.8, 1.2),
7.30 (1H, d, J ) 7.8 Hz), 7.52 (1H, d, J ) 7.8 Hz), 7.84 (1H, bs). 13
C
(1S,5aR,12aR,13aS)-rel-2,3,11,12,12a,13-hexahydro-1,12,12-trim-
ethyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carbazole-
5,14-dione (16) from 12. Yield: 16.5 mg (85%). In this instance, a
small percentage of the ring-opened amine methyl ester was observed
NMR (100 MHz, CDCl3): δ 14.3, 25.3, 28.2, 29.2, 30.7, 34.1, 35.0,
45.1, 45.2, 47.6, 51.9, 57.1, 68.5, 105.4, 110.5, 118.5, 119.4, 121.8,
128.1, 136.5, 139.8, 172.4, 173.3. IR (NaCl, neat): 3301, 2961, 1732,
1637, 1451, 1220, 735 cm-1. HRMS (FAB+) Calcd for C23H30N3O3:
396.2287. Found 396.2282 (M + H). Rf 0.6 (eluted with 4% MeOH/
CH2Cl2). The ring-opened amine methyl ester 22 was cyclized to the
1
by H NMR analysis and TLC. The mixture of ring-opened product
and the desired piperazinedione was refluxed overnight in toluene and
1
purifed by PTLC. H NMR (300 MHz, CDCl3): δ 1.08 (3H, s), 1.20
1
piperazinedione 23 as described above. Data for 23: H NMR (400
(3H, d, J ) 6.6 Hz), 1.33 (3H, s), 1.67 (1H, m), 1.91 (1H, dd, J ) 5.0,
13.4 Hz), 2.03 (1H, dd, J ) 10.1, 13.4 Hz), 2.16 (1H, m), 2.25 (1H,
dd, J ) 3.9, 10.1 Hz), 2.60 (1H, d, J ) 15.4 Hz), 3.01 (1H, m), 3.28
(1H, m), 3.58 (1H, m), 3.87 (1H, d, J ) 15.4 Hz) 5.84 (1H, bs), 7.11
(2H, m), 7.27 (1H, d, J ) 7.3 Hz), 7.50 (1H, d, J ) 7.7 Hz), 7.72 (1H,
bs). 13C NMR (100 MHz, DMSO-d6) δ 13.8, 21.3, 23.8, 24.8, 28.1,
31.3, 34.2, 34.7, 42.5, 48.2, 59.2, 66.9, 103.4, 110.7, 117.5, 118.2, 120.6,
126.5, 136.5, 140.7, 168.3, 173.4. IR (NaCl, neat): 3313, 3066, 2960,
MHz, CDCl3): δ 1.29 (3H, s), 1.32 (3H, s), 1.50 (3H, d, J ) 7.0 Hz),
1.82 (1H, m), 1.99 (1H, dd, J ) 10.5, 13.6 Hz), 2.10 (1H, m), 2.22
(1H, dd, J ) 3.5, 13.6 Hz), 2.31 (2H, m), 2.85 (1H, d, J ) 17.9 Hz),
3.44 (1H, m), 3.61 (1H, m), 3.87 (1H, d J ) 17.9 Hz), 5.73 (1H, bs),
7.10 (1H, t, J ) 7.4 Hz), 7.16 (1H, t, J ) 7.8 Hz), 7.30 (1H, d, J ) 7.8
Hz), 7.50 (1H, d, J ) 7.4 Hz), 7.85 (1H, bs). 13C NMR (100 MHz,
CDCl3): δ 13.6, 23.9, 25.4, 29.0, 32.5, 32.8, 34.5, 41.1, 43.3, 45.7,
61.2, 67.7, 103.7, 110.7, 118.4, 119.7, 122.2, 127.1, 136.4, 139.6, 170.2,
172.8. IR (NaCl, neat): 3228, 2925, 1684, 1670, 1570, 1453, 1406,
1291, 871, 737 cm-1. HRMS (FAB+) Calcd for C22H25N3O2: 363.1946.
Found 363.1953 (M+). Rf 0.5 (eluted with 4% MeOH/CH2Cl2).
Racemic VM55599 and Diastereoisomers (18, 19, 21). General
Procedure for DIBAH Reduction of 16, 18, 20, and 23. The
cycloadduct (16, 18, 20, or 23) (0.005 M in toluene) was stirred at 0
°C under an atmosphere of argon, and DIBAH (20 equiv as a 1.0 M
solution in toluene) was added. The reaction was allowed to come to
room temperature and stirred for 24 h. The reaction was again cooled
to 0 °C and Na2SO4‚10H2O was added slowly until bubbling subsided.
The mixture was stirred an additional 30 min and then filtered through
a fritted funnel. The solid residue was rinsed with ethyl acetate, and
the combined filtrates were evaporated under reduced pressure. The
product was isolated via flash silica gel column chromatography or
PTLC using 2% MeOH/CH2Cl2.
2913, 1681, 1455, 1404, 1257, 1187, 1088, 1022, 795, 734, 693 cm-1
.
HRMS (FAB+) Calcd for C22H25N3O2: 363.1946. Found 363.1943
(M+). Rf 0.40 (eluted twice with 4% MeOH/CH2Cl2).
(1R,5aR,12aR,13aS)-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-tri-
methyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carbazole-
5,14-dione (18) from 13 via 17. Yield: 26 mg (63%). Data for 17:
1H NMR (400 MHz, CDCl3): δ 0.95 (3H, d, J ) 6.6 Hz), 1.28 (1H,
d, J ) 13.7 Hz), 1.31 (1H, d, J ) 13.3 Hz), 1.39 (3H, s), 1.53 (1H, m),
1.56 (3H, s), 1.76 (1H, d, J ) 13.3 Hz), 2.00 (3H, m), 2.82 (1H, d, J
) 16.4 Hz), 2.88 (1H, d, J ) 13.7 Hz), 2.93 (1H, d, J ) 16.4 Hz),
3.68 (2H, m), 3.74 (3H, s), 7.02 (1H, m), 7.08 (1H, m), 7.25 (1H, d,
J ) 7.8 Hz), 7.35 (1H, d, J ) 7.8 Hz), 8.11 (1H, s). 13C NMR (100
MHz, CDCl3): δ 14.2, 28.4, 29.4, 31.5, 31.9, 33.2, 33.9, 45.2, 45.4,
47.5, 52.3, 58.2, 71.3, 103.5, 110.6, 118.1, 119.3, 121.5, 127.6, 136.3,
138.9, 172.2, 174.4. IR (NaCl, neat): 3352, 2964, 1725, 1681, 1455,
1392, 1262, 1115, 1020, 808, 761, 732 cm-1. HRMS (FAB+) Calcd
for C23H30N3O3: 396.2287. Found 396.2281 (M + H). Rf 0.30 (eluted
twice with 4% MeOH/CH2Cl2). The ring-opened amine methyl ester
17 was cyclized to the piperazinedione 18 as described above. Data
for 18: 1H NMR (400 MHz, CDCl3): δ 1.07 (3H, s), 1.29 (3H, s).
1.52 (3H, d, J ) 7 Hz), 1.80 (1H,dd, J ) 5.0, 13.5 Hz), 1.85 (1H, m),
2.09 (1H, m), 2.30 (1H, m), 2.41 (1H, dd, J ) 10.5, 13.5 Hz), 2.58
(1H, dd, J ) 5.0, 10.5 Hz), 2.59 (1H, d, J ) 15.2 Hz), 3.25 (1H, m),
3.65 (1H, m), 3.83 (1H, d, J ) 15.2 Hz), 5.95 (1H, bs), 7.08 (1H, ddd,
J ) 1.2, 7.8, 7.8 Hz), 7.13 (1H, ddd, J ) 0.8, 7.8, 7.8 Hz), 7.26 (1H,
d, J ) 7.8 Hz), 7.48 (1H, d, J ) 7.8 Hz), 7.78 (1H, bs). 13C NMR (100
(()-VM55599. Yield: 13.5 mg, (86%); obtained as an amorphous
1
powder. H NMR (400 MHz, CDCl3 + 1 drop DMSO-d6): δ 1.00
(3H, d, J ) 7.0 Hz), 1.31 (3H, s), 1.37 (1H, m), 1.39 (3H, s), 1.73
(1H, dd, J ) 11.7, 13.2 Hz), 1.96 (1H, dd, J ) 4.3, 13.2 Hz), 2.13
(3H, m), 2.24 (1H, dd, J ) 1.6, 10.1 Hz), 2.76 (1H, d, J ) 15.2 Hz),
2.90 (1H, d, J ) 15.2 Hz), 2.96 (2H, m), 3.45 (1H, d, J ) 10.1 Hz),
6.28 (1H, bs), 7.04 (1H, ddd, J ) 0.8, 7.8, 7.8 Hz), 7.11 (1H, ddd, J
) 1.2, 7.0, 7.0 Hz), 7.29 (1H, d, J ) 7.3 Hz), 7.39 (1H, d, J ) 7.8
Hz), 8.40 (1H, bs). 13C NMR (100 MHz, CDCl3 + 1 drops DMSO-
d6): δ 17.5, 24.0, 26.8, 30.1, 30.2, 30.5, 33.0, 34.2, 46.6, 53.6, 55.7,

Total Synthesis of VM55599
J. Am. Chem. Soc., Vol. 122, No. 8, 2000 1683
58.9, 66.4, 104.1, 110.6, 117.7, 119.2, 121.5, 126.8, 136.4, 141.1, 174.8.
I. R. (NaCl, neat): 3303, 3048, 2920, 1650, 1454, 1296, 779, 734, 695
cm-1. HRMS (FAB+) Calcd for C22H28N3O: 350.2232. Found
350.2235 (M + H). Rf 0.38 (eluted with 4% MeOH/CH2Cl2). This
synthetic compound was identical to natural VM55599 (obtained from
Penicillium sp. IMI332995) by TLC (silica gel, eluted with 4% MeOH/
1462, 133, 1123, 1009, 740, 702 cm-1. HRMS (FAB+) Calcd for
C22H28N3O: 350.2232. Found 350.2233 (M + H). Rf 0.17 (eluted with
4% MeOH/CH2Cl2).
(1R,5aR,12aS,13aS)-(()-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-
trimethyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carba-
zol-14-one (24). Yield: 2.4 mg (54%) obtained as an amorphous
1
1
CH2Cl2), H NMR, and 13C NMR.
powder. H NMR (400 MHz, CDCl3): δ 1.24 (3H, s), 1.29 (3H, s),
(1R,5aR,12aR,13aS)-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-tri-
methyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carbazol-
14-one (19). Yield: 19 mg (79%) obtained as an amorphous powder.
1H NMR (400 MHz, CDCl3): δ 1.30 (3H, s), 1.38 (3H, s), 1.39 (3H,
d, J ) 7.3 Hz), 1.66 (1H, m), 1.91 (3H, m), 2.19 (3H, m), 2.28 (1H,
m), 2.78 (1H, d, J ) 15.2 Hz), 2.89 (1H, d, J ) 15.2 Hz), 3.19 (1H,
m), 3.45 (1H, d, J ) 10.2 Hz), 5.91 (1H, bs), 7.08 (1H, ddd, J ) 1.2,
7.4, 7.4 Hz), 7.15 (1H, ddd, J ) 1, 7.8, 7.8 Hz), 7.30 (1H, d, J ) 7.8
Hz), 7.40 (1H, d, J ) 7.4 Hz), 7.86 (1H, bs). 13C NMR (100 MHz,
CDCl3): δ 13.0, 24.0, 29.9, 30.1, 30.5, 30.6, 34.0, 40.4, 46.3, 53.8,
56.7, 59.8, 65.5, 104.6, 110.6, 117.9, 119.5, 121.8, 126.9, 136.3, 140.8,
173.7. IR (NaCl, neat): 3305, 3060, 2924, 1667, 1455, 1368, 1261,
1.41 (3H, d, J ) 6.6 Hz), 1.89 (3H, m), 2.07 (1H, dd, J ) 4, 13 Hz),
2.07 (1H, m), 2.14 (1H, m), 2.43 (1H, m), 2.57 (1H, d, J ) 10.5 Hz),
2.77 (1H, d, J ) 17.4 Hz), 2.94 (1H, d, J ) 17.4 Hz), 3.19 (1H, d, J
) 10.5 Hz), 3.22 (1H, m), 5.48 (1H, bs), 7.09 (1H, ddd, J ) 0.8, 7.4,
7.4 Hz), 7.15 (1H, ddd, J ) 1.2, 7.6, 7.6 Hz), 7.31 (1H, d, J ) 7.8
Hz), 7.40 (1H, d, J ) 7.8 Hz), 7.81 (1H, bs). 13C NMR (100 MHz,
CDCl3 + 1 drop DMSO-d6): 11.9, 24.5, 25.3, 27.5, 27.9, 28.8, 29.5,
34.4, 37.8, 43.8, 52.7, 54.6, 59.2, 69.5, 101.5, 110.9, 117.8, 119.1, 121.6,
136.5, 140.6, 176.8. IR (NaCl, neat): 3213, 2959, 1694, 1454, 1259,
1022, 797, 702 cm-1. HRMS (FAB+) Calcd for C22H28N3O: 350.2232.
Found 350.2235 (M + H). Rf 0.23 (eluted with 4% MeOH/CH2Cl2).
Acknowledgment. This work was supported by the NIH
(Grant CA 70375 to RMW) and the DGICYT of Spain
(Proyecto PB95-1089 to JFS-C). We also acknowledge the ACS
Division of Organic Chemistry Fellowship (sponsored by
SmithKline Beecham) and the Pharmacia-Upjohn Company for
Fellowship support for Emily M. Stocking. Mass spectra were
obtained on instruments supported by the NIH Shared Instru-
mentation Grant GM49631.
1109, 1014, 801, 741, 706 cm-1
. HRMS (FAB+) Calcd for
C22H28N3O: 350.2232. Found 350.2235 (M + H). Rf 0.40 (eluted with
4% MeOH/CH2Cl2).
(1S,5aR,12aS,13aS)-(()-rel-2,3,11,12,12a,13-Hexahydro-1,12,12-
trimethyl-5H,6H-5a,13a-(iminomethano)-1H-indolizino[7,6-b]carba-
zol-14-one (21). Yield: 8.5 mg, (61%); obtained as an amorphous
powder. 1H (300 MHz, CDCl3): δ 1.02 (3H, d, J ) 7.3 Hz), 1.19 (3H,
s), 1.29 (3H, s), 1.42 (1H, m), 1.69 (1H, m), 2.14 (2H, m), 2.22 (1H,
m), 2.35 (1H, dd, J ) 8.6, 17.2 Hz), 2.67 (1H, d, J ) 10.2 Hz), 2.79
(1H, d, J ) 17.2 Hz), 2.92 (1H, d, J ) 17.2 Hz), 2.96 (1H, m), 3.06
(1H, ddd, J ) 2.5, 8.6, 8.6 Hz), 3.14 (1H, d, J ) 10.6 Hz), 5.67 (1H,
bs), 7.08 (1H, ddd, J ) 1.2, 7.4, 7.4 Hz), 7.15 (1H, m), 7.31 (1H, dd,
J ) 1.1, 7.1 Hz), 7.41 (1H, dd, J ) 1.0, 7.7 Hz), 7.87 (1H, bs). 13C
NMR (100 MHz, CDCl3): δ 17.4, 24.8, 27.3, 28.0, 29.2, 30.6, 32.7,
34.3, 45.7, 53.0, 55.2, 62.4, 65.9, 103.2, 110.8, 117.9, 119.6, 121.9,
127.0, 136.3, 141.5, 174.2. IR (NaCl, neat): 3281, 3060, 2960, 1668,
Supporting Information Available: General experimental
considerations, tables of complete 1H NMR NOE data for
cycloadducts 12-15, and 1H NMR spectra of natural and
synthetic VM55599. This material is available free of charge
via the Internet at http://pubs.acs.org.
JA993791H