Putative Diels-Alder-catalyzed cyclization during the biosynthesis of lovastatin

Article abstract of DOI:10.1021/jo952117p

A Diels-Alder cyclization proposed to occur during polyketide synthase assembly of the bicyclic core of lovastatin (1) (mevinolin) by Aspergillus terreus MF 4845 was examined via the synthesis of the N-acetylcysteamine (NAC) thioester of [2,11-¹³C₂]-(E,E,E)-(R)-6-methyldodecatri-2,8,10-enoate (5a). In vitro Diels-Alder cyclization of the corresponding unlabeled NAC ester 5b, ethyl ester 18b, and acid 20b yielded two analogous diastereomers in each case, under either thermal or Lewis acid-catalyzed conditions. The reaction of thioester 5 proceeds readily at 22°C in aqueous media. For 18b, one product is trans-fused ethyl (1R,2R,4aS, 6R,8aR)-1,2,4a,5,6,7,8,8a-octahydro-2,6-dimethylnaphthalene-1-carboxyl ate (30) (endo product), and the other is cis-fused ethyl (1R,2S,4aR,6R,8aR)-1,2,4a,5,6,7,8,8a-octahydro-2,6-dimethylnaphthalene -1-carboxylate (31) (exo product). Isomer 21 with stereochemistry analogous to 4a,5-dihydromonacolin L (2), a precursor of 1, was made by transformation of a tricyclic lactone, (1S,2S,4aR,6S,8S,8aS)-1-(ethoxycarbonyl)-1,2,4a,5,6,7,8,8a-octahydro-2 -methyl-6,8-naphthalenecarbolactone (22) using reduction and Barton deoxygenation. Comparison of 21 with 30 and 31 confirmed the structural assignments and showed that the nonenzymatic 4 + 2 cyclizations of 5, 18, and 20 proceed via chairlike exo and endo transition states with the methyl substituent pseudoequatorial. The proposed biosynthetic Diels-Alder leading to lovastatin (1) would require an endo conformation with the methyl substituent pseudoaxial. Intact incorporation of the labeled hexaketide triene 5a into 1 was not achieved because of rapid degradation by A. terreus cells.

Full text of DOI:10.1021/jo952117p

J . Org. Chem. 1996, 61, 2613-2623
2613
P u ta tive Diels-Ald er -Ca ta lyzed Cycliza tion d u r in g th e
Biosyn th esis of Lova sta tin
David J . Witter and J ohn C. Vederas*
Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2G2
Received November 30, 1995X
A Diels-Alder cyclization proposed to occur during polyketide synthase assembly of the bicyclic
core of lovastatin (1) (mevinolin) by Aspergillus terreus MF 4845 was examined via the synthesis
1
3
of the N-acetylcysteamine (NAC) thioester of [2,11- C
2
]-(E,E,E)-(R)-6-methyldodecatri-2,8,10-enoate
(
1
5a ). In vitro Diels-Alder cyclization of the corresponding unlabeled NAC ester 5b, ethyl ester
8b, and acid 20b yielded two analogous diastereomers in each case, under either thermal or Lewis
acid-catalyzed conditions. The reaction of thioester 5 proceeds readily at 22 °C in aqueous media.
For 18b, one product is trans-fused ethyl (1R,2R,4aS, 6R,8aR)-1,2,4a,5,6,7,8,8a-octahydro-2,6-
dimethylnaphthalene-1-carboxylate (30) (endo product), and the other is cis-fused ethyl (1R,2S,-
4
aR,6R,8aR)-1,2,4a,5,6,7,8,8a-octahydro-2,6-dimethylnaphthalene-1-carboxylate (31) (exo product).
Isomer 21 with stereochemistry analogous to 4a,5-dihydromonacolin L (2), a precursor of 1, was
made by transformation of a tricyclic lactone, (1S,2S,4aR,6S,8S,8aS)-1-(ethoxycarbonyl)-1,2,-
4
a,5,6,7,8,8a-octahydro-2-methyl-6,8-naphthalenecarbolactone (22) using reduction and Barton
deoxygenation. Comparison of 21 with 30 and 31 confirmed the structural assignments and showed
that the nonenzymatic 4 + 2 cyclizations of 5, 18, and 20 proceed via chairlike exo and endo
transition states with the methyl substituent pseudoequatorial. The proposed biosynthetic Diels-
Alder leading to lovastatin (1) would require an endo conformation with the methyl substituent
pseudoaxial. Intact incorporation of the labeled hexaketide triene 5a into 1 was not achieved
because of rapid degradation by A. terreus cells.
In tr od u ction
keto, hydroxy, olefinic, or fully reduced carbons. The
intermediates remain bound to the synthase complex
until chain assembly is complete. After release from the
protein, the product (e.g. 2) may undergo further trans-
formation (often oxidation or alkylation) by separate
enzymes to produce the final metabolite (i.e. 1). Genetic
Lovastatin (1) (also known as mevinolin, monacolin K,
Mevacor) is a widely-prescribed cholesterol-lowering drug
which was originally isolated from Aspergillus terreus by
Merck researchers and from Monascus ruber by Endo and
co-workers.1 Studies on the biosynthesis of this fungal
-3
6,7
studies and incorporations of partially-assembled in-
1
3
2
18
metabolite using incorporations of C-, H-, and O-
termediates (as their N-acetylcysteamine (NAC) thio-
labeled acetates, 13C-methionine, and
18
2
O show that it
8
esters) support this hypothesis and promise rationally-
is a polyketide having a main chain of nine intact acetate
units linked head to tail (Scheme 1).4 Examination of
late stage transformations suggests that the first isolable
intermediate and product of the lovastatin polyketide
engineered routes to a host of new biologically active
metabolites. Cell-free biosynthesis of fully constructed
natural polyketides has not been reported thus far, but
5
synthase (PKS) may be 4a,5-dihydromonacolin L (2). The
(
6) For recent reviews see: (a) Hopwood, D. A.; Sherman, D. H.
large array of structurally diverse polyketide metabolites
produced by microorganisms all originate biochemically
from repetitive connection of short chain fatty acids (e.g.
Annu. Rev. Genet. 1990, 24, 37-66. (b) Katz, L.; Donadio, S. Annu.
Rev. Microbiol. 1993, 47, 875-912. (c) Rohr, J . Angew. Chem., Int. Ed.
Engl. 1995, 34, 881-885. (d) O’Hagan, D. Nat. Prod. Rep. 1995, 12,
1
2
-32. (e) Hutchinson, C. R.; Fujii, I. Annu. Rev. Microbiol. 1995, 47,
01-238. (f) McDaniel, R.; Ebert-Khosla, S.; Hopwood, D.; Khosla, C.
acetate or propionate) by pathways that are very similar
to fatty acid biosynthesis.6
-8
However, the polyketide
Nature 1995, 375, 549-554.
(7) For some leading references to genetic studies see: (a) Wies-
synthases (PKS) can bypass particular reductive steps
to generate a functionalized carbon skeleton containing
mann, K. E. H.; Cort e´ s, J .; Brown, M. J . B.; Cutter, A. L.; Staunton,
J .; Leadlay, P. F. Chem. Biol. 1995, 2, 583-589. (b) Pieper, R.; Luo,
G.; Cane, D. E.; Khosla, C. J . Am. Chem. Soc. 1995, 117, 11373-11374.
(
c) Brown, M. J . B.; Cortes, J .; Cutter, A. L.; Leadlay, P. F.; Staunton,
X
Abstract published in Advance ACS Abstracts, March 15, 1996.
J . J . Chem. Soc., Chem. Commun. 1995, 1517-1518. (d) McDaniel,
R.; Hutchinson, C. R.; Khosla, C. J . Am. Chem. Soc. 1995, 117, 6805-
6810. (e) Yu, J .-H.; Leonard, T. J . J . Bacteriol. 1995, 177, 4792-4800.
(f) Revill, W. P.; Bibb, M. J .; Hopwood, D. A. J . Bacteriol. 1995, 177,
3946-3952. (g) Kao, C. M.; Luo, G.; Katz, L.; Cane, D. E.; Khosla, C.
J . Am. Chem. Soc. 1995, 117, 9105-9106.
(
1) Endo, A.; Hasumi K. Nat. Prod. Rep. 1993, 10, 541-550.
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Chem. Soc. 1987, 109, 1253-1255. (b) Cane, D. E.; Yang, C.-C. J . Am.
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J . Chem. Soc., Chem. Commun. 1988, 4-6. (d) Yoshizawa, Y.; Li, Z.;
Reese, P. B.; Vederas, J . C. J . Am. Chem. Soc. 1990, 112, 3212-3213.
(e) Staunton, J .; Sutkowski, A. C. J . Chem. Soc., Chem. Commun. 1991,
1110-1112. (f) Li, Z.; Martin, F. M.; Vederas, J . C. J . Am. Chem. Soc.,
1992, 114, 1531-1533. (f) Cane, D. E.; Lambalot, R. H.; Prabhakaran,
P. C.; Ott, W. R. J . Am. Chem. Soc. 1993, 115, 522-526. (g) Patzelt,
H.; Robinson, J . A. J . Chem. Soc., Chem. Commun. 1993, 1258-1260.
(h) Hailes, H. C.; J ackson, C. M.; Leadlay, P. F.; Ley, S. V.; Staunton,
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Chem. Soc. 1995, 117, 6633-6634.
(
3) Endo, A. J . Antibiot. 1979, 32, 852-854.
(4) (a) Moore, R. N.; Bigam, G.; Chan, J . K.; Hogg, A. M.; Nakashima,
T. T.; Vederas, J . C. J . Am. Chem. Soc. 1985, 107, 3694-3701. (b)
Greenspan, M. D.; Yudkovitz, J . B. J . Bacteriol. 1985, 162, 704-707.
(c) Endo, A.; Negishi, Y.; Iwashita, T.; Mizukawa, K.; Hirama, M. J .
Antibiot. 1985, 38, 444-448. (d) Shiao, M.-S.; Don, H.-S. Proc. Natl.
Sci. Counc. Repub. China [B] 1987, 11, 223-231. (e) Yoshizawa, Y.;
Witter, D. J .; Liu, Y.; Vederas, J . C. J . Am. Chem. Soc. 1994, 116,
2
693-2694.
(
5) (a) Kimura, K.; Komagata, D.; Murakawa, S.; Endo, A. J .
Antibiot. 1990, 43, 1621-1622 and references therein. (b) Treiber, L.
R.; Reamer, R. A.; Rooney, C. S.; Ramjit, H. G. J . Antibiot. 1989, 42,
3
0-36.
0
022-3263/96/1961-2613$12.00/0 © 1996 American Chemical Society

2
614 J . Org. Chem., Vol. 61, No. 8, 1996
Witter and Vederas
Sch em e 1
nonenzymatically to give racemic mixtures of endo and
exo adducts if the participating double bond is made
electron deficient by chemical oxidation of the allylic
alcohol to an aldehyde. It thus appears that biological
Diels-Alder reactions may be triggered by generation of
reactive triene systems on an enzyme surface. Applica-
tion of such reasoning to probable enzyme-bound PKS
intermediates likely to be involved in formation of 2
enroute to 1 suggests that a hexaketide triene thioester
A could undergo spontaneous 4 + 2 cyclization.4 In the
present work, we describe the synthesis of such enan-
tiomerically pure hexaketide derivatives of A and exam-
ine both their propensity to cyclize and the factors which
influence the stereochemical outcome of the nonenzy-
matic Diels-Alder process. The synthetic approach is
devised for facile formation of these hexaketides in doubly
a,e
1
3
C labeled form to test whether cultures of A. terreus to
are able to incorporate them intact into lovastatin (1).
Resu lts a n d Discu ssion
Hexaketide analogue 5 was chosen as the principal
target for cyclization as well as incorporation studies with
A. terreus cultures because N-acetylcysteamine (NAC)
thioesters are known to enhance loading of precursors
8
into PKS systems, where they exist as enzyme-bound
thioesters. Recently, R,â-unsaturated thioesters contain-
ing a diene system have been reported to undergo facile
9
i
nonenzymatic intramolecular Diels-Alder reaction. Since
â-oxidation11 can rapidly degrade labeled precursors to
short-chain fatty acids (e.g. acetate) prior to incorpora-
tion, double labeling with 13C (as in 5a ) is essential to
detect intact utilization of the entire advanced precursor
12
8
by fungal cultures. Thus the synthesis of 5 was designed
to incorporate 13C labels at opposite ends of the molecule
at a late stage in such a fashion that biological cyclization
of the intact precursor would give rise to readily-detect-
able12 coupling in the C NMR spectrum of the product-
13
(s).
Syn th esis of th e Hexa k etid e P r ecu r sor 5. Com-
mercially available (R)-citronellol (6) is a readily available
starting material that can be modified and extended in
a two directional approach (Scheme 2). Its enantiomeric
excess was determined to be g98% via derivatization to
N-[(1S)-phenylethyl]-(R)-citronellamide and subsequent
GC-MS.13 Protection of the alcohol 6 as an acetate 7,
14
in vitro formation of a triketide lactone has recently been
1
5
followed by sequential treatment with ozone and tri-
phenylphosphine, delivers the aldehyde 8. A second
protection16 with trimethyl orthoformate yields the acetal
demonstrated using a genetically-modified protein de-
7
a,b
rived from 6-deoxyerythronolide B synthase.
The stereochemistry of 2 supports the intriguing idea
1
7
9
. Hydrolysis of the acetate 9 and oxidation of the
that an enzyme-catalyzed Diels-Alder reaction may
occur during assembly of the polyketide chain.4
a,e
Such
(10) Oikawa, H.; Katayama, K.; Suzuki, Y.; Ichihara, A. J . Chem.
reactions have been proposed during biosynthesis of a
number of other metabolites,8 and recent work shows
that a partially purified oxidizing enzyme can catalyze
exo-selective formation of optically active solanapyrone
Soc., Chem. Commun. 1995, 1321-1322.
i,9
(11) For some leading references to â-oxidation enzymes see: (a)
Schulz, H. In Fatty Acids and Lipids: Biological Aspects; Galli, C.;
Simopoulos, A. P.; Tremoli, E., Eds.; Karger: Basel, Switzerland, 1994;
pp 18-21. (b) Elgersma, Y.; Vanroermund, C. W. T.; Wanders, R. J .
A.; Tabak, H. F. EMBO J . 1995, 14, 3472-3479. (c) Yang, S.-Y.; He,
X.-Y.; Schulz, H. Biochemistry 1995, 34, 6441-6447. (d) Fossa, A.;
Beyer, A.; Pfitzner, E.; Wenzel, B.; Kunau, W. H. Mol. Gen. Genet. 1995,
(
4) from an achiral precursor, prosolanapyrone II (3), with
1
0
high enantiomeric excess. The process also proceeds
2
47, 95-104.
(9) (a) Tamm, Ch. In Biosynthesis of Mycotoxins, Steyn, P. S., Ed.;
(12) For a review of labeling methods see: Vederas, J . C. Nat. Prod.
Academic Press: New York, 1980; pp 269-299. (b) Roush, W. R.;
Peseckis, S. M.; Walts, A. E. J . Org. Chem. 1984, 49, 3429-3432. (c)
Oikawa, H.; Murakami, Y.; Ichihara, A. J . Chem. Soc., Perkin Trans.
Rep. 1987, 4, 277-337.
(13) Huffer, M.; Schreier, P. J . Chromtatogr. 1990, 519, 263-267.
(14) Stork, G.; Nakahara, Y.; Nakahara, Y.; Greenlee, W. J . J . Am.
Chem. Soc. 1978, 100, 7775-7777.
1
1992, 21, 2955-2959. (d) Sanz-Cervera, J . F.; Glinka, T.; Williams,
R. M. J . Am. Chem. Soc. 1993, 115, 347-348. (e) Cane, D. E.; Tan,
W.; Ott, W. R. J . Am. Chem. Soc. 1993, 115, 527-535. (f) Oikawa, H.;
Suzuki, Y.; Naya, A.; Katayama, K.; Ichihara, A. J . Am. Chem. Soc.
(15) Clive, D. L. J .; Keshava Murthy, K. S.; Wee, A. G. H.; Siva
Prasad, J .; da Silva, G. V. J .; Majewski, M.; Anderson, P. C.; Evans,
C. F.; Haugen, R. D.; Heerze, L. D.; Barrie, J . R. J . Am. Chem. Soc.
1990, 112, 3018-3028.
(16) Wenkert, E.; Goodwin, T. E. Synth. Commun. 1977, 6, 409-
415.
1
994, 116, 3605-3606. (g) Hano, Y.; Nomura, T.; Ueda, S. Can. J .
Chem. 1994, 72, 12-14. (h) Kato, N.; Wu, X.; Nishikawa, H.; Nakan-
ishi, K.; Takeshita, H. J . Chem. Soc. Perkin Trans. 1 1994, 1047-
1
1
053. (i) Hart, D. J .; Wu, W.-L.; Kozikowski, A. P. J . Am. Chem. Soc.
995, 117, 9369-9370.
(17) Mancuso, A. J .; Huang, S.-L.; Swern, D. J . Org. Chem. 1978,
43, 2480-2482.

Diels-Alder Cyclization during Biosynthesis of Lovastatin
Sch em e 2a
J . Org. Chem., Vol. 61, No. 8, 1996 2615
a
+
Reagents: (a) AcCl, Et3N (95%); (b) O3, Ph3P (85%); (c) (CH3O)3CH, H (91%); (d) NaOMe, MeOH (89%); (e) Swern oxid (85%); (f)
+
13
-
Ph3PdCHCO2Et (78%); (g) DIBAL (80%); (h) Swern oxid (94%); (i) (Ph3P ) CH2CH3(I ) and PhLi and then PhLi, HCl, (t-BuOK/t-BuOH)
85%); (j) saturated oxalic acid, THF; (k) Ph3Pd CHCO2Et (59%); (l) KOH (74%); (m) DCC, NAC, DMAP (61%).
1
3
(
resultant alcohol 10 generates the aldehyde 11. The first
olefin of the diene system is introduced by the reaction
with Wittig reagent18 to produce a mixture of E- and
Z-isomers (87:13) (12 and 13, respectively) in 78% total
yield. Separation of the major R,â-unsaturated ester 12
from the minor Z-isomer 13 and reduction19 with DIBAL
to the allylic alcohol 14, followed by Swern oxidization17
gives the aldehyde 15. At this point the second E-double
bond is introduced using the Schlosser modification20 of
the Wittig reaction.
Wittig reagent gives a mixture of triene isomers. The
major trans-isomer 18a can be partially purified by flash
chromatography, but it is still contaminated by the
2E,8E,10Z-triene 19a . This unwanted byproduct, orig-
inating from cis-olefin formation in the Schlosser reac-
tion, can be separated from the desired 2E,8E,10E-isomer
23
18a by MPLC with AgNO
3
impregnated silica gel.
Hydrolysis24 of the ester 18a with aqueous KOH in THF
followed by simultaneous treatment of the resulting
25
acid 20a with N-acetylcysteamine (NAC)26 and a mixture
of dicyclohexylcarbodiimide (DCC) and dimethylami-
nopyridine (DMAP) affords the NAC ester of the hexa-
ketide 5a . The methodology used to generate labeled
hexaketide 5a was accomplished initially with unlabeled
reagents. This allowed the construction of unlabeled
ethyl ester 18b, free acid 20b, and NAC triene 5b.
Compound 5a from the NAC coupling reaction con-
tained small amounts of an impurity which displays two
Use of Schlosser conditions with aldehyde 15 and 13C-
labeled phosphonium iodide21 produces the 2E,4E-diene
1
6 with typically 5-15% of the 2Z,4E-isomer. The E,E
geometry of 16 is evident from the characteristic 14.2 Hz
coupling constants seen between both H-2 and H-3, as
well as H-4 and H-5. This mixture is difficult to separate
and is therefore best carried through to the subsequent
steps. The removal of the acetal in saturated aqueous
oxalic acid and THF22 gives the highly volatile aldehyde
13
sets of doublets in the C NMR spectrum at 58.6 and
1
7. It is most effective to condense this directly with the
C-labeled Wittig reagent without purification. Initially,
33.6 ppm (J ) 31.2 Hz) and 56.5 and 34.2 ppm (J ) 33.2
Hz). The quantity of this material increased after normal
flash chromatography and the impurity still persisted in
samples purified by reverse phase HPLC. Although the
1
3
the full synthetic scheme involved a 1,2-ethylene glycol
acetal as the protecting group for the original aldehyde
8
1
, but acetal deprotection at the diene stage generates a
H NMR spectrum appears to indicate presence of only
high preponderance of decomposition products. The side
reactions were circumvented using the more labile
dimethoxy acetal. Condensation of 17 with C-labeled
a single product 5a , it is clear that small amounts of the
triene cyclized spontaneously at room temperature to
1
3
(23) (a) McInnes, A. G.; Walter, J . A.; Wright, J . L. C. Tetrahedron
Lett. 1979, 35, 3245-3248. (b) Rawlings, B. J .; Reese, P. B.; Ramer, S.
E.; Vederas, J . C. J . Am. Chem. Soc. 1989, 111, 3382-3390.
(24) Hungerb u¨ hler, E.; Seebach, D.; Wasmuth, D. Helv. Chim. Acta
1981, 64, 1467-1487.
(25) Parker, D. J . Chem. Soc., Perkin Trans. 2 1983, 83-88.
(26) (a) Gerstein, J .; J encks, W. P. J . Am. Chem. Soc. 1964, 86,
4655-4663. (b) Schwab, J . M.; Klassen, J . B. J . Am. Chem. Soc. 1984,
106, 7217-7227. (c) For some recent alternative approaches not
involving free acids see: Gilbert, I. H.; Ginty, M.; O’Neill, J . A.;
Simpson, T. J .; Staunton, J .; Willis, C. L. Bioorg. Med. Chem. Lett.
1995, 5, 1587-1590.
(
18) Isler, O.; Gutmann, H.; Montavon, M.; R u¨ egg, R.; Ryser, G.;
Zeller, P. Helv. Chim. Acta 1957, 40, 1242-1249.
19) Nicolaou, K. C.; Daines, R. A.; Uenishi, J .; Li, W. S.; Papahatjis,
D. P.; Chakraborty, T. K. J . Am. Chem. Soc. 1988, 110, 4672-4685.
20) Schlosser, M.; Christmann, K. F. Angew. Chem., Int. Ed. Engl.
966, 5, 126.
21) Barnhardt, R. G. J r.; McEwen, W. E. J . Am. Chem Soc. 1967,
9, 7009-7014.
22) Roush, W. R.; Hall, S. E. J . Am. Chem. Soc. 1981, 103, 5200-
211.
(
(
1
8
5
(
(

2
616 J . Org. Chem., Vol. 61, No. 8, 1996
Witter and Vederas
Sch em e 3a
a
Reagents: (a) LiBEt3H, 0 °C (85%); (b) Bu4N+ -HSO4, CS2, CH3I, NaOH, PhH (85%); (c) PDC (80%); (d) Bu3SnH, cymene, 150 °C
(63%); (e) HS(CH2)2SH, BF3‚OEt2 (66%); (f) neat Bu3SnH, AIBN, 120 °C, 4 days (99%).
form two bicyclic Diels-Alder reaction products. This
process is much more easily detectable with the 13C-
labeled material 5a than with unlabeled 5b.
methyl xanthate 25 via a two-phase system30 followed
by oxidation of the troublesome secondary alcohol to the
corresponding ketone 26. Deoxygenation31 of the methyl
xanthate 26 occurs at 150 °C to produce the keto-ester
2
2
unsuccessful. Activated Raney-nickel causes over-reduc-
tion whereas deactivated reagent leads to either complex
mixtures or isolation of starting material. However,
reaction of dithioketal 28 in neat tri-n-butyltin hydride
and catalytic AIBN at elevated temperatures33 produces
the desired reference compound 21 in good yield along
with a partially-reduced species 29 which could be
converted into desired reference compound 21 by further
reaction.
Bicyclic Refer en ce Com p ou n d 21. To unambigu-
ously confirm whether one of the spontaneous cyclization
products is the endo Diels-Alder adduct with stereo-
chemistry analogous to that in the lovastatin precursor,
7, which is subsequently protected32 as the dithioketal
8. Attempts to remove the ketal with Raney-nickel were
4
a,5-dihydromonacolin L (2), the corresponding decalin
derivative 21 was synthesized by an independent route
Scheme 3). The total synthesis of 2 represents a
(
2
7
challenge undertaken by many research groups, and our
route to reference compound 21 begins with the tricyclic
lactone 22 employed as an intermediate by Lewis and
co-workers.2 Compound 22 contains the required ster-
eochemistry and requires only cleavage of the lactone and
its oxygens from the A-ring to give 21. Reduction of 22
with lithium triethylborohydride generates the diol 23
and the triol 24. However, the seemingly straightforward
removal of the hydroxyl groups proved unexpectedly
difficult. Several unsuccessful routes were tried involv-
ing formation of the tosylate or halogenation and subse-
quent elimination, as well as Barton deoxygenation via
7e
In Vitr o In tr a m olecu la r Diels-Ald er Rea ction s.
The intramolecular Diels-Alder reactions of 1,7,9-dec-
atrienes forming bicyclo[4.4.0]decenes have been re-
viewed by several authors in the past 10 years3 and are
4
3
5
actively being used to make natural products. Com-
parison of the exo and endo transition states with the
connecting chain in its chairlike form can sometimes
allow prediction of the stereochemical outcome, with the
major adducts being those generated from the conformers
methyl xanthates2 and phenoxythiocarbonates.
8
29
The
hydroxyl group at C-8 is axial, and this orientation
presents a problem. Its juxtaposition with the axial
methylene group at C-6 facilitates the formation of a
tricyclic ether, and the antiperiplanar relationship to
vicinal hydrogens allows facile elimination to an olefin.
with the least bond-angle strain and fewest nonbonded
34b
interactions.
Consideration of transition states for
intramolecular Diels-Alder reaction of unlabeled triene
ethyl ester 18b, free acid 20b, and NAC thioester 5b
suggests that both endo and exo cyclizations are likely
and could each give two diastereomers (i.e. four isomers
total), depending on whether the methyl at C-6 is
The interference caused by the C-8 hydroxyl could be
circumvented first protecting the primary alcohol as a
(
30) Di Cesare, P.; Gross, B. Synthesis 1980, 714-715.
(27) (a) Hecker, S. J .; Heathcock, C. H. J . Am. Chem. Soc. 1986,
(31) Barton, D. H. R.; Motherwell, W. B; Stange, A. Synthesis 1981,
1
08, 4586-4594. (b) Narasaka, K.; Saitou, M.; Iwasawa, N. Tetrahe-
743-745.
dron: Asymmetry 1991, 2, 1305-1318. (c) Falck, J . R.; Yang, Y.-L.
Tetrahedron Lett. 1984, 25, 3563-3566. (d) Hanessian, S.; Roy, P. J .;
Petrini, M.; Hodges, P. J .; Di Fabio, R.; Carganico, G. J . Org. Chem.
(32) Yang, Y.-L.; Manna, S.; Falck, J . R. J . Am. Chem. Soc. 1984,
106, 3811-3814.
(33) Gutierrez, C. G. Stringham, R. A.; Nitasaka, T.; Glasscock, K.
G. J . Org. Chem. 1980, 45, 3393-3395.
1
990, 55, 5766-5777. (e) Blackwell, C. M.; Davidson, A. H.; Launch-
bury, S. B.; Lewis, C. N.; Morrice, E. M.; Reeve, M. M.; Roffey, J . A.
R.; Tipping, A. S.; Todd, R. S. J . Org. Chem. 1992, 57, 5596-5606. (f)
Schnaubelt, J .; Reissig, H.-U. Synlett. 1995, 452-454.
(34) (a) Ciganek, E. Org. React. 1984, 32, 1-374. (b) Fallis, A. G.
Can. J . Chem. 1984, 62, 183-234. (c) Craig, D. Chem. Soc. Rev. 1987,
16, 187-238. (d) Roush, W. R. Adv. Cycloaddit. 1990, 2, 91-146.
(35) For selected recent examples see citations in references 9i, 10,
27f as well as: Shing, T. K. M.; Yang, J . J . Org. Chem. 1995, 60, 5785-
5789.
(
(
28) Fuller, T. S.; Stick, R. V. Aust. J . Chem. 1980, 33, 2509-2515.
29) Robins, M. J .; Wilson, J . S.; Hansske, F. J . Am. Chem. Soc.
1
983, 105, 4059-4065.

Diels-Alder Cyclization during Biosynthesis of Lovastatin
Sch em e 4
J . Org. Chem., Vol. 61, No. 8, 1996 2617
pseudoaxial or pseudoequatorial (Scheme 4). To deter-
mine the outcome, the trienes were individually subjected
Sch em e 5
3
6
to both thermal and Lewis acid (EtAlCl
cyclization conditions.
2
)
-catalyzed
A solution of the triene ethyl ester 18b in toluene
heated to 160 °C for 4 days in a sealed tube generated a
1
:1 mixture of 30 and 31 (72%), which are separable by
flash chromatography, along with a small amount (6%)
of unreacted starting material 18b. The free acid triene
2
0b and NAC ester triene 5b also cyclize under the
thermal conditions and produce 1:1 mixtures of cycload-
ducts 32-33 (83%), and 34-35 (81%), respectively. Both
mixtures are difficult to separate. Hence, the cyclized
ethyl esters 30 and 31 were reduced37 to their corre-
sponding alcohols 36 (80%) and 37 (86%), respectively,
with lithium aluminum hydride (Scheme 5). The two
mixtures of cyclized products from the NAC ester 5b and
treated with 0.9 equiv of ethylaluminum dichloride
(
2
EtAlCl ) at room temperature. Each reaction was
complete in less than 3 h. Each ester generates the same
two cycloadducts as produced from their respective
thermal reactions, except the product mixtures are no
longer in a 1:1 ratio. The ethyl ester produces a 9:1
mixture of 30:31 (58%), and the NAC ester affords 34:35
free acid 20b reactions were also reduced to their alcohols
1
(
79% and 81%, respectively). Comparison of the H NMR
data of the alcohols 36 and 37, generated from each ethyl
ester reduction, with the mixture of alcohols from the
NAC ester and free acid reductions demonstrates that
each of the Diels-Alder reactions (of 18b, 20b, and 5b)
has the same stereochemical outcome and produces two
analogous products. To further confirm this, the purified
(80%) in a ratio of 19:1. Although the same two products
result from the thermal reaction, the Lewis acid cycliza-
tions proceed rapidly at room temperature and show
significant endo selectivity.
Since incorporation experiments were done with cells
in aqueous media (see below), which are known to
accelerate Diels-Alder reactions,38 the influence of sol-
vent on cyclization of 18b was also examined. The half-
life for cyclization of 18b to 30 and 31 in chloroform at
alcohol 36 derived from the higher R
oxidized to the acid 32 (51%).
f
ethyl ester 30 was
To examine the effects of a Lewis acid on the cycliza-
tion, both the ethyl ester 18b and NAC ester 5b were
2
(
2 °C is about 10 days. In water:acetonitrile:methanol
ca 5:5:1) at 28 °C (fermentation temperature for A.
terreus) the half-life for reaction drops to about 2 days
(
(
36) Roush, W. R.; Gillis, H. R. J . Org. Chem. 1982, 47, 4825-4829.
37) VanMiddlesworth, F. L. J . Org. Chem. 1986, 51, 5019-5021.

2
618 J . Org. Chem., Vol. 61, No. 8, 1996
Witter and Vederas
F igu r e 1. Possible conformations adopted by cycloadducts 21, 30, 31, and B. Relevant NMR data is indicated.
at either pH 5 or 7. Nearly identical accelerations are
seen in cell-free fermentation media used for incorpora-
tion experiments.
NMR analysis of the cis-fused exo adduct 31 is hin-
dered by the complexity of the angular hydrogen signals
which obscures the magnitude of the coupling constant
1
Chromatographic analysis of Diels-Alder reactions
and spectral comparison with 30 and 31 clearly demon-
strate that compound 21 is not one of the products of
thermal or Lewis acid cyclization of ethyl ester 18b. The
full stereochemical assignments of the cycloadducts
between H-4a and H-8a. However, difference H NOE
spectra in which the overlapping C-6 and C-2 methyl
protons are irradiated show an enhancement of the H-4a
but not the H-8a signal, which clearly eliminates the cis-
fused product B as a possible structure, since no confor-
mation of this isomer would allow an NOE enhancement
between the H-4a and H-8a and the bicyclic methyl
substituents. This enhancement also indicates the cis-
fused conformer 31 is preferred over 31′.
In cor p or a tion Exp er im en ts a n d Con clu sion s. In
order to test whether the labeled hexaketide derivative
a could be incorporated intact into lovastatin (1), it was
added under a variety of conditions to growing cultures
1
13
1
required combined use of H, C, COSY, HMQC, and H-
decoupled NMR experiments.39 The H COSY spectrum
1
f
of the higher R diastereomer 30 allows complete assign-
1
ment of the H signals. Irradiation experiments show a
characteristic trans-fused coupling constant of 12.0 Hz
between H-8a and H-4a, thus eliminating the two po-
tential cis-fused adducts as possible structures (Figure
5
1
). The distinction between the trans-fused adducts, 21
and 30, is also clear from the coupling pattern seen for
H-5ax. In the two most probable conformations for the
trans-fused products, only the conformer shown in Figure
of Aspergillus terreus MF 4845 both in the presence and
8d,f
absence of various ω- and â-oxidation inhibitors.
Use
40
41
of protoplasts, saponin, and 2,6-O-dimethyl-â-cyclo-
1
allows for an axial-axial arrangement between H-5ax
8g
dextrin to assist the incorporation was also attempted.
and H-4a, and H-5ax and H-6. The H-5ax signal is an
apparent quartet arising from three doublets each having
a coupling constant of 12.0 Hz. Two of the three doublets
result from two axial-axial vicinal couplings and the
third is from geminal coupling with H-5eq. Spectral
analysis of 21 exhibits two large coupling constants for
H-4a which suggests a 180° relationship with H-5ax and
H-8a as shown. The peak for its H-5ax proton displays
only two large coupling constants corresponding to the
axial-axial relationship of H-5ax and H-4a and the
geminal coupling with H-5eq. The third doublet in this
signal shows only a 4.8 Hz coupling constant between
H-5ax and H-6. The results define the preferred confor-
mations of 21 and 30.
None of these experiments gave any detectable carbon-
13
carbon coupling in the C NMR spectrum of the resulting
1
. In addition, incoporations with intact cells typically
allowed less than 1% recovery of the labeled precursor
a . Attempts to incorporate a number of shorter NAC
5
esters which would be expected to be on the pathway,
13
including doubly C-labeled diketides such as NAC
acetoacetate, NAC â-hydroxybutyrate, and NAC
42
2
-butenoate, gave similar results.
Apparently very
rapid degradation of the precursors in A. terreus prevents
their loading into the lovastatin PKS system. Fortu-
nately, the availability of both nonenzymatic Diels-Alder
products 34 and 35 from cyclization of 18 as well as the
putative enzymatic product, the NAC ester corresponding
(
38) For discussion of media effects on Diels-Alder reactions see:
a) Blake, J . F.; Lim, D.; J orgensen, W. L. J . Org. Chem. 1994, 59,
03-805 and references therein. (b) Pai, C. K.; Smith, M. B. J . Org.
(
8
(40) Ventura, L.; Ram o´ n, D. FEMS Microbiol. Lett. 1991, 82, 189-
194.
(41) Meiners, S.; Gharyal, P. K.; Schlinder, M. Planta 1991, 184,
443-447.
(42) Attempts to incorporate the stereochemically correct triketide,
NAC (E,E)-2,4-hexadienoate, and the tetraketide, NAC (2S)-(E,E)-2-
methyl-4,6-octadienoate, into 1 were also unsuccessful: Yoshizawa,
Y.; Vederas, J . C. unpublished results.
Chem. 1995, 60, 3731-3735. (c) Otto, S.; Engberts, J . B. F. N.
Tetrahedron Lett. 1995, 36, 2645-2648. (d) Faita, G.; Righetti, P.
Tetrahedron 1995, 51, 9091-9102. (e) Breslow, R.; Zhu, Z. J . Am.
Chem. Soc. 1995, 117, 9923-9924.
(
39) NMR of Macromolecules, a Practical Approach; Roberts, G. C.
K., Ed.; Oxford University Press: Oxford, 1993.

Diels-Alder Cyclization during Biosynthesis of Lovastatin
to diastereomer 21, affords a convenient assay for the
J . Org. Chem., Vol. 61, No. 8, 1996 2619
solution was stirred overnight at room temperature. The
mixture was concentrated in vacuo to afford a yellow oil, which
“Diels-Alderase” (most likely a lovastatin PKS) in cell
was purified by flash chromatography (SiO
00% EtOAc, R 0.33) to yield 5b (970 mg, 76%) as a white
solid: mp 38-39 °C; [R]
Cl cast) 3321 (br s), 1657 (s), 1626 (s) cm ; H NMR (400
MHz, CDCl ) δ 6.90 (dt, 1Η, J ) 15.5, 7.4 Ηz), 6.11 (dt, 1H, J
2
; 40 × 120 mm,
free experiments that are currently being investigated.
In summary, the synthesis of doubly 1 C-labeled hexa-
ketide NAC thioester 5a in optically pure form can be
achieved in 13 steps from commercially available (R)-
citronellol with an overall yield of 4.0%, with the labels
added at a late stage. The nonenzymatic Diels-Alder
reactions of unlabeled triene NAC thioester 5b, its ethyl
ester 18b, and its free acid 20b yield two analogous
diastereomers in each case (derived from exo and endo
transition states), under either thermal or Lewis acid
conditions. As expected,9 reaction is most facile for the
thioester 18b in aqueous media. The synthesis of refer-
ence compound 21, which has stereochemistry analogous
to lovastatin precursor 2, confirms the configurational
assignment and demonstrates that 21 is not produced
in the nonenzymatic cyclization. Thus, the conventional
exo and endo Diels-Alder reactions both proceed through
chairlike transition states that place the side chain
methyl pseudoequatorial, whereas the proposed enzy-
matic Diels-Alder enroute to lovastatin (1) requires an
intermediate endo conformation with this methyl group
pseudoaxial. One function of the PKS enzyme may be
to correctly control the conformation of the acyclic triene
thioester, which is naturally prone to cyclization, through
hydrophobic interactions. It may also assist the ring
formation through Lewis acid catalysis, either by simple
1
f
3
2
0
D
-7.41° (c 0.081, CH Cl ); IR (CH -
2
1
2
2
-1
2
3
) 15.5, 1.5 Hz), 6.01 (ddq, 1H, J ) 14.2, 10.2, 1.5 Hz), 5.95
(ddt, 1H, J ) 14.2, 10.2, 1.2 Hz), 5.57 (dq, 1H, J ) 14.2, 6.6
Hz), 5.91-5.83 (br s, 1H), 5.48 (dt, 1H, J ) 14.2, 7.2 Hz), 3.44
(
(
7
(
dt, 2H, J ) 6.3, 5.9 Hz), 3.07 (t, 2H, J ) 6.3 Hz), 2.31-2.12
m, 2H), 2.06 (ddd, J ) 13.7, 7.2, 6.1 Hz), 1.94 (ddd, J ) 13.7,
.4, 7.2 Hz), 1.95 (s, 3H), 1.72 (d, 3H, J ) 6.6 Hz), 1.55-1.45
13
m, 2H), 1.30-1.25 (m, 1H), 0.87 (d, 3H, J ) 6.6 Hz); C NMR
i,38
(
100 MHz, CDCl ) δ 190.36, 170.34, 146.77, 131.97, 131.55,
3
129.66, 128.27, 127.20, 39.86, 39.84, 34.55, 32.87, 29.92, 28.26,
+
+
23.20, 19.32, 17.99; MS (CI, NH
3
) 327 (MNH
4
, 8), 310 (MH ,
14). Anal. Calcd for C17
C, 66.24; H, 9.03.
H26NO S: C, 65.98; H, 8.79. Found:
2
1
3
[
2
2,11- C ]-La beled 5 (5a ). The same method as for the
preparation of 5b was employed. The spectral and chromato-
graphic properties were similar to those of 5b except for the
1
following: H NMR (200 MHz, CDCl
3
) δ 6.92 (dtd, 1H, J )
1
5.6, 7.0, 1.0 Hz), 6.12 (ddt, 1H, J ) 161.0, 15.6, 1.5 Hz), 6.05-
5.85 (m, 2H), 5.57 (ddq, 1H, J ) 150.2, 14.2, 6.7 Hz), 1.73 (dd,
3H, J ) 6.7, 6.7 Hz); 13C NMR (50 MHz, CDCl
) δ 190.30 (d,
3
J ) 76.0 Hz), 146.72 (d, J ) 70.0 Hz), 131.66 (d, J ) 72.5 Hz),
3
3
3
4.50 (d, J ) 3.5 Hz), 17.96 (d, J ) 44.3 Hz); MS (CI, NH )
+
+
29 (MNH
4
,
25), 312 (MH , 100). Anal. Calcd for
12C15
C H26NO S: C, 66.20; H, 8.74. Found: C, 66.00; H, 9.08.
2 2
1
3
(
3R)-Meth yl-6-oxoh exyl Aceta te (8). Ozonized oxygen,
cooled by passage through a glass coil immersed in a cold-
4
3
site specific protonation or coordination of a metal ion.
bath at -78 °C, was bubbled into a cold (-78 °C) solution of
It seems likely that the cyclization occurs spontaneously
upon formation of the appropriate skeleton A in the
polyketide synthase active site and may be enhanced by
14
7
2 2
(6.17 g, 31.1 mmol) in dry CH Cl (60 mL) until the solution
1
5
had acquired a blue tint (1.5 h). Triphenylphosphine (8.98
g, 34.2 mmol) was added, and the reaction mixture was stirred
at -78 °C for a further 30 min. The cooling bath was then
removed and stirring was continued for 2 h. The solution was
evaporated in vacuo, and the residue was purified by flash
carbonyl protonation expected to assist normal acyl
transfer reaction required by PKS enzymes.6
,7
This,
together with the observation that enzymatic oxidation
of 3 spontaneously incites formation of chiral solanapy-
rone (4),10 suggests that other proteins normally not
involved in cyclization reactions may become “Diels-
Alderases” by formation of the appropriate precursor on
chromatography (SiO
(4.56 g, 85%) as a colorless volatile oil: [R]
3 3
CHCl ); IR (CHCl cast) 1740 (s), 1725 (s) cm ; H NMR (400
2
; 30% Et
2
O in pentane, R
f
0.25) to yield
20
8
D
-1
+1.87° (c 7.18,
1
MHz, CDCl ) δ 9.74 (br s, 1H), 4.12-4.01 (m, 2H), 2.50-2.35
3
(m, 2H), 2.00 (s, 3H), 1.71-1.57 (m, 2H), 1.60-1.50 (m, 1H),
1
3
an enzyme surface.4
4,45
1.49-1.40 (m, 2H), 0.89 (d, 3H, J ) 6.4 Hz); C NMR (100
Experiments to test this hypoth-
MHz, CDCl
2
3
) δ 202.34, 171.09, 62.53, 41.44, 35.19, 29.43, 28.69,
esis and to confirm the involvement of hexaketide A in
formation of 2 are in progress.
+
+
0.59, 19.08; MS (CI, NH
Anal. Calcd for C
H, 9.59.
3
) 190 (MNH
9 16 3
H O : C, 62.77; H, 9.36. Found: C, 62.72;
4
, 100), 173 (MH , 4.2).
Exp er im en ta l Section
(
3R)-6,6-Dim eth oxy-3-m eth ylh exyl Aceta te (9). A solu-
tion of aldehyde 8 (24.3 g, 141 mmol), trimethyl orthoformate
100 mL), and acetyl chloride (0.26 mL, 3.62 mmol) in dry
Gen er a l. General procedures and instrumentation can be
obtained from the supporting information. Published proce-
dures, which have been modified and used, are cited.
(
MeOH (100 mL) was heated to reflux overnight using a Dean-
Stark apparatus.16 After the solution was allowed to cool, and
(
6R)-E,E,E-6-Meth yld od eca -2,8,10-tr ien oic Acid N-Ace-
tylcystea m in e Th ioester (5b). A solution of the acid 20b
860 mg, 4.13 mmol) in dry CH Cl (7 mL) was treated
simultaneously with a solution of N-acetylcysteamine (581 mg,
brine (75 mL) and 5% aqueous NaHCO
Most of the MeOH was removed in vacuo, and the mixture
was extracted with Et O layer was
O (2 × 350 mL). The Et
washed with H O (50 mL) and brine (50 mL), and these
aqueous washings were combined and back-extracted with
Et O fractions were dried
O (3 × 250 mL). The combined Et
MgSO ) and concentrated in vacuo to a clear colorless oil (31.8
3
(75 mL) were added.
(
2
2
2
2
2
4
.87 mmol) in CH
2 2
Cl (7 mL), and a solution of DCC (1.02 g,
4
Cl
.95 mmol) and 4-(dimethylamino)pyridine (19.6 mg) in CH -
2
2
5
2
2
2
(7 mL) over 5 min at -15 °C. The resultant cloudy white
43) For selected references to Lewis acid catalysis of Diels-Alder
(
4
g), which can be used without further purification. A small
portion (0.638 g) was removed and purified by Kugelrohr
distillation to yield a clear oil 9 (0.580 g, 91%): bp 109-111
(
reactions see: (a) Kagan, H. B.; Riant, O. Chem. Rev. 1992, 92, 1007-
1
1
020. (b) Haase, C.; Sarko, C. R.; DiMare, M. J . Org. Chem. 1995, 60,
777-1787. (c) Seebach, D.; Dahinden, R.; Marti, R. E.; Beck, A. K.;
2
0
°
1
C (0.2 mmHg); [R]
D
+2.23° (c 2.38, CHCl
3
); IR (CHCl
) δ 4.32 (t, 1H, J )
3
cast)
-
1
1
741 (s) cm ; H NMR (400 MHz, CDCl
3
Plattner, D. A.; K u¨ hnle, N. M. J . Org. Chem. 1995, 60, 1788-1799.
(d) Evans, D. A.; Murry, J . A.; von Matt, P.; Norcross, R. D.; Miller, S.
4.7 Hz), 4.13-4.04 (m , 2H), 3.30 (s, 6H), 2.03 (s, 3H), 1.70-
J . Angew. Chem., Int. Ed. Engl. 1995, 34, 798-800. (e) J ohannsen,
M.; J ørgensen, K. A. J . Org. Chem. 1995, 60, 5757-5762. (f) Yamabe,
S.; Dai, T.; Minato, T. J . Am. Chem. Soc. 1995, 117, 10994-10997.
1.50 (m, 4H), 1.49-1.32 (m, 2H), 1.24-1.14 (m, 1H), 0.90 (d,
13
3
6
H, J ) 6.5 Hz); C NMR (100 MHz, CDCl ) δ 171.18, 104.86,
3
2.91, 52.69, 35.47, 31.58, 30.01, 29.81, 21.02, 19.41; MS (CI,
(44) (a) Hilvert, D.; Hill, K. W.; Nared, K. D.; Auditor, M.-T. M. J .
+
3 4 22 4
) 236 (MNH , 6). Anal. Calcd for C11H O : C, 60.51;
NH
H, 10.16. Found: C, 60.20; H, 10.40.
3R)-6,6-Dim eth oxy-3-m eth ylh exa n ol (10). Freshly pre-
Am. Chem. Soc. 1989, 111, 9261-9262. (b) For a review of catalytic
antibodies see: Schultz, P. A.; Lerner, R. A. Science 1995, 269, 1835-
1
842.
(
(45) For selected references to Diels-Alder studies with nonprotein
pared sodium methoxide (0.47 g, 8.7 mmol) was added to a
solution of acetate 9 (20.0 g, 91.5 mmol) in dry MeOH (200
mL). After stirring at room temperature overnight, the
hosts see: (a) Morris, K. N.; Tarasow, T. M.; J ulin, C. M.; Simons, S.
L.; Hilvert, D.; Gold, L. Proc. Natl. Acad. Sci. U.S.A. 1994, 91, 13028-
1
3032. (b) Walter, C. J .; Sanders, J . K. M. Angew. Chem., Int. Ed. Engl.
995, 34, 217-219.
1
2
mixture was treated with H O (120 mL), and the solvent

2
620 J . Org. Chem., Vol. 61, No. 8, 1996
Witter and Vederas
volume was reduced in vacuo to give an aqueous residue, which
was extracted with Et
O (3 × 330 mL). The combined Et
fractions were washed with H
O (2 × 100 mL) and brine (200
mL), and these aqueous washings were back-extracted with
Et O fractions were again
O (3 × 50 mL). The combined Et
washed with H
O (2 × 50 mL) and brine (50 mL), dried
MgSO ), and concentrated in vacuo to yield a clear colorless
oil (14.4 g), which could be used without further purification.
A small portion (0.948 g) was removed and purified by
Kugelrohr distillation to yield a clear oil 10 (0.844 g, 89%):
50 × 250 mm, 75% Et
g, 80%) as a clear oil: [R]
Cl cast) 3420 (br m), 2952 (s) cm
benzene-d ) δ 5.55-5.49 (m, 2H), 4.27 (t, 1H, J ) 5.6 Hz), 3.91
2
O in pentane, R
f
0.31) to yield 14 (0.85
2
0
2
2
O
D
+3.42° (c 2.93, CHCl
3
); IR (CH
2
-
-
1
1
2
2
; H NMR (400 MHz,
6
2
2
(br s, 2H), 3.14 (s, 6H), 1.99-1.90 (m, 1H), 1.84-1.75 (m, 1H),
2
1.70-1.50 (m, 3H) 1.45-1.33 (m, 2H), 1.22-1.12 (m, 1H), 0.82
(
4
(d, 3H, J ) 6.5 Hz); 13C NMR (100 MHz, benzene-d
) δ 131.57,
6
130.19, 104.88, 63.41, 52.24, 39.91, 33.12, 31.53, 30.36, 19.59;
+
+
MS (CI, NH ) 220 (MNH
22 3
for C11H O : C, 65.30; H, 10.97. Found: C, 65.40; H, 11.07.
4
4
, 6), 203 (MH , 0.3). Anal. Calcd
2
0
bp 125 °C (0.1 mmHg); [R]
D
+2.75° (c 2.22, CHCl
cast) 3409 cm ; H NMR (400 MHz, benzene-d ) δ 4.26 (t, 1H,
J ) 5.7 Hz), 3.57-3.40 (m, 2H), 3.13 (s, 6H), 1.70-1.49 (m,
3
); IR (CHCl
3
(5R)-(E)-8,8-Dim eth oxy-5-m eth yloct-2-en a l (15). The
same method as for the preparation of aldehyde 11 was
employed. Thus, oxidation of alcohol 14 (4.66 g, 23.1 mmol)
with DMSO (3.11 mL, 43.8 mmol) and oxalyl chloride (2.41
mL, 27.7 mmol) afforded 15 (4.04 g, 94%, based on 7%
-1 1
6
4
6
5
1
H), 1.49-1.32 (m, 1H), 1.32-1.12 (m, 2H), 0.83 (d, 3H, J )
.6 Hz); C NMR (100 MHz, benzene-d
2.25, 52.10, 40.10, 32.08, 30.24, 29.60, 19.73; MS (CI, NH
13
6
) δ 104.91, 60.61,
3
)
recovered starting material) as a clear oil after flash chroma-
+
+
20
94 (MNH
: C, 61.31; H, 11.44. Found: C, 60.94; H, 11.29.
3R)-6,6-Dim eth oxy-3-m eth ylh exa n a l (11). Dry DMSO
2.11 mL, 29.8 mmol) was added dropwise over 5 min to a
stirred cooled (-78 °C) solution of distilled oxalyl chloride (1.36
4
, 2.7), 177 (MH , 0.1), 130 (100). Anal. Calcd for
tography (SiO ; 50% Et O in pentane, R 0.35): [R]
-0.20°
2
2
f
D
C
9
H
20
O
3
3 2 2
(c 2.50, CHCl ); IR (CH Cl cast) 2725 (w), 1693 (s), 1636 (w)
cm ; H NMR (400 MHz, benzene-d ) δ 9.30 (d, 1H, J ) 7.7
6
-
1
1
(
(
Hz), 6.09 (dt, 1H, J ) 15.5, 7.7 Hz), 5.90 (ddt, 1H, J ) 15.5,
7.7, 1.0 Hz), 4.21 (t, 1H, J ) 5.5 Hz), 3.14 (s, 6H), 1.76 (ddd,
1H, J ) 13.4, 7.7, 5.8 Hz), 1.59 (ddd, 1H, J ) 13.4, 7.7, 7.4
Hz), 1.56-1.40 (m, 2H), 1.28-1.15 (m, 2 H), 1.10-0.99 (m, 1H),
1
7
mL, 15.6 mmol) in CH
2
Cl
2
(30 mL). After 10 min, a solution
Cl (15 mL) was
of the alcohol 10 (2.72 g, 15.6 mmol) in CH
2
2
0.61 (d, 3H, J ) 6.3 Hz); 13C NMR (100 MHz, benzene-d
) δ
added over 30 min. The resultant slurry was stirred for 20
min at -78 °C, and then dry triethylamine (7.43 mL, 53.3
mmol) was injected dropwise over 30 min. Stirring was
continued at -78 °C for 20 min and the cold bath removed,
6
192.57, 155.71, 134.51, 104.65, 52.35, 39.77, 32.37, 31.53,
+
+
30.76, 19.33; MS (CI, NH
3
) 218 (MNH
4
, 25), 201 (MH , 0.1),
75 (100). Anal. Calcd for C11
Found: C, 65.81; H, 10.09.
H
20
O
3
:
C, 65.95; H, 10.07.
and after a further 30 min, H
mixture was stirred for a further 10 min, and the aqueous
layer was extracted with CH Cl
(2 × 10 mL). The combined
organic layers were washed with 10% v/v aqueous HCl (2 × 7
mL), saturated aqueous NaHCO
(2 × 10 mL), and brine (1 ×
0 mL), dried over MgSO , and evaporated in vacuo. The
residue was purified by flash chromatograghy (SiO ; 25% Et
in pentane, R 0.25) to yield 11 (2.31 g, 85%) as an oil: [R]
13.23° (c 1.41, CHCl
NMR (300 MHz, CDCl
J ) 5.6 Hz), 3.27 (s, 6H), 2.38 (ddd, 1H, J ) 16.0, 5.8, 2.2 Hz),
.21 (ddd, 1H, J ) 16.0, 7.8, 2.2 Hz), 2.04 (br p, 1H, J ) 6.6
2
O (10 mL) was added. The
(7R)-(E,E-10,10-Dim eth oxy-7-m eth yldeca-2,4-dien e (16b).
Ethyltriphenylphosphonium iodide (0.295 g, 0.704 mmol) was
2
2
suspended in dry THF (1.4 mL) and dry Et O (1.1 mL) and
2
3
was stirred with phenyllithium (1.8 M in cyclohexane/Et O (70/
2
2
0
1
4
30); 0.39 mL, 0.704 mmol) for 10 min at room temperature.
2
2
O
After the white slurry turned a clear red color, the solution
2
0
f
D
was cooled to -78 °C and a solution of the aldehyde 15 (0.140
+
3
); IR (neat) 2720 (m), 1725 (s) cm ; 1H
-1
g, 0.704 mmol) in Et O (0.6 mL) was added with vigorous
2
3
) δ 9.71 (t, 1H, J ) 2.2 Hz), 4.30 (t, 1H,
stirring. After 5 min, an additional portion of phenyllithium
(1.8 M in cyclohexane/Et O (70/30); 0.39 mL, 0.704 mmol) was
2
2
added to the orange slurry. The red solution was stirred for
5 min, and ethereal HCl (5.3 M, 0.147 mL, 0.775 mmol) was
slowly added followed by a quick addition of potassium tert-
butoxide (1:1 complex with tert-butyl alcohol, 0.197 g, 1.06
mmol). The mixture was warmed to room temperature, stirred
for 2 h, and then diluted with Et O (50 mL), washed with H O
Hz), 1.71-1.48 (m, 2H), 1.41-1.12 (m, 2H), 0.93 (d, 3H, J )
1
3
6
5
.7 Hz); C NMR (75 MHz, CDCl
3
) δ 202.54, 104.55, 52.78,
) 192
2.74, 50.92, 31.52, 29.99, 27.94, 19.80; MS (CI, NH
3
+
+
(MNH
4
, 8), 175 (MH , 0.4), 75 (100). Anal. Calcd for
C
9 18 3
H O : C, 62.04; H, 10.41. Found: C, 61.89; H, 10.14.
2
2
Eth yl (5R)-(E)-8,8-Dim eth oxy-5-m eth yloct-2-en oate (12).
(4 × 10 mL) until neutral and then brine (1 × 10 mL), and
dried (MgSO ). Concentration of the filtrate in vacuo gave a
yellow liquid, which was purified by flash chromatography
(SiO ; 20% Et O in pentane, R 0.50) to afford 16b (111 mg,
A solution of aldehyde 11 (31.2 g, 179 mmol) and (carbeth-
oxymethylene)triphenylphosphorane (77.1 g, 221 mmol) in dry
benzene (1500 mL) was heated to reflux at 85 °C for 12 h.24
The mixture was cooled, the solvent was removed in vacuo,
and the residue was purified by repeated flash chromatography
4
2
2
f
75%) as a clear oil. This material was contaminated with
2Z,4E-isomer (8% by 1H NMR integration), which was not
2
0
(
(
(
SiO
29.9 g, 68%, R
5.29 g, 10%, R
2
; 15% Et
2
O in pentane) to yield the desired E-isomer 12
0.14) together with the minor, Z-isomer 13
0.19). A small portion of the E-isomer 12 was
easily removed. Data for the mixture: [R]
CHCl ); IR (CH Cl cast) 3016 (m), 2952 (s) cm ; MS (CI, NH
24
230 (MNH , 1.0), 213 (MH , 0.1). Anal. Calcd for C13H O
D
-3.22° (c 2.39,
-1
3
2
2
3
)
:
f
+
+
f
4
2
removed and purified further by Kugelrohr distillation to give
C, 73.52; H, 11.40. Found: C, 73.76; H, 11.46. Data for the
2
0
1
a clear oil: bp 148-151 °C (0.75 mm Hg); [R]
D
+2.98° (c 1.98,
6
desired E,E-diene: H NMR (400 MHz, benzene-d ) δ 6.12-
-
1 1
CHCl
3
); IR (CH
2
Cl
) δ 7.03 (dt, 1H, J ) 15.5, 7.7 Hz), 5.87 (dt,
H, J ) 15.5, 1.2 Hz), 4.19 (t, 1H, J ) 5.7 Hz), 4.04 (q, 2H, J
2
cast) 1722 (s), 1654 (m) cm ; H NMR (400
5.99 (m, 2H), 5.53-5.43 (m, 2H), 4.28 (t, 1H, J ) 5.7 Hz), 3.15
(s, 6H), 2.04 (ddd, 1H, J ) 13.8, 7.0, 7.0 Hz), 1.87 (ddd, 1H, J
) 13.8, 7.0, 6.5 Hz), 1.72-1.52 (m, 2H), 1.62 (d, 3H, J ) 6.7
Hz), 1.48-1.38 (m, 2H), 1.27-1.18 (m, 1H), 0.83 (d, 3H, J )
MHz, benzene-d
6
1
)
7.1 Hz), 3.12 (s, 6H), 1.85 (dddd, 1H, J ) 13.0, 7.7, 5.3, 1.2
6.6 Hz); 13C NMR (100 MHz, benzene-d
) δ 132.46, 132.40,
Hz), 1.67 (dddd, 1H, J ) 13.0, 7.7, 7.7, 1.2 Hz), 1.60-1.50 (m,
6
1
0
(
5
2
6
H), 1.50-1.40 (m, 1H), 1.32-1.20 (m, 2H), 1.11-1.02 (m, 1H),
130.34, 126.68, 104.88, 52.07, 40.38, 33.55, 31.73, 30.48, 19.65,
18.08.
.98 (t, 3H, J ) 7.1 Hz), 0.67 (d, 3H, J ) 6.6 Hz); 13C NMR
1
3
100 MHz, benzene-d
6
) δ 166.05, 147.70, 123.11, 104.66, 59.97,
[2- C]-La beled 16 (16a ). The same method as for the
preparation of 16b was employed. The spectral and chro-
matographic properties were similar to those of 16b except
3
2.21, 39.57, 32.49, 31.64, 30.26, 19.39, 14.30; MS (CI, NH )
+
4 24 4
, 27), 198 (100). Anal. Calcd for C13H O : C,
62 (MNH
3.89; H, 9.91. Found: C, 63.92; H, 9.92.
5R)-(E)-8,8-Dim eth oxy-5-m eth yloct-2-en ol (14). A solu-
tion of ester 12 (1.27 g, 5.24 mmol) in CH Cl (13 mL) was
treated with DIBAL (2.23 g, 15.7 mmol) in CH Cl (9 mL) over
0 min at -78 °C. The reaction mixture was stirred for 2 h
at -78 °C and then for 30 min at -30 °C, at which point MeOH
2 mL) was added to quench the excess of DIBAL. The mixture
was diluted with Et O (300 mL), and the layers were sepa-
rated. The Et O phase was washed with saturated aqueous
potassium-sodium tartrate (4 × 100 mL) and brine (2 × 100
mL), dried (MgSO ), and concentrated in vacuo to give a clear
oil. The residue was purified by flash chromatography (SiO
1
for: Data for the E,E-isomer: H NMR (400 MHz, benzene-
(
d ) δ 5.49 (m, 1H, H-5), 5.52-5.46 (ddd, 1H, J ) 150.5, 14.1,
6
13
6.7 Hz), 1.62 (dd, 3H, J ) 6.7, 6.7 Hz); C NMR (100 MHz,
benzene-d
18.03 (d, J ) 43.3 Hz).
2
2
6
) δ 132.42 (d, J ) 71.4 Hz), 130.30 (d, J ) 10.0 Hz),
2
2
1
9
1
(4R)-E,E-4-Meth yld eca -6,8-d ien a l (17b). Distilled THF
(21.5 mL) and saturated aqueous oxalic acid (15 mL) were
added to the protected aldehyde 16b (1.07 g, 5.02 mmol), and
the resultant mixture was stirred at room temperature for 24
h, then treated with a further portion of saturated aqueous
(
2
2
2
2
4
oxalic acid (2 mL), and stirred for an additional 6 h. The
reaction mixture was partitioned between Et O (250 mL) and
2
;
2

Diels-Alder Cyclization during Biosynthesis of Lovastatin
J . Org. Chem., Vol. 61, No. 8, 1996 2621
organic fractions were dried (MgSO ) and concentrated to yield
a clear oil (855 mg, 97%), which could be used without further
purification. The acid could be purified by flash chromatog-
H
(
5
2
O (50 mL), and the aqueous layer was extracted with Et
3 × 40 mL). The combined organic layers were washed with
% NaHCO (50 mL), H O (50 mL), and brine (50 mL) and
then dried (MgSO ). Partial concentration in vacuo gave a
2
O
4
3
2
4
raphy (SiO
(656 mg, 75%) as a clear oil: [R]
IR (CHCl cast) 3400-2300 (br), 1696 (s), 1650 (s) cm ; H
NMR (400 MHz, CDCl ) δ 11.2 (br s, 1H), 7.08 (dt, 1Η, J )
2
; 20 × 250 mm, 100% Et
2
O, R
f
0.50) to yield 20b
2
0
volatile pale yellow liquid (0.794 g), which could be used
without further purification. A portion of the crude aldehyde
D
-7.37° (c 0.095, CHCl
3
1
);
-
1
3
(
0.200 g) was chromatographed on silica (30% CH
pentane, R 0.25) to afford 17b (0.156 g, 78%), which still
contained the difficult to separate 2Z,4E-isomer (8% by
2
Cl
2
in
3
f
15.6, 6.9 Hz), 6.02 (ddq, 1H, J ) 14.2, 10.4, 1.5 Hz), 5.98 (ddt,
1H, J ) 14.4, 10.4, 1.3 Hz), 5.82 (dt, 1H, J ) 15.6, 1.6 Hz),
5.58 (dq, 1H, J ) 14.2, 6.9 Hz), 5.50 (dt, 1H, J ) 14.4, 7.3 Hz),
2.34-2.14 (m, 2H), 2.05 (ddd, 1H, J ) 13.8, 7.3, 6.4 Hz), 1.93
(ddd, 1H, J ) 13.8, 7.3, 7.0 Hz), 1.73 (d, 3H, J ) 6.9 Hz), 1.57-
1
H
2
0
NMR integration). Data for the mixture: [R]
CH Cl ); IR (CH Cl cast) 3015 (m), 2725 (m), 1723 (s) cm
MS (EI) calcd for C11
09.0650 (24). Anal. Calcd for C11
Found: C, 79.25; H, 10.88. Data for the desired E,E-dienal:
D
-8.27° (c 1.28,
-
1
2
2
2
2
;
+
H
18O 166.1358, found 166.1357 (M , 21),
18O: C, 79.45; H, 10.92.
1
3
1
H
1.45 (m, 2H), 1.32-1.22 (m, 1H), 0.88 (d, 3H, J ) 6.6 Hz);
NMR (100 MHz, CDCl ) δ 172.28, 152.49, 131.96, 131.59,
129.76, 127.16, 120.63, 39.90, 34.50, 32.82, 29.98, 19.30, 18.03;
C
3
1
H NMR (400 MHz, benzene-d ) δ 9.30 (t, 3H, J ) 1.7 Hz),
6
+
+
6
1
.10-5.95 (m, 2H), 5.49 (dq, 1H, J ) 14.2, 7.1 Hz), 5.38 (dt,
H, J ) 14.1, 7.0 Hz), 1.95-1.67 (m, 4H), 1.58 (d, 3H, J ) 7.0
MS (CI, NH
20 2
for C13H O : C, 74.96; H, 9.68. Found: C, 74.61; H, 9.73.
3
) 226 (MNH , 81), 209 (MH , 6). Anal. Calcd
4
Hz), 1.44-1.35 (m, 1H), 1.25-1.15 (m, 1H), 1.15-1.07 (m, 1H),
[2,11-13C ]-La beled 20 (20a ). The same method as for the
2
0
2
2
.67 (d, 3H, J ) 6.7 Hz); 13C NMR (100 MHz, benzene-d
6
) δ
preparation of 20b was employed. The spectral and chro-
matographic properties were similar to those of 20b except
00.62, 132.55, 132.25, 129.76, 126.98, 41.58, 40.31, 32.97,
8.58, 19.19, 18.04.
1
for the following: H NMR (300 MHz, CDCl ) δ 7.07 (dtd, 1Η,
3
1
3
[
9- C]-La beled 17 (17a ). The same method as for the
J ) 15.6, 7.0, 1.9 Ηz), 6.05-5.92 (m, 2H), 5.82 (ddt, 1H, J )
preparation of unlabeled aldehyde 17b was employed. Thus,
reaction of labeled acetal 16a (2.39 g, 11.2 mmol) with
saturated aqueous oxalic acid (10 mL) in THF (30 mL) afforded
crude 17a still containing some solvent. Due to the high
volatility of the product, this material was used in the next
reaction without further purification.
162.8, 15.6, 1.4 Hz), 5.59 (ddq, 1H, J ) 150.2, 14.2, 6.9 Hz),
1.73 (dd, 3H, J ) 6.9, 6.8 Hz); 13C NMR (75.5 MHz, CDCl
) δ
3
172.30 (d, J ) 69.8 Hz), 152.46 (d, J ) 69.4 Hz), 131.64 (d, J
) 70.9 Hz), 34.54 (d, J ) 3.5 Hz), 18.01 (d, J ) 43.8 Hz); MS
(EI) calcd for 13
C C H O 210.1536, found 210.1527 (M , 4.3),
2
+
2 11 20 2
1
3
2
165.1548 (19). Anal. Calcd for
9.59. Found: C, 74.91; H, 9.44.
2 11 20 2
C C H O : C, 75.20; H,
E t h yl (6R)-(E,E,E)-6-Met h yld od eca -2,8,10-t r ien oa t e
(
(
18b). A solution of aldehyde 17b (1.25 g, 7.54 mmol) and
carbethoxymethylene)triphenylphosphorane (3.15 g, 9.04 mmol)
in dry benzene (150 mL) was heated to reflux at 85 °C for 17
Eth yl (1S, 2S, 4a R, 6R, 8a S)-1,2,4a ,5,6,7,8,8a -Octa h y-
d r o-2,6-d im eth yln a p h th a len e-1-ca r boxyla te (21). Tribu-
tyltin hydride (0.5 mL, 1.86 mmol) and freshly recrystallized
R,R′-azobis(isobutyronitrile) (AIBN, ∼5 mg) were stirred with
2
4
h. The mixture was cooled to room temperature, and the
solvent was removed in vacuo to give a pale yellow slurry. The
slurry was partially purified on flash silica (40 × 220 mm, 5%
3
3
thioketal 28 (55.4 mg, 0.170 mmol) at 120 °C for 4 days. After
cooling, the mixture was purified by flash chromatograghy
(SiO ; 2% Et O in pentane) to give the desired fully reduced
2 f
Et O in pentane, R 0.41) to yield a mixture of trienes (1.42 g,
2
2
8
0%). This oil was separated using MPLC with AgNO
silica gel, to give the 2E,8E,10Z-triene impurity 19b (126 mg,
%), the desired E,E,E-triene 18b (1.11 g, 62%), and some
mixed material (95.8 mg, 5%). The E,E,E-triene was further
purified on flash silica (5% Et O in pentane, R 0.41) to yield
-7.59° (c 1.1, CHCl );
cast) 1722 (s), 1655 (m) cm ; H NMR (400 MHz,
3
-stained
product 21 (30.8 mg, 77%, R 0.20), and the mercapto inter-
f
mediate 29 (8.8 mg, 22%, R 0.16) as clear oils. Data for 21:
f
2
0
-1
7
[R] +143.3° (c 0.72, CHCl ); IR (CH Cl cast) 1736 (s) cm ;
D
3
2
2
1
H NMR (400 MHz, CDCl ) δ 5.56 (ddd, 1H, J ) 9.9, 4.0, 3.0
3
2
f
Hz), 5.33 (br d, 1H, J ) 9.9 Hz), 4.15 (q, 2H, J ) 7.2 Hz), 2.61-
2.49 (m, 2H), 2.10-2.00 (m, 1H), 1.93 (dddm, 1H, J ) 12.8,
12.5, 2.6 Hz, 1.72 (dddd, 1H, J ) 12.5, 3.9, 3.9, 3.7 Hz), 1.63
(dddd, 1H, J ) 13.2, 12.9, 3.9, 3.9 Hz), 1.56-1.53 (m, 1H),
1.53-1.48 (m, 1H), 1.45-1.35 (m, 1H), 1.33 (ddd, 1H, J ) 13.0,
12.8, 4.8 Hz), 1.26 (t, 3H, J ) 7.0 Hz), 1.10 (dddd, 1H, J )
12.9, 12.9, 11.9, 3.7 Hz), 0.99 (d, 3H, J ) 7.2 Hz), 0.91 (d, 3H,
1
8b (834 mg, 47%) as a clear oil: [R]20
D
-
3
1 1
IR (CHCl
CDCl
1
3
3
) δ 6.95 (dt, 1Η, J ) 15.6, 6.9 Ηz), 6.02 (ddq, 1H, J )
4.2, 10.4, 1.5 Hz), 5.97 (ddt, 1H, J ) 14.4, 10.4, 1.3 Hz), 5.81
(
(
(
dt, 1H, J ) 15.6, 1.6 Hz), 5.58 (dq, 1H, J ) 14.2, 6.9 Hz), 5.50
dt, 1H, J ) 14.4, 7.3 Hz), 4.17 (q, 2H, J ) 6.5 Hz), 2.29-2.10
m, 2H), 2.06 (ddd, 1H, J ) 13.5, 7.3, 6.1 Hz), 1.93 (ddd, 1H,
J ) 7.0 Hz); 13C NMR (100 MHz, CDCl ) δ 173.88, 131.24,
3
J ) 13.5, 7.3, 7.0 Hz), 1.73 (d, 3H, J ) 6.9 Hz), 1.56-1.43 (m,
2
131.21, 59.76, 49.60, 38.73, 37.20, 35.66, 32.35, 31.94, 27.63,
24.49, 18.37, 17.79, 14.40; MS (EI) calcd for C H O 236.1776,
H), 1.28-1.19 (m, 1H), 1.28 (t, 3H, J ) 6.5 Hz), 0.90 (d, 3H,
1
5
24
2
1
3
+
J ) 6.4 Hz); C NMR (100 MHz, CDCl
3
) δ 166.81, 149.48,
31.89, 131.60, 129.90, 127.14, 121.23, 60.16, 39.93, 34.69,
2.79, 29.86, 19.32, 18.02, 14.31; MS (EI) calcd for C15
found 236.1778 (M , 10), 191.1428 (7), 162.1410 (100). Anal.
Calcd for C H O : C, 76.23; H, 10.23. Found: C, 76.50; H,
1
3
2
1
5
24
2
H
24
O
2
10.19.
+
36.1776, found 236.1775 (M , 9), 190.1353 (6). Anal. Calcd
Eth yl (1S,2S,4aR,6S,8S,8aS)-1,2,4a,5,6,7,8,8a-Octah ydr o-
-h yd r oxy-6-(h yd r oxym et h yl)-2-m et h yln a p h t h a len e-1-
for C15
H
24
O
2
: C, 76.21; H, 10.24. Found: C, 76.40; H, 10.22.
]-La beled 18 (18a ). The same method as for the
8
1
3
[
2,11- C
2
ca r boxyla te (23). A solution of the lactone 22 (0.476 g, 1.80
mmol) in dry THF (20 mL) was cooled to 0 °C, and lithium
preparation of 18b was employed. The spectral and chro-
matographic properties were similar to those of 18b except
for: H NMR (400 MHz, benzene-d
7
J ) 161.1, 15.7, 1.6 Hz), 5.52 (ddq, 1H, J ) 150.5, 14.2, 7.0
Hz), 1.92-1.70 (m, 4H, H-4 and H-7), 1.60 (dd, 3H, J ) 7.0,
7
triethylborohydride (LiBEt H; 1.0 M in THF, 2.34 mmol) was
3
1
27e
6
) δ 7.03 (dtd, 1Η, J ) 15.7,
added slowly (15 min). Further portions of lithium trieth-
.0, 2.0 Ηz), 6.08-5.96 (m, 2H, H-9 and H-10), 5.87 (ddt, 1H,
ylborohydride (1.26 mmol) were added to react with intermedi-
ates generated, but the additions were limited so as to not fully
reduce the starting material (TLC monitoring). H O (0.5 mL)
2
1
3
.0 Hz, H-12); C NMR (100 MHz, benzene-d
6
) δ 166.81 (d, J
was carefully added to destroy excess reagent, followed by 2
N NaOH(aq) (1 mL) and 30% H O solution (1 mL) dropwise.
)
75.5 Hz), 149.10 (d, J ) 69.4 Hz), 132.31 (d, J ) 71.4 Hz),
2
2
3
4.88 (d, J ) 3.0 Hz), 18.04 (d, J ) 43.3 Hz); MS (EI) calcd for
The resulting cloudy mixture was stirred vigorously for 1 h at
1
3
2
+
C
2
C
13
H
24
O
2
238.1843, found 238.1834 (M , 7), 209.1447 (1.1),
room temperature and then poured into Et O (100 mL). The
2
1
3
2
1
1
65.1548 (30). Anal. Calcd for
0.15. Found: C, 76.21; H, 10.14.
6R)-E,E,E-6-Meth yld od eca -2,8,10-tr ien oic Acid (20b).
C
2
C
13
H
24
O
2
: C, 76.42; H,
organic layer was washed with brine (20 mL), and the aqueous
layer was extracted with Et O (2 × 50 mL). The combined
2
(
4
organic layers were dried (MgSO ) and evaporated in vacuo.
A solution of triene ethyl ester 18b (1.00 g, 4.23 mmol) in
Flash chromatography (SiO
resultant oil yielded two products: the desired product 23 (312
mg, 65%, R 0.20) and the triol 24 (9.88 mg, 20%, R 0.07) as
a colorless solids. Data for diol 23: mp 100-100.5 °C; [R]
+142.0° (c 0.29, CHCl ); IR (CH Cl cast) 3350 (br m), 1732
(s), 1712 (s) cm ; H NMR (400 MHz, CDCl ) δ 5.53 (ddd, 1H,
2 2
; 10% pentane in Et O) of the
distilled THF (50 mL) was treated with aqueous 3 M KOH
(
10 mL), and the mixture was stirred for 18 h at 50 °C.24 Most
f
f
2
0
of the THF was removed in vacuo, and the cloudy solution was
stirred at 50 °C until it became clear. After cooling, H O (500
mL) was added and this solution was washed with pentane (2
50 mL). The aqueous layer was acidified to pH 7 (1 N HCl)
and then extracted with Et
O (3 × 200 mL). The combined
D
2
3
2
2
-
1 1
3
×
J ) 9.8, 4.2, 2.8 Hz), 5.39 (br d, 1H, J ) 9.8 Hz), 4.24 (br d,
1H, J ) 2.1 Hz), 4.20-4.10 (m, 2H), 3.78-3.69 (m, 2H), 3.30
2

2
622 J . Org. Chem., Vol. 61, No. 8, 1996
Witter and Vederas
(
2
1
(
(
1
3
1
9
br s, 2H), 2.82 (dd, 1H, J ) 11.9, 6.6 Hz), 2.65-2.54 (m, 1H),
12.9, 12.9, 4.9 Hz), 1.69 (dp, 1H, J ) 13.3, 1.8 Hz), 1.26 (t, 3H,
J ) 7.1 Hz), 0.98 (d, 3H, J ) 7.2 Hz), 0.89 (d, 3H, J ) 7.1 Hz);
.52-2.42 (m, 1H), 2.01-1.95 (m, 1H), 1.95-1.83 (m, 2H),
.83-1.75 (m, 1H), 1.47 (ddd, 1H, J ) 11.4, 11.0, 1.7 Hz), 1.35
1
3
C NMR (100 MHz, CDCl ) δ 210.50, 173.76, 132.15, 128.87,
3
ddd, 1H, J ) 13.4, 13.4, 6.1 Hz), 1.26 (t, 3H, J ) 7.1 Hz), 0.91
60.22, 49.53, 48.11, 42.83, 38.01, 37.82, 31.35, 31.26, 19.39,
17.80, 14.28; MS (EI) calcd for C15 250.1569, found
250.1567 (M , 22), 204.1149 (76), 176.1198 (100). Anal. Calcd
for C15 : C, 71.97; H, 8.86. Found: C, 71.71; H, 8.87.
d, 3H, J ) 7.1 Hz); 13C NMR (100 MHz, CDCl
) δ 173.91,
3
22 3
H O
+
31.19, 130.54, 67.98, 65.69, 60.00, 45.12, 39.85, 35.73, 35.04,
+
3.99, 32.46, 30.52, 17.63, 14.31; MS (CI, NH
3
) 286 (MNH
4
,
22 3
H O
+
0), 269 (MH , 100). Anal. Calcd for C15
.01. Found: C, 66.74; H, 9.36.
H
24
O
4
: C, 67.14; H,
E t h yl (1S,2S,4a R ,6S,8a S)-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-
8-(d im et h ylen ed it h io)-2,6-d im et h yln a p h t h a len e-1-ca r -
boxyla te (28). A solution of ketone 27 (0.244 g, 0.977 mmol)
Eth yl (1S,2S,4aR,6S,8S,8aS)-1,2,4a,5,6,7,8,8a-Octah ydr o-
-h yd r oxy-2-m eth yl-6-[[[(m eth ylth io)th ioca r bon yl]oxy]-
8
2 2
in dry CH Cl (8 mL) was treated with 1,2-ethanedithiol (0.164
m eth yl]n a p h th a len e-1-ca r boxyla te (25). A solution of
tetrabutylammonium hydrogen sulfate (691 mg, 2.04 mmol)
and diol 23 (497 mg, 1.85 mmol) in distilled benzene (15 mL)
was treated (with vigorous stirring), with 4 N NaOH(aq) (15
mL), followed by a quick addition of carbon disulfide (0.23 mL,
mL, 1.95 mmol) and boron trifluoride etherate (0.120 mL, 0.977
mmol) at 0 °C.32 The mixture was then warmed to room
temperature and stirred overnight. The reaction mixture was
diluted with CH
mL), 1 N HCl (10 mL), H
solvent was removed in vacuo, and the resultant oil was
purified by flash chromatography (SiO ; 10% Et O in pentane,
0.22) to yield 28 (212 mg, 66%) as a clear colorless oil: [R]
2
Cl
2
(50 mL) and washed with 1 N NaOH (10
2
O (10 mL), and brine (10 mL). The
3
1
.70 mmol) and methyl iodide (0.173 mL, 2.78 mmol).30 After
0 min, ice (30 g) and Et
2
O (30 mL) were added to quench the
O (2 × 30
mL) and the combined organic layers were washed with brine
15 mL), dried (MgSO ), and evaporated in vacuo. The
resultant pale orange oil was purified by flash chromatograghy
SiO ; 30% Et O in pentane, R 0.30) to give 25 (564 mg, 85%)
as a colorless oil: [R] +162.9° (c 0.42, CHCl ); IR (CH Cl
cast) 3500 (br m) 1731 (s), 1714 (s) cm ; H NMR (400 MHz,
CDCl ) δ 5.57 (ddd, 1H, J ) 9.8, 4.6, 2.6 Hz), 5.39 (br d, 1H, J
2
2
2
0
reaction. The aqueous layer was extracted with Et
2
R
f
D
-1 1
+84.3° (c 0.22, CHCl
(300 MHz, CDCl ) δ 5.64 (br s, 2H), 4.11 (dm, 2H, J ) 3.0 Hz),
3 2 2
); IR (CH Cl cast) 1730 (s) cm ; H NMR
(
4
3
3.43-3.33 (m, 1H), 3.33-3.28 (m, 1H), 3.28-3.20 (m, 2H), 3.10
(dd, 1H, J ) 7.8, 6.2 Hz), 2.47 (br dq, 1H, J ) 7.8, 7.4 Hz),
2.32-2.22 (m, 2H), 2.21 (dd, 1H, J ) 10.9, 6.2 Hz), 2.20-2.10
(m, 1H), 2.12-2.00 (br t, 1H, J ) 12.3 Hz), 1.75 (dm, 1H, J )
13.3 Hz), 1.62 (ddd, 1H, J ) 12.7, 12.7, 5.3 Hz), 1.24 (t, 3H, J
) 7.2 Hz), 1.21 (d, 3H, J ) 7.5 Hz), 1.10 (d, 3H, J ) 7.4 Hz);
(
2
2
f
2
0
D
3
2
2
-
1 1
3
)
9.8 Hz), 4.90 (dd, 1H, J ) 10.9, 9.3 Hz), 4.76 (dd, 1H, J )
1
3
1
2
0.9, 6.1 Hz), 4.31 (dd, 1H, J ) 2.6, 2.5 Hz), 4.21-4.09 (m,
H), 2.85 (dd, 1H, J ) 11.6, 5.9 Hz), 2.66-2.59 (m, 1H), 2.56
C NMR (100 MHz, CDCl
59.95, 54.22, 50.91, 49.74, 40.97, 38.50, 37.99, 33.71, 31.93,
29.50, 20.46, 17.64, 14.33; MS (EI) calcd for C17
3
) δ 176.20, 134.15, 133.37, 72.68,
(
2
1
s, 3H), 2.51-2.41 (m, 1H), 2.41-2.33 (m, 1H), 1.92-1.76 (m,
26 2 2
H O S
+
H), 1.60-1.50 (m, 2H), 1.33 (ddd, 1H, J ) 13.4, 13.4, 5.1 Hz),
326.1374, found 326.1369 (M , 60), 298.1050 (68), 219.0844
.26 (t, 3H, J ) 7.1 Hz), 0.92 (d, 3H, J ) 7.1 Hz); 13C NMR
(27), 159.1168 (100). Anal. Calcd for C17
H, 8.03. Found: C, 62.81; H, 8.34.
26 2 2
H O S : C, 62.54;
(
3
100 MHz, CDCl ) δ 215.94, 173.69, 131.14, 130.66, 77.18,
6
1
6
6.52, 60.04, 44.99, 40.02, 34.86, 33.42, 32.48, 31.88, 28.98,
Diels-Ald er Cycliza tion . (A) Th er m a l Rea ction : A
solution of triene (20.0 mg, 84.9 µmol) in dry toluene (1 mL)
+
+
8.96, 17.54, 14.33; MS (CI, NH
3
) 376 (MNH
: C, 56.95; H, 7.31.
4
, 10), 359 (MH ,
), 268 (100). Anal. Calcd for C17
H
26
O
4
S
2
was placed in a thick-walled glass tube, and the solution was
Found: C, 57.16; H, 7.52.
degassed with a stream of argon for 5 min.36 The tube was
E t h yl (1S,2S,4a R ,6S,8a S)-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-
-m et h yl-6-[[[(m et h ylt h io)t h ioca r b on yl]oxy]m et h yl]-8-
sealed under argon and then heated in an oil bath at 160 °C
for 43 h. The tube was cooled and opened, and the solvent
was removed in vacuo. Flash chromatography of the clear
residue yielded the trans-fused product, the cis-fused product,
and unreacted starting material as clear oils. (B) Lew is Acid -
Ca ta lyzed Rea ction : Ethylaluminum dichloride (240 µL, 54.0
µmol, 1.8 M in toluene) was slowly added to a solution of triene
2
oxon a p h th a len e-1-ca r boxyla te (26). A solution of alcohol
5 (1.14 g, 3.18 mmol) and pyridinium dichromate (1.79 g, 4.76
mmol) in dry CH Cl (5 mL) was stirred at room temperature
for 36 h. The solvent was removed in vacuo, and the residue
was purified by flash chromatography (SiO ; 40% Et O in
0.30) to yield the desired keto-ester 26 (0.908 g,
2
2
2
4
6
2
2
pentane, R
f
(13.4 mg, 56.8 µmol) in dry toluene (0.5 mL), and the mixture
2
0
36
8
1
0%) as a colorless waxy solid: mp 82-82.5 °C; [R]
D
+160.5°(c
) 1733 (s), 1717 (s) cm ; H NMR (400
) δ 5.66 (ddd, 1H, J ) 9.8, 4.5, 2.6 Hz), 5.45 (ddd,
H, J ) 9.8, 1.6, 1.5 Hz), 4.52 (dd, 1H, J ) 11.1, 7.9 Hz), 4.45
dd, 1H, J ) 11.1, 6.9 Hz), 4.22-4.09 (m, 2H), 2.90-2.60 (m,
was stirred at room temperature for 3 h.
The reaction
-1
1
.19, CHCl
3
); IR (CH
3
2
Cl
2
mixture was then poured into 1 N HCl (1.0 mL), and the
MHz, CDCl
aqueous layer was extracted with Et O (3 × 10 mL). The
2
1
(
5
(
(
(
2
4
combined organic layers were washed with saturated aqueous
NaHCO (2 mL), dried (MgSO ), and concentrated in vacuo.
3
4
H), 2.73 (s, 3H), 2.38 (ddd, 1H, J ) 13.7, 1.8, 1.8 Hz), 2.29
The residue was purified by flash chromatography to yield the
trans-fused product, the cis-fused product, and unreacted
starting material.
ddm, 1H, J ) 11.8, 11.8 Hz), 1.97 (dm, 1H, J ) 14.1 Hz), 1.79
ddd, 1H, J ) 13.4, 13.4, 5.3 Hz), 1.26 (t, 3H, J ) 7.2 Hz), 0.89
d, 3H, J ) 7.2 Hz); 13C NMR (100 MHz, CDCl
) δ 215.75,
3
Eth yl (1R,2R,4a S, 6R,8a R)-1,2,4a ,5,6,7,8,8a -octa h yd r o-
09.03, 173.51, 132.50, 128.03, 74.47, 60.30, 49.24, 42.99,
2.63, 38.03, 35.40, 32.92, 31.15, 19.15, 17.72, 14.26; MS (EI)
20
2
,6-d im eth yln a p h th a len e-1-ca r boxyla te (30): [R]
D
-89.5°
-1
1
(c 1.40, CHCl
3
); IR (CH
2 2
Cl cast) 1737 (s) cm ; H NMR (400
+
calcd for C17
10.0696 (16), 202.0989 (100). Anal. Calcd for C17
C, 57.28; H, 6.79. Found: C, 56.98; H, 6.42.
24 4 2
H O S 356.1116, found 356.1110 (M , 18),
MHz, CDCl
3
) δ 5.53 (ddd, 1H, J ) 9.9, 4.0, 2.7 Hz), 5.38 (br d,
3
24 4 2
H O S :
1
H, J ) 9.9 Hz), 4.13 (q, 2H, J ) 7.1 Hz), 2.55 (m, 1H), 2.53
(m, 1H), 1.95 (dddd, 1H, J ) 12.0, 3.2, 3.0, 2.9 Hz), 1.77-1.68
E t h yl (1S,2S,4a R ,6S,8a S)-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-
,6-d im eth yl-8-oxon a p h th a len e-1-ca r boxyla te (27). A so-
(m, 3H), 1.52-1.43 (m , 1H), 1.35 (dddd, 1H, J ) 12.0, 12.0,
12.0, 3.0 Hz), 1.27 (t, 3H, J ) 7.1 Hz), 1.04 (dddd, 1H, J )
12.0, 12.0, 12.0, 3.0 Hz), 1.00-0.95 (m, 1H), 0.93 (d, 3H, J )
6.7 Hz), 0.90 (d, 3H, J ) 6.8 Hz), 0.80 (ddd, 1H, J ) 12.0, 12.0,
2
lution of tributyltin hydride (0.133 mL, 0.496 mmol) in dry
cymene (2 mL) was slowly added to a solution of methyl
xanthate 26 (22.1 mg, 0.062 mmol) in cymene (2 mL) at 150
12.0 Hz); 13C NMR (100 MHz, CDCl ) δ 173.97, 130.93, 130.92,
3
3
1
°
C over 1.25 h. The mixture was allowed to stir overnight
at 150 °C. The solvent was removed in vacuo and the resultant
oil purified by flash chromatography (SiO ; 40% Et O in
pentane, R 0.40) to yield 27 (9.75 mg, 63%) as a clear colorless
solid: mp 34-35 °C; [R]
59.79, 49.51, 41.89, 41.63, 36.20, 35.27, 33.10, 32.46, 29.94,
22.53, 17.75, 14.40; MS (EI) calcd for C H O 236.1776, found
1
5
24
2
+
2
2
236.1775 (M , 11), 191.1429 (7), 162.1406 (100). Anal. Calcd
for C H O : C, 76.23; H, 10.23. Found: C, 76.20; H, 10.20.
f
1
5
24
2
2
0
D
+80.8° (c 0.13, CHCl
3
); IR (CH
cast) 1736 (s), 1718 (s) cm ; H NMR (400 MHz, CDCl ) δ 5.64
ddd, 1H, J ) 9.8, 4.5, 2.8 Hz), 5.45 (ddd, 1H, J ) 9.8, 1.6, 1.5
Hz), 4.23-4.09 (m, 2H), 2.82 (dd, 1H, J ) 11.5, 6.6 Hz), 2.75
dd, 1H, J ) 12.6, 6.6 Hz), 2.70-2.50 (m, 3H), 2.37-2.28 (m,
2 2
Cl
Eth yl (1R,2S,4a R,6R,8a R)-1,2,4a ,5,6,7,8,8a -octa h yd r o-
-
1 1
20
3
2
,6-d im eth yln a p h th a len e-1-ca r boxyla te (31): [R]
+6.95°
cast) 1732 (s) cm ; H NMR (400
) δ 5.54 (ddd, 1H, J ) 10.0, 4.2, 2.4 Hz), 5.42 (ddd,
D
(
-1 1
3 2 2
(c 0.81, CHCl ); IR (CH Cl
MHz, CDCl
3
(
1
1
1
2
H, J ) 10.0, 1.6, 1.5 Hz), 4.17 (q, 1H, J ) 7.1 Hz), 4.16 (q,
H), 2.15 (ddd, 1H, J ) 12.6, 1.9, 1.9 Hz), 1.78 (ddd, 1H, J )
H, J ) 7.07 Hz), 2.52 (dqdd, 1H, J ) 8.7, 6.1, 4.2, 1.6 Hz),
.37 (dd, 1H, J ) 10.0, 8.7 Hz), 2.29 (m, 1H), 2.05 (m, 1H),
(
46) Furber, M.; Mander, L. N. J . Am. Chem. Soc. 1987, 109, 6389-
1.94-1.83 (m , 1H), 1.74-1.64 (m, 2H), 1.48-1.35 (m, 2H),
1.38-1.29 (m, 1H), 1.28 (t, 3H, J ) 7.1 Hz), 1.18 (ddd, 1H, J
6
396.

Diels-Alder Cyclization during Biosynthesis of Lovastatin
9.7, 6.0, 5.0 Hz), 0.99 (d, 3H, J ) 6.1 Hz), 0.97 (d, 3H, J )
J . Org. Chem., Vol. 61, No. 8, 1996 2623
)
(1R ,2R ,4a S, 6R ,8a R )-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-2,6-
d im eth yl-1-(h yd r oxym eth yl)n a p h th a len e (36). The same
method as for the preparation of alcohol 37 was employed.
Thus, reduction of ethyl ester 30 (40.8 mg, 0.173 mmol) with
1
3
6
6
2
2
.0 Hz); C NMR (100 MHz, CDCl
0.08, 47.92, 36.31, 36.19, 34.04, 31.01, 28.06, 27.41, 24.03,
0.61, 18.66, 14.43; MS (EI) calcd for C15 236.1776, found
36.1776 (M , 13), 162.1410 (100). Anal. Calcd for C15
3
) δ 176.22, 131.31, 130.83,
24 2
H O
+
H
24
O
2
:
LiAlH
64-65 °C; [R]2
3600-3100 (br m), 3008 (m), 1455 (m) cm ; H NMR (300
MHz, CDCl ) δ 5.58 (ddd, 1H, J ) 9.7, 4.6, 2.7 Hz), 5.36 (br d,
4
(26.2 mg, 0.690 mmol) afforded 36 (26.9 mg, 80%): mp
0
C, 76.23; H, 10.23. Found: C, 76.07; H, 10.01.
D
2 2 2 2
+70.37° (c 0.054, CH Cl ); IR (CH Cl cast)
-
1
1
(
1R ,2R ,4a S, 6R ,8a R )-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-2,6-
2
0
d im eth yln a p h th a len e-1-ca r boxylic a cid (32): [R]
D
-68.0°
3
(
c 0.10, CH
2
Cl
2
); IR (CH
2
Cl
2
cast) 3600-2400 (br m), 1705 (s)
) δ 5.55 (ddd, 1H, J ) 10.0,
1H, J ) 9.7 Hz), 3.83 (dd, 1H, J ) 10.7, 5.5 Hz), 3.53 (dd, 1H,
J ) 10.7, 9.3 Hz), 2.50-2.38 (m, 1H), 1.80-1.63 (m, 5H), 1.51-
1.36 (m , 2H), 1.11-0.92 (m, 3H), 0.92 (d, 3H, J ) 7.1 Hz),
0.89 (d, 3H, J ) 6.6 Hz), 0.73 (ddd, 1H, J ) 12.2, 12.2, 12.2
-
1
1
cm ; H NMR (300 MHz, CDCl
3
4
2
1
.1, 2.8 Hz), 5.39 (br d, 1H, J ) 10.0 Hz), 2.65-2.50 (m, 2H),
.08-2.00 (m, 1H), 1.80-1.68 (m, 3H), 1.54-1.42 (m, 1H),
.42-1.28 (m, 1H), 1.10-0.90 (m, 2H), 0.95 (d, 3H, J ) 6.7
Hz); 13C NMR (75.5 MHz, CDCl
) δ 132.34, 131.19, 63.21,
44.13, 43.34, 41.83, 37.34, 35.53, 32.82, 31.65, 29.16, 22.61,
3
Hz), 0.90 (d, 3H, J ) 6.8 Hz), 0.79 (ddd, 1H, J ) 12.0, 12.0,
1
3
+
1
4
1
2.0 Hz); C NMR (75.5 MHz, CDCl
9.39, 41.73, 41.56, 35.96, 35.22, 33.05, 32.33, 29.91, 22.49,
7.65; MS (EI) calcd for C13 208.1463, found 208.1469
: C, 74.96;
3
) δ 180.77, 130.92, 130.62,
15.51; MS (EI) calcd for C13H22O 194.1671, found 194.1669 (M ,
10), 163.1487 (100). Anal. Calcd for C13
11.41. Found: C, 80.15; H, 11.44.
H22O: C, 80.35; H,
22 2
H O
+
(
M , 26), 163.1488 (100). Anal. Calcd for C13
H, 9.68. Found: C, 74.72; H, 9.41.
1R,2S,4a R,6R,8a R)-1,2,4a ,5,6,7,8,8a -Octa h yd r o-2,6-d i-
m eth yln a p h th a len e-1-ca r boxylic a cid (33): IR (CH Cl
cast) 3600-2400 (br m), 1704 (s) cm ; H NMR (300 MHz,
CDCl ) δ 5.55 (ddd, 1H, J ) 10.0, 4.1, 2.8 Hz), 5.44 (ddd, 1H,
J ) 10.0, 2.0, 2.0 Hz), 2.58-2.50 (m, 1H), 2.39 (dd, 1H, J )
.5, 8.0 Hz), 2.37-2.29 (m, 1H), 2.11-2.05 (m, 1H), 1.89-1.80
m, 1H), 1.75-1.65 (m, 2H), 1.48-1.35 (m, 2H), 1.30-1.25 (m,
H), 1.20.1.11 (m, 1H), 1.04 (d, 3H, J ) 6.2 Hz), 0.98 (d, 3H,
H
22
O
2
(1R,2S,4a R,6R,8a R)-1,2,4a ,5,6,7,8,8a -Octa h yd r o-2,6-d i-
m eth yl-1-(h yd r oxym eth yl)n a p h th a len e (37). A solution of
(
ethyl ester 31 (45.4 mg, 0.192 mmol) in dry Et
2
O (0.7 mL) was
O (1.7
2
2
added to a slurry of LiAlH (29.2 mg, 7.68 mmol) in Et
4
2
-
1
1
37
mL) at 0 °C. After stirring at room temperature for 2 h, the
reaction mixture was cooled to 0 °C and treated sequentially
3
with H
mixture was allowed to warm to room temperature and was
stirred for an additional 1 h. MgSO was added to the stirring
2
O (33 µL), 3 M NaOH (33 µL), and H
2
O (100 µL). The
9
(
1
4
solution, and the mixture was filtered. The filtrate was
concentrated in vacuo, and the residue was purified by flash
1
3
J ) 6.1 Hz); C NMR (100 MHz, CDCl
3
) δ 182.89, 131.17,
30.46, 48.05, 36.43, 36.05, 33.46, 30.91, 28.63, 27.34, 24.37,
0.74, 18.94; MS (EI) calcd for C13 208.1463, found
08.1461 (M , 21), 163.1488 (100). Anal. Calcd for C13
1
2
2
chromatography (SiO
2 2 f
; 30% Et O in pentane, R 0.35) to yield
22 2
H O
the desired alcohol 37 (32.0 mg, 86%) as a colorless solid: mp
+
20
H
22
O
2
:
59.5-60 °C; [R]
D
+28.6° (c 0.028, CH
2 2 2 2
Cl ); IR (CH Cl cast)
-
1 1
C, 74.96; H, 9.68. Found: C, 74.85; H, 9.80.
1R ,2R ,4a S, 6R ,8a R )-1,2,4a ,5,6,7,8,8a -Oct a h yd r o-2,6-
d im et h yln a p h t h a len e-1-ca r b oxylic a cid , N-a cet ylcys-
3600-3100 (br m), 1454 (m) cm ; H NMR (400 MHz, CDCl )
3
(
δ 5.50 (ddd, 1H, J ) 10.1, 2.8, 2.8 Hz), 5.40 (ddd, 1H, J )
10.0, 2.0, 2.0 Hz), 3.58 (d, 2H, J ) 6.8 Hz), 2.35-2.28 (m, 1H),
2.00-1.90 (m, 1H), 1.82-1.75 (m, 1H), 1.69-1.57 (m, 3H),
1.57-1.42 (m, 2H), 1.39-1.32 (m, 1H), 1.25-1.19 (m, 1H), 1.09
(d, 3H, J ) 7.5 Hz), 0.99-0.90 (m, 1H), 0.87 (d, 3H, J ) 6.6
2
0
tea m in e th ioester (34): mp 94-95 °C; [R]
CH Cl ); IR (CH Cl
s) cm ; H NMR (400 MHz, CDCl
ddd, 1H, J ) 10.0, 4.2, 2.7 Hz), 5.38 (br d, 1H, J ) 10.0 Hz),
D
-31.5° (c 0.41,
cast) 3550-3100 (br m), 1687 (s), 1657
) δ 5.83 (br s, 1H), 5.53
2
2
-
2
2
1
1
(
(
3
Hz); 13C NMR (100 MHz, CDCl
) δ 131.60, 129.80, 65.51, 46.13,
39.59, 34.79, 34.35, 31.07, 30.66, 28.15, 27.78, 22.17, 21.92;
3
3
1
4
6
1
1
2
3
.49-3.41 (m, 2H), 3.03 (t, 2H, J ) 6.3 Hz), 2.85 (dd, 1H, J )
1.2, 5.8 Hz), 2.63-2.55 (m, 1H), 1.95 (s, 3H), 1.83-1.65 (m,
H), 1.50-1.45 (m, 2H), 1.09-0.90 (m, 2H), 0.89 (d, 3H, J )
.6 Hz), 0.88 (d, 3H, J ) 7.1 Hz), 0.78 (ddd, 1H, J ) 12.0, 12.0,
+
MS (EI) calcd for C13H22O 194.1671, found 194.1668 (M , 9),
163.1485 (100). Anal. Calcd for C13
Found: C, 80.07; H, 11.52.
H22O: C, 80.35; H, 11.41.
1
3
2.0 Hz); C NMR (100 MHz, CDCl
30.62, 58.62, 42.07, 41.48, 40.23, 36.81, 35.12, 33.59, 33.00,
9.46, 28.28, 23.28, 22.46, 17.42; MS (EI) calcd for C17
3
) δ 202.00, 171.80, 130.92,
Ack n ow led gm en t. We thank Dr. Yuko Yoshizawa
for extensive assistance with incorporation experiments.
Cambridge Isotope Laboratories (Woburn, MA) gener-
ously provided isotopically labeled starting materials
through the CIL Research Grant Program. Merck
Research Laboratories (Rahway, NJ ) donated freeze-
dried specimens of A. terreus MF 4845. These investiga-
tions were supported by the Natural Sciences and
Engineering Research Council of Canada and the Al-
berta Heritage Foundation for Medical Research.
H
2
27NO S
+
09.1763, found 309.1760 (M , 1.4), 190.1359 (24), 163.1487
100). Anal. Calcd for C17 S: C, 65.98; H, 8.79.
(
H27NO
2
Found: C, 65.70; H, 9.28.
(
1R,2S,4a R,6R,8a R)-1,2,4a ,5,6,7,8,8a -Octa h yd r o-2,6-d i-
m eth yln a p h th a len e-1-ca r boxylic a cid , N-a cetylcystea m -
in e th ioester (35): IR (CH Cl cast) 3550-3100 (br m) 1683
s), 1655 (s) cm ; H NMR (500 MHz, CDCl ) δ 5.83 (br s,
2
2
-
1
1
(
3
1
1
2
H), 5.53 (ddd, 1H, J ) 10.0, 4.2, 2.4 Hz), 5.43 (ddd, 1H, J )
0.0, 2.0, 2.0 Hz), 3.50-3.40 (m, 2H), 3.06 (t, 1H, J ) 6.4 Hz),
.62 (dd, 1H, J ) 9.3, 8.4 Hz), 2.61-2.50 (m, 1H), 2.34-2.27
(
2
1
m, 1H), 2.13-2.07 (m, 1H), 1.92-1.82 (m, 1H), 1.73-1.63 (m,
Su p p or tin g In for m a tion Ava ila ble: Full spectral as-
signments for listed compounds, data for compounds 13, 16,
7 (2Z,4E-isomer), 19a ,b, 24, and 29 and general experimental
procedures (13 pages). This material is contained in many
libraries on microfiche, immediately follows this article in the
microfilm version of the journal, and can be ordered from the
ACS; see any current masthead page for ordering information.
H), 1.48-1.35 (m, 2H), 1.38-1.32 (m, 1H), 1.22-1.15 (m, 1H),
.02 (d, 3H, J ) 7.1 Hz), 0.96 (d, 3H, J ) 7.1 Hz); 13C NMR
1
3
(100 MHz, CDCl ) δ 201.42, 170.24, 131.26, 130.62, 56.55,
4
2
0.22, 40.07, 36.99, 36.31, 34.27, 33.59, 31.15, 28.39, 28.28,
3.27, 22.46, 20.61; MS (EI) calcd for C17 S 309.1763,
H
27NO
2
+
found 309.1760 (M , 1.7), 190.1355 (19), 163.1482 (100); MS
+
+
(CI, NH
3
) 327 (MNH
S: C, 65.98; H, 8.79. Found: C, 65.82; H, 8.95.
4
, 21), 310 (MH , 100). Anal. Calcd for
C
17
H
27NO
2
J O952117P