Synthesis of 2, 3-dinor-5, 6-dihydro-15F2t-isoprostane

Article abstract of DOI:10.1021/jo9910738

A concise synthesis of the 15-F_2t-isoprostane urinary metabolite 2, a potential marker for systemic oxidative stress, is described. This synthesis confirmed the structure of this important product of human metabolism. The key transformation is the rhodium-mediated diastereoselective cyclization of diazo ketone 4, followed by cyclopropane ring opening with thiophenol and Lewis acid. Material prepared by the synthesis outlined here will be used to develop an antibody-based assay for clinically quantifying systemic oxidative stress.

Full text of DOI:10.1021/jo9910738

J . Org. Chem. 1999, 64, 7983-7987
7983
Syn th esis of 2,3-Din or -5,6-d ih yd r o-15F 2t-isop r osta n e
Douglass F. Taber* and Kazuo Kanai
Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716
Received J uly 6, 1999
A concise synthesis of the 15-F2t-isoprostane urinary metabolite 2, a potential marker for systemic
oxidative stress, is described. This synthesis confirmed the structure of this important product of
human metabolism. The key transformation is the rhodium-mediated diastereoselective cyclization
of diazo ketone 4, followed by cyclopropane ring opening with thiophenol and Lewis acid. Material
prepared by the synthesis outlined here will be used to develop an antibody-based assay for clinically
quantifying systemic oxidative stress.
In tr od u ction
It is increasingly apparent that hypercholesterolemia
is linked to an increased risk of cardiovascular desease
only under conditions of at least mild oxidative stress.1
Recently, a series of bioactive prostaglandin-like com-
pounds, the isoprostanes,2 produced (as racemates) from
membrane-bound arachidonic acid by free-radical oxida-
tion, have been described.3 These findings suggest that
quantification of these isoprostanes (e.g., 15-F2t-isopros-
tane 1) may provide a practical approach to the assess-
ment of systemic oxidative stress. Recently, Roberts and
co-workers tentatively identified the major urinary me-
tabolite of 15-F2t-isoprostane (1) as 2,3-dinor-5,6-dihydro-
15F2t-isoprostane (2) by HPLC and mass spectrometric
analysis.4a Before clinical assays based on 2 (preferred
over measuring the unmetabolized isoprostanes4b) could
be developed, it was important to first confirm the struc-
ture of this important product of human metabolism.5-11
Resu lts a n d Discu ssion
Based on our prior work,8 we proposed (Scheme 1) to
prepare 2 by aldol condensation of the ketone 5 with the
aldehyde 6, followed by rhodium(II)-catalyzed cyclization
of the silyloxy ketone 4.
The diketone 8 (Scheme 2), prepared from benzoylac-
etone (7) and 5-bromovalerate, was subjected to the diazo-
transfer reaction12 with p-nitrobenzenesulfonyl azide (p-
NBSA) and DBU to give the diazo ketone 5. Aldol
condensation of the diazo ketone 5 with (E,E)-decadienal
6 was successfully achieved by using KHMDS as base in
the presence of triethylchlorosilane (TESCl), to produce
the TES-protected aldol 9 together with a small amount
of the free aldol 10 (hydrolysis on workup) in 70%
combined yield.6d
We had shown8 that the triethylsilyl protecting group
was not stable to the conditions for cyclopropane opening,
so this was changed to tert-butyldiphenylsilyl (TBDPS).
The resulting diazo ketone 4 was then cyclized with
catalytic rhodium(II) octanoate dimer to provide the
bicyclic ketones 11 and 3 in 23% and 68% yields,
respectively. The structures of the bicyclic ketones 3 and
11 were assigned by comparing their 1H and 13C NMR
spectra to those for the analogous bicyclic ketones that
are intermediates in the synthesis of the 15-F2t-isopros-
(1) (a) Patrono, C.; FitzGerald G. A. Arterioscler. Thromb. Vascular
Biol. 1997, 17, 2309. (b) Davi, G.; Alessandrini, P.; Mezzetti, A.;
Minotti, G.; Bucciarelli, T.; Costantini, F.; Cipollone, F.; Bon, G. B.;
Ciabattoni, G.; Patrono, C. Arterioscler. Thromb. Vascular Biol. 1997,
17, 3230. (c) Gniwotta, C.; Morrow, J . D.; Roberts, L. J . II; Ku¨hn, H.
Arterioscler. Thromb. Vascular Biol. 1997, 17, 3236.
(2) For a summary of isoprostane nomenclature, see: Taber, D. F.;
Morrow, J . D. Roberts, L. J ., II. Prostaglandins 1997, 53, 63.
(3) (a) Morrow, J . D.; Hill, K. E.; Burk, R. F.; Nammour, T. M.; Badr,
K. F.; Roberts, L. J ., II. Proc. Natl. Acad. Sci. U.S.A. 1990, 87, 9383.
(b) Awad, J . A.; Morrow, J . D.; Talkahashi, K.; Roberts, L. J ., II. J .
Biol. Chem. 1993, 268, 4161. (c) Lynch, S. M.; Morrow, J . D.; Roberts,
L. J ., II; Frei, B. J . Clin. Invest. 1994, 93, 998. (d) Takahashi, K.;
Nammour, T. M.; Fukunaga, M.; Ebert, J .; Morrow, J . D.; Roberts, L.
J ., II; Hoover, R. L.; Badr, K. F. J . Clin. Invest. 1992, 90, 136. (e)
Morrow, J . D.; Minton, T. A.; Roberts, L. J ., II. Prostaglandins 1992,
44, 155. (f) Wang, Z.; Ciabattoni, G.; Cre´minon, C.; Lawson, J . A.;
FitzGerald, G. A.; Patrono, C.; Maclouf, J . J . Pharmacol. Exp. Ther.
1995, 275, 94. (g) Gopaul, N. K.; Anggard, E. E.; Mallet, A. I.;
Betteridge, D. J .; Wolff, S. P.; Nourooz-Zadeh, J . FEBS Lett. 1995, 368,
225. (h) Morrow, J . D.; Frei, B.; Longmire, A. W.; Graziano, J . M.;
Lynch, S. M.; Shyr, Y.; Strauss, W. E.; Oates, J . A.; Roberts, L. J ., II.
N. Engl. J . Med. 1995, 332, 1198.
(7) For synthetic routes to 15-F2c-isoprostane: (a) Larock, R. C.; Lee,
N. H. J . Am. Chem. Soc. 1991, 113, 7815. (b) Vionnet, J .-P.; Renaud,
P. Helv. Chim. Acta 1994, 77, 1781. (c) Hwang, S. W.; Adiyaman, M.;
Khanapure, S. P.; Rokach, J . Tetrahedron Lett. 1996, 37, 779.
(8) (a) For a synthetic route to 15-E2t-isoprostane: Taber, D. F.;
Hoerrner, R. S. J . Org. Chem. 1992, 57, 441. (b) For an attempted
synthesis of 15-D2t-isoprostane: Taber, D. F.; Kanai, K. J . Org. Chem.
1998, 63, 6607. (c) see also ref 6d.
(4) (a) Roberts, L. J ., II; Moore, K. P.; Zackert, W. E.; Oates, J . A.;
Morrow, J . D. J . Biol. Chem. 1996, 271, 20617. (b) Morrow, J . D.; Awad,
J . A.; Kato, T.; Takahashi, K.; Bard, K. F.; Roberts, L. J ., II; Burk, R.
F. J . Clin. Invest. 1992, 90, 2502.
(5) After the work described here was completed, Durand et al.
reported the first total synthesis of the 15-F2t-isoprostane urinary
metabolite 2, as an inseparable mixture of diastereomers: Guy, A.;
Durand, T.; Roland, A.; Cormenier, E.; Rossi, J .-C. Tetrahedron Lett.
1998, 39, 6181.
(6) For synthetic routes to 15-F2t-isoprostane: (a) Corey, E. J .; Shih,
C.; Shih. N.-Y.; Shimoji, K. Tetrahedron Lett. 1984, 25, 5013. (b)
Hwang, S. W.; Adiyaman, M.; Khanapure, S.; Schio, L.; Rokach, J . J .
Am. Chem. Soc. 1994, 116, 10829. (c) Taber, D. F.; Herr, R. J .; Gleave,
D. M. J . Org. Chem. 1997, 62, 194. (d) Taber, D. F.; Kanai, K.
Tetrahedron 1998, 54, 11767.
(9) For a synthetic route to 5-F2t-isoprostane: Adiyaman, M.;
Lawson, J . A.; Hwang, S.-W.; Khanapure, S. P.; FitzGerald, G. A.;
Rokach, J . Tetrahedron Lett. 1996, 37, 4849.
(10) For a synthetic route to 8-F2t-isoprostane: Adiyaman, M.; Li,
H.; Lawason, J . A.; Hwang, S.-W.; Khanapure, S. P.; FitzGerald, G.
A.; Rokach, J . Tetrahedron Lett. 1997, 38, 3339.
(11) For a synthetic route to 12-F2t-isoprostane: Pudukulathan, Z.;
Manna, S.; Hwang, S.-W.; Khanapure, S. P.; Lawson, J . A.; FitzGerald,
G. A.; Rokach, J . J . Am. Chem. Soc. 1998, 120, 11953.
(12) Taber, D. F.; Gleave, D. M.; Herr, R. J .; Moody, K.; Hennessy,
M. J . J . Org. Chem. 1995, 60, 2283.
10.1021/jo9910738 CCC: $18.00 © 1999 American Chemical Society
Published on Web 09/21/1999

7984 J . Org. Chem., Vol. 64, No. 21, 1999
Taber and Kanai
Sch em e 1
Sch em e 3a
a
Reagents and conditions: (a) m-CPBA, CH2Cl2, -78 °C;
(MeO)3P, EtOH, -78 °C ∼ rt; (b) TBAF, THF; (c) LiOH‚H2O; 1%
Sch em e 2a
aqueous HCl.
inseparable mixture (ca. 3:1) in 97% yield. These were
distinguishable by comparing their 13C NMR spectra with
the spectra for the analogous ketones prepared in the
synthesis of the 15-F2t-isoprostanes.8 Thus, ketone 12a
had methines at δ 52.4, 51.0, and 50.2, comparable to δ
52.4, 51.4, and 50.3 for the previous ketone with cis side
chains, while ketone 12b had methines at δ 53.3, 53.0,
and 50.0, comparable to δ 53.1, 53.0, and 50.0 for the
previous ketone with trans side chains. We assumed that
the opening with thiophenol proceeded with inversion at
the reacting center, as we have previously observed.8
Reduction of the ketone mixture with sodium borohy-
dride produced alcohols 13 and 14 in 45% and 30% yields,
respectively, accompanied by the alcohol from the reduc-
tion of 12b (25%). Again, the relative configurations of
13 (1H NMR δ 4.39, quint, J ) 3.3 Hz, 1H; 3.98, m, 1H)
and 14 (1H NMR δ 4.64, dt, J ) 3.6 and 6.1 Hz, 1H; 4.25,
m, 1H) were assigned by comparison to the chemical
shifts of the H’s at C-9 and C-11 in the corresponding
diastereomers of the 15-isoprostane intermediates (1H
NMR δ 4.38, m, 1H; 3.99, m, 1H) and (1H NMR δ 4.64,
m, 1H; 4.24, m, 1H).8 The undesired â-alcohol 14 was
oxidized by Dess-Martin periodinane13 and then again
reduced with NaBH4 to give 13 and 14 in 58% and 39%
yields, respectively, over two steps.
With the relative configurations around the ring of 13
firmly established, we were prepared to embark on the
synthesis of 2,3-dinor-5,6-dihydro-15F2t -isoprostane (2)
(Scheme 3). Thus, oxidation and Mislow rearrangement14
of the alcohol 13 gave the allylic alcohol 15, which on
treatment with TBAF in THF gave the triol 16 in 93%
overall yield. Finally, the ester 16 was hydrolyzed with
LiOH in THF-H2O (1:1, v/v) to furnish the putative 15-
F2t-isoprostane urinary metabolite 2 in 92% yield.
Synthetic 2 was found15 to co-chromatograph with the
endogenously derived material on silica gel TLC (98:2
ethyl acetate/methanol) with an Rf ) 0.21 when deriva-
tized as the pentafluorobenzyl ester. Epimeric materials
are easily discerned with this solvent system; for in-
stance, the pentafluorobenzyl ester of PGF2R and of its
a
Reagents and conditions: (a) 5-bromovalerate, K2CO3,
n-Bu4NBr, toluene, 100 °C then 60 °C; (b) p-NBSA, DBU, CH2Cl2,
0 °C; (c) KHMDS, (E,E)-decadienal 6, TESCl, toluene, -78 °C (9:
10 ) 3.5:1); (d) TBAF, NH4Cl, THF, 0 °C; (e) TBDPS, imidazole,
4-DMAP, CH2Cl2; (f) Rh2(oct)4, CH2Cl2 (11:3 ) 1:3); (g) PhSH,
BF3‚OEt2, CH2Cl2, -78 °C then -45 °C; (h) NaBH4, MeOH, 0 °C
(13: 45%; 14: 30%); (i) Dess-Martin periodinane, CH2Cl2; (j)
NaBH4, MeOH, 0 °C (13: 58%; 14: 39%).
tanes.8 In particular, the oxygenated methine of 3 (13C δ
69.3, 1H δ 4.45, d, J ) 4.9 Hz, 1H) is almost exactly
congruent with the analogous 15-F2t-isoprostane precur-
(13) (a) Dess, D. B.; Martin, J . C. J . Org. Chem. 1983, 48, 4156. (b)
Ireland, R. E.; Liu, L. J . Org. Chem. 1993, 58, 2899.
(14) (a) Bickart, P.; Carson, F. W.; J acobus, J .; Miller, E. G.; Mislow,
K. J . Am. Chem. Soc. 1968, 90, 4869. (b) Tang, R.; Mislow, K. J . Am.
Chem. Soc. 1970, 92, 650.
(15) We thank Dr. J ason Morrow at Vanderbilt University for
making these comparisons.
sor (13C δ 69.6, H δ 4.41, d, 1H), while the oxygenated
1
methine of 11 (13C δ 67.9, H δ 4.59, dt, J ) 5.2, 7.9 Hz,
1
1H) is quite different.
Cyclopropane ring opening with thiophenol and
BF3‚OEt2 in CH2Cl2 gave the ketones 12a and 12b as an

Synthesis of 2,3-Dinor-5,6-dihydro-15F2t-isoprostane
J . Org. Chem., Vol. 64, No. 21, 1999 7985
958 cm-1; EI MS m/z (rel intensity) 184 (M+ - N2, 100), 142
(57), 138 (89), 110 (82); EI HRMS calcd for C10H16O3 184.1099,
found 184.1087.
8-epi diasteromer separated by 0.10 Rf units. In addition,
synthetic 2 perfectly co-chromatographed with the en-
dogenous metabolite on gas chromatography (DB1701
fused silica capillary column, methane as carrier gas,
temperature ramp 190-300 °C at 20 °C per minute,
retention time of 6 min) after derivatization to the
pentafluorobenzyl ester/tris trimethylsilyl ether deriva-
tive. Again, epimeric materials are easily discerned with
this system; the derivatives of PGF2R and of its 8-epi
diasteromer separated by a full 10 s.
Synthetic 2 was also analyzed15,16 by MS in both the
EI and negative ion CI modes. The synthetic material
had a molecular weight and fragmentation pattern
identical to the endogenously derived metabolite when
analyzed both as the PFB ester TMS ether derivative and
as the methyl ester TMS ether derivative.
TES-P r otected Ald ol 9 a n d F r ee Ald ol 10. To a stirred
solution of the diazo ketone 5 (1.92 g, 9.06 mmol) in toluene
(190 mL) at -78 °C was added dropwise a 0.33 M toluene
solution of KHMDS (30 mL, 9.96 mmol) over 15 min. After 5
min, a solution of (E,E)-decadienal 6 (1.65 g, 10.87 mmol) and
TESCl (1.82 mL, 10.87 mmol) in toluene (40 mL) was added.
Afetr an additional 15 min, the reaction mixture was parti-
tioned between EtOAc and, sequentially, saturated aqueous
NH4Cl and brine. The combined organic extract was dried (Na2-
SO4) and concentrated. The residue was chromatographed to
afford the TES-ether 9 (2.35 g, 54% yield) as a pale yellow oil:
1
TLC Rf (petroleum ether/MTBE ) 85/15) ) 0.55; H NMR δ
6.14 (1H, dd, J ) 10.5, 15.1 Hz), 5.98 (1H, dd, J ) 10.5, 15.1
Hz), 5.68 (1H, dt, J ) 6.9, 15.1 Hz), 5.53 (1H, dd, J ) 6.9, 15.1
Hz), 4.61 (1H, m), 4.12 (2H, q, J ) 7.1 Hz), 2.73 (1H, dd, J )
8.2, 13.5 Hz), 2.46 (1H, dd, J ) 4.6, 13.5 Hz), 2.28-2.43 (2H,
m), 2.31 (2H, t, J ) 7.3 Hz), 2.06 (2H, q, J ) 6.9 Hz), 1.67
(2H, quint, J ) 7.5 Hz), 1.50 (2H, quint, J ) 7.5 Hz), 1.38 (2H,
quint, J ) 7.1 Hz), 1.23-1.37 (4H, m), 1.25 (3H, t, J ) 7.1
Hz), 0.91 (9H, t, J ) 8.0 Hz), 0.88 (3H, t, J ) 7.1 Hz), 0.56
(6H, q, J ) 8.0 Hz); 13C NMR δ up 191.8, 173.3, 68.4, 60.3,
46.9, 33.8, 32.6, 31.4, 28.8, 26.6, 24.1, 22.5, 22.4, 4.8; down
135.7, 132.5, 130.5, 129.1, 71.1, 14.2, 14.0, 6.7; IR (film) 2955,
2875, 2070, 1736, 1631, 1459, 1368, 1239, 1181, 1071, 990, 744
cm-1; FAB MS m/z (rel intensity) 501 (M+ + Na, 48), 473 (50),
267 (100); FAB HRMS calcd for C26H46N2O4NaSi 501.3125,
found 501.3115. This was followed by the free aldol 10 (508
mg, 16% yield) as a colorless oil: TLC Rf (petroleum ether/
MTBE ) 85/15) ) 0.09; 1H NMR δ 6.25 (1H, dd, J ) 10.4,
15.3 Hz), 6.01 (1H, dd, J ) 10.4, 15.3 Hz), 5.72 (1H, dt, J )
7.0, 15.3 Hz), 5.58 (1H, dd, J ) 7.0, 15.3 Hz), 4.63 (1H, q, J )
6.0 Hz), 4.13 (2H, q, J ) 7.1 Hz), 3.30 (1H, br s), 2.67 (2H, br
s, J ) 6.0 Hz), 2.37 (2H, t, J ) 7.4 Hz), 2.33 (2H, t, J ) 7.4
Hz), 2.07 (2H, q, J ) 6.9 Hz), 1.67 (2H, quint, J ) 7.4 Hz),
15.2 (2H, quint, J ) 7.4 Hz), 13.8 (2H, quint, J ) 7.4 Hz), 1.24-
1.34 (4H, m), 1.26 (3H, t, J ) 7.1 Hz), 0.88 (3H, t, J ) 7.1 Hz);
13C NMR δ up 193.1, 173.3, 68.0, 60.4, 44.2, 33.7, 32.6, 31.3,
28.8, 26.5, 23.9, 22.5, 22.1; down 136.2, 131.2, 130.9, 129.1,
69.0, 14.2, 14.0; IR (film) 3437, 1928, 2858, 2074, 1732, 1621,
1462, 1372, 1183, 1076, 991, 875, 729 cm-1; EI MS m/z (rel
intensity) 336 (M+ - N2, 29), 290 (37), 191 (100), 110 (29); EI
HRMS calcd for C20H32O4 336.2301, found 336.2320.
Con clu sion s
We have confirmed the structure of the 15-F2t-isopros-
tane urinary metabolite 2, and have also developed a
practical synthetic route to this important product of
human metabolism. This racemic synthetic material (the
natural metabolite is racemic) will be used to develop an
immunoassay for the metabolite, thus allowing its quan-
tification for an integrated assessment of individual
oxidative stress.
Exp er im en ta l P r oced u r es17
Dik eton e 8. A mixture of benzoylacetone (7) (18.62 g, 114.8
mmol), K2CO3 (52.9 g, 382.6 mmol), and n-BuN4Br (308 mg,
1.0 mmol) in toluene (300 mL) was heated at 100 °C for 3 h.
The mixture was cooled at 60 °C for the addition of ethyl
5-bromovalerate (20 g, 95.7 mmol). The mixture was then
stirred at 60 °C for 48 h. The mixture was further cooled to
room temperature and partitioned between EtOAc and, se-
quentially, saturated aqueous NH4Cl and brine. The organic
extract was dried (Na2SO4) and concentrated. The residue was
chromatographed to afford the diketone 8 (18.8 g, 68% yield)
as a yellow oil; TLC Rf (petroleum ether/MTBE ) 6/4) ) 0.49;
1H NMR δ 7.97-8.00 (2H, m), 7.58-7.63 (1H, m), 7.43-7.51
(2H, m), 4.44 (1H, t, J ) 7.0 Hz), 4.10 (2H, q, J ) 7.1 Hz),
2.29 (2H, t, J ) 7.4 Hz), 2.13 (3H, s), 1.91-2.14 (2H, m), 1.23
(3H, t, J ) 7.1 Hz); 13C NMR δ up 204.1, 196.2, 173.3, 136.3,
60.2, 33.8, 28.5, 27.0, 24.6; down 133.7, 128.8, 128.6, 63.1, 27.8,
14.1; IR (film) 2938, 1731, 1681, 1596, 1448, 1359, 1182, 1096,
TBDP S-P r ot ect ed Ald ol 4. To a stirred solution of the
TES-ether 9 (140 mg, 0.30 mmol) in THF (3 mL) at 0 °C was
added NH4Cl (78 mg, 1.46 mmol) followed by a 1.0 M THF
solution of n-Bu4NF (0.32 mL, 0.32 mmol). After an additional
30 min, the reaction mixture was partitioned between EtOAc
and, sequentially, saturated aqueous NH4Cl and brine. The
organic extract was dried (Na2SO4) and concentrated. The
residue was chromatographed to afford the free aldol 10 (104
mg, 97% yield) as a pale yellow oil. To a stirred solution of the
free aldol 10 (1.85 g, 5.08 mmol) in CH2Cl2 (40 mL) at room
temperature were added imidazole (450 mg, 6.61 mmol),
4-DMAP (124 mg, 1.02 mmol), and tert-butylchlorodiphenyl-
silane (1.59 mL, 6.10 mmol). After 15 h at room temperature,
the reaction mixture was partitioned between CH2Cl2 and,
sequentially, saturated aqueous NH4Cl and brine. The organic
extract was dried (Na2SO4) and concentrated. The residue was
chromatographed to afford the tert-butyldiphenylsilyl ether 4
(2.9 g, 95% yield) as a pale yellow oil: TLC Rf (petroleum ether/
1031, 972, 773, 695 cm-1; CI MS m/z (rel intesity) 291 (M+
+
1, 100), 275 (33), 249 (39), 204 (11), 187 (26), 163 (15), 157
(11), 141 (12), 105 (41); CI HRMS calcd for C17H23O4 291.1596,
found 291.1596.
Dia zo Keton e 5. To a stirred solution of the diketone 8 (5
g, 17.24 mmol) in CH2Cl2 (250 mmol) at 0 °C was added DBU
(4.13 mL, 27.59 mmol). After 5 min, a solution of p-nitroben-
zenesulfonyl azide (5.9 g, 25.86 mmol) in CH2Cl2 (100 mL) was
added dropwise over 30 min. After an additional 1 h, the
reaction mixture was partitioned between CH2Cl2 and, se-
quentially, saturated aqueous NH4Cl and brine. The organic
extract was dried (Na2SO4) and concentrated. The residue was
chromatographed to afford the diazo ketone 5 (2.23 g, 61%
yield) as a pale yellow oil: TLC Rf (petroleum ether/MTBE )
6/4) ) 0.31; 1H NMR δ 4.13 (2H, q, J ) 7.1 Hz), 2.34 (4H, m),
2.24 (3H, s), 1.68 (2H, quint, J ) 7.6 Hz), 1.52 (2H, quint, J )
7.6 Hz), 1.26 (3H, t, J ) 7.1 Hz); 13C NMR δ up 191.0, 173.2,
66.9, 60.3, 33.7, 26.5, 23.9, 22.0; down 25.3, 14.1; IR (film) 2938,
2867, 2070, 1732, 1644, 1446, 1372, 1327, 1195, 1098, 1030,
1
MTBE ) 85/15) ) 0.36; H NMR δ 7.61-7.64 (4H, m), 7.31-
7.42 (6H, m), 5.65-5.80 (2H, m), 5.38-5.48 (2H, m), 4.67 (1H,
q, J ) 6.7 Hz), 4.11 (2H, q, J ) 7.1 Hz), 2.77 (1H, dd, J ) 7.4,
13.6 Hz), 2.50 (1H, dd, J ) 5.7, 13.7 Hz), 2.20-2.35 (4H, m),
1.99 (2H, q, J ) 7.1 Hz), 1.63 (2H, quint, J ) 7.5 Hz), 1.41-
1.49 (2H, m), 1.22-1.37 (6H, m), 1.24 (3H, t, J ) 7.1 Hz), 1.03
(9H, s), 0.88 (3H, t, J ) 7.1 Hz); 13C NMR δ up 191.3, 173.2,
133.8, 133.7, 68.1, 60.2, 46.4, 33.8, 32.5, 31.3, 28.8, 26.5, 24.0,
22.4, 22.2, 19.2; down 135.90, 135.85, 135.7, 131.4, 129.6, 129.6,
129.4, 129.0, 127.4, 127.3, 72.2, 26.9, 14.2, 14.0; IR (film) 2930,
2857, 2068, 1738, 1634, 1463, 1372, 1185, 1112, 1062, 989, 822,
703, 612 cm-1; FAB MS m/z (rel intensity) 625 (M+ + Na, 69),
(16) For full experimental details for the derivitization and analysis,
see: Morrow, J . D.; Roberts, L. J ., II. Methods Enzymol. 1999, 300, 3.
(17) For a summary of general experimental procedures, see: Taber,
D. F.; Meagley, R. P.; Doren, D. J . J . Org. Chem. 1996, 61, 5723.

7986 J . Org. Chem., Vol. 64, No. 21, 1999
Taber and Kanai
598 (46), 597 (100), 529 (45), 517 (21); FAB HRMS calcd for
129.9, 128.8, 127.8, 127.7, 127.0, 126.2, 72.5, 53.3, 53.0, 50.0,
26.9, 14.2, 13.9.
C
36H50N2O4NaS 625.3438.
Bicyclic Keton es 3 a n d 11. To a stirred solution of the
Alcoh ols 13 a n d 14. To a stirred solution of the inseparable
ketone epimers 12a and 12b (3:1) (695 mg, 1.02 mmol) in
MeOH (14 mL) at 0 °C was added NaBH4. After an additional
1 h, the mixture was partitioned between EtOAc and, seqen-
tially, 5% aqueous HCl, saturated aqueous NaHCO3, and
brine. The organic extract was dried (Na2SO4) and concen-
trated. The residue was chromatographed to afford the alcohol
from reduction of the trans ketone 12b (174 mg, 25% yield) as
a colorless oil; TLC Rf (petroleum ether/MTBE ) 8/2) ) 0.44.
This was oxidized by the Dess-Martin periodinane to give a
quantitiative yield of the ketone 12b, characterized above.
Further elution gave the â-alcohol 14 (208 mg, 30% yield) as
a colorless oil: TLC Rf (petroleum ether/MTBE ) 8/2) ) 0.37;
1H NMR δ 7.64-7.69 (4H, m), 7.35-7.44 (6H, m), 7.20-7.25
(5H, m), 5.25 (1H, dd, J ) 9.5, 15.1 Hz), 4.91 (1H, dt, J ) 6.9,
15.1 Hz), 4.64 (1H, dt, J ) 3.6, 6.1 Hz), 4.21-4.28 (1H, m),
4.12 (2H, q, J ) 7.1 Hz), 3.48 (1H, dd, J ) 5.2, 9.5 Hz), 2.37-
2.48 (3H, m), 2.30 (2H, t, J ) 7.5 Hz), 2.05 (1H, ddd, J ) 3.6,
6.3, 15.1 Hz), 1.85 (1H, ddd, J ) 2.8, 6.3, 15.0 Hz), 1.73-1.80
(2H, m), 1.58-1.68 (2H, m), 1.45-1.58 (2H, m), 1.31-1.38 (1H,
m), 1.25 (3H, t, J ) 7.1 Hz), 1.17-1.26 (4H, m), 1.07-1.14 (4H,
m), 1.05 (9H, s), 0.84 (3H, t, J ) 7.1 Hz); 13C NMR δ up 173.8,
134.5, 134.3, 134.0, 60.2, 45.3, 34.2, 32.0, 31.3, 29.0, 28.2, 25.34,
25.27, 22.5, 19.1; down 135.92, 135.86, 133.3, 131.3, 130.4,
129.59, 129.58, 128.6, 127.57, 127.56, 127.4, 76.2, 73.3, 55.1,
52.9, 45.9, 7.0, 14.2, 14.0; IR (film) 3463, 2929, 2856, 1735,
1460, 1427, 1374, 1186, 1112, 1066, 965, 822, 740, 702, 612
cm-1; FAB MS m/z (rel intensity) 709 (M+ + Na, 90), 687 (34),
679 (76), 673 (62), 671 (59), 643 (35), 629 (100), 627 (95); FAB
HRMS calcd for C42H58O4SiSNa 709.3723, found 709.3747.
This was followed by the R-alcohol 13 (313 mg, 45% yield) as
a colorless oil: TLC Rf (petroleum ether/MTBE ) 8/2) ) 0.28;
1H NMR δ 7.69-7.72 (4H, m), 7.35-7.43 (6H, m), 7.17-7.24
(5H, m), 5.16 (1H, dd, J ) 9.5, 15.1 Hz), 4.87 (1H, dt, J ) 6.9,
15.1 Hz), 4.39 (1H, quint, J ) 3.3 Hz), 4.11 (2H, q, J ) 7.1
Hz), 3.96-4.03 (1H, m), 3.42 (1H, dd, J ) 6.4, 9.5 Hz), 2.60
(1H, dt, J ) 3.7, 7.3 Hz), 2.28 (2H, t, J ) 7.5 Hz), 2.09-2.25
(1H, m), 2.13 (1H, dt, J ) 6.8, 14.7 Hz), 1.56-1.77 (6H, m),
1.24 (3H, t, J ) 7.1 Hz), 1.15-1.46 (m, 6H), 1.07 (9H, s), 1.01-
1.07 (4H, m), 0.83 (3H, t, J ) 7.1 Hz); 13C NMR δ up 173.7,
135.3, 134.3, 133.9, 60.2, 43.8, 34.2, 32.0, 31.2, 28.9, 28.3, 27.9,
25.1, 22.4, 19.1; down 136.00, 135.97, 133.2, 132.0, 129.6, 129.3,
128.5, 127.6, 127.0, 76.9, 76.2, 54.1, 52.4, 50.1, 27.0, 14.2, 14.0;
IR (film) 3441, 2929, 2856, 1732, 1634, 1472, 1373, 1257, 1186,
1111, 1026, 822, 740, 703, 612 cm-1; FAB MS m/z (rel intensity)
709 (M+ + Na, 100), 701 (17); FAB HRMS calcd for C42H58O4-
SiSNa 709.3723, found 709.3693.
Con ver sion of â-Alcoh ol 14 to r-Alcoh ol 13. To a stirred
solution of the â-alcohol 14 (88 mg, 0.13 mmol) in CH2Cl2 (4
mL) at room temperature was added Dess-Martin periodinane
(109 mg, 0.26 mmol). After an additional 30 min, the reaction
mixture was cooled in an ice bath. The resulting precipitate
was filtered and washed with Et2O. Evaporation of the
combined filtrate gave a residue that was chromatographed
to afford the ketone 12a (88 mg) as a colorless oil. To a stirred
solution of the ketone 12a (44 mg, 0.06 mmol) in MeOH (2
mL) at 0 °C was added NaBH4. After an additional 1 h, the
reaction mixture was partitioned between EtOAc and, sequen-
tially, 5% aqueous HCl, saturated aqueous NaHCO3, and
brine. The organic extract was dried (Na2SO4) and concen-
trated. The residue was chromatographed to afford the â-al-
cohol 14 (17.2 mg, 39% recovery). This was followed by the
R-alcohol 13 (25.6 mg, 58% yield, 97% based on 14 not
recovered).
Diol 15. To a stirred solution of the thioether 13 (142 mg,
0.21 mmol) in CH2Cl2 (2.5 mL) at -78 °C was added a solution
of m-CPBA (77 mg, 0.31 mmol) in CH2Cl2 (1.5 mL). The
mixture was stirred for 1 h, after which time a solution of
trimethyl phosphite (244 µL, 2.07 mmol) in EtOH (2 mL) was
added. The mixture was stirred at -78 °C for 5 min and then
warmed to room temperature. The reaction mixture was
partitioned between EtOAc and, sequentially, 5% aqueous
NaOH, saturated aqueous NH4Cl, and brine. The organic
diazo ketone 4 (1.5 g, 2.50 mmol) in CH2Cl2 (20 mL) at room
temperature was added a solution of Rh2(oct)4 (0.7 mg, 0.013
mmol) in CH2Cl2 (10 mL). After 30 min at room temperature,
the reaction mixture was concentrated. The residue was
chromatographed to afford the bicyclic ketone 3 (977 mg, 68%
yield) as a colorless oil: TLC Rf (benzene/MTBE ) 97/3) ) 0.50;
1H NMR δ 7.60-7.69 (4H, m), 7.35-7.47 (6H, m), 5.44 (1H,
dt, J ) 8.0, 15.3 Hz), 4.99 (1H, dd, J ) 8.0, 15.3 Hz), 4.45 (1H,
d, J ) 4.9 Hz), 4.11 (2H, q, J ) 7.1 Hz), 2.31 (2H, dt, J ) 2.6,
7.4 Hz), 2.22 (1H, dd, J ) 5.0, 18.6 Hz), 1.95-2.04 (5H, m),
1.65-1.73 (2H, m), 1.43-1.55 (3H, m), 1.19-1.35 (7H, m), 1.24
(3H, t, J ) 7.1 Hz), 1.05 (9H, s), 0.88 (3H, t, J ) 7.1 Hz); 13C
NMR δ up 212.7, 173.6, 133.7, 133.5, 60.1, 43.8, 42.7, 34.2,
32.5, 31.2, 28.9, 27.1, 14.8, 23.1, 22.4, 19.0; down 135.7, 135.6,
134.6, 129.8, 127.7, 124.4, 69.3, 40.2, 31.7, 26.8, 14.2, 14.0; IR
(film) 2930, 2857, 1732, 1463, 1428, 1372, 1302, 1245, 1161,
1112. 1062, 968, 906, 823, 703, 612 cm-1; EI MS m/z (rel
intensity) 574 (M+, 34), 517 (47), 292 (47), 246 (90), 213 (99),
199 (100), 120 (46); EI HRMS calcd for C36H50O4Si 574.3478,
found 574.3503. This was followed by the bicyclic ketone 11
(326 mg, 23%) as a colorless oil: TLC Rf (benzene/MTBE )
97/3) ) 0.48; 1H NMR δ 7.62-7.72 (4H, m), 7.36-7.46 (6H,
m), 5.67 (1H, dt, J ) 7.6 and 15.2 Hz), 5.06 (1H, dd, J ) 6.9
and 15.2 Hz), 4.59 (1H, dt, J ) 5.2 and 7.9 Hz), 4.08 (2H, q, J
) 7.1 Hz), 2.35 (1H, dd, J ) 4.0 and 8.1 Hz), 2.25 (2H, dd, J
) 3.6 and 8.4 Hz), 2.20 (2H, t, J ) 7.9 Hz), 2.07 (2H, q, J )
6.8 Hz), 1.70-1.80 (2H, m), 1.20-1.50 (11H, m), 1.21 (3H, t, J
) 7.1 Hz), 1.05 (9H, s), 0.90 (3H, t, J ) 7.1 Hz); 13C NMR δ up
210.7, 173.6, 133.9, 133.6, 60.1, 46.6, 42.2, 34.1, 32.6, 31.3, 29.0,
25.0, 23.7, 22.5, 19.0; down 135.8, 135.5, 134.6, 129.83, 129.80,
127.8, 127.7, 124.8, 67.9, 38.5, 30.3, 26.8, 14.2, 14.1; IR (film)
2930, 1727, 1463, 1428, 1365, 1245, 1154, 1111, 966, 824, 741,
703, 612 cm-1; EI MS m/z (rel intensity) 574 (M+, 5), 517 (37),
292 (57), 146 (98), 199 (100), 105 (43); EI HRMS calcd for
C
36H50O4Si 574.3478, found 574.3474.
Keton es 12a a n d 12b. To a stirred solution of the bicyclic
ketone 3 (620 mg, 1.08 mmol) and thiophenol (0.22 mL, 2.16
mmol) in CH2Cl2 (5.4 mL) at -78 °C was added BF3‚OEt2 (0.33
mL, 2.70 mmol). After 10 min, the mixture was warmed to
-45 °C. After an additional 2.5 h, the reaction mixture was
partitioned between CH2Cl2 and, sequentially, saturated aque-
ous NaHCO3 and brine. The organic extract was dried (Na2-
SO4) and concentrated. The residue was chromatographed to
afford an inseparable 3:1 mixture of the thioether 12a and its
6-epimer 12b (715 mg, 97% yield) as a colorless oil: TLC Rf
(petroleum ether/MTBE ) 97:3) ) 0.61; IR (film) 2930, 2857,
1739, 1463, 1427, 1372, 1175, 1112, 1053, 823, 740, 702, 612
cm-1; FAB MS m/z (rel intensity) 707 (M+ + Na, 100), 628 (6),
598 (9), 576 (25); FAB HRMS calcd for
C42H56O4NaSiS
707.3566, found 707.3557. Major isomer: 1H NMR δ 7.62-
7.67 (12/4H, m), 7.37-7.47 (18/4H, m), 7.17-7.26 (15/4H, m),
5.02-5.16 (6/4H, m), 4.60 (3/4H, d, J ) 5.8 Hz), 4.12 (6/4H, q,
J ) 7.1 Hz), 3.33 (3/4H, dd, J ) 3.6, 7.9 Hz), 2.76-2.81 (6/4H,
m), 2.51 (3/4H, dd, J ) 6.1, 19.6 Hz), 2.24-2.32 (6/4H, m),
1.87-1.96 (3/4H, m), 1.61-1.84 (12/4H, m), 1.41-1.57 (9/4H),
1.25 (9/4H, t, J ) 7.1 Hz), 1.16-1.26 (6/4H, m), 1.06-1.12 (12/
4H, m), 1.06 (27/4H, s), 0.82 (9/4H, t, J ) 7.1 Hz); 13C NMR δ
up 217.4, 173.6, 134.6, 133.43, 133.41, 60.2, 46.9, 34.1, 32.0,
31.2, 28.8, 28.0, 25.0, 24.3, 22.4, 19.0; down 135.8, 135.7, 133.3,
132.8, 129.89, 129.86, 128.6, 128.2, 127.8, 127.7, 127.3, 70.6,
52.4, 51.0, 50.2, 26.9, 14.2, 14.0. Minor isomer: 1H NMR δ
7.60-7.70 (4/4H, m), 7.35-7.48 (6/4H, m), 7.18-7.26 (5/4H,
m), 5.08 (1/4H, dd, J ) 9.6, 15.2 Hz), 4.96 (1/4H, dt, J ) 6.7,
15.2 Hz), 4.29 (1/4H, dt, J ) 4.7, 6.1 Hz), 4.13 (2/4H, q, J )
7.1 Hz), 3.59 (1/4H, dd, J ) 4.9, 9.6 Hz), 2.44 (1/4H, dt, J )
4.7, 6.7 Hz), 2.33 (1/4H, m), 2.31 (2/4H, t, J ) 7.6 Hz), 2.16-
2.23 (2/4H, m), 1.74-1.82 (4/4H, m), 1.65 (2/4H, quint, J )
6.7 Hz), 1.44-1.53 (1/4H, m), 1.30-1.39 (1/4H, m), 1.25 (3/
4H, t, J ) 7.1 Hz), 1.15-1.22 (3/4H, m), 0.98-1.12 (4/4H, m),
1.07 (9/4H, s), 0.82 (3/4H, t, J ) 7.1 Hz); 13C NMR δ up 217.8,
173.6, 134.7, 133.4, 133.2, 60.2, 47.3, 34.2, 32.0, 31.1, 30.3, 28.7,
26.4, 25.0, 22.4, 19.1; down 135.9, 135.8, 135.3, 132.6, 130.0,

Synthesis of 2,3-Dinor-5,6-dihydro-15F2t-isoprostane
J . Org. Chem., Vol. 64, No. 21, 1999 7987
extract was dried (Na2SO4) and concentrated. The residue was
chromatographed to afford the diol 15 (116.5 mg, 95% yield)
as a colorless oil: TLC Rf (petroleum ether/MTBE ) 55/45) )
0.33; 1H NMR δ 7.62-7.67 (4H, m), 7.35-7.45 (6H, m), 5.04-
5.15 (2H, m), 4.12 (2H, q, J ) 7.1 Hz), 3.99 (1H, quint, J ) 2.6
Hz), 3.80-3.88 (2H, m), 2.66 (1H, m), 2.27 (2H, t, J ) 7.4 Hz),
2.16-2.23 (2H, m), 2.02 (1H, br s), 1.63-1.77 (2H, m), 1.56-
1.62 (2H, m), 1.25 (3H, t, J ) 7.1 Hz), 1.19-1.44 (12H, m),
1.07 (9H, s), 0.87 (3H, t, J ) 7.1 Hz); 13C NMR δ up 173.8,
134.0, 133.8, 60.2, 43.1, 37.0, 34.2, 31.7, 29.1, 27.7, 25.0, 22.6,
19.0; down 135.9, 135.8, 129.72, 129.68, 128.5, 127.6, 78.8, 77.6,
72.7, 53.9, 50.2, 27.0, 14.2, 14.0; IR (film) 3417, 2930, 2857,
1732, 1463, 1428, 1373, 1258, 1188, 1112, 1066, 970, 823, 741,
703, 613 cm-1; EI MS m/z (rel intensity) 594 (M+, 1), 520 (33),
519 (68), 303 (51), 199 (100), 135 (26); EI HRMS calcd for
C36H54O5Si 584.3741, found 594.3787.
184 (10), 133 (42), FAB HRMS calcd for C20H36O5Na 379.2460,
found 379.2445.
2,3-Din or -5,6-d ih yd r o-15F2t -isop r osta n e (2). To a stirred
solution of the ester 16 (20 mg, 0.056 mmol) in THF-H2O (1:
1, 1.5 mL) at room temperature was added LiOH‚H2O (24 mg,
0.56 mmol). After 3 h at room temperature, the reaction
mixture at 0 °C was acidified to pH ) 4 by adding 1% aqueous
HCl (2.2 mL). After the addition of solid NaCl (1 g), the mixture
was extracted with CHCl3-MeOH (95:5, v/v). The organic
extract was dried (Na2SO4) and concentrated. The residue was
chromatographed to afford 2 (17 mg, 92% yield) as a white
solid: TLC Rf (EtOAc/MeOH/AcOH ) 90/10/0.1) ) 0.13; 1H
NMR (CD3OD) δ 5.49 (1H, dd, J ) 6.6, 15.3 Hz), 5.39 (1H, dd,
J ) 9.5, 15.3 Hz), 3.98 (1H, q, J ) 6.6 Hz), 3.91 (1H, dt, J )
4.2, 6.6 Hz), 3.82 (1H, dt, J ) 6.1, 7.6 Hz), 2.60-2.66 (1H, m),
2.46 (1H, m), 2.28 (2H, t, J ) 7.4 Hz), 1.98-2.05 (1H, m), 1.27-
1.60 (15 H, m), 0.90 (3H, t, J ) 6.8 Hz); 13C NMR (CD3OD) δ
up 43.6, 38.4, 33.0, 29.6, 28.7, 26.4, 26.3, 23.7; down 136.8,
130.5, 76.5, 73.8, 54.2, 50.2, 14.4; IR (film) 353, 2929, 2858,
1704, 1462, 1415, 1344, 1260, 1072, 1018, 974, 797 cm-1; FAB
MS m/z (rel intensity) 351 (M+ + Na, 10), 311 (8), 293 (100),
275 (88), 249 (42), 138 (29); FAB HRMS calcd for C18H32O5Na
351.2147, found 351.2145.
Tr iol 16. To a stirred solution of the silyl ether 15 (192 mg,
0.32 mmol) in THF (3 mL) at room temperature was added a
1 M THF solution of n-Bu4NF (2.59 mL, 2.59 mmol). After 12
h at room temperature, the reaction mixture was partitioned
between EtOAc and, sequentially, saturated aqueous NH4Cl
and brine. The organic extract was dried (Na2SO4) and
concentrated. The residue was chromatographed to afford the
triol 16 (113 mg, 98% yield) as a colorless oil: TLC Rf (EtOAc/
1
MeOH ) 95/5) ) 0.5; H NMR δ 5.53 (1H, dd, J ) 7.3, 15.2
Ack n ow led gm en t. We thank Oxford Biomedical
Research, Inc., for support of this work. We also thank
L. J ackson Roberts II and J ason Morrow at Vanderbilt
University for collaborative efforts.
Hz), 5.38 (1H, dd, J ) 9.7, 15.2 Hz), 4.12 (2H, 2H, q, J ) 7.1
Hz), 4.02 (1H, q, J ) 6.8 Hz), 3.89-3.97 (2H, m), 3.35-3.85
(1H, br), 2.75-3.30 (2H, br), 2.69-2.74 (1H, m), 2.42 (1H,
quint, J ) 7.2 Hz), 2.28 (2H, t, J ) 7.4 Hz), 2.02-2.09 (1H,
m), 1.52-1.64 (4H, m), 1.23-1.48 (11H, m), 1.25 (3H, t, J )
7.1 Hz), 0.89 (3H, t, J ) 7.1 Hz); 13C NMR δ up 174.0, 60.3,
42.4, 37.1, 34.1, 31.7, 28.7, 27.5, 25.2, 24.8, 22.6; down 136.1,
129.6, 76.4, 73.1, 53.4, 49.9, 14.2, 14.0; IR (film) 3226. 2925,
2855, 1738, 1463, 1422, 1379, 1305, 1185, 1092, 1035, 976, 730
cm-1; FAB MS m/z (rel intensity) 379 (M+ + Na, 15), 242 (100),
Su p p or t in g In for m a t ion Ava ila b le: 1H and 13C NMR
spectra for all new compounds. This material is available free
of charge via the Internet at http://pubs.acs.org.
J O9910738