Synthesis of sphingomyelin carbon analogues as sphingomyelinase inhibitors

Article abstract of DOI:10.1021/jo025529o

The highly efficient and stereocontrolled syntheses of sphingomyelin carbon analogues 1 and 2 were achieved by effectively utilizing Hofmann rearrangement of enantiomerically pure β-hydroxyamide 7, which was prepared by an asymmetric hydrogenation of α-acyl-γ-butyrolactone 9 and ring opening with NH₃. Intermediary isocyanate 6 was selectively trapped with the vicinal hydroxy group in an intramolecular fashion to produce an oxazolidinone derivative, 5. In the synthesis of a quite polar compound such as 1, a convenient one-pot procedure of the introduction of a benzyloxycarbonyl group into the hydroxy group resulting from the oxazolidinone ring opening is another key point, because, in addition to the efficiency, this protecting group was easily removable by a simple procedure and workup at the final step. Both synthesized compounds 1 and 2 showed moderate inhibitory activity toward sphingomyelinase from B. cereus.

Full text of DOI:10.1021/jo025529o

Syn th esis of Sp h in gom yelin Ca r bon An a logu es a s
Sp h in gom yelin a se In h ibitor s
Toshikazu Hakogi,† Yoshiko Monden,† Misako Taichi,† Seiji Iwama,† Shinobu Fujii,‡
Kiyoshi Ikeda,‡ and Shigeo Katsumura*,†
School of Science, Kwansei Gakuin University, Gakuen, Sanda, Hyogo 669-1337, J apan, and
Department of Biochemistry, Osaka University of Pharmaceutical Sciences, Nasahara, Takatsuki,
Osaka 569-1041, J apan
katsumura@ksc.kwansei.ac.jp
Received J anuary 17, 2002
The highly efficient and stereocontrolled syntheses of sphingomyelin carbon analogues 1 and 2
were achieved by effectively utilizing Hofmann rearrangement of enantiomerically pure â-hy-
droxyamide 7, which was prepared by an asymmetric hydrogenation of R-acyl-γ-butyrolactone 9
and ring opening with NH3. Intermediary isocyanate 6 was selectively trapped with the vicinal
hydroxy group in an intramolecular fashion to produce an oxazolidinone derivative, 5. In the
synthesis of a quite polar compound such as 1, a convenient one-pot procedure of the introduction
of a benzyloxycarbonyl group into the hydroxy group resulting from the oxazolidinone ring opening
is another key point, because, in addition to the efficiency, this protecting group was easily removable
by a simple procedure and workup at the final step. Both synthesized compounds 1 and 2 showed
moderate inhibitory activity toward sphingomyelinase from B. cereus.
In tr od u ction
initiated by hydrolysis of sphingomyelin by SMase, has
been well recognized, none of the tertiary-dimensional
structures of these important enzymes have been deter-
mined, and their hydrolytic mechanism has not been
well-defined. It is, therefore, a very attractive challenge
to reveal the catalytic mechanism of action of this
important enzyme. Strong and selective sphingomyelin-
ase inhibitors would contribute to a better understand-
ing both of the roles of these enzymes and of ceramide
in signal transduction. Some SMase inhibitors have
recently been reported (Figure 2).4 Among them, scypho-
statin, natural product, has been paid much attention
as a powerful inhibitor of N-SMase.4b,c,f Most recently,
compound Z was designed and synthesized as the first
selective irreversible inhibitor of N-SMase.4a Meanwhile,
Bittman’s group reported the preparation of sphingo-
myelin analogues modified at the C-1 and C-3 positions
of the sphingosine backbone, and 3-O-methylsphingo-
myelin showed moderate inhibitory activity toward N-
SMase.5 To elucidate the detailed catalytic mechanism
of SMase, the development of the methods for supplying
the sphingomyelin analogues, which competitively act at
the catalytic site and strongly inhibit the hydrolytic
ability of the enzyme, have strongly been desired.
Sphingolipids have been known as lipid second mes-
sengers in mammalian cells and cell membranes, and a
great deal of attention has been devoted to the studies
of the biological process regulated by sphingolipids.1 Now,
it has been well accepted that the sphingolipids play
key roles in the cellular signal transmission pathway.
Sphingomyelin, which is one of the key sphingolipids,
is a ubiquitous constituent in animal tissue and has
been known to occur in virtually every cell and in cell
membranes. Ceramide and phosphoryl choline, which are
the primary catabolites of sphingomyelin, are generated
in the so-called sphingomyelin cycle through the action
of either lysosomal acid sphingomyelinase (A-SMase) or
membrane-bound neutral sphingomyelinase (N-SMase);
ceramide is believed to display key roles as a signal
transduction factor in cell differentiation and in pro-
grammed cell death (apoptosis) derivation.2 Hydrolysis
of the N-acyl group of ceramide with ceramidase produces
sphingosine. This secondary catabolite of sphingomyelin
strongly inhibits protein kinase C (Figure 1).3 Although
the significance of the sphingomyelin pathway, which is
* To whom correspondence should be addressed. Phone: 81-795-
65-8314. Fax: 81-795-65-9077.
(4) (a) Arenz, C. Giannis, A. Angew. Chem., Int. Ed. 2000, 39, 1440.
(b) Nara, F.; Tanaka, M.; Hosoya, T.; Suzuki-Konagai, K.; Ogita, T. J .
Antibiot. 1999, 52, 525. (c) Nara, F.; Tanaka, M.; Masuda-Inoue, S.;
Yamasato, Y.; Doi-Yoshioka, H.; Suzuki-Konagai, K.; Kumakura, S.;
Ogita, T. J . Antibiot. 1999, 52, 531. (d) Uchida, R.; Tomoda, H.; Dong,
Y.; Omura, S. J . Antibiot. 1999, 52, 572. (e) Tanaka, M.; Nara, F.;
Yamasato, Y.; Masuda-Inoue, S.; Doi-Yoshioka, H.; Kumakura, S.;
Enokita, R.; Ogita, T. J . Antibiot. 1999, 52, 670. (f) Tanaka, M.; Nara,
F.; Suzuki-Konagai, K.; Hosoya, T.; Ogita, T. J . Am. Chem. Soc. 1997,
119, 7871.
† Kwansei Gakuin University.
‡ Osaka University of Pharmaceutical Sciences.
(1) For recent reviews on signal transduction mediated by sphingo-
lipids, see: (a) Hannun, Y. A.; Luberto, C.; Argraves, K. M. Biochem-
istry 2001, 40, 4893. (b) Kolter, T.; Sandhoff, K. Angew. Chem., Int.
Ed. 1999, 38, 1532 and references therein.
(2) (a) Sawai, H.; Hannun, Y. A. Chem. Phys. Lipids 1999, 102, 141.
(b) Tilly, J . L.; Kolesnick, R. N. Chem. Phys. Lipids 1999, 102, 149. (c)
Kronke, M. Chem. Phys. Lipids 1999, 102, 157. (d) Hannun, Y. A.
Science 1996, 274, 1855 and references therein.
(5) Lister, M. D.; Ruan, Z.-s.; Bittman, R. Biochim. Biophys. Acta
1995, 1256, 25.
(3) Vankar, Y. D.; Schmidt, R. R. Chem. Soc. Rev. 2000, 29, 201.
10.1021/jo025529o CCC: $22.00 © 2002 American Chemical Society
Published on Web 05/31/2002
J . Org. Chem. 2002, 67, 4839-4846
4839

Hakogi et al.
F IGURE 1. Sphingomyelin metabolism.
Martin and co-workers reported the phospholipase C
(PLC) substrate analogues, in which the hydrolyzed
phosphonate oxygen was replaced by methylene, difluoro-
methylene, NH, and S groups, and reported that some
of them strongly inhibited the hydrolytic ability of PLC.11
We then designed the substrate analogues 1 and 2 as
inhibitor candidates on the basis of both our previous
results obtained on sphingomyelin analogues and the
results reported by Martin’s group.11 In analogues 1 and
2, one of the oxygen atoms of the phosphoester, at which
sphingomyelin is hydrolyzed by the enzyme, is replaced
by either a methylene or ethylene group, and the relative
configuration of the asymmetric centers is the (D)-erythro
(3S,4R) form. In addition, the double bond in the back-
bone skeleton is saturated (Figure 3). In this paper, we
describe in detail the highly efficient stereocontrolled
syntheses of the short chain substrate methylene ana-
logue 112 and ethylene analogue 2 and their inhibitory
F IGURE 2. Reported inhibitors of sphingomyelinase.
In a number of synthetic efforts for these biologically
significant and enantiomerically pure sphingolipids,6 we
had also established a method for the stereocontrolled
synthesis of sphingomyelin and provided all stereoiso-
mers of the short-chain monodispersed sphingomyelin
analogues from the chiral oxazolidinone synthon,7 which
had been easily prepared from enantiomerically pure
glycidol.8 We then clarified that the initial velocities of
the hydrolysis of (D)-erythro derivatives catalyzed by B.
cereus SMase are more than 10 times faster than those
of the (D)-threo isomers and that the double bond in the
backbone skeleton would not be essential for the hydroly-
sis by this enzyme. Meanwhile, phosphonate analogues,
in which the labile P-O-C bond is displaced by a stable
P-CH2-C bond, have been explored as phosphate sur-
rogates9 and are often utilized as useful bioorganic tools;10
13
activities toward B. cereus SMase.
Resu lts a n d Discu ssion
Our plan to synthesize these kinds of substrate ana-
logues substituted by carbon groups is shown in Scheme
1. The sphingomyelin carbon analogues 1 and 2 might
be, respectively, derived from phosphonate ceramide
analogues 3 and 4 by hydrolysis of the phosphonyl ester
followed by introduction of a choline group. Both cera-
mide phosphonate analogues might be synthesized from
the 2-substituted oxazolidinone 5 by introduction of a
phosphoryl group and then oxazolidinone ring opening.
To achieve the efficient synthesis of 1, therefore, a
convenient method for the preparation of erythro amino
alcohol derivative 5 was required. The Hofmann rear-
(6) Synthesis of sphingomyelin; see review: (a) Weis, A. L. Chem.
Phys. Lipids 1999, 102, 3. (b) Koskinen, P. M.; Koskinen, A. M. P.
Synthesis 1998, 1075. (c) Byun, H.-S.; Erukulla, R. K.; Bittman, R. J .
Org. Chem. 1994, 59, 6495. (d) Kratzer, B.; Mayer, T. G.; Schmidt, R.
R. Tetrahedron Lett. 1993, 34, 6881. (e) Dong, Z.; Butcher, J . A.,
J r. Tetrahedron Lett. 1991, 32, 5291. (f) Bruzik, K. S. J . Chem. Soc.,
Perkin Trans. 1 1988, 423. Recent syntheses of enantiomerically pure
sphingosine and ceramide, see: (g) He, L.; Byun, H.-S.; Bittman, R. J .
Org. Chem. 2000, 65, 7618. (h) He, L.; Byun, H.-S.; Bittman, R. J . Org.
Chem. 2000, 65, 7627. (i) Corey E. J .; Choi, S. Tetrahedron Lett. 2000,
41, 2765. Synthesis of sphingolipid derivatives, see: (j) Triola, G.;
Febrias, G.; Llebaria, A. Angew. Chem., Int. Ed. 2001, 40, 1960. (k)
Chun, J .; He, L.; Byun, H.-S.; Bittman, R. J . Org. Chem. 2000, 65,
7634. (l) He, L.; Byun, H.-S.; Smit, J .; Wilschut, J .; Bittman, R. J . Am.
Chem. Soc. 1999, 121, 3897. (m) Dondoni, A.; Perrone, D.; Turturici,
E. J . Org. Chem. 1999, 64, 5557. (p) Byun, H.-S.; Sadlofsky, J . A.;
Bittman, R. J . Org. Chem. 1998, 63, 2560.
(10) For recent examples, see: (a) Berkowitz, D. B.; Bose, M.;
Pfannenstiel, T. J .; Doukov, T. J . Org. Chem. 2000, 65, 4498. (b)
Viera de Almeida, M.; Cleophax, J .; GateauOlesker, A.; Prestat, G.;
Dubreuil, D.; Gero, S. D. Tetrahedron 1999, 55, 12997. (c) Billault, I.;
Vasella, A. Helv. Chim. Acta 1999, 82, 1137. (d) Kitanuma, H.;
Shimazaki, K.; Takahashi, N.; Takahashi, K.; Niihata, S.; Aoki, Y.;
Hamada, K.; Matsushita, H.; Nishi, Y. Tetrahedron 1999, 55, 25559.
(e) Sachpatzidis, A.; Dealwins, C.; Lubetsky, J .; Liang, P. H.; Anderson,
K. S.; Lolis, E. Biochemistry 1999, 38, 12665. (f) Arni, R. K.; Watanabe,
L.; Ward, R. J .; Kreiman, R. J .; Kumar, K.; Walz, F. G., J r. Biochemistry
1999, 38, 2452.
(11) (a) Martin, S. F.; J osey, J . A.; Wong, Y. L.; Dean, D. W. J . Org.
Chem. 1994, 59, 4805. (b) Martin, S. F.; Wong, Y. L.; Wagman, A. S.
J . Org. Chem. 1994, 59, 4821.
(7) Murakami, M.; Iwama, S.; Fujii, S.; Ikeda, K.; Katsumura, S.
Bioorg. Med. Chem. Lett. 1997, 7, 1725.
(8) Katsumura, S.; Yamamoto, N.; Morita, M.; Han; Q. Tetra-
hedron: Asymmetry 1994, 5, 161.
(9) For reviews on phosphonates, see: (a) Wiemer, D. F. Tetrahedron
1997, 53, 16609. (b) Engel, R. Chem. Rev. 1977, 77, 349.
(12) Hakogi, T.; Monden, Y.; Iwama, S.; Katsumura, S. Org. Lett.
2000, 2, 2627.
(13) During our study, a CF2 analogue having a phenyl derivative
in the backbone skeleton was reported: Yokomatsu, T.; Takeuchi, H.;
Akiyama, T.; Shibuya, S.; Kominato, T.; Soeda, S.; Shimeno, H. Bioorg.
Med. Chem. Lett. 2001, 11, 1277.
4840 J . Org. Chem., Vol. 67, No. 14, 2002

Synthesis of Sphingomyelin Carbon Analogues
SCHEME 2. Syn th esis of 2-Su bstitu ted
Oxa zolid in on ea
F IGURE 3. Designed inhibitors of sphingomyelinase.
SCHEME 1. Retr osyn th esis
a
Reagents and conditions: (i) H2, cat. (R)-BINAP-RuCl2, CH2Cl2,
60 °C, 100 atm, 10 days, 99% de; (ii) concentrated NH3, DME; (iii)
AgOAc, NBS, DMF, 77% for 2 steps.
CH2Cl2 under 100 atm pressure of hydrogen at 60 °C for
10 days according to the literature.16 The diastereoselec-
tivity of 8 was determined to be 98% de by 1H NMR, and
the enantioselectivity was determined to be 95% ee by
HPLC analysis using a chiral column after benzylation
of the secondary hydroxyl group under neutral condi-
tions.17 With enantiomerically pure alcohol 8 in hand, our
attention turned to construction of the amino alcohol
equivalent. Alcohol 8 was then treated with a large excess
of aqueous NH3 to yield amide 7. The Hofmann rear-
rangement of 7 by treatment with silver acetate and NBS
in DMF followed by intramolecular cyclization success-
fully proceeded and produced substituted oxazolidinone
5, which was easily purified by recrystallization, in 77%
yield from alcohol 8 (Scheme 2). The vicinal coupling
1
constant of the H NMR of the oxazolidinone ring thus
generated was J 4,5 ) 7.8 Hz and was compatible with a
cis-orientation of the two substituents.18 The results
obviously meant that the intermediary isocyanate result-
ing from the Hofmann rearrangement was selectively
trapped with the secondary hydroxyl group spontane-
ously to produce the five-membered oxazolidinone ring
with retention of the stereochemistry.19 Thus, the desired
protected amino alcohol 5 was efficiently synthesized.
The next subject was introduction of a phosphoryl
group and then a choline group. After bromination of the
primary alcohol in 5 with carbon tetrabromide and
triphenylphosphine, the phospholyl group was introduced
by the Arbuzov reaction20 with triethyl phosphite under
reflux. The reaction mixture was directly evaporated and
was reacted with tert-butoxycarbonyl anhydride to pro-
duce quantitatively N-tert-butoxycarbonyl derivative 12.
Considering the quite strong polarity of the molecule
after introduction of a choline group at the phosphoryl
group, the protecting group of the secondary hydroxyl
rangement followed by the intramolecular oxazolidinone
ring formation strategy was very attractive for this.14
Thus, stereospecific oxazolidinone formation resulting
from an intramolecular trap with the vicinal hydroxy
group of reactive isocyanate 6, which would be produced
by the Hofmann rearrangement of an amide such as 7,
would be a very preferable strategy for the synthesis of
5. Amide 7 would be easily prepared from enantiomeri-
cally pure γ-butyrolactone derivative 8, which possesses
a hydroxyalkyl group at the R-position.
We then started the synthesis with reduction of R-acyl-
γ-butyrolactone 9 by the method of Noyori’s asymmetric
hydrogenation.15 Thus, hydrogenation of â-ketoester 9
quantitatively yielded the corresponding alcohol, 8, even
if the reaction is on more than a 20-gram scale, in the
presence of a catalytic amount of (R)-BINAP-RuCl2 in
(16) Nishizawa, M.; Garcia, D. M.; Minagawa, R.; Noguchi, Y.;
Imagawa, H.; Yamada, H.; Watanabe, R.; Yoo, Y. C.; Azuma, I. Synlett
1996, 452.
(17) Hatakeyama, S.; Mori, H.; Kitano, K.; Yamada, H.; Nishizawa,
M. Tetrahedron Lett. 1994, 35, 4367.
(14) (a) Murakami, M.; Hinoue, K.; Nakagawa, K.; Monden, Y.;
Furukawa, Y.; Katsumura, S. Heterocycles 2001, 54, 77. (b) Simons,
S. S., J r. J . Org. Chem. 1973, 38, 414.
(15) (a) Noyori, R.; Ikeda, T.; Ohkuma, T.; Widhalm, M.; Kitamura,
M.; Takaya, H.; Akutagawa, S.; Sayo, N.; Saito, T.; Taketomi, T.;
Kumobayashi, H. J . Am. Chem. Soc. 1989, 111, 9134. (b) Kitamura,
M.; Tokunaga, M.; Ohkuma, T.; Noyori, R. Org. Synth. 1993, 71, 1.
(18) Sakaitani, M.; Ohfune, Y. J . Am. Chem. Soc. 1990, 112, 1150.
(19) Recently, the Curtius rearrangement for the similar oxazoli-
dinone formation was reported: (a) Pais, G. C. G.; Maier, M. E. J . Org.
Chem. 1999, 64, 4551. (b) Ghosh, A. K.; Hussain, K. A.; Fidanze, S. J .
Org. Chem. 1997, 62, 6080. (c) Ghosh, A. K.; Liu, W. J . Org. Chem.
1996, 61, 6175.
(20) Arbuzov, B. A. Pure Appl. Chem. 1964, 9, 307.
J . Org. Chem, Vol. 67, No. 14, 2002 4841

Hakogi et al.
SCHEME 3. Syn th esis of th e Meth ylen e An a logu ea
SCHEME 4. Syn th esis of th e Eth ylen e An a logu ea
a
Reagents and conditions: (i) 16, n-BuLi, THF, -78 °C, then
Boc2O, 63%; (ii) BnOH, Cs2CO3, THF, 57%; (iii) (a) TFA, CH2Cl2,
0 °C; (b) aqueous 1 N NaOH, CHCl3; (c) C5H11COCl, 79%; (iv)
TMSBr, CH2Cl2, then MeOH; (v) choline chloride, CCl3CN, pyr.,
50 °C, 17% for 2 steps; (vi) H2, Pd-C, MeOH, 89%.
a
Reagents and conditions: (i) CBr4, PPh3, CH2Cl2, 0 °C; (ii)
(EtO)3P, reflux; (iii) Boc2O, DMAP, Et3N, DMF, 0 °C, quant. for 3
steps; (iv) BnOH, Cs2CO3, THF, 81%; (v) (a) TFA, CH2Cl2, 0 °C;
(b) aqueous 1 N NaOH, CHCl3; (c) C5H11COCl, 85%; (vi) TMSBr,
CH2Cl2, then MeOH; (vii) choline chloride, CCl3CN, pyr., 50 °C,
17% for 2 steps; (viii) H2, Pd-C, MeOH, 77%.
to react bromide 10 with a lithium anion of methyl-
dimethylphosphonate, 15. Unfortunately, the desired
product was not obtained by the usual extraction proce-
dures because of the strong polarity of the products. We
then tried to introduce a tert-butoxycarbonyl group into
the oxazolidinone nitrogen. Thus, the reaction mixture
of the lithium anion of 15 with bromide 10 was succes-
sively treated with di-tert-butyl dicarbonate to produce
the desired 16 in 63% yield. The convenient one-pot
procedure was again tried for the oxazolidinone ring
opening followed by the protection of the alcohol gener-
ated with a benzyloxycarbonyl group. Thus, compound
16 was treated with cesium carbonate in the presence of
benzyl alcohol in THF to successfully produce 17 in 57%
yield. The tert-butoxycarbonyl group was removed with
a trifluoroacetic acid treatment, and then, a hexanoyl
group was introduced into the resulting amine by treat-
ment with sodium hydroxide and hexanoyl chloride in
chloroform to give N-acyl derivative 4 in 79% yield.
Hydrolysis of the phosphonate ester of 4 by treatment
with trimethylsilyl bromide and then methanol gave the
corresponding acid, which was reacted without any
purification with choline chloride and trichloroacetoni-
trile in pyridine at 50 °C for 2 days. The desired choline
derivative, 18, was obtained in 16% yield after purifica-
tion by reversed phase HPLC. Finally, the benzyloxy-
carbonyl group of the secondary hydroxyl group was
removed by hydrogenation over Pd-C, and the synthesis
of (3S,4R) sphingomyelin ethylene analogue 2 was
achieved (Scheme 4). Thus, effective and stereocontrolled
methods of providing the substrate carbon analogues 1
and 2 were established.
group resulting from the oxazolidinone ring opening
should be selected. It should be removed by hydrogenoly-
sis, which workup requires only simple filtration. For-
tunately, reaction of 12 with cesium carbonate in the
presence of benzyl alcohol in THF successfully proceeded,
and benzyl carbonate was introduced to the secondary
alcohol resulting from the opening of the oxazolidinone
ring, which was activated with the tert-butoxycarbonyl
group.21 After removing the tert-butoxycarbonyl group of
13 by an acid treatment, the introduction of the acyl
group at the resulting amino group produced the desired
O-benzyloxycarbonyl-N-acylphosphonate 3 in 71% yield
in two steps. The final part of the synthesis of 1 was the
introduction of a choline group. Treatment of 3 with
bromotrimethylsilane in CH2Cl2 produced the corre-
sponding silylester, which was continuously stirred in
methanol to complete the hydrolysis22 and was followed
by the reaction with choline chloride and trichloroaceto-
nitrile in pyridine23 without any purification. The desired
choline derivative, 14, was obtained in 16% yield after
purification by reverse HPLC. The major byproduct of
this reaction was the vinyl ester resulting from the
elimination of trimethylamine. Finally, the benzyloxy-
carbonyl group of the secondary hydroxyl group was
removed by hydrogenation over Pd-C, and the synthesis
of (3S,4R) sphingomyelin methylene analogue 1 was thus
achieved (Scheme 3).
Next, our interest is the synthesis of ethylene analogue
2. To construct the backbone skeleton of 2, we planned
We tested the ability of compounds 1 and 2 to inhibit
SMase from B. cereus. Enzyme activity was measured
three times at 37 °C and ionic strength 0.2 with a buffer
of 50 mM Tris-HCl buffer (pH 7.5) in the presence of 10
mM MgCl2. The concentrations of SMase and 2-hexade-
(21) Ishizuka, T.; Kunieda, T. Tetrahedron Lett. 1987, 28, 4185.
(22) (a) Pergament, I.; Srebnik, M. Tetrahedron Lett. 1999, 40, 3895.
(b) McKenna, C. E.; Schmidhauser, J . J . Chem. Soc., Chem. Commun.
1979, 739.
(23) Thomas, B. N.; Corcoran, R. C.; Cotant, C. L.; Lindemann, C.
M.; Kirsch, J . E.; Persichini, P. J . J . Am. Chem. Soc. 1998, 120, 12178.
4842 J . Org. Chem., Vol. 67, No. 14, 2002

Synthesis of Sphingomyelin Carbon Analogues
caused by the specific binding of these compounds to the
substrate binding site of the enzyme, although the effect
of surface dilution was partly involved.
Exp er im en ta l Section
All commercially available reagents were used without
further purification. All solvents were used after distillation.
Tetrahydrofuran and diethyl ether were refluxed over and
distilled from sodium. Dichloromethane was refluxed over and
distilled from P2O5. Dimethylformamide (DMF) was distilled
from CaH2 under reduced pressure. Preparative separation
was usually performed by column chromatography on silica
gel. 1H NMR and 13C NMR spectra were recorded, and
chemical shifts were represented as δ-values relative to the
internal standard TMS. IR spectra were recorded on a Fourier
Transform infrared spectrometer.
F IGURE 4. Inhibition activities.
2-(1-Oxooctyl)-4-bu ta n olid e (9). To a solution of lithium
diisopropylamide prepared from di-iso-propylamine (9.7 mL,
69.6 mmol) and 1.6 N n-butyllithium in hexane (39.8 mL, 63.9
mmol) in THF (58 mL) was added a solution of γ-butyrolactone
(5.00 g, 58.1 mmol) in THF (116 mL) at -78 °C, and the
reaction mixture was stirred for 30 min at the same temper-
ature. To this mixture was added dropwise a solution of octanal
(10.9 mL, 69.7 mmol) in THF (116 mL) at -78 °C. After the
reaction mixture was stirred for an additional 1 h, an aqueous
2 N HCl solution was added, and the resulting mixture was
extracted with ethyl acetate. The organic layers were com-
bined, washed with brine, dried over MgSO4, filtered, and
concentrated in vacuo to give the crude products. Column
chromatography on silica gel (from 16% to 50% ethyl acetate
in hexane) gave 2-(1-hydroxyoctyl)-4-butanolide (11.6 g, 93%)
as a colorless oil. To a solution of the alcohol (9.01 g, 42.0 mmol)
obtained in acetone (210 mL) was added J ones reagent at 0
°C until the color of the reaction mixture turned yellow. After
the reaction mixture was stirred for 5 min at the same
temperature, brine was added, and the resulting mixture was
extracted with ethyl acetate. The organic layers were com-
bined, washed with brine, dried over MgSO4, filtered, and
concentrated in vacuo to give the crude products. Column
chromatography on silica gel (from 11% to 17% ethyl acetate
in hexane) gave 9 (8.21 g, 92.0%) as a colorless oil: IR (NaCl
neat) 2930, 1771, 1725, 1410, 1377 cm-1; 1H NMR (CDCl3, 400
MHz) δ 4.39 (ddd, J ) 6.4, 8.1, 9.0 Hz, 1H), 4.32 (ddd, J ) 6.3,
8.1, 8.8 Hz, 1H), 3.69 (dd, J ) 6.3, 9.3 Hz, 1H), 2.96 (ddd, J )
7.3, 7.6, 18.1 Hz, 1H), 2.78 (tdd, J ) 6.3, 8.1, 12.9 Hz, 1H),
2.61 (ddd, J ) 7.1, 7.6, 17.8 Hz, 1H), 2.32 (m, 1H), 1.55-1.67
(m, 2H), 1.20-1.36 (m, 8H), 0.88 (t, J ) 7.1 Hz, 3H); 13C NMR
(CDCl3, 100 MHz) δ 202.6, 172.8, 67.5, 52.2, 42.2, 31.6, 29.0,
28.9, 23.9, 23.2, 22.6, 14.0; FAB HRMS m/z calcd for C12H21O3
(M+ + H) 213.1491, found 213.1519.
F IGURE 5. Results by detergents.
canoylamino-4-nitrophenylphosphocholine used as the
substrate are 6.0 × 10-9 M and 1.0 mM, respectively.24
As shown in Figure 4, compounds 1 and 2 inhibited the
activity of SMase, and their respective IC50 values were
approximately 120 and 78 µM. The experiments were
conducted exactly as shown in Figure 4. To exclude the
factors for whether the inhibitions by compounds 1 and
2 are due to the surface dilution,25 we examined the
effects of three types of amphiphiles (two detergents and
one phospholipid) on the activity of SMase. Under the
present experimental conditions, 2-hexadecanoylamino-
4-nitrophenylphosphocholine used as a substrate is in a
micellar state. Because compounds 1 and 2 are kinds of
amphiphiles having the same length of aliphatic hydro-
carbon chain as those of dihexanoylphosphatidylcholine
(diC6PC), they are thought to have similar inhibitory
power because of the surface dilution. These three types
of amphiphiles inhibited the enzyme activity to a lesser
extent as shown in Figure 5. The observed inhibitions
by compounds 1 and 2 were much greater than those by
three types of amphiphiles. It was thus concluded that
the inhibitions of SMase by compounds 1 and 2 were
(2S)-[(1R)-Hyd r oxyoctyl]-4-bu ta n olid e (8). (R)-BINAP-
RuCl2 catalyst was prepared in situ from (R)-BINAP (212 mg,
0.341 mmol) and benzene ruthenium chloride dimer (86 mg,
0.171 mmol) in degassed DMF (1.0 mL) at 100 °C for 10 min
under argon atmosphere.15b A solution of 9 (24.2 g, 113.8 mmol)
and (R)-BINAP-RuCl2 catalyst prepared in degassed CH2Cl2
(23 mL) was stirred in an autoclave under 100 atm of hydrogen
at 60 °C for 10 days. Column chromatography on silica gel
(from 11% to 17% ethyl acetate in hexane) gave 8 (24.1 g,
99.0%, 94.8% ee, 94.3% de) as a colorless oil. Its enantiomeric
excess was determined by HPLC analysis using a chiral
column (Daicel chiralcel OD) after benzylation of the secondary
(24) Levade, T.; Salvayre, R.; Douste-Blazy, L. J . Neurochem. 1983,
40, 1762.
(25) Most phospholipases including SMase show interesting but
complex kinetics of hydrolysis, because their substrates are not
monodispersed and exist in a two-dimensional array. The catalytic
mechanisms of phospholipases are complicated by an additional
binding event that may be separated from formation of the Michaelis
complex with substrate. This additional factor is mainly due to the
binding of enzymes to the aggregated surface, which can be affected
by the structure of substrates, kinds of detergents, or other amphi-
philes. When the aggregated substrates (e.g., micelles) at a constant
concentration are diluted with detergents or other amphiphiles, surface
dilution refers to the observation that the specific activities of phos-
pholipases decrease. This phenomenon is attributed to the existing
state of substrates and not enzymes.
hydroxy group according to the procedure reported in the
27.0
literature:17 [R]D
-6.87 (c ) 1.06, CHCl3); IR (NaCl neat)
3482, 2901, 1782 cm-1; 1H NMR (CDCl3, 400 MHz) δ 4.40 (ddd,
J ) 2.9, 8.8, 9.0 Hz, 1H), 4.23 (ddd, J ) 7.3, 9.0, 9.5 Hz, 1H),
4.18 (m, 1H), 2.68 (ddd, J ) 2.9, 9.3, 10.2 Hz, 1H), 2.39 (ddd,
J ) 9.5, 12.4, 19.3 Hz, 1H), 2.16-2.24 (m, 2H), 1.72 (br s, 1H),
1.29-1.57 (m, 11H), 0.88 (t, J ) 6.8 Hz, 3H); 13C NMR (CDCl3,
100 MHz) δ 178.8, 69.2, 67.1, 45.5, 34.7, 31.7, 29.3, 29.2, 25.8,
22.6, 21.8, 14.0; FAB HRMS m/z calcd for C12H23O3 (M+ + H)
232.1913, found 232.1905.
J . Org. Chem, Vol. 67, No. 14, 2002 4843

Hakogi et al.
(4S)-(2-Hyd r oxyeth yl)-(5R)-h ep tyloxa zolid in on e (5). To
a solution of 8 (5.0 g, 23.3 mmol) in DME (23 mL) was added
30% aqueous ammonia (35.3 mL, 600 mmol) at room temper-
ature. After the reaction mixture was stirred for 14.5 h,
solvents were removed in vacuo to give crude 7 as a solid. To
a solution of crude 7 thus obtained in DMF (46.7 mL) were
added N-bromosuccinimide (5.4 g, 30.3 mmol) and silver
acetate (5.06 g, 30.3 mmol) at room temperature. After being
stirred for 3 h, the reaction mixture was diluted with methanol,
and the precipitates which appeared were removed by filtra-
tion. The filtrate was concentrated in vacuo, an aqueous 2 N
HCl solution was added to the residue, and the resulting
mixture was extracted with ethyl acetate. The organic layers
were combined, washed with a 10% sodium hydrogen sulfite
solution and brine, dried over MgSO4, filtered, and concen-
trated in vacuo to give crystals, which were purified by
recrystallization from ethyl acetate to produce 5 (4.15 g, 77.8%
for 2 steps): mp 96.0-96.5 °C; [R]D26.0 -0.84 (c ) 0.84, CHCl3);
mg, 0.044 mmol) at room temperature. After the reaction
mixture was stirred for 3 h, an aqueous 2 N HCl solution was
added at 0 °C, and the resulting mixture was extracted with
ethyl acetate. The organic layers were combined, washed with
brine, dried over MgSO4, filtered, and concentrated in vacuo
to give the crude products. Column chromatography on silica
gel (from 33% to 67% ethyl acetate in hexane) gave 13 (100
25.0
mg, 80.6%) as a colorless oil: [R]D
-0.67 (c ) 0.94, CHCl3);
IR (NaCl neat) 3277, 2930, 1746, 1713, 1256, 1030 cm-1; H
NMR (CDCl3, 400 MHz) δ 7.31-7.40 (m, 5H), 5.17 (s, 2H), 4.75
(ddd, J ) 4.1, 4.4, 8.3 Hz, 1H), 4.65 (d, J ) 9.8 Hz, 1H), 4.02-
4.15 (m, 4H), 3.80 (m, 1H), 1.47-1.93 (m, 10H), 1.43 (s, 9H),
1.31 (t, J ) 7.1 Hz, 6H), 1.20-1.32 (m, 6H), 0.87 (t, J ) 6.8
Hz, 3H); 13C NMR (CDCl3, 100 MHz) δ 155.6, 155.0, 135.2,
128.6, 128.5, 128.2, 80.4, 79.6, 69.7, 61.6 (J C-P ) 6.6 Hz), 61.6
(J C-P ) 6.6 Hz), 52.9 (J C-P ) 17.3 Hz), 31.7, 30.6, 29.3, 29.0,
28.3, 25.3, 22.6, 22.4 (J C-P ) 142.4 Hz), 16.4, 16.4, 14.0; FAB
HRMS m/z calcd for C28H49NO8P (M+ + H) 558.3197, found
558.3196.
1
1
IR (KBr disk) 3362, 3260, 2927, 1724 cm-1; H NMR (CDCl3,
400 MHz) δ 6.13 (br s, 1H), 4.60 (ddd, J ) 3.4, 7.8, 10.0 Hz,
1H), 3.96 (ddd, J ) 3.4, 7.8, 10.0 Hz, 1H), 3.89 (ddd, J ) 4.6,
5.6, 10.7 Hz, 1H), 3.76 (ddd, J ) 4.6, 7.8, 10.7 Hz, 1H), 1.85
(br s, 1H), 1.66-1.79 (m, 3H), 1.47-1.62 (m, 2H), 1.24-1.41
(m, 9H), 0.88 (t, J ) 7.3 Hz, 3H); 13C NMR (CDCl3, 100 MHz)
δ 159.5, 80.1, 60.7, 57.5, 54.9, 54.5, 31.7, 31.5, 30.5, 29.5, 29.3,
29.0, 25.8, 22.6, 14.0; FAB HRMS m/z calcd for C12H24NO3 (M+
+ H) 230.1756, found 230.1808.
(4S)-(2-Br om oeth yl)-(5R)-h ep tyloxa zolid in on e (10). To
a solution of 5 (1.0 g, 4.36 mmol) in CH2Cl2 (22 mL) were added
triphenylphosphine (4.58 g, 17.4 mmol) and carbon tetrabro-
mide (2.89 g, 8.72 mmol) at 0 °C. After the reaction mixture
was stirred for 30 min, the solvent was removed in vacuo.
Column chromatography on silica gel (from 25% to 50% ethyl
acetate in hexane) gave corresponding bromide 10 (1.27 g,
100%) as crystals: mp 76.5-77.5 °C; [R]D16.5 -41.03 (c ) 1.12,
CHCl3); IR (KBr disk) 3268, 2922 cm-1; 1H NMR (CDCl3, 400
MHz) δ 6.96 (br s, 1H), 4.64 (ddd, J ) 3.7, 7.6, 9.8 Hz, 1H),
4.00 (m, 1H), 3.55 (ddd, J ) 4.9, 6.1, 10.5 Hz, 1H), 3.41 (ddd,
J ) 5.9, 9.3, 10.5 Hz, 1H), 1.95-2.09 (m, 2H), 1.72 (m, 1H),
1.48-1.61 (m, 2H), 1.20-1.42 (m, 9H), 0.88 (t, J ) 7.3 Hz, 3H);
13C NMR (CDCl3, 100 MHz) δ: 159.8, 80.1, 53.9, 32.7, 31.7,
29.3, 29.242, 29.0, 25.9, 22.6, 14.0.
Dieth yl 2-[3-N-ter t-bu tyloxyca r bon yl-(5R)-h ep tyl-(4S)-
oxa zolid in on yl]et h ylp h osp h on a t e (12). A solution of 10
(1.27 g, 4.36 mmol) in triethyl phosphite (3.74 mL, 21.8 mmol)
was stirred for 3 h under reflux, and then, the excess of triethyl
phosphite was removed in vacuo to give crude 11, which was
used without further purification. To a solution of 11 thus
obtained in DMF (22 mL) were added N,N-(dimethylamino)-
pyridine (160 mg, 1.31 mmol), triethylamine (0.92 mL, 6.55
mmol), and di-tert-butyl dicarbonate (1.43 g, 6.54 mmol) at 0
°C. After the reaction mixture was stirred at room temperature
for 45 min, a saturated ammonium chloride solution was
added, and the resulting mixture was extracted with ethyl
acetate. The organic layers were combined, washed with brine,
dried over MgSO4, filtered, and concentrated in vacuo to give
the crude products. Column chromatography on silica gel (from
Dieth yl (4R)-Ben zyloxyca r bon yloxy-(3S)-N-(1-oxoh ex-
yl)a m in ou n d ecylp h osp h on a te (3). To a solution of 13 (807
mg, 1.45 mmol) in CH2Cl2 (7.2 mL) was added trifluoroacetic
acid (2.9 mL) dropwise at 0 °C. After being stirred for 3 h, the
reaction mixture was poured into a mixture of an aqueous 1
N NaOH solution and CHCl3 at 0 °C, and the resulting mixture
was stirred for 5 min at the same temperature. To this mixture
was added hexanoic chloride (0.40 mL, 2.89 mmol) at 0 °C.
After the reaction mixture was stirred for an additional 15
min, a saturated NH4Cl solution was added, and the resulting
mixture was extracted with CHCl3. The organic layers were
combined, washed with brine, dried over MgSO4, filtered, and
concentrated in vacuo to give the crude products. Column
chromatography on silica gel (from 50% to 65% ethyl acetate
25.0
in hexane) gave 3 (701 mg, 84.5%) as a colorless oil: [R]D
-1.18 (c ) 0.81, CHCl3); IR (NaCl neat) 3281, 2930, 1746, 1651,
1256, 1030 cm-1; 1H NMR (CDCl3, 400 MHz) δ 7.32-7.39 (m,
5H), 5.97 (d, J ) 8.8 Hz, 1H), 5.15 (s, 2H), 4.74 (m, 1H), 4.05-
4.20 (m, 5H), 2.14 (t, J ) 7.6 Hz, 2H), 1.15-1.91 (m, 28H),
0.89 (t, J ) 6.8 Hz, 3H), 0.87 (t, J ) 7.1 Hz, 3H); 13C NMR
(CDCl3, 100 MHz) δ 173.3, 155.1, 135.2, 128.6, 128.5, 128.2,
80.5, 69.7, 61.7, 51.2 (J C-P
) 14.8 Hz), 36.7, 31.7, 31.4, 31.1,
29.3, 29.0, 25.4, 25.3, 22.6, 22.4, 22.3 (J C-P ) 141.4 Hz), 16.4,
14.0, 13.9; FAB HRMS m/z calcd for C28H51NO7P (M+ + H)
556.3403, found 556.3409.
(4R)-Ben zyloxycar bon yloxy-(3S)-N-(1-oxoh exyl)am in o-
u n d ecylp h osp h on och olin e (14). To a solution of 3 (209 mg,
0.365 mmol) in CH2Cl2 (0.4 mL) was added a solution of
trimethylsilyl bromide (0.48 mL, 3.65 mmol) in CH2Cl2 (0.7
mL) dropwise at room temperature. After the reaction mixture
was stirred for 3 h at the same temperature, the solvent was
removed in vacuo. The resulting crude silylester was dissolved
in MeOH (5 mL), and the solution was stirred for an additional
1 h to complete the hydrolysis of the silylester. Then, the
solvent was removed in vacuo to give the crude phosphonic
acid. To a suspension of the crude phosphonic acid thus
obtained and choline chloride (234 mg, 1.68 mmol) in pyridine
(9.86 mL) was added trichloroacetonitrile (1.57 mL, 15.70
mmol) at 60 °C. After the reaction mixture was stirred for 8
h, the solvent was removed in vacuo. The precipitates that
appeared were removed by filtration through Celite, and the
filtrate was concentrated in vacuo. Column chromatography
on silica gel (from 12.5% methanol in chloroform to 4.3% water
and 26.6% methanol in chloroform) gave 14 as a slightly
brownish foam. It was purified again by HPLC (50% acetoni-
trile in water; flow rate, 2 mL/min; retention time, 29 min;
column, Develosil C8-5, 10 × 250, 216 nm detection) to afford
pure 14 (37 mg, 16.1%) as a colorless foam: IR (NaCl neat)
50% to 75% ethyl acetate in hexane) gave 12 (1.96 g, 100% for
25.0
two steps) as a colorless oil: [R]D
+25.80 (c ) 1.11, CHCl3);
IR (NaCl neat) 2932, 1723, 1254, 1026 cm-1; 1H NMR (CDCl3,
400 MHz) δ 4.48 (m, 1H), 4.27 (m, 1H), 4.05-4.15 (m, 4H),
2.09 (m, 1H), 1.70-1.92 (m, 4H), 1.49-1.63 (m, 2H), 1.55 (s,
9H), 1.33 (t, J ) 7.1 Hz, 6H), 1.20-1.38 (m, 9H), 0.89 (t, J )
7.1 Hz, 3H); 13C NMR (CDCl3, 100 MHz) δ 151.8, 149.4, 84.0,
78.3, 61.8 (J C-P ) 5.8 Hz), 61.7 (J C-P ) 5.8 Hz), 57.9 (J C-P
)
19.9 Hz), 31.6, 29.2, 29.0, 28.5, 27.9, 25.8, 22.6, 22.5, 21.9, 21.9,
21.5, 16.5, 16.4, 14.0; FAB HRMS m/z calcd for C16H33NO5P
(M+ + H - (CH3)3COCO-) 350.2096, found 350.2082.
1
3387, 2928, 1738, 1649 cm-1; H NMR (CD3OD, 400 MHz) δ
Dieth yl (4R)-Ben zyloxyca r bon yloxy-(3S)-N-ter t-bu tyl-
oxyca r bon yla m in ou n d ecylp h osp h on a te (13). To a solu-
tion of 12 (100 mg, 0.22 mmol) in THF (2.2 mL) were added
benzyl alcohol (0.05 mL, 0.44 mmol) and cesium carbonate (14
7.31-7.39 (m, 5H), 5.17 (d, J ) 12.2 Hz, 1H), 5.12 (d, J ) 12.2
Hz, 1H), 4.76 (m, 1H), 4.23 (m, 2H), 4.07 (m, 1H), 3.59 (m,
2H), 3.20 (s, 9H), 2.18 (t, J ) 7.1 Hz, 2H), 1.87 (m, 1H), 1.46-
1.69 (m, 7H), 1.22-1.36 (m, 14H), 0.91 (t, J ) 6.8 Hz, 3H),
4844 J . Org. Chem., Vol. 67, No. 14, 2002

Synthesis of Sphingomyelin Carbon Analogues
0.89 (t, J ) 6.8 Hz, 1H); 13C NMR (CD3OD, 100 MHz) δ 175.0,
155.0, 135.0, 128.0, 127.9, 127.6, 79.4, 68.9, 66.3, 57.1, 57.0,
53.1, 51.8, 51.6, 25.5, 31.3, 30.9, 29.7, 28.8, 28.7, 25.3, 24.9,
23.8, 22.6, 22.4, 22.1, 21.9, 12.9, 12.8; FAB HRMS m/z calcd
for C30H54N2O7P (M+ + H) 585.3671, found 585.3673.
(4R)-Hyd r oxy-(3S)-N-(1-oxoh exyl)a m in ou n d ecylp h os-
p h on och olin e (1). To a solution of 14 (37 mg, 0.058 mmol)
in methanol (0.58 mL) was added palladium on carbon (7 mg).
After the reaction mixture was stirred under H2 atmosphere
for 1 h, the catalyst was removed by filtration, and the filtrate
was concentrated in vacuo to give residues, from which title
(317 mg, 0.58 mmol) in CH2Cl2 (2.9 mL) was added trifluoro-
acetic acid (1.2 mL) dropwise at 0 °C. After being stirred for 3
h, the reaction mixture was poured into a mixture of aqueous
1 N NaOH (5 mL) and CHCl3 (5 mL) at 0 °C, and the resulting
mixture was stirred for another 5 min at the same tempera-
ture. To this mixture was added hexanoic chloride (0.16 mL,
1.16 mmol) at 0 °C, and the mixture was stirred for 15 min. A
saturated NH4Cl solution was added, and the resulting
mixture was extracted with CHCl3. The organic layers were
combined, washed with brine, dried over MgSO4, filtered, and
concentrated in vacuo to give the crude products. Column
chromatography on silica gel (from 50% to 75% ethyl acetate
compound
1 (20 mg, 76.9%) was obtained as a soluble
25.0
component in MeOH (1 mL) as a foam: [R]D26.0 +4.65 (c ) 0.72,
in hexane) gave 4 (249 mg, 78.8%) as a colorless oil: [R]D
MeOH); IR (KBr disk) 3420, 2928, 1642, 1053 cm-1; H NMR
1
-14.40 (c ) 0.414, CHCl3); IR (NaCl neat) 3379, 2930, 1744,
1651, 1545, 1458, 1381, 1259, 1035 cm-1; 1H NMR (CDCl3, 400
MHz) δ 7.32-7.39 (m, 5H), 5.71 (br d, J ) 8.8 Hz, 1H), 5.16
(s, 2H), 4.72 (ddd, J ) 3.7, 4.2, 7.8 Hz, 1H), 4.15 (m, 1H), 3.72
(s, 3H), 3.70 (s, 3H), 2.13 (t, J ) 7.3 Hz, 2H), 1.40-1.90 (m,
10H), 1.20-1.40 (m, 14H), 0.89 (t, J ) 7.1 Hz, 3H), 0.87 (t, J
) 7.1 Hz, 3H); 13C NMR (CDCl3, 100 MHz) δ 173.1, 155.1,
135.2, 128.6, 128.5, 128.2, 80.9, 69.7, 52.3, 50.2, 36.7, 31.6, 31.3,
31.0, 29.3, 29.1, 29.0, 25.4, 25.3, 24.0 (J C-P ) 139.9 Hz), 22.4,
22.3, 19.0, 14.0, 13.9; FAB HRMS m/z calcd for C28H49NO7P
(M+ + H) 542.3246, found 542.3288.
(CD3OD, 400 MHz) δ 4.25 (m, 2H), 3.73 (ddd, J ) 3.2, 6.3, 9.5
Hz, 1H), 3.61 (m, 2H), 3.44 (m, 1H), 3.22 (s, 9H), 2.21 (t, J )
7.6 Hz, 2H), 1.97 (m, 1H), 1.30-1.66 (m, 21H), 0.92 (t, J ) 6.6
Hz, 3H), 0.90 (t, J ) 7.1 Hz, 3H); 13C NMR (CD3OD, 100 MHz)
δ 171.7, 73.0, 66.3, 57.1 (J C-P ) 4.9 Hz), 54.6, 54.5, 53.1, 35.7,
33.2, 31.5, 31.1, 29.2, 28.8, 25.4, 25.3, 23.8, 23.4, 22.4, 22.1,
21.9, 12.9, 12.8; FAB HRMS m/z calcd for C22H48N2O5P (M+
H) 451.3303, found 451.3271.
+
Dim et h yl 3-[3-N-ter t-b u t yloxyca r b on yl-(5R)-h ep t yl-
(4S)-oxa zolid in on yl]p r op ylp h osp h on a te (16). To a solu-
tion of dimethyl methylphosphonate 15 (0.85 mL, 7.85 mmol)
in THF (7.8 mL) was added a solution of 1.6 N n-butyllithium
in hexane (4.9 mL, 7.84 mmol) at -78 °C. After the reaction
mixture was stirred for 30 min at -78 °C, a solution of 10
(765 mg, 2.62 mmol) in THF (5.2 mL) was added dropwise at
the same temperature. After the reaction mixture was stirred
for 1 h, di-tert-butyl dicarbonate (1.71 g, 7.84 mmol) was added
at the same temperature, and the resulting mixture was
stirred for an additional 20 min. The reaction was quenched
with a saturated NH4Cl solution, and then, the solvents were
removed in vacuo. Chloroform was added to the obtained
residue, and the chloroform layer was decanted, dried over
MgSO4, and concentrated in vacuo to give the crude products.
Column chromatography on silica gel (from 50% to 75% ethyl
(5R)-Ben zyloxycar bon yloxy-(4S)-N-(1-oxoh exyl)am in o-
d od ecylp h osp h on och olin e (18). To a solution of 4 (507 mg,
0.93 mmol) in CH2Cl2 (0.9 mL) was added a solution of TMSBr
(1.2 mL, 9.34 mmol) in CH2Cl2 (1.9 mL) dropwise at room
temperature. After the reaction mixture was stirred for 25 min,
the solvent was removed in vacuo. The resulting crude
silylester was dissolved in MeOH (5 mL), and the solution was
stirred for an additional 1 h at room temperature to complete
the hydrolysis of the silylester. Then, the solvent was removed
in vacuo to give the crude phosphonic acid. To a suspension of
the crude phosphonic acid thus obtained and choline chloride
(1.09 g, 7.81 mmol) in pyridine (78 mL) was added trichloro-
acetonitrile (3.4 mL, 33.6 mmol) at 50 °C. After the reaction
mixture was stirred for 40 h, the solvent was removed in vacuo.
The precipitates that appeared were removed by filtration
through Celite, and the filtrate was concentrated in vacuo.
Column chromatography on silica gel (from 14% methanol in
chloroform to 4.3% water and 26.6% methanol in chloroform)
gave 18 as a slightly brownish foam. It was purified again by
HPLC (50% acetonitrile in water; flow rate, 2 mL/min; reten-
tion time, 31 min; column, Develosil C8-5, 10 × 250, 216 nm
acetate in hexane) gave 16 (714 mg, 62.5%) as a colorless oil:
23.0
[R]D
+15.24 (c ) 0.592, CHCl3); IR (NaCl neat) 2930, 1815,
1740, 1717, 1248, 1035 cm-1
;
1H NMR (CDCl3, 400 MHz) δ
4.47 (m, 1H), 4.21 (q, J ) 7.6 Hz, 1H), 3.75 (s, 3H), 3.72 (s,
3H), 1.63-1.92 (m, 9H), 1.55 (s, 9H), 1.24-1.45 (m, 9H), 0.89
(t, J ) 7.1 Hz, 3H); 13C NMR (CDCl3, 100 MHz) δ 152.0, 149.5,
83.8, 57.6, 52.3, 31.6, 29.2, 29.0, 28.5, 27.9, 25.8, 23.8, 22.5,
18.93, 18.88, 14.0; FAB HRMS m/z calcd for C15H31NO5P (M+
+ H - (CH3)3COCO-) 336.1940, found 336.1978.
detection) to afford pure 18 (89 mg, 17.0%) as a colorless foam:
20.5
[R]D
-5.06 (c ) 0.937, MeOH); IR (NaCl neat) 3368, 2928,
1742, 1647, 1545, 1460, 1261, 1188, 1053, 966 cm-1; 1H NMR
(CD3OD, 400 MHz) δ 7.31-7.39 (m, 5H), 5.16 (d, J ) 12.4 Hz,
1H), 5.12 (d, J ) 12.4 Hz, 1H), 4.74 (m, 1H), 4.23 (m, 2H),
4.04 (m, 1H), 3.58 (m, 2H), 3.20 (s, 9H), 2.17 (t, J ) 7.1 Hz,
2H), 1.46-1.70 (m, 10H), 1.22-1.36 (m, 14H), 0.91 (t, J ) 6.8
Hz, 3H), 0.89 (t, J ) 6.8 Hz, 1H); 13C NMR (CD3OD, 100 MHz)
δ 176.37, 156.61, 137.24, 129.61, 129.46, 129.16, 81.19, 70.47,
67.83 (m), 58.52 (d, J c-p ) 4.96 Hz), 54.73, 54.70, 54.66, 52.26,
37.11, 32.88, 32.51, 31.30, 30.86 (d, J c-p ) 14.88 Hz), 30.37,
30.23, 27.67 (d, J c-p ) 135.64 Hz), 23.67, 23.46, 21.56 (d, J c-p
) 4.13 Hz), 14.43, 14.34; FAB HRMS m/z calcd for C31H55N2O7P
(M+ + H) 599.3825, found 599.3870.
(5R)-Hyd r oxy-(4S)-N-(1-oxoh exyl)a m in od od ecylp h os-
p h on och olin e (2). To a solution of 18 (80 mg, 0.13 mmol) in
methanol (2.7 mL) was added palladium on carbon (16 mg).
After the reaction mixture was stirred under H2 atmosphere
for 1 h, the catalyst was removed by filtration, and the filtrate
was concentrated in vacuo to give residues, from which title
Dim eth yl (5R)-Ben zyloxyca r bon yloxy-(4S)-N-ter t-bu -
tyloxyca r bon yla m in od od ecylp h osp h on a te (17). To a so-
lution of 16 (100 mg, 0.23 mmol) in THF (1.15 mL) were added
benzyl alcohol (0.12 mL, 1.15 mmol) and cesium carbonate (164
mg, 0.504 mmol) at room temperature. After the reaction
mixture was stirred for 4 h, an aqueous 2 N HCl solution was
added at 0 °C, and then, the solvents were removed in vacuo.
Chloroform was added to the obtained residue, and the
chloroform layer was decanted, dried over MgSO4, and con-
centrated in vacuo to give the crude products. Column chro-
matography on silica gel (from 33% to 60% ethyl acetate in
23.0
hexane) gave 17 (70 mg, 56.5%) as a colorless oil: [R]D
-10.35 (c ) 0.578, CHCl3); IR (NaCl neat) 3277, 2930, 1743,
1711, 1458, 1259, 1172, 1033 cm-1; 1H NMR (CDCl3, 400 MHz)
δ 7.32-7.39 (m, 5H), 5.16 (s, 2H), 4.73 (ddd, J ) 4.2, 4.4, 8.5
Hz, 1H), 4.58 (d, J ) 9.8 Hz, 1H), 3.79 (m, 1H), 3.73 (s, 3H),
3.70 (s, 3H), 1.47-1.85 (m, 9H), 1.43 (s, 9H), 1.24-1.39 (m,
9H), 0.87 (t, J ) 7.1 Hz, 3H); 13C NMR (CDCl3, 100 MHz) δ
155.6, 155.0, 135.2, 128.54, 128.47, 128.2, 80.7, 79.5, 69.6, 52.3,
51.8, 31.7, 30.6, 29.9 (J C-P ) 16.4 Hz), 29.3, 29.0, 28.3, 27.7
(J C-P ) 29.6 Hz), 25.3, 24.1 (J C-P ) 142.4 Hz), 22.5, 18.9, 14.0;
FAB HRMS m/z calcd for C27H47NO8P (M+ + H) 544.3039,
found 544.3019.
compound
2 (54 mg, 88.5%) was obtained as a soluble
21.0
component in MeOH (1 mL) as a foam: [R]D
-9.12 (c )
0.353, MeOH); IR (NaCl neat) 3420, 2928, 1637, 1051 cm-1
;
1H NMR (CD3OD, 400 MHz) δ 4.24 (m, 2H), 3.74 (m, 1H), 3.60
(m, 2H), 3.44 (ddd, J ) 2.4, 5.9, 7.3 Hz, 1H), 3.22 (s, 9H), 2.20
(t, J ) 7.6 Hz, 2H), 1.22-1.74 (m, 24H), 0.92 (t, J ) 7.1 Hz,
3H), 0.90 (t, J ) 7.0 Hz, 3H); 13C NMR (CD3OD, 100 MHz) δ
Dim eth yl (5R)-Ben zyloxyca r bon yloxy-(4S)-N-(1-oxo-
h exyl)a m in od od ecylp h osp h on a te (4). To a solution of 17
J . Org. Chem, Vol. 67, No. 14, 2002 4845

Hakogi et al.
176.2, 74.9, 67.88 (J C-P ) 6.6 Hz), 67.86 (J C-P ) 6.6 Hz), 58.6,
55.0, 54.72, 54.69, 54.65, 37.3, 34.8, 33.0, 32.7, 31.9 (J C-P
the Ministry of Education, Science and Culture, J apan.
T.H. is grateful to the J SPS for a Research Fellowship
for Young Scientists.
)
15.7 Hz), 30.8, 30.4, 28.0 (J C-P ) 135.6 Hz), 27.0, 26.9, 23.7,
23.5, 21.5, 21.4, 14.4, 14.3; FAB HRMS m/z calcd for C27H47
-
NO8P (M+ + H) 465.3457, found 465.3456.
Su p p or tin g In for m a tion Ava ila ble: Selected 400 MHz
1H NMR and 13C NMR spectra of 1, 2, and the synthetic
intermediates. This material is available free of charge via the
Internet at http://pubs.acs.org.
Ack n ow led gm en t. We thank Professors M. Nishi-
zawa at Tokushima Bunri University and H. Yamada
at Kwansei Gakuin University for their kind advice on
Noyori’s asymmetric hydrogenation. This research is
supported by a Grant-in-Aid for Scientific Research from
J O025529O
4846 J . Org. Chem., Vol. 67, No. 14, 2002