Stereochemistry of the DBU/LiCl-Assisted Nucleophilic Substitution at Phosphorus in Nucleoside-3′-O-(Se-methyl Methanephosphonoselenolate)s

Article abstract of DOI:10.1021/jo980225g

The crystal and molecular structure of diastereomerically pure N⁴-benzoyl-2′-deoxycytidine 3′-O-(Se-methyl methanephosphonoselenolate (FAST-eluted) (4, R = H, B = C^Bz) and 5′-O-pixylthymidine 3′-O-(S-methyl methanephosphonothiolate (FAST-eluted) (5) have been elucidated by X-ray crystallography. The absolute configuration at the phosphorus atom in both compounds is Sp. Each FAST-4′ (R = DMT, B = C^Bz) and FAST-5 (R = Px, B = Thy) in the process of DBU/LiCl-assisted condensation with 3′-O-acetyl-A⁴-benzoylcytidine and 3′-O-acetylthymidine gave after deprotection (S_p)-dicytidine-(3′,5′)-methanephosphonate and (S_P)- dithymidine-(3′,5′)-methanephosphonate, respectively. Unambiguous assignment of the absolute configuration at the phosphorus in 4 (R = H, B = C^Bz) and 5 (R = Px, B = Thy) allows for stereochemical correlation and the conclusion that DBU/LiCl-assisted nucleophilic substitution at phosphorus occurs with net inversion of configuration, in contrast to our earlier erroneous deduction.⁵ Moreover, the knowledge of the absolute configuration at the phosphorus atom in both 4 (R = H, B = C^Bz) and 5 (R = Px, B = Thy) allows for assignment of the absolute configuration at phosphorus in precursors of 4′ and 5, such as 5′-O-DMT-nucleoside 3′-O- methanephosphonothioanilidates (6), methanephosphonoanilidates (8), and methanephosphonoselenoanilidates (9).

Full text of DOI:10.1021/jo980225g

J . Org. Chem. 1998, 63, 5395-5402
5395
Ster eoch em istr y of th e DBU/LiCl-Assisted Nu cleop h ilic
Su bstitu tion a t P h osp h or u s in Nu cleosid e-3′-O-(Se-m eth yl
Meth a n ep h osp h on oselen ola te)s†
Lucyna A. Woz´niak,‡ Michał Wieczorek,§ J arosław Pyzowski,‡ Wiesław Majzner,§ and
Wojciech J . Stec*,‡
Polish Academy of Sciences, Centre of Molecular and Macromolecular Studies, Department of Bioorganic
Chemistry, Sienkiewicza 112, 90-363 Ło´dz´, Poland, and Institute of General Food Chemistry, Technical
University of Ło´dz´, Stefanowskiego 4/ 10, 90-924 Ło´dz´, Poland
Received February 6, 1998
The crystal and molecular structure of diastereomerically pure N4-benzoyl-2′-deoxycytidine 3′-O-
(Se-methyl methanephosphonoselenolate (FAST-eluted) (4, R ) H, B ) CBz) and 5′-O-pixylthymidine
3′-O-(S-methyl methanephosphonothiolate (FAST-eluted) (5) have been elucidated by X-ray
crystallography. The absolute configuration at the phosphorus atom in both compounds is SP. Each
F AST-4′ (R ) DMT, B ) CBz) and F AST-5 (R ) Px, B ) Thy) in the process of DBU/LiCl-assisted
condensation with 3′-O-acetyl-N4-benzoylcytidine and 3′-O-acetylthymidine gave after deprotection
(SP)-dicytidine-(3′,5′)-methanephosphonate and (SP)- dithymidine-(3′,5′)-methanephosphonate, re-
spectively. Unambiguous assignment of the absolute configuration at the phosphorus in 4 (R ) H,
B ) CBz) and 5 (R ) Px, B ) Thy) allows for stereochemical correlation and the conclusion that
DBU/LiCl-assisted nucleophilic substitution at phosphorus occurs with net inversion of configuration,
in contrast to our earlier erroneous deduction.5 Moreover, the knowledge of the absolute
configuration at the phosphorus atom in both 4 (R ) H, B ) CBz) and 5 (R ) Px, B ) Thy) allows
for assignment of the absolute configuration at phosphorus in precursors of 4′ and 5, such as 5′-
O-DMT-nucleoside 3′-O- methanephosphonothioanilidates (6), methanephosphonoanilidates (8), and
methanephosphonoselenoanilidates (9).
In tr od u ction
(all-RP)-Oligo(nucleoside methanephosphonate)s (1) and
chimeric oligonucleotide constructs (2) containing incor-
porated (RP)-dinucleoside-(3′,5′)-methanephosphonates
(3) (Figure 1) constitute promising second-generation
antisense mRNA molecules due to their resistance to
nucleolytic degradation and their high affinity for comple-
mentary mRNA.1
Stereocontrolled synthesis of 1 and 3 is still a matter
of challenge in several research establishments, although
the chimeric molecules 2 have become available from
either chromatographic separation of dinucleoside-(3′,5′)-
methanephosphonates (3), followed by incorporation of
(RP)-3 into an oligonucleotide chain,2 or the stereocon-
trolled synthesis of (RP)-3 from diastereomerically pure
precursors such as 5′-O-DMT-nucleoside 3′-O-(4-nitro-
F igu r e 1. Schematic representations of (RP)-dinucleoside
(3′,5′)-methanephosphonates and chimeric constructs.
phenyl methanephosphonates),3 5′-O-DMT-nucleoside-3′-
O-(O-hexafluoroisopropyl methanephosphonates)4 or 5′-
O-DMT-nucleoside-3′-O-(Se-methyl methanephosphono-
selenolates) (4′).5 Recently a novel, cost-effective ste-
reospecific synthesis of (RP)-3 based upon the stereocon-
trolled synthesis of 5′-O-DMT-nucleoside-3′-O-(S-methyl
methanephosphonothiolates) (5) has been reported from
this laboratory.6
The synthesis of (RP)-3, depicted in Scheme 1, relies
upon the synthesis and separation into diastereomers of
protected nucleoside 3′-O-methanephosphonothioanili-
dates (6).
* To whom correspondence should be addressed. Tel.: (48-42)
6819744. Fax: (48-42) 6815483. E-mail: wjstec@bio.cbmm.lodz.pl.
† This paper is dedicated to Professor Marian Mikołajczyk on the
occasion of his 60th Birthday.
§ Technical University of Ło´dz´.
(1) Reynolds, M. A.; Hogrefe, R. I.; J aeger, J . A.; Schwartz, D. A.;
Riley, T. A.; Marvin, W. B.; Daily, W. J .; Vaghefi, M. M.; Beck, T. A.;
Knowles, S. K.; Klem, R. E.; Arnold, L. J ., J r. Nucleic Acids Res. 1996,
24, 4584-4591.
(2) Miller, P. S.; Annan, N. D.; McParland, K. B.; Pulford, S. M.
Biochemistry 1982, 21, 2507-2512.
(3) Les´nikowski, Z. J .; J aworska, M.; Stec, W. J . Nucleic Acids Res.
1990, 18, 2109-2115.
(5) Woz´niak, L. A.; Pyzowski, J .; Wieczorek, M.; Stec, W. J . J . Org.
Chemistry 1994, 59, 5843-5846.
(6) Stec, W. J .; Woz´niak, L. A.; Pyzowski, J .; Niewiarowski, W.
Antisense Nucleic Acid Drug Dev. 1997, 7, 381-395.
(4) Cormier, J . F.; Pannunzio, T. Tetrahedron Lett. 1991, 32, 7161-
7164.
S0022-3263(98)00225-4 CCC: $15.00 © 1998 American Chemical Society
Published on Web 07/15/1998

5396 J . Org. Chem., Vol. 63, No. 16, 1998
Woz´niak et al.
Sch em e 1
F igu r e 3. Molecular structure of F AST-5 (R ) Px, B ) Thy)
(molecule a ) showing 50% probability displacement ellipsoids.
Resu lts a n d Discu ssion
Syn th esis of Selen oloester 4 (R ) H, B ) CBz) a n d
Th ioloester 5 (R ) P x, B ) Th y). The reaction of 5′-
O-DMT-N4-benzoyl-2′-deoxycytidine with MePCl2 was
performed in THF at -40 °C and was followed by
addition of aniline and elemental selenium. After stan-
dard workup, 9 (R ) DMT, B ) CBz) was obtained in 71%
yield as a mixture of two diastereomers (in ∼1:1 ratio),
which were separated into diastereomerically pure spe-
cies by means of column chromatography (silica gel 60,
230-400 mesh) to give F AST-9 (R ) DMT, B ) CBz) and
SLOW-9 (R ) DMT, B ) CBz). Each separated diaste-
reomer 9 (R ) DMT, B ) CBz) in DMF or THF solution
was activated with sodium hydride (1.3 molar equiv), and
into the resulting slurry the stream of dry CO2 was
introduced. The obtained sodium 5′-O-DMT-N4-benzoyl-
2′-deoxycytidine 3′-O-methanephosphonoselenoate was,
without isolation, alkylated with MeI (5 equiv) to give
the corresponding 5′-O-DMT-N4-benzoyl-2′-deoxycytidine
3′-O-(Se-methyl methanephosphonoselenolate) (4′); thus,
from methanephosphonoselenoanilidate F AST-9 (R )
DMT, B ) CBz), slow-eluted ester SLOW-4′ (R ) DMT,
B ) CBz) was obtained, and methanephosphonoselenoa-
nilidate SLOW-9 (R ) DMT, B ) CBz) was the substrate
for preparation of fast-eluted Se-methyl ester F AST-4′.
Ester F AST-4′ (R ) DMT, B ) CBz) was deprotected by
an acid treatment and purified by means of flash silica
gel column chromatography, and pure F AST 4 (R ) H,
B ) CBz) was crystallized in darkness from EtOH-H2O
(95:5 v/v) solution.
F igu r e 2. Molecular structure of F AST-4 (R ) H, B ) CBz
)
showing 50% probability displacement ellipsoids.
The corollary of our current strategy is the use of both
separated diastereomers of each precursor 6 for prepara-
tion of the desired (RP)-diastereomer 5.
Unfortunately, in our earlier work,5 the absolute con-
figuration at the phosphorus in FAST- 5′-O-pixylthymi-
dine 3′-O-(Se-methyl methanephosphonoselenolate) (7)
was erroneously assigned, which led us to the incorrect
conclusion that the process of DBU/LiCl-assisted con-
densation of 5′-O-pixylthymidine 3′-O-(Se-methyl metha-
nephosphonoselenolate) 7 with 3′-O- acetylthymidine
occurred with retention of configuration.
In this report, the results of the detailed X-ray studies
of fast-eluted diastereomers of N4- benzoyl-2′-deoxycyti-
dine-3′-O-(Se-methyl methanephosphonoselenolate) (4) (R
) H, B ) CBz) (Figure 2) and fast-eluted 5′-O-Px-
thymidine 3′-O-(S-methyl methanephosphonothiolate) (5)
(R ) Px, B ) Thy) (Figures 3 and 4) are presented. Both
F AST-5 (R ) Px, B ) Thy) and F AST-4 (R ) H, B )
CBz) appeared to have the (SP)-configuration. Because
DBU/LiCl assisted condensation of FAST-(SP)-5 with 3′-
acetylthymidine gives (SP)-dithymidine-3′,5′-methane-
phosphonate ((SP)-3), this process occurs with inversion
of configuration. Moreover, since each diastereomer 4′
and 5 is prepared in the Wadsworth-Emmons-Horner-
type reaction7a via stereoretentive conversion either from
a single diastereomer of 5′-O-DMT-nucleoside 3′-O-
methanephosphonothioanilidate 6 or methanephospho-
noanilidate 8, followed by alkylation, an unambiguous
assignment of the absolute configuration at phosphorus
in diastereomers 6 and 8 has become feasible.
Analogously, 5′-O-Px-thymidine gave the appropriate
methanephosphonothioanilidate (6) (R ) Px, B ) Thy)
after treatment with MePCl2 at -40 °C, followed by
addition of aniline and elemental sulfur. After standard
workup, 6 (R ) Px, B ) Thy) was obtained in 91% yield
as a mixture of two diastereomers, which were separated
into diastereomerically pure species by silica gel column
chromatography to give F AST-6 (R ) Px, B ) Thy) and
SLOW-6 (R ) Px, B ) Thy).
(7) (a) Stec, W. J . Acc. Chem. Res. 1983, 16, 411-417. (b) Lesiak,
K.; Lesnikowski, Z. J .; Stec, W. J .; Zielinska, B. Pol. J . Chem. 1979,
53, 2041-2050. (c) W. J . Stec, Khimia i Primienienije Fosfororgan-
itcheskich Sojedinienij, Kirsanov, A. V. Ed.; Naukova Dumka: Kiev,
1981; pp 281-289. (d) Lesiak, K.; Stec, W. J . Z. Naturforsch. 1978,
33b, 782-785.
Each separated diastereomer 6 was converted (NaH/
CO2 treatment) into sodium 5′-O-Px-thymidine-3′-O-

DBU/LiCl-Assisted Nucleophilic Substitution
J . Org. Chem., Vol. 63, No. 16, 1998 5397
3′-O > O > Me, the absolute configurations at phosphorus
in compounds F AST-4′ (R ) DMT, B ) CBz) and F AST-5
(R ) Px, B ) Thy) were assigned as (SP). These data
were used as the foundation for reinvestigation of the
stereochemical pathway of the substitution of the alkylse-
leno group in (Se-alkyl methanephosphonoselenolate)s
and the S-alkyl group in (S-alkyl methanephosphonothi-
olate)s by alcohols, in particular by the 5′-OH group of
nucleosides.
Con d en sa tion of F AST-4′ (R ) DMT, B ) CBz) w ith
3′-O-a cetyl-2′-d eoxy-N4-ben zoylcytid in e. FAST-elut-
ed 5′-O-DMT-N4-benzoyl-2′-deoxycytidine 3′-O-(Se-methyl
methanephosphonoselenolate) ((SP)-4′) was condensed
with 3′-O-acetyl-2′-deoxy-N4-benzoylcytidine. The reac-
tion was performed in MeCN in the presence of DBU and
LiCl (Scheme 1). The reaction progress was controlled
by HP TLC. Upon completion (decease of (SP)-4′), the
reaction mixture was diluted with chloroform, washed
twice with 0.05 M citric acid, concentrated to dryness,
and purified by silica gel column chromatography. The
product of condensation, SLOW-N4-benzoyl-5′-O-DMT-
2′-(deoxycytidylyl)-(3′,5′)-N4-benzoyl-3′-O-acetyl-2′-deoxy-
cytidine-3′- methanephosphonate (3, R ) DMT, B ) CBz),
was isolated, fully characterized (31P NMR δ 33.00 ppm;
F igu r e 4. Molecular structure of F AST-5 (R ) Px, B ) Thy)
(molecule b) showing 50% probability displacement ellipsoids.
1H NMR (CDCl3) δ, 1.58 (d, 3H, 2J P-CH )17.6, Hz, PCH3);
3
FAB-MS [M - H] 1067), and deprotected according to
methanephosphonothioate and, without isolation, was
alkylated with MeI (5 equiv) to give the corresponding
5′-O-Px-thymidine-3′-O-(S-methyl methanephosphono-
thiolate (5) (R ) Px, B ) Thy). Compound F AST-6 (R )
Px, B ) Thy) was converted into SLOW-5 (R ) Px, B )
Thy), while SLOW-6 (R ) Px, B ) Thy) was a substrate
for preparation of F AST-5 (R ) Px, B ) Thy). Fully
protected F AST-5 (R ) Px, B ) Thy) was crystallized
from a CH2Cl2-hexane mixture (slow evaporation of
methylene chloride).
the standard procedures.5
After RP-HPLC purification, fully deprotected 3 (R )
H, B ) Cyt) was compared with the authentic sample of
the diastereomeric mixture of d(CPMeC) obtained on a
solid support via nonstereospecific methanephosphonoa-
midite method and by co-injection on RP-HPLC it was
found to be identical with the SLOW-eluted diastereomer,
(SP)-3.9d
In an analogous way, the reaction between FAST-(SP)-5
and 3′-O-acetylthymidine was performed. The resulting
fully protected dimer SLOW-3 (R ) DMT, B ) Thy) was
isolated, characterized (31P NMR δ 34.14; 1H NMR δ
X-r a y stu d ies. The molecular structures of methane-
phosphonoselenoloester F AST-4 (R ) H, B ) CBz) and
methanephosphonothioloester F AST-5 (R ) Px, B ) Thy)
were determined by a single-crystal X- ray analysis and
are presented in Figures 2 [F AST-4 (R ) H, B ) CBz)],
3, and 4 [F AST-5 (R ) Px, B ) Thy)].
2
(CDCl3) 1.57 (d, 3H, J P-CH ) 17.6 Hz, PCH3); FAB-MS
3
[M - H] 887), and after deprotection compared with an
authentic sample of MIX-3 (R ) H, B ) Thy) prepared
via the methanephosphonoamidite method. The product
of condensation was identical, as proved by RP HPLC,
with (SP)-3. The condensation reactions, under the same
conditions, led to the opposite products, namely (RP)-3,
when the opposite diastereomers SLOW-4′ and SLOW-5
were used as substrates.
The compound F AST-4 (R ) H, B ) CBz) consists of
one molecule in the independent part of the elemental
cell, while F AST-5 (R ) Px, B ) Thy) consists of two
formal a and b molecules in the independent part of the
elemental cell, both possessing the absolute configuration
at the chiral centers identical with the configuration of
F AST-4 (R ) H, B ) CBz), and determined as RC1′, SC3′
,
Th e Ster eoch em istr y of Con d en sa tion of Nu cleo-
sid e 3′-O-(Se-Met h yl m et h a n ep h osp h on oselen o-
la te)s (4′) a n d S-Meth yl Meth a n ep h osp h on oth i-
ola t es (5) w it h 3′-O-P r ot ect ed Nu cleosid es: A
Cor r ection . As described above, F AST-4′ and F AST-5
were the precursors of SLOW-3 while the condensation
of SLOW-4′ and SLOW-5 with the appropriately pro-
tected nucleosides, under analogous conditions, gave
F AST-3 (Scheme 1).
Since all data coming from both X-ray and NMR
studies confirmed the absolute configuration at phospho-
rus in F AST-3 as (RP)-isomers and that of the SLOW-3
as (SP)9 (Figure 1), the DBU/LiCl-assisted nucleophilic
RC4, and SP1. Both the substitution of selenium for the
sulfur atom, incorporation of the 5′-Px protecting group
in F AST-5, as well as different crystallization conditions
change significantly the structure of the elemental cell,
thus creating enough space for four molecules of hexane
to cocrystallize inside the elemental cell [structure FAST-5
(R ) Px, B ) Thy)].
Crystal data and structure solution for F AST-4 and
F AST-5 are collected in Table 1. Further detailed
disscussion of these results is presented in the Supporting
Information.
Using the Cahn-Ingold-Prelog rules8 for the descrip-
tion of the absolute configuration at the phosphorus
center, with the priority order MeSe(MeS) > nucleoside
(9) (a) Chacko, K. K.; Lindner, K.; Saenger, W.; Miller P. S. Nucl.
Acids Res. 1983, 11, 2801-2814. (b) Stec, W. J .; Zon, G.; Egan, W.;
Byrd, R. A.; Phillips, L. R.; Gallo, K. A. J . Org. Chem. 1985, 50, 3908-
3913. (c) Han, F.; Watt, W.; Duchamp, D. J .; Callahan, L.; Kezdy, F.
J .; Agarwal, K. Nucl. Acids Res. 1990, 18, 2759-2767. (d) Lo¨schner,
T.; Engels, J . W. Nucl. Acids Res. 1990, 18, 5083-5088.
(8) Cahn, R. S.; Ingold, C.; Prelog, V. Angew. Chem., Int. Ed. Engl.
1966, 5, 385-414. Brill, W. K.-D; Caruthers, M. H. Tetrahedron Lett.
1988, 29, 1227-1230. Caruthers, M. H.; Haltiwanger, R. C. Acta
Crystallogr. 1989, C45, 1639-1641.

5398 J . Org. Chem., Vol. 63, No. 16, 1998
Woz´niak et al.
Ta ble 1. Cr ysta l Da ta a n d Str u ctu r e Refin em en t for F AST-4 (R ) H, B ) CBz) a n d F AST-5 (R ) P x, B ) Th y)
F AST-4
F AST-5
mol formula
formula wt
cryst syst
space grp
a (Å)
C
18H22N3O6PSe
C37H45N2O7PS
692.78
monoclinic
P21
18.828(10)
9.007(4)
23.899(9)
105.89(3)
3897.9(31)
4
486.32
monoclinic
P21
9.419(2)
7.911(2)
13.668(3)
94.69(3)
1025.3(4)
2
b (Å)
c (Å)
â (deg)
V (Å3)
Z
Dc (g/cm3)
1.575
35.81
1.181
15.06
µ (cm-1
)
cryst dimens (mm)
max 2θ (deg)
0.12 × 0.16 × 0.32
0.03 × 0.28 × 0.65
150
146
radiatn, λ (Å)
scan mode
Cu KR, 1.541 84
ω/2θ
Cu KR, 1.541 78
ω/2θ
scan width (deg)
hkl ranges: (h)
(k)
1.27 + 0.14 tan θ
-11 11
-10 10
1.25 + 0.14 tan θ
-23 23
-11
0
(l)
-17
0
-29 29
0.9026
0.9994
0.9782
8256
6979
4267
858
18
EAC correction: (min)
max
avg
0.8731
1.0000
0.9328
4217
4038
3742
337
no. of reflns: (unique)
refine with I > 0σ(I)
obsd with I > 2σ(I)
no. of param refined
no. of restraints
largest diff. peak (e Å-3
0
)
)
0.423
0.430
largest diff. hole (e Å-3
Robs
-1.193
0.0576
0.1434
0.1156
1.088
-0.252
0.0626
0.1379
0.0782
1.144
wRobs
weighting coeff.a (m)
Sobs
shift/esd max
Rint
Tmeas.
-0.001
0.0530
293(2)
496
-0.001
0.0873
293(2)
1472
F 000
abs structure
flack param ø
R
C1′, SC3′, RC4′, SP1
RC1′, SC3′, RC4′, SP1
0.01(3)
0.00(3)
a
2
Weighting scheme w ) [σ2(Fo2) + (mP)2]-1 where P ) (Fo + 2Fc2)/3.
oxathiaphospholane ring-opening condensation process12
occurs according to the adjacent type mechanism13 and
the cleavage of the P-S bond occurs with retention of
configuration. Ab initio studies of this reaction profile
suggest that the ring opening with retention of configu-
ration is energetically favorable.14 In both discussed
reactions the P-S bond is cleaved, and DBU is used as
an activator, most probably being responsible for proton
abstraction from the 5′-OH group of an attacking nucleo-
side. Interestingly, if in the reaction of 4′ or 5 with the
5′-OH of nucleoside DBU is used as a sole activator,
condensation occurs at a much lower rate and is ac-
companied by partial loss of stereospecificity. Such
observation may suggest that DBU, being usually con-
sidered as a strong nonnucleophilic base, without com-
petitive nucleophile may also act as nucleophilic re-
agent.15 However, as demonstrated by Masamune,16 and
substitution at phosphorus in methane phosphonothio-
(seleno) derivatives with methylseleno or methylthio
leaving groups occurs with inversion of configuration.
These results indicate that the process of nucleophilic
substitution at phosphorus in methanephosphonosele-
nolates or methanephosphonothiolates occurs according
to the in-line mechanism, where the transition state
requires the location of the nucleophile attacking at
phosphorus at the face opposite to the leaving group.
Such a stereochemical result is not surprising, although
it contrasts with some earlier results reported by several
authors.10 Although in Inch’s report the stereochemical
diversity of SN2(P) substitution is demonstrated as
depending upon the mode of catalysis,11 in this laboratory
it has been demonstrated that the DBU-assisted 1,3,2-
(10) Several examples of stereoretentive reactions of nucleophilic
substitution in MeP(O)(OR)(SMe) have been reported: (a) DeBruin,
K. E.; Tang, C.-I. W.; J ohnson, D. M.; Wilde, R. L. J . Am. Chem. Soc.
1989, 111, 5871-5879. (b) Moriyama, M.; Bentrude, W. G. J . Am.
Chem. Soc. 1983, 105, 4727-4733. (c) DeBruin, K. E.; J ohnson, D. M.
J . Chem. Soc., Chem. Commun. 1975, 753-754. (d) Van den Berg, G.
R.; Platenburg, D. H. J . M.; Benschop, H. P. Recl. Trav. Chim. Pays-
Bas 1972, 91, 929-934. (e) Donohue, J .; Mandel, N.; Farnham, W. B.;
Murray, R. K., J r.; Mislow, K.; Benschop, H. P. J . Am. Chem. Soc. 1971,
93, 3792-3793. (f) Benschop, H. P.; Platenburg, D. H. J . M. J . Chem.
Soc., Chem. Commun. 1970, 1098-1099. (g) Farnham, W. B.; Murray,
R. K., J r.; Mislow, K. J . J . Chem. Soc., Chem. Commun. 1970, 605. (h)
Benschop, H. P.; Van den Berg, G. R.; Boter, H. L. Recl. Trav. Chim.
Pays-Bas 1968, 87, 387-395.
(12) Uznan´ski, B.; Grajkowski, A.; Krzyz˘anowska, B.; Kaz´mierkows-
ka, A.; Stec, W. J .; Wieczorek, M. W.; Błaszczyk, J . J . Am. Chem. Soc.
1992, 114, 10197-10202.
(13) Stec, W. J .; Grajkowski, A.; Karwowski, B.; Kobylan´ska, A.;
Koziołkiewicz, M.; Misiura, K.; Okruszek, A.; Wilk, A.; Guga, P.;
Boczkowska, M. J . Am. Chem. Soc. 1995, 117, 12019-12029.
(14) Uchimaru, T.; Stec, W. J .; Taira, K. J . Org. Chem. 1997, 62,
5793-5800.
(15) (a) Fujii, M.; Ozaki, K.; Kume, A.; Sekine, M. Tetrahedron 1987,
43, 3395-3407. (b) Schenk, W. A.; Urban, P.; Dombrowski, E. Chem.
Ber. 1993, 126, 679. (c) Reed, R.; Reau, R. W.; Daham, F.; Bertrand,
G. Angew. Chem., Int. Ed. Engl. 1993, 32, 399-401. (d) Chambers, R.
D.; Roche, A. J .; Batsanov, A. B.; Howard, J . A. J . Chem. Soc., Chem.
Commun. 1994, 2055-2056. (e) Lammers, H.; Cohen-Fernandes, P.;
(11) Hall, C. R.; Inch, T. D. Tetrahedron Lett. 1976, 3645-3648.

DBU/LiCl-Assisted Nucleophilic Substitution
J . Org. Chem., Vol. 63, No. 16, 1998 5399
Sch em e 2a
a
Key: (i) MePCl2, Et3N, -40 °C, THF, 1 h; (ii) aniline, elemental selenium or sulfur rt, 1-2 h; (iii) NaH/CO2, DMF, followed by methyl
iodide.
independently by Seebach,17 addition of lithium salts to
lenolate)s (4′, R ) DMT, B ) Thy) were prepared via
procedures analogous to those presented in Scheme 2. If
5′-O-DMT(or Px)-thymidine was used as the substrate
for condensation with methyldichlorophosphine followed
by substitution with aniline and elemental sulfur was
used as an oxidant of transiently formed methane-
phosphonoanilidites, 5′-O-DMT (or Px)-thymidine 3′-O-
methanephosphonothioanilidates 6 were obtained as
major products. Chromatographic separation gave pure
F AST-6 and SLOW-6 isomers.
DBU potentiates its basicity, so DBU/LiCl is a much more
efficient catalyst of nucleophilic substitution at phospho-
rus. This explanation does not elucidate, however, the
essential difference in stereochemistries of both reactions
discussed here. The ring-opening condensations must
occur with participation of the pentacoordinate interme-
diate due to its five-membered ring stabilization, and an
attack of nucleophile at phosphorus occurs from the site
opposite to the most electronegative ligand such as
endocyclic oxygen. The requirement of an apical entry
of nucleophile and an apical departure of the leaving
group18 is violated here because pseudorotation is con-
certed with formation/cleavage of an equatorial bond in
the TBP species. According to such a mechanistic
proposal, nucleophilic substitution at phosphorus in-
volved in the 1,3,2-oxathiaphospholane ring occurs with
retention of configuration. In the process of nucleophilic
substitution at phosphorus in 4′ and 5, an attack of
alkoxide must occur from the site opposite to the -SeMe
or -SMe ligand via an SN2(P)-type mechanism without
involvement of the TBP intermediate. Besides enhance-
ment of DBU basicity (pKa 11.6 in water15 and 23.9 in
MeCN19), the function of LiCl may be also considered in
terms of coordination of lithium cation with methane-
phosphonate oxygen atom, additionally increasing the
electrophilicity of phosphorus center. That assumption
is lacking, however, any experimental evidence.
Reexamination of the results of X-ray analysis, leading
to the correction of our former erroneous assignment of
the absolute configuration of 5′-O-pixylthymidine 3′-O-
(Se-methyl methanephosphonoselenolate) (7)5 (FAST-
eluted isomer has (SP) configuration) and additional
X-ray data of 4 (R ) H, B ) CBz) and 5 (R ) Px, B )
Thy) allow us to correlate the absolute configuration of
compounds 4′ and 5 with the mobility of diastereomers
in HPLC and with the sense of the chemical shift in 31P
NMR. It is also possible to assign the absolute configu-
rations at phosphorus of precursors of compounds 4′ and
5, the corresponding methanephosphonoselenoanilidates
9, and sulfur analogues 6 (Scheme 3).
The competitive route to thioesters 5 involves the
utilization of methanephosphonoanilidates 8, which can
be obtained directly in the reaction of appropriately
protected nucleosides with methanephosphonodichlori-
date, followed by a substitution of chlorine by aniline in
transiently formed methanephosphonochloridates.20 Ad-
ditionally, as demonstrated at Scheme 3, the stereore-
tentive oxidation of methanephosphonothioanilidates 6
and methanephosphonoselenoanilidates 9 to methane-
phosphonoanilidates 8 permits for the correlation of the
absolute configuration of 8 with the data obtained from
X-ray analysis of 4 (R ) H, B ) CBz) and 5 (R ) Px, B )
Thy). The oxidation of F AST-6 or F AST-9, accomplished
under very mild conditions [2-fold excess of 0.1 M
OXONE (potassium peroxymonosulfate), buffered at pH
7 in THF-MeOH solution) within 15 min leads exclu-
sively to the F AST-8, while SLOW-6 and SLOW-9 are
oxidized to SLOW-8. This oxidation proceeds with full
Cor r ela tion a l An a lysis of th e Absolu te Con figu -
r a tion a t P h osp h or u s in Meth a n ep h osp h on oth io-
a n ilid a t es (6), Met h a n ep h osp h on oa n ilid a t es (8),
a n d Meth a n ep h osp h on oselen oa n ilid a tes (9). As
described in our earlier paper,5 both diastereomeric 5′-
O-DMT-thymidine 3′-O-(Se-methyl methanephosphonose-
Habraken, C. L. Tetrahedron 1994, 50, 865-870. (f) Lesnikowski, Z.
J .; Zabawska, D.; J aworska-Mas´lanka, M. M.; Schinazi, R. F.; Stec,
W. J . New J . Chem. 1994, 18, 1197-1204. (g) Bouhadir, G.; Reed, R.
W.; Reau, R.; Bertrand, G. Heteroatom Chem. 1995, 6, 371-375.
(16) Blanchette, M. A.; Choy, W.; Davis, J . T.; Essenfeld; A. P.;
Masamune, S.; Roush, W. R.; Sakai, T. Tetrahedron Lett. 1984, 21,
2183-2186.
(17) Seebach, D.; Thaler, A.; Blaser, D.; Ko, S. Y. Helv. Chim. Acta
1991, 74, 1102-1118.
(18) Westheimer, F. H. Acc. Chem. Res. 1968, 1, 70-78.
(19) Leffek, K. T.; Pruszyn´ski, P.; Thanapaalasingham, K. Can. J .
Chem. 1989, 67, 990-994.
(20) Pyzowski, J .; Woz´niak, L. A.; Stec, W. J . Collect. Czech. Chem.
Commun. 1996, 61, 1, S152-153.

5400 J . Org. Chem., Vol. 63, No. 16, 1998
Woz´niak et al.
Sch em e 3a
Sch em e 4a
a
Key: (i) NaH/CO2, DMF, followed by methyl iodide; (ii) Oxone;
(iii) NaH/CS2, DMF, followed by methyl iodide.
to TMS (1H) and 80% H3PO4 (31P) as external standards. 2D
1H-1H NMR (NOESY) correlations were applied for identifica-
1
tion of signals in H NMR spectra. Unless otherwise specified,
spectra were measured in CDCl3 at 295 K. The following
abbreviations are used: s, singlet; d, doublet; t, triplet; m,
multiplet. Positive chemical shift values were assigned for
compounds resonating at lower fields than standards. Mass
spectra were recorded on a mass spestrometer with a Cs+ gun
operating at 13 keV using a 3-nitrobenzyl alcohol matrix.
High-resolution MS spectra were recorded for 1:1 mixtures of
diastereomers. High-pressure liquid chromatography was
performed by the use of a reversed-phase column (C-18, 5 µm,
25 cm, 4.6 mm) and the following solvent system: 0.1 M
triethylammonium bicarbonate (TEAB) at pH ) 7.0 (A) and
40% MeCN in 0.1 M TEAB (B). Column chromatography and
HP-TLC analyses were performed on silica gel (240-400 mesh)
and precoated F254 silica gel plates, respectively. Solvents and
reagents were purified according to the standard laboratory
techniques and stored under argon. MeCN was distilled from
CaH2 directly to the reaction vessels. LiCl was recrystallized
from MeOH and dried under vacuum (150 °C/0.05 mmHg) for
several days. Reactions involving air- or moisture-sensitive
compounds were carried out in glassware that was dried either
in an oven or under high vacuum and then placed under a
positive pressure of argon.
a
Key: (i) Oxone; (ii) NaH/CO2, DMF, followed by methyl iodide;
(iii) NaH/CS2, DMF, followed by methyl iodide; (iv) 5′-OH nucleo-
side, DBU/LiCl.
retention of configuration, as proven in model studies in
which 1,3,2-dioxaphosphorinanes were used (data not
shown).21
SLOW-eluted diastereomers 4′ and 5 both have the
(RP)-configuration. The reactions of activation of oxo-,
thio, and selenoanilidates by means of NaH and further
reactions with electrophiles (CO2, CS2, and CSe2)7b (PN
f PS, PN f PO, and PN f PSe conversions) leading to
corresponding phosphylthio- and phosphylselenoates,
respectively, are well documented and proceed with full
retention of configuration.7 The S- and Se-alkylations
take place beyond the stereogenic center so they cannot
change the P-absolute configurations in thio- and sele-
noalkyl esters, as related to the precursors 6 and 9.
In the conversion of methanephosphonoanilidates-
(RP)-8 to S-alkyl esters 5 (Scheme 4), sulfur in the ester
5 originates from CS2 introduced during the Wadsworth-
Emmons-Horner type reaction.7a Thus, the sequence of
reactions presented in Scheme 4 correlating in one
stereochemical cycle monomers 4′ or 5 with their precur-
sors 6, 8, and 9, respectively, creates an open, four-
membered, diligostatic, antipodal stereochemical cycle
with three retentions of configuration and one ligand
metathesis involved.22
Yields of pure isomers, obtained during purifications, are
given in brackets.
Gen er a l P r oced u r e for P r ep a r a tion of 5′-O-DMT-(N-
p r otected ) 2′-Deoxyn u cleosid e 3′-O-Meth a n ep h osp h o-
n oth ioa n ilid a tes (6) a n d Meth a n ep h osp h on oselen oa n il-
id a tes (9). To a solution of methyldichlorophosphine (0.23 g,
2 mmol) in THF (10 mL) and triethylamine (0.40 g, 4 mmol)
cooled to -40 °C by external cooling (dry ice-acetone) was
added dropwise, under magnetic stirring, a solution of 5′-O-
DMT (or Px) (N-protected, except thymidine) nucleoside (1
mmol) in THF (10 mL). After removal of external cooling,
stirring was continued until the reaction mixture reached
ambient temperature. Then aniline (0.28 g, 3 mmol) together
with elemental selenium (or sulfur) (2.5 mmol) were added in
one portion. Stirring was continued overnight. Excess sele-
nium (or sulfur) was removed by filtration, and solvents were
partially evaporated to reduce the volume of the reaction
mixture to ca. 5 mL. The residue was diluted with chloroform,
and the resulting solution was washed with saturated aqueous
NaHCO3. The organic fraction was dried over anhyd MgSO4
and concentrated. Product 6 or 9 was isolated by silica gel
chromatography (elution with a gradient of EtOH in CHCl3).
Exp er im en ta l Section
Gen er a l Meth od s. NMR spectra were recorded either at
300.13 MHz (1H) and 121.47 MHz (31P) or at 500.133 MHz (1H)
and 202.47 MHz (31P). Chemical shifts are reported (δ) relative
(21) Woz´niak, L. A.; Koziolkiewicz, M.; Kobylanska, A.; Stec, W. J .
Biochem. Med. Chem. Lett., in press.
(22) Cram, D. J .; Cram, J . M. Top. Curr. Chem. 1972, 31, 1-43.

DBU/LiCl-Assisted Nucleophilic Substitution
J . Org. Chem., Vol. 63, No. 16, 1998 5401
Appropriate fractions were combined and concentrated under
reduced pressure.
(m, 1H, H4′), 4.95 (m, 1H, H3′), 6.3 (t, 6.1, 12.8, 1H, H1′); HR
FAB-MS found [M - H] 785.270 (calcd 785.274). SLOW-
1
(RP ,SP )-5′-O-DMT-t h ym id in e 3′-O-(m et h a n ep h osp h o-
n oth ioa n ilid a te) (6) was obtained from 5′-O-DMT-thymidine
(0.54 g, 1 mmol) as a colorless foam, total yield 93% (0.66 g).
(SP ): yield 40%; 31P NMR δ 29.95; H NMR δ 1.56 (d, 15.6,
3H, PCH3), 2.28 (m, 1H, H2′), 2.50 (m, 1H, H2′′), 3.31 (m, 2H,
H5′, H5′′), 4.15 (m, 1H, H4′), 4.94 (m, 1H, H3′), 6.15 (dd, 7.1,
13.7, 1H, H1′); HR FAB-MS found [M - H] 785.270.
1
F AST-(RP ): yield 45%; 31P NMR δ 79.43; H NMR δ 2.04 (d,
15.3, 3H, PCH3) 2.37 (m, 2H, H2′, H2′′), 3.53 (m, 2H, H5′, H5′′),
4.41 (d, 1.4, 1H, H4′), 5.27 (d, 9.4, 1H), 6.36 (dd, 6.8, 6.8, 1H,
H1′); FAB-MS found [M - H] 712.3 (calcd 712.225). SLOW-
Gen er a l P r oced u r e for P r ep a r a tion of Dia ster eom er i-
ca lly P u r e (RP )- a n d (SP )-5′-O- DMT-(N-p r otected )-2′-
d eoxyn u cleosid e 3′-O-(Se-Meth yl m eth a n ep h osp h on ose-
len ola te)s (4′) or S-Meth yl Meth a n ep h osp h on oth iola tes
(5). To a stirred solution of corresponding 6 or 9 [1 mmol,
dried twice by coevaporation with dry toluene (5 mL)] in DMF
(10 mL) was added NaH (1.2 molar equiv of 50% suspension
in mineral oil) in a few portions. Stirring was continued until
evolution of hydrogen had ceased, and then a stream of gaseous
CO2 (dried over P2O5) was passed through the slurry for ca. 4
h. The resulting sodium salt of the corresponding nucleoside
3′-O-methanephosphonoselenoic(thioic) acid was treated with
MeI (5 equiv), and the progress of alkylation was followed by
TLC. Upon completion of the reaction, solvents and excess
MeI were removed by evaporation under reduced pressure. The
solid residue was dissolved in CHCl3 (20 mL) and washed twice
with saturated NaHCO3. The combined organic layers were
dried over anhydrous MgSO4 and concentrated. Ester 4′ or 5
was purified and separated into (RP)- and (SP)-diastereomers
by silica gel column chromatography (Kieselgel 60, 230-400
mesh, 3 g of 5 per 90 g of silica gel). The appropriate fractions,
eluted with CHCl3-EtOH (1-5% EtOH), were combined and
evaporated under reduced pressure.
1
(SP ): yield 30%; 31P NMR δ 79.58; H NMR δ 2.01 (d, 15.3,
3H, PCH3), 2.42 (m, 4.2, 2.9, 1H, H2′), 2.73 (m, 1H, H2′′), 3.29
(dd, 2.6, 10.6, 1H, H5′), 3.17 (dd, 2.6, 10.7, 1H, H5′′), 4.17 (d,
1.9, 1H, H4′), 6.48 (dd, 9.1, 9.0, 1H, H1′); FAB-MS found [M
- H] 712.3.
5′-O-DMT-N4-ben zoyl-2′-d eoxycytid in e 3′-O-m eth a n e-
p h osp h on oth ioa n ilid a te (6) was obtained from 5′-O-DMT-
N4-benzoyl-2′-deoxycytidine (0.633 g, 1 mmol). A mixture of
diastereomers (about 1:1 ratio) was obtained as a colorless
foam, total yield 70% (0.56 g). F AST-(RP ): yield 28%; 31P
NMR δ (CH2Cl2/C6D6) 79.33, 1H NMR δ 2.05 (d, 15.3, 3H,
PCH3), 2.27 (dd, 6.5, 13.7, 1H, H2′), 2.77 (dd, 5.1, 11.7, 1H,
H2′′), 3.54 (dd, 2.54, 11.1, 1H, H5′), 3.60 (dd, 3.2, 11.1, 1H,
H5′′), 4.59 (m, 2.4, 1H), 6.36 (dd, 6.6 6.2, 1H, H1′); FAB-MS
found [M - H] 801.3 (calcd 802.590). SLOW-(SP ): yield 32%;
1
31P NMR δ (CH2Cl2/C6D6) 79.84; H NMR δ 2.02 (d, 15.3, 3H,
PCH3); FAB-MS found [M - H] 801.3.
(RP ,SP )-5′-O-DMT-N4-b en zoyl-2′-d eoxycyt id in e 3′-O-
m eth a n ep h osp h on oselen oa n ilid a te (9) was obtained from
5′-O-DMT-N4-benzoyl-2′-deoxycytidine (0.633 g, 1 mmol).
A
mixture of diastereomers (1:1.2 ratio) was obtained as a
5′-O-P x-th ym id in e 3′-O-(S-Meth yl m eth a n ep h osp h o-
n oth iola te) (5). SLOW-(RP) was obtained from F AST-6 (R
) Px, B ) Thy): yield 86%; 31P NMR δ 57.76; 1H NMR δ 1.81
(d, 15.6, 3H, PCH3), 2.29 (d, 13.2, 3H, SCH3), 2.45 (m, 1H, H2′),
2.57 (m, 1H, H2′′), 3.47 (m, 2H, H5′, H5′′), 4.38 (d, 1.2, 1H,
H4′), 5.29 (m, 1H, H3′), 6.46 (dd, 5.4, 8.9, 1H, H1′); FAB-MS
found [M - H] 606.1 (calcd). F AST-(SP ) was obtained from
SLOW-6 (R ) Px, B ) Thy): yield 80%; 31P NMR δ 58.61; 1H
NMR δ 1.82 (d, 15.7, 3H, PCH3), 2.18 (d, 3H, SCH3), 2.45 (m,
1H, H2′), 2.62 (m, 1H, H2′′), 3.44 (m, 2H, H5′, H5′′), 4.18 (d,
1.9, 1H, H4′), 5.34 (m, 1H, H3′), 6.47 (dd, 5.6, 8.7, 1H, H1′);
FAB-MS found [M - H] 606.1.
colorless foam, total yield 70% (0.56 g). F AST-(RP ): yield 30%;
1
1
31P NMR δ 76.65, J P-Se ) 820 Hz; H NMR δ 2.06 (d, 14.8,
3H, PCH3), 2.3 (m, 1H, H2′), 2.55 (m, 1H, H2′′), 3.41 (dd, 2.4,
10.7, 1H, H5′), 3.32 (dd, 2.5, 10.6, 1H, H5′′), 4.15 (m, 1H), 5.5
(m, 1H), 6.45 (m, 1H, H1′); FAB-MS found [M - H] 849.4
(80Se) (calcd 849.5). SLOW-(SP ): yield 25%; 31P NMR δ 76.29
1
(1J P-Se ) 820 Hz); H NMR δ 1.96 (d, 13.54, 3H, PCH3), 2.37
(m, 1H, H2′), 2.5 (m, 1H, H2′′), 2.94 (dd, 2.5, 10.5, 1H, H5′),
3.1 (dd, 3.0, 10.5, 1H, H5′′), 4.35 (m, 1H,), 5.25 (m, 1H), 6.38
(m, 1H, H1′); FAB-MS found [M - H] 849.4 (80Se).
Gen er a l P r oced u r e for P r ep a r a t ion of 5′-O-DMT-
n u cleosid e 3′-O-Meth a n ep h osp h on oa n ilid a tes (8) fr om
Meth a n ep h osp h on od ich lor id a te. To a solution of MeP(O)-
Cl2 (2 mmol) in dry pyridine (15 mL) was added a solution of
5′-O-DMT-nucleoside (1 mmol) in pyridine (5 mL). The
reaction mixture was stirred at room temperature for 15 min,
and aniline (5 mmol, 0.465 g) was added. After the reaction
was completed (30 min), all solvents and reagents were
evaporated to ca. 1/3 volume.
The residue was dissolved with CHCl3 (20 mL) and extracted
with citric acid (0.05 M, 10 mL, three times). Combined
organic layers were concentrated and purified by silica gel
column chromatography [eluent: chloroform-EtOH (0-3%)].
Appropriate fractions of F AST-8 and SLOW-8 were collected
and concentrated to give pure 8 as colorless foams.
5′-O-DMT-N4-ben zoyl-2′-d eoxycytid in e 3′-O-(S-Meth yl
m eth a n ep h osp h on oth iola te) (5, R ) DMT, B ) CBz).
SLOW-[Rp ] was obtained from F AST-7 (R ) DMT, B ) CBz
)
(0.82 g, 1 mmol): yield 82% (0.67 g); 31P NMR δ 55.41; 1H NMR
δ 1.82 (d, 15.6, 3H, PCH3), 2.21 (d, 13.2, 3H, SCH3), 2.92 (m,
2H, H2′, H2′′), 3.48 (m, 2H, H5′, H5′′), 4.31 (d, 3.0, 1H, H4′),
5.28 (m, 1H, H3′), 6.39 (t, 12.8, 1H, H1′); HR FAB-MS found
[M - CH3] 726.204 (calcd 726.203). F AST-(SP ) was obtained
from SLOW-7 (R ) DMT, B ) CBz) (0.82 g, 1 mmol): yield
80% (0.65 g); 31P NMR δ 55.41; HR FAB- MS found [M - CH3]
726.204.
5′-O-DMT-th ym idin e 3′-O-(Se-m eth yl m eth an eph osph o-
n oselen ola te). (4′, R ) DMT, B ) Thy) was obtained from 9
(R ) DMT, B ) Thy) (0.760 g, 1 mmol). Total yield of both
isomers (RP) and (SP): 86%. SLOW-(RP ): yield 40%; 31P NMR
δ 49.76 (1J P-Se ) 430 Hz); 1H NMR δ 1.91 (d, 15.1, 3H, PCH3),
2.16 (d, 11.5, 3H, SeCH3); HR FAB-MS found [M - CH3]
685.124 (calcd [M - CH3] 685.140). F AST-(SP ): yield 46%;
31P NMR δ 49.72 (1J P-Se ) 428 Hz); 31P NMR δ 1.93 (d, 15.2,
3H, PCH3), 2.18 (d, 11.6, 3H, PSeCH3); HR FAB-MS found
[M - CH3] 685.124.
5′-O-DMT-t h ym id in e 3′-O-m et h a n ep h osp h on oa n ili-
d a te (8) was obtained from 5′-O-DMT-thymidine (0.54 g, 1
mmol) and purified and separated by column chromatography
(CHCl3-2% EtOH), total yield 91%. F AST-(RP ): yield 45%;
31P NMR δ 30.52; 1H NMR δ 1.66 (d, 16.9, 3H, PCH3), 2.31
(dd, 5.7, 14.3, 1H, H2′), 2.47 (dd, 6.5, 13.7, 1H, H2′), 3.50 (m,
2.3, 2.4, 2H, H5′, H5′′), 6.47 (dd, 8.9, 9.0, 1H, H1′); HR FAB-
MS found [M - H] 696.244 (calcd 696.247). SLOW-(SP ): yield
1
45%; 31P NMR δ 30.12; H NMR δ 1.70 (d, 16.9, 3H, PCH3),
5′-O-DMT-N4-ben zoyl-2′-d eoxycytid in e 3′-O-(Se-Meth yl
2.40 (m, 6.1, 14.6, 1H, H2′′), 2.76 (m, 5.4, 13.8, 1H, H2′′), 3.27
(dd, 10.7, 2.9, 1H, H5′), 4.02 (dd, 10.6, 2.6, 1H, H5′′), 6.52 (dd,
8.9, 9.0, 1H, H1′); HR FAB- MS found [M - H] 696.244 (calcd
696.247).
m eth a n ep h osp h on oselen ola te) (4′, R ) DMT, B ) CBz).
SLOW-(RP ) was obtained from F AST-9 (R ) DMT, B ) CBz
)
1
(1 mmol): yield 70%; 31P NMR δ 49.76 (1J P-Se ) 430 Hz); H
NMR δ 1.92 (d, 15.1, 3H, PCH3), 2.15 (d, 11.6, 3H, SeCH3),
2.40 (m, 1H, H2′), 2.95 (n, 1H, H2′′), 3.50 (m, 2H, H5′, H5′′),
4.32 (dd, 2.9, 1H, H3′), 5.31 (m, 1H, H4′), 6.41 (dd, 6.3, 6.4,
1H, H1′); HR FAB-MS found [M - H] 788.159 (calcd 788.164).
5′-O-DMT-N4-ben zoyl-2′-d eoxycytid in e 3′-O-m eth a n e-
p h osp h on oa n ilid a te (8) was obtained from 5′-O-DMT-N4-
benzoyl-2′-deoxycytidine (0.63 g, 1 mmol) and purified and
separated into diastereomers by single column chromatogra-
phy (CHCl3-3% EtOH), total yield 90%. F AST-(RP ): yield
F AST-(SP ) was obtained from SLOW-9 (R ) DMT, B ) CBz
)
1
(1 mmol): yield 72%; 31P NMR δ 49.72 (1J P-Se ) 428 Hz); H
NMR δ 1.92 (d, 15.1, 3H, PCH3), 2.19 (d, 11.6, 3H, SeCH3),
2.38 (m, 1H, H2′), 2.95 (m, 1H, H2′′), 3.44 (dd, 3.3, 11.3, 1H,
1
45%; 31P NMR δ 30.12; H NMR δ 1.51 (d, 16.5, 3H, PCH3),
2.13 (m, 1H, H2′), 2.36 (1H, H2′′), 3.17 (m, 2H, H5′, H5′′), 4.18

5402 J . Org. Chem., Vol. 63, No. 16, 1998
Woz´niak et al.
H5′), 3.49 (dd, 2.7, 10.7, 1H, H5′′), 4.54 (dd, 2.2, 1H, H3′), 5.26
(m, 1H, H4′), 6.33 (dd, 5.9, 1H, H1′); FAB-MS found [M - H]
788.2 (80Se).
(R ) DMT, B ) Thy) and 3′-O-acetylthymidine: yield 86%;
31P NMR δ 34.14; 1H NMR δ 1.57 (d, 17.6, 3H, PCH3), 2.43
(m, 2H, H2′, H2′′), 2.56 (m, 2H, H22′, H22′′), 3.44 (m, 2H, H15′,
H15′′), 4.01 (m, 2H, H25′, H25′′), 4.16 (m, 1H, H14′), 4.23 (m,
1H, H24′), 5.08 (m, 5.0, 1H, H13′), 5.22 (m, 1H, H23′), 6.34
(dd, 8.7, 3.2, 1H, H11′), 6.58 (dd, 8.5, 2.9, 1H, H21′); FAB-MS
found [M - H] 887.3.
Dep r otection of 4′ (R ) DMT, B ) CBz). F AST-4′ (0.07
g) was dissolved in 3% trichloroacetic acid in CH2Cl2 (2 mL)
and stirred at room temperature for 3 min. The reaction was
quenched with NaHCO3, and 4 (R ) H, B ) CBz) was extracted
with CHCl3, dried with MgSO4, and concentrated under
reduced pressure. Compound F AST-4 (R ) H, B ) CBz) was
purified by silica gel column chromatography and crystallized
from EtOH/H2O solution (95:5 v/v). Obtained crystals (mp
>200 °C dec) were subjected to X-ray and spectrometric
analyses: 31P NMR (DMSO-d6) δ 50.70 (1J P-Se ) 522.6 Hz);
1H NMR δ 1.97 (d, 15.15, 3H, PCH3), 2.19 (d, 11.4, 3H, SeCH3),
2.33 (dd, 6.6, 14.1, 1H, H2′), 2.62 (dd, 6.0, 14.0, 1H, H2′′), 3.67
(m, 2H, H5′, H5′′), 4.26 (dd, 2.2, 1H, H3′), 5.08 (m, 1H, H4′),
6.20 (dd, 7.3, 6.2, 1H, H1′); HR FAB-MS found [M - H]
486.033 (calcd 486.035).
(RP )-N4-Ben zoyl-5′-O-DMT-2′-d eoxycytid ylyl-(3′,5′)-N4-
b en zoyl-3′-O-a cet yl-2′-d eoxycyt id in e 3′-m et h a n ep h os-
p h on a te (3) was obtained from (RP )-4′ (R ) DMT, B ) CBz
)
and 3′-O-acetyl-N4-benzoyl-2′-deoxycytidine: yield 82%; 31P
1
NMR δ 32.98; H NMR δ 1.65 (d, 17.5, 3H, PCH3); FAB-MS
found [M - H] 1067 (calcd 1067.359).
(SP )-N4-Ben zoyl-5′-O-DMT-2′-d eoxycytid ylyl-(3′,5′)-N4-
b en zoyl-3′-O-a cet yl-2′-d eoxycyt id in e 3′-m et h a n ep h os-
p h on a te (3) was obtained from (SP )-4′ (R ) DMT, B ) CBz
)
and 3′-O-acetyl-N4-benzoyl-2′-deoxycytidine: yield 67%; 31P
1
NMR δ 33.06; H NMR δ 1.66 (d, 17.5, 3H, PCH3); FAB-MS
Gen er a l P r oced u r e for P r ep a r a tion of Din u cleosid e
(3′,5′)-Meth a n ep h osp h on a tes (3). The corresponding dias-
tereomerically pure 4′ or 5 (0.3 mmol), and 3′-O-acetyl (N-
protected)-2′- deoxynucleoside (0.1 mmol) were coevaporated
twice with dry pyridine (5 mL) and left overnight under high
vacuum. LiCl (freshly dried at 150 °C/0.1 mmHg, 0.125 g, 3
mmol) was added, and the resulting mixture was dissolved in
dry acetonitrile (5 mL). To this solution was added DBU (0.456
g, 3 mmol) in acetonitrile (1.5 mL) in one portion. After the
reaction was completed (2 h; disappearance of 4′ or 5 was
followed by HP-TLC), the solvent was evaporated, and the oily
residue was added dropwise to cold hexane. The solid pre-
cipitate was collected by centrifugation and redissolved in
CHCl3 and the solution extracted twice with citric acid (0.05
M). The combined organic layers were dried over anhyd
MgSO4 and concentrated, and crude 3 was purified by column
chromatography. The appropriate fractions, eluted with a
gradient of CHCl3-EtOH (3-10% EtOH), were collected,
combined, and concentrated under reduced pressure.
(RP )-5′-O-DMT-t h ym id ylyl-(3′,5′)-3′-O-a cet ylt h ym i-
d in e 3′-m eth a n ep h osp h on a te (3) was obtained from (RP )-5
(R ) DMT, B ) Thy) and 3′-O-acetylthymidine: yield 92%;
31P NMR δ 33.00; 1H NMR δ 1.58 (d, 17.6, 3H, PCH3); FAB-MS
found [M - H] 887.3 (calcd 887.290).
found [M - H] 1067.
Ack n ow led gm en t. These results were obtained
within the project financially assisted by the State
Committee for Scientific Research (KBN, Grant No. 3
TO9A 031 011) and, in part, by FIRCA- PA-91-77 Grant
to Professor L. J en-J acobson (Principal Investigator,
University of Pittsburgh, PA). We are indebted to Dr.
Marek Sochacki for recording the HRMS, Mr. Sławomir
Kaz´mierski for the 500 MHz spectra, and Mrs.Teresa
Ke¸błowska for skillful technical assistance.
Su p p or tin g In for m a tion Ava ila ble: X-ray crystallo-
graphic data for compounds listed in Table 1, including tables
of bond lengths, bond angles, atomic coordinates, temperature
factors, bond distances, angles, intermolecular contacts, and
hydrogen-bond parameters, five additional figures (stereoviews
of accurate positions of molecules in the elemental cells and
the Newmann projections), and disscussion (31 pages). This
material is contained in 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.
(SP )-5′-O-DMT-t h ym id ylyl-(3′,5′)-3′-O-a cet ylt h ym i-
d in e 3′-m eth a n ep h osp h on a te (3) was obtained from (SP )-5
J O980225G