Oligonucleotide Analogues with a Nucleobase-Including Backbone: Part 6. 2-Deoxy-D-erythrose-Derived Phosphoramidites: Synthesis and Incorporation into 14-Mer DNA Strands

Article abstract of DOI:10.1002/1522-2675(20010516)84:5<1000::AID-HLCA1000>3.0.CO;2-S

Two modified DNA 14-mers have been prepared, containing either a 2-deoxy-D-erythrose-derived adenosine analogue carrying a C(8)-CH2O group (deA*), or a 2-deoxy-D-erythrose-derived uridine analogue, possessing a C(6)-CH2O groupp (deU*). These nucleosides are linked via a phosphinato group between O-C(3')(deA* and deU*) and O-C(5') of one neighbouring nucleotide, and between C(8)-CH2O (deA*), or C(6)-CH2O (deU*) and O-C(3') of the second neighbour. N⁶-Benzoyl-9-(β-D-eryhrofuranosyl)adenine (3) and 1-(β-D-erythrofuranosyl)uracil (4) were prepared from D-glucose, deoxygenated at C(2'), and converted into the required phosphoramidites 1 and 2. The modified tetradecamers 31 and 32 were prepared by solid-phase synthesis. Pairing studies show a decrease in the melting temperature of 7 to 8 degrees for the duplexes 31*30 and 32*29, as compared to the unmodified DNA duplex 29*30. A comparison with the pairing properties of tetradecamers similarly incorporating deoxyribose- instead of the deoxyerythrose-derived nucleotides evidences that the CH2OH substituent at C(4') has no significant effect on the pairing.

Full text of DOI:10.1002/1522-2675(20010516)84:5<1000::AID-HLCA1000>3.0.CO;2-S

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Helvetica Chimica Acta ± Vol. 84 (2001)
Oligonucleotide Analogues with a Nucleobase-Including Backbone:
Part 6
2-Deoxy-d-erythrose-Derived Phosphoramidites: Synthesis and Incorporation
into 14-Mer DNA Strands
by Werngard Czechtizky and Andrea Vasella*
Laboratorium für Organische Chemie, ETH-Zentrum, Universitätstrasse 16, CH-8092 Zürich
Two modified DNA 14-mers have been prepared, containing either a 2-deoxy-d-erythrose-derived
adenosine analogue carrying a C(8) CH2O group (deA*), or a 2-deoxy-d-erythrose-derived uridine analogue,
possessing a C(6) CH2O group (deU*). These nucleosides are linked via a phosphinato group between
O
C(3') (deA* and deU*) and O C(5') of one neighbouring nucleotide, and between C(8) CH2O (deA*), or
C(6) CH2O (deU*) and O C(3') of the second neighbour. N6-Benzoyl-9-(b-d-erythrofuranosyl)adenine (3)
and 1-(b-d-erythrofuranosyl)uracil (4) were prepared from d-glucose, deoxygenated at C(2'), and converted
into the required phosphoramidites 1 and 2. The modified tetradecamers 31 and 32 were prepared by solid-
phase synthesis. Pairing studies show a decrease in the melting temperature of 7 to 8 degrees for the duplexes
31 ´ 30 and 32 ´ 29, as compared to the unmodified DNA duplex 29 ´ 30. A comparison with the pairing properties
of tetradecamers similarly incorporating deoxyribose- instead of the deoxyerythrose-derived nucleotides
evidences that the CH2OH substituent at C(4') has no significant effect on the pairing.
Introduction. ± Oligonucleotide analogues with a nucleobase-including backbone
should allow to answer the question whether the structural differentiation between
nucleobase and backbone in DNA, RNA, and their analogues is a necessary
prerequisite for the formation of stable homo- and/or heteroduplexes [1 ± 5]. We have
already described oligonucleotide analogues with a phosphinato group between
*
O
C(3') and either a C(8) CH2O group of 2'-deoxyadenosine (dAn 1); Fig. 1), or a
1
*
C(6) CH2O group of 2'-deoxyuridine (dUn ); Fig. 1) [5]. Force-field calculations [6]
and Maruzen model studies suggested that these analogues may form autonomous
pairing systems, and that the incorporation of single modified dA* or dU* units into
DNA 14-mers is compatible with duplex formation. Pairing studies of singly modified
tetradecamers, however, showed that the introduction of dA* or dU* units into a 14-
mer DNA duplex lowered the melting temperature by 6 ± 78, as compared to the
unmodified DNA duplex [5]. This destabilization may be due to a distortion of the
DNA duplex caused by the syn-conformation [7] of the dA* or dU* units, as enforced
by an unfavourable interaction between the C(8)- or C(6)-substituent and the
carbohydrate moiety ± presumably the O C(5') group ± of dA* and dU*. To reduce
this steric interaction, we decided to remove the CH2OH substituent at C(4') of dA*
1
)
The repeating units of B-DNA are denoted dA, dT, dG, and dC. 2-Deoxyribose-derived units linked via
C(3') and O(10), or O(7) are denoted dA* and dU*, respectively, and 2-deoxy-d-erythrose-derived
units linked via O C(3') and O(10), or O(7), deA* and deU*, respectively (Fig. 1).
O

Helvetica Chimica Acta ± Vol. 84 (2001)
1001
and dU*, and to introduce the 2-deoxyerythrose-derived units deA*1) and deU*1)
(Fig. 1) into a DNA duplex.
NH2
N
N
CH2OH
O
N
N
HO
O
HN
O
CH2OH
NH2
HO
O
O
O
N
O
N
N
N
O
N
P
HN
O
N
O
O
HO
O
O
O
O
HO
O
P
O
NH2
O
N
O
N
N
O
N
HN
P
O
N
O
O
O
O
HO
O
P
HO
O
O
n–1
n–1
dU*n
HO
dA*n
HO
NH2
O
N
NH2
O
N
H3C
N
N
N
O
N
NH
8
N
HN
6
O
10
N
O
N
9
5'
O 5'
O
N
O
O
O
HO
HO
3'
1
1'
7
O
O
1'
2'
3'
3'
3'
2'
O
O
O
O
O
O
O
O
P
P
P
P
O
O
O
O
dA
dT
dA*
dU*
NHBz
O
N
NH2
O
N
N
N
N
O
HN
8
HN
N
6
O
ODMTr
CN
ODMTr
N
N
10
O
O
O
O
9
N
O
N
1
1'
2'
7
O
1'
2'
3'
3'
O
O
O
O
O
O
CN
P
P
P
O
P
O
O
iPr2N
iPr2N
O
deA*
deU*
1
2
*
*
Fig. 1. 2-Deoxyribose-derived oligonucleotide analogues dAn and dUn , DNA units dA and dT, 2-deoxyribose-
derived units dA* and dU*, 2-deoxyerythrose-derived units deA* and deU*, and the phosphoramidites 1 and 2
We report on the syntheses of the deA*- and deU*-derived phosphoramidites 1 and
2 (Fig. 1), the incorporation of a single deA*, or deU* unit into a tetradecamer, and on
the pairing of these tetradecamers with an unmodified complementary strand.
Results and Discussion. ± 1. Synthesis of the Phosphoramidites 1 and 2.
Nucleosidation of 2-deoxy-d-erythrose derivatives with thymine [8] and uracil [9]
proceeds with a low selectivity, affording 2 :1 to 1:1 mixtures of anomers, while
nucleosidation of anomeric mixtures of triacetyl erythrofuranose with N6-benzoylade-
nine or uracil afforded diastereoselectively the b-d-nucleosides 3 and 4 (Schemes 1 and
2) after deprotection [10]. For this reason, we decided to prepare the phosphoramidites

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Helvetica Chimica Acta ± Vol. 84 (2001)
1 and 2 from the 1,2,3-tri-O-acetyl-a,b-d-erythroses 5/6 (Scheme 1) rather than from 2-
deoxy-d-erythrose, and to subsequently remove the OH group at C(2'). Triacetyl-
erythrose is readily accessible by degradation of d-glucose with Pb(OAc)4 [11][12] and
acetylation [13]2). Deoxygenation of 3 and 4, followed by hydroxymethylation,
protection, and phosphitylation, should lead to the phosphoramidites 1 and 2.
Scheme 1
NHBz
NHBz
N
NHBz
N
N
N
N
N
N
N
N
R1
R2
N
N
N
a)
c)
O
O
O
O
iPr3SiO OH
HO OSiiPr3
OAc
AcO
RO OR
7 R = Ac
3 R = H
5 R1 = H, R2 = AcO
6 R1 = AcO, R2 = H
8
9
d)
b)
1:1
2:1
e)
NHBz
NHBz
NHBz
NHBz
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
O
N
O
f)
O
O
OSiiPr3
iPr3SiO
O
OR
OR
O
S
S
12 R = iPr3Si
14 R = H
10
11
13 R = iPr3Si
15 R = H
N
g)
N
g)
N
N
NHBz
NHBz
N
NHBz
N
NHBz
N
N
N
N
N
N
N
ODMTr
CN
ODMTr
j)
OH
N
N
N
N
N
N
N
h)
i)
O
O
O
O
O
iPr3SiO
OR
OR
P
O
18 R = iPr3Si
19 R = H
16 R = iPr3Si
17 R = H
iPr2N
12
g)
g)
1
a) 1. N6-Benzoyladenine, N,O-bis(trimethylsilyl)acetamide (BSA); 2. SnCl4; 80%. b) 1. NaOH, H2O/MeOH/
i
THF 5 :4 :1; 2. NH4Cl; 80%. c) Pr3SiOTf, pyridine, DMF; 92%. d) SiO2, Et3N. e) N,N-(thiocarbonyl)diimi-
dazole, CH2Cl2. f) Bu3SnH, 2,2'-azobis(isobutyronitrile) (AIBN), toluene; 44% of 12 and 20% of 13 (from 8/9
2 :1). g) Bu4NF ´ 3 H2O, THF; 14 (74%); 15 (89%); 19 (76%). h) 1. LDA/THF; 2. DMF; 3. AcOH; 4. NaBH4/
EtOH; 88%. i) Dimethoxytrityl chloride (ˆ bis(4-methoxyphenyl)(phenyl)methyl chloride; DMTrCl),
EtNiPr2, 4-(dimethylamino)pyridine (DMAP), CH2Cl2; 89%. j) (NCCH2CH2O)P(iPr2N)Cl, CH2Cl2, EtNiPr2;
80% of 2 diastereoisomers (1 :1).
2
)
2-Deoxy-d-erythrose has been prepared from 3-deoxy-d-erythropentose [14], dimethyl (S)-malate [15], d-
isoascorbic acid [16], and (S)-solketal [16].

Helvetica Chimica Acta ± Vol. 84 (2001)
1003
Fig. 2. Crystal structure of 3 ´ MeOH
Nucleosidation of the anomeric mixture 5/6 with N6-benzoyladenine in the presence
of SnCl4 and N,O-bis(trimethylsilyl)acetamide (BSA) at 608 (cf. [17][18]) gave the b-d-
configured nucleoside 7 (80%), which was deacetylated with 2m NaOH to yield 80% of
the nucleoside 3. Stannylation of 3 with Bu2SnO in toluene [19], followed by treatment
of the crude Sn-acetal with Et3SiCl, led to a 1:1 mixture of regioisomeric silyl ethers.
Regioselective silylation of 3 at HO C(3') with various trialkylchlorosilanes in the
presence of Ag salts and DABCO as reported for ribonucleosides [20] led again to
unsatisfactory results. Finally, we treated 3 with a 2.5-fold excess of iPr3SiOTf in DMF/
pyridine 16 :1 at 238 to obtain 92% of a 1:1 mixture of the 3'- and 2'-O-iPr3Si-protected
nucleosides 8 and 9. Treatment of this mixture with silica gel in Et3N led to partial
i
migration of the Pr3Si group from O(2') to O(3'), resulting in a 2 :1 mixture 8/9 after
24 h. This mixture was converted to the thiocarbamates 10/11 (2 :1). Treatment of the
crude mixture with Bu3SnH and 2,2'-azobis(isobutyronitrile) (AIBN) in refluxing
toluene yielded a mixture 12/13 (2 :1; 64% from 8/9). The nucleosides 12/13 were
readily separated by FC. Desilylation of 12 and 13 yielded 14 and 15 (80% each),
respectively. Deprotonation of 12 by LDA [21], addition of DMF, and reduction with
NaBH4 gave 88% of the C(8)-CH2OH nucleoside 16 that was desilylated to 17 (80%).
Dimethoxytritylation of 16 gave 18 (89%) that was desilylated to 19 (80%). The
alcohol 19 was converted to the phosphoramidite 1 in the usual way [22]; 1 is stable at
158 for several months.
The configuration of 3 was established by X-ray crystal-structure analysis of 3 ´
MeOH3) (Fig. 2). The erythrofuranosyl moiety adopts the 2T3 conformation, similarly
to the conformation deduced for 3 in D2O solution [10]. HO C(3') forms a weak
intramolecular H-bond to O C(2') (distance O(2') ´´´ O(3') 2.77 Š); intermolecular
H-bonds are found between HO C(3') and N(1), H N(6) and O C(2'), MeOH and
3
)
The crystallographic data have been deposited with the Cambridge Crystallographic Data Centre
(deposition No. CCDC-158435 (3), CCDC-158436 (4), and CCDC-158437 (25)). Copies of the data can
be obtained, free of charge, on application to the CCDC, 12 Union Road, Cambridge, CB21EZ, UK (fax:
‡44(1223)336033; e-mail: deposit@ccdc.cam.ac.uk).

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Helvetica Chimica Acta ± Vol. 84 (2001)
N(7), and between HO C(2') and MeOH. The nucleobase adopts the syn-conforma-
tion.
The structures of 8 and 9 were assigned on the basis of the 1H-NMR spectra of 8/9
(2 :1). The signals of H C(2') and H C(3') of 8/9 were assigned on the basis of their
multiplicity and coupling constants (see Table 3). In CDCl3, H C(2') of the major
compound appears as a dt with J(2',3') ˆ 5.0, J(1',2') ˆ 4.1, and J(H,OH) ˆ 5.3 Hz.
Upon addition of D2O, this signal collapsed to a dd, while the H C(3') q remained
unchanged, evidencing that the major compound possesses structure 8. The broad
H
C(3') signal of the minor compound (9) collapsed to a ddd upon addition of D2O,
while the H C(2') t remained unchanged. The structure determination was corrobo-
1
rated by the H-NMR spectra of a 2 :1 mixture of the thiocarbamates 10/11. The
H
C(2') and H C(3') signals of 10/11 were assigned on the basis of their multiplicity
and coupling constants (see Table 3). Carbamoylation of O C(2') of the major
compound 8, leading to 10, induced a 1.6-ppm downfield shift for H C(2'), while
carbamoylation of O C(3') of the minor compound 9, leading to 11, resulted in a 1.67-
ppm downfield shift for H C(3'). As expected, H C(1') of 12 appears as a t at
6.46 ppm with J(1',2') ˆ 6.6 Hz (see Table 3 in Exper. Part). The Hpro-S C(2') of 12 gives
rise to a ddd at 2.97 ppm, while Hpro-R C(2') appears at 2.60 ppm as a dddd (W coupling
to H C(4'), J(2',4') ˆ 1.0 Hz). The signals of H C(1'), H C(2'), H C(3'), and
H
C(4') of 13 were assigned on the basis of irradiation experiments. H C(1') of 13
resonates at 6.00 ppm as a d, J(1',2') ˆ 1.6 Hz, Hpro-S C(3') of 13 at 2.32 ppm as a dtd,
and Hpro-R C(3') at 2.12 ppm as a dddd. As a rule, the shift from the anti- to the syn-
conformation is indicated by a downfield shift for Hpro-S C(2') and an upfield shift for
C(2') [23]. The significant downfield shifts for Hpro-S C(2') (Dd ˆ 0.18 ± 0.30 ppm) and
the upfield shifts for C(2') (Dd ˆ 1.1 to 2.6 ppm) of the nucleosides 16, 18, 19, and
the phosphoramidite 1, as compared to the corresponding signals of 12, evidence that
all C(8)-substituted nucleosides adopt predominantly the syn-conformation (Table 1).
The ratio J(1',2'pro-S)/J(3'pro-R,4') is considered a measure of the position of the
equilibrium between the S- (sugar-pucker E ± 2T1) and N-conformers (sugar-pucker
3
3T2 ± 3T4) [24]. The J(1',2'pro-S) (6.6 Hz) and J(3',4') (X2.1 Hz) values (Table 1) evidence
a predominance of the S-conformer of the nucleosides 1, 12, 14, 16, 18, and 19.
Table 1. 1H- and 13C-NMR Shift Differences Dd(H C(2')), and Dd(C(2')) [ppm] for the Nucleosides 16, 18, 19,
and 1 as Compared to 12, and for 24, 26 ± 28, and for 2 as Compared to 24, and Ratio of S/N-Conformers of these
Nucleosides, as Deduced from J(1',2')/J(3',4') [Hz]
Nucleoside
Dd(H C(2'pro-S ))
Dd(C(2'))
J(1',2')
J(3',4')
Ratio S/N
12
16
18
19
1
24
26
27
28
2
±
±
6.6
6.6
6.6
6.6
6.6
6.5
6.6
7.0
7.0
6.5
2.1
< 1.0
< 1.0
< 1.0
< 1.0
2.2
< 1.0
< 1.0
< 1.0
< 1.0
76/24
> 87/13
> 87/13
> 87/13
> 87/13
75/25
> 87/13
> 88/12
> 88/12
> 87/13
0.18
0.23
0.23
0.30
±
0.79
0.74
0.71
0.77
1.1
1.1
1.8
2.6
±
2.0
2.1
3.1
4.1

Helvetica Chimica Acta ± Vol. 84 (2001)
1005
To prepare the phosphoramidite 2 (Scheme 2), the anomeric mixture 5/6 was
treated with uracil in the presence of BSA and SnCl4 in MeCN at 608 (cf. [17][18]). We
obtained 80% of the b-d-configured nucleoside 20, which was deacetylated with 2m
NaOH to yield 80% of 4. To deoxygenate 4 at C(2'), we planned to convert 4 to the
corresponding 2,2'-anhydronucleoside [25], to cleave the C(2') O bond with LiBr and
BF3 ´ Et2O [26], and to reductively debrominate the resulting 2'-bromo-2'-deoxynu-
cleoside. The 2,2'-anhydro-1-(b-d-threofuranosyl)uracil (21) was prepared from 4 by
treatment with diphenyl carbonate and NaHCO3 in DMF [25]. However, all attempts
to convert 21 to the 2'-deoxynucleoside 25 failed. Silylation of the 2,2'-anhydronucleo-
side 21 with a 5-fold excess of iPr3SiOTf yielded 22 (61% from 4). Treatment of 22 with
LiBr and BF3 ´ Et2O gave the 2'-bromo-2'-deoxynucleoside 23 (88%) that was
debrominated in 97% yield with Bu3SnH and AIBN. The resulting 2'-deoxynucleoside
Scheme 2
O
O
N
NH
N
O
O
N
R1
R2
c)
a)
O
O
O
HO
AcO OAc
RO OR
5 R1 = H, R2 = AcO
6 R1 = AcO, R2 = H
20 R = Ac
4 R = H
21
b)
d)
O
O
N
O
NH
N
NH
O
N
O
O
O
N
f)
e)
O
O
iPr3SiO Br
iPr3SiO
RO
24 R = iPr3Si
22
23
g)
25 R = H
h)
O
O
O
N
HN
HN
HN
ODMTr
OH
ODMTr
j)
O
N
O
O
N
i)
O
O
O
O
iPr3SiO
iPr3SiO
CN
P
O
27 R = iPr3Si
28 R = H
iPr2N
26
g)
2
a) 1. Uracil, N,O-bis(trimethylsilyl)acetamide (BSA); 2. SnCl4; 80%. b) 1. NaOH, H2O/MeOH/THF 5 :4 :1; 2.
NH4Cl; 80%. c) 1. Diphenyl carbonate, DMF; 2. NaHCO3. d) iPr3SiOTf, pyridine, DMF; 62% (from 4). e) LiBr,
BF3 ´ Et2O, 1,4-dioxane; 88%. f) Bu3SnH, AIBN, toluene; >97%. g) Bu4NF ´ 3 H2O, THF; 25 (88%), 28 (96%).
i
h) 1. LDA/THF; 2. DMF; 3. AcOH; 4. NaBH4/EtOH; 51%. i) DMTrCl, Pr2NEt, DMAP, CH2Cl2; 77%.
j) (NCCH2CH2O)P(iPr2N)Cl, CH2Cl2, iPr2NEt; 69% of 2 diastereoisomers (2 :1).

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Helvetica Chimica Acta ± Vol. 84 (2001)
24 was desilylated to 25 (80%). Hydroxymethylation of 24 to 26 was accomplished in
52% yield similarly as described above for 12. Dimethoxytritylation of 26 to 27 and
desilylation of 27 yielded 73% of the alcohol 28, which was converted to the
phosphoramidite 2 in the usual way (70%); at 158, 2 is stable for several months.
The b-d-configuration of 1-(b-d-erythrofuranosyl)uracil (4) was established by
X-ray crystal-structure analysis (Fig. 3,a).
Fig. 3. Crystal structures of a) 4 and b) 25
Similarly to N6-benzoyl-9-(b-d-erythrofuranosyl)adenine (3), the erythrofuranosyl
moiety of 4 adopts the 3E-conformation. There is a (weak?) intramolecular H-bond be-
tween HO C(3') and O C(2') (distance O(2') ´´´ O(3') 2.69 Š), while intermolecular
H-bonds are found between HO C(2') and O C(2'), HO C(3') and O C(4'), and
H
N(3) and O C(4'). Unlike 3, the uracil derivative 4 adopts the anti-conformation.
The H-atoms H C(1'), H C(2'), H C(3'), H C(5), and H C(6) of the 2,2'-
anhydronucleoside 21 (Table 5 in Exper. Part) resonate at almost the same field as the
corresponding signals of 2,2'-anhydrouridine (Dd ˆÆ 0.04 ppm) [27]. As observed for
5'-protected 2,2'-anhydrouridines [28][29], C(2), C(4), C(5), C(1'), and C(2') of 21 are
shifted significantly downfield, as compared to 4: C(4) and C(2) give rise to two s at
178.7 and 164.2 ppm (4: two s at 163.5 and 151.0 ppm), C(5) resonates as a d at
112.1 ppm (4: d at 102 ppm), and C(1') and C(2') as two d at 93.4 and 90.6 ppm (4: two d
at 88.8 and 74.1 ppm). Bromination of 22 to 23 is indicated by a strong upfield shift for
‡
C(2') of 23 (d ˆ 52.4 as compared to d ˆ 74.1 ppm in 4), and the [M ‡ H] peak in the
FAB-MS of 23 shows the typical isotope pattern for Br compounds.
As expected, H C(1') of 24 appears at 6.15 ppm as a t with J(1',2') ˆ 6.5 Hz. The
Hpro-R C(2') of 24 gives rise to a ddd at 2.50, and Hpro-S C(2') to a dt at 2.11 ppm. The
ratio J(1',2')/J(3',4') indicates that the 2-deoxy-d-erythrofuranosyl moiety of 24 adopts
predominantly the S-conformation (Table 1).
The structure of 25 was established by X-ray crystal-structure analysis (Fig. 3,b).
Two symmetrically independent molecules in the elementary cell form a dimer with
intermolecular H-bonds between H N(3) and O C(4), and between H N(3) and
2
O
C(2). Both independent molecules adopt the T3- and the anti-conformations.

Helvetica Chimica Acta ± Vol. 84 (2001)
1007
Similarly to the adenine series, the downfield shift for Hpro-S C(2') (Dd ˆ 0.71 ±
0.79 ppm) and the upfield shift for C(2') of 2 and 26 ± 28, as compared to 24 (Dd ˆ 2.0
to
4.1 ppm), show the predominance of the syn-conformation for the C(6)-
substituted nucleosides (Table 1). The equilibrium between the S- and the N-
conformers of 2 and 26 ± 28 is clearly in favour of the S-conformer (> 88/12), as
indicated by J(1',2'pro-S) and J(3',4') values of 2 and 26 ± 28 (Table 1).
2. Solid-Phase Synthesis of 14-Mer DNA Strands Containing deA* or deU*, and of
the Complementary DNA Strands. Table 2 shows the 14-mer DNA strands 29 and 30
possessing a random sequence with an internal dA or dTunit flanked by dG or dC units.
Replacement of the internal dA of 29 by deA* (derived from 1) led to the modified 14-
mer 31, while replacement of the internal dTof 30 by deU* (from 2) led to the modified
14-mer 32. The solid-phase synthesis of the oligomers 29 ± 32 was carried out on a
DNA-synthesizer using essentially the protocol for the synthesis of pRNA [30]. The
coupling time for the attachment of 1, 2, and the DNA phosphoramidites following the
incorporation of 1 and 2 was arbitrarily set to 30 min; it led to coupling yields of 82%4)
for 1 and ca. 80 ± 90% for the DNA-phosphoramidites following 1, and of ca. 90% for 2
(99% for all other couplings). Longer coupling times did not improve yields. The DNA
14-mers 29 and 30 were deprotected and cleaved off the solid support by ammonolysis
with saturated aqueous NH3/MeOH 1:1 at 508 within 20 h, while 31 was deprotected
under the same conditions within 3.5 h. Prolonged heating of the modified DNA strand
32 (incorporating deU*) at 508 led to degradation; this strand was deprotected at 358.
The crude products were analyzed by RP-HPLC. The DNA strands 29 and 30 were
pure, the modified tetradecamer 32 was ca. 70 ± 80% pure, while crude 31 contained
three major products A, B, and C in a ca. 1:2 :1 ratio. The products were purified by
RP-HPLC, desalted, and analyzed by RP-HPLC and MALDI-TOF mass spectroscopy;
only fraction C (from crude 31) exhibited a molecular ion in agreement with structure
315). The yield of the oligomers, as determined from the detritylation assay, and their
molecular masses are shown in Table 2.
Table 2. Total Yields, and Calculated and Experimental Molecular Masses of the Oligodeoxynucleotides 29 ± 32
Sequence
Total yield [%]a)
Calc. mass
Exper. mass (MALDI-TOF)
29 5'-d(CGTAAGCTCGATAG)-3'
30 5'-d(CTATCGAGCTTACG)-3'
31 5'-d(CTATCGeA*GCTTACG)-3'
32 5'-d(CGTAAGCeU*CGATAG)-3'
94
99
27
75
4287.8
4238.8
4238.8
4273.8
4287.4
4239.1
4237.7
4276.6
a) Calculated from the yield of the coupling steps, as determined by the detritylation assay.
3. Pairing Studies. The modified 14-mer 31 (Table 2) was hybridized with the
complementary 14-mer 30 at a concentration of (2 ‡ 2) mm and at pH 7 (10 mm
NaH2PO4/Na2HPO4 in H2O) in 0.1m aqueous NaCl. The modified duplex 31 ´ 30
4
)
Each chain elongation cycle began with an acid promoted cleavage of the dimethoxytrityl group. The
‡
efficiency of each coupling step was calculated by UV-determination of the amount of DMTr (e ˆ 70000 l ´
mol 1 ´ cm 1).
5
)
In agreement with this assignment, the fractions A and B did not pair with 30.

1008
Helvetica Chimica Acta ± Vol. 84 (2001)
showed a decrease of the melting temperature by 88, as compared to the non-modified
duplex 29 ´ 30 (see Fig. 4 for the melting curves).
Fig. 4. Temperature-dependent UV spectra (ꢀmelting curvesꢁ) of the duplexes 29 ´ 30, 31 ´ 30, and 32 ´ 29
Similarly, the modified 14-mer 32 (Table 2) incorporating one deU* unit was
hybridized with the complementary 14-mer 29 at a concentration of (2 ‡ 2) mm and at
pH 7 (10 mm NaH2PO4/Na2HPO4 in H2O) in 0.1m aqueous NaCl. Again, the modified
duplex 32 ´ 29 melted 78 lower than the non-modified DNA duplex 29 ´ 30 (see Fig. 4 for
the melting curves). Since the modified 14-mer 32 is degraded during prolonged
heating, the melting process for the duplex 32 ´ 29 was irreversible.
Similarly as reported for the incorporation of the 2-deoxyribose-derived units dA*
and dU* [5], the introduction of the 2-deoxy-d-erythrose-derived units deA* and deU*
into a DNA duplex causes a decrease in melting temperature by 7 ± 88, as compared to
the unmodified duplex. A comparison of the duplex destabilization imposed by dA*,
dU*, deA*, and deU* shows that this destabilization is independent of the CH2OH
substituent at C(4').
We thank Dr. B. Bernet for checking the experimental part, and the Swiss National Science Foundation and
F. Hoffmann-La Roche AG, Basel for generous support.
Experimental Part
General. Solvents were distilled before use: THF from K/benzophenone, toluene from Na, CH2Cl2, DMF,
pyridine, 1,4-dioxane, and EtNiPr2 from CaH2. Reactions were run under Ar. Qual. TLC: precoated silica-gel
plates (Merck silica gel 60 F254); detection by spraying with ꢀmostainꢁ (400 ml of 10% aq. H2SO4, 20 g of
(NH4)6Mo7O24 ´ H2O, 0.4 g of Ce(SO4)2) and heating. Flash chromatography (FC): silica gel Merck 60 (0.04 ±
0.063 mm). Reversed-Phase High-Performance Liquid Chromatography (RP-HPLC): analytical RP column:
Macherey-Nagel; LiChroCartꢂ250-4 HPLC Cartridge filled with LiChrospherꢂ100, RP-18e (5 mm); prep. RP
column: Macherey-Nagel, Hibar ꢂ RT250-10, customized packing: LiChrospherꢂ100, RP-18e (5 mm), eluents:
0.1m aq. Et3NHOAc, MeCN, H2O. Optical rotations: 1-dm cell at 258 and 589 nm. FT-IR: 1 ± 2% soln. in the
indicated solvent. 1H-, 13C-, and 31P-NMR: at 200, 300, or 400 MHz, 50 or 75 MHz, or 121 MHz, resp. MS: fast-
atom bombardment (FAB), matrix-assisted laser-desorption ionization (MALDI), high-resolution (HR).
NOBA: 3-nitrobenzyl alcohol.
N6-Benzoyl-9-(2',3'-di-O-acetyl-b-d-erythrofuranosyl)adenine (7). A suspension of 5/6 (1:4) [11 ± 13]
(4.16 g, 16.9 mmol) and N6-benzoyladenine (4.4 g, 18.4 mmol) in MeCN (18 ml) was treated dropwise with N,O-
bis(trimethylsilyl)acetamide (10 ml, 41 mmol) at 608. The mixture was stirred for 30 min, treated with SnCl4

Helvetica Chimica Acta ± Vol. 84 (2001)
1009
(8 ml, 68 mmol), stirred for 20 min, poured into a stirred mixture of AcOEt (300 ml) and sat. aq. NaHCO3
(300 ml), and stirred for 15 min. The org. layer was dried (Na2SO4) and evaporated. FC (AcOEt/MeOH 100 :1)
gave 7 (5.7 g, 80%). Colourless powder. Rf(AcOEt/hexane 6 :1) 0.40. [a]D25
ˆ
90.1 (c ˆ 0.75, CHCl3). IR
(CHCl3): 3375w (br.), 3007m, 1750s, 1709s, 1612s, 1587s, 1504m, 1480s, 1469s, 1374s, 1072s. 1H-NMR (200 MHz,
CDCl3): see Table 3; additionally, 9.22 (br. s, NH); 8.05 ± 7.95 (m, 2 arom. H); 7.63 ± 7.40 (m, 3 arom. H); 2.16
(s, AcO); 2.05 (s, AcO). 13C-NMR (50 MHz, CDCl3): see Table 4; additionally, 170.0, 169.5 (2s, 2 O CˆO);
164.9 (s, N CˆO); 133.0 (s); 132.9 (d); 128.9 (d, 2 C); 128.0 (d, 2 C); 20.7 (q, Me); 20.4 (q, Me). FAB-MS
‡
‡
‡
(NOBA): 426 (69, M ), 427 (25, [M ‡ H] ). HR-MALDI-MS: 448.123 (C20H19N5NaO6, [M ‡ Na] ; calc.
448.123).
Table 3. Selected 1H-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] for the Nucleosides 7 ± 13, 16 ±
19, and 1 in CDCl3, for 3 in (D6)DMSO, and for 14 and 15 in CD3OD
7
3
8
9
10
11
12
13
H
H
H
C(1')
C(2'pro-S
C(2'pro-R
6.1 ± 6.2
6.1 ± 6.2
±
5.74
±
5.97
4.86
±
4.28
±
4.40
3.84
8.75
8.68
6.6
±
5.94
4.83
±
5.00
±
4.50
4.00
8.76
8.08
4.2
±
5.90
5.56
±
4.45
±
4.58
4.15
8.80
8.00
5.3
±
6.33
6.42
±
5.47
±
4.62
4.00
8.71
8.35
4.1
±
6.00
5.90
±
6.12
±
4.88
4.37
8.76
8.45
6.2
±
6.46
2.97
2.60
4.93
±
4.43
4.03
8.82
8.13
6.6
6.00
5.10
±
2.32
2.12
4.42
4.32
8.80
8.05
1.6
)
)
Ha C(3')
Hb C(3')
Ha C(4')
Hb C(4')
H
H
4.73
4.17
8.75
8.10
C(2)
C(8)
a
J(1',2')
J(2',2')
)
±
13.7
±
J(2'pro-S,3')
J(2'pro-R,3')
J(3',3')
5.1
±
±
4.1
±
±
5.0
±
±
5.0
±
±
4.4
±
±
4.4
±
±
6.6
2.5
±
4.4
2.4
13.0
J(3',4'a)
J(3',4'b)
J(4'a,4'b)
4.1
2.1
10.4
3.7
1.2
9.1
5.0
4.1
9.3
3.4
< 1.0
10.0
5.0
5.0
9.1
3.0
< 1.0
11.5
4.0
2.1
9.1
8.4, 4.0
8.7, 6.2
8.4
14
15
16
17
18
19
1
H
H
H
C(1')
C(2'pro-S
C(2'pro-R
6.57
6.10
4.84
±
2.40
2.10
4.42
4.28
±
6.50
3.15
2.39
4.95
±
4.36
3.89
4.90
4.90
8.68
±
6.68
6.47
3.20
2.43
4.94
±
4.55
3.92
4.55
4.40
8.80
±
6.37
6.46
3.27
2.60, 2.51
4.85
±
)
)
2.95
2.58
4.77
±
4.42
3.96
±
3.25
2.50
4.82
±
4.57
3.96
4.95
4.90
8.66
±
3.20
2.35
4.87
±
4.50
3.92
4.52
4.45
8.76
±
Ha C(3')
Hb C(3')
Ha C(4')
Hb C(4')
Ha C(10)
Hb C(10)
H
H
J(1',2')
4.53
4.08, 4.04
4.53, 4.50
4.45, 4.42
8.78
±
6.6
±
±
C(2)
C(8)
8.70
8.50
6.6
8.70
8.42
1.9
±
6.6
13.7
6.8
14.0
6.6
13.3
6.6
14.0
J(2',2')
14.0
13.3
J(2'pro-S,3')
J(2'pro-R,3')
J(3'a,3'b)
J(3',4'a)
J(3',4'b)
J(4'a,4'b)
J(10a,10b)
5.6
1.2
±
3.7
1.2
9.3
±
5.6
3.1
14.0
8.4, 4.3
8.4, 7.1
8.4
6.8
1.0
±
5.6
1.3
±
3.7
< 1.0
9.0
6.6
< 1.0
±
3.3
< 1.0
8.7
6.0
< 1.0
±
3.7
< 1.0
9.3
6.4
< 1.0
±
3.3
< 1.0
9.3
4.0
< 1.0
8.7
a
±
)
14.3
11.8
11.8
12.1
a) Not assigned.

1010
Helvetica Chimica Acta ± Vol. 84 (2001)
Table 4. Selected 13C-NMR Chemical Shifts [ppm] for 7 ± 13, 16 ± 19, and 1 in CDCl3, for 14 and 15 in CD3OD,
and for 3 in (D6)DMSO
C(1')
C(2')
C(3')
C(4')
C(10)
C(2)
C(4)
C(5)
C(6)
C(8)
7
3
8
87.3
88.0
91.4
91.2
87.7
91.0
86.2
92.9
87.2
93.8
85.6
87.2
86.1
85.7
85.9
85.8
74.2
74.5
75.2
75.9
83.4
74.3
41.7
76.4
41.7
76.7
40.6
40.7
40.6
39.9
39.2
39.0
71.6
70.4
71.7
71.9
70.4
82.4
72.6
33.8
73.0
33.8
73.0
73.3
73.2
72.7
74.4
74.2
72.7
74.0
74.7
74.7
74.4
72.0
77.3
69.3
78.0
70.5
77.6
78.2
77.8
77.4
76.5
76.2
±
±
±
±
±
±
±
±
151.8
151.9
152.9
153.2
152.6
152.7
152.8
152.5
153.0
153.0
154.8
156.5
152.0
152.0
152.2
152.1
149.9
150.6
150.2
150.2
150.3
150.3
149.9
149.6
151.3
151.0
149.3
150.5
149.5
149.4
149.7
149.6
124.1
126.3
124.1
124.1
124.2
124.2
124.2
123.8
125.6
125.6
122.0
124.2
122.5
122.5
122.7
152.9
152.6
151.5
151.5
151.7
151.7
151.8
151.2
142.1
144.0
142.5
143.1
142.3
143.0
142.2
141.3
9
10
11
12
13
14
15
16
17
18
19
1
±
±
153.1
152.8
144.8
144.0
57.8
58.3
59.7
59.5
59.9
153.0a)
154.5a)
152.7a)
152.6a)
152.7a)
152.4a)
152.9a)
152.6a)
152.5a)
152.6a)
a) Assignment may be interchanged.
N6-Benzoyl-9-(b-d-erythrofuranosyl)adenine ´ MeOH (3). At 238, a soln. of 7 (5.2 g, 12.2 mmol) in THF
(190 ml), MeOH (152 ml), and H2O (38 ml) was treated with 2m NaOH in H2O (25 ml, 50 mmol), stirred at 238
for 10 min, treated with NH4Cl (5.26 g, 98.3 mmol), and evaporated after addition of SiO2 (46 g). The resulting
powder was dried at 0.1 mbar for 14 h, filled in a column, and eluted with CHCl3/MeOH 10 :1. The colourless
crystals obtained from a few pooled fractions were submitted to X-ray analysis. Evaporation of the residual
fractions gave 3 (3.2 g, 78%). Colourless powder. Rf (CHCl3/MeOH 10 :1) 0.20. IR (KBr): 3407s (br.), 3237s
(br.), 1685s, 1608m, 1572s, 1518s, 1466m, 1428m, 1396m, 1325m, 1294m, 1248m, 1194m, 1146m, 1107m,
1070s,1025s. 1H-NMR (200 MHz, (D6)DMSO): see Table 3 ; additionally, 11.17 (br. s, NH); 8.06 ± 7.97
(m, 2 arom. H); 7.69 ± 7.47 (m, 3 arom. H); 5.56 (d, J ˆ 6.2, HO C(2')); 5.29 (d, J ˆ 3.7, HO C(3')); 4.01
(q, J ˆ 5.5, MeOH); 3.16 (d, J ˆ 5.5, MeOH). 13C-NMR (50 MHz, (D6)DMSO): see Table 4; additionally, 165.9
‡
(s, CˆO); 133.6 (s); 132.6 (d); 128.7 (d, 4 C); 48.6 (q, MeOH). FAB-MS (NOBA): 342 (100, [M ‡ H] ).
N6-Benzoyl-9-(3'-O-(triisopropylsilyl)-b-d-erythrofuranosyl)adenine (8) and N6-Benzoyl-9-(2'-O-(triiso-
propylsilyl)-b-d-erythrofuranosyl)adenine (9). A soln. of 3 (3.2 g, 9.4 mmol) in pyridine (4.1 ml, distilled from
i
CaH2) and DMF (64 ml, distilled from CaH2) was treated with Pr3SiOTf (3.2 ml, 12 mmol) at 238, stirred for
4 h, treated with iPr3SiOTf (3 ml, 11 mmol), stirred for 4 h at 238, and evaporated. FC (AcOEt/hexane 6 :1) gave
8/9 1:1 (4.3 g, 92%). A soln. of this mixture in 150 ml of Et3N was treated with SiO2 (60 g), stirred at 238 for 16 h,
and evaporated. FC (CH2Cl2/MeOH 10 :1) gave 8/9 2 :1. Colourless foam.
Data of 8/9: Rf(AcOEt/hexane 6 :1) 0.41. [a]D25
ˆ
58.4 (c ˆ 1.135, CHCl3, 8/9 2 :1). IR (CHCl3, 8/9 2 :1):
3411w (br.), 3003m, 2947s, 2893m, 2869s, 1708s, 1671m, 1612s, 1585s, 1503m, 1457s, 1387m, 1354m, 1328m,
1
1141m, 1112m. H-NMR (300 MHz, CDCl3, 8/9 2 :1): see Table 3; additionally, 9.13 (br. s, 0.33 H), 9.10 (br. s,
0.67 H) (NH); 8.05 ± 7.95 (m, 2 arom. H); 7.65 ± 7.40 (m, 3 arom. H); 3.45 (d, J ˆ 5.6, 0.67 H, exch., HO C(2') of
8); 3.02 ± 2.96 (m, 0.33 H, exch., HO C(3') von 9); 1.20 ± 0.80 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3,
8/9 2 :1): see Table 4; additionally, 165.9 (s, CˆO); 134.0 (s); 133.1 (d); 129.0 (d, 2 C); 128.4 (d, 2 C); 17.9 ± 17.5
‡
(4q, (Me2CH)3Si); 12.4 ± 12.0 (3d, (Me2CH)3Si). FAB-MS (NOBA, 8/9 2 :1): 498 (100, [M ‡ H] ). HR-
‡
‡
MALDI-MS (8/9 2 :1): 498.254 (C25H36N5O4Si, [M ‡ H] ; calc. 498.254); 520.236 (C25H35N5NaO4Si, [M ‡ Na] ;
calc. 520.236).
N6-Benzoyl-9-{2'-O-[(imidazol-1-yl)thiocarbonyl]-3'-O-(triisopropylsilyl)-b-d-erythrofuranosyl}adenine
(10) and N6-Benzoyl-9-(3'-O-[(imidazol-1-yl)thiocarbonyl)-2'-O-(triisopropylsilyl)-b-d-erythrofuranosyl}ade-
nine (11). A soln. of 8/9 2 :1 (2 g, 4 mmol) in CH2Cl2 (20 ml) was treated with 1,1'-(thiocarbonyl)diimidazole
(4.6 g, 26 mmol), stirred at 238 for 3 d, extracted once with H2O and brine, and dried (Na2SO4). Evaporation
gave a yellow oil (3.8 g), consisting of 10/11 (2 :1) and 1,1'-(thiocarbonyl)diimidazole. It was used without

Helvetica Chimica Acta ± Vol. 84 (2001)
1011
further purification. Rf(AcOEt/hexane 6 :1, 10/11 2 :1) 0.33. 1H-NMR (300 MHz, CDCl3, 10/11 2 :1): see
Table 3; additionally, 9.40 (br. s, 0.67 H), 9.37 (br. s, 0.33 H) (NH); 8.20 ± 7.80 (m, 3 arom. H); 7.80 ± 7.30
(m, 3 arom. H); 7.20 ± 6.95 (m, 2 arom. H); 1.20 ± 0.80 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see
Table 4; additionally, 183.0, 182.9 (2s, CˆS); 165.3, 165.1 (2s, CˆO); 137.0 (d); 133.7 (s); 133.6 (s); 132.8 ± 128.0
(several d); 118.1 (d); 17.8 ± 17.4 (several q, (Me2CH)3Si), 12.1, 11.9 (2d, (Me2CH)3Si). FAB-MS (NOBA, 10/11
‡
2 :1): 608 (63, [M ‡ H] ).
N6-Benzoyl-9-[2'-deoxy-3'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]adenine (12) and N6-Benzoyl-9-
[3'-deoxy-2'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]adenine (13). A suspension of 10/11 2 :1 (3.8 g) in
toluene (20 ml) and a mixture of Bu3SnH (6 ml, 22.6 mmol) and AIBN (3.5 g, 21 mmol) in toluene (10 ml) were
each degassed. The suspension of 10/11 was heated to 1108, treated with the soln. of AIBN and Bu3SnH in 6
portions over 40 min, and stirred at 1108 for 2 h. The soln. was diluted with CHCl3 (200 ml), extracted once with
H2O and brine, dried (Na2SO4), and evaporated. FC (AcOEt/hexane 1:1) gave 12 (850 mg, 44% from 8/9) and
13 (385 mg, 20% from 8/9) as colourless foams.
Data of 12: Rf(AcOEt/hexane 6 :1) 0.51. [a]D25
ˆ
40.8 (c ˆ 1.07, CHCl3). IR (CHCl3): 3406w (br.), 3003m,
2946m, 2892w, 2868m, 1708m, 1612s, 1583s, 1503m, 1457s, 1386w, 1346w, 1328m, 1071s. 1H-NMR (200 MHz,
CDCl3): see Table 3; additionally, 9.00 (br. s, NH); 8.10 ± 8.00 (m, 2 arom. H); 7.65 ± 7.50 (m, 3 arom. H); 1.20 ±
1.00 (m, Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see Table 4; additionally, 165.2 (s, CˆO); 134.0 (s); 133.1
(d); 129.2 (d, 2 C); 128.3 (d, 2 C); 17.9 (q, (Me2CH)3Si);12.0 (d, (Me2CH)3Si). FAB-MS (NOBA): 482 (77,
‡
‡
[M ‡ H] ). HR-FAB-MS: 482.2585 (C25H36N5O3Si, [M ‡ H] ; calc. 482.2587).
Data of 13: Rf(AcOEt/hexane 6 :1) 0.57. [a]D25
ˆ
51.1 (c ˆ 1.065, CHCl3). IR (CHCl3): 3403w (br.), 3002m,
2946m, 2892w, 2868m, 1707m, 1612s, 1584m, 1501m, 1455s, 1327w. 1H-NMR (300 MHz, CDCl3): see Table 3;
additionally, 9.00 (br. s, NH); 8.07 ± 7.98 (m, 2 arom. H); 7.65 ± 7.45 (m, 3 arom. H); 1.20 ± 1.00 (m, (Me2CH)3Si).
13C-NMR (75 MHz, CDCl3): see Table 4; additionally, 165.0 (s, CˆO); 133.7 (s); 132.7 (d); 128.7 (d, 2 C); 128.0
‡
(d, 2 C); 17.9 (q, (Me2CH)3Si); 12.0 (d, (Me2CH)3Si). FAB-MS (NOBA): 482 (55, [M ‡ H] ). HR-MALDI-
‡
MS: 504.240 (C25H35N5NaO3Si, [M ‡ Na] ; calc. 504.240).
N6-Benzoyl-9-(2'-deoxy-b-d-glycero-tetrofuranosyl)adenine (14). A soln. of 12 (20 mg, 0.042 mmol) in THF
(2 ml) was treated with 1m Bu4NF in THF (0.24 ml, 0.24 mmol) at 238 for 3 h and evaporated after addition of
SiO2 (0.5 g). The resulting powder was dried at 0.1 mbar for 14 h and filled in a column. Elution (CHCl3/MeOH
20 :1) gave 14 (10 mg, 74%) as colourless powder. Rf(CHCl3/MeOH 10 :1) 0.29. 1H-NMR (300 MHz, CD3OD):
see Table 3; additionally, 8.10 ± 8.00 (m, 2 arom. H); 7.70 ± 7.50 (m, 3 arom. H). 13C-NMR (75 MHz, CD3OD):
see Table 4; additionally, 168.0 (s, CˆO); 135.0 (s); 134.0 (d); 130.0 (d, 2 C); 129.8 (d, 2 C). FAB-MS (NOBA):
‡
‡
326 (56, [M ‡ H] ), 348 (16, [M ‡ Na] ).
N6-Benzoyl-9-(3'-deoxy-b-d-glycero-tetrofuranosyl)adenine (15). A soln. of 13 (100 mg, 0.208 mmol) in
THF (2 ml) was treated with 1m Bu4NF in THF (0.62 ml, 0.62 mmol) at 238 for 3 h and evaporated after addition
of SiO2 (0.5 g). The resulting powder was dried at 0.1 mbar for 14 h and filled in a column. Elution (CHCl3/
MeOH 20 :1) gave 15 (60 mg, 89%) as colourless powder. Rf(CHCl3/MeOH 10 :1) 0.42.1H-NMR (300 MHz,
CD3OD): see Table 3; additionally, 8.10 ± 8.00 (m, 2 arom. H); 7.70 ± 7.50 (m, 3 arom. H). 13C-NMR (75 MHz,
CD3OD): see Table 4; additionally, 168.0 (s, CˆO); 135.0 (s); 134.0 (d); 129.9 (d, 2 C); 129.5 (d, 2 C). FAB-MS
‡
(NOBA): 326 (65, [M ‡ H] ).
N6-Benzoyl-9-(2-deoxy-3-O-(triisopropylsilyl)-b-d-erythrofuranosyl)-8-(hydroxymethyl)adenine (16). At
708, a soln. of 12 (800 mg, 1.663 mmol) in THF (20 ml) was treated dropwise with 2m LDA in THF (4.2 ml,
8.4 mmol), stirred at 708 for 2 h, treated with DMF (3.3 ml, 42.8 mmol), stirred at 708 for 2.5 h, treated with
AcOH (1.4 ml), allowed to warm to 238, and diluted with EtOH (30 ml). The resulting soln. was treated with
NaBH4 (220 mg, 5.8 mmol) for 25 min and evaporated. The residue was taken up in CH2Cl2 (100 ml), washed
once with H2O (50 ml) and brine (50 ml), dried (Na2SO4), and evaporated. FC (AcOEt/hexane 6 :1) gave 16
(748 mg, 88%). Colourless foam. Rf(AcOEt/hexane 6 :1) 0.34. [a]D25
ˆ
40.4 (c ˆ 1.095, CHCl3). IR (CHCl3):
3404m (br.), 3006m, 2945s, 2892m, 2868s, 1706s, 1655m, 1614s, 1585s, 1462s, 1436s, 1384m, 1336s, 1072s. 1H-NMR
(300 MHz, CDCl3): see Table 3; additionally, 9.60 (br. s, NH); 8.10 ± 7.90 (m, 2 arom. H); 7.60 ± 7.40 (m, 3 ar-
om. H); 4.15 (br. s, exch., OH); 1.20 ± 0.80 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see Table 4;
additionally, 165.5 (s, CˆO); 134.2 (s); 132.8 (d); 129.0 (d, 2 C); 128.3 (d, 2 C); 17.9 (q, (Me2CH)3Si); 12.0
‡
‡
(d, (Me2CH)3Si). FAB-MS (NOBA): 512 (35, [M ‡ H] ). HR-MALDI-MS: 512.268 (C26H38N5O4Si, [M ‡ H] ;
‡
calc. 512.269), 534.251 (C26H37N5NaO4Si, [M ‡ Na] ; calc. 534.251).
N6-Benzoyl-9-(2-deoxy-b-d-erythrofuranosyl)-8-(hydroxymethyl)adenine (17). A soln. of 16 (20 mg,
0.039 mmol) in THF (1 ml) was treated with 1m Bu4NF in THF (0.12 ml, 0.12 mmol) at 238 for 3 h and
evaporated after addition of SiO2 (0.4 g). The resulting powder was dried at 0.1 mbar for 14 h and filled in a
column. Elution (CHCl3/MeOH 20 :1 to 10 :1) gave 17 (11 mg, 80%) as colourless powder. Rf(CHCl3/MeOH

1012
Helvetica Chimica Acta ± Vol. 84 (2001)
6 :1) 0.51. 1H-NMR (300 MHz, CD3OD): see Table 3; additionally, 8.10 ± 8.00 (m, 2 arom. H); 7.70 ± 7.50
(m, 3 arom. H). 13C-NMR (75 MHz, CD3OD): see Table 4; additionally, 168.3 (s, CˆO); 135.2 (s); 134.0 (d);
‡
‡
129.8 (d, 2 C); 129.5 (d, 2 C). FAB-MS (NOBA): 356 (78, [M ‡ H] ), 378 (41, [M ‡ Na] ).
N6-Benzoyl-9-[2'-deoxy-3'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]-8-[(4,4'-dimethoxytrityloxy)-
methyl]adenine (18). A soln. of 16 (748 mg, 1.46 mmol), EtN(i-Pr)2 (1.03 ml, 6 mmol) and DMAP (50 mg,
0.4 mmol) in CH2Cl2 (15 ml) was treated with DMTrCl (2 g, 5.91 mmol) at 08. The mixture was stirred at 238 for
16 h, and evaporated. The residue was diluted with CH2Cl2, washed once with H2O and brine, dried (Na2SO4),
and evaporated. FC (AcOEt/hexane 1 :2) gave 18 (1056 mg, 89%). Colourless foam. Rf(AcOEt/hexane 2 :1)
0.61. [a]2D5
ˆ
15.2 (c ˆ 2.32, CHCl3). IR (CHCl3): 3409w (br.), 3007m, 2960m, 2867w, 1706m, 1610s, 1584m,
1509s, 1463m, 1447m, 1428m, 1070m, 1034m. 1H-NMR (200 MHz, CDCl3): see Table 3; additionally, 9.06 (br. s,
NH); 8.10 ± 8.00 (m, 2 arom. H); 7.69 ± 7.20 (m, 12 arom. H); 6.92 ± 6.80 (m, 4 arom. H); 3.78 (s, 2 MeO); 1.20 ±
1.00 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see Table 4; additionally, 165.0 (s, CˆO); 159.2 (s, 2 C);
144.4 (s); 135.6 (s); 135.3 (s); 134.4 (s); 132.8 (d); 130.2 ± 127.2 (several d); 113.3 (d, 4 C); 87.9 (s, Ar3C); 55.3
‡
(q, 2 MeO); 18.0 (q, (Me2CH)3Si), 12.0 (d, (Me2CH)3Si). FAB-MS (NOBA): 814 (38, [M ‡ H] ), 303 (100,
‡
‡
DMTr ). HR-MALDI-MS: 836.382 (C47H55N5NaO6Si, [M ‡ Na] ; calc. 836.381).
N6-Benzoyl-9-[2'-deoxy-b-d-glycero-tetrofuranosyl]-8-[(4,4'-dimethoxytrityloxy)methyl]adenine (19). A
soln. of 18 (976 mg, 1.2 mmol) in THF (15 ml) was treated dropwise with 1m Bu4NF in THF (3.55 ml,
3.55 mmol) at 238 for 1.5 h and evaporated. FC (AcOEt/hexane 2 :1) gave 19 (600 mg, 76%). Colourless foam.
Rf(AcOEt/hexane 2 :1) 0.13. [a]D25
ˆ
12.5 (c ˆ 1.69, CHCl3). IR (CHCl3): 3405m (br.), 3007s, 2839w, 1707m,
1613s, 1585m, 1509s, 1462s, 1448s, 1428s, 1409m, 1336s, 1302s, 1065s, 1036s. 1H-NMR (300 MHz, CDCl3): see
Table 3; additionally, 9.20 (br. s, NH); 8.05 ± 7.95 (m, 2 arom. H); 7.60 ± 7.10 (m, 12 arom. H); 6.85 ± 6.75
(m, 4 arom. H); 3.75 (s, 2 MeO); 2.60 ± 2.40 (br. s, exch., HO C(3')). 13C-NMR (50 MHz, CDCl3): see Table 4;
additionally, 164.9 (s, CˆO); 159.0 (s, 2 C); 144.3 (s); 135.3 (s); 135.2 (s); 134.2 (s); 132.9 (d); 130.2 ± 127.3
‡
(several d); 113.6 (d, 4 C); 88.0 (s, Ar3C); 55.3 (q, 2 MeO). FAB-MS (NOBA): 659 (2, [M ‡ H] ), 303 (100,
‡
‡
DMTr ). HR-FAB-MS (NOBA): 658.2663 (C38H36N5O6, [M ‡ H] ; calc. 658.2666).
N6-Benzoyl-9-(2'-deoxy-b-d-glycero-tetrofuranosyl)-8-[(4,4'-dimethoxytrityloxy)methyl]adenine 3'-[(2-Cy-
anoethyl)diisopropylphosphoramidite] (1). A soln. of 19 (395 mg, 0.601 mmol) and EtN(i-Pr)2 (2 ml) in CH2Cl2
(20 ml) was treated with 2-cyanoethyl diisopropylchlorophosphoramidite (184 mg, 0.77 mmol) at 238, stirred for
3 h, and evaporated. FC (AcOEt/hexane 2 :1) gave 1 (413 mg, 80%, 2 diastereoisomers (1:1)). Colourless foam.
Rf(AcOEt/hexane 2 :1, 2 diastereoisomers (1 :1)) 0.34; 0.45. IR (CHCl3, 2 diastereoisomers (1:1)): 3404w (br.),
3007m, 2970m, 2935m, 1707m, 1613s, 1585m, 1509s, 1462m, 1448m, 1428w, 1408m, 1365m, 1337m, 1063m, 1036m.
1H-NMR (300 MHz, CDCl3, 2 diastereoisomers (1:1)): see Table 3; additionally, 9.00 (br. s, NH); 8.10 ± 7.95
(m, 2 arom. H); 7.70 ± 7.20 (m, 12 arom. H); 6.90 ± 6.80 (m, 4 arom. H); 3.90 ± 3.50 (m, OCH2CH2CN,
(Me2CH)2N); 3.78 (s, 2 MeO); 2.63 (t, J ˆ 6.3), 2.55 (t, J ˆ 6.1) (CH2CN); 1.30 ± 1.10 (m, (Me2CH)2N).
13C-NMR (75 MHz, CDCl3, 2 diastereoisomers (1:1)): see Table 4; additionally, 165.1 (s, N CˆO); 159.2
(s, 2 C); 144.5, 144.4 (2s, 1 C); 135.6, 135.5, 135.4, 134.4 (several s, 3 C); 132.9 ± 127.5 (several d); 117.8 (s, CN);
113.7 (d, 4 C); 88.1 (s, Ar3C); 58.5 (dt, 2J(C,P) ˆ 19.0, OCH2CH2CN); 55.4 (q, 2 MeO); 43.4 (dd, 2J(C,P) ˆ 12.0,
(Me2CH)2N); 24.8 ± 24.5 (several q, (Me2CH)2N); 20.5 ± 20.3 (m, OCH2CH2CN). 31P-NMR (121.5 MHz,
‡
‡
CDCl3): 148.9; 148.6. FAB-MS (NOBA): 858 (49, [M ‡ H] ), 303 (100, DMTr ). HR-MALDI-MS: 880.356
‡
(C47H52N7NaO7P, [ M ‡ Na] ; calc. 880.356).
1-(2',3'-Di-O-acetyl-b-d-erythrofuranosyl)uracil (20). A suspension of 5/6 1 :4 (1.8 g, 7.3 mmol) and uracil
(0.92 g, 8.2 mmol) in MeCN (18 ml) was treated dropwise with N,O-bis(trimethylsilyl)acetamide (4.32 ml,
17.7 mmol) at 608, stirred at 608 for 30 min, and treated with SnCl4 (3.47 ml, 29.5 mmol). The mixture was stirred
at 608 for 20 min, poured into a stirred mixture of AcOEt (300 ml) and sat. aq. NaHCO3 (300 ml), and stirred
for 15 min. The org. layer was dried (Na2SO4) and evaporated. FC (AcOEt/hexane 10 :1) gave 20 (1.74 g, 80%).
Colourless powder. Rf(AcOEt/hexane 6 :1) 0.38. 1H-NMR (200 MHz, CDCl3): see Table 5; additionally, 9.48
(br. s, NH); 2.14 (s, AcO); 2.09 (s, AcO). 13C-NMR (75 MHz, CDCl3): see Table 6; additionally, 170.4, 170.2 (2s,
2 O CˆO); 20.7, 20.5 (2q, 2 Me).
1-(b-d-Erythrofuranosyl)uracil (4). At 238, a soln. of 20 (1.1 g, 3.7 mmol) in a mixture of THF (94 ml),
MeOH (75 ml), and H2O (19 ml) was treated with 2m NaOH in H2O (12.4 ml, 25 mmol), stirred at 238 for 8 min,
treated with NH4Cl (2.6 g, 48.6 mmol), treated with SiO2 (22.8 g), and evaporated. The resulting colourless
powder was dried at 0.1 mbar for 14 h, filled in a column, and eluted (CHCl3/MeOH 6 :1). The colourless
crystals obtained from a few pooled fractions were submitted to X-ray analysis. Evaporation of the residual
fractions gave 4 (0.63 g, 80%). Colourless crystals. Rf (CHCl3/MeOH 6 :1) 0.16. [a]D25
(KBr): 3385s (br.), 3186s, 1693s, 1470m, 1404s, 1262m, 1130m, 1065m. H-NMR (300 MHz, (D6)DMSO): see
ˆ
41.2 (c ˆ 0.9, H2O). IR
1

Helvetica Chimica Acta ± Vol. 84 (2001)
1013
Table 5. Selected 1H-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] for 20, 22 ± 24, 26 ± 28, and 2 in
CDCl3, for 4 in (D6)DMSO, and for 21 and 25 in D2O
2
4
20
22
23
24
26
27
28
21
25
H
H
H
H
C(1')
5.87
2.88
2.32, 2.25
4.72
4.34
3.4 ± 3.9
3.98, 3.97
3.98, 3.97
5.76, 5.73 5.60
±
5.70
4.18
±
4.08
4.21
3.67
±
5.86
5.5 ± 5.6
±
5.4 ± 5.5
4.52
4.06
±
6.27 5.90
5.13 4.64
6.15
2.11
2.50
4.65
4.20
3.87
±
6.11
2.90
2.23
4.83
4.42
3.83
4.52
4.45
5.80
±
5.90
2.85
2.18
4.73
4.38
3.73
3.96
3.96
5.80
±
5.85
2.82
2.12
4.65
4.34
3.73
4.02
3.95
5.80
±
6.56
5.45
±
4.70
4.16
3.93
±
6.18
2.19
2.40
4.53
4.25
3.88
±
C(2'pro-S
C(2'pro-R
C(3')
)
)
±
±
4.64 4.53
4.04 4.35
3.80 4.02
Ha C(4')
Hb C(4')
Ha C(7)
Hb C(7)
H
H
±
±
±
±
±
±
±
±
±
C(5)
C(6)
5.77
7.22
5.3
6.06 5.76
7.38 7.20
5.73
7.32
6.5
6.22
7.94
5.3
±
5.69
7.61
7.0
7.65
6.2
±
4.4
±
4.0
2.2
9.1
8.1
±
J(1',2')
J(2',2')
6.5
13.7
6.5
5.0
±
5.6
±
6.6
12.8
7.0
13.0
7.0
13.4
±
14.0
14.0
a
J(2'pro-S,3')
J(2'pro-R,3')
J(3',4'a)
J(3',4'b)
J(4'a,4'b)
J(5,6)
)
±
< 1.0
±
4.7
±
6.2
2.3
4.2
2.2
9.0
8.1
±
6.5
< 1.0
4.4
< 1.0
8.5
6.5
< 1.0
4.1
< 1.0
8.4
6.5
< 1.0
4.0
< 1.0
9.0
< 1.0
5.3
1.6
3.7
1.5
9.3
8.1
±
a
)
±
< 1.0
2.8
11.2
7.5
4.4
< 1.0
8.7
4.5
2.0
10.3
8.1
±
< 1.0
2.5
10.6
7.5
±
4.4
3.7
9.0
8.1
±
±
±
14.7
±
±
13.4
a
a
J(7a,7b)
)
)
±
a) Not assigned.
Table 6. Selected 13C-NMR Chemical Shifts [ppm] for 20, 22 ± 24, 26 ± 28, and 2 in CDCl3, for 4 in (D6)DMSO,
and for 21 and 25 in D2O
C(1')
C(2')
C(3')
C(4')
C(7)
C(6)
C(2)
C(4)
C(5)
20
4
89.9
88.8
93.4a)
90.3a)
95.4
88.0
88.4
87.7
88.1
88.1
88.2
73.8
74.1
90.6a)
87.7a)
52.4
42.4
41.9
40.4
40.3
39.3
38.4
38.3
71.1
70.0
75.7b)
75.0b)
71.7
72.0
72.3
73.6
73.5
73.0
74.5
74.3
72.4
73.6
75.7b)
74.0b)
74.8
77.2
77.5
78.7
78.5
78.0
79.8
79.6
±
±
±
±
±
±
±
61.2
62.8
62.5
62.8
140.7
141.8
141.7
135.2
141.4
140.0
142.3
156.0
153.5
153.3
153.3
150.7
151.0
164.2
160.3
150.1
150.8
152.1
151.2
151.2
151.2
150.6
150.5
163.4
163.5
178.7
172.0
163.8
164.0
166.0
164.0
163.8
163.6
162.6
103.4
102.0
112.1
110.8
102.9
102.6
102.6
101.6
102.5
102.6
102.6
102.5
21
22
23
24
25
26
27
28
2
a
)
b) Assignments may be interchanged.
Table 5; additionally, 11.22 (br. s, NH); 5.34 (d, J ˆ 6.2, exch., HO C(2')); 5.07 (d, J ˆ 3.7, exch., HO C(3')).
‡
13C-NMR (50 MHz, (D6)DMSO): see Table 6. FAB-MS (NOBA): 215 (100, [M ‡ H] ).
2,2'-Anhydro-1-(b-d-threofuranosyl)uracil (21). A suspension of 4 (1 g, 4.67 mmol) and diphenyl
carbonate (1.29 g, 0.61 mmol) in DMF (25 ml) was stirred at 808 for 30 min, treated with NaHCO3 (390 mg)
at 808, stirred at 1508 for 2 h, and evaporated. The residue was diluted with AcOEt and H2O. Evaporation of the
aq. layer gave 998 mg of a colourless powder containing 21 and Na salts. Rf(AcOEt/MeOH 3 :2) 0.57. [a]D25
ˆ
44 (c ˆ 1.05, H2O). IR (KBr): 2923m, 1662s, 1620m, 1533s, 1482s, 1272w, 1241w, 1190w, 1113w, 1062m, 1041m.
1H-NMR (300 MHz, D2O): see Table 5. 13C-NMR (75 MHz, D2O): see Table 6. FAB-MS (NOBA): 219 (100,
‡
‡
‡
[M ‡ Na] ), 197 (24, [M ‡ H] ). HR-MALDI-MS: 219.037 (C8H8N2NaO4, [M ‡ Na] ; calc. 219.038).

1014
Helvetica Chimica Acta ± Vol. 84 (2001)
2,2'-Anhydro-1-[3'-O-(triisopropylsilyl)-b-d-threofuranosyl]uracil (22). A soln. of 21 (1 g, 5.1 mmol) in
i
DMF (16 ml) and pyridine (12.5 ml) was treated with Pr3SiOTf (7 ml, 26 mmol), stirred at 238 for 24 h, and
evaporated. FC (AcOEt/MeOH 30 :1) gave 22 (790 mg, 61.5% from 4). Colourless foam. Rf(AcOEt/MeOH
10 :1) 0.37. [a]2D5
ˆ
40.6 (c ˆ 0.9, CHCl3). IR (CHCl3): 2944s, 1672s, 1644s, 1544m, 1468s, 1387w, 1134m, 1072m,
1042m. 1H-NMR (200 MHz, CDCl3): see Table 5 ; additionally, 1.20 ± 0.90 (m, (Me2CH)3Si). 13C-NMR
(75 MHz, CDCl3): see Table 6; additionally, 18.0 (q, (Me2CH)3Si); 12.0 (d, (Me2CH)3Si). FAB-MS (NOBA):
‡
‡
‡
353 (100, [M ‡ H] ), 705 (8, [2 M ‡ H] ). HR-MALDI-MS: 353.189 (C17H29N2O4Si, [M ‡ H] ; 353.189).
1-[2'-Bromo-2'-deoxy-3'-O-(triisopropylsilyl)-b-d-erythrofuranosyl]uracil (23). suspension of 22
A
(670 mg, 1.9 mmol) and LiBr (215 mg, 2.47 mmol) in 1,4-dioxane (20 ml) was heated to 608 and treated
dropwise with BF3 ´ Et2O (0.62 ml, 4.9 mmol). The mixture was stirred at 608 for 6 h, and taken to dryness. The
residue was diluted with CHCl3 (50 ml), washed once with H2O (20 ml) and brine (20 ml), dried (Na2SO4), and
evaporated. FC (AcOEt/hexane 1:1) gave 23 (720 mg, 88%). Colourless foam. Rf(AcOEt/hexane 1:1) 0.34.
[a]2D5
ˆ
29.9 (c ˆ 1.15, CHCl3). IR (CHCl3); 3391w (br.), 2940m, 2868m, 1695s, 1633w, 1457m, 1384m, 1143m,
1108m, 1068m, 1017m. 1H-NMR (300 MHz, CDCl3): see Table 5; additionally, 8.60 (br. s, NH); 1.15 ± 1.03
(m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see Table 6 ; additionally, 18.0 (q, (Me2CH)3Si); 12.0
‡
(d, (Me2CH)3Si). FAB-MS (NOBA): 433, 435 (23, 24; [M ‡ H] ). HR-MALDI-MS: 375.173 (C17H28N2NaO4Si,
‡
[M HBr‡ Na] ; calc. 375.172).
1-[2'-Deoxy-3'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]uracil (24). A suspension of 23 (696 mg,
1.61 mmol) in toluene (10 ml) and a mixture of Bu3SnH (0.43 ml, 1.62 mmol) and AIBN (100 mg, 0.61 mmol) in
toluene (5 ml) were each degassed. The suspension of 23 was heated to 808 and treated with the soln. of AIBN
and Bu3SnH over 20 min. The mixture was heated to 1108, stirred for 30 min, diluted with CHCl3 (200 ml),
washed with H2O and brine, dried (Na2SO4), and evaporated. FC (AcOEt/hexane 1:1) gave 24 (556 mg, 97%).
Colourless foam. Rf(AcOEt/hexane 1:1) 0.30. IR (CHCl3): 3391w (br.), 2946m, 1690s, 1633w, 1461m, 1390m,
1135m, 1073m. 1H-NMR (300 MHz, CDCl3): see Table 5 ; additionally, 9.67 (br. s, NH); 1.17 ± 0.94
(m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see Table 6 ; additionally, 18.0 (q, (Me2CH)3Si); 12.0
‡
‡
(d, (Me2CH)3Si). FAB-MS (NOBA): 355 (18, [M ‡ H] ). HR-FAB-MS: 355.2052 (C17H31N2O4Si, [M ‡ H] ;
calc. 355.2053).
1-(2'-Deoxy-b-d-glycero-tetrofuranosyl)uracil (25). A soln. of 24 (91 mg, 0.258 mmol) in THF (2 ml) was
treated with 1m Bu4NF ´ 3 H2O in THF (0.76 ml, 0.76 mmol) at 238 for 3 h, and evaporated after addition of SiO2
(0.5 g). The resulting powder was dried at 0.1 mbar for 14 h, filled in a column, and eluted (CHCl3/MeOH 20 :1).
Evaporation gave 25 (45 mg, 88%) as white powder. Recrystallization in MeOH gave colourless crystals used for
X-ray analysis. Rf(CHCl3/MeOH 10 :1) 0.28. 1H-NMR (300 MHz, CD3OD): see Table 5. 13C-NMR (75 MHz,
‡
CD3OD): see Table 6. FAB-MS (NOBA): 199 (28, [M ‡ H] ).
1-[2'-Deoxy-3'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]-6-(hydroxymethyl)uracil (26). At
708,
2m LDA in THF/heptane/ethylbenzene (FLUKA; 1.63 ml, 3.26 mmol) was treated with a soln. of 24
(230 mg, 0.652 mmol) in THF (4 ml). The mixture was stirred at 708 for 30 min, treated with DMF (1.7 ml,
16.5 mmol), stirred at 708 for 2.5 h, and treated with AcOH (0.55 ml). The mixture was allowed to warm to
238, diluted with EtOH (4 ml), and treated with NaBH4 (83 mg, 2.17 mmol) for 25 min. After evaporation, a
soln. of the residue in CH2Cl2 was washed with H2O and brine, dried (Na2SO4), and evaporated. FC (AcOEt/
hexane 4 :1) gave 26 (128 mg, 51%). Colourless foam. Rf(AcOEt/hexane 1:1) 0.15. [a]D25
ˆ
82.2 (c ˆ 0.4,
CHCl3). IR (CHCl3): 3390m (br.), 2946s, 2867s, 1697s, 1602m, 1462m, 1377m, 1094s, 1015s. 1H-NMR (300 MHz,
CDCl3/CD3OD): see Table 5; additionally, 1.16 ± 0.94 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see
‡
Table 6; additionally, 18.0 (q, (Me2CH)3Si); 12.0 (d, (Me2CH)3Si). FAB-MS (NOBA): 385 (21, [M ‡ H] ).
1-[2'-Deoxy-3'-O-(triisopropylsilyl)-b-d-glycero-tetrofuranosyl]-6-[(4,4'-dimethoxytrityloxy)methyl]uracil
(27). A soln. of 26 (117 mg, 0.305 mmol), EtN(i-Pr)2 (0.24 ml, 1.39 mmol) and DMAP (10 mg, 0.4 mmol) in
CH2Cl2 (5 ml) was treated with DMTrCl (413 mg, 1.26 mmol) at 08, stirred at 238 for 16 h, and evaporated. A
soln. of the residue in CH2Cl2 (50 ml) was washed once with H2O (20 ml) and brine (20 ml), dried (Na2SO4),
and evaporated. FC (AcOEt/hexane/Et3N 1:1 :0.02) gave 27 (161 mg, 77%). Colourless foam. Rf(AcOEt/
hexane 1:1) 0.36. [a]D25
ˆ
25 (c ˆ 0.1, CHCl3). IR (CHCl3): 3389m (br.), 2975m, 1695s, 1607m, 1509s, 1457w,
1045m. 1H-NMR (300 MHz, CDCl3): see Table 5; additionally, 9.75 (br. s, NH); 7.50 ± 7.10 (m, 9 arom. H);
6.93 ± 6.70 (m, 4 arom. H); 3.80 (s, 2 MeO); 1.15 ± 0.75 (m, (Me2CH)3Si). 13C-NMR (75 MHz, CDCl3): see
Table 6; additionally, 159.4 (s, 2 C); 144.3 (s); 135.5 (s); 135.1 (s); 130.1 ± 127.6 (several d); 113.6, 113.5 (2d, 4 C);
88.6 (s, Ar3C); 55.4 (q, 2 MeO); 18.0 (q, (Me2CH)3Si); 12.0 (d, (Me2CH)3Si). FAB-MS (NOBA): 686 (7, [M ‡
‡
‡
H] ); 303 (100, DMTr ).
1-(2'-Deoxy-b-d-glycero-tetrofuranosyl)-6-[(4,4'-dimethoxytrityloxy)methyl]uracil (28). A soln. of 27
(161 mg, 0.235 mmol) in THF (6 ml) was treated dropwise at 238 with 1m Bu4NF in THF (0.7 ml, 0.7 mmol)

Helvetica Chimica Acta ± Vol. 84 (2001)
1015
for 1.5 h, and evaporated. FC (AcOEt/hexane/Et3N 10 :1 :0.02) gave 28 (120 mg, 96%). Colourless foam.
Rf(AcOEt/hexane 10 :1) 0.24. [a]D25
ˆ
17.9 (c ˆ 0.9, CHCl3). IR (CHCl3): 3390m (br.), 2961m, 1692s, 1608m,
1584w, 1509s, 1464s, 1410m, 1378m, 1302m, 1066s, 1035s. 1H-NMR (300 MHz, CDCl3): see Table 5; additionally,
9.60 (br. s, NH); 7.50 ± 7.20 (m, 9 arom. H); 6.90 ± 6.70 (m, 4 arom. H); 3.75 (s, 2 MeO). 13C-NMR (75 MHz,
CDCl3): see Table 6; additionally, 159.2 (s, 2 C); 144.2 (s); 135.2 (s, 2 C); 130.1 ± 127.7 (several d); 114.0 (d, 4 C);
‡
‡
88.2 (s, Ar3C); 55.3 (q, 2 MeO). FAB-MS (NOBA): 530 (12, [M ‡ H] , 303 (100, DMTr ). HR-FAB-MS:
‡
530.2053 (C30H30N2O7, M ; calc. 530.2053).
1-(2'-Deoxy-b-d-glycero-tetrofuranosyl)-6-[(4,4'-dimethoxytrityloxy)methyl]uracil 3'-[(2-Cyanoethyl)diiso-
propylphosphoramidite] (2). A soln. of 28 (108 mg, 0.204 mmol) and EtN(i-Pr)2 (0.12 ml, 0.69 mmol, distilled
from CaH2) in CH2Cl2 (2 ml) was treated with 2-cyanoethyl diisopropylchlorophosphoramidite (46 ml,
0.206 mmol) at 238, stirred for 3 h, and evaporated. FC (AcOEt/hexane 2 :1) gave 2 (102 mg, 69%, 2
diastereoisomers (2 :1)). Colourless foam. Rf(AcOEt/hexane 1:1, 2 diastereoisomers (2 :1)) 0.26. IR (CHCl3, 2
1
diastereoisomers (2 :1)): 3391w (br.), 2967s, 2929s, 1693s, 1608s, 1509s, 1463s, 1378s, 1064m, 1035m. H-NMR
(300 MHz, CDCl3, 2 diastereoisomers (2 :1)): see Table 5; additionally, 8.30 ± 8.20 (br. s, NH); 7.45 ± 7.20
(m, 9 arom. H); 6.90 ± 6.80 (m, 4 arom. H); 3.90 ± 3.45 (m, Me2CH)2N, OCH2CH2CN,
H C(4')); 3.80
(s, 2 MeO); 2.62 (t, J ˆ 6.5, 0.67 H), 2.52 (t, J ˆ 6.5, 1.33 H) (CH2CN); 1.36 ± 1.00 (m, (Me2CH)2N). 13C-NMR
(75 MHz, CDCl3, 2 diastereoisomers (2 :1)): see Table 6; additionally, 159.3 (s, 2 C); 144.2, 144.1 (2s, 1 C); 135.2,
135.0 (2s, 2 C); 130.2 ± 127.7 (several d); 113.9 (d, 4 C); 111.3 (s, CN); 88.4 (s, Ar3C); 58.5 (t, 2J(C,P) ˆ 18,
OCH2CH2CN); 55.5 (q, 2 MeO); 43.3 (dd, 2J(C,P) ˆ 13.0, (Me2CH)2N); 24.8 ± 24.4 (several q, (Me2CH)2N);
20.5, 20.4 (2t, OCH2CH2CN). 31P-NMR (121.5 MHz, CDCl3, 2 diastereoisomers (2 :1)): 148.7, 148.2. FAB-MS
‡
‡
(NOBA, 2 diastereoisomers (2 :1)): 513 (40, [M OPR2] ), 303 (100, DMTr ). HR-MALDI-MS: 535.184
‡
(C30H28N2NaO6 , [M HOP(OCH2CH2CN)NiPr2 ‡ Na] ; calc. 535.185), 449.156 (C18H27N4NaO6P,
‡
‡
[M DMTrH ‡ Na] ; calc. 449.157), 303.138 (C21H19O2, DMTr ; calc. 303.139), 241.108 (C9H19N2NaO2P,
‡
[HOP(OCH2CH2CN)NiPr2 ‡ Na] ; calc. 241.108).
Oligonucleotide Synthesis. Oligonucleotide syntheses were performed on a Pharmacia Gene Assembler on a
1.3-mmol scale. The commercial phosphoramidites and the CPG solid-supports were from Glen Research.
Solvents and reagents were prepared according to the protocol for the synthesis of pRNA [30]. Detritylation
was accomplished within 2 min with 3% Cl2CHCOOH in (CH2Cl)2. Couplings (0.16 ml of 0.1m phosphor-
amidite soln. ‡ 0.36 ml of 0.25m 1-(benzylthio)-1H-tetrazole soln. in MeCN) were performed within 6 ± 10 min
(DNA-phosphoramidites coupled on unmodified nucleotides), or 30 min (modified phosphoamidites, and
DNA-phosphoramidites coupled on modified nucleotides, resp.). Capping and oxidation were accomplished
under standard conditions [31].
Deprotection and Purification of Oligonucleotides. Removal of the protecting groups and detachment from
the solid support was effected in conc. aq. NH3 soln./MeOH 1:1 (2.5 ml) at 508 within 20 h (oligomers 29 and
30), or 3.5 h (oligomer 31), or at 358 within 3.5 h (oligomer 32 containing dU*, see Results and Discussion).
After filtration and evaporation of the resulting mixture, the crude oligomers were dissolved in aq. buffer soln.
(0.1m AcOH, 0.1m Et3N), purified by RP-HPLC, and desalted by RP-HPLC. Their compositions were
confirmed by MALDI-TOF mass spectroscopy.
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Received February 23, 2001