Synthesis of a benzophenone C-nucleoside as potential triplet energy and charge donor in nucleic acids

Article abstract of DOI:10.1055/s-0031-1289672

A synthetic route to the C-nucleoside that bears benzophenone as a DNA base substitution directly at the anomeric center of the 2′- deoxyribofuranoside was worked out. Furthermore, the -anomer of this artificial nucleoside was converted synthetically into the corresponding DNA building block and incorporated into two representative oligonucleotides by automated phosphoramidite chemistry. The chromophore-modified DNA was characterized by methods of optical spectroscopy. The absorption band at ～350 nm can be used for selective excitation of the benzophenone chromophore outside the nucleic acid absorption range, which makes the benzophenone nucleoside potentially useful for photochemical and photobiological applications. Georg Thieme Verlag Stuttgart · New York.

Full text of DOI:10.1055/s-0031-1289672

648
PAPER
Synthesis of a Benzophenone C-Nucleoside as Potential Triplet Energy and
Charge Donor in Nucleic Acids
Benzophenone
C
-Nucl
i
eosidechael Weinberger, Hans-Achim Wagenknecht*
Institute for Organic Chemistry, Karlsruhe Institute of Technology, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany
Fax +49(721)60844825; E-mail: wagenknecht@kit.edu
Received 8 November 2011; revised 7 December 2011
synthesized as models for ribonucleotide reductases9 and
Abstract: A synthetic route to the C-nucleoside that bears ben-
zophenone as a DNA base substitution directly at the anomeric cen-
ter of the 2¢-deoxyribofuranoside was worked out. Furthermore, the
a-anomer of this artificial nucleoside was converted synthetically
as photoreactive dyads.10 Bp has been attached to phos-
phorothioates in RNA11 and to 2¢-deoxyuridine in DNA12
to form interstrand crosslinks. 4-Cyanobenzophenone has
into the corresponding DNA building block and incorporated into been extensively used for electron transfer studies with
DNA.13
two representative oligonucleotides by automated phosphoramidite
chemistry. The chromophore-modified DNA was characterized by
methods of optical spectroscopy. The absorption band at ~350 nm
Herein, we present the synthesis of a novel C-nucleoside
1 that bears the Bp chromophore as a DNA base substitu-
tion directly at the anomeric center of the 2¢-deoxyribo-
furanoside, and its incorporation into oligonucleotides by
automated phosphoramidite chemistry. Furthermore, we
evaluate preliminarily the influence of the DNA environ-
ment on the potential energy and charge transfer proper-
ties of 1 by providing the steady-state phosphorescence
and phosphorescence lifetimes of the Bp nucleoside in a
representative oligonucleotide (DNA1).
can be used for selective excitation of the benzophenone chro-
mophore outside the nucleic acid absorption range, which makes
the benzophenone nucleoside potentially useful for photochemical
and photobiological applications.
Key words: DNA, oligonucleotide, phosphorescence, photocataly-
sis, sensitizer
Benzophenone (Bp) plays a central role in photochemistry
and photocatalysis.1–8 In the biological context, the photo-
chemistry of Bp and its derivatives is applied extensively
for photoaffinity labeling1 based on three major advantag-
es:2 (i) Bp derivatives are chemically more stable than al-
ternative labels (aryl azides and diazirines), especially
under conditions of organic synthesis; (ii) the relatively
long-lived triplet state of Bp is able to abstract H atoms
even from unreactive C–H bonds, such as the a-hydrogen
from amino acids3 in proteins; and (iii) the excitation
wavelength of Bp derivatives in the UV-A region does not
interfere with the absorption range of proteins (especially
tryptophane). On the other hand, in catalytic photochemi-
cal reactions, Bp acts as an antenna collecting the light
and transferring it to the substrate via sensitization by en-
ergy or electron transfer.4 Both processes have been used
for the development of photocatalysts.5 Template-assisted
triplet energy transfer yields enantioselective [2+2] cy-
cloaddition.6,7 Photoinduced electron transfer can be ap-
plied for enantioselective cyclizations.8 Both types of C–
C bond formation are synthetically highly valuable.
The synthesis (Scheme 1) starts from commercially avail-
able 4-bromobenzophenone (2), which is converted by
standard procedure to the ethylene glycol ketal 3 (87%
yield) in order to protect the carbonyl group during the
subsequent formation of the nucleosidic bond. On the oth-
er hand, Hoffer’s chlorosugar 4 was prepared according to
the literature14 to equip the 3¢,5¢-protected 2¢-deoxyribo-
furanoside moiety with a reactive anomeric center. The
Bp 3 was converted into a nucleophile by treatment with
magnesium in anhydrous tetrahydrofuran. The subse-
quent reaction with 4 gave the 3¢,5¢-protected C-nucleo-
side 5 in 50% yield, a good value for C-nucleoside
preparation.15 The obtained a-selectivity of a/b = 4:1
seems to be typical for this route of C-nucleoside synthe-
sis since it has also been observed by others.15 The ano-
mers, 5a and 5b, were separated by flash chromatography
and were assigned based on NOE experiments. The NOE
coupling of the conformationally fixed 3¢-H was used to
differentiate the two 2¢-H atoms (Figure 1). The existence
of a NOE between either one of the identified 2¢-H atoms
and the 1¢-H elucidates the corresponding anomeric con-
figuration. Careful deprotection of the ketals in the pres-
ence of potassium hydrogensulfate16 gives 6a and 6b,
respectively. Subsequent treatment with potassium car-
bonate in anhydrous MeOH cleaves off the two p-toluoyl
groups yielding the completely unprotected C-nucleo-
sides 1a and 1b, respectively.
Despite the broad applicability of Bp derivatives, covalent
conjugates with nucleic acids can be found very rarely in
the literature.9–12 This is surprising since this offers the
possibility (i) for new photoaffinity labels to identify
DNA- and RNA-binding proteins, and (ii) for new photo-
catalytically active DNAzymes or ribozymes. Bp-substi-
tuted nucleosides and nucleoside analogues have been
Due to the better synthetic accessibility, DNA building
block 8 was prepared representatively with the a-anomer
and applied in automated oligonucleotide synthesis with
coupling times of 10 minutes. DNA1 and DNA2 were
SYNTHESIS 2012, 44, 648–652
Advanced online publication: 24.01.2012
DOI: 10.1055/s-0031-1289672; Art ID: T105111SS
© Georg Thieme Verlag Stuttgart · New York
x
x
.x
x
.2
0
1
1

PAPER
Benzophenone C-Nucleoside
649
synthesized as oligonucleotides carrying one or two 1a as bottom). The UV/Vis absorption spectra of the Bp conju-
DNA substitutions (Figure 1). The sequences of DNA1 gates reveal the existence of the Bp modification by its
and DNA2 are random, but do not bear guanine in the di- characteristic broad side band at ~350 nm (when com-
rect vicinity of 1a in order to avoid potentially undesired pared to the reference DNA3). This band appears to be
charge transfer reactions leading to oxidized guanine very small for DNA1, which is not surprising with respect
products.17,18 Adenine was placed representatively as the to the ratio between normal DNA bases and the artificial
base opposite to the Bp moiety in DNA1. The melting base 1a in the sample. Therefore, DNA2 was synthesized;
temperature (Tm) of DNA1 occurs at 56.5 °C and reveals here the side band is clearly observable due to the double
a slight destabilization of 4.5 °C compared to the corre- 1a modification. It is important to point out that the ab-
sponding completely unmodified duplex DNA3 sorption band at ~350 nm can be used for selective excita-
(Tm = 61.0 °C, with T instead of 1a). This is a typical val- tion outside the nucleic acid absorption range, which is a
ue for a C-nucleosidic modification.15
prerequisite for photochemical and photobiological appli-
cations.
The optical properties of the nucleoside 1a were charac-
terized in comparison with DNA1 and DNA2 by measure- Upon excitation at 355 nm, the steady-state phosphores-
ments of their absorption in water at room temperature cence spectrum of the nucleoside 1a in aqueous buffer so-
(Figure 2, top), and phosphorescence at 77 K (Figure 2, lution at 77 K shows a similar shape compared to those of
O
Br
2
1) Mg
ethylene glycol cyclohexane
2)
p-TolO
p-TsOH 87%
O
p-TolO
Cl
O
p-TolO
O
O
O
O
4
p-TolO
THF (anhyd)
50%
Br
α/β = 4:1
5
3
p-TolO
p-TolO
HO
O
O
O
p-TolO
p-TolO
OH
KHSO4
K2CO3
MeOH (anhyd)
68%
H2O/CH2Cl2
quant.
O
O
O
O
5α
6α
1α
DMTO
DMTO
N
N
P
O
O
NC
Cl
O
OH
O
DMT-Cl
(i-Pr)2NEt
P
pyridine (anhyd)
72%
O
CH2Cl2
65%
O
O
NC
7
8
O
O
O
O
K2CO3
KHSO4
p-TolO
HO
p-TolO
O
MeOH (anhyd)
68%
H2O/CH2Cl2
quant.
O
O
p-TolO
HO
p-TolO
6β
1β
5β
Scheme 1 Synthesis of the benzophenone C-nucleosides 1a/1b, and the DNA building block 8
© Thieme Stuttgart · New York
Synthesis 2012, 44, 648–652

650
M. Weinberger, H.-A. Wagenknecht
PAPER
2.93 ppm
2.21 ppm
nucleoside 1a in buffer (2.2 ms) is increased in DNA1
(10.2 ms), which excludes any predominant photoredox-
active process from the triplet state in DNA at 77 K.19–22
Of course, at room temperature the situation could be dif-
ferent and a full theoretical and physical-chemical ac-
count on the singlet and triplet state properties of Bp 1a in
DNA (including guanines as adjacent bases) is in prepara-
tion. The preliminary result that the overall lifetime of 1a
is not shortened, but increased in DNA represents an im-
portant feature and advantage of our Bp C-nucleoside and
gives the opportunity to develop photocatalytically active
DNAzymes based on Bp as a DNA base analogue in
aptamers as substrate binding sites. The DNA base envi-
ronment tunes the phosphorescence properties of the Bp
similarly to an organic solvent (MeOH). Using the syn-
thetic Bp C-nucleoside photoaffinity labeling strategies
can be envisioned, for example, with DNA-binding pro-
teins and DNA-processing enzymes. In conclusion, C-nu-
cleosides with Bp as an artificial DNA base offer a
significant potential to apply this chromophore in nucleic
acid related research projects.
O
O
p-TolO
H
H
H
H
O
p-TolO
H
H
H
O
p-TolO
H
O
O
p-TolO
2.31 ppm
2.51 ppm
5β
5α
DNA1
DNA2
5'
5'
G
C
A
G
C
T
C
T
A
A
1α
A
T C A C T G A 3'
A
C
A
C
G
1α
T
1α
A C A C C G A 3'
Figure 1 NMR and NOE assignments for the H-2¢ protons (arrows)
and sequences of DNA1 and DNA2; DNA3 is the reference to DNA1
with T instead of the 1a modification
the DNA, but reveals slightly altered main maxima. The
phosphorescence of 1a has its main maximum at 467 nm,
very similar to that of Bp, whereas the phosphorescence
maximum of single-stranded DNA1 is blue-shifted and
occurs at 440 nm, which shifts further to 444 nm after hy-
bridization with the complementary counterstrand to the
duplex (data not shown). Interestingly, a similar blue shift
from 467 nm to 442 nm is observed when 1a was dis-
solved in MeOH instead of aqueous buffer solution. Ob-
viously, the DNA bases behave as a polar organic solvent
around 1a in oligonucleotides. This is an important result
since it is expected that the photochemical reactivity of
the Bp derivative 1a embedded in nucleic acids is more
similar to organic solvents (like MeOH) than to water.
Chemicals and anhydrous solvents were purchased and were used
without further purification, unless otherwise mentioned. TLC was
performed on silica gel coated aluminum foil. Flash chromatogra-
phy was carried out with silica gel 60. Spectroscopic measurements
were recorded in Na-Pi buffer solution (10 mM, pH 7) with 250 mM
NaCl using quartz glass cuvettes (10 mm). Absorption spectra were
recorded with a spectrometer equipped with a 6 × 6 cell changer
unit at 20 °C. Mass spectra were measured either in the central an-
alytical facility of the Institute of Organic Chemistry of the Univer-
sity of Regensburg, or at the Institute of Organic Chemistry of KIT,
in negative and positive ionization mode. NMR spectra were re-
corded in deuterated solvents (1H at 300 MHz, 13C at 75 MHz).
Chemical shifts are given in ppm relative to TMS. 31P NMR was re-
corded with 85% H3PO4 as external standard.
The phosphorescence lifetimes of 1a and DNA1 were
measured at 77 K and sufficiently fitted by a triexponen-
tial (1a) or biexponential (DNA1) decay. Global fit anal-
ysis reveals that the overall lifetime of the isolated
2-(4-Bromophenyl)-2-phenyl-1,3-dioxolane (3)
In a Dean–Stark apparatus, 4-bromobenzophenone (2; 3.13 g, 12.0
mmol), p-toluenesulfonic acid monohydrate (202 mg, 1.06 mmol),
and ethylene glycol (4.23 mL, 75.6 mmol) were dissolved in cyclo-
hexane (20 mL). The reaction mixture was refluxed for 48 h. After
cooling to r.t., aq 1 M NaOH (40 mL) was added. The mixture was
extracted with Et2O (3 × 30 mL). The combined organic layers were
evaporated affording a white solid; yield: 3.18 g (87%); Rf = 0.68
(hexanes–EtOAc, 5:1).
1H NMR (acetone-d6): d = 4.04 (s, 4 H, 4-H, 5-H), 7.25–7.38 (m, 3
H, aryl-H), 7.40–7.56 (m, 6 H, aryl-H).
13C NMR (acetone-d6): d = 65.7 (4-C, 5-C), 109.5 (2-C), 122.4,
126.8, 129.0, 129.1, 132.0, 143.2, 143.2 (aryl-C).
HRMS (ESI): m/z calcd for C15H14BrO2 [M + H]+: 305.0172; found
[M + H]+: 305.0356.
1¢-[4-(2-Phenyl-1,3-dioxolan-2-yl)phenyl]-3¢,5¢-di-O-p-toluoyl-
1¢,2¢-didesoxyribose (5)
Activated Mg turnings (134 mg, 5.51 mmol) were stirred in anhyd
THF (3 mL) under N2 atmosphere for a few min in a 40 °C warm
ultrasonic bath. An aliquot (1 mL) of the stock solution of 3 [842
mg, 2.76 mmol in anhyd THF (5 mL)] was added slowly. For fur-
ther activation, a catalytic amount of MeI can be added. After the
reaction mixture had turned red, the remaining amount of 3 (dis-
solved in THF, 4 mL) was added. This solution was added dropwise
to a solution of 4 (670 mg, 1.72 mmol) in anhyd THF (28 mL). The
Figure 2 Normalized UV absorption spectra at r.t. (top) and norma-
lized phosphorescence at 77 K (bottom) of nucleosides 1a/1b and
DNA1–DNA3 (5.0 mM in 10 mM Na-Pi buffer, 250 mM NaCl, pH 7,
l
exc 355 nm)
Synthesis 2012, 44, 648–652
© Thieme Stuttgart · New York

PAPER
Benzophenone C-Nucleoside
651
final reaction mixture was stirred at r.t. overnight. Ice water contain-
ing NH4Cl (25 mL) was added. The mixture was extracted with
EtOAc (4 × 150 mL). The solvent was removed to dryness and the
residue was purified by flash chromatography on silica gel (hex-
anes–EtOAc 12:1) to obtain a white solid as an anomeric mixture
a/b = 4:1; yield: 498 mg (50%). Separation of the two anomers can
be achieved by column chromatography on silica gel (hexanes–
EtOAc, 12:1); Rf = 0.26 (a), 0.31 (b) (hexanes–EtOAc, 5:1).
1¢-(4-Benzoylphenyl)-1¢,2¢-didesoxyribose (1a)
Compound 6a (157 mg, 0.29 mmol) was dissolved in anhyd MeOH
(5.5 mL), and K2CO3 (89 mg) was added. The reaction mixture was
stirred at r.t. overnight until K2CO3 was completely dissolved. The
solvent was removed under reduced pressure and the residue was
purified by flash chromatography on silica gel (CH2Cl2–MeOH,
15:1) to afford a white solid; yield: 62 mg (71%); Rf = 0.29 (a), 0.37
(b) (CH2Cl2–MeOH, 9:1).
1H NMR (CDCl3): d (a-anomer) = 2.27–2.34 (m, 1 H, 2¢-H), 2.39
(s, 3 H, ArCH3), 2.41 (s, 3 H, ArCH3), 2.59–2.96 (m, 1 H, 2¢-H),
4.00–4.10 (m, 4 H, dioxolane-H), 4.53–4.62 (m, 2 H, 5¢-H), 4.66–
4.70 (m, 1 H, 4¢-H), 5.36 (t, J = 6.7 Hz, 1 H, 1¢-H), 5.58–5.62 (m, 1
H, 3¢-H), 7.12 (d, J = 8.1 Hz, 2 H, p-Tol-5¢-H,), 7.24 (d, J = 7.9 Hz,
2 H, p-Tol-3¢-H), 7.27–7.34 (m, 3 H, aryl-H), 7.40 (d, J = 8.2 Hz, 2
H, aryl-H), 7.50–7.54 (m, 4 H, aryl-H), 7.69 (d, J = 8.1 Hz, 2 H, p-
Tol-aryl-H), 7.96 (d, J = 8.1 Hz, 2 H, p-Tol-aryl-H); d (b-anomer) =
2.15–2.28 (m, 1 H, 2¢-H), 2.40 (s, 3 H, aryl-CH3), 2.43 (s, 3 H, aryl-
CH3), 2.46–2.55 (m, 1 H, 2¢-H), 4.02–4.08 (m, 4 H, dioxolane-H),
4.49–4.56 (m, 1 H, 4¢-H), 4.59–4.66 (m, 2 H, 5¢-H), 5.23 (dd,
J = 5.0, 10.9 Hz, 1 H, 1¢-H), 5.55–5.63 (m, 1 H, 3¢-H), 7.20 (d,
J = 8.0 Hz, 2 H, p-Tol-5¢-H), 7.27–7.40 (m, 7 H, aryl-H), 7.46–7.53
(m, 4 H, aryl-H), 7.87–8.02 (m, 4 H, aryl-H).
13C NMR (CDCl3): d (a-anomer) = 21.8, 21.8 (aryl-CH3), 40.4 (2¢-
C), 64.7 (5¢-C), 64.9, 65.0 (dioxolane-C), 76.5 (3¢-C), 80.2 (1¢-C),
82.3 (4¢-C), 109.5 (dioxolane-C), 125.6, 126.3, 126.3, 126.4 (aryl-
C), 126.9, 127.2 (p-Tol-C), 128.0, 128.0, 128.3 (aryl-C), 129.2,
129.3, 129.8, 129.9 (p-Tol-aryl-C), 141.5, 142.2, 142.3 (aryl-C),
144.0, 144.0, 166.3, 166.6 (p-Tol-C); d (b-anomer) = 21.8, 21.9 (ar-
yl-CH3), 41.8 (2¢-C), 65.0 (5¢-C), 64.9, 65.0 (dioxolane-C), 77.4 (3¢-
C), 80.8 (1¢-C), 83.1 (4¢-C), 109.4 (dioxolane-C), 125.9, 126.3,
126.5 (aryl-C), 127.1, 127.2 (p-Tol-C), 128.2, 128.3 (aryl-C),
129.3, 129.3, 129.8, 129.9 (p-Tol-aryl-C), 140.6, 142.0, 142.1 (aryl-
C), 143.9, 144.3, 166.3, 166.5 (p-Tol-C).
1H NMR (MeOD): d = 1.89–1.96 (m, 1 H, 2¢-H), 2.70–2.77 (m, 1 H,
2¢-H), 3.64–3.79 (m, 2 H, 5¢-H), 4.01–4.05 (m, 1 H, 4¢-H), 4.37–
4.42 (m, 1 H, 3¢-H), 5.17 (t, J = 7.7 Hz, 1 H, 1¢-H), 7.50–7.55 (m, 2
H, aryl-H), 7.55–7.60 (m, 2 H, aryl-H), 7.61–7.66 (m, 1 H, aryl-H),
7.74–7.79 (m, 4 H, aryl-H).
13C NMR (MeOD): d = 44.7 (2¢-C), 63.3 (5¢-C), 73.5 (3¢-C), 80.4
(1¢-C), 87.7 (4¢-C), 126.8, 129.5, 131.0, 131.3, 133.7, 137.7, 139.0,
150.2 (aryl-C), 198.4 (C=O).
HRMS (ESI): m/z calcd for C18H19O4 [M + H]+: 299.1278; found
[M + H]+: 299.1305.
1¢-(4-Benzoylphenyl)-5¢-O-(dimethoxytrityl)-1¢,2¢-didesoxyri-
bose (7)
Compound 1a (214 mg, 0.72 mmol) was dissolved in anhyd pyri-
dine (17.8 mL) under N2 atmosphere. 4,4¢-Dimethoxytriphenyl-
methyl chloride (267 mg, 0.79 mmol) was added and the reaction
mixture was stirred at r.t. for 45 h. The solvent was removed under
reduced pressure and the residue was purified by flash chromatog-
raphy on silica gel [hexanes–EtOAc (4:1) + 0.01% Et3N] to give a
slightly yellow solid; yield: 311 mg (72%); Rf = 0.15 (hexanes–
EtOAc, 3:1).
1H NMR (MeOD): d = 1.89–1.96 (m, 1 H, 2¢-H), 2.71–2.79 (m, 1 H,
2¢-H), 3.21–3.25 (m, 1 H, 5¢-H), 3.28–3.34 (m, 1 H, 5¢-H), 3.78 (s,
6 H, OCH3), 4.16–4.21 (m, 1 H, 4¢-H), 4.41–4.46 (m, 1 H, 3¢-H),
5.23 (t, J = 7.6 Hz, 1 H, 1¢-H), 6.83–6.89 (m, 4 H, aryl-H), 7.17–
7.81 (m, 18 H, aryl-H).
HRMS (ESI): m/z calcd for C36H35O7 [M + H]+: 579.2377; found
[M + H]+: 579.2419.
13C NMR (MeOD): d = 44.7 (2¢-C), 55.7 (OCH3), 65.5 (5¢-C), 74.4
(3¢-C), 80.8 (1¢-C), 86.9 (4¢-C), 114.0, 114.1, 127.0, 128.7, 128.8,
129.3, 129.4, 129.5, 131.0, 131.3, 131.3, 131.4, 133.8, 137.4, 137.4,
137.4, 137.8, 139.0, 146.6, 150.0, 160.1, 160.1 (aryl-C), 198.4
(C=O).
1¢-(4-Benzoylphenyl)-3¢,5¢-di-O-p-toluoyl-1¢,2¢-didesoxyribose
(6)
Compound 5 (32 mg, 0.06 mmol), KHSO4 (82 mg), and wet silica
gel (117 mg, 60% w/w silica gel + 40% w/w H2O) were added to
CH2Cl2 (2 mL). This suspension was refluxed for 1 h. The silica gel
was filtered off and washed with EtOAc (10 mL). The organic layer
was evaporated affording a slightly yellow solid; yield: 29 mg
(98%); Rf = 0.33 (a), 0.38 (b) (toluene–EtOAc, 15:1).
1H NMR (CDCl3): d (a-anomer) = 2.31–2.46 (m, 7 H, 2¢ + aryl-
CH3), 2.91–3.05 (m, 1 H, 2¢-H), 4.55–4.64 (m, 2 H, 5¢-H), 4.70–4.77
(m, 1 H, 4¢-H), 5.48 (dd, J = 4.9, 7.7 Hz, 1 H, 1¢-H), 5.57–5.66 (m,
1 H, 3¢-H), 7.11–8.01 (m, 17 H, aryl-H); d (b-anomer) = 2.18–2.31
(m, 1 H, 2¢-H), 2.39 (s, 3 H, aryl-CH3), 2.44 (s, 3 H, aryl-CH3),
2.56–2.66 (m, 1 H, 2¢-H), 4.55–4.61 (m, 1 H, 4¢-H), 4.61–4.74 (m,
2 H, 5¢-H), 5.34 (dd, J = 4.9, 10.7 Hz, 1 H, 1¢-H), 5.60–5.66 (m, 1
H, 3¢-H), 7.15–8.02 (m, 17 H, aryl-H).
13C NMR (CDCl3): d (a-anomer) = 21.8, 21.9 (aryl-CH3), 40.5 (2¢-
C), 64.7 (5¢-C), 76.5 (3¢-C), 80.0 (1¢-C), 82.7 (4¢-C), 125.5, 126.8
(aryl-C), 127.2 (p-Tol-C), 128.4, 128.4 (aryl-C), 129.2, 129.2,
129.3, 129.7, 129.89, 130.2, 130.5, 132.6, 136.7, 137.8, 144.1,
144.3 (p-Tol-aryl-C), 147.7 (aryl-C), 166.1, 166.5 (p-Tol-C), 196.5
(C=O); d = (b-anomer) = 21.8, 21.9 (aryl-CH3), 41.9 (2¢-C), 64.8
(5¢-C), 77.4 (3¢-C), 80.5 (1¢-C), 83.4 (4¢-C), 125.8, 128.4 (aryl-C),
129.4, 129.4, 129.8, 129.9, 130.2, 130.5, 131.4, 132.6, 135.1, 137.2,
144.1, 144.4 (p-Tol-aryl-C), 145.7 (aryl-C), 166.3, 166.5 (p-Tol-C),
196.5 (C=O).
HRMS (ESI): m/z calcd for C39H36O6 + K [M + K]+: 639.2143;
found [M + K]+: 639.2177.
1¢-(4-Benzoylphenyl)-3¢-O-(2-cyanoethoxydiisopropylamino-
phosphanyl)-5¢-O-(dimethoxytrityl)-1¢,2¢-didesoxyribose (8)
Compound 7 (45 mg, 0.07 mmol) was dissolved in anhyd CH2Cl2
(4.5 mL) under N2 atmosphere. i-Pr2NEt (45 mL, 0.26 mmol) was
added and the reaction mixture was stirred at r.t. for 10 min. 2-Cy-
anoethoxydiisopropylaminophosphanyl chloride (25 mL, 0.11
mmol) was added and the solution was stirred at r.t. for 2 h. More
2-cyanoethoxydiisopropylaminophosphanyl chloride (12 mL, 0.05
mmol) was added and the mixture was stirred at r.t. for another 2 h.
The mixture was directly transferred on a silica gel column for pu-
rification [hexanes–EtOAc (2:1) + 0.1% Et3N]. Flash chromatogra-
phy afforded a white foam; yield: 39 mg (65%); Rf = 0.38, 0.44
(hexanes–EtOAc, 3:1).
1H NMR (DMSO-d6): d = 0.92–1.00 (m, 6 H, i-Pr-CH3), 1.05–1.10
[m, 6 H, CH(CH3)2], 1.92–2.09 (m, 1 H, 2¢-H), 2.56–2.66 (m, 2 H,
CH2), 2.73–2.81 (m, 1 H, 2¢-H), 3.06–3.12 (m, 1 H, 5¢-H), 3.16–3.24
(m, 1 H, 5¢-H), 3.37–3.48 (m, 2 H, CH2), 3.51–3.58 [m, 2 H,
CH(CH3)2], 3.71–3.76 (m, 6 H, OCH3), 4.21–4.31 (m, 1 H, 4¢-H),
4.44–4.53 (m, 1 H, 3¢-H), 5.25-5.34 (m, 1 H, 1¢-H), 6.87–6.93 (m, 4
H, aryl-H), 7.20–7.77 (m, 18 H, aryl-H).
HRMS (ESI): m/z calcd for C34H31O6 [M + H]+: 535.2115; found
13C NMR (DMSO-d6): d = 19.7, 19.7 (CH2), 24.1, 24.2, 24.2, 24.3,
24.3, 24.3 [CH(CH3)2], 41.9 (2¢-C), 42.4, 42.5 (CH2), 55.0 (OCH3),
[M + H]+: 535.2142.
© Thieme Stuttgart · New York
Synthesis 2012, 44, 648–652

652
M. Weinberger, H.-A. Wagenknecht
PAPER
58.0, 58.1, 58.2 [CH(CH3)2], 63.8 (5¢-C), 74.59, 75.0 (3¢-C), 78.8,
78.9 (1¢-C), 85.5 (4¢-C), 113.2 (aryl-C), 118.9, 118.7 (CN), 125.7,
125.8, 126.7, 127.7, 127.7, 127.8, 128.6, 129.5, 129.5, 129.7, 129.7,
129.7, 129.8, 132.6, 135.6, 135.8, 137.2, 144.9, 158.1 (aryl-C),
195.46 (C=O).
References
(1) See review: Fleming, S. A. Tetrahedron 1995, 51, 12479.
(2) See review: Dormán, G.; Prestwich, G. D. Biochemistry
1994, 33, 5661.
(3) Galardy, R. E.; Craig, L. C.; Jamieson, J. D.; Printz, M. P.
31P NMR (DMSO-d6): d = 148.21, 148.45.
J. Biol. Chem. 1974, 249, 3510.
MS (ESI): m/z calcd for C48H54N2O7P [M + H]+: 801.4; found [M +
(4) See reviews: (a) Fagnoni, M.; Dondi, D.; Ravelli, D.; Albini,
A. Chem. Rev. 2007, 107, 2725. (b) Ravelli, D.; Dondi, D.;
Fagnoni, M.; Albini, A. Chem. Soc. Rev. 2009, 38, 1999.
(5) See reviews: (a) Svoboda, J.; König, B. Chem. Rev. 2006,
106, 5413. (b) Wessig, P. Angew. Chem. Int. Ed. 2006, 45,
2168.
(6) (a) Müller, C.; Bauer, A.; Bach, T. Angew. Chem. Int. Ed.
2009, 48, 6640. (b) Müller, C.; Bauer, A.; Maturi, M. M.;
Cuquerella, M. C.; Miranda, M. A.; Bach, T. J. Am. Chem.
Soc. 2011, 133, 16689.
H]+: 801.4.
Oligonucleotides
Unmodified oligonucleotides were purchased and used without fur-
ther purification. Bp-modified DNA1 and DNA2 were prepared us-
ing standard phosphoramidite chemistry. Reagents and controlled
pore glass (CPG) (1 mmol) were purchased and used without further
purification. The concentration of 8 was increased to 0.1 M
(MeCN). After preparation, the oligonucleotides were cleaved from
the resin and deprotected by treatment with concd NH4OH at 50 °C
for 24 h. The oligonucleotides were purified by semi-preparative
HPLC column (RP-18), using the following gradient, 0–15% B over
50 min (A: 50 mM NH4OAc buffer, B: MeCN), and identified by
MS (ESI).
(7) Kauble, D. F.; Lynch, V.; Krische, M. J. J. Org. Chem. 2003,
68, 15.
(8) Bauer, A.; Westkämper, F.; Grimme, S.; Bach, T. Nature
2005, 436, 1139.
(9) (a) Lehmann, T. E.; Berkessel, A. J. Org. Chem. 1997, 62,
302. (b) Lehmann, T. E.; Müller, G.; Berkessel, A. J. Org.
Chem. 2000, 65, 2508.
DNA1
M = 5222.9.
(10) Paris, C.; Encinas, S.; Belmadoui, N.; Climent, M. J.;
MS (ESI): m/z calcd for (C174H210N60O99P16)4– [M – 4 H]4–: 1304.7;
(C174H211N60O99P16)3– [M – 3 H]3–: 1740.0; found [M – 4 H]4–:
1305.1, [M – 3 H]3–: 1740.5.
Miranda, M. A. Org. Lett. 2008, 10, 4409.
(11) Musier-Forsyth, K.; Schimmel, P. Biochemistry 1994, 33,
773.
(12) Nakatani, K.; Yoshida, T.; Saito, I. J. Am. Chem. Soc. 2002,
124, 2118.
DNA2
M = 5233.0.
(13) Nakatani, K.; Saito, I. Top. Curr. Chem. 2004, 236, 163.
(14) (a) Hoffer, M. Chem. Ber. 1960, 93, 2777. (b) Rolland, V.;
Kotera, M.; Lhomme, J. Synth. Commun. 1997, 27, 3505.
(c) Dhimitruka, I.; SantaLucia, J. Jr. Synlett 2004, 335.
(d) Chin, T.-M.; Huang, L.-K.; Kan, L.-S. J. Chin. Chem.
Soc. 1997, 44, 413.
MS (ESI): m/z calcd for (C180H212N60O95P16)4– [M – 4 H]4–: 1307.2;
(C180H213N60O95P16)3– [M – 3 H]3–: 1743.3.0; found [M – 4 H]4–:
1307.7, [M – 3 H]3–: 1744.0.
Supporting Information for this article is available online at
http://www.thieme-connect.com/ejournals/toc/synthesis. Included
are images of 1H NMR, 13C NMR, 31P NMR and mass spectra of the
synthesized nucleotides, as well as images of HPLC traces and mass
and phosphorescence lifetime spectra of 1a and DNA1.
(15) Štambaský, J.; Hocek, M.; Kočovský, P. Chem. Rev. 2009,
109, 6729.
(16) Mirjalili, B. F.; Zolfigol, M. A.; Bamoniri, A.; Hazar, A.
Bull. Korean Chem. Soc. 2004, 25, 1075.
(17) Adam, W.; Arnold, M. A.; Nau, W. M.; Pischel, U.; Saha-
Möller, C. R. J. Am. Chem. Soc. 2002, 124, 3893.
(18) (a) Douki, T.; Cadet, J. Int. J. Radiat. Biol. 1999, 75, 571.
(b) Morin, B.; Cadet, J. Photochem. Photobiol. 1994, 60,
102.
(19) Delatour, T.; Douki, T.; D’Ham, C.; Cadet, J. Photochem.
Photobiol. 1998, 44, 191.
(20) Hélène, C.; Charlier, M. Biochimie 1971, 53, 1175.
(21) Lamola, A. A. J. Am. Chem. Soc. 1966, 88, 813.
(22) Jennings, B. H.; Pastra, S.-C.; Wellington, J. L. Photochem.
Photobiol. 1970, 11, 215.
Acknowledgment
Financial support by GRK 1626 (Chemical Photocatalysis) and KIT
is gratefully acknowledged. We thank Hartmut Yersin and his
group (Chemistry Department, University of Regensburg, Germa-
ny) for measurement of the phosphorescence spectra and lifetimes.
Synthesis 2012, 44, 648–652
© Thieme Stuttgart · New York