Synthesis and characterization of nucleobase functionalized monothiophenes

Article abstract of DOI:10.1016/j.tetlet.2010.08.034

The synthesis of a series of nucleobase functionalized thiophene monomers has been accomplished through the reaction of 2-bromo-1-thiophen-3-yl-ethanone with the corresponding DNA base anion. The distinctive pK_a values for the various amine groups in the nucleobases provided a pathway for the creation of specific anions through selective deprotonation of these groups. Using the appropriate anion it is possible to create an amine linkage between the thiophene and nucleobase that is, analogous to that found between the deoxyribose sugar and nucleobase, in the biologically occurring nucleoside.

Full text of DOI:10.1016/j.tetlet.2010.08.034

Tetrahedron Letters 51 (2010) 5483–5485
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Synthesis and characterization of nucleobase functionalized monothiophenes
⇑
Christopher D. McTiernan, M’hamed Chahma
Department of Chemistry & Biochemistry, Laurentian University, Sudbury, ON, Canada P3E 2C6
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 8 July 2010
Revised 10 August 2010
Accepted 11 August 2010
Available online 17 August 2010
The synthesis of a series of nucleobase functionalized thiophene monomers has been accomplished
through the reaction of 2-bromo-1-thiophen-3-yl-ethanone with the corresponding DNA base anion.
The distinctive pKa values for the various amine groups in the nucleobases provided a pathway for the
creation of specific anions through selective deprotonation of these groups. Using the appropriate anion
it is possible to create an amine linkage between the thiophene and nucleobase that is, analogous to that
found between the deoxyribose sugar and nucleobase, in the biologically occurring nucleoside.
Ó 2010 Elsevier Ltd. All rights reserved.
There is an ever increasing need to develop new technologies
for the sensing of important biomolecules such as metabolites, pro-
teins, and DNA because of their important role in disease diagnosis.
In the pursuit of this objective various materials, which exhibit
ideal properties for sensing will need to be developed. Recently,
interesting electrochemical and optical properties of conjugated
polymers such as polythiophenes have been exploited for use in
the development of such novel detection techniques.1 It has been
reported that the optical properties of a cationic conjugated poly-
mer (CCP) can be used to probe the hybridization event between
two complementary strands of DNA.2 The positively charged back-
bone of the CCP was able to interact with anionic single stranded
DNA to form a duplex that promotes complementary hybridization
by diminishing electrostatic repulsion between the two DNA com-
plementary anionic strands.3 The transition from duplex to triplex
conformation has been probed coulometrically because of changes
in color associated with conversion of the polymer from its planar,
highly conjugated conformation (duplex) to its slightly conjugated,
non-planar conformation (triplex).3
ones are plagued by solubility and reactivity problems.6 Herein
we present the (i) synthesis of adenine and thymine functionalized
monothiophenes produced through the reaction of thiophene-3-
acetyl bromide and the corresponding purine or pyrimidine anion
(Chart 1) and (ii) discussion regarding the reactivity of the nucleo-
philic sites of thymine. These nucleobase thiophene monomers
may afford the corresponding polymers after oxidation (Chart 1).
There was precedent for using an acetyl bromide functionality to
immobilize DNA nucleobases to an organic molecule.7 Huang and
co-workers incorporate an adenine nucleobase to a calyx[4]arene
using an acetyl bromide functionality.7a They reported a base-med-
iated deprotonation of the nucleobases in DMF allows the solubiliza-
tion of the adenine and other nucleobases. By adaptingthis approach
we were able to introduce nucleobases on thiophene moieties. The
3-acetylbromothiophene required for this synthesis was prepared
following a procedure described in the literature.8 It reacts with
the adenine anion to afford the corresponding functionalized thio-
phene as described in Scheme 1.9
1H and 13C NMR confirm that the amine linkage between the
thiophene and adenine molecule was in a position that was analo-
gous to the linkage between the nucleobase and sugar in the ade-
nine nucleoside.
The electrochemical and electrochromic properties of conju-
gated polymers functionalized with a biomolecule of interest can
be modified by the formation of hydrogen bonds between the teth-
ered biomolecule and a hydrogen bond donor or acceptor that is
free in solution.4 An example of this behavior is reported by
Thymine, being the complementary base to adenine was the
next nucleobase chosen for linkage to the thiophene monomer.
The structure and composition of thymine are much different than
that of adenine. Unlike adenine, which has only the one secondary
amine, thymine has two. This may affect the selectivity of the
nucleophilic attack affording two isomers. In attempting to link
the nucleobase thymine to the thiophene monomer, we found that
5
Bäuerle and Emge in which bi-thiophenes were functionalized
with pyrimidine or purine analogs and then polymerized electro-
chemically onto the surface of platinum electrodes. The resulting
films were characterized in the presence or absence of its comple-
mentary base. They found that the electrochemical properties of
the films were indeed influenced by the presence of complemen-
tary base pairing and these effects were reversible.
DNA base
DNA base
Modified DNA bases are used to facilitate the synthesis of nucle-
obase functionalized materials because the naturally occurring
[Ox]
n
S
S
⇑
Corresponding author. Tel.: +1 705 675 1151x2213; fax: +1 705 675 4844.
E-mail address: mchahma@laurentian.ca (M. Chahma).
Chart 1. DNA base functionalized oligo/polythiophenes.
0040-4039/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2010.08.034

5484
C. D. McTiernan, M. Chahma / Tetrahedron Letters 51 (2010) 5483–5485
O
O
Br
NH2
NH2
N
N
N
N
N
CuBr2/THF
NaH/DMF
N
H
N
N
S
S
4
DMF
68 %
NH2
N
N
O
N
N
1
S
Scheme 1. Synthetic route for thiophene bearing adenine.
Table 1
the base used to deprotonate the amine can have a large influence
on the identity of the final product. This provided us with a way to
selectively deprotonate the secondary amines of thymine and
other nucleobases.
In performing the synthesis of the 3-acetylthyminethiophene
using the same procedure that was used for the 3-acetyladenin-
ethiophene, but using K2CO3 as the base for amine deprotonation,
we found that thymine was attached to thiophene in the N1 posi-
tion producing the product 2 after recrystallization.10 However,
using NaH as the base to deprotonate thymine gives the undesired
N3-linked 3-acetylthyminethiophene (3).11 Scheme 2 shows the
synthetic route for the two thymine functionalized thiophene
monomers.
Yields and optical properties of nucleobase functionalized thiophenes
Monomer
Yields (%)
k (nm)
e )
(cmÀ1 MÀ1
1
2
3
68
40
70
256
260
261
4.96 Â 104
1.46 Â 104
3.54 Â 104
Table 2
Spectroscopic data of 2–3
Spectroscopic data
2
3
1H NMR of NH(1)
1H NMR of NH(3)
10.5 [10.6]
À[11.0]
À[10.6]
12.1 [11.0 ]
13C NMR of CO(2) (thymine)
13C NMR of CO(4) (thymine)
13C NMR of CO (acetyl)
150.3 [151.5]
169.6 [164.9]
192.9
156.6 [151.5]
163.9 [164.9]
187.3
The yields and optical properties of the newly synthesized
nucleobase functionalized thiophene monomers can be found in
Table 1.
The yield of monomer 2 (40%) is lower than the yield of 1 and 3.
The origin of this result is unclear. For deprotonation of thymine
with K2CO3, the reaction did not afford any other isomer, only
monomer 2. The latter may degrade during the reaction. Moreover,
it was reported that the yield of the reaction giving a thymin-1-yl
acetic acid compound using a similar procedure is around 60%.7b
Using 1H NMR of thymine N–H and 13C NMR of thymine car-
bonyl, we were able to successfully characterize both isomers. Ta-
ble 2 summarizes the spectroscopic data of the specific functional
groups (NH and CO) present in the thymine moiety for both iso-
mers 2 and 3.
Values within square braces represents, spectroscopic data of thymine12
.
O
pKa's
9.86
N1
N3
4
NH
3
5
6
13.86
2
1
N
O
H
The selective substitution can be explained through an analysis
Scheme 3. Structure of thymine showing the pKa values of its amine group at N1
and N3 positions.
of the pKa’s of thymine secondary amines as shown in Scheme 3.13
O
N
O
N
O
O
K2CO3
O
4
2
NH
S
N
H
O
NH
3
DMF
O
O
O
1
NH
N
O
4
NaH
NH
H
N
O
S
N
O
3
Scheme 2. Synthetic route for thymine functionalized thiophenes.

C. D. McTiernan, M. Chahma / Tetrahedron Letters 51 (2010) 5483–5485
5485
In the case of thymine both N1 and N3 can act as weak acids and
References and notes
can be deprotonated. Looking at the pKa’s of thymine, N1 will lose
its proton at a lower whereas N3 will lose its proton when the
media becomes more basic. It is through the N1 position that thy-
mine associates with its sugar in DNA, therefore it is at this posi-
tion we wish to substitute the thiophene monomer. The fact that
we obtained the N3-linked thiophene when using NaH as the base
for deprotonation and the N1-linked thiophene when using K2CO3
as the base for deprotonation relates back to the strength of these
bases. Of the two bases used, NaH is the stronger one. Therefore by
using the K2CO3 we were able to selectively deprotonate the thy-
mine molecule at N1.
Attempts to polymerize the prepared monomers via electro-
chemical oxidation in ACN and DMF using cyclic voltammetry at
the platinum electrode were unsuccessful. The electropolymeriza-
tion process to form a carbon–carbon bond via radical cation–rad-
ical cation coupling was not able to occur due to the high oxidation
peak potential of these monomers (the oxidation peak potential is
outside the potential window of the solvent/supporting electro-
lyte). The alternative to the electrochemical polymerization is the
chemical oxidation using either FeCl3 (Ferric chloride) or NOBF4
(Nitronium tetrafluoroborate) as oxidizing agent. Most of these
chemical polymerizations are carried out in chloride solvents such
as chloroform and dichloromethane. Due to the poor solubility of
the prepared monomers 1–3 in these solvents, it is very hard to
have a clear cut statement regarding their chemical polymeriza-
tions. Further investigations concerning the synthesis of (i) terthi-
ophenes bearing nucleobases with low oxidation potential (I) and
(ii) new monothiophenes (II) with enhanced solubility, are cur-
rently underway and will be published in due course.
1. (a) Ban, C.; Chung, S.; Park, D.-S.; Shim, Y.-B. Nucleic Acids Res. 2004, 32, 1–8; (b)
Lee, T.-Y.; Shim, Y.-B. Anal. Chem. 2001, 73, 5629–5632; (c) Higgins, S. J.;
Mouffouk, F.; Brown, S. J.; Sedghi, N.; Eccleston, B.; Reeman, S. Mater. Res. Soc.
Symp. Proc. 2005, 871E, I1.3.1–I1.3.5; (d) Mouffouk, F.; Higgins, S. J.; Brown, S. J.;
Sedghi, N.; Eccleston, B.; Reeman, S. Chem. Commun. 2004, 20, 2314–2315; (e)
Shuhmann, W.. In Enzyme and Microbial Biosensors Techniques and Protocols;
Humana Press: New Jersey, 1998; Vol. 6, pp 143–156; (f) Kim, H.-J.; Lee, K.-P.;
Gopalan, A. I.; Oh, S.-H.; Woo, J.-C. Ultramicroscopy 2008, 108, 1360–1364; (g)
Uygun, A. Talanta 2009, 79, 194–198.
2. (a) Ren, X.; Xu, Q.-H. Langmuir 2009, 25, 43–47; (b) Gaylord, B. S.; Heeger, A. J.;
Bazan, G. C. PNAS 2002, 99, 10954–10957.
3. Pu, K.-Y.; Liu, B. Biosens. Bioelectron. 2009, 24, 1067–1073.
4. Mouffouk, F.; Brown, S. J.; Demetriou, A. M.; Higgins, S. J.; Nichols, R. J.;
Rajapakse, R. M. G.; Reeman, S. J. Mater. Chem. 2005, 15, 1186–1196.
5. (a) Emge, A.; Bäuerle, P. Synth. Met. 1999, 102, 1370–1373; (b) Bäurle, P.; Emge,
A. Adv. Mater. 1998, 3, 324–330.
6. Navacchia, M. L.; Favaretto, L.; Treossi, E.; Palermo, V.; Barbarella, G. Macromol.
Rapid Commun. 2010, 31, 351–355.
7. (a) Zeng, C.-C.; Zheng, Q.-Y.; Tang, Y.-L.; Huang, Z.-T. Tetrahedron 2003, 59,
2539–2548; (b) Dueholm, K. L.; Egholm, M.; Berhens, C.; Christensen, L.;
Hansen, L. F.; Vulpius, T.; Petersen, K. H.; Berg, R. H.; Nielsen, P. E.; Buchardt, O.
J. Org. Chem. 1994, 59, 5767–5773.
8. Behroozi, S. J.; Mandal, B. K. Synth. Met. 1999, 107, 93–96.
9. Synthesis of 2-(6-amino-purin-9-yl)-1-thiophen-3-yl-ethanone (1). Adenine
(0.132 g, 0.975 mmol) and sodium hydride (NaH) (0.045 g, 1.87 mmol) were
added to 10 mL of dimethylformamide (DMF), which had been dried over 4 A°
molecular sieves and degassed for 1 h with N2. The reaction mixture was
allowed to stir for 2–3 h, at this point the adenine had been deprotonated and a
white precipitate had formed. Compound 4 (0.209 g, 1.00 mmol) was then
introduced to the flask. Immediately upon the addition of 4, the precipitate
disappeared and the reaction mixture became a light yellow. As the reaction
progressed the reaction mixture transitioned from yellow to orange and then to
red, once 4 had been consumed (ca. 45 min.). The DMF was then removed
under vacuum, and the desired product was obtained as a white solid after
recrystalization from a 50/50 mixture of water and ethanol. Yield = 68%. 1H
NMR (500 MHz, DMSO-d6): d 4.85 (s, 2H), 6.36 (s, 2H), 6.71 (m, 1H), 6.85 (m,
1H) 7.21 (s, 2H) 7.90 (m, 1H). 13C NMR (125 MHz, DMSO-d6): d 50.0, 118.5,
126.6, 128.3, 134.9, 138.9, 141.8, 150.1, 152.6, 156.2, 187.5. IR t
(cmÀ1) = 1676
(CO). UV–vis (ACN), kmax
C
(e
) = 256 nm (4.96 Â 104 MÀ1 cmÀ1). HRMS (EI) for
DNA base
11H9N5OS [M+]: calcd 259.0522; found 259.0524.
O
DNA base
(CH2)2
O
10. Synthesis of 5-methyl-3-(2-oxo-2-thiophen-3-yl-ethyl)-1H-pyrimidine-2,4-dione
(2). Thymine (0.126 g, 1.00 mmol) and NaH (0.0245 g, 1.02 mmol) were added
to 10 mL of dimethylformamide (DMF), which had been dried over 4 A°
molecular sieves and degassed for 1 h with N2. The reaction mixture was
allowed to stir for 2–3 h, at this point the thymine had been deprotonated and
a white precipitate had formed. Compound 4 (0.209 g, 1.00 mmol) was then
introduced to the flask. Immediately upon the addition of 4, the precipitate
disappeared and the reaction mixture became a light yellow. As the reaction
progressed the reaction mixture transitioned from yellow to orange and then
to red, once 4 had been consumed (ca. 1–2 h). The DMF was then removed
under vacuum, and the desired product was obtained as a white solid after
recrystallization from a 50/50 mixture of water and ethanol. Yield = 40%. 1H
NMR (500 MHz, DMSO-d6): d 0.92 (s, 3H), 4.30 (s, 2H), 6.63 (d, J = 1.2 Hz, 1H),
6.72 (m, 1H), 6.86 (m, 1H), 7.84 (m, 1H), 10.52 (br s, 1H). 13C NMR (125 MHz,
DMSO-d6): d 17.1, 59.0, 113.5, 131.5, 133.2, 139.8, 144.1, 147.26, 156.3, 169.6,
S
S
S
S
I
II
In summary, we have explored the synthesis of a series of new
thiophene monomers bearing the nucleobases adenine and thy-
mine. These new compounds were fully characterized with differ-
ent spectroscopic techniques and they are stable in air and in the
presence of a variety of organic solvents. We have also shown by
exploiting the pKa’s of the various amine groups of the nucleobases
it is possible to selectively deprotonate and functionalize a specific
amino group by using bases of varying strengths such as sodium
hydride and potassium carbonate. The use of thiophenes substi-
tuted with an acetyl bromide functionality shows a great promise
for nucleobase functionalization of a variety of thiophene mono-
mers, which can be chemically/electrochemically oxidized to form
the corresponding polymers.
192.9.
IR
t
(cmÀ1) = 1680
(CO).
UV–vis
(ACN),
kmax(e) = 260 nm
(1.46 Â 104 MÀ1 cmÀ1). HRMS (EI) for C11H10N2OS [M+]: calcd 250.0407;
found 250.0407.
11. Synthesis of 5-methyl-1-(2-oxo-2-thiophen-3-yl-ethyl)-1H-pyrimidine-2,4-dione
(3). Thymine (0.125 g, 0.992 mmol) and potassium carbonate (K2CO3) (0.150 g,
1.09 mmol) were added to 10 mL of dimethylformamide (DMF), which had
been dried over 4 Å molecular sieves and degassed for 1 h with N2. The
reaction mixture was allowed to stir for 2–3 h, at this point the thymine had
been deprotonated and a white precipitate had formed. Compound 4 (0.208 g,
0.995 mmol) was then introduced to the flask. Immediately upon the addition
of 4, the precipitate disappeared and the reaction mixture became a light
yellow. As the reaction progressed the reaction mixture transitioned from
yellow to orange and then to red, once 4 had been consumed (ca. 1–2 h). The
DMF was then removed under vacuum, and the desired product was obtained
as a white solid after recrystalization from a 50/50 mixture of water and
ethanol. Yield = 70%. 1H NMR (200 MHz, DMSO-d6): d 2.43 (s, 3H), 5.85 (s, 2H),
8.20 (s, 1H), 8.28 (m, 1H), 8.42 (m, 1H), 9.41 (m, 1H), 12.1 (s, 1H). 13C NMR
(50 MHz, DMSO-d6): d 11.5, 53.8, 107.6, 125.8, 127.6, 134.1, 138.4, 141.6,
The ability of the nucleobase functionalized thiophene to form
hydrogen bonds with their complementary base could be exam-
ined, since this is the mechanism through which DNA hybridiza-
tion takes place and is the key principle behind the use of these
polymers in DNA biosensors.
150.6, 163.9, 187.3. IR t (e) = 261 nm
(cmÀ1) = 1680 (CO). UV–vis (CH2Cl2), kmax
(3.54 Â 104 MÀ1 cmÀ1). HRMS (EI) for C20H21NO3S3 [M+]: calcd 250.0407;
found 250.0411.
Acknowledgments
12. Thureau, P.; Ancian, B.; Viel, S.; Thévand, A. Chem. Commun. 2006, 17, 1884–
1886.
13. (a) Zimmer, S.; Biltonen, R. J. Solution Chem. 1972, 1, 291–298; (b) Nawrot, B.;
Michalak, O.; Olejniczak, S.; Wieczorek, M. W.; Lis, T.; Stec, W. J. Tetrahedron
2001, 57, 3979–3985.
M.C. thanks the Laurentian University and the Natural Sciences
and Engineering Research Council of Canada (NSERC) for support-
ing this work.