Synthesis and crystal structures of fluorescent receptors for 9-butyladenine

Article abstract of DOI:10.1016/j.tet.2008.10.041

Two pyrene containing fluorescent receptors, 2-(1-pyrenyl)benzoic acid (FR-1) and 8-(1-pyrenyl)-1-naphthoic acid (FR-2), have been designed and synthesized to mimic a pyrene dinucleotide for molecular recognition of 9-butyladenine (9-BuA). The X-ray cryst

Full text of DOI:10.1016/j.tet.2008.10.041

Tetrahedron 65 (2009) 62–69
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Synthesis and crystal structures of fluorescent receptors for 9-butyladenine
,
Bassam Lamale a, William P. Henry b, Lee M. Daniels c, Cungen Zhang d, Suzane M. Klein a, Yu Lin Jiang a
*
a Department of Chemistry, College of Arts and Sciences, East Tennessee State University, Johnson City, TN 37614, USA
b Department of Chemistry, Mississippi State University, Mississippi State, MS 39762, USA
c Rigaku Americas Corporation, 9009 New Trails Drive, The Woodlands, TX 77381, USA
d Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Two pyrene containing fluorescent receptors, 2-(1-pyrenyl)benzoic acid (FR-1) and 8-(1-pyrenyl)-1-
naphthoic acid (FR-2), have been designed and synthesized to mimic a pyrene dinucleotide for molecular
recognition of 9-butyladenine (9-BuA). The X-ray crystal structures of the receptors FR-1 and FR-2 along
with the binding substrate 9-BuA have been determined. FR-1 has the carboxyl group in the same plane
as the phenyl group whereas the pyrenyl group is perpendicular to the phenyl group. However, both
carboxyl and pyrenyl groups in FR-2 are parallel to each other but perpendicular to the naphthyl group.
The binding constant for FR-2 to 9-BuA was found to be 7896ꢀ2187 Mꢁ1, which is 8.3-fold greater than
that for FR-1 (953ꢀ129 Mꢁ1). The results indicate that the complex of 9-BuA with FR-2 is more stable
than that with FR-1 by 1.2 kcal/mol. In addition, the molecular recognition of 9-BuA with the receptors
can also be observed using fluorescence spectroscopy.
Received 6 October 2008
Accepted 15 October 2008
Available online 19 October 2008
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
phosphate backbone, and carboxyl groups were used to resemble
the DNA base thymine, which is a complementary base of adenine in
A recent innovation is the application of fluorescent hydrocar-
bons as DNA base analogues in biological chemistry and drug dis-
covery.1–5 Pyrene is approximately the same size as a DNA base pair
and can therefore mimic an entire DNA base pair and selectively
bind with a DNA abasic site during DNA replications promoted by
DNA polymerases. As a result, pyrene nucleotide was applied as
a molecular wedge, DNA base pusher, and enzyme activity rescuer
and promoter during enzymatic DNA base flipping and DNA re-
pair.6,7 One of the key features of pyrene is its large surface area for
a DNA base pair. Herein, we report the synthesis of these two re-
ceptors by Suzuki coupling and X-ray structures of the receptors and
the DNA adenine analogue, 9-butyladenine (9-BuA).11 Binding
constants of the receptors with the 9-butyladenine have been de-
termined using NMR spectroscopy. Molecular recognition of 9-BuA
has also been carried out using fluorescence spectroscopy.
2. Results and discussion
p–p stacking interactions. Kool and co-workers reported that while
2.1. Synthesis of FR-1, FR-2, and 9-BuA
pyrene was used as a dangling nucleotide base analogue, it con-
tributed more stability to duplex DNA than any DNA base, such as
adenine, cytosine, guanine or thymine.8,9 Although the solution
structures of pyrene-containing DNA have been determined,
attempts to obtain crystal structures have been unsuccessful.10
Another approach to studying the interactions of pyrene di-
nucleotides is to synthesize molecular mimics and study their
binding with a suitable DNA base analogue. Therefore, two fluo-
rescent receptors, 2-(1-pyrenyl)benzoic acid (FR-1) and 8-(1-pyr-
enyl)-1-naphthoic acid (FR-2), were designed based on the structure
of a pyrene dinucleotide using chemical mimicry (Scheme 1).8,9 The
rigid phenyl and naphthyl rings were used to mimic the DNA
A standard Suzuki coupling was applied in the synthesis of both
receptors FR-1 and FR-2.12 In the synthesis of FR-1 (Scheme 2), 2-
bromoiodobenzene (1) was used as starting material because of the
different reactivity for iodine and bromine during palladium cata-
lyzed coupling reactions. Indeed, coupling product 2 was the only
product isolated. Compound 2 was then converted to the carboxylic
acid using n-butyllithium to generate a carbanion, which reacted
with carbon dioxide to form lithium carboxylate. Following acidi-
fication, receptor FR-1 was obtained in moderate yield.13 Receptor
FR-2 was synthesized using a similar procedure with 1,8-dibro-
monaphthalene (3) as starting material (Scheme 3).
9-Butyladenine was synthesized as described in a preliminary
report from adenine and freshly prepared 1-iodobutane
(Scheme 4).14 In the purification step, the product was recrystal-
lized from toluene followed by aqueous ethanol solution (10%).15
* Corresponding author. Tel.: þ1 423 439 6917; fax: þ1 423 439 5835.
E-mail address: jiangy@etsu.edu (Y.L. Jiang).
0040-4020/$ – see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2008.10.041

B. Lamale et al. / Tetrahedron 65 (2009) 62–69
63
DNA
O
O
O
O
P
Chemical Mimicry
O
or
O
O
O
FR-2
O
H
O
H
N
O
O
N
FR-1
DNA
pyrene dinucleotide
Scheme 1. Chemical mimicry of pyrene dinucleotide using receptors FR-1 and FR-2.
Br
1-pyrenyl boronic acid
Pd(PPh3)4, K2CO3
1) n-BuLi
I
2) CO2
3) HCl
58%
OH
Br
O
FR-1
46%
1
2
Scheme 2. Synthesis of receptor FR-1.
Br Br
1-pyrenyl boronic acid
1) n-BuLi
Pd(PPh3)4, K2CO3
2) CO2
3) HCl
48%
OH
Br
O
54%
3
4
FR-2
Scheme 3. Synthesis of receptor FR-2.
Interestingly, the structure of FR-1 shows the carboxyl group in
approximately the same plane as the phenyl group [O]C–
C1(ph)–C2(ph) torsion angle¼161.3(3)ꢂ] and the pyrenyl group
perpendicular to the phenyl group [C2(py)–C1(py)–C2(ph)–
C1(ph) torsion angle¼ꢁ63.5(3)ꢂ].17 In addition, pyrenyl groups
are turned away from each other in the dimer (Fig. 3). The
structure of FR-2 shows carboxyl [O]C–C1(ph)–C2(ph) torsion
NH2
N
NH2
7
6
5
N
N
1
2
N
n-C4H9-I, Cs2CO3
51%
8
N
H
N
9
4
N
N
3
9-BuA
Scheme 4. Synthesis of 9-BuA.
angle¼ꢁ56.4(6)ꢂ]
and
pyrenyl
[C2(py)–C1(py)–C8(naph)–
C7(naph) torsion angle¼ꢁ64.8(6)ꢂ] groups parallel to each other
but perpendicular to the naphthyl group. The distance between
the two pyrenyl groups is 6.79 Å (Fig. 4) indicating that these
two pyrenyl groups should interact with each other through
2.2. Crystallographic study of receptors FR-1, FR-2, and 9-BuA
Crystallographic studies were performed to explore the struc-
tures of the receptors FR-1 and FR-2. Suitable crystals for crys-
tallographic study were grown as follows. Both receptors FR-1 and
FR-2 were dissolved in hot acetonitrile. After cooling to room
temperature and slow evaporation, dark brown crystals formed on
the surface of the containers. The solvent was then decanted, the
crystals dried under vacuum, and carefully scraped from the
containers.15,16 Colorless crystals of 9-butyladenine were obtained
p–p
interactions.8,9 The closeness of the pyrenyl groups in the
dimer provides evidence for the formation of pyrene excimer
by slow cooling of
a saturated hot aqueous alcohol (10%)
solution.15
The molecular structures of FR-1 and FR-2 as determined by
crystallography are shown in Figs. 1 and 2. As expected, both re-
ceptors form dimers through hydrogen bond association of car-
boxyl groups (Figs. 3 and 4). The parameters for these hydrogen
bonds are given in Table 1. The distance from the carbonyl oxygen to
carboxyl proton of another molecule is 1.86 Å for both FR-1 and FR-2.
Figure 1. Thermal ellipsoid drawing of an FR-1 molecule.

64
B. Lamale et al. / Tetrahedron 65 (2009) 62–69
Figure 2. Thermal ellipsoid drawing of an FR-2 molecule.
Figure 5. Thermal ellipsoid drawing of a 9-BuA molecule.
Figure 3. Thermal ellipsoid diagram of FR-1 dimer.
Figure 6. Thermal ellipsoid diagram of 9-BuA dimer.
Figure 4. Thermal ellipsoid diagram of FR-2 dimer.
Table 1
Hydrogen bonding parameters for FR-1, FR-2, and 9-BuA
Compound
D–H/A
D–H (Å)
H/A (Å)
D/A (Å)
D–H/A (ꢂ)
O–H/O]Ca
O–H/O]Cb
N–H/7-Nc
N–H/1-Nd
O–H/3-N
0.83
0.82
0.87
0.87
1.01(5)
1.01(6)
1.86
1.86
2.18
2.09
1.94(5)
2.16(7)
2.684(3)
2.672(3)
3.050(4)
2.928(4)
2.913(4)
3.064(5)
169
172
177
161
163(5)
148(5)
Figure 7. Close contact between C8-H of 9-BuA to a water molecule.
FR-1
FR-2
9-BuA
different, the dimerization constants, 824ꢀ36 and 960ꢀ10 Mꢁ1
respectively, were very similar (Figs. S1 and S2).
,
O–H/1-Ne
a
ꢁx, 2ꢁy, 1ꢁz.
The X-ray structure of 9-butyladenine was also determined
(Fig. 5). In the crystal, a water molecule is present, which forms
hydrogen bonds with N3 of one 9-butyladenine and N1 of another
(Fig. 6). Furthermore, a close contact of 2.69 Å is observed between
H8 of 9-BuA and the oxygen of a water molecule [C–H: 0.94 Å; C/O
(x, 1þy, z): 3.537(5) Å; C–H$$$O: 150ꢂ] (Fig. 7). This observation
suggests that H8 may be involved in a DNA base pair between
adenine and thymine, and provides evidence of the spectator
H-bond (Scheme 5).20–22
b
c
2ꢁx, 2ꢁy, ꢁz.
ꢁx, 1/2þy, ꢁ1/2ꢁz.
ꢁx, ꢁ1/2þy, ꢁ1/2ꢁz.
x, 3/2ꢁy, 1/2þz.
d
e
for FR-2 with the maximum emission at 472 nm in chloro-
form.18,19 However, the pyrenyl groups in the FR-1 dimer do not
overlap explaining why no pyrene excimer emission for FR-1 is
observed. Although the structures of the two dimers are very

B. Lamale et al. / Tetrahedron 65 (2009) 62–69
65
DNA
N
O
H
HN
H
N
N
N
N
CH3
H
N
N
O
O
H
H
N
N
O
C4H9-n
N
N
H8
DNA
+ 0.77 ppm
N
NH2
Scheme 5. Possible involvement of adenine H8 in hydrogen bonding with oxygen of
thymine in a DNA base pair.
N
-0.12 ppm
-0.20 ppm
N
N
-0.26 ppm -0.13 ppm
More importantly, the 9-butyladenine molecules form hydrogen
bonded chains. The adenine moiety forms both Watson–Crick and
Hoogsteen hydrogen bonding with two other molecules (Fig. 8).23
The distances from hydrogen atoms of the amino group to nitrogen
atoms of the purine moiety are short (Table 1) indicating that
strong hydrogen bonding is the key interaction between adenine
molecules.15
Scheme 6. Molecular recognition of 9-BuA with receptor FR-1 and chemical shift
changes for 9-BuA after binding.
H
N
N
O
O
H
H
N
N
C4H9-n
N
+ 0.63 ppm
N
NH2
N
N
-0.02 ppm
-0.26 ppm
N
-0.10 ppm
-0.53 ppm
Scheme 7. Molecular recognition of 9-BuA with receptor FR-2 and chemical shift
changes for 9-BuA after binding.
Job’s plots, generated to identify the binding stoichiometry bet-
ween 9-BuA and the receptors, showed 1:1 binding association for
both receptors FR-1 and FR-2 (Figs. S3 and S4).25 The binding con-
stant for the complex betweenFR-1 and 9-BuAwas determined tobe
953ꢀ129 Mꢁ1 using a 1H NMR titration experiment with deuter-
iochloroform (CDCl3) as solvent (Fig. S5). Using a similar method,
the binding constant for FR-2 and 9-BuA was determined to be
greater than 104, which is the upper limit for reliability of this ex-
periment.26,27 Therefore, the binding constant for the FR-2:9-BuA
complex was estimated to be 7896ꢀ2187 Mꢁ1 using a 1H NMR
competitive titration experiment (Table S1).26,27 The latter value is
8.3-fold greater than the former one, suggesting that receptor FR-2 is
a better receptor than FR-1 for 9-BuA. The complex FR-2:9-BuA is
more stable than FR-1:9-BuA by 1.2 kcal/mol, indicating that the
Figure 8. Thermal ellipsoid diagram of showing four molecules of 9-BuA chain.
2.3. Molecular recognition of 9-BuA with receptors FR-1 and
FR-2
former has stronger
p–p stacking interactions and hydrogen
Both of the receptors FR-1 and FR-2 were applied in the mo-
lecular recognition of a substrate 9-BuA as studied by titrations
using NMR spectroscopy. First, the variation in proton chemical
shifts of the amino and alkyl groups of 9-BuA (1.00 mM) was de-
termined based on NMR experiments in the presence or absence
of the same molar amount of a receptor (1.00 mM) (Scheme 6). In
the presence of the receptor FR-1, the proton chemical shift of the
amino group moved downfield by 0.77 ppm, suggesting that the
amino group was hydrogen bonding with the carboxyl group of
the receptor. All proton chemical shifts of the butyl group moved
upfield. For example, the chemical shift of the N–CH2 protons
moved upfield by 0.26 ppm indicating that the butyl group was in
bonding. Thefreeenergy difference between thetwocomplexes was
calculated using Eq. 1.
DDG [
[ LRT ln KbðFRꢁ2:9ꢁBuAÞ=KbðFRꢁ1:9ꢁBuAÞ
where DDG is the free energy difference between complexes and
GFR-2:9-BuA and GFR-1:9-BuA are the free energies for the com-
D
G
D
FRꢁ2:9ꢁBuAL GFRꢁ1:9ꢁBuA
(1)
D
D
plexes of 9-BuA with receptors FR-2 and FR-1. Kb(FR-2:9-BuA) and
Kb(FR-1:9-BuA) are the binding constants for receptors FR-2 and
FR-1 with 9-BuA.
An important question to resolve is the contribution to the in-
a shielded area and there were
p–
p
stacking interactions between
termolecular interaction that results from p–p stacking in-
the adeninyl group in 9-BuA and the pyrenyl group of the receptor
FR-1. With receptor FR-2, the chemical shift of the amino group
protons of 9-BuA moved downfield by 0.63 ppm suggesting that
the 9-BuA also formed hydrogen bonding with the carboxyl group
of FR-2 (Scheme 7). The chemical shift of the N–CH2 protons in
9-BuA moved upfield by 0.53 ppm, indicating that the 9-BuA is
teractions of the pyrenyl groups in the receptors with 9-BuA. Rao
and co-workers reported a binding constant of 150 Mꢁ1 for com-
plex formation between 9-BuA and an aromatic acid receptor,
benzoic acid.28 The relative values of the binding constants of
receptors FR-1 and FR-2 to that of benzoic acid are 6.4 and 53
(Table 2). Thus, the energy stabilizations from
p–p stacking
involved in stronger
p
–p
stacking interactions with the pyrenyl
interactions were estimated to be 1.1 and 2.3 kcal/mol for the re-
ceptors FR-1 and FR-2, respectively, using a treatment as in Eq. 1.
group in receptor FR-2 than in FR-1.24

66
B. Lamale et al. / Tetrahedron 65 (2009) 62–69
Table 2
The energy stabilizations from p–p stacking interactions besides hydrogen bonding
in the molecular recognition of 9-BuA with the receptors FR-1 and FR-2 in CDCl3
(296 K)
Receptor
Kb (Mꢁ1
)
Kb(receptor)/Kb(benzoic acid
)
DDG (kcal/mol)
Benzoic acid
FR-1
FR-2
150a
953
d
d
6.4
53
ꢁ1.1
ꢁ2.3
7896
Figure 10. Calculated structure of the complex FR-2:9-BuA.
a
Literature data.28
Moreover, the structures of the complexes of 9-BuA with the
receptors FR-1 and FR-2, individually were calculated using Spar-
tan’06 software at the RM1 level (Figs. 9 and 10).29 The distances
and bond angles in the hydrogen bonds were then analyzed (Table
S2). The complex structures show both receptors formed
p–p
stacking interactions with the binding substrate 9-BuA. However,
hydrogen bonding in the complex FR-2:9-BuA was stronger than
that in FR-1:9-BuA based on hydrogen bonding parameters. The
average calculated hydrogen bond length, 1.771 Å in the complex
FR-2:9-BuA was shorter than the average value of 1.813 Å in FR-1:9-
BuA, and the average calculated hydrogen bond angle of 169ꢂ in the
complex FR-2:9-BuA was closer to 180ꢂ than that in FR-1:9-BuA
(157ꢂ). In terms of
p–p stacking interactions, the adeninyl ring of
9-BuA was approximately parallel to the pyrenyl group with a dis-
tance of 5.37 Å for the complex FR-2:9-BuA. For the complex FR-
1:9-BuA, the
p–p stacking interactions were weaker because the
adeninyl ring of 9-BuA was much less parallel to the pyrenyl group
and there was a slightly greater average distance of 5.68 Å (Figs. S6
and S7). The calculated structures explain why protons in the butyl
group of the complex FR-2:9-BuA were shielded more significantly
than those in the FR-1:9-BuA complex. Therefore, theoretical cal-
culations agree with the experimental results in demonstrating
that FR-2 was a better mimic for the pyrene dinucleotide than FR-1.
Figure 11. Titration of FR-1 with 9-BuA monitored by fluorescence.
fluorescence intensity at 441 nm (Fig. 11) while the binding of the
FR-2 receptor by 9-BuA resulted in an increase of pyrene fluores-
cence at the same wavelength (Fig. 12). For FR-2, there was a blue
shift of the maximum pyrene fluorescence peak from 472 to
441 nm with increasing 9-BuA concentration. Remarkably, during
the binding of the receptor FR-2 by 9-BuA, a visual color change of
the fluorescence emission from greenish blue of receptor FR-2 to
dark blue of the complex FR-2:9-BuA (Fig. 13) was observed. The
blue shift and emission color change during molecular recognition
of 9-BuA by FR-2 are consistent with the diminishing of FR-2’s
excimer emission at 472 nm and recovery of the maximum
The results also indicate that both
p–p stacking interactions and
hydrogen bonding were important in the stabilization of DNA du-
plex and should work in concert for maximum stabilization.
2.4. Pyrene fluorescence spectroscopic study of FR-1 and FR-2
in the molecular recognition of 9-BuA
The fluorescence spectra of receptors FR-1 and FR-2 in chloro-
form were obtained. FR-1 has a maximum emission peak at 441 nm,
while for FR-2 the maximum emission peak occurs at 472 nm. The
shift to higher wavelength for FR-2 can be explained because the
emission for this receptor is from an excimer facilitated in the di-
mer through intermolecular interactions.18,19
Molecular recognition studies of 9-BuA by receptors FR-1 and
FR-2 were performed using fluorescence spectroscopy.30 The ex-
periments were carried out with chloroform as solvent at room
temperature (296 K). Pyrene fluorescence changes were monitored
during titration of the receptors FR-1 and FR-2 with 9-BuA
(Schemes 6 and 7). It is interesting to note that the molecular
recognition of 9-BuA by FR-1 resulted in a decrease of pyrene
Figure 9. Calculated structure of the complex FR-1:9-BuA.
Figure 12. Titration of FR-2 with 9-BuA monitored by fluorescence.

B. Lamale et al. / Tetrahedron 65 (2009) 62–69
67
(Merck, Kieselgel 60 F254). The melting points were determined
using a MEL-TEMP apparatus. IR spectra were recorded on a Genesis
II FT-IRÔ spectrometer. The 1H and 13C NMR spectra were recorded
on a 400 MHz Varian instrument. The spectra were recorded in
deuteriochloroform (CDCl3) or hexadeuteriodimethylsulfoxide
(DMSO-d6). Chemical shifts of protons are given in parts per million
with TMS as internal standard. The chemical shifts of carbons are
reported in parts per million with solvent peaks used as internal
standards. Dilution and titration experiments with NMR were
carried out at 296 K. ESI spectra were recorded on an API 150EX. The
complex structures of 9-BuA with the receptors FR-1 and FR-2 were
calculated with Spartan’06 from Wavefunction, Inc.29
X-ray structures were determined using a Rigaku R-AXIS SPIDER
system. Structure solution was accomplished with SIR-9231 while
structure refinement was performed using SHELX-97.32 Thermal
ellipsoid drawings of the molecular structures were generated with
PLATON.33
Fluorescence spectra were measured on a FluoroMax-3 from
Horiba Jobin Yvon. The general setting was increment 1, integration
0.5, slit widths 3 (excitation) and 3 (emission) for both excitation
and emission fluorescence spectra. All fluorescence spectra were
recorded at 296 K and maintained using a circulating water bath.
The fluorescence cuvette used was made of quartz with a volume of
4 mL. The final volume of solutions for all the measurements was
2 mL. The excitation wavelength was 350 nm.
Figure 13. FR-2’s fluorescence change from greenish blue (left, FR-2, 0.343 mM) to
deep blue (right, complex of FR-2 with 9-BuA, FR-2, 0.343 mM, 9-BuA, 3.43 mM).
emission at 441 nm of the complex FR-2:9-BuA.19 The observed
changes in pyrene fluorescence behavior are indicative of the
binding of 9-BuA by the receptors.19
4.2. 1-(2-Bromophenyl)pyrene (2)
To a three-neck round bottom flask were added potassium
carbonate (0.46 g, 3.3 mmol) and water (2.2 mL). The mixture was
purged with nitrogen for 10 min. To the resulting mixture were
added 2-bromoiodobenzene (0.283 g, 1.0 mmol), 1-pyrenyl boronic
acid (0.246 g, 1.0 mmol), and 1,2-dimethoxyethane (5 mL) under
nitrogen with stirring. After 2 min, the catalyst Pd(PPh3)4 (37 mg,
0.035 mmol, 3.5% molar amount) was added. After 5 min, the
resulting mixture was heated at reflux in an oil bath at 120 ꢂC for
14 h. After cooling to room temperature, water (5 mL) was added to
the reaction and it was extracted with dichloromethane (3ꢃ5 mL).
After evaporation of the solvents in vacuo, the residue was purified
by column chromatography using silica gel with hexane as eluent,
affording intermediate 2, 1-(2-bromophenyl)pyrene (0.190 g white
crystals, 47%), after removal of solvent. Mp 129–130 ꢂC. IR (KBr,
cmꢁ1): 513, 629, 650, 682, 721, 752, 847,1005,1023,1047,1115,1175,
1192, 1243, 1422, 1434, 1455, 1466, 1558, 1601, 2854, 3039. 1H NMR
3. Conclusion
In order to mimic pyrene dinucleotide, the receptors FR-1 and
FR-2 were designed and synthesized. Their binding substrate 9-
butyladenine was also synthesized. The three compounds were
crystallized and their crystal structures determined. Both receptors
FR-1 and FR-2 form dimers through hydrogen bonding between
carboxyl groups of two molecules. In the FR-1 dimer, the carboxyl
group is in the same plane as the phenyl group while the carboxyl
group is vertical to the naphthyl group in the FR-2 dimer. In the FR-
1 dimer, pyrenyl moieties are too far separated to interact. On the
other hand, the pyrenyl groups in the FR-2 dimer are parallel with
a distance of 6.79 Å. The crystal structure of 9-butyladenine shows
that it forms an extensive hydrogen bonding network.
The molecular recognition of 9-butyladenine with receptors
FR-1 and FR-2 established that the binding constant for complex
formation of 9-BuA with FR-2 is 8.3-fold greater than with FR-1. The
binding of 9-butyladenine by the receptor FR-2 is more energeti-
(400 MHz, CDCl3, ppm)
d 7.37 (m, 1H), 7.48 (m, 2H), 7.75 (d,
J¼9.2 Hz, 1H), 7.83 (d, J¼8.8 Hz, 1H), 7.92 (d, J¼7.6 Hz, 1H), 8.02 (m,
2H), 8.12 (s, 2H), 8.22 (m, 3H). 13C NMR (100 MHz, CDCl3, ppm)
cally favorable than by FR-1 by 1.2 kcal/mol, suggesting
p–p
d
124.3, 124.7, 124.7, 124.8, 125.1, 125.3, 125.4, 126.2, 127.4, 127.5,
stacking and hydrogen bonding interactions are stronger in the
complexation of 9-butyladenine with FR-2 than FR-1. These results
indicate that the receptor FR-2 is a better mimic for pyrene di-
127.5, 127.8, 128.8, 129.4, 131.0, 131.2, 131.5, 132.4, 132.9, 136.5, 141.8.
HR-ESI-TOF calcd for C22H14Brþ (MþH) 357.027, found 357.028.
nucleotide. The energy stabilization from p–p stacking is estimated
4.3. 2-(1-Pyrenyl)benzoic acid (FR-1)
to be 2.3 kcal/mol in the complex FR-2:9-BuA. Fluorescence spec-
troscopic studies indicate that the binding of 9-BuA by FR-1 results
in a decrease of fluorescence intensity. In contrast, molecular rec-
ognition of 9-BuA by receptor FR-2 results in a blue shift in emission
wavelength and an increase of the fluorescence intensity.
To a one-neck round bottom flask were added 2 (0.122 g,
0.34 mmol) and THF (2 mL). The mixture was cooled to ꢁ78 ꢂC. To
the chilled solution was added n-butyllithium (0.136 mL of 2.5 M in
hexane, 0.34 mmol) with stirring. After 5 min, dry ice (0.03 g,
0.7 mmol) was added. The reaction mixture was then warmed to
room temperature gradually. The solvent was removed in vacuo, the
residue dissolved in sodium carbonate (0.5 M, 40 mL), and extrac-
ted with diethyl ether (2ꢃ20 mL) to remove impurities. The re-
sultant aqueous solution was acidified with hydrochloric acid (6 N)
to pH¼2. The final product FR-1 was obtained by filtration, washing
with water, and drying to constant weight (0.064 g, white solid,
58%). Mp 244–245 ꢂC. UV–vis (CHCl3, nm) lmax 269, 279, 331, 346.
IR (KBr, cmꢁ1): 495, 563, 632, 651, 683, 725, 753, 763, 848, 932,
4. Experimental
4.1. General
All chemicals were purchased from Sigma–Aldrich and used
without further purification. Flash chromatography was performed
with silica gel (70–230 mesh from Sorbent Technologies) and
monitored by thin layer chromatography (TLC) with silica gel plates

68
B. Lamale et al. / Tetrahedron 65 (2009) 62–69
1071, 1133, 1264, 1306, 1403, 1482, 1483, 1604, 1680, 1697, 2816,
2862, 3038. 1H NMR (400 MHz, DMSO-d6, ppm)
evaporation of most of the solvent, the precipitate was collected by
filtration. The crude product was recrystallized from toluene (5 mL)
and then 10% ethanol in water (5 mL) to give pure product (110 mg,
51%). Mp 136–138 ꢂC (lit.34 116 ꢂC). UV–vis (CHCl3, nm) lmax 261. IR
(KBr, cmꢁ1): 1029, 1071, 1491, 1596, 1676, 3286 (br, NH2). 1H NMR
d
7.50 (d, J¼7.6 Hz,
1H), 7.71 (m, 3H), 7.91 (m, 1H), 8.10 (m, 3H), 8.22 (s, 2H), 8.26 (d,
J¼7.6 Hz, 1H), 8.32 (d, J¼7.2 Hz, 2H), 12.48 (s, 1H). 13C NMR
(100 MHz, DMSO-d6, ppm) d 124.3, 124.6, 125.0, 125.2, 125.5, 125.8,
126.9, 127.6, 127.8, 128.00, 128.4, 128.7, 130.4, 130.5, 130.9, 131.5,
132.0, 132.7, 133.3, 137.9, 141.1, 168.8. ESI-MS calcd for C23H14O2Naþþ
(MþNa), 345.3; found, 345.1. HR-ESI-TOF calcd for C23H15O2
(MþH) 323.107, found 323.107.
(400 MHz, CDCl3, ppm)
d
0.96 (t, 3H, J¼7.6 Hz, CH3), 1.36 (m, 2H,
CH2), 1.87 (m, 2H, CH2), 4.20 (t, 2H, J¼7.2 Hz, CH2), 5.71 (bs, 2H,
NH2), 7.79 (s, 1H, Ar-H), 8.37 (s, 1H, Ar-H). 13C NMR (100 MHz,
CDCl3, ppm) d 13.8, 20.1, 32.3, 43.9, 119.9, 140.7, 150.4, 153.2, 155.6.
Mass (ESI) cal. for C9H14N5þ (MþH), 192.2; found, 192.4, and
4.4. 1-(8-Bromo-1-naphthyl)pyrene (4)
C9H13N5Naþ (MþNa), 214.2; found 212.4.
To a three-neck round bottom flask were added potassium
carbonate (0.46 g, 3.3 mmol) and water (2.2 mL). The mixture was
purged with nitrogen for 10 min. To the resulting mixture were
added 1,8-dibromonaphthalene (0.286 g, 1.0 mmol), 1-pyrenyl
boronic acid (0.246 g, 1.0 mmol), and 1,2-dimethoxyethane (5 mL)
under nitrogen with stirring. After 2 min, the catalyst Pd(PPh3)4
(37 mg, 0.035 mmol, 3.5% molar amount) was added and the mix-
ture stirred for 5 min. The resulting mixture was then heated to
reflux in an oil bath at 120 ꢂC for 14 h. After cooling to room tem-
perature, water (5 mL) was added to the mixture and extracted
with dichloromethane (3ꢃ5 mL). The residue from evaporation of
the solvents in vacuo was purified by column chromatography us-
ing silica gel with 1–3% ethyl acetate (volume) in hexane as eluent,
affording intermediate 4, 1-(8-bromo-1-naphthyl)pyrene (0.219 g
white solid, 54%), after removal of solvent. Mp 154–156 ꢂC. IR (KBr,
cmꢁ1): 493, 541, 721, 766, 821, 844, 1119, 1181, 1197, 1244, 1320,
1363, 1441, 1493, 1561, 2921, 3047. 1H NMR (400 MHz, CDCl3, ppm)
4.7. Determination of the crystal structures of receptors FR-1,
FR-2, and 9-BuA by X-ray diffraction
Crystallographic data (excluding structure factors) for structures
FR-1, FR-2, and 9-BuA in this paper have been deposited with the
Cambridge Crystallographic Data Centre as supplementary publi-
cation nos. CCDC 696757, 696758, and 697622, respectively. Copies
of the data can be obtained free of charge, on application to CCDC,
12 Union Road, Cambridge CB2 1EZ, UK (fax: þ44 (0) 1223 336033
or e-mail: deposit@ccdc.cam.ac.uk). Abbreviated crystallographic
data are given below.
FR-1. C23H14O2, MW¼322.345, monoclinic, P21/c, a¼9.2500 Å,
b¼7.0290 Å, c¼25.8570 Å,
a
¼90.00ꢂ,
b
¼101.11ꢂ,
g
¼90.00ꢂ, V¼1649.7
(13) Å3, Z¼4, T¼223 K,
m
¼0.082 mmꢁ1
,
rcalcd¼1.394 Mg mꢁ3, GOF on
F2¼1.100, R¼0.0602, Rw¼0.1465 [I>2
s(I)].
FR-2. C27H16O2, MW¼372.405, monoclinic, P21/n, a¼12.742(3) Å,
b¼11.796(3) Å, c¼13.476(3) Å,
a
¼90ꢂ,
b
¼114.070(5)ꢂ,
g
¼90ꢂ,
V¼1849.3(8), Z¼4, T¼293 K,
m
¼0.083 mmꢁ1
,
rcalcd¼1.338 Mg mꢁ3
,
d
7.33 (t, J¼7.6 Hz, 1H), 7.60 (m, 3H), 7.73 (d, J¼7.2 Hz), 7.90–8.23 (m,
10H). 13C NMR (100 MHz, CDCl3, ppm)
d
120.3, 124.0, 124.5, 124.9,
GOF on F2¼1.091, R¼0.0609, Rw¼0.1309 [I>2
s(I)].
125.0, 125.2, 125.6, 125.9, 126.0, 126.4, 127.4, 127.7, 128.7, 129.3,
129.6, 130.8, 130.9, 131.1, 131.6, 132.4, 133.9, 136.1, 138.3, 138.7. HR-
ESI-TOF calcd for C26H16Brþ (MþH) 407.043, found 407.042.
9-BuA. C19H15N5O (9-BuA hydrate), MW¼209.26, monoclinic, P21/
c, a¼10.981(4) Å, b¼8.240(3) Å, c¼12.733(5) Å,
a
¼90ꢂ,
b
¼109.131(9)ꢂ,
g
¼90ꢂ, V¼1088.5(7), Z¼4, T¼223 K,
m
¼0.089 mmꢁ1
,
rcalcd
¼
1.277 Mg mꢁ3, GOF on F2¼1.097, R¼0.0619, Rw¼0.1789 [I>2
s(I)].
4.5. 8-(1-Pyrenyl)-1-naphthoic acid (FR-2)
Acknowledgements
To a one-neck round bottom flask were added 4 (0.144 g,
0.35 mmol) and THF (2 mL). The mixture was cooled to ꢁ78 ꢂC. To
the resulting cold solution was added n-butyllithium (0.22 mL of
1.6 M in hexane, 0.35 mmol) with stirring. After 5 min, dry ice
(0.031 g, 0.70 mmol) was added. The reaction mixture was then
warmed to room temperature gradually. The solvent was removed
in vacuo, the residue dissolved in sodium carbonate (0.5 M,
2ꢃ20 mL), and extracted with diethyl ether (2ꢃ20 mL) to remove
impurities. The resulted aqueous solution was acidified with
hydrochloric acid (6 N) to pH¼2. The final product FR-2 was
obtained by filtration, washing with water, and drying to constant
weight (0.063 g off-white solid, 48%). Mp >260 ꢂC, decomposed.
UV–vis (CHCl3, nm) lmax 270, 281, 349. IR (KBr, cmꢁ1): 496, 639,
682, 722, 773, 846, 1155, 1183, 1210, 1293, 1413, 1460, 1503, 1692,
This research is supported by the Office of Research and Spon-
sored Programs (RD 09010) and an SFRA Grant for B.L. and Y.L.J.
from the Honors College at East Tennessee State University.
Supplementary data
Dimerization constant determination, Job’s plots, binding con-
stant determination, structural information (hydrogen bonding and
p–p stacking interactions) from calculations using Spartan’06,
calculated complex structures using the Spartan’06, spectral data
(1H NMR and 13C NMR) of the two receptors, their synthesis in-
termediates and 9-butyladenine are provided. Supplementary data
associated with this article can be found in the online version, at
doi:10.1016/j.tet.2008.10.041.
3039. 1H NMR (400 MHz, DMSO-d6, ppm)
d
7.57–8.32 (m, 15H), 11.76
124.7, 124.8, 125.4,
(s, 1H). 13C NMR (100 MHz, DMSO-d6, ppm)
d
125.6,125.7,126.2,126.4,126.7,127.6,127.7,128.1,128.4,129.0,129.1,
129.2, 129.5, 130.7, 131.2, 131.5, 131.6, 132.6, 133.7, 135.0, 137.9, 138.2,
170.2. ESI-MS calcd for C27H16O2Naþ (MþNa), 395.4; found, 395.1.
HR-ESI-TOF calcd for C27H17O2þ (MþH) 373.122, found 373.122.
References and notes
1. Matray, T. J.; Kool, E. T. Nature 1999, 399, 704–708.
2. Wilson, J. N.; Cho, Y.; Tan, S.; Cuppoletti, A.; Kool, E. T. ChemBioChem: 2008, 9,
279–285.
3. Kwon, K.; Jiang, Y. L.; Stivers, J. T. Chem. Biol. 2003, 10, 351–359.
4. Honcharenko, D.; Zhou, C.; Chattopadhyaya, J. J. Org. Chem. 2008, 73, 2829–
2842.
4.6. Synthesis of 9-BuA
5. Johansson, D.; Jessen, C. H.; Pohlsgaard, J.; Jensen, K. B.; Vester, B.; Pedersen, E.
B.; Nielsen, P. Bioorg. Med. Chem. Lett. 2005, 15, 2079–2083.
6. Jiang, Y. L.; Kwon, K.; Stivers, J. T. J. Biol. Chem. 2001, 276, 42347–42354.
7. Jiang, Y. L.; Stivers, J. T.; Song, F. Biochemistry 2002, 41, 11248–11254.
8. Guckian, K. M.; Schweitzer, B. A.; Ren, R. X.-F.; Sheils, C. J.; Paris, P. L.; Kool, E. T.
J. Am. Chem. Soc. 1996, 118, 8182–8183.
Adenine (302 mg, 2.25 mmol) was dissolved in hot DMF
(20 mL). To the warm solution were added freshly prepared
1-iodobutane (205 mg, 1.12 mmol) and cesium carbonate (1.94 g,
3.36 mmol). The resulting mixture was stirred at 50 ꢂC overnight.
The DMF solvent was removed in vacuo. The residue was boiled
with toluene (50 mL) for 5 min and the hot solution filtered. After
9. Guckian, K. M.; Schweitzer, B. A.; Ren, R. X.-F.; Sheils, C. J.; Tahmassebi, D. C.;
Kool, E. T. J. Am. Chem. Soc. 2000, 122, 2213–2222.

B. Lamale et al. / Tetrahedron 65 (2009) 62–69
69
10. Smirnov, S.; Matray, T. J.; Kool, E. T.; de los Santos, C. Nucleic Acids Res. 2002, 30,
5561–5569.
11. Harvey, R. G.; Lim, K.; Dai, Q. J. Org. Chem. 2004, 69, 1372–1373.
12. Amann, N.; Pandurski, E.; Fiebig, T.; Wagenknecht, H.-A. Chem.dEur. J. 2002, 8,
4877–4883.
26. Herrans, F.; Maria, M. D. S.; Claramunt, R. M. J. Org. Chem. 2006, 71, 2944–2951.
27. Tsukube, H.; Furuta, H.; Odani, A.; Takeda, Y,; Kudo, Y.; Inoue, Y.; Liu, Y.;
Sakamoto, H.; Kimura, K. In Comprehensive Supramolecular Chemistry; Atwood,
J. L., Davies, J. E. D., MacNicol, D. D., Vo¨gtle, F., Eds.; Pergamon: London, 1996;
Vol. 8, pp 425–482.
13. Zimmerman,S.C.;Zeng,Z.;Wu,W.;Reichert,D.E. J. Am. Chem. Soc.1991,113,183–196.
14. Klein, S. M.; Zhang, C.; Jiang, Y. L. Tetrahedron Lett. 2008, 49, 2638–2641.
15. Crisp, G. T.; Jiang, Y. L.; Tiekink, E. R. T. Z. Kristallogr. NCS 2000, 215, 83–84.
16. Benson, R. E.; Clearfield, A.; Daniels, L. M.; Wardeska, J. G. Acta Crystallogr. 2006,
E62, m696–m698.
17. Jiang, Y. L.; Daniels, L. M.; Lamale, B. Abstracts of Papers, 234th ACS National
Meeting, Boston, MA, Aug 19–23, 2007; American Chemical Society: Wash-
ington, DC, 2007, ORGN-200.
28. Rao, P.; Ghosh, K.; Maitra, U. J. Phys. Chem. 1999, 103, 4528–4533.
29. Shao, Y.; Fusti-Molnar, L.; Jung, Y.; Kussmann, J.; Ochsenfeld, C.; Brown, S. T.;
Gilbert, A. T. B.; Slipchenko, L. V.; Levchenko, S. V.; O’Neill, D. P.; Distasio, R. A.,
Jr.; Lochan, R. C.; Wang, T.; Beran, G. J. O.; Besley, N. A.; Herbert, J. M.; Lin, C. Y.;
Van Voorhis, T.; Chien, S. H.; Sodt, A.; Steele, R. P.; Rassolov, V. A.; Maslen, P. E.;
Korambath, P. P.; Adamson, R. D.; Austin, B.; Baker, J.; Byrd, E. F. C.; Daschel, H.;
Doerksen, R. J.; Dreuw, A.; Dunietz, B. D.; Dutoi, A. D.; Furlani, T. R.; Gwaltney,
S. R.; Heyden, A.; Hirata, S.; Hsu, C.-P.; Kedziora, G.; Khalliulin, R. Z.; Klunzinger,
P.; Lee, A. M.; Lee, M. S.; Liang, W.; Lotan, I.; Nair, N.; Peters, B.; Proynov, E. I.;
Pieniazek, P. A.; Rhee, Y. M.; Ritchie, J.; Rosta, E.; Sherrill, C. D.; Simmonett, A.
C.; Subotnik, J. E.; Woodcock, H. L., III; Zhang, W.; Bell, A. T.; Chakraborty, A. K.;
Chipman, D. M.; Keil, F. J.; Warshel, A.; Hehre, W. J.; Schaefer, H. F., III; Kong, J.;
Krylov, A. I.; Gill, P. M. W.; Head-Gordon, M. Phys. Chem. Chem. Phys. 2006, 8,
3172–3191.
18. Christensen, U. B.; Pedersen, E. B. Helv. Chim. Acta 2003, 86, 2090–2097.
19. Jun, E. J.; Won, H. N.; Kim, J. S.; Lee, K.-H.; Yoon, J. Tetrahedron Lett. 2006, 47,
4577–4580.
20. Rebek, J., Jr. Acc. Chem. Res. 1990, 23, 399–404.
21. Kim, S.; Schaefer, H. F. III. J. Phys. Chem. A 2007, 111, 10381–10389.
22. Quinn, J. R.; Zimmerman, S. C.; Del Bene, J. E.; Shavitt, I. J. Am. Chem. Soc. 2007,
129, 934–941.
30. Basaric, N.; Wan, P. J. Org. Chem. 2006, 71, 2677–2686.
23. Crisp, G. T.; Jiang, Y. L. Tetrahedron Lett. 2002, 43, 3157–3160.
24. Inouye, M.; Itoh, M. S.; Nakazumi, H. J. Org. Chem. 1999, 64, 9393–9398.
25. Faraoni, R.; Castellano, R. K.; Gramlich, V.; Diederich, F. Chem. Commun. 2004,
370–371.
31. Goswami, S.; Dey, S.; Jana, S. Tetrahedron 2008, 64, 6358–6363.
32. Sheldrick, G. M. Acta Crystallogr. 2008, A64, 112–122.
33. Spek, A. L. J. Appl. Crystallogr. 2003, 36, 7–13.
34. Ghosh, K.; Sen, T.; Froehlich, R. Tetrahedron Lett. 2007, 48, 7022–7026.