Anthryl-substituted heterocycles as acid-sensitive fluorescence probes

Article abstract of DOI:10.1021/jo047841z

Four pH-sensitive fluorescence probes are presented which consist of an anthracene fluorophore and a π-conjugated oxazoline, benzoxazole, or pyridine substituent. The protonation of the heterocycles increases their acceptor properties and results in signi

Full text of DOI:10.1021/jo047841z

Anthryl-Substituted Heterocycles as Acid-Sensitive Fluorescence
Probes
Heiko Ihmels,*,† Andreas Meiswinkel,‡ Christian J. Mohrschladt,‡ Daniela Otto,†
Michael Waidelich,† Michael Towler,§ Rick White,§ Martin Albrecht,| and
Alexander Schnurpfeil|,
Institut fu¨r Organische Chemie, Universita¨t Siegen, Adolf-Reichwein-Str., D-57068 Siegen, Germany,
Institut fu¨r Organische Chemie, Universita¨t Wu¨rzburg, Am Hubland, D-97074 Wu¨rzburg, Germany,
Department of Chemistry, Sam Houston State University, Huntsville, Texas 77341, and Institut fu¨r
Physikalische und Theoretische Chemie, Universita¨t Siegen, Adolf-Reichwein-Str.,
D-57068 Siegen, Germany
ihmels@chemie.uni-siegen.de
Received December 7, 2004
Four pH-sensitive fluorescence probes are presented which consist of an anthracene fluorophore
and a π-conjugated oxazoline, benzoxazole, or pyridine substituent. The protonation of the
heterocycles increases their acceptor properties and results in significant red-shifts of the absorption
and emission maxima of the anthracene chromophore. The comparison between 2-[2′-(6′-methoxy-
anthryl)]-4,4-dimethyl-2-oxazoline and 2-[2′-(anthryl)]-4,4-dimethyl-2-oxazoline reveals that the
donor-acceptor substitution pattern of the fluorophore is not required to achieve a red shift upon
protonation. The benzoxazole and pyridine substituents offer a particular advantage due to their
persistence under acidic conditions. Thus, these compounds may be used as efficient pH-sensitive
fluorescence switches. Nevertheless, the switching of benzoxazole 2c requires relatively strong acidic
conditions. The anthrylpyridinium exhibits a red-shifted emission in chloroform; however, it is
nonfluorescent in aqueous or alcoholic solution. Although the oxazoline is not persistent under
permanent acidic conditions, this heterocycle may be useful as a substituent in fluorescence
indicators since it may be used to detect acid concentrations of 10-4-10-5 M, which are close to the
biologically relevant range.
Introduction
organic fluorophores exhibit a high potential to be part
of fluorescence probes, since they offer the possibility to
connect a fluorophore directly with a particularly de-
signed functionality whose interaction with the analyte
may have a significant influence on the emission proper-
ties such as emission intensity, wavelength, or lifetime.
A survey of known fluorescence probes and switches,
however, reveals that in the majority of systems the
fluorescence intensity, i.e., the decrease or increase of
emission quantum yield, is used to monitor the interac-
tions with an external stimulus or analyte. Especially in
the case of fluorescence quenching, this process is not
Fluorescence spectroscopy has developed to one of the
most important tools in analytical biology and medicine.1
In particular, the high sensitivity of this method (even
down to the single molecule)2 is useful for the detection
of analytes at low concentrations. Along these lines,
† Institut fu¨r Organische Chemie, Universita¨t Siegen.
‡ Institut fu¨r Organische Chemie, Universita¨t Wu¨rzburg.
§ Department of Chemistry, Sam Houston State University.
| Institut fu¨r Physikalische und Theoretische Chemie, Universita¨t
Siegen.
Present address: Institut fu¨r Theoretische Chemie, Universita¨t
Ko¨ln, Greinstr. 4, D-50939 Ko¨ln, Germany.
10.1021/jo047841z CCC: $30.25 © 2005 American Chemical Society
Published on Web 04/13/2005
J. Org. Chem. 2005, 70, 3929-3938
3929

Ihmels et al.
SCHEME 1a
a Key: (a) CO2Cl2, reflux, 18 h; (b) 2-amino-2-methylpropanol, CH2Cl2, rt, 2 h; (c) SOCl2, reflux 2 h; (d) 2-aminophenol, pyridine,
100 °C, 1 h, then 200 °C, 30 min.
very selective, since quenching may also be caused by
other external factors. Therefore, fluorescence probes are
desirable whose emission wavelength changes signifi-
cantly in the presence of an analytesas realized in
fluorescent ED-π-EA systems. In most donor-acceptor-
substituted fluorescence probes, the protonation or com-
plexation of donor substituents leads to a decrease of
their donor ability and to a corresponding blue shift of
the absorption and emission maxima. This concept is very
often realized with amino functionalities or crown and
aza-crown substituents as donor moiety in a donor-
acceptor-substituted chromophore.1c-e In contrast, ex-
amples for complementary fluorescence probes and
switches are rather rare, in which the modification of
acceptor functionalities leads to a red-shift of the absorp-
tion and emission maxima,3 presumably because there
are only few acceptor functionalities which offer the
appropriate properties. Some examples are known in
which a heterocycle with a CN double bond is used as
tunable acceptor moiety, but to the best of our knowledge
a systematic investigation has not been performed.3c-f It
may also be noted that fluorescence probes are very often
used successfully to detect metal cations or inorganic and
organic anions; however, the number of efficient proton-
sensitive fluorescence probes is comparably small.4 In
preliminary work, we have shown that the protonation
of the anthryloxazoline 2b results in a significant red
shift of its emission from 444 to 540 nm.5 In this paper,
we present our studies on the scope and limits of this
probe system on the basis of alkaline anthryl-substituted
heterocycles.
Results
Synthesis. The anthryl-substituted oxazolines 2a and
2b as well as the benzoxazole 2c were synthesized
according to standard procedures (Scheme 1), starting
from anthracene-2-carboxylic acid (1a) or 6-methoxyan-
thracene-2-carboxylic acid (1b).6 Thus, the transforma-
tion of carboxylic acids 1a or 1b to the corresponding acid
chlorides, reaction with 2-methyl-2-aminobutanol, and
subsequent cyclization with thionyl chloride gave the
oxazolines 2a and 2b in 36% and 60% yield. The ben-
zoxazole 2c was obtained in rather low yield (34%) by
the reaction of the acid chloride with 2-aminophenol
(Scheme 1). The pyridyl-substituted anthracene 7 was
synthesized from 2-(4′-methoxyphenyl)-4,4-dimethyl-2-
oxazoline (3),7 which was ortho-lithiated with n-BuLi
followed by the addition of 4-(2′-pyridyl)benzaldehyde to
yield the diarylmethanol derivative 4 quantitatively. The
latter was treated with p-toluenesulfonic acid to give the
phthalide 5 in 62% yield (Scheme 2). Reduction with zinc
dust in acetic acid gave the benzoic acid derivative 6,
which was cyclized with methanesulfonic acid and sub-
sequently reduced to yield the anthracene 7. Unfortu-
nately, the cyclization-reduction sequence from 6 to 7
proceeds in low yields (21% over both steps), and at-
tempts to optimize the reaction conditions did not lead
to higher yields. All new compounds were characterized
by 1H and 13C NMR spectroscopy, mass spectrometry, and
elemental analysis or high-resolution mass spectrometry.
Photophysical Properties. The absorption proper-
ties of compounds 2a-c and 7 are characteristic of
substituted anthracene derivatives (Table 1, data of 2a
not shown); i.e., short-wavelength absorption bands (S0-
S3 transition)8 appear between 250 and 300 nm and long-
wavelength absorption bands (p band: S0-S1 transition)
between 300 and 430 nm. Moreover, the absorption bands
of the donor-acceptor system 2b are shifted ca. 30 nm
to longer wavelength as compared to 2a due to the
presence of the additional donor substituent which is
linearly conjugated with the acceptor moiety. In all cases,
there is no long-wavelength absorption, which reveals the
absence of an ICT absorption. For all compounds, the
absorption properties are almost independent from the
solvent (Table 1).
(1) (a) Optical Sensors; Wolfbeis, O. S., Ed.; Springer-Verlag: Heidel-
berg, 2004. (b) Mohr, G. J. Chem. Eur. J. 2004, 10, 1082-1090. (c) Pu,
L. Chem. Rev. 2004, 104, 1687. (d) Martinez-Manez, R.; Sancenon, F.
Chem. Rev. 2003, 103, 4419. (e) Wiskur, S. L.; Ait-Haddou, H.; Anslyn,
E. V.; Lavigne, J. J. Acc. Chem. Res. 2001, 34, 963. (f) Chemosensors
of Ion and Molecular Recognition; Desvergne, J.-P., Czarnik, A. W.,
Eds.; Kluwer Academic Press: Dordrecht, The Netherlands, 1997. (g)
de Silva, A. P. H.; Gunaratne, Q.; Gunnlaugsson, T.; Huxley, A. J. M.;
McCoy, C. P.; Rademacher, J. T.; Rice, T. E. Chem. Rev. 1997, 97, 1515.
(h) Fluorescent Chemosensors for Ion and Molecule Recognition;
Czarnik, A. W., Ed.; ACS Symposium Series No. 538; American
Chemical Society: Washington DC, 1993. (i) Valeur, B. In Topics in
Fluorescence Spectroscopy, Vol. 4: Probe Design and Chemical Sensing;
Lakowicz, J. R., Ed.; Plenum Press: New York, 1994; Chapter 2.
(2) (a) Sauer, M. Angew. Chem. 2003, 115, 1834; Angew. Chem., Int.
Ed. Engl. 2003, 42, 1790. (b) Ambrose, W. P.; Goodwin, P. M.; Jett, J.
H.; van Orden, A.; Werner, J. H.; Keller, R. A. Chem. Rev. 1999, 99,
2929. (c) Ha, T.; Chemla, D. S.; Enderle, T.; Weiss, S. Bioimaging 1997,
55, 99.
(3) (a) Bourson, J.; Pouget, J.; Valeur, B. J. Phys. Chem. 1993, 97,
4552. (b) Dix, J. P.; Vo¨gtle, F. Chem. Ber. 1981, 14, 638. (c) Jones, G.,
II; Jiminez, J. A. C. J. Photochem. Photobiol. B: Biology 2001, 65, 5.
(d) Fayed, T. A. J. Photochem. Photobiol. A: Chemistry 1999, 121, 17.
(e) Corrent, S.; Hahn, P.; Pohlers, G.; Connolly, T. J.; Scaiano, J. C.;
Fornes, V.; Garcia, H. J. Phys. Chem. B 1998, 102, 5852. (f) Wolfbeis,
O. S.; Marhold, H. Chem. Ber. 1985, 118, 3664.
(4) For recent examples see: (a) Charier, S.; Ruel, O.; Baudin, J.-
B.; Alcor, D.; Allemand, J.-F.; Meglio, A.; Jullien, L. Angew. Chem.
2004, 43, 4785; Angew. Chem., Int. Ed. Engl. 2004, 116, 4889. (b) Cui,
D.; Quian, X.; Liu, F.; Zhang, R. Org. Lett. 2004, 6, 2757. (c) Diwu, Z.;
Chen, C.-S.; Zhang, C.; Klaubert, D. H.; Haugland, R. P. Chem. Biol.
1999, 6, 411. (d) Kermis, H. R.; Kostov, Y.; Harms, P.; Rao, G. Biotech.
Prog. 2002, 18, 1047. (e) Garreis, T.; Huber, C.; Wolfbeis, O. S.; Daub,
J. J. Chem. Soc., Chem. Commun. 1997, 1717.
(5) Ihmels, H.; Meiswinkel, A.; Mohrschladt, C. J. Org. Lett. 2000,
2, 2865.
(6) Ihmels, H. Eur. J. Org. Chem. 1999, 1595.
(7) Mohan, R.; Katzenellenbogen, J. A. J. Org. Chem. 1984, 49, 1238.
(8) Dick, B.; Hohlneicher, G. Chem. Phys. Lett. 1981, 83, 615.
3930 J. Org. Chem., Vol. 70, No. 10, 2005

Heterocycles as Acid-Sensitive Fluorescence Probes
SCHEME 2a
a Key: (a) n-BuLi (1.6 M in hexane), Et2O, 0 °C, 4 h; (b) 4-(2′-pyridyl)benzaldehyde; Et2O, rt, 16 h (97% over both steps); (c) p-TsOH,
toluene, reflux, 2.5 h (62%); (d) Zn, CH3COOH, reflux, 24 h (75%); (e) CH3SO3H, 90 °C, 4 h; (f) NaBH4, 2-PrOH, reflux, 4 d (21% over both
steps).
TABLE 1. Absorption and Emission Data (λ in nm) of the Anthracene Derivatives 2b, 2c, and 7 (Solvents in Order of
Decreasing ET(30) Value)
2b
log ꢀ
2c
log ꢀ
7
log ꢀ
3.73
a
a
a
solvent
λabs
λfl b (φfl)c
421e
(0.30)
430
λabs
λfl b (φfl)c
433f
(0.49)
442
(0.43)
452
(0.44)
453
(0.48)
458
(0.45)
461
(0.43)
456
λabs
λfl d (φfl)c
426g
(0.26)
434
(0.28)
438
(0.26)
c-C6H12
393
396
395
394
396
397
392
395
3.78
3.66
3.64
3.63
3.64
3.64
3.59
3.61
404
407
406
405
407
409
404
403
4.05
3.99
3.97
3.97
3.97
3.97
3.95
4.01
398
402
401
benzene
CH2Cl2
1-BuCN
DMF
3.72
3.69
438
(0.49)
435
(0.42)
441
(0.50)
444
(0.58)
439
(0.42)
444
(0.48)
401
404
398
398
3.65
3.70
3.67
3.70
441
(0.35)
444
(0.34)
438
(0.29)
440
(0.28)
DMSO
MeCN
MeOH
(0.45)
462
(0.43)
a Maximum at lowest energy; c ) 10-4 M. b Emission maximum; c ) 10-5 M, λex ) 390 nm. c Relative to quinine sulfate in 1 N H2SO4.
d Emission maximum, c ) 10-5 M, λex ) 380 nm. e Additional emission band at λ ) 442. f Additional emission bands at λ ) 456 nm.
g Additional emission band at λ ) 408 and 451 nm.
The emission spectra of compounds 2a-c and 7 exhibit
very broad, mostly structureless bands. In contrast to the
absorption data, the emission properties of 2b, 2c, and
7 show a moderate dependence on the solvent properties.
In general, a correlation of the emission wavelength with
the solvent polarity was observed, as expressed by the
empirical parameter ET(30).9 Thus, a red shift of the
emission band results from an increasing solvent polarity.
This effect is especially pronounced for benzoxazole 2c
(Figure 1) and leads to the conclusion that the dipole
moment of the excited state is larger than the one of the
ground state.10 Notably, emission data in methanol do
not fit into the correlation, presumably because along
FIGURE 1. Correlation of the emission maxima (in cm-1) of
with the dipole-dipole interactions between solvent and
2b (O, r2 ) 0.88), 2c (9, r2 ) 0.94), and 7 (+, r2 ) 0.82) with
chromophore hydrogen bonding may have an influence
on the emission properties in methanol.
Dependence of the Electronic Spectra on the
Proton Concentration. Upon acidification of the oxa-
the solvent parameter ET(30) (for the r2 values, methanol was
excluded).
zolines 2a and 2b or pyridine 7 with hydrochloric acid
in methanol solution, a remarkable shift of the absorption
maximum was observed along with a significant loss of
the absorption-band structure (Figures 2-4 and Table
(9) Reichardt, C. Chem. Rev. 1994, 94, 2319.
(10) Suppan, P.; Ghoneim, N. Solvatochromism; The Royal Society
of Chemistry: London, 1997.
J. Org. Chem, Vol. 70, No. 10, 2005 3931

Ihmels et al.
FIGURE 4. Spectrophotometric titration of 7 (10-4 M in
MeOH) with aqueous HCl; arrows indicate the development
of band maxima during titration. Inset: change of absorption
intensity at 360 and 400 nm at varying acid concentrations.
FIGURE 2. Spectrophotometric titration of 2a (10-4 M in
MeOH) with aqueous HCl; arrows indicate the development
of band maxima during titration. Inset: change of absorption
intensity at 355, 390, and 400 nm at varying acid concentra-
tions.
TABLE 2. Emission Data and pKa Values from
Unprotonated and Protonated Anthracene Derivatives
2a-c and 7
b
λfl (no H+) (nm)
λfl (+H+) (nm)
pKa
pKa * c
2aa
2ba
2ca
7a
426d
450e
469e
443f
517d
544e
567e
540f
3.2
3.5
<1
2.7g
10.2
10.1
9.1
a c ) 10-5 M. b Determined from spectrophotometric titration
in MeOH with aq HCl, error ( 0.1. c Calculated according to
Fo¨rster cycle: ∆pK ) 0.00209 [ν˜ (base) - ν˜ (base-H+)]/cm-1 d λex
.
e
f
) 363 nm. λex ) 380 nm. λex ) 370 nm. g Same pK in CHCl3
with trifluoroacetic acid.
7 are significantly less acidic in the excited state as
compared to the ground state (∆pKa of ca. 7).
In contrast, spectrophotometric titrations of acid to
benzoxazole 2c have no significant influence on the
absorption spectrum at moderate acid concentration
(>10-1 M). Nevertheless, at larger acid concentrations,
a decrease of the long-wavelength absorption band was
observed along with the formation of a shoulder from 430
to 500 nm (Figure 5).
The emission maximum of oxazolines 2a and 2b in
methanol solution was significantly quenched upon ad-
dition of acid, and a new maximum appeared at λ ) 517
nm and λ ) 544 nm, respectively (Figures 6 and 7).
Further acidification led to a decrease of the short-
wavelength fluorescence and to an increase of the red-
shifted emission. It was also shown exemplarily for 2b
that the same effect may be achieved in water solution
(Figure 7B); however, the low solubility of the anthracene
derivative requires lower concentrations (10-6 M). Nota-
bly, the change of emission color may be followed by the
naked eye in each case (Figure 8). On neutralization of
the solution, the initial emission spectrum of anthracene
2b was regained. In principle, the benzoxazole 2c exhibits
the same emission behavior as 2a and 2b upon spectro-
fluorimetric titration with hydrochloric acid (Figure 9);
FIGURE 3. Spectrophotometric titration of 2b (10-4 M in
MeOH) with aqueous HCl; arrows indicate the development
of band maxima during titration. Inset: change of absorption
intensity at 372 and 385 nm at varying acid concentrations.
2, cf. Supporting Information for magnified Figures). In
each case, a plot of the absorption at a fixed wavelength
versus the proton concentration exhibits the character-
istic features of a spectrophotometric acid-base titration.
From these isotherms, the pKa values of the conjugated
acids were estimated to be 3.2 (2a), 3.5 (2b), and 2.7 (7)
under these conditions (Table 2). The pKa values of 2a-c
and 7 differ about 2-3 orders of magnitude from the ones
reportedforoxazoline,pyridine,andbenzoxazolederivatives;3d,11
however, the latter have been determined in water,
whereas the data reported herein were determined in
methanol solutions, in which pK values are usually
smaller than in water.12 The excited-state pKa, i.e., pKa*,
were estimated from Fo¨rster-cycle calculations (Table 2).
Thus, as usually observed for N-heterocycles,13 the pro-
tonated oxazolines 2a and 2b and the protonated pyridine
(11) Albert, A. Heterocyclic Chemistry, 2nd ed.; Athlone Press:
London, 1968; Chapter 13.
(12) Reichardt, C. Solvents and Solvent Effects on Organic Chem-
istry, 3rd ed.; Wiley-VCH: Weinheim, 2003; Chapter 4.2.
(13) Ireland, J. F.; Wyatt, P. A. H. Adv. Phys. Org. Chem. 1976, 12,
131.
3932 J. Org. Chem., Vol. 70, No. 10, 2005

Heterocycles as Acid-Sensitive Fluorescence Probes
of the heterocyclic moiety of anthracenes 2a, 2b, and 7
transforms them in stronger acceptor functionalities and
results in a significant red shift of the absorption band
which is assigned to an ICT absorption. The oxazolines
2a and 2b as well as the pyridine 7 are protonated at
moderate acid concentrations, whereas the aromatic
benzoxazole 2c requires significantly higher acid con-
centrations for protonation. These observations are in
good agreement with the reported pKa data for parent
compounds and derivatives thereof.3d,11 Preliminary in-
terpretations of absorption and emission data with the
protonated anthryloxazoline 2b suggested that the re-
sulting “oxazolinium substituent” has a higher π-acceptor
strength than the oxazoline, so that a stronger donor-
acceptor interplay with the conjugated methoxy func-
tionality is established, along with the corresponding red-
shifted absorption and emission maxima relative to the
ones of nonprotonated 2b. However, the same effect was
observed for the oxazoline 2a, which does not have a
conjugated methoxy group attached to the anthracene
fluorophore. This somewhat surprising result reveals that
the donor substituent in the 6 position, and thus a
conjugated donor-acceptor system in general, is not
necessarily required to achieve the drastic red shift upon
protonation. Moreoever, this observation is in agreement
with studies which show that anthracene moieties may
act as efficient electron donors in photoinduced charge-
transfer processes.14
FIGURE 5. Spectrophotometric titration of 2c (10-4 M in
MeOH) with aqueous HCl; arrows indicate the development
of band maxima during titration. Inset: change of absorption
intensity at 380 and 410 nm at varying acid concentrations.
The proposed qualitative explanations of the photo-
physical properties of compounds 2a and 2b along with
the ones of their protonated forms were supported by
density functional theory (DFT) ab initio calculations on
slightly varied systems 2a′ and 2b′. To keep the compu-
tational effort feasible, the two methyl substituents on
the oxazoline moiety were omitted. Note that the purpose
of the calculations was not to simulate the experimental
data including all specific and nonspecific interactions
with the respective solvent but to analyze the observed
trends in the absorption and emission spectra. In each
calculation, the structure was first obtained from a DFT
geometry optimization employing the RI-DFT method
with the B-P86 functional and a cc-pVTZ basis set using
the program package TURBOMOL.15 Subsequently, the
difference between electron affinity and ionization po-
tential was calculated employing GAUSSIAN0316 with
a B3LYP functional in a 6-311+G** basis set. The
resulting difference was used to estimate the HOMO-
LUMO gap and the corresponding absorption (and emis-
sion) wavelength. Thus, for 2a′ and 2b′ a HOMO-LUMO
gap of 364 and 380 nm was calculated, respectively,
whereas the corresponding protonated forms exhibit
HOMO-LUMO gaps of 477 or 502 nm. These theoretical
data confirm the experimental trend, i.e., that the pro-
tonated anthryloxazolines exhibit considerably red-
shifted absorption and emission maxima compared to the
unprotonated molecules (see, e.g., the emission data:
FIGURE 6. Spectrofluorimetric titration of 2a (10-5 M in
MeOH) with aqueous HCl; arrows indicate the development
of band maxima during titration; λex ) 363 nm. Before
titration: c(HCl) ) 0 M. Titration end: c(HCl) ) 3.2 × 10-2
M.
however, the formation of a new red-shifted emission
band requires significantly higher acid concentrations.
In methanol solution, the fluorescence intensity of the
anthryl-pyridine 7 was significantly quenched upon
addition of aqueous hydrochloric acid (Figure 10A), but
in contrast to observations with the oxazolines 2a and
2b, only a very weak red-shifted emission maximum was
observed. Nevertheless, the addition of trichloroacetic
acid to 7 in chloroform solution led to a quenching of the
emission maximum along with the formation of a new
band at λ ) 540 nm (Figure 10B).
(14) See, e.g.: Hirsch, T.; Port, H.; Wolf, H. C.; Miehlich, B.;
Effenberger, F. J. Phys. Chem. B 1997, 101, 4525.
Discussion
(15) TURBOMOLE Version 5, Ahlrichs, R.; M. Ba¨r, M.; Baron, H.-
P.; Bauernschmitt, R.; Bo¨cker, S.; Furche, F.; Haase, F.; Ha¨ser, M.;
Horn, H.; Ha¨ttig, C.; Huber, C.; Huniar, U.; Kattanek, M.; Ko¨hn, A.;
Ko¨lmel, C.; Kollwitz, M.; May, K.; Ochsenfeld, C.; O¨ hm, H.; Scha¨fer,
A.; Schneider, U.; Sierka, M.; Treutler, O.; Unterreiner, B.; Arnim, M.
V.; Weigend, F.; Weiss, P.; Weiss, H. Institut f. Physikalische Chemie,
Universita¨t Karlsruhe, Kaiserstr. 12, D-76128 Karlruhe, 1998.
The donor-acceptor interplay in compounds 2a-c and
7 appears to be weak as deduced from their absorption
and emission spectra, i.e., no ICT absorption bands and
only a moderate solvatochromism of the emission proper-
ties have been observed. Nevertheless, the protonation
J. Org. Chem, Vol. 70, No. 10, 2005 3933

Ihmels et al.
FIGURE 7. (A) Spectrofluorimetric titration of 2b (10-5 M in MeOH) with aqueous HCl; arrows indicate the development of
band maxima during titration. Before titration: c(HCl) ) 0 M. Titration end: c(HCl) ) 3.2 × 10-2 M. (B) Fluorescence spectra of
2b (10-6 M in Britton-Robinson buffer at pH 2, 3, 4, 5, 6, 7; cont. 1% DMSO); λex ) 380 nm; arrows indicate the development of
band maxima upon decreasing pH.
FIGURE 8. Pictures of unprotonated (left) and protonated
(right) forms of 2a (A), 2b (B), and 7 (C), λex ) 366 nm. A and
B: c ) 10-5 M in methanol. C: c ) 10-5 M in CHCl3.
2a′: ∆λ ) 103 nm; 2b′: ∆λ ) 122 nm). Furthermore, it
can be seen that 2b′ is red-shifted with respect to its 2a′
FIGURE 9. Spectrofluorimetric titration of 2c (10-5 M in
counterpart by 16 nm from 364 to 380 nm, which is also
MeOH) with aqueous HCl; arrows indicate the development
in agreement with the experimental data.
To discuss the observed photophysical properties from
of band maxima during titration; λex ) 380 nm. Before
titration: c(HCl) ) 0 M. Titration end: c(HCl) ) 3.2 × 10-1
the point of view of a simplified single electron orbital
M.
picture, the Kohn-Sham HOMO and LUMO orbitals for
neutral and protonated 2a′ and 2b′ were used (Figure
moiety. In addition, this effect is more significant for the
protonated molecules. Moreover, the pictorial representa-
tion of orbital coefficients documents nicely the degree
of charge transfer upon photoexcitation of the protonated
anthryloxazolines, which leads to the significant red shift
of the absorption and emission maxima as compared to
the unprotonated molecule. Thus, the electron density
in the HOMO, which is almost equally distributed all
over the anthracene framework, is significantly shifted
toward the oxazolinium moiety in the LUMO. Notably,
this effect also takes place in 2a′, which confirms our
proposal that anthracene may act as a donor toward the
protonated oxazoline substituent even when there is no
additional π-conjugated donor substituent present. These
results are in good agreement with theoretical studies
on 3-styryl-substituted pyridine and pyridinium deriva-
tives.17
11 and Figure 12). These orbitals reveal that in each case
the LUMO has somewhat more weight on the oxazoline
or the oxazolinium moiety, whereas in the HOMO most
of the electron density is accumulated in the anthracene
(16) Gaussian 03, Revision B.04: Frisch, M. J.; Trucks, G. W.;
Schlegel, H. B.; Scuseria, G. E.; Robb, M. A.; Cheeseman, J. R.;
Montgomery, J. A., Jr.; Vreven, T.; Kudin, K. N.; Burant, J. C.; Millam,
J. M.; Iyengar, S. S.; Tomasi, J.; Barone, V.; Mennucci, B.; Cossi, M.;
Scalmani, G.; Rega, N.; Petersson, G. A.; Nakatsuji, H.; Hada, M.;
Ehara, M.; Toyota, K.; Fukuda, R.; Hasegawa, J.; Ishida, M.; Nakajima,
T.; Honda, Y.; Kitao, O.; Nakai, H.; Klene, M.; Li, X.; Knox, J. E.;
Hratchian, H. P.; Cross, J. B.; Adamo, C.; Jaramillo, J.; Gomperts, R.;
Stratmann, R. E.; Yazyev, O.; Austin, A. J.; Cammi, R.; Pomelli, C.;
Ochterski, J. W.; Ayala, P. Y.; Morokuma, K.; Voth, G. A.; Salvador,
P.; Dannenberg, J. J.; Zakrzewski, V. G.; Dapprich, S.; Daniels, A. D.;
Strain, M. C.; Farkas, O.; Malick, D. K.; Rabuck, A. D.; Raghavachari,
K.; Foresman, J. B.; Ortiz, J. V.; Cui, Q.; Baboul A. G.; Clifford, S.;
Cioslowski, J.; Stefanov, B. B.; Liu, G.; Liashenko, A.; Piskorz, P.;
Komaromi, I.; Martin, R. L.; Fox, D. J.; Keith, T. Al-Laham, M. A.;
Peng, C. Y.; Nanayakkara, A.; Challacombe, M.; Gill, P. M. W.;
Johnson, B.; Chen, W.; Wong, M. W.; Gonzalez, C.; Pople, J. A.
Gaussian, Inc., Pittsburgh, PA, 2003.
(17) Scheiner, S.; Kar, T. J. Phys. Chem. B 2002, 106, 534.
3934 J. Org. Chem., Vol. 70, No. 10, 2005

Heterocycles as Acid-Sensitive Fluorescence Probes
FIGURE 10. (A) Spectrofluorimetric titration of 7 (10-5 M in MeOH) with aqueous HCl; arrows indicate the development of
band maxima during titration; λex ) 370 nm. Before titration: c(HCl) ) 0 M. Titration end: c(HCl) ) 5.0 × 10-2 M. (B)
Spectrofluorimetric titration of 7 (10-5 M in CHCl3) with trifluoroacetic acid; arrows indicate the development of band maxima
during titration; λex ) 370 nm. Before titration: c(HCl) ) 0 M. Titration end: c(HCl) ) 1.6 × 10-2 M.
FIGURE 11. Orbital plots (HOMO: C and D, LUMO: A and B) for 2-[2′-(anthryl)]-2-oxazoline 2a′ (A,C) and its N-protonated
form (B, D); derived from calculations (B3LYP 6-311+G**).
One major drawback of the oxazoline substituent is its
limited persistence at low pH due to the acid-catalyzed
hydrolysis.18 Although the initial emission band of the
unprotonated oxazoline 2b may be regained by neutral-
ization, the protonation-neutralization sequence cannot
be repeated more than five times, presumably due to
substantial decomposition of the oxazoline under these
conditions. Thus, we chose the pyridine and benzoxazole
moieties, which are stable at lower acid concentrations.
Moreover, both heterocycles have already been used
as pH-sensitive acceptor substituents in fluorescence
probes.4c-f Indeed, the benzoxazole 2c may be used along
these lines, since its absorption and emission properties
can also be modified by a change of the acid concentra-
tion. Nevertheless, the significantly lower basicity con-
stitutes a notable disadvantage of this system, because
2c may only be used as an appropriate sensor at
relatively high acid concentrations. In the case of the
pyridyl-substituted anthracene 7, the spectrophotometric
acid titration in aqueous solution reveals similar results
as in the case of the oxazolines 2a and 2c, i.e., a drastic
red shift upon protonation of the nitrogen in the hetero-
cyclic moiety of 7 and the accompanying formation of a
strong electron acceptor, but the expected red-shifted
emission band of its protonated form was not detected
in methanol. This observation is in accordance with
results obtained with a pyridyl-substituted coumarin
derivative, whose fluorescence is also quenched upon
protonation without the progression of a corresponding
red-shifted fluorescence band.3f However, other pyridyl-
substituted fluorophores have been shown to exhibit an
intense red-shifted emission in aqueous acids.4a,19 Thus,
the question remains why protonated 7 is nonfluorescent
(19) (a) Wang, S.-L.; Ho, T.-I. J. Photochem. Photobiol. A: Chem.
2000, 135, 119. (b) Haroutounian, S. A.; Katzenellenbogen, J. A.
Photochem. Photobiol. 1988, 47, 503.
(18) Gant, T. G.; Meyers, A. I. Tetrahedron 1994, 50, 2297.
J. Org. Chem, Vol. 70, No. 10, 2005 3935

Ihmels et al.
FIGURE 12. Orbital plots (HOMO: C and D, LUMO: A and B) for 2-[2′-(6′-methoxyanthryl)]-2-oxazoline and 2b′ (A,C) and its
N-protonated form (B,D); derived from calculations (B3LYP 6-311+G**).
in methanol solution. Although it may be suggested that
in protonated 7 an excited-state proton transfer from the
pyridinium to the solvent takes place, it appears to be
unlikely since pyridines usually exhibit larger basicity
in the excited state than in the ground state. Quenching
processes between ground- and excited-state molecules
of the fluorophore can also be excluded since the emission
spectrum does not change upon dilution from 10-5 to 10-6
M. One possibility for the fluorescence quenching in
methanol may be the photoinduced intramolecular elec-
tron transfer from the anthracene moiety to the pyri-
dinium acceptor, which has been proposed also for
pyridinium-substituted 9-vinylanthracene.14 This PET
reaction, which is more likely to take place in the polar
solvent methanol than in nonpolar ones, may lead to an
intermediate which is radiationless deactivated by the
interaction with the solvent (proton transfer or electron
transfer) or by a fast electron back transfer. The latter
would correspond to a twisted intramolecular charge
transfer (TICT) state.20 Since the starting material is
already ionic, this phenomenon is usally called a photo-
induced charge shift (CS), which is often observed to
produce nonemissive excited intermediates in cationic
dyes.14 This explanation of the radiationless deactivation
of protonated 7 in methanol is in agreement with the
observation that in the nonpolar chloroform emission of
7 is observed upon acidification.
In summary, we investigated the ability of three
different N-heterocycles to act as acid-sensitive acceptor
functionalities in fluorescence probes. All of these het-
erocycles may be used for such purposes since the
protonation of the corresponding fluorogenic probes
results in a significant red shift of the emission maxima.
In the case of the oxazoline substituent it was shown that
its protonated form is such a strong acceptor that even
the anthracene without further donor substituents is a
sufficient donor to establish an ICT upon light absorption.
The benzoxazole and pyridine ring offer a particular
advantage due to their persistence under acidic condi-
tions. Thus, they may be used as efficient fluorescence
switches. Nevertheless, the switching of benzoxazole 2c
requires relatively strong acidic conditions, and the
pyridine-pyridinium emission switch does not work in
aqueous or alcoholic solution. Thus, these systems are
not appropriate for applications in analytical biology or
medicine. The oxazoline does not exhibit either of these
disadvantages, but it is not persistent under permanent
acidic conditions. Nevertheless, for a one-time analysis,
this system may be very useful. The oxazolines may be
protonated in a pH range between 4 and 5 which is very
close to physiologically relevant range. Even more im-
portant, simple donor-substitution may enhance its
basicity, so that proton concentrations of about 6 may
be monitored by its emission properties. This is a rather
important pH range since tumor tissue usually exhibits
similar proton concentrations (pH 6.2-6.6),21 as opposed
to healthy tissue which is usually neutral (pH 7.5).
Further studies directed toward these applications are
currently under investigation in our laboratory.
Experimental Section
1H NMR: 200 and 400 MHz.13C NMR: 50.3 and 101 MHz.
Cq, CH, CH2, and CH3 were determined by the DEPT pulse
sequence. 1H NMR chemical shifts refer to δTMS ) 0.0. 13C NMR
chemical shifts refer to solvent signals (DMSO-d6: δ ) 39.5;
CD3CN: δ ) 1.3, 118.1). Melting points are uncorrected.
Absorption and emission spectra were recorded in deoxygen-
ated spectral grade solvents. Unless noted otherwise, the
solution concentrations were 10-4 M for absorption spectros-
copy and 10-5 M for fluorescence spectroscopy. The relative
fluorescence quantum yields were determined by the standard
method22 with quinine sulfate in 1 N H2SO4 as reference (φFl
) 0.54623). The Britton-Robinson buffer solutions were made
(21) (a) Stubbs, M.; McSheehy, P. M.; Griffiths, J. R.; Bashford, C.
L. Mol. Med. Today 2000, 6, 15. (b) Smith, C. A.; Sutherland-Smith,
A. J.; Keppler, B. K.; Kratz, F.; Baker, E. N. J. Biol. Inorg. Chem. 1996,
1, 424.
(22) Demas, J. N.; Crosby, G. A. Crosby, J. Phys. Chem. 1971, 75,
991.
(23) Eaton, D. F. Pure Appl. Chem. 1988, 60, 1108.
(20) Grabowski, Z. R.; Rotkiewicz, K.; Rettig, W. Chem. Rev. 2003,
103, 3899.
3936 J. Org. Chem., Vol. 70, No. 10, 2005

Heterocycles as Acid-Sensitive Fluorescence Probes
according to literature procedure.24 Because of its low solubility
in aqueous solution, the anthracene 2b was dissolved in DMSO
(c(2b) ) 10-4 M) and subsequently added to the buffer
solutions, so that the buffer also contained 1 vol % of DMSO.
The pKa data for the ground and excited state (see Table 2)
were determined from spectrophotometric titrations according
to literature procedures.25
chloride (19.5 mL) was stirred under reflux for 18 h. After
evaporation of excess oxalyl chloride in a vacuum, the solid
was added to a solution of 2-aminophenol (88.0 mg, 0.80 mmol)
in pyridine (2 mL). The solution was stirred for 1 h at 100 °C.
The pyridine was evaporated, and the remaining dark residue
was heated for 0.5 h at 200 °C under argon-gas atmosphere.
Purification by column chromatography (SiO2, n-hexane/ethyl
acetate 6:1, Rf ) 0.31) and crystallization from n-hexane/
dichloromethane gave 2c (84.0 mg, 0.26 mmol, 34%) as a
yellow solid. Mp: 148-149 °C. 1H NMR (CDCl3, 200 MHz): δ
3.98 (s, 3 H, OCH3), 7.17-7.22 (m, 2 H, Ar-H), 7.34-7.42 (m,
2 H, Ar-H), 7.59-7.66 (m, 1 H, Ar-H), 7.78-7.87 (m, 1 H, Ar-
H), 7.94 (d, J ) 9.8 Hz, 1 H, Ar-H), 8.05 (d, J ) 9.0 Hz, 1 H,
Ar-H), 8.23-8.29 (m, 2 H, Ar-H), 8.48 (s, 1 H, Ar-H), 8.91 (s,
1 H, Ar-H). 13C NMR (CDCl3, 50 MHz): δ 55.4 (CH3), 103.7
(CH), 110.6 (CH), 119.9 (CH), 121.2 (CH), 122.8 (Cq), 123.2
(CH), 124.3 (CH), 124.6 (CH), 125.1 (CH), 128.0 (CH), 128.5
(CH), 128.8 (Cq), 129.1 (CH), 129.4 (Cq), 130.1 (CH), 132.7 (Cq),
134.1 (Cq), 142.2 (Cq), 150.8 (Cq), 158.0 (Cq), 163.4 (Cq). MS
(70 eV, EI) m/z: 325 (100) [M+], 282 (87), 190 (21), 164 (24).
Anal. Calcd for C22H15N1O2‚0.5H2O: C, 79.03; H, 4.82; N, 4.19.
Found: C, 79.10; H, 4.85; N, 4.11.
2-[2′-(4′′-(2′′′-Pyridyl)phenyl)hydroxymethyl-4′-meth-
oxyphenyl]-4,4-dimethyl-2-oxazoline (4). To a cooled solu-
tion (0 °C) of 37 (1.02 g, 5.00 mmol) in dry diethyl ether (50
mL) was added n-butyllithium (7 mL, 1.6 M in hexane, 5.46
mmol). The solution was stirred for 4 h at 0 °C, and the color
changed from yellow to orange and eventually to brown. A
solution of (2-pyridyl)benzaldehyde (1.00 g, 5.46 mmol) in
diethyl ether (20 mL) was added at 0 °C, and a yellow solid
precipitated. The ice bath was removed, and the suspension
was stirred for 16 h at rt and subsequently poured into a
saturated aqueous solution of NH4Cl (50 mL). After 15 min of
thorough stirring, the aqueous layer was separated and
extracted with diethyl ether (20 mL). The combined organic
phases were washed with water (30 mL) and dried with
MgSO4. After removal of the solvent in a vacuum, 4 was
obtained as a yellow solid (2.08 g, >97%) which was spectro-
scopically pure by 1H NMR. Mp: 102-103. °C. 1H NMR (CDCl3,
400 MHz): δ 1.04 (s, 3 H, CH3), 1.34 (s, 3 H, CH3), 3.78 (s, 3
H, OCH3), 3.92 (d, J ) 8 Hz, 1 H, CH2), 3.99 (d, J ) 8 Hz, 1 H,
CH2), 5.94 (s, 1 H, CH), 6.75 (d, J ) 3 Hz, 1 H, Ar-H), 6.86
(dd, J ) 9 Hz, J ) 3 Hz, 1 H, Ar-H), 7.17-7.24 (m, 1 H, Ar-H),
7.44 (d, J ) 8 Hz, 1 H, Ar-H), 7.72-7.74 (m, 2 H, Ar-H), 7.84
(d, J ) 9 Hz, 1 H, Ar-H), 7.94 (d, J ) 9 Hz, 1 H, Ar-H), 8.66-
8.70 (m, 1 H, Ar-H). 13C NMR (CDCl3, 101 MHz): δ 27.9 (CH3),
28.4 (CH3), 55.3 (CH3), 67.7 (Cq), 74.9 (CH), 78.6 (CH2), 112.0
(CH), 116.7 (CH), 118.9 (Cq), 120.4 (CH), 121.9 (CH), 126.3 (2
× CH), 127.1 (2 × CH), 132.6 (CH), 136.7 (CH), 137.8 (Cq),
144.2 (Cq), 146.7 (Cq), 149.6 (CH), 157.4 (Cq), 161.8 (Cq), 162.2
(Cq). MS (EI, 70 eV) m/z: 388 (39) [M+], 317 (100). Anal. Calcd
for C24H24N2O3‚0.5H2O (397.5): C, 72.52; H, 6.34; N, 7.05.
Found: C, 72.59; H, 6.40; N, 6.88.
2-[2′-(Anthryl)]-4,4-dimethyl-2-oxazoline (2a). A solution
of 383 mg (1.72 mmol) of 2-anthracenecarboxylic acid in 10
mL of oxalyl chloride was heated at reflux for 18 h under
argon-gas atmosphere. After evaporation of excess oxalyl
chloride in a vacuum, the solid was dissolved in dichlo-
romethane and added dropwise to a solution of 307 mg (3.44
mmol) of 2-amino-2-methylpropanol in 8 mL of dichlo-
romethane at 0 °C. After being for 2 h at 20 °C, the precipitated
solid was filtered off. To the remaining solution was added
0.5 mL of thionyl chloride, the solution was kept at reflux for
2 h under argon-gas atmosphere, and after that the solvent
was evaporated in a vacuum. The remaining brown solid was
washed with diethyl ether. To the solid were added water,
diethyl ether, and diluted aqueous NaOH until a pH of 8-9
was indicated. The aqueous phase was extracted twice with
diethyl ether, the combined organic phases were dried with
MgSO4, and the solvent was removed in a vacuum. Crystal-
lization from n-hexane/dichloromethane gave 170 mg (0.62
mmol, 36%) of oxazoline 2a as a yellow solid. For further
purification column chromatography (SiO2, n-hexane/ethyl
acetate 1:1 with 1% pyridine, Rf ) 0.70) or sublimation (100
°C, 0.4 mbar) was used. Mp: 107-110 °C dec. 1H NMR (CDCl3,
200 MHz): δ 1.45 (s, 6 H, 2 CH3), 4.20 (s, 2 H, CH2), 7.47-
7.52 (m, 2 H, Ar-H), 8.00-8.04 (m, 4 H, Ar-H), 8.43 (s, 1 H,
Ar-H), 8.52 (s, 1 H, Ar-H), 8.69 (s, 1 H, Ar-H). 13C NMR (CDCl3,
100 MHz): δ 28.5 (2 CH3), 67.7 (Cq), 79.2 (CH2), 124.1 (CH),
125.7 (CH), 126.2 (CH), 127.2 (CH), 127.4 (CH), 127.9 (CH),
128.2 (CH), 128.4 (CH), 129.4 (CH), 130.7 (Cq), 132.0 (Cq), 132.1
(Cq), 132.6 (Cq), 162.3 (Cq). MS (EI, 70 eV) m/z: 275 (78) [M+],
260 (100). HRMS: calcd for C19H17NO (275.1310), found
275.1311.
2-[2-(6-Methoxyanthryl)]-4,4-dimethyl-2-oxazoline (2b).
A solution of 38.0 mg (0.15 mmol) of 6-methoxy-2-anthracene-
carboxylic acid in 3.8 mL of oxalyl chloride was heated at reflux
for 18 h under argon-gas atmosphere. After evaporation of
excess oxalyl chloride in a vacuum, the solid was dissolved in
dichloromethane and added dropwise to a solution of 27.0 mg
(0.30 mmol) of 2-amino-2-methylpropanol in 3.8 mL of dichlo-
romethane at 0 °C. After the mixture was stirred for 2 h at 20
°C, the precipitated solid was filtered off. To the remaining
solution was added 0.1 mL of thionyl chloride, the solution
was kept at reflux for 2 h under argon-gas atmosphere, and
after that the solvent was evaporated in a vacuum. The
remaining brown solid was washed with diethyl ether. To the
solid were added water, diethyl ether, and diluted aqueous
NaOH until a pH of 8-9 was indicated. The aqueous phase
was extracted twice with diethyl ether, the combined organic
phases were dried with MgSO4, and the solvent was removed
in a vacuum. Crystallization from n-hexane/dichloromethane
gave 25.0 mg (0.09 mmol, 60%) of oxazoline 1f as yellow solid.
Mp: 220-225 °C. 1H NMR (CDCl3, 200 MHz): δ 1.44 (s, 6 H),
3.96 (s, 3 H), 4.18 (s, 2 H), 7.14-7.19 (m, 2 H), 7.87-8.01 (m,
3 H), 8.24 (s, 1 H), 8.38 (s, 1 H), 8.55 (s, 1 H). 13C NMR (CDCl3,
50 MHz): δ 28.5 (2 × CH3), 55.3 CH3), 67.7 (Cq), 79.2 (CH2),
103.6 (CH), 120.9 (CH), 123.8 (Cq), 124.1 (CH), 124.3 (CH),
127.7 (CH), 127.8 (CH), 128.5 (Cq), 129.3 (Cq), 129.5 (CH), 130.0
(CH), 132.6 (Cq), 133.8 (Cq), 157.8 (Cq), 162.3 (Cq). MS (70 eV,
EI) m/z: 305 (82) [M+], 290 (100), 233 (71), 219 (5), 207 (9),
176 (4), 164 (21). HRMS: calcd for C20H19N1O2 (305.1416),
found 305.1416.
3-[4′-(2′′-Pyridyl)phenyl]-5-methoxyphthalide (5). A so-
lution of 4 (2.08 g, 5.35 mmol) and p-toluenesulfonic acid (4.78
g, 25.2 mmol) in toluene (150 mL) was stirred under reflux
for 2.5 h. The brown aqueous layer was separated and
extracted three times with toluene (50 mL). The combined
organic phases were washed with brine and dried MgSO4.
After removal of the solvent in a vacuum, 5 was obtained as a
yellow solid (1.05 g, 3.30 mmol, 62%). Mp: 151-152 °C. IR
(KBr): ν˜ 1757 cm-1 (CdO). 1H NMR (CDCl3, 400 MHz): δ 3.82
(s, 3 H, OCH3), 6.36 (s, 1 H, CH), 6.74 (d, J ) 2 Hz, 1 H, Ar-
H), 7.06 (dd, J ) 9 Hz, J ) 2 Hz, 1 H, Ar-H), 7.24-7.27 (m, 1
H, Ar-H), 7.39, 8.01 (AA′BB′ system, 4 H, Ar-H), 7.71-7.79
(m, 2 H, Ar-H), 7.87 (d, J ) 9 Hz, 1 H, Ar-H), 8.69 (dd, J ) 5
Hz, J ) 1 Hz, 1 H, Ar-H). 13C NMR (CDCl3, 101 MHz): δ 55.9
(CH3), 81.8 (CH), 106.5 (CH), 117.1 (CH), 117.8 (Cq), 120.7
(CH), 122.5 (CH), 127.2 (CH), 127.4 (2 CH), 127.6 (2 CH), 137.0
(CH), 137.3 (Cq), 140.2 (Cq), 149.7 (CH), 152.5 (Cq), 156.5 (Cq),
165.0 (Cq), 170.2 (Cq). MS (EI, 70 eV) m/z: 317 (100) [M+]. Anal.
2-[2-(6-Methoxyanthryl)]benzoxazole (2c). Under argon-
gas atmosphere, a solution of 1b (195 mg, 0.77 mmol) in oxalyl
(24) Britton, H. T. S.; Robinson, R. A. J. Chem. Soc. 1931, 458.
(25) Polster, J.; Lachmann H. Spectrometric Titrations - Analysis
of Chemical Equilibria; VCH: Weinheim, 1989.
J. Org. Chem, Vol. 70, No. 10, 2005 3937

Ihmels et al.
Calcd for C20H15NO3 (317.3): C, 75.70; H, 4.76; N, 4.41.
Found: C, 75.56; H, 4.98; N, 4.41.
(30 mL) and methanol (5 mL). Argon gas was bubbled through
the solution for 30 min to remove oxygen gas. Solid NaBH4
(677 mg, 17.9 mmol) was added, and the solution was stirred
under reflux for 4 d. The reaction mixture was poured on ice-
water, and excess NaBH4 was decomposed with concd HCl.
The suspension was extracted three times with dichlo-
romethane (50 mL). The combined organic phases were dried
with MgSO4, the solvent was removed in a vacuum, and the
remaining solid was purified by column chromatography (SiO2,
PE/acetone 3:1, 1% pyridine, Rf ) 0.52) to give 7 as a yellow
powder (157 mg, 550 µmol, 21% over both steps). Mp: 231-
232 °C. 1H NMR (CDCl3, 400 MHz): δ 3.98 (s, 3 H, OCH3),
7.17 (dd, J ) 9 Hz, J ) 2 Hz, 1 H, Ar-H), 7.21 (d, J ) 3 Hz, 1
H, Ar-H), 7.27 (dd, J ) 8 Hz, J ) 5 Hz, 1 H, Ar-H), 7.28 (dd,
J ) 8 Hz, J ) 2 Hz, 1 H, Ar-H), 7.82 (ddd, J ) 8 Hz, J ) 8 Hz,
J ) 2 Hz, 1 H, Ar-H), 7.90-7.93 (m, 2 H, Ar-H), 8.05 (dd, J )
9 Hz, J ) 1 Hz, 1 H, Ar-H), 8.13 (dd, J ) 9 Hz, J ) 2 Hz, 1 H,
Ar-H), 8.29 (s, 1 H, Ar-H), 8.45 (s, 1 H, Ar-H), 8.62 (s, 1 H,
4-Methoxy-2-[(4′-(2′′-pyridyl)phenyl)methyl]benzoic
Acid (6). Zinc dust (16.0 g, 0.25 mol) was activated by
successive treatment with diluted hydrochloric acid, water, and
methanol and added to a solution of 5 (1.05 g, 3.31 mmol) in
acetic acid (150 mL). The suspension was stirred under reflux
for 24 h. The zinc dust was filtered off, and the filtrate was
poured into a water/ice mixture. The solution was neutralized
with aqueous KOH (10%) and extracted four times with
dichloromethane (50 mL). The combined organic phases were
washed with brine (20 mL) and dried with MgSO4. After
removal of the solvent in a vacuum, 6 was obtained as a pale
yellow solid (0.79 g, 2.48 mmol, 75%). Mp: 176-177 °C. IR
(KBr): ν˜ 1700 cm-1 (CdO). 1H NMR (CD2Cl2, 400 MHz): δ
3.81 (s, 3 H, OCH3), 4.48 (s, 2 H, CH2), 6.78 (d, J ) 3 Hz, 1 H,
Ar-H), 6.83 (dd, J ) 9 Hz, J ) 3 Hz, 1 H, Ar-H), 7.23 (ddd, J
) 7 Hz, J ) 5 Hz, J ) 1 Hz, 1 H, Ar-H), 7.27, 7.88 (AA′BB′
system, 4 H, Ar-H), 7.70 (ddd, J ) 8 Hz, J ) 1 Hz, J ) 1 Hz,
1 H, Ar-H), 7.75 (ddd, J ) 8 Hz, J ) 7 Hz, J ) 2 Hz, 1 H,
Ar-H), 8.06 (d, J ) 9 Hz, 1 H, Ar-H), 8.68 (ddd J ) 5 Hz, J )
2 Hz, J ) 1 Hz, 1 H, Ar-H). 13C NMR (CD2Cl2, 101 MHz): δ
40.0 (CH2), 55.8 (CH3), 111.8 (CH), 117.8 (CH), 120.9 (CH),
121.3 (Cq), 122.4 (CH), 127.3 (2 CH), 129.7 (2 CH), 134.5 (CH),
137.3 (Cq), 137.4 (CH), 142.5 (Cq), 146.0 (Cq), 149.8 (CH), 157.6
(Cq), 163.5 (Cq), 170.2 (Cq). MS (EI, 70 eV) m/z: 319 (47) [M+],
274 (100). Anal. Calcd for C20H17NO3‚0.25H2O (289.9): C,
74.17; H, 5.45; N, 4.32. Found: C, 74.13; H, 5.59; N, 4.28.
2-(6-Methoxyanthracen-2-yl)pyridine (7). Under argon-
gas atmosphere, 6 (835 mg, 2.62 mmol) was added to meth-
anesulfonic acid (6 mL) at 90 °C. The solution was stirred at
90 °C for 4 h, and its color changed to a dark green color. The
reaction mixture was cooled to room temperature and poured
into ice-water (200 mL). The solution was stirred for 30 min
and treated with aqueous KOH (10%) until a pH ) 10 was
indicated. The solution was extracted four times with dichlo-
romethane (100 mL). The combined organic phases were
washed with saturated aqueous NaHCO3 (100 mL) and water
(100 mL) and subsequently dried with MgSO4. The solvent was
removed in a vacuum, and the resulting yellow solid (360 mg)
was filtered through silica gel and redissolved in 2-propanol
Ar-H), 8.76 (ddd J ) 5 Hz, J ) 2 Hz, J ) 1 Hz, 1 H, Ar-H). 13
C
NMR (CDCl3, 101 MHz): δ 55.3 (CH3), 103.7 (CH), 120.7 (CH),
122.0 (CH), 124.0 (CH), 124.4 (CH), 126.8 (CH), 127.4 (CH),
128.2 (CH), 128.7 (Cq), 130.0 (CH), 132.2 (Cq), 133.3 (Cq), 135.1
(Cq), 136.9 (CH), 149.7 (CH), 157.4 (Cq), 157.5 (Cq). MS (EI,
70 eV) m/z: 285 (100) [M+]. Anal. Calcd for for C20H15NO‚
0.25H2O (289.9): C, 82.88; H, 5.39; N, 4.83. Found: C, 82.89;
H, 5.4; N, 4.76.
Acknowledgment. This paper is dedicated to Prof.
Hans-Jo¨rg Deiseroth, University of Siegen, on the
occasion of his 60th birthday. This work was generously
financed by the Bundesministerium fu¨r Bildung und
Forschung, the Deutsche Forschungsgemeinschaft, the
Fonds der Chemischen Industrie, and the Welch Foun-
dation of Houston, TX.
Supporting Information Available: Magnified repre-
sentations of Figures 2-7, 9, and 10. This material is available
free of charge via the Internet at http://pubs.acs.org.
JO047841Z
3938 J. Org. Chem., Vol. 70, No. 10, 2005