Cyclopenta[b]naphthalene cyanoacrylate dyes: Synthesis and evaluation as fluorescent molecular rotors

Article abstract of DOI:10.1039/c4ob02563f

We describe the design, synthesis and fluorescent profile of a family of environment-sensitive dyes in which a dimethylamino (donor) group is conjugated to a cyanoacrylate (acceptor) unit via a cyclopenta[b]naphthalene ring system. This assembly satisfies

Full text of DOI:10.1039/c4ob02563f

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Cyclopenta[b]naphthalene cyanoacrylate dyes:
synthesis and evaluation as fluorescent molecular
rotors†
Cite this: DOI: 10.1039/c4ob02563f
Laura S. Kocsis,a Kristyna M. Elbel,b Billie A. Hardigree,c Kay M. Brummond,*a
Mark A. Haidekker*c and Emmanuel A. Theodorakis*b
We describe the design, synthesis and fluorescent profile of a family of environment-sensitive dyes in
which a dimethylamino (donor) group is conjugated to a cyanoacrylate (acceptor) unit via a cyclopenta[b]-
naphthalene ring system. This assembly satisfies the typical D–π-A motif of a fluorescent molecular rotor
and exhibits solvatochromic and viscosity-sensitive fluorescence emission. The central naphthalene ring
system of these dyes was synthesized via a novel intramolecular dehydrogenative dehydro-Diels–Alder
(IDDDA) reaction that permits incorporation of the donor and acceptor groups in variable positions
around the aromatic core. A bathochromic shift of excitation and emission peaks was observed with
increasing solvent polarity but the dyes exhibited a complex emission pattern with a second red emission
band when dissolved in nonpolar solvents. Consistent with other known molecular rotors, the emission
intensity increased with increasing viscosity. Interestingly, closer spatial proximity between the donor and
the acceptor groups led to decreased viscosity sensitivity combined with an increased quantum yield.
This observation indicates that structural hindrance of intramolecular rotation dominates when the donor
and acceptor groups are in close proximity. The examined compounds give insight into how excited state
intramolecular rotation can be influenced by both the solvent and the chemical structure.
Received 8th December 2014,
Accepted 13th January 2015
DOI: 10.1039/c4ob02563f
www.rsc.org/obc
specific analytes) in real time and with high spatial-temporal
resolution.5,6
Introduction
The study of events in chemistry, biology, and materials
Typically, the FMR motif consists of an electron donor
science requires the development of sophisticated sensory group that is in conjugation with an electron acceptor group
devices that can accurately measure changes that occur in (D–π-A motif).7 In this configuration, the probe responds to
their environment.1 For instance, questions related to cell photoexcitation with an intramolecular charge transfer from
function, tissue organization and organ activities in normal the donor to acceptor. Environmental factors (e.g. solvent
and diseased states cannot be answered without the use of polarity) affect the energy levels of either the ground or excited
specifically designed sensors.2 A subclass of these sensors can states and shift the fluorescence emission wavelength accord-
provide dynamic information of their surrounding stimuli by ingly. Moreover, relaxation from the excited state can occur via
accordingly modifying their fluorescence emission due to sol- fluorescence emission and/or via mechanical de-excitation
vatochromism.3 Often referred to as fluorescent molecular (e.g. rotation across the σ-bonds). Thus, the fluorescence
rotors (FMRs), these environment-sensitive dyes4 can be used quantum yield of FMRs depends on the rigidity, i.e. viscosity,
to measure fluctuations in properties of their microenviron- of their surrounding environment.8 Specifically, if the intramo-
ment (e.g. pH, viscosity, polarity, voltage and presence of lecular rotation around the σ-bonds between donor and
acceptor becomes hindered, e.g. within a viscous or rigid
environment, the quantum yield of fluorescence intensity
increases. Thus, photoexcitation of an FMR in the solid state
aDepartment of Chemistry, University of Pittsburgh, 219 Parkman Avenue,
Pittsburgh, PA 15260, USA. E-mail: kbrummond@pitt.edu
bDepartment of Chemistry & Biochemistry, University of California, San Diego, 9500
Gilman Drive MC: 0358, La Jolla, CA 92093-0358, USA. E-mail: etheodor@ucsd.edu;
Fax: +1-858-822-0386; Tel: +1-858-822-0456
leads mainly to fluorescence emission with a quantum yield
close to unity, since the σ-bonds of the molecule cannot rotate.
On the other hand, photoexcitation in a fluid environment
(e.g. low viscosity or high free volume) leads to both fluo-
rescence and non-fluorescence relaxation processes, the ratio
of which is related to the fluidity of the medium.9
cCollege of Engineering, University of Georgia, 597 D. W. Brooks Drive, Athens,
GA 30602, USA. E-mail: mhaidekk@uga.edu
†Electronic supplementary information (ESI) available: 1H and 13C NMR spectra
for all compounds. See DOI: 10.1039/c4ob02563f
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CNCA dyes were designed to test the effect of the relative posi-
tion of the dimethylamino and the cyanoacrylate groups on
solvatochromism and viscosity sensitivity.
Synthesis of CNCA probes
The CNCA dyes 3–5 were synthesized via a four-step protocol
beginning from enyne substrates 6a–c (Scheme 1). A pyridi-
nium chlorochromate oxidation of commercially available
5-hexyn-1-ol, followed by a Horner–Wadsworth–Emmons reac-
tion of the resulting aldehyde with n-butyllithium and chloro-
benzylphosphonate produced the enyne precursors 6a–c, as
previously reported.17a Formylation of the lithium acetylide of
6a–c by addition of N,N-dimethylformamide formed aldehydes
7a–c in 68–80% yield. Naphthalene derivatives 8a–c were then
generated by a microwave-assisted IDDDA reaction of 7a–c at
180 °C for 1 h (73–80% yield).17a The IDDDA reaction of 7b
resulted in formation of 8-chloro-naphthalene derivative 8d in
addition to 8b in a 79% combined yield (1 : 1.5 8b : 8d). Con-
version of chloro-substituted naphthalenes 8a–c to the amine
derivatives 9a–c was conducted via a palladium-catalyzed
Buchwald–Hartwig cross-coupling reaction employing RuPhos
palladacycle, cesium carbonate, and dimethylamine (44–74%
yield).18 Attempts to perform a Buchwald–Hartwig cross-coup-
ling reaction21 on the sterically hindered naphthalene 8d were
unsuccessful. Subsequent Knoevenagel condensation reactions
of 9a–c utilizing cyanoester 1020 and catalytic 1,8-diazabi-
cycloundec-7-ene (DBU) provided CNCA dyes 3–5 in 77–90%
yield. This condensation is known to produce the thermodyna-
mically stable trans alkenes, in which the ester group and the
aromatic ring reside at the opposite face of the newly formed
double bond. Nonetheless, upon purification of 3–5 we
observed partial double bond isomerization (trans/cis ratio:
Fig. 1 Chemical structures of Prodan (1), Laurdan (2), and cyclopenta[b]-
naphthalene cyanoacrylate (CNCA) dyes (3–5).
The fluorescent dye Prodan (6-propionyl-2-dimethylamino-
naphthalene, 1) is one of the best-known FMRs as it exhibits a
strong bathochromic shift of its emission wavelength in polar
solvents.10 Both Prodan and its more lipophilic analogue
Laurdan (6-dodecanoyl-2-dimethylaminonaphthalene, 2) are
frequently used to measure changes in viscosity and polarity
in phospholipid bilayers and cell membranes.11 Various struc-
tural modifications of Prodan that maintain its fluorophoric
motif but have altered substitutions at the alkyl side chain, e.g.
acrylodan,12 badan13 and aladan,14 have been evaluated as
FMRs. In addition, replacement of the naphthalene motif by
an anthracene15 and a fluorene16 ring system has produced
red-shifted dyes suitable for various biological applications.
Inspired by this data, we sought to apply a novel intramolecu-
lar dehydrogenative dehydro-Diels–Alder (IDDDA) reaction17
for the synthesis of a new family of functionalized naphtha-
lenes, referred to here as CNCA dyes, and evaluate them as
FMRs. Importantly, this key reaction allows the incorporation
of the donor–acceptor pair at different positions of the
naphthalene ring thereby permitting a methodical evaluation
of structure–photophysical property relationships.17a,18 Herein,
we present the synthesis, fluorescent profile and viscosity/
polarity sensitivity of these probes (Fig. 1).
Results and discussion
Design criteria of CNCA probes
Our recently developed IDDDA reaction of styrenyl derivatives
provides efficient and de novo access to a variably functiona-
lized cyclopenta[b]naphthalene scaffold. We envisioned that
further substitution of this scaffold with an amine (donor
group) and a cyanoacrylate (acceptor group) would create the
desired D–π-A fluorophore, referred to here as the cyclopenta-
[b]naphthalene cyanoacrylate (CNCA) motif. In accordance Scheme 1 Reagents and conditions: (a) 6a–c (1.0 equiv.), n-butyl-
with previous studies,19 we anticipated that this motif would
exhibit a viscosity-sensitive emission behavior. To increase
lithium (1.0 equiv.), DMF (2.0 equiv.), THF, −78 °C to rt; (b) 7a–c,
o-dichlorobenzene, μW, 180 °C, 1 h; (c) RuPhos palladacycle (3 mol%),
cesium carbonate (2.0 equiv.), 8a–c (1.0 equiv.), dimethylamine (2.0 M in
THF, 3.0 equiv.), THF, 85 °C, 2.5–5 h; (d) 9a–c (1.0 equiv.), 10 (0.8
solubility of the lipophilic motif in polar solvents, we decided
to attach at the acceptor unit a triethylene glycol monomethyl
equiv.), DBU (0.010 equiv.), THF, 50 °C, 3 h. *Formed as a mixture with
ether (TEGME) group at the acceptor unit.20 The synthesized 48% of 8-Cl substituted product 8d.
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2.2/1) that likely occurred due to the acidic nature of the silica Viscosity-dependent behavior
gel. In fact, treating the resulting mixture of isomers with cata-
The emission spectra of 3, 4, and 5 in different mixtures of
methanol, ethylene glycol, and glycerol were obtained by exci-
tation at the optimum wavelength and are shown in Fig. 2.
Compound 5 appeared to emit fluorescence only in one broad
band that showed almost no variability with the solvent. On
the other hand, compounds 3 and 4 exhibited two distinct
emission peaks. The emission intensity of 3 and 4 increased
with solvents of increased viscosity. Only the longer-wave-
length emission peak of 3 exhibited a noticeable hypsochromic
shift in solvents with higher glycerol content.
lytic DBU for 1 h at 50 °C reformed exclusively the trans-
isomer. To avoid this isomerization, we pretreated the silica
gel with 10% triethylamine/hexanes prior to purification of the
crude substrates and were then able to isolate exclusively the
desired trans isomer. It is also worth mentioning that a small
amount of 9 (between 5–10%) was also recovered during purifi-
cation of compounds 3–5. This retro-Knoevenagel reaction is
likely a result of hydrolysis due to water retained by the
silica gel.
When a power-law relationship between viscosity and
quantum yield (and therefore also intensity) exists (eqn (1)),
data points of intensity, drawn over viscosity in a double-
logarithmic scale, should lie on a straight line, and the slope
of the line is the exponent x in eqn (1).
Solvatochromic behavior of CNCA probes
The solvatochromic behavior of compounds 3–5 was studied
in solvents of various polarities (Table 1). A condensed over-
view of the excitation and emission maxima of 3–5 with their
associated approximate peak intensity values (given in 106
photon counts per second, cps) is provided in Table 1.
ꢀ ꢁ
x
η
ϕF ¼ ϕ0 ꢀ
ð1Þ
σ
In general, compound 3, containing a 1,5-substituted cyclo-
penta[b]naphthalene moiety, absorbed light at shorter wave-
The log–log plots of peak intensity over viscosity are shown
lengths (320–395 nm) in most solvents used as compared to 4 in Fig. 3. Straight lines were fitted by linear regression. In the
(292–382 nm) and 5 (280–436 nm). Compound 5, containing a case of 5, the line did not have a slope significantly different
1,7-substituted cyclopenta[b]naphthalene moiety, has the from zero. Conversely, the slopes of the intensity peaks of 4
strongest fluorescence intensity in all solvents. The CNCA dyes were 0.12 and 0.14, respectively. For 3, we found slopes of 0.1
exhibited multiple emission peaks, and the solvent polarity and 0.37, all of them significantly different from zero
influenced the balance between the two peaks. In 3 and 4, the (P < 0.005). Relative intensities (i.e., the y-intercept of the
emission from the red band (emission peak near λ = 610 nm) slope, which is the extrapolated value at a hypothetical vis-
is best visible in toluene and less prominent than that of 5 (λ = cosity of 1 mPa s) were 54 700 cps (red-shifted emission band)
522 nm). A bathochromic shift of both peak excitation and and 234 000 cps (blue-shifted emission band) for compound 3,
peak emission with increasing solvent polarity can most 247 000 (blue-shifted emission band) and 209 000 cps
clearly be seen in compound 3. In addition, 5 (and to a lesser (red-shifted emission band) for compound 4, and 750 000 cps
extend 3 and 4) can be excited at 280 nm. Most likely, this is a for the single emission band of compound 5.
direct excitation of the naphthalene core since naphthalene
itself is fluorescent.23
Fluorescent molecular rotors are generally considered to
have two emission bands: one from the planar state, and one
Table 1 Dominant fluorescence peaks of CNCA dyes 3, 4 and 5 in solvents of different polarity (polarity is given relative to water)22
Compound 3
Compound 4
Compound 5
Solvent
Excitation Emission Peak
Excitation Emission Peak
Excitation Emission Peak
(Polarity)22
λ (nm)
λ (nm)
Intensity (×106) λ (nm)
λ (nm)
Intensity (×106) λ (nm)
λ (nm)
Intensity (×106)
Toluene
(0.1)
395
333
616
414
0.8
0.25
363
306
603
325
0.42
3.0
425
363
302
522
413
413
2.7
2.5
1.4
DMSO
(0.44)
335
280
425
360
0.6
0.95
365
340
430
369
1.7
1.4
436
364
280
562
430
430
2.5
3.6
2.5
Acetonitrile
(0.46)
348
322
434
439
446
0.2
382
314
425
425
0.8
0.3
425
368
280
548
427
427
2.5
2.2
1.4
Methanol
(0.76)
0.65
0.2
360
315
427
433
0.85
0.95
416
365
280
500
422
422
0.6
2.3
1.4
Water/methanol 320
(0.88)
343
292
433
433
0.4
0.3
384
356
295
475
432
432
1.1
1.05
0.7
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Fig. 2 Emission spectra of 3, 4, and 5 in alcohol mixtures of different viscosity. While 5 shows no major change in the different alcohol mixtures, 4
and notably 3 exhibit increased peak emission from the red-shifted band in alcohols with higher viscosity.
Fig. 3 Peak emission intensity plotted over solvent viscosity in a double-logarithmic scale. Compounds 3 and 4 have significantly positive slopes,
and the slope can be used as an indication of the sensitivity towards viscosity. The slope of 5 is not significantly different from zero. Circles indicate
the blue-shifted band (and the single emission of 5) and open squares indicate the red-shifted band.
from the twisted intramolecular charge transfer (TICT) state. 5, containing
a 1,7-substituted cyclopenta[b]naphthalene
A good example is DMABN (dimethylaminobenzonitrile), whose moiety, was found to have the strongest fluorescence emission
photophysical properties are well-explored.24 In the ground and was the least viscosity sensitive among all compounds
state, the energy of the twisted conformation is higher than tested. On the other hand, compound 3, containing a 1,5-sub-
that of the planar state. In the first excited state, we find the stituted cyclopenta[b]naphthalene moiety, was shown to be the
exact opposite. Therefore, excitation takes place preferentially least fluorescent but most viscosity sensitive, while compound
from the planar state, and emission from the TICT state, 4, containing
a 1,6-substituted cyclopenta[b]naphthalene
unless rotation is hindered in some way. A reduction of the moiety, had intermediate values of fluorescence intensity and
intramolecular rotation rate, for example due to viscous micro- viscosity sensitivity. The inverse relationship between fluo-
friction, leads to dual-band emission with increasing intensity rescence emission and viscosity sensitivity is a general charac-
emitted from the planar state (blue-shifted band). It is also teristic of FMRs and has been previously observed for related
possible that the energy gap between excited and ground naphthalene-based fluorophores.26 It is likely that placement
states in the twisted conformation is so low that no photon of the donor (D) and acceptor (A) groups in close proximity
emission occurs. Those molecular rotors, of which a represen- (e.g. in compound 5) decreases their rotational ability and thus
tative example is DCVJ (9-dicyano-vinyl julolidine), do not have increases the energy level of the TICT state (Fig. 4). This effect
any red-shifted emission. Their quantum yield depends on translates into fluorescence deexcitation without significant
emission from the planar state, which increases with increased passage through the environment-sensitive TICT state. Thus,
microfriction. In the case of 3 and 4, we observe a dual emis- compound 5 becomes more fluorescent but less sensitive to
sion. Increase of solvent viscosity leads to a sharp increase of solvent viscosity.
the red-shifted band (most pronounced in compound 3)
whereas the blue-shifted band increases less strongly. One
possible explanation is a reversal of the energetically-preferred
conformations, as found, for example, in BODIPY-related
dyes.25 Since both emission peaks of 3 and 4 increase with
increasing viscosity, it is also possible that a third energy level
exists which is comparatively weak and shows an extreme
Stokes shift. This is consistent with the observation of emis-
sion in the 600 nm range in toluene. If this interpretation is
correct, the dual emission occurs from a planar or almost-
planar state, and twisted-state emission is invisible due to its
low energy gap.
It is interesting to correlate the structural and photo-
physical differences between compounds 3, 4 and 5. CNCA dye
Fig. 4 Comparison of the relative position of donor (D, Me2N) and
acceptor (A, CHvC(CN)CO2TEGME) substituents in CNCA dyes. Placing
the D and A groups in close proximity decreases their rotational ability
thereby allowing emission from the planar excited state.
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solution was cooled at −78 °C (bath temperature) in a dry ice/
acetone bath, and n-butyllithium (1.0 equiv. of a 1.6 M solu-
Conclusion
The intramolecular dehydrogenative dehydro-Diels–Alder reac- tion in hexanes) was added dropwise via syringe. The reaction
tion (IDDDA) of chlorinated styrenes allowed access to was stirred at −78 °C for 45 min, and anhydrous N,N-dimethyl-
cyclopenta[b]naphthalene carbaldehydes containing a chlorine formamide (DMF, 2.0 equiv.) was added dropwise via syringe.
atom in various positions of the aromatic motif. Conversion of The reaction mixture was stirred at −78 °C for 30 min, and was
the aryl chloride to the corresponding dimethylamino group then warmed to rt and stirred for 2 h. The reaction mixture
(donor) under Buchwald–Hartwig cross-coupling conditions was added to a cold solution of ethyl acetate and 10% KH2PO4,
followed by Knoevenagel condensation of the carbonyl group and stirred for 30 min. The aqueous layer was separated and
with a cyanoacrylate motif (acceptor) produced a family of fluo- extracted with ethyl acetate (1×). The organic layer was then
rescent dyes containing the D–π-A array. Fluorescence emis- washed with brine, dried over magnesium sulfate, gravity
sion of these dyes was found to depend on the solvent polarity filtered, and concentrated under reduced pressure. The
and viscosity, a behavior that is typical for molecular rotors. crude product was purified by flash silica gel column
Moreover, placing the donor and acceptor groups in close chromatography.
(E)-8-(2-Chlorophenyl)oct-7-en-2-ynal (7a). Follows general
procedure A: enyne 6a (0.500 g, 2.44 mmol), THF (6.5 mL),
n-butyllithium (1.52 mL, 2.44 mmol), DMF (0.38 mL,
4.88 mmol), ethyl acetate (8 mL), 10% KH2PO4 (15 mL). The
reaction mixture turned orange upon addition of n-butyl-
lithium and yellow upon addition of DMF. The crude product
was purified by flash silica gel column chromatography
(3.0 cm column, 5% ethyl acetate/hexanes) to yield the title
compound as a light yellow oil (0.455 g, 80%). 1H NMR
(400 MHz, CDCl3) δ 9.19 (s, 1H), 7.48 (d, J = 7.8, 1.2 Hz, 1H),
7.34 (d, J = 7.8, 1.2 Hz, 1H), 7.22–7.13 (m, 2H), 6.81 (d, J =
15.6 Hz, 1H), 6.15 (dt, J = 15.6, 7.2 Hz, 1H), 2.49 (t, J = 7.2 Hz,
2H), 2.39 (q, J = 7.2 Hz, 2H), 1.82 (p, J = 7.2 Hz, 2H); 13C NMR
(100 MHz, CDCl3) δ 176.9, 135.4, 132.5, 131.7, 129.5, 128.1,
127.5, 126.7, 126.6, 98.4, 82.0, 32.0, 26.2, 18.4; IR (thin film)
proximity along the π-ring structure (i.e. CNCA dye 5) was
found to influence viscosity sensitivity. In this case, absence of
viscosity sensitivity combined with a high quantum yield
points to the inability of the molecule to perform intramole-
cular twisting. The pattern of viscosity sensitivity in CNCA dyes
3 and 4 alludes to the existence of a twisted ground state with
nonradiative emission from a planar excited state. These obser-
vations help to understand the influence of chemical structure
on viscosity sensitivity of fluorescent molecular rotors.
Experimental
General notes
All reagents were procured from commercial sources and were
used as received. Tetrahydrofuran (THF) was dried by passing
through alumina using the Sol-Tek ST-002 solvent purification
system. All microwave-mediated reactions were conducted
using either a Biotage Initiator Exp or Anton Paar Monowave
300 microwave synthesizer. The microwave parameters were set
to variable power, constant temperature, and a fixed hold time.
Thin-layer chromatography (TLC) analyses were performed on
Silicycle SiliaPlate G silica gel glass plates (60F-254, 0.25 mm
thickness) and visualized under UV light. Purification of sub-
strates by flash column chromatography was performed with
silica gel (40–63 μm particle size, 60 Å pore size) purchased
from Sorbent Technologies. 1H and 13C NMR spectra were
recorded on Bruker Avance 300, 400, or 500 MHz spectro-
meters. Spectra were referenced to residual chloroform
3307, 3066, 2938, 2860, 2740, 2278, 2200, 1667, 796, 751 cm−1
;
LRMS (TOF MSMS ES+) m/z (%) 233 (100), 214 (35), 205 (20);
HRMS (TOF MS ES+) calculated for C14H14OCl (M + H)+
233.0733, found 233.0726; TLC (UV, KMnO4 stain) Rf
=
0.3 (10% ethyl acetate/hexanes).
(E)-8-(3-Chlorophenyl)oct-7-en-2-ynal (7b). Follows general
procedure A: enyne 6b (0.495 g, 2.42 mmol), THF (6.5 mL),
n-butyllithium (1.51 mL, 2.42 mmol), DMF (0.37 mL,
4.84 mmol), ethyl acetate (8 mL), 10% KH2PO4 (15 mL). The
reaction mixture turned yellow upon addition of n-butyllithium
and lighter upon addition of DMF. The crude product was pur-
ified by flash silica gel column chromatography (3.0 cm
column, 5% ethyl acetate/hexanes) to yield the title compound
1
as a light yellow oil (0.426 g, 76%). H NMR (300 MHz, CDCl3)
1
(7.27 ppm, H, 77.16 ppm, 13C). Chemical shifts are reported
δ 9.19 (t, J = 0.9 Hz, 1H), 7.33 (t, J = 1.8 Hz, 1H), 7.23–7.16 (m,
3H), 6.38 (d, J = 15.9 Hz, 1H), 6.18 (dt, J = 15.9, 7.2 Hz, 1H),
2.47 (t, J = 7.2 Hz, 2H), 2.35 (dq, J = 7.2, 1.2 Hz, 2H), 1.79 (p,
J = 7.2 Hz, 2H); 13C NMR (100 MHz, CDCl3) δ 177.1, 139.1,
134.3, 130.3, 129.9, 129.7, 127.0, 125.8, 124.2, 98.4, 81.9, 31.7,
26.9, 18.4; IR (thin film) 3072, 3023, 2938, 2860, 2733, 2277,
2200, 1667, 1592, 1476, 776 cm−1; LRMS (TOF MSMS ES+) m/z
(%) 233 (100), 215 (59), 205 (86), 198 (12), 197 (29), 179 (16);
HRMS (TOF MS ES+) calculated for C14H14OCl (M + H)+
233.0733, found 233.0735; TLC (UV, KMnO4 stain) Rf = 0.4
(10% ethyl acetate/hexanes).
in ppm, and multiplicities are indicated by s (singlet), d
(doublet), t (triplet), q (quartet), p (pentet), m (multiplet), and
b (broad). Coupling constants, J, are reported in hertz (Hz).
Infrared (IR) spectra were obtained using a Nicolet Avatar E.S.
P. 360 FT-IR. Electron spray ionization (ES) spectra were
recorded on a Waters Q-TOF Ultima API, Micromass UK
Limited high resolution mass spectrometer. Substrates 6a–c
were previously synthesized.17a
General procedure A for the synthesis of formyl enynes
7. To a flame-dried two-neck round-bottomed flask equipped
with an argon inlet adapter, a septum, and a stir bar was
added enyne 6a–c (1.0 equiv.) and THF (0.4 M) via syringe. The
(E)-8-(4-Chlorophenyl)oct-7-en-2-ynal (7c). Follows general
procedure A: enyne 6c (0.500 g, 2.44 mmol), THF (6.5 mL),
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n-butyllithium (1.52 mL, 2.44 mmol), DMF (0.38 mL, (M + H)+ 231.0577, found 231.0568; TLC (UV, KMnO4 stain)
4.88 mmol), ethyl acetate (8 mL), 10% KH2PO4 (15 mL). The Rf = 0.4 (10% ethyl acetate/hexanes).
6-Chloro-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-carbalde-
hyde (8c). Follows general procedure B: aldehyde 7c (0.425 g,
1.83 mmol), o-dichlorobenzene (20 mL). The reaction mixture
was first concentrated under high vacuum, and then the crude
product was purified by flash silica gel column chromato-
graphy (3.0 cm column, 5% ethyl acetate/hexanes) to yield the
title compound as a white solid (0.308 g, 73%). MP: 93–96 °C;
1H NMR (300 MHz, CDCl3) δ 10.7 (s, 1H), 9.18 (s, 1H), 7.87 (s,
1H), 7.72 (d, J = 8.7 Hz, 1H), 7.44 (dd, J = 8.7, 1.8 Hz, 1H), 3.47
(t, J = 7.5 Hz, 2H), 3.05 (dq, J = 7.5, 2.7 Hz, 2H), 2.24 (p, J =
7.5 Hz, 2H); 13C NMR (75 MHz, CDCl3) δ 191.8, 153.3, 143.8,
134.1, 131.4, 130.5, 129.2 (2C), 126.7, 124.5, 123.8, 31.6, 31.2,
reaction mixture turned light yellow upon addition of DMF.
The crude product was purified by flash silica gel column
chromatography (3.0 cm column, 5% ethyl acetate/hexanes) to
yield the title compound as a light yellow oil (0.389 g, 68%). 1H
NMR (300 MHz, CDCl3) δ 9.19 (s, 1H), 7.27 (s, 4H), 6.39 (d, J =
15.9 Hz, 1H), 6.15 (dt, J = 15.9, 6.9 Hz, 1H), 2.48 (t, J = 6.9 Hz,
2H), 2.35 (q, J = 6.9 Hz, 2H), 1.80 (p, J = 6.9 Hz, 2H); 13C NMR
(100 MHz, CDCl3) δ 177.0, 135.8, 132.5, 129.9, 129.4, 128.5
(2C), 127.1 (2C), 98.4, 81.9, 31.7, 26.9, 18.4; IR (thin film) 3026,
2936, 2857, 2729, 2200, 1665, 1489 cm−1; LRMS (TOF MSMS
ES+) m/z (%) 233 (100), 232 (92), 215 (35); HRMS (TOF MS ES+)
calculated for C14H13OCl (M)+ 232.0655, found 232.0632; TLC
(UV, KMnO4 stain) Rf = 0.3 (10% ethyl acetate/hexanes).
25.5; IR (thin film) 3096, 2958, 2913, 2868, 2753, 1675 cm−1
;
LRMS (TOF MSMS ES+) m/z (%) 231 (100), 230 (75), 203 (45),
General procedure B for the synthesis of naphthalene carbal-
dehydes 8. To a microwave irradiation vial equipped with a
stir bar was added enyne 7a–c in o-dichlorobenzene (0.06–0.09
M). The reaction mixture was irradiated with stirring at 180 °C
for 60 min. During this time, the reaction mixture turned
golden in color. The reaction mixture was then purified
directly by flash silica gel column chromatography.
195 (32); HRMS (TOF MS ES+) calculated for C14H11OCl (M)+
230.0498, found 230.0471; TLC (UV, KMnO4 stain) Rf
=
0.3 (10% ethyl acetate/hexanes).
5-Chloro-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-carbalde-
hyde (8d). Formed as a mixture with 8b. White solid (0.058 g,
1
48%); MP: 84–86 °C; H NMR (300 MHz, CDCl3) δ 11.2 (s, 1H),
7.77 (s, 1H), 7.73 (dd, J = 7.5, 1.2 Hz, 1H), 7.44 (dd, J = 7.5,
1.2 Hz, 1H), 7.36 (t, J = 7.5 Hz, 1H), 3.19 (t, J = 7.5 Hz, 2H), 3.06
(dt, J = 7.5, 1.2 Hz, 2H), 2.14 (p, J = 7.5 Hz, 2H); 13C NMR
(100 MHz, CDCl3) δ 194.3, 147.0, 144.6, 135.1, 129.8, 129.7,
128.3, 128.1, 127.7, 126.5, 125.6, 33.1, 32.3, 25.6; IR (thin film)
2959, 2927, 2853, 2751, 1682 cm−1; LRMS (TOF MSMS ES+)
m/z (%) 203 (100), 175 (25), 168 (60); HRMS (TOF MS ES+) cal-
culated for C14H12OCl (M + H)+ 231.0577, found 231.0564; TLC
(UV, KMnO4 stain) Rf = 0.5 (10% ethyl acetate/hexanes).
General procedure C for the synthesis of aminonaphtha-
lenes 9. To an oven-dried microwave irradiation vial equipped
with a stir bar was added RuPhos palladacycle (3 mol%),
cesium carbonate (2.0–2.2 equiv.), and naphthalene 8a–c
(1.0 equiv.). The vial was capped and evacuated and refilled
with argon (3×) by piercing the septum of the cap with a small
gauge needle connected to an argon manifold. THF (0.5 M)
was added via syringe with stirring, followed by dimethylamine
(3.0–3.3 equiv. of a 2.0 M solution in THF). The argon inlet
8-Chloro-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-carbalde-
hyde (8a). Follows general procedure B: aldehyde 7a (0.279 g,
1.20 mmol), o-dichlorobenzene (20 mL). The reaction mixture
was purified directly by flash silica gel column chromato-
graphy (2.5 cm column, 0–10% ethyl acetate/hexanes) to yield
the title compound as a white solid (0.222 g, 80%). MP:
92–96 °C; 1H NMR (300 MHz, CDCl3) δ 10.8 (s, 1H), 9.02 (d, J =
8.7 Hz, 1H), 8.41 (s, 1H), 7.58 (d, J = 7.5 Hz, 1H), 7.47 (t, J =
7.5 Hz, 1H), 3.47 (t, J = 7.5 Hz, 2H), 3.13 (dt, J = 7.5, 1.2 Hz,
2H), 2.25 (p, J = 7.5 Hz, 2H); 13C NMR (100 MHz, CDCl3)
δ 192.1, 152.5, 144.9, 132.0, 131.7, 130.5, 127.5, 126.3, 125.7,
125.5, 123.4, 32.1, 31.6, 25.6; IR (thin film) 3080, 2968, 2929,
2868, 2766, 1677, 1615, 1596, 1490 cm−1; LRMS (TOF MSMS
ES+) m/z (%) 231 (42), 203 (100), 195 (30), 175 (44), 168 (50),
167 (20); HRMS (TOF MS ES+) calculated for C14H12OCl (M +
H)+ 231.0577, found 231.0581; TLC (UV, KMnO4 stain) Rf =
0.3 (10% ethyl acetate/hexanes).
7-Chloro-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-carbalde-
hyde (8b). Follows general procedure B: aldehyde 7b (0.122 g, was removed and the reaction mixture was lowered into a pre-
0.52 mmol), o-dichlorobenzene (7 mL). The reaction mixture heated 85 °C oil bath and stirred until complete by TLC. The
was first concentrated under high vacuum, and then the crude reaction mixture was then cooled to rt and quenched with
product was purified by flash silica gel column chromato- aqueous saturated ammonium chloride. The aqueous layer
graphy (3.0 cm column, 5% ethyl acetate/hexanes) to yield the was separated and extracted with ethyl acetate (3×). The com-
title compound as a white solid (0.037 g, 31%). Substrate 8b bined organic layers were washed with brine, dried over mag-
was formed as a 1 : 1.5 mixture with 8d in a total yield of 79%. nesium sulfate, gravity filtered, and concentrated under
1
MP: 66–67 °C; H NMR (300 MHz, CDCl3) δ 10.8 (s, 1H), 9.07 reduced pressure. The crude product was then purified by
(d, J = 9.1 Hz, 1H), 7.80 (s, 1H), 7.77 (d, J = 2.1 Hz, 1H), 7.51 flash silica gel column chromatography.
8-(Dimethylamino)-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-
carbaldehyde (9a). Follows general procedure C: 0.5–2 mL vial,
RuPhos palladacycle (0.004 g, 0.0055 mmol), cesium carbonate
(0.143 g, 0.44 mmol), naphthalene 8a (0.050 g, 0.22 mmol),
THF (0.43 mL), dimethylamine (0.33 mL, 0.66 mmol). The
reaction mixture was heated for 4.5 h and turned from tan to
orange in color over time. The crude product was purified by
(dd, J = 9.1, 2.1 Hz, 1H), 3.45 (t, J = 7.5 Hz, 2H), 3.08 (dt, J =
7.5, 1.2 Hz, 2H), 2.24 (p, J = 7.5 Hz, 2H); 13C NMR (100 MHz,
CDCl3) δ 192.1, 152.4, 144.8, 134.1, 131.7, 128.4 (2C), 128.3,
126.6, 126.2, 125.5, 31.8, 31.3, 25.6; IR (thin film) 2956, 2868,
2843, 2756, 1682, 1621, 1596, 1489 cm−1; LRMS (TOF MSMS
ES+) m/z (%) 230 (10), 229 (63), 212 (40), 201 (48), 175 (100),
168 (71); HRMS (TOF MS ES+) calculated for C14H12OCl
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flash silica gel column chromatography (2.0 cm column, 5%
General procedure D for the synthesis of CNCA dyes 3–5. To
ethyl acetate/hexanes) to yield the title compound as a yellow stirring solution of aldehyde 9a–c (1.0 equiv.) and 10 (0.8
solid (0.032 g, 60%). MP: 59–61 °C; 1H NMR (300 MHz, CDCl3) equiv.) in THF was added DBU (0.01–0.03 equiv.) under argon.
δ 10.8 (s, 1H), 8.72 (d, J = 8.4 Hz, 1H), 8.40 (s, 1H), 7.49 (t, J = The reaction mixture was heated at 50 °C for 2–3 h then cooled
8.4 Hz, 1H), 7.13 (d, J = 7.2 Hz, 1H), 3.45 (t, J = 7.5 Hz, 2H), to rt and concentrated under reduced pressure. The residue
3.10 (dt, J = 7.5, 1.2 Hz, 2H), 2.88 (s, 6H), 2.22 (p, J = 7.5 Hz, was purified by flash silica gel column chromatography to give
2H); 13C NMR (100 MHz, CDCl3) δ 192.6, 151.3, 151.1, CNCA dyes as colored oils.
N5-CNCA (3). Follows general procedure D: 9a (0.020 g,
0.084 mmol), 10 (0.015 g, 0.067 mmol), THF (0.33 mL), and
DBU (0.001 mmol). The reaction mixture was heated at 50 °C
for 2 h. The residue was purified by flash silica gel column
chromatography (0–20% ethyl acetate/hexanes) to yield the
title compound as an orange oil (0.026 g, 86%). 1H NMR
(400 MHz, CDCl3) δ 8.96 (s, 1H), 8.23 (s, 1H), 7.41–7.39 (m,
142.8, 132.1, 128.8, 127.7, 126.0, 125.6, 119.0, 114.5, 45.5 (2C),
32.2, 31.8, 25.7; IR (thin film) 3583, 3072, 2942, 2859,
2828, 2783, 1680, 1616, 1593 cm−1; LRMS (TOF MSMS ES+)
m/z (%) 240 (65), 225 (100), 211 (78), 197 (86); HRMS (TOF MS
ES+) calculated for C16H18NO (M + H)+ 240.1388, found
240.1372; TLC (UV, KMnO4 stain) Rf = 0.3 (10% ethyl acetate/
hexanes).
7-(Dimethylamino)-2,3-dihydro-1H-cyclopenta[b]naphthalene-4- 2H), 7.11 (dd, J = 4.0, 1.2 Hz, 1H), 4.53–4.51 (m, 2H), 3.87–3.85
carbaldehyde (9b). Follows general procedure C: 0.5–2 mL vial,
RuPhos palladacycle (0.005 g, 0.0063 mmol), cesium carbonate
(0.164 g, 0.50 mmol), naphthalene 8b (0.052 g, 0.23 mmol),
THF (0.50 mL), dimethylamine (0.38 mL, 0.75 mmol). The
reaction mixture was heated for 3 h and turned from burnt
orange to fluorescent yellow in color over time. The crude
product was purified by flash silica gel column chromato-
graphy (1.75 cm column, 5% ethyl acetate/hexanes) to yield the
(m, 2H), 3.76–3.74 (m, 2H), 3.71–3.66 (m, 4H), 3.56–3.54 (m,
2H), 3.36 (s, 3H), 3.14–3.09 (m, 4H), 2.88 (s, 6H), 2.20–2.14 (m,
2H); 13C (100 MHz, CDCl3) δ 162.0, 156.8, 151.5, 144.6, 143.0,
131.7, 128.3, 126.8, 124.8, 122.3, 118.7, 115.0, 114.6, 109.6,
72.0, 71.0, 70.8, 70.7, 68.8, 66.2, 59.0, 45.5 (2C), 33.4, 33.3, 29.8
(grease), 26.7; HRMS (ESI) calculated for C26H33N2O5 (M + H)+
453.2384, found 453.2387; TLC (UV) Rf = 0.3 (40% ethyl
acetate/hexanes).
title compound as a yellow solid (0.024 g, 44%). MP:
N6-CNCA (4). Follows general procedure D: 9b (0.0070 g,
1
122–125 °C; H NMR (300 MHz, CDCl3) δ 10.8 (s, 1H), 8.95 (d, 0.030 mmol), 10 (0.0054 g, 0.023 mmol), THF (0.20 mL), and
J = 9.3 Hz, 1H), 7.73 (s, 1H), 7.26–7.21 (m, 1H), 6.88 (s, 1H), DBU (0.001 mmol). The reaction mixture was heated at 50 °C
3.40 (t, J = 7.5 Hz, 2H), 3.05–3.00 (m, 8H), 2.19 (p, J = 7.5 Hz, for 2 h. The residue was purified by flash silica gel column
2H); 13C NMR (100 MHz, CDCl3) δ 192.8, 148.4, 148.2, 143.7, chromatography (0–40% ethyl acetate/hexanes) to yield the
135.1, 128.1, 125.5, 125.2, 123.2, 117.6, 106.8, 40.6 (2C), 31.9, title compound as an orange oil (0.0094 g, 90%). 1H NMR
31.2, 25.8; IR (thin film) 2953, 2921, 2843, 2802, 2745, 1668, (400 MHz, CDCl3) δ 8.92 (s, 1H), 7.61–7.58 (m, 2H), 7.15 (dd,
1618, 1605, 1510 cm−1; LRMS (TOF MSMS ES+) m/z (%) 239 J = 12.0, 4.0 Hz, 1H), 6.89 (d, J = 4.0 Hz, 1H), 4.52–4.50 (m,
(25), 238 (75), 225 (100), 210 (45), 197 (35); HRMS (TOF MS 2H), 3.87–3.85 (m, 2H), 3.75–3.74 (m, 2H), 3.71–3.66 (m, 5H),
ES+) calculated for C16H18NO (M + H)+ 240.1388, found 3.56–3.54 (m, 2H), 3.37 (s, 3H), 3.05 (m, 9H), 2.17–2.10 (m,
240.1399; TLC (UV, KMnO4 stain) Rf = 0.1 (10% ethyl acetate/ 2H); 13C (100 MHz, CDCl3) δ 162.2, 156.7, 148.6, 143.9, 141.2,
hexanes).
134.5, 125.0, 124.8, 124.3, 123.0, 116.2, 115.0, 109.1, 107.3,
72.1, 71.0, 70.8, 70.7, 68.8, 65.9, 59.2, 40.9 (2C), 33.1, 33.0, 29.8
(grease), 26.7; HRMS (ESI) calculated for C26H33N2O5 (M + H)+
453.2384, found 453.2381; TLC (UV) Rf = 0.2 (40% ethyl
acetate/hexanes).
N7-CNCA (5). Follows general procedure D: 9c (0.020 g,
0.084 mmol), 10 (0.015 g, 0.067 mmol), THF (0.33 mL),
and DBU (0.001 mmol). The reaction mixture was heated
at 50 °C for 3 h. The residue was purified by flash silica
gel column chromatography (0–60% ethyl acetate/hexanes)
to yield the title compound as a red oil (0.023 g, 77%).
1H NMR (400 MHz, CDCl3) δ 8.91 (s, 1H), 7.67–7.64 (m,
6-(Dimethylamino)-2,3-dihydro-1H-cyclopenta[b]naphthalene-4-
carbaldehyde (9c). Follows general procedure C: 2–5 mL vial,
RuPhos palladacycle (0.017 g, 0.023 mmol), cesium carbonate
(0.590 g, 1.81 mmol), naphthalene 8c (0.200 g, 0.87 mmol),
THF (1.8 mL), dimethylamine (1.37 mL, 2.73 mmol). The reac-
tion mixture was heated for 2.5 h and turned from tan to
orange in color over time. The crude product was purified by
flash silica gel column chromatography (2.5 cm column, 5%
ethyl acetate/hexanes) to yield the title compound as an orange
solid (0.154 g, 74%). MP: 132–133 °C; 1H NMR (300 MHz,
CDCl3) δ 10.7 (s, 1H), 8.43 (d, J = 2.4 Hz, 1H), 7.74 (s, 1H), 7.64
(d, J = 9.1 Hz, 1H), 7.12 (dd, J = 9.1, 2.4 Hz, 1H), 3.43 (t, J =
7.5 Hz, 2H), 3.11 (s, 6H), 3.01 (t, J = 7.5 Hz, 2H), 2.19 (p, J =
7.5 Hz, 2H); 13C NMR (125 MHz, CDCl3) δ 192.5, 153.4,
150.5, 138.5, 132.2, 129.8, 129.0, 126.6, 123.6, 115.2, 103.6,
40.6 (2C), 31.6, 31.5, 25.7; IR (thin film) 3068, 2952, 2847,
2794, 2751, 1665, 1612, 1511, 1159 cm−1; LRMS (TOF MSMS
ES+) m/z (%) 240 (80), 239 (100), 226 (59), 211 (60); HRMS (TOF
MS ES+) calculated for C16H17NO (M)+ 239.1310, found
239.1289; TLC (UV, KMnO4 stain) Rf = 0.5 (25% ethyl acetate/
hexanes).
2H), 7.11 (dd,
J = 8.0, 4.0 Hz, 1H), 6.63 (bs, 1H),
4.52–4.50 (m, 2H), 3.88–3.85 (m, 2H), 3.76–3.74 (m, 2H),
3.70–3.66 (m, 4H), 3.56–3.54 (m, 2H), 3.36 (s, 3H),
3.10–3.03 (m, 10H), 2.17–2.10 (m, 2H); 13C (100 MHz,
CDCl3) δ 162.5, 156.4, 148.9, 145.9, 139.2, 132.0, 129.4,
126.3, 125.8, 122.7, 115.6, 115.3, 108.0, 103.2, 72.1, 71.0,
70.8, 70.7, 68.8, 65.9, 59.2, 40.9 (2C), 33.8, 32.7, 29.8
(grease), 26.7; HRMS (ESI) calculated for C26H33N2O5 (M +
H)+ 453.2384, found 453.2386; TLC (UV) Rf = 0.34 (50%
ethyl acetate/hexanes).
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thank the National Science Foundation for instrumentation
grants CHE9709183 and CHE0741968. We also thank
Dr Anthony Mrse (UCSD NMR Facility) and Dr Yongxuan Su
(UCSD MS Facility).
Table 2 Mixture ratios and resulting viscosity of the alcohol gradient
Target
viscosity
(mPa s)
Pre-stained
ethylene
glycol
Unstained
ethylene
glycol
Methanol
Glycerol
4
8
16
35
77
167
1 mL
1 mL
1 mL
1 mL
1 mL
1 mL
2 mL
3 mL
4 mL
3 mL
2 mL
1 mL
2 mL
1 mL
—
—
—
—
—
—
1 mL
2 mL
3 mL
Notes and references
1 For recent selected reviews and monographs on this topic
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ill.
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—
Fluorescence spectroscopy
Dye stock solutions were prepared by dissolving each dye in
methanol at a concentration of 5 mM and were kept at 4 °C
until used. Final dye solutions were prepared by pipetting 10
µL of the stock solution into 5 mL of the solvent under vigo-
rous stirring for a resulting concentration of 10 µM. Solutions
of the compounds in toluene, acetonitrile, DMSO, methanol,
and a 50% by volume mix of water and methanol were
analyzed. Viscosity gradients were composed of a mixture of
methanol, ethylene glycol, and glycerol at different concen-
trations as reported elsewhere.27 Specifically, mixtures of
methanol, ethylene glycol, and glycerol were prepared by pre-
staining 10 mL of ethylene glycol with 10 µL dye stock solu-
tion. One mL of prestained ethylene glycol was then mixed
with either methanol or glycerol and unstained ethylene glycol
to afford solutions of different viscosity and constant dye con-
centration at a total volume of 5 mL as detailed in Table 2.
Viscosity values for the individual solvents were taken from
the labels of the stock bottles or from the pertinent literature28
as 0.5 mPa s for methanol, 16 mPa s for ethylene glycol, and
864 mPa s for glycerol, all at 22 °C. The viscosity ηmix of the
mixtures was calculated with eqn (2)
X
lnηmix
¼
wi lnηi
ð2Þ
i
where wi is the volume fraction and ηi the viscosity of the i-th
solvent. Solvent polarity values for methanol of 0.762, for ethyl-
ene glycol of 0.790, and for glycerol of 0.812 were used.22 For
comparison, the relative polarity of toluene is 0.099, of aceto-
nitrile 0.460, and of DMSO 0.444.
Spectroscopy was performed using a Fluoromax-3 fluoro-
photometer (Jobin-Yvon, Edison, NJ) with a temperature-con-
trolled four-sample holder (Turret-400, Quantum Northwest,
Liberty Lake, WA) set to 22.0 °C. Disposable methacrylate
cuvettes were used for the viscosity gradients, and glass cuvettes
for all other solvents. Unless otherwise stated, emission scans
were performed from the optimum excitation wavelength.
8 M. A. Haidekker and E. A. Theodorakis, Org. Biomol. Chem.,
2007, 5, 1669–1678.
9 For selected recent applications of fluorescent molecular
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Acknowledgements
Financial support from the National Institutes of Health (CA
133002, EAT), (1R21 RR025358, MAH) and the Andrew Mellon
Predoctoral Fellowship (LSK) is gratefully acknowledged. We
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Paper
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