Computational Design and Synthesis of a Deeply Red-Shifted and Bistable Azobenzene

Article abstract of DOI:10.1021/jacs.9b10430

We computationally dissected the electronic and geometrical influences of ortho-chlorinated azobenzenes on their photophysical properties. X-ray analysis provided the insight that trans-tetra-ortho-chloro azobenzene is conformationally flexible and thus subject to molecular motions. This allows the photoswitch to adopt a range of red-shifted geometries, which account for the extended n → π? band tails. On the basis of our results, we designed the di-ortho-fluoro di-ortho-chloro (dfdc) azobenzene and provided computational evidence for the superiority of this substitution pattern to tetra-ortho-chloro azobenzene. Thereafter, we synthesized dfdc azobenzene by ortho-chlorination via 2-fold C-H activation and experimentally confirmed its structural and photophysical properties through UV-vis, NMR, and X-ray analyses. The advantages include near-bistable isomers and an increased separation of the n → π? bands between the trans- and cis-conformations, which allows for the generation of unusually high levels of the cis-isomer by irradiation with green/yellow light as well as red light within the biooptical window.

Full text of DOI:10.1021/jacs.9b10430

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Computational Design and Synthesis of a Deeply Red-Shifted and
Bistable Azobenzene
,∇
∇
̈
David B. Konrad,* Gokcen Savasci, Lars Allmendinger, Dirk Trauner, Christian Ochsenfeld,*
and Ahmed M. Ali*
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ABSTRACT: We computationally dissected the electronic and
geometrical influences of ortho-chlorinated azobenzenes on their
photophysical properties. X-ray analysis provided the insight that
trans-tetra-ortho-chloro azobenzene is conformationally flexible and
thus subject to molecular motions. This allows the photoswitch to
adopt a range of red-shifted geometries, which account for the
extended n → π* band tails. On the basis of our results, we designed
the di-ortho-fluoro di-ortho-chloro (dfdc) azobenzene and provided
computational evidence for the superiority of this substitution
pattern to tetra-ortho-chloro azobenzene. Thereafter, we synthesized
dfdc azobenzene by ortho-chlorination via 2-fold C−H activation
and experimentally confirmed its structural and photophysical properties through UV−vis, NMR, and X-ray analyses. The
advantages include near-bistable isomers and an increased separation of the n → π* bands between the trans- and cis-conformations,
which allows for the generation of unusually high levels of the cis-isomer by irradiation with green/yellow light as well as red light
within the biooptical window.
A strategy for generating red-shifted photoswitches entails
the separation of the n → π* absorption bands, which are
typically overlapping for cis- and trans-azobenzene in the visible
spectrum. In this context, a significant advancement was made
through tetra-ortho-substitution with heteroatoms.18−21 Tetra-
ortho-methoxy azobenzenes, for instance, have long-lived cis-
isomers (τ ≈ 2 d) and are readily photoswitched to their trans-
and cis-configurations with blue and green/red light,
respectively.20 Modification of the tetra-ortho-methoxy pattern
with electron-donating substituents implements an azonium
ion, which can be isomerized with near-infrared light (up to
720 nm) but suffers from a significant decrease in the thermal
stability of the cis-isomer (τ = 1 s).22,23 In contrast, the cis-
isomer of the tetra-ortho-fluoro azobenzene has a thermal half-
life on the order of years (t1/2 = 700 d) and can be reliably
photoswitched with blue (trans-configuration) and green (cis-
configuration) light.19,20 Significant photoconversion to the cis-
isomer can be reached with up to 527 nm light.24 The use of
longer wavelengths requires the use of an alternative
substitution pattern such as the tetra-ortho-chloro azoben-
INTRODUCTION
■
Azobenzenes are versatile photoswitches that can be cycled
between their cis- and trans-configurations with light.1,2
Because of their small size, robust photoswitching, synthetic
accessibility, and low rate of photobleaching, they serve as
excellent building blocks for the generation of elaborate optical
tools.3−5 Photopharmaceuticals, for example, contain azoben-
zene fragments as ON- and OFF-switches, which allow for the
control of biological functions with the spatiotemporal
precision of light.6−11 To elicit the full potential of these
optical tools and to apply them to complex animal tissues, it is
crucial to use tissue-penetrating, nonhazardous red and near-
infrared (NIR) light within the biooptical window12 (650−
950 nm).13−15 In addition, it is highly desirable to use
photoswitches with slow thermal relaxation rates for the less
stable isomers and near-quantitative photoconversion for each
isomer. Fast-relaxing photopharmaceuticals require constant
illumination with high intensity light for their photoswitching
to outcompete the thermal relaxation. This becomes
increasingly difficult in deeper tissue layers.16 The batho-
chromic shift, the thermal stability, and the photoconversion
strongly depend on the substitution pattern of the azobenzene
core.7,16,17 Therefore, the design of new photopharmaceuticals
that are operated at the biooptical window relies on the
availability of azobenzene photoswitches with the appropriate
photophysical properties.
Received: September 27, 2019
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zene.24 It is worth noting that certain photoswitches can be
operated with light in the near-infrared region (NIR) by using
two-photon excitation.25 Because of the slow photoswitching
through two-photon excitation, however, this approach
requires the use of photoswitches with long-lived isomers to
reach significant photoconversion.26
Remarkably, tetra-ortho-chloro azobenzenes allow for rapid
and efficient photoswitching by irradiation of the n → π* band
tails. For example, the fastest isomerization rate to the
photostationary state (PSS) with the highest content of the
cis-isomer for alkyl-substituted derivatives, such as red-AzCA-4
(Figure 1), was reached by using green light (550−560 nm),
0 K followed by the computation of the vertical excitation
energies on the TD-PBE0/def2-TZVP level. This gives access
to absorption maxima (λmax) of the π → π* (S0 → S2) as well
as the n → π* (S0 → S1) transitions for optimized trans- and
cis-azobenzene geometries.
EFFECTS OF THE AZOBENZENE
ORTHO-CHLORINATION
■
The geometrical changes in the conformation of azobenzenes
through isomerization or substitution can be subdivided into
three main categories: changes in the N−N bond length, the
C(1)−C(2)−N(3)−N(4) dihedral angles Φ, and the C(2)−
N(3)−N(4)−C(5) dihedral angle Ψ (Figure 2). The CCNN
Figure 2. Numerical descriptors for the dihedral angles.
dihedral angles describe the rotation of the aryl rings out of the
plane. This rotation can proceed from two directions and thus
can be described by two dihedral angles for each aryl ring: ΦS
and ΦL. We focus our discussion on the smaller ΦS angles. The
Ψ dihedral angle describes the distortion of the N−N double
bond.
For the evaluation of the influence of tetra-ortho-
chlorination on the bathochromic shifts of azobenzene, we
computationally optimized structures for trans-1, cis-1, trans-2,
and cis-2 and verified that their conformations are true minima
on the potential energy surface (PES). We found that trans-
azobenzene has a planar structure with an N−N bond length of
1.242 Å and N−N bond geometry (Ψ dihedral angle) of
180.0°. The structure of trans-tetra-ortho-chloro azobenzene is
characterized by the rotation of both aryl rings out of the plane
by 50.8° (ΦS). In addition, the N−N bond length is shortened
to 1.236 Å, and the N−N double bond is slightly distorted to
175.3° (Ψ). Next, we determined their n → π* (S0 → S1)
absorption bands (Table 1) and found that the conversion of
Figure 1. Structures of red-AzCA-4, azobenzene (1), and tetra-ortho-
chloro azobenzene (2).
although the λmax of the trans-isomer (400 nm-adapted) lies
close to 450 nm.27 It was further demonstrated that irradiation
at far ends of the n → π* band can still produce a PSS with a
significant cis-content by using bright light sources and
continuous irradiation. For instance, Woolley and co-workers28
reported photoswitching of tetra-ortho-chloro azobenzenes
with high intensity red light (625 nm) for 1 min, whereas
Feringa and co-workers24 applied longer-wavelength high
intensity red light (652 nm) for 2.5 h to reach a PSS with
the highest levels of the cis-isomer.
Intrigued by the potential of the tetra-ortho-chloro
substitution pattern (Figure 1) as a basis to develop new
photoswitches with long-lived cis-isomers, near-quantitative
PSS, and excitation wavelengths within the biooptical window,
we focus our study on the evaluation of the influence of
azobenzene ortho-chlorination on the photophysical properties.
By dissecting the effects of the chlorination into its geometric
and electronic subcomponents, we aim to identify the origin of
the extended red-shifted n → π* band tails of ortho-chlorinated
azobenzenes as well as the separation of the n → π* excitation
between the trans- and cis-isomers. Therefore, we employed a
combination of quantum-chemical methodologies as well as X-
ray analyses. On the basis of our results, the tetra-ortho
substitution pattern could be optimized to yield a photoswitch
with a highly stable cis-isomer and PSS with high contents for
each isomer while retaining the red-shifted transition.29,30
We first computed the geometries of our studied azobenzene
substitution patterns as well as their π → π* and n → π*
absorption bands. Previous studies have shown that employing
a PBE0 hybrid functional provides the closest correlation
between the computational and experimental results for
determining the λmax of azobenzene-based photoswitches.31
Therefore, we optimized the structures for all studied
photoswitches on the PBE0-D3/def2-TZVP32−35 level of
theory using the TURBOMOLE program package, to obtain
the respective gas-phase ground-state minimum geometry at
Table 1. Optimized Conformations of 1 and 2: Vertical
Excitation Energies (TD-PBE0/def2-TZVP) and
Photoswitch Structures (PBE0-D3/def2-TZVP)
comp.
NN [Å]
ΦS [deg]
Ψ [deg]
S0 → S1 [nm]
trans-1
cis-1
trans-2
cis-2
1.242
1.234
1.236
1.229
0.0
52.0
50.8
60.5
180.0
8.5
175.3
5.1
483.6
470.0
498.5
474.9
azobenzene (1) to 2 induces a bathochromic shift of the n →
π* absorption band of 14.9 and 4.9 nm for the trans- and cis-
isomers, respectively. Because of the higher red-shift for the
trans-isomer, the overall difference between the excitation band
maxima of both configurations increases from 13.6 nm (1) to
23.6 nm (2). This increased band separation allows for the
generation of PSS with higher contents of cis-isomers by
irradiation of the trans-2 n → π* excitation band as compared
to that of trans-1.
To get a more detailed understanding of the effects of the
ortho-chloro substituents, we studied mono-, di-, tri-, and tetra-
chlorinated azobenzenes (Figure 3 and Table 2). Substituting
B
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concomitant effects on the π* molecular orbital (MO), para-
chloro azobenzene 5 should behave in a fashion similar to that
of the ortho-substituted derivative 4. The n → π* vertical
excitation energies of di-para-chloro azobenzene (5), however,
are comparable to those of azobenzene (1). Therefore, the
excitation wavelength shifts are an effect of the substitution at
the ortho-position. Because the ortho-chlorine atoms of trans-4
are facing toward the nitrogen lone pairs, it is reasonable to
assume that their repulsive interaction destabilizes the n
molecular orbital and thus decreases the energy gap to the π*
molecular orbital.18 In contrast, the bent structure of cis-4
allows the chlorine atoms to face in the opposite directions of
the nitrogen lone pairs. We reason that the strong blue-shift of
the n → π* excitation band that is observed by transitioning
from cis-1 to cis-4 results from a withdrawal of electron density
through the electronegative chlorine substituents in proximity
to the N−N bond. This effect has previously been reported for
tetra-ortho-fluoro azobenzenes and leads to a lowering of the n-
orbital energy.19,36
In contrast to 4, the introduction of the second ortho-
chlorine on the same aryl ring (6) twists the chlorinated aryl
group of the trans-isomer out of the plane at a ΦS angle of
53.6° and leads to a hypsochromic shift of the n → π*
excitation energy. On the basis of our prior findings, we
hypothesize that the hypsochromic shift is a result of a
decreased interaction between the chlorine substituent and the
nitrogen lone pairs due to the rotation of the aryl ring.
Changing the substitution pattern from cis-4 to cis-6 only
results in minor changes in the ΦS(avg) angle (63.0 nm for cis-
4 and 62.4 nm for cis-6), which is reflected by the subtle n →
π* band shift from 438.6 to 444.5 nm. It is worth noting that
the rotation around the Φ dihedral angle for trans-6 is
accompanied by a contraction of the N−N bond length from
1.243 to 1.239 Å, which is retained in the more chlorinated
derivatives 7. Adding a third ortho-chlorine onto 6 induces a
red-shift of the n → π* excitation energy of the trans-7 and cis-
7 of 19.2 and 9.4 nm, respectively. Finalizing the transition to
tetra-ortho-chloro azobenzene (2) from 7 strongly increases
the rotational overall twist of the trans structure to 50.8° for
both ΦS angles. This distortion leads to a further contraction of
the N−N bond length from 1.239 Å (trans-7) to 1.236 Å
(trans-2). In addition, a strong contraction of the Ψ dihedral
angle to 175.3° occurs, which ranges between 178.1° and
180.0° in the trans-configuration of less chlorinated substrates
(1, 3−7). The structural and conformational changes were
accompanied by an n → π* hypsochromic shift of 4.3 nm. In
contrast to trans-2, cis-2 is subject to a strong n → π*
bathochromic shift of 21 nm. We reasoned that the increasing
substitution of the ortho-position decreases the rotational
flexibility of the individual aryl rings around the Φ dihedral
angle. As a consequence, the chlorine atoms of cis-2 are forced
into the proximity of the nitrogen lone pairs, which destabilizes
Figure 3. Mono-, di-, and trichlorinated azobenzenes.
Table 2. Mono-, Di-, Tri-, and Tetrachlorinated
Azobenzenes: Vertical Excitation Energies (TD-PBE0/def2-
TZVP) and Photoswitch Structures (PBE0-D3/def2-TZVP)
NN
ΦS(Ar1)
ΦS(Ar2)
S0 → S1
config.
[Å]
[deg]
[deg]
Ψ [deg]
[nm]
1
3
4
5
6
7
2
trans
cis
trans
cis
trans
cis
trans
cis
trans
cis
trans
cis
trans
cis
1.242
1.234
1.242
1.232
1.243
1.231
1.243
1.235
1.239
1.230
1.239
1.230
1.236
1.229
0.0
52.0
0.2
51.7
0.7
63.0
0.1
50.5
0.6
50.4
10.6
51.7
50.8
60.5
0.0
52.0
1.4
63.0
2.7
180.0
8.5
179.9
9.5
179.8
10.5
180.0
9.0
178.1
6.9
483.6
470.0
501.9
454.3
520.6
438.6
480.0
478.7
483.6
444.5
502.8
453.9
498.5
474.9
63.0
0.1
50.5
53.6
74.3
49.8
71.8
50.8
60.5
179.1
7.8
175.3
5.1
one ortho-hydrogen on 1 for chlorine (3) already leads to a
significant separation of the n → π* transition band of the
trans- and cis-isomers. As compared to 1, trans-3 is red-shifted
by 18.3 nm, and cis-3 is blue-shifted by 15.7 nm. Because both
structures trans-1 and trans-3 remain planar, the bathochromic
shift could be attributed to the electronic contribution of the
chlorination. The chlorinated aryl ring of cis-3 has an increased
ΦS angle by 11.0°. Because of the asymmetric structure, both
aryl rings are rotated at different Φ angles. In this case, the
average of both smaller ΦS angles, ΦS(avg), can be used as a
descriptor for the overall rotation. For example, changing the
substitution pattern from cis-1 to cis-3 results in a difference of
5.4° in the ΦS(avg) (52.0 nm for cis-1 and 57.4 nm for cis-3).
Appending a second ortho-chlorine at the opposite aryl ring
follows the same trend and provides absorption maxima at
520.6 nm (trans-4) and 438.6 nm (cis-4). To evaluate the
origin of the strong shifts in the n → π* excitation wavelengths
through ortho-chloro substitution, we investigated the photo-
physical properties of para-chloro azobenzene (5). If the
bathochromic shift is a result of the extended π-system and the
Table 3. Optimized Conformation of 2 Imposed on the Structure of 1 (A,B) and Conformation of 1 Imposed on 2 (C,D):
Vertical Excitation Energies (TD-PBE0/def2-TZVP) and Photoswitch Structures (PBE0-D3/def2-TZVP)
entry
comp.
NN [Å]
ΦS [deg]
Ψ [deg]
S0 → S1 [nm]
A
B
C
D
trans-1 (trans-2-adopted)
cis-1 (cis-2-adopted)
trans-2 (trans-1-adopted)
cis-2 (cis-1-adopted)
1.236
1.229
1.242
1.234
50.8
60.5
0.0
175.3
5.1
180.0
8.5
443.7
471.4
711.3
463.1
52.0
C
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Figure 4. Computed orbital energies and n-MO densities of the optimized 2 conformation (left) and the conformation of 1 imposed on 2 (right).
the n molecular orbital and decreases the energy gap between
the n → π* transition.
EXAMINING THE EFFECT OF THE
TETRA-ORTHO-CHLORO AZOBENZENE
CONFORMATION ON ITS EXCITATION ENERGY
■
The computational study of the stepwise ortho-chlorination of
azobenzene (1) to tetra-ortho-chloro azobenzene (2) high-
lights that the geometry of the respective photoswitch has a
strong influence on the n → π* excitation energies. To
determine whether the conformational changes of the same
photoswitch structures provide shifts in the excitation energies,
we calculated the vertical excitation energies of 1 adopting the
optimized conformations of 2 (Table 3, entries A and B) as
well as the vertical excitation energies of 2 adopting the
Figure 5. Influence of the C−C−N−N dihedral angle ΦS on the
optimized conformations of 1 (entries C and D). Because the
excitation energy: optimized structure (PBE0-D3/def2-TZVP) with
conformations of cis-1 and cis-2 are closely related, imposing
the corresponding vertical excitation energies (TD-PBE0/def2-SVP)
the cis-2 geometry onto cis-1 and vice versa did not provoke
any significant alterations of its photophysical properties. In
contrast, changing the 3D structure of azobenzene to the tilted
geometry of 2 leads to a blue-shift of 39.9 nm for n → π*
transitions. Imposing the planar trans-1 geometry onto trans-2
leads to an outstanding n → π* bathochromic shift of
212.8 nm. This result supports the theoretical hypothesis that
the photophysical properties of an azobenzene-based photo-
switch are determined by both the chemical composition and
the conformational geometry.20 It furthermore showcases the
profound effect a change in the conformation can have on the
n → π* excitation wavelength of tetra-ortho-chloro azoben-
zenes. By computing the MO energies and visualizing the MOs
of the optimized and the planar trans-2 conformations (Figure
4), we show that the strong n → π* bathochromic shift likely
originates from the increased interaction between the chlorine
and the nitrogen lone pair, which leads to a strong
destabilization of the n-MO by 0.60 eV (Figure 4 and Table
3). The planar geometry additionally stabilizes the π*-MO,
which decreases the gap of the n → π* transition.
To gain a more detailed understanding of the influence of
the azobenzene conformation, we evaluated each of the main
geometrical variables for 1 and 2 separately (Figure 5 and
Supporting Information, Chapter 2.8 and Figure S74).
Therefore, we simulated the contraction and elongation of
the N−N bond lengths (Figure S74A and B), the distortion of
the Ψ dihedral angles (Figure S74C), and the selective rotation
of one aryl ring around the Φ dihedral angles (Figures 5 and
S74E). In each case, elongating the optimized N−N bond
of the photoswitches 1 and 2.
lengths (Figure S74A and B) results in a bathochromic shift of
the n → π* excitation bands with the strongest shifts for trans-
tetra-ortho-chloro azobenzene (2). A distortion of the Ψ
dihedral angles (Figure S74C) by contracting the trans-1 and
trans-2 structures as well as bending the cis-1 and cis-2
structures evokes a bathochromic shift. Rotating one aryl ring
of trans-1 around the Φ dihedral angle (Figure 5) has minor
effects on the n → π* transition. The trans-2 n → π* excitation
energies, however, were subject to a strong red-shift when the
Φ angle approached 0° and 180° (Figure 5). In case of cis-1,
rotating one aryl ring (Figure S74E) around the CCNN axis
shows that approaching planarity with the N−N double bond
in either direction provides a red-shift of n → π* excitation
energies. We found that the steric bulk of the ortho-chlorine
atoms hinders the full rotation around the CCNN axis of cis-2
(Figure S74E and F). Slight rotations of the optimized cis-2
geometry have minor effects on the n → π* excitation energy,
whereas approaching the limit of the rotational axis in each
direction strongly decreases the excitation energy barrier.
In general, our quantum-chemical calculations have shown
that the n → π* vertical excitation energy of trans-2 is strongly
dependent on its conformation and that minor structural
changes, such as the elongation of the N−N bond length,
distortion of the Ψ dihedral angle, and rotations of the Φ
dihedral angles can lead to strong bathochromic shifts. To
compare our computational findings to experimental con-
formations of azobenzene (1) and tetra-ortho-chloro azoben-
D
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zene (2), we recrystallized trans-2 as well as cis-2 and analyzed
their structures using X-ray crystallography.
bands at the red end of the spectrum is affected by highly red-
shifted conformations. The low absorption intensity of the tail
regions is a result of the low abundance of the conformations
that are shifted into the red region. According to our
calculations, highly red-shifted conformations are characterized
by long N−N bonds, small ΦS angles, and/or distorted Ψ
angles. In contrast, the strong absorption intensity of the band
maxima is a result of the high abundance of conformations that
are closely related to the optimized structures. Irradiation at
the band tail thus allows for the photoswitching of red-shifted
conformations to the cis-isomer. On the basis of our
hypothesis, trans-2 continuously cycles through a variety of
3D structures, which include these red-shifted conformations.
Therefore, a PSS with a high cis-content can be reached with
red light by the gradual accumulation of the long-lived cis-
isomer, while the flexible trans-2 molecules continuously adopt
red-shifted conformations and are photoswitched.
X-RAY ANALYSIS OF TETRA-ORTHO-CHLORO
AZOBENZENE
To gain insight into the experimental 3D structures of tetra-
ortho-chloro azobenzenes, we investigated trans-2 and cis-2
using X-ray crystallography (Figure 6 and Table 4). In
■
Figure 6. X-ray structures of trans-2 and cis-2.
It is reasonable to assume that a large fraction of the highly
red-shifted conformations is characterized by small average ΦS
angles, due to the outstanding n → π* band shifts that result
from the interaction between the chlorine atoms and the
nitrogen lone pairs. Therefore, we envisage that modifying the
substitution pattern to allow for a more planar structure yields
a photoswitch that could be isomerized using red light within
the biooptical window. Substituting one ortho-chlorine atom
on each aryl ring of 2 for the smaller fluorine atoms, in this
context, may lead to smaller ΦS dihedral angles and thereby
enable a better overlap between the two retained chlorine
atoms and the nitrogen lone pairs. In addition, incorporation of
fluorine atoms leads to a stronger withdrawal of electron
density in proximity to the diazene unit. This effect is known to
produce a hypsochromic shift of the n → π* band of the cis-
isomer, which increases the separation of the excitation
energies between both isomers.19,36 Irradiation of the
individual bands close to the absorption maxima could
consequently result in a PSS with higher isomer levels using
shorter light pulses.
Table 4. X-ray Analysis of the Structural Flexibility
config.
NN [Å] ΦS(Ar1) [deg] ΦS(Ar2) [deg] Ψ [deg]
1
2
trans
cis
trans1
trans2
trans3
cis
1.249
1.251
1.182
1.245
1.248
1.251
9.7
51.6
35.2
51.6
53.1
57.0
9.7
51.6
79.7
65.4
59.3
63.6
180.0
7.7
180.0
178.1
176.3
3.5
accordance with our computational results, we found that
the steric bulk of the ortho-chlorine atoms twists the two aryl
rings of trans-2 out of the plane. Remarkably, the extended cell
of the crystal structure contains three conformations for trans-2
with strong variations between the conformations. For
example, the N−N bond length ranges from 1.182 to
1.248 Å, the ΦS dihedral angles reach from 35.2° to 79.7°,
and the Ψ dihedral angle is distorted from 180.0° to 176.3°.
This result is an essential cornerstone for the development of
new photoswitches, because it provides a new perspective on
tetra-ortho-substituted azobenzenes: the twisted trans-isomers
are not stiff structures but are highly flexible and subject to
molecular motions. In contrast, the trans-azobenzene (1) X-ray
structure contains one conformation, which is near-planar with
a N−N bond length of 1.249, an ideal nondistorted Ψ angle,
and a ΦS dihedral of 9.7°.37 The X-ray structures cis-138 and
cis-2 also contain a single conformation, which suggests that
these structures have a more restricted conformation as
compared to that of trans-2. Conformational NMR studies
by Brittain and co-workers,39 however, have shown that cis-
tetra-ortho-substituted azobenzenes are subject to a certain
degree of structural flexibility. Comparing the structures of the
cis-isomers shows that the relative changes in the conformation
are accurately portrayed by our quantum-chemical calculations.
For instance, the average distortion of the ΦS dihedral angle is
increased from 51.6° (cis-1) to 60.3° (cis-2), and the Ψ
dihedral angle is decreased from 7.7° (cis-1) to 3.5° (trans-2).
We hypothesize that the broad n → π* excitation bands and
the concomitant extended band tail that allow for the
photoswitching to PSS with a high cis-content with red light
are a result of the conformational flexibility of trans-2. We have
provided computational evidence that azobenzene-based
photoswitches are subject to strong changes in their excitation
energies through the slightest alterations in their conformation.
Consequentially, we deduce that the excitation of the n → π*
COMPUTATIONAL OPTIMIZATION OF THE
TETRA-ORTHO SUBSTITUTION PATTERN
■
A computational investigation of the substitution of two ortho-
chlorines of 2 for the more electronegative fluorine (8, Figure
7) shows that, in accordance with our hypothesis, the ΦS
dihedral angles decrease from 50.8° to 37.3°. In addition, the
N−N bond is lengthened from 1.236 to 1.243 Å, and the
distortion of the Ψ angle is nearly retained. The combination
of these structural changes induces a bathochromic shift of
9.1 nm by transitioning from trans-2 to trans-8 (Table 5).
Figure 7. Structures of di-ortho-fluoro di-ortho-chloro (dfdc, 8), tetra-
ortho-fluoro (9) azobenzene, as well as the dfdc derivatives 10 and 11.
E
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ethyl benzoate substituents (10), however, leads to a strong
bathochromic shift of 19.3 nm for trans-10 and a slight
hypsochromic shift of 2.1 nm of cis-10. Implementing an
electron-rich substitution pattern (11) on the dfdc azobenzene
backbone leads to an elongation of the N−N bond from
1.243 Å (trans-8) to 1.248 Å for trans-11, which is accom-
panied by a relaxation of the twisted geometry by decreasing
the ΦS dihedral angle from 37.3° to 27.0°, whereas no
significant alterations are observed for the cis-11 conformation.
For the combination of these structural properties with the
electron-donating effect of the acetamide substituents, we
observe a bathochromic shift of 15.6 nm for trans-11 and of
11.1 nm for cis-11.
Table 5. Exchange of Two Chlorine Atoms for Fluorine on
Tetra-ortho-chloro Azobenzene: Vertical Excitation
Energies (TD-PBE0/def2-TZVP) and Photoswitch
Structures (PBE0-D3/def2-TZVP)
NN
ΦS(Ar1)
ΦS(Ar2)
S0 → S1
[nm]
config.
[Å]
[deg]
[deg]
Ψ [deg]
2
trans
cis
trans
cis
trans
cis
trans
cis
trans
cis
trans
cis
1.236
1.229
1.243
1.232
1.241
1.231
1.246
1.233
1.242
1.233
1.248
1.234
50.8
60.5
37.3
58.6
37.6
56.4
27.7
59.5
40.5
60.9
27.0
57.1
50.8
60.5
37.3
58.6
42.8
66.8
27.7
59.5
40.5
60.9
27.0
57.1
175.3
5.1
175.7
8.4
175.5
6.8
176.7
8.9
175.1
6.6
176.7
9.8
498.5
474.9
507.6
452.7
508.0
452.4
505.5
440.5
526.9
450.6
523.2
463.8
8
8s
9
SYNTHESIS AND PHOTOPHYSICAL
CHARACTERIZATION OF DI-ORTHO-FLUORO
DI-ORTHO-CHLORO AZOBENZENE
■
10
11
To confirm our theoretical results and to evaluate whether the
designed hybrid combines the red-shift of the tetra-ortho-
chloro azobenzene trans-isomer with the blue-shift of the tetra-
ortho-fluoro azobenzene cis-isomer, we synthesized 8. In
addition, we were intrigued by the strong computed batho-
chromic shift that occurred by substituting trans-8 with two
ethyl benzoate substituents (trans-10) and synthesized 10 to
experimentally validate the effect of an electron-withdrawing
para-substituent on the photophysical properties of dfdc
azobenzenes. For these syntheses, we leveraged our previously
established C−H chlorination methodology.27 By optimizing
this methodology for the dichlorination of the di-ortho-fluoro
azobenzenes 12 and 13, we could increase its substrate scope
to mixed ortho-substituted azobenzenes and provide our
desired products 8 and 10 in ample quantities (Figure 8).
Comparing cis-2 to cis-8 showed a stronger distortion of the Ψ
angle and an increased N−N bond length from 1.229 to
1.232 Å, without any significant alteration of the ΦS dihedral
angle. Although these geometric adjustments have been shown
to contribute to a bathochromic shift, the electronic
contribution to the photophysical properties that stem from
the withdrawing effect of the fluorine atoms results in an
overall hypsochromic shift of 22.2 nm. To exclude the
possibility that the di-ortho-fluoro di-ortho-chloro (dfdc)
azobenzene preferentially adopts a conformation, where the
chlorines are arranged on the same face of the molecule (8s),
we computed the total energies of both structures. The
optimized geometries trans-8 and cis-8 are more stable by
4.0 kJ/mol than are trans-8s and cis-8s, respectively (Table 5).
To evaluate the differences between the newly designed tetra-
ortho-fluoro/tetra-ortho-chloro hybrid (8) and the literature-
known tetra-ortho-fluoro structure (9),19,36 we investigated 9
with the same quantum-chemical methodology. As expected,
transitioning from cis-8 to cis-9 does not significantly alter the
conformation, but the increased withdrawal of electron density
in proximity to the diazene unit leads to a hypsochromic shift
of 12.2 nm. In comparison to trans-8, trans-9 is approaching a
planar conformation. The ΦS angle decreased to 27.7°, the Ψ
angle increased to 176.7°, and the N−N double bond is
elongated to 1.246 Å. These combined structural changes lead
to a positioning of the n → π* excitation maxima (trans-9: λmax
= 505.5 nm) similar to that of trans-8. On the basis of our
previous findings, however, we hypothesize that the geometric
flexibility of trans-8 enables an overlap between the bulky
chlorine atoms with the nitrogen lone pairs through molecular
motions. This interaction strongly contributes to the extended
n → π* band tails and allows for photoswitching with red light.
The small fluorine atoms of trans-9, in comparison, are unable
to overlap with the n molecular orbitals to the same extent,
which leads to narrower n → π* absorption bands and
Figure 8. Synthesis of the tetra-ortho-hydrids 8 and 10 using a
palladium-catalyzed C−H chlorination.
In general, the positions of the π → π* and n → π*
excitation bands were determined through UV−vis analysis
using a 50 μM solution in DMSO. Because of the low
absorption of the n → π* transition, the visualization of the full
extent of the band tail required a measurement at higher
concentrations (500 μM solution in DMSO).40 After the
position of the excitation bands was determined, the extent of
the n → π* band tail was verified through photoswitching by
illuminating the compounds with light at the red end of the
visible spectrum (Figure 9). The light-dependent PSS were
then analyzed by NMR spectroscopy using a 500 μM solution
in DMSO-d6 to allow for a direct comparison with the UV−vis
spectra. For example, the n → π* of trans-azobenzene (trans-
1) absorbs light up to 550 nm (Figure 9c). This was
experimentally confirmed by irradiation of the dark-adapted
state with green/yellow light (see the Supporting Information).
photoswitching with wavelengths close to the λmax
.
To account for the influence of electronically active
substituents on the conformation and photophysical properties
of the dfdc azobenzene, we computationally investigated the
addition of ethyl benzoate (10) and acetamide (11) groups to
its para-positions. Transitioning from trans-8 to trans-10 and
from cis-8 to cis-10 does not show any significant geometrical
changes, respectively. The electron-withdrawing effect of the
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Figure 9. (a) Photoswitching of tetra-ortho-chloro azobenzene (2).
UV−vis spectra of 1 and 2: (b) 50 μM in DMSO is used to show the
full spectrum and (c) 500 μM in DMSO is used to visualize the extent
of the n → π* band tails.
Illumination with 590 nm does not effect a change in the UV−
vis spectrum, whereas 530 nm light produces an altered PSS of
the molecule (1). However, due to the substantial overlap of
the n → π* excitation bands of the trans- and cis-isomers, the
application of 530 nm light only leads to minor photo-
conversion in comparison to irradiation of the π → π* band
maximum with UV-A light. UV−vis analysis of tetra-ortho-
chloro azobenzene (2, Figure 9) shows slightly separated n →
π* band tails with a more red-shifted absorption spectrum for
the dark-adapted state (100% trans-isomer). This slight
separation of the n → π* band tails allows one to reach
significant photoconversion by irradiating with green light
(530 nm, trans:cis = 40:60). The extent of the tail was
determined by applying 660 nm red light with an intensity of
24.1 mW/cm2 to the blue-adapted (410 nm) compound,
which produces a trans:cis ratio of 40:60 after 60 min. Both
azobenzene (1) and tetra-ortho-chloro azobenzene (2) have
long-lived cis-isomers with thermal half-lives of 75.5 and 20.7 h
in DMSO at 25 °C, respectively.
Remarkably, by exchanging two chlorine atoms of 2 for
fluorine (8), the separation between the n → π* excitation
bands of the trans-isomer and cis-isomer of 8 strongly
improved, which allows for the generation of near-quantitative
PSS (Figure 10). Irradiation with green light (530 nm), for
example, leads to an excellent PSS with significantly increased
cis-isomer content (trans:cis = 14:86) as compared to 2
(530 nm, trans:cis = 40:60). In addition, high levels of trans-8
(trans:cis = 90:10) could be produced with blue light
(410 nm). The extent of the n → π* band tail was determined
by applying 660 nm light (24.1 mW/cm2) to the blue-adapted
compound, which produces a trans:cis ratio of 41:59 after
60 min, 20:80 after 120 min, and even reaches 3:97 after
300 min. By employing a deeply red LED with higher intensity
(660 nm, 32.3 mW/cm2) under the same setting, photo-
switching could be accelerated to provide trans:cis ratios of
Figure 10. (a) Photoswitching of di-ortho-fluoro di-ortho-chloro
azobenzene (8). UV−vis spectra of 8: (b) 50 μM in DMSO is used to
show the full spectrum; (c) 500 μM in DMSO and 9:1 DMSO:H2O
are used to visualize the extent of the n → π* band tails; and (d) X-
ray structure of trans-8x and its calculated vertical excitation energy
(TD-PBE0/def2-TZVP).
30:70 and 10:90 after 60 and 120 min, respectively. To further
increase the rate of photoswitching, a three-membered LED
array could be used (660 nm; 24.1, 32.3, and 44.3 mW/cm2),
which leads to a trans:cis ratio of 24:76 after 60 min, 13:87
after 90 min, and 5:95 after 120 min. Next, we investigated the
thermal half-lives of 8 and found that it is near-bistable. Full
relaxation of green-irradiated 8 (530 nm) to its trans-
configuration requires heating to 90 °C for 7 h (t1/2 = 2 h,
DMSO). The thermal half-life at 70 °C was determined to be
16 h in DMSO. X-ray analysis of trans-8 provides the
conformation trans-8x where both aryl rings are oriented on
parallel, slightly displaced planes. In comparison to trans-2,
trans-8x has a decreased ΦS angle (ΦS = 35.9°), a diminished
distortion of the Ψ angle (Ψ = 180°), and an elongated N−N
bond length (N−N = 1.254 Å), which is in accordance with
our calculations. By subjecting trans-8x to our quantum-
chemical methodology, we determined that the n → π*
absorption maximum of the X-ray conformation lies at
478.3 nm, which is a hypsochromic shift of 29.3 nm as
compared to trans-8. On the basis of this result, it is likely that
the highly red-shifted conformations are adopted in the process
of transitioning between the twisted trans-8 and trans-8x,
where both aryl rings are oriented on parallel, slightly displaced
planes.
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We assessed the utility of the dfdc azobenzene as a
photoswitch for the development of photopharmaceuticals by
analyzing its photophysical properties in an aqueous solution
(9:1 DMSO:H2O). Overall, we did not observe any significant
changes in the PSS and thermal stability by transitioning from
DMSO to a mixture of DMSO and water. The PSS that is
reached with green light (530 nm, trans:cis = 14:86) remains
unchanged, and irradiation with blue light (410 nm, trans:cis =
85:15) only decreases the content of the trans-isomer by a
small fraction. Prolonged irradiation (60 min) of the blue-
adapted photoswitch (410 nm) with deep red light (660 nm,
24.1 mW/cm2, trans:cis = 40:60) produces a PSS similar to
that from irradiation in DMSO. The thermal half-life of cis-8 at
70 °C is 12 h in 9:1 DMSO:H2O.
To confirm that the dfdc azobenzene (8) is subject to a
stronger bathochromic shift than are both of its parent
azobenzenes, we examined the extent of the n → π* excitation
band of tetra-ortho-fluoro azobenzene (9). UV−vis analysis
shows that the n → π* band tail of 9 does not exceed 590 nm
(see the Supporting Information for experimental details),
which was subsequently confirmed by NMR analysis.
Irradiating the blue-adapted tetra-ortho-fluoro azobenzene
(420 nm, trans:cis = 83:17) with yellow light (595 nm,
10.1 mW/cm2) for 60 min provided a slight change in the PSS
(trans:cis = 76:14). However, subjecting 9 to a 60 min pulse of
red light (625 nm, 26.7 mW/cm2) or deep red light (660 nm,
24.1 mW/cm2) did not provide a significant alteration of the
trans:cis ratio.
visible spectrum and a stronger overlap in the green region.
Therefore, green light (530 nm) produces a trans:cis ratio of
41:59, whereas photoswitching with 410 and 440 nm blue light
results in outstanding PSS with trans-isomer contents of 96%
and 95%, respectively. This effect is further demonstrated by
irradiating the 410 nm-adapted 10 with deep red light
(660 nm, 24.1 mW/cm2), which shows accelerated photo-
switching kinetics as compared to 8 and produces a trans:cis
ratio of 17:83 after 60 min. The only drawback of the electron-
poor 10 as compared to the unsubstituted dfdc azobenzene is
the diminished thermal stability of the cis-isomer, which has a
half-life of 5 h at 25 °C in DMSO.
SIGNIFICANCE OF THE DI-ORTHO-FLUORO
DI-ORTHO-CHLORO AZOBENZENE
■
Azobenzenes are the most commonly used photoswitches to
develop advanced optical tools such as photopharmaceuticals,
due to their small size, robust photoswitching, and low rate of
photobleaching.1−5 To fine-tune the photophysical properties
of these optical devices, it is highly desirable to have access to
an arsenal of azobenzene core structures with defined
photophysical properties. In this regard, the dfdc azobenzene
(8) provides an unprecedented solution to the long-standing
problem to identify a substitution pattern that combines near-
bistable isomers, PSS with high levels of trans- and cis-isomers,
and photoswitching within the biooptical window. Although
photoswitching with deep red light (660 nm) requires long
irradiation times (trans:cis = 41:59 after 60 min with
24.1 mW/cm2), we show that by increasing the light intensity
(trans:cis = 30:70 after 60 min with 32.3 mW/cm2) and/or
using arrays of multiple LEDs (trans:cis = 24:76 after 60 min
with 24.1, 32.3, and 44.3 mW/cm2), the photoswitching rate
can be significantly accelerated. In addition, the installation of
electron-poor substituents on the 8 para-position (10) is
accompanied by a bathochromic shift, which allows for faster
isomerization with deep red light. Given that photodynamic
therapy is carried out with light sources that provide an
intensity of up to 200 mW/cm2 which can, in theory, be
incorporated into an array, we reason that the dfdc azobenzene
could be photoswitched within a photopharmaceutically
relevant time frame in a therapeutic setting.41 The decreased
steric bulk of the ortho-substitution of 8 as compared to 2
allows the relaxation of the twisted structure to a near-planar
geometry, which leads to a closer 3D relationship of dfdc
azobenzene to azobenzene. This factor plays a crucial role in
the process of red-shifting photopharmaceuticals.42 The ability
to add a para-substituent to 8 increases the utility of this
pattern by allowing its anchoring to optical devices through an
electron-active substituent (10). This substitution pattern
leads to a shift of the λmax(n → π*) to the red end of the
spectrum, increases the trans-content of the blue-irradiated
photoswitch, and only decreases the thermal stability of the cis-
isomer to an extent that 10 is still considered a slow-relaxing
azobenzene. These excellent photophysical properties that
were demonstrated by evaluating two dfdc derivatives leave a
great margin for improvement with regard to the substitution
of the meta- and para-positions. Therefore, 8 serves an ideal
basis for the development of a photoswitch that operates with
near-infrared light while retaining highly bistable isomers by
performing an in-depth investigation into the effects of
alternative substitution patterns on its photophysical proper-
ties.
Next, we examined the effect of electron-withdrawing para-
substituents on the photophysical properties of the dfdc
azobenzene by implementing ethyl benzoate groups (10,
Figure 11). As compared to trans-8 and cis-8, the λmax(n → π*)
values of both isomers, trans-10 and cis-10, are subject to a
bathochromic shift (Figure 11c). This provides an excellent
separation of the n → π* excitation bands between the trans-
and cis-isomers in the blue as well as orange/red regions of the
Figure 11. (a) Photoswitching of the electron-poor di-ortho-fluoro di-
ortho-chloro azobenzene 10. UV−vis spectra of 8 and 10: (b) 50 μM
in DMSO is used to show the full spectrum; and (c) 500 μM in
DMSO is used to visualize the extent of the n → π* band tails.
H
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Lars Allmendinger − Department of Pharmacy, Ludwig-
Maximilians-University Munich, Munich 81377, Germany
Dirk Trauner − Department of Chemistry, Ludwig-Maximilians-
University Munich, Munich 81377, Germany; Department of
Chemistry, New York University, New York, New York 10003,
United States; orcid.org/0000-0002-6782-6056
CONCLUSION AND OUTLOOK
■
We have used a combination of quantum-chemical calculations
and X-ray analyses to dissect the effects of the ortho-
chlorination in its electronic and structural subcomponents
and identified the origin of the extended red-shifted n → π*
band tails of tetra-ortho-chloro azobenzenes. On the basis of
our findings, we designed the dfdc azobenzene substitution
pattern and synthesized 8 and 10 by employing a C−H
dichlorination methodology. UV−vis and NMR studies
showed significant improvements of the photophysical proper-
ties of 8 over the tetra-ortho-chloro and tetra-ortho-fluoro
substitution pattern. Photoswitching can be achieved with red
light within the biooptical window and provides a PSS with up
to 97% of the highly stable cis-isomer. These outstanding
properties make the dfdc azobenzene an ideal photoswitch for
the development of photopharmaceuticals that target complex
animal tissues. In addition, the dfdc azobenzene scaffold is an
excellent basis for the development of bistable photoswitches
that can be isomerized with near-infrared light. Implementing
an electron-poor substitution pattern by adding an ethyl
benzoate group to both para-positions (10), for example, leads
to a significant bathochromic shift of the n → π* excitation
bands of both isomers, which results in an accelerated
photoconversion by applying deep red light as compared to
8. To evaluate the full potential of the dfdc azobenzene, studies
on the effect of meta- and para-substituents are currently
underway.
Complete contact information is available at:
https://pubs.acs.org/10.1021/jacs.9b10430
Author Contributions
∇D.B.K. and G.S. contributed equally.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was supported by the SFB749 to D.T. and C.O. as
well as a Georg Forster Research Fellowship (Alexander von
Humboldt Foundation) to A.M.A. C.O. also acknowledges
financial support by the Innovative Training Network
“Computational Spectroscopy in Natural Sciences and
Engineering” (ITN-COSINE). D.B.K. received funding from
the European Union’s Framework Program for Research and
Innovation Horizon 2020 (2014-2020) under the Marie
Skłodowska-Curie Grant agreement no. 754388 (LMURe-
searchFellows) and from LMU Munich’s Institutional Strategy
LMUexcellent within the framework of the German Excellence
Initiative (no. ZUK22). A.M.A. is grateful to Prof. Dr. Klaus
Wanner for hosting the second year of his AvH fellowship. We
thank Prof. Ivan Huc for hosting revision experiments in his
laboratory, Dr. Oliver Thorn-Seshold and Li Gao for their help
with establishing a LED setup, Dr. Peter Mayer for X-ray
analyses, as well as Marvin Thielert for helpful discussions.
ASSOCIATED CONTENT
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/jacs.9b10430.
■
sı
Experimental procedures, spectroscopic data, and copies
of NMR spectra (PDF)
X-ray crystallographic data for compounds trans-2
(CCDC 1954066), cis-2 (CCDC 1954067), and trans-
8x (CCDC 1954065) (CIF)
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