Biomimetic hydrogen-bonding cascade for chemical activation: telling a nucleophile from a base

Article abstract of DOI:10.1039/d0sc05067a

Hydrogen bonding-assisted polarization is an effective strategy to promote bond-making and bond-breaking chemical reactions. Taking inspiration from the catalytic triad of serine protease active sites, we have devised a conformationally well-defined benzimidazole platform that can be systematically functionalized to install multiple hydrogen bonding donor (HBD) and acceptor (HBA) pairs in a serial fashion. We found that an increasing number of interdependent and mutually reinforcing HBD-HBA contacts facilitate the bond-forming reaction of a fluorescence-quenching aldehyde group with the cyanide ion, while suppressing the undesired Br?nsted acid-base reaction. The most advanced system, evolved through iterative rule-finding studies, reacts rapidly and selectively with CN- to produce a large (>180-fold) enhancement in the fluorescence intensity at λmax = 450 nm.

Full text of DOI:10.1039/d0sc05067a

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DOI: 10.1039/D0SC05067A
ARTICLE
Biomimetic hydrogen-bonding cascade for chemical activation:
telling a nucleophile from a base
Hyunchang Park and Dongwhan Lee*
Received 00th January 20xx,
Accepted 00th January 20xx
Hydrogen bonding-assisted polarization is an effective strategy to promote bond-making and bond-breaking
chemical reactions. Taking inspirations from the catalytic triad of serine protease active sites, we have devised a
conformationally well-defined benzimidazole platform that can be systematically functionalized to install multiple
hydrogen bonding donor (HBD) and acceptor (HBA) pairs in a serial fashion. We found that an increasing number
of interdependent and mutually reinforcing HBD–HBA contacts facilitate the bond-forming reaction of fluorescence-
quenching aldehyde group with cyanide ion, while suppressing the undesired Brønsted acid–base reaction. The
most advanced system, evolved through iterative rule-finding studies, reacts rapidly and selectively with CN– to
produce a large (> 180-fold) enhancement in the fluorescence intensity at λmax = 450 nm.
DOI: 10.1039/x0xx00000x
In such cooperative arrangement of paired HBD–HBA units,
the induced polarization at the mediator site renders the
hydrogen bonds shorter and stronger, when a comparison is
made to a single HBD–HBA contact.19 A prominent example of
the cascade hydrogen bonding in action is a class of enzymes
having catalytic triad, such as serine protease, canonical
esterase, and lactonase.23–27 At the active site of these enzymes,
the carboxylate group of aspartate, imidazole group of histidine,
and hydroxyl group of serine constitute an extended hydrogen
bonding array (Fig. 2a).
Introduction
Hydrogen bonding is a versatile functional motif for chemical
structure design. An elaborate arrangement of hydrogen
bonding donor (HBD) and acceptor (HBA) units have been
exploited for molecular recognition,1–5 signaling,6–8 self-
assembly,9–12 and chemical activation.13–15 Beyond the
paradigm of simple HBD–HBA pair (Fig. 1a), a serial array of
multiple HBD–HBA pairs can also be constructed in a cascade
fashion (Fig. 1b).16–18 With the amphoteric proton mediator in
the middle, electronic polarization at one end automatically
increase the donor/acceptor ability of the other end of the
extended network.19–23
Fig. 1 Schematic representation of (a) simple HBD–HBA pair, and (b) cascade
hydrogen bonding with an amphoteric proton mediator, such as imidazole,
between widely separated HBD and HBA to create a larger additive dipole to
promote the addition/substitution reaction shown in (c).
Fig. 2 (a) Chemical structure and X-ray structure of the active site of serine
protease (PDB: 1HXE). (b) Chemical structure and DFT (B3LYP-D3/6-31G(d,p))
energy-minimized structure of a synthetic “triad” 1.
Department of Chemistry, Seoul National University, 1 Gwanak-ro, Gwanak-gu,
Seoul 08826, Korea. E-mail: dongwhan@snu.ac.kr
† Electronic Supplementary Information (ESI) available: Experimental procedures,
characterization data and supplementary figures, including 1H and 13C NMR
spectra,
crystallographic
data,
FT-IR
and
HR-MS
analysis.
See
DOI: 10.1039/x0xx00000x
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Journal Name
In the biological system, the amphoteric imidazole group
functions as a mediator of the cascade hydrogen bonding (Fig. fluorophore
1b). The hydrogen bonding between aspartate and histidine of toxic cyanide ion. From the simplest molecular prototype
In this paper, we report the chemistry of a latent
View Article Online
(Fig. 2 and 3) and its applicaDtOioI:n10f.o10r3t9h/eD0dSeCt0e5c0t6io7An
1
2,
polarizes the amine N–H group to enhance the basicity of the a systematic increase in the number of HBD–HBA units helps
imine nitrogen, which translates to a stronger N···H–O hydrogen polarize the aldehyde carbonyl group (Fig. 3b). The enhanced
bond between histidine and serine. An unusually downfield- electrophilicity of the aldehyde moiety of 1 effectively promotes
shifted proton resonance of the imidazole N–H proton at ca. 15 selective covalent capture of cyanide ion (Fig. 3c), and allows
ppm reflects tight hydrogen bonding, which polarizes the serine rapid fluorescence turn-on detection. More importantly,
O–H group to enhance its nucleophilicity toward electrophilic comparative studies on
1 and its simpler “site models” 2–4
substrates.23 For the enzymes, it would be challenging to control established that multiple interconnected HBD–HBA pairs help
the strength of hydrogen bonds by using limited types of distinguish nucleophile from Brønsted base.
functional groups offered by amino acid residues. As such, the
construction of hydrogen bonding array, in which the whole is
more than the sum of its parts, is an effective strategy to make
the best out of what is naturally available.
Results and Discussion
Synthesis of a Minimalist Prototype
A
single HBD–HBA pair was first installed onto the
benzimidazole platform to prepare (Scheme 1). To maximize
2
Design Principles
Taking inspirations from the chemical activation strategy of
such catalytic triad, we have designed a benzimidazole-based
the effects of chemical transformation on the emissive
properties, it would be ideal if the functional group that is
activated by hydrogen bonding (Fig. 1c) functions also as a
fluorescence quencher. For such purpose, an aldehyde group
was employed as an HBA unit. Through fast intersystem crossing,
an aldehyde group facilitates the quenching of π-conjugated
fluorophore to which it is directly attached.30–33
synthetic surrogate
1 (Fig. 2b). In the case of enzymes, the
three-dimensional polypeptide scaffold enables the precise
positioning and alignment of HBD–HBA units. Instead of using
peptides, we employed the rigid π-conjugated backbone of
benzimidazole as a minimalist artificial platform to build
biomimetic hydrogen bonding network. Through facile
functionalization at the 2-, 4-, and 7-position of benzimidazole
ring (Fig. 3), various HBD and HBA units can be installed around
the imidazole core. Additionally, the rich photophysics of 2-
arylbenzimidazole motif offers excellent opportunities to
develop these biomimetic small molecules into fluorescent
probes.28,29
The chemical structure of
requirement of this design concept. As outlined in Scheme 1,
the synthesis of involved oxidative condensation of diamine
2 (Fig. 3b) satisfies this minimum
2
5
and aldehyde, and subsequent oxidation of the primary alcohol
group. Except for the reduction of the thiadiazole precursor, all
reactions proceeded in moderate to good yield (> 75%). The
poor isolation yield of the intermediate
5 might be due to the
instability of the electron-rich diamine functionality. The
synthetic modularity in the imidazole ring construction aided
structural diversification (vide infra).
Spectroscopic Studies and Response to Cyanide Ion
Compound
UV–vis absorption and fluorescence emission spectra. As shown
in Fig. 4a, the addition of cyanide to a solution of in DMSO led
to a decrease in the absorption at λmax = 345 nm with the
development of a new band at λmax = 390 nm. While is only
2 responds to cyanide anion by large changes in both
2
2
weakly emissive due to the efficient fluorescence quenching by
the aldehyde group, the intrinsic blue emission of
benzimidazole (λmax,em = 465 nm) was restored upon addition of
cyanide (Fig. 4b).
Fig. 3 (a) Cascade hydrogen bonds built on the 4,7-disubstituted benzimidazole
platform. (b) Structural “evolution” from the simplest 2 to the most elaborate 1
by installing an increasing number of HBD and HBA units. (c) Mechanism of
covalent capture and fluorescence turn-on response of 1 toward CN–.
Scheme 1 Synthetic route to a simple HBD–HBA prototype.
2 | J. Name., 2012, 00, 1-3
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In stark contrast, the H NMR spectrum of a solVuietwioAnrticolefO2nlinine
1
DOI: 10.1039/D0SC05067A
CDCl3 (Fig. S2) shows a well-resolved spectral pattern that is
consistent with the chemical structure of 2, thus ruling out the
involvement of impurities in the complicated 1H NMR spectrum
obtained in DMSO-d6.
We suspect that the intramolecular N–H···O bond of
2 does
not provide sufficient thermodynamic bias toward one
tautomer over the other in DMSO-d6 (Fig. 5b and Scheme 2).
Indeed, the two distinct resonances of the benzimidazole N–H
protons at δ = 13.31 and 13.04 ppm converge into a single
averaged feature at δ = 12.93 ppm as the temperature is
increased (Fig. 5b, coalescence at Tc = 343 K). From eq (1), the
corresponding rate constant (kc) can be determined by the ∆ν
value (= 108 Hz) at the slow exchange limit.34 By applying the
Eyring equation in eq (2), the activation energy of ∆G‡ = 16.4
kcal mol-1 was determined, which is typical for the energy
barrier for the tautomerization.35,36 In this process, the
resonances of the aldehyde protons at δ = 10.81 and 10.34 ppm,
each associated with the two tautomers, also converge into a
single resonance at δ =10.54 ppm.
Fig. 4 (a) Absorption and (b) emission spectra (λexc = 400 nm) of 2 (0.100 mM) prior
to (black dashed lines), and after treatment with NaCN (25 equiv, blue lines) or
Et3N (25 equiv, gray lines) in DMSO at T = 298 K.
In the control experiments with other chemical species,
however, we realized that a large enhancement in the emission
intensity also occurs with the addition of Brønsted base, such as
Et3N (Fig. 4b). The excitation spectrum of 2, obtained in the
presence of Et3N, revealed that absorption at λ ≈ 400 nm is
responsible for the emission at λmax,em = 465 nm (Fig. S1a). This
finding implicates that the loss of hydrogen bonding by
deprotonation of the acidic benzimidazole proton could also
contribute to the fluorescence enhancement, presumably by
suppressing the intersystem crossing of aldehyde as the main
quenching pathway. In other words, both addition and
ꢂ
|
∆ꢃ
|
ꢀꢁ =
(1)
(2)
2
√
deprotonation reactions could take place in the reaction of
2
ꢀꢌꢇꢈ
ꢀꢈℎ
with cyanide, but the fluorescence response is essentially
indistinguishable.
ꢄꢅ‡ = ꢆꢇꢈ ꢉꢊ ꢋ
ꢍ
To delineate the nature of the chemical reaction that is
responsible for the fluorescence enhancement (Fig. 4b), we
The addition of cyanide to 2 in DMSO-d6 produced multiple
1
carried out detailed solution H NMR spectroscopic studies. At
products, presumably from the reactions involving nucleophilic
attack as well as simple deprotonation, as deduced from the 1H
NMR spectrum (Fig. 5a, bottom). We tentatively concluded that
1
room temperature (T = 298 K), the H NMR spectrum of
2 in
DMSO-d6 (Fig. 5a, top) shows a pair of broadened resonances
associated with benzimidazole N–H and aldehyde C–H protons.
the weak and solvent-exposed hydrogen bond of
2 is prone to
tautomerization, and subjected to multiple reaction pathways
with cyanide functioning as either nucleophilic Lewis base or
simple Brønsted base.
Structural Evolution of Hydrogen Bonding Network
To suppress the undesired acid–base chemistry observed for
2
(Fig. 4 and 5), additional functional groups were introduced to
construct tighter hydrogen bonding. We postulated that a
stronger bond polarization (Fig. 1) of cascade hydrogen bonding
should shift the tautomer equilibrium to make the hydrogen-
bonded form dominant even in polar solvent environment
(Scheme 3, bottom), thereby enhancing the electrophilicity of
the aldehyde group toward covalent capture of cyanide anion.
As summarized in Scheme 3, probes
prepared from the common diamine intermediate
3
and
4
were readily
(Scheme 1).
5
Fig. 5 (a) 1H NMR spectra of 2 (4.0 mM) prior to (top) and after (bottom) addition
of NaCN (10 equiv) in DMSO-d6; T = 298 K. (b) Variable-temperature (VT) 1H NMR
spectra of 2 (4.0 mM) in DMSO-d6 measured at T = 293–363 K.
Scheme 2 Tautomerization of a simple HBD–HBA pair.
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Cascade Hydrogen Bonds: Effects on the Reactivity and Solution
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DOI: 10.1039/D0SC05067A
Structure
Both
upon treatment with cyanide anion (Fig. 7). Unlike
they remain silent toward Et3N, thus differentiating Brønsted
base from Lewis base. When comparison is made with , slightly
3
and
4
exhibited dramatic fluorescence enhancement
2, however,
2
blue-shifted but almost superimposable emission spectra
(λmax,em = 445 nm) were observed for the cyanide reaction
products of
3 and 4.
Scheme 3 Construction of hydrogen bonding network to shift the tautomer
equilibrium. (a) aldehyde, Na2S2O5, EtOH, H2O, Δ. (b) MnO2, MeCN/CH2Cl2, r.t.
We anticipated that multiple HBD–HBA pairs within
3
and 4
would shift the tautomer equilibrium to benefit from the
additive and reinforcing dipole alignment (Scheme 3, bottom).
The results from 1H NMR spectroscopic studies, however, were
rather inconclusive. As shown in Fig. 8, the coexistence of two
tautomers was still observed at r.t. for both
broadened proton resonances of resemble those of
top), the two tautomers of appear as sharp and well- resolved
For 3, a hydroxyl group is installed as HBD at the upper ring
of the benzimidazole core. Similarly to the catalytic triad (Fig.
2a), the O–H···N hydrogen bonding between the hydroxyl group
and the imine nitrogen atom of benzimidazole is expected to
push the tautomer equilibrium to reinforce the N–H···O
hydrogen bonding between the aldehyde and amine moieties
of the benzimidazole core. Furthermore, the acidic phenolic O–
H (pKa ~ 10)37 could also function as a sacrificial proton donor in
the acid-base reaction to keep the N–H group intact. Even when
the acid-base reaction takes place, the deprotonation reaction
should occur preferentially at the hydroxyl group, so that the
benzimidazole fluorophore would be less perturbed.
3
and
4
. While the
3
2
(Fig. 5a,
4
spectral patterns in ca. 1:1 ratio (Fig. 8a and Fig. 8b, top).
As a further structural elaboration, probe 4 has an additional
hydrogen bonding acceptor, –OMOM group. Here, the
hydrogen bonding between the ether oxygen atom of –OMOM
and the imidazole N–H group is anticipated to help planarize the
π-system and strengthen the bifurcated hydrogen bonding. It
makes intuitive chemical sense that the multiple HBD–HBA pairs
Fig. 7 Emission spectra (λexc = 343 nm) of (a) 3 (0.100 mM) prior to (black dashed
lines), and after treatment with NaCN (25 equiv, blue line) or Et3N (25 equiv, gray
line), and (b) 4 (0.100 mM) prior to (black dashed lines), and after treatment with
NaCN (25 equiv, blue line) or Et3N (25 equiv, gray line) in DMSO at T = 298 K.
within
4 should work cooperatively in the same direction (Fig.
1b) to preferentially stabilize the desired tautomer which
benefits from stronger hydrogen bonding (Scheme 3, bottom).
Indeed, single-crystal X-ray crystallography confirmed the
presence of an extensive hydrogen bonding network within
As shown in Fig. 6, the X-ray structure of revealed three
hydrogen bonds (dashed lines) around the imidazole core with
short Nimidazole···Ophenol (2.568(3) ) and Nimidazole···Oether (2.634(4)
) distances and essentially coplanar arrangement of the
4.
4
Å
Å
extended p-system (torsional angle for N1–C1–C9–C10 = 0.65˚;
C2–C3–C8–O4 = 1.92˚).
Fig. 6 X-ray structure of 4 as ORTEP diagrams with thermal ellipsoids at the 50%
probability level: (a) face-on and (b) edge-on views. Selected interatomic distances
(Å) of the hydrogen bonds (dashed lines): N1···O3, 2.568(3); N2···O1, 2.634(4);
N2···O4, 2.901(3).
Fig. 8 (a) 1H NMR spectra of 3 (4.0 mM) in DMSO-d6 prior to (top) and after
(bottom) addition of NaCN (10 equiv). (b) 1H NMR spectra of 4 (4.0 mM) in DMSO-
d6 prior to (top) and after (bottom) addition of NaCN (10 equiv). T = 298 K.
4 | J. Name., 2012, 00, 1-3
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Similar to the case of
when cyanide was added to
2
, mixtures of products were obtained furnished the target compound
1
. The low isolatVioienw AyrtiieclledOn(clinae.
DOI: 10.1039/D0SC05067A
3 (Fig. 8a, bottom), presumably 17%) in this final step is due to the purification procedure
reflecting the tautomer equilibrium. The sharp and well- involving repetitive recrystallization.
resolved 1H NMR spectrum of
suggests much slower The single-crystal X-ray structure of
4
1 shown in Fig. 9
interconversion between the two tautomers (Scheme 3), which confirmed the presence of multiple hydrogen bonds. The
is consistent with the more extensive hydrogen bonding array interatomic distances of hydrogen bonds, 2.5431(1)–2.9007(1)
in
4
than its simpler analogues
2
and
3
. Nevertheless, the Å, are similar to those of
4
(Fig. 6). The essentially co-planar
(torsional angle
coexistence of the two tautomers for
formation of multiple products upon reaction with CN- (Fig. 8b, for N1–C1–C9–C10 = 4.17°; C2–C3–C8–O4 = 2.03°) further
bottom). Apparently, the presence of three hydrogen bonds (Fig. validates the functional role of hydrogen bonding network as
4
still resulted in the arrangement of the π-conjugated backbone of
1
6) is still insufficient for a complete conversion of
product.
4
to a single conformational lock. Unlike the DFT computational model (Fig.
2b), the amide N–H group of is twisted away from the
1
benzimidazole-N atom. A close inspection of the crystal packing
diagram revealed extensive intermolecular Namide–H···Ocarbonyl
Biomimetic Hydrogen Bonding Network
As described above, studies on 2–4 suggested the need for hydrogen bonding network between adjacent molecules, which
stronger and more tightly regulated hydrogen bonding network is reinforced further by π–π stacking and C–H···π contacts (Fig.
to control both the tautomer equilibrium and the reactivity S5). Apparently, such intermolecular interactions in the
toward nucleophile. We thus decided to install an additional condensed phase prevail over the inherent propensity of
1 to
HBD–HBA motif onto to prepare (Fig. 3b). Within a six- make intramolecular Namide–H···Nimidazole hydrogen bond as
4
1
membered ring setting, the highly polarized N–H bond of an discrete molecular species (Fig 2b).
amide moiety is ideally suited to make a good HBD–HBA pair
Hydrogen Bonding Network: Effects on the Reactivity and Solution
Structure
with the imidazole imine-N atom, as predicted by the energy-
minimized DFT (B3LYP-D3/6-31G(d,p)) model shown in Fig. 2b.
In line with the schematic diagram shown in Fig. 1, comparative As shown in Fig. 10a, the addition of cyanide anion to a solution
DFT studies on and the simple HBD–HBA pair predicted a of elicited a rapid and dramatic (> 180-fold) enhancement in
large difference in the molecular dipole moment of 6.4413 D the emission intensity at λmax,em = 450 nm, whereas no spectral
(for ) vs 0.5614 D (for ). The molecular electrostatic potential change was observed with the Brønsted base Et3N. A large
(MEP) maps of and (Fig. S4) also predict a larger polarization spectral change was also observed in the electronic excitation
1
2
1
1
2
1
2
of electron density across cascading dipoles.
To functionalize the 4-position of the benzimidazole core,
the overall synthetic scheme needed to be modified to
introduce a bromo substituent at the early stage (Scheme 4).
From the bromo-functionalized diamine, sequential oxidative
condensation and oxidation reactions afforded
6 in high yield
(72% for two steps). By cross-coupling reaction onto this bromo
position, various functional groups could be installed. An
aliphatic amide group was chosen in our molecular design to
suppress direct electronic conjugation with the benzimidazole
core, thereby minimizing perturbation of the photophysical
properties. A palladium-catalyzed Suzuki-Miyaura cross-
Fig. 9 X-ray structure of 1 as ORTEP diagrams with thermal ellipsoids at the 50%
probability level: (a) face-on and (b) edge-on views. Selected interatomic distances
(Å) of the hydrogen bonds (dashed lines): N1···O3, 2.5431(1); N2···O1, 2.62995(9);
N2···O4, 2.9007(1).
coupling of
6 with α-(acetylamino)benzylboronic ester
Fig. 10 (a) Emission (λexc = 343 nm) and (b) absorption spectra of 1 (0.100 mM,
black dashed lines), and spectral changes induced by treatment with NaCN (25
equiv, blue lines), and with Et3N (25 equiv, gray lines) in DMSO at T = 298 K. Inset:
photographic images of (a) fluorescence response (under 365 nm UV lamp), and
(b) color change.
Scheme 4 Synthetic route to 1. (a) aldehyde, Na2S2O5, EtOH/H2O, Δ. (b) MnO2,
MeCN/CH2Cl2, r.t. (c) Pd2(dba)3, P(tBu)3·HBF4, KF, dioxane/H2O, Δ.
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upon the addition of cyanide (Fig. 10b) with color change to
Journal Name
1 was established
The high conformational stability of
View Article Online
1
yellow. In contrast, only a slight increase in the absorption at λ further by concentration-dependent HDNOMI: 1R0.1s0t3u9d/Die0sS.CW05i0t6h7iAn
≈ 400 nm region was observed with Et3N, with the rest of the the concentration range of 1.5–4.0 mM, no noticeable change
spectrum remaining essentially superimposable (Fig. 10b). This was observed in the 1H NMR spectrum of
1
in DMSO-d6,
remains folded in solution against
on allow naked-eye detection of cyanide anion (Fig. 10, inset intermolecular hydrogen bonding (Fig. S7). In stark contrast,
pictures) compound having relatively weak hydrogen bonds undergoes
To investigate the solution structure of
1H NMR significant broadening of benzimidazole aromatic proton
spectroscopic studies were carried out. The H NMR spectrum resonances with increasing sample concentration (Fig. S8).
of (4.0 mM) in DMSO-d6 measured at T = 293 K indicates the Furthermore, the addition of a small amount of H2O (2 μL) to a
dominance of one prevailing tautomer (> 90%, based on the solution of in DMSO-d6 (2.0 mM, 500 μL) sharpened the
visually discernible colorimetric change and fluorescence turn- implying that
1
3
1,
1
1
3
peak integration values; Fig. 11a, top), which remain essentially resonances of these aromatic protons by rapid proton exchange
invariant even with increasing temperature up to T = 363 K (Fig. with the N–H group (Fig. S9). No spectral change was observed
11b). The 2D ROESY NMR spectrum of
1
(4.0 mM) obtained in for
1
under the same condition (Fig. S10).
DMSO-d6 at r.t. revealed prominent ROE signals between (i)
With the hydrogen-bonded tautomeric form prevailing for
1
benzimidazole N–H and aldehyde C–H, and (ii) benzimidazole in solution, the addition of cyanide anion resulted in clean and
N–H and methylene protons of the –OMOM group (Fig. S6). complete conversion to the cyanohydrin adduct (Fig. 11a,
These ROE correlations provide compelling evidence for the bottom). To better interpret the 1H NMR spectrum of the
dominant tautomeric form of
computational studies (Fig. 2b). The gradual up-field shifts of shown in Fig. 12, the cross-peaks at
1
,
as predicted by DFT reaction product, we carried out 2D-COSY NMR studies. As
= 9.7 ppm and = 6.6
ꢎ
ꢎ
the phenolic O–H (δ = 13.46 to 13.30 ppm) and amide N–H ppm arise from the J-coupling of O–H and C–H protons,
protons (δ = 8.99 to 8.68 ppm) with increasing temperature (Fig. respectively, of the cyanohydrin.32,33,40–42 In addition, HPLC-MS
11b) also suggest their involvement in intramolecular hydrogen analysis (Fig. S11) confirmed the formation of the cyanohydrin
bonding.38,39
adduct of
Telling a Nucleophile from a Base
To compare the ability of in distinguishing nucleophile (i.e.
1
at m/z = 473.15 corresponding to [M + H]+.
1–4
CN–) from Brønsted base (i.e. Et3N), each of the probe molecules
was treated with either cyanide anion (25 equiv) or Et3N (25
equiv). The emission intensity was measured at λem = 450 nm,
and the ratio ICN/IEt3N was calculated for each molecule. With an
increasing number of HBD–HBA units installed around the same
fluorogenic benzimidazole core (Fig. 3b), a systematic increase
in the ICN/IEt3N ratio was observed along the series 2→3→4→1
(Fig. 13). Stronger and networked hydrogen bonds seem to
promote bond-forming reaction while effectively suppressing
the undesired acid–base reaction. Comparative 1H NMR studies
(Fig. 5, 8, and 11) establish that such hydrogen bonds can also
shift the solution equilibrium toward the more reactive
tautomer toward cyanide anion.
Fig. 11 (a) 1H NMR spectra of 1 (4.0 mM) prior to (top) and after (bottom) addition
of NaCN (10 equiv) in DMSO-d6 at T = 298 K. Asterisks indicate the resonances of
the minor tautomer which is less than 10% based on the peak integration values.
(b) Variable-temperature 1H NMR spectra of 1 (4.0 mM) in DMSO-d6 at T = 293–
363 K.
Fig. 12 Partial 2D-COSY contour plot of 1 (4.0 mM) treated with NaCN (10 equiv)
in DMSO-d6 at T = 298 K. The corresponding 1D NMR spectrum is shown along the
ordinate. Inset: DFT (B3LYP-D3/6-31G(d,p)) energy-minimized structure of the
cyanohydrin adduct of 1.
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A large enhancement in the emission intensity was also
View Article Online
DOI: 10.1039/D0SC05067A
observed at λem = 450 nm when cyanide anion was subsequently
added to the mixture of
We suspect that deprotonation by strong base could disrupt the
cascade hydrogen bonding, thus diminishing the response of
toward subsequently added CN–. The reaction stoichiometry
between and cyanide was determined by Job plot analysis
1
and other anions except OH– (Fig. 14c).
1
1
using UV–vis spectroscopy. A sharp maximum at the mole
fraction of 0.5 provides compelling evidence for the formation
of 1:1 adduct (Fig. 14d), which is also consistent with the results
from 1H NMR (Fig. 11a and 12) and HPLC-MS studies (Fig. S11).
Kinetic Studies: Rapid Detection of Cyanide under Ambient
Conditions
The rate constant for the bimolecular reaction between
1 and
cyanide was determined by taking time-dependent UV–vis
absorption spectra (Fig. S12). A large enhancement in the
absorption at λ = 400 nm brought by the addition of cyanide
anion helped track the progress of the reaction over time (Fig.
10b). The activation of electrophilic aldehyde group by the
hydrogen bonding array led to a rapid chemical transformation.
Fig. 13 Effects of hydrogen bonding on distinguishing Brønsted base from
nucleophilic Lewis base. Each bar denotes the ratio of fluorescence intensity at λem
= 450 nm determined for each probes (1–4, 0.100 mM) after treating with NaCN
(25 equiv, INaCN) or with Et3N (25 equiv, IEt3N) in DMSO at T = 298 K. λexc = 343 nm
for 1, 3, and 4; 400 nm for 2.
Selectivity toward Cyanide and Reaction Stoichiometry
Even at low temperature (T = 15 °C), the reaction of
1 with
cyanide in DMSO–MeCN (1:1, v/v) was completed within < 2
seconds (Fig. S12a). Under pseudo-first-order kinetic reaction
conditions, the kinetic trace was fitted to obtain pseudo-first-
order rate constants, k´ (= k2[CN–]0); the second-order rate
constant k2 was estimated from the linear relationship between
k´ and [CN–]0 (Fig. S12b). While the precise determination of
rate constants was hampered by the fast reaction rate, the
calculated k2 value (1.3 × 103 M–1s–1) is one of the fastest-
responding cyanide probes that operate by covalent capture
strategy.32,43–50 With 20 equiv of cyanide anion, the reaction
half-life t1/2 is as short as 0.53 sec.
To test the selectivity of the probe
1, aqueous solution samples
of 12 different anions, including CN-, F-, Cl-, Br-, I-, N3 , SCN-, OAc-,
-
NO3 , ClO4 , PF6 , and OH- (25 equiv, delivered as sodium salts
except for KPF6 and KOH) were added to (0.100 mM) in
-
-
-
1
DMSO–H2O (99:1, v/v) mixed-solvent system, and the emission
spectra recorded under identical conditions. As summarized in
Fig. 14a and 14b, the fluorescence turn-on response was
observed exclusively for cyanide anion.
Conclusions
As a synthetic mimic of biological hydrogen bonding triad, a T-
shaped π-conjugated platform was structurally elaborated. In
our molecular design, the amphoteric benzimidazole core
reinforces bond-polarizing HBD-HBA networks to activate an
electrophilic aldehyde group for covalent capture of toxic CN–
anion. We found that a systematic increase in the number of
hydrogen bonds allows the molecules to distinguish bond-
making nucleophile from proton-abstracting Brønsted base. The
most advanced molecular probe
1 has four hydrogen bonds
around the fluorogenic benzimidazole core, and detects cyanide
ion by a rapid and selective turn-on response. Efforts are
currently underway in our laboratory to expand the scope of
this non-covalent design strategy to other types of chemical
transformations of relevance to target-specific signal
transduction.
Fig. 14 (a) Emission spectra of 1 after treatment with CN– (25 equiv, blue line) or
other 11 anions (25 equiv, gray lines; see the list in (b) and (c)). (b) Normalized
emission intensity (= I/I0) of 1 in the presence of various anions: 1, CN–; 2, F–; 3, Cl–
; 4, Br–; 5, I–; 6, N3–; 7, SCN–; 8, OAc–; 9, NO3–; 10, ClO4–; 11, PF6–; 12, OH– (delivered
as sodium salts except for KPF6 and KOH). The intensity at λ = 450 nm (= I) after
the treatment of 1 with each anion (25 equiv) is normalized with that of 1 (= I0).
(c) Enhancement of the emission intensity after addition of NaCN (25 equiv) to 1
in the presence of each anion (25 equiv). Conditions: λexc = 343 nm; T = 298 K; [1]
= 0.100 mM in DMSO–H2O (99:1, v/v). (d) Job plot analysis of the reaction of 1 with
CN– in DMSO at T = 298 K. The absorbance at λ = 410 nm was recorded by varying
the mole fraction of CN– while keeping the total concentration constant (= 67 μM).
Conflicts of interest
There are no conflicts to declare.
This journal is © The Royal Society of Chemistry 20xx
J. Name., 2013, 00, 1-3 | 7
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ARTICLE
Journal Name
28 H. J. Kim, C. H. Heo and H. M. Kim, J. Am. Chem. Soc., 2013,
Acknowledgements
This work was supported by the National Research Foundation
(NRF) of Korea (2020R1A2C2006381 and 2017M3D1A1039558)
funded by the Ministry of Science and ICT.
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DOI: 10.1039/D0SC05067A
ARTICLE
Table of Contents
Biomimetic cascade hydrogen bonds promote covalent capture of the nucleophile by polarizing electrophilic reaction site, while
suppressing non-productive acid–base chemistry as the competing reaction pathway.
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