Control over the strength of excitonic coupling in molecular dye aggregates is a substantial factor for the development of technologies such as light harvesting, optoelectronics, and quantum ...computing. According to the molecular exciton model, the strength of excitonic coupling is inversely proportional to the distance between dyes. Covalent DNA templating was proved to be a versatile tool to control dye spacing on a subnanometer scale. To further expand our ability to control photophysical properties of excitons, here, we investigated the influence of dye hydrophobicity on the strength of excitonic coupling in squaraine aggregates covalently templated by DNA Holliday Junction (DNA HJ). Indolenine squaraines were chosen for their excellent spectral properties, stability, and diversity of chemical modifications. Six squaraines of varying hydrophobicity from highly hydrophobic to highly hydrophilic were assembled in two dimer configurations and a tetramer. In general, the examined squaraines demonstrated a propensity toward face-to-face aggregation behavior observed via steady-state absorption, fluorescence, and circular dichroism spectroscopies. Modeling based on the Kühn–Renger–May approach quantified the strength of excitonic coupling in the squaraine aggregates. The strength of excitonic coupling strongly correlated with squaraine hydrophobic region. Dimer aggregates of dichloroindolenine squaraine were found to exhibit the strongest coupling strength of 132 meV (1065 cm–1). In addition, we identified the sites for dye attachment in the DNA HJ that promote the closest spacing between the dyes in their dimers. The extracted aggregate geometries, and the role of electrostatic and steric effects in squaraine aggregation are also discussed. Taken together, these findings provide a deeper insight into how dye structures influence excitonic coupling in dye aggregates covalently templated via DNA, and guidance in design rules for exciton-based materials and devices.
Exciton delocalization plays a prominent role in the photophysics of molecular aggregates, ultimately governing their particular function or application. Deoxyribonucleic acid (DNA) is a compelling ...scaffold in which to template molecular aggregates and promote exciton delocalization. As individual dye molecules are the basis of exciton delocalization in molecular aggregates, their judicious selection is important. Motivated by their excellent photostability and spectral properties, here, we examine the ability of squaraine dyes to undergo exciton delocalization when aggregated via a DNA Holliday junction (HJ) template. A commercially available indolenine squaraine dye was chosen for the study given its strong structural resemblance to Cy5, a commercially available cyanine dye previously shown to undergo exciton delocalization in DNA HJs. Three types of DNA–dye aggregate configurationstransverse dimer, adjacent dimer, and tetramerwere investigated. Signatures of exciton delocalization were observed in all squaraine–DNA aggregates. Specifically, strong blue shift and Davydov splitting were observed in steady-state absorption spectroscopy and exciton-induced features were evident in circular dichroism (CD) spectroscopy. Strongly suppressed fluorescence emission provided additional, indirect evidence for exciton delocalization in the DNA-templated squaraine dye aggregates. To quantitatively evaluate and directly compare the excitonic Coulombic coupling responsible for exciton delocalization, the strength of excitonic hopping interactions between the dyes was obtained by simultaneously fitting the experimental steady-state absorption and CD spectra via a Holstein-like Hamiltonian, in which, following the theoretical approach of Kühn, Renger, and May, the dominant vibrational mode is explicitly considered. The excitonic hopping strength within indolenine squaraines was found to be comparable to that of the analogous Cy5 DNA-templated aggregate. The squaraine aggregates adopted primarily an H-type (dyes oriented parallel to each other) spatial arrangement. Extracted geometric details of the dye mutual orientation in the aggregates enabled a close comparison of aggregate configurations and the elucidation of the influence of dye angular relationship on excitonic hopping interactions in squaraine aggregates. These results encourage the application of squaraine-based aggregates in next-generation systems driven by molecular excitons.
Molecular aggregates exhibit emergent properties, including the collective sharing of electronic excitation energy known as exciton delocalization, that can be leveraged in applications such as ...quantum computing, optical information processing, and light harvesting. In a previous study, we found unexpectedly large excitonic interactions (quantified by the excitonic hopping parameter
J
m
,
n
) in DNA-templated aggregates of squaraine (SQ) dyes with hydrophilic-imparting sulfo and butylsulfo substituents. Here, we characterize DNA Holliday junction (DNA-HJ) templated aggregates of an expanded set of SQs and evaluate their optical properties in the context of structural heterogeneity. Specifically, we characterized the orientation of and
J
m
,
n
between dyes in dimer aggregates of non-chlorinated and chlorinated SQs. Three new chlorinated SQs that feature a varying number of butylsulfo substituents were synthesized and attached to a DNA-HJ
via
a covalent linker to form adjacent and transverse dimers. Various characteristics of the dye, including its hydrophilicity (in terms of log
P
o/w
) and surface area, and of the substituents, including their local bulkiness and electron withdrawing capacity, were quantified computationally. The orientation of and
J
m
,
n
between the dyes were estimated using a model based on Kühn-Renger-May theory to fit the absorption and circular dichroism spectra. The results suggested that adjacent dimer aggregates of all the non-chlorinated and of the most hydrophilic chlorinated SQ dyes exhibit heterogeneity; that is, they form a mixture of dimers subpopulations. A key finding of this work is that dyes with a higher hydrophilicity (lower log
P
o/w
) formed dimers with smaller
J
m
,
n
and large center-to-center dye distance (
R
m
,
n
). Also, the results revealed that the position of the dye in the DNA-HJ template, that is, adjacent or transverse, impacted
J
m
,
n
. Lastly, we found that
J
m
,
n
between symmetrically substituted dyes was reduced by increasing the local bulkiness of the substituent. This work provides insights into how to maintain strong excitonic coupling and identifies challenges associated with heterogeneity, which will help to improve control of these dye aggregates and move forward their potential application as quantum information systems.
Molecular aggregates exhibit collective sharing of electronic excitation energy known as exciton delocalization, that can be leveraged in applications such as quantum computing, optical information processing, and light harvesting.
Unlike cyanine dyes, which are widely used as fluorescent probes and labels for biomedical applications squaraine dyes are less investigated. A series of monoreactive, water-soluble, squaraine dyes ...with two aromatic sulfo groups and up to 3 sulfobutyl groups was synthesized and the spectral properties of these dyes were compared to dicarbocyanines of identical structure. Compared to the cyanines in aqueous solutions the squaraine dyes absorb and emit at shorter wavelengths (630–636 nm/639–645 nm vs. 647–653 nm/665–672 nm), have higher molar absorptivities (284,000–333,000 M−1 cm−1vs. 242,000–260,000 M−1 cm−1), lower fluorescence quantum yields (4.3–9.4% vs. 27–32%) and lower fluorescence lifetimes (0.2–0.3 ns vs. 1.0–1.2 ns) but the quantum yields and lifetimes substantially increase when bound to proteins (Bovine Serum Albumin (BSA) or antibodies, immunoglobulin G (IgG)). Squaraines with two aromatic sulfo groups show no aggregation tendency up to concentrations of 2 × 10−4 M while the corresponding cyanine dye is free of aggregation up to 5 × 10−4 M. The increase in the number of sulfobutyl groups bears a strong influence on the aggregation tendency of both dye classes upon covalent labeling to BSA and IgG resulting in increased quantum yields and lifetimes of the protein conjugates. Compared to cyanines, squaraine dyes exhibit higher photostabilities and much higher sensitivity of the quantum yields and fluorescence lifetimes toward the microenvironment and are therefore better suited as fluorescence sensors.
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•Comparison of squaraines and cyanines for use in biomedical applications.•Dyes exhibit no aggregation in water at c < 2 × 10−4 M.•Sulfo groups increase the quantum yields and lifetimes of protein conjugates.•Squaraines exhibit high photostabilities and high sensitivity to micro-environment.•Compared to cyanines, squaraines are better suited for sensing applications.
While only one enantiomer of chiral biomolecules performs a biological function, access to both enantiomers (or enantiomorphs) proved to be advantageous for technology. Using dye covalent attachment ...to a DNA Holliday junction (HJ), we created two pairs of dimers of bis(chloroindolenine)squaraine dye that enabled strongly coupled molecular excitons of opposite chirality in solution. The exciton chirality inversion was achieved by interchanging single covalent linkers of unequal length tethering the dyes of each dimer to the HJ core. Dimers in each pair exhibited profound exciton-coupled circular dichroism (CD) couplets of opposite signs. Dimer geometries, modeled by simultaneous fitting absorption and CD spectra, were related in each pair as nonsuperimposable and nearly exact mirror images. The origin of observed exciton chirality inversion was explained in the view of isomerization of the stacked Holliday junction. This study will open new opportunities for creating excitonic DNA-based materials that rely on programmable system chirality.
Molecular (dye) aggregates are a materials platform of interest in light harvesting, organic optoelectronics, and nanoscale computing, including quantum information science (QIS). Strong excitonic ...interactions between dyes are key to their use in QIS; critically, properties of the individual dyes govern the extent of these interactions. In this work, the electronic structure and excited-state dynamics of a series of indolenine-based squaraine dyes incorporating dimethylamino (electron donating) and/or nitro (electron withdrawing) substituents, so-called asymmetric dyes, were characterized. The dyes were covalently tethered to DNA Holliday junctions to suppress aggregation and permit characterization of their monomer photophysics. A combination of density functional theory and steady-state absorption spectroscopy shows that the difference static dipole moment (Δd) successively increases with the addition of these substituents while simultaneously maintaining a large transition dipole moment (μ). Steady-state fluorescence and time-resolved absorption and fluorescence spectroscopies uncover a significant nonradiative decay pathway in the asymmetrically substituted dyes that drastically reduces their excited-state lifetime (τ). This work indicates that Δd can indeed be increased by functionalizing dyes with electron donating and withdrawing substituents and that, in certain classes of dyes such as these asymmetric squaraines, strategies may be needed to ensure long τ, e.g., by rigidifying the π-conjugated network.
Novel indolenine based norsquaraine dyes, wherein the oxygen of the squaric acid bridge was substituted with a barbituric or a dicyanomethylene group, were synthesized and their molecular structure, ...spectral and luminescent properties were compared to those of analogous squaraine dyes. The molecular structure was investigated using X-ray analysis, NMR spectroscopy and ab initio DFT B3LYP/6-311G (d, p) simulations. The calculated populations of possible conformers and the barriers of internal rotations were found to be in good agreement with the experimental data. Norsquaraines absorb and emitt light within the same long-wavelength spectral range as the corresponding squaraines but due to intramolecular H-bonds and increased conformational rigidity they were less sensitive to solvent polarity and the presence of protein (BSA).
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•Novel, norsquaraine dyes were synthesized and compared to analogous squaraines.•Squaric oxygen was substituted with a barbituric and dicyanomethylene group.•The molecular structures were investigated using X-ray, NMR and ab initio.•Norsquaraine molecules are more flat, rigid and polar compared to squaraines.•Both dye classes are suitable for fluorescence based sensing applications.
Molecular excitons play a central role in natural and artificial light harvesting, organic electrònics, and nanoscale computing. The structure and dynamics of molecular excitons, critical to each ...application, are sensitively governed by molecular packing. Deoxyribonucleic acid (DNA) templating is a powerful approach that enables controlled aggregation via sub-nanometer positioning of molecular dyes. However, finer sub-Angstrom control of dye packing is needed to tailor excitonic properties for specific applications. Here, we show that adding rotaxane rings to squaraine dyes templated with DNA promotes an elusive oblique packing arrangement with highly desirable optical properties. Specifically, dimers of these squaraine:rotaxanes exhibit an absorption spectrum with near-equal intensity excitonically split absorption bands. Theoretical analysis indicates that the transitions are mostly electronic in nature and only have similar intensities over a narrow range of packing angles. Compared with squaraine dimers, squaraine:rotaxane dimers also exhibit extended excited-state lifetimes and less structural heterogeneity. The approach proposed here may be generally useful for optimizing excitonic materials for a variety of applications ranging from solar energy conversion to quantum information science.
Novel pH-sensitive, water-soluble, long-wavelength fluorescent norsquaraine dyes and their barbituric and dicyanomethylene derivatives were synthesized and investigated. Some of these dyes contain a ...carboxylic functionality that was converted into an NHS ester to facilitate bio-conjugation. The absorption and emission spectra, fluorescence quantum yields, lifetimes, polarization, and photostabilities were measured free in solution and after binding to bovine serum albumin (BSA) and compared to those of conventional squaraines and the norcyanine dye CypHer5. Contrary to squaraines, norsquaraines are pH-sensitive but almost unaffected by the interaction with proteins. These dyes can potentially be used as fluorescent labels for biomedical applications, in particular for protein labeling, polarization-based assays, cell-based and pH-sensing measurements.
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•Novel pH-sensitive, water-soluble, fluorescent norsquaraine dyes were synthesized.•Squaric oxygen was substituted with a barbituric and dicyanomethylene group.•NHS esters of some of these dyes were synthesized to facilitate bio-conjugation.•Spectral and protolytic properties were investigated and compared to squaraines.•The dyes are suitable for protein labeling, polarization and pH-sensing assays.
Advancements in single molecule detection (SMD) continue to unfold powerful ways to study the behavior of individual and complex molecular systems in real time. SMD enables the characterization of ...complex molecular interactions and reveals basic physical phenomena underlying chemical and biological processes. We present here a systematic study of the quenching efficiency of Förster-type energy-transfer (FRET) for multiple fluorophores immobilized on a single antibody. We simultaneously monitor the fluorescence intensity, fluorescence lifetime, and the number of available photons before photobleaching as a function of the number of identical emitters bound to a single IgG antibody. The detailed studies of FRET between individual fluorophores reveal complex through-space interactions. In general, even for two or three fluorophores immobilized on a single protein, homo-FRET interactions lead to an overall non-linear intensity increase and shortening of fluorescence lifetime. Over-labeling of protein in solution (ensemble) results in the loss of fluorescence signal due to the self-quenching of fluorophores making it useless for assays applications. However, in the single molecule regime, over-labeling may bring significant benefits in regards to the number of available photons and the overall survival time. Our investigation reveals possibilities to significantly increase the observation time for a single macromolecule allowing studies of macromolecular interactions that are not obscured by ensemble averaging. Extending the observation time will be crucial for developing immunoassays based on single-antibody.
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