Correction: Miranda et al. Anion Binding by Fluorescent Ureido-Hexahomotrioxacalix[3]arene Receptors: An NMR, Absorption and Emission Spectroscopic Study. Molecules 2022, 27, 3247.

In the original publication [...].

Are "Carbon Dots" Always Carbon Dots? Evidence for their Supramolecular Nature from Structural and Dynamic Studies in Solution and in the Pure Solid.

A model for the morphology (size, shape, and crystallinity) of carbon dots (CDs) in the solid state consistent with the observed photoluminescence in solution is proposed herein. Overwhelming evidence has been collected that links the data coming from solid-state analysis (high-resolution transmission electron microscopy (HRTEM), atomic force microscopy (AFM), and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS)) to that of solution (pulsed-field gradient (PFG)-NMR spectroscopy, time-resolved fluorescence anisotropy (TRFA), and steady-state/time-resolved fluorescence), allowing the establishment of an overall structural model for CDs. According to this model, the so-called carbon dots, observed under HRTEM imaging, are in fact supramolecular organized structures dynamically assembled from small to medium-sized molecular species when the solvent is removed to give the solid form. In this way, the imaged nanoparticles (TEM/AFM) are not covalently bound entities formed during the synthetic process, but instead supramolecular entities formed by noncovalent interactions. These particles, if at all present in solution, have the form of loose associations of relatively small molecules. This study was conducted on CDs obtained from the hydrothermal carbonization (HTC) of a biomass waste (olive wet pomace).

Are "Carbon Dots" Always Carbon Dots? Evidence for Their Supramolecular Nature from Structural and Dynamic Studies in Solution and in the Pure Solid.

Invited for the cover of this issue are José Prata and co-workers at Instituto Superior de Engenharia de Lisboa and Instituto Superior Técnico. The image depicts a model for the supramolecular nature of carbon dots. Read the full text of the article at 10.1002/chem.202302955.

Fluorescent homooxacalixarenes: recent applications in supramolecular systems.

This review covers recent advances (from 2006 to date) in supramolecular systems based on fluorescent homooxacalixarenes, namely hexahomotrioxacalix[3]arenes, dihomooxacalix[4]arenes and tetrahomodioxacalix[4]arenes, focusing on fluorescence sensing using their intrinsic fluorescence (built-in mesitol-like groups) or the extrinsic fluorescence of organic fluorophores, either covalently linked to the calixarenes or forming supramolecular complexes with them. Sensing applications of ions, ion pairs and neutral molecules are discussed, as well as the potential measurement of temperature based on thermally activated delayed fluorescence.

Critical Analysis of Association Constants between Calixarenes and Nitroaromatic Compounds Obtained by Fluorescence. Implications for Explosives Sensing.

The binding behaviour of two ureido-hexahomotrioxacalix[3]arene derivatives bearing naphthyl (1) and pyrenyl (2) fluorogenic units at the lower rim towards selected nitroaromatic compounds (NACs) was evaluated. Their affinity, or lack of it, was determined by UV-Vis absorption, fluorescence and NMR spectroscopy. Different computational methods were also used to further investigate any possible complexation between the calixarenes and the NACs. All the results show no significant interaction between calixarenes 1 and 2 and the NACs in either dichloromethane or acetonitrile solutions. Moreover, the fluorescence quenching observed is only apparent and merely results from the absorption of the NACs at the excitation wavelength (inner filter effect). This evidence is in stark contrast with reports in the literature for similar calixarenes. A naphthyl urea dihomooxacalix[4]arene (3) is also subject to the inner filter effect and is shown to form a stable complex with trinitrophenol; however, the equilibrium association constant is greatly overestimated if no correction is applied (9400 M-1 vs 3000 M-1), again stressing the importance of taking into account the inner filter effect in these systems.

Luminescent Carbon Dots from Wet Olive Pomace: Structural Insights, Photophysical Properties and Cytotoxicity.

Carbon nanomaterials endowed with significant luminescence have been synthesized for the first time from an abundant, highly localized waste, the wet pomace (WP), a semi-solid by-product of industrial olive oil production. Synthetic efforts were undertaken to outshine the photoluminescence (PL) of carbon nanoparticles through a systematic search of the best reaction conditions to convert the waste biomass, mainly consisting in holocellulose, lignin and proteins, into carbon dots (CDs) by hydrothermal carbonization processes. Blue-emitting CDs with high fluorescence quantum yields were obtained. Using a comprehensive set of spectroscopic tools (FTIR, Raman, XPS, and 1H/13C NMR) in combination with steady-state and time-resolved fluorescence spectroscopy, a rational depiction of WP-CDs structures and their PL properties was reached. WP-CDs show the up-conversion of PL capabilities and negligible cytotoxicity against two mammalian cell lines (L929 and HeLa). Both properties are excellent indicators for their prospective application in biological imaging, biosensing, and dynamic therapies driven by light.

Characterization of the Aeration and Hydrodynamics in Vertical-Wheel™ Bioreactors.

In this work, the oxygen transport and hydrodynamic flow of the PBS Vertical-Wheel MINI™ 0.1 bioreactor were characterized using experimental data and computational fluid dynamics simulations. Data acquired from spectroscopy-based oxygenation measurements was compared with data obtained from 3D simulations with a rigid-lid approximation and LES-WALE turbulence modeling, using the open-source software OpenFOAM-8. The mass transfer coefficients were determined for a range of stirring speeds between 10 and 100 rpm and for working volumes between 60 and 100 mL. Additionally, boundary condition, mesh refinement, and temperature variation studies were performed. Lastly, cell size, energy dissipation rate, and shear stress fields were calculated to determine optimal hydrodynamic conditions for culture. The experimental results demonstrate that the kL can be predicted using Sh=1.68Re0.551Sc13G1.18, with a mean absolute error of 2.08%. Using the simulations and a correction factor of 0.473, the expression can be correlated to provide equally valid results. To directly obtain them from simulations, a partial slip boundary condition can be tuned, ensuring better near-surface velocity profiles or, alternatively, by deeply refining the mesh. Temperature variation studies support the use of this correlation for temperatures up to 37 °C by using a Schmidt exponent of 1/3. Finally, the flow was characterized as transitional with diverse mixing mechanisms that ensure homogeneity and suspension quality, and the results obtained are in agreement with previous studies that employed RANS models. Overall, this work provides new data regarding oxygen mass transfer and hydrodynamics in the Vertical-Wheel bioreactor, as well as new insights for air-water mass transfer modeling in systems with low interface deformation, and a computational model that can be used for further studies.

Anion Binding by Fluorescent Ureido-Hexahomotrioxacalix[3]arene Receptors: An NMR, Absorption and Emission Spectroscopic Study.

Fluorescent receptors (4a-4c) based on (thio)ureido-functionalized hexahomotrioxacalix[3]arenes were synthesised and obtained in the partial cone conformation in solution. Naphthyl or pyrenyl fluorogenic units were introduced at the lower rim of the calixarene skeleton via a butyl spacer. The binding of biologically and environmentally relevant anions was studied with NMR, UV-vis absorption, and fluorescence titrations. Fluorescence of the pyrenyl receptor 4c displays both monomer and excimer fluorescence. The thermodynamics of complexation was determined in acetonitrile and was entropy-driven. Computational studies were also performed to bring further insight into the binding process. The data showed that association constants increase with the anion basicity, and AcO-, BzO- and F- were the best bound anions for all receptors. Pyrenylurea 4c is a slightly better receptor than naphthylurea 4a, and both are more efficient than naphthyl thiourea 4b. In addition, ureas 4a and 4c were also tested as ditopic receptors in the recognition of alkylammonium salts.

Reversible Electronic Energy Transfer (Homo-FRET) in Cyclic Molecular and Supramolecular Systems: Fluorescence Anisotropy Decays for the Isotropic Interaction.

Reversible electronic energy transfer (homo-FRET) in cyclic multichromophoric systems is studied for sets of n identical fluorophores arranged in regular polygons (triangle, square, pentagon, etc.). A general analytic expression for the anisotropy decay is obtained for a regular polygon of any order, under the assumptions of isotropic interaction and nearest-neighbor FRET. A graphical way of connecting the decay form and polygon geometry based on the Frost circle is also presented. The consequences of the relaxation of these assumptions on the anisotropy decay are also discussed and analyzed in detail for the heptagon.

Conventional vs. Microwave- or Mechanically-Assisted Synthesis of Dihomooxacalix[4]arene Phthalimides: NMR, X-ray and Photophysical Analysis.

Direct O-alkylation of p-tert-butyldihomooxacalix[4]arene (1) with N-(bromopropyl)- or N-(bromoethyl)phthalimides and K2CO3 in acetonitrile was conducted under conventional heating (reflux) and using microwave irradiation and ball milling methodologies. The reactions afforded mono- and mainly distal di-substituted derivatives in the cone conformation, in a total of eight compounds. They were isolated by column chromatography, and their conformations and the substitution patterns were established by NMR spectroscopy (1H, 13C, COSY and NOESY experiments). The X-ray structures of four dihomooxacalix[4]arene phthalimide derivatives (2a, 3a, 3b and 5a) are reported, as well as their photophysical properties. The microwave (MW)-assisted alkylations drastically reduced the reaction times (from days to less than 45 min) and produced higher yields of both 1,3-di-substituted phthalimides (3a and 6a) with higher selectivity. Ball milling did not reveal to be a good method for this kind of reaction.

Dihomooxacalix[4]arene-Based Fluorescent Receptors for Anion and Organic Ion Pair Recognition.

Fluorescent dihomooxacalix[4]arene-based receptors 5a-5c, bearing two naphthyl(thio)ureido groups at the lower rim via a butyl spacer, were synthesised and obtained in the cone conformation in solution. The X-ray crystal structures of 1,3- (5a) and 3,4-dinaphthylurea (5b) derivatives are reported. Their binding properties towards several anions of different geometries were assessed by 1H-NMR, UV-Vis absorption and fluorescence titrations. Structural and energetic insights of the naphthylurea 5a and 5b complexes were also obtained using quantum mechanical calculations. The data showed that all receptors follow the same trend, the association constants increase with the anion basicity, and the strongest complexes were obtained with F-, followed by the oxoanions AcO- and BzO-. Proximal urea 5b is a better anion receptor compared to distal urea 5a, and both are more efficient than thiourea 5c. Compounds 5a and 5b were also investigated as heteroditopic receptors for biologically relevant alkylammonium salts, such as the neurotransmitter γ-aminobutyric acid (GABA·HCl) and the betaine deoxycarnitine·HCl. Chiral recognition towards the guest sec-butylamine·HCl was also tested, and a 5:2 selectivity for (R)-sec-BuNH3+·Cl- towards (P) or (M) enantiomers of the inherently chiral receptor 5a was shown. Based on DFT calculations, the complex [(S)-sec-BuNH3+·Cl-/(M)-5a] was indicated as the more stable.

Finding Value in Wastewaters from the Cork Industry: Carbon Dots Synthesis and Fluorescence for Hemeprotein Detection.

Valorisation of industrial low-value waste residues was preconized. Hence, carbon dots (C-dots) were synthesized from wastewaters of the cork industry-an abundant and affordable, but environmentally-problematic industrial effluent. The carbon nanomaterials were structurally and morphologically characterised, and their photophysical properties were analysed by an ensemble of spectroscopy techniques. Afterwards, they were successfully applied as highly-sensitive fluorescence probes for the direct detection of haemproteins. Haemoglobin, cytochrome c and myoglobin were selected as specific targets owing to their relevant roles in living organisms, wherein their deficiencies or surpluses are associated with several medical conditions. For all of them, remarkable responses were achieved, allowing their detection at nanomolar levels. Steady-state and time-resolved fluorescence, ground-state UV-Vis absorption and electronic circular dichroism techniques were used to investigate the probable mechanisms behind the fluorescence turn-off of C-dots. Extensive experimental evidence points to a static quenching mechanism. Likewise, resonance energy transfer and collisional quenching have been discarded as excited-state deactivating mechanisms. It was additionally found that an oxidative, photoinduced electron transfer occurs for cytochrome c, the most electron-deficient protein. Besides, C-dots prepared from citric acid/ethylenediamine were comparatively assayed for protein detection and the differences between the two types of nanomaterials highlighted.

Silica nanoparticles with thermally activated delayed fluorescence for live cell imaging.

Thermally activated delayed fluorescence (TADF) has revolutionized the field of organic light emitting diodes owing to the possibility of harvesting non-emissive triplet states and converting them in emissive singlet states. This mechanism generates a long-lived delayed fluorescence component which can also be used in sensing oxygen concentration, measuring local temperature, or on imaging. Despite this strong potential, only recently TADF has emerged as a powerful tool to develop metal-free long-lived luminescent probes for imaging and sensing. The application of TADF molecules in aqueous and/or biological media requires specific structural features that allow complexation with biomolecules or enable emission in the aggregated state, in order to retain the delayed fluorescence that is characteristic of these compounds. Herein we demonstrate a facile method that maintains the optical properties of solvated dyes by dispersing TADF molecules in nanoparticles. TADF dye-doped silica nanoparticles are prepared using a modified fluorescein fluorophore. However, the strategy can be used with many other TADF dyes. The covalent grafting of the TADF emitter into the inorganic matrix effectively preserves and transfers the optical properties of the free dye into the luminescent nanomaterials. Importantly, the silica matrix is efficient in shielding the dye from solvent polarity effects and increases delayed fluorescence lifetime. The prepared nanoparticles are effectively internalized by human cells, even at low incubation concentrations, localizing primarily in the cytosol, enabling fluorescence microscopy imaging at low dye concentrations.

Ditopic Receptors Based on Dihomooxacalix[4]arenes Bearing Phenylurea Moieties With Electron-Withdrawing Groups for Anions and Organic Ion Pairs.

Two bidentate dihomooxacalix[4]arene receptors bearing phenylurea moieties substituted with electron-withdrawing groups at the lower rim via a butyl spacer (CF3-Phurea 5b and NO2 Phurea 5c) were obtained in the cone conformation in solution, as shown by NMR. The X-ray crystal structure of 5b is reported. The binding affinity of these receptors toward several relevant anions was investigated by 1H NMR, UV-Vis absorption in different solvents, and fluorescence titrations. Compounds 5b and 5c were also tested as ditopic receptors for organic ion pairs, namely monoamine neurotransmitters and trace amine hydrochlorides by 1H NMR studies. The data showed that both receptors follow the same trend and, in comparison with the unsubstituted phenylurea 5a, they exhibit a significant enhancement on their host-guest properties, owing to the increased acidity of their urea NH protons. NO2-Phurea 5c is the best anion receptor, displaying the strongest complexation for F-, closely followed by the oxoanions BzO-, AcO-, and HS O 4 - . Concerning ion pair recognition, both ditopic receptors presented an outstanding efficiency for the amine hydrochlorides, mainly 5c, with association constants higher than 109 M-2 in the case of phenylethylamine and tyramine.

Conformational plasticity in the KcsA potassium channel pore helix revealed by homo-FRET studies.

Potassium channels selectivity filter (SF) conformation is modulated by several factors, including ion-protein and protein-protein interactions. Here, we investigate the SF dynamics of a single Trp mutant of the potassium channel KcsA (W67) using polarized time-resolved fluorescence measurements. For the first time, an analytical framework is reported to analyze the homo-Förster resonance energy transfer (homo-FRET) within a symmetric tetrameric protein with a square geometry. We found that in the closed state (pH 7), the W67-W67 intersubunit distances become shorter as the average ion occupancy of the SF increases according to cation type and concentration. The hypothesis that the inactivated SF at pH 4 is structurally similar to its collapsed state, detected at low K+, pH 7, was ruled out, emphasizing the critical role played by the S2 binding site in the inactivation process of KcsA. This homo-FRET approach provides complementary information to X-ray crystallography in which the protein conformational dynamics is usually compromised.

Calixarenes as High Temperature Matrices for Thermally Activated Delayed Fluorescence: C70 in Dihomooxacalix[4]arene.

Thermally activated delayed fluorescence (TADF) of 12C70 and 13C70 was observed up to 140 °C in a p-tert-butyldihomooxacalix[4]arene solid matrix, a temperature range significantly higher than that of previous TADF quantitative studies. An effective singlet-triplet energy gap of 29 kJ/mol and triplet formation quantum yields of 0.97 and 0.99 were measured for 12C70 and 13C70, respectively. The photophysical properties of the two fullerenes in this new matrix are comparable to those obtained in polystyrene at a lower temperature range. Calixarenes are proposed to be suitable matrices for high temperature TADF studies and applications.

Towards the Development of a Low-Cost Device for the Detection of Explosives Vapors by Fluorescence Quenching of Conjugated Polymers in Solid Matrices.

Conjugated polymers (CPs) have proved to be promising chemosensory materials for detecting nitroaromatic explosives vapors, as they quickly convert a chemical interaction into an easily-measured high-sensitivity optical output. The nitroaromatic analytes are strongly electron-deficient, whereas the conjugated polymer sensing materials are electron-rich. As a result, the photoexcitation of the CP is followed by electron transfer to the nitroaromatic analyte, resulting in a quenching of the light-emission from the conjugated polymer. The best CP in our studies was found to be poly[(9,9-dioctylfluorenyl-2,7-diyl)-co-bithiophene] (F8T2). It is photostable, has a good absorption between 400 and 450 nm, and a strong and structured fluorescence around 550 nm. Our studies indicate up to 96% quenching of light-emission, accompanied by a marked decrease in the fluorescence lifetime, upon exposure of the films of F8T2 in ethyl cellulose to nitrobenzene (NB) and 1,3-dinitrobenzene (DNB) vapors at room temperature. The effects of the polymeric matrix, plasticizer, and temperature have been studied, and the morphology of films determined by scanning electron microscopy (SEM) and confocal fluorescence microscopy. We have used ink jet printing to produce sensor films containing both sensor element and a fluorescence reference. In addition, a high dynamic range, intensity-based fluorometer, using a laser diode and a filtered photodiode was developed for use with this system.

The Role of Local Triplet Excited States and D-A Relative Orientation in Thermally Activated Delayed Fluorescence: Photophysics and Devices.

Here, a comprehensive photophysical investigation of a the emitter molecule DPTZ-DBTO2, showing thermally activated delayed fluorescence (TADF), with near-orthogonal electron donor (D) and acceptor (A) units is reported. It is shown that DPTZ-DBTO2 has minimal singlet-triplet energy splitting due to its near-rigid molecular geometry. However, the electronic coupling between the local triplet (3LE) and the charge transfer states, singlet and triplet, (1CT, 3CT), and the effect of dynamic rocking of the D-A units about the orthogonal geometry are crucial for efficient TADF to be achieved. In solvents with low polarity, the guest emissive singlet 1CT state couples directly to the near-degenerate 3LE, efficiently harvesting the triplet states by a spin orbit coupling charge transfer mechanism (SOCT). However, in solvents with higher polarity the emissive CT state in DPTZ-DBTO2 shifts below (the static) 3LE, leading to decreased TADF efficiencies. The relatively large energy difference between the 1CT and 3LE states and the extremely low efficiency of the 1CT to 3CT hyperfine coupling is responsible for the reduction in TADF efficiency. Both the electronic coupling between 1CT and 3LE, and the (dynamic) orientation of the D-A units are thus critical elements that dictate reverse intersystem crossing processes and thus high efficiency in TADF.

Dinuclear Zinc(II) Macrocyclic Complex as Receptor for Selective Fluorescence Sensing of Pyrophosphate.

A new diethylenetriamine-derived macrocycle known as L, bearing 2-methylquinoline arms and containing m-xylyl spacers, was prepared in good yield by a one-pot [2 + 2] Schiff base condensation procedure, followed by reduction with sodium borohydride. Up to now this is the first hexaazamacrocycle with appended fluorophore units. Single-crystal X-ray diffraction determination of the dinuclear zinc(II) complex of L showed that metal centers are located at about 7.20(2) Å from one another. This complex exhibits only weak fluorescence in aqueous solution, but the addition of 1 equiv of pyrophosphate (PPi) caused a 21-fold enhancement of the fluorescence intensity. The sensor response is linear up to a value of 10 µM HPPi(3-) and has a detection limit of 300 nM. The receptor behaves as a highly selective sensor for pyrophosphate as other anions, including phosphate, phenylphosphate (PhP), adenosine monophosphate (AMP), adenosine diphosphate (ADP), and adenosine triphosphate (ATP), failed to induce any fluorescence change and practically do not affect the fluorescence intensity of the sensor in the presence of HPPi(3-). Competition titrations carried out in aqueous solution at pH 7.4 [in 20 mM 3-(N-morpholino)propanesulfonic acid (MOPS) buffer] by spectrofluorometry revealed a high association constant value of 6.22 log units for binding of PPi by the dinuclear zinc(II) receptor, one of the highest reported values for colorimetric/fluorometric sensors able to work under real aqueous physiological conditions, while association constant values for binding of the other phosphorylated substrates are in the 5.51-4.03 log unit range.

Phasor Representation of Monomer-Excimer Kinetics: General Results and Application to Pyrene.

Phasor plots of the fluorescence intensity decay (plots of the Fourier sine transform versus the Fourier cosine transform, for one or several angular frequencies) are being increasingly used in studies of homogeneous and heterogeneous systems. In this work, the phasor approach is applied to monomer-excimer kinetics. The results obtained allow a clear visualization of the information contained in the decays. The monomer phasor falls inside the universal circle, whereas the excimer phasor lies outside it, but within the double-exponential outer boundary curve. The monomer and excimer phasors, along with those corresponding to the two exponential components of the decays, fall on a common straight line and obey the generalized lever rule. The clockwise trajectories described by both phasors upon monomer concentration increase are identified. The phasor approach allows discussing in a single graphic not only the effect of concentration but also that of rate constants, including the evolution from irreversible kinetics to fast excited-state equilibrium upon a temperature increase. The obtained results are applied to the fluorescence decays of pyrene monomer and excimer in methylcyclohexane at room temperature. A straightforward method of monomer-excimer lifetime data analysis based on linear plots is also introduced.