Fluorescent proton sensors based on energy transfer

Photophysical data and orbital energy levels (from electrochemistry) were compared for molecules with the same BODIPY acceptor part (red) and perpendicularly oriented xanthene or BODIPY donor fragments (green). Transfer of energy, hence the photophysical properties of the cassettes, including the pH dependent fluorescence in the xanthene-containing molecules, correlates with the relative energies of the frontier orbitals in these systems. Intracellular sensing of protons is often achieved via sensors that switch off completely at certain pH values, but probes of this type are not easy to locate inside cells in their "off-state". A communication from these laboratories (J. Am. Chem. Soc., 2009, 131, 1642-3) described how the energy transfer cassette 1 could be used for intracellular imaging of pH. This probe is fluorescent whatever the pH, but its exact photophysical properties are governed by the protonation states of the xanthene donors. This work was undertaken to further investigate correlations between structure, photophysical properties, and pH for energy transfer cassettes. To achieve this, three other cassettes 2-4 were prepared: another one containing pH-sensitive xanthene donors (2) and two "control cassettes" that each have two BODIPY-based donors (3 and 4). Both the cassettes 1 and 2 with xanthene-based donors fluoresce red under slightly acidic conditions (pH < ～6) and green when the medium is more basic (>～7), whereas the corresponding cassettes with BODIPY donors give almost complete energy transfer regardless of pH. The cassettes that have BODIPY donors, by contrast, show no significant fluorescence from the donor parts, but the overall quantum yields of the cassettes when excited at the donor (observation of acceptor fluorescence) are high (ca. 0.6 and 0.9). Electrochemical measurements were performed to elucidate orbital energy level differences between the pH-fluorescence profiles of cassettes with xanthene donors, relative to the two with BODIPY donors. These studies confirm energy transfer in the cassettes is dramatically altered by analytes that perturb relative orbital levels. Energy transfer cassettes with distinct fluorescent donor and acceptor units provide a new, and potentially useful, approach to sensors for biomedical applications.     

Synthesis of 2-substituted 1-tosyl-3-(1-tosyliminoalkyl)imidazolidines and-hexahydropyrimidines

Reactions of N-tosylimidoyl chlorides with the Schiff bases of the general formula TsNH(CH₂)_nN=CHR (n = 2 or 3; R = Prⁱ, 4-MeOC₆H₄, 4-Me₂NC₆H₄, and 3-O₂NC₆H₄) afforded 2-substituted 1-tosyl-3-(1-tosyliminoalkyl)imidazolidines (n = 2) or-hexahydropyrimidines (n = 3). Published in Russian in Izvestiya Akademii Nauk. Seriya Khimicheskaya, No. 5, pp. 872–875, May, 2006.     

Chemosensor activity of 2-(anthracen-9-yl)-substituted imidazolidines and hexahydropyrimidines

A number of 2-(anthracen-9-yl)-substituted imidazolidines and hexahydropyrimidines were synthesized by reaction of N,N′-bis[aryl(hetaryl)methyl]ethylene-1,2-diamines and N,N′-bis[aryl(hetaryl)methyl]-propane-1,3-diamines with anthracene-9-carbaldehyde. The obtained compounds showed chemosensor activity toward Cd²⁺, Cu²⁺, and Hg²⁺ ions.     

Fluorescent optical sensors

Optical sensors are prepared by immobilizing an indicating layer on the distal end of a fiber optic cable. Dyes, enzymes, and antibodies can all be incorporated into the layer using a variety of immobilization techniques. Much of the present work is devoted to developing novel indicating schemes by combining appropriate recognition schemes into polymeric matrices.     

Fluorescent sensors based on controllable conformational change for discrimination of Zn2+ over Cd2+

A new coumarin-derived imine IC1 was designed for highly selective sensing of Zn²⁺ over Cd²⁺ ions in aqueous solution based on a controllable CN isomerization mechanism. Methyl as a fine controllable unit was incorporated into IC1 to tune the conformational change of imine and thus significantly improve its selectivity to Zn²⁺ compared with methyl-free imine IC2. Sensor IC1 was also utilized to image intracellular Zn²⁺ ions in HepG2 cells with a good performance.     

Fluorescent DNA-based enzyme sensors

Fluorescent sensors that make use of DNA structures have become widely useful in monitoring enzymatic activities. Early studies focused primarily on enzymes that naturally use DNA or RNA as the substrate. However, recent advances in molecular design have enabled the development of nucleic acid sensors for a wider range of functions, including enzymes that do not normally bind DNA or RNA. Nucleic acid sensors present some potential advantages over classical small-molecule sensors, including water solubility and ease of synthesis. An overview of the multiple strategies under recent development is presented in this critical review, and expected future developments in microarrays, single molecule analysis, and in vivo sensing are discussed (160 references).     

Fluorescent sensors for nitroaromatic compounds based on monolayer assembly of polycyclic aromatics

A class of fluorescent films in which pyrene was assembled, in a monolayer manner, on glass slide surfaces via various flexible spacers of different lengths and substructures was used for the detection of nitroaromatic compounds (NACs) in vapor phase. This design strategy offers several advantages for thin film fluorescent sensory materials. These advantages have been demonstrated experimentally by the sensitive response of the films to the presence of trace amounts of NACs in vapor phase. The fluorescence quenching of the films upon exposure to NACs vapors depends on several factors, including the evaporate rate of the NAC detected, the length of the spacers connecting the sensing element and the substrate surface, and the density of the sensing element on the substrate surface. Further experimentation showed that the sensing process is reversible and free of commonly encountered interference. The sensitive response, reversibility of the sensing process, and freedom from commonly encountered interference of the specially designed films to NACs qualify these materials as promising NACs fluorescent sensory materials.     

Fluorescent sensors for Zn(2+) based on a fluorescein platform: synthesis, properties and intracellular distribution

Two new fluorescent sensors for Zn(2+) that utilize fluorescein as a reporting group, Zinpyr-1 and Zinpyr-2, have been synthesized and characterized. Zinpyr-1 is prepared in one step via a Mannich reaction, and Zinpyr-2 is obtained in a multistep synthesis that utilizes 4',5'-fluorescein dicarboxaldehyde as a key intermediate. Both Zinpyr sensors have excitation and emission wavelengths in the visible range ( approximately 500 nm), dissociation constants (K(d1)) for Zn(2+) of <1 nM, quantum yields approaching unity (Phi = approximately 0.9), and cell permeability, making them well-suited for intracellular applications. A 3- to 5-fold fluorescent enhancement is observed under simulated physiological conditions corresponding to the binding of the Zn(2+) cation to the sensor, which inhibits a photoinduced electron transfer (PET) quenching pathway. The X-ray crystal structure of a 2:1 Zn(2+):Zinpyr-1 complex has also been solved, and is the first structurally characterized example of a complex of fluorescein substituted with metal binding ligands.     

Regioselectivity of the hydrolysis of 2-(1-alkoxyvinyl)-substituted imidazolidines, 1,3-thiazolidines,and 1,3-oxazolidines

2-(1-Alkoxyvinyl)-1,3-thiazolidines reacted with H₂O or D₂O in the presence of 105 mol % of p-toluenesulfonic acid or trifluoroacetic acid (20°C, 1 h) to give 2-acetyl-1,3-thiazolidine in quantitative yield. 2-(1-Alkoxyvinyl)-3,5-diphenylimidazolidines underwent hydrolysis in the presence of 20 mol % of an acid (20°C, 24 h) at the vinyloxy group with high regioselectivity yielding 2-acetylimidazolidines. Hydrolysis of 2-(1-alkoxyvinyl)-3-phenyl-1,3-oxazolidines in the presence of 10 mol % of p-toluenesulfonic acid (20°C, 5 days) takes two pathways, one of which involves the endocyclic C-O bond with ring opening and the other involves the vinyloxy group to produce 2-acetyl-3-phenyl-1,3-oxazolidine. Unlike phenyl-substituted 1,3-thiazolidines and imidazolidines, hydrolysis of their 3-methyl- and 3,5-dimethyl-substituted analogs in acid medium occurs mainly via ring opening. The observed hydrolysis pathways were interpreted in terms of B3PW91/6-311G(d,p) quantum-chemical calculations.     

Fluorescent sensors using DNA-functionalized graphene oxide

In the past few years, graphene oxide (GO) has emerged as a unique platform for developing DNA-based biosensors, given the DNA adsorption and fluorescence-quenching properties of GO. Adsorbed DNA probes can be desorbed from the GO surface in the presence of target analytes, producing a fluorescence signal. In addition to this initial design, many other strategies have been reported, including the use of aptamers, molecular beacons, and DNAzymes as probes, label-free detection, utilization of the intrinsic fluorescence of GO, and the application of covalently linked DNA probes. The potential applications of DNA-functionalized GO range from environmental monitoring and cell imaging to biomedical diagnosis. In this review, we first summarize the fundamental surface interactions between DNA and GO and the related fluorescence-quenching mechanism. Following that, the various sensor design strategies are critically compared. Problems that must be overcome before this technology can reach its full potential are described, and a few future directions are also discussed.     

Fluorescent anion sensors based on 4-amino-1,8-naphthalimide that employ the 4-amino N-H

The new charge neutral 4-amino-1,8-naphthalimide based anion sensors 2 and 3 bind to both acetate and dihydrogenphosphate with 1:1 stoichiometry through hydrogen bonding to both thiourea N-H atoms and in the case of dihydrogenphosphate, the naphthalimide 4 amino N-H group as well. This was clearly established from ¹H NMR titration experiments with in DMSO-d₆ where a substantial shift in the resonance for the naphthalimide N-H was observed concomitant with the expected migration of the thiourea N-H chemical shifts. The binding constants determined from the titration studies indicate that the new sensors bind more strongly than AcO⁻. Fluorescence titrations with sensor 3 indicate quenching of 59% and 36% upon addition of acetate and dihydrogenphosphate, respectively.     

Fluorescent sensors for organophosphorus nerve agent mimics

We present a small molecule sensor that provides an optical response to the presence of an organophosphorus (OP)-containing nerve agent mimic. The design contains three key features: a primary alcohol, a tertiary amine in close proximity to the alcohol, and a fluorescent group used as the optical readout. In the sensor's rest state, the lone pair of electrons of the basic amine quenches the fluorescence of the nearby fluorophore through photoinduced electron transfer (PET). Exposure to an OP nerve agent mimic triggers phosphorylation of the primary alcohol followed rapidly by an intramolecular substitution reaction as the amine displaces the created phosphate. The quaternized ammonium salt produced by this cyclization reaction no longer possesses a lone pair of electrons, and a fluorescence readout is observed as the nonradiative PET quenching pathway of the fluorophore is shut down.     

Fluorescent detection of anions by dibenzophenazine-based sensors

Binding studies of two sulfonamide-functionalized dibenzophenazine-based sensors are reported. Spectroscopic studies showed that both sensors are effective fluorescent turn-on sensors for several anions. Both sensors showed responses to acetate, benzoate, cyanide, and fluoride ions. NMR titrations confirmed the mode of binding of the sensors to be through H-bonding to the sulfonamide groups.     

Fluorescent acrylamide nanoparticles for boronic acid based sugar sensing — from probes to sensors

Fluorescent nanoparticles containing covalently bound phenylboronic acids (~ 250 nm in diameter) are presented that respond to carbohydrates by swelling which is detected using fluorescence resonance energy transfer. The nanoparticles are characterized in terms of kinetics, response time and dynamic range. The response of the particles to glucose at pH 7.5 depends on the kind of phenylboronic acid and on ionic strength. The particles were immobilized in hydrogel sensor layers that enable continuous optical sensing of carbohydrates.     

2-(2-Hydroxyphenyl)imidazolidines and their O-phosphorylated derivatives

2-(2-Hydroxyaryl)imidazolidines were synthesized by reaction of aromatic carbonyl compounds with N,N′-dialkylethylenediamines. The title compounds were also prepared using the corresponding Schiff bases instead of carbonyl compounds. Phosphorylation of 2-(2-hydroxyphenyl)imidazolidines with phosphoryl and phosphorothioyl chlorides and phosphorochloridites was accomplished. The reaction of O-phosphorylsalicylaldehyde with N,N′-dialkylethylenediamines also afforded 2-(2-hydroxyphenyl)imidazolidines.     

Fluorescent sensors for amino acid anions based on calix[4]arenes bearing two dansyl groups

Two novel types of chiral calix[4]arenes containing hydrazide and dansyl groups were synthesized and examined for their enantioselective recognition abilities by the fluorescence and ¹H NMR spectra in CHCl₃. The results indicate that both 4a and 4b have excellent enantioselectivities to the N-protected alanine or phenylalanine anions.     

Fluorescent nano-PEBBLE sensors designed for intracellular glucose imaging

Polyacrylamide-based, ratiometric, spherical, optical nanosensors, or polyacrylamide PEBBLEs (Probes Encapsulated By Biologically Localized Embedding), have been fabricated, aimed at real-time glucose imaging in intact biological systems, i.e. living cells. These nanosensors are prepared using a microemulsion polymerization process, and their average size is about 45 nm in diameter. The sensors incorporate glucose oxidase (GOx), an oxygen sensitive fluorescent indicator (Ru[dpp(SO3Na)2]3)Cl2, and an oxygen insensitive fluorescent dye, Oregon Green 488-dextran or Texas Red-dextran, as a reference for the purpose of ratiometric intensity measurements. The enzymatic oxidation of glucose to gluconic acid results in the local depletion of oxygen, which is measured by the oxygen sensitive ruthenium dye. The small size and inert matrix of these sensors allows them to be inserted into living cells with minimal physical and chemical perturbations to their biological functions. The PEBBLE matrix protects the enzyme and fluorescent dyes from interference by proteins in cells, enabling reliable in vivo chemical analysis. Conversely, the matrix also significantly reduces the toxicity of the indicator and reference dyes to the cells, so that a larger variety of dyes can be used in optimal fashion. Furthermore, the PEBBLE matrix enables the synergistic approach in which there is a steady state of local oxygen consumption, and this cannot be achieved by separately introducing free enzyme and dyes into a cell. The work presented here describes the production and characterization of glucose sensitive PEBBLEs, and their potential for intracellular glucose measurements. The sensor response is determined in terms of the linear range, ratiometric operation, response time, sensor stability, reversibility and immunity to interferences.     

Fluorescent sensors for the detection of chemical warfare agents

Along with biological and nuclear threats, chemical warfare agents are some of the most feared weapons of mass destruction. Compared to nuclear weapons they are relatively easy to access and deploy, which makes them in some aspects a greater threat to national and global security. A particularly hazardous class of chemical warfare agents are the nerve agents. Their rapid and severe effects on human health originate in their ability to block the function of acetylcholinesterase, an enzyme that is vital to the central nervous system. This article outlines recent activities regarding the development of molecular sensors that can visualize the presence of nerve agents (and related pesticides) through changes of their fluorescence properties. Three different sensing principles are discussed: enzyme-based sensors, chemically reactive sensors, and supramolecular sensors. Typical examples are presented for each class and different fluorescent sensors for the detection of chemical warfare agents are summarized and compared.     

Fluorescent indicators based on BODIPY

This critical review covers the advances made using the 4-bora-3a,4a-diaza-s-indacene (BODIPY) scaffold as a fluorophore in the design, synthesis and application of fluorescent indicators for pH, metal ions, anions, biomolecules, reactive oxygen species, reactive nitrogen species, redox potential, chemical reactions and various physical phenomena. The sections of the review describing the criteria for rational design of fluorescent indicators and the mathematical expressions for analyzing spectrophotometric and fluorometric titrations are applicable to all fluorescent probes (206 references).