Acrylamide is an efficient quencher of tryptophanyl fluorescence which we report to be very discriminating in sensing the degree of exposure of this residue in proteins. The quenching reaction involves physical contact between the quencher and an excited indole ring, and can be kinetically described in terms of a collisional and a static component. The rate constant for the collisional component is a kinetic measure of the exposure of a residue in a protein, and values ranging from 4 X 10(9) M-1 S-1 for the fully exposed tryptophan in the polypeptide, adrenocorticotropin, to less than 5 X 10(8) M-1 S-1 for the buried residue in azurin have been found. Static quenching is readily detected in proteins that are denatured, or contain only a single fluorophor. Quenching patterns for most multi-tryptophan containing proteins are difficult to analyze precisely, but qualitative information can, nevertheless, be extracted. Applications of this probing technique for monitoring protein conformational changes, such as the acid-induced expansion of human serum albumin, and inhibitor binding to enzymes, are presented. The value of this method lies in its ability to sense not only the steady-state exposure of a residue in a protein, but also its dynamic exposure.

Exposure of tryptophanyl residues in proteins. Quantitative determination by fluorescence quenching studies.

Absorption and fluorescence properties of fluorescein

Organic light-emitting diodes (OLEDs) have their performance limited by the number of emissive singlet states created upon charge recombination (25%). Recently, a novel strategy has been proposed, based on thermally activated up-conversion of triplet to singlet states, yielding delayed fluorescence (TADF), which greatly enhances electroluminescence. The energy barrier for this reverse intersystem crossing mechanism is proportional to the exchange energy (ΔEST ) between the singlet and triplet states; therefore, materials with intramolecular charge transfer (ICT) states, where it is known that the exchange energy is small, are perfect candidates. However, here it is shown that triplet states can be harvested with 100% efficiency via TADF, even in materials with ΔEST of more than 20 kT (where k is the Boltzmann constant and T is the temperature) at room temperature. The key role played by lone pair electrons in achieving this high efficiency in a series of ICT molecules is elucidated. The results show the complex photophysics of efficient TADF materials and give clear guidelines for designing new emitters.

Triplet harvesting with 100% efficiency by way of thermally activated delayed fluorescence in charge transfer OLED emitters.

Publisher Summary Fluorescence resonance energy transfer (FRET) is a spectroscopic process by which energy is passed nonradiatively among molecules over long distances. The “donor” molecule, which must be a fluorophore, absorbs a photon and transfer this energy nonradiatively to the “acceptor” molecule. Energy can be transferred over distances on the order of common macromolecular dimensions. This chapter discusses the applications of FRET to nucleic acid molecules conjugated to dye molecules; but in general, the donor-acceptor pairs can be free in solution, one or both bound to a macromolecule, or be an inherent part of the structure. The chapter discusses FRET specifying the physical parameters that are available from a variety of fluorescence measurements. In the chapter, some basic considerations and experimental procedures are described that are necessary to make reliable FRET measurements. It also discusses some difficulties that can arise in the experimental proceedings and data analysis and the sources of experimental error are pointed out.

Fluorescence resonance energy transfer and nucleic acids.

Publisher Summary This chapter discusses acid–base properties of electronically excited states of organic molecules. Excited state pK-values are most easily accessible through the use of the Forster cycle. To perform this calculation for a particular molecule, it is necessary to know the ground state equilibrium constant for the reaction in question and to have some measure of the energy difference between the lowest vibrational level of the ground and the excited state in both the B and BH + forms. The effects of solvation on 0–0 energies are discussed. The changes in molecular fluorescence with acidity give information about the protolytic behavior of the excited singlet state of a compound. Two techniques, phase and pulse fluorometry, are used for the direct measurement of fluorescence decay rates. The excited state acid-base behavior of molecules has direct implications in the field of analytical fluorimetry and phosphorimetry.

D. Webb

Papers

Photoluminescence of solutions