Global Compartmental Analysis of the Excited-State Reaction between Fluorescein and (±)-N-Acetyl Aspartic Acid

Abstract

Fluorescein is a complex fluorophore which at the physiological pH can exist as mono-and dianion forms. In a previous article (J. Phys. Chem. A 2001, 105, 6320-6332), we showed that fluorescein displays an excited-state proton transfer reaction which interconverts the mono-and dianion forms in the presence of a suitable proton donor-acceptor. Because fluorescein is a frequently used fluorescent label in biological systems, it is of interest to know if amino acids with acidic side chains are able to induce fluorescein excited-state proton transfer reactions. We have selected (()-N-acetyl aspartic acid, N-AcAsp, as a model donor-acceptor which mimics the interaction of the aspartic acid residues with the fluorescent label in native proteins. We present absorption and emission properties which show that N-AcAsp at 1 M concentration induces the excitedstate proton transfer reaction between the mono-and dianion of fluorescein. The kinetics of this excited-state process was characterized from time-resolved fluorescence measurements, and the analysis was done within the framework of global compartmental analysis. Thus, we have formulated the fluorescence decay analysis of intermolecular two-state excited-state processes in the presence of buffer in terms of compartments. The kinetic equations describing the excited-state species concentrations were derived, and the expressions for the fluorescence decay surface were obtained. The experimental fluorescence decay data surface of fluorescein in the presence of 1 M N-AcAsp concentration at different pH values and excitation and emission wavelengths was analyzed via this new global compartmental analysis method, to recover the rate constants and the spectral parameters related to absorption and emission of the excited-state proton transfer reaction.

Photophysics of the Fluorescent pH Indicator BCECF

“…1 , and c 1 , whereas the only local (nonlinkable) fitting parameters were the scaling factors κ. 24 Due to the imperfection of certain traces, the global reduced g 2 remained somewhat elevated (1.14). To obtain more reliable estimates that would better fit the experimental data, we removed from the decay surface eight traces that had local 2 > 1.2, corresponding to decays that gave poor-quality biexponential fits in the previous global analyses.…”

Solvent and pH Dependent Fluorescent Properties of a Dimethylaminostyryl Borondipyrromethene Dye in Solution

“…Fluorescence decay traces of 1 were recorded at several emission wavelengths by the single-photon timing method. 52 Details of the instrumentation used 53 and experimental procedures 54 have been described elsewhere. The samples were excited at 530 or 580 nm with a repetition rate of 8.18 MHz.…”

Reduction of Self-Quenching in Fluorescent Silica-Coated Silver Nanoparticles

“…1). Fluorescein has been chosen mainly because selfquenching of this dye is a familiar observation in fluorescence spectroscopy, its spectral properties are well documented, and it is commonly used as a fluorescent probe in clinical diagnostics and research [25][26][27][28][29]. Electromagnetic field enhancement effects because of surface plasmonics have been reported for metal core-silica shell nanoparticles [30,31], but no study regarding its relationship with dye concentration and RET has been published to date.…”