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Excitation spectroscopy of the 7F0----5D0 transition of Eu3+ and diffusion-enhanced energy transfer are used to study metal-binding characteristics of the calcium-binding protein parvalbumin from codfish. Energy is transferred from Eu3+ ions occupying the CD- and EF-binding sites to the freely-diffusing Co(III) coordination complex energy acceptors: [Co(NH3)6]3+, [Co(NH3)5H2O]3+, [CoF(NH3)5]2+, [CoCl(NH3)5]2+, [Co(NO2)3(NH3)3], and [Co(ox)3]3-. In the absence of these inorganic energy acceptors, the excited-state lifetimes of Eu3+ bound to the CD and EF sites are indistinguishable, even in D2O; however, in the presence of the positively charged energy acceptor complexes, the Eu3+ probes in the cod parvalbumin have different excited-state lifetimes due to a greater energy-transfer site from Eu3+ in the CD site than from this ion in the EF site. The observation of distinct lifetimes for Eu3+ in the two sites allows the study of the relative binding site affinities and selectivity, using other members of the lanthanide ion series. Our results indicate that during the course of a titration of the metal-free protein, Eu3+ fills the two sites simultaneously. Eu3+ is competitively displaced by other Ln3+ ions, with the CD site showing a preference for the larger Ln3+ ions while the EF site shows little, if any, competitive selectivity across the Ln3+ ion series.

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X-ray photoemission spectra of the core and valence levels for fluorescein are presented. The data were collected on a Perkin-Elmer/Physical Electronics model #5400 photoelectron spectrometer using unmonochromatized Mg x rays. The material, in powder form, was analyzed in the as received condition. Fluorescein is an important prototype organic dye with molecular formula C20H12O5. It consists of four benzene rings modified by the presence of a carboxylic, a carbonyl, a hydroxyl, and an ether type bonding. Due to the specific nature of these carbon chemistries, fluorescein is being used as a reference material to aid in understanding the chemistry of various functional groups present on oxidized activated carbons. Quantitative XPS atomic percent for carbon and oxygen are in excellent agreement with those based on its molecular formula. The various carbon and oxygen chemistries observed in this investigation are also consistent with those based on fluorescein structure.