Biotin as acylating agent in the Friedel–Crafts reaction. Avidin affinity of biotinyl derivatives of ferrocene, ruthenocene and pyrene and fluorescence properties of 1-biotinylpyrene

Abstract

(D)-Biotin was used for Friedel–Crafts acylation of electron-rich aromatic molecules - ferrocene, ruthenocene and pyrene. The reaction carried out in the presence of trifluoroacetic anhydride and trifluoromethanesulfonic acid afforded the corresponding biotinylarenes in moderate yields. These compounds, although lacking an amide bond, exhibited high affinity for avidin, with the ability to displace 2-(4′-hydroxyphenylazo)-benzoic acid (HABA) in its complex with avidin. Their affinity for avidin was determined by a solid-phase competitive enzymatic assay, which gave IC₅₀ values in the range of 33–58 nM (under the same conditions biotin showed IC₅₀ = 24 ± 7 nM). 1-Biotinylpyrene (1c) excited at 355 nm displayed fluorescence emission in aqueous solutions with λ_max = 461 nm. The fluorescence maximum was shifted to 425 nm upon binding of 1c to avidin. Formation of the avidin-1c complex was also evidenced by quenching of the fluorescence from the protein tryptophan residues (342 nm) and appearance of the emission band of the avidin-bound 1c at 430 nm as a result of a Förster resonance energy transfer (FRET) phenomenon.