Abstract: A 2,3-diaminophenazine bis-urea fluorescent probe monomer (1) was developed. 1 responds to phenylphosphate and phosphorylated amino acids in a ratiometric fashion with en-hanced fluorescence accompanied by the development of a red-shifted emission band arising from an excited-state proton transfer (ESPT) process in the hydrogen bonded probe-analyte complex. The two urea groups of 1 form a cleft-like binding pocket (Kb >10^10 L2 mol-2 for 1:2 complex). Imprinting of 1 in presence of ethyl ester- and fluorenylmethyloxycarbon-yl- (Fmoc) protected phosphorylated tyrosine (Fmoc-pTyr-OEt) as the template, methacrylamide as comonomer and ethyleneglycol dimethacrylate as crosslinker yields few-nanometer-thick molecularly imprinted polymer (MIP) shells on silica core microparticles with excellent selectivity for the tem-plate in a buffered biphasic assay. The supramolecular recog-nition features were established by spectroscopic and NMR studies. Rational screening of comonomers and crosslinkers allowed to single out the best performing MIP components, yielding significant imprinting factors (IF>3.5) while retaining ESPT emission and the ratiometric response in the thin polymer shell. Combination of the bead-based detection scheme with the phase-transfer assay dramatically improved the IF to 15.9, allowing for sensitive determination of the analyte directly in aqueous media
Author keywords: Molecular imprinting, Excited-state proton transfer, fluorescence, core-shell particles, Phosphorylated Tyrosin