SYNTHESIS, PURIFICATION, AND STABILITY OF β-DIKETONATE EUROPIUM CHELATE REAGENT, DETERMINED BY RP-HPLC

Abstract

ABSTRACT Various estimation conditions concerned with a europium chelate labeling agent, 4-(1,1,1,2,2-pentafluoro-3,5-dioxo-pentyl)-[1,11′-biphenyl]-sulfonyl chloride (PDB-SCl), was studied, by using reversed-phase HPLC-time-resolved fluorescence detector system. Samples dissolved in acetonitrile were introduced to a TMS column; the eluent was monitored with absorbance at 280 nm, and then collected. To the fractions was added europium (Eu) solution containing tri-n-octylphosphine oxide (TOPO) and Triton X-100, and the time-resolved fluorescence intensity of the PDB-Eu chelate was measured with an off-line time-resolved fluorometer. In time-resolved fluorescence measurement, one major peak of PDB-SCl was observed, but in absorbance measurement, many peaks were observed. The fractions having time-resolved fluorescence were hydrolyzed, and the reaction solutions were analyzed by an HPLC system connected to an on-line time-resolved fluorescence detector (TRFD). There were little byproduct peaks, which were produced by reacting PDB-SCl with contaminants in the synthesized PDB-SCl powder. The stability of chlorosulfonyl group of PDB-SCl in aqueous acetonitrile solutions was also studied, and 80% of chlorosulfonyl group of PDB-SCl dissolved in 90% aqueous acetonitrile solution was hydrolyzed to sulfonic acid. However, PDB-SCl dissolved in 100% acetonitrile was stable at ambient temperature for at least 96 hours in the dark.