Highly sensitive time-resolved fluorometric determination of estrogens by high-performance liquid chromatography using a β-diketonate europium chelate

A new fluorescent europium chelate labeling reagent, 5-(4″-chlorosulfo-1′,1″-diphenyl-4′-yl)-1,1,1,2,2-pentafluoro-3,5-pentanedione (CDPP), was synthesized for the time-resolved fluorometric detection of HPLC. The label can be directly bound to amino or phenolic hydroxyl groups of analytes with its chlorosulfonyl group, and the labeled analytes are separated on a HPLC column. After separation, EuCl3, TOPO (tri-n-octylphosphine oxide), and Triton X-100 were added by post-column introduction to the eluent, and the fluorescence of the europium chelate was measured with the time-resolved fluorometric detector. Estrone (E1), 17β-estradiol (E2), ethynylestradiol (EE2) and estriol (E3) were measured with the detection limits of 0.65, 0.65, 0.65 and 0.60 ng/ml, respectively. The recovery for river water samples was in the range of 86.0–105.1% with the RSD of 1.9–5.8%. The method was applied to the analysis of a river water sample and estrone (E1) was determined to be 2.1 ng/l. The results and processing have been compared with those of a GC–MS method and a high degree of correlation (r≧0.98) was observed.