Simultaneous determination of estriol and estriol 3-sulfate in serum by column-switching semi-micro high-performance liquid chromatography with ultraviolet and electrochemical detection

A column-switching HPLC with semi-microcolumn enabled us a direct and simultaneous analysis of estriol (E3) and estriol 3-sulfate (E3 S) in human serum in combination with ultraviolet (for E3 S) and electrochemical (for E3) detectors. The mobile phases (phosphate buffer pH 7.0) contained 5 mM tetra-n-butylammonium ion (TBA) as a counter ion for E3 S. Serum samples were diluted with 200 mM phosphate buffer (pH 7.0) containing 100 mM TBA, then injected to the pre-column. After serum proteins had flowed out from the pre-column, E3 and E3 S were transferred to the enrichment column. Subsequently the analytes were eluted to the analytical column. Detection limits of E3 and E3 S in human serum were 2.5 ng/ml and 295 ng/ml. Serum E3 and E3 S levels (mean±SD) of umbilical artery from 18 full-term healthy neonates were 33±23 ng/ml and 1.26±0.69 μg/ml, respectively.