A Sensitive method for Simultaneous quantitation of Estradiol and Estrone in human plasma by Liquid chromatography-tandem mass spectrometry with electrospray ionization

Abstract

A rapid and sensitive, sensitive liquid chromatographytandem mass spectrometry method was developed and validated for simultaneous quantitation of Estradiol and Estrone in human plasma. The analyte were extracted by liquid-liquid extraction with dicholormethane. Amlodipine was used as the internal standard. The chromatographic separation was performed with a mobile phase comprising formic acid and methanol in gradient mode. The mass transitions of m/z 506.13/170.66 for 3- dansyl-estradiol, m/z 504.10/170.66 for 3-dansyl estrone were monitored using multipe reactionmonitoring (MRM) modes for quantification. The assay was linear over the concentration range of 10-1080 pg mL-1 for Estradiol and 10-1080 pg mL-1 for estrone. The lower level of quantification in human plasma was obtained at 10.0 pg mL-1 for Estradiol and 10.0 pg mL-1 for estrone using optimum tuning parameters. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. The validated method has been applied to analyze the Estradiol and estrone concentrations in human bioequivalence study. Key Words: LC-MS/MS, Estradiol, Estrone, Amlodipine, Dansylation