Ultrasensitive Measurement of Estrogens

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DESCRIPTION (provided by applicant): The measurement of estrogens is of interest for a better understanding of their role in both health and disease, especially breast cancer. In this project we will develop an ultrasensitive method for measuring estrogens (estradiol and its metabolites) in serum and plasma. The method will basically consist of five steps: (1) extract the estrogens from serum in the presence of stable isotope internal standards; (2) clean up the extract by evaporation and further solvent partitioning and/or solid phase extraction; (3) derivatize the estrogens with an electrophone reagent and capping reagent; (4) conduct post derivatization sample cleanup; (5) detect the estrogen derivatives by gas chromatography electron capture time-of-flight mass spectrometry (GC-EC-TOF-MS), a technique that has emerged recently. Part of the work in the project will involve the preparation of new electrophone derivatizing agents that are anticipated to provide higher sensitivity than current reagents for measurement of estrogens. We seek to measure 15 estrogens in a single procedure in amounts as small as 200 fg in 200 mu l of serum or plasma. A method with this performance could be used to measure estrogens in cohort samples from epidemiology studies, in serum of postmenopausal women, and in tissue microbiopsy samples. While there is a need for such analysis, the sensitivity of current methods fails to reach this level for real samples.