Kwakman P J, Kamminga D A, Brinkman U A, de Jong G J
Department of Analytical Chemistry, Free University, Amsterdam, The Netherlands.
J Pharm Biomed Anal. 1991;9(9):753-9. doi: 10.1016/0731-7085(91)80217-w.
Column liquid chromatography with chemiluminescence detection is used to determine oestradiol in serum. Oestradiol is labelled by means of a two-phase derivatization procedure with either dansyl chloride or laryl chloride. After derivatization the excess reagent is removed on a short amino-bonded column: the sulphonyl chloride functional group reacts with the amino groups causing removal of the label; the oestradiol derivatives are not retained. Chromatography of oestradiol derivatives is performed with methanol-water mixtures as eluent. Chemical excitation is carried out by adding bis(2-nitrophenyl)oxalate and hydrogen peroxide dissolved in acetonitrile to the column eluate. Linearity is observed (r = 0.9998; n = 7) over at least three decades (10(-6)-10(-9) M) for both dansyl chloride and laryl chloride. A detection limit of 5 x 10(-10) M (50 fmol injected) has been achieved and the dansyl derivatization method has been applied to the determination of oestradiol in spiked serum.
