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  • Title: Radioimmunological analysis of ethinylestradiol in human serum. Validation of the method and comparison with a gas chromatographic/mass spectrometric assay.
    Author: Kuhnz W, Louton T, Back DJ, Michaelis K.
    Journal: Arzneimittelforschung; 1993 Jan; 43(1):16-21. PubMed ID: 8447841.
    Abstract:
    The majority of combination oral contraceptives contain ethinylestradiol (EE2, CAS 57-63-6) as estrogenic component at doses between 50 and 20 micrograms/unit. Since the concentrations of EE2 in the serum of women under oral contraceptive (OC) therapy are in the lower pg-range, highly sensitive and specific analytical methods are required. Radioimmunoassay (RIA) has been the method of choice, but evidence of specificity in the presence of the coadministered progestogens and their metabolites has not always been provided. The present study compares two radioimmunological methods, which use the same antiserum but different sample volumes and standard curves (extracted vs. non-extracted), with a newly developed gas chromatographic/mass spectrometric (GC/MS) method, in order to cross-validate the methods. For that purpose, 51 serum samples obtained from women who had been taking two different combination oral contraceptives were analysed independently by all three methods. The specificity of the antiserum was further examined by submitting ex vivo serum samples obtained from OC-users to a combination of high pressure liquid chromatography (HPLC) and RIA. The results obtained by the three methods were very similar and correlation coefficients (r) obtained from linear regression analysis were about 0.7. There was no interference from the coadministered progestins on the analysis of EE2 as carried out by the three methods. There were no metabolites in the extracts of ex vivo serum samples which showed cross-reactivity with the antiserum used. Modifications of the radioimmunoassay procedure did not markedly affect the results of EE2 determination. If properly validated, radioimmunoassay can be used as an alternative to GC/MS in pharmacokinetic studies.
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