[Simultaneous determination of glucocorticoids in bovine plasma by high-pressure liquid chromatography]

A method is described for the simultaneous determination of major glucocorticoids in bovine plasma, that is, Cortisol, Corticosterone and Cortisone, which includes separation by high-pressure liquid chromatography and measurement using a UV detector (254 nm). Chromatography on the Zorbax CN column is carried out with the eluent system, that is, n-hexane/ethanol (=87/13 v/v). First, Dexamethasone was selected as the best internal standard under the previously mentioned conditions. The calibration curves for the glucocorticoids were all clearly linear. The recoveries for each 5 ng of added Cortisol, Corticosterone and Cortisone were 96.7 +/- 3.2%, 100.4 +/- 2.3% and 96.1 +/- 2.2%, respectively. The reproducibility of Cortisol was good and its coefficient of variation of intraassay was small (2.7%), whereas such coefficient was not so in Corticosterone (7.7%) and Cortisone (7.3%). This appears to be due to low concentrations of Corticosterone and Cortisone in the plasma. Specificity of this method to each glucocorticoid was ascertained using plasma from adrenalectomized wethers. In addition, the correlation coefficient between measurements of Cortisol by radioimmunoassay and those by this method was 0.978. In conclusion, the method described in this paper appears to be satisfactory for the simultaneous quantification of plasma glucocorticoids.