Plasma cortisol determination by mass fragmentography

Abstract

A reference method for the estimation of plasma cortisol with high accuracy and specificity is described. A fixed amount of [4- 14C] cortisol (usually 0.2 μg) is added to a fixed amount of plasma (usually 2 ml) and the mixture is extracted with dichloromethane. The extract is treated with methoxylamine hydrochloride and trimethylsilylimidazol to yield the dimethoxime-tri-trimethylsilyl derivative of cortisol. The amount of unlabeled cortisol is determined from the ratio between the recordings at m/ e 605 and m/ e 607 obtained after analysis with a combined gas Chromatograph—mass spectrometer equipped with an MID unit (multiple ion detector). The two ions used correspond to the M-31 peak in the mass spectrum of unlabeled and (4- 14C) labeled cortisol, respectively. The relative standard deviation of the method is about 3.5%. The method was compared with a fluorometric method and a competitive protein binding method used in routine analysis. The results indicate that the competitive protein binding technique is to be preferred in routine work.