Analysis of corticosteroids in biological fluids by high-performance liquid chromatography with fluorometric detection.

strong sulfuric acid solutions, and can be measured by a fluorometer.1) Gotelli et al.2) and Shihabi et al.3) have used high-performance liquid chromatography (HPLC) to separate fluorescent derivatives, and have measured cortisol levels in serum and urine. Since the sulfuric acid-induced fluorescence is unstable, it is necessary to inject the reaction mixture into HPLC immediately after the formation of fluorescent derivatives is completed. The difficulty in timing the manual injection of the sample may be eliminated if an automated procedure is used for the sulfuric acid reaction and sampling in HPLC. For this purpose, we tried to develop a post-label HPLC method in which the steroids are first separated by HPLC and then changed to fluorescent compounds by an automated procedure. Heparinized plasma (0.5 ml) or whole saliva (2 ml) was extracted with 15 ml