GLC analysis of hydrocortisone, triamcinolone acetonide, and desonide in culture media of mouse and human dermal fibroblasts using flame-ionization detection.

A quantitative GLC assay with flame-ionization detection capable of detecting nanogram quantities of hydrocortisone, triamcinolone acetonide, and desonide in biological fluids was developed. This assay consisted of two extractions of the glucocorticoids from 1 N sodium chloride-treated cell culture media into ethyl acetate and subsequent double derivatization with methoxyamine and N-trimethylsilylimidazole. The chemical structures of methoxime-trimethylsilyl derivatives were confirmed by GLC-mass spectrometry. The methoxime-trimethylsilyl derivatives were stable for 24 hr. The applicability of this assay was demonstrated by studies of the glucocorticoid levels in L-929 and human dermal fibroblasts cell culture media over prolonged incubation (0--96 hr).