Determination of Dexamethasone in Plasma by High-Performance Liquid Chromatography with Chemiluminescence Detection

Abstract

KeywordsHigh-performance liquid hydrazide, plasma chromatography, dexamethasone, chemiluminescence, 4,5-diaminophthalDexamethasone, 9a-fluoro-16a-methyl-1 1f3,17a,21-trihydroxy-l,4-pregnadiene-3,20-dione, is a synthetic corticosteroid which has been widely used for the treatment of various kinds of diseases such as endocrinopathy, rheumatism and nephrotic syndorome owing to its anti-allergic and anti-inflammatory actions. Because dexamethasone has strong efficacy against the diseases, the dose of the drug is much smaller (several mg per day) than those of other synthetic corticosteroids. Therefore, a sensitive method is required for the determination of dexamethasone in plasma after administration for efficient and safety use of the drug.Several methods involving radioimmunoassay (RIA)1-3, gas chromatography-mass spectrometry (GC-MS)4 and liquid chromatography (LC)s-12 have been reported for the determination of dexamethasone in plasma/serum. Although the RIA methods are very sensitive, they require chromatographic purification to avoid crossreaction with other endogeneous corticosteroids. The GC-MS method needs a relatively complicated procedure for sample pretreatment. Spectrophotometric LC5-11 are not very sensitive and thus require a large amount of plasma (2 -3 ml). A fluorometric LC12, which involves precolumn derivatization with 9-anthroylnitrile, is fairly sensitive. However, the method requires solidphase extraction cartridges for the clean-up of plasma and the elimination of the excess reagent before and after the derivatization.We have previously reported a chemiluminometric LC method13 for the determination of 3a,5/3-tetrahydroaldosterone in human urine. The method is based on the conversion of the corticosteroid into the corresponding glyoxal compound by copper(II) acetate oxidation. The glyoxal compound is derivatized with 4,5-diaminophthalhydrazide (DPH), a chemiluminescence derivatization reagent for a-dicarbonyl compounds14, to yield a chemiluminescent quinoxaline. The resulting quinoxaline produces chemiluminescence by reaction with hydrogen peroxide in the presence of potassium hexacyanoferrate(III) in alkaline medium after LC separation.Recently, it was found that dexamethasone generates chemiluminescence by the same derivatization and chemiluminescence reactions as described above. Our present objective is to develop a sensitive and selective LC method involving a simple pretreatment of plasma samples by precolumn Chemiluminescence derivatization with DPH for the quantification of dexamethasone in plasma. Beclomethasone (9a-chloro-16/3-methyl-1 1f3,17a,21-trihydroxy-l,4-pregnadiene-3,20-dione), which has a structure similar to that of dexamethasone, was used as an internal standard for the precise determination of dexamethasone.Dexamethasone concentrations in plasma from a healthy volunteer after oral administration of dexamethasone were measured by the established method.

Chemiluminescence detection of nine corticosteroids in liver

“…17 Chemiluminescence has been proposed by other workers. [14][15][16][17][18] for the detection of corticosteroids, but in those cases the corticosteroids were previously derivatized with different compounds. In other work, 16 the chemiluminescence detection system used luminol in the presence of a catalyst [hexacyanoferrate(III) in alkaline solution].…”

Chemiluminescence detection in HPLC and CE for pharmaceutical and biomedical analysis

“…This method was also employed in the determination of N-acetylneuraminic acid in human serum and urine (Ishida et al, 1992b). Dexamethasone and 3a,5b-tetrahydroaldosterone were detected after the conversion of their keto group to adicarbonyl group by the reaction with copper(II) acetate, and the derivatization of the resultant glyoxal compounds with DPH (Ishida et al, 1992c(Ishida et al, , 1993.…”

Determination of the pseudoephedrine content in pharmaceutical formulations and in biological fluids using a microbore HPLC system interfaced to a microfluidic chemiluminescence detector

“…This enables highly sensitive detection without the need for complex and expensive instrumentation. Due to its simplicity, low cost, high sensitivity and selectivity, CL detection has become an attractive tool in combination with many analytical techniques, such as HPLC (6)(7)(8)(9)(10)(11)(12)(13), capillary electrophoresis (CE) (14), flow injection analysis (16,17) and microfluidics (18)(19)(20)(21)(22)(23)(24).…”