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Development and Partial-Validation of a High Throughput HPLC Method for the Quantification of Dexamethasone in Aqueous and Ocular Matrices-Application to a Pilot Pharmacokinetic Study

Abstract

A simple, sensitive and reliable analytical method for the rapid determination of dexamethasone in aqueous and ocular tissue samples by high performance liquid chromatography (HPLC) was developed and validated. Samples were prepared by simple protein precipitation (PP) technique followed by the addition of acidified phosphate buffer. Aqueous samples were loaded directly with appropriate dilution to be in the calibration curve range. Samples were analyzed on a ZORBAX Eclipse XDB-C18 column with a mixture of acetonitrile and 10 mM phosphate buffer containing 0.1% acetic acid as mobile phase in 29:71 ratio. Dexamethasone was quantified in ocular and aqueous samples using triamcinolone acetonide as an internal standard. The method was linear in the range of 0.1 to 10.0 μg/mL. The method is demonstrated to be suitable for the determination of dexamethasone in aqueous and all the ocular tissue samples. The total time required for the analysis of a single sample was about 14 min.

Srinivas Rao Chennamaneni