High-performance liquid chromatographic method for the determination of budesonide in bronchoalveolar lavage of asthmatic patients.

A simple, sensitive and selective method for the determination of budesonide in bronchoalveolar lavage (BAL) using high-performance liquid chromatography (HPLC) with UV detection was developed. BAL samples were extracted twice with methylene chloride, the extracts were centrifuged and the organic layer separated and dried under nitrogen. The samples were reconstituted in the mobile phase and 80 microliters were injected on to a Spherisorb ODS column with UV absorbance detection at 250 nm. The mobile phase was methanol-aqueous buffer (69:31, v/v). Inter-assay coefficients of variation were measured at 7.81 and 500 ng/ml with ranges of 0.89-7.31%. Average recoveries were 97% or greater. This method was successfully implemented for the analysis of BAL from asthmatics, in order to establish the amount of budesonide available to the lung and to investigate the efficacy of inhaler systems. Patients (n = 9) inhaled four puffs of 200 micrograms of budesonide and BAL was performed 10 min after the last inhalation. Only four BAL out of the nine presented detectable amounts of budesonide. The concentrations in BAL in these four patients were 13.44-84.18 ng/ml, corresponding to total amounts of 0.847-7.997 micrograms.