Direct determination of ibuprofen and ibuprofen acyl glucuronide in plasma by high-performance liquid chromatography...

A method for the simultaneous determination of ibuprofen and its labile, reactive metabolite, ibuprofen acyl glucuronide, in plasma is described. Reversed-phase high-performance liquid chromatography (HPLC) employed a C18 column using methanol-10 mM trifluoroacetic acid as the mobile phase with ultraviolet detection at 225 or 214 nm. It is essential that blood is handled rapidly and plasma is acidified upon collection prior to freezing. Plasma samples first are deproteinated with acetonitrile, the supernatant is diluted with phosphate buffer, and ibuprofen, ibuprofen glucuronide, and ibufenac (internal standard) are initially isolated by solid-phase extraction on C18 cartridges. Upon elution, the residue is evaporated, dissolved and injected onto the HPLC system. Recovery is 94 +/- 8 and 70 +/- 9% for ibuprofen glucuronide and ibuprofen, respectively. The measurable concentration range is linear from 0.1 to 10 micrograms/ml for ibuprofen glucuronide and from 0.5 to 100 micrograms/ml for ibuprofen. The method is satisfactory for the analysis of ibuprofen and ibuprofen glucuronide from pharmacokinetic studies in humans. The direct determination of ibuprofen glucuronide allows accurate measurement of this conjugate at low levels relative to the parent compound, ibuprofen, a distinct advantage compared to previously employed indirect methods.