Assay methodology for quantification of the ester and ether glucuronide conjugates of diflunisal in human urine

Diflunisal is a salicylate derivative with analgesic and anti-inflammatory properties. It is excreted in the urine as an ether glucuronide, a 1-O-acyl glucuronide and as unchanged drug. The 1-O-acyl glucuronide rearranges to isomeric esters of glucuronic acid under neutral to alkaline pH conditions. The development of a urine assay for the conjugates enables the elucidation of diflunisal non-linear pharmacokinetics. The assay quantitates the ether and ester glucuronides and free diflunisal in urine at 0.5-1.0 micrograms/ml. Analysis of the glucuronides does not require authentic standards.