Purification of Synthetic Bilirubin Glucuronides By Reversephase Hplc Followed By on Column Solvent Exchange: Purification and stabilization of synthetic bilirubin glucuronides for in vitro assays

Abstract

Impaired bilirubin conjugation, caused by inhibition of UGT1A1, canresult in clinical consequences, including jaundice and kernicterus. Thus,Evaluation of the ability of new drug candidates to inhibit UGT1A1- Catalysedbilirubin glucuronidation in vitro has become common Practice. However, theinstability of bilirubin and its glucuronides presents substantial technicalchallenges to conduct in vitro bilirubin glucuronidation assays. Furthermore,because bilirubin can be diglucuronidated through a sequential reaction,establishment of initial rate conditions can be problematic. To conduct thesestudies need high pure Bilirubin glucuronides. Purification and postpurification concentration of synthesized bilirubin glucuronides is often themost difficult part of the synthetic procedure. In this study we describe thepurification of synthetic bilirubin glucuronides by reverse phase HPLC,minimizing and reducing expose to light and temperature of pure fraction by oncolumn solvent exchange process.