Diastereoisomeric resolution of a pronucleotide using solid phase extraction and high performance liquid
C Foulon1, C Vaccher, A-L Villard
1Laboratoire de Chimie Analytique, EA 4034, Faculté des Sciences Pharmaceutiques et Biologiques, Université de LILLE 2, BP 83, 3 rue du Pr. Laguesse, 59006 Lille Cedex, France.
Abstract:
A stereospecific HPLC methodology has been developed for the diastereoisomeric resolution of a mononucleotide prodrug in cell extracts. This method involves the use of solid phase extraction on a C18 cartridge. Diastereoisomers and internal standard resolutions were performed on a cellulose based chiral column (Chiralcel OD-H) used in the normal phase mode. The method was validated in terms of specificity, recovery, linearity (diasteroisomers mixture concentration: 3-60 micromol L(-1)), precision and accuracy and detection limit (1.67 and 1.33 micromol L(-1) for first and second eluted diastereoisomer). This method was applied to the determination of the apparent rate constants of disappearance and half-lives of each stereoisomers. This permits to conclude to the stereoselectivity of the enzymatic activity involved in the decomposition pathway of 2.
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