The
cholesteryl ester transfer protein modulator
dalcetrapib is currently under development for the prevention of
dyslipidemia and
cardiovascular disease.
Dalcetrapib, a thioester, is rapidly hydrolyzed in vivo to the corresponding
thiophenol which in turn is further oxidized to the dimer and mixed
disulfides (where the
thiophenol binds to
peptides,
proteins and other endogenous
thiols). These forms co-exist in an oxidation-reduction equilibrium via the
thiol and cannot be stabilized without influencing the equilibrium, hence specific determination of individual components, i.e., in order to distinguish between the free
thiol, the
disulfide dimer and mixed
disulfide adducts, was not pursued for routine analysis. The individual forms were quantified collectively as
dalcetrapib-
thiol (dal-
thiol) after reduction under basic conditions with
dithiothreitol to break
disulfide bonds and derivatization with
N-ethylmaleimide to stabilize the free
thiol. The S-methyl and S-
glucuronide metabolites were determined simultaneously with dal-
thiol with no effect from the derivatization procedure. Column-switching liquid chromatography-tandem mass spectrometry provided a simple, fast and robust method for analysis of human and animal plasma and human urine samples. Addition of the
surfactant Tween 80 to urine prevented adsorptive compound loss. The lower limits of quantitation (LLOQ) were 5 ng/mL for dal-
thiol, and 5 ng/mL for the S-methyl and 50 ng/mL for the S-
glucuronide metabolites. Using stable
isotope-labeled internal standards, inter- and intra-assay precisions were each <15% (<20% at LLOQ) and accuracy was between 85 and 115%. Recovery was close to 100%, and no significant matrix effect was observed.