T. Izumi et al., DETERMINATION OF PROTEIN-BINDING OF TROGLITAZONE STEREOISOMERS BY FLUOROMETRIC TITRATION, Biopharmaceutics & drug disposition, 18(3), 1997, pp. 241-257
Determination of the protein binding of troglitazone is difficult beca
use of its high adsorption to filters and membranes and the instabilit
y of the stereoisomers. We attempted to assess the protein binding of
four stereoisomers of troglitazone in the plasma and albumin from seve
ral species by the method using fluorescent probes. The inhibition con
stants (K-i) for the stereoisomers of troglitazone were obtained from
the decreases in fluorescence intensity of dansylsarcosine caused by c
ompetitive inhibition. Each stereoisomer of troglitazone displaced dan
sylsarcosine, a typical specific fluorescent probe for the diazepam bi
nding site on human serum albumin (HSA). The highest binding affinity
for dansylsarcosine was observed with HSA (dissociation constant, K-d,
K-1=0.5 mu M), while it was lowest in the mouse (K-d,K-1=18 mu M) The
K-i values for KK and ddY mouse plasma and mouse and rat albumin were
in the range 2-15 mu M, and there were no large variations among stere
oisomers, the maximum differences being twofold, For human plasma and
albumin, the displacement could not be accounted for by a simple compe
titive inhibition. Comparison between unbound fraction (f(u)) values c
alculated from thus obtained K-i values and those Df a mixture of the
four stereoisomers determined by high-performance frontal analysis sho
wed that the f(u) values obtained by fluorometric titration were highe
r, while the relative differences among the stereoisomers in terms of
animal species and strain were comparable for the two methods, Small d
ifferences in protein binding among stereoisomers of troglitazone may
not be the major reason for their stereoselective pharmacokinetics. (C
) 1997 by John Wiley & Sons, Ltd.