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Vol. 282, Issue 1, 391-396, 1997
Divisions of Clinical Pharmacology and Biopharmaceutics, Center for
Drug Evaluation and Research, Food and Drug Administration, Rockville,
Maryland
The effect of bovine serum albumin (BSA) on human liver metabolism,
in vitro, of 14C-phenytoin (PHT) was
studied. Michaelis Menten parameters were determined for the conversion
of PHT to p-hydroxy phenytoin in seven different microsomal
preparations with the addition of 0, 2, and 4% BSA. The unbound
Km
(Kmu) values were 30.8 ± 18.6, 1.57 ± 0.21 and 1.50 ± 0.17 µM (mean ± S.D.),
respectively; however, there was excellent agreement among the
Vmax values (29.1, 31.8 and 31.5 pmol/min/mg). With intact
tissue slices, BSA (4%) added to incubations of PHT had a minimal
effect on the Vmax values in two of the four livers studied
and resulted in a mean Kmu value of 2.20 ± 0.59 µM, although the
Kmu in the absence of BSA was 6.64 ± 3.17. In scaling-up to the whole body, Vmax
values were 3.9 and 1.0 mg/kg/day for microsomes and slices,
respectively, compared to 5.9 mg/kg/day, in vivo. The
Kmu values determined in the
presence of albumin in both microsomes and slices were similar to those
based on in vivo human steady state data
(Kmu = 2-3 µM), and the
intersubject variation, in vitro, was decreased in the
presence of BSA. These findings for phenytoin metabolism suggest that
the addition of albumin to incubation media for slices or microsome
experiments may yield Km estimates that are
more representative of in vivo values.
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