A population approach to enzyme characterization and identification: application to phenacetin O-deethylation.
OBJECTIVE
To determine the enzyme kinetics (EK) and identify the human cytochrome(s) P450 (CYP) involved in the deethylation of phenacetin to acetaminophen using a population-based method.


METHODS
A sparse data set was generated from incubations containing human liver microsomes (n = 19) with phenacetin. Estimates of the EK parameters were obtained by fitting the concentration-velocity data to Michaelis-Menten models by using nonlinear mixed effects modeling. Relationships between the EK parameters and the CYP activities determined for these liver microsomes were examined.


RESULTS
A two-enzyme kinetic model with a saturated, low KM enzyme and an unsaturated, high KM enzyme capable of forming acetaminophen best fit the data. The population estimates of the EK parameters were Vmax1, 911 pmol/min/mg protein; KM1, 11.3 microM; and Cl(int2), 0.4 microl/min/mg. The coefficients of variation for interliver variability in Vmax1 and residual error of the model were 39% and 15%, respectively. When the selective catalytic activities were examined as potential covariates, 7-ethoxyresorufin O-deethylation (CYP1A2) activity was found to be associated with the low KM enzyme, however, the high KM enzyme(s) could not be identified.


CONCLUSIONS
The population approach characterized the EK parameters and identified the low KM enzyme responsible for phenacetin O-deethylation as CYP1A2. Population modeling of EK provides valuable information on inter- and intraliver variability in CYP dependent activities.
Belle
D J
DJ
Drug Disposition, Eli Lilly and Company, Indianapolis, IN 46285, USA.
Ring
B J
BJ
Allerheiligen
S R
SR
Heathman
M A
MA
O'Brien
L M
LM
Sinha
V
V
Roskos
L K
LK
Wrighton
S A
SA
D000465:Algorithms; D018712:Analgesics, Non-Narcotic / Q000378:metabolism*; D001711:Biotransformation; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D003640:Dealkylation; D006801:Humans; D066298:In Vitro Techniques; D008862:Microsomes, Liver / Q000378:metabolism; D008954:Models, Biological; D010615:Phenacetin / Q000378:metabolism*; D011153:Population
D018712:Analgesics, Non-Narcotic; D003577:Cytochrome P-450 Enzyme System; D010615:Phenacetin