Table 1.

Measured and predicted rates of P450-mediated NADPH oxidation and product formation catalyzed by simple and mixed systems.^a

Product or Substrate	CYP2B4 system	CYP1A2 system	Mixed system (Experimental results)	Predicted Contribution from CYP2B4 to mixed RS b	Predicted Contribution from CYP1A2 to mixed RS c	Mixed system (Theoretical) d
NADPH oxidized	12.7 ± 0.38	7.21 ± 0.23	14.14 ± 0.38	6.79	21.63	28.42
Toluene metabolites	2.61 ± 0.28	0.01 ± 0.002	1.51 ± 0.10	1.40	0.03	1.43
H2O2 formed	2.01 ± 0.63	1.20 ± 0.10	5.23 ± 0.86	1.08	3.60	4.68
Excess H2O formed	4.04 ± 1.29	3.00 ± 0.33	3.70 ± 1.34	2.16	9.00	11.16

^aThe experimentally measured rates of NADPH oxidation and formation of side-products and toluene metabolites by CYP2B4, CYP1A2, and mixed CYP2B4-CYP1A2 reconstituted systems were determined as described in Materials and Methods. The theoretical rates (expressed in nmol/min) of NADPH oxidation and the formation of uncoupled products by the individual enzymes in the mixed system were predicted by using the measured changes in the rates of formation of the P450-specific toluene metabolites by the mixed system as proportion factors for these other parameters. More specifically, Figure 1 indicates that mixing of CYP1A2 and CYP2B4 caused a 47% decrease in CYP2B4-mediated product (p-cresol) formation and a 3.75-fold increase in CYP1A2-mediated product (o-cresol) formation. To calculate the theoretical contribution of each enzyme in the mixed system, the rates of NADPH oxidation and uncoupled product formation by the simple systems were multiplied by 0.47 and 3.75 for CYP2B4^b- and CYP1A2^c-mediated metabolism, respectively. The theoretical rates of metabolism by the mixed system^d are simply the sum of the “predicted contributions” by each of the enzymes. The theoretical rates are calculated from the averages of the measured rates of metabolism and do not include margins of error.