methods of analyses examined. For the SNLR and KM,app methods there are three values of Ki listed under the column headings, Fixed and Varying and the resulting means and S.D.s of those values. The term “fixed” indicates that the control data were analyzed by nonlin- ear regression to obtain the best fitting values for VMAX and KM.
Those values were then fixed in the subsequent SNLR analysis of all of the data (i.e., control and inhibition data). The term “varying” indicates that values for VMAX and KM were obtained from the simultaneous fit of all of the data (including the control experiment) and there was no attempt made to fix the values.
Each of the Ki values listed was obtained from the analysis of a complete data set that included triplicate determinations of metabolite formation rates per substrate concentration. Thus, the values 4.71, 4.60, and 4.92 (under SNLR, Fixed column) are Ki estimates obtained from three different data sets, each involving triplicate estimations of rate versus substrate concentrations for a control and five inhibitor concentrations. Furthermore, the table is divided into rows according to the number of inhibitor concentrations used in the analysis. As an example, 4.74 ( 0.16) and 4.95 ( 0.27) are the means and S.D.s for Ki when the SNLR method is used to analyze the data from five
inhibitor concentrations and one control experiment and when VMAX and KM are either fixed or allowed to vary, respectively.
Considering the SNLR results, the mean estimates of Ki are not very different from the perfect value of 5.0; regardless of whether VMAX and KM were fixed or allowed to vary. Furthermore, the values for Ki do not vary greatly from the correct number as one moves down the table; that is, as fewer inhibition curves are generated. The S.D.s, however, do increase as the total number of experiments are reduced. Thus, using only the control experiment and one inhibition curve, the mean Ki value is 4.71 (range, 3.91–5.61) with a coefficient of varia- tion of 18%. The latter range and variation are considerably greater than those associated with estimates of Ki that rely upon more inhi- bition curves.
The column designated KM,app is divided into two components, as with the SNLR method, one in which the value for VMAX was fixed