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In a previous study, we determined the in vivo binding parameters of valproic acid (VPA) to serum proteins in seven healthy young adults at steady state by using the Scatchard equation. To evaluate the ability of the Scatchard binding equation to predict steady-state unbound serum VPA concentrations (Cf), 39 adult patients receiving VPA monotherapy and ranging in age from 16 to 68 years were studied. The correlation between predicted and observed Cf was high (r = 0.865). Mean prediction error, mean absolute error (MAE), and root mean squared error (RMSE) were calculated, and served as a measure of prediction bias and precision. The MAE and RMSE were low (MAE = 12.9 mumol/L, RMSE = 17.7 mumol/L). It is feasible to use the Scatchard binding equation to predict Cf in patients receiving VPA monotherapy.

Two hundred milligrams of valproic acid (VPA) was administered orally to seven healthy adults at 9:00 and 21:00 h for 5 consecutive days, including the morning dose on day 6. On the sixth day, blood samples were drawn at 0, 0.17, 0.33, 0.5, 0.75, 1, 1.5, 3, and 6 h after the morning dose. Binding of VPA to serum protein was evaluated by ultrafiltration, and total and unbound VPA concentrations were determined by fluorescence polarization immunoassay. Binding parameters of VPA to serum protein were calculated for each subject by the Scatchard analysis. The binding parameters obtained from seven subjects showed that the mean association constant (K) was 2.72 x 10(4) L/mol while the total number of binding sites (nPt) was 789 mumol/L. There were marked interindividual variations and the coefficient of variation was 42% for K and 28% for nPt. These results show that endogenous free fatty acids (FFAs) significantly reduce the binding affinity of VPA to serum albumin (p less than 0.05). In addition, they suggest the possibility that the primary binding sites for VPA can be strongly reduced by FFAs. Therefore, we consider that interindividual differences in binding parameters may be clinically important.

The interindividual relationship between unbound fraction of valproic acid and some factors such as total drug concentration, level of albumin, free fatty acids (FFA) and total bilirubin was investigated after repeated oral doses in 7 healthy adults. All serum samples (n=63) were collected just prior to the last dose and the following times there-after.The correlation between unbound fraction and both corresponding each variable value and ratio of FFA/albumin and total bilirubin/albumin was determined in all samples (method (1)). Secondly, the correlation between individual average unbound fraction and both average values for each variable and ratio of FFA/albumin and total bilirubin/albumin of individual was also determined (method (2)).For applied method (1), there was significant positive correlation between unbound fraction and both total drug concentration (P

Valproic acid is highly bound to serum protein, mainly albumin. Valproic acid exhibits concentration-dependent protein binding, which partly accounts for considerable variability in the unbound fraction, even within the therapeutic range. Therefore, valproic acidprotein interaction and determination of the corresponding binding parameters should be considered. In the present study, we investigated the binding characteristics in epileptic adults and children under valproic acid chronic monotherapy, and determined the binding parameters of valproic acid to serum protein.There was a curvilinear relationship between total and unbound concentrations orunbound fraction, using a second-degree polynomial equation. The unbound fraction increased curvilinearly in proportion to the increase in total concentration . Averages of unbound fraction obtained from adults and children were not significantly different from each other (P>0.05). The unbound fraction ranged from 4.1 to 16 .7% (adults), and from 5.6 to 16.7% (children). There were marked about three-fold variations of unbound fraction in both adults and children.On the other hand, as the Scatchard plot was shown to be linear in both adults and children, valproic acid was bound to only one class of binding sites on the protein molecule. The binding parameters calculated by Scatchard plot were as follows: association constants (Ka) were 1. 14×104l/mol (adults) and 1.44×104l/mol (children), total number of binding sites (n (Pt)) was 1.36×10-3mol/l (adults) and 1. 20×10-3mol/l (children). There was no significant difference in the two parameters between adults and children (P>0 .05). Therefore, we determined the binding parameters from all samples (n=96) by using Langmuir plot. These results showed that dissociation constant (Kd) was 3 .89×10-5mol/l, total number of binding sites (n (Pt)) was 8.46×10-4 mol/l.These results suggested that valproic acid had the same binding characteristics in both adults and children. The molecular interaction between valproic acid and serum albumin involved one class of binding sites within the investigated range.