IN-VITRO INHIBITION OF ALKALINE-PHOSPHATASE ACTIVITIES FROM INTESTINE, BONE, LIVER, AND KIDNEY BY PHENOBARBITAL

A kinetic study of the inhibition of several alkaline phosphatase (AP) isoenzyme activities by phenobarbital was carried out using p-nitrophenylphosphate (10 mM as a substrate at pH 9.8 in a 300-mM Hepes buffer. AP from bovine kidney, calf intestine, bovine liver, and rat bone was used. Over a phenobarbital concentration range of 20-400 mM, all these isoenzymes were inhibited in an uncompetitive manner with a K(i) of 200 mM for intestinal AP, and in a linear mixed-type manner for all the other isoenzymes tested. The K(i) values were 10, 40 and 55 mM for kidney, bone and liver AP, respectively. The use of 15 mM carbonate-bicarbonate or 400 mM diethanolamine buffer did not modify the degree of inhibition of intestinal AP activity. Dixon plots of the reciprocal of reaction velocity versus inhibitor concentration either at different substrate concentration or at different DEA concentration indicate uncompetitive inhibition for the intestinal enzyme. This in vitro inhibitory effect of phenobarbital is in contrast to its in vivo stimulating action on AP. However, in the whole animal, the effects of phenobarbital administration probably represent the sum of multiple effects.