Enzymic assay for serum theophylline.

We describe an original procedure for determination of theophylline in serum. The drug is extracted from 0.4 mL of serum at pH 7.4 with chloroform/isopropanol (20/1 by vol) and back-extracted into sodium hydroxide (1 mmol/L). The inhibition of beef-liver alkaline phosphatases by theophylline in this alkaline phase is measured at 25 degrees C, with p-nitrophenyl phosphate as substrate, in 2-amino-2-methyl-1-propanol buffer, pH 9.4- The reciprocal of enzyme activity and theophylline concentration are linearly related in the range 2 to 60 mg/L. The maximum interference to be expected from 3-methylxanthine would increase apparent theophylline concentration by no more than 1 mg/L. The method is accurate, free of interference by other xanthines and often-coadministered drugs, and results correlate well with those by the immunoenzymic assay. Major advantages are reagent stability, low cost, and simplicity of instrumentation.