Determination of caffeine in saliva by high-performance liquid chromatography: new sampling method for saliva using...

We report an assay method for the determination of salivary caffeine concentrations by high-performance liquid chromatography. A simple, new method for collecting mixed saliva using a piece of filter paper has been developed. An arbitrary amount of saliva is absorbed and a fixed amount of internal standard is added. The mean coefficient of variation of measurements repeated by changing both volumes absorbed and caffeine concentrations of saliva was 5.6% with this method. The detection limit was determined to be 0.5 microgram/ml. Precision of measurements and recoveries did not depend upon the volume of saliva absorbed. Four volunteers were given a capsule of 230 mg caffeine, and saliva was collected by the ordinary method using chemical stimulation with citric acid and by our filter paper absorption methods; the methods were compared. Salivary caffeine concentrations were always lower than plasma concentrations, and there were no big differences between two methods, although salivary concentrations measured by our method was higher, with a significant difference at 1.5 h after dosing. From the practical viewpoint this method would be convenient and sufficiently accurate.