An assay for inorganic orthophosphate in the presence of alkaloids and related drugs

Abstract

There have been many reported techniques for the determination of inorganic orthophosphate, largely utilising the formation of a yellow complex between the phosphate and a molybdenum salt, and the formation of a blue chromophore on reduction of the complex. The method of Fiske and Subbarow (1) employs l-amino-Znaphthol-Csulphonic acid as the reducing agent, and this technique is still widely used. Modifications to the basic procedure have included the direct measurement of the yellow phosphomolybdate complex to avoid reduction of susceptible compounds (2) and the extraction of the phosphomolybdate complex with organic solvents to prevent interference by inorganic ions (3, 4). Atkinson et al. (5), reported that incorporation of the non-ionic detergent Cirrasol ALN-WF, formerly called Lubrol-W, into the acid ammonium molybdate reagent prevented the interference in the phosphate assay by a number of simple compounds that adversely affect the conventional methods. Our studies on the effects of certain pharmacological agents on the active transport of calcium in muscle tissue, have led us to look for a reliable and simple method of determining levels of inorganic phosphate in the presence of alkaloids and related methyl xanthines, since these compounds form a white precipitate in the presence of phosphate and acid ammonium molybdate which renders the conventional assay inoperable.