An enzymatic assay of inorganic phosphate in serum using nucleoside phosphorylase and xanthine oxidase

Abstract

We have developed a new enzymatic assay for the determination of inorganic phosphate (Pi) in serum, using nucleoside phosphorylase (NP) and xanthine oxidase (XOD). Pi and inosine react to form hypoxanthine and ribose-1-phosphate. The hypoxanthine is oxidized to xanthine, which is further oxidized to uric acid. In these two reactions 2,6-dichlorophenol-indophenol (DCIP) is reduced to a colourless compound and the decrease in colour is measured spectrophotometrically at 600 nm. The assay is automated with an RA-XT analyser. The precision of the automated assay is acceptable (C.V. < 3.5%) and results are accurate and linear across a range of values from 0.2-2.5 mmol/l. The assay correlates well with molybdate methods carried out on SMAC III and RA-XT analysers (r values 0.99 and 0.98, respectively), and seems to be less prone to non-specific sample interference than the usual RA-XT method. The enzymatic assay described seems to be suitable for the routine determination of serum Pi.