A method for preventing sorbitol interference with the determination of inorganic phosphate

✍ Scribed by Roger A. Leigh; Robert R. Walker

• Publisher: Elsevier Science
• Year: 1980
• Tongue: English
• Weight: 472 KB
• Volume: 106
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(80)90149-9

✦ Synopsis

A technique is described for preventing interference of sorbitol with the assay of P, by modifying the procedure of B. N. Ames (1966, in Methods in Enzymology, E. F. Neufeld and V. Ginsburg, eds., Vol. 8, pp. 115-118, Academic Press, New York). The new method relies on the ability of precipitated protein to bind phosphomolybdate and so allow separation of the Pi from the soluble sorbitol. The conditions for the formation and precipitation of phosphomolybdate-protein complex and for the subsequent assay of Pi are described. No unique set of conditions could be found which prevented interference at all sorbitol concentrations tested. Instead, conditions for the elimination of interference by particular sorbitol concentration ranges were established. The application of the procedure to samples containing 0-150 nmol of Pi and lo-100 pmol of sorbitol is described. Complete recovery of Pi was achieved after precipitation. Standard plots were linear. Coefficients of variation ranged from 9% with low amounts of Pi (525 nmol) to 2.5% at higher levels (150 nmol). One hundred nanomoles of Pi gave an absorbance at 700 nm of 0.87. Modifications are described to extend the techniaue to different sorbitol concentration ranges and other applications of the method are mentioned.

MATERIALS AND METHODS

Common

laboratory reagents, which were of the highest purity available, were obtained from B.D.H. Chemicals Ltd.