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A colorimetric procedure is described for determination of orthophosphate (0.2-2.5 nmol) in sample volumes up to 400 microliters. Orthophosphate is selectively extracted (in the form of phosphomolybdate) into an organic solvent mixture (2-methylpropan-1-ol and petroleum spirit) leaving interfering substances, such as labile organic phosphates, in the aqueous phase. Orthophosphate is then back-extracted into a small volume of aqueous sodium hydroxide. By keeping this volume small, orthophosphate from large dilute samples can be concentrated into small volumes and assayed colorimetrically in microcuvettes using the dye malachite green. The procedure is highly reproducible and insensitive to interfering substances, as shown by comparison with a conventional malachite green assay without the solvent extraction.

Original publication

DOI

10.1016/0003-2697(89)90405-3

Type

Journal article

Journal

Anal biochem

Publication Date

15/08/1989

Volume

181

Pages

130 - 134

Keywords

Animals, Cell Extracts, Cells, Cultured, Chemical Phenomena, Chemistry, Colorimetry, Mammals, Phosphates, Rosaniline Dyes, Sodium Hydroxide