Improving the stability of the malachite green method for the determination of phosphate using Pluronic F68.

Improving the stability of the malachite green method for the determination of phosphate using Pluronic F68. Anal Biochem. 2020 Mar 09;:113681 Authors: Martínez Gache SA, Recoulat Angelini AA, Sabeckis ML, González Flecha FL Abstract The analytical determination of phosphate constitutes a fundamental step for the evaluation of catalytic activities of many enzymes involved in the hydrolysis of phosphate-containing biomolecules. The most sensitive colorimetric methods to quantify phosphate are based on measuring the spectral changes produced by the adsorption of malachite green to 12-molybdophosphoric acid. Malachite green methods are time-consuming because they require the preparation of color reagent on the day of use, due to its low stability. In this work we propose a modification of the malachite green method that overcomes these problems and only requires a one-step, ready-to-use stock solution including perchloric acid and Pluronic F68. The improved reaction mixture allowed the quantification of phosphate with a limit of detection of 0.22 μM, a dynamical range up to 80 μM depending on the optical path-length, and a molar absorption coefficient at 640 nm of 94000 M-1cm-1. Color development reaches a steady level within 30 min and remains constant for at least 2 h. The high stability of the color reagent allows long-term storage for at least 1 year. The optimized procedure is especially useful to measure phosphate-containing b...
Source: Analytical Biochemistry - Category: Biochemistry Authors: Tags: Anal Biochem Source Type: research
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