I am working on total phytate quantification using the UV-Vis spectrophotometer, and we prepared the phytate standard solutions (0-200 μg/ml phytic acid in deionized water). There was no difference in the standard colors upon adding the wade reagent, then we centrifuged it for 5 min at 4000 rpm. We measure the absorbance at 500 nm wavelength, but there were no differences in the absorbances between the different concentrations which we do not understand why it failed since all our solutions were freshly prepared.
Phytates have no specific absorption spectra. That's why you have seen no differences in color and absorbance. You need to use indirect methods (links to articles below) or commercial kits (as above).
Links to literature:
Article Fluorescence Sensing of Phytate in Water Using an Isothiouro...
Article Analytical Methods for Determination of Phytic Acid and Othe...
Article A turn-on fluorescent probe for phytic acid based on ferric ...
Links to exemplary commrecial kits:
https://www.biorbyt.com/phytic-acid-microplate-assay-k-orb707384.html
https://www.abbexa.com/phytic-acid-assay-kit
https://www.megazyme.com/phytic-acid-assay-kit
My understanding is that the Wade method should work... (the color of iron sulfosalicylate complex should fade in the presence of phytate). There may be some miscalculation or measuring error involved, for example an order of magnitude error. It might help if you show the actual absorption spectra before and after addition of phytate and supply quantitative data (what volume of what concentration was reacted with what volume of what concentration?).
- Badges
- Science method