Validation of a column liquid chromatographic method for phytate

A column liquid chromatographic method that uses a spectrophotometric detector for the analysis of phytate and other inositol phosphates in foodstuffs and seeds is described. It has been tested thoroughly and sent to 6 other laboratories where there was an interest in such an analytical method and the equipment was available for performing it. These samples were blinded to the individual collaborators. Also sent was a standard wheat bran sample available from the American Association of Cereal Chemists with a standard phytate value reported as 3.2%. The postcolumn sulfosalicylic acid reaction was developed in the 1950s as an analytical method for ferric ion. All reagents are aqueous solutions and the specific pH values of each are important. The column separation is at pH 4, which dissociates all phosphate analogs. The ferric sulfosalicylate is at pH 1.8. The postcolumn reaction is then at pH 2.0-2.3. At this pH, the sulfosalicylate complexes one ferric ion and will yield it to a stronger complexing agent such as a phosphate ester. At higher pH values, sulfosalicylate complexes 2 ferric ions and will yield neither to these anions. The method sensitivity is 8-12 nmoles/injection.