Some observations on the role of copper ions in the reduction of phosphomolybdate by ascorbic acid and their application in the determination of inorganic orthophosphate

The reduction of molybdate in the presence of phosphate gives rise to a blue compound, and under suitable conditions the colour intensity is propor- tional to the amount of orthophosphate present. This principle forms the basis of several methods which have been described for the determination of phosphate and a variety of reducing agents has been used for this purpose. In the method of Lowry & Lopez (1946) ascorbic acid is used at pH 4, in contrast to the strongly acid conditions used in most other methods, and in addition a relatively low molybdate concentration is employed. This reduces the breakdown of labile phosphates such as creatine phosphate, acetyl phosphate andribose 1-phosphate, which is otherwise very rapid (Fiske & Subbarow, 1929; Lipmann & Tuttle, 1944; Lowry & Lopez, 1946), to such an extent that it can be corrected for. This method thus permits the determination of inorganic phosphate in the presence of labile phosphate esters.