King-Armstrong Phosphatase Estimation by the Determination of Liberated Phosphate.

As a result of many requests for a phosphatase method which would use the phenyl phosphate substrate of King and Armstrong (1934), but which would substitute a determination of the liberated inorganic phosphate for that of phenol, we have developed two procedures which are described in this paper. The principal reason for a method in which the final measurement is of inorganic phosphate is that an estimation of the plasma phosphate is often required in the study of children's diseases; and it is thought to be desirable to combine this estimation with the phosphatase determination, as may easily be done by making the inorganic phosphate estimation identical with the control for the phosphatase. This is done in some of the glycerophosphate procedures, such as those of Jenner and Kay (1932) and Bodansky (1933). But the use of phenol phosphate for phosphatase determination is greatly to be preferred, because of the greater convenience of the procedure, the much shorter time which is necessary to complete the determination, and the small amounts of blood plasma or serum required. Two procedures are set out below: in one the determination of liberated phosphate is by the molybdate method of Fiske and Subbarow (1925) with aminonaphtholsulphonic acid as reducing agent; in the other, a more micro procedure, a molybdatestannous chloride method is used. Both procedures employ the same substrate, buffer, temperature, and hydrolysis time as the phenyl phosphate method in which phenol is determined, so that the conditions are identical with those of the KingArmstrong method as modified by King (1951). The results are identical in theory, since one molecule each of phenol and phosphate are liberated for each molecule of phenyl phosphate hydrolysed, and the comparisons of results obtained by the different procedures has, in fact, shown them to be in excellent agreement.