The catalytic effect of molybdate on the hydrolysis of organic phosphate bonds.

The great instability of creatine phosphate (Fiske & Subbarow, 1929), acetyl phosphate (Lipmann & Tuttle, 1944) and 1:3-diphosphoglyceric acid (Negelein & Bromel, 1939) in solutions containing molybdate has been known almost as long as these compounds themselves. It could not fail to attract the attention of the analyst who used a method of phosphate estimation ofthe Briggs (1922) or Fiske & Subbarow (1925) type, because these methods entail a period of colour development duringwhich the organic phosphate is in contact with ammonium molybdate concentrations of 0-2-0-7 %. The acid-lability ofaphosphate bond is apparently not invariably linked with the catalytic effect of molybdate. For ribose-1-phosphate, for example, which is intermediate in acid-stability between creatine phosphate and acetyl phosphate, Kalckar (1947) found, in dilute acids, only about 30% increase of the hydrolysis rate in presence of molybdate, and Meyerhof & Lohmann (1928) even made the curious observation that the hydrolysis of arginine phosphate in acid solution was actually inhibited in presence of molybdate. It is perhaps surprising that, apart from these scattered observations, no systematic study of the molybdate effect has yet been made. To some extent this may be due to the fact that with the older methods of phosphate estimation a tedious separation of organic and inorganic phosphate was necessary for its demonstration. In the extraction method of phosphate estimation introduced by Berenblum & Chain (1938) the time of contact between molybdate and phosphate before completion of the phosphomolybdate extraction can be reduced to about 15 sec. This time interval is sufficiently short to reduce the interaction between molybdate and all but the most labile phosphate bonds to insignificant proportions. The method of Berenblum & Chain is, however, somewhat laborious and unsuited to the handling of large series of analyses. The recent modification of Martin & Doty (1949) has greatly simplified it, and with the adaptations which we used in the present study the method is as easy and expeditious to carry out as the method of Fiske & Subbarow without sacrificing any of the advantages of Berenblum & Chain's method. METHODS