A large number of reports have appeared for the determination of keto acids by the DNP* method. The outline of the method is as follows: Keto acid in water solution is converted to DNPH derivative. Then the DNPH, after extraction with water-immiscible organic solvent, is transferred to water solution by shaking the organic solvent solution with carbonate solution to separate DNPH from the unreacted reagent. To the DNPH in the carbonate solution is added sodium hydroxide and the red color developed is measured spectrophotometrically
(1). However, with the sample which contains keto acid as a sole carbonyl compound, such an extraction procedure is unnecessary. In fact, Friedemann and Haugen (1) proposed a direct method for the determination of keto acid. According to their method, DNP reagent is added to the sample solution and the red color produced by the addition of alkali is measured spectrophotometrically.
The control value, which is obtained by using water instead of sample solution, is subtracted from the value obtained with the sample. However, their method did not always give satisfactory results for the determination of some kinds of a-keto acids, because of the neglect of two geometric isomers of their DNPH.
We solved these obstacles by using the previously reported spectrophotometric method, which is valid irrespective of the existence of both isomers of DNPH (2). This paper deals with improvement of the direct method for the determination of keto acids and presents the results of analysis of several kinds of keto acids.