Cxlvi. the Determination of Pyruvic Acid. by Basettihalli Hanumantha Rao Krishna and Motnahalli Sreenivasaya

THE methods employed for the estimation of pyruvic acid are largely based upon the reaction of its carbonyl group with phenylhydrazine. Its estimation in complex biological fluids where there are other compounds which react likewise is therefore difficult and, particularly when pyruvic acid exists in small quantities amidst these compounds, the estimation is unreliable even when controls are run. This difficulty was greatly felt in some of our investigations, and it was thought worth while to study some of the existing methods which could be satisfactorily employed as such or modified for our purpose. The production of an intense blue colour by sodium nitroprusside and ammonia, especially in the presence of a little acetic acid, has been claimed to be specific for pyruvic acid and unaffected by acetaldehyde. The precipitation of pyruvic acid as hydrazone by excess of phenylhydrazine followed by the determination of the excess phenylhydrazine has been the subject of considerable study [Smedley-MacLean, 1913; Simon and Piaux, 1924]. The reduction of pyruvic to lactic acid by zinc and hydrochloric acid and the subsequent estimation of lactic acid according to the method of Fiirth and Charnass has been adopted by Lieben [1923]. The oxidation of pyruvic to acetic acid by a slight excess of hydrogen peroxide has been shown to be quantitative by Holleman [1904], and potassium dichromate has been employed similarly and the excess dichromate estimated. As it is not the purpose of this paper to give the results of the comparative study of these methods, it will be sufficient to say that none of them as such was found suitable in our experiments, and all required some modifications before they could be employed. The necessary conditions for a method which would be suitable to our requirements were that it should be applicable to small quantities of the acid, usually between 0-5 and 15 mg., and in solutions of very low concentration, i.e. 01% to 0-05 %. The method finally developed is a modification of Lieben's technique, and is based upon the following reactions: CH3. CO.COOH + 2H = CH3.CHOH. COOH ...... (i). CH3.CHOH.COOH + 0 = CH3.CHO + CO2 + H20 ...... (ii).