Determination of citric acid in urine and in serum.

Abstract

A method is described for the determination of citric acid in the routine clinical investigation of urine and serum. It is based on a combination of the methods of GEY, LANE and CHEN. The method has been used for several years and has been completed by a statistical analysis. Es wird die Beschreibung einer Zitronensaurebestimmungsmethode im Harn und Serum, die sich gut zu Reihenuntersuchungen in der Klinik eignet, gegeben. Sie beruht auf der Kombination der Methoden vonGEY und LANE und CHEN. Wahrend mehrerer Jahre wurde diese Methode angewendet und durch eine Statistik erganzt. Recent progress in clinical chemical methods has shown the importance of citric acid in the different metabolic pathways. Its storage and release is influenced by vitamin D and parathormone, and it is a metabolite of the Krebs cycle. Citric acid is also a bone component. Moreover, the secretion and tubular reabsorption of renal citric acid are two problems which remain unsolved. Determinations of serum and urinary citric acid are thus not only of interest in the different bone diseases, but also in certain kidney and liver diseases. The clinical laboratory therefore needs a method for the determination of citric acid in urine and in serum, which is precise, rapid and not too complicated. This paper describes such a method which we have used in our laboratory. Our six years of experience have shown it to be capable of fulfilling all the necessary requirements of the clinical laboratory. Above all, we would like to emphasize its accuracy which permits, for example, the differentiation of certain diseases by analysis of the citric acid levels in serum. We have found many methods for the determination of citric acid in the literature, most of them based on the conversion of citric acid into pentabromacetone (Pba), with various modifications for its oxidation and extraction (2—12). We have retained the methods of GEY, and LANE and CHEN. After trying both, we decided that a judicious combination of the two would permit us to shorten the time required for the determination without sacrificing its sensitivity. We therefore kept LANE et aFs sequence of oxidation at low temperature and the subsequent reduction of the excess permanganate by hydrogen peroxide. We then substituted GEY'S technique for extraction with petroleum ether, elimination of traces of acid, and formation of the Pba-thiourea-borax complex. Thus the combination of the two methods permits us to shorten the time required for the determination without affecting the accuracy of the results. By adopting GEY'S method of extraction, we have eliminated certain inherent errors of the Nal-ethanol extraction, for example the influence of light, temperature and the possible contamination of theNal-Pba-ethanol mixture by acids, which would have to be eliminated by washing with water and the phosphate buffer. Furthermore, the Pba-thioureaborax complex is not affected by the presence of s-hydroxybutyric acid, acetone and succinic acid, even in pathological quantities. The Pba-Nal-ethanol reaction is sensitive to the aforementioned compounds, as well as to aspartic acid.