A simplified photometric method for the determination of citric acid in biological fluids.

Studies on the oxidation and bromination of citric acid to pentabromoacetone were reported as early as 1847 by Cahours (1). This reaction has since formed bhe basis for many of the procedures for determining citric acid. A qualitative test by Stahre (2), reported in 1897, was adapted fol quantitative analysis by Kunz (3). Pucher, Sherman, and Vickery (4) described the first microphotometric method, using the reaction of pentabromoacetone with sodium sulfide for color development. Their procedure, measuring 0.1 and 1.0 mg. of citric acid, was later discarded in favor of a titrimetric method (5). More recent photometric modifications include those of Taussky and Shorr (6), color development with sodium iodide, Wolcott and Boyer (7), potassium iodide, and Xatelson, Pincus, and Lugovoy (8), thiourea. Attempts to apply these methods to the measurement of micro quantities of citric acid have prompted search for a technique having greater sensitivity and simplicity. This report describes a simplification of the procedure for the oxidation and bromination of citric acid to pentabrozloacetone. The reaction ‘of trihalogenated hydrocarbons with alkaline pyridine, described by Fujiwara (9), has been adapted to the measurement of the pentabromoacetone. The resulting photometric procedure measures amounts of citric acid in the range of 2 to 40 y and is accurate to f5 per cent. The method may be used on a variety of biological materials and requires only common laboratory equipment. Reagents1. Sulfuric acid, 9 N. A dilut,ion of concentrated sulfuric acid 1: 4 with water. 2. Metaphosphoric acid, 40 per cent. 80.0 gm. in 200 ml. of solution. 3. Potassium bromide, 2 M. 47.6 gm. in 200 ml. of solution. 4. Potassium permanganate, saturated solution. 13 gm. in 200 ml. of solution. 5. Hydrogen peroxide, 6 per cent,, commercial grade. 6. Heptane, practical grade.