A Rabid Method for the Determination of Citric Acid* by Murray Saffran

Numerous methods, enzymatic and chemical, have been reported for the determination of small amounts of citric acid in biological fluids. Of the chemical methods the majority involve the conversion of the citric acid to pentabromoacetone, which may be estimated by various gravimetric, titrimetric, or calorimetric procedures. The more recent modifications (l-3) of the met.hod have simplified it and increased the sensitivity. Even in the simplified methods, however, the procedures are laborious. The method to be described is an application of the Furth-Herrmann reaction (4) and is considerably more rapid and simple to carry out. Purt,h-Herrmann Reaction-The Furth-Herrmann reaction consists simply in taking up the deproteinized sample in warm acetic anhydride and adding pyridine. Citric acid gives a carmine-red color, aconitic acid violet’-red, and tartaric acid emerald-green. The reaction must be carried out under anhydrous conditions. Gronvall (5), in 1937, used it in a roughly quantitative manner for estimating citric acid in the fluids of the eye, the water being removed by evaporation. Although the reaction gives satisfactory results with simple solutions of citric acid, the color development is erratic when the reaction is applied to dried residues of deproteinized serum. This difficulty may be overcome by using an excess of acetic anhydride for dehydration instead of evaporating the sample to dryness. Under these conditions both citric and aconitic acids, on the addition of pyridine, give a yellow color which is nearly proportional to the concentration. By careful control of conditions the modified method can be used to estimate the total of citric and aconitic acids, or aconitic acid alone. Citric acid may be obtained by difference. In practice, however, since the amount of aconitic acid in animal tissues and fluids is negligible, the method may be used for the direct estimation of citric acid.