Of the procedures used for the determination of xanthurenic acid, the calorimetric method of Rosen, Lowy, and Sprince (2) has proved to be by far the most reliable and is therefore the one most frequently applied, even if sometimes with various slight modifications. This method is based on photometric measurement of the green colored xanthurenic acidferrous complex in weak alkaline media. Alt,hough Wachstein and Gudaitis (3) as well as Glazer, Mueller et .aZ. (1) simplified the method without reducing its accuracy, the expenditure of work in routine determinations is still considerable. It is often necessary to deproteinize the urine, since even t,races of protein may interfere with the determination.
We tried therefore to automatize xanthurenic acid determination1 based on the method inaugurated by Wachetein and Gudaitis (3), who add tris(hydroxymethyl)aminomethane (tris) buffer (pH 7.8) to 2 to 10 ml urine and carry out the determination at 610 nm following the addition of 0.1 ml of a 1.770 ammonium ferric sulfate solution. We decided upon a manifold which allows determinations of xanthurenic acid in the urine within a concentration range of 1 to 100 mg/lOO ml. The deproteinization is achieved by the use of a dialyzer, consequently avoiding distortion of the analytical data. The only disadvantage in the original procedure proved to be the weak alkaline buffer which, on prolonged operation of the automatic system, causes sedimentation of ferrous hydroxide. This trouble was eliminated by changing the pH of the buffer to 6.6 and by using a less concentrated ammonium ferric sulfate solution (O.O5%), impairing neither the sensitivity nor the specificity of the method. MATERIALS AND METHOD Reagents 1. Maleic acid-tris (hydroxymethyl) aminomethane buffer, pH 6.6: 58 gm of maleic acid (Merck No. 380) and 61 gm of tris(hydroxymethyl)-' AutoAnalyzer Inanufactured by Technicon.