Elimination of interfering chromogens in the Zimmerman reaction for measuring 17-ketosteroids.

HEN ALCOHOLIC POTASSIUM HYDROXIDE and m-dinitrobenzene are used in the Zimmerman reaction (1) for the determination of 17ketosteroids (17-KS), a purple complex is formed which has a broad absorption band in the green portion of the visible spectrum. Application of this reaction to urine extracts also gives rise to other chromogens which are brown in color. On occasion, the ratio of brown chromogens to Zimmerman color is so high that quantitation of 17KS becomes difficult. A variety of procedures have been proposed for eliminating the interference of these brown chromogens in quantitation of 17-KS. The method of Talbot et al. (2) uses mathematical correction after reading the absorption in both the green and blue areas of the visible spectrum. The accuracy of this method has been improved considerably by the use of Allen’s correction (3). Absorbance readings are made at the absorption maximum of the ketosteroid complex and at appropriate wave lengths equidistant on either side. By assuming linear absorption of interfering chromogens at the 3 points, it is possible to make a simple correction. This method is promising but