Condensed Report on the Use of Enzymatic Cycling for High Specificity and Precision in the Colorimetric Analysis of Ammonia

摘要

A new method is reported for the colorimetric determination of ammonia in water, based on enzymatic cycling using the ammonia-specific glutamate dehydrogenase reaction followed by an enzymatic cycling reaction that provided controlled amplification of the response in ammonia ion. The cycling reaction product (a tetrazolium formazan) had a conveniently-measured visible absorption maximum at 600 nm. With the established procedure, absorbance versus original ammonium ion concentration plots were linear (correlation coefficients: 0.997-0.9998). The new method was simple and precise in the range of 0.0001-0.0001 molar ammonia concentrations. The sensitivity could be extended well below the ten nanomole per sample determination limit of the assay procedure described, using suitable modifications of the procedure. The coefficient of variation was 2% for a series of replicate measurements on samples containing 0.0000441 molar ammonium ion. A calibration curve for the determination of unknown concentrations could be constructed by carrying standard solutions through the procedure. The method only required the use of volumetric equipment and a spectrophotometer or colorimeter. Analysis time per sample was three hours, and all solutions could be prepared in advance and used for at least two weeks.