Use of urease-bromothymol blue-agar method for large-scale testing of urine on grain and seeds

The current AOAC method (963.28) for large-scale (50 g) testing of urine on grain is based on the reaction of sodium in urine with magnesium uranyl acetate. Detection of sodium suggests that urine is present and that a test for urea is appropriate. Urea is detected with urease-bromothymol blue-paper and is confirmed through its reaction with xanthydrol to form dixanthylurea crystals, which are detected microscopically. The initial nonspecific test for sodium can be influenced by the presence of salt or other sodium compounds. Furthermore, the magnesium uranyl acetate spray used in Method 963.28 potentially exposes the analyst to the aerosol of a volatile, toxic uranium compound. Excess reagents and analyzed test portions must be disposed of as radioactive waste. In addition, Method 963.28 requires several steps to determine the presence of urea. The alternative AOAC method (972.41) tests for the presence of urea from urine on individual seeds. Urea is enzymatically decomposed to ammonia and carbon dioxide by urease. Liberated ammonia shifts the pH, changing the color of the indicator in the agar from yellow to blue. This study adapts Method 972.41 to larger test samples. Up to 25 g grains and seeds are sprayed with urease test agar instead of being individually immersed in the urease test agar. The modified method was used to analyze urea on seeds and grains of 24 plants from 4 families. The method has a limit of detection of one seed contaminated with 1 microgram urea.