Cereal Chem 68:81-86 | VIEW
Use of Fluorometry for the Determination of Uric Acid in Grain. Elimination of Interfering Fluorescence.
W. M. Lamkin, N. C. Unruh, and Y. Pomeranz. Copyright 1991 by the American Association of Cereal Chemists, Inc.
The possible use of fluorometry for determining the amount of uric acid in grain, to measure insect contamination or monitor infestation was investigated. In 0.20M sodium phosphate buffer, pH 12.3, the excitation maximum for the fluorescence of uric acid was 342 nm, and the emission peak was at 408 nm. In alkaline solution, the fluorescence at 408 nm was independent of pH over the range of 11-12.5. Fluorescence at 408 nm was a linear function of uric acid concentration to several hundred parts per million with a lower limit of detection in the range of tenths of a part per million, a sensitivity sufficient for the analysis of uric acid in commercial grain samples. Direct fluorometric analysis of uric acid was not successful as a result of the presence in grain extracts of an interfering fluorescence with an excitation maximum at 290 nm and an emission peak at 354 nm. The interfering fluorescence was shown to be due to the extraction, at the pH of approximately 9 used to extract the uric acid, of tryptophan-containing proteins and peptides. Most of the interfering fluorescence was from proteins with molecular weights above 5,000, but a significant portion was due to compounds with molecular weights below 500. Quantitative removal of the interfering fluorescence was achieved by a cation-exchange procedure using Supelclean LC-SCX solid- phase extraction tubes, which contained 3-propylsulfonyl groups chemically bonded to 40-micrometer silica particles with 60-Angstrom pores.