Blanking for urine color was performed by subtracting the absorbance of parallel reactions of specimens added to biuret reagent without copper.
Visual examination of the reaction products of biuret reagent with amino acids, peptides, and proteins showed a variation in color.
It was apparent that amino acids and small peptides reacted with biuret reagent very rapidly (Fig.
Proteins also had an initial rapid reaction with the biuret reagent, but this was followed by a slow reaction phase over minutes (Fig.
The exact magnitude of cross-reactivity of amino acids and other compounds may vary slightly depending on the sample matrix, composition of the biuret reagent, and instrumentation, but however these conditions are varied, there is likely to be substantial cross-reactivity of these compounds in endpoint biuret assays designed to measure polypeptides and proteins.
The very broad absorbance peaks of the biuret reagent and its many reactants suggest that use of alternative wavelengths for detection offer little opportunity for solving problems of cross-reactivity.