Assessment of a method of determining ribonuclease activity employing RNA as substrate.

An evaluation of the analytical performance of a method for determining ribonuclease catalytic activity employing digestion of RNA substrate was made. It was found that the relationship between concentrations of ribonuclease standards and concentrations of the reaction products, expressed as the 260 nm light absorbance values, was nonlinear and fulfils a binomial function. Substituting ribonuclease activities by square roots of ribonuclease activity values, a linear relationship with light absorbance values, ranging from 0.08 to 0.400 was obtained. In the present work one unit of ribonuclease catalytic activity was defined as RNA-degrading activity which in the assay conditions caused A260 equal to 0.332. The general equation of the ribonuclease catalytic activity was: Ribonuclease (U/l) = [(A260 - 0.02)/0.316]2. In spite of the nonlinearity of the standard curve, the actual CV values did not depend essentially on the value of measured activity and were estimated as 13%.