The Nitroprusside Reaction as a Test for Reduced Glutathione

Abstract

THIS test was first introduced by Mörner¹. He added sodium nitroprusside and then alkali to cystein and obtained a deep purple red. This changed into a reddish brown and finally disappeared. Cystin gave no reaction. Mörner used a solution of pure cystein, and whether or not he heated his solution prior to the test he did not state. Heating in this case would not affect the results materially. In 1921 Hopkins² very casually suggested heating tissue in weak acetic acid in order to intensify the reaction before applying the test. This unchallenged suggestion has led other workers to heat tissues with acetic acid prior to the application of the test, thus obtaining wholly meaningless results (see, for example, Fink³, Camp⁴ and Coldwater⁵). In 1923 Harris⁶ showed that native proteins which are non-reactive to nitroprusside become reactive when warmed with acid or base. For example, fresh ovalbumin is placed in three test-tubes and to tube A nothing is added; to tube B acetic acid is added; and to tube C acetic acid is added, and this tube is heated. All tubes are rendered alkaline with ammonium sulphate. The nitroprusside test is then negative in tubes A and B and positive only in the tube that was heated. Heat denatures proteins and liberates free —SH radicals if the protein contains such in combined form.