A Procedure for Application of the Thiobarbituric Acid Test to Milk

Abstract

Nature of the reaction product(s) has not been established. METHOD TBA test procedure. The TBA test is based upon a reaction involving the formation of a red color when oxidized milk is acidified and heated with 2-thio- barbituric acid. The procedure used in applying the test to lipids and tissue suspensions (7, 10) did not give satisfactory results with milk because the colored compound(s) adsorbed to the milk proteins. Greater reproducibility in estimating the intensity of red color was obtained by using a pyridine-isoamyl alcohol mixture to elute and extract the colored compound(s). After detailed study of a number of factors which influence results of the test, the standardized procedure outlined below was adopted for application of the TBA test to milk. Reagents. TBA reagent : 0.025 M 2-thiebarbituric acid in M phosphoric acid, prepared by mixing equal volumes of 0.05 M 2-thiobarbituric acid and 2 M phos- phoric acid. Discard when precipitate forms. Extraction mixture : 2 : 1 mixture of isoamyl alcohol and pyridine. Method. Ten ml. of milk are pipetted into a 40-ml. centrifuge tube (without pourout), 5 ml. of TBA reagent added and mixed well. The tube is placed in a boiling water bath for 10 m~n, and cooled in cold water. Delay in completion