A QUANTITATIVE METHOD FOR THE SEPARATION OF THE IODINATED AMINO ACIDS OF THYROID TISSUE1

Abstract

A kieselguhr column chromatographic procedure which is applicable to the separation of thyroxine, triiodothyronine, iodide, diiodotyrosine and monoiodotyrosine was described and evaluated. When added to the column in pure state, the recovery of each of these compounds in its respective peak area is better than 94% . This chromatographic procedure was used to evaluate the hydrolysis of radioactive (I131) thyroid tissue. A trypsin method was shown to provide more complete hydrolysis with a similar or smaller risk of degradation of the liberated amino acids than the standard sodium hydroxide method. Chroma to-graphically pure radiothyroxine, triiodothyronine, iodide, diiodotyrosine and monoiodotyrosine were added to nonradioactive thyroid tissue which was subsequently homogenized, hydrolyzed with trypsin, evaporated, extracted and subjected to this column procedure. Thyroxine and triiodothyronine were recoverable to 73 and 84%, respectively, and the tyrosines to better than 90%. No appreciable I exchange occurred. The radioactive peaks obtained in chromatographic effluents from runs of trypsin hydrolysates of radioactive thyroid tissue were tested as to identity and homogeneity by paper chromatography. The presence of thyroxine, triiodothyronine, iodide, diiodotyrosine, and monoiodotyrosine in human gland hydrolysates was confirmed. In each of these specific peak areas, no quantitatively serious contamination was found.