A MICROMETHOD FOR QUANTITATIVE-DETERMINATION OF IODOAMINO ACIDS IN THYROGLOBULIN

We describe a new method for quantification of iodoamino acids after e nzymatic hydrolysis of thyroglobulin. The procedure involves separatio n of monoiodotyrosine (MIT), diiodotyrosine, tri-iodothyronine and thy roxine by reverse phase HPLC with a Vydac C-18 stationary phase and a mobile phase of water-acetonitrile-acetic acid. The separation is moni tored by sensitive spectrophotometric detection through a 96-well micr oplate system based on the catalytic Sandell-Kolthoff reaction of iodi de on the oxidation of arsenic(III) by cerium(IV). This new microassay is particularly convenient because of its high sensitivity and its ra pidity (less than 2 h). It can detect 1 pmol MIT and 0.5 pmol of the o ther three iodoamino acids with a recovery higher than 96%. Moreover, the 96-well microplate system allows many samples to be tested simulta neously and avoids the use of radio-labeled iodine.