Analysis of fluorine and iodine derivatives of tyrosine

Abstract

Separation of tyrosine, fluorotyrosine, monoiodotyrosine and diiodotyrosine was achieved by reversed-phase high-performance liquid chromatography (HPLC) using a gradient of acetonitrile with water and using trifluoroacetic acid for ion pairing. No derivatization of the amino acids, prior to separation, was needed. The spectral properties of Tyr and its fluorine and iodine derivatives and the dependence of their absorbance maxima on pH, made it possible to analyze and differentiate between these derivatives in the free amino acid form or in peptides. This analysis was accomplished by adjusting the post column HPLC eluate from two identical runs to different pH values and then comparing the spectra of the peaks from these two runs with a diode array detector. Hydrolysis in 6 M hydrochloric acid was totally destructive to mono- and diiodotyrosine. However, base hydrolysis in 13.5 M sodium hydroxide for 30 min at 121°C in an autoclave caused no destructive and allowed excellent recovery of all of the Tyr derivatives. This is the first report of simple methods for the detection and analysis of these amino acids and of a hydrolytic method which protects against their loss. A method of storage was also proposed.