Tritium -Methylation of -Alkoxy Maleimide Derivatives as Labeling Reagents for Biomolecules.

An efficient procedure to access tritium-labeled maleimide derivatives in a high specific activity has been developed. -Substituted maleimides containing the hydroxy functionality are -methylated in a three-step synthesis route, including (1) Diels-Alder protection of the maleimide core, (2) -methylation by the use of commercially available [H]methyl nosylate, and (3) deprotection by retro-Diels-Alder reaction. With our procedure, -hydroxyalkyl maleimide derivatives can be labeled in overall radiochemical yields of 13-15% and in >98% radiochemical purity. The major advantage of -alkoxy maleimides in comparison to -alkylated maleimides such as -ethylmaleimide is their lower volatility, which enables safer handling with respect to radiation-safety protection. Tritium-labeled maleimide building blocks allow subsequent Michael-type conjugation reactions of thiol-containing biomolecules for mechanistic or studies.