Identification of cyclic nucleotide derivatives synthesized for radioimmunoassay development by fast atom bombardment with collision‐induced dissociation and mass‐analysed ion kinetic energy spectroscopy

Abstract

The syntheses of 2'-O-snccinyl, 2'-O-succinyltyrosinyl methyl ester and 2'-O-succinyliodotyrosinyl methyl ester derivatives of a cyclic nucleotide, derivatives necessary for the successful development of a specific radioimmunoassay, are described. Fast atom bombardment with collision-induced dissociation and mass-analysed ion kinetic energy spectroscopy were used to verify the positions of substitution and the retention of the 3',5'-cyclic phosphate moiety. Comparison of spectra produced after different iodination times permitted the optimization of the reaction conditions.

Kinetic analysis of cyclic CMP-specific and multifunctional phosphodiesterases by quantitative positive-ion fast-atom bombardment mass spectrometry

“…Consequently enzymes concerned with the metabolism and action of cyclic nucleotides are important pharmacological targets. Application of qualitative fastatom bombardment (FAB) mass spectrometry (MS) with mass-analysed ion kinetic energy (MIKE) spectrum scanning has proved invaluable in the identification of endogenous cyclic nucleotides [1][2][3] and of their analogues 4,5 and derivatives, [6][7][8] by virtue of the characteristic fragmentations 9,10 observed in the MIKE spectra obtained after collisionally-induced dissociation (CID) of the protonated molecule in the FAB mass spectrum. The advantage of softionization methods such as FAB is the production of cyclic nucleotide mass spectra without a requirement for derivatization and, combined with CID/MIKE analysis, this permits the differentiation of the 3',5'-cyclic nucleotides, 9,10 which are the biochemical second messengers, from the 2',3'-cyclic nucleotide isomers, which are merely intermediates of nucleic acid catabolism.…”

Qualitative and quantitative MS analysis of cyclic nucleotides and related enzymes

“…T o set up a radioimmunoassay for cyclic nucleotides it is necessary first to synthesize the 2'-O-succinyl derivative; this is used both to link to a carrier protein to generate a cyclic nucleotide-protein conjugate for immunization, and to link to tyrosine methyl ester before labelling with "'1, the latter process generating a high-specificradioactivity labelled antigen. In both cases the preservation of the 3',5'-phosphodiester ring and [22]. Similarly the retention of the phosphodiester ring and the position and number of substitutions can be monitored by FAB/MIKES analysis during the synthesis of dibutyryl cyclic nucleotides, cell-permeating derivatives used to artificially increase intracellular cyclic nucleotide concentrations "231.…”