SEPARATION AND INDIRECT DETECTION OF AMINO-ACIDS AS ACETYLATED DERIVATIVES
Iron(II)-1,10-phenanthroline [Fe(phen)3(2+)] salts are used as mobile phase additives for the separation and indirect photometric detection (IPD) of N-acetylated amino acid (N-Ac-AA) derivatives on a reversed stationary phase. Mobile phase Fe(phen)3(2+) concentration, pH, organic modifier concentration and counter anion affect retention and IPD. Disodium-1,5-naphthalene disulfonate and sodium benzoate as counter anions and buffer anion, respectively, are optimum because of their contribution to eluent strength, pH and location of their system peaks. The retention order of N-Ac-AA derivatives is influenced by AA side-chain structure. Mixtures of AA can also be separated, detected by IPD and determined after conversion of the AA to the N-Ac-AA derivatives via acetylation. The detection limit depends on the AA side-chain. For N-Ac-Val and derivatives of similar types of AA the detection limit was 0.5 nmol for a 10-mu-l injection. A similar detection limit was found for Val when the acetylation step was included in the procedure.