DETERMINATION OF AMINES USING THE FLUORIMETRIC ION-PAIR TECHNIQUE AS A POST-COLUMN REACTOR FOR HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

A post-column derivatization system using the fluorimetric ion-pair technique is described. The separated amines are mixed in an air-segmented flow with 9,10-dimethoxyanthracene-2-sulphonate and the ion pairs formed are extracted and detected by their fluorescence (383/446 nm). The mobile phase will affect the sensitivity by either increasing the blank or reducing the extractability of the ion pairs. This problem was solved by using a LiChrosorb DIOL stationary phase with an exclusively aqueous mobile phase. The reaction system was tested with hyoscyamine, which was moderately retained (k′ = 0.8) under the chosen conditions. The repeatability of the system was checked by multiple injections (n = 10) of a standard solution (200 ng per injection) and a relative standard deviation of 1.7% was found. The regression was linear over the concentration range of 40-600 ng per injection (r = 0.9998). The limit of detection was about 200 pg (signal-to-noise ratio = 3:1), the improvement therefore being at least 200-fold in comparison with UV detection. The peak width increased by about 40%, probably owing to the dead volume of the reactor. Miniaturization of the system using a smaller phase separator and polypropylene mixing coils of 1 mm I.D: did not provide any improvement. The ion pairs may be adsorbed on the surface of the polypropylene, and glass coils as utilized in the newer generation of AutoAnalyzers will therefore solve that problem. © 1979.