Microbore Liquid Chromatography of Tertiary Amine Anticholinergic ...

Published: Journal of Chromatographic Science, Volume 33, Number 6, June 1995, pp. 297-302.

Microbore Liquid Chromatography of Tertiary Amine Anticholinergic Pharmaceuticals with Tris(2,2'-bi- pyridine)ruthenium(III) Chemiluminescence Detection
John A. Holeman and Neil D. Danielson

The post-column chemiluminescent reaction of six anticholinergic alkaloid compounds with tris(2,2'-bipyridine)ruthenium(III) (Ru(bpy)33+) is applied to microbore high-performance liquid chromatography (HPLC). At flow rates less than 200 µL/min, the capillary mixing cell in which Ru(bpy)33+ and the analyte are mixed directly allows for good light detection. In contrast, a diminished signal occurs at these low flow rates with conventional post-column mixing in a tee. Optimal chemiluminescent pH conditions for atropine, scopolamine, dicyclomine, cyclopentolate, cyclobenzaprine, and procyclidine are determined at moderately basic conditions (pH 7 to 9). 2-Butanone is found to be compatible with the chemiluminescent reaction, whereas tetrahydrofuran and propionitrile cause an increase in background noise and a chemiluminescent signal loss. As 2-butanone is more nonpolar than acetonitrile, it assists in the elution of these hydrophobic anticholinergic compounds. Five anticholinergic compounds are resolved successfully with a PRP-1 polymeric column and a slightly basic mobile phase, but a C8 silica column is better suited for the more hydrophobic compounds (cyclobenzaprine, procyclidine, and dicyclomine).

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