Capillary electrophoresis coupled to laser induced fluorescence detection for the analysis of penicillamine in a non-conventional matrix

Penicillamine (DL-2-amino-3-mercapto-3-methyl-butanoic acid) is a chelating agent derived from the hydrolysis of penicillin, lacking any antibiotic properties. The therapeutic form is D-penicillamine, while L-penicillamine is toxic. This drug is used in the treatment of severe active rheumatoid arthritis and acts by reducing the number of T-lymphocytes, inhibiting macrophage function and preventing collagen from cross-linking. It is also used as a chelating agent in Wilson's disease (a rare genetic disorder of copper metabolism), in cystinuria and in the treatment of heavy metal poisoning. Adverse effects are frequent and may include: membranous glomerulonephritis, antibody-mediated myasthenic syndrome, drug-induced systemic lupus erythematosus, toxic myopathies and elastosis perforans serpiginosa. This last one may persist even after the therapy withdrawal. Thus, to evaluate toxic effects in clinical cases we are developing a method based on capillary electrophoresis coupled to laser induced fluorescence detection (CE-LIF) in specific biological matrices such as epithelium. The analysis is carried out in a fused silica capillary, using a carbonate buffer as the background electrolyte. Satisfactory sensitivity was obtained by exciting the molecule at 488 nm after a derivatisation step with 5-(iodoacetamido)fluorescein (IAF). Preliminary results are promising and the validation of the method is in progress. At the same time, we are developing another technique based on reversed phase liquid chromatography with amperometric detection to analyse penicillamine in epithelium samples