Determination of the trypanocidal drug melarsoprol and its conversion products in biological fluids with...

Although melarsoprol, an organoarsenic compound, is widely used for the treatment of trypanosomiasis (human African sleeping sickness), very little is known about its fate in the human body, its active metabolites passing the blood-brain barrier and the mode of action. Previous pharmacological studies based on the determination of melarsoprol by HPLC-UV or by a bioassay method produced different results. We report a HPLC-ICPMS method suitable for determining melarsoprol and its metabolites in biological fluids. The arsenic selective capability of the method allowed the quantitative measurement of melarsoprol and two arsenic-containing conversion products produced when melarsoprol was incubated with human serum and blood. The major product was identified as melarsen [4-[(4,6-diamino-1,3,5-triazin-2-yl)amino]phenyl]arsonic acid by HPLC/electrospray MS, and by accurate mass measurements. Investigations about the stability of melarsoprol in serum showed that within 30 h about 10% of melarsoprol is converted to melarsen. In blood, however, most of the melarsoprol was bound to proteins and only 1% was converted to melarsen after 30 hours. The limit of detection for melarsoprol and its conversion products were in the range of 1 µg AsL(-1) (13 nmol As L(-1)) based on signal to noise ratio of 3 with a 10 µL injection volume allowing direct determination of the compounds in blood and serum (after protein precipitation) at therapeutically realistic concentrations.