High performance liquid chromatographic determination of proguanil after derivatisation with sodium benzoxazole-2-sulphonate

Abstract

A simple, fast and reproducible method for the determination of proguanil using high performance liquid chromatographic with UV/Fluorescence detection is described. Proguanil was derivatised to its corresponding derivative [(N¹-(4-chlorophenyl)-N⁵-(1 -methyl ethyl) imidocarbonimideamide-Nbenzoxazole]. The derivatisation reaction was conducted in methanol at 60°C using sodium benzoxazole-2-sulphonate under alkaline conditions. The resulting derivative was extracted with chloroform after which the extract was observed under UV lamp at 254 nm before TLC and HPLC analysis. Similarly, the derivatisation process was adapted for derivatisation of proguanil in urine sample. The reaction proceeded smoothly and rapidly. The extraction process was not cumbersome and eliminated the need for costly extraction and evaporation equipments. The resulting derivative fluorensced intensely under UV lamp. Direct HPLC analysis of the reaction mixture was found possible without interferences from excess reagent and endogeneous compounds like ammonium salts. The derivative eluted in less than seven minutes thus making the method suitable for routine use. The calibration plot was linear over the concentration range. A correlation regression of the order of 0.94 was obtained from the calibration curve which indicated a strong relationship between the instrument response and the concentration of proguanil. The discussion also summarizes the derivatisation chemistry that have not being fully explored to date but may find utility in future development of highly sensitive analytical methods for biquanide drugs.