Abstract
Combination therapy has remained as standard of care in malaria treatment, since it is a rationale strategy to increase response and tolerability and to decrease resistance. Recent reports in the literature suggest that chloroquinine (CQ) and hydroxychloroquinine (HCQ) could regain its usefulness after a prolonged absence associated with a disappearance of the drug resistance genotype in falciparum malaria and can be combined with artesunate (ART) in formulation to achieve optimum therapeutic response. Separate analysis methods for ART and HCQ are available but simultaneous quantitative estimation of both the drugs is a difficult task as ART lacks extensive chromophores for UV absorption whilst HCQ absorbs highly. In the present study an accurate, precise, simple method has been developed, optimized and validated as per the guidelines of International Conference on Harmonization (ICH) for co estimation of these drugs in formulation as well as for clinical studies. The chromatographic conditions were established by trial and error and were kept constant throughout the experimentation using HPLC isocratic system on C18 column. After trial and error it was found that artesunate peak resolution is maximum in acetonitrile and phosphate buffer (pH 3.0) in the ratio of 50:50 v/v with flow rate 1.0 ml/min at 222nm.Linearity was observed over concentration range of 1-5 μg/ml for ART and 500-2500 μg/ml for HCQ. HCQ and ART were eluted at retention time (Rt) 2.51 and 5.1 min respectively. The Limit of detection and Limit of quantitation were 0.109 and 0.330 μg/ml for ART, and 0.012 and 0.037 μg/ml for HCQ. The method has potential to be applied for the analysis of these antimalarial drugs in combined pharmaceutical formulation with sensitivities in the nano gram-per-milliliter range. The method gave good resolution between ART and HCQ within short analysis time of 14 min with no interfering peaks found in the chromatograms.
