Sensitive radioimmunoassay and enzyme-linked immunosorbent assay for the simultaneous determination of chloroquine and its metabolites in biological fluids

Article abstract of DOI:10.1002/jps.2600790107

Two new methods for the simultaneous determination of chloroquine and its two main metabolites (monodesethylchloroquine and bisdesethylchloroquine) in biological samples, radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA), are described. Antiserum is produced in rabbits immunized with N-(2-carboxyethyl)desethylchloroquine:protein conjugate. Besides chloroquine, this antiserum recognizes with good affinity the two main metabolites, monodesethylchloroquine and bisdesethylchloroquine (70 and 40% of crossreaction, respectively). Amodiaquine cross reacts by 4.5%; cross reactions with monodesethylamodiaquine, bisdesethylamodiaquine, and other antimalarial drugs are <1%. No extraction step or sample preparation is required for either system. Sensitivity limits are, respectively, 0.70 nM (3 pg of chloroquine sulfate measured in 10 μL of plasma sample) for RIA and 10 nM (22 pg of chloroquine sulfate measured in 5 μL of plasma sample) for ELISA. The interassay coefficients of variation are respectively, <10 and <16% for RIA and ELISA in the range 14-410 nM (6-180 ng/mL). The results of both methods are well correlated (r = 0.97) and correlate with spectrophotometry (r = 0.98) and HPLC results (r = 0.93). Because of their high sensitivity, both methods can be used in the case of chloroquine poisoning and in the control of malaria prophylaxis and treatment.