Quantification of pyrethroid insecticides from treated bednets using a mosquito recombinant glutathione S-transferase

Recombinant glutathione S-transferase (agGST1-6) from the malaria vector mosquito Anopheles gambiae Giles (Diptera: Culicidae) was expressed in Escherichia coli using a pET3a vector system. The expressed enzyme was biochemically active with reduced glutathione (GSH) and 1-chloro-2,4-dinitrobenzene (CDNB). Activity of agGST1-6 with GSH and CDNB was inhibited to different degrees by both alpha-cyano and non-alpha-cyano pyrethroid insecticides. This inhibition was used to develop an assay for quantification of pyrethroids. Standard curves of insecticide concentration against percentage of enzyme inhibition or volume of iodine solution were established by spectrophotometry and iodine volumetric titration, respectively, for permethrin and deltamethrin. These assays allowed estimation of pyrethroid concentrations both spectrophotometrically and visually. For the residue assay of each insecticide, a cut-off point of 50% of the initial pyrethroid impregnation concentration was used, which should differentiate between biologically active and inactive treated bednets. The cross-reactivity of the primary permethrin photodegradants (3-phenoxyalcohol and 3-phenoxybenzoic acid) with the recombinant agGST1-6 was assayed in the same system. No agGST1-6 inhibition by the insecticide metabolites was observed, suggesting that the system is unaffected by primary permethrin metabolites and will accurately measure insecticide parent compound concentrations. The estimated pyrethroid insecticide concentrations, given spectrophotometrically and by iodine titration assay, were comparable to those obtained by direct HPLC quantification of residues extracted from bednets. Hence, it should be relatively easy to adapt this method to produce a test kit for residue quantification in the field.