The
cytochrome P450 monooxygenases play a major role in
insecticide detoxification and become a target for development of
insecticide synergists. In this study, a collection of rhinacanthins (
rhinacanthin-D, -E, -G, -N, -Q, and -H/I) purified from Rhinacanthus nasutus, in addition to previously purified
rhinacanthin-B and -C, were isolated. These compounds displayed various degrees of inhibition against
benzyloxyresorufin-O-debenzylation mediated by CYP6AA3 and CYP6P7 which were implicated in
pyrethroid resistance in Anopheles minimus
malaria vector. Inhibition modes and kinetics were determined for each of rhinacanthins. Cell-based inhibition assays by rhinacanthins employing 3-(4, 5-dimethylthiazol-2-y-l)-2, 5-diphenyltetrazolium
bromide (MTT) cytotoxicity test were explored their synergistic effects with
cypermethrin toxicity on CYP6AA3- and CYP6P7-expressing Spodoptera frugiperda (Sf9) cells.
Rhinacanthin-B, -D, -E, -G, and -N exhibited mechanism-based inhibition against CYP6AA3, an indication of irreversible inhibition, while
rhinacanthin-B, -D, -G, and -N were mechanism-based inhibitors of CYP6P7. There was structure-function relationship of these rhinacanthins in inhibition effects against both
enzymes. In vitro enzymatic inhibition assays revealed that there were synergistic interactions among rhinacanthins, except
rhinacanthin-B and -Q, in inhibition against both
enzymes. These rhinacanthins exerted synergism with
cypermethrin toxicity on Sf9 cells expressing each of the two
P450 enzymes via P450 inhibition and in addition could interact in synergy to further increase
cypermethrin toxicity. The inhibition potentials, synergy among rhinacanthins in inhibition against the P450 detoxification
enzymes, and synergism with
cypermethrin toxicity of the R. nasutus constituents of reported herein could be beneficial to implement effective resistance management of mosquito vector control.