title: Inhibition against mosquito cytochrome P450 enzymes by rhinacanthin-A, -B, and -C elicits synergism on cypermethrin cytotoxicity in Spodoptera frugiperda cells.
abstract: Rhinacanthus nasutus (Acanthaceae) is a shrub reported to contain insecticidal activities. The current study was conducted to determine whether R. nasutus constituents could inhibit benzyloxyresorufin O-debenzylation (BROD) mediated by CYP6AA3 and CYP6P7. Both enzymes have shown pyrethroid degradation activity and been implicated to play role in pyrethroid resistance in Anopheles minumus (Theobald) mosquito, a malaria vector. Three compounds, rhinacanthin-A, -B, and -C that exhibited potent inhibitory activity were isolated and purified from aerial part of R. nasutus. Their kinetic parameters and modes of inhibition were determined. Rhinacanthin-B was the most potent inhibitor in in vitro inhibition assay and exhibited mechanism-based inhibition against both CYP6AA3 and CYP6P7. Rhinacanthin-C which is a major compound of R. nasutus reversibly inhibited both enzymes in vitro with 2-4 folds less inhibitory potency than rhinacanthin-B. In contrast, rhinacanthin-A reversibly inhibited CYP6AA3, but inhibition against CYP6P7 was a mechanism-based inhibition type. Where mechanism-based inhibition was found, the inhibition showed characteristic of time-, concentration-dependence, and requirement of NADPH. Using 3-(4, 5-dimethylthiazol-2-y-l)-2, 5-diphenyltetrazolium bromide (MTT) cytotoxicity assay in intact Spodoptera frugiperda (Sf9) cells, the three compounds increased susceptibility of CYP6AA3- and CYP6P7-expressing cells to cypermethrin cytotoxicity because of inhibition effect on mosquito enzymes. The combined inhibition effect on mosquito cytochrome P450 enzyme and synergistic effect on cypermethrin cytotoxicity of the three R. nasutus compounds could be beneficial for resistance management strategies in mosquito vector control.
authors.affiliation: Department of Biochemistry, Faculty of Science, Mahidol University, Rama 6 Road, Phyatai, Bangkok 10400, Thailand.
authors.forename: Sirikun
authors.initials: S
authors.lastname: Pethuan
authors.forename: Panida
authors.initials: P
authors.lastname: Duangkaew
authors.forename: Songklod
authors.initials: S
authors.lastname: Sarapusit
authors.forename: Ekaruth
authors.initials: E
authors.lastname: Srisook
authors.forename: Pornpimol
authors.initials: P
authors.lastname: Rongnoparut
mesh_terms: D027901:Acanthaceae / Q000737:chemistry*; D000818:Animals; D000852:Anopheles / Q000201:enzymology*; D002460:Cell Line; D065607:Cytochrome P-450 Enzyme Inhibitors; D007305:Insecticide Resistance; D007306:Insecticides; D007700:Kinetics; D009285:Naphthoquinones / Q000737:chemistry* / Q000302:isolation & purification; D010078:Oxazines / Q000378:metabolism; D010574:Pesticide Synergists / Q000302:isolation & purification; D011722:Pyrethrins; D018411:Spodoptera
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D007306:Insecticides; D009285:Naphthoquinones; D010078:Oxazines; D010574:Pesticide Synergists; D011722:Pyrethrins; C515073:rhinacanthin-C; C017160:cypermethrin; C057754:benzyloxyresorufin