Polymethoxylated flavones in orange juice are inhibitors of P-glycoprotein but not cytochrome P450 3A4.

The presence in orange juice of compounds that specifically inhibit the P-glycoprotein (P-gp) drug efflux transporter, but not the cytochrome P450 (CYP) isozyme CYP3A4, was investigated. The uptake of [(3)H]vinblastine, a substrate of P-gp, by Caco-2 cells was measured. An ethyl acetate extract of orange juice did not affect the initial uptake rate of [(3)H]vinblastine but significantly increased the steady-state uptake, as did cyclosporin A (20 microM), an inhibitor of P-gp. No significant effect on the uptake of 3-O-[(3)H]methylglucose or [(14)C]phenylalanine by Caco-2 cells was found, compared with the control. When the extract was separated on a Cosmosil column, the eluate with 70% methanol showed the most potent ability to increase [(3)H]vinblastine uptake. Additional separation of the 70% methanol eluate on a silica gel column with hexane-acetone (3:1) gave 3,3',4',5,6,7,8-heptamethoxyflavone (HMF) and 4',5,6,7,8-pentamethoxyflavone (tangeretin). HMF, tangeretin, and 3',4',5,6,7,8-hexamethoxyflavone (nobiletin), another methoxyflavone contained in orange juice, all increased the steady-state uptake of [(3)H]vinblastine by Caco-2 cells in a concentration-dependent manner. The order of potency of these compounds at the concentration of 50 microM was tangeretin > HMF > nobiletin. None of these methoxyflavones inhibited 6beta-hydroxylation of testosterone catalyzed by CYP3A4. The ethyl acetate extract of orange juice and these methoxyflavones also increased steady-state [(3)H]vinblastine uptake by LLC-GA5-COL300 cells (a cell line transfected with human MDR1 cDNA). We conclude that these methoxyflavones enhanced vinblastine uptake by specifically inhibiting drug efflux via P-gp. They may have potential as agents for reversing multidrug resistance or for recovering the bioavailability of certain drugs.