Bioavailability of orally administered drugs is regulated by P-gp, a member of the ATP binding cassette transporter families. It expresses at the apical surface of epithelial cells and effluxs out several clinically important drugs resulting in decreased absorption and bioavailability. In recent years, the utilization of bioenhancer to increase the bioavailability of drugs has extensively studied. The objective of this study was to evaluate the potential of the compounds found in Citrus hystrix as a bioenhancer for orally administered drugs by modulation of P-gp function. The modulation effects of fruit extracts and isolated pure compounds on P-gp were investigated by uptake assay of the P-gp substrate calcein-AM in Caco-2, LLC-PK1 and LLC-GA5-COL300 cell lines. The results show that the extract from the flavedo part remarkably increased calcein-AM uptake in Caco-2 and LLC-GA5-COL300 cell lines. Among five furanocoumarins identified, 6’,7’-epoxybergamottin, 6’,7’-dihydroxybergamottin and oxypeucedanin significantly enhanced calcein-AM uptake in LLC-GA5-COL300 in a concentration-dependent manner, indicating strongly inhibition effects on P-gp function. Taken together, 6’,7’-epoxybergamottin, 6’,7’-dihydroxybergamottin and oxypeucedanin could be employed as the potential intestinal bioenhancer to improve the bioavailability of P-gp substrate drugs. However, further studies including in vivo studies should be performed to confirm these findings.

The oral delivery is the most practical route to deliver drugs into the body, however drug-metabolizing enzymes and drug transporters can play important roles in modulating drug absorption. This study intended to find a natural bioenhancer for improving drug bioavailability. Two limonoids, including limonin deepoxy and nomilin, isolated from pomelo pulp were studied and the inhibition effects on human CYP3A4 and P-gp were investigated. Testosterone 6β-hydroxylation was performed in recombinant human CYP3A4 to discover the effects on CYP activity. Daunorubicin transport in Caco-2 and calcein-AM uptake in LLC-PK1 and LLC-GA5-COL300 were conducted to evaluate the effects on P-gp function. The results show that both limonin deepoxy and nomilin could inhibit CYP3A4 and only nomilin exhibited mechanism-based inhibition. Nomilin was able to inhibit human P-gp in the concentration-dependent manner. Taken together, nomilin demonstrated strong activities on both CYP3A4 and P-gp, indicating that nomilin could possibly be used as a bioavailability enhancer.

Oxypeucedanin hydrate is a furanocoumarin widely found in various fruits and vegetables so it may interact with prescribed drugs leading to pharmacokinetic interaction. This study was conducted using in vitro cell culture model to investigate the role of oxypeucedanin hydrate on P-gp function. To evaluate the role of oxypeucedanin hydrate as a P-gp substrate, the bidirectional transport studies of oxypeucedanin hydrate were performed in LLC-PK1 and LLC-GA5-COL300. The corrected efflux ratio of oxypeucedanin hydrate was 3.3 ± 0.7, indicating that it was a P-gp substrate. Calcein AM uptakes performed in comparison between LLC-PK1 and LLC-GA5-COL300 as well as daunorubicin transport across Caco-2 cell monolayer were conducted to examine the inhibition effect of oxypeucedanin hydrate on P-gp. The results exhibited that oxypeucedanin hydrate significantly increased calcein accumulation in LLC-GA5-COL300 in a concentration dependent manner and, moreover, the b-a daunorubicin transport across Caco-2 cell monolayer was decreased from 7 to 5, implying the role of the compound on P-gp inhibition, although the effect was quite minimal. Collectively, the results suggested that oxypeucedanin hydrate could act as P-gp substrate and be likely to inhibit P-gp function.

Bioavailability of orally administered drugs can be influenced by many factors. Poor drug absorption across the intestinal membrane is one of the factors that contribute to low bioavailability of drugs. It has been suggested that the metabolism/active efflux in the small intestine is involved in the poor absorption of many drugs. Intestinal CYP3A4 and P-gp work coordinately to reduce the intracellular concentration of drugs. Recently, bioenhancers have been identified and extensively studied. The aim of this study was to evaluate natural furanocoumarins found in juices of common lime and kaffir lime as the potential enhancers for oral delivery by means of modulating CYP3A4 and/or P-gp activities. The role of isolated furanocoumarins on CYP3A4 was assessed by testosterone 6β-hydroxylation reaction, while the effect on P-gp was investigated using R123 and CAM uptake studies in Caco-2, as well as LLC-PK1 and LLC-GA5-Col300. In the present study, we demonstrated that isopimpinellin isolated from common lime is the best CYP3A4 inhibitor among 4 isolated furanocoumarins, implying that isopimpinellin would possibly act as a bioenhancer by inhibiting pre-systemic metabolism. 6’,7’-Dihydroxybergamottin found in kaffir lime is a dual inhibitor of CYP3A4 and P-gp, suggest that it could potentially be used as a bioenhancer by inhibiting both pre-systemic metabolism and efflux mechanism. However, in vivo study should be further conducted to confirm these effects in the body.