Transport of active flavonoids, based on cytotoxicity and lipophilicity: An evaluation using the blood–brain barrier cell and Caco-2 cell models
Introduction
Flavonoids are biologically active molecules which are found in foods, and have been studied as potential components of functional foods, owing to their benefits for health (Hooper et al., 2008). These components comprise a major group of plant secondary metabolites containing over 3000 different chemical structures, which occur ubiquitously in food items of plant origin and consider as dietary supplements and functional ingredients in herbal remedies (Peterson and Dwyer, 1998). In recent decades, this class of compounds has attracted substantial attention due to many beneficial effects on human health. Studies demonstrate that the flavonoids have various biological activities such as antibacterial, antifungal, anti-inflammatory, anti-tumor, anti-allergic, antioxidant and lowering lipid activities, especially vasodilatory action and cardio-cerebral vascular protection (Koshy et al., 2001, Youdim et al., 2002, Montoro et al., 2005, Aron and Kennedy, 2008, Kang et al., 2011, Salas et al., 2011). Due to their chemical composition (Li et al., 2007, Li et al., 2009, Tsuchiya, 2010), the flavonoids span the range from fully hydrophilic to lipophilic compounds, allowing them to perform their cardio-cerebral vascular protection both in solutions and in biological membranes (Carmen et al., 2007). Many of these flavonoids with in vitro biological activities failed to generate higher activity in vivo owing to the solubility, permeability, bioavailability and/or toxicity, thus, a number of in vivo and in vitro evaluation methods have been developed to determine drug metabolism and permeability (Konsoula and Barile, 2005; Alvarez et al., 2010, Serra et al., 2012).
Although the pharmacological effects, in vivo and in vitro absorption of some flavonoids have been substantially reported (Barrington et al., 2009, Tian et al., 2009), the transport evaluation and comparison of the active flavonoids based on the cytotoxicity via the rat brain microvessel endothelial cell (BMEC) and lipophilicity by oil–water partition coefficient (log P), simultaneously using the blood–brain barrier (BBB) cell and human colon adenocarcinoma cell line (Caco-2) models have been not reported. Nevertheless, the structure–permeability relationship for these active flavonoids was also not studied. In this paper, we chose eight representative and active flavonoids including puerarin, rutin, hesperidin, quercetin, genistein, kaempferol, apigenin and isoliquiritigenin, which had typical chemical structures, especially vasodilatory action and obviously cerebral vascular protection (Schroeter et al., 2001, Wu et al., 2005), to evaluate the permeation and study the structure–permeability relationship based on the cytotoxicity and log P values by determining the Papp values of the eight active flavonoids with the BBB bilayer cell and Caco-2 cell models. Among the permeation experiments, the log P values were determined via a simultaneous determination of these eight flavonoids using a high-performance liquid chromatography; the cytotoxicity was investigated by rat BMEC viability assay; and the BBB bilayer cells were co-cultured via rat brain microvascular endothelial cells and astrocytes. The findings will provide important information for evaluating the transport ability, oral bioavailability and brain distribution of the flavonoids.
Section snippets
Reagents and chemicals
Dulbecco’s modified eagle’s medium (DMEM) was obtained from Gibco Industries Inc. (Grand Island, NY, USA). N-2-Hydroxyethylpiperazine-N-2-ethanesulfonic acid (HEPES), 3-(4,5-dimethylthiazol-2yl)-2, 5-diphenyl tetrazolium bromide (MTT), non-essential amino acids (NEAA), penicillin, streptomycin and trypsin were obtained from Amresco Chemical Co. Ltd. (Solon, OH, USA). Fetal bovine serum (FBS) was purchased from Hyclone Perbio Scientific Co. Ltd. (Logan, UT, USA). Endothelial Cell Medium was
Effects of flavonoids on rat BMEC viabilities
The effects of the eight active flavonoids on rat BMEC viabilities are shown in Fig. 2. Compared with the control group, the eight tested flavonoids inhibited rat BMEC viability in a dose-dependent manner. The tested concentrations of the eight flavonoids including puerarin and rutin (750 μM), hesperidin (200 μM), quercetin (500 μM), genistein (50 μM), kaempferol (70 μM), apigenin and isoliquiritigenin (50 μM) could significantly inhibit rat BMEC viability (P < 0.05 or 0.01). Generally, the inhibition
Discussion
Lipophilicity or hydrophobicity of drugs could affect the transmembrane transport, bioavailability, pharmacological activity and toxicity of dugs (Chan and Stewart, 1996, Konsoula and Barile, 2005). In particular, the lipophilicity of a drug is related to its ability to cross cell membranes by means of passive diffusion (Platts et al., 2006). Oil–water partition coefficient (P) is the ratio of the equilibrium concentration of a compound dissolved in a two-phase system consisting of two
Conclusion
This is the first report of evaluating and comparing the transmembrane transport of eight cardio-cerebral vascular protection flavonoids using the CaCo-2 and BBB cell models, based on the cytotoxicity and lipophilicity. The cytotoxicity and log P of these active flavonoids modified by the number and position of the glycoside and hydroxyl group were the key determinant for the transmembrane transport. The cytotoxicity and Papp values of the flavonoids reduced adversely when the numbers of
Conflict of interest
There is no conflict of interest.
Acknowledgements
This work was supported by grant from the National Foundation of Natural Sciences of China (Nos. 30472057 & 81173121), Beijing Natural Science Foundation Program (Nos. 7052007 & KZ201110025024) and Funding Project for Academic Human Resources Development in Institutions of High Learning under the Jurisdicrion of Beijing Municipality (PHR201007111).
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