Kinetic parameters of substrate uptake into vesicles in the presence and absence of 0.1% bovine serum albumin (BSA).

<p>Parameters were calculated based on data from 2–4 separate experiments with GraphPad Prism and are reported as the best fit value (± standard error).</p

Relationship between the albumin effect and amount of vesicles used in the assay.

<p>Transport activity with varying amount of vesicles (measured as total protein in μg per well) were examined in the presence and absence of 0.1% BSA. Substrate concentrations in the assay were (left panel) 50 μM for CDCF (MRP2 vesicles) and (right panel) 200 μM for Lucifer Yellow (BCRP vesicles). Data is expressed as relative transport, normalized to the ATP-dependent uptake in the absence of BSA. Each bar represents the mean (± SD), n = 3–6.</p

The effect of 0.1% BSA on uptake kinetics.

<p>Uptake of (A) 5(6)-carboxy-2’,7’-dichlorofluorescein (CDCF) in MRP2 vesicles, (B) Estradiol-17-β-glucuronide (E₂17βG) in MRP2 vesicles and (C) Lucifer Yellow (LY) in BCRP vesicles in the absence (control, open circles) and presence of 0.1% BSA (closed circles). All assays were performed in triplicate and each point represents the mean (± SD) of 2–4 separate experiments normalized to the calculated V_max of the control. Curves represent the results from model fitting (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0163886#sec002" target="_blank">Materials and Methods</a>).</p

The effect of 0.1% bovine serum albumin (BSA) in BCRP vesicles not loaded with cholesterol.

<p>Uptake of Lucifer Yellow (LY) into BCRP vesicles not loaded with cholesterol is shown in the absence (control, open circles) and presence (closed circles) of 0.1% BSA. The V_max of control was set to 100% and the results normalized to this. Each point represents the mean ± SD, n = 3. Black solid curves represent the fitting of the data for vesicles without cholesterol loading and grey dashed curves represent the corresponding fitting from cholesterol loaded vesicles.</p

Substrate binding to 1.0% BSA as measured using equilibrium dialysis.

<p>The f_u is calculated as the ratio of binding in the presence and absence of BSA and is shown with standard deviation (SD) (n = 3).</p

Inhibition of 5 μM CDCF uptake into MRP2 vesicles and 50 μM LY uptake into BCRP vesicles by oleic acid.

<p>Data is presented as a relative transport activity normalized to uptake in the absence of oleic acid. Small, closed circles show results from inhibition studies and larger black/white circles represent the retention of CDCF or LY in vesicles in studies on unspecific membrane effects of oleic acid (for details see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0163886#sec002" target="_blank">Materials and Methods</a>). All data points were assayed in triplicate and curve fitting was performed using the four parameter logistic equation. Graphs show the results from a representative experiment and data is represented as mean ± SD. IC₅₀ (95% confidence interval) for BCRP was 28.2 μM (21.7–37.7). In the case of MRP2, the IC₅₀ was not applicable.</p

The comparison of 0.1% and 1.0% BSA effect on uptake kinetics.

<p>(Left panel) CDCF uptake into MRP2 vesicles, (right panel) Lucifer Yellow (LY) uptake into BCRP vesicles. Each bar represents the mean (± SD) of 2–3 separate experiments. The uptake rate of the control (0% BSA) was set to 100% and uptake rates in the presence of 0.1% and 1.0% BSA were normalized to control. * p < 0.05 and ** p < 0.01 compared to the uptake in the absence of BSA.</p

Enhanced Photocatalytic Performance of Carbon-Coated TiO_2–<i>x</i> with Surface-Active Carbon Species

Carbon (C) coating on the TiO₂ surface has attracted extensive research interest due to the unique properties of the conjugated materials in electron transport and photoelectronic coupling ability. However, owing to the complexity of surface C species, there is no experimental study on their structure and property. Although the C-coated TiO_2–<i>x</i> photocatalyst (C/TiO_2–<i>x</i>) and its corresponding acid-washed sample (C*/TiO_2–<i>x</i>) exhibit similar visible-light absorption, their catalytic activity is quite different. According to high-resolution transmission electron microscopy, X-ray photoelectron spectroscopy, electron spin resonance, and NMR results, the only structural difference between C/TiO_2–<i>x</i> and C*/TiO_2–<i>x</i> lies in the surface C species. Our NMR experimental results show that several C species (including alkoxy and carboxylate, and macromolecular graphitelike C) are present in C/TiO_2–<i>x</i>, whereas only macromolecular graphitelike C exists in C*/TiO_2–<i>x</i>. Combined with the photocatalytic activity measurements, it can be deduced that the surface graphitelike C should be the active C sites, which facilitate the separation of photoinduced electron and hole and lead to the exceptionally high photocatalytic activity for C*/TiO_2–<i>x</i>, whereas the alkoxy and carboxylate C species that should be the recombination centers would poison seriously the surface of C/TiO_2–<i>x</i>. Accordingly, the hole and electron transfer mechanism in the C-coated TiO_2–<i>x</i> photocatalyst is proposed

Interaction of Food Additives with Intestinal Efflux Transporters

Breast cancer resistance protein (BCRP), multidrug resistance associated protein 2 (MRP2) and P-glycoprotein (P-gp) are ABC transporters that are expressed in the intestine, where they are involved in the efflux of many drugs from enterocytes back into the intestinal lumen. The inhibition of BCRP, MRP2, and P-gp can result in enhanced absorption and exposure of substrate drugs. Food additives are widely used by the food industry to improve the stability, flavor, and consistency of food products. Although they are considered safe for consumption, their interactions with intestinal transporters are poorly characterized. Therefore, in this study, selected food additives, including preservatives, colorants, and sweeteners, were studied <i>in vitro</i> for their inhibitory effects on intestinal ABC transporters. Among the studied compounds, several colorants were able to inhibit BCRP and MRP2, whereas P-gp was fairly insensitive to inhibition. Additionally, one sweetener was identified as a potent inhibitor of BCRP. Dose–response studies revealed that the IC₅₀ values of the inhibitors were lower than the estimated intestinal concentrations after the consumption of beverages containing food colorants. This suggests that there is potential for previously unrecognized transporter-mediated food additive–drug interactions

Transfer Channel of Photoinduced Holes on a TiO₂ Surface As Revealed by Solid-State Nuclear Magnetic Resonance and Electron Spin Resonance Spectroscopy

The detailed structure–activity relationship of surface hydroxyl groups (Ti–OH) and adsorbed water (H₂O) on the TiO₂ surface should be the key to clarifying the photogenerated hole (h⁺) transfer mechanism for photocatalytic water splitting, which however is still not well understood. Herein, one- and two-dimensional ¹H solid-state NMR techniques were employed to identify surface hydroxyl groups and adsorbed water molecules as well as their spatial proximity/interaction in TiO₂ photocatalysts. It was found that although the two different types of Ti–OH (bridging hydroxyl (OH_B) and terminal hydroxyl (OH_T) groups were present on the TiO₂ surface, only the former is in close spatial proximity to adsorbed H₂O, forming hydrated OH_B. In situ ¹H and ¹³C NMR studies of the photocatalytic reaction on TiO₂ with different Ti–OH groups and different H₂O loadings illustrated that the enhanced activity was closely correlated to the amount of hydrated OH_B groups. To gain insight into the role of hydrated OH_B groups in the h⁺ transfer process, in situ ESR experiments were performed on TiO₂ with variable H₂O loading, which revealed that the hydrated OH_B groups offer a channel for the transfer of photogenerated holes in the photocatalytic reaction, and the adsorbed H₂O could have a synergistic effect with the neighboring OH_B group to facilitate the formation and evolution of active paramagnetic intermediates. On the basis of experimental observations, the detailed photocatalytic mechanism of water splitting on the surface of TiO₂ was proposed