In-Syringe Electrokinetic Protein Removal from Biological Samples prior to Electrospray Ionization Mass Spectrometry.

Here, an electrokinetic extraction (EkE) syringe is presented allowing for on-line electrokinetic removal of serum proteins before ESI-MS. The proposed concept is demonstrated by the determination of pharmaceuticals from human serum within minutes, with sample preparation limited to a 5× dilution of the sample in the background electrolyte (BGE) and application of voltage, both of which can be performed in-syringe. Signal enhancements of 3.6-32 fold relative to direct infusion of diluted serum and up to 10.8 fold enhancement, were obtained for basic and acidic pharmaceuticals, respectively. Linear correlations for the basic drugs by EkE-ESI-MS/MS were achieved, covering the necessary clinical range with LOQs of 5.3, 7.8, 6.1, and 17.8 ng mL-1 for clomipramine, chlorphenamine, pindolol, and atenolol, respectively. For the acidic drugs, the EkE-ESI-MS LOQs were 3.1 µg mL-1 and 2.9 µg mL-1 for naproxen and paracetamol, respectively. The EkE-ESI-MS and EkE-ESI-MS/MS methods showed good accuracy (%found of 81 % to 120 %), precision (≤20 %), and linearity (r>0.997) for all the studied drugs in spiked serum samples.

Dispersive Liquid-Liquid Microextraction Based on Solidification of Floating Organic Drop with Central Composite Design for the Spectrofluorometric Determination of Naproxen.

A quick, simple and efficient method for extraction, preconcentration, and determination of naproxen in water and plasma specimens with acceptable recovery by dispersive liquid-liquid microextraction based on solidified floating organic drop with spectrofluorimetry is presented. Various parameters affecting the extraction efficiency are optimized by the Central Composite Design. Moreover, under optimal conditions (120 µL 1-Undecanol with 1 mL Ethanol, pH = 3.5, 2 mL KCl 10% solution), the calibration curve was linear in the range 10.0-120.0 ng/mL. Finally, for naproxen, the detection limit was 2.4 ng/mL.

Submicellar liquid chromatography with fluorescence detection improves the analysis of naproxen in plasma and brain tissue.

Rapid, simple, and sensitive submicellar liquid chromatography with fluorescence detection was developed and validated to quantify naproxen in plasma and brain samples after oral administration of Naproxen formulations. The method used tramadol as an internal standard. Different submicellar mobile phases with organic phases ranging from 40 to 60% were studied to improve the native fluorescence of the Naproxen and decrease retention times. Separation was done in a Zorbax SB C8 column (250 × 4.6 mm, 5 µm) with a mobile phase containing acidic 0.007 M sodium dodecyl sulfate/acetonitrile (50:50, v/v) at a flow rate of 1 mL/min. Detection was performed with an excitation wavelength of 280 nm and emission of 310 nm and 360 nm for internal standard and Naproxen, respectively. The method was validated by International Conference of Harmonization standards. The method is specific, accurate, and precise (relative standard deviation <3%). Limits of detection and quantification were 0.08 and 0.25 µg/mL, respectively, for biological samples. This method was applied to analyze brain/plasma ratios in mice that had received oral administrations of Naproxen micellar formulations containing 10% w/w of sodium dodecyl sulfate, Cremophor RH 40, or Tween 80. The sodium dodecyl sulfate micelles were faster and more widely distributed in the mouse brains.

Simultaneous voltammetric determination of acetaminophen, naproxen, and theophylline using an in-situ polymerized poly(acrylic acid) nanogel covalently grafted onto a carbon black/La2O3 composite.

Lanthanum oxide nanomaterials were decorated with carbon black (CB) and grafted with a poly(acrylic acid) nanogel to obtain a composite material (CB-g-PAA/La2O3) for simultaneous determination of acetaminophen (AMP), naproxen (NPX), and theophylline (TPH). The nanogel was synthesized by in-situ free radical polymerization. The composite was dropped onto a glassy carbon electrode (GCE), and the modified GCE displays robust electrocatalytic activity towards AMP, NPX, and TPH, with voltammetric signals that are enhanced compared to a bare GCE. Features of merit for AMP, NPX, and TPH, respectively, include (a) peak potentials of 0.42, 0.85 and 0.12 V (vs. Ag/AgCl), (b) linear ranges from 0.05-887, 0.05-884, and 0.02-888 µM, and (c) detection limits of 20, 35, and 15 nM. The practical applicability of the CB-g-PAA/La2O3/GCE was illustrated by analyzing serum and urine samples. Graphical abstract Schematic presentation of simultaneous electrochemical sensing of acetaminophen (AMP), naproxen (NPX), and theophylline (TPH) in real sample analysis using poly(acrylic acid) nanogel covalently grafted onto a carbon black/La2O3 composite (CB-g-PAA/La2O3/GCE).

Pharmacokinetic studies of naproxen amides of some amino acid esters with promising colorectal cancer chemopreventive activity.

Naproxen (nap) is belonging to Non-steriodal anti-inflammatory drugs (NSAIDs) group of drugs that characterized by their free carboxylic group. The therapeutic activity of nap is usually accompanied by GI untoward side effects. Recently synthesized naproxen amides of some amino acid esters prodrugs to mask the free carboxylic group were reported. Those prodrugs showed a promising colorectal cancer chemopreventive activity. The current study aims to investigate the fate and hydrolysis of the prodrugs kinetically in different pH conditions, simulated gastric and intestinal fluids with pHs of 1.2, 5.5 and 7.4 in vitro at 37 °C. The effect of enzymes on the hydrolysis of prodrugs was also studied through incubation of these prodrugs at 37 °C in human plasma and rat liver homogenates. The pharmacokinetic parameters of selected prodrugs and the liberated nap were studied after oral and intraperitoneal administration in male wistar rats. The results showed the hydrolysis of naproxen amides of amino acid esters to nap through two steps first by degradation of the ester moiety to form the amide of nap with amino acid and the second was through the degradation of the amide link to liberate nap. The two reactions were followed and studied kinetically where K1 and K2 (rate constants of degradation) is reported. The hydrolysis of prodrugs was faster in liver homogenates than in plasma. The relative bioavailability of the liberated nap in vivo was higher in case of prodrug containing ethyl glycinate moiety than that occupied l-valine ethyl ester moiety. Each of nap. prodrugs containing ethyl glycinate and l-valine ethyl ester moieties appears promising in liberating nap, decreasing direct GI side effect and consequently their colorectal cancer chemopreventive activity.

Effect of Disease-Related Changes in Plasma Albumin on the Pharmacokinetics of Naproxen in Male and Female Arthritic Rats.

Naproxen (NPX) is used in the treatment of rheumatoid arthritis (RA) for alleviation of pain and inflammation. In view of the extensive albumin binding of NPX, this study investigates whether chronic inflammation and sex influence the physiologic albumin concentrations, plasma protein binding, and pharmacokinetics (PK) of NPX. The PK of NPX was evaluated in a rat model of RA [collagen-induced arthritis (CIA) in Lewis rats] and in healthy controls. These PK studies included 1) NPX in female and male CIA rats that received 10, 25, or 50 mg/kg NPX i.p.; and 2) NPX in healthy female and male rats after i.p. dosing of NPX at 50 mg/kg. Plasma albumin concentrations were quantified by enzyme-linked immunosorbent assay, and protein binding was assessed using ultrafiltration. The NPX concentrations in plasma and filtrates were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Plasma concentration-time data of NPX were first assessed by noncompartmental analysis (NCA). Nonlinear PK as indicated by dose-dependent NCA clearances and distribution volumes was observed. A two-compartment model with a first-order absorption process incorporating nonlinear protein binding in plasma and tissues jointly described the PK data of all groups. Saturable albumin binding accounts for the nonlinearity of NPX PK in all rats as well as part of the PK differences in arthritic rats. The CIA rats exhibited reduced albumin concentrations, reduced overall protein binding, and reduced clearances of unbound NPX, consistent with expectations during inflammation. The net effect of chronic inflammation was an elevation of the Cmax and area under the plasma concentration-time curve (AUC) of unbound drug.

Development and validation of an LC-ESI-MS/MS method for the simultaneous quantification of naproxen and sumatriptan in human plasma: application to a pharmacokinetic study.

A sensitive and fast liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) method was developed and validated for the simultaneous quantification of naproxen and sumatriptan in human plasma. A simple liquid-liquid extraction procedure, with a mixture of ethyl acetate, methyl tert-butyl ether, and dichloromethane (4:3:3, v/v), was used for the cleanup of plasma. Naratriptan and aceclofenac were employed as internal standards. The analyses were carried out using an ACE C18 column (50 × 4.6 mm i.d.; particle size 5 µm) and a mobile phase consisting of 2 mM aqueous ammonium acetate with 0.025 % formic acid and methanol (38:62, v/v). A triple-quadrupole mass spectrometer equipped with an electrospray source in the positive mode was set up in the selective reaction monitoring mode to detect the ion transitions m/z 231.67 → m/z 185.07, m/z 296.70 → m/z 157.30, m/z 354.80 → m/z 215.00, and m/z 336.80 → m/z 97.94 for naproxen, sumatriptan, aceclofenac, and naratriptan, respectively. The method was validated and proved to be linear, accurate, precise, and selective over the ranges of 2.5-130 µg mL(-1) for naproxen and 1-50 ng mL(-1) for sumatriptan. The validated method was successfully applied to a pharmacokinetic study with simultaneous administration of naproxen sodium and sumatriptan succinate tablet formulations in healthy volunteers.

Validated HPLC-UV method for determination of naproxen in human plasma with proven selectivity against ibuprofen and paracetamol.

Estimating the influence of interfering compounds present in the biological matrix on the determination of an analyte is one of the most important tasks during bioanalytical method development and validation. Interferences from endogenous components and, if necessary, from major metabolites as well as possible co-administered medications should be evaluated during a selectivity test. This paper describes a simple, rapid and cost-effective HPLC-UV method for the determination of naproxen in human plasma in the presence of two other analgesics, ibuprofen and paracetamol. Sample preparation is based on a simple liquid-liquid extraction procedure with a short, 5 s mixing time. Fenoprofen, which is characterized by a similar structure and properties to naproxen, was first used as the internal standard. The calibration curve is linear in the concentration range of 0.5-80.0 µg/mL, which is suitable for pharmacokinetic studies following a single 220 mg oral dose of naproxen sodium. The method was fully validated according to international guidelines and was successfully applied in a bioequivalence study in humans. Copyright © 2015 John Wiley & Sons, Ltd.

SIMPLE, SENSITIVE AND SELECTIVE SPECTROPHOTOMETRIC ASSAY OF NAPROXEN IN PURE, PHARMACEUTICAL PREPARATION AND HUMAN SERUM SAMPLES.

Two simple, rapid and sensitive spectrophotometric methods have been developed for the determination of naproxen in pure, pharmaceutical preparation and human serum samples. These methods are based on the formation of yellow ion-pair complexes between naproxen and two sulfophthalein acid dyes, namely bromocresol green (BCG method) and bromothymol blue (BTB method). The resulting complexes were measured at 424 nm (BCG method) and at 422 nm (BTB method). The effects of variables such as reagent concentration and reaction time were investigated to optimize the procedure. Beer's law was obeyed in the concentration range of 10-105 µg/mL and 5-85 µg/mL and the detection limits were found to be 0.347 and 0.31 µg/mL for BCG and BTB methods, respectively. The developed methods have been successfully applied for the determination of naproxen in bulk drugs, pharmaceutical formulations and human serum samples with good accuracy and precision. The results are comparable to those of reference methods, and hence are recommended for quality control and routine analysis.

Model-based analysis of thromboxane B2 and prostaglandin E2 as biomarkers in the safety evaluation of naproxen.

The assessment of safety in traditional toxicology protocols relies on evidence arising from observed adverse events (AEs) in animals and on establishing their correlation with different measures of drug exposure (e.g., Cmax and AUC). Such correlations, however, ignore the role of biomarkers, which can provide further insight into the underlying pharmacological mechanisms. Here we use naproxen as a paradigm drug to explore the feasibility of a biomarker-guided approach for the prediction of AEs in humans. A standard toxicology protocol was set up for the evaluation of effects of naproxen in rat, in which four doses were tested (7.5, 15, 40 and 80 mg/kg). In addition to sparse blood sampling for the assessment of exposure, thromboxane B2 and prostaglandin E2 were also collected in satellite groups. Nonlinear mixed effects modelling was used to evaluate the predictive performance of the approach. A one-compartmental model with first order absorption was found to best describe the pharmacokinetics of naproxen. A nonlinear relationship between dose and bioavailability was observed which leads to a less than proportional increase in naproxen concentrations with increasing doses. The pharmacodynamics of TXB2 and PGE2 was described by direct inhibition models with maximum pharmacological effects achieved at doses >7.5 mg/kg. The predicted PKPD relationship in humans was within 10-fold of the values previously published. Moreover, our results indicate that biomarkers can be used to assess interspecies differences in PKPD and extrapolated data from animals to humans. Biomarker sampling should be used systematically in general toxicity studies.

Enhanced chemopreventive effects of a hydrogen sulfide-releasing anti-inflammatory drug (ATB-346) in experimental colorectal cancer.

Regular use of nonsteroidal anti-inflammatory drugs is associated with a significantly lower incidence of several types of cancer, particularly those affecting the gastrointestinal tract. However, the propensity of these drugs to cause ulcers and bleeding in the stomach and small intestine limits their utility for chemoprevention of cancer. In the present study, we evaluated the effectiveness of a novel hydrogen sulfide-releasing derivative of naproxen in reducing the incidence of pre-cancerous lesions (aberrant crypt foci) in mice treated with the carcinogen azoxymethane. Weekly administration of azoxymethane over a 4-week period resulted in formation of an average of ∼50 aberrant crypt foci in the colon. Twice-daily treatment with naproxen at high doses significantly reduced the number of aberrant crypt foci. However, a significantly greater effect was observed with ATB-346 (H2S-releasing naproxen) and it was also effective at much lower doses, where naproxen was ineffective. The H2S-releasing moiety of ATB-346 did not significantly affect the number of aberrant crypt foci, suggesting that both the inhibition of cyclooxygenase activity and release of H2S were necessary for the enhanced chemopreventative effect. ATB-346 suppressed colonic prostaglandin synthesis and whole blood thromboxane synthesis as effectively as naproxen, but did not induce any gastrointestinal injury. These results demonstrate that ATB-346 exerts superior chemopreventive effects to those of naproxen, while sparing the gastrointestinal tract of the injury normally associated with use of the parent drug. ATB-346 may therefore be an attractive agent for chemoprevention of colon cancer, and possibly of cancers in other tissues.

Determination of naproxen in human plasma by GC-MS.

This paper describes a GC-MS method for the determination of naproxen in human plasma. Naproxen and internal standard ibuprofen were extracted from plasma using a liquid-liquid extraction method. Derivatization was carried out using N-methyl-N-(trimethylsilyl)trifluoroacetamide. The calibration curve was linear between the concentration range of 0.10-5.0 µg/mL. Intra- and interday precision values for naproxen in plasma were <5.14, and accuracy (relative error) was better than 4.67%. The extraction recoveries of naproxen from human plasma were between 93.0 and 98.9%. The LOD and LOQ of naproxen were 0.03 and 0.10 µg/mL, respectively. Also, this assay was applied to determine the pharmacokinetic parameters of naproxen in six healthy Turkish volunteers who had been given 220 mg naproxen.

Electrospun titania sol-gel-based ceramic composite nanofibers for online micro- solid-phase extraction with high-performance liquid chromatography.

Titanium(IV) tetraisopropoxide was employed as a metal oxide sol-gel precursor to prepare ceramic composite nanofibers by the electrospinning system. To facilitate this process and obtain the desired nanofibers with higher aspect ratios and surface area, poly(vinylpyrrolidone) was added to the sol of titania. Four ceramic nanofibers sheets based on titania were prepared while each sheet contained different transition metals such as Fe-Mn, Fe-Ni, Fe-Co, and Fe-Mn-Co-Ni. The scanning electron microscope images showed good homogeneity for all the prepared ceramic composites with a diameter range of 100-250 nm. The sorption efficiency was investigated by a micro-solid-phase extraction setup in online combination with high-performance liquid chromatography for the determination of naproxen and clobetasol. All the prepared composites exhibited comparable efficiencies for the desired analytes and the type of metal showed insignificant effect. For the selected composite with Fe-Mn, the linearity of the analytes was in the range of 1-1000 µg/L and the limit of detection values were found to be 2 and 0.3 µg/L for naproxen and clobetasol, respectively. The developed method was extended to the analysis of urine and blood plasma samples and acceptable relative standard deviations were obtained at two concentration levels.

A simple high-performance liquid chromatography method for simultaneous determination of three triazole antifungals in human plasma.

A rapid and simple high-performance liquid chromatography (HPLC) assay was developed for the simultaneous determination of three triazole antifungals (voriconazole, posaconazole, and itraconazole and the metabolite of itraconazole, hydroxyitraconazole) in human plasma. Sample preparation involved a simple one-step protein precipitation with 1.0 M perchloric acid and methanol. After centrifugation, the supernatant was injected directly into the HPLC system. Voriconazole, posaconazole, itraconazole, its metabolite hydroxyitraconazole, and the internal standard naproxen were resolved on a C(6)-phenyl column using gradient elution of 0.01 M phosphate buffer, pH 3.5, and acetonitrile and detected with UV detection at 262 nm. Standard curves were linear over the concentration range of 0.05 to 10 mg/liter (r(2) > 0.99). Bias was <8.0% from 0.05 to 10 mg/liter, intra- and interday coefficients of variation (imprecision) were <10%, and the limit of quantification was 0.05 mg/liter.

Development and validation of a sensitive and high-throughput LC-MS/MS method for the simultaneous determination of esomeprazole and naproxen in human plasma.

A sensitive and high-throughput LC-MS/MS method has been developed and validated for the combined determination of esomeprazole and naproxen in human plasma with ibuprofen as internal standard. Solid-phase extraction was used to extract both analytes and internal standard from human plasma. Chromatographic separation was achieved in 4.0 min on XBridge C18 column using acetonitrile-25 mM ammonium formate (70:30, v/v) as mobile phase. Mass detection was achieved by ESI/MS/MS in negative ion mode, monitoring at m/z 344.19 → 194.12, 229.12 → 169.05 and 205.13 → 161.07 for esomeprazole, naproxen and IS, respectively. The calibration curves were linear from 3.00 to 700.02 ng/mL for esomeprazole and 0.50 to 150.08 ng/mL for naproxen. The intra- and inter-batch precision and accuracy across four quality control levels met established criteria of US Food and Drug Administration guidelines. The assay is suitable for measuring accurate esomeprazole and naproxen plasma concentrations in human bioequivalence study following combined administration.

Nanostructured conducting molecularly imprinted polymer for selective uptake/release of naproxen by the electrochemically controlled sorbent.

A conducting molecularly imprinted polymer (CMIP) film, based on polypyrrole, was electrosynthesized for selective uptake/release and determination of naproxen. The film was prepared by incorporation of a template anion (naproxen) during the electropolymerization of pyrrole into a platinum electrode using the cyclic voltammetry method. Overoxidized polypyrrole films with cavities complementary to the template were used as a potential-induced selective recognition element in the solid-phase sorbent. Various important fabricating factors, which control the performance of the CMIP film, were investigated using fluorescence spectroscopy. The measured fluorescence intensities of released solutions were related to the concentrations of naproxen taken up into the films. Several key parameters such as applied potential and time for uptake and release were varied to achieve the optimal sorption procedure. The film template with naproxen exhibited excellent selectivity over some interference. The calibration graphs were linear in the ranges of 5 × 10⁻8 to 3 × 10⁻7 mol ml⁻¹ and 7 × 10⁻6 to 8 × 10⁻4 mol ml⁻¹, and the limit of detection was 1 × 10⁻8 mol ml⁻¹. The CMIP films, as the electrochemically controlled solid-phase sorbent, were applied for the selective cleanup and quantification of trace amounts of naproxen from physiological samples. Scanning electron microscopy confirmed the nanostructure morphology of the films.

Low-dose naproxen interferes with the antiplatelet effects of aspirin in healthy subjects: recommendations to minimize the functional consequences.

OBJECTIVE: To investigate whether low-dose naproxen sodium (220 mg twice a day) interferes with aspirin's antiplatelet effect in healthy subjects. METHODS: We performed a crossover, open-label study in 9 healthy volunteers. They received for 6 days 3 different treatments separated by 14 days of washout: 1) naproxen 2 hours before aspirin, 2) aspirin 2 hours before naproxen, and 3) aspirin alone. The primary end point was the assessment of serum thromboxane B(2) (TXB(2)) 24 hours after the administration of naproxen 2 hours before aspirin on day 6 of treatment. In 5 volunteers, the rate of recovery of TXB(2) generation (up to 72 hours after drug discontinuation) was assessed in serum and in platelet-rich plasma stimulated with arachidonic acid (AA) or collagen. RESULTS: Twenty-four hours after the last dosing on day 6 in volunteers receiving aspirin alone or aspirin before naproxen, serum TXB(2) was almost completely inhibited (median [range] 99.1% [97.4-99.4%] and 99.1% [98.0-99.7%], respectively). Naproxen given before aspirin caused a slightly lower inhibition of serum TXB(2) (median [range] 98.0% [90.6-99.4%]) than aspirin alone (P = 0.0007) or aspirin before naproxen (P = 0.0045). All treatments produced a maximal inhibition of AA-induced platelet aggregation. At 24 hours, compared with baseline, collagen-induced platelet aggregation was still inhibited by aspirin alone (P = 0.0003), but not by aspirin given 2 hours before or after naproxen. Compared with administration of aspirin alone, the sequential administration of naproxen and aspirin caused a significant parallel upward shift of the regression lines describing the recovery of platelet TXB(2). CONCLUSION: Sequential administration of 220 mg naproxen twice a day and low-dose aspirin interferes with the irreversible inhibition of platelet cyclooxygenase 1 afforded by aspirin. The interaction was smaller when giving naproxen 2 hours after aspirin. The clinical consequences of these 2 schedules of administration of aspirin with naproxen remain to be studied in randomized clinical trials.

Pre-cut dried blood spot (PCDBS): an alternative to dried blood spot (DBS) technique to overcome hematocrit impact.

Quantification of analytes by Dried Blood Spots (DBS) on different paper cards has been extensively reported in the past several years. However, some factors limit the robustness of the precision and accuracy of DBS such as: hematocrit level, blood viscosity, analyte nature, spotting technique and spotting conditions. As such, the paper material used for DBS must meet strict quality control criteria to produce reliable quantification of drugs: uniformity, no chemical leaching and no chromatographic effect. To overcome these variables, especially the hematocrit impact, a modification of the traditional DBS, named Pre-Cut Dried Blood Spot (PCDBS), is presented. In contrast to the classical DBS technique, the new PCDBS procedure demonstrates no variation in response, within ±3%, independently of the hematocrit level or of the type of card used. The impact of the hematocrit level on the analyte recovery is discussed for both DBS and PCDBS approaches. Moreover, for quantification of naproxen by liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS), the PCDBS technique was demonstrated to be as precise (%CV ≤3.1%) and accurate (%nominal between 95.4 and 104.4%) as the classical DBS procedure.

Preparation, in vitro characterization and in vivo release of naproxen loaded in poly-caprolactone nanoparticles.

Naproxen was loaded in poly-caprolactone (PCL) nanoparticles as an implantable sustained release system to prolong its anti-inflammatory activity. Naproxen-loaded nanoparticles were produced with the following characteristics: Nanometric size (< 300 nm), negative zeta potential, low polydispersity index (< 0.1), satisfactory encapsulation efficiency, low water content (< 1%), and spherical shape. In vitro naproxen release profile was sustained and the kinetic followed the Higuchi model. The PCL nanoparticles containing about 12.5% (w/w) of the naproxen (sample A3) was chosen for complementary studies of stability and in vivo release in rats. Nanoparticles did not suffer alteration during stability studies. In vivo release was sustained by one month. Thus, nanoparticles showed potential to act as an implantable sustained release system for chronic inflammatory diseases use.

The ethanol metabolite acetaldehyde increases paracellular drug permeability in vitro and oral bioavailability in vivo.

Alcohol consumption leads to the production of the highly reactive ethanol metabolite, acetaldehyde, which may affect intestinal tight junctions and increase paracellular permeability. We examined the effects of elevated acetaldehyde within the gastrointestinal tract on the permeability and bioavailability of hydrophilic markers and drug molecules of variable molecular weight and geometry. In vitro permeability was measured unidirectionally in Caco-2 and MDCKII cell models in the presence of acetaldehyde, ethanol, or disulfiram, an aldehyde dehydrogenase inhibitor, which causes acetaldehyde formation when coadministered with ethanol in vivo. Acetaldehyde significantly lowered transepithelial resistance in cell monolayers and increased permeability of the low-molecular-weight markers, mannitol and sucrose; however, permeability of high-molecular-weight markers, polyethylene glycol and inulin, was not affected. In vivo permeability was assessed in male Sprague-Dawley rats treated for 6 days with ethanol, disulfiram, or saline alone or in combination. Bioavailability of naproxen was not affected by any treatment, whereas that of paclitaxel was increased upon acetaldehyde exposure. Although disulfiram has been shown to inhibit multidrug resistance-1 P-glycoprotein (P-gp) in vitro, our data demonstrate that the known P-gp substrate paclitaxel is not affected by coadministration of disulfiram. In conclusion, we demonstrate that acetaldehyde significantly modulates tight junctions and paracellular permeability in vitro as well as the oral bioavailability of low-molecular-weight hydrophilic probes and therapeutic molecules in vivo even when these molecules are substrates for efflux transporters. These studies emphasize the significance of ethanol metabolism and drug interactions outside of the liver.