Simultaneous determination of eight analytes of Fuzheng Huayu recipe in beagle dog plasma by UHPLC–Q/exactive Orbitrap HRMS and its application to toxicokinetics

Fuzheng Huayu recipe (FZHY) is a Chinese patent medicine for the treatment of liver fibrosis. This study aimed to investigate the toxicokinetics of FZHY in beagle dogs after oral administration. Blood samples were collected on days 1, 15 and 28 after oral gavage of FZHY dosages of 400 or 1,200 mg/kg body weight once a day. A UHPLC–Q-Orbitrap method was developed and validated to simultaneously determine and quantify eight components of FZHY in beagle dog plasma. The times to peak concentration for eight components were18–120 min. The peak concentrations (C_max) of amygdalin, genistein, daidzein and 3,4-dihydroxybenzaldehyde were 1.43–43.50 ng/ml, the areas under the concentration–time curve (AUC_(0–t)) were 2.45–6,098.25 ng min/ml, and the apparent volumes of distribution (V_d) were 0.05–131.23 × 10⁴ ml/kg. The values of C_max of prunasin, schisantherin A, schisandrin A and schisandrin were 7.35–1,450.73 ng/ml, the values of AUC_(0–t) were 3,642.30–330,388.65 ng min/ml, and the values of V_d were 11.15–1,087.18 × 10⁴ ml/kg. No obvious accumulation of the eight compounds was observed in beagle dogs. The results showed that the method is rapid, accurate and sensitive, and is suitable for detecting the eight analytes of FZHY. This study provides an important basis for the assessment of FZHY safety.     

Enhanced Oral Bioavailability of β-Caryophyllene in Healthy Subjects Using the VESIsorb® Formulation Technology,a Novel Self-Emulsifying Drug Delivery System (SEDDS)

β-Caryophyllene (BCP), a common constituent of many spice and food plants, is gaining increased attention due to recent research identifying numerous potential health benefits. Due to limited oral bioavailability observed in preclinical models, the described benefits of BCP may be maximized by using a suitable delivery system. Additionally, human pharmacokinetics (PK) remain unknown. This study evaluates the relative oral bioavailability of BCP formulated in a self-emulsifying drug delivery system (SEDDS) based on VESIsorb^® formulation technology (BCP-SEDDS) compared to BCP neat oil. Hence, a randomized, double-blind, cross-over design, single oral dose study (100 mg BCP) in 24 healthy subjects (12 men/12 women) was performed under fasting conditions. Pharmacokinetic parameters were analyzed from individual concentration-time curves. The data show that BCP-SEDDS resulted in a 2.2/2.0-fold increase in AUC_0–12h/AUC_0–24h and a 3.6-fold increase in C_max compared to BCP neat oil. Moreover, BCP was absorbed faster from BCP-SEDDS (T_max: 1.43 h) compared to BCP neat oil (T_max: 3.07 h). Gender analysis revealed that there is no significant difference between men and women for both the investigated formulations and all investigated PK endpoints. In conclusion, BCP-SEDDS offers a well-tolerated and effective oral delivery system to significantly enhance the oral bioavailability of BCP in humans.     

Comparative pharmacokinetics studies of benzoylhypaconine,benzoylmesaconine, benzoylaconine and hypaconitine in rats by LC‐MS method after administration of Radix Aconiti Lateralis Praeparata extract and Dahuang Fuzi Decoction

A rapid and sensitive high‐performance liquid chromatography–mass spectrometric (HPLC‐MS) method was developed and validated for simultaneous determination of benzoylhypaconine (BHA), benzoylmesaconine (BMA), benzoylaconine (BAC) and hypaconitine (HA) in rat plasma for the first time. The analytes were separated on a Kromasil C₁₈ column with a total running time of 11 min. The validation data demonstrated a sound feasibility for the newly developed method and it was then applied to the pharmacokinetic study of these analytes in rats. Pharmacokinetic behaviors of BHA, BMA, BAC and HA in rats were studied after oral administration of Radix Aconiti Lateralis Praeparata extract (FZ) and Dahuang Fuzi Decoction (DFD). The main parameters for the two groups of subjects were compared, and significant differences between Radix Aconiti Lateralis Praeparata extract group and Dahuang Fuzi Decoction group in calculated parameters, such as the area under the plasma concentration–time from zero to the last quantifiable time‐point (AUC_0–t), the area under the plasma concentration–time curve from zero to infinity (AUC_0–∞), peak plasma concentration (C_max), half‐life of elimination (T_1/2), mean retention time (MRT_0–t), plasma clearance (CL), volume of distribution (V_d) and time to reach C_max (T_max), were found. After oral administration of DFD, the AUC_0–t, AUC_0–∞ and C_max of BHA, BMA, BAC and HA decreased remarkably (p < 0.05) compared with those of the FZ extract group. V_d and CL values of BHA, BMA, BAC and HA increased, two of which showed significant difference (p < 0.05). T_1/2 and MRT_0–t values of BHA, BMA and BAC in the DFD group were significantly delayed compared with those of FZ extract group. Only the T_max of HA, the toxic ingredient in FZ, delayed significantly in DFD group compared with the value of FZ group. All these pharmacokinetic parameters were statistically compared, and the rationality of the combination for DFD was clearly demonstrated. Copyright © 2013 John Wiley & Sons, Ltd.     

LC-ESI-MS/MS method for quantification of ambrisentan in plasma and application to rat pharmacokinetic study

A sensitive high‐performance liquid chromatography–positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of ambrisentan in plasma. The analyte and the internal standard (armodafinil) were extracted from plasma by acetonitrile precipitation and they were separated on a reversed‐phase C₁₈ column with a gradient program. The MS acquisition was performed with multiple reaction monitoring mode using the respective [M + H]⁺ ions, m/z 379–347 for ambrisentan and m/z 274–167 for the IS. The assay exhibited a linear dynamic range of 1–2000 ng/mL for ambrisentan in plasma. Acceptable precision (<10%) and accuracy (100 ± 8%) were obtained for concentrations over the standard curve range. The method was successfully applied to quantify ambrisentan concentrations in a rodent pharmacokinetic study after a single oral administration of ambrisentan at 2.5 mg/kg to rats. Following oral administration the maximum mean concentration in plasma (C_max; 1197 ± 179 ng/mL) was achieved at 1.0 ± 0.9 h (T_max), and the area under the curve (AUC) was 6013 ± 997 ng h/mL. Therefore, development of such a simple and sensitive method in rat plasma should translate into a method for ambrisentan in human plasma for clinical trials. Copyright © 2012 John Wiley & Sons, Ltd.     

Pharmacokinetics of aconitine in rat skin after oral and transdermal gel administrations

The purpose of this study was to evaluate percutaneous penetration and arrhythmogenic effects of aconitine after transdermal administration, compared with the oral route. Skin penetration of aconitine was tested by a microdialysis technique in rats and in vivo recovery was determined by retrodialysis. After oral and transdermal administration of aconitine, dialysate was sampled at 20 min intervals until the end of the experiment for the determination of concentration of aconitine in skin. Blood samples were collected and analyzed using a validated HPLC‐MS/MS method. In addition, we concurrently recorded the electrocardiogram (ECG). The in vivo recovery of aconitine in the skin was calculated to be 39.59%. The C_max values for aconitine absorbed into the skin after oral and transdermal administration were 1.51 ± 0.53 and 2723.8 ± 848.8 ng/mL, respectively, and within the plasma, 215.86 ± 79.29 and 20.92 ± 3.15 ng/mL. The C_max value for the plasma concentration of aconitine after oral administration was approximately 10 times higher than with the transdermal route. For oral administration, the ECG revealed various types of arrhythmias at a period of T_max, which is normal in transdermal gel administration. These results indicate that transdermal aconitine gel is a safe formulation that can deliver the drug in sufficient amounts and safe concentrations to produce therapeutic action in rats. Copyright © 2011 John Wiley & Sons, Ltd.     

The effect of the electrolyte concentration in the solution on the voltammetric response of insertion electrodes

A solid-state redox reaction involving an insertion of ions is analyzed with respect to the influence of the concentration of inserting ions in the solution phase. The voltammetric response is independent of the mass transfer in the solution provided that z = (D _ss/D _aq)^1/2 ρ/[C⁺]* is smaller than 0.1 (D _ss: diffusion coefficient of the cation C⁺ in the crystal; D _aq: diffusion coefficient of the cation C⁺ in the solution; ρ: density of the solid compound; [C⁺]*: concentration of cations in the bulk of the solution). In real cases this condition will be satisfied at solution concentrations above 1 mol/l. Received: 15 December 1997 / Accepted: 5 March 1998     

Hollow-Fiber-Based LPME as a Reliable Sampling Method for Gas-Chromatographic Determination of Pharmacokinetic Parameters of Valproic Acid in Rat Plasma

A sensitive and simple method based on two-phase liquid-phase microextraction in porous hollow fiber followed by gas chromatography-flame ionization detection was developed for quantification and pharmacokinetic study of valproic acid (VPA, an antiepileptic drug) in rat plasma after oral administration of pure sodium valproate (25 mg kg⁻¹). Some parameters such as type of organic solvent, pH of sample solution, stirring speed, salt addition, extraction time, and volume of sample that affected extraction efficiency of VPA were optimized. Under optimized microextraction conditions, VPA was extracted with 10 μL 1-octanol from 0.5 mL rat plasma previously diluted with 4.5 mL acidified and salinated water (pH 2) using 1-octanoic acid as internal standard. The limit of detection was 17 ng mL⁻¹ with linear response over the concentration range of 50–10,000 ng mL⁻¹ with correlation coefficient higher than 0.998. The developed method was successfully applied to determination of pharmacokinetic parameters such as t _max (peak time in concentration–time profile), C _max (peak concentration in concentration–time profile), t _1/2 (elimination half-life), AUC_0–t (area under the curve for concentration versus time), clearance, and apparent distribution volume in rats following oral administration of VPA.

     

Simultaneous in vivo RP-HPLC-DAD quantification of multiple-component and drug-drug interaction by pharmacokinetics, using 6,7-dimethylesculetin, geniposide and rhein as examples

Increasing evidence has demonstrated that multidrug combinations could amplify the therapeutic efficacies of each agent. Interestingly, the pharmacological effect of traditional Chinese medicine (TCM) is usually attributed to the drug‐interaction property (synergism) of multiple active constituents. Pharmacokinetics is a useful means of evaluating the drug interactions of major active compounds in TCM. A simple, sensitive and reliable RP‐HPLC‐DAD method has been developed to simultaneously quantify 6,7‐dimethylesculetin (D), geniposide (G) and rhein (R), which are the active ingredients in Yin–Chen–Hao–Tang (YCHT), performing drug‐interaction pharmacokinetics studies in vivo. Plasma samples were prepared using methanolic precipitation, a filtration step, and then injection of the methanolic extract onto a Nova‐Pak C₁₈ Guard‐Pak? guard column with a gradient mobile phase. Triple‐wavelength diode array detection was set at λ_max values of 343 nm for D, 241 nm for the G, and 259 nm for R. Our results successfully demonstrate that this method has excellent and satisfactory selectivity, sensitivity, linearity, precision, accuracy and recovery. In healthy rats, the estimated pharmacokinetic parameters (i.e. C_max, AUC and Cl) of D, G and R, when administered with COC (a combination of D, G and R), were C_max 16.05 mg/L, AUC 108.96 mg h/L and Cl 0.36 L/h for D; C_max 9.35 mg/L, AUC 64.71 mg h/L and Cl 0.88 L/h for G; and C_max 14.18 mg/L, AUC 57.98 mg h/L and Cl 1.77 L/h for R. Here, we report that the COC combination could significantly increase the plasma level and slow the elimination rate compared with any one or two of the three individual compounds, which may indicate a drug–drug interaction. Copyright © 2011 John Wiley & Sons, Ltd.     

New unsymmetrical difluoroaromatic compounds and estimation of their reactivities in nucleophilic substitution

A series of previously unknown unsymmetrical difluoroaromatic compounds, viz., p-fluorobenzoylphenyl(p-fluorophenyl)-substituted imidazoles, pyrazines, and quinoxalines, were synthesized according to multistep procedures with the use of chloral as the key compound. The reactivities of the resulting difluoroaromatic compounds were estimated based on ¹⁹F and ¹³C NMR spectral data and the results of quantum-chemical calculations. The calculated charge densities on the C_ipso atoms correlate linearly with the experimental chemical shifts in the ¹⁹F and ¹³C NMR spectra. Difluoroaromatic compounds, which are characterized by _F > –110 and _C > 163 and by the charge density on the C_ipso atom higher than 0.08 e, are sufficiently activated to be used for the preparation of high-molecular-weight polyethers.     

Quantification of urapidil, α-1-adrenoreceptor antagonist, in plasma by LC-MS/MS: validation and application to pharmacokinetic studies

A sensitive high‐performance liquid chromatography–positive ion electrospray tandem mass spectrometry method was developed and validated for the quantification of urapidil in plasma. Following liquid–liquid extraction, the analyte was separated using an isocratic mobile phase on a reverse‐phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M + H]⁺ ions, m/z 388 to 205 for urapidil and m/z 452 to 344 for the internal standard. The assay exhibited a linear dynamic range of 0.1–500 ng/mL for urapidil in plasma. Acceptable precision (<7%) and accuracy (100 ± 8%) were obtained for concentrations over the standard curve range. The method was successfully applied to quantify urapidil concentrations in a preclinical pharmacokinetic study after a single oral administration of urapidil at 3 mg/kg to rats. Following oral administration the maximum mean concentration in plasma (C_max; 616 ± 73 ng/mL) was achieved at 0.5 h (T_max) and area under curve (AUC_0–24) was 1841 ± 308 ng h/mL. The half‐life (t_1/2) and clearance (Cl) were 2.47 ± 0.4 h and 1660 ± 276 mL/h/kg, respectively. Moreover, it is plausible that the assay method in rat plasma would facilitate the adaptability of urapidil quantification in human plasma for clinical trials. Copyright © 2011 John Wiley & Sons, Ltd.     

A Conductometric Study of Complexation Reactions Between Dibenzo-18-Crown-6 (DB18C6) with Cu2+, Zn2+, Tl+ and Cd2+ Metal Cations in Dimethylsulfoxide–Ethylacetate Binary Mixtures

The complex formation between Cu²⁺, Zn²⁺, Tl⁺ and Cd²⁺ metal cations with macrocyclic ligand, dibenzo- 18-crown-6 (DB18C6) was studied in dimethylsulfoxide (DMSO)–ethylacetate (EtOAc) binary systems at different temperatures using conductometric method. In all cases, DB18C6 forms 1:1 complexes with these metal cations. The stability constants of the complexes were obtained from fitting of molar conductivity curves using a computer program, Genplot. The non-linear behaviour which was observed for variations of log K _f of the complexes versus the composition of the mixed solvent was discussed in terms of changing the chemical and physical properties of the constituent solvents when they mix with one another and, therefore, changing the solvation capacities of the metal cations, crown ether molecules and even the resulting complexes with changing the mixed solvent composition. The results show that the selectivity order of DB18C6 for the metal cations in pure ethylacetate and pure dimethylsulfoxide is: Tl⁺ > Cu²⁺ > Zn²⁺ > Cd²⁺ but the selectivity order is changed with the composition of the mixed solvents. The values of enthalpy changes (ΔH°_C) for complexation reactions were obtained from the slope of the van’t Hoff plots and the changes in standard enthalpy (ΔS°_C) were calculated from the relationship: ΔG°_C,298.15=ΔH°_C − 298.15 ΔS°_C. The obtained results show that in most cases, the complexes are enthalpy stabilized, but entropy destabilized and the values of ΔH°_C and ΔS°_C depend strongly on the nature of the medium.     

Pharmacokinetics and penetration into synovial fluid of systemical and electroporation administered sinomenine to rabbits

Sinomenine is an anti‐rheumatoid arthritis (RA) drug derived from the Sinomenium acutum. The major site of RA treatment is within the synovial compartment. However, the pharmacokinetic and penetration into synovial fluid (SF) of sinomenine have not been reported. In our study, the pharmacokinetics and penetration into SF of systemic and electroporation administered sinomenine were investigated by microdialysis incorporated with HPLC‐MS/MS. Sinomenine went into plasma and SF more rapidly with higher peak concentration (C_max) by intramuscular injection compared with oral administration. The area under the concentration–time graph (AUC_0–∞) of intramuscularly injected sinomenine was 1,403,294.75 ± 125,534.567 ng min/mL in plasma and 456,116.37 ± 62,648.36 ng min/mL in SF, which were equivalent with those for an oral dose. These results indicated that equal amounts of sinomenine could penetrate into SF by the two administration routes, and the permeation ratios were approximately 1:3. The AUC_0–∞ and C_max were lower with electroporation compared with systemic administration, but the C_SF/C_Plasma (concentration of sinomenine in SF vs that of plasma) at 90, 120, 150, 180, 240 and 480 min by electroporation was 3‐ to 10‐fold higher relative to systemic administration. This illustrated that sinomenine can be targeted into joints by electroporation, and electroporation is a potential technique for sinomenine's transdermal delivery. Copyright © 2014 John Wiley & Sons, Ltd.     

Determination of toosendanin in rat plasma by ultra‐performance liquid chromatography–electrospray ionization–mass spectrometry and its application in a pharmacokinetic study

Toosendanin (TSN) is a major triterpenoid existing in Melia toosendan, which has been used as a digestive tract parasiticide and insecticide but with serious hepatotoxicity. An ultra‐performance liquid chromatography–electrospray ionization–mass spectrometry method was developed for determination of TSN in rat plasma. Plasma samples were separated on Acquity UPLC^TM BEH C₁₈ column with acetonitrile and water as flow phase by gradient elution and determined by quadrupole mass spectrometer in negative selective ion monitoring mode. Usolic acid was used as internal standard. The calibration curves were linear over 0.02–3.0 µg/mL for TSN with a lower limit of quantification (LLOQ) of 20 ng/mL in rat plasma. The extraction recoveries of TSN were within 74.3–80.7% with an accuracy of 94.5–108.9%. The intra‐ and inter‐day precision values of the assay at three quality control levels were 8.8–13.8% and <13.9% at LLOQ level, respectively. The method was successfully applied to a pharmacokinetic study of TSN in rats after a single intravenous and oral administration of 2 and 60 mg/kg. The shorter T_max, higher V_d and Cl of TSN after oral administration indicated that TSN could be absorbed, distributed and eliminated quickly in rats in vivo. The absolute bioavailability of TSN after oral administration was 9.9%. Copyright © 2012 John Wiley & Sons, Ltd.     

SYNTHESIS OF THE FIRST BENZODISELENAGERMOLES AND ONE SPIROBISDISELENAGERMOLE

Abstract

The first examples of compounds R¹R²GeSe₂C₆H₄R³ (R¹,R²=CH³ C₂H₅, C₃H₂, n-C₄H₉, i-C₅H₁₁, Ph, p-CH₃Ph. R³=H, CH₃, OCH₃) were easily obtained (40–80% yield) from electrophilic cleavage of diselenophenylene zirconocenes by dialkyl or diaryl dichlorogermanes. The synthesis of a spirodi-selenagermole was achieved in the same way using germanium tetrachloride. Analytical data, ¹H and ⁷⁷Se NMR. mass spectra are perfectly consistent with the expected structures.     

The density of interface states and their relaxation times in Au/Bi4Ti3O12/SiO2/n‐Si(MFIS) structures

Bismuth titanate (Bi₄Ti₃O₁₂) (BTO) thin films were fabricated on an n‐type Si substrate and annealed by rapid thermal annealing methods. The I‐V measurement shows that the device has properties of Schottky diode with the Φ_B0 of 0.76 eV, n of 2.42, and leakage current of about 10^?7 A at ? 8 V. The experimental C‐V‐f and G/w‐V‐f characteristics of metal‐ferroelectric‐insulator‐semiconductor (MFIS) structures show fairly large frequency dispersion especially at low frequencies due to interface states N_ss. The energy distribution of (N_ss) has been determined by using the high‐low frequency capacitance (C_HF? C_LF) and conductance method. The relaxation time (τ) of interface states was calculated from the conductance method. It has been shown that both the N_ss and relaxation time increase almost exponentially with bias, which activates traps located at deeper gap energies. Copyright © 2011 John Wiley & Sons, Ltd.     

Method development and validation for simultaneous determination of ebastine and its active metabolite carebastine in human plasma by liquid chromatography–tandem mass spectrometry and its application to a clinical pharmacokinetic study in healthy Chinese volunteers

A simple LC–tandem mass spectrometry (MS/MS) method to determine ebastine and carebastine (active metabolite) in human plasma was developed and validated. Analytes and internal standards were precipitated by protein precipitation and separated on Synergi Hydro-RP 80A column (4 μm, 50 mm × 2.0 mm; Phenomenex) by gradient elution with mobile phase A comprising 0.1% formic acid in 5 mm ammonium acetate (NH₄Ac) and B comprising 100% methanol at a flow rate 0.4 mL/min. Ions were detected in positive multiple reaction monitoring mode, and they exhibited linearity over concentration range 0.01–8.0 and 1.00–300 ng/mL for ebastine and carebastine, respectively. A clinical pharmacokinetic study was conducted in healthy Chinese volunteers under fasting and fed conditions after a single oral administration of 10 mg ebastine. The maximum plasma concentration (C_max), time to C_max (T_max) and elimination half-life for ebastine were 0.679 ± 0.762 ng/mL, 1.67 ± 1.43 h and 7.86 ± 6.18 h, respectively, whereas these for carebastine were 143 ± 68.4 ng/mL, 5.00 ± 2.00 h and 17.4 ± 4.97 h, respectively under fasting conditions; the corresponding values under fed conditions were 4.13 ± 2.53 ng/mL, 3.18 ± 1.09 h and 21.6 ± 7.77 h for ebastine and 176 ± 68.4 ng/mL, 6.14 ± 2.0 h and 20.0 ± 4.97 h for carebastine.     

Photochromic Transformations of Spiro Compounds: 1. A New Approach to Determining the Photocoloration Quantum Yield

A new approach to determining a photocoloration quantum yield for photochromic compounds was considered. This approach is based on comparison between the calculated and experimental values of maximum absorbance A ^B _maxof a photocolored form upon monochromatic irradiation. Using spirooxazines as an example, the quantum yield of photocoloration was determined, and A ^B _maxwas examined as a function of a number of parameters that characterize the photocolored form (the quantum yields of photocoloring and photobleaching and the lifetime of the colored form). It was found that A ^B _maxnonlinearly increased with decreasing rate constant (<0.2 s^–1) of the thermal bleaching of spirooxazines.     

Toxicokinetics of strychnine and brucine after the oral administration of Biqi capsule to rats by RRLC–MS/MS

Biqi capsule is a well‐known traditional Chinese medicine formula that has been widely applied for the clinical treatment of such diseases as rheumatoid arthritis, scapulohumeral periarthritis and cervical spondylopathy. However, there is concern regarding the toxicity of Biqi capsule owing to its active ingredients, strychnine and brucine. To investigate the toxicokinetics of strychnine and brucine after oral administration of Biqi capsule to rats, a sensitive and simple rapid‐resolution liquid chromatography/tandem mass spectrometry method was developed to determine the levels of strychnine and brucine in rat plasma. Chromatographic separation was performed on a Capcell Pak C₁₈ MG II (3.0 μm, 2.0 × 35 mm) column by gradient elution with acetonitrile and 0.2% formic acid as the mobile phase. The method was validated over the range of 0.25–250 ng/mL for strychnine and 0.025–25 ng/mL for brucine. The intra‐ and inter‐day accuracies of strychnine and brucine in rat plasma were 100.3–106.6 and 90.75–106.1% respectively, and the precisions were within 14.2%. The established method was successfully applied to the toxicokinetic study of strychnine and brucine after single and multiple oral administration of Biqi capsule to male and female rats at 0.4, 0.8 and 1.6 g/kg doses. The results showed different toxicokinetic characteristics in the different groups.     

A self‐contrast approach to evaluate the inhibitory effect of chrysosplenetin,in the absence and presence of artemisinin,on the in vivo P‐glycoprotein‐mediated digoxin transport activity

In this study, we used a self‐contrast method, which excluded the individual difference, to evaluate the inhibitory effect of chrysosplentin (CHR) in the presence or absence of artemisinin (ART) on the P‐glycoprotein (P‐gp) transport activity. A sensitive and rapid UHPLC–MS/MS method was applied for quantification of digoxin, a P‐gp‐specific substrate, in rat plasma. A pharmacokinetic study was carried out: first after an oral administration of digoxin at a dose of 0.09 mg/kg (first period), followed by a 20‐day wash‐out, then after another administration of digoxin (second period). During the second period, test compounds were orally given three times per day for seven consecutive days. Results showed that the t_1/2 of digoxin in all the groups had no significant difference between the first and second periods. The AUC_0–24, C_max, t_max, and Cl_z/F of the negative control and ART alone groups showed no difference. However, the AUC_0–24 and C_max in the CHR alone, CHR–ART (1:2) and verapamil (positive control) groups showed 2.34‐, 3.04‐, 1.79‐, and 1.81‐, 1.99‐, 2.06‐fold increases along with 3.50‐, 3.84‐ and 4.76‐fold decreases for CL_z/F, respectively. The t_max in the CHR–ART (1:2) group increased 3.73‐fold. In conclusion, our self‐contrast study suggested that CHR, especially when combined with ART in a ratio of 1:2, inhibited P‐gp activity while ART alone has no effect. Copyright © 2016 John Wiley & Sons, Ltd.