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CYP2B subfamily accounts for 2-10% of total hepatic CYP450 enzymes and participate in the metabolism of around 8% of clinical drugs. Borneol has been widely used in traditional Chinese medicine for thousands of years. There are many studies about borneol-induced promoting penetration role for a number of drugs through various physiologic barriers, whereas there is no report involved the effect of borneol on hepatic CYP2B. The present work studied the in vivo effect of borneol on the expression and activity of rat hepatic CYP2B. The results indicated that the oral administration of borneol (33, 100 and 300 mg/kg/d) to rats for consecutive 7 days increased the hepatic CYP2B1/2 activity by 1.4-, 1.7- and 2.8-fold, hepatic CYP2B1 mRNA expression by 6.3-, 8.7- and 18.1-fold, and hepatic CYP2B1/2 protein expression by 1.2-, 1.9- and 2.6-fold, respectively compared to the control. Additionally, in the borneol pre-dosing (300 mg/kg/d for consecutive 7 days) rats, the increased Cl int and decreased AUC 0-24 of bupropion were observed as compared to the control. Moreover, there were no obvious effects on CAR protein level in rat liver microsome and nucleus following the borneol treatment. Taken together, our observations indicate that borneol is an in vivo inducer of rat hepatic CYP2B with different regulatory mechanism from phenobarbital-like inducers which caused CYP2B induction with CAR activation., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Background: Bupropion is used for treatment of depression during pregnancy. However, its use as a smoking cessation aid for pregnant women is currently under evaluation., Objective: The aim of this opportunistic study was to investigate the transfer of bupropion and its major pharmacologically active metabolites, hydroxybupropion and threohydrobupropion, across the placenta in vivo. In addition, the concentrations of the drug and its metabolites were determined in the amniotic fluid., Study Design: The following samples were collected at deliveries from 22 women taking bupropion: maternal blood (n = 22), umbilical cord venous blood (n = 22), and amniotic fluid (n = 9). The concentrations of the drug and its metabolites in blood plasma and amniotic fluid were determined by means of liquid chromatography-mass spectrometry. Placental passage was calculated as a ratio of umbilical cord venous plasma to maternal plasma concentrations., Results: The levels of hydroxybupropion and threohydrobupropion in umbilical cord venous plasma were invariably lower than their corresponding concentrations in maternal plasma. The concentrations of bupropion in umbilical cord plasma were lower than in maternal plasma in the majority of the maternal-cord blood pairs. The median values of the umbilical cord venous plasma to maternal plasma ratios were: bupropion, 0.53 (interquartile range 0.35, n = 18), hydroxybupropion, 0.21 (interquartile range 0.12, n = 18), and threohydrobupropion, 0.61 (interquartile range 0.11, n = 21). In umbilical cord venous plasma, the median concentration of bupropion was 5.3 ng/mL; hydroxybupropion, 103.6 ng/mL; and threohydrobupropion, 59.6 ng/mL. Bupropion and its metabolites were detectable in the amniotic fluid but the concentrations of threohydrobupropion were higher than those in the corresponding umbilical cord venous plasma., Conclusion: Bupropion and its active metabolites cross the placenta to the fetal circulation. The concentrations of hydroxybupropion and threohydrobupropion in umbilical cord venous plasma were higher than bupropion concentrations suggesting a higher fetal exposure to the metabolites than the parent drug. The higher levels of threohydrobupropion in the amniotic fluid than those in umbilical cord venous plasma suggest that enzymes involved in the metabolism of bupropion to threohydrobupropion are most likely active in the fetus. The biological consequences of fetal exposure to maternally administered bupropion and/or its active metabolites via placental transfer and recirculation of the amniotic fluid are yet to be determined., (Copyright © 2016. Published by Elsevier Inc.)

A stereoselective assay was developed for the quantification of bupropion and oxidative, reductive, and glucuronide metabolites (16 analytes total) in human urine. Initially, authentic glucuronide standards obtained from commercial sources were found to be incorrectly labeled with regard to stereochemistry; the correct stereochemistry was unequivocally reassigned. A trifurcated urine sample preparation and analysis procedure was employed for the stereoselective analysis of bupropion, hydroxybupropion, erythrohydrobupropion, and threohydrobupropion enantiomers, and hydroxybupropion, erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers in urine. Method 1 stereoselectively analyzed bupropion (R and S), and unconjugated free hydroxybupropion (R,R and S,S), erythrohydrobupropion (1R,2S and 1S,2R), and threohydrobupropion (1R,2R and 1S,2S) using chiral chromatography with an α1-acid glycoprotein column. Because no hydroxybupropion β-d-glucuronide standards were commercially available, method 2 stereoselectively analyzed total hydroxybupropion aglycones (R,R and S,S-hydroxybupropion) after urine hydrolysis by β-glucuronidase. Hydroxybupropion β-d-glucuronide (R,R and S,S) urine concentrations were calculated as the difference between total and free hydroxybupropion (R,R and S,S) concentrations. Due to incomplete β-glucuronidase hydrolysis of erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers, method 3 stereoselectively analyzed intact erythrohydrobupropion and threohydrobupropion β-d-glucuronide diastereomers using C18 column chromatography. All analytes were quantified by positive ion electrospray tandem mass spectrometry. The assay was fully validated over analyte-specific concentrations. Intra- and inter assay precision were within 15% for each analyte. The limits of quantification for bupropion (R and S), hydroxybupropion (R,R and S,S), threohydrobupropion (1S,2S and 1R,2R), erythrohydrobupropion (1R,2S and 1S,2R) were 10, 50, 100, and 100ng/mL, respectively. The limits of quantification for (1R,2R)-threohydrobupropion β-d-glucuronide, (1S,2S)-threohydrobupropion β-d-glucuronide, and (1R,2R)-erythrohydrobupropion β-d-glucuronide were each 50ng/mL. Due to the abundance of bupropion and metabolites in human urine, no efforts were made to optimize sensitivity. All analytes were stable following freeze thaw cycles at -80°C. This assay was applicable to clinical pharmacokinetic investigations of bupropion in patients and to in vitro metabolism of the primary bupropion metabolites to their glucuronides., (Copyright © 2016 Elsevier B.V. All rights reserved.)

A stereoselective analytical method was developed and validated for the quantification of bupropion, and principle metabolites hydroxybupropion, erythrohydrobupropion and threohydrobupropion in human plasma. Separation of individual enantiomers (R)-bupropion, (S)-bupropion, (R,R)-hydroxybupropion, (S,S-hydroxybupropion), (1S,2S)-threohydrobupropion, (1R,2R)-threohydrobupropion, (1R,2S)-erythrohydrobupropion, and (1S,2R)-erythrohydrobupropion was achieved utilizing an α1-acid glycoprotein column within a 12-min run time. Chromatograph separation was significantly influenced by mobile phase pH and variability between columns. Analytes were quantified by positive ion electrospray tandem mass spectrometry following plasma protein precipitation with 20% trichloroacetic acid. Identification of erythrohydrobupropion enantiomer peaks and threohydrobupropion enantiomer peaks was achieved by sodium borohydride reduction of enantiopure (R)- and (S)-bupropion. Initial assay validation and sensitivity determination was on AB Sciex 3200, 4000 QTRAP, and 6500 mass spectrometers. Accuracy and precision were within 15% for each analyte. The assay was fully validated over analyte-specific concentrations using an AB Sciex 3200 mass spectrometer. Intra- and inter-assay precision and accuracy were within 12% for each analyte. The limits of quantification for bupropion (R and S), hydroxybupropion (R,R and S,S), threohydrobupropion (1S,2S and 1R,2R), and erythrohydrobupropion (1R,2S and 1S,2R) were 0.5, 2, 1, and 1ng/mL, respectively. All analytes were stable following freeze thaw cycles at -80°C and while stored at 4°C in the instrument autosampler. This method was applicable to clinical pharmacokinetic investigations of bupropion in patients. This is the first chromatographic method to resolve erythrohydrobupropion and threohydrobupropion enantiomers, and the first stereoselective LC-MS/MS assay to quantify bupropion, and principle metabolites hydroxybupropion, erythrohydrobupropion, and threohydrobupropion in human plasma., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Bupropion metabolites formed via oxidation and reduction exhibit pharmacological activity, but little is known regarding their stereoselective disposition. A novel stereoselective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to separate and quantify enantiomers of bupropion, 4-hydroxybupropion, and erythro- and threo-dihydrobupropion. Liquid-liquid extraction was implemented to extract all analytes from 50 μL human plasma. Acetaminophen (APAP) was used as an internal standard. The analytes were separated on a Lux 3 μ Cellulose-3 250×4.6 mm column by methanol: acetonitrile: ammonium bicarbonate: ammonium hydroxide gradient elution and monitored using an ABSciex 5500 QTRAP triple-quadrupole mass spectrometer equipped with electrospray ionization probe in positive mode. Extraction efficiency for all analytes was ≥70%. The stability at a single non-extracted concentration for over 48 h at ambient temperature resulted in less than 9.8% variability for all analytes. The limit of quantification (LOQ) for enantiomers of bupropion and 4-hydroxybupropion was 0.3 ng/mL, while the LOQ for enantiomers of erythro- and threo-hydrobupropion was 0.15 ng/mL. The intra-day precision and accuracy estimates for enantiomers of bupropion and its metabolites ranged from 3.4% to 15.4% and from 80.6% to 97.8%, respectively, while the inter-day precision and accuracy ranged from 6.1% to 19.9% and from 88.5% to 99.9%, respectively. The current method was successfully implemented to determine the stereoselective pharmacokinetics of bupropion and its metabolites in 3 healthy volunteers administered a single 100mg oral dose of racemic bupropion. This novel, accurate, and precise HPLC-MS/MS method should enhance further research into bupropion stereoselective metabolism and drug interactions., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Bupropion is widely used as an antidepressant drug and also as a smoking cessation aid. In humans, this drug is extensively metabolized to form several metabolites. Oxidised hydroxybupropion and two reduced metabolites, threohydrobupropion and erythrohydrobupropion, are major metabolites. All of these metabolites are considered to be active. Although the oxidative metabolic pathway and the central role of CYP2B6 are known, the enzymes that participate in the reduction have not been identified to date. The aim of this study was to confirm the role of human liver subcellular fractions in the metabolism of bupropion and elucidate the contribution of particular carbonyl-reducing enzymes. An HPLC method for the determination of bupropion metabolites was utilised. Bupropion is reduced to threohydrobupropion and less to erythrohydrobupropion in human liver cytosol, microsomes and also mitochondria. Surprisingly, intrinsic clearance for formation of both metabolites is the highest in mitochondrial fraction. Moreover this study provides the first direct evidence that 11β-hydroxysteroid dehydrogenase 1, AKR1C1, AKR1C2, AKR1C3 and CBR1 participate in the reducing biotransformation of bupropion in vitro. The enzyme kinetics of all of these reductases was investigated and kinetic parameters were calculated.

Crystal phase transition and isothermal crystallization kinetics of bupropion hydrobromide is studied by thermal analysis, X-ray powder diffractometry, and scanning electron microscopy. As well known, bupropion hydrobromide has two stable polymorphic forms, form I and form II, during the production. Here it is found that form II will convert into form I with thermal treatment. Avrami exponent n is evaluated to be around 2.0 in the range 170°C-190°C, which indicates that the phase transition is a process of one-dimensional nucleation growth. This viewpoint is also confirmed by microscope morphology study. In addition, phase-transition active energy is calculated to be 239.4 kJ/mol, which means that a high energy barrier needs to be overcome to start up the phase transition., (Copyright © 2011 Wiley Periodicals, Inc.)

A liquid chromatography method coupled to mass spectrometry was developed for the quantification of bupropion, its metabolite hydroxy-bupropion, moclobemide, reboxetine and trazodone in human plasma. The validation of the analytical procedure was assessed according to Société Française des Sciences et Techniques Pharmaceutiques and the latest Food and Drug Administration guidelines. The sample preparation was performed with 0.5 mL of plasma extracted on a cation-exchange solid phase 96-well plate. The separation was achieved in 14 min on a C18 XBridge column (2.1 mm×100 mm, 3.5 μm) using a 50 mM ammonium acetate pH 9/acetonitrile mobile phase in gradient mode. The compounds of interest were analysed in the single ion monitoring mode on a single quadrupole mass spectrometer working in positive electrospray ionisation mode. Two ions were selected per molecule to increase the number of identification points and to avoid as much as possible any false positives. Since selectivity is always a critical point for routine therapeutic drug monitoring, more than sixty common comedications for the psychiatric population were tested. For each analyte, the analytical procedure was validated to cover the common range of concentrations measured in plasma samples: 1-400 ng/mL for reboxetine and bupropion, 2-2000 ng/mL for hydroxy-bupropion, moclobemide, and trazodone. For all investigated compounds, reliable performance in terms of accuracy, precision, trueness, recovery, selectivity and stability was obtained. One year after its implementation in a routine process, this method demonstrated a high robustness with accurate values over the wide concentration range commonly observed among a psychiatric population., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A specific and highly sensitive liquid chromatography-electrospray mass spectrometry (LC-ESI-MS) method for the direct determination of bupropion (BUP) and its main metabolite hydroxybupropion (HBUP) in rat plasma and brain microdialysate has been developed and validated. The analysis was performed on a Bonus RP C18 (100 mm × 2.1mm i.d., 3.5 μm particles) column using gradient elution with the mobile phase consisting of acetonitrile and ammonium formate buffer (10mM, pH 4). Plasma samples were analyzed after a simple, one-step protein precipitation clean-up with trichloroacetic acid (TCA), however clean-up for microdialysis samples was not necessary, enabling direct injection of the samples into the LC-ESI-MS system. Signals of the compounds were monitored under the multiple reaction monitoring (MRM) mode of the LC-ESI-MS (ion trap) for quantification. The precursor to product ion transitions of m/z 240-184 and m/z 256-238 were used to measure BUP and HBUP, respectively. The method was validated in both plasma and microdialysate samples, and the obtained lower limit of quantification (LLOQ) was 1.5 ng mL(-1) for BUP and HBUP in both matrices. The intra- and inter-day assay variability was less than 15% for both analytes. This LC-ESI-MS method provided simple sampling, rapid clean-up and short analysis time (<9 min), applicable to the routine therapeutic monitoring and pharmacokinetic studies of BUP and HBUP., (Copyright © 2010 Elsevier B.V. All rights reserved.)

This investigation includes an evaluation of the percutaneous absorption of bupropion (BUP) and hydroxybupropion (BUPOH) in vitro and in vivo. In addition, a carbamate prodrug of BUPOH (But-BUPOH) was evaluated in vitro. In vitro diffusion studies were conducted in a flow-through diffusion cell system. The in vitro mean steady-state flux of BUP was significantly higher (p < 0.001) compared to BUPOH (320 +/- 16 nmol cm(-2) h(-1) vs. 27 +/- 4 nmol cm(-2) h(-1)). Additionally, a good correlation existed between in vitro and in vivo results. Mean steady-state plasma concentrations of 442 +/- 32 ng/mL and125 +/- 18 ng/mL were maintained over 48 h after topical application of BUP and BUPOH in hairless guinea pigs in vivo, respectively. Although BUP traversed human skin at rates sufficient to achieve required plasma levels, it is chemically unstable and hygroscopic, and unsuitable for transdermal formulation. On the other hand, BUPOH is stable but its transport across skin is much slower. Alternatively, the prodrug But-BUPOH was found to be stable, and also provided a 2.7-fold increase in the transdermal flux of BUPOH across human skin in vitro. Thus, But-BUPOH provides a viable option for the transdermal delivery of BUPOH.

A sensitive, stereoselective assay using solid phase extraction and LC-MS-MS was developed and validated for the analysis of (R)- and (S)-bupropion and its major metabolite (R,R)- and (S,S)-hydroxybupropion in human plasma and urine. Plasma or glucuronidase-hydrolyzed urine was acidified, then extracted using a Waters Oasis MCX solid phase 96-well plate. HPLC separation used an alpha(1)-acid glycoprotein column, a gradient mobile phase of methanol and aqueous ammonium formate, and analytes were detected by electrospray ionization and multiple reaction monitoring with an API 4000 Qtrap. The assay was linear in plasma from 0.5 to 200 ng/ml and 2.5 to 1000 ng/ml in each bupropion and hydroxybupropion enantiomer, respectively. The assay was linear in urine from 5 to 2000 ng/ml and 25 to 10,000 ng/ml in each bupropion and hydroxybupropion enantiomer, respectively. Intra- and inter-day accuracy was >98% and intra- and inter-day coefficients of variations were less than 10% for all analytes and concentrations. The assay was applied to a subject dosed with racemic bupropion. The predominant enantiomers in both urine and plasma were (R)-bupropion and (R,R)-hydroxybupropion. This is the first LC-MS/MS assay to analyze the enantiomers of both bupropion and hydroxybupropion in plasma and urine.

Bupropion is used clinically as an antidepressant and in smoking cessation. As it is metabolised to hydroxybupropion specifically by CYP2B6, bupropion has also been used as a probe to assess CYP2B6 activity. A specific and reproducible HPLC assay has been developed to simultaneously quantify bupropion and its major metabolites hydroxybupropion, threohydrobupropion and erythrohydrobupropion in human plasma. The analysis was performed on an Aqua C18 HPLC column, with a mobile phase consisting of 45:55 of methanol:0.05 M phosphate buffer (pH 5.5) and simultaneous UV detection at 214 nm (bupropion metabolites) and 254 nm (bupropion, internal standard timolol maleate). The assay showed a linear response for bupropion (2.5-250 ng/mL), threohydrobupropion (5-250 ng/mL), erythrohydrobupropion (10-250 ng/mL) and hydroxybupropion (10-1000 ng/mL). Extraction recovery was reproducible and greater than 55% for each analyte. The inter- and intra-day assay variability (measured as percent coefficient of variation; %CV) was less than 15% for all analytes. Limit of quantification was 2.5 ng/mL for bupropion, 5 ng/mL for threohydrobupropion and 10 ng/mL for hydroxybupropion and erythrohydrobupropion. This assay is more sensitive than currently published methods using HPLC with UV detection for the simultaneous quantitation of bupropion and metabolites and can be used for assessing CYP2B6 activity in vivo following a single dose of bupropion.

Background: (2S,3S)-2-(3-Chlorophenyl)-3,5,5,-trimethyl-2-morpholinol hydrochloride (radafaxine) is a new antidepressant that blocks dopamine transporters (DAT). A concern with drugs that block (DAT) is their potential reinforcing effects and abuse liability. Using positron emission tomography (PET) we have shown that for DAT-blocking drugs to produce reinforcing effects they must induce >50% DAT blockade and the blockade has to be fast (within 15 minutes). This study measures the potency and kinetics for DAT blockade by radafaxine in human brain., Methods: PET and [11C]cocaine were used to estimate DAT blockade at 1, 4, 8, and 24 hours after radafaxine (40 mg p.o.) in 8 controls. Plasma pharmacokinetics and behavioral and cardiovascular effects were measured in parallel., Results: DAT blockade by radafaxine was slow, and at 1 hour, it was 11%. Peak blockade occurred at about 4 hours and was 22%. Blockade was long lasting: at 8 hours 17%, and at 24 hours 15%. Peak plasma concentration occurred about 4 to 8 hours. No behavioral or cardiovascular effects were observed., Conclusions: The relatively low potency of radafaxine in blocking DAT and its slow blockade suggests that it is unlikely to have reinforcing effects. This is consistent with preclinical studies showing no self-administration. This is the first utilization of PET to predict abuse liability of a new antidepressant in humans based on DAT occupancy and pharmacokinetics.