Sustained release of 3D printed bupropion hydrochloride tablets bearing Braille imprints for the visually impaired.

3D printing has been introduced as a novel approach for the design of personalized dosage forms and support patient groups with special needs that require additional assistance for enhanced medication adherence. In this study liquid crystal display (LCD) is introduced for the development of sustained release bupropion.HCl printed tablets. The optimization of printing hydrogel inks was combined with the display of Braille patterns on the tablet surface for blind or visually impaired patients. Due to the high printing accuracy, the Braille patterns could be verified by blind patients and provide the required information. Further characterization revealed the presence of BUP in amorphous state within the photopolymerized resins. The selection of poly(ethylene glycol) (PEG)-diacrylate (PEGDA) of different molecular weights and the presence of surfactants or solubilizers disrupted the resin photopolymerization, thus controlling the BUP dissolution rates. A small batch scale-up study demonstrated the capacity of LCD to print rapidly a notable number of tablets within 24 min.

Transport and inhibition mechanisms of the human noradrenaline transporter.

The noradrenaline transporter (also known as norepinephrine transporter) (NET) has a critical role in terminating noradrenergic transmission by utilizing sodium and chloride gradients to drive the reuptake of noradrenaline (also known as norepinephrine) into presynaptic neurons1-3. It is a pharmacological target for various antidepressants and analgesic drugs4,5. Despite decades of research, its structure and the molecular mechanisms underpinning noradrenaline transport, coupling to ion gradients and non-competitive inhibition remain unknown. Here we present high-resolution complex structures of NET in two fundamental conformations: in the apo state, and bound to the substrate noradrenaline, an analogue of the χ-conotoxin MrlA (χ-MrlAEM), bupropion or ziprasidone. The noradrenaline-bound structure clearly demonstrates the binding modes of noradrenaline. The coordination of Na+ and Cl- undergoes notable alterations during conformational changes. Analysis of the structure of NET bound to χ-MrlAEM provides insight into how conotoxin binds allosterically and inhibits NET. Additionally, bupropion and ziprasidone stabilize NET in its inward-facing state, but they have distinct binding pockets. These structures define the mechanisms governing neurotransmitter transport and non-competitive inhibition in NET, providing a blueprint for future drug design.

Molecular basis of human noradrenaline transporter reuptake and inhibition.

Noradrenaline, also known as norepinephrine, has a wide range of activities and effects on most brain cell types1. Its reuptake from the synaptic cleft heavily relies on the noradrenaline transporter (NET) located in the presynaptic membrane2. Here we report the cryo-electron microscopy (cryo-EM) structures of the human NET in both its apo state and when bound to substrates or antidepressant drugs, with resolutions ranging from 2.5 Å to 3.5 Å. The two substrates, noradrenaline and dopamine, display a similar binding mode within the central substrate binding site (S1) and within a newly identified extracellular allosteric site (S2). Four distinct antidepressants, namely, atomoxetine, desipramine, bupropion and escitalopram, occupy the S1 site to obstruct substrate transport in distinct conformations. Moreover, a potassium ion was observed within sodium-binding site 1 in the structure of the NET bound to desipramine under the KCl condition. Complemented by structural-guided biochemical analyses, our studies reveal the mechanism of substrate recognition, the alternating access of NET, and elucidate the mode of action of the four antidepressants.

A bupropion modulatory site in the Gloeobacter violaceus ligand-gated ion channel.

Bupropion is an atypical antidepressant and smoking cessation drug that causes adverse effects such as insomnia, irritability, and anxiety. Bupropion inhibits dopamine and norepinephrine reuptake transporters and eukaryotic cation-conducting pentameric ligand-gated ion channels, such as nicotinic acetylcholine and serotonin type 3A receptors, at clinically relevant concentrations. Here, we demonstrate that bupropion also inhibits a prokaryotic homolog of pentameric ligand-gated ion channels, the Gloeobacter violaceus ligand-gated ion channel (GLIC). Using the GLIC as a model, we used molecular docking to predict binding sites for bupropion. Bupropion was found to bind to several sites within the transmembrane domain, with the predominant site being localized to the interface between transmembrane segments M1 and M3 of two adjacent subunits. Residues W213, T214, and W217 in the first transmembrane segment, M1, and F267 and I271 in the third transmembrane segment, M3, most frequently reside within a 4 Å distance from bupropion. We then used single amino acid substitutions at these positions and two-electrode voltage-clamp recordings to determine their impact on bupropion inhibitory effects. The substitution T214F alters bupropion potency by shifting the half-maximal inhibitory concentration to a 13-fold higher value compared to wild-type GLIC. Residue T214 is found within a previously identified binding pocket for neurosteroids and lipids in the GLIC. This intersubunit binding pocket is structurally conserved and almost identical to a binding pocket described for neurosteroids in γ-aminobutyric acid type A receptors. Our data thus suggest that the T214 that lines a previously identified lipophilic binding pocket in GLIC and γ-aminobutyric acid type A receptors is also a modulatory site for bupropion interaction with the GLIC.

Stereoselective Bupropion Hydroxylation by Cytochrome P450 CYP2B6 and Cytochrome P450 Oxidoreductase Genetic Variants.

Bioactivation of the antidepressant and smoking cessation drug bupropion is catalyzed predominantly by CYP2B6. The metabolite hydroxybupropion derived from t-butylhydroxylation is considered to contribute to the antidepressant and smoking-cessation effects of the parent drug. Bupropion hydroxylation is the canonical in vitro and in vivo probe for CYP2B6 activity. P450 also requires obligate partnership with P450 oxidoreductase (POR). Human CYP2B6 and POR genes are highly polymorphic. Some CYP2B6 variants affect bupropion disposition. This investigation evaluated the influence of several human CYP2B6 and POR genetic variants on stereoselective bupropion metabolism, using an insect cell coexpression system containing CYP2B6, POR, and cytochrome b 5 Based on intrinsic clearances (Clints), relative activities for S,S-hydroxybupropion formation were in the order CYP2B6.4 > CYP2B6.1 > CYP2B6.17 > CYP2B6.5 > CYP2B6.6 ≈ CYP2B6.26 ≈ CYP2B6.19 > CYP2B6.7 > CYP2B6.9 > > CYP2B6.16 and CYP2B6.18; relative activities for R,R-hydroxybupropion formation were in the order CYP2B6.17 > CYP2B6.4 > CYP2B6.1 > CYP2B6.5 ≈ CYP2B6.19 ≈ CYP2B6.26 > CYP2B6.6 > CYP2B6.7 ≈ CYP2B6.9 > > CYP2B6.16 and CYP2B6.18. Bupropion hydroxylation was not influenced by POR variants. CYP2B6-catalyzed bupropion hydroxylation is stereoselective. Though Vmax and Km varied widely among CYP2B6 variants, stereoselectivity was preserved, reflected by similar Clint(S,S-hydroxybupropion)/Clint(R,R-hydroxybupropion) ratios (1.8-2.9), except CYP2B6.17, which was less enantioselective. Established concordance between human bupropion hydroxylation in vitro and in vivo, together with these new results, suggests additional CYP2B6 variants may influence human bupropion disposition. SIGNIFICANCE STATEMENT: Bupropion pharmacokinetics, metabolism, and clinical effects are affected by the CYP2B6*6 polymorphism. Other expressed CYP2B6 polymorphisms had diminished (*5, *6, *7, *9, *19, *26) or defective (*16, *18) in vitro bupropion hydroxylation. P450 oxidoreductase genetic variants had no effect on metabolism, suggesting no clinical consequence of this polymorphism. These CYP2B6 polymorphisms may portend diminished in vivo bupropion hydroxylation and predict additional clinically important variant alleles.

Pharmacokinetic and pharmacodynamic of bupropion: integrative overview of relevant clinical and forensic aspects.

Bupropion is an atypical antidepressant of the aminoketone group, structurally related to cathinone, associated with a wide interindividual variability. An extensive pharmacokinetic and pharmacodynamic review of bupropion was performed, also focusing on chemical, pharmacological, toxicological, clinical and forensic aspects of this drug without a limiting period. Bupropion is a chiral, basic, highly lipophilic drug, clinically used as racemate that undergoes extensive stereoselective metabolism. Its major active metabolites, hydroxybupropion, threohydrobupropion, and erythrohydrobupropion reach higher plasma concentrations than bupropion. Bupropion exerts its effects mainly by inhibiting dopamine and norepinephrine reuptake and by blocking several nicotinic receptors. Recent reports highlight recreational use of bupropion via intranasal insufflation and intravenous use. Seizures, insomnia, agitation, headache, dry mouth, and nausea are some of the reported adverse effects. Neurologic effects are major signs of intoxication that should be carefully managed. Finally, the characterization of the polymorphic enzymes involved in the metabolism of bupropion is essential to understand factors that may influence the interindividual and intraindividual variability in bupropion metabolite exposure, including the evaluation of potential drug-drug interactions and pharmacogenetic implications.

Systematic review of preclinical, clinical, and post-marketing evidence of bupropion misuse potential.

Background: Bupropion is a substituted cathinone compound widely used as a first line or add-on treatment for depression, smoking cessation, and more recently in combination with naltrexone for weight loss. As abuse of synthetic cathinone compounds has received more attention in recent years, concern about the misuse potential of bupropion has grown as well. Objectives: We review bupropion pharmacology and assessments of misuse potential including preclinical evidence, human studies, and post-marketing surveillance of bupropion misuse. Methods: This review reports the results of a systematic review of publications evaluating the potential for bupropion to be misused. Publications were identified using PubMed and Medline through Ovid® as well as iterative bibliographic searches. A summary of data from informal sources of information including substance-user experience from online forum entries is included. Results: Preclinical evidence demonstrates some potential for misuse based on psychomotor, discrimination, self-administration, and conditioned place preference tasks. However, this potential is less than that of commonly misused stimulants. Studies in human populations similarly indicate that bupropion shares interoceptive effects with other stimulants, but lacks some key reinforcing effects of other stimulants. In the real-world setting, misuse of bupropion occurs, but is uncommon. Adverse effects of bupropion misuse are frequently cited as significant barriers to obtaining any desired interoceptive effect. Conclusions: While bupropion demonstrates some potential for misuse, pharmacological differences from other structurally-related stimulants limit bupropion's reinforcing effects. Without additional data indicating susceptibility of specific populations to bupropion misuse, there is no empirical data suggesting a need to modify bupropion prescribing patterns.

Common Polymorphisms of CYP2B6 Influence Stereoselective Bupropion Disposition.

Bupropion hydroxylation is a bioactivation and metabolic pathway, and the standard clinical CYP2B6 probe. This investigation determined the influence of CYP2B6 allelic variants on clinical concentrations and metabolism of bupropion enantiomers. Secondary objectives evaluated the influence of CYP2C19 and P450 oxidoreductase variants. Healthy volunteers in specific cohorts (CYP2B6*1/*1, CYP2B6*1/*6, CYP2B6*6/*6, and also CYP2B6*4 carriers) received single-dose oral bupropion. Plasma and urine bupropion and hydroxybupropion was quantified. Subjects were also genotyped for CYP2C19 and P450 oxidoreductase variants. Hydroxylation of both bupropion enantiomers, assessed by plasma hydroxybupropion/bupropion AUC ratios and urine hydroxybupropion formation clearances, was lower in CYP2B6*6/*6 but not CYP2B6*1/*6 compared with CYP2B6*1/*1 genotypes, and numerically greater in CYP2B6*4 carriers. CYP2C19 and P450 oxidoreductase variants did not influence bupropion enantiomers hydroxylation or plasma concentrations. The results show that clinical hydroxylation of both bupropion enantiomers was equivalently influenced by CYP2B6 allelic variation. CYP2B6 polymorphisms affect S-bupropion bioactivation, which may affect therapeutic outcomes.

Comparison of In Vitro Stereoselective Metabolism of Bupropion in Human, Monkey, Rat, and Mouse Liver Microsomes.

BACKGROUND AND OBJECTIVES: Bupropion is an atypical antidepressant and smoking cessation aid associated with wide intersubject variability. This study compared the formation kinetics of three phase I metabolites (hydroxybupropion, threohydrobupropion, and erythrohydrobupropion) in human, marmoset, rat, and mouse liver microsomes. The objective was to establish suitability and limitations  for subsequent use of nonclinical species to model bupropion central nervous system (CNS) disposition in humans. METHODS: Hepatic microsomal incubations were conducted separately for the R- and S-bupropion enantiomers, and the formation of enantiomer-specific metabolites was determined using LC-MS/MS. Intrinsic formation clearance (CLint) of metabolites across the four species was determined from the formation rate versus substrate concentration relationship. RESULTS: The total clearance of S-bupropion was higher than that of R-bupropion in monkey and human liver microsomes. The contribution of hydroxybupropion to the total racemic bupropion clearance was 38%, 62%, 17%, and 96% in human, monkey, rat, and mouse, respectively.  In the same species order, threohydrobupropion contributed 53%, 23%, 17%, and 3%, and erythrohydrobupropion contributed 9%, 14%, 66%, and 1.3%, respectively, to racemic bupropion clearance. CONCLUSION: The results demonstrate that phase I metabolism in monkeys best approximates that observed in humans, and support the preferred use of this species to investigate possible pharmacokinetic factors that influence the CNS disposition of bupropion and contribute to its high intersubject variability.

Interpretation of the binding interaction between bupropion hydrochloride with human serum albumin: A collective spectroscopic and computational approach.

Bupropion hydrochloride (BPH) an antidepressant and widely used to treat addiction of nicotine. The actual protein existing in blood plasma for the vehicle of exogenous and endogenous substances is human serum albumin i.e. HSA. The interaction of BPH with HSA was examined by molecular docking, multiple spectroscopy's such as fluorescence (emission, synchronous and three-dimensional), UV-vis (ultraviolet-visible), FT-IR (Fourier transform infrared) and CD (circular dichroism) at physiological pH 7.40 at 286, 296 and 306 K. BPH was particularly bind to HSA through forces called hydrogen bonds and vander Waals at site I (IIA) which was confirmed from negative values of thermodynamics calculated by van't Hoff equation and docking studies in addition to site marker analysis. This interaction was spontaneous and exothermic process. Secondary structure including conformation of HSA changes after interaction with BPH was revealed from CD and FT-IR (Fourier self-deconvolution to curve fitting), UV-vis, 3D and synchronous florescence techniques. Forster's theory (non-radiation energy transfer) was applied to calculate the distance from tryptophan of HSA to BPH. This interaction involves static quenching (Stern-Volmer and Modified Stern-Volmer equations) with larger binding constant values were in the range 105 confirming that strong interaction was exists between BPH and HSA. The interference of bio-active Mg2+, Cu2+, Zn2+, Ca2+ and Fe2+ metal ions on this interaction was also analysed.

Inhibitory effect of α-terpinyl acetate on cytochrome P450 2B6 enzymatic activity.

Human cytochrome P450 2B6 is an important hepatic enzyme for the metabolism of xenobiotics and clinical drugs. Recently, more attention has been paid to P450 2B6 because of the increasing number of drugs it metabolizes. It has been known to interact with terpenes, the major constituents of the essential oils used for various medicinal purposes. In this study, the effect of monoterpenes on P450 2B6 catalytic activity was investigated. Recombinant P450 2B6 was expressed in Escherichia coli and purified using Ni-affinity chromatography. The purified P450 2B6 enzyme displayed bupropion hydroxylation activity in gas-mass spectrometry (GC-MS) analysis with a kcat of 0.5 min-1 and a Km of 47 µM. Many terpenes displayed the type I binding spectra to purified P450 2B6 enzyme and α-terpinyl acetate showed strong binding affinity with a Kd value of 5.4 µM. In GC-MS analysis, P450 2B6 converted α-terpinyl acetate to a putative oxidative product. The bupropion hydroxylation activity of P450 2B6 was inhibited by α-terpinyl acetate and its IC50 value was 10.4 µM α-Terpinyl acetate was determined to be a competitive inhibitor of P450 2B6 with a Ki value of 7.6 µM. The molecular docking model of the binding site of the P450 2B6 complex with α-terpinyl acetate was constructed. It showed the tight binding of α-terpinyl acetate in the active site of P450 2B6, which suggests that it could be a competitive substrate for P450 2B6.

Chirality and neuropsychiatric drugs: an update on stereoselective disposition and clinical pharmacokinetics of bupropion.

1. Bupropion, an antidepressant drug has been approved as a racemate containing equal amounts of R- and S-enantiomers. Recently, the chirality of bupropion has received significant attention in the delineation of stereoselective pharmacokinetic (PK) and disposition data. Although the non-stereoselective metabolism of bupropion was well established, the emerging data suggest that bupropion exhibits complex stereoselective metabolism, leading to the formation of various stereoisomeric metabolites. Along with the chiral PKs of bupropion, hydroxybupropion, threohydrobupropion and erythrohydrobupropion, the metabolism data also provided insights into the roles of both CYP2B6 and CYP2C19 enzymes in the stereoselective disposition. Furthermore, the metabolism studies also suggested specific involvement of CYP2B6 pathway in the stereoselective hydroxylation of bupropion to R,R-hydroxybupropion, which was considered as a better marker for CYP2B6 activity. 2. Other significant learnings were: (1) understanding the in vivo CYP2D6 inhibitory potential of bupropion with respect to the chirality of parent drug and the metabolites; (2) the potential involvement of bupropion and metabolites towards significant down regulation of CYP2D6 mRNA; (3) significant in vivo CYP2D6 inhibitory activity (86%) exhibited by R,R-hydroxybupropion and threohydrobupropion. 3. The newly published data on chiral PKs and disposition of bupropion and its metabolites can be used to gauge the drug-drug interaction potential when bupropion is combined in clinical therapy. Moreover, such data would be useful to understand the consequences (if any), due to the combination of bupropion with other drugs both from a safety and efficacy perspective because of the prevalence of polypharmacy situations in many therapeutic areas including CNS indications.

Application of wavelet and Fuorier transforms as powerful alternatives for derivative spectrophotometry in analysis of binary mixtures: A comparative study.

Two signal processing methods, namely, Continuous Wavelet Transform (CWT) and the second was Discrete Fourier Transform (DFT) were introduced as alternatives to the classical Derivative Spectrophotometry (DS) in analysis of binary mixtures. To show the advantages of these methods, a comparative study was performed on a binary mixture of Naltrexone (NTX) and Bupropion (BUP). The methods were compared by analyzing laboratory prepared mixtures of the two drugs. By comparing performance of the three methods, it was proved that CWT and DFT methods are more efficient and advantageous in analysis of mixtures with overlapped spectra than DS. The three signal processing methods were adopted for the quantification of NTX and BUP in pure and tablet forms. The adopted methods were validated according to the ICH guideline where accuracy, precision and specificity were found to be within appropriate limits.

Pharmacokinetics and Pharmacogenomics of Bupropion in Three Different Formulations with Different Release Kinetics in Healthy Human Volunteers.

The purpose of this pharmacokinetics (PK) study was to investigate whether different release kinetics from bupropion hydrochloride (HCl) immediate release (IR), sustained release (SR), and extended release (ER) formulations alter its metabolism and to test the hypothesis that the unsuccessful bioequivalence (BE) study of the higher strength (300 mg) of bupropion HCl ER tablets based on the successful BE study of the lower strength (150 mg) was due to metabolic saturation in the gastrointestinal (GI) lumen. A randomized six-way crossover study was conducted in healthy volunteers. During each period, subjects took a single dose of IR (75/100 mg), SR (100/150 mg), or ER (150/300 mg) formulations of bupropion HCl; plasma samples for PK analysis were collected from 0-96 h for all formulations. In addition, each subject's whole blood was collected for the genotyping of various single-nucleotide polymorphisms (SNPs) of bupropion's major metabolic enzymes. The data indicates that the relative bioavailability of the ER formulations was 72.3-78.8% compared with IR 75 mg. No differences were observed for ratio of the area under the curve (AUC) of metabolite to AUC of parent for the three major metabolites. The pharmacogenomics analysis suggested no statistically significant correlation between polymorphisms and PK parameters of the various formulations. Altogether, these data suggested that the different release kinetics of the formulations did not change metabolites-to-parent ratio. Therefore, the differing BE result between the 150 and 300 mg bupropion HCl ER tablets was unlikely due to the metabolic saturation in the GI lumen caused by different release patterns.

Deconstructed Analogues of Bupropion Reveal Structural Requirements for Transporter Inhibition versus Substrate-Induced Neurotransmitter Release.

Bupropion (1), an α-aminophenone uptake inhibitor at plasma membrane transporters for dopamine (DAT) and norepinephrine (NET), is a widely prescribed antidepressant and smoking cessation aid. Cathinone (2), a structurally simpler α-aminophenone, is a substrate-type releasing agent at the same transporters and a recognized drug of abuse. Our goal was to identify the structural features of α-aminophenones that govern the mechanistic transition from uptake inhibition to substrate-induced release. Deconstructed analogues of 1 were synthesized and compared for their ability to interact with DAT, NET, and the serotonin transporter (SERT) using in vitro assay methods. Bulky amine substituents resulted in compounds that function as DAT uptake inhibitors but not release agents, whereas smaller amine substituents result in relatively nonselective releasing agents at DAT and NET. Our findings add to empirical evidence supporting distinct molecular determinants for α-aminophenone- (i.e., cathinone-) related agents acting as transporter inhibitors versus those acting as releasers.

Design of experiment assisted concurrent enantioseparation of bupropion and hydroxybupropion by high-performance thin-layer chromatography.

A simple and efficient high-performance thin-layer chromatographic method was developed for chiral separation of rac-bupropion (BUP) and its active metabolite rac-hydroxybupropion (HBUP). Design of experiment (DoE)-based optimization was adopted instead of a conventional trial-and-error approach. The Box-Behnken design surface response model was used and the operating variables were optimized based on 17 trials design. The optimized method involved impregnation of chiral reagent, L(+)-tartaric acid, in the stationary phase with simultaneous addition in the mobile phase, which consisted of acetonitrile : methanol : dichloromethane : 0.50% L-tartaric acid (6.75:1.0:1.0:0.25, v/v/v/v). Under the optimized conditions, the resolution factor between the enantiomers of BUP and HBUP was 6.30 and 9.26, respectively. The limit of detection and limit of quantitation for (R)-BUP, (S)-BUP, (R,R)-HBUP, and (S,S)-HBUP were 9.23 and 30.78 ng spot-1 , 10.32 and 34.40 ng spot-1 , 12.19 and 40.65 ng spot-1 , and 14.26 and 47.53 ng spot-1 , respectively. The interaction of L-tartaric acid with analytes and their retention behavior was thermodynamically investigated using van't Hoff's plots. The developed method was validated as per the International Conference on Harmonization guidelines. Finally, the method was successfully applied to resolve and quantify the enantiomeric content from marketed tablets as well as spiked plasma samples.

Evaluation of a Potential Metabolism-Mediated Drug-Drug Interaction Between Atomoxetine and Bupropion in Healthy Volunteers.

PURPOSE: To evaluate the impact of bupropion on the pharmacokinetic profile of atomoxetine and its main active metabolite (glucuronidated form), 4-hydroxyatomoxetine-O-glucuronide, in healthy volunteers. METHODS: An open-label, non-randomized, two-period, sequential clinical trial was conducted as follows: during Period I (Reference), each volunteer received a single oral dose of 25 mg atomoxetine, whilst during Period II (Test), a combination of 25 mg atomoxetine and 300 mg bupropion was administered to all volunteers, after a pretreatment regimen with bupropion for 7 days. Next, after determining atomoxetine and 4-hydroxyatomoxetine-O-glucuronide plasma concentrations, their pharmacokinetic parameters were calculated using a noncompartmental method and subsequently compared to determine any statistically significant differences between the two periods. RESULTS: Bupropion intake influenced all the pharmacokinetic parameters of both atomoxetine and its metabolite. For atomoxetine, Cmax increased from 226±96.1 to 386±137 ng/mL and more importantly, AUC0-∞ was significantly increasedfrom 1580±1040 to 8060±4160 ng*h/mL, while the mean t1/2 was prolonged after bupropion pretreatment. For 4-hydroxyatomoxetine-O-glucuronide, Cmax and AUC0-∞  were decreased from 707±269 to 212±145 ng/mL and from 5750±1240 to 3860±1220 ng*h/mL, respectively. CONCLUSIONS: These results demonstrated that the effect of bupropion on CYP2D6 activity was responsible for an increased systemic exposure to atomoxetine (5.1-fold) and also for a decreased exposure to its main metabolite (1.5-fold). Additional studies are required in order to evaluate the clinical relevance of this pharmacokinetic drug interaction.This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Chiral Plasma Pharmacokinetics and Urinary Excretion of Bupropion and Metabolites in Healthy Volunteers.

Bupropion, widely used as an antidepressant and smoking cessation aid, undergoes complex metabolism to yield numerous metabolites with unique disposition, effect, and drug-drug interactions (DDIs) in humans. The stereoselective plasma and urinary pharmacokinetics of bupropion and its metabolites were evaluated to understand their potential contributions to bupropion effects. Healthy human volunteers (n = 15) were administered a single oral dose of racemic bupropion (100 mg), which was followed by collection of plasma and urine samples and determination of bupropion and metabolite concentrations using novel liquid chromatography-tandem mass spectrometry assays. Time-dependent, elimination rate-limited, stereoselective pharmacokinetics were observed for all bupropion metabolites. Area under the plasma concentration-time curve from zero to infinity ratios were on average approximately 65, 6, 6, and 4 and Cmax ratios were approximately 35, 6, 3, and 0.5 for (2R,3R)-/(2S,3S)-hydroxybupropion, R-/S-bupropion, (1S,2R)-/(1R,2S)-erythrohydrobupropion, and (1R,2R)-/(1S,2S)-threohydrobupropion, respectively. The R-/S-bupropion and (1R,2R)-/(1S,2S)-threohydrobupropion ratios are likely indicative of higher presystemic metabolism of S- versus R-bupropion by carbonyl reductases. Interestingly, the apparent renal clearance of (2S,3S)-hydroxybupropion was almost 10-fold higher than that of (2R,3R)-hydroxybupropion. The prediction of steady-state pharmacokinetics demonstrated differential stereospecific accumulation [partial area under the plasma concentration-time curve after the final simulated bupropion dose (300-312 hours) from 185 to 37,447 nM⋅h] and elimination [terminal half-life of approximately 7-46 hours] of bupropion metabolites, which may explain observed stereoselective differences in bupropion effect and DDI risk with CYP2D6 at steady state. Further elucidation of bupropion and metabolite disposition suggests that bupropion is not a reliable in vivo marker of CYP2B6 activity. In summary, to our knowledge, this is the first comprehensive report to provide novel insight into mechanisms underlying bupropion disposition by detailing the stereoselective pharmacokinetics of individual bupropion metabolites, which will enhance clinical understanding of bupropion's effects and DDIs with CYP2D6.

HPLC enantioseparation of racemic bupropion, baclofen and etodolac: modification of conventional ligand exchange approach by pre-column formation of chiral ligand exchange complexes.

Separation of racemic mixture of (RS)-bupropion, (RS)-baclofen and (RS)-etodolac, commonly marketed racemic drugs, has been achieved by modifying the conventional ligand exchange approach. The Cu(II) complexes were first prepared with a few l-amino acids, namely, l-proline, l-histidine, l-phenylalanine and l-tryptophan, and to these was introduced a mixture of the enantiomer pair of (RS)-bupropion, or (RS)-baclofen or (RS)-etodolac. As a result, formation of a pair of diastereomeric complexes occurred by 'chiral ligand exchange' via the competition between the chelating l-amino acid and each of the two enantiomers from a given pair. The diastereomeric mixture formed in the pre-column process was loaded onto HPLC column. Thus, both the phases during chromatographic separation process were achiral (i.e. neither the stationary phase had any chiral structural feature of its own nor did the mobile phase have any chiral additive). Separation of diastereomers was successful using a C18 column and a binary mixture of MeCN and TEAP buffer of pH 4.0 (60:40, v/v) as mobile phase at a flow rate of 1 mL/min and UV detection at 230 nm for (RS)-Bup, 220 nm for (RS)-Bac and 223 nm for (RS)-Etd. Baseline separation of the two enantiomers was obtained with a resolution of 6.63 in <15 min. Copyright © 2016 John Wiley & Sons, Ltd.

Stereoselective method to quantify bupropion and its three major metabolites, hydroxybupropion, erythro-dihydrobupropion, and threo-dihydrobupropion using HPLC-MS/MS.

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.