A Highly Sensitive LC-MS/MS Method for the Simultaneous Quantification of Bupropion, Hydroxy Bupropion, Erythro Hydro Bupropion and Threo Hydro Bupropion in Human plasma
08 Jul 2018
Bupropion is a relatively weak inhibitor of the neuronal uptake of norepinephrine and dopamine, and does not inhibit monoamine oxidase or the re-uptake of serotonin. It is indicated for the treatment of major depressive disorder and prevention of seasonal major depressive episodes in patients with a diagnosis of seasonal affective disorder.
A selective and sensitive method at the LLOQ of about 1.0 ng/mL for Bupropion, 2.0 ng/mL for Hydroxy Bupropion, 0.5 ng/mL for Erythro Hydro Bupropion and 1.5 ng/mL for Threo Hydro Bupropion are required. This was accomplished in the new method developed at our end.
Briefly, the method for the quantification of Bupropion, Hydroxy Bupropion, Erythro Hydro Bupropion and Threo Hydro Bupropion involves solid phase extraction using Bupropion-d9, Hydroxy Bupropion-d6, Erythro Hydro Bupropion-d9 and Threo Hydro Bupropion-d9 as internal standards (ISTD). Chromatographic separation of the drug and all metabolites were achieved on Poroshell 120 SB C-18 2.7µm 100*4.6 mm column with a mobile phase consisting of 5 mM ammonium formate buffer and methanol, with isocratic time program, and a total run-time of 9 minutes with adequate separation between the metabolites. Detection was carried out in a positive electro-spray ionization mode using MRM transitions of 240.04→184.13, 256.06→238.18, 242.05→168.13, 242.10→168.13, 249.09→185.14, 262.08→244.23, 251.11→169.13, and 251.16→169.14 for Bupropion, Hydroxy Bupropion, Erythro Hydro Bupropion, Threo Hydro Bupropion, Bupropion-d9(ISTD), Hydroxy Bupropion-d6 (ISTD), Erythro Hydro Bupropion-d9 (ISTD) and Threo Hydro Bupropion-d9 (ISTD) respectively.
This new method was validated in terms of linearity, stabilities, carry-over effect, reinjection reproducibility, precision, accuracy and selectivity. The linearity ranges from 1 to 300 ng/mL for Bupropion, 2 to 800 ng/mL for Hydroxy Bupropion, 0.5 to 100 ng/mL for Erythro Hydro Bupropion and 1.5 to 600 ng/mL for Threo Hydro Bupropion; and is found to be stable in plasma under the assay conditions. Moreover, the method exhibited good precision and accuracy with precision better than 7.3% and accuracy greater than 98.5% for Bupropion, 12.9% and 96.9% for Hydroxy Bupropion, 12.2% and 93.3% for Erythro Hydro Bupropion and 7.8% and 97.0% for Threo Hydro Bupropion. Also, there was no significant matrix effect observed in normal human lots and no inter-conversion issues.
This validated method is suitable for the analysis of samples from a single oral dose of 150 mg administration to humans under fasting and fed conditions, and will be applied to the analysis of study samples.
To conclude, a selective, simple, precise, accurate and highly sensitive LC-MS/MS method is now available for simultaneous quantification of Bupropion, Hydroxy Bupropion, Erythro Hydro Bupropion and Threo Hydro Bupropion in human plasma.