Fluorescence detection

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Determination of Reboxetine in Rat Brain Microdialysates and Plasma Samples Using Liquid Chromatography Coupled To Fluorescence Detection

Journal Title, Volume, Page: 
Journal of Chromatography B Volume 898, 1 June 2012, Pages 53–61
Year of Publication: 
2012
Authors: 
Naser Shraim
Vrije Universiteit Brussel, Department of Pharmaceutical Chemistry and Drug Analysis, Center for Neurosciences, Laarbeeklaan 103, B-1090 Brussels, Belgium
Current Affiliation: 
Department of Pharmacy, Faculty of Medicine and Health Sciences, An-Najah National University, Nablus, Palestine
Ralph Clinckers
Vrije Universiteit Brussel, Department of Pharmaceutical Chemistry and Drug Analysis, Center for Neurosciences, Laarbeeklaan 103, B-1090 Brussels, Belgium
Sofie Sarre
Vrije Universiteit Brussel, Department of Pharmaceutical Chemistry and Drug Analysis, Center for Neurosciences, Laarbeeklaan 103, B-1090 Brussels, Belgium
Yvette Michotte
Vrije Universiteit Brussel, Department of Pharmaceutical Chemistry and Drug Analysis, Center for Neurosciences, Laarbeeklaan 103, B-1090 Brussels, Belgium
Ann Van Eeckhaut
Vrije Universiteit Brussel, Department of Pharmaceutical Chemistry and Drug Analysis, Center for Neurosciences, Laarbeeklaan 103, B-1090 Brussels, Belgium
Preferred Abstract (Original): 
A liquid chromatographic method with fluorescence detection was developed and validated for the quantification of the antidepressant reboxetine (RBX), a selective noradrenalin reuptake inhibitor, in rat brain microdialysates. After modification of the method in terms of sample preparation and sensitivity, it was also validated for the quantification of RBX in rat plasma samples. To enable fluorescence detection, a pre-column derivatization step with 9-fluorenylmethyl chloroformate was included. Separations were performed on a reversed phase C₁₈ column using gradient elution. The retention time for RBX was found to be 8.8 min. The assay of RBX in brain microdialysis samples showed a linear relationship in the calibration curve from 2 to 200 ng/mL, with a correlation coefficient ≥0.999. The limit of detection (LOD) and the lower limit of quantification (LLOQ) were 0.6 and 2.0 ng/mL respectively. The intra-day and the inter-day precision (RSD %) ranged between 1.5% and 11.7% with an average recovery of 101.2±8.2% (mean±SD, n=40). For the analysis of plasma samples, the calibration curve was linear between 20 and 700 ng/mL with a correlation coefficient ≥0.999. LOD and LLOQ were 6 and 20 ng/mL respectively. The intra-day and the inter-day precision (RSD %) ranged between 1.7% and 11.5% with an average recovery of 98.5±7.3% (mean±SD, n=40). We demonstrated the applicability of the method to determine the concentration-time profiles of RBX in brain and plasma following systemic administration.
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