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Mohamed S.,IRCCS ISNB Institute of Neurological science of Bologna | Riva R.,IRCCS ISNB Institute of Neurological science of Bologna | Riva R.,University of Bologna | Contin M.,IRCCS ISNB Institute of Neurological science of Bologna | Contin M.,University of Bologna
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2016

We present a simple, fast and validated method for the determination of nimodipine in plasma and cerebrospinal fluid (CSF) of patients with subarachnoid haemorrhage using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Plasma or CSF 250 μL aliquots were pretreated with acetonitrile spiked with lacosamide as internal standard. The chromatographic separation was performed on a Fusion (3 μm) 50 × 2.0 mm I.D. column with gradient elution of 0.1% (v/v) formic acid in water and 0.1% (v/v) formic acid in acetonitrile at a flow rate of 0.35 mL/min. The MS/MS ion transitions were 419.1 → 343 for nimodipine and 251.1 → 91 for the internal standard. The linearity was determined from 2.0 to 40.0 ng/mL in plasma and 40.0-800.0 pg/mL in CSF. The lower limit of quantitation (LLOQ) of nimodipine was 0.4 ng/mL in plasma and 40 pg/mL in CSF. The mean recovery for nimodipine was ≥75% in plasma and ≥90% in CSF at all three considered concentrations. Intra- and interassay precision and accuracy were ≤15% at all quality control concentrations in plasma and CSF. The method was applied to measure plasma and CSF concentrations of nimodipine in a series of patients with subarachnoid haemorrhage treated with intravenous nimodipine. The present procedure, omitting time-consuming liquid-liquid extraction and drying steps, is faster, simpler and cheaper than published LC-MS/MS analytical methods for nimodipine in plasma and the first validated one for nimodipine in CSF. © 2016 Elsevier B.V.


Mohamed S.,IRCCS ISNB Institute of Neurological science of Bologna | Riva R.,IRCCS ISNB Institute of Neurological science of Bologna | Riva R.,University of Bologna | Contin M.,IRCCS ISNB Institute of Neurological science of Bologna | Contin M.,University of Bologna
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2016

A simple and validated ultra high pressure liquid chromatography-tandem mass spectrometry method was developed for the simultaneous determination of the dopaminergic agents pramipexole and ropinirole in plasma of patients with Parkinson's disease. Following liquid-liquid extraction with tert-butyl methyl ether from 250 μL plasma, the separation of the analytes was achieved on a Gemini NX3 column using 10 mM pH 6.0 ammonium formate and 10 mM ammonium formate in methanol as binary gradient mobile phase at a flow rate of 0.3 mL/min. The MS/MS ion transitions were 212.10 → 153.03 for pramipexole, 261.2 → 114.2 for ropinirole and 256.1 → 211 for the internal standard (lamotrigine). The lower limit of quantitation (LLOQ) for both analytes was 80 pg/mL and the linearity was determined from 80 to 4000 pg/mL for pramipexole and from 200 to 10000 pg/mL for ropinirole. Mean recoveries were 94% for PRA and 73% for ROP. Both intra- and inter-assay precision and accuracy were ≤20% at LLOQ concentration and ≤15% at other concentrations. The proposed validated method was successfully applied to measure plasma concentrations of pramipexole and ropinirole in a series of patients with Parkinson's disease on chronic treatment. By grouping the two dopaminergic agents in the same assay, the method allows a large series of patient samples to be processed in a single analytical session. © 2016 Elsevier B.V..


Mohamed S.,IRCCS ISNB Institute of Neurological science of Bologna | Caporali L.,IRCCS ISNB Institute of Neurological science of Bologna | De Giorgio R.,University of Bologna | Carelli V.,IRCCS ISNB Institute of Neurological science of Bologna | And 3 more authors.
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2014

We present a simple, fast and validated method for the determination of the two nucleosides thymidine (dThd) and deoxyuridine (dUrd) in plasma of patients with symptoms suggestive of mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), using high performance liquid chromatography coupled with ultraviolet spectrophotometric detection (HPLC-UV). Plasma sample (100. μL) pretreatment was based on simple deproteinization by 1.2. M perchloric acid, using theophylline as internal standard (I.S.). HPLC-UV analysis was carried out on a Synergi 4. μm Hydro-RP, 150 × 4 mm I.D. column, at room temperature. The mobile phase was a mixture of potassium dihydrogen phosphate buffer (20. mM, pH 4.5) and acetonitrile (95:5, v/v), at an isocratic flow rate of 0.7. mL/min. The UV detector was set at 267. nm. The chromatographic run lasted 19. min. Similar pyrimidine nucleotides and nucleosides do not interfere with the assay. Calibration curves were linear for both dThd and dUrd over a range of 0.5 to 5.0. μg/mL. The limit of quantitation was 0.5. μg/mL for both nucleosides and the absolute recovery was >90% for dThd, dUrd and the I.S. Both intra- and inter-assay precision and accuracy were lower than 10% at all tested concentrations. The proposed method was successfully applied to measure plasma concentrations of dThd and dUrd in two MNGIE patients. This assay simplifies both plasma pretreatment and chromatographic conditions of previously reported procedures and describes the first validated method for the determination of the two nucleotides in human plasma. © 2014 Elsevier B.V.

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