Simultaneous UPLC-MS/MS assay for the detection of the traditional antipsychotics haloperidol, fluphenazine, perphenazine, and thiothixene in serum and plasma.

BACKGROUND: Most antipsychotic drugs that are commonly prescribed in the USA are monitored by liquid and gas chromatographic methods. Method performance has been improved using ultra high pressure liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). A rapid and simple procedure for monitoring haloperidol, thiothixene, fluphenazine, and perphenazine is described here. METHOD: Antipsychotic drug concentrations in serum and plasma were determined by LCMS/MS (Waters Acquity UPLC TQD). The instrument is operated with an ESI interface, in multiple reaction monitoring (MRM), and positive ion mode. The resolution of both quadrupoles was maintained at unit mass with a peak width at half height of 0.7amu. Data analysis was performed using the Waters Quanlynx software. Serum or plasma samples were thawed at room temperature and a 100µL aliquot was placed in a tube. Then 300µL of precipitating reagent (acetonitrile-methanol [50:50, volume: volume]) containing the internal standard (0.12ng/µL Imipramine-D3) was added to each tube. The samples were vortexed and centrifuged. The supernatant was transferred to an autosampler vial and 8µL was injected into the UPLC-MS/MS. Utilizing a Waters Acquity UPLC HSS T3 1.8µm, 2.1×50mm column at 25ºC, the analytes were separated using a timed, linear gradient of acetonitrile and water, each having 0.1% formic acid added. The column is eluted into the LC-MS/MS to detect imipramine D3 at transition 284.25>89.10, haloperidol at 376.18>165.06, thiothixene at 444.27>139.24, fluphenazine at 438.27>171.11, and perphenazine at 404.19>143.07. Secondary transitions for each analyte are also monitored for imipramine D3 at 284.25>193.10, haloperidol at 376.18>122.97, thiothixene at 444.27>97.93, fluphenazine at 438.27>143.08, and perphenazine at 404.19>171.11. The run-time is 1.8min per injection with baseline resolved chromatographic separation. RESULTS: The analytical measurement range was 0.2 to 12.0ng/mL for fluphenazine and perphenazine, and was 1 to 60.0ng/mL for haloperidol and thiothixene. Intra-assay and inter-assay imprecisions (CV) were less than 15% at two concentrations for each analyte. CONCLUSIONS: By utilizing a LC-MS/MS method we combined two previously established analytical assays into one, yielding a 75% time-savings on set-up, and a significantly shortened analytical run-time. These changes reduced the turn-around time for analysis and eliminated interference issues resulting in fewer injections and increased column lifetime.

Simultaneous quantification of levetiracetam and gabapentin in plasma by ultra-pressure liquid chromatography coupled with tandem mass spectrometry detection.

INTRODUCTION: Gabapentin (Neurontin) and levetiracetam (Keppra) are anticonvulsants with novel structures and suggested therapeutic ranges of 2-10 mg/L and 6-20 mg/L, respectively. Gabapentin is also used extensively to manage neuropathic pain, and for this indication, wherein higher doses are prescribed, plasma concentrations of 15-30 mg/L are typical. OBJECTIVE: Here, we describe a simple rapid assay to support therapeutic drug monitoring of gabapentin and levetiracetam in plasma by ultra-pressure liquid chromatography couples to tandem mass spectrometry (UPLC-MS/MS) detection. METHODS: After the addition of internal standard and protein precipitation of patient plasma with methanol:acetonitrile in a 50:50 ratio, 1 µL of supernatant sample is injected onto an Acquity UPLC HSS T3, 1.8 µm, 2.1 × 50 mm (Waters) column. Elution occurs using a linear gradient of acetonitrile and water, each having 0.1% formic acid added. The column is eluted into a Waters Acquity UPLC TQD, operating in a positive mode to detect gabapentin at transition 172.18 > 154.11, levetiracetam at 171.11 > 126, and internal standard (3-amino-2-naphthoic acid) at 188.06 > 170. Secondary transitions for each analyte are also monitored for gabapentin at 172.18 > 137.06, levetiracetam at 171.11 > 154, and internal standard at 188.06 > 115. Runtime is 1.5 minutes per injection with baseline resolved chromatographic separation. RESULTS: The analytical measurement ranges were 1-150 mg/L for gabapentin and for levetiracetam. Intra-assay imprecision by the coefficient of variance (CV) was less than 8% and interassay CV was less than 5% for both analytes, at 4 different concentrations. Results obtained from patient samples were compared with results generated by established high-performance liquid chromatography-UV methods with the following regression statistics: y = 1.12x - 0.77, r = 0.996, Sy, x = 0.89, and n = 29 for gabapentin and y = 0.991x + 0.70, r = 0.997, Sy, x = 2.24, and n = 30 for levetiracetam. No analytical interferences were identified. CONCLUSION: : In summary, a simple reliable UPLC-MS/MS method was developed and validated for routine clinical monitoring of gabapentin and levetiracetam.

Drug monitoring and toxicology: a procedure for the monitoring of oxcarbazepine metabolite by HPLC-UV.

This article describes a rapid high-performance liquid chromatographic (HPLC) method for the measurement of the primary metabolite of oxcarbazepine. Following a simple precipitation step, 10,11,-dihydro-10-hydroxy-5H-dibenzo(b,f)azepine-5-carboxamide is quantitated (5-60 microg/mL) by analysis on an HPLC-UV system. The instrument time is less than 5 min per injection, an improvement over most published methods. The assay's limit of quantitation, linearity, imprecision, and accuracy adequately cover the therapeutic range for appropriate patient monitoring. In comparison to other published methods, this procedure would be of interest to clinical laboratories because it employs a precipitation step for sample preparation, instead of conventional yet time-consuming solid-phase extraction.

Drug monitoring and toxicology: a procedure for the monitoring of levetiracetam and zonisamide by HPLC-UV.

This article describes a rapid isocratic high-performance liquid chromatographic (HPLC) method for the simultaneous measurement of the anticonvulsants levetiracetam and zonisamide. Monitoring these drugs is important for detecting potentially toxic concentrations, particularly in patients with renal impairment, but no commercial assays are currently available. Following a liquid-liquid extraction, levetiracetam (5-150 microg/mL) and zonisamide (5-80 microg/mL) are quantitated by HPLC-UV. The assay's limit of quantitation, linearity, imprecision, and accuracy adequately cover the therapeutic range of these drugs. The assay should be attractive to clinical laboratories because the run time for quantification of both drugs is approximately 5 min per sample, and no interferences are currently known.

A rapid procedure for the monitoring of amiodarone and N-desethylamiodarone by HPLC-UV detection.

This article describes a rapid isocratic high-performance liquid chromatographic (HPLC) method for the simultaneous measurement of the antiarrhythmic drug amiodarone and its potentially active metabolite N-desethylamiodarone (DEA). Following a simple liquid-liquid extraction, amiodarone and its metabolite are quantitated (0.3-6.0 mg/L) by analysis on an HPLC-UV system. The analytical time was reduced by 50%, without compromising the assay performance, when Rocket column technology was employed. The assay's limit of quantitation, linearity, imprecision, and accuracy adequately covered the therapeutic range for appropriate patient monitoring. Amiodarone and DEA can be simultaneously and accurately quantitated in serum or plasma by HPLC-UV detection with imprecision < 6% at therapeutic concentrations and a quantitation range from 0.3 to 6.0 mg/L. Monitoring of this drug can allow for effective use, while minimizing serious side effects.