Method validation of a tricyclic antidepressant drug panel in urine by UPLC-MS/MS.

UNLABELLED: Tricyclic antidepressant (TCA) drugs are also used as adjunctive therapy to treat chronic pain. To monitor patient compliance to therapy, urine specimens may be preferred since collection is non-invasive, and the specimen can provide a longer detection window. TCA drugs are frequently monitored by immunoassay; however, poor antibody specificity may compromise results. The purpose for this study was to develop a confirmation method for determining TCA in urine specimens by ultra-pressure liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Our method can quantify 9 TCA drugs in less than 5 min. Method validation experiments were performed, and the coefficient variation for inter- and intra-day precision was within 12% for each analyte at five different concentrations. Accuracy studies had good agreement with another laboratory that performs testing by GC/MS. HIGHLIGHTS: 1. Tricyclic antidepressants are commonly used to treat depression, anxiety, and neuropathic pain. 2. A confirmation method by ultra-pressure liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to determine TCA in urine specimens was developed. 3. Urine testing for TCA is preferred for monitoring adherence to therapy due to ease of collection.

Comparative pharmacokinetics and pharmacodynamics of doripenem and meropenem in obese patients.

BACKGROUND: Antimicrobial pharmacokinetic and pharmacodynamic data are limited in obesity. OBJECTIVE: To evaluate the steady-state pharmacokinetics and pharmacodynamics of doripenem and meropenem in obese patients hospitalized on a general ward. METHODS: Patients with a body mass index (BMI) ≥40 kg/m² or total body weight (TBW) ≥100 pounds over their ideal body weight randomly received doripenem 500 mg (1-hour infusion) or meropenem 1 g (0.5-hour infusion) every 8 hours. Differences in pharmacokinetic parameters were determined by unpaired t test. Monte Carlo simulations were performed for 500 mg and 1 g every 8 hours, infused over 1 and 4 hours for doripenem and 0.5 and 3 hours for meropenem. Probability of target attainment (PTA) was calculated using a pharmacodynamic target of 40% fT > MIC (free drug concentrations above the minimum inhibitory concentration [MIC]), and cumulative fraction of response (CFR) was calculated using MIC data for 8 Gram-negative pathogens. RESULTS: Twenty patients were studied. Volume of distribution at steady state, corrected for TBW, was significantly larger (0.18 ± 0.04 vs 0.13 ± 0.05 L/kg, P = .048) and systemic clearance was significantly faster for doripenem (11.7 ± 4.1 vs 8.1 ± 2.6 L/h, P = .03). PTA was >90% for all regimens at MICs ≤2 µg/mL. CFR was >90% for all regimens against 6 enteric Gram-negative pathogens and for 3 of 4 regimens for each drug against Pseudomonas aeruginosa. CONCLUSIONS: Doripenem and meropenem pharmacokinetics differ in obesity. However, currently approved dosing regimens provide adequate pharmacodynamic exposures for susceptible bacteria in obese patients.

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.

Quantification of tricyclic antidepressants using UPLC-MS/MS.

Depression is a psychiatric condition that affects about 120 million people worldwide and can interfere with independence and productivity in essentially all aspects of daily life. Depression is also associated with risk of self-harm, and ultimately suicide. Antidepressant medications are widely used to treat symptoms of depression. While there are several classes of antidepressants, therapeutic drug management (TDM) is most common for the tricyclic antidepressants (TCAs). TDM of TCAs is important due to wide inter-individual variability in pharmacokinetics, production of active metabolites, and a high risk of drug-drug interactions. In addition, TDM of some TCAs can be used to optimize dose, wherein concentration relationships are recognized for both therapeutic response and potentially life-threatening toxicity. In many clinical scenarios, TDM of TCAs is accomplished by currently available point of care or automated immunoassays that provide a "total" TCA concentration. However, these assays may not be adequately specific to meet the needs of all clinical scenarios, and hence, chromatographic separation and quantification of individual TCA parent drugs and active metabolites that may contribute to the "total" TCA concentration is sometimes required. This chapter describes an analytical method designed to detect and/or quantify clinically significant concentrations of nine TCAs (amitriptyline, nortriptyline, imipramine, desipramine, doxepin, nordoxepin, protriptyline, clomipramine, and norclomipramine) in serum or plasma, using ultra pressure liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The sample preparation employs a rapid protein precipitation with 50:50 MeOH:acetonitrile, high speed centrifugation, and injection of 5 µL of supernatant onto the instrument, with a 5 min run-time.

Steady-state pharmacokinetics of intravenous levetiracetam in neurocritical care patients.

STUDY OBJECTIVES: To characterize the steady-state pharmacokinetics of intravenous levetiracetam in neurocritical care patients requiring seizure prophylaxis after a neurologic injury and to determine which dosing regimens achieve serum concentrations within the recommended therapeutic range of 6-20 µg/ml. DESIGN. Prospective, open-label, steady-state pharmacokinetic study. SETTING: Neurocritical care unit in a tertiary care medical center. PATIENTS. Twelve adults (five men, seven women) admitted to the neurocritical care unit who required prophylactic anticonvulsant therapy after subarachnoid hemorrhage, subdural hematoma, or traumatic brain injury. INTERVENTION: Patients received an intravenous infusion of levetiracetam 500 mg over 15 minutes every 12 hours. MEASUREMENTS AND MAIN RESULTS: Serial blood samples were collected from all patients after a minimum of four doses of therapy. Serum levetiracetam concentrations were determined by ultraperformance liquid chromatography with tandem mass spectrometry detection, and pharmacokinetic data were analyzed by compartmental and noncompartmental methods. Monte Carlo simulations were performed for multiple levetiracetam dosing regimens to determine the probability of achieving a target trough concentration of 6 µg/ml or greater, 20 µg/ml or greater, and 6-20 µg/ml. The mean ± SD levetiracetam maximum serum concentration was 28.0 ± 8.0 µg/ml, minimum serum concentration 3.1 ± 1.8 µg/ml, half-life 5.2 ± 1.2 hours, systemic clearance 5.6 ± 1.8 L/hour, and volume of distribution at steady state 36.8 ± 6.3 L. Increasing the doses of levetiracetam increased the probability of achieving a target trough concentration of 6 µg/ml or greater but also increased the probability of achieving trough concentrations greater than 20 µg/ml. Levetiracetam doses of 1000 mg every 8 hours and 1500-2000 mg every 12 hours provided the highest probability of achieving a target trough concentration between 6 and 20 µg/ml. CONCLUSION: Compared with previously published results in healthy volunteers and adults in status epilepticus, levetiracetam systemic clearance was faster and the terminal elimination half-life was shorter in neurocritical care patients. Higher doses or more frequent dosing may be needed to achieve target trough concentrations of 6-20 µg/ml.

Rapid and specific quantification of ethylene glycol levels: adaptation of a commercial enzymatic assay to automated chemistry analyzers.

Ethylene glycol ingestion, accidental or intentional, can be a life-threatening emergency. Assays are not available from most clinical laboratories, and, thus, results often require many hours or days to obtain. Enzymatic assays, adaptable to automated chemistry analyzers, have been evaluated, but they have been plagued by analytic problems. With an alternative method of data analysis applied to an existing enzymatic assay, a modified assay was developed and validated on 2 different automated chemistry systems. Compared with a previously validated method based on gas chromatography with flame ionization detection, the modified enzymatic assay showed excellent agreement on patient samples (y = 1.0227x -1.24; r(2) = 0.9725), with a large analytic measuring range (2.5-300 mg/dL [0.4-48.4 mmol/L]). Interferences from propylene glycol, various butanediols, and other related compounds were almost entirely eliminated; when present, they generated error flags rather than falsely elevated ethylene glycol results. This modified assay should make it possible for more clinical laboratories to offer ethylene glycol measurements.

Performance characteristics of the ARK diagnostics gabapentin immunoassay.

BACKGROUND: Gabapentin is an antiepileptic drug used as adjunct therapy in the treatment of seizures. Absorption is saturable, and drug clearance can be reduced if patients have impaired renal function. Therapeutic drug monitoring can be useful for optimizing the dose in patients with impaired renal function, for evaluating individual patient absorption thresholds, and for monitoring compliance. Although chromatographic techniques have historically been used to support gabapentin monitoring, an immunoassay was recently introduced by ARK Diagnostics, for use with open channel chemistry analyzers. Here, we evaluated the immunoassay on a random access instrument. METHODS: The ARK gabapentin assay was validated using a Beckman AU400e automated chemistry analyzer. Imprecision was assessed with 5 replicates of 3 concentrations (2.5, 8.0, and 25.0 mg/L), analyzed for 4 days. The analytical measurement range was evaluated with duplicate measurements of a prepared sample (40.0 mg/L) that was serially diluted. Patients' results were compared with the results generated with a previously validated ultra-high performance liquid chromatography coupled to tandem mass spectrometry method (n = 45, range, 1.5-45.6 mg/L). RESULTS: The within-run and between-run coefficients of variation were ≤8.1%. The analytical measurement range was confirmed to be 1.5-40.0 mg/L, as stated by the manufacturer. The Deming regression for the results of 45 patients produced a correlation coefficient of 0.9987, a linear regression slope of 1.01, and an intercept of 0.24 when compared with the ultra-high performance liquid chromatography coupled to tandem mass spectrometry assay. CONCLUSIONS: The ARK immunoassay is suitable for the clinical use of monitoring gabapentin in serum or plasma on the Beckman AU400e.

A comparison of two FDA approved lamotrigine immunoassays with ultra-high performance liquid chromatography tandem mass spectrometry.

BACKGROUND: Lamotrigine is an anti-epileptic drug used as adjunct therapy for seizures. Lamotrigine is commonly used in pregnant women with epilepsy, a population in which therapeutic drug monitoring (TDM) is useful to optimize dose. Drug-drug interactions that can induce or inhibit metabolism or elimination and impaired hepatic function are also possible indications for lamotrigine TDM. Chromatographic techniques are currently used for performing most TDM of lamotrigine, but this may change, as automated immunoassays were recently introduced. METHODS: Immunoassays available through Seradyn and ARK Diagnostics were validated using a Beckman AU400e automated chemistry analyzer. The intra-day precision was accessed with 5 replicates of three quality control materials, and inter-assay precision was estimated by assaying the same material over 4 days. Linearity was evaluated by serially diluting a spiked sample and measuring it in duplicate. The 2 methods were compared with ultra high performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) using 44 authentic patient specimens. RESULTS: The intra-day (n=5) and inter-assay (n=20) coefficients of variation were ≤7.5% for the 3 levels tested. The analytical measurement ranges were confirmed as stated by the manufacturers (0 or 1-40 µg/ml). The percent recovery of the quality control materials and Deming regression for the 44 patient results showed good agreement of both immunoassays when compared to the UPLC-MS/MS assay. CONCLUSION: The lamotrigine assays studied here produced a slightly lower result than UPLC-MS/MS but were precise and easy to perform.

Discordant carbamazepine values between two immunoassays: carbamazepine values determined by ADVIA Centaur correlate better with those determined by LC-MS/MS than PETINIA assay.

Discordant carbamazepine values as determined by two different immunoassays may be due to different cross-reactivities with the active metabolite carbamazepine 10, 11-epoxide and may cause confusion in interpreting carbamazepine serum levels. In this study, we compared carbamazepine values in samples containing carbamazepine and the epoxide metabolite, as determined by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) and by two commercial carbamazepine immunoassays: the PETINIA and the ADVIA Centaur carbamazepine. Clinical specimens were used for the comparative studies wherein we determined carbamazepine concentrations using the PETINIA, ADVIA Centaur, and LC-MS/MS assays. We observed an excellent correlation between carbamazepine concentrations determined by the ADVIA Centaur and LC-MS/MS methods while carbamazepine values were overestimated using the PETINIA assay. When aliquots of drug-free serum were supplemented with clinically relevant concentrations of the carbamazepine epoxide metabolite, we observed negligible cross-reactivity of epoxide with the ADVIA Centaur assay but over 90% cross-reactivity with the PETINIA assay. We conclude that the ADVIA centaur assay accurately measures carbamazepine concentrations in plasma or serum and that the PETINIA assay significantly overestimates true carbamazepine concentration. Such discordance may cause confusion in interpreting serum carbamazepine levels.

An automated method for supporting busulfan therapeutic drug monitoring.

INTRODUCTION: Busulfan is a chemotherapeutic agent commonly used for myeloablative conditioning regimens such as in the treatment of chronic myelogenous leukemia. Busulfan dosing is complex due to wide interpatient variability in pharmacokinetics and a narrow therapeutic range. Although busulfan dose is normalized to body weight, therapeutic drug monitoring (TDM) using area under the plasma concentration curve is recommended after the first dose. A high busulfan area under the plasma concentration curve (>1500 µM·min) is associated with an increased risk for sinusoidal obstruction syndrome, and a suboptimal area under the plasma concentration curve (<900 µM·min) is associated with an increased risk for graft rejection or disease relapse. TDM of busulfan is not widely available due to the lack of commercially available and rapid methods to determine the area under the plasma concentration curve. METHODS: The purpose of this study was to evaluate the Roche cobas c 111 instrument, a photometric automated chemistry analyzer, using the Busulfan PCM assay from Saladax Biomedical Inc. The assay using this instrument was compared with an enzyme-linked immunosorbent assay (ELISA) from Saladax Biomedical Inc and the Olympus AU400e. Linearity and accuracy were evaluated between 175 and 1750 ng/mL. Imprecision was determined by analyzing 5 concentrations of standards twice a day for 20 days. RESULTS: Linearity for the Roche method had a slope and y-intercept of 1.050 and -5.5, respectively, and percent recovery ranged between 95% and 105%. Correlation between the Roche and ELISA platforms was analyzed by linear regression on 26 frozen patient samples. The results from the comparison of the methods based on the Roche and ELISA platforms were as follows: coefficient of determination (R2) was 0.9684, with a slope and y-intercept of 0.752 and 108.41, respectively. Correlation between the Roche and Olympus instruments was analyzed by linear regression and Bland-Altman plots. The coefficient of determination (R2) was 0.9942, with a slope and y-intercept of 1.035 and -41.3326, respectively. CONCLUSIONS: Availability of TDM of busulfan can be improved by the use of commercially available reagents and automated platforms.

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.

Estimation of carbamazepine and carbamazepine-10,11-epoxide concentrations in plasma using mathematical equations generated with two carbamazepine immunoassays.

Carbamazepine is metabolized to an active metabolite known as carbamazepine-10,11-epoxide, or simply the "epoxide" metabolite. The presence of this metabolite can have clinically significant implications in therapeutic drug monitoring of carbamazepine, but accurate quantification of the epoxide metabolite is currently limited to chromatographic techniques. In this study, mathematical equations are proposed for the estimation of carbamazepine and epoxide metabolite concentrations based on values generated by common carbamazepine immunoassays. Three immunoassays were studied: particle-enhanced turbidimetric inhibition immunoassay (PETINIA, Siemens Healthcare Diagnostics, Deerfield, IL), ADVIA Centaur (Siemens Healthcare Diagnostics), and a cloned enzyme donor immunoassay (CEDIA; Roche, Indianapolis, IN). Equations were based on observed cross-reactivity of epoxide with the PETINIA (average, 96.2%; range, 86.6%-105.7%) and epoxide cross-reactivity with the ADVIA Centaur assay (average, 6.5%; range, 5.3%-7.7%). In addition, equations were developed using average cross-reactivity of epoxide with the PETINIA and with the CEDIA. Values determined by calculation correlated well with carbamazepine and epoxide concentrations in supplemented and patient samples, for which values of carbamazepine (2.2-12.0 microg/mL [9-51 micromol/L]) and the epoxide metabolite (0.6-2.4 microg/mL) were also verified by liquid chromatography-tandem mass spectrometry.

Determination of busulfan in human plasma using an ELISA format.

High-dose busulfan is an important component of many bone marrow transplantation-preparative regimens. High busulfan plasma levels have been shown to increase the chance of venoocclusive disease and low levels are associated with recurrence of disease or graft rejection. Currently, busulfan levels are monitored by physical methods that are expensive and time consuming, resulting in relatively low overall use of busulfan testing for dose adjustment. Novel highly selective antibodies for busulfan have been generated and a microtiter plate immunoassay capable of quantifying busulfan levels in plasma has been developed. The assay was configured using a busulfan-horseradish peroxidase (HRP) conjugate as the reporter group and busulfan monoclonal antibodies. The assay requires 30 microL of plasma with no sample preparation. The immunoassay has a standard curve based on busulfan with a range of 75-2000 ng/mL. The time to first result is 30 minutes with up to 40 patient samples in duplicate; multiple plates can be run at once. The coefficient of variation (CV) on signal is <5% for an entire plate, and the 95% confidence interval for negative samples (n = 78) is below the lowest calibrator of 75 ng/mL. Cross-reactivity with the major inactive metabolites (tetrahydrothiophene, tetramethyl sulfone, and tetrahydrothiophene-3-ol-1,1-dioxide) was <0.1%. Results generated with clinical samples (n = 35 and n = 70) correlate well to gas chromatography-mass spectrometry (R = 0.976 and 0.985, respectively) with a slope of 1.05 +/- 0.05. This immunoassay method is suitable for determining levels of busulfan in human plasma. It offers the advantages of using a smaller sample size, does not require sample preparation, and is less labor intensive than other methods. The ability to make 240 determinations per hour enables effective and timely routine monitoring of busulfan levels in clinical practice.

An ImmunoChip prototype for simultaneous detection of antiepileptic drugs using an enhanced one-step homogeneous immunoassay.

The development and characterization of a one-step homogeneous immunoassay-based multiwell ImmunoChip is reported for the simultaneous detection and quantitation of antiepileptic drugs (AEDs). The assay platform uses a cloned enzyme donor immunoassay (CEDIA) and a Beta-Glo assay system for generation of bioluminescent signal. Results of the one-step CEDIA for three AEDs (carbamazepine, phenytoin, and valproic acid), in the presence of serum, correlate well with the values determined by fluorescence polarization immunoassay. CEDIA intra- and interassay coefficients of variation are less than 10%. A microfabrication process, xurography, was used to produce the multiwell ImmunoChip. Assay reagents were dispensed and lyophilized in a three-layer pattern. The multiwell ImmunoChip prototype was used to detect and quantify AEDs in serum samples containing all three drugs. Luminescent signals generated from each well were recorded with a charge-coupled device (CCD) camera. The assays performed on an ImmunoChip were fast (5 min), requiring only small volumes of both the reagents (<1 microl/well) and the serum sample. The ImmunoChip assay platform described in this article may be well suited for therapeutic monitoring of drugs and metabolites at the point-of-care setting.

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.

Quetiapine cross-reactivity with plasma tricyclic antidepressant immunoassays.

BACKGROUND: Toxicology screens obtained on patients who have overdosed on drugs frequently include tricyclic antidepressants (TCAs) as part of the evaluation. Quetiapine is an antipsychotic agent with structural similarity to the TCAs. OBJECTIVE: To determine whether quetiapine may cross-react with plasma TCA immunoassays in vitro using commonly available autoanalyzers. METHODS: Quetiapine stock solution was added to 9 separate samples of pooled drug-free human plasma to produce concentrations ranging from 1 to 640 ng/mL that were verified by gas chromatography. No quetiapine metabolites were present. Each spiked plasma sample was tested in a blinded fashion using the Abbott Tricyclic Antidepressant TDx Assay on the TDxFLx autoanalyzer in 2 separate laboratories, the Syva Emit tox Serum Tricyclic Antidepressant Assay on the AU400 autoanalyzer and the S TAD Serum Tricyclic Antidepressant Screen on the ACA-Star 300 autoanalyzer. The TDx assay is quantitative, while Emit and S TAD are qualitative screening assays with a threshold of 300 ng/mL for TCA positivity. The outcome of interest was a positive TCA result. RESULTS: The quantitative assay showed concentration-related TCA cross-reactivity beginning at quetiapine concentrations of 5 ng/mL. The 640-ng/mL spiked sample produced TCA results of 379 and 385 ng/mL in labs 1 and 2, respectively. The qualitative assays were screened as TCA positive at quetiapine concentrations of 160 and 320 ng/mL for the S TAD and Emit assays, respectively. CONCLUSIONS: Quetiapine cross-reacts with quantitative and qualitative plasma TCA immunoassays in a concentration-dependent fashion. Therapeutic use or overdose of quetiapine may result in a false-positive TCA immunoassay result.

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.