The effect of acidic beverage versus mineral water on the change in serum phenobarbital concentrations: a randomized clinical trial on children with seizure.

OBJECTIVE: To assess the effect of an acidic beverage (Orange juice) on the change in serum Phenobarbital concentrations in children with seizure who take Phenobarbital as the main treatment. METHODS: We did a parallel design and placebo controlled randomized clinical trial. Patients attending Heshmatiyeh Hospital (Iran) were recruited from October 2016 to December 2017. Forty patients randomly assigned to either experimental group or control group. Firstly, 5 mL blood sample was taken from both groups to measure serum Phenobarbital concentration before experiment. Then, one oral dose of Phenobarbital (2.5 mg/kg) with 100 mL of corporate Orange juice (pH = 3.5) (experiment group) or 100 mL of mineral water (neutral pH) (control group) was given to each group, respectively. After 2 h of administration, another blood sample was taken. The high-performance liquid chromatographic system was used for measurement of serum Phenobarbital concentration. RESULTS: There was significant increase in serum Phenobarbital concentrations after taking Phenobarbital in experiment group in comparison to control group. Statistical analysis revealed a significant increase in change of serum Phenobarbital concentrations in experiment group versus control group. CONCLUSION: The results of the current trial indicate that the level of serum Phenobarbital in the experiment group was higher than that of control group.

Analytical validation of a quantitative method for therapeutic drug monitoring on the Alinity®c Abbott.

OBJECTIVE: The aim of this study was to evaluate the analytical performance of the Alinity®c Abbott compared to the Architect® immunoassay system for the determination of drugs having a narrow therapeutic index. METHODS: Valproic acid, amikacin, gentamicin, phenobarbital and vancomycin were analyzed using Particle-Enhanced Turbidimetric Inhibitor Immunoassay (Petinia), phenytoin and theophylline were analyzed using an immunoenzymatic method and a colorimetric method was performed to quantify lithium. The methods were validated according to the total error approach. Seven validation standards were analyzed in quintuplet during four days to establish the limits of the methods. Dilution integrity and interferences (hemolysis and high concentrations of bilirubin and lipids) were also tested. Depending on the analyte, the results obtained for twenty to forty patients on the Alinity® were compared to those obtained on the Architect®. RESULTS: The bias and the coefficients of variation for repeatability and for intermediate precision were lower than 15% for all drugs. Accuracy profiles were acceptable (acceptance limits fixed at 30%) in the validated ranges. The lower limits of quantification (LLOQ) were similar to those determined by Abbott except for gentamicin for which we determined a LLOQ at 1.22 mg/L while Abbott determined it at 0.5 mg/L. All assays diluted linear and analyte concentrations were not affected by interferences. Concentrations obtained for real samples on the Alinity®c are comparable to those obtained on the Architect®ci. CONCLUSIONS: The analytical validation of a method suitable for therapeutic drug monitoring of drugs on the Alinity®c meets the requirements of European Medicines Agency.

Treatment of phenobarbital intoxication using hemodialysis in two dogs.

OBJECTIVE: To describe the use of hemodialysis in 2 dogs with severe clinical signs from phenobarbital intoxication. SERIES SUMMARY: Two dogs ingested a toxic dose of phenobarbital, leading to severe neurological dysfunction and a comatose state. Both dogs received a 3-hour session of hemodialysis with complete resolution of clinical signs and returned to normal mentation by the end of the therapy. No negative side effects occurred and phenobarbital concentrations returned to therapeutic range during treatment. NEW INFORMATION PROVIDED: This is the first report on the utility and safety of using hemodialysis for phenobarbital intoxication in dogs.

Development and validation of a method for phenobarbital in serum: anticonvulsant pharmacotherapy monitoring

The therapeutic drug monitoring (TDM) is an important strategy for the effectiveness and safety of long-term pharmacotherapy, such as the use of phenobarbital as an anticonvulsant drug in epilepsy. In this sense, HLPC has been presented as a technique for the measurement of phenobarbital in serum. However, the ideal conditions for carrying out the method must be established for each laboratory reality. An analytical method using HPLC was developed and validated in order to identify and quantify Phenobarbital in blood. The chromatographic conditions were C-18 column (Shimpack XR-ODS 50L x 3.0), acetonitrile-water mobile phase (30:70, v v-1), 0.2 mL min-1 flow and reading wavelength of 210 nm. Linearity was established in the range of 2.5 to 80 µg mL-1, the linear correlation coefficient was 0.9981. The average of the coefficient of variation of the precision was 5.30%. The relative standard error of the accuracy was -2.17% and of the recovery coefficient was 97.83%. In all eleven patients, phenobarbital concentrations were below the therapeutic range. The tested method was selective, linear, precise, accurate and showed good recovery.(AU)

Dried Blood Microsampling-Based Therapeutic Drug Monitoring of Antiepileptic Drugs in Children With Nodding Syndrome and Epilepsy in Uganda and the Democratic Republic of the Congo.

Nodding syndrome is a highly debilitating, generalized seizure disorder, affecting children in subregions of sub-Saharan Africa. Despite numerous efforts to uncover the etiology, the exact cause of this syndrome still remains obscure. Therefore, to date, patients only receive symptomatic care, including the administration of first-generation antiepileptic drugs for seizure control. As data on the efficacy of drugs within this population are completely lacking, the aim of this study was to explore how therapeutic drug monitoring could help to understand the differential response to therapy. Considering the challenging environment in which sampling had to be performed (remote areas, devoid of electricity, running water, etc), dried blood matrices [ie, dried blood spots (DBSs)] and volumetric absorptive microsampling (VAMS) were considered fit-for-purpose. In addition, owing to the similarities between the syndrome and other forms of epilepsy, samples originating from patients suffering from (onchocerciasis-associated) epilepsy were included. In total, 68 patients with Nodding syndrome from Uganda, 58 Ugandan patients with epilepsy, and 137 patients with onchocerciasis-associated epilepsy from the Democratic Republic of the Congo were included. VAMS samples and DBS were analyzed using validated methods, involving manual extraction or fully automated extraction, respectively, before quantification using liquid chromatography coupled with tandem mass spectrometry. Analysis revealed that serum concentrations (calculated from DBS) within the respective reference ranges were attained in only 52.9% of the 68 Nodding syndrome patients treated with valproic acid, in 21.4% of the 56 Ugandan epilepsy patients treated with carbamazepine, and in 65.7% of the 137 onchocerciasis-associated epilepsy patients from the Democratic Republic of the Congo treated with phenobarbital. In all other instances, concentrations were subtherapeutic. Furthermore, on comparing DBS with VAMS concentrations, an inexplicable overestimation was observed in the latter. Finally, no obvious link could be observed between the obtained drug concentrations and the number of seizures experienced during the last month before sampling, elaborating the fact that the level of improvement in some patients cannot simply be linked to reaching therapeutic concentrations.

Phenobarbital, Midazolam Pharmacokinetics, Effectiveness, and Drug-Drug Interaction in Asphyxiated Neonates Undergoing Therapeutic Hypothermia.

BACKGROUND: Phenobarbital and midazolam are commonly used drugs in (near-)term neonates treated with therapeutic hypothermia for hypoxic-ischaemic encephalopathy, for sedation, and/or as anti-epileptic drug. Phenobarbital is an inducer of cytochrome P450 (CYP) 3A, while midazolam is a CYP3A substrate. Therefore, co-treatment with phenobarbital might impact midazolam clearance. OBJECTIVES: To assess pharmacokinetics and clinical anti-epileptic effectiveness of phenobarbital and midazolam in asphyxiated neonates and to develop dosing guidelines. METHODS: Data were collected in the prospective multicentre PharmaCool study. In the present study, neonates treated with therapeutic hypothermia and receiving midazolam and/or phenobarbital were included. Plasma concentrations of phenobarbital and midazolam including its metabolites were determined in blood samples drawn on days 2-5 after birth. Pharmacokinetic analyses were performed using non-linear mixed effects modelling; clinical effectiveness was defined as no use of additional anti-epileptic drugs. RESULTS: Data were available from 113 (phenobarbital) and 118 (midazolam) neonates; 68 were treated with both medications. Only clearance of 1-hydroxy midazolam was influenced by hypothermia. Phenobarbital co-administration increased midazolam clearance by a factor 2.3 (95% CI 1.9-2.9, p < 0.05). Anticonvulsant effectiveness was 65.5% for phenobarbital and 37.1% for add-on midazolam. CONCLUSIONS: Therapeutic hypothermia does not influence clearance of phenobarbital or midazolam in (near-)term neonates with hypoxic-ischaemic encephalopathy. A phenobarbital dose of 30 mg/kg is advised to reach therapeutic concentrations. Phenobarbital co-administration significantly increased midazolam clearance. Should phenobarbital be substituted by non-CYP3A inducers as first-line anticonvulsant, a 50% lower midazolam maintenance dose might be appropriate to avoid excessive exposure during the first days after birth.

Identification of phenobarbital and other barbiturates in forensic drug screening using positive electrospray ionization liquid chromatography-high resolution mass spectrometry.

Comprehensive drug-screening performed by liquid chromatography-high resolution mass spectrometry (LC-HRMS) enables identification of hundreds to thousands of drug compounds in a single analysis. Forensic drug screening is generally performed with positive electrospray ionization (ESI+ ), targeting basic drugs; however, a few toxicologically important drugs such as barbiturates, may require analysis by negative ESI. In this work, screening targets for barbiturates were determined using our LC-HRMS screening with ESI+ . For several years, our forensic whole blood samples have been analyzed using the LC-HRMS-ESI+ screening in parallel with a multi-target LC-MS/MS-ESI- method. From 2014 to 2018, 23 samples were positive for phenobarbital (0.5-81 mg/kg). Retrospective data analysis of 4816 blood samples (15 positive) revealed several potential screening targets for phenobarbital. The targets were tentatively identified by exact mass and isotopic pattern as uncommon adducts of phenobarbital and as a decomposition product of phenobarbital N-glucoside (C17 H24 N2 O7 ). Analysis of a test set containing eight positive (0.5-65 mg/kg phenobarbital) and 31 negative samples supported the use of the observed target m/z 323.0614 at 5.14 minutes, corresponding to the [M + HCOONa+Na]+ adduct of phenobarbital. The [M + HCOONa+Na]+ adduct was confirmed as a screening target for common barbiturates, by analysis of barbiturate reference standards in ESI+ /ESI- . The [M + HCOONa+Na]+ adduct allowed retrospective analysis with 91% sensitivity (n = 23) and 100% specificity (n = 4855) for phenobarbital in our existing LC-HRMS-ESI+ screening. The two negative results were the two whole-blood samples with the lowest phenobarbital concentration (<1.8 mg/kg). Thus, a specialized screening is not necessary and use of this adduct likely enables screening for other barbiturates.

Blood concentration, efficacy, and adverse events of phenobarbital: A prospective study in rural China.

OBJECTIVE: This study evaluated the relationship between blood concentration of phenobarbital (PB) and its efficacy as well as adverse events in people with epilepsy in rural China. METHODS: People with epilepsy being treated with PB monotherapy were recruited and followed up for averagely 2.5 years. Data of clinical characteristics were collected using a standardized questionnaire by face-to-face interviews both at baseline and follow-up. Plasma concentration of PB was detected by the high-performance liquid chromatography. RESULTS: Data on treatment response and PB blood concentration was obtained from 225 subjects. Among them, 119 (52.9%) were recognized as effective cases and 106 (47.1%) as ineffective cases. In the effective group, the blood concentration of 95% subjects ranged from 1.22 µg/ml to 41.36 µg/ml with a median at 13.18 µg/ml (IQR = 8.32-20.19 µg/ml). The PB concentration of 95% of the subjects in the ineffective group ranged from 2.73 µg/ml to 70.16 µg/ml with a median at 19.80 µg/ml (IQR = 11.30-30.40 µg/ml), which was significantly higher than that of the effective group (p < 0.001). Multivariate logistic regression analysis showed that PB concentration ≥26.38 µg/ml was related to a 4.5-fold (95% confidence interval [CI], 1.85-11.08) higher risk of inefficacy. A receiver operation characteristic curve was performed to determine the cutoff value of concentration for PB efficacy at 19.02 µg/ml. SIGNIFICANCE: Blood concentration may be an important indicator for clinical decision making when PB monotherapy cannot achieve a good efficacy and more attention should be paid on it in clinical practice especially in resource-poor settings.

Reduced Clearance of Phenobarbital in Advanced Cancer Patients near the End of Life.

BACKGROUND AND OBJECTIVES: Little is known about the pharmacokinetics of phenobarbital in terminally ill cancer patients. We investigated whether phenobarbital clearance alters depending on the length of survival. METHODS: We retrospectively reviewed the clinical, laboratory, and therapeutic drug monitoring (TDM) records of patients who received parenteral or oral phenobarbital for 21 consecutive days or longer between 2000 and 2016. Patients were divided into non-cancer and cancer groups. Cancer patients were further stratified according to the survival interval after TDM: those who survived > 3 months were classified as long-surviving and the remainders short-surviving cancer patients. Phenobarbital clearance (CLPB) was calculated at steady state. Multiple comparisons of median CLPB were conducted among the three groups. RESULTS: Data were collected from 44 non-cancer patients and 34 cancer patients comprising 24 long-surviving and 10 short-surviving cancer patients. Among 10 short-surviving cancer patients, 4 had hepatic metastasis. Median CLPB (range) in short-surviving cancer patients [0.076 (0.057‒0.114) L/kg/day] was significantly (p < 0.05) lower than that in non-cancer patients [0.105 (0.060‒0.226) L/kg/day] and in long-surviving cancer patients [0.100 (0.082‒0.149) L/kg/day]. CONCLUSION: Terminally ill patients with advanced cancer may have reduced CLPB, thereby TDM is recommended for these patients particularly near the end of life.

The effectiveness of phenobarbital in patients with refractory status epilepticus undergoing therapeutic plasma exchange.

This study aimed to elucidate the therapeutic concentration range of phenobarbital (PB) for adults, as well as the influence of therapeutic plasma exchange (TPE) on plasma concentration of PB. We retrospectively reviewed consecutive patients diagnosed with refractory status epileptic (RSE) and treated with a bolus injection of PB as well as TPE, admitted to our neurocritical care unit from November 2015 to October 2016. Continuous electroencephalographic monitoring was performed routinely for these patients. TPE was performed using a continuous-flow cell separator. PB concentrations in the plasma and cerebrospinal fluid were measured by gas chromatography-mass spectrometer analysis before and after TPE. A total of seven patients were included; among these, one patient had RSE related to anti-N-methyl-D-aspartate receptor encephalitis, another patient had Hashimoto encephalopathy, and five patients had undetermined etiology. For patients with clinical and electrographic control (n=6), the plasma concentration of PB ranged from 138 to 243.7 µg/ml. In addition, of six paired plasma samples (before and after TPE) obtained from three patients, no significant differences between the concentrations of PB were detected (P=0.6), suggesting that TPE may not significantly affect the plasma concentration of PB. This study confirmed that PB more than 100 µg/ml was effective for adults with RSE. Moreover, TPE may not have an influence on the plasma concentration of PB.Video abstract: http://links.lww.com/WNR/A489.

Comparison of CYP3A4-Inducing Capacity of Enzyme-Inducing Antiepileptic Drugs Using 4ß-Hydroxycholesterol as Biomarker.

BACKGROUND: Enzyme-inducing antiepileptic drugs (EIAEDs) are among the clinically most important inducers of cytochrome P450 (CYP) 3A4, but there is limited evidence regarding the comparative potency of each EIAED in raising CYP3A4 activity. The aim of this study was to estimate CYP3A4-inductive potency of EIAEDs by comparing CYP3A4 activity in patients treated with carbamazepine, phenobarbital, or phenytoin. METHODS: Residual serum samples from patients treated with EIAEDs or levetiracetam were collected from a therapeutic drug monitoring service for analysis of 4ß-hydroxycholesterol (4ßOHC), which is an indicator of CYP3A4 activity. The samples were collected between January and September 2016 at Diakonhjemmet Hospital, Oslo, Norway. Concentration of 4ßOHC, EIAEDs, and levetiracetam was measured by ultra-performance liquid chromatography tandem mass spectrometry. Kruskal-Wallis and Mann-Whitney tests were used for comparison of 4ßOHC levels between the subgroups. RESULTS: In total, 4ßOHC measurements for 343 and 339 patients treated with EIAEDs and levetiracetam, respectively, were included in the study. Compared with levetiracetam-treated patients, the median 4ßOHC concentration was 3.3-fold, 5.8-fold, and 6.9-fold higher in patients using phenobarbital, phenytoin, or carbamazepine, respectively (P < 0.0001). Phenytoin users (n = 65) and carbamazepine users (n = 225) had 1.8- and 2.1-fold higher median 4ßOHC concentration than phenobarbital users (n = 28), respectively (P ≤ 0.0001). CONCLUSIONS: This study shows that phenytoin and carbamazepine have approximately twice the CYP3A4-inducing potency of phenobarbital. The results indicate that 2-fold higher doses of CYP3A4-metabolized drugs may generally be required during concurrent treatment with phenytoin or carbamazepine compared with phenobarbital.

A potential drug interaction between phenobarbital and dolutegravir: A case report.

In this report, we describe a human immunodeficiency virus (HIV)-infected patient in whom changes in phenobarbital (PB) dosage resulted in associated changes in plasma concentrations of dolutegravir (DTG). His plasma concentrations of DTG were 0.934, 0.584, 1.003 and 3.25 µg/mL, respectively, with concomitant daily PB doses of 40, 70, 30 and 0 mg, respectively. This case suggests that PB can lead to a remarkable reduction in the plasma concentration of DTG in a dose-dependent manner.

Comparison of the effects of sodium phenobarbital in wild type and humanized constitutive androstane receptor (CAR)/pregnane X receptor (PXR) mice and in cultured mouse, rat and human hepatocytes.

Phenobarbital (PB), a constitutive androstane receptor (CAR) activator, produces liver tumours in rodents by a mitogenic mode of action involving CAR activation. In this study, the hepatic effects of sodium phenobarbital (NaPB) were compared in male C57BL/6J wild type (WT) mice and in humanized mice, where both the mouse CAR and pregnane X receptor (PXR) have been replaced by their human counterparts (hCAR/hPXR mice). Investigations were also performed in cultured male C57BL/6J and CD-1 mouse, male Sprague-Dawley rat and male and female human hepatocytes. The treatment of WT and hCAR/hPXR mice with 186-984 ppm NaPB in the diet for 7 days resulted in increased relative liver weight, hypertrophy and induction of cytochrome P450 (CYP) enzyme activities. Treatment with NaPB also produced dose-dependent increases in hepatocyte replicative DNA synthesis (RDS), with the effect being more marked in WT than in hCAR/hPXR mice. While the treatment of cultured C57BL/6J and CD-1 mouse, Sprague-Dawley rat and human hepatocytes with 100 and/or 1000 µM NaPB for 4 days induced CYP enzyme activities, increased RDS was only observed in mouse and rat hepatocytes. However, as a positive control, epidermal growth factor increased RDS in hepatocytes from all three species. In summary, although human hepatocytes are refractory to the mitogenic effects of NaPB, treatment with NaPB induced RDS in vivo in hCAR/hPXR mice, which is presumably due to the human CAR and PXR receptors operating in a mouse hepatocyte regulatory environment. As the response of the hCAR/hPXR mouse to the CAR activator NaPB differs markedly from that of human hepatocytes, the hCAR/hPXR mouse is thus not a suitable animal model for studies on the hepatic effects of nongenotoxic rodent CAR activators.

Modified dispersive liquid-phase microextraction based on sequential injection solidified floating organic drop combined with HPLC for the determination of phenobarbital and phenytoin.

A modified dispersive liquid phase microextraction based on sequential injection solidified floating organic drop was developed for simultaneous separation/preconcentration of trace amounts of phenobarbital and phenytoin. The important factors affecting on the extraction recovery including pH, the volume of extraction solvent, ionic strength, and the number of injections were investigated and optimized by Box-Behnken design and desirability function. Under the optimum experimental conditions, the calibration graph was linear in the concentration range of 1.0-300.0 µg/L (r2  = 0.997) for phenobarbital and 2.0-400.0 µg/L (r2  = 0.996) for phenytoin. The limit of detection and limit of quantification were 0.35 and 1.2 µg/L for phenobarbital and 0.65 and 2.2 µg/L for phenytoin, respectively. The relative standard deviation for six replicate determinations at 10 µg/L was 3.3 and 4.1% for phenobarbital and phenytoin, respectively. The developed method was successfully applied to the determination of phenobarbital and phenytoin in urine and plasma samples.

Characterization and application of restricted access carbon nanotubes in online extraction of anticonvulsant drugs from plasma samples followed by liquid chromatography analysis.

Anticonvulsant drugs are often used in the treatment of epilepsy. However, their therapeutic monitoring is often necessary in order to obtain an appropriate dose adjustment, due to the proximity between their therapeutic and toxic ranges. The aim of this study was to carry out the synthesis, characterization and use of restricted access carbon nanotubes (RACNTs) in an online method for the analyses of phenobarbital and carbamazepine and primidone from untreated human blood plasma by column switching liquid chromatography. Therefore, the synthesis of RACNTs was carried out through coating commercial Carbon nanotubes with bovine serum albumin (BSA) to subsequently use them as adsorbents in a column switching system operating in the backflush mode. This material was evaluated through the construction of the kinetic and isotherm curves. The experimental data for the interaction of primidone with RACNTs were adequately adjusted to the chemisorption and Sips models for the kinetic and adsorption studies, respectively. The analytical curves ranged from 2.0 to 40.0mgL-1, with correlation coefficients higher than 0.99, for all the analytes. The LODs of 0.1, 0.1 and 0.01µgmL-1 were defined for PHB, PRM and CBZ, respectively. The relative standard deviation values ranged from 1.0% to 8.4% for the intra assay precision and from 2.7% to 7.6% for inter assay precision. The relative error values ranged from -13.4% to 7.7% for the intra assay accuracy and from -8.6% to 2.5% for the inter assay accuracy. The method was adequately used in the therapeutic monitoring of anticonvulsant drugs in human plasma samples.

Standardized Treatment of Neonatal Status Epilepticus Improves Outcome.

We aimed to decrease practice variation in treatment of neonatal status epilepticus by implementing a standardized protocol. Our primary goal was to achieve 80% adherence to the algorithm within 12 months. Secondary outcome measures included serum phenobarbital concentrations, number of patients progressing from seizures to status epilepticus, and length of hospital stay. Data collection occurred for 6 months prior and 12 months following protocol implementation. Adherence of 80% within 12 months was partially achieved in patients diagnosed in our hospital; in pretreated patients, adherence was not achieved. Maximum phenobarbital concentrations were decreased (56.8 vs 41.0 µg/mL), fewer patients progressed from seizures to status epilepticus (46% vs 36%), and hospital length of stay decreased by 9.7 days in survivors. In conclusion, standardized, protocol-driven treatment of neonatal status epilepticus improves consistency and short-term outcome.

Serum Phenobarbitone Levels in Term and Near Term Neonates with Seizures.

OBJECTIVE: To evaluate serum phenobarbitone levels in neonates with seizures and to evaluate the effect of repeated loading dose on serum phenobarbitone levels. METHODS: In this prospective observational study conducted in a tertiary care centre of Northern India during 2011- 2012, 99 neonates admitted with seizureswere included.Serum phenobarbitone levels in neonates with seizures at 20 minutes and 12 hours after the first loading dose of phenobarbitone were measured. RESULTS: Serum phenobarbitone levels [mean (SD)] at 20 min and 12 hours was 27.3 (28.4) ug/mL and 23 (19.1) ug/mL, respectively (P=0.07). The mean serum phenobarbitone levels 12 hours after the loading dose, and proportion of neonates with toxic levels increased with each loading dose of intravenous phenobarbitone. CONCLUSION: Monitoring of serum level of phonobarbitone may not be essential because seizure control in neonates appears to be independent of whether serum level is subtherapeutic, therapeutic or toxic range.

Development, validation and clinical application of an online-SPE-LC-HRMS/MS for simultaneous quantification of phenobarbital, phenytoin, carbamazepine, and its active metabolite carbamazepine 10,11-epoxide.

A simple and efficient bioanalytical method for simultaneous determination of phenobarbital (PB), phenytoin (PHT), carbamazepine (CBZ), and its active metabolite carbamazepine 10,11-epoxide (CBZE) in human plasma using online solid phase extraction (SPE)-liquid chromatography (LC) coupled with high resolution mass spectrum (HRMS) under targeted MS/MS (t-MS(2)) analysis mode has been developed. The procedure integrated an automated sample clean-up of human plasma by Oasis®HLB SPE cartridge, a separation by ZORBAX SB-C18 analysis column, and a quantification by Q-Exactive Hybrid Quadrupole-Orbitrap. The total running time was 13min. The lower limit of quantification (LLOQ) of PB, PHT, CBZ, and CBZE were 0.008, 0.008, 0.0016 and 0.0016µgmL(-1) respectively and the linearities were in the range of 0.008-2.500, 0.008-2.500, 0.0016-0.500 and 0.0016-0.500µgmL(-1) respectively. The mean recovery was between 91.82% and 108.27% and the matrix effect was between 93.29% and 102.09%. The relative standard deviations of interday and intraday were less than 6.41%. The method has been successfully applied in therapeutic drug monitoring (TDM) of four Chinese epilepsy patients. This fully automated, simple, sensitive and reliable online-SPE-LC-HRMS/MS method serves well for TDM of PB, PHT, CBZ and CBZE at clinics for either single or combination treatment.