Journal of Chromatographic Science Advance Access published October 24, 2015 Journal of Chromatographic Science, 2015, 1–9 doi: 10.1093/chromsci/bmv146 Article
Article
Evaluation of the Effects of Ketoconazole and Voriconazole on the Pharmacokinetics of Oxcarbazepine and Its Main Metabolite MHD in Rats by UPLC–MS-MS Xinxin Chen1, Ermin Gu1, Shuanghu Wang2, Xiang Zheng1, Mengchun Chen1, Li Wang1, Guoxin Hu1, Jian-ping Cai3, and Hongyu Zhou1,* 1
Pharmacology Department, School of Pharmacy, Wenzhou Medical University, Wenzhou 325035, China, 2The Laboratory of Clinical Pharmacy, The People’s Hospital of Lishui, Lishui 323000, China, and 3The Key Laboratory of Geriatrics, Beijing Hospital & Beijing Institute of Geriatrics, Ministry of Health, Beijing 100730, China *Author to whom correspondence should be addressed. E-mail:
[email protected],
[email protected] Received 23 January 2015; Revised 16 July 2015
Abstract Oxcarbazepine (OXC), a second-generation antiepileptic drug, undergoes rapid reduction with formation of the active metabolite 10,11-dihydro-10-hydroxy-carbazepine (MHD) in vivo. In this study, a method for simultaneous determination of OXC and MHD in rat plasma using ultra-performance liquid chromatography with tandem mass spectrometry (UPLC–MS-MS) was developed and validated. Under given chromatographic conditions, OXC, MHD and internal standard diazepam were separated well and quantified by electrospray positive ionization mass spectrometry in the multiple reaction monitoring transitions mode. The method validation demonstrated good linearity over the range of 10–2,000 ng/mL for OXC and 5–1,000 ng/mL for MHD. The lower limit of quantification was 5 ng/mL for OXC and 2.5 ng/mL for MHD, respectively. The method was successfully applied to the evaluation of the pharmacokinetics of OXC and MHD in rats, with or without pretreatment by ketoconazole (KET) and voriconazole (VOR). Statistics indicated that KET and VOR significantly affected the disposition of OXC and MHD in vivo, whereas VOR predominantly interfered with the disposition of MHD. This method is suitable for pharmacokinetic study in small animals.
Introduction Oxcarbazepine (OXC, Figure 1) is a second-generation antiepileptic drug indicated in the treatment of partial seizures and generalized tonic–clonic seizures (1, 2). In a large number of clinical studies, OXC has shown to be equally effective as carbamazepine (CBZ, Figure 1) but with fewer adverse effects and drug interactions (3, 4). It is generally known that epilepsy is a chronic, even lifelong disease, and patients may suffer intercurrent conditions, including fungal diseases (5). Besides that, fungal infections in central nervous system are a major cause of epilepsy (6, 7). Thus, antiepileptic drugs are inevitably prescribed together with antifungal agents and frequently involved in drug interactions in clinical practice. In these scenarios,
the pharmacokinetics of antiepileptic drugs may be affected so that the altered concentrations in plasma or target tissues induce potential risks in therapeutic safety and efficacy (8). It has been documented that ketoconazole (KET, Figure 1) and voriconazole (VOR, Figure 1) interacted with other drugs by altering their pharmacokinetic progress via the hepatic cytochrome P3A4 (CYP3A4) and P-glycoprotein (9–11). In 1997, Spina et al. demonstrated that, an azole antifungal, KET inhibited the CYP3A4 and significantly increased plasma CBZ concentrations in humans (8). OXC is chemically derived from CBZ, but there are significant differences in elimination pathways between the two drugs. Unlike CBZ, which is oxidized by the cytochrome P450 (P450) system, OXC is rapidly reduced by
© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email:
[email protected]
1
2
Chen et al.
Figure 1. Structure of main compounds. OXC, oxcarbazepine; MHD, 10,11-dihydro-10-hydroxy-carbamazepine; IS, internal standard, diazepam; CBZ, carbazepine; KET, ketoconazole; VOR, voriconazole. This figure is available in black and white in print and in color at JCS online.
cytosolic arylketone reductases in liver to form a pharmacologically active metabolite, 10,11-dihydro-10-hydroxy-carbazepine (MHD, Figure 1), and the MHD is subsequently glucuronidated and excreted in the urine. That is to say, the hepatic P450-dependent enzymes are rarely involved in the metabolism of OXC (12). Based on such pharmacokinetic properties, OXC is considered less likely to interact with other drugs. Here, what we are interested is, whether OXC escapes from the impacts of KET or VOR, because of its distinctive elimination from CBZ. In 2010, Bhatt et al. (13) published an ultra-performance liquid chromatography tandem mass spectrometry (UPLC–MS-MS) method in the biomedical chromatography to measure plasma OXC and MHD concentrations in humans, in which UPLC was evaluated as a faster and more efficient analytical tool compared with current HPLC (14). However, the method developed by Bhatt et al. still suffered from many negative factors, such as obvious matrix effects (MEs) and time-consuming pretreatment, which might prevent it from popular application to high-throughput analysis for biological samples. Based on the method of Bhatt et al., we improved the sample preparation and adjusted the testing conditions to develop a simple, rapid and sensitive UPLC–MS-MS method for determination of OXC and MHD in rat plasma. Further, the method was validated and applied to the evaluation for effects of KET and VOR on the pharmacokinetics of OXC and MHD in rats.
Experimental Reagents and materials OXC ( purity >98%), MHD ( purity >99%), diazepam (internal standard, IS, purity >99%, Figure 1) and carboxymethylcellulose sodium
salt (CMC) were purchased from Sigma-Aldrich Company (St. Louis, USA). KET and VOR were purchased from Melone Biotechnology Co., Ltd (Beijing, China). LC-grade acetonitrile was obtained from Amethyst Chemicals, and LC-grade formic acid was purchased from Sigma (St. Louis, MO, USA). All other chemicals were of HPLC or analytical grade. Ultra-pure water prepared by Milli-Q purification system from Millipore (Bedford, MA, USA) was used for the mobile phase and all other solutions.
Instrumentation UPLC–MS-MS with ACQUITY I-Class UPLC and XEVO TQD triplequadrupole mass spectrometer (Waters Corp., Milford, MA, USA) were used to analyze the compounds. The UPLC system was comprised of a binary solvent manager (BSM) and a sample manager with flow-through needle (SM-FTN). Data acquisition and control of the instrument were performed by Masslynx 4.1 software (Waters Corp., Milford, MA, USA).
Liquid chromatographic and mass spectrometric conditions OXC, MHD and IS were separated using a UPLC® BEH C18 column (2.1 × 50 mm, 1.7 μm; Waters, USA) kept at 40°C. The initial mobile phase consisted of acetonitrile and water (containing 0.1% formic acid) with gradient elution at a flow rate of 0.4 mL/min, and the sample volume was 2 μL. Elution was in a linear gradient, with solvent A (acetonitrile) and solvent B (0.1% formic acid in water) as follows: 0– 1 min (20–80% A), 1–1.5 min (80–80% A), 1.5–2 min (80–95% A), 2–2.5 min (95–95% A) and 2.5–2.6 min (95–20% A). After each
Ketoconazole and Voriconazole on the Pharmacokinetics of Oxcarbazepine and MHD in Rats injection, the sample manager underwent a needle wash process, including strong wash (methanol–water, 50/50, v/v) and weak wash (methanol–water, 10/90, v/v). Mass spectrometric detection was performed on a triple-quadrupole mass spectrometer equipped with an electrospray ionization (ESI) interface in a positive mode (Toronto, Canada). The quantitative analysis of OXC, MHD and IS in rat plasma was performed using the multiple reaction monitoring (MRM) method. Nitrogen was used as the desolvation gas (1,000 L/h) and cone gas (50 L/h). The selected ion monitoring conditions were set as follows: capillary voltage 4 kV; source temperature 150°C; and desolvation temperature 400°C. The cone voltage was set at 30 V for OXC, 20 V for MHD and 35 V for IS. The collision energy was set at 13 V for OXC, 8 V for MHD and 35 V for IS. Quantification was performed using MRM with the transitions of m/z 253.2→235.9, m/z 254.87→236.97, m/z 285.1→ 193.1 for OXC, MHD and IS, respectively.
Preparation of standard solutions and quality control samples The primary standard stock solutions of OXC (100 μg/mL), MHD (50 μg/mL) and IS (0.5 mg/mL) were, respectively, prepared by dissolving each standard substance in methanol. A series of working solutions of range from 10 to 2,000 ng/mL for OXC, from 5 to 1,000 ng/mL for MHD and IS solution at 50 ng/mL were prepared by diluting their stock solutions with methanol. Calibration standards and quality control (QC) samples in rat plasma were prepared by diluting the corresponding working solutions with blank plasma samples. The final concentrations were 10, 25, 50, 100, 250, 500, 1,000 and 2,000 ng/mL for OXC, and 5, 12.5, 25, 50, 125, 250, 500 and 1,000 ng/mL for MHD. QC samples were prepared independently in the same way at three concentrations: 10, 100 and 1,000 ng/mL for OXC, and 5, 250 and 500 ng/mL for MHD. The stock solutions and working solutions were stored at 4°C. The QC samples were dispatched in 100 μL aliquots and stored in Eppendorf tubes at −20°C until analysis. All solutions were kept in the refrigerator no more than 10 days. They were brought to room temperature and appropriately vortexed before use.
3
and interday precisions were separately calculated as relative standard deviation (RSD) for one day and day to day, and the accuracy was calculated as relative error (RE) as well. The ME was evaluated by comparing the responds of analytes dissolved in the reconstituted solution of the blank plasma extracts (E) with those dissolved in the mobile phase (M). The ME was calculated by the formula as follow: ME (%) = E/M × 100%. The recoveries at three QC levels of OXC and MHD were evaluated by comparing peak area ratios of QC samples obtained from precipitation of blank plasma spiked with a known concentration QC with from no precipitation at same concentration. Stability studies were performed by detecting OXC or MHD at three concentrations in six replicates under different storage conditions.
Pharmacokinetics Fifteen male Sprague–Dawley rats (200–250 g), purchased from Wenzhou Medical University Laboratory Animal Center, were divided randomly into the control group (group A) and study groups (group B, group C). Group A, control group: animals were treated with 0.5% CMC intragastrically (i.g.) and half an hour later orally administrated OXC at a dose of 125 mg/kg ; Group B, KET-treated group: animals were treated with 30 mg/kg KET i.g. and half an hour later orally administrated OXC at a dose of 125 mg/kg; Group C, VOR-treated group: animals were treated with 30 mg/kg VOR i.g. and half an hour later orally administrated OXC at a dose of 125 mg/kg. The rats were fasted for 12 h before administration but had free access to water. KET and VOR were dissolved in 0.5% CMC. Blood samples were collected via tail vein to 1.5-mL Eppendorf tube at the time point of 0.17, 0.33, 0.67, 1, 1.5, 2, 3, 4, 6, 8, 10, 12, 24, 36 and 72 h, and were immediately centrifugated at 3,000g for 10 min. The plasma samples were stored at −80°C until analysis. The main pharmacokinetic parameters were acquired by DAS software (Version 3.0, Shanghai BioGuide Medicinal Technology, China). The animal protocol was approved by Science and Technology Department of Zhejiang Province (Certificate No. 2203001).
Results Plasma sample pretreatment
Chromatogram
The frozen plasma samples were all thawed at room temperature and completely mixed by vortex. An aliquot 50 μL of plasma samples and 20 μL IS working solution (50 ng/mL) were added to a 1.5-mL Eppendorf tube. Protein precipitation was performed by 0.1 mL of acetonitrile followed by ∼2 min of vortex mixing. Afterward, the mixture was centrifuged for 10 min with 3,000g, and 2 μL supernatant was injected into the UPLC–MS-MS system for analysis. The calibration samples and QC samples were pretreated in the same way as the plasma samples.
Figure 2 shows the typical chromatograms of a blank plasma (A), OXC standard peak (100 μg/mL), MHD standard peak (50 μg/mL) and IS (diazepam; 10 μg/mL) (B), and a plasma sample obtained from a control group at 3 h after oral administration of OXC (125 mg/mL) (C). OXC, MHD and IS were eluted at 1.23 min, 1.11 min and 1.6 min, respectively, and a sample run could be completed within 3 min. The ME of all analytes ranged from 95.3% to 104.7% at three concentrations. No interfering endogenous substances were observed at the retention times of the analytes and IS. The results indicate that ion suppression or enhancement from the plasma matrix was negligible for this analytical method.
Method validation Specificity was evaluated by analysis of blank rat plasma, in which OXC, MHD and IS were not added in order to find possible interferences from backgrounds. The calibration curves were constructed by the peak area ratio (y) of each analyte to IS versus the analyte concentration (x), which was prepared with the weighted (1/x 2) least square linear regression method. The lower limit of quantification (LLOQ) was set according to the signal-to-noise ≥10, by trying a series concentration of analytes. QC samples at three concentrations, 10, 100 and 1,000 ng/mL for OXC, or 5, 250 and 500 ng/mL for MHD, were assayed in six replicates per day, consecutively for 3 days. The intraday
Linearity and the LLOQ The calibration curves were over the range of 10–2,000 ng/mL for OXC and 5–1,000 ng/mL for MHD. The regression equations for the calibration plot were y = 0.00724689*x + 0.00596567 with r 2 = 0.9997 for OXC and y = 0.0236884*x + 0.0398518 with r 2 = 0.9997 for MHD (Figure 3). The LLOQ in the calibration curves were 5 ng/ mL for OXC and 2.5 ng/mL for MHD, respectively.
4
Chen et al.
Figure 2. Representative UPLC–MS-MS chromatograms for OXC, MHD and IS. (A) Chromatograms for blank plasma, (B) chromatograms for standard peak of OXC (m/z 253.2 > 235.9, 100 ug/mL), MHD (m/z 254.86 > 236.97, 50 ug/mL) and IS (m/z 285.1 > 193.1, 10 ug/mL), (C) chromatograms for plasma sample obtained from a control group at 3 h after oral administration of OXC (125 mg/kg). OXC, oxcarbazepine; MHD, 10,11-dihydro-10-hydroxy-carbazepine; IS, internal standard, diazepam. This figure is available in black and white in print and in color at JCS online.
Ketoconazole and Voriconazole on the Pharmacokinetics of Oxcarbazepine and MHD in Rats
5
Figure 2. Continued
Precision, accuracy and recovery
Effects of VOR on pharmacokinetics of OXC and MHD
The assay performance data are presented in Table I. The precision of the method was determined by calculating RSD for QCs at three concentrations over three validation days. The intra- and interday precision is within the range of 1.31–7.26% and 1.49–7.98%. The recovery of OXC and MHD is 78.95–84.3% and 79.19–83.21%, respectively. Statistics satisfactorily meets the criteria for the analysis of biological samples according to the guidance of the US Food and Drug Administration (15).
The concentration vs. time profiles of MHD and OXC after coadministration with VOR are presented in Figures 4 and 5, and the main pharmacokinetic parameters are shown in Tables II and III. Co-administered with VOR, it could be observed that the AUC and Cmax of OXC were increased by 78.8% and 1.8-fold (P < 0.05) compared with the control group, whereas the CL and Vd were reduced by 50 and 52.8% (P < 0.05), respectively (Figure 4 and Table II). While the Cmax of MHD was decreased by 47.1% (P < 0.01), the AUC of MHD is increased, mildly by 21.7% with no statistical significance relative to the control group (P > 0.05), but by 1.1-fold with a significant difference to the KET-treated group (P < 0.01). The CL of MHD was reduced by 72.3%, with a 52% increase in MRT, which was more remarkable than the KET-treated group (P < 0.01). Besides all that, the Vd of MHD in the VOR-treated group was decreased by 78% compared with the control group, more significantly than 55.4% in the KET-treated group (P < 0.05, VOR-treated group vs. KET-treated group, Figure 5 and Table III). The results suggest that VOR influences the disposition of OXC and MHD in vivo, predominantly of MHD.
Effects of KET on pharmacokinetics of OXC and MHD The concentration vs. time profiles of OXC and MHD after co-administration with KET are presented in Figures 4 and 5, and the main pharmacokinetic parameters are shown in Tables II and III. Compared with the control group, the area under curve (AUC) of OXC in the KET-treated group was increased by 1.1-fold and the maximum concentration (Cmax) was increased by 1.5-fold, without change in peak time (Tmax). Accordingly, the clearance (CL) and apparent volume of distribution (Vd) of OXC were reduced by 48.6 and 48.7% respectively. Significant differences could also be observed in major pharmacokinetic parameters of MHD between the control group and the KET-treated group. Relative to the control group, the Cmax and AUC of MHD were decreased by 54.2 and 39.6% (P < 0.01), meanwhile, the CL and Vd were decreased by 49.6% (P < 0.01) and 55.4% (P < 0.05), respectively, with a corresponding increase in MRT of 44.8% (P < 0.05). The results demonstrate that KET impacts on the disposition of OXC and MHD in vivo.
Discussion At present, methods used in determinations of OXC mainly include HPLC–UV spectrometry or LC–APCI–MS mass-spectrometry (16– 18). According to the FDA guidance for industry bioanalytical method validation (15), the optimal detecting technique applied to pharmacokinetic studies should be sensitive, specific and fully validated. Bhatt et al. established the UPLC–MS-MS method for simultaneous determination of OXC and MHD in human plasma (13). Compared
6
Chen et al.
Figure 3. The curves of OXC and MHD. (A) The calibration curves of OXC, (B) the calibration curves of MHD. OXC, oxcarbazepine; MHD, 10,11-dihydro-10-hydroxycarbazepine; IS, internal standard, diazepam. This figure is available in black and white in print and in color at JCS online.
Table I. The Inter- and Intraday Precision, Accuracy and Recovery for OXC and MHD in Rat Plasma Compound
OXC
MHD
Spiked concentration (ng/mL)
Intraday
Interday
Recovery
Measured (ng/mL)
RSD (%)
RE (%)
Measured (ng/mL)
RSD (%)
RE (%)
Mean ± SD (%)
RSD (%)
10 100 1000 5 250 500
10.5 ± 0.8 101.3 ± 1.3 968.9 ± 53.9 5.0 ± 0.1 247.6 ± 11.3 488.9 ± 34.2
7.3 1.3 5.6 1.8 4.6 7.0
4.7 1.3 −3.1 0.7 −0.9 −2.2
10.5 ± 0.9 102.4 ± 1.5 973.9 ± 50.8 5.1 ± 0.1 243.7 ± 10.7 484.8 ± 36.9
8.0 1.5 5.2 2.1 4.3 7.6
5.3 2.4 −2.6 2.1 −0.1 −3.0
84.3 ± 4.6 79.0 ± 6.0 82.4 ± 7.4 83.2 ± 7.0 79.2 ± 4.6 80.3 ± 6.7
5.4 7.6 9.0 8.4 5.8 8.3
OXC, oxcarbazepine; MHD, 10,11-dihydro-10-hydroxy-carbazepine.
with the HPLC reported previously, UPLC–MS-MS detection is quicker, requires less mobile phase and provides better sensitivity. In our study, improvements were made in order to preferably meet the sensitivity and high-throughput required for studies in small animals. We used a simple sample preparation procedure, acetonitrile precipitation, instead of traditional liquid–liquid or solid-phase extraction, which made the whole process faster and allowed high extraction yields with good selectivity. Furthermore, we modulated the
proportion of the mobile phase and column temperature to increase the column efficiency, which made the method more sensitive and less of MEs. The LLOQ was improved, from 9.6 to 5 ng/mL for OXC, and from 19.4 to 2.5 ng/mL for MHD (13). Only 50 μL rat blood sample per time point from oral administration were needed for analysis because of the high sensitivity of the method. Therefore, the deviation from hypovolemia can be avoided, which might happen when small animal was used to derive a complete pharmacokinetic
7
Ketoconazole and Voriconazole on the Pharmacokinetics of Oxcarbazepine and MHD in Rats
Figure 4. Mean plasma concentration–time curve of OXC in control, KET- and VOR-treated rats (n = 5). A Group, control group, 5% CMC plus 125 mg/kg OXC; B Group, KET-treated group, 30 mg/kg KET plus 125 mg/kg OXC; C Group, VOR-treated group, 30 mg/kg VOR plus 125 mg/kg OXC. KET, ketoconazole; OXC, oxcarbazepine; VOR, voriconazole; CMC, carboxymethylcellulose sodium salt.
Figure 5. Mean plasma concentration–time curve of MHD in control, KET- and VOR-treated rats (n = 5). A Group, control group, 5% CMC plus 125 mg/kg OXC; B Group, KET-treated group, 30 mg/kg KET plus 125 mg/kg OXC; C Group, VORtreated group, 30 mg/kg VOR plus 125 mg/kg OXC. MHD, 10,11-dihydro-10hydroxy-carbazepine; OXC, oxcarbazepine; KET, ketoconazole; VOR, voriconazole; CMC, carboxymethylcellulose sodium salt.
profile. The method is convenient and sensitive; especially suitable for simultaneous determinations of OXC and MHD in rat plasma. We consider this method is also valuable for human clinical study, because the target therapeutic concentration of MHD is 3–40 μg/mL (19, 20), far above the LLOQ in this study. The pharmacokinetic studies revealed that KET obviously affected the pharmacokinetics of OXC and MHD in rats. The increased Cmax and AUC of OXC, decreased CL of OXC, along with the reduced Cmax and AUC of MHD meant that the conversion from OXC to MHD had been inhibited. Besides the inhibited production by KET, the CL of MHD was decreased and the MRT was concurrently increased, suggesting that the elimination of MHD was also inhibited. As we know, OXC undergoes reduction to form MHD in liver cells and the enzyme responsible for this step is cytosolic arylketone reductases, which is relatively stable and insusceptible to other drugs. MHD is eliminated by conjugation with glucuronic acid mediated by uridine diphospate-glucuronosyltransferase (UDPGT) in liver, which is also less susceptible to interference than oxidative enzymes (21). Concerning KET, it appeared that no assay existed for special and selective inhibiting effects on hepatic aryl ketone reductases or UDPGT. However, there were certain reports that oral administration of KET had a serious problem of hepatotoxicity, in which the increase of plasma alanine aminotransferase (ALT) was used as the evidence and reporting indicator (22, 23). We suspect that the inhibition on aryl ketone reductases and UDPGT is secondary to the damage of liver cells. The dose of KET in our studies, 30 mg/kg in rat, is approximately equivalent to 336 mg in person of 70 kg body weight, converted by body surface area. In China, KET is still prescribed for oral administration in some non-standard medical institutions and the recommended dose is 200 mg each, two times per day; the liver damage should not be ignored (24). Another remarkable phenomenon was that, the Vds of OXC and MHD were diminished by KET. This was unexpected because KET was generally regarded as inhibitor of P-glycoprotein (25), an efflux multidrug transporter which played an important role in the distribution of OXC and MHD in vivo and should have led to more accumulation of OXC and MHD in tissues (e.g., in brain). The findings remind that some cellular uptake mechanism for OXC and MHD is suppressed by KET. VOR is another member of the azole family of antifungal reagents available as an alternative to KET, with broad-spectrum antifungal effects and improvement in safety related to liver injury (26). According to the official statement of American Thoracic Society, its recommended oral dose is 200 mg each, two times per day (27). In our study, rats were orally administered 30 mg/kg VOR, equivalent to 200–400 mg in person. Results demonstrated a similar modification in OXC
Table II. The Main Pharmacokinetic Parameters of OXC After Co-administration with KET or VOR (n = 5) Parameters
Control group
KET-treated group
VOR-treated group
Cmax (µg/L) t1/2z (h) Tmax (h) Vz/F (L/kg) CLz/F (L/kg) AUC(0–∞) (µg h/L) MRT(0–∞) (h)
875.7 ± 74.5 4.0 ± 0.4 1.9 ± 0.2 81.3 ± 7.6 14.1 ± 1.1 86,438.0 ± 14,886.0 9.7 ± 1.1
2194.7 ± 117.7* 4.0 ± 0.4 2.0 ± 0.3 41.7 ± 4.9* 7.3 ± 0.46* 182,062.0 ± 31,891.0* 10.5 ± 1.0
2428.3 ± 167.3* 3.8 ± 0.6 2.1 ± 0.2 38.3 ± 6.1* 7.1 ± 0.3* 154,624.0 ± 17,929.0* 8.7 ± 0.6
OXC, oxcarbazepine; KET, ketoconazole; VOR, voriconazole; CMC, carboxymethylcellulose sodium salt. Control group, 0.5% CMC plus 125 mg/kg OXC; KET-treated group, 30 mg/kg KET plus 125 mg/kg OXC; VOR-treated group, 30 mg/kg VOR plus 125 mg/kg OXC. *P < 0.05 compared with the control group by the two-tailed t-test.
8
Chen et al.
Table III. The Main Pharmacokinetic Parameters of MHD after Co-administration with KET or VOR (n = 5) Parameters
Control group
KET-treated group
VOR-treated group
Cmax (µg/L) t1/2z (h) Tmax (h) Vz/F (L/kg) CLz/F (L/kg) AUC(0–∞) (µg h/L) MRT(0–∞) (h)
370.9 ± 39.0 13.6 ± 3.2 4.7 ± 2.0 817.4 ± 268.4 41.3 ± 8.5 33360.0 ± 9955.0 9.8 ± 1.7
169.8 ± 49.6** 11.8 ± 5.6 4.9 ± 1.7 417.8 ± 197.3* 20.829 ± 3.0** 20133.0 ± 2765.0** 14.1 ± 3.8*
196.0 ± 29.3** 11.1 ± 3.2 5.2 ± 1.1 179.2 ± 48.8*,▴ 11.5 ± 2.4*,▴▴ 40614.0 ± 11233.0▴▴ 14.8 ± 1.6**
MHD, 10,11-dihydro-10-hydroxy-carbazepine; OXC, oxcarbazepine; KET, ketoconazole; VOR, voriconazole; CMC, carboxymethylcellulose sodium salt. Control group, 0.5% CMC plus 125 mg/kg OXC; KET-treated group, 30 mg/kg KET plus 125 mg/kg OXC; VOR-treated group, 30 mg/kg VOR plus 125 mg/kg OXC. *P < 0.05 compared with the control group by the two-tailed t-test. **P < 0.01 compared with the control group by the two-tailed t-test. ▴ P < 0.05 compared with the KET-treated group by the two-tailed t-test. ▴▴ P < 0.01 compared with the KET-treated group by the two-tailed t-test.
pharmacokinetics with KET. However, there were significant differences in MHD pharmacokinetics between the VOR and KET-treated group. Compared with KET, pretreatment with VOR resulted in a conspicuous increase in AUC (P < 0.01), a sharp reduction in CL (P < 0.01) and a diminution in Vd of MHD (P < 0.05) as well. The results suggested that VOR predominantly interfered with disposition of MHD. Since both the production and metabolism of MHD are accordantly inhibited, the VOR-induced impairment in liver cells should be regarded. Another possible reason was the lessening of Vd, which resulted from the suppressed uptakes in tissues. Thus, the mechanism for VOR on some unaware carrier protein, especially on liver and kidney, is waiting for further experiments.
Conclusion A sensitive and convenient UPLC–MS-MS method for the simultaneous detection of OXC and MHD in rat plasma was developed. The results of method validation meet the criteria of industrial guidance for bioanalysis (15). The method was applied for the OXC pharmacokinetics study in rats, and the pharmacokinetic interaction with KET or VOR was evaluated. The preliminary results demonstrate significant modifications in pharmacokinetics of OXC and MHD when pretreated by KET or VOR. The method is suitable for small animal experiments because of quicker pretreatment and less sample volume. It is speculated to be also valuable for therapeutic drug monitoring of OXC in clinics for lower LLOQ than target concentration of MHD in humans.
Supplementary data Supplementary data are available at Journal of Chromatographic Science online.
Acknowledgments This work was supported by a grant from the National Key Project for New Drug Investigations (2012ZX09303008) founded by the Ministry of Science and Technology of the People’s Republic of China.
References 1. Wellington, K., Goa, K.L.; Oxcarbazepine: an update of its efficacy in the management of epilepsy; CNS Drugs, (2001); 15: 137–163.
2. Flesch, G., Czendlik, C., Renard, D., Lloyd, P.; Pharmacokinetics of the monohydroxy derivative of oxcarbazepine and its enantiomers after a single intravenous dose given as racemate compared with a single oral dose of oxcarbazepine; Drug Metabolism and Disposition, (2011); 39: 1103–1110. 3. Perucca, E.; Clinically relevant drug interactions with antiepileptic drugs; British Journal of Clinical Pharmacology, (2006); 61: 246–255. 4. Larkin, J.G., McKee, P.J., Forrest, G., Beastall, G.H., Park, B.K., Lowrie, J.I., et al.; Lack of enzyme induction with oxcarbazepine (600 mg daily) in healthy subjects; British Journal of Clinical Pharmacology, (1991); 31: 65–71. 5. Feliu, J., Del Pozo, J.L., Azanza, J.R., Garcia-Munoz, R., Zabalza, A., Gorosquieta, A., et al.; Antifungal prophylaxis with anidulafungin to minimize drug interactions with an antiepileptic treatment in a hematopoietic stem cell transplant recipient; Journal of Clinical Pharmacy and Therapeutics, (2015); doi: 10.1111/jcpt.12299. [Epub ahead of print]. 6. Sutherland, A., Ellis, D.; Treatment of a critically ill child with disseminated Candida glabrata with a recombinant human antibody specific for fungal heat shock protein 90 and liposomal amphotericin B, caspofungin, and voriconazole; Pediatric Critical Care Medicine, (2008); 9: e23–e25. 7. Madhugiri, V.S., Bhagavatula, I.D., Mahadevan, A., Siddaiah, N.; An unusual infection, an unusual outcome—Fonsecaea pedrosoi cerebral granuloma; Jouranl of Neurosurgery: Pediatrics, (2011); 8: 229–232. 8. Spina, E., Arena, D., Scordo, M.G., Fazio, A., Pisani, F., Perucca, E.; Elevation of plasma carbamazepine concentrations by ketoconazole in patients with epilepsy; Therapeutic Drug Monitoring, (1997); 19: 535–538. 9. Nivoix, Y., Ubeaud-Sequier, G., Engel, P., Leveque, D., Herbrecht, R.; Drug–drug interactions of triazole antifungal agents in multimorbid patients and implications for patient care; Current Drug Metabolism, (2009); 10: 395–409. 10. Surowiec, D., DePestel, D.D., Carver, P.L.; Concurrent administration of sirolimus and voriconazole: a pilot study assessing safety and approaches to appropriate management; Pharmacotherapy, (2008); 28: 719–729. 11. Yang, Z.H., Liu, X.D.; P-glycoprotein-mediated efflux of phenobarbital at the blood–brain barrier evidence from transport experiments in vitro; Epilepsy Research, (2008); 78: 40–49. 12. Flesch, G.; Overview of the clinical pharmacokinetics of oxcarbazepine; Clinical Drug Investigation, (2004); 24: 185–203. 13. Bhatt, M., Shah, S., Shivprakash; Rapid ultraperformance liquid chromatography–tandem mass spectrometry method for quantification of oxcarbazepine and its metabolite in human plasma; Biomedical Chromatography, (2011); 25: 751–759. 14. de Villiers, A., Lestremau, F., Szucs, R., Gelebart, S., David, F., Sandra, P.; Evaluation of ultra performance liquid chromatography. Part I. Possibilities and limitations; Journal of Chromatography A, (2006); 1127: 60–69. 15. Food and Drug Administration. Guidance for industry; bioanalytical method validation. Department of Health and Human Services, Rockville, USA (2001).
Ketoconazole and Voriconazole on the Pharmacokinetics of Oxcarbazepine and MHD in Rats 16. Kimiskidis, V., Spanakis, M., Niopas, I., Kazis, D., Gabrieli, C., Kanaze, F.I., et al.; Development and validation of a high performance liquid chromatographic method for the determination of oxcarbazepine and its main metabolites in human plasma and cerebrospinal fluid and its application to pharmacokinetic study; Journal of Pharmaceutical and Biomedical Analysis, (2007); 43: 763–768. 17. Juenke, J.M., Brown, P.I., Urry, F.M., McMillin, G.A.; Drug monitoring and toxicology: a procedure for the monitoring of oxcarbazepine metabolite by HPLC–UV; Journal of Chromatographic Science, (2006); 44: 45–48. 18. Klys, M., Rojek, S., Bolechala, F.; Determination of oxcarbazepine and its metabolites in postmortem blood and hair by means of liquid chromatography with mass detection (HPLC/APCI/MS); Journal of chromatography. B, Analytical technologies in the Biomedical and Life Sciences, (2005); 825: 38–46. 19. Johannessen, S.I., Battino, D., Berry, D.J., Bialer, M., Kramer, G., Tomson, T., et al.; Therapeutic drug monitoring of the newer antiepileptic drugs; Therapeutic Drug Monitoring, (2003); 25: 347–363. 20. Neels, H.M., Sierens, A.C., Naelaerts, K., Scharpe, S.L., Hatfield, G.M., Lambert, W.E.; Therapeutic drug monitoring of old and newer antiepileptic drugs; Clinical Chemistry and Laboratory Medicine, (2004); 42: 1228–1255.
9
21. Bock, K.W., Bock-Hennig, B.S.; Differential induction of human liver UDP-glucuronosyltransferase activities by phenobarbital-type inducers; Biochemical Pharmacology, (1987); 36: 4137–4143. 22. Rodriguez, R.J., Buckholz, C.J.; Hepatotoxicity of ketoconazole in SpragueDawley rats: glutathione depletion, flavin-containing monooxygenasesmediated bioactivation and hepatic covalent binding; Xenobiotica, (2003); 33: 429–441. 23. Findor, J.A., Sorda, J.A., Igartua, E.B., Avagnina, A.; Ketoconazole-induced liver damage; Medicina (B Aires), (1998); 58: 277–281. 24. He, Y., Chen, X.M.; Analysis of drug risk management deficiencies from current status of using ketoconazole oral formulations; Chinese Journal of Pharmacovigilance, (2014); 11: 85–87. 25. Siegsmund, M.J., Cardarelli, C., Aksentijevich, I., Sugimoto, Y., Pastan, I., Gottesman, M.M.; Ketoconazole effectively reverses multidrug resistance in highly resistant KB cells; Journal of Urology, (1994); 151: 485–491. 26. Mikus, G., Scholz, I.M., Weiss, J.; Pharmacogenomics of the triazole antifungal agent voriconazole; Pharmacogenomics, (2011); 12: 861–872. 27. Limper, A.H., Knox, K.S., Sarosi, G.A., Ampel, N.M., Bennett, J.E., Catanzaro, A., et al.; An official American Thoracic Society statement: treatment of fungal infections in adult pulmonary and critical care patients; American Journal of Respiratory and Critical Care Medicine, (2011); 183: 96–128.
