Research article Received: 25 December 2013,

Revised: 7 April 2014,

Accepted: 26 April 2014

Published online in Wiley Online Library

(wileyonlinelibrary.com) DOI 10.1002/bmc.3257

UPLC-MS/MS determination of voriconazole in human plasma and its application to a pharmacokinetic study Zhe Wang, Cheng-ke Huang, Wei Sun, Cui Xiao and Zeng-shou Wang* ABSTRACT: A sensitive and rapid ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed to determine voriconazole in human plasma. Sample preparation was accomplished through a simple one-step protein precipitation with methanol. Chromatographic separation was carried out on an Acquity UPLC BEH C18 column using an isocratic mobile phase system composed of acetonitrile and water containing 1% formic acid (45:55, v/v) at a flow rate of 0.50 mL/min. Mass spectrometric analysis was performed using a QTrap5500 mass spectrometer coupled with an electrospray ionization source in the positive ion mode. The multiple reaction monitoring transitions of m/z 351.0 → 281.5 and m/z 237.1 → 194.2 were used to quantify voriconazole and carbamazepine (internal standard), respectively. The linearity of this method was found to be within the concentration range of 2.0–1000 ng/mL with a lower limit of quantification of 2.0 ng/mL. Only 1.0 min was needed for an analytical run. This fully validated method was successfully applied to the pharmacokinetic study after oral administration of 200 mg voriconazole to 20 Chinese healthy male volunteers. Copyright © 2014 John Wiley & Sons, Ltd. Keywords: voriconazole; UPLC-MS/MS; human plasma; pharmacokinetic

Introduction

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Experimental Chemicals materials Voriconazole and carbamazepine (internal standard, IS) were obtained from Sigma (St Louis, MO, USA). LC-grade acetonitrile and methanol were purchased from the Beijing Chemical Reagents Company (Beijing, China). HPLC-grade water was obtained using a Milli Q system (Millipore, Bedford, USA). Blank human plasma used in this study was supplied by The Second Affiliated Hospital of Wenzhou Medical University (Wenzhou, China).

UPLC-MS/MS conditions Liquid chromatography was performed on an Acquity ultraperformance liquid chromatography (UPLC) unit (Waters Corp., Milford, MA, USA) with an Acquity BEH C18 column (2.1 × 50 mm, 1.7 μm particle size) and inline

* Correspondence to: Z.-S. Wang, Department of Pharmacy, The Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou 325027, China. E-mail: [email protected] Department of Pharmacy, The Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, Wenzhou 325027, China Abbreviations used: MRM, multiple reaction monitoring.

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1

Voriconazole, a triazole antifungal agent, was approved for the treatment of invasive fungal infection with a broad spectrum, including Aspergillus Cryptococcus and Candida species. It is used in the treatment of invasive pulmonary aspergillosis and respiratory disorders as an antifungal agent (Steinbach and Stevens, 2003; Walsh et al., 2008). However, a high incidence of adverse reactions, such as liver dysfunction, visual disturbance and neurological toxicity, may occur during the treatment with voriconazole. (Dolton et al., 2012; Luong et al., 2012; Somchit et al., 2012; Tan et al., 2006; Ueda et al., 2009). Owing to the above findings, (Dodds Ashley et al., 2007) have suggested that voriconazole blood concentration should be maintained between 1.0 and 5.5 μg/mL and the measurement of blood levels could assist with decisions about dose adjustment (Pascual et al., 2008). Therefore, therapeutic drug monitoring of voriconazole in plasma is very important (Chu et al., 2013; Hamada et al., 2013; Michael et al., 2010). A number of methods for the quantification of voriconazole in biological fluids have been described. HPLC with UV detection is most widely used (Cendejas-Bueno et al., 2013; Ekiert et al., 2010; Heng et al., 2013; Simmel et al., 2008). For HPLC methods, a rather long chromatographic run time (up to 12 min) is often needed. LC-MS/MS methods permit shorter runs. However, the published analysis times in the literature vary between 3 and 13 min (Alffenaar et al., 2010; Cheng et al., 2011; Farowski et al., 2010; Lin et al., 2013; Xiong et al., 2013). To our knowledge, only one method with a run time of 1.5 min has been described (Xiong et al., 2010). This method, however, necessitates a time-consuming solvent evaporation step. Hence, it is necessary to develop a simple and sensitive method to monitor voriconazole for obtaining optimum therapeutic concentrations in human blood.

This paper describes a fast, selective and highly sensitive approach to the determination of voriconazole at 2.0 ng/mL in plasma with good accuracy using ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). The analysis time was only 1.0 min and the sample preparation comprised only one simple step of protein precipitation. The method was fully validated and applied to the pharmacokinetic study of voriconazole in Chinese healthy volunteers.

Z. Wang et al. 0.2 μm stainless steel frit filter (Waters Corp., Milford, USA). The mobile phase consisted of acetonitrile and water containing 1% formic acid (45:55, v/v). The flow rate was 0.40 mL/min. The overall run time was 1.0 min. An AB Sciex QTRAP 5500 triple quadruple mass spectrometer equipped with an electrospray ionization source (Toronto, Canada) was used for mass spectrometric detection. The detection was operated in the multiple reaction monitoring (MRM) mode under unit mass resolution (0.7 amu) in the mass analyzers. The dwell time was set to 200 ms for each MRM transition. The MRM transitions were m/z 351.0 → 281.5 and m/z 237.1 → 194.2 for voriconazole and IS, respectively (Fig. 1). After optimization, the source parameters were set as follows: curtain gas, 30 psig; nebulizer gas, 55 psig; turbo gas, 60 psig; ion spray voltage, 3.0 kV; and temperature, 400°C. Data acquisition and processing were performed using analyst software (version 1.5, AB Sciex).

Standard solutions, calibration standards and quality control sample The stock solution of voriconazole that was used to make the calibration standards and quality control (QC) samples was prepared by dissolving 10 mg in 10 mL methanol to obtain a concentration of 1.0 mg/mL. The stock solution was further diluted with methanol to obtain working solutions at several concentration levels. Calibration standards and QC samples in plasma were prepared by diluting the corresponding working solutions with blank human plasma. Final concentrations of the calibration standards were 2.0, 5.0, 10, 25, 50, 100, 250, 500 and 1000 ng/mL for voriconazole in human plasma. The concentrations of QC samples in plasma were 4.0, 100 and 800 ng/mL for voriconazole. IS stock solution was made at an initial concentration of 1.0 mg/mL. The IS working solution (100.0 ng/mL) was made from the stock solution using methanol for dilution. All stock solutions, working solutions, calibration standards and QCs were immediately stored at 4°C.

Sample preparation Before analysis, the plasma sample was thawed to room temperature. In a 1.5 mL centrifuge tube, an aliquot of 200 μL of the IS working solution (100.0 ng/mL in methanol) was added to 100 μL of collected plasma sample. The tubes were vortex-mixed for 1.0 min and spun in a centrifuge at 13,000 rmp for 10 min. The supernatant (10 μL) was injected into the UPLC-MS/MS system for analysis.

Method validation Before using this method to determinate voriconazole in human plasma, the method was fully validated for specificity, linearity, precision, accuracy, recovery, matrix effect and stability according the United States Food and Drug Administration bioanalytical method validation guidances (Jamalapuram et al., 2013). Specificity was determined by analysis of blank human plasma samples from six different volunteers. Every blank sample was handled by the procedure described in ‘Sample preparation’ and confirmed that endogenous substances did not interfere with the analyte and the IS. To evaluate the linearity, calibration standards of nine concentrations of voriconazole (2.0–1000 ng/mL) were separately extracted and assayed on three separate days. The linearity for voriconazole was investigated 2 by weighted (1/x ) least-squares linear regression of peak area ratios against concentrations. The sensitivity of the method was determined by quantifying the lower limit of quantification (LLOQ). The LLOQ was defined as the lowest point in the calibration curve determined at an acceptable precision and accuracy. The precision and accuracy of the method were assessed by determination of QC samples in plasma at different concentrations (4.0, 100, 800 ng/ mL) on three separate days. Precision was expressed as percentage relative standard deviation (RSD) and accuracy was expressed as percentage relative error (RE) between the measured and nominal value. The precision for QC samples was within 15%, and accuracy between 15 and 15%. Extraction recovery experiments which showed an ability to extract the analyte from the test biological samples were evaluated by comparing the peak areas obtained from extracted QC samples with nonprocessed standard solutions at three concentrations at the same concentration. Recovery of IS was determined at the working concentration (100.0 ng/mL) similarly. To determine the matrix effect, six different blank human samples were utilized to prepare QC samples and used for assessing the lot-tolot matrix effect. Matrix effect was evaluated at three QC levels by comparing the peak areas of analytes obtained from plasma samples spiked with analytes after extraction to those of the pure standard solutions at the same concentrations. The matrix effect of IS was evaluated at the working concentration (100.0 ng/mL) in the same manner. To ensure the reliability of the results, stability assay comprising freeze–thaw, short-term and long-term stability were carried out. In the protocol, the short-term stability was determined after the exposure of the spiked samples at room temperature for 2 h, and the ready-to-inject samples (after extraction) in the autosampler at 4°C for 12 h. The freeze– thaw stability was evaluated after three complete freeze–thaw cycles ( 20 to 25°C) on consecutive days. The long-term stability was assessed after storage of the standard spiked plasma samples at 20°C for 60 days. Samples were considered to be stable if assay values were within the acceptable limits of accuracy (RE % ≤ ± 15%) and precision (RSD % ≤ 15%).

Application to a pharmacokinetic study

2 Figure 1. The chemical structures and daughter scan ion spectra of voriconazole and IS in the present study: (A) voriconazole; (B) carbamazepine (IS).

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The present method was applied to a pharmacokinetic study after an oral administration of 200 mg voriconazole to Chinese male volunteers. The clinical protocol was approved by Tonji Medical College Huazhong University of Science and Technology prior to the study. Twenty volunteers gave written informed consent to participate in the study according to the principles of the Declaration of Helsinki. The volunteers who submitted the agreements to attend this project were medically examined for the pharmacokinetics study of voriconazole. The subjects were required to abstain from taking any other drug for 7 days prior to the start of the test. They were also required not to smoke or drink alcohol for 24 h before the beginning of the study until its end. All volunteers received an oral dose of 200 mg voriconazole with 200 mL water. Blood samples (3 mL) were collected into heparinized tubes at 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 12 and 24 h after oral administration. Blood samples were centrifuged at 4000g for 10 min and the plasma was separated and kept frozen at 20°C until analysis.

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UPLC-MS/MS determination of voriconazole Data analysis

Results and discussion

Plasma concentration vs time profiles were analyzed using DAS software (version 2.0, Medical University of Wenzhou, China) to estimate the type of compartment model and pharmacokinetic parameters. Data are expressed as means ± SD.

Method development and optimization Liquid–liquid extraction and solid-phase extraction are often used to prepare biological samples owing to their ability to

Figure 2. Representative chromatograms of voriconazole and IS in human plasma samples. (A) A blank plasma sample; (B) a blank plasma sample spiked with voriconazole and IS; (C) a plasma sample from a person 1.0 h after an oral administration 200 mg.

Table 1. Precision and accuracy of method for the determination of voriconazole in human plasma (n = 6) Analytes

Concentration added(ng/mL)

Voriconazole

4.0 100.0 800.0

Intra-day precision

Inter-day precision

Mean ± SD

RSD (%)

RE (%)

4.13 ± 0.36 100.17 ± 8.70 819.83 ± 50.15

8.74 8.69 6.12

3.33 0.17 2.48

Mean ± SD

RSD (%)

RE (%)

9.47 8.87 6.24

1.25 3.17 2.65

4.05 ± 0.38 103.17 ± 9.15 778.83 ± 48.59

Table 2. Recovery and matrix effect of voriconazole and internal standards (n = 6) Analytes

IS

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4.0 100.0 800.0 100.0

Recovery (%)

Matrix effect (%)

Mean ± SD

RSD (%)

Mean ± SD

RSD (%)

88.83 ± 4.36 85.67 ± 3.56 84.11 ± 2.37 84.56 ± 5.48

4.90 4.15 2.82 6.48

101.67 ± 5.68 98.83 ± 5.64 97.83 ± 3.66 97.77 ± 3.65

5.59 5.70 3.74 3.73

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Voriconazole

Concentration added (ng/mL)

4

Recovery and matrix effect The recovery was calculated by comparing the mean peak areas of the analyte spiked before extraction divided by the areas of

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10.98 6.57 3.28 3.93 ± 0.43 90.67 ± 5.96 801.33 ± 26.25 3.33 2.33 0.77 8.19 9.70 3.02 4.13 ± 0.34 97.67 ± 9.48 793.83 ± 24.01 0.83 5.83 0.79 9.39 8.97 2.56 3.97 ± 0.37 94.17 ± 8.45 793.67 ± 20.31 2.92 9.50 0.90 9.13 6.08 2.94

RSD (%) Mean ± SD RSD (%) Mean ± SD Mean ± SD

4°C, 12 h

RE (%) RSD (%) Mean ± SD

3.88 ± 0.35 90.50 ± 5.50 807.17 ± 23.73 4.0 100.0 800.0

The intra- and inter-day precision and accuracy of the method were determined from the analysis of QC samples at three different concentrations for each biological matrix. The results are summarized in Table 1. The method was reliable and reproducible since the RSD was 80%. The protein precipitation sample preparation procedure was much simpler and less expensive than the liquid–liquid extraction method previously used in the literature (Verdier et al., 2010). Thus, this method met the requirements of high sample throughput in bioanalysis. Analyte ionization is affected by the composition of the mobile phase. Ammonium acetate and formic acid were used to augment the ionic strength. Formic acid was better than ammonium acetate at improving the response of voriconazole. The effect of formic acid (0.05, 0.1 and 0.2% in the aqueous phase) on the response to voriconazole was investigated, and 0.1% formic acid was found to be the optimum concentration. Acetonitrile was better than methanol at improving the shapes of the voriconazole peaks. A mixture of water with 0.1% formic acid– acetonitrile was finally adopted as the mobile phase.

1.67 9.33 0.17

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UPLC-MS/MS determination of voriconazole analytes samples spiked after extraction and multiplied by 100%. Results are shown in Table 2. The recovery in plasma ranged from 84.11 to 88.83% for voriconazole. The recovery of IS (100.0 ng/mL) in plasma was 84.56%. The matrix effects in human plasma were all between 97.83 to 101.67% for voriconazole at different QC levels (Table 2). The matrix effect for IS (100.0 ng/mL) was 97.77%. No apparent matrix effect was found to affect the determination of voriconazole and IS in plasma. As a result, the matrix effect from plasma was negligible in this method.

Stability Stability tests were performed at the low, medium and high QC samples with five determinations for each storage condition. The RSDs of the mean test responses were within 15% in all stability tests. Table 3 shows the stability data for voriconazole in plasma under different storage and temperature conditions. There was no effect on the quantitation for plasma samples kept at room temperature for 2 h and at 4°C for 12 h. No significant degradation was observed when samples of voriconazole were taken through three freeze ( 20°C) and thaw (25°C) cycles. As a result, voriconazole in samples were stable at 20°C for 60 days.

Application of the method in a pharmacokinetic study The method described above was successfully applied to determine the concentration of voriconazole in human plasma. After an oral administration of 200 mg voriconazole tablets, the main pharmacokinetic parameters of voriconazole for 20 volunteers were estimated. The plasma samples with analyte concentrations above the upper limit of quantitation were diluted with blank human plasma. The mean plasma concentration–time curve of voriconazole is displayed in Fig. 3, and the main pharmacokinetic parameters of voriconazole were calculated and are summarized in Table 4. Compared with recently published papers describing the pharmacokinetic profiles of voriconazole in healthy volunteers, the pharmacokinetic parameters of voriconazole obtained in the study were generally similar (Lin et al., 2013).

Conclusions A UPLC-MS/MS method for the determination of voriconazole in human plasma was developed and validated. To the best of our knowledge, this is the first report of the determination of voriconazole level in human plasma using an UPLC-MS/MS method. The method affords the sensitivity, accuracy and precision needed for quantitative measurements of voriconazole in human plasma. It was also successfully applied in a pharmacokinetic study.

References

Figure 3. Plasma concentration vs time curves of voriconazole after a single oral administration 200 mg to 20 Chinese healthy male volunteers.

Table 4. Pharmacokinetic parameters of voriconazole after oral administration 200 mg to 20 Chinese healthy male volunteers. Parameter

5.88 ± 0.93 1037.01 ± 81.18 1.70 ± 0.41 6643.92 ± 696.70 7004.10 ± 794.82

t1/2, half-life; Cmax, maximum plasma; Tmax, Peak time; AUC, the area under the plasma concentration-time curve.

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t1/2 (h) Cmax (ng/mL) Tmax (h) AUC0→24 (ng/mL h) AUC0→∞ (ng/mL h)

Voriconazole

Alffenaar JW, Wessels AM, van Hateren K, Greijdanus B, Kosterink JG and Uges DR. Method for therapeutic drug monitoring of azole antifungal drugs in human serum using LC/MS/MS. Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences 2010; 878: 39–44. Cendejas-Bueno E, Rodriguez-Tudela JL, Cuenca-Estrella M and GomezLopez A. Development and validation of a fast HPLC/photodiode array detection method for the measurement of voriconazole in human serum samples. A reference laboratory experience. Enfermedades Infecciosas y Microbiología Clínica 2013; 31: 23–28. Cheng Y, Zhang ZJ, Tian Y, Li WJ and Wei W. Quantification of voriconazole in human plasma by high-performance liquid chromatography–electrospray ionization mass spectrometry: application to a bioequivalence study. Arzneimittel-Forschung 2011; 61: 132–139. Chu HY, Jain R, Xie H, Pottinger P and Fredricks DN. Voriconazole therapeutic drug monitoring: retrospective cohort study of the relationship to clinical outcomes and adverse events. BMC Infectious Diseases 2013; 13: 105. Dodds Ashley ES, Zaas AK, Fang AF, Damle B and Perfect JR. Comparative pharmacokinetics of voriconazole administered orally as either crushed or whole tablets. Antimicrobial Agents 2007; 51: 877–880. Dolton MJ, Ray JE, Chen SC, Ng K, Pont LG and McLachlan AJ. Multicenter study of voriconazole pharmacokinetics and therapeutic drug monitoring. Antimicrobial Agents and Chemotherapy 2012; 56: 4793–4799. Ekiert RJ, Krzek J and Talik P. Chromatographic and electrophoretic techniques used in the analysis of triazole antifungal agents – a review. Talanta 2010; 82: 1090–1100. Farowski F, Cornely OA, Vehreschild JJ, Hartmann P, Bauer T, Steinbach A, Ruping MJ and Muller C. Quantitation of azoles and echinocandins in compartments of peripheral blood by liquid chromatography–tandem mass spectrometry. Antimicrobial Agents and Chemotherapy 2010; 54: 1815–1819. Hamada Y, Tokimatsu I, Mikamo H, Kimura M, Seki M, Takakura S, Ohmagari N, Takahashi Y, Kasahara K, Matsumoto K, Okada K, Igarashi M, Kobayashi M, Mochizuki T, Nishi Y, Tanigawara Y, Kimura T and Takesue Y. Practice guidelines for therapeutic drug monitoring of voriconazole: a consensus review of the Japanese Society of Chemotherapy and the Japanese Society of Therapeutic Drug Monitoring. Journal of Infection Chemotherapy 2013; 19: 381–392.

Z. Wang et al. Heng SC, Nation RL, Levvey B, Snell GI, Slavin MA and Kong DCM. Quantification of voriconazole in human bronchoalveolar lavage fluid using high-performance liquid chromatography with fluorescence detection. Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences 2013; 913: 171–175. Jamalapuram S, Vuppala PK, Abdelazeem AH, McCurdy CR and Avery BA. Ultra-performance liquid chromatography tandem mass spectrometry method for the determination of AZ66, a sigma receptor ligand, in rat plasma and its application to in vivo pharmacokinetics. Biomedical Chromatography 2013; 27: 1034–1040. Lin D, Li G and Chen L. Determination of voriconazole in human plasma by HPLC-ESI-MS and application to pharmacokinetic study. Journal of Chromatographic Science 2013; 51: 485–489. Luong ML, Hosseini-Moghaddam SM, Singer LG, Chaparro C, Azad S, Lazar N, Boutros PC, Keshavjee S, Rotstein C and Husain S. Risk factors for voriconazole hepatotoxicity at 12 weeks in lung transplant recipients. American Journal of Transplantation 2012; 12: 1929–1935. Michael C, Bierbach U, Frenzel K, Lange T, Basara N, Niederwieser D, Mauz-Korholz C and Preiss R. Determination of saliva trough levels for monitoring voriconazole therapy in immunocompromised children and adults. Therapeutic Drug Monitoring 2010; 32: 194–199. Pascual A, Calandra T, Bolay S, Buclin T, Bille J and Marchetti O. Voriconazole therapeutic drug monitoring in patients with invasive mycoses improves efficacy and safety outcomes. Clinical Infectious Diseases 2008; 46: 201–211. Pauwels S, Vermeersch P, Van Eldere J and Desmet K. Fast and simple LC-MS/MS method for quantifying plasma voriconazole. Clinica Chimica Acta 2012; 413: 740–743. Simmel F, Soukup J, Zoerner A, Radke J and Kloft C. Development and validation of an efficient HPLC method for quantification of voriconazole in plasma and microdialysate reflecting an important target site. Analytical and Bioanalytical Chemistry 2008; 392: 479–488.

Somchit N, Chung JH, Yaacob A, Ahmad Z, Zakaria ZA and Kadir AA. Lack of hepato- and nephrotoxicity induced by antifungal drug voriconazole in laboratory rats. Drug and Chemical Toxicology 2012; 35: 304–309. Steinbach WJ and Stevens DA. Review of newer antifungal and immunomodulatory strategies for invasive aspergillosis. Clinical Infectious Diseases 2003; 37(suppl. 3): S157–187. Tan K, Brayshaw N, Tomaszweski K, Troke P and Wood N. Investigation of the potential relationships between plasma voriconazole concentrations and visual adverse events or liver function test abnormalities. Journal of Clinical Pharmacology 2006; 46: 235–243. Ueda K, Nannya Y, Kumano K, Hangaishi A, Takahashi T, Imai Y and Kurokawa M. Monitoring trough concentration of voriconazole is important to ensure successful antifungal therapy and to avoid hepatic damage in patients with hematological disorders. International Journal of Hematology 2009; 89: 592–599. Verdier MC, Bentue-Ferrer D, Tribut O and Bellissant E. Liquid chromatography–tandem mass spectrometry method for simultaneous quantification of four triazole antifungal agents in human plasma. Clinical Chemistry and Laboratory Medicine 2010; 48: 1515–1522. Walsh TJ, Anaissie EJ, Denning DW, Herbrecht R, Kontoyiannis DP, Marr KA, Morrison VA, Segal BH, Steinbach WJ, Stevens DA, van Burik JA, Wingard JR, Patterson TF and Infectious Diseases Society of America. Treatment of aspergillosis: clinical practice guidelines of the Infectious Diseases Society of America. Clinical Infectious Diseases 2008; 46: 327–360. Xiong X, Duan J, Zhai S, Wang L and Lan X. Fast and reliable determination of voriconazole in human plasma by LC-APCI-MS/MS. Bioscience, Biotechnology, and Biochemistry 2010; 74: 2151–2153. Xiong X, Zhai S and Duan J. Validation of a fast and reliable liquid chromatography–tandem mass spectrometry (LC-MS/MS) with atmospheric pressure chemical ionization method for simultaneous quantitation of voriconazole, itraconazole and its active metabolite hydroxyitraconazole in human plasma. Clinical Chemistry and Laboratory Medicine 2013; 51: 339–346.

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Biomed. Chromatogr. 2014

MS determination of voriconazole in human plasma and its application to a pharmacokinetic study.

A sensitive and rapid ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed to determine voriconazole in ...
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