• Home
  • Search Results
  • Simultaneous determination of seven azole antifungal drugs in serum by ultra-high pressure liquid chromatography and diode array detection.

Simultaneous determination of seven azole antifungal drugs in serum by ultra-high pressure liquid chromatography and diode array detection.

Acta clinica Belgica (2014-03-19)
V Mistretta, N Dubois, R Denooz, C Charlier
ABSTRACT

Azole antifungals are a group of fungistatic agents that can be administered orally or parenterally. The determination of the concentrations of these antifungals (miconazole, fluconazole, ketoconazole, posaconazole, voriconazole, itraconazole, and its major active metabolite, hydroxy-itraconazole) in serum can be useful to adapt the doses to pharmacological ranges because of large variability in the absorption and metabolism of the drugs, multiple drug interactions, but also potential resistance or toxicity. A method was developed and validated for the simultaneous determination of these drugs in serum utilizing ultra-high pressure liquid chromatography and diode array detection (UHPLC-DAD). After a simple and rapid liquid-liquid extraction, the pre-treated sample was analysed on an UHPLC-DAD system (Waters Corporation(®)). The chromatographic separation was carried out on an Acquity BEH C18 column (Waters Corporation) with a gradient mode of mobile phase composed of acetonitrile and aqueous ammonium bicarbonate 10·0 M pH10. The flow rate was 0·4 ml/min and the injection volume was 5 μl. The identification wavelength varied according to the drug from 210 to 260 nm. The method was validated by the total error method approach by using an analytical validation software (e•noval V3·0 Arlenda(®)). The seven azole antifungals were identified by retention time and specific UV spectra, over a 13-minute run time. All calibration curves showed good linearity (r(2)>0·99) in ranges considered clinically adequate. The assay was linear from 0·05 to 10 mg/l for voriconazole, posaconazole, itraconazole, hydroxy-itraconazole, and ketoconazole, from 0·3 to 10 mg/l for fluconazole, and from 0·1 to 10 mg/l for miconazole. The bias and imprecision values for intra- and inter-assays were lower than 10% and than 15%, respectively. In conclusion, a simple, sensitive, and selective UHPLC-DAD method was developed and validated to determine seven azole antifungal drugs in human serum. This method is applicable to patient samples, and can be applied successfully to clinical applications and therapeutic drug monitoring.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Tetrahydrofuran, anhydrous, ≥99.9%, inhibitor-free
Sigma-Aldrich
Dichloromethane, anhydrous, ≥99.8%, contains 40-150 ppm amylene as stabilizer
Sigma-Aldrich
Tetrahydrofuran, anhydrous, contains 250 ppm BHT as inhibitor, ≥99.9%
Sigma-Aldrich
Tetrahydrofuran, inhibitor-free, suitable for HPLC, ≥99.9%
Sigma-Aldrich
Dichloromethane, suitable for HPLC, ≥99.8%, contains amylene as stabilizer
Sigma-Aldrich
Dichloromethane, contains 40-150 ppm amylene as stabilizer, ACS reagent, ≥99.5%
Sigma-Aldrich
Tetrahydrofuran, contains 250 ppm BHT as inhibitor, ACS reagent, ≥99.0%
Sigma-Aldrich
Sodium carbonate, ACS reagent, anhydrous, ≥99.5%, powder or granules
Sigma-Aldrich
Ammonium bicarbonate, ReagentPlus®, ≥99.0%
Sigma-Aldrich
Dichloromethane, HPLC Plus, for HPLC, GC, and residue analysis, ≥99.9%, contains 50-150 ppm amylene as stabilizer
Sigma-Aldrich
Pyrrole, reagent grade, 98%
Sigma-Aldrich
Ammonium bicarbonate, BioUltra, ≥99.5% (T)
Sigma-Aldrich
Sodium carbonate, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium carbonate, powder, ≥99.5%, ACS reagent
Sigma-Aldrich
Dichloromethane, puriss. p.a., ACS reagent, reag. ISO, ≥99.9% (GC)
Sigma-Aldrich
Tetrahydrofuran, contains 250 ppm BHT as inhibitor, puriss. p.a., ACS reagent, reag. Ph. Eur., ≥99.9%
Sigma-Aldrich
Sodium carbonate, ReagentPlus®, ≥99.5%
Sigma-Aldrich
Sodium carbonate, ACS reagent (primary standard), anhydrous, 99.95-100.05% dry basis
Sigma-Aldrich
1-Pentanol, ReagentPlus®, ≥99%
Sigma-Aldrich
1-Pentanol, ACS reagent, ≥99%
Sigma-Aldrich
Tetrahydrofuran, suitable for HPLC, ≥99.9%, inhibitor-free
Supelco
Sodium carbonate concentrate, 0.1 M Na2CO3 in water, eluent concentrate for IC
Sigma-Aldrich
Fluconazole, ≥98% (HPLC), powder
Supelco
Methylene Chloride, Pharmaceutical Secondary Standard; Certified Reference Material
Sigma-Aldrich
Dichloromethane, suitable for HPLC, ≥99.9%, contains 40-150 ppm amylene as stabilizer
Supelco
Sodium carbonate concentrate, Na2CO3 72 mM in water, IC eluent concentrate (20x) for Metrosep A Supp 7
Sigma-Aldrich
Sodium carbonate, anhydrous, powder, 99.999% trace metals basis
Sigma-Aldrich
Tetrahydrofuran, ReagentPlus®, ≥99.0%, contains 250 ppm BHT as inhibitor
Supelco
Fluconazole, Pharmaceutical Secondary Standard; Certified Reference Material
Supelco
Dichloromethane, analytical standard