Dataset Information

Quantitation of azoles and echinocandins in compartments of peripheral blood by liquid chromatography-tandem mass spectrometry.

ABSTRACT: A rapid turnaround is a prerequisite of therapeutic drug monitoring (TDM). For antifungals, this need is still unmet, since hardly any method has been established to simultaneously quantitate concentrations of different antifungal classes. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed allowing quantitation of anidulafungin (ANF), caspofungin (CSF), isavuconazole (ISC), micafungin (MCF), posaconazole (PSC), and voriconazole (VRC). Quantitation was successful with diluted plasma samples, peripheral blood mononuclear cells (PBMC), polymorphonuclear leukocytes (PMN), and erythrocytes (RBC). A triple quadrupole mass spectrometer in selected reaction monitoring mode was used with positive electrospray ionization. Cells and calibration standards were extracted with acetonitrile containing internal standard. Internal standards were a CSF derivate for echinocandins and itraconazole for triazoles. Chromatographic separation of the supernatant was achieved by a gradient method facilitating a BetaBasic C4 column. Analytes were quantified in a single 8-min run. Calibration curves were linear and fitted using least squares with a weighting factor of the reciprocal concentration. Limits of detection (ng/ml) were ANF, 8.3; CSF, 31.5; ISC, 1.5; MCF, 97.7; PSC, 3.3; and VRC, 1.4. The lower limits of quantitation (ng/ml) were ANF, 64; CSF, 108; ISC, 4.5; MCF, 160; PSC, 10; and VRC, 4.2. Intraday precisions ranged from 6.3% to 8.8% for azoles and 8.8% to 15.4% for echinocandins. Intraday and interday accuracies (percent bias) of all analytes were within 13.8%. The method was established as standard practice for the quantitation of intracellular antifungal concentrations and optimizes TDM by applying a rapid single method for 6 antifungals.

SUBMITTER: Farowski F

PROVIDER: S-EPMC2863689 | biostudies-literature | 2010 May

REPOSITORIES: biostudies-literature

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Quantitation of azoles and echinocandins in compartments of peripheral blood by liquid chromatography-tandem mass spectrometry.

Farowski Fedja F Cornely Oliver A OA Vehreschild Jörg J JJ Hartmann Pia P Bauer Tim T Steinbach Angela A Rüping Maria J G T MJ Müller Carsten C

Antimicrobial agents and chemotherapy 20100222 5

A rapid turnaround is a prerequisite of therapeutic drug monitoring (TDM). For antifungals, this need is still unmet, since hardly any method has been established to simultaneously quantitate concentrations of different antifungal classes. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed allowing quantitation of anidulafungin (ANF), caspofungin (CSF), isavuconazole (ISC), micafungin (MCF), posaconazole (PSC), and voriconazole (VRC). Quantitation was successful wit ...[more]

PMID: 20176892

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Project description:The consumption of wheat, rye, and barley may cause adverse reactions to wheat such as celiac disease, non-celiac gluten/wheat sensitivity, or wheat allergy. The storage proteins (gluten) are known as major triggers, but also other functional protein groups such as α-amylase/trypsin-inhibitors or enzymes are possibly harmful for people suffering of adverse reactions to wheat. Gluten is widely used as a collective term for the complex protein mixture of wheat, rye or barley and can be subdivided into the following gluten protein types (GPTs): α-gliadins, γ-gliadins, ω5-gliadins, ω1,2-gliadins, high- and low-molecular-weight glutenin subunits of wheat, ω-secalins, high-molecular-weight secalins, γ-75k-secalins and γ-40k-secalins of rye, and C-hordeins, γ-hordeins, B-hordeins, and D-hordeins of barley. GPTs isolated from the flours are useful as reference materials for clinical studies, diagnostics or in food analyses and to elucidate disease mechanisms. A combined strategy of protein separation according to solubility followed by preparative reversed-phase high-performance liquid chromatography was employed to purify the GPTs according to hydrophobicity. Due to the heterogeneity of gluten proteins and their partly polymeric nature, it is a challenge to obtain highly purified GPTs with only one protein group. Therefore, it is essential to characterize and identify the proteins and their proportions in each GPT. In this study, the complexity of gluten from wheat, rye, and barley was demonstrated by identification of the individual proteins employing an undirected proteomics strategy involving liquid chromatography-tandem mass spectrometry of tryptic and chymotryptic hydrolysates of the GPTs. Different protein groups were obtained and the relative composition of the GPTs was revealed. Multiple reaction monitoring liquid chromatography-tandem mass spectrometry was used for the relative quantitation of the most abundant gluten proteins. These analyses also allowed the identification of known wheat allergens and celiac disease-active peptides. Combined with functional assays, these findings may shed light on the mechanisms of gluten/wheat-related disorders and may be useful to characterize reference materials for analytical or diagnostic assays more precisely.

Dataset Information

Quantitation of azoles and echinocandins in compartments of peripheral blood by liquid chromatography-tandem mass spectrometry.

Publications

Quantitation of azoles and echinocandins in compartments of peripheral blood by liquid chromatography-tandem mass spectrometry.

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