Development, validation and application of a novel liquid chromatography tandem mass spectrometry assay measuring uracil, 5,6-dihydrouracil, 5-fluorouracil, 5,6-dihydro-5-fluorouracil, α-fluoro-β-ureidopropionic acid and α-fluoro-β-alanine in human plasma
| Autor: | Christopher M. Florkowski, Peter M. George, Ottiniel Chavani, Berit Packert Jensen, R. Matthew Strother |
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| Rok vydání: | 2017 |
| Předmět: |
Electrospray
Clinical Biochemistry Pharmaceutical Science Tandem mass spectrometry 030226 pharmacology & pharmacy 01 natural sciences Analytical Chemistry 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Tandem Mass Spectrometry Liquid chromatography–mass spectrometry Drug Discovery Dihydropyrimidine dehydrogenase medicine Humans Urea Uracil Chromatography High Pressure Liquid Spectroscopy Chromatography medicine.diagnostic_test 010401 analytical chemistry Extraction (chemistry) Dihydrouracil 0104 chemical sciences chemistry Therapeutic drug monitoring beta-Alanine |
| Zdroj: | Journal of Pharmaceutical and Biomedical Analysis. 142:125-135 |
| ISSN: | 0731-7085 |
| DOI: | 10.1016/j.jpba.2017.04.055 |
| Popis: | The plasma 5,6-dihydrouracil/uracil (UH2/U) ratio is a possible phenotypic marker of dihydropyrimidine dehydrogenase (DPD) activity, hence an index of 5-fluorouracil (5-FU) response and toxicity. Studies have re-affirmed the value of 5-FU and 5,6-dihydro-5-fluorouracil (FUH2) for therapeutic drug monitoring (TDM). However, FUH2 has limited stability in plasma, necessitating expedited plasma separation and freezing, where routine compliance may not be easy. The metabolites α-fluoro-β-ureidopropionic acid (FUPA) and α-fluoro-β-alanine (FβAL) are stable in plasma and are probable candidates for TDM. This paper describes development, validation and application of an LC-MS/MS assay quantifying U, UH2, 5-FU, FUH2, FUPA and FβAL in human plasma. Extraction was by salt-assisted liquid-liquid extraction (LLE) in two-stages with pH adjustment. The supernatants were mixed, dried and reconstituted. Analytes were resolved on the Luna PFP (2) (150×2.00mm, 3μ) column by gradient elution and analyzed by tandem mass spectrometry via electrospray ionisation in positive polarity. The analytical response was linear (r2≥0.99) in the concentration (ng/mL) ranges: 50-10 000 for FβAL and FUH2, 50-5 000 for FUPA, 50-100 000 for 5-FU, 5-200 for U and 10-400 for UH2. Within- and between-run accuracy and precision were ≤ 10.2% and ≤ 9.8% respectively across the QC range and inclusive of LLOQ. The internal standard (IS) normalised matrix effects were within 93-112% with CV of ≤ 9.7% and normalised recoveries were within 91-107% with CV of ≤ 9.8%. This robust assay was successfully applied to samples from rectal and colorectal cancer patients (n=10) on 5-FU. Deviations ≤ 2.0% from the mean values were observed when incurred samples were reanalysed. |
| Databáze: | OpenAIRE |
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