Simultaneous determination of thymine and its sequential catabolites dihydrothymine and beta-ureidoisobutyrate in human plasma and urine using liquid chromatography-tandem mass spectrometry with pharmacokinetic application

Ni, Ming, Duley, John, George, Rani, Charles, Bruce, Shannon, Catherine, McGeary, Ross and Norris, Ross (2013) Simultaneous determination of thymine and its sequential catabolites dihydrothymine and beta-ureidoisobutyrate in human plasma and urine using liquid chromatography-tandem mass spectrometry with pharmacokinetic application. Journal of Pharmaceutical and Biomedical Analysis, 78-79 129-135. doi:10.1016/j.jpba.2013.01.038

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Author Ni, Ming
Duley, John
George, Rani
Charles, Bruce
Shannon, Catherine
McGeary, Ross
Norris, Ross
Title Simultaneous determination of thymine and its sequential catabolites dihydrothymine and beta-ureidoisobutyrate in human plasma and urine using liquid chromatography-tandem mass spectrometry with pharmacokinetic application
Formatted title
Simultaneous determination of thymine and its sequential catabolites dihydrothymine and β-ureidoisobutyrate in human plasma and urine using liquid chromatography-tandem mass spectrometry with pharmacokinetic application
Journal name Journal of Pharmaceutical and Biomedical Analysis   Check publisher's open access policy
ISSN 0731-7085
1873-264X
Publication date 2013-05-01
Sub-type Article (original research)
DOI 10.1016/j.jpba.2013.01.038
Open Access Status Not Open Access
Volume 78-79
Start page 129
End page 135
Total pages 7
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Language eng
Formatted abstract
To study in vivo activities of dihydropyrimidine dehydrogenase, dihydropyrimidinase, and ureidoproprionase, a sensitive, accurate, selective and precise method for the determination of the endogenous pyrimidine thymine (THY) and its successive metabolites dihydrothymine (DHT) and β-ureidoisobutyric (UIB) acid in human plasma and urine has been developed and validated. Plasma or diluted urine (200μL) was mixed with 1mL of deuterated-THY (internal standard) in acetonitrile, then centrifuged, the supernatant evaporated, and the residue reconstituted in 150μL 0.1% (w/w) formic acid in water. Separation was performed on a Waters Symmetry® C8 column (150mm×3.9mm; 5μm particle size), with gradient elution using a mobile phase of 0.1% (w/w) formic acid in water (phase A), and 15% (v/v) methanol in acetonitrile (phase B). The detection system was an Applied Biosystems model 3200 tandem mass spectrometer with atmospheric pressure chemical ionisation, and multiple reaction monitoring mode using the transitions: THY (m/z: 127.1-110.0), DHT (m/z: 129.1-68.9), UIB (m/z: 147.1-86.0), and deuterated-THY (m/z: 131.1-114.0). THY, DHT, and UIB eluted at 5.12min, 5.17min and 5.00min, respectively. Linearity of the calibrations was established from 2 to 500μg/L. The lower limit of quantification was 5μg/L in plasma, and 10μg/L in urine for THY, DHT and UIB. Ion-suppression had negligible effect. A pilot pharmacokinetic study analysed plasmas and urines from 2 healthy male subjects who each received an oral 250mg THY dose. THY was rapidly absorbed and eliminated with an apparent biphasic log-linear profile. DHT and UIB demonstrated apparent formation-rate limited kinetics.
Keyword Thymine
Dihydrothymine
Ureidoisobutyric acid
LC-MS/MS
Pharmacokinetics
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

 
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