HILIC LC-MS for the determination of 2'-C-methyl-cytidine-triphosphate in rat liver.
A very accurate and selective LC-MS/MS method was developed and validated for the quantification of 2'-C-modified nucleoside triphosphate in liver tissue samples. An efficient pretreatment procedure of liver tissue samples was developed, using a fully automated SPE procedure with 96-well SPE plate (weak anion exchange sorbent, 30 mg). Nucleotide hydrophilic interaction chromatography has been performed on an aminopropyl column (100 mm x 2.0 mm, 3 microm) using a gradient mixture of ACN and ACN/water (5:95 v/v) with 20 mM ammonium acetate at pH 9.45 as mobile phase at 300 microL/min flow rate. The 2'-C-modified nucleoside triphosphate was detected in the negative ESI mode in multiple reaction monitoring (MRM) mode. Calibration curve was linear over the 0.05-50 microM concentration range. Satisfying results, confirming the high reliability of the established LC-MS/MS method, were obtained for intraday precision (CV = 2.5-9.1%) and accuracy (92.6-94.8%) and interday precision (CV = 9.6-11.5%) and accuracy (94.4-102.4%) as well as for recovery (82.0-112.6%) and selectivity. The method has been successfully applied for pharmacokinetic studies of 2'-C-methyl-cytidine-triphosphate in liver tissue samples.
Department of Drug Metabolism and Pharmaceuticals, Merck & Co., Inc., Boston, MA 02115, USA. vincenzo_pucci@merck.com
Vincenzo
V
Pucci
Claudio
C
Giuliano
Rena
R
Zhang
Kenneth Alan
KA
Koeplinger
Joseph F
JF
Leone
Edith
E
Monteagudo
Fabio
F
Bonelli
D000818:Animals; D000998:Antiviral Agents / Q000008:administration & dosage / Q000378:metabolism; D002138:Calibration; D002853:Chromatography, Liquid / Q000379:methods; D003570:Cytidine Triphosphate / Q000031:analogs & derivatives* / Q000032:analysis*; D008099:Liver / Q000737:chemistry*; D008297:Male; D011355:Prodrugs / Q000008:administration & dosage / Q000378:metabolism; D011741:Pyrimidine Nucleosides / Q000008:administration & dosage / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D052616:Solid Phase Extraction / Q000379:methods; D012997:Solvents / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*
D000998:Antiviral Agents; D011355:Prodrugs; D011741:Pyrimidine Nucleosides; D012997:Solvents; D003570:Cytidine Triphosphate; C506207:valopicitabine