title: Validation of a hydrophilic interaction ultra-performance liquid chromatography-tandem mass spectrometry method for the determination of gemcitabine in human plasma with tetrahydrouridine.
abstract: A simple and reproducible bioanalytical method for the determination of gemcitabine in human plasma treated with tetrahydrouridine (THU) was developed and validated using a hydrophilic interaction ultra-performance liquid chromatography with tandem mass spectrometry (UPLC-MS/MS). To prevent deamination of gemcitabine, blood was treated with THU, and the plasma samples obtained after centrifugation were used in this study. Gemcitabine and gemcitabine-(13)C, (15)N2 used as an internal standard, were extracted from human plasma treated with THU using a 96-well Hybrid SPE-Precipitation plate. Extracts were chromatographed on a hydrophilic interaction chromatography column with isocratic elution. Detection was performed using Quattro Premier with positive electrospray ionization multiple reaction monitoring mode. The standard curve ranged from 10 to 10,000 ng/mL without carryover. No significant interferences were detected in blank plasma and no interferences by 2'-2'-difluoro-2'-deoxyuridine, a metabolite of gemcitabine. Accuracy and precision in the intra-batch reproducibility study using quality control samples with three THU levels did not exceed ±5.4 and 7.3%, respectively, and the inter-batch reproducibility results also met the criteria. Stability of gemcitabine was ensured in whole blood and plasma as well as stability of THU in solutions. The UPLC-MS/MS method developed was successfully validated and can be applied for gemcitabine bioanalysis in clinical studies.
authors: Drug Metabolism and Pharmacokinetics, Biopharmaceutical Assessment Core Function Unit, Eisai Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Yuji
authors: Y
authors: Mano
authors: Analysis group, Tsukuba division, Sunplanet Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Kenji
authors: K
authors: Sakamaki
authors: Analysis group, Tsukuba division, Sunplanet Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Takuya
authors: T
authors: Ueno
authors: Analysis group, Tsukuba division, Sunplanet Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Kenji
authors: K
authors: Kita
authors: Analysis group, Tsukuba division, Sunplanet Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Takuho
authors: T
authors: Ishii
authors: Drug Metabolism and Pharmacokinetics, Biopharmaceutical Assessment Core Function Unit, Eisai Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Koichiro
authors: K
authors: Hotta
authors: Drug Metabolism and Pharmacokinetics, Biopharmaceutical Assessment Core Function Unit, Eisai Co. Ltd, 1-3, 5-chome, Tokodai, Tsukuba-shi, Ibaraki, 300-2635, Japan.
authors: Kazutomi
authors: K
authors: Kusano
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000097:blood / Q000737:chemistry; D006801:Humans; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013767:Tetrahydrouridine / Q000737:chemistry*; D000093542:Gemcitabine
chemical_list: D003841:Deoxycytidine; D013767:Tetrahydrouridine; D000093542:Gemcitabine
keywords: UPLC-MS/MS; gemcitabine; human; tetrahydrouridine; validation