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| Content Provider | World Health Organization (WHO)-Global Index Medicus |
|---|---|
| Author | Lin, Weisheng Huang, Mike-Qingtao Xue, Xiaohua Bertelsen, Kirk Chen, Guiyan Zhao, Harry Lin, Zhongping John Fourie, Anne de Jong, Jan Weng, Naidong |
| Description | Country affiliation: United States Author Affiliation: Lin W ( Frontage Laboratories Inc., 700 Pennsylvania Drive, Exton, PA 19341, USA.) |
| Abstract | Leukotriene B4 (LTB4) is an important inflammatory component in a number of diseases and has been used as a pharmacodynamic (PD) biomarker. In this report, a highly sensitive and selective ultra fast liquid chromatography-tandem mass spectrometry (UFLC-MS/MS) method for the determination of LTB4 in plasma from ex vivo stimulated human blood, using leukotriene B4-d4 (LTB4-d4, contains four deuterium atoms at the 6, 7, 14, and 15 positions) as the internal standard (IS), was developed and validated. The chromatographic separation of LTB4 from its three isomers and an unknown interference peak from human plasma was crucial to achieve accurate determination of 0.2 ng/mL (LLOQ) of LTB4. LTB4 and the IS were extracted with methyl tertiary butyl ether (MTBE) from 200 µL human plasma. Reversed-phase HPLC separation was carried out with a Phenomenex Synergi Hydro-RP column (100mm×3mm, 2.5 µm). MS/MS detection was set at mass transitions of 335.0â194.9 m/z for LTB4 and 339.0â196.9 m/z for LTB4-d4 in Turbo Ionization Spray (TIS) negative mode. The dynamic range of the method is 0.2-200 ng/mL. LTB4 was found to be stable in human plasma for at least three freeze (-20 °C)/thaw cycles, and on the benchtop (room temperature) for at least 6h. The stock solution storage stability study demonstrated that the LTB4 stock solution, in 50:50 acetonitrile:water, was stable at 4 °C for at least 198 days. The processed samples were found to be stable for at least 72 h at room temperature. The long-term sample storage stability test demonstrated that LTB4 human plasma samples were stable at a storage temperature of -20 °C for at least 198 days. In addition, intraday and interday accuracy and precision, sensitivity, linearity, and recovery were evaluated. An additional partial validation was conducted to decrease the plasma sample volume from 200 to 100 µL. All the data reported in this study fulfilled the requirements and recommendations in the FDA guidance for bioanalytical method validation. Comparison of the validated UFLC-MS/MS method with an ELISA method using ex vivo stimulated samples indicated that although results from the two assays correlated relatively well, the UFLC-MS/MS method has been shown to be superior in selectivity and dynamic range to an ELISA method in our study. The validated UFLC-MS/MS method was successfully used to analyze samples generated from two clinical studies. The excellent assay performance and incurred sample reproducibility (ISR) results obtained from the study sample analysis demonstrated the assay is robust and reliable. |
| File Format | HTM / HTML |
| ISSN | 15700232 |
| Volume Number | 925 |
| e-ISSN | 1873376X |
| Journal | Journal of Chromatography B |
| Language | English |
| Publisher | Elsevier |
| Publisher Date | 2013-04-15 |
| Publisher Place | Netherlands |
| Access Restriction | One Nation One Subscription (ONOS) |
| Subject Keyword | Discipline Analytical Chemistry Chromatography, High Pressure Liquid Methods Leukotriene B4 Blood Tandem Mass Spectrometry Chromatography, Reverse-phase Humans Chemistry Linear Models Reproducibility Of Results Sensitivity And Specificity Journal Article |
| Content Type | Text |
| Resource Type | Article |
| Subject | Cell Biology Medicine Analytical Chemistry Clinical Biochemistry Biochemistry |
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