Quantification of N-acetyl-seryl-aspartyl-lysyl-proline in hemodialysis patients administered angiotensin-converting enzyme inhibitors by stable isotope dilution liquid chromatography–tandem mass spectrometry

Abstract

We developed a sensitive, selective and accurate method based on liquid chromatography with tandem mass spectrometry (LC–MS/MS) to determine N-terminal thymosin-β peptides of Ac-SDKP and Ac-ADKP in human plasma samples. Quantification of Ac-SDKP and Ac-ADKP was performed using solid phase extraction (SPE) based on C18, reversed phase LC separation, and stable isotope dilution electrospray ionization-MS/MS in multiple reaction-monitoring (MRM) mode. The Ac-SDKP-13C6, 15N2 and Ac-ADKP-d7 were synthesized for the internal standards. These MRM monitoring ions were m/z 488→129 (quantitative ion)/226 for Ac-SDKP, m/z 496→137 for Ac-SDKP-13C6, 15N2, m/z 472→129 (quantitative ion)/226 for Ac-ADKP, and m/z 479→129 for Ac-ADKP-d7, respectively. Lower limit of quantitation (LLOQ) of Ac-SDKP and Ac-ADKP was 0.1ng/mL in human plasma. Recovery values were ranged from 94.7% to 106.3% for inter- (RSD: 0.6–3.5%) and intra- (RSD: 0.4–4.9%) day assays. Plasma Ac-SDKP levels were significantly higher in hemodialyzed subjects treated with angiotensin-converting enzyme inhibitors of enalapril (27.3±24.6ng/mL, n=10) and trandolapril (12.3±16.9ng/mL, n=18) than healthy (0.4±0.2ng/mL, n=7) and hemodialyzed subjects (0.6±0.2ng/mL, n=34). This analytical method would be useful to measure N-terminal thymosin-β peptides in human plasma for the clinical study.