An improved on-line solid phase extraction coupled HPLC-MS/MS system for quantification of sifuvirtide in human plasma.
An improved liquid chromatographic method with on-line solid phase extraction (SPE) and tandem mass spectrometric detection was optimised for quantification of the anti-HIV peptide Sifuvirtide in human plasma. The SPE sorbents, loading buffer composition and other aspects of the on-line SPE column were investigated in detail for efficiently extracting the interesting peptides and simultaneously discarding the large amount of proteins. The gradient elution program was optimised on the analysis column to decrease the matrix effect and obtain excellent selectivity. The multiple charge ion at m/z 946.4 of Sifuvirtide was quantified by a linear ion trap mass spectrometer, operating in the positive mode, and selective reaction monitoring (SRM) acquisition. Method validation results demonstrated that the linear calibration curve covered a range of 6.1-6250 ng/mL, and the correlation coefficients (r(2)) were above 0.992. The lower limit of detection (LLOD) with a signal-to-noise (S/N) ratio higher than 10 was 6.1 ng/mL. The accuracy ranged from -7.6 to 10.6%, and the intra- and inter-batch precisions were less than 8.7% and 5.5%, respectively. Finally, more than nine hundred of samples from a clinical trial was completely analyzed using this on-line SPE coupled HPLC-MS/MS system in one single week, due to the rapid run-time of individual sample (6.5 min).
Wang
Qing-Qing
QQ
Laboratory of Drug Metabolism and Pharmacokinetics, Beijing Institute of Radiation Medicine, Beijing 100850, China.
Xiang
Shen-Si
SS
Jia
Yan-Bo
YB
Ou
Lun
L
Chen
Fang
F
Song
Hai-Feng
HF
Liang
Qing
Q
Ju
Dan
D
D000097:Acetonitriles / Q000737:chemistry; D019380:Anti-HIV Agents / Q000097:blood / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D005561:Formates / Q000737:chemistry; D006801:Humans; D016014:Linear Models; D010455:Peptides / Q000097:blood / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
D000097:Acetonitriles; D019380:Anti-HIV Agents; D005561:Formates; D010455:Peptides; C030544:formic acid; C505768:sifuvirtide; C032159:acetonitrile