Quantitative analysis of HIV-1 protease inhibitors in cell lysates using MALDI-FTICR mass spectrometry.
In this report we explore the use of MALDI-FTICR mass spectrometry for the quantitative analysis of five HIV-1 protease inhibitors in cell lysates. 2,5-Dihydroxybenzoic acid (DHB) was used as the matrix. From a quantitative perspective, DHB is usually a poor matrix due to its poor shot-to-shot and poor spot-to-spot reproducibilities. We found that the quantitative precisions improved significantly when DMSO (dimethylsulfoxide) was added to the matrix solution. For lopinavir and ritonavir, currently the most frequently prescribed HIV-1 protease inhibitors, the signal-to-noise ratios improved significantly when potassium iodide was added to the matrix solution. The mean quantitative precisions, expressed as % relative standard deviation, were 6.4% for saquinavir, 7.3% for lopinavir, 8.5% for ritonavir, 11.1% for indinavir, and 7.2% for nelfinavir. The mean quantitative accuracies, expressed as % deviation, were 4.5% for saquinavir, 6.0% for lopinavir, 5.9% for ritonavir, 6.6% for indinavir, and 8.0% for nelfinavir. The concentrations measured for the individual quality control samples were all within 85-117% of the theoretical concentrations. The lower limits of quantification in cell lysates were 4 fmol/microL for saquinavir, 16 fmol/microL for lopinavir, 31 fmol/microL for ritonavir, and 100 fmol/microL for indinavir and nelfinavir. The mean mass accuracies for the protease inhibitors were 0.28 ppm using external calibration. Our results show that MALDI-FTICR mass spectrometry can be successfully used for precise, accurate, and selective quantitative analyses of HIV-1 protease inhibitors in cell lysates. In addition, the lower limits of quantification obtained allow clinical applications of the technique.
van Kampen
Jeroen J A
JJ
Department of Neurology, Laboratory of Neuro-Oncology and Clinical and Cancer Proteomics, Erasmus MC, Rotterdam, The Netherlands.
Burgers
Peter C
PC
de Groot
Ronald
R
Osterhaus
Albert D M E
AD
Reedijk
Mariska L
ML
Verschuren
Esther J
EJ
Gruters
Rob A
RA
Luider
Theo M
TM
D002478:Cells, Cultured; D017320:HIV Protease Inhibitors / Q000032:analysis*; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*
D017320:HIV Protease Inhibitors