title: Lack of specificity for the analysis of raltegravir using online sample clean-up liquid chromatography-electrospray tandem mass spectrometry.
abstract: BACKGROUND
  Raltegravir is the first antiretroviral agent to target the human immunodeficiency virus-1 (HIV-1) integrase. It is indicated, in association with other antiretrovirals, in the treatment of acquired immunodeficiency syndrome (AIDS) in antiretroviral treatment-experienced adult patients with viral resistance. To evaluate the feasibility of raltegravir therapeutic drug monitoring, we developed a rapid and specific analytical method for the quantification of raltegravir in human plasma by online sample clean-up liquid chromatography-tandem mass spectrometry (LC-MS/MS).
  
  
  METHODS
  After protein precipitation (with 100 microL of acetonitrile/methanol (50/50)) of 25 microL of plasma, fast online matrix-clean-up was performed using a column switching program. The chromatographic step was optimized to separate raltegravir and its glucuronide metabolite (G-raltegravir). Multiple reaction monitoring (MRM) was used for detection of raltegravir and G-raltegravir. In the absence of G-raltegravir standard, G-raltegravir identification was confirmed by beta-glucuronidase pre-treatment.
  
  
  RESULTS
  A total analysis of 3.8 min was needed to separate raltegravir to G-raltegravir. The method was linear between 10 and 3000 ng/mL for raltegravir. Analytical recovery was 94+/-1%. Variation coefficients ranged between 5% and 8.4%. Pre-treatment of plasma from a patient under raltegravir treatment with beta-glucuronidase suppressed G-raltegravir peak.
  
  
  CONCLUSIONS
  We describe a fast online LC-MS/MS assay that is valid and reliable for the quantification of raltegravir, despite the lack of specificity that could occur in MRM scanning mode experiments.
authors.affiliation: CHU, Hôpital A. Michallon, Laboratoire de pharmacologie, BP217, Grenoble, F-38043, France.
authors.forename: Jean François
authors.initials: JF
authors.lastname: Jourdil
authors.forename: Mireille
authors.initials: M
authors.lastname: Bartoli
authors.forename: Françoise
authors.initials: F
authors.lastname: Stanke-Labesque
mesh_terms: D019380:Anti-HIV Agents / Q000097:blood*; D002853:Chromatography, Liquid / Q000379:methods*; D011760:Pyrrolidinones / Q000097:blood*; D000068898:Raltegravir Potassium; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D019380:Anti-HIV Agents; D011760:Pyrrolidinones; D000068898:Raltegravir Potassium