title: Simultaneous determination of lamivudine, lopinavir, ritonavir, and zidovudine concentration in plasma of HIV-infected patients by HPLC-MS/MS.
abstract: The nucleoside reverse transcriptase inhibitors lamivudine and zidovudine and the protease inhibitors lopinavir and ritonavir are currently used in anti-human immunodeficiency virus (HIV) therapy. Here, a high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) method, using a hybrid quadrupole time-of-flight mass analyzer, is reported for the simultaneous quantification of lamivudine, lopinavir, ritonavir, and zidovudine in plasma of HIV-infected patients. The volume of plasma sample was 600 μL. Plasma samples were extracted by solid-phase using 1 cc Oasis HLB Cartridge (divinylbenzene and N-vinylpyrrolidone) and evaporated in a water bath under nitrogen stream. The extracted samples were reconstituted with 100-μL methanol. Five microliters of the reconstituted samples were injected into a HPLC-MS/MS apparatus, and the analytes were eluted on a Vydac column (250 × 1.0 mm i.d.) filled with 3-μm C(18) particles. The mobile phase was delivered at 70 μL/min with a linear gradient elution, both acetonitrile and ultrapure water solvents contained 0.2% formic acid. The calibration curves were linear from 0.47 to 20 ng/mL. The absolute recovery ranged between 91 and 107%. The minimal concentration of lamivudine, lopinavir, ritonavir, and zidovudine detectable by HPLC-MS/MS is 0.47, 0.28, 0.30, and 0.66 ng/mL, respectively. The great advantage of the new HPLC-MS/MS method here reported is the possibility to achieve a very high specificity toward the selected anti-HIV drugs, despite the simple and rapid sample preparation. Moreover, this method is easily extendible to the analysis of co-administrated drugs.
authors.affiliation: Istituto Nazionale per le Malattie Infettive I.R.C.C.S. Lazzaro Spallanzani, Roma, Italy.
authors.forename: Stefania
authors.initials: S
authors.lastname: Notari
authors.forename: Manuel
authors.initials: M
authors.lastname: Sergi
authors.forename: Camilla
authors.initials: C
authors.lastname: Montesano
authors.forename: Jelena
authors.initials: J
authors.lastname: Ivanovic
authors.forename: Pasquale
authors.initials: P
authors.lastname: Narciso
authors.forename: Leopoldo P
authors.initials: LP
authors.lastname: Pucillo
authors.forename: Paolo
authors.initials: P
authors.lastname: Ascenzi
mesh_terms: D019380:Anti-HIV Agents / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000592:standards; D015658:HIV Infections / Q000097:blood / Q000188:drug therapy*; D006801:Humans; D019259:Lamivudine / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D057230:Limit of Detection; D061466:Lopinavir / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D012015:Reference Standards; D015203:Reproducibility of Results; D019438:Ritonavir / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use; D053719:Tandem Mass Spectrometry / Q000592:standards; D015215:Zidovudine / Q000097:blood* / Q000302:isolation & purification / Q000627:therapeutic use
chemical_list: D019380:Anti-HIV Agents; D061466:Lopinavir; D019259:Lamivudine; D015215:Zidovudine; D019438:Ritonavir