Simultaneous determination of 16 anti-HIV drugs in human plasma by high-performance liquid chromatography

Stefania Notari, Alessio Bocedi, Giuseppe Ippolito, Pasquale Narciso, Leopoldo Paolo Pucillo, Gianna Tossini, Raffaele Perrone Donnorso, Francesco Gasparrini, Paolo Ascenzi

Research output: Contribution to journalArticle

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Abstract

Therapeutic drug monitoring (TDM) is pivotal to improve the management of HIV infection. Here, a HPLC-UV method has been developed to quantify simultaneously seven HIV protease inhibitors (amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir; PIs), seven nucleoside reverse transcriptase inhibitors (abacavir, didanosine, emtricitabine, lamivudine, stavudine, zalcitabine, and zidovudine; NRTIs), and two non-nucleoside reverse transcriptase inhibitors (efavirenz and nevirapine; NNRTIs) in human plasma. The volume of the plasma sample was 600 μL. This method involved automated solid-phase extraction with Oasis HLB Cartridge 1 cc (divinylbenzene and N-vinylpyrrolidone) and evaporation in a water bath under nitrogen stream. The extracted samples were reconstituted with 100 μL methanol. Twenty microliters of these samples were injected into a HPLC-UV system, the analytes were eluted on an analytical C18 Symmetry™ column (250 mm × 4.6 mm I.D.) with a particle size of 5 μm. The mobile phase (0.01 M KH2PO4 and acetonitrile) was delivered at 1.0 mL/min with linear gradient elution. The total run time for a single analysis was 35 min, the anti-HIV drugs were detected by UV at 240 and 260 nm. The calibration curves were linear up to 10 μg/mL. The absolute recovery ranged between 88 and 120%. The in vitro stability of anti-HIV drugs (0.005-10 μg/mL) in plasma has been studied at 24.0°C. On these bases, a two to four analyte method has been tailored to the individual needs of the HIV-infected patient. The HPLC-UV method here reported has been validated and is currently applied to monitor PIs, NRTIs, and NNRTIs in plasma of HIV-infected patients. It allows to monitor the largest number of anti-HIV drugs simultaneously, appearing useful in a routine laboratory, and represents an essential step to elucidate the utility of a formal therapeutic drug monitoring for the optimal follow-up of HIV-infected patients.

Original languageEnglish
Pages (from-to)258-266
Number of pages9
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume831
Issue number1-2
DOIs
Publication statusPublished - Feb 2 2006

Fingerprint

Plasma (human)
Anti-HIV Agents
High performance liquid chromatography
divinyl benzene
Reverse Transcriptase Inhibitors
efavirenz
High Pressure Liquid Chromatography
Plasmas
Drug Monitoring
HIV
Zalcitabine
Nelfinavir
Saquinavir
Lopinavir
Indinavir
Stavudine
HIV Protease Inhibitors
Didanosine
Nevirapine
Ritonavir

Keywords

  • HIV non-nucleoside reverse transcriptase inhibitors
  • HIV nucleoside reverse transcriptase inhibitors
  • HIV protease inhibitors
  • HPLC-UV
  • Therapeutic drug monitoring

ASJC Scopus subject areas

  • Biochemistry

Cite this

Simultaneous determination of 16 anti-HIV drugs in human plasma by high-performance liquid chromatography. / Notari, Stefania; Bocedi, Alessio; Ippolito, Giuseppe; Narciso, Pasquale; Pucillo, Leopoldo Paolo; Tossini, Gianna; Donnorso, Raffaele Perrone; Gasparrini, Francesco; Ascenzi, Paolo.

In: Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, Vol. 831, No. 1-2, 02.02.2006, p. 258-266.

Research output: Contribution to journalArticle

Notari, Stefania ; Bocedi, Alessio ; Ippolito, Giuseppe ; Narciso, Pasquale ; Pucillo, Leopoldo Paolo ; Tossini, Gianna ; Donnorso, Raffaele Perrone ; Gasparrini, Francesco ; Ascenzi, Paolo. / Simultaneous determination of 16 anti-HIV drugs in human plasma by high-performance liquid chromatography. In: Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences. 2006 ; Vol. 831, No. 1-2. pp. 258-266.
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