title: Determination of total and unbound concentrations of lopinavir in plasma using liquid chromatography-tandem mass spectrometry and ultrafiltration methods.
abstract: Lopinavir is an HIV protease inhibitor with high protein binding (98-99%) in human plasma. This study was designed to develop an ultrafiltration method to measure the unbound concentrations of lopinavir overcoming the non-specific binding issue. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of total concentrations of lopinavir in plasma was developed and validated, and an adaptation was also optimized and validated for the determination of unbound concentrations. The chromatographic separation was performed with a C18 column (100 mm × 2.1mm i.d., 5 μm particle size) using a mobile phase containing deionized water with formic acid, and acetonitrile, with gradient elution at a flow-rate of 350 μL min(-1). Identification of the compounds was performed by multiple reaction monitoring, using electrospray ionization in positive ion mode. The method was validated over a clinical range of 0.01-1 μg/mL for human plasma ultrafiltrate and 0.1-15 μg/mL in human plasma. The inter and intra-assay accuracies and precisions were between 0.23% and 11.37% for total lopinavir concentrations, and between 3.50% and 13.30% for plasma ultrafiltrate (unbound concentration). The ultrafiltration method described allows an accurate separation of the unbound fraction of lopinavir, circumscribing the loss of drug by nonspecific binding (NSB), and the validated LC-MS/MS methodology proposed is suitable for the determination of total and unbound concentrations of lopinavir in clinical practice.
authors: Unité de Recherche Clinique, AP-HP, Hôpital Tarnier, Paris, France. Electronic address: 07silvia@gmail.com.
authors: S M
authors: SM
authors: Illamola
authors: Service de Pharmacologie Clinique, AP-HP, Groupe Hospitalier Paris Centre, Paris, France.
authors: L
authors: L
authors: Labat
authors: Service de Pharmacologie Clinique, AP-HP, Groupe Hospitalier Paris Centre, Paris, France; EAU08 Université Paris Descartes, Sorbonne Paris Cité, Paris, France.
authors: S
authors: S
authors: Benaboud
authors: AP-HP, Hôpital Pitié Salpêtrière, Service de maladies infectieuses, Paris, France; Sorbonne université, UPMC Paris 06, UMR_S 1136 Pierre Louis Institute of Epidemiology and Public Health, F-75013, Paris, France.
authors: R
authors: R
authors: Tubiana
authors: INSERM, Centre for research in Epidemiology and Population Health, U1018, Equipe VIH et IST, Le Kremlin Bicêtre, France; AP-HP, Hôpital Bicêtre, Service d'Epidemiologie et Santé Publique, Le Kremlin-Bicêtre, France; Université Paris Sud, UMRS 1018, Le Kremlin-Bicêtre, France.
authors: J
authors: J
authors: Warszawski
authors: Unité de Recherche Clinique, AP-HP, Hôpital Tarnier, Paris, France; Service de Pharmacologie Clinique, AP-HP, Groupe Hospitalier Paris Centre, Paris, France; EAU08 Université Paris Descartes, Sorbonne Paris Cité, Paris, France; CIC-0901 INSERM, Cochin-Necker, Paris, France.
authors: J M
authors: JM
authors: Tréluyer
authors: Service de Pharmacologie Clinique, AP-HP, Groupe Hospitalier Paris Centre, Paris, France; EAU08 Université Paris Descartes, Sorbonne Paris Cité, Paris, France.
authors: D
authors: D
authors: Hirt
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D017322:Clinical Trials, Phase II as Topic; D017326:Clinical Trials, Phase III as Topic; D005260:Female; D015658:HIV Infections / Q000517:prevention & control / Q000635:transmission; D006801:Humans; D018445:Infectious Disease Transmission, Vertical / Q000517:prevention & control; D061466:Lopinavir / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification; D011247:Pregnancy; D011251:Pregnancy Complications, Infectious / Q000097:blood / Q000188:drug therapy / Q000821:virology; D016032:Randomized Controlled Trials as Topic; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014462:Ultrafiltration / Q000379:methods*
chemical_list: D061466:Lopinavir
keywords: Lopinavir; Nonspecific binding; UPLC–MS/MS; Ultrafiltration; Unbound fraction