title: Validation and clinical application of a method to quantify nevirapine in dried blood spots and dried breast-milk spots.
abstract: OBJECTIVE
  The validation and clinical application of an LC-MS/MS method for the quantification of nevirapine in dried blood spots (DBS) and dried breast-milk spots (DBMS) are presented.
  
  
  METHODS
  DBS and DBMS were prepared from 50 and 30 μL of nevirapine-spiked whole blood and human breast milk, respectively. Chromatographic separation was achieved on a reverse-phase C18 column with 0.1% formic acid in water/acetonitrile using a solvent gradient programme at a flow rate of 400 μL/min, and detection was by a TSQ Quantum Access triple quadrupole mass spectrometer. The clinical application was evaluated in HIV-positive nursing mothers and their breastfed infants.
  
  
  RESULTS
  The assay was validated over the concentration range 50-10,000 ng/mL. Accuracy ranged from 93.3% to 113.4% and precision ranged from 1.9% to 12.0%. The mean (percentage coefficient of variation) recovery of nevirapine from DBS and DBMS was ≥ 70.7% (≤ 8.2) and the matrix effect was ≤ 1.04 (≤ 6.1). Nevirapine was stable in DBS and DBMS for ≥ 15 months at room temperature and -80°C. Mean (SD) AUC0-12, Cmax and Cmin in maternal plasma versus breast milk were 57,808 ng · h/mL (24,315) versus 55,817 ng · h/mL (22,368), 6140 ng/mL (2605) versus 5231 ng/mL (2215) and 4334 ng/mL (1880) versus 4342 ng/mL (2245), respectively. The milk-to-plasma concentration ratio over the dosing interval was 0.94 (0.15). Infant plasma concentrations 2 and 8 h after maternal dosing were 580.6 ng/mL (464.7-1607) and 584.1 ng/mL (381.5-1570), respectively.
  
  
  CONCLUSIONS
  These methods further extend opportunities for conducting clinical pharmacokinetic studies in nursing mother-infant pairs, especially in resource-limited settings.
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK Faculty of Pharmacy, Obafemi Awolowo University, Ile-Ife, Nigeria.
authors.forename: Adeniyi
authors.initials: A
authors.lastname: Olagunju
authors.affiliation: Liverpool Bioanalytical Facility, Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool L69 3GA, UK.
authors.forename: Alieu
authors.initials: A
authors.lastname: Amara
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK.
authors.forename: Catriona
authors.initials: C
authors.lastname: Waitt
authors.affiliation: Liverpool Bioanalytical Facility, Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool L69 3GA, UK.
authors.forename: Laura
authors.initials: L
authors.lastname: Else
authors.affiliation: Liverpool Bioanalytical Facility, Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool L69 3GA, UK.
authors.forename: Sujan D
authors.initials: SD
authors.lastname: Penchala
authors.affiliation: Faculty of Pharmacy, Obafemi Awolowo University, Ile-Ife, Nigeria.
authors.forename: Oluseye
authors.initials: O
authors.lastname: Bolaji
authors.affiliation: Faculty of Pharmacy, Obafemi Awolowo University, Ile-Ife, Nigeria.
authors.forename: Julius
authors.initials: J
authors.lastname: Soyinka
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK.
authors.forename: Marco
authors.initials: M
authors.lastname: Siccardi
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK.
authors.forename: David
authors.initials: D
authors.lastname: Back
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK.
authors.forename: Andrew
authors.initials: A
authors.lastname: Owen
authors.affiliation: Department of Molecular and Clinical Pharmacology, University of Liverpool, 70 Pembroke Place, Liverpool L69 3GF, UK khoo@liverpool.ac.uk.
authors.forename: Saye
authors.initials: S
authors.lastname: Khoo
mesh_terms: D000328:Adult; D019380:Anti-HIV Agents / Q000493:pharmacokinetics*; D002853:Chromatography, Liquid / Q000379:methods; D016903:Drug Monitoring / Q000379:methods* / Q000592:standards; D004355:Drug Stability; D005260:Female; D006801:Humans; D008895:Milk, Human / Q000737:chemistry; D019829:Nevirapine / Q000493:pharmacokinetics*; D010949:Plasma / Q000737:chemistry; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D019380:Anti-HIV Agents; D019829:Nevirapine