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. 2019 May 16;8(1):880.
doi: 10.4102/ajlm.v8i1.880. eCollection 2019.

Development and validation of a high performance liquid chromatography method to determine nevirapine in plasma in a resource-limited setting

Faithful Makita-Chingombe  1 , Andrew J Ocque  2 , Robin DiFrancesco  2 , Charles Maponga  1   2 , Farai Muzambi  1 , Tsitsi G Monera-Penduka  1 , Tinashe Mudzviti  1 , Takudzwa J Mtisi  1 , Gene D Morse  2
Affiliations

Development and validation of a high performance liquid chromatography method to determine nevirapine in plasma in a resource-limited setting

Faithful Makita-Chingombe et al. Afr J Lab Med. .

Abstract

Background: There are several instances where nevirapine pharmacokinetic monitoring may be useful, such as in special populations or pharmacokinetic drug interaction studies that require the ascertainment of nevirapine pharmacokinetics in the sub-Saharan region.

Objectives: The main aim of this study was to produce a validated, sustainable and relevant nevirapine assay method that meets bio-analytical regulatory requirements.

Methods: The developed method utilised a Waters 2795 Alliance high performance liquid chromatography system with a 2996 photo diode array detector, an Atlantis dC18 5 micron, 3.9 mm × 150 mm analytical column and a gradient flow rate of 1 mL/min. Ultraviolet detection data were collected from 210 nm to 400 nm, extracted at 260 nm, and processed for nevirapine and internal standard peak height responses.

Results: The method proved to be linear (R2 0.995), precise (+1.92% - +9.69%) and accurate (-9.70% - 12.0%). Recovery for the analyte and internal standard was between 98.8% and 114%. The method showed good specificity as no interferences were caused by common African traditional medicines, anti-tuberculosis medications or other concomitant antiretrovirals nor endogenous components.

Conclusion: The method is reproducible, relevant to our setting and uses considerably low plasma volumes with preservation of some consumables, a desirable key factor in a resource-limited setting.

Keywords: high performance liquid chromatography; method development and validation; nevirapine determination.

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Conflict of interest statement

The authors declare that they have no financial or personal relationships that may have inappropriately influenced them in writing this article.

Figures

FIGURE 1
FIGURE 1
Chromatograms of blank plasma (a) and blank plasma spiked with internal standard and nevirapine (b) showing no endogenous components interference at nevirapine and internal standard retention times.
FIGURE 2
FIGURE 2
Representative nevirapine calibration curve for inter-day precision and accuracy.
FIGURE 3
FIGURE 3
Chromatogram overlay of LLOQ with antiretrovirals, no interference was observed at nevirapine and internal standard retention times for all antiretrovirals.
FIGURE 4
FIGURE 4
Chromatogram overlay of LLOQ with anti-tuberculosis drugs; no interference was observed at nevirapine and internal standard retention times.
FIGURE 5
FIGURE 5
Chromatogram overlay of LLOQ with herbs, no interference at nevirapine and internal standard retention times was observed in all herbs.
FIGURE 6
FIGURE 6
Chromatogram of patient sample. (Samples collected at Parirenyatwa Opportunistic infections clinic in 2013 and assayed at University of Zimbabwe International Pharmacology laboratory).
See this image and copyright information in PMC

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