HPLC-MS method for simultaneous quantification of the antiretroviral agents rilpivirine and cabotegravir in rat plasma and tissues

The antiretroviral agents rilpivirine (RPV) and cabotegravir (CAB) are approved as a combined treatment
regimen against human immunodeficiency virus (HIV). To fully understand the biodistribution of these agents
and determine their concentration levels in various parts of the body, a simple, selective and sensitive bioanalytical method is essential. In the present study, a high performance liquid chromatography method with mass spectrometry detection (HPLC-MS) was developed for simultaneous detection and quantification of RPV
and CAB in various biological matrices. These included plasma, skin, lymph nodes, vaginal tissue, liver, kidneys
and spleen, harvested from female Sprague Dawley rats. The suitability of the developed method for each matrix
was validated based on the guidelines of the International Council for Harmonisation of Technical Requirements
for Registration of Pharmaceuticals for Human Use (ICH) on bioanalytical method validation. Analytes were
extracted from biological samples employing a simple one-step protein precipitation method using acetonitrile.
Samples were analysed using an Apex Scientific Inertsil ODS-3 column (4.6 mm × 250 mm, 5 µm particle size),
maintained at 40 ◦C, on a HPLC system coupled with a single quadrupole MS detector. RPV was detected at a
mass-to-charge ratio (m/z) of 367.4 and CAB at 406.3. Separation was achieved using isocratic elution at 0.3 mL/
min with a mixture of acetonitrile and 0.1% (v/v) trifluoroacetic acid in water (81:19, v/v) as the mobile phase.
The run time was set at 13 min. The presented method was selective, sensitive, accurate and precise for detection
and quantification of RPV and CAB in all matrices. The developed and validated bioanalytical method was
successfully employed for in vivo samples with both drugs simultaneously.