Title:Development and Validation of A High-Performance Liquid Chromatography-
Mass Spectroscopy Assay for the Bioequivalence Study of
Guanfacine Extended Release (GXR) Tablet in Human Plasma
Volume: 18
Issue: 9
Author(s): Jinying Wang and Qiaogen Zou*
Affiliation:
- Department of Pharmaceutical Sciences, Nanjing Tech University, Nanjing 210009, P.R. China
Keywords:
Guanfacine, guanfacine-13C-15N3, high performance liquid chromatography-mass spectroscopy, human plasma, protein precipitation, bioequivalence, pharmacokinetics.
Abstract: Guanfacine, an α2-adrenergic (α2A) agonist long indicated to treat hypertension, is now
being used to treat attention deficit hyperactivity disorder (ADHD) in adolescents. A simple, rapid
high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) method to detect
and quantify guanfacine provides a basis for studying its bioequivalence and pharmacokinetics in
human plasma. This assay involves quantitation of guanfacine using its stable isotope-labeled internal
standard (IS) guanfacine-13C-15N3 without the impact of ion suppression in the plasma matrix.
Electrospray Ionization (ESI) in positive mode and Multiple Reaction Monitoring (MRM)
was used for guanfacine and guanfacine-13C-15N3 at the transitions m/z 246.1→60.1 and m/z
250.0→159.1 respectively. This method's sample preparation is optimized with an accurate and
simple protein precipitation method employing methanol. Linearity was demonstrated within the
range of 0.0500-10.0 ng/mL for guanfacine in plasma with correlation coefficients greater than
0.99. The method showed excellent reproducibility. The Aaccuracy (Relative Error) ranged from
-2. to 8.9%, even at the lower limit of quantification (LLOQ), and total precision, expressed as
the coefficient of variation was between 1.6% and 10.5%. The average recoveries of guanfacine
at three spiked levels of 0.150, 1.00 and 7.50 ng/mL were 103.93, 97.91 and 100.22%, respectively.
The validated method was applied successfully to a bioequivalence study of a fixed-dose of
extended-release guanfacine hydrochloride (GXR) tablet (Test formulation) and Intuniv® (Reference
formulation) in healthy Chinese subjects, 42 subjects under fasting conditions and 30 subjects
under feeding condition. Pharmacokinetic parameters were calculated using DAS 3.2.8, and
90% confidence intervals (CIs) of AUC0-t, AUC0-∞ and Cmax for guanfacine were all within 80.00-
125.00%, suggesting that the two formulations were bioequivalent in terms of rate and extent of
absorption.
