Brazilian Journal of Analytical Chemistry - BRJAC - Brazilian Journal ...

Br J Anal Chem 2011, 03, 143–147
dE t E r m i n a t i o n o f f l u n a r i z i nE in P l a s m a By liquid
ChromatograPhy-ElECtrosPray t a n d E m m a s s
sPECtromEtry
gi l v a n viEira d o Ca r m o a , fá B i o alEssandro Pr o E n ç a Ba r ro s a , Jo s é PEdrazzoli Jú n i o r a , fl á v i o má rC i o ma C E d o mEndEs a , gi o v a n a
radomillE to f o l i a , ma r C o s no g u E i r a EBErlin B , si l v a n a aP a rE C i d a Ca l a f a t t i a * a n d Ed u a rd o César mEurEr a
A) UNIFAG (Clinical Pharmacology and Gastroenterology Unit), São Francisco University, Av. São Francisco de Assis 218, 12916-900 Bragança
Paulista, SP, Brazil.
B) Thomson Mass Spectrometry Laboratory, Institute of Chemistry, State University of Campinas, 13083-970 Campinas, SP, Brazil
*Corresponding author:
Dr. Silvana Aparecida
Calafatti
e-mail: silfatti@
saofrancisco.edu.br
Phone: +55 11 24548977;
Fax: +55 11 24548974
www.brjac.com.br
ab s t r a c t
This study presents a sensitive and selective liquid chromatography-tandem mass spectrometry
(LC-MS/MS) method for the quantification of flunarizine (FLU) in human plasma.
The mass spectrometer was operated in the selected reaction monitoring (SRM)
mode using the protonated molecule to ionic fragment combinations of m/z 404.50 >
203.00 (FLU) and m/z 369.40 > 167.00 for cinarizine (internal standard-IS) with short
run times (2.5 min). The peak areas ratio of the analyte and IS were used for quantification
of FLU. The limit of quantification (LOQ) was 0.30 ng mL -1 . The assay exhibited
a linear dynamic range of 0.30-150.00 ng mL -1 with a determination coefficient (r 2 ) of
at least 0.98. The precisions were less than 8.42% and 6.36% for the intra-batch and
inter-batch variation at 0.90, 60.00 and 120.00 ng mL -1 , respectively. This method is
suitable for bioequivalence studies accordingly to FDA guidelines.
Ke y w o r d s : flunarizine; cinarizine; LC-MS/MS; bioequivalence; liquid-liquid extraction.
In t r o d u c t Io n
Flunarizine, 1-[bis (4-fluorophenyl)methyl]-4-(3–phenyl–2–propenyl)
piperazine (CAS 52468-60-7] is a derivative
of cinarizine discovered at Janssen Pharmaceutica
in 1967 (Figure 1). Both cinnarizine and flunarizine
are calcium channel blocking drugs, but flunarizine is
2.5 to 15 times stronger than cinarizine in the frequency
of drug-induced Parkinsonism [1].
fi g u rE 1. ChEmiCal struCturE o f (a) f l u n a r i z i nE a n d (B) CinarizinE.
It is effective in the prophylaxis of migraine, occlusive
peripheral vascular disease, and vertigo of central and
peripheral origin and as an adjuvant in the therapy of
epilepsy. Flunarizine is well absorbed after oral administration,
reaching peak plasma levels within 2-4 hours
and reaching steady state at 5-6 weeks. After extensive
hepatic metabolism, flunarizine and its metabolites
are excreted through the feces via the bile. The mean
terminal elimination half-life is about 18 days. Plasma
protein binding is 99% [2-5].
Liquid chromatography-tandem mass spectrometry
(LC-MS/MS) is the state of–the-art in quantification of
drugs in biological matrices due to improved detectivity,
selectivity and speed of this technique. The LC-MS/MS
system used in this study is one of the most utilized analytical
tools for determining pharmacokinetic parameters
of a drug with high precision and reliability [6, 7].
Other techniques have been used previously to analyze
flunarizine in a variety of matrices. These methods
include gas chromatography coupled to mass spectrometry
(GC-MS) [8,9] and LC-MS [10]. Some of these techniques
were not developed for use in biological matrices,
while others have either a very high limit of quantification
(LOQ) or are much too complex, limiting their application
for a large number of samples. The use of LC-MS/
MS to determine flunarizine levels above the lower limit
of quantified plasma levels is already described, but the
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