Transcript Done-poster
Steroselective HPLC assay of donepezil enantiomers in tablets
and plasma and its application to pharmacokinetics in Rats
Bushra T.
Mahasen A.
*, Heba H.
2
Radwan
3
Abdine
, Hassan Y.
4
Aboul-Enein
and Kenichiro
5
Nakashima
of Pharmaceutics, 2Department of Clinical Pharmacy, 3Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, PO Box 22452, Riyadh 11495; 4 Centre for Clinical Research (MBC-03-65), King Faisal
Specialist Hospital and Research Centre, P.O. Box 3354, Riyadh 11211, Saudi Arabia; 5 Department of Clinical Pharmacy, Nagasaki University, 1-14, Bunkyo-machi, Nagassaki 852-8521, Japan.
Introduction
Results and Disscusion
Experimental
Chromatographic system and
conditions:
Donepezil (DH) is a reversible inhibitor of
acetylcholinesterase. It is the second drug
approved by the FDA for the treatment of mild to
moderate dementia of the Alzheimer’s type. DH
was demonstrated to be a potent and selective
inhibitor of brain acetylcholinesterase with less
adverse effects than physostigmine and tacrine
[1-2].
DH has a chiral center adjacent to a carbonyl
group. It is commercially available as the
hydrochloride salt and is used as a racemic
mixture in dosage forms. The donepzil
enantiomers have differing extents of inhibition
against acetylcholinestrerase in vivo and in vitro
[3]; hence it was necessary to clarify the
pharmacokinetics of each isomer during drug
development course. The importance to develop
new chiral discrimination separation systems to
determine enantiomers in bulk substance, drug
formulations and in biological fluids increases.
This is because stereoisomers may have
different pharmacological effects and also show
differences
in
pharmacokinetic
and
pharmacodynamic properties [4].
Several pharmacokinetic studies [3, 5-7]
including drug interactions with donepezil have
been
reported
using
high
performance
chromatographic (HPLC) method with UV
detection [4], but methodology was not
described. It should be mentioned that Matsui et
al. [3] were the only group addressed the
pharmacokinetics of DH enantiomers in human
using
liquid
chromato-graphy–mass
spectrometry,
which
was
sensitive,
but
complicated.
Aim of the work
HPLC System: Waters' system
Column: Chiralcel OD
Mobile phase: n- hexane, isopropyl alcohol &
triethylamine (87:12.9:0.1, v/v/v)
Flow rate: 1.0 ml/min
Detection: ultraviolet detection at 268 nm
Injection volume: 75 µl
0.9
Determination of donepezil in
pharmaceutical dosage forms:
0.9
0.6
Two commercially available formulations
(Aricept® tablets) labelled to contain either 5
or 10 mg donepezil hydrochloride were
analyzed.
0.6
0.3
0.3
0
0
0
Determination of DH in spiked rat
plasma:
B
1.2
10
20
I
0
-0.3
10
20
(-)R-DH
(+)S-DH
0.1
0.01
Intra-day
Nominal
(ng/ml)
Pharmacokinetics of DH in
rats:
4
6
Time, h
8
Mean plasma concentrations (+
SD) time profile of (-)R and
(+)S- donezepil enantiomers in
rats after receiving 3mg/kg oral
dose of racemic donepezil.
A: Plasma samples free of drug (Blank).
B: Plasma samples spiked with 200 ng/ml of
donepezil HCl (B).
I: (-)R-DH
II: (+)S-DH
Concentration
2
-0.3
Time, min
Two hundred L of rat plasma samples
were spiked with 20-50 L of DH stock
solutions.
Protein was precipitated with 600 L of
acetonitrile.
Vortexed for 1 min, & centrifuged at
20,000 rpm for 15 min.
The supernatant was evaporated to
dryness under a stream of N2.
The residue was reconstituted in 100 L
of the mobile phase.
The absolute recoverry, Intra- and Interday
accuracy
and
precision
were
evaluated by assaying quality controls
with differrent concentrations of DH
(0.05 - 2 g /ml).
12
Parameter
(-)R-DH
(+)S-DH
Cmax*, ng/ml
143 ± 66
148 ± 110
Tmax*, h
5.2 ± 4
3.3 ± 2.7
AUC, mg h/L
3.94
2.91
Inter-day
Recovery %
Precision %
10
H. Sugimoto, Y. Yamanishi, Y.
Iimura, Y. Kawakami, Curr. Med.
Chem. 7 (2000) 303– 339.
[2] E. Scarpini, P. Scheltens, H. Feldman,
Lancet Neurology 2 (2003) 539–547.
[3] K. Matsui, Y. Oda, H. Nakata,
T.Yoshimura, J. Chromatography B,
729 (1999) 147–155.
[4] J.P. Mason, A.J.Hutt Sterochemical
Aspects of Drug Metabolism" Chapter
3 in "The impact of Sterochemistry in
Drug Development and Use", H.Y.
Aboul-Enein and I.W. Wainer (eds.)
John Wiley & Sons Inc., New
York,
USA, 1997, pp. 45- 105.
[5] J.W. Lee, S.L. Rogers, L.T. Friedhoff,
M.R. Stiles, N.M. Cooper, Pharm. Res.
9 (1992) S350.
[6] P.J. Tiseo, S.L. Rogers, L.T. Friedhoff,
Clin. Pharmacol. Ther. 61(1997) 184. (PII75).
[7] N. Yasui-Furukori, R. Furuya, T.
Takahata, T. Tateishi, J. Chromatography B, 768 (2002) 261–265.
Accuracy %
Precision %
Accuracy %
(-)R-DH
(+)S-DH
(-)R-DH
(+)S-DH
(-)R-DH
(+)S-DH
(-)R-DH
(+)S-DH
(-)R-DH
(+)S-DH
Cl/F, L/h/kg
0.038
0.051
50
95.7
93.9
5.9
6.4
5.1
4.6
4.8
4.0
4.3
5.1
t½ , h
27.2
16.8
100
89
92
6.5
5.4
5.3
3.7
7.9
8.6
9.7
8
1000
90.7
88.9
7.1
4.4
3.9
2.9
5.7
8.1
9.3
10
Pharmacokinetic parameters of donepezil after
0.3 mg/kg PO dose in the rat (n=6).
* Calculated from the actual data
Single dose oral study has been performed.
An equivalent to 3 mg drug/ Kg body
weight of rats was calculated and as given
as a suspension.
2000
97.3
95.5
4.3
3.1
1.2
2.8
6.5
7.1
2.7
4.5
Precision (RSD%), accuracy (relative error%) and recovery for determination of donepezil (-)R
and (+)S-DH enantiomers in spiked rat plasma samples (n = 6).
(+)S-DH
Dosage Form
Aricept® 5 mg
Aricept® 10 mg
II
[1]
0
(-)R-DH
The
present
study
describes
a
stereospecific, isocratic, normal-phase
HPLC method for the determination of
the enantiomers of DH in tablets and
plasma
and
its
application
to
pharmacokinetic studies in rats.
1.5
A
1.2
Response
Donepezil hydrochloride (DH), {2,3-dihydro-5,6dimethoxy-2-[[1-(phenyl -methyl)-4-piperidinyl]
methyl]-1H-inden-1-one}hydrochloride
1.5
References
1
Enantiomer Concentrations,g/ml
1Department
1
Al-Quadeib ,
Content* (%)
RSD (%)
Content* (%)
RSD (%)
95.6
96.9
8.3
3.2
96.2
97.3
9.6
3.4
Determination of the mean (-)R and (+)S- donezepil enantiomers content in
pharmaceutical dosage forms (n = 6)
Acknowledgment
Special thanks to King Suad University,
Research Center and to King Faisal
Specialist Hospital and Research Center.
Conclusion
A new simple, stereospecific, reproducible,
accurate and sensitive HPLC method has
been successfully developed and applied to
the determination of DH enantiomers in rats
and in its commercial tablets. The results
were used to generate profiles and to
estimate the pharmacokinetic parameters of
DH enantiomers up to 12 h after oral
administration in rats. The disposition of the
two enantiomers seems to be different.