Original Research

Enteric-coated alendronate
sodium nanoliposomes: a novel
formula to overcome barriers for
1. Introduction
2. Experimental methods
the treatment of osteoporosis
3. Results Khaled Mohamed Hosny†, Osama Abdelhakim Aly Ahmed &
4. Discussion Rana Tariq Al-Abdali
†
5. Conclusions King Abdulaziz University, Faculty of Pharmacy, Department of Pharmaceutics, Jeddah,
Saudi Arabia
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13

Background and objectives: Alendronate sodium (ALS) is the most common

drug used for the treatment of osteoporosis. The challenges facing ALS use
include: very poor oral bioavailability (0.6%), esophageal ulcers, and compli-
cated instructions for its use. The objective of this research is to utilize nano-
technology to formulate ALS into enteric-coated nanoliposomes (NLS) to
overcome the previously mentioned drawbacks.
Methods: NLS were prepared with lipid components of phosphatidylcholine
(PC), cholesterol (CH), and lecithin (Lec) in ratios 4:1:1, 4:2:1, 4:3:1, and 4:4:1,
respectively. Formulas that showed the highest entrapment efficiency were
prepared either alone or mixed with positive and negative charge-inducing
For personal use only.

agents and coated with Eudragit L100. Eudragit-coated NLS (EuC-NLS) were
evaluated for particle size, zeta potential, morphological examination, and
drug release in pH 1.2 and pH 7.4 media. The pharmacokinetic study was
carried out in rabbits.
Results: Spherical NLS were successfully developed with a mean size range
from 70 to 150 nm. EuC-NLS with PC:CH:Lec:dicetyl phosphate (4:3:1:1)
successfully resist the release of ALS in acidic environments and enhanced
the bioavailability in rabbits 12-fold compared with the marketed tablets.
Conclusions: EuC-NLS is a promising novel formula for ALS with higher bio-
availability and a lower dose, avoiding the side effects of esophageal
ulceration.

Keywords: alendronate sodium, enteric coating, Eudragit, nanoliposomes

Expert Opin. Drug Deliv. (2013) 10(6):741-746

1. Introduction

Alendronate sodium (ALS) is a nitrogen-containing oral bisphosphonate used for

the treatment of osteoporosis. ALS exhibits a potent and selective inhibitory effect
on osteoclast-mediated bone resorption and was approved for osteoporosis in post-
menopausal females, males and also in osteoporosis due to glucocorticoid use [1,2].
Approximately 50% of the patients treated with ALS showed decrease in the inci-
dence of osteoporotic fracture in the hips and spine by improving bone density
and reducing bone resorption [3]. The first challenge for use of ALS is the low
oral bioavailability, 0.6 -- 0.7% [4]. ALS is best absorbed when taken 2 h before
food or after overnight fasting, as a result of ALS hydrophilicity and the negative
charges that hinder crossing of the lipoid biomembrane of the digestive tract [5].
The second main challenge is the side effects of ALS which include esophageal
ulceration or erosion mostly with bleeding. As a result of these constraints, the
drug is subject to precautions and complicated instructions.

10.1517/17425247.2013.799136 © 2013 Informa UK, Ltd. ISSN 1742-5247, e-ISSN 1744-7593 741
All rights reserved: reproduction in whole or in part not permitted
 K. M. Hosny et al.
Researchers had shown morning dosing is a major compli-
ance problem with osteoporosis therapy [1,6-8]. Proper ALS
administration instructions should be given to the patient,
by the physician, to avoid esophagitis and upper GI tract
lesions. These instructions include: dose of ALS should be
taken with a full glass of water and the patient should remain
upright at least 30 min before the first food or beverage of the
day. Up to 60% of patients stop taking weekly ALS during the
first year as a result of its side effects and complicated
instructions [8]. Different strategies were tried to improve
the bioavailability of ALS. These include the utilization of
nanotechnology to prepare biodegradable ALS nanopar-
ticles [9]. Han et al. [10] delayed the residence time of ALS in
the GI tract through the preparation of a mucoadhesive lipo-
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13
somal formulation that improved the oral bioavailabilty of
ALS in rats by 2.6-fold. Katsumi et al. [11] developed an
ALS microneedle array using hyaluronic acid that enhanced
ALS bioavailability by 90% in rats.
Small liposomes with diameters of the order of 100 nm are
frequently used as a carrier to utilize their better absorption
and distribution [12]. In addition, NLS have the advantages
of nanoparticles, which improve the adhesion to and absorp-
tion into the intestinal epithelial cells. In this case, the NLS
delivery system might be a highly effective way to improve
For personal use only.
ALS absorption [13]. The technology of enteric coating is
used for several applications to protect the drug from gastric
fluids and enzymes, or to protect the esophagus and stomach
from the irritant action of some drugs. Enteric coating pre-
vents the drug in nanoparticles from disintegration in the
stomach and it will increase the intact amount of nanoparticles
delivered to the duodenum of the small intestine [14], which
improves the bioavailability of ALS and reduces the mucosal
irritant effect. The main objective of this work is to formulate
ALS in the form of NLS to improve the bioavailability of the
drug. NLS are then coated with enteric coating material
(Eudragit L100) to reduce side effects on the esophagus and
allow elderly patients to tolerate the drug easily.
2. Experimental methods
2.1 Materials
ALS was kindly supplied by Saja Pharmaceuticals Co. Ltd.
(Jeddah, Saudi Arabia); phosphatidylcholine from egg yolk,
lecithin and cholesterol were purchased from Sigma Chemical
Co. (St. Louis, Missouri, USA); stearylamine and dicetyl
phosphate (DP) were obtained from Fluka Chemical Co.
(Buchs, Germany). Eudragit was kindly supplied by Evonik
Industries AG, (Essen, Germany) and trehalose was purchased
from Caesar and Loretz (Hilden, Germany). Other chemicals
and reagents were of analytical grade.
2.2 Methods
2.2.1 Preparation of enteric-coated NLS
NLS were prepared according to the method described by
Vincourt et al. [15]. Lipid components of NLS were
742 Expert Opin. Drug Deliv. (2013) 10(6)
phosphatidylcholine (PC), cholesterol (CH), and lecithin
(Lec) in gram ratios 4:1:1, 4:2:1, 4:3:1, and 4:4:1, respec-
tively. Trehalose was selected as the lyoprotectant. The mini-
mal trehalose/lipid ratio tested was 3, in order to avoid
lyophilized cake collapse. NLS lipid components (300 mg)
either alone or mixed with a positive charge-inducing agent
stearylamine (SA) or negative charge-inducing agent DP
were dissolved in 10 ml chloroform and a thin lipid film
formed after removal of chloroform by rotary evaporator.
The lipid film was hydrated by 10 ml phosphate buffered
solution (pH 7.4) containing 50 mg ALS and 0.9 mg treha-
lose with or without the charge-inducing agents SA and DP.
Subsequently, the suspensions were sonicated using probsoni-
cator (VCX750, Sonics & Materials, USA) for a time suffi-
cient to obtain nanosized liposomes. The suspensions were
centrifuged for 30 min at 15,000 rpm to remove the un-
entrapped ALS and then the precipitates of certain formulae
were dispersed in buffer solution (pH 7.4) in which Eudragit
L100 was previously dissolved and sonicated for 5 min. The
suspensions were rendered acidic by addition of acidic solu-
tion at pH 2 in order to precipitate the Eudragit. The final
suspensions were centrifuged for 30 min at 15,000 rpm and
the EuC-NLS were then subjected to freeze--drying.
Determination of ALS entrapment efficiency
2.2.2
(EE%) in NLS
The lyophilized NLS were sonicated with methanol for
15 min. The concentration of ALS in methanol was measured
at 333 nm by spectrophotometric method proposed by
Al Deeb et al. [16]. EE% was calculated as shown in the
equation:
(1)
EE% = [ALSE / ALST ] × 100
where ALSE and ALST represent the encapsulated ALS and
total ALS amount, respectively.
2.2.3Examination of surface morphology by scanning
electron microscope
Neutral, negatively and positively charged samples of NLS were
subjected to morphological examination by using SEM. Sam-
ples were reconstituted with deionized water and spread over
carbon tape. Samples were then dried in vacuum and coated
with gold and examined under the electron microscope to deter-
mine the surface morphology of the optimized lyophilized NLS.
Size analysis and zeta potential measurement
2.2.4
of NLS
The zeta potential and the mean particle size were measured
by using dynamic light scattering particle size analyzer
Zetatrac (Microtrac, Inc., USA).
In vitro gastro-resistance
2.2.5
To evaluate the enteric nature of EuC-NLS, equivalent to
10 mg of ALS was suspended in 1 ml of 0.1 N hydrochloric
 Enteric-coated alendronate sodium nanoliposomes

Table 1. EE%, particle size, and zeta potential for different formulations of EuC-NLS.

Formula Lipid composition Ratio EE% Particle size Zeta potential Zeta potential
before coating after coating

F1 PC:CH:Lec 4:1:1 29.88 57 nm - 2.24 26.43

F2 PC:CH:Lec 4:2:1 35.63 59 nm - 3.12 29.25
F3 PC:CH:Lec 4:3:1 49.39 70 nm - 3.35 16.43
F4 PC:CH:Lec 4:4:1 41.76 65 nm - 4.27 26.34
F5 PC:CH:Lec:SA 4:3:1:1 54.16 150 nm 34.65 49.66
F6 PC:CH:Lec:DP 4:3:1:1 41.92 110 nm - 21.45 4.92

A. B. C.
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13

Figure 1. SEM image of freeze--dried ALS NLS, (A) Neutral NLS, (B) Negatively charged NLS, (C) Positively charged NLS.

acid (pH 1.2) in a glass cylinder with a length of 6 cm and a Program. One-way analysis of variance (ANOVA) was
diameter of 2.5 cm. This cylinder was closed from one end by employed to assess the significance of the difference between
For personal use only.

a presoaked dialysis membrane and suspended in a basket in
the (USP) dissolution tester containing 900 ml of 0.1 N
hydrochloric acid (pH 1.2) maintained at 37 C and
100 rpm. Samples were withdrawn after 0.5, 1, 1.5 and 2 h,
and assayed for ALS.
2.2.6In vitro drug release
The in vitro profile for ALS from EuC-NLS was evaluated at
phosphate buffer pH 7.4. ALS-NLS, equivalent to 10 mg of
ALS, were suspended in 1 ml of phosphate buffer (pH 7.4) in
a glass cylinder as described in the previous experiment at
75 rpm. A 5 ml sample was withdrawn after 5, 10, 15, 30, 45,
60, 90, 120, 150 and 180 min, and assayed for ALS at 333 nm.
2.2.7 In vivo pharmacokinetics study
Healthy albino male rabbits weighing 2 -- 2.5 kg were divided
into two groups, six animals each, and were fasted for 12 h
before and during the experiment. The in vivo animal studies
protocol was revised and approved by the ethical committee
of King Abdulaziz University, Jeddah, Saudi Arabia. An oral
ALS formula of freeze--dried EuC-NLS was given to one
group, and the marketed ALS tablets (10 mg) were given to
the other group. The dose of ALS was 1 mg/kg for each ani-
mal group. The collection of blood samples was carried out
at the following time intervals: 0.5, 0.75, 1, 1.5, 2.5, 3.5, 5,
6, 12, and 24 h. Serum samples were then analyzed by
HPLC. Pharmacokinetic parameters were calculated and
presented as mean ± S.D. Cmax and Tmax after oral adminis-
tration were calculated directly from the plasma concentra-
tion--time curve using WinNonlin
Nonlinear Estimation
the tested NLS formulation and the reference at a level
(p £ 0.05) using the SPSS program [17]. AUC0 -- 24 was calcu-
lated using the linear trapezoidal rule [18]. AUC0-¥ was deter-
mined by adding the last measured plasma concentration
divided by the elimination rate constant (Kel) to AUC0 -- 24.
The relative bioavailability of enteric-coated NLS is calculated
by using the following formula [19].
(2)
Relative bioavailablity = [AUC NLS × Dose Tablet ] /
[AUC Tablet × Dose NLS]
3. Results
3.1 Factors affecting entrapment efficiency
Results in Table 1 show that the increase in the CH content of
NLS without charged additives (DP and SA) up to a ratio of
4:3:1 increased the EE% of the prepared NLS. Above this
ratio there was a decrease in the EE% by the increase in CH
level. Thus, the NLS formed from lipid content in a ratio of
4:3:1 (PC:CH:Lec) was selected in regards to the entrapment
efficiency (EE% = 49.39%). Incorporation of SA leads to an
increase in the EE% from 49.39% (for the neutral NLS)
to 54.16%. However, the EE% decreases to 41.92% by
incorporation of DP within the nanoliposomal membrane.
3.2 Morphology and size analysis of liposomes
Figure 1 shows the SEM for freeze--dried ALS-NLS prepared
by the lipid film hydration technique by a molar ratio of
4:3:1 (PC:CH:Lec), revealing the presence of homogenous,
well-identified spheres, which existed in dispersed and

Expert Opin. Drug Deliv. (2013) 10(6) 743

 K. M. Hosny et al.

60
About 30% ALS was released within 15 min, which highlights
the rapid dissolution of the enteric coating material in pH 7.4.
50 In addition, nearly 100% ALS was released within 3 h. These
% ALS released

results indicated that the formula composed of EuC-NLS

40 Marketed ALS product with lipid content PC:CH:Lec:DP (F6) in molar ratio
Positive NLS 4:3:1:1 is the optimum enteric-coated formula, and was
30
Neutral NLS selected for an in vivo pharmacokinetic study.
20 Negative NLS

3.5 In vivo pharmacokinetics study

10
There was a statistically significant difference of the Tmax and
0 Cmax data for NLS compared with the marketed ALS product
0 30 60 90 120 (Table 2). It was observed that the absorption of ALS was rapid
Time (min) and reached its peak plasma concentration in 0.75 h ± 0.09 in
the case of the commercial tablet, whereas the mean Tmax for
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13

the tested NLS was 3.5 h ± 0.61 (Table 2). The mean
Figure 2. In vitro release of ALS from EuC-NLS formulae and AUC0-¥ for the marketed ALS tablet was significantly different
marketed ALS product in pH 1.2.
compared with NLS. These results confirmed the enhance-
ment of ALS bioavailability > 12-fold when formulated as
enteric-coated NLS.
100
4. Discussion
80
% of ALS released

EE% is used to evaluate the use of different molar ratios for

60
PC:CH:Lec (4:1:1, 4:2:1, 4:3:1, and 4:4:1). The effect of add-
For personal use only.

ing negative and positive charge inducers, DP and SA, on the

40
20
0 stable and less permeable, leading to more drug reten-
0 30 60 90
Time (min)
Figure 3. In vitro release of ALS from Eudragit-coated
negatively charged, positively charged and neutral NLS in
pH 7.4.
aggregate collections. The particle size analysis test showed
that all prepared liposomes occurred in the nanosize range,
this highlights the efficiency of the preparation method in
obtaining nanosized liposomes (Table 1). The size analysis
was 70 nm for neutral NLS. In the case of charged NLS,
the size increased from 70 to 150 nm in positive NLS, and
to 110 nm in negative NLS.
3.3 In vitro gastro-resistance
To evaluate the enteric nature of the coated NLS, Figure 2
shows the in vitro release profile of ALS EuC-NLS in pH
1.2. The percentages of ALS released within 2 h were 3, 9,
14 and 65% from negatively, neutral, positively charged
NLS, and marketed ALS tablets, respectively.
3.4 In vitro drug release
Figure 3 shows the in vitro profile for ALS from Eudragit-
coated negatively charged NLS in phosphate buffer pH 7.4.
Positive NLS
EE% of ALS within NLS was investigated. According to the
Neutral NLS results in Table 1, by increasing the CH content in the lipid
Negative NLS bilayer, the nanoliposomal membrane becomes more rigid,
120 150 180 tion [20,21]. Above certain concentrations for CH, the regular
linear structures of the membrane begin to disrupt, which
leads to a reduction in the EE%. Thus, the NLS formed
from lipid content in a ratio of 4:3:1 (PC:CH:Lec) with EE
% value of 49.39% was selected for further evaluations. Incor-
poration of a positively charged inducing agent (SA) increased
the zeta potential of liposomes from - 3.35 to 34.65, which
increased the stability of the system against liposomal aggrega-
tion and increased the EE% as a result of the electrostatic
attraction between the negative phosphate group of ALS and
the positive charge of SA. The incorporation of a negatively
charged inducing agent (DP) within the nanoliposomal mem-
brane reduces the zeta potential from - 3.35 to - 21.45, which
increased the stability against liposomal aggregation, but
reduced the EE%, which could be attributed to the repulsion
of both negative charges for DP and ALS [22].
The increase in liposomal size after incorporation of either
positively or negatively charged inducing agents into the lipo-
somal bilayer could be attributed to an increase in the spacing
between the adjacent bilayers [23], resulting in the formation
of liposomes larger in size compared with the neutral ver-
sions [21,24]. Furthermore, the positively charged lipid, from
the use of SA, electrostatically attracts ALS anions that would
be expected to push phospholipid head groups apart, hence
increasing the particles’ diameters [25]. Benech et al. [26]
reported that the improved encapsulation efficiency in

744 Expert Opin. Drug Deliv. (2013) 10(6)

 Enteric-coated alendronate sodium nanoliposomes

Table 2. Pharmacokinetic parameters after oral administration of EuC-NLS and marketed ALS tablet (n = 6).

Formulation Tmax (h) Cmax (ng/ml) AUC0 -- 24 (ng·h/ml) K (h-1) AUC0-` (ng·h/ml)

EuC-NLS 3.5 ± 0.61 24.76 ± 4.43 4280 ± 304.5 0.187 ± 0.02 6875.4 ± 407.2
Marketed ALS tablet 0.75 ± 0.09 5.43 ± 2.54 340.6 ± 31.23 0.672 ± 0.12 570.2 ± 43.15

The mean difference is significant at the 0.05 level.

charged liposomes increases the effective diameter, promotes
curvature changes and creates a more swollen membrane
structure. In addition, the increase in particle size is attributed
to the formation of multilamellar NLS. As the charge-
inducing agent makes the liposomal membrane suitable to
structural rearrangement due to the repulsion which occurs
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13
mediums to avoid the adverse effects of ALS, NLS composed of
PC:CH:Lec:DP (F6) in molar ratio 4:3:1:1 was selected for fur-
ther evaluations. The mean pharmacokinetic parameter results
showed a delay in Tmax in case of NLS formula compared
with marketed ALS tablets that is attributed to the efficient
coating with Eudragit L100, which hinders the release of ALS

between similarly charged phospholipid head groups, it ulti-
mately leads to structural transformation due to adhesion-
mediated processes such as bilayer rupture and fusion [27,28].
This occurred as a result of cross-linking for the NLS aggre-
gates by the charge-inducing agent. Then, rapid spreading of
the contact area deforms the liposomes as they flatten against
each other, which places the bilayer under increased tension,
which is relieved by fusion and rupture. Upon bilayer rupture,
vesicles collapse, flattening against each other to form
multilamellar liposomes that increased the particle size. Simi-
For personal use only.
in the stomach (acidic pH) and releases the drug later in the
intestine. The enhancement in bioavailability, > 12-fold, could
be attributed to the size of NLS (110 nm), which have the
advantages of nanoparticles, and improve the adhesion to and
absorption into the intestinal epithelial cells [30]. In addition,
the liposomal vesicles may promote the uptake by M cells in
the Peyer’s patches and increase absorption through the lym-
phatic pathway [31]. Permeation of intact NLS through the
intestinal epithelia pathway is considered as an additional pos-
sible mechanism for the enhancement in bioavailability of the

lar findings were also reported for incorporation of charge-
inducing agents into azathioprine [29], acetazolamide [21],
and indomethacin liposomes [22].
In the case of negatively charged NLS, an electrostatic attrac-
tion occurred during coating between the positively charged
Eudragit and the negatively charged NLS leading to improved
efficiency of Eudragit coating compared with neutral and pos-
itively charged NLS. This efficient coat on negative NLS resists
the release of ALS in acidic pH 1.2 medium. This interaction
was also confirmed by the increase in the zeta potential value
of the negatively charged NLS after coating with Eudragit
from - 21.45 to 4.92 mV. In the case of positively charged
NLS, the repulsion of the positive charges, for Eudragit and
NLS, reduced the efficiency of the enteric coating and conse-
quently the gastro-resistance character of the prepared formula
that is considered undesirable in enteric-coated dosage forms.
As the main aim of this work is to prevent ALS release in acidic
prepared NLS formula [32].
5. Conclusions
Formulation of negatively charged enteric-coated ALS-NLS,
as a novel drug delivery system, provided the maximum
gastro-resistant ALS release. The oral bioavailability of ALS
was enhanced by > 12-fold in relation to the commercially
available product. The improved formula of ALS could elim-
inate the major drawbacks of conventionally used tablets, and
allow osteoporotic patients to tolerate the drug without fear of
esophageal inflammation and/or bleeding.
Declaration of interest
The authors state no conflict of interest and have received no
payment in preparation of this manuscript.

Bibliography
Papers of special note have been highlighted as 3. Cranney A. Treatment of 5. Porras AG, Holland SD, Gertz BJ.
either of interest () or of considerable interest postmenopausal osteoporosis. BMJ Pharmacokinetics of alendronate.
() to readers. 2003;327:355-6 Clin Pharmacokinet 1999;36:315-28

1. de Groen PC, Lubbe DF, Hirsch LJ,
et al. Esophagitis associated with the use
of alendronate. N Engl J Med
1996;335:1016-21
2. Olszynski WP, Davison KS. Alendronate
for the treatment of osteoporosis in men.
Expert Opin Pharmacother 2008;9:491-8
4. Zhou Y, Dial EJ, Doyen R, et al.
Effect of indomethacin on bile
acid-phospholipid interactions:
implication for small intestinal injury
induced by nonsteroidal
anti-inflammatory drugs. Am J Physiol
Gastrointest Liver Physiol
2010;298:722-31
6. Cryer B, Bauer DC. Oral
bisphosphonates and upper
gastrointestinal tract problems: what is
the evidence? Mayo Clin Proc
2002;77:1031-43
7. Watts N, Freedholm D, Daifotis A. The
clinical tolerability profile of alendronate.
Int J Clin Pract Suppl 1999;101:51-61

Expert Opin. Drug Deliv. (2013) 10(6) 745

 K. M. Hosny et al.
8. Peters ML, Leonard M, Licata AA. Role
of alendronate and risedronate in
preventing and treating osteoporosis.
Cleve Clin J Med 2001;68:945-51
.. A good comparative study for
alendronate and risedronate.
9. Cenni E, Granchi D, Avnet S, et al.
Biocompatibility of poly(D,L-lactide-co-
glycolide) nanoparticles conjugated with
alendronate. Biomaterials
2008;29:1400-11
10. Han HK, Shin HJ, Ha DH. Improved
oral bioavailability of alendronate via the
mucoadhesive liposomal delivery system.
Eur J Pharm Sci 2012;46:500-7
Expert Opin. Drug Deliv. Downloaded from informahealthcare.com by 109.83.111.141 on 05/28/13
. Revealed the importance of
mucoadhesive liposomes to improve
ALS bioavailability.
11. Katsumi H, Liu S, Tanaka Y, et al.
Development of a novel self-dissolving
microneedle array of alendronate, a
nitrogen-containing bisphosphonate:
evaluation of transdermal absorption,
safety, and pharmacological effects after
application in rats. J Pharm Sci
2012;101:3230-8
For personal use only.
12. Sulkowski WW, Pentak D, Nowak K,
et al. The influence of temperature,
cholesterol content and pH on liposome
stability. J Mol Struct 2005;744:737-47
13. Xia SQ, Xu SY, Zhang XM.
Optimization in the preparation of
coenzyme Q(10) nanoliposomes. J Agric
Food Chem 2006;54:6358-66
14. Sonaje K, Chen YJ, Chen HL, et al.
Enteric-coated capsules filled with
freeze-dried chitosan/poly(gamma-
glutamic acid) nanoparticles for oral
insulin delivery. Biomaterials
2010;31:3384-94
15. Vincourt V, Nguyen L, Chaumeil JC,
et al. Freeze-drying of ATP entrapped in
cationic, low lipid liposomes.
Cryobiology 2010;60:262-70
16. Al Deeb SK, Hamdan II, Al Najjar SM.
Spectroscopic and HPLC methods for
the determination of alendronate in
tablets and urine. Talanta
2004;64:695-702
. Simple method for analysis of ALS by
UV and HPLC.
746
17. Meyyanathan SN, Muralidharan S,
Rajan S, et al. A Simple sample
preparation with HPLC--UV method for
estimation of amlodipine from plasma:
application to bioequivalence study.
Open Chem Biomed Method J
2008;1:22-7
18. Valliappan K, Kannan K, Sivakumar T,
et al. Enantiospecific pharmacokinetic
studies on ketoprofen in tablet
formulation using indirect chiral HPLC
analysis. J Appl Biomed 2006;4:153-61
19. Yang Z, Gao S, Wang J, et al.
Enhancement of oral bioavailability of
20(S)-ginsenoside Rh2 through improved
understanding of its absorption and
efflux mechanisms. Drug Metab Dispos
2011;39:1866-72
20. du Plessis J, Ramachandran C,
Weiner N, et al. The influence of lipid
composition and lamellarity of liposomes
on the physical stability of liposomes
upon storage. Int J Pharm
2007;127:273-8
21. Hathout RM, Mansour S, Mortada ND,
et al. Liposomes as an ocular delivery
system for acetazolamide: in vitro and
in vivo studies. AAPS PharmSciTech
2007;8:1-12
22. Srinath P, Vyas SP, Diwan PV.
Preparation and pharmacodynamic
evaluation of liposomes of indomethacin.
Drug Dev Ind Pharm 2000;26:313-21
23. Nagarsenker MS, Londhe VY,
Nadkarni GD. Preparation and
evaluation of liposomal formulations of
tropicamide for ocular delivery.
Int J Pharm 1999;190:63-71
24. Hosny KM. Preparation and evaluation
of thermosensitive liposomal hydrogel for
enhanced transcorneal permeation of
ofloxacin. AAPS PharmSciTech
2009;10:1336-42
25. Gruner SM. Materials properties of
liposomal bilayers. In: Ostro MJ, editor.
Liposomes from biophysics to
therapeutics. Marcel Dekker, New York;
1997. p. 1-38
26. Benech RO, Kheadr EE, Laridi R, et al.
Inhibition of Listeria innocua in cheddar
cheese by addition of nisin Z in
liposomes or by in situ production in
Expert Opin. Drug Deliv. (2013) 10(6)
mixed culture. Appl Environ Microbiol
2002;68:3683-90
27. Huebner S, Battersby BJ, Grimm R,
Cevc G. Lipid-DNA complex formation:
reorganization and rupture of lipid
vesicles in the presence of DNA as
observed by cryoelectron microscopy.
Biophys J 1999;76:3158-66
28. Kachar B, Fuller N, Rand RP.
Morphological responses to
calcium-induced interaction of
phosphatidylserine-containing vesicles.
Biophys J 1986;50:779-88
29. Gulati M, Grover M, Singh M, Singh S.
Study of azathioprine encapsulation into
liposomes. J Microencapsul
1998;15:485-94
30. Xia S, Xu S, Zhang X. Optimization in
the preparation of coenzyme
Q10 nanoliposomes. J Agric Food Chem
2006;54:6358-66
31. Guo JX, Ping QN, Chen Y.
Pharmacokinetic behavior of cyclosporin
A in rabbits by oral administration of
lecithin vesicle and sandimmun neoral.
Int J Pharm 2001;216:17-21
32. Chen Y, Lu Y, Chen J, et al. Enhanced
bioavailability of the poorly water-soluble
drug fenofibrate by using liposomes
containing a bile salt. Int J Pharm
2009;376:153-60
Affiliation
Khaled Mohamed Hosny†1,2,
Osama Abdelhakim Aly Ahmed1,3 &
Rana Tariq Al-Abdali4
†
Author for correspondence
1
King Abdulaziz University, Faculty of Pharmacy,
Department of Pharmaceutics, Jeddah,
Saudi Arabia
Tel: +966592722634; Fax: +96626951696;
E-mail: Elswaify2000@yahoo.com
2
Beni Sueif University, Faculty of Pharmacy,
Department of Pharmaceutics and Industrial
Pharmacy, Beni Sueif, Egypt
3
Minia University, Faculty of Pharmacy,
Department of Pharmaceutics and Industrial
Pharmacy, Minia, Egypt
4
King Abdulaziz University,
Faculty of Pharmacy, Jeddah,
Saudi Arabia