International Journal of Pharmaceutical Sciences and Nanotechnology
Volume 3 • Issue 2 • July - September 2010
Research Paper
Development and Characterization of Aspirin-Phospholipid Complex for
Improved Drug Delivery
A. Semalty*, M. Semalty, D. Singh and M.S.M. Rawat
H.N.B. Garhwal University, Srinagar, India.
ABSTRACT:
Aspirin (acetylsalicylic acid) is one of the most widely used analgesic. Aspirin is poorly soluble in water
and causes gastrointestinal (GI) irritation. To improve the solubility (and hence the bioavailability) and minimize the GI
irritation, its complexes with soya-phospholipid-80 % (in 1: 1 molar ratio) were prepared in an organic solvent and
evaluated for solubility, drug content, scanning electron microscopy (SEM), FT-IR spectra, X ray diffraction, differential
scanning calorimetry (DSC) and in vitro dissolution study. Aspirin-phospholipid complex were found to be disc shaped with
rough surface in SEM. Drug content in the complex was found to be 95.6 %. DSC thermograms, XRD and FTIR confirmed
the formation of phospholipid complex. Solubility of the prepared complex was found to be improved. Aspirin complex and
aspirin showed 90.93 % and 69.42 % of drug release at the end of 10 h in dissolution study in pH 1.2 acid buffer. It was
concluded that the phospholipid complex of aspirin may be of potential use for improving the solubility of aspirin and hence
its bioavailability. The complexes may also reduce GI toxicity of the drug.
KEYWORDS: Pharmacosomes; Phospholipids complex; Aspirin; NSAIDs
Introduction
Aspirin (acetylsalicylic acid, ASA) is one of the most
widely used therapeutic substances due to its analgesic,
antipyretic and anti-inflammatory properties. Despite the
proliferation in development of new non-steroidal antiinflammatory drugs (NSAIDs), ASA remains one of the
most effective ‘over-the-counter’ drugs in the treatment of
rheumatic diseases. Furthermore, due to its anti-thrombotic
properties, ASA is now prescribed at low doses in the
prevention and treatment of cardiovascular diseases,
strokes and disorders associated with platelet aggregability
(Van and Botting, 1992).
ASA use is commonly associated with gastrointestinal
(GI) side-effects ranging from dyspeptic symptoms and
ulceration to life-threatening episodes of bleeding
(Hollander, 1994). ASA is poorly soluble in the acidic
conditions of the stomach, which can delay absorption of
high doses for 8 to 24 hours. Modifying the water solubility
of aspirin may prove to be beneficial for improving its
absorption.
The rate of release of a drug is a function of its intrinsic
solubility and influenced by particle size, crystallinity, drug
derivatization and formation of more-soluble complexes
* For correspondence: A. Semalty,
E-mail: [email protected]
940
(Leuner and Dressmann, 2002; Rabinow, 2004 ; Patravale
et al.. 2004; Kesisoglon et al. 2007; Nijlen 2003;
Various approaches have been
Nokhodchi, 2005).
investigated to improve the absorption and permeation of
biologically active constituents of synthetic and natural
origin. These include, development of more soluble prodrug, solid dispersions, preparing complexes with
complexing agents like metals, cyclodextrin and
phospholipids. Apart from other methods used for
modifying the solubility, the complexation with
phospholipids have been found to show improvement in
both, absorption as well as permeation of the active
constituents (Semalty et al., 2009a; Semalty et al., 2009b).
Phospholipids (like phosphatidylcholine) play a major
role in drug delivery due to its amphiphilic nature, that can
modify the rate of drug release for the enhancement of drug
absorption across biological barriers. Developing of
amphiphilic drug-lipid complexes or pharmacosomes may
prove to be a potential approach for improving therapeutic
efficacy of the drugs by improving the bioavailability
(through improvement in solubility in GI fluid and
permeation across the biomembranes). Any drug
possessing an active hydrogen atom (-COOH, -OH, -NH2,
etc.) can be esterified to the lipid, with or without spacer
chain. Synthesis of such compound may be guided in such
Semalty et al. : Development and Characterization of Aspirin-Phospholipid Complex for Improved Drug Delivery
a way that strongly results in an amphiphilic compound,
which will facilitate membrane, tissue, or cell wall transfer,
in the organism. Pharmacosomes are defined as colloidal
dispersions of drugs covalently bound to lipids, and may
exist as ultrafine vesicular, micellar, or hexagonal
aggregates, depending on the chemical structure of the drug
lipid complex. The salient features of pharmacosomes are,
increased entrapment efficiency, easy removal of
unentraped drug from the formulation, no loss of drug due
to leakage, no problem of drug incorporation and no
influence of encaptured volume and drug-bilayer
interaction on entrapment efficiency (Semalty et al., 2009a;
Biju et al., 2006). These amphiphilic drug-lipid complexes,
are stable and more bioavailable with low interfacial
tension between the system and the GI fluid, thereby
facilitating membrane, tissue, or cell wall transfer, in the
organism.
Moreover, the incorporation of NSAIDs with
phospholipids has been also suggested to improve GI safety
of these drugs. It has been reported that the diffusion of
NSAIDs across lipid membranes and into target cells is
accelerated when it is present as a complex with
phosphatidylcholine (PC) (Lichtenberger et al., 1995).
Therefore, developing the drugs as lipid complexes
(pharmacosomes) may prove to be a potential approaches
to improve solubility and to minimize the GI toxicity of
NSAIDs.
Therefore, this study aims to develop the amphiphilic lipid
complexes or pharmacosomes of aspirin to improve
solubility and hence its dissolution. The study deals with its
characterization for drug content, solubility, crystallinity
(XRD), chemical interaction (FTIR), phase transition
behaviour (DSC) and in vitro dissolution study.
Materials and Methods
Materials
Aspirin was purchased from E-Merck Mumbai. Soya
phosphatidylcholine (LIPOID S-80) was obtained as a gift
sample from LIPOID Gmbh Germany. All other chemicals
were of analytical grade.
Methods
Preparation of Aspirin- PC Complex
Aspirin-PC complex was prepared by associating aspirin
with an equimolar concentration of PC (80 % purity grade
of soya-phospholipids). The equimolar concentration of PC
(in 1: 1 molar ratio) and aspirin were placed in a 100 mL
round bottom flask and dissolved in dichloromethane. The
solvent was evaporated off under vacuum at 40 oC in a
941
rotary vacuum evaporator (Perfit Modle No. 5600 Buchi
type). The dried residues were collected and placed in
vacuum desiccators overnight and then subjected to
characterization.
Drug content
To determine the drug content in Aspirin-PC complex,
complex equivalent to 100mg were weighed and added in
100mL of pH 1.2 hydrogen chloride buffers. The
volumetric flask was stirred continuously for 24hr on a
magnetic stirrer. Dilutions were made suitably and
measured for the drug content at 228.3nm UV
spectrophotometrically by Lambda25 Perkin Elmer
UV/Visible Spectrophotometer.
Solubility
To determine the change in solubility due to complexation,
solubility of aspirin and aspirin-PC complex was
determined in pH 1.2 HCl buffer and n-octanol by shake
flask method. 50 mg of aspirin (and 50 mg equivalent in
case of complex) was taken in a 100 mL conical flask.
50mL of pH 1.2 HCl buffer was added and then stirred for
15 minutes. The suspension was then transferred to 250mL
separating funnel with 50mL of n-octanol and was shaken
well. Separating funnel was allowed to stand for about 30
minutes. Concentration of the drug was determined from
the aqueous layer spectrophotometrically at 228.3nm.
Scanning electron microscopy (SEM)
To detect the surface morphology of the pharmacosomes,
SEM of complex was performed at UGC-DAE consortium
Indore (India) by Scanning Electron Microscope (JEOL
JSM 5600).
Infrared Spectroscopic Analysis
FTIR spectra for the various powders were obtained on a
Perkin Elmer FTIR spectrometer (Perkin Elmer Life and
Analytical Sciences, MA, USA) in the transmission mode
with the wave number region 4,000- 500 cm-1. KBr pellets
were prepared by gently mixing 1 mg sample powder with
100mg KBr. The results were shown in Fig. 2.
Differential Scanning Calorimetry (DSC)
Thermograms of Aspirin, phosphatidylcholine (80%), and
aspirin-PC complex were recorded using a 2910 Modulated
differential scanning calorimeter V4.4E. The thermal
behavior was studied by heating 2.0±0.2 mg of each
individual sample in a covered sample pan under nitrogen
gas flow (Fig. 3). The investigations were carried out over
the temperature range 25-250º with a heating rate of
10ºC/min.
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International Journal of Pharmaceutical Sciences and Nanotechnology
Volume 3 • Issue 2 • July-September 2010
Fig. 1 SEM of phospholipid complex of aspirin (a) 750 X magnification (b) 85 X magnification.
X-ray Powder Diffraction Analysis
The crystalline state of aspirin in the different samples was
evaluated with X-ray powder diffraction. Diffraction
patterns were obtained on a Bruker Axs- D8 Discover
Powder X-ray diffractometer (Germany). The X-ray
generator was operated at 40 KV tube voltages and 40 mA
of tube current, using the Ka lines of copper as the
radiation source. The scanning angle ranged from 1 to 60o
of 2θ in step scan mode (step width 0.4 o/min). Aspirin,
phosphatidylcholine
(80%),
and
aspirin-PC
phosphatidylcholine complex were analyzed with X-ray
diffractions. The results were shown in Fig. 4.
Dissolution Study
In vitro dissolution studies for aspirin complex as well as
plain aspirin were performed in triplicate in a USP XXIII
six station dissolution test apparatus (Veego Model
No.6DR) at 100 rpm and at 37 oC. An accurately weighed
amount of the complex equivalent to 100 mg of aspirin
was put into 900 ml pH 1.2 HCl buffer. Samples (three ml
each) of dissolution fluid were withdrawn at different
intervals and replaced with the equal volume of fresh
media, to maintain sink conditions. Withdrawn samples
were filtered (through a 0.45 mm membrane filter) and
diluted suitably and then analyzed spectrophotometrically
at 228.3nm.
Results and Discussion
In the present experiment aspirin–phospholipid complex
(pharmacosomes) were prepared by a simple and
reproducible method.
Content of aspirin in phospholipid complex, as
estimated by UV spectrophotometry, was 95.6% (w/w).
Pharmacosomes showed a high percentage of drug loading,
which is peculiar to them. Complexation is giving a good
percent loading of the drug that makes the delivery of drug
clinically feasible.
Solubility of the aspirin complex was found to be much
higher (in water and n octanol) than the aspirin. Table. 1
provides the solubility data. The increase in solubility of
aspirin in the complex can be explained by the
solubilization resulted from the formation of micelle in the
medium and by the amorphous characteristics of the
complex. As an amphiphilic surfactant, phospholipids
could increase the solubility of the drug by the action of
wetting and dispersion. Unlike non-polar nature of aspirin,
the complex showed an amphiphilic nature, which in turn
may prove to be responsible for improved bioavailability
of the drug.
Table 1 Solubility study of aspirin and its complex.
Drug
Statistical analysis
Results are expressed as mean values and standard
deviations (± SD) and the significance of the difference
observed was analyzed by the Student’s t test. In all tests, a
probability value of P<0.05 was considered statistically
significant.
Solubility in
aqueous layer
(in µg/ mL)*
Solubility in nOctanol layer
(in µg/ mL) *
Aspirin
8.45 ± 1.20
24.54 ± 2.46
Aspirin-PC complex
15.98 ± 1.98
31.44 ± 1.86
* Data expressed as mean values and standard
deviations (± SD); n=3
Semalty et al. : Development and Characterization of Aspirin-Phospholipid Complex for Improved Drug Delivery
943
69.0
866.1
65
800
60
750
700
55
650
50
600
45
550
40
500
522.61
%T
%T 450
1417.84
2740.12
2648.60
3051.81
2622.55
2596.54
400
350
1292.74
748.24
773.21
452.50
757.61
424.61
35
423.44
30
25
300
200
150
3397.85
20
2956.47
2850.33
2918.04
1960.36
1637.83
1744.72
896.54
876.65
843.14
824.34
1104.18
1548.98
3267.22
100
619.19
720.74 593.39
647.98
691.55 554.13
15
5
972.78
1181.87
1220.38 1062.14
3000
2000
1371.18
1605.34 1454.70
1697.75
1306.56
1756.01
10
1363.95
50
4000.0
0.0
562.20
542.72
644.40 515.21
476.23
502.03
1314.17
1485.04
1472.16
1582.28
1571.00
250
1500
2832.25
1000
500 400.0
cm-1
4000.0
0.3
3000
2000
1500
598.36
755.06
969.46
665.90
704.09
916.62
839.74
1012.49
803.61
1093.86
1000
500 400.0
cm-1
(a) Aspirin
(b) PC-80
27.0
36.4
26
34
24
32
30
22
28
20
26
18
24
%T
22
16
20
%T 14
18
12
16
10
14
12
8
766.10
10
6
8
1254.52
6
4
1093.08
1740.25
1734.84
4
2
4000.0
0.8
3000
2000
1457.10
1500
1057.71
2
1000
500 400.0
cm-1
(c) APC-80 (complex)
4000.0
0.0
3000
2000
1500
1000
500 400.0
cm-1
(d) Physical mixture
Fig. 2 FTIR spectra of Aspirin, PC-80, phospholipid complex of Aspirin (APC 80) and their Physical mixture.
Scanning Electron Micrographs of the complex are
shown in Fig. 1. Pharmacosomes were found to be of disc
shaped with rough surface morphology. Complexes were
found to be as free flowing particles. As the phospholipids
are natural component their different purity grades may
have different effects in shape and surface morphology.
The surface was found to be sticky in the complexes
prepared with low purity grades (40 %) of phospholipids.
On the other hand the surface of the complexes prepared
with the high purity grades of phospholipids (80 %) show
rough, non-sticky and free flowing nature as in the present
study (Semalty et al., 2009b).
The formation of the complex can be confirmed by the
IR spectroscopy comparing the spectrum of the complex
with the spectra of the individual components and their
mechanical mixtures (Semalty et al., 2009a). FT-IR spectra
for various powders were obtained on a Perkin Elmer FTIR
spectrometer in the transmission mode with the wave
number region 4,000-500 cm-1. FTIR spectra showed the
changes in peaks in complexes and positions from that of
aspirin and PC. FT-IR spectra of complex were
significantly different from that of components and that of
physical mixtures (Fig. 2).
Aspirin showed the characteristic IR (KBr) peaks of
O-CO- stretching at 1756.01, -COO at 1697.75, -OH
bending at 1454.70 and -OH bending at 916 cm-1. The OCO- stretching vibrations at 1697.75 cm-1 in aspirin has
been shifted to higher wave number side in the complex
with medium absorption band. –OH (out of plane) bending
at 916.62 cm-1 in aspirin has merged with broadband at
1057.71 cm-1. –OH (in plane) bending at 1371.18 cm-1 is
missing in the complex. The –CN stretching in the
phosphatidylcholine at 1238 cm-1 (strong band) has shifted
to 1240 cm-1 (broadband) in the complex. Thus the FTIR
spectra indicate the interaction of PC with the aspirin’s –
COOH group.
The differential scanning calorimetry is a tool used to
measure the temperature and energy variation involved in
the phase transitions, which reflects the degree of
crystallinity and stability of the solid state of
pharmaceutical compounds (Sang et al., 2003). The peak
size and shape of the DSC curves are useful in determining
the crystallinity of the drug and the carrier. In order to
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International Journal of Pharmaceutical Sciences and Nanotechnology
substantiate the association of aspirin with PC, DSC
analysis was performed on aspirin, PC, and the aspirin-PC
complex. The results of the DSC test confirmed the
Volume 3 • Issue 2 • July-September 2010
association of aspirin and PC in the complex as both peaks
representing aspirin acid and PC changed position (Fig. 3).
Fig. 3 DSC thermograms of (a) aspirin; (b) PC-80 and; (c) phospholipid complex of aspirin.
Semalty et al. : Development and Characterization of Aspirin-Phospholipid Complex for Improved Drug Delivery
PC showed two major peaks at 83.21o C and 107.90o C
and a small peak at 64.45o C, while aspirin showed a sharp
endothermic peak at 142.6o C. On the other hand complex
showed two unique peaks (at 50.8o and at 63.8o C), which
were different from the peaks of the individual components
of the complex. The DSC thermograms of the phospholipid
complexes of some phytoconstituents like silybin, puerarin,
curcumin and naringenin also revealed the similar results
(Yanyu et al., 2006; Li et al., 2007; Maiti et al., 2007;
Semalty et al., 2009c). In all these studies the thermogram
of the complex also exhibited a single peak which was
different from the peak of phytoconstituent and
phospholipids. It is evident that the original peaks of
phytoconstituent and phospholipids disappear from the
thermogram of complex and the phase transition
temperature is lower than that of phospholipids. The
acemetacin and indomethacin also showed the significant
changes in the DSC thermogram when these arylacetic acid
derivatives
were
complexed
with
the
1,2-dipalmitoylphosphatidylcholine (Lúcio et al., 2008).
To check whether the changes in the aspirin crystal
morphology correspond to a polymorphic transition and to
study the solid state of aspirin phospholipid complex,
XRPD analysis was conducted. From these patterns, the
degree of crystallinity could be evaluated using the relative
integrated intensity of reflection peaks in the given range of
reflecting angle, 2θ. The value of 2θ means the diffraction
angle of ray beams, which is shown in the abscissa of
Fig. 4.
The XRPD of aspirin complex revealed a broad peak
similar to PC indicating that the aspirin was in amorphous
form in phospholipid complex. The disappearance of
aspirin crystalline diffraction peaks confirmed the
formation of phospholipid complex. These results are well
supported by our previous studies done with the
phospholipid complexes of diclofenac and naringenin
(Semalty et al., 2009b; Semalty et al., 2009c).
The phospholipid complexes of puerarin, insulin and
salmon calcitonin also supported the results obtained (Li et
al., 2007; Shi et al., 2006; Cui et al., 2006; Yoo et al.,
2003). Unlike liposomes, bonding between drug and the
phospholipids in development of pharmacosomes (drugphospholipid complex), might have resulted into the
significant change of its X-ray diffraction.
Intensity(a.u)
25000
20000
15000
10000
5000
0
ASP Pure
500 10
400
300
200
100
0
15
10
15
160
140
120
100
80
60
40
20
0
945
20
25
30
35
40
45
50
55
60
55
60
55
60
PC80
20
25
30
35
40
45
50
APC80
10
15
20
25
30
35
40
45
50
2θ
Fig. 4 High resolution X ray powder diffraction (XRPD) study of Aspirin complex (APC-80) and its components.
946
International Journal of Pharmaceutical Sciences and Nanotechnology
The aspirin–phospholipid complex showed better
dissolution profile than the aspirin (Fig. 5). Unlike the free
aspirin (which showed a total of only 69.42 % drug release
at the end of 10 h) aspirin complex showed 90.93 % at the
end of 10 h of dissolution study in pH 1.2 acid (HCl)
buffer. The solid dissolution is a complex operation
influenced by a great number of factors, not only the
particle size. Differences in crystal habit, surface area,
surface energies, particle size and wettability may all play a
role in affecting the dissolution rate of powder (Jarmer et
al., 2005). Phospholipids being an amphiphilic surfactant,
increased the solubility of the drug by the action of wetting
and dispersion. And that’s why the dissolution profile of
the complex was found to be improved.
Volume 3 • Issue 2 • July-September 2010
poor water and/or lipid solubility) may also be developed
for improving their bioavailability.
Acknowledgments
Authors acknowledge the grant provided by the
Department of Science & Technology, Govt. of India for
the research work. Authors are thankful to LIPOID GmbH
Germany for providing the gift sample of
phosphatidylcholine for this work. Facilities provided by
the UGC-DAE Consortium for Scientific Research, Indore
(M.P.) and Department of Chemistry, University of Delhi
are thankfully acknowledged.
Percent Drug Release
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In the present study aspirin–phospholipid complex
(pharmacosomes) were prepared by a simple and
reproducible method. The physicochemical investigations
showed that aspirin formed a complex with phospholipids
with better solubility and dissolution profile. The
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for improving bioavailability. As the phospholipid
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