24 International Journal of Pharmaceutical and Clinical Science 2013; 3(4): 24-28

ISSN 2277– 7202

Original Article

STUDY OF THE SPECTRAL PROPERTIES OF INCLUSION COMPLEX OF ASPIRIN

WITH HYDROXY PROPYL β-CYCLODEXTRIN

Pushpa Rajagopalan1 and Priya Penial

2

1. Dr. Pushpa Rajagopalan- Corresponding author

Associate Professor, Department of Chemistry, Sarah Tucker College,

Manonmaniam Sundaranar University,Tirunelveli-627007, Tamil Nadu, India.

E-mail: pushpa_chem@yahoo.co.in; ph: 9442468914 2. T.Priya Penial, co author

Assistant Professor,

Department of Chemistry, Sardar Raja College of Engineering,

Alangulam, Tirunelveli- 627808.

Received 04 December 2013; accepted 14 December 2013

Abstract

Aspirin (Acetyl Salicylic Acid) is a non steroidal, anti- inflammatory drug, used as an analgesic and an antipyretic.

Cyclodextrins and modified Cyclodextrins are used as complexing agents to increase the aqueous solubility, bioavailability

and stability of the drugs. Inclusion complex formation of aspirin with Hydroxy propyl β- Cyclodextrin (HPβ-CD) resulted

in stability and solubility enhancement of the drug. Complexation was proved by UV-Vis, FT-IR, 1H NMR and XRD

studies. The interaction of aspirin with HP β-CD is conducive to the formation of inclusion complexes in aqueous as well

as in the solid state. An analysis was made on the binding behaviour of aspirin with HP β-CD and the mode of inclusion of

the guest molecule into the host cavity was also envisaged.

© 2013 Universal Research Publications. All rights reserved

Key words: Cyclodextrins, Aspirin, inclusion complexes, β-CD and HP β-CD

1. Introduction

Cyclodextrins are cyclic (α-1, 4) linked oligosaccharides of

α-D-glucopyranose, containing relatively hydrophobic

central cavity and hydrophilic outer surface [1]. Owing to

lack of free rotation about the bonds connecting the

glucopyranose units, the Cyclodextrins are not perfectly

cylindrical molecules but the torroidal or cone shaped.

Based on this architecture the primary hydroxyl groups are

located on the narrow side of the cone shape, while the

secondary hydroxyl groups are located on the wider edge.

Various physico-chemical properties of the organic guest

molecules are altered in presence of Cyclodextrins with

enhanced selectivity, photo reactivity and stability [2, 3, 4].

This unique ability finds applications in various fields such

as pharmaceuticals [5, 6, 7], agriculture [8], cosmetics [9,

10], food [11], drug delivery [12] and industries [13, 14,

15].

During the past two decades, Cyclodextrins and their

derivatives have been of considerable interest in the

pharmaceutical field because of their potential to form

complex formulation. The hydrophobic cavity of

Cyclodextrins is capable of trapping a variety of molecules

within to produce inclusion complexes. Many advantages

of drug- complexation with Cyclodextrins have been

reported in scientific literature which includes increased

solubility, enhanced bioavailability, improved stability,

masking of bad test or odour, reduced volatility,

transformation of liquid or gas into solid form reduced side

effect and the possibility of a drug release system, etc.[16]

Although the solubility of β-CD is smaller, the size of its

cavity is more appropriate to encapsulate a great variety of

molecules with biological and pharmaceutical properties.

Complexation with β-CD enhances the solubility and

permeability of aspirin. Also the pharmacological activity

like analgesic and anti-inflammatory activities are also

enhanced [17]. It also has as an anti platelet or anti-clotting

effect and is used in long term low doses to prevent heart

attack, strokes and blood clot formation in people at high

risk for developing blood clots[18]. Orally administered

aspirin requires high and frequent dosing because it

undergoes extensive pre systematic metabolism. Also

chronic oral aspirin is associated with serious

gastrointestinal side-effects. Complexation with CD

alleviates the side effects to some extent. The

bioavailability and solubility of aspirin has to be increased

to overcome the side-effects of aspirin related to stomach

and gastro intestinal tract (GTI). Though aspirin complexes

with native CDs have been well documented, very little

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Universal Research Publications. All rights reserved

25 International Journal of Pharmaceutical and Clinical Science 2013; 3(4): 24-28

work has been reported with modified CDs. Hence in this

study it is proposed to investigate the spectral properties of

Aspirin complex with Hydroxy propyl β-CD (HP β-CD)

and to analyze how it modifies the stability and solubility

of the drug. This formulation would open new vistas in the

field of drug delivery.

2. Materials and Methods

Aspirin was synthesised in the laboratory by the reaction of

salicylic acid and acetic anhydride using concentrated

sulphuric acid as catalyst. β-Cyclodextrin was purchased

from Aldrich and HP-β-CD was purchased from Spectro-

chem. These chemicals were used as received. NaOH and

HCl used for buffer solutions were all Merck samples.

Double distilled water was used throughout the study.

2.1 Preparation of 1:1 Complex of Aspirin and HP-β-CD

A known quantity of HP-β-CD was taken in a reaction tube

and stirred magnetically for half an hour with small

quantity of water. Then an equimolar amount of aspirin was

added and stirred magnetically for 24 hours. The solid

complex obtained was dried using Wattman-3 filter paper

and kept in an air oven at 50◦C for 1 hour. The dried

complex was collected for further studies.

2.2 Preparation of stock solutions for UV studies

Stock solution of the substrate (1 x 10-3

M) was prepared

by weighing a known amount of the substrate and

dissolving it in minimum amount of doubly distilled water.

Appropriate CD (1 x 10-3

M) stock solutions in water were

prepared. A known volume of the substrate, buffer and CD

solution were mixed and diluted to 10 ml. These solutions

were stirred for 24 hrs. Absorption spectra were recorded to

calculate the binding constants of the complexes.

2.3 Calculation of Binding Constants

From the linear plots obtained using Benesi-Hildebrand

equation in MS Excel, binding constant of the 1:1 complex

between Aspirin and β-CD is calculated from the slope and

intercept. The Benesi-Hildebrand equation is

[CD][Substrate]/ΔOD = [CD] + [Substrate]/Δε + KfΔε

The slope and intercept are evaluated from the linear plots

drawn using MS Excel and the binding constant is

calculated using the formula, Kb = slope/intercept

2.4 Instrumentation methods

Fourier Transformed Infrared (FT-IR) spectra of complexes

were taken with Thermo Nicolet, Avatar 370 FTIR

Spectrophotometer between 4000-400 cm -1 using pressed

pellet technique. Proton Nuclear Magnetic Resonance (1H

NMR) Spectra of complexes of drug with CDs were taken

at 25°C by a Bruker Avance III, 400MHz X with Z-

gradient digital Nuclear Magnetic Resonance Spectrometer

operating at a proton frequency BBO 400MHz using

DMSO-d6 as a Solvent. Systronics-2201 UV-Visible

spectrophotometer was used for recording UV-Visible

spectra.

3. Results and Discussion

The physicochemical properties of the guest molecules

vary upon the formation of the inclusion complexes with

CDs. The orientation of the substrates inside the CD cavity

is governed by the size, substituents, mode of inclusion of

the substrate and also by the interaction between the

substrate and CD. Hence UV, FT-IR, XRD and 1H NMR

techniques were used to study the incorporation of the

guest into the CD cavities. All characterization studies

confirmed the inclusion of aspirin into the CD cavity

forming inclusion complex with it. The gradual increase of

absorbance in the UV spectra with increase in CD

concentration is the evidence for the inclusion and

stabilization of aspirin in the CD cavity. FT-IR spectra of

aspirin and its CD complexes showed different peaks of

varying intensities, confirming complexation between the

two. The NMR spectra of the CD complexes of aspirin

showed significant changes in the chemical shifts of H3

and H5 protons of CD inner cavity revealing inclusion of

the drug molecule in the CD cavity.

3.1 UV/Visible - absorption spectra

UV/Visible - absorption spectra were recorded at two

different pH values 2.5 and 12.5, using buffers. There is an

increase in absorbance with increasing CD concentration

evidencing the inclusion of aspirin more and more into the

CD cavity and its stabilization in the CD cavity. There was

also a red shift by 2nm due to complexation. At lower pH

blue shift and at higher pH red shift were noticed as the

concentration of CD was increased with HPβ-CD. Figures

1 and 2 show the UV spectra of aspirin with HPβ-CD at pH

2.5 and 12.5 respectively. Also the absorption intensities

were increased by increasing the concentration of CD. A

slight red shift could also be noticed. Apparent binding

constants (Kb) were calculated from Benesi-Hildebrand

plots of [G][H]/∆A Vs [G]+[H] shown in figures 3 and 4.

Fig1 UV Spectrum of aspirin and HPβ-CD at pH2.5;

[Guest] =1x10-3

M [Host] =1x10-3

M

Fig2 UV Spectrum of aspirin and HPβ-CD at pH 12.5;

[Guest]=1x10-3

M [Host]=1x10-3

M

Fig 3 Benesi-Hildebrand plot for aspirin- HPβ-CD at pH

2.5([H][G]/∆A vs [H]+[G])

26 International Journal of Pharmaceutical and Clinical Science 2013; 3(4): 24-28

Fig 4 Benesi-Hildebrand plot for aspirin- HPβ-CD at pH

12.5([H][G]/∆A vs [H]+[G])

The binding constant values with HPβ-CD under different

pH conditions were calculated as slope/intercept=Kb have

been tabulated in Table 1.

Table 1 Binding constants (Kb) of complexes of Aspirin

with CD at pH = 2.5 and pH =12.5 from UV data

pH Kb (M

-1) of complexes of

Aspirin in HP β-CD

Neutral

2.5

12.5

14286

2500

5000

Many interesting conclusions can be drawn from stability

studies of aspirin with CDs. Binding constant values show

that aspirin formed stable complexes with HPβ-CD

(kb=14286) under neutral conditions than with buffers. The

binding constants are higher at pH 12.5 compared to pH 2.5

reveals that the inclusion complexes are more stable in

alkaline solution.

3.2. Characterization by FT-IR spectroscopy

Figures 5 and 6 represent the FT-IR Spectra of Aspirin and

1:1 complex of Aspirin- HPβ-CD respectively. The FT-IR

spectrum of 1:1 complex of Aspirin- HPβ-CD shows the

characteristic frequencies of Aspirin having undergone

changes which are an evidence of complexation between

the two. Aspirin has two carbonyl chromophores. This

compound has an ample of scope for intramolecular

Hydrogen bonding as well intermolecular Hydrogen

bonding with the primary hydroxyl groups on the narrower

rim of the torroidal shape of HPβ-CD and with the

secondary hydroxyl groups on the wider rim. Hence this

facility provides good and comfortable inclusion of the

drug into the CD cavity.

Fig 5 FT-IR Spectrum of Aspirin

Fig 6 FT-IR Spectrum of Aspirin - HPβ-CD 1:1 complex

Table 2 FT-IR frequencies (cm-1

) of pure Aspirin and its complex with HP–β CD

Functional Groups Free Aspirin Complex of aspirin- HP β-CD

Carbonyl OH stretching 2698.29 2872.83

Vinyl ester C=O 1754.75 1753.61

Aromatic acid C=O 1688.93 1690.49

Aromatic C=C stretching 1605.24,1574.701484.58,1458.30 1605.35,1575.07, 1484.64, 1458.21

C-O(acid Ester) 1220.18, 1186.75 1220.16

Ortho substituted phenyl C-H bending 754.42 754.90

Table 2 provides the frequencies of aspirin in Free State

and in the complexed state. The IR spectrum of aspirin

reveals the presence of a peak at 2698.29 cm-1

, assigned to

Carbonyl –OH stretching vibration and one at 1754.75 cm-1

corresponding to the vinyl ester carbonyl group and

at1688.93 cm-1

corresponding to aromatic acid carbonyl

group. Upon complexation, the carbonyl band has slightly

shifted towards a lower wave number, due to the

appearance of host-guest interactions. These suggest the

possibility of formation of hydrogen bonds between the

hydroxyl groups of the host cavities and the aspirin

carbonyl group. This peak shifting towards lower frequency

with change in intensity suggests a change in environment

of carbonyl group associated with ester moiety. Slight

shifting of absorption band for carbonyl groups to a lower

frequency can be attributed to breakdown of intramolecular

hydrogen bonds associated with the drug molecule and

formation of intermolecular hydrogen bonding of the drug

with CDS.

3.3 Proton Nuclear Magnetic (1H NMR) Resonance

Spectroscopy

Applications of NMR on CD chemistry are so important

that no other spectroscopic technique can provide the same

wealth of chemical information on the supramolecular

systems. Figures 7-8 depict the NMR spectra of Aspirin, ,

1:1 complex of Aspirin with HP β-CD. In CD molecule,

hydrogen atoms are located in interior of cavity (H3 and

H5) and outer surface cavity (H1,H2,H4, and H6).When

27 International Journal of Pharmaceutical and Clinical Science 2013; 3(4): 24-28

any guest molecule gets incorporated in CD cavity,

hydrogen atoms located inside cavity experience significant

changes in δ ppm( parts per million) values. But in case of

association of guest molecule with CD hydrogen atoms on

exterior surface show smaller shifts in δ ppm values .Thus a

positive sign of Δδ ppm shows a downfield displacement

and a negative sign an upfield displacement. Table 3 shows

the changes in the chemical shift values of HP β-CD in the

free State and in the 1:1 Complex of HPβ-CD – aspirin.

Table 3 shows the changes in the chemical shift values of

β-CD in the free state and in the 1:1 Complex of HPβ-CD –

aspirin. Table 4 shows the changes in the chemical shift

values of Aspirin in the free State and in the 1:1 Complex

of HPβ-CD – aspirin.

Table 3 Chemical shifts (δ) of protons of HPβ-CD in free

host and inclusion complex

HP β-CD

protons

HPβ-CD

( ppm)

HPβ-CD - aspirin complex

( ppm)

H4 3.51(d) 3.326(m)

H2 3.646(d) 3.631(m)

H5 3.77(m) 4.829(d)

H6 3.80(s) 3.81(d)

H3 3.938(d) 5.674(m)

H1 5.086(d) 5.1(s)

δ- Chemical shift in parts per million H3, H5- Protons in

the interior of CD cavity H1, H2, H4, H6- Protons on the

outer surface of CD t-triplet; d- doublet; s- singlet; m-

multiplet

Table 4 Chemical shifts (δ) of protons of aspirin in free

guest and inclusion complex

Aspirin ( ppm) HP β-CD Aspirin complex ( ppm)

12.917(s) 13.027(s)

7.935(m) 7.933(m)

7.638(m) 7.637(m)

7.378(m) 7.377(m)

7.2(m) 7.2(m)

2.243(s) 2.242(s)

Fig 7

1H NMR of Aspirin

3.4 X – Ray diffraction Studies

Free aspirin is a crystalline solid. The XRD pattern of the

inclusion complexes of the drug with HP β-CDs as shown

in figures 9-10 reveals the presence of both guest and host

in the complex, the diffractogram of the CDs dominating

that of the guest confirming inclusion process. The XRD

pattern of aspirin contained a number of sharp peaks which

is indicative of its crystallinity. The diffraction pattern of

the complex is quite different from pattern of the guest and

CD which shows the sum of both the patterns. In XRD of

the complex, most of the characteristic peaks of aspirin

disappeared and some were reduced in intensity. The

changes are most prominent in the complex with β-CD

which indicates a successful inclusion of aspirin in β-CD.

The sharp peaks of complex confirmed its crystalline

nature. Figures 7 and 8 show the XRD patterns of aspirin

and aspirin –HP β-CD complex.

Fig 8

1H NMR of 1:1 complex of Aspirin – HP β-CD

Fig 9 XRD patterns of Aspirin

Fig 10 XRD patterns of Aspirin –HP β-CD complex

4. Conclusions The inferred data from UV-Visible, NMR, FT-IR spectra

and XRD analysis serve as a proof for the formation of

inclusion complex between HP β-CD and aspirin. The

stability studies show that HP β-CD provides the most

favourable environment for inclusion complex formation

with aspirin. Therefore, complexes of aspirin with CDs

would certainly show improvement in bioavailability of

aspirin. The CD complexes of aspirin showing an increased

28 International Journal of Pharmaceutical and Clinical Science 2013; 3(4): 24-28

solubility and stability, is certainly an advantage over the

free drug usage in the field of medicine. As the CDs are

prepared from starch by enzymic conversion, their safety

profile is also assured and this work may as well be

regarded as an important break-through in drug delivery.

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Source of support: Nil; Conflict of interest: None declared