EFFECT OF IONIC CROSSLINKING AGENTS ON SWELLING BEHAVIOUR OF CHITOSAN HYDROGEL MEMBRANES Milena Pieróg, Magdalena Gierszewska-Drużyńska, Jadwiga Ostrowska-Czubenko Chair of Physical Chemistry and Physicochemistry of Polymers, Faculty of Chemistry Nicolaus Copernicus University ul. Gagarina 7, 87-100 Toruń, Poland e-mail:
[email protected]
Abstract
Physically crosslinked chitosan hydrogels were prepared by treating chitosan with sulfuric acid, trisodium citrate, sodium tripolyphosphate and sodium alginate, respectively. The chemical structure of modified chitosan hydrogels as well as unmodified one was analysed by FTIR spectroscopy and conclusions on the formation of the ionic crosslinks between protonated amino groups of chitosan and anionic functional groups of crosslinking agents were drawn out. The swelling behaviour of the membranes formed from modified and unmodified chitosan was studied in buffer solutions at various pH at 37 ºC. It was observed that the swelling degree of chitosan hydrogel membranes depends both on pH of buffer solution as well as on the type of crosslinking agent.
Key words: chitosan membrane, hydrogel, ionic crosslinking, swelling.
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1. Introduction In general, hydrogels are materials composed of three-dimensional hydrophilic polymeric network and water that fills the free spaces inside this network. Hydrogels are able to absorb and retain 10‑20% up to thousand times water or biological fluids than their dry weight. Hydrogels are called as “intelligent materials”, because they respond reversibly to slight changes of the properties of surrounding media. This ability causes that hydrogels found numerous applications in industry and pharmacy, for example as controlled drug release systems [1]. Recently much attention has been focused on the development of hydrogels based on natural, biodegradable and biocompatible polymeric materials such as chitosan. Chitosan is a deacetylated derivative of chitin. It is a copolymer of β‑(1→4)‑linked 2‑acetamido‑2‑de oxy‑β‑D‑glucopyranose and 2‑amino‑2‑deoxy‑β‑D‑glucopyranose (Figure 1) [2].
Figure 1. Chemical structure of chitosan (DDA-degree of deacetylation),
It is well known that pure chitosan hydrogel membranes exhibit a small resistance to acidic media ‑ they undergo dissolution or disintegration. To avoid this processes chitosan modification through crosslinking is widely used. Among all known chitosan crosslinking methods ionic crosslinking is the simplest and mildest one. In ionically crosslinked hydrogels a network is formed in the presence of negatively charged crosslinking agents (CAs), which form bridges between the positively charged chitosan polymeric chains. Both low molecular as well as high molecular ions can be used as CA [1, 3]. Among other significant properties of hydrogels the one of the most importance is their swelling and dehydration behaviour [3]. Schematic representation of hydrogel swelling and deswelling processes is shown in Figure 2.
Figure 2. Schematic representation of hydrogel swelling and deswelling processes
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The swelling behaviour of the ionic hydrogels is unique. It is affected by the ionization of functional groups along the polymer chains and the ionization of crosslinking agent molecules. Other factors that affect the swelling of these materials are: hydrophilicity of materials used in hydrogel network formation, degree of crosslinking, pH, ionic strength and nature of counterions of swelling medium [4]. In the present studies sulfuric acid, trisodium citrate, tripolyphosphate and sodium alginate were used as chitosan crosslinking agents. The chemical structure of uncrosslinked and ionically crosslinked chitosan hydrogel membranes were analysed by FTIR spectroscopy. The influence of chemical structure of crosslinking agent on swelling kinetics and equilibrium swelling degree of chitosan based hydrogel membranes were studied.
2. Materials and methods 2.1. Materials Commercially available chitosan (Ch) from crab shells of medium molecular weight, sodium alginate (NaAlg) and sodium tripolyphosphate (TPP) were purchased from Sigma Aldrich (Germany). Sulfuric acid (H2SO4) and trisodium citrate (CIT) were purchased from POCh (Gliwice). In all swelling experiments the following buffer solutions of constant ionic strength I = 0,145 M were used: hydrochloric buffer (pH 1.2), acetic buffer (pH 7.4) and TRIS buffer (pH 8.5). Acetic acid (HAc), sodium acetate (NaAc), sodium chloride (NaCl), hydrochloric acid (HCl) and sodium hydroxide (NaOH) were analytical grade and were purchased from POCh (Poland) and TRIS of analytical grade was purchased from Sigma Aldrich (Germany). Potassium bromide (KBr) for spectroscopy was purchased from Merck (Germany). 2.2. Polymer characterization Degree of deacetylation, DDA, of chitosan was determined by potentiometric titration method. The viscosity average molecular weight of chitosan and sodium alginate, Mv, was determined by viscometry. The details of above measurements have been presented elsewhere [5]. Degree of deacetylation was equal to 75.72 ± 3.82%. Mv was equal to 730 kDa for chitosan and 100 kDa for sodium alginate. 2.3. Membrane preparation Pure chitosan (Ch) membranes were prepared by casting solution and solvent evaporation technique. 1 wt.% chitosan solution prepared by dissolving chitosan powder in 2 wt.% acetic acid was cast as film on clean glass plate, evaporated to dryness at 37 ºC and further dried under vacuum at 60 ºC. Chitosan/sulfuric acid (Ch/H2SO4) membranes were prepared by immersing of pure chitosan membranes in 2 M sodium hydroxide solution for 5 min to remove of acetic acid residues, thoroughly washed with deionized water and air‑dried. Then the films were immersed in 0.5 M sulfuric acid solution at room temperature for 60 min. The crosslinked membrane was thoroughly washed with deionised water and dried as pure chitosan membrane.
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Chitosan/trisodium citrate (Ch/CIT) membranes were obtained by immersing of pure chitosan membranes in aqueous 5.0% (w/v) sodium citrate solution for 60 min. Following crosslinking conditions were applied: Tcrosslink. = 4 ºC, pH = 5.0 (initial solution acidified with HCl). After crosslinking membranes were repeatedly washed with deionised water and dried as pure chitosan membranes. Chitosan/sodium tripolyphosphate (Ch/TPP) membranes were prepared by immersing of pure chitosan membranes in aqueous 1.3% (w/v) phosphate solution for 60 min. Applied crosslinking conditions were as follows: Tcrosslink. = 4 ºC, pH = 5.5. Chitosan/sodium alginate (Ch/NaAlg) membranes were prepared by casting solution and solvent evaporation technique. 1 wt.% chitosan solution in 2 wt.% acetic acid and 1 wt.% sodium alginate solution were mixed in the 3 : 1 volume ratio, respectively (pHsolution mixture = 3.5). The crosslinked membranes were additionally thoroughly washed with deionised water and dried as pure chitosan films. 2.4. FTIR spectroscopy analysis FTIR spectra of chitosan and ionically crosslinked chitosan (ICCh) samples in KBr disc form were recorded on Perkin-Elmer 2000 FTIR spectrometer from 400 to 4000 cm‑1 with a resolution 4 cm‑1 and at 100 scans. Ch and ICCh polymers were thoroughly powdered and powders dried under vacuum at 60ºC for 24 hours before milling with anhydrous KBr. 2.5. Kinetics swelling experiments Ch and ICCh membranes ability to swell was determined by kinetics swelling experiments in acidic, inert and basic buffer solutions at constant ionic strength I = 0.145 M and at 37 ºC. Solution media of pH 1.2, pH 7.4 and pH 8.5 were used, simulate gastric fluid, plasma blood and small intestinal fluid, respectively. The swelling behaviour was measured by immersing a dry membrane sample of a known weight in buffer solution on definite time period. Membrane was weighed periodically after carefully drying its surface with a filter paper. The degree of swelling (S) was calculated according to the formula: S = (Wt - W0)/W0 where: W0 ‑ weight of the dry sample in g, Wt ‑ weight of the swollen sample in g at time t.
3. Results and discussion FTIR spectra of unmodified (Ch) and modified chitosan (ICCh) are shown in Figure 3. All spectra exhibit a strong and broad nonsymmetric band at about 3430 cm‑1 that results from overlapping of the O‑H and N‑H stretching vibrations of functional groups engaged in hydrogen bonds [6]. The spectrum of Ch exhibits characteristic absorption bands at 1656 cm‑1 (C=O stretching in amide group, amide I vibration), 1598 cm‑1 (N‑H bending in nonacetylated 2‑aminoglucose primary amine) and 1560 cm‑1 (N‑H bending in amide group, amide II vibration). Absorption bands at 1153 cm‑1 (antisymmetric stretching of the C‑O‑C bridge), 1083 cm‑1 and 1031 cm‑1 (skeletal vibrations involving the C‑O stretching) are characteristic of chitosan saccharide structure [7].
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Effect of Ionic Crosslinking Agents on Swelling Behaviour of Chitosan Hydrogel Membranes
Figure 3. FTIR spectra of chitosan and ionically crosslinked chitosan membranes After crosslinking process in the spectra of all ICCh samples appears a new band at 1635 cm‑1. This band, that can be assigned to antisymmetric deformation N‑H vibrations in NH3+ ion [6], was observed earlier by us and others for ionically crosslinked chitosan membranes [6, 8 ‑ 10]. Moreover, additional changes in the spectra of chitosan after its ionic crosslinking can be observed: (i) in the spectrum of Ch/H2SO4 the new absorption band at 617 cm‑1 appears that corresponds to S‑O bending vibration in sulfuric ions [6], (ii) in the spectrum of Ch/CIT the new band at 1380 cm‑1 appears. It corresponds to C‑O symmetric vibrations in COO‑ ions [6], (iii) in the spectrum of Ch/TPP the new band at 1223 cm‑1 appears that corresponds to ‑P=O stretching vibrations in phosphate ions [6], (iv) in the spectrum of Ch/NaAlg the new absorption band at 1411 cm‑1 appears that corresponds to C‑O antisymmetric vibrations in COO‑ ions [5, 6]. Spectral changes characterized above indicate the formation of ionic crosslinks between chitosan and crosslinking agents as presented Figure 4.
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M. Pieróg, M. Gierszewska-Drużyńska, J. Ostrowska-Czubenko CH2OH H
CH2OH O O
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Figure 4. The chemical structure of ionically (A) Ch/H2SO4, (B) Ch/CIT, (C) Ch/TPP, (D) Ch/NaAlg.
crosslinked
chitosan
membranes:
The results obtained for the swelling kinetics at pH 1.2, 7.4 and 8.5 are reported in Figure 5. As can be seen in Figure 5.A, all uncrosslinked and crosslinked chitosan membranes swollen in acidic media underwent disintegration. It has been observed by us that the time needed to membrane disintegration in acidic media increased in the following order: Ch ≈ Ch/H2SO4 ≈ Ch/CIT Ch/ NaAlg > Ch/TPP. Thus, it can be stated that at pH 7.4 the membrane swelling ability changes in the same order. In buffer solution of pH 8.5 (Figure 5.C) all examined membranes were stable and the constant S value was attained. In comparison to results presented in Figure 5.B, the corresponding S values were lower. The decreasing values of equilibrium swelling degree allow putting the Ch and ICCh membranes in following order: Ch
