TJPS Vol.41 (Supplement Issue) 2017

TJPS 2017, 41 (Supplement Issue): 109

Formulation of magnetic-responsive composite hydrogel

Chanipa Siangsanoh1,2, Sarute Ummartyotin1,2, Pleumchitt Rojanapanthu3, Worapapar Treesuppharat3,*

1 Advanced Functional Polymeric Materials Research Group, Faculty of Science and Technology, Thammasat University,

Patumtani 12120, Thailand 2 Materials Research Center, In collaboration with HORIBA Scientific and Thammasat University, Patumtani 12120,

Thailand 3 Drug Discovery and Development Center, Advanced Science and Technology, Thammasat University, Patumtani 12120,

Thailand

* Corresponding author: Tel. +66(0)25643090; Fax. +66(0)25643092; E-mail address: g4736963@gmail.com

Keywords: Composite hydrogel, Magnetic nanoparticles, Gelatin, Bacterial cellulose, Magnetic

responsiveness

Introduction

Hydrogels are three dimensional polymeric networks that can absorb and retain a large quantity of

aqueous solution without dissolving. Hydrogels based on natural polymers are more attractive to use in many

applications such as wound dressings, tissue engineering scaffolds and drug delivery systems. Gelatin is a

natural polymer derived from protein with favourable properties including biodegradability, low immunogenicity

and low cytotoxicity. Amino acids in side chain groups in gelatin are able to easily crosslink with various

chemical reagents such as carbodiimide and glutaraldehyde. However, gelatin hydrogel has poor mechanical

and thermal stability.1 To improve its mechanical and thermal properties, polysaccharides such as cellulose

can be used as reinforcement materials by incorporating into gelatin hydrogel matrix, so-called composite

hydrogel. Meanwhile, smart hydrogels have also gained importance for biomedical applications because their

swelling behaviour can change in response to environmental stimuli such as temperature, pH and glucose.

Furthermore, magnetic nanoparticles (MNPs) can be responsive to an external magnetic field. Most magnetic

iron oxide nanoparticles that are more attractive for biomedical applications include magnetite (Fe3O4) and

maghemite (-Fe2O3) because of their biocompatibility and relative non-toxicity when equipped with

appropriate coatings.2 This research aimed to prepare and characterize magnetic composite hydrogels (Mag-

H) from gelatin, bacterial cellulose (BC) and MNPs using glutaraldehyde as a crosslinking agent. For this

purpose, gelatin-coated MNPs were synthesized by aqueous co-precipitation method and determined for their

polymorphic form, size, zeta potential and magnetic property. The prepared Mag-H were characterized for

their magnetic property and swelling behaviour.

Methods Synthesis of gelatin-coated MNPs

The gelatin-coated MNPs were synthesized by aqueous co-precipitation of FeCl3/FeCl2 (molar ratio of

2:1). Initially, 2.16 g of FeCl3 and 0.8 g of FeCl2 were dissolved in 1 M HCl solution. The resultant solution was

added to 2% w/w gelatin solution (coating solution) at equal volume and stirred at room temperature (RT) for 1

h followed by removing dissolved oxygen with N2 gas bubbling. The resulting mixture (20 g) was added

dropwise to 2.5 M sodium hydroxide solution (30 g) at 90°C under N2 atmosphere with vigorous stirring for 1 h

followed by further continuous stirring at RT for 2 h. The obtained black precipitate was sonicated for 15 min

and then washed with deionized water using a refrigerated centrifuge until neutral pH was achieved in order to

remove impurities and unbound gelatin coating. The final product was sonicated again and stored at 4°C.

Thai Journal of Pharmaceutical Sciences (TJPS) The JSPS-NRCT Follow-Up Seminar 2017 and

33rd International Annual Meeting in Pharmaceutical Sciences

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TJPS 2017, 41 (Supplement Issue): 110

Preparation of magnetic composite hydrogels (Mag-H)

Gelatin-coated MNPs suspension and BC suspension (1% w/w) were added to gelatin solution (20%

w/w) and mixed well at 40°C to obtain homogeneous mixture. Then, 80 l of glutaraldehyde (25% w/w) acted

as a crosslinking agent was added dropwise into the mixture. The resulting mixture was poured into a petri

dish and placed in an oven at 55°C for 4 h. Mag-H were obtained and then removed from the petri dish

followed by washing with deionized water to remove unreacted chemicals. The composite hydrogels were

then dried in an oven at 45°C. The exact compositions of the Mag-H prepared at different ratios of MNPs to

composite hydrogel are listed in Table 1

Table 1. Composition of magnetic composite hydrogels (Mag-H)

Code MNPs:Hydrogel

(mg:g)

20% w/w gelatin

solution (g)

1% w/w BC

suspension (g)

MNPs suspension

[25 mg/ml] (ml)

Deionized

water (g)

Mag-H 0.4 0.4:1 10 1 0.32 8.68

Mag-H 5 5:1 10 1 4 5

Mag-H 10 10:1 10 1 8 1

Measurement of zeta potential and particles size

Hydrodynamic diameter and zeta potential of the MNPs suspension were measure by dynamic light

scattering (DLS) using a nanopartica SZ-100 (Horiba Scientific, Japan) . All DLS measurements were done

with a scattering angle of 90° at 25°C after diluting the suspension to an appropriate volume with deionized

water.

Analysis of crystal structure and size

The dried MNPs powder was analyzed for a crystal structure and size by XRD (D8 Advance, Bruker).

The MNPs powder was placed uniformly on a sample holder to make sure that the sample had the same X -

ray exposure. The sample was exposed to X-ray radiation using CuKα (λ = 1.542 Å) between of 20° and 80°

( 2θ) at 25°C. Average crystallite size of the core of MNPs was calculated from XRD data using Scherrer's

equation.3

Measurement of magnetic property

The magnetic properties of the dried MNPs and swollen Mag-H at different ratios were measured

using an in-house developed vibrating sample magnetometer4 at RT with an applied magnetic field up to

10,000 Oe. For measurement of magnetic responsiveness of Mag-H, the swollen hydrogels were immersed in

deionized water at the right side of a 250-ml beaker. A magnet was then placed at another side of the beaker.

The magnetic attraction of Mag-H was observed.

Swelling test

Before the swelling tests were performed, the dried Mag-H at different ratios were weighed before

immersion into deionized water at 37°C in the presence and absence of the applied magnetic field. After

immersion for 24 h, the swollen hydrogels were taken out from deionized water and the excess water on the

surface of hydrogels were removed with filter paper before weighing them. This study was done in triplicate.

The swelling ratio of the hydrogels was calculated using Eq. (1).5

Swelling ratio (%) = [(Wt - W0)/W0] ×100 (1)

where Wt is the weight of swollen hydrogel and W0 is the initial weight of the hydrogel.

Statistical analysis

The swelling results are expressed as the mean ± standard deviation (SD). Analysis of variance was

performed by ANOVA procedures using IBM SPSS Statistics 21. The significance of the differences between

means was tested using the post hoc Tukey’s test and Dunnett’s T3 pairwise comparison when the variances

were equal and unequal, respectively. A p < 0.05 was considered statistically significant.

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TJPS 2017, 41 (Supplement Issue): 111

Results and discussion Physicochemical characteristics of gelatin-coated MNPs: The XRD pattern for the synthesized gelatin-

coated MNPs is shown in Figure 1. The XRD spectra showed seven characteristic peaks at 2θ of 3 0 .0 5 °,

35.05°, 43.10°, 53.64°, 57.23°, 62.73°and 74.42° that corresponded to the following Miller indices: (220),

(311) , (400) , (422) , (511) , (440) , and (533). These results indicated that the core of the synthesized MNPs

were magnetite (Fe3O4) with a spinel structure.3,6 The average crystallite size of the core was 12.7 nm and

mean hydrodynamic diameter of the gelatin-coated MNPs was 108 ± 9 nm. Due to the presence of gelatin in

their coating, the gelatin-coated MNPs had a negative zeta potential (-23.4 mV).

Magnetic properties of gelatin-coated MNPs and Mag-H: Magnetization curves ( M-H loop) from VSM

analysis showed that both gelatin-coated MNPs ( Figure 2a) and Mag-H ( Figure 2b) have a

superparamagnetic behaviour as indicated by a very narrow hysteresis loop. The superparamagnetism, which

is responsiveness to an applied magnetic field without permanent magnetization, can play a crucial role in the

applications of drug delivery. 3,6 The saturation magnetization of gelatin-coated MNPs was 50 emu/g, which

was higher than that of Mag-H 0.4 (0.03 emu/g), Mag-H 5 (0.28 emu/g) and Mag-H 10 (0.64 emu/g) because

of a small amount of the MNPs incorporating into the composite hydrogels. The magnetic property of Mag-H

tends to increase with the increase of MNPs content in the composite hydrogel. Figure 3 demonstrates the

magnetic responsiveness of Mag-H by placing a magnet near the left side of a beaker. Mag-H 10 was

attracted towards the magnet, whereas the magnetic response was not observed at other ratios. It can be

attributed to the fact that Mag-H 10 had saturation magnetization higher than other ratios. The results

indicated that MNPs to composite hydrogel ratio of 10: 1 is an optimal ratio to formulate a magnetic field-

responsive composite hydrogel for magnetic directing to the targeted site.

Figure 1. XRD patterns of gelatin-coated MNPs and the bottom of the image indicates the JCPDS data of

magnetite (JCPDS no. 01-088-0315)

Figure 2. Magnetization curves (M-H loop) of gelatin-coated MNPs (a) and Mag-H (b)

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TJPS 2017, 41 (Supplement Issue): 112

Figure 3. Magnetic responsiveness of Mag-H in the absence of magnet (a) and in the presence of magnet at

ratio 10:1 (b), 5:1 (c) and 0.4:1 (d)

Swelling test of Mag-H: The swelling tests were conducted to study swelling behaviour of Mag-H at different

ratios as shown in Figure 4. After immersing all Mag-H in water at 37°C for 24 h in the presence of the applied

magnetic field, Mag-H 10 and Mag-H 5 had a maximum swelling ratio of about 600%, which was significantly

higher than Mag-H 0.4 (p < 0.05). However, the significant difference in swelling ratio of the composite

hydrogels between the presence and absence of the applied magnetic field was clearly observed only in Mag-

H 10 (p = 0.002), indicating that Mag-H 10 had a magnetic-sensitive swelling behaviour. The swelling ratio of

Mag-H 10 with the magnetic field application (607 ± 5%) was higher than that without the magnetic field (510

± 16%). This observation can be attributed to the alignment of MNPs within the hydrogel network by the

external magnetic field, which can cause winding of the polymer chains leading to expand the hydrogel

network.7

Figure 4. Swelling behaviour of Mag-H at different ratios

Conclusion Gelatin-coated superparamagnetic nanoparticles with a saturation magnetization of 50 emu/g were

successfully synthesized by aqueous co-precipitation method. The synthesized MNPs can be used as a good

nanomaterials for preparing the magnetic-responsive composite hydrogel. With an optimal ratio of MNPs to

composite hydrogel of 10:1, the Mag-H had a good magnetic property, which affected their magnetic

sensitive-swelling behaviour as well as magnetically targeted movement. Therefore, this formulation could be

a promising candidate to be developed as a magnetically targeted drug carrier.

Acknowledgements The authors would like to acknowledge the financial support provided by Thammasat University under the TU

New Research Scholar, Contract No. 28/ 2557 and Thailand Research Fund ( TRF) , Contract No

TRG5880190.

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