Morphology and gelation of thermosensitive xyloglucan hydrogels

Abstract

Galactose modified xyloglucan is a thermally reversible hydrogel that is increasingly used in the biomedical field due to the ease of altering the gelation time and temperature by modifying the galactose removal ratio. However there is little information concerning the morphology and rheological properties of the hydrogel under physiological conditions. Differential scanning microcalorimetry (DSμC) showed the thermal gelation process to occur over a broad temperature range (5–50 °C). The rheological properties of the hydrogels were investigated as a function of concentration, temperature and ionic strength. The final elastic moduli of the hydrogels increased with increases in concentration. Isothermal rheology suggests that the gelation occurred in two distinct stages, which was influenced by the solution media. Scanning electron microscopy (SEM) was used to characterize the morphology of the xyloglucan which were thermally gelled at 37 °C. The resultant morphology was strongly dependent on the concentration of the hydrogel. Strong hydrogels were only obtained at 3 wt.% at 37 °C, and the morphology characterized by an open 3-dimensional network, comprised of thin membranes. It is proposed that the first stage of the isothermal gelation is the formation and growth of the thin membranes, followed by the formation of a three dimensional network.

Introduction

There has been growing interest in the use of thermally sensitive in situ forming, physical hydrogels for biomedical applications [1], [2] as minimally invasive scaffolds for tissue engineering. Because of their low interfacial tension and high molecular and oxygen permeability, hydrogels are ideal tissue engineering constructs [3]. Consequently, hydrogels have been investigated for cell gene therapy [4], enzyme and cell encapsulation [5], [6], [7], drug delivery [8], [9], [10], [11], joint cushioning and lubrication [8], [12] and as environmental shape memory materials [4]. The utility of hydrogels as tissue engineering scaffolds appears to be related to microscopic [13] and macroscopic [14] porosity within the structure of hydrogels. Microscopic pore interconnectivity and the volume of water phase present within the hydrogel ensures cellular viability, permits cell migration, increases transportation of nutrients, oxygen and metabolites [15], and influences drug release profiles and enzymatic degradation [8]. The shape and size distribution of the pores also influence cellular function [8]. By exploiting these features it may be possible to mimic features of the natural extracellular matrix, and control tissue structure and cellular functions [16].

Xyloglucan is a neutral, non-toxic polysaccharide [17], whose degradation products consist of naturally occurring saccharides and are assumed non-toxic, although the experimental evidence for this conclusion is limited. Xyloglucan is extracted from the tamarind seed and is a major component of higher plant cell walls [18]. It is composed of a β-1,4 linked D-glucan backbone where the O-6 positions of the glucopyranosyl residue are partially substituted with α-D-xylopyranose residue [11], [19]. Fig. 1 shows the backbone structure of xyloglucan and Fig. 2 presents the structure of the galactopyranose, glucopyranose and xylopyranose which make up the polysaccharide.

Thermally responsive xyloglucan is formed using fungal β-galactosidase to remove more than 35% of the galactose residue [17]. Shirakawa et al. [17] found that the optimum removal ratio was between 35% and 50%. Using DSC and rheology they related gelation temperature to the concentration of xyloglucan in the pre-gel solution. Miyazaki et al. [11] found that increasing the concentration of the pre-gel solution from 1% to 2% decreased gel temperature from 27 to 22 °C as well as the gel time. A lower and upper transition from sol–gel and gel–sol, respectively, were found, and the gel was shown to be thermo-reversible upon cooling. It was also determined that with a higher galactose removal ratio there was an increase in temperature range of the gelation peaks, hence a broader gelation range [17].

While tissue engineering applications for xyloglucan are not extensive, it has been used as skin patches [20], oral and rectal delivery of drugs [11], and for intraperitoneal injections [21]. It was drug loaded in the latter two applications and was found to provide stronger bioavailability of the relevant drug and longer residence times than previous commercial suppositories [22]. Importantly there was no apparent tissue damage [11] implying that xyloglucan hydrogel is a biocompatible material that can be implanted non-invasively via injection in tissue engineering applications.

X-ray scattering studies of the gel nano-structure of galactose-modified xyloglucan, showed that flat structures were formed from lateral stacking of rod-like chains [23] whereas the addition of ethanol to the solution created a random structure consisting of condensed phase aggregates amongst dilute, single chain areas [24].

This study aims to investigate the thermal, rheological properties of thermally gelling xyloglucan with a galactose removal ratio of 48% with consideration of its use as an injectable tissue engineering scaffold. The effect of changing the ionic strength of solution media on the properties of the hydrogel were determined by comparing deionised water and phosphate buffered saline (PBS) as solution media. This study also presents for the first time, the morphology of the gel networks as determined using electron microscopy.