A New Chitosan-PEG Wound Dressing with Enhanced Mechanical Properties

A successful wound dressing is required to show acceptable structural, mechanical and biological properties. Here we aimed in developing a new wound dressing using chitosan as a biocompatible material with the antibacterial property and regenerative potentials, in combination with polyethylene glycol polymer (PEG) for mechanical and structural enhancement by crosslinking chitosan to well carboxylated PEG. The percentages of polymers, the carboxylation process and method of mixing were major parameters to obtain a proper porous structure with appropriate biological and mechanical properties and economical advantage. Hence we used wide ranges of chitosan percentage (15, 30, 50, 70, and 85 percent) and molecular weight of PEG (2000, 3350, 4000, 6000, and 10000), and two methods of fabrication through casting and freeze drying to obtain a novel chitosan based wound dressing with enhanced properties. To produce the CS-PEG wound dressing, first two ends of PEG chains were carboxylated in combination with Succinic Anhydride. Carboxylation of PEG was performed in four different methods and the optimized method was selected based on FTIR results (For properly carboxylated PEG a noticeable peak is observed in FTIR graph in 1730-1750 Hz), the toxicity of the final material and facility of synthesis method. Our results indicated that the successful method of fabrication was achieved when the solution of PEG in methylene chloride was combined with succinic anhydride at 60 °C. Then chitosan functional group (NH2) and carboxylated PEG functional group (COOH) were cross-linked during an esterification reaction. The chemical bond affinity of CS – PEG chains increased by use of EDC and NHS in CS – PEG structure. An optimized strategy to achieve enhanced dressings relies not only on selection of proper constituents but also on a suitable method of fabrication with economical advantage. Two methods of fabrication were used to prepare CS-PEG films. In the first method, the synthesized solution was casted and dried at ambient temperature. In the second method, the solution was frozen abruptly by liquid Nitrogen and was dried by a freeze dryer leading to the sponge type highly porous structure. The fabricated films of two methods were examined to compare their chemical, mechanical and biological properties. Different material properties were examined including swelling, biodegradability, water vapor transmission, and porosity. Furthermore, samples were mechanically characterized by elastic modulus, viscoelastic parameters, and dynamic moduli. All tests were repeated at least three times and the average ± standard deviation values were calculated. We performed structural tests on dried samples of two methods to select the optimized method. The processed samples showed a proper swelling behavior, gaining most swelling in PBS within the first 30 minutes. The degree of swelling of examined samples in both PBS and blood decreased by an increase in the percentage of chitosan most likely due to a reduction in cross-linked chain distances, however such trend was not observed by increasing chitosan percentage. The CS-PEG hydrogels are capable of absorbing blood and exudates within the range of 3-10 times of their weights which can facilitate the remodeling and healing of granulated tissues. Besides, the high range of blood absorption (3-5 times of the dressing weight) can assist the homeostasis phase in the injury site. On average samples produced by freeze-drying method were swollen to 5.83 folds by mass during 48 hr incubation in PBS, with 4.78 folds in the first two hours. The synthesized CS-PEG hydrogels showed longer biodegradation compared to pure chitosan. Samples with a higher percentage of chitosan showed slower biodegradability rates, as samples with 30% chitosan were thoroughly decomposed before 28 days. Hence they were not considered as appropriate candidates for wound dressing. On the other hand, the rate of degradation among samples with a high ratio of chitosan (85%) was by far low, such that after 28 days, only 26% of the mass was reduced. It was concluded that samples with chitosan ratios of 50% and 70% were degraded with sustained rate and appropriate time span for wound healing. Although samples produced by freeze-drying showed proper swelling and blood absorption properties, they indicated poor results in biodegradability and mechanical properties compared to samples produced by casting. The majorities of freeze-dried samples were degraded within two weeks and showed low strength with the average value of 0.26 MPa. Both results indicated inadequate features for a suitable wound dressing candidate. Therefore such samples were disregarded from further analyses. Samples with 70% and 50% of chitosan and molecular weight of 2000, synthesized by casting method were selected after meeting the adequate requirements as proper candidates for the wound dressing. The selected samples not only showed adequate elastic characteristics, but they also performed well in viscoelastic behavior. Skin tissue is considered to be viscoelastic; the candidate dressings similarly showed viscoelastic behavior. The stress relaxation data of samples described a viscoelastic solid state similar to soft tissues including skin, with the dominant effect of storage modulus (represented by the elastic element) compared to loss modulus (represented by the viscous element). Furthermore, the storage modulus showed a stable behavior in a wide range of frequencies. CS – PEG films porous structure with small size pores provided an appropriate environment to enable absorbance and drainage of wound exudate and transmission of vapor and passage of Oxygen and CO2 while forbidding the entry of external contamination into the wound environment. It seems that the average pore size of 2.3 mm that was provided by the synthesized dressings of CS70%-PEG2000 through casting method is an appropriate value for the mentioned purposes. In general, the permeability of samples after one week was in the range of 200 - 7300 gr/m2. The degree of permeability in CS-PEG films increased by the decrease of chitosan ratio most probably due to an increase of cross-linked chain distances. The high reduction rate of bacteria colonies (the average 95 percent) of each sample after 24 hours of culturing Escherichia coli and Staphylococcus aureus bacteria shows outstanding antibacterial properties of fabricated CS-PEG wound dressings, likely due to high content of chitosan. The reduction of colony formation was more in CS70 than CS50. This benefit reduces the possibility of infection and wound odor, and subsequently enhances healing and reconstruction of granulated tissue Moreover, they indicated non-toxicity for L929 fibroblast cells with over 98% viability in MTT assay. The selected samples were introduced as wound dressing candidates.

Keywords:

Wound Dressing , Chitosan , Polyethylene Glycol , Mechanical properties , Biocompatibility