Poster N19 - Mechanical performance of 3D-printed nanocomposite skeletal biomaterials exposed to simulated oxidative environments

Introduction: Biopolymer nanocomposites are an exciting class of materials with strong potential as biomaterials for skeletal reconstruction. Incorporating these materials with additive manufacturing technologies allows for complex and customizable products that have great potential as synthetic bone graft biomaterials which could reduce the dependence on natural bone grafts and metal prostheses (Figure 1). Novel nanocomposite biomaterials composed of functionalized triglyceride-based monomers and hydroxyapatite (HA) have been previously developed which exceed the mechanical properties of commercial bone cements. However, these materials have been previously evaluated under room temperature and dry air conditions, which are not indicative of the intended physiological operating conditions of these orthopaedic biomaterials. These testing conditions also neglect any mechanical consequences of biological interactions with these materials, such as oxidation due to inflammation.

In addition, a design goal for these graft materials is to allow for successful remodelling of natural bone tissue which requires susceptibility to degradation and resorption. In vitro oxidative degradation testing can be performed to assess the biomaterials’ susceptibility to oxidation and give an estimate of their in vivo behavior. It was hypothesized that our biopolymer nanocomposite materials would be susceptible to degradation, observed through mass loss and reduced mechanical properties. The objective of this work was to evaluate the mechanical properties of 3D printed biopolymer nanocomposites after exposure to oxidative degradation, and to contrast two in vitro oxidative environments, hydrogen peroxide (H2O2) and hypochlorous acid (HClO) to determine their suitability as biomaterial screening tools.

Materials and

Methods: The biopolymer resins used in this study were composed of methacrylated acrylated epoxidized soybean oil (50 vol%) and triethylene glycol dimethacrylate (50 vol%). Irgacure 819 was added at 1 vol% as a photo-initiator. Two ink compositions were trialled with 0 and 10 vol% calcium-deficient nanoscale HA (0HA and 10HA, respectively). Inks were processed using a Branson Sonifier ultrasonic homogenizer (Emerson Inc., USA) and a Unidrive mechanical homogenizer (M. Zipperer GmbH, Germany).

Mechanical (dog-bone tensile and cylindrical compressive) test specimens were then printed using masked stereolithography (Sonic XL 4K printer, Phrozen Technologies, Taiwan) with 50 μm layer height and 6 s cure time. Specimens were post-print cured with a CureZone MKII curing chamber (Creative CADWORKS, Canada). To induce physiologically relevant conditions, specimens were incubated at 37 °C for periods of 14 and 28 days. To investigate their response to oxidation, specimens were exposed to 20 vol% H2O2 and 5 mM HClO, as commonly done in literature. Phosphate-buffered saline (PBS) was used as a control.

After incubation, specimens were then mechanically tested using a Psylotech μTS mechanical testing machine (1.6 kN load cell; Psylotech Inc., USA) equipped with digital image correlation for strain field measurements. A heat lamp was incorporated to enable mechanical testing at 37 °C. Tensile and compressive testing were performed following ASTM D3039 and ASTM D695, respectively. In addition, the change in the specimens’ dry masses throughout their incubation periods were recorded, and their surface characteristics were evaluated using digital macroscopy and scanning electron microscopy (SEM).

Results, Conclusions, and Discussions: Biopolymer nanocomposite specimens exposed to both oxidative environments displayed significant evidence of degradation (Figure 2). 10HA samples experienced ~ 16 and 38 vol% mass losses after 28 days of incubation in HClO and H2O2, respectively. In contrast, materials incubated in PBS experienced no significant loss of dry mass. In addition, surface imaging of the degraded samples showed significant defects (Figure 2(b)). H2O2 caused pitting in the surface of the 10HA specimens with average areas of 0.01 mm2/pit after 28 days. HClO initiated larger voids in the composites’ surfaces, with average areas of 0.33 mm2/pit. Interestingly, the incorporation of HA appeared to slow the surface degradation from HClO, while it strikingly increased degradation due to H2O2 (Figure 2(a)).

The mechanical performance of these composites was impacted by two major effects: softening due to swelling and water absorption effects, and loss of structure due to surface oxidative degradation. To highlight these observations, the ultimate tensile strengths calculated with their degraded, ‘real’ area and normalized original areas are shown in Figure 3. Similar reductions in ultimate tensile strength were observed between samples incubated in degradation and PBS media, highlighting the overall softening effect from swelling. However, there was also an impact upon the mechanical performance due to loss of mass and structure, highlighted in the differences of the 10HA, H2O2 specimens when normalized to original dimensions. Overall, there is a significant reduction in the mechanical performance of these biopolymer nanocomposites when exposed to model oxidative environments, resulting in mechanical properties considerably less than the bone cement benchmarks these materials exceed under dry conditions.

Clear evidence of the susceptibility of these biopolymer nanocomposites to oxidative degradation was attained. Two methods of oxidative degradation in vitro modelling were contrasted, and HClO was determined as a more consistent method of degradation modelling when evaluating biomaterials containing HA. Overall, the mechanical performance of these biomaterials is susceptible to two factors: bulk swelling effects and loss of structure. This work provided a comprehensive investigation into the effects of oxidative degradation on biomaterial functionality and highlights the importance of exploring relevant physiological effects on mechanical properties when developing biomaterials.

Acknowledgements (Optional): This work was supported by the Natural Science and Engineering Research Council of Canada, the Canadian Institutes of Health Research, the Canadian Foundation for Innovation, Ontario Research Fund, and the University of Waterloo.