Having determined that the 12.4 mg TA/g GE-HA scaffold had the preferred physical, microstructural and mechanical properties it was chosen for biodegradability and bioactivity evaluation studies. The effectiveness of HA based implants relies on the ability of the scaffold to form on its surface a calcium-phosphate layer , as it can initiate bone formation through the activation of signaling proteins . In order to examine the ability of such a layer to form on these scaffolds in vitro studies were performed by immersing them into a simulated body fluid (SBF) solution. It was shown that after 7 days of immersion such a bone-like apatite film formed throughout the scaffold surface. The microstructure depicted in Fig. 4.2a, b illustrates that this film was comprised of spherical nano-plate aggregates with a 150 nm length. After 14 days the microstructure of the film was slightly altered as the spherical aggregates were elongated and eroded (Fig. 4.2c, d). This change in shape was attributed to the initiation of degradation of the scaffold, since as the HA-gelatin material dissolved stresses evolved at the calcium-phosphate/scaffold interface, which resulted in the deformation of the film crystals. Examination of the scaffold 21 days after immersion in the SBF indicated that the bone-like apatite coating was not continuous, since the scaffold supporting it had further disintegrated and therefore the attached Ca/P exfoliated in the solution. It should be noted that throughout the 21 days of immersion the Ca/P ratio that remained attached on the scaffold was at the values present in trabecular and cortical bone and the observed microstructure of the bone-like apatite crystals was similar to that reported for promoting osteo-integration [19, 20].
In addition to examining the effect that SBF fluid had on the scaffold microstructure, their biodegradability was also examined by immersing them in three additional solutions and monitoring their weight loss after 3, 12, 16 and 25 days following immersion. The first solution was comprised of PBS with a pH of 7.4 and corresponded to degradation in physiological fluids, the second solution included acetic acid/sodium acetate buffer solution with a pH of 4.2 and corresponded to the resorption process of incharge osteoblasts and the third solution included lysozyme (4 mg/mL) into the PBS in order to simulate decomposition by enzymatic catalysis. After 25 days the weight loss was 15% for the physiological fluid conditions, 75% for the 4.2 pH solution and 95% for the enzymatic degradation . The ability of the scaffold to degrade under a 7.2 pH is necessary such that the deposition of biogenic hydroxyapatite crystals can occur as described earlier and shown in Fig. 4.2.