Study Of A Novel Biodegradable Piperazine-based Poly (urethane-urea) For Bone Tissue Repairing

Last centuries, bone repair engineering is one of the most important topics for us to explore. Bone, a live biomaterial in the nature, is a good example of a dynamic tissue, since it has a unique capability of self-regenerating or self-remodeling to a certain extent throughout the life. However, many circumstances call for bone grafting owing to bone defects either from traumatic or from non-traumatic destruction. In the case of severe defects and loss of volume, bone cannot be healed by itself and grafting is required to restore function without damaging living tissues. So far, the grafting based biomaterial was attracted more and more attention and had potentiality to solve this clinical puzzle. In the repair, the suitable biomaterial scaffold is need with many excellent properties, such as biodegradablity, appropriate mechanical properties and biocompatibility and so on. In my present, a novel biodegradable piperazine-based poly (urethane urea) (P-PUU) is synthesized to meet the requirement of scaffold material in bone repair. Some methods of fourier transform infrared spectrometry (FTIR), Gel permeation chromatography with multi angle laser light scattering (GPC-MALLS), differential scanning calorimeter (DSC), nuclear magnetic resonance spectrometer (NMR), high resolving capability atomic force microscope(AFM) and classical chemical analysis were used to characterize its structures. Then mechanical tensile testing and dynamic mechanical analysis (DMA) were used to explore mechanical and shape memory properties of P-PUUs. Thirdly, the hydrophilicity/hydrophobicity, biodegradation and thermal degradation of the P-PUUs were investigated. Thereafter, the cell biocompatibility of P-PUUs was systemically evaluated. The main works and conclusions are included as follows:1. Hydroxy group ended poly (D, L-lactide) (PDLLA diol) was synthesized by melt ring-opening polymerization of D, L-lactide using Sn (Oct) 2 as initiator and piperazine as coinitiator. Then, the conditions of ring-opening polymerization of D, L-lactide with piperazine as coinitiator were optimized. An extensive investigation effort was expended in understanding the effects of dosage of coinitiator on the molecular weight and the structure and thermal properties were characterized.①FTIR, 1HNMR revealed that PDLLA diol was successfully obtained by using above-mentioned system. The optimization conditions of ring-opening polymerization of D, L-lactide with piperazine as coinitiator were as follows: reaction time, 24h; reaction temperature, 150℃.②Hydroxyl value, 1H NMR and GPC-MALLS analysis indicated that PDLLA diol with different molecular could synthesized by varying the ratio of D, L-lactide/PP.③The results of DSC indicated that the glass transition temperature (Tg) depended on its molecular weight and the Tg increased from 15.95 to 32.15℃with the increasing of molecular weight.2. A series of P-PUUs materials were prepared based on PDLLA diol as the soft segment, hexamethylene diisocyanate (HDI) and piperazine (PP) as the chain extender. The effects of the reactive temperature and ratio of hard segment/soft segment were discussed, and the structure and thermal properties of P-PUUs were characterized.①FTIR, 1H NMR and 13C NMR exhibited that, PDLLA diol have successfully reacted with HDI, and then P-PUUs with high molecular weight could obtained by chain extending reaction with PP as the chain extender. The optimization extending reaction was 30℃, nearby the room temperature.②By varying the molecular weight of soft segment and the ratio of soft segment/hard segment, the P-PUUs with gradient contents of piperazine could be synthesized. Therefor, those materials would have different physicochemical properties for application in bone repair.③The weight analysis for the degree of cross-linking showed that comparing with BDA as the chain extender, PP as the chain extender can make the synthesis process of polyurethanes easier to control and lead the polyurethanes with low or no degree of crosslinking, which lays a solid foundation for physical properties of the P-PUUs.④The results of DSC showed that Tg could be regulated by the ratio of hard segment/soft segment and with the content of hard segment raising, Tg of P-PUUs rose. In addition, two temperature transitions in DSC curves of P-PUUs were obviouly observed: the slight transition was cosed to the Tg of PDLLA diol, could be attributed to the internal loosening of soft segments, the other one represented the Tg of P-PUUs. This implied that typical phase separation of P-PUUs due to the thermodynamic incompatibility of the two segments might occur.⑤The results of high resolving capability AFM for scanning the monomolecular layer of P-PUUs indicated that the P-PUUs were successfully obtained with typical phase separation.3. The mechanical properties of P-PUUs were characterized by mechanical tensile testing and DMA, and the influences of molecular weight of soft segment, the ratio of soft segment/hard segment were investigated. The shape memory behaviors were also studied by the tensile testing.①The results indicated that P-PUUs have nice mechanical properties. The tensile modulus and tensile strength increased with the increasing of the hard segment content, while the elongation at break had the opposite trend. With the raising of the molecular weight of soft segment, the tensile modulus and tensile strength decressed, but the elongation at break increased. PP as chain extender could improve the mechanical properties with two methods: one is that PP has a six-atom rigid ring, which was introduced into the backbone of the P-PUUs to improve tensile modulus and tensile strength, the other one is that PP as chain extender can reduce the dregree of crosslinking.②The results of DMA showed that the storage modulus and the transitions of Tanδincreased with the hard segment content rising.③P-PUUs had good shape memory properties with large shape fixation ratio (Rf, all above 95%) and shape recovery ratio (Rr, all above 93%). Meanwhile, the shape memory properties were affected by the components and the hard segment content. Rr decreased with the increasing of the hard segment content, however, Rf had the opposite trend.4. The hydrophilicity/hydrophobicity, biodegradation and themal degradation of P-PUUs were investigated. The evaluation indicators of hydrophilicity/hydrophobicity were static water contact angle and water absorption ratio, while the evaluation indicators of degradation behavior were the weight loss ratio and pH value changes, surface topography, moreover, the themal degradation were studied by thermogravimetric analysis and activation energy. In addition, the effect of different chain extenders on degradation properties of segmented polyurethanes was investigated.①The static water contact angle of P-PUUs was smaller than PDLLA controls and the water absorption ratio was more than PDLLA controls. This is mainly due to the fact that the hydrophilic PP segment could form more hydrogen bond with water molecules and improved the hydrophilicity of P-PUUs. Meanwhile, the hydrophilicity had an uptrend with the increasing of hard segment content.②Data of hydrolytic degradation of the polymers during 12 weeks indicated that the in vitro degradation stability of P-PUUs was more than PDLLA control, because the alkaline substance during the degradation of P-PUUs could eliminate or weaken the acid induced auto-catalysis.③The results revealed that chain extender played an important role in biodegradation and thermal degradation of polyurethanes. During the biodegradation, polyurethane with amine style chain extender had more stability than those with hydroxyl chain extender, however, during thermal degradation the carbamate revealed more stability than urea bond. The results of the thermal degradation of polyurethanes were confirmed by the activation energy, which was calculated from the TGA data according to Ozawa-Flynn-Wall method.5. The cytocompatibility of P-PUUs was evaluated by employing primary SD rat osteoblasts as the model cells and poly (DL-lactic acid) (PDLLA) as the control. The osteoblasts morphology, cell attachment and spreading, cell proliferation, cell differentiation and mineralization ability were systemically detected to indicate the cytocompatibility of P-PUUs.①Initial morphology, adhesion, spreading of osteoblasts on all P-PUU films is no better than those on PDLLA film. Furthermore, with the increased amount of hard segments in P-PUUs, both cell doubling rate and migration rate correspondingly increased. Those results revealed that the interaction between cell and material was a complicated process, not only completely depended on the hydrophilicity of matetials.②The results of cell proliferated behaviors on different films showed that P-PUUs could promoted the proliferation of osteoblast, and with the the increasing of the hard segment content, the positive regulation was more and more obvious. These results were confirmed by Boltzmann function sigmoidal fit. Based on it, we hypothesized that the interaction between cell and material could be ascribed to biocenose growth Logistic model.③Osteoblast on different P-PUUs films all exhibited lower physiological functions compared to those on PDLLA films within the first 10 days after seeding. Thereafter, however, the osteoblasts on P-PUUs demonstrated better differentiation and mineralization than those on PDLLA films, and P-PUU-3 > P-PUU-2 > P-PUU-1.