P3HB VE PBA POLİMERLERİ İKİLİ KARIŞIMLARININ DOKU İSKELESİ OLARAK DEĞERLENDİRİLMESİ

Abstract

In the thesis study, to produce a novel tissue engineering material, a biopolyester poly-3- hydroxybutyrate (P3HB) reinforced with poly-β-alanine (PBA), PBA copolymer poly(β- alanine-co-ethyleneoxide (PMBA), PBA derivative poly(N-(3-methoxypropyl)-β-alanine) (PNMPBA) polyamides, which were synthesized by hydrogen transfer polymerization, were used for the first time in the field of tissue engineering. Elimination of negative features of P3HB by reinforcement was also aimed. Freeze drying and electrospinning methods were selected for the production of tissue scaffolds. Five percent P3HB and P3HB reinforced with PBA, PNMPBA and PMBA with 2%, 5% and 10% by mass relative to P3HB 3-dimensional (3D) sponge-like tissue scaffolds were obtained by freeze-drying method. Mechanical compression tests were applied on the scaffolds and the compressive strength values were found as 14.00 ± 0.00 kPa and 5.66 ± 1.04 kPa, the elastic modulus values were found as 27.40 ± 0.00 kPa and 12.06 ± 3.20 kPa for the P3HB and 2% PBA scaffolds, respectively. X-ray diffraction (XRD) analysis was performed to investigate the effect of the reinforcement on the crystal structure of P3HB. The crystallinity of the P3HB is found as 52.70%; and the crystallinity of the 2% PBA, PNMPBA and PMBA doped groups were 48.84%, 42.50% and 46.84%; the crystallinity of the 5% PBA, PNMPBA and PMBA doped groups was 46.46%, 46.01% and 46.61%, the crystallinity of 10% PBA, PNMPBA and PMBA doped groups were 36.91%, 41.21% and 46.27%, respectively. According to the characterization results, it has been determined that the scaffolds which produced with freeze-drying method were not suitable for tissue engineering applications. Two-dimensional (2D) and 3D fibrous tissue scaffolds were obtained by using conventional electrospinning and wet electrospinning techniques, respectively. For conventional electrospinning, 5% (w/v) P3HB and 10% PBA (w/w) were dissolved in hexafluoroisopropanol (HFIP) and 15 kV voltage, 22.5 cm syringe-collector distance and 1.0 mL/h flow rate have been determined as convenient conditions after optimization of electrospinning conditions. For wet electrospinning, 10% (w/v) P3HB and 10% (w/v) PBA were dissolved in HFIP and 15 kV voltage, 16.5 cm syringe-collector distance and 1 mL/h flow rate conditions were selected after optimization of the electrospinning conditions. Diameters of 5% P3HB and 10% PBA doped P3HB fibers produced by conventional electrospinning method were 4.44±1.81 μm and 3.21±1.82 μm, and the diameters of 10% P3HB and 10% PBA doped P3HB fibers produced by wet electrospinning were 5.08±1.57 μm and 5.12±1.63 μm, respectively. Tensile tests were performed on 2D fibrous matrices produced by conventional electrospinning. The tensile strengths of the scaffolds were found as 4.00 MPa and 8.20 MPa, while the elastic modulus values were found as 152.00 MPa and 290.00 MPa, respectively. In addition, water contact angle analysis was performed on 2D fiber scaffolds. The water contact angles of the scaffolds were found 88.30±19.90° for 5% P3HB and 48.90±9.00° for P3HB with 10% PBA, respectively. Water uptake capacity measurements were performed with tissue scaffolds produced by wet electrospinning. The water uptake capacities of three dimensional fibrous tissue scaffolds were found 356.70±25.00% for 10% P3HB and 443.60±101.10% for P3HB doped 10% PBA. Cell culture studies were carried out with P3HB and PBA doped P3HB fibrous 3B scaffolds and MC3T3-E1 preosteoblastic cell line to examine cellular compatibility. For determining cell viability and cell morphology mitochondrial viability assay (MTT) and scanning electron microscope (SEM) analysis were applied, respectively. In conclusion, it has been found that PBA doping enhance P3HB’s some negative properties in terms of tissue engineering such as high crystallinity, high hydrophobicity and it improves cellular compatibility by adding positive mechanical aspects.