Citrate Crosslinked Poly(Glycerol Sebacate) with Tunable Elastomeric Properties

Abstract

Poly(glycerol-sebacate) (PGS) is a biodegradable elastomer known for its mechanical properties and biocompatibility for soft tissue engineering. However, harsh thermal crosslinking conditions are needed to make PGS devices. To facilitate the thermal crosslinking, citric acid is explored as a crosslinker to form poly(glycerol sebacate citrate) (PGSC) elastomers. The effects of varying citrate contents and curing times are investigated on the mechanical properties, elasticity, degradation, and hydrophilicity. To examine the potential presence of unreacted citric acid, material acidity is monitored in relation to the citrate content and curing times. It is discovered that a low citrate content and a short curing time produce PGSC with tunable mechanical characteristics similar to PGS with enhanced elasticity. The materials demonstrate good cytocompatibility with human umbilical vein endothelial cells similar to the PGS control. The research study suggests that PGSC is a potential candidate for large-scale biomedical applications because of the quick thermal crosslink and tunable elastomeric properties.

Keywords: biomaterial; citric acid; elastomers; poly(glycerol sebacate); thermal crosslinking.

Figure 1.

Chemical reaction to form citrate-crosslinked poly(glycerol sebacate) networks. Compared to the PGS alone that requires 12 to 24h for curing, citric acid significantly facilitates esterification with free hydroxyls on the PGS prepolymer to form PGSC networks.

Figure 2.

Tensile tests for PGSC elastomers at varying citrate contents. All PGSC samples are cured for 3 h at 150 °C. PGS-12h and PGS-24h controls are cured for 12 and 24h at the same conditions without citric acid. (A) Representative stress-strain curves. (B) UTS, p < 0.0001. (C) Strain at break, p = 0.0487. (D) E, p < 0.0001. All PGSC samples are quickly crosslinked in 3h. When citrate content increases up to 6 and 8 mol.%, the E and UTS values are comparable to the PGS-12h control. It is, however, difficult to yield a PGSC elastomer as tough as PGS-24h control by only increasing citrate content in crosslinking. * <0.05, ** <0.01, *** < 0.001. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation (n = 5).

Figure 3.

Tensile properties of PGSC and PGS controls at varying curing times. All PGSC samples are fixed at 6 mol.% citrate but varied in curing time. (A) Representative stress-strain curves. (B) UTS, p < 0.0001. (C) Strain at break, p < 0.0001. (D) E, p < 0.0001. By varying curing time from 1.5 to 6h, E values of the PGSC elastomers are elevated from 0.12 ± 0.2 MPa for the PGSC-1.5h to 1.29 ± 0.14 MPa for the PGSC-6h. This modulus range surpasses the range of the PGS controls cured between 12 and 24h, indicating a significant reduction in curing time to mediate the modulus by citric acid crosslinking as compared to PGS alone. * < 0.05, ** < 0.01, *** < 0.001. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation, (n = 5).

Figure 4.

Cyclic tensile tests for PGSC and PGS-24h control with strain between 5% and 50% to examine the elastic performance (n = 3). Representative plots for the cyclic stress-strain curves of (A) PGSC-4%, (B) PGSC-6%, (C) PGSC-8%, and (D) PGS-24h control. All PGSC elastomers could withstand 200 cyclic loading without break and show little hysteresis loops. Whereas, the PGS-24h control undergoes approximately 100 cycles to break at the same strain range. This comparison indicates that all PGSC elastomers with the three different citrate contents demonstrate more durable network structures to dissipate the cyclic loading stress.

Figure 5.

Cyclic tensile tests for the PGSC and PGS-24h control between 5% and 50% strain (n = 3). The curing time for the PGSC samples varies from 1.5 to 6h. Representative plots for the cyclic stress-strain curves of (A) PGSC-1.5h, (B) PGSC-3h, (C) PGSC-4.5h, (D) PGSC-6h and (E) PGS-24h control. (F) The elasticity of the PGSC-6h is further examined between 5% and 20% strain. The PGSC samples cured between 1.5 and 4.5h are more durable to withstand the large deformations compared to the PGS-24h control. When increasing the curing time up to 6h, the PGSC-6h still demonstrates robust elasticity between 5% and 20% strain. These hysteresis test results indicate the curing time varied from 1.5 to 6h are suitable to construct PGSC elastomers for soft tissue engineering applications.

Figure 6.

Water contact angle measurements to evaluate the hydrophilicity among the PGSC elastomers with curing time varied from 3 to 6h versus the PGS-24h control. All PGSC elastomers show a significantly lower water contact angle compared to the PGS-24h control, indicating that the presence of a certain amount of citrate increases the hydrophilicity. p < 0.0001. ** < 0.01, *** < 0.001. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation, (n = 5).

Figure 7.

pH measurements of the solutions extracted from the PGSC samples and PGS controls. (A) PGSC samples crosslinked with citrate contents from 4 to 8 mol.%, p < 0.0001. (B) PGSC with 6 mol.% citrate and curing time varied from 3 to 6h, p < 0.0001. (C) Calibration curve of pH against the citric acid-based carboxylic acid concentrations. * < 0.05, ** < 0.01, *** < 0.001. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation, (n = 3).

Figure 8.

Accelerated degradation of the PGSC elastomers as a function of time in 60 mM NaOH solution at 37 °C. (A) PGSC samples with varying citrate contents. p < 0.0001. (B) PGSC samples with different curing time. p < 0.0001. The PGS cured for 12 and 24h are used as controls. The degradation rate is proportional to the hydrophilicity but inverse to the crosslinking density. These two aspects synergistically affect the PGSC degradation as compared to the PGS controls. * < 0.05, ** < 0.01 and *** < 0.001. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation, (n = 4).

Figure 9.

MTT assay to evaluate the cytocompatibility of PGSC elastomers using HUVEC cells. The cells are incubated on the PGSC layers over 48 h. The normalized MTT values of all PGSC samples remain similar to the PGS and TCPS controls, indicating the cytocompatibility is good as the PGS control. ns, p = 0.8083. p < 0.05 is considered significantly different. Data represent the mean value ± standard deviation, (n = 4).