Development and characterization of a photocurable alginate bioink for three-dimensional bioprinting

Abstract

Alginate is a biocompatible material suitable for biomedical applications, which can be processed under mild conditions on irradiation. This paper investigates the preparation and the rheological behavior of different pre-polymerized and polymerized alginate methacrylate systems for three-dimensional photopolymerization bioprinting. The effect of the functionalization time on the mechanical, morphological, swelling, and degradation characteristics of cross-linked alginate hydrogel is also discussed. Alginate was chemically-modified with methacrylate groups and different reaction times considered. Photocurable alginate systems were prepared by dissolving functionalized alginate with 0.5- 1.5% w/v photoinitiator solutions and cross-linked by ultraviolet light (8 mW/cm² for 8 minutes).

Keywords: 3D bioprinting; Alginate hydrogel; Functionalization; Photopolymerization; Rheology.

Figure 1

Alginate showing a linkage between the mannuronic and guluronic acid[25].

Figure 2

Schematic representation of the photopolymerization process of alginate methacrylate. (a) After exposing the polymer solution to ultraviolet radiation, the photoinitiators generated free radicals that react with the vinyl methylene starting the crosslinking reaction; (b) the reaction propagates with macroradicals reacting with unreacted carbon-carbon double bonds. (c) At the end through a bimolecular termination mechanism, a three-dimensional network of the cross-linked hydrogel is formed [30].

Figure 3

(a) Non-functionalized alginate (1% w/v), (b) functionalized alginate (1% w/v) after 8 h of reaction, (c) functionalized alginate (1% w/v) after 24 h of reaction. The functionalization is confirmed by the presence of new peaks in the spectra at 5.63 ppm and 6.09 ppm attributed to the methylene group and a peak at 1.82 ppm that corresponds to the methyl group.

Figure 4

(a) Non-functionalized alginate (2% w/v), (b) functionalized alginate (2% w/v) after 8 h of reaction, (c) functionalized alginate (2% w/v) after 24 h of reaction. The functionalization is confirmed by the presence of new peaks in the spectra at 5.63 ppm and 6.09 ppm attributed to the methylene group and a peak at 1.82 ppm that corresponds to the methyl group.

Figure 5

(a) Non-functionalized alginate (3% w/v), (b) functionalized alginate (2% w/v) after 8 h of reaction, (c) functionalized alginate (2% w/v) after 24 h of reaction. The functionalization is confirmed by the presence of new peaks in the spectra at 5.63 ppm and 6.09 ppm attributed to the methylene group and a peak at 1.82 ppm that corresponds to the methyl group.

Figure 6

Stress versus shear rate profiles for alginate methacrylate solutions containing different alginate methacrylate concentrations, reacted for 8 h. (a) 1% w/v of alginate, (b) 2% w/v of alginate, (c) 3% w/v of alginate, (d) comparison of all compositions.

Figure 7

Stress versus shear rate for solutions containing different alginate methacrylate concentrations reacted for 24 h. (a) 1% w/v of alginate, (b) 2% w/v of alginate, (c) 3% w/v of alginate, (d) comparison of all compositions.

Figure 8

Viscosity versus shear rate for solutions containing different alginate methacrylate concentrations reacted for 8 h. (a) 1% w/v, (b) 2% w/v, (c) 3% w/v, (d) comparison of all compositions.

Figure 9

Viscosity versus shear rate for solutions containing different alginate methacrylate concentrations reacted for 24 h. (a) 1% w/v, (b) 2% w/v, (c) 3% w/v, (d) comparison of all compositions.

Figure 10

2% wt. alginate solution reacted for 8 h. (a) storage and loss modulus versus strain (b) storage and loss modulus versus frequency, (c) complex modulus versus frequency, (d) tan δ versus frequency.

Figure 11

2% wt. alginate solution reacted for 24 h. (a) storage and loss modulus versus strain, (b) storage and loss modulus versus frequency, (c) complex modulus versus frequency, (d) tan δ versus frequency.

Figure 12

2% wt. methacrylate alginate with different concentration of VA-086 photoinitiators functionalized for 8 h (a) 0.5% w/v of VA-086, (b) 1% w/v of VA-086, (c) 1.5% w/v of VA-086, d) 0.05%.

Figure 13

2% wt. methacrylate alginate with different concentration of VA-086 photoinitiators functionalized for 8 h (a) 0.5% w/v of VA-086, (b) 1% w/v of VA-086, (c) 1.5% w/v of VA-086, (d) 0.05%.

Figure 14

2% wt. methacrylate alginate hydrogel with different concentration of VA-086 functionalized for 8 h. (a) 0.5 w/v % of VA-086, (b) 1 w/v % of VA-086, (c) 1.5 w/v % of VA-086.

Figure 15

2% wt. methacrylate alginate with different concentration of VA-086 functionalized for 24 h. (a) 0.5 w/v % of VA-086, (b) 1 w/v % of VA-086, (c) 1.5 w/v % of VA-086.

Figure 16

Scanning electron microscopy images of cross-linked methacrylate alginate hydrogel structures obtained from alginate-methacrylate at different reaction times: 8 and 24 h.

Figure 17

Compression tests for alginate methacrylate samples (8 h) with different photoinitiator concentrations (a) 0.5% w/v VA-086, (b) 1% w/v VA-086, (c) 1.5% w/v VA-086.

Figure 18

Compression tests for alginate methacrylate samples (24 h) with different photoinitiator concentrations (a) 0.5% w/v VA-086, (b) 1% w/v VA-086, (c) 1.5% w/v VA-086.

Figure 19

Swelling and degradation rate for 2% wt. functionalized alginate (24 and 8 h) reaction time in (a) distilled water (diH₂O), (b) degradation rate in diH₂O, (c) Dulbecco’s Modified Eagle’s Medium (DMEM), (d) degradation rate in DMEM.