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Materials Science in Additive Manufacturing Bioactive hydrogels for 3D bioprinting

viscosity ≥2 Pa·s at 2% in water at 25°C, molecular weight that consisted of square-shaped pores. The print bed
of 200 kDa), bromothymol blue, and anhydrous calcium temperature was set at 19°C, and the biomaterial inks
chloride were purchased from Sigma Aldrich (Missouri, were deposited at a speed of 3600 mm/min for linear
USA). The bioactive borate glass (BBG 1605, microspheres patterns and 1200 mm/min curve patterns. All samples
at particle size <20 µm) was supplied by ETS Wound Care were printed with the same-size nozzles (0.41 mm
(Missouri, USA). Deionized water was used in biomaterial inner diameter, Nordson, USA). The print head nozzle
ink preparation and other experiments. temperature was set at each of 25°C, 30°C, 35°C, and
40°C. Extrusion pressure ranged from 15 to 125 kPa. After
2.2. Biomaterial ink preparation printing 21 layers, the printed scaffold was immersed in
As described in our previous work, biomaterial ink CaCl solution (1 mol/L) for 10 min to form crosslinks,
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formulations were prepared by alginate, gelatin, and BBG. thereby enhancing the scaffold’s mechanical properties.
The 1605 bioactive borate glass (BBG) utilized in this After cross-linking, the scaffold underwent multiple
research is composed of 51.6 wt% B O , 20 wt% CaO, 6 wt% rinses with deionized water to wash away the remaining
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Na O, 5 wt% MgO, 4 wt% P O , 12 wt% K O, 1 wt% ZnO, and calcium ions. The crosslinked scaffolds were stored at 4°C
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0.4 wt% CuO. This BBG demonstrates high solubility and for further characterization.
rapid degradation compared to silicate glass such as Bioglass
45S5 and 1393 borate glass. First, 1 L of 20 mg/mL glass 2.4. Characterization
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suspension was prepared by dissolving 20 g BBG powder in The crosslinked scaffolds were characterized in this study
1 L of deionized water. The obtained suspension was filtered in terms of their density, chemical structure, rheological
(pore size <0.22 µm) and centrifuged at 2000 rpm for behavior, and mechanical properties.
2 min. The obtained BBG suspension was then diluted with
DI water to obtain 0, 10, 15, and 20 mg/mL concentrations 2.4.1. Density
of BBG. Gelatin powder (500 mg) was added to each BBG The density of each biomaterial ink formulation was
suspension (10 mL each) and stirred at 600 rpm at 40°C, measured by weighing 2 mL batches at five replications.
followed by the addition of 2.5 mg of pH-sensitive dye
(bromothymol blue) to the mixture. After 10 min, 300 mg 2.4.2. Chemical structure
sodium alginate was added to the mixture drop-wise The Fourier transform infrared spectroscopy (FTIR)
and stirred at 1200 rpm for 20 min. The concentration of was conducted using a Nicolet iS50 spectrophotometer
gelatin and alginate was set at 50 mg/mL and 30 mg/mL, (Thermo Scientific, USA), outfitted with a mid-IR range
respectively, as our previous research showed that the 5:3 (4000 – 400 cm ) diamond crystal cell for attenuated total
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gelatin-alginate ratio demonstrates Young’s modulus in reflection. Spectra were acquired at a 4 cm resolution,
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the range of human skin with the best biocompatibility totaling 32 scans per spectrum with a data interval of 0.482
and wound healing outcome. The coding of the samples cm . To prepare for measurement, 500 µL of the hydrogel
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was established based on the weight percentage (wt%) of sample was spread thinly over the crystal to form a film
dry material for each component. In the biomaterial inks, on the splitter area. To maintain accuracy, a new reference
the concentrations of gelatin, alginate, and BBG were set at spectrum was captured every five scans to correct the
5%, 3%, and 0 – 2%, respectively. Then each biomaterial ink subsequent spectra. Analysis of the spectra was performed
sample was poured into 3D printing cartridges (Nordson, using the OMNIC software version 9.2.41 from Thermo
Ohio, USA). Cartridges were centrifuged at 2000 rpm for Fisher Scientific (USA). The results were presented as
2 min and incubated at 37°C for 1 h to debubble the inks. a percentage of transmittance versus the wavenumber
We did not include biomaterial inks with BBG content (cm ). Reference IR spectrum data from Sigma Aldrich
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>20 mg/mL in this research because it was found from were employed to pinpoint the characteristic chemical
our preliminary tests that gelatin-alginate hydrogel inks bonds present in gelatin, alginate, and water.
incorporated with BBG higher than 20 mg/mL were not
extrudable. 2.4.3. Rheological behavior
Dynamic viscosity (µ) was determined at various
2.3. Three-dimensional printing temperatures using a rotational rheometer (Kinexus Ultra+,
The 3D-printed scaffolds were fabricated using an Malvern Panalytical Ltd., U.K.) equipped with a cone and
extrusion-based bioprinter (Inkredible+, CellInk Corp., plate setup (CP4/40) and a lower fixture (PED40) for sample
Sweden), which allows control over printing parameters, handling. The apparatus was set to a 40 mm-diameter cone
including nozzle temperature, layer height, and printing with a 4° angle. A solvent trap was employed to minimize
speed. The G-code defined a scaffold printing pattern sample evaporation. The loading and working gaps were

Volume 3 Issue 1 (2024) 4 https://doi.org/10.36922/msam.2845