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Bioink for Reconstruction of Rigid-living Systems
replacements, bone regeneration, drug delivery, and even into two main parts. The initial one is the biopolymer
exciting uses for molecular gastronomy. Gelatin usually base that works as a binder and crosslinking material.
works as a viscous platform to bind other elements of The second is the bioceramics side that will reinforce the
interest. paste and mimic the paste hard-living structure, a standard
Nevertheless, to expand the functionality of these coral. The materials used include GelMA (synthesized),
materials in tissue engineering, the methacrylation gelatin from porcine skin (Sigma-Aldrich), alginic acid
reaction has been studied; in this case, the (Merck), PEGDA (Sigma-Aldrich), lithium phenyl-2,4,6
functionalization of gelatin can be photo-crosslinked by trimethyl-benzoyl phosphinate (Sigma-Aldrich),
different wavelengths, depending on the photoinitiator milli-Q water, hydroxyapatite (Sigma-Aldrich), calcium
(PI) used. This physicochemical improvement has carbonate (Sigma Aldrich), dimethyl sulfoxide (DMSO)
permitted the usage of novel biofrabrication techniques . (Sigma-Aldrich), Dialyzer Maxi, molecular weight cut
[4]
Besides applications in wound dressings and hard off (MWCO) 12 – 14 kDa (Merck), 0.22 µm bottle top
tissues, cartilages or bones have been implemented with vacuum filter (Corning), DMSO for Nuclear Magnetic
gelatin methacrylate (GelMA), with reinforcements with Resonance (NMR) (Sigma-Aldrich), phosphate-buffered
bioceramics as hydroxyapatite and other sort of inorganic saline (PBS), pH 7.4 (Sigma-Aldrich), and syringe pump
particles [5,6] . In addition, poly (ethylene glycol diacrylate) (Harvard Apparatus).
(PEGDA) has been widely implemented due to its fast
end effective crosslinking behavior, which can work as 2.1. High GelMA (H-GelMA) synthesis
a complement to other photo-cross-linkable polymers . For a high degree of methacrylation of GelMA (Figure S1),
[7]
Innovation should not be stuck in just a particular that is, 10 g of gelatin, 100 mL 1 × PBS (sterile) was
direction; conversely, there are other biological issues added. The mixture was dissolved with the aid of a
that our world is currently facing. Therefore, it is crucial heating plate (~240 rpm at 50°C). 8 mL of methacrylic
to take action to adopt eco-friendly applications that could anhydride was added dropwise with the assistance of a
counteract problems derived from climate change. Thus, syringe pump and let emulsion rotate (240 rpm) at 50°C
we aim to expand tissue engineering applications into a for 2 h. 100 mL of sterile PBS (50°C) was preheated to
broader range of goals in this project. For instance, one of dilute GelMA solution for 10 min at 50°C. The dialysis
the most significant burdens from the environmental and membrane (MWCO 12-14,000 kDa) was prepared at
marine sciences is coral bleaching, that in other words can 40°C, and GelMA solution was inserted inside them;
be considered the disruption of a symbiosis that consists dialysis was allowed to go on for a minimum of 5 days
of a robust and rigid system of calcium carbonate with the with constant stirring. The water was changed from 1
living beings, which are mainly species of polyps derived to 2 times a day to eliminate the excess methacrylate
from heat wave and changes in the marine ecosystem ions and dispose of the residue in a regulated container.
that bleaches the colonies and have strong effects in the A sterile vacuum (0.22 µm) filtration cup was used to
marine biota [8,9] . For this reason, many groups around filter the liquid. Sterilized polymer was transferred into
the world has started to take an interest in creating and Falcon tubes. Semi-closed tubes were submerged in
developing new formulations and taking advantage of liquid nitrogen and freeze-dried for at least 5 days to get a
new innovative materials to counteract environmental sponge-like freeze-dried GelMA sample .
[12]
problems as a way of preventing more extensive
problems that will jeopardize the lives of human beings in 2.2. Biopolymer-base preparation
the future [10,11] . Therefore, in this project, biopolymers as Depending on the volume required to prepare and the
gelatin, alginate, gelatin methacrylate (GelMA), and poly percentages stated in Table 1, freeze-dried H-GelMA,
(ethylene glycol diacrylate) (PEGDA) are reinforced with gelatin, alginate, and PEGDA were dissolved in Milli-Q
bioceramics as calcium carbonate and hydroxyapatite. with constant stirring to dissolve the final solution.
This unique formulation can assist the growth of hard- Using a heath bath is recommended to melt the solution
living systems, like corals, as an innovative technology at a temperature of the maximum of 50°C. A higher
that ionic/photo-crosslink, which makes it adaptable to temperature can modify the molecular behavior of the
new 3D manufacturing technologies and can withstand four polymer chains and jeopardize the accuracy of
under wet conditions (Figure 1). printability. Then, the PI was added (Lithium phenyl-2,4,6
2. Materials and methods trimethylbenzoylphosphinate) to the previous solution. To
avoid the interaction with light that triggers crosslinking
The following materials and reactants are necessary for reaction, it is recommended to cover it with aluminum
obtaining the biopolymer base and bioceramics to develop foil. This base can be kept at −20°C for more extended
the final paste and subsequently realize the fundamental periods if there is no interaction with light that could
characterization. The final formulation is homogenized trigger gelation.

66 International Journal of Bioprinting (2021)–Volume 7, Issue 4