Optimization of chitosan-hydroxyapatite-based bioinks for potential 3d bioprinting application in bone tissue regeneration

  • Ana Helena Alves da Costa (Student)

Student thesis: Master's Thesis


With the passing of the years, scientific evolution has contributed many times to medicine by providing solutions to, what seemed to be, life-long problems for patients. Not all health problems can be fixed with traditional treatments, as not all complications are caused by drug treated diseases. This is especially true with bone related issues. Age, accidents or birth defects can all be at fault for serious skeletal defects that cannot be treated on their own. This necessity to create further solutions to aid in medicine, arose an interest in the scientific community. Fast forward to this day and age, we can now count on a vast number of options to help in bone repair and regeneration, particularly 3D bioprinted scaffolds. In this study, the main goal was to create and optimize a biomaterial that could be used as an ink to create 3D bioprinted structures for future bone regeneration application. The material used consisted of a chitosan (Ch) -based hydrogel with the incorporation of hydroxyapatite (HA), which is known as the most common calcium phosphate present in human bone. Synthetic HA (SYN_HA) was synthesized and showed to be a suitable material for hydrogel preparation; natural HA (NAT_HA) from tuna fish bones, however, also looked as a very interesting alternative, although it still requires some additional work. In the future, with optimized preparation conditions, it would be more sustainable option for patients. Several tests were performed on the hydrogel, to optimize its composition, in particular the most appropriate concentration of HA to include in the hydrogel itself. Tests on the nanocrystals to assess their purity and crystallinity as well the particle size were performed. These were followed by experiments to compare hydrogels with two final HA percentages: 0.125% wt/vol and 0.25% wt/vol. The following analysis were performed: a stability test to study in vitro hydrogel degradation kinetics under physiological conditions; pH, homogeneity and polymerization assessments; a compression test to evaluate mechanical properties of the hydrogel, particularly the stiffness; a bioprintability test, by a hand-extrusion method, that assessed its capability to form fibers without merging, to form 3D constructs. All of these were pre-conditions to determine if they would behave well as a biomaterial ink and for future use on cell viability trials. The ultimate test was the bioprinting of the material where the parameters were optimized, to guarantee a successful reproduction of the printed structures every time. Results showed that the biomaterial with the best concentration of SYN_HA was the 0.125% wt/vol one, as it had the most satisfactory results in all tests performed, confirming it can be used for bioprinting purposes.
Date of Award17 Feb 2023
Original languageEnglish
Awarding Institution
  • Universidade Católica Portuguesa
SupervisorClara Piccirillo (Supervisor) & Francesca Scalera (Co-Supervisor)


  • Bioink
  • Bioprinting
  • Bone regeneration
  • Chitosan-based hydrogel
  • Hydroxyapatite


  • Mestrado em Engenharia Biomédica

Cite this