Freeze–Thawing-Induced Macroporous Catechol Hydrogels with Shape Recovery and Sponge-like Properties

Catechol-containing hydrogels have been exploited in biomedical fields due to their adhesive and cohesive properties, hemostatic abilities, and biocompatibility. Catechol moieties can be oxidized to o-catecholquinone, a chemically active intermediate, in the presence of oxygen to act as an electrophile to form catechol-catechol or catechol-amine/thiol adducts. To date, catechol cross-linking chemistry to fabricate hydrogels has been mostly performed at room temperature. Herein, they report large increases in catechol cross-linking reaction kinetics by the freeze–thawing process. The formation of ice crystals during freezing steps spatially condenses catechol-containing polymers into nearly frozen (yet unfrozen) regions, resulting in decreases in the polymeric chain distances. This environment allows great increases in catechol cross-linking kinetics, a phenomenon that can also occur during thawing steps. The increased cross-linking rate and spatial condensation in the cryogels provide unique wall and pore structures, which result in elastic, spongelike hydrogels. The moduli of the cryogels prepared by glycol-chitosan-catechol (g-chitosan-c) were improved by 3–6-fold compared to room temperature-cured conventional hydrogels, and the degree of improvement increased depending on the freezing time and the number of freeze–thawing cycles. Unlike typical cell encapsulations before cross-linking, which have often been a source of cytotoxicity, the macroporosity of cryogels allows nontoxic cell seeding with ease. This research offers a new way to utilize catechol cross-linking chemistry by freeze–thawing processes to simultaneously regulate mechanical strength and porous structures in catechol-containing hydrogels.

Catechol is a widely used moiety that functionally represents adhesive properties originating from the adhesion pads of mussels. At the interface, catechol exhibits adhesion functions with surprising adaptability in surface binding chemistry. Upon the interaction of catechol with noble metals, metal oxides, ceramics, or synthetic or natural polymers, catechol is able to dynamically change binding mechanisms using coordination and hydrogen bonding. In bulk, catechol exhibits cross-linking functions via catechol-quinone oxidation, resulting in the formation of catechol-to-catechol adducts (e.g., hyaluronic acid-catechol, polyethylene glycol-catechol, and alginate-catechol). Another catechol cross-linking chemistry is the formation of a catechol-to-amine adduc via a Michael addition or Schiff-base reaction (e.g., chitosan-catechol. Representatively, their group has mainly studied chitosan-catechol. This adduct has improved water solubility by catechol conjugation and is spontaneously cross-linked under aerobic and mildly alkaline conditions to form a hydrogel. Since it has water-resistant adhesive properties as well as protein binding abilities due to the above mechanisms, it has been found to be a hemostatic needle and a swab, gels with thermosensitive hemostatic properties, and intraperitoneal patches.

Schematic Representations of g-Chitosan-c Hydrogel (Left) by Conventional Methods and Cryogel (Right) Formation by Freeze-Thawing Methods (i.e., Cryogelation)

Preparation of the g-chitosan-c cryogels. (a) The chemical structure of the catechol-conjugated glycol chitosan by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC)-mediated coupling. (b) Demonstration of the gel preparation process (top section) and gelation of g-chitosan (bottom section) with (second column) or without (first column) catechol conjugation. The solutions shown in the top row experienced freeze–thawing steps (freezing: 4 h, thawing: 4 h, repetition: 3 times), and those shown in the bottom row were cured at room temperature. The black arrows indicate the scraping direction with a spatula or forceps. The red arrowheads show brittle points after spatula movement. The photos in the dark blue colored box show elasticity of g-chitosan-c cryogel (a relaxed state for the top and an extended one for the bottom). (c) The force–extension curve of the room temperature-cured hydrogel (black) and the cryogel (blue). A black line indicates the hydrogel, and the blue line indicates cryogel. (d, e) Rheological analysis of g-chitosan-c at room temperature after (c) 12 h of gelation time or (d) 24 h of gelation time. G′ indicates the storage modulus (filled circles), and G′′ indicates the loss modulus (open circles).

The aforementioned catechol-to-catechol and/or catechol-to-amine cross-linking is advantageous because no chemical or biological catalysts or external stimuli, such as UV, are required; it is a spontaneous reaction. Dissolved oxygen acts as an oxidant initiator by taking up electrons from catechol, which, in turn, becomes o-catecholquinone. This electrophile is a reactive species that forms catechol adducts to generate hydrogels. However, catechol-mediated cross-linking is rather slow in hydrogel systems. Typically, gelation time takes 24 h or longer, so chemical oxidants such as sodium periodate (NaIO4) have been used.Additionally, not all catechol groups effectively participate in the gelation reactions, exhibiting weak mechanical properties. Their rationale is that the spontaneous yet slow and inefficient cross-linking can be overcome by freeze–thawing processes. The formation of ice crystals during freezing spatially condenses polymer chains into a nearly frozen solution phase resulting in “proximity effects” for rapid catechol-mediated cross-linking. Thus, local concentrations of polymers during the freezing step become abnormally high, in which oxidized o-catecholquinones might have dramatically increased chances of cross-linking. Similar environments for cross-linking are provided during a thawing step.