## 3 Chemical Modification of Hyaluronan
### Amidation of Hyaluronic Acid
The chemical modification was performed via EDC/NHS using HMW-HA (8.0 \(\times\) 10\({}^{5}\)-1.2 \(\times\) 10\({}^{6}\) g/mol). The degree of substitution was calculated to be 22% \(\pm\) 2.8% by NMR. Three formulations with different crosslinking densities (30, 60, and 90%, respectively) were prepared by varying the FeCl3 content. PRP was encapsulated and delivered via the hydrogel. The HA-DOPA/PRP hydrogel showed adhesive strength as a function of crosslinking density. The hydrogels exhibited a bacteriostatic effect with a killing ratio larger than 95% for both _E. coli_ and _S. aureus_. The in vitro anti-inflammation experiments were carried out by using lipopolysaccharideaccharide (LPS) over RAW264.7 macrophages. The expression of TNF-\(\alpha\), IL-6, and IL-1\(\beta\) was significantly up-regulated, while IL-10 was down-regulated [79]. [PERSON] et al. described the Schiff base bond formation using oxidized dextran (Ox-DEX) and HA (2.0 \(\times\) 10\({}^{5}\) g/mol) modified with an antimicrobial peptide (AMP = SWLSKTAKKLTFKKIPKKKIPKKKRFPRPRPRPWRPNMI-NH\({}_{2}\)) by amidation mediated by EDC/NHS for the loading of PRP [80]. Adipose-derived stem cells (ADSCs) have raised broad interest in therapeutic applications for wound healing [81]. [PERSON] et al. developed an HA-based patch after chemical modification of HA (2 \(\times\) 10\({}^{5}\) g/mol) using DOPA mediated by EDC/NHS (pH 5.0). The authors reported a substitution degree (DS) of up to 8%. The derivative was crosslinked by oxidizing with NaIO\({}_{4}\) under basic conditions. The patches were tested in vivo using a C57 BL/6 male mice model (7 weeks old, 23-26 g). Fluorescently labelled ADSCs (5 \(\times\) 10\({}^{5}\) cells/100 uL) were transplanted into healthy tissues at the wound boundary or deposited at the HA-CA patch at the wound site. The wound area was examined, as well as collagen content, granulation tissue thickness and vascularity, cell apoptosis, and re-epithelialization. The epidermis was regenerated, and skin thickness was restored in mice treated with the scaffold [82]. [PERSON] et al. prepared a nanocomposite hydrogel by loading 0.1 wt.% of graphene oxide (GO) in HA-tyramine derivative combined with gelatin grafted with gallic acid (GGA). The first step consisted of the amidation of HA (un-reported MW) using the EDC/NHS activation reaction. The authors reported a grafting ratio of 4.7% for HA and 7.9% for gelatin. The HA-Tyr/GGA/GO hydrogels presented a porous, crosslinked structure and good stability, biocompatibility, antioxidant, and hemostatic properties. The hydrogel presented photothermal antibacterial activity against _E. coli_ and _S. aureus_, which accelerated healing [83]. However, hydrogels made of aromatic substances usually do not interact well with tissues or biodegradable in vivo. The amidation of the carboxyl group in HA can also be mediated by DMTMMM in water. [PERSON] et al. prepared an HA/AgNPs/gentamicin nanocarrier. HA (Mw = 2.35 \(\times\) 10\({}^{6}\) g/mol) was chemically modified by amidation using DMTMM as an activator for reacting with 3 aminophenylboronic acid (3-APB). The derivative was used for the encapsulation of silver and gentamycin for the construction of disinfect coatings (HA-3 APB/Ag/gentamycin). The scaffold exhibited strong inhibition of the growth and adhesion of bacteria without affecting cell attachment and proliferation. The in vivo results indicated accelerated wound healing on infected wounds with _S. aureus_[84]. However, the Mw of modified HA was unreported. Conditioned medium secreted by stem cells exerts protective effects through stimulation of angiogenesis and improvement of tissue function [85]. [PERSON] et al. prepared double modification of HA. First, HA (3.4 \(\times\) 10\({}^{5}\) g/mol) was esterified with methacrylic anhydride at low temperature (0-4 \({}^{\circ}\)C) with degrees of substitution (DS) of 21, 39, and 75%, respectively.