Improved and Highly Reproducible Synthesis of Methacrylated Hyaluronic Acid with Tailored Degrees of Substitution

Methacrylated hyaluronic acid (HAMA) is a versatile material that has gained significant attention in various pharmaceutical and biomedical applications. This biocompatible material can be photo-cross-linked in the presence of Irgacure 2959 (I2959) to produce hydrogels. Controlling the degree of methacrylation (DM) is crucial since it plays a pivotal role in determining the properties and thus the potential applications of the gels. We report herein a new green approach for the highly controlled and tailored modification of hyaluronic acid (HA) with methacrylic anhydride (MA). The reaction conditions of previously reported procedures were optimized, leading to a decreased reaction time (3 h instead of 24 h) and consumption of fewer equivalents of MA (5 equiv instead of 20) and water as the sole solvent. By changing the amount of base added, HAMA with three different DMs was obtained: 19, 35, and 60%. The influence of the molecular weight of HA, degree of substitution, and concentration of the HAMA solution prior to photo-cross-linking on the rheological, swelling, and degradation properties of HAMA hydrogels was also studied in this work.

Table S5.Summary of the conditions for the screening reactions of HA with 1, 5 or 10 eq of MA under stirring in dark conditions.NaOH (1 M, 2 equiv./addition)was added every 30 minutes; 4 additions in total.

Figure S2 .
Figure S2.Rheological response of HAMA gels of different DM, concentration and molecular weight (A: 8-15 kDa, B: 40-50 kDa and C: 80-100 kDa) to frequency sweep (left) at a constant shear strain (oscillating) of 10% and increasing frequency from 0.1 to 100 Hz, and amplitude sweep (right) constant shear strain (oscillating) of 10% and constant frequency at 1 Hz.Disk shaped hydrogels of 25 mm diameter and 10 mm height were prepared at different polymeric concentrations (3%w and 1%w) in PBS containing 0.3%w I2959.

Figure S3 .
Figure S3.Rheological response of HAMA gels of different DM, concentration and molecular weight (A: 200-500 kDa and B: 600-1000 kDa) to frequency sweep (left) at a constant shear strain (oscillating) of 10% and increasing frequency from 0.1 to 100 Hz, and amplitude sweep (right) constant shear strain (oscillating) of 10% and constant frequency at 1 Hz.Disk shaped hydrogels of 25 mm diameter and 10 mm height were prepared at different polymeric concentrations (2%w, 1%w and 0.5%w) in PBS containing 0.3%w I2959.

Table S1 .
Summary of the conditions for the screening reactions of HA (80-100 kDa) with 5 equiv. of MA under stirring in dark conditions.The pH was corrected prior to MA additions.

Table S2 .
Summary of the conditions for the screening reactions of HA (80-100 kDa) with 5 or 8.3 equiv. of MA under stirring in dark conditions.The pH was corrected prior to MA additions.

Table S3 .
Summary of the conditions for the screening reactions of HA with 1, 5 or 10 equiv. of MA under stirring in dark conditions.NaOH (1M, 1 equiv./addition)was added every 30 minutes; 4 additions in total.

Table S4 .
Summary of the conditions for the screening reactions of HA with 1, 5 or 10 equiv. of MA under stirring in dark conditions.

Table S5 .
Summary of the conditions for the screening reactions of HA with 1, 5 or 10 equiv. of MA under stirring in dark conditions.NaOH (1 M, 2 equiv./addition)was added every 30 minutes; 4 additions in total.

Table S6 .
Summary of the conditions for the screening reactions of HA (80-100 kDa) with1, 2, 3, 4 or 5 equiv. of MA under stirring in dark