Figure 2.

Schematic representation of the sample preparation process for nuclear magnetic resonance (NMR) analysis of the hyaluronic acid (HA) filler sample, which includes the preparation of phosphate-buffered solution (pH 7.0), enzymatic treatment with chondroitinase ABC, incubation in a shaking chamber, filtration through microfilters, and freeze-drying for preservation. For filtration and preservation, the solution is filtered twice using microfilters of different pore sizes (0.45, 0.2, and 0.1 µm) after incubation. The filtered solution is then frozen at −80 °C for 6 h and sealed according to the NMR analysis setup, and the parameters for the HA filler sample are applied. The freeze-dried sample is reconstituted in 600 µL of D₂O solution in a 5 mm NMR tube. The tube is then inserted into an NMR instrument (Ascend™ 600, Bruker) for analysis; the specific parameters used for the ¹H-NMR (32 scans; D1 time, 25 s) and ¹³C-NMR (4096 scans; D1 time, 15 s) experiments are provided. The paraffin is subjected to freeze-drying for 48 h to ensure sample integrity.