Carboxymethyl Chitosan/Sodium Alginate/Chitosan Quaternary Ammonium Salt Composite Hydrogel Supported 3J for the Treatment of Oral Ulcer

Abstract

Oral ulcer is a common inflammatory disease of oral mucosa, causing severe burning pain and great inconvenience to daily life. In this study, compound 3J with anti-inflammatory activity was synthesized beforehand. Following that, an intelligent composite hydrogel supported 3J was designed with sodium alginate, carboxymethyl chitosan, and chitosan quaternary ammonium salt as the skeleton, and its therapeutic effect on the rat oral ulcer model was investigated. The results show that the composite hydrogel has a dense honeycomb structure, which is conducive to drug loading and wound ventilation, and has biodegradability. It has certain antibacterial effects and good anti-inflammatory activity. When loaded with 3J, it reduced levels of TNF-α and IL-6 in inflammatory cells by up to 50.0%. It has excellent swelling and water retention properties, with a swelling rate of up to 765.0% in a pH 8.5 environment. The existence of a large number of quaternary ammonium groups, carboxyl groups, and hydroxyl groups makes it show obvious differences in swelling in different pH environments, which proves that it has double pH sensitivity. It is beneficial to adapt to the highly dynamic changes of the oral environment. Compared with single hydrogel or drug treatment, the drug-loaded hydrogel has a better effect on the treatment of oral ulcers.

Keywords: carboxymethyl chitosan; chitosan quaternary ammonium salt; hydrogel; oral ulcer; sodium alginate.

Figure 1

The ¹H NMR results of 3J. The green vertical line indicates that the peak is integrated, and the blue number is the size of the peak area accumulated by the corresponding peak.

Figure 2

SEM morphological characteristics of composite hydrogels. (A) AmCS−SA hydrogel form under 2.0 microns. (B) AmCS−SA hydrogel form under 20.0 microns. (C) CMCS−SA−AmCS−SA hydrogel form under 2.0 microns. (D) CMCS−SA−AmCS−SA hydrogel form under 20.0 microns. (E) Standard curve of 3J.

Figure 2

SEM morphological characteristics of composite hydrogels. (A) AmCS−SA hydrogel form under 2.0 microns. (B) AmCS−SA hydrogel form under 20.0 microns. (C) CMCS−SA−AmCS−SA hydrogel form under 2.0 microns. (D) CMCS−SA−AmCS−SA hydrogel form under 20.0 microns. (E) Standard curve of 3J.

Figure 3

FT-IR spectra of CMCS, AmCS, SA, 3J, AmCS−SA, CMCS−SA−AmCS−SA, and CMCS−SA−AmCS−SA−3J.

Figure 4

Characterization of swelling and water retention of hydrogels. (A) Swelling properties of CMCS−SA−AmCS−SA hydrogel (mean ± SD, n = 3). (B) Swelling properties of AmCS−SA hydrogel (mean ± SD, n = 3). (C) AmCS−SA dissolution results within 1 h. (D) Water retention of CMCS−SA−AmCS−SA and AmCS−SA (mean ± SD, n = 3).

Figure 5

Degradation rate characterization of (A,B) CMCS−SA−AmCS−SA hydrogel and (C,D) AmCS−SA hydrogel.

Figure 6

Rheological characterization of hydrogels. (A) Surface morphology imaging of AmCS−SA hydrogel under atomic force microscope (AFM). (B) Surface morphology imaging of CMCS−SA−AmCS−SA hydrogel under AFM. (C,D) The determination process of Young’s modulus of (C) AmCS−SA hydrogel and (D) CMCS−SA−AmCS−SA hydrogel.

Figure 7

In vitro release of free 3J and 3J loaded CMCS−SA−AmCS−SA in a PBS buffer solution (pH 7.4) at 37 °C for 48 h (mean ± SD, n = 3).

Figure 8

Evaluation of the antibacterial effect of hydrogels. (A) Survival ratio was calculated via OD₆₀₀ for S. aureus and E. coli (mean ± SD, n = 3). (B) Images of survival bacteria clones after the treatment of hydrogels.

Figure 9

Anti-inflammatory properties of hydrogels and 3J in vitro. (A) IL-6 and (B) TNF-α were detected by an enzyme-linked immunosorbent assay (ELISA) (mean ± SD, n = 3) vs. LPS, * p < 0.05, ** p < 0.01, *** p < 0.001; vs. control, ### p < 0.001.

Figure 10

Analysis of the effect of treating oral ulcer in vivo. (A) Changes of ulcer area in each group at 0−6 days after establishing oral ulcer model induced by phenol (mean ± SD, n = 8). (B) Quantitative results of oral ulcer area at different time points (mean ± SD, n = 8), * p < 0.05, *** p < 0.001. (C) HE staining of the oral mucosa of SD rats (magnification 100×).

Figure 10

Analysis of the effect of treating oral ulcer in vivo. (A) Changes of ulcer area in each group at 0−6 days after establishing oral ulcer model induced by phenol (mean ± SD, n = 8). (B) Quantitative results of oral ulcer area at different time points (mean ± SD, n = 8), * p < 0.05, *** p < 0.001. (C) HE staining of the oral mucosa of SD rats (magnification 100×).

Figure 11

The synthetic route of 3J.