**3.9 Antioxidant properties of hydrogel wound dressings**

In this work, % scavenging capacity of the plain film sample Ch/CD(0) and carbon dots loaded samples Ch/CD(1), Ch/CD(2), Ch/CD(3) and Ch/CD(4) for the various free radicals i. e. DPPHR, SOR and HR are shown in **Figure 9**. It can be seen

#### **Figure 8.**

*Percent scavenging for DDPH, superoxide and hydroxyl free radicals by various film samples.*

#### **Figure 9.**

*Images showing completely undamaged L929 cells after 24 h contact with extract of film samples (a) Ch/CD (0) and (b) Ch/CD(2).*

that plain chitosan film sample shows fair % Scavenging capacity and it increases slightly due to addition of carbon dots in the film matrix.

Owing to the fair stability, DPPH radical is usually employed to determine the antioxidant or free radical scavenging activity of the wound dressing films. The method involves reduction of methanolic DPPH radical by hydrogen donating antioxidant polymeric film. Topical wound healing formulations with DPPH scavenging ability have been reported to improve skin repair and regeneration [34]. The observed increase may probably be due to the contribution of –COOH groups

**107**

*Wound Dressing Application of Ch/CD Nanocomposite Film*

dots into chitosan film matrix improves its antioxidant property.

considered main factor for delayed healing of chronic wounds.

The adhesion of a wound dressing film on the wounded skin is a significant parameter and requires a perfect balance between the adhesion capacity of the dressing film and comfort level of patient. In case, the film has a very strong adhesion tendency, it may cause discomfort and pain during removal of the dressing. However, its poor adhering property may also be uncomfortable from the point of view of wound healing. In this work, maximum detachment force (Fmax) required to detach the film from mucosal surface, was determined for all the film samples namely, Ch/CD(0), Ch/CD(1), Ch/CD(2), Ch/CD(3) and Ch/CD(4). The values of Fmax were found to

In a report, Fmax value for gum acacia-cl-(poly(HEMA-co-carbopol hydrogel film was found to be 70.20 ± 17.57 m N. The higher value was attributed to the fair hydrophilic nature of the film [39]. In the present work, Fmax values for various CDs-loaded chitosan films were relatively low, probably due to the presence of functionalized carbon dots within the film matrix. The poor water absorption tendency of these films did not induce polymeric chain relaxation and hence there were no new active sites available for exposure to mucus surface. In addition, the physical crosslinks between carboxylate groups present on the carbon dots surface (as discussed earlier) and protonated amino groups on chitosan chains also made

A plausible explanation for mucoadhesion behavior of plain chitosan film Ch/ CD(0) and CDs-loaded sample Ch/CD(2) may be given on the basis of the most commonly proposed Diffusion-Interpenetration Theory [40, 41]. In the case of pure chitosan film sample Ch/CD(0), an appreciable water content within the film matrix causes polymeric chains to relax or un-fold,thus resulting in exposure of new active sites. These polar segments come in contact with mucuschains and get entangled with them. This results in stronger bio-adhesion as shown in **Figure 10(a)**. However, in the case of CDs-loaded sample Ch/CD(2), the film absorbs very small quantity of water due to physical crosslinks and therefore polymeric chains remain folded or non-relaxed. This minimizes the interaction with mucus chains as

be 88.22 ± 11.52, 45.15 ± 8.61, 43.29 ± 7, 44.25 ± 6.97, and 46.22 ± 5.94mN.

the chitosan chains rigid and prevented them from relaxation.

shown in **Figure 10(b)**. As a result, value of Fmax is quite low.

**3.10 Ex-vivo mucoadhesion studies of films**

present on the surface of carbon dots towards reduction of DPPH radicals. It is here also worth mentioning that chitosan has already been reported to have free radicals scavenging activity [35]. Therefore it appears that addition of functionalized carbon

Hydroxyl, an oxygen centered radical, attacks proteins, DNA, polyunsaturated fatty acid in membranes, and most biological molecules with which it comes in contact [36]. It abstracts hydrogen atoms from membrane lipids [37] and brings about peroxidic reaction of lipids. As per reports [38], the scavenging action of chitosan against hydroxyl radicals may probably be due to following reactions (i) The hydroxyl groups, present within the chitosan macromolecular chains, may react with. OH via hydrogen abstraction, and (ii). OH can react with the residual free amino groups NH2 to form stable macromolecule radicals. Finally, when superoxide anions come in to contact with a biomolecule, they damage it directly or indirectly by forming H2O2, ˉOH, peroxy nitrite or singlet oxygen during aging and pathological events such as ischemic reperfusion injury. It is also reported that Super oxide radicals initiate lipid peroxidation [39]. Therefore, scavenging of superoxide radicals by chitosan based wound dressing might be helpful for preventing delayed wound healing induced by superoxide radicals in pathological conditions. The scavenging of SOR will also diminish formation of hydroxyl radicals, which is

*DOI: http://dx.doi.org/10.5772/intechopen.95107*

#### *Wound Dressing Application of Ch/CD Nanocomposite Film DOI: http://dx.doi.org/10.5772/intechopen.95107*

present on the surface of carbon dots towards reduction of DPPH radicals. It is here also worth mentioning that chitosan has already been reported to have free radicals scavenging activity [35]. Therefore it appears that addition of functionalized carbon dots into chitosan film matrix improves its antioxidant property.

Hydroxyl, an oxygen centered radical, attacks proteins, DNA, polyunsaturated fatty acid in membranes, and most biological molecules with which it comes in contact [36]. It abstracts hydrogen atoms from membrane lipids [37] and brings about peroxidic reaction of lipids. As per reports [38], the scavenging action of chitosan against hydroxyl radicals may probably be due to following reactions (i) The hydroxyl groups, present within the chitosan macromolecular chains, may react with. OH via hydrogen abstraction, and (ii). OH can react with the residual free amino groups NH2 to form stable macromolecule radicals. Finally, when superoxide anions come in to contact with a biomolecule, they damage it directly or indirectly by forming H2O2, ˉOH, peroxy nitrite or singlet oxygen during aging and pathological events such as ischemic reperfusion injury. It is also reported that Super oxide radicals initiate lipid peroxidation [39]. Therefore, scavenging of superoxide radicals by chitosan based wound dressing might be helpful for preventing delayed wound healing induced by superoxide radicals in pathological conditions. The scavenging of SOR will also diminish formation of hydroxyl radicals, which is considered main factor for delayed healing of chronic wounds.

#### **3.10 Ex-vivo mucoadhesion studies of films**

The adhesion of a wound dressing film on the wounded skin is a significant parameter and requires a perfect balance between the adhesion capacity of the dressing film and comfort level of patient. In case, the film has a very strong adhesion tendency, it may cause discomfort and pain during removal of the dressing. However, its poor adhering property may also be uncomfortable from the point of view of wound healing. In this work, maximum detachment force (Fmax) required to detach the film from mucosal surface, was determined for all the film samples namely, Ch/CD(0), Ch/CD(1), Ch/CD(2), Ch/CD(3) and Ch/CD(4). The values of Fmax were found to be 88.22 ± 11.52, 45.15 ± 8.61, 43.29 ± 7, 44.25 ± 6.97, and 46.22 ± 5.94mN.

In a report, Fmax value for gum acacia-cl-(poly(HEMA-co-carbopol hydrogel film was found to be 70.20 ± 17.57 m N. The higher value was attributed to the fair hydrophilic nature of the film [39]. In the present work, Fmax values for various CDs-loaded chitosan films were relatively low, probably due to the presence of functionalized carbon dots within the film matrix. The poor water absorption tendency of these films did not induce polymeric chain relaxation and hence there were no new active sites available for exposure to mucus surface. In addition, the physical crosslinks between carboxylate groups present on the carbon dots surface (as discussed earlier) and protonated amino groups on chitosan chains also made the chitosan chains rigid and prevented them from relaxation.

A plausible explanation for mucoadhesion behavior of plain chitosan film Ch/ CD(0) and CDs-loaded sample Ch/CD(2) may be given on the basis of the most commonly proposed Diffusion-Interpenetration Theory [40, 41]. In the case of pure chitosan film sample Ch/CD(0), an appreciable water content within the film matrix causes polymeric chains to relax or un-fold,thus resulting in exposure of new active sites. These polar segments come in contact with mucuschains and get entangled with them. This results in stronger bio-adhesion as shown in **Figure 10(a)**.

However, in the case of CDs-loaded sample Ch/CD(2), the film absorbs very small quantity of water due to physical crosslinks and therefore polymeric chains remain folded or non-relaxed. This minimizes the interaction with mucus chains as shown in **Figure 10(b)**. As a result, value of Fmax is quite low.

**Figure 10.**

*Bio adhesion in the film samples Ch/CD(0) and Ch/CD(2).*
