Green synthesis of silver nanoparticles through oil: Promoting full-thickness cutaneous wound healing in methicillin-resistant Staphylococcus aureus infections

Abstract

Due to the emergence of multi-drug resistant microorganisms, the development and discovery of alternative eco-friendly antimicrobial agents have become a top priority. In this study, a simple, novel, and valid green method was developed to synthesize Litsea cubeba essential oil-silver nanoparticles (Lceo-AgNPs) using Lceo as a reducing and capping agent. The maximum UV absorbance of Lceo-AgNPs appeared at 423 nm and the size was 5-15 nm through transmission electron microscopy result. The results of Fourier transform infrared and DLS showed that Lceo provided sufficient chemical bonds for Lceo-AgNPs to reinforce its stability and dispersion. The in vitro antibacterial effects of Lceo-AgNPs against microbial susceptible multidrug-resistant Escherichia coli (E. coli) and methicillin-resistant Staphylococcus aureus (MRSA) were determined. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of Lceo-AgNPs against E. coli were 25 and 50 μg/ml. The MIC and MBC of Lceo-AgNPs against MRSA were 50 and 100 μg/ml, respectively. The results of scanning electron microscopy showed that the amount of bacteria obviously decreased and the bacteria cells were destroyed by Lceo-AgNPs. In vivo research disclosed significant wound healing and re-epithelialization effects in the Lceo-AgNPs group compared with the self-healing group and the healing activity was better than in the sulfadiazine silver group. In this experiment, Lceo-AgNPs has been shown to have effects on killing multidrug-resistant bacteria and promoting wound healing. This study suggested Lceo-AgNPs as an excellent new-type drug for wound treatment infected with multidrug-resistant bacteria, and now expects to proceed with clinical research.

Keywords: AgNPs; L. cubeba oil; antibacterial; green synthesis; multidrugresistance; wound infection.

FIGURE 1

UV-vis wavelength spectra of Lceo-AgNPs with different ratio of Lceo: AgNO₃ in the range of 300–600 nm (A) and the colors of Lceo, AgNO₃, and Lceo-AgNPs (B).

FIGURE 2

Nanosizer of as-prepared Lceo-AgNPs.

FIGURE 3

Zeta potential of as-prepared Lceo-AgNPs.

FIGURE 4

FTIR absorption spectra of Lceo-AgNPs and Lceo.

FIGURE 5

TEM images of dispersed Lceo-AgNPs.

FIGURE 6

XRD analysis spectrum of Lceo-AgNPs.

FIGURE 7

Time-killing curves of E. coli (A) and MRSA (B) treated with Lceo-AgNPs at the concentrations of MIC and MBC.

FIGURE 8

SEM images of E. coli (A,C) and MRSA (B,D) treated with/without Lceo-AgNPs. (A,C) negative control magnified ×10,000; (B,D) AgNPs-treated bacteria magnified ×10,000.

FIGURE 9

Cytotoxicity of Lceo-AgNPs and AgNPs in macrophages. 0 μg/ml as control group and the data were analyzed using t-test to determine significance of difference. a: significant difference when the Lceo-AgNPs-treated group were compared to control group; b: significant difference when the AgNPs-treated group were compared to the control group.

FIGURE 10

Photographs of skin wounds at 4, 7, and 14 days after treatment.

FIGURE 11

Wound-healing rate at different timepoints including days 1, 4, 7, and 14 after different treatments (*p < 0.05).

FIGURE 12

H&E staining of healthy and wound skin sections after 4 and 14 days with different treatments.