Centella Asıatıca Bitki Özütü ile Kaplı Gümüş Nanopartikül İçeren Polimer Liflerin Yara İyileştirici Etkisinin in Vitro ve in Vivo Araştırılması

Abstract

In this study, we synthesized functional silver nanoparticles (AgNPs) from silver ions using the therapeutic plant extract of Centella asiatica (CA). The formation of AgNPs coated with CA functional groups (CA-AgNPs) is evidenced by characteristic surface plasmon resonance (SPR) peaks at a wavelength of about 420 nm. The mean zeta size and zeta potential of CA-AgNPs were found to be 29.5 nm and -24.5 mV, respectively. It was determined that the obtained nanoparticles had a negative surface charge and the results showed that the CA-AgNPs were stable. Functional group analysis of CA-AgNPs was investigated by fourier transform infrared spectroscopy (FTIR) and it was determined that carboxylate and hydroxyl groups play a role in the reduction of silver ions to AgNPs. In addition, CA-AgNPs have been shown nano-sized and spherical by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) examinations. The thermal properties of the CA extract and CA-AgNPs were investigated by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC), and it was physically proven for the first time in the literature that CA-AgNPs contain nano-sized metallic silver. It has been observed that the synthesized CA-AgNPs have antibacterial activity on both gram positive Staphylococcus aureus and gram negative Escherichia coli. The cytotoxicity effects of CA and CA-AgNPs in different concentrations on L929 fibroblast were investigated with the MTT test, and it was determined that CA-AgNPs had a cytotoxic effect at concentrations above 1 mM. In the production of hybrid nanofiber dressing by electrospinning method; before this, solutions of PCL and PEO with different concentrations were prepared with suitable solvents. Chloform/methanol (7/3, v/v) mixture was used for PCL and pure water was used as the solvent system for PEO. Bead-free, smooth, long and uniform fibers were obtained with 3.5% PEO and 12% PCL solutions under optimum electrospinning conditions, and mean fiber diameters of PEO and PCL were determined as 177±21 nm and 412±93 nm, respectively. Before the synthesis of hybrid nanofiber blanket by electrospinning method; Solutions of PCL and PEO in different concentrations were prepared with suitable solvents. Chloroform/methanol (7/3, v/v) mixture was used for PCL and pure water was used as the solvent system for PEO. Bead-free, smooth, long and uniform fibers were obtained with 3.5% PEO and 12% PCL solutions under optimum electrospinning conditions, and mean fiber diameters of PEO and PCL were determined as 177±21 nm and 412±93 nm, respectively. PEO (3.5%) solutions containing 1%, 5% and 10% (v/v) CA-AgNPs were prepared from 1mM CA-AgNPs solution adjusted to pH 7.4 and bead-free and continuous nanofibers (90 nm-160 nm) was synthesized. Hybrid nanofibers obtained by mutual spinning of the prepared CA-AgNPs-PEO and PCL solutions at optimum electrospinning parameters were characterized by FE-SEM, SEM-EDS, FT-IR, XRD, XRF, TGA, DSC, BET methods. In addition, the mechanical properties, air permeability, water uptake capacity, water contact angle, in vitro degradation and in vitro silver release behavior of the synthesized nanofibers were investigated. With antimicrobial studies, PEO/PCL nanofibers containing 5% and 10% CA-AgNPs were found to be effective against S. aureus, E. coli and C. albicans species. The cytotoxic properties of nanofibers were investigated by MTT test and cell viability was determined to be above 70% at all concentrations. Cell adhesion, apoptotic-necrotic, hemolytic and genotoxic effects of 10% CA-AgNPs-PEO/PCL nanofibers, which are preferred as wound dressings, were investigated by in vitro studies. In studies by evaluating the effects of 10% CA-AgNPs-PEO/PCL nanofiber dressing on skin irritation, skin sensitization and healing of burn wounds by in vivo experiments; healthy young adult albino rabbits were used for skin irritation, healthy young adult albino Guinea pigs for skin sensitization, and Sprague-Dawley rats for burn wound. In addition, histological evaluations were made on tissue samples taken from the wound sites of rats, thus the effect of the nanofiber dressing on the epithelialization of the surface on which it was applied was also investigated.