CONICET | Buscador de Institutos y Recursos Humanos

A NEW ANTIMICROBIAL STRATEGY INVOLVING GOLD NANOPARTICLES AND GREEN LEDs Silvero MJ1, Rocca DM1, Aiassa V2, Artur de la Villarmois E 3, Scaiano JC4, Pérez MF3, Becerra MC1*.1 Instituto Multidisciplinario de Biología Vegetal (IMBIV), CONICET, and Departamento de Farmacia, Facultad de Ciencias Químicas, Universidad Nacional de Córdoba, Córdoba X5000HUA, Argentina.2 Unidad de Tecnología Farmacéutica (UNITEFA), CONICET, and Departamento de Farmacia, FCQ, UNC. 3 Instituto de Farmacología Experimental de Córdoba (IFEC), CONICET and Departamento de Farmacología, FCQ, UNC.4 Universidad de Ottawa, Canadá.*becerra@fcq.unc.edu.ar.Abstract The on growing resistance to antibiotics by microorganisms1 has led to the development of a new antimicrobial strategy. Our work has focused in the development of Photodynamic Antimicrobial Chemotherapy (PACT) involving low cost LEDs with appropriate wavelength to excite the plasmon of gold nanoparticles (NPs). Currently, Photodynamic therapy (PDT) is applied to treat cancer and various infections that are not susceptible to antimicrobial drugs. We have concentrated our studies on the photo-antibacterial capacity of gold and gold/silver bimetallic nanoparticles against antibiotic-resistant bacterial strains. Microbiology essays proved that this kind of NP have a bactericidal effect on various reference and clinical strains. Reactive oxygen species (ROS) detection through fluorescent probes demonstrated that oxidative stress is caused after few hours of PACT. We believe that plasmon excitation of the gold core may produce high ROS levels through a photo-thermal process, up to 12x the non-irradiated inoculum without NP, and 5x the non-irradiated inoculum with NP. Further, macromolecular damage was found in both proteins and lipids, which irreversibly led to bacterial death. Indeed, for the same strains, advanced oxidation protein products were more than double with respect to the control at 4 h, whereas the maxima of lipid peroxidation were sensed through a fluorescent probe at 8 h (30000x to 60000x units more than the controls)2. The time needed to achieve complete inhibition of bacterial growth was shorter for combined gold/silver NP than for gold NP alone. These results suggest that the silver shell was contributing as an antibacterial agent, even though it was not significantly excited by the green LEDs3. By contrast, green light did not affect the normal cell survival, neither the photosensitizer, when the proposed PDT treatment was essayed on human fibroblasts and keratinocytes: in fact, the percentage of respiratory-active cells stayed above 95% even after 18 h exposure to green light. Based on the antibacterial effect of irradiated gold NPs, we have recently synthesised ones with an amoxicillin shell in order to make them more selective for the prokaryotic cells. In this case, the antibiotic was used as the reducing agent and as the stabilizer at the same time, leaving no other chemical in the aqueous media. They are stable in many culture media and, moreover, they keep their shape and size in the blood stream and inside the tissues and organs, as the in vivo experiments in male Wistar rats have demonstrated. They are extremely biocompatible and do not produce any alteration in blood cells. Interesting interactions and minor alterations with liver, kidney, brain, heart and spleen cells were observed through TEM images, while their excretion was followed by measuring the absorption spectrum of urine samples. It is worth to remark that the antibacterial activity of photo-activated metal nanostructures was sustained after 30-40 consecutives bacteria generations, which encourage us to believe that PACT with NPs could circumvent problems related to the development of drug resistance by bacteria.References[1] http://www.who.int/mediacentre/news/releases/2014/amr-report/en/[2] M.J. Silvero, M.C. Becerra, RSC Advances 6 (2016) 100203?100208. [3] C. Fasciani, MJ Silvero, MA Anghel, GA Argüello, MC Becerra, JC Scaiano, J Am Chem Soc. 17 (2014) 136(50):17394-17397. AcknowledgementsThis work was supported by grants of FONCYT, CONICET and SECyT.