Elsevier

Applied Surface Science

Volume 507, 30 March 2020, 145032
Applied Surface Science

Full Length Article
Copper nanoparticles obtained by laser ablation in liquids as bactericidal agent for dental applications

https://doi.org/10.1016/j.apsusc.2019.145032Get rights and content

Highlights

  • Copper nanoparticles were obtained by laser ablation without any additional reagent.

  • The obtained NPs showed cytocompatibility and inhibition of A. actinomycetemcomitans.

  • Oxidation state and size are key parameters in the bactericidal process.

  • Ions release is not the main responsible for the bactericidal activity of Cu NPs.

Abstract

The dramatic increase of antibiotic-resistant bacteria is considered one of the greatest threats to human health at global scale. The antibacterial activity of noble metal nanoparticles, could be the solution against bacterial infectious diseases which currently do not respond to conventional treatments.

In this work, copper nanoparticles were produced by laser ablation using two different lasers. A nanosecond laser operating at 532 nm and a picosecond laser at 1064 nm were used to ablate a copper target submerged in water and methyl alcohol. The obtained colloidal solutions consisted of copper oxide nanoparticles in suspension with diameters ranging from few nanometers to 45 nm. The nanoparticles formation process is highly influenced by laser parameters, but the solvent plays a crucial role on their characteristics. Cu oxide nanoparticles obtained in water present chain-like nanostructure, while those obtained in methyl alcohol are spherical with lower presence of oxide. All the obtained nanoparticles are crystalline and noticeably stable.

Microbiology tests confirm their strong activity against Aggregatibacter actinomycetemcomitans. Cytocompatibility with human periodontal ligament stem cells is also confirmed. The biological assays evidence that ions release is not the main parameter responsible for the bactericidal activity of copper nanoparticles. Other factors such as oxidation state, size and crystallographic structure, have a greater influence on the process.

Introduction

Nowadays, more and more infections caused by resistant microorganism, fail to conventional treatments. According to the Centre for Disease Control and Prevention (CDC), antibiotic resistant bacteria cause at least 2 million infections per year with 23,000 deaths in the U.S. and 25,000 deaths in Europe [1]. The excess and improper use of antibiotics, has made the treatment of infections more difficult and expensive; some voices in the health system claim we are facing a new global health crisis [2], [3]. In this sense, Gram-negative bacteria are a huge threat, because of the rapid evolution of their resistance mechanisms what results in insusceptibility to nearly all available antibiotics [4].

In dentistry, some of the more extended diseases caused by bacterial infections are periodontitis and periimplantitis. In both cases, the inflammation and destruction of soft and hard tissues surrounding teeth or dental implants, ultimately leads to loss of teeth or dental implant failure as the most common consequences [5]. Although many risk factors are related to the origin of periimplantitis [6], pathogenic microflora are the main cause of this periodontal disease [5]. Aggregatibacter actinomycetemcomitans is a Gram-negative bacteria and commonly part of the normal flora of human mouths, especially in gingival and supragingival crevices [7].

In light of this situation, it is necessary to explore new alternatives in the treatment of diseases caused by infections such as periimplantitis. Nobel metal nanoparticles have become an attractive alternative source to fight against such resistant microorganisms. The antibacterial activity of noble metal nanoparticles have been extensively studied because of their high surface to volume ratio, that increases reactivity allowing to kill the pathogens efficiently [8]. Particularly, copper nanoparticles are of special interest because they are potentially effective against different bacterial pathogens [8], [9], and are very attractive in terms of cost when compared for example with Ag nanoparticles [10].

Although a great number of studies in the recent years have tried to explain the mechanisms of the bactericidal process, this still represents a gap in our knowledge. Previous researches have addressed that not only the size is behind this remarkable antibacterial activity, also shape and crystallographic structure of the nanoparticles [11], which depend on the fabrication method. In this sense, Laser ablation of solids in liquids (LASL) is a key technique to obtain pure nanoparticles with no need of chemical precursors which can contaminate the obtained material, resulting in a potential harmful agent not only for the target bacteria but also for the healthy tissues.

In previous works, other noble metal nanoparticles such as Ag NPs were obtained by means of laser ablation in open air [12] and water [13], [14] and their bactericidal effects were demonstrated. Copper and copper oxide nanoparticles were already synthesized by laser ablation in different solvents such as distilled de-ionized water [15], acetone [16] or organic solutions such as phenanthroline [17]. These works show that the characteristic features of the obtained nanoparticles can be controlled by varying the laser parameters and the liquid properties. Although all of them demonstrate the bactericidal properties of copper nanoparticles, none of them establishes the relationship between bactericidal activity and physicochemical properties, neither their biocompatibility.

In the present work, the bactericidal properties of the Cu nanoparticles are evaluated against A. actinomycetemcomitans (a Gram negative bacteria) one of the main pathogens responsible for inducing localized aggressive periodontitis [18], peri-implantitis [5] and various non-oral infections [7]. In order to assess their biocompatibility, the cytotoxic effects were evaluated using human periodontal ligament stem cells. The mechanism responsible for the bacterial growth inhibition is also studied and discussed. The present study provides some insights as to the influence of nanoparticle size, morphology, oxidation state, crystallographic structure, stability and ion release on the biocidal process.

Section snippets

Laser ablation

Copper foils with 99.99% of purity (Thermo Fisher Scientific), were used as laser ablation targets. In order to analyze the influence of the laser parameters and the liquid medium used in the process, the targets were submerged in two different solvents and ablated by two different diode-pumped Nd:YVO4 laser sources.

Sample nomenclature with the corresponding assay conditions are listed in Table 1.

The first laser source was a nanosecond laser providing pulses at wavelength of 532 nm (Green –

Results and discussion

Cu-NPs in suspension were produced by laser ablation in two different solvents without using any chemical precursors. Nanoparticle formation was directly detected because of the change in the solvent color, from colorless to the final coloring (see Fig. 2). The colloidal solutions obtained in water (a and c) exhibit greenish color indicating oxidized Cu. The nanoparticles obtained in methanol show lighter color due different level of oxidation and particle size.

Despite the different color of

Conclusions

Feasibility of laser ablation of solids in liquids to produce crystalline copper nanoparticles without any additional chemical compound is demonstrated. The type of solvent has more influence over size and stability of the obtained nanoparticles than the laser source used and determines to a large extent the characteristics of the nanoparticles.

Cu-NPs obtained by laser ablation in methyl alcohol are spherical with low degree of oxidation, while those obtained in water present chain-like

Author Contributions

M. Fernández-Arias: Collected the data, Performed the analysis, Wrote the paper. M. Boutinguiza: Conceived and designed the analysis, Performed the analysis, Wrote the paper. J. Del Val: Contributed data or analysis tools, Other contribution: Graphic design. C. Covarrubias: Conceived and designed the analysis, Collected the data. F. Bastias: Contributed data or analysis tools. L. Gómez: Performed the analysis. M. Maureira: Collected the data. F. Arias-González: Contributed data or analysis

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was partially supported by the EU research project Bluehuman (EAPA_151/2016 Interreg Atlantic Area), Government of Spain [RTI2018-095490-J-I00 (MCIU/AEI/FEDER, UE)], and by Xunta de Galicia (ED431B 2016/042, ED481D 2017/010, ED481B 2016/047-0). The technical staff from CACTI (University of Vigo) is gratefully acknowledged. C. Covarrubias acknowledges support from Project U-Redes NanoBioMat, University of Chile.

References (42)

  • J. Zhu et al.

    Highly dispersed CuO nanoparticles prepared by a novel quick-precipitation method

    Mater. Lett.

    (2004)
  • S. Prakash et al.

    Green synthesis of copper oxide nanoparticles and its effective applications in Biginelli reaction, BTB photodegradation and antibacterial activity

    Adv. Powder Technol.

    (2018)
  • G. Cristoforetti et al.

    Physico-chemical properties of Pd nanoparticles produced by Pulsed Laser Ablation in different organic solvents

    Appl. Surf. Sci.

    (2012)
  • S. Bhattacharjee

    DLS and zeta potential – what they are and what they are not?

    J. Control. Release

    (2016)
  • N. Wang et al.

    Influence of metal oxide nanoparticles concentration on their zeta potential

    J. Colloid Interface Sci.

    (2013)
  • B. Li et al.

    Bacteria antibiotic resistance: new challenges and opportunities for implant-associated orthopaedic infections

    J. Orthop. Res.

    (2018)
  • C.L. Ventola

    The antibiotic resistance crisis Part 1: causes and threats

    P&T

    (2015)
  • A. Raghunath et al.

    Metal oxide nanoparticles as antimicrobial agents: a promise for the future

    Int. J. Antimicrob. Agents.

    (2017)
  • L.J.A. Heitz-Mayfield et al.

    Comparative biology of chronic and aggressive periodontitis vs. peri-implantitis

    Periodontol

    (2000)
  • J. Mouhyi et al.

    The Peri-Implantitis : Implant Surfaces, Microstructure, and Physicochemical Aspects

    Clin. Implant Dent. Relat. Res.

    (2009)
  • L.G. Rubin

    181 – Other Gram-Negative Coccobacilli

    (2018)
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