Graphene Oxide–Silver Nanoparticle Nanocomposites: A Potent Antibacterial and Antifungal Agent

Abstract

Surface‑modified materials featuring biocidal nanoparticles offer a promising strategy against multidrug‑resistant pathogens. We report a graphene oxide (GO) nanocomposite decorated with silver nanoparticles (Ag‑NPs) that exhibits superior antibacterial and antifungal activity compared with either component alone. Using ultrasonic coating, polyurethane foils were functionalised with GO, Ag‑NPs, or a GO–Ag hybrid. Efficacy against Escherichia coli, Staphylococcus aureus, Staphylococcus epidermidis, and Candida albicans was assessed via cell morphology, PrestoBlue viability, lactate dehydrogenase (LDH) leakage, and reactive oxygen species (ROS) production. The GO–Ag composite achieved >88 % inhibition of all strains, markedly outperforming GO or Ag‑NPs alone.

Background

While antibiotics historically curbed bacterial infections, their misuse has accelerated the emergence of multidrug‑resistant organisms such as methicillin‑resistant S. aureus and extended‑spectrum β‑lactamase‑producing E. coli [1–3]. Resistance spreads through horizontal gene transfer, underscoring the urgent need for novel antimicrobials. Nanoparticles (NPs) possess multifaceted mechanisms—membrane disruption, metal‑ion release, ROS generation—that collectively hinder resistance development [4–8]. Among them, silver NPs (Ag‑NPs) have demonstrated activity against >650 microbes, yet their aggregation limits surface area and potency [9–11]. Graphene oxide (GO), an oxidised, water‑soluble carbon allotrope rich in oxygenated functional groups, can serve as a stable scaffold for Ag‑NPs, potentially enhancing dispersion and interaction with microbial membranes [12–14]. The present study evaluates a GO–Ag nanocomposite’s antimicrobial performance and mechanistic basis against representative gram‑positive, gram‑negative, and fungal pathogens.

Methods

Synthesis and Characterisation of Graphene Oxide

Commercial graphite (Acros Organics) was oxidised via a modified Hummers protocol [15]. X‑ray diffraction (XRD), Raman spectroscopy, Fourier‑transform infrared (FT‑IR) analysis, and dynamic light scattering (DLS) provided structural, compositional, and size data. Zeta potential measurements assessed colloidal stability.

Preparation of Nanocomposite‑Coated Foils

Polyurethane foils (15 × 15 × 0.05 mm) were immersed in aqueous suspensions of GO (200 µg/mL), Ag‑NPs (100 µg/mL; HydroSilver1000), or a 1:1 mixture. Ultrasonic coating (20 kHz, 60 % power) for 10 min yielded uniform layers. Surface morphology was examined by SEM, TEM, and atomic force microscopy (AFM). Surface free energy (SFE) was calculated using the Owens–Wendt method with water and diiodomethane probes.

Microbial Cultures and Antimicrobial Assays

Clinical strains E. coli (ATCC 25922), S. aureus (ATCC 25923), S. epidermidis (ATCC 14990), and C. albicans (90028) were grown to mid‑log phase, then applied to foils (10⁶ CFU/mL) on Mueller–Hinton agar. After 24 h at 37 °C, colony counts quantified growth inhibition.

Cell Viability, Membrane Integrity, and ROS Assessment

PrestoBlue™ assays measured metabolic activity; LDH leakage quantified membrane disruption; DCFDA fluorescence detected intracellular ROS. All experiments were performed in triplicate.

Results

GO and Ag‑NP Physicochemical Properties

GO displayed a highly disordered lattice (I_D/I_G = 1.15) and abundant oxygen‑bearing groups (C=O, C–OH, C–O–C). Ag‑NPs averaged 80 nm in suspension, increasing to ~218 nm post‑ultrasonication due to poly(vinyl alcohol) interaction. The GO–Ag mixture showed a negative zeta potential (~–7 mV), indicating moderate colloidal stability.

Surface Morphology and Energy

SEM revealed that GO formed a continuous, flaky coating; Ag‑NPs produced a granular grid; the GO–Ag composite exhibited both features. AFM confirmed GO flakes adhered to the polymer substrate. GO‑coated foils increased polar SFE from 2.3 ± 0.6 to 68.9 ± 2.8 mJ/m², whereas Ag‑NP and GO–Ag coatings did not significantly alter SFE.

Antimicrobial Efficacy

After 24 h exposure, GO–Ag foils inhibited growth by 88.6 % (E. coli), 79.6 % (S. aureus), 76.5 % (S. epidermidis), and 77.5 % (C. albicans). GO or Ag‑NP coatings alone yielded <20 % inhibition. SEM of treated cells showed pronounced membrane deformation and reduced colony size.

Membrane Integrity and ROS Generation

LDH leakage was highest for GO–Ag (66.3 % for E. coli; 59.4 % for S. aureus; 54.8 % for S. epidermidis; 48.5 % for C. albicans), indicating extensive membrane compromise. ROS levels increased markedly for GO–Ag and Ag‑NP foils (p < 0.05), while GO alone modestly elevated ROS in C. albicans.

Discussion

Our findings confirm that decorating GO with Ag‑NPs produces a synergistic antimicrobial effect. The dual mechanisms—physical disruption by GO edges and chemical assault by Ag‑NPs/Ag⁺ ions—result in rapid membrane permeabilisation and oxidative stress, overwhelming microbial defence systems [16–18]. The superior efficacy against gram‑negative bacteria may reflect the thinner peptidoglycan layer, facilitating NP access, whereas gram‑positive strains exhibit partial resistance due to a thicker cell wall. The GO–Ag nanocomposite’s ability to maintain dispersion under ultrasonic treatment addresses the aggregation limitation of bare Ag‑NPs, preserving bioactive surface area.

Conclusions

GO–Ag nanocomposites demonstrate potent, broad‑spectrum antimicrobial activity, outperforming GO or Ag‑NPs alone. Their mechanism involves membrane disruption and ROS‑induced oxidative damage. These materials hold promise for application in medical textiles, wound dressings, and surface coatings where multidrug‑resistant pathogens pose a threat.

Abbreviations

AFM

Atomic force microscopy

Ag‑NPs

Silver nanoparticles

DLS

Dynamic light scattering

GO

Graphene oxide

GO‑Ag

Graphene oxide decorated with silver nanoparticles

LDE

Laser Doppler electrophoresis

LDH

Lactate dehydrogenase

ROS

Reactive oxygen species

SEM

Scanning electron microscopy

XRD

X‑ray diffraction