key: cord-0902314-bz31bds1 authors: Rehman, Saliha ur; Niazi, Robina Khan; Zulqurnain, M.; Mansoor, Qaisar; Iqbal, Javed; Arshad, Aqsa title: Graphene nanoplatelets/CeO(2) nanotiles nanocomposites as effective antibacterial material for multiple drug-resistant bacteria date: 2022-03-14 journal: Appl Nanosci DOI: 10.1007/s13204-022-02422-9 sha: c45a410e28298a52066e73b7f4466fdebb2746ce doc_id: 902314 cord_uid: bz31bds1 Antibacterial agents with low toxicity to normal cells, redox activity and free radical scavenging property are urgently needed to address the global health crisis. The phenomenal conducting nature of graphene is a best fit to enhance the antibacterial properties of metal oxides. In this work, CeO(2) nanotiles and graphene nanoplatelets/CeO(2) nanotiles nanocomposites (G/CeO(2)) have been synthesized by a solvothermal method. The prepared materials have been characterized using XRD, FE-SEM, EDX, and UV–visible spectroscopy techniques to investigate their crystallinity, morphology, composition, and optical bandgap energies. The CeO(2) and G/CeO(2) nanocomposites have also been tested for antibacterial applications. The neat CeO(2) nanotiles sample inhibits the bacterial growth of Pseudomonas aeruginosa and Staphylococcus aureus up to 14.21% and 39.53% respectively. The antibacterial activity was tremendously enhanced using 25% graphene-loaded sample (G/CeO(2)-II) i.e., approximately 83% loss of P. aeruginosa and 89% in case of S. aureus has been observed. This can be attributed to the unique nano-architecture, oxidative stress due to the excellent ability of reversible conversion between the two electronic states of CeO(2) and the stress exerted by the planar graphene and CeO(2) nanotiles. Therefore, the G/CeO(2) nanocomposites can find potential application as nano-antibiotics for controlling pathogens. Antimicrobial drug resistance (ADR) is one of the greatest health issues faced globally (Majumder et al. 2020) . Antibiotics are the human defence armours that fight against pathogens, but the excessive use of antibiotics and rapid transfer of resistance genes has led to ineffective antibiotics. The alarming rate of worldwide antibiotic resistance has threatened the human health by increasing the mortality rate along with an increased treatment cost (AlSheikh et al. 2020) . UN Ad hoc integrity coordinating group of antimicrobial resistance reported that drug resistance-related diseases cause 10 million deaths each year by 2025. Currently, 700,000 people die every year due to drug-resistant diseases, which includes 230,000 deaths due to multidrug-resistant (MDR) tuberculosis (TB). The severity of microbial co-infection is also highlighted in COVID-19 pandemic. Bacterial pathogens are highly probable to be found in viral respiratory infections like COVID-19. The use of antibiotics is essential in such cases (Langford et al. 2020) . Therefore, the current global pandemic along with possibility of multiple drug resistance in the human body has posed major health challenges. TB, MRSA and VRE infections, diarrhoea, severe sexual complications and some CRE infections are caused by mycobacterium, Vancomycin-resistant Enterococci, Methicillin-resistant Staphylococcus aureus (MRSA), Neisseria gonorrhoea, Clostridium difficile and Carbapenem-resistant Enterobacteriaceae, i.e., Klebsiella species and Escherichia coli respectively. These bacteria have developed resistance to antibacterial drugs over the years. In many parts of the world, the high resistance of E. coli against fluoroquinolone antibiotics has made it ineffective in half of the patients suffering with urinary tract infections. The Staphylococcus aureus has also been resistant to first-line drugs. People with MRSA are 64% more probable to die than people with nonresistant form of bacteria (AL-Saadi 2016; Hibbitts et al. 2019) . Therefore, some immediate steps are needed to prevent a potentially disastrous drug resistance, e.g., it is necessary to develop novel nano-medicines that have tendency to fight against bacterial infection-related morbidity. Metal oxides have been a promising candidate (He et al. 2015) for the development of novel nano-antibiotics. They have other potential applications like photocatalysis, superconductivity, electronics, nanorobotics, and automobile industry Huang and Chen 2009; Kim et al. 2010; Halder and Sun 2019; Meixner et al. 1995; Kummer 1986) . Many transition metal oxides show significant antibacterial performance towards a wide range of Gramnegative and Gram-positive bacteria (Paul et al. 2020; Talat et al. 2020) . These properties are further enhanced if the same materials with reduced dimensions are employed, which mainly is attributed to distinct physicochemical properties appearing at nanoscale (Sanpo et al. 2013; Wang et al. 2010) . Metal oxides have a common feature of producing oxidative stress due to generation of reactive oxygen species (ROS) in presence of UV light. CeO 2 is unique as it can generate ROS even without UV light exposure (Jones et al. 2008) . Moreover, the oxidative stress generation is a unique property of nano-CeO 2 because it has an excellent ability of reversible conversion between its two electronic states in an auto-regenerative cycle (Ce 4+ → Ce 3+ → Ce 4+ ) (Stoianov et al. 2014) . CeO 2 being a natural scavenger of ROS and nitrogen species, oxidizes from Ce (III) to Ce (IV) in the presence of ROS and then undergoes back to Ce (III) through a regeneration process. Therefore, CeO 2 NPs can reduce oxidative stress by acting as antioxidants and may also be toxic to cancerous and bacterial cells by generating ROS under some conditions (Zhang et al. 2019) . The ROS generating capability of CeO 2 contributes towards inhibition of the bacterial growth, which is due to the oxidative stress on lipids and proteins (that are found in plasma membrane of microorganisms) caused by the reduction of Ce (IV) into Ce (III) (Gliga et al. 2017) . Therefore, the antibacterial properties of CeO 2 based materials are expected to be extraordinary as compared to other metal oxides, based on its naturally free radical scavenging property. Also, metal oxide nanostructures with higher surface area than their bulk counterparts lead to higher chemical and biological activity. With these characteristics, metal oxide nanostructures can efficiently target cell membranes for better inactivation of bacteria (Subbiahdoss et al. 2012 ). These properties are also expected to enable nano-CeO 2 to uniquely take up antibiotic drug resistance. However, it is always needed to add highly efficient antibiotic materials to the existing ones to combat ever increasing drug resistance (Bellio et al. 1860) . Therefore, new strategies are required to improve antibacterial efficiency of CeO 2 . Some methods include its modification using dopants and (or) by compositing it with other chemical species. An effective strategy can be the formation of its composites with graphene family nano-materials. Graphene and CeO 2 nano-composites have found various applications, e.g., as biosensors (Yang et al. 2016) , cholesterol detectors (Zhang et al. 2013) , catalysts (Yang et al. 2017) , super-capacitive electrodes (Xie et al. 2018) , anodes in lithium ion battery (Wang et al. 2011 ) and microwave absorbers (Wu et al. 2020) . Graphene-based metal oxide nano-composites have emerged as an extremely promising class of antibacterial materials due to their distinct electrical properties and low toxicity towards mammalian cells (Joung et al. 2011; Rojas-Andrade et al. 2017; Mathew et al. 2020; Alhazmi et al. 2022) . When compared to published reports on biomedical applications of CeO 2 , very limited information is available on the biomedical applications of graphene-based CeO 2 nanocomposites. Some related materials have been studied, e.g., antimicrobial efficiency of CeO 2 /GO nano-composites has been tested (Kashinath et al. 2019; Bharathi and Stalin 2019) . However, the anti-bacterial activity of CeO 2 and neat graphene nanocomposites has not been studied so far. Graphene-based composites are phenomenally effective in bacterial killing due to their bio-compatibility, sharp edgesbased sheets like morphology and high specific surface area (Ni et al. 2010) . Therefore, it can be anticipated that the antibacterial potential of CeO 2 may possibly be enhanced by the formation of a unique nano-architecture of G/CeO 2 nanocomposites. Keeping in view the mentioned discussion, we aim to boost the antibacterial performance by synthesizing cutterlike morphology of CeO 2 nanostructures so that it can act synergistically with the sharp edges of graphene to achieve an outstanding bactericidal or bacteriostatic behaviour against both Gram-negative and Gram-positive bacteria. Therefore, in this work, we present synthesis of CeO 2 nanotiles and G/CeO 2 nanocomposites and study their structural, morphological, optical, and antibacterial properties against two model pathogenic bacterial strains S. aureus and P. aeruginosa. S. aureus is found in skin and respiratory tract, causes a wide range of clinical infections i.e., pneumonia, pimples etc. It is a Gram-positive bacterium (Tong et al. 2015) . P. aeruginosa causes pneumonia, septic shock and many blood infections in humans (Todar 2006) . To the best of our knowledge, this is the first report on the synthesis of CeO 2 nanotiles and the investigation of antibacterial behaviour of G/CeO 2 nanocomposites. The materials used for making CeO 2 and G/CeO 2 nanocomposites are: cerium nitrate hexahydrate Ce(NO 3 ) 3 .6H 2 O, 5 M aqueous sodium hydroxide solution (NaOH), isopropyl alcohol (99%, Sigma Aldrich), ethyl alcohol (99%, Merck), citric acid (C 6 H 8 O 7 ), distilled water and graphene nanoplatelets (100%, KNANO) with the thickness ≤ 15 nm and size ca. 15 μm. All the chemicals were used as obtained. A simple, surfactant-free and cost-effective solvothermal method was used to fabricate CeO 2 and G/CeO 2 . The reaction took place in the beakers first and then it was shifted to an autoclave. Initially 8.642 g Ce (NO 3 ) 3 .6H 2 O and 11.52 g C 6 H 8 O 7 were dispersed in 100 ml distilled water separately in two beakers. These two solutions were mixed. This solution with pH 1 was stirred magnetically. A 5 M aqueous NaOH solution was then added drop-wise to the precursor solution to adjust its pH to 2. This solution was then shifted to an autoclave, which was heat-treated at 90 ℃ for 24 h. The solid was retained after washing it for three times in a centrifuge machine using distilled water and ethanol. The precipitates so collected were dried in oven at 120 ℃ overnight and were ground. The powder was heat-treated in a box furnace at 400 ℃ for 3 h to get a phase pure form. Two different samples of nanocomposites were synthesized using different quantities of graphene, i.e., 12% and 25% using a similar method depicted above, except the addition of GNPs. GNPs were dispersed ultrasonically in a solution containing distilled water and isopropyl alcohol (equal volume). This solution was then added to the CeO 2 solution followed by similar procedure as depicted above. The process is presented schematically in Fig. 1 . The effect of graphene-based CeO 2 nanocomposites on P. aeruginosa and S. aureus can be studied by making disc diffusion assays. Disc diffusion method or Kirby-Bauer test method is used to check the effect of antibiotics on bacteria. We have followed the same protocol as described in previous work (Luc 2015; Arshad et al. 2017a) . Briefly, in this testing method, bacteria are kept in agar plates and antibiotic wafers are placed on them. 10 mg/L of each test sample (CeO 2 , G/ CeO 2 -I and G/CeO 2 -II) are sonicated in sterile water. 5 mL of LB medium (Luria-Bertani Broth) is mixed with 200 μL of each sonicated solution which is then inoculated into 100 μL of the bacterial culture in LB and incubated at 37 °C for 24 h. The structural and phase analysis was performed using powder X-ray diffraction technique at PANalytical X'Pert PRO diffractometer. The surface, substructure and shape of prepared nanomaterials were obtained using SEM (MIRA3-TESCAN) that has a coupled electron dispersive X-ray (EDX) spectrometer. The absorbance profiles were studied using UV-1700 UV-visible spectrophotometer (SHI-MADZU PharmaSpec). The Raman spectra were obtained using Raman spectrophotometer (Nost). The phase purity of the samples is examined by X-ray diffraction. The XRD results of CeO 2 , G/CeO 2 -I and G/CeO 2 -II are shown in Fig. 2 . The XRD peaks of prepared samples have been obtained from 20° to 80°. Results show that CeO 2 nanotiles can be indexed to the pure fluorite cubic structure with a lattice constant a = 5.41 Å. These results match well with JCPDS card no. 00-004-0593 and literature (Kannan and Sundrarajan 2014; Hirano and Kato 1999) . Diffraction peaks at 28.55°, 33.07°, 47.49°, 56.328°, 76.73° and 79.079° can be assigned to (111), (200), (220), (311), (331) and (420) planes of CeO 2 respectively. No impurity peaks are observed in the pure CeO 2 sample. In G/CeO 2 -I and G/ CeO 2 -II, presence of (002) carbon peak at 2θ = 26.228° proves the composition of graphene with CeO 2 . This confirms the formation of biphasic G/CeO 2 nanocomposites. We have also determined the interlayer spacing of neat GNPs and G/CeO 2 nanocomposites. The interlayer spacing for neat GNPs is 3.35 Å, whereas it has increased for G/CeO 2 nanocomposites to 3.37 Å. This increase in interlayer spacing may be attributed to the presence of CeO 2 along with multiple graphene sheets. The average crystallite size is calculated using Scherrer's equation, D = k cos . The crystallite size of neat CeO 2 sample is estimated to be 14.14 nm which is reduced to 13.57 nm and 2.16 nm for G/CeO 2 -I and G/ CeO 2 -II, as determined using (111) peak only. The morphology of prepared samples has been studied by scanning electron microscope (SEM). Figure 3a-c shows the morphology details of all the three samples. Figure 3a reveals that the CeO 2 sample has nano-tiles like structure with the thickness ranging from 76 to 113 nm. Figure 3b , c depict SEM micrographs of G/CeO 2 -I and G/CeO 2 -II respectively. Graphene nanopatches plastered on CeO 2 nanotiles can be seen clearly, that confirm the formation of bi-phase G/CeO 2 nanocomposites. The TEM images of G/ CeO 2 -II nanocomposite are presented in Fig. 3d-f . These images depict the transparent sheet-like structures are graphene sheets as observed in Fig. 3d , e. The images also show the random presence of CeO 2 nanotiles on the graphene sheets. The semi-transparent to transparent graphene nanosheets are clearly visible in Fig. 3e . Figure 3f shows that the thickness of one of the CeO 2 nanotiles is 144.4 nm. The TEM images provide further insight into the morphology of the prepared samples and confirm the formation of G/CeO 2 nanocomposite. The energy-dispersive X-ray spectroscopy has been used to investigate the elemental composition of the synthesized samples. Figure 4a shows the elemental composition of CeO 2 nanotiles. The elemental presence of Ce and O ensures the formation of CeO 2, and no impurity elements are observed. Carbon peaks can be observed in G/CeO 2 -I and G/CeO 2 -II nanocomposite samples, in addition to Ce and O peaks, shown in Fig. 4b , c which are due to presence of graphene. However, making the sample's surface conductive is important, which has been achieved by carbon coating on the sample's surface. Therefore, a low intensity peak related to carbon has also been observed in neat CeO 2 , which is due to carbon coating. No impurity peaks have been observed. The absorbance of prepared samples is investigated using UV-vis spectrophotometer. Figure 5 shows the absorbance spectra of CeO 2 and its composites with graphene. Since, all the samples show a definite absorption peak, therefore, in this case, it is preferred to find the bandgap energies using the relation, E g = 1240 eV . Here λ is the wavelength associated with the prominent peak in respective absorbance profiles of all the three samples. The bandgap energies so obtained are 4.14 eV (for CeO 2 ), 3.86 eV (for G/CeO 2 -I) and 3.80 eV (for G/CeO 2 -II). The observed bandgap of CeO 2 is slightly higher than the reported values for bulk CeO 2 , but it agrees well with a study where 4.65 eV bandgap has been reported for CeO 2 nanoparticles (Safat et al. 2021) . The quantum size effects that occur in materials with nanoscale dimension lead to an increase in bandgap energies, observed in this case (Srdanov et al. 1994 ). It has been observed that the bandgap energies show a decreasing trend in their value due to the inclusion of graphene, in G/CeO 2 -I and G/CeO 2 -II nanocomposites, respectively. The decreasing trend of energy bandgap values is attributed to new energy levels. The existence of new energy levels between valence and conduction bands of CeO 2 in the nanocomposite systems are due to the modification in the electronic band structure in the composite material. Electronic interaction between CeO 2 and graphene sheets may result in the introduction of energy states associated with defects, and modification of band structure due to inclusion of graphene in the vicinity of CeO 2 (Arshad et al. 2017b ). The study of phonon modes of G/CeO 2 nanocomposite using Raman spectroscopy is presented in Fig. 6 . The symmetric stretching mode of Ce-O vibrations (Fluorite, F 2g ) is observed at 461.39 cm −1 . The three primary features of the graphene Raman spectra are D, D' and G band. The G peak in G/CeO 2 -II nanocomposite is associated with E 2g vibrational mode of ordered in-plane sp 2 carbons and is a characteristic for all sp 2 -hybridized carbon structures. The structural defects and impurities express themselves as D band. GNPs and CeO 2 nanotiles in the nanocomposite give rise to D band with enhanced intensity, as compared to that of neat GNPs. This also confirms the formation of nanocomposite and the surface modification of neat GNPs in the G/CeO 2 nanocomposite. The increased width of D band also quantifies the induced disorder in GNPs which is due to CeO 2 presence in the nanocomposite. The D and D' peaks exhibit themselves at 1342 cm −1 and 2695 cm −1 , respectively in G/ CeO 2 -II nanocomposite. The time kill assay is used to study the effect of CeO 2 and G/CeO 2 nanocomposites on Gram-negative and Grampositive bacterial strains. The growth inhibition curves are obtained by recording optical density at 600 nm, at different intervals of time during incubation period 24 h. The control sample presents untreated bacterial strains. The bacterial growth inhibition curves of S. aureus and P. aeruginosa are shown in Fig. 7a , c, whereas Fig. 7b , d depicts the respective cell viabilities. It can be observed from antibacterial experiment that the bacterial growth is significantly inhibited in the presence of prepared materials. For P. aeruginosa, the growth inhibition rates are 14.21%, 61.58% and 82.67% for CeO 2 , G/CeO 2 -I nanocomposite and G/CeO 2 -II nanocomposite, respectively. The growth of S. aureus has been controlled up to 39.53% by CeO 2 , whereas G/CeO 2 -I and G/CeO 2 -II have inhibited the bacterial growth up to 62.4% and 89.48% respectively. It can be concluded from experimental data that neat CeO 2 nanotiles have decreased the cell growth to some extent, but the inclusion of GNPs in the nanocomposites further decreases the number of viable bacterial cells. The nanocomposite with a maximum GNPs content has achieved maximum bacterial growth inhibition rate. It is also evident that the prepared materials show selective cytotoxicity towards the Gram-negative and Gram-positive bacteria. The exact mechanism for the antibacterial activity of metal oxides and graphene-based systems is still controversial; however, several hypothetical mechanisms have been proposed to account for the control of bacterial growth (Ji et al. 2016; Hegab et al. 2016) . The cell composition of bacteria plays a substantial role to determine the toxicity level of prepared materials. Gram-positive bacteria have a single membrane encased by a thick peptidoglycan layer with attached proteins and different glycol polymers like polysaccharides, whereas in Gram-negative bacterial cell, the cytoplasmic membrane is covered by a thin peptidoglycan layer which is over-layered by an asymmetrical phospholipids bilayer having some polysaccharides and proteins (Shahriari et al. 2018) . This difference accounts for the difference in functioning of active nanomedicine on P. aeruginosa and S. aureus as observed in the results depicted above. S. aureus being Gram-positive is easily targeted by G/CeO 2 nanocomposite as compared to P. aeruginosa which is Gram-negative (Arshad et al. 2017a) . The selective cytotoxicity is due to different outer layers of Gram-negative and Gram-positive bacteria, as discussed above, and observed in this study. The interfusion of planar graphene on bacteria can be one of factors that is responsible for inducing the irreversible annihilation of bacterial membrane. It can be speculated form the morphology (Fig. 3) that the rough sharp edges of planar graphene exert stress on bacterial cell membrane which can cause damage to bacterial cell membrane and leakage of cytoplasmic content (Arshad et al. 2019 ). This disruption of membrane is not only limited to the graphene, but the unique morphology of CeO 2 , prepared in this work, also contributes towards bacterial membrane rupture. CeO 2 nanotiles enter the bacterial cell by rupturing its membrane. The sharp blade like sides of CeO 2 nanotiles make it easy to break the cell membranes, thus directly allowing the nanomedicine to interact with the bacterial cytoplasm. This dissolution of bacterial membrane proves to be fatal for bacteria. Also, when bacteria are exposed to CeO 2 nanotiles, the electrostatic charge attraction between negatively charged bacterial cell membrane and positively charged CeO 2 nanotiles causes its adhesion to the bacteria (Kuang et al. 2011 ). Moreover, graphene being an excellent electron acceptor creates a charge imbalance on the cell membrane of bacteria (Harris et al. 2010 ). The bacterial cell membrane and cell wall are typically negatively charged. The contact between graphene and bacterial membrane causes charge flow from membrane to the graphene conducting sheets. This charge imbalance leads to an interruption in bacterial respiratory activity that can cause bacterial cell death. The charge variation at bacterial surface may induce ROS production. The study of graphene toxicity reveals that graphene-based materials mediate oxidative stress. When cellular components are in direct contact with graphene sheets, superoxide anions are generated due to conducting graphene bridge. A related case is depicted for metallic SWCNTs, where the electron transfer from bacterial intracellular components to external environment takes place through metallic SWCNTs (Vecitis et al. 2010) . Graphene can also produce similar effects by modifying bacterial DNA, lipids, and proteins by following the Fig. 7 a Growth profile of P. aeruginosa in the presence of CeO 2 and G/CeO 2 composites, b cell viability of P. aeruginosa, c growth profile of S. aureus in the presence of CeO 2 and G/ CeO 2 composites and d cell viability of S. aureus similar analogy. Strong oxidation by RGO is also observed in a study (Liu et al. 2011) which also supports that rGO nanosheets can oxidize thiol groups of proteins and other bacterial cellular components. The unique composition of prepared nanocomposites is also an important factor that determines the bacterial death. CeO 2 has a very important feature related to Ce 3+ to Ce 4+ ratio. CeO 2 nanotiles when enter inside bacterial cell may allow redox conversion of Ce ions. Ce 4+ atoms may undergo a reduction process from Ce 4+ to Ce 3+ and vice versa in the cell environment. Generally, Ce 3+ to Ce 4+ ratio is maintained by CeO 2 (IV) + e − ⟶ CeO 2 (III). The ratio of Ce 3+ also affects the bandgap values which ultimately influence the antioxidant ability of CeO 2 . This transformation to Ce 3+ is the key parameter to control the ROS generation for effective antibacterial activity (Ansari et al. 2014) . Generally, the metal oxide nanostructures (CeO 2 nanotiles, in this case) can affect the cell viability into two ways either by influencing nutrient transport or by inducing oxidative stress. NPs have a high tendency to interact with biomolecules at sub-cellular level. There are proteins present on the cell membrane that extrude from bacterial membrane to perform nutrients transportation. Ce 4+ nanotiles bind with thiol group (-SH) of these proteins resulting in enlarged proteins and decreased membrane permeability. This leads to the bacterial cell death due to lacking nutrients (Zhang et al. 2019) . The Ce 4+ nanotiles interacting with periplasm and DNA may cause difficulty in cell respiration and interferes with DNA replication and cell division, which in turn hinders the normal function of bacteria. The other factor, i.e., oxidative stress is mainly being developed by ROS generation. ROS are generated as by product of normal metabolic reaction and mitochondrion is the leading organelle responsible for its production. While generating ATP, various ROS like free radicals (i.e., OH − , ·OH, 1 O 2 ), superoxide ions (i.e., O 2 − ) and nonradical molecules (i.e., H 2 O 2 ) are formed. Also, the inter-conversion of Ce 3+ and Ce 4+ amplifies ROS generation. ROS-induced bacterial cell death is composed of oxidative stress and membrane lipid peroxidation. These highly reactive oxygen species can harm the individual bacterial components, such as proteins, mitochondria and nucleic acids, which leads to the interruption of bacterial cell cycle and ultimately induces cell death (Stankic et al. 2016) . A detailed version of reactions taking place between G/CeO 2 and bacteria is demonstrated below (Korsvik et al. 2007; Banavar et al. 2021) : Another factor that may govern the antibacterial activity is the optical bandgap energy of CeO 2 . The suitable light with energy enough to generate the charge carriers (i.e., electrons and holes) when falls on the material, helps the electrons and holes to be available for the generation of ROS. The ROS are essential for effective antibacterial activity. The observed bandgap for CeO 2 , and G/CeO 2 nanocomposites falls in the UV range. Therefore, only a small fraction of light photons may contribute towards the generation of reactive oxygen species, in this case. From the above discussion, we conclude that both graphene and the CeO 2 nanotiles, synergistically restrict the bacterial proliferation. A hypothetical mechanism based on the toxicity of planar graphene sheets and CeO 2 nanotiles against bacterial pathogens is shown in the Fig. 8 . All the possible factors that may influence the bacterial cell death in the presence of G/CeO 2 are presented schematically below. Various metal nanoparticles, metal oxide nanoparticles and their graphene-based composites have been employed to control bacterial growth. Biosynthesized CeO 2 NPs and their nanocomposites have been previously demonstrated as good antibacterial agents. Also, some graphene family materials have also shown significant antibacterial properties. These results are tabulated in Table 1 . CeO 2 and GO/ CeO 2 have been analysed as an antibacterial material previously (Bharathi and Stalin 2019; Kannan and Sundrarajan 2014; Arunachalam et al. 2018; Surendra and Roopan 2016; Hassan et al. 2012; Magdalane et al. 2017; Ravishankar et al. 2015; Yadav et al. 2017; Arumugam et al. 2015; Babu et al. 2014; Khadar et al. 2019; Wang et al. 2013; Moongraksathum and Chen 2018) , but this work, for the first time evaluates the antibacterial activity of G/CeO 2 nanocomposites. The results are promising. Almost 83% growth inhibition of P. aeruginosa and 89% growth inhibition of S. aureus by the G/CeO 2 has been achieved in this work. The unique morphology of CeO 2 nanotiles has been one of the helpful factors towards achieving an excellent antibacterial performance as demonstrated in our work. The work suggests G/CeO 2 as an effective antibacterial material. However, the performance of this material can be further investigated by studying a dose-dependent response of the prepared material. This study only addresses the use of 10 mg/L of prepared samples, which has achieved 83% and 89% growth inhibition. The work can be extended to get the response of prepared materials towards growth inhibition of other bacterial strains (not included in this work). G/CeO 2 nanocomposites are successfully synthesized using a hydrothermal process. The XRD results reveal that CeO 2 exists as fluorite crystal structure in G/CeO 2 nanocomposite, thus confirming the bi-phase nature of the prepared nanocomposites. The morphology analysis obtained using FE-SEM shows that CeO 2 exists as nanotiles, while semitransparent sheets like graphene are also clearly observed in the nanocomposites. The EDX results confirm the purity and elemental composition of the samples. The optical properties as obtained from UV-vis spectroscopy indicate that the inclusion of graphene into neat CeO 2 decreases the bandgap up to 3.80 eV in G/CeO 2 -II nanocomposite. The G/CeO 2 -II nanocomposite shows excellent antibacterial activity, i.e. up to 83% loss of P. aeruginosa and approximately 89% in case of S. aureus. The antibacterial performances of all the prepared nanomaterials reveal that G/CeO 2 -II can be considered as promising antibacterial material. It is noteworthy that the antibacterial activity of these nanomaterials may be attributed to the reversible conversion of Ce (III) and Ce (IV). The unique cutter-like morphology of CeO 2 and graphene inclusion synergistically enhances the activity. These findings can be a landmark to combat ADR and developing PPE (personal protective equipment) in current global health crisis. Biosynthesis, characterization and antimicrobial activity of copper oxide nanoparticles (CONPs) produced using brown alga extract (Bifurcaria bifurcata) Graphene-based biosensors for disease theranostics: development, applications, and recent advancements Estimation of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of cell-free extracts of bifidobacterium species against methicillinresistant Staphylococcus aureus in vitro Plant-based phytochemicals as possible alternative to antibiotics in combating bacterial drug resistance Band gap engineering of CeO 2 Nanostructure by electrochemically active biofilm for visible light applications Graphene/SiO 2 nanocomposites: the enhancement of photocatalytic and biomedical activity of SiO 2 nanoparticles by graphene Graphene nanoplatelets induced tailoring in photocatalytic activity and antibacterial characteristics of MgO/ graphene nanoplatelets nanocomposites Graphene/Fe3O4 nanocomposite: solar light driven Fenton like reaction for decontamination of water and inhibition of bacterial growth Synthesis of cerium oxide nanoparticles using Gloriosa superba L. leaf extract and their structural, optical and antibacterial properties Ultrasound assisted green synthesis of cerium oxide nanoparticles using Prosopis juliflora leaf extract and their structural, optical and antibacterial properties Cytotoxicity and antibacterial activity of gold-supported cerium oxide nanoparticles Ceria nanoparticle theranostics: harnessing antioxidant properties in biomedicine and beyond Cerium oxide nanoparticles as potential antibiotic adjuvant: effects of CeO 2 nanoparticles on bacterial outer membrane permeability Cerium oxide and peppermint oil loaded polyethylene oxide/graphene oxide electrospun nanofibrous mats as antibacterial wound dressings Cerium oxide nanoparticles inhibit differentiation of neural stem cells Biocompatible propulsion for biomedical micro/nano robotics Electrokinesis is a microbial behavior that requires extracellular electron transport Toxicity of Ce 2 O 3 /TiO 2 composite nanofibers against S. aureus and S. typhimurium: a novel electrospun material for disinfection of food pathogens Metal oxide nanomaterials in nanomedicine: applications in photodynamic therapy and potential toxicity The controversial antibacterial activity of graphenebased materials Co-delivery of free vancomycin and transcription factor decoy-nanostructured lipid carriers can enhance inhibition of methicillin resistant Staphylococcus aureus (MRSA) Hydrothermal synthesis of nanocrystalline cerium (IV) oxide powders Rapid removal of heavy metal cations and anions from aqueous solutions by an amino-functionalized magnetic nano-adsorbent Synthesis of nitrogen-doped ceria nanoparticles in deep eutectic solvent for the degradation of sulfamethaxazole under solar irradiation and additional antibacterial activities Antibacterial applications of graphene-based nanomaterials: recent achievements and challenges Antibacterial activity of ZnO nanoparticle suspensions on a broad spectrum of microorganisms Anchoring ceria nanoparticles on reduced graphene oxide and their electronic transport properties A green approach for the synthesis of a cerium oxide nanoparticle: characterization and antibacterial activity Microwave mediated synthesis and characterization of CeO 2 -GO hybrid composite for removal of chromium ions and its antibacterial efficiency Synthesis, characterization and antibacterial activity of cobalt doped cerium oxide (CeO 2 : Co) nanoparticles by using hydrothermal method Optical and electronic properties of post-annealed ZnO: Al thin films Superoxide dismutase mimetic properties exhibited by vacancy engineered ceria nanoparticles Comparison study on the antibacterial activity of nano-or bulkcerium oxide Use of noble metals in automobile exhaust catalysts Bacterial coinfection and secondary infection in patients with COVID-19: a living rapid review and meta-analysis Antibacterial activity of graphite, graphite oxide, graphene oxide, and reduced graphene oxide: membrane and oxidative stress Fabrication of CeO 2 nanoparticle-modified silk for UV protection and antibacterial applications A comparison of disc diffusion and microbroth dilution methods for the detection of antibiotic resistant subpopulations in Gram negative bacilli Facile synthesis of heterostructured cerium oxide/yttrium oxide nanocomposite in UV light induced photocatalytic degradation and catalytic reduction: synergistic effect of antimicrobial studies Tackling antimicrobial resistance by promoting antimicrobial stewardship in medical and allied health professional curricula Graphene-based functional nanomaterials for biomedical and bioanalysis applications. Flat Chem Thin-film gas sensors based on semiconducting metal oxides Anatase TiO 2 co-doped with silver and ceria for antibacterial application Anisotropic mechanical properties of graphene sheets from molecular dynamics Antibacterial study of CuO-NiO-ZnO trimetallic oxide nanoparticle Synthesis and characterization of CeO 2 nanoparticles via solution combustion method for photocatalytic and antibacterial activity studies Photodynamic antibacterial effect of graphene quantum dots Antibacterial mechanisms of graphene-based composite nanomaterials Enhanced sunlight photocatalytic activity and biosafety of marine-driven synthesized cerium oxide nanoparticles Transition metal-substituted cobalt ferrite nanoparticles for biomedical applications Bactericidal effect of graphene oxide and reduced graphene oxide: influence of shape of bacteria, Colloid and Interface Science Increasing the efficiency of some antibiotics on penetrating bacteria cell membrane, Ukrainian Alkali metal and semiconductor clusters in zeolites Pure and multi metal oxide nanoparticles: synthesis, antibacterial and cytotoxic properties Determination of cerium (III) and cerium (IV) in nanodisperse ceria by chemical methods Magnetic targeting of surfacemodified superparamagnetic iron oxide nanoparticles yields antibacterial efficacy against biofilms of gentamicin-resistant staphylococci Photocatalytic and antibacterial properties of phytosynthesized CeO 2 NPs using Moringa oleifera peel extract Graphene nanoplatelets/Cr 2 O 3 nanocomposites as novel nanoantibiotics: towards control of multiple drug resistant bacteria Todar's online textbook of bacteriology Staphylococcus aureus infections: epidemiology, pathophysiology, clinical manifestations, and management Electronicstructure-dependent bacterial cytotoxicity of single-walled carbon nanotubes Anti-microbial activities of aerosolized transition metal oxide nanoparticles Prepartion and electrochemical performance of a cerium oxide-graphene nanocomposite as the anode material of a lithium ion battery Inhibited growth of Pseudomonas aeruginosa by dextran-and polyacrylic acid-coated ceria nanoparticles Oxygen vacancies regulated microwave absorption properties of reduced graphene oxide/multi-walled carbon nanotubes/cerium oxide ternary nanocomposite Graphenecerium oxide/porous polyaniline composite as a novel electrode material for supercapacitor Synthesis of CeO 2 nanoparticles: photocatalytic and antibacterial activities In situ & controlled preparation of platinum nanoparticles dopping into graphene sheets@ cerium oxide nanocomposites sensitized screen printed electrode for nonenzymatic electrochemical sensing of hydrogen peroxide Size controllable preparation of gold nanoparticles loading on graphene sheets@ cerium oxide nanocomposites modified gold electrode for nonenzymatic hydrogen peroxide detection Long-term antibacterial stable reduced graphene oxide nanocomposites loaded with cuprous oxide nanoparticles Cerium oxidegraphene as the matrix for cholesterol sensor Antibacterial mechanism and activity of cerium oxide nanoparticles Production of metal oxides nanoparticles based on poly-alanine/chitosan/ reduced graphene oxide for photocatalysis degradation, antipathogenic bacterial and antioxidant studies The similarity index checked via Turnitin ID: