Single-Atom Enhanced Membrane for Simultaneous Bacteria and Heavy Metal On-Site Water Treatment.

Article Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment Graphical abstract Highlights •A portable graphene-based membrane removes bacteria and heavy metals in one step •Single-atom engineering enhances bacterial capture and binding efficiency •High reusability with >90% efficiency after multiple regeneration cycles •Simple, electricity-free operation enables safe water in remote environments Authors David Pana ´� cek, Renata Ve� ce� rova ´, Zden� ek Ba � dura, ..., Ale� s Pana ´� cek, Aristides Bakandritsos, Radek Zbo� ril Correspondence david.panac[email protected] (D.P.), a.bakandrits[email protected] (A.B.), [email protected] (R.Z.) In brief Clean water access remains a global challenge, especially in resource-limited areas. Pana ´� cek et al. present a graphenebased filtration membrane engineered with single-atom manganese that simultaneously removes bacteria and heavy metals in one step. The system requires only a hand-powered vacuum, achieves outstanding filtration efficiency across diverse water sources, and maintains reusability for many cycles. This portable, sustainable approach highlights how advanced nanomaterials can be translated into simple and impactful water purification technologies. Pana ´� cek et al., 2026, Chem 12, 102785 March 12, 2026 ©2025 The Author(s). Published by Elsevier Inc. https://doi.org/10.1016/j.chempr.2025.102785 ll Article Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment David Pana ´� cek, 1,2,3, *Renata Ve� ce� rova ´, 4 Zden� ek Ba � dura, 1,2 Vı´t� ezslav Hruby´ , 1 Luka ´� s Zdra� zil, 1,2 Milan Kola ´� r, 4 Thamraa Alshahrani, 5 Ale� s Pana ´� cek, 6 Aristides Bakandritsos, 1,2, *and Radek Zbo� ril 1,2,7, * 1 Regional Centre of Advanced Technologies and Materials, Czech Advanced Technology and Research Institute (CATRIN), Palacky´ University, � Slechtitel� u 241/27, Olomouc – Holice, 783 71 Olomouc, Czech Republic 2 Nanotechnology Centre, Centre of Energy and Environmental Technologies, V � SB–Technical University of Ostrava, 17. listopadu 2172/15, 708 00 Ostrava-Poruba, Czech Republic 3 Center for Advanced Technologies and Engineering (CATEN), Technologicka ´ 375/3, 708 00 Ostrava-Pustkovec, Czech Republic 4 Department of Microbiology, Faculty of Medicine and Dentistry, Palacky´ University, Hn� evotı´nska´ 3, 775 15 Olomouc, Czech Republic 5 Department of Physics, College of Science, Princess Nourah Bint Abdulrahman University, P.O. Box 84428, Riyadh 11671, Saudi Arabia 6 Department of Physical Chemistry, Faculty of Science, Palacky´ University, 17 listopadu 1192/12, 771 46 Olomouc, Czech Republic 7 Lead contact *Correspondence: david.pana[email protected] (D.P.), [email protected] (A.B.), [email protected] (R.Z.) https://doi.org/10.1016/j.chempr.2025.102785 SUMMARY Access to clean water remains a major global challenge, especially in remote and disaster-affected areas, where centralized water treatment is often unavailable. This study introduces a single-atom engineering approach to designing a filtration system capable of simultaneously purifying water from bacteria and heavy metals on site, thus providing a sustainable and user-friendly method for water purification. The system demonstrates exceptional efficiency in removing bacterial and heavy metal contaminants from various water sources. Our findings show remarkable filtration efficiency (>99.999%) against a broad spectrum of microorganisms in distilled, tap, and river water. Moreover, the membrane demonstrates a high adsorption capacity for heavy metals, specifically 661 and 248 mg g −1 for Pb 2+ and Cd 2+ , respectively. The simple operation and high membrane permeability, requiring only a hand-powered vacuum, ensure applicability in off-grid settings. In addition, the membrane material maintains more than 90% efficiency after 20 regeneration cycles, addressing both cost and sustainability issues. INTRODUCTION The lack of access to clean water and sanitation remains a pressing global challenge; consequently, ensuring water security is a primary objective of the United Nations Sustainable Development Goals 1 and the Horizon Europe mission. 2 Water scarcity is expected to deteriorate in the coming decades, even in water-rich regions. 3 Despite advances in water treatment methods, currently, 2.2 billion people still lack access to safe drinking water, and more than half of the world’s population lacks safe sanitation, according to a World Health Organization (WHO) report. The increasing negative impacts of human activity are making the provision of quality drinking water more challenging and fundamentally damaging the global economy, THE BIGGER PICTURE Access to safe drinking water remains an urgent global challenge, particularly in regions affected by disasters, conflict, or inadequate infrastructure. Conventional purification technologies often depend on electricity, complex facilities, or expensive materials, which limit their use in remote or resource-poor areas. Here, we present a straightforward yet powerful approach that combines graphene chemistry with single-atom engineering to create a lightweight, portable membrane. This material removes bacteria and toxic heavy metals in a single step using only a hand-powered vacuum. It delivers high purification efficiency across diverse water sources, including rivers and tap water, and retains performance over many reuse cycles, addressing both affordability and sustainability. This distinctive strategy unites microbial and heavy metal removal within one user-friendly platform. Built from benign materials and free from electricity or harsh reagents, it has clear potential for safe water delivery in off-grid communities and emergency settings. Chem 12, 102785, March 12, 2026 © 2025 The Author(s). Published by Elsevier Inc. 1 This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 ll OPEN ACCESS with an estimated 100 billion USD lost annually due to water contamination. 4 Rivers such as the Mississippi, the Ganges, and the Yellow River are among the world’s most polluted water sources, contaminated with bacteria and heavy metals from various sources, including sewage discharges, agricultural runoff, and industrial waste. 5 The coexistence of both contaminants in the rivers poses a significant risk to public health and the environment, underlining the crucial need for effective water management strategies. Disinfection techniques, including chlorination, ultraviolet radiation, and ozonation, have helped eliminate waterborne pathogens and improve health standards. 6 However, current water treatment processes rely on porous filters, 7,8 strong oxidants, 9 or other harsh conditions, 10,11 leading to a high carbon footprint, high energy consumption, and unpredictable health risks (e.g., carcinogenic by-products 12 or development of microbial resistance due to insufficient disinfection and thus the regrowth of pathogens. 13,14 ) The development of advanced materials and technologies is pivotal for efficient remediation and user-friendly water purification to secure our future water sustainability. Advances in the design of tailored inorganic and carbonbased materials have undoubtedly provided access to new and effective nanostructures for purification technologies, which, due to their simplicity and efficiency, are among the simplest and most effective methods of water disinfection 15–22 and heavy metals decontamination. 23–27 A prerequisite for the widespread use of these sophisticated and costly materials is the integration of high-value-added features such as sustainability (i.e., sorbent regeneration and reuse 20,28 ), user-friendliness (e.g., simple filtration 7,8,15 ), or rapid material efficiency (effective purification in seconds 11,29 ), which reduces processing time and, thus, final costs. A variety of nanomaterials, such as nanofibrous silica, 15,30 carbon nanotubes, 31 or hydrogels based on reduced graphene oxide functionalized with silver nanoparticles (AgNPs-rGO) 32 have been utilized to develop efficient filtration devices to sanitize water from bacteria. However, microorganisms have evolved defense mechanisms over billions of years, 33 even in the case of the potent antibacterial silver ions and AgNPs. 34–36 Other decontamination systems rely on the catalytic effects of a material, such as AgPd nanoparticles 10 or MoS 2 , 11,18 which effectively decontaminate water via forming reactive oxygen species (ROS). Although these materials appear to be highly effective in eliminating bacteria, the radicals generated display broad, non-specific toxicity. 35,37 In addition, both systems require light irradiation for ROS formation, further limiting their practical application. Methods using electric fields 38 or capacitive deionization 39 are effective in terms of purification speed. Functionalized graphene oxide with chitosan (CS), 39 deposited on a carbon electrode, exhibits strong antibacterial effects after applying an electrical voltage. However, this method achieved only 97.000% disinfection efficiency, which may subsequently lead to bacterial regrowth. 13,14 A novel method of rapid bacteria disinfection that outperforms traditional electric field treatment was reported, relying on nano-sized electrodes, allowing for significant amplification of the local electric field, achieving fast and effective elimination of bacteria in the order of nanoseconds. 29 The need for power, however, can be a significant limitation in remote and off-grid areas. Exploring new approaches for the effective purification of contaminated water is a key research area. 40–42 Cuttingedge technologies for bacteria decontamination are related to establishing a strong binding between the filtration material and bacteria (mainly to the organic groups on the outer layer of the bacterial membrane. 21,43,44 ) Among such sophisticated purification methods is a system inspired by the water transport and transpiration of trees. 45 The material (composed of CS, hydroxyapatite nanowires, and palladium nanoparticles) functions via the interaction between the negatively charged bacteria and the positively charged CS component, leading to the arrest of their vital functions (e.g., movement and reproduction) and thus to their death. Another approach involves the use of superparamagnetic nickel nanocrystals, rendering the ability to bind to Gram-positive and Gram-negative bacteria. 21 However, a potential limitation of such systems is the use of toxic elements such as palladium and nickel. Moreover, achieving an integrated system that combines effective and safe bacterial disinfection with heavy metal decontamination in a single step remains an unmet challenge. Although nanoparticles have enabled major advances in water treatment, 10,21,32 they often face inherent limitations arising from heterogeneous surfaces, wide size distributions, and aggregation under operating conditions. 46,47 These issues reduce accessible surface area, limit catalytic efficiency, and introduce variability, undermining reproducibility and long-term performance. 46 In contrast, single-atom catalysts (SACs) represent a fundamental advance, delivering maximal atomic efficiency, uniform coordination, and precisely localized active sites on suitable supports. 48,49 This atomic dispersion prevents aggregation and enables tunable local chemistry, affording precise control over binding affinity, redox behavior, and reaction selectivity. 40 These features enable high catalytic performance in decontamination processes, including Fentonlike mechanisms, 50 ROS production, 51 and selective multivalent coordination to pollutants. 44 As a result, SACs combine catalytic efficiency with enhanced sustainability and safety by minimizing toxic metal use and leaching. Their development represents a key opportunity toward precision-designed water purification systems that integrate performance, selectivity, and environmental compatibility. Inspired by these exceptional properties, we engineered a dual-functionalized graphene derivative loaded with single atoms of manganese and carboxylic groups (Mn-NGCOOH) as a biocompatible and very efficient trap for heavy metals (Cd 2+ and Pb 2+ ) and bacteria (Figure 1). Membranes prepared from this material were applied as a filtration system exhibiting exceptional efficiency in removing bacterial and heavy metal contaminants from various water sources, including distilled, tap, and river water. The membrane demonstrated exceptional filtration efficiency (>99.999%) for bacterial removal across various water sources while also exhibiting high adsorption capacity for heavy metals. Furthermore, the filtration system retains over 90% efficiency after six regeneration cycles, ensuring cost-effectiveness and long-term sustainability. The simple operation and high membrane permeability, requiring only a hand-powered vacuum, ensure its applicability in remote areas without any power source, representing Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 2 Chem 12, 102785, March 12, 2026 Article ll OPEN ACCESS a substantial advancement in portable water purification technologies. RESULTS Single-atom-enhanced filtration: Synthesis, structure, and filter construction The filtration device was designed in three key steps: (1) synthesis of nitrogen-doped graphene equipped with carboxyl groups (NGCOOH), (2) coordination of Mn 2+ ions with NGCOOH (MnNGCOOH), (3) assembly of the filtration membrane by integrating NGCOOH and Mn-NGCOOH (Figure 1 and Methods). Briefly, fluorographene reacted with sodium azide and then with nitric acid to afford the NGCOOH. X-ray photoelectron spectroscopy (XPS) showed extensive defluorination (0.37 atom %) and nitrogen doping (5.52 atom %) upon the transformation of fluorographene to NGCOOH. In addition, NGCOOH exhibits a high oxygen content (23.73%), predominantly in the form of carboxyl groups (Figures S1A and S2A), which are essential for the binding of metal atoms to its surface. 25,28 These structural features were also confirmed by Fourier transform infrared spectroscopy (FTIR; Figures 2A and 2B). The FTIR spectra of NGCOOH and Mn-NGCOOH exhibited features similar to common organic carboxylic acids, particularly the broad absorption band of O–H stretching between 2,700 and 3,700 cm −1 due to excessive hydrogen bonding of the groups. The absorption bands at 3,474 and 3,240 cm −1 correspond to various H-bonding configurations. 52 The strong band at 1,720 cm −1 is typical of the carboxylic groups. 52 Symmetric stretching vibration bands at 1,430 and 1,345 cm −1 , as well as C–O stretching at 1,245 cm −1 , appear at the same region with a broad feature of skeletal aromatic C=C vibrations. 52 Immobilization of Mn 2+ on NGCOOH resulted in a loading of 2 wt %, as determined by inductively coupled plasma mass spectrometry (ICP-MS). The carboxyl groups detected by FTIR were partially ionized to carboxylates after interaction with Mn 2+ , as indicated by the occurrence of vibrations at 1,600 and 1,430/1,345 cm −1 in Mn-NGCOOH (Figures 2A and 2B). Detailed characterization of NGCOOH and Mn-NGCOOH is available in our previous work by Pana ´� cek et al. 44 Figure 1. Schematic of the one-step water filtration process using Mn-NGCOOH and NGCOOH layers The illustration shows water flowing through the membrane with simultaneous removal of bacteria and heavy metals. Insets highlight carboxyl-metal binding, manganese coordination on the graphene surface, bacterial membrane structure, and the trapping of bacteria through Mn 2+ binding to outer-membrane polysaccharides. To shed more light on understanding the interaction of membranes with contaminants, the FTIR and XPS analyses were further used to observe changes in the signal of NGCOOH and MnNGCOOH after interaction with heavy metals and bacteria. The FTIR spectrum of bare NGCOOH showed pronounced peaks at 1,720 and 1,600 cm −1 with an intensity ratio of I 1600 /I 1720 = 1.03 (NGCOOH-HM, Figure 2AI). This ratio increases significantly upon interaction with heavy metals in the sample (NGCOOH-HM, Figure 2AII), reaching a value of 1.14. The increase in the COO − band at 1,600 cm −1 , together with the decrease in the intensity for the COOH region at 1,720 cm −1 , indicate strong interactions between the carboxyl groups on NGCOOH and the heavy metal ions. The binding of heavy metals to NGCOOH involves not only electrostatic interactions but also coordination via carboxyl groups to form stable complexes, exhibiting high affinity and selectivity of NGCOOH for metal ions. 28 While NGCOOH alone has limited efficiency in binding to bacteria, as has been found experimentally (see ‘‘one-step bacteria and heavy metals decontamination’’), functionalization with manganese atoms significantly enhanced its interaction with bacteria. The FTIR spectrum of Mn-NGCOOH (Figure 2BI) showed an intensity ratio of I 1600 /I 1720 = 1.08. When MnNGCOOH interacted with Escherichia coli (Mn-NGCOOHE. coli, Figure 2BII), this ratio increased significantly to 1.27. The higher intensity of the COO − –Mn bands can be attributed to the increased dipole moment of the metal carboxylate bond 53 after the coordination of multiple hydroxyl groups from the bacterial polysaccharides, which reside on the outer membrane. 44 In this case, the metal becomes even more electron-deficient due to the high electronegativity of the oxygen from the bacterial hydroxyl groups. The XPS analysis proved that heavy metals were anchored to NGCOOH after filtration due to the emergence of Cd 3d and Pb 4f spectral lines in the NGCOOH-HM survey spectrum (Figure S1B). In contrast, the C 1s and O 1s spectral regions of NGCOOH-HM (Figures 2C and S2, respectively) were technically identical to those of pristine NGCOOH. This implies that, despite their strong attachment, the heavy metals do not chemically alter the nature of the NGCOOH material, making the material recyclable upon desorption of the bound metal ions. The N 1s envelope of NGCOOH-HM (Figure 2D) was affected by its overlay with the Cd 3d 5/2 line, making the nitrite and nitrate species of NGCOOH unobservable. Otherwise, the rest of the N 1s line Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 Chem 12, 102785, March 12, 2026 3 Article ll OPEN ACCESS Figure 2. FTIR and XPS characterization (A) FTIR spectra of fresh NGCOOH (I) and of NGCOOH after heavy metals filtration (II). (B) FTIR spectra of fresh Mn-NGCOOH (III) and of MnNGCOOH after bacteria filtration (IV). (C and D) Deconvoluted HR-XPS of the C 1s region (C) and of the N 1s region (D) for pristine NGCOOH (I) and for NGCOOH after heavy metals filtration (II). (E and F) Deconvoluted HR-XPS of the C 1s region (E) and of the N 1s region (F) for Mn-NGCOOH (III) and for Mn-NGCOOH after bacteria filtration (IV). The deconvolution details can be found in Tables S1–S6 and the supplemental information. Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 4 Chem 12, 102785, March 12, 2026 Article ll OPEN ACCESS shape indicated an unchanged character of N-doping of the NGCOOH backbone after the filtration of the Cdand Pb-containing solution. After bacterial adsorption onto Mn-NGCOOH, no changes in the composition of the material have been detected (Figures S1C and S1D). Preservation of the levels of manganese in Mn-NGCOOH-E. coli indicates that its leaching is non-existent or very limited, ensuring its non-toxic concentrations in the filtrate. Furthermore, the presence of heavy metals in the filtrated water did not lead to their exchange with Mn. In this case, although Cd and Pb were anchored by the Mn-NGCOOH in small quantities, its Mn contents remained consistent (Figure S3). The C 1s spectral waveforms were practically identical in the cases of pristine NGCOOH, Mn-NGCOOH, and Mn-NGCOOH-E. coli (Figure 2E). Since Mn(NO 3 ) 2 was used for the preparation, nitrate contents were higher in the case of Mn-NGCOOH with respect to pristine NGCOOH (Figure 2F). The charge of Mn 2+ ions in MnNGCOOH is, therefore, still compensated by the nitrate ions in the antibacterial material. After the filtration, the nitrate components slightly decreased, indicating the ion exchange of nitrates with other ions, e.g., bicarbonates or chlorides, since these are the most prevalent anions in water. 54 The morphology and microstructure of Mn-NGCOOH were analyzed by high-resolution transmission electron microscopy (HR-TEM, Figure 3A), showing that Mn-NGCOOH comprised few-layered graphene flakes with a size of around 2 μm. The elemental mapping (Figures 3B–3F) shows the homogeneous coverage of the flakes by nitrogen and oxygen, corresponding to the spatial distribution of carbon. In particular, the Mn mapping (Figure 3F) shows the absence of dense Mn regions, indicating the absence of Mn-based nanoparticles. The filter membrane was prepared by controlled deposition of NGCOOH and Mn-NGCOOH on the surface of cellulose paper (see the schematic illustration of the final filter membrane in Figure S5 and more details in section ‘‘membrane construction’’ in the supplemental information). The surfaces were investigated by scanning electron microscopy (SEM, Figures 3G–3N), showing the homogeneous and dense coverage of the cellulose paper by the Mn-NGCOOH flakes (Figures 3G and 3H), which are distinctly different from the structure of neat cellulose paper (Figures 3J and 3K), showing a typical fibrous pattern. An optical photograph showing the Mn-NGCOOH membrane can be seen in Figure 3I, also showing macroscopically the homogeneous cellulose paper coverage. The cross section SEM images of the Mn-NGCOOH membrane (Figures 3M and 3N) deposited over the cellulose paper substrate (Figure 3L) showed a coating thickness of 10 μm. One-step bacteria and heavy metal decontamination With a focus on addressing the alarming effects of bacterial water contamination, NGCOOH and Mn-NGCOOH were evaluated for water decontamination from both Gram-positive and Gram-negative bacteria. The bacterial strains were selected among strains well-known to significantly contribute to water pollution and pose a health risk. E. coli and Enterococcus faecalis were included as major indicators of fecal contamination of water systems. 55 Staphylococcus aureus was selected for its ability to survive in water environments, even in the presence of antibiotics or under harsh conditions, such as higher temperatures. 56 Pseudomonas aeruginosa was chosen due to its significant resistance to disinfection agents and its tendency to form biofilms, complicating its elimination from water infrastructure. 57 In order to determine the effectiveness of the new graphenebased filtration system in removing bacterial and heavy metal contaminants from the water, the study included a series of methodological steps to verify the benefits of each component of the filtration membrane. The first step was to select the optimal metal atom based on the benchmarking experiment, in which six candidates (Mn(II), Fe(II), Cu(II), Zn(II), Ni(II), and Co (II)) were evaluated. Manganese atoms exhibited the highest filtration efficiency (see Figure S4), consistent with previous findings showing that manganese forms strong bonds with polysaccharides in the bacterial outer membrane. 44 In the second step, a conventional cellulose filter paper with a pore size of 2 μm was used to evaluate the filtration efficiency of the filter paper alone, which was then used as a substrate for the deposition of NGCOOH and Mn-NGCOOH. This initial test (Figure 4A) showed a low filtration efficiency of 63.3600% against E. coli since the effective pore size for bacteria exclusion is 100 nm and below. 58 Very high filtration efficiencies are critical since fast bacteria regrowth takes place. 13,14 When NGCOOH was deposited on the cellulose filter paper, the filtration efficiency reached 96.4900%, which was still insufficient to ensure complete bacteria removal to safe levels (Figure 4A). When Mn-NGCOOH was deposited on the cellulose membrane and tested on three types of contaminated water (distilled, tap, and river water), the filtration efficiency against E. coli was remarkably high, reaching 99.9999% in distilled, 99.9984% in tap, and 99.9994% in river water. These results demonstrate the superior properties of the Mn-NGCOOH membrane compared with the pristine NGCOOH, thus highlighting the key role of the manganese atoms in strongly binding bacteria 44 (Figure 4A). Importantly, the present membrane design outperforms stateof-the-art methods for water treatment (Table 1), such as nickel nanocrystals 21 functioning via bacterial binding. In other examples, carbon coated with Cu(OH) 2 nanowires 19 or graphene oxide functionalized with CS 39 displayed insufficient bacterial removal efficiency values below 97.0000%. Although methods using a locally enhanced electric field treatment 29 are attractive in terms of speed (in the order of nanoseconds), removal efficiencies of 95.0000% were still inferior and insufficient due to bacterial regrowth. 13,14 A composite consisting of MoS 2 nanoflakes on transparent Al 2 O 3 , equipped with Cu and magnetic Fe 3 O 4 nanoparticles (Al 2 O 3 @MoS 2 /Cu/Fe 3 O 4 ), represents a highly promising approach for water disinfection. 18 This advanced system achieves rapid and effective decontamination of E. coli, with a 99.9997% efficiency within 1 min. However, its mechanism of action, which is based on generating ROS, presents certain limitations. The formation of ROS is driven by sunlight, making the system dependent on unpredictable environmental conditions (e.g., variable light intensity/effectiveness) or electricity for artificial light. To further verify the versatility and efficiency of the MnNGCOOH filtration system, the experiments were conducted with water contaminated with additional bacterial strains, which Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 Chem 12, 102785, March 12, 2026 5 Article ll OPEN ACCESS Figure 3. The morphology and microstructure of Mn-NGCOOH (A) High-angle annular dark-field (HAADF) image of Mn-NGCOOH flake. (B–E) Energy dispersive X-ray spectroscopy (EDS) chemical mapping of Mn-NGCOOH for (B) carbon, (C) oxygen, and combined chemical mapping of (D) carbon and nitrogen and (E) carbon and oxygen. (F) EDS chemical mapping of Mn-NGCOOH for manganese. (legend continued on next page) Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 6 Chem 12, 102785, March 12, 2026 Article ll OPEN ACCESS significantly contribute to the contamination of both natural and hospital waters. 55,57 The filtration efficiency was 99.9997% for S. aureus, 99.9999% for P. aeruginosa, and 99.9995% for E. faecalis. These outstanding results for different bacterial strains highlight the broad-spectrum filtration efficiency of the Mn-NGCOOH, rendering it suitable for a broad range of decontamination scenarios (Figure 4B). A critical test involved evaluating the performance of the filtration system under real conditions (Figure 4C). Therefore, river water contaminated with bacteria and heavy metals (Cd 2+ and Pb 2+ at 1 ppm) was tested. This concentration of heavy metals was chosen because it significantly exceeds the allowed European limits for drinking water, currently set at 15 and 5 ppb for Pb 2+ and Cd 2+ , respectively. A critical test involved evaluating the performance of the filtration system under real conditions (Figure 4C). Therefore, river water contaminated with bacteria and heavy metals (Cd 2+ and Pb 2+ at 1 ppm) was tested. To exclude the possibility that the exceptional filtration capacity of the membrane arose from synergistic or antagonistic effects of toxic heavy metals, we exposed E. coli to cadmium and lead in the absence of any other material. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) tests confirmed that neither Cd 2+ nor Pb 2+ inhibited or killed E. coli at concentrations ≥2 ppm when applied individually, and that their 1:1 mixture at 1 ppm of each showed no detectable antibacterial activity (see supplemental information). Therefore, bacterial removal with the Mn-NGCOOH membrane takes place effectively even in the presence of heavy metals. Furthermore, in the absence of heavy metals, the removal is also similarly effective, also excluding synergy effects. Despite these challenging conditions, the final design of the filtration membrane, designed for maximum adsorption capacity of heavy metals and bacteria (see the schematic illustration of the final filter membrane in Figure S5 in the supplemental information), maintained high efficiency: 99.9995% for E. coli, 99.9950% for S. aureus, and 99.9978% for E. faecalis. In addition to Mn-NGCOOH, the NGCOOH membrane was used because carboxyl groups are very strong metal-coordinating ligands and show particularly high adsorption capacities for Cd 2+ and Pb 2+ . 25,28 A batch test with NGCOOH showed a high adsorption capacity (Figure 4D), reaching 661 mg g −1 for Pb 2+ (at 100 ppm Pb 2+ feed) and 248 mg g −1 for Cd 2+ (at 100 ppm Cd 2+ feed), outperforming state-of-the-art heavy metal sorbents. 26,59–61 The corresponding distribution coefficients were determined to be 12.8 L⋅g −1 for Pb 2+ and 3.8 L⋅g −1 for Cd 2+ , highlighting the notably stronger affinity of NGCOOH to Pb 2+ . Additional details on the adsorption isotherms and kinetic behavior of both metal ions on NGCOOH are provided in Tables S7 and S8 and Figure S6 (supplemental information). The adsorption properties of NGCOOH were further tested in the presence of competing ions or chemicals that are found in tap water (Ca 2+ , Mg 2+ , Na + , and K + ) and in river water (organic compounds or bacteria, Figure 4C). The results highlighted a very low decrease in the adsorption capabilities (21.9% and 12.9% decrease for Pb 2+ and Cd 2+ , respectively, in river water), suggesting the high potential of NGCOOH. According to the principle of hard and soft acids and bases, the selectivity can be explained as H 2 O molecules are considered as ‘‘hard base’’ ligands, therefore coordinating and keeping in solution the ‘‘hard acid’’ metal ions of Mg 2+ and Ca 2+ (which is also reflected by the high hydration energy of these cations), rather than the intermediate ‘‘soft acid’’ cations of Cd 2+ and Pb 2+ , which coordinate better with the carboxyl groups. 62 Natural organic matter (NOM) in river water can complex Pb 2+ /Cd 2+ and contribute to fouling. In our three-layer architecture, the top NGCOOH layer acts as a front-end scavenger, adsorbing both free metal ions (via carboxylate complexation; consistent selectivity with the hard-soft acid-base principle 63 ) and NOM-metal complexes through hydrogen bonding, π-π interactions, and coordination with the unoccupied part of the coordination sphere of the metals in the NOM-metal complexes. 64 This decouples NOM management from the Mn-NGCOOH middle layer, where bacterial capture proceeds via specific coordination to outer-membrane ligands, and the bottom NGCOOH layer provides polishing of residual species. This stepwise design explains the similar performance observed in distilled and river water and is consistent with established NOM-metal complexation and grapheneNOM adsorption behavior. 65 To evaluate the cost-wise applicability of NGCOOH and MnNGCOOH with respect to the low-cost activated carbons, complementary properties such as reusability were studied. Owing to its covalently bound, non-labile metal-coordination functionalities, NGCOOH could be treated even under harsh acidic conditions (2% v/v HNO 3 ) without affecting its structure and, thus, its adsorption properties. Therefore, the NGCOOH and MnNGCOOH collected after adsorption experiments were regenerated by acid washing to desorb metals and bacteria and then treated with 1% w/v NaOH to establish the carboxylate form, washed, dried, and reused for further filtration experiments. After each 1 L filtration of water containing 1 ×10 6 colony-forming units (CFUs)/mL bacteria and 1 ppm Pb 2+ and Cd 2+ , the active materials of Mn-NGCOOH and NGCOOH were regenerated by acid washing and redeposited onto fresh, low-cost cellulose paper to mitigate support clogging, sustaining more than 90% removal efficiency for at least twenty cycles. The characterization of the recovered material showed preservation of the chemical groups, with only minor changes (Figures S7 and S8; Tables S9–S11, see supplemental information). Indeed, NGCOOH and Mn-NGCOOH retained their adsorption capacities for Pb 2+ , Cd 2+ , and even bacteria for at least twenty regeneration cycles, maintaining over 90% efficiency, eliminating the relatively increased costs of the sorbent (Figure 4E; details of the recycling process can be found in the supplemental information). (G and H) SEM images of top view for Mn-NGCOOH. (I) A detailed photograph showing the Mn-NGCOOH membrane. (J and K) SEM images of top view for cellulose paper substrate. (L–N) Cross section of the (L) cellulose paper substrate, (M) Mn-NGCOOH, and (N) Mn-NGCOOH with higher magnification. Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 Chem 12, 102785, March 12, 2026 7 Article ll OPEN ACCESS The results of this study highlight the exceptional performance and versatility of a filtration system based on a combination of NGCOOH and Mn-NGCOOH. The significant improvement in bacterial filtration efficiency after functionalization with manganese single atoms underscores the critical role of these cations in the capture and subsequent removal of bacteria. The high efficiency of this membrane across various water types and bacterial strains in the presence of heavy metals demonstrates its potential for practical applications. Moreover, the operational simplicity of this on-site filtration system, which is based on an electricity-free, hand-powered filtration, makes it a feasible solution for water purification in remote, off-grid areas. DISCUSSION This study presents the development of a single-atom graphenebased filtration membrane that simultaneously removes bacteria and heavy metals, particularly effectively, from contaminated water. The graphene derivative equipped with carboxyl groups (NGCOOH) and manganese atoms (Mn-NGCOOH) exhibits a high adsorption capacity for heavy metals and bacteria. It has the ability to effectively filter various bacteria in the presence of heavy metals in distilled, tap, and river water, making it suitable for various pollution scenarios. The membrane can be regenerated and reused for at least twenty cycles while maintaining more than 90% of its initial adsorption capacity, thus addressing cost and sustainability issues. It is designed for user-friendly, onsite decontamination even in off-grid areas, eliminating the need for any electrical supply or specialized training, making it practical and cost effective. This study underscores the potential of single-atom engineering in producing advanced materials for environmental remediation and represents a significant step toward highly efficient, sustainable, and user-friendly water treatment solutions. Notably, it brings a previously untapped concept for single-step bacteria and heavy metal removal technologies, opening the door for future advances in filtration materials through their simple modification with benign metal cations. METHODS Further details regarding the methods can be found in the supplemental information. RESOURCE AVAILABILITY Lead contact Requests for further information and resources should be directed to and will be fulfilled by the lead contact, Radek Zbo� ril ([email protected]). Materials availability This study did not generate new unique materials. The graphene-based membranes described here can be prepared using published procedures, and detailed synthetic methods are provided in the Methods and supplemental information sections. Data and code availability All data supporting the findings of this study are available within the article and its supplemental information. Raw datasets are available at the Zenodo Figure 4. Bacteria and heavy metal decontamination (A) Filtration efficiency of distilled, tap, and river water contaminated with E. coli for cellulose paper (three cellulose papers on top of each other), bare NGCOOH (three NGCOOH membranes on top of each other), and NGCOOH decorated with manganese atoms: Mn-NGCOOH (three Mn-NGCOOH membranes on top of each other), represented by a logarithmic reduction in bacterial count compared with the initial concentration of bacteria (contaminated water without treatment). (B) Filtration efficiency of distilled water contaminated with S. aureus, P. aeruginosa, and E. faecalis for Mn-NGCOOH (three Mn-NGCOOH membranes on top of each other), represented by a logarithmic reduction in bacterial count compared with the initial concentration of bacteria (contaminated water without treatment). (C) Filtration efficiency of river water in the presence of Cd 2+ and Pb 2+ (at 1 ppm metal feed) contaminated with E. coli, S. aureus, and E. faecalis for the final filter membrane design (three graphene membranes on top of each other: Mn-NGCOOH/NGCOOH/Mn-NGCOOH, see the schematic illustration of the final filter membrane in Figure S5 in the supplemental information), represented by a logarithmic reduction in bacterial count compared with the initial concentration of bacteria (contaminated water without treatment). The initial bacterial concentration was 1 ×10 6 CFU/mL, and each experiment was performed in triplicate. (D) Adsorption capacities for Pb 2+ and Cd 2+ (heavy metals) in distilled, tap, and river water, as well as in combined Pb 2+ and Cd 2+ solutions in river water and combined Pb 2+ and Cd 2+ in the presence of E. coli in river water for the final filter membrane design. (E) Recycling experiments of the NGCOOH and Mn-NGCOOH for the sorption of Pb 2+ , Cd 2+ , and E. coli with 1% NaOH regeneration of the sorbent (initial [Men+] = 1 ppm). Recovery of the metals and bacteria by desorption from NGCOOH and Mn-NGCOOH using 2% v/v of HNO 3 (more details in Methods in the supplemental information). Please cite this article in press as: Pana ´� cek et al., Single-atom-enhanced membrane for simultaneous bacteria and heavy metal on-site water treatment, Chem (2025), https://doi.org/10.1016/j.chempr.2025.102785 8 Chem 12, 102785, March 12, 2026 Article ll OPEN ACCESS