Fe/Au Galvanic Nanocells to Generate Self-Sustained Fenton Reactions Without Additives at Neutral pH

The generation of reactive oxygen species (ROS) via the Fenton reaction has received significant attention for widespread applications. This reaction can be triggered by zero-valent metal nanoparticles by converting externally added H2O2 into hydroxyl radicals (˙OH) in acidic media. To avoid the addition of external additives or energy supply, developing self-sustained catalytic systems enabling onsite production of H2O2 at a neutral pH is crucial. Here, we present novel galvanic nanocells (GNCs) based on metallic Fe/Au bilayers on arrays of nanoporous silica nanostructures for the generation of self-sustained Fenton reactions. These GNCs exploit the large electrochemical potential difference between the Fe and Au layers to enable direct H2O2 production and efficient release of Fe2+ in water at neutral pH, thereby triggering the Fenton reaction. Additionally, the GNCs promote Fe2+/Fe3+ circulation and minimize side reactions that passivate the iron surface to enhance their reactivity. The capability to directly trigger the Fenton reaction in water at pH 7 is demonstrated by the fast degradation and mineralization of organic pollutants, by using tiny amounts of catalyst. The self-generated H2O2 and its transformation into ˙OH in a neutral environment provide a promising route not only in environmental remediation but also to produce therapeutic ROS and address the limitations of Fenton catalytic nanostructures.


Fabrication of Fe/Au galvanic nanocells to generate Fenton reactions
The Fe/Au GNCs were fabricated by combining colloidal self-assembly and physical vapor deposition techniques as shown schematically in Fig. 1a.Silicon wafers (Siegert Wafer GmbH) were used as substrates for the colloidal self-assembly of the porous silica nanoparticles.Initially, the substrate was thoroughly cleaned several times with acetone and isopropanol, then rinsed with Milli-Q water and dried with a nitrogen gun.The silicon wafer was immersed inside a water filler container and the aqueous suspension of colloidal LPSNPs was initially deposited at the liquid/air interface over the silicon wafer.Once a close-packed monolayer array of LPSNPs was formed at the air/water interface, the water was drained, thereby depositing the LPSNPs array on the silicon wafer.Then, the Si substrate with the array of LPSNP was cleaned with oxygen plasma (400 W, O 2 60 sccm) for five minutes (PS210, PVA Tepla America, Inc.) to fully remove any leftover surfactants.This process avoids the need for calcination and leaves a highly hydrophilic silica surface.Finally, the Fe (60 nm)/Au (20 nm) bilayer was deposited on the array of nanoporous SiO 2 beads using electron beam evaporation (UNIVEX 450, Leybold).Arrays in which only a Fe (60 nm) layer was deposited were used for comparison to reveal the galvanic effects.

Characterization of the Galvanic nanocells to generate Fenton reactions.
The morphology of the GNCs was analyzed by scanning electron microscopy (FEI Magellan 400L XHRSEM), and energy-dispersive x-ray spectroscopy (EDX) elemental mapping was monitored in the same microscope by an Oxford Instruments Ultim Extreme EDX detector system (Fig. S2).

Element
Atomic % Si 17.85 Fe 51.50 Au 30.65 Total: 100.00 We also carried out high resolution SEM of the similar samples with only the deposited Fe layer to show the high roughness of the Fe layer (Fig. S3).

Detection of the produced H 2 O 2
The combination of Amplex Red (10-acetyl-3,7-dihydroxyphenoxazine) and horseradish peroxidase (HRP) has been used as ultrasensitive assay to detect hydrogen peroxide (H 2 O 2 ), with a detection limit of around 50 nM.In the presence of HRP, the Amplex Red reagent reacts with H 2 O 2 in a 1:1 stoichiometry to produce the red-fluorescent oxidation product, resorufin.The concentration of H 2 O 2 is proportional to the generated resorufin.The fluorescence emission peak of resorufin is at 587 nm upon excitation at 488 nm.This process was monitored using confocal fluorescence microscopy (Leica, SP5) by focusing on the interface of the galvanic Fenton nanogenerators.
The procedure started with the preparation of fresh stock solutions: 10 mM Amplex Red reagent in dimethyl sulfoxide (DMSO), 1X

Detection of the produced Fe 2+ /Fe 3+ cations
Fingerprints of the iron ion release can be detected spectrophotometrically by measuring the absorbance of the water solution in contact with the nanostructures at different reaction times within a wavelength range of 200 to 350 nm.However, discrimination between Fe 2+ and Fe 3+ cannot be accomplished.To discriminate and quantify the released concentration of Fe 2+ and Fe 3+ , a colorimetric assay selective to Fe 2+ was used.The spectrophotometric measurements were performed with a UV-vis spectrophotometer (Cary 4000, PerkinElmer).In this assay, Fe 2+ reacted with o-phenanthroline to form a colored complex with an absorption peak at 511 nm.
The absorption at this specific wavelength was utilized for constructing a calibration curve, employing standard Fe 2+ solutions within the concentration range of 1.0 x 10 -7 M to 1.10 x 10 -4 M (Fig. S6 a and b).Hydroxylamine was added as reducing agent to the standard solutions to ensure that iron was in its 2+ oxidation state.To determine the Fe 2+ and Fe 3+ concentrations released by the Fe/Au and Fe nanogenerators, two aliquots of the water solution in contact with the Au/Fe and Fe nanostructured surfaces were extracted after 15 minutes of reaction.The first aliquot was complexed with o-phenanthroline and used to determine the Fe 2+ concentration produced in the unknown sample.The second aliquot was put in contact with o-phenanthroline and the reducing agent hydroxylamine.The use of the reducing agent allowed for obtaining the total concentration of iron cations in the form of Fe 2+ in the unknown sample.This includes the Fe 2+ actually produced during the catalytic process and the Fe 3+ that was produced during the catalytic process but converted into Fe 2+ with the reducing agent.The concentration of Fe 3+ was then obtained by subtracting the iron content of both aliquots.The total iron concentration for Au/Fe and Fe nanostructured surfaces was also obtained by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) measurements at different reaction times.

Catalytic activity tests
The catalytic performance of the Fe/Au GNCs was tested by analyzing the degradation and mineralization of two organic pollutants, Methylene Blue (MB) and Tetracycline (TC).For the degradation assays the evolution of the absorbance of MB and TC was analyzed, following the absorption peaks located at 664 and 362 nm, respectively.In these tests, the Fe and Fe/Au nanostructured catalytic surfaces deposited onto the silicon substrates (size 0.7x2 cm 2 ) were immersed in the pollutant solution with initial concentrations of 3.2 mg/mL MB or 17.7 mg/mL TC at room temperature (25 °C).The color change and decrease of the absorbance were monitored in 15 min intervals using UV-vis spectroscopy (Cary 4000, PerkinElmer) in quartzcuvettes.The degradation percentage (D%) of the pollutants was calculated by the following expression: D%=100 x (A 0 -A t )/A 0, where A 0 is the initial pollutant absorbance and A t the pollutant absorbance at the final time point.Total Organic Carbon Content TOC) was used to evaluate the mineralization efficiency and confirm the degradation of the contaminants using TOC-VCSH equipment (Shimadzu) with a high-sensitivity column.The catalytic tests were conducted in the presence of the oxygen naturally dissolved in water by the simple contact with atmospheric air.To determine the influence of the Fe and Au thickness in the reactivity, MB degradation tests for the following configurations were carried out: Fe 20 nm / Au 20 nm; Fe 40 nm / Au 20 nm; Fe 60 nm / Au 20 nm, and Fe 60 nm / Au 10 nm (Figs.S8 a and b).These assays showed a significant decrease in the degradation efficiency when either the Fe or the Au layers thickness were reduced.In addition, the effect of the Fe and Au layers position was also analyzed, showing that the degradation efficiency dropped when the Fe layer was deposited on top of the Au layer due to the fast passivation of the Fe surface.To better assess the influence of dissolved oxygen in water, additional experiments were carried out in an argon (Ar) atmosphere using a glove box (Fig. S9).

Scavenger experiments
Radical scavenging tests were carried out with the addition of scavenging agents in the MB solutions to identify the reactive oxygen species involved in the pollutant degradation.Accordingly, two different scavengers were used in presence of MB: benzoquinone (1 mM) as quencher of the superoxide radical (•O 2 -), and isopropanol (1 mM) as a quencher of the hydroxyl radical (•OH).

Figure S1 .
Figure S1.STEM image of the large porous silica particles.Scale bar a) 200 nm, b) 100 nm

Fig. S3 :
Fig. S3: High Resolution SEM image of the porous SiO 2 covered by the 60 nm thick Fe layer.

Figure S6 .
Figure S6.(a) Absorbance of the different standard Fe 2+ concentrations as a function of the wavelength, (b) calibration curve at 511 nm (the dashed line indicates the linear range of the calibration) and (c) Absorbance of the water samples undergoing reaction with Fe/Au and Fe nanogenerators.The absorbance peaks of the aliquots of the Fe 2+ /phenanthroline complex corresponding to the actual release of Fe 2+ by the Au/Fe and Fe nanogenerators are depicted by light blue and light red solid lines, respectively.Additionally, the absorption curves of the Fe 2+ /phenanthroline complex representing the total release of iron into the solution by the interaction with both nanogenerators (Au/Fe blue line and Fe red line) are displayed.As pointed

Figure S8 .
Figure S8.MB degradation rates for samples with different thickness and position of the Fe and Au layers.a) Degradation rates for Fe/Au layers with different Fe thickness.b) Degradation rates for Fe/Au layers with different Au thickness.c) Degradation rates for Fe/Au (Au on top) and Au/Fe (Fe on top) configurations.

Figure S9 .
Figure S9.Absorbance measurements of MB before undergoing degradation and after interacting with Fe and Fe/Au nanostructures for 90 minutes in Ar atmosphere.
Reaction Buffer, 10 U/mL HRP, and 20 mM H 2 O 2 in 1X buffer.The H 2 O 2 calibration curve was prepared by diluting the appropriate amount of 20 mM H 2 O 2 into 1 X buffer to produce H 2 O 2 concentrations of 0.5, 2.5, 5, 7.5, and 10 μM, including a no-H 2 O 2 control.Appropriate amounts of Amplex red and HRP were added to the standard solutions of H 2 O 2 to achieve a final concentration of 100 μM in the case of Amplex Red and 0.2 U/mL in the case of HRP.The reaction between H 2 O 2 , Amplex Red, and HRP was incubated to produce the resorufin, as a fluorescent probe of the H 2 O 2 standard solutions.The resulting fluorescence intensity of resorufin as a function of the concentration of the standard H 2 O 2 solutions is plotted in Fig.S5.Each value is the average of three fluorescence measurements.Then, the concentration of H 2 O 2 at the GNCs was determined by the addition of 100 μM Amplex Red and 0.2 U/mL HRP.The produced H 2 O 2 concentration was determined by fitting the linear part of the calibration plot (dashed line).The calibration curve was obtained by measuring resorufin fluorescence as a function of standard solutions of H 2 O 2 .Background fluorescence, determined for a no-H 2 O 2 control reaction, has been subtracted from each value as well as any background fluorescence from the support substrate of the Au/Fe and Fe nanostructures.The dashed line marks the linear range of the calibration.