Ricardo Alvarado Meza

Tom Santori

Xavier Cattoën
email: xavier.cattoen@neel.cnrs.fr

A kinetic approach to the mechanism of formation of mesoporous silica nanoparticles

Keywords: kinetic studies, mechanism, Mesoporous Silica Nanoparticles, hydrolysis, condensation



The synthesis of mesoporous silica nanoparticles was investigated by pH monitoring, which gives valuable complementary information to the currently accepted mechanism.

Introduction

Mesoporous silica nanoparticles (MSNs) are being widely investigated for applications in nanomedicine, owing to their elevated porosity, rigidity, biocompatibility and ease for further surface modifications [START_REF] Vallet-Regí | Engineering mesoporous silica nanoparticles for drug delivery: where are we after two decades?[END_REF]. Their synthesis is usually performed by sol-gel reactions from tetraethoxysilane (TEOS) in the presence of cetyltrimethylammonium bromide (CTAB, CTA + , Br -) as structuredirecting agent. Though multiple preparation methods have been reported to date in order to control their size and texture, [START_REF] Vallet-Regí | Engineering mesoporous silica nanoparticles for drug delivery: where are we after two decades?[END_REF][START_REF] Kachbouri | Synthesis and characterization of a new silica nanoparticles using APG/CTAB as modified agent[END_REF][START_REF] Beltrán-Osuna | Colloidal and spherical mesoporous silica particles: synthesis and new technologies for delivery applications[END_REF][START_REF] Wu | Synthesis of mesoporous silica nanoparticles[END_REF][START_REF] Pal | Recent Trends in Morphology-Controlled Synthesis and Application of Mesoporous Silica Nanoparticles[END_REF][START_REF] Florensa | Key Parameters for the Rational Design, Synthesis, and Functionalization of Biocompatible Mesoporous Silica Nanoparticles[END_REF] two main systems are employed: (i) ammonia-based recipes, where ammonia acts both as basic and nucleophilic catalyst; (ii) sodium hydroxide-based recipes, where the hydroxide ions mainly act as basic catalysts for the hydrolysis and condensation reactions. In the latter case, the usual protocol consists in adding TEOS to a basic solution of CTAB at 80 °C. [START_REF] Lai | A Mesoporous Silica Nanosphere-Based Carrier System with Chemically Removable CdS Nanoparticle Caps for Stimuli-Responsive Controlled Release of Neurotransmitters and Drug Molecules[END_REF] Multiple parameters have been varied to control the size and dispersity of the MSNs, and the use of statistical design of experiments helped to find optimal conditions. [START_REF] Catalano | Design Rules for Mesoporous Silica toward the Nanosize: A Systematic Study[END_REF][START_REF] Kim | Optimization of Mesoporous Silica Nanoparticles through Statistical Design of Experiment and the Application for the Anticancer Drug[END_REF][START_REF] Castillo | Production of MCM-41 Nanoparticles with Control of Particle Size and Structural Properties: Optimizing Operational Conditions during Scale-Up[END_REF] In particular, the sodium hydroxide initial concentration has been found to play a key role in controlling the size, a higher initial pH leading to larger MSNs. [START_REF] Catalano | Design Rules for Mesoporous Silica toward the Nanosize: A Systematic Study[END_REF][START_REF] Kim | Optimization of Mesoporous Silica Nanoparticles through Statistical Design of Experiment and the Application for the Anticancer Drug[END_REF][START_REF] Castillo | Production of MCM-41 Nanoparticles with Control of Particle Size and Structural Properties: Optimizing Operational Conditions during Scale-Up[END_REF][START_REF] Varache | Optimization of MCM-41 type silica nanoparticles for biological applications: Control of size and absence of aggregation and cell cytotoxicity[END_REF][START_REF] Ribeiro | Silica nanocarriers with user-defined precise diameters by controlled template self-assembly[END_REF][START_REF] Varache | Improving structural stability of waterdispersed MCM-41 silica nanoparticles through post-synthesis pH aging process[END_REF] Accordingly, the determining factor governing the MSNs' size is indeed the ionic strength at which silicate-micelles composites aggregate, a larger ionic strength leading to larger particles as a result of increased charge screening between individual linear CTA +silicate micelles, enabling larger assemblies. [START_REF] Ribeiro | Silica nanocarriers with user-defined precise diameters by controlled template self-assembly[END_REF] This was indeed shown by systematically varying the ionic strength using sodium chloride with identical initial pH. pH plays a crucial rule in silica syntheses, affecting both the kinetics of hydrolysis and condensation. The hydrolysis reaction, enabled both by acids and bases is the slowest under neutral conditions, [START_REF] Mcneil | Kinetics and mechanism of hydrolysis of a silicate triester, tris(2-methoxyethoxy)phenylsilane[END_REF] whereas the condensation reactions are the slowest near the isoelectric point of silica at pH~2. Under basic conditions, the hydrolysis rate increases linearly with the hydroxide concentration according to equation [START_REF] Vallet-Regí | Engineering mesoporous silica nanoparticles for drug delivery: where are we after two decades?[END_REF]. This allows monitoring the hydrolysis reaction kinetics by pH measurements. [START_REF] Qiao | Synthesis of Mesoporous Silica Nanoparticles via Controlled Hydrolysis and Condensation of Silicon Alkoxide[END_REF] In turn, the condensation reactions (equations ( 2) and ( 3)) release hydroxide anions, resulting in a pH increase. The balance between hydrolysis and condensation reactions can thus be easily monitored during the reaction. However, this simple technique has never been applied to understand the initial stages of the MSNs synthesis. [START_REF] Qiao | Synthesis of Mesoporous Silica Nanoparticles via Controlled Hydrolysis and Condensation of Silicon Alkoxide[END_REF][START_REF] Issa | Kinetics of Alkoxysilanes and Organoalkoxysilanes Polymerization: A Review[END_REF] Hydrolysis: ≡Si-OEt + OH - ≡Si-OH + EtO - ≡Si-O -+ EtOH (1)

Condensation: ≡Si-O -+ ≡Si-OEt + H 2 O  ≡Si-O-Si≡ + EtOH + HO -(2) ≡Si-O -+ ≡Si-OH  ≡Si-O-Si≡ + HO - (3)
Besides, TEM at various stages has been used to follow the initial stages of MSNs formation in the case of a synthesis using the dilution of a concentrated mixture right after nucleation. Under those conditions, small (ca 5 nm) ellipsoidal clusters were first observed, which then aggregated to form larger objects (ca 50 nm) which internally reorganized to yield surfactant-silicate nanocomposites with an internal 2D symmetry. [START_REF] Sadasivan | Nucleation of MCM-41 Nanoparticles by Internal Reorganization of Disordered and Nematic-Like Silica-Surfactant Clusters[END_REF] Small angle neutron scattering was used [START_REF] Hollamby | Growth of Mesoporous Silica Nanoparticles Monitored by Time-Resolved Small-Angle Neutron Scattering[END_REF] for systems employing tetramethoxysilane as precursor and triethanolamine as basic and nucleophilic catalyst, but which also forms atrane complexes thus involving a substantially different mechanism compared to the simple MSN synthesis. [START_REF] Cabrera | Generalised syntheses of ordered mesoporous oxides: the atrane route[END_REF] Interestingly, this study revealed that the mechanism likely begins with the adsorption of hydrolyzed or slightly condensed silanolates at the surface of positively charged CTA + micelles, followed by the aggregation of those micelles to form nanoobjects as the charge per micelle decreases. These objects likely further aggregate to form larger nanoparticles. SAXS studies were also employed which allowed to understand the evolution of the micellar phase during the process. [START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF] This study gave insights into the evolution of the CTA + micelles during the synthesis, but again for conditions (pH or temperature) significantly different from those usually applied for the synthesis of MSNs and with an elevated concentration of CTAB (82 mM) likely necessary to get sufficient scattering signal. Interestingly, this study revealed that the addition of TEOS would first lead to a swelling of the initially ellipsoidal CTA + micelles, with the hydrolysis of TEOS occurring at their surface. During the reaction, the micelles would then shrink as the TEOS is consumed and the obtained anionic silicates move towards the aqueous phase.

So far, the mechanistic investigations rely either on the initial and final state of the system varying synthesis parameters such as base concentration and salt addition, [START_REF] Catalano | Design Rules for Mesoporous Silica toward the Nanosize: A Systematic Study[END_REF][START_REF] Varache | Optimization of MCM-41 type silica nanoparticles for biological applications: Control of size and absence of aggregation and cell cytotoxicity[END_REF][START_REF] Ribeiro | Silica nanocarriers with user-defined precise diameters by controlled template self-assembly[END_REF][START_REF] Varache | Improving structural stability of waterdispersed MCM-41 silica nanoparticles through post-synthesis pH aging process[END_REF] and on SAXS or SANS in a flow cell and under conditions that slow down the processes. [START_REF] Hollamby | Growth of Mesoporous Silica Nanoparticles Monitored by Time-Resolved Small-Angle Neutron Scattering[END_REF][START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF][START_REF] Blin | Mechanism of self-assembly in the synthesis of silica mesoporous materials: in situ studies by X-ray and neutron scattering[END_REF] In the present study, we decided to monitor the formation of MSNs within the reaction vessel, trying not to disturb the reacting system. In particular we decided to use pH monitoring and light scattering to investigate the early stages of this sol-gel reaction. Together with the data found in the literature, the obtained results enable a deeper understanding on the interplay between physical processes and chemical reactions in these syntheses that lack of reproducibility, and for which scale up is still a significant challenge [START_REF] Castillo | Production of MCM-41 Nanoparticles with Control of Particle Size and Structural Properties: Optimizing Operational Conditions during Scale-Up[END_REF][START_REF] Varache | Optimization of MCM-41 type silica nanoparticles for biological applications: Control of size and absence of aggregation and cell cytotoxicity[END_REF].

Experimental section

Hexadecyltrimethylammonium bromide (CTAB, H5882), tetraethyl orthosilicate (TEOS), and all other reagents were purchased from Sigma-Aldrich. Fresh sodium hydroxide solutions were prepared every day (2.0 M, NaOH). Gradient HPLC grade water (Fisher) was used in all syntheses. The reaction temperature was controlled within the reaction medium and was set to 80 °C ± 1 °C unless otherwise stated.

The pH was monitored using a SevenEasy pHmeter from Mettler Toledo, with a pH inlab expert electrode working from 0 to 95 °C. pH values were transferred every second to a computer for further treatment. Scattering was measured using a VascoKin DLS equipment (Cordouan). The probe, which collects light backscattered at 173°, was located at a distance such that the focal point of the laser lied <1 mm from the wall of the reaction vessel. Field effect scanning electron microscopy (SEM) images were obtained on a Zeiss Gemini ULTRA plus electron microscope operating at 3.0 kV. The samples for SEM measurements were dropped and dried on a piece of silicon wafer. Mean particle sizes and standard deviations were obtained by measuring the dimensions of at least 80 particles in the SEM images. XRD data of the pure NPs were collected on a D8-Endeavor diffractometer with a mixed Kα1 and Kα2 Cu (λ = 1.5406 Å) source radiation, operated in a Bragg-Brentano geometry.

Typical MSN synthesis: A three-necks 250 mL round bottom flask containing a 20×40 mm olive stir bar was equipped with a thermometer and a pH probe. CTAB (333 mg, 0.92 mmol 5,7 mM), water (160 mL) and freshly prepared NaOH solution (2.0 M, 1.17 mL, 2.33 mmol, 14.6 mM) were added. The reaction mixture was heated to 80 °C ± 1 °C, with stirring at 700 rpm. After the temperature had stabilized, TEOS (1.67 mL, 7.50 mmol, 46.8 mM) was added at once. After 15 minutes, a condenser was adapted. The reaction mixture was stirred for a total duration of 2 h at 80 °C. After cooling, the MSNs were collected by centrifugation, resuspended in ethanol then centrifuged. The latter operation was repeated twice. Aliquots for SEM were taken at this stage. After drying (60 °C overnight), the surfactant was removed by calcination (2 h at 550 °C, heating rate 10 °C/min).

MSN synthesis with carbonated sodium hydroxide: The above protocol was followed, substituting a volume V of the NaOH solution (2.0 M) by 4V of NaHCO 3 solution (0.5 M).

Results and discussion

The classical MSN synthesis protocol was first evaluated, by adding TEOS (46.8 mM) to a solution of CTAB (5.7 mM), NaOH (14.6 mM) in water at 80 °C with stirring at 700 rpm. The measured starting pH value at 80 °C was 10.5 instead of the 12.1 value that would be expected at 25 °C, which is explained by the high self-ionization constant of water at 80 °C when compared to its value at room temperature (pK w 12.6 and 13.9 at 75 and 25 °C, respectively). [START_REF] Bandura | The Ionization Constant of Water over Wide Ranges of Temperature and Density[END_REF] Immediately after the one-shot addition of TEOS (t 0 = 0 s), a fast pH decrease occurs within the first minute, reaching a minimum at pH 9.3 before slowly raising to reach a final value of 9.5 at the end of the synthesis (Figure 1). This low final value is indeed due to the presence of deprotonated silanols within the nanoparticles, which consumed hydroxide anions from the reaction medium. Overall, these results tend to indicate that the first minute is dominated by the hydrolysis reactions, and that the condensation slowly takes over afterwards. It is noteworthy that the nucleation of NPs also takes place during this first minute, the mixture becoming cloudy after ca 45 s. Furthermore, no detectable temperature variation occurs during the first minutes, thus thermal effects can be neglected in this study. Considering the time resolution of the light scattering and pH measurements is close to the second, we decided to investigate in more details the kinetics of the early stages of the process (Figure 1). During the first seconds, a linear decrease of the pH is observed, suggesting that the hydrolysis of TEOS is the only process occurring with a partial first order relative to the hydroxide ions (Phase Ia). [START_REF] Issa | Kinetics of Alkoxysilanes and Organoalkoxysilanes Polymerization: A Review[END_REF][START_REF] Osterholtz | Kinetics of the hydrolysis and condensation of organofunctional alkoxysilanes: a review[END_REF] Then, and unexpectedly, a more pronounced decrease is observed after time t 1 = 17 s (Phase Ib), with a sudden change of slope at t 1 . At t 2 = 40 s, the pH stops to decrease, and reaches a plateau or even increases slightly for a few seconds (Phase II), before starting to decrease again, but with a much lower slope. It is noteworthy that a sharp increase in scattered light intensity occurs at t 2 , concomitantly with the plateau of the pH (Figure 2). This scattered intensity then reaches a plateau at the end of Phase II. As the scattered light intensity varies linearly with the concentration of scattering species and their diameter at the power 6, this result suggests that the nanoparticles have reached their final size yet at this stage. The pH variation between t 0 and t 2 enables determining that the (EtO) 3 SiO -concentration is 13 mM, ie 28% of the initial TEOS amount at nucleation, under the hypothesis that only singly hydrolyzed species are present at t 2 . This synthesis resulted in roughly spherical NPs with a diameter of 89±12 nm, with a 2D hexagonal structure of the pores (Figure 2 andS1c). Notably, the same reaction conducted in the absence of CTAB is characterized by a much slower kinetics, with a factor of ca 8 between the initial hydrolysis rates (Figure 1). To better understand the role of the surfactant, experiments were performed by varying its concentration between 2.9 and 7 mM (Figure 3 and S1), ie above the first critical micelle concentration determined at 80 °C and 14.8 mM NaOH to be 1.4 mM. A clear increase in rate constant is observed when the CTAB concentration is increased up to 6 mM, with no more increase above this value. Similarly, the stirring speed plays an important role, faster kinetics being observed when the stirring speed is increased. These observations suggest that the hydrolysis reaction mainly takes place at the surface of CTAB micelles, as TEOS is not miscible with water. Such a micellar catalysis was already suggested for the room-temperature hydrolysis and condensation of TEOS at high concentrations of CTAC (0.12 M), with an acceleration factor close to 2000 under such conditions. [START_REF] Cheng | The Role of Surfactant Micelles in the Synthesis of the Mesoporous Molecular Sieve MCM-41[END_REF] Accordingly, CTAB is able to encapsulate TEOS within inflated micelles, smaller micelles with higher surface over volume ratio being obtained when the stirring speed is increased, and more micelles being present as the concentration is increased, yielding an overall faster hydroxide consumption. The presence of micelles swollen with TEOS was indeed characterized by SAXS investigations, but for a much higher CTAB concentration. [START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF] Replacing CTAB by CTAC under identical concentrations led to slower kinetics. CTAC is known to form micelles with lower aggregation number, as a result of weaker coordination of chloride anions with respect to bromide. [START_REF] Zana | Fluorescence Probing Investigations of the Mechanism of Formation of Organized Mesoporous Silica[END_REF] Using CTAC, larger and more polydisperse NPs were obtained compared with CTAB, while keeping a good pore organization (Figure S2). The strength of stirring (Figure 3), which depends on the stir bar size, stirring speed and shape of the reaction vessel was found to play an important role on the reaction kinetics. Under otherwise constant conditions, we found that the reaction rate increases with the stirring speed between 200 and 400 rpm, being then constant above this value. An important factor leading to experimental discrepancies is the amount of carbonates in the sodium hydroxide solutions, which is reported to reach up to 40% mol . [START_REF] Varache | Optimization of MCM-41 type silica nanoparticles for biological applications: Control of size and absence of aggregation and cell cytotoxicity[END_REF] Thus we tested the influence of substituting part of the sodium hydroxide by sodium hydrogen carbonate to simulate the carbonatation of soda (Figure 4 andS3). Unexpectedly, we found that the hydrolysis kinetics were strongly enhanced by the presence of low amounts of sodium carbonate, the rate constant being almost doubled at 10% mol of carbonatation. This increase in rate constant is accompanied by a strong decrease in the size of the final MSNs. Indeed, starting from 90 ± 12 nm for freshly prepared sodium hydroxide, the diameter decreases down to 37±5 nm for sodium hydroxide bearing 40% of NaHCO 3 . These results corroborate those by Varache et al [START_REF] Varache | Optimization of MCM-41 type silica nanoparticles for biological applications: Control of size and absence of aggregation and cell cytotoxicity[END_REF] who reported that aggregated small MSNs were obtained for a highly carbonated sodium hydroxide solution. Altogether, these results show that freshly prepared sodium hydroxide solutions must be used for this kind of synthesis. The TEOS concentration was varied in the 10-80 mM range (Figure 5 and S4). A strong effect is observed on the hydrolysis kinetic constant as the TEOS concentration varies, with a linear increase at low to moderate concentrations, which is indeed coherent with first order kinetics with respect to TEOS [START_REF] Osterholtz | Kinetics of the hydrolysis and condensation of organofunctional alkoxysilanes: a review[END_REF]. However, the increase of kinetic constant stops for higher concentrations. The nucleation time t 2 is also strongly affected by the TEOS concentration, with no nucleation being observed for the lowest concentration of 10 mM. Indeed, this value is lower than the threshold of ca 12-13 mM of silicates at which the aggregates start to form for higher TEOS concentrations. At low TEOS concentration (20 mM), the MSNs feature a large size dispersion and a bad organization of the pores. For higher concentrations (46-80 mM), the diameter is constant (90 ±12 nm) and the MSNs feature a P6mm pore organization (Figure S4). Temperature has been reported as a determining factor for the pore organization and size of the MSNs [START_REF] Catalano | Design Rules for Mesoporous Silica toward the Nanosize: A Systematic Study[END_REF]. As expected, the reaction rate increases when the temperature increases over the range 60-90 °C (Figure S5), and the diameter also increases as already reported. However, and by contrast with the published results [START_REF] Catalano | Design Rules for Mesoporous Silica toward the Nanosize: A Systematic Study[END_REF], we found that the organization of the pores within the structure was worse at high temperature, a good pore organization after calcination being observed between 60 and 80 °C (Figure S6).

Discussion:

The formation mechanism of MCM-41 materials has been studied by various groups since the first proposals from the seminal 1992 articles [START_REF] Beck | A New Family of Mesoporous Molecular Sieves Prepared with Liquid Crystal Templates[END_REF][START_REF] Kresge | Ordered mesoporous molecular sieves synthesized by a liquid-crystal template mechanism[END_REF]. Two kinds of mechanisms were proposed: in the first one, called true liquid crystal templating, a liquid crystal of 2D hexagonal symmetry would be present in solution then hydrolyzed TEOS would hydrolyze and condense in the voids between the rod-like micelles to form the walls of the MCM-41 structure around the micellar template. In the second mechanism, referred to as cooperative self-assembly, the CTAB surfactant would be initially present as spherical micelles and it is only as the TEOS hydrolyzes that the micelles would undergo a transition from spherical to rod-like then to a 2D-hexagonal arrangement. Though numerous reviews and research articles inappropriately mention the true liquid crystal templating as the actual mechanism for MSN synthesis, the CTAB concentrations at which MSNs are prepared (2-20 mM, 0.7-7 g/L) are much closer to the first critical micellar concentration than to the concentration required to achieve liquid crystals in solution (ca 40 % wt) or even ellipsoidal micelles (0.1-0.4 M). [START_REF] Cheng | The Role of Surfactant Micelles in the Synthesis of the Mesoporous Molecular Sieve MCM-41[END_REF][START_REF] Raman | Template-Based Approaches to the Preparation of Amorphous, Nanoporous Silicas[END_REF][START_REF] Goyal | Shapes and sizes of micelles in CTAB solutions[END_REF] Even at a high surfactant concentration of 30 g/L, ie 80 mM, Yi et al showed by SAXS that only ellipsoidal micelles of CTAB swollen with TEOS were present at the beginning of the synthesis [START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF]. The formation of rod-like micelles was found to occur above 0.2 M at 30 °C, and at higher concentrations when the temperature increases, according to viscosity measurements. [START_REF] Kuperkar | Formation and Growth of Micelles in Dilute Aqueous CTAB Solutions in the Presence of NaNO3 and NaClO3[END_REF] In all above-mentioned mechanistic studies, the focus was placed on the evolution of the micellar component, but much less on the silicate species. Indeed, under typical conditions, [START_REF] Lai | A Mesoporous Silica Nanosphere-Based Carrier System with Chemically Removable CdS Nanoparticle Caps for Stimuli-Responsive Controlled Release of Neurotransmitters and Drug Molecules[END_REF] most of the structure is formed within 2-3 minutes which precludes the use of analytical tools such as 29 Si NMR in solution. The pH monitoring having a time response of ca 1 s, it makes possible to monitor in situ the early stages of hydrolysis and condensation by pH changes. Furthermore, in situ light scattering measurements allow to precisely determine the turning point at which the nanoparticles start to form. In Figure 2, the first 15 s of the reaction are marked by a linear decrease of the pH, which indicates that the hydrolysis is the only reaction taking place. The much slower hydrolysis of TEOS in the absence of CTAB indicates that TEOS hydrolysis occurs at the interface between the micelle and the aqueous domain, but not at the surface of hypothetical TEOS droplets. However, an efficient mixing is required in order to efficiently encapsulate the TEOS within the micelles, as evidenced by the decrease of the rate constant when the stirring speed is reduced to less than 400 rpm (Figure 3). The first hydrolysis regime (Phase Ia) can be adequately described as pseudo first-order. However, an abrupt transition is observed at t 1 towards a new pseudo first order regime, characterized by a faster pH decrease, which rules out the occurrence of condensation at this stage, which would tend to increase the pH. The hydrolysis of mono-hydrolyzed TEOS to dihydrolyzed TEOS [START_REF] Mcneil | Kinetics and mechanism of hydrolysis of a silicate triester, tris(2-methoxyethoxy)phenylsilane[END_REF] with a rate constant k 2 would only have an influence on the pH if a doubly deprotonated silanediol would be formed, according to equations ( 4) and [START_REF] Pal | Recent Trends in Morphology-Controlled Synthesis and Application of Mesoporous Silica Nanoparticles[END_REF].

=Si(OEt)(O) -+ OH - = + EtOH (4) =Si(OEt)(O) -+ H 2 O  =Si(OH)(O) -+ EtOH (5)
However, numerical simulations at various k 2 /k 1 ratio (Figure S7) show that in neither case would an abrupt change of slope occur. Therefore, we conclude that t 1 may correspond to a physical change in the shape of the CTAB micelles swollen with TEOS, that leads to more accessible TEOS molecules at their surface, and hence to faster kinetics. We therefore propose that a change from spherical to elongated micelles occurs at t 1 , after a concentration of ca 9 mM of singly-hydrolyzed TEOS has been reached. When the silicate concentration reaches ca 13 mM at t 2 , the nucleation of a new phase occurs, which is indicated by the strong raise of the turbidity detected by light scattering measurements (Figure 2). This phenomenon can be attributed to the aggregation of wormlike micelles into micelle bundles. [START_REF] Ribeiro | Silica nanocarriers with user-defined precise diameters by controlled template self-assembly[END_REF] Interestingly, the appearance of the new phase occurs simultaneously with a strong change in pH evolution. Indeed, the pH reaches a plateau around t 2 , which means that either the hydrolysis stops or that both hydrolysis and condensation reactions occur at the same rate from this time, which seems more likely. Indeed, as silicate-covered micelles aggregate, the condensation between adjacent silicates in the interstices between micelles becomes favored, with the release of hydroxide anions. These anions are then used in the hydrolysis of unreacted ethoxysilane moieties.

Based on our experimental results and on those published in the literature, [START_REF] Ribeiro | Silica nanocarriers with user-defined precise diameters by controlled template self-assembly[END_REF][START_REF] Sadasivan | Nucleation of MCM-41 Nanoparticles by Internal Reorganization of Disordered and Nematic-Like Silica-Surfactant Clusters[END_REF][START_REF] Hollamby | Growth of Mesoporous Silica Nanoparticles Monitored by Time-Resolved Small-Angle Neutron Scattering[END_REF][START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF] the mechanism depicted in Figure 6 can be proposed. TEOS droplets are first dispersed in water thanks to the CTAB surfactant within spherical micelles. [START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF] TEOS then gets hydrolyzed by the hydroxide ions (Equation ( 1)) at the interface between the hydrophobic core in the micelles and the aqueous medium, following first order kinetics on both TEOS and OH -(Phase Ia). The resulting singly-hydrolyzed silanolates adsorb at the micelles surface by strong electrostatic forces. At t 1 , a sudden change from spherical to short rod-like micelles occurs, as ca 8 mM of TEOS is hydrolyzed. This results in the redistribution of the TEOS molecules within the micelles, with an increase in hydrolysis rate (Phase Ib). At t 2 , the micelles aggregate as the hydrolyzed TEOS concentration reaches 13 mM and condensation between neighboring silanes start to occur, at a similar rate (Phase II). Then the pH decreases again slowly, with concomitant fast hydrolysis and condensation reactions. The pH reaches a minimum after 5 minutes, and slowly raises again as the condensation rate becomes faster than the hydrolysis. The pH reaches an asymptotic value of ca 9.5 after 2 hours of reaction, which is explained by the presence of deprotonated silanol functions at the surface of the MSNs and of their pores. Unfortunately, it was not possible to measure the amount of ethanol released over time, which would be a good indicator of the progress of the reaction, as ethanol is formed by both hydrolysis and

Figure 1 :

 1 Figure 1: Evolution of the pH during the MSN synthesis: (left) full view over the two hours of synthesis; (right) zoom on the first minute. Reaction performed (black) in the presence and (pink) in the absence of CTAB; The green and blue lines depict two linear regimes of the pH decrease. The red line depicts the calculated pH evolution in the case of a first order reaction for both sodium hydroxide and TEOS.

Figure 2 :

 2 Figure 2: (left) Reaction kinetics monitored by (black dots) pH and (purple squares) light scattering. The red line shows the evolution of the triethoxysilanolate concentration, calculated from the pH decrease. (right) SEM micrograph of the MSNs obtained after synthesis, with a diameter of 89±12 nm.

Figure 3 :

 3 Figure 3: (Left) Dependence of the pH decrease on the CTAB concentration with [CTAB]= (black) 0.0; (red) 2.9; (blue) 4.0; (pink) 5.7; (purple) 7.0 mM. Inset: Evolution of the kinetic constant depending on the CTAB concentration. (Right) Dependence of the pH decrease on the stirring speed with r = (black) 200 rpm; (red) 400 rpm; (blue) 700 rpm. Inset: relative kinetic constant depending on the stirring rate.

Figure 4 :

 4 Figure 4: Dependence of the size and rate constant on the carbonatation of sodium hydroxide. (black squares) diameter; (red dots) relative rate constant.

Figure 5 :

 5 Figure 5: Effect of the TEOS concentration on the reaction kinetics. (black) 10 mM; (red) 20 mM; (blue) 47 mM; (pink) 60 mM; green 80 mM. Inset: dependency of the first relative kinetic constant on the TEOS concentration. The blue line depicts the linear fit on the first data points.

Acknowledgements

Sabine Douillet and Dr Jérôme Kieffer (ESRF) are gratefully acknowledged for their help.

Funding: This work was supported by the Agence Nationale de la Recherche (ANR-20-CE09-0017-03).

Competing Interests:

The authors have no relevant financial or non-financial interests to disclose.

condensation reactions. Nevertheless, we can assume that the hydrolysis reaction accelerates after t 2 , as the micelles are in close contact and hydroxide anions are liberated at the surface of the CTAB micelles swollen with TEOS. The proposed mechanism is in good agreement with the swellingshrinking model proposed by Yi et al. [START_REF] Yi | A New Insight into Growth Mechanism and Kinetics of Mesoporous Silica Nanoparticles by in Situ Small Angle X-ray Scattering[END_REF] It is also fully compatible with the nucleation model proposed by Sadasivan et al [START_REF] Sadasivan | Nucleation of MCM-41 Nanoparticles by Internal Reorganization of Disordered and Nematic-Like Silica-Surfactant Clusters[END_REF], though the experimental conditions are substantially different. In conclusion, monitoring the pH during the synthesis of MSNs enables to understand the physicochemical processes occurring during the early stages of their formation. The hydrolysis mechanism is bimolecular with a first order on both TEOS and OH -at the early stages, with a strong dependence of the rate constant on the CTAB concentration which is a signature of micellar catalysis. Importantly, we found that sol-gel condensation reactions only start after the nucleation of CTA + -silicate micelles as hexagonal 2D assemblies of elongated micelles. The nucleation indeed occurs at low TEOS conversion. Though no correlation could be found between the hydrolysis rate and the NPs size or pore organization, some factors strongly affecting the kinetics such as stirring speed and carbonatation of the sodium hydroxide solution may explain the lack of reproducibility of the MSNs synthesis depending on the authors. Much care must therefore be taken to control well all these factors to reproduce or scale up such MSNs synthesis.

Author Contributions: All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Ricardo Alvarado Meza, Tom Santori and Xavier Cattoën. The first draft of the manuscript was written by Xavier Cattoën and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.


Dr Daniele S Oliveira

Dr Thibaud Aumond

Dr Lola Loupias

Dr Leonardo L Santos

Dr Vinícius P S Caldeira

Dr Christine A Canaff

Dr Sibele B C Pergher

Dr Aurélien Habrioux
email: aurelien.habrioux@univ-poitiers.fr

Alexander Sachse
email: alexander.sachse@univ-poitiers.fr

Sachse

Bio-Sourced Zeolite-Templated Carbons for the Development of Efficient ORR Catalysts

Keywords: Zeolite-templated carbons, Glycerol, Carbon, ORR, Graphene

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

The production of bio-derived glycerol has been steadily growing over the past 20 years and production exceeds currently global demand by a factor of six. [1] This explains the quest for sound glycerol valorization strategies. Numerous approaches aim at the conversion of glycerol for the production of added-value chemicals and fuels. Catalytic strategies based on steam reforming, aqueous phase reforming, dry reforming, hydrogenolysis and selective oxidation are largely covered topics in the scientific literature. [2] The development of biomass-derived carbon materials is currently one of the hot topics in advanced materials design. [3] The use of lignocellulose-derived precursors [4] or tannins [5] is widely studied for the production of sustainable materials with defined textural properties. The transformation of glycerol into added-value materials has only scarcely been reported. A limited number of reports present the development of activated carbons from glycerol. [6][7][8] Gonçalves et al. [9] recently reported the use of crude glycerol for the synthesis of activated carbons, which proved promising for the development of bio-derived supercapacitor electrodes. Activated carbons yet feature highly disordered structures of cross-linked, randomly packed graphene sub-units and hence present unstructured porous topologies. [10] Since the beginning of the current century a new class of carbon materials was disclosed and named Zeolite-Templated Carbons (ZTCs). [11] The first synthesis of high quality ZTC was described using a twostep method in which the zeolite is firstly impregnated with polymer, and secondly followed by chemical vapor deposition of a gas. [12] Later, a one-step method was described allowing the formation of high-quality ZTC through the use of ethylene or acetylene, which diffuse and polymerize within the microporous network of a sacrificial zeolite. [13,14] After carbon formation within the zeolite the template is dissolved and the ZTC is recovered. ZTCs are a negative copy of the zeolite texture and hence feature tailored porous topologies. They have been described as 3D graphenes. [15] The combination of excellent electrical conductivity and unparalleled textural properties makes ZTCs very promising for the design of energy conversion and storage devices. [16][17][18][19][20] The use of alternative carbon sources for the synthesis of ZTCs was recently proposed. [21,22] Oxygen containing hydrocarbons, such as methanol, ethanol, isopropanol, acetone, tetrahydrofuran, and diisopropyl ether were tested in the synthesis of ZTCs by Ryoo and co-workers. [23] The authors found that these lead to the generation of olefins at the applied synthesis condition, i.e. 650-700 °C. It is these olefins that then react within the zeolite microporosity and allow for the formation of the carbon skeleton. In the present contribution we firstly study the synthesis of biosourced ZTCs from glycerol synthesized in the temperature range from 300 to 800 °C. The use of glycerol permits the synthesis of structured ZTCs even at temperatures as low as 400 °C. The thorough characterization of the hybrids (zeolite/carbon) and pure carbons allowed to assess that the synthesis temperature importantly impacts textural and chemical properties. After functionalization with thiourea the bio-sourced ZTCs allow for conspicuous ORR activities.

Results and Discussion

Characterization of the hybrid materials

The structural packing density (SPD) gives the amount of carbonaceous material per unit of zeolite mass and can be calculated from the TGA mass loss curves of the hybrid materials. The SPD increases from 0.16 to almost 0.32 gC gZ -1 for the hybrids achieved at 300 and 500 °C, respectively (Figure 1a). This suggests that at 300 °C the zeolite is only partially filled with carbonaceous compounds. Values of SPDs from 0.30 to 0.34 gC gZ -1 are typically observed for hybrid materials leading to high quality ZTCs achieved from ethylene in FAU structured zeolites. [24] With increasing synthesis temperature the SPD slightly decreases to reach 0.27 gC gZ -1 at 800 °C. It is further to note that the mean temperature of combustion is impacted by the synthesis temperature (Figure 1b). From 300 to 500 °C the mean temperature of combustion decreases slightly from 540 to 532 °C, respectively. This value increases thereafter importantly and reaches 565 °C for the material achieved at 800 °C. This variation in the value for the mean combustion temperature indicates that the carbon synthetized at higher temperatures presents a higher degree of condensation of the carbonaceous species in the zeolite channels. [25] From the nitrogen physisorption isotherms at 77 K it is possible to observe that only the hybrid achieved at 300 °C presents incomplete zeolite micropore obstruction (Figure 2a). The sample features a remaining micropore volume of 0.10 cm 3 g -1 , hence 62% of the zeolite micropore volume is blocked. For hybrids achieved above 400 °C the entirety of the micropore volume is inaccessible to nitrogen. It is interesting to note that hybrid materials present, independently of the synthesis temperature, a similar apparent density of the carbonaceous phase between 1 and 1.1 gC cm -3 (Figure 2b). This finding indicates similar topology of the carbonaceous species forming within the zeolite micropores.

Characterization of the carbon materials

From the nitrogen physisorption isotherm of the ZTC-300 sample it is interesting to observe that the desorption branch does not loop the adsorption branch at the relative pressure of 0.43 (Figure 3a). This is classically observed for not fully crosslinked polymers and can be ascribed to deformation/swelling of the network during adsorption. [26] This hence indicates that the carbon skeleton is not fully condensed at 300 °C. For higher synthesis temperatures this phenomenon is much reduced, suggesting that the carbon skeleton presents higher rigidity. The micropore volume is lowest for the ZTC-300 material (0.65 cm 3 g -1 ) (Figure 3b, Table S1). Likewise this sample does not allow for observing any peaks in the XRD powder pattern, indicating that no structural long-range order is present in this sample (Figure 3c). For carbons achieved at 400 °C (ZTC-400 sample) and above the micropore volume amounts to approximately 1 cm 3 g -1 and for all the samples a characteristic XRD peak at approximately 6.5° 2can be observed which corresponds to the channelto-channel structure of the zeolite template. [27] The position of the peak shifts from 7.0° at 400 °C to 6.4° 2at 700 and 800 °C, indicating expansion of the pore-to-pore distance in the ZTC with increasing synthesis temperature. Similar can be inferred from the NLDFT pore size distribution (Figure S5). The absence of a peak at approximately 25° 2in the XRD patterns indicates the absence of extra ZTC carbon on the external surface. This confirms the good quality of the ZTCs obtained using glycerol as carbon source. The SEM images allow to observe that for all synthesis times replicas of the zeolite morphology are achieved. By closer observing the sample achieved at 300 °C one can note that the edges of the particles are smoother than for samples synthetized at higher temperatures (Figure 4). Further insights on the texture were achieved from TEM, which allows for the observation of individual particles that present similar textural properties compared to the template zeolite (Figure 5). For the sample achieved at 300 °C (ZTC-300) unstructured microporosity can be observed. It is further interesting to note the absence of mesoporosity for this sample. The low SPD and the incomplete filling of the zeolite microporosity suggest that these mesopores are probably lost during the zeolite dissolution process by the reaction of radical polyaromatic species. [25] This hence indicates that the carbon species developing at 300 °C rather consist of smaller, uncondensed polyaromatic hydrocarbons. The TEM images of the ZTC-400 sample present the morphological features of the zeolite and the transcription of steamed mesopores. The high resolution TEM image of this sample allows for observing long-range structured microporosity. The TEM image of the ZTC-800 material allows to observe next to the carbon particles resulting from the zeolite templating the presence of turbostratic carbon. [28] This high temperature synthesis hence leads to the presence of extra-ZTC carbon species. The zeolite dissolution allows to remove the entirety of inorganic species from the carbon compounds. The TGA curves present complete weight loss, indicating full oxidation of the carbonaceous species extracted from the hybrid materials (Figure S4). The mean temperature of combustion presents a similar tendency as for the hybrid materials. Its value is minimal for ZTC-500 and maximal for ZTC-800 and amounts to 515 and 558 °C, respectively (Figure 6). This suggests that the carbons achieved at temperature below 500 °C present a higher oxygen content, whilst carbons prepared at higher temperatures present a higher degree of condensed aromatic cycles. [29] In the synthesis of ZTCs using glycerol as carbon precursor, glycerol was directly injected to the reactor at the target temperature. Hence, transformation of glycerol as a function of the temperature probably occurs. Dou et al. [30] studied the thermal degradation of glycerol under nitrogen atmosphere by TGA. Complete degradation was observed for pure glycerol in a temperature range of 130 to 250 °C. The authors studied the composition of the degradation products by FTIR and reported the presence of compounds containing C=C and C=O bonds next to H2O, CO2 and CH4 in the outflow. The pyrolysis of glycerol in the range of 650 to 700 °C has been described by Dalai and co-workers. [31] The authors detected the presence of essentially CO, H2, CO2, CH4 and C2H4. It can hence be assumed that degradation products presenting olefinic and carbonylic functions constitute the precursor mixture of the carbons achieved at temperature below 400 °C. At higher syntheses temperatures (500 °C and above) it can be assumed that the carbons are majorly formed from ethylene. This is confirmed by the C/H and the C/O ratios of the carbons (Figure 7). Indeed, the C/O ratio increases from 9 to 17 for ZTC-300 and ZTC-800. The materials achieved at lower synthesis temperatures present thus a higher degree of oxygenated carbon. The C/H ratios only little evolve as a function of the synthesis temperature from 3.0 to 4.5, indicating a higher degree of condensed carbon skeleton with increasing temperature. To further investigate the chemical nature of the samples Raman spectra were recorded at 532 nm (Figure 8a). Three features can be distinguished in the Raman shift range between 1100 and 1700 cm -1 and are ascribed to the G, D and S bands. The G band can be related to the vibrational modes associated with the asymmetric stretching of all sp 2 carbon atoms in the sample, while the D band is attributed to the breathing mode in aromatic rings. [32-34] The G band is centered at 1600 cm -1 for all samples, which is a typical value for large polyaromatic hydrocarbons or nanocrystalline graphite. [35] It is interesting to note that the D band position is impacted by the synthesis temperature and red-shifts for materials achieved with increasing temperature (Figure 8b). This suggests that the cluster size of polyaromatic entities increases with increasing temperature, [36] which might hint the presence of sp 3 carbons in samples achieved at lower temperatures. The apparition of the S band in all of the samples is related to the topology of the carbon species that can develop in zeolite pores and are composed of elongated polyaromatic units, and is typically observed in rylenetype molecules or carbon nanoribbons. [16,28] The intensity ratio of the D and G bands increases slightly with increasing synthesis temperature, indicating a higher degree of edges in carbons achieved at higher temperatures (Figure 8c). A similar tendency is observed for the intensity ratio of S and G bands, which suggests a higher amount of condensed aromatic units with pseudo one-dimensional D2h point symmetry. [37] Figure 8. Raman spectra of materials synthesized at different synthesis temperatures (a). Raman D band position (b) and ID/IG and IS/IG intensity ratios as a function of the synthesis temperature (c). Color code: 300 °C (black), 400 °C (red), 500 °C (blue), 700 °C (green) and 800 °C (purple).

Chemical evolution of materials was further studied through XAES (Figure S6). From the first derivative of Auger CKLL peak the energy difference between the positive and negative maximum can be measured, which allows to calculate the D parameter (DP) value (Table S2). From this latter the sp 2 /sp 3 ratio of carbons can be estimated, [38,39] and amounts to 0.80, 0.83 and 0.87 for ZTC-300, -400 and -800, respectively. Similar information can be deduced from the C1s photoelectron peak position, which are centered at 284.78, 284.65 and 284.48 eV for ZTC-300, -400 and -800, respectively (Figure S7). Pure graphene features a characteristic C1s peak at 284.40 eV (aromatic sp 2 carbon), whilst for graphene presenting structural defects and higher hydrogen content the peak shifts to higher binding energies. [40] Hence, the shift to lower energies with growing synthesis temperature indicates decreasing sp 3 character of the sample. The evolution of C/O ratio observed from XPS is in agreement with what was deduced from elemental analysis, even if absolute values are higher in XPS (Table S3).

Functionalization of carbon materials with thiourea

Glycerol derived ZTCs were treated with thiourea with the aim to functionalize them with nitrogen and sulfur atoms. The textural properties are slightly impacted by the treatment and a reduction of the equivalent BET surface area and of the micropore volume of 5% was observed for all samples. No impact of the XRD long range order was observed upon functionalization. It appears that the oxygen content in the ZTCs plays a crucial role in the concentration of N and S atoms in functional materials (Table S2). Approximately 10at% and 4at% of N and S were detected by XPS, for ZTC-300-TU and ZTC-400-TU, which both feature a rather high amount of oxygen in the unfunctionalized samples (ZTC-300 and ZTC-400). By revenge, ZTC-800, which features a low amount of oxygen only permits for the functionalization with 2.6 and 1.8at.% of N and S, respectively. This hence indicates that N/S insertion requires the presence of oxygenated groups at the surface of the carbon backbone. Furthermore, after thiourea doping the C/O atomic ratio strongly decreases for all samples (Table S3). This could result from the substitution of some carbons with S and/or N atoms during the thermal treatment performed at 700 °C in the presence of thiourea. To gather further information concerning the speciation of N and S atoms in thiourea-doped samples, XPS spectra of the S2p and N1s regions were recorded (Figure 9a). N1s spectra recorded with thiourea-doped ZTC materials were decomposed using four photopeaks, which are respectively assigned to pyridinic nitrogen (located at ca. 398.2 ± 0.1 eV), pyrrolic nitrogen (located at ca. 399.9 ± 0.1 eV), graphitic nitrogen (located at ca. 401.8 ± 0.5 eV) and oxidized nitrogen species (located at ca. 404.5 ± 0.5 eV). [41] This latter contribution may be attributed to N atoms bonded with two C atoms and one O atom. [42] From values reported in Table S4, it can be observed that the speciation of nitrogen only slightly depends on the ZTC synthesis temperature. For all materials, pyrrolic and pyridinic groups are mainly encountered. The main evolution concerns graphitic speciation with 7.8, 10.8 and 12.8atN.% for ZTC-300, ZTC-400 and ZTC-800, respectively. In addition, no covalent bond between nitrogen and sulfur atoms could be detected. The high resolution spectrum of S2p region can be decomposed using multiple lines (Figure 9b). Two major peaks are located at ca. 163.9 and 165.1 eV, which represent the S2p3/2 and S2p1/2 doublet, with spin orbit splitting at ca. 1.2 eV. This doublet is associated with the presence of C-S like groups (C-S-C and C-S-H groups). [43,44] The contribution at higher binding energy can be ascribed to the presence oxidized S atoms bounded to carbon atoms (-C-SOn-). As described in Table S5, C-S-X (with X = C or H) functionalities are the most abundant speciation for sulfur atoms whatever the considered sample. It can be however noticed that for ZTC-800-TU a higher amount of C-S-X groups can be distinguished.

Electrocatalytic tests

Polarization curves were recorded at 1600 rpm for each ZTCbased catalyst (Figure 10). Measurements with Pt/C and rGO-TU (i.e. reduced graphene oxide functionalized with thiourea in the same way as for the ZTCs) were realized as reference catalysts.

As far as the unfunctionalized ZTC samples are concerned, the synthesis temperature greatly affects the ORR activity (Figure 10a). The half wave potential increases from 0.66 to 0.82 V vs. RHE for ZTC-300 and ZTC-800, respectively (Figure 10b). Two different phenomena are responsible for the observed difference in activity. Firstly, the material synthesized at 300 °C (ZTC-300) is not fully condensed, which most probably affects the electrical conductivity of the material. In addition, ZTC-300 presents a low microporous volume (0.65 cm 3 g -1 ), hence the active surface area is lower than for ZTC-400 for which a microporous volume of around 1 cm 3 g -1 was calculated. ZTC-800 material presents a rigid carbon skeleton mainly composed of sp 2 hybridized carbon atoms with high micropore volume. The sample moreover features turbostratic carbon on the external surface, which favors electrical conductivity. The doping with thiourea improves the ORR activity of materials synthesized at 300 and 400 °C (ZTC-300-TU and ZTC-400-TU), (Figure 10). The interplay between nitrogen and sulfur atoms is known to allow for increasing the spin and charge densities of graphene networks, thus enabling a large number of carbon atoms to become active sites. [45] The first synthesis step involves a heat treatment at 300 °C and 400 °C, respectively. During the second step, the doping with N and S atoms is performed at 750 °C. This step does not only engender the insertion of heterotatoms in the carbon backbone but also probably implies an increase in the graphitization degree of the carbon backbone. This is probably responsible for a higher electronic conductivity and thus for an improved charge transfer. This strongly enhances the reaction kinetics and thus the half-wave potential is shifted towards higher electrode potentials for ZTC-300-TU and ZTC-400-TU catalysts in comparison with ZTC-300 and ZTC-400 materials. For these two materials, the chemical composition, the amounts of sulphur and nitrogen atoms as well as their speciation are rather similar as shown by XPS measurements. For these two materials, half-wave potentials of 0.79 and 0.81 V vs. RHE were measured, which indicates that chemical functionalization allows to overcome low structural and textural quality in ZTC materials. The ORR activity of ZTC-800 is very similar to the one of ZTC-800-TU material (for both a halfwave potential of 0.82 V vs. RHE), probably in reason of the low amount of S and N atoms inserted in the carbon backbone (see XPS results). Moreover, the second step (enabling S and N insertion in the carbon backbone) is performed at a temperature lower than 800°C (i.e. 750 °C). The graphitization degree of the ZTC backbone is in this case is thus not affected by the second heat treatment. Thus the electronic conductivity of ZTC-800-TU and ZTC-800 samples must be close from each other. The activity of N,S functionalized ZTC materials is higher to that observed for the N,S-doped reduced graphene oxide sample (rGO-TU) and close to the activity of a reference Pt/catalyst (half-wave potential of 0.85 V vs. RHE). This catalyst is also one of the most efficient N and/or S-doped carbon-based catalysts reported to date (see Table S6). This highlights the interest in using ZTC-based catalysts as ORR catalysts, with respect to doped graphene-based samples, which suffer from sheet stacking. [46,47] From RRDE experiments complementary information on the selectivity of the reduction process was obtained. The percentage of peroxide species formed during ORR as well as the number of electrons exchanged per oxygen molecule were determined from measured disk and ring currents (Figure 11). The hydroxyl selectivity of the reduction process is greatly improved after the insertion of nitrogen and sulfur atoms into the carbon backbone. For all N,S-doped ZTC catalysts, the amount of peroxide produced during the reduction process remains lower than 20%, and even below 12% for ZTC-800-TU in the whole potential range. This indicates that dual doping with nitrogen and sulfur atoms plays a critical role in directing the oxygen reduction process toward the four-electron pathway. Highest hydroxyl selectivity was observed for ZTC-800-TU, which might be associated to the higher number of graphitic N atoms (see XPS results). Finally, in this latter sample it is mostly probable that charge delocalization allows tuning the chemisorption mode of the O2 molecule, favoring the side-on adsorption mode and thus facilitating the four-electron pathway. [48] It is worth to note that the number of electrons exchanged per oxygen molecule and thus the hydroxyl selectivity of ORR is higher for ZTC-800-TU than for the graphene-based rGO-TU sample.

Conclusion

The synthesis of ZTCs using glycerol as carbon source was studied using synthesis temperatures from 300 to 800 °C. The textural and chemical properties were importantly impacted by the synthesis temperature. At 300 °C carbon materials without microporous long-range order were achieved featuring a flexible carbon network. At 400 °C high quality ZTCs featuring structured microporosity and high sp 2 carbon hybridization of the skeleton could be obtained. Materials synthesized at 800 °C present turbostratic carbon on the external surface of the ZTC. High ORR activity was observed even for unfunctionalized ZTC synthesized at 800 °C, yet with low selectivity towards the fourelectron process. Nitrogen and sulfur functionalization of ZTCs was achieved through thiourea treatment. The amount of S and N functionalization was evidenced to depend on the oxygen content of the ZTC. ORR activities were substantially improved after functionalization even for the material synthetized at 300 °C. High quality N,S functionalized ZTCs derived from glycerol allowed for achieving superior ORR activity and hydroxyl selectivity in comparison to functional graphene-based materials. To complete this study, stability tests have to be performed and an in-depth analysis of structural changes affecting both the carbon backbone and the speciation of heteroatoms under working conditions should be performed.

Experimental Section Materials

Glycerol (Sigma Aldrich, >99%), hydrofluoric acid (49% in water, Fisher Scientific), sodium bicarbonate (Sigma Aldrich, 99%), boric acid (Sigma Aldrich, 99%), nitrogen (99.995%, Air Liquide), thiourea (Sigma Aldrich, 99%) were purchased and used as received.

Synthesis of ZTC using glycerol

A similar reactor as previously described was used for the synthesis of ZTCs from glycerol. [49] 1 g of the zeolite powder was introduced into a cylindrical quartz reactor containing a frit and activated at 150 °C for 1 hour under a nitrogen flow of 150 mL min -1

. The oven temperature was then raised to the target temperature (300 -800 °C) and kept at this temperature for 1 hour. Glycerol feeding was started through a constant injection of 0.033 cm 3 min -1 to the nitrogen flow. After 4 h of feeding glycerol injection was ceased and the temperature was raised to 900 °C for 2 hours under nitrogen flow and then cooled down at room temperature. The recovered materials are named hybrid materials. In order to remove the zeolite template 0.7 g of the hybrid materials was placed in a Teflon beaker and 7 mL of HF were added and kept under stirring for 4 h. Thereafter 42 mL of a saturated aqueous H3BO3 solution was added. The mixture was then neutralized through adding 42 mL of a saturated aqueous NaHCO3 solution. After 1 hour of stirring at room temperature, the carbon materials was recovered by filtration using a polymer filter and washed using hot distilled water. After drying at 80 °C overnight, the final carbons were achieved. The samples were labelled ZTC-X where X corresponds to the synthesis temperature.

Thiourea treatment

The synthesis of N and S dual doped ZTC was performed by using thermally treated ZTC powders and thiourea as precursors. In a typical synthesis, 100 mg of ZTC powder was dispersed into 100 mL of ultrapure water. Then 1.6 g of thiourea are dissolved in the solution. The solution is homogenized by magnetic stirring for 30 minutes. Water is evaporated at 80 °C overnight before the powder is recovered and thermally treated at 700 °C for 2 h under a nitrogen flux (100 mL min -1

). The furnace is then cooled down at room temperature before the catalytic powder is recovered. The catalysts were labelled ZTC-X-TU where X corresponds to the synthesis temperature of ZTC material.

Characterization

The structural packing density (SPD) was determined via thermogravimetric analysis (TGA) using a SDT Q600 Waters device under air with a flow of 100 mL min -1

. Sample was heated to 900 °C with a ramp of 10 °C min -1

. The mean temperature of combustion was calculated from differential temperature analysis and corresponds to the value for which the differential temperature analysis signal reached a maximum. The SPD corresponds to the mass of carbonaceous species per gram of zeolite (

). [25] Nitrogen physisorption isotherms were measured on a Micromeritics 3Flex at 77 K. Approximately 50 mg of sample were outgassed at 300 °C for 12 h before the measurement. Microporous volumes were calculated through the t-plot method. The thickness of the nitrogen layer was calculated using a reference isotherm of (i) non-porous silica (used for the t-plot of zeolites) [50] and (ii) graphene (used for the t-plot of carbons). [49] The apparent density of hybrid compounds (ρApp) is a function of the SPD and of the occupied zeolite micropore volume and was calculated using Eq. ( 1) and is given as the ratio between the mass of carbonaceous species and its volume in the hybrid material (gC cm 3

).

(

) 1
where Vmicroi corresponds to the micropore volume of the zeolite template and Vmicrof to the micropore volume of the hybrid material. X-Ray powder diffraction (XRD) patterns were recorded on a PANalytical Empyrean X-Ray diffractometer using CuKα radiation (1.54059 Å) in the range of 5 -50 2θ. Elemental analysis was performed using a Flash EA 1112/Flash 2000 Thermo instrument. Scanning Electron Microscopy (SEM) Transmission Electron Microscopy (TEM) was achieved on a JEOL JSM-790CF microscope and JEOL 2100 instrument, respectively. Raman spectra were obtained using 532 nm excitation wavelength on a LabRam HR 800-UV Horiba Jobin Yvon confocal microscope. XPS analyses were carried out with a Kratos Axis Ultra DLD spectrometer using a monochromatic Al Kα source (1486.6 eV) operating at 180W (12 mA, 15 kV).

Electrochemical measurements

Electrochemical experiments were carried out using a home-made three electrodes Teflon cell at room temperature. A commercial hydrogen reference electrode (purchased by Gaskatel) and a glassy carbon slab were respectively used as reference and counter electrodes. The catalytic ink was performed by mixing 10 mg of catalytic powder, 750 µL of ultra-pure water, 250 µL of isopropanol and 100 µL of Nafion solution (5wt.% in a solution of aliphatic alcohols). The dispersion was finally dispersed by sonication to obtain a homogeneous solution. All experiments were carried out using a rotating ring disk electrode (Pine). All electrodes were connected to a biologic SP-300 potentiostat. The electrolyte was a 1 mol L -1 NaOH solution saturated with nitrogen or dioxygen. The working electrode was a glassy carbon disk (geometric surface area of 0.196 cm 2 ). For each experiment 7.5 µL of catalytic ink was deposited onto the glassy carbon disk and allowed to dry under nitrogen. Linear sweep voltammograms were recorded using the disk electrode at 1600 rpm by applying a scan rate of 5 mV s -1 to be under steady-state conditions. For all experiments the platinum ring was polarized at 1.2 V vs. RHE. The geometric surface of the ring is 0.11 cm 2 .

From these experiments, the fraction of water formed (p(H20)) as well as the number of electrons (n) exchanged per oxygen molecule during the reduction process can be calculated according to equations ( 2) and ( 3): [51,52] (

(3) Where N is the collection efficiency (21.8%), ID and Ir correspond to the intensities measured with disk and ring currents, respectively.

Figure 1 .

 1 Figure 1. Evolution of SPD (a) and mean temperature of combustion (b) as a function of the synthesis temperature.

Figure 2 .

 2 Figure 2. Nitrogen physisorption isotherms at 77 K (a) and apparent density as a function of the synthesis temperature (b). Color code: template USY zeolite (black hollow symbols), 300 °C (black symbols), 400 °C (red symbols), 500 °C (blue symbols), 700 °C (green symbols) and 800 °C (purple symbols).

Figure 3 .

 3 Figure 3. Nitrogen physisorption isotherms at 77 K (a). Evolution of the micropore volume as a function of the synthesis time (b). X-ray diffraction patterns of materials achieved at different temperature (c). Color code: 300 °C (black), 400 °C (red), 500 °C (blue), 700 °C (green) and 800 °C (purple).

Figure 4 .

 4 Figure 4. SEM images of ZTCs achieved at different synthesis temperatures.

Figure 5 .

 5 Figure 5. TEM images of ZTC-300 (scale bars from right to left: 200, 100 and 20 nm), ZTC-400 (scale bars from right to left: 500, 50 and 20 nm) and ZTC-800 (scale bars from right to left: 500, 100 and 10 nm).

Figure 6 .

 6 Figure 6. Mean temperature of combustion of ZTCs synthetized at different temperatures.

Figure 7 .

 7 Figure 7. Atomic C/O (a) and C/H ratio (b) of ZTCs as a function of the synthesis temperature.

Figure 9 .

 9 Figure 9. N1s (a) and S2p (b) XPS spectra of ZTC-300-TU, ZTC-400-TU, ZTC-800-TU.

Figure 10 .

 10 Figure 10. a) RRDE polarization curves recorded with all ZTC-based catalysts and Pt/C reference catalyst. b) Half-wave potential measured on polarization curves recorded in Figure a. The half-wave potential obtained with rGO-TU catalyst is shown for comparison.

Figure 11 .

 11 Figure 11. Number of electrons transferred per oxygen molecule (top) as well as fraction of peroxide formed during the reduction process (bottom). Calculations were done according to equations 2 and 3 using results shown in Figure 10a. Results obtained with Pt/C and rGO-TU materials are shown for comparison.

Acknowledgements

Authors acknowledge financial support from the European Union (ERDF) and "Région Nouvelle Aquitaine" and from the CAPES-COFECUB program (#88881.370977/2019-01 and #45020TE). The authors acknowledge the European Union (ERDF) and Région Nouvelle Aquitaine for the funding of MABATRI and PLATABAT projects (reference 2021-16148810 and reference 2022-23222510).

Entry for the Table of Contents

Bio-sourced synthesis of Zeolite-Templated Carbons (ZTCs) was achieved through the use of glycerol as carbon source. Oxygen content depends on synthesis temperature and importantly impacts the post-synthetic functionalization with N and S atoms. N,Sfunctionalized ZTCs allow for achieving conspicuous activities and selectivity towards the four-electron process.