Patricia García-Muñoz
email: patricia.gmunoz@upm.es

Paul-Henri Henri Allé

Calogera Bertoloni

Alvaro Torres

María Ulagares De La Orden

Joaquín Martínez Urreaga

Marie-Antoinette Dziurla

Fernando Fresno

Didier Robert
email: didier.robert@univ-lorraine.fr

Nicolas Keller
email: nkeller@unistra.fr

Photocatalytic degradation of polystyrene nanoplastics in water. A methodological study

Keywords: Nanoplastics, TiO2 photocatalysis, characterization tools, total organic carbon, methodology, water treatment

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

Introduction

Nowadays, over 300 million tons of plastic are produced worldwide annually, with ca. 13 million tons being released into rivers and oceans, and it is predicted that 250 million tons will have accumulated by 2025 [1]. Release and accumulation of plastic wastes to the environment is consequently of prime importance. Small plastics particulates enter surface water bodies and sea water mainly through a wide span of complex transport routes and via the fragmentation of larger debris down to the micro and nano size, that is defined as the environment-driven breakdown of larger plastic items by physical, chemical and biological processes [START_REF] Law | Plastics in the Marine Environment[END_REF][START_REF] Galgani | Marine litter within the European Marine Strategy Framework Directive[END_REF][START_REF] González-Pleiter | Secondary nanoplastics released from a biodegradable microplastic severely impact freshwater environments[END_REF].

The microplastics, mostly categorized as anthropogenic, manufactured, or processed items with particle size smaller than 5 mm and larger than 1 µm, has been the most studied size category of plastic debris for several decades [START_REF] Wright | The physical impacts of microplastics on marine organisms: A review[END_REF]. However, the terminology remains ambiguous and conflicting regarding the size classes. There is no standardized definition of the size below which the term microplastic is used [START_REF] Hartmann | Are We Speaking the Same Language? Recommendations for a Definition and Categorization Framework for Plastic Debris[END_REF], and microplastics (primary-or secondary-type) were defined also as being within the 1 to 1000 µm range.

More recently, focus has been made on nanoplastics that became a proper research field as such [START_REF]Questions on nanoplastic[END_REF], regardless they are secondarily produced through the further fragmentation of microplastics, or directly released as primary items by human activities with the increasing incorporation of manufactured nanoplastics within products from the daylife. Although no universal definition of the term 'nanoplastics' has been standardised yet, they are usually defined as items with particle sizes smaller than 1 µm, mostly due to the radical change in behavior of particles at the colloidal scale [START_REF] Hartmann | Are We Speaking the Same Language? Recommendations for a Definition and Categorization Framework for Plastic Debris[END_REF][START_REF] Gigault | Current opinion: What is a nanoplastic?[END_REF][START_REF]ISO/TT 21960 Plastics Environmental Aspects State of Knowledge and Methodologies[END_REF][START_REF] Braun | Status Report: Analysis of Microplastics Sampling, Preparation andDetection Methods within the Scope of the Bmbf Research Focus Plastics inthe Environment: Sources[END_REF].

The release of micro-and nano-sized plastic particulate waste in marine and (inland) freshwater bodies has become a major concern [START_REF] Schirinzi | Cytotoxic effects of commonly used nanomaterials and microplastics on cerebral and epithelial human cells[END_REF][START_REF] Kögel | Micro-and nanoplastic toxicity on aquatic life: Determining factors[END_REF]. They are considered as environmental contaminants of emerging concern (CECs), namely pollutants that have been detected in the environment and that may cause adverse ecological or human health impacts through the alteration of the quality of the ecosystems and of the entire food chain, while being at a given time outside any official regulation by authorities. They add to the long (and moving) list of substances that are (were) collectively falling within this definition, compounds such as (phyto)pharmaceuticals or engineered nanomaterials being among the most recent ones.

Wastewater treatment plants, as the receiver of discharged wastewaters from industrial, commercial and domestic sources, as well as less frequently surface run-off, are not suitably designed for the removal of microplastics, and a fortiori nanoplastics. Their elimination rate in the treatment plants is nowadays not high enough for completely preventing their entry into the natural aqueous ecosystems, and they can circumvent to a certain extent all treatment stages of municipal plants [START_REF] Sun | Microplastics in wastewater treatment plants: Detection, occurrence and removal[END_REF][START_REF] Ngo | Pathway, classification and removal efficiency of microplastics in wastewater treatment plants[END_REF][START_REF] Salvador Cesa | Synthetic fibers as microplastics in the marine environment: A review from textile perspective with a focus on domestic washings[END_REF]. Investigating remediation strategies aiming at reducing or removing MPs/NPs in water is however a very recent research field, with e.g. the first studies for the removal of nanoplastics being published less than 5 years ago [START_REF] Karimi Estahbanati | Treatment processes for microplastics and nanoplastics in waters: State-of-the-art review[END_REF].

Most of the chemical, biological and physical methods studied for removing micro and nanoplastics from water (e.g. dynamic membranes, membrane bioreactors, classical coagulation, electro-coagulation or biological degradation), have to deal with intrinsic drawbacks that limit their viability. The non-destructive nature of many of them is central, as they operate by transferring the plastic targets onto adsorbents or filters, and e.g. coagulation/ flocculation or electrocoagulation processes form large volumes of sludge highly concentrated in plastics [START_REF] Padervand | Removal of microplastics from the environment. A review[END_REF]. By contrast, biodegradation processes are highly dependent on the microbial species and the soil environment, and require more stringent storage conditions [START_REF] Inkinen | From Lactic Acid to Poly(lactic acid) (PLA): Characterization and Analysis of PLA and Its Precursors[END_REF].

In the wake of the development of advanced oxidation processes (AOPs) as high-prospect remediation tools to degrade dissolved pollutants in water, their application to the destructive removal of microplastics and nanoplastics appeared as an emerging research field. Among them, precursor works were conducted on the use of heterogeneous photocatalysis in water for the degradation and mineralization of high-and low-density polyethylene microplastics [START_REF] Ariza-Tarazona | New strategy for microplastic degradation: Green photocatalysis using a protein-based porous N-TiO2 semiconductor[END_REF][20][START_REF] Tofa | Visible light photocatalytic degradation of microplastic residues with zinc oxide nanorods[END_REF][START_REF] Tofa | Enhanced visible light photodegradationof microplastic fragments with plasmonic platinum/zinc oxide nanorod photocatalysts[END_REF] as well as polystyrene and poly(methyl methacrylate) nanoplastics [START_REF] Allé | Efficient photocatalytic mineralization of polymethylmethacrylate and polystyrene nanoplastics by TiO2/β-SiC alveolar foams[END_REF][START_REF] Domínguez-Jaimes | Degradation of primary nanoplastics by photocatalysis using different anodized TiO2 structures[END_REF].

In regards of the degradation of refractory industrial pollutants or micropollutants dissolved in water, the assessment of the efficiency of a remediation technique relies on the implementation of protocols well-established since decades. They consist, a minima, in the monitoring as a function of the treatment time of the concentration of the targeted substrate and of the organic carbon load, through the implementation of off-line techniques such as UV-Vis spectrophotometry, liquid phase chromatography or total organic carbon analysis. To date, those protocols are not adaptable yet for nanoplastics, first because of the non-dissolved, solid nature of the nanoplastic samples to be degraded, that in addition show dispersity in size and properties. Secondly the protocols have to deal with the complexity of the water matrix during the AOP treatment, that consists of a complex mixture of solid plastic particles with different sizes/shapes and of a wide span of organic by-products/intermediates with different functional groups resulting from the polymer oxidation. Both aspects are strongly complexifying the methodology to follow for assessing the efficiency of photocatalysis as degradation process, and for unravelling the degradation mechanisms.

The main mass-based (destructive) and particle-based (non-destructive) methods used for the detection, identification, and quantification of microplastics and (if applicable) of nanoplastics, were recently reviewed by Ivleva [START_REF] Ivleva | Chemical Analysis of Microplastics and Nanoplastics: Challenges, Advanced Methods, and Perspectives[END_REF]. Decreasing the particle size from the micro to the nano range gives rise to a huge methodological gap that constitutes an additional analytical challenge, as the transfer of the analytical tools used for microplastics to the realm of nanoplastics is not straightforward. Characterization of nanoplastics is facing critical challenges, associated with the insufficient sensitivity of the mass-based analysis tools that implies the need of a preconcentration/enrichment step due to the very low concentration of the suspensions, and with the limited particle size detection limit of most the non-destructive particle-based techniques, usually at about 300 nm. FTIR and SEM were recently used for characterising the photocatalytic degradation of microplastics in terms of chemical and morphology changes upon the treatment, but only weight loss analysis allowed for a quantitative follow up of the degradation efficiency and the assessment of the degradation kinetics [START_REF] Ariza-Tarazona | New strategy for microplastic degradation: Green photocatalysis using a protein-based porous N-TiO2 semiconductor[END_REF]20]. This paper aims therefore at studying the ability and relevance of different characterization and analytical tools to provide reliable qualitative or quantitative data allowing the monitoring of the efficiency of photocatalysis towards the removal of nanoplastics in water. To achieve this, photocatalytic degradation tests were implemented in both slurry and immobilized modes using calibrated polystyrene (PS) nanobeads of different characteristic sizes as model compounds.

Experimental

Materials

Commercial monodisperse suspensions of calibrated PS (C8H9)n nanobeads with high optical transparency and high monodispersity (CV < 3.8%) were supplied by Microparticles GmbH (Berlin, Germany). They consist in 5% (w/v) aqueous suspensions of spherical shape and low density (1.05 g/cm 3 ) polystyrene nanoparticles with a nominal mean size of 140 nm, 300 nm and 508 nm, and a standard deviation of 0.005 μm, 0.010 μm and 0.015 μm, respectively. PS NPs are labelled here as PS-140, PS-300 and PS-525 samples.

Commercially available powdery Aeroxide TiO2 P25 (hydrophilic fumed titanium oxide, >99.5%) was obtained from Evonik (Germany). Open-cell, self-bonded β-SiC alveolar foams with medium specific surface area manufactured by the SICAT SARL Company (Strasbourg, France) as cylinders (diameter 38 mm; length 40 mm) were used as support for conducting the degradation experiment in a recirculation mode. They were according to the Shape Memory Synthesis replica method, in which a pre-shaped polyurethane foam is transformed into the corresponding β-SiC carbide [26,[START_REF] Rico-Santacruz | Coating-free TiO2@β-SiC alveolar foams as a ready-to-use composite photocatalyst with tunable adsorption properties for water treatment[END_REF]. In this work, 8 pore per inch (PPI) β-SiC alveolar foams with an open porosity of about 0.91 and characterised by a mean cell size of 5400 +/-700 μm, a mean window size of 2300 +/-600 μm, and a mean bridge diameter of 575 +/-80 μm have been used.

In regards of the TiO2-P25/β-SiC foam photocatalysts, an homogeneous TiO2-P25 Aeroxide© coating was obtained at the foam surface by implementing a classical dip-coating technique followed by thermal annealing to obtain a high adhesion of the catalyst coating on the substrate. The β-SiC foam samples were calcined at 1000°C for 2 h to remove residual organic carbon, before being completely immersed for 5 min in an ethanolic TiO2-P25 suspension (10 g of TiO2-P25 in 200 mL) supplemented with 4 mL of titanium tetraisopropoxide (TTIP, Sigma-Aldrich, 97%). The TTIP was used as binder for the TiO2-P25 nanoparticles through sol-gel formation and allows the enhancement of the stability and the homogeneity of the coating, as previously optimized in Marien et al. [28]. The coated foams were dried at room temperature for 20 min with a fan for avoiding the clogging of the cells, and the dip-coating protocol was triplicated. The TiO2-P25/β-SiC foams were dried in an oven at 110°C overnight to evaporate residual organic compounds, and were further calcined under static air at 450°C for 2 h with a temperature ramp of 5°C.min -1 . The average wt. content of TiO2-P25 per foam was determined at 7.5 % by weight difference, which corresponded to 1.5 g of TiO2-P25 fixed on each β-SiC foam cylinder.

Photocatalytic reactor and analysis

Independently of the test mode, tests were performed with a TOC load higher than that in wastewater and ambient water, in order to compare a wide span of analytical tools and characterization techniques.

Batch slurry reaction mode. The photocatalytic degradation experiments were carried out in a batch beaker-type glass reactor at atmospheric pressure, room temperature (20°C), natural pH, and using O2 from the atmosphere as oxidant, air being pumped into the reactor at 150 mL/min for preventing any oxygen deficiency in the reaction media. The experiments have been performed under stirring at 600 rpm with a reaction volume of 250 mL at a 1 g/L TiO2-P25 catalyst load, and with a concentration of PS-140, PS-300 and PS-525 NPs that corresponds to an initial TOC load of about 100 mg/L. The suspension was stirred for 30 min in the dark in order to establish the dark adsorption/desorption equilibrium before turning on the light. The suspension was further exposed to a 60 W/m 2 irradiance UV-A light centred at 365 nm, provided by Philips 24 W/10/4P lamps.

The samples containing the polymer nanobeads and the titania particles were centrifuged in order to separate the aqueous suspension with the suspended polymer from the powdery catalyst. The supernatant was recovered for DLS analysis while the powder was used for FTIR and SEM analyses after drying overnight at 100°C.

Immobilized reaction mode.

The photocatalytic experiments in an immobilised reaction mode were carried out in a tubular quartz photoreactor operating in a recirculation mode, described in detail in a previous work [START_REF] Allé | Efficient photocatalytic mineralization of polymethylmethacrylate and polystyrene nanoplastics by TiO2/β-SiC alveolar foams[END_REF]. The tubular reactor was filled with three cylinders of TiO2-P25/β-SiC foams and surrounded by 4 UV-A lamps (Philips T5 15W 10 Actinic BL) to provide a 110 W/m 2 irradiance UV-A light centred at 365 nm. Tests were mostly performed with a TOC load of 20 mg/L. A volume of 600 ml of aqueous nanopolymer suspension to be treated was recirculating within the photoreactor for 30 min in the dark to allow the establishment of the equilibrium of the system, before the irradiation was switched on. During the experiments, 10 mL of suspension were taken at regular time-intervals for analysis.

Characterization techniques

Fourier Transform Infrared (FTIR) spectra of the materials were recorded in a Nicolet iS10 spectrometer (Thermo Scientific, Waltham, MA, USA) equipped with a Smart iTR Attenuated Total Reflectance (ATR) accessory controlled by OMNIC Software Ver. 9.9.473. The resolution used was 4 cm -1 , and 16 spectra were recorded for each sample. In the case of PS samples, all the spectra were normalized at 2850 cm -1 , a commonly used internal standard for PS that corresponds to the vibration bands of -CH2- [START_REF] Mylläri | A comparison of rheology and FTIR in the study of polypropylene and polystyrene photodegradation[END_REF]. The carbonyl (CI) and peroxyl (PI) indexes were calculated according to the following equations [START_REF] Mylläri | A comparison of rheology and FTIR in the study of polypropylene and polystyrene photodegradation[END_REF][START_REF] Kuzina | Photo-oxidation of polymers-2. Photo-chain reaction of peroxide radicals in polystyrene[END_REF][START_REF] Yousif | Photodegradation and photostabilization of polymers, especially polystyrene: review[END_REF][START_REF] Domínguez-Jaimes | Degradation of primary nanoplastics by photocatalysis using different anodized TiO2 structures[END_REF]:

CI=h(1750)/hstd (1) PI=h1150/hstd (2)
where h refers to the corrected height of the peak at 1750 cm -1 and 1150 cm -1 associated to the carbonyl and the peroxyl groups, respectively, and hstd refers to the corrected height of the peak at the standard wavenumber for PS, ie. 2850 cm -1 .

Dynamic Light Scattering (DLS) measurements were performed on a Malvern Zetasizer (Nano-ZS Model) with a He-Ne laser at 632 nm wavelength, at a scattering angle of 173° (back angle).

Field emission scanning electron microscopy (FE-SEM) characterization was performed by

means of a JEOL JSM-7900F microscope using selective secondary electrons detection. An accelerating voltage of 3.0 kV was used in order to obtain high-resolution images.

Pyrolysis coupled with gas chromatography/mass spectrometry (py-GC/MS) was performed using a 6150 pyroprobe (CDS Analytical, Quad Service, France), combined online with a Trace 1300 gas chromatograph and ISQ 7000 quadrupole mass spectrometer (Thermoscientific, France). Pyrolysis was performed in the case of PS-140 using a sample volume of 10 µL introduced in a quartz capillary (6201-3004, CDS Analytical, Quad Service, France) in the pyroprobe. Two pyrolysis temperatures were applied, namely 500°C/30 sec for the PS quantification in combination with GC analysis in the split mode, and 250°C/15 sec for the analysis of PS photo-oxidation products, in combination with GC analysis in the splitless mode.

GC-MS analysis was performed using a TG-5SilMS column (30m length X 0. The quantification of PS in water was assessed by measuring the area of the main pyrolysis products, namely styrene, styrene dimer and styrene trimer. Their detection was performed with m/z 78 and 104 for styrene, m/z 91 and 208 for styrene dimer, and m/z 91 and 312 for styrene trimer, with bold values corresponding to the respective fragment ion used for quantification [START_REF] Fabbri | Off-line analytical pyrolysis GC-MS to study the accumulation of polystyrene microparticles in exposed mussels[END_REF]. PS quantification was performed using two standard curves without internal standard, the first one with PS concentrations ranging between 0 and 4 mg/L TOC, analysed with a split ratio 5:1, and the second one with PS concentrations between 4 and 20 mg/L TOC, analysed with a split ratio 50:1.

Total Organic Carbon (TOC) measurements were performed using a Shimadzu TOC-L analyzer to determine the organic carbon load of the aqueous suspensions as a function of the treatment time. Analysis was realised without any prior filtration using suspension volumes of 10 mL sampled at specific time intervals.

Liquid phase UV-Vis spectrophotometry was performed using a double beam CARY 100 Scan UV-visible spectrophotometer (Varian).

Transmission electron microscopy (TEM) was performed using a JEOL 2100 F with a point resolution of 0.2 nm. A drop of the aqueous suspension was deposited onto a copper grid covered by a holey carbon membrane. Grids were hydrophilized prior to any observation through Plasma cleaning for 30 s (Gatan solarus 950).

Results and discussion

Batch slurry reaction mode

The implementation of degradation tests in the batch slurry mode and the necessary monitoring of suited degradation and mineralization indicators are facing a significant challenge in comparison to the analogous tests targeting dissolved pollutants. Indeed, the sub-micronic filters classically used during the sampling for removing the powdery photocatalysts from the aqueous reaction media in order not to distort the analysis (eg. for UV-vis spectrophotometry) and additionally for not damaging sensitive analytics (eg. for TOC analyser, high-performance liquid chromatography, ionic or size-exclusion chromatography), cannot be applied in the case of nanoplastics. This results from the identical characteristic sizes of both the plastic nanobeads and the TiO2 nanopowders, that do not allow for a selective filtration of the photocatalyst.

Independently of the porosity characteristics of the filters, the two kinds of objects are irremediably filtered or released.

These considerations require to turn to unusual characterization tools able to operate reliably and safely, even in the presence of the TiO2 nanoparticles, whether their concentration is significant or residual. Concretely, the ability of IR, DLS and SEM characterization techniques to get rid of separation issues and to be used efficiently for assessing the efficiency of photocatalysis in the degradation of NPs in suspension were here challenged and probed.

First, FE-SEM study allowed for the determination of the impact of the photocatalytic treatment on the morphological features of the PS-300 nanoparticles. Figure 1 shows representative SEM images of the PS-300 nanoparticles at different stages of the process, from which a statistical size analysis allowed size distribution histograms to be derived (Figure 2). Figures 1A and2A show that the PS-300 sample consisted in spherical PS nanoparticles with a relatively narrow size distribution centred at ca. 325 nm, in good agreement with the manufacturer data sheet. Once in contact with TiO2 in dark conditions, it is apparent from SEM images, as exemplified in Figure 1B, that adsorption occurred, with no change in the size distribution (Figure 2B). We have to stress that stricto sensu the term adsorption is not welladapted, as the nanobeads are in the solid phase, so they are not adsorbing on the TiO2 surface, but rather directly contacting with the surface. Upon UV-A irradiation, a reduction in the mean PS nanoparticle size together with a slight broadening of the size distribution was observed, the effect being all the more pronounced that the irradiation time increased, to reach a mean size of 270 nm, while PS nanoparticles as small as 200 nm were observed (Figures 1C-D and2C-D).

Here, it is worth noticing the difficulty of measuring PS particle sizes once they are covered by TiO2, what resulted in a lower number of measurable particles and hence a lower quality of the histograms as observed in Figure 2. However, the size reduction observed was a fingerprint of the degradation of the PS nanoparticles upon the irradiation of the TiO2 dispersion.

Independently of the reaction time, the images evidenced that all the PS nanoparticles observed were surrounded by smaller size TiO2 nanoparticles, no isolated PS nanoparticles being observed. This multi-scale aggregation or attachment phenomena was comparable to that observed by TEM during the inactivation of bacteria by photocatalysis using TiO2 nanopowders in water, for which the crucial role of the contact or adhesion between the TiO2 particles and the bacterial cells in suspension was demonstrated [START_REF] Foster | Photocatalytic disinfection using titanium dioxide: spectrum and mechanism of antimicrobial activity[END_REF][START_REF] Gumy | Catalytic activity of commercial of TiO2 powders for the abatement of the bacteria (E. coli) under solar simulated light: Influence of the isoelectric point[END_REF]. Similarly to the bactericidal action of TiO2 dispersions, the enhanced contact between the PS nanobead targets and the TiO2 photocatalyst grains in the slurry mode favours a multiple attack of HOx° radicals simultaneously on the whole external surface of the PS nanoparticles.

In addition to size effects, SEM images shown in Figure 3 visualise that some of the PS nanoparticles have lost their initial morphology upon the photocatalytic degradation. Indeed, the images reveal the presence of PS nanoparticles with a faceted aspect that differs from the (isotropic) round shape of the starting PS substrate. The photocatalytic degradation of the polystyrene nanobeads was also monitored by FTIR analysis in the case of the PS-140 sample (Figure 5). In addition to the peak at 2850 cm -1 assigned to the C-H stretching of -CH2-groups and used as internal standard for PS, the qualitative analysis of FTIR spectra pointed out several peaks of interest corresponding to characteristic vibration modes [START_REF] Mylläri | A comparison of rheology and FTIR in the study of polypropylene and polystyrene photodegradation[END_REF], namely:

• at 3025 cm -1 , that is the characteristic peak of the stretching mode of aromatic C-H bonds in the polystyrene.

• at 1750 cm -1 , that reflects the presence of carbonyl groups associated with the degradation of the PS nanoparticles.

• at 1600 cm -1 and 1492 cm -1 , and 1450 cm -1 , that can be assigned to C-C stretching modes of aromatic C-C bonds and to δ (CH2) bending, respectively.

• at 1150 cm -1 , that can be associated with C-O-O stretching vibrations of peroxyl groups.

After contacting under stirring the initial PS nanoparticles with TiO2 in the dark, the characteristic polystyrene peaks were observed, confirming the presence of PS nanoparticles within the TiO2 nanoparticle aggregates prior to the irradiation. Figure 6 shows the evolution with the reaction time of both the carbonyl (CI) and peroxyl (PI) indices derived from FTIR spectra. It evidenced the increase of both indices with the reaction time, although to a lesser extent and with a lag period of about 1 h for the PI evolution in comparison to the substantial CI evolution.

Although the evolution of both PI and CI values with the treatment time remains semiquantitative, this points out that a certain oxidation degree was achieved during the reaction and provides information on the degradation pathway. This reflects the efficiency of the photocatalytic process and can be considered as a fingerprint of the degradation of the PS nanobeads over the time.

Immobilized mode in recirculation

The major advantage of the use of supported (immobilized) catalysts compared to suspensions lies in the elimination of the filtration step of the aqueous sample before analysis.

In this case, the suspensions to be analyzed contain the polymer nanoparticles and their degradation by-products without catalyst particles. It is therefore possible to use analytical techniques (UV-Visible spectroscopy, TOC analysis, py-GC/MS, etc.) which cannot be implemented with suspensions containing the photocatalyst powders.

Direct TOC analysis

Total Organic Carbon (TOC) measurement is not directly related to the concentration of polymer in the aqueous suspensions. However, it can demonstrate the transformation of the organic carbon contained in the polymeric nanomaterials into CO2 under the action of UV

Electron microscopy characterizations and morphological aspects

SEM and TEM are intuitively potentially interesting techniques for a visual assessment of the efficiency of the photocatalytic treatment. However, only TEM might maybe provide a quantitative evaluation through a statistical analysis of the size reduction of the nanoparticles, as classical FE-SEM suffers from a lack of resolution at the nanoscale.

Both SEM and TEM images shown in Figure 8 and Figure 9, respectively, do not evidence a clear decrease in the mean diameter of the nanoparticles, independently of the technique resolution, the statistical analysis of diameters being not significative. However Figure 8C shows the presence of polymeric debris with a large variety of sizes and shapes, that probably resulted from the loss of the spherical shape during the oxidative attack by HO°x radicals. This effect was confirmed by the TEM study. Figure 9 reveals the shape transformation of the starting spherical, well-defined PS-140 nanoparticles. Figure 9D shows to which extent the morphology of the nanoparticles was affected by the multiple attacks of the radicals, while the release of polymeric materials from the spheres was clearly evidenced in Figure 9C. This effect was observed without the beads completely losing their spherical morphology, so that a statistical analysis of the particle size was not relevant for expressing the degradation efficiency. This behaviour differs from Nabi et al. who observed a progressive reduction in size of PS nano and microbeads during the photocatalytic treatment on TiO2 films [START_REF] Nabi | Complete Photocatalytic Mineralization of Microplastic on TiO2 Nanoparticle Film[END_REF].

Pyrolysis coupled with gas chromatography/mass spectrometry (py-GC/MS)

Pyrolysis coupled with gas chromatography/mass spectroscopy (Py-GC/MS) is a promising thermoanalytical method for the direct characterization and quantification of MPs and NPs in environmental samples through the analysis of their specific pyrolysis products [37][START_REF] La Nasa | A review on challenges and developments of analytical pyrolysis and other thermoanalytical techniques for the quali-quantitative determination of microplastics[END_REF][START_REF] Gigault | Nanoplastics are neither microplastics nor engineered nanoparticles[END_REF]. At high temperature, the major pyrolysis products of polystyrene are styrene (60-80%), as well as styrene dimer and trimer (<10%) [START_REF] Sullivan | Detection of trace sub-micron (nano) plastics in water samples using pyrolysis-gas chromatography time of flight mass spectrometry (PY-GCToF)[END_REF]. Although styrene is a non-specific product of the polystyrene degradation, this marker was used for quantifying polystyrene in the present work since polystyrene was the only polymer present in our experiments. Temperatures applied for quantifying polystyrene through pyrolysis vary from 500°C [START_REF] Fabbri | Off-line analytical pyrolysis GC-MS to study the accumulation of polystyrene microparticles in exposed mussels[END_REF] to 600°C [START_REF] Fischer | Microplastics analysis in environmental samplesrecent pyrolysis-gas chromatography-mass spectrometry method improvements to increase the reliability of mass-related data[END_REF][START_REF] Dehaut | Handbook of Microplastics in the Environment[END_REF]. In the present work, a temperature of 500°C was selected, with suited variation coefficients of 11 % and 8 % (n=3) for styrene and styrene trimer production, respectively (Supporting Information SI2). In terms of methodology, the analysis of the Py-GC/MS data after pyrolysis at 500°C requires the establishment of a calibration curve to quantitatively relate both styrene trimer and styrene MS area to the TOC quantity of the polystyrene NP. Calibration performed without internal standard within the 0-20 mg/L TOC range showed suited linear correlations for both styrene and styrene trimer areas versus TOC of the PS nanoparticles (Supporting Information SI3).

Figure 10 compares the TOC values during the photocatalytic degradation of PS-140 nanoparticles with those derived from the py-GC/MS measurements after pyrolysis at high temperature thanks to the standard curves (TOC-py). TOC values do not account only for the PS nanoparticles but for the overall carbon load of the aqueous suspension (including all kinds of reaction intermediates). However, the results revealed a good agreement between the (direct)

TOC values and the (indirect) TOC values derived from the py-GC/MS analyses, and that account only for the polystyrene macromolecules. This behaviour could be assigned to the (by far) faster mineralization kinetics of molecular reaction intermediates in comparison to the polystyrene macromolecular objects, independently of the polystyrene nature or morphology, namely as original nanobeads or as released polymeric fragments. It could not be ruled out that the starting PS nanobeads were first transformed throughout the photocatalytic attack into polystyrene of lower molecular weight, as they have the same py-GC/MS signature than the original PS substrate. HPLC measurements performed on filtered (0.22 µm pore-filters) suspensions evidenced the presence of solubilized benzoic acid at a concentration close to the quantification limit (0.1 mg/L, corresponding to only 0.07 mg/L TOC), while no benzaldehyde was detected, both products being amongst the PS photodegradation products [START_REF] Mailhot | Polystyrene photooxidation. 2. A pseudo wavelength effect[END_REF]. Low-temperature pyrolysis (i.e. 250°C for 15 sec) was applied to the aqueous suspensions sampled during the photocatalytic degradation experiments for identifying low molecular weight degradation products (Table 1, Figure 11). No styrene could be detected after pyrolysis at 250°C, indicating the absence of polystyrene depolymerisation at this temperature. While the analysis of the un-treated PS-140 nanoparticle suspensions did not reveal the presence of any degradation products, several volatile compounds were detected on the pyrograms, mainly after 24 h photocatalysis, namely acetic acid, benzoic acid, benzophenone and 1,2-ethanediol monobenzoate. Molecules such as acetic acid, benzoic acid and benzophenone have been been produced during the photocatalytic treatment, and released to the aqueous environment.

They might also remained trapped within the nanoparticles or adsorbed at on the polymer surface. As underlined by Reichel et al. [44], low temperature py-GCMS is useful for studying the capture of pollutants by nanoparticles, which is of main interest in ecotoxicological studies on solid particles such as soil, plastic nanoparticles present in water samples, or fine air-borne particles.

Analysis of the photocatalyst surface

The surface of the TiO2/β-SiC foam photocatalysts has been characterized by SEM as a function of the reaction time (Figure 10). First, the images did not allow any significant decrease in the mean size of the PS nanoparticles to be observed, as mean sizes of 136 nm ± 9 nm, 134 nm ± 13 nm and 130 nm ± 16 nm were determined after stirring in the dark, and for 2 h and 4 h of reaction, respectively. It is worth noting that intact nanobeads with no apparent morphological changes were observed in contact with the TiO2 surfaces independently of the reaction time, even when the remaining TOC concentration in the suspension accounted for only 20% of the initial TOC content (at 4 h, see experiment at 20 ppm of TOC in Figure 11). By contrast, Figure 10F did not evidence the presence of any residual nanobeads after 6 h of reaction when the TOC concentration in water was close to zero. This showed that the PS nanoparticles do not accumulate within the alveolar foam-based structures that do not act as long-term reservoirs, what would distort the analysis and the results.

Advantages and drawbacks

We clearly evidenced the difficulty in obtaining quantitative (and even semi-quantitative) data about the efficiency of photocatalysis for degrading and mineralizing PS nanobeads when the reaction operates in the batch slurry mode. At best, the results revealed the existence of a photocatalytic activity towards PS nanobeads, with no capacity of reliable quantification.

The immobilized reaction mode showed an unequivocal advantage vs. the slurry mode, and allowed the assessment of relevant, quantitative efficiency indicators. Finding the suited counterpart of the indicator 'concentration' used in the case of the degradation of dissolved pollutants is not an easy task. Indeed, TEM results revealed that the PS nanobeads can lose their nanobead morphology, in parts or entirely, while their chemical nature is probably not strongly altered yet, with the release to the water of polystyrene macromolecules. The logical corollary of this is that the parameter size or size distribution, although intuitively of prime importance to monitor as a function of processing time, is not very suitable as an indicator.

Py-GC/MS showed a promising interest in the case of nanoplastics, as it allows the polystyrene concentration to be followed with time. However, it does not provide any reliable information on the concentration of the nanobeads themselves cannot be followed. However, it should be underlined that the technique is prone to contaminations, due to incomplete sample pyrolysis and sample accumulation in the system without reaching the GC column [44]. Also, it requires frequent instrument and material cleaning for obtaining a high sensitivity and accuracy [START_REF] Ivleva | Chemical Analysis of Microplastics and Nanoplastics: Challenges, Advanced Methods, and Perspectives[END_REF]44]. The use of an internal standard such as styrene-d8 is a strategy that might allow the results to be improved [START_REF] Funck | Identification of microplastics in wastewater after cascade filtration using Pyrolysis-GC-MS[END_REF]. However Py-GC/MS does not provide any information to which extent the macromolecular weight was impacted or not during the treatment. Size-exclusion chromatography could be a suited tool to get information on the macromolecular weight and dispersity. However, this technique requires usually large amounts of polymers, what restricts its application to degradation experiments with high concentrations in PS nanobeads.

When the photocatalytic degradation tests can be implemented in the immobilised mode, it appears clear that TOC analysis remains the optimal off-line chemical technique to evaluate the performances of the removal process by quantifying the mineralization of the PS substrates, both in terms of mineralization yield and kinetics. In addition, it was worth noting that the TOC monitoring is apparently a technique that is not dependent on the nature or size of the polymer nanoparticles. Indeed, TOC analysis was also successfully used in Allé et al. for quantifying the mineralization of calibrated polymethylmethacrylate (PMMA) nanoparticles [START_REF] Allé | Efficient photocatalytic mineralization of polymethylmethacrylate and polystyrene nanoplastics by TiO2/β-SiC alveolar foams[END_REF].

TOC analysis vs. UV-Vis spectrophotometry. TOC analysis is destructive by nature and requires large sample volumes, what is not always suited to the experimental setup. UV-Vis spectrophotometry can be considered as an appropriate alternative to TOC analysis for monitoring the mineralization of PS nanobeads. Indeed, the absorbance peak maximum at low wavelength can be considered as a fingerprint of the PS structure, and was already successfully used for the quantitative assessment of the removal of PS nanoplastics through flocculation [START_REF] Chen | Sedimentation of nanoplastics from water with Ca/Al dual flocculants: Characterization, interface reaction, effects of pH and ion ratios[END_REF][START_REF] Chen | Phase transition of Mg/Al-flocs to Mg/Allayered double hydroxides during flocculation and polystyrene nanoplastics removal[END_REF]. The evolution of the TOC concentration as a function of the time under irradiation was derived from the UV-Vis absorbance curves, and could be monitored in an quantitative way by using a suited calibration curve built from the analysis of calibrated PS-140 suspensions with known TOC concentrations (Figure 11A). Indeed, regardless of whether the degradation test was efficient (at 20 mg/L) or not (at 40 mg/L), as depicted in Figure 11B, the TOC concentration profile indirectly obtained through UV-Vis spectrophotometry was in both cases in agreement with that directly obtained using TOC analyser.

UV-Vis spectrophotometry takes advantage of its non-destructive nature and does not consume the reaction media that can be fed back to the reactor after analysis. The method might therefore be adapted for conducting experiments with extended reaction time or with smallscale (low water volume) reactors. However, we have to stress that the analysis is subjected to non-negligible variability, as visible on the graphs. Secondly, the method remains highly TiO2 P25). Upon UV-A light irradiation, adsorbed oxygen and water molecules are reacting with the photogenerated electrons and holes, respectively, and give rise to superoxide and hydroxyl radicals, respectively, with subsequent interaction with the PS nanoparticle substrates. The photocatalytic reaction starts via the attack of polystyrene particle by OH° radical. (Eq. 1 and 5).

This attack reported in Eq. 3 and 7 generates carbon-centred radicals in the polymer structure [START_REF] Domínguez-Jaimes | Degradation of primary nanoplastics by photocatalysis using different anodized TiO2 structures[END_REF], and can be initiated on multiple carbon atoms, simultaneously [START_REF] Kuzina | Photo-oxidation of polymers-2. Photo-chain reaction of peroxide radicals in polystyrene[END_REF]. Then those radicals could react with the dissolved oxygen giving rise to peroxide molecules (Eq. 4 and 8). The third step might involve the production of hydroperoxide compounds when peroxide interacts with a molecule of water (Eq. 5 and 9). Further reactions (Eq. 6 and 10) can give rise to chain scission and formation of carbonyl or ketone compounds like those detected by FTIR. Those intermediates can be subjected to further, consecutive, stepwise degradation to CO2 and water.

It must be stressed that UV photons might also be directly absorbed by the polymeric chains to create excited states that generate chain scission, cross-linking reactions, and oxidation in the polymer [START_REF] Yousif | Photodegradation and photostabilization of polymers, especially polystyrene: review[END_REF][START_REF] Bottino | A study on chemical modifications, mechanical properties and surface photo-oxidation of films of polystyrene (PS) stabilised by hindered amines (HAS)[END_REF][START_REF] Kaczmarek | Photoinitiated degradation of polystyrene in the presence of low-molecular organic compounds[END_REF] Below can be seen two proposed degradation routes depending on the position of the initial OH° attack:

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Conclusion

We have shown that the way of implementing the photocatalytic tests for degrading calibrated nanoplastic nanobeads in water was strongly impacting the ability to monitor and quantify their degradation and mineralization. Testing in the immobilized mode showed an unequivocal advantage over the slurry mode and should be preferred to get rid of the restrictive presence of the nanoparticulate photocatalyst in the samples to be analysed. This resulted from the impossibility to efficiently, selectively filtrate the photocatalyst when operating in the slurry mode. At best, the results in the slurry mode revealed the existence of a photocatalytic activity towards PS nanobeads, with no capacity of reliable quantification.

The nanoplastic size distribution, whether obtained by indirect light-scattering or direct imaging techniques, was not found to be a relevant indicator of the photocatalytic efficiency, as the PS substrate was shown to lose its nanobead morphology during the photocatalytic attack, in parts or entirely, with the simultaneous release to the water of PS macromolecules.

Direct TOC analysis that accounts for the overall carbon load of the aqueous suspension (including the PS nanoparticles and all kinds of reaction intermediates) remains the optimal chemical technique to evaluate the performances of the photocatalytic treatment by quantifying the mineralization of the PS substrates, both in terms of yield and kinetics. The results revealed a good agreement between the TOC analysis and the (indirect) TOC values derived from py-GC/MS analyses, that account only for the PS macromolecules, regardless of their morphology and macromolecular weight. This was assigned to the faster mineralization macromolecular PS objects, whether in the form of nanobeads or released polymeric fragments.

UV-Vis spectrophotometry was reported to be a fast, non-destructive alternative technique to TOC analysis in the specific case of PS.

  25 mm internal diameter and 0.25 mm film thickness ; Thermoscientific, France). The analyte ionization was performed by electron ionization (70 eV), with the MS transfer line set at 280°C, and the ion source set at 250°C. The signal acquisition was performed in scan mode, with a scan time of 0.2 sec, in the 40-350 m/z and 40-250 m/z ranges, for the quantification of PS and the identification of the polymer photo-oxidation products, respectively.

Figure 1 .

 1 Figure 1. FE-SEM images of PS-300 nanoparticles: (A) the starting PS nanobeads, (B) after the

Figure 2 .

 2 Figure 2. Size distributions of the PS nanoparticles derived from the FE-SEM images: (A) the

Figure 3 .

 3 Figure 3. FE-SEM images of PS nanoparticles after irradiation in close contact with TiO2 for (A) 2

Figure 4 .Figure 5 .

 45 Figure 4. Evolution of the particle size distribution as a function of the process time for (A) PS-357

Figure 6 .

 6 Figure 6. Evolution with the reaction time of both carbonyl (CI) and peroxyl (PI) indices derived

  photocatalysis. Indeed, the residual concentration of organic carbon in the suspensions sums up

Figure 7 .

 7 Figure 7. a) Mineralization kinetic of PS-140 nm and PS-525 nm nanobeads on TiO2-P25/β-SiC

Figure 8 .

 8 Figure 8. FE-SEM images of (A) starting PS-140 sample and (B-C) the PS-140 sample after 48 h of

Figure 9 .

 9 Figure 9. TEM images of (A-B) starting PS-140 sample and (C-D) the PS-140 sample after 48 h of

Figure 10 .

 10 Figure 10. Comparison between the overall (direct) TOC values and the (indirect) TOC values

Figure 11 .

 11 Figure 11. Pyrogram recorded when applying low-temperature pyrolysis on the treated PS-140

Figure 10 .

 10 Figure 10. SEM images of the surface of the TiO2-P25/β-SiC foams as a function of the process stage. (A) clean (naked) surface, (B) after contacting with the PS-140 nanoparticles for 1 h in the dark, (C) after 2 h of reaction, (D-E) after 4 h of reaction and (F) after 6 h of reaction when full PS mineralization was

Table 1 .

 1 Molecules observed during the low-temperature pyrolysis applied on the aqueous samples of treated PS-140 nanoparticles.

	Reaction intermediate	Characteristic fragment ions (m/z) Reaction time (h)
	Acetic acid	43, 45, 60	24
	Benzoic acid	51, 77, 105, 122	24, 48, 72
	Benzophenone	51, 77, 105, 182	24, 48, 72
	1,2-ethanediol monobenzoate	51, 77, 105, 123	24

Acknowledgements

Patricia García-Muñoz thanks Escuela Superior de Ingenieros Industriales (UPM) for the project ETSII-UPM20-PU03. Technical assistance of Dr. F. Picó for FE-SEM characterization is gratefully acknowledged. The authors of this article thank Campus France for the grant awarded to Paul Henri Allé.

dependent on the UV-Vis signature of the polymer nanobeads targeted, as for instance the technique was inefficient on PMMA nanobeads. Reaction mechanism. The increase of both carbonyl and peroxyl indexes as a function of the time under irradiation, and the different reaction by-products observed through different analytical tools are in good agreement with the degradation pathway proposed in Domínguez-Jaimesa et al. [START_REF] Domínguez-Jaimes | Degradation of primary nanoplastics by photocatalysis using different anodized TiO2 structures[END_REF] and in Yousif et al. [START_REF] Yousif | Photodegradation and photostabilization of polymers, especially polystyrene: review[END_REF](Eqs. [START_REF] Galgani | Marine litter within the European Marine Strategy Framework Directive[END_REF][START_REF] González-Pleiter | Secondary nanoplastics released from a biodegradable microplastic severely impact freshwater environments[END_REF][START_REF] Wright | The physical impacts of microplastics on marine organisms: A review[END_REF][START_REF] Hartmann | Are We Speaking the Same Language? Recommendations for a Definition and Categorization Framework for Plastic Debris[END_REF][START_REF]Questions on nanoplastic[END_REF][START_REF] Gigault | Current opinion: What is a nanoplastic?[END_REF][START_REF]ISO/TT 21960 Plastics Environmental Aspects State of Knowledge and Methodologies[END_REF][START_REF] Braun | Status Report: Analysis of Microplastics Sampling, Preparation andDetection Methods within the Scope of the Bmbf Research Focus Plastics inthe Environment: Sources[END_REF][START_REF] Schirinzi | Cytotoxic effects of commonly used nanomaterials and microplastics on cerebral and epithelial human cells[END_REF][START_REF] Kögel | Micro-and nanoplastic toxicity on aquatic life: Determining factors[END_REF].

The photocatalytic degradation of PS was proposed to follow a typical degradation route observed for polymers in water, following a first surface contact with the catalyst surface (here