Andrea Carboni
email: andrea.carboni9@unibo.itorcid.org

Danielle L Slomberg

Amazigh Ouaksel

Lenka Brousset

Andrea Campos

Bernard Angeletti

Bahareh Zareeipolgardani

Gael Depres

Alain Thiéry

Jerome Rose

Laurent Charlet

Melanie Auffan

Matrix driven environmental fate and effects of silver nanowires during printed paper electronics end of life

Keywords: Nano-enabled products, printed paper electronics, mesocosms, silver nanomaterials, aquatic ecosystem

The release of engineered nanomaterials (ENMs) during manufacturing, use and disposal of nanoenabled products (NEPs) is a potential route of environmental nano-pollution. To date, there is limited knowledge on the ecological impact of ENMs incorporated in NEP matrices and how they compare to pristine nanoparticles. Here, we examined the fate and effect of silver nanowires (AgNWs) embedded in a cellulose matrix for the production of printed paper electronics (PPE) (nano)technologies. The fate and impact of fragmented AgNWs-PPE was monitored for 21 days in freshwater mesocosms mimicking a pond ecosystem. The Ag release and AgNWs-PPE behavior in water was further characterized in abiotic batch incubations. Qualitative and quantitative analysis were carried out to estimate the Ag partitioning between the environmental compartments and the aging of AgNWs-PPE as well as bioaccumulation and behavior responses in biota. In contaminated pond mesocosms, the NEP cellulose/ polyvinylidene chloride matrix resulted in a rapid settling of the Ag onto the sediments and prevented Ag release into the water column. The highest aqueous concentration measured corresponded to less than 0.5% of the total Ag introduced. The AgNWs-PPE fragments accumulated at the water sediment interface, where they were rapidly (bio)degraded and became bioavailable for benthic organisms. Aquatic snails accumulated a significant fraction of the Ag (1.4±0.5%) and displayed enhanced burrowing behavior in comparison to controls. In batch experiments, alteration of the ENMs morphology was evident at the NEPs surface. Here, the colocalization of Ag and S clusters, suggest an aging via sulfidation similar to other pristine Ag-ENMs. However, the cellulose matrix prevented weathering of the AgNWs within the NEP, which presented a near pristine state even after 21 days incubation. Overall, these results indicate that the fate and impact of Ag embedded in the AgNWs-PPE was driven by the cellulose matrix. In particular, given the specific properties and behavior of paper-based (nano)products, these may constitute a unique category when evaluating NEPs environmental impact. The data gathered in this study can help in defining the environmental fate of such materials and provide useful information to address future studies focused on ENM environmental fate and risk assessment in a life-cycle perspective.

Introduction

Nanotechnologies are expected to impact our future with the development of safer and sustainable solutions in the context of the environmental and energetic transition [START_REF] Rose | The SERENADE project; a step forward in the safe by design process of nanomaterials: The benefits of a diverse and interdisciplinary approach[END_REF]. In this framework, green (nano)technologies such as printed paper electronics (PPE) devices are an emerging and potentially game-changing technology, due to ease of recycling, economics of manufacture and applicability to flexible electronics [START_REF] Liao | Flexible and printable paper-based strain sensors for wearable and large-area green electronics[END_REF]. However, the environmental safety and sustainability aspects of nano-enabled products (NEPs) are being investigated at a relatively slow pace compared to their development and commercialization [START_REF] Mitrano | Review of nanomaterial aging and transformations through the life cycle of nano-enhanced products[END_REF]. Recently, silver nanowires (AgNWs) have gathered attention in PPE due to their excellent optoelectronic properties, ease of processing, and large-scale fabrication capability [START_REF] Li | Screen printing of silver nanowires: balancing conductivity with transparency while maintaining flexibility and stretchability[END_REF]. An expected promising application of AgNWs-based PPE is the production of sensors and electric device displays of high transparency, flexibility, and stretchability [START_REF] Li | Highly transparent and conductive electrodes enabled by scalable printing-and-sintering of silver nanowires[END_REF]. Although AgNWs-based PPE can represent a robust, more sustainable and eco-friendly alternative to conventional ones, knowledge on their environmental risk is largely unknown(2) , [START_REF] Liu | Future paper based printed circuit boards for green electronics: fabrication and life cycle assessment[END_REF]. Furthermore, the existing data sets generated from exposure and hazard studies with pristine Ag-based ENMs could not be representative of Ag-based NEPs behavior, fate and hazard and are inadequate to support meaningful regulation and assessment tasks [START_REF] Moeta | Nano-enabled products in South Africa and the assessment of environmental exposure potential for engineered nanomaterials[END_REF]. Previous studies on Ag-NEPs showed that their environmental fate and impact strongly depends on manufacturing processes (i.e. with Ag ENMs applied as surface coating or embedded in the product matrix), the initial ENM load and the Ag speciation( 8) , ( 9) , [START_REF] Lorenz | Characterization of silver release from commercially available functional (nano)textiles[END_REF] , [START_REF] Lombi | Silver speciation and release in commercial antimicrobial textiles as influenced by washing[END_REF]. The NEPs matrix is also an important factor driving ENMs fate, and was proposed as a main criteria when grouping NEPs eco-toxicity during the use phase [START_REF] Amorim | Environmental Impacts by Fragments Released from Nanoenabled Products: A Multiassay, Multimaterial Exploration by the SUN Approach[END_REF]. Pourzahedi et al., [START_REF] Pourzahedi | Life Cycle Assessment and Release Studies for 15 Nanosilver-Enabled Consumer Products: Investigating Hotspots and Patterns of Contribution[END_REF] performed a life cycle assessment of 15 Ag-NEP (textiles and medical fabrics) and highlighted the role of the matrix, with solid polymeric matrices releasing more silver during washing compared to fibrous materials. A recent study investigated the fate of AgNWs printed on PPE (AgNWs-PPE) during a pilot-scale recycling process [START_REF] Zareeipolgardani | Multiscale investigation of the fate of silver during printed paper electronics recycling[END_REF], showing that the released effluents were free from Ag, which was retained in the pulp conserved for recycling. The main aim of the present study was to mimic a chronic contamination of an aquatic ecosystems by AgNWs-PPE to assess, at environmentally relevant conditions, the fate and effects upon release in the environment. Given the expected long PPE lifetime, compared to the experiment duration, a pre-aging protocol was applied. In particular, the AgNWs-PPE samples were fragmented in order to obtain a fragmented product (FP), resulting in an increased surface area at the NEP/ecosystem interface [START_REF] Nowack | Meeting the Needs for Released Nanomaterials Required for Further Testing-The SUN Approach[END_REF]. Mesocosms allow for simulating realistic ecological conditions, and experiments can be designed to study the aging of ENMs while characterizing the associated exposure and hazard [START_REF] Masion | Monitoring the Environmental Aging of Nanomaterials: An Opportunity for Mesocosm Testing?[END_REF] , [START_REF] Auffan | Contribution of mesocosm testing to a single-step and exposure-driven environmental risk assessment of engineered nanomaterials[END_REF] , [START_REF] Carboni | Aquatic Mesocosm Strategies for the Environmental Fate and Risk Assessment of Engineered Nanomaterials[END_REF]. In the last years, such experimental systems have been extensively applied for the study of ENM fate e.g. ( 19) , ( 20) , ( 21) , [START_REF] Avellan | Gold nanoparticle biodissolution by a freshwater macrophyte and its associated microbiome[END_REF] , but to a lesser extent to NEP [START_REF] Cleveland | Pilot estuarine mesocosm study on the environmental fate of Silver nanomaterials leached from consumer products[END_REF] , [START_REF] Auffan | Environmental exposure of a simulated pond ecosystem to a CuO nanoparticle-based wood stain throughout its life cycle[END_REF]. Using freshwater mesocosms, we first characterized the partitioning of Ag between the environmental compartments. The results obtained with AgNWs-PPE were compared to pristine AgNWs in order to identify the potential role of the matrix in determining the Ag partitioning. Complementary mechanistic abiotic batch experiments were used to further characterize the fate of the NEP and incorporated ENMs. Eventually, we also investigated the bioaccumulation into benthic grazers and assessed behavioral markers, which can be sensitive tools at concentrations far below the lethal effect [START_REF] Amiard-Triquet | Behavioral Disturbances: The Missing Link between Sub-Organismal and Supra-Organismal Responses to Stress? Prospects Based on Aquatic Research[END_REF]. This study provides realistic environmental exposure data, which are needed to inform decisionmaking to mitigate or manage the risks associated to ENMs and NEPs. 2.1. PPE and AgNWs The AgNW u pen ion were obtained by diluting a commercial tandard (nominal 120 μm AgNW length, 20 g[Ag].L -1 in water; ACS material, CA, US) in ultrapure water (UPW; Milli-Q®, Merck, Germany). The AgNWs, characterized by scanning electron microscopy (SEM) coupled to Energy Dispersive X-Ray Spectroscopy (EDX), displayed an average width of 94.3±8.5 nm and an average length of 13.1±4.3 μm (n=40) (Fig. 1a). The AgNWs-PPE samples were received from Arjowiggins now Fedicroni Co. (Voiron, France), as 210mm x 297mm sheets consisting of three distinct layers, namely made of cellulose, an organic coating and conductive ink, respectively. Briefly, a 60 µm thick mixed-cellulose nanofibrils (CNF) matrix was coated, on one side, with a layer of polyvinylidene chloride (PVDC, 5 µm thickness). A conductive ink layer (200-300 nm in thickness) incorporating nano-cellulose fibers and the AgNWs was applied on top of the organic coating, resulting in a sheet resistance of approximately 30 Ω/sq across the samples. The concentration of Ag in the final AgNWs-PPE samples was 1.38±0.07 mg.g -1 .

For both the mesocosm and abiotic batch experiments, the original AgNWs-PPE sheet samples were fragmented, in order to mimic the potential phy ical-chemical form at which the NEP are relea ed in the environment during the use phase. The protocol for fragmentation was optimized from procedures reported in the literature( 12) , (15) , [START_REF] Auffan | Environmental exposure of a simulated pond ecosystem to a CuO nanoparticle-based wood stain throughout its life cycle[END_REF]. First, the PPE samples were cut with a ceramic blade to approximately 2 x 2 mm fragments and transferred into 2 ml plastic vials (Eppendorf, Hamburg, Germany). Then, two zirconium marbles were added in each of the vials, which were loaded into a Teflon sample holder and soaked into liquid nitrogen. The frozen samples underwent two 30 sec cycles of ball-milling at 30 Hz (Mixer Mill MM400, Retsch). Liquid nitrogen was poured between the two fragmentation cycles to prevent unfreezing. Finally, a sieving at 0.5 mm was performed in order to remove the largest PPE fragments. The AgNWs-PPE fragments size, assessed with optical micro copy wa 107.2±72.2 μm on a erage (n=180). A hown in Fig. 1b, in AgNWs-PPE fragments the ENMs were homogeneously distributed onto the NEPs, and displayed average diameter of 81.0 ± 11.1 nm and a erage length of 15.6 ± 3.5 μm (n=40). At the AgNW -PPE surface, the AgNWs formed entanglements, displaying protruding nano-structures as well as portions merged within the cellulose matrix (Fig. 1c). 2.2. Mesocosm set-up and operation Nine indoor aquatic mesocosms were set up to mimic a natural pond ecosystem in the preserved Natura 2000 reserve network, southern France (43.34361 N, 6.259663 E, altitude 107 m a.s.l., Mediterranean climate). Natural sediments, natural water and organisms were collected at the site within 3 days before setting up the mesocosms. Natural sediments were consecutively sieved at 2 mm and 250 μm in order to obtain a homogeneous inoculum containing primary producers (e.g., algae, bacteria). Natural water samples were first sieved at 250 μm and then filtered at 0.2 μm in order to collect suspended matter and biota onto the glass fiber filters. The filters were then placed into 1.5 L Volvic ® water under constant agitation for 48 hrs. Each mesocosm consisted of a glass tank (750 × 200 × 600 mm) consecutively filled with a layer of artificial sediment (79% SiO 2 , 20% kaolinite, and 1% CaCO 3 ) covered with 300 g of water-saturated natural sediment inoculum (48% water content) and 46 L of Volvic ® water, with pH and conductivity values similar to those of the natural pond water. After one week, 0.1 L of water inoculum was added to each aquarium. The mesocosms operated on a 14 h day, 10 h night lighting period. Additional details about mesocosm set-up and monitoring can be found in [START_REF] Auffan | An adaptable mesocosm platform for performing integrated assessments of nanomaterial risk in complex environmental systems[END_REF] [START_REF] Auffan | An adaptable mesocosm platform for performing integrated assessments of nanomaterial risk in complex environmental systems[END_REF]. The set-up was followed by an equilibration period of two weeks, in order to allow the settling of suspended matter, stabilization of the physico-chemical parameters (pH, oxidation-reduction potential [ORP], dissolved O 2 ) and the development of the primary producers [START_REF] Tella | Transfer, Transformation, and Impacts of Ceria Nanomaterials in Aquatic Mesocosms Simulating a Pond Ecosystem[END_REF]. After the equilibration period, organisms were introduced at a density matching the one observed in the natural biotope. In each mesocosm, 70 mollusks (A. leucostoma (Millet 1813), benthic grazers) and 90 micro-crustaceans (Daphnia sp., (Muller 1785), planktonic filter feeders) were added. Three days after the addition of the organisms, the mesocosms were randomly assigned to either contaminated or non-contaminated (control) treatments. Contamination was achieved by chronic (press) addition of either fragmented AgNWs-PPE or pristine AgNWs three times per week (9 injections in total) during 21 days. Spiking suspensions were prepared in UPW immediately before addition, vigorously stirred and homogeneously distributed at the water surface. In AgNWs-PPE treatments, each contamination consisted of 175 mg of AgNWs-PPE fragments, corresponding to single Ag doses of 241 µg and a final concentration of 46.7 µg[Ag].L -1 in the mesocosms. Such a dose was selected as a compromise between the low levels expected in the real ecosystems and the possibility to analyze and characterize the AgNWs-PPE in complex environmental matrices. For AgNWs treatments, single Ag doses of 2.42 mg led to concentration of 470 µg[Ag].L -1 at the end of the experiment. Such a dose corresponded to a ENMs-surface to mesocosm water ratio of 1.9 x 10 -3 m 2 .L -1 (at day 21), and was chosen to match the values recently used for other Ag-ENMs in mesocosms [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF].

Mesocosm sampling and analysis

Several abiotic and biotic parameters were monitored to inform about the exposure and effects of contamination on the aquatic ecosystem. These included the continuous measurements of water physical-chemical properties (pH, oxidation-reduction potentiel (ORP), electrical conductivity, temperature, dissolved oxygen) as well as punctual measurements of biotic parameters (Ag concentration in water column, sediment and organisms, bacterial counting, total organic carbon content (TOC), number of suspended particles and chlorophyll concentration( 20) , ( 26)). A first aim of the present study was to quantify the distribution of Ag in the mesocosm following AgNWs or AgNWs-PPE contamination. For this purpose, superficial sediments, the water column and the aquatic snails A. leucostoma were sampled prior to contamination and then after 7, 14 and 21 days. For water column samples, 10 mL were collected at approximately 10 cm below the water surface, transferred into 15 mL plastic tubes and acidified with 2% ultrapure HNO 3 (Normatom ® ; SCPScience, Quebec, Canada). Additional 5 mL water samples underwent ultrafiltration at 3 KDa (Amicon Millipore; Merck KgaA, Darmstadt, Germany) prior to analysis, for the assessment of dissolved Ag concentrations. Surficial sediment and A. leucostoma samples were dried for 3 days at 65 °C, manually ground in an agate mortar, and then transferred into 15 mL Teflon tubes. 50 mg of each sample was processed as follows. For sediments, 1 mL HCl (34%) (Normatom ® ), 2 mL HNO 3 (67%) and 0.5 mL HF (47-51%, PlasmaPure ® ) were added. For A. leucostoma, an additional 0.5 mL of H 2 O 2 (30-32%, PlasmaPure ® ) was added. Then, acid digestion was performed at 180 °C in a Milestone UltraWAVE microwave digestion system for 1 h. The metal analysis was performed with an inductively coupled plasma mass spectrometer (ICP-MS, NexIon 300X, Perkin Elmer ® ).

Abiotic batch aging experiments

In parallel to the mesocosm experiment, an abiotic batch experiment was carried out for 21d in 250 mL glass beakers, filled with 200 ml Volvic ® water to gather mechanistic information on the aging of the AgNWs-PPE fragments in abiotic conditions close to those in the mesocosm (e.g. water composition, temperature, day/night cycle). Dosing was carried out with a single addition (pulse dosing) of fragmented AgNWs-PPE at day 0 to match the final dose used in AgNWs-PPE mesocosms (50 µg[Ag].L -1 ). In the abiotic batch experiment the water was sampled similar to the mesocosms for total and dissolved (<3KDa) Ag concentrations. In addition, at the end of the experiment the AgNWs-PPE were recovered from the beakers, rinsed with UPW and transferred onto glass petri dishes. The excess water was removed with a pipette and the samples were let to dry at room temperature in the dark. The dried samples were stored at 4 °C in the dark for elemental and SEM analysis. SEM analysis was performed with a Zeiss GeminiSEM 500 ultra-high resolution Field Emission Electron Microscope. For imaging, this system was equipped with a column-mounted detector for high efficiency surface secondary electron detection (In Lens SE) as well as variable pressure secondary electron (VPSE) and variable pressure backscattered electron (VP BSE) detectors to observe non-conductive samples. The surface analysis of a region of interest was acquired first at low voltage (0.5 kV, In Lens SE detector) to increase topographic contrast and reduce specimen charging, and then at higher energy (7 kV, VP BSE detector), to analyze a deeper fraction of the samples and for Energy Dispersive X-ray Spectroscopy (EDX) acquisition. The electron acceleration tension was set at 7 kV in order to achieve sufficient X-ray emission to detect Ag L lines at 3 keV, whereas analysis at higher energies was not possible due to irreversible damage of the PPE matrix. For the chemical imaging, EDX was performed using an EDAX Octane Silicon drift detector (129 eV energy resolution for Manganese). During SEM-EDX mapping, the dwell time for each point was set to 50 µs per pixel and frames were accumulated with drift correction. Reference images taken at the beginning of the acquisition were periodically checked to monitor the potential alteration of the samples during measurements. In AgNWs-PPE contaminated mesocosms, the Ag concentration in the sediments increased with the time to values significantly higher than those of the controls (Fig. 2a). In detail, following chronic exposure of AgNWs-PPE fragments, the Ag in the sediments was 6.4 ± 1.8 mg.kg -1 after 7 days and reached 13.8 ± 3.9 mg.kg -1 at the end of the experiment. These results indicate that the benthic compartment was the main sink for Ag in the simulated pond ecosystem, accumulating up to 97±28% of the total Ag injected at 21d. The Ag partitioning toward the sediments was accompanied by its minor presence in the water column (Fig. 2b) with a total aqueous concentration of Ag reaching 0.10 ± 0.03 μg.L -1 after 7d. This concentration in the water column remained stable until the end of the experiment with 0.19 ± 0.08 μg.L -1 at 14d and 0.14 ± 0.01 μg.L -1 at 21d, corresponding to approximately 0.5% of the Ag initially introduced. This is in agreement with the macroscopic observation of the fast settling down of the AgNWs-PPE fragments right after the injection in the mesocosm surface water. Furthermore, no significant Ag was measured in water after ultrafiltration, indicating either the lack of Ag dissolution or the rapid removal of Ag dissolved species from the water column via reprecipitation or complexation, which was already observed in previous studies( 26) , [START_REF] Furtado | The persistence and transformation of silver nanoparticles in littoral lake mesocosms monitored using various analytical techniques[END_REF]. It must be noted that the Ag species released form the AgNWs-PPE may also be retained by the ultrafiltration membrane applied, because associated to NEP product matrix (e.g. Ag-cellulose complexes) or due to absorption onto the membrane. In freshwater mesocosms, important ENM removal from the water column is well known and has been reported for a variety of ecosystem settings and nanomaterials tested. Previous studies, focused on pristine Ag-ENMs with a similar contamination scenario (i.e. chronic dosing at similar Ag total doses), reported comparable accumulation in sediment for a range of Ag-ENMs differing in coatings and morphology. For instance, in pond indoor mesocosms contaminated with Ag-ENM plates, the surficial sediments accumulated 85±19% of the Ag after 28 d of chronic exposure [START_REF] Colman | Dosing, Not the Dose: Comparing Chronic and Pulsed Silver Nanoparticle Exposures[END_REF] , [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF]. In these cases, compartmentalization into sediments was attributed to ENMs homo-aggregation phenomena as well as hetero-aggregation with natural organic matter and suspended solids [START_REF] Lowry | Long-Term Transformation and Fate of Manufactured Ag Nanoparticles in a Simulated Large Scale Freshwater Emergent Wetland[END_REF] , [START_REF] Tella | Transfer, Transformation, and Impacts of Ceria Nanomaterials in Aquatic Mesocosms Simulating a Pond Ecosystem[END_REF]. In this study, AgNWs-PPE fragments were observed to settle rapidly after spiking into the mesocosm surface water, suggesting that the sediment accumulation was likely driven by the cellulose matrix. Nonetheless, the Ag released in water from the AgNWs-PPE was unexpectedly low for such a surface coated NEP. Indeed, the silver metal and ENM species release from NEPs is known to rely on the product matrix and manufacturing(8) , [START_REF] Lombi | Silver speciation and release in commercial antimicrobial textiles as influenced by washing[END_REF]. In general, high release rates can be expected for ENMs present at the surface, compared to those incorporated into the NEP bulk(3) , [START_REF] Amorim | Environmental Impacts by Fragments Released from Nanoenabled Products: A Multiassay, Multimaterial Exploration by the SUN Approach[END_REF]. For instance, in freshwater aquatic mesocosm studies the Ag released from Ag-NEP surface coated textiles displayed higher aqueous concentration and persistence compared to pristine Ag-ENMs [START_REF] Cleveland | Pilot estuarine mesocosm study on the environmental fate of Silver nanomaterials leached from consumer products[END_REF]. On the contrary, ENMs included into materials such as acrylic paint displayed diminished release in water [START_REF] Auffan | Environmental exposure of a simulated pond ecosystem to a CuO nanoparticle-based wood stain throughout its life cycle[END_REF]. Here, although the AgNWs were present at the surface of the PPE, the Ag partitioning resembled that observed for ENMs incorporated into NEPs such as paint nanocomposite [START_REF] Auffan | Environmental exposure of a simulated pond ecosystem to a CuO nanoparticle-based wood stain throughout its life cycle[END_REF]. Such behavior could be due to the ecosystem physical-chemical properties but also to the low hydrolysis of the cellulose matrix preventing the short-term Ag release in water as well as the intrinsic physical-chemical properties of AgNWs (e.g. size, morphology and surface chemistry) that can drive their environmental fate. Figure 3. Comparison between the Ag partitioning in the aqueous phase (% of total Ag introduced) of AgNWs-PPE (red) and pristine AgNWs (wires, blue), Ag nano-plates (plates, white) and Ag nano-

spheres (spheres, black). Ag nano-plates and Ag nano-spheres data with permission from Auffan et al. (2020)(27).

To elucidate the role of the PPE matrix in determining the Ag behavior, the Ag partitioning following AgNWs-PPE treatment was compared to that of pristine AgNWs (this study) as well as other pristine Ag nano-plates and nano-spheres recently investigated in freshwater mesocosms by our team [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF]. For this purpose, the AgNWs were dosed into mesocosms at similar surface area-based concentrations as the ones used for Ag nano-plates and nano-spheres (Auffan et al., 2020) [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF]. As shown in Fig. 3, in mesocosms contaminated with pristine AgNWs, the percentage of injected Ag found in the water column was in the same range of magnitude than those previously observed for Ag nano-sphere and nano-plates. The average Ag aqueous fraction of all the pristine Ag-ENMs was 9±1% at 14d and 4±2% on average at 21d. Hence, for a given surface area-based concentration the Ag release and water persistence are similar between Ag-ENMs of different shape and size. In mesocosms contaminated with AgNWs-PPE, the water Ag fraction was lower than that of pristine Ag-ENMs at all the time intervals (0.5±0.2% in average). In particular, when compared to pristine AgNWs, the AgNWs-PPE aqueous Ag was 20-fold lower at 7d and 6-fold lower at 21d. These results support the hypothesis that the Ag partitioning observed in AgNWs-PPE treatments was driven by the NEP matrix and not by the specific properties of the AgNWs contained therein.

Mechanisms of AgNMs-PPE aging in freshwater environments

When assessing the AgNWs-PPE and Ag fate in freshwater mesocosms, a major limitation was due to the impossibility to recover the aged NEP fragments. In particular, the AgNWs-PPE accumulated onto the sediment formed flocs, which mixed with and were degraded by me oco m' abiotic (mineral sediment matrix) and biotic (biofilm and benthic organisms) components. In order to monitor the AgNWs-PPE transformation and obtain quantitative and qualitative information, we carried out batch experiments in simplified abiotic conditions. Here, the fragmented AgNWs-PPE was incubated in static conditions, similar to the mesocosm experiment with regard to all the abiotic parameters such as water chemistry, irradiation and temperature. In the batch experiment, the concentrations of Ag in the water increased during the first week and remained stable until the end of the incubation (Supplementary Information, S1). At day 21 the water [Ag] reached 0.93 ± 0.47 μg.L -1 , which was 5-fold higher than the maximum water concentration measured in the mesocosms. This could be related to the lack of natural suspended matter (e.g. organic matter, colloids) and biota (micro-and macro-organisms), in the batch system, which likely play a role in the removal of dissolved or particulate Ag from the mesocosm water column. At the end of the abiotic incubation up to 90.6 ± 3.1 % of the total Ag was found in the aged NEP fragments recovered and the [Ag] measured after ultrafiltration was negligible indicating no free Ag ions in solution.

Overall, these results suggest that the Ag remained associated with the NEP matrix, and highlight a matrix driven Ag fate, suggesting that the contamination of aquatic ecosystems to such cellulose/PVDC-based NEPs predominantly involves the exposure to Ag-PPE complexes rather than free-ENMs and metal species.

Of course, aging under abiotic conditions was a simplified exposure scenario compared to mesocosms, as it did not account for all biodegradation mechanisms. On the other hand, it allowed for an easier recovery of aged AgNWs-PPE fragments, which were analyzed to gather qualitative information about the distribution, morphology and speciation of the AgNWs in the NEP during aging. Before aging, the AgNWs were homogeneously distributed onto one side of the NEP (Fig. 1b) where they formed entanglements displaying protruding nano-structures as well as portions merged within the cellulose matrix. The analysis at higher magnification (Fig. 1c) revealed an irregular (nonhexagonal) morphology of the AgNWs, as well as the presence of NaCl nano-crystals (approx. 10 nm diameter), confirmed by EDX analysis. The latter could be due to the presence of endogenous Cl species in the samples owing to the PVDC coating. As shown in Fig. 4, after 21d of batch incubation, changes in both the NEP matrix and AgNWs morphology and distribution were observed. It must be noted that because of the drying process prior to SEM analysis, an alteration of the cellulose matrix and of the AgNWs could have occurred. With regard to the ENMs, as shown in Fig. 4a the AgNWs could still be identified onto the aged NEPs surface, but presented extensive breakages and changes in morphology. Furthermore, the AgNWs displayed a lower surface density compared to non-aged NEPs, and were all bound to the cellulose matrix, with no protruding nanostructures. The AgNWs density and distribution at the aged surfaces were further investigated by analyzing larger NEPs areas. The detection at low energy (focused on the surface) showed that in aged samples, the AgNWs were not homogeneously distributed onto the cellulose surface (Fig. 4b). Large areas appeared empty and clusters of nanostructures could only be identified in hotspots. Interestingly, the analysis of the same regions at higher energy (both surface and bulk detection) revealed the presence of ENMs incorporated into the cellulose matrix, with distribution similar to that of the original samples (Fig. 4c). These findings suggest that aging of the AgNWs-PPE affected only the AgNWs immediately at the NEP surface, thus directly exposed to the outer environment. However, the majority of the ENMs partially or totally incorporated in the matrix, not directly exposed to weathering, presented a near original state. Hence, our hypothesis is that most of the Ag recovered in the cellulose after 21 d of incubation was in the form of ENM, with morphology and distribution resembling those of the original non-aged NEP. In general, these results are in accordance with a recent study on AgNWs-PPE recycling, in which most of the ENMs were retained in the matrix pulp due to embedding by cellulose nanofibrils [START_REF] Zareeipolgardani | Multiscale investigation of the fate of silver during printed paper electronics recycling[END_REF].

Ag-ENMs are known to undergo transformation processes in environmental media, with the formation of novel Ag species (e.g. Ag sulfuration, AgCl complexes or precipitates)(10) , ( 11) , ( 19) , [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF]. Understanding the occurrence of such processes in NEPs is fundamental, since they can affect ENMs stability, behavior and metal release in aqueous media( 29) , [START_REF] Lead | Nanomaterials in the environment: Behavior, fate, bioavailability, and effects-An updated review: Nanomaterials in the environment[END_REF]. Here, we collected EDX spectra of the AgNWs at the AgNWs-PPE surface in order to investigate the potential changes in the chemical composition and silver speciation. As shown in Fig. 5, in aged AgNWs-PPE we observed an important sulfur signal in AgNWs hotspots, which was not recorded in the cellulose/PVDC NEPs background. Further EDX mapping clarified the colocalization of Ag and S in AgNWs hotspots. In freshwater, Ag sulfidation is a major transformation pathway for pristine Ag-ENMs. It has been extensively reported in mesocosm studies( 19) , [START_REF] Auffan | The shape and speciation of Ag nanoparticles drive their impacts on organisms in a lotic ecosystem[END_REF] and was linked to a reduced toxicity toward biota( 31) , [START_REF] Lehmann | Crumpling of silver nanowires by endolysosomes strongly reduces toxicity[END_REF]. It must be noted that sulfidized Ag-ENMs are often less soluble than their pristine counterparts, though Ag + ion solid diffusion may be high in the Ag 2 S surface layer [START_REF] Simonnin | Phase Transition and Liquid-like Superionic Conduction in Ag 2 S[END_REF], and their formation in the PPE samples may have enhanced their persistence in the nanoproduct. The results obtained in the abiotic batch experiment presented here suggest that similar sulfidation processes could also occur when the Ag-ENMs are incorporated in a NEPs, such as paper-based PPE. However, future works will need to further characterize these processes in biotic experimental conditions and in the natural environment. 3.3 Ecosystem effects induced by AgNW-based PPE Figure 6. Percentage of benthic organisms A. leucosoma displaying a burrowing trait following exposure to fragmented AgNMs-PPE (red circles) versus controls (black triangles). * highlights the differences in mean ± standard deviation (n=3) between the two treatments. The environmental risk posed by a contaminant depends on its toxicity (hazard), speciation and concentration (exposure) toward target ecological niches. In the water column of the mesocosms exposed to AgNWs-PPE, the chronic contamination did not induce significant effects on most biotic and abiotic parameters. Briefly, no significant changes were observed in pH, redox potential and conductivity following AgNWs-PPE addition. The water TOC was not significantly different, over the course of the incubation, between controls (1.71±0.16 mg.L -1 in average) and contaminated mesocosms (1.75±0.25 mg.L -1 on average). The same was valid for chlorophyll and suspended particle amounts as well as planktonic organism (Daphnia sp.) and bacteria populations (Supplementary Information, S2). In general, the low impact on water parameters can be attributed to the Ag speciation and the low Ag water concentration in the mesocosms (Fig. 2b and Fig. 5, respectively). On the other hand, the Ag accumulation in sediments implied a high exposure of the benthic compartment, which was evaluated by monitoring the Ag bioaccumulation and burrowing behavior of the aquatic snail A. leucostoma. Herein, Ag concentrations in A. leucostoma increased up to 43.4 ± 15.1 mg.kg -1 (dw) at 14d and stabilized at 33.7 ± 8.2 mg.kg -1 at 21d (Fig. 2c) following AgNWs-PPE treatments. Interestingly, up to 96 ± 5% of the Ag was not leached from the snails after acidic wash of the external bodies, suggesting that the Ag was ingested by the snail and not adsorbed onto the shell surface. The stabilization of the Ag concentration taken up by the organism between day 14 and 21 could be due to the combination of both ingestion and depuration processes that are well known in mollusks [START_REF] Cleveland | Pilot estuarine mesocosm study on the environmental fate of Silver nanomaterials leached from consumer products[END_REF] , [START_REF] Oliveira-Filho | Effects of AgNPs on the Snail Biomphalaria glabrata: Survival, Reproduction and Silver Accumulation[END_REF]. Such metals uptake/ingestion in benthic organisms has already been reported after contamination with several pristine ENMs such as Ce-ENMs [START_REF] Tella | Transfer, Transformation, and Impacts of Ceria Nanomaterials in Aquatic Mesocosms Simulating a Pond Ecosystem[END_REF], Cu-ENMs [START_REF] Auffan | Environmental exposure of a simulated pond ecosystem to a CuO nanoparticle-based wood stain throughout its life cycle[END_REF] and Ag-ENMs [START_REF] Cleveland | Pilot estuarine mesocosm study on the environmental fate of Silver nanomaterials leached from consumer products[END_REF]. In the case of NEPs, the Ag bioaccumulation in mud snails exposed in freshwater mesocosms was attributed to a trophic transfer route from Ag-enriched biofilm [START_REF] Cleveland | Pilot estuarine mesocosm study on the environmental fate of Silver nanomaterials leached from consumer products[END_REF]. During the present mesocosm experiment, we observed the direct feeding of snails on the AgNWs-PPE fragments accumulated at the sediment/water interface and we hypothesize that the majority of Ag bioaccumulation in snails was due to a direct uptake of AgNWs-PPE. This may have been enhanced by both the cellulose in the NEP matrix, representing a potential food source for the organisms, and the size of fragmented AgNWs-PPE directly bioavailable for the snails. It is noteworthy that the concentrations in the snails observed in this study (~40 mg [Ag].Kg -1 at 21d) were relatively high in consideration of the low dose of total Ag introduced in the mesocosms (~50 ug [Ag].L -1 at 21d). For instance, similar levels (~100 mg [Ag].kg -1 ) were observed in ecotoxicity tests with aquatic snails (B. glabrata) exposed to Ag-ENMs at water concentrations up to 50 times higher [START_REF] Oliveira-Filho | Effects of AgNPs on the Snail Biomphalaria glabrata: Survival, Reproduction and Silver Accumulation[END_REF]. Remarkably, despite significant Ag uptake no enhanced mortality of A. leucostoma adults was observed during the experiment, suggesting no acute toxicity of the Ag from aged AgNWs-PPE for the snails. However, the Ag species taken up did not necessarily enter the animal's systemic circulation, and a low toxicity could be also related to sulfidation of the AgNWs at the surface of the aged AgNWs-PPE (Figure 5), which is known to reduce noxious effects to aquatic biota [START_REF] Carboni | Aquatic Mesocosm Strategies for the Environmental Fate and Risk Assessment of Engineered Nanomaterials[END_REF] , [START_REF] Yuan | Stress Responses of Aquatic Plants to Silver Nanoparticles[END_REF]. In addition, no significant effect was observed on benthic bacteria population density (Supplementary information, S3). However, aquatic organisms can respond to stress with a combination of physiological adaptation and behavior responses, which are known to occur after heavy metal contamination [START_REF] Mcinnes | Effects of metals on the behaviour and oxygen consumption of the mud snail[END_REF], but are less studied in the case of ENMs and NEP [START_REF] Buffet | Behavioural and biochemical responses of two marine invertebrates Scrobicularia plana and Hediste diversicolor to copper oxide nanoparticles[END_REF]. In freshwater snails, such traits are observed as a response to drought and predation but also result as a consequence of contamination [START_REF] Hazelton | Chronic fluoxetine exposure alters movement and burrowing in adult freshwater mussels[END_REF]. In the present study, we noticed an increased burrowing of the A. leucostoma after one week of chronic contamination and monitored it until the end of the experiment. As shown in Fig. 6, in mesocosms treated with AgNWs-PPE, the snail burrowing activity was observed in up to 50±7% of the snails, on average, during the second week of experiment and 30±10% of the snails, on average, during the third week. This was significantly higher than controls at d8, d9 and d15. Similar stress responses, in mesocosms contaminated with ENMs, were reported by Buffet et al., [START_REF] Buffet | Behavioural and biochemical responses of two marine invertebrates Scrobicularia plana and Hediste diversicolor to copper oxide nanoparticles[END_REF] , [START_REF] Buffet | Biochemical and behavioural responses of the endobenthic bivalve Scrobicularia plana to silver nanoparticles in seawater and microalgal food[END_REF] who showed behavioral impairment (burrowing kinetic, feeding rate) of bivalve exposed to Cu-and Ag-ENMs in seawater ecosystems. It is worth comparing these biological effects with those observed in the mesocosm contaminated with pristine AgNWs, which will be discussed in detail in a future study. Here, at d21 the sediments were also the main sink for Ag, accumulating 92±5% of the total dose injected. Furthermore, the Ag concentrations in snails were comparable to those observed in NEP treatments (33±8 mg [Ag].kg -1 for AgNWs-PPE and 69±4 mg [Ag].kg -1 for pristine AgNWs), despite the 10-fold higher total Ag introduced in the mesocosms. However, in the pristine AgNWs treatment, no significant burrowing alteration was observed in comparison to controls. These interesting results suggest that the altered behavior following AgNWs-PPE treatments was not due to the absolute amount of Ag uptake. Rather, it could be induced by a different physical-chemical state and speciation of the AgNWs in the ingested AgNWs-PPE fragment compared to pristine ENMs, or due to a direct positive effect of the PPE matrix uptake on the mollusc metabolism and behavior. Future studies will need to explore the role of the cellulose matrix in the biodistribution and biotransformation of ENMs in aquatic ecosystems more deeply, e.g. by including matrix-only treatments. A better understanding of AgNWs-PPE effects on biota will also require the combination of behavioral traits observations, such as burrowing, with other biological response analysis (e.g. biochemical markers analysis). 5. Environmental implications of paper-based NEPs ENMs are being produced and applied in a wide range of NEPs [START_REF] Nowack | Meeting the Needs for Released Nanomaterials Required for Further Testing-The SUN Approach[END_REF], where they are incorporated into a variety of liquid or solid formulations(39) , ( 40) , [START_REF] Scifo | Non-linear release dynamics for a CeO2 nanomaterial embedded in a protective wood stain, due to matrix photo-degradation[END_REF]. Several studies recognize that the ENMs integration in such matrices determines the properties of the materials that may be released throughout the life cycle [START_REF] Liu | Continued Efforts on Nanomaterial-Environmental Health and Safety Is Critical to Maintain Sustainable Growth of Nanoindustry[END_REF]. Hence, the matrices can affect the behavior, fate and impact of ENMs when compared to pristine nanoparticles, e.g. with regard to their partitioning in the ecosystem, the metal release rates and speciation, and ecotoxicity effects( 12) , [START_REF] Wohlleben | The nanoGRAVUR framework to group (nano)materials for their occupational, consumer, environmental risks based on a harmonized set of material properties, applied to 34 case studies[END_REF]. Here, we investigated the fate and effects of AgNWs incorporated into a cellulose/PVDC matrix for the construction of novel AgNWs-PPE nanotechnologies. In this study, the NEP matrix drove the fate of the incorporated ENMs, influencing their environmental behavior and interaction with biota. In general, the PPE exposure potential (defined as the potential to release ENMs and metals( 7)) appeared to be low, and pond ecosystems contaminated with AgNWs-PPE were exposed to ENMs-matrix complexes rather than free ENMs and metals. Considering an environmental fate and risk perspective, we observed that the PPE accumulated in the pond sediments, resulting in a fast removal of the Ag from the water column, but also an enhanced exposure of the benthic compartment. A lower distribution in water does not necessarily translate to a diminished environmental risk, but to a change in the magnitude to which target ecological niches will be exposed. Indeed, a higher accumulation of the paper-based NEPs in sediments may imply a higher ENMs exposure and bioavailability for benthic organisms, such as the aquatic snails in the present study. Here, no acute toxicity was observed toward the snails, at concentrations higher than those expected in real ecosystems [START_REF] Giese | Risks, Release and Concentrations of Engineered Nanomaterial in the Environment[END_REF]. However, paper-based NEPs may represent a food source for aquatic organisms, resulting in an enhanced bioaccumulation and behavioral changes, which must be monitored for a proper assessment of NEPs impact and should be explored more thoroughly by future studies. We also observed that paper-based NEPs represent a very peculiar material, whose behavior could be hard to predict compared to other matrices such as plastic or cement nano-composites [START_REF] Amorim | Environmental Impacts by Fragments Released from Nanoenabled Products: A Multiassay, Multimaterial Exploration by the SUN Approach[END_REF]. In particular, the fragmented AgNWs-PPE is in principle a (bio)degradable product, but formed flocs that accumulated at the sediment surface in the static mesocosm conditions. Due to the ease of resuspension of such flocs, an enhanced transport may be expected in dynamic lotic environments, such as rivers and flowing water. Future studies will need to address these questions, and to further investigate the state at which paper-based NEPs enter and are transformed in the environment, as well as their transport and removal from aquatic systems. Accordingly, when grouping NEP fate and impact based on the matrix material, paper-based NEPs may constitute a category of their own. This is also supported by recent studies testing food contact release of nano-forms from packaging NEPs, which highlighted a different behavior of paper-based NEPs compared to other materials such as polymeric matrices [START_REF] Ruggiero | Food contact of paper and plastic products containing SiO2, Cu-Phthalocyanine, Fe2O3, CaCO3: Ranking factors that control the similarity of form and rate of release[END_REF]. The data and information reported in this study will be useful for tracking and predicting the fate of ENMs embedded in similar matrices. Acknowledgement

3 .

 3 Figure 2. (a) Total concentration of Ag in the superficial sediments, (b) total and dissolved (<3KDa) Ag concentrations in the water column, and (c) total concentration of Ag in A. leucostoma in mesocosms contaminated with AgNW-PPE versus controls. Dotted-line in (A) represents the concentration expected if all the Ag added to the mesocosms settled at the surface of the sediments. Error bars indicate the standard deviation between mesocosms in the same treatment (n=3).

This work was supported by the Excellence Initiative of Aix-Marseille University -A*MIDEX, a French "In e ti ement d'A enir" program, through it a ociated Labex SERENADE project. Thi work i also a contribution to the OSU-Institut Pythéas. The authors acknowledge the CNRS funding for the IRP iNOVE.


M Ernoult
email: ernoult@ijclab.in2p3.fr

X Doligez

N Thiolliere

Advanced Tools for Uncertainties Visualization and Analysis in Fuel Cycle Simulations

Keywords: M. Ernoult, X Doligez, Nicolas Thiollière nuclear fuel cycle, uncertainties, visualization

  L'archive ouverte pluridisciplinaire

INTRODUCTION

Facilities of the nuclear fuel cycle have long operation time of several decades and the time between their design and the start of their operation is often also more than a decade. To be able to compare designs and sizing of those facilities that will allow future strategies, fuel cycle simulation tools have been developed [START_REF] Mouginot | Core Library for Advanced Scenario Simulation[END_REF][START_REF] Coquelet-Pascal | COSI6: A tool for nuclear transition scenario studies and application to SFR deployment scenarios with minor actinide transmutation[END_REF]. They allow the simulation of the evolution of materials in the fuel cycle. Thanks to them, when a set of strategies for possible fuel cycle transitions is considered, the performance and possible outcomes can be evaluated through scenario studies. These Scenario studies are not at all necessarily simulations of a coherent futures, but more importantly, thought experiments to generate and observe potential consequences of possible future orientations of interest. However, even with the most care and study, the simulation of a transition strategy is necessarily subject to numerous uncertainties.

The majority of the scenarios developed by nuclear institutions and industry players are operational proposals based on feedback from the nuclear industry. They rely on technologies already developed or in use and draw primarily on hypotheses supporting the continued use of nuclear energy, with an unchanged installed capacity [START_REF] Martin | French Scenarios Towards Fast Plutonium Multi-recycling in PWR[END_REF]. The scenarios proposed by the research world can sometimes free themselves from obligation of practical possibility to explore futures that disrupt the current system. Examples include research carried out on transmutation in ADS (accelerator-driven reactors) [START_REF] Romanello | Comparative study of fast critical burner reactors and subcritical accelerator driven systems and the impact on transuranics inventory in a regional fuel cycle[END_REF], actinides recycling in MSR (molten salt reactors) [START_REF] Heuer | Towards the thorium fuel cycle with molten salt fast reactors[END_REF], and studies on the thorium cycle [START_REF] Ernoult | Gestion avancée du plutonium en REP -Complémentarité des cycles thorium et uranium[END_REF].

Scenarios often act as "boundary objects" providing an opportunity to bring together different communities of stakeholders, with various knowledge and different, and sometimes opposing, interests in order to compare their visions for the future, organize their strategies and even cooperate. To facilitate this role of scenario study as boundary objects, and embrace the part of this role that makes them object of communication, producers of scenarios have to pay attention to the method they use to present their results, keeping them intelligible to the other communities.

In depth analysis have been made of the different type of uncertainties in electro-nuclear scenario studies [START_REF] Krivtchik | Analysis of uncertainty propagation in nuclear fuel cycle scenarios[END_REF][START_REF] Zhou | Resilience analysis of nuclear fuel cycle scenarios[END_REF]. They have brought to light six families of uncertainties: Base Physics Data (including Nuclear data), Physics modeling, Historical data, The choice of parameter values for the future strategy, Formulation of criteria to satisfy, allowing to figure out the relevant trajectories and strategies of interest as well as Hypotheses in the political, economic, industrial and societal decisions that may impact the nuclear fleet. [START_REF] Krivtchik | Analysis of uncertainty propagation in nuclear fuel cycle scenarios[END_REF] has represented the uncertainties of nuclear data can be, for the major part of nuclide decays and nuclear reactions in electro-nuclear scenario studies,and analyzed their propagation. The Fit Project [START_REF] Thiollière | Impact of fresh fuel loading management in fuel cycle simulators: A functionality isolation test[END_REF] has started to tackle the question of uncertainties of physics modeling caused by essential simplifications or approximations in the resolutions of equations made to achieve acceptable computational costs. Several work dedicated to one specific modeling assumption have also be done in this direction [START_REF] Doligez | Advanced Modeling of the historical French Nuclear fleet with DONJON5 -CLASS copled simulations[END_REF][START_REF] Ernoult | Automated selection of nuclides and reactions of interest in a depletion simulation[END_REF]. [START_REF] Courtin | Etude de l'incinération du plutonium en REP MOX sur support d'uranium enrichi avec le code de simulation dynamique du cycle[END_REF] has studied the uncertainties of historical data. And [START_REF] Liang | Robustness assessment in electro-nuclear scenario studies[END_REF] and [START_REF] Zhou | Resilience analysis of nuclear fuel cycle scenarios[END_REF] have developed new methods and tools to take into account uncertainties of external decision and evaluate their impacts. All theses analysis either present their result using complex mathematical objects, like Sobol indices or chaos polynomial coefficients [START_REF] Skarbeli | Uncertainty quantification on advanced fuel cycle scenario simulations applying local and global methods[END_REF][START_REF] Courtin | Etude de l'incinération du plutonium en REP MOX sur support d'uranium enrichi avec le code de simulation dynamique du cycle[END_REF], or using a very long series of one-to-one comparisons [START_REF] Edwards | The Effects of the Uncertainty of Input Parameters on Nuclear Fuel Cycle Scenario Studies[END_REF]. They allow in-depth understanding of interaction of uncertainties with scenario study results, but are not easy to share with other stakeholders and used as a basis of fruitful exchanges. The indices ask for a math background that is rare in communities without a strong scientific background, and the length series of dual figures are difficult to digest for strategic actors that have less time. Both are not easy to share with other stakeholders and used as a basis of fruitful exchanges. This lead to uncertainties not being well communicated during discussion with different communities of stakeholders and often not taken into consideration as much as their level would require.

We therefore developed dedicated associated tools to calculate easily and represent uncertainties linked to our fuel studies inspired by the graph with colored area used in IPCC summaries for policy-makers.

NEW TOOLS FOR VISUALIZATION OF UNCERTAINTIES

To help the calculation and representation of uncertainties, a new tool has been developed. It takes the form of a python package, providing many easy-to-use function for launching of post-treatment scripts for the fuel cycle simulation tool. Currently only CLASS [START_REF] Mouginot | Core Library for Advanced Scenario Simulation[END_REF] is supported, but the functions have been designed with modularity in mind and other simulation codes would be easy to plug as long as their input files are available in text form. This package can read an input file created for the fuel cycle simulator and auto-extract the variable. It can then change them depending on the need of the user or the algorithm and launch the simulation or just produce a new input file to be launched by the user. The new values for the variable can be explicitly given or sampled following a random distribution. Most common random distributions are available, notable uniform, normal, triangle and log-normal. With those integrated functions, a whole script of analysis takes only a few lines. Once the calculations are done, outputs from a large number of simulations coming from the same initial input are easily visualized together, and distribution and statistical indicators are available. All figures presented in this paper are a direct result of the package.

APPLICATION TO HISTORICAL FRENCH FLEET SIMULATION

Characterization of uncertainties

To demonstrate the capabilities of this tool, we applied it on the last published scenario study made with CLASS, the simulation of a scenario closely inspired by the history of the French Fleet from [START_REF] Doligez | Advanced modeling of the historical French nuclear fleet with DONJON5/CLASS coupled simulations[END_REF]. This study is a relatively precise simulation of the deployment of a fleet of PWR using first only UOX fuel and starting later to use MOX fuel recycling plutonium from spent UOX assemblies. This scenario is closely inspired by the French nuclear fuel cycle from its start to 2021, and takes into account individual reactor and the specific fuel management strategies that have been used in each of them in the past and does every simulation using both MLP [START_REF] Leniau | A neural network approach for burn-up calculation and its application to the dynamic fuel cycle code CLASS[END_REF], and DONJON [START_REF] Guillet | Coupled CLASS and DONJON5 3D full-core calculations and comparison with the neural network approach for fuel cycles involving MOX fueled PWRs[END_REF] physics models in order to check if the result are still significant when considering uncertainties.

An input based on [START_REF] Doligez | Advanced modeling of the historical French nuclear fleet with DONJON5/CLASS coupled simulations[END_REF] has been created with options for MLP and DONJON physics models. The autoanalyze from the package found the 321 variables. Uncertainties about a lot of these variables have then been characterized. For example, the initial input used the nominal burn-up for each campaign in each reactor, but data show that there is variation of the burn-up between irradiation cycles. For example, uncertainty has been associated with all burn-up following a triangular distribution peaking at the nominal and using the minimum and maximum observed burn-up in the campaign as limits. Uncertainties on load factors that are subject to the need of the electricity network have been modeled by a normal distribution with the observed averaged and a relative standard deviation of 0.015 allowing to reach the most extreme observed annual value. However due to limitation in CLASS, load-factors are only sampled once per reactor and kept constant during the whole life of the reactor. The parameters of the physics models have also been associated with uncertainties coming from the comparison of these simplified model with reference calculation. For example, the reactivity margin (threshold k) for UOX fuel has been associated with a normal distribution of average 1.034 and stdev of 0.005 coming from the result of [START_REF] Guillet | Prise en compte des calculs de coeur via l'utilisation de DONJON5 pour le code de scénario nucléaire CLASS et évaluation de l'impact sur les inventaires isotopiques[END_REF]. Uncertainties have also be associated with starting and stopping date of reactor to represent the fact that in reality, reactor do not start at full power from day one like we do in our CLASS models.

In total 260 of the 321 variables have been attributed uncertainties.

Visualization of operational parameter uncertainties

After the application of the package to extract and replace the variables with sampled one, 100 fuel cycle simulations have been done with the MLP physics models. The agglomeration of the result allowed to produce the figure 1 in which we can see the evolution of the plutonium inventory in the total cycle, in the storage for spent UOX fuels and spent MOX fuels. The middle line represent the average value and the colored area the standard deviation calculated on the 100 simulated trajectories and the total range observed in these trajectories. The figure shows that the uncertainty on the global plutonium inventory is small but the one for individual stock is far higher, up to 50% for the UOX spent fuel at some time. By looking at the value of an output at a set time, we get distributions that represent visually the level of knowledge extracted from the simulation. Outside of the average, standard deviation and limits that were also visible in figure 1, the distribution of figure 2 allows this way to see that the distribution is asymmetric and that the average is not the most probable value. The same tools providing easy access to uncertainties on plutonium inventories presented in the previous section can provide, by adjusting an option, the distribution of plutonium isotopic composition in various places. Figure 3 shows the agglomeration of such isotopic composition in the total cycle, in the storage for spent UOX fuels and spent MOX fuels. In this section, compared to previous figures, the times after 2025 are cut out because after this date the simulated fleet does not represent any realistic trajectory. Our scenario inspired by the Historic French fleet start to phase out, but no new reactors are build. For accumulation of inventories, this quick decrease in active nuclear power-plant had little impact because almost new spent fuel is created, but time without active reactor have an impact on isotopic composition of plutonium that is not of interest for fuel cycle analysis.

In the figure, we can see that after a couple of initial year during which the relative uncertainties are really high due to some different synchronization of the first spent fuel discharge, the uncertainties on isotopic composition averaged on all the plutonium present in the fuel cycle is very low, but when looking as the isotopic composition of the plutonium in specific spent fuel storages the uncertainty, even if it tends to decrease with time, stay high during the whole simulation. The uncertainty is specially high for the content of lighter plutonium isotopes with the standard deviation on the plutonium 239 in the plutonium inside the UOX spent fuels being close to 10% in 2020. This high uncertainty leads plutonium qualities that could go from 40 to 80%, a huge difference that would definitely impact which recycling strategies are applicable to these spent fuel or not. For plutonium in spent MOX fuel, uncertainties are slightly lower once the spent MOX fuel start to be available in reasonable number after 2010, but the standard deviation on the plutonium 239 and plutonium 240 in its plutonium is still higher than 5%, opening the way at very high (over 60%) and very low (under 40%) quality of plutonium. Designing a recycling strategy working for this kind of large range, would be really challenging, so reducing uncertainties on this aspect would really help future fuel cycle studies.

Inputs uncertainties with impact on plutonium-239 inventories

In the tools presented in this paper, several statistical tools are included to quickly and easily analyze the set of trajectory obtained exploring the uncertain values of inputs.

By calculating correlation between the change of each input and the change of one output of interest, Pearson coefficients have been determined help to determine the uncertainties of which input we need to decrease if we want to decrease on the output. An example of such result in table 1 which show the ten inputs (out of the 260 considered) contributing the most to the uncertainties on the total inventory of plutonium-239 in the whole cycle in 2020. In these output, association of city name with number represent French reactor. The city name is the city next to which the power plant site lies and the number is the place in chronological order of the construction of reactor in this power-plant. Fessenheim 2 is the second reactor built at Fessenheim power station. The error on correlation is here linked to the process of calculating the coefficient and is an algorithmic and statistical not, not an uncertainties like the the one presented in graphs elsewhere in this paper.

The MOX reactivity margin is used in our MLP-based models to compensate the fact only assembly-level simulation are used. In order to insure that the reactor could stay critical even with the neutron leakage that are not taken into account in these simulation, the model demands the capacity to maintain a multiplication factor slightly higher than 1, adding to it the MOX reactivity margin. A higher margin implies that more plutonium need to be loaded into the MOX assemblies, leading to an increase in plutonium burned. The uncertainty carried by this margin has thus logically an impact on the total plutonium-239 inventory in 2020

The MOX mass equivalence, is a compensation used in the current modeling of our scenario inspired by the historical French Fleet to adjust the total number of MOX assembly irradiated to take into account the fact this amount is lower than the initially planned one. It is lower than 1. The uncertainty carried by this factor directly impact the amount of MOX fuel loaded into reactor and such impacts the total plutonium-239 inventory in the cycle.

Uncertainties in cooling time will impact the minimum decay time of plutonium before being recycled. Plutonium with less decay time have higher quality and lead to lower amount loaded in MOX assemblies.

Load factor and starting date impact the amount of energy produced by the reactor because no reactor have reached the end of their lifetime in 2020.

All these links between inputs and the plutonium inventory can be expected, but their ranking is not obvious because of the interaction of the sensibility of the output to each input and the size of the uncertainty associated with it. The new tools give access to these value with minimal human work for any trajectories implemented with the CLASS code.

Impact of Missed loadings of MOX fuels

During the simulation of the fuel cycle, it happens that the simulation code is not able to build a MOX fuel according to fulfill the expressed needs. It usually happens when the quantity or the quality of the available plutonium is too low. These failure to build the wanted fuel lead to missed loadings of MOX fuels [START_REF] Thiollière | A methodology for performing sensitivity analysis in dynamic fuel cycle simulation studies applied to a PWR fleet simulated with the CLASS tool[END_REF].

Some of the trajectories simulated by our new tool to explore the space of uncertainty of the inputs have experience these kind of missed loadings. This happened in 19% of the explored space. This percentage does not means in any way that there were 19% of chance to have missed loadings in the historically planned French fleet and that the French nuclear sector took the bet. Indeed our scenario inspired by the historical French fleet consider uncertain a lot of factors that would be known by people managing a real nuclear fleet. Furthermore in a real fuel cycle, if no acceptable plutonium is available to build MOX fuel for a reactor, adaptation can be made, either by using mixes of plutonium or by using UOX assemblies, to ensure that the reactor is loading and is able to produce the electricity it should. However these adaptations lead to a difference in the effective MOX fraction and missed loading are an indicator to easily detect that.

When detecting missed loading, we could put constraint on input, and check which uncertainties we should reduce to ensure that the fuel cycle will be able to support the planned MOX assemblies. To do that we could use our tools and repeat analysis done on plutonium-239 and presented in the previous section but changing the output of interest to be the number of missed loading in the trajectory. The selection and representation of the data could be done as easily, however the statistical tools would need adaptation because they have been developed for continuous outputs and the number of missed loadings is a discrete one, specifically a discrete one where many trajectory will have the same value. Due to the need of tools adaptation, this study has yet to be done.

Another concern that these missed loading raise is the bias they can cause in our other and previous analysis. All analysis done before have been done using all trajectories, including those with missed loading and therefore not respecting the wanted power history and MOX fraction. To evaluate the bias that the inclusion of these particular trajectories in our analysis, we've redone it using only the 81% without missed loading and compared the result.

Most result are indistinguishable, specifically the graphs. How ever we observed some discrepancies in the evaluation of the impact of singular inputs that are presented in table 2. This table present the change in the correlations calculated with the two sets of trajectories, the one with all the trajectories and the one where the trajectories with missed loadings have been removed. The only change with a effect higher than three standard deviation is the change on the observed cooling time link with the plutonium-239 inventory, probably linked to an over selection of shorter cooling time in the set where the trajectories with missed loadings have been removed.

Visualization of the impact of model choice

To compare directly and visually the result of two while taking into account their uncertainties. In this section we use this opportunity to check is the result of [START_REF] Doligez | Advanced modeling of the historical French nuclear fleet with DONJON5/CLASS coupled simulations[END_REF] and the difference they saw between the two models are still significant when uncertainties are considered. We created a new input differing only by the use of DONJON-based physics models and uses the newly developed package with the same characterization of uncertainties to run 30 sampled fuel cycle simulations. And two sets of outputs have been compared with the visualization tools. Figure 4 present inventories of plutonium to result presented in figure 1 but with one color per model, blue for MLP and orange for DONJON. We see that for the global plutonium inventory, the evaluation of the average value and the range of uncertainties are really similar with both models. However for the inventories inside the stocks, even when taken into account the large uncertainties associated, the differences observed in [START_REF] Doligez | Advanced modeling of the historical French nuclear fleet with DONJON5/CLASS coupled simulations[END_REF] are significant.

CONCLUSIONS

Fuel cycle scenarios are "boundary objects" providing an opportunity to bring together different communities of stakeholders. These decision-maker can have various knowledge and different, and sometimes opposing, interests. Scenarios can help them to compare their visions for the future, organize their strategies and even cooperate. Uncertainties analysis is often difficult to discuss during this boundary interactions. New tools have been developed allowing calculating easily and represent uncertainties linked to our fuel cycle studies in a an easy to read fashion. It also allows representing evolution of uncertainties with times and full distribution of outputs. The application of this method to a scenario closy inspired by the historical French fleet allows showing limited impact of uncertainties on global output like global plutonium inventory, but a significant one for inventories in specific stocks like UOX or MOX spent fuel stockpiles. Used as a support to the analysis of the impact of a new model for reactor evolution, the method confirms the significant of results from previous studies, notably those for the global plutonium inventory, the evaluation of the average value and the range of uncertainties are really similar for the two studied models. However for the inventories inside the stocks, even when taken into account the large uncertainties associated, the differences observed are significant.

Figure 1 :Figure 2 :

 12 Figure 1: Representation of plutonium inventory evolution with the associated uncertainty Blue : Total Inventory, Orange : Inventory in UOX Spent Fuel storage, Green : Inventory in MOX Spent Fuel storage

( a )

 a Pu isotopy in the whole cycle (b) Pu isotopy in UOX spent fuels (c) Pu isotopy in MOX spent fuels

Figure 3 :

 3 Figure 3: Ratio of each plutonium isotopes to the global plutoniu minventory, for the full cycle, the stocks of spent UOX Fuel and the stock of spent MOX fuels Blue : Pu238, Orange : Pu239, Green Pu240, Red : Pu241, Lila : Pu242

( a )

 a Impact on total Pu inventory (b) Impact on Pu inventory in UOX spent fuels (c) Impact on Pu inventory in MOX spent fuels

Figure 4 :

 4 Figure 4: Impact of model choice on plutonium inventory in stocks with the associated uncertainty Blue : using MLP Assembly-based models, Orange : Using DONJON core-based models

Table 1 :

 1 Correlation between change of values in inputs within their uncertainties and change in total plutonium-239 inventory in 2020 : 10 most impactfull inputs Input Pearson Coeff. Error on Pearson Coeff.

	MOX reactivty margin	-0.71	2E-16
	Cooling Time	-0.33	7E-4
	MOX mass equivalence	-0.31	2E-3
	Load factor of Bugey 4	0.28	4E-3
	Start of Bugey 4	-0.28	0.01
	Start of Fessenheim 1	0.26	0.01
	Load Factor of Paluel 3	0.25	0.01
	Start of Paluel 2	0.24	0.02
	Start of moxing at Dampierre 3	-0.23	0.02
	Start of Blayais 1	-0.22	0.03

Table 2 :

 2 Change in calculated correlation depending of inclusion or exclusion of trajectories with missed loadings (only changes with error on the change of correlation correlation 100%)

	Input	Change of Pearson Coeff. Error on the change
	Cooling Time	-0.12	0.05
	Start of Paluel 2	-0.07	0.02
	Start of Blayais 1	0.05	0.04
	Load Factor of Bugey 4	0.05	0.04
	Load Factor of Paluel 3	-0.04	0.02
	Start of Blayais 4	-0.03	0.03
	MOX mass equivalence	-0.02	0.01

ACKNOWLEDGEMENTS

We would like to express our gratitude to the NEEDS program of the Energy Unit of the CNRS for the financial support through the CINEASTE project. We also thank Dr. Baptiste Mouginot, for his major work as the main developer of the fuel cycle simulator CLASS