Ali Bakiri

Bertrand Plainchont

Vicente De

Paulo Emerenciano

Romain Reynaud

Jane Hubert

Jean- Hugues Renault

Jean-Marc Nuzillard
email: jm.nuzillard@univ-reims.fr

Keywords: Artificial Intelligence, Computer-Assisted Structure Elucidation, Dereplication, Liquid Chromatography, NMR Spectroscopy [

Natural product chemistry began in Reims, France, in a pharmacognosy research laboratory whose main emphasis was the isolation and identification of bioactive molecules, following the guidelines of chemotaxonomy. The structure elucidation of new compounds of steadily increasing complexity favored the emergence of methodological work in nuclear magnetic resonance. As a result, our group was the first to report the use of proton-detected heteronuclear chemical shift correlation spectra for the computer-assisted structure elucidation of small organic molecules driven by atom proximity relationships and without relying on databases. The early detection of known compounds appeared as a necessity in order to deal more efficiently with complex plant extracts. This goal was reached by an original combination of mixture fractionation by centrifugal partition chromatography, analysis by 13 C NMR, digital data reduction and alignment, hierarchical data clustering, and computer database search.

Introduction

The chemistry research teams at the University of Reims have developed analytical skills for the structure identification of small synthetic and natural organic molecules during the last four decades. Natural product chemistry has evolved from the study of indole alkaloid-containing plants, [START_REF] Sapi | Monoterpenoid Indole Alkaloids[END_REF] following the discovery of the bis-indole anti-cancer drugs such as vinblastine. In the '80s, the isolation, purification and structure analysis of saponins encouraged the acquisition of higherperformance NMR instrumentation as well as the implementation of the then most recent NMR spectroscopy methods. [START_REF] Massiot | [END_REF][START_REF] Massiot | Structure elucidation of plant secondary products. Chemical from plants[END_REF] At the same time, the investigation of other natural product classes such as terpenes and phenolic compounds took a growing importance in our laboratory.

The general approach to the structure elucidation of plant secondary metabolites involved the determination of a gross molecular formula using mass spectrometry (MS) and the atom assembly under constraint of NMR, UV, IR spectral features as well as MS fragmentation. Working in compound series such as indolomonoterpenic alkaloids offered the availability of a vast corpus of physicochemical data, including the coloration of compounds upon spraying of specific reagents on Thin Layer Chromatography (TLC) plates. The quick identification of the compounds that were already reported in the literature was carried out with great reliability by the comparison with archived data of TLC chromatographic Rf values, coloration on TLC plates and low-resolution electron impact mass spectra. A 1 H NMR spectrum could help to secure the certainty of an identification, especially when alternative structures existed due to position isomerism. The structure of the few isolated compounds that were not readily identified by these means was determined by de novo structure determination.

Structure elucidation by NMR underwent a radical change in the 1980's with the availability of 1 H-detected 2D heteronuclear (meaning 1 H-13 C or 1 H-15 N) chemical shift correlation spectra, namely HMQC (and later HSQC) for directly bound nuclei pairs and HMBC for remotely bound nuclei pairs. [START_REF] Mueller | [END_REF] Pulsed static field gradient enhancement of these techniques delivered high-quality 2D NMR spectra that were well suited to the de novo computer-assisted structure elucidation. [5] The use of computers to deal with molecular structures and NMR spectra was initiated at the University of Reims in 1988 by the corresponding author of this article. As an occasional teacher of computer programming at the Reims Institute of Technology, he attended an introductory course on Artificial Intelligence (AI) and on the Prolog (Programmation en Logique) AI language. The newly acquired knowledge was quickly put into action for the design of CASA, an automated system for 13 C NMR spectra assignment whose goal was to pair spectral resonances with the atoms of a known or postulated structure. [6] The conceptual gap between spectral assignment and structure generation was then quickly bridged, still using the Prolog AI language as programming tool, and led to the LSD (Logic for Structure Determination) software. [7] The computer code was later rewritten in the C language for speed and portability reasons and is available as free software (www.univ-reims.fr/LSD).

CASA and LSD were designed for the analysis of pure compounds. Purification is a mandatory step for organic chemists. It is nearly impossible to publish the structure of a new compound without its isolation at the highest possible purity level. The thorough purification of the compounds of an extract requires an important investment in equipment and human time. Therefore, the early identification of known compounds, a task also known as dereplication, when carried out in mixtures, is a way to save time and to concentrate on the structure elucidation of new compounds. Recent developments in this field led to the creation of the workflow called CARAMEL (after CARActérisation de MÉLanges, in French, meaning mixture characterization), which combines spectroscopic, data classification, and databasing tools. [8] 2 Dereplication

Natural product (NP) chemists often feel the unpleasant feeling of rediscovering compounds that have already been discovered. However, the study of the chemical content of an extract (from plant, bacteria, mycete, insect...) generally starts with the exact determination of the species of the studied organism and with bibliographic work on what has been already published about the same species or the same genus or even the same family, depending on the number of related species. The exploitation of such chemical studies allows one to define a list of compounds that could be present in the extract under investigation because a taxonomic proximity is presumably correlated with a chemical proximity. In a formal way, bibliography established a relationship between the species space and the molecular structure space (Figure 1). A given species is related to a set of molecular structures (pharmacognosy) and a given molecular structure is related to a set of species (reverse pharmacognosy). [9] The commonly adopted workflow in NP chemistry involves the isolation of the compounds from organism extracts. Extraction might be more or less selective and/or carried out on particular parts of organisms (such as barks or flowers in plants), when possible, in order to simplify compound isolation and possibly to relate organs and their function. A compound is generally purified by a NP chemist in order to study its biological activity. In the same way that a plant specimen needs to be related to the name of a living species, an isolated compound needs to be related to a molecular structure in order to give value to a biological test.

Dereplication, in one of the acceptations of the term, consists in finding the molecular structure of an isolated compound, pure or in mixture, among those already reported. With this meaning, dereplication is the way to assign a chemical structure to a compound using the knowledge that accumulated during decades of NP research. The main difficulty here is to declare that a sample that has just been isolated in the lab next door is made of the same molecules as the one isolated thirty years ago in a lab on another continent. Physico-chemical analysis is the way to do it, by connecting the space of physical samples and that of analytical data. A relationship between the latter and the space of molecular structures is needed in order to connect isolated compounds with their molecular structure (Figure 1).

Two samples with identical analytical data are presumed to be identical, assuming that identical compounds present identical analytical data. The last part of this assertion is subjected to the repeatability of the analytical process and on the identity of the samples, while the first part really depends on the concept of analytical data. Leaving this question aside for the moment, dereplication boils down to acquisition of analytical data and to comparison between actual and published analytical data, a task that seems easy and instantaneously feasible across time and continents in the era of the Internet. [10] Figure 1. Relationships between different spaces involved in natural product chemistry. The three boxes enclosed by the dotted contour line correspond to physical objects and the others to intellectual objects. Relationship A connects species and molecule structures and is reported in publications whose t itles are like "New sesquiterpene lactones from the flowers of Genusname species-name". Relationship B connects species and biological data as reported in publications with titles like "Biological Activities of Aerial Parts Extracts of Genus-name species-name". Many NP publications belong to the A and to the B type simultaneously, reporting structure and activity of plant parts.

Chemistry journals and publishers require authors to deposit analytical data as Supplementary Information (SI) documents attached to articles, while the visible part of analytical studies most often reduces to lists of analytical parameters. For example, a UV-visible spectrum, acquired through the measurement of hundreds of light absorption values (the analytical data) ends up in a publication as a list of a few maximum absorbance positions (the analytical parameters). Therefore, effective dereplication is carried out by comparison between actual and published analytical parameters. Comparison between spectra is sometimes possible but the plots in most SI documents are of too poor quality to be really useful, a situation that could change with the development of infrastructures for public storage of spectral data and spectral parameters.

Dereplication clearly depends on analytical methods. Fusion temperature might be useful to declare that two samples are made of different molecules (not considering sample purity and the role of molecular packing in crystals), but is hardly convincing as a validation tool for structure identity. Intuitively, combining analytical methods that rely on unrelated physico-chemical processes increases the probability of being sure that two substances are identical. From a practical point of view, it is not possible for a laboratory to invoke all existing methods, and a single method is often selected.

Mass spectrometry (MS) and Nuclear Magnetic Resonance (NMR) are the two main analytical methods that come into play for dereplication purposes, either in NP chemistry or in metabolomics studies. [11] The pros and cons of each technique are known and, to summarize, NMR is poorly sensitive but is quantitative, reproducible and reveals the finest structural details, while MS is utterly sensitive, but hardly reproducible and gives limited access to structural features. No single technique presently combines the advantages of NMR and MS, even though each of them is still the object of steady improvement efforts in order to reach the advantages of the other.

Dereplication of NPs is commonly carried out all over the world using a combination of High-Performance Liquid Chromatography (HPLC, briefly LC) and MS, thus allowing in one LC-MS sequence the isolation of tiny amounts of pure compounds (nanograms, if not picograms) and measurement of their mass spectra. In particular research fields such as marine products and micro-organism chemistry, specialized databases exist and make it possible to propose molecular structures from mass spectral data. The concept of molecular networking in MS tends to increase its analytical power toward structure elucidation, beyond simple structure recognition.

The emergence of our original approach toward dereplication in Reims is the result of many factors. i) A long tradition of NP studies exists there, in which NMR played a decisive role and was the subject of methodological research in the fields of data acquisition and processing (JMN). ii) LC-MS have become available and open to NP research only recently. iii) Preparative separation methods based on centrifugal partition chromatography (CPC) has undergone tremendous development during the last 25 years (JHR). CPC offers the means to isolate pure compounds or simplified fractions in amounts that are compatible with analysis by 13 C NMR spectroscopy and biological activity research (milligrams to tens of grams on a laboratory scale CPC system). iv) An assistant professor (JH) with a background in metabolomics and statistical data analysis was recruited about six years ago. v) Our university and an international company found good reasons to support our research efforts (RR). vi) People there decided to work together.

The CARAMEL dereplication workflow deals with complex mixtures such as plant or microbial extracts (but not limited to these) and starts with extract fractionation by CPC. About 20 simplified fractions are obtained and analysed by 13 C NMR (discussion hereafter). The spectra, recorded over a 240 ppm wide chemical shift window, are submitted to an automatic peak search and therefore reduced to a list of peak coordinates. The peak intensity values are stored in 0.2 ppm wide, initially empty, chemical shift intervals called bins, so that each one corresponds to a chemical shift value. The bins that are empty over the whole series of spectra are left aside. The content of the bins is stored in a table: bin index in rows, fraction index in columns, peak intensities for values in table. The table is then converted into a human-readable text format. Considering two molecules A and B with nonoverlapping peaks and for which the CPC emergence profiles are different, for example with A less retained in the CPC column than B. Considering all the bins of compounds A and B, the content of some of them will have non zero values only for low-index fractions, others for high-index fractions only. The former will correspond to compound A and the latter to compound B. In this simple trivial context, the bins that correspond to compounds A and B are easily differentiated (see Figure 2). A way to automate this process in real-life situations is to group the lines of the table according to similarity. The classification of rows by hierarchical clustering achieves this goal and is implemented using the freely available PermutMatrix software. [12] The textual table of peak intensities is used as software input. The resulting rearranged table is organized into contiguous blocks (or clusters) of bins whose associated chemical shift values correspond to the different compounds that constitute the initial complex mixture. The association between chemical shift lists and molecular structures is achieved by database search. We filled our own database (approximatively 2100 entries at manuscript submission time) with molecular structures collected through the bibliographic work carried out at the beginning of each new extract analysis project. Initially, molecular structures were associated in our database with published experimental chemical shift values. It turned out then that it was equally efficient to store the values calculated by the ACD 13 C NMR predictor. [13] The ACD databasing system also allows one to retrieve compounds from an incomplete list of chemical shift values. The discrepancy between ACD-calculated and experimental values and the search for a compound in an incomplete list of values results in a series of candidate structures that has to be checked manually. Checking is carried out by inspection of 1D 1 H, 1 H-13 C, 2D 1 H-1 H COSY, 2D 1 H-13 C HSQC and HMBC spectra of the fraction(s) in which the 13 C NMR peaks are the most intense and provides a molecular structure that is the most likely. Even though checking is still a mandatory step, the whole process greatly speeds up the search for the structure of known molecules. Additionally, the initial fractionation step delivers samples that may be involved in biological activity tests and further purified when a fraction shows some interesting activity.

The present version of the CARAMEL workflow is based on 1D 13 C NMR spectroscopy. This choice results from practical considerations. i) Even extremely hydrogen-poor compounds (such as ellagic acid and its derivatives) contain carbon atoms. Apart from 13 C NMR spectroscopy, the only practical way to obtain information on quaternary carbons is the recording of 2D HMBC spectra. ii) The 13 C resonances appear in our standard 1 H decoupled spectra as sharp singlets (typically 1 Hz width at half height) within a 36 kHz spectral window (240 ppm from a base frequency of 150 MHz), thus minimizing the probability of peak superimposition. iii) Spectral parameter extraction is easily carried out by automatic peak peaking using the spectrometer software and the result is easily exported as a text file. iv) Apart from (rare) symmetrical molecular structures and (rare) coincidences, there is a one-to-one relationship between carbon positions in molecular structures and chemical shift values revealed as peak positions. v) We compensate the commonly mentioned lack of sensitivity of 13 C NMR by the use of a high static magnetic field (14.1 T, resonance of 1 H nuclei at 600 MHz) and of a Helium-cooled cryo-probe. Spectrum acquisition time is kept under these conditions around one hour for each fraction, during which 1024 transients are co-added, with a 3.6 s repetition time. vi) Prediction of 13 C NMR chemical shifts is more reliable than the one of 1 H, for which variations of experimental conditions may induce chemical shift variations greater than the expected uncertainty on predicted values.

At this stage, one can see that the CARAMEL workflow has some obvious limitations. Two compounds with identical emergence profiles are of course not distinguishable, but this situation is not likely. Compounds in small amounts for which spectral peaks do not significantly emerge from noise are not visible, so CARAMEL misses the minor components of a mixture. Compounds for which 13 C NMR spectra are partly superimposed (collected in the same bin) may be left classified together but not with the other nonsuperimposed peaks, resulting in incomplete chemical shift lists for the concerned compounds. Figure 3 illustrates the CARAMEL procedure on a published example. [14a] Possible improvements of the analytical part of the CARAMEL workflow may include the spectral edition of 13 C resonances according to the parity of the number of attached 1 H nuclei (DEPT spectra), 1 H pure shift spectra, 2D HSQC (pure shift or not) and HMBC spectra. Numerous applications of the CARAMEL workflow have already been reported. [14] A manuscript reporting the direct search for the compounds stored in our database within a crude extract, without fractionation, and on the sole basis of 13 C NMR data, is currently under review.

Structure elucidation by the LSD software

First, and as a rule of thumb: "dereplication precedes structure elucidation". Structure elucidation of new or presumably new compounds is at best performed on pure compounds because this is the best way to avoid spectral data misinterpretation due to confusion between data arising from different compounds. The following lines will assume compound purity.

Computer-assisted structure elucidation (CASE) has been one of the first playgrounds of artificial intelligence. CASE concepts, software and their application have been reviewed in articles, book chapters and a recent book. [15] Structure elucidation is the process of establishing bonds between atoms whose number and individual chemical elements are obtained as a gross formula by high resolution mass spectrometry. Hydrogen atoms play a particular role in molecular structures because they are all the same and always share a single bond with any other atom. As a consequence, chemical structure drawings of organic molecules omit them when they are bound to carbon atoms unless some particular stereochemical feature needs to be displayed. The LSD software (in short: LSD) differentiates between light atoms (all the hydrogens) and heavy atoms (all the others) by describing only the latter by an explicit atom status.

An atom status consists of an atom index, a chemical element symbol, a multiplicity, a hybridization state, and an electric charge. Carbon indexes are, by tradition, assigned to atoms in the decreasing order of the corresponding 13 C chemical shift values. Atoms other than hydrogens and carbons, referred to as heteroatoms, receive arbitrarily given atom indexes. All parts of the status of all atoms must be completely defined at the beginning of the resolution of a problem by LSD. The status defines for each heavy atom the number of directly attached heavy atoms and this number must be accurately known in order to efficiently carry out structure generation. A chemical element symbol may include a valence suffix for non-usual valence values such as S4 for tetravalent sulfur, while S is sufficient for divalent sulfur. The multiplicity of a heavy atom is the number of directly attached hydrogen atoms; this is easily determined for carbons and for heteroatoms when only a single kind of the latter is present. Hybridization states are only sp 3 , sp 2 and sp. For commodity, the sulfur atom of a sulfoxide is declared as sp 2 , even though this does not correspond to the actual bond electronic structure. This example touches the border of the usual "atom and bond" representation of organic molecules that excludes bonds over more than two atoms. Hybridization state of carbons may be unambiguously deduced as being sp 2 or sp 3 from 13 C chemical shift values when they fall in high-value or low-value ranges.

In some cases, different atom status sets are compatible with the available data and more than one data set must be prepared for LSD in order to solve a single problem. This inconvenience is eliminated in pyLSD, but computer system compatibility issues still must be solved in order to make its use of general interest. PyLSD is a driver software for LSD written in the Python programming language that expands all alternatives on individual atom status and retains the status sets compatible with the molecular formula. PyLSD also allows the user to introduce some flexibility in the gross molecular formula of the unknown molecule. [16] Ideally, 13 C-13 C INADEQUATE spectra should be able to delineate the carbon skeleton of an organic molecule, thus simplifying a big part of structure elucidation problems. [17] This approach was considered in the first Computer-Aided Structure Elucidation programs in which 2D NMR was taken into account. [18] The insensitivity of INADEQUATE, that requires neighbouring pairs of 13 C nuclei while the natural abundance of the nuclei is only about 1%, makes it rarely applicable, even though this is the only way to directly prove by NMR that two quaternary carbon atoms are bound together. LSD accepts INADEQUATE data as predefined bonds between atoms. All other forms of atom connectivity involve hydrogen atoms and their 1 H NMR resonances.

Hydrogen atom indexing is preferentially related to heavy atom indexing, according to the HSQC spectrum that correlates the chemical shifts of 1 H and 13 C (or 15 N) nuclei that are directly bound, thus defining with a single index what chemists call a chemical position (C-1 is bound to H-1 at position 1). Inequivalent hydrogens in methylene groups are therefore identically indexed. The COSY and HMBC spectra provide proximity relationships that LSD exploits in order to set bonds between atoms. In the simplest vision, a 1 H-1 H COSY spectrum reveals magnetic couplings between 1 H nuclei through 2 or 3 bonds. Twobond (or 2 J) couplings only occur within methylene groups with inequivalent hydrogen atoms. The 2 J COSY couplings are quickly identified because they happen between identically indexed atoms; they do not bring any information about heavy atom connectivity. A 3 J H-a/H-b coupling arises through the H-a/C-a/C-b/H-b coupling path and therefore proves the existence of the C-a/C-b bond that is translated by LSD into the setting of a firmly established, predefined bond (Figure 4). A HMBC spectrum reveals 2 J or 3 J couplings between 1 H and 13 C (or 15 N) nuclei, thus proving that the corresponding heavy atoms are either directly bound or two bonds away, as if a 1 J or 2 J 13 C-13 C INADEQUATE spectrum were recorded. [7] There is no general and reliable method to decide upon which alternative is correct. [19] The intermediate heavy atom, see Figure 4, in the 3 J hypothesis can be any atom in the molecule, either a carbon or a heteroatom.

Low resolution along the 13 C chemical shift axis adds a supplementary indeterminacy level in the interpretation of an HMBC spectral peak because more than a single carbon atom may be responsible for it. LSD offers the possibility of handling such alternatives on close resonance groups. Resolution enhancement in HMBC may be obtained without increasing recording times by means of strategies such as folding or non-uniform sampling. [20] Groups of poorly distinguishable 1 H resonances may also be exploited by LSD.

Writing datasets for LSD implies that 2D spectra may adequately be reduced, a problem that remains difficult to automate. Results, as chemical shift pair lists, still need to be edited by hand in order to avoid the introduction of contradictory input data in LSD input files that would lead to empty solution lists. Efforts for effective reduction of 2D NMR data have been carried out and were integrated into the CMC-se module of the commercial Bruker TopSpin software. CMC-se integrates a company-developed structure elucidation algorithm as well as that of LSD. [21] The length of the coupling paths revealed by COSY and HMBC spectra is not limited to 2 and 3, even though the initial algorithm was designed as if it were the case. Longer length COSY and HMBC data caused LSD to fail. They originate from coupling constants of low magnitude and give rise to low intensity peaks. A good practice consists in considering first only the most intense 2D peaks. LSD was later modified so that a limited userdefined number of long range couplings may be handled. Additionally, each individual HMBC may be associated with a minimum and a maximum coupling path length.

LSD was designed to mimic the way chemists build structures from spectra. Chemical shift values and 1 H NMR line splitting patterns are related to immediate environments of atoms and therefore LSD accepts constraints, called "atom properties", on the nature of the neighbouring atoms a given heavy atom may have. Atom properties are taken into account by the LSD algorithm during structure generation. Of course, invalid constraints may result in incorrect structures or in no structure at all. User-provided substructure elements may also be given and are used for the solution filtering step that takes place after structure generation.

Natural products may be categorized in classes that correspond to various skeletons, another word for substructures. Their identification was one of the motivations for starting a collection that is included in LSD. The present collection was drawn from the SISTEMAT knowledge base and was established for mono-, sesqui-, and diterpenes. SISTEMAT contains databases (50000 compounds, overall) for various natural product classes and software dedicated to their management and exploitation. A SISTEMAT database contains records for natural products as reported in literature. A record connects the molecular structure of a compound with its spectroscopic data, including those of 13 C NMR, and with botanical data on the plant from which it originates. In this way a species may be immediately related to the probability of the presence of the skeletons of the compounds that were isolated in plants of this species. [22] SISTEMAT may be used for dereplication from 13 C NMR data of mixture. [23] It can also assist elucidation by proposing fragments of known molecules as possible fragments of an unknown molecule. [24] Structure generation by LSD has been successfully guided by SISTEMAT proposals for natural compounds from various classes. [25] The structure generation process in LSD starts (phase 1) with a set of well-defined status atoms connected by explicitly user-supplied bonds and by those arising for 3 J COSY data ( 2 J COSY data is automatically discarded). Atom proximity information from HMBC is then selected and exploited in order to build bonds. The selectionexploitation process (phase 2) is repeated until all the proximity information is processed. The order in which proximity data are taken into account greatly impacts the computation times. A heuristic criterion was implemented, stating that the heavy atoms for which the number of missing bonds is the lowest must have their proximity data exploited first. Atoms for which bonds are missing at the end of phase 2 are systematically paired to produce a meaningful molecular structure during phase 3. Phase 4 includes a set of acceptance tests comprising the determination of the order of bonds, the compliance of structures to Bredt's rule, [26] and the validation of substructure constraints, if any.

The exploitation of proximity data in step 2 involves the selection of a hypothesis in the case of close resonance groups, the choice between a one-bond and a two-bond distance between heavy atoms, and, in the last case, the choice of the index of the intermediate atom.

The solution search process has therefore been organized as a depth-first tree exploration algorithm (Figure 5). [27] The resulting solutions are stored in a text file, from which conversion to various standard formats is carried out by a program named outlsd. The depictions in 2D SD files are most often very crude and can be manually improved using the m_edit utility program or any other multi-structure 2D structure editor, or regenerated from scratch using any available structure drawing generator and depictors, like those from the RDKit, [START_REF]RDKit: Cheminformatics and Machine Learning Software[END_REF] OpenBabel, [START_REF] O'boyle | [END_REF] and CDK [30] chemoinformatic toolkits. The final goal of the structure elucidation process is to find the structure of a molecule. The generation of more than one solution, all compatible with input data, sets the problem of solution ranking. Ranking is provided in pyLSD by a score calculated as a distance between the set of the experimental 13 C NMR chemical shifts and those predicted for each solution structure. The reliability of this approach naturally depends on the quality of the prediction algorithm. The only standalone predictor we had at hand was the one provided by the nmrshiftdb2 free software project. [31] The LSD software played an important role in the structure of natural and sometimes synthetic compounds, as exemplified by published articles (see Figure 6). [32] Future developments of LSD include a better separation between structure generation and validation/filtering/ranking tasks. The structure generator should be modified so that atoms with alternative status are handled during resolution and not before it, as presently achieved in pyLSD for the combinatorial enumeration of status indetermination-free LSD problems. The application of substructure constraints should also be integrated in the bond-building processes, if desired. Molecules that contain parts with symmetry or even symmetrical molecules are not presently easily handled by LSD and solving this problem would enlarge the capabilities of the software. Tasks such as substructure search and depiction generation should be carried out by thoroughly tested free chemoinformatic toolkits. The efficiency of dereplication and of automatically generated structure ranking should improve through the public availability of an increasing amount of experimental chemical shift data, in a format that may facilitate their transfer from laboratory notebooks to free public databases. [10]

Figure 2 .

 2 Figure 2. Principle of data handling in the CARAMEL dereplication workflow. The 13 C NMR spectra of the fractions contain seven peaks. The evolution of intensity for the different peaks characterized by their bin index (line index in data table, see text) is reported as a function of retention time or fraction index (column index in data table). a) original data, b) rearranged data that permit the appearance of the grouping of the resonances arising from two compounds with different chromatographic behaviours.

Figure 3 .

 3 Figure 3. Dereplication of an extract of Tephrosia purpurea, as reported in reference 14a. The PermutMatrix software represents data as heat maps, the brightest colors corresponding to the highest spectral peak intensities. (left) Data before hierarchical clustering. (right) Result of peak clustering according to similarity of fractionation profiles. Each cluster (green ellipses) corresponds to a chemical shift list that serves as key for database search. The structure elucidation of compounds D, H and J, unknown to the database, was assisted by the LSD software.

Figure 4 .

 4 Figure 4. Deduction of bonds and of alternative proximity relationships from 2D HSQC, COSY, and HMBC NMR data. In the case of a 3 J (H-X) coupling, the intermediate atom noted as a question mark may be any atom of the studied molecule.

Figure 5 .

 5 Figure 5. Solution search by depth-first tree exploration. The analysis of a correlation potentially opens a range of interpretation choices. An invalid interpretation may lead to the impossibility of interpreting a still-unexploited correlation (failure).

Figure 6 .

 6 Figure 6. Examples of molecular structures, whose determination was assisted by the LSD software.

Acknowledgements

Financial support by CNRS, Conseil Regional Champagne Ardenne, Conseil Général de la Marne, French Ministry of Higher Education and Research (MESR), FAPESP, CNPq, the Soliance-Givaudan Company and EU-programme FEDER to the PlAneT CPER project is gratefully acknowledged. We thank Dr. Mark Andrew Gannon for his stylistic and grammatical improvement of the manuscript.

Full Paper

C. Author et al.


S Baraka

K Bouearan

L Caner

C Fontaine

F Epron

R Brahmi

N Bion

Catalytic performances of natural Ni-bearing clay minerals for production of syngas from dry reforming of methane

Keywords: Dry Reforming of Methane, Clay, Smectites, Nickel, Biogas, Iron

The catalytic activity of Ni-bearing clay minerals obtained from natural nickel lateritic ores from Niquelândia, Brazil was evaluated in methane dry reforming reaction at 800°C. These phyllosilicates materials are Ni/Mg smectites with a Ni content of about 23 wt %. The mineralogy and the crystallochemistry of the clay minerals were characterized using X-Ray diffraction and FTIR spectroscopy respectively. Prior to any catalytic test, the clay fraction was extracted from the lateritic ore to concentrate the smectites and remove primary minerals (quartz, pyroxenes) or impurities. In these samples, the Ni is localized in the octahedral layers (structural Ni) and in the interlayer space (exchangeable Ni). To evidence the impact of octahedral or exchangeable Ni, the clays were saturated either with Mg or with Ni. The activity in methane dry reforming is higher with the clay minerals saturated with Ni compared to those saturated with Mg. These results support the hypothesis that both octahedral and exchangeable Ni play a role in the conversion of CH 4 and CO 2 into H 2 and CO. Finally, in situ XRD and TEM analyses were crucial to explain the strong influence of the reduction temperature on the H 2 yield. In these natural nickel lateritic ores, the presence of Fe leads to strong interaction with Ni during the reduction step, which depending on its temperature can result in the loss of catalytic activity.

Introduction

Methane dry reforming (MDR) is a widely studied reaction that is of both scientific and industrial importance as one of the most promising routes for biogas valorization because of the economic and environmental benefits of this process [START_REF] Pakhare | A review of dry (CO2) reforming of methane over noble metal catalysts[END_REF][START_REF] Gao | A review of recent developments in hydrogen production via biogas dry reforming[END_REF][START_REF] Wang | Low-temperature catalytic CO2 dry reforming of methane on Ni-based catalysts: A review[END_REF]. It involves the consumption of two important greenhouse gases, methane and carbon dioxide following the Eq. [START_REF] Pakhare | A review of dry (CO2) reforming of methane over noble metal catalysts[END_REF]. MDR results in the formation of syngas (CO + H 2 gas mixture) having a low H 2 /CO ~1 ratio, a resource for high added-value products such as long chain hydrocarbon through Fischer-Tropsch (F-T) reaction [START_REF] Dry | The Fischer-Tropsch process: 1950-2000[END_REF] or oxygenated chemicals like acetic acid [START_REF] Knifton | Syngas reactions: IX. Acetic acid from synthesis gas[END_REF]. MDR is highly endothermic (ΔH 0 298K = +247 kJ.mol -1 ) and therefore thermodynamically favorable only at high temperatures. Production of syngas by MDR is also governed by the parallel reverse water-gas shift reaction (Eq. ( 2)), which consumes part of the produced H 2 resulting in a decrease of the H 2 /CO ratio. In the appropriate temperature range, MDR reaction is finally affected by competing side reactions, such as methane decomposition (Eq. ( 3)) and Boudouard reaction (Eq. ( 4)) which both promote carbon formation and deposition on catalyst surface. The high temperature required to obtain high syngas yield is also responsible for the sintering of the active site and/or the support [START_REF] Xu | One-Pot Synthesis of Ordered Mesoporous NiO-CaO-Al2O3 Composite Oxides for Catalyzing CO2 Reforming of CH4[END_REF][START_REF] Bradford | CO2 Reforming of CH4[END_REF][START_REF] Yu | The promoting role of Ag in Ni-CeO2 catalyzed CH4-CO2 dry reforming reaction[END_REF][START_REF] Paksoy | A study on characterization and methane dry reforming performance of Co-Ce/ZrO2 catalyst[END_REF][START_REF] Aramouni | Catalyst design for dry reforming of methane: Analysis review[END_REF][START_REF] Marinho | Highly active and stable Ni dispersed on mesoporous CeO2-Al2O3 catalysts for production of syngas by dry reforming of methane[END_REF]. MDR was widely studied on supported noble metal (Ru, Rh, Pd, Pt, Ir) catalysts, which are active and generally resistant to the formation of carbon, but their high costs restrict their industrial application [START_REF] Pakhare | A review of dry (CO2) reforming of methane over noble metal catalysts[END_REF][START_REF] Gao | A review of recent developments in hydrogen production via biogas dry reforming[END_REF][START_REF] Drif | Study of the dry reforming of methane and ethanol using Rh catalysts supported on doped alumina[END_REF][START_REF] Marinho | Pt nanoparticles embedded in CeO2 and CeZrO2 catalysts for biogas upgrading: Investigation on carbon removal mechanism by oxygen isotopic exchange and DRIFTS[END_REF][START_REF] Rezaei | Nanocrystalline zirconia as support for nickel catalyst in methane reforming with CO2[END_REF][START_REF] Hou | Production of synthesis gas via methane reforming with CO2 on noble metals and small amount of noble-(Rh-) promoted Ni catalysts[END_REF]. Non-noble metal (Ni, Mg, Co, Fe…) catalysts have therefore concentrated much attention as low-cost materials in the last decade. Ni-containing catalysts are of particular interest from the industrial point of view, due to the low cost, availability and high activity of Ni, but suffer the critical problem of the deactivation during the reaction due to carbon deposition and sintering of the active components under high temperature [START_REF] Aramouni | Catalyst design for dry reforming of methane: Analysis review[END_REF][START_REF] Fang | Methane dry reforming over Ni/Mg-Al-O: On the significant promotional effects of rare earth Ce and Nd metal oxides[END_REF][START_REF] Marinho | Embedded Ni nanoparticles in CeZrO2 as stable catalyst for dry reforming of methane[END_REF][START_REF] Shi | A review of different catalytic systems for dry reforming of methane: Conventional catalysis-alone and plasma-catalytic system[END_REF][START_REF] Akri | Atomically dispersed nickel as coke-resistant active sites for methane dry reforming[END_REF][START_REF] Ha | Development of Active and Stable Low Nickel Content Catalysts for Dry Reforming of Methane[END_REF].

To improve the catalytic performances and to overcome the sintering of nickel nanoparticles and carbon deposits, the influence of the supports was deeply investigated [START_REF] Gao | A review of recent developments in hydrogen production via biogas dry reforming[END_REF][START_REF] Akri | Highly Active and Carbon-Resistant Nickel Single-Atom Catalysts for Methane Dry Reforming[END_REF]. In addition to good thermal stability and mechanical resistance, the support must have low acidic properties to avoid the deposition of carbon which leads to deactivation of the catalyst [START_REF] Yu | The promoting role of Ag in Ni-CeO2 catalyzed CH4-CO2 dry reforming reaction[END_REF][START_REF] Roger | CHAPTER 1 Design of Advanced Catalysts for Natural Gas Reforming Reactions[END_REF][START_REF] Zhou | A single source method to generate Ru-Ni-MgO catalysts for methane dry reforming and the kinetic effect of Ru on carbon deposition and gasification[END_REF][START_REF] Fan | Tuning the composition of metastable COxNiyMg100-x-y(OH)(OCH3) nanoplates for optimizing robust methane dry reforming catalyst[END_REF][START_REF] Li | Loading Ni/La2O3 on SiO2 for CO methanation from syngas[END_REF][START_REF] Li | Dry reforming of methane over Ni/La2O3 nanorod catalysts with stabilized Ni nanoparticles[END_REF][START_REF] Lovell | Enhancing Ni-SiO2 catalysts for the carbon dioxide reforming of methane: Reduction-oxidation-reduction pre-treatment[END_REF]. MgO, CaO and CeO 2 rank among the best supports to prevent carbon formation as confirmed by the study of Yamazaki et al. [START_REF] Yamazaki | Reduction of by Methane with Nion-Mgo-Cao Containing Catalysts[END_REF] on the Ni/MgO-CaO catalyst which reported that carbon deposition can be avoided by using metal oxides with Lewis basicity as supports.

Among the materials containing both Ni and basic properties, the clay minerals appear as an attractive option for MDR reaction. Such materials were already investigated in green and sustainable catalysis processes because they are naturally available, green, and inexpensive [START_REF] Lafaye | Impact of cerium-based support oxides in catalytic wet air oxidation: Conflicting role of redox and acid-base properties[END_REF][START_REF] Hadjltaief | Efficient removal of cadmium and 2-chlorophenol in aqueous systems by natural clay: Adsorption and photo-Fenton degradation processes[END_REF][START_REF] Hao | Zr-Laponite pillared clay-based nickel catalysts for methane reforming with carbon dioxide[END_REF]. Iwasa et al. studied the performances for MDR reaction of catalysts based on smectite containing metals such as Mg, Fe, Co or Ni, obtained by hydrothermal synthesis method [START_REF] Iwasa | Preparation and application of nickel-containing smectite-type clay materials for methane reforming with carbon dioxide[END_REF]. The results showed that smectite-based catalysts containing Ni are logically more active for the reforming reaction than smectites containing Fe, Co or Mg, for which conversion rates were less than 2%. In contrast, the smectite-type catalyst containing Ni exhibits a conversion five times greater than that of the reference Ni/SiO 2 catalyst (35% wt. % Ni) prepared by an impregnation process. Gamba et al. [START_REF] Gamba | Catalytic performance of Ni-Pr supported on delaminated clay in the dry reforming of methane[END_REF] used nickel-based catalysts promoted with praseodymium (1, 3 or 5 wt. % Pr) supported on Al-intercalated clay materials.

The intercalation was assisted by microwave and polyvinyl alcohol (PVA). The results

showed that Pr has a promoting effect on the MDR activity of the catalysts by increasing the conversion of CH 4 and CO 2 specifically for 3 wt % nominal content. However, coke formation correlated with praseodymium content and was detrimental when catalysts were calcined at 500°C. Finally, a calcination at 800°C resulted in a good stability for the most active sample. Wang et al. [START_REF] Wang | Preparation, characterization, and catalytic properties of clay-based nickel catalysts for methane reforming[END_REF] studied montmorillonite intercalated by single aluminum atoms (Al-PILC) and mixed lanthanum and aluminum atoms with La-Al = 1: 5 (LaAl PILC)

as the catalyst support with 5% nickel. The catalytic tests in MDR, carried out between 500 and 800 ° C with CO 2 /CH 4 = 1 revealed the beneficial effect of adding lanthanum. The introduction of lanthanum improved the basicity of the catalyst resulting in an enhancement of the catalytic activity, whereas no improvement was observed in terms of stability. Similarly, the beneficial effect on the catalytic activity of the cerium content (with optimum at 3wt % Ce) was investigated by Daza et al. for nickel (10 wt. %) supported on delaminated natural clay material [START_REF] Daza | High-Stable Mesoporous Ni-Ce/Clay Catalysts for Syngas Production[END_REF]. Liu et al. [START_REF] Liu | Natural clay-based Ni-catalysts for dry reforming of methane at moderate temperatures[END_REF] showed an effect of reduction temperature on natural Nibased clay minerals (from Tunisia) modified with Fe and Cu. The catalysts were reduced at 800 or 900°C before MDR tests. The results of the different studies showed that the reduction temperature has a great influence on the catalytic activity, the better performances were obtained with the catalysts reduced at 800°C. The increase in the reduction temperature to 900°C resulted in a decrease in Ni particles size from 15.2 to 8.1 nm; the largest particle size being observed for the most active catalyst. Among the catalysts reduced at 800°C, the most active sample was obtained for Ni-Ce/Cu-Clay composition. The authors emphasized for this sample the importance of the presence of either weak or medium-strength basic sites.

In the auto-thermal reforming of methane, Akri et al., presented the influence of lanthanum and cerium supported on natural illite [START_REF] Akri | Novel nickel promoted illite clay based catalyst for autothermal dry reforming of methane[END_REF][START_REF] Akri | Development of nickel supported La and Ce-natural illite clay for autothermal dry reforming of methane: Toward a better resistance to deactivation[END_REF]. The results showed the promoting effect of Ce on the activity of the catalysts; the addition of cerium significantly increased the specific surface area of the catalyst and the nickel dispersion at the illite surface. Adding cerium to illite also increased the resistance to carbon deposition [START_REF] Liu | Natural clay-based Ni-catalysts for dry reforming of methane at moderate temperatures[END_REF]. On the other hand, the addition of La had a positive effect on activity and stability by inhibiting the formation of FeNi 3 alloy (from the illite support), this alloy being less active than metallic Ni [START_REF] Akri | Novel nickel promoted illite clay based catalyst for autothermal dry reforming of methane[END_REF]. It is then worth noting that the main clay minerals used in catalytic reforming of methane are modified natural clays with addition of active element or synthetic clays [START_REF] Wang | Recent advances during CH4 dry reforming for syngas production: A mini review[END_REF].

The present work aimed at understanding the origin of the performance (activity and stability) of natural Ni-rich clays minerals (originated from Niquelândia in Brazil). The Niquelândia deposit, located in Goiás state, is one of the best known because of its very large quantity of nickel, contained in silicate and in oxidized ores (Figure . S1) [START_REF] Colin | The Initial Lateritic Weathering of Pyroxenites from Jacuba, Niquelandia, Brazil[END_REF][START_REF] Deoliveira | Lateritic Nickel Deposits of Brazil[END_REF]. In Niquelândia deposits, nickel is present in the structure of the clay minerals of the smectite group in substitution to Mg. The clays minerals correspond to a complex mixture of di-and tri-octahedral smectites [START_REF] Colin | The Initial Lateritic Weathering of Pyroxenites from Jacuba, Niquelandia, Brazil[END_REF][START_REF] Deoliveira | Lateritic Nickel Deposits of Brazil[END_REF][START_REF] Mano | MINERALOGICAL CHARACTERIZATION OF Ni-BEARING SMECTITES FROM NIQUELANDIA[END_REF][START_REF] Raous | Mobility of metals in nickel mine spoil materials[END_REF]. The di-octahedral smectites (Figure . 1b) are found in small quantities in the limonite ore corresponding to the upper part of the deposit. These smectites are iron-rich montmorillonite containing Fe 3+ as the main cation in the octahedral sheet and Si 4+ substituted in a small proportion by Al 3+ cation in the tetrahedral sheets. These smectites contain low Ni content as Fe 3+ substitution. The tri-octahedral smectites of the stevensite group are present in silicate ores in the bottom of the deposit (Figure . 1a). Si 4+ cations are dominant in the tetrahedral sheet (no substitution). These smectites contain various amounts of Ni 2+ in the octahedral sheets in substitution of Mg 2+ due to a similar valence and ionic radius [START_REF] Mano | MINERALOGICAL CHARACTERIZATION OF Ni-BEARING SMECTITES FROM NIQUELANDIA[END_REF][START_REF] Mano | Ni-smectitic ore behaviour during the Caron process[END_REF]. It is worth noting that small content of Fe 3+ may be also present in the octahedral sheets of trioctahedral smectites. In both di-octahedral and tri-octahedral smectites, Mg 2+ , Ni 2+ , and Ca 2+ are the main exchangeable cations. As a result, in the smectite of the Niquelândia deposits, nickel is located in the octahedral sheet (octahedral Ni 2+ ) or as an exchangeable cation in the interlayer space (exchangeable Ni 2+ ).

The present study was focused on the behavior of Ni 2+ cations present in tri-octahedral smectite in the conditions of MDR reaction. The reducibility of this element and the influence of the presence of exchangeable Ni 2+ were particularly investigated. The interactions between Ni 2+ and Fe 3+ under reducing atmosphere were also studied and correlated with the catalytic performances.

Experimental

Materials.

The raw tri-octahedral smectite used in the present study is denoted hereafter as AS-ANi3

sample with reference to a previous reported work [START_REF] Mano | Ni-smectitic ore behaviour during the Caron process[END_REF].

Purification of natural clay sample

The clay fraction (<2µm) of the lateritic ore is extracted to concentrate the smectite and remove primary minerals (quartz, pyroxenes) or impurities. Indeed the natural ore is constituted of a very complex matrix of minerals as observed by XRF analysis (Table S1).

The practical method for achieving this goal involves three steps:

(1) The dispersion of the samples by saturation with Na + was performed from 10 g of sample brought into contact with 100 mL of a 1 M NaCl solution to replace the exchangeable cations of the samples by Na + followed by dispersion with ultrasound for 30 seconds. A total of three contacts of at least 24 hours each were made with stirring.

(2) The removal of the excess of sodium chloride by washings carried out with Milli-Q® ultrapure water (18 MΩ.cm) followed by centrifugation (at 4500 rpm). A silver nitrate test was used to verify the effectiveness of the washing procedure. At least four washing steps were necessary to remove the excess of salts.

(3) Then the separation of the <2 µm fraction was carried by centrifugation according to the Stockes law. The parameters of the extractions were adjusted according to the centrifuge used. (Sorvall Legend XFR centrifuge, 1000 rpm, 2 min 30 s). Finally, the clay suspensions were dried in an oven, heated at 60 ° C for two days.

The clay fraction (<2 µm) obtained from this extraction procedure is hereafter denoted: AS-F (Na). Such a fraction only contained Na + in the interlayer space.

The extraction of <2 µm fraction was also done after saturation with a solution of nickel chloride NiCl 2 (1 mol L -1 ) or magnesium chloride solution MgCl 2 (1 mol L -1 ) following the same protocol. The corresponding samples that only contained Ni 2+ or Mg 2+ in the interlayer space will be denoted AS-F (Ni) and AS-F (Mg) respectively.

Cation exchange in the interlayer space

To study the effect of the nickel position in the structure of the clay minerals (i.e. octahedral vs. exchangeable), the AS-F (Na) clay fraction was subsequently saturated with nickel. In the same way as described above, the samples were brought into contact with NiCl 2 (0.5 mol L -1 ) solution, dispersed with ultrasound and stirred one hour before being centrifuged (Centrifuge Sigma 3-16 L centrifuge, 3.300 rpm, 10 min). Five saturation steps were carried out followed by five stages of washing with osmosed water until the absence of chlorides. Finally, the Nisaturated sample is dried at 60°C for 1 day. The sample obtained is noted: AS-F (Na-Ni).

Characterization

The metal loadings present in the clay materials were analyzed by inductively coupled plasma optical emission spectroscopy (ICP-OES) on a PerkinElmer Optima 2000 DV apparatus. For ICP measurements, the samples were mineralized using a mixture of HNO 3 and HCl as well as microwave activation.

The mineralogy of the clay fraction was characterized by X-ray Diffraction using a Bruker D8 advance diffractometer equipped with a copper tube (CuKα, 40 kV, 40 mA). The powder diffractograms were acquired between 2 and 65 ° 2θ with a step of 0.025 ° 2θ and a counting time of 0.6 s. To better identify clay minerals, oriented preparations were prepared by depositing a small amount of suspension prepared with 25 mg of dried samples brought into contact with 3 mL of ultra-pure water and then disaggregated by ultrasound for 30 s. The suspension was stirred for two days before being deposited on a glass slide. The clay mineral film should be sufficiently thick (10 mg/cm 2 ) to ensure infinite thicknesses for X-rays. The oriented preparation enhances 00l reflections series and measurement of layer thickness that is specific of clay mineral groups. The XRD patterns were recorded both in an air-dried (AD) state and after ethylene glycol solvation (EG, vapour at 50ºC overnight) using the same equipment with steps of 0.025 ° 2θ and a counting time of 0.6 s over the range 2 -35 ° 2θ.

The in situ X-ray diffraction was performed in a Bruker D8 Advance X-ray powder The specific surface areas (S BET ) and pore volume were determined by nitrogen adsorptiondesorption isotherm obtained at -196°C using a Tristar 3000 Micromeritics apparatus. Prior to adsorption experiments, the samples were outgassed at 250°C under vacuum overnight. The specific surface area was calculated using the BET equation and pore size distribution was obtained using the BJH method.

Hydrogen Temperature-Programmed Reduction (H 2 -T ) e periments were performed with a Micromeritics AutoChem . The samples ( 120 mg) were first outgassed under pure Ar (30 mL/min) at 300°C for 30min with the heating rate of 10°C min -1 . The samples were then cooled to 50°C under Ar (30 mL min -1 ) for 10 min. Finally, TPR was performed from 50°C to 700 or 900°C for 1 hour under a flow of diluted hydrogen (10% H 2 /Ar, 30 mL min -1 ) with a heating rate of 5°C min -1 .

Transmission Electron Microscopy (TEM) observations were done using a TEM/STEM JEOL 2100 UHR 0.19 nm resolution, equipped with an Energy-Dispersive spectrometer EDS and High-Angle Annular Dark-Field HAADF. The samples were either dispersed in ethanol or dispersed in Milli-Q® ultra-pure water (18 MΩ.cm) by ultrasound. TEM images were then processed using ImageJ software.

A scanning electron microscope (SEM JEOL/MP JSM-IT500 15 kV, probe current 1nA, 11mm working distance), equipped with an Energy Dispersive Spectrometer (EDS BRUKER X-Flash 4010) was used in backscattered electron mode The chemical analyses were (counting time 60 s.) carried out on the raw sample (AS-ANi3) previously compressed to form a pellet 5 mm in diameter and 1 mm thick coated with a layer of carbon.

Methane Dry Reforming: Catalytic test

The catalysts were tested in catalytic methane dry reforming under different experimental conditions in order to study their performances in a pilot described in detail in [START_REF] Gomes | Study of the main reactions involved in reforming of exhaust gas recirculation (REGR) in gasoline engines[END_REF]. In a typical experiment, 150 mg of catalyst (with a diameter of grains between 40 and 100µm) diluted with 150 mg SiC and packed between quartz wool plugs were introduced in a quartz fix-bed reactor at atmospheric pressure. The pressure of the reactor was controlled online by a pressure sensor (Keller PR21S, 0-5 bar) to verify the absence of pressure drop. In a first step, the catalyst sample was reduced in situ under a flow of pure H 2 with a ramp rate of 10 °C min - 1 . A reactant mixture containing CH 4 /CO 2 /N 2 = 1/1/0.5 molar ratio was switched into the catalyst bed. The temperature of reduction as well as the flow-rate of each reactant and the temperature of reaction varied according to the studied parameters and will be detailed later in the text. The outlet gas is analyzed online using two Varian GC: one, with He as carrier gas, equipped with three columns (13X Molecular Sieve, two Porapak Q) and two FID and one TCD detectors to determine the concentration of CH 4 , CO 2 , CO and N 2 ; the other GC was dedicated to the H 2 analysis, with Ar as carrier gas, equipped with a 5A Molecular Sieve column and a TCD detector.

The calculations for the quantification of CH 4 conversion (X CH4 ), CO 2 conversion (X CO2 ), the H 2 yield (Y H2 ) and H 2 /CO molar ratio are described in Eq. 5, Eq. 6, Eq.7 and Eq. 8

respectively where F x is the molar flow of component x. The molar flow was obtained by an external calibration of each gas, using the total flow-rate analyzed online by a flow-meter (Ritter) installed in the outlet gas position and confirmed by the N 2 used as an internal standard. CH

H 2 CO = (F H 2 ) out (F CO ) out Eq. ( 8
)
The yield of C in the dry reforming of methane (considered as the amount of coke formed) was calculated by closing the C balance

Results and discussion

Characterization of materials

Chemical composition

The elemental analysis was determined using ICP-OES analysis of Ni, Na, Mg and Fe elements (Table 1). Raw AS-ANi3 fraction exhibits a high content of Ni (23.8 wt. %) which can be located in the octahedral sheets, the interlayer spaces and the primary minerals as well.

The success of the cation exchange in the interlayer space is verified since the Na and Mg contents are respectively higher in AS-F (Na) (0.76 wt. % Na) and AS-F (Mg) (

Structural and textural characterizations

The The IR spectrum of AS-ANi3 sample, recorded in the wavenumbers range between 400 and 4000 cm -1 , is shown in Figure . 3. The bands at 710 cm -1 and 660 cm -1 , respectively assigned to Ni 3 OH and Mg 3 OH, confirm the tri-octahedral character of the smectite and the presence of Mg and Ni in the octahedral sheet (Mg/Ni stevensite) as described by Mano et al. [START_REF] Mano | MINERALOGICAL CHARACTERIZATION OF Ni-BEARING SMECTITES FROM NIQUELANDIA[END_REF].

In the range 3000 -3800 cm -1 corresponding to the stretching vibrational region of the hydroxyl groups (Ni and /or Fe and/or Mg)-OH, the absorption bands at 3575 and 3624 cm -1 are assigned to Fe 3+ Mg 3 OH and Ni 3 OH respectively. rigid aggregates of plate-like particles [START_REF] Thommes | Physisorption of gases, with special reference to the evaluation of surface area and pore size distribution (IUPAC Technical Report)[END_REF]. The specific surface area (S BET ) is similar for both samples with 68 and 66 m 2 g -1 for AS-ANi3 and AS-F (Na) respectively (Table 2). H 2 -TPR was also performed on AS-F (Na) sample (Figure . S6). The similarity between the H 2 -TPR profiles observed for AS-F (Na) and AS-ANi3 samples demonstrates that the absence of Ni 2+ in the interlayer space does not influence the redox behavior of the clay minerals. The reducible species (mainly Fe and Ni) at the origin of the H 2 consumption are therefore located in the octahedral sheets of the clay material. The absence of peak at high temperature on the H 2 -TPR profile of AS-F (Na) fraction for which primary minerals are supposed to be eliminated confirms the assignment of the peak at 810°C observed for AS-ANi3 sample. In-situ XRD patterns, recorded at room temperature (RT), 500, 600 and 700 °C, were analyzed to complete the understanding of AS-ANi3 reducibility (Figure. Transmission electron microscopy was used to compare the morphology of the Ni particles size depending on the temperature of reduction. AS-F (Na) sample was chosen for this characterization to limit the presence of primary minerals for the analysis. The pictures of AS-F (Na) reduced at 500°C and 700°C are presented on the Figure 6(a) and 6(b) respectively.

After reduction at 500°C, the crystalline structure of the smectite is preserved (upper image showing veil-like particles typical of smectite material) and a small quantity of particles is formed. Conversely, for the sample reduced at 700°C, the results showed that the crystalline structure of the smectite is altered and that the sample exhibits a large number of particles.

These results, therefore, suggest that a reduction at 700°C induces the destruction of the smectite structure and the formation of larger particles. It is confirmed with the zoom made on various particles even if small particles below 10 nm are visible for both temperature of reduction. The EDS analysis was used on various spots (white circles on the pictures) and relative mass Ni/Fe ratios were obtained. The corresponding values are given in the tables below the pictures of the Figure 6. On the basis of ICP result, the global mass Ni/Fe ratio expected for AS-F (Na) fraction is 5.15. It matches well with the ratio obtained for EDS5

analysis which corresponds to a very large spot (1 µm diameter) on AS-F (Na). However, by focusing the EDS analyses on small particles (10-20 nm), one can see that the Ni/Fe ratios obtained on AS-F (Na) reduced at 500 °C are high (EDS 1-4) whereas the ratios are around 3

for the sample reduced at 700°C (EDS 6-8). These results corroborate the hypothesis that increasing the temperature of reduction would promote the formation of FeNi 3 alloy clusters.

Catalytic performances

Behavior of the bare AS-ANi3 material

In a first set of experiments, AS-ANi3 sample (150 mg) was reduced at 700°C during 1 h under pure H 2 flow, heated under pure N 2 flow to 800°C and tested at 800°C with 87.5 mL.min -1 of gas mixture containing CH 4 /CO 2 /N 2 = 1/1/0.5 (Weight Hourly Space Velocity, WHSV = 35000 mL h -1 g -1 ). Between 700°C (plateau of reduction) and 800°C (temperature of reaction) the sample was thus flushed with pure N 2 . The results obtained in terms of reactant conversion and yield in H 2 are reported in the Figure . 7. One can see that the catalytic performances obtained in these conditions of reaction are quite low. AS-ANi3 fraction exhibits 29% conversion of CH 4 and 0.4 mol H 2 /mol CH 4 introduced after 3 hours of time on stream. A slight deactivation is observed during these 3 hours, the materials presenting 35% conversion after the first 30 min of reaction. In addition, it should be noted that the conversion of carbon dioxide (X CO2 = 44%) is greater than that of methane (X CH4 = 29%). This shows that the ore promotes the reverse water gas shift (RWGS) Eq. ( 2), which is consistent with the observed H 2 /CO ratio inferior to 1. The improvement of the catalytic performances was obtained by increasing the contact time between the reactant mixture and the catalyst. The total flow rate of the reactants was decreased from 87.5 to 43.7 mL min -1 and the mass of catalyst kept constant at 150 mg. As a consequence by decreasing the WHSV value from 35000 to 17500 mL h -1 g -1 , both CH 4 and CO 2 conversions increased from 29 to 49% and 44

to 63 % respectively. The difference of contact time has no influence on the occurrence of the RWGS reaction since the H 2 /CO ratio is preserved. It is worth noting that at WHSV = 17500 mL h -1 g -1 , the H 2 yield obtained is close to 0.8 mol per mol of CH 4 introduced which is still far from the expected yield at the thermodynamic equilibrium (1.6 mol or 1.8 mol of H 2 depending on the consideration of C as a product in the thermodynamic calculations; Table .   S2). The effect of the reaction temperature on the AS-ANi3 catalyst has been studied by varying the temperature of reaction between 700, 750, and 800°C with WHSV maintained at 17500 mL h -1 g -1 (Figure . S8). The highest conversion of CH 4 is logically reached at the highest temperature 65% at 800°C. The lowest deactivation is also observed for the test at 800°C. Indeed the formation of carbon is classically decreased at high temperature where the conversion is high [START_REF] Roger | CHAPTER 1 Design of Advanced Catalysts for Natural Gas Reforming Reactions[END_REF].

On the basis of the results obtained from H 2 -TPR and in situ XRD, a study comparing the catalytic performances depending on the pre-reduction temperature was performed. Figure . 8

shows a remarkable impact of the temperature of pre-reduction under pure H 2 on the conversion of the reactants and product yields: an optimum of activity appears with a reduction at 500°C. The sample pre-reduced at 700°C exhibits a similar behavior as the one directly heated under reactant mixture, with 50% of CH 4 conversion. When pre-reduced at 400 or 600°C, the conversion reaches 70% while after reduction at 500°C a maximum of methane conversion is obtained with 80%. In fact, by increasing the reduction temperature from 500°C to 700°C, the conversion of the reactants decreases by 30% whereas the yield in H 2 decreases by 50%. The difference between the drop of methane conversion and H 2 yield is explained by the higher activity in RWGS reaction when the sample is reduced at 700°C. This could be explained by the fact that at 500°C the structure of the clay is very little altered.

Moreover, on the basis of the TPR results, it was shown that 500°C corresponded to the

Behavior of the -AS-ANi3 saturated with different interlayer cations

The AS-ANi3 sample was then modified to better understand the location of the Ni active sites inside this natural ore. The impact of the reduction temperature was also investigated by pre-treating the materials using two temperatures of reduction, which gave strong differences on AS-ANi3 sample: 700 °C and 500°C. A first step consisted in treating the sample by replacing interlayer cations with Na + in order to recover the smectite fraction (without secondary phases) with particle size inferior to 2 µm. As shown by elemental analyses ( although sodium is rather presented as a promoter in the literature in particular since it would limit the coke deposition [START_REF] Nemeth | Na-promoted Ni/ZrO2 dry reforming catalyst with high efficiency: details of Na2O-ZrO2-Ni interaction controlling activity and coke formation[END_REF]. Then, for the sample saturated with Ni 2+ two protocols were The best catalytic performances over AS-ANi3 (raw ore) being obtained for a reduction pretreatment at 500°C, catalytic tests over clay fractions saturated with various cations were repeated in order to study the effect of the interlayer cation under the optimal conditions determined previously (P = 1 atm, F = 43. These new results of catalytic tests, therefore, seem to show that after pre-reduction at 500 ° C, the exchangeable nickel does not have a tremendous impact on the catalytic activity of the catalysts contrary to the conclusion obtained after reduction to 700 °C. Indeed, a noticeable difference is observed in the catalytic performances of the AS-F (Na) clay fraction containing sodium in the interlayer space and in which the Ni is only present in the octahedral sheets.

While it is only very slightly active in the case of a pre-reduction at 700 °C ( CH 4 = 13%, C = 24% and H 2 = 0.01 mol /mol of CH 4 ), it shows, after reduction at 500 °C, the best catalytic performances CH 4 = 84%, C = 95% and H 2 = 1.61 mol /mol of CH 4 (Table S2). In view of the high conversion values, close to those expected at thermodynamic equilibrium, when pre-reduction at 500°C is undertaken, a new set of experiments was carried out by decreasing the contact time (75 mg catalyst, WHSV = 35000 mL g -1 h -1 ). The results of the tests, presented in Figure . 10, logically reveal that, whatever the sample tested, the catalytic performances decrease when the catalyst mass decreases and consequently for The TEM analysis was also performed for the AS-F (Na) fraction (reduced at 500°C and 700°C) after the MDR reaction (Figure . 11). The crystalline structure of the smectite is completely altered which is expected since the temperature of the reaction was 800°C.

Furthermore, the Ni-Fe-containing particles noticeably grew compared with the pictures collected before the reaction (Figure . 6) with size estimated between 20 nm and 80 nm for the AS-F (Na) sample reduced at 700 °C and between 15 nm and 50 nm when reduced at 500 °C.

The TEM images also show some carbon nanotubes present after the catalytic test for the catalyst reduced at 500°C. This is a well-known drawback for active catalyst in MDR when [START_REF] Kustov | CO methanation over supported bimetallic Ni-Fe catalysts: From computational studies towards catalyst optimization[END_REF][START_REF] Huynh | Promoting effect of Fe on supported Ni catalysts in CO2 methanation by in situ DRIFTS and DFT study[END_REF]. It is reinforced by the previous studies which clearly reported that FeNi 3 alloy is less active than the dispersed particles of metallic Ni [START_REF] Akri | Novel nickel promoted illite clay based catalyst for autothermal dry reforming of methane[END_REF]. In the present study, the different characterization techniques corroborate that the reduction at 700°C which can be the subject of a future investigation.

Conclusion

The catalytic performances of Ni-bearing clay minerals obtained from natural nickel lateritic ores from Niquelândia (Brazil) were investigated in methane dry reforming reaction. The raw material, AS-ANi3, directly extracted from the Niquelândia deposit was studied. XRD, FTIR, and XRF/ICP analyses showed that the material is composed of tri-octahedral smectites of the stevensites group with 23.8 wt% Ni and 1.67 wt% Mg located within the octahedral sheets (octahedral Ni) and the interlayer space (exchangeable Ni). The ore also contained 5.20 wt% Fe present in the octahedral sheets. This variety of elements can also be present in other primary minerals that composed the raw ore. An extraction procedure enabled the

CO 2 +

 2 CH 4 ↔ 2H 2 + 2CO (ΔH 0 298K = +247 kJ mol -1 )Eq.

Figure. 1 .

 1 Figure. 1. Tri-octahedral (a) and di-octahedral (b) structures of smectite from Niquelândia deposit [42].

  diffractometer operated at 40 kV and 40 mA, using CoKα radiation (λ = 1.790307 Å), equipped with a Position Sensitive Detector, VANTEC-1, kβ filter (Ni), used in scanning mode. The 2θ range analyzed was 3 to 70°, step of 0.025°, step time equal to 2 s. The flow of reduction was composed of 10 % H 2 /He and the XRD patterns were collected at different temperatures (room temperature, 300 °C, 500 °C, 600 °C, 700 °C and room temperature after reduction).

  powder XRD pattern of the AS-ANi3 sample shows the presence of three major phases, enstatite (MgSiO 3 ), quartz (SiO 2 ) and smectite (Figure. S2). The position of the reflection at 1.52-1.53 Å (60-61 ° 2) indicates that the smectites are tri-octahedral smectite which is in good agreement with the chemical composition obtained by SEM-EDS plotted in Si -Mg+Ni -Fe+Al ternary diagram (Figure. S3). The XRD oriented pattern of the air-dried AS-F (Na) fraction, shown in Figure. S4, reveals an intense and wide (001) reflection at 12.5 Å and a less intense (003) reflection at 3.15 Å, characteristic of the smectite sample saturated with Na + . After solvation with ethylene glycol (EG), the basal spacing expanded from 12.5 Å to 16.5 Å due to the incorporation of two layers of EG in their interlayer space corresponding to a typical behavior of smectite [42]. The peak at 8.5 Å was attributed to the (002) reflection of smectite in EG-solvated state. The other diffraction peaks related to quartz or enstatite are almost undetectable. This is indicative of the removal of primary minerals and the concentration of the smectite in this AS-F (Na) sample. The XRD oriented patterns for AS-ANi3 and fractions with different exchangeable cations (AS-F (Na), AS-F (Mg), AS-F (Ni), AS-F (Na-Ni)) are represented in Figure.2. As expected, the position of the characteristic (001) reflection for the smectite depends on the cation present in the interlayer space. It is observed at 12.5 Å for the AS-F (Na) and at 15 Å for the samples saturated with divalent cations (Ni 2+ , Mg 2+ ). The (001) reflection at 15 Å for AS-F (Na-Ni) shows that Na + is re-exchanged by Ni 2+ in the second step of cation exchange. The presence of peaks characteristic of amphibole, enstatite and quartz on As-F (Mg) and AS-F (Ni) diffractogramms indicates that the extraction/concentration of < 2 µm smectites is more efficient with NaCl 2 than with MgCl 2 or NiCl 2 .

Figure. 2 :

 2 Figure. 2: XRD oriented patterns of (a) AS-ANi3; (b) AS-F (Na); (c) AS-F(Mg); (d) AS-F (Ni) and (e) AS-F (Na-Ni); A: Amphibole (Actinolite: Ca 2 (Mg,Fe II ) 5 Si 8 O 22 (OH) 2 ,PDF n°01-080-0521 -Tremolite: Ca 2 (Mg) 5 Si 8 O 22 (OH) 2 , PDF n°00-009-0437) ; Q : Quartz (SiO 2 , PDF n°01-085-1054) ; E : Enstatite (MgSiO 3 , PDF n°00-022-0714)

Figure. 3 :

 3 Figure. 3: FTIR spectrum of AS-ANi3 sample

Figure. 4 :

 4 Figure.4: H 2 -TPR profile of AS-ANi3 sample.

  S7 and Figure. 5). The in situ XRD patterns were registered using a cobalt anode and the sample support of the device is made of Kanthal (FeCrAl alloy) characterized by an intense peak at 2= 51.9° that shifts to lower angle values with increasing temperatures. The diffraction pattern recorded at room temperature (RT) presents the peaks related to the smectite, amphibole (actinolite: Ca 2 (Mg,Fe II ) 5 Si 8 O 22 (OH) 2 -Tremolite: Ca 2 (Mg) 5 Si 8 O 22 (OH) 2 ), pyroxene (enstatite: Mg 2 Si 4 O 10 ). In these minerals part of Mg can be substituted by Ni. The 5-35 ° region reported in the Figure. S7 displays the thermal behavior of the smectite. The diffractograms collected at higher temperature show a progressive shift of the peak 001 of the smectite from 12.5 Å at room temperature towards 10 Å at 500°C. This corresponds to a collapse of the smectite by dehydration of the interlayer space and partial dehydroxylation. There is also a decrease in the diffracted intensities at higher temperatures which corresponds to a destruction of the structure of the smectite (formation of amorphous phases). The range 2= 30-65° is shown on the Figure 5 to have a focus of the diffraction peaks expected for metallic Ni. The diffractogramm monitored at RT after heating was also added to take into account the angle shift due to the temperature. The comparison of the two diffractograms collected at RT before and post-heating ramp under H 2 clearly reveals that the reduction process leads to the Sample H 2 -consumption (mmol H 2 /g) estimated on the basis of NiO and Fe2O3 reduction to Ni 0 and Fe 0 taking into account the Ni and Fe contents determined by ICP-OES broadening of the X-ray peak of Kanthal support around 51.9° and the appearance of a new wide and less intense peak around 2= 61°. These peaks are characteristic of the formation of metallic Ni clusters but the reflections expected for FeNi 3 alloy are very close[START_REF] Marinho | Highly active and stable Ni dispersed on mesoporous CeO2-Al2O3 catalysts for production of syngas by dry reforming of methane[END_REF][START_REF] Kim | Cooperativity and Dynamics Increase the Performance of NiFe Dry Reforming Catalysts[END_REF].Considering the high content of Ni compared to Fe loading in AS-ANi3, it is reasonable to conclude that isolated small metallic Ni particles appears from 600°C (peaks at 52.1° (100% relative intensity) and 60.9° (45%), ICDD-PDF n°01-070-1849). Because of the width of the diffraction peaks, the formation of FeNi 3 is also speculated ((peaks at 51.5° (100%) and 60.3° (45%) ICDD-PDF n°03-065-3244).

Figure. 5 :

 5 Figure. 5: In situ XRD patterns of AS-ANi3 sample under H 2 from room temperature (RT) to 700°C and return to RT. Q : Quartz (SiO 2 , PDF n°01-085-1054) ; E : Enstatite (MgSiO 3 , PDF n°00-022-0714); Ni (metallic Ni, PDF n°01-070-1849); FeNi 3 (PDF n°03-065-3244) ; * : Kanthal support of environmental chamber.

Figure. 6 :

 6 Figure. 6: TEM pictures associated to EDS analysis of AS-F (Na) sample reduced under H 2 at (a) 500 °C and (b) at 700°C.

Figure. 7 :

 7 Figure. 7: Catalytic performances of AS-ANi3 ore in MDR reaction depending on WHSV at 800°C (m = 150 mg and P = 1 bar); (a): CH 4 conversion (%); (b): CO 2 conversion (%); (c): yield in H 2 (mol/mol CH 4 ) and (d): yield in CO (mol/mol CH 4 ).

  beginning of the reduction peak for the reduction of Ni while at 700°C reduction of both Ni and Fe cation was reached with the possible formation of Fe-Ni alloy at high temperatures 700°C. A 7h duration catalytic test was performed after pre-reduction of the AS-ANi3 material at 500°C which confirms the high activity in these conditions of pre-treatment even if a slight deactivation is noticeable with time-on-stream (Figure. S9).

Figure. 8 :

 8 Figure. 8: Catalytic performances of AS-ANi3 sample in MDR reaction depending on the temperature of reduction under flow of pure H 2 (150 mg catalyst, WHSV = 17500 mL g -1 h -1 , T = 800°C; comparison is made with heating under the reactant mixture); (a) CH 4 conversion; (b) CO 2 conversion; (c) yield in H 2 and (d) yield in CO.

  used, (i) a direct saturation of raw AS-ANi3 sample with NiCl 2 and extraction of the < 2 µm fraction (AS-F (Ni)); an extraction of the < 2 µm fraction after Na saturation followed by an exchange of Na + by Ni 2+ (AS-F (Na-Ni)). The catalytic behavior of AS-F (Ni) sample is very similar to the one of the raw AS-ANi3 ore despite the small decrease of Ni content likely due to the removal of Ni-containing primary minerals. AS-F (Na-Ni) sample exhibits the highest conversion of CH 4 with 64% and the highest H 2 yield with 1.15 mmol H 2 produced because of a better initial activity and a better stability during the 2h of time on stream. The H 2 /CO ratio calculated for AS-F (Na-Ni) sample is very close to the ratio expected at the thermodynamic equilibrium. It means that on this sample, the RWGS reaction is not favored.

Figure. 9 :

 9 Figure. 9: Catalytic performances of all samples in MDR reaction at 800°C after reduction at 700°C (150 mg catalyst, WHSV = 17500 mL g -1 h -1 ) (a) CH 4 conversion; (b) CO 2 conversion; (c) yield in H 2 and (d) yield in CO.

  7 mL/ min, CH 4 /CO 2 /N 2 = 1/1/0.5, T (reduction) = 500°C, T (reaction) = 800°C, WHSV= 17500 mL.h -1 .g -1 ). The results presented in Figure. S10 and in Table. S2, show that the catalytic performances completely differ when the temperature of reduction is limited to 500°C with high CH 4 conversion values around 85 % for all the samples close to the values expected at the thermodynamic equilibrium. The distribution of products obtained from raw ore and clay fractions containing nickel and magnesium (AS-F (Ni), AS-F (Mg), and AS-F (Na-Ni)) are almost similar.

  shorter contact time. However, the interest of these last experiments with conversion and yield values which are moving away from thermodynamic equilibrium is to better observe the differences between the samples. It is then confirmed that the best performances are observed for the sample AS-F (Na-Ni), i.e. the fraction which contains Ni cations within the interlayer space and the octahedral sheets.

Figure. 10 :

 10 Figure. 10: Catalytic performances of all samples in MDR reaction at 800°C after reduction at 500°C (75 mg catalyst, WHSV = 35000 mL g -1 h -1 ) as a function of time; (a) conversion of CH 4 and (b) yield in H 2.

  followed by the reaction at 800°C results in a very strong interaction between Ni and Fe with possible but not demonstrated formation of Ni-Fe alloy. Deeper investigation is still needed, for instance using XAS-XRD analysis, to confirm the formation of FeNi 3 alloy or other Fe-Ni alloy compositions.

Figure. 11 :

 11 Figure. 11: TEM pictures obtained after the MDR test for AS-F (Na) sample pre-reduced (a) at 500°C and (b) at 700°C.

Table . 1

 . : Chemical composition of the different fractions determined by ICP-OES.

	AS-ANi3	0.05	23.80	1.67	5.20
	AS-F (Na)	0.76	22.24	1.79	4.39
	AS-F (Ni)	0.05	22.81	1.78	4.36
	AS-F (Mg)	0.05	20.38	2.42	4.43
	AS-F (Na-Ni)	0.04	22.66	1.35	4.32
		Na	Ni	Mg	Fe

Sample

Chemical analysis: ICP-OES (wt. %)

Table 2 :

 2 Textural properties of AS-ANi3 and AS-F (Na) samples RT to 900°C with a plateau of reduction at 700 °C during 1 h exhibits three main reduction peaks (Figure. 4). The amounts of H 2 consumed for each of the three peaks were determined by area integration and the corresponding values are reported in Table3. A first

	Sample		N 2 -Physisorption
		S BET (m 2 /g)	V P (cm 3 /g)	D P (nm)
	AS-ANi3	68	0.07	1.37
	AS-F (Na)	66	0.08	1.28

3.1.3 Study of the reducibility

As the catalyst has to be reduced before the MDR reaction, H 2 -TPR and in-situ XRD under H 2 were used to evaluate the reducibility of the Ni ore. The H 2 -TPR profile of AS-ANi3 sample, from presence of other oxidized forms for one or both elements. It is worth noting that other reducible elements present in the ore such as Cu or Mn, might also be implied in the amount of H 2 consumed. On the basis of previous literature

[START_REF] Liu | Natural clay-based Ni-catalysts for dry reforming of methane at moderate temperatures[END_REF][START_REF] Kim | Cooperativity and Dynamics Increase the Performance of NiFe Dry Reforming Catalysts[END_REF]

, the first peak can be assigned to the partial reduction of Fe III into Fe II while the reduction to metallic Fe would occur around 575 °C. The larger peak at 700°C would be due to the reduction of Ni II into metallic Ni.

Finally, the peak of H 2 consumption centered at 810°C could be due to the reduction of reducible primary minerals like amphibole (actinolite: Ca 2 (Mg,Fe II ) 5 Si 8 O 22 (OH) 2 ) or pyroxene (enstatite: Mg 2 Si 4 O 10 ) in which part of Mg can be substituted by Ni. Fe II and Ni II in these structures could require higher temperature to be reduced into metallic form.

Table 3 :

 3 Quantification of H 2 consumption during TPR on AS-ANi3 sample.

Table . 1

 . ), the obtained sample, denoted AS-F (Na), contains 22.24 wt% Ni and 0.76 wt% Na. It indicates that Ni present in the interlayer and the secondary phases of AS-ANi3 sample (primary minerals) represents a very minor part of the Ni. Surprisingly, despite the high remaining content in Ni, the catalytic activity of the AS-F (Na) sample was dramatically inhibited after pre-reduction of the sample at 700°C (Figure. 9). Indeed compared with AS-ANi3, the conversion of CH 4 decreases from 49 to 13 % and the yield in H 2 from 0.76 to 0.01 mol/molCH 4 (Table. 4). If Mg 2+ substitutes Na + in the saturation process, part of the activity observed for AS-ANi3 is recovered with 35% conversion of CH 4 and 0.44 mol H 2 produced per mol of CH 4 introduced. It shows that the loss of activity observed for AS-F (Na) is not only due to the absence of Ni in the smectite interlayer but also to the presence of Na + ,

Table . 4

 . : Reactant conversion and product distribution for reaction at 800°C on samples prereduced at 700°C (CH 4 /CO 2 /N 2 = 1/1/0.5, 150 mg catalyst, WHSV = 17500 mL g -1 h -1 )

  Ni particles size are growing above 5 The carbon deposit is almost absent for the postmortem catalyst which was reduced at 700°C. It is explained by the very low activity of this sample which exhibited 13% conversion of CH 4 and no production of H 2 (Table.4).Moreover one can see that the conversion of CO 2 is 24 % and CO is the unique product detected. The results allow suggesting that with large Ni-Fe-containing particles the syngas and C which are produced by the MDR reaction are further reacting with CO 2 via RWGS (CO 2 + H 2  CO + H 2 O) and the carbon gasification (CO 2 + C  2 CO). The Sabatier reaction (CO 2 + 4 H 2  CH 4 + 2 H 2 O) can also explain the absence of hydrogen production for this sample. Indeed it has been well reported that 25Fe75Ni alloy was very active for the methanation of CO 2

Acknowledgements S. B. thanks AUIF, ERASMUS + Global Morocco-Finland (2016-2020, 4FBB97CE477374B5) Ministère de l'Éducation nationale, de la Formation professionnelle, de l'Enseignement supérieur et de la Recherche scientifique (Morocco) for fellowships. Department of Chemistry of University of Oulu is gratefully acknowledged for XRF analysis as well as Stéphane Pronier for TEM analysis and Lilian Eloy for ICP measurements. European Union (ERDF) and Région Nouvelle Aquitaine region are also gratefully acknowledged their financial support.

concentration of the smectite phase. Supplementary saturation steps the smectite with NaCl, MgCl 2 or NiCl 2 were undertaken to study the behavior of the AS-ANi3 depending on the exchangeable cation present in the interlayer space. The corresponding AS-F (Na), AS-F (Mg), AS-F (Ni) were consequently obtained. A last sample AS-F (Na-Ni) was prepared by using two consecutive saturation steps with NaCl followed by NiCl 2 . The temperatureprogrammed reduction performed under a flow containing 10% H 2 combined with situ XRD on reduced samples showed that the Ni started to be reduced at around 500 °C with possible formation of Ni-Fe alloy at higher temperature. Complementary TEM associated with EDX analyses corroborated the presence of Ni-rich Ni-Fe alloy with heterogeneous particle size.

The catalytic tests were performed at atmospheric pressure and 800°C with two different weight hourly space velocities (WHSV) of 35 000 and 17 500 mL h -1 g -1 and a gas composition of CH 4 /CO 2 /N 2 = 1/1/0.5. After reduction of the raw AS-ANi3 at 700°C, the lower WHSV value permitted the conversion of 49% CH 4 and 63 % CO 2 with a yield in H 2 of 0.76 mol per mol of CH 4 introduced. This result shows that the natural clay without any purification step was able to produce syngas with reasonable yield and limited deactivation.

With the same conditions of test but decreasing the pre-reduction from 700°C to 500°C, the conversions and yield obtained indicated that the thermodynamic equilibrium was reached.

The very strong influence of the reduction temperature was confirmed with the tests performed on the exchanged clay fractions. In particular AS-F (Na) was inactive for H 2 production after reduction at 700°C and reached the thermodynamic equilibrium expected in terms of syngas production when the sample was pre-reduced at 500°C. The highest efficiency was obtained for AS-F (Na-Ni) pre-reduced at 500°C which showed that in these conditions of reduction, there is a benefit to have Ni within both structural and exchangeable locations. Finally, the loss of activity resulting from the reduction at 700°C was related to the formation of large Fe-Ni containing particles which are very less active for dry reforming reaction and that favors the consumption of H 2 via the methanation of CO 2 and/or the reverse water gas shift reaction.

Conflicts of interest

The authors do not have any conflicts to declare.