Marie Lacan

Catherine Thèves
email: catheves@toulouse.inserm.fr

Sylvain Amory

Christine Keyser

Eric Crubézy

Jean-Pierre Salles

Bertrand Ludes

Norbert Telmon

• Sylvain

Jean-Pierre Salles

J-P Salles

Detection of the A189G mtDNA heteroplasmic mutation in relation

Keywords: Mitochondrial DNA, Heteroplasmic point mutation, Ageing, Modern and ancient bones

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Introduction

The ageing theory presented by Harman [START_REF] Harman | The biologic clock: the mitochondria?[END_REF] and supported by Beckman and Ames [START_REF] Beckman | The free radical theory of aging matures[END_REF] postulates that the production of free radicals rises with age, and plays a key role in the degenerative processes of senescence. This oxidative stress increase could thus be at the origin of all cellular molecule damage and in particular, could induce the accumulation of lesions on mitochondrial DNA (mtDNA). These lesions were found in numerous tissues with high metabolism, especially in post-mitotic tissues [START_REF] Michikawa | Aging-dependent large accumulation of point mutations in the human mtDNA control region for replication[END_REF][START_REF] Wang | Muscle-specific mutations accumulate with aging in critical human mtDNA control sites for replication[END_REF][START_REF] Bo | Evidence and age-related distribution of mtDNA D-loop point mutations in skeletal muscle from healthy subjects and mitochondrial patients[END_REF]. They took several forms -mostly insertions, deletions and heteroplasmic point mutations. The presence of this latter mutation type has never been closely studied in bone, for which only deletions have been highlighted [START_REF] Pahipa | Age related somatic mitochondrial DNA deletions in bone[END_REF]. However, observing the presence or absence of heteroplasmic mutations in bones could be useful in skeleton identification or in mitochondrial analysis of archaeological bones. Indeed, when heteroplasmic mutations are found, they could have been inherited from the mother (because of mtDNA maternal inheritance) or induced by oxidative stress and the lack of lesion repair. In this latter case, the mutations are somatic and accumulate during the natural ageing process.

Consequently, their study might be helpful in age evaluation (e.g forensic sciences), where burnt or damaged bones prevent the use of traditional osteology markers. In the same way, it could be an important tool in anthropological studies where the detection of these possible age-related mutations could be used along with osteology markers and provide an interesting approach to determining the state of health of past populations.

The general aim of this study was to show that mtDNA heteroplasmic point mutation can be linked with ageing in DNA extracted from bone. The mutation chosen for this was an adenine to guanine transition at position 189 (A189G). This transition presents high, easily detectable heteroplasmic mutation rates in muscle tissue. Its level been detected at an increasing rate in muscle tissue from older individuals [START_REF] Wang | Muscle-specific mutations accumulate with aging in critical human mtDNA control sites for replication[END_REF][START_REF] Bo | Evidence and age-related distribution of mtDNA D-loop point mutations in skeletal muscle from healthy subjects and mitochondrial patients[END_REF][START_REF] Murdock | The age-related accumulation of a mitochondrial DNA control region mutation in muscle, but not brain, detected by a sensitive PNA-directed PCR clamping based method[END_REF][START_REF] Cormio | Mitochondrial DNA mutations in RRF of healthy subjects of different ageNeurobiol[END_REF][START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF].

In the present study, this mutation was investigated in both muscle and bone from the same individuals, thus providing a comparison of heteroplasmic variability between the two tissues. Heteroplasmy detection requires a very sensitive method able to detect mutations at low levels, hence the selection of PCR with PNA/DNA sequencing method which provides an age threshold for the appearance of the mutation [START_REF] Orum | Single base pair mutation analysis by PNA directed PCR clamping[END_REF][START_REF] Hancock | A standard reference material to determine the sensitivity of techniques for detecting low-frequency mutations, SNPs, and heteroplasmies in mitochondrial DNA[END_REF]. The Peptid Nucleic Acid (PNA) molecule is a DNA mimic, in which the negatively charged sugarphosphate DNA backbone is replaced by an achiral, neutral polyamide backbone formed by repetitive units of N-(2-aminoethyl) glycine. The PNA probe perfectly hybridizes to a target DNA region and prevents PCR primer binding, whereas single base-pair mismatches containing PNA exhibit minimal primer binding inhibition [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF][START_REF] Hancock | A standard reference material to determine the sensitivity of techniques for detecting low-frequency mutations, SNPs, and heteroplasmies in mitochondrial DNA[END_REF]. Thus, incorporating the PNA probe in the PCR reaction mixture allows preferential amplification of the mutated fragments and is more sensitive in detecting the mutation during sequencing.

Thus the three specific objectives of this study were (i) to detect the mutation in the mtDNA extracted from bone tissues (ii) to confirm that it is a somatic mutation, to determine the age at appearance and its possible variations depending on the tissues under consideration, and then (iii) to show that these somatic heteroplasmy types can also be evaluated in bone from ancient skeletal remains.

To this end, bone and muscle from the same individuals (called "modern samples") as well as bones from archeological specimens (called "ancient bones") were analysed. In addition, individuals were described as young or old at death to distinguish between the modern and ancient bones.

Materials and methods

Samples

Modern samples were collected during medico-legal autopsies from accidental death victims, secondarily identified and exempt from any specific macroscopically detectable diseases. The sample from each individual consisted of a sternal fragment extremity of the fourth rib, and a part of the associated intercostal muscle. This double sampling had a dual purpose: firstly, to check the exact correlation between sequences of different tissues from the same individual and secondly, to specifically evaluate the level of the 189 heteroplasmy in the bones. In a laboratory dedicated to modern samples, 34 unrelated individuals ranging in age from 1-87 years at death were thus analysed, including 7 individuals from whom only muscle was collected. First, the muscle was carefully removed from the bone with a scalpel. Then, the bones were placed in a microwave oven (5 x 1 min) to eliminate soft tissues [START_REF] Steadman | The effects of chemical and heat maceration techniques on the recovery of nuclear and mitochondrial DNA from bone[END_REF]. The marrow remaining in the bone matrix was cleaned with distilled water and the bone was then crushed with a mortar.

The six ancient samples dating from the 17 th to the early 19 th century were collected during excavations performed by the MAFSO (French Archaeological Mission in Oriental Siberia) team in 2003 and 2004 in Central Yakutia (north-eastern Siberia). The extremely cold, dry climate of this region generally provided a well-preserved nucleic acid chain [START_REF] Amory | STR typing of ancient DNA extracted from hair shafts of Siberian mummies[END_REF]. Of the six specimens studied, two were identified as newborns using the method described by Scheuer and Black [START_REF] Scheuer | Developmental juvenile osteology[END_REF]. Two adults and two old adults were identified by the ageing process of the human skeleton as presented elsewhere [START_REF] Schmitt | Variability of the pattern of aging on the human skeleton: evidence from bone indicators and implications on age at death estimation[END_REF][START_REF] Corsini | Aging process variability on the human skeleton: artificial network as an appropriate tool for age at death assessment[END_REF].

Ancient DNA samples were collected from femur, tibia, humerus and patella bones.

The protocol to eliminate surface contamination and to generate the powdered bone from bone fragments is described elsewhere [START_REF] Keyser-Tracqui | Nuclear and mitochondrial DNA analysis of a 2,000-year-old necropolis in the Eygin Gol Valley of Mongolia[END_REF]. The ancient bones were treated with extreme precautions during each step of sample preparation. DNA extraction and PCR amplification were carried out in a DNA laboratory specifically for ancient bones as described elsewhere [START_REF] Amory | STR typing of ancient DNA extracted from hair shafts of Siberian mummies[END_REF][START_REF] Keyser-Tracqui | Nuclear and mitochondrial DNA analysis of a 2,000-year-old necropolis in the Eygin Gol Valley of Mongolia[END_REF]. In both laboratories, the technical personnel handling bone material were genotyped for the neighbouring 189 mtDNA region. Extraction and amplification blanks were used as negative controls for both modern and ancient samples

Digestion and extraction protocol

DNA from approximately 1cm 3 of muscle and bone samples were extracted with buffer as described in Thèves et al. [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. The aqueous phase was purified with the Cleanmix kit (Talent, Trieste, Italy) and eluted in 100 µl of sterile water. DNA extracted from the bones was then concentrated in Microcon YM30 filters (Millipore, Billerica, MA) in 40µl for the modern samples and 30µl for the ancient ones.

Amplification, purification and sequencing

For each sample, two conditions of PCR were carried out with and without a PNA probe using the same reaction volume as applied elsewhere [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF], with additional PNA of 8 µM final when amplifications were performed with this probe (CAGGCGAACATACTT). Amplification reactions were carried out in a thermocycler T3 (Biometra, Archamps, France) using the following steps: initial denaturation (95°C, 10 min), 35 cycles of denaturation (95°C, 30 s), primer hybridization (60°C, 30 s), elongation (72°C, 1 min) and a final elongation (72°C, 7 min). PCR with PNA was carried out with the additional step of probe hybridization (69°C, 30 s) between the denaturation and primer hybridization steps. For DNA extracted from the ancient bones, the number of cycles was raised to 40, and probe concentrations of 2 and 8µM

were tested, in order to obtain a sufficient quantity of PCR products before sequencing.

All PCR products were purified with QIAquick ® kit (Qiagen GmbH, Hilden, Germany).

The sequencing reactions were performed on both forward and reverse DNA strands for each sample and for both conditions using the BigDye  Terminator v1.1 kit (PE Applied Biosystems, Courtabeuf, France). The products were purified as recommended by the manufacturer and submitted to an ABI Prism 310 Genetic Analyser (PE Applied Biosystems). The electropherograms were analysed using the Sequencing Analysis 5.2

Software (PE Applied Biosystems).

Southern blotting

To confirm the results obtained after sequencing, mtDNA from ancient bones was also tested by Southern blotting. The amplification of PCR products from the samples and the chemiluminescent technique were performed under the same conditions as described in Thèves et al. [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. However, the quantity of ancient DNA was increased: 40 cycles of PCR were carried out and a total of 25 µl PCR products were blotted onto membranes.

Results

The sequences obtained are located between positions 133 and 252 in the HVR-II region of mtDNA. Figure 1 shows the results obtained at position 189 with and without PNA on three bone samples, representing a young individual (Fig. 1a), an old individual from modern bones (Fig. 1b), and an old subject from ancient bones (Fig. 1c).

As described in the guidelines of Brandstätter et al [START_REF] Brandstätter | Monitoring the inheritance of heteroplasmy by computer-assisted detection of mixed basecalls in the entire human mitochondrial DNA control region[END_REF], at position 189, the G nucleotide was considered heteroplasmic if a secondary peak of more than about 10% of the height of the primary peak was present. This was confirmed for both the forward and reverse sequencing reactions.

In muscle tissues: A total of 34 DNA sequences were studied from muscles (Table 1). Of these sequences, 10 taken from people over 39 years of age presented a heteroplasmy at position 189 with conventional sequencing. After the addition of the probe in the PCR mixture before sequencing, only 3 of the samples did not carry the heteroplasmic mutation (1, 14 and 16-year-old). All remaining muscle samples showed the coexistence of the two nucleotides (A/G noticed R) at position 189or only the presence of the G peak in the electropherogram.

In modern bones: Without using the probe, no heteroplasmy could be highlighted (Table 1), which could mean that the mutation never occurs in this tissue at a rate higher than 20-30 %. In fact, the sequencing technique alone detected no secondary peak below this threshold according to the neighboring sequence [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF][START_REF] Hancock | A standard reference material to determine the sensitivity of techniques for detecting low-frequency mutations, SNPs, and heteroplasmies in mitochondrial DNA[END_REF][START_REF] Chinnery | Point Mutations of the mtDNA control region in normal and neurodegenerative human brains[END_REF].

When a PNA probe was added during amplification it showed that 9 out of the 27 individuals tested carried the heteroplasmic mutation in bones, all of whom were over 38 years of age (Table 1). For bone from individuals over 50 years of age, the mutation was only present in 40 % of cases, whereas for muscle tissue the mutation was always found.

In ancient bones: No A189G heteroplasmy was detected in any of the 6 ancient samples analysed using the conventional sequencing technique (Table 1). On the other hand, the sequencing of PCR products with PNA revealed that 3 of them (YAK20, YAK31 and YAK32) showed a second fluorescence peak at position 189, which is clearly visible in Fig. 1c for YAK32. In the two newborn individuals and in one of the two "adult" individuals, the 189R heteroplasmy was not found. These results were obtained with both probe concentrations (2 and 8 µM), the G peak being higher and more clearly identifiable for the latter PNA concentration. Since all the ancient bone samples were identical to CRS [START_REF] Anderson | Sequence and organization of the human mitochondrial genome[END_REF] next to the probe's hybridization region, they could be analysed by Southern blotting. The 189G mutant molecules were only detected on two old individuals (the detection limit being 10 % with the protocol applied [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]). The 189G mutation was not detected in subjects belonging to "newborn" and "adult" age classes by the Southern blot technique.

Discussion

The study of adjacent polymorphisms -flanking position 189 in the electropherograms -confirmed that all samples were identical to the CRS sequence [START_REF] Anderson | Sequence and organization of the human mitochondrial genome[END_REF] next to the probe hybridization region, except for one sample from a 36-year-old individual (Table 1, subject identified by an asterisk). This subject presented one heteroplasmy (G→A) at position 185, visible by sequencing without probe in both tissues. Present in a young individual, this mutation was probably not somatic and was of no interest in the context of our study. Furthermore, this mutation did not interfere with the probe hybridization since the 185A mutated peak is preferentially amplified with the 189G peak after the addition of the PNA probe.

In Table 1, the observation of only a G peak after additional PNA may generate discussion. For example, in the case of a female subject of 21 years old, the identification of only one G peak could be explained in two ways. Firstly: the level of heteroplasmy present in this subject is not known. As described in our previous study [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF], for position 189, heteroplasmy is not detectable at level ≤ 30% for conventional sequencing, and the possible level of heteroplasmy could be between 0-15 % in muscle for subjects of 20-30 years old (see Fig. 5 in [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]). Secondly, in Hancock et al. [START_REF] Hancock | Design and use of a peptide nucleic acid for detection of the heteroplasmic low-frequency mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) mutation in human mitochondrial DNA[END_REF], their Fig. 4 shows the sequence analysis of a representative A3243G sample amplified with no PNA or with various concentrations of PNA. They found that a PNA concentration of 2 µM was optimal for the greatest suppression of the wild-type 3243 mtDNA.

Moreover the mutant 3243 is preferentially amplified and is the dominant species with all PNA concentrations. For 8 µM of PNA, the G mutant peak is heavily predominant, whereas the A wild-type peak is at a low level, and it may be identified as baseline noise. In the majority of the electropherograms in our study, the 189A peak is absent or at a low level for 8 µM of 189A PNA probe.

To evaluate the detection level of the PNA/DNA sequencing method, five DNA samples from individuals aged 1, 13, 14, 16 and 19 years old whose mutation level had been estimated by Real-Time PCR (qPCR) in a previous muscle study [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF], were analysed. For the 1, 14, and 16-year-old individuals -whose rate was less than 0.3 %no heteroplasmy was detected with the PNA/DNA sequencing method (Table 1).

Above this percentage, heteroplasmy was always seen in the electropherograms.

Considering a possible range of error inherent in the qPCR technology, the minimum detection threshold of heteroplasmy may be estimated at 1 % in the conditions applied using this PNA/DNA method. This minimum was also found in other studies which used the same technology [START_REF] Hancock | Design and use of a peptide nucleic acid for detection of the heteroplasmic low-frequency mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) mutation in human mitochondrial DNA[END_REF].

An observation can be made about the quality of the sequences generated in this study. For modern samples, the quantity of DNA was always sufficient to obtain high quality electropherograms. As in the work of Brandstätter and Parson [START_REF] Brandstätter | Mitochondrial DNA heteroplasmy or artefactsa matter of the amplification strategy?[END_REF], for low quality sequences or in the case of sequences with very high background noise, detecting a low level of heteroplasmy is almost impossible. This could also be a problem for ancient bones. PCR and sequencing with or without PNA were repeated two or three times to obtain a good quality sequences. It was observed that with PNA, the 189G peak is better individualized and there is no uncertainty in the detection of heteroplasmy (see Fig. 1c). However, while this PNA/DNA technique is certainly applicable to ancient DNA, a very large quantity is required.

The principal findings are that somatic 189 heteroplasmy can be visualised from bones and that its appearance occurs later when compared to muscle tissue. These facts lead to several discussion points:

Firstly, the difference in (a) the appearance age and (b) the heteroplasmy rate variability between muscle tissue and bone for each individual demonstrate that the independent extraction of each tissue was properly carried out: consequently, the heteroplasmy found in bone DNA can only be from bone cells.

Secondly, this study emphasizes that tissue specificity is present within the same individuals for the 189 mutation. Similar cases had already been found in other tissues [START_REF] Michikawa | Aging-dependent large accumulation of point mutations in the human mtDNA control region for replication[END_REF][START_REF] Wang | Muscle-specific mutations accumulate with aging in critical human mtDNA control sites for replication[END_REF] and for other somatic mutations [START_REF] Chomyn | MtDNA mutations in aging and apoptosis[END_REF]. In the present case, the A189G tissue variability (that is, the difference in both appearance age and heteroplasmy rate variability), could be explained by a difference in the metabolism of muscle and bone [START_REF] Dimauro | Mitochondrial Respiratory-Chain Diseases[END_REF].

Finally, this study demonstrates for the first time, that this type of somatic point mutation exists in bone tissue and is linked to age. The age threshold for the appearance of the mutation A189G in muscle is about 20 years [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. In addition, although the sample is reduced for this tissue, a later age for the appearance threshold in modern bone is suggested: 38 years. However, while the presence of the 189 mutation seems fairly well correlated with age in the muscle, it is more difficult to demonstrate a direct correlation in the bone. In fact, only 9 modern individuals out of 22 over 38 years old (or approximately 41 %) carried the heteroplasmic mutation in bone at a rate higher than 1 %.

In Table 1, bone tissue was not collected from 6 of the youngest individuals.

These samples came from a previous study in which the heteroplasmy level was quantified using PNA/qPCR [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. The autopsies on these individuals were performed before the beginning of this current work, and the extraction of bone was not part of the autopsy protocol [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. Consequently, no bone tissue samples from young subjects (less than 20 years old) were available for testing the occurrence of bone heteroplasmy.

However, the 189 muscle heteroplasmy was only detected in two subjects (13 and 19 years old) at a low level of 0.3%. So, in bones, where heteroplasmy occurs later than in muscle, bone heteroplasmy for these youngest subjects should, in principle, be absent.

These results obtained using PNA/DNA sequencing appear consistent with those obtained from muscle using qPCR [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF]. Indeed in Thèves et al. [START_REF] Thèves | Detection and quantification of the age-related point mutation A189G in the human mitochondrial DNA[END_REF], where the heteroplasmy rate was evaluated in 50 individuals, the mutation was present at a rate of 5% from 20 years old and increased above this age. So in both cases (with the two methods), heteroplasmy was detected in individuals over 20 years of age. Therefore, the technique described in the present study appears to be a fast and reliable method to determine the presence of these somatic mutations. A multi-factorial approach combining this mutation detection method with other markers currently employed to study skeletal remains (such as bone remodeling, cranial suture synostosis, degenerative diseases, dental indicators) is likely to increase the reliability of age evaluation.

For samples taken from ancient bone remains, cold conservation in permafrost provided a sufficient quantity of good quality DNA to analyse the position 189 in the ancient Yakut bones. Our study revealed that only one adult and the two oldest individuals carried the mutation detected using the PNA/DNA sequencing method.

Furthermore, the fact that the 189 heteroplasmy was only detected in the oldest individuals, and not in the youngest, demonstrates that this mutation is not due to oxidation reactions which could start after the individual's death. Actually, modifications of transition types could occur at death in certain sites known to have high mutation rates [START_REF] Gilbert | Characterization of Genetic Miscoding Lesions Caused by Postmortem Damage[END_REF]. In addition, by providing new indicators for age evaluation, this mutation study could also be very helpful during anthropological studies. In such studies, the state of health of past populations could be analyzed by comparing the mutation appearance age with modern populations (still living in a traditional way).

Outlook: The presence of the 189 mutation in bone is not systematic even in old individuals. The study of a single mutation is obviously not sufficient. It would be interesting to relate the presence of this mutation in bone tissue to other mutations known to be highly polymorphic. Specifically, the mutations T408A, C150T, or others in a control region whose presence seems to be clearly correlated with ageing [START_REF] Chomyn | MtDNA mutations in aging and apoptosis[END_REF] could be studied together with the SNaPshot technology [START_REF] Cassandrini | A new novel method for analysis of mitochondrial DNA point mutations and assess levels of heteroplasmy[END_REF]. These mutation studies can also be applied to ancient DNA [START_REF] Bouakaze | First successful assay of Y-SNP typing by SNaPshot minisequencing on ancient DNA[END_REF]. If the combination of these point mutations is not sufficient to obtain accurate ageing indicators, other mutations in the coding region, whose rate also seems to increase with age in muscle tissues, may possibly complete the study. More specifically, the 8344, 3243 and 10006 [START_REF] Chomyn | MtDNA mutations in aging and apoptosis[END_REF][START_REF] Kadenbach | Human aging is associated with stochastic somatic mutations of mitochondrial DNA[END_REF] positions as well as modification types like mini-duplications could be also tested [START_REF] Tengan | Frequency of duplications in the D-loop in patients with mitochondrial DNA deletions[END_REF].

Conclusion:

This study demonstrates for the first time that the 189 heteroplasmy correlated with age in muscle, is also found in bone, despite strong tissue specificity for the appearance of this mutation. Furthermore, it is shown that this mutation is found in archaeological specimens and therefore probably in old and degraded bones studied in forensic cases.

This first result should open up many research lines in the biological field, in the study of ageing mitochondrial DNA, and more specifically in musculo-skeletal tissue.

Pending the discovery of new age-related mtDNA positions, determining the 189 heteroplasmy point could be added to the current osteology markers. Since the presence of this heteroplasmic mutation in bone gives a first indication of the age at death (38 years old or more), the position 189 on the mtDNA can be considered as an interesting potential marker for age estimation in forensic identification. New information about its presence, gathered in a common database for example, could permit researchers to improve its use in identification.

Fig. 1

 1 Fig. 1

Table 1 :

 1 Detection of the A189G heteroplasmy for each individual according to different techniques used.

	Modern samples		Muscle		Bone
	n: 34		189 position	189 position
	Age (years)	sex	Simple	Sequencing	Simple	Sequencing
			Sequencing	with PNA probe	Sequencing	with PNA probe
	1	Y	189 A	189 A	ns	ns
	13	Y	189 A	189 R	ns	ns
	14	Y	189 A	189 A	ns	ns
	16	X	189 A	189 A	ns	ns
	19	Y	189 A	189 R	ns

Acknowledgements

We are grateful to Doctor David Gainza from the CHU Toulouse Rangueil Forensic Laboratory, (France) for sampling the muscle and bone of modern tissues. Thanks are also due to Professor Eric Crubézy and his staff of the MAFSO (French Archaeological Mission in Oriental Siberia) for collecting the ancient samples. We also thank Dr.

Fabrice Dedouit for his valuable comments on the study and Victoria McBride and François-Xavier Ricault for help with English writing.


domain: chim phys spi

text:
Pierre Gibot
email: pierre.gibot@isl.eu

Virginie Goetz
email: virginie.goetz@isl.eu

Aluminium/Tin (IV) oxide thermite composite: Sensitivities and reaction propagation

Keywords: Nanothermites, SnO2, Sensitivities, Combustion velocity Nanothermites, SnO 2, Sensitivities, Combustion velocity

Thermites are energetic composites made of metal and oxidizer particles that react according to an exothermic reaction and release large amounts of energy. Many possible metal and oxidizer couples exist in the literature. Here, the performance of the aluminium reducing metal and tin (IV) oxide (SnO 2 ) combination was reported as a function of the fuel/oxidizer mass ratio. The Al/SnO 2 energetic composite, in stoichiometric conditions, was insensitive to impact, sensitive to friction and extremely sensitive to electrostatic discharge. The combustion velocity was determined to be close to 600 m/s. Changing the stoichiometry affected both the sensitivity thresholds to the friction test (160 N to > 360 N) and flame propagation velocity (480-588 m/s) of the Al/SnO 2 composite. These results are reported for the first time.

Introduction

Within the energetic materials family, thermites are mixtures constituted of two main micronscale components, a fuel metal and an oxidizer (metal oxide), reacting according to a redox reaction characterized by a high energy production (kJ/cm 3 ) coupled to extremely high flame temperatures (> 2000 K) [START_REF] Fischer | Theoretical energy release of thermites, intermetallics, and combustible metals[END_REF]. Fischer et al. listed various reacting pairs by varying either the metal (aluminium (Al), magnesium (Mg), zirconium (Zr) and bore (B)) and/or the metal oxide (iron (III) oxide (Fe 2 O 3 ), molybdenum (VI) oxide (MoO 3 ), tungsten (VI) oxide (WO 3 ), copper (II) oxide (CuO), bismuth (III) oxide (Bi 2 O 3 )), demonstrating the possibility of tailoring the thermochemical proprieties of such energetic composites [START_REF] Fischer | Theoretical energy release of thermites, intermetallics, and combustible metals[END_REF]. Over the past two decades, these energetic mixtures have attracted growing interest due to the use of nanopowders as alternatives to micron-sized reactants [START_REF] Pantoya | Combustion behaviour of highly energetic thermites: Nano versus Micron composites[END_REF][START_REF] Valliapan | Reactivity of aluminum nanopowders with metal oxides[END_REF][START_REF] Sun | Combustion Behaviour of Highly Energetic Thermites: Nano versus Micron Composites[END_REF]. These new advanced nanocomposites, called metastable intermolecular composites (MICs) or nanothermites, exhibit improved energetic properties such as higher combustion velocities and reduced ignition delay times in a more homogeneous combustion mode, making them potential candidates as primers or pyrotechnic time delays.

Among the list of reacting pairs, Al/Fe 2 O 3 , Al/MoO 3 , Al/WO 3 and Al/CuO thermites seem to be the most investigated systems, probably because they exhibit high-energy properties [5-9; 10-14; 15-18; 19-24]. To the best of our knowledge, the aluminium-tin (IV) oxide (Al/SnO 2 ) energetic system has been poorly investigated compared with the above-mentioned ones and scant related information is available concerning the reactive behaviour, pressure released during the combustion and possible mechanism implied in the reaction (nature of the chemical species produced) [START_REF] Ferguson | SnO 2 atomic layer on ZrO 2 and Al nanoparticles: pathway to enhanced thermite materials[END_REF][START_REF] Qin | Core-shell structured nanothermites synthesized by atomic layer deposition[END_REF][START_REF] Sullivan | Simultaneous pressure and optical measurements of nanoaluminium thermites: Investigating the reaction mechanism[END_REF][START_REF] Wang | Synthesis of nano-nickel coated micro-aluminium and thermal reactivity of aluminium/nickel stannic oxide thermite[END_REF]. Other work has focused on the use of tin (IV) oxide in energetic composites but rather as a third component at a low amount (additive, dopant) to control the reactivity of the investigated composite (Al/MnO 2 ) [START_REF] Yang | Hierarchical MnO 2 /SnO 2 heterostructures for a novel free-standing ternary thermite membrane[END_REF]. This existing situation is surprising because the thermochemical properties of the Al/SnO 2 energetic composite are quite interesting and demonstrate a performing system, similar to the Al/WO 3 energetic composite, with a heat of 3 (2.88 kJ/g), an adiabatic reaction temperature of 2876 K and an amount of gas released of 0.34 g per 100 g of composite [START_REF] Fischer | Theoretical energy release of thermites, intermetallics, and combustible metals[END_REF].

Herein, the detailed reactive behaviour, namely the sensitivity thresholds and combustion reaction speed, of the Al/SnO 2 thermite was investigated as a function of the fuel/oxidizer equivalence ratio within the composite. The results showed interesting and promising characteristics of the Al/SnO 2 energetic nanocomposite, equivalent to other classical nanothermites, with potential civil and military emerging applications.

Experimental section

Materials

Tin (IV) oxide material (SnO 2 , < 10 µm, 99%) was purchased from Sigma Aldrich. Aluminium nanopowder (Al, Al-50P, 50 nm, 99%) and acetonitrile (CH 3 CN, 99%) were obtained from Nanotechnologies Inc. and Prolabo, respectively. All chemicals were used without any further purification.

Preparation of the Al/SnO 2 thermites

The general combustion reaction of the Al/SnO 2 energetic system can be described as follows:

4Al + 3SnO 2 → 2Al 2 O 3 + 3Sn (1)
Typically, in pyrotechnics and thermite composites, the reactants are mixed according to a defined fuel/oxidizer weight ratio labelled the equivalence ratio (Φ). The corresponding formula is given in (2):

(2)

𝛷 = ( 𝐹 𝑂 )𝑒𝑥𝑝. ( 𝐹 𝑂 )𝑠𝑡.
where F and O are the quantities of fuel (Al) and oxidizer (SnO 2 ) and the subscripts exp. and st.

represent the experimental and stoichiometric values. For example, at the stoichiometry, the equivalence ratio takes the value of 0.239 for the considered energetic composite. reactive aluminium in the commercial passivated aluminium powder (Al-50P) was determined to be equal to 52.2 wt.% using thermogravimetric analysis.

The Al/SnO 2 energetic composite was prepared as follows [START_REF] Gibot | Safer and performing energetic materials based on polyaniline-doped nanocomposites[END_REF]. Briefly, considering an equivalence ratio of 1, 0.157 g of passivated-Al and 0.343 g of SnO 2 reactants were suspended in a round-bottomed flask containing 100 mL of acetonitrile. The solution was stirred for 1 hour interrupted by ultrasound periods (3 × 5 min.) to break agglomerates and obtain intimate mixing.

Finally, the fuel/oxidizer mixture was recovered by evaporation of the solvent at 80 °C under vacuum (20 kPa) and was placed in an oven at 353 K for 4 hours.

The pyrotechnic behaviour (sensitivity, combustion velocity) of the Al/SnO 2 energetic composite was evaluated as function of the equivalence ratio varied between 0.8 -1.8 (each 0.2).

Characterization techniques.

Tin (IV) oxide (SnO 2 ) and the Al/SnO 2 composite were characterized using standard laboratory characterization techniques. The crystallographic structure was verified by X-ray diffraction analysis achieved using a Bruker D8 to preserve a constant loading density throughout the tube. A schematic illustration of the set-up is given in [START_REF] Gibot | Safer and performing energetic materials based on polyaniline-doped nanocomposites[END_REF]. The combustion phenomena were recorded using a Photron FASTCAM highspeed camera (5×10 4 frames/s) placed in front of the PMMA tube. The combustion speeds were determined using a series of three burn tests.

The bulk densities of the different energetic compositions were determined by the ratio of the amount of matter on the volume of the PMMA tube. Next, the bulk densities were expressed as a function of the theoretical maximum densities (TMD) of the considered mixture calculated from Eq. ( 3).

(

) 𝑇𝑀𝐷 = 1 ∑ 𝑛 𝑖 = 1 𝑚𝑖 𝜌𝑖 3
where i is the considered chemical species and m and  are its mass and density, respectively. For example, considering the amount of Al ( = 2.70 g/cm 3 ), Al 2 O 3 ( = 3.05 g/cm 3 ) and SnO 2 ( = 6.95 g/cm 3 ) in a typical Al/SnO 2 thermite prepared with an equivalence ratio of 1, the TMD was determined to be equal to 4.794 g/cm 3 . powder failed to observe micron-sized primary particles, suggesting that the particle size distribution, described above (Figure 1A), reflected a high degree of agglomeration of the powder.

Additionally, the specific surface area of the SnO 2 powder, by applying the BET model, was determined to be equal to 8 m 2 /g. The crystal structure of the Al/SnO 2 composite was investigated using X-ray diffraction (Supporting Information 1). The diffractogram showed diffraction peaks that can be indexed to a tetragonal lattice associated with a P4 2 /mnm space group describing the SnO 2 Cassiterite phase (JCPDS card 41-1445) and to a face-centred cubic structure with a Fm-3m space group as expected for the aluminium phase (JCPDS card 65-2869). There are no other diffraction peaks confirming the amorphous character of the Al 2 O 3 alumina layer covering the reactive aluminium.

Supporting Information 1: X-ray diffraction pattern of the Al/SnO 2 thermite ( = 1) indexed with the SnO 2 (tetragonal lattice, P4 2 /mnm space group) and Al (face-centred cubic structure; Fm-3m space group) phases. The corresponding Miller planes were given.

Sensitivities of the Al/SnO 2 thermite

The sensitivity level/threshold knowledge of the energetic materials and composites (thermites) is a key parameter for their future deployment both for civilian and military applications. Among the external stresses that can (accidentally) ignite the thermite mixtures, the mechanical and electrostatic ones are the most investigated. For example, in the case of mechanical solicitations, the ignition of energetic materials and thermite composites is commonly interpreted by the "hot spots" theory. This theory is based on the formation of local hot spots, having as origin interparticular friction, an adiabatic compression of gases or chemical decomposition, within the powder subjected to the considered stress [START_REF] Field | Hot-spot ignition mechanisms for explosives and Propellants[END_REF]. When the hot spot temperature exceeds the decomposition/melting points of the components constituting the powder, the (accidental) initiation becomes. for comparison [START_REF]Recommendations on the transport of dangerous goods: Manual of tests and criteria, 4 th revised edition[END_REF][START_REF] Greason | Electrostatic discharge characteristics for the human body and circuit pack[END_REF][START_REF] Talawar | Primary explosives: Electrostatic discharge initiation, additive effect and its relation to thermal and explosive characteristics[END_REF]. Note that the electrostatic discharge sensitivity standard given in the present work corresponds to the maximum energy that can be dissipated from a human being.

First, compared with the standards, a lower sensitivity to mechanical stimuli such as friction stress and an extreme sensitivity to electrostatic discharge (≤ 0.21 mJ vs. 8.33-20 mJ) were noted for all the as-prepared Al/SnO 2 thermites. This behaviour is typical of fuel/oxidizer mixtures in particular regarding the ESD sensitivity [START_REF] Puszynski | Processing and ignition characteristics of aluminium-bismuth trioxide nanothermite system[END_REF]. Concerning the different Al/SnO 2 energetic composites, because the impact threshold was constantly superior to 49.6 mJ-i.e., the maximum value that can be determined from our fallhammer apparatus-it is difficult to estimate a possible influence of the equivalence ratio.

However, based on a previous study [START_REF]STANAG 4489. In: Standardization Agreement (STANAG) on Explosives, Impact Sensitivity Tests[END_REF], all of the composites can be described as insensitive.

Regarding the friction sensitivity, its threshold increases from 160 to > 360 N with the increase in the equivalence ratio from 0.8 to 1.8, allowing classification of the corresponding thermites as moderately sensitive to insensitive [START_REF]STANAG 4487. In: Standardization Agreement (STANAG) on Explosives, Friction Sensitivity Tests[END_REF]. In scrutinizing the evolution of the sensitivity threshold, a "parabolic profile" seems to draw with a minima at 160 N for the Al/SnO 2 mixture compliant to the stoichiometry (ϕ = 1). This minimum can be justified such that, in the whole volume of the thermite, at the micro-and nanoscale, the ideal chemical composition facilitating the initiation was observed. By varying the equivalence ratio (ϕ), one of the main points responsible for the ESD sensitivity is the aluminium fuel particle size. For example, considering Al/MoO 3 as an energetic composite, it was demonstrated that downsizing the Al particle size (from µm to nm) increases the electrical conductance of the mixture; accordingly, the minimum ignition energy is required for ESD ignition. This trend is supported by a spark energy density higher for small-particle-sized media that may easily lead to joule heating at the surface of the particles. This would lead to the formation of hot spots and trigger the initiation of the composite [START_REF] Weir | The role of aluminum particle size in electrostatic ignition sensitivity of composite energetic materials[END_REF][START_REF] Weir | Electrostatic discharge sensitivity and electrical conductivity of composite energetic materials[END_REF]. Thus, nanothermites are more sensitive to electrostatic discharge than their micron-sized counterparts. In the present work, all the investigated Al/SnO 2 mixtures were also exclusively made of nanosized Al particles, explaining the extreme sensitivity of the mixtures. By varying the equivalence ratio from 0. impoverishes itself in the insulating species (SnO 2 , electrical resistivity = 3.10 6 Ω/m). Thus, the conducting character of the energetic mixture is improved and better aligned with the abovementioned principle. The increase in alumina (Al 2 O 3 , electrical resistivity = 10 12 Ω/m), as a protective layer of the aluminium particles, with an increase in the equivalence ratio, was noted by Weir et al. [START_REF] Weir | The role of aluminum particle size in electrostatic ignition sensitivity of composite energetic materials[END_REF] as a benefit to the global conductivity of the mixture by producing more capacitive impedance.

Combustion velocity of the Al/SnO 2 thermite

The combustion velocities of the different Al/SnO 2 thermites by modifying the equivalence ratio (ϕ = 0.8 to 1.8) were determined in a one-dimensional cylindrical design as described in section 2.3.

Before the burn experiment, the loading densities of the test tubes filled with the diverse Al/SnO 2 formulations were calculated. The average values of 0.422 g.cm -3 , 0.456 g.cm -3 , 0.431 g.cm -3 , 0.419 g.cm -3 , 0.415 g.cm -3 and 0.416 g.cm -3 were established based on a series of three tubes, determined for the Al/SnO 2 energetic formulation with an equivalence ratio ranging from 0.8 to 1.8 (every 0.2), respectively. The low standard deviations of the loading densities and linear regressions of the plots of the height filling as a function of the mass of matter added (Supporting Information 2) highlight a reliable and reproducible filling process. The loading densities correspond to 8.4 -9.9% of the respective theoretical maximum densities (TMD) of each formulation considered (Table 2). These data indicate that both the different energetic compositions can be characterized as being loose powders because of apparent densities lower than 10 % of the TMD [START_REF] Walters | Manufacturing and performance of nanometric Al/MoO 3 energetic materials[END_REF] and that a direct comparison of the combustion properties of the thermites can be performed because the gap between the percentages of TMD is negligible [START_REF] Pantoya | Effect of bulk density on reaction propagation in nanothermites and micron thermites[END_REF]. For all the Al/SnO 2 energetic composites, the first flame or spark within the tube was observed in the second image of the sequence considered, suggesting that a globally similar ignition delay time was recorded for all investigated compositions. However, for the very Al-rich mixtures ( = 1.6 and 1.8)-i.e., the energetic composites exhibiting a large deviation from the stoichiometric conditions defined by equation ( 1)-the propagation of the front flame was highly decreased in contrast to that of the other systems ( = 0.8 to 1.4). A delay of approximately 120 µs was observed previously to observe the displacement of the luminous signal in the experiment tube.

Thereafter, the burn rates progress normally within the thermite formulations. At this stage of the investigation, this behaviour was not well understood. The residual chemicals collected at the end of the combustion tests were analysed using X-ray diffraction to determine the nature of the reaction products. The diffractogram is displayed in The combustion velocities of the energetic formulations were determined from the plot of the distance travelled as a function of the time and then performing a linear regression fit to these points (Figure 5). The initial and final extremities of the plots-i.e., the first 40 mm and last 20 mm with respect to the displacement (y axis)-were voluntarily discarded to eliminate possible artefacts or transient zones such as the acceleration phase and loss of confinement, respectively, thus allowing the acquisition of possible steady combustion velocities. The combustion speeds of the different Al/SnO 2 energetic composites were determined and presented in Table 2. The combustion rates varied between 480 m/s and 588 m/s with an optimal propagation speed recorded when the energetic composite exhibited an equivalence ratio ()

ranging from 1 and 1.4 to even 1.6. For fuel-lean Al/SnO 2 composites ( = 0.8), the propagation velocity was slightly delayed by 10%; however, for the highest fuel-rich formulation ( = 1.8), the burn rate value was reduced to more than 18%, establishing itself as the lowest combustion speed recorded. One possible explanation for the higher combustion velocities of the slightly fuel-rich thermite mixtures ( = 1 to 1.4) may be the presence of the metal component (Al) in a significant amount.

The metal, with good thermal properties, such as thermal conductivity and diffusivity, may promote a better heat transfer within the mixture, leading to enhanced combustion velocities [START_REF] Pantoya | Combustion behaviour of highly energetic thermites: Nano versus Micron composites[END_REF].

For example, the thermal conductivities and diffusivity of Al are 236 W/m.K and 9.7.10 -5 m 2 /s, respectively, clearly higher than those of Al 2 O 3 (36 W/m.K and 1.5.10 -7 m 2 /s, respectively) and However, when the amount of Al is very high in the thermite formulation ( = 1.8), the default oxidizer (SnO 2 ) amount is overpowered and becomes prejudicial to a quick reaction. By analogy, in the fuel-lean Al/SnO 2 thermite-i.e.,  = 0.8-the lack of metal fuel may support the low value of the burn rate (548 m/s).

Table 3 gathers the properties of some of thermites reactions mentioned in the literature such as Al/Fe 2 O 3 , Al/MoO 3 , Al/WO 3 , Al/CuO and Al/Bi 2 O 3 [START_REF] Bezmelnitsyn | Modified Nanoenergetic Composites with Tunable Combustion Characteristics for Propellant Applications[END_REF][START_REF] Son | Combustion of nanoscale Al/MoO 3 thermite in microchannels[END_REF][START_REF] Sanders | Reaction propagation of four nanoscale energetic composites (Al/MoO 3 , Al/WO 3 , Al/CuO and Bi 2 O 3 )[END_REF][START_REF] Puszynski | Processing and ignition characteristics of aluminium-bismuth trioxide nanothermite system[END_REF][START_REF] Weir | The role of aluminum particle size in electrostatic ignition sensitivity of composite energetic materials[END_REF][START_REF] Bach | Modulation of the reactivity of a WO 3 /Al energetic material with graphitized carbon black as additive[END_REF][START_REF] Foley | Development of Nanothermite Composites with Variable Electrostatic Discharge Ignition Thresholds[END_REF][START_REF] Pichot | Nanodiamond for tuning the properties of energetic composites[END_REF]. Regarding the theoretical thermochemical data as the adiabatic reaction temperature and the heat of reaction, the Al/SnO 2 thermite exhibits rather low values compared to the other systems. For example, its adiabatic temperature and heat of reaction are of 2876 K and 15.39 kJ/cm 3 , respectively that is relatively far away the 3253 K and 20.82 kJ/cm 3 for the Al/MoO 3 and Al/CuO composites, respectively. In term of gas releasing, the Al/SnO 2 reaction produces a significant amount of gas (0.29 mol./100 g of thermite), as the Al/CuO and Al/Bi 2 O 3 formulations, which would make it interesting for applications where a mechanical work (transport, inflating) is needed such as in airbags or torch-type output. Then, from an experimental point of view, it is always difficult to compare performance (sensitivities, combustion velocity, and pressurization rate) of energetic systems between themselves (data against data). This is due to the fact that these data strongly depend on the physicochemical properties of the components constituting the systems considered (e.g., the morphology, granulometry, textural properties as well as the fuel/oxidizer equivalence ratio). However, the idea of positioning the Al/SnO

Conclusion

In this investigation, the performance of the Al/SnO 2 energetic formulation, in terms of sensitivity and reactive behaviour, has been described. The fuel:oxidizer weight ratio, or equivalence ratio (), was selected as the adjusting parameter. Globally, the energetic composite related to the stoichiometric ( = 1) is insensitive to shock (> 49.6 mJ), moderately sensitive to friction (160 N) and, classic for such composites, extremely sensitive to electrostatic discharge (0.21 mJ). The variation in the equivalence ratio, regardless of the trend (decrease or increase), does not modify the ESD and the impact sensitivities, while a mitigation of the friction sensitivity was recorded until the mixture was insensitive (> 360 N for  = 1.8). Such evolution was interpreted by the loss of the stoichiometry locally and the non-pyrotechnic character of the alumina (the amount increases with  in contrast to the SnO 2 oxidizer. The combustion speed of the powdered Al/SnO 2 mixture, in confined configuration (tubes), ranged from 480 to 588 m/s with the ceiling value within the 1-to 1.6-equivalence ratio domain.

These results showed that the Al/SnO 2 thermite, poorly investigated until now, exhibits energetic performances (sensitivities and combustion velocity) in the same order of magnitude as those of the most investigated energetic systems in the literature (Al/Fe

1 .

 1 Characterizations of the SnO 2 oxidizer and Al/SnO 2 energetic composite.

Figure 1

 1 Figure1shows the particle size distribution (PSD) and scanning electron microscopy (SEM)

Figure 1 : 8 the

 18 Figure 1: (A) Particle size distribution (PSD) curve and (B) SEM images of the SnO 2 material used as an oxidizer in the Al/SnO 2 energetic composites.

Figure 2 :

 2 Figure 2: SEM views of the Al/SnO 2 thermite composite ( = 1) at (A) high magnification and (B) lower magnification with the corresponding elemental mapping where aluminium and tin are reported in green and blue plots, respectively.

  SnO 2 -rich (ϕ = 0.8) and Al-rich (ϕ = 1.2 to 1.8) energetic mixtures were elaborated and synonymous with a stoichiometry default between the Al and SnO 2 reactants for the energetic mixtures considered. Next, the probability of finding at any points of the energetic mixture the ideal chemical composition decreases. Accordingly, the the initiation are no longer met and it becomes necessary to provide a higher friction force (168 N, 216 N, 288 N, 324 N and > 360 N for ϕ = 0.8, 1.2, 1.4, 1.6 and 1.8, respectively) to create hot spots in the energetic composites to ignite them. Moreover, the force intensity was higher for Al-rich (ϕ = 1.2 to 1.8) energetic mixtures because the increase in the aluminium amount equals the increase in the amount of alumina species within the mixture as Al nanoparticles are chemically composed of pure aluminium and alumina according to a core-shell structure (alumina surrounds the Al metal core). Alumina is a pyrotechnical inert material compared with tin (IV) oxide used initially as an oxidizer[START_REF] Fischer | Theoretical energy release of thermites, intermetallics, and combustible metals[END_REF], explaining why the friction sensitivity thresholds for Al/SnO 2 are higher with ϕ superior or equal to 1.2 than for  equal to 0.8.Concerning the electrostatic discharge sensitivity (ESD), the different Al/SnO 2 energetic composites (ϕ ranging from 0.8 to 1.8) exhibited a very high sensitivity to spark with values varying from 0.14 to 0.21 mJ, one order of magnitude lower than what a human body can dissipate(8.33-20 mJ). These very low thresholds are characteristic of extremely sensitive thermites for which an accidental reaction may occur during handling. As demonstrated by Weir et al.[START_REF] Weir | The role of aluminum particle size in electrostatic ignition sensitivity of composite energetic materials[END_REF],

Figures 3 Supporting Information 2 :

 32 Figures 3 shows the sequences of combustion frames extracted from the video recordings of the

Figure 3 :

 3 Figure 3: Sequences of the combustion images (top to down) of the Al/SnO 2 thermite with an equivalence ratio (ϕ) of (A) 0.8, (B) 1, (C) 1.2, (D) 1.4, (E) 1.6 and (F) 1.8. The interframe time was 60 µs, the scale bar reported in the inset of each first image represents 10 mm, and the vertical dotted lines delimit the tube extremities.

Figure 4 .

 4 Figure 4. The diffraction peaks were attributed to the tin metal (Sn, JCPDS card 89-4898) and

Figure 4 :

 4 Figure 4: X-ray diffraction pattern of the reaction products of the Al/SnO 2 thermite system. ( glass threshold)

Figure 5 :Table 2 :

 52 Figure 5: Front reaction position as a function of time elapsed for the Al/SnO 2 energetic composites (plot based on one burn tube experiment for each composite). For clarity, the lines are plotted.

14 SnO 2 (

 142 40 W/m.K and 1.45.10 -7 m 2 /s, respectively) that comprise the composites[START_REF] Powell | Thermal conductivity of selected materials[END_REF][START_REF] Turkes | Thermal conductivity of SnO 2 single crystals[END_REF].

Figure 1 :Figure 1 :Figure 2 :Figure 2 :Figure 3 :

 11223 Figure 1: (A) Particle size distribution (PSD) curve and (B) SEM images of the SnO2 material used as an oxidizer in the Al/SnO2 energetic composites. 120x89mm (150 x 150 DPI)

Figure 4 :

 4 Figure 4: X-ray diffraction pattern of the reaction products of the Al/SnO2 thermite system. ( * glass threshold) 186x160mm (150 x 150 DPI)

Figure 5 :

 5 Figure 5: Front reaction position as a function of time elapsed for the Al/SnO2 energetic composites (plot based on one burn tube experiment for each composite). For clarity, the lines are plotted. 197x162mm (150 x 150 DPI)

Supporting Information 2 :Supporting Information 2 :

 22 Supporting Information 1: X-ray diffraction pattern of the Al/SnO2 thermite ( = 1) indexed with the SnO2 (tetragonal lattice, P42/mnm space group) and Al (face-centred cubic structure; Fm-3m space group) phases. The corresponding Miller planes were given. 140x120mm (150 x 150 DPI) Page 30 of 32 URL: http://mc.manuscriptcentral.com/uegm Email: james.short@cecd.umd.Filling up height as a function of the mass added for (A) the thermite Al/SnO2 ( = 1, series of 3 tubes) and for (B) all the Al/SnO2 energetic composites (average filling up based on a series of 3 tubes for each composite). For clarity, the lines are plotted. 198x163mm (150 x 150 DPI) Page 31 of 32 URL: http://mc.manuscriptcentral.com/uegm Email: james.short@cecd.umd.Filling up height as a function of the mass added for (A) the thermite Al/SnO2 ( = 1, series of 3 tubes) and for (B) all the Al/SnO2 energetic composites (average filling up based on a series of 3 tubes for each composite). For clarity, the lines are plotted. 198x162mm (150 x 150 DPI) Page 32 of 32 URL: http://mc.manuscriptcentral.com/uegm Email: james.short@cecd.umd.edu Journal of Energetic Materials

  The sensitivity test data for Al/SnO 2 with different fuel/oxidizer weight ratio energetic composites are presented in Table1. Standards, either established by the United Nations intergovernmental organization and relating to the transport of dangerous goods or based on the literature, are given

	F
	o r
	P
	e e r
	R e v
	i e w
	O n
	l
	y
	9

Table 1 : Sensitivity thresholds of the Al/SnO 2 thermite as a function of the fuel/oxidizer mass ratio (equivalence ratio ϕ).

 1

Table 3 : Properties relating to thermite reactions. (a issued from [1], b determined in our laboratory but not yet published and c for not found in the literature). The combustion velocities were determined from burn experiments performed in a confined mode with hollow plastic tubes with an inner diameter of approximately 3 mm.

 3

  2 O 3 , Al/MoO 3 , Al/WO 3 , Al/CuO and Al/Bi 2 O 3 ). Thus, this material can be considered an emerging solution in terms of energetic composites to help with successful deployment of thermites in many energetic applications.

	F o r P e e r R e v i e w F o r P e e r R e v i e w F o r P e e r R e v i e w F F o r o r P e P e e r e r R e R v e v i e i e w w O
	O n O n O n O n l n y
	l l l l
	y y y y

URL: http://mc.manuscriptcentral.com/uegm Email: james.short@cecd.umd.eduJournal of Energetic Materials

Acknowledgements

Page 15 of 32 URL: http://mc.manuscriptcentral.com/uegm Email: james.short@cecd.umd.edu

Journal of Energetic Materials

The authors gratefully acknowledge the French National Centre for Scientific Research (CNRS), French German Research Institute of Saint-Louis (ISL, Saint-Louis, France) and University of Strasbourg (UNISTRA, Strasbourg, France) for funding and D. Spitzer (director of NS3E, Saint-Louis, France) for his support and help in obtaining the funding of the second-year Master's Degree internship of V. Goetz. The authors thank M. Comet (NS3E, Saint-Louis, France), F. Schnell (NS3E, Saint-Louis, France) and Y. Boehrer (ISL, Saint-Louis, France) for the igniters' preparation, scanning electron microscopy analysis coupled to energy dispersive X-ray spectroscopy and combustion video recordings, respectively.

F o r P e e r R e v i e w

O n l y