Murat Kaloğlu

Nevin Gürbüz

David Sémeril

İsmail Özdemir
email: ismail.ozdemir@inonu.edu.tr

Hemilabile Ruthenium(II)-(p-cymene)-N-Heterocyclic Carbene Complexes for the N-Alkylation of Amine Using the Green Hydrogen Borrowing Methodology

Six ruthenium(II) complexes with the general molecular formula [RuCl2(NHC)(η 6 -p-cymene)] (NHC = N-heterocyclic carbene) were synthesized by the transmetalation method from [RuCl2(η 6 -pcymene)]2 and silver(I)-NHC complexes. All complexes were fully characterized by analytical and spectral (FT-IR, elemental analysis and 1 H and 13 C NMR) methods. The solid-state structure of one of the ruthenium complexes {dichloro-[1-(2-(2-ethoxyphenoxy)ethyl)-3-(3,5-dimethylbenzyl)benzimidazol-2-ylidene](p-cymene) ruthenium(II)} has been established by single-crystal X-ray diffraction study, which revealed that the ruthenium atom adopt a classical piano-stool coordination geometry. Under the optimised conditions, these ruthenium complexes were found to be efficient catalysts for N-alkylation of aniline with arylmethyl alcohols using the hydrogen borrowing strategy, which is a cost-effective and environmentally attractive reaction for the preparation of N-alkylated amines.

Introduction

Alkyl amines are widely used in both the bulk and fine chemical industries as biologically active compounds, agrochemicals, functionalized materials and dyes [1]. Due to the importance of these products, alkylation of primary amines to secondary or tertiary amines is one of the most fundamental and important reactions in organic chemistry [2]. Alkylated amines are traditionally synthesized by using alkylating agents, such as alkyl halides in the presence of stoichiometric amounts of inorganic bases. However, these procedures are often associated with environmental problems, such as the toxic nature of alkylating agents. In addition, these reactions generate equimolar amounts of wasteful salts as by-products (Figure 1a) [3]. Thus, valuable methods have been developed for the catalytically synthesis of secondary or tertiary amines, including Buchwald-Hartwig [4],

Ullmann [5], hydroamination [6] and hydroaminovinylation [7]. In this context, the creation of C-N bonds from primary or secondary amines from alcohol as alkylating agent using the "hydrogen borrowing" or "hydrogen auto-transfer" methodology has more recently attracted attention (Figure 1b) [8]. In this onepot reaction, the alcohol is dehydrogenated in situ to give the corresponding aldehyde or ketone by the catalyst, which "borrows" hydrogen from the substrate. Subsequent condensation with the amine and final rehydrogenation leads to alkylated amine. This transformation is highly atom economical and environmentally friendly, since water was produced as the only by-product. At the beginning of the 1980s, pioneering reports dealing with the rhodium and ruthenium catalysed N-alkylation of amines by alcohols were described independently by the groups of Grigg ([RhH(PPh3)4]) [9], Watanabe ([RuCl2(PPh3)3]) [10] and

Murahashi

([RuH2(PPh3)4]) [11]. These first reports demonstrated that noble metals such as rhodium, ruthenium could catalyse the hydrogen borrowing reaction, but with a lack of selectivity, formation of both secondary and tertiary amines, due to the hard catalytic conditions. Then new catalysts were developed to carry out this reaction under milder conditions, especially lower temperature [12]. Recent of applications catalysed by mainly ruthenium-or iridium-based complexes have been described by the groups of Williams [13], Fujita [14],

Crabtree [15], Peris [16] and from Beller [17].

Over the last 20 years, N-heterocyclic carbenes (NHCs)

have become an ubiquitous alternative to phosphine ligands in homogeneous catalysis [18] in particularly in association with ruthenium salts [19], which have been reported to be active catalysts for the N-alkylation of amines with alcohols by hydrogen borrowing strategy [20].

In a recent study we have reported the synthesis of benzimidazolium salts and their silver(I) complexes, which were tested as potential antimicrobial agents [21]. In the present article, we now described the synthesis and characterization of two benzimidazolium salts and six novel ruthenium(II) complexes of the general formula [RuCl2(NHC)(η 6 -p-cymene)]

1a-f (Figure 2). These complexes were tested in N-alkylation of aniline with arylmethyl alcohols using the hydrogen borrowing strategy. The performances and stabilities of the catalytic systems may be attributed to the ability of the ether or the arylmethyl substituents of the benzimidazol moiety to temporarily interact, as hemilabile ligand, with the metal's first coordination sphere [22].

Results and Discussion

Preparation of Ruthenium(II) Complexes

The synthesis of ruthenium complexes 1a-f required that of the benzimidazolium salts 3a-f, respectively (Scheme 1). The two new benzimidazolium salts 3a and 3b were obtained by alkylation (RBr; R = CH2-C6H3-3,5-(CH3)2 and CH2-C6-(CH3)5) of benzimidazole 2 in 87 and 83 % isolated yield, respectively. The salts 3a and 3b were fully characterised by elemental analysis, 1 H and 13 C{ 1 H} NMR and infrared spectroscopies (Table 1). The FT-IR spectra of the free ligands showed a broad band at 1572 and 1556 cm -1 , which corresponds to the -C=N-bond vibration of 3a and 3b, respectively. The signal of the NCHN proton appears in the expected range, at  = 11.42 and 10.33 ppm for 3a and 3b, respectively. Their 13 C NMR spectra display the characteristic singlet at 143.1 and 142.1 ppm for the NCHN carbon for 3a and 3b, respectively [21,23]. The silver complexes 4a and 4b were synthesized according to the general method described by Wang and Lin [24] by reacting the corresponding benzimidazolium salts with Ag2O in dichloromethane under exclusion of light at room temperature for 24 h. The spectroscopic data are similar to those found for other silver(I)-NHC complexes [21,25]. The two silver(I) complexes exhibit a characteristic v(NCN) band at 1442 and 1451 cm -1 for 4a and 4b, respectively. In their 1 H NMR spectra, the NCHN protons were disappeared upon complexation with silver.

As previously mentioned by several authors [26], the Ag-C atom could not being observed by 13 C{ 1 H} NMR spectroscopy in complexes 4a and 4b. Note that, benzimidazolium salts 3c-f and their corresponding silver complexes 4c-f were conveniently prepared according to a method reported earlier [21].

The ruthenium(II) arene complexes 1a-f were obtained in 57-70 % yields by transmetalation of the corresponding silver adducts 4a-f and [RuCl2(p-cymene)]2 (Scheme 1). The air-and moisture-stable ruthenium complexes are brown in colour and soluble in common chlorinated organic solvents. They are found to be diamagnetic and they were fully characterised by elemental analysis, 1 H and 13 C NMR and infrared spectroscopies (Table 1). Their FT-IR data each show a band at 1450-1454 cm -1 (v(NCN)) and their 13 C{ 1 H} NMR spectra each display a singlet in the range 189. 9-191.5 [a] As previously reported by several groups [26], for compounds 4a and 4b, the characteristic C-Ag picks were not observed.

Structural Characterization of Complex 1a

The solid-state structure of the mononuclear ruthenium complex 1a was established by a single-crystal X-ray diffraction study (Figure 3 and Table 2). The complex crystallised with one molecule of chloroform. The half-sandwich arene ruthenium(II) complex adopts a classical piano-stool geometry with a typical Ru-Ccarbene bond length (2.060(4) Å) [22]. To determine the optimal catalytic conditions, ruthenium complex 1c (5 mol %) was used in the alkylation of aniline with benzyl alcohol at 120 °C in toluene. Each experiment was stopped after 24 h. As attempt, no products were formed without the addition of ruthenium complex (Table 3, entry 1). In the first series of reactions, we determined the optimal base by employing either t BuOK, Cs2CO3 or KOH. As can be inferred from the results (Table 3, entries 2-4), the most efficient base was t BuOK, which led to a full conversion with an A/B ratio of 85/15. Reducing the catalyst loading to 1 mol % decreased the conversion and the selectivity toward the formation of compound A (Table 3, entries 4 and5). A good compromise is the use of 2.5 mol % of 1c, which led to a full conversion after 24 h with an A/B ratio of 80/20 (Table 3,entry 6). Operating at lower temperature or for a shorter time reduced both the conversion and the chemioselectivity (Table 3, entries 7 and8). Note that, under our catalytic conditions, only mono-alkylated compounds were formed, no bis-alkylated derivatives were observed. [a] Reagents and conditions: 1c, benzyl alcohol (1.0 mmol), aniline (1.1 mmol), base (2.5 mmol), toluene (3 mL). The conversions and the selectivities were determined by GC and GC-MS analysis with the calibrations based on dodecane.

Under the determined optimal conditions, t BuOK as base in toluene at 120 °C for 24 h, the ruthenium complexes 1a-f (2.5 mol %) were further examined in the N-alkylation of aniline with four arylmethyl alcohols, namely, benzyl alcohol, ptolylmethanol, (4-methoxyphenyl)methanol and (3,4dimethoxyphenyl)methanol (Table 4). In all tests, conversions in arylmethyl alcohol higher than 78 % were measured. Due to the similar nature of the NHC moieties, small differences in reactivities were observed. Nevertheless, by comparison with common NHCs such as 1,3-bis (2,4,6trimethylphenyl)imidazolylidene [19f], the presence of benzimidazolylidene contributes to promote the formation of amines A rather than imines B.

The two more efficient ruthenium precatalysts were complexes 1c and 1d. With the latter complexes, full conversions were reached for benzyl alcohol (1c), ptolylmethanol ( 1d) and (4-methoxyphenyl)methanol (1d) (Table 4, entries 3, 10 and 16). A conversion of 95 % for the more sterically hindered (3,4-dimethoxyphenyl)methanol was observed when ruthenium complex 1e was employed (Table 4, entry 23). In all catalysis, high chemioselectivities toward the formation of amines A were obtained. In fact, A/B ratios of 95/5

were measured for alkylation with benzyl alcohol and ptolylmethanol when complexes 1a and 1f were used, respectively (Table 4, entries 1 and 12). For the two alcohols bearing methoxy substituents, (4-methoxyphenyl)methanol (A/B = 97/3) and (3,4-dimethoxyphenyl)methanol (A/B = 94/6), the higher proportions of amines A were formed when ruthenium complex 1e, was employed (Table 4, entries 17 and 23). [a] Reagents and conditions: [Ru] (0.025 mmol, 2.5 mol %), arylmethyl alcohol (1.0 mmol), aniline (1.1 mmol), t BuOK (2.5 mmol), toluene (3 mL), 120 o C, 24 h. The conversions and the selectivities were determined by GC and GC-MS analysis with the calibrations based on dodecane.

Ruthenium complexes 1a-f were further assessed in the Nalkylation of aniline with furfuryl alcohol or 2-thiophene methanol (Table 5). Both alcohols were efficiently converted in 24 h when the reactions were performed in the presence of complexes 1c (99 % for furfuryl alcohol and 97 % for 2-thiophene methanol; Table 5, entries 3 and9). With the latter precatalyst, the proportions of amines products A formed were 91 % with furfuryl alcohol and 98 % with 2-thiophene methanol. Repeating the run with furfuryl alcohol and complex 1f increase the chimioselectivity toward the formation of A to 93 % (Table 5, entry 6). [a] Reagents and conditions: [Ru] (0.025 mmol, 2.5 mol %), heteroaromatic alcohol (1.0 mmol), aniline (1.1 mmol), t BuOK (2.5 mmol), toluene (3 mL), 120 o C, 24 h. The conversions and the selectivities were determined by GC and GC-MS analysis with the calibrations based on dodecane.

Conclusions

In this study, we prepared six new ruthenium complexes in which a nitrogen atom of the N-heterocyclic carbene ligand is substituted either with an ether or an arylmethyl chain, which may act as a hemilabile ligand. These complexes were all found to be suitable for N-alkylation of aniline with a range of alcohols including arylmethyl alcohols and heteroaromatic alcohols.

These catalytic results reflect the high stability of the ruthenium active species probably due to the ability of the hemilabile ligands to transitory coordinate to the metal center. Future work will aim at exploiting the stabilisation of active species with such hemilabile ligands in catalytic applications.

Experimental Section

General Methods: All reactions performed to prepare the benzimidazolium salts and their metal complexes were carried out under argon in flame-dried glassware using standard Schlenk techniques. The benzimidazolium bromides 3c-f and the silver(I) complexes 4c-f were prepared according to literature procedures [21].

General

Figure 1 .

 1 Figure 1. (a) Traditional approach using alkyl halides and (b) hydrogen borrowing strategy for C-N bond formation.

Figure 2 .

 2 Figure 2. Ruthenium complexes 1a-f synthesised and assessed in the present study.

Figure 3 .

 3 Figure 3. ORTEP drawing illustrating the solid state structure of complex 1aCHCl3 (displacement ellipsoids are drawn at the 50 % probability level).

Table 2 .

 2 Selected bond lengths [Å] and angles [°] in complex 1aCHCl3. of Aniline with Arylmethyl Alcohols The ruthenium complexes 1a-f were evaluated as catalysts for N-alkylation of aniline with arylmethyl alcohols. The corresponding tests were performed in the presence of a base in toluene at 120 °C for 24 hour. In the present catalytic conditions, two products may be formed, namely, a N-(arylmethyl)aniline (A) or a N-phenyl-arylmethylanimine (B) (Scheme 2).

Scheme 2 .

 2 Scheme 2. N-alkylation of aniline with arylmethyl alcohol.

  Chemicals and solvents were purchased from Sigma-Aldrich and Merck. Dichloromethane, dimethylformamide, toluene and diethyl ether were of anhydrous quality and were used as received. The solvents used were purified by distillation over the drying agents indicated and were transferred under argon. All reagents were purchased from commercial sources and used without further purification. Microanalyses were performed by İnönü University Scientific and Technological Research Center (Malatya, Turkey). IR spectra were recorded on ATR unit in the range of 400-4000 cm -1 with Perkin Elmer Spectrum 100 Spectrofotometer. Melting points were measures in open capillary tubes with an Electrothermal-9200 melting points apparatus. Routine 1 H NMR and 13 C NMR spectra were recorded using a Bruker Avance AMX spectrometer operating at 300, 400 and 500 MHz for 1 H NMR, and at 75, 100 and 125 MHz for 13 C NMR in CDCl3 with tetramethylsilane as an internal reference. The NMR studies were carried out in high-quality 5 mm NMR tubes. Chemical shifts (δ) and coupling constants (J) are reported in ppm and in Hz, respectively. 1 H NMR spectra are referenced to residual protiated solvents (δ = 7.26 ppm for CDCl3), 13 C chemical shifts are reported relative to deuteriated solvents (δ = 77.16 ppm for CDCl3). The catalytic solutions were analysed with an Agilent 6890N GC and Schimadzu 2010 Plus GC-MS system by GC-FID with an HP-5 column of 30 m length, 0.32 mm diameter, and 0.25 μm film thickness.

  Procedure for the Preparation of BenzimidazoliumBromides 3a and 3b: A chloroform (10 mL) solution of 1-(2-(2ethoxyphenoxy)ethyl)benzimidazole (0.282 g, 1.0 mmol) and alkyl bromide (1.0 mmol) was stirred at 60 °C for 24 h. After completion of the reaction, the solvent was removed by vacuum and diethyl ether(15 mL) was added to obtain a white crystalline solid, which was filtered off. The solid was washed with Et2O (3 × 10 mL) and dried under vacuum. The crude product was recrystallized from EtOH/Et2O mixture (1:2, v/v) and dried under vacuum. δ = 1.33 (t, 3 J = 7.0 Hz, 3H, OCH2CH3), 2.27 (s, 6H, C6H3(CH3)2), 3.95 (q, 3 J = 6.9 Hz, 2H, OCH2CH3), 4.56 (t,3 J = 4.5 Hz, 2H, Six ruthenium(II) complexes with the general molecular formula [RuCl2(NHC)(η 6 -p-cymene)], (NHC = N-heterocyclic carbene) were synthesized. The obtained complexes were fully characterized by analytical and spectral (FT-IR, elemental analysis and 1 H and 13 C NMR) methods. The solid-state structure of one of the ruthenium complexes {dichloro-[1-(2-(2-ethoxyphenoxy)ethyl)-3-(3,5dimethylbenzyl)benzimidazol-2-ylidene](p-cymene) ruthenium(II)} has been established by single-crystal X-ray diffraction study. The catalytic activity of the all ruthenium(II) complexes have been evaluated in the N-alkylation of aniline using the green hydrogen borrowing methodology

Table 1 .

 1 Physical and spectroscopic properties of new compounds.

	Compound	Formula	Isolated yield (%)	M.p. ( o C)	v(CN) (cm	-1 )	H(2)	1 H NMR (ppm)	C(2)	13 C{ 1 H} NMR (ppm)
	3a	C26H29BrN2O2	87	178-179	1572		11.42	143.1
	3b	C29H35BrN2O2	83	169-170	1556		10.33	142.1
	4a	C26H28AgBrN2O2	81	206-207	1442		-		not observed [a]
	4b	C29H34AgBrN2O2	74	151-152	1451		-		not observed [a]
	1a	C36H42RuCl2N2O2	65	187-188	1452		-		191.2
	1b	C39H48RuCl2N2O2	57	165-166	1450		-		189.9
	1c	C30H38RuCl2N2O3	62	136-137	1452		-		190.6
	1d	C37H44RuCl2N2O2	70	191-192	1450		-		190.2
	1e	C38H46RuCl2N2O2	63	157-158	1454		-		189.9
	1f	C38H46RuCl2N2O2	68	159-160	1452		-		191.5

ppm (NCN), two characteristics of the formation of a Ru-carbene bond [22]. Scheme 1. Synthesis of ruthenium complexes 1a-f.

Table 3 .

 3 N-alkylation of aniline with benzyl alcohola search for optimal catalytic conditions.[a]

							Selectivity [%]
	Entry	1c [mol %]	Base	Time [h]	Temp. [ o C]	Yield [%]	A	B
	1	No	t BuOK	24	120	0	-	-
	2	5	Cs2CO3	24	120	32	89	11
	3	5	KOH	24	120	71	72	28
	4	5	t BuOK	24	120	100	85	15
	5	1	t BuOK	24	120	70	70	30
	6	2.5	t BuOK	24	120	100	80	20
	7	2.5	t BuOK	16	120	80	69	31
	8	2.5	t BuOK	24	90	74	73	27

Table 4 .

 4 Ruthenium-catalysed N-alkylation of aniline with arylmethyl alcohols.

	[a]

Table 5 .

 5 Ruthenium-catalysed N-alkylation of aniline with heteroaromatic alcohols.

	[a]

Acknowledgements

This work was financially supported by the Technological and Scientific Research Council of Turkey TÜBİTAK (112T303).

Keywords: N-Heterocyclic carbene • ruthenium • amine • Nalkylation • hydrogen borrowing strategy

temperature and filtered through a short pad of SiO2 and the filtrate was analyzed by and GC-MS with the calibrations based on decane. 17208(4) Å 3 ,Z = 18,F(000) = 7632,T = 173(2) K. The crystal size (0.160 × 0.140 × 0.100 mm) was studied with a Bruker APEX2 DUO Kappa-CCD diffractometer using Mo-Kα radiation (λ = 0.71073 Å). The structure was solved by using SHELXS-2013 [27], which revealed the non-hydrogen atoms of the molecule. After anisotropic refinement, all the hydrogen atoms were found by Fourier difference. The whole structure was refined with SHELXL-2013 [28] by using the full-matrix least-squares technique (use of F2; x, y, z, ij for C, Cl, N, O and Ru atoms, x, y, z in the riding mode for hydrogen atoms); 436 variables and 6199 observations with I > 2.0σ(I); calcd. w = 1/[σ 2 (Fo 2 ) + (0.0801P) 2 ] in which P = (Fo 2 + 2Fc 2 )/3. Extra solvent accessible voids were detected in 1aCHCl3, therefore, a SQUEEZE/PLATON technique [29] was applied to remove the unidentified solvent contributions. Final results: R = 0.0526, Rw = 0.1455, Sw = 1.057 and Δρ < 0.961 e Å -3 . CCDC 1580353 contains the supplementary crystallographic data for 1a. These data can be obtained free of charge via http://www.ccdc.cam.ac.uk/conts/retrieving.html, or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (+44) 1223-336-033; or e-mail: deposit@ccdc.cam.ac.uk.

X-ray Crystallographic

Supporting Information (see footnote on the first page of this article):

Full 1 H NMR, 13 C NMR and FT-IR spectra for the of benzimidazolium bromides (3a and 3b), silver(I)-NHC complexes (4a and 4b) and ruthenium(II)-NHC complexes (1a-f) .


Marie Perrin
email: mariexuanperrin@yahoo.fr

Aurore Schmitt

Yann Ardagna

From early to late modern societies (late 16th -early 20th century): Shifts in dental health status in two populations from southeastern France

Keywords: Dental health, early and late modern periods, Industrial Revolution, southeastern France, dental caries, hypoplasia, bioarchaeology

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

INTRODUCTION

In France, early and late modern societies have been scarcely investigated by archaeologists and have long been overlooked in bioarcheological research. However, this period covering five centuries, from the Renaissance to the 20 th century, witnessed extensive transformations, that would change population structure, leading to the dawn of contemporary industrialized societies. Changes occurred in socioeconomic areas as the Industrial Revolution took off, but also in political and cultural fields. These changes led to massive consequences on living conditions and health, as scientific and technical progress triggered major medical advances. Hence, such a context would perfectly illustrate the influences and constant interactions between cultural aspects, lifestyles and anatomy and biology. Indeed, the Industrial Revolution constitutes a major and profound shift on human biology, eventually transforming the pathologies from which populations suffer. Notably, the industrial period saw the onset of the epidemiological transition (Omran, 1983), characterised by a radical change in the pathocenosis (Grmek, 1969), modifying the epidemiological structure of populations. This slow process would involve the decline of all infectious and deficiency diseases, which were initially the main causes of death, in favour of chronic and degenerative diseases (Meslé and Vallin, 2000 ;Picheral, 1996).

As rescue archaeology developed, this period slowly entered the scope of bioarcheological studies. The recent excavation of two cemeteries in southeastern France has provided a large skeletal assemblage, including 1 777 individuals from primary burials. This sample offers a unique opportunity to investigate on a large scale the evolution of health during this particular period and specifically at the beginning of industrialization.

The study aims to define and compare the dental health of early and late modern populations in order to determine the impact on health and living conditions brought along by the Industrial Revolution. The many available archaeological and historical sources for such recent historical periods provide a fully documented context in which to interpret and discuss the biological analysis, thus enabling an inclusive bioarcheological approach, with the desire to understand "the causes and consequences of adaptative and behavioral shift in the [recent] past" (Larsen 1997), following carefully the evolution of the pathocenosis throughout History, and especially during this important transition period.

MATERIAL

The samples for this research come from two cemeteries, one in La Ciotat and the other in Marseille, two cities located in southeastern France and about 30 km distant from each other (Figure 1).

2.1.The selected populations

The first collection comes from the 2009 archaeological excavation of the Cimetière St Jacques in La Ciotat (LC HOP), mainly in use during the Ancien Régime, from 1681 to 1831 [START_REF] Richier | Bouches-du-Rhône[END_REF]. The assemblage includes 1245 individuals from primary burials but only 423 were included in this study because they were the ones that still retained their dentition.

The second collection is from the Cimetière des Crottes in Marseille (MPC), chiefly in use during the 19 th century, from 1784 to 1905 [START_REF] Richier | Bouches-du-Rhône, Marseille, prolongement de la ligne 2 du métro de Bougainville vers Capitaine Gèze -72 avenue Félix Zoccola. Ancien cimetière des Crottes. Rapport final d'opération préventive[END_REF]. The excavation led to the discovery of 532 individuals buried in wooden coffins of which 219 were included in this study because they had preserved dental remains.

Both populations are complementary chronologically, allowing the reconstruction of a complete sequence from the end of the 16 th to the early 20 th century. Besides, thanks to the stratification of the archaeological sites, distinct burial phases were defined and precisely dated. It was therefore possible to carefully study the changes over time in dental health indicators (Figure 1).

Both samples share common features: they are located in a close geographical area, they come from urban contexts and mainly represent modest socio-economic backgrounds. Demographic and economic take off occurred in both cities, in La Ciotat through maritime commerce and in Marseille during the 19 th century with the Industrial Revolution.

2.2.The bone assemblage

The complete sample studied includes 642 individuals, of which 430 are adults and 212 juveniles (aged < 20 years old). The age at death of adults was estimated through changes in the pubic symphysis [START_REF] Schmitt | Une nouvelle méthode pour discriminer les individus décédés avant ou après 40 ans à partir de la symphyse pubienne[END_REF] and auricular surface [START_REF] Lovejoy | Chronological metamorphosis of the auricular surface of the ilium: A new method for the determination of adult skeletal age at death[END_REF][START_REF] Schmitt | Une nouvelle méthode pour estimer l'âge au décès des adultes à partir de la surface sacro-pelvienne iliaque[END_REF]. For juveniles, age at death was evaluated through dental development (Moorrees et al., 1963a and b;Ubelaker, 1989), diaphyseal length [START_REF] Adalian | Nouvelle formule de détermination de l'âge d'un foetus[END_REF][START_REF] Maresh | Measurements from roentgenograms, heart size, long bone lengths, bone, muscles and fat widths, skeletal maturation[END_REF] and epiphyseal fusion [START_REF] Scheuer | Developmental juvenile osteology[END_REF][START_REF] Veschi | Recherches sur la collection d'enfants et d'adolescents d'âge et de sexe connus de Bologne (Italie): Diagnose de l'âge sur la base du degré de maturation osseuse[END_REF]. According to their estimated age at death, individuals were placed into one of the age categories listed in Table 1. Sex was determined for adults only, using coxal bones, through the combination of measurements defined in the DSP method [START_REF] Murail | DSP: A tool for probabilistic sex diagnosis using worldwide variability in hip-bone measurements[END_REF] and the five morphological criteria described by [START_REF] Bruzek | A method for visual determination of sex, using the human hip bone[END_REF]. Statistical discriminant function analyses were also performed on long bone measurements to assess sex when the hip bones were not preserved [START_REF] Murail | A new approach to sexual diagnosis in past populations. Practical adjustments from Van Vark's procedure[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales, interactions bio-culturelles[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales[END_REF]. Basic demographic information concerning the individuals under study is provided in Tables 1 and2.

In La Ciotat, three phases were outlined thanks to a very legible stratification of the cemetery backfill and dated through the study of grave goods and archival data. In Marseille, four phases were identified by the stratigraphic study, combined with the analysis of grave management and funerary organisation. They were dated via historical sources, and especially the dates of the successive extensions of the cemetery.

METHODS: DENTAL INDICATORS OF HEALTH

Dental status analysis is considered a valuable tool for the evaluation of health and seven markers or pathologies, routinely used in bioarchaeology, were selected and used to compare the two samples. Through their various aetiologies and pathogenesis, they altogether provide an overview of diet, biological stress, daily habits, hygiene and dental care, some of the key features of the health concept [START_REF] Rose | Histological enamel indicators of childhood stress in prehistoric skeletal samples[END_REF][START_REF] Goodman | The chronological distribution of enamel hypoplasias from prehistoric Dickson Mounds populations[END_REF][START_REF] Nourrisson | Tabagisme et antitabagisme en France au XIX e siècle[END_REF][START_REF] Irish | A Companion to dental anthropology[END_REF][START_REF] Hillson | Dental Anthropology[END_REF]2019). The methodology's originality relies on adapting the study of dental markers to early and late modern populations. Furthermore, medical, medical-historical or historical studies have contributed to preconceived ideas about the secular changes of these markers at the dawn of the 19 th century, which should be tested here thanks to the available bone assemblage, considered as a more direct source of information. Besides, the use of well-documented historical and archaeological sources available for such recent archaeological sites was necessary to the bioarcheological approach.

Dental wear, the progressive loss of the constituent tissues of the tooth, through abrasion, attrition and erosion (Burnett, 2016[START_REF] Hillson | Dental Anthropology[END_REF][START_REF] Scheuer | Developmental juvenile osteology[END_REF]2019 ;Kaidonis, 2008) was recorded according to the eight stages described by [START_REF] Smith | Patterns of molar wear in hunger-gatherers and agriculturalists[END_REF]. Its interest lies in the information it can provide on cultural practices [START_REF] Bocquentin | Pratiques funéraires, paramètres biologiques et identités culturelles au Natoufien : une analyse archéo-anthropologique[END_REF]Henderson and Walker, 2012;Kvaal and Derry, 1996), diet reconstruction and changes (Molnar, 1971;[START_REF] Smith | Patterns of molar wear in hunger-gatherers and agriculturalists[END_REF]Larsen, 1995), but also in its interaction with oral pathologies. Worldwide, the general drastic decrease in tooth wear would have occurred recently (Kaidonis et al., 1992). It will be interesting to document this possible change with a precise chronological frame and its possible impact on dental pathologies, in particular on caries.

Ante-mortem tooth loss (AMTL) was considered through the level of alveolar resorption. Despite various aetiologies (pathological, mechanical, curative or cultural), the quality of oral hygiene and access to care remain crucial factors (Larsen, 1997) which fully justify their use in this work.

Carious lesions are considered a privileged marker to reconstruct human feeding behavior as the nature of the diet remains the main aetiology (Hillson, 1979;Cohen and Armelagos, 1984). Indeed, carbohydrates are particularly cariogenic (Powel, 1985 ;[START_REF] Hillson | Dental Anthropology[END_REF]Lucaks, 2008 ;Hillson, 2018). Caries has been extensively used to track possible dietary changes, particularly with the consumption of cultivated cereals from the adoption of agriculture (Larsen, 1997). For modern periods, the major dietary changes mainly concern the increase of sugar consumption or processed food (Moore and Corbett 1975;[START_REF] Corbett | Distribution of dental caries in ancient British populations. IV-The 19 th century[END_REF]Flandrin and Montanari, 1996) and many anthropological studies have shown an increase in this type of lesion from the modern period onwards (Muller and Hussein, 2017). Carious lesions were recorded in accordance with [START_REF] Hillson | Recording dental caries in archaeological human remains[END_REF] and the protocol is presented in Table 3 [START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales, interactions bio-culturelles[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales[END_REF].

The presence of periapical lesions, usually resulting from dental caries, pronounced attrition or tooth fracture [START_REF] Dias | Abscess cavity'-a misnomer[END_REF][START_REF] Hillson | Dental Pathology[END_REF] was evaluated and the different types of lesions [START_REF] Dias | Abscess cavity'-a misnomer[END_REF] have been grouped under the generic term abscess.

Calculus formation is multifactorial, but anthropological studies have focused on two etiologies. The first one is the level of dental hygiene, since calculus deposit refers to the mineralisation of dental plaque [START_REF] Hillson | Dental Anthropology[END_REF]2019). Its recording could highlight possible dental hygiene practices, at a time when the use of toothbrushes and toothpastes are being introduced (Clément, 2008;2010).The second one lies in the nature of the diet: a high protein intake would promote its formation [START_REF] Hillson | Dental Anthropology[END_REF]2019 ;Lieverse, 1999) and the 19 th century is supposed to witness a growing increase in meat consumption (Flandrin, 1996c).

Periodontal disease is "an inflammatory condition that affects any or all portions of the periodontium" (Wasterlain et al., 2011), caused mainly by the presence of dental plaque [START_REF] Hillson | Dental Anthropology[END_REF]2018 ;Wasterlain et al., 2011 ;Nelson, 2016) and the aggravating factors are those that influence its presence, quantity and composition, including oral hygiene, diet or the presence of calculus. Calculus deposits and periodontal disease were recorded using the three stages defined by [START_REF] Brothwell | Digging up Bones: The excavation, treatment, and study of human skeletal remains[END_REF].

Finally, linear enamel hypoplasia (LEH) was assessed on each observable tooth, following [START_REF] Bocquentin | Pratiques funéraires, paramètres biologiques et identités culturelles au Natoufien : une analyse archéo-anthropologique[END_REF] Table 3). These defects form bands of decreased enamel thickness and provide a nonspecific indicator of stress, during tooth crown formation: during childhood. A wide range of aetiologies have been recognised, but the main ones are nutritional or vitamin deficiencies (chronic or episodic malnutrition or undernutrition) and infectious or parasitic diseases [START_REF] Goodman | Assessment of systemic physiological perturbations from dental enamel hypoplasias and associated histological structures[END_REF][START_REF] Hillson | Dental Anthropology[END_REF][START_REF] Schmitt | Une nouvelle méthode pour estimer l'âge au décès des adultes à partir de la surface sacro-pelvienne iliaque[END_REF]2018;Lewis, 2007). They are then often linked to socio-economic status and considered a good indicator of the health status of former populations.

Combining the number of linear defects on every tooth and their age of appearance, a minimal number of stress episodes was defined for each individual.

A population-based approach involving the calculation of frequencies per tooth and prevalence per individuals [START_REF] Waldron | Counting the dead, the epidemiology of skeletal populations[END_REF][START_REF] Waldron | Palaeoepidemiology: The measure of disease in the human past[END_REF] allows for a paleoepidemiological analysis. Comparisons between frequency and prevalence in the two samples relied on Chi² independent tests, Fisher's exact test and odd ratio analysis. To investigate the severity of the different pathologies and the number of stress episodes, mean scores of severity and mean number of episodes were created for each individual, then compared using Student's t test. Statistical analysis was performed using R.

RESULTS

4.1.Common features

In both samples, frequencies per teeth exhibit similar patterns [START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales, interactions bio-culturelles[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales[END_REF] following a classical distribution of dental lesions according to the different types of teeth [START_REF] Irish | A Companion to dental anthropology[END_REF][START_REF] Hillson | Dental Anthropology[END_REF]2019). Molars are the teeth most lost ante-mortem and most affected by carious lesions. The lower incisors bear a larger amount of the calculus deposits, particularly on the lingual surfaces, whereas canines and upper incisors mainly show LEH.

When comparing total frequencies, nearly every dental marker reveals differences (Table 4), suggesting that, except for AMTL and abscesses, dental pathologies are more frequent in the late modern sample of Le Cimetière des Crottes. Teeth from this population also exhibit less intense dental wear: stages 6 to 8 from the Smith recording system (1984) seem rarer. However, even if differences are statistically significant, frequencies remain quite close between both samples. These results, which are based on the calculation of frequencies per tooth, could be partially influenced by the potential presence of few individuals whose teeth bear many and/or severe pathologies. Hence, this set of data must be weighted, or rather supplemented by an individual-based approach. Therefore, we will retain differences only when expressed in both frequencies per tooth and prevalence per individual.

Considering the prevalence per individual, no difference is to be observed in the expression of the majority of dental markers between both samples (Figure 2; Figure 3). Moreover, the distribution of dental lesions according to age at death and sex are quite similar. Except for LEH, dental pathology prevalence and severity tend to increase with age (Figure 2; Figure 3; [START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales, interactions bio-culturelles[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales[END_REF]. In both samples, no significant difference between sexes was noticed regarding each dental indicator [START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales, interactions bio-culturelles[END_REF][START_REF] Perrin | État sanitaire entre Ancien Régime et révolution industrielle: Étude paléoépidémiologique de deux populations provençales[END_REF].

Results emphasize similar tendencies. Indeed, both samples exhibit very limited dental wear. Stages 2 and 3 (from [START_REF] Smith | Patterns of molar wear in hunger-gatherers and agriculturalists[END_REF] are the most common and mean scores by individuals are quite low (Figure 2). Both populations experienced a very large amount of AMTL. Around 30 % of teeth were lost ante-mortem (Table 4) and most adult individuals are concerned by this pathology (Figure 3). A very high presence of dental caries was recorded. Between 13 % and 23 % of teeth are affected (Table 4). Carious lesions develop early in life (from 1-4 years old in LC HOP, and before the end of the first year of life in MPC) and, when reaching adulthood, most individuals exhibit at least one dental caries (Figure 3). Paradoxically, dental abscesses are quite rare, considering the high rate of dental decay (Table 4), but they are well distributed among the adults. Thirty percent show at least one periapical lesion (Table 4). Calculus deposits and periodontal disease are frequent and age correlated, but no actual changes between early and late modern periods was observed (Table 4; Figure 3). Finally, LEH are very frequent: more than half of both groups are affected by this general stress marker (Figure 3).

4.2.Differences between the early and the late modern samples

Despite many common characteristics, significant differences were observed for three indicators in both analyses (per tooth and per individual).

Dental caries and dental wear

Results emphasize a significant increase in dental caries frequencies and prevalence (Table 4; Figure 3), associated with a decrease in the rate of dental wear (Table 4; Figure 3). A closer look at these two parameters shows how they changed over time.

When considering the distinct burial chronological phases, results show the constant rise of dental decay from the end of 16 th century to the beginning of the 20 th century (Figure 4). Meanwhile, a decrease in the severity of carious lesions simultaneously occurred. The mean score of severity, related to the extent of the lesion, declined and the proportion of cavities located on the dental crown tended to rise at the expense of those caries developing around the cervix (Figure 4). Finally, these shifts in the severity and location of dental caries are combined with a progressive reduction of dental wear (Figure 4). Moreover, these changes appear to have occurred simultaneously: they took place or intensified during the second phase of Le Cimetière des Crottes in Marseille, around the 1830-1850.

Linear enamel hypoplasia

The high frequencies and prevalence of LEH in both sites highlights the omnipresence of biological stressors, which increase even further during the 19 th century (Table 4; Figure 3). It may indicate the upsurge of stressful events during that period. Moreover, individuals from the 19 th century in the Cimetière des Crottes in Marseille, tend to have survived more stress episodes than those from the early modern period in La Ciotat (Figure 5b). When taking age at death into account, two differences stand out. Prevalence among adults tends to decrease with ageing in La Ciotat but remains stable in Marseille (Figure 5a). Considering juveniles, prevalence is higher for the youngest children (under 5 years old at death) in La Ciotat (Figure 5a). These differences may indicate a change in mortality patterns as a consequence of stressful events. Finally, the age of formation of LEH show that the earliest stress periods (before 3 years old) are significantly more frequent during the 19 th century.

DISCUSSION

Results mostly show similarities between both samples, which highlight common features in health status during the early and late modern periods in southeastern France. Hence, both populations' health was characterized by limited dental wear, many dental caries and LEH, high rates of AMTL, few abscesses and moderate levels of calculus and periodontitis. A few differences were also observed for dental wear, caries and LEH, revealing changes over time. However, these changes are evident on traits already documented for earlier periods and further accentuate some trends. Considering the etiologies of these dental indicators and including archaeological and historical data available, a number of assumptions can be discussed in terms of diet, dental care and the infectious environment. Cross-referencing these different kinds of data allows the construction of a precise framework to interpret the biological characteristics of health status at the time.

5.1.Everyday diet during the early and late modern periods

Ubiquitous cereal consumption

The high level of dental decay in both samples (Table 4; Figure 3) suggests that daily diet heavily depended on carbohydrates. Indeed, diet is a key factor in the development of carious lesions and carbohydrates are known to be particularly cariogenic (Powel, 1985;[START_REF] Lukacs | Fertility and Agriculture Accentuate Sex Differences in Dental Caries Rates[END_REF][START_REF] Hillson | Dental Anthropology[END_REF][START_REF] Hillson | Recording dental caries in archaeological human remains[END_REF]2019).

From modern times onward, subsistence has massively relied on a single type of resource: cereals, consumed mainly as bread, gruels or porridge (Flandrin, 1996a;[START_REF] Lachiver | Les années de misère: La famine au temps du Grand Roi[END_REF][START_REF] Morineau | Croître sans savoir pourquoi: structures de production, démographie et rations alimentaires[END_REF]. This prominence reflected new feeding strategies, in relation to the European demographic expansion of the end of the Middle Ages [START_REF] Morineau | Croître sans savoir pourquoi: structures de production, démographie et rations alimentaires[END_REF]. Agriculture mostly turned to cereal production because it provided more calories per equal surface of allocated land than did animal husbandry, hunting or gathering (Flandrin, 1996a).

During the 19 th century, cereals were still essential in everyday diet. While the structure of the food portion underwent some modifications, the caloric intake first increased only through the consumption of cereals and other starchy foods (Flandrin, 1996b and c). Hence, the highest cereal consumption in France was reached between 1885 and 1894 (Flandrin, 1996c).

Paradoxical subsistence strategies: diet and stress episodes

Such a diet, mainly based on one type of nutrient, could have noxious consequences both from a qualitative and quantitative perspective. Poor diet, one of the main etiologies for the formation of LEH, could explain the widespread occurrence of stress periods in the populations under study, illustrated by the high frequencies of these linear defects (Table 4; Figure 3).

Under normal circumstances, food should have been sufficient thanks to new feeding strategies and the progress in agricultural practices. Nevertheless, the popular diet remained little diversified for both the early and late modern periods, which were still marked by monotonous meals. This kind of diet would not have met sufficiently the body's needs [START_REF] Lachiver | Les années de misère: La famine au temps du Grand Roi[END_REF]Flandrin, 1996 b and c) and this widespread phenomenon of malnutrition could explain the high prevalence of LEH [START_REF] Goodman | Nutritional supplementation and the development of linear enamel hypoplasias in children from Tezonteopan, Mexico[END_REF].

The high consumption of cereals in everyday diet can have dramatic effects, leading to periods of food shortages when harvests were poor. During the early modern period, food crises reached an unprecedented scale [START_REF] Lachiver | Les années de misère: La famine au temps du Grand Roi[END_REF]Flandrin, 1996a and b). The subsistence economy of the Ancien Régime, influenced by inefficient agronomic techniques and founded on self-based consumption, prevented long-term storage [START_REF] Lachiver | Les années de misère: La famine au temps du Grand Roi[END_REF]. Inadequate means of transport complicated the supply chains during shortage events. The end of the 17 th and beginning of the 18 th centuries were periodically marked by several episodes of drought, heavy rains, late frosts, etc. For instance, the severe winters of 1680 and 1709, directly mentioned in the archives of La Ciotat [START_REF] Reboul | La Ciotat: Essai de géographie urbaine[END_REF], had a serious impact on harvests, causing periods of famine, which in turn caused terrible epidemics. These food crises, followed by epidemic outbreaks, had a strong effect on morbidity and mortality [START_REF] Lachiver | Les années de misère: La famine au temps du Grand Roi[END_REF]Flandrin, 1996a and b). They could explain the recurrence of stress episodes experienced by the inhabitants of La Ciotat and materialized in LEH. More than 60% of the people buried in the Saint-Jacques cemetery display these defects (Figure 3). However, by the 19 th century, the subsistence economy gradually disappeared in favor of a market economy. With the onset of global commerce, promoted by the transport revolution (steam navigation, extensive railway networks) and agricultural advances, these periods of shortage tended to end (Flandrin, 1996c;[START_REF] Daumalin | Marseille (1831-1865). Une révolution industrielle entre Europe du Nord et Méditerranée[END_REF][START_REF] Bertrand | Histoire d'une ville : Marseille[END_REF]. Other factors may explain the predominance of LEH and their increasing frequencies. Among them, the social and financial status of the population must certainly be taken into account. Despite the shift towards a market economy, both populations under study mostly came from the lowest or modest classes, who may have not had the financial means to flourish yet and did not benefit from the positive effects of the growing economy.

5.2.The Industrial Revolution and technical progress

Introduction and consumption of new products

The synchronized changes in dental caries and dental wear chronologically coincide with the early to mid-19 th century, the take-off of the Industrial Revolution in Marseille. This period was marked by important changes in everyday diet (Figure 4). With the development of more modern transports and world trade, new foodstuffs were introduced and distributed on a large scale, especially overseas products rich in carbohydrates, such as potatoes or refined sugar (Flandrin, 1996c). While potatoes would only become popular in central and northern Europe (Flandrin, 1996c), refined sugar had a different fate worldwide and particularly in southeastern France. Sugar derived from the transformation of sugar cane, imported from the West Indies, appeared in France during the 17 th century, but remained an expensive commodity, reserved for the highest spheres (Flandrin, 1996b). Until the 1830s, its consumption remained quite limited (Huetz de [START_REF] De Lemps | Boissons coloniales et essor du sucre[END_REF][START_REF] Daumalin | Marseille (1831-1865). Une révolution industrielle entre Europe du Nord et Méditerranée[END_REF]. However, thanks to the transformation of beetroots, progressively produced in greater amounts and increasingly popular in northern France, the price of refined sugar decreased drastically. Marseille became the largest sugar cane warehouse and refineries multiplied: sugar was no longer a luxury product, progressively freeing the offer and demand [START_REF] Daumalin | Marseille (1831-1865). Une révolution industrielle entre Europe du Nord et Méditerranée[END_REF]. In France, in only 70 years, sugar consumption exploded: from 1.2 kg per individual in 1815 to 12 kg in 1894 (Flandrin, 1996c).

Over the long term, starch slightly decreases buccal acidity. Its regular consumption favors the development of carious lesions, leading to moderate prevalence [START_REF] Hillson | Dental Anthropology[END_REF], as obtained at La Ciotat. However, sugar leads to a dramatic drop of the oral pH, but not necessarily over a long time. Hence, the combination of both types of food (starch and sugar), could explain the considerable rise in the prevalence of caries [START_REF] Hillson | Dental Anthropology[END_REF] observed in the Marseille sample. Such an increase in caries from the 18 th century has been documented across all of Europe and is associated with the advent of sugar [START_REF] Müller | Meta-analysis of teeth from European populations before and after the 18th century reveals a shift towards increased prevalence of caries and tooth loss[END_REF].

Finally, one last overseas product must have had great consequences on dental health: tobacco. At first, the prerogative of men, it was consumed in particular through the pipe. Its success, from the second half of the 19 th century onwards, was due to a new medium: the cigarette, dating back to 1830 and gradually spreading to all levels of society including women [START_REF] Nourrisson | Tabagisme et antitabagisme en France au XIX e siècle[END_REF]2010). Throughout the 19 th century, tobacco consumption exploded, rising from 9000 tons in 1815 to 48000 tons in 1914 [START_REF] Nourrisson | Tabagisme et antitabagisme en France au XIX e siècle[END_REF]. Smoking increases the risk of certain oral pathologies, particularly caries, periodontosis and abscesses (de Araùjo Nobre and Malo, 2017). The increase of tooth decay and to a lesser extent of periodontosis coincided with the expansion of cigarette smoking and, more generally, with tobacco consumption in France (Figure 4; Table 4).

New production techniques

Alongside the introduction of new dietary products, industrialization innovated with new techniques for food production, especially in refineries and flour mills. As a result, food and more specifically flour were more refined, producing a less abrasive food and influencing the rate of dental wear. In Marseille, the technological modernization of grain milling occurred between 1830 and 1848 [START_REF] Daumalin | Les difficiles débuts de la minoterie à vapeur à Marseille (1779-1835) : Blocage technologique ou problème de marché ?[END_REF]. This period seems to match the decrease of dental wear documented in the studied samples (Figure 4). These new food processing techniques marked the end of the regularly observed competition between dental wear and caries [START_REF] Hillson | Recording dental caries in archaeological human remains[END_REF]. Indeed, heavy wear obliterates morphological irregularities on posterior teeth and erases early lesions as the tooth crown is progressively destroyed. Moreover, hard food is a protective and cleansing asset through attrition and the stimulation of saliva production [START_REF] Corbett | Distribution of dental caries in ancient British populations. IV-The 19 th century[END_REF]. In Provençal samples, the progressive decrease of dental wear may have played a part in the gradually rising prevalence of dental caries, in the decreasing severity of these lesions and also in the type of lesion itself (Figure 4). Indeed, data show a shift in the initiation site: a relative regression of lesions located around the tooth cervix, in favor of those developing on the crown. This gradual change over time has already been observed elsewhere [START_REF] Corbett | Distribution of dental caries in ancient British populations. IV-The 19 th century[END_REF][START_REF] Hillson | Recording dental caries in archaeological human remains[END_REF][START_REF] Müller | Meta-analysis of teeth from European populations before and after the 18th century reveals a shift towards increased prevalence of caries and tooth loss[END_REF]. Cervix area lesions are mainly characteristic of older periods, when tooth wear was still high. In populations where wear is less intense, caries more likely initiate in occlusal fissures and in interproximal contact areas (Figure 4). Such patterns of caries and dental wear have already been identified in England in a similar time frame [START_REF] Corbett | Distribution of dental caries in ancient British populations. IV-The 19 th century[END_REF], where this transition was initiated at the end of Middle Ages, asserted during the 17 th century and intensified during the 19 th century.

5.3.Industrial Revolution: evolution of curative and social practices

Dental health and hygiene practices

From the 18 th century, dentistry witnessed many medical improvements [START_REF] David | Hygiène bucco-dentaire du XVII e au XIX e siècle en France[END_REF]. However, the high rate of carious lesions may also indicate limited dental hygiene. Furthermore, no evidence of dental treatment was found in both sites, in contrast to other contemporary contexts [START_REF] Valentin | Anthropologie, pathologie et soins dentaires au XVIII e siècle: Découverte exceptionnelle à Saint-Martin-des-Champs de Paris[END_REF][START_REF] Molleson | The Spitalfields Project[END_REF][START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF][START_REF] Miles | St Marylebone Church and Burial Ground in the 18th to 19th centuries: Excavations at St Marylebone School 1992 and 2004-6[END_REF]. In those cemeteries, prostheses, dentures, or fillings sometimes made of expensive materials have been discovered. The social composition of samples under study may likely explain this absence: the relatively modest populations buried in the Saint-Jacques and Les Crottes cemeteries had probably little access to this type of sophisticated care.

In both series, AMTL are the only element suggesting dental care. In ancient populations, caries and wear were the major causes for tooth loss [START_REF] Lukacs | The 'caries correction factor': A new method of calibrating dental caries rates to compensate for antemortem loss of teeth[END_REF]. Considering the high prevalence of caries and the low wear rate observed in the studied samples, tooth loss might be mainly related to carious lesions. Besides, severe caries are rare and AMTL were also observed on young individuals (Figure 3). Most serious lesions could have led to tooth extractions for therapeutic purposes given that caries unlikely cause a spontaneous tooth loss [START_REF] Hillson | Dental Pathology[END_REF]. Tooth removal could therefore have been seen as the only effective and affordable dental pathology treatment.

New feeding practices for young infants

During the 19 th century, women started to work outside the domestic sphere more often and this called for new arrangements, such as the increased need of nurses for the youngest children and babies. Initially reserved for the elites of the 17 th and 18 th centuries, the employment of child nurses became a common practice and gradually spread to the lowest social categories [START_REF] Rollet | Allaitement, mise en nourrice et mortalité infantile en France à la fin du XIX e siècle[END_REF][START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF][START_REF] Fildes | Breasts, bottles and babies: A history of infant feeding[END_REF][START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF]. Wet nurses were quite expensive and it is more likely that in the working-class environment of Provence, dry nurses were privileged [START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF]. Moreover, breastfeeding declined across Europe either as a fad or out of necessity [START_REF] Riordan | Basics of breastfeeding. Part I: Infant feeding patterns past and present[END_REF]. This phenomenon could generate a new kind of systemic stress as it would demand artificial feeding and early weaning.

Artificial breastfeeding was being developed and recipes varied from one mother/nurse to another. They may have been largely unsuitable for infant feeding: often a mixture of groats and animal milk [START_REF] Rollet | Allaitement, mise en nourrice et mortalité infantile en France à la fin du XIX e siècle[END_REF][START_REF] Riordan | Basics of breastfeeding. Part I: Infant feeding patterns past and present[END_REF][START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF]. In Marseille, sweet bread soup seems to be preferred, replacing milk, often adulterated in this region where it is not produced [START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF].

Those practices might have created early biological stress as the artificial "milk" or weaning food (that is mainly water and flour) did not meet the nutritional needs of infants. This could be one of the recurrent childhood stressors during the 19 th century, illustrated by the increase of both frequencies in LEH and in individually-experienced stress episodes (Figure 5). Starch-rich weaning food and artificial [START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF][START_REF] Sorcinelli | L'alimentation et la santé[END_REF] provide many calories, but few of the essential elements for infant survival (proteins, vitamins, calcium, minerals) and therefore would cause nutritional defects. Moreover, hand feeding was spreading while at the same time sterilization and pasteurization, supposed to eliminate the pathogens responsible for many deaths, were neither mastered nor generalized and caused serious health issues [START_REF] Rollet | Allaitement, mise en nourrice et mortalité infantile en France à la fin du XIX e siècle[END_REF][START_REF] Riordan | Basics of breastfeeding. Part I: Infant feeding patterns past and present[END_REF][START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF].

Moreover, the mean age at weaning seemed to decrease in Provence and generally across Europe [START_REF] Rollet | Allaitement, mise en nourrice et mortalité infantile en France à la fin du XIX e siècle[END_REF][START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF]. The development of LEH has often been related to stress occurring during the weaning process [START_REF] Moggi-Cecchi | Enamel hypoplasia and age at weaning in 19 th century Florence, Italy[END_REF]. In the Provençal assemblages, early weaning or artificial breastfeeding could partly explain the shift seen in the age at which episodic stress events appear (Lewis, 2002a). Indeed, LEH formed between 6 months and 3 years were more frequent in the 19 th century assemblage (24 %) compared to the Ancien Régime one (18.7%; Figure 5).

Furthermore, an early introduction of carbohydrate-rich cereals or starchy food easily explains the high prevalence of caries on deciduous teeth and the increasing prevalence among the youngest children in the population of the Cimetière des Crottes (Table 4; Figure 3). Finally, artificial breastfeeding could deprive the newborn of colostrum, rich in vitamin A and important for developing immune protection especially against infectious diseases [START_REF] Rollet | Allaitement, mise en nourrice et mortalité infantile en France à la fin du XIX e siècle[END_REF][START_REF] Cox | Life and death at Spitalfields, 1700-1850[END_REF][START_REF] Sorcinelli | L'alimentation et la santé[END_REF].

5.4.Industrial Revolution: evolution of the infectious environment

In this general context of malnutrition, which seems to initiate early in the studied population (from early childhood), metabolic /deficiency or infectious diseases could easily spread [START_REF] Sorcinelli | L'alimentation et la santé[END_REF]. Infectious diseases and epidemic outbreaks, particularly prevalent in the modern periods, are a main cause of death [START_REF] Harrison | Disease and the Modern World: 1500 to the Present Day[END_REF]. Until the mid-19 th century, Marseille was still marked by a high infant mortality rate and infectious diseases accounted for at least 30 % of the deaths of the entire population [START_REF] Aubert | La médecine à Marseille au XIX e siècle[END_REF][START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF]. Children are especially vulnerable to this infectious context. The high prevalence of hypoplasia could be indirect evidence for systemic stress caused by infectious diseases. Indeed, infections remain a key factor for the development of LEH [START_REF] Goodman | Assessment of systemic physiological perturbations from dental enamel hypoplasias and associated histological structures[END_REF][START_REF] Hillson | Dental Anthropology[END_REF]2019;[START_REF] Lewis | Paleopathology of children[END_REF] and the general environment, witnessing massive urban and commercial growth, could have favored their development and spread.

Urbanization

Both populations come from urban contexts. In La Ciotat, urban development appeared quite rapidly at the beginning of the 16 th century, but before the end of the 18 th century the city declined soon after its apogee, and the population fell drastically [START_REF] Reboul | La Ciotat: Essai de géographie urbaine[END_REF][START_REF] Deidier | Le terroir de La Ciotat. Sa géographie et son histoire[END_REF][START_REF] Deidier | Histoire de La Ciotat : Des origines à nos jours[END_REF] [START_REF] Reboul | La Ciotat: Essai de géographie urbaine[END_REF][START_REF] Deidier | Le terroir de La Ciotat. Sa géographie et son histoire[END_REF][START_REF] Deidier | Histoire de La Ciotat : Des origines à nos jours[END_REF][START_REF] Castrucci | Les habitats du faubourg et les maisons à double affectation[END_REF]. In this context, urbanization could then have had a limited or short-term effect on morbidity or mortality. Moreover, studies by Lewis (2002a and b) tend to minimize the influence of urbanization on health before the industrial period in England.

However, the development of cities and their transformation during the Industrial Revolution could have had a huge impact on the infectious environment, through an immense and unprecedented concentration of people in urban centers. Such a situation would favor the transmission and spread of pathogens. The city of Marseille experienced significant demographic growth, due to rural exodus and immigration [START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF][START_REF] Temime | Histoire de Marseille de la révolution à nos jours[END_REF][START_REF] Dell'umbria | Histoire universelle de Marseille: De l'an mil à l'an deux mille[END_REF][START_REF] Daumalin | La grande mutation 1815-1870[END_REF]. Although rapid, urban development was slower than the pace of the massive influx of new people, leading to overcrowded houses in the old town and to the hasty creation of new districts on the outskirts, built without any consideration for hygiene [START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF]. The infrastructure, especially for the water supply and drainage, could barely cope with such an expansion [START_REF] Gaillard | La Vie quotidienne des ouvriers provençaux au XIX e siècle[END_REF][START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF]. This urbanization, of an unprecedented scale in History, certainly played a role in the frequency of epidemics [START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF] and in the increased prevalence of LEH.

Globalization

In urban centers international trade expanded exponentially with the Industrial Revolution. While the success of La Ciotat, during the 17 th century relied on its commercial exchanges with other Mediterranean cities [START_REF] Reboul | La Ciotat: Essai de géographie urbaine[END_REF][START_REF] Deidier | Le terroir de La Ciotat. Sa géographie et son histoire[END_REF][START_REF] Deidier | Histoire de La Ciotat : Des origines à nos jours[END_REF], the expansion of colonial empires and the development of means of transport, during the 19 th century, opened a new and significant perspective for a globalized and massive trade. National and international exchanges intensified in Marseille [START_REF] Daumalin | Les difficiles débuts de la minoterie à vapeur à Marseille (1779-1835) : Blocage technologique ou problème de marché ?[END_REF] : the flow of goods, but also of people, carriers of diseases, was more fluid, therefore stimulating the spread of pathogens.

The city faced the arrival of new diseases, such as cholera, which broke out in 1834 and became almost endemic in between periodical and regular massive epidemics [START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF]. However, cholera was not the only epidemic disease present: there were also smallpox (which was also endemic), typhus, typhoid fever, influenza, diphtheria and measles [START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF], as well as residual diseases such as tuberculosis. This specific context would promote both the apparition of new pathogens and their transmission, which could explain the rising frequencies of LEH, revealing increased childhood stress. It could also account for the increased number of periods of biological stress experienced during the industrial transition for those individuals who had reached adulthood. Finally, the virulence of these epidemic diseases could also explain the lower survival rate of children under 4 years old in Les Crottes, who died before hypoplastic lines formed (Figure 5). Even though the first campaigns of vaccination against smallpox took place in Marseille in 1801, it was incomplete and would have had very little effect on morbidity and mortality [START_REF] Aubert | La médecine à Marseille au XIX e siècle[END_REF][START_REF] Mafart | Les épidémies à Marseille au XIX e siècle[END_REF].

CONCLUSION

This study presents a first set of references to characterize the post-medieval health status of French populations. Large and significant differences between the early and late modern samples were expected and were supposed to highlight the changes occurring during the Industrial transition. However, health was also characterized by common features in both populations. The observed differences were less obvious than originally presumed and concerned mostly dental decay, dental wear and hypoplasia. Results rather display a continuity of dental health features than a sharp break. Moreover, the specific characteristics of the health status observed were first established during the early modern period and only exacerbated during late modern times. In the light of these new results and concerning the secular changes of health in Provence, it would seem more relevant to refer to an industrial transition, rather than an Industrial Revolution.

Technical and medical progress, social changes, public and sanitation policies set during the 19 th century are often considered one of the main factors improving living conditions. However, this study shows that far from improving, health seemed to deteriorate in southeastern France. Indeed, large hygienist investment might first benefit to the upper classes of the society. Moreover, long debated, health and hygiene measures would effectively be introduced and enforced only at the end of the 19 th century, having real and large consequences only from the beginning of the 20 th century onwards.

The authors have no competing interests to declare This research did not receive any specific grant from funding agencies in the public, commercial, or notfor-profit sectors.