Xiongbin Wang

Junjie Hao

Jiaji Cheng

Junzi Li

Jun Miao

Ruxue Li

Yiwen Li

Jiagen Li

Yuhui Liu

Xi Zhu

Yanjun Liu

Xiao Wei Sun

Zikang Tang

Marie-Hélène Delville

Tingchao He

Rui Chen

Chiral CdSe Nanoplatelets as an Ultrasensitive Probe for Lead Ion Sensing

As opposed to traditional photoluminescence and ultra-violet based optical sensing, we present here a sensing system based on resolved optically active polarization with promising applications. It is based on the of ultrathin CdSe nanoplatelets (NPLs) when modified with either L or D -cysteine molecules (L/D-cys) as bio-to-nano ligands. The chiral ligand transfers its chiroptical activity to the achiral nanoplatelets with an anisotropy factor ~10 -4 , which unlocks the chiral exitonic transitions and allows lead ions detection with a limit of detection (LOD) as low as 4.9 nM. Simulations and modelling based on timedependent density function theory (TD-DFT) reveal the chiral mechanism of L/D-cys capped CdSe NPLs. The presented CDbased sensing system illustrates an alternative possibility of using chiral CdSe NPLs as competitive chiral sensors for heavy metal ion detection.

Introduction

Colloidal semiconductor nanoparticles (NPs) with chirality have attracted tremendous interest due to their unique electronic and optical properties, which open the floodgate for novel sensitive chiroptical platforms as compared to their pristine achiral counterparts. [1][2][3][4] Generally, the induction of chirality in semiconductor NPs can be classified as: (i) NPs with intrinsically chiral dislocations and defects, [5][6][7][8] (ii) chiral surfaces formed by the chiral ligands or chiral interactions between chiral ligands and achiral semiconductor core, [9][10][11][12] and (iii) chiral assemblies consisting in achiral nanocrystals. [13][14][15][16][17][18] For instance, G. Markovich's group has accomplished enantioselective synthesis of intrinsically chiral mercury sulfide NPs. [8] More recently, N. A. Kotov et al., 19 reported the preparation of chiral tungsten oxide hydrate NPs with chirality transferred from the chiral ligands proline (Pro) and aspartic acid (Asp) to the tungsten oxide NPs via bio-to-nano chirality transfer. Such NPs can facilitate the formation of peptide bonds, leading to Asp-Asp and Asp-Pro dipeptides. Moreover, M. Liu and his colleague reported the synthesis of chiral assembled perovskite nanostructures through a supramolecular self-assembly approach and the perovskite NPs with circularly polarized luminescence (CPL) were successfully endowed. However, semiconductor NPs with intrinsic chirality or assembled chiral geometries often suffer from chemical laborious procedures or sophisticated fabrication processes, leading to large scale, high cost and timeconsuming production. Currently, the most universal method is the chirality transfer which is based on the electronic interactions between chiral molecules and the achiral semiconductor core. As a result, the intriguing circular dichroism (CD) peaks occur in the vicinity of the absorption bands of the colloidal NPs. 21 Recently, chiral ligand capped semiconductor nanodots, [22][23][24][25] nanorods 10,26 and NPLs [27][28][29][30] as well as their core-shell nanostructures have been extensively studied. Like in the case of metallic plasmonic NPs such as Au or Ag, the strong coupling between chiral ligand and achiral semiconductor can induce subsequent chirality transfer from molecules to semiconductor core with an anisotropic factor close to 10 -4 . 2,4,[31][32][33] Such an emerging property can easily be tuned by a simple control of the size, composition and shapes of the semiconductor NPs. Therefore, it could provide great potential for applications in chiral catalysis, 19,34 stereoselective synthesis, 8,35 and more importantly, in chiral sensing. [36][37][38][39][40] Heavy metal ions spread over in water sources do significantly affect human health and global environments. It is necessary to develop reliable and low-cost methods with high sensitivity for their detection as well as remediation. The most utilized approach for heavy metal ions detection up-to-now has been fluorescence-based color probe, during which semiconductor NPs would undergo emission quenching with the introduction of heavy metal ions. [41][42][43][44][45][46][47] These ions exfoliate capping ligands around the semiconductor NPs interact with their surface and generate a photoluminescence "on-off "sensing detectable by a colorimetric method. However, such a fluorescence detection strategy suffers from limitations such as the lack of specificity of the fluorescent carrier materials and their relatively low sensitivity in water. However, using the emerging chirality in semiconductor NPs induced by capping ligands could be a suitable alternative for a fast and accurate heavy metal ion sensing thanks to the follow-up of the chiral response of the NPs while interacting with the targeted ions. The detection limits and sensitivity are then sensed via CD spectra since CD signals are quenched with increasing metal ions concentration. In addition, compared to conventional CdSe quantum dots or quantum rods, 2D CdSe NPLs with few layers exhibit unique absorption and emission properties, including extremely narrow and separated absorption peaks resulting from heavy and light hole excitations. Herein, we synthesized cysteine-capped four layers CdSe NPLs via proper ligand exchange reaction between oleate anion and chiral cysteine molecules. The chirality transfer was monitored via electronic CD measurements and the chiral genesis was investigated through theoretical modelling based on the density functional theory (DFT). Finally, the as-synthesized chiral CdSe NPLs were applied as a chiral probe to detect lead ion, and it shows high sensitivity with a LOD close to 4.9 nM. The strong interaction between the thiol groups and lead ions results in a high selectivity for chiral sensing among seven kinds of metal ions. The linear relationship between the CD intensity, the gfactor and the ion concentrations demonstrates an ultrasensitive lead ion detection based on such a system. These results facilitate scalable chiroptical semiconductor nanostructures toward potential applications in chiral sensing, chiral separations and design of chiral devices.

Results and discussion

First, the zinc-blende (ZB) CdSe two-dimension (2D) NPLs were synthesized directly from cadmium myristate, during which oleic acid (OA) was used as the capping agent to avoid aggregation. The CdSe NPLs with few atomic layers were prepared undergoing precursor self-assembling followed with successive purification processes (Fig. 1). The transmission electron microscope (TEM) image (Fig. 1b) presents the 2D morphology of NPLs with an average size of 5.1±1.4 nm in width and 25.1±2.7 nm in length, (Fig. S1). Fig. 1c shows the highresolution TEM image, where the 3.23 Å lattice spacing corresponds to the (111) adjacent plane for ZB crystal structure can be clearly seen. The powder X-ray diffraction (XRD) patterns also indicate that the CdSe NPLs are in ZB phase. Fig. 1d shows the sharp reflection at 2θ = 25 o , 42 o and 49 o , which can be assign to the (111), ( 220) and (311) planes of ZB phase, respectively. The results are consistent with previous reports, which demonstrate the appropriate growth CdSe NPLs. The most intriguing properties for 2D NPLs could be attributed to their unique optical response. The absorption spectrum of CdSe NPLs capped with OA (Fig. 2d) exhibits two sharp excitonic peaks located in 484 and 513 nm. These two excitonic transitions are induced by different recombination, namely the light hole-electron for the higher energy, and the heavy holeelectron one for the lower energy absorption. The full widths at half maximum (FWHM) are 14 and 9 nm for light and heavy hole bands, respectively. The extremely narrow peaks are attributed to the strong quantum confinement effect in ultrathin 2D nanostructures. The observed absorption peaks indicate the thickness of NPLs herein is around four monolayers according to previous studies. 27,49 After the preparation and characterization of OA-capped NPLs, the subsequent ligand exchange reaction was performed using either the L-or D-form of the chiral cysteine (Fig. S2). Due to the thiol groups in cysteine molecules, a strong Cd-S bond is formed Please do not adjust margins Please do not adjust margins and replaces the pre-existing interaction of the CdSe surface with OA molecules (Fig. 2a-2c). After the ligand exchange, a redshift in the absorption spectrum for both light and heavy holeelectron transitions is observed to 498 and 528 nm respectively. The red-shift originates from the partial delocalization of the carrier wave function from the NPLs to the ligands. 27 Besides, the evident broadening of the two excitonic peaks demonstrates the change of ligand density, as well as the dipole-dipole coupling after ligand exchange and the new aqueous dispersion. Electronic CD measurements were then performed to investigate the chiroptical response of the cysteine functionalized CdSe NPLs. Indeed, bipinnate appearance of CD signals with multiple peaks arises in the vicinity of the exciton absorption of the NPLs, and a nearly mirrored CD line-shape is recorded when L-and D-cys are used as surface ligands respectively (Fig. 2e, black and red lines). Whereas in contrast, when achiral ligand (OA) or equal amount of each enantiomer of cysteine (enantiomer excess, ee =0) is used, the CD observations exhibit weak or even silent responses (pink and blue lines). All these results indicate that the induced chiroptical properties are mainly attributed to the chirality transfer from the chiral surface ligand to the achiral semiconductor core. Specifically, at the band-edge position close to 520 nm, the obtained CD shows the strongest value of the signal with an anisotropy factor close to 3×10 -4 , which is in good agreement with previous studies. 6,10 To clarify the CD line shape and the chirality transfer mechanisms in these NPLs nanostructures, simulations about the molecular structure and orbital wave function in hybrid systems based on TD-DFT method were performed. The ground state geometries of L-cys and D-cys capped CdSe nanoclusters are shown in Fig. S3. Fig. 3a and 3b illustrate the optimized ground state geometries of L-cys and D-cys capped CdSe nanoclusters, which were utilized as the prototypes with much smaller size than in reality. All the calculations were conducted in red hat linux system. We applied four ligands to incorporate to the bare CdSe nanocluster with ZB crystal structure. For the chiral activity transfer mechanism, the surface chiral ligand would hybridize with the electronic state of achiral inorganic nanoparticle core (the excitonic band in this system), leading to the strong wave function overlap in orbital coupling. The calculated frontier molecular orbitals hybridized in the lowest unoccupied molecular orbitals (LUMO) and highest occupied molecular orbitals (HOMO) for L-cys capped CdSe nanocluster are shown in Fig. 3c Please do not adjust margins Please do not adjust margins organic ligand capped complexes. In contrast, the LUMO exhibits missing overlaps of the wave function of nanocluster and chiral ligands. Fig. S4 shows similar electronic state coupling tendency for the HOMO-1 to HOMO-3 and LUMO+1 to LUMO+3. These observations are consistent with previous simulation results for CdSe quantum dots and quantum rods. 10,25 Therefore, the intrinsic chirality of bare nanoclusters were ruled out. The induction of chirality onto the achiral CdSe nanocluster could then be ascribed to the orbital coupling effect. Based on the discussions above, it is noted that the chirality of NPLs is due to the introduction of chiral ligand. The affinity of lead for sulfur is such, that once the external lead ions are introduced, the Pb-S bonding forms reducing the amount of chiral cysteine linked to the CdSe NPLs surface and inducing a quenching of the CD signal. The higher concentration of lead ions then results in a larger chiral signal quenching through weakening the electron state interaction between cysteine and CdSe NPLs. This provides a possible prerequisite for construction of highly sensitive heavy metal ion sensing. In this work, we utilized the lead (II) phosphate, Pb3(PO4)2, as the lead source for detection. All the aqueous analyte samples were adjusted into pH = 10 environment. Fig. 4a exhibits the CD peak response after adding various concentrations of lead ions. Their chiroptical activity showed significant decrease with increasing lead ion concentrations. The CD intensity around 520 nm in L-cys capped CdSe NPLs decreased from 20 mdeg to less than 1 mdeg under lead ions concentration between 0.01 to 1 μM. Similarly, the D-cys capped NPLs shows a signal decrease from 27 mdeg to less than 1 mdeg with the same ion concentration procedures. The UV-vis. absorption spectra in the meantime only exhibit a slight variation of the absorption intensity with the same ion concentration inductions (Fig. 4b). Fig. 4c reveals that chiral NPLs exhibit a clear linear relationship of CD intensity response at 520 nm versus logarithmic lead ion concentration in the range of 0.01 to 1 μM, with correlation coefficients of 0.998 and 0.994for L-and D-cys capped NPLs, respectively. The LOD for lead ion sensing is determined to be 4.9 ± 0.3 nM, from the straight line obtained in Fig. 4c.

To probe the specificity of the lead ion detection, we prepared a large enough quantity of NPLs starting material in one single batch to be able to also test six other metal ions at a 0.1 μM concentration for a comparison purpose. As shown in Fig. 4d, compared to the control sample (CD intensity at around 520 nm (L-cys capped NPLs) 33 mdeg), the addition of Ca 2+ , Mg 2+ , Fe 3+ , Fe 2+ resulted in the decrease of CD intensity in different extents in a range less than 15% of the control. The L-cys capped NPLs solutions seemed to be a bit more sensitive to the addition of Cu 2+ and Zn 2+ with respective decreases of 18 and 25% of CD signals compared to ion free control sample. Remarkably, among all the ions, the lead ions lead to the strongest CD signal quenching (58%), of intensity of control sample. A summary of the relationship between CD intensity versus ions concentrations for the seven kinds of metal ions are shown in Fig. 5. The expected linear relation significant of an effective sensing ability is present only in the case of lead ions detection. The slopes of fitted curves are summarized in Table S1. These evaluations indicate that the CdSe NPLs capped with chiral ligand are appropriate only for lead ions detection with both specificity and selectivity. As summarized in Table 1, our chiral detected performance was comparable (or even better) to the conventional optical heavy or not metal ions sensor based on photoluminescence methods. The CdSe NPLs could not be used for detecting lead ions through photoluminescence methods due to the non-luminous behavior after ligand exchange. However, the ion detection process could be realized through CD spectrum detection. Our work then provides a new and simple strategy for these non-luminous semiconductors in the field of heavy metal ion detection.

CONCLUSIONS

In summary, ultrathin CdSe NPLs with four monolayers were prepared in this work. The ligand exchange treatment was conducted to achieve the aqueous cysteine capped NPLs. The chiral ligand has induced the chiroptical activity unlocking the exitonic transition peaks of CdSe NPLs. Moreover, we have discussed the chiral signal originated from ligand transfer mechanism according to orbital coupling simulation based on TD-DFT calculation. Taking advantage of this mechanism, highly sensitive chiroptical sensing for lead ions detection with high selectivity was demonstrated. These results revealed that such chiral super-nanostructures may serve as a versatile platform for heavy metal ions detection in waste water process.

Conclusions

In conclusion, optical characteristics of CdSe/CdS/ZnS QDs were investigated through a comprehensive steady-state spectroscopic study. By analyzing the influence of ZnS shell on CdSe/CdS QDs, photostable and temperature-insensitive QDs have been obtained. Thanks to the improved optical properties of QDs, laser action is successfully obtained based on whispering gallery mode (WGM) at room temperature by using QDs as optical gain media. Furthermore, lasing at higher temperatures up to 312.6 K was demonstrated, which results from the superior photostability of the samples. Our result shows that CdSe/CdS/ZnS QDs have great potential as a laser material for better temperature stability and higher temperature operation. CdTe-TGA QDs Pb 2.0×10 -6 -1.0×10 -4 2.7×10 -7 [46] Graphene oxide Pb, K, Cu etc. 5.0×10 -9 -3.0×10 -7 3.0×10 -9 [51] CdZnSe-GSH QDs Pb, Zn, Co etc. 5.0×10 -9 -2.0×10 -5 2.7×10 -7 [47] CdSe-DAB QDs Cd, Pb, Mg etc. 1.0×10 -5 -1.5×10 -4 6.0×10 -5 [45] CdSe/CdS-XO QDs Pb, Cu etc. 5.0×10 -8 -6.0×10 -6 2.2×10 -8 [44] CdTe-TGA QDs Pb, Zn, Co etc. 2.0×10 -8 -3.6×10 -6 8.0×10 -8 [43] GSH-Mn-ZnS QDs Pb, Na, Cd etc. 4.8×10 -9 -4.8×10 -7 2.2×10 -9 [42] MPA-Mn-ZnS QDs Pb, Mg, Zn etc. 4.0×10 -8 -6.0×10 -6 3.7×10 -8 [41] CdSe-Cys NPLs Pb, Fe, Cu etc. 1.0×10 -8 -1.0×10 -6 4.9×10 -9 This work *TGA: Thioglycolic acid; GSH: Glutathione; DAB: Diaminobutane-based poly(propyleneimine) dendrimers; XO: Xylenol orange; MPA: Mercaptopropionic acid; Cys: Cysteine.

EXPERIMENTAL SECTION

Materials: Cadmium nitrate tetrahydrate (Cd(NO3)2 • 4H2O, 99.997%), cadmium acetate dihydrate (Cd(Ac)2•2H2O, 98.0%), oleic acid (OA, 90%), tetramethylammonium hydroxide pentahydrate (TMAH, ≥ 97%) were purchased from Sigma-Aldrich. L-cysteine hydrochloride monohydrate (99%), Dcysteine hydrochloride monohydrate (98%), and selenium powder (Se, ≥ 99.999%) were purchased from aladdin. 1octadecene (ODE, 90%) and sodium myristate (98 %) were purchased from J & K. Pure water was purchased from Wahaha, China. All chemicals were used as received without further purification.

Preparation of Cadmium Myristate: Cadmium myristate was prepared according to the recipe given in the literature. For a typical synthesis, 1.23 g of Cd(NO3)2•4H2O was dissolved in 40 mL of methanol and 3.13 g of sodium myristate was dissolved in 250 mL of methanol. After complete dissolution of Cd(NO3)2•4H2O and sodium myristate powders, solutions were mixed and stirred vigorously around 1 h. Then, the cadmium myristate (Cd(myristate)2) powders were precipitated by centrifugation. and re-dispersed in methanol. The washing step with methanol was performed at least three times to remove any unreacted and/or excess precursors. After successive washing steps, the precipitated part was completely dried under vacuum overnight. Synthesis of the CdSe Core NPLs: CdSe nanoplatelets (NPLs) having four monolayers (4 MLs) were prepared according to the procedure described in the literature with little modifications. 52 For a typical synthesis, 0.17 g (0.3 mmol) of Cd(myristate)2, 0.012 g (0.15 mmol) of Se and 15 mL of ODE were introduced into a three-neck flask. The solution was degassed at 100 °C for an hour under vacuum. After degassing step, the solution was heated to 240 °C under argon atmosphere. 0.055 g (0.2 mmol) of Cd(Ac)2•2H2O was swiftly added to the reaction solution. After 10 min growth at 240 °C, 1 mL of OA was injected, and the temperature of the solution was decreased to room temperature. The cadmium precursor injection temperature and the growth time were found to be important for the resulting shape and size of CdSe core NPLs. The resulting 4 ML CdSe NPLs are separated from other reaction products with successive purification steps. First, the resulting mixture is centrifuged at 14,500 rpm for 10 min, and the supernatant is removed from the centrifuge tube. The precipitate is dried under nitrogen, re-dispersed in hexane, and centrifuged again at 4,500 rpm for 5 min. In the second step, the supernatant is separated into another centrifuge tube, and 30% ethanol is added into the supernatant solution. In the last step, after the turbid solution is centrifuged at 4,500 rpm for 5 min, the precipitate is dissolved in hexane and filtered with a 0.20 μm filter. Ligand Exchange of the CdSe NPLs with Chiral Cysteine Molecules: The L-and D-cysteine ligand exchange was carried out using a previously reported method. 10,27 D-or L-cysteine hydrochloride monohydrate (0.2 M) was dissolved in DI water. The pH of solution was adjusted to 12 by TMAH to form the cysteine solution. 2 mL hexane solution containing CdSe NPLs was added into 2 ml of cysteine solution. The mixture was stirred at room temperature for 24 h. The reaction mixture was left to stand for 1 h to allow the phases to separate. The bottom aqueous layer was removed with a syringe, and the Cys-CdSe NPLs were purified by precipitation with ethanol/DI water (4:1, two times). The purified Cys-NPLs were re-dispersed in DI H2O and stored at room temperature in the dark (O.D.=0.26 for 2 mm optical length after ligand exchange).

Structural and Optical Characterization:

The UV/vis absorption measurements were performed using a TU-1901 double-beam UV/vis spectrophotometer (Beijing Purkine General Instrument Co. Ltd., China), and the emission spectra were recorded on a fluoroSENS spectrophotometer (Gilden Photonics).

Please do not adjust margins Please do not adjust margins CD measurements: The measurements were conducted on a JASCO J-1500 CD spectrometer at a scan rate of 20 nm/min. All CD experiments were carried out in Milli-Q water with a quartz cuvette (1 cm path length, Hellma). TEM images were collected using a Tecnai F30 microscope working at 300 kV. Powder X-ray diffraction (XRD) was carried out by a Rigaku Smartlab system. All the samples were at concentrations leading to peak absorbance of 1.15 O.D.. Calculation of g-Factor: In designing chiral nanoparticles for application, the anisotropy or g-factor is a key parameter to determine the optical activity due to its readily availability by comparing CD spectrum with absorption spectrum. The equation is as follows: g = ɵ /3.298 × 10 4 × A, where ɵ is the vertical coordinate of CD spectrum, and A is the value in the absorption spectrum at the same concentration used to measure the CD spectrum. Simulation method: Gaussian 09 software is used for all chemical calculations. Ground state geometries were optimized at the DFT. UV and CD spectra were calculated with TD-DFT. B3LYP and LanL2DZ basis sets were used for all the elements in the calculations. The small CdSe clusters usually are in the form of a tetrahedron; herein, we constructed a tetrahedron-like Cd4Se9 cluster to mimic the interactions between the chiral light and the nanoplatelets. We take the charge states of Cd and Se as 2 + and 2 -. All the geometries optimized with the lowest energy were convergence to make sure available model. [53][54][55][56][57][58][59]

48

 48

Fig. 1 .

 1 Fig. 1. (a) Schematic of ligand capping of CdSe NPLs with oleic acid. (b) TEM and (c) HRTEM image of OA capped CdSe NPLs. (d) XRD spe ctra of CdSe NPLs.

Fig. 2 .

 2 Fig. 2. (a-c) Ligand exchange process for phase transformation from oil to water with cysteine ligand replacement. (d) UV -Vis. absorption spectra of CdSe NPLs before and after ligand exchange. (e) Chirality performance using electronic CD measurements for OA, L-and D-cys as well as L+D (ee = 0) mixed capped NPLs.

2 S

 2 and 3d. The HOMO showing strong wave function overlaps between ZB CdSe nanocluster and thiol group in chiral ligand could be clearly observed. The results demonstrate the delocalized orbitals in CdSe and cysteine in the This journal is © The Royal Society of Chemistry 20xx

Fig 3 .

 3 Fig 3. (a) L-(b) D-cys capped CdSe nanocluster with optimized geometries. (c) Calculated frontier molecular orbitals of LUMO and (d) HOMO for L-cys capped CdSe nanocluster.

Fig. 4 .

 4 Fig. 4. (a) CD and (b) absorption measurements for lead ions detection ranging from 0.01 to 1 μM, both for L-and D-cys capped NPLs. (c) linear relationship between chiral signal at 520 nm and various lead ion concentrations. (d) Examination of the highly selectivity of CdSe NPLs chiral se nsor with seven kinds metal ion target analytes, the concentrations of all metal ions are settled in 0.1 μM. Statistics were performed on at least 5 times for the control at an OD of 1.2 and 3 times for each io n.

Fig. 5 .

 5 Fig. 5. Relationship between CD intensity and ion concentrations for the seven kinds of metal ions.

Table 1 .

 1 Lead ions limit of detection (LOD) and linear range.

	Sensing system	Detected metal ions	Range(M) (Pb)	LOD(M) (Pb)	Ref.
	Au nanoparticles	Pb, Fe, Mn etc.	1.0×10 -7 -1.0×10 -5	1.0×10 -7	[50]

This journal is © The Royal Society of Chemistry 20xx J. Name., 2013, 00, 1-3 | 7

Acknowledgements

This work is supported by the National Natural Science Foundation of China (11574130), Shenzhen Science and Technology Innovation Commission (Projects Nos. KQJSCX20170726145748464, JCYJ20150930160634263, JCYJ20170302142433007, and KQTD2015071710313656).

Conflicts of interest

The authors declare no competing financial interest.


Francis Albarède
email: albarede@ens-lyon.fr

Janne Blichert-Toft

François De Callataÿ

Gillan Davis

Pierluigi Debernardi

Liesel Gentelli

Haim Gitler

Fleur Kemmers

Sabine Klein

Chloé Malod- Dognin

Jean Milot

Philippe Télouk

Markos Vaxevanopoulos

Katrin Westner

From commodity to money: the rise of silver coinage around the ancient Mediterranean (6 th -1 st century BCE)

The reasons why the Western Mediterranean, especially Carthage and Rome, resisted monetization relative to the Eastern Mediterranean are still unclear. Here, we address this question by combining lead and silver isotope abundances in silver coinage from the Aegean, Magna Graecia, Carthage, and the Roman Republic. The clear relationships observed between 109 Ag/ 107 Ag and 208 Pb/ 206 Pb reflect mixing of silver ores or silver objects with lead metal used for

cupellation. The combined analysis of Ag and Pb isotopes reveal important information about the technology of smelting. The Greek world extracted Ag and Pb from associated ores, whereas, on the Iberian Peninsula, Carthaginians and Republican era Romans applied Phoenician cupellation techniques and added exotic Pb to lead-poor Ag ores. Massive silver re-cupellation is observed in Rome during the Second Punic War. After defeating the Carthaginians and the Macedonians in the late 2 nd century BCE, the Romans brought together the efficient, millenniumold techniques of silver extraction of the Phoenicians, who considered this metal a simple commodity, with the monetization of economy introduced by the Greeks.

Silver has been highly prized in the Mediterranean and Near East region for millennia. The most obvious traces of its extraction can be seen in the contamination of ice and peat bogs all the way to the Arctic [START_REF] Cortizas | Atmospheric Pb deposition in Spain during the last 4600 years recorded by two ombrotrophic peat bogs and implications for the use of peat as archive[END_REF][START_REF] Hong | Greenland ice evidence of hemispheric lead pollution two millennia ago by Greek and Roman civilizations[END_REF]. The rise of monetized silver in the late 6 th and the 5 th century BCE in the Eastern Mediterranean world can be interpreted in different ways. Some consider that minting facilitated trade [START_REF] Davis | Dating the drachmas in Solon's laws[END_REF][START_REF] Kim | A hoard of archaic coins of Colophon and unminted Silver (CH I. 3)[END_REF], and, indeed, the abundance of shipwrecks in the Mediterranean as a proxy for economic performance correlates with lead peaks observed in Arctic ice [START_REF] Parker | Ancient Shipwrecks of the Mediterranean and the Roman Provinces (BAR Int. S[END_REF]. In the long run, however, the spread of coinage is also undoubtedly linked to the explosion of Greek mercenary service in the late 6 th -5 th centuries, but to an extent that varies with the period and the local political circumstances [START_REF] De Callataÿ | Money and its ideas[END_REF][START_REF] Krasilnikoff | Aegean mercenaries in the fourth to second centuries BC: a study in payment, plunder and logistics of ancient Greek armies[END_REF][START_REF] Trundle | Greek mercenaries: From the late archaic period to Alexander[END_REF]. The present work uses Ag and Pb isotopes to further inform the debate by investigating the transition between two modes of silver utilization around the Mediterranean: an early mode in which silver was considered as a simple commodity and traded by weight, and a later monetary mode.

Silver isotopes hold the potential of adding a new dimension to the understanding of early economies first and foremost because silver is a metal of monetary interest and not only another isotopic tracer. As a fingerprinting tool for historical and archeological applications, Ag isotopes are free from the uncertainties often afflicting Pb isotopes when it comes to untangling silver extraction and recycling [START_REF] Brill | Lead isotopes in ancient coins: Methods of chemical and metallurgical investigation of ancient coinage: a symposium[END_REF]. Silver and Pb isotope variations reflect processes that are unrelated in nature. Silver isotope variations are due to the minute differences in the thermodynamic properties of the two stable isotopes 107 Ag and 109 Ag, known as the 'isotope effect' (Fujii and [START_REF] Evans | 109Ag-107Ag fractionation in fluids with applications to ore deposits, archeometry, and cosmochemistry[END_REF]. The range is very small (on the order of per mil) and essentially nil at the temperatures of magmatic and other high-temperature (such as cupellation) processes. The latter is true for Pb isotopes as well. In contrast, owing to the radiogenic nature of 206 Pb, 207 Pb, and 208 Pb, Pb isotope abundances vary with geological time and tectonic processes. Because Pb isotope ratios change by several percent upon decay of U and Th, they vary over a much larger range than the per mil variations created by the isotope effect, which affects Pb isotopes the same way as Ag isotopes, but is drowned in the radiogenic ingrowth not affecting the stable Ag isotopes.

The present work assesses the contribution of combining Ag and Pb isotopic data as a marker of silver extraction and metallurgy in the mining districts which contributed to the monetary mass of the ancient Mediterranean world through time. Much is already known about silver mining techniques and metallurgy [START_REF] Domergue | Les Mines Antiques. La Production des Métaux aux Epoques Grecques et Romaines[END_REF]. Cupellation, the technique of silver purification by smelting in the presence of metallic lead, was known from the early Bronze Age in the Middle East [START_REF] Pernicka | Silver production by cupellation in the fourth millennium BC at Tepe Sialk, in The Potters of Sialk[END_REF][START_REF] Pernicka | Late Uruk silver production by cupellation at Habuba Kabira, Syria[END_REF], Attica [START_REF] Kakavogianni | Silver metallurgical finds dating from the end of the Final Neolithic period until the Middle Bronze Age in the area of Mesogeia: Aegean metallurgy in the Bronze Age, proceedings of an international symposium held at the University of Crete[END_REF], and the island of Thasos [START_REF] Bassiakos | Late Neolithic/Early Bronze Age metallurgical practices at Limenaria, Thasos: Evidence for silver and copper production[END_REF]. On the Iberian Peninsula, however, evidence for cupellation, often in the form of litharge (lead oxide) is lacking before the Phoenician colonization in the early 1 st millennium BCE [START_REF] Bartelheim | The silver of the South Iberian El Argar Culture: A first look at production and distribution[END_REF][START_REF] Pérez Macías | Metalurgia extractiva prerromana en Huelva[END_REF][START_REF] Rovira Lloréns | Innovación y continuidad en la metalurgia prehistórica de la Península Ibérica: Presente y futuro de los paisajes mineros del pasado: Estudios sobre minería[END_REF], which opened up vigorous exchanges between Iberia and the Near East [START_REF] Neville | Mountains of silver and rivers of gold: the Phoenicians in Iberia[END_REF]. From the Second Punic War through to the 2 nd century AD, Romans took over the silver mining districts of southern Iberia first exploited by the Celtiberians and the Phoenicians from the 8 th century BCE [START_REF] Avery | Not on Queen Victoria's birthday: the story of the Rio Tinto mines[END_REF][START_REF] Jurado | The Tartessian economy: mining and metallurgy[END_REF][START_REF] Mata | The beginnings of the Phoenician presence in Southwestern Andalusia, The Phoenicians in Spain: An Archaeological Review of the Eighth-sixth Centuries BCE: a Collection of Articles Translated from Spanish[END_REF], and briefly later on during the mid-3 rd century by the Carthaginians. In the Greek states, the monetary economy was based largely on exploitation of ores in the Aegean and Thraco-Macedonia from the second half of the 6 th century through to the Roman conquest in 148 BCE.

A key question for determining the origin of bullion is the nature of ores used as the primary source (galena, sulfides, sulfosalts, chlorides, or their alteration products). If Ag and Pb are extracted from the same ore, such as galena, anglesite, or cerussite, their isotopes in the purified bullion will reflect the common metal source. If, however, cupellation involves exotic Pb, isotope systematics will instead reflect mixing between the two metal sources. Combined Ag and Pb isotopes can also address whether a particular metal used for coinage was extracted from a primary ore or rather was the product of the re-melting of mixed batches of silver fragments, such as silver plates, jewelry, or silverware.

Silver isotope variation in coinage (Fujii and [START_REF] Evans | 109Ag-107Ag fractionation in fluids with applications to ore deposits, archeometry, and cosmochemistry[END_REF] is much narrower than in ores [START_REF] Arribas | The isotopic composition of Ag in ore minerals[END_REF][START_REF] Mathur | Fractionation of silver isotopes in native silver explained by redox reactions[END_REF], which potentially allows for a high selectivity of provenance. Correlating minting ages and Pb isotopes have been able to uphold the proposition of a major shift of silver ores shortly after the Second Punic War [START_REF] Albarède | A miner's perspective on Pb isotope provenances in the Western and Central Mediterranean[END_REF][START_REF] Albarède | A glimpse into the Roman finances of the Second Punic War through silver isotopes[END_REF] . Here we add to this suggestion by describing correlations between Ag and Pb isotopes on 92 literature and newly acquired data on silver coinage from the Aegean world, Magna Graecia, Carthage, and Rome (Figure 1). With this new expanded study, we reverse Desaulty et al.'s (2011) conclusion that Ag and Pb isotopes are uncorrelated and discuss the bearings of the new data set and observed Ag-Pb correlations on our understanding of silver sources and extraction techniques in the pre-Roman Empire Mediterranean world.

The present work combines literature data with 14 new Pb isotope data and 42 new Ag isotope data (Supplementary Table 1). The Pb isotope compositions are reported in Figure 2 with symbols for proper identification and dates for minting age (the median of the uncertainty range). The fields of volcanic rocks ( [START_REF] Albarède | A miner's perspective on Pb isotope provenances in the Western and Central Mediterranean[END_REF]; http://georoc.mpch-mainz.gwdg.de/georoc/) genetically related to magmatic hydrothermal ore deposits (light blue for Southern Iberia, pink for the Aegean) from the literature are also plotted to show how closely the main group comprising most samples from Thrace, Greece, and the Greek colonies (Magna Graecia) track potential lead sources. Also plotted is the field of Pb isotopes in the Near Eastern silver hoards of hacksilber (chopped silver plates and jewelry) from Dor, Akko, and Ein Hofez (10 th -9 th century BCE), which are contemporaneous with the first Phoenician settlements around the Mediterranean, especially in Southern Iberia [START_REF] Eshel | Lead isotopes in silver reveal earliest Phoenician quest for metals in the west Mediterranean[END_REF][START_REF] Neville | Mountains of silver and rivers of gold: the Phoenicians in Iberia[END_REF]. Particularly striking is a major difference in the apparent source of the metal used to mint the denarii from the Roman Republic [START_REF] Albarède | A miner's perspective on Pb isotope provenances in the Western and Central Mediterranean[END_REF][START_REF] Westner | Rome´s rise to power. Geochemical analysis of silver coinage from the Western Mediterranean (4th to 2nd centuries BCE)[END_REF]. The majority of late denarii (in black) plot to the right of the coins struck before the Second Punic War (240-200 BCE) to slightly after and conspicuously overlap the fields of Phoenician hoards [START_REF] Eshel | Lead isotopes in silver reveal earliest Phoenician quest for metals in the west Mediterranean[END_REF] and the field of the Phoenician and Ibero-Punic slags and silver objects found in Southern Iberia [START_REF] Anguilano | The importance of lead in the silver production at Riotinto (Spain). L'importance du plomb dans la production d'argent à Riotinto (Espagne)[END_REF][START_REF] Murillo-Barroso | The Macro-Regional Scale of Silver Production in I beria During the First Millennium BC in the Context of Mediterranean Contacts[END_REF]. Although more data clearly are needed, Pb isotopes from three Carthaginian and two Gaulish coins also plot to the right but at higher 208 Pb/ 206 Pb values.

Silver isotope compositions are reported as e109Ag values (the deviation in parts per 10,000 of the 109 Ag/ 107 Ag ratio of any given sample from that of the Ag standard SRM NIST 978a) [START_REF] Desaulty | Isotopic Ag-Cu-Pb record of silver circulation through 16th-18th century Spain[END_REF]. All the data fall into four well-defined groups. We found that the clearest representation is e109Ag vs 208 Pb/ 206 Pb (Figure 3). Model ages could have been used as well but, since 208 Pb/ 206 Pb ratios strongly correlate with model ages (Figure 4), we opted for the former which is more familiar to the community of Archaeometry. In the e109Ag vs 208 Pb/ 206 Pb plot, the data form a bundle of arrays radiating from e109Ag~0 and 208 Pb/ 206 Pb =~2.065:

(1) Group 1 slightly trends to the right of e109Ag ~ -0.25

(2) The well-populated Group 2 trends more steeply towards a value of e109Ag ~ -1.0;

(3) Group 3 trends towards positive values.

These groups are well delineated at high 208 Pb/ 206 Pb. A few samples cannot be attributed to one group or another, while some rogue points are arguably difficult to explain, such as the low e109Ag-Abydos drachma. With these restrictions in mind, most data points of Group 1 are Roman Republic denarii minted before the Second Punic War to slightly after. Most data points of Group 2 are coinage from Carthage and Roman Republic denarii minted after the Second Punic War. The two Gaulish samples belong to this group. Five coins from Group 3 were minted in Southern Italy and Sicily (Thurium, Gela, Syracuse, and Selinus). Most coins from classic Greece and Magna Graecia plot at the apex of the bundle ( 208 Pb/ 206 Pb~2.065) and can be considered as either pertaining to all groups or not pertaining to any group, in which latter case they can be considered to define a fourth group.

The coins from Groups 1-3 plot within a narrow range of ±1 e109Ag. This range is conspicuously smaller (Fuji and Albarède, 2018) than the spread of Ag ore deposits [START_REF] Arribas | The isotopic composition of Ag in ore minerals[END_REF][START_REF] Mathur | Fractionation of silver isotopes in native silver explained by redox reactions[END_REF]. Group 2 shows a well-defined negative correlation between e109Ag and 208 Pb/ 206 Pb. This group, with few exceptions, corresponds to the field to the right of the magmatic trend in Figure 2. Given that silver and lead isotope fractionation is due to processes physically independent of each other, this array attests to mixing between two end-members, e.g., an Ag ore and Pb metal used for cupellation. The idea that lead in silver coins comes from a mixture of Pb and Ag ores is not new [START_REF] Anguilano | The importance of lead in the silver production at Riotinto (Spain). L'importance du plomb dans la production d'argent à Riotinto (Espagne)[END_REF][START_REF] Brill | Lead isotopes in ancient coins: Methods of chemical and metallurgical investigation of ancient coinage: a symposium[END_REF][START_REF] Eshel | Lead isotopes in silver reveal earliest Phoenician quest for metals in the west Mediterranean[END_REF], but Pb isotopes by themselves do not suffice to substantiate mixing, first because ores often form from mixed sources, and second because lead is a metallurgical additive to the silver ore indispensable for Ag purification.

At this stage, it is essential to emphasize the non-linear property of mixing in ratio-ratio plots when the denominators, here 107 Ag and 206 Pb, are different (Appendix A). This can be simply cast by reasoning that adding pure lead to a mixed Ag-Pb ore leaves silver with the e109Ag signature of the ore. In contrast, mixing changes 208 Pb/ 206 Pb and the other isotopic ratios, but only if Pb from the ore and the cupellation metal have different origins. The curvature of mixing hyperbolae increases with the contrast of 107 Ag/ 206 Pb, or accepting, for sake of simplification, a negligible inaccuracy, the Ag/Pb ratios between the two end-members. Reference hyperbolae going through two geologically and historically acceptable end-members have been calculated and drawn in Figure 4 using the simple and useful relationship derived in Appendix A. In the e109Ag vs 208 Pb/ 206 Pb plot, the slope of each array at the Ag-rich end-member (Figure 5) is a particularly useful parameter. When comparing two Groups, say 1 and 2, the Pb/Ag ratio of the silver-rich group relates to the corresponding slope through:

(Pb/Ag) 2 (Pb/Ag) 1 = slope 1 slope 2 (1)
Note that the hyperbolae drawn in Figure 4 were not fitted through the actual data but were drawn for reference only. Silver ore smelted for the peri-Punic Group 1 is clearly much richer in Ag than that used for Group 2. Mixing within Group 2 (in blue) corresponds to a Pb/Ag ratio in the Ag ore only a factor of ~4 times lower than in the Pb metal used for cupellation. The ore was therefore fairly poor in silver. In contrast, the Ag ore of Group 1 (in green) has a Pb/Ag ratio 20 times smaller than in the Pb additive. Combining Ag and Pb isotope analysis of Roman coins confirms evidence from archeological excavations [START_REF] Anguilano | The importance of lead in the silver production at Riotinto (Spain). L'importance du plomb dans la production d'argent à Riotinto (Espagne)[END_REF][START_REF] Eshel | Lead isotopes in silver reveal earliest Phoenician quest for metals in the west Mediterranean[END_REF][START_REF] Murillo-Barroso | The Macro-Regional Scale of Silver Production in I beria During the First Millennium BC in the Context of Mediterranean Contacts[END_REF] that the two metals came from distinct sources and had to be combined for efficient silver extraction. The range in Pb isotope compositions of Roman ingots from Iberia is rather narrow [START_REF] Trincherini | The identification of lead ingots from the Roman mines of Cartagena: the role of lead isotope analysis[END_REF], while the e109Ag values of Iberian galenas, although more variable, are negative [START_REF] Arribas | The isotopic composition of Ag in ore minerals[END_REF].

Most of the data in Figure 4 cluster around e109Ag ~ -0.25 and 208 Pb/ 206 Pb=2.06-2.08. The 207 Pb/ 206 Pb range is ambiguous. But, as pointed out by Fujii and Albarède (2018), the narrow range of e109Ag (-1 to +2 per 10,000) in silver coinage, whether minted around the Mediterranean during the Antiquity, in medieval or pre-modern Europe, or in the Spanish Americas, contrasts markedly with the broad scatter of several per mil observed for potential silver ores [START_REF] Arribas | The isotopic composition of Ag in ore minerals[END_REF]. It is remarkable that the mean value of e109Ag in silver coinage differs from the (rare) mantle values but is indistinguishable from the value of the organic-rich Cody Shale value [START_REF] Theis | Palladium-silver chronology of IAB iron meteorites[END_REF] deposited some 85 Ma ago. This observation is consistent with the suggestion that the metals of Pb-Zn(-Ag) ores were eventually derived not from the mantle, but from oceanic black shales deposited in anoxic environments (Milot et al., submitted).

The range of e109Ag values among the silver-rich end-members is very narrow, within ±0.2 epsilon units of the SRM NIST 978a value. Such striking lack of fractionation of Ag isotopes suggests that the source ores formed primarily in a high-temperature environment, usually qualified as mesothermal (200-400°C) [START_REF] Guilbert | The geology of ore deposits[END_REF]. It shows that the silver ores, presumably sulfides, did not go through intense weathering or any other low-temperature process that could have readily fractionated Ag isotopes [START_REF] Evans | 109Ag-107Ag fractionation in fluids with applications to ore deposits, archeometry, and cosmochemistry[END_REF]. The ores used as lead sources could be galena and cerussite, as is the case for the Athenian Lavrion district [START_REF] Conophagos | Le Laurium antique et la technique grecque de la production de l'argent[END_REF] and Southern Iberia [START_REF] Baron | The Cabezo del Pino workshops complex (Sierra Minera de Cartagena-La Union, Murcia) and the mining activities organisation at Carthago Noua at the end of the Roman Republic. Cross-contributions of archaeology and geochemistry[END_REF]. The contrast between the Aegean and non-Aegean coins implies that the source of silver of the former went through cupellation using geologically young (Cenozoic), and probably cogenetic, lead and silver. Extraction of the two metals in the Aegean must have taken place at a short distance from each other.

The new perspective provided by Ag isotopes and derived from the curvature of Ag-Pb mixing hyperbolae allows pursuing the suggestion based on archeometallurgical observations [START_REF] Rowan | Coinage as commodity and bullion in the western Mediterranean, ca. 550-100 BCE[END_REF], statistics on triumph and temple treasure composition [START_REF] De Callataÿ | Reflexions quantitatives sur l'or et !'argent non monnayés a l'epoque hellénistique[END_REF], and Pb isotopes on silver coinage (Birch et al., 2020b) that the Western Mediterranean used Greek coinage as a source of bullion. Silver-lead isotopic correlations show that silver from Group 1, most likely delivered by Carthage after the First Punic War, was reprocessed by the Romans. In contrast, the two Gaulish coins belong to Group 2 and, therefore, were not re-cupellated from pre-existing bullion.

Silver ores smelted for Group 3 (Gela, Thurium, Selinus, Syracuse, and cities from Magna Graecia with a long history of war and shared interests) were cupellated with unusual Pb ores containing Ag with positive e109Ag values, but unfortunately of so-far unknown origin.

The corollary of these observations is: a) Silver from most Aegean and Magna Graecia samples was smelted with lead from local ores. Having both Ag and Pb sources at disposal locally, possibly from the same mining district, perhaps the Lavrion, seems to have been key to secure silver production. b) In contrast, silver mined by Carthage in the 4 th century BCE and by the Romans in the second half of the 2 nd century BCE was cupellated with geologically older lead extracted at some distance from the silver ores. c) Metal used for Roman coins at the time of the Second Punic War was obtained from recalled silver [START_REF] Albarède | A glimpse into the Roman finances of the Second Punic War through silver isotopes[END_REF] and purified with Pb unrelated to any identifiable silver mining districts.

Silver used for minting Roman coinage during the Second Punic War (218-201 BCE, Group 1) was minted only during a short time period consistent with a discontinuity in the Ag isotopic composition noted by [START_REF] Albarède | A glimpse into the Roman finances of the Second Punic War through silver isotopes[END_REF] at the time of the Roman monetary reform of 213 BCE (this date is used here for convenience; the exact date is still a matter of debate though not too far off 213 BCE). This change in silver source most likely corresponds to domestic remelting of massive amounts of silver metal from the war penalties paid by Carthage after the First Punic War and the Mercenary wars. Given the apparent success of the silver recall in Rome at the climax of the 2 nd Punic War (Livy, XXIII,48.9), Carthaginian silver must have been partly in private hands, mostly as silverware and jewelry, and partly in the form of debased quadrigati. Cupellation using scrap Pb of nondescript origin, possibly from Sardinia, which Rome annexed in 238 BCE, must have followed in the wake of the unsuccessful debasement attempted during the monetary reform of 213 BCE.

The narrow range of Pb/Ag ratios displayed by each group requires explanation. The answer lies in the metallurgical process. Silver lost to the litharge by oxidation steeply increases with metal fineness [START_REF] Swinbourne | Thermodynamic modelling of cupellation: activity of AgO0• 5 in molten PbO at 1000ºC[END_REF][START_REF] Ueda | Thermodynamic study on the Ag-Pb-O system at 1273 K[END_REF]. In other words, making very pure silver comes at the price of wasting more of it. The metallurgist therefore has to play with two parameters, the oxygen flow and the Pb/Ag ratio of the silver ore-lead metal mixture to find the optimum ore-lead mixture that will provide the purest silver without losing too much of it to the slag (see Appendix B). Two-step cupellation as proposed for Lavrion [START_REF] Pernicka | Archäometallurgische Untersuchungen zur antiken Silbergewinnung in Laurion. III. Das Verhalten einiger Spurenelemente beim Abtreiben des Bleis[END_REF] would manifest itself by sharply curved mixing hyperbolae unless Ag and Pb ores are very similar in both components. Archeological finds from many Phoenician settlements attest that the art of cupellation applied to low-grade ores, typically jarosite earth from Rio Tinto in the lower Guadalquivir valley near the modern town of Huelva, using metallic Pb had been mastered to the highest degree well before the first Lydian (late 7 th century BCE) and Aegean (late 6 th century BCE) coinages were struck [START_REF] Blanco | Pre-Roman Silver Miners at Riotinto[END_REF]. The narrow range of Pb concentrations in coins from the Roman Republic [START_REF] Hollstein | Metallanalytische Untersuchungen an Münzen der Römischen Republik[END_REF][START_REF] Westner | Rome´s rise to power. Geochemical analysis of silver coinage from the Western Mediterranean (4th to 2nd centuries BCE)[END_REF], which is always less than solubility in silver (1-3 wt.% between 300 and 900°C, [START_REF] Karakaya | The Ag-Pb (Silver-Lead) system[END_REF]), and the low vapor pressure of Pb, leaves no doubt that Pb was not added to the metal after purification.

Once the Carthaginians were defeated in Zama in 202 BCE, it took the Romans several decades to control the land of the Celtiberian tribes (Appian, 6; [START_REF] Richardson | BC[END_REF]) and organize mining and silver extraction. The present data provide evidence that the Romans only recommenced large-scale exploitation of jarositic Ag ores of the Iberian Pyrite Belt and Rio Tinto [START_REF] Domergue | Les mines de la Péninsule Ibérique dans l'antiquité romaine[END_REF][START_REF] Westner | Rome´s rise to power. Geochemical analysis of silver coinage from the Western Mediterranean (4th to 2nd centuries BCE)[END_REF] in the late 2 nd century BCE. They clearly did not adopt the Greek method of local mining of high-grade ores, but used the Phoenician protocol, also mastered by the Celtiberian tribes, of combining low-grade Ag ores with galena-sourced Pb from the entire Iberian Peninsula.

The Eastern and Western Mediterranean civilizations looked at the same metal but from different perspectives. Phoenicia [START_REF] Elayi | The coinage of the Phoenician city of Tyre in the Persian period (5th-4th cent. BCE)[END_REF][START_REF] Gitler | The coinage of Philistia of the fifth and fourth centuries BC: a study of the earliest coins of Palestine[END_REF] and Carthage [START_REF] Jenkins | Coins of Punic Sicily. Part 2, Carthage Series I[END_REF], although well aware of coined silver in the Aegean world, primarily considered this metal as a precious commodity, just as ivory, myrrh, and amber. Only with the arrival of Greek mercenaries in the second half of the 5 th century BCE did these authorities adopt coinage. Not all the Greeks, however, struck silver as intensively as did Athens and a few other city-states later on and, later on again, Hellenistic kingdoms. The Western Mediterranean, especially Carthage and Rome, also resisted monetization for longer than did the Greek world. From the inventories of Roman triumphs and temples in the 2 nd century BCE, de Callataÿ ( 2006) concluded that at least 80 wt.% of available silver was left as bullion (infecti). Monetized silver does not seem to have become prevalent relative to jewelry and bullion, as attested to by persistent references to large sums in weight units (37). The overall coincidence between the isotopic differences and the historical gap between Groups 1 and 2 testifies to the reluctance of the Roman Republic to use coined money for larger transactions. Indemnities inflicted on the defeated Carthaginians reported by Polybius in his Histories and on Macedonians reported by Livy (XXXIII,30.7), always labelled in talents and not in any sort of monetary denomination, attest to the Romans being uneasy about pervasive monetization until reliable sources of bullion were safely established. Numismatic evidence supporting the rapid expansion of silver coinage in the second half of the 2 nd century BCE and its potential causes, such as pressure on the copper supply for as minting [START_REF] Bransbourg | La politique monétaire de la République romaine[END_REF], are reviewed in detail by [START_REF] Kay | Rome's Economic Revolution[END_REF].

Materials and Methods

Silver isotopes were analyzed on series of coins from the Roman Republic [START_REF] Westner | Rome´s rise to power. Geochemical analysis of silver coinage from the Western Mediterranean (4th to 2nd centuries BCE)[END_REF] and Magna Graecia (Birch et al., 2020a;Birch et al., 2020b). Drill cores previously analyzed for Pb isotopes were re-used for Ag isotopes. Drill cores of Punic War denarii from Spain (provided by Pierluigi Debernardi) and of ten drachmas from different periods of Thrace, Athens, Miletus, Rhodes, Cilicia, and Gela (Birch et al., 2020a) were analyzed for both Pb and Ag isotopes.

Literature data [START_REF] Albarède | A glimpse into the Roman finances of the Second Punic War through silver isotopes[END_REF][START_REF] Desaulty | Isotopic Ag-Cu-Pb record of silver circulation through 16th-18th century Spain[END_REF] complemented the data set. Analytical techniques have been described elsewhere [START_REF] Desaulty | Isotopic Ag-Cu-Pb record of silver circulation through 16th-18th century Spain[END_REF].

slag is approximately

𝑓 XYZ[ 𝐾 XYZ[/]^_ `[ ≈ 𝐶 M TU 𝐾 XYZ[/]^_ b[ (B2)
where

𝐾 XYZ[/]^_ `[
is the coefficient of Ag fractionation between the slag and purified Ag.

Reducing the amount of lead in the original ore-lead mixture both reduces 𝐶 M TU and increases 10 [START_REF] Ueda | Thermodynamic study on the Ag-Pb-O system at 1273 K[END_REF], which requires the proportions to be optimized before extraction. This result justifies that the Pb/Ag ratio during cupellation is constrained within a relatively narrow range. Graecia coinage (primary ores). The points to the right attest to addition of exotic Pb to Ag ores during cupellation. The overlap between these points, the Phoenician hoards from the 10 th -9 th 10 century BCE, and the Ibero-Punic slags and silver objects found in southern Iberia [START_REF] Anguilano | The importance of lead in the silver production at Riotinto (Spain). L'importance du plomb dans la production d'argent à Riotinto (Espagne)[END_REF][START_REF] Murillo-Barroso | The Macro-Regional Scale of Silver Production in I beria During the First Millennium BC in the Context of Mediterranean Contacts[END_REF], show that first Carthage, then Rome in the late 2 nd century BCE, had adopted the Phoenician smelting process. the ore (or recycled bullion) and (Pb/Ag)Pb in the lead used for smelting and q is the ratio (Pb/Ag)ore /(Pb/Ag)Pb. For Group 1 (green mixing hyperbola), (Pb/Ag)ore = 20 (Pb/Ag)Pb, which points to a Ag-rich silver source. In contrast, for Group 2 (blue mixing hyperbola), (Pb/Ag)ore = 4 (Pb/Ag)Pb, which suggests a Ag-poor source. Group 1, consisting of coins minted around the Second Punic War (see panel b), clearly represents re-melting of silver bullion, jewelry, or debased coinage. Group 2, comprising coins minted by Carthage, some Magna Graecia colonies, and the late Roman Republic, represents coins minted from Iberian ores after cupellation with exotic Pb. In contrast, coins pertaining to the largest group of Greek and Magna Graecia ores and plotting near the apex of this bundle was cupellated using Pb cogenetic with Ag ores. Five Magna Graecia coins (Group 3) definitely have been cupellated with Pb containing Ag with positive e109Ag values. The dashed grey hyperbolae show more potential mixing arrays: shallow slopes, both positive and negative, indicate probable multiple events of cupellation.

𝐾 XYZ[/]^_ b[ (

Fig. 1 .

 1 Fig. 1. Map of the Mediterranean with names of localities (cities and major ore districts) referred to in the text. In the interest of clarity, the toponymy used is a mix of names from different periods (e.g., Greece, Asia Minor).

Fig. 2 .

 2 Fig. 2. Lead isotope relationships for the samples analyzed in this study and literature data. Panels (a) and (b) show the same samples with symbols (panel a) and minting ages (BCE) (panel b), respectively. In spite of a substantial overlap, panel b was meant to reduce the number of

Fig. 3 .

 3 Fig. 3. Correlation between 109 Ag/ 107 Ag, expressed using the epsilon notation (deviation from a reference ratio in parts per 10,000), and 208 Pb/ 206 Pb for 92 silver coins from the Aegean world, Magna Graecia, Carthage, and Rome. In spite of a substantial overlap, panel b was meant to reduce the number of different symbols and improve legibility, while helping with the age of minting. Groups are referred to with light grey roman numerals. The same data are represented with symbols (panel a) and minting ages BCE (panel b). (a) Mixing lines do not display as linear arrays but as hyperbolae (see Supplementary Information). The curves are shown as references and were not obtained by a fit to the data. The curvature is a function of the ratio (Pb/Ag)ore in

Fig. 4 .

 4 Fig. 4. Correlation between model ages (in million years) as calculated inAlbarède et al. (2012)

Fig. 5 .

 5 Fig. 5.Hyberbolae representing mixing between a silver-rich ore a and metallic lead b used for smelting for different values of q = (Pb/Ag) b /(Pb/Ag) a . The thin colored lines represent tangents to the corresponding mixing hyperbolae at x = x a and y = y a (see text). 𝑠 = 7𝑦 ) -𝑦 ' 8/7𝑥 ) -𝑥 ' 8.

  Table 1). Right: minting ages BCE.

									RdN
								Mi
	Model Age (Ma)		RdSy Rd Rd Se Rd Sy rQ rQ Sy Hi Rd Rd Rd Rv Rq RdC Rd RDSp Rd R GS Rd Ci Ca AP Rdca	RQ Rd rd Ca Rd Rd GA	Rd Ca
		Sy Sy Sy Sy Th Ta Ta Ta Ta Me Me Me R R Th r Ma Hi Sy Me R R Rq Rv RvLu Ma Is AP At Ge Io Rh At At ND AP At Ph Rv Rdsy RqLu Rd Rv Ab Ma Ly Rca Ab Is Is Ma	Rd Ab	Ta
			Si
	2.05	2.06	2.07		2.08	2.09	2.1	2.11	2.05	2.06	2.07	2.08	2.09	2.1	2.11
			208 Pb/	206 Pb

Acknowledgments:

The authors thank the curators of the coin collections of Düsseldorf, Giessen, Heidelberg, Münster, and Tübingen for giving permission to sample their coin collections. We further thank Yigal Erel and Ryan Mathur for useful and courteous reviews. Funding: This is a contribution of Advanced Grant 741454-SILVER-ERC-2016-ADG 'Silver Isotopes and the Rise of Money' awarded to FA by the European Research Council; Author contributions: FA, JBT, FdC, GD, HG, PD, SK conceived the scientific project. JBT, CMD, SK, FK, and KW produced the data. FA, JBT, FdC, GD, PD, LG, HG, FK, SK, JM, MV, and KW contributed to the interpretations and writing; Competing interests: Authors declare no competing interests; Data and materials availability: All data are available in the supplementary table.

Appendix A: The mixing equation for ratios

Mass balance of isotope ratios x and y upon mixing of end-members a and b requests [START_REF] Albarède | Introduction to Geochemical Modeling[END_REF]:

where

is the mass fraction of Pb contributed by end-member b. f a and f b stand for the weight fraction of end-members a and b, respectively, and C for the weight concentration of the species at the denominator ( 206 Pb for x and 107 Ag for y). Similar definitions apply to the other variables.

Replacing 𝜑 𝑃𝑏 𝛼 by 1 -𝜑 𝑃𝑏 𝛽 , we obtain

The last equation can be rearranged, with a similar equation for y as At the expense of a negligible error, 206 Pb/ 107 Ag for y ratios have, for sake of simplicity, been replaced by the elemental weight ratios Pb/Ag. Expressing the isotopic ratios, we finally get:

The last equation is represented by a hyperbola when the Pb/Ag ratios of the two end-members are different and by a straight-line when they are identical (Figure 5). An alternative equation can be derived:

where

The value of q is inferred from the slope of the mixing hyperbola (dy/dx)a at x = x a . After tedious, but straightforward manipulations, the relationship

is obtained, where 𝑠 = 7𝑦 ) -𝑦 ' 8/7𝑥 ) -𝑥 ' 8.

A useful expression relating q, jPb, and jAg can also be derived: Table ST1