R Echevarria

G Gley

S Landrot

F Pollastri

Emmanuelle Fraysse

Montargès-Pelletier

I O Zelano

C Cloquet

A Van Der Ent

G Echevarria

Coupling nickel chemical speciation and isotope ratios to decipher nickel dynamics in the Rinorea cf. bengalensis-soil system in Malaysian Borneo

Keywords: Ni, ultramafic soil, hyperaccumulator plants, isotopic fractionation, chemical speciation

Coupling nickel chemical speciation and isotope ratios to

decipher nickel dynamics in the Rinorea cf. bengalensis-soil system in Malaysian Borneo

Introduction

Ultramafic rocks are mantle rocks characterized by a specific composition with low silica, high contents of mafic minerals (Fe and Mg oxides). Soils derived from the alteration of ultramafic rocks pose edaphic stresses for vegetation, including a low Ca/Mg ratio, lack of essential nutrients such N, P, and K, and a high concentrations of Mn, Ni, Cr and Co [START_REF] Whittaker | The Ecology of Serpentine Soils[END_REF][START_REF] Bonifacio | Pedogenesis in a soil catena on serpentinite in north-western Italy[END_REF][START_REF] Becquer | Sources of trace metals in Ferralsols in New Caledonia[END_REF]). These soils are known for their hosting adapted vegetation, which includes the occurrence of hyperaccumulator plants that can achieve extraordinary levels of metal accumulation due to enhanced uptake and translocation mechanisms from the roots to the shoots [START_REF] Baker | Accumulators and excluders strategies in the response of plants to heavy metals[END_REF]. On ultramafic soils developed from serpentinized ultramafic rocks (which represent the most important terrestrial Ni reservoirs), numerous Ni hyperaccumulator plant species have been reported over the past 40 years (Reeves et al. 2018a). Hyperaccumulators can attain exceptionally high concentrations of Ni in their living tissues, with up to several weight percent Ni (e.g. 25 Wt% in latex or 16 Wt% in phloem sap) [START_REF] Jaffre | Sebertia acuminata: A Hyperaccumulator of Nickel from New Caledonia[END_REF][START_REF] Baker | Botanical Exploration for Minerals in the Humid Tropics[END_REF][START_REF] Reeves | Tropical hyperaccumulators of metals and their potential for phytoextraction[END_REF][START_REF] Van Der Ent | Hyperaccumulators of metal and metalloid trace elements: Facts and fiction[END_REF][START_REF] Mesjasz-Przybylowicz | Extreme nickel hyperaccumulation in the vascular tracts of the tree Phyllanthus balgooyi from Borneo[END_REF]Reeves et al. 2018b;[START_REF] Jaffré | The discovery of nickel hyperaccumulation in the New Caledonian tree Pycnandra acuminata 40 years on: an introduction to a Virtual Issue[END_REF]. Nickel hyperaccumulator plants are known to transfer most of the Ni into their aerial parts (shoots) of the plant, instead of using root sequestration for metal detoxification [START_REF] Baker | Accumulators and excluders strategies in the response of plants to heavy metals[END_REF]. Via leaf litter deposition, hyperaccumulators can increase Ni concentrations of the soil in a surface area surrounding the plant [START_REF] Boyd | The significance of metal hyperaccumulation for biotic interactions[END_REF]. In tropical Southeast Asia, Rinorea cf. bengalensis (Violaceae) is one of more than 40 different Ni hyperaccumulator species identified from the Malaysian state of Sabah on Borneo Island (van der Ent et al. 2018b;2019a). Rinorea cf. bengalensis is a large tree (up to 25m tall) that contain up to 4.77 kg of Ni (van der Ent and Mulligan 2015). Substantial Ni enrichment under the canopy of R.

cf. bengalensis has been reported due to the degradation of leaf litter (van der Ent et al. 2015). However, it is still an open question to what extent Ni is recycled in the natural habitat of R. cf. bengalensis through Ni uptake, leaf shedding, re-uptake of Ni from decomposed leaf litter, and the Ni release in surface soil (Nkrumah et al. 2019b, d).

The interest toward Ni cycling at the Earth's surface has rapidly increased during the last decades but only a few scientific studies have been published proposing the use of stable Ni isotopes to elucidate the biogeochemical cycle of Ni in the upper soil horizons and surface waters [START_REF] Elliott | The Isotope Geochemistry of Ni[END_REF]. The degree of Ni isotope fractionation between the different surface compartments are a practical approach to identify and characterise the Ni biogeochemical processes. In order to better understand Ni isotopic fractionation on a larger scale, in catchments or soil-plant systems, the processes that fractionate Ni isotopes at the molecular scale need to be elucidated. Recent studies contributed to this understanding by showing Ni isotopic fractionation due to sorption onto mineral phases under laboratory controlled conditions [START_REF] Wasylenki | Ni isotope fractionation during sorption to ferrihydrite: Implications for Ni in banded iron formations[END_REF][START_REF] Wang | Experimental constraints on reconstruction of Archean seawater Ni isotopic composition from banded iron formations[END_REF][START_REF] Gueguen | Variable Ni isotope fractionation between Feoxyhydroxides and implications for the use of Ni isotopes as geochemical tracers[END_REF][START_REF] Spivak-Birndorf | Nickel isotope fractionation during continental weathering[END_REF], and due to complexation with organic acids relevant for the soil-plant system [START_REF] Zelano | The influence of organic complexation on Ni isotopic fractionation and Ni recycling in the upper soil layers[END_REF]. Nickel is involved in various biological reactions that might be implicated in isotopic fractionation [START_REF] Cameron | A biomarker based on the stable isotopes of nickel[END_REF]) but, to date, only few works have been focused on the impact of hyperaccumulator plant homeostasis on the Ni isotopic signature in surface soils [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF][START_REF] Estrade | Weathering and vegetation controls on nickel isotope fractionation in surface ultramafic environments (Albania)[END_REF][START_REF] Zelano | The influence of organic complexation on Ni isotopic fractionation and Ni recycling in the upper soil layers[END_REF][START_REF] Ratié | The behavior of nickel isotopes at the biogeochemical interface between ultramafic soils and Ni accumulator species[END_REF]). However, isotopic fractionation of metals has been observed in higher plants and has been interpreted as a consequence of physiological processes involved in maintaining homeostasis, especially with regard to Fe [START_REF] Guelke | Fractionation of Stable Iron Isotopes in Higher Plants[END_REF], Zn [START_REF] Weiss | Isotopic discrimination of zinc in higher plants[END_REF][START_REF] Tang | Fractionation of Stable Zinc Isotopes in the Field-Grown Zinc Hyperaccumulator Noccaea caerulescens and the Zinc-Tolerant Plant Silene vulgaris[END_REF][START_REF] Tang | Zinc Isotope Fractionation in the Hyperaccumulator Noccaea caerulescens and the Nonaccumulating Plant Thlaspi arvense at Low and High Zn Supply[END_REF][START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF][START_REF] Caldelas | Zinc Homeostasis and isotopic fractionation in plants: a review[END_REF], Cd [START_REF] Imseng | Fate of Cd in Agricultural Soils: A Stable Isotope Approach to Anthropogenic Impact, Soil Formation, and Soil-Plant Cycling[END_REF][START_REF] Imseng | Towards an understanding of the Cd isotope fractionation during transfer from the soil to the cereal grain[END_REF] and Cu [START_REF] Jouvin | Stable Isotopes of Cu and Zn in Higher Plants: Evidence for Cu Reduction at the Root Surface and Two Conceptual Models for Isotopic Fractionation Processes[END_REF]. For example, in the case of Zn, the preferential uptake at root level of heavier or lighter Zn isotopes has been associated with the presence of high-and low-affinity transport systems, respectively [START_REF] Weiss | Isotopic discrimination of zinc in higher plants[END_REF][START_REF] Caldelas | Zinc Homeostasis and isotopic fractionation in plants: a review[END_REF]. The type of transport system is dependent on the bioavailability of the metal, e.g. a high level of available Zn favours lighter isotopes uptake through low affinity transports, while a lower availability of the metal induces uptake of heavy isotopes through high affinity transporters [START_REF] Tang | Zinc Isotope Fractionation in the Hyperaccumulator Noccaea caerulescens and the Nonaccumulating Plant Thlaspi arvense at Low and High Zn Supply[END_REF].

For Zn, it has been suggested that further fractionation from roots to higher parts of the plant can take place during the xylem loading process and subsequent transportation [START_REF] Tang | Zinc Isotope Fractionation in the Hyperaccumulator Noccaea caerulescens and the Nonaccumulating Plant Thlaspi arvense at Low and High Zn Supply[END_REF]).

In the case of Ni, the physiological mechanisms regulating metal homeostasis are still largely unknown [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF], but few studies posit that Ni sorption and translocation are regulated by low-affinity transport systems, and no high-affinity transport systems have been discovered hitherto in higher plants [START_REF] Cataldo | Nickel in Plants: I. Uptake Kinetics Using Intact Soybean Seedlings[END_REF][START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF][START_REF] Deng | Effects of the interactions between nickel and other trace metals on their accumulation in the hyperaccumulator Noccaea caerulescens[END_REF]. Studying Ni isotopic fractionation and the homeostasis of the hyperaccumulator Alyssum murale grown in controlled hydroponic conditions, a preferential sorption of lighter isotopes from solution was observed (Δ 60 Ni plant-solution = -0.9 to -0.63 ‰) [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF]. It is still unclear, however, how and at which stage Ni isotopic fractionation exactly takes place. Studying plants collected in the temperate ultramafic area of Albania, [START_REF] Estrade | Weathering and vegetation controls on nickel isotope fractionation in surface ultramafic environments (Albania)[END_REF] reported an enrichment in heavier isotopes of Ni in the plants compared to the bulk soil, Δ 60 Ni whole plant-soil up to 0.40 ‰. Nevertheless, the Ni taken up by plants was lighter than the bioavailable Ni, extracted by diethylenetriaminepentaacetic acid (DTPA), and the difference Δ 60 Ni available-rhizo soil was up to 0.89 ‰. This result suggests that hyperaccumulating plants take up lighter isotopes from the heavier bioavailable Ni pool in soils. However, a recent study on hyperaccumulators in the tropical ultramafic region of Brazil, did not report on any significant Ni isotopic fractionation neither during Ni uptake, nor during Ni translocation within the plants [START_REF] Ratié | The behavior of nickel isotopes at the biogeochemical interface between ultramafic soils and Ni accumulator species[END_REF]. In our previous work, we have studied the remobilization of Ni by simulating the effect of leaf litter degradation of A. murale and R. cf. bengalensis. Obtained results showed that, in the case of R. cf. bengalensis, depending on the entity of the fractionation factor between roots and leaves, Ni recycling can result in a lighter isotope input into the surface soil [START_REF] Zelano | The influence of organic complexation on Ni isotopic fractionation and Ni recycling in the upper soil layers[END_REF]. Nickel hyperaccumulators, therefore, are responsible not only for "phyto-enrichment" of Ni concentration in surface soil, but they can also influence its isotopic composition. However, a clear mechanistic understanding of the observed Ni isotopic signatures in the soil-plant system and the links with the processes responsible for Ni isotopic fractionation have not been attained thus far. Only few publications have attempted to decipher the metal dynamics in the soil-rootshoot continuum by combining stable isotope tool and speciation data [START_REF] Aucour | Dynamics of Zn in an urban wetland soil-plant system: Coupling isotopic and EXAFS approaches[END_REF][START_REF] Aucour | Zn Speciation and Stable Isotope Fractionation in a Contaminated Urban Wetland Soil-Typha latifolia System[END_REF]. In this work, we developed an analytical strategy to follow Ni transfer from soil to plant shoots, considering the soil characteristics and the vertical distribution of Ni in the different horizons. Furthermore, in order to better understand the dynamics of Ni during the growth of a woody tropical hyperaccumulator tree, we investigated two differently aged plants. As such, this study focuses on two related soil-plant systems of R. cf. bengalensis naturally occurring on ultramafic soils, in the tropical rainforest of Kinabalu Park (Sabah, Borneo Island, Malaysia) (van der Ent and Mulligan 2015; van der Ent et al. 2015Ent et al. , 2018c;;[START_REF] Van Der Ent | X-ray fluorescence elemental mapping of roots, stems and leaves of the nickel hyperaccumulators Rinorea cf. bengalensis and Rinorea cf. javanica (Violaceae) from Sabah (Malaysia), Borneo[END_REF]. Two differently sized/aged plants were sampled on two distinct soils having two different stages of pedogenesis. On the basis of Ni concentrations, Ni speciation and δ 60 Ni in soils and plants, first results on the Ni dynamics in the soil-plant system are provided and interpreted.

Materials and methods

All reagents were prepared using ultrapure water (18.2 MΩ cm). Nitric and hydrochloric acids were of suprapure grade. Standard solutions were supplied by TechLab (1000 µg mL -1 , in a 2-5% HNO 3 ).

Plant material and soil collection

Two individuals of R. cf. bengalensis were sampled in the native equatorial rainforest of Sabah, (Borneo Island, Malaysia). The plants naturally developed on soils derived from serpentinized and non-serpentinized ultramafic bedrock (for a detailed overview of the characteristics of the local soils, see van der Ent et al. 2018a;2019b). The first specimen, R 150 , measuring 150 cm high, was collected on a deeply weathered Ferralsol near Serinsim (Nkrumah et al. 2018). The other individual, R 600 , is a tree of approximatively 6 m tall, and was collected near Lompoyou Hill, on an eroded hypermagnesian Cambisol on strongly serpentinized bedrock (Nkrumah et al. 2018;van der Ent et al. 2018a). According to a recent study (Nkrumah et al. 2019a,b), the R 150 and R 600 trees are estimated to be less than 1 year and about 3 years old, respectively. Although such trees are common in this area of the world, trees of different age were only found in distinct areas during our one-week campaign. This study is, therefore, focused on two trees of different age, respectively grown on a Ferralsol and on a Cambisol.

The R 150 plant was harvested in its entirety and was subsequently divided into roots, trunk (later subdivided in bark and wood samples) and leaves. For R 600 , multiple leaf and bark samples were collected along the entire height of the plant, from 1-6 m. Two phloem tissue samples were extracted by stripping the inner bark from the wood core at 5 cm under and 5 m above soil level. In the young tree, the phloem tissue is thin and nearly impossible to sample (van der Ent and Mulligan 2015), therefore samples were collected only for R 600. Two root samples, considered to be surface roots, were also collected at about 5 cm and 15 cm under soil level. Deeper root samples could not be collected.

The surface soils, between 0 cm and 15 cm depth, were collected at the base of R 150 and R 600 and referred to as SR 150 H1 (0-5 cm horizon), SR 150 H2 (5-15 cm horizon), and SR 600 H1 (0-5 cm horizon), SR 600 H2 (5-10 cm horizon) and SR 600 H3 (10-15 cm horizon) respectively. About 500 g for each sample were collected. Several rain forest species, including R. cf. bengalensis, are very shallow rooting and most of the nutrient cycling happens in the top layer of the soil [START_REF] Vitousek | Litterfall, Nutrient Cycling, and Nutrient Limitation in Tropical Forests[END_REF][START_REF] Vitousek | Nutrient Cycling in Moist Tropical Forest[END_REF], so only superficial soil horizons were collected. Soils and plant samples were air-dried immediately after field collection, at temperature below 40°C. This preparation mode was selected due to the sampling conditions and the sampling location was not compatible with the use of liquid nitrogen to preserve biological samples in situ. This mode of preparation is not usually recommended for XAS measurements, and the drying step may have influenced the chemical speciation of Ni, replacing water molecules by other ligands. The breaking of cell walls and membranes during the drying could have induced the dissociation of Ni complexes with low stability. However, XAS results obtained on plants show that the Ni speciation was not drastically modified by drying and the results are consistent with published data (van der Ent et al. 2017).

Due to their apparent difference in particle size distribution, soils were sieved to compare similar size fractions.

Each soil sample was, then, subdivided in two equivalent parts; one was kept and referred to as 'bulk sample', whereas the second part was sieved into four fractions, i.e. >1.5 mm; 1.5 mm -250 µm; 250-50 µm and <50 µm.

Element concentration

All soil and plant samples were dried in a dehydrating oven at 110˚C overnight, and manually ground using an agate mortar. About 50-70 mg of ground plant material was weighed in Savilex® beakers and left overnight in 3-5 mL of concentrated (70%) HNO 3 at room temperature. Samples were, then, heated to 140˚C on a hot plate to complete the acid digestion. When necessary, a second digestion step was performed, using 3 mL of Aqua Regia and 1-2 ml of HF (32%) to allow for complete sample dissolution. Solutions were finally dried at 110˚C, recovered with 0.3 M HNO 3 and measured with Inductively Coupled Plasma Mass Spectrometer (ICP-MS Quadrupole, X-Series Thermo Fisher Scientific).

Major and trace elements in soil samples were analysed by Service d'Analyse des Roches et des Minéraux (SARM), at Centre de Recherches Pétrographiques et Géochimiques, Nancy, France. The content of organic carbon in soils was measured using the EMIA-Analyser Carbon/Sulfur supplied by HORIBA, after decarbonation with HCl and filtration.

To estimate the available Ni concentration in soil samples, an aliquot of bulk soil was suspended in ultra-pure water for 24h, in triplicate, using 100 mg soil and 40 mL of milli-Q water. The final suspension was centrifuged for one hour at 52000 g and the resulting supernatant was filtered through 0.22 µm with cellulose syringe filters.

Nickel concentration was measured in solution and, as an approximation, will be indicated hereafter as 'available Ni'. Reported results of available Ni concentration correspond to the average values of experimental triplicates.

The bioavailable Ni pool in soils is operationally determined by chemical extraction using organic ligands, e.g. DTPA [START_REF] Echevarria | Assessment and control of the bioavailability of nickel in soils[END_REF]. By performing, however, a water extraction we exclude any potential isotopic fractionation arising from the Ni-ligand complex formation.

The cation exchange capacity (CEC) of the bulk soils was determined using cobalt hexammine. Briefly, 30 mL of 0.05 N hexammine cobalt(III) chloride were added to 1 g of dry soil, shaken for two hours at 30 ˚C and centrifuged at 52000 g. Supernatants were filtered through 0.22 µm and the residual concentration of hexammine analysed with UV spectrometry. This makes possible to determine the total CEC of the soil. Furthermore, element concentrations (Na, Ca, K, Mg, Fe, Al, Si and Ni) in the supernatants were measured using a Thermo Fischer ICAP 6500 Inductively Coupled Plasma Optical Emission Spectrometer (ICP-OES).

Nickel isotope measurements

Prior to Ni isotopic measurement, a two-step chromatography separation was performed to remove sample matrix. The reported procedure is based on a previously developed protocol [START_REF] Cameron | A biomarker based on the stable isotopes of nickel[END_REF][START_REF] Gueguen | Nickel Isotope Variations in Terrestrial Silicate Rocks and Geological Reference Materials Measured by MC-ICP-MS[END_REF]) and modified as described by [START_REF] Estrade | Weathering and vegetation controls on nickel isotope fractionation in surface ultramafic environments (Albania)[END_REF] and [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF]. Briefly, soil and plant samples were digested, as described above, to determine, Ni content. Depending on the measured Ni concentration (verified after digestion using a Q-ICP-MS X series 2 Thermo-Scientific), a known volume of dissolved sample was evaporated to give a mass of Ni between 1 and 2 µg. After evaporation, the residue was dissolved again in 2 mL of 3 M HNO 3 . In order to correct for any isotope composition variation potentially induced by the chemistry process, and for an instrumental mass bias, the technique of a double-spike addition was adopted. For that, a double-spike solution containing an equal amount of 61 Ni and 62 Ni isotopes was added to samples. The volume of double-spike solution to add was adapted for each sample to obtain a spike/sample ratio of 1.15. The mix was left equilibrating overnight, evaporated on a hot plate at 110˚C and dissolved in 1 mL of 6 M HCl. Solution was, then, passed on the first chromatographic column filled up with 2 ml of the anion-exchange resin AG®1-X8 and recovered. The resin, which is in chloride form, is able to retain Fe and Zn present in the sample matrix, while Ni is eluted with the solution. Additional 15 mL of 6 M HCl were passed through the column to recover all Ni. The final volume was successively evaporated on a hot plate at 110˚C and dissolved again in 1 mL HCl 1 M. The second chromatographic separation to complete the sample purification is based on the use of a Ni specific resin, supplied by Triskem Inc., France. The used resin is able to retain Ni through chelation with dimethylglyoxime at pH value 8-9. Before the second chromatographic separation, 0.2 mL of ammonium citrate ((NH 4 ) 6 C 6 H 6 O 7 ) were added to the samples, the pH was adjusted at 8-9 with 100-150 μL of ammonia, and samples were left overnight for equilibration. Solution was loaded on the column and Ni retained by the resin was successively recovered by 4 mL of 3 M HNO3 and put to evaporate on a hot plat at 110˚C. To eliminate any potential residue, acid digestion with aqua regia were performed, before recovery the sample with 0.3 M HNO 3 for measurement.

Procedural blanks were prepared in each sample batch, resulting in 0.058-0.256 nmol of Ni content that has a maximum value of 1.5 % of the total Ni, and a corresponding negligible influence on reported isotopic values, within uncertainties. The solutions were measured by Multi-Collector Inductively Coupled Plasma Mass Spectrometer (MC-ICP-MS, Neptune plus, Thermo Scientific) in medium resolution mode and using an Apex HF desolvation introduction system in a 0.3 M HNO 3 media. Ni masses (58,60,61,62) were measured simultaneously as well as Cu isotopes and one Fe isotope at masses 63, 65 and 57 respectively. The 57 Fe intensity was monitored continuously to correct for any 58 Fe contribution to 58 Ni. The correction is effective only for a signal lower than 10 mV. When the signal was higher than 10 mV, samples were processed and eluted again through the first column. A standard bracketing method was used during sample measurements and δ 60 Ni in samples were normalized to NIST-986.

Sample δ 60 Ni values were calculated as:

(1)

The analytical reproducibility was monitored by processing and measuring a Ni ICP-MS standard solution for each measurement session. Since no value exist for this solution, the standard subjected and not-subjected to the entire sample preparation procedure was measured and compare to itself, in order to control for isotopes composition modification during the process. The obtained average value was δ 60 Ni = -0.11 ± 0.05 ‰ (2σ, n = 25). The reference material BHVO-2 was processed and measured ten times in different measurement sessions, obtaining δ 60 Ni = 0.05 ± 0.04 ‰ (2σ), in agreement with published data [START_REF] Gall | Nickel isotopic compositions of ferromanganese crusts and the constancy of deep ocean inputs and continental weathering effects over the Cenozoic[END_REF][START_REF] Gueguen | Nickel Isotope Variations in Terrestrial Silicate Rocks and Geological Reference Materials Measured by MC-ICP-MS[END_REF][START_REF] Estrade | Weathering and vegetation controls on nickel isotope fractionation in surface ultramafic environments (Albania)[END_REF][START_REF] Ratié | Nickel isotope fractionation during tropical weathering of ultramafic rocks[END_REF]. Estimated recovery from the measured intensities is always higher than 90%. However, recovery is not a crucial parameter as the double spike addition technique was used.

In addition, in order to have a matrix match reference for plant samples, the Oak Leaves reference material V464 from the French National Institute for Agricultural Research (INRA) was also digested, purified and measured.

The value obtained for this material is δ 60 Ni= -0.02±0.05‰ (2σ, N=6).

All samples were measured three times. The soil-water suspension samples to determine available Ni were prepared in triplicate. For those samples, the reported δ 60 Ni values correspond to the average X of experimental triplicates x m . The associated standard error (2σ) is calculated as:

! = !" ! , ( 2
)
where sd is the standard deviation of the sample mean, and N is the number of samples.

For bulk soil and plant material samples, no experimental triplicates are available and the reported 2σ represent the analytical error deriving from three measurements of the sample. The 2σ = 0.05 ‰ obtained for the Ni ICP-MS standard (digested and measured 25 times) was applied to all sample results presenting analytical 2σ < 0.05 ‰.

The Ni isotopic fractionation between two samples A and B, Δ 60 Ni A-B , was calculated as the difference between the isotopic signatures measured in A and B, as reported below:

Δ 60 Ni A-B = δ 60 Ni A -δ 60 Ni 2+ B ( 3
)
The standard deviation σ associated to Δ 60 Ni includes the error propagation and was calculated as:

y = a±b, σ! = σ! ! + σ! ! (4)

Soil mineralogical composition

The mineralogical composition of the samples was determined by X-ray diffraction (XRD, Bruker D8 Advance)

with an X-ray tube producing Co Kα radiation (1.78886 Å) operated at 35 kV and 45 mA. The XRD patterns were obtained by scanning crushed powders of the sieved fractions and of the bulk soil, from 3° to 64° 2θ with a 0.035° 2θ step and with a 3s counting time per step. The diffracted signal was measured using a LynxEye detector with an energy discrimination window adjusted to minimize the high fluorescence background signal coming from Fe to improve the peak-to-background ratio.

Soil mineralogy was also investigated with electron microscopy, and transmission electron images were obtained with a Philips CM20 transmission electron microscope (TEM), operating at 200 kV, coupled to an Energy-Dispersive X-ray Spectrometer (EDXS). For TEM imaging and microanalysis, a few milligrams of sample were re-suspended in ethanol using ultrasonication (10 min) and a drop of suspension was evaporated on a carboncoated copper grid. The EDX spectra were obtained in nanoprobe mode (20-30 nm) using a counting time of 40-70 seconds.

Chemical speciation of nickel (X-ray absorption spectroscopy)

X-ray absorption spectroscopy (XAS) experiments were performed at two beamlines, in fluorescence mode at the SAMBA beamline (SOLEIL synchrotron, France), and in transmission mode at the XAFS beamline (ELETTRA synchrotron, Italy). In both cases, Ni K-edge X-ray absorption spectra (both in the XANES and EXAFS regions) were recorded at low temperature (using a liquid He cryostat 25 K and liquid N 2 cryostat, 77 K respectively) with a Si( 111 The XANES and EXAFS data were reduced using standard normalization procedures performed with the ATHENA and ARTEMIS programmes [START_REF] Ravel | HEPHAESTUS: data analysis for X-ray absorption spectroscopy using IFEFFIT[END_REF]. The spectra were background subtracted, and normalized. A spline function was fit through the absorption envelope and subtracted from each spectrum.

To transform data from energy space E (eV) to wave vector space k (Å -1 ), E 0 edge energy was chosen in the higher part of the edge step, at 8343 eV. The resulting χ function was weighted by k 3 to avoid oscillations damping at high χ values. The EXAFS signals obtained from plant and soil samples were fitted as linear combinations of the standard EXAFS spectra collected on solutions and solid samples. The number of components was set to a maximum of three on the basis of TEM and XRD observations. The fitting parameters were then selected on the basis of quality indicators (χ 2 , r-factor and reduced χ 2 ). Shell by shell fitting was also performed using theoretical scattering paths calculated from the unit cell structure of distinct Ni-bearing phases (Artemis code on the basis of FEFF6) [START_REF] Ravel | HEPHAESTUS: data analysis for X-ray absorption spectroscopy using IFEFFIT[END_REF][START_REF] Rehr | Parameter-free calculations of X-ray spectra with FEFF9[END_REF].

Results

Chemical and mineralogical composition of soils

The elemental concentrations were determined in the different horizons of soils associated with plants, SR 150 and SR 600 , for the bulk samples and for the size fractions i) >1.5 mm, ii) 250-50 µm, and iii) <50 µm, of both H1 horizons (Table SI 1 %) respectively. However, this content tends to increase with depth for SR 600 , reaching up to 36.9 and 35.4 % in the H2 and H3 horizons respectively. The third major element discriminating the soils is Mg, which reaches up to 1.6% (MgO %) in SR 150 and which ranges between 10.9 and 13.4 % in the SR 600 soil horizons. Organic carbon is higher in RS 600 (11.84% in H1) than in RS 150 (7.44% in H1) and in both cases organic carbon decreases with depth. The trace elements are slightly higher in SR 150 than in SR 600. In general, no marked changes within the soil profile can be observed for all trace elements measured. These results are consistent with previously published data of ultramafic rhizosphere soils, collected in the same area in Sabah, that constitute the habitat of R. cf.

bengalensis (van der Ent et al. 2015(van der Ent et al. , 2017;;[START_REF] Van Der Ent | X-ray fluorescence elemental mapping of roots, stems and leaves of the nickel hyperaccumulators Rinorea cf. bengalensis and Rinorea cf. javanica (Violaceae) from Sabah (Malaysia), Borneo[END_REF].

Nickel concentrations (Table 1) are higher in SR 150 than in SR 600 , reaching 113 µmol g -1 and only 61 µmol g -1 , respectively. In both cases, Ni concentration remains rather constant in the different horizons, but decreases with soil size fraction (from fraction >1.5 mm to fraction <50 µm). Available Ni in bulk samples was estimated by measuring Ni concentration in the supernatant from soil-aqueous suspension. For SR 150 H1 and -H2, the concentration of available Ni is about 0.35 µmol g -1 , while it is 0.40 µmol g -1 in SR 600 H1 and decreases in SR 600 H2 and SR 600 H3 to 0.19 µmol g -1 and 0.09 µmol g -1 , respectively.

The highest cationic exchange capacity (CEC) in both bulk soil samples (Table SI 2) is observed in the H1 horizons (54.6 mEq per 100 g and 22.7 mEq per 100 g in SR 600 H1 and SR 150 H1, respectively). In both cases, CEC decreases with depth as a result of the decrease of organic matter. As a general trend, SR 600 has a higher CEC than SR 150 and the difference in CEC between the two soils, about 32 meq 100g -1 , mainly derives from the higher concentration of exchangeable Mg that reaches 42.4 mEq per 100 g in SR 600 H1, but only 5.8 mEq per 100 g in SR 150 H1. Exchangeable Fe was under the detection limit in all of the samples. Exchangeable Ni is similar in both H1 horizons, but its relative contribution to total CEC is 6% in SR 150 and is less than 1 % in SR 600 .

Moreover, it slightly increases in SR 150 H2, while it decreases in SR 600 H2-H3.

The mineralogy, as revealed by both XRD and TEM data, supports the discrimination of SR 150 and SR 600 soils (Fig. 1 The XRD analysis is sensitive to crystalline phases only and soils are commonly constituted of poorly crystalline or amorphous phases. To enhance the mineralogical investigations, TEM images and combined spectra (EDX)

were acquired on the fine fraction (<50 µm) of the topsoil horizons from SR 150 and SR 600 (Figure 1). The TEM micrographs for SR 150 reveal mainly talc and goethite particles. Associated EDX spectra show that in SR 150 Ni is mainly associated with Fe-oxyhydroxides, including goethite. In SR 600 , different types of clay minerals are observed, as well as several poorly defined phases. For SR 150 , goethite appears to be the predominant Fe-bearing phase, while for SR 600 , TEM analyses reveal the presence of amorphous Fe-oxyhydroxides, frequently containing Ni (shown by EDX analysis). In the latter sample, Ni was detected in most of the investigated particles, ranging from 0.3 to 0.9% (atomic percentage, EDX data) including silicate particles.

The XAS spectra and extracted EXAFS oscillations were used to determine the solid speciation of Ni in the soils (Fig. 2). The comparison of bulk and fine fractions in the two soils show that the solid speciation of Ni was similar for both size fractions. XANES spectra from SR 150 and SR 600 appear very similar, mainly due to the fact that Ni has the same oxidation state (2+) and octahedral coordination. Some differences, however, can be observed in the XANES region and on EXAFS oscillations that are consistent with mineralogy results (TEM-EDXS and XRD). Superimposition of XANES spectra reveals slight differences in the position of the main edge and of the first oscillation (Fig. SI.3). Differences were also noticed in the EXAFS oscillations and associated Fourier transforms (Fig. 2). Fourier transform curves (imaginary and magnitude parts) show that the slight differences between these two spectra come from the nature and distribution of atoms in the 2 nd and further coordination spheres (Fig. 2). Shell by shell fitting shows that for both soil samples, six atoms of oxygen at 2.03-2.1 Å surround Ni atoms, and dominate the backscattering signal (Table 2).

Shell-by shell fitting ( respectively, and Ni in serpentine (26% for SR 600 ) (Fig. 2).

Element concentrations and Ni chemical speciation in plant samples

Multi-elemental analyses were performed on different plant material of R 150 and R 600 which conform with concentrations ranges previously reported for R. cf. bengalensis (van der Ent et al. 2017;[START_REF] Van Der Ent | X-ray fluorescence elemental mapping of roots, stems and leaves of the nickel hyperaccumulators Rinorea cf. bengalensis and Rinorea cf. javanica (Violaceae) from Sabah (Malaysia), Borneo[END_REF]. Results and detailed discussion are provided in Supplementary Information Table SI 3-4 and associated notes.

The R 150 specimen was entirely collected and the mass of plant material was weighed. The whole dry plant weight was 91.96 g. The plant was then divided in leaf, root, bark and wood samples and, except for leaves, the quantification of their relative mass was difficult to achieve. However, an estimation of the relative mass of bark and wood was made, and on the basis of the corresponding Ni concentration, the total Ni content has been calculated to be 8018 µmol. The weighted average concentration in the whole plant was 87.20 ± 70.65 µmol of Ni per g of plant. The weighted standard deviation has been calculated as reported in equation 5:

sd w = !"(!"!!") ! ! !!! (!!!) !" ! !!! ! , ( 5
)
where wi is the weight for the ith observation, N is the number of non-zero weights, and xw is the weighted mean of the observations.

In line with data previously published for R. cf. bengalensis (van der Ent et al. 2017;[START_REF] Van Der Ent | X-ray fluorescence elemental mapping of roots, stems and leaves of the nickel hyperaccumulators Rinorea cf. bengalensis and Rinorea cf. javanica (Violaceae) from Sabah (Malaysia), Borneo[END_REF], Ni concentrations leaves are high, between 134 µmol g -1 and 221 µmol g -1 in R 150 and, with the exception of leaves at 80 cm height, Ni slightly decreases from the bottom to the top of the plant (Table 4). In contrast, no clear trend can be observed in the bark samples where Ni concentrations range between 43 µmol g -1 and 98 µmol g -1 , decreasing from 20 cm to 60 cm and increasing again from 60 cm to 90 cm. Nickel concentrations in the wood samples have an increasing trend from the bottom to the top, from 6.68 µmol g -1 to 70 µmol g -1 . A different trend is observed in R 600 , where Ni concentrations, both in leaf and bark samples, clearly decreases as a function of plant height: from 522 µmol g -1 in older leaves at 2 m, to 156 µmol g -1 in younger leaves at 6 m, and Ni concentrations in bark at the same height decreases from 133 µmol g -1 to 53 µmol g -1 . Two root samples per plant were collected at about 5 cm and 15 cm under the soil surface and Ni concentrations have similar range, 154 µmol g -1 and 81.7 µmol g -1 in R 150 , and 63.2 µmol g -1 and 111 µmol g -1 in R 600 .

Nickel chemical speciation in R. cf. bengalensis leaves was investigated (Fig. 3, Table 3) and the XAS spectra are rather similar evidencing that Ni speciation is predominated by Ni complexed with low molecular weight carboxylic acids (malate and citrate). These results are consistent with those previously reported (van der Ent et al. 2017), and suggest that Ni chemical speciation was not drastically changed by the drying step. XAS spectra could not reveal any differences of chemical speciation of Ni in the investigated R. cf. bengalensis specimen.

Linear combination fitting suggests the contribution of hydrated cations Ni 2+ (H 2 O) 6 at 33% and 31% for R 600 and R 150 respectively, the presence of Ni-malate complexes at 41% and 42% for R 600 and R 150 respectively, and Nicitrate complexes at 26% and 27 % for R 600 and R 150 respectively.

The chemical speciation of Ni in R. cf. bengalensis leaves supports the idea that Ni is not strongly chelated by specific organic molecules and is not considered as a contaminant or as xenobiotic by the plants. The apparent weakly bound state of Ni makes it readily mobile, and Ni might be not permanently stored within the leaves. XAS data do not reveal any differences between the R 600 and R 150 leaves, suggesting that Ni remains relatively mobile, whatever the age of the tree.

Nickel isotopic composition in soils and plants

The The Ni isotopic composition of the two R. cf. bengalensis specimens R 150 and R 600 are reported on Table 4. The root samples from R 150 plant at -5 cm and -15 cm under the surface level has δ 60 Ni roots = -0.35± 0.05 ‰ and δ 60 Ni roots = -0.42 ± 0.05 ‰, respectively. Similarly, the surface roots of R 600 plant, both collected at -5 and -15 cm under surface level, has negative values, -0.58± 0.05‰ and -0.59 ± 0.06 ‰ respectively (Table 4).

In the aerial parts, δ 60 Ni in leaves of R 150 is between -0.66 and -0.89 ‰, clearly showing a preferential translocation of light Ni isotopes from the roots to the leaves (Tables 4 and5) and, among the leaves, from the bottom to the top, with the presence of lighter Ni isotopes in the younger apical leaves. Moreover, it can be seen from Fig. 4 that leaves, which constitute the Ni-enriched parts of the plants (table 4), have δ 60 Ni values lighter than the ligneous material, i.e. wood and bark. The wood and bark samples have relatively wide-ranging values.

However, with the exception of the wood sample at 100 cm, a preferential concentration of heavier Ni isotopes can be observed from the bottom to the top of the plant. The δ 60 Ni value of the whole plant R 150 was estimated by calculating the weighted mean of the δ 60 Ni in the different parts of the plant, resulting in a value of -0.65 ± 0.23 ‰. The weighted standard deviation was calculated as reported in eq 5.

The δ 60 Ni values in R 600 leaves range between -0.19 ‰ and -0.33 ‰, but no systematic variation is observed between the basal and apical leaves, collected from 2 to 6 m. Moreover, bark samples also have variable δ 60 Ni values, from -0.29 ‰ to 0.15 ‰, in some cases heavier and in others lighter than the leaves collected at the same height, without any clear trend. The phloem tissue sample collected from beneath the bark at -5 cm has a similar isotopic composition than root collected at the same level, δ 60 Ni phloem = -0.60 ± 0.08 ‰. The phloem tissue at 5 m high has, on the contrary, a heavier Ni isotopic composition (δ 60 Ni phloem = -0.10 ± 0.06 ‰), than leaf and bark at the same height, δ 60 Ni leaf = -0.30 ± 0.05 ‰ and δ 60 Ni bark = -0.13 ± 0.09 ‰, respectively. However, the anomalously low Ni concentration in this sample collected at 5 m suggests a collection error and, therefore, it

will not be included in the further discussion.

Discussion

Nickel isotopic composition in soil

The studied soil samples have two distinct stages of pedogenesis, which are reflected not only by the overall element concentrations and associated mineralogy, but also by their Ni isotopic composition. Weathering processes are expected to influence δ 60 Ni by depleting the heavier fraction of Ni in soil [START_REF] Ratié | Nickel isotope fractionation during tropical weathering of ultramafic rocks[END_REF] 5), suggesting that R. cf. bengalensis takes up lighter Ni isotopes. This hypothesis can be reasonably extrapolated to the case of R 600 , for which the calculation of a mass balance was not possible, but in which all plant material have an isotopic signature lighter than the available Ni in soil. In a previous study it has been shown that during leaf litter degradation process, about the 80 % of Ni released has the same δ 60 Ni value of the original leaves, and that after a few days, a release of light isotopes occurs [START_REF] Zelano | The influence of organic complexation on Ni isotopic fractionation and Ni recycling in the upper soil layers[END_REF]. Therefore, the enrichment of lighter isotopes in the whole plant likely produces a release of lighter Ni isotopes during leaf litter degradation to topsoil, H1, compared to the deeper horizons, H2 and H3. However, this contribution might also vary depending on the potential isotopic fractionation between leaves and roots.

Nickel fractionation within plants

From the isotopic composition of both specimen R 150 and R 600 , it can be concluded that R. cf. bengalensis preferentially takes up the lighter Ni isotopes from the available Ni fraction in soil through absorption by root cells, as reported in Table 5. This result is consistent with previously published data for Ni hyperaccumulators grown in hydroponics, where it was also shown that the entity of the fractionation was due to a competition effect, induced by the concomitant presence of other metals, e.g. Zn [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF]). In the case of Zn accumulation in wheat, a non-hyperaccumulating plant, it has been reported that the variation of Zn supply can be a source of different trends of Zn isotopic fractionation in the soil-plant system. The lower availability of Zn could enhance the release of organic ligands, such as phytosiderophores by the plant [START_REF] Arnold | Evidence for the mechanisms of zinc uptake by rice using isotope fractionation[END_REF], and thus the Zn uptake from soil, with subsequent enrichment of the plant in heavy isotopes [START_REF] Wiggenhauser | Zinc isotope fractionation during grain filling of wheat and a comparison of zinc and cadmium isotope ratios in identical soil-plant systems[END_REF]). However, hyperaccumulators often have more specific processes of uptake and translocation of metals than non-accumulating plants. For example, the Zn hyperaccumulator N. caerulescens has the ability to use both high-and low-affinity transport systems depending on Zn availability, and this behaviour probably induces different trends of Zn isotopic fractionation in the plants [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF]). In our case, the investigated R. cf.

bengalensis grew on ultramafic soils, and neither the scarcity of Ni nor the competition effect with other metals can be considered as relevant factors determining Ni isotopic fractionation in the soil-plant system. In hyperaccumulating plants, it has been assumed that Ni is taken up through a symplastic pathway, and that Ni isotopic composition in whole plant derives from the uptake mechanisms of root cell membrane [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF], promoting the uptake of lighter isotopes. Different pathways of fractionation within the plant parts can, then, take place, but they are far from being fully understood. It has been recently reported that in wheat plants, important Zn isotopic fractionation within plant parts can be attributed to the absorption of heavy isotopes to cell walls in the apoplastic space [START_REF] Wiggenhauser | Zinc isotope fractionation during grain filling of wheat and a comparison of zinc and cadmium isotope ratios in identical soil-plant systems[END_REF]). However, through experiments based on the use of radioactive 63 Ni, it was reported that Ni is transferred from roots to shoots via xylem, and also that its mobility is greater than that of Zn, Mn, Co and Cd [START_REF] Page | Selective transport of zinc, manganese, nickel, cobalt and cadmium in the root system and transfer to the leaves in young wheat plants[END_REF]. These observations suggest that in hyperaccumulators, Ni undergoes a very efficient translocation process that might not be the only source of isotopic fractionation, or at most, it is responsible for the smaller isotopic fractionation observed for Ni compared to e.g. Zn [START_REF] Deng | Nickel and Zinc Isotope Fractionation in Hyperaccumulating and Nonaccumulating Plants[END_REF][START_REF] Wiggenhauser | Zinc isotope fractionation during grain filling of wheat and a comparison of zinc and cadmium isotope ratios in identical soil-plant systems[END_REF] and Cd [START_REF] Wiggenhauser | Cadmium Isotope Fractionation in Soil-Wheat Systems[END_REF]. Whereas some publications report demonstrations or assumptions about the translocation mechanisms of Ni [START_REF] Kerkeb | The Role of Free Histidine in Xylem Loading of Nickel in Alyssum lesbiacum and Brassica juncea 1[END_REF][START_REF] Alves | Nickel speciation in the xylem sap of the hyperaccumulator Alyssum serpyllifolium ssp. lusitanicum growing on serpentine soils of northeast Portugal[END_REF], nothing is known about this mechanism for R. cf. bengalensis. In this study, reported XAFS spectra on R. cf. bengalensis (Fig. 3, Table 3) show that Ni in leaves is unequivocally present as a complex with low molecular weight carboxylic acids (i.e. malate and citrate), confirming results reported by van der Ent et al. ( 2017).

In the case of Zn and Cd, the isotopic fractionation in wheat shoots has been attributed to their different affinity for functional groups of organic ligands [START_REF] Wiggenhauser | Zinc isotope fractionation during grain filling of wheat and a comparison of zinc and cadmium isotope ratios in identical soil-plant systems[END_REF], or different chemical speciation. The hypothesis about the possible influence of complexation with organic ligands on Ni isotopic fractionation was previously investigated [START_REF] Zelano | The influence of organic complexation on Ni isotopic fractionation and Ni recycling in the upper soil layers[END_REF]). However, a rather low potential of fractionation induced by complexation reaction between Ni and citric acid, oxalic acid and purified humic substances was reported with a maximum value of Δ 60 Ni bond-free < 0.2 ‰. Such a small fractionation can be excluded as a source of Δ 60 Ni within plant material, discarding the hypothesis raised up for Zn fractionation in wheat shoots. Ni is never stored in roots, but it is rapidly transferred to them, as it expected for Ni hyperaccumulators that do not require defence mechanisms against high Ni concentration in soil. In the early stage of growth of the plant (R 150 ), 80 % of the total amount of Ni is mobilized and more than 70 % is concentrated in leaves. This transfer produces a remarkable isotopic fractionation between leaves and roots, from which a preferentially translocation of lighter isotopes, Δ 60 Ni root-leaf up to 0.51 ± 0.07 ‰, can be observed (Table 5). At the same time, an enrichment of heavier isotopes in the ligneous parts of the plant is present. As the total amount of Ni in leaves is definitively higher than in wood and bark, these results strongly suggest that lighter isotopes are transferred first, going from roots to leaves. Data reported in Fig. 4 suggest, moreover, that the fractionation is correlated to the rate of Ni translocation (lighter isotopes move faster) as an important fractionation is also observed as a function of height, between younger (apical) and older (basal) parts of the plant. This implies that as soon as new leaves appear, Ni is mobilized through leaves, and lighter isotopes arrive first. This trend is no longer observed in the older specimen (R 600 ), where the isotopic fractionation between basal and apical leaves is not evident anymore. This result is surprising as Ni concentration still shows a clear trend between the apical and basal leaves, higher in basal, and lower in apical ones. Moreover, the isotopic fractionation between leaves and other plant material is less pronounced in the older specimen than in the younger plant (see Fig. 4). This observation suggests that the fractionation observed in R 150 is not necessarily attributable to different transport or retention mechanisms, as it is hypothesized for other metals, e.g. Cd [START_REF] Wiggenhauser | Cadmium Isotope Fractionation in Soil-Wheat Systems[END_REF]. If the fractionation were due to transport or retention mechanisms, it would be accentuated in the older plant, but the opposite result is observed. In addition, in R 600 , all plant material has lighter isotopes than soil available Ni, but all aerial parts (leaves, bark, wood) are enriched in heavier isotopes compared to roots.

Taken together, the available data suggest that during the first period of growth, when the plant starts to take up Ni, the transfer of light isotopes from roots to leaves can be due to kinetic effects. Once the amount of Ni accumulated in the plant has increased (see Ni concentration in R 600 , Table 4), the Ni already present in the plant circulates through the phloem tissue into all plant parts. Successive loading and unloading cycles would have, therefore, the effect to homogenize the isotopic fractionation among leaves, and between leaves and the other plant components, erasing the difference between new and old leaves. In old specimens, the consequent increasing number of cycles that Ni makes within the plant through the circulation of phloem sap from top to bottom, and inversely, would reduce the isotopic fractionation. The observed data in R 600 are probably the results of several additive processes and support the idea that in the hyperaccumulator R. cf. bengalensis, Ni is involved in plant homeostasis as any other cation. However, to confirm this hypothesis, supplementary data should be collected on a wider population of trees of R. cf. bengalensis, collected in their natural habitat and at different stage of growth.

Conclusions

This study aimed to unravel the biogeochemical processes resulting in Ni isotopic fractionation in two different soil-plant systems associated with the Ni hyperaccumulator plant R. cf. bengalensis. The investigated soils have distinct stages of pedogenesis, which are both highlighted by the mineralogical composition and the isotopic signature of the available pool of Ni. The Ferralsol SR 150 reflects a stable system, which has reached a steadystate equilibrium, subsequent to the complete weathering of primary minerals and for which Ni is mainly associated to poorly crystalline Fe-oxyhydroxides. The isotopic signature has no variation, neither in soil bulk surface horizons, nor in the available pool of Ni. In the partially weathered hypermagnesian Cambisol SR 600 , Ni is associated not only to Fe-oxyhydroxides, but also to the identified phyllosilicates, and even though the Ni isotopic composition of bulk soils does not vary, an enrichment in heavy isotopes is observed with depth for the Ni available fraction. In both soil systems, the two investigated specimens of the hyperaccumulator R. cf.

bengalensis accumulate lighter Ni isotopes from the soil available Ni pool. The Ni isotopic composition in the different parts of the plants were investigated and correlated to the plant height and age. Despite the limited number of investigated samples, the study highlights the potential effect of plant age on Ni isotopic fractionation in hyperaccumulator plants. A translocation of lighter isotopes of Ni from roots to leaves was observed in the young plant, Δ 60 Ni root-leaf up to 0.51 ± 0.07 ‰, and isotopic fractionation was also unravelled between the apical and basal leaves, Δ 60 Ni basal-apical up to 0.23 ± 0.07 ‰ (Table 5). However, this effect was not detected in the older plant, where no isotopic fractionation was observed as a function of plant height, and where the Ni isotopic fractionation seems to be mask by successive cycles of Ni circulation through the plant. These results suggest that once Ni is taken up from the soil by roots, it is rapidly transferred to leaves, without being permanently stored there. Instead, it has an active and continuous cycle within the plant through phloem-mediated redistribution. Once the plant has accumulated a substantial quantity of Ni, the isotopic fractionation induced by

Ni translocation among aerial plant material is masked by an isotopic homogenization, and the observed isotopic signature is the result of consecutive internal translocation cycles. However, it is still not clear why the roots of the older specimen have a lighter isotopic signature than in aerial parts.

The acquired results suggest that it is inordinately difficult to discriminate between specific, individual processes contribution N°2567. The authors also would like to thank the two anonymous reviewers for their useful suggestions that helped improving the quality of the manuscript. The amplitude reduction factor S 0 2 was fixed to 0.9 as well as the energy shift on the basis of reference spectra analysis. The number of atoms of the first coordination shell was fixed to 6, but the number of atoms of other coordination shells was released. The distance R and disorder Δ 2 were varied for all the coordination shells. 2-Linear combination fitting was obtained on the fitting range Δk using as a maximum three components: Ni sorbed onto a purified clay in position of exchangeable cation, Ni sorbed onto goethite and Ni as structural cation in serpentine mineral. 3 for fitting parameters).

Notes

The concentrations of Ca and K in the plant material are relatively high compared to the amount of exchangeable cations in the corresponding soils. In both plants, the highest Ca concentration is in the bark, where it reaches 2000 µmol g -1 and 1700 µmol g -1 in R 150 and R 600 , respectively. The highest K concentrations is recorded in the leaves, 546 µmol g -1 in R 150 and 686 µmol g -1 R 600 . Despite the fact that exchangeable Mg in the soil is 10-fold higher in SR 600 than in SR 150 , Mg concentrations are similar in both plants, e.g. ~150 µmol g -1 in the leaves. Other elemental concentrations, such as Fe, Mn, Cu and Zn are in the ranges previously reported for R. bengalensis (van der Ent et al., 2017). Some clear trends are found in R 150 , the Mg and K concentrations in the bark and wood increase with height, from the bottom to the top. This trend is also observed for Mg in bark of R 600 , but no trend is observed for K. For the trace elements no clear trends is found. Cobalt is more concentrated in R 150 leaves than in R 600 leaves, although Co contents in SR 150 and SR 600 soils are in the same range of order.

δ

  60 Ni = ( 60/58 Ni sample 60/58 Ni NIST 986 -1) ×1000 in (‰)

  ) double crystal monochromator. A Ni foil was positioned after the sample and collected simultaneously to each spectrum in order to have perfect energy calibration; the first inflection point of metal Ni spectrum was calibrated at 8333 eV. Soil samples were scanned at SAMBA in fly scan mode from 8000 to 9500 at velocity of 5 eV/s, resulting in energy step of 0.5 eV, using an X-ray fluorescence detector (VORTEX, Hitachi). For each soil sample, between 22 and 35 spectra were recorded. Dried leaf samples were measured at the XAFS beamline in transmission detection mode, with three scans recorded in a step by step mode, with 5 eV steps for the first 200 eV, 0.2 eV for the XANES range and with a k-constant step of 0.03 (Å -1 ) in the EXAFS range. In all cases, the samples were ground and pelletized before the XAS measurements. Reference spectra were obtained on the SAMBA and XAFS beamlines from previous sessions, under the same acquisition conditions. These standards included Ni aqueous solution with and without organic ligands, Ni incorporated into goethite, Ni sorbed onto phyllosilicate (charged and non-charged minerals), Ni sorbed onto calcite, Ni sorbed onto iron oxyhydroxide (goethite), and natural Ni containing minerals including chrysoprase, nimite, and Nibearing serpentine (clay mineral).

  and Fig.. SR 600 XRD patterns (in all horizons and size fractions) show the presence of phyllosilicate phases, commonly encountered in ultramafic soils. The clay minerals are represented by chlorite, talc and 7Å minerals (i.e. serpentine and kaolinite) and are associated with other silicates such as quartz, feldspars and amphiboles, as well as Fe-oxyhydroxide (goethite). The mineralogy of SR 600 relates to the presence of the serpentinized bedrock beneath and to a less advanced weathering of this soil. The XRD patterns have intense diffraction peaks from amphibole and chlorite minerals (Fig.SI 2). For the second soil SR 150 , quartz, talc and goethite diffraction lines dominate XRD patterns. Those patterns are all drastically different from those obtained for the SR 600 samples, with a smaller number of crystalline phases (Fig.SI 1), and the presence of goethite is consistent with the higher Fe content in the SR 150 soil samples. For both soils, XRD data do not reveal any variation in the different horizons and size fractions. The XRD analysis of the clay fraction of SR 600 shows the predominance of chlorite minerals, and additional 7Å minerals (from the serpentine or kaolinite groups).

  leading to Ni isotopic fractionation during Ni sorption and translocation within the plant, especially for hyperaccumuating plants. The comparison of two plants with a different stage of growth seems to highlight the role of plant`s age on Ni isotopic fractionation. However, to better constrain Ni homeostasis and the contribution of R. cf. bengalensis to Ni isotopic signature in surface soil, more specimens should be investigated and compared, both in the early and in the advanced stages of growth.

Fig. 1

 1 Fig.1 XRD patterns obtained on bulk soils, H1 horizons of SR 150 and SR 600 (A). TEM micrographs of the fine fraction <50 µm of SR 150 (B) and SR 600 (C) soils, H1 horizons.

  Fig.2EXAFS oscillations obtained at Ni-K edge, Linear combination fitting (left graphs) and theoretical fitting (or shell by shell fitting, right graphs), for SR 150 (up) and SR 600 (down). Fitting parameters are reported in Table2.

Fig. 3

 3 Fig.3 EXAFS oscillations at Ni K edge of R.cf. bengalensis leaves from R 150 and R 600 specimen. Oscillations are fitted as a combination of reference spectra (see Table3for fitting parameters).

Fig. 4

 4 Fig.4 Nickel isotopic composition, δ 60 Ni, in leaf, root, bark and wood samples of R. cf. bengalensis specimen R 150 , and in leaf, bark and phloem tissue samples of the R 600 specimen. All results are the average of three measurements for each sample and the relative standard deviations correspond to the instrumental error. When the 2σ associated to δ 60 Ni values are ˂ 0.05 ‰, the value of 2σ = 0.05 ‰ obtained for Oak Leaves reference material V464 (δ 60 Ni = -0.02 ± 0.05 ‰, N=6) is used.

Figure SI 1 .

 1 Figure SI 1. XRD patterns obtained on SR 150 bulk soil, horizon H1 (0-5 cm) and H2 (5-15 cm). The pattern demonstrates the predominance of goethite and quartz, associated to one phyllosilicate (talc), no clay minerals or other silicate could be evidenced.

Figure

  Figure SI 2. XRD patterns obtained on SR 600 bulk soil, horizon H1 (0-5 cm), H2 (5-10 cm) and H3 (10-15 cm). The patterns evidence a predominance of phyllosilicates, chlorite, talc and 7Å phyllosilicate (serpentine or kaolinite). Primary minerals such as amphibole and feldspars were also detected. Quartz and goethite appear as minor phases.

Figure

  Figure SI 3. XANES spectra at Ni-K edge from SR 150 and SR 600 soils (surface horizon H1, bulk soil samples).

  ). The major element contents are typical for soils developed on ultramafic bedrock (van derEnt et al. 2018a). The Fe content is relatively high, especially in SR 150 , where it reaches up to 34 and 39.8 % (Fe 2 O 3 %), while it ranges from 4.6 and 22.1 % in SR 600 , with a clear increasing trend with depth in the latter case. The Si content is rather similar in the surface horizons of SR 150 and SR 600 , with up to 28.9 and 29.2 % (SiO 2

  Table2) also indicates the presence of Fe atoms at 3.07 and 3.54 Å in the case of SR 150 , supporting the hypothesis that a fraction of Ni is adsorbed onto Fe oxyhydroxides, or is inserted in the crystal lattice of Fe oxyhydroxides. This conclusion is consistent with the higher concentration of Fe (i.e. goethite) in this sample SR 150 (See tableSI 1). For SR 600 , FEFF fitting also shows the presence of low Z atoms at 3.26 Å, supporting the hypothesis that a part of Ni is incorporated in the silicate structure. This result is also correlated with the high proportion of phyllosilicates in sample SR 600(XRD and TEM data). Linear combination fitting (Table2) reveals matches with reference spectra in the samples SR 600 and SR 150 : Ni sorbed onto smectite (59 % and 50% for SR 600 and SR 150 respectively), Ni sorbed onto goethite (16% and 50% for SR 600 and SR 150

  δ 60 Ni values in bulk soils (δ 60 Ni bulk ) and in the available fraction (δ 60 Ni available ) are shown in Table 1. The bulk soil horizons of SR 150 and SR 600 have slightly different Ni isotopic signatures, δ 60 Ni bulk_RS150_H1 =δ 60 Ni bulk_RS600_H2 = 0.01 ± 0.06 ‰, respectively. In both cases, δ 60 Ni does change neither with depth, nor in the <50 µm fractions of H1 compare to the bulk. The SR 150 H1 available Ni (δ 60 Ni available_RS150_H1 ) is slightly enriched in lighter Ni isotopes in comparison with the corresponding bulk soil SR 150 (δ 60 Ni available_RS150_H1 = -0.32 ± 0.09 ‰), obtaining Δ 60 Ni bulk-available_RS150_H1 = 0.15 ± 0.10 ‰. However, this result is not observed for the lower horizon SR 150 H2, where available Ni is similar to the bulk soil, δ 60 Ni available_RS150_H2 = -0.14± 0.05 ‰, and

	0.17 ± 0.05 ‰,	δ 60 Ni bulk_SR150_H2 = -0.16 ± 0.05 ‰,	and	δ 60 Ni bulk_RS600_H1 = -0.01 ± 0.05 ‰,

Δ 60 Ni bulk-available_RS150_H2 = -0.02 ± 0.07 ‰. On the contrary, SR 600 available Ni has an enrichment in heavier Ni isotopes, increasing with soil depth, δ 60 Ni available_RS600_H1 = 0.32 ± 0.05 ‰, δ 60 Ni available_RS600_H2 = 0.52 ± 0.05 ‰ and δ 60 Ni available_RS600_H3 = 0.62 ± 0.05 ‰. The isotopic difference with the corresponding bulk soils were calculated as Δ 60 Ni bulk-available_RS600_H1 = -0.33 ± 0.06 ‰ and Δ 60 Ni bulk-available_ RS600_H2 = -0.51 ± 0.07 ‰.

  positions and/or adsorbed onto the surface forming outer-sphere complexes[START_REF] Chardot | Nickel bioavailability in an ultramafic toposequence in the Vosges Mountains (France)[END_REF][START_REF] Raous | Potentially toxic metals in ultramafic mining materials: Identification of the main bearing and reactive phases[END_REF][START_REF] Bani | Pedogenesis and nickel biogeochemistry in a typical Albanian ultramafic toposequence[END_REF], are the main crystalline phases. To a lesser extent, Ni is also associated to amorphous Fe oxides (Fig.1). Both total Ni concentration and δ 60 Ni values in bulk soils remain constant in the three horizons of SR 600 , i.e. H1, H2 and H3. However, from H1 to H3, available Ni concentration decreases with depth and has a clear enrichment of heavy isotopes, with δ 60 Ni values increasing from 0.32 ‰ to 0.62 ‰ from H1 to H3, respectively. These values are directly correlated with a decreasing of exchangeable Ni (see CEC values in TableSI2), a decreasing of organic C, and an increasing of Fe content. Performing water extraction provides an estimation of the available Ni mainly involved in weak bonds with mineral surface groups or organic matter. Such a contribution to the available pool of Ni in surface horizon of ultramafic soils has already been noted[START_REF] Zelano | Study of Ni exchangeable pool speciation in ultramafic and mining environments with isotopic exchange kinetic data and models[END_REF]. The contribution of organic matter (OM) to available Ni is, therefore, likely more important in H1 and H2 than in H3, and it can be hypothesized that the increasing positive values of δ 60 Ni available in H2 and H3, are due to a lower proportion of weakly bound Ni in those horizons compared to SR 600 H1. In the light of the results, it can be assumed that the lighter pool of available Ni in SR 600 H1 is the result of combined

	first SR 150 soil horizons H1 and H2, which have similar total and available Ni concentrations, and similar
	isotopic composition. The available Ni from H1 is slightly enriched in light isotopes (Δ 60 Ni bulk-
	available = 0.15±0.10 ‰), while no fractionation is observed for H2 (Δ 60 Ni bulk-available = -0.03 ±0.07 ‰). Such a
	stable isotopic composition of Δ 60 Ni bulk-available ~ 0, can be the result of the loss of the heavier water soluble Ni

. This hypothesis is consistent with the lighter Ni isotopic signature of SR 150 bulk (δ 60 Ni = -0.17 ± 0.05) compare to SR 600 bulk (δ 60 Ni = -0.01 ± 0.05). The SR 150 is a Ferralsol, collected near Serinsim region, and can be considered to have reached steady state equilibrium, as in Ferralsols all the alterable minerals have been weathered and secondary minerals carry most of the Ni in this soil. This assumption is supported by the soil mineralogy, characterized by a quasi-absence of phyllosilicates, and by the Ni solid speciation, showing that Ni is mainly bound to poorly crystalline Fe oxyhydroxides. Moreover, no difference is detected between the two from the soil, due to full weathering of the primary minerals, achieved after thousands of years. In contrast, the soil SR 600 is a partially weathered hypermagnesian Cambisol and the soil isotopic composition suggests the contribution of other processes than in SR 150 . Primary and secondary phyllosilicates, in which Ni is often present in structural processes, such as the prolonged Ni uptake by plant and the leaf litter degradation, during which Ni is released to surface soil, in primis to H1. Reported results have shown that Ni isotopic composition of the whole plant R 150 is lighter than the available Ni in soil (Table

  The observed Δ 60 Ni values within the two investigated specimen of R. cf. bengalensis cannot be fully explained by a complexation reaction or by different affinity towards organic molecules. Despite only two specimens have been investigated in this work, one possible interpretation for the observed Δ 60 Ni might come from the different age of the two R. cf. bengalensis plants. Even though they grew on different soils, both plants take up lighter Ni isotopes and the reported results suggest a correlation between the plant age and the observed Δ 60 Ni values. Both specimens have the highest concentration of Ni in leaves, confirming that

Table 1 .

 1 Nickel concentration, µmol g -1 , and isotopic composition δ 60 Ni, in: i) bulk soil horizons collected at the bottom of R. cf. bengalensis R 150 and R 600 ; reported results are the average of three measurements for each sample with the corresponding instrumental standard deviation σ. The standard deviation obtained for the Ni ICP-MS standard solution (δ 60 Ni = -0.11 ± 0.05 ‰) was applied to all sample results presenting analytical error 2σ < 0.05 ‰. ii) Concentration and isotopic composition of Ni in solution after 24h of soil suspension in ultrapure water. Results are the average of triplicate samples and σ values were calculated as described in eq. 2, ! =

	!" !	, where sd is the standard deviation of the sample mean,

Table 2 .

 2 XAS data on SR 150 and SR 600 bulk soils, H1 horizons. 1-Summary of the local structural parameters derived from EXAFS data analysis (shell by shell fitting). The Ni EXAFS data were Fourier transformed over the k range Δk and fitted over the ΔR range to four single scattering paths involving O, Fe or Si atoms.

Table 3 .

 3 XAS data on R. bengalensis leaves -Linear combination fitting was obtained on the fitting range Δk using as a maximum three components from a library of reference spectra obtained from organic complexes of Ni as well as from Ni aqueous solution. Ni-malate, Nihistidine and Ni-citrate solutions were prepared at pH 6 with a metal/ligand ratio of 7.

	1-Structural parameters

Table 4 .

 4 Plant mass (g), Ni concentration (µmol g -1 ) and isotopic composition (δ 60 Ni) of R. bengalensis R 150 and Ni concentration (µmol g -1 ) and δ 60 Ni values of R 600 specimens are reported, in blue and green, respectively. All results are the average of three measurements for each sample and the relative standard deviations correspond to the instrumental error. When the 2σ associated to δ 60 Ni values are ˂ 0.05 ‰, the value of 2σ = 0.05 ‰ obtained for Oak Leaves reference material V464 (δ 60 Ni = -0.02 ± 0.05 ‰, N=6) is used. The Ni concentration and the δ 60 Ni of the whole plant have been calculated as the weighted average values. The corresponding 2σ values are calculated according to equation 5.

	\

Table 5 .

 5 Nickel isotopic fractionation Δ 60 Ni, calculated between bulk soil and average δ 60 Ni in roots, Δ 60 Ni bulk -root , between available Ni and roots, Δ 60 Ni available -root , and between roots and leaves, Δ 60 Ni root-leaf . Reported 2σ values were calculated applying the error propagation formulas reported in equation 4.

	SR 150 Δ 60 Ni bulk soil-root	2σ	Δ 60 Ni available -root	2σ Plant height Δ 60 Ni root-leaf	2σ
	H1	0.21	0.12	0.06	0.12	50cm	0.28	0.08
	H2	0.22	0.09	0.24	0.09	60 cm	0.37	0.08
						80 cm	0.40	0.08
						110 cm	0.44	0.08
						150 cm	0.51	0.07
	SR 600	Δ 60 Ni bulk -root	2σ	Δ 60 Ni available -root	2σ Plant height Δ 60 Ni root-leaf	2σ
	H1	0.58	0.08	0.91	0.09	2 m	-0.26	0.08
	H2	0.60	0.09	1.11	0.08	3 m	-0.40	0.08
	H3			1.21	0.07	4 m	-0.39	0.08
						5 m	-0.29	0.07
						6 m	-0.34	0.11

Table 2 .

 2

Table SI 2

 SI TableSI 1. Elemental composition of SR 150 and SR 600 soil samples in µmol kg -1 for trace elements, oxide percentages for major elements and percentage of organic carbon. The range of standard deviation associated to reported results is between 5% and 20% for trace elements (5% for Ni), between 2% and 10% for major and minor elements, and 2 % for organic carbon, as reported by the Service d'Analyse des Roches et des Minéraux, at Centre de Recherches Pétrographiques et Géochimiques, Nancy, France. . Exchangeable cations and total soil cation exchange capacity (CEC mEq per 100 g of soil), determined in bulk samples of different horizons of SR 150 and SR 600 . = below the limit of detection Table SI 3. Element concentrations (µmol g -1 ) in leaf, bark, wood and root samples of R. bengalensis R 150 plant. Results are the average of three measurements for each samples. Standard deviation correspond to the triplicate of the measurements. = below the limit of detection Table SI 4. Element concentrations (µmol g -1 ) in leaf, bark, and phloem samples of R. bengalensis R 600 plant. Results are the average of three measurements for each samples. Standard deviation correspond to the triplicate of the measurements.

	µmol kg -1 and % SR150 H1 bulk SR150 H2 bulk SR150 H1 >1.5mm SR150 H1 250 -50µm SR150 H1 < 50µm SR600 H1 bulk SR600 H2 bulk SR600 H3 bulk SR600 H1 >1.5mm SR600 H1 250 -50µm SR600 H1 < 50µm <d.l. = below the limit of detection Cu Pb Sc 591 24.71 663 556 23.36 773 517 24.07 686 818 33.18 630 1037 24.40 598 573 12.79 424 510 13.48 566 548 13.70 596 586 11.57 463 902 28.74 370 1695 41.18 380 Soil horizon Na + K + SR 150 H1 0.08 0.79 SR 150 H2 0.03 0.24 SR 600 H1 0.09 0.59 SR 600 H2 0.05 0.17 SR 600 H3 0.05 0.06 <d.l. Tree part Height Mg K Leaf cm 132±0.9 483±3.8 Co 7.46 8.89 7.67 6.91 5.46 4.65 6.74 8.39 6.12 2.97 2.84 Ca 2+ 14.4 3.66 10.0 3.22 1.21 Ca Cr 283 364 302 313 206 276 358 381 296 245 278 749±22.4 Zn 3.59 3.66 3.62 3.80 3.36 3.95 4.89 4.70 3.83 3.32 3.82 Mg 2+ 5.8 2.08 42.4 20.1 16.1 cm 131±0.24 459±2.0 785±21.3 cm 85±0.73 516±1.4 621±19.7 110 cm 139±0.60 366±3.5 691±22 150 cm 172±1.78 546±6.3 651±31.0 Bark cm 41.5±0.17 244±1.2 1697±38.4 2.0±0.03 %SiO2 28.9 30.5 30.2 26.7 34.6 29.2 36.9 35.4 31.2 26.5 35.8 Fe 3+ %Al2O3 4.36 5.15 4.53 4.20 3.80 4.96 6.58 7.02 5.40 4.40 4.55 <d.l. <d.l. <d.l. <d.l. <d.l. Mn Fe 12.7±0.14 10.6±0.16 171±1.10 2.56±0.01 0.1±0.01 %Fe2O3 %MnO %MgO %CaO %Na2O 34.0 0.47 1.43 0.49 < d.l. 39.8 0.52 1.61 0.14 < d.l. 35.1 0.48 1.46 0.41 < d.l. 31.5 0.43 1.42 0.62 < d.l. 29.9 0.35 1.16 0.55 < d.l. 14.6 0.35 10.77 1.06 0.21 19.3 0.51 13.21 1.09 0.28 22.1 0.64 13.41 0.97 0.29 16.3 0.49 11.48 1.03 0.21 11.3 0.24 9.61 1.26 0.22 10.8 0.24 10.25 1.33 0.28 Al 3+ Si 4+ Total Ni 2+ CEC <d.l. 1.24 0.30 22.7 0.07 1.15 0.49 7.72 <d.l. 1.09 0.38 54.6 <d.l. 1.15 0.23 24.9 0.05 1.17 0.12 18.7 Ni Zn Cr 12.5±0.06 13.3±13.6 156±0.62 2.83±0.03 0.2±0.01 10.1±0.02 8.37±0.37 221±16.0 6.73±0.07 0.1±0.01 14.8±0.12 10.6±0.85 134±2.40 3.03±0.05 0.1±0.01 14.8±0.31 9.08±1.85 138±2.75 3.61±0.05 0.1±0.01 19.9±1.16 68.7±0.78 6.76±0.10 0.2±0.01 cm 41.1±0.12 203±1.2 2093±10.4 2.3±0.01 27.2±0.83 62.1±0.49 7.42±0.06 0.2±0.0 cm 62.9±0.13 221±2.4 1669±41.6 4.4±0.06 17.6±1.48 58±0.37 3.54±0.07 0.0±0.0 cm 46.1±0.26 243±1.0 1519±32.4 5.4±0.06 14.5±1.24 43.0±0.46 2.38±0.01 0.0 cm 71.2±0.03 344±3.3 1404±48.6 3.0±0.04 18±2.15 67.5±1.07 3.69±0.07 0.7±0.64 cm 107±1.28 507±2.3 1364±25.4 2.01±0.02 15.5±1.42 98.6±0.73 4.43±0.01 0.0±0 Wood cm 22.6±0.15 128±2.0 333±20.9 0.2±0.01 4.47±0.58 6.7±0.04 0.25 <d.l. cm 22.9±0.13 154±2.8 454±26.1 0.3±0.01 7.48±0.62 9.4±0.12 0.36±0.01 <d.l. cm 34.4±0.08 224±37.3 666±32.9 0.4±0.02 8.96±1.02 15.6±0.15 0.59±0.01 <d.l. cm 36.1±0.37 205±3.2 673±20.2 0.4±0.01 8.14±0.80 11.9±0.11 0.89±0.02 <d.l. cm 47.9±0.66 305±1.7 623±18.3 0.5±0.01 8.16±0.86 28.1±0.38 0.88±0.01 <d.l. 100 cm 65.7±0.30 412±3.3 629±34.1 0.8±0.02 10.0±1.08 69.9±0.62 1.67±0.02 <d.l. Root -5 cm 99.1±1.25 237±2.7 963±31.7 4.5±0.04 44.2±1.66 154±1.93 2.07±0.03 0.8±0.01 -15 cm 85.7±0.66 155±1.4 414±16.4 3.2±0.05 169±2.78 81.7±0.95 2.37±0.02 4.1±0.35 part Height Mg K Ca Mn Fe Ni Zn Cr Leaf 2m 149±0.75 684±10.7 995±36.8 1.98±0.03 9.99 ±1.18 522±7.83 3.76±0 0.06±0.01 0.05±0.01 0.32±0.01 %K2O %TiO2 %Corg 0.10 0.15 7.44 0.07 0.18 2.91 0.09 0.16 / 0.10 0.14 / 0.10 0.14 / 0.04 0.20 11.84 < d.l. 0.24 3.01 < d.l. 0.23 1.64 0.04 0.20 / 0.05 0.20 / 0.06 0.16 / Cu Co 0.07±0.01 1.78±0.03 0.07±0.01 1.74±0.01 0.06±0.01 1.59±0.01 0.09±0.01 1.68±0.03 0.09±0.01 1.47±0.04 0.06±0.01 0.18±0.0 0.09±0.0 0.19±0.0 0.11±0.01 0.27±0.01 0.12±0.01 0.24±0.01 0.15±0.01 0.23±0.01 0.13±0.73 0.3±0.01 0.02±0 0.02±0.0 0.05±0.01 0.03±0 0.08±0.01 0.05±0 0.04±0.01 0.04±0 0.20±0.02 0.07±0.0 0.20±0.02 0.16±0.0 0.06±0.0 0.38±0.0 0.07±0.0 0.47±0.01 Cu Co 3m 183±1.42 463±3.37 923±22.1 1.83±0.02 8.93±0.44 349±5.50 2.15±0 0.09±0.01 0.05±0.0 0.19±0.01 4m 144±1.77 552±13.7 854±28.3 1.86±0.03 8.95±0.80 309±5.94 1.31±0.01 0.07±0.00 0.06±0.0 0.20±0.0 5m 209±3.22 686±21.5 1180±221 1.86±0.11 20.2±7.51 220±6.85 2.13±0.01 0.21±0.15 <d.l. 0.12±0.02 6m 97±0.93 436±26.5 873±151 2.15±0.02 13.7±2.02 156±1.17 1.15±0.01 0.21±0.05 <d.l. 0.17±0.01 Bark 1m 36±0.22 278±3.95 1565±35.7 0.53±0.01 14.2±0.45 133±1.23 1.94±0.0 0.02±0.00 0.06±0.01 0.03±0 2m 33±0.37 305±4.98 1544±25.2 0.66±0.02 14.1±0.68 149±2.37 1.60±0.0 0.02±0.0 0.05±0.01 0.04±0 3m 33±0.18 287±4.81 1486±35.1 0.71±0.01 14.2±1.42 108±1.13 0.66±0.0 0.01±0.0 0.07±0.01 0.04±0 4m 33±0.37 450±4.62 1553±14.8 1.49±0.01 14.0±0.5 105±1.49 0.87±0.0 0.01±0.0 0.07±0.0 0.08±0 5m 50±0.47 293±6.26 1770±39.2 2.21±0.02 16.4±0.9 93±0.96 1.40±0.0 0.01±0.0 0.08±0.0 0.10±0 6m 78±0.30 155±1.22 742±23.3 0.49±0.0 6.8±0.5 53±0.38 0.78±0.0 0.01±0.0 0.03±0.02 0.02±0 Phloem -5 cm 127.1±0.74 510±5.22 1488±35.1 10.6±0.09 16.33±1.09 545±7.05 19.4±0.02 0.26±0.01 0±0 0.6±0 5 m 71±0.76 344±10.4 580±79.0 0.39±0.04 3.50±2.97 12±0.24 0.09±0.00 0.01±0.01 0.07±0.03 0.02±0 Root -5 cm 91.2±1.09 228±3.40 301±12.9 1.37±0.01 22.3±0.38 63.2±0.24 3.03±0.01 1.50±0.07 0.11±0.0 0.09±0 -15 cm 186±0.61 188±10.4 447±232 2.60±0.08 41.9±8.77 111±0.4 5.57±0.00 4.35±0.07 0.22±0.08 0.17±0.01 <d.l. Tree <d.l. = below the limit of detection

Acknowledgements

We thank Sabah Parks for granting permission to conduct research in Kinabalu Park, and the Sabah Biodiversity Council for research permits. A. van der Ent is the recipient of a Discovery Early Career Researcher Award (DE160100429) from the Australian Research Council. We thank Emile Benizri, Séverine Lopez, Celestino Quintela-Sabaris and Sukaibin Sumail for their support during sampling campaign. We acknowledge IJL and Mrs. Migot for access to TEM facilities. Parts of this research were carried out at SAMBA beamline at SOLEIL, and at XAFS beamline from ELETTRA Synchrotron facility. We would like to thank Emiliano Fonda (SOLEIL), Luca Olivi (ELETTRA) for assistance during the experiments. The research leading to this result has been supported by the project CALIPSOplus under the Grant Agreement 730872 from the EU Framework Programme for Research and Innovation HORIZON 2020. The authors would like to thank Agence Nationale de la Recherche (ANR) project number ANR-14-CE04-0005-03 (AGROMINE) for funding. This is CRPG