In response to new European Union regulations, studies are underway to mitigate accumulation of toxic cadmium (Cd) in cacao (Theobroma cacao, Tc). This study advances such research with Cd isotope analyses of 19 genetically diverse cacao clones and yeast transformed to express cacao natural resistance-associated macrophage protein (NRAMP5) and heavy metal ATPases (HMAs). The plants were enriched in light Cd isotopes relative to the hydroponic solution with Δ 114/110 Cd tot-sol = −0.22 ± 0.08‰. Leaves show a systematic enrichment of isotopically heavy Cd relative to total plants, in accord with closed-system isotope fractionation of Δ 114/110 Cd seq-mob = −0.13‰, by sequestering isotopically light Cd in roots/stems and mobilisation of remaining Cd to leaves. The findings demonstrate that (i) transfer of Cd between roots and leaves is primarily unidirectional; (ii) different clones utilise similar pathways for Cd sequestration, which differ from those of other studied plants; (iii) clones differ in their efficiency of Cd sequestration. Transgenic yeast that expresses TcNRAMP5 (T. cacao natural resistance-associated macrophage gene) had isotopically lighter Cd than did cacao. This suggests that NRAMP5 transporters constitute an important pathway for uptake of Cd by cacao. Cd isotope signatures of transgenic yeast expressing HMA-family proteins suggest that they may contribute to Cd sequestration. The data are the first to record isotope fractionation induced by transporter proteins in vivo.

We have prepared a large volume of pure, concentrated and homogenous zinc standard solution. This new standard solution is intended to be used as a primary reference standard for the zinc isotope community, and to serve as a replacement for the nearly exhausted current reference standard, the so-called JMC-Lyon Zn. The isotopic composition of this new zinc standard (AA-ETH Zn) has been determined through an inter-laboratory calibration exercise, calibrated against the existing JMC-Lyon standard, as well as the certified Zn reference standard IRMM-3702. The data show that the new standard is isotopically indistinguishable from the IRMM-3702 zinc standard, with a weighted δ66/64Zn value of 0.28 ± 0.02‰ relative to JMC-Lyon. We suggest that this new standard be assigned a δ66/64Zn value of +0.28‰ for reporting of future Zn isotope data, with the rationale that all existing published Zn isotope data are presented relative to the JMC-Lyon standard. Therefore our proposed presentation allows for a direct comparison with all previously published data, and that are directly traceable to a certified reference standard, IRMM-3702 Zn. This standard will be made freely available to all interested labs through contact with the corresponding author

This work critically reviews stable isotope fractionation of essential (B, Mg, K, Ca, Fe, Ni, Cu, Zn, Mo), beneficial (Si), and non-essential (Cd, Tl) metals and metalloids in plants. The review (i) provides basic principles and methodologies for non-traditional isotope analyses, (ii) compiles isotope fractionation for uptake and translocation for each element and connects them to physiological processes, and (iii) interlinks knowledge from different elements to identify common and contrasting drivers of isotope fractionation. Different biological and physico-chemical processes drive isotope fractionation in plants. During uptake, Ca and Mg fractionate through root apoplast adsorption, Si through diffusion during membrane passage, Fe and Cu through reduction prior to membrane transport in strategy I plants, and Zn, Cu, and Cd through membrane transport. During translocation and utilization, isotopes fractionate through precipitation into insoluble forms, such as phytoliths (Si) or oxalate (Ca), structural binding to cell walls (Ca), and membrane transport and binding to soluble organic ligands (Zn, Cd). These processes can lead to similar (Cu, Fe) and opposing (Ca vs. Mg, Zn vs. Cd) isotope fractionation patterns of chemically similar elements in plants. Isotope fractionation in plants is influenced by biotic factors, such as phenological stages and plant genetics, as well as abiotic factors. Different nutrient supply induced shifts in isotope fractionation patterns for Mg, Cu, and Zn, suggesting that isotope process tracing can be used as a tool to detect and quantify different uptake pathways in response to abiotic stresses. However, the interpretation of isotope fractionation in plants is challenging because many isotope fractionation factors associated with specific processes are unknown and experiments are often exploratory. To overcome these limitations, fundamental geochemical research should expand the database of isotope fractionation factors and disentangle kinetic and equilibrium fractionation. In addition, plant growth studies should further shift toward hypothesis-driven experiments, for example, by integrating contrasting nutrient supplies, using established model plants, genetic approaches, and by combining isotope analyses with complementary speciation techniques. To fully exploit the potential of isotope process tracing in plants, the interdisciplinary expertise of plant and isotope geochemical scientists is required.

Cd isotope composition in cacao seems to be cultivar-specific whereas Cd in soil is probably due to tree litter recycling.

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