1155 Environmental Toxicology and Chemistry, Vol. 22, No. 5, pp. 1155–1161, 2003  2003 SETAC Printed in the USA 0730-7268/03 $12.00 + .00 ACCUMULATION OF CADMIUM BY DURUM WHEAT ROOTS: BASES FOR CITRATE-MEDIATED EXCEPTIONS TO THE FREE ION MODEL EDWARD BERKELAAR and BEVERLEY A. HALE* Department of Land Resource Science, University of Guelph, Guelph, Ontario N1G 2W1, Canada ( Received 7 December 2001; Accepted 31 October 2002) Abstract—The accumulation of Cd in durum wheat (Triticum turgidum) roots from hydroponic solutions, with the proportion of total Cd (8.9–445 nM Cd) as Cd 2+ varied by the addition of citrate, was determined to test the free-ion model (FIM) of metal bioavailability for higher plants. Calcium, Mg, and K were also varied. Citrate enhanced root-Cd accumulation at higher Cd 2+ concentrations but not lower relative to the same Cd 2+ concentrations in solutions containing 0 mM citrate. Elevating Ca 2+ and Mg 2+ concentrations in the citrate solution to the same as those in control solutions alleviated some of the citrate-mediated enhancement but not all. Solutions containing 66% less Ca or Mg than control but the same Cd 2+ concentration and no citrate also resulted in increased root Cd. Elevated K + did not influence Cd accumulation. Regression relationships between root-Cd accumulation and total Cd in solution were similar for the control and pooled amended solutions, whereas they were different for root-Cd accumulation and solution Cd 2+ . These results contribute to the growing body of evidence that the FIM alone is likely insufficient to predict plant accumulation of metals from soils, although it may be a useful probe for the mechanistic bases of metal bioavailability. Keywords—Bioavailability Cadmium Citrate Durum wheat Speciation INTRODUCTION The free-ion model (FIM) suggests that for both aquatic organisms and higher plants, accumulation and/or toxicity of dissolved metals such as Cd correlates better with the con- centration of the free ion (Cd 2+ ) than with the total concen- tration of the dissolved metal in the response medium [1,2]. The FIM in its simplest form is cautioned to apply only when the effect or accumulation of the metal is proportional to the extent of occupancy of cell surface binding sites by the free ion and not by a complexed form, when no other metals exist in the exposure solution that interact with dissolved ligands or with cell surface binding sites, and when the rate-limiting step in the process is metal transport across the biological barrier (diffusion of the metal to the cell surface and binding to this surface are not rate limiting). The first assumption is consistent with the physiology of ion uptake by plants: In the case of cations, primary capture by the negative charges on root cells is followed by active or passive transport of the cation across plasma membrane. Thus, FIM does not allow for the uptake of a metal-ligand complex. The second assumption means that FIM in its simplest form will not apply to most biological situations, as soil solutions and surface waters con- tain some concentrations of Ca 2+ and Mg 2+ ; thus, the second assumption is never met. The third assumption can be a lim- itation to the application of FIM to plant roots, as for some elements supply of the element to the root surface by diffusion can be the rate-limiting step to uptake. Additional to that limitation to the use of FIM in biological situations, some recent studies have indicated that the presence of hydrophilic, low-molecular-weight compounds [3] may lead to exceptions to FIM. Increasing the concentration of Cl in the exposure solution resulted in enhanced accumulation of Cd (at similar solution Cd 2+ concentrations) by Swiss chard * To whom correspondence may be addressed (bhale@uoguelph.ca). [4,5]. More Cl - in solution resulted in a greater concentration of CdCl n 2-n species, and the authors suggested that Cd accu- mulation was increased because of accumulation of these spe- cies or because of enhanced diffusion of Cd 2+ to the uptake sites. In a study of SO 4 in solution and accumulation of Cd, plant tissue Cd concentrations were unaffected by increasing solution SO 4 concentrations, even though the concentration of Cd 2+ in solution was reduced significantly; the authors con- cluded that CdSO 4 0 (aq) was taken up as readily as Cd 2+ or that diffusion of Cd to the root surface was enhanced [6]. Addition of ethylenediaminetetraacetic acid (EDTA) to a solution con- taining Cd significantly reduced the Cd 2+ concentration in so- lution and also reduced the accumulation of Cd by duckweeds (Lemnaceae) [7]. However, the reduction in accumulation was not as great as predicted by the reduction in solution Cd 2+ concentration, and the authors attributed this discrepancy to uptake of CdEDTA species through breaks in the root endo- dermis or to CdEDTA as a buffer of Cd 2+ concentration during treatment, resulting in enhanced supply of Cd to the root sur- face. Evidence for accumulation of Zn-EDTA complexes in xylem sap arises from a study where Zn-EDTA complexes were measured in several species; its concentration correlated with total Zn in shoot tissue and varied among plant species [8]. A recent study with unicellular algae has demonstrated that the toxicity of Cd and Zn is not solely dependent on their free-ion (Cd 2+ and Zn 2+ ) concentrations. The copresence of citrate (a low-molecular-weight metabolite) resulted in greater Cd and Zn toxicities than predicted for similar free-ion activ- ities [9]. Citrate was also accumulated by the algae, leading the authors to conclude that the accidental transport of a Cd- citrate complex by the citrate transporter could account for the enhanced toxicity in the presence of citrate. Malonate and malate alleviated some Cu toxicity to wheat root elongation, although not to the extent predicted by FIM [10]. No such alleviation of toxicity was observed for Cu and citrate or for any of these ligands and Zn.