1. Comparison of Toxic Effects of Chromium III and VI on Tomato Plants R. Silva 1, R. Quaresma 1, R. P. Coelho 1, A. R. Costa 1, L. L. Martins 2, M. P. Mourato 2, F. S. Henriques 3 Introduction (1) Departamento de Qu í mica Agr í cola e Ambiental, ISA, Tapada da Ajuda, 1349-017 Lisboa, Portugal (2) Instituto de Ciências Agr á rias Mediterrânicas, Universidade de É vora Apartado 94 7002 - 554 É vora, Portugal (3) Plant Biology Unit, Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa), 2829 Caparica, Portugal Materials and methods Results and Discussion References Chromium (Cr) is a relatively abundant element with a high potential for plant toxicity. A survey is currently underway in Portugal to evaluate the extent of Cr pollution resulting from mining activities, and this prompted our study. Cr is most frequently present in soils in either its trivalent or hexavalent forms, depending on the soil´s pH. The element´s availability for plants depends on a number of interactions with other soil constituents, mainly with Mn and Fe-oxides/hydroxides. Although Cr is known to be essential for animals and micro-organisms, apparently it plays no beneficial role in plant metabolism; on the contrary, elevated Cr concentrations are toxic to cultivated species. For a same concentration, Cr VI has been found to be more toxic than Cr III; the main reasons for this difference are still being evaluated, but the high solubility and oxidative capacity of Cr VI as well as its ability to interfere with essential nutrients´ uptake and distribution by the plant are certainly major contributing factors. In this work, we examine the effects of Cr III and VI on the plant water relations and photosynthetic metabolism as well as on the plant nutritional status (1). Plant material: One-week old tomato (Lycopersicon esculentum Mill. cv Juncal) seedlings were grown for two weeks in hydroponics (full-strength Hoagland’s solution) and then subjected to Cr III or Cr VI concentrations of 10, 20, 30, 40 and 50 μM. Experiments were performed in duplicate using 7 plants, in a total of 14 plants for each treatment (1) A.Zayed and N.Terry. Plant and Soil 249 (2003) 139-156 (2) R.J.Porra et al.. BBA 975 (1984) 384-389 (3) P.Vajpyee et al.. Chemosphere 41 (2000) 1075-1082 Analytical determinations: Measurements of studied parameters were carried out one week later, as follows:  chlorophyll concentration was measured spectrophotometrically using the equations of Porra et al (2) based on the measurement with Hansatech portable chlorophyll meter;  water potential was determined using a Scholander´s bomb;  net photosynthetic rates and stomatal conductance were measured with an open flow-through portable system (LCi, Hansatech);  fluorescence parameters were recorded with a portable fluorometer (PEA, Hansatech)  Mineral content were obtained after sample microwave digestion, and analysed by flame atomic absorption spectrophotometer (Unicam Solaar M) for As, Ca, Mg, Mn, Fe, Cu, Zn, Na. Phosphorus was determined by the molybdovanadate colorimetric method (using an Hitachi U-2000 UV/Vis Spectrophotometer); these analytical determinations were performed in triplicate Total dry weight loss caused by Cr III was relatively small: only a 30% loss relative to the control was measured for the 50μM treatment. On the contrary, Cr VI caused a large decrease in the plants´ dry weight that remained roughly constant at 70% of the control from the 20 μM treatment upwards. However, there appeared to be some dry weight recovery for the plants grown in 50 μM Cr VI solution, which was found to correlate positively with other measured parameters. The shoot-root weight ratio was largely decreased in the Cr VI treatments above 20 μM, again showing slight recovery at the 50 μM treatment. Plant dry weight: Cr symptoms: Cr VI at 20 μM concentrations and above caused marked leaf chlorosis and plant stunting, the shoot part being more strongly affected than the root. Cr III failed to induce visible toxic symptoms on the leaf, even at the highest concentration tested, although slight overall stunting could be detected. Cr III 40μMCr VI 20 μM Control Chlorophyll content: Dry weight (g) [Cr] (  M) shoot-root weight ratio [Cr] (  M) Leaf chlorophyll content was drastically reduced at the Cr VI 20 μM concentration and above, thus explaining the extensive leaf chlorosis caused by this Cr form; the 50 μM Cr VI treatment exhibits a visible recovery, as we found for dry weight and shoot-root weight ratio. Cr III also caused a significant reduction in leaf chlorophyll content but only for the two highest concentrations tested. Cr is a known inhibitor of ALA de-hidratase (3), thus preventing leaf chlorophyll formation. Chlorophyll (r.u.) [Cr] (  M) No significant differences on the plants´ relative water content were found between controls and the Cr III/Cr VI treatments. Water potential variations showed no trend that could be correlated with Cr concentrations used in these experiments. Water relations: [Cr] (  M) RWC (%) Cr VI caused marked reductions in stomatal conductance of increasing magnitude from the 20μM treatment onwards, whereas the Cr III caused only small reductions at the three highest treatments. Cr VI also significantly reduced net photosynthetic rates in a manner parallel to its effect in stomatal conductance, demonstrating the direct correlation between the two. Additionally, there must be an effect resulting from the decreased leaf chlorophyll content of plants treated with Cr VI. Fluorometric measurements showed only minor effects of either Cr III or VI on PS II function, in spite of major re-organisation of chlorophyll arrangements around this photosystem. Stomatal conductance and photosynthetic rate: From the data gathered, it is concluded that Cr VI is more toxic than Cr III for plants, thus confirming previous observations. The 20μM Cr VI concentration represents a threshold beyond which the harmful effects of this element becime drastic and compromise the plant´s viability. It is also concluded that the marked effect of Cr VI on net photosynthesis is mediated mostly by stomatal closure and only secondarily by PS II dysfunction. Cr interactions: [Cr] (  M) IRGA A (mmol CO 2 m -2 s -1 ) stomatal conductance (gs) 13h [Cr] (  M) Both Cr III and Cr VI accumulate preferentially in the roots, only a minor fraction being transferred to the shoots. However, Cr VI shows a much higher shoot-root ratio than Cr III, which partially accounts for its predominant effects on the shoot part of the plant. In the roots, Cr VI significantly decreases Mn and Ca concentrations whereas Cr III decreases that of Mg. In the shoots, Cr VI decreases P, Mn and Zn and Cr III decreases P and Mn. Conclusions: