Variability of trace element distribution in Noccaea spp., Arabidopsis spp., and Thlaspi arvense leaves: the role of plant species and element accumulation ability.

Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) was applied for the determination of Cd and Zn distributions within the leaves of Cd- and Zn-hyperaccumulating plants, Noccaea caerulescens, N. praecox, and Arabidopsis halleri, in contrast to nonaccumulator species, Thlaspi arvense and A. thaliana. The elemental mapping of the selected leaf area was accomplished via line scans with a 110-µm-diameter laser beam at a 37-µm s-1 scan speed and repetition rate of 10 Hz. The lines were spaced 180 µm apart and ablated at an energy density of 2 J cm-2. The elemental imaging clearly confirmed that Cd was predominantly distributed within the parenchyma of the T. arvense, whereas in the Noccaea spp. and A. halleri, the highest intensity Cd signal was observed in the veins of the leaves. For Zn, higher intensities were observed in the veins for all the plant species except for A. thaliana. Close relationships between Zn and Ca were identified for the Noccaea spp. leaves. These relationships were not confirmed for A. halleri. Significant correlations were also proved between the Cd and Zn distribution in A. halleri, but not for the Noccaea spp. For both T. arvense and A. thaliana, no relevant significant relationship for the interpretation of the results was observed. Thus, the LA-ICP-MS imaging is proved as a relevant technique for the description and understanding of the elements in hyperaccumulating or highly accumulating plant species, although its sensitivity for the natural element contents in nonaccumulator plant species is still insufficient.

Nutritive value and the maximum inclusion level of pennycress meal for broiler chickens.

Two experiments were conducted to evaluate the nutritive value and maximum safe level (MSL) of pennycress meal (PM) for broiler chicks. In experiment 1, a total of 480 chicks was fed either mash or crumbled diets containing zero, 5, 10, or 15% PM for 18 d (8 diets; 6 replications per diet). In experiment 2, a total of 660 chicks was fed mash diets containing zero, 3, 6, 9, 12, or 15% of either PM or canola meal (CM; a comparative reference) for 14 d (11 diets; 6 replications per diet). Analytical results show that PM is a good source of protein (∼31% CP) and it is very comparable to CM (∼36% CP). However, it contains higher erucic acid (∼1.68 vs. < 0.021%), glucosinolates (sinigrin) (∼63.5 vs. <0.163 µmol /g), and crude fiber (18.60 vs. 9.27%) compared to CM. In experiment 1, increasing PM from zero to 15% resulted in linear reductions (P < 0.05) in FI, BWG, and FCR at 10 days. Above 10%, performance responses were affected for FI and BWG at 18 d, respectively. An estimated MSL of 10% PM based on orthogonal contrast was optimal for satisfactory FI and BWG. The MSL as estimated by broken-line linear (BLL) and broken-line quadratic (BLQ) models was 9.12 ± 0.50 and 7.0 ± 1.27%, respectively. In experiment 2, growth performance at 14 d was reduced above 9% due to PM inclusion. CM inclusion did not affect growth performance at 14 d, suggesting 15% to be safe. The MSL for maximum growth performance varied depending on the statistical analysis as follows: 12% by orthogonal contrast and LSD, 15% by the Scheffé test, 10.84 ± 0.57 by BLL, and 8.61 ± 1.29 by BLQ. In conclusion, PM can be included in broiler starter diets as a protein source but its inclusion should be limited to no more than 8.5%. Different statistical procedures give different MSL and this should be considered when interpreting the data.

Towards practical cadmium phytoextraction with Noccaea caerulescens.

A series of field trials were conducted to investigate the potential of Noccaea caerulescens F.K. Mey [syn. Thlaspi caerulescens J &C Presl. (see Koch and Al-Shehbaz 2004)] populations (genotypes) derived from southern France to phytoextract localized Cd/Zn contamination in Thailand. Soil treatments included pH variation and fertilization level and application of fungicide. N. caerulescens populations were transplanted to the field plots three months after germination and harvested in May, prior to the onset of seasonal rains. During this period growth was rapid with shoot biomass ranging from 0.93-2.2 g plant(-1) (280-650 kg ha(-1)) DW. Shoot Cd and Zn concentrations for the four populations evaluated ranged from 460-600 and 2600-2900 mg kg(-1) DW respectively. Cadmium and Zn Translocation Factors (shoot/root) for the populations tested ranged from 0.91-1.0 and 1.7-2.1 and Bioaccumulation Factors ranged from 12-15 and 1.2-1.3. We conclude that optimizing the use of fungicidal sprays, acidic soil pH, planting density and increasing the effective cropping period will increase rates of Cd and Zn removal enough to facilitate practical Cd phytoextraction from rice paddy soils in Thailand.

A thiocyanate-forming protein generates multiple products upon allylglucosinolate breakdown in Thlaspi arvense.

Glucosinolates, amino acid-derived thioglycosides found in plants of the Brassicales order, are one of the best studied classes of plant secondary metabolites. Together with myrosinases and supplementary proteins known as specifier proteins, they form the glucosinolate-myrosinase system that upon tissue damage gives rise to a number of biologically active glucosinolate breakdown products such as isothiocyanates, epithionitriles and organic thiocyanates involved in plant defense. While isothiocyanates are products of the spontaneous rearrangement of the glucosinolate aglycones released by myrosinase, the formation of epithionitriles and organic thiocyanates depends on both myrosinases and specifier proteins. Hydrolysis product profiles of many glucosinolate-containing plant species indicate the presence of specifier proteins, but only few have been identified and characterized biochemically. Here, we report on cDNA cloning, heterologous expression and characterization of TaTFP, a thiocyanate-forming protein (TFP) from Thlaspi arvense L. (Brassicaceae), that is expressed in all plant organs and can be purified in active form after heterologous expression in Escherichia coli. As a special feature, this protein promotes the formation of allylthiocyanate as well as the corresponding epithionitrile upon myrosinase-catalyzed hydrolysis of allylglucosinolate, the major glucosinolate of T. arvense. All other glucosinolates tested are converted to their simple nitriles when hydrolyzed in the presence of TaTFP. Despite its ability to promote allylthiocyanate formation, TaTFP has a higher amino acid sequence similarity to known epithiospecifier proteins (ESPs) than to Lepidium sativum TFP. However, unlike Arabidopsis thaliana ESP, its activity in vitro is not strictly dependent on Fe²âº addition to the assay mixtures. The availability of TaTFP in purified form enables future studies to be aimed at elucidating the structural bases of specifier protein specificities and mechanisms. Furthermore, identification of TaTFP shows that product specificities of specifier proteins can not be predicted based on amino acid sequence similarity and raises interesting questions about specifier protein evolution.

Nicotianamine forms complexes with Zn(II) in vivo.

The non-proteinogenic amino acid nicotianamine (NA) is a major player in plant metal homeostasis. It is known to form complexes with different transition metals in vitro. Available evidence associates NA with translocation of Fe, and possibly other micronutrients, to and between different plant cells and tissues. To date, however, it is still extremely challenging to detect metal-ligand complexes in vivo because tissue disruption immediately changes the chemical environment and thereby the availability of binding partners. In order to overcome this limitation we used various Schizosaccharomyces pombe strains expressing a plant NAS gene to study formation of metal-NA complexes in vivo. Tolerance, accumulation and competition data clearly indicated formation of Zn(ii)-NA but not of Cu(ii)-NA complexes. Zn(ii)-NA was then identified by X-ray absorption spectroscopy (XAS). About half of the cellular Zn was found to be bound by NA in NAS-expressing cells while no NA-like ligands were detected by XAS in control cells not expressing NAS. Given the high conservation of eukaryotic metal homeostasis components, these results strongly suggest the possible existence of Zn(ii)-NA complexes also in planta. Reported observations implicating NA in plant Zn homeostasis would then indeed be attributable to direct interaction of Zn(ii) with NA rather than only indirectly to perturbations in Fe metabolism. Re-evaluation of extended X-ray absorption fine structure (EXAFS) spectra for the Zn hyperaccumulator Thlaspi caerulescens showed that NA is as expected not a major storage ligand for Zn. Instead it is hypothesized to be involved in efficient translocation of Zn to above-ground tissues in hyperaccumulators.

Proteomics of Thlaspi caerulescens accessions and an inter-accession cross segregating for zinc accumulation.

Metal hyperaccumulator plants have previously been characterized by transcriptomics, but reports on other profiling techniques are scarce. Protein profiles of Thlaspi caerulescens accessions La Calamine (LC) and Lellingen (LE) and lines derived from an LCxLE cross were examined here to determine the co-segregation of protein expression with the level of zinc (Zn) hyperaccumulation. Although hydrophobic proteins such as membrane transporters are not disclosed, this approach has the potential to reveal other proteins important for the Zn hyperaccumulation trait. Plants were exposed to metals. Proteins were separated using two-dimensional electrophoresis and those showing differences among accessions, lines or metal exposures were subjected to mass-spectrometric analysis for identification. Crossing decreased the number of different proteins in the lines compared with the parents, more so in the shoots than in the roots, but the frequencies of Zn-responsive proteins were about the same in the accessions and the selection lines. This supports the finding that the Zn accumulation traits are mainly determined by the root and that Zn accumulation itself is not the reason for the co-segregation. This study demonstrates that crossing accessions with contrasting Zn accumulation traits is a potent tool to investigate the mechanisms behind metal hyperaccumulation. Four tentatively identified root proteins showed co-segregation with high or low Zn accumulation: manganese superoxide dismutase, glutathione S-transferase, S-formyl glutathione hydrolase, and translation elongation factor 5A-2. However, these proteins may not be the direct determinants of Zn accumulation. The role of these and other tentatively identified proteins in Zn accumulation and tolerance is discussed.

Decrease of labile Zn and Cd in the rhizosphere of hyperaccumulating Thlaspi caerulescens with time.

By using a rhizobox micro-suction cup technique we studied in-situ mobilization and complexation of Zn and Cd in the rhizosphere of non-hyperaccumulating Thlaspi perfoliatum and two different Thlaspi caerulescens ecotypes, one of them hyperaccumulating Zn, the other Zn and Cd. The dynamic fraction (free metal ions and small labile complexes) of Zn and Cd decreased with time in the rhizosphere solution of the respective hyperaccumulating T. caerulescens ecotypes, and at the end of the experiment, it was significantly smaller than in the other treatments. Furthermore, the rhizosphere solutions of the T. caerulescens ecotypes exhibited a higher UV absorptivity than the solution of the T. perfoliatum rhizosphere and the plant-free soil. Based on our findings we suggest that mobile and labile metal-dissolved soil organic matter complexes play a key role in the rapid replenishment of available metal pools in the rhizosphere of hyperaccumulating T. caerulescens ecotypes, postulated earlier.

Metallothioneins 2 and 3 contribute to the metal-adapted phenotype but are not directly linked to Zn accumulation in the metal hyperaccumulator, Thlaspi caerulescens.

To study the role of metallothioneins (MTs) in Zn accumulation, the expression of TcMT2a, TcMT2b, and TcMT3 was analysed in three accessions and 15 F(3) families of two inter-accession crosses of the Cd/Zn hyperaccumulator Thlaspi caerulescens, with different degrees of Zn accumulation. The highest expression levels were found in the shoots of a superior metal-accumulating calamine accession from St Laurent le Minier, with >10-fold TcMT3 expression compared with another calamine accession and a non-metallicolous accession. Moreover, F(3) sibling lines from the inter-accession crosses that harboured the MT2a or MT3 allele from St Laurent le Minier had higher expression levels. However, there was no co-segregation of TcMT2a or TcMT3 expression and Zn accumulation. To examine the functions of TcMTs in plants, TcMT2a and TcMT3 were ectopically expressed in Arabidopsis. The transformant lines had reduced root length in control medium but not at high metal concentrations, suggesting that the ectopically expressed proteins interfered with the physiological availability of essential metals under limited supply. The Arabidopsis transformant lines did not show increased tolerance to Cd, Cu, or Zn, nor increased Cd or Zn accumulation. Immunohistochemical analysis indicated that in roots, MT2 protein is localized in the epidermis and root hairs of both T. caerulescens and Arabidopsis thaliana. The results suggest that TcMT2a, TcMT2b, and TcMT3 are not primarily involved in Zn accumulation as such. However, the elevated expression levels in the metallicolous accessions suggests that they do contribute to the metal-adapted phenotype, possibly through improving Cu homeostasis at high Zn and Cd body burdens. Alternatively, they might function as hypostatic enhancers of Zn or Cd tolerance.

Feasibility of phytoextraction to remediate cadmium and zinc contaminated soils.

A Cd and Zn contaminated soil was mixed and equilibrated with an uncontaminated, but otherwise similar soil to establish a gradient in soil contamination levels. Growth of Thlaspi caerulescens (Ganges ecotype) significantly decreased the metal concentrations in soil solution. Plant uptake of Cd and Zn exceeded the decrease of the soluble metal concentrations by several orders of magnitude. Hence, desorption of metals must have occurred to maintain the soil solution concentrations. A coupled regression model was developed to describe the transfer of metals from soil to solution and plant shoots. This model was applied to estimate the phytoextraction duration required to decrease the soil Cd concentration from 10 to 0.5 mg kg(-1). A biomass production of 1 and 5 t dm ha(-1) yr(-1) yields a duration of 42 and 11 yr, respectively. Successful phytoextraction operations based on T. caerulescens require an increased biomass production.

Comparison of essential and non-essential element distribution in leaves of the Cd/Zn hyperaccumulator Thlaspi praecox as revealed by micro-PIXE.

A detailed localization of elements in leaf tissues of the field-collected Cd/Zn hyperaccumulator Thlaspi praecox (Brassicaceae) growing at a highly metal-polluted site was determined by micro-proton-induced X-ray emission (micro-PIXE) in order to reveal and compare nutrient and non-essential element accumulation patterns in the case of multiple metal accumulation within particular leaf tissues, including the detailed distribution between apoplast and symplast regions. On the larger scans, the highest concentrations of metals were observed in the epidermis, S and Ca in the palisade mesophyll, Cl in the spongy mesophyll and vascular bundles, and P and K in the vascular bundles. On the more detailed scans, the highest Cd, Pb, Cl and K concentrations were observed in vascular bundle collenchyma. The relative element distribution (%) was calculated based on concentrations of elements in particular leaf tissues and their relative weight portions, indicating that most of the accumulated Zn was located in epidermises, while the majority of Cd and Pb was distributed within the mesophyll. Detailed scans of epidermal/mesophyll tissues revealed that Zn was mainly accumulated and detoxified in the symplast of large vacuolated epidermal cells, Cd in the mesophyll symplast, and Pb in the mesophyll symplast and apoplast.

Spatial distribution of cadmium in leaves of metal hyperaccumulating Thlaspi praecox using micro-PIXE.

* Localization of cadmium (Cd) and other elements was studied in the leaves of the field-collected cadmium/zinc (Cd/Zn) hyperaccumulator Thlaspi praecox from an area polluted with heavy metals near a lead mine and smelter in Slovenia, using micro-PIXE (proton-induced X-ray emission). * The samples were prepared using cryofixation. Quantitative elemental maps and average concentrations in whole-leaf cross-sections and selected tissues were obtained. * Cd was preferentially localized in the lower epidermis (820 microg g(-1) DW), vascular bundles and upper epidermis, whereas about twice the lower concentrations were found in the mesophyll. * Taking into account the large volume of the mesophyll compared with the epidermis, the mesophyll is indicated as a relatively large pool of Cd, possibly involved in Cd detoxification/dilution at the tissue and cellular level.

Identification of nickel chelators in three hyperaccumulating plants: an X-ray spectroscopic study.

We have investigated the accumulation of nickel in a hyperaccumulating plant from the Brassicacae family Leptoplax emarginata (Boiss.) O.E. Schulz. Two supplementary hyperaccumulating plants, which have been the subject of a high number of publications, Alyssum murale Waldst. & Kit and Thlaspi caerulescens J.&C. Presl, and a nonaccumulating species Aurinia saxatilis were also studied for reference. The plants were grown during 4 months in specific rhizoboxes with Ni-bearing minerals as a source of nickel. Nickel speciation was analyzed through X-ray absorption spectroscopy at Ni K-edge (X-ray absorption near edge spectroscopy and extended X-ray absorption fine structure spectroscopy) in the different parts of the plants (leaves, stems and roots) and compared with aqueous solutions containing different organo-Ni(II) complexes. Carboxylic acids (citrate, malate) appeared as the main ligands responsible of nickel transfer within those plants. Citrate was found as the predominant ligand for Ni in stems of Leptoplax and Alyssum, whereas in leaves of the three plants, malate appeared as the chelating organic acid of accumulated metal. Histidine could not be detected either in leaves, stems nor roots of any studied plant sample.

Fast, sensitive, and inexpensive alternative to analytical pigment HPLC: quantification of chlorophylls and carotenoids in crude extracts by fitting with Gauss peak spectra.

Quantification of pigments in complex mixtures is an important task in the physiology of photosynthetic organisms, because pigment composition differs depending on the species, tissue, and physiological state. Currently available methods, however, are either limited to very few pigments (classical UV/vis spectroscopic methods), or they are time-consuming, labor intensive, or costly (e.g., HPLC). Here we describe a UV/vis spectrophotometric method that is capable of a rapid (approximately 1 min/sample) and inexpensive (<1 euro/sample) quantification of more than a dozen pigments in a crude extract, which means it is suitable for high-throughput screening applications. A detection limit of <1 pmol for each pigment allows for determining the pigment composition in only 0.5 microg of lyophilized leaves or algae. The method is based on the description of each pigment spectrum by a series of Gaussian peaks. A sample spectrum is then fitted by a linear combination of these "Gauss peak spectra" including an automatic correction of wavelength inaccuracy, baseline instability, sample turbidity, and effects of temperature/water content. Here we present the Gauss peak spectra from 350 to 750 nm for acetone solutions of all chlorophyll and carotenoid derivatives that are abundant (including conditions of Cd, Cu, or Zn stress) in leaves of higher plants, Euglena, brown algae, and various cyanobacteria like Anabaena and Trichodesmium: [Mg]-Chl a, b, c1, c2; pheophytin a, b; [Cd]-Chl a, b; [Cu]-Chl a, b; [Zn]-Chl a, b; antheraxanthin, aurochrome, beta-carotene, beta-cryptoxanthin, cis- and trans-canthaxanthin, diadinochrome (=diadinoxanthin 5,6-epoxide), cis- and trans-diadinoxanthin, diatoxanthin, cis- and trans-echinenone, fucoxanthin, lutein, myxoxanthophyll, neoxanthin, violaxanthin, and all three stereoisomers of zeaxanthin in acetone. We present extensive tests of our new quantification method for determining optimal and limiting conditions of its performance and for comparison with previous methods. Finally, we show application examples for Thlaspi fendleri (Chlorophyta), Euglena gracilisc (Euglenophyta), Ectocarpus siliculosus (Phaeophyta), and Trichodesmium erythraeum IMS101 (cyanobacteria).

A comparison of phytoremediation capability of selected plant species for given trace elements.

In our experiment, As, Cd, Pb, and Zn remediation possibilities on medium contaminated soil were investigated. Seven plant species with a different trace element accumulation capacity and remediation potential were compared. We found good accumulation capabilities and remediation effectiveness of Salix dasyclados similar to studied hyperaccumulators (Arabidopsis halleri and Thlaspi caerulescens). We have noticed better remediation capability in willow compared to poplar for most of the elements considered in this experiment. On the contrary, poplar species were able to remove a larger portion of Pb as opposed to other species. Nevertheless, the removed volume was very small. The elements found in plant biomass depend substantially on the availability of these elements in the soil. Different element concentrations were determined in natural soil solution and by inorganic salt solution extraction (0.01 molL(-1) CaCl(2)). Extracted content almost exceeded the element concentration in the soil solution. Element concentrations in soil solution were not significantly affected by sampling time.

Thermal treatment of metal-enriched biomass produced from heavy metal phytoextraction.

Phytoextraction is an environmentally sound method for cleaning up sites that are contaminated with toxic heavy metals. However, the method has been questioned because it produces a biomass-rich secondary waste containing the extracted metals. Therefore, further treatment of this biomass is necessary. In this study, we investigated whether thermal treatment could be a feasible option for evaporatively separating metals from the plant residues. We used a laboratory scale reactor designed to simulate the volatilization behavior of heavy metals in a grate furnace. The evaporation of alkali and heavy metals from plant samples was investigated online, using a thermo-desorption spectrometer (TDS). Experiments were performed in the temperature range of 25-950 degrees C with leaves of the Cd and Zn hyperaccumulator Thlaspi caerulescens and of the high biomass plant Salix viminalis (willow), both grown on contaminated soils. Gasification (i.e., pyrolysis), which occurs under reducing conditions, was a better method than incineration under oxidizing conditions to increase volatilization and, hence subsequently recovery, of Cd and Zn from plants. It would also allow the recycling of the bottom ash as fertilizer. Thus, our investigations confirmed that incineration (or co-incineration) is a viable option for the treatment of the heavy metal-enriched plants.

Identification of the form of Cd in the leaves of a superior Cd-accumulating ecotype of Thlaspi caerulescens using 113Cd-NMR.

Thlaspi caerulescens (Ganges ecotype) is a known Cd hyperaccumulator, however, the ligands which coordinate to Cd ions in the leaves have not been identified. In the present study, the chemical form of Cd was investigated by using 113Cd-nuclear magnetic resonance (NMR) spectroscopy. Plants were grown hydroponically with a highly enriched 113Cd stable isotope. Measurements of 113Cd-NMR with intact leaves showed a signal at the chemical shift of around -16 ppm. Crude leaf sap also gave a similar chemical shift. Purification by gel filtration (Sephadex G-10), followed by cationic and anionic exchange chromatography, showed that Cd occurred only in the anionic fraction, which gave the same chemical shift as intact leaves. Further purification of the anionic fraction, combined with 113Cd- and 1H-NMR studies, revealed that only the fraction containing malate showed a chemical shift similar to the intact leaves. These results indicate that Cd was coordinated mainly with malate in the leaves of T. caerulescens. The malate concentration in the leaves was not affected by increasing Cd concentration in the solution, suggesting that malate synthesis is not induced by Cd. Because the Cd-malate complex is relatively weak, we suggest that the complex forms inside the vacuoles as a result of an efficient tonoplast transport of Cd and a constitutively high concentration of malate in the vacuoles, and that the formation of the Cd-malate complex may lead to a decrease of subsequent Cd efflux to the cytoplasm.

Biofumigant compounds released by field pennycress (Thlaspi arvense) seedmeal.

Defatted field pennycress (Thlaspi arvense L.) seedmeal was found to completely inhibit seedling germination/emergence when added to a sandy loam soil containing wheat (Triticum aestivum L.) and arugula [Eruca vesicaria (L.) Cav. subsp. sativa (Mill.) Thell.] seeds at levels of 1.0% w/w or higher. Covering the pots with Petri dishes containing the soil-seedmeal mixture decreased germination of both species at the lowest application rate (0.5% w/w), suggesting that the some of the phytotoxins were volatile. CH2Cl2, MeOH, and water extracts of the wetted seedmeal were bioassayed against wheat and sicklepod (Senna obtusifolia (L.) H. S. Irwin & Barneby) radicle elongation. Only the CH2Cl2 extract was strongly inhibitory to both species. Fractionation of the CH2Cl2 extract yielded two major phytotoxins, identified by gas chromatography-mass spectrometry and NMR as 2-propen-1-yl (allyl) isothiocyanate (AITC) and allyl thiocyanate (ATC), which constituted 80.9 and 18.8%, respectively, of the active fraction. When seeds of wheat, arugula and sicklepod were exposed to volatilized AITC and ATC, germination of all three species was completely inhibited by both compounds at concentrations of 5 ppm or less. In field studies, where seedmeal was applied at 0.50, 1.25, and 2.50 kg/m2 and tarped with black plastic mulch, all of the treatments significantly reduced dry weight of bioassay plants compared to the tarped control, with the highest seedmeal rate decreasing dry matter to less than 10% of the control 30 d after seedmeal application. Field pennycress seedmeal appears to offer excellent potential as a biofumigant for high-value horticultural crops for both conventional and organic growers.

Thlaspi caerulescens on nonmetalliferous soil in Luxembourg: ecological niche and genetic variation in mineral element composition.

Forty-seven populations of Thlaspi caerulescens in Luxembourg were characterised for population size, soil mineral element composition and other habitat characteristics. Foliar concentrations of eight elements were assessed in 15 populations in the field and in eight populations cultivated in zinc (Zn)-cadmium (Cd)-nickel (Ni)-enriched soil. T. caerulescens favoured stony soil developed on steep, south-facing Emsian shale outcrops. All soil samples were nonmetalliferous. Soil pH ranged from 4.2 to 6.9. Field-growing plants had very high concentrations of heavy metals in the leaves (Zn, 3000-13 000 mg kg(-1); Cd, 11-44 mg kg(-1); Ni, 38-473 mg kg(-1)). Positive soil-plant correlations existed for Zn and Mn. In cultivation, significant genetic variation was found for biomass and six of eight mineral elements. For Cd and Zn, variation range among 48 half-sib families was two-fold (Cd, 183-334 mg kg(-1); Zn, 8030-16 295 mg kg(-1)). Most of the variation occurred among populations, consistent with the selfing mating system of those populations. There was a tight Zn-Cd genetic correlation (r = +0.83, P < 0.0001). The significance of the results to the conservation of T. caerulescens in Luxembourg is briefly discussed.

Zn, Cd and Pb accumulation and arbuscular mycorrhizal colonisation of pennycress Thlaspi praecox Wulf. (Brassicaceae) from the vicinity of a lead mine and smelter in Slovenia.

Significant hyperaccumulation of Zn, Cd and Pb in field samples of Thlaspi praecox Wulf. collected from a heavy metal polluted area in Slovenia was found, with maximal shoot concentrations of 14,590 mg kg(-1) Zn, 5960 mg kg(-1) Cd and 3500 mg kg(-1) Pb. Shoot/root ratios of 9.6 for Zn and 5.6 for Cd show that the metals were preferentially transported to the shoots. Shoot bioaccumulation factors exceeded total soil Cd levels 75-fold and total soil Zn levels 20-fold, further supporting the hyperaccumulation of Cd and Zn. Eighty percent of Pb was retained in roots, thus indicating exclusion as a tolerance strategy for Pb. Low level colonisation with arbuscular mycorrhizal fungi (AMF) of a Paris type was observed at the polluted site, whereas at the non-polluted site Arum type colonisation was more common. To our knowledge this is the first report of Cd hyperaccumulation and AMF colonisation in metal hyperaccumulating T. praecox.

Phytoalexins from Thlaspi arvense, a wild crucifer resistant to virulent Leptosphaeria maculans: structures, syntheses and antifungal activity.

Phytoalexins are inducible chemical defenses produced by plants in response to diverse forms of stress, including microbial attack. Our search for phytoalexins from cruciferous plants resistant to economically important fungal diseases led us to examine stinkweed or pennycress (Thlaspi arvense), a potential source of disease resistance to blackleg. We have investigated phytoalexin production in leaves of T. arvense under abiotic (copper chloride) and biotic elicitation by Leptosphaeria maculans (Desm.) Ces. et de Not. [asexual stage Phoma lingam (Tode ex Fr.) Desm.], and report here two phytoalexins, wasalexin A and arvelexin (4-methoxyindolyl-3-acetonitrile), their syntheses and antifungal activity against isolates of P. lingam/L. maculans, as well as the isolation of isovitexin, a constitutive glycosyl flavonoid of stinkweed, having antioxidant properties but devoid of antifungal activity.