1. Case Study: Heavy metal bioavailability in a soil affected by mineral sulphides contamination following the mine spillage at Aznalcóllars (Spain) Clemente et al., Biodegradation, 2003 Aryani Sumoondur Environmental Geosciences, Spring 2005

2. Los Frailes tailings dam failure, Aznalcóllar, Spain (April, 1998)

3. Overview April 1998: 5 million m 3 of an acidic highly toxic pyrite waste spread along the Guadiamar river and 45 km 2 of arable land April 1998: 5 million m 3 of an acidic highly toxic pyrite waste spread along the Guadiamar river and 45 km 2 of arable land solid phase (9 × 105 m 3 ) spread 37 km downstream solid phase (9 × 105 m 3 ) spread 37 km downstream Minerals% Pyrite(FeS 2 ) 83.1 Sphalerite (Zn,Fe)S 5.4 Galena( PbS ) 2.1 Chalcopyrite(CuFeS 2 ) 1.4 Arsenopyrite(FeAsS ) 0.9 Several trace metals N/A Table 1: Composition of Sludge

4. Effect on Soil In some areas, heavy metal levels (esp. Zn, Cd, Cu) still present at phytotoxic levels even though most of the sludge and the topsoil was removed In some areas, heavy metal levels (esp. Zn, Cd, Cu) still present at phytotoxic levels even though most of the sludge and the topsoil was removed Source (Zn, Cd, Cu) : solution phase of spill and solid phase for the other elements Source (Zn, Cd, Cu) : solution phase of spill and solid phase for the other elements Under suitable aeration + moisture conditions, Under suitable aeration + moisture conditions, sulphides are oxidised to H 2 SO 4 (lower pH!) 4FeS 2 + 14H 2 O + 15O 2 → 4Fe(OH) 3 + 8SO 4 2- + 16H +

5. Aim of Study Assess effect of organic amendment and lime (CaO) addition on the bioavailability of heavy metals in soils contaminated by the mine spill Assess effect of organic amendment and lime (CaO) addition on the bioavailability of heavy metals in soils contaminated by the mine spill Factors controlling the solubility and bioavailability of heavy metals Factors controlling the solubility and bioavailability of heavy metals 1) Soil pH2) Redox potential 3) Soil texture 4) Electrical Conductivity 5) Organic matter (OM) content 14 months field experiment where the evolution of soil pH and sulphate formation were monitored in particular 14 months field experiment where the evolution of soil pH and sulphate formation were monitored in particular

6. How to study bioavailabilty? Metal fractions are bioavailable when they are in chemical forms which can be taken up by soil organisms and plants Metal fractions are bioavailable when they are in chemical forms which can be taken up by soil organisms and plants Common method: use a chemical extractant or sequential leaching to predict bioavailability of toxic metals in soils Common method: use a chemical extractant or sequential leaching to predict bioavailability of toxic metals in soils Particular chemical phases of metals in the soil are extracted, which correlate well with amounts of metals taken up by plants grown in the soil Particular chemical phases of metals in the soil are extracted, which correlate well with amounts of metals taken up by plants grown in the soil

7. Methods and Sampling Soil type: non-calcareous, 19.7% clay, 34.3% silt, 46% sand and ~ 1.1% OM Soil type: non-calcareous, 19.7% clay, 34.3% silt, 46% sand and ~ 1.1% OM Treatment :12 plots of 32m 2 Treatment :12 plots of 32m 2 3 plots: cow manure (soluble and easily mineralisable OM) 3 plots: cow manure (soluble and easily mineralisable OM) 3 plots: mature compost with highly humified OM 3 plots: mature compost with highly humified OM rest: control rest: control lime: applied to highly acidic plots lime: applied to highly acidic plots 2 crops of Brassica juncea were grown 2 crops of Brassica juncea were grown 2 organic amendments were added 1 month before each sowing and fertilized 2 organic amendments were added 1 month before each sowing and fertilized After 1st crop, all plots were divided into 2-3 subplots due to the great variation of contamination and pH within plots After 1st crop, all plots were divided into 2-3 subplots due to the great variation of contamination and pH within plots Plots showing excessive soil acidification were limed pH to about 6.0 Plots showing excessive soil acidification were limed pH to about 6.0 0–20 cm deep samples were taken on March, May and Dec 2000 and April 2001 0–20 cm deep samples were taken on March, May and Dec 2000 and April 2001 Samples were air dried and sieved at <2 mm Samples were air dried and sieved at <2 mm

8. Analytical Methods Total metal conc. in plant material and soil were determined following HNO 3 /HClO 4 digestion Total metal conc. in plant material and soil were determined following HNO 3 /HClO 4 digestion Bioavailable metals were analysed after extraction with DTPA-CaCl 2 -triethanolamine Bioavailable metals were analysed after extraction with DTPA-CaCl 2 -triethanolamine Analysis: Atomic Absorption Spectrometry (AAS) Analysis: Atomic Absorption Spectrometry (AAS) Soil pH was measured in a saturated soil paste Soil pH was measured in a saturated soil paste EC was determined in a 1:5 aqueous soil extract EC was determined in a 1:5 aqueous soil extract SO 4 2- content was determined by turbidimetry with BaCl 2 SO 4 2- content was determined by turbidimetry with BaCl 2 Plant growth(fresh and dry weight) were also determined Plant growth(fresh and dry weight) were also determined

9. Results Wide variation in total metal conc. between and within plots Wide variation in total metal conc. between and within plots Zn, Pb and Cu were principal pollutants Zn, Pb and Cu were principal pollutants Removal of sludge was not effective Removal of sludge was not effective

10. pH levels during experiment Mar00: wide range Mar00: wide range May00: lower pH (1st harvest) due to sulphide oxidation May00: lower pH (1st harvest) due to sulphide oxidation Dec00: higher pH values, adequate for plant growth ( liming and dry summer conditions ) Dec00: higher pH values, adequate for plant growth ( liming and dry summer conditions ) April01: low OM and CaCO 3,, limited buffering, soil pH changes drastically April01: low OM and CaCO 3,, limited buffering, soil pH changes drastically

11. [SO 4 2- ]affected pH values of the soil [SO 4 2- ] affected pH values of the soil pH decreased due to sulphide oxidation pH decreased due to sulphide oxidation [SO 4 2- ] [SO 4 2- ] show a close relationship with EC Plots with pH 7 have lowest [SO 4 2- ] Plots with pH 7 have lowest [SO 4 2- ] pH and precipitation of soluble SO 4 2- as CaSO 4 Liming decreased [SO 4 2- ] by increasing pH and precipitation of soluble SO 4 2- as CaSO 4 SO 4 2-, EC and pH

12. In April 2001, sulphate concentrations were at the lowest level With time, the concentration of oxidisable sulphides decreased, which contributed to pH stabilisation OM which is more readily oxidised could also have affected the redox conditions by reducing sulphide oxidation

13. B. juncea survival and biomass production pH < 3.0, plant survival and biomass production is zero pH < 3.0, plant survival and biomass production is zero Addition of organic amendments improved production Addition of organic amendments improved production Type 1st harvest 2nd harvest Cow manure 2869 ± 507 kg ha −1 518 ± 135 kg ha −1 Compost 977 ± 195 kg ha −1 2186±56 3 kg ha −1 Control 1354 ± 470 kg ha −1 1631±94 6 kg ha −1

14. DTPA-extracted heavy metals May 2000 April 2001

15. Behaviour of different heavy metals Zn, Cu, Fe, Mn are in a wide range in all samplings due to the differing total metal concentrations in each plot Zn, Cu, Fe, Mn are in a wide range in all samplings due to the differing total metal concentrations in each plot After 1st harvest, highest values of Zn and Cu were found in zones of very low pH After 1st harvest, highest values of Zn and Cu were found in zones of very low pH After the 2nd harvest, soil conc. of Zn, Fe and Mn decreased, even in zones where pH was low, indicating immobilisation of metals After the 2nd harvest, soil conc. of Zn, Fe and Mn decreased, even in zones where pH was low, indicating immobilisation of metals [Zn],[Mn] were directly correlated with [SO 4 2− ] [Zn],[Mn] were directly correlated with [SO 4 2− ] No correlation for [Fe] and [SO 4 2− ], as Fe forms secondary minerals No correlation for [Fe] and [SO 4 2− ], as Fe forms secondary minerals

16. Behaviour of different heavy metals % Pb extracted as low, (0.8%) although total [Pb] is high % Pb extracted as low, (0.8%) although total [Pb] is high Pb shows inverse relationship with [SO4 2− ] due to formation of insoluble Pb cpds and adsortion on surfaces of Fe-oxides Pb shows inverse relationship with [SO4 2− ] due to formation of insoluble Pb cpds and adsortion on surfaces of Fe-oxides OM generally promoted fixation of heavy metals in non-available soil fractions (Zn decreased from 44.2% to 26.7%) OM generally promoted fixation of heavy metals in non-available soil fractions (Zn decreased from 44.2% to 26.7%) Cu bioavailability did not decrease after second harvest due to formation of stable Cu complexes with soluble OM Cu bioavailability did not decrease after second harvest due to formation of stable Cu complexes with soluble OM

17. Conclusions Soil was highly contaminated by Zn, Cu and Pb, with a wide range of pH Soil was highly contaminated by Zn, Cu and Pb, with a wide range of pH Plant survival, biomass production and heavy metal contents and bioavailability were conditioned by soil pH Plant survival, biomass production and heavy metal contents and bioavailability were conditioned by soil pH Effect of the organic amendments on the bioavailability of metals was difficult to observe (great variability of total metal concentration and pH) but OM improved plant growth Effect of the organic amendments on the bioavailability of metals was difficult to observe (great variability of total metal concentration and pH) but OM improved plant growth Liming successfully controlled soil acidification Liming successfully controlled soil acidification

18. Effect of OM and lime on soil Lime: Raises soil pH Lime: Raises soil pH Humified OM and lime immobilise heavy metals, improving soil quality Humified OM and lime immobilise heavy metals, improving soil quality Soluble OM in fresh manure increases short-term solubility of heavy metals Soluble OM in fresh manure increases short-term solubility of heavy metals However, effect of OM on heavy metal bioavailability in calcareous soils is not related to the OM composition or degree of humification However, effect of OM on heavy metal bioavailability in calcareous soils is not related to the OM composition or degree of humification