The occurrence of Itai-Itai disease in Japan, the long half-life of Cd in humans, and evidence that Cd causes kidney damage and other medical conditions in humans are reasons why contamination of soils and the wider environment by Cd has attracted international attention. An additional reason for concern arises because concentrations of Cd in food associated with undesirable accumulations in human tissues, particularly the kidneys, rarely affects the growth and appearance of food plants and animals. Standards that place limits on permitted concentrations (maximum levels) of Cd in the edible portions of various foods have helped in efforts to manage for Cd minimisation (Rayment 1991; Abbott 2007; Warne et al. 2007).
Cadmium is a rare Group IIB heavy metal (67th element in order of abundance). In common with Hg and Zn, Cd readily loses two electrons to form dipositive ions (Aylett 1971), its predominant oxidation state in nature (WHO 1985). Its inorganic chemistry (not its biochemistry) follows closely that of Zn (Fergusson 1990), except that unlike Zn, Cd is insoluble in alkalis (Aylett 1971; 1979).
Within soils, Cd is extensively associated with colloidal and particulate matter, with speciation in oxic-soil solutions confined predominantly to (in decreasing order) free Cd2+, CdSO40 and CdCl1+. In oxic, alkaline soils the predominant water-soluble species are Cd2+, CdCl1+, CdSO40 and CdHCO31+.
The proportion of organically-bound Cd in soil solution, even following sewage sludge addition, is relatively small (Alloway 1990). Moreover, the amounts of electrically neutral CdSO40 and CdCl20 species increase beyond pH 6.5. Adsorption processes rather than precipitation reactions control the distribution of Cd between soil-bound and soluble forms at the concentrations normally encountered in soils, including polluted soils (Alloway 1990). The distribution diagram for Cd/oxy/hydroxide species indicates that most free Cd2+ disappears from aqueous solutions by pH 10.
It is known that the uptake of Cd by vegetables such as Swiss chard and potatoes (Bingham et al. 1983; McLaughlin et al. 1993, 1997) is enhanced by the presence of elevated concentrations of soil and water Cl–. Also, a wide array of soil tests for Cd have emerged, often to provide guidance on the phyto-availability of soil Cd (e.g. Clayton and Tiller 1979; Sposito et al. 1982; Brams and Anthony 1988). The task is challenging, because in addition to Cl–, soil properties such as pH, plus the extent and location of Cd additions from impurities in phosphatic fertilisers, sewage sludge, industrial wastes, etc all influence plant Cd uptake.
European specialists report a preference for CaCl2 as the extractant for routine use in assessing plant-available Cd (and Zn) in soil (Salt and Kloke 1986), although soil/solution ratios, extracting times, and concentrations of CaCl2 vary, the latter commonly from 0.01 M to 0.1 M. Theoretical, practical and analytical reasons account for the various concentrations of CaCl2 selected (e.g. Sauerbeck and Styperek 1985, Novozamsky et al. 1993). Others to report the successful use of dilute solutions of CaCl2, as indicators of Cd phyto-bioavailability across soil types, include Morgan and Alloway (1984), König (1986) and Alloway et al. (1990) for vegetables, and Whitten and Richie (1991) for subterranean clover tops.
This method is based on that of Sauerbeck and Styperek (1985) and Smilde et al (1992). It involves extraction of air-dry soil with 0.1 M CaC12 for 2 h at a soil/solution ratio of 1:2.5, with subsequent instrumental analysis for Cd by ICP-MS (preferred) or graphite furnace, or by ICPAES when solution concentrations are expected to exceed ≈0.05 mg Cd/L. Analytical attributes of the method include: (i) it extracts similar forms but higher amounts of Cd than more-dilute solutions of CaCl2 under otherwise similar conditions, simplifying the subsequent analytical measurement of Cd; (ii) higher amounts of Cd in soil extracts minimise the consequences of accidental contamination in the laboratory, and (iii) it has been shown to be a useful predictor of Cd concentrations in edible plant tissues in Europe (Sauerbeck and Styperek 1985) and in commercially grown vegetables from Queensland (Rayment 1994). Expect concentrations (mg Cd/kg air-dry) in ‘normal’ horticultural soils to average around 0.032 ± 0.028 at 0–100 mm and 0.019 ± 0.016 at 200–300 mm (n ≈190; Rayment 1994), while values of at least 2.5 mg Cd/kg have been obtained from contaminated soils (Sauerbeck and Styperek 1985). Total and extractable levels of Cd commonly decline with increasing soil depth (e.g. Williams and David 1973; Pierce et al. 1982; Rayment et al. 1997).
0.1 M CaCl2 Extracting Solution
Dissolve 14.70 g calcium chloride (CaCl2.2H2O) and make to 1.0 L with high-purity water (DDW). Store in a clean (Cd-free), covered plastic container.
2% Nitric Acid
Dilute 20 mL of nitric acid (HNO3, 15 mol/L; high quality or distilled) to 1.0 L with DDW and mix well. Store in a clean (Cd-free), covered plastic container.
Cadmium Primary Standard
1 L contains 100 mg of Cd.
Dilute a certified, multi-element standard containing Cd (and often other metals) or dissolve 0.2282 g high grade cadmium sulfate (3CdSO4.8H2O) and make volume to 1.0 L with DDW and 2% Nitric Acid in the proportion 1+4 in an acid-cleaned and rinsed borosilicate volumetric flask. Store in a clean polypropylene reagent bottle.
Cadmium Secondary Standard
1 L contains 2 mg of Cd.
Dilute 10.0 mL Cd Primary Standard, while swirling/stirring, to 500 mL with 0.1 M CaC12 Extracting Solution–2% Nitric Acid in the proportion 1+4. The borosilicate volumetric flasks should be acid-cleaned and rinsed before use. This solution is best prepared each time working standards are required but may be stored for up to 2 months if necessary.
Cadmium Working Standards
Using a combination of a calibrated digital burette and a calibrated burette, add 0.2, 0.4, 2.0, 3.0, 4.0, 10.0, 20.0, 40.0, 60.0, 80.0 and 100.0 mL Cd Secondary Standard to separate 200 mL volumetric flasks and make to volume with 0.1M CaCl2 Extracting Solution–2% Nitric Acid in the proportion 1+4. The borosilicate volumetric flasks should be acid-cleaned and rinsed before use. These working standards contain Cd concentrations of 0.002, 0.004, 0.02, 0.04, 0.10, 0.20 ... 0.8 and 1.0 mg Cd/L. For a 1:2.5 soil/extract ratio, these correspond to soil concentrations of 0.005, 0.01, 0.05, 0.10, 0.25, 0.5 ... 2.0 and 2.5 mg Cd/kg. Store in clean polypropylene reagent bottles and use within one month.
10 μg/L Mix v Tuning Solution (for ICP-MS analysis)
Weigh 0.500 g of multi-element Analytika standard (or equivalent) (10 mg/L Ba, Be, Bi, Ce, Co, In, Li, Ni, Pb and U) into a clean 500 mL polypropylene bottle, add 10 mL HNO3, (15 mol/L; high quality) while mixing, and then make to 500.0 g with DDW.
Tellurium Internal Standard (for ICP-MS analysis)
Initially prepare a Te Solution containing 10 mg Te/L by pipetting 10 mL of a commercially obtained 1000 mg Te/L Standard. Add 20 mL distilled HNO3 (15 mol/L), mix well then make to 1.0 L with DDW. This solution may be stored for up to 12 months. Next weigh into a clean 500 mL polypropylene bottle 3.50 g of this 10 mg Te/L Solution, add 10 mL of HNO3 (15 mol/L; high quality), mix well and make to 500 g with DDW. This solution may be stored for up to 2 months.
Matrix modifier (1.2% w/v ammonium dihydrogen orthophosphate) (for Graphite Furnace AAS) Dissolve 1.2 g of NH4H2PO4 in water and dilute to 100 mL.
The sieves, other apparatus, glassware and filter papers used for soil preparation, soil extractions, filtration and instrumental analysis should be acid washed (e.g. 10% v/v HNO3/DDW) and rinsed with DDW before use. Occasional tests should also be made to ensure all sample bags, equipment and the DDW contain no measurable Cd contamination. See Notes 1 and 2 for more details.
Dry soil samples in a clean, dust-free environment, ideally in a Class 1000 Clean Room or within a Class 100 Laminar Flow Hood. When air-dry (40–45°C max.), crush/mill the soil to <2 mm in a stainless steel mill and store in sealed, uncontaminated containers.
Subsequently, weigh 20.0 g of air-dry soil into clean, dry, 100 mL, wide-neck polyethylene flasks, then add 50 mL 0.1 M CaCl2 Extracting Solution to each, stopper, then mechanically shake end-over-end for 2.0 h at around 22–25°C. After shaking, allow solutions to stand for ≈15 min. Centrifuge in 10 mL centrifuge tubes (subsequently remove any floating material by vacuum tube) or filter (e.g. Whatman No. 542) prior to instrumental analysis by ICP-MS (preferred) or GFAAS. Only use ICPAES if elevated concentrations are expected. Refer to Note 3 for guidance on how to store and how long the extracts can be held before instrumental analysis. Follow manufacturer’s recommendations with respect to instrument parameters. A reagent blank should also be measured and adjustments made as necessary. Also, dilute any over-range samples with extracting solution.
For determination of Cd by ICP-MS, initially check for instrument response/performance, using the 10 μg/L Mix v Tuning Solution. All instrument performance criteria should be confirmed at this point. Next dilute 2 mL of each clarified soil extract solution with 8 mL of 2% HNO3 into a 10 mL polyethylene centrifuge tube. Cap and shake and make allowance for this dilution when calculating results. Mix the Tellurium Internal Standard (this is to compensate for drift and matrix effects) with each standard and diluted sample extract in the proportion 0.42 mL/min of Te Internal Standard to 0.6 mL/min of diluted sample extract during operation of the ICP-MS. Also, check a reference solution every 10 samples and recalibrate if drift exceeds 10%. If analyte concentration exceeds the standard calibration range, dilute the clarified soil extracts with calibration blank and repeat relevant determinations. The optimum batch size (samples, including checks) is ≈48–50.
For ICPAES, noting that this technology lacks sensitivity at low Cd concentrations, the preferred wave length is either 226.5 nm or 361.05 nm, with no interference to Cd from Fe and Al expected. For GFAAS, the preferred wave length is 228.8 nm. Also for a Perkin ElmerTM 4100ZL GFAAS, use a sample volume of 20 μL (normal) or 10 μL or 5 μL (if calibration limit is exceeded) together with a Matrix Modifier (5 μL of 1.2% w/v ammonium dihydrogen orthophosphate).
Determine Cd concentrations of sample extracts directly or from a graph (or from a regression equation), taking account of any dilutions.
Report 0.1 M CaC12-extractable Cd as mg Cd/kg on an air-dry soil basis.
1. (a) As recommended by Murphy (1976), stoppers and tubing made of rubber and PVC should be avoided, as these are ‘dirty’ with respect to trace metal impurities.
(b) To clean glassware and plastic-ware, initially soak for about 2 days in 2% Decon, a non-ionic detergent. Afterwards, rinse with DDW, then soak in a 10% v/v HNO3/DDW solution for up to 7 days, followed by rinsing. Soaking-baths must be replenished periodically with relevant solutions. When the apparatus is dry and/or when not required for immediate use, store in clean, sealed, polyethylene bags, or invert on a polyethylene sheet in a dust-free cupboard. If laboratory-ware is known to contain insignificant Cd contamination due to prior use, the prescribed cleaning procedures may be expedited but not overlooked. Periodically monitor the effectiveness of these measures via the use of blanks and quality-control reference samples.
(c) Periodically tests the purity of the DDW using electrical conductivity (this should be ≤0.5 mS/m) and the dithizone test of Stout and Arnon (1939). Key points of the dithizone test are:
• Prepare dithizone reagent by dissolving 100 mg high-grade diphenylthiocarbazone in 10 mL of redistilled chloroform. Store this stock solution in metal free glassware and a sub-volume in a clean dropping bottle. [Since chloroform reacts slowly with oxygen/oxidizing agents to form phosgene, chlorine, and HCl, particularly when exposed to light and air (Riddick and Toops 1955), it must be stabilised if lengthy storage is envisaged. This can be achieved by adding 0.5–1.0% of redistilled ethanol.]
• Prepare a separating funnel by dispensing into it (in a fume cupboard) in about 1.0 mL DDW, 5 mL of redistilled chloroform, 5 drops of 6 M NH4OH and 3 drops of dithizone reagent, followed by vigorously shaking by inverting at regular intervals for 1 min or more. Next allow the chloroform layer to settle out (any build up of vapour must be released periodically through the stop-cock during shaking). (Prepare the 6 M NH4OH solution by diluting high purity NH4OH reagent with DDW.)
• Drain the chloroform layer to waste and retain the dithizone contained in the ammoniacal water in the separating funnel.
• Add 200 mL of the water to be tested and 5 mL of chloroform into the separating funnel containing the purified dithizone, shake vigorously for 1 min or more, then allow the chloroform layer to settle out. Any red or purple colour indicates the presence of one or more of the following metals: Zn, Cu, Pb, Ni, Co, Hg, Cd, Tl, Bi. For precise studies, confirm the presence or absence of Cd (and any other metals of interest) by scanning the concentrated aqueous dithizone extract on an ICP at recommended wave lengths for these metals. If there is no response when the ICP is operated at maximum sensitivity, the relevant metal can be assumed to be absent for the purposes of this soil test. A larger volume of water may be tested if more accuracy is thought necessary.
(d) Occasionally test for the presence of heavy metal contamination on the surface of items such as plastic tubing, gloves, etc with a few drops of the dithizone reagent. The formation of a red or purple colour indicates undesirable heavy metal contamination. If the dithizone’s colour is too intense, a sub-sample can be diluted before use with redistilled chloroform. Replace or decontaminate items if evidence of significant contamination is apparent.
2. Patterson and Settle (1976) recommend use of laboratory ware made of teflon, ultra-pure quartz and conventional polyethylene in preference to conventional borosilicate glass, polycarbonate, methacrylate, linear polyethylene, polypropylene, nylon, PVC, and platinum.
3. The 2% HNO3 solution is used to help stabilise the Cd Standard Solutions. At the final concentration, there is insufficient matrix effect to require 2% HNO3 (1+4) addition to unknown extract solutions (A Jeffrey, pers. comm.). According to Dr Sauerbeck (pers. comm.), the particulate-free soil extracts may be held for up to 10 days at 5°C if rapid analysis is not immediately possible. Re-extract if ‘flakes’ should form on the surface of these extract solutions during this period of delay.