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A comparative analysis of gold-rich plant material using various analytical methods
Microchemical Journal 81 (2005) 81 – 85 www.elsevier.com/locate/microc
A comparative analysis of gold-rich plant material using various analytical methods Christopher Andersona,b,T, Fabio Morenoa, Frank Geurtsc, Carel Wreesmannc, Mory Ghomsheid, John Meechd a
Institute of Natural Resources, Massey University, Palmerston North, New Zealand b Tiaki Resources Ltd., Palmerston North, New Zealand c Akzo Nobel Chemicals bv, Research-dept. CFC, 6800 SB Arnhem, The Netherlands d The Centre for Environmental Research in Minerals, Metals and Materials (CERM3), The University of British Columbia, Vancouver, Canada Received 22 January 2004; accepted 21 January 2005 Available online 5 March 2005
Abstract During 2003 a field demonstration of gold phytoextraction technology was conducted in Brazil. As there is no commercially available biogeochemical standard reference material with an elevated concentration of gold, the trial biomass was analysed for its gold content using five analytical methods, at five laboratories, to confirm the concentration of gold in the harvested plant material. Nitric and hydrochloric acid digest followed by ICP-OES solution analysis of a dore´ bead prepared through fire assay of vegetative material was considered the benchmark analytical method to which the other results were compared. The gold concentrations reported by the five laboratories varied widely. Flame atomic absorption analysis of a solution prepared through the digest of plant ash by aqua regia proved the most accurate analytical method relative to fire assay. Gold concentrations reported by a New Zealand commercial laboratory using ICP-MS and a standard dbiological materials digestT procedure were affected by the digest method employed. X-ray fluorescence results may have been affected by synthetic standards that were prepared specifically for this investigation. Alternatively, matrix effects may not have been fully corrected for using XRF. Analysis of metal-rich biomass is becoming more common due to the popularity of studies that use plants to absorb heavy metals. The results of our comparative investigation emphasise that due care and consideration must be given to the selection of the analytical method chosen to analyse such plant material. Our results also highlight the need for standard reference materials that are suitably enriched in metals, such that these may be of use to phytoextraction studies. D 2005 Elsevier B.V. All rights reserved. Keywords: Gold; Vegetation; Analytical method; Standard reference materials
1. Introduction Gold has been the focus of vegetation studies since the beginning of the 20th century. The quest for gold in plants has been driven not only by curiosity, but also by biogeochemistry. This is an exploration technique that seeks to link the concentration of gold in a plant to that in the underlying soil. Gold concentrations higher than an T Corresponding author. Institute of Natural Resources, Massey University, Palmerston North, New Zealand. Tel.: +64 6 356 9099; fax: +64 6 350 5632. E-mail address: [email protected] (C. Anderson). 0026-265X/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.microc.2005.01.004
expected background can indicate mineralisation. Biogeochemistry is particularly well suited in areas of disturbed land, where the provenance of soil collected during a geochemical soil sampling survey cannot always be known. Brooks et al. [1] published a thorough review of biogeochemistry, and recent applications of the technique have been described for mineral exploration in Australia [2]. Gold is not the only metal that has been studied in plants. Since the late 19th century scientists have recorded high concentrations of metals such as Cd, Cu, Co, Mn, Ni, Se, Tl and Zn in plants [3]. Today, plants that are enriched in metals are termed hyperaccumulators [4] and these plants
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can be used to recover metals from contaminated or metalrich soils (phytoextraction). There is a significant volume of literature published on the wider application of plants to clean up the environment. Robinson et al. [5] published recently an up to date review on phytoremediation. In 1998 New Zealand scientists reported that they had induced common mustard plants to hyperaccumulate gold [6]. Biogeochemical studies in North America over the previous decades had identified a background level of gold in plants as 0.2 Ag/kg [7], hence Anderson et al. [6] defined the hyperaccumulation concentration of gold as 1 mg/kg. Any report of a metal concentration in plants is dependant upon a reliable and accurate analytical procedure. A plethora of analytical techniques are today available to determine metal concentrations. Techniques range from relatively inexpensive flame atomic absorption spectrometry (FAAS), to costly neutron activation analysis (NAA). Atomic absorption depends upon a wet digest procedure to release metal cations into an acid solution, as do inductively coupled plasma optical emission spectrometry (ICP-OES), ICP mass spectrometry (ICP-MS), and graphite furnace AAS (GFAAS). Techniques such as NAA and X-ray fluorescence work on a solid sample. Hall et al. [8,9] described the importance of choosing the correct analytical technique for gold determinations in biogeochemical samples, and stressed the necessity of a suitable sample preparation method. The complete procedure of sample preparation followed by elemental quantification using a suitable analytical technique is described as the analytical method. To manage the quality of analytical data, standard reference materials (SRM) are commercially available. These are analytical samples with a standard metal concentration. The field of biogeochemistry is, however, poorly represented by such reference materials. One commercial source, for example, is The National Institute of Standards and Technology in the USA. This companies website reports one SRM that has an indicated gold concentration: 0.001 mg/kg in SRM 1515, apple leaves. Note this is only an indicated and therefore uncertified concentration that is 1000 times lower than the defined limit of gold hyperaccumulation in plants. The authors of this current work have been studying gold hyperaccumulation for several years. In 2003 a field demonstration for gold phytoextraction was carried out at a gold mine in Brazil [10]. The plant species that showed the greatest ability to concentrate gold reported an average gold concentration of 39 mg/kg as determined by FAAS. But due to the lack of a suitable SRM, how were the authors to know whether this gold assay result was accurate? A comparative investigation of the gold and copper concentration in the harvested biomass was conducted to answer this question. The following techniques were used to assay the plant material: FAAS, GFAAS, ICP-MS, XRF and fire assay followed by ICPOES. Dry plant material was submitted to five different
laboratories as part of this investigation. The results of the comparative study are presented in this paper.
2. Methodology 2.1. Preparation of the plant material for analysis Between April and June 2003 a field demonstration of gold phyto-reclamation technology was conducted at the Fazenda Brasileiro mine, in Bahia, Brazil [10]. The trial was a collaborative effort involving Massey University, The University of British Columbia and the Brazilian mining company CVRD. No known plant species will naturally hyperaccumulate gold, however plants will accumulate metals that are made soluble using an appropriate lixiviant. A mini ore pad 15 m15 m0.5 m deep was prepared by CVRD and hand seeded with two plant species (Brassica juncea and Zea mays). The ore pad was underlain with geotextile and fertilised with a commercial agricultural fertiliser. The pad was irrigated with a sprinkler system twice daily. Six weeks after seeding, the plot was treated with cyanide and a combination of thiocyanate and hydrogen peroxide to induce the uptake of gold (Table 1). Chemicals were hand irrigated onto the plot as 400 L of solution using a small electric pump at an approximate flow rate of 0.55 L/ s. Two elevated 1000 L tanks provided the necessary head pressure. One week after treatment all above-ground plant material was harvested, air-dried, packaged and shipped to New Zealand. Upon receipt at Massey University the biomass was unpackaged and dried at 70 8C. A bulk sample (100 g) for each treatment and species combination was then ground using a Cyclotec 1093 Sample Mill. Subsamples of ground biomass were distributed for analysis at internal (Massey University) and external laboratories. Ten samples were analysed in total, each representing a different plant and chemical combination. Sample 10 was a sample of nontreated, control, biomass. Three replicates for each sample were analysed at Massey University and the results presented are the mean concentration for the three analyses. For all external analyses only one replicate was analysed.
Table 1 Description of lixiviant treatments Treatment
Chemical
Application
Solution concentration
1
NH4SCN/ peroxide NaCN KCN
0.3 g/kg (0.23 g/kg SCN) as a 3.7% peroxide solution 0.15 g/kg (0.08 g/kg CN) 0.15 g/kg (0.06 g/kg CN)
4.3 g/L of SCN 1.40 g/L CN 1.03 g/L CN
2 3
The chemical application rate is presented in two ways in this table. The total mass of chemical applied per treatment is recorded, as well as the calculated mass of anion. Peroxide in combination with thiocyanate has been shown to be a more efficient lixiviant for gold than thiocyanate alone (Massey University unpublished study).
C. Anderson et al. / Microchemical Journal 81 (2005) 81–85
2.2. Semi-quantitative ICP-MS analysis (designated ICP-MS) A 0.5 g subsample of plant material was accurately weighed into a capped 50 mL centrifuge tube. Water (2 mL of type 1) was added to assist with sample wetting. Concentrated nitric acid (2.5 mL) and concentrated hydrochloric acid (0.5 mL) were added, and the sample was digested for 3 h at 100 8C. The digest solution was made to a volume of 50 mL giving a final concentration of 5% nitric and 1% hydrochloric acids. The sample solutions were then analysed using ICP-MS. 2.3. XRF laboratory one (XRF1) A 10 g subsample of oven re-dried (60 8C) and finely ground plant material was combined with 2.5 g of wax binder to prepare a sample briquette. Each briquette was then exposed to primary X-rays. Secondary X-ray counts yielded Au and Cu metal concentrations. A set of new analytical programs were constructed for this analysis due to the lack of a commercial method at this laboratory to analyse plant samples with high levels of gold and copper. Synthetic standards were generated by spiking pure cellulose with standard solutions of Au and Cu.
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The plant material was ashed at 530 8C for 14 h to liberate volatile plant components. The ash was transferred into plastic beakers and digested with 5 mL of aqua regia on a water bath to low volume. The digestion residue was made to 10 mL with 2 M hydrochloric acid. Solutions were then analysed for Au and Cu by flame atomic absorption spectrometry on a GBC Avanta spectrometer operated by the Institute of Natural Resources, Massey University, New Zealand. No loss of gold has ever been attributed to the ashing process during 6 years of analysis of biological samples at Massey University. Standards were prepared through dilution of a 1000 mg/L Spectrosol gold standard solution. 2.7. Graphite furnace atomic absorption spectrometry (GFAA) A 2 mL aliquot of the digest solution described for flame atomic absorption was extracted with 5 mL of methyl isobutyl ketone (MIBK). The organic phase was then analysed for Au by graphite furnace atomic adsorption spectrometry using a GBC 909AA spectrometer operated by the Institute of Natural Resources, Massey University, New Zealand. Standards were prepared through dilution of a 1000 mg/L Spectrosol gold standard solution.
2.4. Semi-quantitative XRF laboratory two (XRF2) A weighed subsample for each sample of ground and dry plant material was analysed as received using a Philips PW 1404 X-ray Fluorescence spectrometer with a Cr tube. UniQuantk was used to perform a semi-quantitative analysis, yielding the concentration of all elements in the sample. The reported detection limit for Au and Cu using this method was 10 mg/kg. UniQuantk software allows standardless semi-quantitative to quantitative XRF analysis and requires no sample preparation.
3. Results and discussion 3.1. Gold Analytical results for the Au concentration in plant material are summarised in Table 2. Fire assay is regarded as a standard technique for the precious metal analysis of geological samples. The laboratory that conducted the fire assay analysis is a reputable and certified laboratory and therefore, the fire assay result is considered a reliable
2.5. Fire assay and ICP-OES (fire assay) Analysis by fire assay was carried out by the Acme Analytical Laboratory, Vancouver, Canada. A weighed subsample of plant material was custom blended with fire assay fluxes, PbO litharge and an Ag inquart. The charge was fired at 1050 8C to liberate Au into a molten Pb-metal phase. After cooling the Pb button was recovered, placed in a cupel and fired to generate a dore´ bead. The bead was weighed, leached with hot nitric acid (1 mL) then dissolved in hydrochloric acid (10 mL). The digest solution was then analysed for Au by Acme Analytical Laboratories, Vancouver, Canada, using a Jarrel-Ash Atomcorp model 975 ICP emission spectrometer. 2.6. Flame atomic absorption spectrometry (FAA) Replicate subsamples of plant material (approximately 0.2 g) were accurately weighed into borosilicate test tubes.
Table 2 The Au concentration in 10 samples (different plant and treatment combinations) harvested from the phytoextraction field trial as reported by five analytical laboratories (five analytical methods) Sample
1 2 3 4 5 6 7 8 9 10
FAA
GFAA
Mean (S.D.)
Mean (S.D.)
b1 b1 b1 19.6 (1.5) 29.7 (2.7) 38.9 (1.1) 8.1 (0.7) 10.4 (2.9) 30.4 (3.4) b1
2.4 (0.5) b1 b1 13.0 (0.3) 12.3 (2.0) 29.2 (2.7) 6.3 (0.2) 7.7 (0.8) 23.1 (3.1) 0.2 (0.1)
XRF1
XRF2
ICP-MS
Fire assay
b1 b1 b1 5 6 9 5 6 10 nd
b10 b10 b10 b10 b10 40 b10 b10 b10 nd
1.44 0.56 0.23 3.79 3.75 8.33 1.95 2.93 7.14 0.09
5.3 1.2 1.0 17.2 16.1 35.9 10.7 15.3 35.1 nd
All concentrations are mg/kg. nd signifies no analysis of sample 10, the control treatment. For FAA and GFAA, n=3. For other methods, n=1.
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determination of the gold concentration in the plant material. FAA analysis of the digested ash proved to be an accurate method when compared with fire assay. GFAA of the MIBK extracted digest solution reported concentrations approximately 25% below those reported by FAA. Two explanations can be proposed for this 25% discrepancy: (1) gold may not have been quantitatively extracted with MIBK from the digest solutions or (2) the quality of GFAA determinations may have been hampered by the presence of MIBK that was not sufficiently considered during calibration. The low gold concentrations reported through external ICP analysis of the digest solution were surprising, and are attributed to the digest technique employed. The acid strength was 50% when the water dilution (wetting agent) is taken into account. Research experience at Massey University shows that dilute acids will not digest plant material (unpublished). We therefore assume that 50% aqua regia did not completely oxidise the plant material, and therefore did not reliably release the gold particles enclosed within the cellular tissues for dissolution as a chlorocomplex. The laboratory that conducted this analysis used a standard biological materials digest procedure. Clearly, digestion of biogeochemical samples for Au analysis is not a standard procedure. It is unlikely that this laboratory had previously received a biological sample with an elevated Au concentration. XRF analysis of the plant material by XRF Laboratory 1 reported a very low gold concentration for all samples. The limit of detection for the method at this laboratory is reported as 1 mg/kg. The low concentrations were surprising and the reason is unknown. The error may lie in the standards generated for the work or in matrix effects that were not corrected for during analysis. The laboratory prepared synthetic standards due to a lack of commercially available herbage with a high gold concentration. Interpretation of the data from XRF Laboratory 2 is complicated by a high limit of detection for gold in the plant material (10 mg/kg). One sample (number 6) is in good agreement with the FAA and fire assay result, however no conclusion can be drawn from this one value. Semi-quantitative XRF as used by Laboratory 2 has an error of F50%. 3.2. Copper Analytical results for the Cu concentration in plant material are summarised in Table 3. GFAA and ICP analysis of the fire assay dore´ bead were not used to determine Cu concentrations. The concentrations determined through FAA and ICP analysis of plant digests are in good agreement. This is in contrast to the Au assay results reported for these two analytical methods that employed digest preparation techniques. Copper is more reactive than Au and will presumably have been digested during the procedure used by the external laboratory.
Table 3 The Cu concentration in 10 samples (different plant and treatment combinations) harvested from the phytoextraction field trial as reported by four analytical laboratories (three analytical methods) Sample
FAA
XRF1
XRF2
ICP-MS
29 21 22 67 40 132 38 65 134 nd
100 40 50 140 100 360 50 70 380 nd
164 82 85 261 154 583 91 202 603 75
Mean (S.D.) 1 2 3 4 5 6 7 8 9 10
133 57 62 137 100 541 61 112 571 50
(11.0) (6.3) (8.7) (7.3) (2.2) (14.0) (5.7) (4.8) (36.0) (36)
All concentrations are mg/kg. nd signifies no analysis of sample 10, the control treatment. For FAA, n=3. For other methods, n=1.
Of the two XRF techniques, the results obtained from Laboratory 2 agree reasonably well with FAA and ICP. Results for XRF conducted by Laboratory 1 report concentrations that are much lower than the other three methods. Again, it seems reasonable to assume that the error may lie with the synthetic standards generated for this work.
4. Summary and conclusions For the analytical comparison described in this paper, ICP analysis of a fire assay dore´ bead was taken as a benchmark method to determine the gold concentration in plant material. Fire assay is regarded by the resource industry as an accurate and reliable assay technique. Flame atomic absorption at Massey University of a solution prepared by the aqua regia digest of ashed plant samples proved an accurate analytical method relative to the chosen benchmark. Fortunately, FAA could be employed during this work due to the high gold concentration in the plant material. Lower concentrations would have necessitated reliance on the more sensitive GFAA technique. Based on this conclusion FAA data were used by Anderson et al. [10] to describe the outcomes of the phytoextraction field trial. FAA is routinely used at Massey University to determine the gold concentration in hyperaccumulator plants. The analytical comparison emphasises the importance of the digest procedure employed. The low Au concentrations in the experimental samples reported by the ICPMS laboratory are explained through the standard dbiological materials digestT used by this laboratory. There is, however, no standard digest technique for gold hyperaccumulators. The analytical technique used by the ICPMS laboratory proved reliable for Cu, but must be modified for future Au determinations. Ashing prior to digestion appears to be the most accurate method for sample preparation. FAA analysis of digested ash is now
C. Anderson et al. / Microchemical Journal 81 (2005) 81–85
adopted as the analytical method of choice for gold phytoextraction studies at Massey University. Results from the two XRF laboratories are conflicting. Concentrations reported by XRF Laboratory 2 appears to agree with FAA, however, interpretation is limited due to the low limit of detection (10 mg/kg for Au) and 50% error on the semi-quantitative analysis. Concentrations reported by XRF Laboratory 1 are much lower than FAA. This discrepancy is attributed to the make up of synthetic standards that were prepared specifically for this work. Laboratories that analyse plant samples enriched in gold should consider the wide variation of concentrations reported using the five different analytical techniques presented in this paper. This is especially relevant for laboratories that are ready to embark on new gold phytoextraction studies. Conventional and standard analytical methods may not be suitable for such new studies. Accuracy of results is hampered by a lack of gold-enriched reference standard. Fortunately, as a result of the research at Massey University described in this paper, a plant standard with a reliable gold concentration of approximately 39 mg/ kg has now been prepared that is routinely used during analytical determinations.
Acknowledgements The funding agencies for this research are acknowledged: CERM3 at the University of British Columbia (through the
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Canada Foundation for Innovation Project Grant 2545); Akzo Nobel Chemicals Pte. Ltd. (Singapore); the New Zealand Foundation for Research Science and Technology (Contract MAUX0020); and the National Council for Scientific and Technological Development of Brazil (CNPq).
References [1] R.R. Brooks, C.E. Dunn, G.E.M. Hall (Eds.), Biological Systems in Mineral Exploration and Processing, Ellis Horwood, Hemel Hempstead, 1995, 538 pp. [2] D.C. Arne, J.E. Stott, H.M. Waldron, J. Geochem. Explor. 67 (1999) 1. [3] R.R. Brooks (Ed.), Plants that Hyperaccumulate Heavy Metals, CAB International, Wallingford, 1998, 380 pp. [4] R.R. Brooks, J. Lee, R.D. Reeves, T. Jaffre, J. Geochem. Explor. 7 (1977) 49. [5] B. Robinson, S. Green, T. Mills, B. Clothier, M. van der Velde, R. Laplane, L. Fung, M. Deurer, S. Hurst, T. Thayalakumaran, C. van den Dijssel, Aust. J. Soil Res. 41 (2003) 599. [6] C.W.N. Anderson, R.R. Brooks, R.B. Stewart, R. Simcock, Nature 395 (1998) 553. [7] C.E. Dunn, in: R.R. Brooks, C.E. Dunn, G.E.M. Hall (Eds.), Biological Systems in Mineral Exploration and Processing, Ellis Horwood, Hemel Hempstead, 1999, p. 371. [8] G.E.M. Hall, J.-C. Pelchat, C.E. Dunn, J. Geochem. Explor. 37 (1990) 1. [9] G.E.M. Hall, A.N. Rencz, A.I. Maclaurin, J. Geochem. Explor. 41 (1991) 291. [10] C. Anderson, F. Moreno, J. Meech, Miner. Eng. 18 (2005) 385.
A comparative analysis of gold-rich plant material using various analytical methods Christopher Andersona,b,T, Fabio Morenoa, Frank Geurtsc, Carel Wreesmannc, Mory Ghomsheid, John Meechd a
Institute of Natural Resources, Massey University, Palmerston North, New Zealand b Tiaki Resources Ltd., Palmerston North, New Zealand c Akzo Nobel Chemicals bv, Research-dept. CFC, 6800 SB Arnhem, The Netherlands d The Centre for Environmental Research in Minerals, Metals and Materials (CERM3), The University of British Columbia, Vancouver, Canada Received 22 January 2004; accepted 21 January 2005 Available online 5 March 2005
Abstract During 2003 a field demonstration of gold phytoextraction technology was conducted in Brazil. As there is no commercially available biogeochemical standard reference material with an elevated concentration of gold, the trial biomass was analysed for its gold content using five analytical methods, at five laboratories, to confirm the concentration of gold in the harvested plant material. Nitric and hydrochloric acid digest followed by ICP-OES solution analysis of a dore´ bead prepared through fire assay of vegetative material was considered the benchmark analytical method to which the other results were compared. The gold concentrations reported by the five laboratories varied widely. Flame atomic absorption analysis of a solution prepared through the digest of plant ash by aqua regia proved the most accurate analytical method relative to fire assay. Gold concentrations reported by a New Zealand commercial laboratory using ICP-MS and a standard dbiological materials digestT procedure were affected by the digest method employed. X-ray fluorescence results may have been affected by synthetic standards that were prepared specifically for this investigation. Alternatively, matrix effects may not have been fully corrected for using XRF. Analysis of metal-rich biomass is becoming more common due to the popularity of studies that use plants to absorb heavy metals. The results of our comparative investigation emphasise that due care and consideration must be given to the selection of the analytical method chosen to analyse such plant material. Our results also highlight the need for standard reference materials that are suitably enriched in metals, such that these may be of use to phytoextraction studies. D 2005 Elsevier B.V. All rights reserved. Keywords: Gold; Vegetation; Analytical method; Standard reference materials
1. Introduction Gold has been the focus of vegetation studies since the beginning of the 20th century. The quest for gold in plants has been driven not only by curiosity, but also by biogeochemistry. This is an exploration technique that seeks to link the concentration of gold in a plant to that in the underlying soil. Gold concentrations higher than an T Corresponding author. Institute of Natural Resources, Massey University, Palmerston North, New Zealand. Tel.: +64 6 356 9099; fax: +64 6 350 5632. E-mail address: [email protected] (C. Anderson). 0026-265X/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.microc.2005.01.004
expected background can indicate mineralisation. Biogeochemistry is particularly well suited in areas of disturbed land, where the provenance of soil collected during a geochemical soil sampling survey cannot always be known. Brooks et al. [1] published a thorough review of biogeochemistry, and recent applications of the technique have been described for mineral exploration in Australia [2]. Gold is not the only metal that has been studied in plants. Since the late 19th century scientists have recorded high concentrations of metals such as Cd, Cu, Co, Mn, Ni, Se, Tl and Zn in plants [3]. Today, plants that are enriched in metals are termed hyperaccumulators [4] and these plants
82
C. Anderson et al. / Microchemical Journal 81 (2005) 81–85
can be used to recover metals from contaminated or metalrich soils (phytoextraction). There is a significant volume of literature published on the wider application of plants to clean up the environment. Robinson et al. [5] published recently an up to date review on phytoremediation. In 1998 New Zealand scientists reported that they had induced common mustard plants to hyperaccumulate gold [6]. Biogeochemical studies in North America over the previous decades had identified a background level of gold in plants as 0.2 Ag/kg [7], hence Anderson et al. [6] defined the hyperaccumulation concentration of gold as 1 mg/kg. Any report of a metal concentration in plants is dependant upon a reliable and accurate analytical procedure. A plethora of analytical techniques are today available to determine metal concentrations. Techniques range from relatively inexpensive flame atomic absorption spectrometry (FAAS), to costly neutron activation analysis (NAA). Atomic absorption depends upon a wet digest procedure to release metal cations into an acid solution, as do inductively coupled plasma optical emission spectrometry (ICP-OES), ICP mass spectrometry (ICP-MS), and graphite furnace AAS (GFAAS). Techniques such as NAA and X-ray fluorescence work on a solid sample. Hall et al. [8,9] described the importance of choosing the correct analytical technique for gold determinations in biogeochemical samples, and stressed the necessity of a suitable sample preparation method. The complete procedure of sample preparation followed by elemental quantification using a suitable analytical technique is described as the analytical method. To manage the quality of analytical data, standard reference materials (SRM) are commercially available. These are analytical samples with a standard metal concentration. The field of biogeochemistry is, however, poorly represented by such reference materials. One commercial source, for example, is The National Institute of Standards and Technology in the USA. This companies website reports one SRM that has an indicated gold concentration: 0.001 mg/kg in SRM 1515, apple leaves. Note this is only an indicated and therefore uncertified concentration that is 1000 times lower than the defined limit of gold hyperaccumulation in plants. The authors of this current work have been studying gold hyperaccumulation for several years. In 2003 a field demonstration for gold phytoextraction was carried out at a gold mine in Brazil [10]. The plant species that showed the greatest ability to concentrate gold reported an average gold concentration of 39 mg/kg as determined by FAAS. But due to the lack of a suitable SRM, how were the authors to know whether this gold assay result was accurate? A comparative investigation of the gold and copper concentration in the harvested biomass was conducted to answer this question. The following techniques were used to assay the plant material: FAAS, GFAAS, ICP-MS, XRF and fire assay followed by ICPOES. Dry plant material was submitted to five different
laboratories as part of this investigation. The results of the comparative study are presented in this paper.
2. Methodology 2.1. Preparation of the plant material for analysis Between April and June 2003 a field demonstration of gold phyto-reclamation technology was conducted at the Fazenda Brasileiro mine, in Bahia, Brazil [10]. The trial was a collaborative effort involving Massey University, The University of British Columbia and the Brazilian mining company CVRD. No known plant species will naturally hyperaccumulate gold, however plants will accumulate metals that are made soluble using an appropriate lixiviant. A mini ore pad 15 m15 m0.5 m deep was prepared by CVRD and hand seeded with two plant species (Brassica juncea and Zea mays). The ore pad was underlain with geotextile and fertilised with a commercial agricultural fertiliser. The pad was irrigated with a sprinkler system twice daily. Six weeks after seeding, the plot was treated with cyanide and a combination of thiocyanate and hydrogen peroxide to induce the uptake of gold (Table 1). Chemicals were hand irrigated onto the plot as 400 L of solution using a small electric pump at an approximate flow rate of 0.55 L/ s. Two elevated 1000 L tanks provided the necessary head pressure. One week after treatment all above-ground plant material was harvested, air-dried, packaged and shipped to New Zealand. Upon receipt at Massey University the biomass was unpackaged and dried at 70 8C. A bulk sample (100 g) for each treatment and species combination was then ground using a Cyclotec 1093 Sample Mill. Subsamples of ground biomass were distributed for analysis at internal (Massey University) and external laboratories. Ten samples were analysed in total, each representing a different plant and chemical combination. Sample 10 was a sample of nontreated, control, biomass. Three replicates for each sample were analysed at Massey University and the results presented are the mean concentration for the three analyses. For all external analyses only one replicate was analysed.
Table 1 Description of lixiviant treatments Treatment
Chemical
Application
Solution concentration
1
NH4SCN/ peroxide NaCN KCN
0.3 g/kg (0.23 g/kg SCN) as a 3.7% peroxide solution 0.15 g/kg (0.08 g/kg CN) 0.15 g/kg (0.06 g/kg CN)
4.3 g/L of SCN 1.40 g/L CN 1.03 g/L CN
2 3
The chemical application rate is presented in two ways in this table. The total mass of chemical applied per treatment is recorded, as well as the calculated mass of anion. Peroxide in combination with thiocyanate has been shown to be a more efficient lixiviant for gold than thiocyanate alone (Massey University unpublished study).
C. Anderson et al. / Microchemical Journal 81 (2005) 81–85
2.2. Semi-quantitative ICP-MS analysis (designated ICP-MS) A 0.5 g subsample of plant material was accurately weighed into a capped 50 mL centrifuge tube. Water (2 mL of type 1) was added to assist with sample wetting. Concentrated nitric acid (2.5 mL) and concentrated hydrochloric acid (0.5 mL) were added, and the sample was digested for 3 h at 100 8C. The digest solution was made to a volume of 50 mL giving a final concentration of 5% nitric and 1% hydrochloric acids. The sample solutions were then analysed using ICP-MS. 2.3. XRF laboratory one (XRF1) A 10 g subsample of oven re-dried (60 8C) and finely ground plant material was combined with 2.5 g of wax binder to prepare a sample briquette. Each briquette was then exposed to primary X-rays. Secondary X-ray counts yielded Au and Cu metal concentrations. A set of new analytical programs were constructed for this analysis due to the lack of a commercial method at this laboratory to analyse plant samples with high levels of gold and copper. Synthetic standards were generated by spiking pure cellulose with standard solutions of Au and Cu.
83
The plant material was ashed at 530 8C for 14 h to liberate volatile plant components. The ash was transferred into plastic beakers and digested with 5 mL of aqua regia on a water bath to low volume. The digestion residue was made to 10 mL with 2 M hydrochloric acid. Solutions were then analysed for Au and Cu by flame atomic absorption spectrometry on a GBC Avanta spectrometer operated by the Institute of Natural Resources, Massey University, New Zealand. No loss of gold has ever been attributed to the ashing process during 6 years of analysis of biological samples at Massey University. Standards were prepared through dilution of a 1000 mg/L Spectrosol gold standard solution. 2.7. Graphite furnace atomic absorption spectrometry (GFAA) A 2 mL aliquot of the digest solution described for flame atomic absorption was extracted with 5 mL of methyl isobutyl ketone (MIBK). The organic phase was then analysed for Au by graphite furnace atomic adsorption spectrometry using a GBC 909AA spectrometer operated by the Institute of Natural Resources, Massey University, New Zealand. Standards were prepared through dilution of a 1000 mg/L Spectrosol gold standard solution.
2.4. Semi-quantitative XRF laboratory two (XRF2) A weighed subsample for each sample of ground and dry plant material was analysed as received using a Philips PW 1404 X-ray Fluorescence spectrometer with a Cr tube. UniQuantk was used to perform a semi-quantitative analysis, yielding the concentration of all elements in the sample. The reported detection limit for Au and Cu using this method was 10 mg/kg. UniQuantk software allows standardless semi-quantitative to quantitative XRF analysis and requires no sample preparation.
3. Results and discussion 3.1. Gold Analytical results for the Au concentration in plant material are summarised in Table 2. Fire assay is regarded as a standard technique for the precious metal analysis of geological samples. The laboratory that conducted the fire assay analysis is a reputable and certified laboratory and therefore, the fire assay result is considered a reliable
2.5. Fire assay and ICP-OES (fire assay) Analysis by fire assay was carried out by the Acme Analytical Laboratory, Vancouver, Canada. A weighed subsample of plant material was custom blended with fire assay fluxes, PbO litharge and an Ag inquart. The charge was fired at 1050 8C to liberate Au into a molten Pb-metal phase. After cooling the Pb button was recovered, placed in a cupel and fired to generate a dore´ bead. The bead was weighed, leached with hot nitric acid (1 mL) then dissolved in hydrochloric acid (10 mL). The digest solution was then analysed for Au by Acme Analytical Laboratories, Vancouver, Canada, using a Jarrel-Ash Atomcorp model 975 ICP emission spectrometer. 2.6. Flame atomic absorption spectrometry (FAA) Replicate subsamples of plant material (approximately 0.2 g) were accurately weighed into borosilicate test tubes.
Table 2 The Au concentration in 10 samples (different plant and treatment combinations) harvested from the phytoextraction field trial as reported by five analytical laboratories (five analytical methods) Sample
1 2 3 4 5 6 7 8 9 10
FAA
GFAA
Mean (S.D.)
Mean (S.D.)
b1 b1 b1 19.6 (1.5) 29.7 (2.7) 38.9 (1.1) 8.1 (0.7) 10.4 (2.9) 30.4 (3.4) b1
2.4 (0.5) b1 b1 13.0 (0.3) 12.3 (2.0) 29.2 (2.7) 6.3 (0.2) 7.7 (0.8) 23.1 (3.1) 0.2 (0.1)
XRF1
XRF2
ICP-MS
Fire assay
b1 b1 b1 5 6 9 5 6 10 nd
b10 b10 b10 b10 b10 40 b10 b10 b10 nd
1.44 0.56 0.23 3.79 3.75 8.33 1.95 2.93 7.14 0.09
5.3 1.2 1.0 17.2 16.1 35.9 10.7 15.3 35.1 nd
All concentrations are mg/kg. nd signifies no analysis of sample 10, the control treatment. For FAA and GFAA, n=3. For other methods, n=1.
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determination of the gold concentration in the plant material. FAA analysis of the digested ash proved to be an accurate method when compared with fire assay. GFAA of the MIBK extracted digest solution reported concentrations approximately 25% below those reported by FAA. Two explanations can be proposed for this 25% discrepancy: (1) gold may not have been quantitatively extracted with MIBK from the digest solutions or (2) the quality of GFAA determinations may have been hampered by the presence of MIBK that was not sufficiently considered during calibration. The low gold concentrations reported through external ICP analysis of the digest solution were surprising, and are attributed to the digest technique employed. The acid strength was 50% when the water dilution (wetting agent) is taken into account. Research experience at Massey University shows that dilute acids will not digest plant material (unpublished). We therefore assume that 50% aqua regia did not completely oxidise the plant material, and therefore did not reliably release the gold particles enclosed within the cellular tissues for dissolution as a chlorocomplex. The laboratory that conducted this analysis used a standard biological materials digest procedure. Clearly, digestion of biogeochemical samples for Au analysis is not a standard procedure. It is unlikely that this laboratory had previously received a biological sample with an elevated Au concentration. XRF analysis of the plant material by XRF Laboratory 1 reported a very low gold concentration for all samples. The limit of detection for the method at this laboratory is reported as 1 mg/kg. The low concentrations were surprising and the reason is unknown. The error may lie in the standards generated for the work or in matrix effects that were not corrected for during analysis. The laboratory prepared synthetic standards due to a lack of commercially available herbage with a high gold concentration. Interpretation of the data from XRF Laboratory 2 is complicated by a high limit of detection for gold in the plant material (10 mg/kg). One sample (number 6) is in good agreement with the FAA and fire assay result, however no conclusion can be drawn from this one value. Semi-quantitative XRF as used by Laboratory 2 has an error of F50%. 3.2. Copper Analytical results for the Cu concentration in plant material are summarised in Table 3. GFAA and ICP analysis of the fire assay dore´ bead were not used to determine Cu concentrations. The concentrations determined through FAA and ICP analysis of plant digests are in good agreement. This is in contrast to the Au assay results reported for these two analytical methods that employed digest preparation techniques. Copper is more reactive than Au and will presumably have been digested during the procedure used by the external laboratory.
Table 3 The Cu concentration in 10 samples (different plant and treatment combinations) harvested from the phytoextraction field trial as reported by four analytical laboratories (three analytical methods) Sample
FAA
XRF1
XRF2
ICP-MS
29 21 22 67 40 132 38 65 134 nd
100 40 50 140 100 360 50 70 380 nd
164 82 85 261 154 583 91 202 603 75
Mean (S.D.) 1 2 3 4 5 6 7 8 9 10
133 57 62 137 100 541 61 112 571 50
(11.0) (6.3) (8.7) (7.3) (2.2) (14.0) (5.7) (4.8) (36.0) (36)
All concentrations are mg/kg. nd signifies no analysis of sample 10, the control treatment. For FAA, n=3. For other methods, n=1.
Of the two XRF techniques, the results obtained from Laboratory 2 agree reasonably well with FAA and ICP. Results for XRF conducted by Laboratory 1 report concentrations that are much lower than the other three methods. Again, it seems reasonable to assume that the error may lie with the synthetic standards generated for this work.
4. Summary and conclusions For the analytical comparison described in this paper, ICP analysis of a fire assay dore´ bead was taken as a benchmark method to determine the gold concentration in plant material. Fire assay is regarded by the resource industry as an accurate and reliable assay technique. Flame atomic absorption at Massey University of a solution prepared by the aqua regia digest of ashed plant samples proved an accurate analytical method relative to the chosen benchmark. Fortunately, FAA could be employed during this work due to the high gold concentration in the plant material. Lower concentrations would have necessitated reliance on the more sensitive GFAA technique. Based on this conclusion FAA data were used by Anderson et al. [10] to describe the outcomes of the phytoextraction field trial. FAA is routinely used at Massey University to determine the gold concentration in hyperaccumulator plants. The analytical comparison emphasises the importance of the digest procedure employed. The low Au concentrations in the experimental samples reported by the ICPMS laboratory are explained through the standard dbiological materials digestT used by this laboratory. There is, however, no standard digest technique for gold hyperaccumulators. The analytical technique used by the ICPMS laboratory proved reliable for Cu, but must be modified for future Au determinations. Ashing prior to digestion appears to be the most accurate method for sample preparation. FAA analysis of digested ash is now
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adopted as the analytical method of choice for gold phytoextraction studies at Massey University. Results from the two XRF laboratories are conflicting. Concentrations reported by XRF Laboratory 2 appears to agree with FAA, however, interpretation is limited due to the low limit of detection (10 mg/kg for Au) and 50% error on the semi-quantitative analysis. Concentrations reported by XRF Laboratory 1 are much lower than FAA. This discrepancy is attributed to the make up of synthetic standards that were prepared specifically for this work. Laboratories that analyse plant samples enriched in gold should consider the wide variation of concentrations reported using the five different analytical techniques presented in this paper. This is especially relevant for laboratories that are ready to embark on new gold phytoextraction studies. Conventional and standard analytical methods may not be suitable for such new studies. Accuracy of results is hampered by a lack of gold-enriched reference standard. Fortunately, as a result of the research at Massey University described in this paper, a plant standard with a reliable gold concentration of approximately 39 mg/ kg has now been prepared that is routinely used during analytical determinations.
Acknowledgements The funding agencies for this research are acknowledged: CERM3 at the University of British Columbia (through the
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Canada Foundation for Innovation Project Grant 2545); Akzo Nobel Chemicals Pte. Ltd. (Singapore); the New Zealand Foundation for Research Science and Technology (Contract MAUX0020); and the National Council for Scientific and Technological Development of Brazil (CNPq).
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