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Revista Brasileira de Ciência do Solo

Print version ISSN 0100-0683On-line version ISSN 1806-9657

Rev. Bras. Ciênc. Solo vol.39 no.3 Viçosa May/June 2015 


Methods to Quantify Nickel in Soils and Plant Tissues

Métodos de Quantificação de Níquel em Solo e Tecido Vegetal

Bruna Wurr Rodak (1)   *  

Milton Ferreira de Moraes (2)   (3)  

João Augusto Lopes Pascoalino (3)  

Adilson de Oliveira Junior (4)  

Cesar de Castro (4)  

Volnei Pauletti (3)  

(1)Universidade Federal de Lavras, Departamento de Ciência do Solo, Lavras, Minas Gerais, Brasil.

(2)Universidade Federal de Mato Grosso, Instituto de Ciências Exatas e da Terra, Campus Universitário do Araguaia, Barra do Garças, Mato Grosso, Brasil.

(3)Universidade Federal do Paraná, Departamento de Ciência do Solo e Engenharia Agrícola, Curitiba, Paraná, Brasil.

(4)Empresa Brasileira de Pesquisa Agropecuária, Centro Nacional de Pesquisa em Soja, Londrina, Paraná, Brasil.


In comparison with other micronutrients, the levels of nickel (Ni) available in soils and plant tissues are very low, making quantification very difficult. The objective of this paper is to present optimized determination methods of Ni availability in soils by extractants and total content in plant tissues for routine commercial laboratory analyses. Samples of natural and agricultural soils were processed and analyzed by Mehlich-1 extraction and by DTPA. To quantify Ni in the plant tissues, samples were digested with nitric acid in a closed system in a microwave oven. The measurement was performed by inductively coupled plasma/optical emission spectrometry (ICP-OES). There was a positive and significant correlation between the levels of available Ni in the soils subjected to Mehlich-1 and DTPA extraction, while for plant tissue samples the Ni levels recovered were high and similar to the reference materials. The availability of Ni in some of the natural soil and plant tissue samples were lower than the limits of quantification. Concentrations of this micronutrient were higher in the soil samples in which Ni had been applied. Nickel concentration differed in the plant parts analyzed, with highest levels in the grains of soybean. The grain, in comparison with the shoot and leaf concentrations, were better correlated with the soil available levels for both extractants. The methods described in this article were efficient in quantifying Ni and can be used for routine laboratory analysis of soils and plant tissues.

Key words: micronutrient; availability; Mehlich-1; DTPA; ICP-OES


Em comparação a outros micronutrientes, os teores de níquel (Ni) disponíveis no solo e tecido vegetal são bastante baixos, com alto grau de dificuldade para a quantificação deles. O objetivo deste estudo foi apresentar métodos otimizados de determinação da disponibilidade de Ni em solos com extratores e teor total em tecido vegetal para análises de rotina em laboratórios comerciais. Foram processadas e analisadas amostras de solos naturais e agrícolas com extração pelo extrator Mehlich-1 e por DTPA. Para quantificar o Ni no tecido vegetal, as amostras foram digeridas com ácido nítrico em sistema fechado em forno micro-ondas. A determinação foi realizada em espectrômetro de emissão óptica com plasma induzido (ICP-OES). Houve correlação positiva e significativa entre os teores disponíveis de Ni nos solos extraídos em Mehlich-1 e DTPA, enquanto, para o tecido vegetal os teores de Ni recuperados foram altos e similares aos materiais de referência. A disponibilidade de Ni em algumas amostras de solos naturais e tecido vegetal foram inferiores aos limites de quantificação. Os teores desse micronutriente foram maiores em amostras de solo em que houve fornecimento de Ni. A concentração de Ni difere nas partes vegetais analisadas, e os grãos de soja apresentaram os maiores teores. Os grãos, em relação às partes aérea e foliar, têm melhor correlação com os teores disponíveis no solo para ambos extratores. Os métodos descritos nesta pesquisa foram eficientes na quantificação de Ni e podem ser adotados em laboratórios de rotina de solos e tecido vegetal.

Palavras-Chave: micronutriente; disponibilidade; Mehlich-1; DTPA; ICP-OES


Nickel (Ni) is the 23º most abundant element in the Earth’s crust. The total concentration of Ni in soils varies widely, from 5 to 500 mg kg-1, with an average value of 40 mg kg-1 (Liu et al., 2011). Uren (1992) reported that the available amounts only correspond to 0.001 % of the total amounts, and according to Vanselow (1966), most of the time the available levels are lower than 1 mg dm-3. The leaf concentrations of Ni in plants grown in uncontaminated soils are generally between 0.05 and 5 mg kg-1, but most frequently nearer to the lower limit of this range (Brooks, 1980; Welch, 1981). Therefore, Ni can be classified as a trace element in both soils and plants.

The difficulty of quantification due to the lack of devices that are sufficiently sensitive to detect Ni is one of the reasons this element was the last one included in the list of micronutrients. Its essentiality, as demonstrated by Brown et al. (1987) and Eskew et al. (1983, 1984), is due to the fact that Ni is a structural constituent of the enzyme urease, which hydrolytically turns urea [CO(NH2)2] into ammonia (NH3) and carbon dioxide (CO2) (Dixon et al., 1975).

Nickel was recently included in Brazilian regulations on fertilizers by the Normative Instruction Nº 5 of February 23, 2007, issued by the Ministry of Agriculture (Brasil, 2014), but there are so far no official recommendations for its use in fertilization programs. Formulating adequate recommendations depends on understanding the effects of the available levels in agricultural soils and plant tissues, and thus on standardization of suitable methods to measure these levels.

There are various analytic methods to measure Ni concentrations. Among them, the most promising are inductively coupled plasma with mass spectrometry (ICP-MS), inductively coupled plasma with optical emission spectrometry (ICP-OES), atomic fluorescence spectrometry (AFS), X-ray fluorescence (XRF), polarography, voltammetry, flame atomic absorption spectrometry (FAAS), and graphite furnace atomic absorption spectrometry (GFAAS) (Freschi et al., 2000). Among those techniques, some are very expensive and also involve instruments not commonly found in laboratories for routine analyses of soils and plant tissues. Therefore, it is necessary to develop less expensive and more accessible methods for extraction and digestion, to enable Ni quantification in laboratories and provide farmers with well-based recommendations and technical assistance.

The objective of this scientific note is to present optimized methods for Ni quantification in plant tissues and its availability in soils, using extraction and digestion techniques commonly employed in routine laboratory analyses in Brazil.


The study was conducted at the Soil and Plant Tissue Laboratory of Brazilian Corporation of Agricultural Research, National Soybean Research Center, located in Londrina, Paraná, Brazil.

Samples of 14 soils were collected from the 0-20 cm layer (Table 1), as well as samples of soybean plant tissues (Glycine max [L.] Merrill) (grains, shoot and leaves). Some of these soil and plant samples came from experiments under controlled conditions where the plants were treated with Ni doses of 0.0, 0.2, 0.4, 0.5, 0.8, 1.0, and 5.0 mg dm-3 incorporated in the soil, in which a compilation of the mean Ni levels from these experiments was performed (Rodak, 2014).

Table 1 Classification and location of soil samples from the 0-20 cm layer 

Soil classification(1) Location
County State
Latossolo Vermelo-Amarelo distrófico típico (LVAd) Balsas Maranhão
Latossolo Amarelo distrófico típico (LAd) Luiz Eduardo Magalhães Bahia
Latossolo Vermelho distrófico típico (LVd) [1] Primavera do Leste Mato Grosso
Latossolo Vermelho distrófico típico (LVd) [2] Rio Verde Goiás
Latossolo Bruno alumínico típico (LBa) Campo Novo Rio Grande do Sul
Latossolo Vermelho distrófico húmico (LVd) [3] Coxilha Rio Grande do Sul
Latossolo Vermelho distroférrico típico (LVdf) [1] Londrina Paraná
Latossolo Vermelho eutroférrico típico (LVef) Palotina Paraná
Latossolo Vermelho distrófico típico (LVd) [4] Iporã Paraná
Cambissolo Háplico alumínico típico (CXa) Ponta Grossa Paraná
Argissolo Vermelho distrófico arênico (PVd) [1] Umuarama Paraná
Argissolo Vermelho distrófico latossólico (PVd) [2] Paranavaí Paraná
Neossolo Regolítico eutrófico típico (RRe) Diamante do Sul Paraná
Latossolo Vermelho distroférrico típico (LVdf) [2] Ampére Paraná

(1)According to Embrapa (2006). For more details, see Rodak (2014).

The soil and plant tissue samples were dried in a chamber with forced air circulation, at approximately 60 to 65 °C, for 72 h. Dried samples were then ground, sieved, and stored in plastic containers until analysis.

The Ni concentrations in the samples were determined by ICP-OES with a Perkin Elmer Optima 8300 DV spectrometer at a wavelength of 231.604 nm. Calibration solutions were prepared from suitable dilutions of a stock solution containing 1,000 mg L-1 of Ni. Calibration curves were plotted from Ni concentrations of 0.015, 0.050, 0.10, 0.25, 0.50, 1.00, and 2.00 mg L-1 to quantify the availability in the soil, and 0.0075, 0.015, 0.075, 0.15, 0.25, 0.50, and 1.00 mg L-1 for the plant tissue samples. The smaller the concentration range of the calibration curve, the more sensitive will be the determination of low Ni concentrations. Thus, when the samples did not tend to high Ni concentrations, the soil curve was used up to a concentration of 1.0 mg L-1.

Reference samples of soil (BCR® - 142R) and plant tissue (Trifolium repens L. BCR® - 402 and Lichen BCR® - 482), certified by the European Commission - Joint Research Centre, Institute for Reference Materials and Measurements, and grain samples (Vicia fava IPE 903 and Phaseolus vulgaris IPE 192), certified by the Wageningen Evaluating Programs for Analytical Laboratories, International Plant-Analytical Exchange, were used to ensure the quality control of the analyses and adjustment of the methods.

The Pearson linear test was used to calculate the correlations of the available levels of Ni in the soil and concentrations in the shoots, grains and leaf samples, using Statistica 7 software (Stat Soft, 2004).

Soil analysis

Availability of Ni with Mehlich-1 digestion

Extraction method as described by Embrapa (2009). Samples (volume of 5 cm3) were added to 50 mL of Mehlich-1 extractant solution. Then the samples were stirred for 10 min at 200 rpm, after which the suspension was left to rest for 16 h to decant and an aliquot of the sample was submitted to ICP-OES to quantify the Ni content. Mehlich-1 extractant solution consisted of a mixture of hydrochloric acid (HCl 0.05 mol L-1) and sulfuric acid (H2SO4 0.012 mol L-1).

Availability of Ni with DTPA digestion

Extraction method as described by Raij et al. (2001). Samples of a volume of 5 cm3 were added to 50 mL of DTPA extractant solution. Then the samples were stirred for 2 h at 220 rpm, after which the suspension was immediately passed through quantitative filter paper (blue band) and subjected to ICP-OES for determination of Ni content. DTPA solution consisted of a mixture of diethylenetriamine pentaacetic acid (DTPA 0.005 mol L-1), triethanolamine (TEA 0.1 mol L-1) and calcium chloride (CaCl2.2H2O 0.01 mol L-1) at pH 7.3, corrected with a hydrochloric acid solution (HCl 4 mol L-1).

Analysis of plant tissue samples

To determine Ni concentration in the plant parts, the samples were digested with nitric acid (HNO3) in a closed system in a Mars Xpress microwave oven (CEM), with subsequent determination by ICP-OES (Malavolta et al., 1997). To measure Ni concentrations in the plant tissue, the analytical method was optimized to concentrate the extract for determination. For this, 6 mL of HNO3 was added to 0.5 g of ground dry plant matter and the samples were pre-digested for 30 min before being placed in the microwave oven. The heating program was: 10 min to reach 170 ºC, 15 min at 170 ºC, and 20 min of cooling. After the microwave digestion, the samples were diluted to 15 mL with ultra-pure water.


The average recovery values obtained by fitting the method for plant tissue and grains were high, with values between 86.54 to 93.90 % in relation to the certified values for the samples (Table 2), indicating successful Ni determination with adjustment of the method. However, the recovery values for the available levels in the soil were low, averaging 2.06 % when using Mehlich-1 extraction and 0.79 % with DTPA (Table 2). These recovery percentages can be explained by the fact that the method used was adjusted to measure available levels rather than total levels.

Table 2 Average of reference values and recovery of certified samples used to set the methods of nickel quantification 

Sample Ni concentration
Reference value Quantified value Recovery
  Soil (mg dm-3)(1)
BCR® - 142 R 64.5(2) 1.33 / 0.51 2.06 / 0.79
  Shoot (mg kg-1)
BCR® - 402 8.25 7.17 86.91
BCR® - 482 2.47 2.14 86.64
  Grain (mg kg-1)
IPE 903 2.13 2.0 93.90
IPE 192 1.04 0.9 86.54

(1) Available values extracted by Mehlich-1 and DTPA, respectively; (2) Total concentration.

The average concentrations of available Ni in the soil samples varied from <0.10 to 1.39 mg dm-3 with Mehlich-1 extraction and from <0.013 to 0.97 mg dm-3 with DTPA (Table 3). In Ni fertilized soil samples, the element availability increased, to at most 3.24 mg dm-3 Ni with Mehlich-1 and 1.87 mg dm-3 with DTPA. The soil samples LVAd, LAd and LVd [4] without Ni applications contained available levels below the quantification limits of <0.10 and <0.013 mg dm-3 of Ni with Mehlich-1 and DTPA, respectively. This clearly shows the difficulty of quantifying this micronutrient, especially in natural soils.

Table 3 Available levels of Ni extracted by DTPA and Mehlich-1 in 14 soil samples collected from the 0-20 cm layer 

Classification Ni concentration
Minimum Maximum Average Minimum Maximum Average
  mg dm-3
  Without Ni application
LVAd - - <0.10 - - <0.013
LAd - - <0.10 - - <0.013
LVd [1] - - 0.14 - - 0.09
LVd [2] - - 0.27 - - 0.11
LBa - - 0.16 - - 0.08
LVd [3] - - 0.13 - - 0.07
  With Ni application(1)
LVdf [1] 0.44 1.12 0.75 0.18 0.54 0.34
LVef 0.27 3.24 0.81 0.14 1.87 0.44
LVd [4] <0.10 1.78 0.44 <0.013 1.13 0.24
Cxa 0.07 0.43 0.21 0.07 0.37 0.17
PVd [1] 0.13 0.56 0.30 0.09 0.47 0.22
PVd [2] 0.17 0.55 0.30 0.10 0.48 0.25
RRe 0.90 1.41 1.10 0.64 1.37 0.94
LVdf [2] 1.25 1.65 1.39 0.73 1.25 0.97

(1)Rates of 0.0, 0.2, 0.4, 0.5, 0.8, 1.0 and 5.0 mg dm-3 of Ni incorporated into the soil; LVAd - Latossolo Vermelho-Amarelo distrófico típico (Balsas - MA); LAd - Latossolo Amarelo distrófico típico (Luiz Eduardo Magalhães - BA); LVd - Latossolo Vermelho distrófico típico (Primavera do Leste - MT [1] and Rio Verde - GO [2]) or Latossolo Vermelho distrófico húmico (Coxilha - RS [3] and Iporã - PR [4]); LBa - Latossolo Bruno alumínico típico (Campo Novo - RS); LVdf - Latossolo Vermelho distroférrico típico (Londrina [1] and Ampére [2] - PR); LVef - Latossolo Vermelho eutroférrico típico (Palotina - PR); Cxa - Cambissolo Háplico alumínico típico (Ponta Grossa - PR); PVd - Argissolo Vermelho distrófico arênico (Umuarama [1]) and Argissolo Vermelho distrófico latossólico (Paranavaí [2] - PR); RRe - Neossolo Regolítico eutrófico típico (Diamante do Sul - PR). For more details, see Rodak (2014).

To ensure soil quality and prevent problems of food grown on contaminated soils, the National Environmental Council of Brazil (Conama, 2009) set a deadline of 2014 to establish Reference Quality Values (RQV) for potentially toxic elements, including Ni, for each state of the country. Table 4 presents the RQV proposed to date in some Brazilian states. There are no ranges and critical values in agricultural or natural soils regarding available Ni concentrations, evidencing that the analysis of total levels alone is not sufficient, but that policies for available levels must be set.

Table 4 Reference quality values (RQV) for nickel in the Brazilian states 

State RQV Literature
  mg kg-1  
Minas Gerais 21.50 Caires (2009)
Espírito Santo 9.17 Paye et al. (2010)
Rondônia and Mato Grosso 2.10 Santos (2011)
São Paulo 13.00 Cetesb (2005)
Coastal plain of Paraná 17.22 Buschle (2013)
Paraná (B horizon) 17.00 Licht et al. (2006)
Conama(1) 30.00 Conama (2009)

(1)National Environmental Council of Brazil.

Concentrations of Ni in the plant tissue varied from <0.084 to 14.26 mg kg-1 (Table 5). As stated for the available levels (Table 3), plants cultivated in soils without Ni fertilization contained the lowest levels, below the quantification limit of <0.084 mg kg-1. Concentrations in soybean plants were generally higher in the leaves than shoots, but lower than in the grains, with or without Ni fertilization (Table 5), evidencing that plant parts differ regarding Ni levels. The average values found in the plant tissue in this study were within the maximum tolerance range established by the National Sanitary Surveillance Agency (Anvisa, 1965), i.e., 0.1 to 4.0 mg kg-1 Ni.

Table 5 Nickel concentration in plant tissues of soybean 

Plant tissue(1) Ni concentration
Minimum Maximum Average
  mg kg-1
Leaf <0.084 3.02 0.58
Shoot <0.084 1.49 0.33
Grain 0.28 14.26 3.22

(1)Including samples with and without Ni application.

Correlations (r) between the available Ni levels in the soil and in the three plant parts were positive and significant for extraction with Mehlich-1 and DTPA (Table 6). Among the Ni levels in the shoot, leaf and grain, the correlation with the soil concentration was highest in the latter, by both extractants. The less significant correlation for leaf in relation to grains levels can be related to the mobility of Ni in plant tissue. Findings of Cataldo et al. (1978) confirm this assumption. They observed that 70 % of the Ni present in soybean leaves in the senescent state was remobilized from the leaf tissue to the grains.

Table 6 Correlations between Ni in soil (Mehlich-1 and DTPA), leaf, grain and shoot of soybean plants grown in soils with different levels of this micronutrient 

  DTPA Shoot Grain Leaf
Mehlich-1 0.95** 0.10ns 0.91** 0.47**
DTPA - 0.11ns 0.91** 0.53**
Shoot - - 0.90** 0.14ns
Grain - - - 0.47**

Including samples with and without Ni application; ** and ns: significant and non-significant at p<0.01 by the Pearson correlation coefficient.


The tested methods were successful in quantifying Ni and can be used in laboratories for routine analysis of soil and plant tissue samples. There was a positive and significant correlation between the available Ni levels extracted from the soil with Mehlich-1 and DTPA; in the plant tissue the recovered levels were high in relation to the corresponding certified samples.

Nickel availability in some natural soil and plant tissue samples were lower than the quantification limit.

Nickel concentrations were higher in the soil samples that had received Ni fertilization.

Nickel levels differed in the plant parts, and were highest in the grains.

Correlations between Ni levels in the grains and available levels in the soil extracted with Mehlich-1 and DPTA were stronger than the correlations with Ni levels in shoot and leaves.


The authors are indebted to the Graduate Program of Soil Science of the Federal University of Paraná - UFPR; to the Coordination for the Improvement of Higher Education Personnel, CAPES, and to Embrapa Soybean, especially to the technicians Mr. Joviano Kleinert and Ms. Mariana Germano; to the Prof. Dr. Luiz Roberto G. Guilherme for provide samples of certified reference materials; to Dr.a Aline R. Coscione for the suggestions on digestion of plant tissues.


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Received: August 22, 2014; Accepted: January 26, 2015

* Corresponding author.E-mail:

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