Cloud point extraction to avoid interferences by structured background on nickel determination in plant materials by FAAS

Abstract

An analytical procedure based on microwave-assisted digestion with diluted acid and a double cloud point extraction is proposed for nickel determination in plant materials by flame atomic absorption spectrometry. Extraction in micellar medium was successfully applied for sample clean up, aiming to remove organic species containing phosphorous that caused spectral interferences by structured background attributed to the formation of PO species in the flame. Cloud point extraction of nickel complexes formed with 1,2-thiazolylazo-2-naphthol was explored for pre-concentration, with enrichment factor estimated as 30, detection limit of 5 µg L⁻¹ (99.7% confidence level) and linear response up to 80 µg L⁻¹. The accuracy of the procedure was evaluated by nickel determinations in reference materials and the results agreed with the certified values at the 95% confidence level.

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1. Introduction

Metal analyses in plant materials are usually required for agronomical studies. These determinations have often been carried out by spectrometric techniques following closed-vessel microwave-assisted acid digestion,^1,2 with advantages over procedures based on convective heating. Excellent results have been obtained with diluted acids,^1–3 often combined with H₂O₂ as the auxiliary oxidant agent, exploiting the high efficiency of water to convert microwave radiation in heat. This alternative method yields greener procedures by drastic reduction of wastes and allows minimizing the risks of sample contamination and damage to the instrument.

Cloud point phenomenon occurs when aqueous solutions of non-ionic surfactants above the critical micelle concentration become turbid upon modification of temperature or introduction of a suitable additive.⁴ A surfactant-rich phase can then be separated by centrifugation, removing the hydrophobic species from the aqueous solution. This process has been used in trace metal determination by spectroscopic techniques mainly for analyte pre-concentration after reaction with a suitable hydrophobic complexing agent.⁵ However, this strategy also has the potential for separation of components of the sample matrix, as demonstrated previously for sample clean up to the analytical measurement by spectrophotometry,⁶chromatography⁷ or capillary electrophoresis.⁸

Nickel is an essential micronutrient, which can be toxic to plants in high concentrations.⁹ Its determination in plant materials by atomic absorption spectrometry is susceptible to spectral interferences caused by structured background.^10–12 This interference, attributed to the formation of PO species in the atomizer, can not be suitably compensated for by a continuous source background corrector.

In this work, an analytical procedure based on a double cloud point extraction for sample clean up and analyte pre-concentration is proposed for nickel determination in plant materials, following microwave-assisted digestion with diluted acids.

2. Experimental

2.1. Apparatus

An AAS vario 6 flame atomic absorption spectrometer (Analytikjena, Germany) equipped with deuterium background correction and nickel hollow-cathode lamps as radiation sources was used for absorbance measurements at 232.0 nm. The instrumental parameters were adjusted according to the manufacturer's recommendations. Acid decomposition was carried out in closed perfluoroalkoxy (PFA) vessels by using a microwave oven (Anton Paar, Multiwave 3000). A centrifuge (Quimis Q222T, Brazil) was used to speed up the phase separation in the cloud point extraction procedures.

A sliding-bar injector-commutator designed for flow injection analysis¹³ was employed to insert discrete volumes of the extracts in the FAAS nebulizer, as previously described.¹⁴ Measurements were based on peak height and carried out in triplicate.

2.2. Reagents and solutions

All solutions were prepared with analytical grade chemicals and deionized water. A stock 1000 mg L⁻¹ Ni solution was prepared by suitable dilution of Titrisol^® (Merck, Germany). A 2.0 × 10⁻³ mol L⁻¹1,2-thiazolylazo-2-naphthol (TAN, Across Organics, USA) solution was prepared in ethanol. Solutions of the nonionic surfactant Triton X-114 (Sigma, USA) were prepared at the concentration 5% (w/w). Acetate buffer solutions were prepared from 1 mol L⁻¹acetic acid with pH adjusted to 4.8 with sodium hydroxide.

2.3. Procedures

The microwave-assisted acid digestion procedure was carried out in PFA vessels with 250 mg of each plant material and different amounts of HNO₃ and H₂O₂ (30% v/v). The heating program was performed in 4 steps: 1—ramp (140 °C/5 min)/hold (1 min); 2—ramp (180 °C/4 min)/hold (5 min); 3—ramp (220 °C/4 min)/hold (10 min); 4—cooling to 25 °C (20 min).

Sample digests were submitted twice to the cloud point extraction procedure, after partial neutralization of the excess of acid with 2 mol L⁻¹NaOH. For sample clean up, 1 mL of Triton X-114 solution and 1 mL acetate buffer were transferred to 15 mL graduate tubes containing a suitable amount of the digested sample. Phase separation was induced by heating the tubes into a water bath at 60 °C for 15 min. The mixture was then centrifuged for 20 min, using a rotation of 3000 rpm and the aqueous phase was quantitatively transferred to another 15 mL graduate tube. Aliquots of 450 µL of TAN solution, 1 mL of Triton X-114 solution and 1 mL acetate buffer were added for nickel pre-concentration. The phase separation procedure was carried out as previously described, the aqueous phase being removed with a Pasteur pipette. The surfactant-rich phase was then diluted with ethanol to 0.5 mL by using the tube graduation, in order to decrease its viscosity, making feasible the introduction in the FAAS by pneumatic nebulization.

3. Results and discussion

Nickel is usually found in low concentrations in plant materials (usually in the 0.05–5 µg g⁻¹ range) and its determination by FAAS requires pre-concentration. A procedure based on cloud point extraction was developed, exploiting the formation of complex with TAN and extraction with Triton X-114, similar to the procedure previously employed for determination of copper and iron.¹⁴ The analytical response was linear up to 80 µg L⁻¹ nickel, with a detection limit of 5 µg L⁻¹, estimated according to IUPAC recommendations¹⁵ at 99.7% confidence level. The enrichment factor was estimated as 30 and the extraction was quantitative, as evaluated by measurements in the supernatant solution after extraction. The procedure consumes 100 mg Triton X-114 and 150 µg TAN per determination.

In order to minimize waste generation in the sample digestion and to make feasible the pH adjustment required to complex formation and analyte extraction, samples were digested with a mixture of diluted HNO₃ and H₂O₂ peroxide by microwave-heating. Transparent and colorless digests were obtained by using 5 mL HNO₃ 2.8 mol L⁻¹ and 1.1 mL 30% H₂O₂ for a sample mass of 250 mg, employing the heating program previously described (section 2.3.). In previous work, it was estimated for a plant material, by using a similar acid concentration, that the residual carbon content was (11.3 ± 2.0)% and ca. 30% of the acid rested after digestion.²

Inorganic species in the higher concentration expected in digests of plant materials did not interfere in nickel determination (Table 1). However, positive interferences were observed in analyte determination without the clean up step, yielding analytical signals up to twice higher than expected. In addition, it was observed that the digests of certified reference materials (NIST 1547 peach leaves and NIST 1515 apple leaves) presented a yellow color after pH adjustment to 4.8 (absorbance signals measured by UV-vis spectrophotometry at 380 nm were higher than 1.2 for a 10-fold diluted sample digest), indicating incomplete sample decomposition. Acid digests of plant materials can contain residual amounts of organic phosphorus compounds—in the evaluated samples, the phosphorus amount may reach concentrations as high as 0.4% (w/w). The presence of phosphorus species in the samples can cause spectral interferences, due to generation of structured background,^10–12 usually related to the formation of PO species in the flame, uncorrectable with the deuterium source employed in the FAAS measurements. Similar interference has been observed in nickel determination by GFAAS.^10–12 Hydrophobic organic compounds containing phosphorus can also be extracted to the surfactant-rich phase, then generating spectral interferences due to molecular absorption caused by the PO species in the same wavelength recommended for nickel determination. This hypothesis was confirmed by a study carried out with 50 µg L⁻¹ Ni solutions, in the absence and presence of 50, 150 and 200 mg L⁻¹phytic acid, which is usually found in vegetables and difficult to decompose by acid digestion procedures. Interferences higher than 50% were observed when solutions containing phytic acid were submitted to the cloud-point pre-concentration procedure without the clean up step. A similar study was carried out in the presence of 1 mmol L⁻¹inorganic phosphorus (PO₄³⁻). However, significant differences in the analytical signal were not observed because these hydrophilic species were not extracted to the surfactant-rich phase.

Table 1 Effect of concomitant species on the determination of 25 µg L⁻¹ nickel by cloud point extraction

Element	Concentration/µg L^−1^a	Signal variation (%)	Element	Concentration/µg L^−1^a	Signal variation (%)
a Defined from the maximum concentration found in plant certified reference materials.
Al	9 × 10^3	+2.1	Mn	4 × 10^3	+1.3
Ca	8 × 10^5	+4.6	Ni	26	__
Cd	25	+4.7	P	1 × 10^4	+0.6
Co	9	+2.5	Pb	14	+4.8
Fe	6 × 10^3	+3.9	V	14	+3.3
Hg	0.7	+4.9	Zn	500	+3.2
Mg	2 × 10^4	+4.9	Cu	95	+4.0

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A clean up step based on cloud point extraction without addition of TAN was included for the preliminary removal of the organic matter remaining after acid digestion. Adopting this procedure, the species that caused a yellow color in the samples were completely removed to the surfactant rich phase that was further discarded. The aqueous phase was then submitted to the cloud point preconcentration of the analyte with the addition of the complexing agent. The certified reference materials were then analyzed by FAAS and the results (Table 2) were in agreement with the certified values at the 95% confidence level, indicating the efficiency of the clean up step for separation of the interfering species.

Table 2 Results of nickel determination in certified reference materials by FAAS after clean up and pre-concentration by cloud point extraction

Certified reference material	Ni/µg g^−1
	Certified value	Proposed procedure
Peach leaves NIST 1547	0.69 ± 0.09	0.74 ± 0.04
Apple leaves NIST 1515	0.91 ± 0.12	1.03 ± 0.29

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4. Conclusions

Organic species containing phosphorous that can not be effectively decomposed by microwave-assisted sample digestion with diluted acids can generate PO in flame atomizers and cause spectral interferences by structured background in nickel determination. A sample clean up step, based on cloud point extraction, is a fast and simple alternative to remove the organic phosphorus compounds avoiding the spectral interferences. The enrichment factor achieved by the quantitative extraction of nickel complexes formed with TAN allowed the analyte determination by FAAS.

Acknowledgements

The authors acknowledge the fellowships and financial support from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP).

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