Ingeniatrics

ICP-MS and ICP-OES Analysis of Multi-Elemental Profile of Red and White Wines using MultiNeb® nebulizer

1. Introduction

Wine is one of the most popular alcoholic beverages in the world. Wine is a complex matrix and, it contains low-level concentrations of mineral elements in major (Ca, K, Mg, and Na), minor (Fe, Cu, Mn and Zn) and trace levels (As, Cd, Co, Cr, Ni, Pb, and Hg). The elemental composition of wines and sparkling wines depends on natural and anthropogenic sources. Endogenous elements come from the absorption of minerals from the soil and can be found in different parts of the plant, including the grapes. Since the composition of the soils from the vineyards, usually calcareous and clayey, grapes are rich in metals like Ca, Mg, Fe, Al, or Mn. Anthropogenic elements come from agricultural practices in the vineyard as well as from winemaking processes. In this sense, wine elemental composition varies by cultivar, geographic origin, viticultural and enological practices, and is often used for authenticity validation. Elemental analysis of wine by Coupled Plasma Optical Emission Spectroscopy (ICP-OES) or Mass Spectrometry (ICP-MS) is challenging due to the potential for non-spectral interferences and plasma instability arising from organic matrix components. ICP-MS and ICP-OES analysis of wines using MultiNeb.

Several elements, such as As, Cd, Cr, Hg and Pb, can have adverse health effects in humans even at low concentrations (<100 µg/L), while others (i.e., Cu) are essential nutrients that only become toxic at levels greater than typically encountered in the human diet, or are of importance for wine stability or development and other elements, such as Cu, Fe, Ni, Zn related with haze formation causing changes of taste and smell properties.

In addition, minerals can contribute to stability and clarity in the wine and its color, and they may affect the organoleptic characteristics of the wine, mainly Zn and, or wine conservation, i.e. precipitation of K and Ca tartrates.

Wine elemental analysis is usually performed using atomic absorption and emission spectroscopy techniques such as Flame Atomic Absorption/Emission Spectroscopy (FAAS and FAES) and Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) or Mass Spectrometry (ICP-MS). ICP-MS reports of sub-parts per billion detection limits, multielement analysis capabilities, and a wide linear dynamic range have made it an often-used technique for wine elemental analysis. However, elemental determination in wine, via plasma sources, presents a challenge analytically because of non-spectral interferences by organic matrix components, primarily ethanol when present in concentrations from 7 to 20 %. In this sense, direct analysis of wine by ICP plasma efficiency and sample transport thus causing perturbations in signal intensity and stability. Modifications on the instrumental configurations, e.g., using a collision reaction cell (CRC) technology in combination with matrix-matched calibration are used to mitigate the effects and allow for direct wine analysis by ICP-MS. Collision/reaction cell technology (CRC) has been demonstrated to reduce the effects of spectral interferences in elemental analysis of wine by ICP-MS and allowed for the application of lower dilution factors.

Wine

For the determination of trace and ultra-trace elements from ppq to ppb levels, undoubtedly inductively couple plasma mass spectrometry (ICPMS) is the most powerful among atomic spectrometry techniques due to its high sensitivity, low detection limits and multi-element detection capability. Major ions of Ca, K, Mg and Na at ppm levels in wine are exceeding the linear dynamic range of the ICP-MS instruments which employ electron multiplier (EM) detector, preventing their direct measurements. Moreover, polyatomic interferences arising from sample matrix such as 12C12C+ and 48Ca2+ on 24Mg+, 12C13C+ on 25Mg+, 38Ar1H+ and 23Na16O+ on 39K+ and 12C16O16O+ on 44Ca+ encountered in ICP-MS need to be resolved or removed for their accurate determination.

For this purpose, the combination of Inductively-coupled plasma mass spectrometry (ICP-MS) and Inductively-coupled plasma optical emission spectrometry (ICP-OES) provides extensive capability for food control and guarantee a high level of quality.

Here we describe a simple, quick and robust method for elements in red and white wine using the multiple inlet nebulizer MultiNeb® (Ingeniatrics Tecnologías S.L.) by a simple online dilution for small volume of sample and ICP-OES in combination with ICP-MS analysis, equipped with CRC was used to reduce polyatomic interferences and internal standards were used to counter the matrix effects and instrument drift. Conventionally, the internal standard is mixed with the calibration standards and samples using a Y connection, when conventional nebulizer is employed. However, the novel MultiNeb® (Ingeniatrics Tecnologías S.L.) has been developed which allows a high mixing efficiency between two liquids, miscible or immiscible, since the mixing takes place under turbulent conditions of high pressure at the tip of the nebulizer.

2. Experimental

Reagents and solutions

All reagents used were of the highest available purity. Helium was used as collision gas, respectively, in an ICP-MS system, were of high-purity grade (>99.999%). Water was purified with a Milli-Q Gradient system (Millipore, Watford, UK). The aqueous levels of calibration standards were prepared by appropriate dilution of a Multi-element calibration standard-2A, contains 10 mg.L-1 of Ag, Al, As, Ba, Be, Ca, Cd, Co, Cr, Cs, Cu, Fe, Ga, K, Li, Mg, Mn, Na, Ni, Pb, Sb, Se, Sr, Tl, U, V, Zn (and Hg) in a matrix of 5% HNO3 (Agilent Technologies, Part Number: 8500-6940). All aqueous solutions are acidified using 5% (v/v) high purity nitric acid (Suprapur®, Merck, Darmstadt, Germany). As internal standard, a mixture of Scandium (Sc), Rhenium (Re) and Rhodium (Rh) was employed for ICP-MS analysis and Yttrium (Y) was employed for ICP-OES analysis. High purity ethanol and hydrogen peroxide (Aldrich, 99.999% trace metals basis) and high purity D-glucose (>99.9%, Merck) as a matrix-matching component were employed.

Instrumentation

A heating block system type DigiPREP MS (SCP Science, Courtaboeuf, France) was employed for sample preparation. All measurements were carried out using an Agilent 7900 ICP-MS (Agilent Technologies) and Agilent 5110 ICP-OES (Agilent Technologies) equipped with a SPS4 autosampler (Agilent Technologies) with 0.5 mm ID sampling probe (Agilent Technologies, Part No.: G8410-80101). The instruments operating conditions are shown in Table I. 

The choice of an appropriate calibration method is critical for the compensation of these matrix effects for obtaining accurate results. Conventionally, the internal standard is mixed with the calibration standards and samples using a Y connection, when OneNeb® nebulizer in employed. However, the novel MultiNeb® has been developed which allows a high mixing efficiency between two liquids, miscible or immiscible, since the mixing takes place under turbulent conditions of high pressure at the tip of the nebulizer.

The MultiNeb® nebulizer used in this study consists of two independent liquid inlets and a common gas inlet in a single nebulizer body of polytetrafluoroethylene (Figure 1). All peristaltic pump tubings employed were of 0.25 mm i.d., normally used for ISTD solutions (blue/orange taps).

Additionally, it was demonstrated that the spectralinterferences caused by rich-organic matrices can be eliminated by MultiNeb® nebulizer. Traditionally, the carbon deposition in cones, can be avoided by adding a small amount of oxygen to the intermediate gas flow. However, the use of oxygen to support complete combustion of carbon further increases the complexity of the experimental setup and the cost per analysis.

Multineb nebulizer

Figure 1. MultiNeb® nebulizer

Related to this, adding aqueous solution in the aerosol, through MultiNeb’s second channel, will help the complete combustion of carbon, avoiding its byproducts and the subsequent clogging of the torch’s injector, cones and will, therefore prevent the spectral interferences, providing a better sensitivity, reproducibility and precision. In this sense, in this study a solution with internal standards in 20 % (v/v) of hydrogen peroxide was employed to obtain better background in analytes interfered by carbon matrix, by increasing the contribution of oxygen in the aerosol compared to water, favoring the formation of CO2 and reducing carbon deposition in torch’s injector and cones, and reducing carbon interferences in plasma. In addition, this solution contains the internal standards with order to evaluate the long-term stability of the signal and matrix effects.

Operational conditions for 7900 ICP-MS and 5110 ICP-OES

Table I. Operational conditions for 7900 ICP-MS and 5110 ICP-OES using online internal calibration

The MultiNeb®-based configuration is composed by the MultiNeb® nebulizer associated with a spray chamber without any additional modification required, as the MultiNeb® is built on the right dimensions to allow easy connection to any commercial spray chamber conventionally used in ICP-based (Figure 2). This implies an important advantage over conventional systems since it does not require the continuous cleaning of ICP components or the use of expensive additional components such as cooled spray chambers or an auxiliary oxygen supply. This simple and powerful alternative to remove spectral interference caused by organic matrix enables to analyze organic samples with confidence.

Schematic representation of MultiNeb®-

Figure 2. Schematic representation of MultiNeb®-based configuration employed in this Application Note.

Matrix-matched calibration and samples

Several calibration strategies have been employed for elemental wine analysis in ICP-based techniques for wine samples. Among them, external calibration using aqueous, and matrix matched standards and internal standardization were evaluated in the present work since both methodologies are the most convenient for multi-elemental analysis: they are easy to apply, consume a low sample volume and the analysis throughput is high.

Interferences due to wine  matrix components in ICP techniques can be divided in two groups: spectral and non-spectral. The former are related to the limited resolution capability of the spectrometer. Thus, for instance, the analysis of Se in carbon-rich matrices using ICP-OES is interfered by the CO molecular band. In ICP-MS, the analysis of Cr and Cu using the most abundant isotopes (i.e. 52Cr+ and 63Cu+, respectively) in matrices with high levels of carbon and/or sodium could be hindered due to the 52ArC+ and 63ArNa+ polyatomic interferences. Non-spectral interferences are defined as any signal variation induced by the matrix components.

Thus, the signal in both ICP-OES and ICP-MS could be enhanced or depressed when carbon or easily ionisable elements are present in the matrix. These interferences are usually generated in the sample introduction system and/or in the processes of excitation of the atoms in ICP-OES and the transport of the ions in ICP-MS. The analysis of wine samples is usually performed by means of matrix matched standards but, when matrix effects are strong, alternative approaches such as standard addition and internal standardization give rise to more accurate results.

In this sense, wine samples were analyzed using ICP-MS and ICP-OES using different amounts of ethanol 10-14 % (v/v) and glucose (1 – 4 g.L-1) as a major matrix components. In addition, internal standardization was applied to evaluate the efficiency in signal response. Table II summarized the total content in ethanol and sugars containing in wines employed in this study.

Spanish Wines samples

Table II. Wine samples analyzed in this study

A total of three internal standards have been used depending on the mass of the isotope in ICP-MS and yttrium as internal standard in ICP-OES. The Table III summarizes the internal standard employed for each analyte throughout this work.

Internal standard using ICP-MS and ICP-OES.

Table III. Internal standard used for each analyte throughout this work using ICP-MS and ICP-OES

The best results were obtained using 12 % (v/v) of ethanol and 2 g.L-1 of glucose, as a matrix matched components, in terms of sensitivity, reproducibility and precision in recoveries results for each analyte quantified using ICP-MS and ICP-OES analytical techniques.

Sample preparation

Figure 3 shows the workflow followed for standard calibration levels, QC samples, blanks and samples preparation for elemental determination using ICP-MS and ICP-OES.

Brieftly, 10 mL of standard calibration solutions, QC samples and blanks prepared in ethanol 12% (v/v) and glucose 2 g.L-1 and 10 mL of white and red wines samples were incubated with 2 mL of HNO3 65% (w/w) in a Digiprep digestor. Then, resulting solutions were diluted with ultrapure water to 25 mL and divided in two aliquots of 10 mL. One of this solution was directly analyzed by ICP-OES, and the other one was diluted two times with 5% HNO3 for ICP-MS analysis.

3. Results and Discussion

Sensitivity and signal stability

Nebulizers designed by Ingeniatrics Tecnologías SL, such as MultiNeb®, use Flow Blurring nebulization technology instead of the traditional Venturi effect, as conventional concentric nebulizer. This allows the generation of a very fine droplet aerosol with a narrow size distribution (most droplets are smaller than 10 μm), which improves efficiency over a wide range of nebulization gas flow rates, especially 0.60-0.75 L min-1 (150-250 kPa nebulization pressure).

The method detection limits (MDLs) were established by analyzing five of the calibration blank and multiplying the obtained standard deviation by three. The results obtained are show in Table IV

The ICP-CRC-MS was optimized for the removal of all common polyatomic overlaps using helium (He) collision mode and kinetic energy discrimination (KED).

When high matrix samples are measured using ICP source, the matrix elements can affect analyte signals in several distinct ways. Probably the most widely recognized is the gradual downward drift that typically occurs due to the build-up of matrix deposits on the ICP-MS interface components (sampling cone, skimmer cone and torch injector) when high matrix samples are measured over an extended period. In this study, following the workflow for sample preparation previously described, standard calibration levels, QC samples, blanks and samples preparation were only diluted 1:2.5 for ICP-OES analysis and 1:5 for ICP-MS analysis. After this treatment, resulting digested solutions are directly aspirated for one channel of MultiNeb® nebulizer, using peristaltic pump tubings with 0.25 mm i.d., normally employed for ISTD solutions, which in turn is diluted with the solution aspirated through the second channel of the nebulizer containing internal standards solutions in 20 % H2O2 (v/v) to correct instrument drift. In this sense, the resulting aerosol is diluted two times prior to plasma, reducing the carbon content and benefiting the formation of carbon dioxide.

On the other hand, elements such as Ca, K, Na and Mg, occurring at concentrations in the order of magnitude of mg L−1 and higher, could be better measured by ICP-OES since this technique was less affected by matrix effect while the rest of elements should be determined by ICP-MS. As a result, the combination of both techniques in a complementary way for profiling trace and sub-trace elements was the option of choice for further quantitative analysis.

Workflow used for sample preparation

Figure 3. Workflow used for sample preparation.

Multineb nebulizer experimental values

Table IV. Experimental values for each analyte monitored, LOD as well as the SD and recoveries results obtained for 5 replicates of the different samples analyzed using MultiNeb® nebulizer employed in this work by ICP-OES and ICP-MS detection. ND: Not detected. LODs are expressed in µg.L-1 and mg.L-1 for ICP-MS and ICP-OES, respectively.

For the study of signal stability and plasma drift, a monitoring standard solution as quality control (QC) was analyzed once every 5 samples, in order to evaluate the stability of the signal. The recoveries must fall within the limits of 97-103 % and 98-101 % for ICP-OES and ICP-CRC-MS, respectively (Table V). The concentration of the different elements in QC sample using as a monitoring control solution is shown in Table V.

Multineb nebulizer experimental values

Table V. Experimental values for recoveries results each analyte in QC sample analyzed for 5 replicates of the different samples using MultiNeb® nebulizer employed in this work by ICP-OES and ICP-MS detection in a sequence of 50 samples in the same analytical batch. ND: Not detected.

Based on the results shown in Table V, the use of the multiple inlet nebulizer, MultiNeb®,  which allows the simultaneously nebulization of two liquid flows, it demonstrates the long signal stability along the sequence in the same analytical batch. Because of a supplementary addition of oxygen in plasma, as a hydrogen peroxide solution, a resulting aerosol is diluted two times prior to plasma, reducing the carbon content and benefiting the formation of carbon dioxide is minimized.

Precision and reproducibility

As certified reference materials were not available, precision was evaluated based on recovery assays, spiked twice time the concentration of each analyte in each sample matrices employed in this study. The results are shown in Table IV. Additionally, each sample was analyzed using 5 replicates,  and the results obtained, expressed as standard destination (SD) in the same table, demonstrates the notable reproducibility obtained using the proposed methodology in this technical application note for multielemental quantification of 24 elements in wine matrices.

4. Conclusions

The results obtained in this study using MultiNeb® nebulizer provides higher precision, sensitivity and reproducibility for multielement determination in wine samples by ICP-MS and ICP-OES simultaneously determination.

On the other hand, the enhanced precision results obtained with the MultiNeb® nebulizer are related to the higher sensitivity and reproducibility obtained in comparison with the Y connection normally employed for this purpose, what demonstrates that the mixing of the internal standard dissolved in hydrogen peroxide solution, as a supplementary oxygen supply when carbon matrices are analyzer using plasma source, minimizes the effects on the nebulization process and therefore improves the analytical operation and results, , favorizing the complete combustion of carbon, avoiding its byproducts and the subsequent clogging of the torch’s injector, cones and will, therefore prevent the spectral interferences, providing a better sensitivity, reproducibility and precision. Additionally, the proposed analytical methodology prolongs the life time of torches compared to the direct supply of oxygen to the plasma.

Finally, we concluded that elements such as Ca, K, Na and Mg, occurring at concentrations in the order of magnitude of mg L−1 and higher, could be better measured by ICP-OES since this technique was less affected by matrix effect while the rest of elements should be determined by ICP-MS. As a result, the combination of both techniques in a complementary way for profiling trace and sub-trace elements was the option of choice for further quantitative analysis and validation of the results obtained using both analytical techniques.

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