Deborah K. Bradshaw

Using internal standards is a common technique to correct for variations in sample matrices and the affect this has on analyte intensities. There are several basic criteria to be considered when using internal standards: selection of appropriate internal standards, the concentration added to the solutions analyzed, setting up in the correct view (axial vs. radial), how to introduce the internal standard to the solutions to be analyzed, and evaluating the resulting data. Each of these topics are considered and suggestions presented.

In the analysis by plasma techniques, the percentage of each solution analyzed that reaches the plasma can vary due to differences in sample matrices, particularly when compared to simple standards. When this occurs, sample results can be incorrect. Compensating for this problem can be done by matrix matching by preparing calibration solutions in the same type of matrix as the samples; by using the method of standard additions for calibration; or by using internal standards as part of the method. Note that matrix matching and method of additions may not always be practical.

 are elements that are added to all analytical solutions. Their intensities are monitored during analysis. As intensities fluctuate in each solution, most instrument software can correct the readings of the analytes to be determined and report percent recoveries of the internal standards. The use of internal standards has become an integral part of most methods for analysis by plasma techniques for obtaining accurate data. While this has become a common correction technique, the analyst must be aware of several key points to consider when using internal standards.

When selecting elements to be used as internal standards, there are a few basic rules. First, internal standards are elements that don’t need to be determined. Second, they have to be elements that won’t be found in any measurable concentration in any of the samples. Third, be sure the internal standards won’t spectrally interfere with the elements to be determined, or that the other constituents of the samples won’t spectrally interfere with the internal standards. Fourth, internal standards should never be elements that are common environmental contaminants, even if these elements are not in any of your samples. Both yttrium (Y) and scandium (Sc) are chosen for many applications following the above criteria, though they may not be the correct choices for all types of analyses. While choosing internal standards for most types of samples is generally not difficult, for the analysis of some types of samples, selection can be challenging, and may require more rigorous method development.

The actual concentration of the internal standard should be based on the intensity produced by the internal standard. There should be enough intensity from the internal standard that precision of the replicates is optimum (better than 2% relative standard deviation [RSD] in calibration solutions). Poor precision should not play a major role in the correction of the analyte readings. The correction itself is a linear function, so the maximum intensity of the internal standard must be in the linear range for the wavelength chosen.

Setting up the Internal Standard in the Correct “View” 

Since many inductively coupled plasma–optical emission spectroscopy (ICP–OES) devices have dual viewing capabilities, it is important to note that the internal standards must be in the same “view” as the analytes to be determined. If the analyte is viewed axially (down the plasma), the internal standard must be in that view also. If the analytes are viewed radially (across the plasma), so must the internal standard be viewed. This might require the use of multiple internal standards if the method has some elements in the axial view and some in the radial view.

How the internal standard will be added to each solution is of key importance. Concentrations of the internal standards must be the same in all analytical solutions in order for the correction to be accurate. Addition can be made manually by the analyst or automated using either another channel on a peristaltic pump or a valve system to introduce the internal standard to the analytical solutions. Both addition techniques have advantages and disadvantages, as shown in Table I. Whichever way is chosen, evaluation of the resulting data is essential.

There are three areas to consider when evaluating data:

While variations of internal standard recoveries from solution to solution is normal, samples with excessively low or high recoveries should always be investigated before reporting data. Excessive changes of the internal standard intensities may indicate that it was not added correctly to that solution, whether the addition is done manually (due to pipetting or analyst errors) or automated (due to improper mixing). If the recovery in a sample is very high, this may also indicate the internal standard was in the original sample. Some regulatory agencies suggest internal standard recoveries of samples should be within 20% as compared to calibration solutions. While perhaps a good rule of thumb, the actual recovery acceptability can be analysis-specific. Once established, if the internal standard recovery is out of the expected range, it is always a good idea to view the spectral data first to determine if a spectral interference has occurred.

Paying close attention to the precision of internal standard replicates is critical and can be more important that the actual recovery. RSDs of the internal standard replicates greater than 3% should be further investigated as this can cause incorrect analyte results. Poor precision on the internal standard replicates can indicate poor mixing, so this is particularly important when using automated mixing.

Concentrations greater than 1% of easily ionized elements in samples can reduce the ion population and enhance the atom population of analytes to be determined when compared to the calibration solutions. This can lead to incorrect results for some elements, depending on the analyte wavelengths chosen and the internal standards used. This effect is more pronounced in the axial view. One remedy for this is to reduce the sample matrix by diluting the sample, thus reducing the concentration of the easily ionized elements. This is only appropriate if the concentrations of the analytes are still sufficient to be reportable after dilution. Another approach includes optimizing the instrumental parameters (gas flows and sample introduction parameters) (1). A third is to add an ionization buffer to all solutions, which adds an excess of an easily ionized element to all analytical solutions, including standards. A fourth approach is to use internal standards that mimic the wavelengths for the elements of interest. If ion wavelengths are used for the elements, use an ion wavelength for the internal standard. If the wavelength for the analyte is an atom wavelength, use an internal standard with an atom wavelength. Table II shows some comparative data for cadmium (Cd) at two wavelengths, one atom and one ion, in a 2% sodium chloride (NaCl) solution. Four different internal standards were investigated: Y371 and Sc361, both ion wavelengths, and Ge265 and Ga417, both atom wavelengths. When no internal standard correction is made, Cd recoveries are very low for both wavelengths. For Cd228, an atom wavelength, germanium (Ge) and gallium (Ga) provide the most accurate recoveries. For Cd214, an ion wavelength, Y and Sc provide the most accurate recoveries. It may be necessary to use multiple internal standards when the sample matrix has more than 1% total dissolved solids in order to obtain the most accurate results.

Using internal standards for analysis by ICP-OES is an essential part of good method development for most types of samples, but the analyst must be aware of several points in order to insure the reporting of accurate data.

(1) Mermet, J. M. Use of Magnesium as a Test Element for Inductively Coupled Plasma Atomic Emission Spectrometry Diagnostics. 

Identification and Correction of Interferences in Practical ICP-OES

. Short course, Winter Plasma Conference 2022.

 is an analytical chemist who has been working the field of atomic spectroscopy for over 35 years. For the past 20 years, she has been working as a consultant in the field of atomic spectroscopy, conducting training classes and giving technical support for atomic absorption (AA) spectroscopy, inductively coupled plasma–optical emission spectroscopy (ICP–OES), and inductively coupled plasma–mass spectrometry (ICP–MS). Direct correspondence to: 

Articles

Using internal standards is a common technique to correct for variations in sample matrices and the effect this has on analyte intensities. There are several basic criteria to be considered when using internal standards: selection of appropriate internal standards, the concentration added to the solutions analyzed, setting up in the correct view (axial vs. radial), how to introduce the internal standard to the solutions to be analyzed, and evaluating the resulting data. Each of these topics are considered and suggestions presented.

Key Points to Remember When Using Internal Standards for Sample Analysis by ICP-OES 

A common misconception among users of inductively coupled plasma optical emission spectrometry (ICP–OES) is that good “spike” recoveries automatically indicate that the original sample results are accurate. Another common misconception is that the use of method of standard additions (MSA) as a means of calibration to correct for interferences will also always produce accurate results. This tutorial shows that neither of these techniques will produce accurate results if the interference is from spectral overlaps that have not been corrected by other appropriate means.

	When using any atomic spectroscopy technique for elemental analysis, it is easy to obtain results. Validating that the results are accurate is not always so easy. There are many practices used to help validate that results are accurate. The analysis of calibration verification solutions, reference materials, interference check solutions, samples prepared in duplicate, and analyte addition techniques are some ways used to verify the data is accurate.

	There are two types of analyte addition techniques commonly used in atomic spectroscopy. The first is to fortify a portion of a sample with a known additional concentration of analyte, or what is termed to “spike” the sample. There are also times when, instead of using simple calibration standards and reading the analyte concentration from the calibration curve, a technique of method of standard additions (MSA) is used. In this scenario, the calibration curve is achieved in the presence of the sample matrix.

	There are three basic types of interferences that can be experienced using this inductively coupled plasma optical emission spectrometry (ICP-OES): 1) physical interferences that have to do with how much sample is transported into the plasma with each solution analyzed; 2) interferences from the sample matrix having an effect on the population of excited atoms and ions within the plasma; and 3) spectral interferences where something other than the analyte of interest produces spectra different from the plasma at the measured wavelength. Acceptable spike recoveries (typically 85% to 115% in a real sample matrix) is a good indicator of physical and matrix interferences. Using MSA can help alleviate both of these problems. What neither of the techniques properly addresses are spectral interferences.

	Unfortunately, a common mistake is believing that achieving good “spike” recovery or obtaining results using MSA is a guarantee for reporting accurate results. While obtaining poor spike recoveries is an indicator of results that are probably not accurate, good recoveries do not guarantee compensation for all types of interferences. While using MSA rather than simple calibration can compensate for some types of interferences, it does not compensate for all interferences for the same reason. Neither of these techniques will indicate or compensate for spectral interferences.

	To show the problem, the determination of phosphorus in the presence of a high concentration of Cu is used as an example. Two very commonly used phosphorus wavelengths, 213.617 and 214.914, both suffer from overlaps from nearby copper wavelengths, 213.597 (and 213.599) and 214.898, respectively. A third wavelength, 177.434, also will show overlap from Cu 177.427. The P 178.221 wavelength does not suffer from this same problem. In this study, the P 178 wavelength is used as a control.

	A method was set up for the determination of P at the four wavelengths cited above. Calibration was achieved using standards prepared in 1% HNO

 at 5, 10, and 20 mg/L P. All calibration curves had correlation coefficient of 0.998 or 0.999 with linear regression type curve. A solution containing 10 mg/L P and 200 mg/L Cu was analyzed. A portion of this sample was then spiked with an extra 10 mg/L P, and analyzed.

	MSA was then used for sample analysis. Four portions of the sample were spiked with the calibration blank and the three standards, 5, 10, and 20 mg/L, to achieve calibration. Correlation coefficients were again 0.998 and 0.999 for the wavelengths used.

	Generally, acceptable “spike recoveries” are considered to be within 15%. When the sample and its spike were analyzed, the recoveries were calculated. The results are shown in Table I. Calculated spike recoveries for all the wavelengths fall within acceptable (15%) criteria. However, the only wavelength to show correct results for the known concentration of 10 mg/L is P 178.221. The results for the sample when using MSA are also shown in this table. When using MSA, that is also the only wavelength to show correct results.

	The problem is that Cu provides a spectral interference on all wavelengths except the P 178, contributing to the P concentration, and this has not been compensated. Figure 1 shows examples of the spectral peaks for the unknown at the four wavelengths. Only P 178.221 is free from the spectral overlap.

	To correct for the spectral overlaps, interelement correction was applied for the P 213, P 214, and P 177 wavelengths. Simple calibration was once again used and not MSA. Table II shows the corrected data for these wavelengths. Now all wavelengths are in concentration agreement of the known value of 10 mg/L, as well as good recoveries of the fortified portion of the sample.

	It is important for the analyst to use all quality control tools when analyzing sample with complex matrices. Good spike recoveries and using MSA alone may end up in the reporting of inaccurate data. Analyte addition techniques are helpful for compensation of physical and matrix issues, but are not sufficient for correcting for spectral interferences. Viewing the spectral data is a good troubleshooting tool to determine if these problems exist and need further correction for the wavelengths chosen.

is an analytical chemist who has been working the field of atomic spectroscopy for over 35 years. For the past 20 years, she has been working as a consultant in the field of atomic spectroscopy, conducting training classes and giving technical support for atomic absorption (AA) spectroscopy, inductively coupled plasma-optical emission spectroscopy (ICP-OES), and inductively coupled plasma-mass spectrometry (ICP-MS). Direct correspondence to: 

Contrary to popular belief, neither good “spike” recoveries nor the use of the method of standard additions (MSA) will guarantee accurate results in ICP-OES.

Deborah K. Bradshaw

Articles

Key Points to Remember When Using Internal Standards for Sample Analysis by ICP-OES

Influence of Spectral Interferences on the Reliability of Data When Using Analyte Addition Techniques with ICP-OES