Online Standard Additions Calibration of Transient Signals for

An online standard additions calibration method for transient signals in ... and the precision achieved was comparable to that achieved by the certify...
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Anal. Chem. 2007, 79, 688-694

Online Standard Additions Calibration of Transient Signals for Inductively Coupled Plasma Mass Spectrometry Margaret Antler,† E. Jane Maxwell,‡ David A. Duford,† and E. D. Salin*,†

Department of Chemistry, McGill University, 801 Sherbrooke Street West, Montreal, QC, Canada H3A 2K6, and Department of Chemistry, University of British Columbia, 6174 University Boulevard, Vancouver, BC, Canada V6T 1Z3

An online standard additions calibration method for transient signals in ICPMS is demonstrated in which a small volume of standard is injected as a spike into the sample/carrier stream, overlaying the analyte peak. This technique provides the advantages of conventional standard additions but requires only a single sample run. The method corrects for matrix effects and is suitable for transient signals in which the severity of the matrix effect changes over the analyte peak. The method uses a peakfitting program to determine the area of the underlying peak and is shown to be effective for the determination of trace metal concentrations in both a high ionic strength matrix and in a biological matrix (urine). Eight analytes with concentrations in the range of 0.82-233.2 µg L-1 in urine were simultaneously determined using a standard spiking solution of 75 µg L-1 injected through a 100-µL loop. The measured concentrations for analytes free of spectral interferences agreed with the certified values, and the precision achieved was comparable to that achieved by the certifying agency. Using a conventional cross-flow nebulizer and Scott-type spray chamber, the accuracy obtained for online standard additions calibration was within 2%, and the precision was within 5%. Due in large part to the emerging fields of metallomics and metalloproteomics,1-4 many new techniques have been developed recently for studying metal species in biological and environmental systems. One of the most important tools employed is inductively coupled plasma mass spectrometry (ICPMS), often used in combination with separation techniques5-12 such as liquid chromatography (LC) or capillary electrophoresis, which are used to * To whom correspondence should be addressed. E-mail: eric.salin@ mcgill.ca. † McGill University. ‡ University of British Columbia. (1) Haraguchi, H. J. Anal. At. Spectrom. 2004, 19, 5-14. (2) Szpunar, J. Anal. Bioanal. Chem. 2004, 378, 54-56. (3) Szpunar, J. Analyst 2005, 130, 442-465. (4) Sanz-Medel, A.; Montes-Bayo´n, M.; Luisa´ Ferna´ndez Sa´nchez, M. Anal. Bioanal. Chem. 2003, 377, 236-247. (5) Kolbl, G. Mar. Chem. 1995, 48, 185-197. (6) Michalke, B. Trends Anal. Chem. 2002, 21, 142-153. (7) Michalke, B. Trends Anal. Chem. 2002, 21, 154-165. (8) Montes-Bayon, M.; DeNicola, K.; Caruso, J. A. J. Chromatogr., A 2003, 1000, 457-476. (9) Thompson, J. J.; Houk, R. S. Anal. Chem. 1986, 58, 2541-2548.

688 Analytical Chemistry, Vol. 79, No. 2, January 15, 2007

separate the analytes according to their chemical forms before online coupling to ICPMS. Compared to softer ionization sources such as electrospray ionization and MALDI, where the degree of ionization depends on the nature of the analyte, ionization in the plasma source is virtually independent of molecular form,3 allowing for quantitative analysis without the need for additional treatments. As a detection system, ICPMS offers low detection limits, a large linear range, and multielement capabilities. When ICPMS is interfaced to a separation system, the signal measured is usually transient, which can lead to a more complex calibration procedure. The calibration of the various chemical species present in a sample analyzed by LC-ICPMS is also difficult, because commercial standards are not available for many complex analyte species.13 Additionally, the solvents or buffers used in the separation step can cause serious matrix effects in the ICPMS.14 By adding methanol to a sample, Larsen and Stu¨rup demonstrated that the presence of carbon enhances the signal intensity of arsenic in ICPMS.15 Similarly, Heumann and coworkers showed similar results with dissolved organic carbon present as humic acid.16 Alternately, if a buffer is used, signal suppression may develop over the course of an analysis due to salt build-up on the nebulizer tip, sampler, and skimmer cones and ion optics or due to defocusing of the ion beam.14 Calibration of transient signals such as those obtained with LC-ICPMS may be performed using either the method of isotope dilution, external standards, or standard additions. Isotope dilution has been widely reported in the literature for online calibration of LC-ICPMS.16-21 While the method of isotope dilution is accurate, there are drawbacks such as the unsuitability of the method to elements with only one stable isotope, and the expense (10) Caruso, J. A.; Montes-Bayon, M. Ecotoxicol. Environ. Saf. 2003, 56, 148163. (11) Schaumloffel, D. Anal. Bioanal. Chem. 2004, 379, 351-354. (12) Waddell, R.; Lewis, C.; Hang, W.; Hassell, C.; Majidi, V. Appl. Spectrosc. Rev. 2005, 40, 33-69. (13) Hirner, A. Anal. Bioanal. Chem. 2006, 385, 555-567. (14) Evans, E. E.; Giglio, J. J. J. Anal. At. Spectrom. 1993, 8, 1-18. (15) Larsen, E. H.; Sturup, S. J. Anal. At. Spectrom. 1994, 9, 1099-1105. (16) Heumann, K. G.; Rottmann, L.; Vogl, J. J. Anal. At. Spectrom. 1994, 9, 13511355. (17) Beauchemin, D.; Specht, A. A. Anal. Chem. 1997, 69, 3183-3187. (18) Rottmann, L.; Heumann, K. G. Anal. Chem. 1994, 66, 3709-3715. (19) Rottmann, L.; Heumann, K. G. Fresenius J. Anal. Chem. 1994, 350, 221227. (20) Di Marco, V. B.; Bombi, G. G. J. Chromatogr., A 2001, 931, 1-30. (21) Heumann, K. Anal. Bioanal. Chem. 2004, 378, 318-329. 10.1021/ac061616v CCC: $37.00

© 2007 American Chemical Society Published on Web 11/17/2006

Table 1. Solutions and Loop Volumes by Experiment experiment

carrier

peak solution

spiking solution

mL-1

mL-1

loop volumes

matrixfree

1% nitric acid

145 ng multielement standard in carrier

150 ng multielement standard + Rh standard in carrier

1.394 mL (peak) 328.0 µL (spike)

constant matrix

500 µg mL-1 Na in 1% nitric acid

20 ng mL-1 multielement standard in carrier

40 ng mL-1 multielement standard + Rh standard in 1% nitric acid

1.394 mL (peak) 328.0 µL (spike)

gradient matrix

1% nitric acid

20 ng mL-1 multielement standard in (1000 µg mL-1 Na in 1% nitric acid)

40 ng mL-1 multielement standard + Rh standard in carrier

1.394 mL (peak) 328.0 µL (spike)

biological matrix

1% nitric acid

SRM 2670a: toxic elements in urine, high level

75 ng mL-1 multielement standard + Rh standard in carrier

821 µL (peak) 172 µL (spike)

of enriched isotope standards. External standards or standard additions may be employed confidently only when the identities of all species of interest are known and standards in the same chemical form can be acquired. Our laboratory has been investigating the method of standard additions for ICPMS22,23 in particular to promote the application of automated “intelligent” procedures.24 Others have demonstrated the online application of standard additions as a useful technique to reduce analysis time and the amount of sample preparation required.25-31 In particular, Wiederin et al. proposed an interesting method for online standard additions for ICPMS.30 The technique involved the use of two injection loops and a direct injection nebulizer (DIN). The sample is loaded into a large injection loop, and the standards are loaded into a small injection loop. As the sample is injected into the ICPMS, the standards are injected on top of the sample plug. As such, the sample can be calibrated from a single sample injection. Because of the use of the DIN, the sample and standard peaks are rectangular, with flat tops. The method of online standard additions, applied to transient signals, offers the potential of allowing the complete analysis to be done immediately as the analytes elute, eliminating the need for a second standard addition run. In the traditional application of standard additions in chromatography, the spiked standard needed to be in the same chemical form as the analyte so that it appeared with the analyte during the elution process. This limitation is avoided in our technique. This paper describes a calibration technique for use with transient signals, such as those obtained with LC-ICPMS. In this technique, online standard additions are made as spikes to a bolus of analyte. This procedure produces peaks that closely resemble chromatographic peaks, allowing the technique to be evaluated for its applicability in analyses involving liquid separations techniques in general. (22) Abbyad, P.; Tromp, J. W.; Lam, J.; Salin, E. D. J. Anal. At. Spectrom. 2001, 16, 464-469. (23) Hamier, J.; Salin, E. D.; Huxter, V. J. Anal. At. Spectrom. 2003, 18, 71-75. (24) Branagh, W.; Whelan, C.; Salin, E. D. J. Anal. At. Spectrom. 1997, 12, 13071315. (25) Agudo, M.; Rı`os, A.; Valca´rcel, M. Anal. Chim. Acta 1995, 308, 77-84. (26) Beauchemin, D. Anal. Chem. 1995, 67, 1553-1557. (27) Campı´ns-Falco´, P.; Bosch-Reig, F.; Blasco-Go´mez, F. Anal. Chim. Acta 1999, 379, 89-97. (28) Coedo, A. G.; Dorado, M. T.; Ruiz, J.; Escudero, M.; Rubio, J. C. J. Mass Spectrosc. 1996, 31, 427-432. (29) Israel, Y.; Barnes, R. M. Analyst 1989, 114, 843-848. (30) Wiederin, D. R.; Smyczek, R. E.; Houk, R. S. Anal. Chem. 1991, 63, 16261631. (31) Huang, C.; Beauchemin, D. J. Anal. At. Spectrom. 2003, 951-952.

Table 2. Instrument Conditions and Experimental Parameters instrument ICP rf power plasma gas flow auxiliary gas flow aerosol carrier gas flow rate sample liquid uptake mass spectrometer detector ion lens voltage auto lens peak scan parameters scan mode replicates dwell time resolution sweeps per reading readings per replicate

PE-Sciex Elan 6000 ICPMS 1.0 kW 15 L min-1 1.2 L min-1 0.825 L min-1 1.8 mL L min-1 pulse mode 7.5 V off peak hopping 1000 20 ms 0.1 amu 1 1

EXPERIMENTAL SECTION Solutions. All solutions were prepared gravimetrically, using a solution of 1% nitric acid (Fluka TraceSelect, Buchs, Switzerland) in Milli-Q purified water (Millipore, Billerica, MA) for all dilutions. A 7 µg mL-1 multielement stock solution was prepared by diluting a 100 µg mL-1 multielement standard (SCP Science, Baie d’Urfe´, QC, Canada). This stock solution was used in the preparation of all other multielement solutions. Where required, a sodium matrix was prepared by dissolving high-purity sodium nitrate (Alfa Aesar, Ward Hill, MA) in a solution of 1% nitric acid. In all cases, the peak solution and the spike solution contained the same elements with the exception of rhodium internal standard (SCP Science), which was added to the spike solution in order to later determine the duration of the spike peaks. The solutions used in each experiment are described in Table 1. Apparatus. A PE Sciex Elan 6000 ICPMS with a cross-flow nebulizer and Scott-type spray chamber was used for the experiments. Typical operating parameters for the ICPMS are listed in Table 2. Four sample injection loops with approximate volumes of 1 mL, 500 µL, 250 µL, and 100 µL were calibrated gravimetrically in order to determine their total volume, including that of the tubing used to attach them to the injection valves. The total volumes of the loops were determined to be 1.394 ( 0.002 mL, 821 ( 5 µL, 328.0 ( 0.6 µL, and 172 ( 2 µL, respectively, based on the average of 10 replicate measurements. The injection loops used in each experiment are listed in Table 1. The flow injection loops and valves were connected to the ICPMS as illustrated in Figure 1. Both injection valves were Analytical Chemistry, Vol. 79, No. 2, January 15, 2007

689

Figure 1. Schematic diagram of the online standard additions apparatus. Sam, sample; Std, standard; C, carrier; I, internal standard (optional). Sample injection loop volume 1.3940 mL, spike injection loop volume 0.3280 mL, sample carrier flow rate 0.9 mL min-1, and spike carrier flow rate 0.9 mL min-1.

arranged so that, when in the “load” position, carrier passed to the nebulizer and peak or spike solution filled the injection loop with the excess passing to waste. In the “inject” position, carrier was directed through the injection loop, flushing the loop contents to the nebulizer, and the spike or peak solutions were directed to waste. A 38-cm length of knotted 3.00-mm-inner diameter Tygon tubing was used as a mixing coil and positioned between the injection valve of the large sample loop and the T-junction to promote dispersion of the sample. This created a Gaussian-like analyte peak rather than a flat-topped peak. The length of tubing joining the T-junction to the nebulizer was kept as short as possible in order to minimize dispersion of the spike peaks and, thus, minimize the area of the underlying peak which is obscured by the spikes. For experiments involving a gradient matrix effect a solution of 10 ng mL-1 yttrium was mixed into the peak carrier after the large loop, but before the mixing coil, so that a constant concentration of yttrium was delivered to the ICPMS. The yttrium signal could therefore be used as an internal standard to gauge the effect of the sample matrix on the ionization of all elements. Elements were determined in peak-hopping mode with a 20ms dwell time per element. The data acquisition was started 30 s after the large loop was switched to the inject position. Additional details are provided in Table 2. Spikes were added at a regular, 60-s interval to the main carrier stream by manually switching between the “inject” position and the “load” position. CALCULATIONS Overview. In simplistic terminology, the technique of standard additions involves measuring the analyte solution once and then measuring it a second time after adding a known amount of analyte. To achieve this goal with a transient signal, we have elected to add a series of spikes overlaying the analyte peak. The first step in the data-handling procedure involves ignoring the analyte spike information and using a curve-fitting routine to fit the remaining data, which consists of the peak shape with spike time segments missing. A simplex-based curve fit is used to make an estimate of the underlying peak. The underlying peak is then integrated to obtain the “analyte solution” measurement. The 690

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Figure 2. (a) Area of peak from simplex curve fit. (b) Area of peak and standard additions spikes.

entire peak, with spikes, is integrated to obtain the second “spiked solution” measurement. The process is presented in more detail below. Standard Additions. A two-point standard additions calibration was performed. Because of the large linear range of the ICPMS, it has been shown that a two-point standard additions calibration is as effective as a multiple-point standard additions calibration.22 Since the fit peak obtained from the simplex optimization represents only the sample peak with no spikes added, the baseline-subtracted area under the best-fit peak was used as the first point (Figure 2a). The area of the second point was calculated as the baseline-subtracted total area under the actual analysis data, including the spikes (Figure 2b). The added mass, m, of the second point was calculated as

m ) ncspikevloop

(1)

here n is the number of spikes, cspike is the concentration of the spike solution, and vloop is the gravimetrically calibrated total volume of the injection loop.

Data Analysis by Curve Fitting. To fit the underlying analyte peak, a simplex optimization program32 was written in the Matlab programming language. The peak shape was described as a hybrid of Gaussian and truncated exponential functions.20,33 This function requires only four parameters to describe the shape of the peak, according to the equation

f(x) )

{

(

h exp

2

-(x - z)

)

2w2 + s(x - z)

0

when 2w2 + s(x - z) > 0 when 2w2 + s(x - z) e 0

}

(2)

where h represents the maximum peak height, z is the position of the peak maximum, w is the width of the peak, and s is related to the symmetry of the peak. The simplex searched for the combination of parameters that yielded the smallest difference between the experimental data and the curve fit function. This was evaluated as the sum of squares of the residuals. The simplex optimization ended when the curve fit showed an improvement of less than 0.01% for 20 consecutive iterations of the simplex. In order to fit the underlying peak despite the presence of the spikes, it was necessary to determine the time index of the beginning and end points of each spike. This was easily accomplished by examining the data for the Rh spike signal, which was present only in the spiking solution and not in the underlying peak. The beginning and end points of each spike were the points at which the Rh increased from and returned to its baseline value. All data points that lay in the range of the spikes were ignored when calculating the peak fit function, and the fit therefore was calculated using only data from the underlying analyte peak. In the presence of the biological matrix (urine), it was found that the original (eq 2) peak function was not capable of accurately describing the observed peak shape for the large sample loop. In this case, the following exponentially modified Gaussian function34 was used to fit the underlying peaks:

f(x) )

(

)[

(

c2 d b-x b-x c acx2π exp + 2 - erf + 2d d 2d |d| x2c x2d

)]

(3)

where a and b are the amplitude and center, respectively, of the deconvolved Gaussian, c relates to the peak width, and d is the exponential time constant. The change of peak functions required only a small modification to the simplex program, since both functions have only four parameters requiring optimization. RESULTS AND DISCUSSION Curve Fitting of Modeled Data. Data were simulated in order to test the simplex program. Simulated peaks were created in Matlab using eq 2. Random noise (5%) was added to the peak in order to simulate experimental conditions. The simplex program was applied to the simulated peaks, evaluating all data points in the measure of goodness of fit, since no spikes were present. The values of h, z, w, and s were compared for the original simulated (32) Deming, S. N.; Morgan, S. L. Anal. Chem. 1974, 46, 1170-1181. (33) Li, J. J. Chromatogr., A 2002, 952, 63-70. (34) Tablecurve 2D 5.01 for Windows User’s Manual; Systat Software Inc.: Richmond, CA, 2002.

Table 3. Comparison of Curve Parameters Errors for Modeled and Optimized Best-Fit Curves average error (%) curve parameter

curve fit of entire peak

curve fit eliminating the spike areas of the peak

height (h) translation (z) skew (s) width (w)

0.2 0.1 1 0.3

-1 -0.02 5 1

peak and the best-fit curve generated by the simplex optimization. An average error of 0.52% between the true values and best-fit data was obtained, indicating that the simplex program can be used to provide an accurate fit for a peak created using eq 2. The results are presented in Table 3. For the purposes of this experiment, the curve-fitting optimization needed to be capable of determining the best fit of a peak where only a portion of the data was used, since areas where spikes appeared were ignored. Simulated data for a large peak with short spikes added was created using Matlab. A peak was created using eq 2, and 5% random noise was added. A rectangular spiking pattern was created with a 50% duty cycle, and 5% random noise was added to the spikes. The peak and the spikes were added together, and the simplex peak-fitting optimization was performed on the simulated spiked data using only the regions between spikes in order to test the fit. The four curve-fitting parameters of the simulated spiked data and best-fit data for the underlying curve were compared. All four parameters were within 5% of the actual value of the peak parameters. The results are also presented in Table 3. The results indicate that ignoring some regions of the peak did not adversely affect the quality of the bestfit curve. Reproducibility of Spikes and Peaks. The reproducibility of the areas of the peaks created by injecting either loop into the carrier flow will determine the precision of the standard additions technique; therefore, the areas of several injections for both loops were compared. The area of each peak or spike was calculated by subtracting the baseline signal from the data and summing the signal intensity for each data point. For five injections of the small sample loop in a single analysis, the average relative standard deviation of the area was 1.01%. For the larger loop, the average relative standard deviation of five injections, each from a separate analysis, was 0.5%. From these experiments, it can be seen that reproducibility of the injections will not limit the ability to obtain good precision with this standard additions calibration technique. Curve Fitting of Experimental Data. The peak function shown in eq 2 was selected because it required only four parameters, rather than seven or more for many other modified Gaussian peak functions.22 Before applying the simplex optimization to the standard additions procedure, it was necessary to verify that eq 2 could accurately describe the peaks created by the flow injection of the larger loop with additional tubing for dispersion. Peaks injected from the large sample loop with no spikes added were analyzed using the ICPMS. The simplex program was applied to the output data, and the areas of the actual and fit peaks were compared for 11 elements. Figure 3 shows a plot of the experimental data for silver and its optimized best-fit curve. The Analytical Chemistry, Vol. 79, No. 2, January 15, 2007

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Table 5. Results of Online Standard Additions Calibration in 0.5% HNO3 element 107Ag 138Ba 9Be 209Bi 7Li 139La 98Mo 121Sb 88Sr

mass injected to the ICPMS (ng)

determined massa (ng)

199.6 199.4 201.8 199.4 201.0 199.6 200.2 200.0 199.6

197 ( 9 199 ( 10 205 ( 11 203 ( 10 201 ( 6 201 ( 10 202 ( 8 201 ( 10 201 ( 9

a Average values for four replicate measurements ( standard error at 95% confidence level.

Figure 3. 107Ag analyte peak with simplex best-fit results. A total mass of 40 ng was injected into the carrier stream (1.3940 mL of 28.72 ng mL-1 solution) at a flow rate of 0.9 mL min-1. Table 4. Accuracy of Optimized Best-Fit Curve Areas by Element element 107Ag

average error (%)

88Sr

0.79 0.91 0.86 0.75 1.53 0.89 1.04 0.79 0.24 0.87 0.91

average

0.87

138Ba 9Be 209Bi 39K 7Li 139La 98Mo 31P 121Sb

results of the comparison of the calculated (best fit) and actual areas are summarized in Table 4. The average error between the actual and calculated peak areas was 0.87%. Therefore, the bestfit curve generated by the simplex optimization can be used as a reliable representation of experimental peak, and the quality of the fit lies well within the acceptable range. Standard Additions. The flow injection peaks were spiked with the standard at a regular interval. The spiking pattern is shown in Figure 2. A summary of the standard additions results for four replicates is provided in Table 5. The overall average error was 0.85%, showing that the method is not biased. The relative standard deviation between the results of the four replicates was close to 5% for most elements. These results show that this method is capable of performing an accurate and precise calibration. The spiked standard additions method relies on at least one spike overlaying the analyte peak, which means that the spikes must be narrower and more frequent than the underlying peak. This could become problematic for very narrow peaks, as the spike may miss the peak entirely or be completely obscured by the spike. Decreasing the width of the spikes will allow the application of the method to narrow analyte peaks. The frequency of the spikes may be increased by performing more injections with a smaller injection loop, increasing the speed of the pump for the spikes, or using a direct injection nebulizer to minimize the 692 Analytical Chemistry, Vol. 79, No. 2, January 15, 2007

Table 6. Results for the “Steady-State Constant Matrix Effect Experiment” (Standard Additions Calibration in the Presence of a 500 µg mL-1 Na Matrix) element 107Ag 75As 114Cd 59Co 55Mn 98Mo 58Ni 121Sb 205Tl 51V

mass injected to ICPMS (ng)

determined mass fittinga (ng)

27.94 27.52 27.83 27.49 27.94 27.80 27.69 27.52 27.77 27.55

27.8 ( 0.4 27.4 ( 0.4 28.1 ( 0.5 27.6 ( 0.4 28.1 ( 0.4 27.8 ( 0.4 27.7 ( 0.5 27.4 ( 0.4 27.4 ( 0.5 27.7 ( 0.4

a Average values for eight replicate measurements ( standard error at 95% confidence level.

diffusion of the spike plug. Steady-State Constant Matrix Effect Experiment. A 500 µg mL-1 Na solution was used as the carrier solution to test the calibration method in an environment that induces a severe matrix effect. Although Na is not a matrix that is commonly problematic for LC-ICPMS analysis, it was selected as the matrix element for several reasons. Na is known to cause signal suppression for most analytes with relatively low concentrations, it is available in a very pure form (as NaNO3) suitable for ICPMS analysis, and the spectroscopic interferences arising from the NaNO3 species are minimal. If the proposed calibration method can compensate for signal suppression in the presence of Na, it should extend to other nonspectroscopic interferences as well. A 30 ng mL-1 solution prepared in a 500 µg mL-1 Na matrix was analyzed by external standards, and an average signal suppression of 22 ( 6% was observed. The spikes were positioned as shown in Figure 2. The results of eight trials of the standard additions calibration in the Na matrix are summarized in Table 6. Despite the strong matrix effect, very accurate and precise results were obtained using the simplex curve fit. The overall average error was -0.03% and the average absolute error was 1.4%, again demonstrating accurate results and no bias in the method. The reproducibility of the determinations was good at 1.7% RSD. Transient (Gradient) Matrix Effect Experiment. In the case of online separation methods coupled to ICPMS, the matrix effect will not necessarily remain constant throughout the analyte peak.

Table 7. Results for the Standard Additions Calibration for the “Transient (Gradient) Matrix Effect Experiment” element (isotope) 107Ag 138Ba 59Co 52Cr 65Cu 55Mn 98Mo 58Ni 208Pb 121Sb

injected mass (ng)

spike position 1 resultsa (ng)

spike position 2 resultsb (ng)

with Y internal stdc (ng)

27.07 26.93 26.63 26.74 27.01 27.07 26.93 26.82 26.63 26.66

25 ( 1 26 ( 1 25 ( 1 25.5 ( 0.6 29 ( 3 25.8 ( 0.2 26 ( 1 25.0 ( 0.3 25 ( 1 26 ( 1

26 ( 1 26 ( 1 27 ( 1 26 ( 2 27 ( 2 27 ( 1 26 ( 2 26 ( 2 26 ( 1 26 ( 1

26.3 ( 0.9 26 ( 1 26.1 ( 0.9 26.1 ( 0.5 28 ( 2 26.6 ( 0.7 27 ( 1 25.9 ( 0.9 25.9 ( 0.7 26 ( 1

a For spike positions as shown in Figure 5a, four replicate measurements. b For spike positions as shown in Figure 5b, four replicate measurements. c Same measurements as (b), with an yttrium internal standard.

Figure 4. (a) Analyte peak with spikes for a transient matrix effect. (b) Spikes with the analyte peak removed, showing change in matrix effect. The dotted line indicates the top of the first spike, without a matrix effect present.

Figure 5. (a) Spike position 1, overlaying sides of analyte peak. (b) Spike position 2, overlaying the apex of the analyte peak.

In this experiment, the carrier composition was 1% nitric acid, and the sample matrix was 1000 µg mL-1 Na. Figure 4a shows the signal obtained for Ag with the underlying peak and spikes, and Figure 4b shows the analyte spikes with the underlying peak subtracted from the total signal. The spike areas are not constant from spike to spike, indicating that the matrix effect was not constant throughout the underlying peak. During the first four trials, the spikes were started at the same time and were added every 60 s over the analyte peak. For the last four trials, the spiking frequency remained the same, but the first spike was added 30 s later than the first four trials, so that all of the spike positions were shifted by 30 s. This is illustrated in Figure 5. The results of these experiments are shown in Table 7. It can be seen in the cases of 59Co and 55Mn that both the precision and accuracy of the analysis are slightly improved when the leading edge of the peak is not obscured by the spiking pattern, because the signal in this region changes more rapidly than in any other area of the

peak, making it a crucial area for fitting. For all other analytes, the results for the two different spike patterns were not significantly different at the 95% confidence level. A different approach was selected. An yttrium internal standard was added to the peak carrier stream, and the ratio of the analyte signal to the internal standard was calculated. The peak-fitting and calibration calculations were performed, and the results are shown in Table 7. It can be seen that the use of an internal standard does not significantly improve the accuracy or precision of the method. However, under severe drift conditions, the method of standard additions with an internal standard has been shown to be accurate, independent of the internal standard selected.35 The addition of an internal standard in the carrier stream allows the correction for any distortion in the shape of the analyte peak due to the changing matrix effect. The method of spiked standard additions with an internal standard is suitable for transient matrix effects and should also apply to linear drift or matrix effects (as in certain types of HPLC with carrier gradients), although this was not verified experimentally. Standard Reference Solution with a Biological Matrix. Validation of the online standard additions method was performed using a certified reference material of toxic metals in human urine. Eight elements were determined with concentrations ranging from 0.82 to 233.2 µg L-1. The results of the measurements are presented in Table 8. It can be seen that, with the exception of cadmium, the determined values agree well with the certified values. The discrepancies in the cadmium values are due to a number of possible spectral interferences with the oxides of major matrix components that are not present in the spiking solution.36 This type of interference cannot be corrected by using standard additions calibration. Although the concentrations of the measured analytes ranged over more than 2 orders of magnitude, all analytes in the spiking solution were present at 74.4-75.4 µg L-1. This ability to determine a large range of analyte concentrations with a common (35) Salin, E. D.; Antler, M.; Bort, G. J. Anal. At. Spectrom. 2004, 19, 14981500. (36) Murphy, K. E.; Beary, E. S.; Rearick, M. S.; Vocke, R. D. Fresenius J. Anal. Chem. 2000, 368, 362-370.

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Table 8. Results for the Standard Additions Calibration for the “Standard Reference Solution with a Biological Matrix” (Urine)

element 55Mn 59Co 82Se 98Mo 114Cd 121Sb 205Tl 207Pb

certified concn (µg L-1)

concn determined by simplex fittinga (µg L-1)

99 ( 12 51.2 ( 3.2 229.5 ( 8.3 114.1 ( 4.8 4.862 ( 0.084 0.824 ( 0.070 5.417 ( 0.064 233.2 ( 9.4

97 ( 11 48 ( 3 210 ( 28 114 ( 6 8.0 ( 0.5 0.9 ( 0.2 5.3 ( 0.4 220 ( 18

a Average values for g3 replicate measurements ( standard error at 95% confidence level.

spiking concentration offers two advantages: (1) no knowledge of the analyte concentrations is required prior to analysis, and (2) the preparation of the spiking solution can be done quickly and simply using an appropriate multielement standard. CONCLUSIONS The calibration method demonstrated appears to offer considerable advantages for the analysis of transient signals. There should be at least a factor of 2 improvement in throughput as only one run is needed for analysis and calibration. Good accuracy was obtained in the presence of a strong matrix effect. The

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Analytical Chemistry, Vol. 79, No. 2, January 15, 2007

placement of the standard addition spikes does not appear to have a significant effect on the accuracy of the method; however, it was observed that in some cases spikes positioned over either the apex or leading edge of the underlying peak led to slightly decreased precision. The addition of an internal standard to the carrier stream appears to be necessary only in cases with severe drift or drastic signal distortion. The calibration of transient signals by this method can rival the accuracy obtained with steady-state analysis. The added standard spikes do not need to be in the same chemical form as the analytes, and the concentrations of the spikes do not need to be matched to the analyte concentrations, allowing for greater versatility. The method is also widely applicable as it requires only basic equipment that is standard to most ICPMS instruments, within the limitation that the spike peaks must remain significantly shorter than the underlying peak. This calibration is suitable for hyphenated liquid separation-ICPMS techniques and has the potential to simplify and reduce the costs of speciation analysis for complex biological and environmental samples. ACKNOWLEDGMENT The authors thank David H. Burns for helpful conversations regarding peak fitting and optimization.

Received for review August 29, 2006. Accepted October 11, 2006. AC061616V