Anal. Chem. 1999, 71, 3497-3502
Determination of Continuous Size and Trace Element Distribution of Colloidal Material in Natural Water by On-Line Coupling of Flow Field-Flow Fractionation with ICPMS Martin Hassello 1 v,*,† Benny Lyve´n,*,† Conny Haraldsson,‡ and Waraporn Sirinawin†
Analytical and Marine Chemistry, Go¨teborg University SE-412 96 Go¨teborg, Sweden, and Swedish National Testing and Research Institute, P.O. Box 857 SE-501 15 Borås, Sweden
The coupled technique here presented utilizes the sizeseparating capabilities of flow field-flow fractionation (FlFFF) and the high-sensitivity, multielement detection capabilities of inductively coupled plasma mass spectrometry (ICPMS). The system enables determinations of colloidal size and element distributions for 28 elements (including carbon) in natural waters. To cope with the low concentrations often found for colloids and trace elements in natural waters, the conventional FlFFF system has been modified to include an on-channel preconcentration procedure allowing injection of up to 50 mL of sample. The performance of the FlFFF-ICPMS system is further evaluated in terms of reproducibility and detection limits. The size distributions of six selected elements are given, and different colloidal distributions for the elements are discussed. The mobility and bioavailability of trace elements in natural systems are to a large extent determined by the physical and chemical forms in which the metals occur. An element in a natural water system is partitioned between different physical states such as truly dissolved or complexed or associated with organic and inorganic colloids and with mineral particles.1,2,3 The distribution between these different forms will determine whether an element is available for uptake via membrane transport mechanisms and will affect its function, either as a toxin or as a nutrient.2 Furthermore, the physical form of an element will affect its transport properties and hence its rate of removal from a natural water system.4,5 The size of the molecule or colloid to which an element is bound provides important information about the behavior of that element. Methods to study the size distributions of various elements include size fractionation using dialysis, ultrafiltration,
or size exclusion chromatography (SEC) with subsequent metal determination.6 Currently, the most common method for colloid size studies in natural waters is cross-flow ultrafiltration.7 There has been an extensive debate about the reliability of cross-flow ultrafiltration for size fractionation of natural colloids, which led to an intercalibration study. The outcome was reported in a recent special issue8 and the conclusion was that ultrafiltration fractionation has to be interpreted with great caution. The problems include imprecise molecular weight cutoff of available membranes and risk of perturbation of the sample colloids by processes such as aggregation or adsorption during the rather slow (up to several hours) concentration enrichment procedure.9 Cross-flow ultrafiltration followed by metal determination has been widely used in colloid-metal interaction studies despite the likelihood of changes in speciation during the concentration or fractionation procedure. Field-flow fractionation (FFF) is a family of relatively new separation techniques which build on the principle of combining a field perpendicular to a laminar flow in thin channels (100500-µm thickness). The diffusion coefficient will determine to what extent colloids and particles in the flow channel will be able to diffuse against the perpendicular field. The parabolic flow profile will then cause smaller particles to travel in faster flow vectors, thus eluting more quickly than larger particles.10 FFF techniques are classified according to the field used, the major variants being sedimentation FFF (SedFFF) with a centrifugal field and flow FFF (FlFFF) with a cross-flow field. Flow FFF is the method with the widest operating size range spanning approximately from molecular weight 1000 (a few nm) to 1 µm. SedFFF on the other hand is the FFF technique with the highest selectivity, but it is not applicable to colloids smaller than about 0.05 µm and is therefore mainly suitable for suspended or sedimented particles. The advantages of FFF include minimal exposure of the colloids to surfaces during the separation, particularly in comparison to
†
Goteborg University. Swedish National Testing and Research Institute. (1) Stumm, W.; Morgan, J. J. Aquatic Chemistry, 3rd ed.; John Wiley: New York, 1996. (2) Tessier, A., Turner, D. R., Eds. Metal Speciation and Bioavailability in Aquatic Systems; John Wiley: Chichester, 1995; p 679. (3) Stumm, W. Colloids Surf. A 1993, 73, 1-18. (4) McCarthy, J. F.; Zachara, J. M. Environ. Sci. Technol. 1989, 23, 496-502. (5) Batley, G. E., Ed. Trace element speciation: Analytical methods and problems; CRC Press: Boca Raton, 1989; Vol. 1, p 350. ‡
10.1021/ac981455y CCC: $18.00 Published on Web 06/30/1999
© 1999 American Chemical Society
(6) Batley, G. E. In Trace element speciation: Analytical methods and problems; Batley, G. E., Ed.; CRC Press: Boca Raton, 1989; Vol. 1, pp 43-76. (7) Buffle, J.; Perret, J.; Newman, J. In Environmental Particles I; Buffle, J., van Leeuwen, H. P., Eds.; Lewis: Chelsea, 1992; pp 171-230. (8) Buesseler, K. O. Ed., The use of cross-flow filtration (CFF) for the isolation of marine colloids Mar. Chem. 1996, 55, 1-204. (9) Gustafsson, O ¨ .; Buesseler, K. O.; Gschwend, P. M. Mar. Chem. 1996, 55, 93-111. (10) Giddings, J. C. Analyst 1993, 118, 1487-1494.
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ultrafiltration and SEC, and the ability to adjust the system to different particle size ranges by changing the perpendicular field. A variety of methods is available for studying the properties of trace elements in natural waters, e.g., electrochemical techniques,11 methods based on ion exchangers,12 and chromatographic methods.13,14 However, ICPMS is gaining in popularity due to its high sensitivity, multielement capability, and also its ability to be coupled on-line to suitable separation methods. The use of SEC coupled with ICPMS for metal speciation in humic isolates has been demonstrated.14,15 The problem associated with this technique is mainly the small injection volume, which necessitates the use of a preconcentration step (e.g., ultrafiltration or solid-phase extraction) with concentration factors of 50-500, possibly disturbing the ratio between adsorbed and free metal ions. Due to the large surface area of the packing material to which the sample colloids are exposed in the SEC column, there is a significant risk of adsorption.6 Furthermore, the large surface area will also be a source of contamination.16 Separation using SedFFF followed by ICPMS determination has been demonstrated for mineral and suspended riverine particles17,18 as well as for soil particles.19 In this paper, the use of FlFFF with on-channel preconcentration and on-line ICPMS detection is demonstrated for simultaneous determinations of colloidal size distributions and metal content from ultratrace levels to major components for 28 elements. EXPERIMENTAL SECTION Reagents. All buffer reagents used were of analytical grade dissolved in Milli-Q water (Millipore). High-purity acid was prepared in a clean lab by sub-boiling point quartz distillation of analytical grade HNO3 (Merck). The molecular weight standards used were polystyrenesulfonates (PSS) ranging from 1100 to 356 000 (Phenomenex, Torrance, CA). Metal standards were prepared by dilution of 1000 mg L-1 stock standard solutions (Merck). Sampling and Filtration. The freshwater samples were taken from Delsjo¨ba¨cken (Go¨teborg, Sweden), a small creek with moderate colloid concentrations. The samples were collected in precleaned polyethylene bottles, transported to the laboratory, where large particles were removed by two different methods. The first was filtration through a precleaned 0.45-µm cellulose acetate filter (Millipore), and the second was settling for 24 h at 4 °C, after which the overlying 50% of the sample was pumped to a clean bottle. These two pretreatment procedures were carried out in order to remove particles that would otherwise cause steric interferences in the FFF separation,10 i.e., roll quickly along the (11) Buffle, J. In Complexation Reactions in Aquatic Systems: An Analytical Approach; Ellis Horwood: Chichester, 1988; pp 467-562. (12) Haraldsson, C.; Lyve´n, B.; Pollak, M.; Skoog, A. Anal. Chim. Acta 1993, 284, 327-335. (13) Heumann, K. G.; Gallus, S. M.; Ra¨dlinger, G.; Vogl, J. Spectrochim. Acta Part B 1998, 53, 273-287. (14) Itoh, A.; Haraguchi, H. Anal. Sci. Suppl. 1997, 13, 393-397. (15) Vogl, J.; Heumann, K. G. Fresenius J. Anal. Chem. 1997, 359, 438-441. (16) Landing, W. M.; Haraldsson, C.; Paxe´us, N. Anal. Chem. 1986, 58, 30313035. (17) Taylor, H. E.; Garbarino, J. R.; Hotchin, D. M.; Beckett, R. Anal. Chem. 1992, 64, 2036. (18) Murphy, D. M.; Garbarino, J. R.; Taylor, H. E. J. Chromatogr. 1993, 642, 459. (19) Chittleborough, D. J.; Hotchin, D. M.; Beckett, R. Soil Sci. 1992, 153, 341.
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membrane, coelute with much smaller particles, and give rise to an artifact in the results. The filtered pretreated samples were kept at 4 °C, and analyzed a soon as possible, and never stored for more than 2 weeks since some change in colloidal size was observed in this study when the samples wereas stored for a long time, 2 months. Instrumentation. A flow FFF system (Figure 1) (FFFractionation, Salt Lake City, UT), equipped with an 1000 MWCO ultrafilter membrane (Omega, Pall-Filtron), modified to allow injection of large sample volumes, was used.20 To be able to determine trace metal distributions, considerable effort was devoted to making the entire system metal free to avoid contamination. This was done by replacing as many of the metal parts as possible with plastic materials, e.g., PTFE, PEEK, PMMA, and acetal. The UV detector cell and some of the parts for one of the LC pumps, which could not be replaced by plastic materials, were instead replaced by titanium. Initially, the FlFFF system had an unacceptably high background for some metals and therefore a cleaning procedure was necessary every time a new membrane was installed in the FlFFF channel (typically every 3 months). Effective leaching of trace metals is usually carried out with a combination of fairly strong acids, but since some parts of the system, such as the membrane, will be damaged by high acid concentrations a mild precleaning procedure was adopted. The leaching began by pumping 1.4 mmol L-1 nitric acid for 4 days, followed by 1.5 mmol L-1 hydrochloric acid for 2 days, and finally 2 days with 1.4 mmol L-1 nitric acid. After the acid wash, the system was equilibrated with the buffer solution for 10 h and blanks were determined. The instrument setup is shown in Figure 1, and optimized operating conditions are presented in Table 1. The FlFFF analysis procedure consists of three steps shown in Figure 2: sample loading and focusing (A), relaxation (B), and finally elution (C). The sample was loaded through the backward end of the channel, and a flow ratio of 1:12.5 between the forward (0.4 mL min-1) and backward (5 mL min-1) focusing flows was used. During sample loading and focusing, the carrier solution is allowed to pass out of the channel only through the cross-flow outlet, which is connected to waste via a three-way valve.3 This bypass is used in order to lower the back pressure that otherwise occurs due to high flow rates during sample loading and focusing in combination with the long and narrow connecting tubing. During relaxation, the only flow through the channel is the cross-flow. This is achieved by stopping the focus pump and switching valves 1, 2, and 3. The relaxation step is long enough for approximately two void volumes to pass through the channel. The third step is elution, where the channel flow and crossflow revert to the standard FlFFF configuration and data acquisition is started. The resulting fractogram is acquired, and the retention time is corrected for the dead volume between the channel outlet and the detectors. Although the volume of the sample injected is not a limiting factor when on-channel preconcentration is used, there is a limitation in the amount of colloids that can be injected before retention times start to decrease due to repulsion between the colloids, i.e., the channel becomes overloaded. When an unknown (20) Lyve´n, B.; Hassello¨v, M.; Haraldsson, C.; Turner, D. R. Anal. Chim. Acta 1997, 357, 187-196.
Figure 1. Instrumental setup: configuration of pumps, switching valves, FlFFF channel, ICPMS, and connections. Table 1. Operating Conditions for the FlFFF-ICPMS System channel flow (mL min-1) cross-flow (mL min-1) dilution pump flow (mL min-1) split ratio (waste:ICPMS) sample volume (mL) sample load and focusing time (min) ICPMS nebulizer flow (L min-1) ICPMS auxiliary flow (L min-1) ICPMS cool gas flow (L min-1) rf power (W) UV detector wavelength (nm)
0.50 2.96 1.00 0.8:0.7 46.8 20 0.72 0.8 15 1300 270
type of sample is injected, it is necessary to inject different volumes and to check whether the retention times differ significantly. To achieve more reproducible retention times, it was found necessary to incorporate a cross-flow suction pump which resulted in more stable flows. Both this pump and the cross-flow pump are double-piston pumps with continuous piston speed, thus also having a continuous flow rate on the suction side of the pump (LKB-2150, Pharmacia Instruments). This is an unusual property of modern LC pumps but essential for a cross-flow suction pump if stable flows are to be achieved. Previous studies of the size determinations of polymers and natural colloids have shown that the buffers have to contain a fairly high ionic strength (15-30 mmol L-1) to give an acceptable separation.20,21 However, long runs with high salt contents will degrade the performance of the ICPMS by lowering the sensitivity. It was found that a pH 8.1 buffer containing 5 mmol L-1 borate (21) Hassello ¨v, M.; Hulthe, G.; Lyve´n, B.; Stenhagen, G. J. Liq. Chromatogr. Relat. Tech. 1997, 20, 2843-2856.
and 10 mmol L-1 sodium chloride could still achieve good separation and low sample loss rates. The ICPMS stability was improved by diluting the outlet from the FlFFF system by a factor of 3 followed by splitting (Figure 1), giving a final flow rate of 0.7 mL min-1 to the ICPMS. To obtain the best performance from the nebulizer-spray chamber system, the sample was acidified by diluting the FlFFF outflow with 1.5% (v/v) nitric acid, which also contained 7.5 µg L-1 indium, used as internal standard for the ICPMS measurements. The indium added was also used for tuning the ICPMS since it proved to be important to optimize the signal maximum in the same matrix as the samples. The ICPMS (VG-PQ 1 upgraded with new electronics and the PQ 2+ interface) was equipped with a Meinhard nebulizer and standard Scott spray chamber. Data acquisition was carried out with Time-Resolved software (VG elemental) using the isotopes listed in Table 2 with 3 s/scan. The sensitivity was typically around 40 MHz ppm-1 for indium. All elements were measured in pulse counting mode, except for carbon which was acquired in analog mode. Metal Quantification. The signal obtained from the ICPMS was converted to concentration using blanks and standards prepared in the same matrix as the sample, 1.7 mmol L-1 borate, 3.3 mmol L-1 sodium chloride, and 1% (v/v) nitric acid, reaching the ICPMS. The concentration versus retention time curve was integrated to give the total peak area. The area was then converted to the amount of metal by multiplying by the flow rate and the dilution factor in the FFF-ICPMS interface. The amount of metal found in the fractionated peaks was then compared with the total amount of metal injected (sample volume multiplied by filtered total sample concentration). The filtered sample concentration was Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
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Table 2. Detection Limits and Reproducibility element C Mg Al Ca Mn Fe Co Ni Cu Zn Sr Y Zr Mo Sb Ba La Ce Pr Nd Gd Dy Ho W Pb Bi Th U
isotope detection limita colloidal metal retention time (m/z) (µg L-1) conc RSDb,d (%) RSDc,d (%) 12 26 27 44 55 57 59 60 65 66 88 89 90 95 121 137 139 140 141 147 157 162 165 182 208 209 232 238
0.2 0.3 1 0.006 0.8 0.001 0.01 0.1 0.06 0.0004 0.9 0.1 0.01 0.01 0.04 0.009 0.03 0.008 0.01 0.007 0.006 0.002 0.006 0.01 0.02 0.007 0.006
8.0 28 40 22 42 15 11 16 26 12 6.0 8.1 12 25 8.7 15 4.2 5.1 11 6.6 7.2 8.8 12 13
2.7
6.4 0.81
0.57 3.9
3.1
18 16
a Defined as 3 times the standard deviation of 10 blanks runs over a 3 days. b Relative standard deviation in the colloidal concentration found for five replicate runs of a natural sample. c Relative standard deviation in the retention time for peak maximum for three replicate runs of a natural sample. d Sample from Delsjo ¨ba¨cken, Go ¨teborg.
Figure 2. Schematics of pump and valve settings for on-channel preconcentration; sample loading and focusing (A), relaxation (B), and elution modes (C).
determined by measuring an acidified 0.45-µm filtered sample. Size and Molecular Weight Distributions. The fractograms obtained from the UV or ICPMS signal as a function of the retention time are converted into frequency as a function of size or molecular weight in order to facilitate interpretation of the results.22 This was done either as the hydrodynamic diameter distribution calculated from the FFF theory or as the molecular weight distribution obtained from a calibration with reference PSS standards. The theory for converting the retention time to diameter was first developed by Giddings23,24 showing the relationship between the measured retention ratio R and the theoretically expressed retention parameter λ shown in eq 1. Equation 1 then gives the relationship between the Stokes (hydrodynamic) diameter d, retention time tr, and void time t0 shown in eqs 2, where channel thickness w, width b, length l, volumetric cross-flow rate V˙ c, temperature T, Boltzmann constant k, and buffer viscosity η are used to calculate the diameter. The diffusion coefficient of the PSS standards (molecular weights 1100, 13 000, 150 000, and 356 000) were determined and (22) Beckett, R.; Jue, Z.; Giddings, J. C. Environ. Sci. Technol. 1987, 21, 289295. (23) Giddings, J. C. Sep. Sci. Technol. 1966, 1, 123. (24) Giddings, J. C. Science 1993, 260, 1456-1465.
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calculated according to eq 3, and the relation between the diffusion coefficient and molecular weight is given in eq 4.22,25
R ) t0/tr ) 6λ[coth(1/2λ) - 2λ]
(1)
d ) 2kTbltr/t0wV˙ cηπ
(2)
D ) λ(V˙ c/bL)w
(3)
log D ) -β log Mw + log R
(4)
RESULTS AND DISCUSSION Previous studies7 have suggested that filtration could cause colloids to adsorb or aggregate. In this study, no differences were found between the filtered and settled samples. Since the smallest “steric” particles, and particles with a density close to that of water, may not sediment and consequently cause problems during the settling procedure, filtration was chosen as the best method of pretreatment. A feature that required investigation was the washout time for the ICPMS nebulizer. If there is a long washout time, due to wall effects or large dead volumes in the nebulizer, the element size distributions will be distorted toward larger colloids by signal tailing. The washout time was determined by installing a lowvolume injection valve between the FFF channel and the ICPMS (25) Tanford, C. Physical Chemistry of Macromolecules; John Wiley: New York, 1961; pp 1-710.
Figure 3. FlFFF-ICPMS raw data for the iron signal using three different cross-flows: (A) 2.06, (B) 2.96, and (C) 3.88 mL min-1. The calculated hydrodynamic diameter distribution for these runs is shown in (D).
interface. The washout time was determined to be about 40 s, which is satisfactory, although a sample injection system with lower dead volume could reduce band broadening. This would in practice only affect small particle sizes. With the on-channel preconcentration system, different concentration factors are obtained, depending on the sample volume and how large a volume the specific element is eluting in. Extreme examples of concentration factors in this work using the sample loop of 46.8 mL are 9.0 for nickel and 3.5 for lead (Figure 4). To examine the performance of the method, repeated analyses of a natural sample were carried out. Five replicates over 2 days were used to determine the reproducibility in colloidal metal concentration presented in Table 2; the reproducibility of the peak retention time is also presented. Since the repeated measurements were made over a 2-day period, and the cross-flow was not recorded on the second day, only three replicates were used for the retention time calculations. The colloidal metal determinations show a variability of 1015%, which is satisfactory, bearing in mind that the variability of ICPMS determination of total concentrations is typically in the order of 5-10%.26 For some elements (Mn, Ca, Mg, Cu, Al), we see a considerably higher variation, due to factors such as low concentrations in the colloidal fraction, spectroscopic interferences, or contamination. The good reproducibility obtained (26) Van Leuven, L.; Taylor, P. D. P.; O ¨ rnemark, U.; Moody, J. R.; Heumann, K. G.; De Bie`vre, P. Accred. Qual. Assur. 1998, 3, 56-68.
Figure 4. Calculated hydrodynamic diameter distributions for six elements and for UV detection presented together with a molecular weight scale obtained from calibrations with PSS standards.
otherwise is an indication that the preconcentration procedure is very reproducible. Detection limits were determined for 27 elements as 3 times the standard deviation of the peak area determined for 10 blank injections (MilliQ-water). The peak area was integrated over the retention time where the colloidal material in the natural water samples was eluted (Table 2). For most elements, the detection limits are well below the colloidal concentrations found in the samples, using a 46.8mL sample volume. Some elements could not be detected due to high detection limits, caused by large isobaric interferences from the buffer (As, Cr, V) or due to very low natural concentrations (Ag, Cd, Pt). In Figure 3A-C, raw data are presented for the iron signal from three different sets of FlFFF-ICPMS conditions. The crossflow was varied from 2.06 (Figure 3A) to 2.96 mL min-1 (Figure 3B) and 3.88 mL min-1 (Figure 3C); the different elution patterns illustrate the flexibility of FlFFF, i.e., the ability to adjust the separation parameters for application to different size ranges. Furthermore, this shows that the second peak for iron is really a population of large colloids and not an artifact since the results agree with FFF theory. This is illustrated in Figure 3D, where the calculated size distribution is plotted for the three run conditions. It can be seen that the two runs with the lower crossflows match, but in the run with the highest cross-flow, there is Analytical Chemistry, Vol. 71, No. 16, August 15, 1999
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Table 3. Filtered Samplea Concentration and Colloidal Metal Fraction element sample conc (µg L-1) colloidal fraction (%)
C 2.5
Fe 900 89
Ni 3.0 12
Mo 0.47 15b
La 1.2 85
Pb 0.6 83
a Sample from Delsjo ¨ba¨cken, Go ¨teborg, 0.45-µm filtered and 1% HNO3 acidified. b Not colloidal but dissolved fraction (molybdate).
Figure 5. Example of a molecular weight frequency function (dm/ dMw) versus molecular weight for carbon (A), nickel (B), and iron (C) from calibration with PSS molecular weight standards.
a shift toward larger colloid sizes. This is probably due to colloidmembrane interactions occurring when too high a cross-flow field is applied. On the basis of these results, a cross-flow of 2.96 mL min-1 was chosen for the subsequent experiments. Size distributions for six elements with different behaviors are presented in Figure 4, together with a nonlinear molecular weight scale. Results from metal quantification are shown in Table 3, where filtered sample total concentration and colloidal fraction are presented. Figure 5 shows an example of a molecular weight distribution for the elements carbon, nickel, and iron. Since the FlFFF retention time has a close to linear dependence on the hydrodynamic diameter,24 the size distributions of the elements show patterns similar to the raw data, while conversion to molecular weight distributions results in a significant change in
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the shape of the fractogram. This is because the molecular weight is exponentially proportional to the retention time and, hence, the diameter. However, molecular weight distributions are more easily interpreted in the smallest size ranges, so that both forms of distribution can be useful in different conditions. If the size distribution approach is used, it is very important to accurately determine the channel dimensions and flows since the size conversion relies on these properties. Results for six elements from a FlFFF-ICPMS determination of an natural sample are shown in Figure 4. The major components of the colloidal material, e.g., carbon and iron, can be determined for different size fractions as well as the elements associated with the specific size fractions. It can be noted that the UV and carbon signal show good agreement. For the selected elements present in Figure 4, it can be seen that colloidal nickel is mainly associated with the carbon peak, but it should be noted that there is a certain overlap between the carbon peak and the first iron peak. Lead is entirely bound to the larger, iron-based colloids while lanthanum shows an intermediate pattern, i.e., associated to both the organic and inorganic colloids but to a larger extent to the smaller size fractions. Molybdenum, on the other hand, shows a completely different pattern, eluting before the colloids, which is interpreted as showing that molybdenum is present as dissolved molybdate. Although most dissolved ions escape through the membrane in the FlFFF channel during the preconcentration step, negatively charged ions like molybdate are partly retained, probably due to charge repulsion from the negatively charged polyether sulfone ultrafilter membrane. CONCLUSIONS The use of FlFFF in combination with a suitable detector, especially ICPMS, is a valuable tool for studying trace element speciation in natural waters as well as in other situations, e.g., industrial processes, where the form in which an element occurs affects its function. The technique is highly flexible and applicable to various size ranges. Furthermore, it has a relatively simple theoretical background which makes it possible to compare theoretically obtained sizes with sizes or molecular weights obtained from calibrations. Using the optimized conditions, detection limits and reproducibility are sufficient for metal speciation in natural freshwater samples. Further advantages of the method, not within the scope of this paper, are the possibility of using enriched isotope materials for studies of adsorption processes. ACKNOWLEDGMENT The authors thank the Knut and Alice Wallenberg Foundation for Financial Support. Valuable discussions with Professors David Dyrssen and David Turner are gratefully acknowledged.
Received for review December 31, 1998. Accepted April 27, 1999. AC981455Y
