Biogeochemistry of Chelating Agents - American Chemical Society

Chapter 5

Speciation of Aminopolycarboxylate and Aminophosphonate Metal Complexes by AEX ICP-MS in Environmental Water Samples Downloaded by TUFTS UNIV on November 28, 2015 | http://pubs.acs.org Publication Date: July 21, 2005 | doi: 10.1021/bk-2005-0910.ch005

Adrian A. Ammann Analytical Chemistry of the Aquatic Environment, ΕAWAG, Swiss Federal Institute for Environmental Science and Technology, P.O. Box 611, CH-8600 Dübendorf, Switzerland

The widespread diverse applications, the persistency and fate of chelating agents in the environment are inevitably associated with metal binding. Depending on the metal bound to a chelating agent, such a species may be well or not at all bio-degradable. So biodegradability of free chelator is not a criterion to assess whether chelating agents are dangerous pollutants or not. Direct and reliable observation of single metal chelates is required in order to gain adequate knowledge on persistency, mobility, chemical transformation, uptake and effects of metal chelates. This is provided by sensitive species specific analytical methods as the one described here. Determination of so called total chelator concentrations is not intended to and cannot provide such a realistic picture since free chelators do not exist in the environment and speciation cannot be calculated from the total amount of chelators. A low hydrophobic narrow bore anion exchange column coupled to an ICP-MS provided new exceptional selectivity and sensitivity in metal speciation analysis of anionic synthetic and biogenic chelates and oxo-species in environmental waters. Moreover the selectivity produced individual metal separation patterns unique for many chelators which make it possible to identify them independent of retention times.

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© 2005 American Chemical Society In Biogeochemistry of Chelating Agents; Nowack, B., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2005.

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Downloaded by TUFTS UNIV on November 28, 2015 | http://pubs.acs.org Publication Date: July 21, 2005 | doi: 10.1021/bk-2005-0910.ch005

Introduction Chelating agents form stable metal complexes that keep metals in solution and prevent the formation of undesired aggregates and precipitates. This sequestering effect leads to their widespread application in consumer goods, fertilizers and production processes. In cases of insufficient degradation or elimination chelators enter environmental water bodies through waste water treatment plants (1,2,3) and through soil water (4) where they efficiently increase the mobility of heavy metals and dominate its speciation in the final receiving waters. Investigations in to the biodégradation of free chelators have revealed structural aspects (secondary or primary amine) that are more susceptible to degradation but these also give rise to less stable metal chelates (5). Once released to the environment, uncontrolled and innumerable conditions for species formation and partial degradation occur, which make metal chelator species and chelator degradation unpredictable. For this reason research on environmental behavior of metals and of chelating agents are closely interrelated. Speciation of chelating agents in the environment cannot be carried out without considering complexing metals (6) and environmental metal speciation cannot be done without considering strongly binding biogenic or anthropogenic chelators (i). Accordingly, this requires speciation methods that can determine individual chelating agents including the metals bound to it (species). One example that illustrates how single metal chelates are related to real world problems is the non biodegradability of metal-EDDS (metal = Co, Cu, N i , Hg, Zn) species (7) whereas EDDS itself is a biogenic and biodegradable chelator recommended to replace EDTA in many applications. So, unfortunately, biodegradability of free or weakly bound chelator is not a criterion to assess whether chelating agents are dangerous pollutants or not. Whatever the source of EDDS, i f non-biodegradable metal complexes are formed before degradation, EDDS certainly will contribute to the mobility of Cu, N i , Zn and Hg. Research in this area strongly depend on analytical speciation methods (6). In order to meet the research requirements for multiple aspects of environmental chelation chemistry (6) a selective anion exchange (AEX) procedure was developed to separate metal chelates of several of the most often used chelators. With respect to above problems it makes sense to classify published analytical methods according to the ability to determine individual metal chelates, that is species specific (speciation) and non species specific (non speciation) methods. They were developed in different times and nowadays fulfill different tasks. Both methods use diverse separation techniques such as

In Biogeochemistry of Chelating Agents; Nowack, B., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 2005.

110 gas chromatography (GC), capillary electrophoresis (CE), and several liquid chromatographic (LC) methods such as ion exchange chromatography (IEC) and ion pair chromatography (IPC) on reversed phase HPLC columns. These techniques are linked to diverse detectors according to the analytical task the system has to provide. Traditionally, non speciation analyses are conveniently performed by non specific but universal detectors like flame ionization, U V , and conductivity etc. However, research and validation requires more species specific information which can be obtained from molecule or element specific detectors e.g. mass spectrometers.


Non species specific analytical methods Non speciation methods are used to determine so called "total" concentration of a chelator. Total implies here that a chelating agent can be isolated and determined quantitatively irrespective of its binding to metals, which is at least a delicate task and in some cases even impossible (see below). Weiss (8) and Schwedt (9) were among the first who separated free polyaminocarboxylates (NTA, EDTA, DTPA), polyaminophosphonates (ATMP; EDTMP, DTPMP), polyphosphonic and polyphosphinic acids. These non UV-absorbing chelators were transformed into UV-absorbing iron chelates which were successfully detected by these authors. Later this derivatisation step became the most often used for U V detection frequently in combination with ion pair separation procedures (a detailed review is given by Schmidt & Brauch in this book). However there are several crucial points which need careful attention and optimization (10). Environmental samples especially, usually contain the most refractory (non biodegradable) metal bound chelators. Among them are metals that are not readily exchangeable. But all the metals bound to a particular chelator including the slow reacting have to be quantitatively exchanged (11) by iron since free chelators are exclusively used for calibration. Beside different reaction velocities, the stability of die Fe-chelates has to be considered, e.g. Fe(III)EDTA is photo-labile (12,13) and is destroyed by day light or intense lab light. In fact differences in reaction speed (14) and reaction conditions (15) have been used to differentiate metal chelates. The derivatisation step was also performed after separation (16) as post column in line reaction with higher detection limits (DL) (> 60 nmolL" ) because of dilution and shorter reaction times available to slowly reacting metal chelates. The most severe limitation of this derivatisation arises from the fact that some chelates do not form complexes with iron; either because they are not stabile enough or they undergo side reactions (17). Additionally, the method cannot tell the analyst i f there are metal (e.g. Crin, Com) complexes in the sample that will not exchange with iron (11) except under condition that destroys the chelator. Usually not all 1


Ill these conditions are checked for in "total" chelator determinations and it can be concluded that not in all cases a "total" value corresponds to the real total amount of a chelator.


Spedes specific analytical methods Gains in sensitivity and information on composition and structure are the main reasons for the use of mass spectrometers as detectors in chelator separation. Some information on elemental and isotopic composition of chelates especially those containing transition or heavy metals are most conveniently obtained from inductively coupled plasma mass spectroscopy (ICP-MS) (18,19). The high temperature in the plasma (5000-7000°) produces ultra fast and efficiently elemental ions almost independent of any molecule binding. As the elements are atomized information on molecule structure is lost except in cases where a structure specific separation is coupled to the ICP-MS (20). Such hyphenated techniques (21) can identify and quantitate known metal chelates based on reference materials. Structural details of unknown chelates might be obtained from low energy ion source MS like e.g. electro spray ionization M S (ESI-MS) (22). But ESI mass spectra of metal chelates can be extremely complicated caused by metal redox reaction and/or many additional fragment ations of the organic molecule catalyzed by the metal. Among the separation techniques used C E is a newer method providing superior separation efficiency. Application of C E connected to non specific detectors is reviewed extensively in the following two papers. However, the inherently higher D L in CE is due to the low flow (-10 nLmin" ) available. But such low flow rates are also a problem for sample introduction systems of MS. There are no nebulizers that can directly convert such a low flow into an aerosol. A n additional flow (make-up or sheath flow ~ 10-20 μίνηιίη" ) is required which has caused additional problems in the past (24). Recent developments (25) in C E coupling to ICP-MS, however, seem to omit these problems since D L in the range 10-100 nmolL" for determination of metallothioneins (26,27) and Se- and As-species (28) have been reported. No investigation of metal chelates by CE-ICP-MS has been published so far but EDTA was used as a competitive ligand in determining rare earth metals fulvic acid binding constants by this speciation method (29). Little is known (30 31) on the applicability of CE-ESI-MS to metal chelate speciation. 1

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IÇ is an older and established separation technique that was more often applied in speciation analysis. Coupling IC to ESI-MS with a carbonate eluent as commonly used in IC was preferred over other inorganic eluents since carbonate can be chemically suppressed providing much better signal to noise ratios (32,33). However carbonate eluents have two main drawbacks. Firstly, the


112 anionic MeEDTA species eluted at similar retention times like the high concentrated anions CI", N 0 " , H C 0 " and SO4 " and secondly, a carbonate eluent needs a high pH (>9) to reduce the amount of H C 0 " in order to increase C 0 " and the elution strength. But at such high pH's some metal (Fe, A l , Zn) complexes are no longer stable. Other eluents can be used in IC coupled to ICP-MS. NH4NO3 was preferably applied because it exhibits the best plasma compatibility and has the lowest interferences with elements to be detected. A l l other eluents usually used in IEC and IPC would produce deposits in the MS and interferences that prevent sensitive determination of some metals. NH4NO3 was used to separate CniiEDTA in a Crlii/Crvi speciation (34). As metal chelates exhibit a high affinity for polymeric column materials, high eluent salt concentrations have to be applied. This generates a high salt load in the plasma that can be lowered by chemical suppression of the eluent. On the other hand, the plasma load was also considerably reduced by utilizing a micro bore system that requires lower eluent concentrations and lower flow rates which introduce lower amounts of salt into the plasma. This setup was specifically developed (35) to determine low concentrated metal chelates of commonly used chelators. 2

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Materials and Methods The chromatography and the coupling to ICP-MS have been described (35) in details. In brief, an all PEEK micro bore system and a low capacity anion exchange column ( A S H , 250 χ 2 mm, Dionex) was used. The high selectivity observed was due to lowest hydrophobicity and high efficiency in mass transfer of the column material. As an eluent NH4NO3 was chosen because of its ideal plasma compatibility and eluent properties (35). The pH of the eluent was adjusted (HNO3/NH4OH) and controlled at the column exit. In a direct coupling without splitting the eluent flow (440 μίΛηίη" ) was optimized for fast separation and to the highest nebulizer to plasma mass transfer. On column sample preconcentration was achieved either by a sample loop or, for larger volumes (1-5 mL), by injection on a preconcentrator column (AGI 1, 50x4 mm, Dionex). A l l given stability constants (1=0.1, 25°) and p K values were taken from (36) if not otherwise indicated. 1

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Results and Discussion The investigations (35) of free chelators like E D A , H E D A , N T A , EDDS, EDTA, CDTA, DTPA, NTMP, EDTMP and their metal (Fe, Mn, Co, N i , Cu, 3

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113 Zn, Cd, Pb) complexes comprised of several aspects of complexation chemistry (charge density, equilibrium stability constants, reaction kinetics) and environmental concern. Application of a particular isocratic eluent concentration provided the high selectivity for the separation of several metal chelates that do not differ by more than one charge unit. A pure anion exchange mechanism was shown to be active in separation (35). Multiply charged species required increasing eluent concentrations of about 32 mmolL" N 0 " per formal charge unit. It allowed separation of several metal species of the same chelator which was used for chelator identification (see below). However, in environmental waters chelators were found that differ by more than one charge unit. In order to separate as many as possible of these higher charged chelates in the same run, gradient elution had to be applied. It was found that gradients (20-200 mmolL" NH4NO3, 0-8 min) up to such high concentrations were well tolerated by the plasma and they separated in the same run anionic species that differ approximately by five charge units. The selectivity was still excellent because the gradient narrowed the peak width considerably so that small structural changes which affected charge density resulted in retention time (t ) shifts. The selectivity by gradient elution was further investigated using standard solutions of MeEDTA, MeCDTA and MeDTPA. The separation achieved for the metal chelates (Me= Cu, Ni, Zn, Mn, 0.1 μπιοΐΐ/ each) of these three most often applied polyaminocarboxylates is shown in Figure 1. The chelates of each ligand are well separated from each other. This is even the case for MeEDTA ' and MeCDTA * which both bear the same negative formal charge. A n explanation for the elution order is given in (35). Moreover, the four metals resulted in individual metal separation patterns which are unique for each chelator. This allows identification of each ligand structure independently of the actual t by simply measuring only three commonly occurring metals (e.g. Cu, Zn and Mn). It was also noticed that a loss of a carboxylic or a phosphonate group altered the metal chelate separation pattern as well, beside a large shift in retention time. 1

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Influence of pH As a master variable the pH strongly influences die stability and other properties of metal species. However, die pH of the so far most often used eluents in IC (HN0 , C 0 " and OH") cannot be altered without changing the eluent strength. However a change in eluent strength vastly changes ion chromatography. This is not the case for the eluent chosen. Here the eluent pH can be adjusted in the range of highest metal complex stabilities (37,7) without changing the eluent strength. This means that the pH of the NH4NO3 eluent can be used as an independent variable to control the separation selectivity. 2

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Figure L Separation and ICP MS detection ofpolyaminocarboxylate metal chelates. The sample was injected by a loop (250 pL) and a NH4NO3 gradient (20-150 mmoUL' ) was applied. 1

Tunable Selectivity Since the pH of this eluent can be varied in the most relevant range (pH 6-8) conditions were established that separated free DTPAH " (pK =8.6) from EDTAH " and CDTAH " at pH 8.0. Such a partial deprotonation was also helpful in separation of MeEDTA " from MeEDDS ". Since EDTA and EDDS are built from the same number and type of atoms (constitution isomers) similar size and charge density can be expected. However, the deprotonation of a coordinated H 0 molecule differs by one pK-unit which was used to separate CuEDTA from CuEDDS at pH = 8.7 (see Figure 2). Since the p K of their N i analogues is higher their separation is only slightly improved by the applied increase in pH. The protonation degree is not the only reason for different charge densities of metal complexes. Oxidation state, composition and stereo electronic effects cause also differences in charge densities which result in different t (35). This is illustrated in Figure 3 for MeEDTA " (Me = Fe, Co, N i , Cu, Cd, Pb). Retention factors (k') were lower (=shorter ta) for die larger metals (Cd, Pb) complexes having lower charge densities, and there was almost no pH 3

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115 dependency of k' except for Fe(III)EDTA which exhibited a large increase in k' between pH 7 and 8.

3.0E+03 «»

H 8 1

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NiEDTA NiEDDSss *

2.0E+031

NiEDTA

CuEDTA CuEDDSss

NiEDDSss

CuEDTA i\ pKa=U.4 II

CuEDDSss pKa=10.4


1.0E+03

\ 14r66

100

100 200 300 400 Retention time [sec]

200 300 400 Retention time [sec]

Figure 2. Separation of CuEDTAfromthe natural enantiomer CuEDDSss (both 0.08 μηοΐϋ ,100 pL injected) occurs by a gradient run with an eluentpH closer to the pK of the coordinated H 0 in the molecules. 1

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This is in agreement with spectroscopic investigations (38) which showed that the coordinated H 0 molecule in these complexes can be deprotonated below pH=8 only in case of Fe(H 0)EDTA (pK =7.5). The p K of all other species are above 10 so they do not change charge density by varying the pH below 8. 2

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Figure 3. Variation of retention factors k* ( ~(t-t )/t ) at different eluent pH. 0

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116 DL and Sensitivity The sensitivity depends on a variety of factors, mainly on species stability, preconcentration capability of the procedure, mass transfer efficiency of the sample introduction system, salt load in the plasma and the isotope measured. The chelate stability is an important prerequisite since there is competition in metal binding between the chelator and the column. With die column used chelate stabilities pK>10 are required for concentrations below micro molar. The greater the stability, the lower the amount of the metal taken up by the column and the lower the concentration that can be determined. The ion content is another important factor in species preconcentration. The higher the concentration e.g. of doubly charged anions C 0 " and S0 " in a sample, the lower the preconcentration capability of a column. Ion exchange is especially suited to preconcentrate selectively multiply charged ionic species by a factor 10-100. As in this case, the matrix is separated and at the same time exchanged by the eluent with approximately similar ionic strength and a minimal shift in pH (ΔρΗ

Biogeochemistry of Chelating Agents - American Chemical Society

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