Structure and Dynamics of Chromatographically Relevant Fe (III

Article pubs.acs.org/JPCB

Structure and Dynamics of Chromatographically Relevant Fe(III)Chelates Christoph B. Messner,*,† Oliver M. D. Lutz,† Matthias Rainer,† Christian W. Huck,† Thomas S. Hofer,‡ Bernd M. Rode,‡ and Günther K. Bonn† †

Institute of Analytical Chemistry and Radiochemistry, Leopold-Franzens University, Innsbruck, Innrain 80-82, 6020 Innsbruck, Austria ‡ Theoretical Chemistry Division, Institute of General, Inorganic and Theoretical Chemistry, Leopold-Franzens University, Innsbruck, Innrain 80-82, 6020 Innsbruck, Austria S Supporting Information *

ABSTRACT: Immobilized metal ion affinity chromatography (IMAC) is an important chromatographic technique for biomolecules. In order to get a detailed understanding of the hydration of immobilized Fe(III), complexes of Fe(III) with methyl substituted iminodiacetate ([Fe(MSIDA)(H2O)3]+) as well as with methyl substituted nitrilotriacetate ([Fe(MSNTA)(H2O)2]) were simulated in aqueous solutions with the quantum mechanical charge field molecular dynamics (QMCF MD) approach. The simulations were carried out at the Hartree−Fock (HF) level of theory, since cluster calculations at the HF, MP2, and B3LYP levels of theory showed that this method results in a good compromise between computational effort and accuracy. None of the coordinating water molecules were exchanged during the simulation period of 15 ps. The Fe−OH2O bond distances as well as the Fe−OH2O stretching motions differed among the coordinating water molecules, indicating different bond strengths. For the water molecules in the second hydration layer, mean residence times of 2.7 and 1.9 ps were obtained for [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2], respectively. Furthermore, infrared measurements were carried out to characterize the most prominent bond features of aqueous Fe(III)−NTA and to discuss these results in conjunction with the computationally derived frequencies.

1. INTRODUCTION Iminodiacetic acid (IDA) and nitrilotriacetic acid (NTA) are the most common chelating groups in immobilized metal ion affinity chromatography (IMAC). This technique was introduced by Porath et al. in 19751 and is today regarded as a versatile and powerful purification method for biomolecules,2 that is widely used in industry as well as in scientific research. The approach is based on affinities of certain biomolecules toward metal ions that are bound to a chromatographic resin. Such stationary phases are usually silica or agarose beads to which the chelating groups are bound via an alkyl chain. The affinity of a ligand toward such a stationary phase depends mainly on the immobilized ion as well as on the utilized chelating agent,3 whereas the type of resin (silica or agarose) does not significantly influence the selectivity.4 For the purification of phosphoproteins/phosphopeptides, Fe(III) is the most prominent immobilized ion,5,6 but a number of other ions are used as well depending on the application. Although the hydration of the immobilized ions plays a crucial role in their interactions with ligands,7 not much is known about their solvation properties. IDA and NTA metal complexes have been studied by X-ray as well as infrared (IR) spectroscopy solely in the solid state.8−11 © XXXX American Chemical Society

The experimental investigation of the structure and dynamics of such complexes in aqueous solutions is challenging. One difficulty is that ligand exchange processes can occur on extremely short time scales. Experimental methods, however, can often not resolve such short time periods, and therefore, only averaged structures are measured. Another problem, in particular in spectroscopy, is that experimental data are fitted to models in order to obtain results and hence the quality of the results strongly depends on the applied model. Due to such limitations, there has always been a high demand for reliable computational approaches. The theoretical treatment of systems including highly polarizing ions as Fe(III) requires QM methods for a sufficiently accurate description. However, static QM calculations reflect the situation at 0 K and are restricted to systems that are too small to include a representative amount of solvent molecules. Furthermore, such static calculations cannot describe dynamical processes. MD simulations on the other hand are being successfully used to study not only the structure but also the dynamics of Received: May 27, 2014 Revised: September 25, 2014

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dx.doi.org/10.1021/jp505193y | J. Phys. Chem. B XXXX, XXX, XXX−XXX

The Journal of Physical Chemistry B

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r is the distance of a given particle from the center of the QM zone, and r1 and r0 define the inner and outer borders of the smoothing region. This formalism has proven its applicability to anions,15 cations,16 small organic molecules,17 and recently also to metal−organic complexes.18 A detailed description of the QMCF MD approach can be found elsewhere.14,20−22 2.2. Simulation Protocol. Simulations were conducted in cubic boxes with a side length of 39.2 Å, containing Fe(III), MSIDA, or MSNTA and 2000 surrounding water molecules. Periodic boundary conditions were applied, and the density was fixed to 0.997 g/cm3. The temperature was kept around 298 K by using the Nosé−Hoover chain dynamics,23 with a chain length of 5. The Coulombic cutoff was set to 15 Å and longrange electrostatic interactions were taken into account via the reaction field method (ε = 78.36).24 For the integration of the equations of motion, the velocity-Verlet algorithm25 was applied with a time step of 0.2 fs. Fe(III) was placed in the center of the QM region, and the radii of the core and layer zone were set to 1.0 and 6.5 Å, respectively. Smoothing was applied on the atoms of the molecules located in between a distance of 6.3 and 6.5 Å from the center. The well established LANL2DZ ECP26,27 was used for Fe(III), and the 631G(d,p)28 basis set was applied on all C, N, H, and O atoms. A multiplicity of 6 was assigned to Fe(III). All QM calculations were carried out with the Turbomole software.29 The flexible SPC-mTR30 water model was applied, and the non-Coulombic interactions between the particles in the layer zone and the particles in the MM region were described using the generalized Amber force field (GAFF).31 The restrained electrostatic potential (RESP)32 charges of the MSIDA and MSNTA complexes were obtained with Gaussian 09.33 These calculations were performed at the HF level of theory and the same basis sets as used for the simulations. To obtain an appropriate starting structure, an MM simulation was carried out for 100 ps, applying the previously mentioned generalized Amber force field (GAFF). Prior to the 15 ps sampling period, the system was re-equilibrated for 3 ps. To estimate the error associated with the neglect of electron correlation, geometry optimizations of [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] clusters at the HF, B3LYP, and MP2 levels of theory were carried out with the Gaussian 09 software.33 The aforementioned LANL2DZ ECP26,27 and the 6-31G(d,p)28 basis sets were applied on Fe(III) and C, N, H, and O, respectively. The interaction energies (EIA) are listed in Table 1

complexes in solution. However, as such simulations are one of the most computer-time-consuming calculations in computational chemistry, a full QM treatment with nonempirical methods of a reasonably sized system is unfeasible, and therefore, QM/MM MD approaches have become a popular and powerful tool.12,13 These methods are based on a partitioning of the molecular system into regions where QM and MM methods are applied and hence enable the simulation of relatively large systems within reasonable computation times, while still providing high accuracy for the region of main interest. In the quantum mechanical charge field molecular dynamics (QMCF MD) approach, the QM region is relatively large and further partitioned into subregions.14 This has the advantage that metal ions and metal complexes can be simulated without the need of constructing metal ion−solvent potentials, which is a tedious and sometimes even not feasible task, especially for highly charged ions such as Fe(III). Several investigations have shown that the Hartree−Fock method with a double-ζ basis set delivers good results for various systems and is a good compromise between computational effort and accuracy.15−18 In the present study, complexes of Fe(III) with methyl substituted IDA and NTA (MSIDA and MSNTA) serve as model systems for IDA and NTA IMAC phases and were studied within the QMCF MD formalism to get insight into their structure and dynamics in aqueous solution. Furthermore, an IR spectrum of the aqueous Fe(III)−NTA complex was recorded for the first time and the results were compared to the computationally derived frequencies.

2. METHODS 2.1. Simulation Method. In the QMCF MD formalism, the system is divided into a QM region and a MM region, with the QM region being relatively large and further partitioned into an inner (core) region and an outer (layer) region. Coulombic interactions of the MM particles with the QM atoms are evaluated dynamically, and in order to obtain the QM charges, Mulliken population analysis19 is performed in every step of the MD simulation for all QM atoms. The electrostatic interactions of the MM particles with the QM atoms are taken into account via a perturbation term of the core Hamilton operator (eqs 1 and 2), where qJ are the partial charges and M the number of particles in the MM region. HCF = HHF + V ′ M

V′ =

∑

(1)

qJ

r J = 1 iJ

Table 1. Interaction Energies (EIA) in kcal/mol and Various Bond Distances in Å of [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2], Obtained from Geometry Optimizations at Different Levels of Theory

(2)

To ensure a continuous transition of particles migrating between the QM region and the MM zone, a smoothing function is introduced: F jSmooth = S(r )(F jlayer − F jMM) + F jMM

HF

(3) EIA (kcal/mol) Fe−OH2O (Å)

where

S(r ) = 1, S(r ) =

for r ≤ r1

(r0 2 − r 2)2 (r0 2 + 2r 2 − 3r12) (r0 2 − r12)3

Fe−OCOO (Å) Fe−N (Å)

(4)

,

for r1 < r ≤ r0 (5)

S(r ) = 0,

for r > r0

(6) B

B3LYP

[Fe(MSIDA)(H2O)3]+ −1192 −1308 2.12/2.14/2.19 2.16/2.18/2.18

MP2 −1231 2.14/2.14/2.17 1.87/1.87 2.17

EIA (kcal/mol) Fe−OH2O (Å)

1.86/1.86 1.87/1.87 2.18 2.26 [Fe(MSNTA)(H2O)2] −1450 −1568 2.11/2.32 2.16/2.34


1.90/1.91/1.92 2.26

1.91/1.92/1.93 2.21

1.90/1.92/1.94 2.28

−1488 2.13/2.30



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was observed. After a minute, a very fine yellow powder precipitated from the previously clear solution. The product was suction filtered and washed extensively with boiling water and hot ethanol. While the treatment with hot water significantly reduces the yield, this is a necessary measure to quantitatively remove unreacted NTA. Noteworthy, remainders of NTA are easily detected spectroscopically via the intense band at 1745 cm−1. Finally, the purified complex was ground in a mortar and dried at 80 °C for 24 h. The product was then characterized IR spectroscopically in accordance with the report by Rajabalee.11 Furthermore, the pale yellow solution of the complex in water was found to be slightly acidic (i.e., pH 5−6), thus confirming the findings by Brintzinger and Hesse.38 Equally, we were able to observe the precipitation of iron hydroxide upon treatment of the solution with sodium hydroxide or sodium carbonate. The photosensitive yellow complex was found to change color to gray-green upon several days of storage in clear vials. A saturated solution of Fe(III)−NTA in water was obtained by thoroughly mixing the solid complex with water in an ultrasonic bath. After centrifuging the emulsion at 13000 rpm (Eppendorf centrifuge 5415 R) for 15 min, the supernatant was carefully isolated from the solid remainder. The measurement of the solutions was performed on a universal attenuated total reflectance (uATR) equipped PerkinElmer Spectrum 400 FT IR spectrometer (PerkinElmer, Seer Green, U.K.). For that, 20 μL of the sample was directly pipetted onto the diamond/zinc selenide uATR crystal and 20 scans were accumulated at a maximum interferometer retardation of 0.125 cm. The scan speed of the interferometer was chosen as 0.1 cm s−1 in order to optimize the signal-to-noise ratio and the scan-to-scan wavenumber accuracy. The spectrometer’s Jacquinot stop has been fully opened (ø = 8.92 mm), and strong Norton−Beer apodization39 has been applied to the IR spectrum recorded in the region between 1800 and 800 cm−1. A spectrum of pure water was deducted from the solution spectrum in order to facilitate the identification of the analyte’s IR absorptions.

and were calculated with eq 7, where A are the MSIDA and MSNTA ligands. The number of water molecules (n) and the charges of the complexes (m) are 3 and 1 for the MSIDA complex and 2 and 0 for the MSNTA complex. OPT OPT m + − (E E IA = E[Fe(III)A(H + n(H 2OOPT )) Fe(III) + EA 2O)n ]

(7)

The differences between the HF and correlated MP2 method are rather small, indicating a minor influence of electron correlation, which is generally observed for strongly polarized systems.16 Furthermore, the bond distances show small differences between these two methods, and as, unfortunately, simulations of such systems are not feasible at the MP2 level of theory at present, the HF method in combination with a double-ζ basis set resulted as a good compromise between accuracy and computation time. 2.3. Analysis. The structural evaluations are based on radial distribution functions (RDFs) and angular distribution functions (ADFs). The Fe−OH2O power spectra were extracted by analyzing the autocorrelation of the particles’ velocity vectors.34 The velocity autocorrelation function (VACF) C(t) can be written as N

C(t ) =

N

∑i t ∑ j vj⃗(ti)vj⃗(ti + t ) N

N

∑i t ∑ j vj⃗(ti)vj⃗(ti)

(8)

In eq 8, Nt is the number of time origins in time ti utilized for the autocorrelation of all the velocity components v of a particle j, while N is the number of particles under investigation. A Fourier transform (FT) of the obtained data is realized via eq 9: F(ω) =

1 2π

∫0

tcorr

C(t )eiω(ti + t )e−at dt

(9)

Herein, tcorr corresponds to the size of the correlation window and ω is the frequency. e−at with a = 4 is a commonly employed exponential window function which is used in order to avoid a discontinuous cutoff of the signal at t = tcorr.35 For both the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] simulations, C(t) has been computed using a correlation length of 2.0 ps. Avoiding an artificial overcorrelation between the windows,34 the gap size between correlation windows was set to five sample points, corresponding to a simulation time of 5 fs. The IR spectrum of [Fe(MSNTA)(H2O)2] was calculated with the IR Spectral Density Calculator Plugin implemented in the Visual Molecular Dynamics (VMD) software package.36 This tool computes an autocorrelation and a subsequent FT of the time series of the dipole moment, where the dipole moment was calculated from the positions and the respective charges (representative Mulliken partial charges from the simulation) of the atoms. The mean residence time (MRT) of a ligand in a certain hydration layer was calculated with the direct method.37 2.4. Chemicals and Reagents. Nitrilotriacetic acid (>99.5%), ammonium iron(III) sulfate dodecahydrate (>99%), water (LC/MS grade), and ethanol (>96%) were purchased from Sigma-Aldrich Chemical Co., Vienna, Austria, and used without further purification to synthesis. 2.5. Synthesis and IR Spectroscopy of Fe(III)NTA·H2O. The monohydrate of nitrilotriacetatoferrate was synthesized in accordance with an established protocol.38 To a boiling 0.5 M solution of NH4Fe(SO4)2·12H2O, crystalline NTA was added carefully until a change in color from orange to green-yellow

3. RESULTS AND DISCUSSION The starting structures, which were obtained from classical preequilibrations, were 6-fold coordinated complexes which can be denoted as [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2], and these complexes were stable throughout the QMCF MD simulations. Starting QMCF MD simulations from either 5- or 7-fold coordinated structures revealed 6-fold coordinated complexes within a few hundred femtoseconds, justifying the 6-fold coordinated complex as the starting structure. The MSIDA and MSNTA ligands coordinate with the carboxylate groups and the nitrogen atom toward Fe(III), resulting in triand tetradentate coordinations, respectively (Figure 1). Furthermore, three ([Fe(MSIDA)(H2O)3]+) and two ([Fe(MSNTA)(H2O)2]) water molecules occupy the free coordination sites of Fe(III) (Figure 1). The Fe−OH2O RDFs of both complexes are shown in Figure 2. Values of zero between the first and second peaks indicate no exchanges between the first and second hydration layer during the simulation period. The maxima of the first peaks are observed at a distance of 2.07 Å for both complexes (Table 2), which is slightly larger than in the octahedral [Fe(H2O)6]3+ complex, where a Fe−OH2O distance of 2.03 Å was reported by the QMCF MD study of Moin et al.40 The hydration of Fe(III) in aqueous solution has been the subject of many C



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Water molecules form hydrogen bonds to the noncoordinating oxygens of the carboxylate groups. These water molecules are at a distance of 6.0−6.5 Å from the Fe(III) ion. The Fe− MSNTA complex, which has one carboxylate goup more than the Fe−MSIDA complex, shows a feeble peak in the RDF (Figure 2) at this distance. The average MRT values for these water molecules are 1.48 and 1.63 ps for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively, which are lower as for the second shell water molecules. The distances between Fe(III) and the coordinating OCOO and N are 1.93 Å/1.95 Å (MSIDA/MSNTA) and 2.23 Å/2.23 Å (MSIDA/MSNTA), respectively (Table 2). The average Fe(III) partial atomic charges (Mulliken) are 1.73 (1.55−1.85) and 1.71 (1.61−1.86) for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively, indicating a higher ligand−ion charge transfer than observed for Fe(III) in pure water, where a Mulliken charge of 1.85 has been reported for the Fe(III) ion.40 A smaller partial charge results in a smaller contribution of electrostatic interactions to the binding energy and hence, possibly, in the aforementioned elongated Fe−OH2O distances. In order to estimate the covalent character of the ion−ligand interactions, Wiberg bond indices49 were calculated with the Gaussian 09 software,33 using the HF method and the same basis sets as employed during the simulations. Total Wiberg bond indices of 1.63 and 1.66 were obtained for the Fe(III) ions of the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively. The highest contributions to these values have the Fe−OCOO bonds with 0.38 (MSIDA) and 0.35 (MSNTA) per bond. The Fe−OH2O bonds have a much lower covalent character, with values of 0.19 and 0.17 for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively. The Wiberg indices of the Fe−N bonds are quite similar to those of the Fe−OH2O bonds with 0.19 and 0.15 for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively. Although the total Wiberg bond index for Fe(III) is higher for [Fe(MSNTA)(H2O)2] compared to [Fe(MSIDA)(H2O)3]+, the covalent character of each bond is lower, due to the coordination of the additional carboxylate group. The X−Fe−X ADFs of [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2], with X being all atoms within a distance of 3.5 Å to the Fe(III) ion, are shown in Figure 3. The [Fe(MSIDA)(H2O)3]+ complex shows peaks at 80 and 90° and a broad peak between 150 and 180°. The peak at 80° results from the N−Fe−OCOO angle, whereas the peak at 90° is due to the water coordination. The ADF of [Fe(MSNTA)(H2O)2] has its dominating peak at 80°, as it has an additional carboxylate group. However, a large shoulder remains at about 90°, due to the coordination of two water molecules. In an ideal octahedral geometry, four atoms are lying in a plane, however, in the case of [Fe(MSIDA)(H2O)3]+ and especially in that of [Fe(MSNTA)(H2O)2], the two OCOO, Fe(III), and N do not form a planar surface, indicated by a OCOO−Fe−N−OCOO dihedral angle of about 170° ([Fe(MSIDA)(H2O)3]+) and 160° ([Fe(MSNTA)(H2O)2]). In addition, the distance to one coordination site is elongated, as can be shown by calculating the individual Fe−OH2O RDFs of each coordinating water molecule (Figure 4). The water molecules denoted as 3 ([Fe(MSIDA)(H2O)3]+) and 4 ([Fe(MSNTA)(H2O)2]) in Figure 1 exhibit such elongated distances from the coordination center.

Figure 1. Snapshots taken from the QMCF MD simulations of (a) [Fe(MSIDA)(H2O)3]+ and (b) [Fe(MSNTA)(H2O)2] via the VMD Version 1.9.1.36 Cyan, C; white, H; blue, N; red, O; gray, Fe.

Figure 2. Fe−OH2O radial distribution functions and corresponding integration for (a) the [Fe(MSIDA)(H2O)3]+ and (b) [Fe(MSNTA)(H2O)2] complexes in aqueous solution, obtained from the QMCF MD simulations.

Table 2. Various Bond Distances (Peak Maxima of RDFs) in Å of the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] Complexes in Aqueous Solution, Obtained from the QMCF MD Simulations [Fe(MSIDA)(H2O)3]+

[Fe(MSNTA)(H2O)2]

Fe−OH2O (Å)

2.07

2.06


1.93 2.23

1.95 2.23

experimental41−45 and theoretical46−48 studies; however, the work of Moin et al. is best comparable with the present study. A second hydration layer is observed between 3.5 and 4.8 Å in both simulations (Figure 2). The average number of water molecules in this region is 7.9 and 6.4 for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively. The MSNTA complex has a lower number of second shell water molecules than the MSIDA complex, as the additional carboxylate group of the MSNTA ligand occupies coordination sites. The MRTs of the second shell water molecules are 2.7 and 1.9 ps for the [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] complexes, respectively. Both complexes exhibit shorter MRTs than Fe(III) in pure water, where a value of 3.1 ps was reported.40 Furthermore, the second shell water molecules of the [Fe(MSIDA)(H2O)3]+ complex show shorter MRTs compared to the [Fe(MSNTA)(H2O)2] complex. This may be due to the lower number of first shell water molecules, resulting in fewer hydrogen bonds formed between the first and second hydration layers. D



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Figure 5. (a) Experimental spectrum of Fe−NTA, measured with a uATR-FTIR spectrometer and (b) theoretical spectrum derived from the QMCF MD simulation of the [Fe(MSNTA)(H2O)2] complex via autocorrelation of the dipole moment.

Figure 3. X−Fe−X angular distribution functions of (a) ([Fe(MSIDA)(H2O)3]+) and (b) ([Fe(MSNTA)(H2O)2]), with X being all atoms within a distance of 3.5 Å to the Fe(III) ion.

calculations (see the Supporting Information for this data) to help in band assignment. The experimental and theoretical frequencies are listed in Table 3, and they are in reasonable Table 3. Simulation Derived Absorption Band Locations of the [Fe(MSNTA)(H2O)2] Complex in Comparison to the Experimental Values Obtained for Aqueous Fe(III)−NTA Complexes, Measured with a uATR-FTIR Spectrometer mode

QMCF-MD (cm−1)

experiment (cm−1)

νCOO,as νCOO,s νCN δCOO

1870 1540 1180 1015

1616 1388 1112 914

agreement. The theoretically obtained frequencies are blueshifted, which may be a result of the relatively strong influence of electron correlation on the C−N as well as on the C−OCOO bond. No IR measurements of aqueous Fe(III)−IDA complexes have been carried out, since bis-IDA iron complexes ([Fe(IDA)2]−) are already formed in a 1:1 solution of Fe:IDA.8 Figure 6 shows the vibrational density of states, resembling the Fe−OH2O stretching motions of [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] derived via VACFs. The bands in the 450 cm−1 region may result from the Fe−OH2O stretching frequencies, which is in agreement with a Raman spectroscopic study of concentrated Fe−NTA solutions, where a band at 410−416 cm−1 was observed which was assigned to a coupling of FeO stretching vibrations and ligand deformations.52 The Fe−OH2O stretching motions of the studied complexes are slightly lower as for Fe(III) in pure water (513 cm−1),40 indicating reduced bond strengths and higher exchange rates. However, the exchange kinetics of Fe(III) are in general very slow, as water substitution rate constants of 1.6 × 102 s−1 have been observed experimentally for the [Fe(H2O)6]3+ complex.53 This is far beyond the time scale of our simulations, explaining why no water exchange processes have been observed in our study. Furthermore, the Fe−OH2O stretching frequencies differ among the coordinating water molecules (Figure 6), as can be expected due to the different Fe−OH2O distances (Figure 4). The frequencies of the water molecules with the elongated

Figure 4. Fe−OH2O radial distribution functions for the water molecules denoted as (a) 1/2/3 ([Fe(MSIDA)(H2O)3]+) and (b) 4/5 ([Fe(MSNTA)(H2O)2]), obtained from the QMCF MD simulations.

In IMAC stationary phases, the IDA and NTA group is bound to the resin via an alkyl chain, which is modeled in our simulations by the methyl group. In the [Fe(MSNTA)(H2O)2] complex, the methyl group does not have an influence on the hydration of the Fe(III) ion. In the case of the [Fe(MSIDA)(H2O)3]+ complex, however, the water molecule denoted with 1 (Figure 1) is influenced by the nucleic repulsion of the methyl group, resulting in N−Fe−OH2O angles of ∼100° and hence a further deviation from the ideal octahedral structure. Figure 5 shows the IR spectrum of the [Fe(MSNTA)(H2O)2] complex derived from the simulation as a comparison to the experimental spectrum of aqueous Fe−NTA, measured with a uATR-FTIR spectrometer. The assignment of the absorption bands in the experimental spectrum was done by comparing them with the bands of the solid Fe−NTA complex11 as well as of other metal−NTA chelates.50,51 The theoretically observed frequencies were assigned by performing additional calculations, solely considering parts of the molecule in the autocorrelation functions (see the Supporting Information for these spectra). Furthermore, the theoretical spectrum was compared with frequencies obtained via static E



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of analytical techniques may subsequently promote the development of new and innovative analytical approaches.

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ASSOCIATED CONTENT

S Supporting Information *

Theoretical spectra obtained by solely considering parts of the [Fe(MSNTA)(H2O)2] complex in the autocorrelation of the dipole moment. Harmonically approximated spectroscopic data for [Fe(MSNTA)(H2O)2] obtained from static calculations at the HF, B3LYP, and MP2 levels of theory. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Notes

Figure 6. Fe−OH2O frequencies obtained via VACFs and subsequent FT from the QMCF MD simulation of (a) [Fe(MSIDA)(H2O)3]+ and (b) [Fe(MSNTA)(H2O)2] in aqueous solution.

The authors declare no competing financial interest.

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distances (3 and 4) are slightly shifted to lower values, indicating a reduced bond strength. This can be related to steric effects of the MSNTA and MSIDA ligands and implies a higher tendency to undergo exchange processes. Therefore, these water molecules may be preferentially exchanged with other ligands (e.g., phosphorylated macromolecules) in IMAC approaches. The electronic structure (partial charges) and the Fe−OH2O force constants are similar for [Fe(MSNTA)(H2O)2] and [Fe(MSIDA)(H2O)3]+. Hence, differences in performance of Fe(III)−IDA and Fe(III)−NTA stationary phases may be attributed to the different number of free coordination sites (steric effects) as well as to the net charge of the complex, which is +1 and 0 for Fe(III)−IDA and Fe(III)−NTA, respectively (at neutral pH). This is in agreement with experimental observations, as phosphorylated macromolecules are equally enriched on Fe(III)−IDA and Fe(III)−NTA agarose.54 However, very acidic proteins were found to have a stronger affinity toward Fe(III)−IDA stationary phases, which may be a consequence of the positive net charge and, hence, the pseudoanion exchange processes.54

4. CONCLUSION [Fe(MSIDA)(H2O)3]+ and [Fe(MSNTA)(H2O)2] were studied by the well established QMCF MD approach, and due to the ab initio treatment and the substantially large QM region, accurate data were obtained. Furthermore, IR measurements of aqueous Fe(III)−NTA complexes were performed and the results were in reasonable agreement with the computationally derived frequencies. Deviations of the theoretical from the experimental values may be attributed to the neglect of electron correlation. However, a treatment of such systems with correlated methods is still not feasible with today’s computational facilities. This work serves as an example of how theoretical studies and spectroscopic measurements can be combined in order to improve the interpretation of structure and binding in complex systems in solution. Furthermore, the findings of this work emphasize the significance of theoretical approaches, as they can help to understand experimental approaches such as IMAC. This is of particular interest, since an improved understanding F



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Structure and Dynamics of Chromatographically Relevant Fe (III

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