Short communication
Experimental and Theoretical Study on the Complexation of Beauvericin with the Ammonium Cation
Emanuel Makrlik,1'* Petr Toman2 and Petr Vanura3
1 Faculty of Environmental Sciences, Czech University of Life Sciences, Prague, Kamyckâ 129,
165	21 Prague 6, Czech Republic
2 Institute of Macromolecular Chemistry, Academy of Sciences of the Czech Republic, Heyrovského sq. 2,
162 06 Prague 6, Czech Republic
3 Department of Analytical Chemistry, Institute of Chemical Technology, Technickâ 5,
166	28 Prague 6,Czech Republic
* Corresponding author: E-mail: makrlik@centrum.cz Received: 07-11-2011
Abstract
From extraction experiments and y-activity measurements, the extraction constant corresponding to the equilibrium NH+(aq) + NaL+ (nb) o NH4L+(nb) + Na+ (aq) taking place in the two-phase water-nitrobenzene system (L = beauvericin; aq = aqueous phase, nb = nitrobenzene phase) was evaluated as log Kex (NH+, NaL+) = 1.5 ± 0.1. Further, the stability constant of the NH4L+ complex in water-saturated nitrobenzene was calculated for a temperature of 25 °C as log Pnb (NH4L+) = 4.6 ± 0.2. Finally, by using quantum mechanical DFT calculations, the most probable structure of the NH4L+ cationic complex species was derived. In this complex having C3 symmetry, the ammonium cation NH+ is bound by three strong linear hydrogen bonds to the three corresponding oxygen atoms of the parent beauvericin ligand L. The interaction energy of the resulting complex NH4L+ was found to be -828.8 kJ/mol, confirming the formation of the considered complex NH4L+.
Keywords: Beauvericin, ammonium cation, complexation, water-nitrobenzene system, extraction and stability constants, DFT calculations, complex structure
1. Introduction
Beauvericin (abbrev. L; see Scheme 1) is a cyclic hexadepsipeptide with alterning methyl-phenylalanyl and hydroxyl-iso-valeryl residues.1 For the first time, beauvericin was isolated from the fungus Beauveria bassiana. It was further found that the antibiotic beauvericin was produced by many other fungi (e. g., by some Fusarium species) so that this compound, which belongs to the enni-atin family, can occur very often in grain contaminated with these fungi.2-4
The dicarbollylcobaltate anion (DCC-)5 and some of its halogen derivatives are very useful reagents for the extraction of various metal cations (especially Cs+, Sr2+, Ba2+, Eu3+ and Am3+) from aqueous solutions into a polar organic phase, both under laboratory conditions for purely theoretical or analytical purposes,6-22 and on the technological scale for the separation of some high-activity iso-
Scheme 1. Structural formula of beauvericin (abbrev. L).
topes in the reprocessing of spent nuclear fuel and acidic
radioactive waste.23,24
Recently, the complexation of the ammonium cation with valinomycin has been investigated in methanolic
medium.25 On the other hand, in the current work, the sta-
in the aqueous and nitrobenzene phases, respectively. For
bility constant of the beauvericin complex with NH+ (i. e., the constant Kex (NH+, Na+) one can write5,28,29 NH4L+) was determined in the organic phase of the waterlog Kex(NH+, Na+) = log KNH+ - log K^
nitrobenzene extraction system. Moreover, applying quantum mechanical DFT calculations, the most probable structure of the NH4L+cationic complex species was predicted.
2. Experimental
Beauvericin (L; see Scheme 1) was purchased from Aldrich. Cesium dicarbollylcobaltate (CsDCC) was synthesized by means of the method published by Hawthorne et. al.26 The other chemicals used (Lachema, Brno, Czech Republic) were of reagent grade purity. A nitrobenzene solution of hydrogen dicarbollylcobaltate (HDCC)5 was prepared from CsDCC by the procedure described else-where.27 The equilibration of the nitrobenzene solution of HDCC with stoichiometric NaOH, which was dissolved in an aqueous solution of NaCl (0.20 mol/L), yielded the corresponding NaDCC solution in nitrobenzene. The radionuclide 22Na+ (DuPont, Belgium) was of standard ra-diochemical purity.
The extraction experiments were carried out in 10 mL polypropylene test-tubes with polypropylene stoppers: 2 mL of an aqueous solution of NH4Cl (1 x 10-2 to 3 x 10-2 mol/L) and microamounts of 22Na+ were added to 2 mL of a nitrobenzene solution of L and NaDCC, whose initial concentrations varied also from 1 x 10-2 to 3 x 10-2 mol/L (in all experiments, the initial concentration of L in nitrobenzene, CLin,nb, was equal to the initial concentration of NaDCC in thLis medium, CNina,nDbCC). The test-tubes filled with the solutions were shaken for 2 h at 25 ± 1 °C, using a laboratory shaker. Then the phases were separated by centrifugation. Afterwards, 1 mL samples were taken from each phase and their y-activities were measured by means of a well-type NaI(Tl) scintillation detector connected to a /-analyzer NK 350 (Gamma, Budapest, Hungary).
The equilibrium distribution ratios of sodium, DNa, were determined as the ratios of the measured radioactivities of 22Na+ in the nitrobenzene and aqueous samples.
3. Results and Discussion
With respect to the results of previous papers,5,28,29 the two-phase water-NH4Cl-nitrobenzene-sodium dicar-bollylcobaltate (NaDCC) extraction system can be described by the following equilibrium
(2)
NH+ (aq) + Na+ (nb) «
NH+ (nb) + Na+ (aq); Kex (NH+, Na+)
(1)
where KNi H+ and KNi a+are the individual extraction constants for NH4+ an4 d Na+, respectively, in the water-nitrobenzene system.28 Knowing the values log KNH+ = -4.7 28 and log KNa+ = -6.0,28 the exchange extraction constant Kex (NH+, Na+) was simply calculated from Eq. (2) as log Kex (NH+, Na+) = 1.3.
Previous results30-33 indicated that the two-phase water-NH4Cl-nitrobenzene-NaDCC-L (beauvericin) extraction system (see Experimental), chosen for determination of the stability constant of the complex NH4L+ in water-saturated nitrobenzene, can be characterized by the main chemical equilibrium
NH+(aq) + NaL+ (nb) « NH4L+ (nb) + Na+(aq); Kex (NH4+, NaL+)
(3)
with the respective equilibrium extraction constant Ke (NH+, NaL+):

[nh.l+urtl,
[NH4+]aij[NaL+]nh
(4)
with the c,orresponding exchange extraction constant Kex (NH+, Na+); aq and nb denote the presence of the species
It is necessary to emphasize that L is a considerably hydrophobic ligand, practically present in the nitrobenzene phase only, where this ligand forms - with NH4+ and Na+ - the relatively stable complexes NH4L+ and NaL+.
Taking into account the conditions of electroneutral-ity in the organic and aqueous phases of the system under study, the mass balances of NH4+ and Na+ cations at equal volumes of the nitrobenzene and aqueous phases, as well as the measured equilibrium distribution ratio of sodium, DNa = [NaL+]nb / [Na+]aq, combined with Eq. (4), we get the final expression for Kex(NH4+, NaL+) in the form
(5)
where CNinH,aqCl is the initial concentration of NH4Cl in aqueous phase a4 nd CiNna,nDbCC denotes the initial concentration of NaDCC in the organic phase of the system under consideration.
In this study, from the extraction experiments and y-activity measurements (see Experimental) by means of Eq. (5), the following value of the constant Kex (NH+, NaL+) was determined as log Kex (NH+, NaL+) = 1.5 ± 0.4.
Furthermore, with regard to previous papers,30-33 for the extraction constants Kex (NH+, Na+) and Kex (NH+, NaL+) defined above, as well as for the stability constants of the complexes NH4L+ and NaL+ in nitrobenzene saturated with water, denoted by finb (NH4L+) and jinb (NaL+), respectively, one can formulate
log ¿3nb (NH4L+) = log (NaL+) + log Kex (NH+,NaL+) - log Kex (NH4+,Na+)
(6)
Using the constants log Kex(NH+,Na+) and log Kex(NH+,NaL+) given above, the value log finb (NaL+) = 4.4 ± 0.1,34 and applying Eq. (6), we obtain the stability constant of the NH4L+ complex in water-saturated nitrobenzene at 25 °C as log ¡3nb (NH4L+) = 4.6 ± 0.2. This means that the stability constants of the cationic complex species NH4L+ and NaL+ in the mentioned nitrobenzene medium are comparable. In this context it should be noted that the stability constant of the complex NH4L+, where L is valinomycin, in nitrobenzene saturated with water is log £nb(NH4L+) = 8.4 ± 0.1.35 Thus, in this medium, the stability of the NH4L+ complex (L = valinomycin) is substantially higher than that of the species NH4L+ (L = beau-vericin) under study.
The quantum mechanical calculations were carried out at the density functional level of theory (DFT, B3LYP functional) using the Gaussian 03 suite of programs.36 The 6-31G(d) basis set was used and the optimizations were unconstrained. In order to increase the numerical accuracy and to reduce oscillations during the molecular geometry optimization, two-electron integrals and their derivatives
were calculated by using the pruned (99,590) integration grid, having 99 radial shells and 590 angular points per shell, which was requested by means of the Gaussian 03 keywords "Int = UltraFine".
Although a possible influence of a polar solvent on the detailed structures of L and NH4L+ complex species could be imagined, our quantum mechanical calculations in similar cases, performed in an analogous way, showed very good agreement of experiment with theory.37-44
In the model calculations, we optimized the molecular geometries of the parent beauvericin ligand L and its complex species with NH+. The optimized structure of the free ligand L with C3 symmetry is illustrated in Figure 1.
In Figure 2, the structure obtained by the full DFT optimization of the NH4L+ complex having also C3 symmetry is depicted, together with the lengths of the corresponding hydrogen bonds (in A ; lA = 0.1 nm). As follows from this figure, the complexation with the NH4+ cation changes the overall shape of the parent ligand L only slightly. In the resulting NH4L+ cationic complex species, which is most energetically favoured, the "central" cation NH4+ is bound by three strong linear hydrogen bonds to the corresponding carbonyl oxygen atoms (1.72, 1.72 and 1.72 A) of the parent beauvericin ligand L.
Figure 1. Two projections of the DFT optimized structure of free beauvericin ligand L [B3LYP/6-31G(d)].
Figure 2. Two projections of the DFT optimized structure of the NH4L+ complex [B3LYP/6-31G(d)]. Hydrogen bond lengths of NH+ to the three corresponding carbonyl oxygens of L are 1.72, 1.72 and 1.72 Â.
Finally, the interaction energy, E(int), of the NH4L+ complex [calculated as the difference between the pure electronic energies of the complex NH4L+ and isolated L and NH+ species: E(int) = E(NH4L+) - E(L) - E(NH+)] was found to be -828.8 kJ/mol, which confirms the formation of the cationic complex NH4L+.
4. Conclusions
In summary, we have demonstrated that a complementary experimental and theoretical approach can provide important information on the beauvericin ligand L complexation with the ammonium cation. From the experimental investigation of the resulting complex NH4L+ in the two-phase water-nitrobenzene extraction system, the strength of the considered NH4L+ cationic complex species in nitrobenzene saturated with water was characterized quantitatively by the stability constant, log finb (NH4L+) = 4.6 ± 0.2 (for a temperature of 25 °C). By ming the theoretical quantum mechanical DFT calculations, the structural details of the NH4L+ complex, such as position of the NH+ cation with regard to the parent beau-vericin ligand L as well as the significant interatomic distances within the complex species under study, were obtained.
5. Acknowledgements
This work was supported by the Grant Agency of Faculty of Environmental Sciences, Czech University of Life Sciences, Prague, Project No.: 42900/1312/3114 "Environmental Aspects of Sustainable Development of Society," by the Czech Ministry of Education, Youth and Sports (Project MSM 6046137307) and by the Czech Science Foundation (Project P205/10/2280). The computer time at the MetaCentrum (Project LM 2010005), as well as at the Institute of Physics (computer Luna/ Apollo), Academy of Sciences of the Czech Republic, is gratefully acknowledged.
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Povzetek
Z raziskavami ekstrakcije in meritvami y-aktivnosti smo določili konstanto ekstrakcije za ravnotežje NH+(aq) + NaL+ (nb) o NH4L+(nb) + Na+ (aq) v dvofaznem sistemu voda-nitrobenzen (L = beauvericin; aq = vodna, nb = nitrobenzen), ki znaša Kex (NH+, NaL+) = 1.5 ± 0.1. Nadalje smo v vodi, nasičeni z nitrobenzenom, pri 25 °C določili konstanto stabilnosti kompleksa NH4L+, log Pnb (NH4L+) = 4.6 ± 0.2. Z uporabo kvantno mehanskih DFT računov smo predvideli najbolj verjetno strukturo NH4L+ kationskega kompleksa.