Introduction

The biological activity of organotin(IV) compounds and their use as metallopharmaceuticals has been reported [¹-¹³]. A tremendous research is directed towards the design of non-platinum chemotherapeutics with the aim to optimize the features of classical platinum drugs constituting the basic cisplatin framework viz. their toxic side effects, inherent intrinsic resistance and high cost [¹⁴]. Diorganotin(IV) complexes are acting as potential antitumours. They exhibit promise activity against many tumour cell lines [¹⁵-¹⁹]. Numerous diorganotin(IV) derivatives have been found to exhibit high in vivo cytotoxicity against P388 lymphocytic leukaemia.

The antitumour action mechanism is still unclear, although some evidence is consistent with their binding directly to DNA. The antituour activity of the organotin(IV) complex R₂SnX₂L is depending on the group attached to the tin atom, leaving groups (X) and the coordinated ligand (L). The coordinated ligand favours in some way the transport of the drug into cells, and the antitumour activity would be exerted by the diorganotin(IV) ion released from the complex [²⁰]. The released diorganotin(IV) species would interact with nucleic acids, similarly as in the case of the widely used anticancer drug cisplatin. Therefore, there is a relationship between the stability of the organotin(IV) complexes and their antitumour activity. In conjunction with our previous studies on organotin(IV) complexes [²¹-²⁶] the present paper aims to study the dimethyltin(IV) complexes with N,N,N’,N’-tetraethylethylene diamine to have information regarding the plausible interaction mechanism toward DNA fragment. The four ethyl groups attached to ethylenediamine nitrogen atoms may undergo hydrophobic interaction with the methyl groups of tin(IV) moiety. Such interaction may increase the stability of the formed complex. Also, the four ethyl groups of Et₄en will increase the hydrophobic nature of the complex, which will facilitate the dimethyltin(IV) complex to get into the cell. Density functional theory (DFT) calculations for geometry optimization and vibrational frequencies of all complexes and ligands were reported.

Experimental

Computational Details

For geometries optimization, the density functional theory was applied using the Gaussian09 program [³²]. A full optimization of all complexes and ligands geometries were obtained using the hybrid functional B3LYP (Becke's, three-parameter exchange functional, in combination with Lee-Yang-Parr correlation function) [³³-³⁵]. The basis set used is LANL2DZ (Los Alamos National Laboratory 2 double-zeta) [³⁶-³⁸] for Sn metal, while C, H, N, O and Cl were described by 6-311G* standard basis set in optimization. The GaussView 5.0 by Gaussian Inc. program was used for inspecting the input and output files generated by Gaussian09, for preprocessing, structure modification, and post processing analyses of structures, frequencies, and forces. To identify the most stable structure, the minima, a frequency analysis was performed for each stationary point. These analyses are performed to ensure that all minima have no imaginary frequencies in the vibrational mode calculations.

Results and discussion

Dimethyltin(IV) complex formation equilibria

The protonation constants of N,N,N’,N’-tetraethylethylenediamine were determined using the same experimental conditions as solvent composition, temperature and ionic strength which are used for the investigation of organotin complex equilibria. The overall protonation constants (logβ₁₁ and logβ₁₂) for Et₄en were estimated. The values obtained agree with the literature data [³⁹] considering changes in experimental condition.

The acid-base equilibria of dimethyltin(IV) ion was investigated previously in aqueous media [⁴⁰-⁴⁴]. The potentiometric data in the mixed solvents used were fitted assuming a model composed of the species 10-1, 10-2, 10-3, 10-4, 10-4, 20-3 and 20-4, Table 1. The aqua-hydroxo species (10-1) may undergo dimerization according to the general equilibrium

2[(DMT)(H2O)3(OH)]+⇌ [(DMT)2(HO)2(H2O)4]2++ 2H2O+ 3

The dimerization constant (K_d) can be determined by Eq. 4.

log Kd= log β202- 2logβ10-1  4

Following the same way the dimerization constant of the species 20-4 is determined by the Eq. 5 [²⁴].

logKd= log β20-4 - 2logβ10-2      5

The potentioetric titration curves of the protonated N,N,N’,N’-tetraethylethylenediamine and its dimethyltin(IV) complex are compared. The complex titration curve is lower than the Et₄en curve, Fig. 1. This corresponds to the formation of a complex species through the ionization of hydrogen ion. The titration data were fitted considering the formation of the 110, 111, 11-1 and 11-2 species, according to scheme 1.

Fig. 1 Concentration distribution of various species as a function of pH in the DMT- Et₄en system. 

Scheme 1 Formation equilibria of dimethyltin(IV)-Et₄en complex. 

Eq. 6 and 7 [²⁶] are used to calculate the pK_a of coordinated water molecules.

pKa1= log β110- logβ11-1 6

pka2= log β11-1- logβ11-2  7

The obtained values are 5.57 and 9.68 for pK_a1 and pk_a2 respectively. These are higher than those of water molecules coordinated to the free dimethyltin(IV) ion. This may be explained on the premise that coordination of Et₄en will decrease the electrophilicity of tin. This will result in the coordinated water molecule will be weakly bound and consequently will be less acidic.

The speciation diagram of DMT-Et₄en complexes is used to provide a useful picture of the coordination of organotin with Et₄en as a function of pH, Fig. 1. The protonated complex with stoichiometric coefficients 111, predominates at lower pH. It attains a maximum concentration of 38% at pH = 3.8. The deprotonated complex 110 starts to form at pH 3.0 and reaches the maximum concentration of 39% at pH = 5.7. The hydrolyzed form 11-1 predominates between pH = 5.7 and 8.0 with maximum formation degree of 27% at pH = 6.9. It is interesting to note that the complex species 110 and 11-1 are predominating in the physiological pH range. Therefore the binding of DMT complex with DNA constituent, the main target in the chemotherapy is possible. This may be explained on the premise that the DNA constituents coordinate in the negatively charged anionic form.

Effect of temperature

The thermodynamic parameters ΔH° and ΔS° associated with the protonation of N,N,N`,N`-tetraethylethylendiamine (Et₄en) and its complex formation with the dimethyltin(IV) species were calculated from the temperature dependence of the data in Table 3. The plots are given in figures 2-3. These data can be employed to extrapolate the equilibrium constants to other temperatures. The main conclusions from the data can be summarized as follows:

Fig. 2 Plot of log β₁ vs. 1/T for (CH₃)₂Sn-Et₄en complex. 

Fig. 3 Plot of log β₂ vs. 1/T for (CH₃)₂Sn-Et₄en complex. 

Effect of solvent

It is reported that the effective dielectric constant of the surrounding of active sites cavities of enzymes and proteins [⁴⁵,⁴⁶] is smaller , compared to in bulk water. Dielectric constants in such sites were found to be in range from 30 to 70 [⁴⁷, ⁴⁸]. Hence by using solution media containing around 10 to 50% dioxane, one may expect to approach to some extent the situation in these biological sites. By this way one may extrapolate the data to physiological condition. Careful examination of the medium effect on the equilibrium constants reveal the following features:

Fig. 4 Plot of log β₁ vs. dioxane percentage for (CH₃)₂Sn-Et₄en complex. 

Fig. 5 Plot of Plot of log β₂ vs. dioxane percentage for (CH₃)₂Sn-Et₄en complex. 

Displacement reaction of coordinated Et₄en by DNA constituents

It was reported that (CH₃)₂Sn(IV) has high affinity for N-donor ligands such as DNA constituents. Such interaction with DNA is thought to be responsible for the anti-tumour activity of related complexes [⁵⁰]. The mechanism of antitumor activity of (CH₃)₂Sn-(amine) complex is based on the displacement of the amine by DNA. Consequently, the equilibrium constant for such reaction is of biological importance. Consider inosine as a typical DNA constituent (presented by HA) and Et₄en (presented by B). The equilibria of complex-formation and displacement reactions may be presented introductory as:

The equilibrium constant for the displacement reaction is expressed by:

Substitution results in:

The formation constant value for Et₄en complex, [(CH₃)₂SnB]²⁺ , amounts to log β₁₁₀ = 11.31 at 25^oC (Table 1). The log β₁₁₀ values [⁵¹] for (CH₃)₂Sn-DNA constituents complexes [(CH₃)₂SnA]⁺ together with the equilibrium constants values (log K_eq ) of displacement reaction calculated by Eq. 12, are given in Table 4. The K_eq values clearly indicate the ability of DNA to displace the coordinated Et₄en from its dimethyltin(IV) complex. Also, K_eq value reveals to what extent the organotin-amine complex could interact with the DNA constituents, the main target in chemotherapy of tumours. The values of equilibrium constant of the displacement reaction for the different DNA constituents are compared. The nucleotide IMP and GMP have the highest values. The extra stability is due to the different columbic force of attraction between the dipositively charged dimethyltin(IV) ion and the nucleotides IMP or GMP having extra negative charges on the phosphate group. Also, the phosphate group of IMP and GMP is acting as anchoring site for tin [⁵²]. This will facilitate the release of coordinated Et₄en.

Geometry optimiation

Calculations of the equilibrium geometries of the complexes and the ligands were carried out by the program Gaussian 09 at the B3LYP level of theory using the bases set LANL2DZ for Sn metal and standard basis set 6-311G* for C, H, N, O and Cl atoms. The optimized structures of the ligands and the complexes are given in Fig. 6. The geometric computational parameters for the optimized molecular geometry as bond angles, atomic charges and bond lengths are determined and given in (Tables 5 and 6). The results are interpreted as given in the following:

Fig. 6 The optimized structure of ligands and complexes according to B3LYP level of theory using the bases set LANL2DZ for Sn metal and standard basis set 6-311G* for C, H, N, O and Cl atoms. 

Based on the DFT calculation, the polarity of the complexes, is much larger than that for the free ligands as indicated by the dipole moment of the complexes and the ligands (Table 6). On the other hand, the dipole moment of the hydroxo-complexes (CH₃)₂Sn(Et₄en)(OH)₂ and (CH₃)₂Sn(en)(OH)₂ are lower than that of the corresponding dichloro-complexes (CH₃)₂Sn(Et₄en)Cl₂ (Table 7). This may be described on the assumption that the O-Sn(IV) bond is shorter than the Cl-Sn(IV) bond as given in Table 5.

The binding energy (BE) of each complex was calculated according to the following equation:

In equation 13, E_AB, E_A and E_B is the energy of the complex, dimethyltin(IV) species and ligand respectively. The calculated BE (in kJ/mol) of the complexes (CH₃)₂Sn(Et₄en)Cl₂, (CH₃)₂Sn(en)Cl₂, (CH₃)₂Sn(Et₄en)(OH)₂, (CH₃)₂Sn(en)(OH)₂, (CH₃)₂Sn(Et₄en) (H2O)22+ and (CH₃)₂Sn(en) (H2O)22+ are -52.4, -99.2, -4.2, -31.0, -400.1 and -382.8, respectively. The negative values of the BE indicate that the complexes are stable.

Calculated infrared spectra of ligands and complexes

The experimental counterparts of the calculated IR spectra results are currently not available for comparison and it will be considered in our future work. Therefore, we will discuss the calculated IR spectra of the complexes from the theoretical point of view only. The calculated IR spectra of the free ligands and the complexes are shown in Fig. 7. There are no imaginary vibrational frequencies observed in all calculated spectra of the ligands and complexes. The assignments of the calculated vibrational frequencies of the free ligands and their complexes are listed in Table 8. The theoretical assignments are easily obtained by the visualization of the normal mode displacement vectors utilizing the GaussView program. The assignments listed in Table 8 are only for the vibrational modes of the amino group vibrations because they are the most sensitive to changes upon the metal-nitrogen interactions. Also, the Sn-N, Sn-OH, Sn-Cl and Sn-OH₂ stretching vibrations are identified. In the spectra of the CH₃)₂Sn(en)Cl₂, (CH₃)₂Sn(en) (H2O)22+ and (CH₃)₂Sn(en)(OH)₂ complexes the β_s(NH₂) vibration is shifted to higher wave frequency in comparison with that found in the free en ligand (Table 8). Also for the same complexes the v (C−N) vibration mode is shifted to lower wave frequency in comparison with that of free en ligand. However, in the case of (CH₃)₂Sn(Et₄en)Cl₂, (CH₃)₂Sn(Et₄en)(H₂O)₂ ²⁺ and (CH₃)₂Sn(Et₄en)(OH)₂ complexes this vibrational mode (v(C−N)) is shifted to higher frequency in comparison with that of free Et₄en ligand. In the spectra of all complexes the v _as(C‒C‒N vibration mode is shifted to lower frequency in comparison with those of free en and Et₄en ligands. The above mentioned shifts of these vibrational modes can be attributed to the coordination of the nitrogen atom with the metal.

Fig. 7 Calculated Infrared spectra of the (a) Et₄en ligand, (b) En ligand, (c) (CH₃)₂Sn(Et₄en)Cl₂ complex, (d) (CH₃)₂Sn(en)Cl₂ complex, (e) (CH₃)₂Sn(Et₄en) (H2O)22+ complex, (f) (CH₃)₂Sn(en) (H2O)22+ complex, (g) (CH₃)₂Sn(Et₄en)(OH)₂ complex and (h) (CH₃)₂Sn(en)(OH)₂ complex according to B3LYP level of theory using the bases set LANL2DZ for Sn metal and standard basis set 6-311G* for C, H, N, O and Cl atoms. 

Conclusion

The present investigation reports the complex formation equilibria of dimethyltin(IV) with N,N,N’,N’-tetraethylethylenediamine and its parent ethylenediamine. The results show the formation of 1:1 complex and the mono- and dihydroxo- species. The effect of solvent polarity on the stability of the complexes was discussed. The thermodynamic parameters of the complexes as ΔH° and ΔS° were determined and discussed. The displacement reaction of Et₄en coordinated to dimethyltin (IV) by DNA constituents was investigated. IMP and GMP have the highest tendency to displace Et₄en. The geometric parameters, bond angle, atomic charge and bond lengths were computed and interpreted. The results of the present study are expected to contribute to the chemistry of diorgantin (IV) as potential antitumours.