THE COMPLEXING ABILITY OF N-SUBSTITUTED THIOUREA DERIVATIVES AS CHELATING LIGANDS IN THE REACTION WITH PdCl2

The complexation reactions of N-substituted thiourea derivatives with PdCl2 have been investigated in the present work. The functionally substituted thiourea derivatives are found to be effective chelating agents, in which the nature of substituents has a significant impact to the compositions and structures of complexes. Thus, (N-pyridine-2-yl)thioureas in the interaction with PdCl2 form two types of complexes in the molar ratio of M:L 1:1 and 1:2, in which they act as S,N-bidentate ligands coordinated to the palladium ion by thione sulphur and the nitrogen atom of the pyridine ring. The reaction of N-allylthioureas with PdCl2 in the equimolar ratio results in formation of the π-complexes where the ligands are coordinated by the thione sulphur and the C=C double bond of the allylic moiety. The preparative methods for the synthesis of this type of complexes have been developed. The composition of the products synthesized and the ligands coordination mode have been determined by elemental analysis and 1H NMR spectroscopy. Furthermore, the structure of compounds 4, 5 has been confirmed by the X-ray diffraction study. The biomedical studies have proven that the complex compounds 5 and 6 in vitro have the cytostatic and cytotoxic activity against tumour HeLa cells.

Thiourea and its N-substituted derivatives are efficient complexing agents for metal ions, which are widely used as ligands in coordination chemistry [1][2][3]. The presence of unshared electron pairs in the sulphur atom and two nitrogen atoms allows to form the complex compounds with transition metal salts of the different type, many of which possess valuable applied properties (optical, semiconductor, biological, etc.) and are employed in various fields of science and technology [4][5][6][7]. Thioureas are of prime importance as anticancer drugs due to their ability to inhibit the enzymes that are involved in the malignant tumour formation processes (tyrosine kinase, protein tyrosine kinase and NADH oxidase) [8][9][10].
In recent 15-20 years much attention has been given to the study of the biological activity of complex compounds of transition and platinum metals [11][12][13][14][15]. Many of them exhibit a high pharmacological activity, and as therapeutic agents are often superior to starting metal salts because the complex formation prevents hydrolysis in the physiologic medium, reduces toxicity and facilitates penetration of medicines through the cell membrane. Moreover, the use of biologically active ligands, an additive or synergistic effect manifests itself between the constituents of complexes [16,17]. Therefore, development of methods for the synthesis of different types of complexes based on thiourea derivatives is a promi-sing trend in creating novel therapeutic and diagnostic agents [3,[18][19][20][21].
The introduction of additional functional groups into the structure of thioureas (such as allyl, pyridine, morpholine) allows to expand the range of their biological activity and to increase the denticity of these ligand systems, it leads to their competitive coordination to metal ions. On the other hand, palladium as a "soft Lewis acid" has also the competing ability in the interaction with ambidentate ligands by the «soft-soft» type thereby resulting in formation of complexes with different structures depending on the synthetic conditions and the geometrical arrangement of donor atoms in the ligands.

Results and Discussion
The complexation reactions were carried out in the acidic medium according to the Scheme. Thioureas (HL 1 , HL 2 ) contain nucleophilic pyridine nitro- ISSN 2308-8303 gen and sulphur atoms of the carbothioamide group at the stoichiometrically advantageous positions for formation of metallochelates; it results in their coordination to the central metal ion as S,N-bidentate ligands thereby forming complexes 1-4. At the same time the acid medium contributes to their stay in thionic tautomeric form. Changing the reaction conditions such as temperature, the heating time and the molar ratio affect the number of coordinated ligand molecules and chlorine anions. Unlike HL 1 , HL 2 , thioureas HL 3 , HL 4 contain only one nucleophilic atom (sulphur) capable to coordination, which makes them potential monodentate reagents. However, the ability of palladium to the "softsoft" interaction leads to formation of n,π-chelate complexes 5, 6. All attempts to synthesize complexes with the molar ratio of 1:2 under the same conditions were unsuccessful. It may be due to the antisymbiosis effect of "soft" allyl and thiourea groups, which makes unfavourable further substitution of "hard" Clions to "soft" donor atoms of another ligand molecule [22].
All the complexes are soluble in DMSO and DMF, whereas complexes 2, 4-6 are also soluble in water and alcohols.
The asymmetric unit of compound 5 involves one solvation water molecule (Fig. b). The central palladium ion forms a square planar coordination unit Pd(C=C) SCl 2 by atom S1, two chloride ions Cl1, Cl2 and double bond C1=C2 of the allyl fragment. The midpoint of the C1=C2 double bond is considered as the point ligand with the distance of 2.0451(3) Ǻ to the central metal ion. The average deviation from the absolute planar configuration is 0.0325 Ǻ. The angle between the polyhedral plane and С1-С2 bond is 179º. The bond length С4−S1 (1.741(4) Ǻ) indicates to the thione tautomeric form of the coordinated thiourea HL 3   of the signal for N 2 H is due to its participation in forming the six-membered chelate ring Pd-N 1 -C 1 -N 2 H-C 6 -S (Scheme). The fairly substantial downfield shifts of the signals for the pyridine protons can also be attributed to the coordination of HL 1 by the pyridine N atom. It should be noted that the protons C 5 H Py (Δδ=0.766), C 2 H Py (Δδ=0.296), C 3 H Py (Δδ=0.266) and C 4 H Py (Δδ=0.192 ppm), which are in ortho and para position to the coordination bond (N Py →Pd) are most sensitive to the complexation. This is in full agreement with the current approach to location of the coordination bond using the heteronuclear NMR method [26].
The 1 H NMR spectrum of the HL 2 shows the signals at 7.05, 7.60, 7.70 and 8.28 ppm, which are assigned to the pyridine ring protons. A broad singlet corresponding to one proton at 9.85 ppm is assigned to the NH group. The morpholine protons give two triplets at 3.64 and 3.89 ppm, which correspond to -N-CH 2 and -O-CH 2 groups, respectively.
In the spectra of complexes 3, 4 all multiplets of the pyridine ring are shifted downfield and observed at 7.42-8.97 ppm, wherein the doublet H 5 located near the nitrogen atom undergoes significantly more downfield shift (∆δ=0.69 ppm); it indicates the participation of the pyridine ring in the coordination to the metal ion. The N 2 H group is included in the six membered metallocycle, and it causes a significant downfield shift of a broad singlet of this proton by 1.56/1.53 ppm and indicates inclusion of the carbothioamide group in the coordination to the metal. The morpholine proton signals are observed at 3.77/3.75 (-N-CH 2 ) and 4.03/4.00 ppm (-O-CH 2 ).
The comparison of the 1H NMR spectra of thioureas HL 3 , HL 4 and complexes 5, 6 shows that the proton signals of NH (the thioureate group) and =C2Hallyl by ∆δ=+1.289/+1.648, +0.301/+0.332 ppm undergo the most significant downfield shifts while the multiplet of the allyl fragment =С1Н2 is significant upfield by -0.544/-0.824 ppm. The most characteristic for the 1H NMR spectra of these complexes is splitting of the C3H2 signal to the two doublets.
The highest index ІС 50 (1.5•10 -4 M) was determined for the palladium complex with HL 3 (5). Both compounds studied had the proapoptotic effect, but in contrast to the highest ІС 50 index, cytostatic and proapoptotic effects for complex compound 5 were more pronounced. For complexes 5, 6 the apoptotic indexes exceeded the control rate sevenfold and 2.5 times, respectively.
Both compounds possessed the cytostatic and antisynthetic effect. The highest inhibition of cells in both synthetic and mitotic phases (G 2 /M) was found for compound 5: the growth of the cell subpopulation in the G 1 /G 0 -phase (1.5 times) was accompanied with reduction of the cell subpopulation in synthetic (1.3 times) and G 2 /М phases (in 3.3 times). The effect of compounds 6 on the cell cycle phases in the concentration of C complex =IC 50 /10 does not differ significantly from the control.