Synthesis, Crystal Structures, and Anti-Liver Cancer Activity Studies on Three Similar Coordination Polymers

Abstract

By employment of a pre-designed molecular building block [(pzTp)Fe(CN)₃], one cyano-bridged trinuclear complex [MnMe₄TACD][(pzTp)Fe(CN)₃]₂(H₂O)₁₆ 1, [pzTp = tetrakis(pyrazoly)borate, Me₄TACD = 1,4,7,10-tetramethyl-1,4,7,10-tetraazacyclododecane] and two 1D chain-like coordination polymers {[(pzTp)Fe(CN)₃]₂[M(Bim)₂](H₂O)₂}_n [M = Cu(II) for 2 and Mn(II) for 3, Bim = bis(1-imidazolyl)methane] were synthesized by making use of different nitrogen-containing organic linkers. Furthermore, in vitro cytotoxicity of compounds 1–3 was studied against four human liver cancer cells (HepG2, Bel-7402, SMMC7721, and MHCC97) via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) assay experiments.

1. Introduction

Nowadays, it has been widely accepted that cancer is one of the major threat to the health of people worldwide which could cause millions of deaths every year [1]. In spite of great efforts have been devoted to the creation of therapeutic tools, chemotherapy is still the dominant treatment method. In this case, the search of effective chemotherapy drugs with low cost, high selectivity, lack of undesirable side effects, and low cytotoxicity is of high importance [2,3]. The cancer cell resistance to anticancer agents and the toxicity of modern chemotherapy make the sciences worldwide search for new drugs and cure methods for such an insidious disease [4].

Metal-organic coordination polymers have been rapidly developed in the past decades due to their fascinating structures or topologies, especially their potential use as functional materials in various key domains such as asymmetrical catalysis, energy gas storage and separation, molecular and ionic magnetism, guest-dependent luminescence, and so on [5,6,7,8,9,10]. Many efforts have been devoted to selecting suitable ligands because they play important roles in the guiding the formation of the obtained produces with targeted chemical and physical characters [11]. The organic ligands containing N atoms have received considerable attention for the versatile binding modes as a bridging linker between inorganic moieties [12,13]. In recent years, [(pzTp)Fe(CN)₃] has been used as a pre-designed molecular building block to construct heterometallic coordination polymers, which show interesting structural diversity and outstand properties [14]. For instance, Oshio and co-workers reported a series of heterometallic coordination polymers constructed from the [(pzTp)Fe(CN)₃] building blocks, which demonstrate both electric and magnetic bistability [15]. In this work, we chose [(pzTp)Fe(CN)₃] to react with different metal ions and different N-donor organic ligands to afford one cyano-bridged trinuclear complex [MnMe₄TACD][(pzTp)Fe(CN)₃]₂(H₂O)₁₆ 1, (pzTp = tetrakis(pyrazoly)borate, Me₄TACD = 1,4,7,10-tetramethyl-1,4,7,10-tetraazacyclododecane) and two 1D chain-like coordination polymers {[(pzTp)Fe(CN)₃]₂[M(Bim)₂](H₂O)₂}_n (M = Cu(II) for 2 and Mn(II) for 3, Bim = Bis(1-imidazolyl)methane, Figure 1) with interesting structural diversity. Additionally, in vivo anticancer activity of compounds 1–3 was then evaluated.

2. Materials and Methods

2.1. Instruments and Chemicals

A Rigaku D/MAX 2500V diffractometer (Rigaku Corporation, Tokyo, Japan) was used to collect the powder X-ray diffraction (PXRD) curves. An Oxford Xcalibur E diffractometer with Mo-Kα radiation (λ = 0.71073) was employed to collect the single crystal X-ray diffraction data. A Perkin Elmer model 240C (PerkinElmer, Norwalk, CT, America) instrument was employed to obtain the element analyses (C, H, and N) results. All the chemicals in this study were commercially available and used as received. Bu₄N[pzTpFe(CN)₃], Me₄TACD, and Bim ligands were bought from Ji’nan Heng Hua Technology Co., Ltd. (Ji Nan, China) at analytical grade. Mn(NO₃)₂ (aq, 50% w/w) and Cu(NO₃)₂·6H₂O were purchased from Alfa Aesar Chemical Reagent company (Tianjin, China) at analytical grade.

2.2. Preparation and Characterization for Compounds 1–3

A 10.0 mL ethanol solution containing 0.10 mmol (65.5 mg) of Bu₄N[pzTpFe(CN)₃] and 0.20 mmol (45.0 mg) of Me₄TACD was added dropwise to a 10.0 mL aqueous solution containing 0.10 mmol of Mn(NO₃)₂ (aq, 50% w/w). The as-made suspension was mixed tempestuously via stirring for 3 h and then filtrated to remove the solid. After filtration, slow evaporation of the filtrate in air gave black crystals after several days. Analytical calcd for compound 1 (C₄₂H₉₀B₂MnFe₂N₂₆O₁₉): H, 6.25; C, 34.75; N, 25.09%; found: H, 6.11; C, 35.02; N, 25.26%.

A 10.0 mL methanol solution containing 0.10 mmol (65.5 mg) of Bu₄N[pzTpFe(CN)₃] and 0.20 mmol (29.0 mg) of Bim ligand was added dropwise to a 10.0 mL aqueous solution containing 0.10 mmol (24.0 mg) of Cu(NO₃)₂·6H₂O. The as-made suspension was mixed tempestuously via stirring for 3 h and then filtrated to remove the solid. After filtration, slow evaporation of the filtrate in air gave black crystals after several days. Analytical calcd for 2 (C₇₂H₆₈B₂CuFe₂N₂₈O₁₀): H, 3.63; C, 43.25; N, 34.39; found: H 3.29, C 43.11, N 34.08.

The synthesis process of complex 3 was similar to that of 2, except that the equal amount of Mn(NO₃)₂ (aq, 50% w/w) was used to replace Cu(NO₃)₂·6H₂O. Analytical calcd for 3 (C₇₂H₆₈B₂MnFe₂N₂₈O₁₀): H, 3.66; C, 43.56; N, 34.63; found: H 3.49, C 43.42, N 34.55.

2.3. Crystal Structure Determination

An Oxford Xcalibur E diffractometer with graphite monochromatized Mo K radiation (0.71073 Å) was used to collect the reflection data of 1–3 at 298 K. The direct method in the XS program was used to get the initial structural modes and the least-squares method in XL program was used to refine the structural models. All the H atoms on the organic ligands were generated geometrically, and all non-hydrogen atoms in the two structures were treated anistropically. For the highly-disordered nature of the seven water molecules in 1, they could not be figured out appropriately from the Fourier map, so the SQUEEZE manipulation in the software PLATON(version: 1.18) was used to remove their electronic contribution. The SQUEEZE calculation result reveals that each unit cell contains 1256 electrons, corresponding to 78.5 electrons (7.85 H₂O) per molecular unit, which is near to the value derived from the element analyses (eight H₂O per molecular unit).

Table 1. The crystalline data and refinement parameter for complexes 1–3.

	1	2	3
Formula	C48H72B2Fe2MnN22	C44H44B2CuFe2N30O2	C44H44B2Fe2MnN30O2
Mr	1145.53	1221.95	1213.32
T/K	298 (3)	298 (3)	298 (3)
Crystal system	Orthorhombic	Monoclinic	Monoclinic
Space group	Fddd	P21/n	P21/n
a/Å	16.8839 (6)	8.4126 (3)	8.6043 (11)
b/Å	25.7793 (6)	16.1784 (5)	16.1504 (19)
c/Å	62.8204 (17)	19.4159 (6)	19.470 (2)
α/°	90	90	90
β/°	90	92.007 (2)	92.126 (9)
γ/°	90	90	90
V/Å3	27,342.9 (14)	2640.93 (15)	2703.7 (6)
Z	16	2	2
Dcalc/g·cm−3	1.113	1.537	1.4902
μ(Mo Kα)/mm−1	0.646	1.012	0.828
Rint	0.0632	0.0920	0.0493
R1	0.0433	0.0570	0.0417
ωR2	0.1303	0.1649	0.1201
CCDC number	1,832,283	1,832,284	1,832,285

shows the crystal data and the structure refinements. Pertinent crystal data and structural refinement results for compounds 1–3 were summarized in Table 1.

2.4. Antitumor Activity

Four human liver cancer cell (HepG2, Bel-7402, SMMC7721, and MHCC97) lines were developed in a RPMI 1460 medium supplemented with 10% fetal calf serum, 0.1 g/L streptomycin and 0.1 g/L penicillin. The above systems were incubated in a humidified incubator with air and CO₂ (95:5, v:v) at 37 °C. RPMI 1640 was used to dilute the cell to a concentration of 5 × 10⁴ cells/mL. The 96-well cell culture at a volume of 100 μL per cell was used to seed the cells. Then the system was incubated for 24 h at 37 °C in 5% CO₂. The MTT solution (0.5 mg/mL, 150 μL) was added to each well tested and the plates were cultured for 4 h. Afterwards, 100 μL of dimethyl sulfoxide (DMSO) was transferred into each well to dissolve the samples. A microplate reader (Per-kinElmer, Waltham, MA, USA) was used to record the absorbance at a wavelength of 450 nm, which is further used in the calculation of the cell relative viability. Each parallel experiment was carried out for at least three times to obtain the mean values.

3. Results and Discussion

3.1. Molecular Structure

An evaporation method was used in the synthesis of the single crystals of 1 in a beaker. The X-ray single-crystal investigation demonstrates that 1 locates in the orthorhombic crystal system, the Fddd space group with a large cell volume of 27,342 ų. The molecular unit of 1 is composed of neutral [MnMe₄TACD][(pzTp)Fe(CN)₃]₂ cluster as well as the lattice water molecules. In the structure of the trcore unit, the [Fe^III(CN)₃(pzTp)]⁻ building block behaves as a monodentate linker using its one cyano group to connect with the [MnMe₄TACD] unit, while each [MnMe₄TACD]²⁺ building block is coordinated to two [Fe^III(pzTp)(CN)₃]⁻ building blocks, leading to the formation of a centrosymmetric neutral trcore [MnMe₄TACD][(pzTp)Fe(CN)₃]₂ molecular structure. Within the structure of the trcore unit, there is one crystallographically independent Fe(III) ion (labeled as Fe1) and one Mn(II) ion (labeled as Mn1). In the [Fe^III(pzTp)(CN)₃]⁻ anion, the six-coordinated Fe(III) ion is bonded to three N donors from there pz linkers with the leaving coordinating sites completed via the C donors on three CN ions (Figure 2a). The Fe-N bond distances locate in the region of 1.990(2) to 2.013(2) Å while the Fe-C bond distances range from 1.918(3) to 1.932(3) Å, respectively, which are comparable to those reported in the literature which are also formed as Fe^III tricyanide metal-organic complexes. As for the Mn²⁺ ion in the [MnMe₄TACD] unit, it six-connects with the N donors on the four Me₄TACD ligands and two N donors on two independence [(pzTp)Fe(CN)₃] units with a N-Mn lengths ranging from 2.195(3) to 2.391(3) Å. Two Fe(III) centers and one Mn(II) center were arranged in a winding array with a Fe–Mn–Fe angle of 87.5° (Figure 2b). Furthermore, the adjacent trinuclear cluster are interacted with each other though the π–π interaction of the pz rings to form a 1D chain-like supermolecular structure (Figure 2c).

Complexes 2 and 3 could be obtained in a moderate yield using a slow volatilization method, and their structures were fully studied by the X-ray single crystalline diffraction technology, which shows that both complexes are isomeric and possess similar 1D chain-like framework. Thus, herein we only describe the structure of complex 2 in detail. X-ray single-crystal diffraction analyses revealed that complex 2 locates in the monoclinic crystal system, P2₁/n space group and demonstrates a 1D chain-like structure. The crystal structure comprises of two components: one is the [Fe^III(pzTp)(CN)₃]⁻ anion and the other is the 1D Cu(II) chains that formed by the connection of the Cu(II) ions by the Bim ligands along the a axis. As shown in Figure 3a, in the [Fe^III(pzTp)(CN)₃]⁻ anion, the six-coordinated Fe(III) atom is coordinated to three N donors from three independence pz linkers with the leaving binding sites completed via the C donors on the three CN ions. The Fe-N bond lengths are in the region of 1.953(3) to 1.995(3) Å and the Fe-C bond distance varies from 1.925(4) to 1.932(4) Å, respectively, which are comparable to those reported in the literature, which are also formed as Fe^III tricyanide metal-organic complexes. As for the Cu²⁺ ion in the 1D chain (Figure 3b), it binds with six nitrogen donors on four Bim linkers and two nitrogen donors from the [Fe^III(pzTp)(CN)₃]⁻ anion with a N-Cu distances varying from 2.243(3) to 2.270(3) Å, forming a distorted octahedral geometry. Each Cu²⁺ ion is connected with two [Fe^III(pzTp)(CN)₃]⁻ anions though the CN group on the [Fe^III(pzTp)(CN)₃]⁻ anion, which is further linked with the adjacent Cu²⁺ ions though four Bim ligands in a double-walled model. Such connection modes results in the formation of the 1D chain-like framework of 2. The 1D chains are further stacked in an ABAB fashion in the three-dimensional direction to form a 3D supermolecular network (Figure 3c). We carried out the power X-ray diffraction measurement for the three compounds, and the results are shown in Figure 4. The well match between their simulated peaks with the peaks of their prepared samples confirmed the phase purity of their prepared samples.

3.2. In Vitro Cytotoxicity Study against the Cancer Cells

In order to preliminarily evaluate the anticancer efficacies of the three complexes (1–3), their tumor cell growth inhibition activity was assessed in vitro against four human liver cancer cells (HepG2, Bel-7402, SMMC7721, and MHCC97) by MTT assay with the cisplatin drug as the refer. Expose time was set to 48 h and the cell growth inhibition activity effects of cisplatin were used to make a comparison with the results of 1–3. DMSO was used to dissolve the compounds at the concentration of 1 μM, and the diluted solution was added into the cell culture.

The data, which represent means ± SEM of three independent tests carried out in duplicate, are plotted in Figure 5a. A descending order could be observed for the anti-proliferative activity of the test complexes toward each type of the tumor cell lines used, which might be ascribed to the measured concentration required to inhibit tumor cell proliferation by 50% (IC₅₀ μg/mL). From the results above, we can conclude that the new compounds 1–3 demonstrate comparable growth inhibition activity toward all four types of cancer cells (IC₅₀ = 34–56 μg/mL) with the reference drug.

To probe the cytotoxicity of the as-prepared three compounds toward the human normal cells in vitro, oral epidermal cells (human normal cells) were selected to incubate with 1–3 for 24 h under different concentrations (0–30 μg/mL). Figure 5b shows the cell viabilities of the three complexes, which are all above 84% even at a concentration of 30 μg/mL. The above experimental results indicate that the three complexes showed low cytotoxicity toward the human normal cells.

4. Conclusions

The X-ray single crystal study demonstrates reveals that the metal-organic complex 1 is a trinuclear cluster that consists of two (pzTp)Fe(CN)₃ units and one MnMe₄TACD unit that connected by the CN ions, while compounds 2 and 3 are isostructural and demonstrate 1D chain-like structure that formed by the connection of the (pzTp)Fe(CN)₃ units and the 1D M(II)-chains though the CN ions. Furthermore, the in vitro cytotoxicity of compounds 1–3 was studied against four human liver cancer cells (HepG2, Bel-7402, SMMC7721, and MHCC97) via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) assay experiments, which reveals their potential use as anticancer drugs.