Synthesis, Crystal Structure, and CT-DNA Binding Activity of a 3-Methylpyridyl Pendant-Arms Cu(II)-Zn(II) Hetero-Binuclear Complex

, , , , , ,

(1.School of Chemical and Environmental Engineering, Wuhan Polytechnic University, Wuhan 430023, China; 2.Sichuan Institute of Product Quality Supervision and Inspection, Chengdu 610100, China)

Abstract:A new 3-methylpyridyl pendant-arms Cu(II)-Zn(II) hetero-binuclear complex was synthesized by condensation reaction of 3,3′-((ethane-1,2-diylbis((pyridin-2-ylmethyl)azanediyl))bis(methylene))bis(2-hydroxy-5-methylbenzaldehyde) (H2L) and propane diamine in the presence of metal ions. The structure was characterized by IR spectroscopy, UV-Vis spectroscopy, ES-MS spectroscopy and X-ray single-crystal diffraction. The crystal structure reveals that the complex crystallizes in hexagonal, space group P63/m, with a=1.982 18(17) nm, b=1.982 18(17) nm and c=1.839 4(2) nm. The coordination environment of Zn(II) and Cu(II) in complex can be described as approximately square pyramid, where the apical positions are occupied by acetate radical. Two metal centers are equivalently bridged by the phenolic oxygens and an acetate radical with the intermetallic separation of 0.289 7 nm. The interaction between the complex and calf thymus DNA (CT-DNA) was measured by cyclic voltammetry and viscosity studies, which shows a weak binding activity of 6.92×103mol/L.

Key words:3-methylpyridyl pendant-arm; Cu(II)-Zn(II) hetero-binuclear complex; coordination environment; CT-DNA binding

0 Introduction

Over the years, Schiff base complexes have received wide attention for their easy preparation and structural variety, and many of them have been synthesized[1-6]. Schiff base complexes are considered to be one type of the most important stereochemical models in transition metal coordination chemistry. Diamines with different branch or pendant-arms are usually selected to synthesize polynitrogen Schiff base macrocyclic complexes with different metal cavity sizes, hoping to achieve similar structure of metalloenzymes and metalloproteins. Pan et al[7]. have obtained a series of 28-memebered Schiff base macrocyclic complexes with two 2-thiophenoethyl pendant-arms using diethylene triamine, which shows good DNA cleavage and antibacterial activities againstE.coli. Kou et al[8-9]. have acquired two 28-memebered schiff base benzyl pendant-arms macrocyclic complexes with 1,3-propanediamine, which reveals an efficient catalytic activity of phosphoester bond cleavage.

A lot of macrocyclic binuclear transitional metal complexes owning DNA binding activities, and it's important in further study of DNA structural probes, phosphatase mimics, sequence-specific cleaving agents and potential anticancer drugs. So, the investigation of interaction between calf thymus DNA (CT-DNA) and transition metal complex is an important and active research area[10-11]. Cu(II) and Zn(II) play a critical role in diverse biological processes in organisms. In previous work, we reported an unsymmetrical 2-methylpyridyl macrocyclic with heterobinuclear Zn(II)Ni(II) complex[12], which shows good binding activity to CT-DNA and cleavage to pBR322 DNA. In order to further clarify the influence of pendant-arms in the complex during DNA binding process, a new Cu(II)Zn(II) heterobinuclear macrocyclic complex bearing two 3-methylpyridyl pendant-arms was prepared, shown in Fig. 1. The interaction between complex and CT-DNA was investigated by cyclic voltammetry and viscosity studies.

Fig.1 Synthetic route of complex

1 Expertimental section

1.1 Materials and instruments

The chemicals such as 5-methylbenzaldehyde, 3-pyridinecarboxaldehyde, propane diamine, ethanediamine and triethylamine are of analytical grade, and used as purchased. The high-purity solvents such as ethanol, acetonitrile and tetrahydrofuran (THF) are obtained from Sigma. Tetra(nbutyl) ammonium perchlorate (TBAP) was redistilled three times and then dried in a vacuum before use. 3-(Bromomethyl)-2-hydroxy-5-methylbenzaldehyde was prepared according to the literature methods[13].

The crystal structure of H2L and complex were carried out on a Bruker AXS SMART diffractometer using graphite monochromatized Mo-Kradiation (Mo Kαradiation monochromator). Data reduction and cell refinement were reported by SMART and SAINT programs. The structure was solved by direct methods (Bruker SHELXTL) and refined on F2by the full-matrix least-squares method[14]. Hydrogen atoms were located geometrically and refined in riding mode. The non-H atoms were refined with anisotropic displacement parameters. Calculations were performed using the SHELX-97 crystallographic software package.

IR spectra was recorded using KBr disc by a vector 22 FT-IR spectrophotometer. UV-Vis spectra was recorded on an UV-2450 spectrophotometer. The contents of carbon, hydrogen, and nitrogen element were determined on a PerkinElmer 240 element automatic analyzer. ESMS spectra was recorded on a Finnigan LCQ ESMS mass spectrograph using methanol as the mobile phase with an approximate concentration of 1.0 mmol·L-1. Cyclic voltammograms were run on a CHI model 750B electrochemical analyze with a three-electrode cell, which was equipped with a glassy carbon-working electrode, a platinum wire as the counter electrode and an Ag/AgCl electrode as the reference electrode. Viscosity experiments were performed on an Ubbelohde viscometer.

1.2 Synthesis of the complex

1.2.1 Synthesis of N1, N2-bis(pyridin-3-ylmethyl)ethane-1,2-diamine

To an ethanol (20 mL) solution of 3-pyridinecarboxaldehyde (10.8 g, 0.1 mol), a solution of ethanediamine (3.0 g, 0.05 mol) in ethanol (20 mL) was added dropwise. magnetic stirred for 10 h at room temperature, and white solid was formed. Solid mass was separated from the reaction solution and dissolved in 50 mL methanol, followed by reduction with NaBH4(7.5 g, 0.2 mol)[15]. Light yellow, oily liquid was obtained. Yield: 10.94 g (88.4%).1H NMR (300 MHz, CDCl3) δ 8.62-8.41 (4H, m, Py-H), 7.66 (2H, d, Py-H), 7.38-7.17 (2H, m, Py-H), 3.79 (4H, s, CH2), 2.76 (4H, s, CH2), 1.76 (2H, s, N-H).

1.2.2 Synthesis of 3,3′-((ethane-1,2-diylbis((pyridin-2-ylmethyl)azanediyl)) bis(methylene)) bis (2-hydroxy-5-methylbenzaldehyde) (H2L)

3-(Bromomethyl)-2-hydroxy-5-methylbenzaldehyde (5.2 g, 0.022 mol) was dissolved in 30 mL THF, N1, N2-bis(pyridin-3-ylmethyl)ethane-1,2-diamine (2.5 g, 0.01 mol) and triethylamine (4.2 g, 0.04 mol) as dried by potassium hydroxide was added under magnetic stirring. The mixture was stirred at room temperature for 24 h, then removed the solvent by spinning evaporator. The remaining yellow oil was recrystallized in acetonitrile, yield white solid. Yield: 3.6 g (66.8%). Some of the white solid dissolved in acetonitrile, light yellow cubic crystals suitable for the X-ray measurement were obtained.1H NMR (300 MHz, CDCl3) δ 10.19 (s, 2H, CHO), 8.54 (d, J=4.6 Hz, 2H, Py-H), 7.61 (d, J=1.1 Hz, 2H, Py-H), 7.37 (s, 2H, Py-H), 7.33-7.07 (m, 6H, Py-H &Ph-H), 3.74 (s, 4H, Py-CH2), 3.66 (s, 4H, Ph-CH2), 2.75 (s, 4H,NCH2), 2.23 (s, 6H, CH3).13C NMR (75 MHz, CDCl3) δ 194.33 (s), 158.44 (s), 150.59 (s), 149.08 (s), 138.00 (s), 136.92 (s), 133.54 (s), 130.82 (s), 128.89 (s), 125.35 (s), 123.60 (s), 121.49 (s), 56.10 (s), 53.09 (s), 50.83 (s), 20.48 (s).

1.2.3 Synthesis of complex ([CuZnL(OAc)])

1.3 DNA binding experiments

The binding activity of the complex towards CT-DNA were analyzed by cyclic voltammetry and viscosity methods. The CT-DNA solution was prepared by dissolving CT-DNA in Tris-HCl buffer (100 mL, 50 mmol/L Tris-HCl, 50 mmol/L NaCl, pH=7.2). The concentration of CT-DNA solution was determined by employing an extinction coefficient of 6 600 mol/L·cm-1at 260 nm[16]. The complex was dissolved in DMF solutions containing tetra(nbutyl)ammonium perchlorate (TBAP) as the supporting. The solution was deaerated for 15 min before measurements.

The viscosity studies were carried out using a capillary viscometer at (25.0±0.1) ℃. Each set of data measured in triplicate, averages are presented as (η/η0)1/3versus molar ratio of complex to DNA, whereηis the viscosity of DNA in the presence of the complex andη0is the viscosity of DNA in the absence of complex[17].

2 Results and discussion

2.1 Structural characterization of the complex

Fig.2 IR spectrum of the complex

The UV-Vis spectra of complex shows two sharp absorptions in the range of 300～800 nm. The spectral features of the complex is similar to those macrocyclic metal complexes bearing 2-methylpyridyl pendant-arms[20]. The sharp absorptions around 258 nm region is assigned to π-π*transition of benzene ring, and the moderate absorptions at 367 nm are attributed to the charge transfer (CT) transition of ligand-to-metal. The IR and UV-Vis spectra data of the complexes are in good agreement with their structural features.

The ES-MS spectra of the complex in methanol solution was shown in Fig.3. The dominant molecular ion peaks appear at m/z 762.17 corresponding to [CuZnL(OAc)]+(calc. 762.71), indicating that the cation is stable in methanol solution. The results are proved by the good agreement with the theoretical and experiment isotope distributions.

Fig.3 ES-MS spectra of complex (inset the isotopic distribution of the peak at m/z 762.17). (a) Experimental pattern; (b) calculated pattern

Two single crystals of H2L and complex with dimensions of 0.16 mm×0.12 mm×0.10 mm and 0.22 mm×0.24 mm×0.28 mm were selected for X-ray structure analysis. The results indicate that H2L crystallizes in monoclinic, space groupC2/c, witha=2.072 9(2) nm,b=0.701 24(8) nm,c=1.945 2(2) nm, while the complex crystallizes in Hexagonal, space groupP63/m, witha=1.982 18(17) nm,b=1.982 18(17) nm,c=1.839 4(2) nm. Crystallographic data and details about the data collection of H2L and complex are presented in Table 1. Selected bond lengths and angles relevant to the Zn(II) and Cu(II) coordination spheres of the complex are listed in Table 2.

Table 1 Crystal data and details of the structure determination for H2L and complex

The perspective view of H2L and complex are given in Fig. 4. The formula of H2L is C32H34N4O4, the basal phenyl plane and pyridyl plane are approximately vertical. The molecular structure of the complex contains a heterodinuclear [CuZnL]2+cation and an OAc-anion, which is rotational symmetry along the Cu-Zn axis. The complex is a 19-membered [1+1] Schiff-base macrocycle, with two metal ions bridged by two 2-phenoxy oxygens and one acetate. The two 3-methylpyridyl pendant-arms of the complex are situated in the same sidepiece of the mean molecular plane, different from the reported 3-pyridyl pendant-arms binuclear Ni(II) complex, probably due to the distinctive synthetic route[21]. The Zn(II) center coordinated with O and N atoms belonging to ligand (O1, N1) and acetate(O3) yield an overall metal coordination geometry of five froming three six-membered chelate rings. (ring 1: Zn1-O1-C1-C2-C7-N1; ring 2: Zn1-N1-C17-C18-C17-N1; ring 3: Zn1-N1-C7-C2-C1-O1). The Cu(II) center bounds by O1, N2 and O2, forming two same six-membered chelate rings (Cu1-O1-C1-C6-C9-N2) and a five-membered chelate ring (Cu1-N2-C16-C16-N2). The average distances of Zn(II) or Cu(II) ions deviating from the coordination plane of the respective N2O2composition are 0.047 0 nm and 0.061 3 nm, respectively. The coordination geometry of Zn(II) and Cu(II) may be considered as approximately square pyramid. The Zn—Cu distance is 0.289 7 nm, which is smaller than the dinickel(II) macrocyclic complex with 2-thiophenoethyl pendant-arms[7].The distance of Cu(II) or Zn(II) ions and corresponding coordination atoms are within 0.202 0～0.210 4 and 0.198 4～0.202 5 nm, respectively. The angle between the two pyridine rings is 25.4°.

Fig.4 Perspective view of H2L (a) and complex (b) together with the atom numbering schemes

The cyclic voltammograms of the complex were recorded in DMF solution in the scan range of -1.2～-0.4 V. The scan rates were varied in the range of 50～200 mV·s-1, shown in Fig.5. A pair of redox peak was found at every experimental scan rate, which is considered as Cu(II)Zn(II)/Cu(I)Zn(II) redox couple. For the scan rate of 100 mV·s-1, the anodic peak potential (Epa) and the cathodic peak potential (Epc) are -0.617 and -0.812 V, respectively. Half-wave potentials,E1/2, taken as the average ofEpaandEpc, is -0.715 V, which is 0.046 V higher than that ofE1/2in 2-methylpyridyl pendant-arms Zn(II)Ni(II) complex[2]. The difference ofE1/2may be from the different solution composition in the experiments and the coordination environment around metal ions. The separation of the anodic and cathodic peak potentials(ΔEp) is 0.195 V, and the current intensity of cathodic and anodic peaks are nonequivalent(ipc/ipa=2.6), suggesting it’s a quasi-reversible electrochemical process[22]. The separation ofipc/υ1/2values in the scan rates of 50～200 mV·s-1is slightly (<0.001), indicative of diffusion controlled.

Fig.5 Cyclic voltammograms of the complex in DMF solution

2.2 DNA binding activity

At present, the electrochemical study of complex and DNA includes cyclic voltammetry, DC polometry, linear scanning voltammetry, etc. Among these methods, the cyclic voltammetry is used the most extensive and advantaged[23]. The complex solution interacts with DNA may change redox performance of the cyclic voltammetric curve evidently. Accompany with the presence of DNA, the peak current of the complex shifts positive, indicating that the complex binds with DNA in intercalation mode; the peak current of the complex shifts negative, means the interaction between complex and DNA is grooving or electrostatic interaction[12].

The cyclic voltammograms of the complex in Tris-HCl buffer(100 mL, 50 mmol/L Tris-HCl, 50 mmol/L NaCl, pH = 7.2) at a scan of 100 mV/s are shown in Fig.6. Upon the addition of 100 μL DNA, it exhibits a pair of anodic and cathodic peaks at -0.789 and -0.550 V, which can be attributed to the Cu(II)/Cu(I) redox couple[2]. The single reduction peak at 1.148 V is caused by strong electron-donor capacity of the pyridyl arms in complex, which bind to CT-DNA and lead to electronic transfer. Meanwhile,Epawas shifted by 0.019 V toward anode andipawas dropped by 4.53%, the minor changes compare to the other complexes indicate the present complex bind to CT-DNA through weak intercalation[24]. The following equation[25]was applied to further testify the binding affinity of complex to CT-DNA.

Fig.6 Cyclic voltammograms of the complex in the absence (1) and increasing amount (2～5) of DNA

(1)

1/[DNA] =K(1-A)/[1-(i/i0)]-K

(2)

where [DNA] is the concentration of DNA,Kis the binding constant,iandi0are the peak currents with and without DNA,Ais the proportionality constant.

According to the calculation, a ratio ofKred/Koxwas calculated as 2.4, which indicates the reduced form of the complex bind to DNA is 2.4 times stronger than the oxidized form. Through the plot of 1/[DNA] versus 1/(1-i/i0) (see Fig. 7), the value ofKis 6.92×103L·mol-1, which is accord with intercalation mode. TheKvalue reveals the CT-DNA binding activity of the complex is weaker compare to the 2-methylpyridyl Zn(II)Ni(II) complex.

Fig.7 Plot of 1/[DNA] versus 1/(1-i/i0)

The viscosity method is one of the most direct and effective methods to study the interaction of complex with DNA[26]. When the complex binds to the DNA in the interaction mode, the double helix structure of DNA is opened and DNA chain becomes longer, which resulted in the viscosity of DNA solution increases[27]. The effect of the H2L and its Cu(II)Zn(II) complex on the viscosity of CT-DNA are shown in Fig.8. With the addition of increasing amounts of H2L or complex, CT-DNA viscosity increases slightly. The results indicate that the binding mode of the complex with CT-DNA is weak intercalation, which is according with the cyclic voltammogram studies. The CT-DNA binding activity of 2-methylpyridyl macrocyclic Zn(II)Ni(II) complex is notable higher than the present 3-methylpyridyl complex, maybe the reason of different metal coordination environment.

Fig.8 Effects of increasing amounts of H2L (1) and complex (2) on the relative viscosities of CT-DNA at (25.0±0.1) ℃; [DNA]=200 μmol/L

3 Conclusions

In summary, H2L and its Cu(II)Zn(II) complex were synthesized and structurally characterized. The X-ray diffraction results indicate that H2L and its Cu(II)Zn(II) complex crystallizes in different crystal systems and space group. The CT-DNA binding experiments were carried out through cyclic voltammetry and viscosity studies, which suggest that the complex bind to CT-DNA in a weak intercalation mode. The binding activity of the reported 2-pyridylmethyl macrocyclic complex is evidently higher than the present 3-pyridylmethyl complex, maybe caused by different metal coordination environment.