Synthesis , Structural Diversity and Mimic Superoxide Dismutase of Mn ( II ) Complexes Derived from N , O-donor Schiff bases

Two new potentially tetradentate Schiff base ligands N’-(pyridin-2-ylmethylene)nicotinohydrazide (L1), and N’-(pyridin-2-ylmethylene)isonicotinohydrazide (L2) were synthesized. Reactions of hydrazone ligands L1 and L2 with Mn(NO3)2 afford two mononuclear Mn(II) complexes, [Mn(L1)(NO3)(H2O)2]NO3 (1) and [Mn(L2)2(NO3)(H2O)]NO3 (2). For complexes 1 and 2, L1 and L2 act as pincer-like tridentate or bidentate ligands, respectively. The Mn(II) ions in the two compounds are both in hepta-coordinated environment, while the two molecules display diverse solid-state supramolecular structures because of the different orientation of Npyridine and hydrogen bonding patterns of nitrate anions. Complex 1 features 2D supramolecular sheet, while complex 2 has double-chain supramolecular structure. Both of the complexes exhibit moderate superoxide dismutase (SOD) mimetic activity.


Introduction
Schiff base compounds derived from the condensation reaction of aldehydes with amines are very attractive for their coordination ability to metal atoms, [1][2][3] and excellent biological applications. 4,5Among the variety of aldehydes, salicylaldehyde derivatives are mostly used.7][8] Aygün et al. have also reported tridentate Schiff base ligand derived from phenylalanine and 2,4-dihydroxybenzaldehyde. 9 While Schiff base ligands based on pyridylaldehyde derivatives are still rare or lacking. 10,11Aroyl hydrazones are an important class of Schiff base ligands, coordinating through amide oxygen and the imine nitrogen of hydrazone moiety. 10,12Moreover, N and O heteroatoms of the aroyl hydrazones with free electron pairs can also be considered as potential hydrogen bond acceptors to expand polymeric frameworks with hydrogen-bonding interactions.
Numerous diseases, such as amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease are considered to be connected with the over production of superoxide radical anion.Superoxide dismutase (SOD) can accelerate the dismutation of superoxide anion into hydrogen peroxide and dioxygen, which play an important role in defending life from the diseases mentioned above. 13Three isoforms of SOD have been found, including Cu/Zn-SOD (SOD 1), Mn-SOD (SOD 2) and extracellular SOD (SOD 3).Compared with native SOD enzymes, low molecular weight SOD mimics have advantages in lack of immunogenic responses, improved access to lipid layer of the cell membrane, and low cost. 14Therefore the rational design and synthesis of SOD mimics has potential in metal-based therapeutic agents.Mn complexes derived from Schiff base, porphyrin and their derivatives have been studied as Mn-SOD mimics. 15n this contribution, we present the synthesis of two new hydrazone Schiff base ligands N'-(pyridin-2ylmethylene)nicotinohydrazide (L 1 ) and N'-(pyridin-2ylmethylene)isonicotinohydrazide (L 2 ) (see Scheme 1), and two novel mononuclear Mn(II) complexes 1 and 2 generated from these two hydrazone ligands and Mn(NO 3 ) 2 , the structures and IR spectra, as well as the SOD mimetic activities.

X-ray Crystallography
Diffraction intensities for complexes 1 and 2 were collected at 298 K on a Bruker Smart Apex CCD diffractometer equipped with graphite-monochromated Mo Kα (λ = 0.71073 Å) radiation using a ω-2θ scan mode.The collected data were reduced using the SAINT program, 17 and multi-scan absorption corrections were performed using the SADABS program. 18The structures were solved by direct methods and refined based on F 2 by full-matrix least-squares methods were performed with the SHELXTL program packages. 19All non-hydrogen atoms were refined with anisotropic displacement parameters.All the hydrogen atoms bonded to C atoms were generated geometrically and refined isotropically using the riding model.While hydrogen atoms bond to N or O atoms were first found in the Fourier map and then fixed at their ideal positions.Water H atoms were refined with distance restraints of O-H = 0.85(1) Å, H•••H = 1.44(2)Å and U iso (H) = 1.5U eq (O).Imine H atoms were refined with distance restraints of N-H = 0.89(1) Å, and U iso (H) = 1.5U eq (N).In complex 2, O5 was disordered over two positions that refined to a ratio of 0.50(4):0.50(4).Further refinement details of the structural analysis for the two complexes are given in Table 1.Selected bond lengths and angles of the complexes are listed in Table 2.

3. Superoxide Dismutase Assay
The SOD activity of the title complexes was studied by indirect method using Superoxide Dismutase Assay Kit according to the instruction of the manufacturer.Briefly, the xanthine-xanthine oxidase system was used for the production of superoxide.For the detection of superoxide the absorbance increases at 510 nm arising from the reduction of iodophenyl nitrophenyl phenyltetrazolium salt (INT).The reaction mixture contained 4.2 • 10 -5 M xanthine and 8.25 • 10 -5 M INT in 0.05 M phosphate buffer solution at pH 7.8.The reaction was started by the addition of xanthine oxidase in the amount needed to generate the absorbance change 0.03-0.04min -1 .The SOD mimic activity of the complexes 1 and 2 was evaluated from the absorbance decrease comparing to the blank after the 10 min incubation at 37 °C.The IC 50 value was defined as the concentration of complex (μM) necessary for the 50% inhibition of the INT reduction.

2. X-ray Crystallographic Analysis
Complex 1 crystallized in the triclinic space group Pl -.L 1 serves as tridentate chelating ligand, the pyrimidine N-atom from nicotinoylhydrazine is not implicated in complexation.The coordination geometry around the Since the anionic moieties NO 3 ¯possess the hydrogen bond acceptors 22 and the water molecules possess the hydrogen bond donors, 23 complex 1 presents enhanced hydrogen-bonding framework in the solid state (Table 3).The adjacent mononuclear cations are seized together through the strong hydrogen bonding interactions (O8-H8B•••O2 i , symmetry code: (i) -x + 1, -y, -z + 1) between coordinated water molecules and NO 3 ¯into a 8membered dimeric unit with the Mn•••Mn separation of 5.407(1) Å.As shown in Figure 2, the free nitrate anions are located between these dimeric units, and serve as hydrogen bonding acceptor linking these dinuclear units into infinite double-chain structures along a axis via O8-H8A•••O5 and O9-H9B•••O6 ii (symmetry code: (ii) x -1, y, z) hydrogen bonds.These supramolecular chains stack in a face-to-face fashion in ac plane, the hydrogen bonds exist between the amido group of ligand L 1 and the oxygen atom of free nitrate to form an intermolecular N3-H3A•••O7 iii (symmetry code: (iii) -x + 1, -y, -z) hydrogen bonding interaction, leading to the construction of 2D supramolecular sheet in the ac plane (Figure 3).
The idea behind the use of ligand L 2 is to control supramolecular motifs through a 4-pyridine-type ligand.It is known that the relative different orientation of the nitrogen donors on the pyridyl rings could result in unusual building blocks, which can lead to the construction of versatile supramolecular motifs.Crystallization of L 2 with Mn(NO 3 ) 2 for the self-assembly reaction in the CH 2 Cl 2 /EtOH solvent system afforded [Mn(L 2 ) 2 (NO 3 ) (H 2 O)]NO 3 (2).Complex 2 also crystallized in triclinic    space group Pl -.An ORTEP view of 2 together with the atom numbering scheme is given in Figure 4.The complex 2 also showed pentagonal bipyramid geometry with a slight distortion as evidenced from the sum of five equatorial angles of 361.74° compared to the ideal value of 360°f or planar geometry, and the axial O9-Mn1-O3 angle of 164.05( 8) is also smaller than the ideal value of 180°.The Mn-O bond distance (2.228(2)--2.4148(16)Å) and Mn-N bond distance (2.2894(18)-2.3915(18)Å) are comparable to those found in complex 1.

IR Spectroscopy
The spectra of complexes 1 and 2 exhibit broad bands at 3350 and 3383 cm -1 , respectively, which can be attributed to v(OH) of the crystallized water molecules, and sharp peaks at 1556 and 1553 cm -1 , respectively, due to H-O-H bending vibrations. 24The characteristic v(NH) stretches are shown at 3220 cm -1 for 1 and 3178 cm -1 for 2. 25 The sharp absorption bands of NO 3 -in 1 are clearly visible at 1381 and 823 cm -1 , while in 2 are at 1384 and 850 cm -1 . 26In the complexes 1 and 2, the involvement of the carbonyl O and azomethine N in coordination with Mn atom weakens the C=O band and C=N bond leads to the v(C=O) and v(C=N) towards lower frequencies in the region of ca.1648 cm -1 and 1598 cm -1 , respectively. 24

4. SOD Mimetic Property
Interest in Mn(II) complexes has been amplified due to their application in biology, 15,27 e.g., Mn-MRI (magnetic resonance imaging) contrast, Mn-fluorescence agent, and Mn-SOD biomimetics.In previous reports, Mn(II) complexes of pentagonal bipyramidal geometry are functional mimics of SOD. 14,15,28In this paper, we studied the SOD mimetic activity of complexes 1 and 2. The reported manganese complexes with various organic ligands (such as macrocyclic penta-amine, tripodal polyimidazole, di-Schiff base) showed IC 50 values rank from 0.30 to 35 μM. 14,29In our study, 1 and 2 possess moderate activity of dismutation of superoxide with IC 50 values of 32.5 and 21.0 μM, respectively.The IC 50 values reported here are much larger than those of Mn(II) complex generated from di-Schiff base ligand (2.85 μM). 14This one order of magnitude difference may be ascribe to the rigid planar structure of 1 and 2, which possess small tendency to fold. 28herefore the Mn(II) center is constrained in a geometry that limits fast electron-transfer, i.e., a pseudo octahedral geometry preferred by Mn(III).

Conclusion
In summary, nicotinoylhydrazine and isonicotinoylhydrazine were used to synthesize hydrazone Schiff base ligands L 1 and L 2 .Further coordination reactions of these ligands with Mn(NO 3 ) 2 afforded mononuclear complexes 1 and 2, respectively.Although L 1 and L 2 are potential NNNO tetradentate ligands, in complexes 1 and 2, The crystal packing of complex 2 is stabilized by means of hydrogen bonding (Table 3) in combination with π-π stacking interactions.Similar to complex 1, in complex 2, as shown in Figure 5a, the adjacent molecules are linked through hydrogen bonding interactions to form dimers with Mn•••Mn separation of 10.562(6) Å.The intermolecular hydrogen bond exists between the hydroxyl group of coordinated H 2 O molecules and nitrogen atom of pyridine ring (O9-H9B•••N7 iv , symmetry code: (iv) -x + 1, -y + 1, -z).Meanwhile, the centroid-to-centroid separation of two parallel pyridyl rings is found to be 3.

Supplementary Information
CCDC files 1023096 (1) and 1023097 (2) contain the supplementary crystallographic data for this paper.These data can be obtained free of charge from The Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/data_request/cif.

Table 3 .
Geometrical parameters for hydrogen bonds