Binding of permanganate anion to pentaammineazidocobalt(III) cation in solution and solid phases: synthesis, characterization, X-ray structure, and genotoxic effects of [Co(NH3)5N3](MnO4)2⋅H2O

A pentaammineazidocobalt(III) complex, [Co(NH3)5N3](MnO4)2XH2O has been synthesized by an one-pot synthesis method. It was characterized by studies such as infrared (IR) and UV-visible spectroscopy. The single crystal X-ray structure analysis revealed that the title complex crystallizes in space group Cc. The cobalt center is six coordinated with slightly octahedral geometry. The supramolecular architecture is also formed by intermolecular N-H…O (anion and H2O) and Mn-O…O-H hydrogen bonds. The binding property of the cation, [Co(NH3)5N3]2+ with the anion, MnO4– has also been determined (in solution phase) with the help of UV-visible spectroscopic titrations. Further, the genotoxic effects of KMnO4, [Co(NH3)5N3]Cl2 and [Co(NH3)5N3](MnO4)2XH2O were studied using Allium cepa root chromosomal aberration assay and it revealed that the genotoxicity of the newly synthesized complex is 1.97–1.76 fold, which is less compared to KMnO4. The order of genotoxic potential has been observed to be KMnO4 > [Co(NH3)5N3](MnO4)2XH2O > [Co(NH3)5N3]Cl2.

[Co(NH 3 ) 5 2 .H 2 O have been determined using genotoxicity assay in Allium cepa root tip cells, which also has shown the importance to bind MnO 4 in aqueous medium.

Materials and methods
The reagents (AR grade of Merck) were used as such without any additional purification. [Co(NH 3 ) 5 N 3 ]Cl 2 was prepared according to the method reported by Linhard et al. [27]. Cobalt was determined by standard method [28] and H, N was estimated microanalytically by the automatic Eager 300 elemental analyzer. Solubility of the newly synthesized complex was measured at temperature 25 ± 2 ºC. The Shimadzu 1800 spectrophotometer was used for UV-visible spectra using a quartz cuvette (water as the solvent). For infrared spectra, a Varian Resolution Pro 660 FT/IR spectrophotometer was utilized as KBr pellets. XRD was recorded on a x-pert-pro PANalytical (Cu-Kα radiation, λ = 0.15418 nm) having angle range 5-60 o . The thermal behavior was recorded in temperature range from 25 to 500 ºC (with ramp of 10 ºC rate) using a TGA/DTA/NETZSCH STA 449F1 instrument, with ramp of 10 ºC .

Synthesis of [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O
Hot (60-70 o C temperature) aqueous solution (30 mL) of pentaammineazidocobalt(III) chloride (0.50 g, 0.0019 mol) was reacted with a hot (60-70 o C temperature) aqueous solution (15 mL) of potassium permanganate (0.615 g, 0.0077 mol) at room temperature. The mixture of solutions was allowed to cool slowly by keeping it overnight which resulted in formation of crystals. The crystals were filtered and air dried at room temperature. The violet colored clear supernatant solution (after 1 day) gave second crop of crystals. The overall yield was 0.774 g (90%). The melting point of the newly synthesized complex was observed to be 415 K (dec.

General procedure for Job's plot by UV-vis method
The stock solutions (1×10 -5 M) of receptor, [Co(NH 3 ) 5 N 3 ]Cl 2 and the guest, KMnO 4 were prepared in an aqueous medium. The absorbance in each case with different receptor (10:0)-guest (0:10) ratio but equal in volume (10 mL) was recorded. The Job's plots were drawn to find out the stoichiometries using continuous variation method at λ max 313 nm for manganate.

UV-visible titrations
The binding tendency of complex cation for manganate anion was determined using UV-visible titrations. The stock solutions of [Co(NH 3 ) 5 N 3 ]Cl 2 and KMnO 4 were prepared in concentration 1X10 -4 M and 1 X 10 -2 M respectively in aqueous medium. The titration were performed by adding increments of 5 μL solution of anion (each addition was made after 1 min) into stock solution (2 mL) of [Co(NH 3 ) 5 N 3 ]Cl 2 in a quartz cuvette (optical path length 1 cm). All absorption spectra were recorded, saved, and used to find the binding constant by fitting the data with the global analysis program Hypspec2014 11 [29] 2.4. Crystal structure determination Single-crystal diffraction data for [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O have been collected on a Bruker AXS SMART diffractometer equipped with CCD detector. More than a hemisphere of data was collected on each crystal over three batches of exposure using MoK a radiation (l = 0.71073 Å). A fourth set of data was measured and compared to the initial set to monitor and correct for decay, which was negligible. Data processing was performed using the program SAINT [30]. The absorption correction was done using an empirical method (SADABS) [31]. The structure was solved by the direct method and refined by the full-matrix least-squares method on all F 2 data using SHELX-97 [32]. All other information regarding the refinement is also recorded in Table 1. . The slides were prepared using squash method and were screened under microscope to score different types of aberrations. Three slides were scored for each concentration with at least 50-70 dividing cells.

Genotoxic potential of lead acetate
Estimation of genotoxic potential lead (0.5 ppm) was evaluated as per the protocol mentioned in section 2.5.1.

Antigenotoxicity of [Co(NH 3 ) 5 N 3 ]Cl 2 , [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O against lead
In order to see the sequential effects of newly synthesized compounds against lead induced genotoxicity, two modes of treatment i.e. pre and post were followed after germination of roots. In pretreatment, the onion roots were treated with [Co(NH 3 ) 5 N 3 ]Cl 2 for 3h followed by treatment with lead while in post treatment, roots were first exposed to lead solution (0.5 ppm) and then to [Co(NH 3 ) 5 N 3 ]Cl 2 . After treatment, the slides were prepared using standard protocol and screened for chromosomal aberrations in Allium root tip cells.

Synthesis
The single pot synthetic approach was used for this synthesis of title complex. Under this approach pentaammineazidocobalt(III) chloride and potassium permanganate were reacted in 1:2 molar ratio in hot aqueous medium with the expectation of [Co(NH 3 ) 5  containing complexes [20][21][22][23][24][25][26]. Solubility product: The newly synthesized complex is insoluble in organic solvent (CCl 4 , CHCl 3 ) but soluble in inorganic solvent (DMSO, MeCN) indicating the ionic behavior. Solubility of ionic complex in water differs to a great extent and on the basis of

Jobs plot and UV-visible titrations
After taking the UV-visible spectra of complex cation and anion (Figure 2a), the stoichiometry was determined by Job's plot (using continuous variation method) at different λ max 313 nm. Jobs plot is plotted between ΔA* XR or XG (mole fraction of receptor or guest) (along y-axis) and XR or XG (along x-axis) in the mixtures where the total concentrations of receptor and guest remain constant. It is used to identify the stoichiometry of the complex. The maximum were observed at 0.35 mole fraction with respect to receptor respectively, which implied the stoichiometry to be 1:2 (see Figure 2b).

Infrared spectrum
The infrared spectrum of [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O has been recorded in the range 400 to 4000 cm -1 and interpretations have been made on the basis of earlier reported literature [25,26,34]. In new cobalt(III) complex, the stretching vibration frequencies were lower for the coordinated NH 3 molecules than those of the free NH 3 . This lowering might be due to the formation of N-H…O type of hydrogen bonds which weaken the N-H bonds. It has been observed that the rocking vibrations (r r ), symmetric deformation (d s ), degenerate deformation (d d ), and antisymmetric (n as ) and symmetric (n s ) stretches appeared in the regions of 800-900, 1370-1200, 1650-1550, and 3400-3000 cm -1 respectively [25] [33].

TG/DT/DTG analysis
Thermal stability of title complex was examined between 20 and 800 ºC under nitrogen flow (Figure 4). The decomposition of complex started with the loss of water and azide in two steps, one at 117 ºC and other at 125 ºC . The complete loss (water and azide) occurs at 159 ºC (13.6 %) followed by loss of ammonia and decomposition of MnO 4 up to temperature 183 ºC . Both these decomposition steps were exothermic in nature (see Figure 4). After that not much change in weight loss was observed.

Single crystal X-ray diffraction (SCXRD)
The X-ray crystal structure of the [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O was unambiguously determined by single crystal X-ray crystallography. The crystal structure conclusively established the existence of single complex of composition, [Co(NH 3 ) 5 N 3 ] (MnO 4 ) 2 ×H 2 O and also ruled out the possibility of a double salt and mixture of salts. Furthermore, this study revealed for the first time that it is an ionic complex which contains discrete ions, [Co(NH 3 ) 5 N 3 ] 2+ and two MnO 4 in addition to one lattice water molecule in the solid state. The numbering scheme and ORTEP diagram of [Co(NH 3 ) 5 N 3 ](MnO 4 ) 2 ×H 2 O is shown in Figure 5a.
In [Co(NH 3 ) 5 N 3 ] 2+ , the cobalt(III) metal ion is surrounded by six nitrogen atoms originating from five coordinating ammonia molecules and one from azide molecule resulting in a nearly octahedral geometry. The Co-NH 3 [35].
In the crystal lattice of the title complex, a supramolecular architecture is also formed (see Table 3). It is formed by intermolecular hydrogen bonding between: 1) cation and anion (N-H(NH 3

)…O (MnO 4 -)), 2) solvent molecule (water) and cation (H-O (water)…H-N (cation)) and 3) solvent molecule (water) and anion (H-O-H (water)…O-Mn(anion)).
The hydrogen bonding parameters are given in Table 3. This supramolecular architecture stabilizes the crystal lattice along with electrostatic forces of attractions between cations and anions. All kinds of hydrogen bonding interactions are shown in Figure 5b.

Powder X-ray diffraction (PXRD)
The simulated PXRD pattern of title compound is given in Figure 6a. The simulated XRD pattern was compared with the SCXRD pattern obtained from the cif file (Figure 6b) of the compound and they are similar to one another. This indicates that SCXRD pattern obtained from cif file have same crystalline phase with respect to their bulk materials.