Molecular Structure, Spectroscopic, Frontier Molecular Orbital Analysis, Molecular Docking Studies, and In Vitro DNA-Binding Studies of Osmium(II)-Cymene Complexes with Aryl Phosphine and Aryl Phosphonium Assemblies

X-ray crystallography, spectroscopy, computational methods, molecular docking studies, and in vitro DNA-binding studies have been useful in the investigations of intermolecular and intramolecular interactions of osmium-cymene oxalato complexes with aryl phosphine and aryl phosphonium groups in both primary and secondary coordination spheres, respectively. Molecular structures of the novel complexes PPh4[Os(η6-p-cymene)Br(κ2-O,O′-C2O4)] (1) and [Os(η6-p-cymene) (κ2-O,O′-C2O4)PPh3] (2) were resolved by single-crystal X-ray diffraction (XRD). Primary and secondary coordination sphere contacts were investigated using Hirshfeld surface analysis which was supported by molecular docking (MD) studies. The MD data obtained predicted significant differences in binding energy across three receptors for the two osmium complexes. An in vitro DNA-binding study was accomplished using UV-Vis spectroscopy which showed that both 1 and 2 bond with DNA through an intercalation approach. The optimized molecular geometry, frontier molecular orbital (EHOMO and ELUMO) energies, global electrophilicity index (ω), chemical hardness (η), chemical potential (µ), and the energy band gap (EHOMO–ELUMO) were calculated utilizing density functional theory (DFT) methods. Computed structural parameters (bond lengths and angles) support the experimental single-crystal XRD data.


Introduction
Within cancer chemotherapy treatment, platinum-based complexes, such as Cisplatin, Carboplatin, and Oxaliplatin, have achieved remarkable success and constitute one of the best extensively utilized classes of metallodrugs [1][2][3][4][5].Tese complexes have a similar chemical structure, efecting a square planar geometry with the central Pt(II) ion bearing a pair of N-donor moieties as the stable or nonleaving group ligands and two labile ligands such as halides or an O,O′chelator (Figure 1).However, despite the success of these metallodrugs in treating cancer, their clinical application is constrained by undesirable efects, specifcally neuro-, hepatic-, and nephrotoxicity as well as inherent or acquired resistance.To overcome the limitations of platinum-based metallopharmaceuticals, numerous research initiatives to fnd more metal complexes, which have potential applications in cancer treatment, have improved signifcantly in the area of medicinal inorganic chemistry [6][7][8].Most notable is the development of ruthenium-based RAPTA-type complexes that have 1,3,5-triaza-7-phosphaadamantane (PTA) group, including the corresponding osmium analogues, which both exhibit promising anticancer properties [9][10][11][12].
Te oxalato ligand performs a critical function in the anticancer activity of the platinum-based drug, Oxaliplatin.Te O,O′-chelator remains bound to platinum in vivo until the complex enters the cell cytoplasm, where the oxalato ligand is then replaced by chloride ligands to activate the complex [1,13].Te activated complex can then covalently bind to the imidazole N7 of guanine.Hydrogen bonding interactions have been found to perform a critical task in the stabilization of the metal-DNA adducts formed by the activated complex [14].However, when bound to a diferent metal, such as osmium, the oxalato moiety could function as an intercalating agent, inducing conformation changes to the DNA, and consequently disrupting replication and transcription.
In coordination complexes, ligands connected directly to the metal centre comprise the primary coordination while ligands which are not directly linked but are bound to the metal through noncovalent interactions comprise the secondary coordination sphere.Te outer coordination sphere can be manipulated through ligand modifcation to direct the reactivity of the metal complex and has also been shown to be key in the functioning of metalloproteins [15].Te secondary coordination sphere in metalloproteins is controlled by weak electrostatic forces and plays a key role in molecular recognition as well as in infuencing the reactivity and stability of the molecule [16,17].
Hirshfeld surface analysis is rapidly gaining traction in molecular structure research.Tis tool ofers novel and key understanding of the intermolecular interactions in molecular crystals.Hirshfeld surfaces are particularly valuable for complexes where the surface morphology is not just a consequence of intermolecular packing but also refects the delicate balance of forces between individual atoms within the molecules, making them powerful tools for deciphering the intricate interplay of these interactions within the crystal lattice [18].Besides the visual map, fngerprint plots ofer a quantifable breakdown of the diferent intermolecular relations, revealing their relative infuences concerning the molecule's stability [19].
Molecular docking analysis is a key tool for simulating the interactions between proteins and transition metal complexes [20].Tis computational tool gives information about the potential for binding between proteins and complexes, the binding energies, the binding positions on the protein, and the nature of interactions [21].
DNA is the key carrier of genetic material and has been broadly investigated as a primary target for numerous metallodrugs.Generally, the binding of metal complexes to DNA arises via three approaches of connecting that is: intercalation, electrostatic attraction, and groove binding [22][23][24][25][26]. Terefore, an in vitro DNA binding study is key to understanding interactions with metal compounds [27].Te most widely used method for probing in vitro DNA binding with metal complexes is with electronic absorption spectroscopy [28,29].
Tis study investigates the infuence of primary and secondary coordination sphere phosphine and phosphonium groups on the vibrational spectroscopy, molecular docking, and hydrogen bonding contacts of osmium complexes PPh 4 [Os(η 6 -p-cymene)Br(κ 2 -O,O′-C 2 O 4 )] (1) and [Os(η 6 -p-cymene) (κ 2 -O,O′-C 2 O 4 )PPh 3 ] (2) (Scheme 1).For the complexes reported in this project, the designed and prepared Os(II) complexes contain structural features found in the platinum and ruthenium-based metallodrugs cited above.In this study, the structural motif of the PTA ligand has been emulated by the triphenylphosphine ligand which has been shown to enhance the hydrophobicity of the metalarene complex, efecting increased levels of cytotoxicity in vitro [30].
In our study, the osmium complexes were docked against the human serum transferrin, human serum albumin, and DNA duplex.Te proteins were selected based on their involvement in cancer tumour growth or their role as transporting agents that infuence drug movement within the body.In addition, the molecular structures have been resolved utilizing single-crystal X-ray difraction while the molecular docking fndings were confrmed through conducting in vitro DNA-binding studies for complexes in this study.

Instrumentations.
Te solid-state IR spectroscopy data were collected from a Bruker Vertex 70 Fourier Transform-Infrared instrument employing an attenuated total refectance (ATR) element, with a resolution of 2 and 32 number of scans with a range of 4,000-400 cm −1 .Raman spectral data were collected on a Bruker Raman II instrument with a resolution of 4 and 128 number of scans in the range of 5000-0 cm −1 .UV-Visible spectroscopy information was collected using a Shimadzu UV-Vis 1800 instrument between wavelength range 250 and 800 nm.Melting point data were collected from Mettler Toledo MP50 Melting Point System.
Nuclear Magnetic Resonance (NMR) information was obtained from 500 MHz Agilent Technologies instrument.Tetramethyl silane (SiMe 4 ) was used as an external reference standard for both 1 H and 13 C NMR studies, whereas for 31 P studies, phosphoric acid (H 3 PO 4 ) was employed for the external reference standard.Te Agricultural Research Council-Institute for Soil, Climate and Water (ARC) used the Carlo Erba NA 1500 (Nitrogen, Carbon and Sulfur) to collect microanalysis data.
Difraction data for the molecular structures of 1 and 2 were obtained from a Bruker D8 Venture Photon CCD area detector difractometer at 173(2) K. Data reduction was executed by SAINT-Plus and XPREP and structure solutions were solved by SHELXS97 [31].Structure refnements were performed using SHELXL2014/7 [32] and molecular graphics were performed by ORTEP for Windows [33] while WinGX publication routine software [33] was used to prepare material for publication.

Molecular Hirshfeld Surfaces Calculations.
Hirshfeld surface plots of complexes 1 and 2 were created using Crystal Explorer 17 [34][35][36].Hirshfeld surfaces were utilized to establish intermolecular contacts involving H Tree-dimensional (3D) Hirshfeld surface diagrams were produced with the d norm (normalized for the atom size) surfaces mapped over a static red-, whiteblue colour system signifying short interactions, van der Waals interactions, and longer interactions sequentially.Te typical 0.6-2.6Å view was utilized to create the twodimensional (2D) fngerprint maps, with the plot axes displaying d e and d i distance scales.Two-dimensional fngerprint plots for 1 and 2 were defned for several contact types, including the H to evaluate and illustrate the infuence of polar and nonpolar contacts towards the crystal packing forces.

Computational Experimental Section.
All the computations were computed by DMol 3 DFT program as employed in the Accelrys Material Studio ® version 2018 software package [37,38].All geometry optimizations were accomplished using the nonlocal generalized gradient approximation (GGA) utilizing the Perdew-Burke-Ernzerhof (PBE) exchange-correlation functional [39].In this study, core electrons of the Os were taken into consideration using a DFT semi-core pseudopotential in conjunction with double numeric, polarised split valence (DNP) basis set.While the DNP basis set is equivalent in size to the Gaussian 6-31 G * * basis set, the DNP is utmost exact [40].Optimizations of the geometries were done with unrestricted spins and no symmetry constraints.Tese optimizations' convergence criterion included the following threshold values: a self-consistent feld density convergence threshold of 1 × 10 −5 Ha was provided, whereas the following values were given for energy, gradient, and displacement convergence: 2 × 10 −5 Ha, 0.004 Ha Å−1 , and 0.005 Å, respectively.To authenticate the nature of the stationary positions, a comprehensive frequency analysis using the equivalent theoretical level (GGA/PBE/DNP) was performed on all optimized geometries.Te absence of imaginary frequencies was a characteristic of the optimized geometries.

Molecular Docking Study.
Te rigid molecular docking studies on the osmium complexes were conducted following a method described by Atlam and co-workers [41] employing Hex 8.0 software [41].Te structural coordinates of the complexes were obtained from the crystallographic information fles (CIF Files) and then geometrically optimized by GGA/PBE/DNP using DMol 3 density functional theory (DFT) software, followed by converting the fle format to PDB.Te structure of the receptors, human serum transferrin (PDB ID: 1D3K), DNA duplex (PDB ID: 1XRW), and human serum albumin (PDB ID: 1H9Z) were obtained from Protein Data Bank (https://www.pdb.org/pdb/home/home.do).All cocrystallized water molecules, ligands, and co-factors were eliminated from the protein structure before molecular docking computations were embarked on with the studied complexes.In the molecular docking calculation, molecules were displayed using 3D parametric functions that determine both surface shape and electrostatic charge.Te docked poses were visualized using Discovery Studio 2020.

Assessment of DNA-Binding Activity by UV-Visible
Spectroscopy.Electronic absorption spectra for 1 and 2 were analysed in DMSO using the range of 250-800 nm.Stability studies for both complexes using UV-Vis spectroscopy were conducted over the period of 3 hours, at 15-minute intervals, to examine the activities and stability of the 1 and 2 in the chosen solvent system (DMSO and Tris bufer) prior to performing DNA titrations.Te DNA-binding studies of reported complexes were accomplished using tris (hydroxymethyl) aminomethane bufer (5 mM Trizma base, 50 mmol NaCl, pH 7.2).Te DNA stock solution was produced by diluting 200 microlitres of CT-DNA in 10 mL of tris bufer solution.Molarity of CT-DNA was recorded spectrophotometrically at UV 260 , using molar absorptivity (ε 260 � 6600 M −1 cm −1 ) [22,28,42] and was found to be 6.20 × 10 −5 M. Te DNA stock solution was preserved in a freezer below 15 °C and used in less than 96 hours.Stock solutions of reported complexes in dimethyl sulfoxide were made and diluted further using the bufer to the necessary concentration (1 × 10 −4 M) [42].To regulate possible interactions of CT-DNA with the complexes, a fxed concentration of the compounds were used, with varying increments of DNA stock solution being augmented to the sample and reference chambers, to eradicate possible absorption of free CT-DNA [28].Te combination was nurtured for 15 minutes ahead of the analysis of absorption spectra at ambient temperature.

4
Bioinorganic Chemistry and Applications

Vibrational Spectroscopy.
Te uncoordinated oxalato anion adopts a nonplanar conformation with approximate D 2d point group symmetry which has the irreducible representation given by Γ � 3A1 + B1 + 2B1 + 3E.However, upon coordination as a bidentate ligand (κ 2 -O,O′-C 2 O 4 ), a planar conformation is adopted and the symmetry of the oxalato ligand is reduced to C 2v where the irreducible representation is given by Γ � 6A 1 + 2A 2 + 5B 1 + 2B 2 .In this case, the Raman and Infrared modes were all active [43,44].
Te CO symmetric and asymmetric stretching bands of the carboxylate groups were found in the ranges 1 500-1 400 cm −1 and 1 700-1 500 cm −1 , respectively [45].In this study, complexes 1 and 2 show some diferences in the CO stretching bands which may be attributed to the complex charge and the ancillary ligand.However, both Infrared and Raman bands of 1 and 2 exhibit some inclusions with the occurrence of nearcoincidence, which is associated with the C 2v point group of the oxalato ligand [46].Despite the similarities in spectral appearances, we have observed some diferences in the peak splitting in the Infrared and Raman data.

Vibrational Spectroscopy of COO Bands.
Te Infrared and Raman bands of complexes 1 and 2 were assigned with reference to other previously reported transition metal oxalato complexes [47][48][49][50][51][52][53][54][55].Te Infrared (2000-400 cm −1 ) and Raman data (2000-0 cm −1 ) for 1 and 2 are presented in Supplementary Figures S1-S4 in the supplementary data section and the proposed assignments are reported in Table 1.Te IR data of 1 and 2 exhibit strong stretching bands at 1695s, 1674s, 1653 s cm −1 and 1706sh, 1693s, 1669s respectively, which have been assigned to the ] asym (OCO) mode.Te Raman spectra exhibit weak bands for this stretching frequency in both 1 and 2 which correlate with the IR data.Te stretching bands assigned as ] sym (OCO) + ](CC) mode in the Infrared spectra appeared at 1483w, 1436m cm −1 and 1482w, 1467vw, 1433m cm −1 for 1 and 2 successively.Corresponding Raman vibrational modes were observed at 1483vw, 1460vw, 1440vw cm −1 for 1 and 1483vw, 1455vw, 1440vw cm −1 for 2. Te symmetric ](OCO) bands in the Infrared spectra of 1 were observed at 1378s/ 1317w cm −1 as both strong and weak bands, whereas for 2 these bands are observed as a single strong band with two shoulder bands at 1371sh, 1363sh, and 1356s cm −1 .Raman spectra show both weak and medium bands for 1 at 1384m, 1317vw cm −1 ; however, a single weak vibration at 1364w cm −1 was detected for 2 corresponding to the ] sym (OCO) mode.Te mode of coordination of the O,O′-chelating ligand in these complexes is consistent with a ∆ value > 200 cm −1 where ∆ � ] asym (OCO) − ] sym (OCO) [56].Te coordination mode of the oxalato ligand in 1 and 2 was additionally supported by X-ray difraction information.
Te ](CC) mode for 1 was observed as three bands in the Infrared spectrum at 910vw, 886vw, and 859vw cm −1 .Two signals at 904vw and 875w cm −1 observed for 2 were assigned as the ](CC) mode for this complex.Te Raman spectra showed very weak single bands for both 1 and 2 at 907vw and 889vw cm −1 for the ](CC) mode.Weak and strong bands at 808w and 787s cm −1 in the IR spectra for 1 was recognised as ](CC) + δ(OCO), whereas for 2 these peaks coalesce to a single strong band at 786s cm −1 attributed to electronic efects of ancillary ligands.Raman spectra exhibit a medium intensity band corresponding to the ](CC) + δ(OCO) mode at 807m cm −1 for 1 and 797m cm −1 for 2. Stretching bands were designated as ](OsO) + ](CC) for 1 at 530vs, 521vs cm −1 which for 2 similarly coalesce to a single band at 530vs cm −1 .Te ](OsO) + ](CC) mode in the Raman data shows a band at 537vw cm −1 for 1 whereas at 534w cm −1 for 2.

Vibrational Spectroscopy of M-P, M-O, and M-X
Bands.Te comparison of 1 and 2 indicates that the Infrared vibrations at 510s and 495m cm −1 can be attributed to ] asym (Os-P) which, in contrast, exhibited as a very weak band at 495vw cm −1 in the Raman data.Te symmetric bands ] sym (Os-P) assigned at 437m cm −1 in the IR showed a corresponding broad weak band at 437w cm −1 in the Raman spectrum.Furthermore, the Raman spectra show the Os-O bands at 415sh, 391m cm −1 for 1, and 405m cm −1 for 2 (see Table 1) [57].Te infuence of intramolecular interactions in 2 may account for the alteration of the Os-O medium band by ca. 15 cm −1 towards higher energies than that observed at ca. 390 cm −1 for 1. Te Os-Br band in 1 is found within the expected range in literature [58].

Multinuclear Magnetic Resonance Spectroscopy.
Te 1 H NMR data of 1 and 2 agree with the proposed structures.Two pairs of doublets attributed to the p-cymene aromatic protons appear for complex 1 at δ 5.94 and 5.67 ppm.Tese two sets of doublets for 2 appear at δ 5.54 and 5.32 ppm.Surprisingly, these 1 H signals of 2 are shielded relative to 1 despite the negative charge on the latter complex ion.Te reduced electron density in the p-cymene ring is attributed to the π-acceptor properties PPh 3 group.
Te 13 C resonance peaks on the ring of the p-cymene ligand of 1 were observed in the range of δ 89.05-68.86ppm, whereas for 2 these signals were found in the range of 98.56-78.93ppm.As expected, 1 and 2 exhibit diferent oxalato CO signals, at δ 166.89 and 164.11 ppm, respectively, due to changes in the ligand system of the two complexes.
Te 31 P NMR information of 1 and 2 further confrmed the positions of the PPh 4 + and PPh 3 groups in the outer sphere and inner sphere, respectively.Complex 1 gave a 31 P signal at 24.07 ppm slightly shifted from 23.09 ppm of the PPh 4 Br precursor.Coordination of PPh 3 to the metal ion was observed to have shifted the 31 P signal of 2 from -5.53 ppm to 1.78 ppm.In addition, the 187 Os satellite peaks confrming the direct Os-P bond were observed with J( 187 Os-31 P) � 309 Hz which is consistent with previously reported osmium(II) complexes [59].

Bioinorganic Chemistry and Applications
Molecular structures of 1 and 2 have been elucidated using single-crystal X-ray difraction (see Figures 2 and 3).Complexes 1 and 2 are both pseudo-octahedral, with the hexahaptic p-cymene group dominating three coordination positions, and crystalizing in the monoclinic crystal system, with space group P2 1 (no.4).Te cationic counterion of 1 displays a distorted tetrahedral geometry.
Selected bond lengths, bond, and torsion angles comparing the single-crystal XRD data and DFT-calculated geometrical parameters of complexes 1 and 2 are included in Table S1.Te Os-Br bond distance in 1 at ca. 2.53 Å is within the range of terminal bromide ligands in previously reported Os(II)-arene complexes [59][60][61].Te Os-O bond distances in 1 were measured at 2.099(4) Å and 2.100(4) Å, whereas the corresponding Os-O bond distances in 2 were determined to be 2.093(3) Å and 2.078(3) Å.For 2, the oxalato ligand was found to bind asymmetrically to the osmium centre.Tis may be due to the larger steric requirement of the PPh 3 ligand.Te Os-P bond distance at ca. 2.35 Å observed for 2 is similar to related Os(II) complexes [59][60][61][62].
Te osmium-cymene centroid distance in 2 was found to be slightly longer than in 1 which was attributed to the inner sphere infuence of the osmium bound PPh 3 ligand.Consequently, the osmium-carbon bond lengths of 1 were found to be marginally shorter relative to 2 due to the sterically demanding coordinated PPh 3 group.Te strong σ-donor and π-acceptor ability of PPh 3 efected elongated Os-C bonds trans to the P-donor atom.Both 1 and 2 showed loss of aromaticity of the p-cymene group evidenced by varying shorter and longer carbon-carbon bond lengths within the ring.
Te bite angle O1-Os-O3 of 1 is slightly reduced compared to 2. Tis is unexpected as complex 2 would be expected to have a large bite angle due to the steric demand of the large PPh 3 .However, the observed increase in bite angle may be due to electronic factors generated by O-atom lone pair repulsions within the chelate ring of 2. Te combined covalent radii of the Os(II) ion and O donor atom at 2.1 Å are consistent with Os-O single bonds in the metallacycle moiety of both 1 and 2 in this study [63].Lack of planarity of the metallacycle in both 1 and 2 is indicated by the nonzero torsion angle O1-C11-C12-O3 which is large for 2 because of the steric requirements of the bulky PPh 3 ligand.

Hirshfeld Surface Calculations.
From the XRD data, the structure of 1 shows the PPh 4 + cation and the anionic complex [Os(η 6 -p-cymene)Br(κ 2 -O,O′-C 2 O 4 )] that are held jointly via two C-H . . .O and one C-H . . .Br interactions.Furthermore, one C-H . . .O intramolecular hydrogen bond exists (see Figure 2).Tis C-H. ..O contact defned by the osmium coordinated oxygen atom and the methine hydrogen of the p-cymene isopropyl group was measured with a distance of 2.673 Å and an angle of 149.01 °.Te C-H . . .O intermolecular hydrogen bond observed links at the same O atom and an H atom of a phenyl group measured 2.479 Å at 149.22 °.Te second C-H . . .O intermolecular hydrogen bond observed between a carbonyl O atom and a hydrogen of the phenyl group measured 2.582 Å at 145.22 °.Te third C-H . . .Br intermolecular hydrogen bond detected between the Br atom and a hydrogen of the phenyl substituent measured 2.996 Å at 150.08 °.Te latter intermolecular hydrogen bonding is classifed as a weaker hydrogen bond.
Complex 2 shows a C-H . . .O intramolecular hydrogen bonding owing to one of the O atoms coordinated to the Os metal centre and a C(sp 2 )-H group of one of the phenyl groups measured 2.808 Å at 128.39 °(see Figure 3).Tis interaction is considered a weak contact because of the long range and the angle which deviates signifcantly from linearity [64].A second intramolecular hydrogen bond between a metal-coordinated O atom and a C(sp 3 )-H of the propan-2-yl substituent on the p-cymene fragment measured 2.545 Å at 133.46 °.
To obtain a greater understanding into the efect of intermolecular forces on the geometry of the osmium complexes, a comparative Hirshfeld surface study of the two complexes was conducted.Hirshfeld surfaces mapped over d norm functions of 1 and 2 are presented in Figures 4 and 5 successively.
In the two-dimensional fngerprint plots, it is understood that the molecule functions as an acceptor if d i > d e but the molecule is a donor if d i < d e .Complex 1 exhibits three short contacts C-H . . .Br, C-H . . .O, and C-H . . .H in XRD.Consistency is observed between the difraction data and Hirshfeld Surface calculations for 1. Mapping of 1 as visualized in Figure 4(a) shows several contacts and their contributions (see Figure S5(a) in supplementary data section) were calculated as follows: hydrogen bonds [H• • •H where X cal.and X exp .are the computed and the experimental information, in sequence.RMS errors of the bond lengths and internal angles are 0.0918 Å and 0.7412 °for complex 1 and 0.0974 Å and 0.6327 °for complex 2 successively.

Frontier Molecular Orbital (FMO) Analysis.
To gain an understanding of the behaviour of the electronic transitions within the complexes investigated in this study, a quantitative analysis of key quantum chemical parameters of the molecular species was carried out.DFT simulations provided insights into the electronic structures of 1 and 2, including their energy levels, bonding patterns, and reactivity.Analysing the DFT-derived global reactivity descriptors, particularly the interplay between chemical hardness (η) and electrophilicity (ω), revealed fascinating insights into the relative stability and reactivity profles of the complexes.Tis information provided valuable clues for understanding their potential applications and behaviour.Te frontier molecular orbital (FMO) energies are widespread quantum mechanical descriptors since the orbitals were illustrated to perform the key function towards infuencing various chemical reactions.Te highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) energies are associated to gas phase ionization energies (IP) and electron afnities (EA) according to Koopmans' theorem [66,67].Te computed reactivity indices are summarized in Table 3.
Chemical hardness (η), a concept rooted in frontier molecular orbitals, quantifes a molecule's resistance to changes in its electron distribution or charge transfer.Tis resistance is directly connected to the energy gap involving the HOMO and the LUMO, as described by equation (2).A wider HOMO-LUMO gap signifes a higher η, implying greater stability and reduced reactivity.
Designated as the negatively charged electronegativity and computed by equation ( 3), electronic chemical potential (μ) captures the ease with which electrons can escape from a molecule.A higher μ indicates a greater tendency for electron loss, making the molecule less stable and more reactive [68].
Introduced by Parr and calculated using equation ( 4), the electrophilicity index (ω) refects the molecule's tendency to accept electrons.It combines the stabilizing efect of gaining an electron (μ) with the resistance to electron redistribution (η), providing a quantitative measure of electron-loving power.
It can be seen from the molecular orbital diagrams in Table 4 that the HOMO and LUMO of 1 are confned to the osmium anionic and phosphonium aryl moieties, respectively, whereas in 2 the HOMO is mainly scattered over the oxalato ligand and the LUMO is extensively dispersed over the osmium-phosphine segment of the complex.Te HOMO signifes the dissemination and energy of the least tightly held electrons in the compound while the LUMO identifes the moiety of the compound where the addition of electrons is most probable.Based on the relative HOMO energies of the complexes, it is expected that 1 would more readily donate its electrons and undergo reduction.According to the HOMO-LUMO band gap values, 1 also has the lower excitation energy and would thus exhibit a higher chemical reactivity and lower kinetic stability.A small global hardness (η) means that the complex has high polarizability.Te smaller η value of 1 indicates that the two-component ionic osmium moiety has higher polarizability relative to the neutral complex 2, which is expected to exhibit greater resistance towards deformation of its electron cloud under small perturbations.Te negligible diference in electronegativity indicates that both 1 and 2 have a similar capacity for attracting electrons from the neighbouring molecules.Te electronic chemical potential (µ) is a property of an equilibrium state which indicates that both complexes have comparable capacities for changes in electron density and are expected to undergo similar electron density fux in an interacting system.

Molecular Docking Studies.
Molecular docking results of 1 and 2 are reported in Table 5. Te Hex docking results reveal that 1 has an improved binding potential than 2 due to the lower free energy indicating better interaction.Across diferent receptors, complex 1 showed stable binding energies with an average of −284.1 ± 4.56 kJ/mol while 2 showed variations between diferent receptors with an average energy of −214.9 ± 57.99 kJ/mol.Tus, 1 was found to have better binding energy than 2 against all the evaluated receptors.Interestingly, no halogen-type interactions were Bioinorganic Chemistry and Applications observed with the bromo group in complex 1.However, the methyl groups and the phosphonium aryl group have contributed mostly to the hydrophobic interaction, while the oxygen atoms play a role in attractive (dipole-dipole) interactions.
Te molecular docking studies of 1 and 2 against human serum transferrin (1D3K) exhibited some occurrences of hydrogen bonds.For complex 1, the osmium complex formed alkyl-π interactions with Cys137(A), Tyr136(A), and Cys331(A) attributed to methyl group of the p-cymene, as shown in Figure 6.Te oxygen within the chelate ring interacted with one of the phenyl rings on the phosphonium moiety through a π-anion interaction.Te phosphonium moiety also formed van der Waals interaction with Gly133(A), Asn325(A), Tr321(A), Tyr317(A), Tyr319(A), and Ala244(A).Te phenyl ring of PPh 4 + cation also formed π-anion interactions with Glu318(A) whilst also playing a role in π-σ and π-alkyl interaction with Ala322(A).For complex 2, there are strong van der Waals force interactions with various amino acids [Lys291(A), Ser189(A), Gly190(A), Tyr185(A), Phe186(A), Lys193(A), Asn183(A), Gly187(A), Gln184(A), Ser180(A), Leu182(A), and His289(A)].A carbon-oxygen contact occurred via the oxygen of the coordinated oxalato ligand interacting with Gly290(A), whereas the methyl group of the cymene ligand exhibits a πalkyl contact with Leu293(A) and His14(A) as illustrated in Figure 7.In addition, one of the aromatic rings of the metalcoordinated PPh 3 exhibited π-π stacking with His14(A).
Te interactions of the complexes were also evaluated against DNA Duplex (1XRW).In complex 1, the osmium complex moiety formed π-anion interaction with the phosphonium moiety through the oxygen, as shown in Figure 8. Te oxalato ligand carbonyl group interacted with DG5(A), via a carbon-hydrogen bond interaction.Te complex also forms van der Waals interactions with DC7(A) and DT6(A).Te phosphonium moiety of 1 also participates by having van der Waals interaction with DA6(B), DT6(A), and DC7(A).Te phenyl rings of PPh4 play a role in π-π stacking with DC4(B) and DG5(A) while also interacting via π-σ interaction with DG5(B).For complex 2, it was observed that the oxygen of the carbonyl played a role in hydrogen bonding with DG5(A), with a ring oxygen having a negativenegative interaction with DC7(A), as shown in Figure 9. Te methyl group formed π-alkyl interaction primarily with DC4(B), whereas the PPh 3 ligand played a role in π-π stacking with DG5(B).Te interaction of complexes 1 and 2 against human serum albumin (1H9Z) was investigated.In complex 1, the Os-O moiety was observed to form an attractive charge interaction with Lys190(A) while simultaneously interacting with the phosphonium moiety via a π-anion interaction, as shown in Figure 10.A hydrogen bond between the oxygen of the carbonyl with Lys190(A) was detected.Te methyl substituent on the cymene ring contributed towards π-alkyl interaction with Leu463(A).Te phosphonium moiety was observed to form a π-cation interaction with Arg197(A), while a π-π T-shaped contact occurred via one of the phenyl rings with His146(A) and a second phenyl ring interacted with Lys190(A) via π-alkyl interaction.Complex 2 exhibits a carbon-hydrogen interaction with His440(A) via the oxalato carbonyl group.In addition, the PPh 3 has a π-sulfur interaction with Cys448, π-π T-shaped stack interaction with Tyr452(A), and π-anion interaction with Asp451(A), as shown in Figure 11.A second phenyl ring participates in π-cation contact involving Arg218(A) and π-alkyl interaction involving Pro447(7).

Assessment of DNA-Binding Activity by UV-Visible
Measurements.UV-Vis absorption information of 1 and 2 are accessible in the supplementary data section, Supplementary Figures S9 and S10.Complex 1 has four observable absorption peaks, in the range of 250 nm to 350 nm.Te signals around 267 nm and 269 nm correspond to n − π * electronic transitions; 276 nm corresponds to a π − π * transition, and 340.25 nm is associated with an MLCT transition of the complex [69].In comparison, complex 2 has only three peaks appearing at 268 nm, 277 nm, and 340.5 nm consistent to n − π * , π − π * , and MLCT transitions, respectively [69].
Te stability test results for 1 and 2 in the DMSO and Tris bufer binary solvent system are reported in the supplementary data section Supplementary Figures S11 and S12.Te data illustrate that both complexes are stable in DMSO and Tris bufer solvent systems, as there were no observable changes over time.Tere are no ligand exchange reactions or precipitation occurring in the solvent system since a change in the ligation of the metal results in observable changes in the electronic spectrum.

Bioinorganic Chemistry and Applications
In vitro experimentations on the possible interaction of 1 and 2 to DNA were carried out by observing changes in the UV-Vis spectra.When DNA interacts with metal fragments, it could result in hypochromic or hyperchromic changes.Hyperchromic efects are due to an electrostatic binding mode with DNA, whereas hypochromic efects are as a result of an intercalative binding mode [70].From Supplementary Figures S13 and S14 in the supplementary data section, both complexes show a signifcant hypochromic efect (decrease in peak intensities).Hypochromic efect is caused by signifcant damage to the DNA double helical structure, which causes the π * orbitals of the ligands in the complex to interact with the π orbitals in the DNA base pairs [71].Tis results in an enhanced π − π * stacking assembly involving DNA base pairs with conjugated planar ring systems of the complex [28].Tus, the resulting coupled π * orbitals become partially flled, efecting a decrease in the possible electron transitions [71].In addition, slight bathochromic changes were detected in the data of both complexes, which has been reported to be due to intercalative bonding of the complex with DNA [28,42].It is worth noting that complexes under study, based on the DNAbinding study, show diverse mechanism of action compared to the standard drug, Oxaliplatin.Research shows that Oxaliplatin bond to DNA covalently by attaching through N7 of the guanine base [13,72].Contrarily, the complexes in this study were able to function as an intercalating agent, inducing conformation changes to the DNA, which would disrupt replication and transcription.

Conclusions
Tis study provides an insight into the identifcation of structural parameters that infuence intermolecular and intramolecular interactions in complexes of this type, which may assist in the design of new potential metallodrugs.Two novel osmium-cymene complexes containing phosphine as well as phosphonium aryl assemblies in both primary and secondary coordination domains have been prepared and structurally characterized utilizing singlecrystal XRD, FT-Raman, FT-IR, UV-Vis, and NMR spectroscopy.From a computational DFT study, HOMO-LUMO orbitals for both complexes have been determined and the relative molecular stability evaluated using FMO analysis.Te chemical reactivity descriptors' values highlight that osmium-oxalato complex with the phosphonium aryl moiety in the second coordination sphere exhibits a higher chemical reactivity and lower kinetic stability.Te Hirshfeld surface analysis discloses that the transposition of the phosphine aryl group from the inner coordination sphere to a phosphonium aryl group in the outer coordination sphere also has a signifcant infuence on the intra-and intermolecular bonding capabilities of the osmium(II)-oxalato moiety.Hirshfeld surface analysis interactions are supported by molecular docking study results which show several interactions between the complexes and selected receptors.Complex 1 shows lower free energy with stable binding energy across the three receptors compared to complex 2. Due to the bulkiness and the charged nature of complex 1, it is more likely to dock in larger pockets and where there are charged species than complex 2. Complex 2 exhibits variations in binding energies which could be because of its neutral state and hydrophobic nature which leads to interaction with amino acids that favour hydrophobic interactions.Te stability of both complexes was evaluated in DMSO and Tris bufer to warrant the use of this solvent system for DNA titrations.From the DNA-binding study, it can be concluded that both 1 and 2 bond to CT-DNA in vitro, possibly applying the intercalation approach of binding.Te behaviour is attributed to efective π − π * stacking connections involving the DNA base pairs and conjugated planar ring system chromophores of the complex.Tese results agreed with in silico docking studies that were executed to develop an understanding of the interactions in 1 and 2 against DNA Duplex (1XRW), as both complexes showed possible π-π interactions with DNA Duplex.

Data Availability
Additional crystallography information is accessible without restrictions on the Cambridge Crystallography Data Centre at https://www.ccdc.cam.ac.ac.uk/data and the deposition numbers for the complexes in this study are CCDC-1909507 for 1 and CCDC-1909506 for 2.

Table 2
contains the crystal data and structural refnement details of complexes 1 and 2.

Table 1 :
Assignment of vibrational (Infrared and Raman) data of complexes 1 and 2.

Table 2 :
Crystal data and structural refnement of 1 and 2.

8
Bioinorganic Chemistry and Applications packing diagrams of complexes 1 and 2 (see Supplementary FiguresS7 and S8in the supplementary data section).

Table 4 :
Global chemical reactivity indices for 1 and 2.

Table 5 :
Binding energies of complexes 1 and 2 with receptors showing E-value (kJ/mol).