Ru(II)-p-Cymene Complexes of Furoylthiourea Ligands for Anticancer Applications against Breast Cancer Cells.

{"full_text": " pubs.acs.org/IC Article\n\n\n\n Ru(II)\u2011p\u2011Cymene Complexes of Furoylthiourea Ligands for\n Anticancer Applications against Breast Cancer Cells\n Dorothy Priyanka Dorairaj, Jebiti Haribabu, Mahendiran Dharmasivam, Rahime Eshaghi Malekshah,\n Mohamed Kasim Mohamed Subarkhan, Cesar Echeverria, and Ramasamy Karvembu*\n Cite This: Inorg. Chem. 2023, 62, 11761\u221211774 Read Online\n\n\n ACCESS Metrics & More Article Recommendations *\n s\u0131 Supporting Information\nSee https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.\n Downloaded via MOSCOW STATE UNIV on May 12, 2026 at 11:22:45 (UTC).\n\n\n\n\n ABSTRACT: Half-sandwich Ru(II) complexes containing nitro-substituted furoylthiourea ligands, bearing the general formula\n [(\u03b76-p-cymene)RuCl2(L)] (1\u22126) and [(\u03b76-p-cymene)RuCl(L)(PPh3)]+ (7\u2212-12), have been synthesized and characterized. In\n contrast to the spectroscopic data which revealed monodentate coordination of the ligands to the Ru(II) ion via a \u201cS\u201d atom, single\n crystal X-ray structures revealed an unusual bidentate N, S coordination with the metal center forming a four-membered ring.\n Interaction studies by absorption, emission, and viscosity measurements revealed intercalation of the Ru(II) complexes with calf\n thymus (CT) DNA. The complexes showed good interactions with bovine serum albumin (BSA) as well. Further, their cytotoxicity\n was explored exclusively against breast cancer cells, namely, MCF-7, T47-D, and MDA-MB-231, wherein all of the complexes were\n found to display more pronounced activity than their ligand counterparts. Complexes 7\u221212 bearing triphenylphosphine displayed\n significant cytotoxicity, among which complex 12 showed IC50 values of 0.6 \u00b1 0.9, 0.1 \u00b1 0.8, and 0.1 \u00b1 0.2 \u03bcM against MCF-7, T47-\n D, and MDA-MB-231 cell lines, respectively. The most active complexes were tested for their mode of cell death through staining\n assays, which confirmed apoptosis. The upregulation of apoptotic inducing and downregulation of apoptotic suppressing proteins as\n inferred from the western blot analysis also corroborated the apoptotic mode of cell death. The active complexes effectively\n generated reactive oxygen species (ROS) in MDA-MB-231 cells as analyzed from the 2\u2032,7\u2032-dichlorofluorescein diacetate (DCFH-\n DA) staining. Finally, in vivo studies of the highly active complexes (6 and 12) were performed on the mice model. Histological\n analyses revealed that treatment with these complexes at high doses of up to 8 mg/kg did not induce any visible damage to the tested\n organs.\n\n\n \u25a0 INTRODUCTION\n The prevalence of cancer has increased substantially, claiming\n metals exhibit almost the same kinetic rate in the order of 10\u22123\u2212\n 10\u22122 s\u22121.4 Organometallic Ru compounds possess remarkable\n millions of lives and impacting many across the globe. Based on activity against several cancer cell lines. Clarke\u2019s group, in 1980,\n the American Cancer Society statistics, breast cancer has synthesized the first Ru-based classical Werner-type complex\n [RuCl3(NH3)3], which was tested for its anticancer activity.5\n emerged as the most frequently reported one and also the\n Thereafter, some polypyridyl complexes of the type cis-\n second highest cause of mortality in women.1 Despite cisplatin\u2019s\n [RuCl2(bpy)2] and mer-[RuCl3(tpy)] were reported by\n enormous success as an anticancer drug, its limitations and side\n Novakova et al., which exhibited good cytotoxicity on leukemia\n effects have forced the scientific community to focus on alternate\n and colon cancer cells. 6 Importantly, trans-[imi-H]-\n metallodrugs with minimal side effects and enhanced anticancer\n [RuCl4(imi)2] and trans-[ind-H][RuCl4(ind)2] reported by\n efficacy.2 In the search for developing anticancer agents\n containing metals other than Pt, Ru complexes have emerged\n as the most promising ones in recent years. The ligand exchange Received: March 7, 2023\n kinetics for metal complexes in an aqueous medium is one of the Published: July 17, 2023\n crucial factors influencing anticancer activity.3 Coordination\n compounds having slow ligand exchange rates, similar to the cell\n cycle processes, appear to be highly effective in targeting cancer\n cells. This is especially obvious in Pt and Ru complexes. These\n\n \u00a9 2023 American Chemical Society https://doi.org/10.1021/acs.inorgchem.3c00757\n 11761 Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n\n\n\nFigure 1. Examples of some well-known Ru(II)/Ru(III) potential anticancer drugs.\n\nScheme 1. Synthesis of Ligands (L1\u2212L6)\n\n\n\n\nKeppler\u2019s group exhibited remarkable activity on colorectal presence of the hydrophobic arene, labile chloride, lipophilic\ncancer cells, which were considered as a breakthrough in the area PPh3, and chelating ligands contribute to their overall anticancer\nof Ru complexes as anticancer agents. Further, substantial activity. The ligands present in the Ru\u2212arene system are\ninsights into the anticancer potential of NAMI-A have been important in controlling anticancer efficacy. One such vital\nreported by Alessio\u2019s, Mestroni\u2019s, and Sava\u2019s groups.7\u221212 ligand system is aroylthiourea, which has been found to exhibit\nRuthenium-based compounds such as NAMI-A, KP1019, an array of biological properties.24 They are air- and moisture-\nKP1339, and TLD1433 had initially entered clinical trials. stable and have the ability to exhibit versatile coordination\nOwing to less activity and solubility limitations, NAMI-A and modes with metals due to the presence of O, N, and S donor\nKP1019 were dropped out of the trials. Currently, TLD1433 has atoms.25 Furans are well-known heterocyclic compounds\nentered phase IIa human clinical trials (ClinicalTrials.gov, possessing a wide range of therapeutic properties and constitute\nidentifier NCT03053635) and has shown to be effective against an important part of natural products such as furanoflavinoids,\nhuman non-musculoinvasive bladder cancer.13\u221215 Some of the furanolactones, furanocoumarins, and some terpenoids. In the\nwell-known Ru anticancer drugs are illustrated in Figure 1. present work, we report twelve Ru(II)-arene-based furoylth-\n The anticancer activity of Ru\u2212arene complexes was first iourea complexes, their interactions with biomolecules, and their\nreported by Tocher\u2019s group in 1992, wherein the activity of well- cytotoxic properties exclusively against breast cancer cells.\nknown [Ru(\u03b76-benzene)(metronidazole)Cl2] was described.16\nSubsequently, Sadler\u2019s and Dyson\u2019s groups investigated the\nactivity of some Ru\u2212arene complexes.11,17\u221219 Recently, Ru-\n \u25a0 RESULTS AND DISCUSSION\n Synthesis of Ligands and Complexes. Refluxing furoyl\n(II)\u2212arene complexes containing PPh3 have been developed, chloride and potassium thiocyanate in acetone for 1 h, followed\nwhich displayed superior activity against cancer cells.20\u221223 The by the addition of the corresponding aniline (2-nitroaniline, 4-\n 11762 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\nScheme 2. Synthesis of Complexes (1\u22126)\n\n\n\n\nScheme 3. Synthesis of Complexes Containing PPh3 (7\u221212)\n\n\n\n\nmethyl-2-nitroaniline, 4-methoxy-2-nitroaniline, 5-chloro-2-ni- carbonyl- and thiocarbonyl-attached N\u2212H protons, respec-\ntroaniline, 5-chloro-4-methyl-2-nitroaniline or 4,5-dimethyl-2- tively. Aromatic protons of phenyl and furoyl groups resonated\nnitroaniline) at room temperature (Scheme 1) resulted in the at 6.62\u22128.67 ppm. In the 1H NMR spectra of complexes, there\nformation of ligands L1\u2212L6. Among them, ligand L1 was was an insignificant change in the signal of the carbonyl-attached\nalready reported.26 The treatment of ligands with [RuCl2(\u03b76-p- N\u2212H proton, whereas the thiocarbonyl-attached N\u2212H proton\ncymene)]2 and [RuCl2(\u03b76-p-cymene)(PPh3)] in 2:1 and 1:1 appeared to be more deshielded compared to that of the ligands,\nratios in toluene for 12 and 24 h, respectively, yielded the which indicated the coordination of S to the Ru(II) ion. Two\ncorresponding complexes (Schemes 2 and 3). sets of doublets at 5.74\u22125.19 and 5.45\u22124.98 ppm, a septet at\n Spectroscopic Characterization. The ligand and complex 3.04\u22122.61 ppm, a singlet at 2.50\u22121.89 ppm, and a doublet at\nformation were confirmed by ultraviolet\u2212visible (UV\u2212vis), 1.62\u22121.08 ppm validated the presence of p-cymene in the\nFourier transform infrared (FT-IR), nuclear magnetic resonance complexes. The 13C NMR spectra of ligands displayed two\n(NMR), and electrospray ionization-mass spectrometry (ESI\u2212 signals in the ranges of 179.0\u2212181.1 and 155.9\u2212158.1 ppm, that\nMS) spectroscopic techniques. The bands appearing in the UV\u2212 belonged to thiocarbonyl and carbonyl carbons, respectively,\nvis spectra of the ligands at 261\u2212279 and 276\u2212296 nm were\n while the aromatic carbons resonated at 109.1\u2212156.2 ppm. The\ncharacteristic of \u03c0 \u2192 \u03c0* and n\u2192 \u03c0* transitions, respectively.27\n new signals appearing at 109.3\u221293.5, 100.5\u221288.9, 91.0\u221281.2,\nThe additional bands seen at 347\u2212365 and 425\u2212459 nm in the\n 89.7\u221279.7, 30.9\u221230.3, 22.4\u221221.2, and 19.7\u221217.2 ppm in the\nspectra of complexes were due to MLCT and d \u2192 d transitions,\nrespectively. FT-IR spectra of the ligands exhibited four sharp spectra of complexes verified the existence of p-cymene.29 In\npeaks at 3431\u22123459, 3081\u22123189, 1679\u22121691, and 1248\u22121269 addition, 2D NMR techniques such as 1H\u221213C HSQC and\ncm\u22121 that belonged to the amide N\u2212H, thiourea N\u2212H, C\ufffdO, NOESY for representative ligand (L1) and complex (1) were\nand C\ufffdS stretching vibrations, respectively. In the spectra of employed to better understand the proton and carbon\ncomplexes, a noticeable shift (from 1248\u22121269 to 1171\u22121186 interactions. The appearance of a singlet in the 31P NMR\ncm\u22121) in the stretching frequency of C\ufffdS was seen, whereas spectra at 29.43\u221231.87 ppm proved the coordination of PPh3 to\nalmost no change in the C\ufffdO stretching frequency was Ru, while a septet found in the region \u2212146.5 to \u2212135.5 ppm in\nobserved, indicating the coordination of S to the Ru(II) ion. The the case of complexes 7\u221212 was due to PF6.30 31P NMR\nstretching frequencies observed at 710\u2212721, 1076\u22121092, and chemical shift values for complexes 7\u221212 are listed in Table S1.\n1437\u22121452 cm\u22121 confirmed the presence of PPh3 in the The NMR spectra for L1\u2212L6 and 1-\u221212 are depicted in Figures\ncomplexes.28 FT-IR spectra of the complexes are depicted in S2\u2212S43. The 2D NMR spectra for L1 and complex 1 are shown\nFigure S1. 1H NMR spectra of the ligands showed two singlets at in Figures S44\u2212S47. ESI\u2212MS spectra of complexes 1\u22126\n13.33\u221212.54 and 11.69\u22129.30 ppm, which were due to the displayed an m/z value corresponding to [M \u2212 HCl \u2212 Cl\u2212]+,\n 11763 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n\n\n\nFigure 2. X-ray structures of L2, L3, 1, 3, 4, and 6.\n\nwhile complexes 7\u221212 displayed a molecular ion peak due to [M while crystals of the complexes were obtained by evaporation of\n\u2212 PF6\u2212]+ (Figures S48\u2212S59). their DMF-dichloromethane (2:98) solutions. In contrast to the\n Solid-State Structures. Single crystals of the ligands were results obtained from spectroscopy, an unusual bidentate\nobtained by slow evaporation of their acetonitrile solutions, coordination of the furoylthiourea ligands through N, S atoms\n 11764 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\nwith the Ru(II) ion was seen in complexes 1 and 3, whereas the decrease in viscosity as covalent binders bind to the sides of the\nmonodentate coordination involving only S was observed in DNA helix that results in a breakage/kinking of the helical\ncomplexes 4 and 6. Such N, S coordination of aroylthiourea strand. The changes in the viscosity of CT DNA upon the\nligands with Ru(II)-arene in the solid state has already been incremental addition of Ru(II) complexes are depicted in Figure\nreported by Cunha, Parveen, Obradovic\u0301, and Swaminathan et S64. EB was used as a positive control while [Co(NH3)6]Cl3 was\nal.21,31\u221233 Figure 2 illustrates the thermal ellipsoid plots for the chosen as a negative control.41 An increase in viscosity was seen\nligands and complexes, as well as the atomic labeling schemes. with the addition of complexes 1\u221212 to CT DNA, suggesting\nCrystal data and selected bond lengths and angles for the ligands intercalation. However, the effect of intercalation was less when\nand complexes are summarized in Tables S2\u2212S5. Ligands L2 compared to EB, and greater when compared to [Co(NH3)6]-\nand L3 crystallized in a monoclinic fashion with the P21/c space Cl3.\ngroup. Complexes 1 and 3 adopted a pseudo-tetrahedral piano- Complex\u2212BSA Interaction. To understand the complex\u2212\nstool geometry encompassing N, S-coordinated aroylthiourea, protein interaction, a binding study with BSA was performed. An\np-cymene, and chlorido ligands. In the complexes, an intra- enhancement in absorbance was seen upon the addition of the\nmolecular hydrogen bonding was observed between the complexes at 280 nm, which indicated the static nature of\nthioamide N\u2212H and carbonyl oxygen. Also, an intramolecular binding between BSA and complexes (Figure S65).42 Further,\nhydrogen bonding between amide N\u2212H, carbonyl oxygen, and the interaction between the complexes and BSA was studied\nNO2 was observed in the crystal pattern of complexes 1, 3, 4, and through fluorescence spectroscopy. The fluorescence spectra\n6. In addition, a hydrogen-bonding interaction was observed were recorded in the range of 290\u2212500 nm upon excitation at\nbetween the amide N\u2212H and one of the chlorido ligands. The 280 nm. An incremental addition of the Ru(II) complexes (0\u2212\nRu\u2212C bond lengths were in the region 2.152\u22122.233 \u00c5, while the 18 \u03bcM) to BSA (1 \u03bcM) resulted in the quenching of\nRu\u2212S, Ru\u2212Cl, and Ru\u2212N bond distances were in the ranges of fluorescence at 345 nm, which was accompanied by a minor\n2.326\u22122.360, 2.378\u22122.396, and 2.254\u22122.274 \u00c5, respectively, hypsochromic shift (Figure S66). This quenching was due to the\nwhich were in line with the literature values.34,35 active sites of the protein being buried inside a hydrophobic\n Interaction with Biomolecules. Complex\u2212CT DNA pocket.43 In addition, the inner filter effect of the Ru(II)\nInteraction. Figure S60 depicts the absorption spectra of the complexes was corrected using the equation, Fcorr = Fobs \u00d7\nRu(II) complexes with and without CT DNA. The addition of e(Aex+Aem)/2, where Fcorr is the corrected fluorescence intensity, Fobs\nCT DNA to the Ru(II) complexes led to a decrease in is the observed intensity, and Aex and Aem are the absorbances of\nabsorbance, creating a hypochromic effect accompanied by a the compounds at excitation and emission wavelengths,\nsmall red shift (2\u22125 nm). Compounds exhibiting noncovalent respectively.44 From the Stern\u2212Volmer equation, the plot of\ninteractions with DNA generally display hypochromism owing F\u00b0/F versus [Q] was deduced as shown in Figure S67. By\nto the stacking interactions between the aromatic chromophore employing the Scatchard equation, the plot of log [(Fo\u2212F)/F]\nand the DNA base pairs. The extent of binding was understood versus log[Q] was deduced (Figure S68), from which the\nfrom the intrinsic binding constant (Kb) values deduced from number of binding sites (n) and binding constant (Kb) values\nthe Wolfe\u2212Shimmer equation.36 The plots of [DNA]/(\u03b5a \u2212 \u03b5f) were obtained. The values of Kb, Kq, and n are listed in Table S7.\nversus [DNA] are shown in Figure S61, and the Kb values are Docking of the Ru(II) Complexes with Biomolecules (DNA\nlisted in Table S6. The complexes holding PPh3 (7\u221212) and BSA). The interactions of DNA and BSA with the Ru(II)\u2212\ndisplayed better binding than complexes 1\u22126. This could be arene complexes were further visualized by molecular docking\nlinked to the presence of phenyl rings (of PPh3), resulting in studies using Molegro software with the chosen targets DNA\nenhanced planarity and \u03c0\u2212\u03c0 aromatic stacking interactions with (PDB ID: 1Z3F) and BSA (PDB ID: 3V03). From the docked\nthe DNA base pairs.37 Among the different substituents, the images of complexes (Figure S69), we could infer that the\ncomplexes bearing an electron-donating substituent (methyl or complexes, by and large, got inserted between the DNA helical\nmethoxy) exhibited the highest binding ability with DNA, which strands, confirming intercalation on par with the results obtained\nagreed with the literature.38 from absorption and emission spectral studies. Complexes 7\u221212\n As the Ru(II) complexes remained nonfluorescent, their containing PPh3 exhibited better docking energy scores than\nbinding with CT DNA could not be directly predicted from complexes 1\u22126, which could be due to the enhanced stacking\nemission studies. Hence, competitive binding with ethidium interactions between the phenyl rings and DNA base pairs.\nbromide (EB) was performed. Figure S62 depicts the emission Based on the docking energy scores (Table S8), the complexes\nspectra of the EB\u2212DNA mixture in the absence and presence of showed good interaction with BSA as well (Figure S70).\nthe complexes wherein a decrease in the fluorescence intensity at Lipophilicity Measurements. Lipophilicity is a key pharma-\n610 nm was observed upon the addition of Ru(II) complexes codynamic and pharmacokinetic characteristic that impacts\n(0\u221250 \u03bcM) to EB\u2212DNA. The slope of the plot of Fo/F versus drug absorption, metabolism, distribution, excretion, and\n[Q] gave Kq (Figure S63). The extent of quenching (Kq) was toxicity (ADMET). The partition coefficient, Log P, which can\ncalculated from the Stern\u2212Volmer equation.39 The Kq and Kapp be defined as the propensity of a neutral (uncharged) compound\nvalues are listed in Table S6. Viscosity studies were carried out to to dissolve in an immiscible biphasic system of lipid (fats, oils, or\nunderstand the nature of binding interactions. This hydro- organic solvents) and water or in simple terms, the ratio of\ndynamic experiment is sensitive to changes in the DNA chain partitioning between octanol and water, can be used to describe\nlength, and binding interactions such as covalent and non- a drug\u2019s lipophilicity.45 A negative Log P value indicates that the\ncovalent can be distinguished based on the changes in viscosity compound has a higher affinity for the aqueous phase (it is more\nlevels.40 Intercalators generally tend to slide in between the hydrophilic); when Log P = 0, the compound is equally\nDNA base pairs, leading to elongation of the helical strand, partitioned between the lipid and aqueous phases; and a\nresulting in increased viscosity. In the case of noncovalent major positive Log P value indicates that the compound has a higher\nand minor groove binding, negligible changes in viscosity are concentration in the lipid phase (it is more lipophilic). Log P = 1\nseen. In contrast, the covalent binding interactions result in a indicates a 10:1 partitioning in the organic\u2212aqueous phases.\n 11765 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\nTable 1. IC50 (\u03bcM) Values and Selectivity Index (S.I) of the Ru(II) Complexes and Cisplatin after 72 h Incubation at 37 \u00b0C\n IC50 (\u03bcM) S.I\n compound MCF-7 T47-D MDA-MB-231 MCF-10a MCF-7 T47-D MDA-MB-231\n L1 39.2 \u00b1 0.5 36.4 \u00b1 0.6 36.8 \u00b1 0.8 >50 1.27 1.37 1.35\n L2 29.5 \u00b1 0.1 26.8 \u00b1 0.8 27.1 \u00b1 0.5 >50 1.69 1.86 1.84\n L3 27.6 \u00b1 0.9 23.6 \u00b1 0.5 24.4 \u00b1 0.5 >50 1.81 2.11 2.04\n L4 43.0 \u00b1 0.5 39.7 \u00b1 1.0 40.2 \u00b1 0.7 >50 1.16 1.25 1.24\n L5 34.4 \u00b1 0.2 30.1 \u00b1 0.8 31.2 \u00b1 0.6 >50 1.45 1.66 1.60\n L6 25.3 \u00b1 0.1 23.5 \u00b1 0.3 23.9 \u00b1 1.0 >50 1.97 2.12 2.09\n 1 19.3 \u00b1 0.6 16.2 \u00b1 0.4 16.7 \u00b1 0.4 >50 2.59 3.08 2.99\n 2 13.6 \u00b1 0.8 11.4 \u00b1 0.2 11.4 \u00b1 1.2 >50 3.67 4.38 4.38\n 3 12.5 \u00b1 0.5 10.0 \u00b1 0.8 10.3 \u00b1 0.3 >50 4.00 5.00 4.85\n 4 28.6 \u00b1 0.3 24.2 \u00b1 0.9 21.6 \u00b1 0.4 >50 1.74 2.06 2.31\n 5 14.6 \u00b1 0.3 12.1 \u00b1 0.2 12.9 \u00b1 0.9 >50 3.42 4.13 3.87\n 6 9.8 \u00b1 0.2 8.5 \u00b1 0.4 8.6 \u00b1 0.6 >50 5.10 5.88 5.81\n 7 2.4 \u00b1 0.1 1.2 \u00b1 0.9 1.2 \u00b1 0.4 >50 20.83 41.66 41.66\n 8 1.5 \u00b1 0.8 0.4 \u00b1 0.6 0.5 \u00b1 0.5 >50 33.33 125.00 100.00\n 9 1.1 \u00b1 0.6 0.1 \u00b1 0.8 0.1 \u00b1 0.2 >50 45.45 500.00 500.00\n 10 3.9 \u00b1 0.5 2.8 \u00b1 0.7 2.8 \u00b1 1.2 >50 12.82 17.85 17.85\n 11 1.6 \u00b1 0.7 0.9 \u00b1 0.5 0.8 \u00b1 0.3 >50 31.25 55.55 62.5\n 12 0.6 \u00b1 0.9 0.1 \u00b1 1.0 0.1 \u00b1 0.2 >50 83.33 500.00 500.00\n cisplatin 19.4 \u00b1 0.5 15.8 \u00b1 0.1 7.1 \u00b1 0.8 28.7 \u00b1 0.8 1.47 1.81 4.04\n\nHigh lipophilicity (Log P > 5) often contributes to high S75). The isobestic points observed around 325 and 350 nm\nmetabolic turnover, low solubility, and poor oral absorption, were chosen for monitoring the kinetics for half an hour. The\nthereby hampering the drug\u2019s bioavailability. Compounds with a plots of absorbance versus time are depicted in Figure S76. By\nLog P index higher than 1 or less than 4 are considered to have performing a nonlinear regression analysis using Origin 8.5 pro\ngood physicochemical properties suitable for oral consumption. software, it was understood that the complexes (1\u221212) obeyed a\nIn the present work, the Log P values of complexes (1\u221212) were pseudo-first-order kinetics. The corresponding rate constants\ndetermined by the shake flask procedure. Figure S71 depicts the (Kobs) and half-life time (t1/2) were calculated from the formulae\nlipophilicity diagram of the Ru(II) complexes, while Table S9 k = 1/t and t1/2 = 0.693/k. From the calculated values (Table\ndepicts their Log P values. All of the complexes showed a Log P S10), it is inferred that complexes 6 and 12 displayed the highest\nvalue in the range of 1.2\u22123.1. Compared to the dichlorido rate constants and shortest half-life. The relative Kobs and t1/2\ncomplexes, complexes 7\u221212 bearing PPh3 had a slightly greater order of the complexes was 6 > 3 > 5 > 1 > 4 (complexes bearing\nlipophilicity index. two chlorido ligands) and 12 > 9 > 8 > 11 > 7 > 10 (complexes\n Solution Behavior of the Complexes. The hydrolysis process bearing one PPh3 and one chlorido ligands). The inclusion of\nis widely thought to be a crucial activation step inside the cell electron-donating group(s) in the phenyl rings might result in an\nbefore the drug reaches the intracellular DNA target. It has been increase in electron density around the Ru(II) ion, which might\nwell documented in the literature that for the [Ru(\u03b76- have increased the lability of chlorido and PPh3, thereby\narene)Cl(en)]+ (arene = p-cymene, benzene or dihydroan- facilitating hydrolysis. Hence, the complexes with electron-\nthracene) systems, the hydrolysis of Ru\u2212Cl bond to form donating group(s) such as methyl and methoxy exhibited a\n[Ru(\u03b76-arene)(H2O)(en)]+ aqua species may activate the higher rate of hydrolysis than the ones with electron-with-\ncomplex for DNA binding.46 In the present case, the stability drawing group(s).\nassessment of the complexes in 1:99 (v/v) DMSO\u2212water In Vitro Cytotoxicity of the Ru(II) Complexes. Based on the\nmedium through absorption spectroscopy for 72 h ensured that binding interactions with DNA and BSA, we further explored the\nall of the complexes were stable with no alteration or formation cytotoxicity of Ru(II) complexes against breast cancer cells,\nof new bands (Figure S72). Then, the hydrolysis of active namely, MCF-7, MDA-MB-231, and T47-D. To ensure their\ncomplexes 6 and 12 in the presence of chloride ions (4 mM and selectivity, the activity on a normal MCF-10a cell line was\n100 mM NaCl) was studied. Almost no suppression of evaluated. From the MTT assay results, we observed that,\nhydrolysis was seen at a 4 mM concentration of NaCl, but the surprisingly, both the ligands and their Ru(II) complexes\nextent of hydrolysis was considerably suppressed at 100 mM displayed remarkable cytotoxicity against the three breast cancer\nNaCl (Figure S73). These results positively indicated that the cell lines in a dose-dependent manner. The IC50 values of ligands\ncomplexes were capable of surviving in bloodstream conditions ranged 27.65\u221243.08, 23.52\u221239.71, and 23.98\u221240.82 \u03bcM\nwhere a high concentration of chloride ions is present.46 against MCF-7, MDA-MB-231, and T47-D cancer cells,\n To evaluate the kinetics of hydrolysis, time-dependent respectively, after 72 h of incubation. The complexes portrayed\nabsorption spectra for the complexes were recorded in 100 much higher cytotoxicity than their ligands, highlighting the role\nmM NaClO4 solution, from which the plot of \u0394A versus K was of Ru(II) ion and arene moiety. Compared to complexes 1\u22126\nobtained (Figure S74), where \u0394A = \u0394t \u2212 \u0394o (\u0394t = absorbance at which contain two labile chlorido ligands, complexes 7\u221212\ntime t ; \u0394o = absorbance at t = 0.2 min). The presence of containing one chlorido and one PPh3 exhibited greater\nisobestic points as seen from the spectra implied that an cytotoxicity portraying IC50 values of 0.62\u22123.98, 0.15\u22122.81,\nequilibrium existed between the aqua and chlorido/PPh3 and 0.17\u22122.88 \u03bcM against MCF-7, MDA-MB-231, and T47-D\ncomplexes, and this equilibrium was attained in 1 h (Figure cell lines, respectively. It was concluded that the presence of\n 11766 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\nTable 2. Anticancer Activity of the Reported Ru(II)\u2212Arene Aroylthiourea Complexes against Breast Cancer Cells\n literature MCF-7 MDA-MB-231 MCF-10a\n Rohini et al.48 15.2\u221248.0 \u00b1 0.5 \u03bcM\n Rohini et al.34 53.0 and 98.1 \u00b1 0.5 \u03bcM\n Cunha et al.22 0.2\u22120.7 \u00b1 0.5 \u03bcM 3.14\u22126.67 \u00b1 0.95 \u03bcM\n Oliveira et al.23 0.1\u22120.2 \u00b1 0.5 \u03bcM 0.5\u22120.7 \u00b1 0.5 \u03bcM\n Barolli et al.49 8.9 and 12.2 \u03bcM\n Becceneri et al.50 9.3\u221221.9 \u00b1 0.5 \u03bcM 7.9\u221229.4 \u00b1 0.5 \u03bcM\n Colina-Vegas et al.51 9.3\u221231.0 \u00b1 0.5 \u03bcM\n Jeyalakshmi et al.52 52.3 \u2192 500 \u03bcM\n Jeyalakshmi et al.53 151.2\u2212162.9 \u00b1 0.5 \u03bcM\n Becceneri et al.54 33.4 \u03bcM\n this work 0.6\u22123.9 \u03bcM 0.1\u22122.8 \u03bcM >50 \u03bcM\n\nlipophilic PPh3 and electron-donating substituents such as\nmethyl and methoxy enhanced the activity of complexes.47\nComplexes 6 and 12 bearing two methyl groups exhibited the\nhighest anticancer activity against MDA-MB-231 cancer cell\nline. Notably, the cytotoxicity of the complexes was much\ngreater than cisplatin, and positively, they exhibited less toxicity\non the normal MCF-10a cell line. The IC50 values of the ligands\nand complexes along with their selectivity index are shown in\nTable 1. The selectivity index (S.I) was calculated by using the\nformula IC50 of MCF-10a/IC50 of the cancer cells. Compared to Figure 4. MDA-MB-231 cancer cells stained with DAPI after being\nthe literature results, the synthesized Ru(II)\u2212arene complexes treated with the active complexes at a concentration of 25 \u03bcM.\ndisplayed superior activity toward the breast cancer cells (Table\n2).\n Validation of Apoptosis. Based on the encouraging results fluorescence, blue fluorescence was seen in MDA-MB-231 cells\nobtained from the MTT assay, we investigated the mode of cell upon treatment with complexes 2, 3, and 6 (20 \u03bcM), indicating\ndeath in the MDA-MB-231 cancer cell line upon treatment with that these complexes were capable of inducing apoptosis. On the\ncomplexes 2, 3, 6, 8, 9, 11, and 12 at 20 \u03bcM concentration. From other hand, bright fluorescent fetches in MDA-MB-231 cells\nthe color transformation visualized in AO/EB staining and treated with complexes 8, 9, 11, and 12 (20 \u03bcM) were observed,\nnuclear fragmentation pattern observed in DAPI staining, the indicating that PPh3 complexes were able to promote apoptosis\nmode of cell death could be predicted. In AO/EB staining, live in a much better way than the former ones.\ncells emit green fluorescence while early apoptotic cells display Intracellular ROS Generation. Mitochondria, \u201cpower-\nyellow fluorescence; late apoptotic cells exhibit orange house of the cells\u201d, is the primary site for intracellular reactive\nfluorescence, and necrotic cells give out red fluorescence.55 As oxygen species (ROS) production. Although optimum levels of\nshown in Figure 3, yellow fluorescence exhibited by complexes ROS can be beneficial, excessive accumulation can promote\n cancer due to oxidative stress and cellular dysfunction in the\n cell.57 A characteristic trait that distinguishes a cancer cell from a\n normal cell is its ability to generate excess ROS and its increased\n dependence on an antioxidant defense system.58 Here, the ROS\n levels were detected using the DCFH-DA staining assay. Upon\n entering the cells, DCFH-DA undergoes deacetylation to form\n DCFH which is nonfluorescent in nature. Further, upon\n oxidation by ROS, dichlorodihydrofluorescein (DCFH) gets\n converted to dichlorofluorescein (DCF), which emits bright\n green fluorescence at 520 nm upon excitation at 495 nm.59 The\nFigure 3. MDA-MB-231 cancer cells stained with AO/EB after being images depict ROS generation in MDA-MB-231 cells on\ntreated with the active complexes at a concentration of 25 \u03bcM. treatment with active complexes 2, 3, 6, 8, 9, 11, and 12 at 20\n \u03bcM concentration (Figure 5). No fluorescence was seen in the\n control cells, but a considerable extent of green fluorescence was\n2, 3, and 6 indicated that these complexes induced early emitted by MDA-MB-231 cells treated with complexes 2, 3, and\napoptosis in MDA-MB-231 cells. Complexes 8, 9, 11, and 12 6. However, intense green fluorescence fetches were seen with\nbearing PPh3 emitted bright orange fluorescence, which revealed complexes 8, 9, 11, and 12, indicating that PPh3 complexes had a\nthat these complexes induced late apoptosis in the MDA-MB- greater ability to generate ROS in the cancer cell line, thereby\n231 cancer cell line. To visualize the nuclear alterations taking promoting apoptosis.60 The enhanced ability of PPh3-bearing\nplace in MDA-MB-231 cells, DAPI staining was employed. complexes (8, 9, 11, and 12) to generate excess ROS could be\nDAPI is a cell-permeable, fluorescent dye that predominantly attributed to the charge of these complexes and mitochondria.\nbinds to the adenine-thymine regions of DNA. Upon detection Since this organelle is negatively charged, positively charged\nof apoptosis, the dye emits a bright blue fluorescence at 457 cations could easily permeate through the mitochondrial\nnm.56 From the confocal microscopic images (Figure 4), we membrane, leading to better drug absorption.61 Since complexes\ncould see that compared to the control cells that exhibited no 7\u221212 contain an overall positive charge in contrast to complexes\n 11767 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n Table 3. Percentage of Cells at the G0/G1, S and G2/M\n Phases\n compound G0/G1 S G2/M\n control 59.78 \u00b1 0.76 15.84 \u00b1 0.29 19.91 \u00b1 0.31\n 6 47.51 \u00b1 1.05 12.18 \u00b1 0.48 17.79 \u00b1 0.92\n 12 40.95 \u00b1 0.11 8.71 \u00b1 0.36 10.83 \u00b1 1.08\n\n\n caspase-3 and caspase-9 proteins and a downregulation of Bcl-\nFigure 5. MDA-MB-231 cells stained with DCFH-DA upon treatment 2 and Bax proteins in the cancer cell line were observed. The\nwith the active complexes at 25 \u03bcM. expression of these apoptotic markers inferred that both the\n active complexes (6 and 12) promoted apoptosis.\n1\u22126 which are neutral, they could permeate easily across the In Vivo Study. As complexes 6 and 12 emerged as the most\nmitochondrial membrane.62 Parallelly, 2\u2032,7\u2032-dichlorodihydro- active ones as inferred from in vitro experiments, their in vivo\nfluorescein diacetate (H2DCFH-DA) is said to be negatively potential in the healthy ICR models was analyzed. The mice (n =\ncharged due to the presence of two acetyl units (CH3COO\u2212), 8, four females and four males in each group) were intra-\nwhich could make these complexes interact strongly with this peritoneally injected with complexes 6 or 12 (2, 4, 6, 8, and 16\ndye, leading to an excess generation of ROS in MDA-MB-231 mg/kg in DMSO) 5 times on alternate days. The animal models\ncancer cells. were also administered with controls such as DMSO and saline\n Flow Cytometry Analysis. Flow cytometry analysis for the for a comparative study. Upon visualization of the results, it was\nmost active complexes (6 and 12) on MDA-MB-231 cells was inferred that all of the mice treated with cisplatin (4 mg/kg) did\ncarried out to analyze the cell count at each stage of the cell cycle. not survive, whereas the mice administered with complexes 6\nUpon treatment with different concentrations of complexes 6 and 12 with dosages of 8 and 16 mg/kg exhibited survival rates\nand 12 (10, 25, and 50 \u03bcM), the corresponding change in the of 100 and 80%, and 100 and 74%, respectively. Further,\ncell count at the G0/G1, S, and G2/M phases was seen after 48 h hematoxylin and eosin staining analyses were performed for\n(Figure 6a). The control cells showed a percentage count of complexes 6 (8 mg/kg), 12 (8 mg/kg), and cisplatin (4 mg/kg)\n59.78, 15.84, and 19.91 in the G0/G1, S, and G2/M phases, to analyze the morphological alterations in kidneys, lungs,\nrespectively. Complex 6 displayed 47.51 (G0/G1), 12.18 (S), spleen, liver, and heart. Figure 8 picturizes the microscopic\nand 17.79% (G2/M), while complex 12 exhibited a count of images of the organ tissues, which revealed that the organs,\n40.95 (G0/G1), 8.71 (S), and 10.78% (G2/M). Based on the especially lungs, heart, and kidneys administered with complexes\nchange in the cell count at the three stages of the cell cycle, it was 6 and 12, displayed the normal presence of the glomerular\nconcluded that both the complexes induced the cell cycle arrest capsules, cytoplasm, alveolar sacs, cardiac muscle fibers, and\nin MDA- MB-231 cells at the G0/G1 phase. The percentage of tubular epithelial cells. On the contrary, photomicrographs of\ncells at the three phases is listed in Table 3. the organs treated with cisplatin showed severe inflammation\n Intrinsic Mediated Apoptosis Pathway. Lastly, western and edema of the kidneys, degeneration of the distal and\nblotting was performed to understand the occurrence of proximal epithelial cells, accumulation of vesicles in the\napoptosis in MDA-MB-231 cells by measuring the levels of cytoplasm, and damage to the alveolar sacs. From these\napoptotic markers such as caspase-3, caspase-9, Bcl-2, and Bax. histological images, it is evident that the administration of\nThe Bcl-2 protein, situated in the outer mitochondrial region, is Ru(II) complexes 6 and 12 in the mice models did not cause any\nknown to control the intrinsic apoptosis pathway by suppressing obvious damage to the organ tissues.33\napoptosis.63 Caspase-3 and caspase-9 proteins are members of\nthe aspartate-specific cysteine proteases that are known to\npromote apoptosis. The Bax is a nuclear-encoded protein\n \u25a0 CONCLUSIONS\n Furoylthiourea ligands and their Ru(II)-p-cymene complexes\nlocated in the outer mitochondrial membrane, which is said to (1\u221212) were synthesized and characterized by spectroscopic\nbe anti-apoptotic.64 From Figure 7, an overexpression of methods. Although all of the complexes (1\u221212) displayed a\n\n\n\n\nFigure 6. (a) Cell cycle arrest in MDA-MB-231 cells with control, complex 6, and complex 12 and (b) percentage cell count at each phase of the cell\ncycle.\n\n 11768 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n\n\n\nFigure 7. (a) Expression levels of pro- and anti-apoptotic proteins in MDA-MB-231 cancer cell line upon treatment with complexes 6 and 12 and (b)\nthe percentage expression of protein levels in the cancer cell treated with complexes 6 and 12 for 48 h.\n\n this work conclude that half-sandwich Ru(II)\u2212arene aroylth-\n iourea complexes continue to serve as potential anticancer\n agents and in future, their anticancer activity could be further\n enhanced by tuning the mode of coordination, the choice of\n terminal substituents, and the presence of other biologically\n important heterocycles apart from furan.\n\n \u25a0 EXPERIMENTAL SECTION\n Furoylthiourea Ligand Synthesis. Ligands L1\u2212L6 were\n synthesized according to literature procedures.65,66 In a 500 mL\n round-bottom flask, furoyl chloride (1.984 g, 10 mmol) and potassium\n thiocyanate (0.971 g, 10 mmol) were taken in acetone and refluxed for\n an hour. After cooling the reaction mixture, the corresponding aniline\n (0.841\u22121.229 g, 10 mmol) was added, and the mixture was stirred for 4\n h. The progress of the reaction was monitored by thin-layer\n chromatography. On completion, HCl (0.1 N, 500 mL) and distilled\n water (500 mL) were added to the mixture, which led to the formation\n of a yellow precipitate, which was washed well with water and dried in\nFigure 8. Mice organs (kidneys, lungs, spleen, liver, and heart) treated vacuo.\nwith saline, cisplatin (4 mg/kg), complex 6 (8 mg/kg), and complex 12 N-((2-nitrophenyl)carbamothioyl)furan-2-carboxamide (L1).\n(8 mg/kg) were stained with hematoxylin and eosin. Yield: 82%. Light yellow solid. Mp: 162 \u00b0C. Anal. Calcd. for\n C12H9N3O4S (%): C, 49.48; H, 3.11; N, 14.43; S, 11.01. Found: C,\n 49.51; H, 3.15; N, 14.46; S, 11.05. UV\u2212vis (CH3CN): \u03bbmax, nm (\u03b5, dm3\nmonodentate coordination of the furoylthiourea ligands with the mol\u22121 cm\u22121) 271 (7535), 292 (9939). FT-IR (KBr, cm\u22121): 3431 (m;\nRu(II) ion via S in their solution state, X-ray structures of \u03bd(amide N\u2212H)), 3081 (s; \u03bd(thioamide N\u2212H)), 1682 (s; \u03bd(C\ufffdO)),\n 1264 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 13.18 (s, 1H,\ncomplexes 1 and 3 revealed a bidentate N, S coordination of the\n O\ufffdCNH), 9.35 (s, 1H, S\ufffdCNH), 8.48 (d, J = 9.3 Hz, 1H, phenyl),\nligands with the Ru(II) ion, and those of 4 and 6 showed a 8.12 (d, J = 8.3 Hz, 1H, phenyl), 7.67 (dd, J = 16.4, 7.9 Hz, 2H, furoyl),\nmonodentate coordination of the ligands. Further, biomolecular 7.49\u22127.37 (m, 2H, phenyl), 6.66 (dd, J = 3.6, 1.7 Hz, 1H, furoyl). 13C\n(DNA and BSA) interaction studies (absorption, emission, and NMR (100 MHz, CDCl3): \u03b4, ppm 179.0 (C\ufffdS), 156.3 (C\ufffdO), 146.6\ndocking) with the complexes revealed that complexes of the type (furoyl), 144.9 (O2N\u2212C), 142.3 (furoyl), 133.4, 132.3, 128.2, 126.8,\n[(\u03b76-p-cymene)RuCl(L)(PPh3)]+ (7\u221212) displayed greater 125.1 (phenyl), 119.7, 113.8 (furoyl).\nbinding affinity than [(\u03b76-p-cymene)RuCl2(L)] (1\u22126). Non- N-((4-methyl-2-nitrophenyl)carbamothioyl)furan-2-carbox-\ncovalent binding interactions (intercalation) of the complexes amide (L2). Yield: 78%. Bright yellow solid. Mp: 164 \u00b0C. Anal. Calcd.\n for C13H11N3O4S (%): C, 51.14; H, 3.63; N, 13.76; S, 10.50. Found: C,\nwith CT DNA were inferred from the spectroscopic analysis.\n 51.10; H, 3.60; N, 13.72; S, 10.48. UV\u2212vis (CH3CN): \u03bbmax, nm (\u03b5, dm3\nThereafter, in vitro cytotoxicity against breast cancer cells, mol\u22121 cm\u22121) 274 (7539), 296 (9942). FT-IR (KBr, cm\u22121): 3449 (m;\nnamely, MCF-7, T47-D, and MDA-MB-231, and a normal cell \u03bd(amide N\u2212H)), 3094 (s; \u03bd(thioamide N\u2212H)), 1690 (s; \u03bd(C\ufffdO)),\nline (MCF-10a) was evaluated. The IC50 values revealed that the 1256 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 13.03 (s, 1H,\ncomplexes possessed greater activity than their ligand counter- O\ufffdCNH), 9.30 (s, 1H, S\ufffdCNH), 8.27 (d, J = 8.4 Hz, 1H, phenyl),\nparts. Positively, they remained noncytotoxic on the normal 7.90 (d, J = 1.5 Hz, 1H, furoyl), 7.61 (d, J = 1.4 Hz, 1H, furoyl),7.50\u2212\nMCF-10a cell line. Complexes 6 and 12 emerged as the most 7.33 (m, 2H, phenyl), 6.64\u22126.62 (dd, J = 3.6, 1.8 Hz, 1H, furoyl), 2.44\nactive ones, displaying remarkable IC50 values against the MDA- (s, 3H, methyl). 13C NMR (100 MHz, CDCl3): \u03b4, ppm 179.1 (C\ufffdS),\nMB-231 cancer cell line (8.5 and 0.1 \u03bcM). Further, these two 156.0 (C\ufffdO), 146.5 (furoyl), 144.9 (O2N\u2212C), 142.2 (furoyl), 137.7,\n 134.1, 129.7, 128.0, 125.3 (phenyl), 119.4, 113.6 (furoyl), 20.9\ncomplexes induced apoptosis on the above cancer cell line,\n (methyl).\nwhich was inferred from AO/EB staining, DAPI staining, N-((4-methoxy-2-nitrophenyl)carbamothioyl)furan-2-car-\nwestern blot, and flow cytometry analyses. Lastly, in vivo results boxamide (L3). Yield: 75%. Bright yellow solid. Mp: 168 \u00b0C. Anal.\nindicated that these two complexes did not cause any noticeable Calcd. for C13H11N3O5S (%): C, 48.60; H, 3.45; N, 13.08; S, 9.98.\ndamage to the organs of the mice. The findings obtained from Found: C, 48.63; H, 3.47; N, 13.11; S, 10.01. UV\u2212vis (CH3CN): \u03bbmax,\n\n 11769 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\nnm (\u03b5, dm3 mol\u22121 cm\u22121) 279 (7542), 293 (9939). FT-IR (KBr, cm\u22121): UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 265 (7539), 285\n3451 (m; \u03bd(amide N\u2212H)), 3135 (s; \u03bd(thioamide N\u2212H)), 1691 (s; (7691), 352 (8509), 425 (9648). FT-IR (KBr, cm\u22121): 3442 (m;\n\u03bd(C\ufffdO)), 1249 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm \u03bd(amide N\u2212H)), 3139 (s; \u03bd(thioamide N\u2212H)), 1681 (s; \u03bd(C\ufffdO)),\n12.89 (s, 1H, O\ufffdCNH), 9.32 (s, 1H, S\ufffdCNH), 8.20 (d, J = 9.1 Hz, 1183 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 13.15 (s, 1H,\n1H, phenyl), 7.63 (d, J = 1.3 Hz, 1H, furoyl), 7.59 (d, J = 3.0 Hz, 1H, O\ufffdCNH), 11.25 (s, 1H, S\ufffdCNH), 8.12 (d, J = 8.2 Hz, 1H, phenyl),\nphenyl),7.45 (d, J = 3.6 Hz, 1H, phenyl), 7.22 (dd, J = 9.1, 3.0 Hz, 1H, 7.99 (t, J = 5.0 Hz, 2H, phenyl), 7.69 (dd, J = 16.4, 8.1 Hz, 2H, furoyl),\nfuroyl), 6.65 (dd, J = 3.6, 1.7 Hz, 1H, furoyl), 3.90 (s, 3H, methoxy). 13C 7.51 (t, J = 7.7 Hz, 1H, phenyl), 6.53 (dd, J = 3.6, 1.5 Hz, 1H, furoyl),\nNMR (100 MHz, CDCl3): \u03b4, ppm 179.2 (C\ufffdS), 157.9 (C\ufffdO), 156.2 5.40 (d, J = 5.9 Hz, 2H, aromatic H of p-cymene), 5.24 (d, J = 5.9 Hz,\n(C\u2212OCH3), 146.6 (furoyl), 144.9 (O2N\u2212C), 143.5 (furoyl), 129.7, 2H, aromatic H of p-cymene), 2.89 (sept, J = 6.9 Hz, 1H, CH(CH3)2),\n125.2, 120.0 (phenyl), 119.3, 113.6 (furoyl), 109.1 (phenyl), 56.1 2.21 (s, 3H, CH3), 1.28 (d, J = 4.2 Hz, 6H, CH(CH3)2). 13C NMR (100\n(OCH3). MHz, CDCl3): \u03b4, ppm 181.0 (C\ufffdS), 158.3 (C\ufffdO), 148.4 (furoyl),\n N-((5-chloro-2-nitrophenyl)carbamothioyl)furan-2-carboxa- 144.4 (O2N\u2212C), 143.9 (furoyl), 133.6, 130.8, 128.6, 125.4, 122.4\nmide (L4). Yield: 79%. Brownish-yellow solid. Mp: 172 \u00b0C. Anal. (phenyl), 112.9 (furoyl), 104.0, 100.0, 84.2, 82.8 (p-cymene), 30.5\nCalcd. for C12H8ClN3O4S (%): C, 44.25; H, 2.48; N, 12.90; S, 9.84. (CH(CH3)2), 22.1 (CH(CH3)2), 18.3 (CH3). ESI\u2212MS (m/z) [Found\nFound: C, 44.21; H, 2.45; N, 12.88; S, 8.82. UV\u2212vis (CH3CN): \u03bbmax, (Calcd.)]: 526.0383 (526.0375) [M \u2212 HCl \u2212 Cl\u2212]+.\nnm (\u03b5, dm3 mol\u22121 cm\u22121) 269 (7531), 282 (9930). FT-IR (KBr, cm\u22121): [Dichloro(p-cymene)(N-((4-methyl-2-nitrophenyl)-\n3459 (m; \u03bd(amide N\u2212H)), 3119 (s; \u03bd(thioamide N\u2212H)), 1691 (s; carbamothioyl)furan-2-carboxamide)ruthenium(II)] (2). Yield:\n\u03bd(C\ufffdO)), 1248 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 82%. Orange solid. Mp: 195 \u00b0C. Anal. Calcd. for C23H25Cl2N3O4RuS\n13.33 (s, 1H, O\ufffdCNH), 9.34 (s, 1H, S\ufffdCNH), 8.67 (d, J = 2.1 Hz, (%): C, 45.18; H, 4.12; N, 6.87; S, 5.24. Found: C, 45.21; H, 4.14; N,\n1H, phenyl), 8.06 (dd, J = 12.2, 9.0 Hz, 2H, phenyl), 7.66\u22127.39 (m, 2H, 6.90; S, 5.28. UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 269\nfuroyl), 6.65\u22126.63 (dd, J = 4.7, 1.8 Hz, 1H, furoyl). 13C NMR (100 (7540), 288 (7693), 359 (8514), 431 (9651). FT-IR (KBr, cm\u22121):\nMHz, CDCl3): \u03b4, ppm 179.0 (C\ufffdS), 155.9 (C\ufffdO), 146.6 (furoyl), 3438 (m; \u03bd(amide N\u2212H)), 3089 (s; \u03bd(thioamide N\u2212H)), 1683 (s;\n144.7 (O2N\u2212C), 141.8 (furoyl), 140.0, 133.7, 127.6, 126.3, 119.8 \u03bd(C\ufffdO)), 1171 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm\n(phenyl), 117.8, 113.6 (furoyl). 13.06 (s, 1H, O\ufffdCNH), 11.23 (s, 1H, S\ufffdCNH), 7.99 (d, J = 3.6 Hz,\n N-((5-chloro-4-methyl-2-nitrophenyl)carbamothioyl)furan- 1H, phenyl), 7.93 (s, 1H, phenyl), 7.85 (d, J = 8.3 Hz, 1H, furoyl), 7.66\n2-carboxamide (L5). Yield: 84%. Orange-yellow solid. Mp: 178 \u00b0C. (d, J = 1.5 Hz, 1H, phenyl), 7.49 (d, J = 9.9 Hz, 1H, furoyl), 6.54 (dd, J =\nAnal. Calcd. for C13H10ClN3O4S (%): C, 45.96; H, 2.97; N, 12.37; S, 3.7, 1.7 Hz, 1H, furoyl), 5.40 (d, J = 6.0 Hz, 2H, aromatic H of p-\n9.49. Found: C, 45.93; H, 2.92; N, 12.34; S, 9.46. UV\u2212vis (CH3CN): cymene), 5.24 (d, J = 6.1 Hz, 2H, aromatic H of p-cymene), 2.91 (sept, J\n\u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 261 (7526), 276 (9922). FT-IR (KBr, = 6.9 Hz, 1H, CH(CH3)2), 2.50 (s, 3H, CH3), 2.22 (s, 3H, CH3), 1.28\ncm\u22121): 3451 (m; \u03bd(amide N\u2212H)), 3129 (s; \u03bd(thioamide N\u2212H)), 1679 (d, J = 7.0 Hz, 6H, CH(CH3)2). 13C NMR (100 MHz, CDCl3): \u03b4, ppm\n(s; \u03bd(C\ufffdO)), 1251 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, DMSO-d6): \u03b4, 181.1 (C\ufffdS), 158.6 (C\ufffdO), 148.4 (furoyl), 144.7 (O2N\u2212C), 143.8\nppm 12.64 (s, 1H, O\ufffdCNH), 11.69 (s, 1H, S\ufffdCNH), 8.17 (s, 1H, (furoyl), 139.4, 134.5, 129.6, 128.4, 125.6 (phenyl), 122.4, 112.9\nphenyl), 8.11\u22127.98 (m, 2H, furoyl), 7.91 (s, 1H, phenyl), 6.78 (dd, J = (furoyl), 104.2, 100.4, 84.9, 82.9 (p-cymene), 30.6 (CH(CH3)2), 22.4\n3.6, 1.7 Hz, 1H, furoyl), 2.44 (s, 3H, CH3). 13C NMR (100 MHz, (CH(CH3)2), 21.1 (CH3), 18.3 (CH3). ESI\u2212MS (m/z) [Found\nDMSO-d6): \u03b4, ppm 181.1 (C\ufffdS), 158.1 (C\ufffdO), 149.2 (furoyl), (Calcd.)]: 540.0561 (540.0531) [M \u2212 HCl \u2212 Cl\u2212]+.\n144.8 (O2N\u2212C), 142.7 (furoyl), 138.7, 136.2, 131.5, 130.1, 127.3 [Dichloro(p-cymene)(N-((4-methoxy-2-nitrophenyl)-\n(phenyl), 119.9, 113.1 (furoyl), 19.0 (CH3). carbamothioyl)furan-2-carboxamide)ruthenium(II)] (3). Yield:\n N-((4,5-dimethyl-2-nitrophenyl)carbamothioyl)furan-2-car- 78%. Light orange solid. Mp: 199 \u00b0C. Anal. Calcd. for\nboxamide (L6). Yield: 88%. Pale yellow solid. Mp: 171 \u00b0C. Anal. C23H25Cl2N3O5RuS (%): C, 44.02; H, 4.02; N, 6.70; S, 5.11. Found:\nCalcd. for C14H13N3O4S (%): C, 52.66; H, 4.10; N, 13.16; S, 10.04. C, 44.04; H, 4.05; N, 6.73; S, 5.16. UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3\nFound: C, 52.68; H, 4.14; N, 13.19; S, 10.08. UV\u2212vis (CH3CN): \u03bbmax, mol\u22121 cm\u22121) 272 (7542), 294 (7697), 362 (8516), 436 (9655). FT-IR\nnm (\u03b5, dm3 mol\u22121 cm\u22121) 266 (7529), 281 (9928). FT-IR (KBr, cm\u22121): (KBr, cm\u22121): 3445 (m; \u03bd(amide N\u2212H)), 3123 (s; \u03bd(thioamide N\u2212\n3450 (m; \u03bd(amide N\u2212H)), 3189 (s; \u03bd(thioamide N\u2212H)), 1691 (s; H)), 1691 (s; \u03bd(C\ufffdO)), 1178 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz,\n\u03bd(C\ufffdO)), 1269 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, DMSO-d6): \u03b4, CDCl3): \u03b4, ppm 12.92 (s, 1H, O\ufffdCNH), 11.23 (s, 1H, S\ufffdCNH),\nppm 12.54 (s, 1H, O\ufffdCNH), 11.59 (s, 1H, S\ufffdCNH), 8.10 (s, 1H, 7.99 (d, J = 3.6 Hz, 1H, furoyl), 7.81 (d, J = 9.0 Hz, 1H, phenyl), 7.68\u2212\nphenyl), 7.99\u22127.82 (m, 2H, furoyl), 7.68 (s, 1H, phenyl), 6.78 (dd, J = 7.58 (m, 2H, phenyl), 7.15 (s, 1H, furoyl), 6.54 (dd, J = 3.6, 1.4 Hz, 1H,\n3.5, 1.6 Hz, 1H, furoyl), 2.50 (s, 3H, CH3), 2.33 (s, 3H, CH3). 13C furoyl), 5.39 (d, J = 5.9 Hz, 2H, aromatic H of p-cymene), 5.23 (d, J =\nNMR (100 MHz, DMSO-d6): \u03b4, ppm 180.9 (C\ufffdS), 158.1 (C\ufffdO), 5.9 Hz, 2H, aromatic H of p-cymene), 3.93 (s, 3H, OCH3), 2.98 (sept, J\n149.1 (furoyl), 144.9 (O2N\u2212C), 144.1 (furoyl), 142.1, 137.3, 130.9, = 7.4 Hz, 1H, CH(CH3)2), 2.34 (s, 3H, CH3), 1.31 (d, J = 8.0 Hz, 6H,\n130.1, 125.5 (phenyl), 119.5, 113.1 (furoyl), 19.9 (CH3), 19.1 (CH3). CH(CH3)2). 13C NMR (100 MHz, CDCl3): \u03b4, ppm 180.8 (C\ufffdS),\n Synthesis of the Ru(II)\u2212Furoylthiourea Complexes. 158.5 (C\ufffdO), 148.2 (furoyl), 144.3 (O2N\u2212C), 131.1 (furoyl), 129.2,\n[RuCl2(\u03b76-p-cymene)]2 (0.122 g, 0.2 mmol) and the ligand (0.116\u2212 128.1, 125.2 (phenyl), 121.7 (furoyl), 120.1 (phenyl), 112.7 (furoyl),\n0.136 g, 0.4 mmol) (L1\u2212L6) were taken in 10 mL of toluene and stirred 109.6, 103.7, 100.2, 84.4, 82.7 (p-cymene), 56.3 (OCH3), 30.4,\nfor 12 h at 27 \u00b0C, forming a clear orange solution. After the completion (CH(CH3)2), 22.2 (CH(CH3)2), 18.2 (CH3). ESI\u2212MS (m/z) [Found\nof reaction as inferred from TLC, the volume of solution was reduced to (Calcd.)]: 556.0461 (556.0482) [M \u2212 HCl \u2212 Cl\u2212]+.\n2 mL and the addition of petroleum ether (60\u221280 \u00b0C) (10 mL) [Dichloro(p-cymene)(N-((5-chloro-2-nitrophenyl)-\nresulted in an orange solid, which was washed with hexane and dried in carbamothioyl)furan-2-carboxamide)ruthenium(II)] (4). Yield:\nvacuo to yield complexes 1\u22126. 83%. Bright orange solid. Mp: 205 \u00b0C. Anal. Calcd. for\n [RuCl2(\u03b76-p-cymene)(PPh3)] (0.113 g, 0.1 mmol) and the ligands C22H22Cl3N3O4RuS (%): C, 41.82; H, 3.51; N, 6.65; S, 5.07. Found:\n(0.029\u22120.033 g, 0.1 mmol) were dissolved in 15 mL of toluene and C, 41.78; H, 3.50; N, 6.61; S, 5.05. UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3\nstirred at room temperature for 20 h. After this, 0.015\u22120.0025 g of mol\u22121 cm\u22121): 275 (7546), 287 (7694), 365 (8519), 442 (9659). FT-IR\nNH4PF6 dissolved in methanol was added to the reaction mixture, (KBr, cm\u22121): 3430 (m; \u03bd(amide N\u2212H)), 3159 (s; \u03bd(thioamide N\u2212\nwhich was stirred for 4 h. The resultant orange solution was H)), 1691 (s; \u03bd(C\ufffdO)), 1178 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz,\nconcentrated to 2 mL, and the addition of cold hexane gave a mild CDCl3): \u03b4, ppm 13.29 (s, 1H, O\ufffdCNH), 11.24 (s, 1H, S\ufffdCNH),\norange precipitate, which was filtered, washed with petroleum ether, 8.26\u22128.02 (m, 2H, phenyl), 7.66 (s, 1H, furoyl), 7.45 (d, J = 8.7 Hz, 1H,\nand dried to yield complexes 7\u221212. furoyl), 7.20 (d, J = 36.0 Hz, 1H, phenyl), 6.53 (dd, J = 4.8, 2.4 Hz, 1H,\n [Dichloro(p-cymene)(N-((2-nitrophenyl)carbamothioyl)- furoyl), 5.43 (d, J = 3.7 Hz, 2H, aromatic H of p-cymene), 5.31 (d, J =\nfuran-2-carboxamide)ruthenium(II)] (1). Yield: 91%. Light orange 5.6 Hz, 2H, aromatic H of p-cymene), 3.04 (sept, J = 6.8 Hz, 1H,\nsolid. Mp: 191 \u00b0C. Anal. Calcd. for C22H23Cl2N3O4RuS (%): C, 44.23; CH(CH3)2), 2.23 (s, 3H, CH3), 1.30 (d, J = 6.8 Hz, 6H, CH(CH3)2).\n 13\nH, 3.88; N, 7.03; S, 5.37. Found: C, 44.25; H, 3.92; N, 7.06; S, 5.40. C NMR (100 MHz, CDCl3): \u03b4, ppm 181.1 (C\ufffdS), 158.5 (C\ufffdO),\n\n 11770 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n148.6 (furoyl) 144.8 (O2N\u2212C), 140.0 (furoyl), 132.0, 128.9, 128.1, (II)]hexaflurophosphate (8). Yield: 77%. Orange solid. Mp: 215 \u00b0C.\n126.9, 125.5 (phenyl), 122.3, 112.4 (furoyl), 101.4, 97.1, 81.2, 80.6 (p- Anal. Calcd. for C41H40ClN3O4PRuS (%): C, 58.74; H, 4.81; N, 5.01; S,\ncymene), 30.5 (CH(CH3)2), 22.0 (CH(CH3)2), 19.0 (CH3). ESI\u2212MS 3.82. Found: C, 58.71; H, 4.76; N, 4.98; S, 3.78. UV\u2212vis (CH3CN):\n(m/z) [Found (Calcd.)]: 560.0255 (559.9985) [M \u2212 HCl \u2212 Cl\u2212]+. \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 268 (7539), 281 (7690), 359 (8514), 459\n [Dichloro(p-cymene)(N-((5-chloro-4-methyl-2-nitrophenyl)- (9648). FT-IR (KBr, cm\u22121): 3430 (m; \u03bd(amide N\u2212H)), 3159 (s;\ncarbamothioyl)furan-2-carboxamide)ruthenium(II)] (5). Yield: \u03bd(thioamide N\u2212H)), 1700 (s; \u03bd(C\ufffdO)), 1183 (s; \u03bd(C\ufffdS)), 1445,\n79%. Orange solid. Mp: 194 \u00b0C. Anal. Calcd. for C23H24Cl3N3O4RuS 1076 and 721 (s; \u03bd(PPh3)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 13.17\n(%): C, 42.77; H, 3.75; N, 6.51; S, 4.96. Found: C, 42.74; H, 3.72; N, (s, 1H, O\ufffdCNH), 9.32 (s, 1H, S\ufffdCNH), 8.46 (d, J = 8.3 Hz, 1H,\n6.48; S, 4.92. UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 270 phenyl), 8.10 (d, J = 8.3 Hz, 1H, furoyl), 7.83 (s, 1H, phenyl), 7.78 (s,\n(7541), 281 (7690), 358 (8514), 434 (9651). FT-IR (KBr, cm\u22121): 1H, phenyl), 7.62 (d, J = 2.5 Hz, 1H, furoyl), 7.40\u22127.29 (m, 15H, H of\n3442 (m; \u03bd(amide N\u2212H)), 3139 (s; \u03bd(thioamide N\u2212H)), 1681 (s; PPh3), 6.71 (dd, J = 3.9, 1.4 Hz, 1H, furoyl), 5.19 (dd, J = 9.9, 6.3 Hz,\n\u03bd(C\ufffdO)), 1179 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 2H, aromatic H of p-cymene), 4.98 (dd, J = 10.2, 8.4 Hz, 2H, aromatic\n13.18 (s, 1H, O\ufffdCNH), 12.19 (s, 1H, S\ufffdCNH), 8.05 (d, J = 24.6 Hz, H of p-cymene), 2.82 (sept, J = 5.9 Hz, 1H, CH(CH3)2), 1.85 (s, 3H,\n1H, phenyl), 7.99 (s, 1H, phenyl), 7.72 (d, J = 2.8 Hz, 1H, furoyl), 7.65 CH3), 1.08 (d, J = 7.0 Hz, 9H, CH(CH3)2; CH3). 13C NMR (100 MHz,\n(d, J = 1.5 Hz, 1H, furoyl), 6.53 (dd, J = 3.7, 1.6 Hz, 1H, furoyl), 5.42 (d, CDCl3): \u03b4, ppm 179.3 (C\ufffdS), 156.1 (C\ufffdO), 146.5 (O2N\ufffdC),\nJ = 6.1 Hz, 2H, aromatic H of p-cymene), 5.26 (d, J = 5.9 Hz, 2H, 144.9, 134.5 (furoyl), 134.3 (phenyl), 134.1 (furoyl), 133.6 (phenyl),\naromatic H of p-cymene), 2.90 (sept, J = 5.7 Hz, 1H, CH(CH3)2), 2.49 130.3, 130.2, 128.1 (C of PPh3), 127.9 (phenyl), 127.0 (C of PPh3),\n(s, 3H, CH3), 2.23 (s, 3H, CH3), 1.25 (d, J = 7.0 Hz, 6H, CH(CH3)2). 125.2 (furoyl), 119.6, 113.6 (phenyl), 111.0 (furoyl), 96.2, 88.9, 87.1,\n13\n C NMR (100 MHz, CDCl3): \u03b4, ppm 181.0 (C\ufffdS), 158.5 (C\ufffdO), 79.7 (p-cymene), 22.5 (CH(CH3)2), 22.3 (CH(CH3)2), 17.6 (CH3).\n 31\n148.7 (furoyl), 144.4 (O2N\u2212C), 141.6 (furoyl), 140.1, 137.0, 129.7, P NMR (162 MHz, CDCl3): \u03b4, ppm 26.81 (PPh3), \u2212131.5, \u2212134.3,\n127.1 (phenyl), 122.8, 112.9 (furoyl), 104.1 (phenyl), 101.2, 100.5, \u2212138.3, \u2212143.8, \u2212149.1, \u2212151.8, \u2212157.6 (PF6). ESI\u2212MS (m/z)\n84.2, 82.8 (p-cymene), 30.7 (CH(CH3)2), 22.0 (CH(CH3)2), 20.1 [Found (Calcd.)]: 838.1241 (838.1249) [M \u2212 PF6\u2212]+.\n(CH3), 18.2 (CH3). ESI\u2212MS (m/z) [Found (Calcd.)]: 574.0125 [Chlorotriphenylphosphine(p-cymene)(N-((4-methoxy-2-\n(574.0142) [M \u2212 HCl \u2212 Cl\u2212]+. nitrophenyl)carbamothioyl)furan-2-carboxamide)ruthenium-\n [Dichloro(p-cymene)(N-((4,5-dimethyl-2-nitrophenyl)- (II)]hexafluorophosphate (9). Yield: 81%. Orange solid. Mp: 221\ncarbamothioyl)furan-2-carboxamide)ruthenium(II)] (6). Yield: \u00b0C. Anal. Calcd. for C41H40ClN3O5PRuS (%): C, 57.64; H, 4.72; N,\n86%. Dark orange solid. Mp: 201 \u00b0C. Anal. Calcd. for 4.92; S, 3.75. Found: C, 57.60; H, 4.70; N, 4.88; S, 3.72. UV\u2212vis\nC24H27Cl2N3O4RuS (%): C, 46.08; H, 4.35; N, 6.72; S, 5.13. Found: (CH3CN): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 260 (7530), 275 (7686), 354\nC, 46.12; H, 4.36; N, 6.74; S, 5.17. UV\u2212vis (CH2Cl2): \u03bbmax, nm (\u03b5, dm3 (8510), 452 (9644). FT-IR (KBr, cm\u22121): 3426 (m; \u03bd(amide N\u2212H)),\nmol\u22121 cm\u22121) 275 (7546), 286 (7694), 362 (8517), 439 (9658). FT-IR 3140 (s; \u03bd(thioamide N\u2212H)), 1685 (s; \u03bd(C\ufffdO)), 1184 (s; \u03bd(C\ufffdS)),\n(KBr, cm\u22121): 3438 (m; \u03bd(amide N\u2212H)), 3115 (s; \u03bd(thioamide N\u2212 1451, 1092 and 715 (s; \u03bd(PPh3)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm\nH)), 1691 (s; \u03bd(C\ufffdO)), 1187 (s; \u03bd(C\ufffdS)). 1H NMR (400 MHz, 12.83 (s, 1H, O\ufffdCNH), 11.84 (s, 1H, S\ufffdCNH), 7.85 (s, 1H,\nCDCl3): \u03b4, ppm 12.94 (s, 1H, O\ufffdCNH), 11.19 (s, 1H, S\ufffdCNH), phenyl), 7.82 (d, J = 4.6 Hz, 1H, furoyl), 7.80 (s, 1H, phenyl), 7.76 (s,\n7.93 (d, J = 3.6 Hz, 1H, furoyl), 7.87 (s, 1H, phenyl), 7.64 (s, 1H, 1H, phenyl), 7.62 (s, 1H, furoyl), 7.45\u22127.33 (m, 15H, H of PPh3), 6.69\nfuroyl), 7.60 (s, 1H, phenyl), 6.49 (dd, J = 3.6, 1.6 Hz, 1H, furoyl), 5.35 (dd, J = 3.6, 1.6 Hz, 1H, furoyl), 5.20 (d, J = 6.1 Hz, 2H, aromatic H of p-\n(d, J = 5.9 Hz, 2H, aromatic H of p-cymene), 5.19 (d, J = 5.9 Hz, 2H, cymene), 5.00 (d, J = 5.5 Hz, 2H, aromatic H of p-cymene), 3.95 (s, 3H,\naromatic H of p-cymene), 2.93 (sept, J = 5.2 Hz, 1H, CH(CH3)2), 2.32 OCH3), 2.93 (sept, 1H, J = 5.8 Hz, CH(CH3)2), 1.89 (d, J = 10.4 Hz,\n(d, J = 5.3 Hz, 6H, CH3), 2.16 (s, 3H, CH3), 1.23 (d, J = 6.9 Hz, 6H, 6H, CH(CH3)2), 1.18 (s, 3H, CH3). 13C NMR (100 MHz, CDCl3): \u03b4,\nCH(CH3)2). 13C NMR (100 MHz, CDCl3): \u03b4, ppm 180.7 (C\ufffdS), ppm 181.3 (C\ufffdO), 159.9 (C\ufffdS), 147.9 (O2N\u2212C), 144.8 (furoyl),\n158.1 (C\ufffdO), 148.1 (furoyl), 144.5 (O2N\u2212C), 143.9 (furoyl), 141.4, 144.4 (phenyl), 134.3 (furoyl), 133.7 (phenyl), 131.6, 131.0 (C of\n137.8, 130.5, 128.3, 126.0 (phenyl), 121.8, 113.0 (furoyl), 103.7, 100.0, PPh3), 130.3 (phenyl), 128.6, 128.0 (C of PPh3), 122.2 (phenyl), 113.7,\n84.1, 82.8 (p-cymene), 30.4 (CH(CH3)2), 22.2 (CH(CH3)2), 19.9 111.2 (furoyl), 110.6 (phenyl), 102.3, 95.7, 88.9, 87.1 (p-cymene), 56.7\n(CH3), 19.4 (CH3), 18.2 (CH3). ESI\u2212MS (m/z) [Found (Calcd.)]: (OCH3), 30.7 (CH(CH3)2), 21.6 (CH(CH3)2), 17.2 (CH3). 31P NMR\n554.0729 (554.0688) [M \u2212 HCl \u2212 Cl\u2212]+. (162 MHz, CDCl3): \u03b4, ppm 30.78 (PPh3), \u2212130.9, \u2212135.4, \u2212139.9,\n [Chlorotriphenylphosphine(p-cymene)(N-((2-nitrophenyl)- \u2212144.4, \u2212148.3, \u2212152.3, \u2212157.0 (PF6). ESI\u2212MS (m/z) [Found\ncarbamothioyl)furan-2-carboxamide)ruthenium(II)]- (Calcd.)]: 854.1289 (854.1297) [M \u2212 PF6\u2212]+.\nhexaflurophosphate (7). Yield: 81%. Pale orange solid. Mp: 210 \u00b0C. [Chlorotriphenylphosphine(p-cymene)(N-((5-chloro-2-\nAnal. Calcd. for C40H38ClN3O4PRuS (%): C, 58.28; H, 4.65; N, 5.10; S, nitrophenyl)carbamothioyl)furan-2-carboxamide)ruthenium-\n3.89. Found: C, 58.32; H, 4.67; N, 5.13; S, 3.92. UV\u2212vis (CH3CN): (II)]hexaflurophosphate (10). Yield: 76%. Bright orange solid. Mp:\n\u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 263 (7536), 275 (7685), 350 (8510), 449 216 \u00b0C. Anal. Calcd. for C40H37Cl2N3O4PRuS (%): C, 55.95; H, 4.34;\n(9642). FT-IR (KBr, cm\u22121): 3430 (m; \u03bd(amide N\u2212H)), 3166 (s; N, 4.89; S, 3.73. Found: C, 55.99; H, 4.37; N, 4.93; S, 3.76. UV\u2212vis\n\u03bd(thioamide N\u2212H)), 1695 (s; \u03bd(C\ufffdO)), 1186 (s; \u03bd(C\ufffdS)), 1437, (CH3CN): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 250 (7523), 262 (7679), 343\n1092 and 710 (s; \u03bd(PPh3)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm 12.97 (8501), 444 (9638). FT-IR (KBr, cm\u22121): 3446 (m; \u03bd(amide N\u2212H)),\n(s, 1H, O\ufffdCNH), 11.83 (s, 1H, S\ufffdCNH), 7.89 (d, J = 1.2 Hz, 1H, 3125 (s; \u03bd(thioamide N\u2212H)), 1683 (s; \u03bd(C\ufffdO)), 1187 (s; \u03bd(C\ufffdS)),\nfuroyl), 7.75 (dd, J = 1.7, 0.7 Hz, 1H, phenyl), 7.60 (dd, J = 3.7, 0.7 Hz, 1452, 1081 and 715 (s; \u03bd(PPh3)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm\n1H, furoyl), 7.52\u22127.38 (m, 15H, H of PPh3), 7.36 (dd, J = 2.8, 1.2 Hz, 12.97 (s, 1H, O\ufffdCNH), 11.85 (s, 1H, S\ufffdCNH), 7.90 (s, 1H,\n3H, phenyl), 6.68 (dd, J = 3.7, 1.7 Hz, 1H, furoyl), 5.62 (d, J = 6.4 Hz, phenyl), 7.85\u22127.78 (m, 1H, furoyl), 7.77 (s, 1H, phenyl), 7.60 (d, J =\n2H, aromatic H of p-cymene), 5.40 (dd, J = 78.0, 6.3 Hz, 2H, aromatic 2.9 Hz, 1H, phenyl), 7.49 (s, 1H, furoyl), 7.49\u22127.19 (m, 15H, H of\nH of p-cymene), 2.67 (sept, J = 4.4 Hz, 1H, CH(CH3)2), 2.49 (s, 3H, PPh3), 6.67 (dd, J = 16.0, 2.9 Hz, 1H, furoyl), 5.57 (d, J = 7.6 Hz, 2H,\nCH(CH3)2), 1.89 (s, 3H, CH(CH3)2), 1.62 (s, 3H, CH3). 13C NMR aromatic H of p-cymene), 5.10 (d, J = 5.9 Hz, 2H, aromatic H of p-\n(100 MHz, CDCl3): \u03b4, ppm 181.4 (C\ufffdS), 158.3 (C\ufffdO), 147.8 cymene), 2.99 (sept, J = 5.4 Hz, 1H, CH(CH3)2), 2.50 (s, 3H, CH3),\n(O2N\u2212C), 144.4 (furoyl), 143.6 (phenyl), 140.7 (furoyl), 135.5 1.14 (d, J = 7.6 Hz, 6H, CH(CH3)2). 13C NMR (100 MHz, CDCl3): \u03b4,\n(phenyl), 131.6, 130.2, 129.7 (C of PPh3), 128.6 (phenyl), 127.0 (C of ppm 178.9 (C\ufffdS), 155.9 (C\ufffdO), 146.7 (O2N\u2212C), 144.6 (furoyl),\nPPh3), 125.8 (furoyl), 121.2, 115.7 (phenyl), 113.8 (furoyl), 93.6, 92.1, 140.0 (phenyl), 134.4 (furoyl), 134.3, 133.6 (phenyl), 130.3, 130.2 (C\n91.0, 89.7 (p-cymene), 30.8 (CH(CH3)2), 21.2 (CH(CH3)2), 17.4 of PPh3), 128.0 (phenyl), 127.9 (C of PPh3), 119.8 (phenyl), 113.6,\n(CH3). 31P NMR (162 MHz, CDCl3): \u03b4, ppm 30.82 (PPh3), \u2212130.7, 111.4 (furoyl), 95.9 (phenyl), 89.1, 89.0, 87.2, 87.0 (p-cymene), 30.3\n\u2212135.2, \u2212139.3, \u2212143.7, \u2212148.0, \u2212152.7, \u2212156.2 (PF6). ESI\u2212MS (m/ (CH(CH3)2), 22.0 (CH(CH3)2), 17.8 (CH3). 31P NMR (162 MHz,\nz) [Found (Calcd.)]: 824.3133 (824.3148) [M \u2212 PF6\u2212]+. CDCl3): \u03b4, ppm 28.48 (PPh3), \u2212132.5, \u2212136.4, \u2212139.3, \u2212143.8,\n [Chlorotriphenylphosphine(p-cymene)(N-((4-methyl-2- \u2212147.8, \u2212152.3, \u2212155.7 (PF6). ESI\u2212MS (m/z) [Found (Calcd.)]:\nnitrophenyl)carbamothioyl)furan-2-carboxamide)ruthenium- 858.7560 (858.7568) [M \u2212 PF6\u2212]+.\n\n 11771 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\fInorganic Chemistry pubs.acs.org/IC Article\n\n [Chlorotriphenylphosphine(p-cymene)(N-((5-chloro-4-\nmethyl-2-nitrophenyl)carbamothioyl)furan-2-carboxamide)-\nruthenium(II)]hexaflurophosphate (11). Yield: 80%. Bright orange\n \u25a0 AUTHOR INFORMATION\n Corresponding Author\nsolid. Mp: 224 \u00b0C. Anal. Calcd. for C41H39Cl2N3O4PRuS (%): C, 56.42; Ramasamy Karvembu \u2212 Department of Chemistry, National\nH, 4.50; N, 4.81; S, 3.67. Found: C, 56.38; H, 4.48; N, 4.77; S, 3.63. Institute of Technology, Tiruchirappalli 620015, India;\nUV\u2212vis (CH3CN): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 258 (7529), 265 orcid.org/0000-0001-8966-8602; Email: kar@nitt.edu\n(7680), 347 (8506), 449 (9640). FT-IR (KBr, cm\u22121): 3434 (m;\n\u03bd(amide N\u2212H)), 3131 (s; \u03bd(thioamide N\u2212H)), 1689 (s; \u03bd(C\ufffdO)), Authors\n1171 (s; \u03bd(C\ufffdS)), 1441, 1087 and 720 (s; \u03bd(PPh3)). 1H NMR (400 Dorothy Priyanka Dorairaj \u2212 Department of Chemistry,\nMHz, CDCl3): \u03b4, ppm 13.11 (s, 1H, O\ufffdCNH), 11.96 (s, 1H, S\ufffd National Institute of Technology, Tiruchirappalli 620015,\nCNH), 8.02 (s, 1H, phenyl), 7.81 (dd, J = 12.8, 7.7 Hz, 1H, furoyl),\n India\n7.70\u22127.59 (m, 1H, phenyl), 7.60\u22127.35 (m, 15H, H of PPh3), 6.74 (dd,\nJ = 10.0, 2.4 Hz, 1H, furoyl), 5.74 (d, J = 13.8 Hz, 2H, aromatic H of p- Jebiti Haribabu \u2212 Faculty of Medicine, University of Atacama,\ncymene), 5.45 (d, J = 21.9 Hz, 2H, aromatic H of p-cymene), 2.96 (sept, 1532502 Copiapo, Chile; orcid.org/0000-0001-8855-\nJ = 5.9 Hz, 1H, CH(CH3)2), 2.50 (s, 3H, CH3), 1.97 (s, 3H, CH3), 1.14 032X\n(d, J = 6.7 Hz, 6H, CH(CH3)2). 13C NMR (100 MHz, CDCl3): \u03b4, ppm Mahendiran Dharmasivam \u2212 Department of Chemistry,\n179.5 (C\ufffdS), 157.8 (C\ufffdO), 146.5 (O2N\u2212C), 145.0 (furoyl), 134.4 Griffith Institute for Drug Discovery, Griffith University,\n(phenyl), 134.2 (furoyl), 131.2 (phenyl), 130.2, 129.9 (C of PPh3), Brisbane, Queensland 4111, Australia\n128.5 (phenyl), 128.5 (C of PPh3), 127.9 (phenyl), 127.9, 125.3 Rahime Eshaghi Malekshah \u2212 Medical Biomaterial Research\n(furoyl), 119.9 (phenyl), 119.4 (furoyl), 109.3, 95.8, 89.1, 87.3 (p- Centre (MBRC), Tehran University of Medical Sciences,\ncymene), 30.7 (CH(CH3)2), 30.4 (CH(CH3)2), 21.5 (CH3), 17.4\n Tehran 1416634793, Iran\n(CH3). 31P NMR (162 MHz, CDCl3): \u03b4, ppm 29.83 (PPh3), \u2212133.5,\n\u2212137.1, \u2212140.7, \u2212144.3, \u2212147.7, \u2212151.2, \u2212154.7 (PF6). ESI\u2212MS (m/ Mohamed Kasim Mohamed Subarkhan \u2212 The First Affiliated\nz) [Found (Calcd.)]: 872.0833 (872.0840) [M \u2212 PF6\u2212]+. Hospital, Key Laboratory of Combined Multi-Organ\n [Chlorotriphenylphosphine(p-cymene)(N-((4,5-dimethyl-2- Transplantation, Ministry of Public Health, School of\nnitrophenyl)carbamothioyl)furan-2-carboxamide)ruthenium- Medicine, Zhejiang University, Hangzhou 310018, P. R.\n(II)]hexaflurophosphate (12). Yield: 83%. Light orange solid. Mp: China; orcid.org/0000-0002-1164-478X\n219 \u00b0C. Anal. Calcd. for C42H42ClN3O4PRuS (%): C, 59.18; H, 4.97; N, Cesar Echeverria \u2212 Faculty of Medicine, University of Atacama,\n4.93; S, 3.76. Found: C, 59.20; H, 5.00; N, 4.97; S, 3.79. UV\u2212vis\n 1532502 Copiapo, Chile\n(CH3CN): \u03bbmax, nm (\u03b5, dm3 mol\u22121 cm\u22121) 261 (7531), 272 (7685), 351\n(8509), 454 (9645). FT-IR (KBr, cm\u22121): 3454 (m; \u03bd(amide N\u2212H)), Complete contact information is available at:\n3131 (s; \u03bd(thioamide N\u2212H)), 1691 (s; \u03bd(C\ufffdO)), 1171 (s; \u03bd(C\ufffdS)), https://pubs.acs.org/10.1021/acs.inorgchem.3c00757\n1441, 1089 and 720 (s; \u03bd(PPh3)). 1H NMR (400 MHz, CDCl3): \u03b4, ppm\n13.00 (s, 1H, O\ufffdCNH), 11.91 (s, 1H, S\ufffdCNH), 7.92 (s, 1H, Notes\nphenyl), 7.84 (d, J = 10.1 Hz, 1H, furoyl), 7.77 (s, 1H, phenyl), 7.63 (d,\n The authors declare no competing financial interest.\nJ = 3.6 Hz, 1H, furoyl), 7.50\u22127.33 (m, 15H, H of PPh3), 6.70 (dd, J =\n3.7, 1.7 Hz, 1H, furoyl), 5.66 (d, J = 10.7 Hz, 2H, aromatic H of p-\ncymene), 5.41 (d, J = 6.2 Hz, 2H, aromatic H of p-cymene), 2.61 (sept, J\n= 6.4 Hz, 1H, CH(CH3)2), 2.40 (s, 3H, CH3), 2.30 (s, 3H, CH3), 1.93\n \u25a0 ACKNOWLEDGMENTS\n D.P.D. thanks the Department of Science and Technology,\n(s, 3H, CH3), 1.12 (d, J = 6.9 Hz, 6H, CH(CH3)2). 13C NMR (100 Ministry of Science and Technology, Government of India, for\nMHz, CDCl3): \u03b4, ppm 181.4 (C\ufffdS), 158.2 (C\ufffdO), 147.8 (O2N\u2212C), the DST-INSPIRE doctoral fellowship (IF170457). J.H. thanks\n145.2 (furoyl), 144.1 (phenyl), 141.7 (furoyl), 139.3 (phenyl), 131.8,\n131.2 (C of PPh3), 130.6 (phenyl), 128.6 (C of PPh3), 128.5 (phenyl),\n the Fondo Nacional de Ciencia y Tecnologia (FONDECYT,\n128.4, 126.4 (furoyl), 113.7 (phenyl), 102.3 (furoyl), 93.5, 92.3, 89.2, Project Nos. 3200391 and 11170840) for the post-doctoral\n87.2 (p-cymene), 30.9 (CH(CH3)2), 22.2 (CH(CH3)2), 21.5 (CH3), fellowship. R.K. thanks the DST-SERB for financial assistance\n19.9 (CH3), 19.7 (CH3). 31P NMR (162 MHz, CDCl3): \u03b4, ppm 30.3 (CRG/2022/003145).\n(PPh3), \u2212130.5, \u2212135.6, \u2212139.3, \u2212144.5, \u2212147.3, \u2212152.6, \u2212156.6\n(PF6). ESI\u2212MS (m/z) [Found (Calcd.)]: 852.3681 (852.3688) [M \u2212\nPF6\u2212]+.\n \u25a0 REFERENCES\n (1) Fahad Ullah, M. Breast cancer: Current perspectives on the disease\n\n\u25a0 ASSOCIATED CONTENT\n* Supporting Information\n s\u0131\n status. 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Three-dimensional cell culture models\nfor metallodrug testing: induction of apoptosis and phenotypic\nreversion of breast cancer cells by the trans-[Ru(PPh3)2(N, N-\ndimethyl-N-thiophenylthioureato-k2O,S)(bipy)]PF6 complex. Inorg.\nChem. Front. 2020, 7, 2909\u22122919.\n (55) Kasibhatla, S.; Amarante-Mendes, G. P.; Finucane, D.; Brunner,\nT.; Bossy-Wetzel, E.; Green, D. R. Acridine orange/ethidium bromide\n(AO/EB) staining to detect apoptosis. Cold Spring Harbor Protoc. 2006,\n2006, No. 4493.\n\n 11774 https://doi.org/10.1021/acs.inorgchem.3c00757\n Inorg. Chem. 2023, 62, 11761\u221211774\n\f", "pages_extracted": 14, "text_length": 112046}