Synthesis, photoluminescence and electrochemiluminescence of ruthenium(II) polypyridyl complexes based on bipyridine derivatives with carbazole moieties

Six complexes (1-6) with the type of [Ru(bpy)₂L]X₂ (1-3: L = L1-L3, X = Cl⁻; 4-6: L = L1-L3, X = PF₆⁻) were synthesized based on 2,2′-bipyridine and three 2,2′-bipyridine derivatives L1, L2 and L3 (L1 = 5,5′-dibromo-2,2′-bipyridine, L2 = 5-bromo-5′-carbazolyl-2,2′-bipyridine, L3 = 5,5′-dicarbazolyl-2,2′-bipyridine). The complexes 1-6 were characterized by ¹H NMR, MS(ESI) and IR spectra, along with the X-ray crystal structure analysis for 1, 5 and 6. Their photophysical properties and electrochemiluminescence (ECL) properties were investigated in detail. In the UV-Vis absorption spectra, all complexes 1-6 show strong intraligand (π → π^∗) transitions and metal-ligand charge transfer (MLCT, dπ (Ru) → π^∗) bands. Upon the excitation wavelengths at ∼508 nm, all complexes 1-6 exhibit typical MLCT emission of ruthenium(II) polypyridyl complexes. The introduction of carbazole moieties improves the MLCT absorption and emission intensity. The ruthenium(II) complexes 1-6 exhibit good electrochemiluminescence (ECL) properties in [Ru(bpy)₂L]²⁺/tri-n-propylamine (TPrA) acetonitrile solution and the complexes with PF₆⁻ showed higher ECL emission intensity than that of the complexes with Cl⁻ based on the same ligands.     

Analysis in vivo of antitumor activity, cytotoxicity and Interaction between plasmid DNA and the cis-dichloro-tetra-amine-ruthenium(III) chloride

Several metallic compounds recognized as potent antitumor agents, have been developed and tested in vivo and in vitro. In this work, we evaluated the toxic, therapeutic, and cytotoxic properties of the cis-dichloro-tetra-amine-ruthenium(III) chloride. Transplanted animals with Sarcoma 180 cells were treated with ruthenium(III) complex and injected i.p., at different time intervals. After the 15th day, tumoral postimplant, the animals were sacrificed and their lungs, kidneys, liver, and tumors were removed and processed for histopathological analysis. Blood samples were also taken for haematological and biochemical analyses. Interaction between the ruthenium complex and the DNA was also investigated. Besides being cytotoxic for the S180 cells, the metallic compound induced tumoral volume reduction and increased survival time of the animals treated. Serum levels of LDH, creatinine, and bilirubin increased, but no serious irreversible histopathological alterations were observed in the analyzed tissues. The compound did not cause anemia, but reduced the number of leukocytes in the treated animals. The absence of viable S180 cells, necrotic cells, and the presence of granulation tissue were observed in tumor tissue of treated animals. The Ru(III) complex, in the presence of the reduction agent, caused plasmid DNA to fragment. These results suggest that cis-RuCl(2)(NH(3))(4)Cl compound is a potent antitumoral drug in vitro and in vivo, which seems to involve binding to DNA molecule.     

DNA-binding studies of mixed-ligand (ethylenediamine)ruthenium(II) complexes

A series of mixed-ligand ruthenium(II) complexes of the type [Ru(en)(2)bpy](2+) (bpy=2,2-bipyridine; 1), [Ru(en)(2)phen](2+) (phen=1,10-phenantroline; 2), [Ru(en)(2)IP](2+) (IP=imidazo[4,5-f][1,10]phenanthroline; 3), and [Ru(en)(2)PIP](2+) (PIP=2-phenylimidazo[4,5-f][1,10]phenanthroline; 4) have been isolated and characterized by UV/VIS, IR, and (1)H-NMR spectral methods. The binding of the complexes with calf thymus DNA has been investigated by absorption, emission spectroscopy, viscosity measurements, DNA melting, and DNA photo-cleavage. The spectroscopic studies together with viscosity measurements and DNA melting studies support that complexes 1 and 2 bind to CT DNA (=calf thymus DNA) by groove mode. Complex 2 binds more avidly to CT DNA than complex 1, complexes 3 and 4 bind to CT DNA by intercalation mode, 4 binds more avidly to CT DNA than 3. Noticeably, the four complexes have been found to be efficient photosensitisers for strand scissions in plasmid DNA.     

DNA-binding property and antitumor activity of a cyclam bridged dinuclear platinum(II) complex

The design of multinuclear Pt(II) complexes with novel structural feature is very important in the search for new anticancer agents. In this work, a dinuclear platinum(II) complex [Pt₂(DTBPA)Cl₂] (II) [DTBPA = (2,2′-(4,11-dimethyl-1,4,8,11-tetraazacyclotetradecane-1,8-diyl)bis(N-(2-(pyridin-2-yl)ethyl)acetamide))] was synthesized via two different methods and characterized by NMR, IR, electrospray mass spectrometry and elemental analysis. It binds to calf thymus DNA (CT-DNA) and induces its conformational changes. Gel electrophoresis data show that complex II leads to a clear decrease of migration rate of the negatively supercoiled band (form I) of supercoiled pUC19 plasmid. The cytotoxic activity of the complex II was tested against human cervical cancer cell line (Hela) and human ovarian carcinoma cell line (Caov-3) and compared with cisplatin. It displays more potent cytotoxicity against Hela cell line than cisplatin at low concentration range.     

A comparative study of PCR product detection and quantitation by electrochemiluminescence and fluorescence

Amplification and detection of target DNA sequences are made possible in a polymerase chain reaction (PCR) by using a mixture of biotinylated and ruthenium(II) trisbipyridal (Ru(bpy)₃²⁺)-end-labelled primers. In this way, biotin for capture and Ru(bpy)₃²⁺ for detection are directly incorporated into the PCR product obviating subsequent probe hybridization. PCR of a bacterial DNA template from Alteromonas species strain JD6.5 using a cocktail of biotin- and Ru(bpy)₃²⁺-labelled primers amplified a 1 kilobase region. Serial dilution of PCR product followed by magnetic separation with Streptavidin (SA)-coated magnetic beads and an electrochemiluminescence (ECL) assay using the semi-automated QPCR System 5000 demonstrated sensitive (pg range) DNA detection. ECL assay of probe hybridization to a human immunodeficiency virus (HIV) sequence also produced pg level sensitivity. Quantitative DNA determination by ECL assay correlated well with visual detection of DNA in electrophoretic gels. However, DNA detection by ECL assay was 10 to 100 times more sensitive than conventional ethidium bromide staining. The combination of DNA-based magnetic separation with ECL assay provides a very sensitive and rapid method of quantitating DNA which, owing to its rapid and facile nature, may have many applications in the research, environmental monitoring, industrial and clinical fields.     

Synthesis and characterization of the ligand based on benzoxazole and its transition metal complexes: DNA-binding and antitumor activity

A new ligand 2-((2-((benzo[d]oxazol-2-yl)methoxy)phenoxy)methyl)benzoxazole (L) and its four transition metal complexes M(NO₃)₂L (M = Cu, Co, Ni, Zn), have been synthesized and investigated. The single crystal structures of the complexes show that all of them have similar molecular structure and the ligand exhibits good coplanarity after coordination with the metal ions. Further investigation of DNA binding indicates that both the ligand L and the complexes can bond to DNA by intercalation mode, and the latter possesses much stronger binding affinity. Antitumor activity of these compounds tested on the four cancer cell lines, follows the order: Cu-L > Ni-L ≈ Co-L > Zn-L ? L, which are thought to be related with their DNA-binding affinity.     

DNA- and RNA-binding and enhanced DNA-photocleavage properties of a ferrocenyl-containing ruthenium(II) complex

The interaction of [Ru(bpy)₂(fip)](PF₆)₂ {bpy = 2,2′-bipyridine, fip = 2-ferrocenyl-1H-imidazo[4,5-f][1,10]-phenanthroline} with calf thymus DNA and yeast tRNA was investigated comparatively by UV-visible absorption and luminescence spectrophotometric titrations, steady-state emission quenching by [Fe(CN)₆]^4 −, ethidium bromide competition experiment, DNA thermal denaturation, viscosity measurements and salt effect studies. The results suggest that the complex binds to the DNA more strongly than to the RNA. The density functional theory calculations were also carried out in order to better understand the nucleic acid binding properties. Agarose gel electrophoresis showed that the complex exhibited enhanced DNA-photocleavage capacity on pUC 18 plasmid DNA under irradiation at 360 nm as compared with a ferrocenyl-free analogous complex.     

DNA conformational changes and cleavage by ruthenium(II) nitrofurylsemicarbazone complexes

Metal complexes that establish interactions with DNA are being studied not only because of their potential use as therapeutic agents but also as tools for biochemistry and molecular biology. Searching for drugs with anti-trypanosome activity, we previously synthesized a series of ruthenium mixed ligand dimethyl sulfoxide complexes of the type [Ru(II)Cl(2)(DMSO)(2)L], where L is 5-nitrofurylsemicarbazone derivatives and DMSO is dimethyl sulfoxide. Though they present the ability to bind DNA, no activity against parasites in cell culture was observed. Considering their potential application as molecular tools we further analyzed the interactions with DNA through an electrophoretic approach. Non covalent withdrawal of superhelicity and a rapid nicking activity upon covalent interaction was observed. Inhibition of both effects was observed in the presence of distamycin suggesting the involvement of the DNA minor groove in the interaction with the nitrofurylsemicarbazone ruthenium complexes. In addition cleavage inhibition by dimethyl sulfoxide suggests an oxidative mechanism of action.     

pH effects on optical and DNA binding properties of a thiophene-containing ruthenium(II) complex

A new ruthenium(II) complex, [Ru(bpy)₂(Htip)]Cl₂ {where bpy = 2,2′-bipyridine and Htip = 2-(thiophen-2-yl)-1H-imidazo[4,5-f][1,10]phenanthroline}, has been synthesized and characterized by ¹H NMR spectroscopy, elemental analysis, and mass spectrometry. The pH effects on UV-Vis absorption and emission spectra of the complex have been studied, and the ground- and excited-state acidity ionization constant values have been derived. The calf thymus (ct) DNA binding properties of the complex have been investigated with UV-Vis absorption and luminescence titrations, steady-state emission quenching by [Fe(CN)₆]⁴⁻, DNA competitive binding with ethidium bromide, DNA melting experiments, and viscosity measurements. The molecular structures and electronic properties of [Ru(bpy)₂(Htip)]²⁺ and deprotonated form [Ru(bpy)₂(tip)]⁺ have also been investigated by means of density functional theory calculations in an effort to understand the DNA binding properties. The results suggest that the complex undergo three-step successive protonation/deprotonation reactions with one of which occurring over physiological pH region, and act as a ct-DNA intercalator with an intrinsic DNA binding constant value on 10⁵ M⁻¹ order of magnitude that is insensitive to pH.     

Catalysis of aerobic olefin oxidation by a ruthenium perhaloporphyrin complex

The perhalogenated porphyrin ruthenium complex (TFPPCl₈)Ru(CO) (TFPPCl₈ = octachlorotetrakis(pentafluorophenyl)porphyrin) catalyzes aerobic oxidation of olefins at room temperature. Cyclohexene is oxidized primarily at the allylic position, and styrene primarily to benzaldehyde, indicating a radical autoxidation mechanism. Reactions are enhanced by visible light. Reaction with _m-chloroperbenzoic acid converts the ruthenium complex to (TFPPCl₈)Ru(O)₂, but such oxo complexes do not appear to participate in catalytic aerobic oxidation.     

pH- and DNA-induced dual molecular light switches based on a novel ruthenium(II) complex

A novel Ru(II) complex, [Ru(bpy)₂(btppz)]Cl₂, where bpy = 2,2′-bipyridine and btppz = benzo[h]tripyrido[3,2-a:2′,3′-c:2″,3″-j]phenazine, has been synthesized and characterized. The pH effects on UV-visible (UV-vis) absorption and emission spectra of the complex have been studied and ground- and excited-state ionization constants of the complex have been derived. The calf thymus DNA (ct-DNA) binding properties of the complex were investigated with UV-vis absorption and luminescence spectrophotometric titrations, steady-state emission quenching by [Fe(CN)₆]⁴⁻, DNA competitive binding with ethidium bromide, DNA melting experiments, reverse salt titrations and viscosity measurements. The complex was demonstrated to act as dual molecular switches: pH-induced “on-off” emission switch with an on-off intensity ratio of ∼54 which is favorably compared with those reported for structurally analogous Ru(II) complexes, and a DNA molecular light switch with a luminescence enhancement factor of 22 as it intercalatively bound to the DNA.     

Synthesis,Structure, DNA‐Binding Properties,and Cytotoxicity of Ruthenium(II) Polypyridyl Complexes

Many ruthenium(II) complexes show high antitumor activities, and the in vitro antitumor activities are usually related to DNA binding. We designed and synthesized two Ru^II polypyridyl complexes, [Ru(dmp)₂(fpp)]²⁺ (dmp=2,9‐dimethyl‐1,10‐phenanthroline; fpp=2‐[3,4‐(difluoromethylenedioxy)phenyl]imidazo[4,5‐f] [1,10]phenanthroline and [Ru(phen)₂(fpp)]²⁺ (phen=1,10‐phenanthroline). The DNA‐binding properties of these complexes have been investigated by spectroscopic titration, DNA melting experiments, viscosity measurements, and photoactivated cleavage. The mechanism studies of photocleavage revealed that singlet oxygen (¹O₂) and superoxide anion radical (O$\rm{{_{2}^{{^\cdot} -}}}$ ) may play an important role in the photocleavage. The cytotoxicity of complexes 1 and 2 have been evaluated by MTT (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl‐2H‐tetrazolium bromide) method; complex 2 shows slightly higher anticancer potency than 1 does against all the cell lines screened.     

Assay of antioxidants by the quenching of the anthracene-sensitized electrochemiluminescence

The kinetic and spectral characteristics (chemiluminescence and fluorescence spectra) of ultraweak luminescence accompanying the electrolysis of a sodium citrate–methanol–dissolved O₂ solution and its application for the determination of antioxidants were examined. The energy transfer-assisted luminescence sensitized by anthracene provides a fast and sensitive assay for the determination of the concentration and kinetic parameters of antioxidants and free radical scavengers.     

DNA binding and oxidative DNA damage induced by a quercetin copper(II) complex: potential mechanism of its antitumor properties

The interaction of a quercetin copper(II) complex with DNA was investigated using UV–vis spectra, fluorescence measurement, viscosity measurement, agarose gel electrophoresis, and thiobarbituric acid reactive substances assay. The results indicate that the quercetin copper(II) complex can promote the cleavage of plasmid DNA, producing single and double DNA strand breaks, and intercalate into the stacked base pairs of DNA. Moreover, the complex can induce oxidative DNA damage involving generation of reactive oxygen species such as H₂O₂ and Cu(I)OOH. In addition, the cytotoxicity experiments carried out with A549 cells confirmed its apoptosis-inducing activity. And we also demonstrate that the levels of survivin protein expression in A549 cells decreased, and that relative activity of caspase-3 increased significantly after treatment with the complex. So our results suggest that the antitumor mechanism of the quercetin copper(II) complex involves not only its oxidative DNA damage with generation of reactive oxygen species but also its specific interaction with DNA. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Synthesis and evaluation of modified chalcone based p53 stabilizing agents

Tumor suppressor protein p53 induces cell cycle arrest and apoptotic cell death in response to various cellular stresses thereby preventing cancer development. Activation and stabilization of p53 through small organic molecules is, therefore, an attractive approach for the treatment of cancers retaining wild-type p53. In this context, a series of nineteen chalcones with various substitution patterns of functional groups including chloro, fluoro, methoxy, nitro, benzyloxy, 4-methyl benzyloxy was prepared using Claisen-Schmidt condensation. The compounds were characterized using NMR, HRMS, IR and melting points. Evaluation of synthesized compounds against human colorectal (HCT116) and breast (CAL-51) cancer cell lines revealed potent antiproliferative activities. Nine compounds displayed GI₅₀ values in the low micromolar to submicromolar range; for example (E)-1-phenyl-3-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (SSE14108) showed GI₅₀ of 0.473 ± 0.043 µM against HCT116 cells. Further analysis of these compounds revealed that (E)-3-(4-chlorophenyl)-1-phenylprop-2-en-1-one (SSE14105) and (E)-3-(4-methoxyphenyl)-1-phenylprop-2-en-1-one (SSE14106) caused rapid (4 and 8-h post-treatment) accumulation of p53 in HCT116 cells similar to its induction by positive control, Nutlin-3. Such activities were absent in 3-(4-methoxyphenyl)propiophenone (SSE14106H2) demonstrating the importance of conjugated ketone for antiproliferative and p53 stabilizing activity of the chalcones. We further evaluated p53 levels in the presence of cycloheximide (CHX) and the results showed that the p53 stabilization was regulated at post-translational level through blockage of its degradation. These chalcones can, therefore, act as fragment leads for further structure optimization to obtain more potent p53 stabilizing agents with enhanced anti-proliferative activities.     

Formation of DNA-adducts and induction of DNA-crosslinks and chromosomal aberrations by the new potent anthracycline antitumor antibiotics: morpholinodaunomycin,cyanomorpholinodaunomycin and cyanomorhpolinoadriamycin

The DNA-interaction of three newly developed semisynthetic anthracyclines with high antitumor potency MoDNM³, CNMoDNM, and CNMoADM, was investigated. When primary rat hepatocytes were incubated with tritium labeled MoDNM and CNMoDNM and their DNA was purified and enzymatically hydrolized, the formation of DNA-adducts could be demonstrated by the HPLC chromatography of the resulting mononucleoside mixtures. The parent compound, daunomycin (DNM), also formed covalent adducts with hepatocyte DNA, but to a lesser extent. These findings correlate well earlier observaitons that MoDNM and CNMoDNM are potent inducers of DNA-repair in primary rat hepatocytes, whereas DNM is only weakly active in this regard. Aklaline elution studies were performed with L 1210 mouse leukemia cells and V79 Chinese hamster fibroblasts. The cyanomorpholinyl derivatives showed dose-dependant DNA crosslinking activities in both cell lines at concentrations 5 nMol/l. The formation of crosslinks began a few minutes after treatment of the cells and reached a maximum after 1 hr. In contrast, MoDNM, at concentrations of up to 10 Mol/l, had only a limited capacity to induce single strand breaks in L 1210 cells but did not induce DNA-crosslinks. In addition, chromosomal aberrations (chromatid breaks and translocations) were induced by the treatment of Friend and L 1210 leukemia cells with CNMoADM at concentrations between 0.07–0.6 n Mol/l. At higher doses, chromosome clumping was observed. These results indicate that the high capacity of MoDNM, CNMoDNM and CNMoADM to induce DNA repair in primary rat hepatocytes is due to the formation of covalent adducts with DNA. The cyanomorpholino compounds have alkylating capacities also in cell lines such as L 1210 and V79, whereas MoDNM requires rat hepatocytes for activation. The ready formation of DNA crosslinks and chromosomal aberrations could be responsible for the high cytotoxicity of these compounds.Abbreviations ADM adriamycin - CNMoADM cyanomorpholinoadriamycin - CNMoDNM cyanomorpholinodaunomycin - DNM daunomycin - FLC Friend leukemia cells - (G³H) generally tritium labeled - HPLC high performance liquid chromatography - MoDNM morpholinodaunomycin - Rf retention factor - (Mo³H) tritium labelled at morpholinyl site - Rad radiation unit - RT retention time - SDS sodium dodecylsulphate - Tris tris (hydroxymethyl)aminomethan     

Novel thiosemicarbazone derivatives as potential antitumor agents: Synthesis, physicochemical and structural properties, DNA interactions and antiproliferative activity

The paper describes synthesis of several novel thiosemicarbazone derivatives. Furthermore, crystal and molecular structure of 4-diethylamino-salicylaldehyde 4-phenylthiosemicarbazone revealed planarity of conjugated aromatic system, which suggested the possibility of DNA binding by intercalation, especially for here studied naphthalene derivatives. However, here presented DNA binding studies excluded this mode of action. Physicochemical and structural properties of novel derivatives were compared with previously studied analogues, taken as reference compounds, revealing distinctive differences. In addition, novel thiosemicarbazone derivatives (1, 2 and 5–8) clearly display stronger antiproliferative activity on five tumor cell lines than the reference compounds 3 and 4, which supports their further investigation as potential antitumor agents.     

A sensitive method for the determination of levamisole in serum by electrochemiluminescence

A novel method was developed for the determination of levamisole by electrochemiluminescence. The method was based on electrochemiluminescence signal enhancement produced by Ru(bpy)₃²⁺, which reacted with the tertiary amine group of levamisole on a platinum electrode in 12 mmol/L borate buffer (pH 9). A linear relationship between the luminous intensity and concentration of levamisole in the range 0–1 × 10^–7 mol/L was obtained and the detection limit was 1.76 × 10^–11 mol/L. The method is sensitive, selective, simple and convenient. The method has been successfully applied to the analysis of levamisole in serum. Copyright © 2013 John Wiley & Sons, Ltd.     

Formation and molecular structure of novel Ru(III) dimers of a symmetric antitumor drug analogue

The symmetrical anionic and neutral dimers [H(TMSO)₂]₂trans-[{RuCl₄(TMSO)}₂](μ-pyz) (1), and mer-[{RuCl₃(TMSO)₂}₂](μ-pyz) (2) were isolated by the reaction of [H(TMSO)] trans-[RuCl₄(TMSO)₂] and mer-[RuCl₃(TMSO)₃] with heterocyclic nitrogen donor ligand pyrazine (pyz) at room temperature. These complexes can be regarded as unprecedented examples in the general Creutz-Taube family of ruthenium dimers. Each ruthenium center in 1 and 2 has a coordination environment akin to that of known anionic and neutral monomeric Ru(III) complexes. Crystals of 1 · acetone are orange, needle like, space group , a=10.419(3) Å, b=10.539(3) Å, c=12.595(5) Å, α=69.837(16)°, β=69.968(15)°, γ=74.330(15)° and crystals of 2 · 4TMSO are orange prisms, trigonal, space group , a=33.971(5) Å, b=33.971(5) Å, c=12.210(2) Å, α=90°, β=90° and γ=120°.     

Synthesis, DNA-binding and photocleavage studies of ruthenium complexes [Ru(bpy)2(mitatp)] and [Ru(bpy)2(nitatp)]

Two new ruthenium complexes [Ru(bpy)₂(mitatp)](ClO₄)₂1 and [Ru(bpy)₂(nitatp)](ClO₄)₂2 (bpy = 2,2′-bipyridine, mitatp = 5-methoxy-isatino[1,2-b]-1,4,8,9-tetraazatriphenylene, nitatp = 5-nitro-isatino[1,2-b]-1,4,8,9-tetraazatriphenylene) have been synthesized and characterized by elemental analysis, ¹H NMR, mass spectrometry and cyclic voltammetry. Spectroscopic and viscosity measurements proved that the two Ru(II) complexes intercalate DNA with larger binding constants than that of [Ru(bpy)₂(dppz)]²⁺ (dppz = dipyrido[3,2-a:2′,3′-c]phenazine) and possess the excited lifetime of microsecond scale upon binding to DNA. Both complexes can efficiently photocleave pBR322 DNA in vitro under irradiation. Singlet oxygen (¹O₂) was proved to contribute to the DNA photocleavage process, the ¹O₂ quantum yields was determined to be 0.43 and 0.36 for 1 and 2, respectively. Moreover, a photoinduced electron transfer mechanism was also found to be involved in the DNA cleavage process.