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Metal-based drugs represent a rich source of chemical substances of potential interest for the treatment of COVID-19. To this end, we have developed a small but representative panel of nine metal compounds, including both synthesized and commercially available complexes, suitable for medical application and tested them in vitro against the SARS-CoV-2 virus. The screening revealed that three compounds from the panel, i.e., the organogold(III) compound Aubipyc, the ruthenium(III) complex KP1019, and antimony trichloride (SbCl 3 ), are endowed with notable antiviral properties and an acceptable cytotoxicity profile. These initial findings prompted us to perform a computational study to unveil the likely molecular basis of their antiviral actions. Calculations evidenced that the metalation of nucleophile sites in SARS-CoV-2 proteins or nucleobase strands, induced by Aubipyc, SbCl 3 , and KP1019, is likely to occur. Remarkably, we found that only the deprotonated forms of Cys and Sec residues can react favorably with these metallodrugs. The mechanistic implications of these findings are discussed.

The known ruthenium complex [Ru(tpy)(bpy)(Hmte)](PF 6 ) 2 ([1](PF 6 ) 2 , where tpy = 2,2':6',2″-terpyridine, bpy = 2,2'-bipyridine, Hmte = 2-(methylthio)ethanol) is photosubstitutionally active but non-toxic to cancer cells even upon light irradiation. In this work, the two analogs complexes [Ru(tpy)(NN)(Hmte)](PF 6 ) 2 , where NN = 3,3'-biisoquinoline (i-biq, [2](PF 6 ) 2 ) and di(isoquinolin-3-yl)amine (i-Hdiqa, [3](PF 6 ) 2 ), were synthesized and their photochemistry and phototoxicity evaluated to assess their suitability as photoactivated chemotherapy (PACT) agents. The increase of the aromatic surface of [2](PF 6 ) 2 and [3](PF 6 ) 2 , compared to [1](PF 6 ) 2 , leads to higher lipophilicity and higher cellular uptake for the former complexes. Such improved uptake is directly correlated to the cytotoxicity of these compounds in the dark: while [2](PF 6 ) 2 and [3](PF 6 ) 2 showed low EC 50 values in human cancer cells, [1](PF 6 ) 2 is not cytotoxic due to poor cellular uptake. While stable in the dark, all complexes substituted the protecting thioether ligand upon light irradiation (520 nm), with the highest photosubstitution quantum yield found for [3](PF 6 ) 2 (Φ [3]  = 0.070). Compounds [2](PF 6 ) 2 and [3](PF 6 ) 2 were found both more cytotoxic after light activation than in the dark, with a photo index of 4. Considering the very low singlet oxygen quantum yields of these compounds, and the lack of cytotoxicity of the photoreleased Hmte thioether ligand, it can be concluded that the toxicity observed after light activation is due to the photoreleased aqua complexes [Ru(tpy)(NN)(OH 2 )] 2+ , and thus that [2](PF 6 ) 2 and [3](PF 6 ) 2 are promising PACT candidates., (© 2021. The Author(s).)

The clinical success of cisplatin, carboplatin, and oxaliplatin has sparked the interest of medicinal inorganic chemistry to synthesize and study compounds with non-platinum metal centers. Despite Ru(II)-polypyridyl complexes being widely studied and well established for their antitumor properties, there are not enough in vivo studies to establish the potentiality of this type of compound. Therefore, we report to the best of our knowledge the first in vivo study of Ru(II)-polypyridyl complexes against breast cancer with promising results. In order to conduct our study, we used MCF7 zebrafish xenografts and ruthenium complexes [Ru(bipy) 2 (C 12 H 8 N 6 -N,N)][CF 3 SO 3 ] 2 Ru1 and [{Ru(bipy) 2 } 2 (μ-C 12 H 8 N 6 -N,N)][CF 3 SO 3 ] 4 Ru2 , which were recently developed by our group. Ru1 and Ru2 reduced the tumor size by an average of 30% without causing significant signs of lethality when administered at low doses of 1.25 mg·L -1 . Moreover, the in vitro selectivity results were confirmed in vivo against MCF7 breast cancer cells. Surprisingly, this work suggests that both the mono- and the dinuclear Ru(II)-polypyridyl compounds have in vivo potential against breast cancer, since there were no significant differences between both treatments, highlighting Ru1 and Ru2 as promising chemotherapy agents in breast cancer therapy.

Flavonoids are a class of natural polyphenolic compounds sharing a common 2-phenyl-3,4-dihydro-2 H -1-benzopyran (flavan) backbone. Typically known for their antioxidant activity, flavonoids are also being investigated regarding antitumour and antimicrobial properties. In this review, we report on the complexation of both natural and synthetic flavonoids with ruthenium as a strategy to modulate the biological activity. The ruthenoflavonoid complexes are divided into three subclasses, according to their most prominent bioactivity: antitumour, antimicrobial, and protection of the cardiovascular system. Whenever possible the activity of the ruthenoflavonoids is compared with that of commercial drugs for a critical assessment of the feasibility of using them in future clinical applications.

This review concentrates on recent developments in ruthenium Schiff bases, whose steric and electronic characteristics can be manipulated easily by selecting suitable condensing aldehydes or ketones and primary amines, and their metal complexes. Ruthenium metal-based complexes and Schiff base ligands are rapidly becoming conventionally considered for biological applications (antioxidant, anticancer, antimicrobial), in catalysis, in functional materials, in sensors, and as pigments for dyes. Ruthenium complexes exhibit a broad variety of activities concerning simple Schiff base ligands. This may be due to the octahedral bonding of both Ru(II) and Ru(III) complexes, which acquire an extended reservoir of a three-dimensional framework, providing the potential for an elevated degree of site selectivity for binding to their biological targets. This review provides an overview of this field, and intends to highlight both ligand design and synthetic methodology development, as well as significant applications of these metal complexes. In this review, we summarize our work on the development of ruthenium complexes, which was performed over the last few years.

Cisplatin and derivatives are highly effective in the treatment of a wide range of cancer types; however, these metallodrugs display low selectivity, leading to severe side effects. Additionally, their administration often results in the development of chemoresistance, which ultimately results in therapeutic failure. This scenario triggered the study of other transition metals with innovative pharmacological profiles as alternatives to platinum, ruthenium- (e.g., KP1339 and NAMI-A) and gold-based (e.g., Auranofin) complexes being among the most advanced in terms of clinical evaluation. Concerning the importance of improving the in vivo selectivity of metal complexes and the current relevance of ruthenium and gold metals, this review article aims to survey the main research efforts made in the past few years toward the design and biological evaluation of target-specific ruthenium and gold complexes. Herein, we give an overview of the inorganic and organometallic molecules conjugated to different biomolecules for targeting membrane proteins, namely cell adhesion molecules, G-protein coupled receptors, and growth factor receptors. Complexes that recognize the progesterone receptors or other targets involved in metabolic pathways such as glucose transporters are discussed as well. Finally, we describe some complexes aimed at recognizing cell organelles or compartments, mitochondria being the most explored. The few complexes addressing targeted gene therapy are also presented and discussed.

Procoagulant snake venoms have been inhibited by the ruthenium containing compounds CORM-2 and RuCl 3 separately, presumably by interacting with critical histidine or other sulfur-containing amino acids on key venom enzymes. However, combinations of these and other platinoid containing compounds could potentially increase, decrease or not affect the procoagulant enzyme function of venom. Thus, the purpose of this investigation was to determine if formulations of platinoid compounds could inhibit venom procoagulant activity and if the formulated compounds interacted to enhance inhibition. Using a human plasma coagulation kinetic model to assess venom activity, six diverse venoms were exposed to various combinations and concentrations of CORM-2, CORM-3, RuCl 3 and carboplatin (a platinum containing compound), with changes in venom activity determined with thrombelastography. The combinations of CORM-2 or CORM-3 with RuCl 3 were found to enhance inhibition significantly, but not in all venoms nor to the same extent. In sharp contrast, carboplatin-antagonized CORM-2 mediated the inhibition of venom activity. These preliminary results support the concept that platinoid compounds may inhibit venom enzymatic activity at the same or different molecular sites and may antagonize inhibition at the same or different sites. Further investigation is warranted to determine if platinoid formulations may serve as potential antivenoms.

Metal complexes are currently potential therapeutic compounds. The acquisition of resistance by cancer cells or the effective elimination of cancer-affected cells necessitates a constant search for chemical compounds with specific biological activities. One alternative option is the transition metal complexes having potential as antitumor agents. Here, we present the current knowledge about the application of transition metal complexes bearing nickel(II), cobalt(II), copper(II), ruthenium(III), and ruthenium(IV). The cytotoxic properties of the above complexes causing apoptosis, autophagy, DNA damage, and cell cycle inhibition are described in this review.

Anticancer potential of ruthenium complexes has been widely investigated, but safety evaluation studies are still scarce. Despite of ruthenium-based anticancer agents are known to cause fewer side effects compared to other metal-based drugs, these compounds are not fully free of toxicity, causing mainly nephrotoxicity. Based on the promising results from antitumor activity of the complexes [Ru(L-Met)(bipy)(dppb)]PF 6 (RuMet) and [Ru(L-Trp)(bipy)(dppb)]PF 6 (RuTrp), for the first time we investigated the toxicity profile of these complexes in rodent and zebrafish models. The acute oral toxicity was evaluated in Swiss mice. The mutagenic and genotoxic potential was determined by a combination of Micronucleus (MN) and Comet assay protocols, after exposure of Swiss mice to RuMet and RuTrp in therapeutic doses. Zebrafish embryos were exposed to these complexes, and their development observed up to 96 h post-fertilization. RuMet and RuTrp complexes showed low acute oral toxicity. Recorded behavioral changes were not recorded, nor were macroscopic morphological changes or structural modifications in the liver and kidneys. These complexes did not cause genetic toxicity, presenting a lack of micronuclei formation and low DNA damage induction in the cells from Swiss mice. In contradiction, cisplatin treatment exhibited high mutagenicity and genotoxicity. RuMet and RuTrp showed low toxicity in the embryo development of zebrafish. The RuMet and RuTrp complexes demonstrated low toxicity in the two study models, an interesting property in preclinical studies for novel anticancer agents., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Ruthenium-based compounds represent a class of potential antineoplastic drugs. Recently, we designed, synthesized, and identified the Ru(II)-thymine complex [Ru(PPh₃)₂(Thy)(bipy)]PF₆ (where PPh = triphenylphosphine, Thy = thymine and bipy = 2,2'-bipyridine) as a potent cytotoxic agent with the ability to bind to DNA and human and bovine serum albumins. In this study, the underlying cytotoxic mechanism of the [Ru(PPh₃)₂(Thy)(bipy)]PF₆ complex was assessed. This complex displayed potent cytotoxicity in different cancer cell lines; the morphology that is associated with apoptotic cell death, increased internucleosomal DNA fragmentation without cell membrane permeability, loss of the mitochondrial transmembrane potential, increased phosphatidylserine externalization, and caspase-3 activation were observed in human promyelocytic leukemia HL-60 cells that were treated with the complex. Moreover, pretreatment of HL-60 cells with Z-VAD(OMe)-FMK, a pan-caspase inhibitor, partially reduced the apoptosis that was induced by the complex, indicating that the apoptotic cell death occurred through a caspase-mediated pathway. In conclusion, the [Ru(PPh₃)₂(Thy)(bipy)]PF₆ complex displays potent cytotoxicity to different cancer cells and induces caspase-mediated apoptosis in HL-60 cells.

RuP x nanoparticles (NPs) encapsulated in uniform N,P-codoped hollow carbon nanospheres (RuP x @NPC) have been synthesized through a facile route in which aniline-pyrrole copolymer nanospheres are used to disperse Ru ions followed by a gas phosphorization process. The as-prepared RuP x @NPC exhibits a uniform core-shell hollow nanospherical structure with RuP x NPs as the core and N,P-codoped carbon (NPC) as the shell. This strategy integrates many advantages of hollow nanostructures, which provide a conductive substrate and the doping of a nonmetal element. At high temperatures, the obtained thin NPC shell can not only protect the highly active phase of RuP x NPs from aggregation and corrosion in the electrolyte but also allows variation in the electronic structures to improve the charge-transfer rate greatly by N,P codoping. The optimized RuP x @NPC sample at 900 °C exhibits a Pt-like performance for the hydrogen evolution reaction (HER) and long-term durability in acidic, alkaline, and neutral solutions. The reaction requires a small overpotential of only 51, 74, and 110 mV at 10 mA cm -2 in 0.5 m H 2 SO 4 , 1.0 m KOH, and 1.0 m phosphate-buffered saline, respectively. This work provides a new way to design unique phosphide-doped carbon heterostructures through an inorganic-organic hybrid method as excellent electrocatalysts for HER., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Peritoneal carcinomatosis is considered as a potentially lethal clinical condition, and the therapeutic options are limited. The antitumor effectiveness of the [Ru(l-Met)(bipy)(dppb)]PF 6 (1) and the [Ru(l-Trp)(bipy)(dppb)]PF 6 (2) complexes were evaluated in the peritoneal carcinomatosis model, Ehrlich ascites carcinoma-bearing Swiss mice. This is the first study that evaluated the effect of Ru(II)/amino acid complexes for antitumor activity in vivo. Complexes 1 and 2 (2 and 6 mg kg -1 ) showed tumor growth inhibition ranging from moderate to high. The mean survival time of animal groups treated with complexes 1 and 2 was higher than in the negative and vehicle control groups. The induction of Ehrlich ascites carcinoma in mice led to alterations in hematological and biochemical parameters, and not the treatment with complexes 1 and 2. The treatment of Ehrlich ascites carcinoma-bearing mice with complexes 1 and 2 increased the number of Annexin V positive cells and cleaved caspase-3 levels and induced changes in the cell morphology and in the cell cycle phases by induction of sub-G1 and G0/G1 cell cycle arrest. In addition, these complexes reduce angiogenesis induced by Ehrlich ascites carcinoma cells in chick embryo chorioallantoic membrane model. The treatment with the LAT1 inhibitor decreased the sensitivity of the Ehrlich ascites carcinoma cells to complexes 1 and 2 in vitro-which suggests that the LAT1 could be related to the mechanism of action of amino acid/ruthenium(II) complexes, consequently decreasing the glucose uptake. Therefore, these complexes could be used to reduce tumor growth and increase mean survival time with less toxicity than cisplatin. Besides, these complexes induce apoptosis by combination of different mechanism of action.

Photodynamic therapy (PDT) is a treatment that uses photosensitizing agents to kill cancer cells. Scientific community has been eager for decades to design an efficient PDT drug. Under such purview, the current report deals with the computational photodynamic behavior of ruthenium(II) nitrosyl complex containing N, N'-salicyldehyde-ethylenediimine (SalenH 2 ), the synthesis and X-ray crystallography of which is already known [Ref. 38,39]. Gaussian 09W software package was employed to carry out the density functional (DFT) studies. DFT calculations with Becke-3-Lee-Yang-Parr (B3LYP)/Los Alamos National Laboratory 2 Double Z (LanL2DZ) specified for Ru atom and B3LYP/6-31G(d,p) combination for all other atoms were used using effective core potential method. Both, the ground and excited states of the complex were evolved. Some known photosensitizers were compared with the target complex. Pthalocyanine and porphyrin derivatives were the compounds selected for the respective comparative study. It is suggested that effective photoactivity was found due to the presence of ruthenium core in the model complex. In addition to the evaluation of theoretical aspects in vitro anticancer aspects against COLO-205 human cancer cells have also been carried out with regard to the complex. More emphasis was laid to extrapolate DFT to depict the chemical power of the target compound to release nitric oxide. A promising visible light triggered nitric oxide releasing power of the compound has been inferred. In vitro antiproliferative studies of [RuCl 3 (PPh 3 ) 3 ] and [Ru(NO)(Salen)(Cl)] have revealed the model complex as an excellent anticancer agent. From IC 50 values of 40.031mg/mL in former and of 9.74mg/mL in latter, it is established that latter bears more anticancer potentiality. From overall study the DFT based structural elucidation and the efficiency of NO, Ru and Salen co-ligands has shown promising drug delivery property and a good candidacy for both chemotherapy as well as light therapy., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The ruthenium(II) compounds cis-[Ru(bpy) 2 (4-bzpy)(CO)](PF 6 ) 2 (I) and cis-[Ru(bpy) 2 (4-bzpy)(Cl)](PF 6 ) (II) (4-bzpy=4-benzoylpyridine, bpy=2,2'-bipyridine) were synthesized and characterized by spectroscopic and electrochemical techniques. The crystal structure of II was determined by X-ray diffraction. The photochemical behavior of I in aqueous solution shows that irradiation with ultraviolet light (365nm) releases both CO and 4-bzpy leading to the formation of the cis-[Ru(bpy) 2 (H 2 O) 2 ] 2+ ion as identified by NMR and electronic spectroscopy. Carbon monoxide release was confirmed with the myoglobin method and by gas chromatographic analysis of the headspace. CO release was not observed when aqueous I was irradiated with blue light (453nm). Changes in the electronic and 1 H NMR spectra indicate that I undergoes photoaquation of 4-bzpy to form cis-[Ru(bpy) 2 (CO)(H 2 O)] 2+ . Blue light irradiation of aqueous II released the coordinated 4-bzpy to give the cis-[Ru(bpy) 2 (H 2 O)(Cl)] 2+ ion. When the latter reaction was carried out in the presence of the nucleobase guanine, Ru-guanine adducts were formed, indicating that the metal containing photoproduct may also participate in biologically relevant reactions. The photochemical behavior of I indicates that it can release either CO or 4-bzpy depending on the wavelength chosen, a feature that may have therapeutic application., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Organometallic metal(arene) anticancer agents require ligand exchange for their anticancer activity and this is generally believed to confer low selectivity for potential cellular targets. However, using an integrated proteomics-based target-response profiling approach as a potent hypothesis-generating procedure, we found an unexpected target selectivity of a ruthenium(arene) pyridinecarbothioamide (plecstatin) for plectin, a scaffold protein and cytolinker, which was validated in a plectin knock-out model in vitro. Plectin targeting shows potential as a strategy to inhibit tumor invasiveness as shown in cultured tumor spheroids while oral administration of plecstatin-1 to mice reduces tumor growth more efficiently in the invasive B16 melanoma than in the CT26 colon tumor model., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Ruthenium is seldom mentioned in microbiology texts, due to the fact that this metal has no known, essential roles in biological systems, nor is it generally considered toxic. Since the fortuitous discovery of cisplatin, first as an antimicrobial agent and then later employed widely as an anticancer agent, complexes of other platinum group metals, such as ruthenium, have attracted interest for their medicinal properties. Here, we review at length how ruthenium complexes have been investigated as potential antimicrobial, antiparasitic and chemotherapeutic agents, in addition to their long and well-established roles as biological stains and inhibitors of calcium channels. Ruthenium complexes are also employed in a surprising number of biotechnological roles. It is in the employment of ruthenium complexes as antimicrobial agents and alternatives or adjuvants to more traditional antibiotics, that we expect to see the most striking developments in the future. Such novel contributions from organometallic chemistry are undoubtedly sorely needed to address the antimicrobial resistance crisis and the slow appearance on the market of new antibiotics., (© 2017 Elsevier Ltd. All rights reserved.)

Ruthenium-based complexes have emerged as promising anticancer, especially antimetastatic agents. Among them, NAMI-A (trans-[Ru(III)Cl 4 (DMSO)(Im)][ImH], Im = imidazole, DMSO = dimethyl sulfoxide) was well studied. In this study, we studied the antimetastatic activities of two novel NAMI-A derivatives containing pyridine, G26b and G94a, using cultured cells and tumor-bearing mice. Same to NAMI-A, these two complexes displayed little direct cytotoxicity to the cancer cells in vitro and in vivo, but they, especially G26b, significantly reduced the occurrence and development of lung metastases in mice bearing the 4T1 mammary carcinoma. In vitro, these two complexes displayed significant suppressive effect on invasion and migration of cells and tube formation of human umbilical vein endothelial cell, to the same extent of NAMI-A. The transcription of important molecules involved in metastasis, matrix metalloproteinase 2 and 9 (MMP-2 and -9), and vascular endothelial growth factor, was suppressed by the two complexes, as well as NAMI-A. Plasma atomic emission spectrometer showed G26b had a longer Ru-elimination time in lung, which may be a reason for better antimetastatic effect of G26b than NAMI-A. Our results have demonstrated that G26b is a more effective antimetastatic agent than NAMI-A., (© 2016 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

2,5-Diphenyl-3,4-dicyanothiophene (1) and phthalonitrile (2) were mixed and treated with ruthenium (III) trichloride, 4-methylpyridine, and 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) in 2-ethoxyethanol at 135°C, to produce low-symmetrical tetraazaporphyrins (TAPs) (3), (4), (5), and (6) with one to three thiophene rings. Two thiophene-annelated tetraazaporphyrins were isolated as opposite and adjacent isomers 4 and 5. The structure of 3 was determined by X-ray crystallography, showing that the thiophene ring linked at the 3,4-positions on the tetraazaporphyrin scaffold deviates from the mean plane of the four central pyrrole nitrogen atoms (N1-N3-N5-N7). Optical and electrochemical properties of the products were examined by UV-vis and magnetic circular dichroism (MCD) spectroscopy, together with cyclic voltammetry. In the (1)H NMR spectra, the signals of 4-methylpyridine coordinating to the central ruthenium atom appeared at a higher magnetic field than those of uncoordinated 4-methylpyridine itself due to the shielding effect of the TAP ring. Increasing the number of fused thiophene rings resulted in 1) lower magnetic field shifts of the signals of axially coordinated 4-methylpyridine in the (1)H NMR spectra, 2) lower energy shifts of the Q band absorption in the UV-vis spectra, and 3) decreasing (cathodic shift) of the first oxidation potentials. The structures of simplified model compounds were optimized using the DFT method with the Gaussian 09 program at the B3LYP/LANL2DZ level for the Ru atom and the B3LYP/6-31G (d, p) level for the C, H, N, and S atoms. The optimized structures were utilized to calculate the NMR shielding constants, the HOMO and LUMO orbital energies, and the electronic absorption spectra., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Toward the development of visible-light-driven molecular rotary motors, an overcrowded alkene-based ligand and the corresponding ruthenium(II) complex is presented. In our design, a 4,5-diazafluorenyl coordination motif is directly integrated into the motor function. The photochemical and thermal isomerization behavior has been studied by UV/Vis and NMR spectroscopy. Upon coordination to a Ru(II) bipyridine complex, the photoisomerization process can be driven by visible (λmax = 450 nm) instead of UV light and furthermore, a large increase of the speed of rotation is noted. DFT calculations point to a contraction of the diazafluorenyl lower half upon metal-coordination resulting in reduced steric hindrance in the "fjord region" of the molecule. Consequently, it is shown that metal-ligand interactions can play an important role in the adjustment of both photophysical and thermodynamic properties of molecular motors., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The structural changes and electrochemical behavior of RuO2 are investigated by using in situ XRD, X-ray absorption spectroscopy, and electrochemical techniques to understand the electrochemical reaction mechanism of this metal oxide anode material. Intermediate phase-assisted transformation of RuO2 to LiRuO2 takes place at the start of discharge. Upon further lithiation, LiRuO2 formed by intercalation decomposes to nanosized Ru metal and Li2 O by a conversion reaction. A reversible capacity in addition to its theoretical capacity is observed on discharging below 0.5 V during which no redox activity involving Ru is observed. TEM, X-ray photoelectron spectroscopy, and the galvanostatic intermittent titration technique are used to probe this additional capacity. The results show that the additional capacity is a result of Li storage in the grain boundary between nanosized Ru metal and Li2 O. Findings of this study provide a better understanding of the quantitative share of capacity by a unique combination of intercalation, conversion, and interfacial Li storage in a RuO2 anode., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The rational design of metal-based complexes is an effective strategy for the discovery of potent sensitizers for use in cancer radiotherapy. In this study, we synthesized three ruthenium complexes containing bis-benzimidazole derivatives: Ru(bbp)Cl3 (1), [Ru(bbp)2 ]Cl2 (2 a) (in which bbp=2,6-bis(benzimidazol-1-yl)pyridine), and [Ru(bbp)2]Cl2 (2 b) (where bbp=2,6-bis-(6-nitrobenzimidazol-2-yl)pyridine). We evaluated their radiosensitization capacities in vitro and mechanisms of action. Complex 2 b was found to be particularly effective in sensitizing human melanoma A375 cells toward radiation, with a sensitivity enhancement ratio of 2.4. Along with this potency, complex 2 b exhibited a high degree of selectivity between human cancer and normal cells. Mechanistic studies revealed that 2 b promotes radiation-induced accumulation of intracellular reactive oxygen species (ROS) by reacting with cellular glutathione (GSH) and then causing DNA stand breaks. The subsequent DNA damage induces phosphorylation of p53 (p-p53) and upregulates the expression levels of p21, which inhibits the expression of cyclin-B, leading to G2M arrest. Moreover, p-p53 activates caspases-3 and -8, triggers cleavage of poly(ADP-ribose) polymerase (PARP), finally resulting in apoptosis. Taken together, the results of this study provide a strategy for the design of ruthenium-based radiosensitizers for use in cancer therapy., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The reaction of Na[RuCl4(SO(CH3)2)2], 1, with one equivalent of FcCONHCH2C6H4N (Fc=FeC10H9), L1, FcCOOCH2CH2C3H3N2, L2, FcCOOC6H4N, L3, afforded the dinuclear species, Na[FcCONHCH2C6H4N[RuCl4(SO(CH3)2)]], RuL1, Na[FcCOOCH2CH2C3H3N2[RuCl4(SO(CH3)2)]], RuL2, Na[FcCOOC6H4N(RuCl4(SO(CH3)2))], RuL3, respectively, yielding, in each case, a ferrocene moiety bridged to a ruthenium center. The complexes were characterized by NMR, IR, and XRD (X-ray diffraction). The sulfoxide ligands are bonded to the metal through the sulfur atom. The complexes were evaluated for their biological activity with pBluescript DNA plasmid, and the protein BSA (bovine serum albumin). These reactions were monitored by XAS (X-ray absorption spectroscopy), EXAFS (extended X-ray Absorption Fine Structure), NMR, UV/visible, emission spectroscopy, and gel electrophoresis. Donor atoms from the biomolecules substitute for the chloride ligands in the parent complexes., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Magnetic γ-Fe2 O3 nanocrystallites encapsulated by hydroxyapatite (HAP), HAP@γ-Fe2 O3 , were prepared followed by cation exchange of Ca(2+) on the external HAP surface with Ru(3+) to give the γ-Fe2 O3 @HAP-Ru catalyst. The structure of the as-prepared catalyst was characterized, and its catalytic activity was studied in the aerobic oxidation of 5-hydroxymethylfurfural (HMF). γ-Fe2 O3 @HAP-Ru showed a high catalytic activity for the aerobic oxidation of HMF into 2,5-diformylfuran (DFF). A high DFF yield of 89.1 % with an HMF conversion of 100% was obtained after 4 h at 90 °C. Importantly, the synthesis of DFF from fructose was realized by two consecutive steps. The dehydration of fructose in the presence of a magnetic acid catalyst (Fe3 O4 @SiO2-SO3 H) produced HMF in a yield of 90.1%. Then the Fe3 O4 @SiO2 SO3 H catalyst was removed from the reaction solution with a permanent magnet, and HMF in the resulting solution was further oxidized to DFF with a yield of 79.1% based on fructose. The synthesis of DFF from fructose by two steps avoids the tedious separation of the intermediate HMF, which saves time and energy., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A series of [{(terpy)(bpy)Ru}(μ-O){Ru(bpy)(terpy)}](n+) ([RuORu](n+), terpy=2,2';6',2''-terpyridine, bpy=2,2'-bipyridine) was systematically synthesized and characterized in three distinct redox states (n=3, 4, and 5 for Ru(II,III)2, Ru(III,III)2, and Ru(III,IV)2, respectively). The crystal structures of [RuORu](n+) (n=3, 4, 5) in all three redox states were successfully determined. X-ray crystallography showed that the Ru-O distances and the Ru-O-Ru angles are mainly regulated by the oxidation states of the ruthenium centers. X-ray crystallography and ESR spectra clearly revealed the detailed electronic structures of two mixed-valence complexes, [Ru(III)ORu(IV)](5+) and [Ru(II)ORu(III)](3+), in which each unpaired electron is completely delocalized across the oxo-bridged dinuclear core. These findings allow us to understand the systematic changes in structure and electronic state that accompany the changes in the redox state., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Evolution can increase the complexity of matter by self-organization into helical architectures, the best example being the DNA double helix. One common aspect, apparently shared by most of these architectures, is the presence of covalent bonds within the helix backbone. Here, we report the unprecedented crystal structures of a metal complex that self-organizes into a continuous double helical structure, assembled by non-covalent building blocks. Built up solely by weak stacking interactions, this alternating tread stairs-like double helical assembly mimics the DNA double helix structure. Starting from a racemic mixture in aqueous solution, the ruthenium(II) polypyridyl complex forms two polymorphic structures of a left-handed double helical assembly of only the Λ-enantiomer. The stacking of the helices is different in both polymorphs: a crossed woodpile structure versus a parallel columnar stacking., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Chelated amino acid ester enolates are excellent nucleophiles for ruthenium-catalyzed allylic alkylations. Although [Cp*Ru(MeCN)3 ]PF6 was found to be the most reactive catalyst investigated, with the resulting allyl complexes reacting at temperatures as low as -78 °C, unfortunately the process took place with only moderate regio- and diastereoselectivity. In contrast, [(p-cymene)RuCl2 ]2 allowed allylations to be performed with a high degree of regioretention. Secondary allyl carboxylates with a terminal double bond were found to be the most reactive substrates, giving rise to the branched amino acids with perfect regioretention and chirality transfer. In this case, no isomerization of the Ru-allyl complex formed in situ was observed, in contrast to the analogues palladium complexes. This isomerization-free protocol can also be used for the synthesis of (Z)-configured γ,δ-unsaturated amino acid derivatives, starting from (Z)-allylic substrates. Here, the more reactive phosphates were found to be superior to the carboxylates, providing the required amino acids in almost quantitative yield with perfect regio- and stereoretention. Therefore, the Ru-catalyzed allylation reactions are well positioned to overcome the drawbacks of Pd-catalyzed processes., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A series of pyridine-based derivatives of the clinically successful Ru(III)-based complexes indazolium [trans-RuCl4(1H-indazole)2] (KP1019) and sodium [trans-RuCl4(1H-indazole)2] (KP1339) have been synthesized to probe the effect of hydrophobic interactions with human serum albumin (hsA) on anticancer activity. The solution behavior and protein interactions of the new compounds were characterized by using electron paramagnetic resonance (EPR) and UV/Vis spectroscopy. These studies have revealed that incorporation of hydrophobic substituents at the 4'-position of the axial pyridine ligand stabilizes non-coordinate interactions with hsA. As a consequence, direct coordination to the protein is inhibited, which is expected to increase the bioavailability of the complexes, thus potentially leading to improved anticancer activity. By using this approach, the lifetimes of hydrophobic protein interactions were extended from 2 h for the unsubstituted pyridine complex, to more than 24 h for several derivatives. Free complexes were tested for their anticancer activity against the SW480 human colon carcinoma cell line, exhibiting low cytotoxicity. Pre-treatment with hsA improved the solubility of every compound and led to some changes in activity. Particularly notable was the difference in activity between the methyl- and dibenzyl-functionalized complexes. The former shows reduced activity after incubation with hsA, indicating reduced bioavailability due to protein coordination. The latter exhibits little activity on its own but, following treatment with hsA, exhibited significant cytotoxicity, which is consistent with its ability to form non-coordinate interactions with the protein. Overall, our studies demonstrate that non-coordinate interactions with hsA are a viable target for enhancing the activity of Ru(III)-based complexes in vivo., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

As a proof-of-principle, two hetero-bimetallic PNA oligomers containing a ruthenium(II) polypyridyl and a cyclopentadienyl manganese tricarbonyl complex have been prepared by serial combination of solid-phase peptide coupling and in-solution thiol chemistry. Solid-phase N-terminus attachment of Ru(II)-polypyridyl carboxylic acid derivative, C1, onto the thiol-functionalized PNA backbone (H-a-a-g-t-c-t-g-c-linker-cys-NH 2) has been performed by standard peptide coupling method. As two parallel approaches, the strong affinity of thiols for maleimide and haloacetyl group has been exploited for subsequent post-SPPS addition of cymantrene-based organometallic cores, C2 and C3. Michael-like addition and thioether ligation of thiol functionalized PNA1 (H-gly-a-a-g-t-c-t-g-c-linker-cys-NH 2) and PNA2 (C1-a-a-g-t-c-t-g-c-linker-cys-NH 2) to cymantrene maleimide and chloroacetyl derivatives, C2 and C3, respectively, has been performed. The synthesized ruthenium(II)-cymantrenyl PNA oligomers have been characterized by mass spectrometry (ESI-MS) and IR spectroscopy. The distinct Mn-CO vibrational IR stretches, between 1,924-2,074 cm (-1) , have been used as markers to confirm the presence of cymantrenyl units in the PNA sequences and the purity of the HPLC-purified PNA thioethers assessed using LC-MS.

We report the development of an enhanced algorithm for the calculation of collision cross-sections in combination with Travelling-Wave ion mobility mass spectrometry technology and its optimisation and evaluation through the analysis of an organoruthenium anticancer complex [(eta6-biphenyl)Ru(II)(en)Cl]+. Excellent agreement was obtained between the experimentally determined and theoretically determined collision cross-sections of the complex and its major product ion formed via collision-induced dissociation. Collision cross-sections were also experimentally determined for adducts of this ruthenium complex with the single-stranded oligonucleotide hexamer d(CACGTG). Ion mobility tandem mass spectrometry measurements have allowed the binding sites for ruthenium on the oligonucleotide to be determined., (Copyright 2009 John Wiley & Sons, Ltd.)

The localization of proteoglycans in rat epiphyseal growth plate cartilage was investigated immunoelectron microscopically by the post-embedding method, using mouse monoclonal antibody (2-B-6) which specifically recognizes 4-sulphated chondroitin or dermatan sulphate after digestion of proteoglycans with chondroitinase ABC. Fixation with ruthenium hexamine trichloride (RHT) and embedding in LR White served to preserve chondrocytes in the expanded state and matrix proteoglycans were observed as a reticular network of filaments. Immunoelectron microscopy revealed gold labelling of the secondary antibodies for the demonstration of proteoglycans on these filamentous structures and in elements of the Golgi apparatus. Filaments associated with matrix vesicles were also labelled. After fixation in the presence of RHT, it was clearly demonstrated that cartilage matrix proteoglycans are retained approximately in their original spatial distribution and their antigenicity is well preserved.

We tested various cationic dyes chemically related to ruthenium hexaammine trichloride (RHT) [i.e., the RHT-cyclohexanedione complex (RHT-CC), pentaamine ruthenium N-dimethylphenylenediimine trichloride (PRT), tris-(bipyridyl)ruthenium (II) chloride (TRC), tris (bipyridyl) iron (II) chloride (TIC), and cobalt hexaammine trichloride (CHT)] for their effectiveness in precipitating cartilage matrix proteoglycans in situ. Dyes were introduced into media at the onset of processing and were present throughout both aldehyde fixation and osmium tetroxide post-fixation. Contrary to expectation, most of the dye-proteoglycan complexes generated and stable under aldehyde fixation conditions were found to be unstable during post-fixation despite the continuing presence of the dye. A similar phenomenon was also found for the cationic dyes commonly used for precipitation of proteoglycans in cartilage tissue sections (such as Acridine Orange, Alcian Blue, Azure A, Methylene Blue, and Ruthenium Red). Only two dyes, i.e., RHT and the newly tested RHT-CC, formed proteoglycan precipitates sufficiently stable to resist disruption and extraction during osmium tetroxide post-fixation. The latter may be particularly useful in semiquantitative analyses of proteoglycan content in unstained tissue sections owing to its intense brown-black color. For applications in which the osmium tetroxide post-fixation step may be omitted, TRC and PRT may also be valuable for semiquantitative histochemistry by virtue of their stable fluorescence and intense violet color signals, respectively.

Ruthenium tetroxide fixation has permitted the electron microscopic visualization of intercellular lipid lamellae in thin sections of stratum corneum. This development complements prior freeze-fracture studies of lipid lamellae and has advanced our knowledge about the ultrastructure of epidermal lipids in several ways. We have demonstrated a continuous lipid envelope that surrounds each differentiated stratum corneum cell and the presence of lipid lamellae throughout the entire stratum corneum of three mammalian species, including humans. Wherever lamellae are seen, they are present in multiples of one, two, or more pairs of bilayers, consistent with their formation from fused, flattened lipid vesicles. A unique pattern of lipid monolayers intervening between each pair of bilayers, based on sharing lipid chains between bilayers, has been proposed. In regions where there are no intercellular lamellae between corneocytes, intervening monolayers are in contact with adjacent lipid envelopes that might be involved in stratum corneum cohesion. However, limitations to the ruthenium technique must be overcome before changes in lamellar patterns can be accurately attributed to, or correlated with, changes in permeability brought about by experimental procedures or in diseased states.

The ultrastructural appearance of different types of basement membrane was studied using histochemical methods for visualizing glycosaminoglycans. Samples of rat gingiva and mouse molar germ tissue were fixed either with glutaraldehyde, glutaraldehyde-ruthenium hexammine trichloride (RHT), glutaraldehyde-Cuprolinic Blue (CB) or cetylpyridinium chloride-glutaraldehyde (CPC). Ultrathin sections were stained with uranyl acetate and lead citrate. The results showed that the conventional trilaminar structure of the basement membrane was observed after glutaraldehyde and CB fixation. In contrast, after CPC or RHT fixation, the appearance of the basement membrane was homogeneous without any evidence of a lamina lucida. Furthermore, after single fixation with CPC, the ultrastructure of different basement membranes from oral tissues showed some differences in appearance which were related to their localizations, functions, or both.

The polymorphism of a long-chain cycloparaffin (CH2)120 and chain packing in its crystals were discussed on the basis of some results obtained mainly by transmission electron microscopy. Monoclinic and orthorhombic single crystals of (CH2)120 were isothermally grown together from a dilute solution in p-xylene. Lozenge-shaped orthorhombic single crystals were more frequently observed than lath-shaped monoclinic ones. The basal surfaces of orthorhombic and monoclinic single crystal platelets were decorated with vapor-deposited polyethylene [PE]. Orthorhombic single crystals of (CH2)120 with the (110) twin boundary and monoclinic ones with the (100) twin boundary were also observed. Rod-like edge-on crystals of (CH2)120 were grown from a dilute p-xylene solution onto the (001) surface of alkali halides. The crystal system of the (CH2)120 edge-on crystals depended on the kind of substrate. The monoclinic crystal was grown on NaCl, the orthorhombic one on KBr and KCl. The monoclinic form of (CH2)120 edge-on crystal was transformed to the orthorhombic one by annealing on NaCl. In both monoclinic and orthorhombic edge-on crystals, the molecular plane determined by two zigzag stems in a molecule of (CH2)120 was parallel to the substrate surface and the molecular axis (crystallographic c-axis) was perpendicular to the longer side of the rod-like edge-on crystals. The sub-cell dimensions of the stem chains in both forms of (CH2)120 crystals were very similar to those of monoclinic and orthorhombic PE crystals, respectively.

Ruthenium hexammine trichloride (RHT) and acridine orange were used to preserve and visualize anionic groups in human plaque and dental calculus. RHT-reacting material was present on the membrane of micro-organisms and in intermicrobial spaces of the calcifying areas, and seems to correspond to, and derive from, acidic glyco- and phospholipids of the plasma membrane of the micro-organisms. However, the presence of acidic salivary peptidoglycans cannot be ruled out. Two types of calcification were found: extramicrobial and intramicrobial. The former consisted of calcified deposits irregularly scattered in the intermicrobial matrix. They were in close relationship with RHT-reacting material, or were placed inside vesicular structures delimited by a membrane. Intramicrobial calcification consisted of small aggregates of needle-shaped crystals and/or of granular deposits; in both cases, they either masked the whole cytoplasm of the micro-organisms, or were located only over the plasma membrane. These results suggest that mineral deposition occurs in connection with acidic components of intermicrobial matrix, microbial plasma membranes, and cytoplasms. The addition of RHT and acridine orange to fixing and decalcifying solutions yields satisfactory preservation of dental calculus and plaque, and apparently reduces loss of their anionic organic components and increases their electron density. However, these substances are not sufficient to preserve all ultrastructural details in decalcified areas, probably because the inorganic substance prevents reaction of acridine orange and RHT with the organic components of the calcified matrix.

Several methods of chemical fixation of avian physeal cartilage were compared. The Ruthenium hexammine trichloride method was compared to isotonic glutaraldehyde and neutral buffered formalin for light microscopy and paraffin embedment, and to two osmium-ferrocyanide methods and a combination of 1% glutaraldehyde and 4% formaldehyde for electron microscopy. Only the Ruthenium hexammine trichloride method prevented the loss of matrix proteoglycans and shrinkage of chondrocytes. In undecalcified paraffin-embedded cartilage, preservation of matrix and cellular detail was excellent, but Ruthenium hexammine trichloride interfered with Haematoxylin and Eosin staining. Glutaraldehyde gave more intense eosinophilia than neutral buffered formalin. Ultrastructurally, the Ruthenium hexammine trichloride method was the most consistent and gave the best overall fixation. Matrix elements and cellular and nuclear membranes were well preserved. It did result in vacuolation of the cytoplasm and mitochondria, and it increased granularity of the cytoplasm, chromatin, and rough endoplasmic reticulum. Other fixatives produced minimal vacuolation and finer granularity, but preservation was less consistent, cell/matrix contrast was often excessive, and they caused shrinkage of all chondrocytes. Large dilatations of the rough endoplasmic reticulum that appear to be cytoplasmic inclusions by light microscopy are described for the first time in avian cartilage.

Despite the importance of intercellular lamellar bilayers for stratum corneum (SC) barrier function, knowledge about the structure of these bilayers is limited due to their poor visualization and/or retention. Whereas substitution of ruthenium tetroxide (RuO4) for osmium tetroxide fixation provides clear images of these bilayers, the usefulness of RuO4 has been limited by its slow penetration and cytotoxicity. Utilizing a new fixation protocol for RuO4, we obtained clear images of lamellar domains at all levels of murine SC. Computer-aided image reconstructions demonstrated a lamellar spacing of 129 +/- 2 A, which agreed with x-ray diffraction data from parallel, unfixed samples (131 +/- 2 A), a spacing not affected by hydration. Furthermore, novel structures were seen in the intercellular spaces of normal SC. Finally, in murine essential fatty acid deficiency (EFAD), the overall lamellar spacing is comparable to normal [127 +/- 7 A by computer transform vs. 131.9 +/- 2 A (hydrated) and 129.6 +/- 2.2 A (dry) by x-ray diffraction]. Yet, these domains are structurally abnormal, displaying regions with either an excess or absence of lamellae. The new RuO4 protocol provides quantitative information about SC lamellar dimensions and morphologic abnormalities in bilayer distribution and substructure in EFAD stratum corneum that are not detected by either x-ray diffraction or computer-aided image reconstruction. Thus, the barrier abnormality in EFAD stratum corneum can be ascribed either to focal depletion of lamellae or abnormalities in lamellar substructure.

Bone marrow from domestic fowls was initially fixed for electron microscopy in glutaraldehyde and post-fixed with either ruthenium tetroxide or osmium tetroxide. Inclusions with distinct outlines were revealed in the secondary granules of heterophil leucocytes after ruthenium tetroxide but not with osmium tetroxide fixation. In longitudinal orientation, these inclusions were rod-shaped and composed of microfilaments measuring 3.7 nm in diameter. In transverse section, the outline of some of these inclusions was hexagonal and therefore the inclusions may be crystalline in nature.

The epidermis of avians and terrestrial mammals has evolved distinct, but related mechanisms to survive in a terrestrial environment. In both phyla, stratum corneum lipids form the basis of the cutaneous permeability barrier, but barrier function is less efficient in avians. Whereas in mammals the epidermal lamellar body (LB) secretes its contents into the intercellular spaces, in the feathered epidermis of avians, its distinctive secretory organelle, the multigranular body (MGB), does not secrete its contents into the stratum corneum intercellular spaces. Yet, neutral lipid-enriched droplets, derived from the cytosolic breakdown of MGB, ultimately are squeezed through membrane pores into the stratum corneum interstices. In this study we determined: a) using ruthenium tetroxide (RuO4) fixation, whether these droplets form membrane structures after deposition in the stratum corneum interstices; and b) the similarities and differences between avian MGB and mammalian LB, using enzyme cytochemistry as a marker for secretion, and optical diffraction computer-aided image analysis and reconstruction to compare the internal structure of MGB vs. LB. MGB were shown to possess a similar lamellar substructure to LB in RuO4-fixed specimens, exhibiting comparable dimensions on optical diffraction and computer transform analysis. Moreover, the intercellular lipids of avian stratum corneum lacked membrane-substructure, as was present in parallel samples of mammalian stratum corneum. Thus, both the absence of MGB secretion, and the failure of intercellular lipids to form membrane bilayers may explain the inherent differences in barrier function in these two taxa.

The application of ruthenium dioxide (RuO2) modified electrodes to the electrocatalytic detection of the saccharide-related antibiotics streptomycin, novobiocin and neomycin, at low fixed potentials, was investigated. The RuO2-modified graphite - epoxy composite electrodes give extremely stable and reproducible catalytic oxidation currents for these antibiotics at potentials as low as +0.2 V (versus Ag - AgCl). Rapid quantification at the micromolar level is therefore possible. Standard calibration graphs for streptomycin and neomycin yielded slopes of 4.43 and 0.08 nA microM-1 over the linear ranges of 1.5 x 10(-6) - 2.5 x 10(-4) and 1 x 10(-5) - 2 x 10(-3) M, respectively. Owing to its catalytic oxidation by the RuIII - RuIV couple, rather than the RuIV - RuVI transition (which catalyses the oxidation of streptomycin and neomycin), novobiocin could be detected at a lower (+0.2 V) potential, with a sensitivity of 1.31 nA microM-1. Detection limits of 1.5, 6.0 and 10 microM were obtained for streptomycin, novobiocin and neomycin, respectively. These catalytic surfaces can be renewed (by polishing), with a surface-to-surface reproducibility of 6.5% for the detection of 5 x 10(-5) M streptomycin. The analytical application of RuO2-modified carbon paste electrodes to the analysis of these antibiotics by flow injection was investigated, with a view to liquid chromatographic separation with electrochemical detection applications.

Ruthenium dioxide (RuO2) containing carbon paste electrodes exhibiting electrocatalytic response toward carbohydrates are described. The electrocatalytic behavior is exploited for developing a highly stable and sensitive flow detection scheme for carbohydrates at a low and fixed potential (+0.4 V vs Ag/AgCl). The effects of pH, flow rate, operating potential, surface "loading", concentration, and other variables are explored. The electrode response was stable for more than 48 h, with a signal loss of less than 10% over this period. The detection limits at the picomole level and a relative standard deviation of 1.2% (n = 72) are reported. Electrocatalytic oxidation is described also for related polyhydroxyl compounds (aldonic and aldaric acids and alditols).

The purpose of this investigation was to examine the extracellular matrix of rabbit ligament before and after digestion with glycosaminoglycan degrading enzymes. In order to preserve and enhance the visibility of negatively charged tissue components, particularly the glycosaminoglycan-containing proteoglycans, the cationic stains ruthenium red (RR) and ruthenium hexamine trichloride (RHT) were used. Cross-sections of the midsubstance of 10-month-old (mature) rabbit medial collateral ligaments fixed using conventional procedures revealed a sparse population of stellate-shaped cells that did not appear to be interconnected. Similar tissue fixed in either RR or RHT showed an extensive network of thin, electron-dense "seams" that interconnected cells and appeared to irregularly subdivide the extracellular matrix (ECM). These seams mainly consisted of a meshwork of microfilaments throughout which small granules were dispersed. Numerous 14-nm microfibrils, as well as mature elastic fibers were also present within the seams. The size and shape of the microfilaments, together with their threadlike, beaded appearance suggested that they could be Type VI collagen. The seam granules were easily removed with chondroitinase ABC, chondroitinase AC II, and mild (0.18 M) salt treatment. Only chondroitinase ABC succeeded in removing additional granules, tentatively identified as proteodermatan sulphate molecules, that were periodically located at d band sites along the Type I collagen fibrils. These results suggest that the seam granules are not dermatan sulphate containing proteoglycans, and further, that these proteoglycans may be sequestered into specific zones within the ECM through loose association with the seam microfilaments. While the functional significance of the seams remains unknown and their specific composition clearly requires further study, it is likely that they represent important functional (e.g., viscoelastic) or biological (e.g., nutritional) subdivisions of ligament substance.

Steady-state spectra of cytochrome oxidase in phospholipid vesicles were obtained by using hexaammineruthenium(II) and ascorbate as reductants. Cytochrome a was up to 80% reduced in the steady state in coupled vesicles. Upon addition of nigericin or acetate, which decrease delta pH, resulting in an increase in delta psi, cytochrome a became more oxidized in the steady state with no change in the rate of respiration. On the other hand, uncouplers or valinomycin plus nigericin, which lower both delta psi and delta pH, stimulated respiration 2-8-fold and also lowered the steady-state level of reduction of cytochrome a. These experiments indicate that electron transfer between cytochromes a and a 3 is sensitive primarily to the pH gradient. Studies with the reconstituted and the soluble enzyme at various pH values indicated that the pH on the matrix side of the membrane, rather than delta pH, controlled the steady-state level of reduced cytochrome a. Hexaammineruthenium(II) substituted for cytochrome c in measurements of proton pumping by cytochrome oxidase. Dicyclohexylcarbodiimide, which eliminated proton pumping by cytochrome oxidase, decreased the effect of ionophores on the steady-state level of reduced cytochrome a.

The epidermal permeability barrier necessary for terrestrial life resides in the intercellular spaces of the stratum corneum and is composed of lipids. Membrane coating granules (MCGs), small intracellular organelles found in the uppermost layers of the living epidermis, contain stacks of membranous disks which are extruded into the intercellular space and undergo both biochemical and physical changes to form the lipid sheets which constitute this barrier. Using ruthenium tetroxide as a secondary fixative, we are able to demonstrate stacks of lamellae filling the intercellular spaces in the uppermost layers of the stratum corneum. The structure of these lipid lamellae is consistent with the proposed derivation of MCG lipid disks and also suggests that the lipid bilayer adjacent to the corneocyte cell envelope may be assembled from lipids not derived from MCGs.

Fixation-dependent selective contrast-enhancement of the cochlear duct (or endolymphatic) glycocalyx was observed in the guinea pig inner ear after post-fixation with osmium tetroxide-potassium rutheniumcyanide. Contrast of the perilymphatic glycocalyx, however, was not significantly enhanced. The possible factors underlying this phenomenon are discussed.

A comparative study of the binding of square planar cis- and trans-[Pt(NH3)2Cl2] complexes and the octahedral [Ru(NH3)5(H2O)]3+ complex to tRNAphe from yeast was carried out by X-ray crystallography. Both of the carcinostatic compounds, cis-[Pt(NH3)2Cl2] and [Ru(NH3)5(H2O)]3+ show similarities in their mode of binding to tRNA. These complexes bind specifically to the N(7) positions of guanines G15 and G18 in the dihydrouridine loop. [Ru(NH3)5(H2O)]3+ has an additional binding site at N(7) of residue G1 after extensive soaking times (58 days). A noncovalent binding site for ruthenium is also observed in the deep groove of the acceptor stem helix with shorter (25 days) soaking time. The major binding site for the inactive trans-[Pt(NH3)Cl2] complex is at the N(1) position of residue A73, with minor trans-Pt binding sites at the N(7) positions of residues Gm34, G18 and G43. The similarities in the binding modes of cis-[Pt(NH3)2Cl2] and [Ru(NH3)5(H2O)]3+ are expected to be related to their carcinostatic properties.

Ruthenium hexammine trichloride (RHT) used as a probe to visualize anion groups in the predentine and dentine of rat incisors, showed a complex distribution pattern including: intercellular proteoglycans (Pg), detected in the predentine as granules 10-15 nm in diameter and as filaments. Non-aggregating Pg was observed in the spaces between collagen fibres as an amorphous group substance. The dentine included smaller RHT-positive granules, observed after thin-section demineralization; pericellular aggregates, 30-50 nm in diameter, which were absent at the onset of the cell coat along the plasma-membrane of the odontoblast process and of the membrane itself. All these RHT-positive components might be Pg and/or sialoglyconjugates and glyco- or phospholipids present on the plasma membrane.

Articular-epiphyseal cartilage complexes from the distal femur and humerus of five 3-month-old pigs were collected and preserved using either a conventional or a ruthenium hexammine trichloride (RHT)-supplemented fixation technique. Lesions were similar regardless of the fixation technique. Areas of necrosis were in epiphyseal but not articular cartilage from both sites of all pigs. Cartilage canals were confined to epiphyseal cartilage and contained vessels which had endothelial cells in varying stages of degeneration and necrosis. Areas of necrotic cartilage often were adjacent to or surrounded degenerate cartilage canals. Lipid emboli (up to 40.0 micron in diameter) were infrequently located in vessels within cartilage canals. Associated with the lipid emboli were leukocytes, erythrocytes, necrotic cell remnants, and flocculent material. Restriction of necrosis to epiphyseal cartilage and the association of these necrotic areas with degenerating vessels in cartilage canals strongly implicate a defect in cartilage canal blood supply in the pathogenesis of osteochondrosis. The RHT fixation technique resulted in excellent cellular detail at the light microscopic level, but ultrastructurally there was marked vacuolation of chondrocytes and matrix. The conventional technique caused shrinkage of all chondrocytes, resulting in a wide halo of pericellular matrix surrounding each cell.