The authors confirm that all data underlying the findings are fully available without restriction. All relevant data are within the paper and its Supporting Information files, except for Xray structural data deposited in the CSD. CCDC Nos. 1010556 and 1010557 contain the supplementary crystallographic data for 1, and 3, respectively. These data can be obtained free of charge via <http://www.ccdc.cam.ac.uk/conts/retrieving.html>, or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (44) 1223-336-033; or email: <deposit@ccdc.cam.ac.uk>.

Introduction {#s1}
============

Gold-based medication was used for a wide range of ailments already in the distant history of ancient China 2500 BC [@pone.0107373-BernersPrice1], [@pone.0107373-Ho1]. Thus, it is quite extraordinary that even current clinical practice still recognizes chrysotherapy (*chrysos*, gold in Greek), i.e. the treatment of diseases by the administration of gold-based therapeutic agents, as an integral part of modern medicine [@pone.0107373-ShawIII1], [@pone.0107373-BernersPrice2]. During the last eighty years of clinical use of gold-based metallotherapeutics, two major groups of complexes have been introduced. The first one is represented by the parenteral preparations, containing relatively simple gold(I)-complexes, such as sodium aurothiomalate (Myochrysin, sodium ((2-carboxy-1-carboxylatoethyl)thiolato)gold(I)) and aurothioglucose (Solganol, {(2 *S*,3*R*,4 *S*,5 *S*,6*R*)-3,4,5-trihydroxy-6-(hydroxymethyl)-oxane-2-thiolato}gold(I)) [@pone.0107373-Sigler1], and the second one comprises the orally administered drug Auranofin (Ridaura) [@pone.0107373-Williams1]--[@pone.0107373-Madeira1] (triethylphosphine-(2,3,4,6-tetra-*O*-acetyl-1-D-thiopyranosato-*S*)gold(I)) (See [Figure 1](#pone-0107373-g001){ref-type="fig"}). Nowadays, the gold-based metallotherapeutics do not represent antiarthritic agents of the first choice, since new highly potent and more specific anti-rheumatic compounds have been developed (i.e. biopharmaceuticals, novel non-steroidal anti-inflammatory drugs, NSAIDs, or corticosteroids), however, they are still indicated in specific conditions, including moderately to severely active rheumatoid arthritis [@pone.0107373-Madeira1], [@pone.0107373-Rau1]. Although the clinical application of Auranofin has declined in recent years, there are several attempts for repurposing it in other significant indications, such as cytoprotective agents and in the treatment of HIV, severe microbial infections and cancer [@pone.0107373-BernersPrice1], [@pone.0107373-Ho1], [@pone.0107373-BernersPrice2], [@pone.0107373-Madeira1], [@pone.0107373-Ott1].

![Schematic representations of gold(I) complexes used as anti-inflammatory drugs.\
**A**: Sodium aurothiomalate (Myochrysin, sodium {(2-carboxy-1-carboxylatoethyl)thiolato}gold(I)); **B**: Aurothioglucose (Solganol, {(2 *S*,3*R*,4 *S*,5 *S*,6*R*)-3,4,5-trihydroxy-6-(hydroxymethyl)-oxane-2-thiolato}gold(I)); **C**: Auranofin (Ridaura, triethylphosphine-(2,3,4,6-tetra-*O*-acetyl-1-D-thiopyranosato-*S*)gold(I)).](pone.0107373.g001){#pone-0107373-g001}

Several Auranofin inspired gold(I)-complexes have been studied for significant anti-inflammatory, and/or antitumor activities. Among linear gold(I) phosphine complexes, those involving ligands as e.g. dithiocarbamates [@pone.0107373-BernersPrice1], sulfanylpropenoates [@pone.0107373-Barreiro1], naphthalimide derivatives [@pone.0107373-Ott2], imidazole, pyrazole [@pone.0107373-Gallassi1], [@pone.0107373-Abbehausen1] as well as purine derivatives [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1] have been investigated. It has been established that the gold(I)-phosphine moiety is responsible for the actual interactions [@pone.0107373-ShawIII1], [@pone.0107373-Craig1] with the target sites of the biological molecules, i.e. mostly the selanyl-groups of enzymes [@pone.0107373-BernersPrice1], [@pone.0107373-Gandin1], while the other, weaker bonded ligand influences the kinetic profile of the compounds. This principle has been employed by our group in several recently published works, describing highly antitumor active as well as anti-inflammatory potent gold(I) complexes of the type \[Au(L)(PPh~3~)\] involving various *N*6-benzyladenine derivatives (L) [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1].

In this work, we report a series of gold(I) complexes of the general formula \[Au(L~n~)(PPh~3~)\], where HL~n~ represents variously mono- and disubstituted derivatives of hypoxanthine (See [Figure 2](#pone-0107373-g002){ref-type="fig"}). The rationale for the selection of these *N*-donor ligands is based on the fact that variously substituted derivatives of hypoxanthine have been identified as promising inhibitors of diverse essential enzymes, as cyclin-dependent kinases [@pone.0107373-Gibson1], [@pone.0107373-Arris1] and *O*6-alkylguanine-DNA-alkyltransferase [@pone.0107373-Griffin1], [@pone.0107373-Schirrmacher1], and therefore in combination with the {Au-PPh~3~} moiety could bring additional means of influencing the cellular metabolism both on the level of cell division, and modulation of cellular responses to the inflammatory stimuli.

![Schematic representation of the prepared gold(I) complexes 1--9.](pone.0107373.g002){#pone-0107373-g002}

The complexes were fully characterized, including high resolution NMR spectroscopy and single crystal X-ray analysis (for complexes **1** and **3**), and were evaluated for their cytotoxicity against a panel of nine human cancer cell lines (MCF7, HOS, THP-1, A549, G361, HeLa, A2780, A2780R (*cisplatin* resistant), and 22Rv1, and also for their *in* *vitro* anti-inflammatory effect on the model of LPS-stimulated human monocytic leukaemia (THP-1) macrophage-like cell line and the mechanisms of interaction of the complexes with sulfur-containing biomolecules (i.e. L-cysteine and reduced glutathione) were studied by means of mass spectrometry. Moreover, this work fits within the focus of our long term study dedicated to research and development of new transition metal complexes showing various types of biological effects, represented dominantly by the anticancer {Pt(II) [@pone.0107373-tarha1], Pd(II) [@pone.0107373-Vrzal1], Cu(II) [@pone.0107373-Buchtk1], [@pone.0107373-Trvnek2] or Ru(III) [@pone.0107373-Trvnek3]}, anti-inflammatory {Au(I/III)} [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1], [@pone.0107373-Kikavov1], antidiabetic and cytoprotective {Cu(II) [@pone.0107373-Vano1]} activities.

Materials and Methods {#s2}
=====================

Chemicals and Biochemicals {#s2a}
--------------------------

All the chemicals, involving H\[AuCl~4~\]·3H~2~O (Acros Organics, Pardubice, Czech Republic), triphenylphosphine (PPh~3~; Sigma-Aldrich Co., Prague, Czech Republic), NaOH (Sigma-Aldrich Co., Prague, Czech Republic) and used solvents (acetone, diethyl ether, dimethyl sulfoxide (DMSO), toluene, *n*-hexane and *N, N^'^*-dimethylformamide (DMF); Fisher-Scientific, Pardubice, Czech Republic) were obtained from the commercial sources and were used without any further purification. The *N*-donor ligands derived from hypoxanthine, HL~1--9~ (HL~1~ = 6-ethoxy-9*H*-purine; HL~2~ = 6-butyloxy-9*H*-purine; HL~3~ = 6-allyloxy-9*H*-purine; HL~4~ = 6-benzyloxy-9*H*-purine; HL~5~ = 6-phenetyloxy-9*H*-purine; HL~6~ = 2-chloro-6-ethoxy-9*H*-purine; HL~7~ = 2-chloro-6-butyloxy-9*H*-purine, HL~8~ = 2-chloro-6-allyloxy-9*H*-purine, HL~8~ = 2-chloro-6-benzyloxy-9*H*-purine) were prepared according to the previously published procedure [@pone.0107373-Gibson1] and characterized by elemental analysis, FT-IR and ^1^H and ^13^C NMR spectroscopy, which gave evidence about their composition and purity. The starting \[AuCl(PPh~3~)\] complex was synthesized as described previously [@pone.0107373-Mann1], [@pone.0107373-Bruce1].

The RPMI 1640 medium and penicillin-streptomycin mixture (Lonza, Verviers, Belgium), phosphate-buffered saline (PBS), foetal bovine serum (FBS), phorbol myristate acetate (PMA), prednisone (≥98%), Auranofin (≥98%), erythrosin B, and *Escherichia coli* 0111:B4 lipopolysaccharide (LPS) (Sigma-Aldrich, Steinheim, Germany), as well as a Cell Proliferation Reagent WST-1, and cOmplete Proteinase Inhibitor Cocktail (Roche, Mannheim, Germany) were obtained from commercial sources.

A RealTime Ready Cell Lysis Kit (Roche, Mannheim, Germany) served for isolation of RNA from cells and Transcriptor Universal cDNA Master (Roche, Mannheim, Germany) was used for reverse transcription of RNA to cDNA. Specific primers and probes (Gene Expression assays) for polymerase chain reaction (PCR) were obtained from Applied Biosystems (Foster City, CA, USA). The following assays were chosen for the quantification of gene expression: Hs00174128_m1 for tumour necrosis factor-α (TNF-α), Hs01555410_m1 for interleukin-1β (IL-1β), and 4326315E for β-actin, which served as an internal control of gene expression. Quantitative PCR (qPCR) was performed with Fast Start Universal Probe Master (Roche, Mannheim, Germany). Instant ELISA Kits (eBioscience, Vienna, Austria) were used to evaluate the production of TNF-α and IL-1β by the enzyme linked immunosorbent assay (ELISA). The Immun-Blot PVDF (polyvinylidene fluoride) membrane 0.2 µm (Bio-Rad, Hercules, CA, USA) and albumin bovine fraction V (BSA) (Serva, Heidelberg, Germany) were used for Westernblot. Murine monoclonal anti-I*κ*B-α (Cell Signaling, Danvers, MA, USA), murine monoclonal anti-β-actin (Abcam, Cambridge, UK) and goat polyclonal anti-mouse IgG (with the conjugated peroxidase) antibodies (Sigma-Aldrich, Saint Louis, MO, USA) were applied for immunodetection. The activity of the conjugated peroxidase was detected by an Opti-4CN Substrate Kit (Bio-Rad, Hercules, CA, USA).

Chemistry {#s2b}
---------

Gold(I) complexes of the general formula \[Au(L~n~)(PPh~3~)\] (**1**--**9**), where L~n~ stands for a deprotonated form of a hypoxanthine-derived compound, were prepared by a slight modification of the previously reported procedure for the synthesis of similar \[Au(L)(PPh~3~)\] complexes, in which HL stands for 6-benzylaminopurine derivatives [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1]. Concretely, the reactions of the acetone solutions of \[AuCl(PPh~3~)\] (1 mmol in 10 mL) were mixed with the corresponding hypoxanthine derivative (HL~n~; 1 mmol in 20 mL), and subsequently, 1 mL of 1 M NaOH was added into the reaction mixture. The mixture was stirred for 3 h, after which precipitated NaCl was filtered off. The colourless filtrate was evaporated to dryness and the residue was collected. Recrystallization was carried out from acetone. In the case when the residue was of gel-like consistency, 20 mL of diethyl ether was added and the suspension was sonicated to obtain a powder product. Crystals of complexes **1** and **3**, suitable for single crystal X-ray analysis, were prepared by a slow diffusion of *n*-hexane into a saturated solution of the appropriate complex in toluene. The purity and composition of the obtained gold(I) complexes **1**--**9** were established based on the methods of elemental analysis, electrospray-ionization mass spectrometry (ESI-MS), ^1^H and ^13^C NMR, and FT-IR spectroscopies, and thermal analysis (TG/DTA) (characterization data are given in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}). The molecular and crystal structures were determined by single crystal X-ray analysis for complexes **1** and **3**.

Physical Measurements {#s2c}
---------------------

Elemental analyses (C, H, N) were carried out on a Flash 2000 CHNO-S Analyser (Thermo Scientific, USA). FT-IR spectra were measured on a Nexus 670 FT-IR spectrometer (ThermoNicolet, USA) by the ATR technique in the 200--4000 cm^−1^ range. ^1^H, ^13^C and ^31^P NMR spectra were measured in DMF-*d* ~7~ on a Varian 400 MHz NMR spectrometer at 300 K using tetramethylsilane (SiMe~4~) (for ^1^H and ^13^C spectra) and 85% H~3~PO~4~ (for ^31^P) as an internal reference standard. Mass spectra of methanol solutions of complexes were recorded on an LCQ Fleet Ion-Trap mass spectrometer using the positive mode electrospray ionization (Thermo Scientific, USA). Thermogravimetric (TG) and differential thermal (DTA) analyses were performed on a thermal analyser Exstar TG/DTA 6200 (Seiko Instruments Inc., Japan) in dynamic air conditions (50 mL min^−1^) between room temperature (ca 25°C) and 950°C (gradient 2.5°C min^−1^). Single crystal X-ray analyses were performed on an Xcalibur2 diffractometer equipped with a CCD detector Sapphire2 (Oxford Diffraction Ltd., Oxford, UK) using MoKα radiation (monochromator Enhance, Oxford Diffraction Ltd.), and using the ω-scan technique at 120 K. Data collection, data reduction and cell parameter refinements were performed by the *C[rys]{.smallcaps}A[lis]{.smallcaps}* software package [@pone.0107373-Oxford1]. The molecular structures were solved by direct methods and all non-hydrogen atoms were refined anisotropically on *F^2^* using the full-matrix least-squares procedure (*SHELX-97*) [@pone.0107373-Sheldrick1]. H-atoms were found in difference maps and refined by using the riding model with C−H = 0.95, 0.98 and 0.99 Å, with *U* ~iso~(H) = 1.2*U* ~eq~(CH, CH~2~) and 1.5*U* ~eq~(CH~3~). The highest residual peaks of 2.623 e Å^--3^ and 1.792 e Å^--3^ were located 0.85 Å from Au1 (for complex **1**), and 0.88 Å from Au1 (for complex **3**), respectively. Molecular graphics were drawn and additional structural parameters were interpreted in *DIAMOND* [@pone.0107373-Brandenburg1] and *Mercury* [@pone.0107373-Macrae1].

Maintenance and Preparation of Macrophages {#s2d}
------------------------------------------

The human monocytic leukaemia cell line THP-1 (ECACC, Salisbury, UK) was used for the *in* *vitro* anti-inflammatory activity evaluation. The cells were cultivated at 37°C in the RPMI 1640 medium supplemented with 2 mM [l]{.smallcaps}-glutamine, 10% FBS, 100 U/mL of penicillin and 100 µg/mL of streptomycin in humidified atmosphere containing 5% CO~2~. Stabilized cells (3^rd^--15^th^ passage) were split into microtitration plates to get the concentration of 500 000 cells/mL and the differentiation to macrophages was induced by addition of phorbol myristate acetate (PMA) dissolved in DMSO at the final concentration of 50 ng/ml and the cells were incubated for 24 h. Unlike monocytes, differentiated macrophages tend to adhere to the bottom of the cultivation plates. For the subsequent 24 h, the cells were incubated with the fresh complete RPMI medium, i.e. containing antibiotics and FBS, without PMA. Then, the medium was aspirated, and the cells were washed with PBS and cultivated in the serum-free RPMI 1640 medium for next 24 h. These prepared macrophages were used for the detection of inflammatory response.

*In Vitro* Cytotoxicity Testing {#s2e}
-------------------------------

Cytotoxicity in THP-1 cells was determined by the WST-1 assay. The THP-1 cells (floating monocytes, 500 000 cells/mL) were incubated in 100 µL of the serum-free RPMI 1640 medium and seeded into 96-well plates in triplicate at 37°C. Measurements were taken 24 h after the treatment with the tested compounds dissolved in 0.1% DMSO in the concentration range of 0.16--10 µM. Viability was determined by the WST-1 test according to the manufacturer's manual. The amount of created formazan (correlating with the number of metabolically active cells in the culture) was calculated as a percentage of the control cells, which were treated only with 0.1% DMSO and was set-up as 100%. The cytotoxic IC~50~ values of the tested compounds were calculated from the obtained data. The WST-1 assay was performed spectrophotometrically at 440 nm (FLUOstar Omega, BMG Labtech, Germany).

*In* *vitro* cytotoxicity of the presented compounds was further determined by the MTT assay in a wide range of human cancer cell lines, i.e. human breast adenocarcinoma (MCF7; ECACC no. 86012803), human osteosarcoma (HOS; ECACC no. 87070202), lung carcinoma (A549; ECACC no. 86012804), malignant melanoma (G-361; ECACC no. 88030401), cervix epitheloid carcinoma (HeLa; ECACC no. 93021013), ovarian carcinoma (A2780; ECACC no. 93112519), *cisplatin*-resistant ovarian carcinoma (A2780R; ECACC no. 93112517) and prostate carcinoma (22Rv1; ECACC no 105092802) cancer cell lines purchased from European Collection of Cell Cultures (ECACC). The cells were cultured according to the ECACC instructions. They were maintained at 37°C and 5% CO~2~ in a humidified incubator. The cells were treated with the complexes **1**--**9**, free HL~1--9~ molecules, HAuCl~4~, AuCl and *cisplatin* (applied up to 50 µM) for 24 h, using multi-well culture plates of 96 wells. In parallel, the cells were treated with vehicle (DMF; 0.1%, v/v) and Triton X-100 (1%, v/v) to evaluate the minimal (i.e. positive control), and maximal (i.e. negative control) cell damage, respectively. The MTT assay was performed spectrophotometrically at 540 nm (TECAN, Schoeller Instruments LLC, Prague, CZ).

Drug Treatment and Induction of Inflammatory Response {#s2f}
-----------------------------------------------------

Differentiated macrophages were pretreated with 300 nM solutions of the complexes **1**--**9**, HL~1--9~, AuCl, \[AuCl(PPh~3~)\], PPh~3~ and Auranofin dissolved in DMSO (the final DMSO concentration was 0.1%) and with 0.1% DMSO solution (vehicle) for 1 h; the given concentrations of the tested compounds lack the cytotoxic effect (cell viability \>94%). The inflammatory response was triggered by addition of 1.0 µg/mL LPS dissolved in water to the pretreated macrophages, control cells were without the LPS treatment.

RNA Isolation and Gene Expression Evaluation {#s2g}
--------------------------------------------

For the evaluation of the expression of TNF-α, IL-1β, and β-actin mRNA, the total RNA was isolated directly from the THP-1 cells in cultivation plates using a RealTime Ready Cell Lysis Kit, according to the manufacturer's instructions.

The gene expression was quantified by two-step reverse-transcription quantitative (real-time) PCR (RT-qPCR). The reverse transcription step was performed by Transcriptor Universal cDNA Master using cell lysate as a template. The reaction consists of 3 steps: (1) primer annealing, 29°C for 10 min; (2) reverse transcription, 55°C for 10 min; and (3) transcriptase inactivation, 85°C for 5 min. FastStart Universal Probe Master and Gene Expression assays were used for qPCR. These assays contain specific primers and TaqMan probes that bind to an exon-exon junction to avoid DNA contamination. The parameters for the qPCR work were adjusted according to the manufacturer's recommendations as follows: 50°C for 2 min, then 95°C for 10 min, followed by 40 cycles at 95°C for 15 s and 60°C for 1 min. The results were normalized to the amount of ROX reference dye, and the change in gene expression was determined by the ΔΔC~T~ method. Transcription of the control cells was set as 1 and other experimental groups were multiples of this value.

Evaluation of Cytokine Secretion by ELISA {#s2h}
-----------------------------------------

Macrophages, which were pretreated with the tested compounds for 1 h, were incubated with LPS for next 24 h. After this period, the medium was collected and the concentration of TNF-α and IL-1β was measured by an Instant ELISA kit according to the manufactures' manual.

Determination of IκB Degradation by Western Blot {#s2i}
------------------------------------------------

Macrophage-like THP-1 cells were pretreated with the tested compounds and stimulated by LPS as described above. Thirty minutes after the addition of LPS, the medium was aspirated and cells were washed by cold PBS. Subsequently, the cells were collected using the lysis buffer \[50 mM Tris-HCl pH 7.5, 1 mM EGTA, 1 mM EDTA, 1 mM sodium orthovanadate, 50 mM sodium fluoride, 5 mM sodium pyrophosphate, 270 mM sucrose, 0.1% (v/v) Triton X-100, and cOmplete Protease Inhibitor Cocktail (Roche, Germany)\] and scraper. The lysis of cells was facilitated by short (ca. 30 s) incubation in the ultrasonic water bath. The protein concentration was determined according to Bradford's method. For protein separation, 30 µg of protein was loaded onto 12% polyacrylamide gel. Then, they were electrophoretically transferred on the PVDF membranes, which were subsequently blocked by 5% BSA dissolved in TBST buffer \[150 mM NaCl, 10 mM Tris base pH 7.5, 0.1% (v/v) Tween-20\]. The membranes were incubated with the primary anti-IκB-α antibody at the concentration of 1∶500, or with the primary anti-β-actin at the concentration of 1∶5000 at 4°C for 16 h. After washing, the secondary anti-mouse IgG antibody diluted 1∶2000 was applied on the membranes and incubated for 1 h at laboratory temperature (∼22°C). The amount of the bound secondary antibody was detected colorimetrically by an Opti-4CN kit according to the manufacturer's manual.

Statistical Evaluation {#s2j}
----------------------

The cytotoxicity data were expressed as the percentage of viability, when 100% represent the treatment with vehicle (DMF or DMSO). The experiments were conducted in triplicate using cells from different passages. The IC~50~ values were calculated from viability curves. The results are presented as arithmetic mean ± standard error of the mean (SE). The significance of the differences between the results was evaluated by the ANOVA analysis with p\<0.05 considered to be significant (QC Expert 3.2, Statistical software, TriloByte Ltd., Pardubice, CZ).

The statistically significant differences between individual groups during anti-inflammatory activity testing were assessed by the one-way ANOVA test, followed by Tukey's *post-hoc* test for multiple comparisons. GraphPad Prism 5.02 (GraphPad Software Inc., San Diego, CA, USA) was used for the analysis.

Interactions with L-Cysteine and Reduced Glutathione Analyzed by Mass Spectrometry {#s2k}
----------------------------------------------------------------------------------

The interaction experiments between the selected representative complexes **1** and **6**, bearing the same ethoxy-substitution on the C6 atom, but differing in the substituents in the C2 position, and the mixture of physiological levels of cysteine and glutathione were performed on a Thermo Scientific LTQ Fleet Ion-Trap mass spectrometer, using the positive ionization mode. The reaction system contained the physiological concentrations of L-cysteine (290 µM) and reduced glutathione (6 µM) [@pone.0107373-Salemi1] and the tested complex (20 µM) in the methanol:water (1∶1, v/v) mixture. The reference system was comprised of the solution of complex (20 µM) in the methanol:water (1∶1, v/v) mixture. The flow injection analysis (FIA) method was used to introduce the reaction system (5 µL spikes) into the mass spectrometer, while pure acetonitrile was used as a mobile phase. The ESI-source was set up as follows: source voltage was 4.5 kV, the vaporizer temperature was 160°C, the capillary temperature was 275°C, the sheath gas flow rate was 30 L/min, and auxiliary gas flow rate was 10 L/min. The system was calibrated according to the manufacturer specifications and no further tuning was needed.

Results and Discussion {#s3}
======================

General Properties of the Au(I) complexes {#s3a}
-----------------------------------------

This work reports on the preparation, thorough characterization, and *in* *vitro* cytotoxic and anti-inflammatory activities of a series of gold(I)-triphenylphosphine complexes \[Au(L~1--9~)(PPh~3~)\] involving mono- or disubstituted derivatives of hypoxanthine (HL~1--9~). The complexes were synthesized by a modification of a previously published method [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1], i.e. by a reaction of the precursor complex \[AuCl(PPh~3~)\] with an equimolar amount of the corresponding organic molecule HL~1--9~ in acetone with the addition of 1 M NaOH. The prepared complexes **1**--**9** are very well soluble in acetone, alcoholic solvents, toluene, DMSO and DMF, and partially soluble in water at laboratory temperature. The complexes were prepared non-solvated, as evidenced by the results of simultaneous TG/DTA analysis (see Figure S3 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}). The complexes were further characterized as chemical individuals by elemental analysis, ^1^H, ^13^C and ^31^P NMR spectroscopy, FT-IR spectroscopy and ESI--MS. Single crystals suitable for X-ray analysis were prepared in the cases of **1** and **3**, thus providing information about the molecular and crystal structures of the complexes (discussed in detail below).

^1^H and ^13^C NMR spectra were measured for all the presented complexes. The obtained results were very beneficial for the confirmation of the purity and composition of **1**--**9**. The observed signals unambiguously proved the presence of both hypoxanthine and phosphine derivatives in the structures of the Au(I) complexes as well as gave information about the coordination mode of these ligands. The most conclusive evidence concerning the identification of the donor atom of HL~1--9~ was obtained from the ^13^C NMR spectra by comparing the chemical shifts in the uncoordinated and coordinated hypoxanthine derivative. The resulting differences, calculated as coordination shifts; *Δδ* = *δ~complex~*--*δ~ligand~*, ppm; [Table 1](#pone-0107373-t001){ref-type="table"}), showed that the greatest changes occurred for the signals corresponding to the carbons C4 and C8, shifted by 2.02--4.82 ppm, and 3.57--6.89 ppm, respectively. This observation indirectly pointed to N9 as the coordination site, as the two mentioned carbons lie in the direct vicinity to this nitrogen. The set of the most intensive signals at around 130 ppm, characteristic of all the spectra of **1**--**9**, was assigned to the carbon atoms of triphenylphosphine. Accordingly, the ^1^H NMR spectra of all the nine Au(I) complexes showed very strong multiplet signals at around 7.70 ppm with the relative integral intensity well corresponding to the calculated value of 15 hydrogens of PPh~3~. Additionally, the C8 H signal of the corresponding HL~1--9~ ligand was shifted by 0.08--0.27 ppm upfield in the spectra of the complexes with respect to the spectra of the uncoordinated HL~1--9~, corresponding with the results following from ^13^C NMR spectroscopy about N9 being the site of coordination to gold. What should be also pointed out is the absence of the signal assignable to the proton N9 H in the spectra of **1**--**9**, which well agrees with the presence of deprotonated hypoxanthine-based ligands in the studied complexes symbolized as L~1--9~. All the ^31^P NMR spectra of the complexes were characteristic of the presence of one singlet at 31.67--33.13 ppm (see Figure S5 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}), which is significantly shifted as compared to the signal of free PPh~3~ (--5.96 ppm), which confirms the coordination of PPh~3~ to gold(I) atom through phosphorus, thus forming the Au--P bond. In summary, the presence and chemical shift of all the detected signals in ^1^H, ^13^C and ^31^P NMR spectra, as well as the relative intensity of the peaks in the proton spectra, indicate that the gold(I) atom in **1**--**9** is coordinated by one deprotonated hypoxanthine-derived compound (L~1--9~) binding via N9 and one PPh~3~ molecule. These conclusions are in good agreement with the results following from single crystal X-ray analysis of complexes **1** and **3** discussed below.

10.1371/journal.pone.0107373.t001

###### ^1^H and ^13^C NMR coordination shifts (Δδ = δ~complex~--δ~ligand~; ppm) of calculated for **1**--**9**.

![](pone.0107373.t001){#pone-0107373-t001-1}

           ^1^H NMR   ^13^C NMR
  ------- ---------- ----------- -------- ------ -------- -------- ------
  **1**     --0.05     --0.08     --0.90   4.80   --1.14   --1.04   4.40
  **2**     --0.05     --0.10     --0.60   4.82   --0.59   --0.21   3.57
  **3**     --0.11     --0.23      0.98    2.20   --0.56   --0.24   5.11
  **4**     --0.13     --0.23      0.86    3.00   --0.81    0.50    5.03
  **5**     --0.12     --0.25     --0.01   2.16   --1.29   --0.32   5.85
  **6**      n.a.      --0.27     --0.16   2.02   --1.17    0.05    6.02
  **7**      n.a.      --0.10     --0.04   4.01   --0.54   --0.73   6.89
  **8**      n.a.      --0.12     --0.49   2.38   --0.74   --0.35   4.89
  **9**      n.a.      --0.24     --0.53   3.22   --1.01   --0.18   5.70

n.a. -- not available.

Mass spectra measured in the positive ionization mode also indirectly confirmed the composition of the presented complexes, as all the spectra of **1**--**9** contained the \[M+H\]^+^ molecular peaks. The presence of the hypoxanthine-based derivatives was demonstrated by the observed peaks corresponding to the adducts of \[HL~n~+Na/K\]^+^ (for details of ESI+MS characterization, see [File S1](#pone.0107373.s001){ref-type="supplementary-material"}).

Further properties of the presented Au(I) complexes were studied by FT-IR spectroscopy. The courses of the spectra of **1**--**9** in the mid-IR region were qualitatively very similar to those of the uncoordinated HL~1--9~. The very intensive peaks at 1600--1578 cm^−1^ could be assigned to the ν(C--^...^N)~ring~ stretching vibrations characteristic of heterocyclic molecules with nitrogen ring atoms. Further assigned characteristic vibrations could be found at 3057--3048 cm^−1^ for v(C-H)~ar~, 2986--2889 cm^−1^ for v(C-H)~aliph~, and also 1480--1436 cm^−1^ for ν(C--^...^C)~ring~. The typical vibrations of the ether functional group found in the *O*6-substituted hypoxanthine derivatives were detected at 1339--1272 cm^−1^ for the aromatic C6--O stretch and at 1118--1087 cm^−1^ for the O--C10 stretch. The medium to strong intensity bands found between 800--700 cm^−1^, and at ca. 690 cm^−1^ can be attributed to the out-of-plane C--H bending vibrations, and ring bending, respectively, determining the presence of an aromatic ring in all the complexes **1**--**9** [@pone.0107373-Nakamoto1], [@pone.0107373-Smith1]. In the far spectra of the Au(I) complexes, new bands were identified as compared with the spectra of free HL~1--9~. These maxima at 506--492 cm^−1^, and 329--321 cm^−1^ could be assigned to v(Au--N), and v(Au--P) stretching vibrations, respectively, although it should be noted that triphenylphosphine alone has intensive vibrations in this region of IR spectra, so these bands could be overlapped [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1], [@pone.0107373-Nakamoto1], [@pone.0107373-Faggianhi1].

Molecular and Crystal Structures of \[Au(L~1~)(PPh~3~)\](1) and \[Au(L~3~)(PPh~3~)\] (3) {#s3b}
----------------------------------------------------------------------------------------

The crystallization method of slow *n*-hexane diffusion into the saturated toluene solution of the complexes allowed the preparation of single crystals suitable for X-ray analysis of two complexes, \[Au(L~1~)(PPh~3~)\](**1**) and \[Au(L~3~)(PPh~3~)\] (**3**), where HL~1~ = 6-ethoxy-9*H*-purine and HL~3~ = 6-allyloxy-9*H*-purine. The molecular structures are depicted in [Figure 3](#pone-0107373-g003){ref-type="fig"}, and [4](#pone-0107373-g004){ref-type="fig"}, respectively. Crystal data and structure refinement parameters (Table S1 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}), selected bond lengths and angles (Table S2 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}), as well as non-covalent interaction parameters (Tables S3 and S4 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}) are given in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}.

![Molecular structure of \[Au(L~1~)(PPh~3~)\] (1).\
The molecular structure of **1** showing the atom numbering scheme. Non-hydrogen atoms are displayed as ellipsoids at the 50% probability level.](pone.0107373.g003){#pone-0107373-g003}

![Molecular structure of \[Au(L~3~)(PPh~3~)\] (3).\
The molecular structure of **3** showing the atom numbering scheme. Non-hydrogen atoms are displayed as ellipsoids at the 50% probability level.](pone.0107373.g004){#pone-0107373-g004}

In both complexes **1** and **3**, the central gold(I) atom is two-coordinated in the distorted linear arrangement of the {NP} donor set formed by one *N*-donor deprotonated hypoxanthine derivative and the electroneutral molecule of triphenylphosphine bonded via phosphorus. Comparing the coordination bonds in **1** and **3**, both bond lengths are mutually comparable between the complexes, as they were found to be 2.046(3), and 2.042(3) Å for Au--N; and 2.2344(9), and 2.2356(8) Å for Au--P, respectively. On the other hand, it is evident that the Au--P bonds are significantly longer than the Au--N bonds, which is most likely connected with the bulky phenyl moieties surrounding the phosphorus donor atom in PPh~3~. This bond distance difference is not uncommon in similar complexes. The search in the Cambridge Structural Database (CSD ver. 5.35, February 2014 update) [@pone.0107373-Allen1] within the deposited 113 mononuclear compounds involving the N-Au-PPh~3~ motif with the central gold atom in the linear geometry proved that the Au--N and Au--P bond lengths in **1** and **3** fall within the interval of such bond lengths found in the above specified set of compounds, i.e. 1.971--2.134 Å, and 2.22--2.278 Å, respectively. The distortion of the linear geometry is well demonstrated on the dimensions of the N9--Au1--P1 angles, which are equal to 172.73(10)° in **1** and 175.65(8)° in **3**. These values also clearly reveal that the distortion from the ideal angle of 180° is significantly higher in complex **1** than in **3**. This difference could most likely be connected with a somewhat different array of non-covalent contacts found in the complexes. In general, the crystal structures of both complexes **1** and **3** are characterised by absence of strong non-covalent contacts, such as typical hydrogen bonds. The crystal structures of both the complexes are stabilized by C--H···C, C--H···N and C--H···O non-covalent contacts. For more details, see Supplementary Information, Tables S3 and S4 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}, Figures S1 and S2 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}.

*In Vitro* Cytotoxicity {#s3c}
-----------------------

For the evaluation of *in* *vitro* cytotoxicity, the presented complexes were tested by the MTT assay and the results were compared with the clinically applied metallodrug *cisplatin*, used as the reference standard, as well as with the starting compounds, i.e. hypoxanthine derivatives HL~1--9~, and gold-containing inorganic compounds AuCl or HAuCl~4~ (applied in the concentration range of 0.01--50.0 µM, unless their solubility was found to be lower). The first phase of testing involved screening of all the prepared complexes **1**--**9** against two human cancer cell lines, breast adenocarcinoma (MCF7) and osteosarcoma (HOS). The starting compounds were found inactive up to 50 µM or to the limiting concentration of their solubility against these cells. Then, this testing showed a varied effect of the tested complexes on the viability of the selected cancer cell lines. Complex **7** showed to be inactive up to the tested concentration range on both cell lines (IC~50~\>50 µM). Complexes **8** and **9** were found to be cytotoxic inactive against the MCF7 cells in the used concentration ranges given by their solubility, therefore their cytotoxicity can be evaluated as \>25 µM. Complex **2** was moderately active on both cell lines, however, the IC~50~ values, i.e. 27.1±0.2 µM (MCF7) and 30.0±2.1 µM (HOS), were higher than those for *cisplatin* (in the case of MCF7, significantly higher, ANOVA, p\<0.05). This pointed to the fact that the *n*-butyl group as the R1 substituent showed to be the least beneficial for the resulting cytotoxic activity against these cell lines within the tested series of compounds (*n*-Bu present in **2** and **7**). The best results were observed for the complexes **4**--**6**, all of which possessed significantly better cytotoxicity than *cisplatin* (ANOVA, *p*\<0.05) against both cell lines, concretely the IC~50~ values were found to be 2--5-times lower. The most promising data were found for complex **4** against the MCF7 cell line, i.e. IC~50~ = 3.7±0.5 µM (17.9±1.2 µM for *cisplatin*), and for complex **6** against HOS cells, i.e. 4.0±0.3 µM as compared with 20.5±0.1 µM for *cisplatin* ([Table 2](#pone-0107373-t002){ref-type="table"}).

10.1371/journal.pone.0107373.t002

###### *In vitro* cytotoxicity of complexes 1--9 and *cisplatin* against MCF7 and HOS cancer cell lines.

![](pone.0107373.t002){#pone-0107373-t002-2}

                                 Cell Line
  ------------- ------------------------------------------- ------------ -----------
  **1**                         15.22±0.78                   19.20±0.71   1.17±0.04
  **2**                         27.10±0.21                   30.02±2.13   1.03±0.04
  **3**                         12.40±1.07                   15.60±1.10   1.87±0.09
  **4**                          3.66±0.48                   11.30±0.67   2.15±0.19
  **5**                          6.30±0.80                   6.56±0.97    1.89±0.11
  **6**                          5.23±0.68                   3.96±0.29    1.94±0.12
  **7**                            \>50                         \>50      1.97±0.14
  **8**                            \>25                      18.80±0.70   5.28±0.21
  **9**                            \>25                      16.80±0.27     \>10
  *cisplatin*                   17.90±1.17                   20.50±0.10      \-
  *Auranofin*    1.10±0.30[a](#nt103){ref-type="table-fn"}      n.d.      0.88±0.04

The results of the *in* *vitro* cytotoxic activity testing of **1**--**9** and *cisplatin* against human breast adenocarcinoma (MCF7) and osteosarcoma (HOS): cells were treated with the tested compounds for 24 h; measurements were performed in triplicate, and cytotoxicity experiment was repeated in three different cell passages; data are expressed as IC~50~±SE (µM).

adopted from Ref. [@pone.0107373-Ott3]; n.d. -- not determined.

Before using the THP-1 cell line for testing of the anti-phlogistic effect, cytotoxicity of complexes **1**--**9** and uncoordinated hypoxanthine derivatives HL~1--9~ was also tested on this cell line ([Table 2](#pone-0107373-t002){ref-type="table"}). The IC~50~ values for **1**--**7** were between 1.03--2.15 µM, which are significantly lower than for the MCF7 and HOS cell lines. Interestingly, **8** has the IC~50~ only 5.3±0.2 µM and **9** does not even have any cytotoxic effect up to the concentration of 10 µM, whereas their structural analogues without chlorine at C2 (**3** and **4**) have the IC~50~ values equal to 1.87±0.09 µM, and 2.15±0.19 µM, respectively. In all the cases, cytotoxicity was lower (i.e. the IC~50~ values were higher; 1.03--5.28 µM) than for commercially used drug Auranofin (IC~50~ = 0.88±0.04 µM). Recently, the gold(I) complexes with the general composition \[Au(PPh~3~)(Y)\], where Y represents an *S*-coordinated thioamide ligand, showing remarkable cytotoxicity towards leiomyosarcoma cells with the IC~50~ values in the comparable range as **1**--**7** (0.7--2.1 µM) have been reported [@pone.0107373-Kouroulis1]. Complexes **1**--**8** additionally demonstrate the hormesis effect (higher viability; in this case up to 168%) around the concentration of 0.3 µM.

Based on the promising results for complexes **4**--**6** against all the three above mentioned cell lines, further testing was performed for these representatives on a panel of human cancer cell lines involving lung carcinoma (A549), malignant melanoma (G361), cervix epitheloid carcinoma (HeLa), ovarian carcinoma (A2780), ovarian carcinoma resistant to *cisplatin* (A2780R) and prostate carcinoma (22Rv1). The testing again revealed that the starting compounds (HL~n~ and AuCl, HAuCl~4~) were not toxic in the tested concentration range, only a weak antiproliferative effect was observed for HAuCl~4~ on the G361 cell line (38.1±0.8 µM). The IC~50~ values determined for **4**--**6** showed that viability of all the employed cell lines was reduced by the tested complexes comparably or better than by the metallodrug *cisplatin* ([Table 3](#pone-0107373-t003){ref-type="table"}, [Figure 5](#pone-0107373-g005){ref-type="fig"}). Complexes **5** and **6** were significantly more *in* *vitro* antitumour active (ANOVA, *p*\<0.05) against all the cell lines as compared with *cisplatin*, which is valid also for complex **4** with the exception of the G361 cells, against which its activity was found comparable with *cisplatin* (3.9±0.5 µM, and 5.3±0.2 µM, respectively). Focusing on individual cell lines, the lowest antiproliferative effect of **4**--**6** was observed for A549 and HeLa, which in the testing did not respond to the treatment by *cisplatin* up to 50 µM. The IC~50~ values for the tested Au(I) complexes showed moderate activity in the micromolar range (≈15--20 µM). On the other hand, low micromolar IC~50~ values (≈3--5 µM) resulted from the cytotoxicity testing against all the other cancer cell lines, *i.e.* G361, 22Rv1, A2780 as well as A2780R. It should be also pointed out that complexes **4**--**6** showed to be ca. 3-times (A2780), 5-times (A2780R) and 7-times (22Rv1) more effective than *cisplatin*. Moreover, the evaluation of cytotoxicity of **4**--**6** on both A2780 cell lines sensitive and resistant to *cisplatin* allowed the calculation of the resistance factors (RF), *i.e.* the ratio of IC~50~(A2780R)/IC~50~(A2780). The RF values are equal to 1.19 (**4**), 1.14 (**5**), 1.21 (**6**) and 2.25 (*cisplatin*), which unambiguously shows that **4**--**6** are able to circumvent the acquired resistance of cancer cells to *cisplatin*. It can be additionally pointed out that the tested complexes are comparably active on both A2780 and A2780R cell lines, as there is no statistically significant difference in their cytotoxic activity against these cancer cells (ANOVA, *p*\<0.001).

![The results of *in*\
***vitro*** **cytotoxicity against selected cancer cell lines for complexes 4--6 and** ***cisplatin*** **.** The cells were exposed to the employed complexes for 24 h. Measurements were performed in triplicate and each cytotoxicity experiment was repeated three times. The given IC~50~±SE (µM) values represent an arithmetic mean. The asterisk (\*) denotes significant difference (ANOVA, p\<0.05) between **4**--**6** and *cisplatin*.](pone.0107373.g005){#pone-0107373-g005}

10.1371/journal.pone.0107373.t003

###### *In* *vitro* cytotoxicity of complexes 4--6 and *cisplatin* against a panel of cancer cell lines.

![](pone.0107373.t003){#pone-0107373-t003-3}

                 Cell Line
  ------------- ----------- --------- ---------- ---------- ---------- ----------
  **4**          16.8±0.7    3.9±0.5   14.2±0.5   3.6±0.1    4.3±0.2    3.8±0.1
  **5**          19.3±0.8    3.4±0.1   19.8±0.9   4.2±0.2    4.8±0.6    4.4±0.3
  **6**          21.7±0.4    3.0±0.1   21.6±0.3   4.2±0.3    5.1±0.4    3.6±0.1
  *cisplatin*      \>50      5.3±0.2     \>50     12.0±0.3   27.0±1.5   26.9±1.2

The results of the *in* *vitro* cytotoxic activity testing of **4**--**6** and *cisplatin* against the human cancer cell lines. Cells were treated with the tested compounds for 24 h; measurements were performed in triplicate, and cytotoxicity experiments were repeated in three different cell passages; data are expressed as IC~50~±SE (µM).

*In Vitro* Anti-Inflammatory Activity {#s3d}
-------------------------------------

To evaluate the anti-inflammatory effect of the presented complexes *in* *vitro*, their ability to diminish the production of pro-inflammatory cytokines TNF-α and IL-1β in LPS-stimulated macrophages derived from the THP-1 cell line was determined. The results of this study showed that complexes **1**--**7** significantly decreased secretion of TNF-α in the LSP-stimulated cells, while complexes **8** and **9** exhibited no effect, as the extent of their influence on TNF-α secretion was comparable to the vehicle alone (0.1% DMSO) ([Figure 6A](#pone-0107373-g006){ref-type="fig"}). It is interesting to note, that the comparison of complexes **1**--**4** and their analogues **6**--**9** differing only in the substitution at C2 (hydrogen, and chlorine, respectively) showed that in all the cases, the complexes **1**--**4** were more effective in the TNF-α level attenuation. Moreover, all the active complexes (**1**--**7**) had comparable activity as the commercial reference drug Auranofin, but importantly they possessed a lower cytotoxic effect in the employed THP-1 cell line. To elucidate the role of each constituent part of the complexes (*N*-donor ligand, PPh~3~ or gold(I) species) in diminishing TNF-α secretion, free hypoxanthine derivatives (HL~1--9~), \[AuCl(PPh~3~)\], PPh~3~, and AuCl were also tested. It was found that none of these starting materials showed the anticipated effect. Moreover, the compound HL~7~ significantly increased the production of TNF-α. These results indicated that in order to show the anti-inflammatory effect, the corresponding ligand has to be bonded into the Au(I)-complex. This observation is in agreement with the previous studies, where no anti-phlogistic effect was observed for free and uncoordinated purine-derived compounds applied as ligands in Au(III)-complexes [@pone.0107373-Kikavov1].

![Effects of the Au(I) complexes 1--9, reference drug Auranofin, \[AuCl(PPh~3~)\], AuCl, PPh~3~ and HL~1--9~ on LPS-induced TNF-α (A) and IL-1β (B) secretion.\
The cells were pretreated with the tested compounds (300 nM) or the vehicle (0.1% DMSO) only. After 1 h of the incubation, the inflammatory response was induced by LPS \[except for the control cells (CTRL)\]. The secretion was determined 24 h after the LPS addition. The results are expressed as means±SE of three independent experiments. Significant difference in comparison to: \*vehicle-treated cells (p\<0.05), \*\*vehicle-treated cells (p\<0.01), \*\*\*vehicle-treated cells (p\<0.001).](pone.0107373.g006){#pone-0107373-g006}

The extent of the influence of complexes **1**--**9** on IL-1β secretion was very similar as for TNF-α ([Figure 6B](#pone-0107373-g006){ref-type="fig"}), but it should be noted that IL-1β production was affected slightly more than TNF-α. The same observation was also made in previous studies [@pone.0107373-Trvnek1], [@pone.0107373-Seitz1], which showed that the tested gold(I) complexes preferentially inhibit IL-1β production as compared to TNF-α. The results proved that similarly to TNF-*α*, compounds **1**--**7** significantly decreased the level of IL-1β, whereas complex **9** showed again no effect. The only difference as compared to the above-described TNF-α production influence study was found for complex **8**, which was inactive in the case of TNF-α secretion, but it significantly reduced the production of IL-1β. However, its inhibitory activity was still lower as compared with the activity of **1**--**7**. Testing of the constituent parts of the complexes as well as of starting materials revealed the element necessary for diminishing the production of IL-1β. The free HL~n~ molecules were again found to be inactive, with the exception of HL~6~ which reduced the level of the cytokine significantly as compared with the vehicle-treated cells (*p*\<0.05). On the other hand, unlike in the case of TNF-α, AuCl significantly decreased the level of this pro-inflammatory cytokine (*p*\<0.01) as well as the \[AuCl(PPh~3~)\] complex, which even curtailed its level as effectively as complexes **1**--**9**. These results indicate that the presence of the {Au-PPh~3~} moiety in the complexes is crucial for the reduction of IL-1β secretion, but the coordinated *N*-donor organic ligand can dramatically change the activity of such a complex as it is visible on the example of the complexes **8** and **9** with low or no anti-inflammatory activity.

To determine whether the production of the pro-inflammatory cytokines TNF-α and IL-1β is regulated by a post-translation or post-transcription mechanism in the presence of the Au(I) complexes, the level of the corresponding mRNA was determined. The obtained results showed that selected representative complexes **2** and **7** were able to decrease the transcription of these cytokines after the LPS stimulation ([Figure 7](#pone-0107373-g007){ref-type="fig"}). This finding indicates that the target site for the Au(I) complexes should be up-stream the transcription, which is in accordance with previous studies [@pone.0107373-Trvnek1], [@pone.0107373-Han1].

![Effects of the Au(I) complexes 2 and 7, and reference drug Auranofin on gene expression of TNF-α (A) and IL-1β (B).\
THP-1 macrophages were pretreated with complexes **2**, **7** and Auranofin at the concentration of 300 nM or the vehicle (0.1% DMSO) only. After 1 h of the incubation, the inflammatory response was induced by LPS \[except for the control cells (CTRL)\]. After 2 h, the level of TNF-α and IL-1β mRNA was evaluated by RT-qPCR. The amount of cytokine mRNA was normalised to β-actin mRNA. The results are expressed as means±SE of three independent experiments. A.U.  = arbitrary unit. Significant difference in comparison to: \*vehicle-treated cells (p\<0.05), \*\*vehicle-treated cells (p\<0.01).](pone.0107373.g007){#pone-0107373-g007}

The transcription of the cytokines TNF-α and IL-1β is controlled by transcription factor NF-κB. This protein is kept in cytoplasm by its inhibitor IκB. After the activation of the signalling pathway, this inhibitor is degraded [@pone.0107373-Hayden1]. Gold-containing compounds are known for their ability to inhibit this signalling pathway [@pone.0107373-Jeon1]. The results from the herein presented study are in agreement with the suggested mechanism. Complexes **2** and **7**, as well as Auranofin, were able to attenuate the degradation of IκB nonsignificantly (Figure S4 in [File S1](#pone.0107373.s001){ref-type="supplementary-material"}). This blocking of the NF-κB signalling pathway leads to lower transcription of TNF-α and IL-1β genes and thus, decreases their secretion.

Interactions with a Mixture of Cysteine and Glutathione Analysed by ESI-MS {#s3e}
--------------------------------------------------------------------------

As soft Lewis acids, Au(I) species prefer the formation of strong coordination bonds with soft Lewis base ligands, i.e. thiolate or selenolate ions, or phosphine derivatives, while the latter ones form the most stable bonds. It is a known fact, that Au(I) complexes behave likewise, as they bind to sulfanyl-containing substances, such as amino acid cysteine (Cys) or small proteins, such as glutathione (GSH), and with high molecular weight proteins (*e.g.* albumin or globulins [@pone.0107373-Shaw1]), in the biologically relevant environments (*i.e.* blood or serum) by the ligand exchange mechanism. The complexes formed this way can be considered as transport intermediates. The exchange of *N*-ligands for *S*-ligands occurs relatively quickly (within 20 minutes when interacting with albumin and globulins in the blood [@pone.0107373-Iqbal1]), while the *P*-ligand exchange proceeds much more slowly. It seems that in this mechanism the cooperative effects of adjacent thiolato or selenolato ligands in the neighbourhood of the interaction site play an important role. In connection with the above mentioned, the ligand exchange is interpreted as one of the molecular mechanisms of incorporation of gold into the active site of selenium-containing flavoreductases, such as thioredoxin reductase [@pone.0107373-Saccoccia1]. In the scope of our work, we strived to uncover the molecular behaviour of selected complexes **1** and **6** (applied in the concentration of 20 µM, which approximately corresponds to the highest therapeutic blood levels of gold during chrysotherapy [@pone.0107373-Lewis1]) in biologically relevant conditions using a mixture of cysteine (at the 290 µM concentration) and reduced glutathione (at the 6 µM concentration) [@pone.0107373-Salemi1].

Based on the results of the ESI-MS experiments, we confirmed that both complexes **1** and **6** react with the used sulfhydryl-containing substances in time-independent manner by the ligand-exchange mechanism based on the substitution of the *N*-ligand (L~n~) by the cysteine or glutathione molecule. This mechanism was confirmed by the emergence of ions at 1224.24 *m/z*, corresponding to the \[(Au-PPh~3~)~2~+GSH\]^+^ intermediate, and an ion at 1355.25 *m/z*, corresponding to the ionic species \[Cys+(Au-PPh~3~)~2~+PPh~3~+CH~3~OH+Na\]^+^ (see [Figure 8](#pone-0107373-g008){ref-type="fig"}). Unlike our previous reports on the biological activities of Au(I) complexes [@pone.0107373-Trvnek1], [@pone.0107373-Hoek1], in this interaction experiments we observed both the intermediates involving cysteine and glutathione molecules. This might indicate that the herein reported compounds are much more susceptible to the sulfur-containing molecules with several donor atoms, like glutathione.

![The results of the ESI-MS study of complex 1 (A) and 6 (B) solutions and interacting systems involving the mixture of physiological levels of cysteine and reduced glutathione and complex 1 (C) or complex 6 (D).](pone.0107373.g008){#pone-0107373-g008}

In concordance with the above mentioned suggestion as well as in accordance with the previously reported behaviour of some Au(I) complexes in water-containing solutions [@pone.0107373-Hoek1], the mass spectra of the reacting systems involving sulfur-containing molecules and also the reference solutions of complexes revealed a considerable instability of the complexes demonstrated by the appearance of the intensive ion at 721.41 *m/z*, corresponding to the \[Au(PPh~3~)~2~\]^+^ intermediate, and other ionic species involving the residue Au-PPh~3~ (i.e. \[M+(Au-PPh~3~)\]^+^; 1081.33 *m/z* for complex **1**, and 1115.51 *m/z* for complex **6**), the free HL~n~ molecules (\[HL~1~+H\]^+^ at 165.10 *m/z* and (\[HL~6~+H\]^+^ at 199.12 *m/z*), or the free triphenylphosphine residue (i.e. \[PPh~3~+H\]^+^ at 263.24 *m/z* and \[Au+(PPh~3~)~3~+CH~3~OH+Na\]^+^ at 1038.30 *m/z*).

Conclusions {#s4}
===========

A series of gold(I) complexes of the general composition \[Au(L~n~)(PPh~3~)\] (**1**--**9**), involving a combination of *N*-donor (HL~n~) and *P*-donor triphenylphosphine ligands, was prepared and thoroughly characterized. The *in* *vitro* cytotoxicity results against a panel of nine human cancer cell lines (MCF7, HOS, A549, HeLa, A2780, A2780R, 22Rv1, G-361 and THP-1) revealed the complexes as more anticancer active than *cisplatin*, with the best IC~50~ ≈ 1--5 µM. Moreover, the calculated resistance factors, i.e. the ratio of the IC~50~ values found for the *cisplatin* resistant and sensitive cell lines (A2780R)/IC~50~(A2780) showed that the complexes are able to circumvent the acquired resistance of cancer cells to *cisplatin*, as the resistance factor values are equal to 1.19 (**4**), 1.14 (**5**), 1.21 (**6**) as compared to the value of 2.25 for *cisplatin*. Further, the complexes **1**--**9** were evaluated for their *in* *vitro* anti-inflammatory activity on the model of the LPS-activated THP-1 monocytes. It was found out that the complexes **1**--**7** exhibited the ability to influence the cell-cycle of THP-1 cells resulting in the hormetic effect, and they were also able to significantly attenuate the production of pro-inflammatory cytokines TNF-α and IL-1β at comparable levels as gold(I)-based reference drug Auranofin, but with lower toxicity than this metallodrug. Based on the results of the ESI-MS experiments may be concluded that representative complexes **1** and **6** react with the sulfur-containing substances (cysteine and reduced glutathione) in time-independent manner by the ligand-exchange mechanism based on the substitution of the *N*-ligand (L~n~) by the cysteine or glutathione molecule.

Supporting Information {#s5}
======================

######

**Supporting Information.** The results of elemental analysis, FTIR, ^1^H and ^13^C NMR, and ESI--MS experiments for **1**--**9**. **Table S1.** Crystal data and structure refinements for \[Au(L1)(PPh3)\] (**1**) and \[Au(L3)(PPh3)\] (**3**). **Table S2.** Selected bond lengths and angles in complexes **1** and **3. Figure S1.** Parts of the crystal structure of complex \[Au(L~1~)(PPh~3~)\] (**1**). **Table S3.** Selected non-covalent contacts and their parameters for **1**. **Figure S2.** Parts of the crystal structure of complex \[Au(L~3~)(PPh~3~)\] (**3**). **Table S4.** Selected non-covalent contacts and their parameters for **3**. **Figure S3.** TG/DTA curves of the complexes **1** and **4**. **Figure S4.** Effects of the Au(I) complexes, and Auranofin on the LPS-induced degradation of IκB-α. **Figure S5.** ^31^P NMR spectrum of complex **6**. (DOCX) CCDC Nos. 1010556 and 1010557 contain the supplementary crystallographic data for **1**, and **3**, respectively. These data can be obtained free of charge via <http://www.ccdc.cam.ac.uk/conts/retrieving.html>, or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (44) 1223-336-033; or email: <deposit@ccdc.cam.ac.uk>.

(DOCX)

######

Click here for additional data file.

The authors are grateful to Ms. Kateřina Kubešová for help with the cytotoxicity testing.

[^1]: **Competing Interests:**The authors have declared that no competing interests exist.

[^2]: Conceived and designed the experiments: RK J. Hosek JV ZD ZT. Performed the experiments: RK J. Hosek JV J. Hutyra ZD ZT. Analyzed the data: RK J. Hosek JV ZD ZT. Contributed to the writing of the manuscript: RK J. Hosek JV ZD ZT.


UP worst, Kerala best on health indicators: NITI Aayog

New Delhi, Feb 9: Despite being among the fastest improving states, Uttar Pradesh ranked at the bottom among 21 large states on health indicators as per NITI Aayog's Health Index Rankings announced on Friday. Kerala emerged on top followed by Punjab and Tamil Nadu.

In terms of annual incremental performance, UP ranked third behind Jharkhand and Jammu and Kashmir.

"Jharkhand, Jammu and Kashmir and UP showed the maximum gains in improvement of health outcomes from base (2014-15) to reference year (2015-16) in indicators such as neonatal mortality rate, under-five mortality rate, full immunization coverage, institutional deliveries, and People Living with HIV (PLHIV) on anti-retroviral therapy," the "Healthy States, Progressive India" report said.

However, it noted that while states that start at lower levels of development are generally at an advantage in notching up incremental progress over states with high health index scores, it is a challenge for states with high index scores to even maintain their performance levels.

In overall rankings among the larger states, Punjab jumped three places to knock off Tamil Nadu from the second place. Both Jammu & Kashmir and Jharkhand jumped four places to be placed at 7th and 14th places respectively.

However, despite improving its overall score on the index, Uttar Pradesh could not improve its ranking and was placed at the bottom below Bihar and Rajasthan.

Among the smaller states, Mizoram ranked first followed by Manipur on overall performance. In terms of annual incremental performance, Manipur came out on top followed by Goa.

Among the union territories, Lakshadweep showed both the best overall performance, as well as the highest annual incremental performance.

The NITI Aayog said the index would be linked to incentives under the National Health Mission, which underlines the importance of such an exercise.

While releasing the report, NITI Aayog CEO Amitabh Kant said it would serve as a tool to leverage co-operative and competitive federalism to accelerate the pace of achieving health outcomes.

It would also serve as an instrument for "nudging" states and union territories to put much greater focus on output and outcome-based measurement of annual performance, he said.

Kant added that the next such report on the states' performance will be released in June.

He added that NITI Aayog would also bring out a ranking of 730 district hospitals based on their performance.

"The intention is to give incentives to good performers while shaming those who do not improve by naming them and putting out their details in public domain," he said.

The health ranking of states has been done in three categories namely larger states, smaller states and union territories to ensure comparison among similar entities.

The NITI Aayog said it is a weighted composite index based on indicators in three domains: health outcomes (70 per cent weightage), governance and information (12 per cent), and key inputs and processes (18 per cent).

According to the report, the incremental measurement reveals that about one-third of the states have registered a decline in their performance in 2016 as compared to 2015, stressing the need to pursue domain-specific, targeted interventions.


NORFOLK, Va. (WAVY) — Police say a man was charged over the weekend after he accidentally shot a woman in Norfolk.

Norfolk police said 20-year-old Damon S. Keene is charged with reckless handling of a firearm.

The charge stems from a Saturday incident inside a home in the 1400 block of Gabriel Drive, police said.

Police said that Keene tried to take his gun out of his pocket when he accidentally fired a bullet, which struck a 20-year-old woman. She was taken to the hospital and was expected to be OK.

Keene fled the scene before officers arrived, but returned later and was interviewed by detectives. Keene was formally charged on Sunday, police said.


King County is looking at expanding water taxi service to Ballard, and are asking for feedback to gauge interest.

Currently, the passenger-only water taxi operates only from downtown Seattle to West Seattle and Vashon Island. Now, through a budget proviso, the King County Council has asked Metro to outline a plan for potential service to Ballard and Kenmore.

To that end, Metro has developed a survey to find out whether a ferry service would provide a better commuting option for people traveling between Ballard and downtown Seattle.

The routes being considered are:

Ballard to downtown Seattle and/or the Expedia campus (Interbay)

Kenmore to Seattle (University of Washington, Madison Park, Leschi Park, or the Portage Bay area)

The Metro Water Taxi expansion survey is live now, and will be open until Dec. 23rd.

Photo: KingCounty.gov


Interactions of ficolin-3 with ovarian cancer cells.
Ficolin-3 is a pattern-recognition molecule with the ability to activate the lectin pathway of complement. It is found in lung, liver and blood, but its physiological role is unclear. We have investigated interaction of recombinant ficolin-3 with malignant cells and tissues. Cells of various lines of human origin as well as ovarian tissue sections have been studied with the use of flow cytometry and immunohistochemistry. Recombinant (but not serum-derived) ficolin-3 was found to bind strongly to the ovarian cancer cell lines, SKOV-3, OVCAR-3 and ES-2, at concentrations of 2.5 μg/ml and above. Moreover, His-tagged recombinant ficolin-3 (10 μg/ml) preferentially stained ovarian tissue sections from patients with malignant tumours compared with those from patients without. Binding to cell lines was inhibited by EDTA and specific carbohydrate ligands, indicating involvement of the fibrinogen-like domain. Binding was enhanced under mildly acidic conditions and at physiological pH after pre-incubation of cells with mildly acidic buffer. Basing on data concerning recombinant protein, it may be suggested that ficolin-3 is involved in immune response in ovarian cancer. However, unidentified serum factor(s) seem(s) to protect cancer cells from recognition by natural or rficolin-3.


Q:

GNU Make: how to join list and separate it with separator?

I have this:
FOO = foo1 foo2 ... fooN

and want to get join all these string and separate it with, for instance, colong:
foo1:foo2:foo3:...:fooN

How to do this in GNU Make, without using external UNIX tools?

A:

See the code below.
# A literal space.
space :=
space +=

# Joins elements of the list in arg 2 with the given separator.
#   1. Element separator.
#   2. The list.
join-with = $(subst $(space),$1,$(strip $2))

Usage:
FOO = foo1 foo2 ... fooN

COLON_SEPARATED_FOO := $(call join-with,:,$(FOO))

A:

You can simply replace spaces with colon:
EMPTY :=
SPACE := $(EMPTY) $(EMPTY)
FOO = foo1 foo2 ... fooN
FOO_LIST = $(subst $(SPACE),:,$(FOO))

FOO_LIST is foo1:foo2:...:fooN.


Business

UFC’s celebrity investors have a chokehold on $300M dividend

Ultimate Fighting Championship is draining its cash reserves to fund big payouts to celebrity investors — even as its fighters gripe that they are vastly underpaid, The Post has learned.

The mixed martial arts giant has approved a massive $300 million dividend to UFC’s investors — a star-studded list that includes Mark Wahlberg, Charlize Theron, Gisele Bündchen, Ben Affleck and tennis stars Serena and Venus Williams, sources said.

Roughly half of the $300 million will go to Endeavor, the entertainment holding company run by Hollywood superagent Ari Emanuel, which bought a 50 percent stake in UFC in 2016 for $4 billion. Endeavor also owns Hollywood talent agency WME, fashion and sports media agency IMG and the Miss Universe Pageant.

Wahlberg, the Hollywood producer behind “Entourage” and “Boardwalk Empire,” is slated to nab a roughly half-million-dollar dividend, sources said, while Brazilian model Bündchen — wife of Patriots quarterback Tom Brady — is on track to get $145,000.

Other beneficiaries include UFC president Dana White, Endeavor CEO Emanuel — the inspiration for the Ari Gold character in HBO’s “Entourage” — and Endeavor executive chairman Patrick Whitesell, ex-husband of Jeff Bezos gal pal Lauren Sanchez.

They’re slated to pocket more than $3 million each, sources said.

UFC private equity backers KKR and Silverlake, which own a 42 percent stake in Endeavor, will also cash in, sources said.

The $300 million in payments marks the first time UFC has issued a dividend payment since Endeavor bought a 50 percent stake in 2016, sources said. And its sheer size has jaws agape.

“Most of the time when you do a dividend, you don’t distribute your entire balance sheet,” said a financial source. “If you were going to dividend out 10 percent of your cash, that would be an ordinary course. If you were going to dividend 90 percent of your cash, that’s not.”

Fight for higher pay

The payments will leave UFC with just $50 million as it faces new MMA rivals and allegations by fighters that they’re underpaid compared to athletes of other major sports teams.

Sources tell The Post UFC’s fighters cost the Las Vegas promotions company less than $150 million last year — or under 16 percent of its $900 million in revenue. By contrast, Major League Baseball, the National Basketball Association and the National Football League all share between 48 percent and 50 percent of revenues with their players, data shows.

“It’s different league by league,” Endeavor president Mark Shapiro said. “We pay our fighters significantly more than any other MMA organization. They deserve it. Fighter compensation has gone up commensurately with the success of UFC.”

While UFC fighters might be more appropriately compared to boxers, who get paid per bout, they have argued in legal papers that they’re often prohibited from negotiating for more money because UFC dominates its sport— and uses that leverage to keep wages down.

In 2014, a group of UFC fighters, including Cung Le, Jon Fitch and Kyle Kingsbury, sued UFC’s then-parent company, Zuffa LLC, claiming it engaged in anti-competitive practices intended to drive down their compensation costs, including forcing fighters into long-term contracts that prevent them from fighting elsewhere, and “using its market dominance to coerce fighters to re-sign contracts.”

The case has been slowly winding its way through a Las Vegas federal court, where a judge is weighing whether to grant it class-action status. If that happens, UFC could be on the hook for over $1 billion in damages to over 1,200 fighters who risked getting their faces bashed in on a regular basis, including Conor McGregor and Ronda Rousey. Both McGregor and Rousey were UFC fighters between the class period of 2010 through 2017, lawyers for the plaintiffs have said.

Endeavor IPO flop

The $300 million dividend follows Endeavor’s failed effort to raise $600 million through an initial public offering last fall. Endeavor’s Shapiro denies that the payment has anything to do with the flopped IPO, calling it the “normal course of business” after an “incredible year” at UFC.

But sources tell The Post the dividend is directly tied to Endeavor having yanked its IPO last September amid weak demand that was first reported by The Post.

“Because an idea did not work, the UFC is now going to be the next source of cash until Endeavor goes back out again to try to go public,” said a source close to Endeavor. “It’s now draining the UFC in order to prop up the rest of the company.”

The company has a $4.6 billion debt load and, as The Post reported in December, it faces a horde of dispirited WME talent agents who were counting on the IPO to cash in on stock options they’ve been accepting in exchange for smaller bonuses. Agents have been demanding they be made whole, and Endeavor has been searching for ways to compensate them at least in part before April, as The Post has reported.

Shapiro on Thursday confirmed that Endeavor will use some of its UFC money for a stock buyback program, but said the dividend issuance is “not directly linked to liquidating employees.”

Endeavor also plans to use the money for acquisitions, Shapiro said without providing specifics.

Rivals on horizon

UFC has proven a bright spot in Endeavor’s portfolio with cash flow of more than $200 million a year, sources said. Still, it will take the lucrative fighting club a good year to rebuild its coffers — hindering its ability to expand even as the competition ramps up, sources said.

Rival Bellator, which is owned by ViacomCBS, is seeking to expand in the US, as is One Championship, a Singapore-based league backed by Sequoia Capital that is valued at more than $1 billion due in part to its aggressive footprint in Asia.

Experts say UFC could give up ground if it can’t continue to invest heavily in Asia, make meaningful acquisitions or expand into other areas, like boxing.

But Shapiro tells The Post that there are no plans to move into boxing — and he nixed recent reports that Endeavor might spin off UFC or sell shares of the MMA organization to the public through an initial stock offering.

“We never had any plans to go public. The UFC is a major and valuable interest in Endeavor. It is not in our interest to spin it off or IPO,” he said.

That jibes with what sources have told The Post — namely, that Emanuel doesn’t want to lose his cash-cow UFC business to an IPO or spinoff.

“Ari is adamantly against UFC going public. Endeavor wouldn’t be worth as much,” the source said before adding that they also “don’t want the fighters to know exactly how profitable the UFC is.”

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Confederation Building (Winnipeg)

The Confederation Building is a building in Winnipeg, Manitoba, built by architect J. Wilson Gray. The ten-story office building stands forty-one meters tall (135 ft), and was built in 1913. It was designated a National Historic Site of Canada in 1976, for its Chicago school-influenced architecture.The Confederation Building

This ten story steel-framed office block is representative of early high-rise building construction technology in Winnipeg. Designed in the Chicago style of architecture by J. Wilson Gray of Toronto, it was erected in 1912 by the Carter-Halls-Aldinger Company of Winnipeg at a cost of $400,000. The building was owned and occupied by the Confederation Life Association for over 50 years. Its style, use, and placement within Winnipeg's commercial core make it an enduring symbol of the city's great economic and spatial growth in the early twentieth century - Historic Sites and Monuments Board of Canada

English text from the plaque on the front of the building.

External links
 Confederation Building

References

Category:Buildings and structures in Winnipeg
Category:National Historic Sites in Manitoba
Category:Office buildings completed in 1913
Category:Chicago school architecture in Canada
Category:1913 establishments in Manitoba


A hero lorry driver swerved across the road to block the escape of two violent London moped criminals, moments after they attacked and injured a policeman.

Two males, including a 15-year-old boy, had been speeding through Catford in south London on the stolen vehicle, before driving at the officer and crushing his leg.

The officer and a colleague had spotted the criminals on their rampage and attempted to intervene before the brutal ramming attack.

After the moped was halted by the vigilante lorry driver, the two males split up and escaped on foot before the teenager was apprehended later.

Officers said the Metropolitan Police’s Commercial Vehicle Unit inspected the lorry driver and his vehicle, but he was not arrested or reported for any offences.

Kay Sloane, 36, told The Sun: “It was really frightening. I was putting the children to bed at around 8:15 when I heard this almighty bang. My husband thought it was thunder.

“The moped crashed into the policeman on his bike first – how he’s not killed anyone or himself I don’t know. What would have happened if I was going out or coming in with the children?

“The Waitrose driver swerved into them to stop them. The driver’s intentions weren’t malicious he just wanted to stop them,” the mother added.

https://twitter.com/DanielleDASH/status/1024752937053167617

Twitter user and passerby Danielle DASH captured the aftermath of the incident on video and posted it online.

The Waitrose lorry can be seen angled across the road, blocking most of the pavement, too. A moped is seen on the ground inches from it, along with police cars and an ambulance.

Danielle’s main concern, however, was the wellbeing of the attackers. She wrote: “Hi Waitrose. Is it the policy of your company to encourage your HGV lorry drivers to use their vehicles to crash into alleged moped thieves?

“The two young men fled the scene after the violent act and we are unsure of the severity of their injuries.

She added: “The Waitrose driver acted in a way that could have resulted in these young men’s deaths.”


Chitosan refers to a biodegradable and natural polymer derived from the shells of shrimp and other crustaceans such as lobsters, squids, and crabs. Chemically, it is an organic carbohydrate polymer containing chitin. Chitosan is also derived from fungal source through substrate fermentation. The global chitosan market is growing rapidly due to the wide applications of the polymer across various industries. The global market for chitosan was worth US$1.35 bn in 2013. Expanding robustly at a CAGR of 17.7% during the period between 2014 and 2020, the market is estimated to reach a valuation of US$4.22 bn by 2020.

In terms of applications, the global chitosan market has been segmented into food and beverages, industrial, water treatment, biomedicines and pharmaceuticals, agrochemical, cosmetics, and others including photographic products and fuel cells. The market is driven by the demand from the water treatment application segment, especially across the regions such as North America, Japan, and Europe.

Chitosan is used as a flocculent to treat wastewater. It is an effective coagulant for both organic and inorganic compounds present in water. It is also used as a chelating agent to bind with the highly toxic heavy metals such as lead present in industrial wastewater. Owing to its biodegradability, biocompatibility, and non-toxic nature, the demand for chitosan in wastewater treatment will surge in the coming years, particularly across Asia Pacific where China and Japan are the key markets.

Chitosan is also used in agrochemicals and in the near future, the demand for chitosan for application in agrochemicals is expected to be the highest. The growing demand for organic constituents in cosmetics and wide applications of chitosan in hair care, skin care, and oral care cosmetic formulations are anticipated to define the future outlook of the market. Lately, researchers have found out innovative applications of chitosan. Chitosan-based dietary supplements are being increasingly used due to its weight reduction and fat binding properties. Chitosan nanoparticles are widely used across the pharmaceutical and biomedical sectors owing to their higher reactivity.

Chitosan-based Eco-Friendly Food Packaging Material Developed at National University of Singapore

Researchers from the National University of Singapore have developed an environment-friendly food packaging material derived from chitosan. Free from chemical additives, the natural chitosan-based composite film with grapefruit seed extract can slow down fungal growth, thereby increasing the shelf life of perishable food products such as bread. While chitosan has inherent antifungal and antimicrobial properties, grapefruit seed extract is antioxidant and has anti-viral properties. The novel composite film has flexibility and mechanical strength that are comparable to synthetic polythene film commonly used for food packaging. The film blocks UV light, thereby slowing down oxidation and photochemical reactions that increases the shelf life of food products.


How beta2-selective is the adrenoceptor antagonist drug, IPS 339?
Schild plots for the compound (t-butylamino-3-ol-2-propyl)oximino-9-fluorene (IPS 399) have been obtained on isolated intrinsic tone trachea and atria (rate) og guinea-pigs. Alpha-Adrenoceptors and uptakes were inhibited. The Schild plots for IPS 339 on trachea (fenoterol as agonist) and on atria (noradrenaline as agonist) were not superimposed suggesting that IPS 339 was beta2-selective. The slopes of the Schild plots obtained on intrinsic tone tracheal preparations (isoprenaline or fenoterol as agonist), although greater than 1.0, were not significantly different from that on atria (noradrenaline as agonist). From the average separation of these Schild plots on trachea and atria IPS 339 was assessed to be only 3.3 times more active on beta2- than on beta1-adrenoceptors. The experiments in the literature which showed a high beta2-selectivity for IPS 339 (155 fold) were carried out on carbachol-contracted tracheal preparations (isoprenaline as agonist) and the Schild plot obtained had a very low slope which was quite different from that on atria. Therefore, the results illustrate how the quantitative estimate of the selectivity of a beta-adrenoceptor antagonist can be misleading when Schild plots with different slopes are compared.


On or about 815am on Wednesday morning, May 9, 2013, I approached the tram with my daughter as we do each morning as I take her to preschool in Manhattan as I go to work. That morning, like many mornings, there was no attendant stationed at the booth to open the security gate for parents who have young children with strollers.

That morning, there was a female Public Safety Officer stationed in the booth.... I swiped my card and I asked the Officer if she would open the gate so I could enter with the stroller. My daughter wanted to go in to be with her friends who were waiting for the tram with their mother and she also wanted me to come with her.

The Officer said it was not her job to open the gate. I then asked her if she would contact someone who would and she refused to do that. Meanwhile my daughter was getting anxious to enter to be with her friends and asking me to come.

I told the Officer that other Public Safety Officers open the gate when the tram employees are not around but she still refused and became very un professional and hostile towards me. My daughter started to cry and panic reacting to the Officer’s un professional conduct and temper.

Finally the tram came and the operator opened the gate but the Officer continued her hostility towards me and my poor daughter cried all the way over on the tram. The Officer blamed me for causing my daughter to cry when in fact it was her misconduct which created the situation. Public Safety is to assist the public. It is not their job to treat us like thugs or criminals but to help us.

A few months ago on behalf of the Roosevelt Island Parents Network and the RIRA Island Services Committee I brought up the problem of the access to the Tram boarding area for parents with strollers, which also affects people in wheelchairs and bikers.

Parents with strollers have to wait until someone from the Tram personnel, usually the Tram conductor, comes to open the gate for them. While the cabins are not in the station, all other passengers can go through the turnstiles and pre-load. Especially during long winter months like this year, this can affect the health of the residents and their babies. For example, mothers who just gave birth and their newborns have to wait in the cold in front of the gate and cannot get to the heaters of the pre-boarding area. But even in warmer times, the current situation is unfair and affects timing: Especially in rush hour a parent or another care-giver can miss the Tram even though she or he was the first one at the station – but had to wait for the gate to be opened while everybody else pre-loaded – causing the parent to be late for work or another appointment.

We suggested a number of solutions to RIOC and we are very pleased to have received a positive progress report last week by RIOCʼs Community Relations Specialist Erica Spencer-El. As a short-term solution, an additional staffer has been assigned who should now assist with opening the gate during rush hours. Unfortunately there is still feed-back from parents that this is not always the case. In the long run the plan is to hire an extra person designated to opening the gate and supervising the whole area permanently. We hope this can happen as soon as possible. On behalf of the many parents who live on Roosevelt Island or visit it, as well as the wheelchair- users and bikers we would like to thank RIOC for the progress already made and for further acting on this issue!

The issue of Baby Stroller access to the Roosevelt Island Tram entrance gate

Written by Kim Esop-Wylie and Directed by Mihaela Mihut and Cornelius Horgan, Mythery & Mayhem is set in ancient Greece. According to MST&DA:

To humans 3000 years ago the world was a strange and mystical place. What is lightening? Why are there seasons? Where does fire come from? The ancient Greeks had an answer - it must be the Gods. Travel back to ancient Greece and Mount Olympus to discover the Magic and Mayhem of Mythology. Adventure around in Kim Esop-Wylie's newest creation, a funny and fast paced exploration of nature, love and life in the stories that shaped civilization itself.

... The Cornell NYC Tech ULURP application will now proceed to the full NYC Council and Mayor Bloomberg for expected approval. However, even after final approval from NYC, Cornell NYC Tech still needs to negotiate and come to an agreement with the Roosevelt Island Operating Corp (RIOC) for the approximately 3 acres of land controlled by RIOC within the planned Cornell NYC Tech campus on the site of the current Goldwater Hospital....

This morning, Ms Lappin, NYC Council Speaker and Mayoral Candidate Christine Quinn, Cornell NYC Tech Vice President Cathy Dove, Cornell Dean Dan Huttenlocher, Roosevelt Island Community Coalition (RICC) Co-Chairs Ellen Polivy and Jonathan Kalkin as well as Roosevelt Island resident and Community Board 8 Member Larry Parnes appeared at a Press Conference to announce the vote to approve Cornell NYC Tech coming to Roosevelt Island and to congratulate each other on working together to bring the project to this stage of the process.

followed by questions about barging construction materials, ferry service, ongoing negotiations with the Roosevelt Island Operating Corp (RIOC) for land controlled by RIOC, financial contributions to Roosevelt Island by NYC and more.

Click here for theLetter Agreement detailing the commitments made by Cornell NYC Tech to City Council Member Jessica Lappin, the entire City Council and the Roosevelt Island Community which are included as part of the lease agreement between the NYC and Cornell for the Goldwater Hospital site. In addition to the commitments included in the lease agreement, NYC Deputy Mayor’s Robert Steel sent this letter dated May 2 2012 to Ms. Lappin regarding NYC contributing operating funds for a pilot Roosevelt Island ferry service and NYPD monitoring security concerns for the new campus.

Here's the Press Release from Ms. Lappin's office:

Today, City Council Member Jessica Lappin, Council Speaker Christine C. Quinn, Cornell NYC Tech Vice President Cathy Dove, Cornell NYC Tech Dean Dan Huttenlocher, and Roosevelt Island community members announced that the City Council will vote to approve the Cornell NYC Tech campus on Roosevelt Island.

“Cornell NYC Tech is about the future of New York—it will drive innovation and economic growth for generations to come. When this idea was first proposed, I formed a community coalition to champion building the campus in Manhattan, and specifically Roosevelt Island. The new school will be a tremendous boon, not only for my City Council district, but for the entire Big Apple,” said Council Member Jessica Lappin, whose district includes Roosevelt Island.

Today’s City Council vote is the final step in the city’s land use review process for Cornell NYC Tech, two million-square-foot applied sciences and engineering campus that will be located at the Goldwater Hospital site on Roosevelt Island in Manhattan. The project is expected to have a $33 billion economic impact, creating 48,000 new jobs and 1,000 spin-off companies over the next three decades.[1]

Council Member Jessica Lappin negotiated to secure many commitments from the Administration and Cornell NYC Tech. The city has pledged to contribute funding to a one-year summer ferry service pilot program and has designated an NYPD liaison to ensure continued safety and security on Roosevelt Island. The university has committed to:

· Undertake the most aggressive voluntary use of barging in New York City, which will reduce the number of construction vehicles on Roosevelt Island by more than half, as compared to traditional building techniques.

· Partner with New York City middle schools to create Science, Technology, Education, and Mathematics (STEM) programs, with a special focus on girls. Initial pilot programs will begin at P.S./I.S. 217 on Roosevelt Island, M.S. 406 in East Harlem, and I.S. 204 and P.S. 111 in Long Island City, Queens. Starting this summer, Cornell NYC Tech will sponsor an immersion program for girls in these schools to learn coding, web design, robotics and mobile development under the mentorship of some of the industry’s top engineers and entrepreneurs.

· "Adopt" the local school on Roosevelt Island, PS/IS 217, by providing programming such as teacher training and support, after school programming, tech events, career day options, and hardware and software programming development.

· Conduct air monitoring throughout the project, including demolition, excavation and construction. The results of this monitoring will be posted on the construction website.

· Engage extensively with the Roosevelt Island community by creating a construction task force with residents and others; researching ways that technology can enhance the lives of the disabled and the older adults on the Island; providing computers and training to senior center members and the disabled community; providing a tech fair to advance technology education for the disabled; working with the community to program outdoor space for children.

Speaker Christine C. Quinn said “Cornell NYC Tech is a win for the City of New York, Roosevelt Island community and our local economy. This campus will help New York City continue to attract more of the brightest minds, which in turn, will attract more of the best businesses. The City Council is proud to approve the final steps necessary for construction to commence, and we look forward to the completion of this new state-of-the-art campus.”

Cathy Dove, Vice President, Cornell NYC Tech said “We are extremely grateful for the leadership of Council Member Lappin and Speaker Quinn, for the support of the City Council, and for Mayor Bloomberg's vision. We worked throughout the public review process to incorporate input from the community into our approach– that input has resulted in a stronger campus plan. We are confident the result is a Cornell Tech campus we can all be proud of and that will benefit both the Roosevelt Island community and the entire city. We look forward to breaking ground next year and continuing to work closely with the community as our campus develops.”

Ellen Polivy, Co-Chair of the Roosevelt Island Community Coalition (RICC) said “Council Member Lappin did an amazing job fighting for us. Cornell NYC Tech’s “adoption” of our local school was much appreciated by everybody. I want to thank Cornell for all their work to plan for the future, and we look forward to a successful partnership going forward.”

Jonathan Kalkin, Co-Chair of Roosevelt Island Community Coalition (RICC) said “It’s been exciting to work with Council Member Lappin and Cornell. Cornell NYC Tech will be a game changer for Roosevelt Island and for New York City. We can’t wait to see what innovations will come from the school, and we look forward to greeting our new neighbors.”

The goslings hatched! We got a few snaps of them when we were walking in Four Freedoms Park. We could not get too close because papa gosling was hissing at us mad-like.

Irina Hage snapped this photo and the one at top of page

Image From Irina Hage

remarking:

The new Resident on our Island. They are so cute!

Denise Shull shares these pictures and adds:

Can you believe these and the twisted priority of Four Freedoms Park Conservancy - "Please excuse our barricades" - REALLY?? What happened to Please marvel at this miracle of nature? (Look closely in the 3rd picture to see just that).

Images From Denise Shull

The FDR Park barricade sign says:

Please excuse our temporary barricades.

A mother Canada goose is currently nesting under one of our Littleleaf Linden Trees. The father Canada goose is protecting the nest and can be very aggressive.

To protect both the goose family and our visitors we have barricaded the nest until the eggs hatch. Please respect the barricades and follow the directions of Four Freedoms Park Rangers during this time.

Dogs can be in strollers but not babies Incredible!! RIOC allows dogs in strollers to ride the Red Bus (and they sure can walk), but they make parents close the strollers and carry their babies during rush hours.... So now dogs have more rights than persons?

Hand baggage, instrument cases, bundles and parcels may be carried onto RIOC buses by customers provided that these articles can be carried on without inconveniencing other customers and/or the driver and does not obstruct the bus aisle or doors. Persons with unfolded strollers and/or shopping carts are not permitted to board RIOC buses unless the driver, at his or her sole discretion, determines that it will not cause an inconvenience or safety hazard to other customers or the driver. Orthopedic (wheelchair type) strollers for children with disabilities are permitted on buses just as wheelchairs are and must be secured in the wheelchair area. Bus operators must check strollers carefully to ensure that children with disabilities are not denied accessible service.

Updated signage has been posted on buses stating:

"Strollers and Shopping Carts must be folded upon operator’s request".

PLEASE TAKE NOTICE that a meeting of the Real Estate Development Advisory Committee of the RIOC Board of Directors will be held on Monday, May 6, 2013 at 5:30 p.m. at the RIOC administrative office, 591 Main Street, Roosevelt Island, New York.

The NYC Council Land Use Committee approved the Roosevelt Island Cornell NYC Tech ULURP application last Thursday May 2. The Cornell NYC Tech ULURP application will now proceed to the full NYC Council and Mayor Bloomberg for expected approval. However, even after final approval from NYC, Cornell NYC Tech still needs to negotiate and come to an agreement with the Roosevelt Island Operating Corp (RIOC) for the approximately 3 acres of land controlled by RIOC within the planned Cornell NYC Tech campus on the site of the current Goldwater Hospital.

Click here for theLetter Agreement detailing the commitments made by Cornell NYC Tech to City Council Member Jessica Lappin, the entire City Council and the Roosevelt Island Community which are included as part of the lease agreement between the NYC and Cornell for the Goldwater Hospital site. In addition to the commitments included in the lease agreement, NYC Deputy Mayor’s Robert Steel sent this letter dated May 2 2012 to Ms. Lappin regarding NYC contributing operating funds for a pilot Roosevelt Island ferry service and NYPD monitoring security concerns for the new campus.

As previously reported, the NYC Land Use Committee held Public Hearing on the Cornell NYC Tech application on April 30. Witnesses at the hearing were representatives from NYC, Cornell NYC Tech and Roosevelt Island residents. Click here for the full video of the hearings. (the video is from the NYC Council and includes all Land Use Committee hearing for the day. The Roosevelt Island Cornell NYC Tech hearing begins at the 2 hour 28 minute mark).

Just about everything you want to know about the Cornell NYC Tech project and how it will impact Roosevelt Island is discussed in the video. Watch it.

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WELCOME TO ROOSEVELT ISLAND

Welcome to the Roosevelt Islander Online!

Roosevelt Island is a mixed income, racially diverse waterfront community situated in the East River of New York City between Manhattan and Queens and is jurisdictionally part of Manhattan. The Roosevelt Island Tramway, which connects Roosevelt Island to the rest of Manhattan, has become the iconic symbol of Roosevelt Island to its residents.

The Purpose of this Blog is to provide accurate and timely information about Roosevelt Island as well as a forum for residents to express opinions and engage in a dialogue to improve our community.