This work describes several synthetic approaches to append organic functional groups to gold and silver = 7. 16.19) is cytotoxic to the human being leukemia cell collection CCRF-CEM but the ligand 11 alone (IC50 = 981.65) is not. The low toxicity Raddeanin A of the ligand compared to the complex reconfirms the Au ion is the active agent. (7) Conjugation to the alkyne group on 13 is possible via copper catalyzed azide-alkyne cycloaddition. A test reaction Raddeanin A with benzyl azide was carried out inside a NMR tube and monitored via 1H NMR and IR spectroscopy. Complex 13 benzyl azide 10 PPh3CuOAc and CDCl3 were added to an NMR tube. After intervals of 10 min 24 h and 48 h IR and 1H NMR spectra were acquired. The IR spectra show the absorption at 2100 cm? 1 attributable to the Raddeanin A azide stretching mode diminishes overtime and disappears after 48 h. In addition fresh resonances attributable to the triazole-NHC-Au complex 14 appear in the 1H NMR spectra; most notable are the methylene resonances at 5.49 ppm Raddeanin A 4.77 ppm Raddeanin A and 3.62 ppm. Conclusions This statement recounts attempts in developing carboxylic acid and alkyne functionalized NHC-metal complexes for biomolecule conjugation. The carboxylic acid functionalized mono NHC-AuI complex was not isolable but led to the reliable isolation of the water stable and water soluble Na[(NHC)2AuICl] complex 4. Although separation from gold particles generated in the reaction is a challenge the = 1.9 Hz HC5) 7.38 (d 2 = 1.9 Hz HC4) 6.95 (s 4 = 7.6 Hz 4 = 7.7 4 = 6.0 Hz 2 = 6.3 2.5 Hz 2 HC7) 2.31 (s 3 = 3.4 = 1.5 Hz 1 = 1.7 Hz 1 = 6.4 Hz 2 = 6.1 Hz 2.5 Hz 2 = 2.6 Hz 1 = 6.2 Hz 2 = 6.4 1.7 Hz 2 = 1.9 Hz 1 = 1.6 Hz HC5) 6.94 (s 2 = 1.6 Hz = 6.2 Hz 2 = 6.7 2.4 Hz 2 = 2.9 = 1.8 Hz 1 = 1.7 Hz 1 = 6.4 Hz 2 = 6.3 2.4 Hz 2 = 2.6 Hz 1 = 1.9 Hz = 1.9 Hz = 6.4 Hz 2 = 6.4 2.6 Hz 2 = 2.5 = 7.0 Hz = 7.0 Hz 4 2 Hz = 2.0 Hz = 7.0 Hz 2 = 7.0 Hz 2 HC11) 3.49 (s 3 HC4). Biological Studies Cell tradition The cell lines DLD-1 (human being colorectal adenocarcinoma) CEM (T cell leukemia) and Ramos (Burkitt’s Lymphoma) were cultured relating to ATCC specifications in RPMI-1640 medium. The cell lines MCF-7 (human being breast adenocarcinoma) HEK (human being embryonic kidney) and Hep-G2 (human being hepatocellular carcinoma) were cultured relating to ATCC specifications in DMEM medium. Both media were supplemented with 10% fetal bovine serum (Invitrogen) and the cells were incubated at 37 °C in 5% CO2. Cytotoxicity studies using MTS Mouse monoclonal to CD4.CD4 is a co-receptor involved in immune response (co-receptor activity in binding to MHC class II molecules) and HIV infection (CD4 is primary receptor for HIV-1 surface glycoprotein gp120). CD4 regulates T-cell activation, T/B-cell adhesion, T-cell diferentiation, T-cell selection and signal transduction. assay All cell lines analyzed were treated at the following concentrations of 9 (dissolved in DMSO to obtain a stock concentration of 20 mM): 0.5 μM 1 μM 2 μM 4 μM 8 μM 16 μM 32 μM and 64 μM respectively for 48 h at 37 °C. In addition to treatments cells were treated with two settings: (a) cells in press and (b) DMSO (0.01%). Approximately 100 uL of 2 0 0 (depending on the growth characteristics of the cell collection) freshly collected cells were added to the inner 60 wells of its own 96-well plate. After 24 h incubation 80 μL of older media was removed from each well and then 100 uL of each concentration of 9 was added to six wells of each of the 96-well plates. Each of the plates was subjected to this treatment and incubated for 48 h at 37 °C in 5% CO2. After 48 h 100 μL of the drug treatment was eliminated and replaced with 120 uL of MTS dye (MTS = 3-(4 5 diluted with PBS buffer (to 15% MTS). The assay was allotted 2 h at 37 Raddeanin A °C for development. After incubation with the MTS dye the 490 nm absorbance of each well was read on a Tecan plate 110 reader. Each cell measurement experienced the treatment background subtracted before analysis. Quantitative and statistical analyses were performed using the Origin 8.5 software. The same methods were used to study 6 to assess whether the proligand displayed any cytotoxicity. ? Fig. 1 A few examples of the cytotoxicity of metal-NHC complexes towards breast tumor cell lines.2 10 Fig. 2 Solid state structure of 13 with ellipsoids drawn and the 50% probability level. Supplementary Material ESIClick here to view.(2.8M pdf) Acknowledgements The authors thank UF for monetary support. The Paul Tarrant Faculty Fellowship awarded to ASV contributed monetary support to the work. This work is also supported from the National Institutes of Health (GM079359 and CA133086). MEG thanks the UF Technology for Life System and Howard Hughes Medical Institute for providing.