Olvents: CDCl3 (seven.26 ppm), d6-DMSO (two.49 ppm) for 1H NMR spectra; CDCl3 (77.0 ppm) or d6-DMSO (39.five ppm) for 13C NMR spectra (see also Figures S3-S6). 1H- and 13C-assignments had been determined by COSY and HSQC experiments. MS experiments have been carried out on the Finnigan LCQ Advantage MAX ion trap instrument. Analytical thin-layer chromatography (TLC) was carried out on Marchery-Nagel Polygram SIL G/UV254 plates. Flash column chromatography was carried out on silica gel 60 (70-230 mesh). All reactions had been carried out below argondx.doi.org/10.1021/bc400513z | Bioconjugate Chem. 2014, 25, 188-required for multicolor single-molecule FRET studies and is currently undertaken in our laboratory.Bioconjugate Chemistry environment. Chemical reagents and solvents were bought from business suppliers and made use of with out even more purification. Natural solvents for reactions were dried overnight in excess of freshly activated molecular sieves (four ?. 2-O-(2-Azidoethyl)uridine (2). 2,2-Anhydrouridine one (565 mg, two.five mmol) was coevaporated with dry pyridine 3 times and stored over P2O5 in the desiccator for four hrs prior to use. Then, compound 1 was suspended in DMA (4 mL) and BF3 Et2 (785 L, 6.25 mmol) was added below argon and heated to 120 . 2-Azidoethanol (1250 mg, 14.3 mmol) was injected in to the option as well as the mixture was refluxed for sixteen h. Following the response was finished solvents were removed in vacuo, and the oily residue was redissolved in methanol and adsorbed on silica gel.Formula of 3-Bromopiperidine-2,6-dione Compound 2 was purified by column chromatography on SiO2 with CHCl3/CH3OH, 95:5. Yield: 431 mg of two like a white reliable (fifty five ). TLC (CH2Cl2/CH3OH = 85:15): Rf = 0.XantPhos Pd G4 Order 51. 1H NMR (300 MHz, DMSO): 3.17 (m, 2H, H1-C(2) H2-C(two)); 3.58 (m, 2H; H1-C(five) H2- C(five)); three.86 (m, 2H, H1-C(one) H2-C(one)); 3.88 (m, 1H, H- C(4)); 4.04 (m, 1H, H-C(2)); 4.60 (dd, J = four.eight Hz, J = 9.eight Hz, 1H, H-C(three)); 5.14 (m, 2H, HO-C(three), HO-C(5)); five.72 (d, J = 8.0 Hz, 1H, H-C(5)); five.88 (d, J = 4.8 Hz, 1H, H- C(1)); 7.94 (d, J = 8.0 Hz, 1H, H-C(six)); eleven.29 (s, 1H, NH) ppm. 13C NMR (150 MHz, DMSO): 49.93 (C(two)); 60.39 (C(5)); 68.two (C(3)); 68.PMID:23892746 86 (C(one); 81.31 (C(2); 84.93 (C(four)); 86.15 (C(1)); 101.79 (C(five)); 140.32 (C(six)); 150.56; 163.ten ppm. ESI-MS (m/z): [M-H]- calcd for C11H15N5O6, 312.11; observed 312.46. 2-O-(2-Azidoethyl)-5-O-(4,4-dimethoxytrityl)uridine (2a). Compound two (372 mg, 1.19 mmol) was coevaporated with dry pyridine three times and dissolved in pyridine at room temperature and under argon atmosphere. four,4-Dimethoxytrityl chloride (443 mg, 1.31 mmol) was added in two portions more than a period of two h. Stirring was continued overnight, and when TLC showed total reaction, methanol was extra as well as the alternative was evaporated in vacuo. The residue was dissolved in CH2Cl2 and extracted with 5 citric acid, water, and saturated NaHCO3. The organic layer was dried more than Na2SO4 and evaporated. The crude product or service was purified by column chromatography on SiO2 with CH2Cl2/CH3OH, a hundred:0 to 98:2. Yield: 549 mg of 2a as a white foam (75 ). TLC (CH2Cl2/CH3OH = 92/8): Rf = 0.54. 1H NMR: (300 MHz, CDCl3): two.58 (d, J = 9.7 Hz, 1H, HO-C(three)); three.49 (m, 2H, H1-C(two), H2-C(2)); three.58 (m, 2H; H1-C(5), H2-C(5)); three.80 (s, 6H, H3CO) 3.96 (m, 1H, H2-C(one)); 3.96 (m, 1H, H-C(two)); four.04 (m,1H, H-C(4)); four.19 (m, 1H, H1-C(1)); 4.51 (m, 1H, H-C(3)); five.thirty (d, J = eight.one Hz, 1H, H-C(5)); 5.93 (s, 1H, H-C(1)); six.85 (m, 4H, H-C(ar)); seven.31 (m, 9H, H-C(ar)); 8.09 (d, J = 8.one Hz, 1H, H-C(six)); 9.sixteen (s, 1H, N- H) ppm. 13C NMR (150 MHz, CDCl3): 50.99 (C(two)); 55.forty (CH3O); 61.03 (.
