Ocetaxel (2-Br-C16DX)[7] A flame-dried round-bottom flask was charged with (-
Ocetaxel (2-Br-C16DX)[7] A flame-dried round-bottom flask was charged with (-2-bromohexadecanoic acid (0.62 g, 1.85 10-3 mol, 1.5N) and DCC (0.five g, 2.47 10-3 mol, 2N) in dry CH2Cl2 (200 mL) beneath argon. The resolution was stirred for ten min at space temperature. DX (1.0 g, 1.24 10-3 mol, 1N) was added as well as a catalytic level of DMAP (0.15 g, 1.24 10-3 mol, 1N) plus the reaction mixture was stirred at area temperature for an extra five min. The reaction was monitored by TLC (CH2Cl2: MeOH 95:5 vv; Rf = 0.58) for completion. The white precipitate of dicyclohexyl urea byproduct was filtered by means of a fritted funnel, as well as the filtrate was evaporated under vaccuo. The crude solution was purified by preparative TLC in CHCl3: MeOH (95:five). The silica gel was removed by filtration via a fine fritted funnel plus the filtrate was evaporated beneath vaccuo to give the desired solution as a white powder (0.4 mg, 86 ). 1H NMR (400 MHz, CDCl3): (ppm) = 0.eight (t, 3H, H3(CH2)14), 1.05 (s, 6H, 16,17), 1.16 (s, 9H, 7”), 1.19 (s, 3H, 19), 1.23 (m, 28H, (CH2)14CH3), 1.68 (s, 3H, 18), 1.78 (m, 2H, 14), 1.67 (d, 2H, H2C1″), 1.87 (s, 3H, H22), two.24 (m, 1H, 3), two.38 (s, 1H, 7), 3.86 (d, 1H, 4), four.12 (d, 1H, two), four.two (t, 1H, HBrC1″), 4.26 (t, 2H, 13), four.88 (d, 1H, 10), five.2 (d, 2H, 20), 5.22 (d, 1H, 2′),Adv Healthc Mater. Author manuscript; offered in PMC 2014 November 01.Feng et al.Page5.62 (d, 1H, 3′), 7.22.53 (m, 8H, r-H268 and Ar-H305), 8.05 (d, 2H, rH25,29). 13C NMR (one hundred MHz, CD3OD): (ppm) = 8.9 ( 19), 14.1 ( H3(CH2)20), 20.9 (C18), 22.6 ( 22), 23.7 (CH2)19CH2CH3), 27 ( 16,17), 28.1 ( 7”), 29.six ((CH2)14C1″), 31.9 ( six,14), 43.1 ( 15), 44.5 ( three), 45 ( HBr), 46.4 ( 3′), 57.5 ( eight), 71.eight ( 13), 72.1 ( 7), 74.4 ( 2), 75 ( 10), 75.3 ( 20), 78.9 ( 6′), 79.9 ( 1), 80.9 (C4), 84.two ( five), 126.three ( 31,33,35), 128.9 ( 32,34), 129.2 ( 26,28), 130.2 ( 24,25,29), 133.6 ( 27), 135.5 ( 11), 138.9 ( 12), 154.2 ( 5′), 167 ( 23), 167.three ( 21), 169 ( 1), 169.7 ( 1″), 211.five ( 9). Characterization of DX and DX conjugates Electrospray Ionization (ESI) coupled with direct injection was employed to determine the mz with the final synthetic conjugate solution by Thermo Scientific TSQ Quantum Access with constructive ionization. The mz from the observed molecular ion was 1125, which clearly corresponded to the H adduct of 2-Br-C16-DX. The 2-Br-C16-DX concentrations had been quantified by HPLC utilizing a Finnigan LPAR1 Species Surveyor HPLC program using a Photodiode Array (PDA) detector, autosampler and LC pump plus using a InertsilODS-3 column (four , 4.6 150 mm, GL Sciences) at 25 . Chromatographic HSP70 web separation was accomplished by gradient elution using mobile phase 2-propanol, acetonitrile (ACN) and water (5: 55: 40 vvv). The flow price was 1.0 mLmin as well as the total run time was 25 min for each 25 injection. The wavelength was 230 nm. The DX concentration was quantified by LCMSMS as described previously.[4] 2-Br-C16-DX digestion in fresh mouse plasma The esterase digestion study was performed in fresh BALBc mouse plasma. The 2-Br-C16DX NPs (0.5 mgmL) have been spiked in to the plasma to create a final concentration of ten mL. The mixture was incubated at 37 within a water bath shaker. At designated time points, 100 of digestion mixture was removed. The concentration of 2-Br-C16-DX was determined by Hybrid-SPE precipitate system as described previously followed by HPLC analysis.[4] The 2-Br-C16-DX remaining at any time point was calculated as one hundred the ratio of remaining drug amount towards the total drug spiked into this.