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 (0.5 g, 2.47 10-3 mol, 2N) in dry CH2Cl2 (200 mL) under argon. The resolution was stirred for ten min at area temperature. DX (1.0 g, 1.24 10-3 mol, 1N) was added in conjunction with a catalytic volume of DMAP (0.15 g, 1.24 10-3 mol, 1N) and also the mAChR2 Storage & Stability reaction mixture was stirred at space temperature for an added 5 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 via a fritted funnel, along with the filtrate was evaporated beneath vaccuo. The crude solution was purified by preparative TLC in CHCl3: MeOH (95:five). The silica gel was removed by filtration through a fine fritted funnel as well as the filtrate was evaporated below vaccuo to give the preferred 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), three.86 (d, 1H, four), 4.12 (d, 1H, 2), four.2 (t, 1H, HBrC1″), 4.26 (t, 2H, 13), four.88 (d, 1H, 10), five.2 (d, 2H, 20), five.22 (d, 1H, 2′),Adv Healthc Mater. Author manuscript; available in PMC 2014 November 01.Feng et al.Page5.62 (d, 1H, 3′), 7.22.53 (m, 8H, r-H268 and Ar-H305), eight.05 (d, 2H, rH25,29). 13C NMR (one hundred MHz, CD3OD): (ppm) = eight.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 ( 6,14), 43.1 ( 15), 44.five ( 3), 45 ( HBr), 46.4 ( 3′), 57.5 ( 8), 71.eight ( 13), 72.1 ( 7), 74.four ( 2), 75 ( 10), 75.3 ( 20), 78.9 ( 6′), 79.9 ( 1), 80.9 (C4), 84.two ( five), 126.3 ( 31,33,35), 128.9 ( 32,34), 129.2 ( 26,28), 130.two ( 24,25,29), 133.6 ( 27), 135.5 ( 11), 138.9 ( 12), 154.two ( 5′), 167 ( 23), 167.3 ( 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 identify the mz of your final synthetic conjugate solution by Thermo Scientific TSQ Quantum Access with constructive ionization. The mz with the observed molecular ion was 1125, which clearly corresponded for the H adduct of 2-Br-C16-DX. The 2-Br-C16-DX concentrations have been quantified by HPLC utilizing a Finnigan Surveyor HPLC technique using a Photodiode Array (PDA) detector, autosampler and LC pump plus with a InertsilODS-3 column (four , four.six 150 mm, GL Sciences) at 25 . Chromatographic separation was achieved by gradient elution working with mobile phase 2-propanol, acetonitrile (ACN) and water (5: 55: 40 vvv). The flow rate was 1.0 mLmin and the total run time was 25 min for every single 25 injection. The wavelength was 230 nm. The DX Cathepsin B site 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 make a final concentration of 10 mL. The mixture was incubated at 37 within a water bath shaker. At designated time points, one hundred 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 evaluation.[4] The 2-Br-C16-DX remaining at any time point was calculated as 100 the ratio of remaining drug amount towards the total drug spiked into this.
