Waters 1515 2414

The transactivating transcriptional activator (TAT) peptide (YGRKKRRQRRRC) with purity ≥97% was purchased from ChinaPeptides Co., Ltd. (Shanghai, China). Taxol injection was obtained from Yangtze River Pharmaceutical (Group) Co., Ltd. (Jiangsu, China). Hochest33342 and 4% paraformaldehyde were provided by Solarbio Science and Technology Co., Ltd. (Beijing, China). Acetonitrile for high-performance liquid chromatography (HPLC) was obtained from Tedia Company, Inc (Fairfeld, OH, USA). Anhydrous pyridine, 1-(2-Hydroxyethyl)-1H-pyrrole-2,5-dione (HEMI), PEG (Mw2000) and D, L-lactide were obtained from Aladdin Bio-Chem Technology Co., LTD (Shanghai, China). Dulbecco’s Modified Eagle Medium (DMEM) (high glucose) cell culture medium, penicillin/streptomycin stock solutions, fetal bovine serum (FBS) and Trypsin were all bought from Gibco BRL (Gaithersberg, MD, USA). Protonic nuclear magnetic resonance (¹H-NMR) (400 MHz) was used to confirm the obtained compounds (Bruker AVVANCE DRX-400 NMR spectrometer, Bruker, Switzerland). In addition, gel permeation chromatography (GPC) (Waters 1515–2414, Waters, USA) was used to analyze all polymers by using tetrahydrofuran as the mobile phase (flow rate: 1 mL/min) at 30 °C. Polystyrene was used as standards.

¹H NMR spectra were recorded on a Bruker AVANCE III 400 MHz superconducting Fourier (Bruker, Billerica, MA, USA) in deuterium generation reagent with tetramethyl silane (TMS) as the internal standard for structure characterization of polymers. The number average molecular weight (M_n) was detected by gel permeation chromatography (GPC) (Waters 1515/2414, Waters, Milford, MA, USA), using THF as the mobile phase with a flow rate of 1.0 mL/min and polystyrene (PS) as the standard for calibration. The morphological characterization of the polymeric micelles was characterized with a HT7700 transmission electron microscopy (TEM, Hitachi, Japan). The sizes and zeta potential of the block polymers were measured by dynamic light scattering (DLS) with a Zeta PALS zeta potential and granularity analyzer (Brookhaven, New York, NY, USA). The fluorescence spectra were examined on a FluoroMax-4 fluorescence spectrometer (HORIBA Jobin Yvon, Clifton Park, NY, USA). UV–Vis spectra were performed on a UV2450 spectrophotometer (Shimadzu, Kyoto, Japan). The copper residues in the polymer were characterized by an Escalab 250Xi X-ray photoelectron spectroscopy (XPS, Thermo Fisher, West Sussex, UK).

¹H NMR spectra were recorded at 400 MHz (Bruker AVVANCE DRX-400 NMR spectrometer, Bruker, Switzerland). CDCl₃ and DMSO-d₆ were used as deuterated solvents. Chemical shifts (δ, ppm) were determined with internal solvent signal as reference (CHCl₃: δ = 7.26, DMSO-d₆: δ = 2.50). Monomer conversion during polymerization was determined based on the concentration of residual monomer using gas chromatography (GC) (FULI9720 series, RBX-5 capillary column 30 m × 0.32 mm × 1.0 μm). The column temperature was 150 °C (15 min). N₂ was used as the eluent at a flow rate of 1.0 mL min⁻¹. Gel permeation chromatography (GPC) (Waters 1515–2414, Waters, Milford, MA, USA) was used to determine the number- and weight-average molecular weights (M_n and M_w) and polydispersity (M_w/M_n) of copolymers in THF at 30 °C with 1 mL min⁻¹ flow rate. Polystyrene standards were used to calibrate the instrument. Transmission electron microscopy (TEM) images were taken with a Hitachi HT7700 instrument operating at an accelerating voltage of 120 KV. The particle size and polydispersity index (PDI) of polymeric nanoparticles were determined by dynamic light scattering (DLS) equipped with a Malvern Nano ZS90 Zetasizer, a 632.8 nm He-Ne laser, and a 173° backscatter detector.