Louis, MO, USA). ε-caprolactone (CL) were obtained from Acros Organics (Geel, Belgium). this website thiolated chitosan Selleckchem NVP-BGJ398 (Mw 33000 Da) was from NanoMed Biotech Co. Ltd (Shenzhen, China). Poly(ε-caprolactone) (PCL) (MW 42000 Da), and stannous octoate (Sn(OOCC7H15)2) were also purchased from Sigma (St. Louis, MO, USA). Paclitaxel powder of purity 99.9% was from BioOne Biotech Co. Ltd (Shenzhen, China). Fetal bovine serum was received from Gibco (Life Technologies, AG, Switzerland).
Methanol and acetonitrile were obtained from EM Science (Mallinckrodt Baker, USA). Deionized (DI) water produced by Millipore Water Systems (Millipore Corporation, Billerica, USA) was utilized throughout all experiments. Synthesis and characterization of PLA-PCL-TPGS random copolymer PLA-PCL-TPGS random copolymers were synthesized from ε-caprolactone, lactide, and TPGS in the presence of stannous octoate as a catalyst via ring opening polymerization. In short, pre-weighted amounts of ε-caprolactone, ACY-1215 cell line lactide, TPGS, and one drop of stannous octoate were added in a flask. The mixture was heated to 145°C and allowed to react for approximately 16 h. Synthesis was carried out under an oxygen- and moisture-free environment. The product was dissolved
in dichloromethane (DCM) and then precipitated in excess cold methanol to remove unreacted monomers and TPGS. The final product was collected by filtration and dried under vacuum. The TPGS content and number-averaged molecular weight of the copolymer was determined by 1H NMR in CDCl3 at 300 Hz (Bruker ACF300, Bruker AXS Pte Ltd., Singapore). Preparation of thiolated chitosan-modified all paclitaxel-loaded nanoparticles Nanoparticles were prepared by a modified solvent extraction/evaporation technique [29, 30]. In brief, 11 mg of paclitaxel powder and 100 mg of PLA-PCL-TPGS copolymer were weighed and dissolved in 6 ml DCM. The organic solution was immediately poured into 100 ml of 0.03% (w/v) TPGS solution under mild stirring. The mixture was then sonicated for
90 s at 30 W output to form water-in-oil emulsion. The emulsion was further evaporated under ambient conditions overnight to remove DCM. The nanoparticles were harvested by centrifugation at 80,000×g for 15 min and then washed three times to remove the emulsifiers and unentrapped drug. The resulting nanoparticles were finally resuspended in 5 ml of deionized water and lyophilized. The PLA-PCL-TPGS nanoparticles was further modified by thiolated chitosan using a method described previously . Preweighed thiolated chitosan was dissolved in deionized water at a concentration of 0.5 mg/ml. The nanoparticles were suspended in thiolated chitosan solution at a concentration of 9.5 mg/ml by sonication at 30 W power output for 30 s in an ice bath, and then were collected by centrifugation at 80,000×g for 15 min. The coumarin-6-loaded nanoparticles were prepared by encapsulation of 0.1% (w/v) coumarin-6 instead of paclitaxel.