Synthesis of Azido-Functionalized Folate
Preparation of PLA-PEG-FA copolymer was started with the synthesis of OH-PLA-PEG-COOH according to the method described by Hami et al (Hami, Amini, Ghazi-Khansari, Rezayat, & Gilani, 2014b). Briefly, definite amounts of lactide monomers was added to HO-PEG-COOH under dry argon, then the reaction was continued at 140 °C for 24 h in the presence of stannous octoate (Sn(Oct)2) as catalyst. The mixture was dissolved in chloroform and precipitated by a mixture of ether and methanol (v/v= 1:1). Then, the HO-PLA-PEG-COOH obtained as precipitate was dried. This obtained polymer was dissolved in DCM with DCC (1.2 mmol) and NHS (0.6 mmol). The product was stirred at 0 °C and 25°C for 1 h and 24 h respectively. Triethylamine (TEA) and propargylamine (0.6 mmol) were added into the above product. Next, the mixture was stirred at 25°C for another 24 h. The product (HO-PLA-PEG-alkyne copolymer) was precipitated in cold ether, filtered and dried at 30 °C under vacuum. Then, the HO-PLA-PEG-alkyne copolymer (0.1 mmol) was conjugated with acryloyl chloride (C3H3ClO) (0.2 mmol) in dry toluene containing triethylamine (C6H15N) (0.2 mmol). The sample was stirred for 10 h at 80 °C. After that, the product was cooled to 25°C, filtered and precipitated in n-hexane then dried in vacuum oven to produce acrylate-PLA-PEG-alkyne copolymer. Acrylate-PLA-PEG-Folic acid and Acrylate-PLA-PEG-Glucose were synthesized stepwise by the addition of Acrylate-PLA-PEG-alkyne copolymer (0.1 mmol) and Azido-functionalized folate and Azido-functionalized glucose in 30 mL aq. NH4HCO3 (10 mM) separately. Then, sodium ascorbate and CuSO4 solutions were added to acrylate-PLA-PEG-alkyne copolymer and stirred with a rotator at 25 °C for 24 h. Next, the product was filtered with a 0.22 μm membrane filter. The reaction solutions were diluted with sufficient NaCl aqueous solution, and then extracted several times by dichloromethane. The resulting products were concentrated under vacuum and precipitated in cold ether. Finally, acrylate-PLA-PEG-FA copolymer and acrylate-PLA-PEG-Glu copolymers were dried under vacuum.