Paragon 1000 spectrometer

The click reaction between Cs-N₃ and PLLD-G3 was carried out as follows: 0.515 g of PLLD-G3 (0.29 mmol) and 0.05 g of Cs-N₃ were added successively with magnetic stirring to 10 mL of DMSO. After a clear solution was obtained, 18 mg of CuSO₄·5H₂O and 28 mg of sodium ascorbate were added successively under a nitrogen atmosphere. The resultant reaction mixture was heated to 40°C for 48 hours. After the reaction, the product was dialyzed in distilled water for 3 days (MWCO =8,000) and lyophilized to obtain the dendronized chitosan derivative (Cs-g-PLLD, yield 73%). FTIR (Perkin–Elmer Paragon 1000 spectrometer) and ¹H NMR (Bruker DPX-300 NMR spectrometer) analyses were used to confirm the click reaction between Cs-N₃ and PLLD-G3. Based on elemental analysis, the degree of substitution (the number of polyamidoamine dendrons per 100 anhydroglucose units of chitosan) of PLLD-G3 on chitosan was determined to be 10.35.

Conjugation between Cs-g-PLLD and FA was carried out as follows: 10 mL of DMSO, 0.359 g of FA (0.81 mmol), 0.168 g of dicyclohexylcarbodiimide, and 0.102 g of N-hydroxysuccinimde were added successively with magnetic stirring for 5 hours at 25°C. After filtration, a clear solution was obtained, and then, 0.5 mL of solution and 30 mg of Cs-g-PLLD were added successively with magnetic stirring for 48 hours. After the reaction, the product was dialyzed in distilled water for 3 days (MWCO =8,000) and was lyophilized to obtain the dendronized chitosan derivative (Cs-g-PLLD-FA, yield 61%). FTIR (Perkin–Elmer Paragon 1000 spectrometer) and ¹H NMR (Bruker DPX-300 NMR spectrometer) analyses were used to confirm this conjugation between Cs-g-PLLD and FA.