LabBook

Aim:

As described above, we wanted to synthesize different types of peptide-based nanoparticles based on different amino acids. The steps are as followed:

Figure. F-1. The procedure of synthesis.

Step 1: Synthesis of a-amino acid NCAs (N-carboxyanhydrides) monomer

The process of synthesizing each monomer is slightly different from each other due to different chemical structures. Here we take the synthesis of modified glutamate* NCA as an example.

  1. In a glove box, modified glutamate* NCA and triphosgene were weighted out and dissolved in anhydrous THF.
  2. The resulting mixture was taken out the glove box and stirred at 55 ℃ for 2 hours.
  3. Then the reaction mixture was added to anhydrous hexane, causing the precipitation of NCA, and recrystallized at -20℃.
  4. The NCA powder was obtained by removing hexane via filtration, followed by drying in vacuo.
    5. *: patent confidentiality

Step 2: Synthesis of PEG-block-peptide block copolymer

Figure. F-2. Ring-opening polymerization.

We will synthesize PEG-block-peptide block copolymer via ring-opening polymerization (ROP) using amine modified polyethylene glycol (PEG -NH2) as the initiator as shown above (Figure. F-2.).

  1. In a glove box, PEG-NH2 and NCA were weighted out and placed in two different flasks. Anhydrous DMF was added to the two flasks.
  2. The PEG-NH2 solution was then transferred to the NCA solution and the resulting mixture was taken out from the glove box and stirred at room temperature under argon atmosphere.
  3. After stirring for 72 h, the reaction mixture was dialyzed against methanol for 2 days, followed by dialyzing against deionized (DI) water for 1 day.
  4. The solution was freeze-dried to yield the product as a white powder.

Figure. F-3. Drug solution preparation.

Step 3: Synthesis and Analysis of Polymeric Nanoparticles

We used different processes including film rehydration, nanoprecipitation, and solvent displacement methods to prepare the polymeric nanoparticles. The block copolymer would self-assemble to form nanoparticles in aqueous solutions due to the hydrophobic interactions exerted by the polypeptide segments.