Synthesis PEGu2k

1. Dissolve PEI 2K (0.010 mol amine residues/L) in potassium carbonate aqueous solution. The concentration of potassium carbonate is 0.050 mol/L.

2. Add pyrazole-1-carboxamidine (5 eq. per amine residue) to the above solution.

1H-Pyrazole-1-carboxamidine hydrochloride, 402516-10G, $54.70

3. Stire the reaction mixture for 20 h.

4. Purify by dialysis (M.W.C.O = 1,000 Da against distilled water. The conversion yield is 12% and 19% for PEI 25 kDa and PEI 2 kDa, respectively. Considering that the primary amine groups are 25% of total amine groups in PEI, the primary amine-guanidine conversion yields are 48% (25kDa) and 76% (2 kDa).

5. lyophilize

The transfection efficiency of the unmodified PEI 2 kDa is only one tenth of that of PEI 25 kDa at their optimal conditions. However, after the guanidylation, PEGu 2 kDa showed 20 times higher efficiency than PEI 2 kDa and two times higher than PEI 25 kDa. Moreover, the transfection efficiency of PEGu 2 kDa was eight times higher than that of PEI 25 kDa in presence of 10% fetal bovine serum condition, which was more similar to the in vivo environment.

IC 50 value of PEGu 2 kDa is more than five times of that of PEI 25 kDa. Considering that the transfection efficiency of the PEGu 2 kDa was two times higher than PEI 25 kDa, the guanidylation of low-molecular weight PEI could be a good way to obtain an efficient and safe gene delivery carrier.

In summary, the trasnfection efficiency of PEI 2 kDa can be improved 20 times by guanidylation as well as maintain its low cytotoxicity.