Biodegradable crosslinked polyethylenimine and its uses
A polyethyleneimine, biodegradable technology, applied in the biological field, can solve problems such as cell membrane damage, and achieve the effect of low cytotoxicity and reduced cytotoxicity
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Embodiment 1
[0044] Synthesis and properties of embodiment 1 cross-linked PEI
[0045] 1 gram of PEI (Mw: 2000Daltons) was dissolved in 3ml of freshly distilled dichloromethane, and then an appropriate proportion of diglycidyl adipate and hexanediol diacrylate was added. After a proper reaction time at 40°C, the solution gradually turn yellow. The solution was then transferred to a Spectra / Por Mwco 10,000 membrane and dialyzed against double distilled water for 4 days at 4°C. It was then lyophilized to remove water to obtain a light-colored solid (CLPEI). Store at -70°C. Various cross-linked PEI polymers (Table 1) can be obtained by using different cross-linking agents and PEI combinations. The reaction combination is shown in Formula 1, and the obtained polymer structure is shown in Formula 2. Product carries out HNMR (varian 300MHz, D 2 (0) analysis, it was found that there was no double bond hydrogen (5.5-6.0ppm CH in the raw material) in the product 2 =CH-) absorption peak, but th...
Embodiment 2
[0055] Embodiment 2 Lactobionic acid and folic acid modification of cross-linked PEI
[0056] The cross-linked PEI is connected with lactobionic acid by amide bond through EDC (1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide) activated amino group. 200 mg of the cross-linked PEI synthesized in Example 1 was dissolved in 6 ml of 10 mM TEMED / HCl buffer. Then 180 mg of EDC was added and stirred at 25°C for 24 hours. Then add an appropriate amount of lactobionic acid and stir for 72 hours at 25°C. Finally, the obtained product was dialyzed with double distilled water for 4 days, lyophilized to remove water, and galactosyl-modified cross-linked PEI (Gal-PEI) was obtained.
[0057] 10 mg of folic acid and an appropriate amount of DCC were dissolved in freshly distilled DMSO and reacted at low temperature for a period of time, and then 200 mg of the PEI synthesized in Example 1 was added. After reacting for 12 hours under stirring, dialyze with double distilled water for 4 days at ...
Embodiment 3
[0058] Example 3 Preparation and Characterization of Crosslinked PEI and pEGFP-C1 Complex
[0059] Preparation of complex
[0060] The biodegradable cross-linked PEI or PEI 25kDa synthesized in Example 1 were respectively dissolved in PBS (140mM NaCl, 2.7mM KCl, 10mM NaCl 2 HPO 4 , 1.8mM KH 2 PO 4 , pH 7.4), made into a 1mg / ml stock solution. ddH 2 O diluted to 1 mg / ml stock solution. According to a certain PEI / DNA (mass ratio), prepare a complex of cross-linked PEI or PEI 25kDa PEI and DNA, for example, prepare a complex of cross-linked PEI / DNA=2:1, the specific process is as follows: Take 1 μl of plasmid DNA solution (1mg / ml) was dissolved in 50μl PBS, mixed gently, then 2μl cross-linked PEI was dissolved in 50μl PBS, mixed gently, then the two were mixed, oscillated for 10 seconds, and stood at room temperature for 10-15 Minutes, ready for experiments characterizing its performance and cell transfection.
[0061] Electrophoretic retardation experiment
[0062] Prep...
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