Synthesis, Characterization And Biomedical Applications Of Branched Polyethylenimine-and Dendrimer-Based Functionalized Nanomaterials

Because of their unique properties of sound, optics, electronics, magnetism, heat and mechanics, nanomaterials have been widely applied in biomedical fields and become the hot spots of research. In recent years, the nanomaterials synthesis using polymers has proven to offer them unique chemical and physical properties, which significantly expanded their applications. This paper will be focused on the synthesis, characterization and biomedical applications of polyethylenimine (PEI) and amine-terminated generation five poly(amidoamine) (PAMAM) dendrimer for the functionalization or synthesis of gold nanoparitcles or functionalization of carbon nanotubes. All the relative studies are outline as follows:(1) The amine groups of PEI were modified with acetyl, carboxyl, hydroxyl groups, and PEG chains and these modified PEI derivatives were characterized using 1H NMR, FTIR, and zeta-potential measurements. The results show the PEI surface charge can be transformed to be positive, negative, and neutral, forming PEI with desired surface functional groups. In addition, acetylation of PEI is a controllable, stoichiometric reaction and partial acetylation of PEI may enable multiple modification of PEI with different funetionalities. Importantly, the formed PEI derivatives (PEI-Ac, PEI-PEG-Ac, PEI-SAH, and PEI-Gly) do not display apparent cytotoxicity at a concentration up to 200μg/mL, while the pristine PEI is cytotoxic at a concentration as low as 10μg/mL. Our study indicates that through relatively simple modification strategies to neutralize the surface amines, PEI can be functionalized to have improved biocompatibility.(2) We demonstrated the green synthesis of PEI-stabilized gold nanoparticles (PEI-AuNPs) and the use of the particles for the highly selective and sensitive colorimetric detection of heparin for the first time. Due to the electrostatic interaction between the negatively-charged heparin and the positively-charged PEI-AuNPs, PEI-AuNPs undergo aggregation and a color change from red to blue in solution in the presence of heparin, thus providing a simple and visual approach for colorimetric detection of heparin. The prepared PEI-AuNPs has been successfully applied to the colorimetric detection of heparin with a concentration rang of 3-11μg/mL and with high sensitivity (with a detection limit of 1.5μg/mL). The results showed that heparin can be detected quickly and accurately in lower ppm level with excellent discrimination against other biopolymers. This method can be potentially applied in clinic medicine.(3) The multifunctional dendrimer-based gold nanoparticles were synthesized, characterized and applied as a dual modality contrast agent for computed tomography and magnetic resonance (CT/MR) molecular imaging applications. Amine-terminated generation five PAMAM dendrimers (G5-NH2) were multifunctionalized through grafting gadolinium chelate (DOTA-NHS) and polyethylene glycol monomethyl ether with one end of carboxyl group (mPEG-COOH) onto their surface. Using functional dendrimers as templates, gold nanoparticles were prepared with sodium borohydride reduction chemistry, and then Gd(III) was chelated into DOTA molecules on the dendrimer surface. Subsequently, the remaining amine groups of dendrimer were acetylated to neutralize the positive charges and finally got the multifunctional dendrimers containing both CT and MR imaging elements. The formed{(Au0)250-G5-DOTA(GdⅢ)10-PEG20-Ac} nanoparticles (G5-Gd/Au DENPs) were characterized using'H NMR, transmission electron microscopy (TEM), UV-Vis absorption spectrometry, MR T1 relaxometry, and X-ray attenuation measurements. In vitro and in vivo CT and MR imaging and the biodistribution were also investigated. The results indicated that the numbers of DOTA, PEG, Gd(Ⅲ) and Au atom onto each dendrimer were 8.8, 18.0,27 and 250, respectively. We also showed that the formed G5-Gd/Au DENPs are colloidally stable and biocompatible. The synthesized G5-Gd/Au DENPs displayed the reasonable r1 relaxivity and enhanced X-ray attenuation property in vitro, while in vivo CT and T1-weighted MR images show that the G5-Gd/Au DENPs can used to for dual modality blood pool imaging of rats or mice. Furthermore, the in vivo biodistribution analysis reveals that the nanoparticles have the prolonged blood circulation time and have a metabolism process in organs (eg. liver and spleen) with the increase of time. Therefore, the synthesized multifunctional dendrimer-based composite particles may be used as a bimodal imaging contrast agent for biomedical imaging.(4) Through the optimization of pH value, the concentration of doxorubicin (DOX) and multiwalled carbon nanotubes (MWCNTs), the mass ratio of DOX and MWCNTs, we were able to get the high drug loadings (1d) and encapsulation efficiency (ee) of DOX within MWCNTs. The optimized conditions were pH 9.5, concentration of 1 mg/mL for both DOX and MWCNTs, and mass ratio of 1:1. In addition, G5-NH2 were sequentially modified with fluorescein isothiocyanate (FITC) and folic acid (FA) via covalent conjugation, followed by covalent grafting on the surface of MWCNTs. Finally an acetylation reaction was applied to neutralize the remaining surface amines of dendrimers. The synthesized multifunctional MWCNTs (MWCNTs-G5-FI-FA-Ac and MWCNTs-G5-FI-Ac) were then used to load DOX for targeted and pH-sensitive delivery of the drugs to cancer cells expressing high-level folic acid receptors (FAR). We showed that the formed drug delivery system has the high Id and ee both with 98%. In vitro DOX release tests show that the formed DOX/MWCNT-G5-Fl-FA-Ac and DOX/MWCNT-G5-FI-Ac display a pH-responsive release property. DOX can be fast released from this nanocomposite materials under acidic environment (pH= 5.4 acetic acid salt buffer, which refers to the tumor tissue environment), while it is released slowly under physiology environment (pH=7.4 PBS buffer). In vitro cytotoxicity experiments indicated that the DOX loaded onto the multifunctional MWCNTs were able to effectively kill KB cells. Also, the cell morphology, MTT assay, and laser confocal microscope studies showed that the formed DOX/MWCNT-G5-FI-FA-Ac were able to target to KB cells overexpressing high-level folic acid receptors and can selectively and effectively kill the specific KB cells.