Synthesis of PLA-PEG-FA and PLA-PEG-Glu copolymers
The chemical structure of the PLA-PEG-FA and PLA-PEG-Glu copolymers were examined with proton nuclear magnetic resonance spectroscopy (1H NMR). Fig. 1 showed the 1H-NMR spectrum of PEI-PLA-PEG-FA copolymer, where the presence of CH2 protons of PEG in PEI-PLA-PEG-FA copolymer was observed around 3.6 ppm (-CH2CH2O-). Moreover, the appearance of signals at 5.1 and 1.6 ppm are attributed to the methine (CH) protons and methyl protons – (CH3) of PLA, respectively (Fig. 1 A) (Heald et al., 2002; Sim et al., 2017).
The appearance of chemical shifts at 6.5-8.6 ppm (aromatic protons associated with FA) and 7.9 ppm (s, weak, 1H, triazoles) provided evidence for successfully obtaining PEI-PLA-PEG-FA copolymer (S.-J. Yang et al., 2010)(Fig. 1 A).
Also in the Fig. 1 B The multiple sharp peaks at 3.6-3.8 ppm and weak peak at 5.4 ppm which were assigned to methylene protons in PEG and aromatic proton in glucose respectively, appeared on the spectrum after the reaction of PLA-PEI with PEG-Glu (Fig. 1 B)(Sadeghi et al., 2015).
The structures of PLA-PEG-FA and PLA-PEG-Glu were ascertained by the FTIR spectrum (Fig. 2). The sharp peak appearing at 1767 cm-1 was assigned to the carbonyl (C=O) group in the PLA-PEG copolymer (Xiong et al., 2011).
The peaks that appeared in the regions of 1193 cm-1 and 1460 cm-1 are related to stretching C-O and the bending of -CH2- groups in PLA-PEG copolymer, respectively (Amani, Kabiri, Shafiee, & Hamidi, 2019; Danafar, Rostamizadeh, Davaran, & Hamidi, 2017).
Moreover, the peaks at 1640 and 1095 cm-1 corresponds to the C=O and C-O-C stretching of COOH in the FTIR spectrum of PEG, PLA- PEG-FA and PLA-PEG-Glu suggested that PEG was grafted to the nanoparticles (Ahmed, Arfat, Castro-Aguirre, & Auras, 2016; Rafat et al., 2010).
The FTIR spectra of PLA-PEG-FA copolymer showed that there is also a broad peak in the region between 1580 and 1648 cm−1 belonging to the amine group of FA. Moreover, the peak observed around 2108 and 2116 cm-1 were derived from azido group of FA and Glu respectively. The peak at 3580 cm-1 corresponds to the O-H stretching of OH in the FTIR spectrum of glucose suggested that glucose was grafted to the PLA-PEG.
The FT-IR spectra of FeCo, FeCo-PEI, FeCo-PEI-PLA-PEG-FA, and FeCo-PEI-PLA-PEG-Glu are illustrated in Fig. 2.
After conjugation of PEI with FeCo nanoparticles, the PEI peaks from 1050 cm-1 to 1250 cm-1 were attributed to the amide and amide amino groups , which indicated that the PEI was attached on the surface of FeCo nanoparticles (Fig. 2) (Gultekinoglu et al., 2015).
As also evident in Fig. 2, almost all peaks related to PLA-PEG, FeCo-PEI, glucose and folic acid were observed in the NPsA and NPsB, therefor it is indicated the successful synthesis of these nanoparticles.
Thermogravimetric analysis (TGA) of NPsA and NPsB was performed to find out different chemical compounds in the samples. The TGA curves of PEG, PEI, PLA, PLA-PEG, PLA-PEG-FA and CBP-FA copolymers, are shown in Fig. 3.
TGA analysis of the pure polymers (PEI, PLA and PEG) shows single step degradation while the spectrum of PLA-PEG copolymer exhibits three steps degradation related to the presence of three components in the PLA-PEG copolymer. The initial mass loss in the range of 175 °C to 305 °C, may be due to the loss of adsorbed water molecules from the copolymer (Jadhav et al., 2013).
The second stage of the PLA-PEG degradation occurs between 320 °C and 380 °C, which is related to the thermal degradation reaction of the PLA polymer (Shih & Huang, 2011). The third stage of thermal degradation occurs at temperatures above 380 °C, is associated with degradation of PEG polymer (Kwon & Kim, 2006).
The thermogram of the –FA and –Glu shows several mass losses during the temperature rise from 200 to which proposes that the glucose has higher temperature resistance, compared to other pure PEI, , and folic acid and therefore, temperature resistance of PLA-PEG copolymer increased after binding to glucose.
As it is seen in Fig. 3 the FeCo-PEI nanoparticles has a much better thermal stability than that of PLA-PEG-FA and PLA-PEG-Glu. Therefore, improvement in temperature resistance of PLA-PEG-FA and PLA-PEG-Glu was observed after modification with FeCo nanoparticles, which indicated the successful conjugation of PLA-PEG-FA and PLA-PEG-Glu with FeCo.