*2.10. Statistical Analysis*

All results, expressed as the mean ± SD, were analyzed using a two-tailed Student's t-test or one-way analysis of variance (ANOVA). The acceptable level of significance was *p* < 0.05.

#### **3. Results and Discussion**

#### *3.1. Structural Study of PNIPAm-co-PAAm-Mela HG System*

The 1H NMR spectrum analysis confirmed the structure of the prepared PNIPAm-co-PAAm-Mela HG. The 1H NMR spectra of the hydrogel system in CDCl3 solvent are shown in Figure 1. Because of the solubility of hydrophilic PAAm and hydrophobic PNIPAm and Mela segments, all of the relevant proton signals for PNIPAm-co-PAAm-Mela HG block copolymer are presented in Figure 1a. The resonance peaks for methyl (-CH2) and -OCH2 groups were δ0.7 ppm, δ1.2 ppm and δ1.8 ppm, respectively. The significant resonance signal at δ3.2 ppm indicates that the copolymer included the PNIPAm polymer segment in the PNIPAm-co-PAAm copolymer HG. The resonance peaks of methyl protons in isopropyl units are indicated by the peak at δ3.6 ppm. Furthermore, after Mela groups cross-linking, the resonance signals at δ2.4 ppm and δ1.82 ppm indicated that the melamine groups were cross-linked via glutaraldehyde units Figure 1b.

**Figure 1.** Characterization of PNIPAm-co-PAAm copolymer samples. (**a**,**b**) 1H NMR spectrum; and (**c**) FT-IR analysis of PNIPAm-co-PAAm copolymer; and PNIPAm-co-PAAm-Mela HG samples, respectively. (**d**) SEM images of the (**i**) PNIPAm-co-PAAm copolymer; (**ii**) PNIPAm-co-PAAm-Mela HG; (**iii**) PNIPAm-co-PAAm-Mela/Cur HG samples, respectively. The red circles in figure (**d**) (**iii**) indicate the presence of Cur molecules in the hydrogel samples.

The structure of the PNIPAm-co-PAAm-Mela HG system was determined using FT-IR spectroscopy. The FTIR spectrum is shown in Figure 1c, with the characteristic vibration peak at 1542 cm−<sup>1</sup> corresponding to the imine group (-C=N-) vibration, the peak at 1453 cm−<sup>1</sup> indicating the stretching bands of C-N groups, and the sharp band at 1417 cm−<sup>1</sup> indicating the N-H groups of NIPAm units in the PNIPAm-co-PAAm copolymer sample. The stretching mode of C=O groups in PNIPAm and PAAm segments was attributed to the absorption peak at 1727 cm−1. Additionally, the intense stretching peak appeared at 2836 cm−<sup>1</sup> and 2912 cm−<sup>1</sup> of the PNIPAM-co-PAAM-Mela copolymer system's alkyl carbon C-C stretching modes.

SEM analysis was used to examine the surface morphology of the prepared copolymer samples. SEM analysis was carried out on dried powder samples of PNIPAm-co-PAAm copolymer, PNIPAm-co-PAAm-Mela HG, and Cur loaded PNIPAm-co-PAAm-Mela/Cur HG, respectively. The PNIPAm-co-PAAm copolymer exhibited flake-like particles with an average particle size of around 2–3 μm, as shown in Figure 1d(i). In contrast, the Mela-Glu precursor cross-linked PNIPAm-co-PAAm HG sample (Figure 1d(ii)) exhibited slight aggregation with interconnected particles with appropriate pores with an average diameter of about 1 μm, which could be attributed to the cross-linking of PNIPAm-co-PAAm polymer chains via Mela-Glu precursor. Furthermore, the Cur drug-loaded PNIPAm-co-PAAm-Mela/Cur HG sample exhibited a similar aggregated morphology with micropores (Figure 1d(iii)).

The composition of PNIPAm-co-PAAm-Mela HG was determined by XPS analysis. As seen in Figure 2, the XPS spectra with prominent signals for carbon (C1s), nitrogen (N1s), and oxygen (O1s) peaks suggested that the presence of N-isopropyl acrylamide and acrylamide monomer segments existed in the PNIPAm-co-PAAm-Mela HG system. The high-resolution C1s spectra of PNIPAm-co-PAAm-Mela HG were shown in Figure 2a, with the peak at 284.5 eV and −289.2 mV, representing the aliphatic C-C binding mode. The nitrogen peak at 403.3 eV indicates the C-N groups of N-isopropyl acrylamide and acrylamide monomers in the PNIPAm-co-PAAm-Mela HG. The N1s spectra in Figure 2b revealed two peaks at 398.4 eV and 399.25 eV, evidenced that the C-N and C=N bonds of the amide and imine groups of N-isopropyl acrylamide and acrylamide monomer, as well as the melamine units of PNIPAm-co-PAAm-Mela HG [29]. Furthermore, the O1s signal at 402.9 eV represents the C=O groups of N-isopropyl acrylamide and acrylamide monomer in the PNIPAm-co-PAAm-Mela HG (Figure 2c). This supports that the PNIPAm-co-PAAm-Mela copolymer material was successfully synthesized [30].
