Paragon 500

Fourier transform infrared (FTIR) was employed to analyze samples in the powdered state in order to identify functional groups and host–guest complexes formation. Spectra were obtained using a PerkinElmer spectrometer (Paragon 500, Milan, Italy), equipped with a ZnSe attenuated total reflectance (ATR) crystal accessory. Spectra were acquired in the 4000–650 cm⁻¹ range, at a resolution of 4 cm⁻¹ (16 scans).

The prepared samples were characterized using XRD, FETEM, FESEM, FTIR, XPS, and PL techniques. Phase-purity and crystallinity were examined by X-ray diffraction (XRD) (PanAnalytic X’Pert Pro, Malvern Instruments, Malvern, UK) with Cu-Kα radiation (λ = 0.15405 nm, at 45 kV and 40 mA) and an angle range from 30° to 80°. Morphological and elemental mapping were identified using field emission transmission electron microscopy (FETEM, JEM-2100F, JEOL, Inc., Tokyo, Japan) and field emission scanning electron microscopy (FESEM, JSM-7600F, JEOL, Inc. Tokyo, Japan). Energy-dispersive X-ray spectrometry (EDS) was employed to determine the elemental composition. The chemical states of elements were investigated by X-ray photoelectron spectroscopy (XPS) (PHI-5300 ESCA PerkinElmer, Boston, MA, USA). Photoluminescence (PL) spectra were measured by a fluorescent spectrometer (Hitachi F-4600). Fourier transform infrared spectroscopy (FTIR) (PerkinElmer Paragon 500, Woonsocket, RI, USA) was used to identify the functional group of the prepared samples.

A series of analytical methods were used to confirm the successful formation of CS-LO-PEG-HER NPs. The loading of LO and, functionalization of mAb (Anti-HER) were analyzed by FTIR (Fourier-transform infrared spectroscopy; PerkinElmer Paragon 500, Waltham, MA, USA) and NMR (Nuclear Magnetic Resonance; FT-NMR, Bruker, 600 MHz, MA, USA). For the FTIR analysis, the nanoparticle samples, including CS, PEG6000, LO, HER, CS-LO NPs, CS-LO-PEG NPs, and CS-LO-PEG-HER NPs, were prepared as KBr pellets using the standard sample preparation method, and the IR scanning was performed from the 400–4000 cm⁻¹. To determine the size and zeta potential of nanoparticles, DLS (Dynamic light scattering; zeta potential particle size analyzer Malvern, Eindhoven, The Netherland) and ELS (Electrophoretic light scattering) analysis were used. For the DLS/ELS analysis, 10 µg of each NPs was dissolved in 3 mL of DI water, sonicated for 2 min, and filtered using a 2 µm syringe filter. The filtered nanoparticle suspension was used for DLS analysis. Transmission electron microscopic (JEOL-JSM 1200EX, Tokyo, Japan) analysis was used to observe the morphology of nanoparticles. For TEM analysis, 10 µg of each NPs was dissolved in 1 mL of ethanol, sonicated for 2 min, and negative stained with 2% uranyl acetate then loaded into a copper grid for TEM observation [27 (link)].

The micromorphology and elemental content of S-AgNP NPs were analyzed by transmission electron microscopy (FE-TEM) and energy-dispersive X-ray spectroscopy (EDS) (JEOL-JSM, Akishima, Japan). The particle size distributions and potentials of the nanoparticles were determined by a zeta potential particle size analyzer (Malvern PANalytical, Worcestershire, UK). The crystallinity of the starch, S-AgNP NPs, and AgNP NPs were determined using X-ray powder diffraction (XRD, X’pert-pro MPD-PANalytical, Worcestershire, UK). The functional characteristics of the nanoparticles were detected by Fourier-transform infrared spectroscopy (FTIR, PerkinElmer Paragon 500, Waltham, MA, USA). For the XRD and FTIR analysis, the starch, S-AgNP NPs, and AgNP NPs were analyzed after sufficient drying and grinding into powder. The silver content of S-AgNP NPs was analyzed using ICP-MS (PerkinElmer (NextION 300D), Waltham, MA, USA); in brief, 10 mg of sample was dissolved in 150 μL of HNO₃ and 350 μL of HCl, and after 3 h of full acid digestion, 20 μL was diluted in 10 mL of 0.2% HNO₃ solution. The silver and iron contents of AgNP NPs were determined according to standard calibration curves for silver and iron (1, 2.5, 5, 10, 25, 50, 100 μg/L) [16 (link)].

The presence of the functional group in the polysaccharides (EPS1, EPS2, and IPS1-4) was determined by Fourier transform infrared (FT-IR) spectroscopy (PerkinElmer Paragon 500, Waltham, MA, USA) analysis using the KBr pellets of polysaccharides. Then, the linkage of the polysaccharides was determined by ¹H, ¹³C and 2D NMR analysis. A total of 20 mg of each polysaccharide (EPS1, EPS2, and IPS1-4) was dissolved in 0.5 mL of D₂O at ambient temperature. The NMR was recorded using an FT-NMR spectrometer (Bruker, 600 MHz, Fällanden, Switzerland).

The FTIR-ATR spectra of the obtained films were recorded using a Perkin-Elmer FTIR spectrometer (model Paragon 500 equipment) with the aim to assess the completeness of the photopolymerization process. The spectra (32 scans) were recorded in a 4000–400 cm⁻¹ wavelength range at 4 cm⁻¹ resolution.