Ir solution software

All samples were analyzed by a Shimadzu IRAffinity-1 FT IR Spectrophotometer (Shimadzu, Japan) with a DLATGS detector (Deuterated Triglycine Sulfate Doped with L-Alanine). Scan range was set at 4000−500 cm⁻¹ with 20 scans at 4 cm⁻¹ resolution. Spectra were obtained with an Attenuated Total Reflectance (ATR) accessory with a ZnSe crystal, at 20 °C and 50% relative humidity. The spectra were corrected for CO₂ and spectral noises using the IR-Solution software (version 1.50—Shimadzu).

Filtrate samples after incubation were diluted 1:10 in 0.05 M phosphate buffer at pH 7.5, then equilibrated at room temperature for 10 minutes and finally examined with an electron microscope (JEOL/JEM 1200 EX II, Japan), operated at 120 kV, atomic force microscope (Dimension 3100, Veeco, USA) and FT-IR spectrometer (IRPrestige-21, Shimadzu, Japan).
TEM. Samples for transmission electron microscopy were prepared according to the following sequence: incubation, filtration and dilution 1:1 with a fixative (1.5% glutaraldehyde in 0.1 M sodium cacodylate buffer) and stored at 4°C.
AFM. The solution of α-LaNTs was diluted 1000 times in the same buffer used for preparation of the nanotubes. A sample of 20 μL of this solution was incubated at room temperature for 10 minutes on a glass surface previously immersed for 5 minutes in a solution of 1% DETA (Diethylenetriamine, Sigma-Aldrich, USA). The surface was washed with buffer to remove excess material and allowed to dry at room temperature. AFM was used in tapping mode.
FT-IR. An attenuated total reflectance (ATR) sampling accessory in combination with the FT-IR spectrometer provided the best means for determining the composition of the α-LaNTs. Infrared spectra were obtained over the range 1100–1800 cm⁻¹ with a resolution of 1 cm^-1 and 256 scans. The absorbance values were determined by mean of the IRsolution software (Shimadzu, Japan).

An FTIR spectrometer (PerkinElmer, USA) equipped with a universal ATR Miracle accessory was used to analyze the leaves of plants obtained from inoculated seeds and the control. The leaf sample was placed on a ZnSe crystal plate (PerkinElmer) and FTIR spectra were recorded over the region of 4000–380 cm–1 at room temperature, with a resolution of 4 cm–1 and 64 scans. First, raw data (first derivates) were smoothed using IRSolution software (Shimadzu, Japan). The second derivative and vector-normalized IR spectra were used in all data analyses while the signals of amide and lipid bands were analyzed from the vector-normalized absorbance IR spectra. Peak intensity was determined using the Quant option in the IRSolution software. Baseline corrected spectra were normalized according to the following formula: (x – min(x))/max (x – min(x)), with x as the absorbance. Principal component analysis (PCA) was performed using MATLAB software. Each experiment was repeated three times.

To obtain tablets the C. cupressoides SPs (5 mg) were mixed thoroughly with dry potassium bromide (KBr). Then, infrared spectra of these tablets were obtained using Fourier Transform Infrared Spectroscopy (IRAffinity-1, Shimadzu Corp., Kyoto, Japan) equipped with IRsolution software (version 1.20, Shimadzu Corp., Kyoto, Japan). The analysis frequency range was 4000 to 400 cm⁻¹. Three independent analyses were performed.

1 g sodium hyaluronate (2.5 mmol) was dissolved in water (100 mL) at RT. A sodium periodate solution (0.982 mmol) was added dropwise to the dissolved hyaluronate, after which 500 µL ethylene glycol was added while stirring for 1 h. The reaction mixture (i.e., oxidized hyaluronic acid, OHA) was purified by dialysis, lyophilized, and stored at 4 °C. OHA was dissolved in distilled water, and the PEI solution was added. The final mass ratio was 1:1. The reaction mixture (i.e., OHA‐PEI) was lyophilized and stored at 4°C until use. The synthesis of OHA‐PEI was determined using attenuated total reflection‐Fourier transform infrared spectroscopy (ATR‐FTIR, Shimadzu 8400S, Shimadzu 605 Corp., Kyoto, Japan). Spectra were taken from 4000 to 400 cm⁻¹, using 32 scans at a resolution of 4 cm⁻¹. Spectral analysis was performed using IR solution software (Shimadzu). The surface topography of OHA‐PEI was examined by SEM.