Unity plus 400 spectrometer

The kinetics of chitosan in solution was determined by ¹H NMR relaxation [longitudinal (T₁) and transverse (T₂) relaxation times and NOE] as the degree of acetylation (DA), and degree of polymerization, temperature, concentration and ionic strength. This analysis indicates that chitosan is a semi-rigid polymer with higher flexibility at higher DA, consistent with reduced electrostatic repulsion between protonated amino groups [32 (link)]. The ¹H NMR method was used to determine the structural characterization and degree of deacetylation (DD) of chitosan. A brief description of the experimental steps is as follows: after dissolving the chitosan in the solvent (HOD) containing 1% acetic acid-d₄ (10 mg/mL) at 70 °C, 1 mL of the chitosan solution was transferred into a 5 mm NMR tube. ¹H NMR spectra were recorded on a Varian-Unity-Plus-400 spectrometer operating at 400 MHz. Chemical shifts (δ) are reported in ppm and referenced to the residual solvent signal of D₂O at δ_H 4.80.
The integrals of the characteristic signals were used to calculate the degree of deacetylation (DD). The DD value is calculated according to the following formula: $$\mathrm{DD}\%=\left\{1-\left[{\mathrm{I}}_{\mathrm{CH}3}/\left(3\times {\mathrm{I}}_{\mathrm{H}1-\mathrm{GlnNAc}}\right)\right]\right\}\times 100$$
where the I_H1-GlnNAc is the integral for H1 (GlcNAc) and I_CH3 is the integral for—CH3 signal.

The solvents were purified according to the standard procedures. The ¹H, ¹³C spectra were recorded on a Varian Unityplus-400 spectrometer (400 and 125 MHz, respectively) in a DMSO-d₆ solution. Chemical shifts are reported in ppm downfield from TMS as internal standards. Mass spectra were recorded on an LC-MS instrument with chemical ionization (CI). LC-MS data were acquired on an Agilent 1200 HPLC system equipped with DAD/ELSD/LSMS-6120 diode matrix and mass-selective detector. Melting points were determined using a Fischer Johns instrument. Elemental analysis was performed at an analytical laboratory of the Institute of Bioorganic Chemistry and Petrochemistry, National Academy of Sciences of Ukraine.