Nanoscope 9

The samples were prepared by incubation on mica as for the imaging experiments above. Images and mechanical properties were obtained simultaneously using “peak force QNM” on a Multimode AFM (Bruker), under ambient conditions. Tap525A probes, with a nominal spring constant (k) of 200 N/m or RTESP-300-30 (Bruker) with a calibrated spring constant of 47.1 N/m probes were used. The effective tip radius was calibrated on a sample of HOPG with a modulus of 18 GPa or on PolyStyrene (PS) with a modulus of 2.7 GPa. Deformation depths were kept to about 1 nm. The Bruker software (Nanoscope 9.2) was used to compute the elastic modulus maps using the DMT model to fit the force vs. deformation curves.

AFM analysis was performed on a Bruker Dimension Icon using NanoScope 9.1 (Karlsruhe, Germany) for measurements and NanoScope Analysis 1.5 for image processing. We measured in ScanAsyst air mode using a ScanAsyst air tip with a spring constant of ~0.4 N/m and a resonant frequency of 70 Hz.3.2.4. NMR and ESI MS NMR spectra were recorded on a Bruker Avance 300 MHz Spektrometer (Ettlingen, Germany). Mass spectra were recorded on FlexarTM SQ 300 MS Detector (PerkinElmer, Rodgau, Germany).

For the viscosity
measurements, a DV2TLV viscometer (Brookfield) was used along with
Rheocalc software. Each CNF sample (0.4% CNF suspension) was evaluated
in three parallels, and the viscosity of each parallel was measured
from 0.1 to 100 rpm. For the nanostructures, 0.01% nanocellulose suspensions
of each sample (25 μL) was added to clean the mica surface,
left to dry in air, and then imaged with a Dimension ICON atomic force
microscope (AFM) using NanoScope 9.4 Software (Bruker). Surface roughness
(R_a) was obtained from images (10 μm
× 10 μm) using NanoScope Analysis software (version 1.9).
For the quantification of residual fiber content, a Fiber Tester
912 Plus (ABB AB/Lorentzen Wettre) was used. CNF suspensions were
pumped through a flow cell where the fibers were photographed with
a resolution of 4 μm. The images were further used to analyze
the mean length and width of the fibers. To evaluate the microstructure
of all cellulosic materials, tissue culture plates (24-well clear
flat bottom polystyrene; NUNC, Denmark) were covered with cellulosic
suspensions and stored at 37 °C for 24 h to dry. Films were analyzed
using an optical microscope (Nikon Eclipse 80i, Tokyo, Japan) after
staining with crystal violet.

A 5 µL volume of the freshly prepared NFPh emulsion, prepared as described in Section 4.1, was dispensed onto freshly cleaved mica. Mica surface (0.15 mm thick, sized 15 × 15 mm, TipsNano, Zelenograd, Russia) was used as a substrate for non-covalent adsorption.
The emulsion droplet was incubated on the mica substrate surface for 10 min. Then, the substrate was washed with 1 mL of deionized water, which was obtained using a Simplicity UV system (Millipore, Molsheim, France). The washed substrate was dried in air and subjected to AFM scanning.
The AFM images of nanosized particles were obtained with a Dimension atomic force microscope equipped with an Icon^™ scanner (Bruker, Billerica, MA, USA). The instrument is a part of the Avogadro unique research facility (http://avo.ibmc.msk.ru/ (accessed on 1 October 2023)). Scanning was carried out in the tapping mode. A short cantilever holder was used; the measurements were conducted in air. The images were recorded using the NanoScope 9.4 software (Bruker, Billerica, MA, USA). AFM images were processed using the standard NanoScope Analysis 2.0 software (Bruker, Billerica, MA, USA), Gwyddion 2.62, and Femtoscan Online software 4.8 (LLC Scientific and Production Enterprise “Center for Advanced Technologies”, Moscow, Russia; www.nanoscopy.net/en/Femtoscan-V.shtm (accessed on 6 May 2021)).