*2.3. Raw Material and Cellulosic Pulp Characterization*

The olive tree pruning biomass and the cellulosic pulp obtained were characterized in terms of the chemical composition of the lignocellulose matrix. Both were characterized according to their content of ethanol extractables, hot water extractables, ashes, lignin, hemicelluloses and α-cellulose according to the TAPPI standards T-204, T-435, T-211, T-222, T-9m-54 and T-203 cm-09, respectively. The determination of each component of the chemical characterization was performed in triplicate and the means and standard deviations were calculated.

### *2.4. Cellulose Nanofiber Production*

To obtain cellulose nanofibers (CNFs), two independent pretreatments were used, mechanical beating and TEMPO-mediated oxidation, both followed by a high-pressure homogenization treatment.

The mechanical pretreatment consisted of a mechanical refining (PFI beater) according to the ISO 5264-2:2002 standard, during 40,000 revolutions, to reach a Schopper–Riegler Degree (ºSR) of 90 [26]. This pretreatment allows the fibrillation of the cellulose fibers by shear forces to facilitate nanofibrillation in the subsequent treatment. The TEMPO-mediated oxidation was carried out following the methodology described by Saito et al. [29]. The reaction was carried out at pH 10 and started with the addition of a specific amount of NaClO solution in order to obtain an oxidative power of 5 mmol per g of fiber. Once the addition of NaClO was complete, the pH was maintained by adding a 0.5 M NaOH solution. The reaction was finished when the pH remained stable.

A 1% pretreated fiber suspension was subjected to a nanofibrillation process in a high-pressure homogenizer (PandaPlus 2000, GEA Niro, Düsseldorf, Germany) in order to isolate the nanofibers that form the cellulose fibers. To avoid the occlusion of the homogenizer, gradual fibrillation was performed in the following sequence: 4 passes at 300 bars, 3 passes at 600 bars and 3 passes at 900 bars. This treatment has been demonstrated as an effective way of obtaining cellulose nanofibers from different raw materials and pretreatments [34].

By means of the mechanical and TEMPO-mediated oxidation pretreatments, CNFs were obtained, although in the case of unbleached pulp, residual lignin content remained in the final product (lignocellulose nanofibers; LCNF).
