Mechanically Robust Gels Formed from Hydrophobized Cellulose Nanocrystals.

Cellulose nanocrystals (CNCs) that bind to each other through associative hydrophobic interactions have been synthesized by modifying sulfated CNCs (sCNCs) with hydrophobic moieties. These octyl-CNCs form gels at significantly lower concentrations than parent sCNCs, producing extremely strong hydrogels. Unlike sCNCs, these octyl-CNCs do not form ordered liquid crystalline phases indicating a random association into a robust network driven by hydrophobic interactions. Furthermore, involvement of the octyl-CNCs into multicomponent supramolecular assembly was demonstrated in combination with starch. AFM studies confirm

2 favorable interactions between starch and octyl-CNCs, which is thought to be the source of the dramatic increase in gel strength.
The formation of strong gels from cellulose nanofibrils are well-documented in the literature, and are potentially useful as rheological modifiers in food, paints and in homecare and beauty products. [1][2][3] Their stability in each of these applications environments is predicated on their mechanical integrity. Cellulose nanocrystals (CNCs), a form of highly crystalline rod-like cellulose 4 are also reported to form gels. [5][6][7] It is generally agreed that rheological properties of CNC suspensions are defined by their ability to form biphasic systems of isotropic and chiral nematic phases. 5,6,[8][9][10] At low concentrations, CNC suspensions are practically isotropic and behave almost as Newtonian fluids. With an increase in the CNC concentration these suspensions exhibit the properties of viscoelastic fluids due to the formation and growing fraction of the liquid crystal phase. Further increases in concentration lead to the formation of randomly entangled gels with dominating elastic properties. Typically, these gels are relatively weaker than those formed of fibrils due to relatively fewer inter-fiber crossings. Critical concentrations at the transition between these states of CNC suspensions depend on their aspect ratio and surface charge, and the ionic strength of the aqueous media. CNCs have been explored as reinforcing additives for hydrogels both as a component of supramolecular hydrogels [11][12][13] and a reactive component incorporated into polymer networks via covalent binding [14][15][16] . Herein we report the formation of extremely strong gels based on CNCs using chemical modification of their surface to introduce hydrophobic alkyl chains on their surface (octyl-CNCs). These strong gels form at much lower concentrations than typical CNC materials, which holds potential for their use for a wide range of applications where stability is required. Moreover, we demonstrate that octyl-CNCs induce the formation of strong supramolecular hydrogels in combination with starch. The work represents a new form of 3 nanocellulose based colloidal or hybrid supramolecular and colloidal hydrogel. Preservation of the anionic charges of parent CNCs enables high water intake while the addition of hydrophobic groups enhances network formation via fibrils self-association or supramolecular binding of water soluble polymers.
Modified CNCs were synthesized from materials produced by hydrolysis with sulfuric acid.
From here we denote the initial CNCs as sCNCs to indicate that their surface is functionalized with sulfate half-ester groups. Octyl-CNCs were synthesized in two steps: periodate oxidation of sCNCs followed by the reductive amination reaction between aldehyde groups of the oxidized sCNCs and octylamine. The modification of the CNCs with octyl chains has been confirmed by FTIR and NMR (Supporting information, Figures S2, S3). From the analysis of 1 H-13 C CP-MAS NMR spectra degree of surface functionalization was found to be around 4% (Supporting information, Table S1). Both sCNCs and octyl-CNCs were used in sodium form as never dried materials.
The viscoelastic properties of octyl-CNCs and sCNCs over a wide range of concentration are compared in Figure 1. In contrast to sCNCs, G' is found to be higher than G'' (tan  < 1) for octyl-CNCs over the whole range of concentrations, showing the dominance of elastic properties. At concentrations above 2.0 wt.% octyl-CNCs formed strong self-supporting gels with a value of G' larger than G''; by a factor of 3 or more. Suspensions of octyl-CNCs however did not form invertible gels at concentrations below 2.0 wt.% (Supporting information, Figure S4) and there was less than a decade difference in G' and G'' for a 1.2 wt.% suspension (tan  > 0.18). These values are characteristic for "weak gels" or "structured fluids" which are usually formed by a tenuous association of mesoscopic domains. Naturally G' increased with an increase in concentration for both types of CNCs. However, for suspensions of the same concentrations G' is significantly higher for octyl-CNCs. For example, at 5 wt.% G' for the octyl-CNCs gel is almost five orders of magnitude higher than for sCNCs; c.f. 4300 and 0.05 Pa, respectively. Moreover, in contrast to octyl-CNCs, tan  is strongly dependent on the concentration of the sCNCs suspension. The storage modulus also increased significantly for the sCNCs suspension, and tan  only fell below 1.0 when concentrations were higher than 5 wt.%. Thus, the elastic properties of sCNC suspensions only became significant when concentrations exceeded 5 wt.%. inclusion complexes with hydrophobic and amphiphilic ligands such as fatty acids, lactones, aldehydes, etc. 20 In such complexes a hydrophobic cavity of a single amylose helix accommodates a hydrophobic guest or hydrophobic part of an amphiphilic ligand. Inclusion complexes of large ligands such as hydrophobic synthetic polymers 21,22 or Single Walled Carbon Nanotubes (SWCNTs) 23 have been also assembled with amylose. Starch properties vary depending on the source. However, starches of any source are known to form gels only at relatively high concentrations (above 6 wt.%). 24,25 We attempted to achieve gelation in starch/CNC systems, when individual concentrations are lower than those required for the formation of invertible gels. Figure 3 compares the rheological properties of gels prepared with sCNCs and octyl-CNCs. Both gels showed a weak dependence on frequency, responding as solid-  Normalization of adhesive interaction forces to the force of tip interaction with the background mica allows a simple comparison of results obtained with different tips and substrates as adhesive interaction forces can prove complex in ambient environments and are typically dominated by the formation of a capillary bridge. 28 In such a way, each sample was characterized using a fresh cantilever which ensures sharp tips (maintaining a tip radius of ~2 nm for different samples) for maximum image resolution and tip surfaces conditioned in the same environment for comparative force measurements.
As can be seen from Figure 4b, bare sCNC and octyl-CNC significantly differ in their affinity towards the AFM probe; relative adhesive responses of 24  5% and 69  8% were respectively obtained. AFM probe adhesion to starch (42  7%) is lower than adhesion to bare octyl-CNCs but higher than to bare sCNCs. Mixing starch with octyl-CNCs leads to a significant drop in the AFM probe adhesion to octyl-CNCs (38  6%). In fact, adhesion between the probe and octyl-CNCs in a starch/octyl-CNC mix is the same as the probe/starch adhesion within their respective error bounds (cf. 38  6% and 42  7%). This further verifies that octyl-CNCs are coated with starch.
However, the presence of starch does not influence the interaction between the sCNCs and the AFM probe (relative adhesion 24  9% compared to 24  5% for bare sCNC). This confirms the lack of affinity between starch and sCNCs and sCNCs remain uncoated when mixed with starch.
Thus, AFM allows clear identification of different types of CNCs even when they are combined with starch.
To further asses the effect of supramolecular association of different types of CNCs on the properties of starch/CNC gels, a series of systems were produced with a mix of both hydrophobized and non-hydrophobized CNCs. It was found that sCNCs weakened interactions, thereby decreasing the elastic properties of starch/CNC gels (Supporting Information, Figure S6a). Despite the increase in the total CNC content, starch/CNC gels had a lower yield stress and viscosity when an additional sCNC content (25 and 50wt.% with respect to octyl-CNC) was added to the gels. In fact, gels with a higher CNC content (3 wt.%) were weaker than gels containing 2 wt.% octyl-CNCs or 2 wt.% octyl-CNCs with 0.5 wt.% sCNCs. Thus, different combinations of sCNCs and octyl-CNCs allow the tuning of rheological properties of gels according to a desired application.

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Poor starch/sCNC interactions and lack of sCNC integration into a multicomponent supramolecular network was confirmed by the particle analysis of the topographic images of starch combined with both sCNCs and octyl-CNCs (Figure 4c). An increase in content of octyl-CNC content causes drop in number of thin (less than 3 nm) particles, which are loose starch particles in starch/CNC mixtures. Thus, a growing fraction of starch adheres to CNC surfaces when the octyl-CNCs content increases. Simultaneously, thicker particles become dominant with an increase in octyl-CNC content. These particles represent octyl-CNCs coated with starch reflecting re-distribution of the loose starch to octyl-CNC-bound states.
In summary, we have produced gels using hydrophobically modified CNCs. These gels show very high storage moduli compared to non-functionalized sCNCs. Associative hydrophobic CNCs form gels at lower concentrations. Furthermore, it is shown that surface functionalization increases the affinity of CNCs to starch, enabling the formation of strong gels at relatively low concentrations. Hydrophobized CNCs are expected to advance rheological modifying and gel forming formulations for food, cosmetic and pharmaceutical products. Properties of materials based on hydrophobic CNCs and thereby applicable to such materials, and can be further diversified in combination with other hydrocolloids such as starch and other polysaccharides.
Hydrophobically derivatized CNCs can enable further progress in polymer nanocomposites due to potentially enhanced compatibility of modified CNCs with hydrophobic polymer matrices.
Specifically, the demonstrated high affinity between octyl-CNC and starch can pave the way to composites combined with thermoplastic starch.

ASSOCIATED CONTENT
The Supporting Information is available free of charge on the ACS Publications website at DOI: Synthesis, experimental details, FTIR, NMR results, optical images of gel formation, determination of CNC sizes from AFM, rheology results for starch/CNC gels (PDF). All data from this paper can be found on the University of Bristol repository at https://data.bris.ac.uk/data/.

Author Contributions
The manuscript was written through contributions of all authors led by R.Nigmatullin and S.J.
Eichhorn. All authors have given approval to the final version of the manuscript.

Funding Sources
The Engineering and Physical Sciences Research Council (EPSRC) is acknowledged for provision of financial support (EP/N03340X/2, EP/N033337/1). PeakForce atomic force microscopy was carried out in the Chemical Imaging Facility, University of Bristol with equipment funded by EPSRC under Grant "Atoms to Applications" (EP/K035746/1).