Unraveling the Supramolecular Structure and Nanoscale Dislocations of Bacterial Cellulose Ribbons Using Correlative Super-Resolution Light and Electron Microscopy

Abstract

Cellulose is a structural linear polysaccharide that is naturally produced by plants and bacteria, making it the most abundant biopolymer on Earth. The hierarchical structure of cellulose from the nano- to microscale is intimately linked to its biosynthesis and the ability to process this sustainable resource for materials applications. Despite this, the morphology of bacterial cellulose microfibrils and their assembly into higher order structures, as well as the structural origins of the alternating crystalline and disordered supramolecular structure of cellulose, have remained elusive. In this work, we employed high-resolution transmission electron and atomic force microscopies to study the morphology of bacterial cellulose ribbons at different levels of its structural hierarchy and provide direct visualization of nanometer-wide microfibrils. The non-persistent twisting of cellulose ribbons was characterized in detail, and we found that twists are associated with nanostructural defects at the bundle and microfibril levels. To investigate the structural origins of the persistent disordered regions that are present along cellulose ribbons, we employed a correlative super-resolution light and electron microscopy workflow and observed that the disordered regions that can be seen in super-resolution fluorescence microscopy largely correlated with the ribbon twisting observed in electron microscopy. Unraveling the hierarchical assembly of bacterial cellulose and the ultrastructural basis of its disordered regions provides insights into its biosynthesis and susceptibility to hydrolysis. These findings are important to understand the cell-directed assembly of cellulose, develop new cellulose-based nanomaterials, and develop more efficient biomass conversion strategies.

Exploring Microemulsion Systems for the Incorporation of Glucocorticoids into Bacterial Cellulose: A Novel Approach for Anti-Inflammatory Wound Dressings

“…To the best of our knowledge, Figure 9 shows the first FF-TEM image of the hierarchical structure of ~ 80 × 4 nm BC ribbon-like microfibrils ( K. xylinus ) and their assembly by smaller protofibrils, allowing a very interesting insight into the BC ribbon nanostructure. Important to mention is the recent work of Babi et al [ 96 ], who published TEM images of the BC supramolecular structure as well. Nevertheless, the FF-TEM technique in combination with plunge freezing is likely to better preserve the native microstructure, as well as show the structure of never-dried BC, in contrast to the drying involved in other imaging techniques, such as atomic force microscopy (AFM) and TEM.…”

Structural Color from Cellulose Nanocrystals or Chitin Nanocrystals: Self-Assembly, Optics, and Applications

“…Acid hydrolysis is thought to attack the glycosidic bonds at the periodically arranged disordered regions of the fibers (section ), leading to a rapid reduction in the molecular weight followed by a leveling off of the degree of polymerization (LODP). − This breakage, along with cellulose affinity for the solvent, induces the separation of laterally assembled fiber bundles. In accordance with the lower critical solution temperature (LCST) behavior of cellulose in sulfuric acid, , the use of low hydrolysis temperature (<50 °C), favors the presence of residual oligosaccharide chains (7–20 DP) solubilized from the crystallite surfaces .…”

“…Native cellulose, while having defects and dislocations in the crystal structure, contains no extensive amorphous regions . However, there is evidence of periodic disorder along the microfibril chain axis of cellulosic fibers, as observed by small-angle neutron scattering and, more recently, super-resolution microscopy. , While this would explain the leveling-off degree of polymerization (LODP) observed for the hydrolysis of cellulose microfibrils, the origin of these periodic disorders along the microfibrils is a matter of debate …”

“…38 However, there is evidence of periodic disorder along the microfibril chain axis of cellulosic fibers, as observed by small-angle neutron scattering 55 and, more recently, super-resolution microscopy. 56,57 While this would explain the leveling-off degree of polymerization (LODP) observed for the hydrolysis of cellulose microfibrils, the origin of these periodic disorders along the microfibrils is a matter of debate. 38 There are several ways to laterally associate cellulose chains, which result in different crystal structures, known as allomorphs (Figure 2).…”

Grafting polyanhydride polymers to cellulose nanofibers

“…This form of CNF is a broad mixture (Arteaga-Pérez et al 2017) (Kelly et al 2021) of spaghetti-like (Youse Shivyari et al 2016) micro brils, micro bril bundles, micrometer-wide ribbons (Jiang et al 2013) and larger aggregates. The hieratical nomenclature herein follows recent literature (Song et al 2020) (Babi et al 2023) that describe micro brils as having widths of 2-5 nm; micro ber bundles are tens of nm wide and ribbons greater than 100 nm wide.…”