Technical field

This disclosure relates generally to methods for treating bacterial nanocellulose, such as dissolution of bacterial nanocellulose

Background

The global textile industry is reported to be one of the most polluting industries in the world; from deforestation of old-growth woodlands, to the overuse of natural freshwater supplies, and the non-biodegradability of synthetic fibres. New and environmentally sustainable alternatives for producing textiles are needed to overcome these global challenges and take some of the environmental pressures of the textile industry.

Currently, there are three major fibres used in the textile industry: protein-based (wool, silk, etc); synthetic (polyester, nylon, etc); and cellulosic (cotton, regenerated cellulose, etc). Protein-based fibres, such as wool, are associated with large quantities of water and chemical usage from the irrigation of the pastoral land, drinking supply, the post-harvest processing of the fibres, and the clearing of vast native bushlands to seed monocultural pastoral species. Synthetic fibres are generally formed from non-renewable crude oil-based feedstocks and, after the use of some harmful chemicals in manufacturing, these synthetic fibres can remain in the environment for several hundred years once discarded. Cotton, a cellulosic fibre, is biodegradable, but cotton’s environmental impacts include the huge amount of land, water, and chemicals used for horizontal farming techniques. Regenerated fibres, such as viscose and lyocell, are also biodegradable cellulosic fibres and are regenerated from trees and other plant species. However, large amounts of woodlands are cut down in the production of cellulose-derived fibres.

The industrial processes that are used to dissolve and regenerate these fibres, for example, to produce fibres for the textile industry, come with a very large environmental cost. Large amounts of energy are required to break down trees and other plant materials to make it suitable for the regenerating process. Excessive amounts of sodium hydroxide, in the case of viscose, are used in the dissolution process and discarded into the environment as waste. Other polluting chemicals such as carbon disulphide and ionic liquids are used to assist dissolution of the cellulose polymer and are discarded as waste. Xanthation is one common method used to help dissolve and regenerated cellulose fibres, where xanthation forms xanthated cellulose. Irrespective of the fibre type, fibres can be used to make woven and non-woven materials. Woven materials are initially made by first making a fibre, and are either spun as a raw material (e.g. cotton and wool) or regenerated via chemical dissolution and extruded into a filament (e.g. viscose and polyester). These spun fibres are then woven or knitted into a woven material e.g. denim and cardigans. Woven materials are often made on a loom where fibres are arranged at a right angle to each other; the weft and the warp. These fibres can also be made into a nonwoven by bonding the fibres together using chemical, mechanical, heat or solvent treatments. Examples of nonwovens are medical gowns, filters and absorbency materials such those used in nappies.

Bacterial nanocellulose (BNC; also referred to as bacterial cellulose, microbial nanocellulose and microbial cellulose) is a type of cellulose that is formed from cellulose nanofibrils, and are produced by non-hazardous bacteria belonging to the Acetic Acid Bacteria (AAB) family such as, but not limited to, Komagataeibacter, Gluconacetobacter and Acetobacter. AAB’s are Gram-negative, rod-shaped, strictly aerobic bacteria that are best known for their prolific production of BNC. The structure of the BNC fibrils is chemically and physically similar to plant-based cellulose fibres in many aspects except having a much smaller fibril diameter. Plant-based cellulose has fibres with a diameter in the range of 10-20pm, whereas BNC has been reported to have a diameter ranging from 20-1 OOnm. The other major differences between BNC and many plant-based celluloses is that BNC has a higher degree of polymerisation (DP) and a lower polydispersity (PD).

The DP of cellulose polymers depends greatly on its source. The DP for cellulose from wood pulp is between 300 - 1 ,700, from cotton is 800 - 10,000, and from regenerated fibres 250 - 500. The DP of BNC polymers can reach levels of 8,000 - 10,000. The high DP makes BNC more difficult to defibrillate and solubilise compared to lower DP cellulose sources.

There are two main methods of dissolving plant-based cellulose, either derivative or direct dissolution. Derivative dissolution involves the production of an intermediate, like alkali cellulose and sodium cellulose xanthate in the viscose process, before the cellulose is dissolved. However, these intermediates can cause environmental issues. They also require additional processing steps, such as xanthation to form sodium cellulose xanthate followed by regeneration to reform cellulose and carbon disulfide. Direct dissolution of cellulose using chemicals similar to N-Methylmorpholine N-oxide (NMMO) dissolves cellulose without any intermediates and is believed to be less damaging to the environment. An increase in DP makes direct dissolution more difficult due to increased entanglement of individual fibres. Plant-based cellulose sources including, wood pulp, cotton linters or bamboo, that are commonly used to regenerate cellulose fibres are generally still present in their lumen state before entering the regeneration process. The lumen state of plant-based cellulose is regarded as still retaining its cellular structure. This cellular structure and state may possibly affect the reactivity of the cellulose polymers in the dissolution procedure.

Viscose processes to regenerate fibres from plant-based sources have been used for over one hundred years. Cellulose polymers are macerated with a high concentration of sodium hydroxide to form alkali cellulose that causes the cellulose polymers to swell. Swelling makes xanthation with carbon disulphide easier to form a sodium cellulose xanthate that can then be completely dissolved in a weaker sodium hydroxide solution to form a dope. The aging or reduction of the cellulose polymer length (DP) is also another critical factor for complete dissolution using the viscose process. If the polymers are too short the resulting regenerated fibres may not have adequate tenacity and/or suitable properties for the textile industry, whereas if the cellulose polymers are too long they may not dissolve. The general rule of thumb is that the smaller the cellulose polymer’s DP, the easier it is to dissolve.

Typically, the cellulose polymers used in the viscose process are pre-aged to reduce the length of the cellulose polymer to assist in dissolution in the final dope. However, the downside of reducing the cellulose polymer’s DP is that it can reduce the desirable properties of regenerated fibres such as fibre strength.

In the case of ionic liquids and other organic solvents (e.g. NMMO), for the direct dissolution of plant-based cellulose polymers, it is important that the polymer’s DP are at a specific length before the process begins. If the cellulose polymers are too short the resulting regenerated fibres may not have the required tenacity, and if the cellulose polymers are too long they may not dissolve. The mechanisms and understanding of direct dissolution methods of plant-based cellulose fibres are not as well understood as the much older viscose process.

Direct dissolution of cellulose with a high DP is desired by the textile industry due to the minimal use of hazardous chemicals, reduced processing times, and the ability to form fibres with desirable properties. However, to date, direct dissolution of cellulose has not been possible and instead has required xanthation to form xanthate intermediates. Xanthation increases production costs and has a number of significant environmental issues. It is to be understood that, if any prior art publication is referred to herein, such reference does not constitute an admission that the publication forms a part of the common general knowledge in the art, in Australia or any other country.

Summary

Disclosed is a process of defibrillating bacterial nanocellulose, comprising: providing a pulp of bacterial nanocellulose and an alkaline solution; and freezing the pulp to form a frozen pulp.

Disclosed is a process of defibrillating bacterial nanocellulose, comprising: providing a pulp of bacterial nanocellulose and an alkaline solution; freezing the pulp to form a frozen pulp; and allowing the frozen pulp to thaw to form a solution having at least partially defibrillated bacterial nanocellulose fibres.

By “defibrillating” and its variants such as “defibrillation” and “defibrillate”, it is meant at least partially separating fibrils of nanocellulose, for example from bacterial nanocellulose fibres into bacterial nanocellulose fibrils, bacterial nanocellulose macrofibrils into microfibrils, and/or into bacterial nanocellulose polymer chains. Defibrillation may result in nanocellulose fibres being dispersed and/or solubilised in a solution.

The process of defibrillating bacterial nanocellulose may be a direct process. By “direct process” it is meant that chemical modification of BNC to form an intermediate cellulose species, such as xanthation to form cellulose xanthate, is not required to defibrillate the BNC. In this way, an embodiment of the process may only require use of an alkaline solution as chemical treatment to dissolve or at least partially defibrillate bacterial nanocellulose.

The pulp may comprise up to 4 wt.% bacterial nanocellulose. The alkaline solution may comprise a metal hydroxide. The metal of the metal hydroxide may be a monovalent cation. The pulp may comprise 0.1 wt.% up to 11 wt.% metal hydroxide. The pulp may comprise 0.1 wt.% up to 3.0 wt.% metal hydroxide. The at least partially defibrillated bacterial nanocellulose fibres may be in the form of a mass of non-woven bacterial nanocellulose fibres. The mass of non-woven bacterial nanocellulose may be in the form of a sheet. A shape of the mass of non-woven bacterial nanocellulose may be determined by a shape of a mould in which the pulp is frozen. The pulp may comprise 3.0 wt.% up to 11.0 wt.% metal hydroxide. The at least partially defibrillated bacterial nanocellulose fibres may be solubilised. The process may further comprise diluting the solution of having at least partially defibrillated bacterial nanocellulose fibres with water to cause the at least partially defibrillated bacterial nanocellulose fibres to fall out of the solution and collapse in on themselves to form a solid material. After drying, the solid material may have a density ranging from about 0.03 g/cm ³ to about 1 .5 g/cm ³ .

The metal hydroxide may be a monovalent metal hydroxide. The monovalent metal hydroxide may be sodium hydroxide, lithium hydroxide, or potassium hydroxide. The step of providing the pulp may include forming the pulp by macerating solid bacterial nanocellulose in an alkaline solution. The process may further comprise washing the solid bacterial nanocellulose prior to forming the pulp. The pulp may be formed at a temperature of 50 ^e C or less. The solid bacterial nanocellulose may have a degree of polymerisation (DP) >500 or >1000. The solid bacterial nanocellulose may have a degree of polymerisation ranging from 1 ,000-10,000. The disclosed process may not need to “age” the nanocellulose to reduce the degree of polymerisation prior to freezing. The solid bacterial nanocellulose may be in the form of flakes.

The pulp may be frozen at a temperature ranging from -30 ^e C to -10 ^e C. The frozen pulp may be thawed at a temperature of 30 ^e C or less.

The process may further comprise increasing a solids content of the pulp prior to forming the frozen pulp, for example by removal of at least some of the alkaline solution from the pulp. The process may further comprise degassing the pulp prior to forming the frozen pulp.

At least some of the defibrillated nanocellulose fibres may be dissolved after thawing. The defibrillated nanocellulose fibres may be completely dissolved. An embodiment of the process may be free from using pressure vessels and/or the use of sulphates/sulphites. The steps of forming the pulp and freezing the pulp may be free from using pressure vessels and/or the use of sulfate/sulphites. The step allowing the frozen pulp to thaw may be free from using pressure vessels and/or the use of sulfate/sulphites. The steps of forming the pulp, freezing the pulp, and allowing the frozen pulp to thaw, may be free from using heating steps. The defibrillated nanocellulose fibres may form part of a dispersion.

Disclosed is a process of defibrillating bacterial nanocellulose, comprising: providing a pulp comprising bacterial nanocellulose and an alkaline solution; freezing the pulp to form a frozen pulp; and allowing the frozen pulp to thaw to form a solution of bacterial nanocellulose polymer chains. Disclosed is a process of defibrillating bacterial nanocellulose, comprising: providing a pulp comprising bacterial nanocellulose and an alkaline solution; freezing the pulp to form a frozen pulp; and allowing the frozen pulp to thaw to form a mass of non-woven material of bacterial nanocellulose fibres. The mass of non-woven material of bacterial nanocellulose fibres may be a sponge.

Disclosed is a process of defibrillating bacterial nanocellulose, comprising:

(i) providing a pulp comprising bacterial nanocellulose and a monovalent metal hydroxide solution; and

(ii) freezing the pulp to form a frozen pulp. An embodiment of the process may consist only of steps (i) and (ii).

Disclosed is a process of defibrillating bacterial nanocellulose, comprising:

(i) providing a pulp comprising bacterial nanocellulose and a monovalent metal hydroxide solution;

(ii) freezing the pulp to form a frozen pulp; and

(iii) allowing the frozen pulp to thaw to form a solution of at least partially defibrillated bacterial nanocellulose fibres. An embodiment of the process may consist only of steps (i)- ('")

Disclosed is a solution of dissolved bacterial nanocellulose formed using the process as set forth above.

Disclosed is the use of the solution as set forth above. Disclosed is a cellulosic material formed using the solution of dissolved bacterial nanocellulose. Disclosed is a cellulosic material comprising defibrillated bacterial nanocellulose, the at least partially defibrillating bacterial nanocellulose may be formed using the process as set forth above.

Brief description of figures

Embodiments of the disclosure will now be described, by way of example only, with reference to the following non-limiting Figures.

Figure 1 shows a photo of 1 wt/wt% bacterial nanocellulose in 0 wt/wt% NaOFI solution after maceration for 3 minutes and using polarising microscopy, where the BNC fibre was combed in one direction (a) and at 90 degrees to the first combing (b) showing no alignment. Figure 2 shows a photo of a1 wt/wt% bacterial nanocellulose in 6 wt/wt% NaOH solution after maceration for 3 minutes and using polarising microscopy, where the BNC fibre was combed in one direction (a) and at 90 degrees to the first combing (b) showing alignment in the direction of the combing.

Detailed description of embodiments

Embodiments of the disclosure are directed to processes that can be used to defibrillate bacterial nanocellulose (BNC). In an embodiment, the process includes the steps of: providing a pulp comprising bacterial nanocellulose and an alkaline solution; freezing the pulp to form a frozen pulp; and allowing the frozen pulp to thaw to form a solution of at least partially defibrillated bacterial nanocellulose fibres. The defibrillated bacterial nanocellulose fibres may be dissolved after thawing.

BNC polymers generally have a higher molecular weight, giving rise to a higher DP, than many plant-based cellulose polymers and regenerated cellulose polymers. A high DP typically decreases the solubility of cellulose. However, the inventor has discovered that the different properties of BNC compared to plant-based cellulose affects its dissolution.

Without being bound by theory, it is thought that defibrillation of bacterial nanocellulose behaves in a similar way to the dispersion of very long clays such as kaolinite. In particular, without being bound by theory, it is thought that the use of an alkaline solution, such as a metal hydroxide, means that at least some of the hydrogens on the hydroxyl groups on the cellulose polymer chains are replaced with the metal ion (i.e. a cation) of the metal hydroxide forming a metal-substituted hydroxyl group (e.g. a metal alkoxide) on the cellulose polymer chains, where the metal cation provides electrostatic repulsion between adjacent cellulose polymer chains which causes adjacent cellulose polymer chains to move away from one another to facilitate hydration of the cellulose polymer chains. Freezing is thought to help cause the nanocellulose fibres to defibrillate further due to the formation of ice crystals between adjacent cellulose polymer chains, causing the adjacent cellulose polymer chains to move ever further apart and become even more hydrated. A cellulose polymer chain first becomes sufficiently hydrated to become dispersed. Further hydration of the polymer chain solubilises the polymer chain. The cations associated with the substituted hydroxyl groups provides sufficient electrostatic repulsion to keep cellulose polymer chains defibrillated (e.g. dispersed and/or dissolved). It should be noted that freezing to cause defibrillation of nanocellulose (or any type of cellulose) is opposite to heating processes typically employed to defibrillate cellulose and the xanthation of cellulose. In an embodiment, a dissolution of BNC may be achieved with no need for solubilising derivatives and additives, shortening of the cellulose polymers DP and/or use of harmful chemicals, such as carbon disulfide used in xanthation, as required for existing processes used to dissolve cellulose. Thus, an embodiment may use considerably less energy, water and chemicals with minimal waste management than the conventional techniques used to regenerate a cellulosic fibre.

Throughout this disclosure, the terms “pulp” and “slurry” are used interchangeably.

Currently, the majority of the global BNC that is industrially grown occurs in South East Asia by Acetic Acid Bacteria (AAB) under static conditions to produce nata de coco cellulose. Static cultures are used for the formation of thick, smooth BNC pellicles at the liquid/air interface and are harvested and diced into cubes of nata de coco cellulose. AAB’s have optimal growing conditions of 25-30 °C, low pH and no light requirements capable of growing in a vertically stacked liquid vat system. BNC derived from AAB’s may be suitably produced by vertical farming techniques giving rise to a lower environmental footprint. In an embodiment, the BNC is derived from nata de coco cellulose.

An embodiment of the process to defibrillate bacterial nanocellulose is not limited by the higher DP or other properties such as the hydrophilicity and difficult filterability of BNC. The BNC used in the disclosed embodiments generally has cellulose polymers with a higher molecular weight and higher DP compared to conventional plant-based cellulose sources. The disclosed process does not defibrillate other sources of plant-based lumen state cellulose commonly used to regenerate cellulose fibres.

In an embodiment, the BNC polymer chains in the pulp prior to freezing may have an average molecular weight (Mw) greater than 100,000 g/mol (DP> 600). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 100,000 g/mol (DP> 1 ,200). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 300,000 g/mol (DP> 1 ,800). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 400,000 g/mol (DP 2,400). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater 500,000 g/mol (DP>3,000). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 600,000 g/mol (DP 3,600). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 700,000 g/mol (DP 4,200). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 800,000 g/mol (DP 4,800). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 900,000 g/mol (DP 5,400). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,000,000 g/mol (DP 6,400). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,100,000 g/mol (DP 7,000). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,200,000 g/mol (DP 7,600). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,300,000 g/mol (DP 8,200). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,400,000 g/mol (DP 8,800). In an embodiment, the BNC polymer chains in the pulp prior to freezing can have an average molecular weight (Mw) greater than 1 ,500,000 g/mol (DP 9,400).

The BNC polymer chains can have an average molecular weight less than 2,000,000 g/mol (DP<~12,000), In an embodiment, the BNC polymer chains have an average molecular weight ranging from about 500,000-600,000 g/mol (DP 3,000-4,000). In an embodiment, at least prior to freezing, the BNC polymer chains in the pulp have a DP ranging from 1 ,000- 10,000. After thawing, the BNC may have a DP ranging from 1 ,000-10,000.

A concentration of BNC in the pulp may be less than 4 wt/wt%. A concentration of BNC in the pulp may be less than 3.5 wt/wt%. A concentration of BNC in the pulp may be less than 3 wt/wt%. A concentration of BNC in the pulp may be less than 2.5 wt/wt%. A concentration of BNC in the pulp may be 2 wt/wt% or less. A concentration of BNC in the pulp may be about 1.9 wt/wt% or less. A concentration of BNC in the pulp may be about 1.8 wt/wt% or less. A concentration of BNC in the pulp may be about 1.7 wt/wt% or less. A concentration of BNC in the pulp may be about 1.6 wt/wt% or less. A concentration of BNC in the pulp may be about 1 .5 wt/wt% or less. A concentration of BNC in the pulp may be about 1.4 wt/wt% or less. A concentration of BNC in the pulp may be about 1.3 wt/wt% or less. A concentration of BNC in the pulp may be about 1.2 wt/wt% or less. A concentration of BNC in the pulp may be about 1 .1 wt/wt% or less. A concentration of BNC in the pulp may be about 1.0 wt/wt% or less. A surprising difference between BNC and other plant-based cellulose sources is that a BNC concentration in the pulp greater than 2 wt/wt% tends to exhibit a non-fluid pulp which may be an undesirable characteristic in processing cellulose fibres. By way of comparison only, a 3 wt/wt% cellulose pulp made from dissolving wood pulp remains a free-flowing fluid pulp. A difference in the DP of the cellulose polymer chains may affect fluidity of the pulp compared to non-bacterial cellulose sources.

The pulp may be formed using an alkaline solution. The alkaline solution may be formed from a metal hydroxide. The metal hydroxide may have a monovalent cation. The metal hydroxide may include sodium hydroxide, lithium hydroxide and/or potassium hydroxide. A concentration of the metal hydroxide may be about 11 wt/wt%. A concentration of the metal hydroxide may be about 10 wt/wt%. A concentration of the metal hydroxide may be about 9 wt/wt%. A concentration of the metal hydroxide may be about 8 wt/wt%. A concentration of the metal hydroxide may be about 7 wt/wt%. A concentration of the metal hydroxide may be about 6 wt/wt%. A concentration of the metal hydroxide may be about 5 wt/wt%. A concentration of the metal hydroxide may be about 4 wt/wt%. A concentration of the metal hydroxide may be about 3 wt/wt%. A concentration of the metal hydroxide may be about 2 wt/wt%. A concentration of the metal hydroxide may be about 1 wt/wt%.

A concentration of the metal hydroxide may be equal to or greater than 0.001 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 0.01 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 0.1 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 1 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 2 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 3 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 4 wt/wt%. A concentration of the metal hydroxide may be equal to or greater than 5 wt/wt%.

A concentration of the metal hydroxide may be equal to or less than 15 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 14 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 13 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 12 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 11 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 10 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 9 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 8 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 7 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 6 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 5 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 4 wt/wt%. A concentration of the metal hydroxide may be equal to or less than 3 wt/wt. In an embodiment, a concentration of metal hydroxide ranges from 0.001 wt/wt% to 3.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 0.01 wt/wt% to 3.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 0.1 wt/wt% to 3.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 1 .0 wt/wt% to 3.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 3.0 wt/wt% to 11.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 3.0 wt/wt% to 9.0 wt/wt%. In an embodiment, a concentration of metal hydroxide ranges from 6.0 wt/wt% to 8.0 wt/wt%. The alkaline solution may be formed from metal hydroxide with a concentration of about 6 wt/wt%.

A metal hydroxide concentration greater than about 3 wt/wt% may form a solution of dispersed and/or solubilised nanocellulose fibres (e.g. dissolved cellulose polymer chains) after the freeze-thaw steps. A metal hydroxide concentration up to about 3 wt/wt% may form a dispersion of BNC polymer chains. A metal hydroxide concentration up to about 3 wt/wt% may form a non-woven material after the freeze-thaw steps. In an embodiment, sodium hydroxide, potassium hydroxide or lithium hydroxide may be used to form a non-woven material of BNC. A combination of sodium hydroxide and/or potassium hydroxide and/or lithium hydroxide may be used to form a non-woven material of BNC.

The non-woven material may act as a sponge. The non-woven material may act as a cell scaffold. When a non-woven material is formed, an embodiment of the disclosed process may further comprise blending or crushing or grinding the dried non-woven material to form flocks, fleece and/or microcarriers. Generally, increased blending or crushing or grinding decreases a particle size of the non-woven material. The non-woven material may have a density of about 0.02 to about 0.09 g/cm ³ . In an embodiment the non-woven material has a density of about 0.03 g/cm ³ . The non-woven material may hold about 30-40 times its weight in water. For example, about 1 g of non-woven material may hold about 30 g - 40 g of water.

Without being bound by theory, for the formation of a non-woven material such as a sponge, it is thought that upon freezing the nanocellulose polymer chains are separated due to the expansion of water between adjacent cellulose chains which causes hydrogen bonds between adjacent cellulose chains to be broken. The presence of metal hydroxides during freezing means that at least some of the hydrogens on the hydroxyl groups on the cellulose are replaced with the metal ion of the metal hydroxide forming a metal-substituted hydroxyl group on the cellulose polymer chains. The electrostatic charge(s) of the metal-substituted hydroxyl group on the cellulose, in addition to the frozen state of the pulp of cellulose, prevents the reformation of hydrogen bonding between adjacent nanocellulose polymer chains. However, if there is an insufficient amount of metal hydroxide and/or cations, the nanocellulose polymer chains are not sufficiently hydrated to allow dissolution, which results in a dispersion of nanocellulose polymer chains. The formation of ice crystals between adjacent polymer chains is thought to form a network of interconnected pores through the dispersion of nanocellulose polymer chains. It is thought that upon thawing, hydrogen bonding between adjacent nanocellulose polymer chains reforms thereby forming the non- woven material (e.g. sponge) of nanocellulose polymer chains. It should be noted that the disclosed process of forming a non-woven material differs from freeze-drying and conventional methods for forming porous materials. Specifically, the disclosed process does not require a drying step, such as freeze-drying, to form a non-woven sponge-like material.

An alkaline solution concentration greater than 3 wt/wt% may result in the formation of a solution of dissolved BNC after the freeze-thaw steps. Generally, plant-based cellulose polymers of any known source do not dissolve in a solution of sodium hydroxide under normal conditions and ambient temperature. Surprisingly, the inventor has discovered that in some embodiments freezing and then thawing the cellulose pulp can completely dissolve the BNC cellulose polymer chains to produce a viscous gel using processing conditions that otherwise would not have formed a viscous gel for plant-based cellulose. The gel may be used in the production of a regenerated fibre, sheet or other cellulosic composites. In an embodiment, an alkaline solution comprising sodium hydroxide or lithium hydroxide may be used to form a solution of BNC. An alkaline solution comprising a combination of sodium hydroxide and lithium hydroxide may be used to form a solution of BNC.

Following the formation of a solution of BNC, the process may further comprise diluting the solution of BNC to cause the at least partially defibrillated bacterial nanocellulose fibres to fall out of solution (e.g. precipitate) and collapse in on themselves to form a solid material. The solid material may be dried to form a hard material. The dried solid material may be considered a solid mass of BNC. A density of the dried solid material may be about 1.5g/cm ³ . Without being bound by theory, it is thought that dilution reduces a concentration of metal hydroxide to allow re-protonation of the hydroxyl groups on the cellulose which is accompanied by an increase in hydrogen bonding between adjacent cellulose polymer chains. Without being bound by theory, it is also thought that dilution causes a decrease in metal ion concentration which allows adjacent polymer chains to stack together due to the decrease in electrostatic repulsion provided by the metal ion.

Unlike BNC, hemicellulose and lignin are a major component in plant-based cellulose sources and are required to be removed before the regeneration process. Accordingly, in an embodiment, the disclosed process does not require any pre-treatment step prior to forming the pulp. However, the pulp may be subject to a cleaning step to remove minor components, such as fats and proteins.

The pulp may be formed by homogenising solid BNC in an alkaline solution. Homogenising the BNC may include mechanical, ultrasound or pressure. Homogenising may include a process that applies high shear mechanical force. Homogenising may include maceration. Homogenising may be performed at room/ambient temperature. The pulp may be formed at a temperature of 50 ^e C or less. A temperature of the solution used to form the pulp may be 50 ^e C or less during homogenising. The pulp may be formed at a temperature of 40 ^e C or less. The pulp may be formed at a temperature ranging from about 10 ^e C to about 40 ^e C.

Mechanical homogenisation may be best suited to macerate the BNC into the pulp using high speed rotating blades. The high-speed rotating blades collide with the BNC, shearing and defibrillating the BNC flakes into the solution. One small-scale example of mechanical homogenisation using high speed rotating blade is a kitchen blender/juicer, e.g. NutriBullet Rx.

Maceration of the BNC to produce a homogenised cellulose pulp results in a decrease in a particle size of the BNC. A properly homogenised cellulose pulp is one that has totally macerated all of the densely packed BNC flakes or sheets into cellulose fibrils.

Without being bound by theory, it is thought that mechanical homogenisation, such as maceration, in the presence of an alkaline solution defibrillates and disperses the BNC fibrils in the pulp. It is thought that the use of divalent ion may act as a crosslinking agent in some circumstances which could prevent dispersion and dissolution.

The time of mechanical homogenisation using high speed, high shear force blades to macerate the BNC flakes may be greater than 30 seconds. The time of mechanical homogenisation using high speed, high shear force blades to macerate the BNC flakes may be greater than 1 minute. The time of mechanical homogenisation using high speed, high shear force blades to macerate the BNC flakes may be greater than 2 minutes. The time of mechanical homogenisation using high speed, high shear force blades to macerate the BNC flakes may be greater than 3 minutes. The time taken to homogenise the BNC generally depends on the type of homogeniser used. Polarising lenses can be used to analyse the extent of maceration. The homogenised cellulose pulp formed after maceration disperses the cellulose fibres/polymers which can expose the reactive hydroxyl groups to metal hydroxides to assist in defibrillation and dissolution.

Once homogenised, the cellulose pulp can optionally be filtered to remove some of the alkaline solution to concentrate the BNC content of the final cellulose pulp prior to freezing. Removal of some of the alkaline solution to concentrate the BNC content of the final cellulose pulp may act as a concentration step. The concentration step may increase the concentration of BNC to be 1 .0 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 1 .25 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 1 .5 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 1 .75 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 2.0 wt/wt% or greater The concentration step may increase the concentration of BNC to be 2.25 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 2.5 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 2.75 wt/wt% or greater. The concentration step may increase the concentration of BNC to be 3.0 wt/wt% or greater. The concentration of BNC after the concentration step may be less than 4.0 wt/wt%. The concentration of BNC after the concentration step may be less than 3.0 wt/wt%. The concentration of BNC after the concentration step may be less than 2.0 wt/wt%.

The concentration step may be performed at a temperature ranging from about 5 ^e C to about 50 ^e C. The concentration step may be performed at a temperature ranging from about 5 ^e C to about 40 ^e C. The concentration step may be performed at a temperature ranging from about 10 ^e C to about 40 ^e C. In an embodiment, the concentration step may be performed at room temperature.

Prior to forming the pulp of alkaline solution and BNC, the BNC may optionally be purified in a washing step using a cleaning agent. The cleaning agent may help to remove non- cellulosic material such as proteins and fats produced by AABs. The cleaning agent may include ionic surfactants and/or non-ionic surfactants. The detergent may include laundry detergents. The detergent may include an alkyl benzene sulfonate, a quaternary ammonium species, a polyoxyethylene detergent, or a glycoside detergent. The cleaning agent may include hydroxide species. Hydroxide species may be used in combination with a detergent. The cleaning agent may be provided as a solution. The washing step may include two or more steps. The first washing step may include washing with a cleaning agent and the second or more washing steps may include washing with purified water to remove the cleaning agent. The washing step may be performed at room temperature. The washing step may be performed at elevated temperatures. The washing step may be performed at a temperature up to 50 °C. The washing step may be performed at a temperature up to 60 °C. The washing step may be performed at a temperature up to 70 °C. The washing step may be performed at a temperature up to 80 °C. The washing step may be performed at a temperature up to 90 °C. The washing step may be performed at a temperature up to 100 °C. The washing step may be performed at a boiling point of the cleaning agent solution. The washing step may be performed for up to 5 minutes. The washing step may be performed for up to 10 minutes.

The washing step may be performed for up to 15 minutes. The washing step may be performed for up to 20 minutes. The washing step may be performed for up to 25 minutes. The washing step may be performed for more than 30 minutes. Following washing, the BNC may be dried. The BNC may be macerated immediately following washing.

The pulp is frozen in embodiments of the disclosed process. The pulp may be flash frozen. The pulp may be frozen using a recirculated freezing solution. The pulp may be frozen in a freezer. The pulp may be frozen by immersing the pulp in a freezing fluid. A freezing fluid may include liquid nitrogen. The pulp may be frozen in a container. The container may be a mould. The mould may be used to form a non-woven BNC material.

Once the pulp is frozen, it is then thawed. The combination of freezing and thawing is referred to as the freeze/thaw steps or freeze/thaw treatment. The pulp may remain frozen for less than 30 seconds. The pulp may remain frozen for less than 1 minute. The pulp may remain frozen for less than 5 minutes. The pulp may remain frozen for less than 10 minutes. The pulp may remain frozen for less than 20 minutes. The pulp may remain frozen for less than 30 minutes. The pulp may remain frozen for less than 40 minutes. The pulp may remain frozen for less than 50 minutes. The pulp may remain frozen for less than 1 hour. The pulp may remain frozen for less than 2 hours. The pulp may remain frozen for less than 3 hours. The pulp may remain frozen for less than 4 hours. The pulp may remain frozen for less than 5 hours. The pulp may remain frozen for less than 10 hours. The pulp may remain frozen for less than 20 hours. The pulp may remain frozen for more than 20 hours. The pulp may be stored in a frozen state until thawing. The pulp may be stored frozen for up to two weeks.

The pulp may be frozen at a temperature less than 0 °C. The pulp may be frozen at a temperature down to -5 ^e C. The pulp may be frozen at a temperature down to -10 ^e C. The pulp may be frozen at a temperature down to -20 ^e C. The pulp may be frozen at a temperature down to -30 ^e C. The pulp may be frozen at a temperature down to - 0 °C. The pulp may be frozen at a temperature down to -50 °C. The pulp may be frozen at a temperature down to -60 °C. The pulp may be frozen at a temperature down to -50 °C. The pulp may be frozen at a temperature down to -70 °C. The pulp may be frozen at a temperature down to -80 °C. The pulp may be frozen at a temperature down to -90 °C. The pulp may be frozen at a temperature down to -100 °C. The pulp may be frozen at a temperature less than -100 °C. The pulp may be frozen at a temperature ranging from about -20 ^e C to about -30 ^e C. The pulp may be frozen at a temperature ranging from -60 ^e C to -10 ^e C.

Thawing of the pulp may be performed at a melting point of the solution used to form the slurry. Thawing of the pulp may be performed at a temperature less than 0 ^e C. Thawing of the pulp may be performed at a temperature of between 0 ^e C and room temperature. Thawing of the pulp may be performed at a temperature ranging from about 5 ^S C to room temperature. In an embodiment, the frozen pulp is thawed at a temperature ranging from about 5 ^e C to about 10 ^e C. Room temperature as used herein means approximately 15 ^e C - 30 ^e C. Thawing of the pulp may be performed at a temperature greater than room temperature. The pulp may be thawed at a temperature ranging from about 30 °C to about 100 °C The pulp may be thawed at a temperature of 100 °C or less. The frozen pulp may be thawed using a thawing solution. The thawing solution may be an aqueous-based solution. The thawing solution may have a temperature greater than room temperature. The thawing solution may have a temperature of about 40 °C. The thawing solution may have a temperature of about 50 °C. The thawing solution may have a temperature of about 60 °C. The thawing solution may have a temperature of about 70 °C. The thawing solution may have a temperature of about 80 °C. The thawing solution may have a temperature of about 90 °C. The thawing solution may have a temperature of about 100 °C. The thawing solution may have a temperature greater than 100 °C. The frozen pulp may be immersed in the thawing solution. The frozen pulp may be in a container and the thawing solution may be used to heat the container to thaw the frozen pulp contained in the container. The container may be a mould. The pulp may be heated using a heating fluid. The heating fluid may be steam. The heating fluid may be heated air.

In an embodiment, thawing of the frozen pulp may form a solution of at least partially defibrillated BNC fibres when the metal hydroxide concentration is greater than 3 wt./wt.%.

In an embodiment, thawing of the frozen pulp may form a solution of at BNC polymer chains when the metal hydroxide concentration is greater than 3 wt./wt.%. Thawing processes may result in a solution where the defibrillated BNC fibres are dissolved. However, it should be appreciated that after thawing the solution of defibrillated BNC fibres or solution of BNC polymer chains may include BNC particles, aggregates or structures rather than individual polymer chains, where the particles, aggregates or structures have dimensions in the nanometre scales that are small enough to prevent the scattering of light to give the visual perception of a solution of fully dissolved BNC fibres. The solution of at least partially defibrillated BNC fibres may have BNC fibres that are dissolved and/or suspended. After thawing, the solution of defibrillated BNC fibres may be in the form of a gel, e.g. a BNC gel. The gel may be flowable. The solution of BNC fibres (e.g. the BNC gel) may optionally be filtered. Filtering may remove particulate matter present in the solution of BNC fibres. In an embodiment, thawing of the frozen pulp may form a material formed from non-woven BNC fibres when the metal hydroxide concentration of the pulp is up to 3 wt./wt.%. A shape of the non-woven material may be determined by a mould used to freeze/thaw the pulp. For example, a thin layer of pulp frozen in a tray may produce a sheet of non-woven material after freezing and thawing.

The solution of defibrillated BNC fibres (e.g. a solution of BNC polymer chains) may be used as a feedstock. The feedstock may be used to prepare cellulose-based materials. The cellulose-based materials may include fibres, sheets, sponges, foams and other cellulosic materials. The solution of defibrillated BNC fibres may be filtered prior to use of the solution of defibrillated BNC as a feedstock.

In some embodiments, the pulp is degassed prior to freezing. Degassing may help to reduce the occurrence of outgassing of any dissolved gas during thawing and/or when the solution of defibrillated BNC is used as a feedstock. For example, when the defibrillated BNC solution is used as a feed stock to prepare cellulose fibres, degassing of the pulp prior to freezing may help to reduce or eliminate outgassing of the gas dissolved in the feedstock during fibre formation.

Examples

Embodiments will now be described by way of illustration only with reference to the following examples.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOFI/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solutions (wt/wt%). Each wash was for 10 minutes using boiling water.

The BNC flakes were placed in a NutriBullet Rx (high shear and cutting, mechanical homogenisation) kitchen type blender with water at different ratios of 0.1 wt/wt%, 0.5 wt/wt%, 1 wt/wt %, 2 wt/wt%, 3 wt/wt%, 4 wt/wt% and 5 wt/wt% BNC content.

The BNC flakes were macerated for 1 minute and assessed for defibrillation. Further defibrillation assessment was completed after 2 and 3 minutes. Complete defibrillation of the BNC fibrils occurred in the 0.1 -2.0 wt/wt% BNC content slurries after 3 minutes. Slurries containing 0.1 -2.0 wt/wt% BNC content did not defibrillate well at less than 2 minutes maceration in the NutriBullet Rx.

Slurries with contents greater than 2 wt/wt% BNC did not adequately defibrillate even after 3 minutes or maceration leaving unmacerated flakes in the slurry. Maceration of greater than 2% BNC slurries was not fluid enough after 0.5 - 1 minute for the slurry to hit the blades adequately to continue maceration.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solutions (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 0 wt/wt%, 1 wt/wt%, 2 wt/wt%, 3 wt/wt%, 4 wt/wt%, 5 wt/wt%, 6 wt/wt%, 7 wt/wt%, 8 wt/wt%, 9 wt/wt%, 10 wt/wt% or 11 wt/wt% NaOH solution. To make a 1 wt/wt% BNC in 1 wt/wt% NaOH: 3g of NaOH dissolved in 294g of deionised water, and 3 grams of dry BNC flakes were added to 297g of the NaOH solution. Complete maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes. This procedure was followed for the remaining reactions with increasing NaOH concentrations.

After 3 minutes maceration, each of the 1 wt/wt% BNC in the 0 wt/wt%, 1 wt/wt%, 2 wt/wt%, 3 wt/wt%, 4 wt/wt%, 5 wt/wt%, 6 wt/wt%, 7 wt/wt%, 8 wt/wt%, 9 wt/wt%, 10 wt/wt% or 11 wt/wt% solutions was analysed using a microscope with a polarising lens. The BNC fibrils were combed using a plastic pointer in one direction and then combed at 90 degrees to the original combing (Figure 1 and Figure 2).

No alignment of the BNC fibres was observed in the 0 wt/wt% NaOFI solution.

After 3 minutes maceration, the samples were frozen until solid in a -20 °C freezer. Once frozen, the samples were thawed to a final temperature of 5 °C. This is referred to as the “freeze/thaw” treatment. Visual observations of the transparency of the thawed solution and dissolution of the BNC were made by eye. The dissolution characteristics at different NaOFI concentrations is outlined in Table 1 .

At 4-5 wt/wt% NaOFI the 1 wt/wt% BNC was moderately dissolved with some gelling observed. At 6-8 wt/wt% NaOFI the BNC dissolved to form a clear solution. At 9-11 wt/wt% NaOFI the BNC showed less dissolution and appeared to flocculate with increasing concentration of NaOFI.

1 wt/wt% BNC macerated in 0 wt/wt% NaOFI showed no dissolution of the fibrils after a freeze/thawed treatment was applied. 1 wt/wt% BNC macerated in 6 wt/wt% NaOFI showed complete dissolution of the fibrils after a freeze/thawed treatment was applied.

Table 1 : A dissolution scale used to assess the dissolution of a 1% BNC pulp in various concentrations of sodium hydroxide after thawing.

^* Dissolution scae: 1 : many small separated white clumps of undissolved BNC; 2: an undissolved mass of fibrous BNC mass; 3: an undissolved mass of gelled BNC; 4: a clear mass of gelled BNC; 5: clear solution of dissolved BNC.

A 1 wt/wt% BNC in 6 wt/wt% NaOH solution was made: 18g of NaOH was added into 279g of deionised water; 3 grams of dried BNC was added into 297g of the NaOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

Once macerated, the BNC/NaOH slurry was filtered through a mesh made from nylon fibres, with 100g of the NaOH solution being removed. The remaining BNC/NaOH slurry had a final concentration of 3g BNC and 200g of NaOH solution (1 .5 wt/wt% BNC in 6 wt/wt% NaOH).

The 1.5 wt/wt% BNC in 6 wt/wt% NaOH slurry was frozen until solid at -20 °C. After frozen to a solid state, the BNC/NaOH solid was thawed to a final temperature of 5 °C to form a solution of dissolved BNC.

The solubility of the BNC was tested by visual observation. The BNC was totally dissolved forming a flowable gel-like substance.

Example 4

This example acts as a control using an embodiment of the disclosed process on wood pulp- derived cellulose.

A 1 wt/wt% wood pulp in 6 wt/wt% NaOH solution was made. Adding 18g of NaOH into 279g of deionised water. 3 grams of dried wood pulp was added into 297g of the NaOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

After 3 minutes maceration, the sample was frozen until solid in a -20 °C freezer. Once frozen the samples were allowed to thaw to a final temperature of 5 °C. Visual observations on the transparency of the thawed solution and dissolution of the wood pulp were made by eye showed the wood pulp-derived cellulose did not dissolve forming a suspension of fibres.

Example 5 BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 0.1 , 0.2, 0.4, 0.6, 0.8, 1 wt/wt% NaOH solution. To make a 1 wt/wt% BNC in 0.2 wt/wt% NaOH: 2g of NaOH dissolved in 988g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the NaOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes. This same procedure was followed for each of 0.1 , 0.2, 0.4, 0.6, 0.8 and 1 wt/wt% NaOH solutions.

After 3 minutes of maceration, 500ml of each the 1 wt/wt% BNC in the 0.1 wt/wt% - 1.0 wt/wt% NaOH mixtures were poured into a plastic container with a size of 33 cm length x 21 cm width and frozen until solid in a -20 °C freezer. Once frozen the samples were removed from the freezer and allowed to thaw at room temperature. The samples were then washed three times with water to neutral pH and then dried to provide a sheet formed from non- woven BNC fibres.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 0.4 wt/wt% NaOH solution. To make a 1 wt/wt% BNC in 0.4 wt/wt% NaOH: 4g of NaOH dissolved in 986g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the NaOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes.

After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in the 0.4 wt/wt% NaOH solutions were poured into a plastic container with a size of 33 cm length x 21 cm width and placed in a -20 °C freezer. Once some of the sample had frozen so that a portion was still unfrozen, the sample was removed from the freezer and allowed to thaw at room temperature. The sample was then washed three times with water to neutral pH and then dried to provide a sheet formed from non-woven BNC fibres. The portion of the sample that did not freeze during the freezing step did not form a non-woven material.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 0.4 or 2.0 wt/wt% Ca(OH) ₂ solution. To make a 1 wt/wt% BNC in 0.4 wt/wt% Ca(OH) ₂ : 4g of Ca(OH) ₂ dissolved in 986g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the Ca(OH) ₂ solution. The same procedure was followed for 2 wt/wt% Ca(OH) ₂ . Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes.

After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in either the 0.4 and 2.0 wt/wt% Ca(OH) ₂ mixtures were poured into a plastic container with a size of 33cm length x 21cm width and frozen until solid in a -20 °C freezer. Once frozen the sample was removed from the freezer and allowed to thaw at room temperature. After thawing the BNC was in the form of aggregated clumps and did not form a non-woven mat.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 0.6 wt/wt% KOH solution. To make a 1 wt/wt% BNC in 0.6 wt/wt% KOH: 6g of KOH dissolved in 984g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the KOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes. After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in the 0.6 wt/wt% KOH mixture was poured into a plastic container with a size of 33 cm length x 21 cm width and frozen until solid in a -20 °C freezer. Once frozen the samples were removed from the freezer and allowed to thaw at room temperature. The samples were then washed three times with water to neutral pH and then dried to provide a sheet formed from non-woven BNC fibres.

A 1 wt/wt% BNC in 6 wt/wt% NaOH solution was made: 18g of NaOH was added into 279g of water; 3 grams of dried BNC was added into 297g of the NaOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

Once macerated, the BNC/NaOH slurry was filtered through a mesh made from nylon fibres, with 10Og of the NaOH solution removed. The remaining BNC/NaOH slurry had a final concentration of 3g BNC and 200g of NaOH solution (1 .5 wt/wt% BNC in 6 wt/wt% NaOH).

The 1.5 wt/wt% BNC in 6 wt/wt% NaOH slurry was frozen until solid at -20 °C. After frozen to a solid state, the BNC/NaOH solid was thawed to a final temperature of 5 °C to form a solution of dissolved BNC. The solution of dissolved BNC was then diluted with water to cause the dissolved BNC to fall out of solution and aggregate to form a solid mass. Upon drying, the solid mass formed a hard material.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 7.0 wt/wt% KOH solution. To make a 1 wt/wt% BNC in 7.0 wt/wt% KOH: 70g of KOH dissolved in 920g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the KOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes.

After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in the 7.0 wt/wt% KOH mixture was poured into a plastic container with a size of 33 cm length x 21 cm width and frozen until solid in a -20 °C freezer. Once frozen the samples were removed from the freezer and allowed to thaw at room temperature. The samples were then washed three times with water to neutral pH and then dried to provide a sheet formed from non-woven BNC fibres.

BNC sheets were produced using Komagataeibacter spp., a Gram-negative rod-shaped bacterium, in liquid static culture conditions using white wine as a culture medium. Once harvested and dried, the BNC sheets were shredded into flakes and washed with a 1 .0% NaOH/1% Biozet washing detergent (Kao Corporation) before rinsing three times with water and finally dried. All washes were a ratio of 1 BNC:50 washing solution (wt/wt). Each wash was for 10 minutes using boiling water.

A solution of 1 wt/wt% BNC was macerated in a 12 wt/wt% KOH solution. To make a 1 wt/wt% BNC in 12 wt/wt% KOH: 120g of KOH dissolved in 870g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the KOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes.

After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in the 12 wt/wt% KOH mixture was poured into a plastic container with a size of 33 cm length x 21 cm width and frozen until solid in a -20 °C freezer. Once frozen the samples were removed from the freezer and allowed to thaw at room temperature. The samples were then washed three times with water to neutral pH and then dried to provide a sheet formed from non-woven BNC fibres.

This example acts as a control using an embodiment of the disclosed process on paper pulp-derived cellulose.

A 1 wt/wt% paper pulp in 0.4 wt/wt% NaOH solution was made. Adding 4.0g of NaOH into 986g of water. 10 grams of dried paper pulp was added into 990g of the NaOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

After 3 minutes maceration, the sample was frozen until solid in a -20 °C freezer. Once frozen the samples were allowed to thaw at room temperature. Visual observations on the transparency of the thawed solution and dissolution of the wood pulp were made by eye showed the wood pulp-derived cellulose did not form a non-woven material.

Example 13

This example acts as a control using the disclosed process on paper pulp-derived cellulose. A 1 wt/wt% paper pulp in 6.0 wt/wt% NaOH solution was made. Adding 60g of NaOH into 930g of water. 10 grams of dried paper pulp was added into 990g of the NaOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

After 3 minutes maceration, the sample was frozen until solid in a -20 °C freezer. Once frozen the samples were allowed to thaw to a final temperature of 5 °C. Visual observations on the transparency of the thawed solution and dissolution of the wood pulp were made by eye showed the wood pulp-derived cellulose did not form a solution of dissolved cellulose.

This example acts as a control using the disclosed process on cotton-derived cellulose.

Examples 13 and 14 were repeated but the paper pulp was replaced with cotton (from cotton balls). Following the freeze-thaw treatment, the cotton did not dissolve or form a non-woven material.

A 1 wt/wt% BNC in 6 wt/wt% LiOH solution was made: 60g of LiOH was added into 930g of water; 10 grams of dried BNC was added into 990g of the LiOH solution and macerated for 3 minutes in a NutriBullet Rx kitchen juicer.

The 1 wt/wt% BNC in 6 wt/wt% LiOH slurry was frozen until solid at -20 °C. After frozen to a solid state, the BNC/LiOH solid was thawed to a final temperature of 5 °C to form a solution of dissolved BNC.

The solubility of the BNC was tested by visual observation. The BNC was totally dissolved forming a flowable gel-like substance.

A solution of 1 wt/wt% BNC was macerated in a 1 wt/wt% LiOH solution. To make a 1 wt/wt% BNC in 1 wt/wt% LiOH: 10g of LiOH dissolved in 980g of deionised water, and 10 grams of dry BNC flakes were added to 990g of the LiOH solution. Maceration was achieved using a NutriBullet Rx kitchen blender for 3 minutes.

After 3 minutes of maceration, 500ml of the 1 wt/wt% BNC in the 1 wt/wt% LiOH mixture was poured into a plastic container with a size of 33 cm length x 21 cm width and frozen until solid in a -20 °C freezer. Once frozen the samples were removed from the freezer and allowed to thaw at room temperature. The samples were then washed three times with water to neutral pH and then dried to provide a sheet formed from non-woven BNC fibres. It will be understood to persons skilled in the art of the disclosure that many modifications may be made without departing from the spirit and scope of the disclosure.

In the claims which follow and in the preceding description, except where the context requires otherwise due to express language or necessary implication, the word “comprise” or variations such as “comprises” or “comprising” is used in an inclusive sense, i.e. to specify the presence of the stated features but not to preclude the presence or addition of further features in various embodiments.