Bone analogue

Abstract

A vegetarian bone analogue including non-animal derived protein and a plasticizer that is mimetic of animal bone is provided. Also provided are methods of forming the bone analogue and uses of the bone analogue for example, in a meat analogue.

Description

Bone analogue

[0001] The present invention relates to bone analogues. In particular, vegetarian bone analogues, methods of making such analogues and uses thereof. In addition, the invention relates to meat analogues that include the bone analogues and methods of making meat analogues including the bone analogues.BACKGROUND

[0002] Meat production and consumption can be harmful for the environment and health, so meat alternatives produced from non-animal-derived ingredients are of growing importance and popularity. Some technological solutions have been developed to provide plant-based meat alternatives. Plant-based meat alternatives beneficially provide consumers with similar products and experiences to meat and so reduce the need to change consumer behaviour.

[0003] Part of providing a more realistic meat analogue is the inclusion of animal bone alternatives (referred to as bone analogues). Previously wood and resin bone analogues have been described such as those in US20210298325A1 . However, these bone analogues do not realistically mimic animal bone and can have detrimental effects on the properties of meat analogues including such bone analogues. For example, detrimental effects on the taste and smell of the meat analogues.

[0004] There is a need for improved bone analogues. There is also a need for bone analogues that better mimic animal bones.

[0005] There is also a need for improved bone analogues that do not have detrimental effects on products they are incorporated into. There is a need for bone analogues that are tasteless and / or odourless or have a taste and / or odour similar to animal bones.

[0006] There is also a need for bone analogue that can be more easily incorporated into a meat analogue.BRIEF SUMMARY OF THE DISCLOSURE

[0007] In a first aspect there is provided a vegetarian bone analogue for inclusion in a meat analogue, the meat analogue for human consumption, the vegetarian bone analogue comprising: a. a texturized non-animal derived protein; and b. a plasticizer;wherein the vegetarian bone analogue comprises an anisotropic structure.

[0008] In certain embodiments, the vegetarian bone analogue has a hardness of at least about 500 N.

[0009] In certain embodiments, the vegetarian bone analogue has a moisture content of at most about 15% w / w.

[0010] In certain embodiments, the anisotropic structure comprises an anisotropic fibrous structure.

[0011] In certain embodiments, the vegetarian bone analogue comprises at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent.

[0012] In certain embodiments, the anisotropic fibrous structure comprises denatured nonanimal derived protein.

[0013] In certain embodiments, the vegetarian bone analogue comprises a shape that is mimetic of an animal bone.

[0014] In certain embodiments, the texturized non-animal derived protein is crosslinked.

[0015] In certain embodiments, the vegetarian bone analogue comprises a film disposed on at least a portion of the outer surface of the vegetarian bone analogue.

[0016] In certain embodiments, the film comprises a cartilage analogue.

[0017] In certain embodiments, the film comprises at least one hydrocolloid and / or at least one gum.

[0018] In certain embodiments, the film comprises gellan gum, optionally highly acetylated gellan gum.

[0019] In certain embodiments, the vegetarian bone analogue comprises an internal hollow structure.

[0020] In certain embodiments, the internal hollow structure comprises a vegetarian bone marrow analogue, optionally, wherein the bone marrow analogue comprises a vegetarian liquid composition for mimicking the flavour of bone marrow and a gelling agent.

[0021] In certain embodiments, the vegetarian bone analogue is vegan.

[0022] In certain embodiments, the vegetarian bone analogue has been pasteurized.

[0023] In another aspect there is provided a method of forming a vegetarian bone analogue, the method comprising: a. mixing a non-animal derived protein with a plasticiser to form a bone base material; b. heating and applying a pressure to the bone base material to form a gel and to texturize the non-animal derived protein; c. drying the gel at a temperature and for a time period sufficient to provide a vegetarian bone analogue having a hardness of at least 500N; and d. wherein the vegetarian bone analogue comprises an anisotropic structure.

[0024] In certain embodiments, the temperature and time period are sufficient to provide a bone analogue with a moisture content less than 15%. In certain embodiments, the temperature is less than 100°C. In certain embodiments, the temperature is about 85°C.

[0025] In certain embodiments, the time period is at least 0.5 hours (30 minutes). In certain embodiments, the time period is at most 24 hours.

[0026] In certain embodiments, drying comprises: a. a first drying step comprising drying the gel at a first temperature and for a first time period sufficient to provide a pre-bone analogue having a hardness of less than 500N; and b. a second drying step comprising drying the pre-bone analogue at a second temperature and for a second time period sufficient to provide a vegetarian bone analogue having a hardness of at least 500N.

[0027] In certain embodiments, the method further comprises cooling the bone analogue. In certain embodiments, cooling comprises maintaining the bone analogue at ambient temperature. In some examples, the bone analogue is maintained at ambient temperature until the hardness of the bone analogue has reached a maximum.

[0028] In certain embodiments, step (a) further comprises mixing at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent;with the non-animal derived protein and plasticiser.

[0029] In certain embodiments, the first temperature and first time period are sufficient to provide a pre-bone analogue with a moisture content of less than the bone base material and greater than the bone analogue; and / or the second temperature and second time period are sufficient to provide a bone analogue with a moisture content less than 15%.

[0030] In certain embodiments, step (b) comprises: a. using an extrusion process to heat and apply pressure to the bone base material; b. using a shear cell process to heat and apply pressure to the bone base material.

[0031] In certain embodiments, the extrusion process comprises using a twin-screw extruder.

[0032] In certain embodiments, a. the non-animal derived protein and plasticizer are mixed at a 1 :1 ratio; b. the first temperature is at least 100°C; c. the first time period is of at least one hour; d. the second temperature is less than 100°C or at least 100°C; e. the second time period is at least 5 hours when the second temperature is less than 100°C or the second time period is at least 2 hours when the second temperature is at least 100°C; and / or f. the pressure is around 6x106Pa (60 bar).

[0033] In certain embodiments, the method further comprises shaping the gel after step (b) and before step (c).

[0034] In certain embodiments, the method further comprises crosslinking the gel after step (b) and before step (c).

[0035] In certain embodiments, the method further comprises applying a film to at least a portion of the outer surface of the pre-bone analogue or the formed vegetarian bone analogue.

[0036] In certain embodiments, a. the film comprises a cartilage analogue; and / or b. the film comprises gellan gum, optionally highly acetylated gellan gum.

[0037] In certain embodiments, applying the film comprises:a. depositing a solution comprising at least one hydrocolloid and / or gum onto the portion of the outer surface; and b. drying the deposited solution on the outer surface to form the film.

[0038] In certain embodiments, applying the film further comprises preparing the solution, preparing comprising: a. dissolving the at least one hydrocolloid and / or gum in a solvent to form the solution; and b. cooling the solution to a temperature suitable for increasing the viscosity of the solution before depositing the solution.

[0039] In certain embodiments, drying the gel at the first temperature and for the first time period provides a pre-bone analogue comprising an internal hollow structure and the method further comprises depositing a bone marrow analogue within the internal hollow structure of the pre-bone analogue or the formed vegetarian bone analogue.

[0040] In certain embodiments, the vegetarian bone analogue comprises an anisotropic fibrous structure.

[0041] In another aspect there is provided a vegetarian bone analogue formed by the methods described herein.

[0042] In another aspect there is provided use of a vegetarian bone analogue according as described herein as a bone mimetic in a meat analogue.

[0043] In another aspect there is provided a method of forming a meat analogue comprising a bone analogue, the method comprising: a. providing a first fibrous muscle tissue analogue; b. applying a binder to an upper surface of the first fibrous tissue analogue; c. applying a binder to a vegetarian bone analogue as described herein; d. depositing the vegetarian bone analogue onto the upper surface; e. applying a second analogue on top of the bone analogue and the upper surface of the first fibrous muscle tissue analogue.

[0044] In certain embodiments, the method further comprises setting the binder.

[0045] In certain embodiments, the second analogue comprises a second fibrous muscle tissue analogue and / or fat tissue analogue.

[0046] In certain embodiments, the second analogue comprises a binder on at least one surface of the second analogue, wherein the at least one surface contacts the vegetarian bone analogue.

[0047] In certain embodiments, the first fibrous muscle tissue analogue comprises a fat analogue.

[0048] In certain embodiments, the binder comprises a fat analogue.

[0049] In certain embodiments, the first fibrous muscle tissue analogue and / or the second fibrous muscle tissue analogue comprise texturized non-animal derived protein, wherein at least some of the fibres of the fibrous muscle tissue analogue are partially separated;

[0050] In certain embodiments, the first fibrous muscle tissue analogue and / or the second fibrous muscle tissue analogue comprise a marinade absorbed into the fibres; and / or

[0051] In certain embodiments, the first fibrous muscle tissue analogue and / or the second fibrous muscle tissue analogue comprise a or the binder interspersed between at least some of the fibres.

[0052] In certain embodiments, a. the fibres of the second fibrous muscle tissue analogue are substantially parallel to the fibres of the first fibrous muscle tissue analogue; or b. the fibres of the second fibrous muscle tissue analogue are substantially nonparallel to the fibres of the fibrous muscle tissue analogue.

[0053] In another aspect there is provided a meat analogue formed by a method described herein.

[0054] In another aspect there is provided a meat analogue comprising a vegetarian bone analogue as described herein.

[0055] Throughout the description and claims of this specification, the words “comprise” and “contain” and variations of them mean “including but not limited to”, and they are not intended to (and do not) exclude other moieties, additives, components, integers or steps.

[0056] Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[0057] Features, integers, characteristics, compounds, chemical moieties or groups described in conjunction with a particular aspect, embodiment or example of the invention areto be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith.BRIEF DESCRIPTION OF THE FIGURES

[0058] Embodiments of the invention are further described hereinafter with reference to the accompanying drawings, in which:

[0059] Figure 1 shows schematic diagrams of bone analogues. A shows a rib bone analogue and B shows a chicken leg bone analogue.

[0060] Figure 2 shows exemplary bone analogues. A shows a rib bone analogue. B shows a “T-bone” analogue. C shows a bone analogue cut through to show the internal hollow structure. D shows a further example shape of a bone analogue.

[0061] Figure 3 shows a meat analogue including bone analogues in situ. A shows an example of a rack of ribs meat analogue including rib bone analogues. B shows a rib meat analogue with bone analogues placed onto a layer of binder on a muscle tissue analogue with further binder applied on top of the bone analogues. C shows a final product of a rack of ribs analogue with the bone analogues disposed between layers of muscle tissue and fat tissue analogues with a fat tissue analogue disposed on the topmost surface.

[0062] Figure 4 shows texturized bone base material (i.e. texturized vegetable protein or gel) after extrusion. As seen, the texturized bone base material has a firm texture, is recovered as a single piece and has an anisotropic structure.

[0063] The patent, scientific and technical literature referred to herein establish knowledge that was available to those skilled in the art at the time of filing. The entire disclosures of the issued patents, published and pending patent applications, and other publications that are cited herein are hereby incorporated by reference to the same extent as if each was specifically and individually indicated to be incorporated by reference. In the case of any inconsistencies, the present disclosure will prevail.

[0064] Various aspects of the invention are described in further detail below.DETAILED DESCRIPTIONBone Analogue

[0065] Provided herein are bone analogues. The term “bone analogue” refers to an article that is a mimetic of a naturally occurring bone, such as animal bone. For example, a bone analogue is an article that may have one or more properties that are similar to or the same as an animal bone. For example, a bone analogue may have physical properties such as hardness and / or hardness that is similar to or the same as naturally occurring bone. Bone analogues may also have structural properties that are mimetic of naturally derived animalbones. For example, shape, colour, appearance and / or texture that is similar to animal bone. Bone analogues may also have organoleptic properties such as aroma (odour), and / or taste (flavour) similar to or the same as animal bone.

[0066] The bone analogues described herein are suitable for use in meat analogues. For examples, in meat analogues that are for consumption by humans. For example, meat analogues as described herein.

[0067] One aim of the bone analogues described herein is to provide an alternative to animal bones. As such the bone analogues described herein may be vegetarian. That is to say, the components are not made from or with the aid of products or components derived from animals that have died, have been slaughtered, or animals that die as a result of being eaten. The bone analogues described herein may be vegan. That is to say; the components are not sourced from or derived from an animal or animal product. Food products that are "vegan" are free of any animal products or animal by-products.

[0068] The bone analogues described herein include a texturized non-animal derived protein. That is to say that the bone analogues described herein include vegetarian or vegan protein. The bone analogues may be made from a bone base material that includes a nonanimal derived protein; the non-animal derived protein may comprise a plant protein, such as a vegetable protein, in particular soy protein. In other examples, the non-animal derived protein may additionally or alternatively comprise a fungal protein, a protein extracted from a microorganism, or a recombinantly produced protein, for example, a microbially produced protein. In examples, the non-animal derived protein may comprise two or more different non- animal derived proteins. The non-animal derived protein may be in pure form of protein isolate, or a protein concentrate. In other examples, the protein may be a defatted meal with a high protein content, such as soybean meal, providing a protein content of greater than about 55%.

[0069] The bone base material may be processed, in particular texturized. Texturization of a non-animal derived protein may create a protein texturate having anisotropically distributed interconnected protein. For example, by applying pressure and / or heat to denature the non- animal derived protein and applying shear forces to form anisotropically distributed interconnected protein. In some examples, texturization of the protein leads to formation of fibres. Therefore, the bone analogues may include a fibrous texturized non-animal derived protein. The fibres may be generally aligned. In some examples, the fibres may be substantially parallel, for example, parallel.

[0070] The texturized non-animal derived protein has an anisotropic structure. The anisotropic structure may provide a bone analogue that more closely mimics animal bone. For example, the anisotropic structure may provide a bone analogue that has a hardness that issimilar to or the same as animal bone. In some examples, the texturized non-animal derived protein has an anisotropic fibrous structure. This textural anisotropy not only sets the bone analogues described herein apart from other high-protein products but also highlights their novelty and significance, especially in contexts where texture plays a pivotal role in product functionality and consumer perception.

[0071] The anisotropic structure of the bone analogues and / or gel they are formed from may be determined by methods known in the art. For example, by microstructural analysis and / or mechanical testing methods of gels used to form the bone analogues or of the formed bone analogues.

[0072] Microstructural analysis methods, such as microscopy methods, may allow visualisation of the internal structure of a bone analogue as described herein or of the gel used to form the bone analogue. For example, anisotropic structure may be determined by scanning electron microscopy (SEM), transmission electron microscopy (TEM), or confocal laser scanning microscopy (CLSM). Such methods may provide information about network structure and direction-dependent characteristics of the bone analogues and / or gel. Further image analysis may also identify patterns of anisotropy indicated by features such as pore size, distribution, and orientation of structural elements.

[0073] Mechanical testing methods include texture analysis and / or rheological analysis. For example, the gels from which the bone analogues are formed or bone analogues may be analysed using a texture analyser to measure properties such as hardness, elasticity, and / or adhesiveness from different directions. Such analysis methods help identify how the structure varies with direction. Rheological analysis may provide insight into the gel’s viscoelastic properties. For example, rheological analysis may include oscillatory rheology, where the gel is subjected to small deformations. Such methods may reveal anisotropies in viscoelastic parameters.

[0074] For example, the bone analogue may have hardness of at least 500 N (about 51 kilogram-force) as measured by a texture analyser such as TA.XTplusC (Stable Micro Systems). In some examples, the bone analogue has a hardness of at least about, 500 N, 600 N or more. In some examples, the bone analogue has a hardness of about 500 N. In some examples, the bone analogue has a hardness of about 600 N. The hardness of the bone analogue of the bone may be dependent on the shape of the bone. For example, a curved or arcuate bone analogue fracture at a lower force than a substantially flat bone analogue. Hardens may be defined as a measure of how mechanically resistant a material is to the mechanical penetration of an indenter or as the resistance of a specific material to localised plastic deformation.

[0075] Methods for measuring hardness are known in the art. In some examples, the hardness is measured using an instrument such as a texture analyser. For example, a TA.XTplusC (Stable Micro Systems). The following setting may be used: load cell: 50Kg pretest speed: 1 mm / sec; test speed: 5mm / sec; strain: 50%. Hardness may be measured and expressed using any one of a shore hardness test, Vickers hardness test or Brinell hardness test. When measuring hardness a force is usually applied whereby either the penetration depth or the size of the indentation caused by an indenter is measured. Using a Texture Analyser the force to go to a set distance in the material may also indicate and be a suitable way to assess hardness. Brinell, Vickers and Shore hardness are all methods that measure material hardness and can be easily determined using a Texture Analyser.

[0076] In some examples, the bone analogue may have a shore hardness of at least 50D (i.e. 50 on shore D scale). The bone analogue may have a shore hardness of at 60D, 70D, 80D, 90D or more. For example a shore hardness of between 50D and 100D. Shore hardness may be measured using any known methods in the field. For example, using a durometer.

[0077] The bone analogue also includes at least one plasticizer. A plasticizer refers to a substance can be used to reduce viscosity which is normally a liquid. In some examples, the plasticizer is water. Other suitable plasticizer include glycerol, polyols and vegetable oils.

[0078] The bone analogue may include a plasticizer at a level of at most 15%. For example, the bone analogue may have a moisture content of at most 15%. For example, at most 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 % moisture content. A moisture content of less than 15% may provide a hard and stiff bone analogue that can be used in meat analogues to recreate a better eating experience by realistically mimicking animal bone.

[0079] The bone analogue may include at least 50% w / w of the texturized non-animal derived protein. In some examples, the bone analogue includes at least 85% w / w texturized non-animal derived protein. For example, at least 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, 59%, 60%, 61 %, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81 %, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% w / w texturized non-animal derived protein.

[0080] Methods that may be used to measure moisture content are well known and include, for example, spectroscopic methods, chemical assays, conductivity assays and thermogravimetric analysis, loss on drying methods and Karl Fischer titration methods.

[0081] In some examples, the bone analogue consists of a texturized non-animal derived protein and water.

[0082] In some examples, the bone analogue has a relatively low fat or oil content in comparison to a meat analogue the bone analogue may be used in. That is to say the bone analogue has a fat content less than a meat analogue. For example, the bone analogue may have a fat content of at most 5%. In some examples, the bone analogue may have a fat content of at most 2%. In some examples, the bone analogue does not have any additional fat (i.e. oils) added to the bone analogue. That is to say the body of the bone analogue, not including any additional components added to the surface and / or within the bone analogue (e.g. within a cavity such as the hollow internal structure) has a fat content as described above.

[0083] The bone analogue may further include one or more additional agents. For example one or more flavouring agents, colouring agents, flavour precursors, Maillard reaction precursors, preservatives, spices, seasonings, hydrocolloids, gums, and / or fibres.

[0084] The hydrocolloids may be selected from one or more of sodium alginate, xanthan gum, carrageenan, gellan gum, konjac glucomannan, citrus fibre, Puratein G®, methylcellulose, hydroxypropylmethyl cellulose or similar. Advantageously, providing a hydrocolloid may help with fibre separation and providing an anisotropic fibrous structure.

[0085] The term hydrocolloid refers to long chain polymers (polysaccharides and proteins) characterised by their property of forming viscous dispersions and / or gels when dispersed in water. Presence of a large number of hydroxyl (-OH) groups markedly increases their affinity for binding water molecules rendering them hydrophilic compounds. Further, they produce a dispersion, which is intermediate between a true solution and a suspension, and exhibits the properties of a colloid. Considering these two properties, they are aptly termed as ‘hydrophilic colloids’ or ‘hydrocolloids’.

[0086] The hydrocolloid may be at least one reversible thermoset hydrocolloid. Reversible or reversibly thermoset hydrocolloid refers to a hydrocolloid which upon heating becomes reversibly more viscous (i.e. sets upon application of heat) and when cooled become less viscous. Examples of such hydrocolloids include modified celluloses such as methylcellulose. Other examples include Avebe (potato protein) Solanic 200 or 300..

[0087] Methylcellulose is a semiflexible cellulose ether derivative. Methylcellulose, as a gel, has sets when hot and melts when cold. The gelation is concurrent with fibril formation upon heating, whereby the Methylcellulose polymer chains self-assemble into fibrils with a relatively consistent mean diameter, independent of polymer concentration, molecular weight, and temperature of gelation.

[0088] The hydrocolloid may be or include at least one thermoreversible hydrocolloid. Thermoreversible refers to a substance that has reversibly decreased viscosity when heated (i.e. melts) and has reversibly increased viscosity when cooled (i.e. gels). For example thehydrocolloid may include any one or more of agar, a carrageenan, gellan gum, or hydroxypropylmethyl cellulose.

[0089] The hydrocolloid may be or include carrageenan. Carrageenans are a family of natural linear sulphated polysaccharides that are extracted from red edible seaweeds. The most common seaweed used to obtain carrageenans is red seaweed Chondrus crispus (Irish moss). Carrageenans contain 15-40% ester-sulphate content, which makes them anionic polysaccharides. They can be mainly categorized into three different classes based on their sulphate content. Kappa-carrageenan has one sulphate group per disaccharide, iota- carrageenan has two, and lambda-carrageenan has three. In some examples, the second hydrocolloid comprises Kappa-carrageenan and / or iota-carrageenan. In some examples, the second hydrocolloid comprises Kappa-carrageenan. The carrageenan may be refined, semirefined or unrefined or combinations thereof.

[0090] In some examples the bone analogue may include at least one gum. For example, guar gum, locust bean gum, carob bean gum, tara gum, acacia gum, xanthum gum, carrageenan, lecithin gum, and / or gellan gum.

[0091] In examples, bone analogues may additionally comprise a fibre, for example pectin or cellulose fibre. In examples, the bone analogues may additionally comprise a carbohydrate, for example starch. Advantageously, addition of a fibre or carbohydrate may help with formation of fibres during texturization processes.

[0092] In some examples, the bone analogue includes a flavouring agent. A "flavouring agent" refers to a compound or salt or solvate thereof, that can be acceptably ingested, that induces a taste and / or smell in an animal or human. The flavouring agent may be natural, semi-synthetic or synthetic.

[0093] The flavouring agent may be a water soluble flavouring agent. The flavouring agent may be an oil soluble flavouring agent. The flavouring agent may be commercially available flavouring agent and may be selected depending on the use. Examples of flavouring agents include acids, amino acids, sugars, reducing sugars (e.g., ribose), vitamins, and minerals. Flavouring agents may also include microorganisms and fungi, such as recombinant microorganisms and yeast.

[0094] For example, the flavouring agent may be selected to mimic the taste and / or aroma of animal bone.

[0095] Colouring agents may be any suitable colouring that is safe for consumption by animals and humans. Examples of suitable colouring agents include titanium dioxide or calcium carbonate.

[0096] Examples of spices and seasonings include salts (such as sodium or calcium chloride), pepper and / or garlic. The amounts and types of spices and seasonings added may be determined by the type of meat analogue the bone analogue is intended for use with.

[0097] In some examples, salts may act as texturization aids. For example, calcium chloride can modify the amount of free water in the final product. In other instances salts may also be used as a colouring agent. For example, calcium phosphate could be used to obtain a whiter colour.

[0098] In some examples, the bone analogue includes a flavouring precursor. A flavour precursors may include one or more of an amino acid, a reducing sugar (e.g., ribose), and a vitamin, for example, vitamin B12. The flavour precursors may be configured to transform or react when heated (i.e. cooked) to release flavours or aromas that mimic animal bone when cooked.

[0099] Maillard reaction, also called the non-enzymatic browning reaction, plays a role in food technology, on the one hand for the typical brown colour of roasted, baked or fried food products and on the other hand for the associated typical aroma. The Maillard reaction is not a single, specific chemical reaction, but a complex set of many reactions that lead to a large number of reaction products. A so-called reaction product is mainly responsible for the desired effects of melanoidins. These always occur in foods when compounds with free carbonyl groups, such as reducing sugars or lipid oxidation products or products from the degradation of polyphenols (o-quinones) and fermentative processes (methylglyoxal) with proteins, peptides and I or amino acids are heated together.

[0100] Examples of Maillard reaction precursors include ribose, xylose, glucose, fructose, arabinose, glucose-6-phosphate, fructose 6-phosphate, fructose 1 ,6-diphosphate, inositol, maltose, sucrose, maltodextrin, glycogen, nucleotide-bound sugars, molasses, a phospholipid, a lecithin, inosine, inosine monophosphate (IMP), guanosine monophosphate (GMP), pyrazine, adenosine monophosphate (AMP), lactic acid, succinic acid, thiamine, creatine, pyrophosphate, vegetable oil, algal oil, corn oil, soybean oil, palm fruit oil, palm kernel oil, safflower oil, flaxseed oil, cottonseed oil, sunflower oil, canola oil, olive oil, a free fatty acid (arachidonic, oleic, palmitoleic fatty acid etc.), cysteine, methionine, isoleucine, leucine, lysine, phenylalanine, threonine, tryptophan, valine, arginine, histidine, alanine, asparagine, aspartate, glutamate, glutamine, glycine, proline, serine, tyrosine, glutathione, an amino acid derivative, hydrolysed vegetable proteins, malt extract, and yeast extract.

[0101] In some examples, the bone analogue consists of a texturized non-animal derived protein, water and / or one or more additional agents as described herein.

[0102] The bone analogue may include one or more cross-linking agents, for example a hydrocolloid, a protein, or a carbohydrate. The cross-linking agent may form cross-links within the bone analogue to bind the fibres together.

[0103] In some examples, the texturized non-animal derived protein may be crosslinked. For example, by a crosslinking agent. For example, by an enzymatic crosslinking agent such as transglutaminase. Transglutaminases (EC 2.3. 2.13) are a class of enzymes that catalyse the formation of a covalent bond between the y-carboxamide group of protein- or peptide- bound glutamine (acyl donors) and the free amine group of protein- or peptide-bound lysine (acyl acceptors).

[0104] The bone analogue may be shaped to any desired shape. In particular, the bone analogue may be shaped to mimic the shape of an animal bone. For example, a shape that is mimetic of a rib or leg bone of an animal such as a chicken or pig. The shape may be a shape mimetic of any bone that may be found in a cut of animal meat. For example, for cuts of beef the shape may be similar to that of bone found in a T-bone steak, a rib steak, a porterhouse steak, shank crosscut, back ribs, rib roast, ox tail, or bone in top loin steak. For example, for cuts of pork the bone analogue may have a shape mimetic of the bone found in a rib chop, a T-bone chop, shank or knuckle, shoulder steak, loin rack joint, spareribs, ribs or a leg joint. For example, for cuts of lamb the shape may be similar to that of bone found in a leg joint, shoulder joint, shank, hind shank, double loin cop, arm chop, foreshank, ribs or riblets.

[0105] Shape not only refers to the outline of the bone analogue but also the curvature, thickness and other structural characteristics. Therefore, the bone analogue may be curved or arcuate.

[0106] The bone analogue may be at least partially covered or coated with a film that mimics animal cartilage (e.g. a cartilage analogue). Such a film may help provide a bone analogue that more closely mimics and animal bone. The cartilage analogue may contain a hydrocolloid and / or a gum as described herein. For example, the film may include one or more of guar gum, locust bean gum, carob bean gum, tara gum, acacia gum, xanthum gum, carrageenan, lecithin gum, gellan gum, sodium alginate, carrageenan, konjac glucomannan, citrus fibre, Avebe (potato protein) Solanic 200 or 300methylcellulose, and / or hydroxypropylmethyl cellulose or similar.

[0107] In some examples, the film includes gellan gum. Gellan gum is a water-soluble anionic polysaccharide produced by the bacterium Sphingomonas elodea. The repeating unit of the polymer is a tetrasaccharide, which consists of two residues of D-glucose and one of each residues of L-rhamnose and D-glucuronic acid. In some examples, the film includeshighly acylated gellan gum. Highly acylated gellan gum has two acyl substituents acetate at C6 and glycerate at C2 on the first glucose unit of the repeating unit of tetrasaccharide, and on average, there is one glycerate per repeat and one acetate per every two repeats.

[0108] In some examples, the film covers only part of the bone analogue. For example, the film may coat parts of the bone analogue that would have cartilage attached for an animal bone. Therefore, mimicking an animal bone and cartilage attached thereto. In some examples, the entirety of the external surface of the bone analogue is coated with the film.

[0109] In some examples, the bone analogue has a hollow internal structure. That is to say that the bone analogue has a void encased by the external surfaces of the bone analogue. In some examples, the bone analogue has at least one opening that extends through at least one surface into the hollow internal structure. In some examples, the bone analogue may include two openings that extend into the hollow internal structure. For example, openings within opposing surfaces of the bone analogue.

[0110] The internal hollow structure may be filled with a substance. For example, the hollow structure may be filled with a bone marrow analogue. This may help provide a bone analogue that more closely resembles or mimics an animal bone. The bone marrow analogue may also provide organoleptic properties.

[0111] A bone marrow analogue may be a substance that consists of one or more hydrocolloids or gels as described herein and a fat. For example, bone marrow analogues may have a high fat content that replicates the fat content of animal derived bone marrow. In some examples, a bone marrow analogue may have a fat content that replicates the mouthfeel and texture of animal derived bone marrow. For example, one or more additional agents as described herein. For example, seasonings, flavourings agents, colouring agents flavour precursors, Maillard reaction precursors, preservatives, and / or spices as described herein.

[0112] Examples of bone marrow analogues include a gum (e.g. konjac, xanthan gum, agar, guar gum) mixed with a vegan broth. For example, a liquid composition that mimics the flavor of bone marrow or meat that is gelled with a gum. In some examples, the liquid composition may be in the form of an emulsion gel or oleo gel which comprises a gelling agent such as a gum or hydrocolloid as described herein.

[0113] The inclusion of the film may also provide the advantageous effect of improving adhesion of the bone analogue to a meat analogue when in use.

[0114] In some examples, the bone analogue may be pasteurized. Pasteurized refers to a method of treating edible materials, generally by heating them (alternatively in some instances by gamma irradiating) to a certain point to kill pathogenic microorganisms but not harm theflavour or quality of the bone analogue. Pasteurization may help reduce the risk of any adverse effects on a person who consumes a meat analogue including the bone analogue.Method of forming bone analogue

[0115] Also provided herein are methods of forming a bone analogue as described. The methods include providing a non-animal derived protein such as a plant protein, fungal protein and / or microbial protein (such as a recombinant protein). The non-animal derived protein may comprise a plant protein, such as a vegetable protein, in particular soy protein. In other examples, the non-animal derived protein may additionally or alternatively comprise a fungal protein, a protein extracted from a microorganism, or a recombinantly produced protein, for example, a microbially produced recombinant protein such as beef myoglobin. In examples, the non-animal derived protein may comprise two or more different non-animal derived proteins. The non-animal derived protein may be in pure form of protein isolate, or a protein concentrate. In other examples, the protein may be a defatted meal with a high protein content, such as soybean meal, providing a protein content of greater than about 55%.

[0116] The non-animal derived protein is mixed with a plasticizer such as water to form a bone base material. In some examples, the plasticizer and non-animal derived protein are mixed at a ratio of 1 : 1. For example, the method may involve mixing 50% w / w of a non-animal derived protein with 50% water.

[0117] In some examples, the other additional agents as described herein may be mixed with the non-animal derived protein and plasticizer. For example, one or more hydrocolloids, gums, fibres, and / or flavouring agents. For example, one or more of sodium alginate, xanthan gum, carrageenan, gellan gum, konjac glucomannan, citrus fibre, Puratein G®, methylcellulose, hydroxypropyl methyl cellulose or similar may be mixed with the non-animal derived protein and plasticizer. In some examples, a fibre, for example, pectin or cellulose fibre may be mixed with the non-animal derived protein and plasticizer. In some examples, a carbohydrate, for example, starch may be mixed with the non-animal derived protein and plasticizer. Advantageously, addition of a fibre or carbohydrate may help with formation of protein fibres during a texturization process. In examples, the bone base material may not include any additional oil. For example, no additional oil may be mixed with the non-animal derived protein and plasticizer. As such, the oil content of the bone base material may be the same as the oil content of the non-animal derived protein.

[0118] The non-animal derived protein and plasticizer (bone base material) may then be processed in order to texturize the non-animal derived protein. For example, the process may include applying pressure and heat to the bone base material. The application of heat and pressure may lead to the denaturation of the non-animal derived protein and lead to theformation of fibres. In some examples, the fibres may be aligned parallel or substantially parallel to each other.

[0119] Heating and applying pressure to the bone base material may also lead to the bone base material transitioning to a gel. A gel refers to a colloidal system in which a network of particles spans the volume of a liquid medium. Although gels mainly are composed of liquids, and thus exhibit densities similar to liquids, gels have the structural coherence of solids due to the network of particles that spans the liquid medium. For this reason, gels generally appear to be solid, jelly-like materials.

[0120] The bone base material may be heated to a “high-temperature” that is greater than the glass transition temperature of the bone base material. In examples of the bone base material may be heated to at least 50°C. In examples of the bone base material may be heated to at least 60°C. In examples of the bone base material may be heated to at least 90°C. In some examples, the bone base material is heated to about 50°C to 170°C, for example 60°C to 160°C In some examples, the bone base material is heated to about 90°C to 170°C, for example 90°C to 160°C. For example about, 50, 51 , 52, 53, 54, 55, 56, 5, 58, 59, 60, 61 , 62, 63, 64, 65, 66, 67, 68, 69, 70, 71 , 72, 73, 74, 75, 76, 77, 78, 79, 80, 81 , 82, 83, 84, 85, 86, 87,88, 89, 90, 91 , 92, 93, 94, 95, 96, 97, 98, 99, 100, 101 , 102, 103, 104, 105, 106, 107, 108,109, 110, 111 , 112, 113, 114, 115, 116, 117, 118, 119, 120, 121 , 122, 123, 124, 125, 126,127, 128, 129, 130, 131 , 132, 133, 134, 135, 136, 137, 138, 139, 140, 141 , 142, 143, 144,145, 146, 147, 148, 149, 150, 151 , 152, 153, 154, 155, 156, 157, 158, 159, 160°C.

[0121] Pressure may be applied mechanically, for example by a screw drive in an extrusion process. Alternatively or additionally, the pressure may be generated by expansion of the bone base material within a constrained volume, such as in a shear cell. Alternatively or additionally, the pressure may be generated by water vapour created by the heating. For example, water vapour pressure. Heat may be applied directly, for example by heating a part of the texturization apparatus, and / or heat may be generated by applying pressure and / or shear forces to the bone base material due to friction.

[0122] In some examples, the pressure may be from about 4x106Pa to about 1.45x108Pa (i.e. 40 to 100 Bar). In some examples, the pressure may be about 6x106Pa (about 60 bar).

[0123] In some examples, the method involves the use of an extrusion apparatus to apply heat and pressure to the bone base material. Extrusion apparatus may include a material feed, such as a hopper, that communicates with a generally tubular barrel. The barrel includes an elongate tube having an inlet corresponding to the material feed and an outlet provided by a die. One or more screws, rotatably driven by a motor, may be positioned in the barrel and acts to push bone base material from the inlet towards the die and out of the outlet. The screw orscrews may have a variable pitch screw thread that acts to compress the bone base material as it moves along the barrel. The barrel may be heated. The bone base material is thereby heated and pressurised in the barrel and forced out of the die. The heat and pressure act to denature the protein and as the denatured protein cools and anisotropically gels in the die it forms into an anisotropic pre-bone analogue. In some examples, when the textured protein includes fibres or is formed into a fibrous texturized protein the denatured protein may gel to form into an anisotropic fibrous pre-bone analogue.

[0124] In some examples, a twin screw extruder may be used. The screws of a twin-screw extruder can rotate within a barrel in the same or opposite directions. Rotation of the screws in the same direction is referred to as single flow, whereas rotation of the screws in opposite directions is referred to as double flow or counter rotating. The speed of the screw or screws of the extruder may vary depending on the particular apparatus.

[0125] The extrusion apparatus (e.g. twin screw extruder) generally comprises a plurality of heating zones through which the bone base material is conveyed through under mechanical pressure prior to exiting the extrusion apparatus through an extrusion die assembly. The temperature in each successive heating zone (i.e. in the barrel) may be different to a previous zone, barrel or part of a single barrel.

[0126] In some examples, the bone base material may be conveyed or passed through a plurality of different zones or barrels and thus heated to a plurality of temperatures. For example, heated to a plurality of temperatures between 50°C and 160°C. In some examples, the first temperature in the extrusion apparatus (i.e. in a first zone or barrel) may be 50°C, the second temperature may be 60°C, the third temperature may be 110°C, the fourth temperature may be 130°C, the fifth temperature may be 140°C, the sixth temperature may be 150°C, the seventh temperature may be 160°C, and the eight temperature may be 130°C. In some examples, different zones may have the same temperature as a previous zone, for example a first zone at a first temperature, a second zone at a third temperature, a fourth zone at the second temperature and fifth zone at a third or the first temperature.

[0127] In some examples, the die temperature where the texturized bone base material exits the extrusion apparatus may be about 60°C.

[0128] In some examples, the screw speed of a twin screw extruder may be around 60 to 600 rpm. In some examples, the screw speed of a twin screw extruder may be around30 Hz. In some examples, the screw diameter may be about 50mm.

[0129] The low levels of oils or fat in the bone base material helps prevent slippage of the screws when texturizing the bone base material and allows for more regular formation of anisotropic fibrous material. The generation of anisotropic structure in high-protein extrudedproducts (such as the bone analogues described herein) can be accomplished by adjusting one or more parameters such as shear, heat, moisture, pressure differentials, and / or by rapid cooling. These parameters may contribute to distinctive directional and varied texture associated with anisotropic products. By aligning protein structures and establishing moisture and pressure gradients during extrusion, a structured matrix with directional properties may be formed, which is further emphasized by cooling and solidification post-extrusion. This textural anisotropy not only sets extruded high-protein products such as the bone analogues described herein) apart but also highlights their novelty and significance, especially in contexts where texture plays a pivotal role in product functionality and consumer perception.

[0130] The moisture of the bone base material during the extrusion may be from about 40 to about 70%. The moisture of the bone base material may vary throughout the extrusion process in the range of about 40 to 70%.

[0131] As detailed above, pressure may be applied to the bone base material. In some examples, the pressure may be a differential pressure that varies throughout the process. For example, at the start of the process, the bone base material may be at ambient pressure. For example, around 100 kPa (i.e. 1 bar). In some examples, while within the plurality of different zones or barrels, as described above, the bone base material may be at a pressure of about 2000 kPa to about 10,000 kPa (i.e. 20 to 100 bar). In some examples, while within the plurality of different zones or barrels, as described above, the bone base material may be at a pressure of about 4000 kPa to about 10,000 kPa (i.e. 40 to 100 bar). In some examples, the bone base material is then transferred to a cooling die. The pressure of the bone base material in the cooling die may be greater than ambient pressure. For example, about 101 , 102, 103, 104, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150 kPa. By using a slightly higher pressure than ambient pressure in the cooling die before exiting the extruder, the textured vegetable protein may not expand and / or has a firm texture, allowing for the textured vegetable protein to be recovered from the extruder as a solid piece (see Figure 4).

[0132] In some examples, heat and pressure may be applied by the use of a shear cell apparatus. Shear cell apparatuses and methods are well known in the art. In one example, a shear cell apparatus may include a first portion defining a first surface and a second portion defining a second surface, wherein the first portion and / or the second portion is rotatable. The bone base material comprising the non-animal derived protein is positioned in the space between the first and second surfaces and one or both of the first and second portions is rotated so as to apply shear forces to the bone base material between the first and second surfaces. The first and / or second portions are heated to apply heat to the bone base material.

[0133] In other examples, heat and pressure may be applied by use of an autoclave. If such a method is used to apply heat and pressure, further processing of the bone base material in order to texturize (i.e. form fibres in) the non-animal derived protein.

[0134] In some examples, the gel formed by the heat and pressure applied to the bone base material is crosslinked. For example, by using crosslinking agents as described herein. For example by the use of transglutaminase.

[0135] Optionally, once the bone base material has been formed into a gel having an anisotropic fibrous structure, the gel may be shaped. For example, shaped to have an appearance similar to an animal bone. Shaping may be done by cutting or manipulating the gel. For example, by rotating, twisting and / or bending the gel so as to form a shape that is mimetic of an animal bone. For example, if the bone analogue is to mimic a rib bone, the gel may be bent or twisted to provide an inclination of about 10%. The size of the bone analogue may be any suitable size. For example, the bone analogue may be made to have the dimensions of 6 cm (length) x 1 cm (width) x 0.5 cm (height) or 6 cm (length) x 2 cm (width) x 0.5 cm (height).

[0136] After the optional shaping or after texturization, the texturized non-animal derived protein (e.g., gel) is then dried. In some examples, the method includes a single continuous drying step. The single continuous drying step may be carried out at a temperature and time period that are configured to lead to formation of a bone analogue that has a hardness of at least 500N. For example, at least 500N, 600N, 700N, 800N, 900N or 1000N.

[0137] The temperature of the single continuous drying step may be at most 100°C. In some examples, the temperature may be less than 100°C. In some examples, the temperature may be from 100°C to 50°C. In some examples, the temperature may be from 100 to about 70°C. In some examples, the temperature of the single continuous drying step may be at most 100°C. In some examples, the temperature may be from 100°C to 50°C. In some examples, the temperature may be from 90 to about 80°C. For example, the temperature may be about 100, 99, 98, 97, 96, 95, 94, 93, 92, 91 , 90, 89, 88, 87, 86, 85, 84, 83, 82, 81 , 80, 79, 78, 77, 76, 75, 74, 73, 72, 71 , 70, 69, 68, 67, 66, 65, 64, 63, 62, 61 , 60, 59, 58, 57, 56, 55, 54, 53, 52, 51 , 50, 49, 48, 47, 46, 45, 44, 43, 42, 41 , 40, 39, 38, 37, 36, 35, 34, 33, 32, 31 , 30, 29, 28, 27, 26, 25, 24, 23, 22, 21 , 20, 19, 18, 17, 16, 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1°C. In some examples, the temperature may be about 85°C.

[0138] The time period of the single continuous drying step may be determined based on the hardness or moisture content of the bone analogue at various time points. It will be understood that the time to provide a bone analogue as described depends on the drying temperature. Higher temperatures will require shorter drying times. In some examples, thetime period of the single continuous drying step is sufficient to provide a bone analogue having a hardness of at least 500N. In some examples, the time period of the single continuous drying step is sufficient to provide a bone analogue with a moisture content of at most 15%. In some examples, the time period of the single continuous drying step is at least about 30 minutes. In some examples, the time period of the single continuous drying step is at most about 24 hours. In some examples, the time period of the single continuous drying step is from about 0.5 hours to about 24 hours. In some examples, the time period of the single continuous drying step is from about 12 hours to about 24 hours. For example, the time period of the single continuous drying step may be about 0.5, 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, or 24 hours. In some examples, the time period of the single continuous drying step is about 12 hours. In some examples, the time period of the single continuous drying step is about 12 hours and the temperature is about 85°C.

[0139] In some examples, the bone base material may be dried to form a pre-bone analogue using a first drying step carried out at a first temperature and first time period. The bone analogue may be dried for a time to provide a pre-bone analogue having a hardness of less than 500N. The bone analogue may be dried for a time to provide a pre-bone analogue having a hardness of about 300N. That is to say that the bone base material is dried to a point that provides a pre-bone analogue that has a hardness less than the bone analogue.

[0140] In some examples, the bone base material may be dried to provide a pre-bone analogue that has a moisture content less than the bone base material but greater than the bone analogue. For example, the bone base material may be dried at a temperature and time period to evaporate at most 86% of the plasticizer in the bone base material.

[0141] The first drying temperature may be at least 100°C. In some examples, the drying may be at a first temperature of from 100°C to about 130°C. For example, 100, 101 , 102, 103, 104, 105, 106, 107, 108, 109, 110, 111 , 112, 113, 114, 115, 116, 117, 118, 119, 120, 121 , 122, 123, 124, 125, 126, 127, 128, 129, 130°C. In some examples, the first drying temperature does not exceed 130°C. Temperatures higher than this may lead to burning or charring of the pre-bone analogue.

[0142] The time period of the first drying step may be determined based on the hardness or moisture content of the pre-bone analogue at various time points. It will be understood that the time to provide a pre-bone analogue depends on the drying temperature. Higher temperatures will require shorter drying times. In some examples, the time period of the first drying step is at least one hour. In some examples, the time period of the first drying step (i.e. the first time period) is from 1 hour to 5 hours. For example, 1 , 2, 3, 4, or 5 hours.

[0143] The method then may include a second drying step. The second drying step is carried out at a temperature and for a time that provides a bone analogue that has a moisture of at most 15%. For example, the pre-bone analogue may be dried for a time period and at a temperature sufficient to reduce the moisture content to 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13 or 14%.

[0144] In some examples, the second drying step may be carried out for a time period and at a temperature sufficient to provide a bone analogue having a hardness greater than the pre-bone analogue. In some examples, the second drying step may be carried out for a time period and at a temperature sufficient to provide a bone analogue having a hardness of more than 300N. In some examples, the second drying step may be carried out for a time period and at a temperature sufficient to provide a bone analogue having a hardness of more than 500N. In some examples, the second drying step may be carried out for a time period and at a temperature sufficient to provide a bone analogue having a hardness of at least 500N.

[0145] The second drying step may be carried out at a temperature of less than 100°C or at least 100°C. The second drying temperature (i.e. second temperature) may be selected based on the desired appearance of the final bone analogue. A second drying temperature of less than 100°C may provide a bone analogue that has a lighter colour. A second drying temperature of at least 100°C may provide a bone analogue that has a darker colour. In some examples, the second drying temperature is from 100 to 130°C.

[0146] It will be understood that the time period for the second drying step (second time period) may be altered depending on the second drying temperature used. When the second drying temperature is less than 100°C the second drying time period may be at least 5 hours. In some examples, when the second drying temperature is less than 100°C the second drying temperature may be from 5 to 12 hours. In some examples, when the second drying temperature is less than 100°C the second drying temperature may be from 5 to 10 hours. For example, 5, 6, 7, 8, 9, 10, 11 or 12 hours.

[0147] In some examples, when the second drying temperature is at least 100°C the second drying temperature may be at most 10 hours. In some examples, when the second drying temperature is at least 100°C the second drying temperature may be at most 6 hours. In some examples, when the second drying temperature is at least 100°C the second drying temperature may be from 2 to 10 hours. In some examples, when the second drying temperature is at least 100°C the second drying temperature may be from 2 to 6 hours. For example, 2, 3, 4, 5, 6, 7, 8, 9 or 10 hours.

[0148] After the single continuous drying step or second drying step, the bone analogue may be cooled. For example, the bone analogue may be cooled to ambient or room temperature.This may be done by any suitable method. For example, the bone analogue may be maintained at a desired temperature (e.g. ambient or room temperature) until the temperature of the bone analogue has reduced. For example, reduced to the same temperature as the desired temperature. For example, the bone analogue may be maintained at ambient or room temperature until the bone analogue reaches a temperature substantially equal to or equal to ambient or room temperature. In some examples, the bone analogue may be cooled till a maximum hardness has been reached.

[0149] The method may also optionally include applying a film onto at least a part of the outer surface of the pre-bone analogue or bone analogue. For example, a film may be applied after the first drying step or may be applied after the second drying step. The film may be a cartilage analogue as described herein.

[0150] In some examples, the film includes at least one hydrocolloid and / or gum as described herein. For example, the film may include one or more of guar gum, locust bean gum, carob bean gum, tara gum, acacia gum, xanthum gum, carrageenan, lecithin gum, gellan gum, sodium alginate, carrageenan, konjac glucomannan, citrus fibre, Puratein G®, methylcellulose, and / or hydroxypropylmethyl cellulose or similar. In some examples, the film includes gellan gum. In some examples, the film includes highly acylated gellan gum.

[0151] The film may be deposited onto at least a part of the outer surface by any suitable method, such as spraying, painting or immersing the pre-bone analogue or bone analogue in a solution including the components of the film. Once applied to the surface the film may be dried.

[0152] In some examples, the method includes preparing the film solution by dissolving the at least one hydrocolloid and / or gum in a solvent. In some examples, the solvent is water.

[0153] In some examples, the film includes 0.8% w / w highly acylated gellan gum. The method may include heating the at least one hydrocolloid and / or gum in the solvent to a temperature sufficient to dissolve the at least one hydrocolloid and / or gum. For example, to a temperature of about 85°C. Before depositing the solution, it may be cooled so as to increase the viscosity of the solution. For example, the solution may be cooled to about 50°C.

[0154] Once cooled, the solution is applied to the pre-bone analogue or bone analogue. The film is then formed by drying the film in situ on the pre-bone or bone analogue. For example, at a temperature of around 60 to 70°C. In the case of the film being applied to a pre-bone analogue the pre-bone analogue and solution deposited thereon may be dried with an additional drying step at 60 to 70°C for at least 2 hours before the second drying step, or the film may be dried during the second drying step. In the case of the film being applied to aready formed bone analogue (i.e. after the second drying step), the film may be dried onto the outer surface by a further drying step at 60 to 70°C for at least 2 hours.

[0155] In some examples, the first drying step may lead to the formation of an internal hollow structure in the pre-bone analogue. The internal hollow structure may be filled with a substance such as a bone marrow analogue as described herein.

[0156] The drying steps and heating may be carried out using any suitable methods. For example, heating may be carried out in a convection oven or other heating apparatus.Uses of Bone analogue and Meat analogues

[0157] The bone analogues described herein may be used for a number of different purposes. In particular, the bone analogues may be used in meat analogues to provide a consumer with a more realistic meat analogue. The bone analogues may also provide a consumer with a means to more easily handle and eat a meat analogue by providing a solid entity which can be gripped by the consumer.

[0158] As such, there is provided the use of a bone analogue as described herein as a bone mimetic in a meat analogue. There is also provided the use of a bone analogue as described herein for improving the appearance of a meat analogue.

[0159] As such, there is also provided a method of producing a meat analogue including a bone analogue as described herein.

[0160] A meat analogue refers to a food product that is not produced by the slaughter of an animal but has structure, texture, aesthetic qualities, and / or other properties comparable or similar to those of slaughtered animal meat, such as livestock (e.g., beef, pork), game (e.g., venison), poultry (e.g., chicken, turkey, duck), and / or fish or seafood. The term refers to uncooked, cooking, and cooked meat-like food products. Seafood refers to the marine and freshwater species in Phylum Arthropoda, Class Malacostraca, Orders Decapoda and Euphausiacea (e.g. shrimp, crayfish, lobsters and crabs); Phylum Mollusca, Classes Bivalvia, Gastropoda and Cephalopoda (e.g. shellfish) Phylum Echinodermata, Classes Echinoidea and Holothuroidea (e.g. sea urchins and sea cucumbers); Phylum Chordata, Class Actinopterygii, Orders Pleuronectiformes, Perciformes, Scorpaeniformes, Gadiformes, Anguilliformes (e.g. pelagic fish, demersal fish and reef fish).

[0161] In some examples, a meat analogue may be a plant-based meat analogue. The meat analogue may be made from a base material that includes a non-animal derived protein; the non-animal derived protein may comprise a plant protein, such as a vegetable protein, in particular soy protein. In other examples, the non-animal derived protein may additionally or alternatively comprise a fungal protein, a protein extracted from a microorganism, or arecombinantly produced protein, for example, a microbially produced recombinant protein such as beef myoglobin. In examples, the non-animal derived protein may comprise two or more different non-animal derived proteins. The non-animal derived protein may be in pure form of protein isolate, or a protein concentrate. In other examples, the protein may be a defatted meal with a high protein content, such as soybean meal, providing a protein content of greater than about 55%.

[0162] Meat analogues as used herein also refers to meat analogues formed from cultured cells. Cultured cell based meat analogues may also be known as cultured meat, in vitro meat, cellular agriculture products, or artificial meat. Such products are formed by in vitro culturing of non-human animal cells (for example, non-human animal myocytes) to form a structure that resembles cuts of meat obtained from a farmed animal. For example, see "Ng, Ee Theng, et al. "Cultured meat-a patentometric analysis." Critical Reviews in Food Science and Nutrition (2021): 1-11." and the references included therein.

[0163] The base material may additionally comprise water. For example, the base material may comprise up to about 90% water by weight, for example, up to about 80% water by weight, for example, about 70%-75% water by weight. In some examples, the base material may additionally comprise a fibre, for example, pectin or cellulose fibre. In some examples, the base material may additionally comprise a carbohydrate, for example, starch. Advantageously, addition of a fibre or carbohydrate may help with formation of fibres during a texturization process, such as high moisture extrusion. In examples, the base material may additionally comprise an oil, such as a vegetarian oil.

[0164] The base material may be processed, in particular texturized. For example, to create a fibrous protein texturate that may be referred to as a fibrous muscle tissue analogue. For example, by applying pressure and / or heat to denature the non-animal derived protein and applying shear forces to form fibres. The fibres may be at least partially separated to better mimic the texture of meat and improve the cooking and eating characteristics. Moreover, separation of the fibres also provides advantages for marinating and other processes that may be performed to produce the meat analogue, as described below. The fibres may be generally aligned. In some examples, the fibres may be substantially parallel, for example, parallel. The fibrous muscle tissue analogue may be a generally planar slab or sheet having two major surfaces and side surfaces, and the fibres of the fibrous muscle tissue analogue extend in a direction between the two major surfaces.

[0165] The base material may additionally comprise one or more of a fibre, such as a plant fibre. For example pectin or cellulose fibre, a carbohydrate, for example starch, a salt, for example sodium chloride or calcium chloride and / or an oil, such as a vegetable oil.Advantageously, addition of a fibre or carbohydrate may help with formation of fibres during the texturization process.

[0166] In some examples, the protein texturate may be a simple textured protein that does not include fibres, for example, tofu or soy-free textured proteins such as gluten based textured protein (e.g., seitan), lentil based textured protein, or bean based textured protein.

[0167] The base material may additionally comprise a hydrocolloid, for example, sodium alginate, xanthan gum, carrageenan, gellan gum, konjac glucomannan or similar. Advantageously, providing a hydrocolloid can help with fibre separation for improving the texture of the meat analogue.

[0168] In some examples, the base material may additionally comprise a salt, for example, sodium chloride or calcium chloride.

[0169] The protein texturate may also be marinated. For example, by applying, for example, spraying or brushing, a marinade on the protein texturate. The marinade may comprise one or more of a colouring, a flavouring, a flavour precursor, a preservative, a spice, and / or an oil. In some examples, particularly for red meat analogues, the colouring may comprise a beetroot-based thermolabile colouring. In some examples, the flavourings may comprise a meat flavouring. In some examples, the flavour precursors may comprise one or more of an amino acid, a reducing sugar (e.g., ribose), and a vitamin, for example, vitamin B12. The flavour precursors may be configured to transform or react during cooking of the meat analogue to release flavours or aromas like those of cooked animal meat. In some examples, the marinade may comprise a browning precursor such as lysin, and polyphenols (e.g., from apple extract). In some examples, the flavour precursor may additionally act as a browning precursor, for example, an amino acid or reducing sugar.

[0170] The meat analogues described herein may include one or more additional agents such as at least one Maillard reaction precursor, at least one mineral (such as such as an iron in the form of Fe-lactate, Fe-gluconate); at least one vitamin (such as ascorbic acid, and thiamine); at least one haem-containing protein (such as haemoglobin, myoglobin and leghaemoglobin); and / or at least one unsaturated fatty acid (for example palmitoleic acid, oleic acid, linoleic acid, linolenic acid, DHA and EPA).

[0171] A "flavouring agent" refers to a compound or salt or solvate thereof, that can be acceptably ingested, that induces a taste and / or smell in an animal or human. The flavouring agent may be natural, semi-synthetic or synthetic.

[0172] The flavouring agent may be a water soluble flavouring agent. The flavouring agent may be an oil soluble flavouring agent. The flavouring agent may be commercially availableflavouring agent and may be selected depending on the use of the composition. Examples of flavouring agents include acids, amino acids, sugars, reducing sugars (e.g., ribose), vitamins, and minerals. Flavouring agents may also include microorganisms and fungi, such as recombinant microorganisms and yeast.

[0173] For example, the flavouring agent may be selected to mimic the taste and / or aroma of animal-derived meat when the composition is for use in a meat analogue such as meat flavouring agents.

[0174] In some examples, the flavouring agent may be a sweet flavouring agent such as a sugar, or a compound that activates a T1 R2 / T1 R3 receptor in vitro.

[0175] Colouring agents may be any suitable colouring that is safe for consumption by animals and humans. Examples of suitable colouring agents include titanium dioxide, calcium phosphate or calcium carbonate.

[0176] Examples of spices and seasonings include salts (such as sodium or calcium chloride), pepper and / or garlic. The amounts and types of spices and seasonings added may be determined by the type of meat analogue intended.

[0177] In some examples, the meat analogue may include a binder. Binder may be applied after marinating the fibrous muscle tissue analogue, or in some examples, the binder may be applied before marinating the fibrous muscle tissue analogue. Binders include water-soluble or water-dispersible, non-proteinaceous film-formers such as synthetic and / or the natural polysaccharide film formers, proteinaceous film-formers, and mixtures thereof. Common binders used in plant-based meat include soy protein isolate, methylcellulose, carrageenan, and modified starches. Examples of binders include those described in EP0031622A1 and US4125630A.

[0178] The binder may be sprayed or brushed onto the fibrous muscle tissue analogue. In other examples, a binder bath may be provided, similar to the marinade bath, and the fibrous muscle tissue analogue may be passed through the binder bath to submerge the fibrous muscle tissue analogue in the binder.

[0179] In some examples, the binder is a fluid (in particular a liquid) that is sprayed or brushed onto the meat analogue, for example, onto the fibrous muscle tissue analogue. In other examples, a binder bath may be provided, similar to the marinade bath, and the fibrous muscle tissue analogue may be passed through the binder bath to submerge the fibrous muscle tissue analogue in the binder. 1

[0180] In some examples, the binder may include one or more of a hydrocolloid, a protein, or a carbohydrate. In some examples, the binder may comprise the same hydrocolloid, protein, or carbohydrate as provided in the base material.

[0181] In various examples, the binder may comprise a cold-set binder, for example, a sodium alginate solution or a salt mixture solution or BDF Binder 12.0. A cold-set binder can be set by storing the meat analogue at a cold temperature, for example, in a refrigerator at about 4°C, for a period of time to set the cold-set binder. The cold-set binder may be set by cooling the cold-set binder to below about 10°C, for example, to between about 2°C and about 8°C.

[0182] In other examples, the binder may comprise a heat-set binder, for example, a protein that denatures when heated. For example, the heat-set binder may comprise a soy protein, egg protein, potato protein, rubisco or a hydrocolloid, such as methyl cellulose. In examples where a heat-set binder is used, the meat analogue is heated to set the heat-set binder.

[0183] In some examples, the binder may alternatively comprise an enzymatic binder such as a transglutaminase enzyme. An enzymatic binder may form enzymatic crosslinking during heat setting. In a particular example, heat setting with an enzymatic binder (e.g., transglutaminase) may comprise a binding phase and a denaturing phase. During the binding phase, the enzymatic binder is activated and acts to bind the fibres together, and in the denaturing phase, the enzymatic binder denatures, leaving an inactive enzyme.

[0184] In other examples, the binder may additionally or alternatively comprise a konjac glucomannan that may provide a fatty mouthfeel for the meat analogue. Optionally, xanthan gum may be included in the binder to impart additional fatty mouthfeel.

[0185] In some examples, the binder comprises carrageenan. In some examples, the binder comprises methylcellulose. For example, the binder may comprise 10% carrageenan.

[0186] The binder acts to re-join the separated fibres of the fibrous muscle tissue analogue in a manner akin to intramuscular fat and extracellular matrix in animal meat. During cooking of the meat analogue, the binder may loosen or soften to mimic the cooking process of animal meat. The binder also acts to seal in the marinade to ensure that the marinade does not bleed out of the meat analogue during storage and transport before and during cooking.

[0187] The binder may comprise one or more of a hydrocolloid, a protein, or a carbohydrate. The binder may comprise the same hydrocolloid, protein, or carbohydrate as provided in the base material. The binder may comprise a mixture of a hydrocolloid and a protein isolate in the water phase. The binder may comprise more than 90% water by weight, for example 94% water by weight.

[0188] The binder may comprise a fat analogue. Advantageously, providing the binder with a fat analogue provides for a fat marbling on the meat analogue. In particular, the binder and fat analogue will penetrate and set in spaces between and around the separated fibres in the fibrous muscle tissue analogue, allowing the fat analogue to mimic intramuscular fat of animal meat. Addition of a fat analogue to the fibrous muscle tissue analogue is particularly advantageous for fatty meat analogues such as beef, mutton, lamb and pork meat analogues.

[0189] The meat analogues described herein may also include a fat analogue as described herein that is separate from the binder. In some examples the meat analogues may comprise a fat analogue such as plant oils and oil gels formed from hydrophobic polymers, such as those described in Dreher, Johannes, et al. "Formation and characterisation of plant-based emulsified and crosslinked fat crystal networks to mimic animal fat tissue". Journal of food science 85.2 (2020): 421-431. and Patel A.R., Nicholson R.A., Marangoni A.G. 2020. Applications of fat mimetics for the replacement of saturated and hydrogenated fat in food products. Current Opinion in Food Science, 33: 61-68. In addition, fat analogues include those described in W02014110539A1 which describes an adipose tissue analogue including one or more isolated plant proteins, and one or more plant or algal derived oils as well as those described in W02008047012 which describes a fat analogue that is a mixture of semi-solid palm oil representing and at least one vegetable oil which is fluid at ambient temperature.

[0190] The fat analogues described herein may comprise an oleogel (for example ethylcellulose or monoglyceride stabilised oleogel), an oil in water hydrocolloid stabilised emulsion, an oil in water emulsion, a water in oil emulsion, or a white coloured water-based hydrocolloid. The oil may comprise a plant oil, for example, sunflower oil, canola oil, or coconut oil. In examples, the fat analogue may additionally or alternatively comprise Maillard reaction precursors as described herein.

[0191] In some examples, the meat analogue is vegetarian. In some examples, the meat analogue is vegan. In some examples, all of the components of the meat analogue described herein may be vegetarian. That is to say, the components are not made from or with the aid of products or components derived from animals that have died, have been slaughtered, or animals that die as a result of being eaten. In some examples, the meat analogue described herein may be vegan. That is to say; the components are not sourced from or derived from an animal or animal product. Food products that are "vegan" are free of any animal products or animal by-products.

[0192] In some examples, the fibrous muscle tissue analogue may be shaped, for example, by placing the fibrous muscle tissue analogue in mould and setting the binder. The binder maybe set by heating or cooling the mould and the fibrous muscle tissue analogue within. The binder may help to retain the shape of the fibrous muscle tissue analogue.

[0193] In some examples, the fibrous muscle tissue analogue may be rolled before being shaped and before the binder is set. Rolling the fibrous muscle tissue analogue may increase the thickness of the fibrous muscle tissue analogue while keeping the fibres substantially parallel to each other.

[0194] Provided herein in one aspect is a method of producing a meat analogue comprising the steps of: forming a fibrous muscle tissue analogue by high-temperature texturization of a base material comprising a non-animal derived protein, the high-temperature texturization configured to cause denaturing of the non-animal derived protein and formation of substantially parallel fibres in the fibrous muscle tissue analogue; compressing the fibrous muscle tissue analogue to partially separate at least some of the fibres of the fibrous muscle tissue analogue; and applying a binder to the fibrous muscle tissue analogue after compressing the fibrous muscle tissue analogue, the binder being configured to set to bind the separated fibres together.

[0195] In another aspect there is provided a method producing a meat analogue comprising the steps of: forming a fibrous muscle tissue analogue by high-temperature texturization of a base material comprising a non-animal derived protein, the high-temperature texturization configured to cause denaturing of the non-animal derived protein and formation of substantially parallel fibres in the fibrous muscle tissue analogue; compressing the fibrous muscle tissue analogue to partially separate at least some of the fibres of the fibrous muscle tissue analogue; and applying a binder to the fibrous muscle tissue analogue after compressing the fibrous muscle tissue analogue, the binder being configured to set to bind the separated fibres together and subsequently applying a fat analogue.

[0196] Forming the fibrous muscle tissue analogue may comprise an extrusion process or a shear cell process. These processes apply pressure and heat to the base material to denature the non-animal derived protein and apply shear forces to form the protein into fibres.

[0197] During the high-temperature texturization process the base material may be heated to a “high-temperature” that is greater than the glass transition temperature of the basematerial. A “high temperature” may be about 90°C or higher, depending on the non-animal derived protein used in the base material.

[0198] Pressure may be applied mechanically, for example by a screw drive in an extrusion process. Alternatively or additionally, the pressure may be generated by expansion of the base material within a constrained volume, such as in a shear cell. Alternatively or additionally, the pressure may be generated by water vapour created by the heating. For example, water vapour pressure may generate about 3-4 bar of pressure. Heat may be applied directly, for example by heating a part of the texturization apparatus, and / or heat may be generated by applying pressure and / or shear forces to the base material due to friction.

[0199] The method includes partially separating the fibres of the fibrous muscle tissue analogue. In particular, at least some of the fibres of the fibrous muscle tissue analogue are partially separated. In examples, at least some of the fibres are only partially separated (i.e., not fully separated) such that the fibrous muscle tissue analogue retains an integral form, for example a slab-like form. The fibres may be partially separated by compressing the fibrous muscle tissue analogue to prise apart some of the fibres. Compression acts to separate some of the fibres due to the anisotropic nature of the fibrous muscle tissue analogue in which the fibres are substantially parallel to each other and so more readily separate from each other rather than break fibres. In some examples, the further fibrous muscle tissue analogue is bound such that the fibres of the fibrous muscle tissue analogue are substantially parallel to the fibres of the further fibrous muscle tissue analogue. By orientating the fibres of the first and second fibrous muscle tissue analogues in this way may provide a meat analogue with a texture, appearance and / or consistency similar to an animal-derived meat product.

[0200] Advantageously, partially separating at least some of the fibres of the fibrous muscle tissue analogue greatly improves the texture of the meat analogue. In particular, separation of the fibres makes the texture of the meat analogue more similar to that of animal meat. In addition, partial separation of the fibres of the fibrous muscle tissue analogue permits faster and more effective marinating to provide flavours and colouring, and also permits a fat analogue as described herein to be applied between the fibres to more closely mimic fat marbling in some animal meats, notably beef.

[0201] Optionally the method further includes marinating the fibrous muscle tissue analogue after the fibres have been partially separated, for example by separation rollers. Marinating may provide flavour and / or colouring for the meat analogue, particularly for producing a beef meat analogue, a lamb or mutton meat analogue, a chicken meat analogue, a fish meat analogue or a pork meat analogue.

[0202] In particular, the fibrous muscle tissue analogue may be marinated by applying, for example spraying or brushing, a marinade on the fibrous muscle tissue analogue. In examples, a powder flavour marinade may be applied by rubbing the powder flavour marinade onto the surfaces of the fibrous muscle tissue analogue and between at least some of the separated fibres. Alternatively, the fibrous meat tissue analogue may be marinated by passing the fibrous muscle tissue analogue through a marinade bath. The marinade may comprise one or more of a colouring, a flavouring, a flavour precursor, a preservative, a spice, and an oil. In some examples, the fibrous muscle tissue analogue has a high moisture content, for example, 50% or more moisture and the marinade may be applied to the fibrous muscle tissue analogue in an amount of up to about 10% of the weight of the fibrous muscle tissue analogue, for example, about 7% of the weight of the fibrous muscle tissue analogue. In examples, the marinade may comprise one or more binding components, for example, soluble binding components. In other examples, the fibrous muscle tissue analogue has a low moisture content, for example, below 50% or less moisture. For a low moisture fibrous muscle tissue analogue, the fibrous muscle tissue analogue may absorb from 50% of the weight of the fibrous muscle tissue analogue. In some examples a low moisture fibrous muscle tissue analogue, may absorb up to 4-times the dry weight of a low moisture fibrous muscle tissue analogue.

[0203] A fat analogue as described herein may be applied after applying the binder. That is to say a fat analogue may be applied to a meat analogue having a binder that comprises a fat analogue as described herein, or to a meat analogue that does not include a fat analogue. The fat analogue may be applied randomly to the fibrous muscle tissue analogue to provide a marbling effect. For example, the fat analogue may be applied to the fibrous muscle tissue analogue and then the fibrous muscle tissue analogue can be scraped or brushed to remove some of the fat analogue and leave remaining fat analogue in spaces between the separated fibres near the surface of the fibrous muscle tissue analogue.

[0204] The fat analogue may be configured to melt during cooking, for example, at between 50°C and 200°C. The fat analogue may oxidise during cooking such that degradation of triglycerides provides flavour.

[0205] The fat analogue may be applied before the binder has set. The fat analogue may be cold-set or heat-set and penetrate the fibrous muscle tissue analogue in the same manner as the binder, to provide marbling and intramuscular fat between the fibres.

[0206] In some examples, the meat analogue may include a further analogue. The further analogue may comprise a second fibrous muscle tissue analogue the same or similar to the fibrous muscle tissue analogue described above, and optionally further fibrous muscle tissueanalogues. In some examples, the further analogue may comprise a fat tissue analogue. The fat tissue analogue may comprise the same fat analogue as set out above, that can be bound to the fibrous muscle tissue analogue to provide a fat tissue layer on the meat analogue. In this way, a meat analogue can be assembled from a fibrous muscle tissue analogue and at least one further analogue, for example, a further fibrous muscle tissue analogue and / or a fat tissue analogue.

[0207] Binding the further analogue to the fibrous muscle tissue analogue may comprise using a binder as described above, which may be set to bind the further analogue to the fibrous muscle tissue analogue. A press may be used to facilitate assembly of the fibrous muscle tissue analogue and the further analogue. The combined fibrous muscle tissue analogue and further analogue may be added to a mould, as described above, and the binder may be set within the mould so as to hold the combined analogues in the desired shape. In examples, a plurality of fibrous muscle tissue analogues may be bound to each other with binder and optionally a fat tissue analogue therebetween to assemble a thicker meat analogue.

[0208] The fat analogue applied subsequently to the binder may be in the form of a fat tissue analogue.

[0209] In some examples, the fibrous muscle tissue analogue is bound to a fat tissue analogue. The fat tissue analogue may be a homogenous fat analogue that can be bound to a fibrous muscle tissue analogue to provide a fat tissue layer on the meat analogue.

[0210] The fat tissue analogue for example, may be applied to one side of a meat analogue, or between first and second fibrous muscle tissue analogues, analogous to intermuscular fat or subcutaneous fat layer.

[0211] A bone analogue as described herein may be disposed between a first fibrous muscle tissue analogue and a second analogue. For example between a first fibrous muscle tissue analogue and a second fibrous muscle tissue analogue or a fat tissue analogue.

[0212] The first fibrous muscle tissue analogue may first have a binder applied to a surface onto which the bone analogue is to contact. In some examples, the binder may be the same as the binder used to form the fibrous muscle tissue analogue or may be a different binder. In some examples, the binder comprises carrageenan. For example, comprises about 10% carrageenan in water.

[0213] A binder is also applied to the outer surface of the bone analogue and the bone analogue with binder thereon is deposited onto the surface of the fibrous muscle tissue analogue having the binder thereon. In some examples, the binder comprises an enzymatic binder such as a transglutaminase enzyme. An enzymatic binder may form enzymaticcrosslinking during heat setting. In a particular example, heat setting with an enzymatic binder (e.g., transglutaminase) may comprise a binding phase and a denaturing phase. During the binding phase, the enzymatic binder is activated and acts to bind the bone analogue and fibrous muscle tissue analogue, and in the denaturing phase, the enzymatic binder denatures, leaving an inactive enzyme. The employment of an enzyme may be advantageous for facilitating the binding of partially separated fibres to each other and the adhesion of partially separated fibres of the fibrous muscle tissue analogue to a bone analogue. The bone analogue includes a non-derived animal protein which can be affixed to the separated fibres of the fibrous muscle tissue analogue through the application of, for example, transglutaminase.

[0214] The use of enzymatic binders may help improve the organoleptic properties of a meat analogue, including a bone analogue as described herein. Such implementation enhances consumer satisfaction by providing a final product, such as a ribs analogue, that securely adheres to the bone. Consequently, this may provide consumers with a more realistic eating experience akin to consuming ribs directly from the bone, thereby improving overall product appeal.

[0215] A second analogue, such as a further fibrous muscle tissue analogue or fat tissue analogue with binder applied to at least one surface which contacts the bone analogue is then applied on top of the bone analogue and first fibrous muscle tissue analogue. Thereby encasing the bone analogue between two analogues.

[0216] In some examples, the binding the second fibrous muscle tissue analogue to the fibrous muscle tissue analogue comprises orientating the second fibrous muscle tissue analogue such that the fibres of the second fibrous muscle tissue analogue are substantially non-parallel to the fibres of the fibrous muscle tissue analogue, for example substantially perpendicular.

[0217] In some examples, the method further comprises at least one step of setting the binder. For example, the binder may be set after the meat analogue comprising the bone analogue has been assembled or after each application of the binder. Methods of setting the binder depends on the binder used. For example, for a cold set binder, the meat analogue and bone analogue deposited thereon may be set by maintaining the partially assembled or assembled meat analogue comprising the bone analogue at room or ambient temperature or in a reduced temperature compared to ambient or room temperature. If the binder is a heat set binder the partially assembled or assembled meat analogue comprising the bone analogue may be maintained at a temperature greater than ambient or room temperature.

[0218] Such a method advantageously provides for producing a meat analogue having fibres oriented in different directions in different parts of the meat analogue, permitting a range of different meat analogues to be produced with unique appearances and textures.

[0219] In examples, the fibres of the second fibrous muscle tissue analogue are substantially parallel to the fibres of the first fibrous muscle tissue analogue.

[0220] Accordingly, the size of the meat analogue can be increased by binding together two or more fibrous muscle tissue analogues. In examples, layers of fat analogue may be provided between the fibrous muscle tissue analogue and the second fibrous muscle tissue analogue, analogous to intermuscular fat in the meat analogue along with a bone analogue between at least two tissue analogue layers.

[0221] As the bone analogues and meat analogues described herein are for use in food products which are to be ingested by an animal, such as by humans, all the components are preferably food grade and / or food safe. Food grade refers to components that meet government food safety standards and have been approved for animal and / or human consumption. For example see the Food Chemicals Codex (FCC). In examples, “food grade” refers to an agent or substance classified by the United States Food and Drug Administration (US FDA) as generally regarded as safe (GRAS). In some examples, “food grade” refers to an agent or substance that is a “feed material” or “food additive” as these terms are defined by the European Union. Food materials are defined, for example, by Regulations (EC) Nos 1831 / 2003, 767 / 2009, and 575 / 2011 of the European Parliament and of the Council. Food additives are defined, for example, by European Parliament and Council Directive 95 / 26 EC. As such, each of these agents or substances can be added directly to food or a feedstock, or used in food contact applications. Accordingly, their use is considered safe for animal and human consumption.

[0222] In addition one of the aims of the bones analogues described herein are to provide an alternative to animal derived fat tissue and / or cells. As such, all of the components of the food products described herein may be vegetarian. That is to say the components are not made from or with the aid of products or components derived from animals that have died, have been slaughtered, or animals that die as a result of being eaten. In some examples, one or more of the components may be vegan. That is to say the components are not sourced from or derived from an animal or animal product. As such, the components that are “vegan” are free of any animal products or animal by-products.

[0223] Unless defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. For example, Singleton and Sainsbury, Dictionary of Microbiology andMolecular Biology, 2d Ed., John Wiley and Sons, NY (1994); and Hale and Marham, The Harper Collins Dictionary of Biology, Harper Perennial, NY (1991) provide those of skill in the art with a general dictionary of many of the terms used in the invention. Although any methods and materials similar or equivalent to those described herein find use in the practice of the present invention, the preferred methods and materials are described herein. Accordingly, the terms defined immediately below are more fully described by reference to the Specification as a whole. Also, as used herein, the singular terms “a”, “an,” and “the” include the plural reference unless the context clearly indicates otherwise. Unless otherwise indicated, nucleic acids are written left to right in 5’ to 3’ orientation; amino acid sequences are written left to right in amino to carboxy orientation, respectively. It is to be understood that this invention is not limited to the particular methodology, protocols, and reagents described, as these may vary, depending upon the context they are used by those of skill in the art.

[0224] Aspects of the invention are demonstrated by the following non-limiting examples.EXAMPLESExample 1

[0225] The bone analogues are a way of substituting the real animal bones with a more sustainable option. This invention increases the appearance of plant-based products making them more appealing for the final consumer. The hyper-realistic bones provide a better eating experience to meat analogues as it will be more akin to the animal derived counterpart.

[0226] The plant-based bones are a really versatile product. They can be shaped with any desired form to mimic any animal-based bone found in nature. A twin-screw extruder or shear cell technology is used to create an anisotropic structure that allows for a higher hardness of the end product thanks to the fibres created.

[0227] To create an even more realistic eating experience when eating meat analogues containing bones analogues described herein, bones are further processed to obtain a cartilage-like structure around them and improve adhesiveness to plant-based meat analogues.

[0228] The method comprises the following steps: mixing a plant protein (either soy, pea, wheat, lupin... ) with water and possibly hydrocolloids, gums, fibres, flavourings to obtain a better texture and flavour. This mixture is then heated up with pressure to create a gel-like structure. It can also be crosslinked with enzymes to get to a firmer gel. A twin-screw extruder with minimum temperatures of 60°C and maximum temperatures of 160°C with a pressure of around 60 bars is used to obtain a fibrousmaterial. Alternatively, shear cell technology can also be used to create an anisotropic structure similar to the extrusion.

[0229] Once the protein gel / texturate is obtained, it is shaped and cut to obtain the final desired bone product (ribs, chicken legs, chicken wings, fish...). The protein gel / texturate can be reshaped modifying its three-dimensional shape to add special features to the bone, such as inclination, bending or twisting. The drying step is done in a convection oven. Temperatures higher than 100°C for a couple of hours allow water evaporation inside the bone which results in a puffed bone with a bigger thickness and a more rounded shape. This hollow inside allows the addition of a paste to recreate the bone marrow found in animal bones. After this time, bones are dried at temperatures lower than 100°C until the moisture content of the bone is minimum. This results in a hard and stiff product that can be used in meat analogues to recreate a better eating experience. Temperatures above 100°C result in a darker bone analogue, whereas temperatures lower than 100°C result in a lighter beige colour.

[0230] Bones may be coated with a solution of hydrocolloids or gums (such as methylcellulose, alginate, guar gum, locust bean gum...) to create a film around it with a cartilage-like structure. This film mimics the cartilage found around animal bones and will also help to stick the bone to the meat analogue. With this solution, the consumer will have the full eating experience as they will be able to hold the meat just by grabbing the bone analogue and biting around it.

[0231] The film is a solution of 0.8 % w / w of high acetylated gellan gum. The gellan gum is dissolved in water and stirred on a magnetic stirrer while heating to 85°C for complete dissolution. Then the solution is cooled down to 50°C, so it thickens, which helps the solution not to drip down from the bone. Bone analogue is dipped into the solution and placed on baking sheet paper and the bones are dried at 60-70°C. Drying time depends on the dryness level of the bone. If they have already been dried prior to making the film, 2 hours of drying the film is enough, otherwise drying time lengthens.

[0232] The last step to manufacture the final product is the assembling step. Shortly, a pulled fibrous material is bonded to the bones using a binder. The binder may be a paste composed of a mixture of 10 % carrageenan and water.

[0233] This binder is applied in between the different layers of partially separated fibres. Between two layers of textured plant protein, bones are placed with binder around them to ensure a better adhesiveness to the meat analogue. Later the final product is pasteurised so that it sets after cooling - then the product can be handled similar to raw or sous-vide cooked meat product. Pasteurisation that can prolong the shelf life. The processing temperatures need to be higher than the gelling temperature of the binding hydrocolloids (for instance 69°C forkappa carrageenan). The temperature can also be as high as 121 °C and the process can result in shelf stable product if the heating is done in the final packaging.

[0234] To create a fattier and more succulent product, a fat system can be included in the product. It can be a mixture of different saturated and unsaturated fats such as sunflower oil, olive oil, coconut butter, shea butter. This fat system would be applied in between the partially separated fibres to obtain a homogeneous mouthfeel.Example 2Production of bone analogue1 . Plant protein was mixed with the water in a 50-50% ratio.2. The mixture was placed in a twin-screw extruder.Settings of the extruder:Barrels temperature: 50, 60, 110, 130, 140, 150, 160, 150, 130 °CScrew speed: 60-600 rpmCooling die temperature: 60 °C3. The texturized gel was cut into the desired shape.4. The texturized and shaped gel was dried at a temperature higher than 100°C for 2h or until the water inside the gel has partially evaporated and a hollow structure is formed.5. The bone analogue was then further dried at a temperature lower than 100°C (around 70°C) for around 8h or until the moisture content was lower than 15%.

[0235] Examples of the bone analogues can be seen in figure 2A to D.Meat analogue protocol including bone analogue (for example ribs)1 . a layer of fibrous muscle tissue analogue was prepared.2. binder was applied on top of the first layer of muscle tissue analogue.(Repeat the procedure as many times as needed to reach the desired thickness).3. The bone analogues were coated with binder and placed them on top of the previous layers of muscle tissue analogue with partially separated fibres.4. The bone analogues were coated with one more layer binder and of muscle tissue analogue with partially separated fibres was placed on top.

[0236] Examples of the bone analogues in situ with a meat analogue can be seen in figure 3A to C.Example 3Additional Protocol for Production of Bone analogue1 . Plant protein was mixed with water in a 1 :1 ratio.2. The mixture was introduced into a twin extruder and extruded reaching maximum temperatures of 170°C.3. The resulting non-expanded protein gel was then cut to the desired size to mimic the pork ribs bones (95x20x10 mm).4. Bones were then placed on a tray and dried for 12 hours at 85°C in a convection oven.5. After drying, the bones were cooled down at room temperature until the maximum hardness was obtained.Application of Bone analogues1 . A layer of fibrous muscle tissue analogue was prepared.2. Binder was applied on top of the first layer of muscle tissue analogue, (repeat this process as many times as needed to achieve the desired thickness)3. The number of bone analogues desired (in this case 6) were placed in parallel on top of the fibrous muscle tissue, with the tips sticking out of the fibrous tissue analogue.4. The bone analogues were coated with at least one more layer of binder and another layer of partially separated fibres on top.5. Finally, the pork ribs analogue product is placed in a sous vide bath / convection oven to set the gel binder. As carrageenan was present in the binder, the temperature used was 85°C for 30 min.

[0237] The reader's attention is directed to all papers and documents which are filed concurrently with or previous to this specification in connection with this application and which are open to public inspection with this specification, and the contents of all such papers and documents are incorporated herein by reference.

[0238] All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and / or steps are mutually exclusive.

[0239] Each feature disclosed in this specification (including any accompanying claims, abstract and drawings), may be replaced by alternative features serving the same, eguivalent,or similar purpose, unless expressly stated otherwise. Thus, unless expressly stated otherwise, each feature disclosed is one example only of a generic series of equivalent or similar features.

[0240] The invention is not restricted to the details of any foregoing embodiments. The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.NUMBERED CLAUSES1. A vegetarian bone analogue for inclusion in a meat analogue, the meat analogue for human consumption, the vegetarian bone analogue comprising: a. a texturized non-animal derived protein; and b. a plasticizer; wherein the vegetarian bone analogue comprises an anisotropic structure.2. The vegetarian bone analogue of clause 1 , wherein the vegetarian bone analogue has a hardness of at least about 500 N.3. The vegetarian bone analogue of clauses 1 or 2, wherein the vegetarian bone analogue has a moisture content of at most about 15% w / w.4. The vegetarian bone analogue of any of clauses 1 to 3, wherein the anisotropic structure comprises an anisotropic fibrous structure.5. The vegetarian bone analogue of any of clauses 1 to 4, wherein the vegetarian bone analogue comprises at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent.6. The vegetarian bone analogue of any preceding claim, wherein the anisotropic fibrous structure comprises denatured non-animal derived protein.7. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises a shape that is mimetic of an animal bone.8. The vegetarian bone analogue of any preceding claim, wherein the texturized non- animal derived protein is crosslinked.9. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises a film disposed on at least a portion of the outer surface of the vegetarian bone analogue.10. The vegetarian bone analogue of clause 9, wherein the film comprises a cartilage analogue.The vegetarian bone analogue of clauses 9 or 10, wherein the film comprises at least one hydrocolloid and / or at least one gum. The vegetarian bone analogue of any of clauses 9 to 11 , wherein the film comprises gellan gum, optionally highly acetylated gellan gum. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises an internal hollow structure. The vegetarian bone analogue of clause 13, wherein the internal hollow structure comprises a vegetarian bone marrow analogue, optionally, wherein the bone marrow analogue comprises a vegetarian liquid composition for mimicking the flavour of bone marrow and a gelling agent. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue is vegan. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue has been pasteurized. A method of forming a vegetarian bone analogue, the method comprising: a. mixing a non-animal derived protein with a plasticiser to form a bone base material; b. heating and applying a pressure to the bone base material to form a gel and to texturize the non-animal derived protein; c. drying the gel at a first temperature and for a first time period sufficient to provide a pre-bone analogue having a hardness of less than 500N; and d. drying the pre-bone analogue at a second temperature and for a second time period sufficient to provide a vegetarian bone analogue having a hardness of at least 500N. The method of clause 17, wherein step (a) further comprises mixing at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent; with the non-animal derived protein and plasticiser. The method of clause 17 or 18, wherein a. the first temperature and first time period are sufficient to provide a pre-bone analogue with a moisture content of less than the bone base material and greater than the bone analogue; and / or b. the second temperature and second time period are sufficient to provide a bone analogue with a moisture content less than 15%. The method of any of clauses 17, 18 or 19 wherein step (b) comprises: a. using an extrusion process to heat and apply pressure to the bone base material;b. using a shear cell process to heat and apply pressure to the bone base material. optionally wherein the extrusion process comprises using a twin-screw extruder. The method of any of clauses 17 to 20, wherein: a. the non-animal derived protein and plasticizer are mixed at a 1 :1 ratio; b. the first temperature is at least 100°C; c. the first time period is of at least one hour; d. the second temperature is less than 100°C or at least 100°C; e. the second time period is at least 5 hours when the second temperature is less than 100°C or the second time period is at least 2 hours when the second temperature is at least 100°C; and / or f. the pressure is around 6x106Pa (60 bar). The method of any of clauses 17 to 21 , wherein the method further comprises shaping the gel after step (b) and before step (c) of clause 17. The method of any of clauses 17 to 22, wherein the method further comprises crosslinking the gel after step (b) and before step (c) of clause 17. The method of any of clauses 17 to 23, the method further comprises applying a film to at least a portion of the outer surface of the pre-bone analogue or the formed vegetarian bone analogue; optionally: wherein the film comprises a cartilage analogue; and / or wherein the film comprises gellan gum, optionally highly acetylated gellan gum. The method of clause 24, wherein applying the film comprises: a. depositing a solution comprising at least one hydrocolloid and / or gum onto the portion of the outer surface; and b. drying the deposited solution on the outer surface to form the film. The method of clauses 24 or 25, wherein applying the film further comprises preparing the solution, preparing comprising: a. dissolving the at least one hydrocolloid and / or gum in a solvent to form the solution; and b. cooling the solution to a temperature suitable for increasing the viscosity of the solution before depositing the solution. The method of any of clauses 17 to 26, wherein drying the gel at the first temperature and for the first time period provides a pre-bone analogue comprising an internal hollow structure and the method further comprises depositing a bone marrow analogue within the internal hollow structure of the pre-bone analogue or the formed vegetarian bone analogue. The method of any of clauses 17 to 27, wherein the vegetarian bone analogue comprises an anisotropic fibrous structure. A vegetarian bone analogue formed by the method of any of clauses 17 to 28. Use of a vegetarian bone analogue according to any one of clauses 1 to 16 and clause 29, as a bone mimetic in a meat analogue.A method of forming a meat analogue comprising a bone analogue, the method comprising: a. providing a first fibrous muscle tissue analogue; b. applying a binder to an upper surface of the first fibrous tissue analogue; c. applying a binder to a vegetarian bone analogue according to any of clauses 1 to 16 and clause 29; d. depositing the vegetarian bone analogue onto the upper surface; e. applying a second analogue on top of the bone analogue and the upper surface of the first fibrous muscle tissue analogue. The method of clause 31 , wherein the method further comprises setting the binder. The method of clause 32, wherein the second analogue comprises a second fibrous muscle tissue analogue and / or fat tissue analogue. The method of clause 31 or 32, wherein the second analogue comprises a binder on at least one surface of the second analogue, wherein the at least one surface contacts the vegetarian bone analogue. The method of any of clauses 31 to 34, wherein the first fibrous muscle tissue analogue comprises a fat analogue; optionally wherein the binder comprises a fat analogue. The method of any of clauses 31 to 35, wherein the first fibrous muscle tissue analogue and / or the second fibrous muscle tissue analogue comprise texturized non-animal derived protein, wherein at least some of the fibres of the fibrous muscle tissue analogue are partially separated; optionally comprising a marinade absorbed into the fibres; and / or comprising a or the binder interspersed between at least some of the fibres. The method of any of clauses 33 to 36, wherein: a. the fibres of the second fibrous muscle tissue analogue are substantially parallel to the fibres of the first fibrous muscle tissue analogue; or b. the fibres of the second fibrous muscle tissue analogue are substantially nonparallel to the fibres of the fibrous muscle tissue analogue. A meat analogue formed by a method according to any of clauses 31 to 37. A meat analogue comprising a vegetarian bone analogue according to any one of clauses 1 to 16 and clause 29.

Claims

CLAIMS1. A vegetarian bone analogue for inclusion in a meat analogue, the meat analogue for human consumption, the vegetarian bone analogue comprising: a. a texturized non-animal derived protein; and b. a plasticizer; wherein the vegetarian bone analogue comprises an anisotropic structure.

2. The vegetarian bone analogue of claim 1 , wherein the vegetarian bone analogue has a hardness of at least about 500 N.

3. The vegetarian bone analogue of claims 1 or 2, wherein the vegetarian bone analogue has a moisture content of at most about 15% w / w.

4. The vegetarian bone analogue of any of claims 1 to 3, wherein the anisotropic structure comprises an anisotropic fibrous structure.

5. The vegetarian bone analogue of any of claims 1 to 4, wherein the vegetarian bone analogue comprises at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent.

6. The vegetarian bone analogue of any preceding claim, wherein the anisotropic fibrous structure comprises denatured non-animal derived protein.

7. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises a shape that is mimetic of an animal bone.

8. The vegetarian bone analogue of any preceding claim, wherein the texturized non- animal derived protein is crosslinked.

9. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises a film disposed on at least a portion of the outer surface of the vegetarian bone analogue.

10. The vegetarian bone analogue of claim 9, wherein the film comprises a cartilage analogue.

11. The vegetarian bone analogue of claims 9 or 10, wherein the film comprises at least one hydrocolloid and / or at least one gum.

12. The vegetarian bone analogue of any of claims 9 to 11 , wherein the film comprises gellan gum, optionally highly acetylated gellan gum.

13. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue comprises an internal hollow structure.

14. The vegetarian bone analogue of claim 13, wherein the internal hollow structure comprises a vegetarian bone marrow analogue, optionally, wherein the bone marrow analogue comprises a vegetarian liquid composition for mimicking the flavour of bone marrow and a gelling agent.

15. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue is vegan.

16. The vegetarian bone analogue of any preceding claim, wherein the vegetarian bone analogue has been pasteurized.

17. A method of forming a vegetarian bone analogue, the method comprising: a. mixing a non-animal derived protein with a plasticiser to form a bone base material; b. heating and applying a pressure to the bone base material to form a gel and to texturize the non-animal derived protein; c. drying the gel at a temperature and for a time period sufficient to provide a vegetarian bone analogue having a hardness of at least 500N; and wherein the vegetarian bone analogue comprises an anisotropic structure.

18. The method of claim 17, wherein the temperature is less than 100°C; optionally about 85°C.

19. The method of claim 17 or 18 wherein time period is at least 0.5 hours.

20. The method of claim 17, wherein drying comprises: a. a first drying step comprising drying the gel at a first temperature and for a first time period sufficient to provide a pre-bone analogue having a hardness of less than 500N; and b. a second drying step comprising drying the pre-bone analogue at a second temperature and for a second time period sufficient to provide a vegetarian bone analogue having a hardness of at least 500N.

21. The method of any one of claims 17 to 20, wherein the method further comprises: e. cooling the bone analogue; optionally wherein cooling comprises maintaining the bone analogue at ambient temperature until the hardness of the bone analogue reaches a maximum value.

22. The method of any of claims 17 to 21 , wherein step (a) further comprises mixing at least one or more of: a. at least one hydrocolloid; b. at least one gum; c. at least one fibre; and / or d. at least one flavouring agent with the non-animal derived protein and plasticiser.

23. The method of any of claims 17 to 19, 21 and 22, wherein the temperature and time period are sufficient to provide a bone analogue with a moisture content less than 15%.

24. The method of any of claims 20 to 22, wherein a. the first temperature and first time period are sufficient to provide a pre-bone analogue with a moisture content of less than the bone base material and greater than the bone analogue; and / or the second temperature and second time period are sufficient to provide a bone analogue with a moisture content less than 15%.

25. The method of any of claims 20 to 24, wherein: a. the first temperature is at least 100°C; b. the first time period is at least one hour; c. the second temperature is less than 100°C or at least 100°C; and / ord. the second time period is at least 5 hours when the second temperature is less than 100°C or the second time period is at least 2 hours when the second temperature is at least 100°C.

26. The method of any of claims 17 to 25, wherein step (b) comprises: a. using an extrusion process to heat and apply pressure to the bone base material; optionally wherein the extrusion process comprises using a twin- screw extruder; or b. using a shear cell process to heat and apply pressure to the bone base material.

27. The method of any of claims 17 to 26, wherein: a. the non-animal derived protein and plasticizer are mixed at a 1:1 ratio; and / or b. the pressure is around 6x106Pa (60 bar).

28. The method of any of claims 17 to 27, wherein the method further comprises shaping the gel after step (b) and before step (c) of claim 17.

29. The method of any of claims 17 to 28, wherein the method further comprises crosslinking the gel after step (b) and before step (c) of claim 17.

30. The method of any of claims 17 to 29, the method further comprises applying a film to at least a portion of the outer surface of the pre-bone analogue or the formed vegetarian bone analogue; optionally: wherein the film comprises a cartilage analogue; and / or wherein the film comprises gellan gum, optionally highly acetylated gellan gum.

31. The method of claim 30, wherein applying the film comprises: a. depositing a solution comprising at least one hydrocolloid and / or gum onto the portion of the outer surface; and b. drying the deposited solution on the outer surface to form the film.

32. The method of claims 30 or 31 , wherein applying the film further comprises preparing the solution, preparing comprising: a. dissolving the at least one hydrocolloid and / or gum in a solvent to form the solution; andb. cooling the solution to a temperature suitable for increasing the viscosity of the solution before depositing the solution.

33. The method of any of claims 17 to 32, wherein drying the gel at the first temperature and for the first time period provides a pre-bone analogue comprising an internal hollow structure and the method further comprises depositing a bone marrow analogue within the internal hollow structure of the pre-bone analogue or the formed vegetarian bone analogue.

34. The method of any of claims 17 to 33, wherein the vegetarian bone analogue comprises an anisotropic fibrous structure.

35. A vegetarian bone analogue formed by the method of any of claims 17 to 34.

36. Use of a vegetarian bone analogue according to any one of claims 1 to 16 and claim 35, as a bone mimetic in a meat analogue.

37. A method of forming a meat analogue comprising a bone analogue, the method comprising: a. providing a first fibrous muscle tissue analogue; b. applying a binder to an upper surface of the first fibrous tissue analogue; c. applying a binder to a vegetarian bone analogue according to any of claims 1 to 16 and claim 35; d. depositing the vegetarian bone analogue onto the upper surface; e. applying a second analogue on top of the bone analogue and the upper surface of the first fibrous muscle tissue analogue.

38. The method of claim 37, wherein the method further comprises setting the binder.

39. The method of claim 38, wherein the second analogue comprises a second fibrous muscle tissue analogue and / or fat tissue analogue.

40. The method of any of claims 37 to 39, wherein the second analogue comprises a binder on at least one surface of the second analogue, wherein the at least one surface contacts the vegetarian bone analogue.41 . The method of any of claims 37 to 40, wherein the first fibrous muscle tissue analogue comprises a fat analogue; optionally wherein the binder comprises a fat analogue.

42. The method of any of claims 37 to 41 , wherein the first fibrous muscle tissue analogue and / or the second fibrous muscle tissue analogue comprise texturized non-animal derived protein, wherein at least some of the fibres of the fibrous muscle tissue analogue are partially separated; optionally comprising a marinade absorbed into the fibres; and / or comprising a or the binder interspersed between at least some of the fibres.

43. The method of any of claims 39 to 42, wherein: a. the fibres of the second fibrous muscle tissue analogue are substantially parallel to the fibres of the first fibrous muscle tissue analogue; or b. the fibres of the second fibrous muscle tissue analogue are substantially nonparallel to the fibres of the fibrous muscle tissue analogue.

44. A meat analogue formed by a method according to any of claims 37 to 43.

45. A meat analogue comprising a vegetarian bone analogue according to any one of claims 1 to 16 and claim 35.

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