Introduction

Overfishing is a major threat to marine fish stocks worldwide. The World Economic Forum estimated that 90% of the world’s stocks were either fully exploited, overexploited or depleted in 2018.

In terms of public health, there is also a growing awareness and concern about the accumulation of heavy metals and microplastics in fish. About 2% of the global population are thought to suffer from sea food allergies.

If consumers were encouraged to make the switch to fish analogues, this could help to address the sustainability and public health issues. One of the most popular types of fish is salmon which is enjoyed in many countries. Salmon analogue products do exist but they are generally of very low quality and lack the taste, texture, and nutrition of real salmon.

There is a clear need to provide consumers with salmon analogue products which address the sustainability and public health issues, and which more closely resemble the qualities of real salmon.

Summary of invention

The inventors have developed a method for gelation of fibers to form viscoelastic and translucent gels which mimic raw fishlike texture and appearance. Specific starches, flours, proteins, and salts are used to modulate the in-mouth creamy perception, color effect and thermal stability. The insertion of white connective tissue layers between orange or pink layers mimics the appearance of animal salmon and provides textural differentiation between the orange or pink layers and the white connective tissue layer. Embodiments of the invention

The invention relates to a method of preparing a fish analogue, said method comprising the steps of hydrating a mixture comprising a glucomannan source and a carrageenan source.

The invention relates to a method of preparing a fish analogue having a layered structure, said method comprising the steps of hydrating a first mixture comprising a glucomannan source, a starch source, a salt source, and a carrageenan source; and hydrating a second mixture comprising a glucomannan source, a salt source, and a whitening agent; and forming a layered structure which comprises a layer of the second mixture between two layers of the first mixture.

The invention further relates to a method of preparing a fish analogue having a layered structure, said method comprising the steps a. Hydrating a first mixture comprising a glucomannan source, a starch source, a salt source, a carrageenan source, and optionally one or more of a colorant, a plant based oil comprising omega 3 or omega 6, a flavor, and a pectin source, c. Hydrating a second mixture comprising a glucomannan source, a salt source, a whitening agent, and optionally a starch source, a carrageenan source, wherein the whitening agent is a protein, a fiber, an insoluble salt, or an oil-in-water emulsion; d. Forming a layered structure which comprises a layer of the second mixture between two layers of the first mixture; and wherein the first and second hydrated mixtures are heated before or after layering, for example at a minimum temperature of 75 °C.

In some embodiments, the heating step is before layering. In some embodiments, the heating step is after layering.

In some embodiments, the glucomannan source in the first mixture is a konjac flour, wherein the first mixture comprises up to 10 % konjac flour, and wherein said konjac flour comprises up to 5% glucomannan. In some embodiments, the starch source is derived from a cereal, tuber or legume source that is rich in starch, preferably with more than 50% starch.

In some embodiments, the starch source is a native potato starch, a tapioca starch, a potato flour, a tapioca flour, or their combination. In some embodiments, the starch source is a native potato starch or a tapioca starch. Typically, the first mixture comprises between 1 to 7wt% starch, preferably 2-5wt% starch, for example between 2 to 3 wt% potato starch or tapioca starch.

In some embodiments, both first and second mixture layers each comprise between 0.1 to 1 wt% carrageenan source, preferably a seaweed flour comprising carrageenan. In some embodiments, both first and second mixture layers each comprise between 0.5 - 5 wt% seaweed flour, preferably between 1 to 3 wt%. This provides improved flavor, texture and stability, for example a reduction in syneresis.

In some embodiments, both first and second mixture layers each comprise konjac glucomannan. In some embodiments, both first and second mixture layers each comprise seaweed flour. In some embodiments, both first and second mixture layers each comprise potato starch.

In some embodiments, sodium chloride is a salt source in the first mixture layer. In some embodiments, between 0.5 to 3 wt%, or about 1 .5 wt% sodium chloride is a salt source in the first mixture layer. In some embodiments, sodium chloride is a salt source in the second mixture layer. In some embodiments, between 0.5 to 3 wt%, or about 1.5 wt% sodium chloride is a salt source in the second mixture layer.

In some embodiments, the salt source comprises a combination of sodium chloride and calcium chloride in the first mixture layer. In some embodiments, the salt source comprises a combination of between 0.5 to 3 wt%, or about 1 .5 wt% sodium chloride and between 0.05 to 0.5 wt%, or about 0.2 wt% calcium chloride in the first mixture layer.

In some embodiments, the salt source comprises a combination of sodium chloride and calcium chloride in the second mixture layer. In some embodiments, the salt source comprises a combination of between 0.5 to 3 wt%, or about 1 .5 wt% sodium chloride and between 0.05 to 0.5 wt%, or about 0.2 wt% calcium chloride in the second mixture layer. In some embodiments, the salt source comprises a combination of sodium chloride, calcium chloride, and potassium carbonate in the first mixture layer. In some embodiments, the salt source comprises a combination of between 0.5 to 3 wt%, or about 1 .5 wt% sodium chloride and between 0.05 to 0.5 wt%, or about 0.2 wt% calcium chloride and between 0.05 to 0.3 wt%, or about 0.15 wt% potassium carbonate in the first mixture layer.

In some embodiments, the salt source comprises a combination of sodium chloride, calcium chloride, and potassium carbonate in the second mixture layer. In some embodiments, the salt source comprises a combination of between 0.5 to 3 wt%, or about 1 .5 wt% sodium chloride and between 0.05 to 0.5 wt%, or about 0.2 wt% calcium chloride and between 0.05 to 0.3 wt%, or about 0.15 wt% potassium carbonate in the second mixture layer.

In some embodiments, the first mixture layer comprises konjac glucomannan, seaweed flour, potato starch, and citrus fiber.

In some embodiments, the first and second mixtures comprise a pectin-rich fiber, for example citrus fiber at between 0.1 to 1 wt%, for example about 0.5 wt% citrus fiber. This improves the water holding capability and reduces syneresis during storage and improves the freeze-thaw stability of the final product.

In some embodiments, the first mixture and/or the second mixtures comprises alkali agent, for example sodium carbonate or potassium carbonate. This improves heat stability of the final product.

In some embodiments, the first mixture in step (a) is hydrated for at least 5 minutes, preferably for at least 10 minutes.

In some embodiments, the whitening agent comprises insoluble protein, fiber, or salt particles, or oil droplets in an oil-in-water emulsion, and wherein said particles are between 0.1 to 200 microns.

In some embodiments, the whitening agent is insoluble protein, for example rice protein. In some embodiments, the whitening agent is an insoluble fiber, for example pea fiber, bamboo fiber, wheat fiber, oat fiber, cellulose powder, or mixtures thereof, preferably pea fiber.

In some embodiments, the whitening agent is an insoluble salt, for example an insoluble calcium such as calcium carbonate, calcium sulphate, calcium phosphate, or tricalcium citrate, preferably calcium carbonate.

In some embodiments, the whitening agent is an oil-in-water emulsion. In one embodiment, the oil-in-water emulsion has been prepared by adding plant oils and plant proteins as surfactant during hydration with high shear for emulsification.

The invention further relates to a salmon analogue having a layered structure comprising a first layer and a second layer, wherein the first layer comprises a glucomannan source, a starch source, a carrageenan source, and optionally one or more of a colorant, a plant based oil comprising omega 3 or omega 6, a flavor, a pectin source; and the second layer comprises a glucomannan source, a starch source, a salt source, a whitening agent, and optionally a carrageenan source.

Detailed description of the invention

First layer or first mixture layer

The first layer or first mixture layer comprises a glucomannan source, a starch source, a salt source, a carrageenan source, and optionally one or more of a colorant, a DHA oil and flavor

When the glucomannan source is konjac flour, then the glucomannan content of the konjac flour is between 40-80% the glucomannan content in the first layer or first mixture layer is between 0.5 to 3 wt%. The konjac flour can be natural konjac flour, for example between 1 to 6 wt% konjac flour in the recipe. Other glucomannan sources are purified konjac gum or glucomannan, or deacetylated konjac glucomannan. The carrageenan source can be purified carrageenan, or a seaweed flour containing carrageenan. The seaweed can be Irish Moss or cottonii. Seaweed flour was found to provide the best freeze thaw stability and syneresis.

The salt source can be NaCI, KCI, CaCl2, an alkali agent such as Na2CO3, K2COs or a mixture thereof. When the salt source is NaCI, then the first layer or first mixture layer comprises between 0.5 to 3% NaCI. When the salt source is KCI, then the first layer or first mixture layer comprises between 0.05 to 1 % KCI. When the salt source is CaCl2, then the first layer or first mixture layer comprises between 0.05 to 2% CaCh The salt source can be a combination of NaCI and CaCh When the salt source is an alkali agent, then the first layer or first mixture layer comprises between 0.05 to 1 % Na2COs or K2CO3. The alkali salt is to control the pH and induce konjac heat set gelation which forms an elastic and heat stable texture and mimics the properties of the animal connective tissue.

The starch source and be a flour rich in starch, for example potato flour and tapioca flour. The starch source can be isolated starch, preferably potato starch or tapioca starch, or a mixture thereof. The starch source could be native or pregelatinized. Potato starch reduces gumminess and provides the creamy and soft texture close to animal raw and smoked salmon.

The oil is a plant based oil comprising omega 3 or omega 6. For example, the oil is a plant based oil comprising a combined total of 40% omega 3 and/or omega 6.

The pectin-rich fiber, for example citrus fiber at between 0.1 to 1 % improves the water holding capability and reduces syneresis during storage.

The colorant can have an orange, red, pink, or purple color. It can be a plant material or its extract, for example from carrot or paprika or pumpkin, or red lentil flour.

Second layer or second mixture layer

The second layer or second mixture layer can also be referred to as a white layer. It is located between the first layers or first mixture layers, which can be referred to as the orange or pink layers. The white layer comprises a glucomannan source, a starch source, a salt source, a whitening agent, and optionally a carrageenan source and a pectin source, wherein the whitening agent is a protein, a fiber or an insoluble salt

The whitening agent may be an insoluble fiber powder, for example bamboo fiber, cellulose fiber, or oat insoluble fiber, in combination with a white insoluble salt suspension, for example a calcium carbonate suspension.

The average diameter of the salt and protein particles can be in the range 0.1 to 200 microns. The average length of the fiber can be in the range 0.1 to 200 microns. For the emulsion, the average oil droplet size can be in the range 0.1 to 200 microns.

The white layer can be a liquid dough with a formulation similar to an orange first layer except for the orange color. The first layer and second layer connected with each other due to their ingredients in common.

The white layer can be injected, poured, sprayed or co-extruded with the first layer. The white layer can be a liquid at temperatures above 50°C when applying. This is referred to as a cold-set white layer. The white layer can be a dough containing konjac flour and alkali agent. This is referred to as a heat-set white layer. This dough is liquid at room temperature, and forms an elastic gel when heat is applied, for example at 80°C for Wminutes.

Protein, insoluble fiber and/or insoluble salt, oil in water emulsion are used to adjust the whiteness of the white layer.

Definitions

As used herein, the singular forms "a," "an" and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "an insoluble fiber source" or "the insoluble fiber source" includes two or more insoluble fiber sources.

The words "comprise," "comprises" and "comprising" are to be interpreted inclusively rather than exclusively. Likewise, the terms "include," "including" and "or" should all be construed to be inclusive, unless such a construction is clearly prohibited from the context.

The compositions disclosed herein may lack any element that is not specifically disclosed. Thus, a disclosure of an embodiment using the term "comprising" includes a disclosure of embodiments "consisting essentially of and "consisting of the components identified. Similarly, the methods disclosed herein may lack any step that is not specifically disclosed herein. Thus, a disclosure of an embodiment using the term "comprising" includes a disclosure of embodiments "consisting essentially of” and "consisting of” the steps identified.

The term "and/or" used in the context of "X and/or Y" should be interpreted as "X," or "Y," or "X and Y." Where used herein, the terms "example" and "such as," particularly when followed by a listing of terms, are merely exemplary and illustrative and should not be deemed to be exclusive or comprehensive. Any embodiment disclosed herein can be combined with any other embodiment disclosed herein unless explicitly stated otherwise.

As used herein, "about" and "approximately" are understood to refer to numbers in a range of numerals, for example the range of -10% to +10% of the referenced number, preferably within -5% to +5% of the referenced number, more preferably within -1% to +1 % of the referenced number, most preferably within -0.1 % to +0.1 % of the referenced number.

As used herein, a product “substantially devoid” of an ingredient means that none of that ingredient is added as such to the product, and that any of the ingredient present originates from minor traces or impurities present in other ingredients.

A vegan product is defined as being devoid of animal products, for example devoid of dairy products and meat products. A vegan fish analogue product of the invention has the look, taste, and texture which is close to the animal version. Preferably, the vegan fish analogue is a vegan salmon analogue.

The skilled person will appreciate that specific embodiments described herein may be used interchangeable between the product, method, and use aspects of the invention. The invention will now be illustrated by way of examples, which should in no way be thought to limit the scope of the invention as herein described.

EXAMPLES Example 1

Recipe and method to prepare a salmon analogue

Recipe for first layer (orange layer):

Recipe for second layer (white layer):

The first layer mixture is prepared by hydrating the dry mix of konjac flour, seaweed flour, protein powder, potato starch, and salts in water for 20min at room temperature while mixing, and then the mixture is heated at 85°C for 20min, colorant and flavors were added in the end of the heating. This dough is used to mimic muscle of fish fillet, for example salmon, tuna or white fish. The heating of the hydrated mixture can be done by a heating die with double jacket, in this case, the heating temperature is about 85-90°C, and the dough is cooled down to about 70°C for layering. The mass is functionalized when it is heated to more than 80°C.

The liquid white dough is prepared by hydrating the dry mix of konjac flour, seaweed flour, rice protein, potato starch, salts and pea fiber at room temperature for 20min, then the mixture is heated at 85°C for 20min before it is ready to use for layering. This while dough is to mimic the connective tissue of a fish fillet. Rice protein and pea fiber are used to provide the whiteness.

The first layer mixture is pushed through a die to generate the first layer which is loaded in a mould, and the liquid white dough is kept hot with temperature above 60°C before spraying as a thin layer on top of the first layer. On top of the white layer, another first layer mixture was added as the second colored layer. The layering is repeated to generated multiple colored and white layers to mimic animal salmon or other fish fillet. Instead of spraying, the liquid white dough can be injected by injectors.

After layering, the whole sample together with the mold is sealed in a package and heated in an oven at 75°C for 15min without shaking to increase the shelf-life. After the heating, the sample is cooled down in a fridge to set. The final sample can be cut into slides or blocks mimicing fish fillet. The white layer is designed to have stronger texture than orange dough as observed in raw animal fish fillet.

Example 2 Effect of citrus fiber and different starches on hardness and firmness

Recipes comprising citrus fiber and different starches were prepared according to the recipes in the tables below. Waxy starch, pea starch, and potato starch were tested.

The reference sample (without starch neither citrus fiber) was too gummy and chewy, which was reduced by the addition of potato starch. As shown in the figure below, the hardness from cutting test was decreased by addition of potato starch and it was increased by addition of pea starch and waxy starch. This correlates to tasting results which showed reduction of gumminess and improved softness and creaminess when potato starch was used. Similar observation with tapioca starch was captured. Potato starch and tapioca starch behaved similarly likely due to its medium ratio of amylose and amylopectin because pea starch is high in this ratio and waxy starch is very low in this ratio.

Citrus fiber at 0.5 wt% increased the hardness and especially the firmness, due to its soluble pectin fraction. Use at lower dosage e.g. at 0.3% together with potato or tapioca starch provided good texture, improved heat stability and reduced syneresis during storage.

Figure 1 shows the textural changes when adding different starches and citrus fiber in konjac-carrageenan base recipe (ref).

When potato starch was increased from 2 to 4%, the hardness of the sample was largely reduced (figure below) and the final product was too soft when 4% potato starch was used. A preferred concentration of starch with the selected base was with 3% which provided balanced softness. A native potato starch from different supplier (potato starch II) and tapioca starch were tested in the recipe at 3 wt% and showed good texture with balance softness and creaminess as well.

Figure 2 shows the impact of potato starch concentration and tapioca starch on hardness in cutting test.

Figure 3 shows that addition of starches and citrus fiber also improved the freeze-thaw stability, less water release and minimal textural changes were observed. Only pea starch showed large changes in hardness after freeze-thawing. Example 3

Impact of seaweed flour, citrus fiber on heat stability and reduction of syneresis of the final product

A comparison was made of the addition of seaweed flour at 1 .5 wt% (20% soluble fiber, commercially available, from Irish moss) with purified kappa carrageenan at 0.3 wt% with the base recipe described in example 1. Sample containing 1.5 wt% seaweed flour in recipe had similar texture to samples containing 0.3% kappa carrageenan, however the seaweed flour significantly improved the water holding capacity with less syneresis of water while chilled and frozen storage. When samples with the same shape and weight were heated in the pan at the same time, carrageenan sample melted fast and the seaweed flour sample had a considerably delayed melting. This improved heat stability is preferred to allow it to be used in some hot applications.

The addition of citrus fiber at 0.5 wt% in base recipe containing 0.3% kappa carrageenan was tested. Significant improvement on reduction of syneresis and heat stability was observed when comparing to with the recipe without citrus fiber.

Example 4

Impact of salt combination on heat stability of the final product

Different salt combinations were tested in base recipe containing 2% potato starch (recipe in example 2)

Samples prepared from NaCI and KCI presented the worst heat and freeze-thaw stability i.e. they melted fast and showed important syneresis during thawing. At room temperature, these samples were firmer than the other salt mixtures. Higher NaCI concentrations gave unstable and incompletely set gels.

Potassium- and sodium-induced gelation undergo different mechanisms, displayed by different gel structures, which may relate to the differences in stability observed. When both salts are mixed, a mixture of both gelation mechanisms occur, which could contribute to the lower stability of K+/Na+ gels compared to only K+ gel.

Calcium-induced gelation of kappa-carrageenan is less efficient than with monovalent ions, commonly due to a tighter packing of the polymer chain leading to phase separation as the calcium concentration increases. This is consistent with the lower strength of Na/Ca gels compared to Na/K.

Example 5

Heat stable gel recipes containing K2CO3

Recipes were prepared using konjac heat set gelation under alkaline conditions. The ingredient amounts are shown in the table below. Dough was hydrated at room temperature or at higher temperature to reduce hydration time. K2CO3 was added and the mixture was layered into a mold. The layered product was then heated to over 80°C for 5 min. A thermo-irreversible gel forms due to the alkaline and heat conditions. This makes the product suitable for hot application options. In comparison to the cold set carrageenan based gelation for gels containing only NaCI and/or KCI, the thermo-irreversible gel with K2CO3 is harder in first bite, firmer while chewing and overall less flexible and has a more smooth and slippery surface.

When varying the concentration of K2CO3 from 0.1 wt% to 0.4 wt%, keeping all other ingredients constant as given in the table below, lower concentrations of K2CO3 led to weaker gels and softer texture. For higher K2CO3 contents the texture is affected vice versa. Furthermore, it could be identified, that higher konjac powder concentrations enhances firmness and the melting perception in mouth when K2CO3 concentration was kept constant.