Arabinose-containing and galactose-containing food composition and process of preparation

Abstract

An arabinose-containing and galactose-containing food composition is disclosed. Said food food composition is liquid or semi-liquid or powdered. It comprises plant proteins, less than 5wt.%free uronicacid by dry weight and a free arabinose to free galactose weight ration of from 1:1.5 to 1:5. A process for preparing such a food composition is also disclosed. A method for selecting an enzyme comprising a pectinase activity suitable for the invention is also disclosed.

Description

[0001] ARABINOSE-CONTAINING AND GALACTOSE-CONTAINING FOOD COMPOSITION AND PROCESS OF PREPARATION

[0002] TECHNICAL FIELD

[0003] The present invention relates generally to the field of manufacturing a food composition in liquid, semi-liquid or powder format starting from low-refined plant proteincontaining ingredients. For example, the present invention relates to an arabinose-containing and galactose-containing food composition, a process for preparing such a food and a method for selecting an enzyme comprising a pectinase activity suitable for the invention.

[0004] BACKGROUND OF THE INVENTION

[0005] There is a growing consumer interest in plant-based food products, including plantbased beverages (PBBs), driven by various factors such as environmental sustainability and health considerations like lactose intolerance. PBBs, which are in their sensorial and / or textural characteristics, close to dairy beverages made from cow milk, are particularly sought after. Such PBBs are generally called plant-based dairy analogues (PBDAs) and are prepared with plant-based materials.

[0006] Starting from plant-based materials, the food industry is challenged to research processes that enable the production of plant-based dairy analogues that are as close as possible to the cow milk in their nutritional quality and sensorial experience.

[0007] Currently, plant protein isolates are often used in PBDAs to ensure to keep a significant protein content in the final product and to limit the technical challenges which are associated with the addition of less refined plant ingredients, such as plant flours. However, the use of refined plant ingredients, such as plant protein isolates, comes with challenges too. For example, the use of additives may be required to increase the stability, sensory and / or nutritional properties of the final PBDA product prepared with such isolates. Nonetheless, long ingredient lists that include additives may be perceived and associated by consumers with low naturality.

[0008] Considering the aforementioned challenges, there is interest to develop plant-based dairy analogues (PBDAs) using less refined plant ingredients. In particular, plant flours, such as wholegrain flours and legume flours, appear as promising ingredients for this purpose. However, less refined ingredients, like plant flours, are especially rich in protein, but also fibres, so called non-starch polysaccharides (NSP). NSPs are polysaccharides, like cellulose, hemicellulose, pectin and lignin, that are unique to plant-based foods, due to the cell wall structure that is part of the plant cells. The presence of NSPs leads to an increase in the soluble, but also insoluble fraction of the PBDA, depending on the plant source used.

[0009] The presence of NSPs, especially the insoluble fraction of NSPs, in less refined plant ingredients may lead to sensory, processability and / or stability challenges in the preparation of PBDAs. For example, they may lead to excessive increase in viscosity, upon heating, which may hindertheir processability and may lead to undesirable sensory properties. They may also lead to significant sedimentation in the final product which is undesirable on a sensory standpoint.

[0010] Hence, this would be desirable to provide plant-based liquid or semi-liquid food composition which has reduced level of non-starch polysaccharides, including insoluble nonstarch polysaccharides, in particular pectic polysaccharides.

[0011] It would also be desirable that the plant-based liquid or semi-liquid food composition is prepared from less refined plant ingredient such as plant flour.

[0012] It would also be desirable that the plant-based liquid or semi-liquid food composition has good nutritional properties, in particular a significant source of fibers and / or other nutritional compounds.

[0013] Any reference to prior art documents in this specification is not to be considered an admission that such prior art is widely known or forms part of the common general knowledge in the field.

[0014] SUMMARY OF THE INVENTION

[0015] It has been discovered that the insoluble fraction of NSPs (Non-Starch Polysaccharides) of less-refined plant ingredients, such as plant flours are rich in pectic polysaccharides. The pectic polysaccharides identified in these ingredients, in particular flours primarily consist of arabinose, galactose, and uronic acid units. This discovery is contrary to expectations as pectic polysaccharides are typically associated with the soluble fraction of NSPs and not with the insoluble fraction. This discovery highlights the significance of identifying and use enzymes that can degrade these pectic polysaccharides in an effective way. It has been discovered that the use of such enzymes significantly reduces the quantity of insoluble NSPs. In addition, the efficient enzymatic degradation of these pectic polysaccharides leads to an increased release of free arabinose and free galactose, further improving the overall composition of the product. The object of the present invention is to improve the state of the art, and in particular to provide a food composition, a process for preparing food composition and a method for selecting enzyme that overcome the problems of the prior art and addresses the needs described above, or at least to provide a useful alternative.

[0016] The inventors were surprised to see that the object of the present invention could be achieved by the subject matter of the independent claims. The dependent claims further develop the idea of the present invention.

[0017] Accordingly, a first aspect of the invention proposes an arabinose-containing and galactose-containing food composition, which is liquid or semi-liquid or powdered and which comprises:

[0018] - plant proteins selected from the list consisting of legume proteins, seed proteins and mixture thereof,

[0019] -less than 5wt.% free uronic acid by dry weight,

[0020] -a weight ratio of free arabinose to free galactose (free arabinose: free galactose) of from 1:1.5 to 1:5, preferably 1:3.5 to 1:4.5.

[0021] A second aspect of the invention proposes a method for selecting an enzyme comprising a pectinase activity which is high at a pH from 6 to 7.5, preferably at a pH of 7:

[0022] (a) providing one or several enzymes, providing a liquid substrate medium A comprising a substrate A and providing a liquid substrate B comprising a substrate B, wherein the substrate A is a molecule comprising an a-arabinofuranosyl-residue attached to a chromogenic compound, and wherein the substrate B is molecule comprising a p-galactosyl-residue attached to a chromogenic compound,

[0023] (b) contacting the liquid substrate medium A with said one or several enzymes for an incubation time of 1 to 15 minutes, at a temperature of 20°C to 60°C, preferably 30°C to 50, more preferably 37°C and at a pH from 6 to 7.5, preferably at a pH of 7 to obtain a treated liquid substrate medium A,

[0024] (c) contacting the liquid substrate medium B with said one or several enzymes for an incubation time of 1 to 15 minutes, at a temperature of 20°C to 60°C, preferably 30°C to 50, more preferably 37°C and at a pH from 6 to 7.5, preferably at a pH of 7 to obtain a treated liquid substrate medium B, (d) measuring the lytic activity towards pNP-a-L-arabinofuranoside in the treated liquid substrate medium A: after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute, after 1 minute, if the incubation time used in step (b) is equal to 1 minute,

[0025] (e) measuring the lytic activity towards pNP-|3-D-galactopyranoside in the treated liquid substrate medium B: after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute, after 1 minute, if the incubation time used in step (b) is equal to 1 minute,

[0026] (f) selecting the one or more enzymes that have:

[0027] - a lytic activity towards pNP-a-L-arabinofuranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium A based on the lytic activity measured in step (d), and

[0028] - a lytic activity towards pNP-|3-D-galactopyranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium B based on the lytic activity measured in step (e).

[0029] A third aspect of the invention proposes a process for preparing an arabinose- containing and galactose-containing food composition which is liquid or semi-liquid or powdered comprising the steps of:

[0030] (a) providing a low-refined plant protein-containing ingredient, wherein the low-refined plant protein-containing ingredient comprises pectic polysaccharides,

[0031] (b) dispersing the low-refined plant protein-containing ingredient in an aqueous liquid to form a plant dispersion,

[0032] (c) optionally, heat treating the plant dispersion,

[0033] (d) optionally, adjusting the pH of the plant dispersion to a pH of 6 to 7.5,

[0034] (e) contacting the plant dispersion which has a pH from 6 to 7.5 with an enzyme preparation comprising at least one enzyme comprising a pectinase activity to provide an enzymatically-treated arabinose-containing and galactose-containing liquid or semi-liquid food composition,

[0035] (f) heat-treating the enzymatically-treated arabinose-containing and galactose- containing liquid or semi-liquid food composition to inactivate the at least one enzyme comprising a pectinase activity and to form an arabinose-containing liquid or semi-liquid food composition,

[0036] (g) optionally, drying of the arabinose-containing and galactose-containing liquid or semiliquid food composition to form an arabinose-containing and galactose-containing powdered food composition, wherein the amount of free arabinose by dry weight is increased by at least 0.25wt% and the amount of free galactose by dry weight is increased by at least 0.3wt.% in the enzymatically- treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

[0037] These and other aspects, features and advantages of the invention will become more apparent to those skilled in the art from the detailed description of embodiments of the invention, in connection with the attached drawings.

[0038] BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 shows flow diagram of EtOH (A), hot water (B), acid (C) and alkaline (D) extraction procedures of pea flour including and yields thereof.

[0040] Figure 2 shows the release of free arabinose in a) pea flour and b) destarched pea flour after the incubation with various carbohydrate-degrading enzyme preparations before and after a protease (=Alcalase®) treatment.

[0041] Figure 3 shows the release of free galactose in pea flour after the incubation with various carbohydrate-degrading enzyme preparations before and after a protease (=Alcalase®) treatment.

[0042] Figure 4 shows the release of free arabinose (a - c) and free galactose (d - f) after the incubation different pea flour extracts (alkaline AIS fraction: a and d, Acid extract AIS fraction: b and e, water extract supernatant: c and f) with different expressed enzymes in absence or presence of a pectinase (Enzyme Preparation 16).

[0043] Figure 5 shows the release of free arabinose from destarched pea flour extract using pectinase Preparation 20 and a-L-arabinofuranosidase B (SEQ ID NO: 1).

[0044] Figure 6 shows the release of free arabinose in destarched pea flour after the incubation with a-L-arabinofuranosidase B (SEQ ID NO: 1) alone or both a-L- arabinofuranosidase B (SEQ ID NO: 1) and arabinan endo-l,5-a-L-arabinosidase C (SEQ ID NO: 4).

[0045] DETAILED DESCRIPTION OF THE INVENTION

[0046] As used in the specification, the words "comprise", "comprising" and the like are to be construed in an inclusive sense, that is to say, in the sense of "including, but not limited to", as opposed to an exclusive or exhaustive sense.

[0047] As used in the specification, the word "about" should be understood to apply to each bound in a range of numerals. Moreover, all numerical ranges should be understood to include each whole integer within the range.

[0048] As used in the specification, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise.

[0049] As used in the specification, the term "substantially free" means that no more than about 10 weight percent, preferably no more than about 5 weight percent, and more preferably no more than about 1 weight percent of the excluded material is present. In a preferred embodiment, "substantially free" means that no more than about 0.1 weight percent of the excluded material remains. "Entirely free" typically means that at most only trace amount of the excluded material is present, and preferably, no detectable amount is present. Conversely, "substantially all" typically means that at least about 90 weight percent, preferably at least about 95 weight percent, and more preferably at least about 99 weight percent of the material is present.

[0050] Unless noted otherwise, all percentages in the specification refer to weight percent, where applicable.

[0051] Unless defined otherwise, all technical and scientific terms have and should be given the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0052] As used herein, the term "acidifying ingredient" refers to an ingredient that comprises organic acids and / or inorganic acids and whose primary function is to lower the pH of the products it is incorporated into.

[0053] As used herein, the term "alkalinizing ingredient" refers to an ingredient that comprises organic bases and / or inorganic bases and whose primary function is to raise the pH of the products it is incorporated into.

[0054] As used herein, the term "pectic polysaccharides" are a group of carbohydrates derived from plant and comprising homogalacturonan compartments, xylogalacturonan compartments, rhamnogalacturonan-l compartments, rhamnogalacturonan-ll compartments, arabinan compartments, arabinogalactan compartments, arabinogalactan- protein complexes and combinations thereof.

[0055] As used herein, the term "polysaccharides" refers to polysaccharides having a DP of 10 or above different from starch.

[0056] As used herein, the term "oligosaccharides" refers to polysaccharides having a DP of 10 or above different from starch.

[0057] As used herein, the term "non-starch polysaccharides" refers to polysaccharides or above different from starch.

[0058] As used herein, the term "carbohydrates" refers to monosaccharides, disaccharides, oligosaccharides and polysaccharides.

[0059] As used herein, the term "whole plant material", "whole seed" or "whole legume", it is understood respectively a plant material, seed or legume which is integral and comprises the germ and the endosperm, optionally the bran and the hull. Preferably, the whole plant material, whole seed or whole legume is integral and comprises the germ, the endosperm, and the bran, and optionally the hull. More preferably, the whole plant material, whole seed or whole legume is integral and comprises the germ, the endosperm, the bran, and the hull. By "integral", it is understood that the plant material, seed or legume is intact and has not undergone any step of mechanical disruption.

[0060] As used herein, the term "ground whole plant material", "ground whole seed" or "ground whole legume" refers to whole plant materials, whole seeds or whole legumes who have been ground by milling, crushing, mixing and / or grinding, in presence or absence of an aqueous liquid, preferably water, to form a suspension, liquid or powder.

[0061] As used herein, the term "ground heated whole plant material", "ground heated whole seed" or "ground heated whole legume" refers to whole plant materials, whole seeds or whole legumes who has been heated above 60°C, for example by boiling in water after being ground by milling, crushing, mixing and / or grinding, in presence or absence of an aqueous liquid, preferably water. The heat-treatment step may contribute to reduce or remove undesirable microorganisms from the whole plant materials, whole seeds or whole legumes. This heattreatment step may also contribute to starch gelatinization in whole plant materials, whole seeds or whole legumes that may facilitate starch degradation through enzymatic treatment.

[0062] In a first aspect, the invention relates to an arabinose-containing and galactose- containing food composition. The arabinose-containing and galactose-containing food composition food composition is liquid or semi-liquid or powdered. In a preferred embodiment, the arabinose-containing and galactose-containing food composition is liquid.

[0063] In some embodiment, the arabinose-containing and galactose-containing food composition may be selected from the list consisting of beverage, yogurt, pudding, custard, porridge, cocoa-containing beverage, malt-containing beverage, confectionery, creamer, sauce, dressing, spreads, bouillon, condensed milk analogue, cream, infant formula, baby food, food for special medical purpose (FSMP), soup, dietary supplement, nutritional composition, complete nutritional composition, oral nutritional supplement, medical food, nutraceutical and mixture thereof.

[0064] In a preferred embodiment, the arabinose-containing and galactose-containing food composition is a beverage. In a more preferred embodiment, the beverage is selected from the list of plain or flavoured milk beverage, plain or flavoured plant-based milk beverage analogue, plain or flavoured plant-based milk analogue, plain or flavoured drinkable yogurt, plain or flavoured plant-based drinkable yogurt analogue, smoothies, milkshake, juice, cocoacontaining beverage, malt-containing beverage, coffee, tea, carbonated beverage, energy drink, and mixture. In a most preferred embodiment, the beverage is plain or flavoured plantbased milk analogue.

[0065] In an embodiment, the arabinose-containing and galactose-containing food composition is vegetarian. In some other embodiment, the arabinose-containing and galactose-containing food composition is vegan.

[0066] The arabinose-containing and galactose-containing food composition comprises plant proteins. The plant proteins are selected from the list consisting of legume proteins, seed proteins and mixture thereof.

[0067] In some embodiment, the arabinose-containing and galactose-containing food composition comprises at least 0.5wt.%, more preferably 0.5wt% to 60wt%, even more preferably 0.5wt% to 20wt.%, even more preferably lwt.% to 15wt.%, even more preferably lwt.% to 6wt.% plant proteins.

[0068] When the arabinose-containing and galactose-containing food composition is liquid or semi-liquid, the arabinose-containing and galactose-containing food composition comprises at least 0.5wt.%, more preferably 0.5wt% to 25wt%, even more preferably 0.5wt% to 20wt.%, even more preferably lwt.% to 15wt.%, even more preferably lwt.% to 6wt.% plant proteins.

[0069] When the arabinose-containing and galactose-containing food composition is powdered, the arabinose-containing and galactose-containing food composition comprises at least 4wt.%, more preferably 4wt% to 60wt%, even more preferably 4wt% to 20wt.%, even more preferably 4wt.% to 15wt.%, even more preferably 4wt.% to 6wt.% plant proteins.

[0070] In some embodiment, the arabinose-containing and galactose-containing food composition comprises at least 0.5wt.%, more preferably 0.5wt% to 25wt%, even more preferably 0.5wt% to 20wt.%, even more preferably lwt.% to 15wt.% proteins. In some embodiment, the proteins comprise milk proteins and plant proteins. In some other embodiment, the proteins consist only of plant proteins.

[0071] In some embodiment, the legume proteins of the arabinose-containing and galactose- containing food composition come from the legume selected from the list consisting of pea, cowpea, split pea, peanut, lentil, faba bean, chickpea, bean, soybean and combination thereof. For example, the bean may be selected from the list consisting of navy bean, black bean, butter bean, red bean, green bean, kidney bean, pinto bean, lima bean, cannellini bean, adzuki bean, mung bean, cranberry bean, Great Northern bean, yellow eye bean, black turtle bean, calypso bean, tongue of fire bean, and mixture thereof.

[0072] In some embodiment, the seed proteins of the arabinose-containing and galactose- containing food composition come from the seed selected from the list consisting of almond, rapeseed, sunflower seed, hemp seed, flaxseed, linseed, sesame seed, pumpkin seed, chia seed and mixture thereof.

[0073] In a more preferred embodiment, the plant proteins are legume proteins. The legume proteins are legume proteins that come from legumes as provided above. In an even more preferred embodiment, the legume proteins come from the legume selected from the list consisting of pea, cowpea, split pea, chickpea, bean, soybean and combination thereof. For example, the bean may be selected from the list consisting of navy bean, black bean, butter bean, red bean, green bean, kidney bean, pinto bean, lima bean, cannellini bean, adzuki bean, mung bean, cranberry bean, Great Northern bean, yellow eye bean, black turtle bean, calypso bean, tongue of fire bean, and mixture thereof.

[0074] In some embodiment, the plant proteins are provided as low-refined plant proteincontaining ingredient. The low-refined plant protein-containing ingredient is selected from the list consisting of plant flour, plant protein concentrate, ground whole legume, ground whole seed and mixture thereof. In some embodiment, the ground whole legume may be ground heated whole legume. In some embodiment, the ground whole seed may be ground heated whole seed. These low-refined plant protein-rich ingredients contain a substantial quantity of plant protein and retain a greater amount of macro- and micro-nutrients naturally found in seeds and legumes, compared to their more refined counterparts like plant protein isolates. They also typically have fewer off-notes than their more refined counterparts, like plant protein isolates. Therefore, using these low-refined plant protein-rich ingredients is beneficial as they improve the nutritional profile and sensory experience of the food composition in which they are incorporated compared to more processed ingredients like plant protein isolates. Moreover, due to their minimal processing, these ingredients contribute to waste reduction and may enhance the perceived naturalness of the food composition. In a most preferred embodiment, the low-refined plant protein-rich ingredient is plant flour. Plant flours are highly favored as they strike an optimal balance between providing a substantial amount of protein and fibers (higher than in ground seeds / legumes) and other micro- and macro-nutrients simultaneously (higher than in plant protein concentrate). In some embodiment, the low-refined protein-rich ingredient is treated enzymatically with at least one enzyme comprising a pectinase activity. The at least one enzyme comprising a pectinase activity may be as disclosed in the third aspect of the invention. The enzymatic treatment may be performed under the same conditions (T°C, time, pH, enzyme concentration) as provided in the third aspect of the invention. The benefits of such enzymes are disclosed in the third aspect of the invention.

[0075] In some embodiment, the low-refined protein-rich ingredient may be further treated enzymatically with at least one starch-degrading enzyme and / or at least one cellulosedegrading enzyme and / or protein-degrading enzyme. The at least one starch-degrading enzyme may be as disclosed in the third aspect of the invention. The at least one cellulosedegrading enzyme may be as disclosed in the third aspect of the invention. The at least one protein-degrading enzyme may be as disclosed in the third aspect of the invention. The enzymatic treatment with the starch-degrading enzyme and / or cellulose-degrading enzyme and / or protein-degrading enzyme may be performed under the same conditions (T°C, time, pH, enzyme concentration) as provided in the third aspect of the invention. The benefits of such starch-degrading enzymes and cellulose-degrading enzymes are disclosed in the third aspect of the invention.

[0076] The use of low-refined protein-rich ingredient results in the presence of significant amount of carbohydrates. Especially, in some embodiment, the arabinose-containing and galactose-containing food composition may comprise at least 15wt.%, preferably 15% to 60 wt.% carbohydrate by dry weight.

[0077] The arabinose-containing and galactose-containing food composition has a limited amount of free uronic acid. Specifically, the arabinose-containing and galactose-containing food composition further comprises less than 5wt.%, preferably less than 3wt.% free uronic acid by dry weight. In some embodiment, it comprises more than 0.5wt.% free uronic acid by dry weight.

[0078] The term "free uronic acid" refers to uronic acid that is present in a composition or formulation without being chemically bound or conjugated to other components. It signifies the availability of uronic acid as a separate entity within the composition, which may have specific implications for its functionality, bioavailability, or targeted applications.

[0079] Free uronic acid is a by-product of the pectinase activity of enzymes on the pectic polysaccharides found in the non-starch polysaccharides, including the insoluble ones, of low- refined protein-containing ingredients coming from seeds and legumes (hereinafter "low- refined seed / legume ingredients"). Free uronic acid is released but remain limited as only pectin from the low-refined seed / legume ingredients is hydrolysed. No pectin ingredient, e.g. pure pectin, is added before hydrolysis as it will result in an important amount of free uronic acid. The addition of pectin is not advantageous because it is an additive as it is not inherently present in the low-refined seed / legume ingredients and would increase the length of the ingredient list. Without wishing to be bound by theory, the generation of excessive amount of free uronic acid is undesirable as it can impact the pH and the sensory properties, including the taste of the final product.

[0080] In some embodiment, the arabinose-containing and galactose-containing food composition has a weight ratio of free arabinose to free galactose (free arabinose: free galactose) of from 1:1.5 to 1:5, preferably 1:3.5 to 1:4.5.

[0081] The term "free arabinose" refers to arabinose that is present in a composition or formulation without being chemically bound or conjugated to other components. It signifies the availability of arabinose as a separate entity within the composition, which may have specific implications for its functionality, bioavailability, or targeted applications.

[0082] Likewise, the term "free galactose" refers to galactose that is present in a composition or formulation without being chemically bound or conjugated to other components. It signifies the availability of galactose as a separate entity within the composition, which may have specific implications for its functionality, bioavailability, or targeted applications.

[0083] The use of enzymes with pectinase activity on low-refined seed / legume ingredients result in the release of free arabinose and free galactose in specific ratio. This release of free arabinose and free galactose, at the expense of pectic polysaccharides, enhances the overall carbohydrate composition of the food composition. Indeed, the breakdown of pectic polysaccharides into smaller carbohydrates reduces considerably the amount of non-starch polysaccharides, including insoluble ones and mitigates, limits or reduces the issues associated with such non-starch polysaccharides, such as viscosity and stability challenges. Moreover, without wishing to be bound by theory, the release of free galactose and free arabinose (i.e. in an abound form) may provide further properties and functionalities, including sensory and nutritional properties. For example, it can increase the taste properties, in particular sweetness of the final food composition.

[0084] In some embodiment, the arabinose-containing and galactose-containing food composition has a weight ratio of endogenous free arabinose to endogenous free galactose (endogenous free arabinose: endogenous free galactose) of from 1:1.5 to 1:5, preferably 1:3.5 to 1:4.5.

[0085] The term "endogenous free arabinose" refers to free arabinose that come from within the low-refined plant protein-containing ingredients as provided above and used in the food composition. It is not added separately but it is naturally occurring in these ingredients or is released as a result of the reaction of the enzyme(s) with pectinase activity on these ingredients. The term distinguishes the free arabinose present inherently in the low-refined plant protein-containing ingredient (i.e. naturally occurring or released from treatment with the enzyme(s) with pectinase activity) which is herein considered "endogenous" from any free arabinose that may be added as an ingredient or additive which is herein considered "exogenous".

[0086] The term "endogenous free galactose" refers to free galactose that come from within the low-refined plant protein-containing ingredients as provided above and used in the food composition. It is not added separately but it is naturally occurring in these ingredients or is released as a result of the reaction of the enzyme(s) with pectinase activity on these ingredients. The term distinguishes the free galactose present inherently in the low-refined plant protein-containing ingredient (i.e. naturally occurring or released from treatment with the enzyme(s) with pectinase activity) which is considered herein "endogenous" from any galactose that may be added as an ingredient or additive which is considered herein "exogenous".

[0087] The terms "endogenous free arabinose" and "free endogenous arabinose" are used interchangeably. Likewise, the terms "endogenous free galactose" and "free endogenous galactose" are used interchangeably.

[0088] In some embodiment, the arabinose-containing and galactose-containing food composition may further comprise less than 20wt.%, preferably less than 15wt.%, more preferably less than 10wt.%, even more preferably less than 5wt.%, even more preferably less than lwt.% pectic polysaccharides by dry weight. In some embodiment, the arabinose- containing and galactose-containing food composition may be free from pectic polysaccharides. Despite the use of low-refined protein-containing ingredient coming from seed and / or legumes, the use of enzyme comprising pectinase activity results in significant reduction of pectic polysaccharides in food composition with such ingredients, even at neutral or nearly neutral pH. It has been discovered that the non-starch polysaccharides of low- refined protein-rich ingredient coming seeds and / or legumes are abundant in pectic polysaccharides. Hence, the hydrolysis of pectic polysaccharides result in a significant limitation of non-starch polysaccharides, including their insoluble fractions. This significantly limit the issues associated with such non-starch polysaccharides, including their insoluble fractions in the food composition.

[0089] In some embodiment, the pectic polysaccharides of the arabinose-containing and galactose-containing food composition consist only of endogenous pectic polysaccharides. The term "endogenous pectic polysaccharides" refers to pectic polysaccharides that come from within the low-refined plant protein-containing ingredients as provided above and used in the food composition. They are not added separately but they are naturally occurring in these ingredients. The term distinguishes the pectic polysaccharides present inherently in the low-refined plant protein-containing ingredient from any pectic polysaccharides that may be added as an ingredient or additive.

[0090] In some embodiment, the arabinose-containing and galactose-containing food composition comprises at least 0.25wt.%, preferably at least 0.4wt free arabinose by dry weight. In some embodiment, the arabinose-containing and galactose-containing food composition comprises at most 5wt.%, preferably at least 3wt.%, more preferably at most 1.5wt.% free arabinose by dry weight. In some embodiment, the free arabinose consists only of endogenous free arabinose.

[0091] In some embodiment, the free arabinose of the arabinose-containing and galactose- containing food composition comprises or consists only of endogenous free arabinose and at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the endogenous free arabinose of the arabinose-containing and galactose-containing food composition is derived from a legume and / or a seed. In some preferred embodiment, the free arabinose of the arabinose-containing and galactose-containing food composition comprises or consists only of endogenous free arabinose and all of the endogenous free arabinose of the arabinose-containing and galactose-containing food composition is derived from a legume and / or a seed. In a preferred embodiment, the endogenous free arabinose and the vegetable protein and / or seed proteins are derived from the same legume and / or seed. Accordingly, the legume is selected from the list consisting of pea, cowpea, split pea, peanut, lentil, faba bean, chickpea, bean, soybean and combination thereof. For example, the bean may be selected from the list consisting of navy bean, black bean, butter bean, red bean, green bean, kidney bean, pinto bean, lima bean, cannellini bean, adzuki bean, mung bean, cranberry bean, Great Northern bean, yellow eye bean, black turtle bean, calypso bean, tongue of fire bean, and mixture thereof. The seed is selected from the list consisting of almond, rapeseed, sunflower seed, hemp seed, flaxseed, linseed, sesame seed, pumpkin seed, chia seed and mixture thereof.

[0092] The process of the invention generates arabinose from the plant protein ingredients. The arabinose is therefore endogenous and derived essentially from the plant protein ingredients. This advantageously enables to provides arabinose while limiting the addition of exogenous ingredient, in particular exogenous arabinose.

[0093] In some embodiment, the free galactose of the arabinose-containing and galactose- containing food composition comprises or consists only of endogenous free galactose and at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the endogenous free galactose of the arabinose-containing and galactose-containing food composition is derived from a legume and / or a seed. In some preferred embodiment, the free galactose of the arabinose-containing and galactose-containing food composition comprises or consists only of endogenous free galactose and all of the endogenous free galactose of the arabinose-containing and galactose-containing food composition is derived from a legume and / or a seed. In a preferred embodiment, the endogenous free arabinose and the vegetable protein and / or seed proteins are derived from the same legume and / or seed. Accordingly, the legume is selected from the list consisting of pea, cowpea, split pea, peanut, lentil, faba bean, chickpea, bean, soybean and combination thereof. For example, the bean may be selected from the list consisting of navy bean, black bean, butter bean, red bean, green bean, kidney bean, pinto bean, lima bean, cannellini bean, adzuki bean, mung bean, cranberry bean, Great Northern bean, yellow eye bean, black turtle bean, calypso bean, tongue of fire bean, and mixture thereof. In particular, the seed is selected from the list consisting of almond, rapeseed, sunflower seed, hemp seed, flaxseed, linseed, sesame seed, pumpkin seed, chia seed and mixture thereof. The process of the invention generates arabinose from the plant protein ingredients. The galactose is therefore endogenous and derived essentially from the plant protein ingredients. This advantageously enables to provides galactose while limiting the addition of exogenous ingredient, in particular exogenous galactose.

[0094] In some embodiment, the arabinose-containing and galactose-containing food composition comprises at least 0.3wt.%, preferably at least 0.7wt.%, more preferably at least 1.5wt.% free galactose by dry weight. In some embodiment, the arabinose-containing and galactose-containing food composition comprises at most 5wt.%, preferably at most 4wt.%, more preferably at most 3wt.% free galactose by dry weight. In some embodiment, the free arabinose consists only of endogenous free galactose.

[0095] In some embodiment, the arabinose-containing and galactose-containing food composition comprises less than 30wt.%, preferably less than 15wt.%, more preferably less than 10wt.%, even more preferably less than 5wt.%, even more preferably less than 3wt.%, even more preferably lwt.% starch by dry weight. In some embodiment, the arabinose- containing and galactose-containing food composition may be free from starch.

[0096] In some embodiment, the arabinose-containing and galactose-containing food composition may comprise less than 30wt%, preferably less than 20wt.%, more preferably 1 to 10wt.%, even more preferably 5 to 10wt.% non-starch polysaccharides by dry weight.

[0097] In some embodiment, the arabinose-containing and galactose-containing food composition may comprise less than 25wt%, preferably less than 20wt.%, more preferably 2wt.% to 20wt.%, even more preferably 5wt.% to 10wt.% insoluble non-starch polysaccharides by dry weight.

[0098] The invention provides food compositions with limited content of non-starch polysaccharides, including insoluble non-starch polysaccharides. This limited content is ensured despite the use of low-refined plant protein-containing ingredients. In particular, the non-starch polysaccharides, including insoluble non-starch polysaccharides are abundant in pectic polysaccharides. The effective hydrolysis of pectic polysaccharides contributes to limit considerably the total content in non-starch polysaccharides, including insoluble non-starch polysaccharides.

[0099] Ultimately, the limited content in non-starch polysaccharides, including insoluble non- starch polysaccharides results in the limitation of problems associated with non-starch polysaccharides, including insoluble non-starch polysaccharides.

[0100] In some embodiment, the arabinose-containing and galactose-containing liquid or semi-liquid or powdered food composition may further comprise fat. In particular, the arabinose-containing and galactose-containing liquid or semi-liquid or powdered food composition may comprise 0-40wt.%, preferably 0.5-40wt.% fat, more preferably 0.5-15wt.% fat. The fat may be animal fat and / or vegetable fat. In a particular, the animal fat may be milk fat. In some embodiment, the fat consists only of vegetable fat.

[0101] In some embodiment, the arabinose-containing and galactose-containing liquid or semiliquid or powdered food composition may further comprise an ingredient selected from the list consisting of flavouring ingredient, color, vitamins, minerals, emulsifier, bioactive compound, texturizing agent, buffer salts, sweetening agent, bulking agent, and mixture thereof.

[0102] In some embodiment, the arabinose-containing and galactose-containing liquid or semiliquid or powdered food composition may be obtained or obtainable by a process according to the third aspect of the invention.

[0103] In some embodiment, the arabinose-containing and galactose-containing food composition may be free from camomille, xylose, ramnose, nettle, myristic acid, cinnamic acid and mixtures thereof.

[0104] In a second aspect, the invention relates to a method for selecting an enzyme comprising a pectinase activity which is high at a pH from 6 to 7.5. Preferably, it relates to a method for selecting an enzyme comprising a pectinase activity which is high at a pH of 7.

[0105] The method comprises a step (a) of providing one or several enzymes, providing a liquid substrate medium A and providing a liquid substrate medium B.

[0106] The liquid substrate medium A comprises a substrate A. The substrate A is a molecule comprising an arabinofuranosyl residue attached to a chromogenic compound. In particular, the arabinofuranosyl residue is attached to a chromogenic compound by a bond which is cleavable by an enzyme comprising a pectinase activity, in particular comprising an a- arabinofuranosidase activity. The chromogenic compound may be any chromogenic compound known in the art. In one embodiment, the liquid substrate medium A comprises at least 0.5 mM, preferably 0.5-100 mM, more preferably 0.5 to 10 mM, even more preferably 0.5 to 5 mM, most preferably 2.4 mM of substrate A. In a preferred embodiment, the substrate A is pNP-a-L-arabinofuranoside. pNP-a-L-arabinofuranoside corresponds to p-Nitrophenyl-a- L-arabinofuranoside.

[0107] The liquid substrate medium B comprises a substrate B. The substrate B is a molecule comprising a p-galactosyl residue attached to a chromogenic compound. In particular, the |3- galactosyl residue is attached to a chromogenic compound by a bond which is cleavable by an enzyme comprising a pectinase activity, in particular comprising a p-galactanase activity. The chromogenic compound may be any chromogenic compound known in the art. In one embodiment, the liquid substrate medium B comprises at least 0.5mM, preferably 0.5- lOOmM, more preferably 0.5 to 10 mM, even more preferably 0.5 to 5 mM, most preferably 2.4mM of substrate B. In a preferred embodiment, the substrate B is pNP- -D- galactopyranoside. pNP-|3-D-galactopyranoside corresponds to p-Nitrophenyl-beta-D- galactopyranoside.

[0108] In some embodiment, the one or several enzymes provided in step (a) may be diluted in an aqueous liquid before step (b) and before step (c). In one embodiment, the aqueous liquid has a pH of 6 to 7.7, preferably of 7. In one embodiment, the aqueous liquid comprises a buffer salt. The buffer contributes to maintain neutral or nearly neutral pH. For example, the buffer salt may be Bis-Tris buffer, preferably O.IM Bis-Tris buffer. In some embodiment, the one or several enzymes may be diluted at least 2 times, preferably 2 times to 500 times. In some embodiment, the one or several enzymes provided in step (a) is not diluted in an aqueous liquid before step (b) and before step (c).

[0109] The method further comprises a step (b) of contacting the liquid substrate medium A with said one or several enzymes to obtain a treated liquid substrate medium A. The liquid substrate medium A is contacted with said one or several enzymes for an incubation time of 1 to 15 minutes, at a temperature of 20°C to 60°C, preferably of 30°C to 50, more preferably of 37°C and at a pH from 6 to 7.5 preferably at a pH of 7.

[0110] The method further comprises a step I of contacting the liquid substrate medium B with said one or several enzymes to obtain a treated liquid substrate medium B. for an incubation time of 1 to 15 minutes, at a temperature 20°C to 60°C, preferably of 30°C to 50, more preferably of 37°C and at a pH from 6 to 7.5, preferably at a pH of 7.

[0111] The steps (b) and (c) may be performed in parallel (i.e. simultaneously) or sequentially.

[0112] The method further comprises a step (d) of measuring the lytic activity towards pNP-a-L- arabinofuranoside in the treated liquid substrate medium A. The lytic activity towards pNP-a- L-arabinofuranoside in the treated liquid substrate medium A is measured:

[0113] -after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute,

[0114] -after 1 minute, if the incubation time used in step (b) is equal to 1 minute.

[0115] In a preferred embodiment, the lytic activity towards pNP-a-L-arabinofuranoside in the treated liquid substrate medium A may be measured by measuring the absorbance with a spectrophotometer or photometric analyzer at a wavelength of 380-420 nm, preferably of 405 nm. For example, the absorbance may be measured with the following photometric analyzer: semi-automatic GalleryTM Plus system (Thermo Fisher Scientific). The method further comprises a step (e) of measuring the lytic activity towards pNP-|3-D- galactopyranoside in the treated liquid substrate medium B. The lytic activity towards pNP-|3- D-galactopyranoside in the treated liquid substrate medium B is measured: after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute, after 1 minute, if the incubation time used in step (b) is equal to 1 minute.

[0116] The steps (d) and I may be performed in parallel (i.e. simultaneously) or sequentially.

[0117] The lytic activity towards pNP-|3-D-galactopyranoside in the treated liquid substrate medium B may be measured by measuring the absorbance with a spectrophotometer or photometric analyzer at a wavelength of 380-420 nm, preferably of 405nm. For example, the absorbance may be measured with the following photometric analyzer: semi-automatic GalleryTM Plus system (Thermo Fisher Scientific).

[0118] The method further comprises a step (f) of selecting the one or more enzymes that have:

[0119] - a lytic activity towards pNP-a-L-arabinofuranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium A based on lytic activity measured in step (d), and

[0120] - a lytic activity towards pNP-|3-D-galactopyranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium B based on the lytic activity measured in step I.

[0121] The term "undiluted enzyme" refers to the calculation of lytic activity based on the initial concentration of the enzyme before any dilution is applied. In simpler terms, if the enzyme was diluted before steps (b) and (c), the dilution factor is reversed to determine the lytic activity per gram of undiluted enzyme, rather than per gram of the diluted enzyme.

[0122] Enzymes comprising pectinase activity are generally suitable for the treatment fruitbased products that are very acidic. Hence, such enzymes have an optimal activity at very acidic pH. However, they are generally not suitable for enzymatic treatment at neutral or nearly neutral pH and their enzyme activity in such pH conditions is generally unsatisfactory. Such enzymes are therefore not adapted for the treatment of other plant materials that has generally a neutral or nearly neutral pH, such as low-refined plant protein-containing ingredients as provided in the first aspect of the invention. Likewise, such enzymes are therefore not adapted for the preparation of food compositions whose process is mainly performed at nearly neutral or neutral pH, for example plant-based milk analogues. The present method allows to identify enzymes having pectinase activity which have an important lytic activity under nearly neutral or neutral pH conditions. Such enzymes allow the effective removal of pectic polysaccharides in low-refined plant protein-containing ingredients as provided in the first aspect of the invention.

[0123] In a third aspect, the invention relates to a process for preparing an arabinose- containing and galactose-containing food composition which is liquid or semi-liquid or powdered.

[0124] The arabinose-containing and galactose-containing food composition may be arabinose-containing and galactose-containing food composition as provided in the first aspect of the invention.

[0125] The process comprises a step (a) of providing a low-refined plant protein-containing ingredient. The low-refined plant protein-containing ingredient comprises pectic polysaccharides.

[0126] In some embodiment, the low-refined plant protein-containing ingredient may comprise at least 5wt.%, preferably 5wt.% to 25wt.%, more preferably 5wt.% to 20wt.%, even more preferably 5wt.% to 15wt.%, even more preferably 5wt.% to 10wt.% pectic polysaccharides.

[0127] In some embodiment, the proteins of the low-refined plant protein-containing ingredient consist only of plant proteins. In some embodiment, the low-refined plant proteincontaining ingredient may comprise at least 5wt.%, preferably from 5 to 75wt.%, more preferably 15 to 50wt.%, even more preferably 15wt.% to 25wt.% plant proteins.

[0128] In some embodiment, the low-refined plant protein-containing ingredient may comprise at least 15wt.%, preferably 15wt.% to 70wt.%, more preferably 40wt.% to 65wt.% carbohydrates.

[0129] In some embodiment, the low-refined plant protein-containing ingredient may comprise at least 2wt.%, preferably 2 to 65wt.% starch.

[0130] In some embodiment, the low-refined plant protein-containing ingredient may comprise at least 5wt.%, preferably 5 to 40wt.% non-starch polysaccharides.

[0131] In some embodiment, the low-refined plant protein-containing ingredient may comprise at least 20%, preferably 20 to 75% of the non-starch polysaccharides are insoluble non-starch polysaccharides.

[0132] In some embodiment, the proteins of the low-refined plant protein-containing ingredient consist only of plant proteins. In some embodiment, the low-refined plant protein- containing ingredient is vegetarian. In some embodiment, the low-refined plant proteincontaining ingredient is vegan.

[0133] In an embodiment, the low-refined plant protein-containing ingredient is selected from the list consisting of plant flour, plant protein concentrate, ground whole plant material, and mixture thereof. The ground whole plant material may be ground heated whole plant material. The advantage of the use of low-refined plant protein-containing ingredient is provided in the first aspect of the invention.

[0134] In an embodiment, the low-refined plant protein-containing ingredient comes from seed and / or legume. The seed and legume may be respectively seeds and legumes as provided in the first aspect of the invention.

[0135] When the low-refined plant protein-containing ingredient is ground whole plant material and comes from seed, the ground whole plant material is ground whole seed. Likewise, when the low-refined plant protein-containing ingredient is ground heated whole plant material and comes from seed, the ground whole plant material is ground heated whole seed.

[0136] When the low-refined plant protein-containing ingredient is ground whole plant material and comes from legume, the ground whole plant material is ground whole legume. Likewise, when the low-refined plant protein-containing ingredient is ground heated whole plant material and comes from legume, the ground whole plant material is ground heatedwhole legume.

[0137] In a particular embodiment, the seed may be selected from the list consisting of almond, rapeseed, sunflower seed, hemp seed, flaxseed, linseed, sesame seed, pumpkin seed, chia seed and mixture thereof.

[0138] In a particular embodiment, the legume may be selected from the list consisting of pea, cowpea, split pea, peanut, lentil, faba bean, chickpea, bean, soybean and combination thereof. More preferably, the legume may be selected from the list consisting of pea, cowpea, split pea, chickpea, bean, soybean and combination thereof. For example, the bean may be selected from the list consisting of navy bean, black bean, butter bean, red bean, green bean, kidney bean, pinto bean, lima bean, cannellini bean, adzuki bean, mung bean, cranberry bean, Great Northern bean, yellow eye bean, black turtle bean, calypso bean, tongue of fire bean, and mixture thereof. The process comprises a step (b) of dispersing the low-refined plant protein-containing ingredient provided in step (a) in an aqueous liquid to form a plant dispersion. In some embodiment, the plant dispersion of step (b) has a pH of 6 to 7.5, preferably of pH of 7.

[0139] In some embodiment, before step (e), the plant dispersion may be further mixed with ingredients selected from the list consisting of flavouring ingredient, color, vitamins, minerals, emulsifier, bioactive compound, texturizing agent, buffer salts, sweetening agent, bulking agent and mixture thereof.

[0140] In some embodiment, before step (e), the plant protein dispersion may be further mixed with a milk protein-containing ingredient. For example, the milk protein-containing ingredient may be selected from the list consisting of buttermilk, condensed milk, liquid milk, milk powder, cream, acid whey, whey, milk protein concentrate, whey protein concentrate, casein concentrate, micellar casein contrate, whey protein isolate, casein isolate, yogurt, and mixture thereof. The milk protein-containing ingredient is derived from non-human mammal milk, preferably cow milk. The milk protein-containing ingredient may comprise at least 2wt.%, preferably 2 to 99wt.%, more preferably 3 to 99wt.% milk protein.

[0141] In some embodiment, the process does not comprise any step of addition of a milk protein-containing ingredient as provided above.

[0142] The process further comprises an optional step (e) of heat treating the plant dispersion. In some embodiment, this step is not optional.

[0143] In some embodiment, the heat treatment of step (e) may be performed at a temperature of at least 60°C, preferably of 60°C to 100°C, more preferably 80°C to 100°C. In addition, the heat treatment of the heat treatment of step (e) may be performed for a time of at least 1 minute, preferably 1 minute to 15 minutes, more preferably 5 minutes to 15 minutes.

[0144] This heat treatment step contributes to gelatinization of the starch and improve solubility of non-starch polysaccharides that would facilitate subsequent enzymatic treatment. In addition, this heat treatment contributes to inactivate endogenous enzyme in the low-refined plant protein-containing ingredient that may negatively impact the sensory properties of the final food composition.

[0145] The process further comprises an optional step (d) of adjusting the pH of the plant dispersion to a pH of 6 to 7.5, preferably to a pH of 7. In some embodiment, this step is not optional. This step (d) may be needed when the pH of plant dispersion of step (b) is different from pH of 6 to 7.5, preferably pH of 7. The pH adjustment of step (d) may be performed by adding an acidifying ingredient or adding an alkalinizing ingredient to the plant dispersion until reaching a pH of 6 to 7.5, preferably a pH of 7.

[0146] In particular, when the pH of the plant dispersion is below a pH of 6, the pH adjustment of step (d) may be performed by adding an acidifying ingredient until reaching a pH of 6 to 7.5, preferably a pH of 7.

[0147] More particularly, when the pH of the plant dispersion is below a pH of 6 to 7.5, preferably below a pH of 7, the pH adjustment of step (d) may be performed by adding an acidifying ingredient until reaching a pH of 6 to 7.5, preferably a pH of 7.

[0148] In particular, when the pH of the plant dispersion is above a pH of 7, the pH adjustment of step (d) may be performed by adding an alkalinizing ingredient until reaching a pH of 6 to 7.5, preferably a pH of 7.

[0149] In particular, when the pH of the plant dispersion is above a pH of 6 to 7.5, preferably above a pH of 7, the pH adjustment of step (d) may be performed by adding an alkalinizing ingredient until reaching a pH of 6 to 7.5, preferably a pH of 7.

[0150] The alkalinizing ingredient may be any alkalinizing ingredient suitable for food application, i.e. any alkalinizing ingredient that are edible. Examples of alkalinizing ingredient include sodium bicarbonate, potassium bicarbonate, magnesium hydroxide, calcium carbonate, potassium carbonate, sodium carbonate, ammonium carbonate, sodium citrate, potassium citrate, magnesium carbonate, sodium hydroxide, and combination thereof.

[0151] The acidifying ingredient may be any acidifying ingredient suitable for food application, i.e. any acidifying ingredient that are edible. Examples of acidifying ingredient include glucono delta-lactone (GDL), citrus juice, citric acid, lactic acid, phosphoric acid, acetic acid, tartaric acid, malic acid, fumaric acid, ascorbic acid, gluconic acid, succinic acid, hydrochloric acid, sulfuric acid, and combination thereof. Preferably, the citrus juice is lemon juice.

[0152] The step I may be before or after step (d). Preferably, the step I is before step (d).

[0153] In some embodiment, the process further comprises a step (e) of contacting the plant dispersion with an enzyme preparation to provide an enzymatically-treated arabinose- containing and galactose-containing liquid or semi-liquid food composition.

[0154] The enzyme preparation comprises at least one enzyme comprising a pectinase activity. In some embodiment, the enzyme preparation consists only of at least one enzyme comprising a pectinase activity. The pectinase activity of the at least one enzyme comprising a pectinase activity may be the main activity of the enzyme or a side activity of the enzyme, preferably main activity. In a preferred embodiment, the pectinase activity of said at least one enzyme comprising a pectinase activity comprises a-arabinofuranosidase activity and / or |3- galactanase activity. The at least one enzyme comprising a pectinase activity may be selected from the list consisting of pectinase, cellulase, hemicellulase, amylase, amyloglucosidase, endo-polygalacturonase, exo-polygalacturonase, pectate lyase, pectin lyase, arabinogalactan endo-|3-l,4-galactanase, p-galactosidase, endo-arabinase, exo-arabinase, rhamnogalacturonan lyase, rhamnogalacturonan hydrolase , rhamnogalacturonan acetylesterase, a-l-arabinofuranosidase, carbohydrate esterase, and mixture thereof. In one embodiment, the carbohydrate esterase may be selected from the list consisting of pectin methylesterase, acetylesterase, ferulic acid esterase and mixture thereof.

[0155] It has been discovered that enzymes with notable pectinase activity at neutral or nearly neutral pH levels can effectively eliminate pectic polysaccharides in compositions containing low-refined plant protein-containing ingredients. This leads to a substantial decrease in non-starch polysaccharides, including insoluble ones. As a result, the processing of these compositions is enhanced, and the properties of the final food composition prepared from said compositions are improved. Indeed, the reduction of non-starch polysaccharides, including insoluble ones minimize the challenges associated with non-starch polysaccharides, including insoluble ones.

[0156] In a preferred embodiment, the enzyme comprising a pectinase activity, when it is subjected to a method comprising step (a) to (e) according to the second aspect of the invention, has:

[0157] - a lytic activity towards pNP-a-L-arabinofuranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium of step (b), and

[0158] - a lytic activity towards pNP- -D-galactopyranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium of step (b).

[0159] The term "undiluted enzyme" has the same meaning as in the second aspect of the invention.

[0160] As mentioned in the second aspect of the invention these enzymes have been identified to effectively degrade pectic polysaccharides under neutral or nearly neutral pH.

[0161] In a preferred embodiment, the at least one enzyme comprising a pectinase activity has at least 60% sequence identity with one of the amino acid sequences SEQ ID NO: 1-3. More preferably, the at least one enzyme comprising a pectinase activity has at least 70%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% with one of the amino acid sequences SEQ ID NO: 1-3. Most preferably, the at least one enzyme comprising a pectinase activity has the amino acid sequence of any one of SEQ ID NO: 1-3.

[0162] These enzymes having sequence identity with the above amino acid sequences have been identified to effectively degrade pectic polysaccharides under neutral or nearly neutral pH.

[0163] In a preferred embodiment, the at least one enzyme comprising a pectinase activity is derived from the species Neosartorya or Aspergillus. In a more preferred embodiment, the at least one enzyme comprising a pectinase activity is derived from Neosartorya fumigata or Aspergillus niger.

[0164] It has been observed that the enzymes derived from the species Neosartorya or Aspergillus, in particular derived from Neosartorya fumigata or Aspergillus niger are advantageous for the invention as they effectively degrade pectic polysaccharides under neutral or nearly neutral pH.

[0165] The plant dispersion in step (e) has a pH from 6 to 7.5, preferably a pH of 7.

[0166] In some further embodiment, the plant dispersion is contacted with the enzyme preparation in step (e) such that the content of the enzyme comprising a pectinase activity in the plant dispersion is of at least 0.1%, preferably 0.1 to 0.7% (w / w). In particular, the content of the enzyme comprising a pectinase activity contacted with the plant dispersion in step I is of at least 0.1%, preferably 0.1 to 0.7% (w / w).

[0167] In some further embodiment, the plant dispersion may be contacted with the enzyme preparation in step (e) at a temperature of 20 to 60°C.

[0168] In some further embodiment, the plant dispersion may be contacted with the enzyme preparation in step (e) for at least 15 minutes, preferably 15 minutes to 5 hours, more preferably 15 minutes to 2 hours.

[0169] These different conditions ensure optimal degradation of pectic polysaccharides in the plant dispersion by the enzyme comprising pectinase activity.

[0170] The step (e) effectively degrades of the pectic polysaccharides and result in the release of free arabinose and free galactose. The amount of free arabinose by dry weight is increased by at least 0.25wt.%, preferably at least 0.4wt.% in the enzymatically-treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

[0171] In some embodiment, the amount of free arabinose by dry weight is increased by at most 5wt.%, preferably at most 3wt.%, more preferably at most 1.5wt.% in the enzymatically- treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

[0172] The amount of free galactose by dry weight is increased by at least 0.3wt.%, preferably at least 0.7wt.%, more preferably at least 1.5wt.% in the enzymatically-treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

[0173] In some embodiment, the amount of free galactose by dry weight is increased by at most 5wt.%, preferably at most 4wt.%, more preferably at most 3wt.% in the enzymatically- treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

[0174] In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (e) may have a weight ratio of free arabinose to free galactose (free arabinose: free galactose) of from 1:1.5 to 1:5, preferably 1:3.5 to 1:4.5.

[0175] In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (e) may comprise less than 20wt.%, preferably less than 15wt.%, more preferably less than 10wt.%, even more preferably less than 5wt.%, even more preferably less than lwt.% pectic polysaccharides by dry weight.

[0176] In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (e) may comprise less than 30wt.%, preferably less than 15wt.%, more preferably less than 10wt.%, even more preferably less than 5wt.%, even more preferably less than 3wt.%, even more preferably lwt.% starch by dry weight. In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (d) may be free from starch. Low starch content may be achieved in the presence of a starch-degrading enzyme in step I.

[0177] In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (e) may comprise less than 30wt%, preferably less than 20wt.%, more preferably 1 to 10wt.%, even more preferably 5 to 10wt.% non-starch polysaccharides by dry weight. In some embodiment, the enzymatically-treated arabinose and galactose-containing food composition obtained after step (e) may comprise less than 25wt%, preferably less than 20wt.%, more preferably 2wt.% to 20wt.%, even more preferably 5wt.% to 10wt.% insoluble non-starch polysaccharides by dry weight.

[0178] In an advantageous embodiment, the plant dispersion is further contacted in step (e) or is contacted before step (e) with at least one starch-degrading enzyme and / or at least one cellulose-degrading enzyme and / or at least one protein-degrading enzyme. In a more advantageous embodiment, the plant dispersion is further contacted in step (e) or is contacted before step (e) with at least one starch-degrading enzyme and / or at least one cellulose-degrading enzyme.

[0179] While not being bound by theory, it is believed that the inclusion of a starch-degrading enzyme and / or cellulose-degrading enzyme and / or protein-degrading enzyme facilitates the release of non-starch polysaccharides that are trapped and hardly accessible within the complex polysaccharide structure of low-refined plant protein-containing ingredients. This enzymatic action further helps to expose and make accessible the pectic polysaccharides, allowing the pectinase activity of the enzyme to hydrolyze them. Starch-degrading enzyme and / or cellulose-degrading enzyme are preferable to protein-degrading enzyme because protein-degrading enzyme may lead to the formation of off-notes.

[0180] The starch-degrading enzyme may be selected from the list consisting of a-amylase, |3- amylase, glucoamylase, amyloglucosidase, pullulanase, isoamylase, cyclodextrin glucanotransferase, transglucosidase, dextrinase, limit dextrinase, and mixture thereof.

[0181] The cellulose-degrading enzyme may be selected from the list consisting of cellulase, cellobiohydrolase, endoglucanase, exoglucanase, p-glucosidase and mixture thereof.

[0182] The protein-degrading enzyme may be selected from the list consisting of endoprotease, exo-protease, endo-peptidase, exo-peptidase, and mixture thereof.

[0183] In a preferred embodiment, the starch-degrading enzyme is different than the enzyme comprising pectinase activity.

[0184] In a preferred embodiment, the cellulose-degrading enzyme is different than the enzyme comprising pectinase activity.

[0185] In a preferred embodiment, the protein-degrading enzyme is different than the enzyme comprising pectinase activity.

[0186] When the plant dispersion is contacted with the at least one starch-degrading enzyme and / or at least one cellulose-degrading enzyme and / or at least one protein-degrading enzyme during step (e), the at least one enzyme comprising pectinase activity and the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme may be added together or separately. When the plant dispersion is contacted with the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme during step (e), the enzyme preparation of step (e) may comprise at least one enzyme comprising pectinase activity and the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme. In this case, the enzyme comprising pectinase activity and the starch-degrading enzyme and / or the cellulose-degrading enzyme and / or the protein-degrading enzyme are added together in step (e) as they are part of the same ingredient.

[0187] When the the plant dispersion is contacted with the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one proteindegrading enzyme during step (e) or before step (e), the enzymatic reactions are performed under the same conditions, i.e. time, temperature and pH, as the conditions provided above for the enzymatic reaction with the enzyme comprising pectinase activity.

[0188] When the the plant dispersion is contacted with the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one proteindegrading enzyme during step (e) or before step (e), the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme is / are contacted with the plant dispersion at the following content in the plant dispersion : 0.01-2.0wt.%, preferably 0.02-1.0wt.%, more preferably 0.05-0.2wt.% by dry weight.

[0189] When the plant dispersion is contacted with the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme before step (e), this step of contacting the plant dispersion with the at least one starch-degrading enzyme and / or the at least one cellulose-degrading enzyme and / or the at least one protein-degrading enzyme may be performed after step (b) or step (c) or step (d).

[0190] The process further comprises a step (f) of heat-treating the enzymatically-treated arabinose-containing and galactose-containing liquid or semi-liquid food composition to inactivate the at least one enzyme comprising a pectinase activity and to form an arabinose- containing liquid or semi-liquid food composition. This heat treatment step may also inactivate enzymes other than enzyme comprising a pectinase activity (if any), such as starch- degrading enzymes and cellulose-degrading enzymes and protein-degrading enzymes.

[0191] The heat treatment of step (f) may be performed at a temperature of at least 75°C, preferably of at least 80°C, more preferably at least 90°C. In addition, the heat treatment of step (f) is performed at a temperature of at most 140°C, preferably at most 135°C, more preferably at most 125°C. In addition, the heat treatment of the heat treatment of step (f) may be performed for a time of at least 3 seconds, preferably 3 seconds to 15 minutes, more preferably 3 seconds to 90 seconds.

[0192] The heat treatment step (f) can also serve the purpose of eliminating undesired microorganisms and extending the shelf life of the food composition. In other words, this heat treatment can also act as a pasteurization or sterilization, ensuring the safety and stability of the food product. As a result, the shelf life of the food composition can be extended from few days to weeks / months, allowing for a longer duration of storage and consumption without compromising its quality or safety.

[0193] In some embodiment, the process may further comprise a step of homogenizing the enzymatically-treated arabinose-containing and galactose-containing liquid or semi-liquid food composition before step (f) and after step (e) and / or a step of homogenizing the arabinose-containing liquid or semi-liquid food composition after step (f) and before step (g).

[0194] The homogenization step(s) described just above may be performed at a pressure above 50 bar. Preferably, the homogenizing step may be performed at a pressure of 50 bar to 700 bar. Further preferably, the homogenizing step may be performed at a pressure of 50 bar to 500 bar. More preferably, the homogenizing step may be performed at a pressure of 50 to 300 bar, from 100 to 300 bar or from 150 to 300 bar.

[0195] The process may comprise the optional step (g) of drying of the arabinose-containing and galactose-containing liquid or semi-liquid food composition to form an arabinose- containing and galactose-containing powdered food composition. The drying step (g) may be performed by any well-known drying method for food products. For example, the drying step (g) may be performed by spray drying, vacuum band drying, roller drying or freeze drying.

[0196] In some embodiment, the process may comprise a step of evaporating and / or concentrating the arabinose-containing liquid or semi-liquid food composition before step (g). When a homogenization step is applied after step (f), the step of evaporation and / or concentration is after said homogenization step. In some embodiment, the process may further comprise a step of decantation and / or filtration to remove remaining pectic polysaccharides after step (f). If any step (g) of drying, the step of decantation and / or filtration is before step (g) of drying. This particular step can result in a food composition which is free from pectic polysaccharides. In some embodiment, the process does not comprise any step of decantation and / or filtration to remove pectic polysaccharides. SEQUENCES

[0197] Those skilled in the art will understand that they can freely combine all features of the present invention disclosed herein. In particular, features described for the composition of the present invention may be combined with the method and process of the present invention and vice versa. Further, features described for different embodiments of the present invention may be combined.

[0198] Furthermore, where known equivalents exist to specific features, such equivalents are incorporated as if specifically referred in this specification. Further advantages and features of the present invention are apparent from the figures and non-limiting examples.

[0199] EXAMPLES

[0200] Example 1: Methods for the determination of the carbohydrate composition

[0201] Total sugar analysis Total sugar analysis of flours and fractions thereof was performed based on the methanolysis method by Quemener & Thibault (1990). In brief, samples were weighed in 5 ml screw-top V- Vials® (Merck, Darmstadt, Germany) and dried under vacuum in the presence of phosphorus pentoxide (P2O5) at 40°C for 16 h. Whilst being stirred, samples were dissolved to 5 mg ml1in anhydrous methanol (CH3OH) containing 2 M HCI. The tubes were capped and incubated under continuous stirring at 80°C for 16 h. Samples were cooled and evaporated under a draft of N2. To each tube, 2 ml of 2 M Trifluoroacetic acid (CF3COOH) was added, after which the tubes were capped and incubated under continuous stirring at 121°C for 1 h. The tubes were cooled, evaporated under a draft of N2 and redissolved in 25 ml of MilliQ water (Merck Millipore, Burlington, MA, USA). The monosaccharide composition of the samples was analyzed using a DionexTM ICS-6000 from Thermo Fisher Scientific (Waltham, MA, USA) using a CarboPacTM PA-1 (2 X 250 mm) column linked to an electrochemical detector with an Ag / AgCI reference electrode. Samples were injected into the system at a volume of 10 pl. A gradient elution was applied. The flowrate was 0.25 ml min-1. The following eluents were used: Solvent A: MilliQ water, Solvent B: 300 mM NaOH, Solvent C: 150 mM NaOH and 500 mM NaOAc, and Solvent D: NaOH 100 mM. For the post column, 300 mM NaOH was used at a flowrate of 0.15 ml min-1. Calibration standards were comprised of 1-20 pg ml1fucose, rhamnose, xylose and mannose, 1.5-60 pg ml1arabinose and galacturonic acid, and 1-40 pg ml1galactose, glucose, and glucuronic acid.

[0202] Free sugar analysis

[0203] To determine the free sugar content, l g of dried sample was extracted using in 60 mL of MilliQ water at 70°C for 30 min under mild stirring. After cooling, the volume was adjusted at 100 mL. Prior to analysis, the obtained solutions were diluted according to the calibration standards.

[0204] The free sugars were quantified by High Performance Anion Exchange Chromatography coupled to a Pulsed Amperometric Detection (HPAEC-PAD) using an Ion Chromatograph ICS- 6000 (Thermo Fisher Scientific, SARL, Ecublens, Switzerland) equipped with a CarboPac™ PA20 column (3 x 150 mm, 6.5 pm). The system was operated at a flow rate of 0.5 mL min1with MilliQ water, 300 mM NaOH and 150 mM NaOH containing 500 mM NaOAc by applying the following gradients: Starting from 98% MilliQ water and 2% 300 mM NaOH, the concentration was gradually increased to 5% 300 mM NaOH over a period of 12 min; afterwards the gradient was altered towards 46% MilliQ water, 34% 300 mM NaOH and 20% of 150 mM NaOH containing 500 mM NaOAc over a period of 15 min; then, a gradient of 100% 150 mM NaOH containing 500 mM NaOAc was applied for 5 min followed by a gradient of 100% 300 mM NaOH for 5 min; finally, the column was re-equilibrated using the starting gradient of 98% MilliQ water and 2% 300 mM NaOH for 5 min. For quantification, the standards glucose, sucrose, fructose, maltose (Sigma-Aldrich Chemie GmbH, Buchs, Switzerland) and leucrose (Biosynth Carbosynth® Berkshire, UK) were used at a concentrations range between 1 and 50 pg mL-1. The injection volume was 25 pL. The sugars content in g 100 g1(w / w) was calculated based on the prepared standard solutions.

[0205] Starch Analysis

[0206] The starch content of pea flour, de-starched pea flour and fractions thereof were analysed using the Total Starch Assay kit from Megazyme (Wicklow, Ireland) according to method A in the supplier's handbook. In brief, the samples were weighted into tubes (16 x 120 mm) and dissolved in 10 ml of 100 mM NaOAc buffer (pH 5) containing 5 mM CaCL The sample preparation was performed in duplicate. To one sample, 1% v / v of thermostable a-amylase was added and to the other one 1% v / v of 100 mM NaOAc buffer (pH 5) containing 5 mM CaCl2. All tubes were immediately transferred to a 95°C water bath for 15 min. The samples were vortexed after 2, 5, and 10 min during the heating step. After boiling, the samples were transferred to a heating plate which was set to 50°C. Samples were cooled to 50°C under continuous stirring. To the a-amylase treated sample, 1% v / v of amyloglucosidase was added to the other one 1% v / v of 100 mM NaOAc buffer (pH 5) containing 5 mM CaCL Samples were incubated at 50°C for 30 min. Afterwards, the samples were cooled to room temperature. A volume of 2 ml of each treated sample was transferred into a 2 ml Eppendorf tube and centrifuged at 20,817 x g for 5 min. From each sample, 0.1 ml of aliquot was transferred to tubes (16 x 120 mm) containing 3 ml of GOPOD reagent. This step was performed in duplicate. The above-described incubation with GOPOD reagent was repeated using 100 mM NaOAc buffer (pH 5) containing 5 mM CaCl2 as a reagent blank and a 1 mg ml1glucose solution in quadruplicate (glucose control). All tubes were incubated at 50°C for 20 min. Within 60 min after the last incubation, the absorbance of the samples was measured at 510 nm against the reagent blank using an Agilent HP 8453 spectrophotometer (Santa Clara, CA, USA). If the obtained absorbance value was higher than the glucose control, the sample was diluted 5 times in 100 mM NaOAc buffer (pH 5) containing 5 mM CaCl2 before performing the GOPOD reaction step. Total starch content (% (w / w)) was determined according to the supplier's handbook using the following formula: Starch (%(w / w)) = AA*F*10.2*D / W*0.9; wherein AA is the absorbance value, F the GOPOD conversion factor (100 / average glucose control absorbance), D the Dilution factor, and W the Sample weight in mg.

[0207] Example 2 Method for the determination of the protein content

[0208] Protein determination by Kjeldahl

[0209] The total N2 content of the different flours, concentrates and all obtained fractions after processing was determined by an automated Kjeldahl Method using Buchi KjelDigest automat K-449 which was connected to a Buchi Scrubber B-415 (BUCHI Labortechnik AG, Flawil, Switzerland). Samples were weighed in a Kjeldahl weighing boat at 100-200 mg per sample, if the expected N2 content was >7%, otherwise the sample weight ranged from 500- 1.000 mg. Samples were placed into a mineralization vessel, to which 10 ml of concentrated sulfuric acid and 1 tablet of Missouri catalyst (4.98 g K2SO4 + 0.02 g CuSC x 5 H2O) was added. The mineralization vessels were placed Into the mineralization device and empty spaces were closed with small closing tubes. Mineralization was done according to the following steps: 1, heating to 300 °C; 2, heating to 350°C for 15 min; 3, heating to 420°C for 105 min; 4, cooling for 35 min. After cooling down, samples were immediately distilled in a Buchi Kjelmaster unit K-375. Titration was performed using 0.1 M HCI until reaching pH 4.65. Total N2 (TN) content was determined using the Formula below and expressed in g / 100 g product. Conversion to protein was done by multiplying the TN by factor 6.25. g V * 0.1 * 14 * 100 1000

[0210] V: Volume of 0.1 M HCI used m: Sample weight in g

[0211] Example 3: Method for the preparation of pea flour extracts

[0212] De-starching of pea flour and fraction thereof

[0213] A 0.1 g ml1solution of yellow pea flour in water was prepared in a Thermomix® TM6 (Vorwerk International & Co. KmG, Wollerau, Switzerland) and heat-treated under continuous stirring (Stirring Speed 3) at 95°C for 10 min. The mixture was cooled down to 50°C, after which 1% w / w a-amylase and 1% w / w glucoamylase was added. The mixture was kept under continuous stirring at 50°C for 2 h. Afterwards, the mixture was heat-treated under continuous stirring at 95°C for 10 min and then cooled down to room temperature. The mixture was dialysed using + / - 80 cm long cellulose dialysis bags from SpectrumTM Spectra / Por® (Repligen, Massachusetts, United States) with a diameter of 76 mm and a 6-8 kDa cut-off. Dialysis was performed in MilliQ water under continuous stirring using a magnetic stirrer at 4°C for four days. MilliQ water was exchanged twice a day during this period. After dialysis, the destarched pea flour (DPF) was freeze dried using a Telstar® (Barcelona, Spain) LyoQuest freeze dryer.

[0214] Chemical extraction ofde-starched pea flour (DPF)

[0215] In total, four different extraction procedures were applied to isolate pectic polysaccharides from DPF, which included EtOH, water, acid and alkaline extraction. The amount of DPF as a starting material for each fractionation and mass balance is presented in Figure 1. EtOH extraction - A 96% EtOH suspension containing 23 mg ml1DPF was solubilized under continuous stirring at 70°C for 30 min to obtain the alcohol insoluble solids (AIS) fraction. Water extraction - A concentration of 6.67 mg ml1of DPF was dissolved in MilliQ and placed in an 80°C water bath for 30 min under continuous stirring to obtain the hot water extract. Alkaline Extraction - DPF was dissolved in 50 mM NaOH (pH 12.3) yielding to a concentration of 10 mg ml1under continuous stirring at 4°C for 17 h to obtain the alkaline extract. Acid Extraction - A concentration of 10 mg ml1of DPF was dissolved in MilliQ at a set pH of 2.5 using 1 M HCL. The extraction was performed at 80°C under continuous stirring for 1 h to obtain the acid extract.

[0216] To obtain the AIS of the latter two extracts, 96% EtOH was added to the alkaline and acid extraction yielding to a final concentration of 70% EtOH. Precipitation of pectic polysaccharides and insoluble fibres in the alkaline and acid extract was done overnight. Samples were centrifuged at 24,470 x g at 4°C for 15 min. The supernatant was collected, and the pellet was washed another 3-times using 96% EtOH. The pellets of the EtOH, alkaline, and acid extracts were dried in the fume hood to evaporate the remaining EtOH. To dry the 96% EtOH extract supernatants, the supernatants were diluted with MilliQ water to obtain a 70% EtOH concentration. Afterwards, the supernatant of the EtOH, alkaline, and acid extract were concentrated using a Buchi rotor evaporator R-210 which was equipped with a Buchi vacuum controller V-850, a Buchi heating bath B-491, and a LKB Bromma 2,219 Multitemp II Thermostatic Circulator with a cooling solution. The water bath was set to 40°C and a vacuum of 72 mbar was applied until the supernatants were concentrated to about 25% of the starting volume. The water of the hot water extract supernatant was concentrated using a rotor evaporator in a water bath temperature of 60°C and the vacuum was slowly turned down to 86 mbar until the supernatant was concentrated to about 30% of the starting volume. All samples were freeze dried using a Telstar® (Barcelona, Spain) LyoQuest, weighted and stored at room temperature.

[0217] Yields and composition of the de-starched pea flour and fractions thereof

[0218] A chemical extraction was performed on destarched pea flour by EtOH, hot water, alkaline and acid extraction. There was no centrifugation step before the EtOH precipitation during the alkaline and acidic extraction procedure. Thus, the insoluble fraction including the extracted pectin fraction were both precipitated in 70% EtOH. The latter means that the pellet did not only consist of precipitated pectic polysaccharides, but also the other remaining insoluble fibres as the extracted pectic polysaccharides were not separated. Therefore, for the alkaline and acid extract AIS, this fraction refers to a combination of acid and alkaline soluble pectic polysaccharides and insoluble fibres, including as well pectic polysaccharides. To investigate the impact of the chemical extraction procedures on the composition of the obtained fibre fractions, the monosaccharide content was determined (cf. Table l).ln particular, it can be observed in table 1 in the water pellet fraction that corresponds to the insoluble compounds, it comprises 70% of pectic polysaccharides, including arabinose, galactose, rhamnose and uronic acid. Hence, it appears that the insoluble fraction of nonstarch polysaccharides is abundant in pectic polysaccharides.

[0219] Table 1: Total monosaccharide composition of pea flour, insoluble fraction found in current product, and the chemical treatments of destarched pea flour*.

[0220] *The exact carbohydrate fraction of the specific sample was not known, therefore the carbohydrate fraction as given by the manufacturer is used to give estimations %w / dw of the total sample.

[0221] AIS=Alcohol insoluble solids. The monosaccharide composition was determined by Methanolysis using TFA and the quantity was determined using HPAEC-PAD (Example 1). The value without brackets gives % w / w based on the total sample while the value between brackets indicates the % w / w based only on the total amount of carbohydrates.

[0222] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Man, Mannose; Gal Ac, Galacturonic Acid; Glc Ac, Glucuronic Acid

[0223] Example 4 Method for selecting a pectinase

[0224] Activity screening using pNP-substrates

[0225] Enzyme activities towards para-nitrophenyl (pNP)-substrates were measured using a spectrophotometric assay which was supported by a semi-automatic Gallery™ Plus system (Thermo Fisher Scientific). Quantification was based on the release of pNP from pNP-a-L- arabinofuranoside, pNP-|3-D-glucopyranoside, pNP-|3-D-galactopyranoside, and pNP-a-L- rhamnopyranosidel. From each pNP-substrate, a stock solution of 240 mM pNP-substrate in DMSO was prepared and stored in a dark place at 4°C. Each stock solution was freshly diluted to 2.4 mM in 0.1 M Bis-Tris buffer (pH 7.0) using a 10 ml volumetric flask 1 h prior to use. Enzymes were freshly dissolved at a concentration of 50 mg ml1in 0.1 M Bis-Tris buffer (pH 7.0) using a volumetric flask on the day of use. The enzyme solutions were kept on ice water during the day of use. For powdered enzymes, the 50 mg ml1Bis-Tris solutions were placed in a Branson 5210 Sonicator (Branson, CT, USA) for 15 min and 2 ml aliquots were centrifuged at 14000 x g for 2 min. The supernatants of the aliquots were used for the enzyme activity screening.

[0226] The enzyme activity screening was performed using a Gallery™ Plus and a reaction temperature of 37°C. All enzyme solutions were put in 1.5 ml cuvettes and placed in the sample rack of the Gallery™ Plus. The 2.4 mM pNP-substrate solution was put in a 20 ml reagent tube and placed in the reagent rack of the Gallery™ Plus. Enzyme activities were screened for non-diluted, 10 times, and 100 times diluted enzyme solutions. Twelve measurements were taken every 1.05 min until the last measurement at 11.55 min. For the calibration curve, a stock solution of 1.2 M pNP in DMSO was freshly prepared on the day of use. The stock solution was diluted to 12 mM in 0.1 M Bis-Tris buffer (pH 7.0) using a volumetric flask. The 12 mM pNP solution was put into a 1.5 ml cuvette, which was placed in the sample rack of the Gallery™ Plus. The absorption was measured photometrically at a wavelength of 405 nm. A tube containing 0.1 M Bis-Tris (pH 7.0) was placed in the reagent rack to use for the dilutions in the calibration curve. A new calibration curve was prepared prior to each measurement. The enzyme activity was calculated by converting the absorbance values of the samples to molarity by using the calibration curve and multiplying it with the dilution factor if applicable (pM min1). Considering the enzyme concentration, the corresponding enzyme activity in pmol / min / g (or U / g) was determined.

[0227] Polygalacturonase (PG) activity screening using the DNS reducing end assay

[0228] The polygalacturonase activity that was present in the enzyme preparations was determined using the reducing end assay. Therefore, a DNS reagent solution consisting of 0.06 M 3,5- dinitrosalicylic acid (DNS), 0.05 M potassium sodium tartrate tetrahydrate solution in 0.5 M NaOH, was prepared. As substrate, a 5.0 mg ml1polygalacturonic acid solution in 0.1 M Bis- Tris buffer (pH 7.0) was prepared. To ensure microbial safety, 0.01% w / v of sodium azide was added to the substrate solution. For each enzyme preparation, a 50 mg ml1solution in 0.1 M Bis-Tris buffer (pH 7.0) was prepared. 112.5 pl of the substrate solution was put into six 2 ml tubes, which were placed in an Eppendorf ThermoMixer® (Eppendorf SE, Hamburg, Germany) which was set to 40°C. To each sample, 12.5 pl of enzyme solution was added and the samples were incubated for exactly 2, 4, 6, 8, 10, or 12 min under stirring. After the incubation time, 375 pl of DNS was added to the samples and all samples were immediately heat-treated in a water bath at 95°C for 10 min. After the heat treatment, samples were directly placed on ice water. To each sample, 1 ml of MilliQ water was added and the samples were poured into 1.5 ml cuvettes. The absorbance was determined at 540 nm using a Gallery™ Plus system.

[0229] For the calibration, a standard stock solution of 10 mM D-(+)-galacturonic acid monohydrate (>97% purity) was prepared. The stock solution was diluted to a concentration of 1, 2, 3, 4, 5, 6, 7, 8, and 9 mM. A 1:3 solution of standard stock solutio DNS reagent was prepared in 2 ml Eppendorf tubes for each calibration curve dilution. The tubes were heated for 10 min at 95°C and then stored on ice water. The samples were diluted 3 times in MilliQ water and poured into separate 1.5 ml cuvettes. The cuvettes were placed into the sample rack of the Gallery™ Plus. A new calibration curve was prepared on each measuring day. The enzyme activity was quantified based on the calibration curve of each standard and expressed as pmol min1g1(or U g-1).

[0230] The results of the enzyme activities towards pNP-a-L-arabinofuranoside, pNP-6-D- glucopyranoside, pNP-6-D-galactopyranoside, pNP-a-L-rham nopyranoside, and polygalacturonic acid are summarized in Table 2.

[0231] Table 2: Enzyme activities towards pNP-substrates and polygalacturonic acid of different enzyme preparations.

[0232] Example 5 Expression of pectinases Pichia pastoris transformation

[0233] The protein sequences of various pectinases were reverse-translated and codon-optimized for optimal expression in Pichia pastoris. Subsequently, gene fragments were ordered from Twist Bioscience (San Francisco, CA, USA). Employing a commercially available Pichia pastoris strain as the expression host, plasmids were linearized using Sac I HF (NEB, Ipswich, MA, USA) according to NEB's protocol. Prior to transformation, chemically competent Pichia pastoris cells were prepared. Following this preparation, these cells underwent transformation using the Frozen-EZ Yeast Transformation II™ Kit (Zymo Research, Murphys, CA, USA) and were selected on YPD plates containing 100 pg ml1Zeocin® (Invitrogen, Thermo Fisher Scientific, Dreieich, Germany). Subsequently, an overnight culture originating from a single yeast colony was inoculated in BMGY medium. Positive transformants (500 pl) were mixed with 500 pl of 30% (w / v) glycerol to create 15% (w / v) glycerol stocks, which were then stored at -80 °C. Recombinant expression of pectinases in Pichia pastoris in shake flasks

[0234] For the recombinant expression of pectinases in shake flasks, Pichia pastoris transformants were cultured on YPD plates (10 g 11yeast extract, 20 g 11peptone, 20 g 11glucose, 20 g 11agar-agar) and incubated at 30 °C for three to four days. BMGY medium (10 g 11yeast extract, 20 g 11peptone, 100 mM potassium phosphate pH 6.5, 1.34% (w / v) yeast nitrogen base, 4 x 10-5% (w / v) biotin, 2.0% (v / v) glycerol) was utilized for both pre- and main cultures. The main culture (400 ml working solution) was inoculated in a 2 L shake flask with 5% (v / v) of the preculture and shaken at 28°C at 110 rpm for two days without the addition of methanol. Subsequently, 0.5% (v / v) methanol was introduced to induce protein expression via the AOX1 promoter. Methanol addition (0.5% (v / v)) was repeated every 24 hours. After 72 hours, yeast cells were harvested through centrifugation at 13,400 x g at 4 °C for 10 minutes, and the supernatant was sterile filtered (0.22 pm, TTP®, Trasadingen, Switzerland).

[0235] The following sequences were used and expressed in P. pastoris (the signal peptide is underlined): a-L-arabinofuranosidase B from Neosartorya fumigate CBS 101355 (Uniprot ID: Q4WL66);

[0236] 52,5 kDa based on the amino acid sequence (SEQ ID NO: 1)

[0237] MLPQLSIERASVFALGLIATGSLVVAGPCDIYSAGGTPCVAAHSTTRALYSSYSGPLYQVKRGSDGATADIA PLSAGGVANAAAQDSFCDGTTCLITIIYDQSGRGNHLTQAPPGGFSGPESNGYDNLASAIGAPVTLNGQK AYGVFISPGTGYRNNAASGTATGDAPEGMYAVLDGTHYNDACCFDYGNAETSSRDTGNGHMEAIYFG DNTIWGTGSGSGPWIMADLENGLFSGSSPDNNSGDPSISYRFLTAVVKGKQNQWAIRGANAASGSLST FYNGARPSVSGYNPMSKEGAIILGIGGDNSNGAQGTFYEGVMTSGYPSDATENSVQANIVAAKYATASL TSGPKLTVGSSISLQATTPGYTTRYIAHSGSTVNTQVVSSSSSTTLKQQASWTVRTGLANSDCFSFESRDTP GSFLRHYNFVLQLSANDGTKQFHEDATFCPQAGLNGQGNSIRSWNYPTRYFRHYNNVLYAASNGGVHT FDATSSFNNDVSWVISTGFA

[0238] 0-galactosidase A from Neosartorya fumigata CBS 101355 (Uniprot ID: Q4WS33); 110,4 kDa based on the amino acid sequence (SEQ ID NO: 2) MKLLSVCAIALLAAQAAGASIKHMLNGFTLMEHSDPAKRELLQKYVTWDEKSLFVNGERIMIFSGEVHPF RLPVPSLWLDVFQKIKALGFNCVSFYVDWALLEGKPGEYRAEGNFALEPFFDVAKQAGIYLLARPGPYINA EASGGGFPGWLQRVNGTLRTSDPAYLKATDNYIAHVAATIAKGQITNGGPVILYQPENEYSGACCDATFP DGDYMQYVIDQARNAGIVVPLINNDAWTGGHNAPGTGKGEVDIYGHDSYPLGFDCGHPSVWPKGNLP TTFRTDHLKQSPTTPYSLIEFQAGSFDPWGGPGFAACAALVNHEFERVFYKNDLSFGAAILNLYMTFGGT NWGNLGHPGGYTSYDYGSPLTESRNVTREKYSELKLIGNFVKASPSYLLATPGNLTTSGYADTADLTVTPLL GNGTGSYFVVRHTDYTSQASTPYKLSLPTSAGRLTVPQLGGTLTLNGRDSKIHVVDYNVAGTNIIYSTAEVF TWKNFGDSKVLILYGGPGEHHELAVSLKSDVQVVEGSNSEFKSKKVGDVVVVAWDVSPSRRIVQIGDLKI FLLDRNSVYNYWVPQLDKDDSSTGYSSEKTTASSIIVKAGYLVRTAYTKGSGLYLTADFNATTPVEVIGAPS NVRNLYINGEKTQFKTDKNGIWSTEVKYSAPKIKLPSMKDLDWKYLDTLQEVQSTYDDSAWPAADLDTT PNTLRPLTTPKSLYSSDYGFHTGYLIYRGHFVADGSETTFDVRTQGGSAFGSSVWLNESFLGSWTGLNAN ADYNSTYKLPQVEQGKNYVLTILIDTMGLNENWVVGTDEMKNPRGILSYKLSGRDASAITWKLTGNLGG EDYQDKIRGPLNEGGLYAERQGFHQPQPPSQKWKSASPLDGLSKPGIGFYTAQFDLDIPSGWDVPLYFN FGNSTKSAYRVQLYVNGYQYGKFVSNIGPQTSFPVPQGILNYQGTNWVALTLWALESDGAKLDDFELVN TTPVMTALSKIRPSKQPNYRQRKGAY

[0239] Arabinogalactan endo- - 1,4-galactanase A from Neosartorya fumigata CBS 101355 (Uniprot ID: Q4WJ80); 39,1 kDa based on the amino acid sequence (SEQ ID NO: 3) MLGKTVLLPLLVLLCHSLASASLVYRGADISSLLI EEKAGI EYKN VNGQTQPLENI LKANGVNSVRQRVWV NPSDGSYNLDYNVKLAKRVKAAGMSVYLDLHFSDTWADPSHQTTPRGWSTNDIGTLTWQVYNYTMEV CNTFASNGIDVSIVAIGNEIRNGLLWPLGKPDNYANIANILHSAAFGVKDSTLSPKPKIMIHLDNGWDWS AQKFFYNRVLSSGANLVKSDFDLIGVSYYPFYNPSATLSALTTSLKNLRSTYGKDVLVVETDWPVSCPNPAY

[0240] AFPSDLKDIPFSVAGQTTFVQRVANIVAQTPGGIGLYYWEPAWVQNAALGSSCADNLMVDWSTRQAR TSLSVFATI

[0241] Arabinan enc / o-l,5-a-L-arabinosidase C from Neosartorya fumigata CBS 101355 (Uniprot ID:

[0242] Q4W930); 35,1 kDa based on the amino acid sequence (SEQ ID NO: 4)

[0243] MYLYTLILLFLASANVNAYANPGACSGNCWTHDPGLYQRKSDGKYFRFATGGGIHIASADSLEGPWTDD GYVLPSGSIIDLDGKTNLWAPDLHYHDGTYYLYYAVSSLGSQNSATGVATSKTMEAGSWTDHGTTGIEST PSSPYNTIDANWIAVGGTQYVNFGSYWNNLFQVEMENGLKVKSGATPHQIAYNASGIHRQEAAFMFER NNYFYLTFSGGIALGYNDTWPAPGEEYFIAVCRSTSATGGFVDKNGVSCLNSGGSLLLSSHDFVYGPGGQ GILQDSSKGFVLYYHYADTRIGKAVEDYQFGWNQLKWENDWPSV

[0244] Example 5 Determination of the arabinose and galactose release from pea flour and destarched pea flour using selected enzymes and pectinase preparations.

[0245] Pea flour and destarched pea flour was incubated with various carbohydrate-degrading enzyme preparations before and after a protease treatment (Alcalase® from Sigma Aldrich). Therefore, a 5 mg ml1(destarched) pea flour suspension in MilliQ water was incubated with or without a 1% (w / w) alcalase® (=protease) preparation at 50°C for 4 h. After inactivation at 95°C for 10 min, the 5 mg ml1(destarched) pea flour suspension (with and without Alcalase® pre-treatment) were incubated with a cellulase preparation (Enzyme Preparation 25; 2% (w / w)) and a mixture of three pectinase preparations (Enzyme Preparation 6, 15, 16; 2% (w / w) per preparation) at 20°C for 21.5 h. After heat-inactivation at 95°C for 10 min, the suspensions were incubated with a p-glucosidase (1% (w / w)) at 55°C for 1 h, followed by another heattreatment to deactivate the enzymes. The p-glucosidase had an activity of 10 U g1.

[0246] The results are shown in figures 2 and 3. About 30% of the total arabinose content was released from pea flour after the enzymatic treatment (Figure 2a and 2b). In comparison, 30 to 60% of the total arabinose was released if destarched pea flour was incubated with the selected enzymes. The incubation of pea flour with the enzymes let to a release of about 30 - 60% of the total galactose present in the substrate. The release of arabinose and galactose from pea flour and destarched pea flour incubated with carbohydrate-degrading enzyme preparations was slightly higher after a pre-incubation with an Alcalase® compared to a preincubation without this protease (Figure 3).

[0247] Example 6 Method for the determination of arabinose and galactose released from neutral side chains pea flour extracts using selected enzymes and pectinase preparations.

[0248] To determine the effectiveness of the expressed enzymes towards neutral side chains of pea flour extracts, the release of arabinose and galactose was quantified. First, alkaline extract AIS, acid extract AIS and water extract supernatant were added at a concentration of 5 mg ml1to MilliQ water. Afterwards, the expressed a-L-arabinofuranosidase B (SEQ ID NO: 1), arabinan endo-l,5-a-L-arabinosidase C (SEQ ID NO: 4), p-galactosidase A (SEQ ID NO: 2) or arabinogalactan endo-|3-l,4-galactanase A (SEQ ID NO: 3) were added (2,5% v / v enzyme solution per substate preparation) and samples were incubated at 40°C for 24 h. The reaction was stopped by incubating the samples at 95°C for 15 min using a water bath. After centrifugation (4°C, 20817 x g for 15 min), the amount of free galactose and free arabinose was measured using HPAEC-PAD (Example 1). As a second step, 0.5 mL of the supernatant was transferred into a new vial and incubated for another time using a pectinase (Preparation 16) at a concentration of 1% v / v. The incubation was performed at 40°C for 24 h. The reaction was stopped by incubating the samples at 95°C for 15 min using a water bath. The resulting supernatant was analyzed by HPAEC-PAD.

[0249] The results are shown in Figure 4.

[0250] Except for the incubation of the alkaline extract AIS, the incubation of the acid extract AIS and the water extract supernatant with a-L-arabinofuranosidase B led to an almost complete free arabinose release of 12.7% (w / w) and 2.9% (w / w), respectively (Figure 4a - c). When compared to the amount determined by the total sugar analysis, even a surplus of 1.73% for the acid extract AIS and 0.68% for the water extract supernatant of the arabinose in the substrates was measured in HPAEC. A possible reason is that the TFA hydrolysis used to determine the total sugar composition of the corresponding fraction could have led to an underestimation of monosaccharides present in the fractions (Quemener and Thibault, 1990). Arabinose was only determined for the incubation of the pea fractions that were incubated with a-L-arabinofuranosidase B, whereas the use of the expressed Arabinan endo-l,5-a-L- arabinosidase C, P-Galactosidase A, Arabinogalactan endo-|3-l,4-galactanase A did not lead to a release of this monosaccharide (Figure 4a - c).

[0251] The incubation of the supernatants, which were obtained after the first enzymatic treatment using expressed enzymes, with a pectinase (Preparation 16) led to a release of arabinose in all fractions including the blank, which indicated that arabinose-rich oligomers, like arabinogalactans, were present in the soluble supernatant of all samples (Figure 4a - c). As mentioned above, only the treatment of the fraction with a-L-arabinofuranosidase B led to a release of arabinose and the additional treatment of the supernatant thereof with a pectinase (Preparation 16) did not or only slightly increase the amount of released arabinose. As the total amount of released arabinose was higher in the fractions treated with a-L- arabinofuranosidase B compared to the hydrolysis of the soluble supernatant using pectinase preparation 16, it is likely that a-L-arabinofuranosidase B can also release arabinose from the insoluble part of the pea fractions.

[0252] The incubation of the alkaline extract AIS, acid extract AIS and the water extract supernatant with P-Galactosidase A and arabinogalactan endo-|3-l,4-galactanase A led to the release of free galactose (Figure 4d - f). However, the free galactose release was very low if the water extract supernatant was incubated with the latter two enzymes. Based on sequence homology, endo-|3-l,4-galactanase A was identified as an endo-active enzyme, but the release of galactose monomers also indicated the presence of an exo-activity. No galactose was released after the incubation of the three pea fractions with a-L-arabinofuranosidase B, arabinan endo-l,5-a-L-arabinosidase C or without any enzyme.

[0253] The incubation of the supernatants, which were obtained after the first enzymatic treatment using expressed enzymes, with a commercial pectinase (Preparation 16) led to a release of galactose in all fractions including the blank, which indicated that galactose-rich oligomers, like arabinogalactans, were present in the soluble supernatants (Figure 4d - f). In general, the release of galactose in the blank of the latter incubation withing the same range as for the fraction that were pre-incubated using the expressed enzymes. Only the incubation of water extract supernatant with a-L-arabinofuranosidase B led to an increase in the arabinose release after the soluble supernatant with the pectinase Preparation 16 (Figure 4f). Example 7 Method to determine the release of arabinose from destarched pea flour extract using an expressed a-arabinofuranosidase and a pectinase that comprises an arabinan- degrading activity

[0254] The release of free arabinose from destarched pea flour was determined after an enzymatic treatment using pectinase Preparation 20 and an a-L-arabinofuranosidase B (Figure 5). Pectinase preparation 20 was chosen for this comparison as it contained a high activity towards pNP-a-L-arabinofuranoside (Table 2; Example 4).

[0255] In brief, destarched pea flour was dissolved at a concentration of 5 mg ml1in MilliQ water. Depending on the experiment, the added enzyme concentration was 1% (w / w) and 1% (v / v) for Enzyme Preparation 20 and a-L-arabinofuranosidase B, respectively. After enzyme addition, samples were incubated at 40°C for 24 h. Samples were taken at various time points, followed by a rapid enzyme deactivation at 95°C for 15 min. After centrifugation (4°C, 20817 x g for 15 min), the arabinose content in the supernatants was analyzed using HPAEC (Example 1).

[0256] The results are shown in Figure 5.

[0257] Both pectinase Preparation 20 and a-L-arabinofuranosidase B were active towards destarched pea flour. After an incubation time of 24 h, the released arabinose content after the enzymatic treatment was about as much as the arabinose content which was present in the soluble fraction (1 %(w / w)) of destarched pea flour.

[0258] Example 8: Method for the determination of the release of free arabinose from destarched destarched pea flour extract using an expressed a-arabinofuranosidase and arabinose

[0259] The release of free arabinose from destarched pea flour was determined after an enzymatic treatment using an a-L-arabinofuranosidase B and / or an arabinan endo-l,5-a-L-arabinosidase C (Figure 6). Therefore, destarched pea flour was dissolved at a concentration of 5 mg ml1in MilliQ water. Depending on the experiment, the added enzyme concentrations were 1% (v / v) of a-L-arabinofuranosidase B (SEQ ID NO: 1), 0.5% (v / v) of a-L-arabinofuranosidase B (SEQ ID NO: 1) and 0.5% (v / v) of arabinan endo-l,5-a-L-arabinosidase C (SEQ ID NO: 4) leading to a total concentration of 1% (v / v), and 1% (v / v) of a-L-arabinofuranosidase B (SEQ ID NO: 1) and 1% (v / v) of arabinan endo-l,5-a-L-arabinosidase C (SEQ ID NO: 4) leading to a total concentration of 2% (v / v). After enzyme addition, samples were incubated at 40°C for 24 h. Samples were taken at various time points, followed by a rapid enzyme deactivation at 95°C for 15 min. After centrifugation (4°C, 20817 x g for 15 min), the free arabinose content in the supernatants was analyzed using HPAEC (Example 1).

[0260] The results are shown in Figure 6.

[0261] The highest release of free arabinose was determined after the incubation of destarched pea flour with the a-L-arabinofuranosidase B (1% (v / v)), which yielded to an arabinose release of 178 pg ml1. Addition of an arabinan endo-l,5-a-L-arabinosidase C to a-L-arabinofuranosidase B did not lead to an increase in the amount of released arabinose from destarched pea flour. The arabinose release after the incubation of destarched pea flour with the enzymes reached a maximum of 57% of the total amount of arabinose present in the samples after 24 h.

[0262] Example 9 Method for the preparation of an arabinose-containing liguid or semi-liguid food composition from low-refined plant protein-containing ingredients at pilot plant scale.

[0263] To obtain a food grade product, legume flours and concentrates, as well as whole grain flours were used. If the grains were not milled yet, a Retsch Mill (Retsch GmbH, Haan, Germany) was used to obtain a homogenous flour.

[0264] First, the flours were reconstituted in 20 L tap water to yield a final TS of 10% using a Stephan Mixer (Stephan universal mixer LIMSK 24E) (22 Kg in total, 25% mixing capacity for 3 min). Afterwards, oil was added, and the flour suspension was heated to 90-95°C and the temperature was kept for 15 min to ensure a complete gelatinization of the starch and a full deactivation of intrinsic enzymes. The resulting sample was a non-enzymatically-treated liquid or semi-liquid food composition and cooled down to a temperature of about 50°C. After enzyme addition, this batch was incubated at 50°C for 60 min applying the same stirring speed as described above. After enzyme hydrolysis, the hydrolysate was sieved, using a mesh size of 1 mm and homogenized at 50 and 250 bar (GEA Suisse AG, Kirchberg, Switzerland), yielding to an enzymatically-treated liquid or semi-liquid food composition. The cooled food composition was kept in a fridge and a UHT treatment was performed. The food composition sample was heat-treated (UHT treatment, OMVE Lab & Pilot Equipment, De Meern, The Netherlands) and aseptically filled in PP bottles.

[0265] Alternatively, if the Stephan Mixer was not always available, the hydration of the flours was done by slowly adding the flours to a bucket which was pre-filled with tap water under strong agitation (for 5 min) using an Ultra-Turrax. Afterwards, the flour suspension was added to a double jacket mixing tank and the treatment was performed the same way as described above.

[0266] The following enzyme preparations were used for the enzymatic treatment of pea flour, pea protein concentrate, defatted soy flour, soy flour concentrate:

[0267] Table 3: Enzyme preparation used for the Pilot Plant Trials

[0268] *Substrate weight refers to the weight of the liquid / semi-liquid food product containing all ingredients

[0269] Example 10 Sugar compositions including free and bound monosaccharides present in pea flour and pea flour concentrate-based liguid or semi-liguid food composition before and after enzymatic treatment.

[0270] Based on the carbohydrate composition analysis, the carbohydrate content accounted up to 48,4% and 16.9% of the pea flour and pea flour concentrate-based liquid or semi-liquid food composition based on dry matter, respectively (Table 5a). The total amount of pectic polysaccharides, which included quantified amounts of rhamnose, arabinose and galactose, was 2.3 and 1.6% of the pea flour and pea flour concentrate-based liquid or semi-liquid food composition based on dry matter, respectively (Table 5a). The free and bound uronic acid content was less than 5wt% in the pea flour and pea flour concentrate-based liquid or semiliquid food composition based on dry matter. Pea flour and pea flour concentrate was enzymatically-treated as described under Example 9. The selected enzyme preparations and concentrations, which were based on the screening (Example 4), are summarized in Table 3 (Example 9). The unbound monosaccharide composition was determined as described under Example 1. The determined free arabinose : free galactose ratio was 1:4,8 and 1:3,9 in the pea flour and pea flour concentrate-based liquid or semi-liquid food composition after enzymatic treatment , respectively (Table 5b).

[0271] Based on the carbohydrate composition analysis, the enzymatic treatment let to a release of 20.7-24.8% of the bound arabinose content as free arabinose and 69.0-74.7% of the bound galactose content as free galactose of the pea flour-based liquid or semi-liquid food composition, respectively (Table 5a & 5b). The soluble fraction after centrifugation of the latter samples accounted up for 86.3 - 93.4%. In comparison, the enzymatic treatment let to a release if 11.7-16.5% of the bound arabinose content as free arabinose and 36.5-51.6% of the bound galactose content as free galactose as free galactose of the pea flour concentratebased liquid or semi-liquid food composition, respectively (Table 5a & 5b). The soluble fraction after centrifugation of the latter sample accounted up for 49.4 - 69.9%. The amount of free arabinose and free galactose content that was present in the pea flour and pea flour concentrate-based liquid or semi-liquid food composition before enzymatic treatment was subtracted prior to calculating the effect of the enzyme preparation on the release of arabinose and galactose (Table 5a &b). Table 5a: Total monosaccharide composition including free and bound monosaccharides present in the pea flour and pea flour concentrate-based liquid or semi-liquid food composition *

[0272] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Sue, Sucrose; Man, Mannose; Fru, Fructose; Sum, sum of monosaccharides based on g / lOOg 'on dry matter'; Sum pectic polysaccharides including Rha, Ara, & Gal (% - percentage of pectic polymers per 100 g of DM). Table 5b: Unbound (i.e. free) monosaccharides present in the pea flour and pea flour concentrate-based liquid or semi-liquid food composition before and after the enzymatic treatment*

[0273] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Sue, Sucrose; Man, Mannose; Fru, Fructose; Sum, sum of monosaccharides based on g / lOOg 'on dry matter'.

[0274] Example 11 Sugar compositions including free and bound monosaccharides present in in enzymatically-treated defatted soy flour and soy flour concentrate-based liguid or semi-liguid food composition before and after enzymatic treatment.

[0275] Based on the carbohydrate composition analysis, the carbohydrate content accounted up to 44,2% and 39.2% of the defatted soy flour and soy flour concentrate-based liquid or semiliquid food composition based on dry matter, respectively (Table 6a). The total amount of pectic polysaccharides, which included quantified amounts of rhamnose, arabinose, galactose and galacturonic acid, was 5.1 and 4.4% of the defatted soy flour and soy flour concentratebased liquid or semi-liquid food composition based on total dry matter, respectively (Table 6a). Accordingly, the free and bound uronic acid content was less than 5wt% in the defatted soy flour and soy flour concentrate-based liquid or semi-liquid food composition based on dry matter. Defatted soy flour and soy flour concentrate was enzymatically-treated, as described under Example 9. The selected enzyme preparations and concentrations, which were based on the screening (Example 4), are summarized in Table 3 (Example 9). The unbound monosaccharide composition was determined as described under Example 1. The determined arabinose : galactose ratio was 1:3,4 and 1:1,5 in the defatted soy flour and soy flour concentrate-based liquid or semi-liquid food composition after enzymatic treatment, respectively (Table 6b).

[0276] Based on the results summarized in Table 6b, significant amounts of pectic polysaccharide, including free arabinose and free galactose, were released after the enzymatic treatment of the defatted soy flour and soy flour concentrate-based liquid or semi-liquid food composition compared to the preparation without enzymatic treatment. In the latter untreated compostion, no free arabinose and no free galactose was detected.

[0277] Table 6a: Total monosaccharide composition including free and bound monosaccharides present in the defatted soy flour and soy flour concentrate-based liquid or semi-liquid food composition *

[0278] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Sue, Sucrose; Man, Mannose; Fru, Fructose; GalA, Galacturonic acid; Sum, sum of monosaccharides based on g / lOOg 'on dry matter'; Sum pectic polysaccharides including Rha, Ara, Gal & GalA (% - percentage of pectic polymers per 100 g of DM).

[0279] Table 6b: Unbound (i.e. free) monosaccharides present is a defatted soy flour and soy flour concentrate-based liquid or semi-liquid food composition before and after the enzymatic treatment *

[0280] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Sue, Sucrose; Man, Mannose; Fru, Fructose; GalA, Galacturonic acid; Sum, sum of monosaccharides based on g / lOOg 'on dry matter'.

[0281] Example 12 Sugar compositions including free and bound monosaccharides present in a chickpea flour-based liguid or semi-liguid food composition before and after enzymatic treatment

[0282] Chickpea flour was prepared as described in Example 9 yielding to an chickpea flour-based liquid or semi-liquid food composition sample without enzymatic treatment. Afterwards, the composition was incubated using a ASGIP® Parallel Bioreactor System. The following process conditions were used: 1. Add 200 g per food composition sample + 200 mL of demineralized water to each chamber in the DasGip System, 2. Ensure good stirring (300 rpm), 3. Equilibrate the temperature to 50°C, 4. Add enzymes (See below for concentrations), 5. Incubate the food composition for 1 h at 50°C, 6. After incubation, fill the hydrolysate into a glass flask and place them into a water bath which was pre-heated to 95°C, 7. Heat up the samples to reach 90°C and keep them at 90°C for 10 min to deactivate all enzymes, 8. Analyze the unbound (i.e. free) monosaccharide composition as described in Example 1. The following enzymes and concentrations thereof were used (Enzyme Mix 1):

[0283] Cellulase (Preparation 25), 0.2%(w / w) (weight Enzyme Preparation / weight food composition)

[0284] Amylase (Preparation 37), 0.1%(w / w) (weight Enzyme Preparation / weight food composition)

[0285] Amyloglucosidase (Preparation 38), 0.1%(w / w) (weight Enzyme Preparation / weight food composition)

[0286] In addition of the above-described incubation, the chickpea flour-based liquid or semi-liquid food composition was also incubated with Enzyme Mix 1 and pectinase Preparation 12 and Preparation 20 at a concentration of 0.4%(w / w) (weight Enzyme Preparation / weight food composition). Based on the results summarized in Table 7, significant amounts of pectic polysaccharide, including free arabinose and free galactose, were released afterthe enzymatic treatment that included the use of a pectinase of the chickpea flour-based liquid or semi-liquid food composition compared to the composition a pectinase-containing enzymatic treatment. The determined free arabinose : free galactose ratio was 1:4.3 (cf. Table 7). The free and bound uronic acid content was less than 5wt% in the chickpea flour-based liquid or semi-liquid food composition based on dry matter.

[0287] Table 7: Unbound (=free) monosaccharide composition of a treated chickpea flour-based liquid or semi-liquid food composition using an enzyme mix without and with pectinase addition*

[0288] *Fuc, Fucose; Rha, Rhamnose; Ara, Arabinose; Gal, Galactose; Glc, Glucose; Xyl, Xylose; Sue, Sucrose; Man, Mannose; Fru,

[0289] Fructose

[0290] Although the invention has been described by way of example, it should be appreciated that variations and modifications may be made without departing from the scope of the invention as defined in the claims.

[0291] References

[0292] Quemener, B., Thibault, J.-F., (1990). Assessment of methanolysis for the determination of sugars in pectins, Carbohydrate Research, 206(2), 277-287. DOI: 10.1016 / 0008-6215(90)80067-D

Claims

CLAIMS1. An arabinose-containing and galactose-containing food composition, which is liquid or semi-liquid or powdered and which comprises:- plant proteins selected from the list consisting of legume proteins, seed proteins and mixture thereof,-less than 5wt.% free uronic acid by dry weight,-a weight ratio of free arabinose to free galactose (free arabinose: free galactose) of from 1:1.5 to 1:5, preferably 1:3.5 to 1:4.5.

2. The arabinose-containing and galactose-containing food composition according to claim 1, which comprises less than 30wt.% starch.

3. The arabinose-containing and galactose-containing food composition according to claim 1 or 2, which comprises at least 0.25wt.% free arabinose by dry weight, preferably free endogenous arabinose by dry weight.

4. The arabinose-containing and galactose-containing food composition according to any of the preceding claims, which comprises less than 30wt% non-starch polysaccharides by dry weight.

5. The arabinose-containing and galactose-containing food composition according to any of the preceding claims, which comprises less than 20wt.% pectic polysaccharides by dry weight.

6. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, wherein the legume proteins come from the legume selected from the list consisting of pea, cowpea, split pea, peanut, lentil, faba bean, chickpea, bean, soybean and combination thereof.

7. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, wherein the seed proteins come from the seed selected from thelist consisting of almond, rapeseed, sunflower seed, hemp seed, flaxseed, linseed, sesame seed, pumpkin seed, chia seed and mixture thereof.

8. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, wherein the plant proteins are provided as plant flour, plant protein concentrate, ground whole legume, ground whole seed and mixture thereof.

9. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, which is vegetarian or vegan.

10. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, which is a beverage, preferably plant-based milk analogue.

11. The arabinose-containing and galactose-containing food composition according to any one of the preceding claims, wherein the free arabinose comprises or consists of free endogenous arabinose and all of the free endogenous arabinose is derived from a legume and / or a seed.

12. The arabinose-containing and galactose-containing food composition according to claim 11, wherein the free endogenous arabinose and the vegetable protein and / or seed proteins are derived from the same legume and / or seed.

13. A method for selecting an enzyme comprising a pectinase activity which is high at a pH from 6 to 7.5, preferably at a pH of 7:(a) providing one or several enzymes, providing a liquid substrate medium A comprising a substrate A and providing a liquid substrate B comprising a substrate B, wherein the substrate A is a molecule comprising an a-arabinofuranosyl-residue attached to a chromogenic compound, and wherein the substrate B is molecule comprising a |3- galactosyl-residue attached to a chromogenic compound,(b) contacting the liquid substrate medium A with said one or several enzymes for an incubation time of 1 to 15 minutes, at a temperature of 20°C to 60°C, preferably 30°Cto 50, more preferably 37°C and at a pH from 6 to 7.5, preferably at a pH of 7 to obtain a treated liquid substrate medium A,(c) contacting the liquid substrate medium B with said one or several enzymes for an incubation time of 1 to 15 minutes, at a temperature of 20°C to 60°C, preferably 30°C to 50, more preferably 37°C and at a pH from 6 to 7.5, preferably at a pH of 7 to obtain a treated liquid substrate medium B,(d) measuring the lytic activity towards pNP-a-L-arabinofuranoside in the treated liquid substrate medium A: after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute, after 1 minute, if the incubation time used in step (b) is equal to 1 minute,(e) measuring the lytic activity towards pNP-|3-D-galactopyranoside in the treated liquid substrate medium B: after one or several time-points between 1 minute and the incubation time used in step (b), if the incubation time used in step (b) is different than 1 minute, after 1 minute, if the incubation time used in step (b) is equal to 1 minute,(f) selecting the one or more enzymes that have:- a lytic activity towards pNP-a-L-arabinofuranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium A based on the lytic activity measured in step (d), and- a lytic activity towards pNP-|3-D-galactopyranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium B based on the lytic activity measured in step (e).

14. A process for preparing an arabinose-containing and galactose-containing food composition which is liquid or semi-liquid or powdered comprising the steps of:(a) providing a low-refined plant protein-containing ingredient, wherein the low-refined plant protein-containing ingredient comprises pectic polysaccharides,(b) dispersing the low-refined plant protein-containing ingredient in an aqueous liquid to form a plant dispersion,(c) optionally, heat treating the plant dispersion,(d) optionally, adjusting the pH of the plant dispersion to a pH of 6 to 7.5,(e) contacting the plant dispersion which has a pH from 6 to 7.5 with an enzyme preparation comprising at least one enzyme comprising a pectinase activity to provide an enzymatically-treated arabinose-containing and galactose-containing liquid or semi-liquid food composition,(f) heat-treating the enzymatically-treated arabinose-containing and galactose- containing liquid or semi-liquid food composition to inactivate the at least one enzyme comprising a pectinase activity and to form an arabinose-containing liquid or semiliquid food composition,(g) optionally, drying of the arabinose-containing and galactose-containing liquid or semiliquid food composition to form an arabinose-containing and galactose-containing powdered food composition, wherein the amount of free arabinose by dry weight is increased by at least 0.25wt% and the amount of free galactose by dry weight is increased by at least 0.3wt.% in the enzymatically-treated arabinose and galactose-containing food composition of step (e) compared to the plant dispersion of step (b).

15. A process according to claim 14, wherein the low-refined plant protein-containing ingredient is a plant flour or plant protein concentrate or ground whole plant material.

16. A process according to any one of claim 14 or 15, wherein the low-refined plant protein-containing ingredient comes from seed and / or legume.

17. A process according any one of claims 15-16, wherein the enzyme comprising a pectinase activity, when it is subjected to a method comprising step (a) to (e) of claim 11, has:- a lytic activity towards pNP-a-L-arabinofuranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium of step (b), and- a lytic activity towards pNP-|3-D-galactopyranoside of at least 1000.0, preferably at least 5000.0 units per gram of undiluted enzyme in the treated liquid substate medium of step (b).

18. A process according to any one of claims 14-17, wherein the enzyme comprising a pectinase activity has at least 60%, preferably 80%, more preferably 90% sequence identity with one of the amino acid sequences SEQ ID NO: 1-3.

19. A process according any one of claims 14-18, wherein the enzyme comprising a pectinase activity is derived from Neosartorya fumigata or Aspergillus niger.

20. A process according to any one of claims 14-19, wherein the content of enzyme comprising a pectinase activity contacted with the plant dispersion in step (e) is of at least 0.1%, preferably 0.1 to 0.7% (w / w).

21. A process according to any one of claims 14-20, wherein the plant dispersion is further contacted in step (e) or before step (e) with at least one starch-degrading enzyme and / or at least one cellulose-degrading enzyme and / or at least one protein-degrading enzyme.

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