Enzymatic production of soluble fiber

Abstract

The present invention relates to a method for preparing a mixture of indigestible α-glucans from a substrate rich in oligosaccharides having a degree of polymerization (DP) of 4.

Description

[Technical Field]

[0001] The present invention relates to a method for preparing a mixture of indigestible α-glucans from a substrate rich in oligosaccharides having a degree of polymerization (DP) of 4. In this application, this substrate refers to an oligosaccharide-containing syrup having a content of at least 40%, preferably at least 45%, and even more preferably at least 50% of oligosaccharides having a degree of polymerization (DP) of 4.

[0002] The present invention also relates to a mixture of indigestible α-glucans.

[0003] The present invention also relates to the use of an α-glucanotransferase capable of generating α(1,6) glycosidic bonds to reduce the digestibility of a mixture of α-glucans. [Background technology]

[0004] Dietary fiber plays an important role in human nutrition. Among dietary fibers, soluble fiber, which is water-soluble and has gelling properties, is distinguished from insoluble fiber. Soluble fiber, including branched maltodextrins, is particularly advantageous because it is indigestible. Therefore, incorporating them into the diet can reduce the glycemic index of food and prolong satiety. They also have prebiotic properties for the intestinal flora. In other words, they can selectively promote the growth of specific probiotic-type bacteria or the activity of the microflora, thereby providing health benefits.

[0005] To date, soluble fibers, including branched maltodextrins, have been obtained primarily through physicochemical methods.

[0006] This is particularly the case for the maltodextrin sold by the Applicant Company under the trade name NUTRIOSE® FM10 as a water-soluble fibre.

[0007] There are other soluble fibres obtained physicochemically, such as PROMITOR® sold by the company Tate and Lyl, FIBERSOL® or LITESSE® sold by Dupont Nutrition and Biosciences.

[0008] Many studies have demonstrated that digestibility properties are directly related to the proportion of different glycosidic bonds within soluble fiber.

[0009] In fact, standard maltodextrins are defined as rapidly digestible, purified and concentrated mixtures of glucose and glucose polymers essentially linked via α1→4 (hereinafter 1→4 or α(1,4)) linkages, with only 4-5% of α1→6 glycosidic bonds (hereinafter 1→6 or α(1,6)), of very different molecular weights, completely water-soluble and with low reducing power.

[0010] By increasing the proportion of α1→6 or α1→3 linkages, the degree of branching of the maltodextrin increases, which makes the maltodextrin more resistant to digestion.

[0011] Enzymatic approaches using enzymes that can promote the creation of "branched" type bonds have many advantages in terms of safety and environmental protection, and also offer better specificity.

[0012] Originally, most enzymatic methods for producing soluble fiber are carried out using sucrose as a substrate for the enzyme to create new bonds. For example, WO2015183714 describes an enzymatic reaction from a mixture of sucrose and an α-glucan type substrate.

[0013] Currently, most enzymatic processes use amylomaltase to produce soluble fiber from starch.

[0014] It is desirable to enzymatically obtain soluble fiber from a substrate in the absence of sucrose. DETAILED DESCRIPTION OF THE INVENTION

[0015] The Applicant Company has discovered that it is possible to enzymatically obtain fibers of interest in human and animal nutrition from syrups rich in oligosaccharides with a degree of polymerization (DP) of 4. The Applicant Company has therefore developed a method using specific enzymes capable of generating α(1,6) bonds from syrups rich in DP4 oligosaccharides.

[0016] In a first aspect, the present invention relates to a method for preparing a mixture of α-glucans, preferably a mixture of branched maltodextrins, the method comprising the step of combining a substrate and an enzyme, wherein the substrate is a syrup rich in oligosaccharides having a degree of polymerization (DP) of 4, and the enzyme is an α-glucanotransferase capable of cleaving α(1,4) glycosidic bonds to generate α(1,6) glycosidic bonds.

[0017] According to the present invention, the terms "α-glucan", "soluble fiber" and "food soluble fiber" are used interchangeably. They define oligosaccharides composed of at least three glucose units linked together by α-glycosidic (or α-glucosidic) bonds.

[0018] The classification of α-glucans is primarily based on the measurement of their reducing power, conventionally expressed by the concept of "Dextrose Equivalent" (DE). In this particular respect, the definition of maltodextrin given in the Monograph Specifications of the Food Chemical Codex specifies that the DE value of maltodextrins must not exceed 20. Above 20, they are glucose syrups.

[0019] However, such DE measurements are insufficient to accurately represent the molecular distribution of α-glucan: indeed, acid hydrolysis of starch, which is completely random, or its enzymatic hydrolysis, which is slightly more ordered, cannot be accurately defined by a single DE measurement and provides a mixture of glucose and glucose polymers containing short-sized molecules with low DP and very long-sized molecules with high DP.

[0020] Measurement of DE actually only gives an approximate idea of the average DP of a mixture of glucose and the constituent glucose polymers of α-glucan and therefore their number-average molecular weight (Mn). To complete the characterization of the molecular weight distribution of α-glucan, it is important to determine another parameter, the weight-average molecular weight (Mw).

[0021] In practice, (Mn) and (Mw) are determined experimentally by different analytical techniques, such as a measurement method suitable for glucose polymers, based on gel permeation chromatography on a chromatography column calibrated with pullulan of known molecular weight.

[0022] The Mw / Mn ratio, known as the polymolecularity index (PI), allows characterizing the overall molecular weight distribution of a polymer blend. As a rule, the molecular weight distribution of standard maltodextrins results in an PI value between 5 and 10.

[0023] These various parameters also reflect the α-glycosidic bond profile of the α-glucan: indeed, a standard α-glucan mixture has a very high proportion of "linear" α(1,4) bonds (more than 90%) and a low proportion of so-called "branched" (α(1,2), α(1,3) and α(1,6) bonds).

[0024] The method according to the invention makes it possible to reduce the proportion of α(1,4) bonds in favor of α(1,6) bonds, which has the advantage of reducing the digestibility of the mixture of α-glucans obtained by this method.

[0025] The mixture of α-glucans prepared according to the method of the present invention is preferably a mixture of branched maltodextrins.

[0026] For the purposes of the present invention, branched maltodextrins are understood to be maltodextrins which have a higher content of α(1,6) glycosidic bonds than standard maltodextrins.

[0027] Standard maltodextrins are defined as purified and concentrated mixtures of glucose and essentially α(1,4)-linked glucose polymers with only 4-5% α(1,6) glycosidic bonds, of very different molecular weights, completely water-soluble, and with low reducing power.

[0028] According to one embodiment of the present invention, the syrup rich in oligosaccharides with DP4 comprises at least 40%, preferably at least 45%, even more preferably at least 50% oligosaccharides with DP4.

[0029] According to one embodiment of the present invention, the syrup rich in oligosaccharides having a DP4 has a dextrose equivalent (DE) of more than 20.

[0030] According to a preferred embodiment of the present invention, the DP4 enriched syrup is a syrup having the properties set out in Table 1 below.

[0031] In a preferred embodiment of the present invention, the substrate is present in the reaction medium at a concentration of 50 g / L to 500 g / L, preferably 100 g / L to 200 g / L.

[0032] In a preferred embodiment of the present invention, the α-glucanotransferase capable of cleaving an α(1,4) glycosidic bond to generate an α(1,6) glycosidic bond is a protein having the sequence of SEQ ID NO: 1 or a protein having at least 90% identity to a protein having the sequence of SEQ ID NO: 1 (hereinafter known as GT#19). Preferably, the α-glucanotransferase is a protein having at least 91%, even more preferably at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identity to a protein having the sequence of SEQ ID NO: 1. The sequence of SEQ ID NO: 1 corresponds to Genbank accession number WP_053069107.1.

[0033] As shown in the Examples, the inventors demonstrated that enzyme GT#19 can modify DP4-rich syrup to make it resistant to digestion (percent hydrolysis ≤ 45% according to method AOAC 2002.02).

[0034] According to one embodiment of the present invention, the enzyme is added at a concentration of 0.01 to 1 mg / mL of reaction medium, preferably 0.05 to 0.5 mg / mL, and even more preferably about 0.1 mg / mL of reaction medium.

[0035] According to one embodiment of the present invention, the substrate and the enzyme are contacted for 12 to 48 hours, preferably about 24 hours.

[0036] According to one embodiment of the present invention, the substrate and enzyme are combined at a temperature of 20-40°C, preferably about 37°C.

[0037] According to one embodiment of the present invention, the substrate and enzyme are combined at a pH of 5 to 6.5, preferably 5.5 to 6, and even more preferably about 5.75.

[0038] In one embodiment of the present invention, the method further comprises a step of enzymatic treatment with an α-glucanotransferase capable of cleaving an α(1,4) glycosidic bond to generate an α(1,3) glycosidic bond. For example, the α-glucanotransferase can be a protein having the sequence of SEQ ID NO:2 or a protein having at least 90% identity to a protein having the sequence of SEQ ID NO:2 (hereinafter known as GT#11). Preferably, the α-glucanotransferase is a protein having at least 91%, even more preferably at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identity to a protein having the sequence of SEQ ID NO:2. The sequence of SEQ ID NO:2 corresponds to Genbank accession number AOR73699.1.

[0039] According to one aspect, the present invention also relates to a mixture of α-glucans, preferably a mixture of branched maltodextrins, obtainable by the method described above.

[0040] This mixture of α-glucans is characterized by a low digestibility according to the AOAC 2002.02 method. Advantageously, the method according to the invention makes it possible to reduce the hydrolyzable fraction, measured according to the AOAC 2002.02 method, by at least 2-fold, preferably by at least 2.5-fold, and even more preferably by at least 3-fold relative to the starting substrate.

[0041] The AOAC 2002.02 method can be performed using, inter alia, the "HPAEC-PAD Assay" portion of the "Starch Resistance, K-RSTAR06 / 18" kit sold by Megazyme®, Inc., as described in Example 1, Part 5 below.

[0042] The method according to the invention makes it possible to increase the proportion of α(1,6) bonds by at least 3-fold, preferably by at least 3.5-fold, and even more preferably by at least 4-fold relative to the starting substrate.

[0043] The percentage of α(1,6) linkages can be measured by the Hakomori method (1964 HAKOMORI A Rapid Permethylation of Glycolipid, and Polysaccharide Catalyzed by Methylsulfinyl Carbanion in Dimethyl Sulfoxide) as described in Example 1, Part 8 below, or by proton NMR as described in Example 1, Part 7 below.

[0044] According to one aspect, the present invention relates to an α-glucan mixture, preferably a mixture of branched maltodextrins, comprising: - a content of hydrolyzable fibers of less than 55%, preferably less than 50% and even more preferably less than 45%; - and / or at least 20% α(1,6) bonds characterized in that it has The fiber content corresponds to the hydrolyzable (i.e., non-resistant) fraction according to the AOAC 2002.02 method, and the percentage of α(1,6) bonds represents the molar percentage of α(1,6) bonds relative to the total number of glycosidic bonds, as determined by the Hakomori method.

[0045] Preferably, the hydrolyzable fiber content is less than 44%, preferably less than 43%, even more preferably less than 42%, 41%, 40%, 39%, 38%, 37%, 36%, 35%, 34%, 33%, 32%, 31%, 30%.

[0046] Preferably the hydrolyzable fiber content is greater than 5%, preferably greater than 10%, 11%, 12%, 13%, 14%, 15%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%.

[0047] Preferably, the content of hydrolyzable fibers is 5% to 45%, preferably 10% to 45%, preferably 20% to 44%, and even more preferably 30% to 45%.

[0048] Preferably, the percentage of α(1,6) linkages is at least 21%, preferably at least 22%, even more preferably at least 23%, at least 24%, at least 25%, at least 26%, at least 27%, at least 28%, at least 29%, at least 30%, at least 31%, at least 32%, at least 33%, at least 34%, at least 35%.

[0049] Preferably, the proportion of α(1,6) bonds is at most 40%, preferably at most 36%, at most 35%, at most 34%, at most 33%, at most 32%, at most 31%.

[0050] Preferably, the proportion of α(1,6) bonds is 20% to 40%, preferably 20% to 35%, more preferably 25% to 35%.

[0051] Preferably, the proportion of α(1,3) bonds is at least 2%, preferably at least 3%.

[0052] Preferably, the proportion of α(1,3) bonds is at most 6%, preferably at most 5%.

[0053] Preferably, the proportion of α(1,3) bonds is 2% to 8%, more preferably 3% to 5%.

[0054] Preferably, the proportion of α(1,2) bonds is at least 1%, preferably at least 2%.

[0055] Preferably, the proportion of α(1,2) bonds is at most 8%, preferably at most 6%, at most 5%, at most 4% or at most 3%.

[0056] Preferably, the proportion of α(1,4) bonds is at most 80%, preferably at most 70%, at most 65%.

[0057] Preferably, the proportion of α(1,4) bonds is at least 50%, preferably at least 55%, at least 60%.

[0058] Preferably, the proportion of α(1,4) bonds is 50% to 80%, preferably 55% to 70%, more preferably 55% to 65%.

[0059] Preferably, the mixture of α-glucans is preferably a mixture of branched maltodextrins, the mixture comprising: - characterized in that it has a content of hydrolyzable fibres of less than 55%, preferably less than 50% and even more preferably less than 45%; - and - 20% to 40% α(1,6) bonds, - 20% to 40% α(1,3) bonds, - 2% to 8% α(1,2) bonds, - 50% to 80% α(1,4) bonds, The fiber content corresponds to the hydrolyzable (i.e. non-resistant) fraction according to the AOAC 2002.02 method, and the proportion of α(1,6), α(1,3), α(1,2) or α(1,4) bonds represents the molar percentage of such types of bonds relative to the total number of glycosidic bonds, as determined by the Hakomori method.

[0060] Preferably, the content of α(1,6), α(1,3), α(1,2) or α(1,4) bonds is such that the sum of their molar percentages equals 100%.

[0061] The present invention also relates to the use of a mixture of α-glucans obtained according to the aforementioned method and having the aforementioned properties for preparing a nutritional food product for humans or animals.

[0062] Typically, the mixture of alpha glucans according to the present invention can be used to promote gut health, glycemic control, satiety and weight management, and sustained energy release.

[0063] Finally, in another aspect, the present invention relates to the use of a glucanotransferase capable of cleaving α(1,4) glycosidic bonds to generate α(1,6) glycosidic bonds to reduce the digestibility of a mixture of α-glucans, the glucanotransferase having the sequence of SEQ ID NO: 1 or a protein having at least 90% identity to a protein having the sequence of SEQ ID NO: 1.

[0064] In this aspect of the invention, the mixture of α-glucans is preferably an oligosaccharide-rich syrup, in particular an oligosaccharide-rich syrup having DP4 as described in the first aspect of the invention.

[0065] Preferably, the reduction in digestibility is at least a 2-fold, preferably at least a 2.5-fold, even more preferably at least a 3-fold reduction in the hydrolyzable fraction measured according to the AOAC 2002.02 method relative to the starting substrate.

[0066] The present invention will be better understood with the aid of the following examples, which are intended to be illustrative and non-limiting.

[0067] Example 1: Preparation of branched maltodextrins from DP4-rich syrup: Materials and methods 1. Preparation of DP4 Substrate Solution The starting substrate used was a DP4-rich syrup with the properties listed in Table 1.

[0068] [Table 1]

[0069] Solutions of the various substrates in pH 5.75, 50 mM sodium acetate buffer (DP4-rich syrup) were prepared at concentrations of 100 g / L, 200 g / L, or 400 g / L.

[0070] 2. Production of recombinant enzymes. The following enzymes were recombinantly produced: - Enzyme GT#11: α-4,3 glucanotransferase from Lactobacillus fermentum NC2970 (SEQ ID NO: 2) which has as its amino acid sequence the sequence listed in Genbank under the reference AOR73699.1. - Enzyme GT#19: glycoside hydrolase GH70 from Lactobacillus mucosae, having as amino acid sequence the sequence listed in Genbank under the reference number WP_053069107.1 (SEQ ID NO: 1).

[0071] E. coli BL21 star cells containing the plasmid pET-21 a-enzyme no.X (to produce various enzymes, including GT#11 and GT#19) were cultured in ZYM-5052 medium containing 1% glycerol and 1% lactose. At the end of the culture, the cells were centrifuged at 6,500 g for 10 minutes, and the cell pellet was resuspended in 20 mM phosphate buffer (pH 7.4) containing 300 mM NaCl and 20 mM imidazole at a DO of 80. The cells were lysed by cold sonication using four cycles of 20 seconds at 30% amplitude followed by a 4-minute pause. Cell debris was separated from the solubilized protein by centrifugation at 10,000 g for 30 minutes.

[0072] 3. Enzyme Purification Purification of the target protein was performed on a cobalt resin (Invitrogen) loaded with divalent cobalt ions (CO2+), for which the polyhistidine tag has affinity. Elution was performed by competition between the polyhistidine tag and increasing concentrations of imidazole. Briefly, 10–35 mL of E. coli cell extract was contacted with 1 mL of cobalt resin pre-equilibrated with 25 mL of 20 mM phosphate buffer (pH = 7.4) containing 300 mM NaCl and 20 mM imidazole for 1 h. All unbound protein was removed by filtering the resin through a sintered glass filter. The resin was then washed five times with 40 mL of 20 mM phosphate buffer (pH = 7.4) containing 300 mM NaCl and 20 mM imidazole. Finally, the target enzyme was isolated by elution with 3 mL of 20 mM phosphate buffer (pH 7.4) containing 300 mM NaCl and 250 mM imidazole for 5 min. The enzyme solution was then dialyzed (10 kDa molecular weight cutoff membrane) against 5 L of 50 mM sodium acetate buffer (pH 5.75) overnight at 4 °C with stirring to remove NaCl and imidazole. The various protein solutions were assayed by measuring their absorbance at 280 nm using a 2000 Nanodrop spectrophotometer (Thermofisher). The molar extinction coefficient ε was determined using the ProtParam tool application on the ExPASy bioinformatics resource portal.

[0073] 4. Enzyme reaction Reactions were performed with 0.1 mg / mL of purified enzyme and dialyzed in the presence of 10%, 20%, or 40% substrate in 50 mM sodium acetate buffer (pH = 5.75). The reactions were incubated at 37 °C for 24 h with stirring. The reactions were stopped by heating (95 °C for 5 min). Samples were taken at the beginning and end to analyze the specificity of the enzyme using different analytical techniques (HPAEC-PAD, NMR).

[0074] 5. Digestibility test The transfection reaction was frozen at -80°C for 24 hours and then lyophilized. 25 mg of the lyophilized product was dissolved in 1 mL of 100 mM sodium maleate buffer containing 30 U of pancreatic α-amylase and 3 U of amyloglucosidase (starch resistance kit, Megazyme K-STAR 06 / 18, which implements the AOAC 2002.02 method). The reaction was incubated at 37°C for 16 hours. The product was diluted with water prior to HPAEC PAD analysis.

[0075] 6. Chromatographic Analysis The resulting product was analyzed by anion exchange chromatography coupled to a pulsed amperometric detector (HPAEC PAD - HIGH Performance Anion Exchange Chromatography with Pulsed Amperometric Detection). The analysis was performed on a Thermo ICS6000 system equipped with a CarboPac™ PA100 analytical column (2 mm x 250 mm) coupled to a CarboPac™ PA100 guard precolumn (2 mm x 50 mm). A gradient of sodium acetate in 150 mM sodium was applied at a flow rate of 0.250 mL / min according to the following profile: 0-5 min, 0 mM; 5-35 min, 0-300 mM; 35-40 min, 300-450 mM; 40-42 min, 450 mM. Detection was performed using a gold working electrode and a pH Ag / AgCl reference cell. Samples were diluted to a total dry mass of 1 g / L before injection.

[0076] 7. NMR. Spectrum 1 H, 13 C and HSQC were recorded on a Bruker Avance 500 MHz instrument using a 5 mm Z-gradient H-BB-D BBI probe at 298 K. Data were acquired and processed using TopSpin 3 software.

[0077] 8.Hakomori method The Hakomori method (1964 HAKOMORI A Rapid Permethylation of Glycolipid, and Polysaccharide Catalyzed by Methylsulfinyl Carbanion in Dimethyl Sulfoxide) allows for the chemical characterization of glycosidic bonds by distinguishing between free OH groups and bonded groups. It is a destructive method that includes the steps of methylation, hydrolysis, reduction with NaBD4, acetylation, and analysis by mass spectrometry.

[0078] Example 2: Preparation of branched maltodextrins from DP4-rich syrup: Results The results of the various enzymatic reactions are shown in Table 2 below, which shows the proportions of α-1,6, α-1,3 and α-1,4 linkages measured by proton NMR or the Hakomori method, as well as the proportion of hydrolysis (AOAC2002.02) in the resulting reaction products.

[0079] [Table 2] RM: Raw material = DP4 syrup

[0080] The inventors have demonstrated that enzyme GT#19 can modify DP4-rich syrup to make it resistant to digestion (percentage of hydrolysis ≦45% according to method AOAC 2002.02).

[0081] The product obtained by this enzymatic treatment with enzyme GT#19 contains significantly fewer α-1,4 linkages and more α-1,6 linkages than the starting material. The number of α-1,2 and α-1,3 linkages remains unchanged. Therefore, enzyme GT#19 is a 4,6-α-glucanotransferase.

[0082] Conversely, treatment with another GT, enzyme GT#11, results in a decrease in the proportion of α-1,4 linkages and the appearance of α-1,3 linkages. The digestibility of the product obtained by treatment with enzyme GT#11 is also reduced (53 and 55% for initial concentrations of 100 and 200 mg / mL, respectively, compared to 84 and 88% for the untreated substrate).

[0083] Advantageously, the mixture of α-glucans according to the invention also has an interesting in vitro digestibility profile according to the Englyst method.

Claims

1. A method for preparing a mixture of α-glucans, comprising the step of combining a substrate and an enzyme, wherein the substrate is an oligosaccharide-rich syrup having a degree of polymerization (DP) of 4, and the enzyme is an α-glucanotransferase capable of cleaving α(1,4) glycosidic bonds to produce α(1,6) glycosidic bonds.

2. The method according to claim 1, wherein the syrup rich in oligosaccharides having DP4 contains at least 40%, preferably at least 45%, and more preferably at least 50% of oligosaccharides having DP4.

3. The method according to claim 1, wherein the syrup, which is rich in oligosaccharides having DP4, has more than 20 dextrose equivalents (DE).

4. The method according to claim 1, wherein the substrate is present in the reaction medium at a concentration of 50 g / L to 500 g / L, preferably 100 g / L to 200 g / L.

5. The method according to claim 1, wherein the α-glucanotransferase capable of cleaving the α(1,4) glycosidic bond to generate an α(1,6) glycosidic bond is a protein having the sequence of SEQ ID NO: 1, or a protein having at least 90% identity with the protein having the sequence of SEQ ID NO:

1.

6. The method according to claim 1, wherein the enzyme is present in the reaction medium at a concentration of 0.01 to 1 mg / mL, preferably 0.05 to 0.5 mg / mL, and more preferably about 0.1 mg / mL of the reaction medium.

7. The method according to claim 1, characterized in that the substrate and the enzyme are brought together for a period of 12 to 48 hours, preferably about 24 hours, and / or at a temperature of 20 to 40°C, preferably about 37°C, and / or at a pH of 5 to 6.5, preferably about 5.

75.

8. The process involves cleaving an α(1,4) glycosidic bond to create an α(1,3) glycosidic bond. The method according to claim 1, further comprising a step of enzymatic treatment with α-glucanotransferase that can produce [the product].

9. The method according to claim 8, wherein the α-glucanotransferase capable of cleaving the α(1,4) glycosidic bond to generate an α(1,3) glycosidic bond is a protein having the sequence of SEQ ID NO: 2, or a protein having at least 90% identity with the protein having the sequence of SEQ ID NO:

2.

10. A mixture of α-glucans that can be obtained by the method described in claim 1.

11. A mixture of α-glucans, wherein the mixture of α-glucans is - Content of hydrolyzable fibers less than 55%, preferably less than 50%, and more preferably less than 45%, - and / or at least 20% α(1,6) bonds It is characterized by having, The fiber content corresponds to the hydrolyzable fraction according to the AOAC2002.02 method, and the proportion of α(1,6) bonds represents the molar percentage of α(1,6) bonds relative to the total number of glycosidic bonds, as measured by the Hakomori method, in a mixture of α-glucans.

12. Use of the α-glucan mixture according to any one of claims 10 or 11 for preparing a food for the nutrition of humans or animals.

13. The use of a glucanotransferase capable of cleaving α(1,4) glycosidic bonds to generate α(1,6) glycosidic bonds in order to reduce the digestibility of a mixture of α-glucans, wherein the glucanotransferase has the sequence of SEQ ID NO: 1 or a sequence having at least 90% identity with a protein having the sequence of SEQ ID NO: 1.