WO2018215450A1

WO2018215450A1 - Konjac glucomannan hydrolysates

Info

Publication number: WO2018215450A1
Application number: PCT/EP2018/063362
Authority: WO
Inventors: Bernard Pora; Véra YANG; Jovin Hasjim
Original assignee: Roquette Freres
Priority date: 2017-05-23
Filing date: 2018-05-22
Publication date: 2018-11-29
Also published as: CN108949855A; CN110637037A; CN108949855B

Abstract

The present invention relates to a process for preparing konjac glucomannan hydrolysates said process comprising the following steps: a. hydrolyzing a konjac glucomannan flour in the presence of a food-grade acid to provide a partially hydrolyzed konjac glucomannan, the moisture content at the beginning of said acid hydrolysis not exceeding 25 % by weight, preferably between 15 and 22 % by weight, more preferably between 18 and 20 % by weight; b. further hydrolyzing the partially hydrolyzed product of step a) in the presence of at least one enzyme chosen in the group consisting of transglucosidase, β-amylase, pullulanase, maltogenase glucoamylase, cyclodextrin glycosyltransferase, β-glucanase, cellulase, xylanase and polygalacturonase, preferably in the group consisting of transglucosidase, β-amylase, pullulanase and maltogenase, more preferably in the group consisting of transglucosidase and pullulanase; c. recovering the konjac glucomannan hydrolysates of step b).

Description

KONJAC GLUCOMANNAN HYDROLYSATES

Field of the invention

The present invention relates to a novel process for preparing konjac glucomannan hydrolysates from a konjac glucomannan flour. The present invention also relates to the hydrolysates obtainable by said process. The present invention further relates to the use of said hydrolysates for the preparation of a food product or beverage.

Background

Konjac glucomannan is a water soluble dietary fiber obtained from konjac plant root powder.

Konjac glucomannan exerts many beneficial physiological functions. For example, it is a beneficial dietary adjunct for treatment of metabolic syndromes, such as hypercholesterolemia, glucose intolerance, obesity and diabetes. Konjac glucomannan is also a natural prebiotic and can be used to alleviate constipation.

Thus, as a kind of healthy ingredient, konjac glucomannan flour found interest from food and nutraceutical industries, especially in Asia. In food industries, konjac glucomannan flour is commonly used as a gelling agent, a thickening agent. However, its high viscosity limits its application. Indeed, konjac glucomannan flour swells instantly upon contact with water and its viscosity increases rapidly, causing lump formation and preventing the complete dispersion of the glucomannan in water.

In nutraceutical industries, konjac glucomannan flour is mainly used for weight control. It is usually placed in hard capsules or used in a small quantity in powdered drink mix due to the limitation of its high viscosity. The viscosity of konjac glucomannan (≥ 90% purity) solution at 1% concentration can reach above 30,000 cP at 30°C. At this low concentration, some of the health benefits of konjac glucomannan may be limited, such as to increase the intakes of dietary fiber and prebiot ic .

It is therefore very difficult to make homogeneous solution of konjac glucomannan above 1% by weight. In addition, solution of konjac glucomannan is not stable; it tends to gel upon storage, especially at low temperature and/or at basic pH .

So it is the merit of the Applicant to propose a rapid and efficient process for preparing konjac glucomannan hydrolysates , said hydrolysates exhibiting better wettability and higher solubility than parent konjac glucomannan (or native konjac glucomannan) and having high stability (little or no change in viscosity and no gelling upon storage) . In addition the hydrolysates show a small amount of simple sugars (mono and disaccharides ) and high dietary fibers content, which is particularly advantageous for food application.

Summary of the invention

A first object of the present invention is a process for preparing konjac glucomannan hydrolysates. The said process comprises the following steps:

a. hydrolyzing a konjac glucomannan flour in the presence of a food-grade acid, preferably a strong food-grade acid, to provide a partially hydrolyzed konjac glucomannan, the moisture content at the beginning of said acid hydrolysis not exceeding 25 % by weight, preferably between 15 and 22 % by weight, more preferably between 18 and 20 % by weight; b. further hydrolyzing the partially hydrolyzed product of step a) in the presence of at least one enzyme chosen in the group consisting of transglucosidase, β-amylase, pullulanase, maltogenase, glucoamylase, cyclodextrin glycosyltransferase, β-glucanase, cellulase, xylanase and polygalacturonase, preferably in the group consisting of transglucosidase, β- amylase, pullulanase and maltogenase, more preferably in the group consisting of transglucosidase and pullulanase; c. recovering the konjac glucomannan hydrolysates of step b) .

A second object of the present invention is the konjac glucomannan hydrolysate obtained or obtainable by the said process .

A third object of the present invention is the use of the konjac glucomannan hydrolysates for the preparation of a food product or beverage.

As used herein the expression "hydrolysate" means material of lower molecular weight than the parent polysaccharide, and includes, but is not limited exclusively to oligosaccharides and sugars.

Detailed description of the invention Step a) of the process according to the present invention consists of hydrolyzing the konjac glucomannan flour in the presence of a food-grade acid, preferably a strong food-grade acid, under dry conditions.

As used herein the expression "food-grade acid" means that the acid doesn't contaminate food with harmful materials on coming in direct contact or lying nearby.

As used herein the expression "dry conditions" or "dry acid hydrolysis" means that the water content at the beginning of the acid hydrolysis does not exceed 25 % by weight, preferably between 15 % and 22 % by weight, more preferably between 15 % and 20 % by weight.

The dry acid hydrolysis of the present invention is particularly advantageous because there is no need to dissolve the flour to perform the acid hydrolysis, there is less water to remove and no organic solvent is used.

The konjac glucomannan flour useful for the process according to the present invention typically comprises between 60 % and 95 % by weight of glucomannan, for example between 75 and 95% by weight of glucomannan. In a particular embodiment of the present invention, the flour is sold by Hubei Yizhi Konjac Biotechnology Co., Ltd. comprising about 85% by weight of glucomannan with a ratio of glucose to mannose of 1:0.68, 0.6% by weight of proteins, 1.5% by weight of ash, 6.2% by weight of crude cellulose and 0.04% by weight of crude fat.

The food-grade acid useful for the present invention is typically chosen in the group consisting of hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, acetic acid and citric acid, preferably in the group consisting of hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid. The food-grade acid is more preferably hydrochloric acid .

The step a) of dry acid hydrolysis of the present invention more particularly comprises a first step al) of heating the konjac glucomannan flour in the presence of the food-grade acid, preferably the strong food-grade acid, at a temperature between 50 and 100°C, preferably between 60 and 80°C, for at least 1 hour, preferably between 1 and 5 hours, more preferably between 2 and 4 hours.

The temperature during this step al) is preferably at moderately high temperatures, but strictly—preferably below 100°C, in order to avoid an undesirable coloration of the partially hydrolyzed product and/or an excessive depolymerisation or excessive hydrolysis of the konjac glucomannan .

Typically the temperature during this step al) is from 50 to about 100 °C, preferably between 60 and 80 °C. The duration of this step al) is function of the degree of low molecular weight konjac glucomannan of interest, which is also indicated by its solubility and viscosity. In the present invention, the duration of the acid hydrolysis is typically of at least 1 hour, preferably between 1 and 5 hours, more preferably between 2 and 4 hours.

Further, at the end of this step of hydrolysis al) the water content or moisture content is typically between 4 and 8 % by weight, preferably between 4 and 5% by weight so as to avoid an excessive depolymerisation or excessive hydrolysis of the konjac glucomannan. The resulting partially hydrolyzed powder is soluble at and above 20 % solid concentration without forming gel and has pale amber color. Longer acid hydrolysis can result in undesirable darker color.

In a particular embodiment of the present invention, the konjac glucomannan flour is pre mixed with starch before being partially hydrolyzed by means of food-grade acid, preferably strong acid.

The addition of starch exhibits positive effects on the final konjac glucomannan hydrolysates of the present invention, more particularly on the viscosity and solubility when the mixture is dissolved in aqueous solution.

The step a) of dry acid hydrolysis of the present invention further comprises a step a2) of adding water and basic reactant, preferably alkaline reactant, to quench the acid hydrolysis in step al) and to get the partially hydrolyzed product resulting in the form of an aqueous solution with a concentration between 1 and 20 % by weight, preferably between 5 and 15% by weight and a pH between 5 and 6. Above 20% by weight the solution is too thick for subsequent enzyme hydrolysis.

The process of the present invention comprises a further step of hydrolysis in presence of specific enzymes, step b) . This further step, step b) , consists of further hydrolyzing the partially hydrolyzed product of step a) in the presence of at least one enzyme chosen in the group consisting of transglucosidase, β-amylase, pullulanase, maltogenase, glucoamylase, cyclodextrin glycosyltransferase, β-glucanase, cellulase, xylanase and polygalacturonase, preferably in the group consisting of transglucosidase, β-amylase, pullulanase and maltogenase, more preferably in the group consisting of transglucosidase and pullulanase.

Such specific enzymes are particularly interesting since they significantly reduce the molecular weight, thus the viscosity of the partially hydrolyzed product of step a) , without producing too much reducing sugars, more particularly without producing too much monosaccharides (DPI) or disaccharides (DP2) .

Thus, the konjac glucomannan hydrolysates obtained by the process according to the present invention have low amount of monosaccharide/disaccharide (DP1+DP2) and high amount of oligosaccharide/polysaccharide (DP≥3). In other words, the konjac glucomannan hydrolysates comprise low simple sugar content and high fiber content, which is particularly advantageous for healthy and/or nutritious food application.

As used herein the notation DPI, DP2, and DP≥3 refers to the degree of polymerization of the saccharide, i.e., DPI is a degree of polymerization of 1 (monosaccharide), DP2 is a disaccharide, and DP≥3 is a degree of polymerization of 3 or higher .

More precisely, the enzymatic hydrolysis step b) of the present invention comprises the following steps:

bl) bringing the partially hydrolyzed product of step a) (more precisely the partially hydrolyzed product in the form of an aqueous solution of step a2)) into contact with an effective amount of the at least one enzyme; b2) permitting the at least one enzyme to further hydrolyze the partially hydrolyzed product of step a) so as to obtain the konjac glucomannan hydrolysates. The partially hydrolyzed product of step a) is brought into contact with an effective amount of the at least one enzyme. Typically the at least one enzyme is added at a concentration between 0.5 and 5 pL/g, preferably between 0.5 and 1.5 pL/g.

The temperature during the enzymatic hydrolysis (step b) is preferably not high, in order to prevent the denaturation of the enzymes and to operate at a temperature wherein the enzymes exhibit their optimum activities.

Typically the temperature during the enzymatic hydrolysis is from room temperature to about 65 °C, preferably between 50 and 60 °C.

The duration of the enzymatic hydrolysis is function of the degree of low molecular weight konjac glucomannan of interest, which is also indicated by its solubility and viscosity. In the present invention, the duration of the enzymatic hydrolysis (step bl) is typically, of at least 5 hours, preferably between 6 and 30 hours, more preferably between 6 and 20 hours. The process according to the present invention comprises a step c) of recovering the konjac glucomannan hydrolysates.

Such step c) usually or advantageously comprises a step of removing the at least one enzyme, color, and other contaminants such as salts, proteins, ashes, furfurals coming from the acid reaction of sugar or combination thereof. This removal of impurities is typically performed by using filtration, centrifugation, adsorption, distillation, chromatography, washing treatment, or a combination thereof. In a particularly preferred embodiment, step c) of the process of the present invention comprises a first step of filtration the konjac glucomannan hydrolysates resulting from step b) on an activated carbon followed by a filtration on an ion-exchange resin. The activated carbon enables to remove the coloration of the hydrolysates obtained during the acid hydrolysis, the at least enzyme as well as organic impurities such as furfurals from the acid reaction of sugar. The ion- exchange resin removes salts and proteins present in the hydrolysates obtained in step b) .

In a very preferred embodiment, step c) of the process of the present invention comprises or further comprises a step of drying the konjac glucomannan hydrolysates, preferably by spray-drying .

Spray drying is a particularly interesting drying technique in the process because it is a rapid process to dry soluble polysaccharide solution into powder.

According to this very preferred embodiment, the konjac glucomannan hydrolysates are therefore recovered in the form of a powder.

The konjac glucomannan hydrolysates obtained by the process of the present invention exhibit better wettability and higher solubility in water than the parent konjac glucomannan. Therefore aqueous solutions can be prepared at higher solids contents (i.e. with a concentration of konjac glucomannan hydrolysate of more than 1%, preferably more than 2%, more preferably between 2 and 6% by weight), within shorter mixing time, with significantly lower viscosities and without forming gel.

The resulting solutions also show high stability (little or no change in viscosity and no gelling upon storage) .

In addition the hydrolysates show a small amount of simple sugars (mono and disaccharides ) and high dietary fibers content, which is particularly advantageous for healthy and/or nutritious food application.

In a particular embodiment, dextrin or maltodextrin, preferably having a low DE between 10 and 20, can be added to the konjac glucomannan hydrolysates of step b) before the step c) of recovering, more particularly before the step of drying. The dextrin or maltodextrin can be originated from, but not limited to, normal maize starch, waxy maize starch, tapioca starch, wheat starch, potato starch, pea starch, rice starch, etc. Such addition increases the solubility of the konjac glucomannan hydrolysates and allows the adjustment of the viscosity when they are dissolved in aqueous solution.

A second object of the present invention is the konjac glucomannan hydrolysate obtainable or obtained by the process according to the first object.

The konjac glucomannan hydrolysate of the present invention is preferably in the form of a powder.

The konjac glucomannan hydrolysates of the present invention may have typically a mass average molar mass Mw comprised between 24000 and 30000 g/mol, a number average molar mass Mn comprised between 15000 and 20000 g/mol.

As mentioned above, the konjac glucomannan hydrolysates according to the present invention exhibit a good solubility in aqueous medium, in particular in water and the resulting aqueous solution shows low viscosity.

The konjac glucomannan hydrolysates of the present invention when dissolved in an aqueous medium, typically in water at a concentration of 3% by weight show a viscosity not exceeding 50 cP and 35 cP, for example not exceeding 35 cP and 20 cP, said viscosity being measured using a Rapid Visco Analyser (RVA) at 25°C and 65°C, respectively.

The konjac glucomannan hydrolysates of the present invention when dissolved in an aqueous medium at a concentration of 3% by weight, show a viscosity that does not increase more than 15%, preferably does not increase more than 10%, over one month period of time at a temperature of 4°C.

In addition, the konjac glucomannan hydrolysates of the present invention comprise more than 65 % by weight, preferably between 70 and 92 %, for example between 75 and 85%, based on the dry matter of saccharides of degree of polymerization ≥ 3 and a proportion of saccharides of degree of polymerization < 3 less than 20 % by weight, preferably less than 15%, more preferably between 5 and 15% based on the dry matter.

Thus the hydrolysates show a small amount of simple sugars (mono and disaccharides ) and high dietary fibers content, which is particularly advantageous for healthy and/or nutritious food application.

A third object of the present invention is the use of the konjac glucomannan hydrolysates according to the second object for the preparation of a food product or beverage.

The present invention makes now possible the formulation of food products containing high levels of glucomannan, such as nutritional beverages, which could not have been made before because native glucomannans are known to form extremely viscous solutions or gels at concentration higher than 1%. It is also difficult to dissolve a moderate to large amount of konjac flour in water without forming lumps.

The konjac glucomannan hydrolysates of the present invention can be used to stabilize or texturize food and beverage products, such as in the production of ice cream, and as a fat substitute in a variety of reduced fat, low fat and fat free foods, such as soups, porridges, mochi sweets, noodles, cakes, pudding type desserts, sauces, margarine, butter, peanut butter, cream cheese and other spreads, salad dressings, snack dips, mayonnaise, sour cream, yogurt, ice cream, frozen desserts, fudge, soft candies and other confections, skim milk, fruit juices, fruit preserves, jellies, smoothies and sport drinks.

EXAMPLES

Comparative Example 1 : dry acid hydrolysis

A commercially available konjac glucomannan flour was sprayed with 5% HC1 solution to a concentration of 1% (w/w) of pure HC1. The moisture content of said acid sprayed konjac glucomannan flour before hydrolysis was about 15% by weight.

The konjac glucomannan flour used was a flour sold by Hubei Yizhi Konjac Biotechnology Co., Ltd comprising more than 85.3% by weight of glucomannan with a ratio of glucose to mannose of 1:0.68 and further comprises 0.6% by weight of proteins, 1.5% by weight of ash, 6.2% by weight of crude cellulose and 0.04% by weight of crude fat.

The acid sprayed konjac glucomannan flour was well mixed, so as to increase homogeneity of the mixture, and then heated at 70 °C for 3 hours. Then, a solution of the resulting konjac glucomannan hydrolysate was prepared at a concentration of 10% by weight. The pH of the solution was adjusted to 5.8 with 5% NaOH solution to quench the acid hydrolysis.

The hydrolyzed konjac glucomannan solution was purified with activated carbon and ion-exchange resins. The purified konjac glucomannan hydrolysate was spray dried, so as to obtain konjac glucomannan hydrolysate in powder form.

The konjac glucomannan hydrolysate powder was re-dissolved in water at a concentration of 12% by weight. The viscosity of the resulting solution was measured using Rapid Visco-Analyser (RVA) after the time of preparation and after being stored at 4°C 45 days. The RVA heating started with isothermal heating at 25°C for 10 mins, followed by heating from 25°C to 65°C at 2°C/min (a total of 20 min) and finally ending with isothermal heating at 65°C for 10 mins. The stirring speed was kept constant at 160 rpm.

The konjac glucomannan hydrolysate resulting from dry acid hydrolysis produces low viscosity solution at 12% concentration by weight and exhibits no gelling. However, the viscosity increases with storage time, showing that this hydrolysate is not stable during storage time.

Directly after the preparation, the resulting solution had a viscosity (initial viscosity) of about 250 cP at 25°C and 100 cP at 65 °C.

After being stored for 45 days, the resulting solution had a viscosity of about 1000 cP at 25°C and 500 cP at 65°C.

Comparative Example 2 : enzymatic hydrolysis / enzyme selection

Two konjac glucommanan flours with purity of 72.5% and 85.3% were hydrolyzed with a number of enzymes. Each flour was mixed with water to create 1% (w/w) solutions. This concentration was chosen due the difficulty to completely dissolve native konjac flour in water above 1% concentration. Each konjac glucomannan solution (200 g) was equilibrated at 55°C. Then, the pH and viscosity of the konjac glucomannan solution were measured immediately and used as time zero (before enzyme addition) . The viscosity was measured with a Brookfield viscometer using Spindle #4 at 12 rpm.

The characteristics of the two konjac flours are showed in the following table:

size content

91.2% (>120 72.5%

01 mesh, 125 (Standard ≥ 1000 cp 6.0

microns ) 70% )

92.8% (>120 85.3%

02 mesh, 125 (Standard ≥ 21000 cp 6.1

microns ) 85% )

Table 1

Then, 2 pL of enzyme (not diluted) was added to each konjac glucomannan solution.

Eight enzymes used for each of the solutions were respectively maltogenase, pullulanase, β-amylase, transglucosidase, cylodextrin glycosyltransferase, β- glucanase, cellulose and xylanase mixture, and polygalacturonase .

The solutions were incubated at 55°C for 24 hours. The pH and viscosity of the konjac glucomannan solutions were monitored at different time points up to 24 hours. The viscosity was measured with a Brookfield viscometer using Spindle #4 at 12 rpm.

7 7 7

Cyclodetrin pH 5.8 5.91 5.93 5.97 6 6 5.4 glycosyltrans 866. 616. 183.

Viscosity 1433 800 750 550

ferase 7 7 3 pH 5.8 5.87 5.82 5.94 6.01 6.05 5.87 β-Glucanase

Viscosity 1433 33.3 16.7 16.7 16.7 16.7 16.7

Cellulase and pH 5.8 5.86 5.83 5.84 5.89 5.97 5.87 xylanase 233. 133.

mixture Viscosity 1433 100 83.3 66.7 16.7

3 3

Polygalacturo pH 5.8 5.7 5.65 5.74 5.84 5.89 5.76 nase Viscosity 1433 33.3 33.3 16.7 16.7 16.7 16.7

Table 2

Konjac flour type 02 (85.3% purity)

Table 3

From the obtained experimental results, the pH of konjac glucomannan solutions did not show large changes during the incubation. The viscosity of solution from konjac flour with 72.5% purity after incubation is in the following order: control > cyclodextrin glycosyltransferase, β-amylase > maltogenase, pullulanase > transglucosidase, cellulase and xylanase mixture > polygalacturonase, β-glucanase. Whereas, that from konjac flour with 85.3% purity is in the following order: control > cyclodextrin glycosyltransferase > cellulase and xylanase mixture, β-amylase > maltogenase, pullulanase, transglucosidase > polygalacturonase, β-glucanase. Although maltogenase, pullulanase, β-amylase and transglucosidase are commonly used to modify starch and starch derivatives, it appears that these enzymes can be used to hydrolyze konjac glucomannan flour to produce konjac hydrolysates with high solubility, without sacrificing too much of the viscosity and molecular size.

However it is difficult to prepare 1% konjac glucomannan flour solution or higher concentrations because the flour swells instantly when it contacts with water. The flour needs to be added slowly to the water to avoid lump formation, which is not practical for industrial process. Furthermore, the viscosity of konjac glucomannan flour solution at higher concentrations can be too high for effective enzyme reaction. Whereas at 1% solution, there is too much water to be removed to produce konjac glucomannan hydrolysate powder.

Comparative Example 3 : enzymatic hydrolysis / enzyme selection

Konjac glucomannan flour with 72.5% purity was mixed with water to create 0.6% (w/w) solutions. Each solution (100 g) was equilibrated at 55°C for 10 minutes. Then, 40 pL of enzyme (not diluted) was added to each konjac glucomannan solution. Thirteen enzymes used were three types of -amylases (Liquozyme® Supra, Termamyl® 120 L and BAN® 480 L) , maltogenase, pullulanase, glucoamylase, β-amylase, transglucosidase, cyclodextrin glycosyltransferase, β- glucanase, pectinase, cellulose and xylanase mixture, and polygalacturonase .

After the addition of one of the thirteen enzymes, the solutions were incubated at 55°C for 30 minutes in a shaking water bath. The solutions were cooled down at room temperature; then their pH, degree Brix and viscosity were measured. The degree Brix was measured using a refractometer . The viscosity was measured with a Brookfield viscometer using Spindle #3 at 100 rpm. Each solution was also mixed with Fehling's solution (5 mL Fehling's A and 5 mL Fehling's B) and heated at 70°C for 30 mins . The reducing sugars in the solution can react with the Fehling's solution and change the color of Fehling's solution from blue to brown. All three a-amylases and pectinase did not result in any noticeable changes to the viscosity of the konjac glucomannan solution and did not produce substantial amounts of reducing sugars, indicating that these enzymes are not effective to hydrolyze konjac glucomannan solution. The results of other enzymes are represented in the following table .

Glucoamylase 5.99 181 cP +++ β-Amylase 6.05 68 cP None

Transglucosidase 5.99 16 cP +++

Cyclodextrin

6.12 88.0 cP None

glycosyltransferase β-Glucanase 5.34 11.0 cP +++++

Cellulase and xylanase mixture 6.09 17.0 cP None

Polygalacturonase 5.56 11.0 cP +++++ ^a "Unable to detect" means the liquid is too thick for filtration to measure the %Brix. ^b For reducing sugar test, "+" indicates the concentration of reducing sugar. "None" means no any visible reaction.

Table 4

The pH of konjac glucomannan solutions were slightly lower with the addition of enzymes (the decrease in pH is between 0.3 and 1.1) . The viscosity of konjac flour solution was reduced substantially by one of the enzymes of the group consisting of maltogenase, pullulanase, β-amylase, transglucosidase, cyclodextrin glycosyltransferase, and cellulase and xylanase mixture. On the other hand, glucoamylase produced a large amount of reducing sugars without obvious decrease in the viscosity of konjac glucomannan flour solution. Although β- glucanase and polygalacturonase were effective to decrease the viscosity of the konjac glucomannan solution, they produced too much reducing sugars.

Example 1 : dry acid hydrolysis followed by enzymatic hydrolysis Konjac glucomannan flour was sprayed with 5% HC1 solution to a concentration of 1% (w/w) of pure HC1. The flour used was the same as in comparative example 1.

The acid sprayed konjac glucomannan flour was well mixed, so as to increase homogeneity of the mixture, and then heated at 70 °C for 3 hours. Then, a solution of the resulting konjac glucomannan flour was prepared at a concentration of 10% by weight. The pH of the solution was adjusted to 5.8 with 5% NaOH solution to quench the acid hydrolysis. The solution was equilibrated at 55°C for 10 minutes before adding lpL of the enzyme transglucosidase (not diluted) per gram of dry konjac glucomannan hydrolysate.

The resulting solution was incubated at 55°C for 20 hours in a shaking water bath and then cooled down at room temperature.

The hydrolyzed konjac glucomannan solution was purified with activated carbon and ion-exchange resins. The purified konjac glucomannan hydrolysate was spray dried, so as to obtain konjac glucomannan hydrolysate in powder form. The konjac glucomannan hydrolysate powder was re-dissolved in water at a concentration of 3% by weight. The viscosity of the 3% by weight of konjac glucomannan hydrolysate solution was measured before storage and after storage time at 4°C from 1 to 90 days using RVA. The RVA heating started with isothermal heating at 25°C for 10 mins, followed by heating from 25°C to 65°C at 2°C/min (a total of 20 min) and finally ending with isothermal heating at 65°C for 10 mins. The stirring speed was kept constant at 160 rpm.

Before storage, the resulting solution had a viscosity of about 32 cP at 25°C and 18 cP at 65°C (initial viscosity) . After being stored, the viscosity of the konjac flour hydrolysate slightly increased to about 37 cP at 25°C and 20 cP at 65°C (low viscosity) . The viscosity profile showed no obvious changes between 1- and 90-day storage and did not show the presence of gel.

The simple sugar composition of the resulting konjac glucomannan hydrolysate was determined using HPLC. Results are represented in Table 5.

The resulting konjac glucomannan hydrolysate solution (lOOpL) around 0.02 g/mL concentration was injected into an HPLC system equipped with TSK guard column PWXL (Tosoh Corp., 6.0 mm, id 4 cm) and two TSK-GEL G2500PWXL LC columns (Tosoh Corp., 7.8 mm, id 30 cm) connected in series and an RI detector. Glycerol was added into the solution at 1% concentration as an internal standard. Four peaks were observed from the chromatogram for glycerol (internal standard), DPI, DP2, and DP>3. The peak areas of DPI, DP2, and DP≥3 were calculated as a ratio to the peak area of glycerol. The response factor (RF) , which is the peak area ratio between glucose and glycerol as a function of the mass ratio between glucose and glycerol was determined from the chromatograms of 100 pL dextrose and glycerol solution at five different ratios (lO.Omg, 20.0mg, 30.0mg ,40.0mg, and 50.0mg glucose standard in 4 mL 1% glycerol standard solution) . The peak positions of DPI, DP2, and DP≥3 were determined from the chromatogram of 100 pL 1% standard maltodextrin solution.

Table 5 From the HPLC results, it appears that the konjac glucomannan hydrolysate obtained from the process according to the invention has a low amount of monosaccharide/disaccharide (DP1+DP2<9%) and a high amount of oligosaccharide/polysaccharide (DP3>91%) . In other words, it means that the konjac glucomannan hydrolysate obtained from the process according to the invention comprises low sugar content. The konjac glucomannan hydrolysate also has high fiber content (-81% according to AOAC Method 2001.03). The properties of the konjac glucomannan hydrolysate obtained from the process according to the invention are particularly advantageous for food application and more particularly on its viscosity stability during processing and storage.

Example 2 : dry acid hydrolysis followed by enzymatic hydrolysis / maltodextrin

Konjac glucomannan hydrolysates were prepared according to the process of the invention in which maltodextrin was added after the purification step of the konjac glucomannan hydrolysate, in the following way:

Konjac glucomannan flour was sprayed with 5% HC1 solution to a concentration of 1% of pure HC1. The flour used was the same as in example 1.

The acid sprayed konjac glucomannan flour was well mixed, so as to increase homogeneity of the mixture, and then heated at 70 °C for at least 3 hours. Then, a solution of the resulting konjac glucomannan flour was prepared at a concentration of 10% by weight. The pH of the solution was adjusted to 5.8 with 5% NaOH solution to quench the acid hydrolysis. The solution was equilibrated at 55°C for 10 minutes before adding 1 pL of the enzyme transglucosidase (not diluted) per gram of dry konjac glucomannan hydrolysate.

The resulting solution was incubated at 55°C for 20 hours in a shaking water bath and then cooled down at room temperature. The hydrolyzed konjac glucomannan solution was purified with activated carbon and ion-exchange resins. The purified konjac glucomannan hydrolysate was mixed with maltodextrin having DE of 12 at a ratio of 1:1. The maltodextrin-konj ac glucomannan hydrolysate mixture solution was spray dried, so as to obtain the mixture in powder form. The konjac glucomannan hydrolysate powder mixture was re-dissolved in water at a concentration 6% by weight.

The viscosity of the 6% by weight of resulting mixture solution was measured after storage time at 4°C from 1 to 45 days using RVA. The RVA heating started with isothermal heating at 25°C for 10 mins, followed by heating from 25°C to 65°C at 2°C/min (a total of 20 min) and finally ending with isothermal heating at 65°C for 10 mins. The stirring speed was kept constant at 160 rpm.

From the viscosity results, it appeared that the konjac glucomannan hydrolysate - maltodextrin mixture had more or less the same viscosity profile before storage than after storage. The konjac glucomannan hydrolysate - maltodextrin mixture remained therefore stable over time.

The konjac glucomannan hydrolysate - maltodextrin mixture exhibited very low viscosity and no gelling over time.

The viscosity showed that the konjac glucomannan hydrolysate - maltodextrin mixture has a viscosity between 20 and 25 cP at 25°C and a viscosity between 15 and 20 cP at 65°C. Example 3 : dry acid hydrolysis followed by enzymatic hydrolysis from a mixture of corn starch - konjac glucomannan flour in proportion 9 : 1 Konjac glucomannan flour was mixed with corn starch in proportion of 1:9.

Corn starch - konjac glucomannan flour mixture was sprayed with 5% HC1 solution to a concentration of 1% of pure HC1. The moisture content during said acid sprayed mixture is maintained at about 20 % by weight.

The acid sprayed corn starch - konjac glucomann flour mixture was well mixed, so as to increase homogeneity of the mixture, and then heated at 70 °C for 3 hours. Then, a solution of the resulting corn starch - konjac glucomannan flour mixture was prepared at a concentration of 30% by weight. The pH of the mixture solution was adjusted to 5.8 with 5% NaOH solution to quench the acid hydrolysis. The mixture solution was equilibrated at 55°C for 10 minutes before adding the enzymes -amylase and transglucosidase (not diluted) at a concentration of lpL of each enzyme per gram of dry corn starch - konjac glucomannan mixture.

The resulting solution was incubated at 55°C for 20 hours in a shaking water bath and then cooled down at room temperature . The hydrolyzed corn starch - konjac glucomannan mixture solution was purified with activated carbon and ion exchange resins. The purified corn starch - konjac glucomannan mixture hydrolysate was spray dried, so as to obtain corn starch - konjac glucomannan mixture hydrolysate in powder form. The corn starch - konjac glucomannan mixture hydrolysate powder was re-dissolved in water at a concentration of 10% by weight. The viscosity of the 10% by weight of corn starch - konjac glucomannan mixture hydrolysate solution was measured before storage and after storage time at 4°C for 2 to 42 days by RVA. The RVA heating started with isothermal heating at 25°C for 10 mins, followed by heating from 25°C to 65°C at 2°C/min (a total of 20 min) and finally ending with isothermal heating at 65°C for 10 mins. The stirring speed was kept constant at 160 rpm .

From the viscosity results, it appeared that the corn starch - konjac glucomannan mixture hydrolysate at 10% concentration had approximatively the same viscosity profile before storage than after short-term storage, which showed viscosity of about 20 cP at 25°C and about 15 cP at 65°C. The viscosity after long-term storage (e.g. after 42 days) was slightly lower showing viscosity of about 15 cP at at 25°C and about 12 cP at at 65°C. The corn starch - konjac glucomannan mixture hydrolysate solution in general remained stable over cold storage time. The corn starch - konjac glucomannan mixture hydrolysate also exhibited low viscosity and no gelling over storage time.

The polymerization degree (DP) of corn starch - konjac glucomannan mixture hydrolysate was determined by HPLC (using the same method as described in example 1) . Results are shown in Table 6.

Table 6

From the HPLC results, it appears that the corn starch - konjac glucomannan mixture hydrolysate obtained from the process according to the invention had lower amount of monosaccharide/disaccharide (DP1+DP2<35%) and higher amount of oligosaccharide/polysaccharide (DP3>65%) . In other words, it means that the konjac glucomannan flour can be hydrolyzed with acid and enzyme in the presence of starch using the process according to the invention to produce konjac glucomannan hydrolysate and maltodextrin mixture, which is particularly advantageous for food application and more particularly on its viscosity stability during processing and storage.

Claims

1. A process for preparing konjac glucomannan hydrolysates said process comprising the following steps: a. hydrolyzing a konjac glucomannan flour in the presence of a food-grade acid, preferably a strong food-grade acid, to provide a partially hydrolyzed konjac glucomannan, the moisture content at the beginning of said acid hydrolysis not exceeding 25 % by weight, preferably between 15 and 22 % by weight, more preferably between 18 and 20 % by weight ; b. further hydrolyzing the partially hydrolyzed product of step a) in the presence of at least one enzyme chosen in the group consisting of transglucosidase, β-amylase, pullulanase, maltogenase, glucoamylase, cyclodextrin glycosyltransferase, β-glucanase, cellulase, xylanase and polygalacturonase, preferably in the group consisting of transglucosidase, β-amylase, pullulanase and maltogenase, more preferably in the group consisting of transglucosidase and pullulanase; c. recovering the konjac glucomannan hydrolysates of step b) .

2. The process according to claim 1, wherein said acid hydrolysis of step a) comprises the following steps: al . heating the konjac glucomannan flour, optionally pre mixed with starch, in the presence of the food-grade acid at a temperature between 50 and 100°C, preferably between 60 and 80°C, for at least 1 hour, preferably between 1 and 5 hours, more preferably between 2 and 4 hours;

a2. adding water and basic reactant, preferably alkaline reactant, to quench the acid hydrolysis and to get the partially hydrolyzed product resulting in the form of an aqueous solution with a concentration between 1 and 20 % by weight, preferably between 5 and 15% by weight and a pH between 5 and 6.

3. The process according to either claim 1 or 2, wherein said enzymatic hydrolysis of step b) comprises the following steps: bl . bringing the partially hydrolyzed product of step a) into contact with an effective amount of the at least one enzyme; b2. permitting the at least one enzyme to further hydrolyze the partially hydrolyzed product of step a) so as to obtain the konjac glucomannan hydrolysates.

4. Process according to anyone of claims 1 to 3, wherein the konjac glucomannan flour comprises between 60% and 95% by weight of glucomannan.

5. Process according to anyone of claims 1 to 4, wherein the food-grade acid of step a) is chosen in the group consisting of hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, acetic acid and citric acid, preferably in the group consisting of hydrochloric acid, sulfuric acid, nitric acid phosphoric acid, more preferably is hydrochloric acid.

6. The process according to anyone of claims 1 to 5, wherein step c) of recovering the konjac glucomannan hydrolysates comprises a step of removing the at least one enzyme, color, and other contaminants from the konjac glucomannan hydrolysates by filtration, centrifugation, adsorption, distillation, chromatography, washing treatment, or a combination thereof, for example by filtration on activated carbon and ion-exchange resins.

7. The process according to anyone of claims 1 to 6, wherein step c) of recovering the konjac glucomannan hydrolysates comprises a step of drying the konjac glucomannan hydrolysates, preferably by spray-drying.

8. The process according to claim 7, wherein dextrin or maltodextrin is added to the konjac glucomannan hydrolysates before the step of drying.

9. Konjac glucomannan hydrolysates obtainable by the process as claimed in anyone of claims 1 to 8.

10. The konjac glucomannan hydrolysates as claimed in claim 9 being in the form of a powder.

11. The konjac glucomannan hydrolysates as claimed in anyone of claims 9 to 10 or obtained by the process as claimed in anyone of claims 1 to 8 comprising more than 65 % by weight, preferably between 70 and 92%, for example between 75 and 85 % based on the dry matter of saccharides of degree of polymerization ≥ 3 and a proportion of saccharides of degree of polymerization < 3 less than 20 % by weight, preferably less than 15 %, more preferably between 5 and 15% based on the dry matter.

12. The konjac glucomannan hydrolysates as claimed in anyone of claims 9 to 11 or obtained by the process as claimed in anyone of claims 1 to 8 when dissolved in an aqueous medium, typically in water at a concentration of 3% by weight show a viscosity not exceeding 50 cP and 35 cP, said viscosity being measured using a Rapid Visco Analyzer at 25°C and 65°C, respectively .

13. The konjac glucomannan hydrolysates as claimed in anyone of claims 9 to 12 or obtained by the process as claimed in anyone of claims 1 to 8 when dissolved in an aqueous medium at a concentration of 3% by weight, typically in water show a viscosity that does not increase more than 15% over one month period of time at a temperature of 4°C.

14. Use of the konjac glucomannan hydrolysates as claimed in anyone of claims 9 to 13 or obtained by the process as claimed in anyone of claims 1 to 8 for the preparation of a food product or beverage.