JP7325821B2 - METHOD FOR EVALUATING THE BODIES OF DIETARY FIBER IN MAMMALIAN GUT OR ORGANIC ACID PRODUCTION - Google Patents

Description

TECHNICAL FIELD The present invention relates to a method for evaluating the ability of dietary fiber to decompose in mammalian intestines or the ability to produce organic acids. More specifically, the present invention can evaluate the intestinal kinetics of dietary fiber, which is one of the food ingredients, in the intestines of mammals such as humans. It relates to a method for evaluating acid-producing ability.

In the human intestinal tract, a wide variety of bacteria are constantly proliferating, and these are called intestinal microflora. In the study of intestinal microflora, culture using an intestinal mimetic culture device that reproduces the human intestinal environment is used. A culturing method and an apparatus capable of culturing while maintaining the structural balance of the intestinal flora have been proposed (Patent Documents 1 and 2, and Non-Patent Document 1).

Gut-mimetic cultures can be used, for example, to assess the effects of food ingredients (eg, dietary fiber such as inulin) on the intestinal flora (Patent Document 1 and Non-Patent Document 1). In addition, examination of ulcerative colitis and screening of therapeutic agents for ulcerative colitis using intestinal mimetic culture have also been reported (Patent Document 2).

By the way, prebiotic foods are attracting attention as useful foods for adjusting the intestinal microflora so as to constitute healthy human intestinal microflora. Prebiotic foods "feed" indigenous intestinal bacteria (beneficial bacteria) that favor the host, and by regulating their growth or activity, they have a beneficial effect on the host and improve the host's health. .

However, there are still many unknowns about the intestinal dynamics of such prebiotic foods, and various technological developments that are useful for application to commercialization of foods, pharmaceuticals, etc. are desired.

WO2015/136916 JP 2018-198560 A

Enterobacteriology journal 31 (2) 104 (2017) "Effect of indigestible dietary fiber on intestinal bacteria using cultured human intestinal model"

An object of the present invention is to solve the above-mentioned problems, and an object of the present invention is to easily and efficiently grasp the intestinal dynamics of dietary fiber in mammals. To provide a method for evaluating the decomposition performance or organic acid production performance of

The present invention provides a method for evaluating the degradation performance of dietary fiber in the mammalian intestine, the method comprising:
(a) a step of anaerobic culture using feces from a healthy individual to construct an intestinal flora;
(b) adding the dietary fiber to the culture obtained in the step (a) and further anaerobic culturing; and (c) adding the dietary fiber in the culture obtained in the step (b) Including measuring the degradation rate and/or degradation rate.

In one embodiment, the anaerobic culture of step (a) above is performed for 16 to 24 hours.

In one embodiment, said mammal is a human.

The present invention provides a method for evaluating the ability of dietary fiber to produce organic acids in the intestine of mammals, the method comprising:
(i) a step of anaerobic culture using feces from a healthy individual to construct an intestinal flora;
(ii) adding the dietary fiber to the culture obtained in step (i) and further anaerobic culturing; and (iii) adding the organic acid in the culture obtained in step (ii). including the step of measuring the amount;
The organic acid is acetic acid and/or propionic acid.

In one embodiment, the anaerobic culture of step (i) above is performed for 16 to 24 hours.

In one embodiment, said mammal is a human.

The present invention provides a method for screening dietary fiber having organic acid-producing ability in the intestine of mammals, the method comprising:
Determining the total ratio (A) of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber, and the number of 1→2 glycosidic bonds to the total number of glycosidic bonds of the standard dietary fiber group and Using a relative curve showing the correlation between the total ratio of 1→3 glycosidic bond numbers (B) and the production of organic acids in the mammalian intestine of the standard dietary fiber group, from the ratio (A) a step of estimating the ability of the dietary fiber to produce organic acids in the intestine of the mammal;
including
The organic acid is acetic acid and/or propionic acid.

The present invention provides a method for predicting the degradation performance of dietary fiber in the mammalian intestine, the method comprising:
A step of determining the total ratio (A) of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the dietary fiber, and the number of 1→2 glycosidic bonds to the total number of glycosidic bonds of the standard dietary fiber group and 1 → 3 glycosidic bond number total percentage (B) and the degradation rate and / or degradation rate in the mammalian intestine of the standard dietary fiber group using a relative curve showing the correlation, the percentage ( estimating the intestinal degradation rate and/or degradation rate of the dietary fiber from A);
including.

INDUSTRIAL APPLICABILITY According to the present invention, the intestinal degradation rate and/or degradation rate and organic acid production ability can be easily grasped as the dynamics of dietary fiber in the large intestine in mammals. The kinetics of dietary fiber obtained in this way can be compared with the kinetics of other dietary fibers in the same mammal, and these results can be used to predict the intestinal fermentability of dietary fiber and its functionality as a prebiotic food. can be used as information for

It is a graph which shows the production amount of (a) acetic acid, (b) propionic acid and (c) butyric acid when cultured for 24 hours after adding various dietary fibers (42 hours after culture). It is a graph which shows (a) decomposition rate and (b) decomposition rate of various dietary fibers. It is a graph which shows the relationship of (a) decomposition rate and (b) decomposition rate of dietary fiber with respect to acetic acid concentration. It is a graph which shows the relationship between (A) decomposition rate and (B) decomposition rate of dietary fiber with respect to propionic acid concentration. It is a graph which shows the relationship of (A) decomposition rate and (B) decomposition rate of dietary fiber with respect to butyric acid concentration. A graph showing the relationship between (a) acetic acid concentration, (b) propionic acid concentration and (c) butyric acid concentration with respect to the ratio of the total number of 1 → 2 glycosidic bonds and the number of 1 → 3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. be. 1 is a graph showing the relationship between (a) degradation rate and (b) degradation rate of dietary fiber with respect to the ratio of the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. The results of (a) number of sequences (number of reads) obtained by next-generation sequencing, (b) number of OTUs, (c) Shannon-Wiener diversity index and (d) Simpson index are shown. 1 is a graph showing abundance ratios of various enterobacteriaceous phyla relative to total enterobacteria in stool samples and culture solutions.

(Definition of terms)
“Prebiotic food” is defined as “food” for indigenous bacteria (beneficial bacteria) that work favorably for the host, and by regulating their growth or activity, it has a beneficial effect on the host, indigestible foods that improve health”. Prebiotic foods include oligosaccharides (including indigestible oligosaccharides), resistant starch, dietary fiber, and the like.

"Mammals" include humans; pets such as dogs and cats; animals for testing and research such as mice and rats; and farm animals such as pigs. In the present invention, "mammals" preferably do not include herbivores that mainly eat herbs. A "mammal" in the present invention is preferably a human.

In the present invention, the term "dietary fiber" refers to a single substance containing a glycosidic bond in its constituent molecule, which is not digested by digestive enzymes possessed by at least some animals included in the above mammals, or Refers to substances that are difficult to digest. It is also called indigestible component or indigestible substance. Typical examples of dietary fibers include polysaccharides. Dietary fiber may be, for example, a substance naturally derived from plants, animals, microorganisms, etc., or a chemically synthesized substance. Glycosidic bonds contained in molecules constituting dietary fiber include, for example, in the case of polysaccharides, 1→4 glycosidic bonds, 1→6 glycosidic bonds, 1→2 glycosidic bonds, and A 1→3 glycosidic bond is included. When dietary fiber is decomposed in the large intestine of mammals, it becomes a substrate for anaerobic fermentation by intestinal bacteria, and organic acids are produced as final metabolites. Such organic acids include short chain fatty acids such as acetic acid, propionic acid and butyric acid. The property of dietary fiber to become a fermentation substrate for intestinal bacteria in the large intestine is also called "fermentability".

"Total number of glycosidic bonds" refers to the total number of glycosidic bonds contained in a molecule (one molecule) constituting dietary fiber. The "total number of glycosidic bonds" can be calculated from, for example, the type and structure of molecules that constitute dietary fiber, the structure of constituent sugars, the molecular weight, and the like. Alternatively, the "total number of glycosidic bonds" can be obtained by, for example, using a gas chromatograph-mass spectrometer to determine the binding positions of constituent sugars through methylation analysis in the molecule, even if the structures of the constituent molecules are unknown. can be obtained by

"The ratio of the total number of 1 → 2 glycosidic bonds and the number of 1 → 3 glycosidic bonds to the total number of glycosidic bonds" refers to the total number of glycosidic bonds in a molecule (one molecule) constituting dietary fiber, 1 in the molecule → Refers to the total ratio of the number of 2 glycosidic bonds and the number of 1→3 glycosidic bonds. The "number of 1→2 glycosidic bonds" and "number of 1→3 glycosidic bonds" can be calculated from, for example, the types and structures of molecules constituting dietary fiber, the structures of constituent sugars, molecular weights, and the like. can be obtained, for example, by ascertaining the binding positions of constituent sugars by methylation analysis in the molecule using a gas chromatograph-mass spectrometer, even if the structure of the constituent molecule is unknown.

“Intestinal flora” refers to a group of bacteria that normally reside in the large intestine of mammals such as humans. Bacteria that constitute the intestinal flora of healthy humans include, for example, bacteria belonging to the following phyla at the phylum level: Verrucomicrobia, Proteobacteria, Fusobacteria ( Fusobacteria), Firmicutes, Bacteroidetes, Actinobacteria, and the like. At a further level, bacteria belonging to the following genera or families are included: Bifidobacterium, Collinsella, Bacteroides, Parabacteroides, Prevotella, Rikenellaceae, Lactobacillales, etc.

A "healthy individual" refers to a mammalian individual who is not afflicted with disease. Individuals who have not received antibiotics for at least two months prior to the date of stool collection are preferred.

The “standard dietary fiber group” means that the organic acid production amount or the decomposition rate and / or decomposition rate of dietary fiber has been measured under the same culture conditions or is known, and 1 → 2 Refers to a group composed of a plurality of dietary fibers for which the ratio of the total number of glycosidic bonds and 1→3 glycosidic bonds has been calculated or can be calculated.

Both "degradation rate" and "degradation rate" serve as indices representing the degree of reduction in dietary fiber caused by bacterial degradation of dietary fiber in the presence of intestinal flora. "Degradation rate" is the amount of dietary fiber (weight) _D1 at the start time _T1 when dietary fiber is placed in the presence of intestinal flora, and the amount of dietary fiber remaining at time _T2 after a certain period of time. (Weight) Refers to the ratio (%) of _D2 , which is represented by the formula _D2 / _D1x100 . The "degradation rate" can be expressed, for example, as a ratio of the final dietary fiber amount (weight) remaining at the end of the culture to the initial dietary fiber amount (weight) at the start of the culture in the presence of intestinal flora. When dietary fiber is placed in the presence of intestinal flora, the "degradation rate" is the amount of dietary fiber (weight) at the start time _T1 , and the amount of dietary fiber (weight) at time _T2 after a certain period of time has elapsed from _D1 . The amount of change per unit time (for example, the amount of decrease) up to D ₂ is represented by the formula (D ₁ -D ₂ )/(T ₂ -T ₁ ). The "degradation rate" can also take into account the volume of the culture solution when culturing in the presence of intestinal flora. "Degradation rate" is, for example, the amount of change from the initial amount of dietary fiber (weight) at the start of culture in the presence of intestinal flora to the final amount of dietary fiber (weight) remaining at the end of culture per volume of culture solution and It can be expressed as an amount per unit time (for example, the amount decreased by decomposition of dietary fiber).

For example, according to the evaluation method of the present invention, the production amount of dietary fiber organic acid or the decomposition rate and/or decomposition rate of dietary fiber is measured, and can be used as the dietary fiber constituting the standard dietary fiber group.

(Method for Evaluating Decomposition Performance of Dietary Fiber in Mammal Intestines)
The method of evaluating the degradation performance of dietary fiber in mammalian intestines of the present invention includes the steps of (a) performing anaerobic culture using feces derived from a healthy individual to construct an intestinal flora; (b) the step (a) adding the dietary fiber to the culture obtained in step (a) and further anaerobic culturing; and (c) the decomposition rate and/or of the dietary fiber in the culture obtained in step (b) Including the step of measuring the decomposition rate.

The culturing of step (a) establishes an intestinal flora that mimics the mammalian large intestine environment of the feces used. In step (a), a culture may be prepared by adding feces from a healthy individual to the culture medium. Any culture medium may be used as long as it can be used for anaerobic culture. Stool can be prepared in advance as a suspension in a predetermined solvent and added to the medium. GAM medium, for example, is used as the anaerobic culture medium. GAM medium includes, for example, GAM agar medium, modified GAM agar medium, GAM semi-fluid stratified medium, GAM bouillon and modified GAM bouillon. Before culturing, the medium can be sterilized (sterilization), for example using an autoclave. Liquid culture is preferred, and the liquid culture is also referred to as "culture solution". During culturing, the culture medium can be appropriately agitated. After collection, feces can be stored in a container, such as an anaerobic culture swab, until culturing is initiated.

The anaerobic environment of the culture can be created by aerating the medium with anaerobic gases. Anaerobic gases are, for example, nitrogen, nitrogen and carbon dioxide, or nitrogen and carbon dioxide and hydrogen. Anaerobic gas aeration is performed constantly or intermittently at a predetermined flow rate (eg, 0.1 to 1.0 dL/min). Further, the anaerobic gas is preferably a mixed gas composed of nitrogen and carbon dioxide, since intestinal gas may contain, for example, nitrogen and carbon dioxide. In order to maintain highly anaerobic conditions, it is preferable to constantly aerate anaerobic gas.

In the present invention, in anaerobic culture, the pH of the culture solution is preferably set to 6.2 to 6.7, more preferably 6.2 to 6.5 at the start of the culture. By adjusting the pH within the above range at the start of culture (for example, when the culture solution containing feces is placed in an anaerobic environment), it is possible to match the pH in the large intestine of mammals corresponding to the feces used. can. It is sufficient that the pH of the culture solution at the start of the culture is within the above range, and thereafter the pH may be left as it is without any particular adjustment of the pH, or the pH may be adjusted as necessary to prevent an extreme drop in the pH. The pH may be adjusted within the above range by agents. Preferably, the pH is left as it is after the start of the culture. "Leave alone" with respect to pH means that no operation intended to maintain the pH within the above range (for example, pH adjustment by addition of a pH adjuster or alkali) is performed during cultivation.

As the culture temperature, a temperature close to the body temperature of the mammal is preferably used because it mimics the environment inside the large intestine of the mammal corresponding to the feces used. For example, when using human feces, the culture temperature is preferably 36° C. to 38° C., more preferably 36° C. to 37° C., because it is close to the body temperature of healthy humans.

The method of culturing is not limited, but a one-time batch method is preferred.

The culture period in step (a) is preferably a period during which intestinal flora can be established. By constructing the intestinal flora, the ability of bacteria of the intestinal flora to decompose dietary fiber can be exhibited. The culture period in step (a) of addition is, for example, greater than 12 hours, preferably 16-24 hours, more preferably 18-24 hours. When the culture period of step (a) is 12 hours or less, the growth of intestinal bacteria is insufficient and the bacterial flora is not sufficiently established, and the decomposition of dietary fiber or the production of organic acids by bacteria in the bacterial flora is observed. It may not be done as much as possible. When the culture period of step (a) exceeds 24 hours, the intestinal flora is sufficiently established within that time period to the extent that the decomposition of dietary fiber or the production of organic acids by bacteria can be observed, and the evaluation method It may lengthen the entire period of

In step (b), dietary fiber is added as a test substance to the culture obtained in step (a), and anaerobic culture is performed. Dietary fiber, which is a test substance, is also referred to as “test dietary fiber”. The dietary fiber to be tested is added, for example, at 6 g to 12 g, preferably 8 g to 10 g, per 1 L of culture medium. The above addition amount range is an amount more suitable for measuring the decomposition of dietary fiber and the amount of organic acid produced. In step (b), the anaerobic culture of step (a) is continued except for the addition of dietary fiber. The culture period in step (b) can be appropriately determined based on the length of stay of solid food in the digestive tract of the subject animal. For example, when the mammal is a human (adult), the culture period in step (b) is, for example, 20 to 30 hours, preferably 20 to 30 hours, taking into account the residence time of food (solid matter) in the human digestive tract. is 23-25 hours.

With respect to step (c), the degradation rate and percent can be determined by contrasting the amount of dietary fiber added and the amount of dietary fiber in the culture obtained in step (b). The culture may be solid-liquid separated using techniques known in the art (eg, filtration) and the supernatant obtained may be used for measurement. The degradation rate can be expressed, for example, as the amount of dietary fiber in the culture (culture solution) per hour (g/L/h). The decomposition rate can be expressed, for example, as the ratio (%) of the amount of dietary fiber remaining at the end of culture with added dietary fiber to the amount of added dietary fiber. The amount of dietary fiber in fecal samples or cultures can be measured using methods well known to those skilled in the art. Such methods include, for example, an enzyme-gravimetric method and a method combining an enzyme-gravimetric method and high performance liquid chromatography (HPLC) (an enzyme-HPLC method).

Thus, by determining the degradation rate and/or degradation rate of dietary fiber in vitro by the method of the present invention, the rate of intestinal fermentation by that dietary fiber in vivo and/or the amount of dietary fiber utilized for intestinal fermentation You can guess the percentage. A high decomposition rate of dietary fiber is an index of high intestinal decomposition performance. When the intestinal decomposition performance is high, the dietary fiber is digested and decomposed in the intestine, and anaerobic fermentation can be promoted. As shown in the examples below, in the fecal culture of the present invention, the higher the rate of decomposition of dietary fiber, the higher the amount of organic acid produced. A low rate of dietary fiber degradation is an indicator of low intestinal degradation performance in that mammal. When the intestinal decomposition performance is low, the dietary fiber remains in the intestine of the mammal without being decomposed, and therefore tends to be excreted as stool without being digested in the body. Therefore, dietary fiber with a high decomposition rate is expected to increase the production of organic acids in intestinal fermentation, and dietary fiber with a slow decomposition rate is expected to increase defecation.

(Method for Evaluating Organic Acid Producing Ability of Dietary Fiber in Mammal Intestines)
The method for evaluating the organic acid-producing ability of the dietary fiber in the mammalian intestine of the present invention includes the steps of (i) anaerobic culture using feces from a healthy individual to construct an intestinal flora; (ii) adding the dietary fiber to the culture obtained in step (i), and further anaerobic culturing; and (iii) measuring the amount of the organic acid in the culture obtained in step (ii). including the step of Here, "organic acid" is acetic acid and/or propionic acid.

Steps (i) and (ii) are the same as steps (a) and (b) above.

Regarding step (iii), the amount of organic acid produced by adding dietary fiber is, for example, the amount of organic acid at the end of culture in step (ii), and the amount of organic acid at the end of culture under the same conditions except for the addition of dietary fiber. can be expressed as a percentage of The organic acid concentration in the supernatant obtained by solid-liquid separation of the culture by techniques known in the art (eg, filtration) may be measured. The organic acid concentration can be measured, for example, by high performance liquid chromatography (HPLC).

By determining the amount of organic acid produced by the addition of dietary fiber in vitro in this way, the intestinal fermentability of the dietary fiber (for example, the ability to produce organic acid by intestinal fermentation) can be estimated.

(Method for Screening Dietary Fiber Having Ability to Produce Organic Acid in Mammalian Intestine)
INDUSTRIAL APPLICABILITY According to the present invention, there is provided a method for screening dietary fibers having an organic acid-producing ability in mammalian intestines. Organic acids are acetic acid and/or propionic acid.

In the above screening method, the ability to produce organic acids is estimated based on the ratio of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. This inference is, for example, as a relative evaluation to the value measured under the same culture conditions as the standard dietary fiber group, as a result of the tendency to show the high or low organic acid productivity (for example, the amount of organic acid produced per unit time). can get. "High and low" can be classified, for example, based on the intermediate value between the highest organic acid concentration and the lowest organic acid concentration of all dietary fibers constituting the standard dietary fiber group.

The total number of glycoside bonds, the number of 1→2 glycoside bonds and the number of 1→3 glycoside bonds of dietary fibers and dietary fibers constituting the standard dietary fiber group are analyzed by sugar chain structure analysis, for example, using a gas chromatograph mass spectrometer. can be determined by doing. Then, based on the total number of glycosidic bonds, the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds thus obtained, the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds relative to the total number of glycosidic bonds A percentage of the total can be calculated.

The intestinal organic acid-producing ability of dietary fibers constituting the standard dietary fiber group can be estimated, for example, based on the evaluation method of the organic acid-producing ability of the present invention. The organic acid production ability is evaluated by culturing a plurality of dietary fibers constituting the standard dietary fiber group under the same conditions.

The relationship between the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in dietary fiber and the ability of the dietary fiber to produce organic acids is, for example, Acetic acid and propionic acid levels may decrease. For this reason, the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the organic acid productivity of the standard dietary fiber group If a "relative curve" indicating the correlation is created, the production amount of organic acids for a given dietary fiber can be confirmed using a computational method based on the "relative curve" without necessarily using an experimental method. can be inferred. Examples of such a "relative curve" include, for example, diagrams and graphs showing the relationship between the ratio (B) and the production amount of organic acids in the standard dietary fiber group; A computer program that incorporates

The "relative curve" is created, for example, as follows. First, for multiple types of dietary fibers that have been measured or are known for their ability to produce organic acids, their ratios of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of those dietary fibers Organic acid productivity is plotted. Then, based on this plotted information, an approximate curve or approximate expression is created or calculated using techniques known to those skilled in the art. It is not particularly limited whether the approximated curve obtained in this way has linearity or nonlinearity. In the present invention, for example, the approximated curve (approximated straight line) and the determinant (R ² ) may be determined based on the method of least squares. In the present invention, when an approximate curve based on the method of least squares is used as the relative curve, the obtained determinant (R ² ) is preferably 0, because it is possible to grasp more accurate organic acid productivity. It is preferred to use an approximation curve in the range of 0.7 to 1, more preferably 0.8 to 1. Using the relative curve created in this way, the ratio (A) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber is used to calculate the organic acid production capacity of the dietary fiber. is presumed.

From the organic acid-producing ability estimated based on the relative curve, it is possible to predict the tendency of the organic acid-producing ability in the intestine of the mammal to increase or decrease. As a result, it becomes possible to screen for dietary fibers capable of producing organic acids in the intestines of mammals, and to predict the fermentability of dietary fibers in the intestines of mammals.

(Method for Predicting Decomposition Performance of Dietary Fiber in Mammalian Intestines)
According to the present invention, a method for predicting the degradation performance of dietary fiber in mammalian intestines is provided.

In the above prediction method, the decomposition rate and/or decomposition rate of dietary fiber in mammalian intestines is estimated based on the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. be done. The value obtained by this estimation can be used, for example, as a relative value to the value measured under the same culture conditions as the standard dietary fiber group.

The decomposition rate and decomposition rate of the dietary fibers constituting the standard dietary fiber group can be determined, for example, based on the decomposition performance evaluation method of the present invention. The decomposition performance can be evaluated by, for example, culturing a plurality of dietary fibers constituting the standard dietary fiber group under the same conditions.

The ratio of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fibers and dietary fibers constituting the standard dietary fiber group is determined in the same manner as described above.

The relationship between the ratio of the total number of 1 → 2 glycosidic bonds and the number of 1 → 3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber and the decomposition rate and decomposition rate of the dietary fiber is, for example, as the ratio increases The rate and rate of degradation of dietary fiber may be decreased. For this reason, the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group, and the decomposition rate and decomposition rate of dietary fiber in the standard dietary fiber group If a "relative curve" showing the correlation between can be inferred using a computational method by Examples of such a "relative curve" include diagrams and graphs showing the relationship between the ratio (B) and the decomposition rate and decomposition rate of the dietary fiber of the standard dietary fiber group, and mathematical representations thereof. Examples include mathematical formulas and computer programs incorporating them.

The "relative curve" is created, for example, as follows. First, for multiple types of dietary fiber that have been measured or known regarding the degradation rate and rate of dietary fiber, the ratio of the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds to the total number of glycosidic bonds of those dietary fibers Degradation rates and percentages of those dietary fibers are plotted against . Then, based on this plotted information, an approximate curve or approximate expression is created or calculated using techniques known to those skilled in the art. It is not particularly limited whether the approximated curve obtained in this way has linearity or nonlinearity. In the present invention, for example, the approximated curve (approximated straight line) and the determinant (R ² ) may be determined based on the method of least squares. In the present invention, when the approximate curve based on the method of least squares is used for the relative curve, the determinant (R ² ) obtained is because it is possible to grasp the decomposition rate and decomposition rate of dietary fiber more accurately. Preferably, an approximation curve within the range of 0.7 to 1, more preferably 0.8 to 1 is used. Using the relative curve created in this way, the ratio (A) of the total number of 1 → 2 glycosidic bonds and the number of 1 → 3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber is calculated. and decomposition rates are estimated.

From the prediction of the degradation rate and/or degradation rate obtained based on the relative curve, it is possible to predict the trend of high or low degradation rate and/or degradation rate of dietary fiber in the mammalian intestine. This makes it possible to screen for dietary fibers that have the ability to produce organic acids in the intestines of mammals, to predict the intestinal fermentability of dietary fibers, or to increase defecation.

INDUSTRIAL APPLICABILITY According to the present invention, the degradation rate and/or decomposition rate in the intestine of a mammal and the ability to produce organic acids can be easily evaluated as the kinetics of dietary fiber in the large intestine of the mammal. Therefore, it is expected to be used to predict the intestinal fermentability of dietary fiber and the functionality of prebiotic foods.

The present invention will be described in detail below with reference to examples. However, the present invention is not limited to these.

(Example 1: Dietary fiber addition culture)
Fecal samples were collected from 8 healthy volunteers. The experimental participants had not received any antibiotics for more than 2 months. After the fecal samples were collected, they were stored in anaerobic culture swabs (212550 BD BBL Culture Swab; manufactured by Becton, Tickinson and Company) and sent to the laboratory.

A single-batch type culture apparatus simulating human large intestine was prepared using a multichannel incubator (Bio Jr. 8; manufactured by ABLE Co., Ltd.). This in vitro model apparatus consists of eight parallel independent vessels, each containing 100 mL of autoclaved (115° C. for 15 minutes) Gifu University-prescribed anaerobic medium (GAM medium [Code 05422]; Nissui Pharmaceutical Co., Ltd.). company) and adjusted the pH to 0.1 M phosphate buffer (68.5:31.5 (molar ratio) mixture of pH _6.5 , 0.1 M _NaH2PO4 and 0.1 _{M Na2HPO4 )} . ) was adjusted to 6.5 by addition. Aeration (15 mL) in nitrogen and carbon dioxide mixed gas (N ₂ : CO ₂ =80:20 (volume ratio)) filtered through a 0.2 μm PTFE membrane (manufactured by Pall Corporation) at 37° C. for 1 hour before culturing /min) to establish anaerobic conditions in the culture vessel. For the preparation of inoculum, fecal samples were added to 0.1 M phosphate buffer (pH 6.5, _0.1 M NaH PO _and suspended in ₂ mL of a 68.5:31.5 (molar ratio) mixture of 0.1M _Na2HPO4 . Anaerobic culture was initiated by inoculating 100 μL of the fecal suspension into each medium-containing container. During cultivation, anaerobic conditions were maintained by aerating the medium with a filter-sterilized gas mixture.

After 18 hours of culture, dietary fiber was added at an addition concentration of 1.0 (w/v)% (1.0 g/100 mL), and cultured for an additional 24 hours (42 hours in total). At 0 hour, 3 hours, 6 hours, 9 hours, 12 hours and 24 hours after the addition of dietary fiber, a portion of the culture solution was collected from the projecting portion on the side of the container. The harvested culture fluid was analyzed. Fecal samples and cultures were stored at -20°C until analysis.

The following dietary fibers were used: polydextrose (PDS: "Lytes II", manufactured by Dupont Nutrient & Biosciences), resistant glucan (RGN: "Fit Fiber #80", manufactured by Nihon Shokuhin Kako Co., Ltd.), indigestible Dextrin (DEX-1: "Nutriose FM06", manufactured by Rocket), indigestible dextrin (DEX-2: "Promita 85", manufactured by Rocket), indigestible dextrin (DEX-2: "Fibersol 2" , manufactured by Matsutani Chemical Industry Co., Ltd.), isomaltodextrin (IMD: “Fiberixa”, manufactured by Hayashibara Co., Ltd.).

Table 1 shows the glycosidic bond percentage (%) and molecular weight of each dietary fiber. The ratio (%) of various glycosidic bonds is obtained by determining the bonding carbon positions of the constituent sugars of the sugar chain by methylation analysis and calculating the number of 1→4, 1→6, 1→2, and 1→3 glycosidic bonds. Asked and calculated. Methylation analysis was according to the method of Cuine and Kerek (Carbohydr Res. Doi: 10.1016/0008-6215(84)85242-8). Gas chromatograph-mass spectrometer (GC (7890A; manufactured by Agilent Technologies)-MS (Jms-Q1000GC; manufactured by JEOL Ltd.)) was performed under the following conditions: Column: BD-225 capillary column (30 mL × 0.25 mm ID×0.15 μm film; manufactured by Agilent Technologies); column temperature: heated at 170° C. for 1 minute, then increased to 210° C. at 3° C./min and maintained at 210° C. for 20 minutes; temperature: 210° C.; eluent: helium). Molecular weights were determined by high performance liquid chromatography (HPLC) based on the average value of one sugar molecule. HPLC measurements were performed under the following conditions: Column: TSKgel G6000PWXL, G3000PWXL, G2500PWXL (300 mm X 7.8 mm ID) (manufactured by Tosoh Corporation); column temperature: 80°C; eluent: distilled water; flow rate: 0.5 mL. /min; detection: differential refractive index; injection volume: 100 μL. Samples were prepared in distilled water at a final concentration of 1% (w/w). Standard P-82, maltose and glucose were used as standard samples.

Hereinafter, various dietary fibers are represented by "PDX", "RGN", "DEX-1", "DEX-2", "DEX-3" and "IMD".

(Example 2: Organic acid analysis)
The culture fluid collected at 0 and 24 hours after the addition of dietary fiber in Example 1 was used for measuring the concentration of organic acids (short-chain fatty acids: acetic acid, propionic acid and butyric acid). The filtrate obtained by filtering the culture solution was subjected to high performance liquid chromatography (HPLC) equipped with an Aminex HPX-87H column (manufactured by Bio-Rad Laboratories) and a RID-10A differential refractive index detector (manufactured by Shimadzu Corporation). (manufactured by Shimadzu Corporation) to measure the amounts of acetic acid, propionic acid, and butyric acid. The HPLC was operated at 65° C. with 5 mM H ₂ SO ₄ as mobile phase at a flow rate of 0.6 mL/min.

FIG. 1 is a graph showing the production amounts of (a) acetic acid, (b) propionic acid and (c) butyric acid when cultured for 24 hours after adding various dietary fibers (42 hours after culture). The amount of production was expressed as a relative ratio of the amount of each organic acid in the case of adding dietary fiber to that in the control without dietary fiber. In the control, (a) acetic acid, (b) propionic acid and (c) butyric acid were produced at 90.3±15.0 mM, 28.9±8.95 mM and 13.5±12.0 mM, respectively (mean ±SD, n=8). * in the graph indicates a significant difference from the control value (n=8) by Dunnett's test (*p<0.05 and **p<0.01). The data are presented as medians and interquartile ranges (25% tile to 75% tile).

(Example 3: Decomposition rate and decomposition rate of dietary fiber)
The culture fluid collected at 0 and 24 hours after the addition of dietary fiber in Example 1 was used for determining the degradation rate and degradation rate of dietary fiber. In order to determine the degradation rate and degradation rate, the filtrate obtained by the enzymatic-gravimetric method was subjected to HPLC to determine the amount of dietary fiber.

Culture medium (5 mL) was mixed with phosphate buffer (15 mL, 0.08 M, final pH 6.0). The total amount of dietary fiber was measured according to the procedure described in the Total Dietary Fiber Assay Kit (manufactured by Sigma-Aldrich, TDF-100A). Gelation with thermostable α-amylase followed by digestion with protease and amyloglucosidase removed proteins and starch in the sample. Amberlite IRA67 and 200CT(H)HG were used to remove residual impurities. The indigestible portion of the dietary fiber was quantified by HPLC (manufactured by Shimadzu Corporation) equipped with two TSKgel G2500PWxl columns (manufactured by Tosoh Corporation).

FIG. 2 is a graph showing (a) decomposition rate and (b) decomposition rate of various dietary fibers. The degradation rate (g/L/h: concentration per hour) was calculated using linearly degrading values. Data are presented as median and interquartile range (25% tile to 75% tile). The decomposition rate (%) was calculated as the ratio of the amount of dietary fiber 24 hours after addition of dietary fiber (after 42 hours of culture) to the amount of dietary fiber initially added (after 18 hours of culture). * in the graph indicates a significant difference from the control value (n=8) by Wilcoxon signed rank sum test (*p<0.05 and **p<0.01).

(Example 4: Correlation analysis of organic acid concentration and dietary fiber decomposition)
The decomposition rate and rate of dietary fiber were plotted against the concentration of various organic acids to investigate the correlation. These results are shown in Figures 3-5.

FIG. 3 is a graph showing the relationship between (A) decomposition rate and (B) decomposition rate of dietary fiber with respect to acetic acid concentration. FIG. 4 is a graph showing the relationship between (A) degradation rate and (B) degradation rate of dietary fiber with respect to propionic acid concentration. FIG. 5 is a graph showing the relationship between (A) decomposition rate and (B) decomposition rate of dietary fiber with respect to butyric acid concentration. Acetic acid and propionic acid showed correlations with the degradation rate and rate of dietary fiber, respectively. Faster decomposition rates and higher decomposition rates increased the production of acetic acid and propionic acid. As the decomposition rate of dietary fiber increased, the organic acid productivity also increased for acetic acid and propionic acid. Butyric acid did not show any correlation between the rate and rate of degradation of dietary fiber.

(Example 5: Correlation analysis between organic acid concentration and total number of 1→2 glycosidic bonds and number of 1→3 glycosidic bonds)
For various dietary fibers, the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber was plotted against the organic acid concentration to investigate the correlation. "Total glycosidic bond number" is the sum of 1→2 glycosidic bond number, 1→3 glycosidic bond number, 1→4 glycosidic bond number and 1→6 glycosidic bond number.

FIG. 6 shows the relationship between (a) acetic acid concentration, (b) propionic acid concentration and (c) butyric acid concentration with respect to the ratio of the total number of 1 → 2 glycosidic bonds and 1 → 3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. is a graph showing Acetic acid and propionic acid showed a correlation to the ratio of the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds to the total number of glycosidic bonds. In particular, propionic acid showed a high correlation. The larger the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds, the lower the amounts of acetic acid and propionic acid produced. No correlation was observed with butyric acid.

(Example 6: Correlation analysis between the degradation rate and degradation rate of dietary fiber and the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds)
For various dietary fibers, the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber was plotted against the organic acid concentration to investigate the correlation.

FIG. 7 is a graph showing the relationship between (a) decomposition rate and (b) decomposition rate of dietary fiber with respect to the ratio of the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds to the total number of glycosidic bonds of dietary fiber. be. The degradation rate and degradation rate showed a correlation with the ratio of the total number of 1→2 glycosidic bonds and 1→3 glycosidic bonds to the total number of glycosidic bonds. The greater the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds, the lower the rate of degradation and the rate of degradation. and the degradation rate increased, suggesting that it is susceptible to degradation in the human large intestine. No such correlation was observed for butyric acid.

(Reference Example 1: Analysis of bacteria in feces)
Genomic DNA of bacteria in the bacterial flora was extracted from the culture medium collected 24 hours after the addition of dietary fiber in Example 1. The V3-V4 region of the bacterial 16S rRNA gene was amplified from the extracted genomic DNA and subjected to sequence analysis using a next-generation sequencer to perform bacterial diversity analysis and bacterial composition analysis. The procedure is shown below.

Using the primer pair S-D-Bact-0341-b-S-17 (SEQ ID NO: 1) and S-D-Bact-0785-a-A-21 (SEQ ID NO: 2), the extracted genomic DNA was used as a template to amplify the bacterial 16S rRNA gene. did. An Illumina adapter overhang nucleotide sequence (manufactured by Illumina Inc.) was added to the gene-specific sequence. PCR cycling reactions were performed according to the manufacturer's instructions. Verified amplicons were purified using AMPure XP DNA purification beads (Beckman Coulter, Inc.) and eluted in 25 μl of 10 mM Tris (pH 8.5). Amplicons were quantified on an Agilent Bioanalyzer 2100 DNA 1000 chip (manufactured by Agilent Technologies) and pooled at equimolar concentrations. The 16S rRNA gene product (along with an internal control (PhiX Control V3; Illumina Inc.)) was subjected to paired-end sequencing using a MiSeq sequencer (Illumina Inc.) with a 600 cycle MiSeq reagent kit (Illumina Inc.). . PhiX sequences were excised and paired-end reads with Q scores >20 were spliced using the software package QIIME version 1.9.1. Chimeric sequences were identified and eliminated from the library. Paired-end reads were phylogenetically classified by the GreenGenes phylogenetic database using the Ribosomal Database Project (RDP) Classifier. OTUs (Operational Taxonomic Units) that reached 97% similarity were used for diversity comparison of Shannon-Wiener and Simpson indices.

Real-time PCR was performed using TP700 Thermal Cycler Dice Real Time System Lite (manufactured by Takara Bio Inc.). Amplification with a primer set targeting whole gut bacteria was performed as described in Takagi, R et al., PLoS One 11, e0160533 (2016).

The Shannon-Wiener Diversity Index and Simpson Index were calculated using the QIIME software package. The non-parametric Kruskal-Wallis test was used to determine significant differences for OTU numbers, Shannon-Wiener diversity index and Simpson index. P-values less than 0.05 were considered significant.

FIG. 8 shows the results of (a) the number of sequences (number of reads) obtained by the next-generation sequencer, (b) the number of OTUs, (c) the Shannon-Wiener diversity index, and (d) the Simpson index. The stool sample at the start of culture is represented as "FEC", and the culture solution without dietary fiber added is represented as "CUL". Data are presented as median and interquartile range (25% tile to 75% tile). There was no significant difference in the number of OTUs, the Shannon-Wiener diversity index and the Simpson index between the cultures with no dietary fiber added and those with dietary fiber added.

FIG. 9 is a graph showing the abundance ratio of various enterobacteriaceous phyla to total enterobacteria in stool samples and culture solutions. From sequence analysis of the 16S rRNA gene products, the bacteria detected include Unclassified Bacteria, Verrucomicrobia, Proteobacteria, Fusobacteria, Firmicutes, Bacteroidetes. Bacteroidetes) and Actinobacteria. In FIG. 9, “HS1” to “HS8” indicate feces from eight healthy subjects, and within each “HS”, FEC, CUL, PDX, RGN, DEX-1, DEX-2, DEX-3 and IMD The results are shown in order of There was no significant difference in the proportions of various enterobacteriaceae between the culture solution with no dietary fiber added and the culture solution with dietary fiber added.

Therefore, there was no significant difference in the intestinal microflora formed by fecal culture between the case of adding dietary fiber and the case of not adding dietary fiber.

(Example 7: Analysis of bacteria in feces)
Cultivation was carried out in the same manner as in Example 1, except that only IMD was used as dietary fiber and the culture time before addition of dietary fiber was either 12 hours, 18 hours or 24 hours. IMD-added and non-IMD-added cultures were collected at the end of the dietary fiber IMD-added culture to determine the rate of IMD degradation, with the following results:
Degradation rate when the culture time before adding dietary fiber was 12 hours: 0.48 g / L / h
Degradation rate when culture time before dietary fiber addition was 18 hours: 0.86 g / L / h
Degradation rate when culture time before dietary fiber addition was 24 hours: 0.91 g / L / h

When the same dietary fiber was cultured for a different period of time before adding dietary fiber, the degradation rate of dietary fiber was low in 12-hour culture, intestinal bacteria did not grow sufficiently in the culture medium, and the decomposition performance was low. I wanted to think In contrast, there was almost no difference in the rate of decomposition of dietary fiber between the 18-hour culture and the 24-hour culture. From this result, it was considered that 18-hour culture was optimal for examining the ability to decompose dietary fiber.

INDUSTRIAL APPLICABILITY The present invention is useful for the production of food, beverages and pharmaceuticals.

Claims (22)

A method for screening water-soluble dietary fiber having an organic acid-producing ability in mammalian intestines, comprising:
A step of determining the ratio (A) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber, and 1→2 glycoside bonds to the total number of glycosidic bonds of the standard dietary fiber group. Using a relative curve showing the correlation between the total ratio (B) of the number of bonds and the number of 1→3 glycosidic bonds and the production of organic acids in the mammalian intestine of the standard dietary fiber group, the ratio ( A step of estimating the ability of the water-soluble dietary fiber to produce organic acids in the intestine of the mammal from A);
including
The method wherein the organic acid is acetic acid and/or propionic acid. A computer-implemented program for screening a water-soluble dietary fiber capable of producing an organic acid in mammalian intestines, the method comprising:
Determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of water-soluble dietary fiber, and the step of determining the ratio (A) of the total number of 1→2 glycosidic bonds to the total number of glycosidic bonds of the standard dietary fiber group and 1→3 glycosidic bond number (B) and the production of organic acids in the mammalian intestine of the standard dietary fiber group, the ratio (A ), the step of estimating the ability of the water-soluble dietary fiber to produce organic acids in the intestine of the mammal;
including
A program wherein the organic acid is acetic acid and/or propionic acid. A method for predicting the degradation performance of water-soluble dietary fiber in mammalian intestines, comprising:
A step of determining the ratio (A) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber, and 1→2 glycoside bonds to the total number of glycosidic bonds of the standard dietary fiber group. using a relative curve showing the correlation between the total ratio (B) of the number of bonds and the number of 1→3 glycosidic bonds and the degradation rate and/or degradation rate in the mammalian intestine of the standard dietary fiber group, estimating the intestinal degradation rate and/or degradation rate of the water-soluble dietary fiber from the ratio (A);
A method, including A computer-implemented program for predicting the degradation performance of water-soluble dietary fiber in a mammalian intestine, the method comprising:
A step of determining the ratio (A) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber, and 1→2 glycoside bonds to the total number of glycosidic bonds of the standard dietary fiber group. using a relative curve showing the correlation between the total ratio (B) of the number of bonds and the number of 1→3 glycosidic bonds and the degradation rate and/or degradation rate in the mammalian intestine of the standard dietary fiber group, estimating the intestinal degradation rate and/or degradation rate of the water-soluble dietary fiber from the ratio (A);
program, including A method for evaluating whether a substance containing water-soluble dietary fiber functions as a prebiotic in a mammal, comprising:
Determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group A step of estimating the organic acid-producing ability of the substance in the mammal from the ratio (A) using a relative curve showing the correlation, and whether it functions as a prebiotic based on the organic acid-producing ability including the step of evaluating
The method wherein the organic acid is acetic acid and/or propionic acid. 6. The method of claim 5, wherein said prebiotic is a food or pharmaceutical. A computer-implemented program for evaluating whether a substance containing soluble dietary fiber functions as a prebiotic in a mammal, the method comprising:
Determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group A step of estimating the organic acid-producing ability of the substance in the mammal from the ratio (A) using a relative curve showing the correlation, and whether it functions as a prebiotic based on the organic acid-producing ability including the step of evaluating
A program wherein the organic acid is acetic acid and/or propionic acid. A method for evaluating or predicting the intestinal dynamics and/or intestinal fermentability of a substance containing water-soluble dietary fiber in a mammal,
Determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group a step of estimating the ability of the substance to produce an organic acid in the mammal from the ratio (A) using a relative curve showing the correlation; including the step of evaluating or predicting endofermentability;
The method wherein the organic acid is acetic acid and/or propionic acid. A computer-implemented program for evaluating or predicting the intestinal dynamics and/or intestinal fermentability of a substance containing water-soluble dietary fiber in mammals, the method comprising:
Determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group a step of estimating the ability of the substance to produce an organic acid in the mammal from the ratio (A) using a relative curve showing the correlation; including the step of evaluating or predicting endofermentability;
A program wherein the organic acid is acetic acid and/or propionic acid. A culture device for evaluating or predicting the degradation performance of water-soluble dietary fiber in mammalian intestines and/or the organic acid production performance,
A means for anaerobic culture using feces to construct an intestinal flora;
a means for determining the total number of glycosidic bonds ;
means for determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber;
The ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group, and (i) the amount of organic acids in the intestine of the mammal in the standard dietary fiber group using a relative curve showing the correlation between production and/or (ii) the degradation rate and/or degradation rate in the mammalian intestine of the standard dietary fiber group, From said ratio (A), (i) a means of inferring the ability of said water-soluble dietary fiber to produce organic acids in said mammalian intestine, and/or (ii) the rate of degradation of said water-soluble dietary fiber in said intestine. and/or a means of estimating the degradation rate
a culture device. A combination for evaluating or predicting the degradation performance of water-soluble dietary fiber in mammalian intestines and / or organic acid production performance,
A culture device for anaerobic culture using feces to build an intestinal flora;
a means for determining the total number of glycosidic bonds ;
means for determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber;
The ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group, and (i) the amount of organic acids in the intestine of the mammal in the standard dietary fiber group using a relative curve showing the correlation between production and/or (ii) the degradation rate and/or degradation rate in the mammalian intestine of the standard dietary fiber group, From said ratio (A), (i) a means of inferring the ability of said water-soluble dietary fiber to produce organic acids in said mammalian intestine, and/or (ii) the rate of degradation of said water-soluble dietary fiber in said intestine. and/or a means of estimating the degradation rate
combination of. A system for evaluating whether a substance containing soluble dietary fiber functions as a prebiotic in a mammal, comprising:
A means for anaerobic culture using feces to construct an intestinal flora;
a means for determining the total number of glycosidic bonds ;
means for determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group means for estimating the ability of the substance to produce an organic acid in the mammal from the ratio (A) using a relative curve showing the correlation;
and a means for evaluating whether it functions as a prebiotic based on the ability to produce the organic acid
system, including A system for evaluating or predicting the intestinal dynamics and/or intestinal fermentability of a substance containing water-soluble dietary fiber in a mammal,
A means for anaerobic culture using feces to construct an intestinal flora;
a means for determining the total number of glycosidic bonds ;
means for determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the substance containing the water-soluble dietary fiber;
Between the ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the mammal in the standard dietary fiber group means for estimating the ability of the substance to produce an organic acid in the mammal from the ratio (A) using a relative curve showing the correlation;
and means for evaluating or predicting intestinal dynamics and/or intestinal fermentability based on the ability to produce the organic acid
system, including A system for evaluating or predicting the degradation performance and / or organic acid production performance of water-soluble dietary fiber in mammalian intestines,
A means for anaerobic culture using feces to construct an intestinal flora;
a means for determining the total number of glycosidic bonds ;
means for determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber;
(B) the ratio of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group, and (i) the amount of organic acids in the intestine of the mammal in the standard dietary fiber group using a relative curve showing the correlation between production and/or (ii) the degradation rate and/or degradation rate in the mammalian intestine of the standard dietary fiber group, From said ratio (A), (i) a means of inferring the ability of said water-soluble dietary fiber to produce organic acids in said mammalian intestine, and/or (ii) the degradation rate of said water-soluble dietary fiber in said intestine. and/or a means of estimating the degradation rate
system, including A method for evaluating the degradation performance of water-soluble dietary fiber in mammalian intestines, comprising:
(a) a step of anaerobic culture using feces from a healthy individual to construct an intestinal flora;
(b) adding the water-soluble dietary fiber to the culture obtained in step (a) and further anaerobic culturing; and (c) the water in the culture obtained in step (b) measuring the degradation rate and/or degradation rate of soluble dietary fiber , wherein the water-soluble dietary fiber is
determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber; and
The ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the degradation rate and/or degradation rate of the standard dietary fiber group in the intestine of the mammal estimating the intestinal degradation rate and/or degradation rate of the water-soluble dietary fiber from the ratio (A) using a relative curve showing the correlation between
wherein the degradation performance in the intestine of said mammal is predicted by a method comprising : 16. The method of claim 15, wherein the anaerobic culture of step (a) is performed for 16 hours to 24 hours. 17. The method of claim 15 or 16, wherein said mammal is a human. The construction of the intestinal flora is carried out by anaerobically culturing the feces for at least 12 hours or more in an anaerobic environment created by aerating an anaerobic gas containing nitrogen and carbon dioxide into the medium. ,
The method according to any one of claims 15 to 17, wherein said anaerobic environment is 36°C to 38°C and pH 6.2 to 6.7. A method for evaluating the organic acid-producing ability of water-soluble dietary fiber in mammalian intestines, comprising:
(i) a step of anaerobic culture using feces from a healthy individual to construct an intestinal flora;
(ii) adding the water-soluble dietary fiber to the culture obtained in step (i) and further anaerobic culturing; and (iii) the organic matter in the culture obtained in step (ii) measuring the amount of acid, the water-soluble dietary fiber comprising:
determining the ratio (A) of the sum of the number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds of the water-soluble dietary fiber; and
The ratio (B) of the total number of 1→2 glycosidic bonds and the number of 1→3 glycosidic bonds to the total number of glycosidic bonds in the standard dietary fiber group and the production amount of organic acids in the intestine of the mammal in the standard dietary fiber group estimating the ability of the water-soluble dietary fiber to produce an organic acid in the intestine of the mammal from the ratio (A) using a relative curve showing the correlation between
was screened by a method comprising
The method wherein the organic acid is acetic acid and/or propionic acid. 20. The method of claim 19, wherein the anaerobic culture of step (i) is performed for 16 hours to 24 hours. 21. The method of claim 19 or 20, wherein said mammal is human. The construction of the intestinal flora is carried out by anaerobically culturing the feces for at least 12 hours or more in an anaerobic environment created by aerating an anaerobic gas containing nitrogen and carbon dioxide into the medium. ,
The method according to any one of claims 19-21, wherein the anaerobic environment is 36°C-38°C and pH 6.2-6.7.

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