UA65532C2 - Absorbable dietary composition for improving biological balance of intestinal tract flora - Google Patents

Abstract

An absorbable dietary composition for improving the biological balance of intestinal tract flora in humans and mammals is disclosed. Said composition consists of a dehydrated preparation containing, propionic bacteria and bifid bacteria in an amount of at least 109 cells/g, preferably 1010-1012 cells/g.

Description

The present invention relates to an easily digestible dietary composition for improving the biological balance of the intestinal flora of humans and other mammals.

It is known that the human digestive tract, the capacity of which is approximately 1 to 1.5 kg of food during its digestion, is inhabited by a large population of microorganisms, which in the colon is estimated to be about 1079 cells per gram.

This population consists of different groups of bacteria, some of which are harmless or even beneficial, while others, especially enteric bacteria and putrefactive bacteria, lead to the production of toxic substances and have a detrimental effect on health.

Since the beginning of the twentieth century, many significant studies have been conducted to demonstrate the fact that the presence of a large population of lactic acid bacteria very dramatically limits the development of putrefactive bacteria and, therefore, the production of toxic substances.

It is therefore quite natural that the idea of introducing a large population of these bacteria into the body either with certain foodstuffs or by direct contamination with these microbes through the mouth has been proposed; among the acceptable types of lactic acid flora, the following were recommended for use:

And asiorasiyshe asidorniyshee or even bigeriosossivye Taesiyt (they were mentioned only as an example, but this list is not exhaustive).

However, the introduction of these bacteria did not lead to the expected results, because, on the one hand, the population of microbes in the digestive tract is a mass, the balance of which is difficult to change radically and for a long time, and on the other hand, it was found that the use of lactic acid bacteria does not have much effect on putrefactive flora, which causes known diseases and disorders.

Later - in the seventies - the idea of introducing bifidobacteria into the human diet, which do not belong to the lactic acid genus and which proved to be beneficial for health, appeared. Leavens of this type can be used either with milk puddings, or with other dairy products partially fermented by bifidoflora, or by direct ingestion of live cells in an acceptable form.

However, in practice, such introduction of bacteria did not give the desired results, and in particular because it is very difficult to successfully transplant exogenous bifidobacteria into the human intestine for a long period, especially if the balance of its intestinal flora is not ideal.

Based on these discovered general facts, the main idea of the invention appeared - to find a means that would allow to increase the population of bifidobacteria.

In the process of this work, the authors came across the work of Professor Henri Berran from Lille, who managed to discover that propionic acid is a specifically selective and elective element of bifidoflora.

It is known that this organic acid is one of the products that arise mostly as a result of the fermentation of exclusively lactate by bacteria of a certain type - propionic bacteria, which, although not belonging to lactic acid bacteria, have been present in human food products for several centuries; Emmental cheese owes its holes to these bacteria: they are formed because, at the end of the cooking process, this cheese contains about 109 propionic bacteria cells per gram.

It should be noted that, in addition to propionic acid, the fermentation of propionic bacteria is also caused by acetic acid and carbonic acid.

In parallel with this work, thanks to the publication of Tita Capeko, Nigopagy Moi, Mediti maga apa bZaspiki

Medigo, "Sgomp egitshayog og Viidobasiegia Rhodisei Bu Rgoriopirasiepiyit netsdepgeisnii apa bZemegai Ipieziipa!

Vasiepa", 1994, ). Oaigu Zsi. 77:393-404, it was found that, on the one hand, propionic bacteria, especially bacteria of the type Rhoriopyrasiyut igetsdepgeisnii, are capable of producing the growth factor of bifidobacteria, and on the other hand - short-chain fatty acids , especially propionic acid, have a clearly demonstrated inhibitory effect on the development of many harmful intestinal bacteria, but stimulate the development of bifidobacteria.

In the patent UR-A-07 227207, it was already proposed to add propionic bacteria to food products in order to increase the viability of bifidobacteria, especially to fermented milk containing these bacteria.

In addition, it should be noted that from the patent Sh5-A-4 8063689, a diet pill created on the basis of dietary fibers, in particular apple fibers, which may contain bifidobacteria, as well as, in a smaller amount, propion bacteria, which act as an antifungal agent, is already known .

Based on these preliminary discoveries, the idea of this invention arose, namely, the idea of finding ways to directly use a large amount of combined bifidoenzymes and propionic enzymes within the limits of a measurable dose that can be easily swallowed.

It was clear that this method of consumption has advantages in many aspects, in particular it allows to significantly reduce the usual attacks of putrefactive bacteria, even in the case of a possible negative effect, taking into account the large amount of carbon dioxide released by propionic fermentation, as occurs when formed by these enzymes cavities in cheese.

Therefore, the task of this invention is to provide an easily digestible dietary composition for improving the biological balance of the flora of the digestive tract of humans and other mammals, which is characterized by the fact that it consists of an anhydrous preparation containing propion bacteria and bifidobacteria in the amount of at least 109 cells per gram, in the optimal version - in the range from 1019 to 1012 cells per gram, that is, a high concentration of bacteria.

In general, these bacteria must be alive, but they can be killed, if measures are taken to preserve the content of enzymes in them.

In the optimal version, these bacteria should be in the following ratio: from 80 to 9995 propionic bacteria and from 20 to 195 bifidobacteria.

Thus, the composition according to the present invention contains, on average, ten times more propionic bacteria than bifidobacteria.

The bifidobacteria used belong, as a rule, to the genus Voydobasiepyt and, in the optimal version, are selected from the strains of V. rbibydyt, V. Iopdit, V. Adoiezsepiiv, V. bheme, V. ipgapii5 and V. rzendoiopdit.

Propionic bacteria also, as a rule, belong to the genera Rhoriopyrasieist iecydepgeisp».

Rgoriopirasiegt 5Peptapii, Rgoriopirasiepst (Shoepii, Rgoriopirasijeitst |epzepii or Rgoriopirasieiet asiairgoriopsii.

These bacteria can be used as a pure strain or, preferably, as a mixture of strains; as for propionic bacteria, the simultaneous combination of highly autolytic and not very autolytic strains is preferred.

The invention also relates to the use of an anhydrous preparation containing propionic bacteria in the amount of at least 109 cells per gram, in the optimal version - in the range from 1019 to 1012 cells per gram, for obtaining an easily digestible dietary composition for improving the biological balance of the flora of the digestive tract of humans and other mammals , in particular to increase the population of bifidobacteria.

It should be noted that this population of bifidobacteria can originate either from endogenous bacteria or from exogenous bacteria added to the preparation.

This composition optimally exists in the form of separate fractions in the amount of approximately 100 mg to 1 g, better - from 200 to 500 mg, and contains the necessary dose of bacteria for daily use.

These fractions are intended either for direct oral administration, or they are first diluted in a liquid; they can have any form convenient for such reception: tablets, sachets of granulated or non-granulated powder, etc.

It was confirmed that such concentrated anhydrous preparations of propionic bacteria, stored for two years at 14"C, show such a decrease in concentration that does not exceed the natural logarithm.

Experience shows that the best form is capsules, both gastro-resistant and other.

According to another criterion of the invention, each separate fraction contains at least 109 bacteria, in the optimal version - in the range from 1019 to 1012 bacteria.

It should be noted that with the minimum concentration of bacteria, which is 1079, it is not always possible to achieve the desired effect; this is apparently because the bacteria are then destroyed by stomach acid or bile salts before they reach the colon.

Thus, according to the present invention, each person is suggested to consume daily with food a dose of propionic bacteria, which is approximately one thousandth of the entire flora of the digestive tract (about 10!7 units); this number is significantly higher than previously suggested for other types of bacteria.

It was confirmed that such a dose according to the present invention does not have a harmful effect and, in addition to the above-mentioned advantages, provides the following: - a noticeable decrease in the production of gases with an unpleasant smell, characteristic of putrefactive bacteria; - change in stool color and smell.

These findings certainly also highlight the "symbiotic" effect of propion bacteria and bifidobacteria, where propion bacteria stimulate the development of bifidobacteria.

This result was confirmed by a study conducted to find out whether the strains actually

Rgoriopirasiegts are able to facilitate the adaptation of the Vijdobasiegisht riidit strain in the digestive tract of mice or to increase the level of their activity in different segments of the digestive tract.

In the initial phase, this study showed that it is impossible to achieve long-term adaptation

Rgoriopyrasiegut in the digestive tract of mice, even mice that are sterile, that is, in which the flora of the digestive tract is absent.

On the contrary, it was demonstrated that the addition of a culture of R. goriopyrasipyst to the drinking water of these mice allows to stimulate the formation of the strain Viyidobasiegisht riiiaiSht colonies in the digestive tract.

Thus, this scientific study proves the existence of a direct synergy between the action of propionic bacteria and the action of bifidobacteria.

In addition, these results were confirmed by a series of experiments conducted in both ip mimo in healthy humans and ip myo.

Try ip mimo

The purpose of these experiments was to study in the clinical conditions of the University Hospital in Caen (France) the viability and colonization of strains of propionic bacteria in the digestive tract environment, as well as their effect on bifidoflora in a group of nineteen healthy male volunteers.

The experiments included daily oral intake for two weeks by each of the experimental capsules containing 5 x 1019 propionic bacteria selected at IMVA (National Agricultural Research Institute) from the Bank of strains used in the cheese industry and therefore completely harmless to humans.

Assigned propionic bacteria were divided into two strains of approximately 5095 each, one of which was not very autolytic, while the other became apolitic in the stomach and early small intestine when exposed to bile salts.

Faecal flora from stool samples before administration of propionic bacteria were examined to determine the baseline flora, and then during and after administration to assess how long exogenous bacteria were able to survive in the digestive tract of subjects.

This study included counting propionate bacteria and bifidobacteria consumed.

On the basis of these experiments, it was confirmed that already from the first sample during the intake of propionic bacteria by the subjects, the population of propionic bacteria in the stool increased significantly, but this effect was temporary and lasted for a short time after the cessation of taking the bacteria.

These results indicate that a fraction of propionic bacteria strains are able to survive in the human digestive tract, but they are not able to colonize there for a long time.

To assess the effect of propionic bacteria on the population of bifidobacteria, attention was also paid to the samples taken (a) before the administration of propionic bacteria; (b) during their reception; and after admission (two samples (c and d), taken with an interval of one week).

It should be noted that the indicator of the sample (b) is the arithmetic mean of the indicators of two samples taken with an interval of one week.

The counting of bifidobacteria made it possible to calculate the average number (in natural logarithms) of populations during these four periods.

The corresponding mean values are: (a) 8.40; (b) 8.90; (c) 9.43 and (d) 8.89. Evolution over time was studied using a non-parametric statistical test for compared populations, the Wilcox pairwise relative rank test. It shows that during the administration of propionic bacteria (sample b), the population of bifidobacteria increases significantly compared to the baseline (sample a) (p-0.0096) and remains at a high level at the first check (sample c) after stopping their administration ( p-0.031). There is no difference between the number of bifidobacteria during the reception of propionic bacteria (sample b) and at the above-mentioned first check after its termination (1 week) (sample c) (p-0.19); the population of bifidobacteria decreases at the second check after stopping the intake (sample d) (p-0.049) compared to the first check (sample c) and returns to the level that was determined before taking propionic bacteria (sample a) (p-0.25), in the period between the second inspection (sample d) and the inspection that precedes it (sample c).

These results show that taking propionic bacteria in high doses promotes the development of bifidoflora; the effect was rapid, it was observed from the very first week and continued for at least a week after stopping the propionic bacteria.

Experiments ip could

The task of these experiments was: - to study the resistance of propion bacteria to digestive "stresses", that is, gastric acidity and bile salts, - to study the effect that propion bacteria have on the development of bifidobacteria, - to identify, if possible, the agent responsible for this effect.

Resistance of propionic bacteria to digestive "stress"

Between the time of ingestion and the time of arrival in the colon, where they come into contact with bacteria in the digestive tract, especially bifidobacteria, propionic bacteria undergo numerous "stresses" that prevent their survival, the main ones being gastric acidity and contact with bile salts in the small intestine.

To evaluate the resistance of propionic bacteria to these "stresses", two strains belonging to the genus Rgoriopirasiegsht pgecaepgeisni were selected, namely: strain 5410, which is not very autolytic, and strain

Y 5 2501, which is highly autolytic. The pH of the human stomach, after the appearance of food in it, is from 2 to 3, and the average time required for emptying the stomach is approximately 90 minutes.

To evaluate the resistance of the above strains to gastric acidity, these strains were tested at different pH levels (2, C and 4) for 90 minutes at 3720.

As a result of the conducted observations, it was established that:

At pH 4, the viability of the two strains does not change at all;

At pH 3, strain I 5 410 remains intact, but strain I 5 2501 shows a slight decrease in viability; At pH 2, both strains almost completely die after 90 minutes of incubation.

The resistance of two strains to bile salts at 37"С was measured with concentrations of bile acids of 1.2 and 5 g/l, which, as a rule, is the concentration at the entrance to the small intestine.

Thanks to these experiments, it was found that at the weakest concentration (1 g/l), strain 15 410 completely retains its resistance even after 5 and a half hours of incubation. In contrast, strain I 5 2501 shows less resistance.

In the presence of 2 to 5 g/l of bile acids, the viability of the two strains drops sharply, and the strain

Y 5 2501 is more sensitive than strain Y 5 410.

So, these experiments showed that the two investigated strains clearly demonstrate great resistance to moderate digestive "stresses". However, most bacteria are unable to withstand conditions of extreme "stress", and the more sensitive strain is highly autolytic (I 5 2501).

These results indicate the need for oral administration of a large number of bacteria to achieve a noticeable effect on the biological balance of the flora of the human digestive tract to ensure the necessary survival of the strains at the entrance to the colon.

The influence of cultures of propionic bacteria on the development of bifidobacteria

To carry out these experiments, two strains of bifidobacteria, B. Iopdit and B. Biiidit, were sown in a medium containing in equal proportions: - culture broth suitable for the development of bifidobacteria, - a culture of propionic bacteria (strains I 5 410 or I 5 2501), either young (48 h) and, therefore, relatively unlysed, or 11 days old and, therefore, highly lysed; this was especially true of the autolytic strain

And 5 2501.

Accordingly, it was observed that the presence of young culture I 5410 and, to a lesser extent, 15 2501, as a result gives a higher level of the total population of B. biidite at the end of the exponential phase: the number of living bacteria is 3-4 times higher in the presence of I 5 410 and 2-3 times higher - in the presence of I 5 2501.

As for B. Iopdit, culture I 5 410 at the end of development doubles the population of B. Iopdit.

In cases with old cultures, a noticeable beneficial effect of the two strains was observed during the development of B. bydit during the exponential phase (the population doubled or tripled).

Therefore, both groups of propionic bacteria tested in this experiment affect the development of B.

log out their effect depends on the age of the culture: young cultures cause an increase in the viability of bifidobacteria at the end of development, as a result of which the population increases 2-4 times (a more noticeable effect when using I 5 410), while old cultures act only at the beginning of development, reducing the time , necessary for the formation of a population, but without changing the level of its final number.

Identification of the agent responsible for the "bifidogenic" effect - Effect of the upper layers of cultures of propionic bacteria - The role of propionic acid

Experiments similar to those mentioned above were carried out, using, first, only the upper layers of the cultures, which make up the extracellular fractions of the bacteria, and, second, a control medium with propionic acid.

Accordingly, it was observed that the upper layers of the culture of propionic bacteria compared to the control have a noticeable stimulating effect on the development of V. Viydyt (increasing the viability of the bacterium by 3-4 times after 9 hours in the culture); this effect is more pronounced at the end of development with strain I 5 2501.

These stimulatory effects were confirmed to be due in part to the effects of propionate. However, it appears that propionate alone is not responsible for these effects, especially in the case of strain I 5 2501.

Propionic acid has a stimulating effect on metabolic activity V. Iopdit. - Influence of the intracellular environment:

It was observed that the presence of the intracellular environment of two strains of propionic bacteria leads to a significant increase in optical density and dry cell mass of both strains of bifidobacteria.

However, these effects, which are not related to the activation of the development of V. Biydit or V. I opdit, are caused by the fact that their viability and their metabolic activity during development do not change. - Influence of insulated walls:

It was observed that the isolated walls of propionic bacteria, hydrolyzed in one way or another, have no effect on the development of B. Iopdite and B. rijdite.

So, it turns out that the effect of promoting the development of bifidobacteria by propionic bacteria is not related either to their intracellular content or to their walls. However, the compounds responsible for development are mostly found in the intracellular environment. One of these compounds is propionate.

Claims (8)

1. Dehydrated preparation, characterized by the fact that it contains propionic bacteria, in the amount of at least 109 cells per gram, preferably from about 1079 to 1012 cells per gram, suitable for obtaining an easily digestible composition for improving the biological balance of the flora of the intestinal tract of humans and other mammals by stimulating the development of bifidobacteria in the intestines. 2. The drug according to claim 1, which differs in that the composition contains bifidobacteria. 3. The preparation according to claim 2, which differs in that the composition contains from 80 to 99 95 propion bacteria and from 20 to 1 95 bifidobacteria. 4. The drug according to any one of claims 1-3, which differs in that the propionic bacteria belong to the genus Rgoriopyrasiegicht iecaepgeispii or Rgoriopirasiegisht 5pegtapia or Rgoriopirasieuyut (poveii or Rgogiopirasiegiit )epzepia or Rgoriopirasiegicht asidirgoriopisi. 5. The drug according to any one of claims 2-4, which is characterized by the fact that the bifidobacteria belong to the genus Viiidobasiegiot and are selected from the lines of V. Biyaut, V. Iopdit, V. adoiezsepii5, V. bheme, V. iptapiiz and V. Rzendoiopdit . 6. The drug according to any of claims 1-5, which differs in that the composition is presented in the form of individual fractions of approximately 100 mg to 1 g, preferably from 200 to 500 mg, and contains a dose of bacteria for daily intake. 7. The drug according to p.b., which differs in that the composition is presented in the form of capsules. 8. The preparation according to any one of claims 6 and 7, characterized in that each individual fraction contains at least 109, preferably from about 10192 to 1012 bacteria.