Combination of probiotics and butyric acid to improve health of intestinal epithelial cells, thereby improving health of pets and livestock

Abstract

Described herein is the use of a probiotic, in particular an Enterococcus faecium strain NCIMB 10415, named # imgabs0 #, as a probiotic active ingredient in combination with butyric acid or a salt thereof to improve the health of intestinal epithelial cells, thereby improving the health of pets and livestock.

Description

[0001] illustrate

[0002] The present invention relates to the field of animal nutrition, and more particularly to the use of probiotics in combination with butyric acid or its salts to improve the health of intestinal epithelial cells, thereby improving the health of pets and livestock.

[0003] Butyric acid and its salts, especially sodium butyrate, have been used in animal feed for decades. For optimal use, animals need to absorb butyric acid or its salts into the intestinal epithelial cells through this short-chain fatty acid (SCFA) absorption mechanism.

[0004] However, absorption of butyrate or its salts at the intestinal level may be impaired by inflammation, which results in or is characterized by damage to the intestinal tissue and epithelial barrier.

[0005] For example, we observed that the absorption of butyrate or its salts is impaired in mice that were fed a high-fat diet and developed obesity.

[0006] Therefore, there is a need to optimize the use of butyrate and its salts in animal husbandry, especially for pets (dogs, cats) and livestock (fattening turkeys, chickens, hens, sows and piglets (lactation and weaning), finishing pigs, dairy cows, calves, etc.) to improve the health of intestinal epithelial cells.

[0007] We have now found that by co-administering probiotics to animals, in particular strains of Enterococcus faecium, more specifically strain NCIMB 10415 (which is designated ) as a probiotic active ingredient, can improve the absorption of butyric acid or its salts at the intestinal level in livestock and pets.

[0008] It is therefore an object of the present invention to use Enterococcus faecium in combination with butyric acid or its salts as a feed supplement to improve the health of intestinal epithelial cells and thereby increase the health of pets and livestock.

[0009] In this context, "improving the health of the intestinal epithelium" means improving the ability of the intestinal epithelium to absorb butyrate. This can make the animal healthier, which from a practical point of view means weight gain in livestock and improvements in physiological and behavioral parameters known as pet health indices, such as appetite, sleep, skin / hair condition, etc. These health indices of animals are well known to experts in the field.

[0010] Preferably, the Enterococcus faecium strain with access number NCIMB 10415, which is designated as SF68, is used as the probiotic active ingredient. More preferably, SF68 is used in the form of microcapsules, for example, on the market as They are sold in microcapsule form under the names LBC ME10, LBC ME20 plus and LBC ME5 PET.

[0011] Butyric acid is preferably used in the form of a salt, in particular the sodium salt. It is particularly preferably used in the form of a capsule, for example, available on the market as The recipe sold by name.

[0012] In a preferred embodiment of the present invention, both probiotics and butyric acid or its salts are administered to animals mixed with a standard diet.

[0013] The amounts of probiotics and butyric acid or its salts will vary depending on the animal species and applicable local regulations.

[0014] For reference only and without any limitation, we report some butyrates in Table 1 below Examples of dosages for LBC ME10 and LBC ME20 plus for livestock and LBC ME5 PET for pets.

[0015] Table 1

[0016]

[0017] In its preferred embodiment, the invention consists in feeding animals with a standardized feed, selected by an expert in the field according to the type of animal and its reproductive characteristics (dairy, meat, etc.), supplemented with the above-mentioned amounts of butyric acid or its salts and the probiotic SF68, these amounts also being selected according to the type of animal.

[0018] The animals' physiological growth and / or weight gain is improved compared to animals fed only a standard diet.

[0019] The idea of ​​combining the probiotic SF68 with butyrate or its salts originated from experimental studies on groups of mice treated with a high-fat diet, where it was found that butyrate was better utilized / absorbed by intestinal epithelial cells when administered together with SF68 (see Example 1).

[0020] In line with the results in mice, we could observe that SF68 leads to normalization of the expression of the apical transporter of butyrate, whereas a high-fat diet (HFD) alters this expression.

[0021] In further studies on livestock, particularly chickens, it has been observed that the administration of SF68 together with butyrate or its salts resulted in an increase in the animals' body weight compared to a control group receiving only a standard diet.

[0022] This further feature of the addition of butyric acid or its salts and the probiotic Enterococcus faecium (SF68) to animal feed makes the present invention particularly advantageous.

[0023] The utility, effects and advantages of the present invention will now be described in more detail by the following examples, which are not intended to limit the scope of the present invention in any way.

[0024] Example 1

[0025] SF60 reduces fecal butyrate levels in mice fed a high-fat diet (HFD)

[0026] C57BL / 6 mice (weight 20-22 g), 5 weeks old, were provided by Envigosrl (San Pietro al Natisone, Udine, Italy). Six mice were housed in cages placed in a constant temperature room with a 12-h light cycle, a temperature of 22-24°C, and a humidity of 50-60%, and allowed to acclimate for at least one week. Animal handling followed Directive 2010 / 63 / UE.

[0027] During the adaptation period, all mice consumed a standard diet (SD, 18% of calories from fat; TD. 2018). The animals were then randomly divided into six groups of 10 mice each, as follows:

[0028] -SD lasts for 8 weeks

[0029] -HFD for 8 weeks

[0030] -SD+SF68(P) starting from week 4 (treatment group 4+4)

[0031] -HFD+SF68(P) starting from week 4 (treatment group 4+4)

[0032] -SD+SF68(P) for 8 weeks

[0033] -HFD+SF68(P) for 8 weeks

[0034] A high-fat diet (HFD) provides 60% of calories from fat (TD.06414).

[0035] The calorie content of the two diets compares as follows:

[0036]

[0037] The body weight of the animals was measured weekly starting from the first day of the study. At the end of 8 weeks, the animals were anesthetized and sacrificed. Blood samples and tissue samples were collected and stored at -80°C for further analysis.

[0038] For the analysis of short-chain fatty acids (SCFA), stool samples were lyophilized and subjected to gas chromatography. The lyophilizate was dissolved in 100 ml of 5 M formic acid and 400 ml of acetone and centrifuged (5 min at 4000 rpm). A GC column equipped with a flame ionization detector and a thin-film capillary column was used. The SCFA concentration in the supernatant was determined using a GC2010 Plus gas chromatograph (Shimadzu Deutschland GmbH, Duisburg, Germany) equipped with a GC-2010 Plus gas chromatograph (Shimadzu Deutschland GmbH, Duisburg, Germany). The samples were developed by split injection using an automatic sampler AOC-20s / I (Shimadzu Deutschland GmbH). A GC solution chromatography data system (Shimadzu Deutschland GmbH) was used for data processing. For the quantification of SCFA, an external standard (Supelco TM WSFA-1Mix, Supelco Sigma-Aldrich Co., Bellefonte PA).

[0039] The results are reported in Figure 1 middle.

[0040] Mice fed a high-fat diet (HFD) had significantly enhanced fecal butyrate levels after being fed the HFD for 8 weeks. This enhancement was counteracted by supplementation with SF68, both in the 4+4 treatment group and in the 8-week treatment group.

[0041] Example 2

[0042] Study on the effect of increasing weight of chicken

[0043] The pen numbers of the five treatment groups of birds were randomly assigned to 12 replications for a total of 60 study pens.

[0044] Table 2 - Study design

[0045]

[0046]

[0047] Each pen is equipped with a Plasson drinker and a feeding tube with a capacity of 20.4 kg. Each pen measures 5'x4' and the stocking density is 622 cm2 per bird, with 30 broilers in each pen.

[0048] The animals began to feed on the new wood chips. On day 5, all 60 pens received 226 g of litter waste from chickens that had not been exposed to dietary DFM or enzymes.

[0049] A total of 1800 (30 / pen) one-day-old Ross 308AP direct chicks were used.

[0050] On the first day of the study, 200 randomly captured chicks were weighed and the mean body weight was determined. The chicks were then distributed to pens starting from pen 1 (Table 2), and 30 chicks were randomly removed from the crates and weighed in groups so that the body weights of all 60 groups of 30 chicks were within ±10% of the determined mean body weight.

[0051] The chicks were kept illuminated throughout the study. All pens were checked at least once a day during the study to control feed and water availability and temperature.

[0052] Each feed type (day-old chicks, grower chicks, finisher chicks) was made from a single large basal diet that was formulated for each diet and treatment group to keep the sodium content of each treatment group as equal as possible.

[0053] The diet is provided according to the age of the chickens (newborn chicks from 0 to 14 days old, grower chicks from 15 to 28 days old, and finisher chicks from 29 to 42 days old). Feed formulations are reported in Figure 2a (newborn chicks), 2b (growing chicks) and 2c (finishing chicks).

[0054] All feeds are mixed on site and the expected allotment per bird for each feed type is a maximum of 567g for day-old chicks, 1590g for growers and 2270g for finishers.

[0055] Table 3 below reports a description of the supplementary diets provided to the five study groups.

[0056] Table 3

[0057]

[0058]

[0059] ButiPEARL - Butyric Acid Encapsulated Commercial Formula

[0060] ME10-E.faecium NCIMB 10415 Commercial formula

[0061] All broilers were weighed pen by pen at 0, 14, 28 and 42 days of age, and feed was weighed at 14, 28 and 42 days of age.

[0062] During the study, body weight and feed intake and associated feed conversion values ​​(FCV) of broiler chickens were measured on days 0, 14, 28 and 42.

[0063] The results obtained are reported in the table below.

[0064] Table 4: 0-14 days

[0065]

[0066] *P value from RCB ANOVA.

[0067] FCV = Feed Conversion Value

[0068] Superscript "a" and "b" = There is a significant difference between the two (p ≤ 0.05)

[0069] Table 5: 0-21 days

[0070]

[0071] *P value from RCB ANOVA.

[0072] FCV = Feed Conversion Value

[0073] Superscript "a" and "b" = there is a significant difference between the two (p ≤ 0.05)

[0074] Table 6: 0-28 days

[0075]

[0076] *P value from RCB ANOVA.

[0077] FCV = Feed Conversion Value

[0078] Superscript "a" and "b" = There is a significant difference between the two (p ≤ 0.05)

[0079] Table 7: 0-42 days

[0080]

[0081] *P value from RCB ANOVA.

[0082] FCV = Feed Conversion Value

[0083] Superscript "a" and "b" = there is a significant difference between the two (p ≤ 0.05)

[0084] Significant differences between the treated and control groups were observed starting from day 28. At the end of the study (42 days), the combination of probiotics and butyrate resulted in a numerically higher mean weight gain than the butyrate group.

Claims

1. Use of a combination of Enterococcus faecium and butyric acid or its salts as a feed supplement for improving intestinal epithelial cell health and thereby improving the health of livestock and pets.

2. The use according to claim 1, wherein Enterococcus faecium is a strain with a deposit number of NCIMB 10415.

3. The use according to claim 1, wherein The butyric acid or its salt is sodium butyrate.

4. The use according to any one of the preceding claims, wherein Enterococcus faecium is in microencapsulated form.

5. The use according to any one of the preceding claims, wherein The butyric acid or its salt is in capsule form.

6. The use according to any one of the preceding claims, wherein The livestock are fattening turkeys, chickens, laying hens, sows and lactating and weaned piglets, fattening pigs, dairy cows, calves, and pets are dogs and cats.

7. A feed supplement containing Enterococcus faecium and butyric acid or its salts for improving the health of intestinal epithelial cells and thus improving the health of livestock and pets.

8. The feed supplement according to claim 7, wherein Enterococcus faecium is a strain with a deposit number of NCIMB10415.

9. The feed supplement according to claim 7, wherein The butyric acid or its salt is sodium butyrate.

10. The feed supplement according to any one of claims 7 to 9, wherein Enterococcus faecium is in microencapsulated form.

11. The feed supplement according to any one of claims 7 to 10, wherein The butyric acid or its salt is in non-encapsulated form.

12. The feed supplement according to any one of claims 7 to 11, wherein The livestock are fattening turkeys, chickens, laying hens, sows and lactating and weaned piglets, fattening pigs, dairy cows, calves, and pets are dogs and cats.

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