Synbiotic compositions and uses

Abstract

The invention relates to a synbiotic composition comprising Bifidobacterium ssp., non- digestible oligosaccharides, and pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260. The invention further relates to a nutritional composition comprising the synbiotic composition. The invention further relates to the use of the synbiotic composition for treating or preventing gut dysbiosis, preferably antibiotic-induced gut dysbiosis.

Description

[0001] SYMBIOTIC COMPOSITIONS AND USES

[0002] FIELD OF THE INVENTION

[0003] The invention relates to a synbiotic composition comprising Bifidobacterium ssp., non- digestible oligosaccharides, and pectin polysaccharides having a molecular weight from 50 to 2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260. The invention further relates to a nutritional composition comprising the synbiotic composition of the invention, and the use of the compositions for treating or preventing gut dysbiosis, preferably antibiotic-induced gut dysbiosis.

[0004] BACKGROUND OF THE INVENTION

[0005] Dysbiosis is defined by an imbalance in bacterial composition, changes in bacterial metabolic activities, or changes in bacterial distribution within the gut. The three types of dysbiosis are: 1) loss of beneficial bacteria, 2) overgrowth of potentially pathogenic bacteria, and 3) loss of overall bacterial diversity. In most cases, these types of dysbiosis occur at the same time. The causes often relate to impairment of the microbiota, antibiotic treatment, alcohol, inappropriate diet, malnutrition, infections (including viral infections), and the like. Antibiotic-induced dysbiosis affects the microbiota resilience and poses a challenge even for healthy adult subjects in returning to normobiosis (normal diversity of gut microbiome). The effects of antibiotics on gut microbiota composition and diversity can last from weeks to months.

[0006] One of the features exhibited in a dysbiotic gut microbiome is the shift towards lower anaerobic microorganisms and higher aerotolerant microorganisms. This tends to occur because of the increased oxidative stress (Winter, S. and Baumler, A., Gut dysbiosis: Ecological causes and causative effects on human disease, PNAS 2023 vol. 120(50) https: / / doi.org / 10.1073 / pnas.2316579120). It is known that a positive fecal redox potential has been correlated to the ratio of relative abundance of aerotolerant to strict anaerobes (Million M and Raoult D, Unking gut redox to human microbioma. Human Microbiome Journal, vol 10 (27-32), December 2018).

[0007] Probiotic bacteria and prebiotic fibres are the most used tool in treating gut microbiota dysbiosis, alone or in combination. There is great variability in the modulatory and reparatory activity of probiotics and prebiotics, which is highly dependent on their inherent structural features. For instance, polymeric pectin with a complex structure has been described as a promoter of proliferation of their utilizing microbes (Cantu-Jungles & Hamakera, New View on Dietary Fiber Selection for Predictable Shifts in Gut Microbiota. mBio. 2020 Feb 18;11(1):e02179-19. doi: 10.1128 / mBio.02179-19. Erratum in: mBio. 2020 May 26;11(3): PMID: 32071263; PMCID: PMC7029134.) whereas some, but not all, oligomeric pectin has been reported to stimulate gut bifidogenic effects.

[0008] WO2021151956 relates to compositions for treating and / or preventing intestinal and / or pulmonary disease associated with gut dysbiosis, the compositions comprising acetate- overproducing bacteria hypersensitive to oxygen (AOHO bacteria).

[0009] Onumpai, Kolida, Bonnin, & Rastall (Microbial utilization and selectivity of pectin fractions with various structures. Appl Environ Microbiol. 2011 Aug 15;77(16):5747-54. doi: 10.1128 / AEM.00179-11. Epub 2011 Jul 1. PMID: 21724897; PMCID: PMC3165253.) reported that the arabino-oligosaccharides (DP2-11) and galacto-oligosaccharides (DP2-9) components of pectin produced bifidogenic effect, while homogalacturonan-oligosaccharides (DP1-23) and rhamnogalacturonan-oligosaccharides (DP2-19) do not.

[0010] WO2018 / 067002 describes synbiotic compositions for the prevention of dysbiosis. The compositions comprise Bifidobacterium and dietary fiber (Fructo-oligosaccharides).

[0011] W02009 / 067000 describes compositions comprising Lactobacillus and Bifidobacterium ssp. DN-173010, GOS, a pectin-derived product (galacturonic acid oligosaccharides) and inulin. The DP of the galacturonic acid oligosaccharide used is in the range of 2 - 250. The pectinderived fraction is described as useful to reduce adhesion of pathogenic microorganisms to the intestinal epithelial cells.

[0012] W02006 / 112714 describes compositions comprising Lactobacillus or Bifidobacterium, pectin derivatives and prebiotic indigestible fibres such as GOS and / or FOS for prevention of pathogenic infections. The pectin derivatives have a DP of 4.

[0013] Fermentability and prebiotic properties of pectin oligomers are thus not yet fully explored nor can it be readily predictable.

[0014] The overall health of an individual is linked to an optimal gut microbiota state. It is thus relevant to support the quick re-establishment of normobiosis after a dysbiotic assault. Therefore, the search for compositions capable of optimizing the gut microbiota restoration after dysbiotic state is important for the development of microbiome-based personalized nutrition. In particular, there remains a need to provide a composition capable of aiding recovery from dysbiosis particularly caused by antibiotic-therapy.

[0015] SUMMARY OF THE INVENTION

[0016] The present inventors have observed that the addition of Bifidobacterium ssp. to a combination of prebiotic fibres comprising non-digestible oligosaccharides and pectin polysaccharides with a high degree of methylation is capable of optimizing the recovery from dysbiotic state, such as that caused after treatment with antibiotics.

[0017] Accordingly, a first aspect of the invention relates to a synbiotic composition comprising:

[0018] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0019] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0020] • 0.5-4 wt.% on dry basis of a pectin polysaccharide having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35%.

[0021] The invention also relates a synbiotic composition comprising:

[0022] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0023] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0024] • 0.5-4 wt.% on dry basis of a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0025] The present invention also relates to a nutritional composition comprising the synbiotic composition of the invention, macro ingredients selected from digestible carbohydrates, fats, proteins and preferably further comprising a therapeutically effective combination of:

[0026] • omega-3 fatty acids;

[0027] • vitamin B2;

[0028] • vitamin D3, and

[0029] • optionally a choline source selected from choline or a salt thereof and / or phosphatidylcholine. A further aspect of the invention is the use of the synbiotic composition or nutritional composition comprising the synbiotic composition according to the invention in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably an antibiotic-induced dysbiosis.

[0030] It was unexpectedly found that the synbiotic composition or nutritional composition comprising scGOS, IcFOS, pectin polysaccharide and Bifidobacterium ssp have an increased effect on the relative abundance (%) of Bifidobacteriaceae, whereas the composition comprising galacturonic acid oligosaccharides (AOS) did not provide an increase in the relative abundance (%) of Bifidobacteriaceae to the same extent. Moreover, it was surprisingly found that the composition according to the invention synergistically increases both acetic acid and butyric acid production. The composition according to the invention provided a much stronger effect compared to a composition comprising AOS.

[0031] Furthermore, the effect of the composition according to the invention was explored on epithelial barrier function. A significant increase in trans epithelial electrical resistance (TEER) was observed for the composition comprising the pectin according to the invention compared to a composition comprising AOS. An increase in TEER is indicative of an improvement of gut barrier function.

[0032] DRAWINGS

[0033] Figure 1 : MAPI (Metagenomic Aerotolerant Predominance Index) in all treatment groups and starting inoculum (I NO).

[0034] Figure 2: Comparison of relative abundance (%) of Bifidobacteriaceae family at 72h between treatments.

[0035] Figure 3A: Acetic acid finding at 72 hours of anaerobic fermentation with GOS, FOS, LvPectin 9:1 :2 with or without BB536, GOS, FOS, AOS 9:1 :2 with or without BB536 and BB536 alone. Figure 3B: Butyric acid finding at 72 hours of anaerobic fermentation with GOS, FOS, LvPectin 9:1 :2 with or without BB536, GOS, FOS, AOS 9:1 :2 with or without BB536 and BB536 alone. Figure 4: TEER values after 21 h treatment of the Caco-2 cells with the fermented samples of GOS, FOS, LvPectin 9:1 :2 + BB536 and GOS, FOS, AOS 9:1 :2 + BB536.

[0036] DETAILED DESCRIPTION OF THE INVENTION

[0037] In a first aspect, the present invention relates to a synbiotic composition comprising:

[0038] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.; • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0039] • 0.5-4 wt.% on dry basis of a pectin polysaccharide having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35%.

[0040] In a second aspect, the present invention relates to a synbiotic composition comprising:

[0041] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0042] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0043] • 0.5-4 wt.% on dry basis of a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0044] In a third aspect, the present invention relates to a nutritional composition comprising the synbiotic composition of the invention. The invention further relates to the use of the synbiotic composition of the invention or nutritional composition comprising the same in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis.

[0045] In some jurisdictions, the third aspect of the invention can be defined as the use of the synbiotic composition according to the invention and / or nutritional composition comprising the same for the manufacture of a product for use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis, the synbiotic composition comprising:

[0046] • 0.5-2.5 x 108. cfu / gram on dry basis of Bifidobacterium ssp.;

[0047] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0048] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0049] The invention may likewise be defined as the use of the Bifidobacterium ssp., non-digestible oligosaccharides and a pectin polysaccharide in the manufacture of a synbiotic composition or nutritional composition comprising the same for use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis, the synbiotic composition comprising:

[0050] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0051] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0052] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0053] The invention may also be defined as the use of the Bifidobacterium ssp., non-digestible oligosaccharides and a pectin polysaccharide in the manufacture of a synbiotic composition or nutritional composition comprising the same for use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis, the synbiotic composition comprising:

[0054] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0055] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0056] • 0.8-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0057] In yet other jurisdictions, the invention may be defined as a method of preventing and / or treating gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis, the method comprising administration of a synbiotic composition and / or nutritional composition comprising the same, the synbiotic composition comprising:

[0058] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0059] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0060] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0061] The present invention also encompasses the non-therapeutic use of the synbiotic composition of the invention and / or nutritional composition comprising the same for repairing the gut dysbiotic state of an adult subject after being subjected to antibiotic therapy. It may further relate to the non-therapeutic use of the synbiotic composition and / or nutritional composition according to the invention for improving the gut barrier function in a healthy adult subject. The invention may further relate to the non-therapeutic use of the synbiotic composition and / or nutritional composition of the invention for treating a compromised gut barrier in a healthy adult subject.

[0062] The concentration of pectin polysaccharides and their monosaccharide composition can be determined by analytical techniques known to the skilled person. After acid hydrolysis, the monosaccharide composition can suitably be determined by High Performance Anion Exchange Chromatography combined with Pulse Amperometric Detection (HPAEC-PAD).

[0063] The molecular size distribution can be determined by High Performance Size-Exclusion Chromatography using refractive index (Rl) detection (concentration), light scattering detection (molecular mass detection), UV detection (indicative for presence of proteins) and differential pressure detection (intrinsic viscosity detection).

[0064] The above mentioned analytical methods are described in: Analytical Biochemistry Vol. 207, Issue 1 , 1992, p. 176 (for neutral sugar analysis) and in Mol. Nutr. Food Res., Vol 61 , Issue 1 , 2017, 1600243 (for the galacturonic acid analysis and the molecular size distribution).

[0065] Unless indicated otherwise, all weight percentage figures are provided herein on dry basis.

[0066] List of embodiments

[0067] 1. A synbiotic composition comprising:

[0068] ■ 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0069] ■ at least 5wt.% on dry basis of non-digestible oligosaccharides, selected from galactooligosaccharides and / or fructooligosaccharides; and

[0070] ■ 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35%. 2. The synbiotic composition of claim 1 , wherein the probiotic bacterial strain is selected from Bifidobacterium spp., preferably Bifidobacterium longum, more preferably Bifidobacterium longum BL999.

[0071] 3. The synbiotic composition of any of the preceding claims, wherein it comprises 0.8 - 2.0 x 108cfu / gram on dry basis of Bifidobacterium spp.

[0072] 4. The synbiotic composition of any of the preceding claims, wherein it comprises 5 - 15 wt.% on dry basis of non-digestible oligosaccharides, preferably 7 - 12 wt.% on dry basis of non- digestible oligosaccharides.

[0073] 5. The synbiotic composition of any of the preceding claims, wherein it comprises galactooligosaccharides and fructooligosaccharides.

[0074] 6. The synbiotic composition of any of the preceding claims, wherein the pectin polysaccharides have a molecular weight of 60-1000 kDa, more preferably 70-500 kDa, even more preferably 75 - 250 kDa.

[0075] 7. The synbiotic composition of any of the preceding claims, wherein the pectin polysaccharides comprise a substantially unbranched backbone of galacturonic acid residues, preferably wherein less than 20% molar weight of the backbone of galacturonic acid residues is branched.

[0076] 8. The synbiotic composition of any of the preceding claims, wherein the backbone of the pectin polysaccharides comprise galacturonic acid residues and rhamnose residues in a molar ratio of more than 10:1 , preferably more than 20:1.

[0077] 9. The synbiotic composition of any of the preceding claims, the pectin polysaccharides having a degree of acetylation (DA) of less than 18%, preferably less than 15%, more preferably less than 10%, preferably less than 5%, even more preferably less than 3%.

[0078] 10. The synbiotic composition of any of the preceding claims, wherein the weight ratio of the sum of non-digestible oligosaccharides to pectin polysaccharides is between 2 : 1 to 30 : 1 , preferably 3 : 1 to 20 : 1. 11. The synbiotic composition of any of the preceding claims, wherein it comprises less than 20 wt.% on dry basis of inulin, preferably less than 10 wt.%, more preferably less than 5 wt.%.

[0079] 12. A nutritional composition comprising the synbiotic composition according to any one of the preceding claims, macro ingredients selected from digestible carbohydrates, fats, proteins, and preferably further comprising a therapeutically effective combination of:

[0080] ■ omega-3 fatty acids;

[0081] ■ vitamin B2;

[0082] ■ vitamin D3, and

[0083] ■ optionally a choline source selected from choline or a salt thereof and / or phosphatidylcholine.

[0084] 13. A composition according to any one of the preceding claims for use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis.

[0085] 14. The composition for use of claims 13, wherein the subject is or has been submitted to antibiotic therapy for at least 4 days, preferably at least 7 days.

[0086] 15. The composition for use of claims 13 or 14, wherein the composition is for use after interruption of antibiotic therapy, preferably at least 4 days after interruption of antibiotic therapy.

[0087] Bifidobacterium strains

[0088] Bifidobacterium strains are herein provided as a probiotic bacterial strain in the compositions of the invention. The term “probiotic” as used herein refers to micro-organisms which, when orally administered in adequate amounts, provide a health benefit. As such, the amounts of Bifidobacterium ssp. strains according to the invention are effective for providing a health benefit. Preferably, the strains used herein are selected from viable micro-organisms, non- viable micro-organisms, fragments of micro-organisms and combinations thereof, preferably viable micro-organisms.

[0089] The synbiotic composition preferably comprises 0.6 x 108to 2.3 x 108cfu / gram on dry basis of Bifidobacterium spp, more preferably 0.8 x 108to 2.0 x 108cfu / gram on dry basis of Bifidobacterium spp. Expressed in calories, the synbiotic composition according to the invention preferably comprises from 1.0 x 108to 7.5 x 108cfu per 100 kcal, more preferably from 2.0 x 108to 6.0 x 108cfu per 100 kcal, most preferably from 2.5 x 109to 5.0 x 109cfu per 100 kcal based on total energy content of the synbiotic composition. Preferably, the Bifidobacterium spp. is selected from the group consisting of B. longum, B. breve, B. infantis, B. animalis, B. lactis and B. bifidum, most preferably B. longum. In one preferred embodiment, the species is Bifidobacterium longum, even more preferably Bifidobacterium longum, official deposit no.: ATCC BAA-999; NCC No.: NCC 3001; “BL999”. B. longum (ATCC BAA-999), also referred to as BL999, may be obtained from Morinaga Milk Indus try Co. Ltd. of Japan under the trade mark BB536 which is commercially available.

[0090] “Non-digestible oligosaccharides’’ (NDO)

[0091] The term “non-digestible oligosaccharides” (NDO) as used in the present invention refers to oligosaccharides which are not digested in the intestine by the action of acids or digestive enzymes present in the human upper digestive tract, e.g. small intestine and stomach, but which are preferably fermented by the human intestinal microbiota. For example, glucose, galactose, sucrose, lactose, maltose and maltodextrins are considered digestible.

[0092] In a preferred embodiment, the synbiotic composition comprises at least 8 wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides (GOS) and / or fructooligosaccharides (FOS). The synbiotic composition may comprise for instance 5 - 15 wt.% on dry basis of non-digestible oligosaccharides, preferably 7 - 12 wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides (GOS) and / or fructooligosaccharides (FOS).

[0093] The non-digestible oligosaccharide is selected from the group consisting of fructooligosaccharide and galacto-oligosaccharide, and combinations thereof. Preferably, the composition comprises non-digestible oligosaccharides with a degree of polymerization (DP) in the range of 2 to 250, more preferably 3 to 60.

[0094] Suitable galacto-oligosaccharides (GOS) include, but are not limited to, beta-galacto- oligosaccharide and alpha-galacto-oligosaccharide. Preferably, the galacto-oligosaccharide has beta-(1 ,4), beta-(1 ,3) and / or beta-(1 ,6) glycosidic bonds and a terminal glucose. According to one embodiment, the composition comprises non-digestible galactooligosaccharide comprising beta1 ,4 linkages, in particular beta1 ,4 linkages between the galactose units, and having a degree of polymerization of at least 4, preferably with a degree of polymerization from 2 to 10, more preferably in the range of 3 to 7. A commercially available source of GOS is VivinalOGOS from FrieslandCampina Domo (Amersfoort, The Netherlands). Preferably the synbiotic composition according to the present invention comprises at least 5 wt.% GOS, on dry basis of the composition, more preferably from 6 to 12 wt%, most preferably between 7 and 10 wt% on dry basis of the composition.

[0095] The preferred fructo-oligosaccharides (FOS) have a DP or average DP in the range of 2 to 250, more preferably 3 to 100, even more preferably above 20, most preferably 10 to 60. FOS suitable for use in the composition of the invention is also readily commercially available, e.g. RaftilineHP® (Orafti). Preferably the synbiotic composition according to the present invention comprises at least 0.1 wt.% FOS, on dry basis of the composition, more preferably between 0.2 and 3 wt%, most preferably between 0.5 and 2 wt%.

[0096] In one embodiment, based on calories, preferably the synbiotic composition according to the invention may comprise based on dry weight from 0.5 to 6 grams GOS per 100 kcal, more preferably from 1 to 4 grams per 100 kcal, most preferably from 1.5 to 3 grams GOS per 100 kcal based on total energy content of the synbiotic composition. In another embodiment, based on calories, preferably the synbiotic composition according to the invention may comprise based on dry weight from 0.1 to 3 grams FOS per 100 kcal, more preferably from 0.1 to 2 grams FOS per 100 kcal, most preferably from 0.1 to 1 grams FOS per 100 kcal based on total energy content of the synbiotic composition.

[0097] In a further embodiment, based on calories, preferably the synbiotic composition according to the invention may comprise based on dry weight from 0.5 to 6 grams non-digestible oligosaccharides per 100 kcal, more preferably from 1 to 4 grams per 100 kcal, most preferably from 1.5 to 3.5 grams non-digestible oligosaccharides per 100 kcal based on total energy content of the synbiotic composition.

[0098] Preferably, the synbiotic composition comprises galactooligosaccharides and fructooligosaccharides. Preferably the combination of GOS and FOS is present in a weight ratio of from 1 :99 to 99: 1 , more preferably from 1 : 19 to 19: 1 , more preferably from 1 : 1 to 19: 1 , more preferably from 2:1 to 15:1 , more preferably from 5:1 to 12:1 , even more preferably from 8:1 to 10:1 , even more preferably in a ratio of about 9:1.

[0099] Pectin polysaccharides

[0100] The term “pectin polysaccharides” as used herein refers to optionally branched polysaccharides comprising a backbone that substantially consists of galacturonic acid residues. The terms “backbone chain” and “backbone” are synonyms. Preferred pectin polysaccharides according to the invention are derived from fruits, preferably derived from citrus, apple, grapes, berries , more preferably derived from apple.

[0101] The pectin polysaccharides according to the invention have a preferred molecular weight between 60-1000 kDa, more preferably 70-500 kDa, even more preferably 75 - 250 kDa.

[0102] The backbone of the pectin polysaccharides according to the invention preferably comprises at least 70% molar weight of 1 ,4-linked a-D-galacturonic acid residues, more preferably at least 80% molar weight of 1 ,4-linked a-D-galacturonic acid residues, even more at least 90% molar weight of 1 ,4-linked a-D-galacturonic acid residues.

[0103] Accordingly, in a preferred embodiment, the pectin polysaccharides comprise a substantially unbranched backbone of galacturonic acid residues. In other words, the backbone of the pectin polysaccharides preferably comprises less than 20% by weight of the polysaccharide of branched galacturonic acid residues, more preferably less than 15% by weight of the polysaccharide of branched galacturonic acid residues.

[0104] Preferably, the pectin polysaccharides of the invention comprises less than 30% by weight of the polysaccharide of rhamnose residues, preferably less than 20%, more preferably less than 10%, even more preferably less than 5%, most preferable less than 3% by weight of the polysaccharide of rhamnose residues. Herein, the rhamnose residues are in the backbone of the pectin polysaccharides.

[0105] The low abundance of rhamnose residues in the backbone of the polysaccharides of the invention may also be expressed by the ratio between galacturonic acid residues and rhamnose residues. Accordingly, in a preferred embodiment, the backbone of the pectin polysaccharides comprises galacturonic acid residues and rhamnose residues in a molar ratio of more than 10: 1 , preferably more than 20: 1.

[0106] The carboxyl groups of the galacturonic acid residues within the backbone of the pectin polysaccharides of the invention are typically esterified. Esterified galacturonic acid may occur in the form of the methyl ester or acetyl ester. The pectin polysaccharides according to the invention have a high degree of methylation (DM). Preferably, the DM of the pectin polysaccharides in the present composition is of at least 45%, more preferably of at least 50%, even more preferably at least 60%. The degree of acetylation (DA) of the pectin polysaccharides in the present composition is typically very low. Accordingly, the preferred DA is less than 18%, more preferably less than 15%, even more preferably less than 10%, most preferably less than 5%, most preferably less than 3%.

[0107] Pectin polysaccharides are formed by domains. The term “domain” as used herein refers to the backbone with any sidechains that may attached thereto. Backbone refers to the sequence of glycosidically linked monosaccharides within the backbone of a polysaccharide, excluding any sidechains that are attached thereto. Preferably, the pectin polysaccharides of the invention are rich in a domain called homogalacturonan (HG). Typically, HG domains do not contain any sidechains. Besides HG-I domains, the pectin polysaccharides of the present invention may contain small amounts of one or more of the following domains:

[0108] • xylogalacturonan (XG),

[0109] • apiogalacturonan (AG),

[0110] • rhamnogalacturonan-l (RG-I),

[0111] • rhamnogalacturonan-l I (RG-I I).

[0112] The backbone of the pectin polysaccharides of the invention is majorly comprised of homogalacturonan (HG) domains, preferably more than 35%, more preferably more than 40% by weight of the polysaccharide is HG domain. HG domains are substantially free of rhamnose residues in the polysaccharide backbone and are substantially unbranched.

[0113] According to the invention, the sum of XG, AG, RG-I and RG-I I domain represent no more than 45 wt.% on dry basis of the polysaccharide, preferably no more than 30 wt%, more preferably no more than 20 wt.% on dry basis of the polysaccharide.

[0114] In a preferred embodiment, the pectin polysaccharide of the invention has a degree of polymerisation (DP) of less than 6000 and / or a molecular weight of less than 150 kDa. Commercially available pectins according to the invention include e.g., Herbapekt LV and Pectin GENU® from CP Kelco, Europe.

[0115] Degree of polymerization (DP) as used herein refers to the number of monomeric saccharide units in a (poly- or oligo-)saccharide that are linked by glycosidic bonds.

[0116] In a preferred embodiment the low viscosity pectin has a degree of polymerisation of less than 6000 or a molecular weight of less than 150 kDa. The pectin polysaccharides according to the invention have a preferred molecular weight of 60-1000 kDa, more preferably 70-500 kDa, more preferably 75 - 250 kDa, more preferably less than 150 kDa, even more preferably 50 - 150 kDa, most preferably 80 - 120 kDa.

[0117] In a preferred embodiment, the pectin polysaccharide of the invention has a degree of polymerisation (DP) of less than 6000, and / or a molecular weight of less than 150 kDa. The DP is preferably at least 260, more preferably at least 300, even more preferably at least 450. Preferably the pectin polysaccharides have a DP of less than 6000, more preferably less than 4000, more preferably less than 2000, even more preferably less than 1000.

[0118] The carboxyl groups of the galacturonic acid residues within the backbone of the pectin polysaccharides of the invention are typically esterified. The pectin polysaccharides according to the invention have a high degree of esterification. Preferably, the esterification of the pectin polysaccharides is more than 50%, preferably more than 60% even more preferably more than 65%, most preferably more than 70%. The esterification of the pectin polysaccharides is preferably 50 - 80%, more preferably 50 - 75%, even more preferably 60 - 72%.

[0119] Esterified galacturonic acid may occur in the form of the methyl ester or acetyl ester. The pectin polysaccharides according to the invention have a high degree of methylation (DM). Preferably, the DM of the pectin polysaccharides in the present synbiotic composition and / or nutritional composition is of at least 45%, more preferably of at least 50%, even more preferably at least 55 or at least 58%. The degree of acetylation (DA) of the pectin polysaccharides in the present synbiotic composition and / or nutritional composition is typically very low. Accordingly, the preferred DA is less than 18%, more preferably less than 15%, even more preferably less than 10%, most preferably less than 5%, most preferably less than 3%.

[0120] Preferably, the pectin polysaccharide of the invention comprises less than 30% by weight of the polysaccharide of rhamnose residues, preferably less than 20%, more preferably less than 10%, even more preferably less than 5%, most preferable less than 4% by weight of the polysaccharide of rhamnose residues. Herein, the rhamnose residues are in the backbone of the pectin polysaccharides.

[0121] The present inventors have surprisingly found that the addition of Bifidobacterium ssp. to prebiotic fibres particularly selected from NDO (GOS and / or FOS) and pectin polysaccharides as described herein is superior in recovering the gut microbiome of an adult from antibiotic- induced dysbiosis as compared to the addition of the same species to other known prebiotic fibres, such as inulin. Inulin is a type of fructopolysaccharide wherein at least 75% of the glycosidic linkages are 3(2,1) linkages. Inulin is known to be effective to mitigate dysbiosis. The inventors have surprisingly shown that adding a probiotic Bifidobacterium ssp. to inulin as prebiotic fiber did not lead even to an additive effect on mitigating dysbiosis, whereas the combination of probiotics and prebiotics of the invention provides such unexpected effect. Typically, inulin has an average chain length between 8 and 60 monosaccharide units, and is commercially available under the trade name RaftilineOHP (Orafti). Accordingly, in a preferred embodiment, the synbiotic composition and / or nutritional composition comprises less than 20 wt.% on dry basis of fructopolysaccharides, preferably less than 10 wt.%, more preferably less than 5 wt.%, most preferably the synbiotic composition and / or nutritional composition does not comprise fructopolysaccharides.

[0122] Preferably, the synbiotic composition and / or nutritional composition comprises a weight ratio of the sum of non-digestible oligosaccharides to pectin polysaccharides of between 2 : 1 to 30 : 1 , more preferably 3 : 1 to 20 : 1 , even more preferably between 4 : 1 to 10 : 1.

[0123] In an embodiment, the synbiotic composition and / or nutritional composition comprises a weight ratio of the sum of non-digestible oligosaccharides to pectin polysaccharides of between 5 : 1 to 40 : 1 , more preferably between 10 : 1 to 30 : 1 , most preferably between 15 : 1 to 25 : 1.

[0124] In a further preferred embodiment, the non-digestible oligosaccharides are galactooligosaccharides and fructooligosaccharides in a weight ratio of 2:1 to 15:1 , more preferably 9:1. In some embodiments, the weight ratio of galactooligosaccharides to fructooligosaccharides to pectin polysaccharides is in the range of 5 - 15 : 0.5 - 5 : 1 - 10, more preferably 5 - 13 : 0.7 - 2 : 1.5 - 4, most preferably 7 - 10 : 0.8 - 1.5 : 1.8 - 3.

[0125] In one embodiment, based on calories, preferably the synbiotic composition according to the invention may comprise based on dry weight from 0.1 to 3 grams pectin polysaccharides per 100 kcal, more preferably from 0.1 to 2 grams per 100 kcal, most preferably from 0.2 to 1 grams pectin polysaccharides per 100 kcal based on total energy content of the synbiotic composition.

[0126] The nutritional composition according to the invention preferably comprises at least 30 mg pectin polysaccharides per 100 kcal, more preferably 50 - 350 mg, even more preferably 60 - 300 mg pectin polysaccharides per 100 kcal of the composition. In a further preferred embodiment, the composition comprises at least 0.1 wt.% pectin polysaccharides based on dry weight of the composition, more preferably 0.1 - 1 wt.%, most preferably 0.2 - 0.8 wt% pectin polysaccharides. In one embodiment, when the composition is in liquid form, preferably the composition according to the invention comprises at least 100 mg pectin polysaccharides per 100 ml of the composition, even more preferably 200 - 800 mg, most preferably 250 - 700 mg based on 100 ml of the composition.

[0127] Synbiotic compositions

[0128] The term “synbiotic composition” as used herein refers to a combination of probiotics and prebiotics, preferably that work together to promote healthy gut microbiome.

[0129] The synbiotic composition according to the invention may in a preferred aspect be used as a nutritional supplement, i.e., as an additive to a diet. The supplement, preferably for enteral application, may be a solid or liquid galenical formulation. Examples of solid galenical formulations are tablets, capsules (e.g. hard or soft shell gelatine capsules), pills, sachets, powders, granules and the like which contain the active ingredient together with conventional galenical carriers. Any conventional carrier material can be utilized. The carrier material can be organic or inorganic inert carrier material suitable for oral administration. Suitable carriers include water, gelatine, gum Arabic, lactose, starch, magnesium stearate, talc, vegetable oils, and the like. Additionally, additives such as flavouring agents, preservatives, stabilizers, emulsifying agents, buffers and the like may be added in accordance with accepted practices of nutritional and pharmaceutical compounding.

[0130] Typically, the synbiotic compositions of the invention are in powder form for reconstitution or in liquid form, preferably in powder from for reconstitution. The compositions are preferably for enteral administration, more preferably oral administration.

[0131] The synbiotic composition according to the invention preferably comprises, based on dry weight, at least 0.1 grams of pectin polysaccharides per 100 kcal more preferably 0.1 to 2 g, even more preferably 0.2 - 1 g pectin polysaccharides per 100 kcal based on total energy content of the synbiotic composition. Based on dry weight, the synbiotic composition according to the invention preferably comprises at least 0.5 g pectin polysaccharides per 100 g, more preferably from 0.5 to 4 g, even more preferably from 1to 3 g per 100 g.

[0132] Nutritional composition

[0133] The invention also relates to a nutritional composition comprising the synbiotic composition of the invention and other ingredients. In a preferred embodiment, the nutritional composition comprises in addition to the synbiotic composition, a therapeutically effective combination of omega-3 fatty acids; vitamin B2 vitamin D3 and optionally a choline source selected from choline or a salt thereof and / or phosphatidylcholine (PC).

[0134] If a choline source is present in the nutritional composition preferably at least 40 mol % choline is provided in the form of phosphatidylcholine, preferably the molar ratio of choline or a salt thereof to PC is in the range from 40:60 to 60:40. As herein referred to, the molar ratio of choline and PC is referred to as the mole% choline derived from a choline salt and PC.

[0135] In a further preferred embodiment the nutritional composition further comprises one or more components selected from vitamin A, C and E, phospholipids, selenium, magnesium and further B vitamins.

[0136] The nutritional composition according to the invention may be used as a pharmaceutical product or preferably a nutritional product. In one embodiment, the present nutritional composition is a solid (typically a powder or tablet, preferably a powder) which is reconstitutable with a liquid, preferably with water, to obtain a liquid composition. Preferably the composition is administered orally.

[0137] In one aspect, the nutritional composition according to the invention may be used as a pharmaceutical product comprising one or more pharmaceutically acceptable carrier materials. Any conventional carrier material can be utilized. The carrier material can be organic or inorganic inert carrier material suitable for oral administration. Suitable carriers include water, gelatine, gum Arabic, lactose, starch, magnesium stearate, talc, vegetable oils, and the like. Additionally, additives such as thickeners, flavouring agents, preservatives, stabilizers, emulsifying agents, buffers and the like may be added in accordance with accepted practices of pharmaceutical compounding. While the individual active ingredients are suitably administered in a single composition, they may also be administered in individual dosage units.

[0138] In a preferred aspect, the nutritional composition according to the invention may be used as a nutritional product, for example as a nutritional supplement, e.g. as an additive to a normal diet, as a fortifier, to add to a normal diet, or as a complete nutrition. The nutritional product preferably comprises at least one component, preferably all components, selected from the group of fats, proteins, and carbohydrates. It is understood that a nutritional product differs from a pharmaceutical product by the presence of nutrients which provide nutrition to the subject to which the composition is administered, in particular the presence of protein, fat, and digestible carbohydrates. It may further contain ingredients such as minerals, vitamins, organic acids, and flavouring agents. Although the term "nutraceutical product" is often used in literature, it denotes a nutritional product with a pharmaceutical component or pharmaceutical purpose. Hence, the nutritional composition according to the invention may also be used in a nutraceutical product.

[0139] In one embodiment, the nutritional composition comprises a lipid fraction, wherein the lipid fraction provides between 15 and 50 energy% of the composition. In one embodiment, the nutritional composition is in the form of an oral nutritional supplement having a caloric density of between 2.2 and 2.6 kcal per ml. The energy provided by nutrients is calculated using the Atwater calculation factors of 9 kcal per g lipid, 4 kcal per gram protein or gram digestible carbohydrate, 2 kcal per gram fiber and zero kcal for the other components in the product.

[0140] The amount of lipid fraction can be determined by applying the methods known in the art for measuring fat content in the food matrix as applicable. For example, fat content for general foods is determined by applying AOAC(R) official method 983.23, while the Roese-Gottlieb method (AOAC(R) 932.06) is better applicable for products based on dried milk (Lehner, R., Estoppey, A., (1954) Mitt. Lebensmitteluntersuchung Hyg. 54:183-185). The amount of individual lipid components can be determined by applying methods specifically designed for measuring that specific component or by fractionating the fat fraction isolated from the extraction of the chloroform-niethanol fraction as given in the 983.23 method.

[0141] Suitable sources of omega-3 fatty acids for use in the composition include but are not limited to fish oil, krill oil, algae oil, preferably fish oil.

[0142] Suitable sources of further lipids for use in the nutritional composition include but are not limited to soybean oil, rape seed oil (such as colza oil, low erucic acid rape seed oil and canola oil), high oleic sunflower oil, coconut oil, high oleic safflower oil and olive oil. An alternative option are lipids obtained from milk from non-human mammals, preferably cow's milk, goat milk, sheep milk, horse milk, buffalo milk, yak milk, reindeer milk, donkey milk and camel milk, particularly cow's milk and / or goat milk. Milk lipid is sometimes also referred to as milk fat or butter fat.

[0143] The nutritional composition according to the invention thus preferably comprises a lipid fraction, preferably a lipid fraction suitable for nutrition as known in the art. Based on calories, preferably the nutritional composition according to the invention may comprise 3 to 5.5 g of lipid per 100 kcal, preferably 3.5 to 5 g per 100 kcal, more preferably 3.8 to 4.5 g of lipid per 100 kcal based on the total energy content of the composition. The lipid fraction includes polyunsaturated fatty acids and optionally also mono-unsaturated fatty acids (MLIFAs) and saturated fatty acids (SFA).

[0144] The total protein that is present in the nutritional composition, i.e., the combination of all proteins present, may also be referred to as the “protein fraction” of the nutritional composition. Based on calories, preferably the nutritional composition according to the invention may comprise 4 to 7 g of protein fraction per 100 kcal, preferably 4.5 to 6.5 g of protein fraction per 100 kcal, more preferably 5 to 6 g of protein fraction per 100 kcal based on the total energy content of the composition.

[0145] Suitable protein sources may be based on cows' milk proteins such as whey, casein, and mixtures thereof and vegetable proteins such as those based on soy, potato, pea, rapeseed and the like. The nutritional composition preferably further comprises a carbohydrate fraction. Based on calories, preferably the nutritional composition according to the invention may comprise 8.5 to 10.5 g of carbohydrates per 100 kcal, preferably 9 to 10 g per 100 kcal, more preferably 9.2 to 9.6 g of carbohydrates per 100 kcal based on the total energy content of the composition. The composition according to the invention preferably comprises 4 to 7 g protein per 100 kcal, 3 to 5.5 g lipids per 100 kcal and 8.5 to 10.5 g carbohydrates per 100 kcal.

[0146] Suitable carbohydrate sources include lactose, glucose, sucrose, fructose, galactose, maltose, starch, isomaltulose, and maltodextrin.

[0147] Administration of the nutritional composition occurs preferably at least one time per day, more preferably one or two times per day, although alternative dosage regimes can be determined from these numbers.

[0148] Omega-3 fatty acids

[0149] The present nutritional composition according to the invention may comprise omega-3 polyunsaturated fatty acids (PLIFA), wherein the PLIFAs are n-3 LC-PUFAs (long-chain PLIFAs). In an embodiment, the nutritional composition comprises at least docosahexaenoic acid (22:6, co-3; DHA). Preferably the nutritional composition comprises two omega-3 long- chain polyunsaturated fatty acids (LC-PUFA; having a chain length of 18 and more carbon atoms) wherein one omega-3 fatty acid is DHA and the second omega-3 fatty acid is selected from eicosapentaenoic acid (20:5, co-3; EPA) and docosapentaenoic acid (22:5 co-3; DPA), preferably the second omega-3 fatty acid is EPA. EPA is converted to DPA (w-3), increasing subsequent conversion of DPA to DHA in the brain. In an alternative embodiment, the nutritional composition comprises only DHA as omega-3 fatty acid and preferably comprises no EPA.

[0150] The DHA, EPA and / or DPA may be provided in any form such as, but not limited to, triglycerides, diglycerides, monoglycerides, free fatty acids or their salts or esters, phospholipids, lysophospholipids, glycerol ethers, lipoproteins, ceramides, glycolipids or combinations thereof. Preferably, the present composition comprises at least DHA in triglyceride form. Suitable co-3 LCPLIFA and / or sources of DHA and EPA include tuna oil, (other) fish oils, DHA-rich alkyl esters, algae oil, egg yolk, krill oil or phospholipids enriched with w-3 LCPLIFA e.g. phosphatidylserine-DHA. Preferably, the nutritional composition or composition for use according to the invention comprises fish oil providing the omega-3 LCPUFA(s).

[0151] DHA is preferably administered in an amount of 100 to 1250 mg per day, more preferably 150 to 1200 mg per day, even more preferably 200 to 1100 mg per day, most preferably 250 to 1000 mg per day. In one embodiment when the nutritional composition is in the form of a oral nutritional supplement the daily amount of DHA administered is preferably 150 to 850 mg per day, more preferably 200 to 800 mg per day, most preferably 250 to 750 mg per day.

[0152] Based on calories, preferably the nutritional composition according to the invention comprises 20 to 100 mg DHA per 100 kcal, preferably 25 to 90 mg DHA per 100 kcal, more preferably 30 to 80 mg DHA per 100 kcal based on the total energy content of the nutritional composition. The above numbers are based on the molar weight of DHA.

[0153] EPA is preferably administered in an amount of 40 to 350 mg, more preferably 50 to 300 mg, most preferably 60 to 250 mg EPA per day. In one embodiment when the nutritional composition is in the form of an oral nutritional supplement the daily amount of EPA administered is preferably 40 to 220 mg, more preferably 50 to 200 mg per day, most preferably 60 to 180 mg per day.

[0154] Based on calories, preferably the nutritional composition according to the invention comprises 5 to 30 mg, more preferably 6 to 25 mg, most preferably 8 to 20 mg EPA per 100 kcal based on the total energy content of the nutritional composition. The above numbers are based on the molar weight of EPA. If EPA is present in the nutritional composition, the composition preferably comprises more DHA than EPA. i.e., the weight ratio of DHA to EPA is preferably more than 1 , more preferably 1.5:1 to 4:1 even more preferably 2:1 to 4:1.

[0155] In terms of daily dosage, the present use and method preferably comprises the administration of preferably comprises 200 to 2000 mg omega-3 fatty acids (preferably DHA+EPA+DPA, most preferably DHA+EPA) per day, more preferably 230 to 1750 mg per day, even more preferably 250 to 1600 mg EPA per day.

[0156] The present nutritional composition preferably comprises 0.3 to 3 wt% omega-3 fatty acids (more preferably DHA+EPA+DPA, most preferably DHA+EPA) on the total fatty acids, more preferably 0.5 to 2.5 wt%, even more preferably 0.75 to 2.0 wt% omega-3 fatty acids based on total fatty acids. The present nutritional composition preferably comprises 0.1 to 3.5 wt% DHA based on total fatty acids, preferably 0.3 to 3 wt% DHA based on total fatty acids, more preferably 0.5 to 2.5 wt% DHA based on total fatty acids. The present nutritional composition preferably comprises between 0.05 to 0.30 wt% EPA based on total fatty acids, more preferably 0.1 to 0.25 wt%, even more preferably 0.15 to 0.20 wt% EPA based on total fatty acids.

[0157] The above-mentioned ratios and amounts take into account and optimize several aspects, including taste (too high n-3 LC-PUFA levels reduce taste, resulting in a reduced compliance), balance between DHA and precursors thereof to ensure optimal effectiveness while maintaining low-volume formulations.

[0158] Vitamin B2 and further B vitamins

[0159] The nutritional composition may comprise vitamin B2. In some embodiments, the nutritional composition may comprise further B vitamins, preferably the composition comprises at least two B vitamins, wherein one B vitamin is B2 and the second B vitamin is selected from the group of vitamin B1 (thiamine), B3 (niacin or niacinamide), vitamin B5 (pantothenic acid), B6 (pyridoxine, pyridoxal, or pyridoxamine, or pyridoxine hydrochloride), vitamin B7 (biotin) vitamin B9 (folic acid or folate), and vitamin B12 (cobalamins). Functional equivalents are encompassed within these terms. The term “vitamin B12” incorporates all cobalamin equivalents known in the art.

[0160] B-vitamins are known as nutritional co-factors that act as biochemical “spark plugs” in mitochondria, and therefore act to further nurture and support optimal neural metabolism. In an embodiment the nutritional composition comprises DHA, vitamin D and vitamin B2. In a further embodiment the nutritional composition comprises DHA, vitamin D, vitamin B2 and optionally together with vitamin B6 and / or vitamin B9.

[0161] Preferably, B vitamin in the context of the nutritional composition according to the invention comprises at least three B vitamins, wherein the first B vitamin is B2 and the second and third B vitamin are selected from the group of vitamin B1 , B3, B5, B6, B7, vitamin B12 and vitamin B9. Preferably the nutritional composition comprises vitamin B2, B6 and B9, more preferably vitamin B2, B6, B9 and B12, even more preferably vitamin B1 , vitamin B2, vitamin B6, vitamin B9 and vitamin B12. In one embodiment the nutritional composition according to the invention may include all of vitamin B1 (thiamine), B2 (riboflavin), B6 (pyridoxine, pyridoxal, or pyridoxamine, or pyridoxine hydrochloride), vitamin B9 (folic acid or folate), and vitamin B12 (cyanocobalamin). In some embodiment the nutritional composition may comprise further B vitamins, further B vitamins as used herein are vitamin B1 , B3, B5 and B7 which the composition may comprise in addition to vitamin B2 and one or more of vitamin B6 and vitamin B9.

[0162] If present in the nutritional composition, the vitamin B1 is preferably present in an amount to provide a daily dosage in the range of 0.7 to 5 mg, preferably in the range of 0.8 to 4.9 mg, more preferably in the range of 0.9 to 4.8 mg.

[0163] In a preferred embodiment, the vitamin B1 is preferably present in a therapeutic effective amount if the nutritional supplement or composition provides at least 1.2, preferably at least 1.3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1 .3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. The RDI is a common reference in the art, for example herewith is referred to the population reference intake (PRI) such as according to the Summary of dietary reference values for the Ell population as derived by the European Food Safety Authority as determined in September 2017. In the Ell the RDI based on the ESFA for vitamin B1 is 0.1 mg per 238.83 calories intake.

[0164] In the nutritional composition according to the invention, the vitamin B2 is preferably present in an amount to provide a daily dosage in the range of 0.8 to 7.6 mg, preferably in the range of 1 to 7.4 mg, more preferably in the range of 1.2 to 7.2 mg per day, most preferably in the range of 1 .4 to 7.2 mg per day.

[0165] In a further preferred embodiment, vitamin B2 is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1 .3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. The RDI is a common reference in the art, for example herewith is referred to the population reference intake (PRI) such as according to the Summary of dietary reference values for the Ell population as derived by the European Food Safety Authority as determined in September 2017. In the Ell the RDI based on the ESFA for vitamin B2 is 1.6 mg per day.

[0166] If present in the nutritional composition, the vitamin B3 is preferably present in an amount to provide a daily dosage in the range of 3 to 45 mg, preferably in the range of 3.5 to 42 mg, more preferably in the range of 3.8 to 40 mg niacin equivalent per day.

[0167] If present in the nutritional composition, the vitamin B5 is preferably present in an amount to provide a daily dosage in the range of 0.8 to 15 mg, preferably in the range of 1 to 14 mg, more preferably in the range of 1.2 to 13 mg per day.

[0168] If present in the nutritional composition, the vitamin B6 is preferably present in an amount to provide a daily dosage in the range of 0.7 to 9 mg, preferably in the range of 0.8 to 8 mg, more preferably in the range of 0.9 to 7.5 mg.

[0169] In a preferred embodiment, the vitamin B6 is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for vitamin B6 is 1.65 mg per day.

[0170] If present in the nutritional composition, the vitamin B7 is preferably present in an amount to provide a daily dosage in the range of 8 to 95 pg, preferably in the range of 9 to 90 pg, more preferably in the range of 9.5 to 85 pg per day.

[0171] Vitamin B9 or folic acid is an essential nutrient that is converted by the body into folate, which is required for DNA and RNA biosynthesis, DNA repair and one-carbon metabolic reactions. If present in the nutritional composition, the vitamin B9 is preferably present in an amount to provide a daily dosage in the range of 150 to 900 pg, preferably in the range of 175 to 800 pg, more preferably in the range of 200 to 750 pg per day.

[0172] In a preferred embodiment, the vitamin B9 is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1.3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for vitamin B9 is 330 pg dietary Folate Equivalents per day.

[0173] If present in the nutritional composition, the vitamin B12 is preferably present in an amount to provide a daily dosage in the range of 2 to 20 pg, preferably in the range of 2.5 to 19 pg, more preferably in the range of 3 to 18 pg per day.

[0174] In a preferred embodiment, the vitamin B12 is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably

[0175] 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for vitamin B12 is 4 pg per day.

[0176] Vitamin D3

[0177] The present nutritional composition may comprise vitamin D3. Vitamin D is a group of fatsoluble secosteroids, the two major physiologically relevant forms of which are vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). These are known collectively as calciferol. Vitamin D without a subscript refers to all forms of vitamin D, either D1 , D2, D3, or D4, in particular D2 and D3, or any mixture thereof. Vitamin D may be provided in an active (1 ,25(OH)2D) or unactive (vitamin D3 or D2) form. The nutritional composition according to the invention preferably comprises vitamin D3.

[0178] According to one embodiment, vitamin D3 is supplemented in an amount of 5 to 80 pg per day, preferably 6 to 75 pg per day, more preferably 8 to 70 pg per day, even more preferably 10 to 65 pg per day. Based on calories, preferably the nutritional composition according to the invention comprises 1.3 to 4.5 pg vitamin D3, per 100 kcal, preferably 1.5 to 4 pg vitamin D3 per 100 kcal, more preferably 2 to 3.5 pg per 100 kcal based on the total energy content of the nutritional composition. The above numbers are based on the molar weight of vitamin D3.

[0179] In a further preferred embodiment, vitamin D3 is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1.3 times the recommended daily intake (RDI), and preferably not more than 7 times the RDI. In a preferred aspect from 1 .2 to 5 times, preferably 1 .3 to 4.5 times, more preferably

[0180] 1.4 to 4 times the recommended daily intake is provided. The RDI is a common reference in the art, for example herewith is referred to the population reference intake (PRI) such as according to the Summary of dietary reference values for the Ell population as derived by the European safety Authority as determined in September 2017. In embodiments, the RDI of vitamin D3 is 15 pg per day. Choline

[0181] The present nutritional composition preferably further comprises choline. Choline may be present as such, or in the form of a choline salt and / or choline ester. The choline salt is preferably selected from choline chloride, choline bitartrate, choline butyrate or choline stearate. The choline ester is preferably selected from a phosphatidylcholine and lyso- phosphatidyl choline, preferably phosphatidylcholine. In a preferred embodiment the present nutritional composition comprises both a) choline as such or a choline salt and b) a choline ester, preferably phosphatidylcholine.

[0182] The present nutritional composition preferably comprises the administration of more than 70 mg of a) choline or choline salt per day, preferably 70 to 1300 mg per day, more preferably 80 to 1200 mg per day, most preferably 100 to 1100 mg per day. Based on calories, preferably the nutritional composition according to the invention comprises 25 to 110 mg choline per 100 kcal, preferably 30 to 105 mg choline per 100 kcal, more preferably 33 to 100 mg per 100 kcal based on the total energy content of the nutritional composition.

[0183] The above numbers are based on choline, the amounts of choline salts or esters can be calculated taking the molar equivalent to choline into account.

[0184] The present nutritional composition may comprise choline in the form of a choline ester, preferably provided in the form of phospholipids. Preferably, one or more phospholipid(s) is / are present in the nutritional composition according to the invention. The one or more phospholipid(s) is / are selected from the group consisting of phosphatidic acid (PA), phosphatidylethanolamine (PE), phosphatidylcholine (PC), phosphatidylserine (PS) and phosphoinositides (PI), preferably at least phosphatidyl choline. Phospholipids are in a preferred embodiment a source of the choline ester phosphatidylcholine.

[0185] Suitable sources of phospholipids herein are soya lecithin, egg lecithin as well as rapeseed and sunflower lecithin. Milk contains phospholipids in the milk fat globule membrane (MFGM), suitable sources include milk phospholipid concentrate such as those commercially available from Lecico GmbH, whey protein concentrate Lacprodan® MFGM from Aria Foods. Depending on the source, phospholipids comprise about 10 to 40 wt% of phosphatidylcholine.

[0186] If present, the nutritional composition and use or method of use of the nutritional composition preferably comprises the administration of phospholipids in an amount of 0.6 to 6.6 g mg per day, preferably 0.8 to 6.4 g of phospholipids per day, more preferably 0.9 to 6.2 g of phospholipids per day, most preferably 1 to 6 g of phospholipids per day. In an aspect the amount of phospholipids is about 200 mg per day. The phospholipids preferably at least provide phosphatidylcholine. In one embodiment when the nutritional composition is in the form of a oral nutritional supplement the daily amount of phospholipids administered is preferably 0.5 to 4 g per day, more preferably 0.75 to 3.5 g per day, most preferably 1 to 3 g per day. Based on calories, preferably the nutritional composition according to the invention comprises 100 to 500 mg of phospholipids per 100 kcal, preferably 150 to 400 mg of phospholipids per 100 kcal, more preferably 200 to 350 mg of phospholipids per 100 kcal based on the total energy content of the nutritional composition.

[0187] In a specific embodiment, the nutritional composition according to the invention, comprises a) choline or choline salts and b) phosphatidylcholine (PC). In such embodiment the weight ratio phosphatidylcholine to choline is usually more than 0.1 , preferably more than 0.26, in particular 0.30-6, more preferably 0.36-3. Most preferably, the weight ratio phosphatidylcholine to choline is in the range of 40:60 to 60:40. Herein, the amount of choline in grams is to be calculated as the molar contribution of choline as provided by all choline sources (when orally digested and assuming 100% bioavailability, including PC), times the molecular weight of choline (104 g / mol). Herein, the molecular weight of PC is 810 gram / mol. So for example, including 400 mg choline chloride (having a molecular weight of 139.6g / mol) and 200 mg phosphatidylcholine and 200 mg phospholipids other than PC would then result in a weight ratio of PC to choline of 200 / [(104 / 139.6)x400 + (104 / 810)x200] = 200 / [298 + 25.7] = 0.62.

[0188] Preferably the nutritional composition according to the invention comprises choline for at least 20 mole%, preferably at least 30 mole%, more preferably at least 40 mole% provided in the form of phosphatidylcholine, preferably the molar ratio choline (salts) and PC is in the range from 40:60 to 60:40, i.e. wherein 40 to 60 mole% choline is derived from phosphatidylcholine based on total choline in the composition. Preferably, the nutritional composition comprises a combination of omega-3 fatty acids DHA and EPA, vitamin B2, vitamin D3, choline and the synbiotic composition according to the invention, wherein choline is for at least 40 mole% provided in the form of phosphatidylcholine, preferably the molar ratio choline and PC is in the range from 40:60 to 60:40 and further comprises one of more selected from A, further B, C, and E vitamin(s), phospholipids, selenium and magnesium. More preferably the composition contains two or more of these further ingredients. The further ingredients and their respective amounts will be discussed hereinafter.

[0189] Vitamin A

[0190] The present nutritional composition may further comprise vitamin A. Vitamin A is a fat-soluble vitamin which comes in several forms (as retinol, retinal, retinoic acid or retinyl ester). Any functional form of vitamin A known in the art is suitable to be used herein, including retinol (in particular retinol esters), retinal, retinoic acid, beta-carotene, provitamin A, or any combination thereof. Preferably, the nutritional composition comprises retinol, in particular retinyl acetate and / or retinyl palmitate.

[0191] Vitamin A intakes or requirements are generally expressed in terms of retinol equivalents (RE). One RE is defined as the biological activity associated with 1 pg of all-trans retinol. Hence 1 pg Retinol Equivalent is similar to 1 pg of all-trans retinol. 1 International Unit (IU) retinol corresponds to 0.3 pg Retinol Equivalents.

[0192] If present in the nutritional composition, vitamin A is preferably administered in an amount of 300 to 2800 pg Retinol Equivalents per day, more preferably 350 to 2700 pg Retinol Equivalents per day, even more preferably 650 to 900 pg Retinol Equivalents per day, most preferably 375 to 2600 pg Retinol Equivalents per day. In a preferred embodiment, the vitamin A is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1.2, preferably at least 1.3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the EU the RDI based on the ESFA for vitamin A is 700 pg Retinol Equivalents per day.

[0193] Vitamin C

[0194] The present nutritional composition may further comprise vitamin C. Vitamin C is an antioxidant that may provide beneficial characteristics. Vitamin C includes functional equivalents thereof including sodium ascorbate and calcium ascorbate, and may be present in an amount to provide a daily dosage in the range of 75 to 700 mg, preferably in the range of 85 to 600 mg, more preferably in the range of 95 to 550 mg per day.

[0195] When the nutritional composition is in the form of an oral nutritional supplement the daily amount of vitamin C administered is preferably 80 to 320 mg per day, more preferably 90 to 300 mg per day, most preferably 95 to 290 mg per day. Based on calories, preferably the nutritional composition according to the invention may comprise 12 to 40 mg vitamin C per 100 kcal, preferably 14 to 35 mg vitamin C per 100 kcal, more preferably 16 to 32 mg vitamin C per 100 kcal based on the total energy content of the composition. The above numbers are based on the molar weight of ascorbic acid.

[0196] In a preferred embodiment, the vitamin C is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1.3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for vitamin C is 102.5 mg per day.

[0197] Vitamin E

[0198] The nutritional composition according to the invention may comprise vitamin E. Vitamin E refers to compounds having vitamin E activity as known in the art, typically tocopherol and / or an equivalent thereof.

[0199] If included, vitamin E may be present in the nutritional composition in an amount to provide a daily dosage in the range of 8 to 75 mg, preferably in the range of 10 to 70 mg, more preferably in the range of 12.5 to 70 mg alpha-TE.

[0200] When the nutritional composition is in the form of an oral nutritional supplement the daily amount of vitamin E administered is preferably 8 to 42 mg per day, more preferably 10 to 40 mg per day, most preferably 12.5 to 37.5 mg alpha-TE per day. Such amounts of vitamin E prevent oxidative damage.

[0201] Based on calories, preferably the nutritional composition according to the invention may comprise 1 to 5.5 mg alpha-TE per 100 kcal, preferably 1.5 to 5 mg alpha-TE per 100 kcal, more preferably 2 to 4.5 mg alpha-TE per 100 kcal based on the total energy content of the nutritional composition.

[0202] In a preferred embodiment, the vitamin E is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for vitamin E is 12 mg per day.

[0203] The term “tocopherol and / or an equivalent thereof”, as used in this description, comprises tocopherols (e.g. alpha- and gamma-), tocotrienols, pharmaceutical and / or nutritional acceptable derivatives thereof and any combination thereof. The above numbers are based on alpha-tocopherol equivalents (alpha-TE) and the molecular weight thereof, as recognized in the art.

[0204] Selenium

[0205] The present nutritional composition may comprise selenium. The antioxidant activity of selenium may advantageously prevent and / or inhibit damages to intestines. If present in the nutritional composition, selenium is preferably present in an amount to provide a daily dosage in the range of 30 to 275 pg, preferably in the range of 35 to 250 pg, more preferably in the range of 38 to 225 pg per day. When the nutritional composition is in the form of an oral nutritional supplement the daily amount of selenium administered is preferably 25 to 140 pg per day, more preferably 30 to 130 pg per day, most preferably 35 to 120 pg per day. Based on calories, preferably the nutritional composition according to the invention may comprise 4 to 18 pg selenium per 100 kcal, preferably 6 to 16 pg selenium per 100 kcal, more preferably 8 to 14 pg selenium per 100 kcal based on the total energy content of the nutritional composition.

[0206] In a preferred embodiment, the selenium is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for selenium is 70 pg per day.

[0207] Magnesium

[0208] The present nutritional composition may comprise magnesium. If present in the nutritional composition, magnesium is preferably present in an amount to provide a daily dosage in the range of 125 to 850 mg, preferably in the range of 150 to 800 mg, more preferably in the range of 170 to 750 mg per day.

[0209] When the nutritional composition is in the form of an oral nutritional supplement the daily amount of magnesium administered is preferably 150 to 600 mg per day, more preferably 160 to 550 mg per day, most preferably 170 to 525 mg per day. Based on calories, preferably the nutritional composition according to the invention may comprise 10 to 70 mg magnesium per 100 kcal, preferably 15 to 65 magnesium per 100 kcal, more preferably 20 to 60 mg magnesium per 100 kcal based on the total energy content of the nutritional composition.

[0210] In a preferred embodiment, the magnesium is preferably present in a therapeutic effective amount if the nutritional supplement or nutritional composition provides at least 1 .2, preferably at least 1 .3 times the recommended daily intake (RDI) and preferably not more than 20 times, preferably not more than 10 times the RDI. In a preferred aspect from 1.2 to 5 times, preferably 1.3 to 4.5 times, more preferably 1.4 to 4 times the recommended daily intake is provided. In the Ell the RDI based on the ESFA for magnesium is 325 mg per day.

[0211] The nutritional composition may comprise further minerals including but not limited to sodium, potassium, chloride, calcium, phosphor, iron, zinc, cupper, manganese, fluor, molybdenum, chrome and iodium. Amount thereof comply with general recommendations for nutritional requirements of the FSMP.

[0212] LA / ALA

[0213] The lipid fraction in the nutritional composition may in addition to the omega-3 fatty acid(s) further comprise lipids that are a source of essential fatty acids alpha-linolenic acid (ALA, C- 18:3) and linoleic acid (LA, C-18:2). LA and / or ALA may be provided as free fatty acids, in triglyceride form, in diglyceride form, in monoglyceride form, in phospholipid form, or as a mixture of one of more of the above. Preferably the present nutritional composition contains at least one, preferably at least two lipid sources selected from the group consisting of soybean oil, rape seed oil (such as colza oil, low erucic acid rape seed oil and canola oil), high oleic sunflower oil, high oleic safflower oil and olive oil. The LA : ALA weight ratio is preferably in the range of 8:1 to 3:1 , preferably 7:1 to 4:1 , more preferably 6:1 to 5:1.

[0214] Based on calories, preferably the nutritional composition according to the invention may comprise 0.1 to 1.5 g LA per 100 kcal, preferably 0.2 to 1.1 g LA per 100 kcal, more preferably 0.25 to 1 g LA per 100 kcal based on the total energy content of the composition. Based on calories, preferably the composition according to the invention may comprise 0.03 to 0.08 g ALA per 100 kcal, preferably 0.04 to 0.07 g ALA per 100 kcal, more preferably 0.05 to 0.065 g ALA per 100 kcal based on the total energy content of the composition.

[0215] In a particular embodiment there is provided a nutritional composition comprising

[0216] (i) the synbiotic mixture according to the invention

[0217] (ii) DHA and EPA;

[0218] (iii) vitamin D3;

[0219] (iv) vitamin B2;

[0220] (vi) choline, choline salts and / or choline ester, preferably at least phosphatidylcholine;

[0221] (vi) vitamin B6;

[0222] (vii) vitamin B9; and at least one or more of the following ingredients

[0223] (viii) Retinol Equivalents of vitamin A;

[0224] (ix) vitamin B1 ;

[0225] (x) vitamin B12;

[0226] (xi) vitamin C;

[0227] (xii) alpha-TE;

[0228] (xiii) selenium; and,

[0229] (xiv) magnesium Application

[0230] The synbiotic composition and / or nutritional composition according to the invention relate to the use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic-induced gut microbial dysbiosis.

[0231] In another aspect of the invention, the synbiotic composition and / or nutritional composition according to the invention relate to the use in preventing and / or treating intestinal barrier disruption in a subject.

[0232] The term “treatment” as used herein encompasses:

[0233] (a) inhibiting the disease, e.g. halting or slowing down its progression (includes maintenance therapy);

[0234] (b) relieving the disease, e.g. causing regression thereof; and

[0235] (c) prolonging survival as compared to expected survival if not receiving treatment.

[0236] As the skilled person will appreciate, antibiotics are used against infectious pathogens but also as in prophylactic uses when no infection is installed in the host. Accordingly, “antibiotictherapy” as used herein refers to the prophylactic or therapeutic use of antibiotics in a subjects in need thereof. Preferably, the adult subject is undergoing or has been submitted to antibiotic therapy, more preferably the subject is undergoing antibiotic therapy.

[0237] In some embodiments, the subject is or has been submitted to antibiotic therapy for at least 4 days, preferably at least 7 days, more preferably at least 10 days.

[0238] Preferably, the synbiotic composition and / or the nutritional composition of the invention is for use after interruption of antibiotic therapy, preferably at least 2 days after interruption of antibiotic therapy, more preferably at least 5 days after interruption of antibiotic therapy. Alternatively, gut microbial dysbiosis can persist up to months after antibiotic treatment, thus in a further preferred embodiment, the synbiotic composition and / or the nutritional composition of the invention is for use after interruption or after finishing the antibiotic treatment, preferably up to six months after interruption or after finishing the antibiotic therapy, more preferably up to three months after interruption or after finishing the antibiotic therapy

[0239] In some embodiments, antibiotic-induced gut microbial dysbiosis may be defined as the remodelling of the gut microbiome towards a higher relative abundance of strict anaerobes and lower relative abundance of aerotolerant microbes. Accordingly, in one embodiment, the composition of the invention is for use in antibiotic-induced gut microbial dysbiosis wherein the use comprises increasing the relative abundance of strictly anaerobic gut microorganisms and / or to reduce the relative abundance of aerotolerant gut microorganisms. Herein, the terms increasing and reducing are in comparison with the relative abundance before initiating the intervention with the synbiotic composition of the invention. Examples of strictly anaerobic gut microorganisms are from bacterial families like Oscillospiraceae, Lachnospiraceae, Ruminococcaceae and Bacteroidaceae.. Examples of aerotolerant microorganisms are Enterobacteriaceae and Enterococcaceae.

[0240] Remodelling of the gut microbiome and relative abundance of anaerobic and aerotolerant microorganisms can be measured using the Metagenomic Aerotolerant Predominance Index (MAPI) as described by Million & Raoult, 2018 (Million & Raoult . “Linking gut redox to human microbiome”. Human Microbiome Journal, v.10, Dec 2018, pages 27-32, https: / / doi.Org / 10.1016 / j.humic.2018.07.002).

[0241] The intestinal (gut) barrier is a critical interface of the human body - on the one hand it provides for a strong defence against foreign substances while on the other hand it regulates the absorption of essential nutrients, water and electrolytes. The intestinal barrier is composed of multiple layers. The outer one comprises the mucus layer, the commensal gut microbiota and defence proteins such as antimicrobial proteins (AMPs) and secretory immunoglobulin A (slgA). Intestinal epithelial cells (lECs) are the middle layer, while the inner part is composed of e.g. immune cells of innate and adaptive immunity.

[0242] The term “intestinal barrier disruption” as used herein refers to any damage or wounding of the intestinal barrier that interferes with proper functioning of the intestinal barrier and results in reduced intestinal barrier integrity.

[0243] The term “preventing intestinal barrier disruption” as used herein refers to promoting intestinal conditions wherein the intestinal barrier is in optimal condition and thereby less vulnerable to damage or wounding.

[0244] The term “treating intestinal barrier disruption” as used herein refers to promoting intestinal conditions where the intestinal barrier can recover faster after being exposed to damage or wounding.

[0245] Experiments showed that the synbiotic composition and / or nutritional composition comprising the same according to the invention has a beneficial effect on the production of microbial metabolites which upon addition to epithelial tubules lead to increased TEER values, implying an improvement of the intestinal barrier.

[0246] In a preferred embodiment, the invention relates to the use of the synbiotic composition and / or nutritional composition comprising the synbiotic composition of the invention for use in preventing and / or treating intestinal barrier disruption in a subject, wherein the composition comprises:

[0247] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0248] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0249] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260

[0250] In a preferred embodiment, the invention relates to the use of the synbiotic composition and / or nutritional composition comprising the synbiotic composition of the invention for use in improving gut barrier function in a subject.

[0251] In a preferred embodiment, the invention may also be worded as a method for preventing and / or treating intestinal barrier disruption in a human subject, wherein the method comprises administration of a synbiotic composition and / or nutritional composition comprising the synbiotic composition to said subject, wherein said synbiotic composition and / or nutritional composition comprises:

[0252] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0253] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0254] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0255] In a preferred embodiment, the invention may also be worded as a method for improving gut barrier function in a subject, wherein the method comprises the administration of the synbiotic composition and / or nutritional composition comprising the synbiotic composition according to the invention to said subject.

[0256] In another preferred embodiment, the invention may also be worded as the use of Bifidobacterium ssp., non-digestible oligosaccharides, and pectin polysaccharides in the manufacture of a synbiotic composition and / or nutritional composition for preventing and / or treating intestinal barrier disruption in a human subject, wherein the synbiotic composition and / or nutritional composition comprising the synbiotic composition comprises:

[0257] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0258] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0259] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0260] In another preferred embodiment, the invention may also be worded as the use of Bifidobacterium ssp., non-digestible oligosaccharides, and pectin polysaccharides in the manufacture of a synbiotic composition and / or nutritional composition for improving gut barrier function in a subject, wherein the synbiotic composition and / or nutritional composition is according to the invention.

[0261] In yet another preferred embodiment, the invention may also be worded as the use of a synbiotic composition and / or nutritional composition comprising the synbiotic composition for preventing and / or treating intestinal barrier disruption in a human subject, wherein the use comprises administration of a synbiotic composition or nutritional composition to said subject, wherein the synbiotic composition and / or nutritional composition comprising the synbiotic composition comprises:

[0262] • 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;

[0263] • at least 5wt.% on dry basis of non-digestible oligosaccharides selected from galactooligosaccharides and / or fructooligosaccharides; and

[0264] • 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

[0265] In an embodiment, the subject is human, more preferably, the human subject is at risk of intestinal barrier disruption. Intestinal barrier disruption may for example be caused by inflammatory bowel disease (IBD), celiac disease, irritable bowel syndrome (IBS), infections and / or autoimmune diseases. In an embodiment, the human subject at risk of intestinal barrier disruption suffers from inflammatory bowel disease (IBD), celiac disease, irritable bowel syndrome (IBS), infections and / or autoimmune diseases.

[0266] The invention is further illustrated by the following non-limiting examples.

[0267] EXAMPLES

[0268] Example 1

[0269] A synbiotic composition according to the invention was prepared by dry blending commercially available pectin polysaccharides (LvPectin) with GOS and FOS in a weight ratio of 2:9:1. The pectin polysaccharide used for the analysis was Herbapekt SF 50-LV from Herbafood Ingredients GmbH, Germany. The molecular weight of the pectin polysaccharide is about 100 kDa and it has a DP of about 500. The pectin polysaccharides are rich in 1 ,4-linked a-D- galacturonic acid residues.

[0270] The monosaccharide composition of pectin polysaccharides was analysed by methanolysis and is provided in Table 1. The degree of methyl esterification was 59.2% and acetyl content was DA=1 .2%, on basis of a moles of ester per 100 mol of galacturonic acid for pectin.

[0271] Table 1 This profile confirms that the pectin of the invention is rich in HG-I, particularly in view of the low DM and DA and very high GalA content.

[0272] Example 2

[0273] Fecal samples of adult volunteers in recovery following antibiotic therapy have been collected. The two key selection criteria of the donors for the current study were; age >50 years (preferably > 60) and recent intake of amoxicillin in combination with clavulanic acid (sample donation during first week after cessation of the antibiotic treatment). The average age of the test subjects was 63 and the fecal samples were collected between 1 to 7 days after cessation of antibiotic treatment.

[0274] Gut microbiome of the subjects enrolled in this study was analyzed using a model as described in Van den Abbeele et al. (Bridging preclinical and clinical gut microbiota research using the ex vivo SIFR® technology, Front. Microbiol.,, 2023, 14, p.1 -11) and Systemic Intestinal Fermentation Research (SIFR®) technology. SIFR® technology involves the use of sets of small individual bioreactors that are handled in a custom-built processing device allowing for temperature control as well as headspace control and monitoring. This technology minimises the bias in microbiota composition between in vivo derived microbiota and the one colonizing the bioreactors.

[0275] Remodelling of the gut microbiome and relative abundance of anaerobic and aerotolerant microorganisms was measured using the Metagenomic Aerotolerant Predominance Index (MAPI) as described by Million & Raoult (Linking gut redox to human microbiome. Human microbiome journal, v.10, Dec 2018, pages 27-32, https: / / doi.Org / 10.1016 / j.humic.2018.07.002). According to MAPI, the value above 0 (zero) indicates a higher abundance of aerotolerant microbes and below zero, higher relative abundance of strict anaerobes.

[0276] Prebiotics, probiotics and synbiotic mixtures were compared. The test products were inulin (Raftiline®HP (Orafti)) and a fibre blend comprising short chain GOS (Vivinal®GOS from FrieslandCampina Domo, Amersfoort, The Netherlands), long chain FOS (oligofructose, Orafti®) and pectin polysaccharide as described under example 1 in a ratio of 9:1 :2 and the probiotic Bifidobacterium longum, BL999 (BB536 from Morinaga). The test fibre products were tested individually and combined with the probiotic and compared to the inoculum (control). Fibre products were tested at a dose equivalent of 5 grams per day and the probiotic was dosed at a final concentration of 2 x 107cfu / mL. An ex vivo SIFR® study was performed, simulating the colonic fermentation carried out for 48 hours of test products by the gut microbiota samples derived from dysbiotic human adults . SIFR technology has been validated by studying the impact of carbohydrates for which clinical data is available. Microbial composition analysis was carried out by quantitative 16S rRNA gene profiling.

[0277] The effect in ameliorating the dysbiotic effect is shown in Figure 1 , and summarized in Table 2 below.

[0278] Table 2 - Median MAPI (Metagenomic Aerotolerant Predominance Index) values

[0279] These results demonstrate the ability of synbiotic composition of the invention (BB536-G) to boost the relative abundance of strict anaerobes compared to not only the synbiotic mixture BB536+INUL, but also to the probiotics and prebiotics (GFLvp ; INIIL) alone.

[0280] Example 3

[0281] To study the effect of the composition on antibiotic induced gut microbiome dysbiosis, the gut microbiome composition is analyzed by metagenomic sequencing and short chain fatty acid profile. To study the effect of the composition according to the invention in respect to galacturonic acid oligosaccharides, a composition comprising scGOS, IcFOS and galacturonic acid oligosaccharides (AOS) in a weight ratio of 9:1 :2 was taken along in the experiments. AOS are produced by hydrolyzing pectin and thus contain a similar percentages of monosaccharide units as pectin, however in are present in either oligomers or monomers instead of polymers. The DP of AOS is in general lower than 20.

[0282] Samples

[0283] To induce dysbiosis, fecal samples were preincubated with Amoxicillin trihydrate: potassium clavulanate (4: 1 ):final concentration 500ug / ml in 5x diluted fecal slurry has been done with 12 hours incubation time. After dysbiosis, fecal samples were fermented with five different compositions:

[0284] 1. scGOS, IcFOS, LvPectin in a 9:1 :2 ratio;

[0285] 2. scGOS, IcFOS, AOS in a 9:1 :2 ratio.

[0286] 3. scGOS, IcFOS, LvPectin in a 9:1 :2 ratio and BB356;

[0287] 4. scGOS, IcFOS, AOS in a 9:1 :2 ratio and BB536; and

[0288] 5. BB536 alone (only measured in SOFA determination).

[0289] The components used in the compositions were a fibre blend comprising short chain GOS (VivinalOGOS from FrieslandCampina Domo, Amersfoort, The Netherlands), long chain FOS (oligofructose, Orafti®) and pectin polysaccharide (LvPectin) as described under example 1 (Herbapekt SF 50-LV from Herbafood Ingredients GmbH, Germany, MW~100 kDa.) in a ratio of 9:1 :2 and the probiotic Bifidobacterium longum, BL999 (BB536 from Morinaga). AOS are produced from pectin (Sudzucker AG, Mannheim, Germany) with a DP of 1 - 20 and comprises of approximately 75% galacturonic acid oligomers based on total weight. An oligomer has around 2 - 20 monomeric units.

[0290] The control in all experiments is a blanc fermentation sample.

[0291] Material and methods

[0292] Fecal samples were collected after 72 hours of fermentation for metagenomic analysis and supernatant of fermentation was used for SOFA analysis. Briefly, 6 ml of the faeces / substrate suspension was put in a dialysis tube and air was removed in the empty space. The dialysis tube was put in a 100 ml Scott bottle filled with 100 ml dialysis medium. The Scott bottles were closed and incubated at 37°C. Samples of the dialysis medium (dialysate) and faecal suspension (lumen) were taken after 72 hours for determination of short chain fatty acids (SOFA).

[0293] Fermentation medium (Me Bain and MacFarlane) contains buffered peptone water 3,0 g / l, Yeast Extract 2,5 g / l, Tryptone 3,0 g / l, L-Cysteine-HCI 0,4 g / l, Bile salts 0,05 g / l, K2HPO4.3H2O 2,6 g / l, NaHCO30,2 g / l, NaCI 4,5 g / l, MgSO4.7H2O 0,5 g / l, CaCI2. 2H2O 0,3 g / l, FeSO4.7H2O 0,005 g / l. Ingredients were added one by one in 800 ml water, pH was adjusted to 6.3±0.1 with K2HPO4or NaHCO3and volume was filled up to 1 litre. Medium was sterilized for 15 minutes at 121 °C and put in the anaerobic cabinet at least 16 hours before use.

[0294] Dialysis medium contains K2HPO4.3H2O 2,6 g / l, NaHCO30,2 g / l, NaCI 4,5 g / l, MgSO4.7H2O 0,5 g / l, CaCh.2H2O 0,3 g / l, FeSO4.7H2O 0,005 g / l. pH was adjusted to 6.3 ±0.1 with K2HPO4 or NaHCOs. Medium was not sterilized because of forming of sediment. The medium was put in the anaerobic cabinet at least 16 hours before use.

[0295] The relative abundance of Bifidobacteriaceae family was measured by shotgun metagenomics. Briefly, samples were subject to DNA extraction using a commercial DNA isolation kit followed by shot gun metagenome sequencing using Illumina sequencing. The sequencing data was quality checked to remove low quality sequences and taxonomic profiling was done using metaphlan 4. Data was aggregated at microbial family level and compared between groups.

[0296] The SCFA acetic acid and butyric acid values were quantitatively determined after 72 hours using a Shimadzu- GC2025 gas chromatograph with a flame ionization detector. Hydrogen was used as mobile phase. The levels of SCFA were determined using 2-ethylbutyric acid as an internal standard. 2-Ethylbutyric acid is used as internal standard because it is chemically synthesized and is not present in the original fecal samples. From the peak area a calibration curve was constructed and the concentration in the samples was calculated.

[0297] Results

[0298] The relative abundance (%) of the Bifidobacteriaceae family was measured after 72 hours of fermentation. The results are shown in Figure 2 and Table 3. The combination of scGOS, IcFOS, LvPectin and BB536 shows an improvement in abundance of Bifidobacteriaceae compared to the composition without BB536. More importantly, the addition of BB536 to the composition comprising scGOS, IcFOS and LvPectin provides a higher increase in Bifidobacteriaceae abundance compared to the addition of BB536 to the composition comprising AOS.

[0299] The addition of BB536 to the composition comprising scGOS, IcFOS and LvPectin led to an increase of 19%, whereas the addition of BB536 to the composition comprising AOS instead of LvPectin only led to a 3% increase.

[0300] Table 3. Mean (S.D.) relative abundance of Bifidobacteriaceae family at 72 hours in the tested treatments.

[0301] The combination of scGOS, IcFOS, LvPectin and BB536 resulted in an increase in both acetic acid and butyric acid production compared to the composition without BB536. The increase in both acetic acid and butyric acid was higher than the theoretical value. The theoretical value 5 is the additive value of the measurements of scGOS, IcFOS, LvPectin and BB536. Hence, the composition of the invention synergistically increases both acetic acid and butyric acid production.

[0302] The effect on acetic acid and butyric acid production is shown in Figures 3A and 3B and T ables 0 4 and 5.

[0303] Table 4. Effect of scGOS, IcFOS, LvPectin and BB536 on acetic acid production after 72 hours. 5 Table 5. Effect of scGOS, IcFOS, LvPectin and BB536 on butyric acid production after 72 hours.

[0304] Conclusion

[0305] These results demonstrate the ability of the synbiotic composition / nutritional composition of 0 the invention to synergistically increase the relative abundance of Bifidobacteriaceae. More importantly, this synergistic effect on Bifidobacteriaceae was only observed in the composition according to the invention. If galacturonic acid oligosaccharides (hydrolysed pectin) were used instead of LvPectin the relative abundance in Bifidobacteriaceae did increase, however not to a similar extent. Hence, it is the specific combination of scGOS, IcFOS, LvPectin and BB536 that synergistically increases the relative abundance in Bifidobacteriaceae.

[0306] These results demonstrate the ability of the synbiotic composition / nutritional composition of the invention (scGOS, IcFOS, LvPectin and BB536) to significantly increase both acetic acid and butyric acid production compared to the individual components, hence illustrating a synergistic effect. Also, the increase in acetic acid and butyric acid production was much higher when the composition comprising LvPectin was used compared to the composition comprising AOS. Indicating that the specific combination of BB536 and the prebiotic combination with LvPectin according to the invention are responsible for the observed synergistic effect.

[0307] Example 4

[0308] To study the effect of the composition on epithelial barrier function trans epithelial resistance (TEER) was measured. TEER is a widely accepted quantitative technique to measure the integrity of tight junction dynamics in cell culture models of endothelial and epithelial monolayers. TEER values are strong indicators of the integrity of the cellular barriers and thus gut barrier function, see for example Srinivasan et al., TEER measurement techniques for in vitro barrier model systems, J Lab Autom, 2015, 20(2), 107 -126.

[0309] Samples

[0310] The dysbiosis induced fecal samples of Example 3 were also used in the TEER measurements. After the induction of dysbiosis the fecal samples were fermented with compositions 3 and 4 as described in Example 3. Samples were collected after 72 hours of fermentation. For this experiment 50 pl of dialysate was diluted 1 / 20 with Caco-2 medium (DMEM) without FCS and filtered (0.2um) and subsequently was added to the apical upper left channel A1 , 50 pl was added to apical channel A3. Also in the basolateral channel dialysates (1 / 20 diluted in medium and filtered (0.2 urn)) were added, 50 pl in C1 and 50 pl in C3.

[0311] Material and methods

[0312] Experiments were performed in the microfluidic OrganoPlate platform of Mimetas (Oegstgeest, Netherlands) and performed according to method as described in Nicolas et al., High throughput transepithelial electrical resistance (TEER) measurements on perfused membrane-free epithelia, LAB CHIP, 2021 , 21 , 1676-1685. In the experiment, Caco-2 ready plates were provided by Mimetas. The plates contained two tubules grown directly against an extracellular matrix without the use of artificial membranes. The TEER was measured by a four-point measurement in a single chip with electrodes contacting the cell culture medium that addresses either the luminal or basil side of the epithelium. The epithelial tubules in the Caco-2 plates were seeded in the upper channel of the plate (day 0) as described in Nicolas et al. (2021). Upon arrival (day 4) the plates were incubated horizontally and static for 1 h at 37 °C, whereafter the transport medium was removed and fresh Caco-2 medium (DM EM with FCS) was added according to standard protocols provided by Mimetas and as described in Nicolas et al. (2021). The plates were placed on a rocker in the incubator for 24 h at 37 °C. On day 5 the medium was refreshed and on day 7 the dialysates were added to the plate.

[0313] 50 pl of the dialysate was added to the top perfusion inlet wells (left and right channel A1 and A3). In the basolateral channels (C1 and C3) 50 pl of dialysates were added as described in the protocol by Mimetas and in Nicolas et al., (2021).

[0314] Each sample was added in five different chips on the plate (n=5) and TEER was measured over time according to the protocols of Mimetas as described in Nicolas et al., (2021).

[0315] Results

[0316] The effect on TEER values after 21 hours of Caco-2 treatment are shown in Figure 4.

[0317] The dialysates derived from the fecal samples treated with a composition comprising scGOS, IcFOS and LvPectin in a 9:1 :2 ratio and BB536 resulted in a significant increase in TEER compared to dialysates derived from the fecal samples treated with the composition comprising scGOS, IcFOS and AOS in a 9:1 :2 ratio and BB536.

[0318] Conclusion

[0319] These results demonstrate the ability of the nutritional composition of the invention (scGOS, IcFOS, LvPectin in a 9:1 :2 ratio and BB536) to beneficially affect the production of microbial metabolites which upon incubation with epithelial tubules lead to an improved effect on gut barrier function compared to a composition which comprises AOS instead of LvPectin, thus illustrating a synergistic effect of the composition on improving gut barrier function.

[0320] Example 5

[0321] An exemplary nutritional composition is provided. The exemplary nutritional composition is provided as a powder in a pack with instructions to reconstitute with water to obtain a ready- to-drink formulation. The composition comprises per 125 ml: 150 kcal, 20g protein, 3 g fat, 2.5 g digestible carbohydrates and 2.5 x 108 cfu / gram Bifidobacterium longum BL999. The composition further comprises:

Claims

CLAIMS1 . A synbiotic composition comprising:• 0.5-2.5 x 108cfu / gram on dry basis of Bifidobacterium ssp.;• at least 5wt.% on dry basis of non-digestible oligosaccharides, selected from galactooligosaccharides and / or fructooligosaccharides; and• 0.5-4 wt.% on dry basis of a pectin polysaccharides having a molecular weight between 50-2000 kDa, the backbone thereof comprising at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and having a degree of methylation (DM) of at least 35% and / or a pectin polysaccharide that has a degree of polymerisation of at least 260.

2. The synbiotic composition of claim 1 , wherein the pectin polysaccharide preferably have a molecular weight between 50-2000 kDa, the backbone thereof comprise at least 55% by weight of the polysaccharide of 1 ,4-linked a-D-galacturonic acid residues and have a degree of methylation (DM) of at least 35%.

3. The synbiotic composition of claim 1 , wherein the pectin polysaccharide preferably has a degree of polymerisation of at least 260.

4. The synbiotic composition of any of the preceding claims, wherein the probiotic bacterial strain is selected from Bifidobacterium spp., preferably Bifidobacterium long urn, more preferably Bifidobacterium longum BL999.

5. The synbiotic composition of any of the preceding claims, wherein it comprises 0.8 - 2.0 x 108cfu / gram on dry basis of Bifidobacterium spp.

6. The synbiotic composition of any of the preceding claims, wherein it comprises 5 - 15 wt.% on dry basis of non-digestible oligosaccharides, preferably 7 - 12 wt.% on dry basis of non-digestible oligosaccharides.

7. The synbiotic composition of any of the preceding claims, wherein it comprises galactooligosaccharides and fructooligosaccharides.

8. The synbiotic composition of any of the preceding claims, wherein the pectin polysaccharides have a molecular weight of 60-1000 kDa, more preferably 70-500 kDa, even more preferably 75 - 250 kDa.

9. The synbiotic composition of any of the preceding claims, wherein the pectin polysaccharides has a degree of polymerisation of at least 300, preferably at least 450.

10. The synbiotic composition of any of the preceding claims, wherein the pectin polysaccharides comprise one or more of the following:• a substantially unbranched backbone of galacturonic acid residues, preferably wherein less than 20% molar weight of the backbone of galacturonic acid residues is branched; and / or• wherein the backbone of the pectin polysaccharides comprises galacturonic acid residues and rhamnose residues in a molar ratio of more than 10:1 , preferably more than 20:1 ; and / or• the pectin polysaccharides having a degree of acetylation (DA) of less than 18%, preferably less than 15%, more preferably less than 10%, preferably less than 5%, even more preferably less than 4%.11 . The synbiotic composition of any of the preceding claims, wherein the weight ratio of the sum of non-digestible oligosaccharides to pectin polysaccharides is between 2 : 1 to 30 : 1 , preferably 3 : 1 to 20 : 1 .

12. The synbiotic composition of any of the preceding claims, wherein it comprises less than 20 wt.% on dry basis of inulin, preferably less than 10 wt.%, more preferably less than 5 wt.%.

13. A nutritional composition comprising the synbiotic composition according to any one of the preceding claims, macro ingredients selected from digestible carbohydrates, fats, proteins, and preferably further comprising a therapeutically effective combination of:• omega-3 fatty acids;• vitamin B2;• vitamin D3, and• optionally a choline source selected from choline or a salt thereof and / or phosphatidylcholine.

14. A composition according to any one of the preceding claims for use in the prevention and / or treatment of gut microbial dysbiosis in an adult subject, preferably antibiotic- induced gut microbial dysbiosis, preferably wherein the subject is or has been submitted to antibiotic therapy for at least 4 days, preferably 7 days.

15. A composition according to any one of claims 1 - 13 for use in preventing and / or treating intestinal barrier disruption in a subject

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