EP3968787A1

EP3968787A1 - Fermented formula for improving intestinal development

Info

Publication number: EP3968787A1
Application number: EP20724882.4A
Authority: EP
Inventors: Mona MISCHKE; Ingrid Brunhilde RENES; Isabelle VAN SEUNINGEN; Audrey Vincent
Original assignee: Nutricia NV
Current assignee: Nutricia NV
Priority date: 2019-05-15
Filing date: 2020-05-15
Publication date: 2022-03-23
Also published as: CN113163835A; WO2020229689A1

Abstract

The present invention concerns inducing a gut maturation pattern in formula fed infants that is more similar to that of human milk fed infants, by administering a nutritional composition that is at least partly fermented by lactic acid producing bacteria. The present invention also relates to prevention precocious maturation of the intestine of an infant.

Description

FERMENTED FORMULA FOR IMPROVING INTESTINAL DEVELOPMENT

FIELD OF THE INVENTION

The current invention is in the field of infant nutrition and its effect on the maturation of the intestinal tract.

BACKGROUND OF THE INVENTION

The gastrointestinal tract (gut) undergoes significant and well-timed growth and maturation during the perinatal period, leading to the establishment of proper digestive and absorptive functions, intestinal barrier and immune homeostasis. These are crucial factors for health and well-being for the infant, and also may have effects later in life.

Early life nutrition is known to affect this developmental trajectory of the gut. Compared to mother’s milk, formula concepts have been shown to change the timing and outcome of gut maturation in clinical and preclinical studies, and are indicated to induce precocious maturation of the small intestine. For example Cummins and Thompson (Gut, 2002, 51 :748-754) found that breast fed infants have smaller crypts in the intestine than bottle fed infants. Bhinder et al (Scientific Reports, 2017, 7: 45275) found that control formula yielded significant deficits in intestinal development as compared to mother milk fed littermates, and that addition of milk fat globule membrane (MFGM) to formula restored intestinal growth, Paneth and goblet cell numbers, and tight junction protein patterns to that of mother milk fed pups. Dvorak et al (JPGN, 2000, 31 : 162-169) discloses that formula concept with cow’s milk protein induced intestinal overgrowth and precocious maturation of the small intestine compared to rats fed pooled rat milk. DeVos et al (Livestock Sci 2014, 165-173) show that formula fed piglets, compared to mothers’ milk fed piglets until day 28 had greater absorptive area, deeper crypts and increased maltase and sucrase activities in the intestine.

An intact gut epithelial barrier is key in maintaining intestinal immune homeostasis and can be modulated by the gut microbiota. Moreover, environmental factors like stress and antibiotics negatively impact the gut barrier, which results into a proinflammatory immune response and can also lead to visceral hypersensitivity. But a precociously decreased intestinal permeability in infants could affect health long- lastingly by altering immune maturation. It is thought that the elevated intestinal permeability in neonates facilitates the development of food tolerance and immune function by enabling the access of antigens to immune cells of the gut.

WO 2004/1 12509 discloses a composition for inducing a pattern of gut barrier maturation similar to that observed with breast-feeding and able to improve gut barrier maturation, e.g. during neonatal stress. Maternal separation increased in rats the intestinal permeability and a blend containing LC-PUFA, Lactobacillus paracasei and non-digestible oligosaccharides restored the intestinal permeability to normal levels. WO 2014/160150 discloses a method of enhancing maturation of a lung, gut, or both in an infant, the method comprising the step of administering to the infant a nutritional composition comprising from about 3 weight % phospholipids to about 20 weight % phospholipids, based on total fat of the nutritional composition.

WO 2009/047754 discloses a high palmitic acid lipid composition high in palmitic acid at the sn-2 position for the prevention and treatment of gastrointestinal diseases and disorders, and for promoting intestinal development, maturation, adaptation and differentiation.

Human milk is the preferred food for infants. However, it is not always possible or desirable to breastfeed an infant. In such cases infant formulae or follow on formulae are a good alternative. These formulae should have an optimal composition in order to mimic the beneficial effects of human milk as close as possible. For the benefit of infants that will not be completely breast fed, there is a continuing need to develop infant formulas which will replicate human milk as close as possible, both in terms of its nutritional and its bioactive properties.

SUMMARY OF THE INVENTION

Employing a mouse model, the inventors surprisingly found that administration of a nutritional composition which was partly fermented stimulated a gut maturation development more similar to the mother-fed situation, when compared to a nutritional composition not being fermented or partly fermented.

Morphologically, the crypt-villus length in the intestine of mice fed non-fermented formula was significantly greater than that of mother fed mice, while in mice fed the experimental partly fermented nutritional composition the crypt-villus length resembled that of mother fed mice. The intestinal permeability in mice fed the experimental partly fermented composition was also more similar to mother fed-mice, while it was significantly lower in mice receiving the control formula.

So, standard formula feeding induced intestinal overgrowth, when compared to the mother-fed situation, but feeding a partly fermented formula partly prevented this morphological effect. Furthermore, feeding a partly fermented formula supported an intestinal barrier function maturation pattern more similar to the breast-fed situation, which was not precocious as seen in the standard formula group. Additionally, under non-neonatal conditions, or under conditions where stress induced an increase in the intestinal permeability, the partly fermented formula decreased the intestinal permeability when compared with a non-fermented formula. Therefore partly fermented infant formula prevent precocious maturation while simultaneously protecting the gut barrier function once the barrier has matured and is under stress conditions. DETAILED DESCRIPTION OF THE INVENTION

The present invention thus relates in a first aspect to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants.

The invention relates in a second aspect to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in preventing precocious maturation of the intestine in an infant.

In some jurisdictions the first aspect of the invention can be worded as the use of a fermented composition for the manufacture of a nutritional composition selected from an infant formula and a follow on formula for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In these jurisdictions the second aspect of the invention can be worded as the use of a fermented composition for the manufacture of a nutritional composition selected from an infant formula and a follow on formula for preventing precocious maturation of the intestine in an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In some jurisdictions the first aspect of the present invention can be worded as a method for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In these jurisdictions the second aspect of the present invention can be worded as a method for preventing precocious maturation of the intestine in an infant, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In some jurisdictions administering a nutritional composition to an infant is considered non-therapeutic. In these jurisdictions, the first aspect of the present invention can be worded as a non-therapeutic method for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria. In these jurisdictions the second aspect of the present invention can be worded as a non-therapeutic method for preventing precocious maturation of the intestine in an infant, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

Alternatively, the first aspect of the invention can also be worded as the use of a nutritional composition selected from an infant formula and a follow on formula for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

The second aspect of the invention can also be worded as the use of a nutritional composition selected from an infant formula and a follow on formula for preventing precocious maturation of the intestine in an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In the context of the present invention the term “more similar to the intestinal maturation pattern observed in human milk fed infants” is as compared to when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered.

In the context of the present invention the term“prevention” means“reducing the risk of” or“reducing the severity of.

In the context of the present invention the term“precocious maturation of the intestine” refers to a premature maturation of the intestine, in other words to a maturation of the intestine that occurs too early or too rapidly.

In a preferred embodiment of the composition for use, the method or the use according to the first and second aspect of the invention, the infant has an age of 0 to 6 months. Preferably the infant has an age of 0 to 4 months.

In another preferred embodiment of the composition for use, the method or the use according to the first and second aspect of the invention, the infant is a term infant. Preferably the infant is a healthy term infant.

In this document and in its claims, the verb "to comprise" and its conjugations is used in its non-limiting sense to mean that items following the word are included, but items not specifically mentioned are not excluded. In addition, reference to an element by the indefinite article "a" or "an" does not exclude the possibility that more than one of the element is present, unless the context clearly requires that there be one and only one of the elements. The indefinite article "a" or "an" thus usually means "at least one. Wt% means weight percentage. Fermented composition

The nutritional composition for use, and the nutritional composition in the methods or uses according to the present invention, hereafter also referred to as the present nutritional composition, or nutritional composition of the invention or final nutritional composition, is at least party fermented. A partly fermented nutritional composition comprises at least for a part a composition that was fermented by lactic acid producing bacteria. It was shown that the presence of fermented composition in the final nutritional composition, upon administration, induced an intestinal maturation pattern - in particular the growth and morphological development of the intestine, more in particular the intestinal crypt-villus length - which was more similar to the intestinal maturation pattern observed in mother-fed, when compared to administration of a non-fermented infant formula or follow on formula. In addition, precocious maturation of the intestine, in particular of the intestinal barrier function, was prevented.

The fermentation preferably takes place during the production process of the nutritional composition. Preferably, the nutritional composition does not contain significant amounts of viable bacteria in the final product, and this can be achieved by heat inactivation after fermentation or inactivation by other means. Preferably the fermented composition is a milk-derived product, which is a milk substrate that is fermented by lactic acid producing bacteria, wherein the milk substrate comprises at least one selected from the group consisting of milk, whey, whey protein, whey protein hydrolysate, casein, casein hydrolysate or mixtures thereof. Suitably, nutritional compositions comprising fermented compositions and non-digestible oligosaccharide and their way of producing them are described in WO 2009/151330, WO 2009/151331 and WO 2013/187764.

The fermented composition preferably comprises bacterial cell fragments like glycoproteins, glycolipids, peptidoglycan, lipoteichoic acid (LTA), lipoproteins, nucleotides, and/or capsular polysaccharides. It is of advantage to use the fermented composition comprising inactivated bacteria and/or cell fragments directly as a part of the final nutritional product, since this will result in a higher concentration of bacterial cell fragments. When commercial preparations of lactic acid producing bacteria are used, these are usually washed and material is separated from the aqueous growth medium comprising the bacterial cell fragments, thereby reducing or eliminating the presence of bacterial cell fragments. Furthermore, upon fermentation and/or other interactions of lactic acid producing bacteria with the milk substrate, additional bio-active compounds can be formed, such as short chain fatty acids, bioactive peptides and/or oligosaccharides, and other metabolites, which may also result in an intestinal microbiota- function more similar to the intestinal microbiota-function of breastfed infants. Such bioactive compounds that are produced during fermentation by lactic acid producing bacteria may also be referred to as post-biotics. A composition comprising such post-biotics is thought to be advantageously closer to breast milk, as breast milk is not a clean synthetic formula, but contains metabolites, bacterial cells, cell fragments and the like. Therefore the fermented composition, in particular fermented milk-derived product, is believed to have an improved effect compared to non-fermented milk-derived product without or with merely added lactic acid producing bacteria on the prevention of precocious maturation of the intestine in an infant, and inducing, in an infant, an intestinal maturation pattern which is more similar to the intestinal maturation pattern observed in human milk fed infants.

Preferably the final nutritional composition comprises 5 to 97.5 wt% of the fermented composition based on dry weight, more preferably 10 to 90 wt%, more preferably 20 to 80 wt%, even more preferably 25 to 60 wt%. As a way to specify that the final nutritional composition comprises at least partly a fermented composition, and to specify the extent of fermentation, the level of the sum of lactic acid and lactate in the final nutritional composition can be taken, as this is the metabolic end product produced by the lactic acid producing bacteria upon fermentation. The present final nutritional composition preferably comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate based on dry weight of the composition, more preferably 0.1 to 1 .0 wt%, even more preferably 0.2 to 0.5 wt%. Preferably at least 50 wt%, even more preferably at least 90 wt%, of the sum of lactic acid and lactate is in the form of the L(+)-isomer. Thus in one embodiment the sum of L(+)-lactic acid and L(+)-lactate is more than 50 wt%, more preferably more than 90 wt%, based on the sum of total lactic acid and lactate. Herein L(+)-lactate and L(+)-lactic acid is also referred to as L-lactate and L-lactic acid.

Lactic acid producing bacteria used for producing the fermented ingredient

Lactic acid producing bacteria used for preparing the fermented ingredient, in particular for fermentation of the milk substrate are preferably provided as a mono- or mixed culture. Lactic acid producing bacteria consists of the genera Bifidobacterium, Lactobacillus, Carnobacterium, Enterococcus, Lactococcus, Leuconostoc, Oenococcus, Pediococcus, Streptococcus, Tetragenococcus, Vagococcus and Weissella. Preferably the lactic acid producing bacteria used for fermentation comprises bacteria of the genus Bifidobacterium and/or Streptococcus.

Preferably the Streptococcus is a strain of S. thermophilus. Selection of a suitable strain of S. thermophilus is described in example 2 of EP 778885 and in example 1 of FR 2723960. In a further preferred embodiment according to the present invention, the nutritional composition comprises 10²-10⁵ cfu living bacteria of S. thermophilus, per g dry weight of the final nutritional composition, preferably the final nutritional composition comprises 10³-10⁴ cfu living bacteria of S. thermophilus per g dry weight.

Preferred strains of S. thermophilus to prepare the fermented ingredient for the purpose of the present invention have been deposited by Compagnie Gervais Danone at the Collection Nationale de Cultures de Microorganismes (CNCM) run by the Institut Pasteur, 25 rue du Docteur Roux, Paris, France on 23 August 1995 under the accession number 1-1620 and on 25 August 1994 under the accession number 1-1470. Other S. thermophilus strains are commercially available.

Bifidobacteria are Gram-positive, anaerobic, rod-shaped bacteria. Preferred Bifidobacterium species to prepare the fermented ingredient for the purpose of the present invention preferably have at least 95 % identity of the 16 S rRNA sequence when compared to the type strain of the respective Bifidobacterium species, more preferably at least 97% identity as defined in handbooks on this subject for instance Sambrook, J., Fritsch, E.F., and Maniatis, T. (1989), Molecular Cloning, A Laboratory Manual, 2nd ed., Cold Spring Harbor (N.Y.) Laboratory Press. The bifodobacteria preferably used are also described by Scardovi, V. Genus Bifidobacterium p.1418 - p.1434. In: Bergey’s manual of systematic Bacteriology. Vol. 2. Sneath, P.H.A., N.S. Mair, M.E. Sharpe and J.G. Holt (ed.). Baltimore: Williams & Wilkins. 1986. 635 p. Preferably the lactic acid producing bacteria used for fermentation comprises or is at least one Bifidobacterium selected from the group consisting of B. breve, B. infantis, B. bifidum, B. catenulatum, B. adolescentis, B. thermophilum, B. gallicum, B. animalis or lactis, B. angulatum, B. pseudocatenulatum, B. thermacidophilum and B. longum more preferably B. breve, B. infantis, B. bifidum, B. catenulatum, B. longum, more preferably B. longum and B. breve, even more preferably B. breve, more preferably B. breve selected from the group consisting of B. breve Bb-03 (Rhodia/Danisco), B. breve M-16V (Morinaga), B. breve R0070 (Institute Rosell, Lallemand), B. breve BR03 (Probiotical), B. breve BR92 (Cell Biotech) DSM 20091 , LMG 1 1613 and B. breve 1-2219 deposited at the CNCM, Paris France. Most preferably, the B. breve is B. breve M-16V (Morinaga) or B. breve 1-2219, even more preferably B. breve 1-2219.

Most preferably the nutritional composition of the invention comprises a fermented composition that is fermented by lactic acid producing bacteria comprising both B. breve and S. thermophilus. In one embodiment the fermentation by lactic acid producing bacteria is fermentation by Streptococcus thermophilus and Bifidobacterium breve. In one embodiment, the final nutritional composition comprises fermented composition wherein the lactic acid producing bacteria are inactivated after fermentation.

Preferably the fermented composition is not fermented by Lactobacillus bulgaricus. L. bulgaricus fermented products are considered not suitable for infants, since in young infants the specific dehydrogenase that converts D-lactate to pyruvate is far less active than the dehydrogenase which converts L-lactate.

Preferably the nutritional composition of the invention comprises inactivated lactic acid producing bacteria and/or bacterial fragments derived from lactic acid producing bacteria being the equivalent of more than 1x10⁴ cfu lactic acid producing bacteria per g based on dry weight of the final composition, more preferably 1x10⁵ cfu, even more preferably 1x10⁶ cfu. Preferably the inactivated bacteria or bacterial fragments are the equivalent of less than 1x10¹³ cfu lactic acid producing bacteria per g based on dry weight of the final composition, more preferably 1x10¹¹ cfu, even more preferably 1x10¹⁰ cfu. The correlation of inactivated lactic acid bacteria and the equivalence with cfu can be determined by molecular techniques, known in the art, or by checking the production process.

Process of fermentation

Preferably the fermented composition is a milk-derived product, which is a milk substrate that is fermented by lactic acid producing bacteria, and said milk substrate comprising at least one selected from the group consisting of milk, whey, whey protein, whey protein hydrolysate, casein, casein hydrolysate and mixtures thereof. The milk derived product or milk substrate to be fermented is suitably present in an aqueous medium. The milk substrate to be fermented comprises at least one selected from the group consisting of milk, whey, whey protein, whey protein hydrolysate, casein, casein hydrolysate and mixtures thereof. Milk can be whole milk, semi-skimmed milk, skimmed milk or mixtures thereof. Preferably the milk substrate to be fermented comprises skimmed milk. Whey can be sweet whey, and/or acid whey. Preferably the whey is present in a concentration of 3 to 80 g dry weight per I aqueous medium containing milk substrate, more preferably 40 to 60 g per I. Preferably whey protein hydrolysate is present in 2 to 80 g dry weight per I aqueous medium containing milk substrate, more preferably 5 to15 g/l. Preferably lactose is present in 5 to 50 g dry weight per I aqueous substrate, more preferably 1 to 30 g/l. Preferably the aqueous medium containing milk substrate comprises buffer salts in order to keep the pH within a desired range. Preferably sodium or potassium dihydrogen phosphate is used as buffer salt, preferably in 0.5 to 5 g/l, more preferably 1 .5 to 3 g per I. Preferably the aqueous medium containing milk substrate comprises cysteine in amount of 0.1 to 0.5 g per I aqueous substrate, more preferably 0.2 to 0.4 g/l. The presence of cysteine results in low redox potential of the substrate which is advantageous for activity of lactic acid producing bacteria, particularly bifidobacteria. Preferably the aqueous medium containing milk substrate comprises yeast extract in an amount of 0.5 to 5 g/l aqueous medium containing milk substrate, more preferably 1 .5 to 3 g/l. Yeast extract is a rich source of enzyme co-factors and growth factors for lactic acid producing bacteria. The presence of yeast extract will enhance the fermentation by lactic acid producing bacteria.

Suitably the milk substrate, in particular the aqueous medium containing milk substrate, is pasteurised before the fermentation step, in order to eliminate the presence of unwanted living bacteria. Suitably the product is pasteurised after fermentation, in order to inactivate enzymes. Suitably the enzyme inactivation takes place at 75 °C for 3 min. Suitably the aqueous medium containing milk substrate is homogenised before and/or the milk-derived product is homogenised after the fermentation. Homogenisation results in a more stable substrate and/or fermented product, especially in the presence of fat.

The inoculation density is preferably between 1x10² to 5x10¹⁰, preferably between 1x10⁴ to 5x10⁹ cfu lactic acid producing bacteria/ml aqueous medium containing milk substrate, more preferably between 1x10⁷ to 1x10⁹ cfu lactic acid producing bacteria/ml aqueous medium containing milk substrate. The final bacteria density after fermentation is preferably between 1x10³ to 1x10¹⁰, more preferably between 1x10⁴ to 1x10⁹ cfu/ml aqueous medium containing milk substrate.

The fermentation is preferably performed at a temperature of approximately 20 °C to 50 °C, more preferably 30 °C to 45 °C, even more preferably approximately 37 °C to 42 °C. The optimum temperature for growth and/or activity for lactic acid producing bacteria, more particularly lactobacilli and/or bifidobacteria is between 37 °C and 42 °C. The incubation is preferably performed at a pH of 4 to 8, more preferably 6 to 7.5. This pH does not induce protein precipitation and/or an adverse taste, while at the same time lactic acid producing bacteria such as lactobacilli and/or bifidobacteria are able to ferment the milk substrate.

The incubation time preferably ranges from 10 minutes to 48 h, preferably from 2 h to 24 h, more preferably from 4 h to 12 h. A sufficient long time enables fermentation and the concomitant production of immunogenic cell fragments such as glycoproteins, glycolipids, peptidoglycan, lipoteichoic acid (LTA), flagellae, lipoproteins, DNA and/or capsular polysaccharides and metabolites (postbiotics) to take place at a sufficient or higher extent, whereas the incubation time needs not be unnecessarily long for economic reasons.

Preferably, a milk derived product or milk substrate, preferably skimmed milk, is pasteurized, cooled and fermented with one or more lactic acid producing strains, preferably a strain of S. thermophilus, to a certain degree of acidity, upon which the fermented product is cooled and stored. Preferably a second milk-derived product is prepared in a similar way using one or more Bifidobacterium species for fermentation. Subsequently, the two fermented products are preferably mixed together and mixed with other components making up an infant formula, except the fat component. Preferably, the mixture is preheated, and subsequently fat is added in-line, homogenized, pasteurized and dried. Alternatively the fermentation takes place having both Bifidobacterium, preferably B. breve, and S. thermophilus in the fermentation tank.

Procedures to prepare fermented composition suitable for the purpose of the present invention are known per se. EP 778885, which is incorporated herein by reference, discloses in particular in example 7 a suitable process for preparing a fermented ingredient. FR 2723960, which is incorporated herein by reference, discloses in particular in example 6 a suitable process for preparing a fermented ingredient. Briefly, a milk substrate, preferably pasteurised, containing lactose and optionally further macronutrients such as fats, preferably vegetable fats, casein, whey protein, vitamins and/or minerals etc. is concentrated, e.g. to between 15 to 50% dry matter and then inoculated with S. thermophilus, for example with 5% of a culture containing 10⁶ to 10¹⁰ bacteria per ml. Preferably this milk substrate comprises milk protein peptides. Temperature and duration of fermentation are as mentioned above. Suitably after fermentation the fermented ingredient may be pasteurised or sterilized and for example spray dried or lyophilised to provide a form suitable to be formulated in the end product.

A preferred method for preparing the fermented composition to be used in the nutritional composition of invention is disclosed in WO 01/01785, more particular in examples 1 and 2. A preferred method for preparing the fermented composition to be used in the nutritional composition of invention is described in WO 2004/093899, more particularly in example 1 .

Living cells of lactic acid producing bacteria in the fermented composition are after fermentation preferably eliminated, for example by inactivation and/or physical removal. The cells are preferably inactivated. Preferably the lactic acid producing bacteria are heat killed after fermentation of the milk substrate. Preferable ways of heat killing are (flash) pasteurization, sterilization, ultra-high temperature treatment, high temperature/short time heat treatment, and/or spray drying at temperatures bacteria do not survive. Cell fragments are preferably obtained by heat treatment. With this heat treatment preferably at least 90 % of living microorganisms are inactivated, more preferably at least 95 %, even more preferably at least 99 %. Preferably the fermented nutritional composition comprises less than 1x10⁵ colony forming units (cfu) living lactic acid bacteria per g dry weight. The heat treatment preferably is performed at a temperature ranging from 70 to180 °C, preferably from 80 to 150 °C, preferably for about 3 minutes to 2 hours, preferably in the range of 80 to 140 °C for 5 minutes to 40 minutes. Inactivation of the lactic acid bacteria advantageously results in less post acidification and a safer product. This is especially advantageous when the nutritional composition is to be administered to infants. Suitably after fermentation the fermented ingredient may be pasteurised or sterilized and for example spray dried or lyophilised to provide a form suitable to be formulated in the end product.

Non-digestible oligosaccharides

The present nutritional composition preferably comprises non-digestible oligosaccharides and preferably comprises at least two different non-digestible oligosaccharides, in particular two different sources of non-digestible oligosaccharides. The presence of non-digestible oligosaccharides advantageously further improves the prevention of precocious maturation of the intestine in an infant, and further induces, in an infant, an intestinal maturation pattern which is more similar to the intestinal maturation pattern observed in human milk fed infants.

The term“oligosaccharides” as used herein refers to saccharides with a degree of polymerization (DP) of 2 to 250, preferably a DP 2 to 100, more preferably 2 to 60, even more preferably 2 to 10. The term “non-digestible oligosaccharides” 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, sucrose, lactose, maltose and maltodextrins are considered digestible.

Preferably the present non-digestible oligosaccharides are soluble. The term "soluble" as used herein, when having reference to a polysaccharides, fibres or oligosaccharides, means that the substance is at least soluble according to the method described by L. Prosky et al., J. Assoc. Off. Anal. Chem. 71 , 1017- 1023 (1988).

Preferably, the nutritional composition for use or the nutritional compositions used in the methods and uses of the present invention comprises 2.5 to 15 wt.% non-digestible oligosaccharides, based on dry weight of the formula. If present, the non-digestible oligosaccharides included in the present composition for use and in the present nutritional compositions in the methods or uses according to the invention preferably include a mixture of different non-digestible oligosaccharides. The non-digestible oligosaccharides are preferably selected from the group consisting of fructo-oligosaccharides, such as inulin, non-digestible dextrins, galacto-oligosaccharides, such as transgalacto-oligosaccharides, xylo-oligosaccharides, arabino- oligosaccharides, arabinogalacto-oligosaccharides, gluco-oligosaccharides, gentio-oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan-oligosaccharides, isomalto- oligosaccharides, nigero-oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, fuco-oligosaccharides, sialyloligosaccharides and mixtures thereof. Such oligosaccharides share many biochemical properties and have similar functional benefits including improving the intestinal microbiota-function. Yet it is understood that some non-digestible oligosaccharides and preferably some mixtures have an even further improved effect. Therefore more preferably the non-digestible oligosaccharides are selected from the group consisting of fructo- oligosaccharides, such as inulin, and galacto-oligosaccharides, such as betagalacto-oligosaccharides, and mixtures thereof, even more preferably betagalacto-oligosaccharides and/or inulin, most preferably betagalacto-oligosaccharides. In one embodiment in the nutritional composition according to the present invention, the non-digestible oligosaccharides are selected from the group consisting of galacto- oligosaccharides, fructo-oligosaccharides and mixtures of thereof, more preferably betagalacto- oligosaccharides, fructo-oligosaccharides and mixtures thereof. In a preferred embodiment the non- digestible oligosaccharides are a mixture of betagalacto-oligosaccharides and fructo-oligosaccharides.

The non-digestible oligosaccharides preferably comprise galacto-oligosaccharides. The galacto- oligosaccharides are preferably selected from the group consisting of betagalacto-oligosaccharides, alphagalacto-oligosaccharides, and galactan. According to a more preferred embodiment galacto- oligosaccharides are betagalacto-oligosaccharides. Preferably the galacto-oligosaccharides comprise galacto-oligosaccharides with beta(1 ,4), beta(1 ,3) and/or beta(1 ,6) glycosidic bonds and a terminal glucose. Transgalacto-oligosaccharides is for example available under the trade name Vivinal®GOS (Domo FrieslandCampina Ingredients), Bi2muno (Clasado), Cup-oligo (Nissin Sugar) and Oligomate55 (Yakult).

The non-digestible oligosaccharides preferably comprise fructo-oligosaccharides. Fructo- oligosaccharides may in other context have names like fructopolysaccharides, oligofructose, polyfructose, polyfructan, inulin, levan and fructan and may refer to oligosaccharides comprising beta- linked fructose units, which are preferably linked by beta(2,1) and/or beta(2,6) glycosidic linkages, and a preferable DP between 2 and 200. Preferably, the fructo-oligosaccharides contain a terminal beta(2,1) glycosidic linked glucose. Preferably, the fructo-oligosaccharides contain at least 7 beta-linked fructose units. In a further preferred embodiment inulin is used. Inulin is a type of fructo-oligosaccharides wherein at least 75% of the glycosidic linkages are beta(2,1) linkages. Typically, inulin has an average chain length between 8 and 60 monosaccharide units. A suitable fructo-oligosaccharides for use in the compositions of the present invention is commercially available under the trade name Raftiline®HP (Orafti). Other suitable sources are Raftilose (Orafti), Fibrulose and Fibruline (Cosucra) and Frutafit and Frutalose (Sensus).

Preferably the present nutritional composition comprises a mixture of galacto-oligosaccharides and fructo-oligosaccharides. Preferably the mixture of galacto-oligosaccharides and fructo-oligosaccharides 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 . This weight ratio is particularly advantageous when the galacto-oligosaccharides have a low average DP and fructo-oligosaccharides have a relatively high DP. Most preferred is a mixture of galacto-oligosaccharides with an average DP below 10, preferably below 6, and fructo-oligosaccharides with an average DP above 7, preferably above 1 1 , even more preferably above 20. Therefore, in a preferred embodiment the nutritional composition for use according to the present invention and the nutritional composition in the uses and methods of the present invention is at least partly fermented by lactic acid producing bacteria and comprises galacto-oligosaccharides and fructo-oligosaccharides, wherein the galacto-oligosaccharides and fructo-oligosaccharides are present in a weight ratio of from 2/1 to 15/1 , and wherein the galacto- oligosaccharides have an average DP below 10 and the fructo-oligosaccharides have an average DP above 7. In this embodiment it is further preferred that the galacto-oligosaccharides and fructo- oligosaccharides are present in a weight ratio of from 5/1 to 12/1 , the galacto-oligosaccharides have an average DP below 10 and the fructo-oligosaccharides have an average DP above 1 1 . In this embodiment it is particularly preferred that the galacto-oligosaccharides and fructo-oligosaccharides are present in a weight ratio of from 8/1 to 10/1 , the galacto-oligosaccharides have an average DP below 6 and the fructo-oligosaccharides have an average DP above 20. In these embodiments it is further preferred that the nutritional composition comprises 2.5 to 15 wt.% of the galacto-oligosaccharides and fructo-oligosaccharides, based on dry weight of the formula, more preferably 3.0 to 10 wt%, and most preferably 5.0 to 7.5 wt%, all based on dry weight of the nutritional composition.

Preferably the present nutritional composition comprises a mixture of short chain (sc) fructo- oligosaccharides and long chain (lc) fructo-oligosaccharides. Preferably the mixture of short chain fructo-oligosaccharides and long chain fructo-oligosaccharides is present in a weight ratio of from 1/99 to 99/1 , more preferably from 1/19 to 19/1 , even more preferably from 1/10 to 19/1 , more preferably from 1/5 to 15/1 , more preferably from 1/1 to 10/1 . Preferred is a mixture of short chain fructo- oligosaccharides with an average DP below 10, preferably below 6 and long chain fructo- oligosaccharides with an average DP above 7, preferably above 1 1 , even more preferably above 20. Therefore, in another preferred embodiment the nutritional composition for use according to the present invention and the nutritional composition in the uses and methods of the present invention is at least partly fermented by lactic acid producing bacteria and comprises short chain fructo-oligosaccharides and long chain fructo-oligosaccharides, wherein the short chain fructo-oligosaccharides and long chain fructo-oligosaccharides are present in a weight ratio of from 1/10 to 19/1 , and wherein the short chain fructo-oligosaccharides have an average DP below 10 and the long chain fructo-oligosaccharides have an average DP above 7. In this embodiment it is further preferred that the short chain fructo- oligosaccharides and long chain fructo-oligosaccharides are present in a weight ratio of from 1/5 to 15/1 , the short chain fructo-oligosaccharides have an average DP below 10 and the long chain fructo- oligosaccharides have an average DP above 1 1 . In this embodiment it is particularly preferred that the galacto-oligosaccharides and fructo-oligosaccharides are present in a weight ratio of from 1/1 to 10/1 , the short chain fructo-oligosaccharides have an average DP below 6 and the long chain fructo- oligosaccharides have an average DP above 20. In these embodiments it is further preferred that the nutritional composition comprises 2.5 to 15 wt.% of the short chain fructo-oligosaccharides and long chain fructo-oligosaccharides, based on dry weight of the formula, more preferably 3.0 to 10 wt%, and most preferably 5.0 to 7.5 wt%, all based on dry weight of the nutritional composition.

Preferably the present nutritional composition comprises a mixture of short chain fructo- oligosaccharides and short chain galacto-oligosaccharides. Preferably the mixture of short chain fructo- oligosaccharides and short chain galacto-oligosaccharides is present in a weight ratio of from 1/99 to 99/1 , more preferably from 1 /19 to 19/1 , even more preferably from 1 /10 to 19/1 , more preferably from 1/5 to 15/1 , more preferably from 1/1 to 10/1 . Preferred is a mixture of short chain fructo- oligosaccharides and galacto-oligosaccharides with an average DP below 10, preferably below 6. Therefore, in another preferred embodiment the nutritional composition for use according to the present invention and the nutritional composition in the uses and methods of the present invention is at least partly fermented by lactic acid producing bacteria and comprises short chain fructo-oligosaccharides and short chain galacto-oligosaccharides, wherein the short chain fructo-oligosaccharides and short chain galacto-oligosaccharides are present in a weight ratio of from 1/10 to 19/1 , and wherein the short chain fructo-oligosaccharides have an average DP below 10 and the short chain galacto- oligosaccharides have an average DP below 10. In this embodiment it is further preferred that the short chain fructo-oligosaccharides and short chain galacto-oligosaccharides are present in a weight ratio of from 1 /5 to 15/1 , and the short chain fructo-oligosaccharides and short chain galacto-oligosaccharides have an average DP below 10. In this embodiment it is particularly preferred that the galacto- oligosaccharides and fructo-oligosaccharides are present in a weight ratio of from 1/1 to 10/1 , and the short chain fructo-oligosaccharides and short chain galacto-oligosaccharides have an average DP below 6. In these embodiments it is further preferred that the nutritional composition comprises 2.5 to 15 wt.% of the short chain fructo-oligosaccharides and short chain galacto-oligosaccharides, based on dry weight of the formula, more preferably 3.0 to 10 wt%, and most preferably 5.0 to 7.5 wt%, all based on dry weight of the nutritional composition.

The present nutritional composition preferably comprises 2.5 to 20 wt% total non-digestible oligosaccharides, more preferably 2.5 to 15 wt%, even more preferably 3.0 to 10 wt%, most preferably 5.0 to 7.5 wt%, based on dry weight of the nutritional composition. Based on 100 ml the present nutritional composition preferably comprises 0.35 to 2.5 wt% total non-digestible oligosaccharides, more preferably 0.35 to 2.0 wt%, even more preferably 0.4 to 1 .5 wt%, based on 100 ml of the nutritional composition. A lower amount of non-digestible oligosaccharides may be less effective in improving the prevention of precocious maturation of the intestine in an infant, and inducing, in an infant, an intestinal maturation pattern which is more similar to the intestinal maturation pattern observed in human milk fed infants, whereas a too high amount will result in side-effects of bloating and abdominal discomfort.

Infant and follow on formula

The nutritional composition used according to the present invention is an infant formula or a follow on formula. More preferably the nutritional composition is an infant formula. The present nutritional composition can be advantageously applied as a complete nutrition for infants. Preferably the present nutritional composition is an infant formula. An infant formula is defined as a formula for use in infants and can for example be a starter formula, intended for infants of 0 to 6 or 0 to 4 months of age. A follow on formula is intended for infants of 4 or 6 months to 12 months of age. At this age infants start weaning on other food. The present composition preferably comprises a lipid component, protein component and carbohydrate component and is preferably administered in liquid form. The present nutritional composition may also be in the form of a dry food, preferably in the form of a powder which is accompanied with instructions as to mix said dry food, preferably powder, with a suitable liquid, preferably water. Therefore in one embodiment, the nutritional composition for use or the nutritional composition in the methods and uses according to the present invention is in the form of a powder, suitable to reconstitute with water to provide a ready to drink formula. In another embodiment, the nutritional composition for use or the nutritional composition in the methods and uses according to the present invention is in the form of a liquid.

The nutritional composition for use or the nutritional composition in the methods and uses according to the invention preferably comprises other fractions, such as vitamins, minerals, trace elements and other micronutrients in order to make it a complete nutritional composition. Preferably infant formulae and follow on formulae comprise vitamins, minerals, trace elements and other micronutrients according to international directives.

The present nutritional composition preferably comprises lipid, protein and digestible carbohydrate wherein the lipid provides 5 to 50% of the total calories, the protein provides 5 to 50% of the total calories, and the digestible carbohydrate provides 15 to 90% of the total calories. Preferably, in the present nutritional composition the lipid provides 35 to 50% of the total calories, the protein provides 7.0 to 12.5% of the total calories, and the digestible carbohydrate provides 40 to 55% of the total calories. For calculation of the % of total calories for the protein, the total of energy provided by proteins, peptides and amino acids needs to be taken into account. Preferably the lipid provides 3 to 7 g lipid per 100 kcal, preferably 4 to 6 g per 100 kcal, the protein provides 1 .6 to 4 g per 100 kcal, preferably 1 .7 to 2.5 g per 100 kcal and the digestible carbohydrate provides 5 to 20 g per 100 kcal, preferably 8 to 15 g per 100 kcal of the nutritional composition. Preferably the present nutritional composition comprises lipid providing 4 to 6 g per 100 kcal, protein providing 1 .6 to 2.0 g per 100 kcal, more preferably 1 .7 to 1 .9 g per 100 kcal and digestible carbohydrate providing 8 to 15 g per 100 kcal of the nutritional composition. In one embodiment, the lipid provides 3 to 7 g lipid per 100 kcal, preferably 4 to 6 g per 100 kcal, the protein provides 1 .6 to 2.1 g per 100 kcal, preferably 1 .6 to 2.0 g per 100 kcal and the digestible carbohydrate provides 5 to 20 g per 100 kcal, preferably 8 to 15 g per 100 kcal of the nutritional composition and wherein preferably the digestible carbohydrate component comprises at least 60 wt% lactose based on total digestible carbohydrate, more preferably at least 75 wt%, even more preferably at least 90 wt% lactose based on total digestible carbohydrate. The amount of total calories is determined by the sum of calories derived from protein, lipids, digestible carbohydrates and non- digestible oligosaccharide.

The present nutritional composition preferably comprises a digestible carbohydrate component. Preferred digestible carbohydrate components are lactose, glucose, sucrose, fructose, galactose, maltose, starch and maltodextrin. Lactose is the main digestible carbohydrate present in human milk. The present nutritional composition preferably comprises lactose. As the present nutritional composition comprises a fermented composition that is obtained by fermentation by lactic acid producing bacteria, the amount of lactose is reduced compared to its source due to the fermentation whereby lactose is converted into lactate and/or lactic acid. Therefore in the preparation of the present nutritional composition lactose is preferably added. Preferably the present nutritional composition does not comprise high amounts of carbohydrates otherthan lactose. Compared to digestible carbohydrates such as maltodextrin, sucrose, glucose, maltose and other digestible carbohydrates with a high glycemic index, lactose has a lower glycemic index and is therefore preferred. The present nutritional composition preferably comprises digestible carbohydrate, wherein at least 35 wt%, more preferably at least 50 wt%, more preferably at least 60 wt%, more preferably at least 75 wt%, even more preferably at least 90 wt%, most preferably at least 95 wt% of the digestible carbohydrate is lactose. Based on dry weight the present nutritional composition preferably comprises at least 25 wt% lactose, preferably at least 40 wt%, more preferably at least 50 wt% lactose.

The present nutritional composition preferably comprises at least one lipid selected from the group consisting of animal lipid (excluding human lipids) and vegetable lipids. Preferably the present composition comprises a combination of vegetable lipids and at least one oil selected from the group consisting of fish oil, animal oil, algae oil, fungal oil, and bacterial oil. The lipid of the present nutritional composition preferably provides 3 to 7 g per 100 kcal of the nutritional composition, preferably the lipid provides 4 to 6 g per 100 kcal. When in liquid form, e.g. as a ready-to-feed liquid, the nutritional composition preferably comprises 2.1 to 6.5 g lipid per 100 ml, more preferably 3.0 to 4.0 g per 100 ml. Based on dry weight the present nutritional composition preferably comprises 12.5 to 40 wt% lipid, more preferably 19 to 30 wt%. Preferably the lipid comprises the essential fatty acids alpha-linolenic acid (ALA), linoleic acid (LA) and/or long chain polyunsaturated fatty acids (LC-PUFA). The LC-PUFA, 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 or more of the above. Preferably the present nutritional composition comprises at least one, preferably at least two lipid sources selected from the group consisting of rape seed oil (such as colza oil, low erucic acid rape seed oil and canola oil), high oleic sunflower oil, high oleic safflower oil, olive oil, marine oils, microbial oils, coconut oil and palm kernel oil. The present nutritional composition is not human milk. The present nutritional composition preferably comprises protein. The protein used in the nutritional composition is preferably selected from the group consisting of non-human animal proteins, preferably milk proteins, vegetable proteins, such as preferably soy protein and/or rice protein, and mixtures thereof. The present nutritional composition preferably contains casein and/or whey protein, more preferably bovine whey proteins and/or bovine casein. Thus in one embodiment the protein in the present nutritional composition comprises protein selected from the group consisting of whey protein and casein, preferably whey protein and casein, preferably the whey protein and/or casein is from cow’s milk. The present nutritional composition preferably comprises casein and whey proteins in a weight ratio casein : whey protein of 10 : 90 to 90 : 10, more preferably 20 : 80 to 80 : 20, even more preferably 35 : 65 to 55 : 45.

Preferably the protein comprises less than 5 wt% based on total protein of free amino acids, dipeptides, tripeptides or hydrolysed protein.

Preferably the composition comprises at least 90 wt% intact cow’s milk proteins based on total protein. These proteins are superior in delivering the correct profile of essential amino acids that is needed to support the growth and development of an infant. Yet, the precocious maturation of the intestine is sometimes attributed to the presence of cow’s milk protein, but it was now found that in a fermented formula with such proteins this was prevented. Therefore, in one embodiment of the present invention, the nutritional composition for use or the nutritional composition in the methods and uses according to the invention is at least partly fermented by lactic acid producing bacteria and comprises at least 90 wt% intact cow’s milk proteins, based on total protein.

The wt% protein based on dry weight of the present nutritional composition is calculated according to the Kjeldahl-method by measuring total nitrogen and using a conversion factor of 6.38 in case of casein, or a conversion factor of 6.25 for other proteins than casein. The term‘protein’ or‘protein component’ as used in the present invention refers to the sum of proteins, peptides and free amino acids.

The present nutritional composition preferably comprises protein providing 1 .6 to 4.0 g protein per 100 kcal of the nutritional composition, preferably providing 1 .6 to 3.5 g, even more preferably 1 .75 to 2.5 g per 100 kcal of the nutritional composition. In one embodiment, the present nutritional composition comprises protein providing 1 .6 to 2.1 g protein per 100 kcal of the nutritional composition, preferably providing 1 .6 to 2.0 g, more preferably 1 .7 to 2.1 g, even more preferably 1 .75 to 2.0 g per 100 kcal of the nutritional composition. In one embodiment, the present nutritional composition comprises protein in an amount of less than 2.0 g per 100 kcal, preferably providing 1 .6 to 1 .9 g, even more preferably 1 .75 to 1 .85 g per 100 kcal of the nutritional composition. A too low protein content based on total calories will result in less adequate growth and development in infants. A too high amount will put a metabolic burden, e.g. on the kidneys of infants. When in liquid form, e.g. as a ready-to-feed liquid, the nutritional composition preferably comprises 0.5 to 6.0 g, more preferably 1 .0 to 3.0 g, even more preferably 1 .0 to 1 .5 g protein per 100 ml, most preferably 1 .0 to 1 .3 g protein per 100 ml. Based on dry weight the present nutritional composition preferably comprises 5 to 20 wt% protein, preferably at least 8 wt% protein based on dry weight of the total nutritional composition, more preferably 8 to 14 wt%, even more preferably 8 to 9.5 wt% protein based on dry weight of the total nutritional composition.

In order to meet the caloric requirements of an infant, the nutritional composition preferably comprises 45 to 200 kcal/100 ml liquid. For infants the nutritional composition has more preferably 60 to 90 kcal/100 ml liquid, even more preferably 65 to 75 kcal/100 ml liquid. This caloric density ensures an optimal ratio between water and calorie consumption. The osmolarity of the present composition is preferably between 150 and 420 mOsmol/l, more preferably 260 to 320 mOsmol/l. The low osmolarity aims to further reduce the gastrointestinal stress, which may affect intestinal maturation.

When the nutritional composition is in a ready to feed, liquid form, the preferred volume administered on a daily basis is in the range of about 80 to 2500 ml, more preferably about 200 to 1200 ml per day. Preferably, the number of feedings per day is between 1 and 10, preferably between 3 and 8. In one embodiment the nutritional composition is administered daily for a period of at least 2 days, preferably for a period of at least 4 weeks, preferably for a period of at least 8 weeks, more preferably for a period of at least 12 weeks, in a liquid form wherein the total volume administered daily is between 200 ml and 1200 ml and wherein the number of feedings per day is between 1 and 10.

The present nutritional composition, when in liquid form, preferably has a viscosity between 1 and 60 mPa.s, preferably between 1 and 20 mPa.s, more preferably between 1 and 10 mPa.s, most preferably between 1 and 6 mPa.s. The low viscosity ensures a proper administration of the liquid, e.g. a proper passage through the whole of a nipple. Also this viscosity closely resembles the viscosity of human milk. Furthermore, a low viscosity results in a normal gastric emptying and a better energy intake, which is essential for infants which need the energy for optimal growth and development. The present nutritional composition alternatively is in powder form, suitable for reconstitution with water to provide a ready to drink liquid. The present nutritional composition is preferably prepared by admixing a powdered composition with water. Normally infant formula is prepared in such a way. The nutritional composition for use or the nutritional composition in the uses and methods according to the invention thus can also be in the form of a packaged powder composition wherein said package is provided with instructions to admix the powder with a suitable amount of liquid, thereby resulting in a liquid composition with a viscosity between 1 and 60 mPa.s. The viscosity of the liquid is determined at a shear rate of 95 s^_1 at 20 °C. A suitable equipment to measure the viscosity is Physica Rheometer MCR 300 (Physica Messtechnik GmbH, Ostfilden, Germany). Application

In a first aspect the present invention relates to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants.

In the context of the present invention the term “more similar to the intestinal maturation pattern observed in human milk fed infants” is as compared to when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered. In other words, the invention relates to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants when the maturation pattern is compared to an infant that was administered a non-fermented infant formula or follow on formula.

In a second aspect the invention relates to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in preventing precocious maturation of the intestine in an infant. Maturation of the intestine is as compared to an infant that was administered a non-fermented infant formula or follow on formula. The invention thus also relates to a nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in preventing precocious maturation of the intestine in an infant, as compared to an infant that was administered a non-fermented infant formula or follow on formula.

In some jurisdictions the first aspect of the present invention can be worded as a method for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria. In these jurisdictions the second aspect of the present invention can be worded as a method for preventing precocious maturation of the intestine in an infant, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

The methods and uses according to the present invention as described above are considered to be therapeutic methods or uses. However, in some jurisdictions administering a nutritional composition to an infant is considered non-therapeutic. In these jurisdictions, the first aspect of the present invention can be worded as a non-therapeutic method for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In these jurisdictions the second aspect of the present invention can be worded as a non-therapeutic method for preventing precocious maturation of the intestine in an infant, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria.

In the nutritional composition for use according to the invention, and in the nutritional composition in the methods and uses according to the invention, the maturation pattern or maturation is compared to an infant that was administered a non-fermented infant formula or follow on formula.

The methods or uses according to the present invention, comprising administering the present nutritional composition to an infant, also refer to administering an effective amount of the nutritional composition to the infant. The nutritional composition according to the invention, and the nutritional composition in the methods and uses according to the invention, prevents precocious maturation of the barrier function and supports a morphological and functional intestinal maturation that is more similar as to mother fed.

The present invention may relate to the functional properties of the intestine, in particular the intestinal barrier function. It was found that the intestinal barrier function, as determined by the FITC-dextran assay, in mice fed the experimental formula which is at least partly fermented by lactic acid producing bacteria, was more similar to that of mother fed mice. This permeability was higher than in standard formula fed mice.

The present invention may also relate to the structural and/or morphological properties of the intestine, in particular the tissue development. It was found that the crypt-villus length in mice fed the experimental formula which is at least partly fermented by lactic acid producing bacteria, was more similar to that of mother fed mice. This length was shorter than in standard formula fed mice.

The present nutritional composition is for use in, and the present methods and uses are for, inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, and/or for use in preventing precocious maturation of the intestine in an infant. In one embodiment the maturation is the maturation of the intestinal barrier function. In another embodiment the maturation is the growth and morphological development of the intestine, in particular the intestinal crypt-villus length.

The effects on the intestinal maturation pattern or the preventing of precocious maturation are observed when compared to an infant that was administered a non-fermented infant formula or follow on formula.

The growth and morphological development of the intestine in human milk fed infants is lower than the growth and morphological development of the intestine of infants fed standard infant formula or follow on formula, i.e. a non-fermented formula. In the context of the present invention the term“more similar to the intestinal maturation pattern observed in human milk fed infants” means that the growth and morphological development of the intestine in infants fed the infant formula or follow on formula according to the invention is lower as compared to the growth and morphological development of the intestine when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered. More in particular, the intestinal crypt-villus length in human milk fed infants is less than the intestinal crypt-villus length of infants fed standard infant formula or follow on formula, i.e. a non- fermented formula. In the context of the present invention the term “more similar to the intestinal maturation pattern observed in human milk fed infants” means that the intestinal crypt-villus length in infants fed the infant formula or follow on formula according to the invention is lower as compared to the intestinal crypt-villus length when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered. Hence in one embodiment according to the invention, the use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing growth and morphological development of the intestine that is lower as compared to the growth and morphological development of the intestine of an infant that was administered a non- fermented infant formula or follow on formula.

Hence in one embodiment according to the invention, the use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing intestinal crypt-villus length that is lower as compared to the intestinal crypt-villus length of an infant that was administered a non-fermented infant formula or follow on formula.

The intestinal barrier function as determined by intestinal permeability in human milk fed infants is higher than the intestinal barrier function of infants fed standard infant formula or follow on formula, i.e. a non- fermented formula. In the context of the present invention the term “more similar to the intestinal maturation pattern observed in human milk fed infants” means that the intestinal barrier function in infants fed the infant formula or follow on formula according to the invention is higher as compared to the intestinal barrier function when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered. More in particular, the intestinal permeability in human milk fed infants is higher than the intestinal permeability of infants fed standard infant formula or follow on formula, i.e. a non- fermented formula. In the context of the present invention the term “more similar to the intestinal maturation pattern observed in human milk fed infants” means that the intestinal permeability in infants fed the infant formula or follow on formula according to the invention is higher as compared to the intestinal permeability when a standard infant formula or follow on formula, i.e. a non-fermented formula, is administered.

Hence in one embodiment according to the invention, the use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing intestinal barrier function that is higher as compared to the intestinal barrier function of an infant that was administered a non-fermented infant formula or follow on formula.

In one embodiment according to the invention, the use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing intestinal permeability that is higher as compared to the intestinal permeability of an infant that was administered a non-fermented infant formula or follow on formula.

Without wishing to be bound by theory, the inventors believe that an intact gut epithelial barrier is key in maintaining intestinal immune homeostasis and can be modulated by the gut microbiota. Moreover, environmental factors like stress and antibiotics negatively impact the gut barrier, which results into a proinflammatory immune response and can also lead to visceral hypersensitivity. But a precociously decreased intestinal permeability in infants could affect health long lastingly by altering immune maturation. It is thought that the elevated intestinal permeability in breast fed neonates facilitates the development of food tolerance and immune function by enabling the access of antigens to immune cells of the gut. However, a compromised intestinal barrier function, and hence increased intestinal permeability, at adulthood can be associated with adverse effects such as bacterial translocation and nerve endings sensitization contributing to visceral hypersensitivity development in rodents and human.

Therefore it is advantageous that under conditions of physical or psychological stress the intestinal barrier function was not impaired, but strengthened when partly fermented formula is administered. In a mouse model using partial restrained stress indeed the stress induced increase in gut permeability was prevented in mice administered the formula of the invention when compared to mice administered a standard formula.

Therefore, in an embodiment of the composition for use and the methods and uses according to the invention, increase of intestinal barrier permeability induced by physical or psychological stress is prevented.

The present invention is for use in an infant. An infant is a human with an age of 0 to 12 months. Preferably the use is for an infant of 0 to 6 months of age, more preferably 0 to 4 months of age. Although the maturation of the intestine takes place during entire infancy, the largest effects are in the first 6 month of life, and even more in the first 4 months. Preferably the infants are healthy infants. Preferably the infants are term infants. A term infants means an infant born at a gestational age of 37 to 42 weeks. More preferably the infant is a healthy term infant.

EXAMPLES

Example 1: Partly fermented infant formula prevents precocious decrease in gut permeability

Mice & experimental design:

Experimental groups were obtained from C57BL/6 dams and housed in a controlled environment (12- h/12-h light/dark cycle, humidity, and temperature) with ad libitum access to food and water. Dams were fed grain-based standard rodent chow (RM3; Special Diets Services). After birth, litters were left undisturbed with the dams, and litter size was 6-8 pups/litter. At postnatal day (PN) 14.5, mouse pups were separated from the dams and randomized to one of the intervention groups receiving different infant formulas (IF) as diet ad libitum until the end of the study (PN 18.5). During the intervention period, 4-5 mouse pups were housed together. IF diets were prepared from IF powders and water as detailed below and were provided to pups in liquid form in dishes on the cage floor to allow easy access. Dishes were cleaned and IF diets were refreshed every 3 hours to ensure adequate and unsoiled availability of the diets. Randomized, age-matched, non-separated mother-fed pups (MF) served as reference group. Mice were weighed on a daily basis after PN 14.5, and at the end of the intervention period, at day PN 18.5, morphological and functional parameters for gut maturation were assessed. A total of n=35 mouse pups were included in this study (9 for each formula group and 8 for the mother fed mice). Experimental diets:

IF powders were prepared with water as specified below and preparations were provided to pups in liquid form in dishes on the cage floor ad libitum.

Infant formula 1 (G/F) was an infant formula supplemented with non-digestible oligosaccharides (scGos/lcFOS, 9:1 w/w). The infant formula comprised 5.7 g non-digestible oligosaccharides per 100 g dry weight As a source of short chain GOS (scGOS) Vivinal GOS (Friesland Campina, Borculo, Netherlands) was used and as a source of long chain FOS (IcFOS) RaftilineHP (Orafti, Tiense Suikerraffinaderij N.V., Tienen, Belgium) was used.

Infant formula 2 (FM+G/F) was a partly fermented infant formula with non-digestible oligosaccharides (scGos/lcFOS, 9:1 w/w). About 26 wt% of the formula dry weight consisted of fermented infant formula (as a source Lactofidus 1). About 0.3 wt% L-lactic acid was present based on dry weight. The infant formula comprised 5.7 g non-digestible oligosaccharides per 100 g dry weight As a source of short chain GOS (scGOS) Vivinal GOS (Friesland Campina, Borculo, Netherlands) was used and as a source of long chain FOS (IcFOS) RaftilineHP (Orafti, Tiense Suikerraffinaderij N.V., Tienen, Belgium) was used. Infant formula 3 (CTRL) was a control infant formula that was not fermented and did not comprise added non-digestible oligosaccharides.

The three formula were equal in energy content and similar in content of protein (bovine whey and casein), digestible carbohydrates, and fat.

Experimental procedures:

Histology and immunohistochemistry to assess tissue morphology / architecture: haematoxylin staining was performed on six-micron-thick paraffin embedded sections. Crypt-villus length were measured in 5 to 10 well-oriented crypts and villi per section from three small intestinal segments per animal.

Gut permeability was measured by FITC-Dextran Permeability Assay: Intestinal permeability was assessed by luminal enteral administration of FITC-dextran 4000 (Sigma), a nonmetabolizable macromolecule that is used as a permeability probe. Mice were gavaged with FITC-dextran (40 mg/100 g body weight) 4 h before sacrificing. Whole blood was obtained by cardiac puncture at the time of sacrificing, and FITC-dextran measurements were performed in duplicate by fluorometry. Dilutions of FITC-dextran in PBS were used as a standard curve, and absorption of 100 mI_ of serum or standard was measured in a fluorometer at 488 nm.

Results:

Small intestinal architecture / morphology:

Crypt-villus length in small intestine of CTRL and G/F mice was significantly greater than in MF mice (p=0.008 and p=0.039, respectively), while FM+G/F mice did not show a significantly different crypt length compared to the MF mice, see Table 1). Ordinary one-way ANOVA followed by Sidak’s multiple comparisons test was performed, as data of groups G/F (n=5), FM+G/F (n=5), CTRL (n=5) and MF (n=5) passed KS normality test; data shown are mean ± SEM. Table 1 : Effect of diet on intestinal crypt-villus length in 18.5 days old mice

*p<0.05 compared to MF, ** p< 0.01 compared to MF

Gut barrier function:

At the end of the intervention period the intestinal permeability was similar in FM+G/F and MF mice. Gut permeability was significantly lower in mice receiving CTRL or G/F IF compared to both FM+G/F and MF mice (p=0.028 and p=0.032 respectively), as is shown in Table 2. Ordinary one-way ANOVA followed by unpaired, two-tailed t-tests between selected groups were performed, as data of groups G/F (n=8), FM+G/F (n=8), CTRL (n=8) and MF (n=7) passed KS normality test; data shown are mean ± SEM.

Table 2: Effect of diet on gut permeability in 18.5 day old mice

*p<0.05 compared with CTRL

These results support the notion that infant formula (IF) per se can induce morphological overgrowth and precocious functional gut maturation. Precociously decreased permeability as seen in CTRL IF could affect health long lastingly by altering immune maturation. However, IF-induced effects on gut growth and architecture, as well as permeability, seem to be abolished in FM+G/F mice, rendering the gut maturation and its timing in mice fed the nutritional composition of the present invention more similar to mother’s milk fed mice. This is indicative for an effect of administering at least partly fermented infant formula on the prevention of precocious maturation of the intestine, and inducing, an intestinal maturation pattern which is more similar to the intestinal maturation pattern observed in mother fed. Preferably the infant formula also comprises non-digestible oligosaccharides, more preferably a mixture of galacto-oligosaccharides and fructo-oligosaccharides.

Example 2: A partly fermented infant milk formula reduces stress-induced gut permeability in rats The impact of a partly fermented formula with non-digestible oligosaccharides (scGOS/lcFOS 9:1 w/w) on gut permeability as such and on hyper-permeability induced by partial restraint stress (PRS), when compared to NaCI (saline) control was assessed. This investigation was conducted on 4 groups of 10 female Wistar rats with the method as described in Agostini et al., Neurogastroenterol Motil. 2012; 24:376-172.

Group 1 and 2 were saline control (oral gavage 1 mL/day) for 14 days (day 1 to 14).

Group 3 and 4 were administered partly fermented formula with non-digestible oligosaccharides, similar to infant formula 2 of example 1 (oral gavage 1 ml/day) for 14 days. The protein content was 2.0 g/100 kcal (intact bovine whey protein and casein).

Group 1 and 2 were not exposed to PRS.

Group 3 and 4 were exposed to PRS.

All stress sessions were performed at the same time of the day to minimize any influence of circadian rhythms. Stress effects were studied using a 2 h session of PRS considered as a mild non-ulcerogenic model. Under mild anesthesia, upper forelimbs were taped to the thoracic trunk in order to constrain animal body movements. Rats were replaced in their home cages for 2 h.

Gut paracellular permeability was evaluated using ⁵¹Cr-EDTA (Perkin Elmer Life Sciences, Paris, France). The rats were daily administered with the FM + G/F or saline diets for a period of 14 days. At the end of administration (day 15) animals received an oral administration of ⁵¹Cr-EDTA and, immediately they were submitted or not to PRS. ⁵¹Cr-EDTA was diluted in 0.5 ml of saline and administered by gavage. Rats were then placed in metallic cages and radioactivity in urine (the total of 24 h) was measured with a gamma counter. Permeability was expressed as the percentage in urine of total ⁵¹Cr-EDTA administered.

PRS induced intestinal hyperpermeability in rats having consumed saline was substantial. It was found that in the rats having consumed FM+G/F this PRS-induced intestinal hyperpermeability was prevented. Administration of FM+G/F thus limits the increase in gut permeability that is associated with acute stress in rats, see Table 3. These data suggest that the administration of FM+G/F does not impact gut barrier permeability under normal, healthy conditions, but prevents the intestinal epithelial barrier disruption caused by stress.

Table 3: Effect of PRS and a partly fermented formula with non-digestible oligosaccharides on the permeability of the intestine in rats.

*p < 0.05 compared to saline without PRS

These results are thus indicative for the protective effect that FM + G/F has on maintaining the gut permeability. Furthermore, the administration of the FM + G/F formula does not increase the gut permeability as such when compared to control. This is an indication that the increased gut permeability as observed in example 1 , and which was more similar to the mother fed situation, is due to improved intestinal maturation kinetics and prevention of precocious closure of the gut barrier, and not due to an intestinal barrier permeability increasing effect as such.

This is indicative for an effect of administering at least partly fermented infant formula for the prevention of an increase of intestinal barrier permeability induced by physical or psychological stress. Preferably the infant formula also comprises non-digestible oligosaccharides, more preferably a mixture of galacto- oligosaccharides and fructo-oligosaccharides.

Claims

1 . A nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto-oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto-oligosaccharides, gluco-oligosaccharides, gentio- oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan- oligosaccharides, isomalto-oligosaccharides, nigero-oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.

2. A nutritional composition selected from an infant formula and a follow on formula, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, for use in preventing precocious maturation of the intestine in an infant, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition, and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto-oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto-oligosaccharides, gluco-oligosaccharides, gentio- oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan- oligosaccharides, isomalto-oligosaccharides, nigero-oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.

3. The nutritional composition for use according to claim 1 or 2, wherein the maturation is the maturation of the intestinal barrier function.

4. The nutritional composition for use according to claim 1 or 2, wherein the maturation is the growth and morphological development of the intestine, in particular the intestinal crypt-villus length.

5. The nutritional composition for use according to any one of the preceding claims, wherein the maturation pattern or maturation is compared to an infant that was administered a non-fermented infant formula or follow on formula.

6. The nutritional composition for use according to any one of the preceding claims, wherein the infant has an age of 0 to 4 months.

7. The nutritional composition for use according to any one of the preceding claims, wherein the infant is a term infant.

8. The nutritional composition for use according to any one of the preceding claims, wherein increase of intestinal barrier permeability induced by physical or psychological stress is prevented.

9. The nutritional composition for use according to any one of the preceding claims, wherein the nutritional composition comprises lactic acid producing bacteria, preferably selected from the group consisting of Bifidobacterium and Streptococcus, more preferably S. thermophilus and/or B. breve, most preferably S. thermophilus and B. breve.

10. The nutritional composition for use according to any one of the preceding claims, wherein the sum of L-lactic acid and L-lactate is 50 wt% or more, based on the sum of total lactic acid and lactate.

1 1 . The nutritional composition for use according to any one of the preceding claims, wherein the non- digestible oligosaccharides are selected from the group consisting of galacto-oligosaccharides and fructo-oligosaccharides, preferably the non-digestible oligosaccharides are galacto- oligosaccharides and fructo-oligosaccharides.

12. The nutritional composition for use according to any one of the preceding claims, wherein inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing intestinal crypt-villus length that is lower as compared to the intestinal crypt-villus length of an infant that was administered a non-fermented infant formula or follow on formula.

13. The nutritional composition for use according to any one of the preceding claims, wherein inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, is inducing intestinal permeability that is higher as compared to the intestinal permeability of an infant that was administered a non-fermented infant formula or follow on formula.

14. The nutritional composition for use according to any one of the preceding claims, wherein the nutritional composition comprises at least 90 wt% intact cow’s milk proteins, based on total protein.

15. The nutritional composition for use according to any one of the preceding claims, wherein the nutritional composition comprises 1 .7 to 2.1 g protein per 100 kcal, preferably 1 .75 to 2.0 g protein per 100 kcal.

16. The nutritional composition for use according to any one of the preceding claims, wherein the nutritional composition is a powder, suitable to reconstitute with water to provide a ready to drink formula.

17. Use of a fermented composition for the manufacture of a nutritional composition selected from an infant formula and a follow on formula for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto-oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto-oligosaccharides, gluco- oligosaccharides, gentio-oligosaccharides, glucomanno-oligosaccharides, galactomanno- oligosaccharides, mannan-oligosaccharides, isomalto-oligosaccharides, nigero-oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.

18. Use of a fermented composition for the manufacture of a nutritional composition selected from an infant formula and a follow on formula for preventing precocious maturation of the intestine in an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto- oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto- oligosaccharides, gluco-oligosaccharides, gentio-oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan-oligosaccharides, isomalto-oligosaccharides, nigero- oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.

19. A method for inducing an intestinal maturation pattern in an infant which is more similar to the intestinal maturation pattern observed in human milk fed infants, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto- oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto- oligosaccharides, gluco-oligosaccharides, gentio-oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan-oligosaccharides, isomalto-oligosaccharides, nigero- oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.

20. A method for preventing precocious maturation of the intestine in an infant, the method comprising administering a nutritional composition selected from an infant formula and a follow on formula to an infant, wherein the nutritional composition is at least partly fermented by lactic acid producing bacteria, wherein the infant has an age of 0 to 6 months and wherein the nutritional composition comprises 0.1 to 1 .5 wt% of the sum of lactic acid and lactate, based on dry weight of the nutritional composition and wherein the nutritional composition comprises 2.5 to 15 wt% non-digestible oligosaccharides, based on dry weight of the formula, wherein the non-digestible oligosaccharides are selected from the group consisting of fructo-oligosaccharides, non-digestible dextrins, galacto- oligosaccharides, xylo-oligosaccharides, arabino-oligosaccharides, arabinogalacto- oligosaccharides, gluco-oligosaccharides, gentio-oligosaccharides, glucomanno-oligosaccharides, galactomanno-oligosaccharides, mannan-oligosaccharides, isomalto-oligosaccharides, nigero- oligosaccharides, chito-oligosaccharides, soy oligosaccharides, uronic acid oligosaccharides, sialyloligosaccharides and fuco-oligosaccharides, and mixtures thereof.