RU2789158C2 - Compositions containing at least one n-acetylated and at least one fucosylated oligosaccharide for use in activation of digesting ability in infants and young children - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: group of inventions relates to activation of a digesting ability in an infant or a young child. The use of an artificial nutritional composition containing at least one N-acetylated oligosaccharide and at least one fucosylated oligosaccharide in a method for activation of a digesting ability in an infant or a young child is disclosed. The use of a milk formula for children aged 1 to 3 in a method for activation of a digesting ability in a child is also disclosed.

EFFECT: group of inventions provides an increase in an ability to cleave macronutrients, such as lipids, fat-soluble vitamins, proteins, and/or carbohydrates, in infants and young children.

16 cl, 1 dwg, 2 tbl, 3 ex

Description

Scope of the invention

The present invention relates to compositions for use in activating the development and/or increasing the surface area of the small intestine and consequently improving the digestive capacity of an infant or young child.

Prerequisites for the creation of the invention

The development of the intestines in infants and young children is of key importance for the development of the infant and child, in particular for an infant or young child, optimal digestive capacity is very important. Thus, it is desirable to develop compositions that promote the development of the digestive capacity of infants and young children.

Prior to absorption by the body, proteins and carbohydrates are first split in the intestinal lumen into smaller fragments, after which they are further split into amino acids, di- and tripeptides and monosaccharides under the action of various enzymes located on the apical part of the enterocytes of the small intestine.

Like proteins and carbohydrates, dietary lipids and fat-soluble vitamins in the intestinal lumen undergo hydrolysis by various types of lipases. After the first stage of digestion in the stomach under the action of gastric juice lipase, large drops of undigested fat and fat breakdown products enter the intestine, where they are mixed with bile salts to form a fine emulsion that is easily affected by pancreatic lipases.

Further, lipid breakdown products are introduced into the micelles of bile salts in the intestinal lumen and form the so-called mixed micelles. This process significantly increases the solubility of lipid breakdown products, which, in turn, greatly facilitates their absorption by enterocytes. The formation of bile salt micelles in the intestinal lumen requires a minimum concentration of bile salts, referred to as the critical micellar concentration. In healthy adults, the concentration of bile salts in the intestinal lumen in the postprandial state exceeds the critical micellar concentration, which ensures highly efficient breakdown and absorption of lipids. In infants, particularly preterm infants (see Lindqvist et al., Lipid digestion and absorption in early life: an update, Current Opinion in Clinical Nutrition and Metabolic Care; 13(2010): 314–320), and in some pathological conditions (for example, cystic fibrosis), the concentration of bile salts in the intestinal lumen is low, below the critical micellar concentration, which leads to inefficient breakdown and absorption of lipids.

The main consequence of this inefficient lipid breakdown is the loss of nutrients in the faeces, namely the loss of fat-soluble vitamins and fat, such as essential fatty acids and long-chain polyunsaturated fatty acids (LCPUFA), as well as the loss of associated energy. As a result, these nutrients and energy cannot be used for the growth and development of the infant. Additionally, such inefficient digestion is associated, for example, with diarrhea, food intolerance and changes in the intestinal microbiota. Therefore, it is particularly desirable to find solutions to improve digestion in infants and young children, and in particular in preterm or low birth weight infants.

In healthy adults, bile salts in the intestinal lumen travel along the small intestine and are efficiently absorbed (up to 95%) in the ileum. Free bile salts are returned via the portal vein to the liver, where they are stored for re-secretion into the intestinal lumen. Thus, the major part of the pool of bile salts in the intestinal lumen depends on this enterohepatic circulation, and only a small part is formed as a result of de novo synthesis. However, in infants during the first months of life, the enterohepatic circulation process is incomplete (see Belle et al.; Intestinal absorption of bile salts: immature development in the neonate; J Pediatr 94 (1979): 472–476), especially in preterm infants and in certain pathological conditions like short bowel syndrome. In particular, an increase in the absorptive surface of the ileum allows for improved enterohepatic circulation of bile salts, their reuse in the liver, and an increase in the pool of bile salts in the intestinal lumen, which will provide better breakdown of fats and fat-soluble vitamins.

The small intestine plays a leading role in this digestion process. One of the important parameters affecting the digestive capacity is the morphology of the small intestine. The key parameter is the surface area of the small intestine epithelium (see Helander et al.; Surface area of the digestive tract – revisited; Scandinavian Journal of Gastroenterology; 49 (2014): 681–689). The greater the surface area of the epithelium, the greater the pool of bile salts and digestive enzymes available for the ultimate breakdown of nutrients such as lipids, fat-soluble vitamins, carbohydrates, and proteins. An increase in the length of the villi in the small intestine provides a beneficial increase in surface area and a corresponding improved breakdown of macronutrients such as lipids, fat-soluble vitamins, proteins and carbohydrates. This effect of the length of the villi and the corresponding increase in the surface area of the small intestine on the digestive capacity is well described in the literature. For example, L. Montagne et al., A review of interactions between dietary fiber and the intestinal mucosa, and their consequences on digestive health in young non-ruminant animals, Animal Feed Science and Technology 108 (2003) p.95–117 , and in particular in paragraph 4.3 on page 105, several studies are listed that describe the positive effect of increasing the length of the villi on the digestive capacity of animals.

Improving digestion is desirable for all infants and young children. However, certain specific groups of infants and young children are in particular need of compositions capable of activating their development in general and in particular their digestive and/or absorptive capacity. Such infants and young children are, for example, premature infants, low birth weight infants, infants and young children suffering from short bowel syndrome, and/or infants or young children with stunted growth, and in particular, infants with intrauterine growth retardation (IUGR).

Many attempts have been made to improve the digestive capacity of infants and young children, as well as young animals. In particular, several studies have evaluated the effects of different fiber types on intestinal morphology and on the digestive capacity of young animals, infants or young children. In the above review, L. Montagne et al. described a variety of studies demonstrating the variable effect of different types of dietary fiber, some of which were associated with a positive effect on digestion, while others gave the impression of being harmful.

One example of a study in which dietary fiber has shown beneficial effects is P. Liu and al, Effects of chito-oligosaccharide supplementation on the growth performance, nutrient digestibility, intestinal morphology, and fecal shedding of Escherichia coli and Lactobacillus in weaning pigs, J .anim. sci. 86 (2008), p. 2609–2618. In this study, an increase in villus height and villus:crypt ratio in the ileum and jejunum was associated with improved digestibility in weaned piglets when chitooligosaccharides were added to the piglets' diet compared to control diets. In contrast, in a study by van Meer et al.; Prebiotic oligosaccharides and the enterohepatic circulation of bile salts in rats; Am J Physiol Gastrointest Liver Physiol 294 (2008): G540 G547 showed that the addition of galactooligosaccharides to the diet of baby rats did not increase the pool of bile salts or change the kinetics of the pool of bile salts and, therefore, did not provide activation of fat breakdown.

Other ingredients were evaluated for their ability to activate cleavage. For example, WO 2011/65552 describes a composition containing a protein hydrolyzate from a fermented milk product (eg from cottage cheese), lipids and isomaltulose, which effectively increased the length of the villi in the small intestine. However, such compositions are not suitable for infants and young children because isomaltulose is undesirable for use in such compositions.

Of particular advantage would be the identification of ingredients specifically selected for human infants and/or young children and capable of improving the digestive capacity of such infants and/or young children.

Mother's milk is recommended for all babies. However, in some cases, breastfeeding is insufficient or ineffective for medical reasons, or the mother decides not to breastfeed. Infant formulas have been developed for these situations. Fortifiers have also been developed to fortify breast milk or infant formula with certain ingredients.

There is a clear need to develop acceptable ways to increase the ability to break down macronutrients such as lipids, fat soluble vitamins, proteins and/or carbohydrates in infants and young children.

There is also a need to provide such health benefits in a manner that is particularly suitable for younger subjects (infants and young children), the method need not include classical pharmaceutical intervention as these infants or young children are particularly vulnerable.

There is a need to provide these infants or young children with such health benefits in a manner that does not cause side effects and/or that can be easily provided and that is acceptable to parents or healthcare professionals.

There is also a need to provide such benefits in a manner that will keep the cost of such provision reasonable and affordable to the majority.

Thus, there is a clear need to develop alternative methods other than classical pharmaceutical intervention, such as the use of antibiotics, at least because of the associated risk of developing antibiotic resistance.

Statement of the Invention

The present inventors have found that a composition comprising at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide can advantageously be used to activate digestion, preferably the ability to break down fat, fat-soluble vitamins, proteins and carbohydrates, in an infant or toddler. age. Although the authors do not wish to be bound by any theory, it is believed that such oligosaccharides have a synergistic effect in these specific health conditions.

Accordingly, the present invention provides a nutritional composition comprising at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide for use in a method of promoting digestion in an infant or young child.

The present invention also provides a 1 to 3 year old infant formula comprising at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide for use in a method for activating a child's digestive capacity.

Nutritional composition, respectively milk formula for children from 1 to 3 years, in particular, is effective for use in the method

- improving the enterohepatic circulation, in particular the enterohepatic circulation of bile salts, and

- stimulation of an increase in surface area in the small intestine; And

- stimulation of the increase in the length of the villi in the small intestine

infant or young child, respectively child.

In a particularly advantageous embodiment, the nutritional composition or infant formula for children from 1 to 3 years of age contains 2'-fucosyllactose (2-FL) and lacto-N-neotetraose (LNnT).

Brief description of graphic materials

In FIG. 1 shows the length of villi in the following groups of rat pups:

- in animals with normal intrauterine growth (normal group; positive control);

- in animals that suffered intrauterine growth retardation and received a normal diet (IUGR group; negative control);

- in animals with intrauterine growth retardation, which were kept on a diet with the addition of 2-FL (experimental group IUGR/2FL);

- in animals with intrauterine growth retardation, which were kept on a diet with the addition of LNnT (experimental group IUGR/LNnT);

- in animals with intrauterine growth retardation, which were kept on a diet with the addition of a combination of 2-FL and LNnT (experimental group IUGR/mixture of breast milk oligosaccharides (HMO)).

The average length of the villi in each group of rats is represented by a horizontal line. The 2-FL and LNnT groups showed no significant differences from the IUGR group. The 2-FL and LNnT group, in contrast, showed significantly longer villi than the other groups. Thus, synergy between 2-FL and LNnT can be observed.

Detailed description of the invention

In this document, the following terms have the following definitions.

The term "infant" means a child under the age of 12 months. The term "young child" means a child between the ages of one and less than three years (also called a toddler). The expression "child" means between the ages of three and seven years.

"Infant or young child born by caesarean section" means an infant or young child born by caesarean section. This means that the baby or young child was not born through a vaginal delivery.

"Infant or young child born by vaginal delivery" means an infant or young child who was born by vaginal delivery and not by caesarean section.

"Premature" or "premature" means an infant or young child who is born prematurely. Essentially, this term refers to an infant or young child born before the 37th week of pregnancy.

“Low birth weight infant” means a newborn weighing less than 2500 g (5.5 lb) due to either preterm delivery or limited fetal growth. Thus, this term includes:

- an infant or young child who has/had a birth weight between 1500 and 2500 g (commonly referred to as "low birth weight" or LBW);

- an infant or young child who has/had a birth weight between 1000 and 1500 g (commonly referred to as "very low birth weight" or VLBW);

- an infant or young child who has/had a birth weight of less than 1000 g (called "extremely low birth weight" or LUBR).

"Infant born small for gestational age (SGA)" means a baby with birth weight less than the 10th percentile for babies of the same gestational age.

The expression "nutritional composition" means the composition that feeds the subject. This nutritional composition is usually intended for oral or intravenous use and usually includes a lipid or fat source and a protein source.

In a specific embodiment, the composition of the present invention is a hypoallergenic nutritional composition. The expression "hypoallergenic nutritional composition" means a nutritional composition that is unlikely to cause allergic reactions.

In a specific embodiment, the composition of the present invention is an "artificial nutritional composition". The expression "artificial nutritional composition" means a mixture obtained by chemical and/or biological means, which may be chemically identical to the naturally occurring mixture present in mammalian milk (i.e., the artificial composition is not breast milk).

The expression "infant formula" as used herein refers to a food product that is specifically intended to be eaten by infants during the first months of life and that by itself satisfies the nutritional needs of this category of persons (Article 2(c) of European Commission Directive 91/ 321/EEC 2006/141/EC of 22 December 2006 on infant and complementary formula). It also refers to a nutritional composition intended for infants as defined in the Codex Alimentarius (Codex STAN 72-1981) and Infant Specialties (including the entry Food for Special Medical Purpose). The expression "infant formula" covers both "initial formula" and "advance-level formula" or "pre-fed formula".

“Follow-up formula” or “pre-fed formula” is given starting at 6 months. It is the main liquid element in the gradually increasing variety of diets for this category of people.

The expression "infant food" means a food product specially intended for consumption by infants or young children during the first years of life.

The expression "children's cereal-based composition" means a food product specifically intended for consumption by infants or young children during the first years of life.

The expression "milk formula for children 1 to 3 years old" (or GUM) refers to a milk-based drink, usually with added vitamins and minerals, that is intended for young children or children.

The term "fortifier" refers to liquid or solid nutritional compositions suitable for mixing with breast milk or infant formula.

The expression "weaning period" means the period during which the diet of an infant or young child replaces mother's milk with another food.

The expressions "aged days/weeks/months/years" and "days/weeks/months/years after birth" can be used interchangeably.

The expression "activation of the digestive capacity" includes one or more of the following:

- an increase in the surface area of the small intestine, and in particular the ileum;

- an increase in the length of the villi in the small intestine, in particular in the ileum; And

- improvement of enterohepatic circulation, in particular enterohepatic circulation of bile salts.

By "mother's milk" is meant the mother's breast milk or colostrum.

An "oligosaccharide" is a saccharide polymer containing a small number (typically three to ten) of simple sugars (monosaccharides).

The term "HMO" refers to the oligosaccharide(s) of breast milk. These carbohydrates are highly resistant to enzymatic hydrolysis, indicating that they may have significant functions not directly related to their calorie content. First of all, they have been shown to play a vital role in the early development of infants and young children, such as the development of the immune system. Many different types of HMO have been found in breast milk. Each individual oligosaccharide is based on a combination of glucose, galactose, sialic acid (N-acetylneuraminic acid), fucose, and/or N-acetylglucosamine, forming numerous and varied bonds between them, resulting in the presence of a huge number of different oligosaccharides in breast milk - at the moment it was more than 130 such structures have been identified. Nearly all of them have a lactose moiety at their reducing end, while sialic acid and/or fucose (if present) end up at the non-reducing ends. HMOs can be acidic (eg, charged sialic acid-containing oligosaccharide) or neutral (eg, fucosylated oligosaccharide).

"Fucosylated oligosaccharide" is an oligosaccharide having a fucose residue. It has a neutral character. Some examples are 2-FL (2'-fucosyllactose), 3-FL (3-fucosyllactose), difucosyllactose, lacto-N-fucopentaose (e.g. lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto-N- fucopentaose III, lacto-N-fucopentaose V), lacto-N-fucohexaose, lacto-N-difucohexaose I, fucosyl lacto-N-hexaose, fucosyllacto-N-neohexaose, difucosyl lacto-N-hexaose I, difucosyl lacto-N-neohexaose II and any combination of them. Although the authors do not wish to be bound by any theory, it is believed that the fucosyl epitope of fucosylated oligosaccharides may act as a "trap" on the mucosal surface. Through a competitive effect, it can prevent and/or limit the action of infection-causing pathogens (of viral or bacterial origin) or of their secreted components (e.g., toxins), primarily by eliminating their binding to natural ligands, and, without regard to any theory , it is believed that this will reduce the risk of infections/inflammations, and in particular the risk of infections and/or inflammations of the lower respiratory tract (LRT)/ear. Additionally, it is believed that fucosylated oligosaccharides stimulate the growth and metabolic activity of specific symbiotic microbes that reduce the inflammatory response and create an unfavorable environment for pathogens, thereby providing colonization resistance.

The expressions "fucosylated oligosaccharides containing a 2'-fucosyl epitope" and "2-fucosylated oligosaccharides" cover fucosylated oligosaccharides with a certain homology of form, since they contain a 2'-fucosyl epitope, therefore, a certain homology of function can be expected. Although the authors do not wish to be bound by any theory, it is believed that the 2'-fucosyl epitope of these fucosylated oligosaccharides is particularly characteristic of pathogens (or components secreted by them) that cause LRT and/or ear infections.

The term "N-acetylated oligosaccharide(s)" encompasses both "N-acetyllactosamine" and "oligosaccharide(s) containing N-acetyllactosamine". They are neutral oligosaccharides having an N-acetyllactosamine residue. Suitable examples are lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT), para-lacto-N-neohexaose (para-LNnH), disialyl lacto-N-neotetraose (DSLNT), or any combination thereof. Other examples are lacto-N-hexaose, lacto-N-neohexaose, para-lacto-N-hexaose, para-lacto-N-neohexaose, lacto-N-octaose, lacto-N-neo-octaose, iso-lacto-N-octaose , para-lacto-N-octaose and lacto-N-dekaose.

The terms "at least one fucosylated oligosaccharide" and "at least one N-acetylated oligosaccharide" mean "at least one type of fucosylated oligosaccharide" and "at least one type of N-acetylated oligosaccharide".

"HMO precursor" is a key compound that is included in the production of HMO, such as sialic acid and/or fucose.

A "sialylated oligosaccharide" is an oligosaccharide containing a charged sialic acid, i. e. an oligosaccharide having a sialic acid residue. It has an acidic character. Some examples are 3-SL (3'-sialyllactose) and 6-SL (6'-sialyllactose).

The nutritional composition of the present invention may be in solid form (eg powder) or liquid form. The amount of various ingredients (eg, oligosaccharides) may be expressed as g/100 g of the composition on a dry weight basis if presented in solid form, such as powder, or as a concentration in g/l of the composition if it is in liquid form ( the latter case also includes a liquid composition that can be made from a powder after being dissolved in a liquid, e.g. milk, water, etc., e.g. up to 3 years, or baby food cereal, or any other formula intended for baby food).

The term "prebiotic" refers to non-digestible carbohydrates that benefit the host by selectively stimulating the growth and/or activity of beneficial bacteria, such as bifidobacteria, in the human colon (Gibson GR, Roberfroid MB. Dietary modulation of the human colonic microbiota: introducing the concept of prebiotics J Nutr 1995;125:1401–12).

The term "probiotic" means preparations of microbial cells or components of microbial cells that have a beneficial effect on the health or well-being of the host organism. (Salminen S, Ouwehand A. Benno Y. et al. "Probiotics: how should they be defined" Trends Food Sci. Technol. 1999:10 107–10.) Microbial cells are essentially bacteria or yeasts.

The term "CFU" should be understood as a colony-forming unit.

Unless otherwise indicated, all percentages are by weight.

Additionally, in the context of the invention, the terms "comprising" or "comprises" do not exclude other possible elements. The composition of the present invention, including many of the embodiments described herein, may contain the essential elements and limitations of the invention described herein, as well as any additional or optional ingredients, components or limitations described herein or otherwise, depending on needs, consist or essentially consist of them.

Any reference to prior art documents in this specification should not be construed as an admission that such prior art is widely known or forms part of the common knowledge in the art.

A more detailed description of the invention will be presented below. It should be noted that various aspects, features, examples and embodiments described in this application may be compatible and/or combined with each other.

Thus, the first object of the present invention is a nutritional composition containing at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, for use in promoting digestion in an infant or young child.

The second object of the present invention is a milk formula for children from 1 to 3 years, containing at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, for use in activating the digestive capacity of the child.

Preferably, the nutritional composition or infant formula for 1 to 3 years of the present invention is for use in activating the ability to digest fat, fat soluble vitamins, proteins and carbohydrates in an infant or toddler, respectively, in a child.

In order to achieve such advantages, the nutritional composition, respectively milk formula for children from 1 to 3 years old, of the invention is preferably intended for use in the method

- increasing the surface area of the small intestine, preferably the ileum;

- increase in the length of the villi in the small intestine, preferably in the ileum; And

- improvement of enterohepatic circulation, in particular enterohepatic circulation of bile salts;

in an infant or young child, respectively, in a child.

The nutritional composition or infant formula for children from 1 to 3 years of the present invention contains at least one fucosylated oligosaccharide. It may contain one or more types of fucosylated(s) oligosaccharide(s). The fucosylated oligosaccharide(s) may in fact be selected from a list including 2'-fucosyllactose, 3'-fucosyllactose, difucosyllactose, lacto-N-fucopentaose (e.g., lacto-N-fucopentaose I, lacto-N- fucopentaose II, lacto-N-fucopentaose III, lacto-N-fucopentaose V), lacto-N-fucohexaose, lacto-N-difucohexaose I, fucosyllacto-N-hexaose, fucosyllacto-N-neohexaose (e.g. fucosyllacto-N-neohexaose I, fucosyllacto-N-neohexaose II), difucosyl lacto-N-hexaose I, difuco-lacto-N-neohexaose, difucosylacto-N-neohexaose I, difucosyl lacto-N-neohexaose II, fucosyl-para-lacto-N-hexaose, three -fuco-para-lacto-N-hexaose I and any combination thereof.

In some specific embodiments, the fucosylated oligosaccharide comprises a 2'-fucosyl epitope. It may, for example, be selected from a list including 2'-fucosyllactose, difucosyllactose, lacto-N-fucopentaose, lacto-N-fucohexaose, lacto-N-difucohexaose, fucosyllacto-N-hexaose, fucosyllacto-N-neohexaose, difucosyllacto- N-hexaose, difuco-lacto-N-neohexaose, difucosyllacto-N-neohexaose, fucosyl-para-lacto-N-hexaose, and any combination thereof.

In a preferred embodiment, the nutritional composition, respectively milk formula for children from 1 to 3 years, in accordance with the invention contains 2'-fucosyllactose (or 2FL, or 2'FL, or 2-FL, or 2'-FL). In a particular embodiment, no other type of fucosylated oligosaccharide other than 2'-fucosyllactose is used, i. e. nutritional composition or milk formula for children from 1 to 3 years of the invention contains only 2'-fucosyllactose as a fucosylated oligosaccharide.

The fucosylated oligosaccharide(s) may be isolated from a natural source, such as animal milk, by chromatography or filtration methods. In an alternative embodiment, it(them) can be produced by biotechnological means using specific fucosyltransferases and/or fucosidases, either through the use of an enzyme-based fermentation method (recombinant or natural enzymes) or a microbiological fermentation method. In the latter case, either the microorganisms can express their natural enzymes and substrates, or it is possible to create microorganisms that produce the corresponding substrates and enzymes. Single microorganism cultures and/or mixed cultures can be used. Fucosylated oligosaccharide formation can be initiated by acceptor substrates starting at any degree of polymerization (DP) from DP=1 onwards. In an alternative embodiment, fucosylated oligosaccharides can be obtained from lactose and free fucose using chemical synthesis. Fucosylated oligosaccharides are also available from, for example, Kyowa, Hakko, Kogyo, Japan.

The nutritional composition or milk formula for children from 1 to 3 years of the present invention also contains at least one N-acetylated oligosaccharide. It may contain one or more types of N-acetylated oligosaccharides. The N-acetylated oligosaccharide(s) may be, for example, lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT), or any combination thereof. In some specific embodiments, the N-acetylated oligosaccharide is lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT), para-lacto-N-neohexaose (para-LNnH), disialyl lacto-N-neotetraose (DSLNT) or any combination of them. In some specific embodiments, the N-acetylated oligosaccharide is LNnT. In some specific embodiments, the N-acetylated oligosaccharide is LNT. In some other specific embodiments, the N-acetylated oligosaccharide is a mixture of LNT and LNnT. In some specific embodiments, the nutritional composition or milk formula for children 1 to 3 years of age contains both LNT and LNnT in a ratio of LNT : LNnT from 5:1 to 1:2, or from 2:1 to 1:1, or from 2:1.2 to 2:1.6.

In a preferred embodiment, the nutritional composition or milk formula for children from 1 to 3 years in accordance with the invention contains lacto-N-neotetraose (LNnT). In a particular embodiment, no other type of N-acetylated oligosaccharide other than lacto-N-neotetraose (LNnT) is used, i. e. nutritional composition or milk formula for children from 1 to 3 years of the invention as N-acetylated oligosaccharide contains only lacto-N-neotetraose (LNnT).

N-acetylated oligosaccharide(s) can be chemically synthesized by enzymatic transfer of saccharide units from donor moieties to acceptor moieties using glycosyltransferases, as described, for example, in US Pat. No. 5,288,637 and WO 96/10086. In an alternative embodiment, LNT and LNnT can be produced by chemically converting ketohexoses (e.g., fructose), either free or bound to an oligosaccharide (e.g., lactulose), into an N-acetylhexosamine or N-acetylhexosamine-containing oligosaccharide, as described in Wrodnigg, T.M. ; Stutz, A.E. (1999) Angew. Chem. Int. Ed. 38:827-828. The N-acetyllactosamine thus obtained can then be transferred to lactose as an acceptor moiety.

In a particularly advantageous embodiment of the present invention, the nutritional composition or milk formula for children from 1 to 3 years of age contains 2'-fucosyllactose (2FL) and lacto-N-neotetraose (LNnT).

In another specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years of the present invention contains a mixture of oligosaccharides, consisting of 2'-fucosyllactose (2-FL) and lacto-N-neotetraose (LNnT). In other words, the nutritional composition or milk formula for children from 1 to 3 years of the invention contains only 2'-fucosyllactose (2-FL) as a fucosylated oligosaccharide and only lacto-N-neotetraose (LNnT) as an N-acetylated oligosaccharide.

In a particularly advantageous embodiment of the present invention, the fucosylated oligosaccharide(s) and N-acetylated oligosaccharide(s) are present in the nutritional composition or formula for children 1 to 3 years of age in certain specific amounts. The term "amount", unless otherwise indicated, refers to the total amount of each of such two components in a nutritional composition or formula for children from 1 to 3 years of age. Therefore, it does not refer to an individual quantity, unless it refers to a single type of such components (in which case both the total and the individual quantity would be the same). As an illustrative example, if only one fucosylated oligosaccharide (that is, only one type) is used in the composition (for example, 2FL), its individual amount (and therefore the total amount of fucosylated oligosaccharides) will be in the range of 0.75-1, 65 g/l. In the case of several fucosylated oligosaccharides (i.e. several types), their individual amount will be less (for example, in the case of 2 different types of fucosylated oligosaccharide, for example 2FL + 3FL, each of them may, for example, be present in an individual amount of 0.5 g/l) , but the total amount of fucosylated oligosaccharides will be in the range of 0.75–1.65 g/l.

Fucosylated oligosaccharide(s) may be present in the nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the present invention in a total amount of 0.75-1.65 g/l of the composition. In some embodiments, the fucosylated oligosaccharide(s) may be present in a total amount of 0.005-5 g/L of the composition, such as 0.01-3 g/L, or 0.02-2 g/L, or 0.1-2.5 g/l, or 0.15-2 g/l, or 0.25-1.9 g/l of the composition. In a particular embodiment, the fucosylated oligosaccharide(s) are present in a total of 1 g/l of the composition. In another embodiment, the fucosylated oligosaccharide(s) are present in a total amount of 0.26 g/l of the composition.

The fucosylated oligosaccharide(s) may be present in the nutritional composition or milk formula for children from 1 to 3 years of age in a total amount of 0.004-3.8 g/100 g of the composition based on dry weight. The fucosylated oligosaccharide(s) may be present in a total amount of 0.008–2.3 g/100 g of the composition, for example 0.015–1.5 g/100 g, or 0.08–1.9 g/ 100 g, or 0.12-1.5 g/100 g, or 0.15-1.5 g/100 g, or 0.19-1.5 g/100 g of composition. In a particular embodiment, the fucosylated oligosaccharide(s) are present in a total amount of 0.78 g/100 g of the composition. In another specific embodiment, the fucosylated oligosaccharide(s) are present in a total amount of 0.2 g/100 g of the composition.

N-acetylated oligosaccharide(s) may be present in the nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the present invention in a total amount of 0.1-3 g/l of the composition. In some embodiments, the N-acetylated oligosaccharide(s) may be present in a total amount of 0.01-2.5 g/L of the composition, such as 0.05-1.5 g/L, or 0 .05–1 g/l, or 0.05–0.8 g/l, or 0.08–0.8 g/l, or 0.09–0.8 g/l of the composition. In a specific embodiment, the N-acetylated oligosaccharide(s) is(s) present in a total amount of 0.5 g/l of the composition. In another specific embodiment, the N-acetylated oligosaccharide(s) is(are) present in a total of 0.13 g/l of the composition.

N-acetylated oligosaccharide(s) may be present in the nutritional composition or milk formula for children from 1 to 3 years of age in a total amount of 0.008-2.3 g/100 g of the composition based on the dry weight. N-acetylated oligosaccharide(s) may be present in a total amount of 0.008–1.9 g/100 g of the composition, for example 0.04–1.2 g/100 g, or 0.04–0 .8 g/100 g, or 0.04–0.6 g/100 g, or 0.06–0.6 g/100 g, or 0.07–0.6 g/100 g of composition. In a particular embodiment, the N-acetylated oligosaccharide(s) is(are) present in a total amount of 0.38 g/100 g of the composition. In another specific embodiment, the N-acetylated oligosaccharide(s) is(are) present in a total amount of 0.1 g/100 g of the composition.

Therefore, in one embodiment of the present invention, the nutritional composition or milk formula for children from 1 to 3 years old contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, wherein:

- fucosylated oligosaccharide(s) are present in a total amount of 0.005–5 g/l of the composition and/or in a total amount of 0.004–3.8 g/100 g of the composition, based on the weight of dry matter; and/or

- N-acetylated oligosaccharide(s) is present in a total amount of 0.1–2.5 g/l of the composition and/or in a total amount of 0.08–1.9 g/100 g of the composition in terms of dry weight.

In another specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years of the present invention contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, and:

- fucosylated oligosaccharide(s) are present in a total amount of 0.005–5 g/l of the composition and/or in a total amount of 0.004–3.8 g/100 g of the composition, based on the weight of dry matter; and/or

- N-acetylated oligosaccharide(s) is present in a total amount of 0.05–1.5 g/l of the composition and/or in a total amount of 0.04–1.2 g/100 g of the composition in terms of dry weight.

In another specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years of the present invention contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, and:

- fucosylated oligosaccharide(s) are present in a total amount of 0.02–2 g/l of the composition and/or in a total amount of 0.015–1.5 g/100 g of the composition, based on the weight of dry matter ; and/or

- N-acetylated oligosaccharide(s) is present in a total amount of 0.5–1 g/l of the composition and/or in a total amount of 0.4–0.8 g/100 g of the composition, calculated on dry weight.

In another specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years of the present invention contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, and:

- fucosylated oligosaccharide(s) is present in a total amount of 0.1–2.5 g/l of the composition and/or in a total amount of 0.08–1.9 g/100 g of the composition, calculated by weight of dry matter; and/or

- N-acetylated oligosaccharide(s) is (are) present in a total amount of 0.5–0.8 g/l of the composition and/or in a total amount of 0.4–0.6 g/100 g of the composition in terms of dry weight.

In a specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years in accordance with the present invention contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, and:

- fucosylated oligosaccharide(s) are present in a total amount of 1 g/l of the composition and/or in a total amount of 0.77 g/100 g of the composition, based on the weight of dry matter; and/or

- N-acetylated oligosaccharide(s) are present in a total amount of 0.5 g/l of the composition and/or in a total amount of 0.38 g/100 g of the composition, based on the weight of dry matter.

In another specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years in accordance with the invention contains at least one fucosylated oligosaccharide and at least one N-acetylated oligosaccharide, and:

- fucosylated oligosaccharide(s) are present in a total amount of 0.26 g/l of the composition and/or in a total amount of 0.2 g/100 g of the composition, based on the weight of dry matter; and/or

- N-acetylated oligosaccharide(s) are present in a total amount of 0.13 g/l of the composition and/or in a total amount of 0.1 g/100 g of the composition, based on the weight of dry matter.

The fucosylated oligosaccharide(s) and N-acetylated oligosaccharide(s) contained in the nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the invention, as a rule, are present in a ratio of N-acetylated oligosaccharide(s) : fucosylated oligosaccharide(s) from 1 : 20 to 2 : 1, preferably from 1 : 15 to 1 : 1, most preferably from 1 : 10 to 1:2. In a particularly advantageous embodiment, this ratio is 1:2, or about 1:2.

In a specific embodiment, the fucosylated oligosaccharide is contained in the nutritional composition or infant formula for children from 1 to 3 years of the present invention in such an amount that the normal intake of the nutritional composition or formula for children from 1 to 3 years of age will be provided for an infant or young child, respectively a child, when consuming a total daily dose of 0.003-6.5, preferably 0.006-3.9 g, for example 0.012-2.6 g, per day.

For preterm, low birth weight, and small for their gestational age infants, the daily dose of fucosylated oligosaccharides is preferably 0.05 to 1 g/kg body weight per day, preferably 0.06–0.9 g, or 0.07– 0.8 g or 0.08-0.7 g or 0.09-0.6 g or 0.1-0.5 g or 0.2-0.4 g, most preferably 0.34 g per kg of body weight per day.

In a specific embodiment, the N-acetylated oligosaccharide is contained in the infant formula or infant formula of the present invention in an amount such that a normal intake of the infant or toddler formula or infant formula of the present invention is provided. age, respectively, of the child, when consuming a total daily dose of 0.003–3.9 g, preferably 0.006–3.25 g, or 0.03–1.95 g, or 0.03–1.3 g, or 0.03– 1 g, e.g. 0.05-1 g per day.

For preterm, low birth weight and small for their gestational age infants, the daily dose is preferably 0.005 to 0.1 g/kg body weight per day, preferably 0.006 to 0.09 g, or 0.007 to 0.08 g, or 0.008-0.07 g or 0.009-0.06 g or 0.01-0.05 g or 0.02-0.04 g, most preferably 0.034 g per kg body weight per day.

The nutritional composition or milk formula for children from 1 to 3 years in accordance with the present invention may also contain at least other (s) oligosaccharide (s) (i.e., different (s) from the fucosylated (s) oligosaccharide (s) and N-acetylated (s) oligosaccharide (s) necessarily present (s) in the composition), and / or at least fiber (s), and / or at least their precursor (- And). The other oligosaccharide and/or fiber and/or precursor thereof may be selected from a list including galactooligosaccharides (GOS), fructooligosaccharides (FOS), inulin, xylooligosaccharides (XOS), polydextrose, sialylated oligosaccharides, sialic acid, fucose, and any combination thereof. . They may be present in an amount of 0 to 10% by weight of the composition. In a specific embodiment, the nutritional composition or infant formula for children from 1 to 3 years may also include at least one bovine milk oligosaccharide (BMO).

Suitable commercially available products that can be used in addition to the oligosaccharides contained in the oligosaccharide mixture to produce nutritional compositions or infant formula for children from 1 to 3 years of age in accordance with the invention include combinations of FOS with inulin, such as the product sold by BENEO under the trade name Orafti, or polydextrose sold by Tate & Lyle under the trade name STA-LITE®.

In a specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years in accordance with the invention may contain sialylated(s) oligosaccharide(s). It may contain one or more sialylated oligosaccharides. The sialylated oligosaccharide(s) may be selected from the group consisting of 3'-sialyllactose (3-SL), 6'-sialyllactose (6-SL) and any combination thereof. In some embodiments of the invention, the composition contains 3-SL and 6-SL. In some specific embodiments, the ratio of 3'-sialyllactose (3-SL) to 6'-sialyllactose (6-SL) may range from 5:1 to 1:10, or from 3:1 to 1:1, or from 1:1 to 1:10. In some specific embodiments, the sialylated oligosaccharide of the composition is 6'-sialyllactose (6-SL).

The sialylated oligosaccharide(s) may be isolated from a natural source such as animal milk by chromatography or filtration. In an alternative embodiment, they can be produced by biotechnological means using specific sialyltransferases or sialidases, neuraminidase, or by enzyme-based fermentation (recombinant or natural enzymes), by chemical synthesis, or by microbiological fermentation. In the latter case, microorganisms can express their natural enzymes and substrates, or it is possible to create microorganisms that produce the corresponding substrates and enzymes. Single-species cultures or mixed cultures can be used. The formation of sialyl oligosaccharides can be initiated by acceptor substrates starting at any degree of polymerization (DP) from DP = 1 onwards. In an alternative embodiment, sialyllactoses can be obtained by chemical synthesis from lactose and free N'-acetylneuraminic acid (sialic acid). Sialyl lactoses are also commercially available, for example from Kyowa Hakko Kogyo, Japan.

In another preferred embodiment of the invention, the nutritional composition or milk formula for children from 1 to 3 years old may include 0.005-5 g/l of sialylated oligosaccharides, or 0.008-2.5 g/l, or 0.01-1 g/l, or 0.02–0.7 g/l, e.g. 0.03–0.5 g/l.

A nutritional composition or milk formula for children from 1 to 3 years of age according to the invention may include 0.004-3.8 g of sialylated oligosaccharides per 100 g of the composition based on the weight of dry matter, for example 0.006-1.9 g, or 0.008-0. 8 g, or 0.015-0.5 g, for example 0.023-0.4 sialylated oligosaccharides per 100 g of the composition, based on the weight of dry matter.

In some specific embodiments of the present invention, the nutritional composition or milk formula for children from 1 to 3 years old contains sialylated oligosaccharide(s) in an amount of less than 0.1 g/100 g of the composition based on dry weight.

In a particular embodiment, the sialylated oligosaccharide is contained in the infant formula or infant formula of the present invention in such an amount that a normal intake of the infant or toddler formula or formula for infants 1 to 3 years of age is provided, respectively of a child, when consuming a total daily dose of 0.003-6.5 g, preferably 0.005-3.3 g, or 0.006-1.3 g, or 0.01-0.9 g, for example 0.018-0.65 g, in day.

In some specific embodiments, the implementation of the present invention nutritional composition or milk formula for children from 1 to 3 years does not contain any(s) sialylated(s) oligosaccharide(s).

The nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the present invention may also optionally include at least one oligosaccharide precursor. It may contain one or more oligosaccharide precursors. For example, the breast milk oligosaccharide precursor is sialic acid, fucose, or a mixture thereof. In some specific embodiments, the implementation of the composition contains sialic acid.

In specific examples, the nutritional composition or milk formula for children from 1 to 3 years old contains from 0 to 3 g/l of the oligosaccharide precursor(s), or from 0 to 2 g/l, or from 0 to 1 g/l, or from 0 to 0.7 g/l or 0 to 0.5 g/l or 0 to 0.3 g/l or 0 to 0.2 g/l oligosaccharide precursor(s). A composition according to the invention may comprise 0 to 2.1 g of oligosaccharide precursor(s) per 100 g of composition on a dry weight basis, for example 0 to 1.5 g, or 0 to 0.8 g, or 0 to 0.15 g of oligosaccharide precursor(s) per 100 g of composition based on dry weight.

The nutritional composition or milk formula for children from 1 to 3 years of the present invention may further include at least one probiotic (or probiotic strain), such as a probiotic bacterial strain.

The most commonly used probiotic microorganisms, which are mainly bacteria and yeasts of the following genera: Lactobacillus spp., Streptococcus spp., Enterococcus spp., Bifidobacterium spp. and Saccharomyces spp.

In some specific embodiments, the implementation of the probiotic is a probiotic bacterial strain. In some specific embodiments, the implementation is, in particular, strains of Bifidobacteria and/or Lactobacilli.

Suitable probiotic bacterial strains include Lactobacillus rhamnosus ATCC 53103, from Valio Oy, Finland, under the trademark LGG; Lactobacillus rhamnosus CGMCC 1.3724, Lactobacillus paracasei CNCM I-2116, Lactobacillus johnsonii CNCM I-1225, Streptococcus salivarius DSM 13084 sold by BLIS Technologies Limited, New Zealand under the name KI2; Bifidobacterium lactis CNCM 1-3446, sold inter alia by Christian Hansen, Denmark, under the trademark Bb 12; Bifidobacterium longum ATCC BAA-999 sold by Morinaga Milk Industry Co. Ltd., Japan, under the trademark BB536; Bifidobacterium breve sold by Danisco under the trademark Bb-03; Bifidobacterium breve sold by Morinaga under the trademark M-16V; Bifidobacterium infantis marketed by Procter & Gambie Co. under the Bifantis trademark; and Bifidobacterium breve sold by Institut Rosell (Lallemand) under the trademark R0070.

A nutritional composition or milk formula for children from 1 to 3 years of age according to the invention may comprise from 10e3 to 10e12 cfu of the probiotic strain, more preferably from 10e7 to 10e12 cfu, for example from 10e8 to 10e10 cfu, of the probiotic strain per gram of composition, by weight dry matter.

In one embodiment, the probiotics are viable. In another embodiment, the probiotics are non-propagating or inactivated. In some other embodiments, both viable probiotics and inactivated probiotics can be used. You can also add probiotic components and metabolites.

The nutritional composition according to the invention may, for example, be an infant formula, a starter infant formula, a formula for complementary foods or a follow-up formula, an infant formula for children from 1 to 3 years old, an infant formula, a cereal-based infant formula, a fortifier, such as a human breast milk fortifier, or supplement. In some specific embodiments, the composition of the invention is an infant formula, fortifier or supplement, which may be for the first 4 or 6 months of life. In a preferred embodiment, the nutritional composition of the invention is an infant formula.

In some other embodiments, the nutritional composition of the present invention is a fortifier. The fortifier can be a human milk fortifier (eg human breast milk fortifier) or a formula fortifier such as an infant formula fortifier or formula fortifier for formula fed/follow-up formula.

If the nutritional composition is a supplement, it may be provided in unit dosage form. In such cases, it is particularly useful to determine the amount of N-acetylated oligosaccharides and optionally other oligosaccharides in relation to the daily dose for administration to an infant or young child, as described above.

The nutritional composition of the present invention may be in solid (eg powder), liquid or gelatinized form.

In a particular embodiment, the nutritional composition is in the form of a powder supplement and is supplied in a sachet or syrup form. If the additive is in the form of a powder, it may include a carrier. However, it is preferable that the carrier in the additive was absent. If the supplement is in the form of a syrup, the HMO is preferably dissolved or suspended in water acidified with citrate.

The nutritional composition or milk formula for children from 1 to 3 years of age according to the invention usually contains a protein source. Protein can be contained in an amount of 1.6 to 3 g per 100 kcal. In some embodiments, particularly when the composition is for preterm infants, the amount of protein may be from 2.4 to 4 g/100 kcal, or greater than 3.6 g/100 kcal. In some other embodiments, the amount of protein may be below 2.0 g per 100 kcal, such as 1.8 to 2 g/100 kcal, or the protein may be present in an amount below 1.8 g per 100 kcal.

The type of protein is not considered critical to the present invention, provided that the minimum requirements for the content of essential amino acids are met and satisfactory growth is ensured. Thus, protein sources based on whey, casein and mixtures thereof, as well as protein sources based on soy, can be used. With respect to whey proteins, the protein source may be based on acid whey or sweet whey, or mixtures thereof, and may include alpha-lactalbumin and beta-lactoglobulin in any desired proportions.

In some preferred embodiments, the protein source is whey-dominated (ie, more than 50% of the proteins are from whey proteins, eg, 60% or 70%).

The proteins may be native or hydrolysed, or they may be a mixture of native and hydrolyzed proteins. The term "native" means that the bulk of the proteins are native, i. their molecular structure is not altered, e.g. at least 80% of the proteins are not altered, e.g. at least 85% of the proteins are not altered, preferably at least 90% of the proteins are not altered, even more preferably at least 95% of the proteins are not altered, e.g. at least 98% of the proteins are changed. In a specific embodiment, 100% of the proteins are unchanged.

The term "hydrolyzed" in the context of the present invention means a protein that has been hydrolyzed or cleaved into its constituent amino acids. Proteins may be fully or partially hydrolysed. It may be desirable to administer partially hydrolysed proteins (degree of hydrolysis between 2 and 20%), for example, to infants or young children who are at risk of developing cow's milk allergy. If it is necessary to use hydrolyzed proteins, the hydrolysis process can be carried out as desired and in a manner known in the art. For example, whey protein hydrolysates can be prepared by enzymatic hydrolysis of a whey fraction in one or more steps. If the whey fraction used as starting material is substantially free of lactose, it has been found that the blocking of protein lysine occurs to a much lesser extent during hydrolysis. This allows the degree of lysine blockage to be reduced from about 15% by weight of total lysine to less than about 10% by weight of lysine; for example, up to about 7 wt.% lysine, which significantly improves the nutritional quality of the protein source.

In one embodiment of the invention, at least 70% of the proteins are hydrolyzed, preferably at least 80% of the proteins are hydrolyzed, e.g. at least 85% of the proteins are hydrolyzed, even more preferably at least 90% of the proteins are hydrolyzed, e.g. at least 95% of the proteins are hydrolyzed in particular, at least 98% of the proteins are hydrolysed. In a specific embodiment, 100% of the proteins are hydrolysed.

In one specific embodiment, the proteins of the nutritional composition are hydrolyzed, fully hydrolyzed, or partially hydrolyzed. The degree of hydrolysis (DH) of the protein can be from 8 to 40, or from 20 to 60, or from 20 to 80, or more than 10, 20, 40, 60, 80, or 90.

In an alternative embodiment, the protein component may be replaced with a mixture or an artificial amino acid, such as for preterm or low birth weight infants.

In a specific embodiment, the nutritional composition or milk formula for children from 1 to 3 years in accordance with the invention is a hypoallergenic composition. In another specific embodiment, the composition in accordance with the invention is a hypoallergenic nutritional composition or milk formula for children from 1 to 3 years.

The nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the present invention usually contains a source of carbohydrates. This is especially preferred when the nutritional composition of the invention is an infant formula. In this case, any carbohydrate source commonly found in infant formulas can be used, such as lactose, sucrose, sucrose, maltodextrin, starch, and mixtures thereof, although one preferred carbohydrate source is lactose.

The nutritional composition or milk formula for children from 1 to 3 years in accordance with the present invention usually contains a source of lipids. This is especially important if the nutritional composition of the present invention is an infant formula. In this case, the lipid source can be any lipid or fat that is suitable for use in infant formulas. Some suitable fat sources include palm oil, structured triglyceride oil, high oleic sunflower oil, and high oleic safflower oil, medium chain triglyceride oil. You can also add essential fatty acids - linoleic acid and α-linolenic acid, as well as small amounts of oils containing large amounts of arachidonic acid and docosahexaenoic acid in finished form, such as fish oils or microbial oils. The ratio of n-6 to n-3 fatty acids in the fat source may be from about 5:1 to about 15:1; for example, from about 8:1 to about 10:1.

The nutritional composition or milk formula for children from 1 to 3 years of the invention may also contain all the vitamins and minerals considered indispensable in the daily diet, in nutritionally significant amounts. Some vitamins and minerals have minimum requirements. Examples of minerals, vitamins and other nutrients that are optionally present in the composition of the invention include vitamin A, vitamin B1, vitamin B2, vitamin B6, vitamin B12, vitamin E, vitamin K, vitamin C, vitamin D, folic acid, inositol, niacin, biotin, pantothenic acid, choline, calcium, phosphorus, iodine, iron, magnesium, copper, zinc, manganese, chlorine, potassium, sodium, selenium, chromium, molybdenum, taurine and L-carnitine. Minerals are usually added in the form of salts. The presence and amounts of specific minerals and other vitamins may vary depending on the intended population.

If necessary, the nutritional composition or milk formula for children from 1 to 3 years of the invention may contain emulsifiers and stabilizers, such as soy, lecithin, mono- and diglyceryl esters of citric acid, and the like.

The nutritional composition or milk formula for children from 1 to 3 years of the invention may also contain other substances that may have a beneficial effect, such as lactoferrin, nucleotides, nucleosides, and the like.

The nutritional composition or milk formula for children from 1 to 3 years of the invention may also contain carotenoid(s). In some specific embodiments of the invention, the nutritional composition of the invention does not contain carotenoids.

The nutritional composition or milk formula for children from 1 to 3 years of age in accordance with the invention can be obtained by any suitable method. Next, the composition will be described on a specific example.

For example, a formula, such as infant formula, can be made by mixing a protein source, a carbohydrate source, and a fat source in appropriate proportions. At this stage, emulsifiers can be included if they are used. Vitamins and minerals can be added at this stage, but are usually added later to prevent thermal decomposition. Any lipophilic vitamins, emulsifiers, and the like may be dissolved in the fat source prior to mixing. Water, preferably purified by reverse osmosis, can then be mixed in to form a liquid mixture. The temperature of the water conveniently ranges from about 50° C. to about 80° C. to facilitate dispersion of the ingredients. Commercially available thinners can be used to form the liquid mixture.

Fucosylated oligosaccharide(s) and N-acetylated oligosaccharide(s) may be added at this stage, especially if the final product is to be in liquid form. If the end product is to be a powder, these components can also be added at this stage if desired.

The liquid mixture is then homogenized, for example in two stages.

The liquid mixture can then be thermally treated to reduce the bacterial load by rapidly heating the liquid mixture, for example to a temperature in the range of about 80° C. to about 150° C., over a period of time from about 5 seconds to about 5 minutes. The heating can be carried out by steam injection using an autoclave or a heat exchanger such as a plate heat exchanger.

The liquid mixture can then be cooled to a temperature of from about 60° C. to about 85° C., for example by flash cooling. The liquid mixture can then be homogenized again, for example in two stages: under pressure in the range of about 10 MPa to about 30 MPa in the first stage and from about 2 MPa to about 10 MPa in the second stage. The homogenized mixture can then be further cooled to add any heat sensitive components such as vitamins and minerals. At this point, the pH and solids content of the homogenized mixture is adjusted for convenience.

If the end product is to be a powder, the homogenized mixture is transferred to a suitable drying apparatus, such as a spray dryer or freeze dryer, and converted to powder. The moisture content of the powder should be less than about 5% by weight. Fucosylated oligosaccharide(s) and N-acetylated oligosaccharide(s) can also or alternatively be added at this stage by dry mixing or by mixing them in the form of a crystal syrup together with the probiotic (- them) strain(s) (if used), and the mixture is spray dried or freeze dried.

If a liquid composition is preferred, the homogenized mixture may be sterilized and then aseptically filled into suitable containers, or may be first filled into containers and then sterilized by autoclaving.

In another embodiment, the composition of the invention may be an additive. The supplement may be presented, for example, in the form of tablets, capsules, lozenges or liquid. The additive may further contain protective hydrocolloids (such as gums, proteins, modified starches), binders, film-forming agents, encapsulating agents/materials, wall/shell materials, matrix compounds, coatings, emulsifiers, surfactants, solubilizing agents (oils, fats, waxes, lecithins, etc.), adsorbents, carriers, fillers, auxiliary compounds, dispersing agents, wetting agents, processing aids (solvents), anti-caking agents, taste masking agents, weighting agents, gelling agents and gelling agents. The additive may also contain traditional pharmaceutical additives and excipients, excipients and diluents, including, without limitation, water, gelatin of any origin, vegetable gums, lignin sulfonate, talc, sugars, starch, gum arabic, vegetable oils, polyalkylene glycols, flavoring agents, preservatives, stabilizers , emulsifying agents, buffers, lubricants, colorants, wetting agents, fillers, and the like.

Additionally, the supplement may contain an organic or inorganic carrier material suitable for oral or parenteral administration, as well as vitamins, trace minerals and other trace elements in accordance with the recommendations of government agencies, such as the recommended intakes (RDA) of the United States.

The nutritional composition according to the invention is intended for use in infants or young children. Infants or young children may be born at term or prematurely. In a specific embodiment, the nutritional composition of the invention is for use in infants or young children who were born premature, have a low birth weight and/or were born small for their gestational age (SGA). In a particular embodiment, the nutritional composition of the invention is for use in preterm infants, infants of low birth weight and/or infants born small for their gestational age (SGA).

The nutritional composition of the present invention may also be used in infants or young children delivered by caesarean section or by vaginal delivery.

In some embodiments, the implementation of the composition in accordance with the invention may be intended for use before and/or during the period of weaning.

The age at which a nutritional composition is administered (or used) or fed to it, as well as the duration of such administration, depends on the needs.

The nutritional composition may, for example, be given to infants immediately after birth. The composition of the invention can also be given during the first week of an infant's life, or during the first 2 weeks of life, or during the first 3 weeks of life, or during the first month of life, or during the first 2 months of life, or during the first 3 months of life. , or during the first 4 months of life, or during the first 6 months of life, or during the first 8 months of life, or during the first 10 months of life, or during the first year of life, or during the first two years of life, or even longer. In some particularly advantageous embodiments of the invention, the nutritional composition is given (or administered) to an infant during the first 4, 6, or 12 months after the birth of said infant. In some other embodiments, the nutritional composition of the invention is given over several days (e.g., 1, 2, 3, 5, 10, 15, 20, etc.), or several weeks (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, etc.), or several months (for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, etc.) after birth. First of all, this may be relevant, but not necessarily, in the case when the baby was born prematurely.

In one embodiment, the composition of the invention is given to an infant or young child as a composition to supplement mother's milk. In some embodiments, the infant or young child receives breast milk for at least the first 2 weeks, the first 1, 2, 4, or 6 months. In one embodiment, the nutritional composition of the invention is given to an infant or young child after such a breast milk feeding period, or is given during such a breast milk feeding period. In another embodiment of the invention, the composition is given to an infant or young child as the sole or main nutritional composition for at least one period of time, for example after 1, 2 or 4 months of life, for at least 1, 2, 4 or 6 months.

In one embodiment, the nutritional composition of the invention is a complete nutritional composition (satisfying all or most of the subject's nutritional needs). In another embodiment, the nutritional composition is a supplement or fortifier intended, for example, to supplement human breast milk, or to supplement infant formula or complementary formula/follower formula.

Examples

The following examples demonstrate some specific embodiments of the composition for use in accordance with the present invention. The examples are for illustrative purposes only and should not be construed as limitations of the present invention.

Example 1

Table 1 below shows an example of the composition of a nutritional composition (eg, infant formula) in accordance with the present invention. This composition is for illustrative purposes only.

Table 1. Composition of infant formula according to example 1

Nutrients Per 100 kcal per liter Energy value (kcal) 100 670 Protein (g) 1.83 12.3 Fat (g) 5.3 35.7 Linoleic acid (g) 0.79 5.3 α-Linolenic acid (mg) 101 675 lactose (g) 11.2 74.7 Minerals (g) 0.37 2.5 Na (mg) 23 150 K (mg) 89 590 Cl (mg) 64 430 Ca (mg) 62 410 P (mg) 31 210 Mg (mg) 7 50 Mn (mcg) 8 50 Se (mcg) 2 13 Vitamin A (µg RE) 105 700 Vitamin D (mcg) 1.5 10 Vitamin E (mg TE) 0.8 5.4 Vitamin K1 (mcg) 8 54 Vitamin C (mg) 10 67 Vitamin B1 (mg) 0.07 0.47 Vitamin B2 (mg) 0.15 1.0 Niacin (mg) 1 6.7 Vitamin B6 (mg) 0.075 0.50 Folic acid (mcg) 9 60 Pantothenic acid (mg) 0.45 3 Vitamin B12 (mcg) 0.3 2 Biotin (mcg) 2.2 15 Choline (mg) 10 67 Fe (mg) 1.2 8 I (µg) 15 100 Cu (mg) 0.06 0.4 Zn (mg) 0.75 5 Oligosaccharides
(OGM) 2FL(g) 0.15 1 LNnT (g) 0.075 0.5

Table 1. An example of the composition of the nutritional composition (for example, infant formula) in accordance with the present invention

Example 2

Study Description

Three sets of concurrently bred female Sprague-Dawley rats were purchased from Charles River Laboratories. During the last 10 days of pregnancy, one group had dietary restrictions of 60% and their offspring were cross-fed to normal-fed rats. The second group of pregnant females received normal nutrition, and their offspring were cross-fed among the same group of mothers. Immediately after birth (d = 2), the born rat pups — participants in the experiment — were divided into one of the following groups:

• normal group (positive control): normal weight offspring born to unrestricted mothers fed a normal diet without supplements;

• IUGR group (negative control; n = 20): IUGR rats fed a normal diet after birth;

• IUGR + HMO rats (test groups): during the experiment they were fed by their mothers for 21 days and received a supplement of HMO (2FL, LNnT or a mixture of 2FL and LNnT). After weaning, they were fed a diet supplemented with the same HMOs, see details below. There were three different groups:

- group ZVUR/2FL (n = 10): with the addition of only 2-FL;

- group IUGR/LNnT (n = 10): with the addition of only LNnT; And

- group ZVUR/mixture OGM (n = 10): with the addition of LNnT and 2FL in a mass ratio of 1 : 2.

All rat pups were fed from birth (d = 1) to 57 days (d = 57) based on the following protocol:

• d1–d6: all groups of rats were mother-fed (only mother's milk);

• d7–d21: all rats were mother-fed (mother's milk only), but in test groups 3 g/kg body weight of HMO was also administered via gavage:

- 3 g/kg body weight 2FL for the IUGR/2FL group;

- 3 g/kg body weight LNnT for the IUGR/LNnT group;

- 3 g/kg body weight of a mixture of LNnT and 2FL in a mass ratio of 1 : 2 for the IUGR group/a mixture of HMO;

• d22-d57: All rats received the diet as detailed in Table 2 below.

Table 2. Composition of diets received by different groups from d22 to d57

Amount in the diet of the control group [%] Amount in the diet of the 2FL group [%] Amount in the diet of the LNnT group [%] Amount in the diet of the 2FL and LNnT groups [%] Corn starch 53.4 53.4 53.4 53.4 Casein 20 20 20 20 sucrose 10 10 10 10 Soybean oil 7 7 7 7 Mineral Blend AIN-93-G* 3.5 3.5 3.5 3.5 Choline bitartrate 0.25 0.25 0.25 0.25 L-cystine 0.3 0.3 0.3 0.3 tert-butylhydroquinone 0.0014 0.0014 0.0014 0.0014 Vitamin Blend AIN-93-VX* 1 1 1 1 Maltodextrin 4.5 0 0 0 2FL 0 4.5 0 3 LNnT 0 0 4.5 1.5

*from Research Diets, Inc.

Rats were sacrificed on day 57 after a 6 hour fasting period and small intestine samples were collected. The length of the villi was measured in the sample of the ileum.

A single 1 cm sample (1 cm cranially from the caecum) was taken from the ileum and washed with PBS (Sigma D8537). This section was fixed and appropriately processed for histological and morphometric analysis.

Two consecutive sections of fixed tissue, 4 µm thick, were manually stained with hematoxylin and eosin to determine the height of the villi. Villus height was acquired by light microscopy with a dry objective at 10x magnification using the Axion Vision computerized image analysis system (version 4.8.2.0, Zeiss®). To determine the average length of the villi in two different segments, ten randomly selected villi were measured.

conclusions

The villus length measured in each rat is shown in FIG. 1. For each group, the average length of the villi is represented by a horizontal line. As can be seen, the average length of the villi of all groups was similar, with the exception of the group receiving the combination of 2FL and LNnT, which had significantly longer villi in the ileum. This increase in the length of the villi is associated with an increase in the surface area of the ileum, which provides the baby rats with improved digestive capacity, in particular for the breakdown of fats, proteins and carbohydrates. This improved digestion, in particular the breakdown of fats, is facilitated by improved enterohepatic circulation, in particular enterohepatic circulation of bile salts, which is associated with an increase in the surface area of the small intestine, in particular the ileum.

These results indicate that a mixture of 2FL and LNnT provided a successful increase in the length of the villi in the ileum of rats with growth retardation (due to maternal malnutrition). The villi of stunted pups fed the 2FL and LNnT diet were even significantly longer than those of pups that did not experience intrauterine growth retardation and were fed a normal diet.

These results also indicate synergy between 2FL and LNnT, given that neither 2FL nor LNnT produced a statistically significant effect when used alone, while a statistically significant effect was observed when both compounds were used in combination.

Example 3

The supplement for premature infants is prepared to give the following daily dose:

- 0.034 g/kg of body weight per day LNnT; And

- 0.34 g/kg of body weight per day 2-FL.

Claims (25)

1. Use of an artificial nutritional composition containing at least one N-acetylated oligosaccharide and at least one fucosylated oligosaccharide in a method for activating the digestive capacity of an infant or young child. 2. The use of milk formula for children from 1 to 3 years, containing at least one N-acetylated oligosaccharide and at least one fucosylated oligosaccharide, in a method for activating the digestive capacity of a child. 3. Use according to claim 1 or 2, wherein the activation of the digestive capacity includes the activation of the capacity to break down fat, fat-soluble vitamins, proteins and carbohydrates in an infant or young child, respectively, in a child. 4. Use according to any one of the preceding claims, wherein the activation of the digestive capacity includes: improvement of enterohepatic circulation, preferably enterohepatic circulation of bile salts; development and/or increase in the surface area of the small intestine; and/or an increase in the length of the villi in the small intestine. 5. Use according to any one of the preceding claims, wherein at least one N-acetylated oligosaccharide is lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT), para-lacto-N-neohexaose (para-LNnH) , disialyl lacto-N-neotetraose (DSLNT), or any combination thereof. 6. Use according to any one of the preceding claims, wherein at least one fucosylated oligosaccharide is selected from the list consisting of 2'-fucosyllactose, 3'-fucosyllactose, difucosyllactose, lacto-N-fucopentaose I, lacto-N-fucopentaose II, lacto- N-fucopentaose III, lacto-N-fucopentaose V, lacto-N-fucohexaose, lacto-N-difucohexaose I, fucosyllacto-N-hexaose, fucosyllacto-N-neohexaose I, fucosyllacto-N-neohexaose II, difucosyllacto-N-hexaose I, difucosyl lacto-N-neohexaoses I, difucosyl lacto-N-neohexaoses II, fucosyl-para-lacto-N-hexaoses, and any combination thereof. 7. Use according to any one of the preceding claims, wherein the artificial nutritional composition or milk formula for children from 1 to 3 years of age contains 2'-fucosyllactose and lacto-N-neotetraose (LNnT) or contains an oligosaccharide mixture consisting of 2'-fucosyllactose (2 'FL) and lacto-N-neotetraose (LNnT). 8. Use according to any one of the preceding claims, wherein the N-acetylated oligosaccharide and fucosylated oligosaccharide are present in a ratio of N-acetylated oligosaccharide(s):fucosylated oligosaccharide(s) from 1:20 to 2:1, preferably from 1:15 to 1:1, more preferably from 1:10 to 1:2, most preferably the ratio is 1:2. 9. Application according to any of the preceding claims, and the fucosylated oligosaccharide(s) is(are) present in a total amount of 0.005-5 g/l of the composition or in a total amount of 0.004-3.8 g/100 g of the composition based on the weight of dry matter; and/or N-acetylated oligosaccharide(s) are present in a total amount of 0.1-2.5 g/l of the composition or in a total amount of 0.08-1.9 g/100 g of the composition based on the weight of dry matter . 10. Application according to any of the preceding claims, and fucosylated oligosaccharide(s) are present in a total amount of 0.02-2 g/l of the composition or in a total amount of 0.015-1.5 g/100 g of the composition based on the weight of dry matter; and/or The N-acetylated oligosaccharide(s) is(are) present in a total amount of 0.5-1 g/l of the composition, or in a total amount of 0.4-0.8 g/100 g of the composition, based on the weight of dry matter. 11. Application according to any of the preceding claims, and the fucosylated oligosaccharide(s) is(are) present in a total amount of 0.1-2.5 g/l of the composition or in a total amount of 0.08-1.9 g/100 g of the composition based on the weight of dry matter; and/or The N-acetylated oligosaccharide(s) is(are) present in a total amount of 0.5-0.8 g/l of the composition or in a total amount of 0.4-0.6 g/100 g of the composition based on dry weight . 12. Use according to any of the preceding claims, wherein said artificial nutritional composition or milk formula for children from 1 to 3 years of age contains at least other(s) oligosaccharide(s), and/or fiber(s), and/or their precursor (i) selected from the list including galactooligosaccharides (GOS), fructooligosaccharides (FOS), xylooligosaccharides (XOS), inulin, polydextrose, sialylated oligosaccharides, sialic acid, fucose, and any combination thereof. 13. The use according to any one of the preceding claims, wherein said artificial nutritional composition or milk formula for children from 1 to 3 years additionally contains at least one probiotic in an amount of from 10 ³ to 10 ¹² CFU/g of said composition (by weight of dry matter) . 14. Application according to any paragraphs. 1 or 3-13, wherein said artificial nutritional composition is infant formula, initial infant formula, infant formula for formula-fed infants or follow-up infant formula, milk formula for infants from 1 to 3 years old, infant formula, cereal-based infant formula, fortifier or additive. 15. Application according to any one of paragraphs. 1 or 3-14, wherein the infant is a premature infant, a low birth weight infant, an infant born small for gestational age (SGA), and/or a growth retarded infant or young child. 16. Use according to claim 15, wherein the infant is an IUGR infant.

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