Nutritional formulations for promoting healthy gut microbiota and function

Abstract

Disclosed are methods and compositions, including infant formulas, for promoting the growth of beneficial bacteria in the intestine of an infant and / or for promoting a strong intestinal barrier. The compositions include chondroitin sulfate and, in some embodiments, one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

Description

NUTRITIONAL FORMULATIONS FOR PROMOTING HEALTHY GUT MICROBIOTA AND FUNCTION CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to United States Provisional Patent Application No. 63 / 666,976, filed on July 2, 2024. FIELD

[0002] The present disclosure relates to a composition, such as an infant formula, containing chondroitin, optionally in the form of chondroitin sulfate, and optionally one or more human milk oligosaccharides (HMOs), one or more nucelotides, or both. The present disclosure also relates to methods of promoting gut health, such as in an infant, by administering a nutritional composition containing chondroitin, optionally in the form of chondroitin sulfate, and optionally one or more human milk oligosaccharides (HMOs), one or more nucelotides, or both. BACKGROUND

[0003] It is generally accepted that the gut microbiota composition of healthy, full-term, breastfed infants is the standard for healthy infant microbiota development. In particular, bifidobacteria are the dominant microbial group in a healthy breastfed infant and have been shown to play an important role in infant health and development. The levels of bifidobacteria in formula-fed infants has been shown to be lower than that present in healthy breastfed infants. This has led to various attempts at developing infant formulas that promote the colonization of bifidobacteria in the infant gut.

[0004] Further, during postnatal development, a newborn's intestine experiences a process of maturation that ends with the production of gastrointestinal epithelium that functions as a selective barrier (i.e., gut barrier or intestinal barrier). The gastrointestinal epithelium permits the absorption of nutrients, electrolytes and water, while preventing exposure to dietary and microbial antigens, including food allergens. Specifically, this barrier limits the passage of antigens to systemic circulation, thereby preventing infection, inflammatory reactions, and other gastrointestinal diseases and disorders that may occur during infancy and later in life. For veryAttorney Docket No. 15402WOO1 (67909WO01) young infants, and particularly preterm infants, who have an immature immune system and intestinal tract, development of suboptimal intestinal flora and / or breakdown of the intestinal barrier can hinder development and may result in infection, diarrhea, allergies, and food intolerance. SUMMARY

[0005] The present inventors have found that the development of an early bifidogenic intestinal microbiota in formula-fed infants may be enhanced by feeding the infants an infant formula that contains chondroitin, such as in the form of chondroitin sulfate.

[0006] The general inventive concepts are directed to compositions and methods utilizing chondroitin, such as in the form of chondroitin sulfate, alone or in combination with additional functional ingredients such as Human Milk Oligosaccharides (HMOs), to promote gut health and particularly to promote healthy gut function in a formula-fed infant.

[0007] In certain exemplary embodiments, the general inventive concepts relate to a nutritional composition in either reconstitutable powder or liquid form for use in promoting gut health, and in particular infant gut health. When in liquid form, the nutritional composition comprises an effective amount of chondroitin, e.g. chondroitin sulfate. In some embodiments, the nutritional composition may also comprise at least one human milk oligosaccharide and optionally at least one nucleotide. The chondroitin and, if present, human milk oligosaccharides and nucleotides, may be useful to promote the growth of healthy bacteria, such as bifidobacteria, in the gut; strengthen the intestinal barrier; or both.

[0008] In certain exemplary embodiments, the general inventive concepts relate to methods for promoting gut health, and in particular infant gut health, by administering to an individual such as an infant, a nutritional composition that comprises an effective amount of chondroitin, e.g. chondroitin sulfate and, in some embodiments, at least one human milk oligosaccharide and / or at least one nucleotide. The methods for promoting infant gut health may include promoting the colonization of healthy bacteria, such as bifidobacteria, in the gut; strengthening the intestinal barrier; or both.Attorney Docket No. 15402WOO1 (67909WO01)

[0009] The promotion of a healthy gastrointestinal bacteria, e.g. one that is comparable to that of a breast-fed infant, and / or a strong intestinal barrier produces a number of key health benefits, including for instance aiding in digestion and / or nutrient absorption, increasing the production of beneficial metabolites (e.g. short chain fatty acids, vitamins, etc.), reducing the chances of developing allergies and / or food intolerances, and improving immunity and promoting healthy development of the immune system.

[0010] In some embodiments, the nutritional compositions, e.g. infant formulas, may by promoting a healthy balance of beneficial bacterial in the gastrointestinal tract be used to treat gut dysbiosis, thereby treating and / or preventing formula intolerance (e.g., fussiness, gas, crying, colic) or other gastrointestinal diseases and / or disorders resulting from suboptimal gastrointestinal flora population / balance and / or poor intestinal barrier functioning.

[0011] Administration of the nutritional compositions for improving gut maturation and gut health early in life may also reduce the risk of developing associated conditions and diseases of the gastrointestinal tract later in life. Diseases known to be associated with suboptimal gut maturation and / or gut dysbiosis include, but are not limited to, irritable bowel syndrome, colitis (e.g., necrotizing enterocolitis, Crohn's disease, ischemic colitis, cryptosporidium enterocolitis, pseudomembranous colitis, cytomegalovirus, ulcerative colitis), inflammatory bowel disease, celiac disease, colic, food intolerance, and food allergies. Thus, in certain exemplary embodiments, the general inventive concepts also contemplate nutritional compositions, e.g. infant formulas, for treating and / or reducing the risk of any of the above.

[0012] Embodiments of the present disclosure are directed to an infant formula comprising chondroitin, optionally in one or more sulfated forms generally known as chondroitin sulfate. The infant formula may also comprise one or more human milk oligosaccharides (HMOs), one or more nucleotides, or a combination thereof.

[0013] The chondroitin or chondroitin sulfate may be present in the formula in an amount of at least 0.1 g / L, optionally at least 0.2 g / L, optionally at least 0.3 g / L, when the infant formula is a liquid. In some embodiments, for instance, the chondroitin or chondroitin sulfate may be present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L andAttorney Docket No. 15402WOO1 (67909WO01) about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L of the infant formula when in liquid form.

[0014] In embodiments of the infant formula containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof. For instance, the one or more HMOs may comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N- acetylglucosamine oligosaccharide. In some embodiments, the fucosylated oligosaccharide may comprise at least one of 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL); the sialylated oligosaccharide may comprise at least one of 3’-sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide may comprise at least one of lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT).

[0015] In embodiments of the infant formula containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of 2′- fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and any combination thereof. In some embodiments, for instance, the one or more HMOs may comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

[0016] In some embodiments, the infant formula may contain total HMOs in an amount between 0.1 g / L and 10 g / L when the infant formula is a liquid.

[0017] In embodiments of the infant formula containing nucleotides, the one or more nucleotides may comprise at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof. In some embodiments, for example, the infant formula may contain all of adenosine, cytidine, guanosine, and uridine.

[0018] In some embodiments, the infant formula may contain total nucleotides in an amount between 15 mg / L and about 110 mg / L when the infant formula is a liquid.Attorney Docket No. 15402WOO1 (67909WO01)

[0019] In some embodiments, the infant formula may be a ready-to-drink liquid, also referred to as a ready-to-feed formula. In other embodiments, the infant formula may be in the form of a powder that is configured to be reconstituted into a liquid for feeding, i.e. a reconstitutable powder.

[0020] The infant formula may also contain sources of protein, carbohydrate, and fat. In some embodiments, for instance, the infant formula may comprise carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).

[0021] The infant formula may be configured to produce any of a variety of benefits. In some embodiments, for instance, the infant formula may be configured to promote (i.e., increase) the growth of beneficial intestinal bacteria, such as bifidobacteria. In some embodiments, the infant formula may be configured to promote the development of a beneficial intestinal microbiota comparable with that found in breast fed babies. In some embodiments, the infant formula may be configured to improve the feeding tolerance of an infant and / or to treat feeding intolerance in an infant. In some embodiments, the infant formula may be configured to strengthen the intestinal barrier, and thereby promote the development of a healthy gastrointestinal system and / or a healthy immune system.

[0022] Embodiments of the present disclosure are also directed to nutritional compositions for use in promoting the growth of beneficial intestinal bacteria, such as bifidobacteria, in a formula- fed infant. In some embodiments, the nutritional composition may promote, e.g. in a formula- fed infant, the development of a beneficial intestinal microbiota comparable with that found in a breast-fed infant. The nutritional composition comprises chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both. In some embodiments, the nutritional composition may be a follow-on-formula (FOF) or other composition configured to be consumed by an older infant or toddler.

[0023] Embodiments of the present disclosure are also directed to nutritional compositions for use in promoting a bifidogenic intestinal microbiota in a formula-fed infant. The nutritionalAttorney Docket No. 15402WOO1 (67909WO01) composition comprises chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

[0024] Embodiments of the present disclosure are also directed to nutritional compositions for use in promoting a strong intestinal barrier in an infant, such as a formula-fed infant. The nutritional composition comprises chondroitin, which may be in the form of chondroitin sulfate. The nutritional composition may also include one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

[0025] The chondroitin or chondroitin sulfate may be present in the nutritional composition in an amount of at least 0.1 g / L, optionally at least 0.2 g / L, optionally at least 0.3 g / L, when the nutritional composition is a liquid. In some embodiments, for instance, the chondroitin or chondroitin sulfate may be present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L of the nutritional composition when in liquid form.

[0026] In embodiments of the nutritional composition containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof. For instance, the one or more HMOs may comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N-acetylglucosamine oligosaccharide. In some embodiments, the fucosylated oligosaccharide may comprise at least one of 3-fucosyllactose (3-FL) and 2′- fucosyllactose (2′-FL); the sialylated oligosaccharide may comprise at least one of 3’- sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide may comprise at least one of lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT).

[0027] In embodiments of the nutritional composition containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of 2′- fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and anyAttorney Docket No. 15402WOO1 (67909WO01) combination thereof. In some embodiments, for instance, the one or more HMOs may comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

[0028] In some embodiments, the nutritional composition may contain total HMOs in an amount between 0.1 g / L and 10 g / L when the nutritional composition is a liquid.

[0029] In embodiments of the nutritional composition containing nucleotides, the one or more nucleotides may comprise at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof. In some embodiments, for example, the nutritional composition may contain all of adenosine, cytidine, guanosine, and uridine.

[0030] In some embodiments, the nutritional composition may contain total nucleotides in an amount between 15 mg / L and about 110 mg / L when the nutritional composition is a liquid.

[0031] In some embodiments, the nutritional composition may be a ready-to-drink liquid. In other embodiments, the nutritional composition may be in the form of a powder that is configured to be reconstituted into a liquid for feeding, i.e. a reconstitutable powder. In some embodiments, the nutritional composition may be an infant formula.

[0032] In some embodiments, the nutritional composition may also contain sources of protein, carbohydrate, and fat. In some embodiments, for instance, the nutritional composition may comprise carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis)..

[0033] Embodiments of the present disclosure are also directed to methods for promoting the growth of beneficial intestinal bacteria, such as bifidobacteria, in a formula-fed infant, the method comprising administering to an infant a nutritional composition comprising chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both. For instance, embodiments of the present disclosure are directed to method for promoting, in a formula-fed infant, the development of a beneficial intestinal microbiota comparable with that found in a breast-fed infant, the method comprising administering to an infant a nutritional composition comprising chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milkAttorney Docket No. 15402WOO1 (67909WO01) oligosaccharides (HMOs), one or more nucleotides, or both. Embodiments of the present disclosure are directed to methods for promoting a bifidogenic intestinal microbiota in an infant, e.g. in a formula-fed infant, the method comprising administering to an infant a nutritional composition comprising chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

[0034] Embodiments of the present disclosure are also directed to methods for promoting development of a strong intestinal barrier, the method comprising administering to an infant a nutritional composition comprising chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

[0035] Embodiments of the present disclosure are also directed to methods for improving feeding tolerance and / or treating feeding intolerance, the method comprising administering to an infant a nutritional composition comprising chondroitin, which may be in the form of chondroitin sulfate, as well as optionally one or more human milk oligosaccharides (HMOs), one or more nucleotides, or both.

[0036] The chondroitin or chondroitin sulfate may be present in the nutritional composition in an amount of at least 0.1 g / L, optionally at least 0.2 g / L, optionally at least 0.3 g / L, when the nutritional composition is a liquid. In some embodiments, for instance, the chondroitin or chondroitin sulfate may be present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L of the nutritional composition when in liquid form.

[0037] In embodiments of the nutritional composition containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof. For instance, the one or more HMOs may comprises a combination of at least one fucosylated oligosaccharide, at least one sialylatedAttorney Docket No. 15402WOO1 (67909WO01) oligosaccharide, and at least one N-acetylglucosamine oligosaccharide. In some embodiments, the fucosylated oligosaccharide may comprise at least one of 3-fucosyllactose (3-FL) and 2′- fucosyllactose (2′-FL); the sialylated oligosaccharide may comprise at least one of 3’- sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide may comprise at least one of lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT).

[0038] In embodiments of the nutritional composition containing one or more HMOs, the one or more HMOs may, in some instances, comprise at least one selected from the group of 2′- fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and any combination thereof. In some embodiments, for instance, the one or more HMOs may comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

[0039] In some embodiments, the nutritional composition may contain total HMOs in an amount between 0.1 g / L and 10 g / L when the nutritional composition is a liquid.

[0040] In embodiments of the nutritional composition containing nucleotides, the one or more nucleotides may comprise at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof. In some embodiments, for example, the nutritional composition may contain all of adenosine, cytidine, guanosine, and uridine.

[0041] In some embodiments, the nutritional composition may contain total nucleotides in an amount between 15 mg / L and about 110 mg / L when the nutritional composition is a liquid.

[0042] In some embodiments, the nutritional composition may be a ready-to-drink liquid. In other embodiments, the nutritional composition may be in the form of a powder that is configured to be reconstituted into a liquid for feeding, i.e. a reconstitutable powder. In some embodiments, the nutritional composition may be an infant formula.

[0043] In some embodiments, the nutritional composition may also contain sources of protein, carbohydrate, and fat. In some embodiments, for instance, the nutritional composition may comprise carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).Attorney Docket No. 15402WOO1 (67909WO01) BRIEF DESCRIPTION OF THE DRAWINGS

[0044] The general inventive concepts, as well as embodiments and advantages thereof, are described below in greater detail, by way of example, with reference to the drawings in which:

[0045] Figure 1 is a graph showing the results of Example 1, which demonstrate the ability of chondroitin sulfate to promote the adhesion of bifidobacterium, e.g. B. infantis 15697, to intestinal cells, e.g. HT-29 intestinal epithelial cells.

[0046] Figure 2 is a graph showing the results of Example 2, which demonstrate a strong synergistic effect of a combination of chondroitin sulfate, human milk oligosaccharides, and nucelotides on intestinal barrier function.

[0047] Figure 3 is a graph showing additional results of Example 2, which demonstrate that the strong synergistic effect observed in Figure 2 relates to the combination of chondroitin sulfate and human milk oligosaccharides.

[0048] Figure 4 is graph showing the results of Example 3, demonstrating a synergistic effect of a combination of chondroitin sulfate and human milk oligosaccharides on reducing the adhesion of pathogenic bacteria, e.g. E. coli 35401, to intestinal cells. DETAILED DESCRIPTION

[0049] The compositions and methods described herein utilize chondroitin sulfate, optionally in combination with other functional ingredient(s) such as human milk oligosaccharides (HMOs), nucleotides, or a combination thereof, for promoting intestinal, i.e. gut, health as described in greater detail herein. The compositions, including nutritional compositions, described herein include synthetic formulas that comprise chondroitin sulfate, optionally in combination with other functional ingredient(s) such as human milk oligosaccharides (HMOs), nucleotides, or a combination thereof. These and other features of the compositions and methods, as well as some of the many optional variations and additions, are described in detail hereafter.

[0050] The terms “nutritional formulation” or “nutritional composition” as used herein, are used interchangeably and, unless otherwise specified, refer to synthetic formulas including nutritional liquids, nutritional powders, nutritional solids, nutritional semi-solids, nutritionalAttorney Docket No. 15402WOO1 (67909WO01) semi-liquids, nutritional supplements, and any other nutritional food product as known in the art. The nutritional powders may be reconstituted to form a nutritional liquid, all of which comprise one or more of fat, protein and carbohydrate and are suitable for oral consumption by a human and may be used as a sole or supplemental source of nutrition. The terms “nutritional formulation” or “nutritional composition” do not include human breast milk and do not refer to supplemented milk and are generally shelf stable.

[0051] The term “nutritional liquid” as used herein, unless otherwise specified, refers to nutritional compositions in ready-to-drink liquid form, concentrated form, and nutritional liquids made by reconstituting the nutritional powders described herein prior to use.

[0052] The term “nutritional powder” as used herein, unless otherwise specified, refers to nutritional compositions in flowable or scoopable form that can be reconstituted with water or another aqueous liquid prior to consumption and includes both spray dried and dry mixed / dry blended powders.

[0053] The term “nutritional semi-solid,” as used herein, unless otherwise specified, refers to nutritional compositions that are intermediate in properties, such as rigidity, between solids and liquids. Some semi-solids examples include puddings, gelatins, and doughs.

[0054] The term “nutritional semi-liquid,” as used herein, unless otherwise specified, refers to nutritional compositions that are intermediate in properties, such as flow properties, between liquids and solids. Some semi-liquids examples include thick shakes and liquid gels.

[0055] The terms “fat” and “oil” as used herein, unless otherwise specified, are used interchangeably to refer to lipid materials derived or processed from plants or animals. These terms also include synthetic lipid materials so long as such synthetic materials are suitable for oral administration to humans.

[0056] The term “shelf stable” as used herein, unless otherwise specified, refers to a nutritional product that remains commercially stable after being packaged and then stored at 18-24° C. for at least 3 months, including from about 6 months to about 24 months, and also including from about 12 months to about 18 months.Attorney Docket No. 15402WOO1 (67909WO01)

[0057] The term “individual” as used herein, refers generally to a preterm infant, infant, toddler, child, or adult.

[0058] The term “infant” as used herein, refers generally to individuals up to age 12 months of age, actual or corrected.

[0059] The term “preterm infant” as used herein, refers to those infants born at less than 37 weeks gestation, have a birth weight of less than 2500 gm, or both.

[0060] The term “beneficial intestinal microbiota comparable with that found in breast fed babies” means an intestinal microbiota dominated by appreciable populations of Bifidobacterium, Bacteroides, and Lactobacillus species to the exclusion of appreciable populations of such species as Escherichia coli and Streptococci.

[0061] The term “bifidogenic intestinal microbiota” means an intestinal microbiota which is dominated by the Bifidobacterium genus such as Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium longum and which is generally comparable with that found in a breast fed infant of the same age.

[0062] As used herein, all concentrations expressed as either “µg / liter,” “mg / liter,” “mg / L,” “mcg / L,” “g / L,” etc., refer to ingredient concentrations within the described nutritional compositions as calculated on an as-fed basis (e.g., reconstituted for consumption in the case of nutritional powders), unless otherwise specified.

[0063] The term “reconstitute” or various other forms such as “reconstituted” or “reconstituting” all refer to the general act of adding a suitable amount of liquid, typically water, to a form of nutritional formulation that is not in its ready-to-drink liquid form, such as nutritional powder or a concentrated form of a nutritional liquid, thereby making the nutritional composition ready-to-drink.

[0064] The terms “susceptible” and “at risk” as used herein, unless otherwise specified, mean having little resistance to a certain condition or disease relative to the general population, including being genetically predisposed, having a family history of, and / or having symptoms ofAttorney Docket No. 15402WOO1 (67909WO01) the condition or disease. The term refers to those having a vulnerability higher than the general population.

[0065] The terms “treating” and “treatment” as used herein, unless otherwise specified, includes delaying the onset of a condition, reducing the severity of symptoms of a condition, or eliminating some or all of the symptoms of a condition.

[0066] All percentages, parts and ratios as used herein, are by weight of the total composition, unless otherwise specified. All such weights, as they pertain to listed ingredients, are based on the active level and, therefore, do not include solvents or by-products that may be included in commercially available materials, unless otherwise specified.

[0067] Numerical ranges as used herein are intended to include every number and subset of numbers within that range, whether specifically disclosed or not. Further, these numerical ranges should be construed as providing support for a claim directed to any number or subset of numbers in that range. For example, a disclosure of from 1 to 10 should be construed as supporting a range of from 2 to 8, from 3 to 7, from 5 to 6, from 1 to 9, from 3.6 to 4.6, from 3.5 to 9.9, and so forth.

[0068] All references to singular characteristics or limitations of the general inventive concepts shall include the corresponding plural characteristic or limitation, and vice versa, unless otherwise specified or clearly implied to the contrary by the context in which the reference is made.

[0069] All combinations of method or process steps as used herein can be performed in any order, unless otherwise specified or clearly implied to the contrary by the context in which the referenced combination is made.

[0070] The chondroitin sulfate and, if present, HMOs and / or nucleotides, can be formulated in suitable compositions and then, in accordance with the methods of the invention, administered to an individual in a form adapted to the chosen route of administration. The formulations include, but are not limited to, those suitable for oral administration. Oral administration, as defined herein, includes any form of administration in which the composition passes through the esophagus of the patient. For example, oral administration includes nasogastric intubation, inAttorney Docket No. 15402WOO1 (67909WO01) which a tube is run from through the nose to the stomach of the patient to administer food or drugs.

[0071] Oral formulations of the compositions disclosed herein include any solid, liquid, or powder formulation suitable for use herein, provided that such a formulation allows for the safe and effective oral delivery of the chondroitin sulfate and, if present, HMOs and / or nucleotides, and optional nutritive components. In preferred embodiments, the oral formulation is a liquid nutritional composition or reconstitutable powder. Formulations of the present invention suitable for oral administration may be presented as discrete units such as tablets, troches, capsules, lozenges, wafers, or cachets, each containing a predetermined amount of the chondroitin sulfate and, if present, HMOs and / or nucelotides, as a powder or granules or as a solution or suspension in an aqueous liquor or non-aqueous liquid such as a syrup, an elixir, an emulsion, or a draught.

[0072] The nutritional compositions and methods may comprise, consist of, or consist essentially of the essential elements of the compositions and methods as described herein, as well as any additional or optional element described herein or otherwise useful in oral applications. Chondroitin / Chondroitin Sulfate

[0073] In an aspect of the general inventive concepts, the compositions and methods of the present disclosure include chondroitin, which is most commonly found in a one or more of a variety of sulfated forms (including, for example, GlcAβ1-3GalNAc4S (Type A), IdoAα1- 3GalNAc4S (Type B), GlcAβ1-3GalNAc6S (Type C), GlcA2Sβ1-3GalNAc6S (Type D), GlcAβ1- 3GalNAc4,6diS (Type E), etc.) which are all generally and collectively known as chondroitin sulfate. Chondroitin sulfate (CS) is a glycosaminoglycan (GAG) widely present in the extracellular matrix of animal cartilaginous tissues and cell surfaces. It is also the most prominent GAG component of breastmilk (55% of total GAGs), with lower concentrations found in bovine milk (21% of total GAGs). The total chondroitin sulfate concentration of human milk has been reported as 231 mg / L, while the concentration of chondroitin sulfate in fresh bovine milk is 12.9 mg / L. Chondroitin may also be utilized in an unsulfated form.

[0074] The general inventive concepts are based in part on the discovery that chondroitin sulfate shows the ability to promote adhesion of beneficial bacteria, such as bifidobacteria, to theAttorney Docket No. 15402WOO1 (67909WO01) intestinal cell surface in vitro. Adherence to epithelial cell surfaces is a necessary step to initiate colonization of the gastrointestinal tract. Therefore, a promotion in adhesion of this beneficial bacteria would produce a more desirable intestinal microflora population, and may render the intestinal microflora population of a formula-fed infant more comparable to that of a breast-fed infant. Because chondroitin sulfate is known to have an effect of reducing the adhesion of pathogenic bacteria to the intestinal cell surface, the present discovery that chondroitin sulfate increases the adhesion of beneficial bifidobacteria is highly surprising.

[0075] Additionally, Applicants demonstrate the unexpected synergistic effect between chondroitin sulfate and HMOs whereby a greater prevention of pathogenic E. coli adhesion to the intestinal cell surface is observed when used in combination. By reducing the adhesion of pathogenic bacteria such as E. coli and promoting the adhesion of beneficial bacteria, such as bifidobacteria, the administration of a combination of chondroitin sulfate and HMOs may have an even great effect on promoting a healthy intestinal microflora (and achieving the various health benefits associated therewith).

[0076] The general inventive concepts are also based in part on the unexpected synergistic effect between chondroitin sulfate and HMOs on the functioning of the intestinal barrier. By improving the functioning of the intestinal barrier, the administration of a combination of chondroitin sulfate and HMOs reduces the chances of developing allergies and / or food intolerances and promotes a healthy immune system. Strengthening / improving the functioning of the intestinal barrier may also generally be considered improving gut health.

[0077] In certain exemplary aspects the nutritional compositions and methods contemplate providing chondroitin or chondroitin sulfate in an amount of at least 0.1 g / L of the composition, when in liquid form, alternative at least 0.2 g / L of the composition, alternatively at least 0.3 g / L of the composition, alternatively at least 0.4 g / L of the composition, alternatively at least 0.5 g / L of the composition. In some embodiments, the nutritional composition, when in liquid form, may include chondroitin or chondroitin sulfate in an amount between about 0.1 g / L and about 3 g / L, alternatively between about 0.1 g / L and about 2 g / L, alternatively between about 0.1 g / L and about 1.5 g / L, alternatively between about 0.1 g / L and about 1.0 g / L, alternatively between about 0.2 g / L and about 3 g / L, alternatively between about 0.2 g / L and about 2 g / L,Attorney Docket No. 15402WOO1 (67909WO01) alternatively between about 0.2 g / L and about 1.5 g / L, alternatively between about 0.2 g / L and about 1.0 g / L, alternatively between about 0.3 g / L and about 3 g / L, alternatively between about 0.3 g / L and about 2 g / L, alternatively between about 0.3 g / L and about 1.5 g / L, alternatively between about 0.3 g / L and about 1.0 g / L, alternatively between about 0.4 g / L and about 3 g / L, alternatively between about 0.4 g / L and about 2 g / L, alternatively between about 0.4 g / L and about 1.5 g / L, alternatively between about 0.4 g / L and about 1.0 g / L, alternatively between about 0.5 g / L and about 3 g / L, alternatively between about 0.5 g / L and about 2 g / L, alternatively between about 0.5 g / L and about 1.5 g / L, alternatively between about 0.5 g / L and about 1.0 g / L.

[0078] Using a specific reconstitution ratio, one can calculate the amount of chondroitin or chondroitin sulfate in a powdered composition, e.g. by weight percentage. As is understood in the art, higher caloric density formulas can be also prepared by using a higher ratio of powder to water or lower caloric density formulas can be prepared by using a lower ratio of powder to water. The ratio of powder to water to produce a formula of a desired caloric density will also depend on the specific composition, e.g. macronutrient distribution, of the formula.

[0079] Using an example reconstitution ratio of about 145 g / L, one can calculate the amount of chondroitin or chondroitin sulfate in a powdered composition, e.g. by weight percentage, that would provide a liquid concentration of chondroitin or chondroitin sulfate in the above ranges. In certain aspects, the composition of the present disclosure, when in reconstitutable powder form, may comprise chondroitin or chondroitin sulfate of from about 0.06 wt.% to about 2.0 wt.% based on the total weight of the composition. In certain aspects, the composition of the present disclosure, when in powder form, may comprise chondroitin or chondroitin sulfate of from about 0.1 wt.% to about 1.4 wt.% based on the total weight of the nutritional composition, including from about 0.2 wt.% to about 0.7 wt.%, including from about .06 wt.% to about 1.4 wt.%, including from about 0.06 wt.% to about 0.7 wt.%, including from about 0.1 wt.% to about 2.0 wt.%, including from about 0.1 wt.% to about 0.7 wt.%, including from about 0.2 wt.% to about 2.0 wt.%, including from about 0.2 wt.% to about 1.4 wt.%, and also including from about 0.1 wt.% to about 1.0 wt.% and from about 0.2 wt.% to about 1.0 wt.%, based on the total weight of the nutritional composition.Attorney Docket No. 15402WOO1 (67909WO01) Human Milk Oligosaccharides (HMOs)

[0080] In one aspect, the compositions of the present disclosure include an HMO in combination with the chondroitin sulfate. In certain aspects, the compositions include a mixture of HMOs comprising at least one neutral HMO and at least one acidic HMO.

[0081] In certain aspects, the compositions of the present disclosure include an HMO or a mixture of HMOs (e.g., a neutral HMO and an acidic HMO). The HMOs may be isolated or enriched from milk(s) secreted by mammals including, but not limited to: human, bovine, ovine, porcine, or caprine species. The HMOs may also be produced via microbial fermentation, enzymatic processes, chemical synthesis, or combinations thereof.

[0082] Suitable HMOs for use in the compositions of the present disclosure may include neutral oligosaccharides, n-acetyl glucosylated oligosaccharides, acidic HMOs, and HMO precursors. For example, specific non-limiting suitable HMOs that may be included in the compositions according to the present disclosure include: 2’-fucosyllactose (2’-FL), 3-fucosyllactose (3-FL), Difucosyllactose (DFL), 3’-sialyllactose (3’-SL), 6’-sialyllactose (6’-SL), lacto-N-neotetraose (LNnT), and lacto-N-tetraose (LNT).

[0083] Other HMOs that may be included in certain aspects of the compositions of the present disclosure include: N-acetylglucosamine (GlcNAc); L-fucose (L-Fuc); D-fucose (D-Fuc); fucosyl oligosaccharides (i.e., Lacto-N-fucopentaose I; Lacto-N-fucopentaose II; Lacto-N-fucopentaose III; Lacto-N-difucohexaose I; and Lactodifucotetraose); sialyl fucosyl oligosaccharides (i.e., 3′- Sialyl-3-fucosyllactose; Disialomonofucosyllacto-N-neohexaose; Monofucosylmonosialyllacto- N-octaose (sialyl Lea); Sialyllacto-N-fucohexaose II; Disialyllacto-N-fucopentaose II; Monofucosyldisialyllacto-N-tetraose); and sialyl oligosaccharides (i.e., 3-Sialyllactosamine; 6′- Sialyllactosamine; Sialyllacto-N-neotetraose c; Monosialyllacto-N-hexaose; Disialyllacto-N- hexaose I; Monosialyllacto-N-neohexaose I; Monosialyllacto-N-neohexaose II; Disialyllacto-N- neohexaose; Disialyllacto-N-tetraose; Disialyllacto-N-hexaose II; Sialyllacto-N-tetraose a; Disialyllacto-N-hexaose I; and Sialyllacto-N-tetraose b). Also useful are variants in which the glucose (Glc at the reducing end is replaced by N-acetylglucosamine (e.g., 2′-fucosyl-N- acetylglucosamine (2′-FLNac) is such a variant to 2′-fucosyllactose). These HMOs are described more fully in U.S. Patent Application No. 2009 / 0098240, which is herein incorporated byAttorney Docket No. 15402WOO1 (67909WO01) reference in its entirety. Other suitable examples of HMOs that may be included in the compositions according to the present disclosure include lacto-N-fucopentaose V, lacto-N- hexaose, para-lacto-N-hexaose, lacto-N-neohexaose, para-lacto-N-neohexaose, monofucosyllacto-N-hexaose II, isomeric fucosylated lacto-N-hexaose (1), isomeric fucosylated lacto-N-hexaose (3), isomeric fucosylated lacto-N-hexaose (2), difucosyl-para-lacto-N- neohexaose, difucosyl-para-lacto-N-hexaose, difucosyllacto-N-hexaose, lacto-N-neoocataose, para-lacto-N-octanose, iso-lacto-N-octaose, lacto-N-octaose, monofucosyllacto-neoocataose, monofucosyllacto-N-ocataose, difucosyllacto-N-octaose I, difucosyllacto-N-octaose II, difucosyllacto-N-neoocataose II, difucosyllacto-N-neoocataose I, lacto-N-decaose, trifucosyllacto-N-neooctaose, trifucosyllacto-N-octaose, trifucosyl-iso-lacto-N-octaose, lacto-N- difuco-hexaose II, sialyl-lacto-N-tetraose a, sialyl-lacto-N-tetraose b, sialyl-lacto-N-tetraose c, sialyl-fucosyl-lacto-N-tetraose I, sialyl-fucosyl-lacto-N-tetraose II, and disialyl-lacto-N-tetraose, and combinations thereof.

[0084] The HMOs may be present in the composition, when in liquid form (e.g., ready-to-feed form), in total amounts of HMO in the composition (mg of HMO per mL of composition) of at least about about 0.1 g / L, alternatively at least about 0.2 g / L, alternatively at least about 0.5 g / L, alternatively at least about 1.0 g / L, alternatively at least about 1.5 g / L, alternatively at least about 2.0 g / L, alternatively at least about 2.5 g / L. In some embodiments, the nutritional composition may contain total HMOs in an amount of from about 0.1 g / L to about 20 g / L, including about 0.2 g / L to about 10 g / L, about 1 g / L to about 10 mg / mL, about 2 g / L to about 10 g / L, about 2.5 g / L to about 10 g / L, about 0.1 to about 7.5 g / L, about 0.2 g / L to about 7.5 g / L, about 1 g / L to about 7.5 g / L, about 2 g / L to about 7.5 g / L, and about 2.5 g / L to about 7.5 g / L, , about 0.1 to about 5 g / L, about 0.2 g / L to about 5 g / L, about 1 g / L to about 5 g / L, about 2 g / L to about 5 g / L, and about 2.5 g / L to about 5 g / L. The exact amount of HMO in the composition may depend on the specific HMO or HMOs present and the amounts of other components in the composition.

[0085] In certain aspects of the present disclosure, the composition may include an acidic HMO, and a neutral HMO comprising a fucosylated HMO and an N-acetylated HMO. In certain aspects of the present disclosure, the composition comprises a combination of 2’-FL, 3-FL, DFL, 3’-SL, 6’-SL, and LNT. In certain aspects of the present disclosure, the composition (when in ready-to- feed liquid form) may comprise 2’-FL in an amount up to 4.15 mg / mL, LNT in an amount up toAttorney Docket No. 15402WOO1 (67909WO01) 2.11 mg / mL, 3-FL in an amount up to 1.17 mg / mL, 3’-SL in an amount up to 0.36 mg / mL, and 6’-SL in an amount up to 0.44 mg / mL.

[0086] In certain aspects, the composition of the present disclosure, when in powder form, may comprise a total amount of HMOs of from about 1 wt.% to about 10 wt.% based on the total weight of the composition. In certain aspects, the composition of the present disclosure, when in powder form, may comprise a total amount of HMOs of from about 1 wt.% to about 8 wt.% based on the total weight of the nutritional composition, including from about 1.5 wt.% to about 7.5 wt.%, including from about 2 wt.% to about 7 wt.%, including from about 2.5 wt.% to about 6.5 wt.%, and also including from about 3 wt.% to about 6 wt.% based on the total weight of the nutritional composition. Nucleotides

[0087] In addition to the chondroitin sulfate and HMOs, the nutritional compositions according to the general inventive concepts may additionally comprise nucleotides and / or nucleotide precursors selected from the group consisting of nucleosides, purine bases, pyrimidine bases, ribose and deoxyribose. The nucleotide may be in monophosphate, diphosphate, or triphosphate form. The nucleotide may be a ribonucleotide or a deoxyribonucleotide. The nucleotides may be monomeric, dimeric, or polymeric (including RNA and DNA). The nucleotide may be present in the nutritional composition as a free acid or in the form of a salt, preferably a monosodium salt.

[0088] Suitable nucleotides and / or nucleosides for use in the nutritional compositions include one or more of cytidine 5′-monophosphate, uridine 5′-monophosphate, adenosine 5′- monophosphate, guanosine 5′-1-monophosphate, and / or inosine 5′-monophosphate, more preferably cytidine 5′-monophosphate, uridine 5′-monophosphate, adenosine 5′-monophosphate, guanosine 5′-monophosphate, and inosine 5′-monophosphate.

[0089] When present, the nutritional composition may include nucleotides in a total amount of about 15 mg / L to about 150 mg / mL, including about 15 mg / L to about 110 mg / L, including about 20 mg / L to about 110 mg / L of the nutritional composition, including about 30 mg / L to about 100 mg / L of the nutritional composition, including about 50 mg / L to about 100 mg / L of the nutritional composition, including about 15 mg / L to about 100 mg / mL, including about 15 mg / L to about 80Attorney Docket No. 15402WOO1 (67909WO01) mg / mL, including about 30 mg / L to about 90 mg / L, including about 40 mg / L to about 90 mg / mL of the nutritional composition. In order to obtain a concentration within this range when reconstituted with water, the nutritional composition may include nucleotides in an amount of about 0.01 wt. % to about 0.10 wt. % of the nutritional composition when the nutritional composition is a reconstitutable powder.

[0090] In certain exemplary aspects of the present disclosure, the nucleotides may comprise about 40-50% cytidine 5’monophosphate, about 15-20% uridine 5′-monophosphate, about 12-18% adenosine 5′-monophosphate and about 20-25% guanosine 5′-monophosphate by total weight of nucleotides. In certain exemplary aspects of the present disclosure, the nucleotides may comprise about 43% cytidine 5′-monophosphate, about 18.5% uridine 5′-monophosphate, about 16.5% adenosine 5′-monophosphate and about 22% guanosine 5′-monophosphate by total weight of nucleotides.

[0091] In an exemplary aspect, the nucleotides may be present in the nutritional compositions in a total amount of about 50 mg / L to about 150 mg / mL or more of the nutritional composition and comprise about 25 to 39 mg / L of cytidine 5′-monophosphate; 10 to 21 mg / L of uridine 5′- monophosphate; 10 to 15 mg / L of adenosine 5′-monophosphate; and 12 to 20 mg / L of guanosine 5′-monophosphate.

[0092] In an exemplary aspect, the nucleotides may be present in the weight ratio of cytidine 5′- monophosphate: uridine 5′-monophosphate is from about 1.5:1 to about 2.6:1; of cytidine 5′- monophosphate: adenosine 5′-monophosphate is from about 2:1 to about 3.9:1; and of cytidine 5′- monophosphate: guanosine 5′-monophosphate is from about 1.75:1 to about 2.8:1.

[0093] In certain embodiments, the nutritional composition includes a combination of the chondroitin sulfate, HMOs, and nucleotides as described herein such that the composition provides a synergistic benefit to the end user, such as a synergistic benefit in strengthening the intestinal barrier. Macronutrients

[0094] In addition to the specific functional ingredients described herein, the nutritional composition may in many embodiments, such as where the nutritional composition is an infantAttorney Docket No. 15402WOO1 (67909WO01) formula, include one or more ingredients that help satisfy the individual’s nutritional requirements. The optional nutrients can provide up to about 1000 kcal of energy per serving or dose, including from about 25 to about 900 kcal, from about 75 to about 700 kcal, from about 150 to about 500 kcal, from about 350 to about 500 kcal, or from about 200 to about 300 kcal.

[0095] The nutritional compositions may be formulated with sufficient kinds and amounts of nutrients to provide a sole, primary, or supplemental source of nutrition, or to provide a specialized nutritional product for use in individuals afflicted with specific diseases or conditions or with a targeted nutritional benefit as described below.

[0096] The nutritional compositions may be formulated to include at least one of protein, fat, and carbohydrate. In many embodiments, the nutritional compositions will include protein, carbohydrate and fat. Although total concentrations or amounts of the fat, protein, and carbohydrates may vary depending upon the product type (i.e., nutritional formula), product form (i.e., nutritional solid, powder, ready-to-feed liquid, or concentrated liquid) and targeted dietary needs of the intended user, such concentrations or amounts most typically fall within one of the following embodied ranges, inclusive of any other essential fat, protein, and / or carbohydrate ingredients as described herein. Nutritional Compositions

[0097] Nutritional compositions, as discussed herein, include at least one of fat, carbohydrate, and protein in addition to the chondroitin sulfate and optionally HMOs and / or nucleotides. Infant formulas, which are configured to provide sole source nutrition, will typically contain a combination of fat, carbohydrate, and protein.

[0098] Where present, carbohydrate concentrations most typically will range from about 10% to about 65%, including from about 20% to about 60%, including from about 30% to about 60%, by dry weight of the nutritional composition. Where present, fat concentrations most typically range from about 10% to about 45%, including from about 15% to about 40%, and also including from about 20% to about 40%, by dry weight of the nutritional composition. Where present, protein concentrations most typically range from about 5% to about 30%, including from about 10% to about 30%, and also including from about 15% to about 25%, by dry weight of the nutritionalAttorney Docket No. 15402WOO1 (67909WO01) composition. In some embodiments, an infant formula may include from about 40% to about 60% carbohydrates, from about 10% to about 25% protein, and from about 20% to about 40% fat by dry weight of the infant formula.

[0099] The amount of any or all of the carbohydrates, fats, and proteins in any of the nutritional compositions (e.g., infant formula) described herein may also be characterized as a percentage of total calories in the nutritional composition. In some embodiments, the nutritional composition (e.g. infant formula) may contain carbohydrates in an amount of from about 35% to about 50% as a percentage of total calories, protein in an amount of from about 5% to about 20% as a percentage of total calories, and fat in an amount of from about 35% to about 55% as a percentage of total calories. Fat

[0100] The nutritional compositions according to the general inventive concepts may comprise a source or sources of fat. Suitable sources of fat for use herein include any fat or fat source that is suitable for use in an oral nutritional product and is compatible with the essential elements and features of such products. Most typically the fat may be an emulsified fat. In an exemplary embodiment, the fat is derived from short chain fatty acids.

[0101] Non-limiting examples of suitable fats or sources thereof for use in the nutritional products described herein include coconut oil, fractionated coconut oil, soybean oil, corn oil, olive oil, safflower oil, high oleic safflower oil, oleic acids (EMERSOL 6313 OLEIC ACID, Cognis Oleochemicals, Malaysia), MCT oil (medium chain triglycerides), sunflower oil, high oleic sunflower oil, palm and palm kernel oils, palm olein, canola oil, marine oils, fish oils, fungal oils, algae oils, cottonseed oils, and combinations thereof. Lipid sources of arachidonic acid and docosahexaenoic acid include, but are not limited to, marine oil, egg yolk oil, and fungal or algal oil.

[0102] Numerous commercial sources for these fats are readily available and known to one practicing the art. For example, soy and canola oils are available from Archer Daniels Midland of Decatur, Ill. Corn, coconut, palm and palm kernel oils are available from Premier Edible Oils Corporation of Portland, Organ. Fractionated coconut oil is available from Henkel CorporationAttorney Docket No. 15402WOO1 (67909WO01) of LaGrange, Ill. High oleic safflower and high oleic sunflower oils are available from SVO Specialty Products of Eastlake, Ohio. Marine oil is available from Mochida International of Tokyo, Japan. Olive oil is available from Anglia Oils of North Humberside, United Kingdom. Sunflower and cottonseed oils are available from Cargil of Minneapolis, Minn. Safflower oil is available from California Oils Corporation of Richmond, Calif.

[0103] In addition to these food grade oils, structured lipids may be incorporated into the food product if desired. Structured lipids are known in the art. A concise description of structured lipids can be found in INFORM, Vol. 8, No. 10, page 1004; entitled Structured lipids allow fat tailoring (October 1997). Also see U.S. Pat. No. 4,871,768. Structured lipids are predominantly triacylglycerols containing mixtures of medium and long chain fatty acids on the same glycerol nucleus. Structured lipids and their use in enteral formula are also described in U.S. Pat. Nos. 6,194,379 and 6,160,007.

[0104] Optionally, ω-3 fatty acids may comprise up to approximately 5% of the oil blend, preferably the ω-3 fatty acids largely consist of the longer chain forms, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Dietary oils used in the preparation of the nutritional composition generally contain ω-3 fatty acids in the triglyceride form and include, but are not limited to canola, medium chain triglycerides, fish, soybean, soy lecithin, corn, safflower, sunflower, high-oleic sunflower, high-oleic safflower, olive, borage, black currant, evening primrose and flaxseed oil. Optionally, the weight ratio of ω-6 fatty acids to ω-3 fatty acids in the lipid blend according to the invention is about 0.1 to 3.0. The daily delivery of ω-3 fatty acids should be at least 450 mg and may vary depending on body weight, sex, age and medical condition of the individual. As mentioned, higher levels are desired for adult human consumption: for example, from about 0.5 to 50 gm daily, more preferably from about 2.5 to 5 gm daily Protein

[0105] In certain exemplary embodiments, the nutritional compositions according to the general inventive concepts include protein. Any protein source that is suitable for use in oral nutritional compositions and is compatible with the essential elements and features of the general inventive concepts may be used.Attorney Docket No. 15402WOO1 (67909WO01)

[0106] Non-limiting examples of suitable proteins or sources thereof for use in the nutritional compositions include hydrolyzed, partially hydrolyzed or non-hydrolyzed proteins or protein sources, which may be derived from any known or otherwise suitable source such as milk (e.g., casein, whey), animal (e.g., meat, fish), cereal (e.g., rice, corn), vegetable (e.g., soy) or combinations thereof. Non-limiting examples of such proteins include milk protein isolates, milk protein concentrates as described herein, casein protein isolates, extensively hydrolyzed casein, whey protein, sodium or calcium caseinates, whole cow milk, partially or completely defatted milk, soy protein isolates, and soy protein concentrates.

[0107] In an exemplary embodiment, the protein source is a hydrolyzed protein, i.e., a protein hydrolysate. In this context, the terms “hydrolyzed protein” or “protein hydrolysates” are used interchangeably herein and include extensively hydrolyzed proteins, wherein the degree of hydrolysis is most often at least about 20%, including from about 20% to about 80%, and also including from about 30% to about 80%, even more preferably from about 40% to about 60%. The degree of hydrolysis is the extent to which peptide bonds are broken by a hydrolysis method. The degree of protein hydrolysis for purposes of characterizing the extensively hydrolyzed protein component of these embodiments is easily determined by one of ordinary skill in the formulation arts by quantifying the amino nitrogen to total nitrogen ratio (AN / TN) of the protein component of the selected liquid formulation. The amino nitrogen component is quantified by USP titration methods for determining amino nitrogen content, while the total nitrogen component is determined by the Tecator Kjeldahl method, all of which are well known methods to one of ordinary skill in the analytical chemistry art.

[0108] Suitable hydrolyzed proteins include soy protein hydrolysate, casein protein hydrolysate, whey protein hydrolysate, rice protein hydrolysate, potato protein hydrolysate, fish protein hydrolysate, egg albumen hydrolysate, gelatin protein hydrolysate, combinations of animal and vegetable protein hydrolysates, and combinations thereof. Particularly preferred protein hydrolysates include whey protein hydrolysate and hydrolyzed sodium caseinate.

[0109] In some embodiments, the protein source may include at least about 20% (by weight total protein) protein hydrolysate, including from about 30% to 100% (by weight total protein)Attorney Docket No. 15402WOO1 (67909WO01) protein hydrolysate, and including from about 40% to about 80% (by weight total protein) protein hydrolysate, and including about 50% (by weight total protein) protein hydrolysate. Carbohydrate

[0110] In an exemplary embodiment, the nutritional compositions according to the general inventive concepts include carbohydrates that are suitable for use in an oral nutritional composition and are compatible with the essential elements and features of the general inventive concepts.

[0111] Non-limiting examples of suitable carbohydrates or sources thereof for use in the nutritional compositions described herein include maltodextrin, hydrolyzed or modified starch or cornstarch, glucose polymers, corn syrup, corn syrup solids, rice-derived carbohydrates, pea- derived carbohydrates, potato-derived carbohydrates, tapioca, sucrose, glucose, fructose, lactose, high fructose corn syrup, honey, sugar alcohols (e.g., maltitol, erythritol, sorbitol), artificial sweeteners (e.g., sucralose, acesulfame potassium, stevia) and combinations thereof. A particularly desirable carbohydrate is a low dextrose equivalent (DE) maltodextrin. Long Chain Polyunsaturated Fatty Acids (LCPUFAs)

[0112] The nutritional compositions according to the general inventive concepts may also include Long Chain Polyunsaturated Fatty Acids (LCPUFAs). LCPUFAs are included in the nutritional compositions to provide nutritional support and other benefits to the end user.

[0113] Exemplary LCPUFAs for use in the nutritional compositions include, for example, ω-3 LCPUFAs and ω-6 LCPUFAs. Specific LCPUFAs include docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), arachidonic acid (ARA), linoleic acid, linolenic acid (alpha linolenic acid) and gamma-linolenic acid derived from oil sources such as plant oils, marine plankton, fungal oils, krill oil and fish oils. In one particular aspect, the LCPUFAs are derived from fish oils such as menhaden, salmon, anchovy, cod, halibut, tuna, or herring oil. Particularly preferred LCPUFAs for use in the nutritional compositions include DHA, ARA, EPA, DPA, and combinations thereof.Attorney Docket No. 15402WOO1 (67909WO01)

[0114] In order to reduce potential side effects of high dosages of LCPUFAs including DHA, ARA, EPA, DPA, in the nutritional compositions, the content of DHA, ARA, EPA, DPA, preferably does not exceed 5% by weight of the total fat content, including below 2% by weight of the total fat content, and including below 1% by weight of the total fat content in the nutritional composition.

[0115] The LCPUFA may be provided as free fatty acids, in triglyceride form, in diglyceride form, in monoglyceride form, in phospholipid form, in esterified form or as a mixture of one or more of the above, preferably in triglyceride form.

[0116] The nutritional compositions as described herein may comprise total concentrations of ARA, DHA, EPA, and DPA of from about 0.001 g / L to about 1 g / L or more, including from about 0.01 g / L to about 1 g / L, and about 0.1 g / L to about 1 g / L.

[0117] In an exemplary aspect, the nutritional compositions may include a long chain polyunsaturated fatty acid component comprising DHA and ARA in a concentration of from about 0.17 mg / mL to about 0.33 mg / mL, including from about 0.17 mg / mL to about 0.26 mg / mL of ARA and DHA. In an exemplary aspect, the nutritional compositions may include DHA in a concentration of from about 0.025 mg / mL to about 0.130 mg / mL. In another aspect, the nutritional compositions may include ARA in a concentration of from about 0.080 mg / mL to about 0.350 mg / mL. In yet another aspect, the nutritional compositions may include combinations of DHA and ARA such that the ratio of DHA to ARA ranges from about 1:4 to about 1:2. Antioxidants

[0118] Additionally, the nutritional compositions may comprise one or more antioxidants.

[0119] Any antioxidants suitable for oral administration may be included for use in the nutritional compositions according to the general inventive concepts, including, for example, vitamin A, vitamin E, vitamin C, retinol, tocopherol, and carotenoids, including lutein, beta- carotene, zeaxanthin, and lycopene, and combinations thereof, for example.

[0120] The nutritional composition may comprise at least one carotenoid selected from lutein, lycopene, zeaxanthin, and beta-carotene to provide a total amount of carotenoid of from aboutAttorney Docket No. 15402WOO1 (67909WO01) 0.001 μg / mL to about 10 μg / mL. More particularly, the nutritional compositions may comprise lutein in an amount of from about 0.001 μg / mL to about 10 μg / mL, including from about 0.001 μg / mL to about 5 μg / mL, including from about 0.001 μg / mL to about 0.0190 μg / mL, including from about 0.001 μg / mL to about 0.0140 μg / mL, and also including from about 0.044 μg / mL to about 5 μg / mL of lutein. The nutritional composition may comprise from about 0.001 μg / mL to about 10 μg / mL, including from about 0.001 μg / mL to about 5 μg / mL, from about 0.001 μg / mL to about 0.0130 μg / mL, including from about 0.001 μg / mL to about 0.0075 μg / mL, and also including from about 0.0185 μg / mL to about 5 μg / mL of lycopene. The nutritional compositions may comprise from about 1 μg / mL to about 10 μg / mL, including from about 1 μg / mL to about 5 μg / mL, including from about 0.001 μg / mL to about 0.025 μg / mL, including from about 0.001 μg / mL to about 0.011 μg / mL, and also including from about 0.034 μg / mL to about 5 μg / mL of beta-carotene. It should be understood that any combination of these amounts of beta-carotene, lutein, zeaxanthin, and lycopene can be included in the nutritional compositions according to the general inventive concepts. Other carotenoids may optionally be included in the nutritional compositions as described herein. Any one or all of the carotenoids included in the nutritional compositions described herein may be from a natural source or artificially synthesized.

[0121] Each of the carotenoids in the selected combinations can be obtained from any known or otherwise suitable material source for use in nutritional compositions, and each can be provided individually, or all together, or in any combination and from any number of sources, including sources such as multivitamin premixes containing other vitamins or minerals in combination with one or more of the carotenoids as described herein. Non-limiting examples of some suitable sources of lutein, lycopene, beta-carotene, or combinations thereof include LycoVit® lycopene (available from BASF, Mount Olive, N.J.), Lyc-O-Mato® tomato extract in oil, powder, or bead form (available from LycoRed Corp., Orange, N.J.), beta-carotene, lutein, or lycopene (available from DSM Nutritional Products, Parsippany, N.J.), FloraGLO® lutein (available from Kemin Health, Des Moines, Iowa), Xangold® Natural Lutein Esters (available from Cognis, Cincinnati, Ohio), and Lucarotin® beta-carotene (available from BASF, Mount Olive, N.J.). Other Optional IngredientsAttorney Docket No. 15402WOO1 (67909WO01)

[0122] The nutritional compositions according to the general inventive concepts may further comprise other optional components that may modify the physical, chemical, aesthetic or processing characteristics of the products or serve as pharmaceutical or additional nutritional components when used in the targeted population. Many such optional ingredients are known or otherwise suitable for use in medical food or other nutritional products or pharmaceutical dosage forms and may also be used in the compositions herein, provided that such optional ingredients are safe for oral administration and are compatible with the essential and other ingredients in the selected product form.

[0123] Non-limiting examples of such optional ingredients include preservatives, emulsifying agents, buffers, fructooligosaccharides, fiber, galactooligosaccharides, polydextrose, and other prebiotics, probiotics, pharmaceutical actives, anti-inflammatory agents, additional nutrients as described herein, colorants, flavors, thickening agents and stabilizers, emulsifying agents, lubricants, and so forth.

[0124] The nutritional compositions may further comprise a sweetening agent, preferably including at least one sugar alcohol such as maltitol, erythritol, sorbitol, xylitol, mannitol, isolmalt, and lactitol, and also preferably including at least one artificial or high potency sweetener such as acesulfame K, aspartame, sucralose, saccharin, stevia, and tagatose. These sweetening agents, especially as a combination of a sugar alcohol and an artificial sweetener, are especially useful in formulating liquid beverage embodiments according to the general inventive concepts having a desirable favor profile. These sweetener combinations are especially effective in masking undesirable flavors sometimes associated with the addition of vegetable proteins to a liquid beverage. Optional sugar alcohol concentrations in the nutritional product may range from at least 0.01%, including from about 0.1% to about 10%, and also including from about 1% to about 6%, by weight of the nutritional product. Optional artificial sweetener concentrations may range from about 0.01%, including from about 0.05% to about 5%, also including from about 0.1% to about 1.0%, by weight of the nutritional product.

[0125] The nutritional compositions may further comprise a flowing agent or anti-caking agent to retard clumping or caking of the powder over time and to make a powder embodiment flow easily from its container. Any known flowing or anti-caking agents that are known orAttorney Docket No. 15402WOO1 (67909WO01) otherwise suitable for use in a nutritional powder or product form are suitable for use herein, non-limiting examples of which include tricalcium phosphate, silicates, and combinations thereof. The concentration of the flowing agent or anti-caking agent in the nutritional composition varies depending upon the product form, the other selected ingredients, the desired flow properties, and so forth, but most typically range from about 0.1% to about 4%, including from about 0.5% to about 2%, by weight of the nutritional composition.

[0126] The nutritional compositions may further comprise a stabilizer. Any stabilizer that is known or otherwise suitable for use in a nutritional composition is also suitable for use herein, some non-limiting examples of which include gums such as xanthan gum. The stabilizer may represent from about 0.1% to about 5.0%, including from about 0.5% to about 3%, including from about 0.7% to about 1.5%, by weight of the nutritional composition.

[0127] The nutritional compositions may further comprise any of a variety of other vitamins or related nutrients, non-limiting examples of which include vitamin A, vitamin D, vitamin E, vitamin K, thiamine, riboflavin, pyridoxine, vitamin B12, niacin, folic acid, pantothenic acid, biotin, vitamin C, choline, inositol, salts and derivatives thereof, and combinations thereof. The food products preferably include, but are not limited to, the following vitamins and minerals: calcium, phosphorus, sodium, chloride, magnesium, manganese, iron, copper, zinc, selenium, iodine, chromium, molybdenum, conditionally essential nutrients m-inositol, carnitine and taurine, and Vitamins A, C, D, E (including natural vitamin E), K and the B complex, and mixtures thereof.

[0128] The nutritional compositions may further comprise any of a variety of other additional minerals, non-limiting examples of which include calcium, phosphorus, magnesium, iron, zinc, manganese, copper, sodium, potassium, molybdenum, chromium, chloride, and combinations thereof.

[0129] The nutritional compositions also may contain fiber and fiber-like stabilizers. Suitable sources of fiber and / or stabilizers include, but are not limited to, xanthan gum, guar gum, gum arabic, gum ghatti, gum karaya, gum tracacanth, agar, furcellaran, gellan gum, locust bean gum, pectin, low and high methoxy pectin, oat and barley glucans, carrageenans, psyllium, gelatin, microcrystalline cellulose, CMC (sodium carboxymethylcellulose), methylcelluloseAttorney Docket No. 15402WOO1 (67909WO01) hydroxypropyl methyl cellulose, hydroxypropyl cellulose, DATEM (diacetyl tartaric acid esters of mono- and diglycerides), dextran, carrageenans, FOS (fructooligosaccharides), and mixtures thereof. Numerous commercial sources of soluble dietary fibers are available. For example, gum arabic, hydrolyzed carboxymethylcellulose, guar gum, pectin and the low and high methoxy pectins are available from TIC Gums, Inc. of Belcamp, Md. The oat and barley glucans are available from Mountain Lake Specialty Ingredients, Inc. of Omaha, Nebr. Psyllium is available from the Meer Corporation of North Bergen, N.J. while the carrageenan is available from FMC Corporation of Philadelphia, Pa.

[0130] In addition to fiber, the nutritional compositions may also contain oligosaccharides that are not naturally found in human milk such as fructooligosaccharides (FOS) or galactooligosaccharides (GOS). These oligosaccharides are rapidly and extensively fermented to short chain fatty acids by anaerobic microorganisms that inhabit the large bowel. These oligosaccharides are preferential energy sources for most Bifidobacterium species, but are not utilized by potentially pathogenic organisms such as Clostridium perfingens, C. difficile, or Eschericia coli.

[0131] The nutritional products may additionally comprise one or more thickeners (i.e., thickening agents). The addition of thickeners reduces the incidences of paresthesia by inducing the feeling of satiety, which prolongs gastric transit time as discussed above. Product Forms

[0132] The compositions according to the general inventive concepts may be formulated and administered in any known or otherwise suitable oral product form. Any solid, liquid, semi-solid, and semi-liquid, or powder product form, including combinations or variations thereof, are suitable for use herein, provided that such forms allow for safe and effective oral delivery to the individual of the essential ingredients as also defined herein.

[0133] The compositions may be in any product form comprising the ingredients described herein, and which is safe and effective for oral administration. The compositions may be formulated to include only the ingredients described herein, or may be modified with optional ingredients to form a number of different product forms.Attorney Docket No. 15402WOO1 (67909WO01)

[0134] The compositions according to the general inventive concepts are desirably formulated as dietary product forms, which are defined herein as those embodiments comprising the ingredients according to the general inventive concepts in a product form that then contains at least one of fat, protein, and carbohydrate, and preferably also contains vitamins, minerals, or combinations thereof. The nutritional compositions will comprise at least chondroitin, e.g. in the form of chondroitin sulfate; at least chondroitin, e.g. in the form of chondroitin sulfate, and one or more HMOs; or at least chondroitin, e.g. in the form of chondroitin sulfate, one or more HMOs, and one or more nucleotides; desirably in combination with at least one of protein, fat, carbohydrate, vitamins, and minerals, to produce a nutritional composition.

[0135] Liquid compositions include both concentrated and ready-to-feed liquids. These nutritional liquids are most typically formulated as suspensions or emulsions, although other liquid forms are within the scope of the general inventive concepts. Nutritional compositions in the form of emulsions suitable for use may be aqueous emulsions comprising proteins, fats, and carbohydrates. These emulsions are generally flowable or drinkable liquids at from about 1° C. to about 25° C. and are typically in the form of oil-in-water, water-in-oil, or complex aqueous emulsions, although such emulsions are most typically in the form of oil-in-water emulsions having a continuous aqueous phase and a discontinuous oil phase.

[0136] The nutritional emulsions may be and typically are shelf stable. The nutritional emulsions typically contain up to about 95% by weight of water, including from about 50% to about 95%, also including from about 60% to about 90%, and also including from about 70% to about 85%, of water by weight of the nutritional emulsions. The nutritional emulsions may have a variety of product densities, but most typically may have a density greater than about 1.03 g / mL, including greater than about 1.04 g / mL, including greater than about 1.055 g / mL, including from about 1.06 g / mL to about 1.12 g / mL, and also including from about 1.085 g / mL to about 1.10 g / mL.

[0137] The nutritional emulsions may have a caloric density tailored to the nutritional needs of the ultimate user, although in most instances the emulsions may comprise generally at least 19 kcal / fl oz (660 kcal / liter), more typically from about 20 kcal / fl oz (675-680 kcal / liter) to about 25 kcal / fl oz (820 kcal / liter), even more typically from about 20 kcal / fl oz (675-680 kcal / liter) toAttorney Docket No. 15402WOO1 (67909WO01) about 24 kcal / fl oz (800-810 kcal / liter). Generally, the 22-24 kcal / fl oz formulas are more commonly used in preterm or low birth weight infants, and the 20-21 kcal / fl oz (675-680 to 700 kcal / liter) formulas are more often used in term infants. In some embodiments, the emulsion may have a caloric density of from about 50-100 kcal / liter to about 660 kcal / liter, including from about 150 kcal / liter to about 500 kcal / liter. In an exemplary embodiment, the emulsion may have a caloric density of 25, or 50, or 75, or 100 kcal / liter.

[0138] The nutritional emulsion may have a pH ranging from about 2.5 to about 8, but are most advantageously in a range of from about 4.5 to about 7.5, including from about 5.5 to about 7.3, including from about 6.2 to about 7.2.

[0139] Although the serving size for the nutritional emulsion can vary depending upon a number of variables, a typical serving size is generally at least about 1 mL, or even at least about 2 mL, or even at least about 5 mL, or even at least about 10 mL, or even at least about 25 mL, including ranges from about 1 mL to about 475 mL, including from about 30 mL to about 360 mL, including from about 60 mL to about 300 mL, and including from about 60 mL to about 240 mL.

[0140] The compositions in the form of solids may be in any solid form but are typically in the form of flowable or substantially flowable particulate compositions, or at least particulate compositions. Particularly suitable nutritional solid product forms include spray dried, agglomerated and / or dry blended powder compositions. The compositions can easily be scooped and measured with a spoon or similar other device, and can easily be reconstituted by the intended user with a suitable aqueous liquid, typically water, to form a nutritional composition for immediate oral or enteral use. In this context, “immediate” use generally means within about 48 hours, most typically within about 24 hours, preferably right after reconstitution.

[0141] The nutritional powders may be reconstituted with water prior to use to a caloric density tailored to the nutritional needs of the ultimate user, although in most instances the powders are reconstituted with water to form compositions comprising at least 19 kcal / fl oz (660 kcal / liter), more typically from about 20 kcal / fl oz (675-680 kcal / liter) to about 25 kcal / fl oz (820 kcal / liter), even more typically from about 20 kcal / fl oz (675-680 kcal / liter) to about 24 kcal / fl oz (800-810 kcal / liter). Generally, the 22-24 kcal / fl oz formulas are more commonly used inAttorney Docket No. 15402WOO1 (67909WO01) preterm or low birth weight infants, and the 20-21 kcal / fl oz (675-680 to 700 kcal / liter) formulas are more often used in term infants. In some embodiments, the reconstituted powder may have a caloric density of from about 50-100 kcal / liter to about 660 kcal / liter, including from about 150 kcal / liter to about 500 kcal / liter. In an exemplary embodiment, the emulsion may have a caloric density of 25, or 50, or 75, or 100 kcal / liter.

[0142] Compositions, e.g. infant formulas, in the form of a reconstitutable powder may in some embodiments be reconstituted by mixing a measured amount of powder with a measured amount of water to obtain a liquid formula of a desired caloric density. In some embodiments, for instance, a liquid formula having a caloric density of about 20 kcal / fl oz may be obtained by mixing about 8.7 g powder with about 2 fl oz water; a liquid formulation having a caloric density of about 22 kcal / fl oz may be obtained by mixing about 17.4 g powder with about 3.5 fl oz water; a liquid formulation having a caloric density of about 24 kcal / fl oz may be obtained by mixing about 26.2 g powder with about 5 fl oz water; a liquid formulation having a caloric density of about 26 kcal / fl oz may be obtained by mixing about 8.7 g powder with about 1.5 fl oz water; a liquid formulation having a caloric density of about 27 kcal / fl oz may be obtained by mixing about 26.2 g powder with about 4.25 fl oz water; or the like. Methods of Manufacture

[0143] The nutritional compositions according to the general inventive concepts may be prepared by any known or otherwise effective manufacturing technique for preparing the nutritional compositions. Many such techniques are known for any given product form such as nutritional liquids or powders and can easily be applied by one of ordinary skill in the art to the nutritional compositions described herein.

[0144] The nutritional compositions according to the general inventive concepts can therefore be prepared by any of a variety of known or otherwise effective formulation or manufacturing methods. In one suitable manufacturing process, for example, at least three separate slurries are prepared, including a protein-in-fat (PIF) slurry, a carbohydrate-mineral (CHO-MIN) slurry, and a protein-in-water (PIW) slurry. The PIF slurry is formed by heating and mixing the oil (e.g., canola oil, corn oil, etc.) and then adding an emulsifier (e.g., lecithin), fat soluble vitamins, and a portion of the total protein (e.g., milk protein concentrate, etc.) with continued heat and agitation.Attorney Docket No. 15402WOO1 (67909WO01) The CHO-MIN slurry is formed by adding with heated agitation to water: minerals (e.g., potassium citrate, dipotassium phosphate, sodium citrate, etc.), trace and ultra-trace minerals (TM / UTM premix), thickening or suspending agents (e.g., avicel, gellan, and carrageenan). The chondroitin, e.g. in the form of chondroitin sulfate, may be added to the CHO-MIN slurry at this stage or with additional minerals and / or carbohydrates as described in the following step. The resulting CHO-MIN slurry is held for 10 minutes with continued heat and agitation before adding additional minerals (e.g., potassium chloride, magnesium carbonate, potassium iodide, etc.), and / or carbohydrates (e.g., fructooligosaccharide, sucrose, corn syrup, etc.). The PIW slurry is then formed by mixing with heat and agitation the remaining protein, if any.

[0145] The resulting slurries are then blended together with heated agitation and the pH adjusted to 6.6-7.0, after which the composition is subjected to high-temperature short-time (HTST) processing during which the composition is heat treated, emulsified and homogenized, and then allowed to cool. Water soluble vitamins and ascorbic acid are added, the pH is adjusted to the desired range if necessary, flavors are added, and water is added to achieve the desired total solid level. This emulsion can then be further diluted, heat-treated, and packaged to form a ready-to-feed or concentrated liquid, or it can be heat-treated and subsequently processed and packaged as a reconstitutable powder, e.g., spray dried, dry mixed, agglomerated.

[0146] The nutritional solid, such as a spray dried nutritional powder or dry mixed nutritional powder, may be prepared by any collection of known or otherwise effective techniques, suitable for making and formulating a nutritional powder.

[0147] For example, when the nutritional powder is a spray dried nutritional powder, the spray drying step may likewise include any spray drying technique that is known for or otherwise suitable for use in the production of nutritional powders. Many different spray drying methods and techniques are known for use in the nutrition field, all of which are suitable for use in the manufacture of the spray dried nutritional powders herein.

[0148] One method of preparing the spray dried nutritional powder comprises forming and homogenizing an aqueous slurry or liquid comprising predigested fat, and optionally protein, carbohydrate, and other sources of fat as described above, and then spray drying the slurry or liquid to produce a spray dried nutritional powder. The method may further comprise the step ofAttorney Docket No. 15402WOO1 (67909WO01) spray drying, dry mixing, or otherwise adding additional nutritional ingredients, including any one or more of the ingredients described herein, to the spray dried nutritional powder.

[0149] Other suitable methods for making nutritional compositions are described, for example, in U.S. Pat. No. 6,365,218 (Borschel, et al.), U.S. Patent No. 6,589,576 (Borschel, et al.), U.S. Pat. No. 6,306,908 (Carlson, et al.), U.S. Patent Application No. 20030118703 A1 (Nguyen, et al.), which descriptions are incorporated herein by reference to the extent that they are consistent herewith. Methods of Use

[0150] The methods of use according to the general inventive concepts include the oral administration of the compositions that include at least chondroitin, e.g. in the form of chondroitin sulfate; at least chondroitin, e.g. in the form of chondroitin sulfate, and one or more HMOs; or at least chondroitin, e.g. in the form of chondroitin sulfate, one or more HMOs, and one or more nucleotides to improve gut health, particularly in an infant and more particularly in a formula-fed infant. The improvement in gut health may include promoting healthy gut function, e.g. by strengthening the intestinal barrier. The improvement in gut health may include promoting the growth / colonization of beneficial bacteria, such as bifidobacteria, in the gut.

[0151] By promoting a healthy gut microflora and / or strengthening the function of the intestinal barrier, the methods of use according to the general inventive concepts also include the oral administration of the compositions that include at least chondroitin, e.g. in the form of chondroitin sulfate; at least chondroitin, e.g. in the form of chondroitin sulfate, and one or more HMOs; or at least chondroitin, e.g. in the form of chondroitin sulfate, one or more HMOs, and one or more nucleotides to aid in digestion, to reduce the chances of developing allergies and / or food intolerances, to improve immunity and / or promote healthy development of the immune system, to treat guy dysbiosis, to treat or prevent formula intolerance or other gastrointestinal diseases or disorders.

[0152] The compositions as described herein can be administered to individuals including infants, or may, in certain embodiments, be administered to a specific subclass of individuals that are “in need thereof;” that is, to specific individuals that would particularly benefit byAttorney Docket No. 15402WOO1 (67909WO01) administration of the compositions. For example, a specific individual may be “in need of” the composition as described herein if they are susceptible to (i.e., have one or more of a genetic predisposition, a family history of, and symptoms of the disease or condition) diseases and conditions that can impair / reduce function in one or more of the areas discussed herein.

[0153] The individual desirably consumes at least one serving of the composition daily, and in some embodiments, may consume two, three, or even more servings per day. Each serving is desirably administered as a single, undivided dose, although the serving may also be divided into two or more partial or divided servings to be taken at two or more times during the day. The methods according to the general inventive concepts include continuous day after day administration, as well as periodic or limited administration, although continuous day after day administration is generally desirable. The methods according to the general inventive concepts are preferably applied on a daily basis, wherein the daily administration is maintained continuously for at least 3 days, including at least 5 days, including at least 1 month, including at least 6 weeks, including at least 8 weeks, including at least 2 months, including at least 6 months, desirably for at least about 18-24 months, desirably as a long term, continuous, daily, dietary source or supplement.

[0154] Based on the foregoing, because some of the method embodiments according to the general inventive concepts are directed to specific subsets or subclasses of identified individuals (that is, the subset or subclass of individuals “in need” of assistance in addressing one or more specific diseases or specific conditions noted herein), not all individuals will fall within the subset or subclass of individuals as described herein for certain diseases or conditions. For example, in certain instances not all infants will specifically fall within the class. Rather, it is individuals which would be considered to have an increased susceptibility over the general class of individuals sue to one or more of the risk factors described above. Examples

[0155] The following examples illustrate exemplary embodiments and / or features of the methods and nutritional compositions according to the general inventive concepts. The examples are given solely for the purpose of illustration and are not to be construed as limitations of the general inventive concepts, as many variations thereof are possible without departing from theAttorney Docket No. 15402WOO1 (67909WO01) spirit and scope of the general inventive concepts. All exemplified amounts are weight percentages based upon the total weight of the composition, unless otherwise specified. Though each of the examples involves in vitro tests, because those in vitro tests involve contacting intestinal cells with chondroitin sulfate (and in some cases additional components), the concentrations of chondroitin sulfate (and in some cases additional components) utilized in the in vitro tests are believed to correlate with the concentrations of chondroitin sulfate (and in some examples additional components) in a nutritional composition. This is because the concentration of chondroitin sulfate present in an ingested nutritional composition is expected to be generally equivalent to that which comes into contact with the intestinal cells upon digestion of the nutritional composition.

[0156] Example 1: HT-29 colonic epithelial cells employed in this example were purchased from the American Type Culture Collection (ATCC). The cells were routinely maintained in 75 cm2tissue culture flasks and incubated at 37 °C in a 5% CO2 humidified atmosphere. They were passaged when the confluency of the flasks was approximately 80–90%. After being trypsinised, they were seeded onto 12 well PVDF plates (Corning®, NY, USA) at a density of 1 × 105cells / well. The media was changed every other day with McCoy's or DMEM media (10%, v / v, FBS (Merck)) until 100% confluency was reached.

[0157] Chondroitin sulfate at different concentrations was incubated on the intestinal cells for 24hrs. A control was incubated for the same period, but without any chondroitin sulfate. After that time period, the chondroitin sulfate was removed and B. infantis 15697 was added to the intestinal cell line. After 2 hours, the cells were lysed and plated onto agar plates. After 48 hours, colony forming units (CFUs) were counted on the agar plates. The counted number of CFUs correspond to the number of bifidobacterium that attached to the intestinal cell line.

[0158] The results are shown in Figure 1. Differences were observed with the intestinal cells that had no chondroitin sulfate incubation (i.e., the control) vs. the intestinal cells which were incubated with chondroitin sulfate. Namely, as can be seen in Figure 1, incubation with chondroitin sulfate brought about a significant increase in the amount of bifidobacterium that adhered to the intestinal cells, with the number of CFUs increasing with increased amounts of chondroitin sulfate. Indeed, incubation with 300 µg / mL of chondroitin sulfate resulted in theAttorney Docket No. 15402WOO1 (67909WO01) number of colony forming units of the bifidobacterium that adhered to the intestinal cell line increasing by over double compared to that of the control. By increasing the amount of bifidobacterium that attaches to the intestinal cells, it is understood and expected that colonization of bifidobacterium in the intestine would be increased, i.e. promoted.

[0159] These results are highly surprising, particularly since previous studies with chondroitin sulfate demonstrated that it reduced the adhesion of pathogenic bacteria to intestinal cells. It has now surprisingly been found that chondroitin sulfate can enhance the colonization of beneficial bacteria such as bifidobacteria while deterring colonization of pathogenic bacteria.

[0160] Example 2: Primary human jejunum epithelial cells were obtained by Altis Biosystems. The cells were maintained in in Altis Expansion media (EM) in a cell culture incubator at 5% CO2, 37 degree Celsius.

[0161] The cells were treated with Tumor Necrosis Factor alpha (TNF alpha or TNF-a), which is an inflammatory cytokine produced by macrophages / monocytes during acute inflammation and is responsible for a diverse range of signaling events within cells. Unlike Lipopolysaccharides (LPS), which is a bacterial inflammatory stimulant that is often used for testing of this sort, TNF-a is a more generic inflammatory stimulant. As such, the use of TNF-a is believed to represent a broader range of intestinal barrier infection.

[0162] Samples of the cells were then contacted with one of the following (i) chondroitin sulfate at a concentration of 5 mg / mL, (ii) a blend of HMOs (containing 2’-Fucosyllactose, Lacto-N-Tetraose, 3-Fucosyllactose, 6’-Sialyllactose, and 3’-Sialyllactose) at a concentration of 5 mg / mL, (iii) a blend of nucleotides (containing adenosine, cytidine, guanosine, and uridine at a ratio of 0.375:1:0.5:0.425) at a concentration of 1 mg / mL, or (iv) a combination of the chondroitin sulfate at a concentration of 5 mg / mL from sample i, the blend of HMOs at a concentration of 5 mg / mL from sample ii, and the blend of nucleotides at a concentration of 1 mg / mL from sample iii. A sample treated solely with the TNF-a was maintained as a control.

[0163] More particularly, primary human jejunum epithelial cells were plated in Expansion media (EM) using Altis-established protocols onto three 96-well RepliGut® transwell plates coated with a thin layer of hydrogel. Cultures were monitored daily using a bright fieldAttorney Docket No. 15402WOO1 (67909WO01) microscope during these experiments. Media was aspirated and fresh EM was applied 48 hours after plating. Cells became confluent on day 5 post-plating, at which point EM was aspirated and Altis differentiation media (DM) was applied to promote cellular differentiation and polarization. After 48 hours, DM was removed, and fresh DM was applied to the cells. On Day 4 in DM, media was removed, and fresh DM was applied to the basal compartments and fresh DM ± (i) chondroitin sulfate at a concentration of 5 mg / mL, (ii) a blend of HMOs (containing 2’- Fucosyllactose, Lacto-N-Tetraose, 3-Fucosyllactose, 6’-Sialyllactose, and 3’-Sialyllactose) at a concentration of 5 mg / mL, (iii) a blend of nucleotides (containing adenosine, cytidine, guanosine, and uridine at a ratio of 0.375:1:0.5:0.425) at a concentration of 1 mg / mL, or (iv) a combination of the chondroitin sulfate at a concentration of 5 mg / mL from sample i, the blend of HMOs at a concentration of 5 mg / mL from sample ii, and the blend of nucleotides at a concentration of 1 mg / mL from sample iii. was applied to the apical compartment (T = -24 hours before TNFα treatment). At T = 0 hours to TNFα treatment, media was aspirated and replenished ± compounds in the apical compartment and ± TNFα in the basal compartment. Plate 1 received no TNFα and plate 2 received 3 ng / mL TNFα. At T = 24 hours of TNFα treatment, apical and basal media were collected and stored at -80ºC.

[0164] TEER was measured in all transwell cultures prior to addition of compounds to determine baseline TEER using an Epithelial Volt / Ohm Meter (World Precision Instruments, EVOM2) and STX100C96 Electrode. TEER was also measured every 24 hours after the addition of compounds for the remainder of the experiment. The surface area of a 96-well transwell is 0.143 cm2 per well. Corrected TEER was calculated using the formula: (^^^ ^^^^^^^^ ^^^^ ^^ ^^^^^^ ^^^^^^^^^−^^^ ^^^^^^^^ ^^^^ ^^ ^^^^^ ^^^^^^^^^) ^ ^^^^^^^ ^^^^.

[0165] TEER is the measurement of electrical resistance across a cellular monolayer and is a very sensitive and reliable method to confirm the integrity and permeability of the monolayer. A higher TEER result means that the cells are more intact, i.e. form a stronger barrier.

[0166] The results are shown in Figure 2. As can be seen in Figure 2, treatment of the cells with chondroitin sulfate alone (sample i) resulted in little to no increase in TEER over the control. Treatment with the blend of HMOs (sample ii) resulted in little to no effect and actually resulted in a reduction in the TEER. Treatment with the blend of nucleotides (sample iii) resulted in a anAttorney Docket No. 15402WOO1 (67909WO01) increase in the TEER of about 130%, bringing it up to about 200 ohms / cm2. Treatment with the combination of chondroitin sulfate, HMOs, and nucleotides (sample iv), however, resulted in an increase in the TEER of about 600% over the control, bringing it up to over 600 ohms / cm2. Together, the chondroitin sulfate, HMOs, and nucleotides produced a synergistic effect on the epithelial cells, producing an extremely strong intestinal barrier and demonstrating a synergistic and unexpected ability to maintain, and even increase, the function of the intestinal barrier in the face of inflammatory stimulation.

[0167] Figure 3 shows the results of additional testing in which it can be seen that after treatment with the TNF-a, incubation of the epithelial cells with a combination of the chondroitin sulfate at a concentration of 5 mg / mL from sample i. and the blend of nucleotides at a concentration of 1 mg / mL from sample iii. resulted in a synergistic increase in TEER, bringing it up to about 300 ohms / cm2. However, this result also demonstrates that the extremely high increase in TEER produced by sample iv. is at least partly attributed to a synergistic effect between chondroitin sulfate and HMOs. Figure 3 also shows the TEER of the epithelial cells prior to and without any treatment with TNF-a, which is notably lower than that of the cells after treatment with TNF-a and incubation with a combination of chondroitin sulfate, HMOs, and nucleotides.

[0168] Example 3: Caco-2 intestinal epithelial cells employed in this examples were purchased from the American Type Culture Collection (ATCC). The Caco-2 cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% (v / v) FBS. All cells were routinely maintained in 75 cm2tissue culture flasks and incubated at 37 °C in a 5% CO2humidified atmosphere. They were passaged when the confluency of the flasks was approximately 80–90%. After being trypsinised, they were seeded onto 12 well PVDF plates (Corning®, NY, USA) at a density of 1 × 105cells / well. The media was changed every other day with McCoy's or DMEM media (10%, v / v, FBS (Merck)) until 100% confluency was reached.

[0169] Bacterial strains used in this example were stored in their respective broth containing 50% (v / v) glycerol at −20 °C and cultured directly from storage into the same broth (0.2% inoculum) at 37 °C for 24 h.

[0170] Chondroitin sulfate at a concentration of 0.1 g / L was incubated with E. coli 35401 (Enterotoxigenic Escherichia coli; ETEC ATCC 35401, serotype O78:H11) at a concentration ofAttorney Docket No. 15402WOO1 (67909WO01) 1 x 108CFU / mL, under aerobic conditions for 1 hr at 37^C. After the incubation period, the Chondroitin sulfate:bacterial mix was exposed to Caco-2 intestinal epithelial cells for 2 hrs at 37^C in a 5% CO2 incubator. To determine pathogen adhesion, cells were washed with 1 mL PBS 3-4 times and thereafter lysed with 0.2% Triton-X-100 (PBS) for 30 min. Serial dilutions of the cell lysates were plated onto agar plates and incubated at 37 °C for 24 h after which bacterial CFU were counted. For comparison, a control was conducted in which the cells were exposed to pathogen which had not been subjected to incubation with chondroitin sulfate. The experiment was repeated using a blend of HMOs (containing 2’-Fucosyllactose, Lacto-N-Tetraose, 3- Fucosyllactose, 6’-Sialyllactose, and 3’-Sialyllactose) at a concentration of 5 mg / mL and a combination of the chondroitin sulfate and HMOs.

[0171] The results are shown in Figure 4. As shown in Figure 4, pre-incubation of the pathogen with 5HMO at a concentration of 5 g / L or chondroitin sulfate at a concentration of 0.1 g / L reduced adhesion of E. coli 35401 (Enterotoxigenic Escherichia coli; ETEC ATCC 35401, serotype O78:H11) to Caco-2 intestinal epithelial cells individually (30% 5HMO; 29% CS). However, a combination of 5HMO at a concentration of 5 g / L and chondroitin sulfate at a concentration of 0.1 g / L significantly reduced E. coli 35401 adhesion to Caco-2 intestinal epithelial cells greater than either ingredient alone (5HMO:CS mixture 57%).

[0172] Unless otherwise indicated herein, all sub-embodiments and optional embodiments are respective sub-embodiments and optional embodiments to all embodiments described herein. While the present application has been illustrated by the description of embodiments thereof, and while the embodiments have been described in considerable detail, it is not the intention of the applicants to restrict or in any way limit the scope of the appended claims to such detail. Additional advantages and modifications will readily appear to those skilled in the art. Therefore, the application, in its broader aspects, is not limited to the specific details, the representative compositions or formulations, and illustrative examples shown and described. Accordingly, departures may be made from such details without departing from the spirit or scope of the applicant’s general disclosure herein.

Claims

Attorney Docket No. 15402WOO1 (67909WO01) WHAT IS CLAIMED IS:

1. An infant formula comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

2. The infant formula of claim 1, wherein the chondroitin or chondroitin sulfate is present in an amount of at least 0.1 g / L, optionally at least 0.2 g / L, optionally at least 0.3 g / L, when the infant formula is a liquid.

3. The infant formula of claim 2, wherein the chondroitin or chondroitin sulfate is present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L.

4. The infant formula of any preceding claim, wherein the infant formula is a reconstitutable powder and the chondroitin or chondroitin sulfate is present in an amount between about 0.1 wt. % and about 1.0 wt. % of the powder.

5. The infant formula of any preceding claim, further comprising one or more human milk oligosaccharides (HMOs).

6. The infant formula of claim 5, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof.

7. The infant formula of claim 6, wherein the one or more human milk oligosaccharides comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N-acetylglucosamine oligosaccharide.

8. The infant formula of any one of claims 6 and 7, wherein the fucosylated oligosaccharide comprises at least one of 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL); theAttorney Docket No. 15402WOO1 (67909WO01) sialylated oligosaccharide comprises at least one of 3’-sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide comprises at least one of lacto-N-neotetraose (LNnT), lacto-N-tetraose (LNT).

9. The infant formula of claim 5, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of 2′-fucosyllactose, 3-fucosyllactose, 3′- sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and any combination thereof.

10. The infant formula of claim 9, wherein the one or more human milk oligosaccharides comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N- tetraose.

11. The infant formula of any one of claims 5 to 10, wherein the infant formula contains total HMOs in an amount between 0.1 g / L and 10 g / L when the infant formula is a liquid.

12. The infant formula of any one of claims 5 to 11, wherein the infant formula is a reconstitutable powder and contains total HMOs in an amount between about 1 wt. % and about 10 wt. % of the powder.

13. The infant formula of any preceding claim, further comprising one or more nucleotides.

14. The infant formula of claim 13, wherein the one or more nucleotides comprises at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof.

15. The infant formula of any one of claims 13 and 14, wherein the infant formula contains total nucleotides in an amount between about 15 mg / L and about 110 mg / L when the infant formula is a liquid.

16. The infant formula of any one of claims 13 to 15, wherein the infant formula is a reconstitutable powder and contains total nucleotides in an amount between about 0.01 wt. % and about 0.10 wt. % of the powder.

17. The infant formula of any preceding claim, wherein the infant formula is a ready-to-drink liquid.Attorney Docket No. 15402WOO1 (67909WO01) 18. The infant formula of any one of claims 1 to 16, wherein the infant formula is a reconstitutable powder that is configured to be reconstituted into a liquid.

19. The infant formula of any preceding claim, wherein the infant formula comprises protein, carbohydrate, and fat.

20. The infant formula of claim 19, wherein the infant formula comprises carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).

21. The infant formula of any preceding claim, wherein the formula is configured to promote / increase the growth of beneficial intestinal bacteria, optionally wherein the beneficial intestinal bacteria comprises bifidobacteria, optionally wherein the formula is configured to promote the development of a beneficial intestinal microbiota comparable with that found in breast fed babies.

22. The infant formula of any preceding claim, wherein the formula is configured to strengthen the intestinal barrier.

23. The infant formula of any preceding claim, wherein the formula is configured to improve the feeding tolerance of an infant and / or to treat feeding intolerance in an infant.

24. A method of promoting the growth of beneficial intestinal bacteria in an infant, e.g. a formula-fed infant, optionally wherein the beneficial intestinal bacteria comprises bifidobacteria, the method comprising administering to an infant a nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

25. A method of promoting the development of a beneficial intestinal microbiota comparable with that found in a breast-fed infant, e.g. a formula-fed infant, the method comprising administering to an infant a nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.Attorney Docket No. 15402WOO1 (67909WO01) 26. A method of promoting a bifidogenic intestinal microbiota in an infant, e.g. a formula-fed infant, the method comprising administering to an infant a nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

27. A method of promoting a strong intestinal barrier, the method comprising administering to an individual, optionally an infant, a nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

28. A method of improving feeding tolerance and / or treating feeding intolerance, the method comprising administering to an individual, optionally an infant, a nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

29. The method of any one of claims 24-28, wherein the nutritional composition is an infant formula.

30. The method of any one of claims 24-29, wherein the nutritional composition is a ready-to- drink liquid.

31. The method of any one of claims 24-29, wherein the nutritional composition is a liquid that is reconstituted from a reconstitutable powder.

32. The method of any one of claims 24-31, wherein the nutritional composition comprises at least 0.1 g / L chondroitin or chondroitin sulfate, optionally at least 0.2 g / L chondroitin or chondroitin sulfate, optionally at least 0.3 g / L chondroitin or chondroitin sulfate, optionally wherein the chondroitin or chondroitin sulfate is present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L.

33. The method of any one of claims 24-32, wherein the nutritional composition further comprises one or more human milk oligosaccharides (HMOs).Attorney Docket No. 15402WOO1 (67909WO01) 34. The method of claim 33, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof.

35. The method of claim 33, wherein the one or more human milk oligosaccharides comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N-acetylglucosamine oligosaccharide.

36. The method of any one of claims 34-35, wherein the fucosylated oligosaccharide comprises at least one of 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL); the sialylated oligosaccharide comprises at least one of 3’-sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N- acetylglucosamine oligosaccharide comprises at least one of lacto-N-neotetraose (LNnT), lacto- N-tetraose (LNT).

37. The method of claim 33, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′- sialyllactose, and lacto-N-tetraose, and any combination thereof.

38. The method of claim 33, wherein the one or more human milk oligosaccharides comprises 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

39. The method of any one of claims 33-38, wherein the nutritional composition contains total HMOs in an amount between 0.1 g / L and 10 g / L when the infant formula is a liquid.

40. The method of any one of claims 24-39, wherein the nutritional composition further comprises one or more nucleotides.

41. The method of claim 40, wherein the one or more nucleotides comprises at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof.

42. The method of any one of claims 40-41, wherein the nutritional composition contains total nucleotides in an amount between about 15 mg / L and about 110 mg / L when the nutritional composition is a liquid.Attorney Docket No. 15402WOO1 (67909WO01) 43. The method of any one of claims 24-42, wherein the nutritional composition comprises protein, carbohydrate, and fat.

44. The method of claim 43, wherein the nutritional composition comprises carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).

45. The method of any one of preceding claims 24-44, wherein the infant is a formula-fed infant.

46. A nutritional composition for use in promoting the growth of beneficial intestinal bacteria in an infant, e.g. a formula-fed infant, optionally wherein the beneficial intestinal bacteria comprises bifidobacteria, optionally wherein the nutritional composition is configured to promote the development of a beneficial intestinal microbiota comparable with that found in a breast-fed infant, the nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

47. A nutritional composition for use in promoting a bifidogenic intestinal microbiota in an infant, e.g. a formula-fed infant, the nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

48. A nutritional composition for use in promoting a strong intestinal barrier in an individual, optionally an infant, the nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

49. The nutritional composition of any one of claims 46-48, wherein the nutritional composition is an infant formula.

50. The nutritional composition of any one of claims 46-49, wherein the nutritional composition is a ready-to-drink liquid.

51. The nutritional composition of any one of claims 46-49, wherein the nutritional composition is a reconstitutable powder that is configured to be reconstituted into a drinkable liquid.Attorney Docket No. 15402WOO1 (67909WO01) 52. The nutritional composition of any one of claims 46-51, wherein when in liquid form the nutritional composition comprises at least 0.1 g / L chondroitin or chondroitin sulfate, optionally at least 0.2 g / L chondroitin or chondroitin sulfate, optionally at least 0.3 g / L chondroitin or chondroitin sulfate, optionally wherein the chondroitin or chondroitin sulfate is present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L.

53. The nutritional composition of claim 51 wherein the nutritional composition is a reconstitutable powder and wherein the chondroitin or chondroitin sulfate is present in an amount between about 0.1 wt. % and about 1.0 wt. % of the powder.

54. The nutritional composition of any one of claims 46-53, further comprising one or more human milk oligosaccharides (HMOs).

55. The nutritional composition of claim 54, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof.

56. The nutritional composition of claim 55, wherein the one or more human milk oligosaccharides comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N-acetylglucosamine oligosaccharide.

57. The nutritional composition of any one of claims 55 and 56, wherein the fucosylated oligosaccharide comprises at least one of 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL); the sialylated oligosaccharide comprises at least one of 3’-sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide comprises at least one of lacto-N- neotetraose (LNnT), lacto-N-tetraose (LNT).

58. The nutritional composition of claim 54, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of 2′-fucosyllactose, 3-Attorney Docket No. 15402WOO1 (67909WO01) fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and any combination thereof.

59. The nutritional composition of claim 54, wherein the one or more human milk oligosaccharides comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

60. The nutritional composition of any one of claims 54-59, wherein the nutritional composition contains total HMOs in an amount between 0.1 g / L and 10 g / L when the infant formula is a liquid.

61. The nutritional composition of any one of claims 54-59, wherein the nutritional powder is a reconstitutable powder and contains total HMOs in an amount between about 1 wt. % and about 10 wt. % of the powder.

62. The nutritional composition of any one of claims 46-61, further comprising one or more nucleotides.

63. The nutritional composition of claim 62, wherein the one or more nucleotides comprises at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof.

64. The nutritional composition of any one of claims 62-63, wherein the nutritional composition contains total nucleotides in an amount between about 15 mg / L and about 110 mg / L when the nutritional composition is a liquid.

65. The nutritional composition of any one of claims 62-63, wherein the nutritional composition is a reconstitutable powder and contains total nucleotides in an amount between about 0.01 wt. % and about 0.10 wt. % of the powder.

66. The nutritional composition of any one of claims 46-65, wherein the nutritional composition comprises protein, carbohydrate, and fat.

67. The nutritional composition of claim 66, wherein the nutritional composition comprises carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fatAttorney Docket No. 15402WOO1 (67909WO01) in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).

68. A nutritional composition comprising chondroitin, optionally wherein the chondroitin is in the form of chondroitin sulfate.

69. The nutritional composition of claim 68, wherein the chondroitin or chondroitin sulfate is present in an amount of at least 0.1 g / L, optionally at least 0.2 g / L, optionally at least 0.3 g / L, when the nutritional composition is a liquid.

70. The nutritional composition of claim 69, wherein the chondroitin or chondroitin sulfate is present in an amount between about 0.1 g / L and about 3 g / L, optionally between about 0.1 g / L and about 2 g / L, optionally between about 0.1 g / L and about 1.5 g / L, optionally between about 0.2 g / L and about 3 g / L, optionally between about 0.2 g / L and about 2 g / L, optionally between about 0.2 g / L and about 1.5 g / L, optionally between about 0.3 g / L and about 3 g / L, optionally between about 0.3 g / L and about 2 g / L, optionally between about 0.3 g / L and about 1.5 g / L.

71. The nutritional composition of any preceding claim, wherein the nutritional composition is a reconstitutable powder and the chondroitin or chondroitin sulfate is present in an amount between about 0.1 wt. % and about 1.0 wt. % of the powder.

72. The nutritional composition of any preceding claim, further comprising one or more human milk oligosaccharides (HMOs).

73. The nutritional composition of claim 72, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of a fucosylated oligosaccharide, a sialylated oligosaccharide, an N-acetylglucosamine oligosaccharide, and any combination thereof.

74. The nutritional composition of claim 73, wherein the one or more human milk oligosaccharides comprises a combination of at least one fucosylated oligosaccharide, at least one sialylated oligosaccharide, and at least one N-acetylglucosamine oligosaccharide.

75. The nutritional composition of any one of claims 73-74 wherein the fucosylated oligosaccharide comprises at least one of 3-fucosyllactose (3-FL) and 2′-fucosyllactose (2′-FL);Attorney Docket No. 15402WOO1 (67909WO01) the sialylated oligosaccharide comprises at least one of 3’-sialyllactose (3′-SL) and 6′ sialyllactose (6’-SL); and the N-acetylglucosamine oligosaccharide comprises at least one of lacto-N- neotetraose (LNnT), lacto-N-tetraose (LNT).

76. The nutritional composition of claim 72, wherein the one or more human milk oligosaccharides comprises at least one selected from the group of 2′-fucosyllactose, 3- fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose, and any combination thereof.

77. The nutritional composition of claim 76, wherein the one or more human milk oligosaccharides comprise 2′-fucosyllactose, 3-fucosyllactose, 3′-sialyllactose, 6′-sialyllactose, and lacto-N-tetraose.

78. The nutritional composition of any one of claims 72-77, wherein the nutritional composition contains total HMOs in an amount between 0.1 g / L and 10 g / L when the infant formula is a liquid.

79. The nutritional composition of any one of claims 72-78, wherein the nutritional composition is a reconstitutable powder and contains total HMOs in an amount between about 1 wt. % and about 10 wt. % of the powder.

80. The nutritional composition of any preceding claim, further comprising one or more nucleotides.

81. The nutritional composition of claim 80, wherein the one or more nucleotides comprises at least one selected from the group of adenosine, cytidine, guanosine, uridine, and any combination thereof.

82. The nutritional composition of any one of claims 80-81, wherein the nutritional composition contains total nucleotides in an amount between about 15 mg / L and about 110 mg / L when the infant formula is a liquid.

83. The nutritional composition of any one of claims 80-82, wherein the nutritional composition is a reconstitutable powder and contains total nucleotides in an amount between about 0.01 wt. % and about 0.10 wt. % of the powder.Attorney Docket No. 15402WOO1 (67909WO01) 84. The nutritional composition of any preceding claim, wherein the nutritional composition is a ready-to-drink liquid. 85 The nutritional composition of any one of claims 68-83, wherein the nutritional composition is a reconstitutable powder that is configured to be reconstituted into a liquid.

86. The nutritional composition of any preceding claim, wherein the nutritional composition comprises protein, carbohydrate, and fat.

87. The nutritional composition of claim 86, wherein the nutritional composition comprises carbohydrate in an amount of from about 40% to about 60% by weight (on a dry weight basis); fat in an amount of from about 20% to about 40% by weight (on a dry weight basis); and protein in an amount of about 10% to about 25% by weight (on a dry weight basis).

88. The nutritional composition of any preceding claim, wherein the nutritional composition is configured to promote / increase the growth of beneficial intestinal bacteria, optionally wherein the beneficial intestinal bacteria comprises bifidobacteria, optionally wherein the nutritional composition is configured to promote the development of a beneficial intestinal microbiota comparable with that found in breast fed babies.

89. The nutritional composition of any preceding claim, wherein the nutritional composition is configured to strengthen the intestinal barrier.

90. The nutritional composition of any preceding claim, wherein the nutritional composition is configured to improve the feeding tolerance of an infant and / or to treat feeding intolerance in an infant.

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