Methods for modulating gastrointestinal microbial metabolic pathways and metabolites

Abstract

The present disclosure relates to methods of feeding animals by providing feed additives that modulate the gut microbiome to improve health, nutrition, and growth performance. The present disclosure further relates to methods of modulating metabolites present in the gastrointestinal tract of an animal. Such modulation includes, for example, modulating the levels of said metabolites.

Description

Detailed Description of the Invention

[0001] [CROSS-REFERENCE TO RELATED APPLICATIONS]

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 62 / 989,107, filed March 13, 2020, the entire disclosure of which is incorporated herein by reference.

[0002] [background] The gut microbiome, including bacteria, viruses, fungi, molds, and protozoa present in the digestive tract, is responsible for converting undigested and unabsorbed components of an animal's diet into thousands of biologically active metabolites that subsequently interact with the animal's local and systemic physiology and its external environment.

[0003]

[0003] Under normal circumstances, the biochemical output of the microbiome is determined in part by the composition of the food consumed by the animal and in part by the phylogenetic composition of the gut microbiome. In conventional diets, especially those containing plant fiber polysaccharides and seeds, such as cellulose, lignin, hemicellulose, pectin, and starch-binding proteins, a portion of the food ingested by the animal remains undigested and unabsorbed by the primary digestive process. These unabsorbed species reach the lower intestinal system, where they can be processed and utilized by microbiota and converted into metabolites. The resulting gut metabolome, produced by the metabolic action of the gut microbiome on these unabsorbed components of the feed, is influenced by the chemical composition of these various unabsorbed components.

[0004]

[0004] Metabolites produced in the intestine can be absorbed, for example, via the colon or portal venous circulation, and transported to other organs of the animal, where they can affect the structure and / or function of those organs. These biochemicals affect diverse biological functions, such as nutrient absorption, energy regulation, mitochondrial function, systemic inflammation, stress response, liver function, kidney function, cardiometabolic function, satiety, physiological mood, and alertness. Metabolites produced in the intestine can also be excreted by the animal into its external environment.

[0005]

[0005] In some cases, metabolites produced by the gut microbiome are beneficial to the host or otherwise contribute to the productivity, health, welfare, and sustainability of the host animal. In other cases, metabolites produced by the gut microbiome are harmful to the host, resulting in reduced productivity, health, or welfare. Certain metabolites are undesirable because, when excreted, they can be harmful to the animal's external environment and can result in water, soil, and / or air pollution or otherwise increase the environmental footprint of animal farming.

[0006]

[0006] Overall animal productivity and health are key factors in the economics of the animal protein production industry. Consumer and regulatory pressure for improved sustainability is becoming increasingly essential to maintaining competitiveness in the production industry.

[0007]

[0007] Thus, there exists a need to be able to regulate or otherwise control the metabolic pathways and metabolic output of an animal's gut microbiome in order to improve the nutrition, health, welfare, and / or sustainability of production and companion animals. However, a challenge is that animals typically exhibit high taxonomic variability in the phylogenetic composition of their gut microbiome. This is true even in the animal production industry, where thousands of essentially identical animals are often raised in cohorts with the same underlying genetics, a common phylogenetic diet, and a common environment, and significant phylogenetic differences are observed in the microbiomes of apparently identical, healthy animals. Thus, conventional feed additives that target the gut microbiota are generally understood in the industry to not provide a consistent impact on the gut metabolome of animals fed them.

[0008] [overview] In one aspect, provided herein are methods for improving the nutrition, health, welfare, and sustainability of an animal by providing the animal with a feed additive that increases or decreases the expression of one or more metabolic pathways in the metagenome of the animal's microbiome. In certain embodiments, the method for improving animal nutrition comprises increasing the abundance, expression, or flux through a metabolic pathway in the metagenome of gastrointestinal microflora responsible for harvesting nutritional energy from undigested components of the animal's diet. In other embodiments, the method for improving animal health comprises decreasing the expression, expression, or flux through a metabolic pathway in the metagenome of gastrointestinal microflora responsible for the adverse breakdown of aromatic amino acids to biogenic amines. In other embodiments, the method for improving animal health comprises increasing the expression, abundance, or flux through a metabolic pathway in the metagenome of gastrointestinal microflora responsible for maintaining immune-inflammatory homeostasis.

[0009]

[0009] In one aspect, provided herein is a method for improving nitrogen utilization in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydrosubunit-containing oligosaccharide at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of a plurality of metabolites associated with improved nitrogen utilization are higher in a gastrointestinal sample from the animal compared to a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0010] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0011] In some embodiments, the plurality of metabolites are (2S,3S)-3-methylaspartate, (R)-3-(phenyl)lactate, (R)-3-(phenyl)lactoyl-CoA, (S)-3-aminobutanoyl-CoA, 2-oxoglutarate, 3-(4-hydroxyphenyl)pyruvate, 4-aminobutanal, 4-aminobutanoate, 4-guanidinobutanoate, 4-guanidinobutryaldehyde, 4-guanidobutryamide, 4-maleyl-acetoacetate, 5-aminopentanal, 5-aminopentanoate, 5-guanidino-2-oxopentanoate, agmatine, ammonia, and cadaverine. , cinnamate, cinnamoyl-CoA, coenzyme A, formamide, homogentisate, L-β-lysine, L-cystathionine, L-glutamate, L-glutamate-5-semialdehyde, L-histidine, L-homocysteine, L-lysine, L-methionine, L-ornithine, L-proline, L-serine, mesaconate, N-carbamoylputrescine, N-formimino-L-glutamate, N2-succinyl-L-arginine, pyruvate, succincate semialdehyde, succinyl-CoA, or urea.

[0012]

[0012] In some embodiments, the levels of the multiple metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the levels of the multiple metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basic nutritional composition but not the synthetic oligosaccharide preparation.

[0013]

[0013] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold higher than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basic nutritional composition but not the synthetic oligosaccharide preparation.

[0014]

[0014] In some embodiments, the levels of multiple metabolites associated with reduced nitrogen utilization are lower in gastrointestinal samples from the animal compared to gastrointestinal samples from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0015] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0016]

[0016] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (3S,5S)-3,5-diaminohexanoate, (S)-3-methyl-2-oxopentanoate, (S)-5-amino-3-oxohexanoate, 2-oxoglutarate, acetyl-CoA, ammonia, D-alanine, formate, fumarate, glycine, L-2-amino-3-oxobutanoate, L-alanine, L-asparagine, L-aspartate, L-glutamate, L-isoleucine, N-formimino-L-glutamate, N-formyl-L-glutamate, N2-succinylglutamate, and pyruvate.

[0017]

[0017] In some embodiments, the levels of the multiple metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% lower than the levels of the multiple metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basic nutritional composition but not the synthetic oligosaccharide preparation.

[0018]

[0018] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basic nutritional composition but not the synthetic oligosaccharide preparation.

[0019]

[0019] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0020]

[0020] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0021] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0022] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0023]

[0023] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0024] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0025]

[0025] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0026]

[0026] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0027]

[0027] In some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0028] In some embodiments, the nutritional composition comprises an α-tocopherol concentration of about 100 ppm to 2000 ppm, 100 ppm to 1500 ppm, 100 ppm to 1000 ppm, 100 ppm to 900 ppm, 100 ppm to 800 ppm, 100 ppm to 700 ppm, 100 ppm to 600 ppm, 100 ppm to 500 ppm, 100 ppm to 400 ppm, 100 ppm to The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0029]

[0029] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0030]

[0030] In some embodiments, the animal has an increased body weight compared to the animal's body weight prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0031]

[0031] In some embodiments, the body weight of the animal increases by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the body weight of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0032]

[0032] In some embodiments, the weight gain is greater than that of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0033]

[0033] In some embodiments, the body weight of the animal increases by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0034]

[0034] In some embodiments, the animal has increased feed efficiency compared to the animal's feed efficiency prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0035]

[0035] In some embodiments, the feed efficiency of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the feed efficiency of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0036]

[0036] In some embodiments, the animal has an increase in feed efficiency that is greater than an increase compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0037]

[0037] In some embodiments, the increase in feed efficiency of the animal is at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of the equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0038]

[0038] In some embodiments, the animal has a decreased feed conversion ratio (FCR) compared to the FCR of the animal prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0039]

[0039] In some embodiments, the animal's feed conversion rate is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the animal's feed conversion rate before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0040]

[0040] In some embodiments, the animal has a reduction in the feed conversion ratio that is greater than a reduction compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0041]

[0041] In some embodiments, the feed conversion ratio of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the feed conversion ratio of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0042]

[0042] In some embodiments, the life expectancy or survival rate of the animal is increased compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0043]

[0043] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to animals administered a nutritional composition that does not contain the synthetic oligosaccharide preparation.

[0044]

[0044] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to the animal prior to administration of the synthetic oligosaccharide preparation.

[0045]

[0045] In some embodiments, administration results in improved quality of meat from the animal compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0046]

[0046] In some embodiments, the administration results in at least one of a) improved color of the animal's meat, b) improved flavor of the animal's meat, and c) improved tenderness of the animal's meat.

[0047]

[0047] In some embodiments, the animal is poultry, seafood, sheep, cattle, cattle, buffalo, bison, pig, cat, dog, rabbit, goat, guinea pig, donkey, camel, horse, pigeon, ferret, gerbil, hamster, mouse, rat, fish or bird.

[0048] In some embodiments, the animal is poultry. In some embodiments, the poultry is a chicken, turkey, duck, or goose. In some embodiments, the poultry is a chicken. In some embodiments, the chicken is a broiler chicken, layer chicken, or breeder chicken.

[0049] In some embodiments, the animal is a pig. In some embodiments, the pig is a young pig, a growing pig, or a finishing pig.

[0050] In some embodiments, the animal is a fish. In some embodiments, the fish is a salmon, tilapia, or tropical fish.

[0051] In some embodiments, the animal is a livestock animal.

[0052] In some embodiments, the animal is a companion animal, hi some embodiments, the companion animal is a cat, dog, hamster, rabbit, guinea pig, ferret, gerbil, bird, or mouse.

[0053] In some embodiments, the relative abundance of oligosaccharides in DP fractions of at least 5, 10, 20, or 30 decreases monotonically with their degree of polymerization.

[0054] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with its degree of polymerization. In some embodiments, n is at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 1 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100.

[0055]

[0055] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2% or less than 1%.

[0056]

[0056] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0057]

[0057] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0058]

[0058] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0059]

[0059] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0060]

[0060] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2% or less than 1%.

[0061]

[0061] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0062]

[0062] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0063]

[0063] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0064]

[0064] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0065]

[0065] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 15%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2% or less than 1%.

[0066]

[0066] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0067]

[0067] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0068]

[0068] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0069]

[0069] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0070]

[0070] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 2% to about 12%.

[0071] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%.

[0072] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 1% to about 10%.

[0073]

[0073] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharide in a relative abundance of about 5% to about 10%.

[0074]

[0074] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0075]

[0075] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0076]

[0076] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0077]

[0077] In some embodiments, the oligosaccharide preparation comprises anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.5%, 0.6%, 0.8%, 1.0%, 1.5%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, or 12%.

[0078]

[0078] In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 1% to about 40% by weight as determined by liquid chromatography.

[0079]

[0079] In some embodiments, the oligosaccharide preparation has a DP2 fraction content of about 1% to about 35% by weight as determined by liquid chromatography.

[0080]

[0080] In some embodiments, the oligosaccharide preparation has a DP3 fraction content of about 1% to about 30% by weight as determined by liquid chromatography.

[0081]

[0081] In some embodiments, the oligosaccharide preparation has a DP4 fraction content of about 0.1% to about 20% by weight as determined by liquid chromatography.

[0082]

[0082] In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 0.1% to about 15% by weight as determined by liquid chromatography.

[0083] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction is about 0.02 to about 0.40 as determined by liquid chromatography.

[0084] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction is about 0.01 to about 0.30 as determined by liquid chromatography.

[0085] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 40%, or less than 30% as determined by liquid chromatography.

[0086]

[0086] In some embodiments, the oligosaccharide preparation comprises at least 10 3 , at least 10 4 , at least 10 5 , at least 10 6 , or at least 10 9 different oligosaccharide species.

[0087]

[0087] In some embodiments, two or more independent oligosaccharides comprise different anhydro subunits.

[0088]

[0088] In some embodiments, each of the anhydro-subunit-containing oligosaccharides comprises one or more anhydro-subunits that are the thermal dehydration product of a monosaccharide.

[0089]

[0089] In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits selected from anhydro-glucose, anhydro-galactose, anhydro-mannose, anhydro-allose, anhydro-altrose, anhydro-gulose, anhydro-indose, anhydro-talose, anhydro-fructose, anhydro-ribose, anhydro-arabinose, anhydro-rhamnose, anhydro-lyxose, and anhydro-xylose.

[0090]

[0090] In some embodiments, the oligosaccharide preparation comprises one or more anhydro-glucose, anhydro-galactose, anhydro-mannose, or anhydro-fructose subunits.

[0091]

[0091] In some embodiments, the DP1 fraction comprises 1,6-anhydro-β-D-glucofuranose anhydro subunits or 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0092]

[0092] In some embodiments, the DP1 fraction contains both 1,6-anhydro-β-D-glucofuranose anhydro subunits and 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0093] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is In the preparation), the ratio is about 10:1 to 1:10, about 9:1 to about 1:10, about 8:1 to about 1:10, about 7:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 10:1 to about 1:9, about 10:1 to about 1:8, about 10:1 to about 1:7, about 10:1 to about 1:6, about 10:1 to about 1:5, about 10:1 to about 1:4, about 10:1 to about 1:3, about 10:1 to about 1:2, or about 1:1 to about 3:1.

[0094]

[0094] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide preparation is about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9 or about 1:10.

[0095]

[0095] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in the oligosaccharide preparation.

[0096]

[0096] In some embodiments, the DP2 fraction comprises at least five anhydro-subunit-containing oligosaccharides.

[0097]

[0097] In some embodiments, the DP2 fraction comprises about 5-10 anhydro-subunit-containing oligosaccharides.

[0098] In some embodiments, the oligosaccharide preparation comprises one or more sugar caramelization products, which may be selected from the group consisting of methanol, ethanol, furan, methylglyoxal, 2-methylfuran, vinyl acetate, glycolaldehyde, acetic acid, acetol, furfural, 2-furanmethanol, 3-furanmethanol, 2-hydroxycyclopent-2-en-1-one, 5-methylfurfural, 2(5H)-furanone, 2-methylcyclopentenolone, levoglucosenone, cyclic hydroxylactones, 1,4,3,6-dianhydro-α-D-glucopyranose, dianhydroglucopyranose, and 5-hydroxymethylfurfural (5-hmf).

[0099] In some embodiments, greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0100]

[0100] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 5000 g / mol as determined by high performance liquid chromatography (HPLC).

[0101]

[0101] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0102]

[0102] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0103]

[0103] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0104]

[0104] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 5000 g / mol as determined by HPLC.

[0105]

[0105] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0106]

[0106] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0107]

[0107] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0108]

[0108] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 2000 to about 2800 g / mol.

[0109]

[0109] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 1000 to about 2000 g / mol.

[0110]

[0110] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0111]

[0111] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0112]

[0112] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0113]

[0113] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0114]

[0114] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0115] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation, hi some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0116] In some embodiments, the nutritional composition contains about 100 ppm to 2000 ppm, 100 ppm to 1500 ppm, 100 ppm to 1000 ppm, 100 ppm to 900 ppm, 100 ppm to 800 ppm, 100 ppm to 700 ppm, 100 ppm to 600 ppm, 100 ppm to 500 ppm, 100 ppm to 400 ppm, 100 ppm to The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0117]

[0117] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0118]

[0118] In one aspect, provided herein is a method of improving nitrogen utilization in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharide at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of a plurality of metabolites associated with decreased nitrogen utilization are lower in a gastrointestinal sample from the animal compared to a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0119] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0120] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (3S,5S)-3,5-diaminohexanoate, (S)-3-methyl-2-oxopentanoate, (S)-5-amino-3-oxohexanoate, 2-oxoglutarate, acetyl-CoA, ammonia, D-alanine, formate, fumarate, glycine, L-2-amino-3-oxobutanoate, L-alanine, L-asparagine, L-aspartate, L-glutamate, L-isoleucine, N-formimino-L-glutamate, N-formyl-L-glutamate, N2-succinylglutamate, and pyruvate.

[0121]

[0121] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0122]

[0122] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0123]

[0123] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0124]

[0124] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0125]

[0125] In some embodiments, the animals do not develop footpad dermatitis or develop less severe footpad dermatitis (e.g., as measured according to Table 17) compared to the comparable control animals administered an equivalent nutritional composition comprising the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0126]

[0126] In some embodiments, the severity of footpad dermatitis is measured according to the system in Table 17.

[0127]

[0127] In some embodiments, the animals administered the synthetic oligosaccharide preparation do not develop footpad dermatitis of scores 2, 3, or 4 as referenced in Table 17.

[0128]

[0128] In some embodiments, the animals administered the synthetic oligosaccharide preparation do not develop footpad dermatitis scores of 3 or 4 as referenced in Table 17.

[0129]

[0129] In some embodiments, the animal excretes urine and feces in liters, and the liters are in contact with the animal's paws.

[0130]

[0130] In some embodiments, the pH of the liter sample is less than the pH of a liter sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0131]

[0131] In some embodiments, the pH of the liter is at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1 pH unit less than a sample of the liter from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0132]

[0132] In some embodiments, the quality of the liter sample is improved compared to a liter sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation, and the quality of the liter is assessed according to Table 16.

[0133]

[0133] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0134]

[0134] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0135]

[0135] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0136]

[0136] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0137]

[0137] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0138]

[0138] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0139]

[0139] In some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0140] In some embodiments, the nutritional composition contains about 100 ppm to 2000 ppm, 100 ppm to 1500 ppm, 100 ppm to 1000 ppm, 100 ppm to 900 ppm, 100 ppm to 800 ppm, 100 ppm to 700 ppm, 100 ppm to 600 ppm, 100 ppm to 500 ppm, 100 ppm to 400 ppm, 100 ppm to The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0141]

[0141] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0142]

[0142] In some embodiments, the animal has an increased body weight compared to the animal's body weight before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0143]

[0143] In some embodiments, the body weight of the animal increases by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the body weight of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0144]

[0144] In some embodiments, the weight gain is greater than that of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0145]

[0145] In some embodiments, the body weight of the animal increases by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0146]

[0146] In some embodiments, the animal has increased feed efficiency compared to the animal's feed efficiency prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0147]

[0147] In some embodiments, the feed efficiency of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the feed efficiency of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0148]

[0148] In some embodiments, the animal has an increase in feed efficiency that is greater than an increase compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0149]

[0149] In some embodiments, the increase in feed efficiency of the animal is at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of the equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0150]

[0150] In some embodiments, the animal has a decreased feed conversion ratio (FCR) compared to the animal's FCR prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0151]

[0151] In some embodiments, the feed conversion rate of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion rate of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0152]

[0152] In some embodiments, the animal has a reduction in the feed conversion ratio that is greater than a reduction compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0153]

[0153] In some embodiments, the feed conversion ratio of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion ratio of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0154]

[0154] In some embodiments, the life expectancy or survival rate of the animal is increased compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0155]

[0155] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0156]

[0156] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to the animal prior to administration of the synthetic oligosaccharide preparation.

[0157]

[0157] In some embodiments, administration results in improved quality of meat from the animal compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0158]

[0158] In some embodiments, the administration results in at least one of a) improved color of the animal's meat, b) improved flavor of the animal's meat, and c) improved tenderness of the animal's meat.

[0159]

[0159] In some embodiments, the animal is poultry, aquatic animals, sheep, cattle, cattle, buffalo, bison, pig, cat, dog, rabbit, goat, guinea pig, donkey, camel, horse, pigeon, ferret, gerbil, hamster, mouse, rat, fish or bird.

[0160] In some embodiments, the animal is poultry. In some embodiments, the poultry is a chicken, turkey, duck, or goose. In some embodiments, the poultry is a chicken. In some embodiments, the chicken is a broiler chicken, layer chicken, or breeder chicken.

[0161] In some embodiments, the animal is a pig. In some embodiments, the pig is a girl pig, a grower pig, or a finisher pig.

[0162] In some embodiments, the animal is a fish. In some embodiments, the fish is a salmon, tilapia, or tropical fish.

[0163]

[0163] In some embodiments, the animal is a livestock animal.

[0164] In some embodiments, the animal is a companion animal, hi some embodiments, the companion animal is a cat, dog, hamster, rabbit, guinea pig, ferret, gerbil, bird, or mouse.

[0165]

[0165] In some embodiments, the relative abundance of oligosaccharides in DP fractions of at least 5, 10, 20, or 30 decreases monotonically with their degree of polymerization.

[0166] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with its degree of polymerization. In some embodiments, n is at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 1 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100.

[0167]

[0167] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0168]

[0168] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0169]

[0169] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0170]

[0170] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0171]

[0171] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0172]

[0172] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0173]

[0173] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0174]

[0174] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0175]

[0175] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0176]

[0176] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0177]

[0177] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 15%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0178]

[0178] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0179]

[0179] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0180]

[0180] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0181]

[0181] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0182]

[0182] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 2% to about 12%.

[0183]

[0183] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%.

[0184]

[0184] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 1% to about 10%.

[0185]

[0185] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 5% to about 10%.

[0186]

[0186] In some embodiments, the DP2 fraction contains anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0187]

[0187] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0188]

[0188] In some embodiments, the DP3 fraction comprises anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0189]

[0189] In some embodiments, the oligosaccharide preparation comprises an anhydrosubunit-containing oligosaccharides in a relative abundance of greater than 0.5%, 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0190]

[0190] In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 1% to about 40% by weight as determined by liquid chromatography.

[0191]

[0191] In some embodiments, the oligosaccharide preparation has a DP2 fraction content of about 1% to about 35% by weight as determined by liquid chromatography.

[0192]

[0192] In some embodiments, the oligosaccharide preparation has a DP3 fraction content of about 1% to about 30% by weight as determined by liquid chromatography.

[0193]

[0193] In some embodiments, the oligosaccharide preparation has a DP4 fraction content of about 0.1% to about 20% by weight as determined by liquid chromatography.

[0194]

[0194] In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 0.1% to about 15% by weight as determined by liquid chromatography.

[0195]

[0195] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction is about 0.02 to about 0.40 as determined by liquid chromatography.

[0196]

[0196] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction is about 0.01 to about 0.30 as determined by liquid chromatography.

[0197]

[0197] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 40%, or less than 30% as determined by liquid chromatography.

[0198]

[0198] In some embodiments, the oligosaccharide preparation comprises at least 10 3 , at least 10 4 , at least 10 5 , at least 10 6 , or at least 10 9 different oligosaccharide species.

[0199]

[0200] In some embodiments, two or more independent oligosaccharides comprise different anhydro subunits.

[0200]

[0200] In some embodiments, each of the anhydro-subunit-containing oligosaccharides comprises one or more anhydro-subunits that are the thermal dehydration product of a monosaccharide.

[0201] In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits selected from anhydro-glucose, anhydro-galactose, anhydro-mannose, anhydro-allose, anhydro-altrose, anhydro-gulose, anhydro-indose, anhydro-talose, anhydro-fructose, anhydro-ribose, anhydro-arabinose, anhydro-rhamnose, anhydro-lyxose, and anhydro-xylose.

[0202]

[0202] In some embodiments, the oligosaccharide preparation comprises one or more anhydro-glucose, anhydro-galactose, anhydro-mannose, or anhydro-fructose subunits.

[0203]

[0203] In some embodiments, the DP1 fraction comprises 1,6-anhydro-β-D-glucofuranose anhydro subunits or 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0204]

[0204] In some embodiments, the DP1 fraction contains both 1,6-anhydro-β-D-glucofuranose anhydro subunits and 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0205] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide complement is about 10:1 to 1:10, about 9:1 to about 1:10, about 8:1 to about 1:10, about 7:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, or about 6:1 to about 1:10. :10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 10:1 to about 1:9, about 10:1 to about 1:8, about 10:1 to about 1:7, about 10:1 to about 1:6, about 10:1 to about 1:5, about 10:1 to about 1:4, about 10:1 to about 1:3, about 10:1 to about 1:2, or about 1:1 to about 3:1.

[0206]

[0206] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide preparation is about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9 or about 1:10.

[0207]

[0207] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in the oligosaccharide preparation.

[0208]

[0208] In some embodiments, the DP2 fraction comprises at least five anhydro-subunit-containing oligosaccharides.

[0209]

[0209] In some embodiments, the DP2 fraction comprises about 5-10 anhydro-subunit-containing oligosaccharides.

[0210] In some embodiments, the oligosaccharide preparation comprises one or more sugar caramelization products, in some embodiments, the sugar caramelization products are selected from the group consisting of methanol, ethanol, furan, methylglyoxal, 2-methylfuran, vinyl acetate, glycolaldehyde, acetic acid, acetol, furfural, 2-furanmethanol, 3-furanmethanol, 2-hydroxycyclopent-2-en-1-one, 5-methylfurfural, 2(5H)-furanone, 2-methylcyclopentenolone, levoglucosenone, cyclic hydroxylactones, 1,4,3,6-dianhydro-α-D-glucopyranose, dianhydroglucopyranose, and 5-hydroxymethylfurfural (5-hmf).

[0211]

[0211] In some embodiments, greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0212]

[0212] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 5000 g / mol as determined by high performance liquid chromatography (HPLC).

[0213]

[0213] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0214]

[0214] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0215]

[0215] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0216]

[0216] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 5000 g / mol as determined by HPLC.

[0217]

[0217] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0218]

[0218] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0219]

[0219] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0220]

[0220] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 2000 to about 2800 g / mol.

[0221]

[0221] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 1000 to about 2000 g / mol.

[0222]

[0222] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0223]

[0223] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0224]

[0224] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0225]

[0225] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0226]

[0226] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0227] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation, hi some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0228] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0229]

[0229] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0230]

[0230] In one aspect, provided herein is a method of reducing harmful amino acid degradation in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of a plurality of metabolites associated with amino acid degradation are lower in a gastrointestinal sample from the animal compared to a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0231] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0232] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (3S,5S)-3,5-diaminohexanoate, (S)-3-methyl-2-oxopentanoate, (S)-5-amino-3-oxohexanoate, 2-oxoglutarate, acetyl-CoA, ammonia, D-alanine, formate, fumarate, glycine, L-2-amino-3-oxobutanoate, L-alanine, L-asparagine, L-aspartate, L-glutamate, L-isoleucine, N-formimino-L-glutamate, N-formyl-L-glutamate, N2-succinylglutamate, and pyruvate.

[0233]

[0233] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0234] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0235]

[0235] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0236]

[0236] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0237]

[0237] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0238]

[0238] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0239]

[0239] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0240]

[0240] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0241]

[0241] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0242]

[0242] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0243]

[0243] In some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0244] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0245]

[0245] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0246]

[0246] In some embodiments, the animal has an increased body weight compared to the animal's body weight before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0247]

[0247] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the body weight of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0248]

[0248] In some embodiments, the weight gain is greater than that of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0249]

[0249] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0250]

[0250] In some embodiments, the animal has increased feed efficiency compared to the animal's feed efficiency prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0251]

[0251] In some embodiments, the feed efficiency of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed efficiency of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0252]

[0252] In some embodiments, the animal has an increase in feed efficiency that is greater than an increase compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0253]

[0253] In some embodiments, the increase in feed efficiency of the animal is at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of the equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0254]

[0254] In some embodiments, the animal has a decreased feed conversion ratio (FCR) compared to the animal's FCR before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0255]

[0255] In some embodiments, the feed conversion rate of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion rate of the animal before administration of the nutritional composition containing the synthetic oligosaccharide preparation.

[0256]

[0256] In some embodiments, the animal has a reduction in the feed conversion ratio that is greater than a reduction compared to an equivalent control animal administered an equivalent nutritional composition that includes the basal nutritional composition but does not include the synthetic oligosaccharide preparation.

[0257]

[0257] In some embodiments, the feed conversion ratio of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion ratio of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0258]

[0258] In some embodiments, the life expectancy or survival rate of the animal is increased compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0259]

[0259] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0260]

[0260] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to the animal prior to administration of the synthetic oligosaccharide preparation.

[0261]

[0261] In some embodiments, administration results in improved quality of meat from the animal compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0262]

[0262] In some embodiments, the administration results in at least one of a) improved color of the animal's meat, b) improved flavor of the animal's meat, and c) improved tenderness of the animal's meat.

[0263] In some embodiments, the animal is poultry, aquatic, sheep, cattle, cattle, buffalo, bison, pig, cat, dog, rabbit, goat, guinea pig, donkey, camel, horse, pigeon, ferret, gerbil, hamster, mouse, rat, fish or bird.

[0264] In some embodiments, the animal is poultry. In some embodiments, the poultry is a chicken, turkey, duck, or goose. In some embodiments, the poultry is a chicken. In some embodiments, the chicken is a broiler chicken, layer chicken, or breeder chicken.

[0265]

[0265] In some embodiments, the animal is a pig. In some embodiments, the pig is a girl pig, a grower pig, or a finisher pig.

[0266] In some embodiments, the animal is a fish. In some embodiments, the fish is a salmon, tilapia, or tropical fish.

[0267]

[0267] In some embodiments, the animal is a livestock animal.

[0268] In some embodiments, the animal is a companion animal, hi some embodiments, the companion animal is a cat, dog, hamster, rabbit, guinea pig, ferret, gerbil, bird, or mouse.

[0269]

[0269] In some embodiments, the relative abundance of oligosaccharides in DP fractions of at least 5, 10, 20, or 30 decreases monotonically with their degree of polymerization.

[0270] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with its degree of polymerization. In some embodiments, n is at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 1 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100.

[0271]

[0271] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0272]

[0272] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0273]

[0273] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0274]

[0274] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0275]

[0275] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0276]

[0276] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0277]

[0277] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0278]

[0278] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0279]

[0279] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0280]

[0280] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0281]

[0281] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 15%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0282]

[0282] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0283]

[0283] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0284]

[0284] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0285]

[0285] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0286]

[0286] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 2% to about 12%.

[0287]

[0287] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%.

[0288]

[0288] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 1% to about 10%.

[0289]

[0289] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 5% to about 10%.

[0290]

[0290] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0291]

[0291] In some embodiments, the DP1 fraction contains anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0292]

[0292] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0293]

[0293] In some embodiments, the oligosaccharide preparation comprises an anhydro subunit-containing oligosaccharides in a relative abundance of greater than 0.5%, 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0294]

[0294] In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 1% to about 40% by weight as determined by liquid chromatography.

[0295]

[0295] In some embodiments, the oligosaccharide preparation has a DP2 fraction content of about 1% to about 35% by weight as determined by liquid chromatography.

[0296]

[0296] In some embodiments, the oligosaccharide preparation has a DP3 fraction content of about 1% to about 30% by weight as determined by liquid chromatography.

[0297]

[0297] In some embodiments, the oligosaccharide preparation has a DP4 fraction content of about 0.1% to about 20% by weight as determined by liquid chromatography.

[0298]

[0298] In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 0.1% to about 15% by weight as determined by liquid chromatography.

[0299]

[0299] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction is about 0.02 to about 0.40 as determined by liquid chromatography.

[0300]

[0300] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction is about 0.01 to about 0.30 as determined by liquid chromatography.

[0301]

[0301] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 40%, or less than 30% as determined by liquid chromatography.

[0302]

[0302] In some embodiments, the oligosaccharide preparation comprises at least 10 3 , at least 10 4 , at least 10 5 , at least 10 6 , or at least 10 9 different oligosaccharide species.

[0303]

[0303] In some embodiments, two or more independent oligosaccharides comprise different anhydro subunits.

[0304]

[0304] In some embodiments, each of the anhydro-subunit-containing oligosaccharides comprises one or more anhydro-subunits that are the thermal dehydration product of a monosaccharide.

[0305] In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits selected from anhydro-glucose, anhydro-galactose, anhydro-mannose, anhydro-allose, anhydro-altrose, anhydro-gulose, anhydro-indose, anhydro-talose, anhydro-fructose, anhydro-ribose, anhydro-arabinose, anhydro-rhamnose, anhydro-lyxose, and anhydro-xylose.

[0306]

[0306] In some embodiments, the oligosaccharide preparation comprises one or more anhydro-glucose, anhydro-galactose, anhydro-mannose, or anhydro-fructose subunits.

[0307]

[0307] In some embodiments, the DP1 fraction comprises 1,6-anhydro-β-D-glucofuranose anhydro subunits or 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0308]

[0308] In some embodiments, the DP1 fraction contains both 1,6-anhydro-β-D-glucofuranose anhydro subunits and 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0309] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide complement is about 10:1 to 1:10, about 9:1 to about 1:10, about 8:1 to about 1:10, about 7:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, or about 6:1 to about 1:10. :10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 10:1 to about 1:9, about 10:1 to about 1:8, about 10:1 to about 1:7, about 10:1 to about 1:6, about 10:1 to about 1:5, about 10:1 to about 1:4, about 10:1 to about 1:3, about 10:1 to about 1:2, or about 1:1 to about 3:1.

[0310]

[0310] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide preparation is about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9, or about 1:10.

[0311]

[0311] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in the oligosaccharide preparation.

[0312]

[0312] In some embodiments, the DP2 fraction comprises at least five anhydro-subunit-containing oligosaccharides.

[0313]

[0313] In some embodiments, the DP2 fraction comprises about 5-10 anhydro-subunit-containing oligosaccharides.

[0314] In some embodiments, the oligosaccharide preparation comprises one or more sugar caramelization products selected from the group consisting of methanol, ethanol, furan, methylglyoxal, 2-methylfuran, vinyl acetate, glycolaldehyde, acetic acid, acetol, furfural, 2-furanmethanol, 3-furanmethanol, 2-hydroxycyclopent-2-en-1-one, 5-methylfurfural, 2(5H)-furanone, 2-methylcyclopentenolone, levoglucosenone, cyclic hydroxylactones, 1,4,3,6-dianhydro-α-D-glucopyranose, dianhydroglucopyranose, and 5-hydroxymethylfurfural (5-hmf).

[0315]

[0315] In some embodiments, greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0316]

[0316] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 5000 g / mol as determined by high performance liquid chromatography (HPLC).

[0317]

[0317] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0318]

[0318] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0319]

[0319] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0320]

[0320] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 5000 g / mol as determined by HPLC.

[0321]

[0321] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0322]

[0322] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0323]

[0323] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0324]

[0324] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 2000 to about 2800 g / mol.

[0325]

[0325] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 1000 to about 2000 g / mol.

[0326]

[0326] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0327]

[0327] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0328]

[0328] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0329]

[0329] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0330]

[0330] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0331] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation, hi some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0332] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0333]

[0333] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0334]

[0334] In one aspect, provided herein is a method of improving carbon utilization in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of a plurality of metabolites associated with improved carbon utilization in a gastrointestinal sample from the animal are higher compared to a comparable control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0335]

[0335] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0336] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (R)-lactate, (R)-lactoyl-CoA, (S)-lactate, (S)-propane-1,2-diol, 1-propanal, acetate, acetyl-CoA, acryloyl-CoA, propanoate, propanoyl-CoA, and pyruvate.

[0337]

[0337] In some embodiments, the level of at least the metabolite is at least about 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0338] In some embodiments, the level of at least the metabolite is at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0339]

[0339] In some embodiments, levels of multiple metabolites associated with energy metabolism are higher in gastrointestinal samples from the animals compared to gastrointestinal samples from comparable control animals administered an equivalent nutritional composition containing the basic nutritional composition but not the synthetic oligosaccharide preparation.

[0340]

[0340] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0341]

[0341] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of 2-oxoglutarate, fumarate, L-alanine, L-glutamate, oxaloacetate, propanoyl-CoA, pyruvate, and succinate.

[0342]

[0342] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0343] In some embodiments, the level of the plurality of metabolites in the gastrointestinal sample is at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold higher than the level of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0344]

[0344] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0345]

[0345] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0346] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (3R)-3-hydroxybutanoyl-CoA, (R)-lactate, (R)-lactoyl-CoA, (S)-3-aminobutanoyl-CoA, (S)-3-hydroxy-isobutanoate, (S)-3-hydroxy-isobutanoyl-CoA, (S)-3-hydroxybutanoyl-CoA, (S)-5-amino-3-oxohexanoate, (S)-lactate, 4-hydroxybutanoate, acetate, acetoacetate, acetoacetyl-CoA, acetyl-CoA, butanoate, butanoyl-CoA, coenzyme A, crotonyl-CoA, succinate, succinate semialdehyde, and succinyl-CoA.

[0347]

[0347] In some embodiments, the level of at least the metabolite is at least about 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0348] In some embodiments, the level of at least the metabolite is at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0349]

[0349] In some embodiments, the levels of multiple metabolites associated with carbon utilization are lower in gastrointestinal samples from the animals compared to gastrointestinal samples from comparable control animals administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0350]

[0350] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0351]

[0351] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0352] In some embodiments, the levels of the plurality of metabolites in the gastrointestinal sample are at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold lower than the levels of the plurality of metabolites in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition comprising the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0353]

[0353] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0354]

[0354] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0355]

[0355] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0356]

[0356] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0357]

[0357] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0358]

[0358] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0359]

[0359] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0360]

[0360] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0361]

[0361] In some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0362] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0363]

[0363] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0364]

[0364] In some embodiments, the animal has an increased body weight compared to the animal's body weight before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0365]

[0365] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the body weight of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0366]

[0366] In some embodiments, the weight gain is greater than that of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0367]

[0367] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0368]

[0368] In some embodiments, the animal has increased feed efficiency compared to the animal's feed efficiency prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0369]

[0369] In some embodiments, the feed efficiency of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed efficiency of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0370]

[0370] In some embodiments, the animal has an increase in feed efficiency that is greater than an increase compared to an equivalent control animal administered an equivalent nutritional composition that includes the basal nutritional composition but does not include the synthetic oligosaccharide preparation.

[0371]

[0371] In some embodiments, the increase in feed efficiency of the animal is at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of the equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0372]

[0372] In some embodiments, the animal has a decreased feed conversion ratio (FCR) compared to the FCR of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0373]

[0373] In some embodiments, the feed conversion rate of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion rate of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0374]

[0374] In some embodiments, the animal has a reduction in the feed conversion ratio that is greater than a reduction compared to an equivalent control animal administered an equivalent nutritional composition that includes the basal nutritional composition but does not include the synthetic oligosaccharide preparation.

[0375]

[0375] In some embodiments, the feed conversion ratio of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion ratio of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0376]

[0376] In some embodiments, the life expectancy or survival rate of the animal is increased compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0377]

[0377] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0378]

[0378] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to the animal prior to administration of the synthetic oligosaccharide preparation.

[0379]

[0379] In some embodiments, administration results in improved quality of meat from the animal compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0380]

[0380] In some embodiments, the administration results in at least one of a) improved color of the animal's meat, b) improved flavor of the animal's meat, and c) improved tenderness of the animal's meat.

[0381] In some embodiments, the animal is poultry, aquatic, sheep, cattle, cattle, buffalo, bison, pig, cat, dog, rabbit, goat, guinea pig, donkey, camel, horse, pigeon, ferret, gerbil, hamster, mouse, rat, fish or bird.

[0382] In some embodiments, the animal is poultry. In some embodiments, the poultry is a chicken, turkey, duck, or goose. In some embodiments, the poultry is a chicken. In some embodiments, the chicken is a broiler chicken, layer chicken, or breeder chicken.

[0383] In some embodiments, the animal is a pig. In some embodiments, the pig is a girl pig, a grower pig, or a finisher pig.

[0384] In some embodiments, the animal is a fish. In some embodiments, the fish is a salmon, tilapia, or tropical fish.

[0385]

[0385] In some embodiments, the animal is a livestock animal.

[0386] In some embodiments, the animal is a companion animal, hi some embodiments, the companion animal is a cat, dog, hamster, rabbit, guinea pig, ferret, gerbil, bird, or mouse.

[0387]

[0387] In some embodiments, the relative abundance of oligosaccharides in DP fractions of at least 5, 10, 20, or 30 decreases monotonically with their degree of polymerization.

[0388] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with its degree of polymerization. In some embodiments, n is at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 1 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100.

[0389]

[0389] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0390]

[0390] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0391]

[0391] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0392]

[0392] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0393]

[0393] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0394]

[0394] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0395]

[0395] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0396]

[0396] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0397]

[0397] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0398]

[0398] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0399]

[0399] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 15%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0400]

[0400] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0401]

[0401] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0402]

[0402] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0403]

[0403] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0404]

[0404] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 2% to about 12%.

[0405]

[0405] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%.

[0406]

[0406] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 1% to about 10%.

[0407]

[0407] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 5% to about 10%.

[0408]

[0408] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0409]

[0409] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0410]

[0410] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0411]

[0411] In some embodiments, the oligosaccharide preparation comprises an anhydrosubunit-containing oligosaccharides in a relative abundance of greater than 0.5%, 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0412]

[0412] In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 1% to about 40% by weight as determined by liquid chromatography.

[0413]

[0413] In some embodiments, the oligosaccharide preparation has a DP2 fraction content of about 1% to about 35% by weight as determined by liquid chromatography.

[0414]

[0414] In some embodiments, the oligosaccharide preparation has a DP3 fraction content of about 1% to about 30% by weight as determined by liquid chromatography.

[0415]

[0415] In some embodiments, the oligosaccharide preparation has a DP4 fraction content of about 0.1% to about 20% by weight as determined by liquid chromatography.

[0416]

[0416] In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 0.1% to about 15% by weight as determined by liquid chromatography.

[0417]

[0417] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction is about 0.02 to about 0.40 as determined by liquid chromatography.

[0418]

[0418] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction is about 0.01 to about 0.30 as determined by liquid chromatography.

[0419]

[0419] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 40%, or less than 30% as determined by liquid chromatography.

[0420]

[0420] In some embodiments, the oligosaccharide preparation comprises at least 10 3 , at least 10 4 , at least 10 5 , at least 10 6 , or at least 10 9 different oligosaccharide species.

[0421]

[0421] In some embodiments, two or more independent oligosaccharides comprise different anhydro subunits.

[0422]

[0422] In some embodiments, each of the anhydro-subunit-containing oligosaccharides comprises one or more anhydro-subunits that are the thermal dehydration products of a monosaccharide.

[0423] In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits selected from anhydro-glucose, anhydro-galactose, anhydro-mannose, anhydro-allose, anhydro-altrose, anhydro-gulose, anhydro-indose, anhydro-talose, anhydro-fructose, anhydro-ribose, anhydro-arabinose, anhydro-rhamnose, anhydro-lyxose, and anhydro-xylose.

[0424]

[0424] In some embodiments, the oligosaccharide preparation comprises one or more anhydro-glucose, anhydro-galactose, anhydro-mannose, or anhydro-fructose subunits.

[0425]

[0425] In some embodiments, the DP1 fraction comprises 1,6-anhydro-β-D-glucofuranose anhydro subunits or 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0426]

[0426] In some embodiments, the DP1 fraction contains both 1,6-anhydro-β-D-glucofuranose anhydro subunits and 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0427] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide complement is about 10:1 to 1:10, about 9:1 to about 1:10, about 8:1 to about 1:10, about 7:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, or about 6:1 to about 1:10. :10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 10:1 to about 1:9, about 10:1 to about 1:8, about 10:1 to about 1:7, about 10:1 to about 1:6, about 10:1 to about 1:5, about 10:1 to about 1:4, about 10:1 to about 1:3, about 10:1 to about 1:2, or about 1:1 to about 3:1.

[0428]

[0428] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide preparation is about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9 or about 1:10.

[0429]

[0429] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in the oligosaccharide preparation.

[0430]

[0430] In some embodiments, the DP2 fraction comprises at least five anhydro-subunit-containing oligosaccharides.

[0431]

[0431] In some embodiments, the DP2 fraction comprises about 5-10 anhydro-subunit-containing oligosaccharides.

[0432] In some embodiments, the oligosaccharide preparation comprises one or more sugar caramelization products selected from the group consisting of methanol, ethanol, furan, methylglyoxal, 2-methylfuran, vinyl acetate, glycolaldehyde, acetic acid, acetol, furfural, 2-furanmethanol, 3-furanmethanol, 2-hydroxycyclopent-2-en-1-one, 5-methylfurfural, 2(5H)-furanone, 2-methylcyclopentenolone, levoglucosenone, cyclic hydroxylactones, 1,4,3,6-dianhydro-α-D-glucopyranose, dianhydroglucopyranose, and 5-hydroxymethylfurfural (5-hmf).

[0433]

[0433] In some embodiments, greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0434]

[0434] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 5000 g / mol as determined by high performance liquid chromatography (HPLC).

[0435]

[0435] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0436]

[0436] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0437]

[0437] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0438]

[0438] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 5000 g / mol as determined by HPLC.

[0439]

[0439] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0440]

[0440] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0441]

[0441] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0442]

[0442] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 2000 to about 2800 g / mol.

[0443]

[0443] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 1000 to about 2000 g / mol.

[0444]

[0444] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0445]

[0445] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0446]

[0446] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0447]

[0447] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0448]

[0448] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0449] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation, hi some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0450] In some embodiments, the nutritional composition comprises an α-tocopherol concentration of about 100 ppm to 2000 ppm, 100 ppm to 1500 ppm, 100 ppm to 1000 ppm, 100 ppm to 900 ppm, 100 ppm to 800 ppm, 100 ppm to 700 ppm, 100 ppm to 600 ppm, 100 ppm to 500 ppm, 100 ppm to 400 ppm, 100 ppm to The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0451]

[0451] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0452]

[0452] In one aspect, provided herein is a method of improving energy metabolism in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of a plurality of metabolites associated with improved energy metabolism in a gastrointestinal sample from the animal are higher compared to a comparable control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0453]

[0453] In some embodiments, the plurality includes at least 3, 4, 5, 6, 7, 8, 9, or 10 metabolites.

[0454]

[0454] In some embodiments, the plurality includes at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of 2-oxoglutarate, fumarate, L-alanine, L-glutamate, oxaloacetate, propanoyl-CoA, pyruvate, and succinate.

[0455]

[0455] In some embodiments, the level of at least the metabolite is at least about 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0456]

[0456] In some embodiments, the level of at least the metabolite is at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, or 10-fold higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0457]

[0457] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0458]

[0458] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0459]

[0459] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0460]

[0460] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0461]

[0461] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0462]

[0462] In some embodiments, the animal ingests at least a portion of the nutritional composition over at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 24-hour periods.

[0463]

[0463] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0464]

[0464] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm or 2000 ppm of the synthetic oligosaccharide preparation.

[0465]

[0465] In some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0466] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0467]

[0467] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0468]

[0468] In some embodiments, the animal has an increased body weight compared to the animal's body weight before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0469]

[0469] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9% or 10% compared to the body weight of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0470]

[0470] In some embodiments, the weight gain is greater than that of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0471]

[0471] In some embodiments, the body weight of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0472]

[0472] In some embodiments, the animal has increased feed efficiency compared to the animal's feed efficiency prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0473]

[0473] In some embodiments, the feed efficiency of the animal is increased by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed efficiency of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0474]

[0474] In some embodiments, the animal has an increase in feed efficiency that is greater than an increase compared to an equivalent control animal administered an equivalent nutritional composition that includes the basal nutritional composition but does not include the synthetic oligosaccharide preparation.

[0475]

[0475] In some embodiments, the increase in feed efficiency of the animal is at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the body weight of the equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0476]

[0476] In some embodiments, the animal has a decreased feed conversion ratio (FCR) compared to the FCR of the animal prior to administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0477]

[0477] In some embodiments, the feed conversion rate of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion rate of the animal before administration of the nutritional composition comprising the synthetic oligosaccharide preparation.

[0478]

[0478] In some embodiments, the animal has a reduction in the feed conversion ratio that is greater than a reduction compared to an equivalent control animal administered an equivalent nutritional composition that includes the basal nutritional composition but does not include the synthetic oligosaccharide preparation.

[0479]

[0479] In some embodiments, the feed conversion ratio of the animal is reduced by at least 1%, 2%, 3%, 4%, 5%, 5%, 7%, 8%, 9%, or 10% compared to the feed conversion ratio of an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0480]

[0480] In some embodiments, the life expectancy or survival rate of the animal is increased compared to an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0481]

[0481] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0482]

[0482] In some embodiments, administration results in at least one of: a) improved nutrient absorption, b) improved mitochondrial function, c) improved liver function, d) improved renal function, e) improved sociability, f) improved physiological mood, g) improved energy, h) improved satiety; and i) improved alertness, each compared to the animal prior to administration of the synthetic oligosaccharide preparation.

[0483]

[0483] In some embodiments, administration results in improved quality of meat from the animal compared to an animal administered a nutritional composition that does not include the synthetic oligosaccharide preparation.

[0484]

[0484] In some embodiments, the administration results in at least one of a) improved color of the animal's meat, b) improved flavor of the animal's meat, and c) improved tenderness of the animal's meat.

[0485]

[0485] In some embodiments, the animal is poultry, aquatic, sheep, cattle, cattle, buffalo, bison, pig, cat, dog, rabbit, goat, guinea pig, donkey, camel, horse, pigeon, ferret, gerbil, hamster, mouse, rat, fish or bird.

[0486] In some embodiments, the animal is poultry. In some embodiments, the poultry is a chicken, turkey, duck, or goose. In some embodiments, the poultry is a chicken. In some embodiments, the chicken is a broiler chicken, layer chicken, or breeder chicken.

[0487] In some embodiments, the animal is a pig. In some embodiments, the pig is a girl pig, a grower pig, or a finisher pig.

[0488] In some embodiments, the animal is a fish. In some embodiments, the fish is a salmon, tilapia, or tropical fish.

[0489]

[0489] In some embodiments, the animal is a livestock animal.

[0490] In some embodiments, the animal is a companion animal, hi some embodiments, the companion animal is a cat, dog, hamster, rabbit, guinea pig, ferret, gerbil, bird, or mouse.

[0491]

[0491] In some embodiments, the relative abundance of oligosaccharides in DP fractions of at least 5, 10, 20, or 30 decreases monotonically with their degree of polymerization.

[0492] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with its degree of polymerization. In some embodiments, n is at least 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 1 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99 or 100.

[0493]

[0493] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0494]

[0494] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0495]

[0495] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0496]

[0496] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0497]

[0497] In some embodiments, the DP2 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0498]

[0498] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0499]

[0499] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0500]

[0500] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0501]

[0501] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0502]

[0502] In some embodiments, the DP1 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0503]

[0503] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of less than 15%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1%.

[0504]

[0504] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 2% to about 12%.

[0505]

[0505] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 1% to about 10%.

[0506]

[0506] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 10%.

[0507]

[0507] In some embodiments, the DP3 fraction comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 5% to about 10%.

[0508]

[0508] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 2% to about 12%.

[0509]

[0509] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%.

[0510]

[0510] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 1% to about 10%.

[0511]

[0511] In some embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 5% to about 10%.

[0512]

[0512] In some embodiments, the DP2 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0513]

[0513] In some embodiments, the DP1 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0514]

[0514] In some embodiments, the DP3 fraction contains anhydrosubunit-containing oligosaccharides at a relative abundance of greater than 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0515]

[0515] In some embodiments, the oligosaccharide preparation comprises an anhydrosubunit-containing oligosaccharides in a relative abundance of greater than 0.5%, 0.6%, greater than 0.8%, greater than 1.0%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, or greater than 12%.

[0516]

[0516] In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 1% to about 40% by weight as determined by liquid chromatography.

[0517]

[0517] In some embodiments, the oligosaccharide preparation has a DP2 fraction content of about 1% to about 35% by weight as determined by liquid chromatography.

[0518]

[0518] In some embodiments, the oligosaccharide preparation has a DP3 fraction content of about 1% to about 30% by weight as determined by liquid chromatography.

[0519]

[0519] In some embodiments, the oligosaccharide preparation has a DP4 fraction content of about 0.1% to about 20% by weight as determined by liquid chromatography.

[0520]

[0520] In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 0.1% to about 15% by weight as determined by liquid chromatography.

[0521]

[0521] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction is about 0.02 to about 0.40 as determined by liquid chromatography.

[0522]

[0522] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction is about 0.01 to about 0.30 as determined by liquid chromatography.

[0523]

[0523] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 40%, or less than 30% as determined by liquid chromatography.

[0524]

[0524] In some embodiments, the oligosaccharide preparation comprises at least 10 3 , at least 10 4 , at least 10 5 , at least 10 6 , or at least 10 9 different oligosaccharide species.

[0525]

[0525] In some embodiments, two or more independent oligosaccharides comprise different anhydro subunits.

[0526]

[0526] In some embodiments, each of the anhydro-subunit-containing oligosaccharides comprises one or more anhydro-subunits that are the thermal dehydration product of a monosaccharide.

[0527] In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits selected from anhydro-glucose, anhydro-galactose, anhydro-mannose, anhydro-allose, anhydro-altrose, anhydro-gulose, anhydro-indose, anhydro-talose, anhydro-fructose, anhydro-ribose, anhydro-arabinose, anhydro-rhamnose, anhydro-lyxose, and anhydro-xylose.

[0528]

[0528] In some embodiments, the oligosaccharide preparation comprises one or more anhydro-glucose, anhydro-galactose, anhydro-mannose, or anhydro-fructose subunits.

[0529]

[0529] In some embodiments, the DP1 fraction comprises 1,6-anhydro-β-D-glucofuranose anhydro subunits or 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0530]

[0530] In some embodiments, the DP1 fraction contains both 1,6-anhydro-β-D-glucofuranose anhydro subunits and 1,6-anhydro-β-D-glucopyranose anhydro subunits.

[0531] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide complement is about 10:1 to 1:10, about 9:1 to about 1:10, about 8:1 to about 1:10, about 7:1 to about 1:10, about 6:1 to about 1:10, about 5:1 to about 1:10, or about 6:1 to about 1:10. :10, about 4:1 to about 1:10, about 3:1 to about 1:10, about 2:1 to about 1:10, about 10:1 to about 1:9, about 10:1 to about 1:8, about 10:1 to about 1:7, about 10:1 to about 1:6, about 10:1 to about 1:5, about 10:1 to about 1:4, about 10:1 to about 1:3, about 10:1 to about 1:2, or about 1:1 to about 3:1.

[0532]

[0532] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the oligosaccharide preparation is about 10:1, about 9:1, about 8:1, about 7:1, about 6:1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9 or about 1:10.

[0533]

[0533] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in the oligosaccharide preparation.

[0534]

[0534] In some embodiments, the DP2 fraction comprises at least five anhydro-subunit-containing oligosaccharides.

[0535]

[0535] In some embodiments, the DP2 fraction comprises about 5-10 anhydro-subunit-containing oligosaccharides.

[0536] In some embodiments, the oligosaccharide preparation comprises one or more sugar caramelization products selected from the group consisting of methanol, ethanol, furan, methylglyoxal, 2-methylfuran, vinyl acetate, glycolaldehyde, acetic acid, acetol, furfural, 2-furanmethanol, 3-furanmethanol, 2-hydroxycyclopent-2-en-1-one, 5-methylfurfural, 2(5H)-furanone, 2-methylcyclopentenolone, levoglucosenone, cyclic hydroxylactones, 1,4,3,6-dianhydro-α-D-glucopyranose, dianhydroglucopyranose, and 5-hydroxymethylfurfural (5-hmf).

[0537]

[0537] In some embodiments, greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0538]

[0538] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 5000 g / mol as determined by high performance liquid chromatography (HPLC).

[0539]

[0539] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0540]

[0540] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0541]

[0541] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0542]

[0542] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 5000 g / mol as determined by HPLC.

[0543]

[0543] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 300 to about 2500 g / mol as determined by HPLC.

[0544]

[0544] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 2000 g / mol as determined by HPLC.

[0545]

[0545] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 500 to about 1500 g / mol as determined by HPLC.

[0546]

[0546] In some embodiments, the oligosaccharide preparation has a weight average molecular weight of about 2000 to about 2800 g / mol.

[0547]

[0547] In some embodiments, the oligosaccharide preparation has a number average molecular weight of about 1000 to about 2000 g / mol.

[0548]

[0548] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0549]

[0549] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0550]

[0550] In some embodiments, the administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0551]

[0551] In some embodiments, the animal ingests at least a portion of the nutritional composition over a 24-hour period of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 times.

[0552]

[0552] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0553] In some embodiments, the nutritional composition comprises about 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation, hi some embodiments, the nutritional composition comprises about 500 ppm of the synthetic oligosaccharide preparation.

[0554] In some embodiments, the nutritional composition contains between about 100 ppm and 2000 ppm, 100 ppm and 1500 ppm, 100 ppm and 1000 ppm, 100 ppm and 900 ppm, 100 ppm and 800 ppm, 100 ppm and 700 ppm, 100 ppm and 600 ppm, 100 ppm and 500 ppm, 100 ppm and 400 ppm, 100 ppm and The synthetic oligosaccharide preparation may be present in an amount of 300 ppm, 100 ppm to 200 ppm, 200 ppm to 1000 ppm, 200 ppm to 800 ppm, 200 ppm to 700 ppm, 200 ppm to 600 ppm, 200 ppm to 500 ppm, 300 ppm to 1000 ppm, 300 ppm to 700 ppm, 300 ppm to 600 ppm, or 300 ppm to 500 ppm.

[0555]

[0555] In some embodiments, the nutritional composition comprises about 300 ppm to 600 ppm of the synthetic oligosaccharide preparation.

[0556]

[0556] In one aspect, the present disclosure provides a method for improving antibacterial and / or anti-inflammatory responses in an animal, the method comprising administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, wherein the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and wherein each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.5% to about 15% as determined by mass spectrometry, and wherein levels of metabolites associated with improved antibacterial and anti-inflammatory responses in a gastrointestinal sample from the animal are higher compared to a comparable control animal administered an equivalent nutritional composition comprising the basal nutritional composition and not the synthetic oligosaccharide preparation.

[0557]

[0557] In some embodiments, the metabolite is itaconate.

[0558]

[0558] In some embodiments, the level of the metabolite is at least about 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0559]

[0559] In some embodiments, the level of the metabolite is at least about 0.1-fold, 0.2-fold, 0.3-fold, 0.4-fold, 0.5-fold, 0.6-fold, 0.7-fold, 0.8-fold, 0.9-fold, 1-fold, 2-fold, 3-fold, 4-fold, or 5-fold higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

[0560]

[0560] In some embodiments, the gastrointestinal sample is a biopsy of gastrointestinal tissue, a fecal sample, a rumen fluid sample, or a cloacal swab.

[0561]

[0561] In some embodiments, the gastrointestinal tissue is cecal tissue or ileal tissue.

[0562]

[0562] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal for at least 1, 7, 10, 14, 30, 45, 60, 90, or 120 days.

[0563]

[0563] In some embodiments, the nutritional composition comprising the synthetic oligosaccharide preparation is administered to the animal at least once, twice, three times, four times, or five times daily.

[0564]

[0564] In some embodiments, administering comprises providing the nutritional composition to the animal for ad libitum consumption.

[0565]

[0565] In some embodiments, the animal ingests at least a portion of the nutritional composition over a 24-hour period of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 90, or 120 times.

[0566]

[0566] In some embodiments, the nutritional composition comprises at least 100 ppm, 200 ppm, 300 ppm, 400 ppm, 500 ppm, 600 ppm, 700 ppm, 800 ppm, 900 ppm, 1000 ppm, 1500 ppm, or 2000 ppm of the synthetic oligosaccharide preparation.

[0567] [Incorporated by reference]

[0567] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. To the extent that the publications and patents or patent applications incorporated by reference conflict with the disclosure contained in the specification, the specification supersedes and / or is intended to take precedence over such conflicting material.

[0568] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings (also referred to herein as "figures" and "FIG."). [Brief explanation of the drawings]

[0569] [Figure 1] A portion of the 1H, 13C-HSQC NMR spectrum of oligosaccharide preparation 9.2 is shown. [Figure 2] 9 shows a MALDI-MS spectrum of the oligosaccharide preparation from Example 9.7 showing the presence of anhydro subunits. [Figure 3] 1D1H NMR spectrum of the anhydro-DP1 component of the oligosaccharide preparation of Example 9. [Figure 4] 1D APT13C-NMR spectrum of the anhydro-DP1 component of the oligosaccharide preparation of Example 9. [Figure 5] FIG. 1 shows a magnified view of the GC-MS chromatogram (TIC and XIC (m / z 229) plot) for the oligosaccharide preparation of Example 2.9 after derivatization. [Figure 6] 1 shows the 1H,13C-HSQC spectrum of an oligosaccharide preparation with caramelized anhydro subunits. [Figure 7] 1 shows portions of MALDI-MS spectra comparing the oligosaccharide preparation of Example 9 at different laser energies. [Figure 8A]LC-MS / MS detection of anhydroDP2 species at concentrations of 1-80 μg / mL of the oligosaccharide preparation of Example 9 in water. [Figure 8B] Figure 8A shows a linear calibration curve obtained from LC-MS / MS detection. [Figure 9-1] Quantification of the anhydro-DP2 content of various control and treated dietary compositions is shown. [Figure 9-2] Quantification of the anhydro-DP2 content of various control and treated dietary compositions is shown. [Figure 10] 2D-1H JRES NMR spectra of anhydro-subunit-containing gluco-oligosaccharide samples. [Figure 11] 1 is a representative 1H, 13C-HSQC NMR spectrum of an anhydro subunit-containing gluco-oligosaccharide sample with relevant resonances and assignments used for bond distribution. [Figure 12] 1 shows an overlay of 1H DOSY spectra of three anhydrosubunit-containing oligosaccharides. [Figure 13] A comparison of 1,6-anhydro-β-D-glucose (DP1-18), 1,6-anhydro-β-D-cellobiose (DP2-18), and an anhydro-subunit-containing oligosaccharide sample is shown. [Figure 14] Mass chromatograms of anhydro-subunit-containing oligosaccharides (top) and digested anhydro-subunit-containing oligosaccharides (bottom) in selected MRMs are shown. [Figure 15] 1 shows a graph of relative abundance versus degree of polymerization (DP) of oligosaccharides from Example 9. The graph shows that the oligosaccharide preparations have a monotonically decreasing DP distribution. [Figure 16] 1 shows a graph of relative abundance versus degree of polymerization of oligosaccharides from Example 9. The graph shows that the oligosaccharide preparations have a non-monotonic decreasing DP distribution. [Figure 17] 1 shows a dose-dependent improvement in litter quality in broilers fed the oligosaccharide preparation of Example 9, with a statistically significant (P<0.05) improvement in litter quality compared to the control at 500 ppm. [Figure 18] Broilers fed the oligosaccharide preparation of Example 9 showed a dose-dependent reduction (lesion scoring) in footpad dermatitis, accompanied by a statistically significant (P<0.05) improvement in footpad welfare compared to controls at 500 ppm. [Figure 19] The results show a dose-dependent improvement in bird mobility as measured by mean gate scores in broilers fed the oligosaccharide preparation of Example 9, with statistically significant (P<0.05) improvements in welfare compared to controls at both the 250 and 500 ppm dose levels. [Figure 20] FIG. 1 shows the metabolic network used in the microbiome pathway analysis of Example 40, where edges n1 to n143 correspond to metabolites (and intermediates) and edges R1 to R256 correspond to biochemical reactions that can convert one metabolite (or intermediate) to another according to reactions with EC numbers in Table 18. [Figure 21] Two DP1 and one DP2 anhydrosubunit-containing oligosaccharides are shown. [Figure 22] An anhydrosubunit-containing oligosaccharide (cellotriosan) is shown. [Figure 23A] 1 shows a MALDI-MS spectrum of the oligosaccharide preparation from Example 2 showing the presence of anhydro subunits. [Figure 23B] 1 shows a MALDI-MS spectrum of the oligosaccharide preparation from Example 2 showing the presence of anhydro subunits. [Figure 24A] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 1. [Figure 24B] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 1. [Figure 24C] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 1. [Figure 25A] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 3. [Figure 25B] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 3. [Figure 25C] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 3. [Figure 26A] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 4. [Figure 26B] LC-MS / MS detection of anhydro DP2, anhydro DP1 and DP2 species of the oligosaccharide preparation of Example 4. [Figure 26C] LC-MS / MS detection of anhydro DP2, anhydro DP1 and DP2 species of the oligosaccharide preparation of Example 4. [Figure 27A] LC-MS / MS detection of anhydro DP2, anhydro DP1, and DP2 species of the oligosaccharide preparation of Example 7. [Figure 27B] LC-MS / MS detection of anhydro DP2, anhydro DP1 and DP2 species of the oligosaccharide preparation of Example 7. [Figure 27C] LC-MS / MS detection of anhydro DP2, anhydro DP1 and DP2 species of the oligosaccharide preparation of Example 7. [Figure 28A] 1 shows GC-MS spectral detection of DP1, anhydroDP1, DP2 and anhydroDP2 fractions of the oligosaccharide preparation of Example 1. [Figure 28B] A magnified view of the DP2 and anhydroDP2 fractions shown in Figure 28A is shown. [Figure 29A] 1 shows GC-MS spectral detection of DP1, anhydroDP1, DP2 and anhydroDP2 fractions of the oligosaccharide preparation of Example 3. [Figure 29B] A magnified view of the DP2 and anhydroDP2 fractions shown in Figure 29A is shown. [Figure 30A] 1 shows GC-MS spectral detection of DP1, anhydroDP1, DP2 and anhydroDP2 fractions of the oligosaccharide preparation of Example 4. [Figure 30B] A magnified view of the DP2 and anhydroDP2 fractions shown in Figure 30A is shown. [Figure 31A] 1 shows GC-MS spectral detection of DP1, anhydroDP1, DP2 and anhydroDP2 fractions of the oligosaccharide preparation of Example 7. [Figure 31B] A magnified view of the DP2 and anhydroDP2 fractions shown in Figure 31A is shown. [Figure 32] 1 shows the effect of reaction temperature, water content and reaction time on the content of DP2 anhydrosubunit-containing oligosaccharides in oligosaccharide preparations compared to oligosaccharide preparations according to Example 2. [Figure 33] NMR assignments for 1,6-anhydro-β-D-glucofuranose and 1,6-anhydro-β-D-glucopyranose are shown. [Figure 34-1] FIG. 1 shows increased abundance of desirable metabolic pathways in the microbiome metagenome of broiler chickens fed diets containing oligosaccharide preparations. [Figure 34-2] FIG. 1 shows increased abundance of desirable metabolic pathways in the microbiome metagenome of broiler chickens fed diets containing oligosaccharide preparations. [Figure 35] 1 shows MALDI-MS spectra comparing oligosaccharide preparations from Example 9 at different laser energies. [Figure 36-1] We show high variability in microbiome strain composition between otherwise identical broiler chickens fed the same diet and grown under identical conditions in the same broiler house. [Figure 36-2] We show high variability in microbiome strain composition between otherwise identical broiler chickens fed the same diet and grown under identical conditions in the same broiler house. [Figure 37] Figure 36 shows relatively high consistency of key microbiome metabolic functions in the same birds where microbiome phylogenetic composition was variable. [Figure 38]1 shows upregulation of microbiome function associated with health and nutritional benefits by feeding the oligosaccharide preparation from Example 9.

[0570] [Detailed explanation]

[0607] The following description and examples will explain in detail the embodiments of the present disclosure. It should be understood that the present disclosure is not limited to the specific embodiments described herein and may vary as such. Those skilled in the art will appreciate that there are many variations and modifications of the present disclosure that are within its scope.

[0571]

[0608] All terms are intended to be understood as understood by one of ordinary skill in the art. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

[0572]

[0609] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0573]

[0610] While various features of the present disclosure may be described in the context of a single embodiment, the features may also be provided individually or in any suitable combination. Conversely, although the present disclosure may be described herein for clarity in the context of separate embodiments, the present disclosure may also be implemented in a single embodiment.

[0574]

[0611] The following definitions supplement those in the art and are directed to the present application and are not to be construed as a limitation on, for example, commonly owned patents or applications, whether related or unrelated. Although any methods and materials similar or equivalent to those described herein can be used to carry out the testing of the present disclosure, the preferred materials and methods are described herein. Therefore, the terminology used herein is merely for the purpose of describing particular embodiments and is not intended to be limiting.

[0575] [I. Definition]

[0612] The terms used herein are for the purpose of describing particular cases only and are not intended to be limiting. As used herein, the singular forms "a," "an," and "the" are intended to include the plural forms as well, unless the context clearly dictates otherwise. Furthermore, when the terms "including," "comprises," "has," "having," "comprises," or variations thereof are used in either the detailed description and / or claims, such terms are intended to be inclusive in the same manner as the term "comprises."

[0576]

[0613] Terms such as "comprise," "included," "including," and the like are understood to have the meaning ascribed to them in U.S. patent law, i.e., they mean "include," "included," "comprising," and the like, and are intended to be inclusive or open-ended and do not exclude additional, unrecited elements or method steps; and terms such as "consisting essentially of" and "consisting essentially of" have the meaning ascribed to them in U.S. patent law, i.e., they allow for elements not expressly recited, but exclude elements found in the prior art or which affect the basic or novel characteristics of the invention.

[0577]

[0614] The term "and / or" as used herein in phrases such as "A and / or B" is intended to include both A and B; A or B; A alone; and B alone. Similarly, the term "and / or" as used in phrases such as "A, B and / or C" is intended to encompass each of the following embodiments: A, B and C; A, B or C; A or B; A or C; B or C; A and B; A and C; B and C; A alone; B alone; and C alone.

[0578]

[0615] When ranges are used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formula, all combinations and subcombinations of ranges and specific embodiments therein are intended to be included. The term "about" when referring to a numerical value or numerical range means that the referenced numerical value or numerical range is approximate within experimental variation (or within statistical experimental error); thus, in some cases, the numerical value or numerical range varies by 1% to 15% of the stated numerical value or numerical range. In some embodiments, the term "about" means within 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, or 0.05% of the given value or range.

[0579]

[0616] As used herein, the term "administering" includes providing a synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition described herein to an animal so that the animal can ingest the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition. In such embodiments, the animal ingests a portion of the synthetic oligosaccharide preparation, nutritional composition, or animal feed composition. In some embodiments, the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition is provided to the animal so that the animal can ingest the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition ad libitum. In some embodiments, the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition is administered to the animal as a formulated diet. In some embodiments, the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition is administered to the animal via manual feeding, e.g., oral syringe feeding, tube feeding, etc. In some embodiments, the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition is administered to the animal orally, e.g., ad libitum or manually. In some embodiments, animals receive a portion of the synthetic oligosaccharide preparation, nutritional composition, liquid, or animal feed composition every 24 hours or every other day for at least 7, 14, 21, 30, 45, 60, 75, 90, or 120 days. In some embodiments, the oligosaccharide preparation can be dissolved in water or another liquid, and the animal receives a portion of the oligosaccharide preparation by drinking the liquid. In some embodiments, the oligosaccharides are provided to the animal via drinking water. In some embodiments, the oligosaccharide preparation, nutritional composition, liquid, or animal feed composition is consumed ad libitum.

[0580]

[0617] As used herein, the term feed conversion ratio (FCR) refers to the ratio of feed input (e.g., ingested by an animal) to livestock output, where livestock output is the livestock product of interest. For example, livestock output for dairy animals is milk, while livestock output for animals raised for meat is body mass.

[0581]

[0618] As used herein, "feed efficiency" refers to the ratio of feed input (e.g., ingested by an animal) to animal output, where animal output is the animal product of interest.

[0582]

[0619] As used herein, the term "anhydro subunit" refers to a monosaccharide (or monosaccharide subunit) or a thermal dehydration product of a sugar caramelization product. For example, an "anhydro subunit" can be an anhydro-monosaccharide, such as anhydro-glucose. As another example, an "anhydro subunit" can be linked to one or more regular or anhydro-monosaccharide subunits via glycosidic bonds.

[0583]

[0620] The term "oligosaccharide" refers to a monosaccharide or compound containing two or more monosaccharide subunits linked by glycosidic bonds. As such, oligosaccharides include regular monosaccharides; anhydro-monosaccharides; or compounds containing two or more monosaccharide subunits, where one or more monosaccharide subunits are optionally independently replaced with one or more anhydro-subunits. Oligosaccharides can be functionalized. As used herein, the term oligosaccharide encompasses all species of oligosaccharides, where each of the monosaccharide subunits in an oligosaccharide is independently and optionally functionalized and / or replaced with its corresponding anhydro-monosaccharide subunit.

[0584]

[0621] As used herein, the term "oligosaccharide preparation" refers to a preparation that includes at least one oligosaccharide.

[0585]

[0622] As used herein, the term "gluco-oligosaccharide" refers to a compound containing glucose or two or more glucose monosaccharide subunits linked by glycosidic bonds. As such, gluco-oligosaccharides include compounds containing glucose; anhydro-glucose; or two or more glucose monosaccharide subunits linked by glycosidic bonds, wherein one or more of the glucose monosaccharide subunits are each optionally and independently replaced with an anhydro-glucose subunit.

[0586]

[0623] As used herein, the term "galacto-oligosaccharide" refers to galactose or a compound containing two or more galactose monosaccharide subunits linked by glycosidic bonds. As such, galacto-oligosaccharides include galactose; anhydro-galactose; or a compound containing two or more galactose monosaccharide subunits linked by glycosidic bonds, where at least one monosaccharide subunit is optionally replaced with an anhydro-galactose subunit.

[0587]

[0624] As used herein, the term "gluco-galactose-oligosaccharide preparation" refers to a composition produced from a complete or incomplete glycocondensation reaction of glucose and galactose. Thus, in some embodiments, gluco-galactose-oligosaccharide preparations include gluco-oligosaccharides, galacto-oligosaccharides, compounds containing one or more glucose monosaccharide subunits and one or more galactose monosaccharide subunits linked by glycosidic bonds, or combinations thereof. In some embodiments, gluco-galactose-oligosaccharide preparations include gluco-oligosaccharides and compounds containing one or more glucose monosaccharide subunits and one or more galactose monosaccharide subunits linked by glycosidic bonds. In some embodiments, gluco-galactose-oligosaccharide preparations include galacto-oligosaccharides and compounds containing one or more glucose monosaccharide subunits and one or more galactose monosaccharide subunits linked by glycosidic bonds. In some embodiments, the gluco-galactose-oligosaccharide preparation comprises a compound containing one or more glucose monosaccharide subunits and one or more galactose monosaccharide subunits linked by a glycosidic bond.

[0588]

[0625] As used herein, the terms "monosaccharide unit" and "monosaccharide subunit" are used interchangeably. A "monosaccharide subunit" refers to a monosaccharide monomer of an oligosaccharide. For oligosaccharides having a degree of polymerization of 1, the oligosaccharide may be referred to as a monosaccharide subunit or a monosaccharide. For oligosaccharides having a degree of polymerization of 2 or higher, the monosaccharide subunits are linked via glycosidic bonds.

[0589]

[0626] As used herein, the term "normal monosaccharide" refers to a monosaccharide that does not contain an anhydro subunit. The term "normal disaccharide" refers to a disaccharide that does not contain an anhydro subunit. Thus, the term "normal subunit" refers to a subunit that is not an anhydro subunit.

[0590]

[0627] As used herein, the terms "anhydroDPn oligosaccharide," "anhydroDPn species," or "DPn anhydrosubunit-containing oligosaccharide" refer to an oligosaccharide having a degree of polymerization n and containing one or more anhydrosubunits. As such, anhydroglucose is a DP1 anhydrosubunit-containing oligosaccharide, and cellotriosan is a DP3 anhydrosubunit-containing oligosaccharide.

[0591]

[0628] The term "relative abundance" or "abundance," as used herein, refers to the abundance of a species based on how common or rare the species is. For example, a DP1 fraction containing 10% anhydrosubunit-containing oligosaccharides at relative abundance may refer to a plurality of DP1 oligosaccharides, where 10% of the DP1 oligosaccharides are anhydromonosaccharides. For example, for a particular DP fraction of oligosaccharides, the relative abundance can be determined by suitable analytical equipment, e.g., mass spectrometry and liquid chromatography, such as LC-MS / MS, GC-MS, HPLC-MS, and MALDI-MS. In some embodiments, the relative abundance is determined by integrating the area under the peak in a chromatograph (e.g., LC-MS / MS, GC-MS, and HPLC-MS) corresponding to the fraction of interest. In some embodiments, the relative abundance is determined by peak intensity (e.g., MALDI-MS). In some embodiments, the relative abundance is determined by a combination of analytical methods, such as gravimetric determination after liquid chromatographic separation.

[0592]

[0629] As used herein, the singular forms "a," "and," and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, reference to "an agent" includes a plurality of such agents, and reference to "an oligosaccharide" includes one or more oligosaccharides (or oligosaccharides) and equivalents thereof known to those skilled in the art.

[0593] II. Oligosaccharide Preparations

[0630] Disclosed herein are oligosaccharide preparations suitable for use in nutritional compositions. In some embodiments, the oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than or equal to 2. In some embodiments, n is an integer greater than or equal to 3. In some embodiments, each of the fractions 1 to n in the oligosaccharide preparation comprises 1% to 90% anhydrosubunit-containing oligosaccharides in relative abundance as measured by mass spectrometry. In some embodiments, the relative abundance of oligosaccharides in each fraction decreases monotonically with their degree of polymerization.

[0594]

[0631] In some embodiments, n is an integer greater than or equal to 3. In some embodiments, n is an integer in the range of 1 to 100, such as 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, or 50. In some embodiments, each of the fractions 1 to n in the oligosaccharide preparation independently comprises 0.1% to 90% anhydro-subunit-containing oligosaccharides by relative abundance as measured by mass spectrometry, LC-MS / MS, or GC-MS. In some embodiments, each of the fractions 1 to n in the oligosaccharide preparation independently comprises about 0.1% to about 15% anhydro-subunit-containing oligosaccharides. In some embodiments, each of fractions 1 through n in an oligosaccharide preparation independently comprises about 0.5% to about 15% anhydrosubunit-containing oligosaccharides. In some embodiments, the DP1 and DP2 fractions independently comprise about 0.1% to about 15% anhydrosubunit-containing oligosaccharides by relative abundance as measured by mass spectrometry such as MALDI-MS, LC-MS / MS, or GC-MS. In some embodiments, the DP1 and DP2 fractions independently comprise about 0.5% to about 15% anhydrosubunit-containing oligosaccharides. In some embodiments, the DP1 and DP2 fractions each independently comprise anhydrosubunit-containing oligosaccharides at a relative abundance of about 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.8%, 1%, 2%, or 3% to about 8%, 9%, 10%, 11%, 12%, 13%, 14%, or 15% as measured by mass spectrometry, LC-MS / MS, or GC-MS. In some embodiments, the relative abundance of oligosaccharides in each fraction decreases monotonically with their degree of polymerization.

[0595]

[0632] In some embodiments, the oligosaccharide preparation is a synthetic oligosaccharide preparation. In some embodiments, a synthetic oligosaccharide preparation refers to a plurality of oligosaccharides produced by a process that does not require a living organism. In some embodiments, a synthetic oligosaccharide preparation refers to a plurality of oligosaccharides produced by a process that does not require an enzyme. In some embodiments, a synthetic oligosaccharide preparation refers to a plurality of oligosaccharides produced by a chemical process. In certain embodiments, a synthetic oligosaccharide preparation refers to a plurality of oligosaccharides produced by condensation of sugars.

[0596] [A. Animal Microbiome Metabolic Regulators]

[0633] Disclosed herein are microbiome metabolic regulators comprising oligosaccharide preparations containing anhydro-sugar components and / or sugar dehydration product components that exhibit complex functional regulation of microbial communities, such as the gut microbiome of an animal. The oligosaccharide preparations provide utility in regulating, modifying, or controlling fermentable carbon utilization by microflora and directing metabolic flux toward beneficial species, thereby providing microbiome-mediated health or nutritional benefits.

[0597]

[0634] Non-digestible carbohydrates can function as prebiotics by providing microbial communities with a fermentable carbon source. For example, diets rich in soluble plant fiber have been shown to have the ability to nourish the intestinal microflora. Furthermore, bifidobacterial prebiotics support the growth of bifidobacteria (e.g., members of the Bifidobacterium genus), while lactic acid bacteria prebiotics support the growth of Lactobacillus species.

[0598]

[0635] Prebiotic fibers can be fermented into beneficial chemical species such as short-chain fatty acids (SCFAs). Prebiotic fibers include resistant starch; cellulose; pectins such as rhamnogalactan, arabinogalactan, and arabinan; hemicelluloses such as arabinoxylan, xyloglucan, glucomannan, and galactomannan; xylans such as corncob oligosaccharides; β-glucans such as cereal β-glucan, yeast β-glucan, and bacterial β-glucan; polyfructans such as inulin and levan; and gums such as alginates. Inulin is a common bifidobacterial prebiotic fiber.

[0599]

[0636] In other cases, prebiotics act by interfering with the ability of pathogenic bacteria to colonize and thus infect the host organism through antiadhesion mechanisms such as competitive binding of cell surface receptor sites. Certain galacto-oligosaccharides provide effective antiadhesion for various enteropathogenic organisms, such as Escherichia species.

[0600]

[0637] Prebiotics are typically provided to host animals by incorporation into their diets, where they exhibit a dose-dependent response (at least up to a saturation threshold). For example, providing high doses of bifidobacterial prebiotics, such as inulin, tends to significantly increase the population of Bifidobacterium species. High doses of inulin correspond to higher production of SCFAs through fermentation. This is because the prebiotic provides a metabolic carbon source, and more carbon is converted into more fermentation products. Similarly, providing higher doses of anti-adhesion prebiotics offers the potential for competitive binding to surface receptor sites.

[0601]

[0638] Certain carbohydrate species containing modified monomeric subunits may affect how microbial systems utilize other carbohydrates that are otherwise available as prebiotic sources. For example, such carbohydrate species may be modified carbohydrate species that regulate the bacterial starch utilization system (SUS), i.e., proteins involved in cell surface recognition, glycosidic cleavage, and import of starch metabolites.

[0602]

[0639] Carbohydrate compositions capable of complexly modulating animal microbiota have utility as feed additives for improving animal health and nutrition through their effects on the animal's microbiome. For example, modulation of butyrate production by intestinal microflora promotes healthy intestinal mucosa, barrier function, and provides health benefits to animals through anti-inflammatory effects. Modulation of propionate production influences the metabolic energy extracted from an animal's diet through increased host gluconeogenesis. Related microbial communities include, for example, the ileum, jejunum, and cecum and / or fecal microbiota of poultry, pigs, dogs, cats, and horses, or the microbiota of ruminant animals such as cattle, cows, and sheep.

[0603]

[0640] Furthermore, the oligosaccharide preparations disclosed herein are advantageous in that they can be selectively analyzed and quantified in complex nutritional compositions, such as complete animal feeds, due to the presence of anhydrous subunits. It is commercially useful to assess the presence and / or concentration of feed additives, such as oligosaccharide preparations. Such assessments can be performed for quality control purposes to determine whether the additive can be uniformly blended with a base nutritional composition to provide a final nutritional composition containing the additive at the intended dosage or inclusion level.

[0604]

[0641] However, nutritional compositions themselves contain a large amount and variety of carbohydrate structures (e.g., starch, vegetable fiber, and pectin). Therefore, it is particularly difficult to distinguish small amounts of oligosaccharide-based feed additives from the vast ocean of other carbohydrates present as the basis of nutritional compositions. As such, the oligosaccharide preparations disclosed herein provide a means to distinguish themselves from other carbohydrate sources in nutritional compositions via their anhydrous subunits.

[0605] [B. Degree of polymerization (DP) distribution]

[0642] In some embodiments, the oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1-DPn fractions), each having a different degree of polymerization selected from 1 to n. In some embodiments, the oligosaccharide preparation comprises n fractions of oligosaccharides (DP1-DPn fractions), each having a different degree of polymerization selected from 1 to n. For example, in some embodiments, the DP1 fraction comprises one or more monosaccharides and / or one or more anhydro-monosaccharides. As another example, in some embodiments, the DP1 fraction comprises glucose, galactose, fructose, 1,6-anhydro-β-D-glucofuranose, 1,6-anhydro-β-D-glucopyranose, or any combination thereof. As yet another example, in some embodiments, the DP2 fraction comprises one or more normal disaccharides and one or more anhydro-subunit-containing disaccharides. In some embodiments, the DP2 fraction comprises lactose.

[0606]

[0643] In some embodiments, n is at least 2, at least 3, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39, at least 40, at least 41, at least 42, at least 43, at least 44, at least 45, at least 46, at least 47, at least 48, at least 49, at least 50, at least 51 , at least 52, at least 53, at least 54, at least 55, at least 56, at least 57, at least 58, at least 59, at least 60, at least 61, at least 62, at least 63, at least 64, at least 65, at least 66, at least 67, at least 68, at least 69, at least 70, at least 71, at least 72, at least 73, at least 74, at least 75, at least 76, at least 77, at least 78, at least 79, at least 80, at least 81, at least 82, at least 83, at least 84, at least 85, at least 86, at least 87, at least 88, at least 89, at least 90, at least 91, at least 92, at least 93, at least 94, at least 95, at least 96, at least 97, at least 98, at least 99, or at least 100.In some embodiments, n is 2, 3, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51 , 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100. In some embodiments, n is less than 10, less than 11, less than 12, less than 13, less than 14, less than 15, less than 16, less than 17, less than 18, less than 19, less than 20, less than 21, less than 22, less than 23, less than 24, less than 25, less than 26, less than 27, less than 28, less than 29, less than 30, less than 31, less than 32, less than 33, less than 34, less than 35, less than 36, less than 37, less than 38, less than 39, less than 40, less than 41, less than 42, less than 43, less than 44, less than 45, less than 46, less than 47, less than 48, less than 49, less than 50, less than 51, less than 52, less than 53, less than 54 full, less than 55, less than 56, less than 57, less than 58, less than 59, less than 60, less than 61, less than 62, less than 63, less than 64, less than 65, less than 66, less than 67, less than 68, less than 69, less than 70, less than 71, less than 72, less than 73, less than 74, less than 75, less than 76, less than 77, less than 78, less than 79, less than 80, less than 81, less than 82, less than 83, less than 84, less than 85, less than 86, less than 87, less than 88, less than 89, less than 90, less than 91, less than 92, less than 93, less than 94, less than 95, less than 96, less than 97, less than 98, less than 99, or less than 100. In some embodiments, n is 2 to 100, 5 to 90, 10 to 90, 10 to 80, 10 to 70, 10 to 60, 10 to 50, 10 to 40, 10 to 30, 15 to 60, 15 to 50, 15 to 45, 15 to 40, 15 to 35, or 15 to 30.

[0607]

[0644] The degree of polymerization distribution of an oligosaccharide preparation can be determined by any suitable analytical method and instrument, including, but not limited to, end-group analysis, osmotic pressure (osmometry), ultracentrifugation, viscometry, light scattering, size-exclusion chromatography (SEC), SEC-MALLS, field-flow fractionation (FFF), asymmetric flow field-flow fractionation (A4F), high-performance liquid chromatography (HPLC), and mass spectrometry (MS). For example, the degree of polymerization distribution can be determined and / or detected by mass spectrometry, such as matrix-assisted laser desorption / ionization (MALDI)-MS, liquid chromatography (LC)-MS, or gas chromatography (GC)-MS. As another example, the degree of polymerization distribution can be determined and / or detected by SEC, such as gel permeation chromatography (GPC). As yet another example, the degree of polymerization distribution can be determined and / or detected by HPLC, FFF, or A4F. In some embodiments, the degree of polymerization distribution is determined and / or detected by MALDI-MS. In some embodiments, the degree of polymerization distribution is determined and / or detected by GC-MS or LC-MS. In some embodiments, the degree of polymerization distribution is determined and / or detected by SEC. In some embodiments, the degree of polymerization distribution is determined and / or detected by HPLC. In some embodiments, the degree of polymerization distribution is determined and / or detected by a combination of analytical instruments, such as MALDI-MS and SEC. In some embodiments, the degree of polymerization of an oligosaccharide preparation can be determined based on its molecular weight and molecular weight distribution. For example, Figure 2 shows a MALDI-MS spectrum showing the degree of polymerization of various fractions and the presence of anhydrosubunit-containing oligosaccharides (-18 g / mol MW offset peak) in all observed fractions.

[0608]

[0645] In some embodiments, the relative abundance of oligosaccharides in the majority of the fractions decreases monotonically with their degree of polymerization, hi some embodiments, the relative abundance of oligosaccharides in less than 6, less than 5, less than 4, less than 3, or less than 2 fractions of the oligosaccharide preparation does not decrease monotonically with their degree of polymerization.

[0609]

[0646] In some embodiments, the relative abundance of oligosaccharides in at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 DP fractions decreases monotonically with their degree of polymerization. In some embodiments, the relative abundance of oligosaccharides in at least 5, at least 10, at least 15, at least 20, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive DP fractions decreases monotonically with their degree of polymerization. In some embodiments, the relative abundance of oligosaccharides in at least 5, at least 10, at least 20, or at least 30 consecutive DP fractions decreases monotonically with their degree of polymerization. In some embodiments, the relative abundance of oligosaccharides in at least 5, at least 10, at least 20, or at least 30 consecutive DP fractions decreases monotonically with their degree of polymerization.

[0610]

[0647] In some embodiments, the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with their degree of polymerization. For example, Figure 15 provides an example of a DP distribution in which the relative abundance of oligosaccharides in each of the n fractions decreases monotonically with their DP. For example, in some embodiments, only the relative abundance of oligosaccharides in the DP3 fraction does not decrease monotonically with their degree of polymerization, i.e., the relative abundance of oligosaccharides in the DP3 fraction is lower than the relative abundance of oligosaccharides in the DP4 fraction. In some embodiments, the relative abundance of oligosaccharides in the DP2 fraction is lower than the relative abundance of oligosaccharides in the DP3 fraction. For example, Figure 16 shows a degree of polymerization distribution in which the relative abundance of oligosaccharides in the DP2 fraction does not decrease monotonically with their degree of polymerization.

[0611]

[0648] In some embodiments, the oligosaccharide preparations described herein have a DP1 fraction content by weight or relative abundance of about 1% to about 50%, about 1% to about 40%, about 1% to about 35%, about 1% to about 30%, about 1% to about 25%, about 1% to about 20%, about 1% to about 15%, about 5% to about 50%, about 5% to about 40%, about 5% to about 35%, about 5% to about 30%, about 5% to about 25%, about 5% to about 20%, about 5% to about 15%, about 10% to about 50%, about 10% to about 40%, about 10% to about 35%, about 10% to about 30%, about 10% to about 25%, about 10% to about 20%, or about 10% to about 15%. In some embodiments, the oligosaccharide preparation has a DP1 fraction content of about 10% to about 35%, about 10% to about 20%, or about 10% to about 15% by weight or relative abundance. In some embodiments, the DP1 fraction content is determined by MALDI-MS. In some embodiments, the DP1 fraction content is determined by HPLC. In some embodiments, the DP1 fraction content is determined by LC-MS / MS or GC-MS.

[0612]

[0649] In some embodiments, the oligosaccharide preparations described herein have a DP2 fraction content of about 1% to about 35%, about 1% to about 30%, about 1% to about 25%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, about 5% to about 30%, about 5% to about 25%, about 5% to about 20%, about 5% to about 15%, or about 5% to about 10%, by weight or relative abundance. In some embodiments, the oligosaccharide preparations have a DP2 fraction content of about 5% to about 25%, about 5% to about 20%, about 5% to about 15%, or about 5% to about 10%, by weight or relative abundance. In some embodiments, the DP2 fraction content is determined by MALDI-MS. In some embodiments, the DP2 fraction content is determined by HPLC. In some embodiments, the DP2 fraction content is determined by LC-MS / MS or GC-MS.

[0613]

[0650] In some embodiments, the oligosaccharide preparations described herein have a DP3 fraction content of about 1% to about 30%, about 1% to about 25%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, about 5% to about 30%, about 5% to about 25%, about 5% to about 20%, about 5% to about 15%, or about 5% to about 10%, by weight or relative abundance. In some embodiments, the oligosaccharide preparations have a DP3 fraction content of about 1% to about 15%, about 1% to about 10%, about 5% to about 15%, or about 5% to about 10%, by weight or relative abundance. In some embodiments, the DP3 fraction content is determined by MALDI-MS. In some embodiments, the DP3 fraction content is determined by HPLC. In some embodiments, the DP3 fraction content is determined by LC-MS / MS or GC-MS.

[0614]

[0651] In some embodiments, the oligosaccharide preparations described herein have a DP4 fraction content of about 0.1% to about 20%, about 0.1% to about 15%, about 0.1% to about 10%, about 0.1% to about 5%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, or about 1% to about 5%, by weight or relative abundance. In some embodiments, the oligosaccharide preparations described herein have a DP4 fraction content of about 1% to about 15%, about 1% to about 10%, or about 1% to about 5%, by weight or relative abundance. In some embodiments, the oligosaccharide preparations described herein have a DP5 fraction content of about 0.1% to about 15%, about 0.1% to about 10%, about 0.1% to about 5%, about 1% to about 15%, about 1% to about 10%, or about 1% to about 5%, by weight or relative abundance. In some embodiments, the oligosaccharide preparation has a DP5 fraction content of about 1% to about 10% or about 1% to about 5% by weight or relative abundance. In some embodiments, the DP4 and / or DP5 fraction content is determined by MALDI-MS. In some embodiments, the DP4 and / or DP5 fraction content is determined by HPLC. In some embodiments, the DP4 and / or DP5 fraction content is determined by LC-MS / MS or GC-MS.

[0615]

[0652] In some embodiments, the ratio of the DP2 fraction to the DP1 fraction in the oligosaccharide preparation, expressed in terms of weight or relative abundance, is about 0.01 to about 0.8, about 0.02 to about 0.7, about 0.02 to about 0.6, about 0.02 to about 0.5, about 0.02 to about 0.4, about 0.02 to about 0.3, about 0.02 to about 0.2, about 0.1 to about 0.6, about 0.1 to about 0.5, about 0.1 to about 0.4, or about 0.1 to about 0.3. In some embodiments, the ratio of the DP2 fraction to the DP1 fraction in the oligosaccharide preparation, expressed in terms of weight or relative abundance, is about 0.02 to about 0.4.

[0616]

[0653] In some embodiments, the ratio of the DP3 fraction to the DP2 fraction in the oligosaccharide preparation, expressed in terms of weight or relative abundance, is about 0.01 to about 0.7, about 0.01 to about 0.6, about 0.01 to about 0.5, about 0.01 to about 0.4, about 0.01 to about 0.3, or about 0.01 to about 0.2. In some embodiments, the ratio of the DP3 fraction to the DP2 fraction in the oligosaccharide preparation, expressed in terms of weight or relative abundance, is about 0.01 to about 0.3.

[0617]

[0654] In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, or less than 10% by weight or relative abundance. In some embodiments, the aggregate content of the DP1 and DP2 fractions in the oligosaccharide preparation is less than 50%, less than 30%, or less than 10% by weight or relative abundance.

[0618]

[0655] In some embodiments, the oligosaccharide preparations described herein have an average DP value ranging from 2 to 10. In some embodiments, the oligosaccharide preparations have an average DP value of about 2 to about 8, about 2 to about 5, or about 2 to about 4. In some embodiments, the oligosaccharide preparations have an average DP value of about 3.5. The average DP value can be determined by SEC or elemental analysis.

[0619] [C. Anhydro Subunit Concentration]

[0656] In some embodiments, each of the n fractions of oligosaccharides independently comprises an anhydrosubunit concentration. For example, in some embodiments, the DP1 fraction comprises 10% relative abundance of anhydrosubunit-containing oligosaccharides, and the DP2 fraction comprises 15% relative abundance of anhydrosubunit-containing oligosaccharides. For another example, in some embodiments, the DP1, DP2, and DP3 fractions each comprise 5%, 10%, and 2% relative abundance of anhydrosubunit-containing oligosaccharides, respectively. In other embodiments, two or more fractions of oligosaccharides may comprise the same concentration of anhydrosubunit-containing oligosaccharides. For example, in some embodiments, the DP1 and DP3 fractions each comprise about 5% relative abundance of anhydrosubunit-containing oligosaccharides.

[0620]

[0657] In some embodiments, each of the 1-n fractions in an oligosaccharide preparation described herein independently comprises about 0.1% to 15% anhydrosubunit-containing oligosaccharides in relative abundance as measured by mass spectrometry, LC-MS / MS, or GC-MS. In some embodiments, each of the 1-n fractions in an oligosaccharide preparation independently comprises about 0.5% to 15% anhydrosubunit-containing oligosaccharides in relative abundance as measured by mass spectrometry, LC-MS / MS, or GC-MS. In some embodiments, LC-MS / MS is used to determine the relative abundance of oligosaccharides in the DP1, DP2, and / or DP3 fractions. In some embodiments, GC-MS is used to determine the relative abundance of oligosaccharides in the DP1, DP2, and / or DP3 fractions. In some embodiments, MALDI-MS is used to determine the relative abundance of oligosaccharides in the DP4 fraction and higher DP fractions. In some embodiments, the relative abundance of a particular fraction is determined by integrating the area under the peak in an LC-MS / MS chromatogram designated as corresponding to that fraction. In some embodiments, the relative abundance of a particular fraction is determined by integrating the area under the peak in a GC-MS chromatogram designated as corresponding to that fraction.

[0621]

[0658] The concentration of anhydro subunits can be determined by any suitable analytical method, such as nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry, HPLC, FFF, A4F, or any combination thereof. In some embodiments, the concentration of anhydro subunits is determined, at least in part, by mass spectrometry, such as MALDI-MS. In some embodiments, the concentration of anhydro subunits is determined, at least in part, by NMR. In some embodiments, the concentration of anhydro subunit-containing oligosaccharides is determined, at least in part, by HPLC. In some embodiments, the concentration of anhydro subunit-containing oligosaccharides is determined, at least in part, by MALDI-MS, as shown by the -18 g / mol MW offset peak in Figure 2. In some embodiments, the presence and type of anhydro subunit species can be determined and / or detected by NMR, as shown in Example 11, Figures 3, and 4. In some embodiments, the relative abundance of anhydro subunit-containing oligosaccharides is determined by MALDI-MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by LC-MS / MS, as shown in Figures 24A-24C, 25A-25C, 26A-26C, and 27A-27C. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by GC-MS, as shown in Figures 28A-28B, 29A-29B, 30A-30B, and 31A-31B.

[0622]

[0659] In some embodiments, at least one fraction of an oligosaccharide preparation described herein comprises anhydro-subunit-containing oligosaccharides at a relative abundance of less than 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%. In some embodiments, at least one fraction of an oligosaccharide preparation described herein comprises anhydro-subunit-containing oligosaccharides at a relative abundance of less than 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, or 2%. In other embodiments, at least one fraction of the oligosaccharide preparations described herein comprises, by relative abundance, greater than 0.5%, greater than 0.8%, greater than 1%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, greater than 12%, greater than 13%, greater than 14%, greater than 15%, greater than 16%, greater than 17%, greater than 18%, greater than 19%, greater than 20%, greater than 30%, greater than 40%, greater than 50%, greater than 60%, greater than 70%, or greater than 80% anhydro-subunit-containing oligosaccharides. In other embodiments, at least one fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 20%, greater than 21%, greater than 22%, greater than 23%, greater than 24%, greater than 25%, greater than 26%, greater than 27%, greater than 28%, greater than 29%, or greater than 30%. In some embodiments, at least one fraction (such as DP1, DP2, and / or DP3) of the oligosaccharide preparation comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, or about 30%.In some embodiments, at least one fraction (such as DP1, DP2, and / or DP3) of the oligosaccharide preparation comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 1.5%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10%. In some embodiments, at least one fraction of the oligosaccharide preparation (such as DP1, DP2, and / or DP3) has a relative abundance of about 0.1% to about 90%, about 0.5% to about 90%, about 0.5% to about 80%, about 0.5% to about 70%, about 0.5% to about 60%, about 0.5% to about 50%, about 0.5% to about 40%, about 0.5% to about 30%, about 0.5% to about 20%, about 0.5% to about 10%, about 0.5% to about 10%. The anhydrosubunit-containing oligosaccharides are 5% to about 9%, about 0.5% to about 8%, about 0.5% to about 7%, about 0.5% to about 6%, about 0.5% to about 5%, about 0.5% to about 4%, about 0.5% to about 3%, about 0.5% to about 2%, about 1% to about 10%, about 2% to about 9%, about 2% to about 8%, about 2% to about 7%, about 2% to about 6%, about 2% to about 5%, about 2% to about 4%, about 2% to about 3%, or about 5% to about 10%. In some embodiments, the DP1 and DP2 fractions of the oligosaccharide preparation each independently comprise anhydro-subunit-containing oligosaccharides in a relative abundance ranging from about 0.1%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 1.1%, 1.2%, 1.3%, 1.4%, or 1.5% to about 8%, 9%, 10%, 11%, 12%, or 15% as measured by mass spectrometry, LC-MS / MS, or GC-MS. In some embodiments, the DP1 and DP2 fractions each independently comprise anhydro-subunit-containing oligosaccharides in a relative abundance ranging from about 0.5% to about 15% as measured by mass spectrometry, LC-MS / MS, or GC-MS.

[0623]

[0660] In some embodiments, each fraction of the oligosaccharide preparations described herein comprises less than 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, or 2% of anhydrosubunit-containing oligosaccharides by relative abundance. In some embodiments, each fraction of the oligosaccharide preparations described herein comprises less than 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, or 2% of anhydrosubunit-containing oligosaccharides by relative abundance. In other embodiments, each fraction of the oligosaccharide preparations described herein comprises greater than 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, or 80% anhydrosubunit-containing oligosaccharides by relative abundance. In other embodiments, each fraction of the oligosaccharide preparations described herein comprises greater than 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, or 30% anhydrosubunit-containing oligosaccharides by relative abundance. In some embodiments, each fraction of the oligosaccharide preparations described herein comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, or about 30%. In some embodiments, each fraction of the oligosaccharide preparations described herein comprises about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% anhydrosubunit-containing oligosaccharides in relative abundance.In some embodiments, each fraction of the oligosaccharide preparations described herein has a relative abundance of about 0.1% to about 90%, about 0.1% to about 15%, about 0.5% to about 90%, about 0.5% to about 80%, about 0.5% to about 70%, about 0.5% to about 60%, about 0.5% to about 50%, about 0.5% to about 40%, about 0.5% to about 30%, about 0.5% to about 20%, about 0.5% to about 10%, about 0.5% to about 10%, about 0.5% to about 15 ... The anhydrosubunit-containing oligosaccharides are 5% to about 9%, about 0.5% to about 8%, about 0.5% to about 7%, about 0.5% to about 6%, about 0.5% to about 5%, about 0.5% to about 4%, about 0.5% to about 3%, about 0.5% to about 2%, about 2% to about 9%, about 2% to about 8%, about 2% to about 7%, about 2% to about 6%, about 2% to about 5%, about 2% to about 4%, about 2% to about 3%, or about 5% to about 10%.

[0624]

[0661] In some embodiments, the oligosaccharide preparations described herein comprise anhydro-subunit-containing oligosaccharides at a relative abundance of less than 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1%. In some embodiments, the oligosaccharide preparations comprise anhydro-subunit-containing oligosaccharides at a relative abundance of less than 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, or 2%. In other embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.5%, 0.8%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 30%, 40%, 50%, 60%, 70%, or 80%. In other embodiments, the oligosaccharide preparation comprises an anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, or 30%. In some embodiments, the oligosaccharide preparation comprises anhydro-subunit-containing oligosaccharides at a relative abundance of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, or about 30%. In some embodiments, the oligosaccharide preparation comprises an anhydrosubunit-containing oligosaccharides at a relative abundance of about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10%.In some embodiments, the oligosaccharide preparation comprises, in relative abundance, about 0.1% to about 90%, about 0.1% to about 15%, about 0.5% to about 90%, about 0.5% to about 80%, about 0.5% to about 70%, about 0.5% to about 60%, about 0.5% to about 50%, about 0.5% to about 40%, about 0.5% to about 30%, about 0.5% to about 20%, about 0.5% to about 10%, about 0.5% to about 9% , about 0.5% to about 8%, about 0.5% to about 7%, about 0.5% to about 6%, about 0.5% to about 5%, about 0.5% to about 4%, about 0.5% to about 3%, about 0.5% to about 2%, about 2% to about 9%, about 2% to about 8%, about 2% to about 7%, about 2% to about 6%, about 2% to about 5%, about 2% to about 4%, about 2% to about 3%, or about 5% to about 10% anhydrosubunit-containing oligosaccharides.

[0625]

[0662] In some embodiments, the DP1 fraction of the oligosaccharide preparations described herein comprises, by relative abundance, less than 30%, less than 20%, less than 19%, less than 18%, less than 17%, less than 16%, less than 15%, less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% anhydrosubunit-containing oligosaccharides. In some embodiments, the DP1 fraction of the oligosaccharide preparations described herein comprises anhydro-subunit-containing oligosaccharides at a relative abundance of greater than 0.1%, greater than 0.5%, greater than 0.8%, greater than 1%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, greater than 12%, greater than 13%, greater than 14%, or greater than 15%. In some embodiments, the DP1 fraction of the oligosaccharide preparations described herein comprises anhydrosubunit-containing oligosaccharides at a relative abundance of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, or about 20%. In some embodiments, the DP1 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.1% to about 15%, about 0.1% to about 20%, about 0.5% to about 20%, 0.5% to about 10%, about 0.5% to about 15%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, about 2% to about 14%, about 3% to about 13%, about 4% to about 12%, about 5% to about 11%, about 5% to about 10%, about 6% to about 9%, or about 7% to about 8%, or any range therebetween. In some embodiments, the DP1 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by mass spectrometry, such as MALDI-MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by LC-MS / MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by GC-MS.

[0626]

[0663] In some embodiments, the DP2 fraction of the oligosaccharide preparations described herein comprises, by relative abundance, less than 30%, less than 20%, less than 19%, less than 18%, less than 17%, less than 16%, less than 15%, less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% anhydrosubunit-containing oligosaccharides. In some embodiments, the DP2 fraction of the oligosaccharide preparations described herein comprises, by relative abundance, greater than 0.1%, greater than 0.5%, greater than 0.8%, greater than 1%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, greater than 12%, greater than 13%, greater than 14%, or greater than 15% anhydro-subunit-containing oligosaccharides. In some embodiments, the DP2 fraction of the oligosaccharide preparations described herein comprises anhydrosubunit-containing oligosaccharides at a relative abundance of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, or about 20%. In some embodiments, the DP2 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.1% to about 15%, about 0.1% to about 20%, about 0.5% to about 20%, 0.5% to about 10%, about 0.5% to about 15%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, about 2% to about 14%, about 3% to about 13%, about 4% to about 12%, about 5% to about 11%, about 0.5% to about 10%, about 6% to about 9%, or about 7% to about 8%, or any range therebetween. In some embodiments, the DP2 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 5% to about 10%. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by mass spectrometry, such as MALDI-MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by LC-MS / MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by GC-MS.

[0627]

[0664] In some embodiments, the DP3 fraction of the oligosaccharide preparations described herein comprises, by relative abundance, less than 30%, less than 20%, less than 19%, less than 18%, less than 17%, less than 16%, less than 15%, less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% anhydrosubunit-containing oligosaccharides. In some embodiments, the DP3 fraction of the oligosaccharide preparations described herein comprises, by relative abundance, greater than 0.1%, greater than 0.5%, greater than 0.8%, greater than 1%, greater than 1.5%, greater than 2%, greater than 3%, greater than 4%, greater than 5%, greater than 6%, greater than 7%, greater than 8%, greater than 9%, greater than 10%, greater than 11%, greater than 12%, greater than 13%, greater than 14%, or greater than 15% anhydro-subunit-containing oligosaccharides. In some embodiments, the DP3 fraction of the oligosaccharide preparations described herein comprises anhydrosubunit-containing oligosaccharides at a relative abundance of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, or about 20%. In some embodiments, the DP3 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.1% to about 15%, about 0.1% to about 20%, about 0.5% to about 20%, 0.5% to about 10%, about 0.5% to about 15%, about 1% to about 20%, about 1% to about 15%, about 1% to about 10%, about 2% to about 14%, about 3% to about 13%, about 4% to about 12%, about 5% to about 11%, about 5% to about 10%, about 6% to about 9%, or about 7% to about 8%, or any range therebetween. In some embodiments, the DP3 fraction of the oligosaccharide preparations described herein comprises an anhydro-subunit-containing oligosaccharides in a relative abundance of about 0.5% to about 10%. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by mass spectrometry, such as MALDI-MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by LC-MS / MS. In some embodiments, the relative abundance of anhydro-subunit-containing oligosaccharides is determined by GC-MS.

[0628]

[0665] In some embodiments, anhydrosubunit-containing oligosaccharides contain one or more anhydrosubunits. For example, DP1 anhydrosubunit-containing oligosaccharides contain one anhydrosubunit. In some embodiments, DPn anhydrosubunit-containing oligosaccharides can contain 1 to n anhydrosubunits. For example, in some embodiments, DP2 anhydrosubunit-containing oligosaccharides contain one or two anhydrosubunits. In some embodiments, each oligosaccharide in the oligosaccharide preparation independently contains zero, one, or two anhydrosubunits. In some embodiments, more than 99%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, or 30% of the anhydrosubunit-containing oligosaccharides have only one anhydrosubunit. In some embodiments, greater than 99%, 95%, 90%, 85%, or 80% of the anhydro-subunit-containing oligosaccharides have only one anhydro-subunit.

[0629]

[0666] In some embodiments, one or more oligosaccharides in the oligosaccharide preparation or each fraction of the oligosaccharide preparation each comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 anhydro subunits linked via glycosidic bonds, where the glycosidic bond linking each anhydro subunit is independently selected. In some embodiments, one or more oligosaccharides in the oligosaccharide preparation or each fraction of the oligosaccharide preparation each comprise 1, 2, or 3 anhydro subunits linked via glycosidic bonds, where the glycosidic bond linking each anhydro subunit is independently selected. In some embodiments, more than 50%, 60%, 70%, 80%, 90%, or 99% of the oligosaccharides in the oligosaccharide preparation or each fraction each comprise 1, 2, or 3 anhydro subunits linked via glycosidic bonds, where the glycosidic bond linking each anhydro subunit is independently selected. In some embodiments, one or more of the oligosaccharides in the oligosaccharide preparation or each fraction contain one anhydro subunit linked via a glycosidic bond, hi some embodiments, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, or more than 99% of the oligosaccharides in the oligosaccharide preparation or each fraction contain one anhydro subunit linked via a glycosidic bond.

[0630] [D. Anhydro Subunit Species]

[0667] In some embodiments, the oligosaccharide preparation comprises different species of anhydro subunits. In some embodiments, exemplary anhydro subunit-containing oligosaccharides are shown in Figure 33, Figure 21, and Figure 22. In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits that are the thermal dehydration products of monosaccharides, i.e., anhydro-monosaccharide subunits. In some embodiments, the oligosaccharide preparation comprises one or more anhydro subunits that are the reversible thermal dehydration products of monosaccharides.

[0631]

[0668] An anhydro-monosaccharide (or anhydro-monosaccharide subunit) should be understood to refer to the thermal dehydration product of one or more species of monosaccharide. For example, in some embodiments, an anhydro-glucose refers to 1,6-anhydro-β-D-glucopyranose (levoglucosan) or 1,6-anhydro-β-D-glucofuranose. In some embodiments, a plurality of anhydro-glucoses refers to a plurality of 1,6-anhydro-β-D-glucopyranoses (levoglucosan), a plurality of 1,6-anhydro-β-D-glucofuranoses, a plurality of other thermal dehydration products of glucose, or any combination thereof. Similarly, in some embodiments, a plurality of anhydro-galactoses refers to a plurality of any thermal dehydration products of galactose, or any combination thereof.

[0632]

[0669] In some embodiments, the oligosaccharide preparations described herein comprise one or more anhydroglucose, anhydrogalactose, anhydromannose, anhydroallose, anhydroaltrose, anhydrogulose, anhydroindose, anhydrotalose, anhydrofructose, anhydroribose, anhydroarabinose, anhydrorhamnose, anhydrolyxose, anhydroxylose, or any combination of these subunits. In some embodiments, the oligosaccharide preparations comprise one or more anhydroglucose, anhydrogalactose, anhydromannose, or anhydrofructose subunits. In some embodiments, the oligosaccharide preparations described herein include 1,6-anhydro-3-O-β-D-glucopyranosyl-β-D-glucopyranose, 1,6-anhydro-3-O-α-D-glucopyranosyl-β-D-glucopyranose, 1,6-anhydro-2-O-β-D-glucopyranosyl-β-D-glucopyranose, 1,6-anhydro-2-O-α-D-glucopyranosyl-β-D-glucopyranose, 1,6-anhydro- β-D-cellobiose (cellobiosan), 1,6-anhydro-β-D-cellotriose (cellotriosan), 1,6-anhydro-β-D-cellotetraose (cellotetraosan), 1,6-anhydro-β-D-cellopentaose (cellopentaosan), and 1,6-anhydro-β-D-maltose (maltosan).

[0633]

[0670] In some embodiments, the oligosaccharide preparation comprises one or more 1,6-anhydro-β-D-glucofuranose subunits. In some embodiments, the oligosaccharide preparation comprises one or more 1,6-anhydro-β-D-glucopyranose (levoglucosan) subunits. For example, Figure 33 shows two DP1 anhydrosubunit-containing oligosaccharides (levoglucosan and 1,6-anhydro-β-D-glucofuranose) and one DP2 anhydrosubunit-containing oligosaccharide (anhydro-cellobiose).

[0634]

[0671] The presence and level of anhydro subunit species can vary based on the feed sugar used to produce the oligosaccharides, for example, in some embodiments, gluco-oligosaccharides contain anhydro-glucose subunits, galacto-oligosaccharides contain anhydro-galactose subunits, and gluco-galacto-oligosaccharides contain anhydro-glucose and anhydro-galactose subunits.

[0635]

[0672] In some embodiments, the oligosaccharide preparation comprises both 1,6-anhydro-β-D-glucofuranose and 1,6-anhydro-β-D-glucopyranose anhydrous subunits. In some embodiments, at least 0.1%, 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 99% of the anhydrous subunits are selected from the group consisting of 1,6-anhydro-β-D-glucofuranose and 1,6-anhydro-β-D-glucopyranose. In some embodiments, at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% of the anhydrous subunits are 1,6-anhydro-β-D-glucofuranose. In some embodiments, at least 1%, 5%, 10%, 20%, 30%, 40%, 50%, or 60% of the anhydro subunits are 1,6-anhydro-β-D-glucopyranose.

[0636]

[0673] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the preparation is about 10:1 to 1:10, 9:1 to 1:10, 8:1 to 1:10, 7:1 to 1:10, 6:1 to 1:10, 5:1 to 1:10, 4:1 to 1:10, 3:1 to 1:10, 2:1 to 1:10, 10:1 to 1:9, 10:1 to 1:8, 10:1 to 1:7, 10:1 to 1:6, 10:1 to 1:5, 10:1 to 1:4, 10:1 to 1:3, 10:1 to 1:2, or 1:1 to 3:1. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the preparation is about 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:8, 1:9, or 1:10. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in the preparation is about 2:1.

[0637]

[0674] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in each fraction is about 10:1 to 1:10, 9:1 to 1:10, 8:1 to 1:10, 7:1 to 1:10, 6:1 to 1:10, 5:1 to 1:10, 4:1 to 1:10, 3:1 to 1:10, 2:1 to 1:10, 10:1 to 1:9, 10:1 to 1:8, 10:1 to 1:7, 10:1 to 1:6, 10:1 to 1:5, 10:1 to 1:4, 10:1 to 1:3, 10:1 to 1:2, or 1:1 to 3:1. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:8, 1:9, or 1:10 in each fraction. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in each fraction.

[0638]

[0675] In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose in at least one fraction is about 10:1 to 1:10, 9:1 to 1:10, 8:1 to 1:10, 7:1 to 1:10, 6:1 to 1:10, 5:1 to 1:10, 4:1 to 1:10, 3:1 to 1:10, 2:1 to 1:10, 10:1 to 1:9, 10:1 to 1:8, 10:1 to 1:7, 10:1 to 1:6, 10:1 to 1:5, 10:1 to 1:4, 10:1 to 1:3, 10:1 to 1:2, or 1:1 to 3:1. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 10:1, 9:1, 8:1, 7:1, 6:1, 5:1, 4:1, 3:1, 2:1, 1:1, 1:2, 1:3, 1:4, 1:5, 1:6, 1:7, 1:8, 1:8, 1:9, or 1:10 in at least one fraction. In some embodiments, the ratio of 1,6-anhydro-β-D-glucofuranose to 1,6-anhydro-β-D-glucopyranose is about 2:1 in at least one fraction.

[0639]

[0676] In some embodiments, the oligosaccharide preparations described herein comprise anhydro-subunit-containing DP2 oligosaccharides. In some embodiments, the oligosaccharide preparations comprise anhydro-lactose, anhydro-sucrose, anhydro-cellobiose, or a combination thereof. In some embodiments, the oligosaccharide preparations comprise about 2-20, 2-15, 5-20, 5-15, or 5-10 DP2 anhydro-subunit-containing oligosaccharides. In some embodiments, the oligosaccharide preparations described herein are free of cellobiosan or do not contain detectable levels of cellobiosan.

[0640]

[0677] In some embodiments, the oligosaccharide preparations described herein contain one or more anhydrous subunits that are sugar caramelization products. In some embodiments, the oligosaccharide preparations containing one or more anhydrous subunits are sugar caramelization products selected from the group consisting of methanol; ethanol; furan; methylglyoxal; 2-methylfuran; vinyl acetate; glycolaldehyde; acetic acid; acetol; furfural; 2-furanmethanol; 3-furanmethanol; 2-hydroxycyclopent-2-en-1-one; 5-methylfurfural; 2(5H)-furanone; 2-methylcyclopentenolone; levoglucosenone; cyclic hydroxylactone; 1,4,3,6-dianhydro-α-D-glucopyranose; dianhydroglucopyranose; and 5-hydroxymethylfurfural (5-hmf). In some embodiments, the oligosaccharide preparations contain 5-hmf anhydrous subunits.

[0641]

[0678] In some embodiments, in at least one of the oligosaccharide preparations or DP fractions, anhydro subunits that are caramelization products are less abundant than anhydro subunits that are thermal dehydration products of monosaccharides. In some embodiments, in at least one of the oligosaccharide preparations or fractions, anhydro subunits that are caramelization products are more abundant than anhydro subunits that are thermal dehydration products of monosaccharides. In some embodiments, in at least one of the oligosaccharide preparations or fractions, anhydro subunits that are caramelization products and anhydro subunits that are thermal dehydration products of monosaccharides have similar abundance.

[0642]

[0679] In some embodiments, the anhydro subunits in the oligosaccharide preparations described herein are from about 0.01% to about 50%, from about 0.01% to about 40%, from about 0.01% to about 30%, from about 0.01% to about 20%, from about 0.01% to about 10%, from about 0.01% to about 5%, from about 0.01% to about 4%, from about 0.01% to about 3%, from about 0.01% to about 2%, or from about 0.01% to about 10%. Between about 0.01% and about 1%, between about 0.01% and about 0.5%, between about 0.1% and about 50%, between about 0.1% and about 40%, between about 0.1% and about 30%, between about 0.1% and about 20%, between about 0.1% and about 10%, between about 0.1% and about 5%, between about 0.1% and about 4%, between about 0.1% and about 3%, between about 0.1% and about 2%, between about 0.1% and about 1%, or between about 0.1% and about 0.5% of the anhydro subunits in the oligosaccharide preparation are caramelization products. In some embodiments, between about 0.1% and about 5%, between about 0.1% and about 2%, or between about 0.1% and about 1% of the anhydro subunits in the oligosaccharide preparation are caramelization products. In some embodiments, less than 50%, less than 40%, less than 30%, less than 25%, less than 20%, less than 15%, less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the anhydro subunits in the oligosaccharide preparation are caramelized products.

[0643]

[0680] In some embodiments, about 0.01% to about 50%, about 0.01% to about 40%, about 0.01% to about 30%, about 0.01% to about 20%, about 0.01% to about 10%, about 0.01% to about 5%, about 0.01% to about 4%, about 0.01% to about 3% of the anhydro subunits in at least one fraction (e.g., DP1, DP2, and / or DP3 fraction) of the preparations described herein. , about 0.01% to about 2%, about 0.01% to about 1%, about 0.01% to about 0.5%, about 0.1% to about 50%, about 0.1% to about 40%, about 0.1% to about 30%, about 0.1% to about 20%, about 0.1% to about 10%, about 0.1% to about 5%, about 0.1% to about 4%, about 0.1% to about 3%, about 0.1% to about 2%, about 0.1% to about 1%, or about 0.1% to about 0.5% of the anhydro subunits in at least one fraction (e.g., DP1, DP2, and / or DP3) of the preparation is a caramelization product. In some embodiments, about 0.1% to about 5%, about 0.1% to about 2%, or about 0.1% to about 1% of the anhydro subunits in at least one fraction (e.g., DP1, DP2, and / or DP3) of the preparation is a caramelization product. In some embodiments, less than 50%, 40%, 30%, 25%, 20%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of the anhydro subunits in at least one fraction of the preparation are caramelization products. In some embodiments, less than 20%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, or 1% of the anhydro subunits in the DP1, DP2, and / or DP3 fractions of the oligosaccharide preparations described herein are caramelization products.

[0644]

[0681] In some embodiments, the anhydro subunits in each fraction of the oligosaccharide preparations described herein are between about 0.01% and about 50%, between about 0.01% and about 40%, between about 0.01% and about 30%, between about 0.01% and about 20%, between about 0.01% and about 10%, between about 0.01% and about 5%, between about 0.01% and about 4%, between about 0.01% and about 3%, between about 0.01% and about 2%, between about 0.01% and about 3%, between about 0.01% and about 4%, between about 0.01% and about 5%, between about 0.01% and about 6%, between about 0.01% and about 6%, between about 0.01% and about 7%, between about 0.01% and about 8%, between about 0.01% and about 9%, between about 0.01% and about 10 ... Between 0.01% and about 1%, between 0.01% and about 0.5%, between 0.1% and about 50%, between 0.1% and about 40%, between 0.1% and about 30%, between 0.1% and about 20%, between 0.1% and about 10%, between 0.1% and about 5%, between 0.1% and about 4%, between 0.1% and about 3%, between 0.1% and about 2%, between 0.1% and about 1%, or between 0.1% and about 0.5% of the anhydro subunits in each fraction of the preparation are caramelization products. In some embodiments, between 0.1% and about 5%, between 0.1% and about 2%, or between 0.1% and about 1% of the anhydro subunits in each fraction of the preparation are caramelization products. In some embodiments, less than 50%, less than 40%, less than 30%, less than 20%, less than 25%, less than 20%, less than 15%, less than 14%, less than 13%, less than 12%, less than 11%, less than 10%, less than 9%, less than 8%, less than 7%, less than 6%, less than 5%, less than 4%, less than 3%, less than 2%, or less than 1% of the anhydro subunits in each fraction of the preparation are caramelization products.

[0645]

[0682] In some embodiments, each of the oligosaccharides in the oligosaccharide preparations described herein independently and optionally comprises an anhydrosubunit. In some embodiments, two or more independent oligosaccharides comprise the same or different anhydrosubunits. In some embodiments, two or more independent oligosaccharides comprise different anhydrosubunits. For example, in some embodiments, the oligosaccharide preparation comprises an oligosaccharide containing a DP1 anhydrosubunit comprising a 1,6-anhydro-β-D-glucopyranose subunit and an oligosaccharide containing a DP2 anhydrosubunit comprising a 1,6-anhydro-β-D-glucofuranose subunit. In some embodiments, one or more oligosaccharides in the oligosaccharide preparation comprise two or more of the same or different anhydrosubunits.

[0646]

[0683] In some embodiments, in any fraction of the oligosaccharide preparation having a degree of polymerization of 2 or greater (i.e., the DP2-DPn fractions), an anhydro subunit may be linked to one or more regular or anhydro subunits. In some embodiments, in the DP2-DPn fractions, at least one anhydro subunit is linked to one, two, or three other regular or anhydro subunits. In some embodiments, in the DP2-DPn fractions, at least one anhydro subunit is linked to one or two regular subunits. In some embodiments, in the DP2-DPn fractions, at least one anhydro subunit is linked to one regular subunit. In some embodiments, in any of the DP2-DPn fractions, more than 99%, 90%, 80%, 70%, 60%, 50%, 40%, or 30% of the anhydro subunits are linked to one regular subunit. In some embodiments, in each of the DP2 through DPn fractions, more than 99%, 90%, 80%, 70%, 60%, 50%, 40%, or 30% of the anhydro subunits are bound to one regular subunit.

[0647]

[0684] In some embodiments, in any fraction of an oligosaccharide preparation having a degree of polymerization of 2 or greater (i.e., DP2-DPn fractions), anhydro subunits can be located at the chain ends of the oligosaccharides. In some embodiments, in any fraction of an oligosaccharide preparation having a degree of polymerization of 3 or greater (i.e., DP3-DPn fractions), anhydro subunits can be located at positions other than the chain ends of the oligosaccharides. In some embodiments, in DP2-DPn fractions, at least one anhydro subunit is located at the chain end of the oligosaccharides. In some embodiments, more than 99%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, or 30% of the anhydro subunits in DP2-DPn fractions are located at the chain ends of the oligosaccharides. In some embodiments, greater than 95%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, or 10% of the anhydro subunits in the oligosaccharide preparation are located at the chain ends of the oligosaccharides. In some embodiments, greater than 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits. In some embodiments, greater than 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the anhydro subunit-containing oligosaccharides comprise chain-terminal anhydro subunits.

[0648] [E. Glycosidic bond]

[0685] In some embodiments, the oligosaccharide preparations described herein comprise a variety of glycosidic linkages. The type and distribution of glycosidic linkages may depend on the source and method of production of the oligosaccharide preparation. In some embodiments, the type and distribution of glycosidic linkages may be determined and / or detected by any suitable method known in the art, such as NMR. For example, in some embodiments, the glycosidic linkages are 1 H NMR, 13In some embodiments, the glycosidic bond is determined and / or detected by, for example, C NMR, 2D NMR such as 2D JRES, HSQC, HMBC, DOSY, COSY, ECOSY, TOCSY, NOESY, or ROESY, or any combination thereof. 1 In some embodiments, the glycosidic bond is at least partially determined and / or detected by H NMR. 13 In some embodiments, the glycosidic bond is at least partially determined and / or detected by 2D C NMR. 1 H, 13 Determined and / or detected by C-HSQC NMR.

[0649]

[0686] In some embodiments, the oligosaccharide preparations described herein comprise one or more α-(1,2) glycosidic linkages, α-(1,3) glycosidic linkages, α-(1,4) glycosidic linkages, α-(1,6) glycosidic linkages, β-(1,2) glycosidic linkages, β-(1,3) glycosidic linkages, β-(1,4) glycosidic linkages, β-(1,6) glycosidic linkages, α-(1,1)-α glycosidic linkages, α-(1,1)-β glycosidic linkages, β-(1,1)-β glycosidic linkages, or any combination thereof.

[0650]

[0687] In some embodiments, the oligosaccharide preparation has from about 0 to about 60 mol%, about 5% to about 55 mol%, about 5% to about 50 mol%, about 5% to about 45 mol%, about 5% to about 40 mol%, about 5% to about 35 mol%, about 5% to about 30 mol%, about 5% to about 25 mol%, about 10% to about 60 mol%, about 10% to about 55 mol%, about 10% to about 50 mol%, about 10% to about 45 mol%, about 10% to about 40 mol%, about 10% to about 35 mol%, about 15% to about 60 mol%, or about 15% to about The glycoside bond distribution is about 55 mol%, about 15% to about 50 mol%, about 15% to about 45 mol%, about 15% to about 40 mol%, about 15% to about 35 mol%, about 20% to about 60 mol%, about 20% to about 55 mol%, about 20% to about 50 mol%, about 20% to about 45 mol%, about 20% to about 40 mol%, about 20% to about 35 mol%, about 25% to about 60 mol%, about 25% to about 55 mol%, about 25% to about 50 mol%, about 25% to about 45 mol%, about 25% to about 40 mol%, or about 25% to about 35 mol%.

[0651]

[0688] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 50 mol%, about 0 to about 40 mol%, about 0 to about 35 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 5% to about 40 mol%, about 5% to about 35 mol%, about 5% to about 30 mol%, about 5% to about 25 mol%, about 5% to about 20 mol%, about 10% to about 40 mol%, about 10% to about 35 mol%, about 10% to about 20 mol%, about 15% to about 40 mol%, about 15% to about 35 mol%, about 15% to about 30 mol%, about 15% to about 25 mol%, or about 15% to about 20 mol% α-(1,3) glycosidic linkages.

[0652]

[0689] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 40 mol%, about 0 to about 35 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, about 2% to about 30 mol%, about 2% to about 25 mol%, about 2% to about 20 mol%, about 2% to about 15 mol%, about 2% to about 10 mol%, about 3% to about 30 mol%, about 3% to about 25 mol%, about 3% to about 20 mol%, about 3% to about 15 mol%, about 3% to about 10 mol%, about 5% to about 30 mol%, about 5% to about 25 mol%, about 5% to about 20 mol%, about 5% to about 15 mol%, or about 5% to about 10 mol%.

[0653]

[0690] In some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution of about 0 to about 40 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, or about 0 to about 5 mol% α-(1,4) glycosidic linkages. In some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution of less than 40 mol%, less than 30 mol%, less than 20 mol%, less than 15 mol%, less than 10 mol%, less than 9 mol%, less than 8 mol%, less than 7 mol%, less than 6 mol%, less than 5 mol%, less than 4 mol%, less than 3 mol%, or less than 2 mol% α-(1,4) glycosidic linkages.

[0654]

[0691] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 40 mol%, about 0 to about 35 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, about 2% to about 30 mol%, about 2% to about 25 mol%, about 2% to about 20 mol%, about 2% to about 15 mol%, about 2% to about 10 mol%, about 5% to about 30 mol%, about 5% to about 25 mol%, about 5% to about 20 mol%, about 5% to about 15 mol%, about 5% to about 10 mol%, about 8% to about 30 mol%, about 8% to about 25 mol%, about 8% to about 20 mol%, about 8% to about 15 mol%, or about 10% to about 15 mol%.

[0655]

[0692] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 40 mol%, about 0 to about 35 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, about 2% to about 30 mol%, about 2% to about 25 mol%, about 2% to about 20 mol%, about 2% to about 15 mol%, about 2% to about 10 mol%, about 3% to about 30 mol%, about 3% to about 25 mol%, about 3% to about 20 mol%, about 3% to about 15 mol%, about 3% to about 10 mol%, about 5% to about 30 mol%, about 5% to about 25 mol%, about 5% to about 20 mol%, about 5% to about 15 mol%, or about 5% to about 10 mol% β-(1,4) glycosidic linkages.

[0656]

[0693] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 40 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, about 0 to about 5 mol%, about 1% to about 20 mol%, about 1% to about 15 mol%, about 1% to about 10 mol%, about 1% to about 5 mol%, about 2% to about 20 mol%, about 2% to about 15 mol%, about 2% to about 10 mol%, or about 2% to about 5 mol% β-(1,2) glycosidic linkages. In some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution of less than 40 mol%, less than 30 mol%, less than 20 mol%, less than 15 mol%, less than 10 mol%, less than 9 mol%, less than 8 mol%, less than 7 mol%, less than 6 mol%, less than 5 mol%, less than 4 mol%, less than 3 mol%, or less than 2 mol% β-(1,2) glycosidic linkages.

[0657]

[0694] In some embodiments, the oligosaccharide preparation has a glycosidic bond distribution of about 0 to about 40 mol%, about 0 to about 30 mol%, about 0 to about 25 mol%, about 0 to about 20 mol%, about 0 to about 15 mol%, about 0 to about 10 mol%, about 0 to about 5 mol%, about 1% to about 20 mol%, about 1% to about 15 mol%, about 1% to about 10 mol%, about 1% to about 5 mol%, about 2% to about 20 mol%, about 2% to about 15 mol%, about 2% to about 10 mol%, or about 2% to about 5 mol% β-(1,3) glycosidic linkages. In some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution of less than 40 mol%, less than 30 mol%, less than 20 mol%, less than 15 mol%, less than 10 mol%, less than 9 mol%, less than 8 mol%, less than 7 mol%, less than 6 mol%, less than 5 mol%, less than 4 mol%, less than 3 mol%, or less than 2 mol% β-(1,3) glycosidic linkages.

[0658]

[0695] In some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution that differs from that of a non-synthetic oligosaccharide preparation. For example, in some embodiments, the oligosaccharide preparation has a glycosidic linkage distribution that differs from that of a basal nutritional composition. In some embodiments, the basal nutritional composition includes a natural carbohydrate source, such as starch and plant fiber. Some natural carbohydrate sources have a high proportion of α-(1,4), α-(1,6), and / or β-(1,6) glycosidic linkages. Thus, in some embodiments, the oligosaccharide preparation has a lower proportion of α-(1,4) glycosidic linkages than the basal nutritional composition. In some embodiments, the oligosaccharide preparation has a lower proportion of α-(1,6) glycosidic linkages than the basal nutritional composition. In other embodiments, the oligosaccharide preparation has a higher proportion of α-(1,6) glycosidic linkages than the basal nutritional composition. In some embodiments, the oligosaccharide preparation has a lower percentage of β-(1,6) glycosidic linkages than the basal nutritional composition, hi some embodiments, the oligosaccharide preparation comprises glycosidic linkages that are not readily digested or hydrolyzed by enzymes.

[0659]

[0696] Specifically, in some embodiments, the glycosidic linkage distribution of the oligosaccharide preparations described herein has at least 50 mol%, at least 40 mol%, at least 30 mol%, at least 20 mol%, at least 15 mol%, at least 10 mol%, at least 5 mol%, at least 2 mol%, or at least 1 mol% lower than that of the basal nutritional composition. In some embodiments, the glycosidic linkage distribution of the oligosaccharide preparation is at least 50 mol%, at least 40 mol%, at least 30 mol%, at least 20 mol%, at least 15 mol%, at least 10 mol%, at least 5 mol%, at least 2 mol%, or at least 1 mol% higher than that of the basal nutritional composition.

[0660]

[0697] It should be understood by those skilled in the art that a particular type of glycosidic bond may not be applicable to an oligosaccharide containing a particular type of monosaccharide. For example, in some embodiments, the oligosaccharide preparation comprises an α-(1,2) glycosidic bond and an α-(1,6) glycosidic bond. In other embodiments, the oligosaccharide preparation comprises an α-(1,2) glycosidic bond and a β-(1,3) glycosidic bond. In some embodiments, the oligosaccharide preparation comprises an α-(1,2) glycosidic bond, an α-(1,3) glycosidic bond, and a β-(1,6) glycosidic bond. In some embodiments, the oligosaccharide preparation comprises α-(1,2) glycosidic linkages, α-(1,3) glycosidic linkages, α-(1,4) glycosidic linkages, α-(1,6) glycosidic linkages, β-(1,2) glycosidic linkages, β-(1,3) glycosidic linkages, β-(1,4) glycosidic linkages, and β-(1,6) glycosidic linkages.

[0661] [F.Molecular weight]

[0698] The molecular weight and molecular weight distribution of the oligosaccharide preparations can be determined by any suitable analytical means and instrumentation, such as end-group methods, osmotic pressure (osmometry), ultracentrifugation, viscometry, light scattering, SEC, SEC-MALLS, FFF, A4F, HPLC, and mass spectrometry. In some embodiments, the molecular weight and molecular weight distribution are determined by mass spectrometry, such as MALDI-MS, LC-MS, or GC-MS. In some embodiments, the molecular weight and molecular weight distribution are determined by size exclusion chromatography (SEC), such as gel permeation chromatography (GPC). In other embodiments, the molecular weight and molecular weight distribution are determined by HPLC. In some embodiments, the molecular weight and molecular weight distribution are determined by MALDI-MS.

[0662]

[0699] In some embodiments, the oligosaccharide preparations described herein comprise from about 100 to about 10,000 g / mol, from about 200 to about 8,000 g / mol, from about 300 to about 5,000 g / mol, from about 500 to about 5,000 g / mol, from about 700 to about 5,000 g / mol, from about 900 to about 5,000 g / mol, from about 1100 to about 5,000 g / mol, from about 1300 to about 5,000 g / mol, from about 1500 to about 5,000 g / mol, from about 1700 to about 5,000 g / mol, from about 300 to about 4,500 g / mol, from about 500 to about 4,500 g / mol, from about 700 to about 4,500 g / mol, from about 900 to about 4,500 g / mol, / mol, about 1100 to about 4500 g / mol, about 1300 to about 4500 g / mol, about 1500 to about 4500 g / mol, about 1700 to about 4500 g / mol, about 1900 to about 4500 g / mol, about 300 to about 4000 g / mol, about 500 to about 4000 g / mol, about 700 to about 4000 g / mol, about 900 to about 4000 g / mol, about 1100 to about 4000 g / mol, about 1300 to about 4000 g / mol, about 1500 to about 4000 g / mol, about 1700 to about 4000 g / mol, about 1900 to about 4000 g / mol, about 300 to about 3000 g / mol, about 500 to about 3000 g / mol, about 700 to about 3000 g / mol, about 900 to about 3000 g / mol, about 1100 to about 3000 g / mol, about 1300 to about 3000 g / mol, about 1500 to about 3000 g / mol, about 1700 to about 3000 g / mol, about 1900 to about 3000 g / mol, about 2100 to about 3000 g / mol, about 300 to about 2500 g / mol, about 500 to about 2500 g / mol, about 700 to about 2500 g / mol, about 900 to about 2500 g / mol, about 1100 to about 2500 g / mol, about 1300 to about 2500 g / mol, about 1500 to about 2 The weight average molecular weight is about 500 g / mol, about 1700 to about 2500 g / mol, about 1900 to about 2500 g / mol, about 2100 to about 2500 g / mol, about 300 to about 1500 g / mol, about 500 to about 1500 g / mol, about 700 to about 1500 g / mol, about 900 to about 1500 g / mol, about 1100 to about 1500 g / mol, about 1300 to about 1500 g / mol, about 2000 to about 2800 g / mol, about 2100 to about 2700 g / mol, about 2200 to about 2600 g / mol, about 2300 to about 2500 g / mol, or about 2320 to about 2420 g / mol.In some embodiments, the weight-average molecular weight of the oligosaccharide preparation is about 2000 to about 2800 g / mol, about 2100 to about 2700 g / mol, about 2200 to about 2600 g / mol, about 2300 to about 2500 g / mol, or about 2320 to about 2420 g / mol. In some embodiments, the oligosaccharide preparation has a weight-average molecular weight ranging from at least 500 g / mol, 750 g / mol, 1000 g / mol, or 1500 g / mol to a maximum of 1750 g / mol, 2000 g / mol, 2250 g / mol, 2500 g / mol, or 3000 g / mol. In some embodiments, the weight-average molecular weight of the oligosaccharide preparations described herein is determined by HPLC according to Example 9.

[0663]

[0700] In some embodiments, the oligosaccharide preparations described herein comprise from about 100 to about 10,000 g / mol, about 200 to about 8,000 g / mol, about 300 to about 5,000 g / mol, about 500 to about 5,000 g / mol, about 700 to about 5,000 g / mol, about 900 to about 5,000 g / mol, about 1,100 to about 5,000 g / mol, about 1,300 to about 5,000 g / mol, about 1,500 to about 5,000 g / mol, about 1,700 to about 5,000 g / mol, about 300 to about 4,500 g / mol, about 500 to about 4,500 g / mol, about 700 to about 4,500 g / mol, about 900 to about 4,500 g / mol, about 1,100 to about 4500 g / mol, about 1300 to about 4500 g / mol, about 1500 to about 4500 g / mol, about 1700 to about 4500 g / mol, about 1900 to about 4500 g / mol, about 300 to about 4000 g / mol, about 500 to about 4000 g / mol, about 700 to about 4000 g / mol, about 900 to about 4000 g / mol, about 1100 to about 4000 g / mol, about 1300 to about 4000 g / mol, about 1500 to about 4000 g / mol, about 1700 to about 4000 g / mol, about 1900 to about 4000 g / mol, about 300 to about 3000 g / mol, about 500 to about 3000 g / mol, about 700 to about 3000 g / mol, about 900 to about 3000 g / mol, about 1100 to about 3000 g / mol, about 1300 to about 3000 g / mol, about 1500 to about 3000 g / mol, about 1700 to about 3000 g / mol, about 1900 to about 3000 g / mol, about 2100 to about 3000 g / mol, about 300 to about 2500 g / mol, about 500 to about 2500 g / mol, about 700 to about 2500 g / mol, about 900 to about 2500 g / mol, about 1100 to about 2500 g / mol, about 1300 to about 2500 g / mol, about 1500 to about 2500 g / mol, about 1700 to about 2500 g / mol, about 19 00 to about 2500 g / mol, about 2100 to about 2500 g / mol, about 300 to about 2000 g / mol, about 500 to about 300 to 2000 g / mol, about 700 to about 2000 g / mol, about 900 to about 2000 g / mol, about 1100 to about 2000 g / mol, about 300 to about 1500 g / mol, about 500 to about 1500 g / mol, about 700 to about 1500 g / mol, about 900 to about 1500 g / mol, about 1100 to about 1500 g / mol, about 1300 to about 1500 g / mol, about 1000 to about 2000 g / mol, about 1100 to about 1900 g / mol, about 1200 to about 1800 g / mol,The oligosaccharide preparation has a number average molecular weight of about 1300 to about 1700 g / mol, about 1400 to about 1600 g / mol, or about 1450 to about 1550 g / mol. In some embodiments, the number average molecular weight of the oligosaccharide preparation is about 1000 to about 2000 g / mol, about 1100 to about 1900 g / mol, about 1200 to about 1800 g / mol, about 1300 to about 1700 g / mol, 1400 to 1600 g / mol, or 1450 to 1550 g / mol. In some embodiments, the oligosaccharide preparation has a number average molecular weight ranging from at least 500 g / mol, 750 g / mol, 1000 g / mol, or 1500 g / mol to a maximum of 1750 g / mol, 2000 g / mol, 2250 g / mol, 2500 g / mol, or 3000 g / mol. In some embodiments, the number average molecular weight of the oligosaccharide preparations described herein is determined by HPLC according to Example 9.

[0664] [G. Types of Oligosaccharides]

[0701] The species of oligosaccharides present in an oligosaccharide preparation can depend on one or more types of feed sugars. For example, in some embodiments, if the feed sugar comprises glucose, the oligosaccharide preparation comprises gluco-oligosaccharides. For example, in some embodiments, if the feed sugar comprises galactose, the oligosaccharide preparation comprises galacto-oligosaccharides. For another example, in some embodiments, if the feed sugar comprises galactose and glucose, the oligosaccharide preparation comprises gluco-galacto-oligosaccharides.

[0665]

[0702] In some embodiments, the oligosaccharide preparations described herein comprise one or more species of monosaccharide subunits, hi some embodiments, the oligosaccharide preparations comprise oligosaccharides having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more different species of monosaccharide subunits.

[0666]

[0703] In some embodiments, the oligosaccharide preparation comprises oligosaccharides having one, two, three, or four different species of monosaccharide subunits. In some embodiments, the oligosaccharide preparation comprises oligosaccharides having one, two, or three different species of monosaccharide subunits. In some embodiments, the oligosaccharide preparation comprises oligosaccharides having three different species of monosaccharide subunits. In some embodiments, the oligosaccharide preparation comprises oligosaccharides having two different species of monosaccharide subunits. In some embodiments, the oligosaccharide preparation comprises oligosaccharides having one species of monosaccharide subunits.

[0667]

[0704] In some embodiments, the oligosaccharide preparations comprise different oligosaccharide species, where each oligosaccharide molecule independently comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 different species of monosaccharide subunits. 2 , 10 3 , 10 4 , 10 5 In some embodiments, some of the oligosaccharides in a preparation contain monosaccharide subunits of one species, while others of the oligosaccharides in the same preparation contain monosaccharide subunits of two or more species. For example, in some embodiments, if the feed sugars are glucose and galactose, the oligosaccharide preparation can contain oligosaccharides containing only glucose subunits, oligosaccharides containing only galactose subunits, oligosaccharides containing both glucose and galactose subunits in various ratios, or any combination thereof.

[0668]

[0705] In some embodiments, any or all of the n fractions of an oligosaccharide preparation comprise different species of oligosaccharide subunits, where each oligosaccharide independently comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 different species of monosaccharide subunits. In some embodiments, some of the oligosaccharides in a fraction of the preparation comprise monosaccharide subunits of one species, and some other of the oligosaccharides in the same fraction of the preparation comprise monosaccharide subunits of two or more species.

[0669]

[0706] In some embodiments, the oligosaccharide preparations described herein comprise one or more monosaccharide subunits selected from the group consisting of triose, tetrose, pentose, hexose, heptose, and any combination thereof, wherein each of the triose, tetrose, pentose, hexose, or heptose subunits is independently and optionally functionalized and / or substituted with one of its corresponding anhydrous subunits. In some embodiments, the corresponding anhydrous subunit is a thermal dehydration product of the monosaccharide subunit. In some embodiments, the corresponding anhydrous subunit is a caramelization product of the monosaccharide subunit.

[0670]

[0707] In some embodiments, the oligosaccharide preparations described herein comprise pentose subunits, hexose subunits, or any combination thereof, wherein each of the pentose or hexose subunits is independently and optionally functionalized and / or substituted with one of its corresponding anhydro subunits. In some embodiments, the oligosaccharide preparations comprise hexose subunits, wherein each of the hexose subunits is independently and optionally functionalized and / or substituted with one of its corresponding anhydro subunits.

[0671]

[0708] As used herein, tetrose refers to a monosaccharide having four carbon atoms, such as erythrose, threose, and erythrulose. As used herein, pentose refers to a monosaccharide having five carbon atoms, such as arabinose, lyxose, ribose, and xylose. As used herein, hexose refers to a monosaccharide having six carbon atoms, such as allose, altrose, glucose, mannose, gulose, idose, galactose, talose, psicose, fructose, sorbose, and tagatose. As used herein, heptose refers to a monosaccharide having seven carbon atoms, such as sedoheptulose and mannoheptulose.

[0672]

[0709] In some embodiments, the oligosaccharide preparations described herein comprise glucose subunits, wherein at least one glucose subunit is optionally replaced with an anhydro-glucose subunit. In some embodiments, the oligosaccharide preparations described herein comprise galactose subunits, wherein at least one galactose subunit is optionally replaced with an anhydro-galactose subunit. In some embodiments, the oligosaccharide preparations described herein comprise galactose and glucose subunits, wherein at least one galactose subunit or at least one glucose subunit is optionally replaced with one of its corresponding anhydro-subunits. In some embodiments, the oligosaccharide preparations described herein comprise fructose and glucose subunits, wherein at least one fructose subunit or at least one glucose subunit is optionally replaced with one of its corresponding anhydro-subunits. In some embodiments, the oligosaccharide preparations described herein comprise mannose and glucose subunits, wherein at least one mannose subunit or at least one glucose subunit is optionally substituted with one of its corresponding anhydro subunits.

[0673]

[0710] In some embodiments, the oligosaccharide preparations described herein are selected from the group consisting of gluco-galactose-oligosaccharide preparations, gluco-oligosaccharide preparations, galacto-oligosaccharide preparations, fructo-oligosaccharide preparations, manno-oligosaccharide preparations, arabino-oligosaccharide preparations, xylo-oligosaccharide preparations, gluco-fructo-oligosaccharide preparations, gluco-manno-oligosaccharide preparations, gluco-arabino-oligosaccharide preparations, gluco-xylo-oligosaccharide preparations, galacto-fructo-oligosaccharide preparations, galacto-manno-oligosaccharide preparations, galacto-arabino-oligosaccharide preparations, galacto-xylo-oligosaccharide preparations, fructo-manno-oligosaccharide preparations, fructo-arabino-oligosaccharide preparations, fructo-xylo-oligosaccharide preparations. and / or any combination thereof, wherein each of the monosaccharide subunits within the preparation is independently and optionally functionalized and / or substituted with one of its corresponding anhydro subunits.

[0674]

[0711] In certain embodiments, the oligosaccharide preparations described herein comprise greater than 99% glucose subunits by weight, hi some embodiments, the oligosaccharide preparations comprise only glucose subunits.

[0675]

[0712] In some embodiments, the oligosaccharide preparations described herein contain about 45%-55% glucose subunits and about 55%-45% galactose subunits by weight, hi some particular embodiments, the oligosaccharide preparations contain about 50% glucose and 50% galactose subunits by weight.

[0676]

[0713] In some embodiments, the oligosaccharide preparations described herein comprise, by weight, about 80%-95% glucose subunits and about 20%-5% mannose subunits, hi some embodiments, the oligosaccharide preparations comprise, by weight, about 85%-90% glucose subunits and about 15%-10% mannose subunits.

[0677]

[0714] In some embodiments, the oligosaccharide preparations described herein comprise, by weight, about 80%-95% glucose subunits and about 20%-5% galactose subunits, hi some embodiments, the oligosaccharide preparations comprise, by weight, about 85%-90% glucose subunits and about 15%-10% galactose subunits.

[0678]

[0715] In some embodiments, the oligosaccharide preparations described herein comprise, by weight, about 80%-95% glucose subunits, 0%-8% galactose subunits, and 5%-20% mannose subunits. In some embodiments, the oligosaccharide preparations comprise, by weight, about 80%-90% glucose subunits, 1%-5% galactose subunits, and 10%-15% mannose subunits.

[0679]

[0716] In some embodiments, the oligosaccharide preparations described herein contain about 1% to about 100%, about 50% to about 100%, about 80% to about 98%, or about 85% to about 95% by weight of glucose subunits, or any range therebetween. In some embodiments, galactose subunits are present in the oligosaccharide preparations described herein in an amount of about 0% to about 90%, about 1% to about 50%, about 2% to about 20%, or about 5% to about 15% by weight, or any range therebetween. In some embodiments, mannose subunits are present in the oligosaccharide preparations described herein in an amount of about 0% to about 90%, about 1% to about 50%, about 2% to about 20%, or about 5% to about 15% by weight, or any range therebetween.

[0680]

[0717] In some embodiments, the oligosaccharide preparations described herein have a monosaccharide subunit composition as shown in Table 26.

[0681] [Table 1]

[0682] [HD type vs. L type]

[0718] In some embodiments, at least one monosaccharide subunit in the oligosaccharide is in the L-form. In some embodiments, at least one monosaccharide subunit in the oligosaccharide is in the D-form. In some embodiments, the monosaccharide subunits in the oligosaccharide preparations described herein are in their naturally abundant form, such as D-glucose, D-xylose, and L-arabinose.

[0683]

[0719] In some embodiments, the oligosaccharide preparations described herein comprise a mixture of L- and D-type monosaccharide subunits. In some embodiments, the ratio of L- to D- or D- to L-type monosaccharide subunits is about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9, about 1:10, about 1:12, about 1:14, about 1:16, about 1:18, about 1:20, about 1:25, about 1:30, about 1:35, about 1:40, about 1:45, about 1:50, about 1:55, about 1:60, about 1:65, about 1:70, about 1:75, about 1:80, about 1:85, about 1:90, about 1:100, or about 1:150.

[0684] [I. Functionalized Oligosaccharides]

[0720] In some embodiments, one or more oligosaccharides in the preparation are independently functionalized.Functionalized oligosaccharides can be produced, for example, by combining one or more sugars with one or more functionalizing compounds in the presence of a catalyst.Methods for producing functionalized oligosaccharides are described in International Publication Nos. 2012 / 118767, 2014 / 031956, and 2016 / 122887 (their entire disclosures are incorporated herein by reference).

[0685]

[0721] In some embodiments, the functionalized compound contains one or more acidic groups (e.g., —COOH), hydroxyl groups, N-containing groups (e.g., —CN, —NO, and —N(R a )2[wherein, R a is a hydrogen group, an alkyl group, an alkenyl group, an alkynyl group, a haloalkyl group, a heteroalkyl group, a cycloalkyl group, an aryl group, a heterocycloalkyl group, or a heteroaryl group, an S-containing group (e.g., thiol and sulfate), a halide (e.g., -Cl), a P-containing group (e.g., phosphate), or any combination thereof. In some embodiments, the functionalized compound is attached to at least one monosaccharide subunit through an ether bond, an ester bond, an oxygen-sulfur bond, an amine bond, or an oxygen-phosphorus bond. In some embodiments, one or more functionalized compounds are attached to the monosaccharide subunit through a single bond. In some embodiments, at least one functionalized compound is attached to one or two oligosaccharides through two or more bonds.

[0686]

[0722] For each oligosaccharide in the oligosaccharide preparation, each of the described embodiments is independent and can be combined as if each combination were described separately; therefore, it should be understood that any combination of embodiments is included in the present disclosure. For example, various embodiments can be grouped into several categories, including, but not limited to, (i) the presence or absence of anhydrosubunits; (ii) the number and level of anhydrosubunits; (iii) the type of anhydrosubunit species; (iv) the position of anhydrosubunits; (v) the degree of polymerization; (vi) molecular weight; (vii) the presence or absence of optional functional groups; (viii) the type of oligosaccharide; (ix) the type of glycosidic bond; and (x) L-type vs. D-type. Thus, the described oligosaccharide preparations contain multiple oligosaccharides of different species. In some embodiments, the oligosaccharide preparations described herein contain at least 10, 10 2 , 10 3 , 10 4 , 10 5 , 106 , 10 7 , 10 8 , 10 9 or 10 10 In some embodiments, the preparation comprises at least 10 different oligosaccharide species. 3 , 10 4 , 10 5 , 10 6 or 10 9 In some embodiments, the preparation comprises at least 10 different oligosaccharide species. 3 It contains different oligosaccharide species.

[0687] III. Methods for Producing Oligosaccharide Preparations

[0723] In one aspect, provided herein are methods for producing an oligosaccharide preparation. In some embodiments, provided herein are methods for producing an oligosaccharide preparation suitable for use in nutritional compositions, such as animal feed compositions, or fed directly to an animal. In one aspect, provided herein are methods for producing an oligosaccharide preparation, comprising heating an aqueous composition comprising one or more feed sugars and a catalyst to a temperature and for a time sufficient to induce polymerization, wherein the catalyst is selected from the group consisting of (+)-camphor-10-sulfonic acid; 2-pyridinesulfonic acid; 3-pyridinesulfonic acid; 8-hydroxy-5-quinolinesulfonic acid hydrate; α-hydroxy-2-pyridinemethanesulfonic acid; (β)-camphor-10-sulfonic acid; butylphosphonic acid; diphenylphosphinic acid; hexyl Phosphonic acid;Methylphosphonic acid;Phenylphosphinic acid;Phenylphosphonic acid;tert-Butylphosphonic acid;(SS)-VAPOL hydrogen phosphate;6-Quinolinesulfonic acid, 3-(1-pyridinio)-1-propanesulfonate;2-(2-Pyridinyl)ethanesulfonic acid;3-(2-Pyridyl)-5,6-diphenyl-1,2,4-triazine-p,p'-disulfonic acid monosodium salt hydrate;1,1'-Binaphthyl-2,2'-diyl hydrogen phosphate;Bis(4-methoxyphenyl)phosphinic acid ;Phenyl(3,5-xylyl)phosphinic acid;L-Cysteic acid monohydrate;Poly(styrenesulfonic acid-co-divinylbenzene);Lysine;Ethanedisulfonic acid;Ethanesulfonic acid;Isethionic acid;Homocysteic acid;HEPBS (N-(2-hydroxyethyl)piperazine-N'-(4-butanesulfonic acid));HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid);2-Hydroxy-3-morpholinopropanesulfonic acid;2-(N-morpholino)ethanesulfonic acid Sulfonic acid;Methanesulfonic acid;Methaniazid;Naphthalene-1-sulfonic acid;Naphthalene-2-sulfonic acid;Perfluorobutanesulfonic acid;6-sulfoquinovose;Trifluoromethanesulfonic acid;2-aminoethanesulfonic acid;Benzoic acid;Chloroacetic acid;Trifluoroacetic acid;Caproic acid;Enanthic acid;Caprylic acid;Pelargonic acid;Lauric acid;Palmitic acid;Stearic acid;Arachidic acid;Aspartic acid;Glutamic acid;Serine;Threonine;Glutamine;Cysteine;Glycine;Proline;The oligosaccharide preparation is selected from the group consisting of alanine, valine, isoleucine, leucine, methionine, phenylalanine, tyrosine, and tryptophan, and the oligosaccharide preparation comprises at least n fractions of oligosaccharides each having a distinct degree of polymerization selected from 1 (DP1 fraction) to n (DPn fraction), where n is an integer greater than or equal to 3.

[0688]

[0724] In some embodiments, n is an integer greater than or equal to 3. In some embodiments, n is an integer in the range of 1 to 100, such as 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, or 50. In some embodiments, polymerization of the feed sugar is achieved by step-growth polymerization. In some embodiments, polymerization of the feed sugar is achieved by polycondensation.

[0689] [A. Supply sugar]

[0725] In some embodiments, the methods of producing an oligosaccharide preparation described herein include heating one or more types of feed sugars, in some embodiments, the one or more types of feed sugars include monosaccharides, disaccharides, trisaccharides, tetrasaccharides, or any mixture thereof.

[0690]

[0726] In some embodiments, the one or more feed sugars comprise glucose. In some embodiments, the one or more feed sugars comprise glucose and galactose. In some embodiments, the one or more feed sugars comprise glucose, xylose, and galactose. In some embodiments, the one or more feed sugars comprise glucose and mannose. In some embodiments, the one or more feed sugars comprise glucose and fructose. In some embodiments, the one or more feed sugars comprise glucose, fructose, and galactose. In some embodiments, the one or more feed sugars comprise glucose, galactose, and mannose.

[0691]

[0727] In some embodiments, the one or more feed sugars comprise disaccharides, such as lactose, sucrose, and cellobiose. In some embodiments, the one or more feed sugars comprise trisaccharides, such as maltotriose or raffinose. In certain embodiments, the one or more feed sugars comprise glucose, mannose, galactose, xylose, maltodextrin, arabinose, or galactose, or any combination thereof. In certain embodiments, the one or more feed sugars comprise a sugar syrup, such as corn syrup. In some embodiments, the one or more feed sugars comprise glucose and lactose. In some embodiments, the one or more feed sugars comprise glucose and sucrose.

[0692]

[0728] In some embodiments, the type of feed sugar can affect the resulting oligosaccharide preparation produced. For example, in some variations where one or more feed sugars are all glucose, the resulting oligosaccharide preparation comprises a gluco-oligosaccharide preparation. In other embodiments, where one or more feed sugars are all mannose, the resulting oligosaccharide preparation comprises a manno-oligosaccharide preparation. In some embodiments, where one or more feed sugars comprise glucose and galactose, the resulting oligosaccharide preparation comprises a gluco-galacto-oligosaccharide preparation. In yet other embodiments, where one or more feed sugars comprise xylose, glucose, and galactose, the resulting oligosaccharide preparation comprises a gluco-galacto-xylo-oligosaccharide preparation.

[0693]

[0729] In some embodiments, each of the one or more feed sugars may be independently in its dehydrated or hydrated form. In some embodiments, the one or more feed sugars comprise glucose, galactose, fructose, mannose, or any combination thereof, and each of glucose, galactose, fructose, or mannose is independently in its monohydrate or dehydrated form. In some embodiments, the one or more feed sugars comprise a monosaccharide monohydrate, such as glucose monohydrate. In some embodiments, the one or more feed sugars comprise a sugar dihydrate, such as trehalose dihydrate. In some embodiments, the one or more feed sugars comprise at least one sugar in its dehydrate form and at least one sugar in its hydrate form.

[0694]

[0730] In some embodiments, one or more feed sugars can be provided as a sugar solution in which the sugars are combined with water and fed to the reactor. In some embodiments, the sugars can be fed to the reactor in solid form and combined with water in the reactor. In some embodiments, the one or more feed sugars are combined and mixed before adding water. In other embodiments, the one or more feed sugars are combined with water and then mixed.

[0695]

[0731] In some embodiments, the method comprises combining two or more feed sugars with a catalyst to produce an oligosaccharide preparation. In some embodiments, the two or more feed sugars comprise glucose, galactose, fructose, mannose, lactose, or any combination thereof. In some embodiments, the method comprises combining a mixture of sugars (e.g., monosaccharides, disaccharides, and / or trisaccharides) with a catalyst to produce the oligosaccharide preparation. In other embodiments, the method comprises combining a mixture of sugars and sugar alcohols with a catalyst to produce the oligosaccharide preparation.

[0696]

[0732] In some embodiments, one or more of the source sugars comprises a functionalized or modified sugar. The functionalized or modified sugar may comprise an amino sugar, a sugar acid, a sugar alcohol, a sugar amide, a sugar ether, or any combination thereof. In some embodiments, an amino sugar refers to a sugar molecule in which a hydroxyl group has been replaced with an amine group. Exemplary amino sugars include, but are not limited to, N-acetyl-d-glucosamine, mannosamine, neuraminic acid, muramic acid, N-acetyl-neuramine, N-acetyl-muramine, N-acetyl-galactosamine, N-acetyl-mannosa, N-glycolylneuram, acarviosin, D-glucosamine, and D-galactosamine.

[0697]

[0733] In embodiments, sugar acid refers to a sugar having a carboxyl group. Exemplary sugar acids include, but are not limited to, aldonic acids (such as glyceric acid, xylonic acid, gluconic acid, and ascorbic acid), ulosonic acids (such as neuraminic acid and ketodeoxyoctulosonic acid), uronic acids (such as glucuronic acid, galacturonic acid, and iduronic acid), and aldaric acids (such as tartaric acid, mucic acid, and saccharinic acid).

[0698]

[0734] In some embodiments, sugar alcohol refers to a polyol derived from a sugar. Exemplary sugar alcohols include, but are not limited to, ethylene glycol, arabitol, glycerol, erythritol, threitol, xylitol, ribitol, mannitol, sorbitol, galactitol, fucitol, iditol, inositol, and volemitol.

[0699]

[0735] In some embodiments, sugar amide refers to a sugar molecule containing a -C(=O)-N- group. In embodiments, sugar ether refers to a sugar molecule containing an ether bond, such as a glucoside.

[0700]

[0736] In some embodiments, the functionalized or modified sugar comprises glucosamine, N-acetylglucosamine, glucuronic acid, galacturonic acid, glucitol, xylitol, mannitol, sorbitol, hi some embodiments, the one or more feed sugars comprise a deoxy sugar, such as fucose, rhamnose, deoxyribose, or fuculose.

[0701]

[0737] In some embodiments, the methods for producing oligosaccharide preparations described herein are carried out on a gram scale. In some embodiments, the methods for producing oligosaccharide preparations described herein are carried out on a kilogram or larger scale. Thus, in some embodiments, the methods comprise heating an aqueous composition comprising one or more feed sugars in an amount greater than 0.5 kg, greater than 1 kg, greater than 2 kg, greater than 3 kg, greater than 4 kg, greater than 5 kg, greater than 6 kg, greater than 7 kg, greater than 9 kg, greater than 10 kg, greater than 100 kg, or greater than 1000 kg. In some embodiments, the methods comprise heating an aqueous composition comprising one or more feed sugars in an amount equal to or less than 0.5, 1, 2, 3, 4, 5, 6, 7, 9, 10, 100, 1000, or 1500 kg. In some embodiments, the methods comprise heating an aqueous composition comprising one or more feed sugars in an amount equal to or less than 1 kg.

[0702] [B.Catalyst]

[0738] In some embodiments, the catalyst provided herein comprises one or more acids. In some embodiments, the catalyst provided herein comprises one or more acids, such as mineral acids, carboxylic acids, amino acids, sulfonic acids, boronic acids, phosphonic acids, phosphinic acids, sulfuric acids, phosphoric acids, poly(styrenesulfonic acid-co-vinylbenzyl-imidazolium sulfate-co-divinylbenzene), poly(styrenesulfonic acid-co-divinylbenzene), (+)-camphor-10-sulfonic acid, 2-pyridinesulfonic acid, 3-pyridinesulfonic acid, 8-hydroxy-5-quinolinesulfonic acid hydrate, α-hydroxy- 2-Pyridinemethanesulfonic acid;(β)-Camphor-10-sulfonic acid;Butylphosphonic acid;Diphenylphosphinic acid;Hexylphosphonic acid;Methylphosphonic acid;Phenylphosphinic acid;Phenylphosphonic acid;tert-Butylphosphonic acid;(SS)-VAPOL hydrogen phosphate;6-Quinolinesulfonic acid;3-(1-Pyridinio)-1-propanesulfonate;2-(2-Pyridinyl)ethanesulfonic acid;3-(2-Pyridyl)-5,6-diphenyl-1,2,4-triazine-p,p'-disulfonic acid Phosphine acid monosodium salt hydrate;1,1'-binaphthyl-2,2'-diyl hydrogen phosphate;Bis(4-methoxyphenyl)phosphinic acid;Phenyl(3,5-xylyl)phosphinic acid;L-Cysteic acid monohydrate;Acetic acid;Propionic acid;Butanoic acid;Glutamic acid;Lysine;Ethanedisulfonic acid;Ethanesulfonic acid;Isethionic acid;Homocysteic acid;HEPBS (N-(2-hydroxyethyl)piperazine-N'-(4-butanesulfonic acid));HEPES (4-(2-hydroxyethyl)-1-piperazine Perazineethanesulfonic acid; 2-hydroxy-3-morpholinopropanesulfonic acid; 2-(N-morpholino)ethanesulfonic acid; Methanesulfonic acid; Methaniazid; Naphthalene-1-sulfonic acid; Naphthalene-2-sulfonic acid; Perfluorobutanesulfonic acid; 6-sulfoquinovose; Trifluoromethanesulfonic acid; 2-aminoethanesulfonic acid; Benzoic acid; Chloroacetic acid; Trifluoroacetic acid; Caproic acid; Enanthic acid; Caprylic acid; Pelargonic acid; Lauric acid; Pamitic acid; Stearic acid; Arachidic acid; Aspartic acid; Glutamic acid; Serine; Threonine; Glutamine; Cysteine; Glycine; Proline; Alanine; Valineisoleucine; leucine; methionine; phenylalanine; tyrosine; tryptophan; polymeric acids; carbon-bearing acids; or any combination thereof;

[0703]

[0739] In some embodiments, the catalyst provided herein is selected from the group consisting of (+)-camphor-10-sulfonic acid; 2-pyridinesulfonic acid; 3-pyridinesulfonic acid; 8-hydroxy-5-quinolinesulfonic acid hydrate; α-hydroxy-2-pyridinemethanesulfonic acid; (β)-camphor-10-sulfonic acid; butylphosphonic acid; diphenylphosphinic acid; hexylphosphonic acid; methylphosphonic acid; phenylphosphinic acid; phenylphosphonic acid; tert-butylphosphonic acid; (SS)-VAPOL phosphate hydrate Element;6-Quinolinesulfonic acid, 3-(1-pyridinio)-1-propanesulfonate;2-(2-Pyridinyl)ethanesulfonic acid;3-(2-Pyridyl)-5,6-diphenyl-1,2,4-triazine-p,p'-disulfonic acid monosodium salt hydrate;1,1'-Binaphthyl-2,2'-diyl hydrogen phosphate;Bis(4-methoxyphenyl)phosphinic acid;Phenyl(3,5-xylyl)phosphinic acid;L-Cysteic acid monohydrate;Poly(styrenesulfonic acid-co-divinylbenzene);Lysine ;Ethanedisulfonic acid;Ethanesulfonic acid;Isethionic acid;Homocysteic acid;HEPBS (N-(2-hydroxyethyl)piperazine-N'-(4-butanesulfonic acid));HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid);2-Hydroxy-3-morpholinopropanesulfonic acid;2-(N-morpholino)ethanesulfonic acid;Methanesulfonic acid;Methaniazid;Naphthalene-1-sulfonic acid;Naphthalene-2-sulfonic acid;Perfluorobutanesulfonic acid;6- sulfoquinovose; trifluoromethanesulfonic acid; 2-aminoethanesulfonic acid; benzoic acid; chloroacetic acid; trifluoroacetic acid; caproic acid; enanthic acid; caprylic acid; pelargonic acid; lauric acid; palmitic acid; stearic acid; arachidic acid; aspartic acid; glutamic acid; serine; threonine; glutamine; cysteine; glycine; proline; alanine; valine; isoleucine; leucine; methionine; phenylalanine; tyrosine; tryptophan; or any combination thereof.

[0704]

[0740] In some embodiments, the catalyst provided herein is (+)-camphor-10-sulfonic acid. In some embodiments, the catalyst provided herein is 2-pyridinesulfonic acid. In some embodiments, the catalyst provided herein is 3-pyridinesulfonic acid. In some embodiments, the catalyst provided herein is 8-hydroxy-5-quinolinesulfonic acid hydrate. In some embodiments, the catalyst provided herein is α-hydroxy-2-pyridinemethanesulfonic acid. In some embodiments, the catalyst provided herein is (β)-camphor-10-sulfonic acid. In some embodiments, the catalyst provided herein is butylphosphonic acid. In some embodiments, the catalyst provided herein is diphenylphosphinic acid. In some embodiments, the catalyst provided herein is hexylphosphonic acid. In some embodiments, the catalyst provided herein is methylphosphonic acid. In some embodiments, the catalyst provided herein is phenylphosphinic acid. In some embodiments, the catalyst provided herein is phenylphosphonic acid. In some embodiments, the catalyst provided herein is tert-butylphosphonic acid. In some embodiments, the catalyst provided herein is (SS)-VAPOL hydrogen phosphate. In some embodiments, the catalyst provided herein is 6-quinolinesulfonic acid. In some embodiments, the catalyst provided herein is 3-(1-pyridinio)-1-propanesulfonate. In some embodiments, the catalyst provided herein is 2-(2-pyridinyl)ethanesulfonic acid. In some embodiments, the catalyst provided herein is 3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-p,p'-disulfonic acid monosodium salt hydrate. In some embodiments, the catalyst provided herein is 1,1'-binaphthyl-2,2'-diyl hydrogen phosphate. In some embodiments, the catalyst provided herein is bis(4-methoxyphenyl)phosphinic acid. In some embodiments, the catalyst provided herein is phenyl(3,5-xylyl)phosphinic acid.In some embodiments, the catalyst provided herein is L-cysteic acid monohydrate. In some embodiments, the catalyst provided herein is poly(styrenesulfonic acid-co-divinylbenzene). In some embodiments, the catalyst provided herein is lysine.

[0705]

[0741] In some embodiments, the catalyst is ethanedisulfonic acid. In some embodiments, the catalyst is ethanesulfonic acid. In some embodiments, the catalyst is isethionic acid. In some embodiments, the catalyst is homocysteic acid. In some embodiments, the catalyst is HEPBS (N-(2-hydroxyethyl)piperazine-N'-(4-butanesulfonic acid)). In some embodiments, the catalyst is HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid). In some embodiments, the catalyst is 2-hydroxy-3-morpholinopropanesulfonic acid. In some embodiments, the catalyst is 2-(N-morpholino)ethanesulfonic acid. In some embodiments, the catalyst is methanesulfonic acid. In some embodiments, the catalyst is naphthalene-1-sulfonic acid. In some embodiments, the catalyst is methanazid. In some embodiments, the catalyst is naphthalene-2-sulfonic acid. In some embodiments, the catalyst is perfluorobutanesulfonic acid. In some embodiments, the catalyst is 6-sulfoquinovose. In some embodiments, the catalyst is trifluoromethanesulfonic acid. In some embodiments, the catalyst is 2-aminoethanesulfonic acid. In some embodiments, the catalyst is benzoic acid. In some embodiments, the catalyst is chloroacetic acid. In some embodiments, the catalyst is trifluoroacetic acid. In some embodiments, the catalyst is caproic acid. In some embodiments, the catalyst is enanthic acid. In some embodiments, the catalyst is caprylic acid. In some embodiments, the catalyst is pelargonic acid. In some embodiments, the catalyst is lauric acid. In some embodiments, the catalyst is pamitic acid. In some embodiments, the catalyst is stearic acid. In some embodiments, the catalyst is arachidic acid. In some embodiments, the catalyst is aspartic acid. In some embodiments, the catalyst is glutamic acid. In some embodiments, the catalyst is serine. In some embodiments, the catalyst is threonine. In some embodiments, the catalyst is glutamine. In some embodiments, the catalyst is cysteine. In some embodiments, the catalyst is glycine.In some embodiments, the catalyst is proline. In some embodiments, the catalyst is alanine. In some embodiments, the catalyst is valine. In some embodiments, the catalyst is isoleucine. In some embodiments, the catalyst is leucine. In some embodiments, the catalyst is methionine. In some embodiments, the catalyst is phenylalanine. In some embodiments, the catalyst is tyrosine. In some embodiments, the catalyst is tryptophan.

[0706]

[0742] In some embodiments, the catalysts provided herein are polymeric catalysts or carbon-supported catalysts disclosed in WO2016122887, which is incorporated herein by reference in its entirety and for its disclosure.

[0707]

[0743] In some embodiments, the catalysts provided herein are present in an amount of about 0.01% to about 5%, about 0.02% to about 4%, about 0.03% to about 3%, or about 0.05% to about 2% by dry weight of one or more feed sugars. In some embodiments, the catalysts provided herein are present in an amount of about 1% to 2% by dry weight of one or more feed sugars. In some embodiments, the catalyst provided herein is present in an amount of about 0.5%, about 0.6%, about 0.7%, about 0.8%, about 0.9%, about 1.0%, about 1.1%, about 1.2%, about 1.3%, about 1.4%, about 1.5%, about 1.6%, about 1.7%, about 1.8%, about 1.9%, about 2.0%, about 2.1%, about 2.2%, about 2.3%, about 2.4%, about 2.5%, about 2.6%, about 2.7%, about 2.8%, about 2.9%, or about 3.0% by dry weight of the one or more feed sugars.

[0708]

[0744] In some embodiments, the catalysts provided herein are present in an amount of about 0.01% to about 5%, about 0.02% to about 4%, about 0.03% to about 3%, or about 0.05% to about 2% by dry weight of the aqueous composition. In some embodiments, the catalysts provided herein are present in an amount of about 1% to 2% by dry weight of the aqueous composition. In some embodiments, the catalyst provided herein is present in an amount of about 0.8%, about 0.9%, about 1.0%, about 1.1%, about 1.2%, about 1.3%, about 1.4%, about 1.5%, about 1.6%, about 1.7%, about 1.8%, about 1.9%, about 2.0%, about 2.1%, about 2.2%, about 2.3%, about 2.4%, about 2.5%, about 2.6%, about 2.7%, about 2.8%, about 2.9%, or about 3.0% by dry weight of the aqueous composition.

[0709]

[0745] In some embodiments, the catalysts provided herein are combinations of two or more different catalysts. In some embodiments, the catalysts include recyclable catalysts, such as resin and polymer catalysts, and non-recyclable catalysts. In some embodiments, when the catalyst includes at least two different catalysts, each of the catalysts is present in the amounts provided herein. In other embodiments, when the catalyst includes at least two different catalysts, the at least two different catalysts are present in an aggregate in the amounts provided herein.

[0710]

[0746] In some embodiments, the catalyst is added to the aqueous composition in a dry form. In other embodiments, the catalyst is added to the aqueous composition in a wet form, such as an aqueous solution. In some embodiments, the catalyst is combined with one or more feed sugars before adding water. In other embodiments, the catalyst is dissolved in water before combining with the one or more feed sugars. In some embodiments, the methods provided herein include producing an aqueous composition by combining one or more feed sugars in a dehydrated form and a catalyst in a wet form (e.g., an aqueous solution).

[0711] [C. Addition of Water]

[0747] In some embodiments, the method of producing an oligosaccharide preparation includes adding water to form an aqueous composition. In some embodiments, all or a portion of the water in the aqueous composition is added as free water. In other embodiments, all of the water in the aqueous composition is added as bound water, e.g., in a sugar monohydrate or dihydrate. In some embodiments, all of the water in the aqueous composition is added as bound water, e.g., in a monosaccharide monohydrate, such as glucose monohydrate. In certain embodiments, all or a portion of the water in the aqueous composition is added together with a catalyst, i.e., via a catalyst solution.

[0712] [D. Water content]

[0748] As the process for producing an oligosaccharide preparation progresses, water can be generated by reactions. For example, in some embodiments, water is generated (i) with the formation of glycosidic bonds, (ii) with the formation of anhydro subunits, or (iii) by other mechanisms or sources. Because both sugar condensation and dehydration reactions involve water, in some embodiments, the water content affects the composition of the oligosaccharide preparation.

[0713]

[0749] Furthermore, in some embodiments, the water content affects the viscosity of the aqueous composition, which in turn can affect the effectiveness of mixing of the aqueous composition. For example, in some embodiments, an overly viscous aqueous composition may result in undesirable uneven catalyst distribution in the aqueous composition. Moreover, in some embodiments, a very low water content may cause the aqueous composition to solidify, preventing effective mixing. On the other hand, in other embodiments, an extremely high water content may hinder the sugar condensation reaction and reduce the concentration of anhydrous subunits. Therefore, the present disclosure describes a water content suitable for producing oligosaccharide preparations.

[0714]

[0750] In some embodiments, the method of producing the oligosaccharide preparations described herein comprises forming and / or heating an aqueous composition. In some embodiments, the aqueous composition comprises from about 0% to about 80%, from about 0% to about 70%, from about 0% to about 60%, from about 0% to about 50%, from about 0% to about 40%, from about 0% to about 35%, from about 0% to about 30%, from about 0% to about 25%, from about 0% to about 20%, from about 0% to about 19%, from about 0% to about 18%, from about 0% to about 17%, from about 0% to about In some embodiments, the aqueous composition comprises about 1% to about 20%, about 1% to about 18%, about 1% to about 16%, about 1% to about 15%, about 0% to about 14%, about 0% to about 13%, about 0% to about 12%, about 0% to about 11%, about 0% to about 10%, about 0% to about 9%, about 0% to about 8%, about 0% to about 7%, about 0% to about 6%, about 0% to about 5%, about 0% to about 4%, about 0% to about 3%, about 0% to about 2%, or about 0% to about 1% water by total weight. In some embodiments, the aqueous composition comprises about 3% to about 16%, about 3% to about 14%, about 3% to about 12%, about 3% to about 10%, about 3% to about 8%, about 3% to about 6%, about 5% to about 16%, about 5% to about 14%, about 5% to about 12%, about 5% to about 10%, about 7% to about 16%, about 7% to about 14%, about 7% to about 12%, about 7% to about 10%, or about 8% to about 10% water by total weight. In some embodiments, the aqueous composition comprises about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, or about 15% water by total weight. In some embodiments, the aqueous composition comprises about 9% water by total weight. However, it should be understood that the amount of water in the aqueous composition can be adjusted based on the reaction conditions and the particular catalyst used. In some embodiments, the water content in the aqueous composition as disclosed above is measured at the beginning of the reaction, for example, before heating the sugar feed. In some embodiments, the water content in the aqueous composition as disclosed above is measured at the end of the polymerization or condensation reaction. In some embodiments, the water content in the aqueous composition as disclosed above is measured as the average water content at the beginning and end of the reaction.

[0715]

[0751] In certain embodiments, the methods described herein can further include monitoring the water content and / or the ratio of water to sugar or catalyst present in the aqueous composition over a period of time. In some embodiments, the method further includes removing at least a portion of the water in the aqueous composition, for example, by distillation. Water can be removed from the aqueous composition using any method known in the art, including, for example, vacuum filtration, vacuum distillation, heating, steam, hot air, and / or evaporation.

[0716]

[0752] In some embodiments, the oligosaccharide preparations described herein are hygroscopic. Thus, in some embodiments, the hygroscopicity of the feed sugars and oligosaccharides formed in the polymerization can affect the rate at which water can be removed from the aqueous composition.

[0717]

[0753] In some embodiments, the methods described herein include removing at least a portion of the water in the aqueous composition such that the water content in the aqueous composition is about 1% to about 20%, about 1% to about 18%, about 1% to about 16%, about 1% to about 14%, about 1% to about 12%, about 1% to about 10%, about 1% to about 8%, about 2% to about 16%, about 2% to about 14%, about 2% to about 12%, about 2% to about 10%, about 2% to about 8%, about 2% to about 6%, about 4% to about 16%, about 4% to about 14%, about 4% to about 12%, about 4% to about 10%, about 4% to about 8%, about 6% to about 16%, about 6% to about 12%, about 6% to about 10%, or about 6% to about 8%, by total weight. In some embodiments, the method comprises removing at least a portion of the water from the aqueous composition such that the water content of the aqueous composition is about 2% to about 10%, about 2% to about 8%, or about 4% to about 8% by total weight. In some embodiments, the method comprises removing at least a portion of the water from the aqueous composition such that the water content of the aqueous composition is about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% by total weight. In some embodiments, the method comprises removing at least a portion of the water from the aqueous composition such that the water content of the aqueous composition is about 4% to about 8% by total weight. In some embodiments, the method comprises removing at least a portion of the water in the aqueous composition such that the water content of the aqueous composition at the end of the polymerization and / or condensation reaction is as disclosed above. In some embodiments, the method comprises removing at least a portion of the water in the aqueous composition such that the water content of the aqueous composition at the start of the polymerization and / or condensation reaction is as disclosed above. In some embodiments, the method comprises removing at least a portion of the water in the aqueous composition such that the average water content in the aqueous composition at the beginning and end of the polymerization and / or condensation reaction is within the ranges disclosed above. In some embodiments, the method comprises removing at least a portion of the water in the aqueous composition throughout the polymerization and / or condensation reaction such that the water content in the aqueous composition remains within the ranges disclosed above.

[0718]

[0754] In some embodiments, the methods described herein comprise adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition is about 1% to about 20%, about 1% to about 18%, about 1% to about 16%, about 1% to about 14%, about 1% to about 12%, about 1% to about 10%, about 1% to about 8%, about 2% to about 16%, about 2% to about 14%, about 2% to about 12%, about 2% to about 10%, about 2% to about 8%, about 2% to about 6%, about 4% to about 16%, about 4% to about 14%, about 4% to about 12%, about 4% to about 10%, about 4% to about 8%, about 6% to about 16%, about 6% to about 12%, about 6% to about 10%, or about 6% to about 8%, by total weight. In some embodiments, the method comprises adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition is about 2% to about 10%, about 2% to about 8%, or about 4% to about 8% by total weight. In some embodiments, the method comprises adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition is about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, or about 10% by total weight. In some embodiments, the method comprises adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition is about 4% to about 8% by total weight. In some embodiments, the method comprises adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition at the end of the polymerization and / or condensation reaction is as disclosed above. In some embodiments, the method comprises adding at least a portion of water to the aqueous composition such that the water content of the aqueous composition at the start of the polymerization and / or condensation reaction is as disclosed above. In some embodiments, the method comprises adding at least a portion of the water to the aqueous composition such that the average water content in the aqueous composition at the beginning and end of the polymerization and / or condensation reaction is within the ranges disclosed above. In some embodiments, the method comprises adding at least a portion of the water to the aqueous composition throughout the polymerization and / or condensation reaction such that the water content in the aqueous composition remains within the ranges disclosed above.

[0719]

[0755] In some embodiments, the degree of polymerization of the oligosaccharides and / or the amount and type of anhydro subunits in the oligosaccharide preparation can be adjusted by adjusting or controlling the water content present in the aqueous composition throughout the manufacturing process. For example, in some embodiments, the degree of polymerization of the oligosaccharides and / or the amount and type of anhydro subunits is increased by decreasing the water content.

[0720]

[0756] Thus, in some embodiments, the methods described herein include in-process control (IPC) of water content, which may include monitoring the water content, maintaining the water content, increasing the water content, decreasing the water content, or any combination thereof. In some embodiments, the IPC process includes maintaining the water content while the aqueous composition is heated to a temperature described herein. In some embodiments, the method includes maintaining the water content for a time sufficient to induce polymerization. In some embodiments, the method includes maintaining the water content within the disclosed ranges by adding water or removing water from the aqueous composition, or both. In some embodiments, the method includes maintaining the water content within the disclosed ranges by distillation. In some embodiments, the method includes maintaining the water content within the disclosed ranges by vacuum distillation. In some embodiments, the method includes maintaining the water content within the disclosed ranges by distillation at atmospheric pressure.

[0721]

[0757] In some embodiments, the water content of the aqueous composition is maintained within the range of about 1% to about 20%, about 1% to about 18%, about 1% to about 16%, about 1% to about 14%, about 1% to about 12%, about 1% to about 10%, about 1% to about 8%, about 2% to about 16%, about 2% to about 14%, about 2% to about 12%, about 2% to about 10%, about 2% to about 8%, about 2% to about 6%, about 4% to about 16%, about 4% to about 14%, about 4% to about 12%, about 4% to about 10%, about 4% to about 8%, about 6% to about 16%, about 6% to about 12%, about 6% to about 10%, or about 6% to about 8% by total weight. In some embodiments, the water content of the aqueous composition is maintained within a range of about 2% to about 10%, about 2% to about 8%, or about 4% to about 8% by total weight. In some embodiments, the water content of the aqueous composition is maintained within a range of about 2% to about 8% by total weight.

[0722]

[0758] The water content of an aqueous composition can be determined by various analytical methods and instruments. In some embodiments, the water content is determined by evaporation methods (e.g., loss on drying), distillation methods, or chemical reaction methods (e.g., Karl Fischer titration). In some embodiments, the water content is determined by analytical instruments such as a moisture meter. In some embodiments, the water content is determined by Karl Fischer titration.

[0723]

[0759] In some embodiments, the water content of the aqueous composition is measured during the reaction and used to implement in-process control (IPC) of the water content. In certain embodiments, the water content of the reaction is measured by Karl Fischer titration, infrared spectroscopy, near-infrared spectroscopy, conductivity, viscosity, density, torque mixing, or energy mixing. In some embodiments, the measurement of the water content of the reaction is used to control devices that actively regulate the water content of the reaction, such as a water addition pump or a flow valve.

[0724]

[0760] Without being bound by theory, it is believed that the water content during the sugar polymerization and / or condensation reaction can affect the concentration of anhydro subunits in the oligosaccharide preparations described herein. For example, as shown in Figure 21, in some embodiments, higher water content correlates with lower concentrations of anhydro subunits. In some embodiments, lower reaction temperatures can correlate with lower concentrations of anhydro subunits.

[0725] [E. Temperature]

[0761] In some embodiments, the degree of polymerization of the oligosaccharides and / or the amount and type of anhydro subunits in the oligosaccharide preparation can be controlled by adjusting the temperature to which the aqueous composition is heated. In some embodiments, methods for producing the oligosaccharide preparations described herein include heating the aqueous composition to a temperature of about 80°C to about 250°C, about 90°C to about 200°C, about 100°C to about 200°C, about 100°C to about 180°C, about 110°C to about 170°C, about 120°C to about 160°C, about 130°C to about 150°C, or about 135°C to about 145°C. In some embodiments, methods for producing the oligosaccharide preparations include heating the...

Claims

1. 1. A method for improving nitrogen utilization in an animal (excluding a human), comprising: administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharide in a relative abundance of 0.5% to 15% as determined by mass spectrometry; levels of multiple metabolites associated with improved nitrogen utilization are higher in gastrointestinal samples from the animals compared to gastrointestinal samples from comparable control animals administered an equivalent nutritional composition comprising the basal nutritional composition and not comprising the synthetic oligosaccharide preparation; This includes: The metabolites include (2S,3S)-3-methylaspartate, (R)-3-(phenyl)lactate, (R)-3-(phenyl)lactoyl-CoA, (S)-3-aminobutanoyl-CoA, 3-(4-hydroxyphenyl)pyruvate, 4-aminobutanal, 4-aminobutanoate, 4-guanidinobutanoate, 4-guanidinobutraldehyde, 4-guanidobutriamide, 4-maleyl-acetoacetate, 5-aminopentanal, 5-aminopentanoate, 5-guanidino-2-oxopentanoate, agmatine, and cadaverine. , cinnamate, cinnamoyl-CoA, coenzyme A, formamide, homogentisate, L-β-lysine, L-cystathionine, L-glutamate-5-semialdehyde, L-histidine, L-homocysteine, L-lysine, L-methionine, L-ornithine, L-proline, L-serine, mesaconate, N-carbamoylputrescine, N2-succinyl-L-arginine, succinate semialdehyde, succinyl-CoA, or urea.

2. 1. A method for improving carbon utilization in an animal (excluding a human), comprising: administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharide in a relative abundance of 0.5% to 15% as determined by mass spectrometry; The levels of multiple metabolites associated with improved carbon utilization in gastrointestinal samples from the animals are higher compared to matched control animals administered an equivalent nutritional composition comprising the basal nutritional composition and not comprising the synthetic oligosaccharide preparation. This includes:

10. The method of claim 1, wherein the plurality of metabolites comprises at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 metabolites selected from the group consisting of (R)-lactate, (R)-lactoyl-CoA, (S)-lactate, (S)-propane-1,2-diol, 1-propanal, acetate, acetyl-CoA, acryloyl-CoA, propanoate, propanoyl-CoA, and pyruvate.

3. 1. A method for enhancing antibacterial and / or anti-inflammatory responses in an animal (but excluding humans), comprising: administering to the animal a nutritional composition comprising a basal nutritional composition and a synthetic oligosaccharide preparation, the synthetic oligosaccharide preparation comprises at least n fractions of oligosaccharides (DP1 to DPn fractions), each having a different degree of polymerization selected from 1 to n, where n is an integer greater than 3, and each of the DP1 and DP2 fractions independently comprises an anhydro-subunit-containing oligosaccharide in a relative abundance of 0.5% to 15% as determined by mass spectrometry; The levels of metabolites associated with improved antibacterial and anti-inflammatory responses in gastrointestinal samples from the animals are higher compared to matched control animals administered an equivalent nutritional composition comprising the basal nutritional composition and not comprising the synthetic oligosaccharide preparation. This includes: The method, wherein the metabolite is itaconate.

4. 4. The method of claim 3, wherein the level of the metabolite is at least 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15%, 20%, or 25% higher than the level in a gastrointestinal sample from an equivalent control animal administered an equivalent nutritional composition containing the basal nutritional composition but not the synthetic oligosaccharide preparation.

Images