# US 11,297,865 B2 · Methods of treating biomass to produce oligosaccharides and related compositions > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 11,297,865 B2 - Document ID: US11297865B2 - Status: Granted - Kind: B2 - Application number: 17/083,121 - Filed: 2020-10-28 - Published: 2021-12-23 - Granted: 2022-04-12 - Earliest claimed: 2019-08-16 - Assignees: Cambridge Glycoscience Ltd - Applicants: Cambridge Glycoscience Ltd - Inventors: Thomas J. Simmons; Jeremy Bartosiak-Jentys; Aleksandra Wlodek; Gustavo Valente - Web page: https://patentgrep.com/patent/US11297865B2 ## CPC - A23L 29/30: containing carbohydrate syrups; containing sugars; containing sugar alcohols, e.g. xylitol; containing starch hydrolysates, e.g. dextrin (products from apiculture A23L21/20; artificial sweetening agents A23L27/30) - A23L 7/126: Snacks or the like obtained by binding, shaping or compacting together cereal grains or cereal pieces, e.g. cereal bars - A23L 7/135: Individual or non-extruded flakes, granules or shapes having similar size, e.g. breakfast cereals - A23L 27/33: {containing sugars or derivatives} - A23L 27/63: {Ketchup} - A23L 29/206: of vegetable origin - A23L 29/262: Cellulose; Derivatives thereof, e.g. ethers - A23L 33/21: Addition of substantially indigestible substances, e.g. dietary fibres - A23L 33/24: Cellulose or derivatives thereof - C08B 37/0057: {beta-D-Xylans, i.e. xylosaccharide, e.g. arabinoxylan, arabinofuronan, pentosans; (beta-1,3)(beta-1,4)-D-Xylans, e.g. rhodymenans; Hemicellulose; Derivatives thereof} - C08H 8/00: Macromolecular compounds derived from lignocellulosic materials {(pretreatment thereof B27N)} - C08L 5/00: Compositions of polysaccharides or of their derivatives not provided for in groups C08L1/00 or C08L3/00 - C12P 19/02: Monosaccharides (2-ketogulonic acid C12P7/60) - C12P 19/04: Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds - C12P 19/12: Disaccharides - C12P 19/14: produced by the action of a carbohydrase {(EC 3.2.x)}, e.g. by alpha-amylase {, e.g. by cellulase, hemicellulase} - A23V 2002/00: Food compositions, function of food ingredients or processes for food or foodstuffs - A23L: FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF (preservation thereof A23B) - C08B: POLYSACCHARIDES; DERIVATIVES THEREOF (polysaccharides containing less than six saccharide radicals attached to each other by glycosidic linkages C07H; fermentation or enzyme-using processes C12P19/00; sugar industry C13; production of cellulose D21) - C08H: DERIVATIVES OF NATURAL MACROMOLECULAR COMPOUNDS (polysaccharides C08B; natural rubber C08C; natural resins or their derivatives C09F; bituminous materials C10) - C08L: COMPOSITIONS OF MACROMOLECULAR COMPOUNDS (compositions based on polymerisable monomers C08F, C08G; artificial filaments or fibres D01F; textile treating compositions D06) - C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)} - A23V: INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION ## Abstract Methods of forming an ingredient for human consumption are provided herein. The methods may include isolating one or more soluble polysaccharides from a biomass, generating one or more oligosaccharides from the biomass, and combining the one or more isolated soluble polysaccharides with the generated oligosaccharides to form the ingredient. Methods of pretreating a biomass are also provided. The methods may include administering a physical pretreatment to a biomass, administering a gentle pretreatment to the physically pretreated biomass, and administering a strong pretreatment to the gently pretreated biomass. Ingredients for human consumption are also provided. ## Claims ### Claim 1 A method for producing an ingredient for human consumption, the method comprising: (a) subjecting a biomass to a first pretreatment to reduce an average size of the biomass to produce a pretreated biomass; (b) subjecting the pretreated biomass to a second pretreatment comprising: (i) incubating the pretreated biomass in an aqueous solution to solubilize monosaccharides and/or disaccharides from the pretreated biomass; and (ii) removing a portion of the aqueous solution comprising solubilized monosaccharides and/or disaccharides to produce a second pretreated biomass; (c) subjecting the second pretreated biomass to a third pretreatment to increase a digestibility of the biomass to produce a third pretreated biomass, wherein the third pretreatment is performed after the second pretreatment; (d) contacting, in a solution or a suspension, one or more polysaccharide-cleaving enzymes and the third pretreated biomass to form one or more oligosaccharides; and (e) enriching the solution or suspension to increase a concentration of the one or more oligosaccharides to form the ingredient. ### Claim 2 (depends on 1) The method of claim 1, further comprising removing at least 25% vol/vol of the aqueous solution comprising the solubilized monosaccharides and/or disaccharides at (b)(ii). ### Claim 3 (depends on 1) The method of claim 1, wherein (b) and/or (c) is performed at least two or more times. ### Claim 4 (depends on 1) The method of claim 1, wherein the third pretreatment is a thermochemical treatment comprising incubating the gently pre-treated biomass in an alkali solution at a temperature of 50° C. to 150° C. ### Claim 5 (depends on 4) The method of claim 4, wherein the thermochemical treatment is conducted from 5 minutes to 72 hours. ### Claim 6 (depends on 1) The method of claim 1, wherein the solution or suspension comprising the strongly pretreated biomass after (c) comprises less than 10% w/w by dry weight monosaccharides. ### Claim 7 (depends on 1) The method of claim 1, further comprising purifying the one or more oligosaccharides from the solution or suspension. ### Claim 8 (depends on 1) The method of claim 1, wherein the portion of the solubilized monosaccharides and/or disaccharides removed in (b) is not combined with the one or more oligosaccharides of (e) to form the ingredient. ### Claim 9 (depends on 1) The method of claim 1, wherein the ingredient comprises less than 15% by dry weight monosaccharides. ### Claim 10 (depends on 1) The method of claim 1, wherein the ingredient comprises less than 60% by dry weight disaccharides. ### Claim 11 (depends on 1) The method of claim 1, wherein the ingredient is substantially free of monosaccharides. ### Claim 12 (depends on 1) The method of claim 1, wherein the one or more oligosaccharides comprise at least one of: i) a cello-oligosaccharide having a degree of polymerization (DP) of from two to six; ii) a xylo-oligosaccharide having a DP of from two to twelve; iii) an arabinoxylo-oligosaccharide having a DP of from three to fifteen; iv) a manno-oligosaccharide having a DP of from two to twelve; v) a mixed-linkage glucan oligosaccharide having a DP of from two to five; vi) a xyloglucan oligosaccharide having a DP of from four to twelve; or vii) a chito-oligosaccharide having a DP of from two to twelve. ### Claim 13 (depends on 12) The method of claim 12, wherein the ingredient comprises at least two of the oligosaccharides selected from (i) to (vii). ### Claim 14 (depends on 1) The method of claim 1, wherein the ingredient comprises at least 5% w/w of the one or more oligosaccharides. ### Claim 15 (depends on 12) The method of claim 12, wherein the ingredient comprises at least one of sucrose, maltose, lactose, glucose, fructose, or galactose at a concentration less than 50% dry w/w of the total dry w/w of the one or more oligosaccharides. ### Claim 16 (depends on 1) The method of claim 1, wherein the monosaccharides and/or disaccharides comprise at least one of sucrose, maltose, lactose, glucose, fructose, or galactose. ### Claim 17 (depends on 1) The method of claim 1, wherein (b) further comprises solubilizing one or more organic acids. ### Claim 18 (depends on 17) The method of claim 17, wherein the one or more organic acids comprises at least one of oxalate, tartrate, succinate, formate, citrate, malate, lactate, or acetate. ### Claim 19 (depends on 18) The method of claim 18, wherein the total weight of the one or more organic acids is greater than 10% of the total dry weight of the monosaccharides and/or disaccharides in the portion removed in (b)(ii). ### Claim 20 (depends on 1) The method of claim 1, wherein the first pretreatment of (a) comprises at least one of chipping, chopping, milling, ball-milling, grinding, sprucing, or blending the biomass. ### Claim 21 (depends on 1) The method of claim 1, wherein the second pretreatment of (b) occurs in an aqueous solution comprising water. ### Claim 22 (depends on 1) The method of claim 1, wherein the second pretreatment of (b) occurs at a temperature of from 5° C. to 150° C. ### Claim 23 (depends on 22) The method of claim 22, wherein the second pretreatment of (b) is conducted from 15 minutes to 1 hour. ### Claim 24 (depends on 1) The method of claim 1, wherein (c) comprises treating the gently pretreated biomass in an alkali solution having a pH from 8 to 14. ### Claim 25 (depends on 24) The method of claim 24, wherein the alkali solution comprises at least one of sodium hydroxide, potassium hydroxide, sodium carbonate, calcium carbonate, aqueous ammonia, ammonium sulfate, or ammonium hydroxide. ### Claim 26 (depends on 1) The method of claim 1, wherein the biomass is a plant biomass comprising at least one of a grain, sugar cane, corn, wheat, rice, miscanthus, sorghum residue, switch grass, bamboo, monocotyledonous tissue, hardwood tissue, or softwood tissue. ### Claim 27 (depends on 1) The method of claim 1, wherein the biomass comprises at least one of cellulose, chitin, chitosan, xylan, xyloglucan, mixed-linkage glucan, mannan, arabinoxylo-oligosaccharides, or lignocellulose. ### Claim 28 (depends on 1) The method of claim 1, wherein the one or more polysaccharide-cleaving enzymes comprises at least one of cellulase, xylanase, xyloglucanase, endo-glucanase, cellobiohydrolase, mannanase, lichenase, or lytic polysaccharide monooxygenase (LPMO). ### Claim 29 (depends on 1) The method of claim 1, wherein the one or more of the polysaccharide-cleaving enzymes is prepared from a filamentous fungus. ### Claim 30 (depends on 1) The method of claim 1, further comprising incorporating the ingredient into at least one of a baked good, a chocolate, a confectionery, or a drink.