AU2012251903A1 - Process for the production of isomaltulose from plant juices - Google Patents
Process for the production of isomaltulose from plant juices Download PDFInfo
- Publication number
- AU2012251903A1 AU2012251903A1 AU2012251903A AU2012251903A AU2012251903A1 AU 2012251903 A1 AU2012251903 A1 AU 2012251903A1 AU 2012251903 A AU2012251903 A AU 2012251903A AU 2012251903 A AU2012251903 A AU 2012251903A AU 2012251903 A1 AU2012251903 A1 AU 2012251903A1
- Authority
- AU
- Australia
- Prior art keywords
- strain
- klebsiella
- seq
- sucrose
- isomaltulose
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
Links
- 238000000034 method Methods 0.000 title claims abstract description 54
- 235000011389 fruit/vegetable juice Nutrition 0.000 title claims abstract description 51
- PVXPPJIGRGXGCY-TZLCEDOOSA-N 6-O-alpha-D-glucopyranosyl-D-fructofuranose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1OC[C@@H]1[C@@H](O)[C@H](O)C(O)(CO)O1 PVXPPJIGRGXGCY-TZLCEDOOSA-N 0.000 title claims abstract description 35
- 238000004519 manufacturing process Methods 0.000 title claims description 15
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims abstract description 66
- 229930006000 Sucrose Natural products 0.000 claims abstract description 66
- 239000005720 sucrose Substances 0.000 claims abstract description 65
- 108090000790 Enzymes Proteins 0.000 claims abstract description 37
- 102000004190 Enzymes Human genes 0.000 claims abstract description 37
- 108010093096 Immobilized Enzymes Proteins 0.000 claims abstract description 10
- 238000006243 chemical reaction Methods 0.000 claims abstract description 7
- 241001622809 Serratia plymuthica Species 0.000 claims description 28
- 241000556426 Erwinia rhapontici Species 0.000 claims description 13
- 241000588754 Klebsiella sp. Species 0.000 claims description 9
- 241000192130 Leuconostoc mesenteroides Species 0.000 claims description 9
- 241000607715 Serratia marcescens Species 0.000 claims description 9
- 239000000203 mixture Substances 0.000 claims description 9
- 241000589155 Agrobacterium tumefaciens Species 0.000 claims description 8
- 241000588699 Erwinia sp. Species 0.000 claims description 8
- 241001014264 Klebsiella variicola Species 0.000 claims description 8
- 241000611870 Pantoea dispersa Species 0.000 claims description 8
- 241000588746 Raoultella planticola Species 0.000 claims description 8
- 241000588756 Raoultella terrigena Species 0.000 claims description 8
- 241000083686 [Pseudomonas] mesoacidophila Species 0.000 claims description 8
- 241000147019 Enterobacter sp. Species 0.000 claims description 7
- 241000586779 Protaminobacter Species 0.000 claims description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 claims description 6
- 229940072056 alginate Drugs 0.000 claims description 6
- 235000010443 alginic acid Nutrition 0.000 claims description 6
- 229920000615 alginic acid Polymers 0.000 claims description 6
- 241000607694 Serratia odorifera Species 0.000 claims description 5
- 241001631439 Azotobacter vinelandii DJ Species 0.000 claims description 4
- 108010010525 Isomaltulose synthase Proteins 0.000 claims description 4
- 241000588747 Klebsiella pneumoniae Species 0.000 claims description 4
- 241000376410 Klebsiella pneumoniae 342 Species 0.000 claims description 3
- 241001278462 Serratia plymuthica 4Rx13 Species 0.000 claims description 3
- 150000004676 glycans Chemical class 0.000 claims description 3
- 229920001282 polysaccharide Polymers 0.000 claims description 3
- 239000005017 polysaccharide Substances 0.000 claims description 3
- 239000007787 solid Substances 0.000 claims description 3
- 229920001661 Chitosan Polymers 0.000 claims description 2
- 241000607720 Serratia Species 0.000 claims description 2
- 239000000679 carrageenan Substances 0.000 claims description 2
- 235000010418 carrageenan Nutrition 0.000 claims description 2
- 229920001525 carrageenan Polymers 0.000 claims description 2
- 229940113118 carrageenan Drugs 0.000 claims description 2
- 229940045110 chitosan Drugs 0.000 claims description 2
- 235000010987 pectin Nutrition 0.000 claims description 2
- 239000001814 pectin Substances 0.000 claims description 2
- 229920001277 pectin Polymers 0.000 claims description 2
- 229960000292 pectin Drugs 0.000 claims description 2
- 229920002451 polyvinyl alcohol Polymers 0.000 claims description 2
- 235000019422 polyvinyl alcohol Nutrition 0.000 claims description 2
- UHVMMEOXYDMDKI-JKYCWFKZSA-L zinc;1-(5-cyanopyridin-2-yl)-3-[(1s,2s)-2-(6-fluoro-2-hydroxy-3-propanoylphenyl)cyclopropyl]urea;diacetate Chemical compound [Zn+2].CC([O-])=O.CC([O-])=O.CCC(=O)C1=CC=C(F)C([C@H]2[C@H](C2)NC(=O)NC=2N=CC(=CC=2)C#N)=C1O UHVMMEOXYDMDKI-JKYCWFKZSA-L 0.000 claims description 2
- 241000588748 Klebsiella Species 0.000 claims 3
- 238000006317 isomerization reaction Methods 0.000 abstract description 12
- 238000000746 purification Methods 0.000 abstract description 6
- 229960004793 sucrose Drugs 0.000 description 54
- 239000000243 solution Substances 0.000 description 28
- 230000001580 bacterial effect Effects 0.000 description 9
- 210000004027 cell Anatomy 0.000 description 9
- 241000196324 Embryophyta Species 0.000 description 8
- 210000001822 immobilized cell Anatomy 0.000 description 8
- 239000000758 substrate Substances 0.000 description 7
- 241000219310 Beta vulgaris subsp. vulgaris Species 0.000 description 6
- 235000021536 Sugar beet Nutrition 0.000 description 6
- 239000000126 substance Substances 0.000 description 5
- 229920002684 Sepharose Polymers 0.000 description 4
- 230000002255 enzymatic effect Effects 0.000 description 4
- 238000000855 fermentation Methods 0.000 description 4
- 230000004151 fermentation Effects 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- 235000013379 molasses Nutrition 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 235000000346 sugar Nutrition 0.000 description 4
- 230000007306 turnover Effects 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 239000000499 gel Substances 0.000 description 3
- 238000004128 high performance liquid chromatography Methods 0.000 description 3
- 235000015097 nutrients Nutrition 0.000 description 3
- 238000002360 preparation method Methods 0.000 description 3
- 239000001117 sulphuric acid Substances 0.000 description 3
- 235000011149 sulphuric acid Nutrition 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 2
- 229930091371 Fructose Natural products 0.000 description 2
- 239000005715 Fructose Substances 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 240000008042 Zea mays Species 0.000 description 2
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 2
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 2
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 description 2
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 2
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 2
- 239000001166 ammonium sulphate Substances 0.000 description 2
- 235000011130 ammonium sulphate Nutrition 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
- 239000011324 bead Substances 0.000 description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 2
- 239000011942 biocatalyst Substances 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 230000001013 cariogenic effect Effects 0.000 description 2
- 235000005822 corn Nutrition 0.000 description 2
- 238000002425 crystallisation Methods 0.000 description 2
- 230000008025 crystallization Effects 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 238000012262 fermentative production Methods 0.000 description 2
- 102000037865 fusion proteins Human genes 0.000 description 2
- 108020001507 fusion proteins Proteins 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- AJVRSHNXSHMMCH-UHFFFAOYSA-K iron(III) citrate monohydrate Chemical compound O.[Fe+3].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O AJVRSHNXSHMMCH-UHFFFAOYSA-K 0.000 description 2
- CKAODHQJQJOTCB-UHFFFAOYSA-L magnesium;dichloride;heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[Cl-].[Cl-] CKAODHQJQJOTCB-UHFFFAOYSA-L 0.000 description 2
- CNFDGXZLMLFIJV-UHFFFAOYSA-L manganese(II) chloride tetrahydrate Chemical compound O.O.O.O.[Cl-].[Cl-].[Mn+2] CNFDGXZLMLFIJV-UHFFFAOYSA-L 0.000 description 2
- 150000002772 monosaccharides Chemical class 0.000 description 2
- 229920001184 polypeptide Polymers 0.000 description 2
- ILRLTAZWFOQHRT-UHFFFAOYSA-N potassium;sulfuric acid Chemical compound [K].OS(O)(=O)=O ILRLTAZWFOQHRT-UHFFFAOYSA-N 0.000 description 2
- 108090000765 processed proteins & peptides Proteins 0.000 description 2
- 102000004196 processed proteins & peptides Human genes 0.000 description 2
- 239000008399 tap water Substances 0.000 description 2
- 235000020679 tap water Nutrition 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- RJPPRBMGVWEZRR-WTZPKTTFSA-N (3s,4r,5r)-1,3,4,5-tetrahydroxy-6-[(2s,3r,4s,5s,6r)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxyhexan-2-one Chemical compound OC[C@H]1O[C@H](OC[C@@H](O)[C@@H](O)[C@H](O)C(=O)CO)[C@H](O)[C@@H](O)[C@@H]1O RJPPRBMGVWEZRR-WTZPKTTFSA-N 0.000 description 1
- 125000003903 2-propenyl group Chemical group [H]C([*])([H])C([H])=C([H])[H] 0.000 description 1
- 229920000178 Acrylic resin Polymers 0.000 description 1
- 239000004925 Acrylic resin Substances 0.000 description 1
- 229920000936 Agarose Polymers 0.000 description 1
- 108010011170 Ala-Trp-Arg-His-Pro-Gln-Phe-Gly-Gly Proteins 0.000 description 1
- KWIUHFFTVRNATP-UHFFFAOYSA-N Betaine Natural products C[N+](C)(C)CC([O-])=O KWIUHFFTVRNATP-UHFFFAOYSA-N 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- 210000004128 D cell Anatomy 0.000 description 1
- 239000004593 Epoxy Substances 0.000 description 1
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 description 1
- 102220547770 Inducible T-cell costimulator_A23L_mutation Human genes 0.000 description 1
- 241000192132 Leuconostoc Species 0.000 description 1
- KWIUHFFTVRNATP-UHFFFAOYSA-O N,N,N-trimethylglycinium Chemical compound C[N+](C)(C)CC(O)=O KWIUHFFTVRNATP-UHFFFAOYSA-O 0.000 description 1
- 108010073038 Penicillin Amidase Proteins 0.000 description 1
- 239000004952 Polyamide Substances 0.000 description 1
- 229920002873 Polyethylenimine Polymers 0.000 description 1
- 239000004793 Polystyrene Substances 0.000 description 1
- 240000000111 Saccharum officinarum Species 0.000 description 1
- 235000007201 Saccharum officinarum Nutrition 0.000 description 1
- VJHCJDRQFCCTHL-UHFFFAOYSA-N acetic acid 2,3,4,5,6-pentahydroxyhexanal Chemical compound CC(O)=O.OCC(O)C(O)C(O)C(O)C=O VJHCJDRQFCCTHL-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 229960003237 betaine Drugs 0.000 description 1
- 238000010364 biochemical engineering Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 230000036983 biotransformation Effects 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 239000007853 buffer solution Substances 0.000 description 1
- 239000000648 calcium alginate Substances 0.000 description 1
- 235000010410 calcium alginate Nutrition 0.000 description 1
- 229960002681 calcium alginate Drugs 0.000 description 1
- OKHHGHGGPDJQHR-YMOPUZKJSA-L calcium;(2s,3s,4s,5s,6r)-6-[(2r,3s,4r,5s,6r)-2-carboxy-6-[(2r,3s,4r,5s,6r)-2-carboxylato-4,5,6-trihydroxyoxan-3-yl]oxy-4,5-dihydroxyoxan-3-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylate Chemical compound [Ca+2].O[C@@H]1[C@H](O)[C@H](O)O[C@@H](C([O-])=O)[C@H]1O[C@H]1[C@@H](O)[C@@H](O)[C@H](O[C@H]2[C@H]([C@@H](O)[C@H](O)[C@H](O2)C([O-])=O)O)[C@H](C(O)=O)O1 OKHHGHGGPDJQHR-YMOPUZKJSA-L 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 239000003054 catalyst Substances 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- MYSWGUAQZAJSOK-UHFFFAOYSA-N ciprofloxacin Chemical compound C12=CC(N3CCNCC3)=C(F)C=C2C(=O)C(C(=O)O)=CN1C1CC1 MYSWGUAQZAJSOK-UHFFFAOYSA-N 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- ATDGTVJJHBUTRL-UHFFFAOYSA-N cyanogen bromide Chemical compound BrC#N ATDGTVJJHBUTRL-UHFFFAOYSA-N 0.000 description 1
- MNNHAPBLZZVQHP-UHFFFAOYSA-N diammonium hydrogen phosphate Chemical compound [NH4+].[NH4+].OP([O-])([O-])=O MNNHAPBLZZVQHP-UHFFFAOYSA-N 0.000 description 1
- 235000013681 dietary sucrose Nutrition 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000005538 encapsulation Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000006459 hydrosilylation reaction Methods 0.000 description 1
- 229910052738 indium Inorganic materials 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 210000004379 membrane Anatomy 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 150000007522 mineralic acids Chemical class 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 229920000058 polyacrylate Polymers 0.000 description 1
- 229920002647 polyamide Polymers 0.000 description 1
- 229920000728 polyester Polymers 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 229920000193 polymethacrylate Polymers 0.000 description 1
- 229920002223 polystyrene Polymers 0.000 description 1
- 239000013587 production medium Substances 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- LIVNPJMFVYWSIS-UHFFFAOYSA-N silicon monoxide Chemical class [Si-]#[O+] LIVNPJMFVYWSIS-UHFFFAOYSA-N 0.000 description 1
- 229910052814 silicon oxide Inorganic materials 0.000 description 1
- 239000004447 silicone coating Substances 0.000 description 1
- 230000000087 stabilizing effect Effects 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 229920005613 synthetic organic polymer Polymers 0.000 description 1
- 150000003573 thiols Chemical class 0.000 description 1
- 125000000391 vinyl group Chemical group [H]C([*])=C([H])[H] 0.000 description 1
- 229920002554 vinyl polymer Polymers 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/24—Preparation of compounds containing saccharide radicals produced by the action of an isomerase, e.g. fructose
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L27/00—Spices; Flavouring agents or condiments; Artificial sweetening agents; Table salts; Dietetic salt substitutes; Preparation or treatment thereof
- A23L27/30—Artificial sweetening agents
- A23L27/33—Artificial sweetening agents containing sugars or derivatives
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L29/00—Foods or foodstuffs containing additives; Preparation or treatment thereof
- A23L29/30—Foods or foodstuffs containing additives; Preparation or treatment thereof containing carbohydrate syrups; containing sugars; containing sugar alcohols, e.g. xylitol; containing starch hydrolysates, e.g. dextrin
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/12—Disaccharides
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Polymers & Plastics (AREA)
- Food Science & Technology (AREA)
- Nutrition Science (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Molecular Biology (AREA)
- Botany (AREA)
- Mycology (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
- Enzymes And Modification Thereof (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Non-Alcoholic Beverages (AREA)
Abstract
The present invention relates to a process for producing isomaltulose, which comprises the process steps: A) bringing an immobilized enzyme complex capable of catalysing the conversion of sucrose into isomaltulose into contact with a sucrose-containing solution; B) isomerization of at least some of the sucrose to give isomaltulose; C) removing the enzyme complex, and if appropriate D) further purification of the isomaltulose, characterized in that the sucrose-containing solution used is at least one selected from the series consisting of thick juice or thin juice, preferably thick juice.
Description
(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date W I PO I PC T WO 2012/150051 Al 8 November 2012 (08.11.2012) (51) International Patent Classification: (81) Designated states (unless otherwise indicated, for every A23L 1/236 (2006.01) C12P 19/24 (2006.01) kind of national protection available): AE, AG, AL, AM, C12P 19/12(2006.01) AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, DO, (21) International Application Number: PCT/EP2012/052096 DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, (22) International Filing Date: 8 February 2012 (08.02.2012) KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, (25) Filing Language: German NZ, OM, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, (26) Publication Language: German TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (30) Priority Data: 10 2011 100 772.9 5 May 2011 (05.05.2011) DE (84) Designated states (unless otherwise indicated, for every kind of regional protection available): ARIPO (BW, GH, (71) Applicant (for all designated States except US): EVONIK GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, SZ, TZ, DEGUSSA GMBH [DE/DE]; Rellinghauser StraBe 1-11, UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, MD, RU, 45128 Essen (DE). TJ, TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, (72) Inventors; and LU, LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, (75) Inventors/Applicants (for US only): HABERLAND, SI, SK, SM, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, Jinrgen [DE/DE]; Dahlienstr. 26, 45721 Haltern am See GN, GQ, GW, ML, MR, NE, SN, TD, TG). (DE). ZEHNACKER, Olivier [FRIDE]; Von Bruchhausen Str. 1, 45657 Recklinghausen (DE). GRAMMANN, Katrin Published: [DE/DE]; Denningsgraben 16, 45739 Oer-Erkenschwick - with international search report (Article 21(3)) (DE). HOLTKAMP, Manuel [DE/DE]; R6merstr. 85, _ with sequence listingpart of description (Rule S.2(a)) 45721 Haltern am See (DE). WOLTER, Jan [DE/DE]; An den Ksmpen 36, 40489 Dusseldorf (DE). HULLER, Thomas [DE/DE]; RingerottstraBe 97, 45772 Marl (DE). MOHR, Alexander [DE/DE]; B6rster Weg 18, 45657 Recklinghausen (DE). GRIMBERG, Gerlind [DE/DE], Rosenweg 12, 48301 Nottuln (DE). HAAS, Thomas [DE/DE]; Backenkamp 9,48161 Miinster (DE). As printed (54) Tlc: PROCESS FOR THE PRODUCTION OF ISO)MALTULO)SE FROM PLANT JUICES (54) Bezeichnung VERFAHRFN ZUR HERSTELLUNU VON ISOMALTULOSE AUS PFLANZFNSnFTEN (57) Abstract, The present nemien relaei to a process or prode in SonatUKe, Which conpries the process steps: A) bringing an iunobilized enzyme complex capable of catalysmn Theconversion of sucrose mie isomLuu1Lose mt; contact with a Suctose contaming solution; B) isomnerilion of leas1 some of t sucose 1o give iSomahlulose; C) niov ig the enzyNme complex, and if approiate 1)) further purification o: te isomaltulosc. characterized m that the sucroseonng solution used i at least one selected from the series consisting of ihick it. or hbin juice, prefe r Thick juice. (57) Zusammenfassung: Gegentaud der vorliegenden Erfindung it m rfairen zu= lerstellung von Ioomaulos. umfassend die Verfhrensschritte A) in Kontakt hrigen eines imnobiliieren Enzym koplexe welches die Umsezung voi Scharose zu tnmahulose so katalysicren verniag, ni einr haro nugen L&sung, 1) 'omerisierung nindestens eine, lees der Saccharose zu I6-amose; C) Abtrunen t Enzymkomplexe und g egebenenfalb DI) weere: Aufreinet der homahulose due gekenn/eichnet, das a& saccharosehahige Lisung nindstens einer auewahh aus Diksaf: oder Dinn . bevor/tpg hekaLt cingesem witI, WO 2012/150051 PCT/EP2012/052096 Process for the production of isomaltulose from plant juices Field of the invention The present invention relates to a process for isomerizing a 5 sucrose-containing plant juice to give isomaltulose. Prior art The enzymatic isomerization of sucrose in aqueous solution with D the aid of isomaltulose synthases (sucrose glucosylmutases, EC 5.4.99.11) to give isomaltulose is a known process. For example, DE1049800 describes a process for fermentative production of isomaltulose from sucrose-containing plant juices by means of the bacterial strain Protaminobacter rubrum. 5 Alternatively, enzymatic conversion may also be carried out using dead organisms. The cell mass is removed batchwise by centrifugation at the end of the reaction and the product is worked-up further. This process is found in the literature at H. Schiweck, alimenta 19, 5-16, 1980. D EP0001099 describes a process of continuous fermentation of the bacterial strains Protaminobacter rubrum (CBS574.77) and Serratia plymuthica (ATCC 15928) with concomitant isomerization of sucrose to isomaltulose using, inter alia, thick juice or thin juice as substrate, with the substrate solution itself being nutrient medium for culturing the cells. After the reaction the cells are removed from the product solution by means of centrifugation. Cultivating the biocatalytically active organisms separately from the actual isomerization in a medium other than the substrate medium is expressly shown to be disadvantageous. DE3241788 describes a process for preparing 1-0-a-D glucopyranosido-D-fructose from aqueous sucrose solutions with free cells of the bacterial strains from the group consisting of Protaminobacter rubrum (CBD574 .77), Serratia plymuthica (ATCC 15928), Serratia marescens (NCIB 8285), Leuconostoc WO 2012/150051 PCT/EP2012/052096 -2 mesenteroides (NRRL-B-512 F) and Erwinia rhapontici (NCPPB 1578). The process described may use thick juice which also serves as carbon source for the cell-culturing process. In the case of immobilized cells, use is made of pure sucrose solution. 5 With the fermentative production of isomaltulose disclosed in the two aforementioned documents it is possible to employ economically and energetically reasonable thick juice under sterile conditions. It should be mentioned here, however, that, disadvantageously, some of the sucrose is utilized for producing D cell mass, and also complex sterile hardware as well as elaborate removal of the cell suspension from the product stream are required. EP0983374 describes a process for simultaneously preparing isomaltulose and betaine using immobilized bacterial strains from the group consisting of Protaminobacter rubrum (CBD574.77), Serratia plymuthica (ATCC 15928) and Erwinia rhapontici (NCPPB 1578). The production process starts with molasses, a composition which is produced at the end of the sugar manufacturing process. It is not possible to use thick juice as substrate in this process, which requires an explicit depletion of the sucrose by, for example, crystallization. W097/44478 describes a process for preparing isomaltulose using living, immobilized bacterial strains from the group consisting of Protaminobacter rubrum (CBD574.77), Serratia plymuthica (ATCC 15928) and Erwinia rhapontici (ATCC 29284). The production process starts with molasses or pure sucrose solution. EP0028900 describes a process for preparing isomaltulose using immobilized cells of the bacterial strain Erwinia rhapontici (NCPPB 1578). The production process starts with a pure sucrose solution with a maximum of 15% v/v molasses admixed. DE2217628, EP49472, EP3038219 and EP91063 describe processes using immobilized bacterial cells for enzymatic conversion of pure sucrose to isomaltulose. To this end, EP0625578 employs bacterial strains from the group consisting of Protaminobacter * rubrum (CBS 574.77), Serratia plymuthica (ATCC 15928), Serratia marcescens (NCIB 8285), Leuconostoc mesenteroides (NRRL-B 512 F WO 2012/150051 PCT/EP2012/052096 -3 (ATCC 1083 a)) and Erwinia rhapontici (NCPPB 1578). EP0392556 and EP1257638 describe the use of bacterial strains from the group consisting of Klebsiella terrigena JCM 1687, Klebsiella sp. No. 88 (FERM BP-2838) and Klebsiella singaporensis LX3 and 5 LX21. DE3133123 and EP0915986 describe isomerization processes using immobilized cells and employing the enzyme catalysts of calcium alginate or ion exchangers in the immobilization processes. A common feature of all of the known processes for the ) isomerization of sucrose to isomaltulose using immobilized cells is that of only the end products of the sugar manufacturing process being utilized, namely pure sucrose or molasses. It was therefore an object of the present invention to provide a process which enables isomaltulose to be obtained from a readily 5 available sucrose source. Description of the invention Surprisingly we have now found that this object can be achieved by using sucrose-containing plant juices for preparing isomaltulose with the use of immobilized enzyme complexes. The present invention therefore relates to a process for preparing isomaltulose, comprising the process steps of A) contacting an immobilized enzyme complex capable of catalyzing the conversion of sucrose to isomaltulose with a sucrose-containing solution; B) isomerizing at least some of the sucrose to isomaltulose; C) removing the enzyme complex, and optionally D) further purifying the isomaltulose, characterized in that the sucrose-containing solution used is at least one of thick juice or thin juice, preferably thick juice.
WO 2012/150051 PCT/EP2012/052096 -4 In this connection, the invention advantageously provides for the use of sucrose-containing plant juices as substrate for the isomerization. Despite the fact that the sucrose-containing solutions used are unpurified and therefore highly complex substrate mixtures, the present invention surprisingly achieves the same yield as and even a higher selectivity in the isomerization than with high purity sucrose. One advantage of the present invention is that, as a result of the increased residence time of the immobilized cells in combination with a crude plant juice as substrate, a substantially higher resource efficiency is achieved due to the purification step to give the high-purity sucrose being omitted. A further advantage of the present invention is that of being able to save energy and operating means by directly using thick juice. Another advantage of the present invention is that, owing to the use of thick juice, no buffer solutions of any kind or other additives are required for stabilizing the enzymatic activity, the enzymes employed rather being stabilized naturally. Yet another advantage of the present invention is that using sucrose-containing plant juices, in contrast to using purely aqueous sucrose solution, prevents a bleeding of the bed. Another advantage of the present invention is that of enabling the content of the cariogenic monosaccharides, fructose and glucose, to be reduced by using the thick juice during isomerization, compared to using purified sucrose solution. The term "isomaltulose" is to be understood as meaning 6-0-a-D glucopyranosido-D-fructose. The term "enzyme complex" is to be understood as meaning a composition or mixture composition which has at least one active enzyme and which may also be complex in nature, such as, for example, a living cell. Further examples of an enzyme complex are fusion proteins in which the at least one active enzyme is linked to at least one further polypeptide, but also a purified WO 2012/150051 PCT/EP2012/052096 -5 enzyme itself can be an enzyme complex for the purposes of the present invention. In connection with the present invention, the term "immobilized enzyme complex" is to be understood as meaning an enzyme complex which is bound to or enclosed by a matrix in such a way that free diffusion of the enzyme complex or its free movement in an aqueous solution is restricted, for example slowed. In connection with the present invention, the term "thin juice" is to be understood as meaning the product which is present after juice purification in the production of sugar from sugar beets or sugar cane. Thin juice typically comprises 10-20% by weight of sucrose based on total juice. In connection with the present invention, the term "thick juice" is to be understood as meaning the thickened "thin juice" which comprises more than 20% by weight of sucrose based on total juice. Unless stated otherwise, all percentages (%) given are percentages by mass. In the process according to the invention, the sucrose containing solution used is advantageously thick juice; this can, if required, be diluted with water to a volume ratio of 1:5, based on thick juice to water, which results in a better controllable isomerization. More specifically, dilution of the thick juice with water is chosen for the thick juice to contain, at the start of process step A), a sucrose concentration of 20-80% by weight, in particular of 30-50% by weight, based on total juice. In particular, thin juice or thick juice from sugar beets is to be used advantageously, with particular preference being given to thick juice from sugar beets. The enzyme present in the immobilized enzyme complex is preferably at least one sucrose glucosylmutase of enzyme class EC 5.4.99.11. Particular preference is given to using sucrose glucosylmutases from Protaminobacter rubrum, in particular the strain Protaminobacter rubrum CBS 574.77 and those with Seq ID WO 2012/150051 PCT/EP2012/052096 -6 Nos. 5 and 6; Protaminobacter ruber Z12; Serratia plymuthica, in particular the strain Serratia plymuthica ATCC 15928; Serratia odorifera, in particular the strain Serratia odorifera 4Rx13 (Seq ID No. 7); Serratia marcescens, in particular the strain Serratia marcescens NCIB 8285; Leuconostoc mesenteroides, in particular the strain Leuconostoc mesenteroides ATCC 1083 a; Erwinia rhapontici, in particular the strains Erwinia rhapontici ATCC29283 (Seq ID No. 1) , NCPPB 1578, DSM 4484 (Seq ID No. 8), NX-5 (Seq ID No. 9) and WAC2928 (Seq ID No. 10); Erwinia sp., in particular the strain Erwinia sp. D12; Agrobacterium radiobacter, in particular the strain Agrobacterium radiobacter MX-232; Klebsiella terrigena, in particular the strain Klebsiella terrigena JCM 1687; Klebsiella sp., in particular the strains Klebsiella sp. FERM BP-2838, LX3 (Seq ID No. 11) and NK33-98-8 (Seq ID No. 12); Klebsiella pneumoniae, in particular the strain Klebsiella pneumoniae 342 (Seq ID No. 4); Klebsiella singaporensis, in particular the strain Klebsiella singaporensis LX21; Pseudomonas mesoacidophila, in particular the strain Pseudomonas mesoacidophila MX-45 (Seq ID No. 2); Pantoea dispersa, in particular the strain Pantoea dispersa UQ68J (Seq ID NO. 3); Klebsiella planticola, in particular the strains Klebsiella planticola CCRC 19112, MX10 and UQ14S (Seq ID NO. 13); Enterobacter sp., in particular the strain Enterobacter sp. FMB-1 (Seq ID NO. 16), SZ62 and Ejp617 (Seq ID NO. 14); Azotobacter vinelandii DJ (Seq ID NO. 15) in the process according to the invention, with Protaminobacter rubrum CBS 574.77 and Protaminobacter ruber Z12 being particularly preferred. The enzymes may be employed in purified form by way of polypeptides. For ease of purification, these can be in the form of fusion proteins, with, for example, a tag facilitating purification, such as for example a His tag, a Strep tag, a GST tag or an MBP tag, being fused to the enzyme.
WO 2012/150051 PCT/EP2012/052096 -7 In the process according to the invention, preference is given to the enzyme complex being whole cells which are preferably selected from the group consisting of Protaminobacter rubrum, in particular the strain Protaminobacter rubrum CBS 574.77; 5 Protaminobacter ruber Z12; Serratia plymuthica, in particular the strain Serratia plymuthica ATCC 15928; Serratia odorifera, in particular the strain Serratia odorifera 4Rx13; Serratia marcescens, in particular the strain Serratia marcescens NCIB 8285; Leuconostoc mesenteroides, in particular the strain D Leuconostoc mesenteroides ATCC 1083 a; Erwinia rhapontici, in particular the strains Erwinia rhapontici ATCC29283, NCPPB 1578, DSM 4484, NX-5 and WAC2928; Erwinia sp., in particular the strain Erwinia sp. D12; Agrobacterium radiobacter, in particular the strain Agrobacterium radiobacter MX-232; Klebsiella terrigena, in particular the strain Klebsiella terrigena JCM 1687; Klebsiella sp., in particular the strains Klebsiella sp. FERM BP-2838, LX3 and NK33-98-8; Klebsiella pneumoniae, in particular the strain Klebsiella pneumoniae 342; Klebsiella singaporensis, in particular the strain Klebsiella singaporensis LX21; Pseudomonas mesoacidophila, in particular the strain Pseudomonas mesoacidophila MX-45; Pantoea dispersa, in particular the strain Pantoea dispersa UQ68J; Klebsiella planticola, in particular the strains Klebsiella planticola CCRC 19112, MX10 and UQ14S; Enterobacter sp., in particular the strain Enterobacter sp. FMB-1, SZ62 and Ejp617; Azotobacter vinelandii DJ, with Protaminobacter rubrum CBS 574.77 and Protaminobacter ruber Z12 being particularly preferred. The enzyme complexes may be immobilized, for example, in the form of CLEAs (insoluble crosslinked enzyme aggregates) (Cao, L. et al., 2000, Cross-linked enzyme aggregates: a simple and effective method for the immobilization of penicillin acylase, Org. Lett., 2: 1361-1264) or on solid support materials of natural or synthetic origin. Examples of natural materials are polysaccharides such as alginate, agarose, sepharose, cellulose and its derivatives (e.g. DEAE- or CM-cellulose) . It is also WO 2012/150051 PCT/EP2012/052096 -8 possible to use modified sepharoses, for example epoxy activated, bromocyanogen-activated, NHS-activated, thiol activated sepharose. These sepharoses are commercially available for example from the companies GE Healthcare, BioRad, Sigma and Pierce. Synthetic organic polymers which may be employed are polystyrene derivatives, polyacrylates, in particular epoxide-activated acrylic resin beads (Eupergit), polymethacrylates, polyacrylamides, vinyl and allyl polymers, polyesters or polyamides. Possible inorganic carriers are materials based on silicon oxides or aluminium oxides, or mixtures thereof. The enzyme complexes may also be immobilized by encapsulation in polymeric porous gels e.g. hydrophobic sol-gel materials of RSi(OCH 3
)
3 or mixtures of RSi(OCH 3
)
3 and Si(OCH 3
)
4 (Reetz, M.T.; Zonta, A.; Simpelkamp, J.; Rufinska, A.; Tesche, B. J. Sol-Gel Sci. Technol. 1996, 7, p. 35-43) or of porous polymeric silica gels (Elgren, T.M.; Zadvorny, O.A.; Brecht, E.; Douglas, T.; Zorin, N.A.; Maroney, M. J. & Peters, J. W.). Besides immobilization, multiple usage of the enzyme in enzyme membrane reactors is also conceivable. The enzyme complexes employed in the process according to the invention, in particular cells, are preferably immobilized in polysaccharides such as alginate, pectin, carrageenan, chitosan or polyvinyl alcohols, such as lentikats for example, or mixtures thereof, in particular in alginate; cf. in this regard for example Shimizu, H., et al. (1997) Screening of novel microbial enzymes for the production of biologically and chemically useful compounds, in: Advances in biochemical engineering biotechnology, Vol58: New Enzymes for Organic Synthesis (Scheper, T., ed.) pp. 45-88, Springer, New York.
WO 2012/150051 PCT/EP2012/052096 -9 A particularly preferred immobilization to be used for any form of the enzyme complexes employed in the process according to the invention is the process described in EP2011865, in which the enzyme complexes immobilized on an inert support are provided with a silicone coating obtained by hydrosilylation. According to the invention, preference is given to the pH in process step B) being from 4 to 9.5. This pH is advantageously adjusted by means of an acid, in particular an inorganic acid, preferably from the group consisting of sulphuric acid, hydrochloric acid and acetic acid. According to the invention, preference is furthermore given to the temperature, measured in the sucrose-containing solution, in process step B) being from 20 to 400C, preferably from 25 to 350C. In process step C), the enzyme complex is removed from the isomaltulose, for example by filtration, sedimentation or centrifugation. Due to the immobilizate character of the enzyme complex, said removal is easier than with non-immobilized enzyme. For reasons of process economy, it is preferred according to the invention if the immobilized enzyme complex is used in the form of a solid bed through which the sucrose-containing solution flows (H. Schiweck, Zuckerind. (1990), 115 (7), 555-565). Corresponding solid-bed processes are described in A. Liese, et. al. Processes in A. Liese, K. Seelbach, C. Wandrey (Eds.), Industrial Biotransformations 2nd Edition (2006), Wiley-VCH, Weinheim. In such a process, process steps A) to C) are continuous, and thus merging into one another up to being simultaneous. The isomaltulose may be further purified in the optional process step D), in particular by crystallization or chromatography. Corresponding procedures are described in DE3241788 and EP0983374.
WO 2012/150051 PCT/EP2012/052096 - 10 The examples listed below describe the present invention by way of example without any intention of limiting the invention, the scope of which arises from the entire description and the claims, to the embodiments specified in the examples. Examples: Example 1: Preparation of the biocatalyst Cells were washed off a subculture of the strain Protaminobacter rubrum (CBS574.77) with 1-5 ml of a sterile nutrient medium consisting of 50 g/kg sucrose, 15 g/kg corn steep liquor, 7 g/kg ammonium sulphate, 0.5 g/kg yeast extract, 1 g/kg potassium dihydrogen sulphate, 0.41 g/kg magnesium chloride heptahydrate, 0.004 g/kg manganese chloride tetrahydrate, 0.047 g/kg iron citrate monohydrate and 926 g/kg water, if necessary adjusted to pH 7.2. This suspension served as inoculum for the preculture, which comprises 200 ml of the above nutrient solution in a 1 1 shaker flask. After cultivation at 30 0 C for 20 hours, a 2 1 fermenter containing one litre of sterile production medium consisting of 50 g/kg sucrose, 15 g/kg corn steep liquor, 3 g/kg ammonium sulphate, 4 g/kg ammonium hydrogen phosphate, 0.5 g/kg yeast extract, 1 g/kg potassium dihydrogen sulphate, 0.41 g/kg magnesium chloride heptahydrate, 0.004 g/kg manganese chloride tetrahydrate, 0.047 g/kg iron citrate monohydrate and 926 g/kg water, adjusted to pH 7.2, was inoculated with the preculture in such a way that the initial optical density (OD 600 ) was 1. Fermentation was carried out at 30 0 C, pH 7 (regulated) and a P02 of 30% (cascade stirrer, Airflow) . Sucrose was fed in during fermentation. After 15-20 h, the fermentation process was complete and the biomass was harvested for immobilization.
WO 2012/150051 PCT/EP2012/052096 - 11 For this purpose, the suspension obtained was mixed with water and a 4% strength alginate solution with a 1:1 volume ratio. This suspension was then immobilized by adding it dropwise to a 2% strength CaCl 2 solution. The resulting beads were post-cured with polyethylenimine and glutaraldehyde. The resulting biocatalyst can be stored for several weeks at 4-10 0 C. Example 2: Preparation of isomaltulose by isomerizing pure sucrose solution (not according to the invention) Pure sucrose was prepared from a thick juice from sugar beets as described in Sugar in Ullman's Encyclopedia of Industrial Chemisty (2007); the sucrose yield is 88.3 %. The immobilized cells obtained in Example 1 were introduced into a heatable column reactor and heated to 20-35 0 C. A sucrose solution with a dry substance (ds) content of 35 to 45% was prepared from said pure sucrose and tap water and passed through the column reactor in a continuous stream. HPLC analysis of the isomaltulose solution leaving the column reactor revealed in this process step averages for yield, selectivity and turnover of: yield 85% selectivity 91% turnover 94% The yield of this process was 75%, based on sucrose used. Example 3: Preparation of isomaltulose by isomerizing thick juice (according to the invention) The immobilized cells obtained in Example 1 are introduced into a heatable column reactor and heated to 20-35 0 C, and the above described diluted thick juice from sugar beets with a content of 35 to 45% ds sucrose is passed through in a continuous stream.
WO 2012/150051 PCT/EP2012/052096 - 12 HPLC analysis of the isomaltulose solution leaving the column reactor revealed averages for yield, selectivity and turnover of: yield 82% selectivity 91% turnover 90% The yield of this process was 82%, based on sucrose used, and thus is 7 % higher than the yield of a process of the prior art. Example 4: Selectivity comparison of the isomerization of thick juice compared to the isomerization of pure sucrose solution A column reactor heated to 300C was charged with immobilized cells obtained according to Example 1. Purified sucrose solution, the concentration of which was adjusted with tap water to 40% dry substance content and the pH of which was adjusted to 6 with concentrated sulphuric acid, was passed through column reactor 1 in a continuous stream. Thick juice from sugar beets, diluted to 40% sucrose dry substance content and adjusted to pH 6 with concentrated sulphuric acid, was passed through column reactor 2 in a continuous stream. To improve the determinability of the selectivities, the isomaltulose yields in both column reactors were set to an identical value of 79%, based on the particular sucrose dry substance content, by adjusting the volume stream. Analytical determination was carried out by way of HPLC analysis of the isomaltulose solution obtained. ) The selectivities achieved were as follows: selectivity (thick juice) 85 % selectivity (sucrose solution) 83 % In addition, formation of the undesired, since cariogenic, monosaccharides, fructose and glucose, was reduced by approx. 10% (based on dry substance) in the final product by using thick juice compared with the use of purified sucrose solution.
Claims (4)
1. Process for preparing isomaltulose, comprising the process steps of 5 A) contacting an immobilized enzyme complex capable of catalyzing the conversion of sucrose to isomaltulose with a sucrose-containing solution; B) isomerizing at least some of the sucrose to 10 isomaltulose; C) removing the enzyme complex, and optionally D) further purifying the isomaltulose, characterized in that the sucrose-containing solution used is at least 15 one of thick juice or thin juice, preferably thick juice.
2. Process according to Claim 1, characterized in that 20 the sucrose-containing solution used is a thick juice diluted with water and having a sucrose concentration of 20-80% by weight, in particular
30-50% by weight, based on total juice. 25 3. Process according to Claim 1 or 2, characterized in that the enzyme present in the immobilized enzyme complex is at least one sucrose glucosylmutase of enzyme class EC 5.4.99.11, in particular one from 30 Protaminobacter rubrum, in particular the strain Protaminobacter rubrum CBS 574.77 and those with Seq ID Nos. 5 and 6; Protaminobacter ruber Z12; WO 2012/150051 PCT/EP2012/052096 - 14 Serratia plymuthica, in particular the strain Serratia plymuthica ATCC 15928; Serratia odorifera, in particular the strain Serratia odorifera 4Rxl3 (Seq ID No. 7); Serratia marcescens, in particular 5 the strain Serratia marcescens NCIB 8285; Leuconostoc mesenteroides, in particular the strain Leuconostoc mesenteroides ATCC 1083 a; Erwinia rhapontici, in particular the strains Erwinia rhapontici ATCC29283 (Seq ID No. 1), NCPPB 1578, 10 DSM 4484 (Seq ID No. 8), NX-5 (Seq ID No. 9) and WAC2928 (Seq ID No. 10); Erwinia sp., in particular the strain Erwinia sp. D12; Agrobacterium radiobacter, in particular the strain Agrobacterium radiobacter MX-232; Klebsiella terrigena, in 15 particular the strain Klebsiella terrigena JCM 1687; Klebsiella sp., in particular the strains Klebsiella sp. FERM BP-2838, LX3 (Seq ID No. 11) and NK33-98-8 (Seq ID No. 12); Klebsiella pneumoniae, in particular the strain Klebsiella 20 pneumoniae 342 (Seq ID No. 4); Klebsiella singaporensis, in particular the strain Klebsiella singaporensis LX21; Pseudomonas mesoacidophila, in particular the strain Pseudomonas mesoacidophila MX-45 (Seq ID No. 2); Pantoea dispersa, in 25 particular the strain Pantoea dispersa UQ68J (Seq ID NO. 3); Klebsiella planticola, in particular the strains Klebsiella planticola CCRC 19112, MX10 and UQ14S (Seq ID NO. 13); Enterobacter sp., in particular the Enterobacter sp. FMB-1 (Seq ID NO. 30 16), SZ62 and Ejp617 (Seq ID NO. 14); Azotobacter vinelandii DJ (Seq ID NO. 15). WO 2012/150051 PCT/EP2012/052096 - 15 4. Process according to at least one of the preceding claims, characterized in that the enzyme complex is cells, in particular cells 5 which are selected from Protaminobacter rubrum, in particular the strain Protaminobacter rubrum CBS
574.77; Protaminobacter ruber Z12; Serratia plymuthi ca, in particular the strain Serratia plymuthica ATCC 15928; Serratia odorifera, in 10 particular the strain Serratia odorifera 4Rx13; Serratia marcescens, in particular the strain Serratia marcescens NCIB 8285; Leuconostoc mesenteroides, in particular the strain Leuconostoc mesenteroides ATCC 1083 a; Erwinia rhapontici, in 15 particular the strains Erwinia rhapontici ATCC29283, NCPPB 1578, DSM 4484, NX-5 and WAC2928; Erwinia sp., in particular the strain Erwinia sp. D12; Agrobacterium radiobacter, in particular the strain Agrobacterium radiobacter MX-232; Klebsiella 20 terrigena, in particular the strain Klebsiella terrigena JCM 1687; Klebsiella sp., in particular the strains Klebsiella sp. FERM BP-2838, LX3 and NK33-98-8; Klebsiella pneumoniae, in particular the strain Klebsiella pneumoniae 342; Klebsiella 25 singaporensis, in particular the strain Klebsiella singaporensis LX21; Pseudomonas mesoacidophila, in particular the strain Pseudomonas mesoacidophila MX-45; Pantoea dispersa, in particular the strain Pantoea dispersa UQ68J; Klebsiella planticola, in 30 particular the strains Klebsiella planticola CCRC 19112, MX10 and UQ14S; Enterobacter sp. FMB-1, SZ62 and Ejp617; Azotobacter vinelandii DJ. WO 2012/150051 PCT/EP2012/05209 6 - 16 5. Process according to Claim 4, characterized in that the cells are immobilized in polysaccharides such 5 as alginate, pectin, carrageenan, chitosan, or polyvinyl alcohols, such as lentikats for example, or mixtures thereof, in particular in alginate. 6. Process according to at least one of the preceding 10 claims, characterized in that a pH of from 4 to 9.5 is present in process step B). 15 7. Process according to at least one of the preceding claims, characterized in that the temperature in process step B) is from 20 to 40OC. 20 8. Process according to at least one of the preceding claims, characterized in that the immobilized enzyme complex is used in the form 25 of a solid bed through which the sucrose-containing solution flows.
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE102011100772A DE102011100772A1 (en) | 2011-05-05 | 2011-05-05 | Process for the preparation of isomaltulose from plant juices |
| DE102011100772.9 | 2011-05-05 | ||
| PCT/EP2012/052096 WO2012150051A1 (en) | 2011-05-05 | 2012-02-08 | Process for the production of isomaltulose from plant juices |
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| AU2012251903A1 true AU2012251903A1 (en) | 2013-10-24 |
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| AU2012251903A Abandoned AU2012251903A1 (en) | 2011-05-05 | 2012-02-08 | Process for the production of isomaltulose from plant juices |
| AU2012251596A Abandoned AU2012251596A1 (en) | 2011-05-05 | 2012-05-04 | Process for preparing isomaltulose from plant juices |
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| AU2012251596A Abandoned AU2012251596A1 (en) | 2011-05-05 | 2012-05-04 | Process for preparing isomaltulose from plant juices |
Country Status (7)
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| EP (2) | EP2704594B1 (en) |
| CN (2) | CN103501636A (en) |
| AU (2) | AU2012251903A1 (en) |
| BR (2) | BR112013028369A2 (en) |
| DE (1) | DE102011100772A1 (en) |
| TW (1) | TW201307565A (en) |
| WO (2) | WO2012150051A1 (en) |
Families Citing this family (18)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE102012200907A1 (en) | 2012-01-23 | 2013-07-25 | Evonik Industries Ag | Method and absorption medium for absorbing CO2 from a gas mixture |
| DE102012207509A1 (en) | 2012-05-07 | 2013-11-07 | Evonik Degussa Gmbh | Method for absorbing CO2 from a gas mixture |
| DE102015212749A1 (en) | 2015-07-08 | 2017-01-12 | Evonik Degussa Gmbh | Method for dehumidifying moist gas mixtures |
| SG11201804996TA (en) * | 2015-11-12 | 2018-07-30 | Petiva Private Ltd | Food and beverage products comprising low calorie, low glycemic index (gi), and sustained energy release sugar composition |
| DE102016206891B3 (en) | 2016-04-22 | 2017-04-27 | Zf Friedrichshafen Ag | Hydropneumatic strut |
| DE102016210481B3 (en) | 2016-06-14 | 2017-06-08 | Evonik Degussa Gmbh | Process for purifying an ionic liquid |
| DE102016210483A1 (en) | 2016-06-14 | 2017-12-14 | Evonik Degussa Gmbh | Process and absorbent for dehumidifying moist gas mixtures |
| DE102016210478A1 (en) | 2016-06-14 | 2017-12-14 | Evonik Degussa Gmbh | Method for dehumidifying moist gas mixtures |
| EP3257843A1 (en) | 2016-06-14 | 2017-12-20 | Evonik Degussa GmbH | Method of preparing a high purity imidazolium salt |
| EP3257568B1 (en) | 2016-06-14 | 2019-09-18 | Evonik Degussa GmbH | Method for the removal of moisture from moist gas mixtures by use of ionic liquids |
| DE102016210484A1 (en) | 2016-06-14 | 2017-12-14 | Evonik Degussa Gmbh | Method for dehumidifying moist gas mixtures |
| US20210076724A1 (en) | 2018-02-28 | 2021-03-18 | C-Lecta Gmbh | Enzymatic in-situ fortification of food with functional carbohydrates |
| KR101964958B1 (en) * | 2018-04-20 | 2019-04-02 | (주)네오크레마 | Palatinose syrup with inhibited crystalization which has inhibition activity of elevation of blood sugar level |
| BR112021002996A2 (en) * | 2018-08-22 | 2021-05-11 | Dsm Ip Assets B.V. | sucrose isomerases as food and nutritional supplements |
| CN109511849B (en) * | 2018-12-26 | 2022-03-01 | 广西壮族自治区农业科学院农产品加工研究所 | Cassava juice beverage capable of regulating intestinal flora and processing method thereof |
| CN113621601B (en) * | 2021-06-24 | 2023-06-30 | 浙江工业大学 | Sucrose isomerase mutant, coding gene and application thereof |
| CN114703211A (en) * | 2022-02-16 | 2022-07-05 | 李宪臻 | Sucrose isomerase activity inclusion body fused with coiled coil structure domain |
| CN116179631B (en) * | 2023-04-27 | 2023-11-28 | 山东百龙创园生物科技股份有限公司 | Isomaltulose crystal and preparation method thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE1049800B (en) | 1957-10-11 | 1959-01-29 | Süddeutsche Zucker-Aktiengesellschaft, Mannheim | METHOD FOR MANUFACTURING PALATINOSE (6-α-GLUCOSIDO-FRUCTOFURANOSE) |
| DE2217628C2 (en) | 1972-04-12 | 1974-06-06 | Sueddeutsche Zucker Ag | Process for the production of alpha-D-glucopyranosido square bracket on 1-6 square bracket to sorbitol (isomaltite) |
| EP0001099B1 (en) | 1977-09-13 | 1980-08-20 | Bayer Ag | Process for the continuous isomerisation if saccharose to isomaltulose by means of microorganisms |
| DE2741197A1 (en) * | 1977-09-13 | 1979-03-29 | Bayer Ag | Continuous sucrose conversion to iso:maltulose - during continuous fermentation of iso:maltulose-forming microorganisms with addn. of sucrose-contg. solns. |
| DE3066516D1 (en) | 1979-11-07 | 1984-03-15 | Tate & Lyle Plc | Production of isomaltulose |
| JPS5836959B2 (en) | 1980-08-21 | 1983-08-12 | 三井製糖株式会社 | Method for producing palatinose using immobilized α-glucosyltransferase |
| JPS5762310A (en) | 1980-10-02 | 1982-04-15 | Matsushita Electric Ind Co Ltd | Coil combustion wick |
| DE3038219A1 (en) | 1980-10-09 | 1982-04-15 | Süddeutsche Zucker AG, 6800 Mannheim | METHOD FOR PRODUCING ISOMALTULOSE (6-O- (ALPHA) -D-GLUCOPYRANOSIDO-D-FRUCTOSE) WITH THE AID OF IMMOBILIZED BACTERIA CELLS |
| DE3213107A1 (en) | 1982-04-07 | 1983-10-13 | Süddeutsche Zucker AG, 6800 Mannheim | METHOD FOR PRODUCING ISOMALTULOSE (6-O- (ALPHA) -D-GLUCOPYRANOSIDO-D-FRUCTOSE) WITH THE AID OF IMMOBILIZED BACTERIA CELLS |
| DE3241788A1 (en) | 1982-11-11 | 1984-05-17 | Süddeutsche Zucker AG, 6800 Mannheim | METHOD FOR PRODUCING 1-0- (ALPHA) -D-GLUCOPYRANOSIDO-D-FRUCTOSE AND USE AS A SWEETENER |
| DE3725974A1 (en) | 1987-08-05 | 1989-02-16 | Messerschmitt Boelkow Blohm | INSERT FOR A COMPONENT THAT IS AT LEAST PARTLY OF PLASTIC |
| CN1030530C (en) * | 1989-02-03 | 1995-12-20 | 广东省微生物研究所 | Preparation of parakin sugar by solidified alpha-glucosyl group transferase |
| JPH02273192A (en) | 1989-04-13 | 1990-11-07 | Meito Sangyo Kk | Production of isomaltulose |
| EP0625578B2 (en) | 1993-05-06 | 2004-04-28 | Südzucker Aktiengesellschaft Mannheim/Ochsenfurt | Sweetener, process of preparation and use thereof |
| DE9321600U1 (en) * | 1993-05-06 | 2000-04-06 | Suedzucker Ag | Sweeteners |
| FI104563B (en) | 1996-05-17 | 2000-02-29 | Xyrofin Oy | Method and carrier for the production of isomaltulose by immobilized microorganisms |
| FI105048B (en) * | 1997-05-22 | 2000-05-31 | Xyrofin Oy | Process for the preparation of isomaltulose and other products |
| ATE368736T1 (en) | 2000-02-15 | 2007-08-15 | Agency Science Tech & Res | BACTERIAL STRAINS OF THE GENUS KLEBSIELLA AND THEIR GENE CODING ISOMALTULOSE SYNTHASE |
| DE102007031689A1 (en) | 2007-07-06 | 2009-01-08 | Evonik Goldschmidt Gmbh | enzyme preparations |
-
2011
- 2011-05-05 DE DE102011100772A patent/DE102011100772A1/en not_active Ceased
-
2012
- 2012-02-08 BR BR112013028369A patent/BR112013028369A2/en not_active IP Right Cessation
- 2012-02-08 WO PCT/EP2012/052096 patent/WO2012150051A1/en active Application Filing
- 2012-02-08 EP EP12702557.5A patent/EP2704594B1/en not_active Not-in-force
- 2012-02-08 CN CN201280021859.XA patent/CN103501636A/en active Pending
- 2012-02-08 AU AU2012251903A patent/AU2012251903A1/en not_active Abandoned
- 2012-05-04 WO PCT/EP2012/058228 patent/WO2012150332A1/en active Application Filing
- 2012-05-04 EP EP12717789.7A patent/EP2704595A1/en not_active Withdrawn
- 2012-05-04 CN CN201280021863.6A patent/CN103501637A/en active Pending
- 2012-05-04 BR BR112013028410A patent/BR112013028410A2/en not_active IP Right Cessation
- 2012-05-04 AU AU2012251596A patent/AU2012251596A1/en not_active Abandoned
- 2012-05-04 TW TW101116148A patent/TW201307565A/en unknown
Also Published As
| Publication number | Publication date |
|---|---|
| WO2012150332A1 (en) | 2012-11-08 |
| DE102011100772A1 (en) | 2012-11-08 |
| EP2704594B1 (en) | 2016-04-27 |
| AU2012251596A1 (en) | 2013-10-24 |
| BR112013028369A2 (en) | 2017-03-01 |
| EP2704595A1 (en) | 2014-03-12 |
| EP2704594A1 (en) | 2014-03-12 |
| TW201307565A (en) | 2013-02-16 |
| WO2012150051A1 (en) | 2012-11-08 |
| CN103501637A (en) | 2014-01-08 |
| BR112013028410A2 (en) | 2019-09-24 |
| CN103501636A (en) | 2014-01-08 |
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