JPH04112797A - Production of isomaltooligosaccharide - Google Patents
Production of isomaltooligosaccharideInfo
- Publication number
- JPH04112797A JPH04112797A JP23188790A JP23188790A JPH04112797A JP H04112797 A JPH04112797 A JP H04112797A JP 23188790 A JP23188790 A JP 23188790A JP 23188790 A JP23188790 A JP 23188790A JP H04112797 A JPH04112797 A JP H04112797A
- Authority
- JP
- Japan
- Prior art keywords
- reaction
- organic solvent
- maltose
- water
- glucose
- 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.)
- Pending
Links
- DLRVVLDZNNYCBX-RTPHMHGBSA-N isomaltose 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@@H](O)C(O)O1 DLRVVLDZNNYCBX-RTPHMHGBSA-N 0.000 title claims abstract description 17
- 238000004519 manufacturing process Methods 0.000 title claims description 11
- 239000003960 organic solvent Substances 0.000 claims abstract description 16
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 15
- 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 claims abstract description 12
- 239000008103 glucose Substances 0.000 claims abstract description 12
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 claims abstract description 11
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 claims abstract description 11
- 230000002209 hydrophobic effect Effects 0.000 claims abstract description 11
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 claims abstract description 7
- 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 claims abstract description 6
- 238000006276 transfer reaction Methods 0.000 claims abstract description 4
- 229920001542 oligosaccharide Polymers 0.000 claims description 12
- 230000002255 enzymatic effect Effects 0.000 claims description 2
- 238000006243 chemical reaction Methods 0.000 abstract description 22
- SNRUBQQJIBEYMU-UHFFFAOYSA-N dodecane Chemical compound CCCCCCCCCCCC SNRUBQQJIBEYMU-UHFFFAOYSA-N 0.000 abstract description 10
- 108010059892 Cellulase Proteins 0.000 abstract description 7
- 229940106157 cellulase Drugs 0.000 abstract description 7
- 108090000604 Hydrolases Proteins 0.000 abstract description 3
- 239000006185 dispersion Substances 0.000 abstract description 3
- 239000002994 raw material Substances 0.000 abstract description 3
- 108090000790 Enzymes Proteins 0.000 description 10
- 102000004190 Enzymes Human genes 0.000 description 10
- 229940088598 enzyme Drugs 0.000 description 10
- 238000006911 enzymatic reaction Methods 0.000 description 8
- 230000035484 reaction time Effects 0.000 description 8
- 239000007853 buffer solution Substances 0.000 description 7
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 6
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 6
- 238000004128 high performance liquid chromatography Methods 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 239000002904 solvent Substances 0.000 description 6
- 230000000694 effects Effects 0.000 description 5
- 239000010410 layer Substances 0.000 description 5
- 150000002482 oligosaccharides Chemical class 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 238000003756 stirring Methods 0.000 description 5
- AYRXSINWFIIFAE-SCLMCMATSA-N Isomaltose Natural products OC[C@H]1O[C@H](OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O)[C@@H](O)[C@@H](O)[C@@H]1O AYRXSINWFIIFAE-SCLMCMATSA-N 0.000 description 4
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 4
- 244000052616 bacterial pathogen Species 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 239000000203 mixture Substances 0.000 description 4
- 239000000758 substrate Substances 0.000 description 4
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 description 3
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 3
- OFBQJSOFQDEBGM-UHFFFAOYSA-N Pentane Chemical compound CCCCC OFBQJSOFQDEBGM-UHFFFAOYSA-N 0.000 description 3
- 230000007423 decrease Effects 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 3
- 238000000746 purification Methods 0.000 description 3
- 229930195734 saturated hydrocarbon Natural products 0.000 description 3
- -1 2.2-dimethylbutane Chemical compound 0.000 description 2
- QPFMBZIOSGYJDE-UHFFFAOYSA-N 1,1,2,2-tetrachloroethane Chemical compound ClC(Cl)C(Cl)Cl QPFMBZIOSGYJDE-UHFFFAOYSA-N 0.000 description 2
- ZFFMLCVRJBZUDZ-UHFFFAOYSA-N 2,3-dimethylbutane Chemical compound CC(C)C(C)C ZFFMLCVRJBZUDZ-UHFFFAOYSA-N 0.000 description 2
- AFABGHUZZDYHJO-UHFFFAOYSA-N 2-Methylpentane Chemical compound CCCC(C)C AFABGHUZZDYHJO-UHFFFAOYSA-N 0.000 description 2
- 241000228245 Aspergillus niger Species 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- RGSFGYAAUTVSQA-UHFFFAOYSA-N Cyclopentane Chemical compound C1CCCC1 RGSFGYAAUTVSQA-UHFFFAOYSA-N 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- YNQLUTRBYVCPMQ-UHFFFAOYSA-N Ethylbenzene Chemical compound CCC1=CC=CC=C1 YNQLUTRBYVCPMQ-UHFFFAOYSA-N 0.000 description 2
- URLKBWYHVLBVBO-UHFFFAOYSA-N Para-Xylene Chemical group CC1=CC=C(C)C=C1 URLKBWYHVLBVBO-UHFFFAOYSA-N 0.000 description 2
- 150000001720 carbohydrates Chemical class 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- MVPPADPHJFYWMZ-UHFFFAOYSA-N chlorobenzene Chemical compound ClC1=CC=CC=C1 MVPPADPHJFYWMZ-UHFFFAOYSA-N 0.000 description 2
- RWGFKTVRMDUZSP-UHFFFAOYSA-N cumene Chemical compound CC(C)C1=CC=CC=C1 RWGFKTVRMDUZSP-UHFFFAOYSA-N 0.000 description 2
- DIOQZVSQGTUSAI-UHFFFAOYSA-N decane Chemical compound CCCCCCCCCC DIOQZVSQGTUSAI-UHFFFAOYSA-N 0.000 description 2
- DBTMGCOVALSLOR-AXAHEAMVSA-N galactotriose Natural products OC[C@@H]1O[C@@H](O[C@@H]2[C@@H](O)[C@H](CO)O[C@@H](O[C@H]3[C@@H](O)[C@H](O)O[C@@H](CO)[C@@H]3O)[C@@H]2O)[C@H](O)[C@H](O)[C@H]1O DBTMGCOVALSLOR-AXAHEAMVSA-N 0.000 description 2
- 230000003301 hydrolyzing effect Effects 0.000 description 2
- 239000003456 ion exchange resin Substances 0.000 description 2
- 229920003303 ion-exchange polymer Polymers 0.000 description 2
- FBJQEBRMDXPWNX-FYHZSNTMSA-N isomaltotriose 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@@H](O)[C@@H](OC[C@@H]2[C@H]([C@H](O)[C@@H](O)C(O)O2)O)O1 FBJQEBRMDXPWNX-FYHZSNTMSA-N 0.000 description 2
- QWTDNUCVQCZILF-UHFFFAOYSA-N isopentane Chemical compound CCC(C)C QWTDNUCVQCZILF-UHFFFAOYSA-N 0.000 description 2
- IVSZLXZYQVIEFR-UHFFFAOYSA-N m-xylene Chemical group CC1=CC=CC(C)=C1 IVSZLXZYQVIEFR-UHFFFAOYSA-N 0.000 description 2
- BDJAEZRIGNCQBZ-UHFFFAOYSA-N methylcyclobutane Chemical compound CC1CCC1 BDJAEZRIGNCQBZ-UHFFFAOYSA-N 0.000 description 2
- UAEPNZWRGJTJPN-UHFFFAOYSA-N methylcyclohexane Chemical compound CC1CCCCC1 UAEPNZWRGJTJPN-UHFFFAOYSA-N 0.000 description 2
- BKIMMITUMNQMOS-UHFFFAOYSA-N nonane Chemical compound CCCCCCCCC BKIMMITUMNQMOS-UHFFFAOYSA-N 0.000 description 2
- TVMXDCGIABBOFY-UHFFFAOYSA-N octane Chemical compound CCCCCCCC TVMXDCGIABBOFY-UHFFFAOYSA-N 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 238000006116 polymerization reaction Methods 0.000 description 2
- 239000004094 surface-active agent Substances 0.000 description 2
- VZGDMQKNWNREIO-UHFFFAOYSA-N tetrachloromethane Chemical compound ClC(Cl)(Cl)Cl VZGDMQKNWNREIO-UHFFFAOYSA-N 0.000 description 2
- RSJKGSCJYJTIGS-UHFFFAOYSA-N undecane Chemical compound CCCCCCCCCCC RSJKGSCJYJTIGS-UHFFFAOYSA-N 0.000 description 2
- DFKPJBWUFOESDV-NGZVDTABSA-N (2S,3R,4S,5S,6R)-6-[[(2S,3R,4S,5S,6R)-3,4,5-Trihydroxy-6-[[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-[[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxymethyl]oxan-2-yl]oxymethyl]oxan-2-yl]oxymethyl]oxane-2,3,4,5-tetrol 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@@H](O)[C@@H](OC[C@@H]2[C@H]([C@H](O)[C@@H](O)[C@@H](OC[C@@H]3[C@H]([C@H](O)[C@@H](O)[C@@H](O)O3)O)O2)O)O1 DFKPJBWUFOESDV-NGZVDTABSA-N 0.000 description 1
- WSLDOOZREJYCGB-UHFFFAOYSA-N 1,2-Dichloroethane Chemical compound ClCCCl WSLDOOZREJYCGB-UHFFFAOYSA-N 0.000 description 1
- ZPQOPVIELGIULI-UHFFFAOYSA-N 1,3-dichlorobenzene Chemical compound ClC1=CC=CC(Cl)=C1 ZPQOPVIELGIULI-UHFFFAOYSA-N 0.000 description 1
- 108010084185 Cellulases Proteins 0.000 description 1
- 102000005575 Cellulases Human genes 0.000 description 1
- XDTMQSROBMDMFD-UHFFFAOYSA-N Cyclohexane Chemical compound C1CCCCC1 XDTMQSROBMDMFD-UHFFFAOYSA-N 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 108010073178 Glucan 1,4-alpha-Glucosidase Proteins 0.000 description 1
- 102100022624 Glucoamylase Human genes 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical group CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- XSTXAVWGXDQKEL-UHFFFAOYSA-N Trichloroethylene Chemical group ClC=C(Cl)Cl XSTXAVWGXDQKEL-UHFFFAOYSA-N 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 229910052784 alkaline earth metal Inorganic materials 0.000 description 1
- 150000001342 alkaline earth metals Chemical class 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 150000004945 aromatic hydrocarbons Chemical class 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 239000003729 cation exchange resin Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 239000004927 clay Substances 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- 238000004332 deodorization Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000003028 enzyme activity measurement method Methods 0.000 description 1
- 230000002349 favourable effect Effects 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000003599 food sweetener Nutrition 0.000 description 1
- 150000008282 halocarbons Chemical class 0.000 description 1
- DMEGYFMYUHOHGS-UHFFFAOYSA-N heptamethylene Natural products C1CCCCCC1 DMEGYFMYUHOHGS-UHFFFAOYSA-N 0.000 description 1
- 108010025730 hydrolase C Proteins 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 125000003071 maltose group Chemical group 0.000 description 1
- GYNNXHKOJHMOHS-UHFFFAOYSA-N methyl-cycloheptane Natural products CC1CCCCCC1 GYNNXHKOJHMOHS-UHFFFAOYSA-N 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- 239000012044 organic layer Substances 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- BHZOKUMUHVTPBX-UHFFFAOYSA-M sodium acetic acid acetate Chemical compound [Na+].CC(O)=O.CC([O-])=O BHZOKUMUHVTPBX-UHFFFAOYSA-M 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 238000013519 translation Methods 0.000 description 1
- UBOXGVDOUJQMTN-UHFFFAOYSA-N trichloroethylene Natural products ClCC(Cl)Cl UBOXGVDOUJQMTN-UHFFFAOYSA-N 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Description
【発明の詳細な説明】
［産業上の利用分野］
本発明はイソマルトオリゴ糖の製造方法に関するもので
、さらに詳しくは甘味を有する食品や保湿性、老化防止
、制菌性などを有する健康食品に使用されるイソマルト
オリゴ糖の製造方法に関するものである。[Detailed Description of the Invention] [Industrial Application Field] The present invention relates to a method for producing isomalto-oligosaccharide, and more specifically to a method for producing isomaltooligosaccharide. The present invention relates to a method for producing the isomalto-oligosaccharide used.
［従来の技術］
従来より糖類は、食品工業における重要な甘味料として
広く使用されているが、最近は微生物白米の酵素を利用
した生理活性を有する新しい糖類の開発が相次いで行わ
れている。中でも特にオリゴ糖の研究開発は盛んで例え
ば、イソマルトオリゴ糖を製造する方法については種々
の提案がされている。[Prior Art] Saccharides have traditionally been widely used as important sweeteners in the food industry, but recently new saccharides with physiological activity using microbial white rice enzymes have been developed one after another. Among these, research and development on oligosaccharides is particularly active, and various proposals have been made regarding, for example, methods for producing isomalto-oligosaccharides.
例えば特開昭５８−７２５９８にグルコアミラーゼによ
るグルコース逆合成反応過程でイソマルトースを含有し
た原糖液をアルカリまたはアルカリ土類金属型強酸性カ
チオン交換樹脂に通し、その後樹脂に吸着したイソマル
トースを水で溶出し、高純度で得る方法が開示されてい
る。For example, in JP-A-58-72598, in the glucose retrosynthesis reaction process using glucoamylase, a raw sugar solution containing isomaltose is passed through an alkaline or alkaline earth metal type strongly acidic cation exchange resin, and then the isomaltose adsorbed on the resin is hydrated. A method for obtaining high purity by elution is disclosed.
［発明が解決しようとする課題］
ところが、上記特開昭５８−７２５９８の方法は水中で
の酵素反応で、反応速度が遅いため長時間を要し、また
多数の副生成物が生成し、煩雑な精製過程が必要で目的
物の生産効率が悪いという問題があった。[Problems to be Solved by the Invention] However, the method of JP-A-58-72598 involves an enzymatic reaction in water, which requires a long time due to the slow reaction rate, and produces many by-products, making it complicated. There was a problem that the production efficiency of the target product was low because it required a long purification process.
本発明は上記の問題点に鑑み、短い反応時間で、高収率
、高濃度かつ、精製過程が簡単なイソマルトオリゴ糖の
製造方法を提供することを目的とするものである。In view of the above-mentioned problems, it is an object of the present invention to provide a method for producing isomalto-oligosaccharides, which requires a short reaction time, has a high yield, is highly concentrated, and has a simple purification process.
［課題を解決するための手段］
本発明は、水を疎水性有機溶剤中に分散させた系におい
てグルコースまたはマルトースを加水分解酵素による酵
素転移反応で、イソマルトオリゴ糖に変換することを特
徴とするイソマルトオリゴ糖の製造方法を提供するもの
である。すなわち、本発明は反応系が水−疎水性有機溶
剤の二相の撹拌分散系であること、及び加水分解酵素を
使用することを特徴とするものである。本発明において
グルコースを基質として反応させた場合、主としてイソ
マルトオリゴ糖が生成され、またマルトースを基質とし
て反応させた場合、イソマルトトリオースオリゴ糖が生
成される。[Means for Solving the Problems] The present invention is characterized in that glucose or maltose is converted into isomalto-oligosaccharide by an enzymatic transfer reaction using a hydrolase in a system in which water is dispersed in a hydrophobic organic solvent. A method for producing isomalto-oligosaccharide is provided. That is, the present invention is characterized in that the reaction system is a two-phase stirred dispersion system of water and a hydrophobic organic solvent, and in that a hydrolytic enzyme is used. In the present invention, when the reaction is carried out using glucose as a substrate, isomalto-oligosaccharides are mainly produced, and when the reaction is carried out using maltose as a substrate, isomaltotriose-oligosaccharides are produced.
イソマルトオリゴ糖とは、単糖類であるブドウ糖がα−
１，６結合、α−１，２結合、α−１，３結合というα
−１，４結合以外の結合部分を有し、重合度２〜４を主
体とし、重合度１０以下までのオリゴ糖を総称する。Isomalto-oligosaccharide is a monosaccharide glucose that is α-
α 1,6 bond, α-1,2 bond, α-1,3 bond
A general term for oligosaccharides that have a bonding moiety other than a -1,4 bond and have a polymerization degree of 2 to 4, with a polymerization degree of 10 or less.
本発明において、疎水性有機溶剤として水と相溶性ない
ものであればいずれも使用可能であるが、具体的にはｎ
−ペンタン、２−メチルブタン、２−メチルペンタン、
２．２−ジメチルブタン、２，３−ジメチルブタン、ヘ
プタン、オクタン、２−メチルへブタン、２，２．３−
トリメチルへブタン、２．２．４−トリメチルへブタン
、ノナン、デカン、ウンデカン、ドデカンなどの鎖状飽
和炭化水素系溶剤；シクロペンタン、メチルシクロブタ
ン、シクロヘキサン、メチルシクロヘキサンなどの環状
飽和炭化水素系溶剤；ベンゼン、Ｏ−キシレン、ｍ−キ
シレン、ｐ−キシレン、エチルベンゼン、クメンなどの
芳香族炭化水素系溶剤；石油エーテル、軽ベンジン、リ
グロインなどの石油留分として得られる溶剤；ジクロル
メタン、クロロホルム、四塩化炭素、１．２−ジクロル
エタン、１，１，２．２−テトラクロルエタン、トリク
ロルエチレン、クロルベンゼン、２．６−ジクロルベン
ゼンなどのハロゲン化炭化水素系溶剤などが使用可能で
あるが、好ましくはペンタン、ヘキサン、ヘプタン、オ
クタン、２−メチルへブタン、ドデカンなどの鎖状飽和
炭化水素系溶剤である。In the present invention, any hydrophobic organic solvent that is incompatible with water can be used, but specifically, n
-pentane, 2-methylbutane, 2-methylpentane,
2.2-dimethylbutane, 2,3-dimethylbutane, heptane, octane, 2-methylhebutane, 2,2.3-
Chain saturated hydrocarbon solvents such as trimethylhebutane, 2.2.4-trimethylhebutane, nonane, decane, undecane, and dodecane; Cyclic saturated hydrocarbon solvents such as cyclopentane, methylcyclobutane, cyclohexane, and methylcyclohexane; Aromatic hydrocarbon solvents such as benzene, O-xylene, m-xylene, p-xylene, ethylbenzene, and cumene; Solvents obtained as petroleum fractions such as petroleum ether, light benzene, and ligroin; dichloromethane, chloroform, carbon tetrachloride , 1,2-dichloroethane, 1,1,2,2-tetrachloroethane, trichloroethylene, chlorobenzene, 2,6-dichlorobenzene, and other halogenated hydrocarbon solvents can be used, but preferably A linear saturated hydrocarbon solvent such as pentane, hexane, heptane, octane, 2-methylhebutane, and dodecane.
本発明で用いる酵素はセルロース加水分解酵素Ｃ以下セ
ルラーゼという）であり、アスペルギルス・ニガー（Ａ
ｓｐｅｒｇｉｌｌｕｓ ｎｉｇｅｒ）などの由来のセル
ラーゼを使用することができる。特にＡｓｐｅｒｇｉ−
ｆｌｕｓ ｎｉｇｅｒに由来するセルラーゼは安価であ
り、入手し易くかつ加水分解能力も高いため好ましく用
いられる。水層は緩衝溶液を用い緩衝溶液としては、酢
酸−酢酸ナトリウム系が好ましく使用される。緩衝溶液
の疎水性有機溶剤に対する割合は、２０ｖｏ１％以下が
好ましく、２０ｖｏ１％以上になるとイソマルトオリゴ
糖の生成量が低下する。水の容量が少ないほどイソマル
トオリゴ糖の生産比が増加する。緩衝溶液のイオン強度
は１Ｍ付近が好ましいがより低い値でもよく、本発明で
はこの値に制限されるものではない。The enzyme used in the present invention is cellulose hydrolase C (hereinafter referred to as cellulase), which is produced by Aspergillus niger (A
Cellulases derived from, for example, Spergillus niger can be used. Especially Aspergi-
Cellulase derived from S. flus niger is preferably used because it is inexpensive, easily available, and has a high hydrolytic ability. A buffer solution is used for the aqueous layer, and an acetic acid-sodium acetate system is preferably used as the buffer solution. The ratio of the buffer solution to the hydrophobic organic solvent is preferably 20 vol % or less, and when it is 20 vol % or more, the amount of isomalto-oligosaccharide produced decreases. The smaller the water capacity, the higher the production ratio of isomalto-oligosaccharides. The ionic strength of the buffer solution is preferably around 1M, but a lower value may be used, and the present invention is not limited to this value.
酵素反応の至適温度は水系で３７℃付近の狭い温度範囲
であるが、本発明では７０℃以下の広い温度範囲で反応
が進行し、好ましくは１０〜６５℃、より好ましくは３
０〜６０℃である。水の容量が少ない撹拌分散状態では
、０℃以下でも反応系が凍結することがないため、例え
ば−１０℃でも反応させることが可能である。この場合
、水がｂｏｕｎｄ Ｗａｔｅｒとして系内に存在するた
めと考えられる。The optimal temperature for the enzyme reaction is a narrow temperature range of around 37°C in an aqueous system, but in the present invention, the reaction proceeds in a wide temperature range of 70°C or less, preferably 10 to 65°C, more preferably 37°C or less.
The temperature is 0 to 60°C. In a stirred and dispersed state with a small volume of water, the reaction system does not freeze even at 0°C or lower, so it is possible to carry out the reaction even at -10°C, for example. In this case, it is considered that water exists in the system as bound water.
本発明の酵素反応に適したｐｉ（は３．５〜７．０であ
る。ｐＨ値は緩衝溶液によって容易に調整することがで
きる。A suitable pi (pi) for the enzyme reaction of the present invention is 3.5 to 7.0. The pH value can be easily adjusted with a buffer solution.
反応は、例えば疎水性有機溶剤中に撹拌しながらグルコ
ースまたはマルトースを分散し、次いで所定のｐＨ値の
緩衝溶液中にセルラーゼを溶解させ、この酵素溶液を前
記の有機溶剤中に添加して、所定の温度にて撹拌しなが
ら、分散させることにより行われる。The reaction can be carried out, for example, by dispersing glucose or maltose in a hydrophobic organic solvent with stirring, then dissolving cellulase in a buffer solution with a predetermined pH value, and adding this enzyme solution into the organic solvent. It is carried out by dispersing while stirring at a temperature of .
反応後、撹拌を止めて放置すると有機層と水層とが相分
離するので、その水層をとり濃縮することにより生成物
を高濃度で含有する反応液が得られる。従来、このよう
な処理は雑菌の侵入を防止するために密閉系で行わなけ
ればならなかったが、本発明では疎水性有機溶剤を用い
るため、反応系の滅菌の必要がなくなり、反応系が系外
から侵入する雑菌から遮断され、雑菌による汚染が防止
される。また、有機溶剤臭を除（ため分離された水層を
活性炭や活性白土などの吸着処理剤による脱臭処理を施
してもよい。After the reaction, if the stirring is stopped and the mixture is allowed to stand, the organic layer and the aqueous layer will phase separate, and by removing and concentrating the aqueous layer, a reaction solution containing a high concentration of the product can be obtained. Conventionally, such treatment had to be carried out in a closed system to prevent the intrusion of germs, but since the present invention uses a hydrophobic organic solvent, there is no need to sterilize the reaction system. It is blocked from germs that invade from outside, and contamination by germs is prevented. Further, to remove the odor of the organic solvent, the separated water layer may be subjected to deodorization treatment using an adsorption treatment agent such as activated carbon or activated clay.
なお、反応系中に界面活性剤を加えると微細な水滴が疎
水性有機溶剤中に分散されて反応速度が上昇するので界
面活性剤の使用は本発明を妨げるものではない。Note that, when a surfactant is added to the reaction system, fine water droplets are dispersed in the hydrophobic organic solvent and the reaction rate increases, so the use of a surfactant does not hinder the present invention.
また、本発明において、活性炭やイオン交換樹脂を共存
させて酵素反応を進めると、分子量が大きいものほど、
吸着され易いため平衡がイソマルトオリゴ糖生成に有利
な方向に傾き、好ましい結果を与える。この場合反応後
に活性炭やイオン交換樹脂から、水性アルコール溶液な
どでイソマルトオリゴ糖を脱離し回収することができる
。In addition, in the present invention, when the enzyme reaction is carried out in the presence of activated carbon or ion exchange resin, the larger the molecular weight, the more
Because it is easily adsorbed, the equilibrium is tilted in favor of isomalto-oligosaccharide production, giving favorable results. In this case, after the reaction, the isomalto-oligosaccharide can be removed and recovered from the activated carbon or ion exchange resin using an aqueous alcohol solution.
［実施例］ 以下、実施例を挙げて本発明を説明する。[Example] The present invention will be explained below with reference to Examples.
実施例１
５００ｍｌのバッフルプレート付き三角フラスコにドデ
カン２９１ｍ１とグルコース９ｇを入れ撹拌しながら分
散させた。一方、セルラーゼとしては市販のアスペルギ
ルス・ニガー（Ａｓｐｅｒｇｉｌｌｕｓｎｉｇｅｒ ）
由来のセルラーゼを用いて酢酸緩衝溶液（ｐＨ５，０、
イオン強度ＬＭ）９ｍｌ中に０．３ｇ（２４０ｕ）を溶
解させ、この酵素水溶液を上記の三角フラスコに添加し
て、撹拌（６００〜１００１０００ｒｐ、なから４０℃
で酵素反応を行った。生成した糖の組成（％）は高速液
体クロマトグラフィー（ＨＰＬＣ、カラム； Ｂｉｏ
Ｒａｄ ＨＰＸ８７Ｃ）により下記測定条件で定量した
。Example 1 291 ml of dodecane and 9 g of glucose were placed in a 500 ml Erlenmeyer flask with a baffle plate and dispersed with stirring. On the other hand, as a cellulase, commercially available Aspergillus niger
Using cellulase derived from
Dissolve 0.3 g (240 u) in 9 ml of ionic strength LM), add this enzyme aqueous solution to the above Erlenmeyer flask, and stir (600-1001000 rpm, from 40°C
An enzymatic reaction was performed. The composition (%) of the sugar produced was determined by high performance liquid chromatography (HPLC, column; Bio
Rad HPX87C) under the following measurement conditions.
ＨＰＬＣの測定条件
ＨＰＬＣ、水系
カラム；Ｂｉｏ Ｒａｄ ＨＰＸ８７Ｃ移動層；蒸留
水
流 速；０．６ｍｌ／分
カラム温度；８５℃
検出方法、ＲＩ検出器
また酵素活性の測定はｐ−ニトロフェノールで修飾され
たアナログ基質を用いてＵＶ測定により行った。酵素反
応における糖の組成の経時変化を第１図に示した。経過
時間とともにオリゴ糖の生成が増加し、反応時間１６０
ｈｒ経過後、イソマルトースなどオリゴ糖が２３％を示
した（％はＨＰＬＣのピーク面積の比）。また、反応時
間１６６ｈｒ経過後、有機溶剤中の酵素活性を調べた結
果、同じ条件の水層中の酵素活性（１００％）と比べて
８８．４％活性が残存していることがわかった。すなわ
ち、水−疎水性有機溶剤系において長時間にわたって酵
素活性が持続しており、酵素が繰り返し使用できること
がわかった。HPLC measurement conditions HPLC, aqueous column; Bio Rad HPX87C mobile phase; distilled water flow rate; 0.6 ml/min Column temperature: 85°C Detection method, RI detector and enzyme activity measurement using p-nitrophenol-modified analogs. This was carried out by UV measurement using a substrate. Figure 1 shows the change in sugar composition over time during the enzymatic reaction. The production of oligosaccharides increases with the passage of time, and the reaction time is 160
After hours, oligosaccharides such as isomaltose showed 23% (% is the ratio of HPLC peak areas). Furthermore, after a reaction time of 166 hours, the enzyme activity in the organic solvent was examined, and it was found that 88.4% of the enzyme activity remained compared to the enzyme activity (100%) in the aqueous layer under the same conditions. That is, it was found that the enzyme activity persisted for a long time in the water-hydrophobic organic solvent system, and the enzyme could be used repeatedly.
一方、水系中の場合は、このようなオリゴ糖が生成され
なかった。On the other hand, such oligosaccharides were not produced in an aqueous system.
実施例２
ドデカン１９１０１１、セルラーゼ０．２ｇ、緩衝溶液
９ｍｌ、マルトース９ｇ、反応温度４０℃で他の条件は
実施例１と同様にして生成した糖の組成の経時変化を第
２図に示した。反応時間５５時間のＨＰＬＣの結果を第
３図に示した。Example 2 Dodecane 191011, cellulase 0.2 g, buffer solution 9 ml, maltose 9 g, reaction temperature 40° C., and the other conditions were the same as in Example 1. Changes in the composition of sugar produced over time are shown in FIG. The results of HPLC after a reaction time of 55 hours are shown in FIG.
反応時間２０時間後、グルコース３８％、マルトースま
たはイソマルトース３６％、イソマルトトリオース２５
％、イソマルトテトラオース１％を示した（％はＨＰＬ
Ｃのピーク面積の比）。After 20 hours of reaction time, glucose 38%, maltose or isomaltose 36%, isomaltotriose 25%
%, isomaltotetraose 1% (% is HPL
(ratio of peak areas of C).
第２図からイソマルトトリオースは２Ｏｈｒ後にピーク
を示し、その後徐々に減少していき、一方グルコースと
イソアミロテトラオースは増加し続け、マルトースは次
第に減少することがわかる。From FIG. 2, it can be seen that isomaltotriose shows a peak after 2 Ohr and then gradually decreases, while glucose and isoamylotetraose continue to increase, and maltose gradually decreases.
このことから、反応時間をコントロールすることにより
生成物の成分比率を制御することが可能である。From this, it is possible to control the component ratio of the product by controlling the reaction time.
［発明の効果］
本発明の方法での、水と疎水性有機溶剤の二相の撹拌分
散系で反応させることにより、グルコースやマルトース
のように安価な原料からイソマルトオリゴ糖を高収率で
製造することが可能となった。反応速度も短縮され、例
えば基質がマルトースの場合、短時間で、生成物を選択
的に高濃度で得ることができる。それにより煩雑な精製
過程が不要であり操作が簡単にできる。さらに有機溶剤
を用いるため反応系の滅菌の必要がなく、反応系が雑菌
から遮断され汚染される心配がない。また、反応時間に
より生成物の成分比が異なるため、反応の進行に従って
任意の目的物を選んで得ることができる。[Effects of the invention] Isomalto-oligosaccharides can be produced in high yield from inexpensive raw materials such as glucose and maltose by reacting in a two-phase stirred dispersion system of water and a hydrophobic organic solvent using the method of the present invention. It became possible to do so. The reaction rate is also shortened; for example, when the substrate is maltose, the product can be selectively obtained in a high concentration in a short time. This eliminates the need for complicated purification processes and allows for easy operation. Furthermore, since an organic solvent is used, there is no need to sterilize the reaction system, and there is no need to worry about the reaction system being isolated from germs and being contaminated. Furthermore, since the component ratio of the product varies depending on the reaction time, any target product can be selected and obtained according to the progress of the reaction.
第１図は実施例１の酵素反応における各成分の経時変化
を示すグラフであり、第２図は実施例２の酵素反応にお
ける各成分の経時変化を示すグラフである。
第３図は実施例２の反応時間５５ｈｒ後の反応溶液のＨ
ＰＬＣのチャートを示したものである。
出願人 株式会社フォークウェイズ・ジャパン代理人
弁理士 舘 野 公
衡！ｉｇ蛮（訳）
時間（ｈｒ）
保持時間FIG. 1 is a graph showing the change over time of each component in the enzyme reaction of Example 1, and FIG. 2 is a graph showing the change over time of each component in the enzyme reaction of Example 2. Figure 3 shows the H of the reaction solution after a reaction time of 55 hours in Example 2.
This shows a PLC chart. Applicant Folkways Japan Co., Ltd. Agent
Patent attorney Kimihira Tateno! igban (translation) time (hr) retention time
Claims (1)
ースまたはマルトースを加水分解酵素による酵素転移反
応でイソマルトオリゴサッカライドに変換することを特
徴とするイソマルトオリゴ糖の製造方法。1. A method for producing isomalto-oligosaccharides, which comprises converting glucose or maltose into isomalto-oligosaccharide by an enzymatic transfer reaction using a hydrolase in a system in which water is dispersed in a hydrophobic organic solvent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP23188790A JPH04112797A (en) | 1990-08-31 | 1990-08-31 | Production of isomaltooligosaccharide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP23188790A JPH04112797A (en) | 1990-08-31 | 1990-08-31 | Production of isomaltooligosaccharide |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH04112797A true JPH04112797A (en) | 1992-04-14 |
Family
ID=16930590
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP23188790A Pending JPH04112797A (en) | 1990-08-31 | 1990-08-31 | Production of isomaltooligosaccharide |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH04112797A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1078615C (en) * | 1998-08-06 | 2002-01-30 | 福州大学 | Method for continuous preparation of isomalt oligose by use of hollow fiber enzyme membrane reactor |
| KR100387286B1 (en) * | 2000-06-30 | 2003-06-12 | 주식회사 삼양제넥스 | Method for production of isomaltooligosaccharides |
-
1990
- 1990-08-31 JP JP23188790A patent/JPH04112797A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1078615C (en) * | 1998-08-06 | 2002-01-30 | 福州大学 | Method for continuous preparation of isomalt oligose by use of hollow fiber enzyme membrane reactor |
| KR100387286B1 (en) * | 2000-06-30 | 2003-06-12 | 주식회사 삼양제넥스 | Method for production of isomaltooligosaccharides |
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