JPS63216492A - Production of neotrehalose and centose - Google Patents
Production of neotrehalose and centoseInfo
- Publication number
- JPS63216492A JPS63216492A JP4878587A JP4878587A JPS63216492A JP S63216492 A JPS63216492 A JP S63216492A JP 4878587 A JP4878587 A JP 4878587A JP 4878587 A JP4878587 A JP 4878587A JP S63216492 A JPS63216492 A JP S63216492A
- Authority
- JP
- Japan
- Prior art keywords
- neotrehalose
- centose
- starch
- produced
- sugar
- 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.)
- Granted
Links
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 title claims abstract description 42
- HDTRYLNUVZCQOY-BTLHAWITSA-N alpha,beta-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-BTLHAWITSA-N 0.000 title claims abstract description 42
- JKMASURNTJPVIE-UHFFFAOYSA-N Centose Natural products OCC(OC1OC(CO)C(O)C(O)C1O)C(O)C(O)C(OC2OC(CO)C(O)C(O)C2O)C=O JKMASURNTJPVIE-UHFFFAOYSA-N 0.000 title claims abstract description 40
- 238000004519 manufacturing process Methods 0.000 title claims description 15
- 235000000346 sugar Nutrition 0.000 claims abstract description 32
- 229920000858 Cyclodextrin Polymers 0.000 claims abstract description 18
- 239000000203 mixture Substances 0.000 claims abstract description 17
- 108010073178 Glucan 1,4-alpha-Glucosidase Proteins 0.000 claims abstract description 14
- 102100022624 Glucoamylase Human genes 0.000 claims abstract description 14
- 229920002472 Starch Polymers 0.000 claims abstract description 12
- 235000019698 starch Nutrition 0.000 claims abstract description 12
- 239000000758 substrate Substances 0.000 claims abstract description 12
- 239000008107 starch Substances 0.000 claims abstract description 11
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims abstract description 9
- 238000004440 column chromatography Methods 0.000 claims abstract description 8
- 238000000354 decomposition reaction Methods 0.000 claims abstract description 6
- HFHDHCJBZVLPGP-UHFFFAOYSA-N schardinger α-dextrin Chemical compound O1C(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC(C(O)C2O)C(CO)OC2OC(C(C2O)O)C(CO)OC2OC2C(O)C(O)C1OC2CO HFHDHCJBZVLPGP-UHFFFAOYSA-N 0.000 claims description 9
- 239000003513 alkali Substances 0.000 claims description 5
- 108090000790 Enzymes Proteins 0.000 abstract description 13
- 102000004190 Enzymes Human genes 0.000 abstract description 13
- 229940088598 enzyme Drugs 0.000 abstract description 13
- 235000013305 food Nutrition 0.000 abstract description 5
- 229920001592 potato starch Polymers 0.000 abstract description 4
- 102000004139 alpha-Amylases Human genes 0.000 abstract description 2
- 108090000637 alpha-Amylases Proteins 0.000 abstract description 2
- 229940024171 alpha-amylase Drugs 0.000 abstract description 2
- 102000003960 Ligases Human genes 0.000 abstract 2
- 108090000364 Ligases Proteins 0.000 abstract 2
- 150000001875 compounds Chemical class 0.000 abstract 1
- 238000007796 conventional method Methods 0.000 abstract 1
- 239000002994 raw material Substances 0.000 abstract 1
- 150000008163 sugars Chemical class 0.000 description 11
- 238000006243 chemical reaction Methods 0.000 description 10
- 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 8
- 239000008103 glucose Substances 0.000 description 8
- 238000004458 analytical method Methods 0.000 description 7
- 238000004128 high performance liquid chromatography Methods 0.000 description 7
- 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 description 6
- 239000001116 FEMA 4028 Substances 0.000 description 6
- 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 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- WHGYBXFWUBPSRW-FOUAGVGXSA-N beta-cyclodextrin Chemical compound OC[C@H]([C@H]([C@@H]([C@H]1O)O)O[C@H]2O[C@@H]([C@@H](O[C@H]3O[C@H](CO)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](CO)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](CO)[C@H]([C@@H]([C@H]3O)O)O[C@H]3O[C@H](CO)[C@H]([C@@H]([C@H]3O)O)O3)[C@H](O)[C@H]2O)CO)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@@H]3O[C@@H]1CO WHGYBXFWUBPSRW-FOUAGVGXSA-N 0.000 description 6
- 235000011175 beta-cyclodextrine Nutrition 0.000 description 6
- 229960004853 betadex Drugs 0.000 description 6
- 238000004816 paper chromatography 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
- 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 description 4
- 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 description 4
- 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 3
- 238000010586 diagram Methods 0.000 description 3
- 238000010828 elution Methods 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 238000006491 synthase reaction Methods 0.000 description 3
- 150000004043 trisaccharides Chemical class 0.000 description 3
- QIGJYVCQYDKYDW-UHFFFAOYSA-N 3-O-alpha-D-mannopyranosyl-D-mannopyranose Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(CO)OC(O)C1O QIGJYVCQYDKYDW-UHFFFAOYSA-N 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical class OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 2
- -1 G-(141)-β-ri-G Chemical compound 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 102100024295 Maltase-glucoamylase Human genes 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 108010028144 alpha-Glucosidases Proteins 0.000 description 2
- 238000009835 boiling Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000003456 ion exchange resin Substances 0.000 description 2
- 229920003303 ion-exchange polymer Polymers 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 230000011987 methylation Effects 0.000 description 2
- 238000007069 methylation reaction Methods 0.000 description 2
- QIGJYVCQYDKYDW-NSYYTRPSSA-N nigerose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@H]1[C@H](O)[C@@H](CO)OC(O)[C@@H]1O QIGJYVCQYDKYDW-NSYYTRPSSA-N 0.000 description 2
- 229920001542 oligosaccharide Polymers 0.000 description 2
- 150000002482 oligosaccharides Chemical class 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 235000011121 sodium hydroxide Nutrition 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 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 1
- 229920001353 Dextrin Polymers 0.000 description 1
- 239000004375 Dextrin Substances 0.000 description 1
- 101100379080 Emericella variicolor andB gene Proteins 0.000 description 1
- 240000008620 Fagopyrum esculentum Species 0.000 description 1
- 235000009419 Fagopyrum esculentum Nutrition 0.000 description 1
- 229930091371 Fructose Natural products 0.000 description 1
- 239000005715 Fructose Substances 0.000 description 1
- 240000003183 Manihot esculenta Species 0.000 description 1
- 235000016735 Manihot esculenta subsp esculenta Nutrition 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 241000178960 Paenibacillus macerans Species 0.000 description 1
- 244000061456 Solanum tuberosum Species 0.000 description 1
- 235000002595 Solanum tuberosum Nutrition 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- LFYJSSARVMHQJB-QIXNEVBVSA-N bakuchiol Chemical compound CC(C)=CCC[C@@](C)(C=C)\C=C\C1=CC=C(O)C=C1 LFYJSSARVMHQJB-QIXNEVBVSA-N 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 239000000969 carrier Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 235000009508 confectionery Nutrition 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 229940097362 cyclodextrins Drugs 0.000 description 1
- 230000009849 deactivation Effects 0.000 description 1
- 238000010612 desalination reaction Methods 0.000 description 1
- 235000019425 dextrin Nutrition 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 235000021255 galacto-oligosaccharides Nutrition 0.000 description 1
- 150000003271 galactooligosaccharides Chemical class 0.000 description 1
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 235000012907 honey Nutrition 0.000 description 1
- 230000000415 inactivating effect Effects 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 235000014347 soups Nutrition 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Description
【発明の詳細な説明】
［産業上の利用分野コ
本発明は、ネオトレハロースおよびセントースまたは両
者を主として含む糖混合物の製法に関する。DETAILED DESCRIPTION OF THE INVENTION [Field of Industrial Application] The present invention relates to a method for producing a sugar mixture containing primarily neotrehalose and centose or both.
［従来の技術］
最近、マルトオリゴ糖、イソマルトース等の分岐オリゴ
糖、フラクトオリゴ糖、ガラクトオリゴ糖、その他各種
オリゴ糖が開発され、ネオトレハロース、セントースに
ついても、その大量製造法の開発が強く望まれてきた。[Prior Art] Recently, branched oligosaccharides such as maltooligosaccharides and isomaltose, fructooligosaccharides, galactooligosaccharides, and other various oligosaccharides have been developed, and there is a strong desire to develop a method for mass production of neotrehalose and centose. Ta.
これら二種糖の中、ネオトレハロースは還元末端をもた
ず、安定な糖であると考えられ、しょ糖の０−α−Ｄ−
Ｇ−（１→２）−β−Ｄ−Ｆと構造的にも０−α−Ｄ−
Ｇ−（１４１）−β−り−Ｇのように、グルコースとフ
ラクトースの差異はあるものの、類似している。Among these disaccharides, neotrehalose does not have a reducing end and is considered to be a stable sugar.
G-(1→2)-β-D-F and structurally 0-α-D-
Although there are differences between glucose and fructose, such as G-(141)-β-ri-G, they are similar.
一方、セントースはマルトースとコージビオース部分を
一分子内にもつ２Ｉ−α−グルコシル−マルトースの構
造をもっている。On the other hand, centose has a 2I-α-glucosyl-maltose structure containing maltose and cordibiose moieties in one molecule.
これらが大量生産されれば、食品用として有用な素材と
なりうる。If these can be mass-produced, they can become useful materials for food.
しかし、ネオトレハロースは麹汁中に含まれていること
が知られているのみで、その製造法については知られて
いず、ましてや大量に得た例はなく、したがって利用の
可能性については未知であフだ、また、セントースも特
殊な糖であり、これまで調製例が二件知られているにす
ぎない、しかも、セントースは蜂蜜中に含まれているが
、その含有量は０．０６％と少ない（１，Ｒ，５ｉｄｄ
ｉｑｕｉ ａｎｄＢ、Ｆｕｒｇａｌａ ： Ｃａｒｂｏ
ｈｙｄ、Ｒｅｓ、、６（１９６８）２５０−２５２）。However, it is only known that neotrehalose is contained in koji soup, but the production method is not known, much less it has never been obtained in large quantities, so the possibility of its use is unknown. Also, centose is a special sugar, and only two examples of its preparation are known so far.Furthermore, centose is contained in honey, but its content is 0.06%. and less (1,R,5idd
iqui andB, Furgala: Carbo
hyd, Res., 6 (1968) 250-252).
その後、蕎麦のα−グルコシダーゼを用い、マルトース
から１％の収率で得た例が報告されているくに、Ｎ１５
ｈｉ、Ｓ、Ｃｈｉｂａ ａｎｄ Ｔ、Ｓｈｉｍｏａ＋ｕ
ｒａ：Ａｇｒ、Ｂｉｏｌ。Subsequently, it has been reported that buckwheat α-glucosidase was obtained from maltose with a yield of 1%.
hi, S, Chiba and T, Shimoa+u
ra: Agr, Biol.
Ｃｈｅａｐ、 、３９（１９７５）７２７−７２８）が
、生成率が低く、調製法も容易でなかったために、その
応用は考えられなかった。Cheap, 39 (1975) 727-728), but its application was not considered because the production rate was low and the preparation method was not easy.
［発明が解決しようとする問題点］
従来、グルコアミラーゼ、α−グルコシダーゼの逆反応
により、マルトース、イソマルトース、ニゲロース、コ
ージビオースの三糖類が生成することは知られていたが
ネオトレハロースを酵素により大量に生成させることは
知られていなかった。[Problems to be solved by the invention] It has been known that the trisaccharides of maltose, isomaltose, nigerose, and cordibiose are produced by the reverse reaction of glucoamylase and α-glucosidase. It was not known that it could be produced.
さらに、これらの三糖類にサイクロデキストリン合成酵
素のカップリング反応により各種の三糖類が生成される
が、セントースを合成した例は知られていない。Furthermore, various trisaccharides are produced by the coupling reaction of cyclodextrin synthase to these trisaccharides, but there are no known examples of synthesizing centose.
本発明では、これらの生産が困難な糖を可及的容易に生
産して安価となし、その食品用素材としての利用を可能
とすることを目的とする。The purpose of the present invention is to produce these sugars, which are difficult to produce, as easily as possible and make them inexpensive, so that they can be used as food materials.
［問題点を解決するための手段］
そこで本発明者らは、サイクロデキストリン合成酵素を
澱粉等に強い反応条件下で作用させることにより、大量
のネオトレハロースとセントースが生成することを見い
出し、本発明を完成した。[Means for Solving the Problems] The present inventors have discovered that a large amount of neotrehalose and centose can be produced by allowing cyclodextrin synthase to act on starch, etc. under strong reaction conditions. completed.
本発明を以下に示す。The invention is illustrated below.
１）澱粉等の基質にサイクロデキストリン合成酵素を作
用させることを特徴とするネオトレハロースおよびセン
トース、またはこれらを含む糖混合物の製造法。1) A method for producing neotrehalose and centose, or a sugar mixture containing them, which comprises allowing cyclodextrin synthase to act on a substrate such as starch.
２）サイクロデキストリン合成酵素反応液にグルコアミ
ラーゼおよび／または酵母を作用させるネオトレハロー
スとセントースの製造法。2) A method for producing neotrehalose and centose in which glucoamylase and/or yeast are allowed to act on a cyclodextrin synthase reaction solution.
３）アルカリ処理によりネオトレハロース以外の還元糖
を分解除去するネオトレハロースの製造法。3) A method for producing neotrehalose in which reducing sugars other than neotrehalose are decomposed and removed by alkali treatment.
４）ネオトレハロースとセントースの混合物をカラムク
ロマトグラフィーにより分離する各々の製造法。4) Each production method involves separating a mixture of neotrehalose and centose by column chromatography.
本発明に用いる基質としては澱粉の他、その分解生成物
がある。詳しくは馬鈴薯、トウモロコシ、タピオカ等各
種澱粉やこれらの分解生成物、例えばグルコース、各種
デキストリン、サイクロデキストリン等があり、これら
の中の一種または二種以上の混合物を使用することがで
きる。Substrates used in the present invention include starch and its decomposition products. Specifically, there are various starches such as potato, corn, and tapioca, and their decomposition products, such as glucose, various dextrins, and cyclodextrins, and one type or a mixture of two or more of these can be used.
次に、上記基質に作用させる酵素としてはバチルス・マ
セランスの酵素（マセランス酵素）、バチルスΦメガテ
リウム、サーキュランス、ステアロサーモフィラス、好
アルカリ性バチルス属、り。Next, the enzymes to act on the substrate include Bacillus macerans enzyme (macerans enzyme), Bacillus Φ megaterium, Circulans, Stearothermophilus, alkalophilic Bacillus, and Ri.
レブシエラ・ニューモニア等のｙｌの酵素がある。There are yl enzymes such as Lebsiella pneumoniae.
反応条件は用いる酵素剤によって異なるが、例えばマセ
ランス酵素では、基質１ｇ当り１５０〜３００ＴＨＵ
（チルデン・ハドソン単位）を加え４０〜５０℃で２〜
５日反応により両成分で３０〜４０％の収率が得られる
。Reaction conditions vary depending on the enzyme agent used, but for example, for macerans enzyme, 150 to 300 THU per gram of substrate.
(Tilden-Hudson units) and heat at 40-50℃ for 2~
A yield of 30-40% is obtained for both components after 5 days of reaction.
β−サイクロデキストリン合成酵素等の転移作用の強い
酵素を用いれば４０〜５０％の収率が得られ、この他の
サイクロデキストリン合成酵素も同様に、本発明の方法
が適用できる。If an enzyme with a strong transfer effect such as β-cyclodextrin synthase is used, a yield of 40 to 50% can be obtained, and the method of the present invention can be similarly applied to other cyclodextrin synthases.
尚！（質濃度は任意に選べるが経済的には１５％以上の
濃度が好ましい。still! (The concentration can be arbitrarily selected, but from an economic standpoint, a concentration of 15% or more is preferable.
このようにして得られた反応液には目的とするネオトレ
ハロースおよび／またはセントースが含反応液の高速液
体クロマトグラフィー（ＨＰＬＣ）による分析によれば
ネオトレハロース、セントースが各々１８．６％、２０
．０％含まれていたく第１図参照）。The reaction solution thus obtained contains the desired neotrehalose and/or centose.According to analysis of the reaction solution by high performance liquid chromatography (HPLC), neotrehalose and centose are 18.6% and 20%, respectively.
．． (See Figure 1).
さらに、ネオトレハロース画分には、ペーパークロマト
グラフィー（Ｐ Ｃ）での分析によれば、約２０％のニ
ゲロース、コージビオース、イソマルトースが混入して
いるが、アルカリ分解によりネオトレハロース以外の還
元糖は除去できる。Furthermore, according to paper chromatography (PC) analysis, the neotrehalose fraction contains approximately 20% nigerose, cordibiose, and isomaltose, but due to alkaline decomposition, reducing sugars other than neotrehalose are removed. Can be removed.
また、セントース画分にも約２０％の七″ントース以外
の糖が含まれているが、グルコアミラーゼを作用させる
とセントースを水解しない条件でも水解されるので、純
品を得ることができる。The centose fraction also contains about 20% of sugars other than 7'ntose, but when glucoamylase is applied, centose can be hydrolyzed even under conditions that do not hydrolyze centose, so a pure product can be obtained.
サイクロデキストリン合成酵素反応液には五糖類以上の
糖もかなり含まれ、本反応液を熱失活した後、グルコア
ミラーゼを作用させると各種割合でグルコースを含むネ
オトレハロースとセントース含有糖液が得られるので、
目的に応じて、反応の各段階の糖液を製品とすることが
できる。The cyclodextrin synthase reaction solution contains a considerable amount of sugars higher than pentasaccharides, and after heat inactivation of this reaction solution, when glucoamylase is applied, a sugar solution containing neotrehalose and centose containing glucose in various proportions can be obtained. So,
Depending on the purpose, the sugar solution at each stage of the reaction can be used as a product.
また、サイクロデキストリン合成酵素反応液の酵素を失
活させた後、グルコアミラーゼと同時に酵母を添加して
、生成したグルコースを資化除去すると、ネオトレハロ
ースとセントースを８０％以上含む糖液が得られる（第
２図参照）。Furthermore, after inactivating the enzyme in the cyclodextrin synthase reaction solution, yeast is added at the same time as glucoamylase, and the produced glucose is assimilated and removed, resulting in a sugar solution containing more than 80% neotrehalose and centose. (See Figure 2).
尚、この反応で、ネオトレハロースが原糖液の１．３〜
１．５倍に増加するが、セントースは逆に０．８５倍と
なるので、ネオトレハロースの非還元末端の両端または
一端にグルコースが１個以上結合した糖がかなりの量存
在するものと考えられ、サイクロデキストリン合成酵素
の作用形式からも、これらの糖の存在は予想できる。In addition, in this reaction, neotrehalose is 1.3~
It increases by 1.5 times, but centose increases by 0.85 times, so it is thought that there is a considerable amount of sugar with one or more glucose attached to both or one end of the non-reducing end of neotrehalose. The presence of these sugars can also be predicted from the mode of action of cyclodextrin synthase.
さらに、セントースについても非還元末端にグルコース
が一個以上結合した糖の存在が予想される。Furthermore, regarding centose as well, it is expected that there is a sugar in which one or more glucose is bonded to the non-reducing end.
グルコアミラーゼ−酵母処理して得られた糖液から酵母
を除去し、イオン交換樹脂等により脱塩脱色すればネオ
トレハロースとセントースを主として含む糖製品が得ら
れる。本製品の糖組成の一例を示すと、ネオトレハロー
ス６２％、セントース２９％、その他の五糖類以上の糖
１９％であるが、反応条件を変化させることにより比率
を変えることができるので、目的に適合する条件を選択
すべきである。If yeast is removed from the sugar solution obtained by glucoamylase-yeast treatment and the sugar solution is desalted and decolorized using an ion exchange resin or the like, a sugar product containing mainly neotrehalose and centose can be obtained. An example of the sugar composition of this product is 62% neotrehalose, 29% centose, and 19% other sugars higher than pentasaccharides, but the ratio can be changed by changing the reaction conditions, so it is possible to You should choose the conditions that suit you.
また、ネオトレハロースのみを得たい場合には例えば本
製品に終濃度０．１規定の苛性ソーダな加え１５分、オ
ートクレーブした後、中和、カラムクロマトグラフィー
、溶媒沈澱法のいずれかまたは組み合せて分離すること
ができる。尚、加えるアルカリ、アルカリ濃度、加熱条
件に制限はないが、脱塩の負荷が少い方が望ましい、
。If you want to obtain only neotrehalose, for example, add caustic soda at a final concentration of 0.1N to this product, autoclave for 15 minutes, and then separate by neutralization, column chromatography, solvent precipitation, or any combination thereof. be able to. There are no restrictions on the alkali to be added, the alkali concentration, or the heating conditions, but it is preferable that the desalination load is small.
.
アルカリ処理液を中和した後、トヨパールＨＷ−４０Ｓ
のカラムクロマトグラフィーを行うと単一ピークが得ら
れ、着色物を完全に分離することができ（第３図参照）
、この画分を集めて濃縮すればＨＰＬＣ，ＰＣ分析で単
一ピークを示す純品が得られる。After neutralizing the alkaline treatment liquid, use TOYOPEARL HW-40S.
When column chromatography is performed, a single peak is obtained, and the colored substances can be completely separated (see Figure 3).
If these fractions are collected and concentrated, a pure product that shows a single peak in HPLC and PC analysis can be obtained.
尚、カラム担体としては、この他活性炭、イオン交換樹
脂等、各種のものが考えられる。In addition, various other types of column carriers can be considered, such as activated carbon and ion exchange resins.
セントースについては、グルコアミラーゼ処理液または
グルコアミラーゼ−酵母処理液をカラムクロマトグラフ
ィーにより分離しＨＰＬＣ，ＰＣ分析で単一ピークを示
す純品が得られる。Regarding centose, a pure product that shows a single peak in HPLC and PC analysis can be obtained by separating a glucoamylase-treated solution or a glucoamylase-yeast-treated solution by column chromatography.
このようにして得られた純品についてＳＩＭＳ。SIMS for the pure product thus obtained.
メチル化、ＮＭＲ分析、その他の一般分析をした結果、
ネオトレハロース画分（Ｉ）については、分子量３４２
、セントース画分（１１）については、５０４の値を得
た。メチル化分析では、■からは１．５−Ｄｉ−０−Ａ
ｃ−２，３，４，８−ｔｅｔｒａ−０−Ｍｅ−ｇｌｕｃ
ｉｔｏｌのみが生成し、この結果からゲルコニ糖類のｌ
、ｌ結合が確認された。１１からは２，４−Ｄｉ−０−
Ａｃ−１，３，５，６−ｔｅｔｒａ−０−Ｍｅ−ｇｌｕ
ｃｉｔｏｌと１．５−Ｄｉ−０−Ａｃ−２，３，４，６
−ｔｅ−ｔｒａ−０−Ｍｅ−ｇｌｕｃｉ ｔｏｌが１＝
２の割合で生成した。As a result of methylation, NMR analysis, and other general analyses,
For neotrehalose fraction (I), molecular weight 342
, a value of 504 was obtained for the centose fraction (11). In the methylation analysis, 1.5-Di-0-A from ■
c-2,3,4,8-tetra-0-Me-gluc
Only itol was produced, and from this result it was found that l of gelconisaccharides was
, l bond was confirmed. 11 to 2,4-Di-0-
Ac-1,3,5,6-tetra-0-Me-glu
citol and 1,5-Di-0-Ac-2,3,4,6
-te-tra-0-Me-glucitol=1
It was produced at a ratio of 2.
［α］。はＩで９１．６°、１１で１１９．８゜であり
、文献値と近似していた。[α]. were 91.6° for I and 119.8° for 11, which were close to the literature values.
以上の結果からｌはネオトレハロース、１１はセントー
スと決定された。From the above results, 1 was determined to be neotrehalose and 11 was determined to be centose.
この他の性質としては、■は全く還元力を示さず酸に対
して安定で塩酸終濃度０．０６規定４０℃、４時間放置
しても分解せず、沸騰水中１５分間放置で１１％、０．
５規定、沸騰水中１５分放置により６６％の分解率であ
る。As for other properties, (■) shows no reducing power at all, is stable against acids, has a final concentration of hydrochloric acid of 0.06N, does not decompose even after being left at 40°C for 4 hours, and 11% when left in boiling water for 15 minutes. 0.
5N, the decomposition rate is 66% when left in boiling water for 15 minutes.
グルコアミラーゼの作用に対しても抵抗性があり、イソ
マルトースより作用を受は難い。It is also resistant to the action of glucoamylase, and is less susceptible to the action than isomaltose.
１１は！よりアルカリ、酸に対する抵抗性は弱いが、グ
ルコアミラーゼに対する抵抗性は同等である。11 is! The resistance to alkaline and acid is weaker, but the resistance to glucoamylase is the same.
［実施例］ 次に本発明の実施例を示す。[Example] Next, examples of the present invention will be shown.
実施例１
馬鈴薯澱粉を耐熱性α−アミラーゼで液化し基質終濃度
１６％とし、β−サイクロデキストリン合成酵素（土壌
より分離した菌株の）を、基質ｇ当り１５０〜７５０Ｔ
ＨＵ加えて、ｐＨ６，０，４０℃で２日間反応した後失
活処理し、グルコアミラーゼを基質ｇ当り１００ＩＵ、
市販酵母２００ｍ ｇ 、炭酸カルシウム２０ ｍ ｇ
を加え、４０℃、２４時間反応し、生成したネオトレハ
ロースとセントースをＨＰ Ｌ Ｃで定Ａ（シた結果、
第４図のように３００ＴＨＵで両成分の生成率は４５％
に達っした。Example 1 Potato starch was liquefied with heat-stable α-amylase to give a final substrate concentration of 16%, and β-cyclodextrin synthase (from a strain isolated from soil) was added at a concentration of 150 to 750 T per g of substrate.
After adding HU and reacting at pH 6, 0 and 40°C for 2 days, deactivation treatment was performed, and glucoamylase was added at 100 IU per g of substrate.
Commercially available yeast 200 mg, calcium carbonate 20 mg
was added and reacted at 40°C for 24 hours, and the generated neotrehalose and centose were analyzed by HPLC at a constant A (as a result of
As shown in Figure 4, the production rate of both components is 45% at 300 THU.
reached.
実施例２
β−サイクロデキストリン＋グルコースまたはマルトー
ス（等量混合）、グルコース、マルトースまたは可溶性
澱粉各々単独の基質を用い、かつ酵素濃度を３００ＴＨ
Ｕとした以外は実施例１と同様にして酵素を作用させた
。Example 2 β-cyclodextrin + glucose or maltose (mixed in equal amounts), glucose, maltose or soluble starch were used as substrates, and the enzyme concentration was 300TH.
The enzyme was allowed to act in the same manner as in Example 1 except that U was used.
その結果、第５図に示したように、β−サイクロデキス
トリン＋マルトースの組合せのとき最も高い収率５５％
で糖混合物を得た。As a result, as shown in Figure 5, the combination of β-cyclodextrin and maltose had the highest yield of 55%.
A sugar mixture was obtained.
尚、市販の各種粉飴では可溶性澱粉と同等の収率であっ
た・
実施例３
マセランス酵素を用い、実施例２と同様に各種基質に作
用させたところ、可溶性澱粉の場合に最も高い収率４１
％でネオトレハロースとセントースを含む糖混合物を得
た。In addition, the yield of commercially available powdered candy was equivalent to that of soluble starch.Example 3 When the macerans enzyme was used to act on various substrates in the same manner as in Example 2, the highest yield was found for soluble starch. 41
A sugar mixture containing neotrehalose and centose in % was obtained.
実施例４
実施例１で得たネオトレハロースとセントースの混合物
（糖濃度１０％）に０．２規定の苛性ソーダを等量加え
、１５分間オートクレーブし、中和した後トヨパールＨ
Ｗ−４０Ｓに負荷して第３図のように分離し、ＨＰＬＣ
，ＰＣ分析で単一のピークを示すネオトレハロースの純
品を得た。Example 4 Add an equal amount of 0.2 N caustic soda to the mixture of neotrehalose and centose obtained in Example 1 (sugar concentration 10%), autoclave for 15 minutes, neutralize, and then use Toyo Pearl H.
Load it on W-40S, separate it as shown in Figure 3, and analyze it by HPLC.
, a pure product of neotrehalose showing a single peak in PC analysis was obtained.
実施例６
実施例１で得たネオトレハロースとセントースの混合物
をトヨパニルＨＷ−４Ｏ９のカラム（２，６ｃｍ ｘ
１００ｃｍ ｘ ２本）に負荷し、ＨＰＬＣで単一ビ［
発明の効果］
本発明によれば、これまで大量生産が困難であったネオ
トレハロースとセントースが一段階で生産され、しかも
これらの糖は安定性にも優れているので、食品素材とし
て広く利用できるものと期待される。Example 6 The mixture of neotrehalose and centose obtained in Example 1 was applied to a toyopanil HW-4O9 column (2.6 cm x
100cm
Effects of the invention] According to the present invention, neotrehalose and centose, which have been difficult to produce in large quantities, can be produced in one step, and these sugars have excellent stability, so they can be widely used as food materials. It is expected that
また目的に応じて、純品から、これら二成分を主として
含む製品、各種糖との混合物の形でも製品化できる。Furthermore, depending on the purpose, it can be commercialized in the form of a pure product, a product mainly containing these two components, or a mixture with various sugars.
第１図は液化馬鈴薯澱粉にマセランス酵素を作用させた
ときに生成する糖成分を示したＨＰＬＣ図である。
第２図はマセランス酵素反応液にグルコアミラーゼと酵
母を加えて処理した後の糖成分を示したＨＰＬＣ図であ
る。
第３図はグルコアミラーゼと酵母処理液にアルカリを加
え、ネオトレハロース以外の糖を分解した後のカラムク
ロマトグラフィーによる溶出図を示す。
第４図は液化馬鈴薯澱粉に各種濃度のβ−サイクロデキ
ストリン合成酵素を作用させたときの、ネオトレハロー
スとセントースの収率な示す。
第５図は各種糖にβ−サイクロデキストリン合成酵素を
作用させたときの収率を示す。
第６図はグルコアミラーゼと酵母処理液からのカラムク
ロマトグラフィーによるネオトレハロースとセントース
の分離例を示す。
特許出願人 農林水産省食品総合研究所長第１図
０４１２２０ ４゜
保持時間（分）
第２図
０４ １２ ２０ ２Ｂ
保持時間（分）
第３図
溶出ｆｆｉ（ｍｌ）
第４図
ＴＨＵ／ｇＷ質
第５図
Ｇ＋ ０１＋ＣＤ ０２ Ｇ２＋ＣＤ
ＳＳ第６図
溶出ｆｆｉ（ｍｌ）FIG. 1 is an HPLC diagram showing sugar components produced when liquefied potato starch is treated with macerans enzyme. FIG. 2 is an HPLC diagram showing the sugar components after the macerans enzyme reaction solution was treated with glucoamylase and yeast. FIG. 3 shows an elution diagram obtained by column chromatography after adding alkali to the glucoamylase and yeast treatment solution to decompose sugars other than neotrehalose. Figure 4 shows the yields of neotrehalose and centose when liquefied potato starch was treated with various concentrations of β-cyclodextrin synthase. FIG. 5 shows the yield when β-cyclodextrin synthase was allowed to act on various sugars. FIG. 6 shows an example of separation of neotrehalose and centose by glucoamylase and column chromatography from a yeast treatment solution. Patent applicant Director, Food Research Institute, Ministry of Agriculture, Forestry and Fisheries No. 1 041220 4゜Retention time (min) Fig. 2 04 12 20 2B Retention time (min) Fig. 3 Elution ffi (ml) Fig. 4 THU/gW quality No. 5 Figure G+ 01+CD 02 G2+CD
SS Figure 6 Elution ffi (ml)
Claims (1)
クロデキストリン合成酵素を作用させることを特徴とす
るネオトレハロースおよびセントースを含む糖混合物の
製造法。 ２、ネオトレハロースおよびセントースを含む糖混合物
にグルコアミラーゼおよび／または酵母を作用させるこ
とを特徴とする主としてネオトレハロースおよびセント
ースの混合物の製造法。 ３、主としてネオトレハロースおよびセントースの混合
物をアルカリ処理することを特徴とするネオトレハロー
スの製造法 ４、主としてネオトレハロースおよびセントースの混合
物をカラムクロマトグラフィーにかけ両者を分離するこ
とを特徴とするネオトレハロースとセントースの製造法
。[Scope of Claims] 1. A method for producing a sugar mixture containing neotrehalose and centose, which comprises causing a cyclodextrin synthase to act on a substrate selected from starch and its decomposition products. 2. A method for producing a mixture mainly of neotrehalose and centose, which comprises allowing glucoamylase and/or yeast to act on a sugar mixture containing neotrehalose and centose. 3. A method for producing neotrehalose, which is mainly characterized by treating a mixture of neotrehalose and centose with an alkali. 4. A method for producing neotrehalose and centose, which is mainly characterized by subjecting a mixture of neotrehalose and centose to column chromatography to separate them. manufacturing method.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4878587A JPS63216492A (en) | 1987-03-05 | 1987-03-05 | Production of neotrehalose and centose |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4878587A JPS63216492A (en) | 1987-03-05 | 1987-03-05 | Production of neotrehalose and centose |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPS63216492A true JPS63216492A (en) | 1988-09-08 |
| JPH0317477B2 JPH0317477B2 (en) | 1991-03-08 |
Family
ID=12812896
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP4878587A Granted JPS63216492A (en) | 1987-03-05 | 1987-03-05 | Production of neotrehalose and centose |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPS63216492A (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5403727A (en) * | 1990-11-15 | 1995-04-04 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Process for preparing neotrehalose and its uses |
| EP0619951A3 (en) * | 1993-03-16 | 1995-08-02 | Hayashibara Biochem Lab | Energy-supplementing saccharide source and its uses. |
| US5578469A (en) * | 1992-02-25 | 1996-11-26 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Process for preparing neotrehalose, and its uses |
| US5604211A (en) * | 1991-09-20 | 1997-02-18 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Saccharide for supplementing energy to living body, and uses |
| CN106381347A (en) * | 2016-11-22 | 2017-02-08 | 保龄宝生物股份有限公司 | Crystallization technical method for industrially producing trehalose |
-
1987
- 1987-03-05 JP JP4878587A patent/JPS63216492A/en active Granted
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5403727A (en) * | 1990-11-15 | 1995-04-04 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Process for preparing neotrehalose and its uses |
| US5604211A (en) * | 1991-09-20 | 1997-02-18 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Saccharide for supplementing energy to living body, and uses |
| US5578469A (en) * | 1992-02-25 | 1996-11-26 | Kabushiki Kaisha Hayashibara Seibutsu Kagaku Kenkyujo | Process for preparing neotrehalose, and its uses |
| EP0619951A3 (en) * | 1993-03-16 | 1995-08-02 | Hayashibara Biochem Lab | Energy-supplementing saccharide source and its uses. |
| CN106381347A (en) * | 2016-11-22 | 2017-02-08 | 保龄宝生物股份有限公司 | Crystallization technical method for industrially producing trehalose |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH0317477B2 (en) | 1991-03-08 |
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