JPH02222693A - Enzymatic hydrolysis of cellulose-containing material - Google Patents
Enzymatic hydrolysis of cellulose-containing materialInfo
- Publication number
- JPH02222693A JPH02222693A JP4425289A JP4425289A JPH02222693A JP H02222693 A JPH02222693 A JP H02222693A JP 4425289 A JP4425289 A JP 4425289A JP 4425289 A JP4425289 A JP 4425289A JP H02222693 A JPH02222693 A JP H02222693A
- Authority
- JP
- Japan
- Prior art keywords
- cellulose
- containing material
- cellulase
- saturated hydrocarbon
- hexane
- 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
- 229920002678 cellulose Polymers 0.000 title claims abstract description 22
- 239000001913 cellulose Substances 0.000 title claims abstract description 22
- 239000000463 material Substances 0.000 title claims abstract description 17
- 230000007071 enzymatic hydrolysis Effects 0.000 title claims description 5
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 title claims description 5
- 238000000034 method Methods 0.000 claims abstract description 12
- 239000007853 buffer solution Substances 0.000 claims abstract description 11
- 229930195734 saturated hydrocarbon Natural products 0.000 claims abstract description 11
- 239000000243 solution Substances 0.000 claims description 11
- 238000003756 stirring Methods 0.000 claims description 5
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 abstract description 33
- 108010059892 Cellulase Proteins 0.000 abstract description 7
- 229940106157 cellulase Drugs 0.000 abstract description 7
- 230000003301 hydrolyzing effect Effects 0.000 abstract description 5
- 238000005422 blasting Methods 0.000 abstract description 4
- 239000007788 liquid Substances 0.000 abstract description 4
- BHZOKUMUHVTPBX-UHFFFAOYSA-M sodium acetic acid acetate Chemical compound [Na+].CC(O)=O.CC([O-])=O BHZOKUMUHVTPBX-UHFFFAOYSA-M 0.000 abstract description 4
- 241000228245 Aspergillus niger Species 0.000 abstract description 3
- 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 abstract description 3
- 239000008103 glucose Substances 0.000 abstract description 3
- 229920001542 oligosaccharide Polymers 0.000 abstract description 3
- 150000002482 oligosaccharides Chemical class 0.000 abstract description 3
- 239000006185 dispersion Substances 0.000 abstract 1
- 239000011369 resultant mixture Substances 0.000 abstract 1
- 235000000346 sugar Nutrition 0.000 description 19
- 108090000790 Enzymes Proteins 0.000 description 6
- 102000004190 Enzymes Human genes 0.000 description 6
- 229940088598 enzyme Drugs 0.000 description 6
- 108090000604 Hydrolases Proteins 0.000 description 4
- 102000004157 Hydrolases Human genes 0.000 description 4
- IMNFDUFMRHMDMM-UHFFFAOYSA-N N-Heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 4
- 230000000052 comparative effect Effects 0.000 description 4
- 239000000872 buffer Substances 0.000 description 3
- 125000004432 carbon atom Chemical group C* 0.000 description 3
- 230000007062 hydrolysis Effects 0.000 description 3
- 238000006460 hydrolysis reaction Methods 0.000 description 3
- 239000007864 aqueous solution Substances 0.000 description 2
- SNRUBQQJIBEYMU-UHFFFAOYSA-N dodecane Chemical compound CCCCCCCCCCCC SNRUBQQJIBEYMU-UHFFFAOYSA-N 0.000 description 2
- 238000010025 steaming Methods 0.000 description 2
- 150000008163 sugars Chemical class 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 108010084185 Cellulases Proteins 0.000 description 1
- 102000005575 Cellulases Human genes 0.000 description 1
- NHTMVDHEPJAVLT-UHFFFAOYSA-N Isooctane Chemical compound CC(C)CC(C)(C)C NHTMVDHEPJAVLT-UHFFFAOYSA-N 0.000 description 1
- AFCARXCZXQIEQB-UHFFFAOYSA-N N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]-2-[[3-(trifluoromethoxy)phenyl]methylamino]pyrimidine-5-carboxamide Chemical compound O=C(CCNC(=O)C=1C=NC(=NC=1)NCC1=CC(=CC=C1)OC(F)(F)F)N1CC2=C(CC1)NN=N2 AFCARXCZXQIEQB-UHFFFAOYSA-N 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- JVSWJIKNEAIKJW-UHFFFAOYSA-N dimethyl-hexane Natural products CCCCCC(C)C JVSWJIKNEAIKJW-UHFFFAOYSA-N 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- TVMXDCGIABBOFY-UHFFFAOYSA-N octane Chemical compound CCCCCCCC TVMXDCGIABBOFY-UHFFFAOYSA-N 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
Description
【発明の詳細な説明】
Ｕ産業上の利用分野］
本発明は飽和炭化水素の液中にセルロース含有材料を分
散させ、該液中に緩衝液およびセルロース加水分解酵素
を添加してセルロース含有材料を酵素加水分解する方法
に関する６
［従来の技術］
セルロース含有材料をグルコースまたはそのオリゴ糖に
セルロース加水分解酵素を用いて加水分解しようとする
試みは、従来から多数提案されている。しかしセルロー
ス含有材料は難溶性基質であり、これをセルロース加水
分解酵素で加水分解させるためには、爆砕または蒸煮処
理等の複雑な前処理が必要であり、また加水分解速度が
遅く酵素を用いた実用的な加水分解方法はいまだに提案
されていない。Detailed Description of the Invention U Industrial Field of Application] The present invention involves dispersing a cellulose-containing material in a saturated hydrocarbon solution, and adding a buffer and a cellulose hydrolase to the solution to disperse the cellulose-containing material. 6 Related to Enzymatic Hydrolysis Method [Prior Art] Many attempts to hydrolyze cellulose-containing materials into glucose or its oligosaccharides using cellulose hydrolase have been proposed in the past. However, cellulose-containing materials are poorly soluble substrates, and in order to hydrolyze them with cellulose hydrolyzing enzymes, complex pretreatments such as blasting or steaming are required, and the hydrolysis rate is slow, making it difficult to use enzymes. No practical hydrolysis method has yet been proposed.
［発明が解決しようとする課題］
本発明は、爆砕処理等の複雑な前処理を必要とせず、加
水分解速度が速く、かつグルコースまたはそのオリゴ糖
をセルロース含有材料から高収率で得られるセルロース
含有材料の酵素による加水分解方法の提供を目的とする
。[Problems to be Solved by the Invention] The present invention provides cellulose that does not require complicated pretreatment such as blasting treatment, has a high hydrolysis rate, and can obtain glucose or its oligosaccharides in high yield from a cellulose-containing material. The purpose of the present invention is to provide a method for enzymatic hydrolysis of contained materials.
［課題を解決するための手段］
本発明は、セルロース含有材料を分散させた飽和炭化水
素溶液中に、綬ｉ液に溶解させたセルロース加水分解酵
素を添加し撹拌することからなるセルロース含有材料の
酵素加水分解方法を提供する。[Means for Solving the Problems] The present invention provides a method for preparing a cellulose-containing material, which comprises adding a cellulose hydrolase dissolved in a ribbon liquid to a saturated hydrocarbon solution in which the cellulose-containing material is dispersed, and stirring the mixture. An enzymatic hydrolysis method is provided.
本発明で用いる酵素はセルロース加水分解酵素（以下セ
ルラーゼという）であり、Ａｓｐｅｒｇｉｌｌｕｓｎｉ
ｇｅｒ、Ｐｅｎｉｃｉｉｌｕｍ ｒｕｎｉｃｕｌｏｓ
ｕｍ、Ｔｒｉｃｈｏｄｅｒｇ＋ａｖｉｒｉｄｅ等各種産
生菌に由来するセルラーゼを使用することができる。と
くにＡｓｐｅｒｇｉｌｌｕｓ 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。The enzyme used in the present invention is cellulose hydrolase (hereinafter referred to as cellulase),
ger, Peniciilum runiculos
Cellulases derived from various producing bacteria such as um, Trichoderg+aviride, etc. can be used. In particular, cellulase derived from Aspergillus niger is preferably used because it is inexpensive, easily available, and has a high hydrolytic ability.
本発明で用いる飽和炭化水素としては、ヘキサン、ヘプ
タン、オクタン、イソオクタン、ドデカン等各種のもの
を例示できるが、炭素数が多い飽和炭化水素はヘキサン
、ヘプタンに比して還元糖収率が低下するため、ヘキサ
ンを最も好ましいものとして例示できる。Examples of saturated hydrocarbons used in the present invention include hexane, heptane, octane, isooctane, and dodecane, but saturated hydrocarbons with a large number of carbon atoms have a lower reducing sugar yield than hexane and heptane. Therefore, hexane can be cited as the most preferred example.
緩衝液としては、酢酸−酢酸ナトリウム系を好ましい例
として例示できる０Ｍ街衝液飽和炭化水素液中における
割合は、２０容量％以下が好ましく、２０容量％以上で
は還元糖の生成量も２０容量％のそれに比して増加しな
いため好ましくない。As a buffer solution, an acetic acid-sodium acetate system can be exemplified as a preferable example.The ratio in a 0M saturated hydrocarbon solution is preferably 20% by volume or less, and if it is 20% by volume or more, the amount of reducing sugar produced will also be 20% by volume. It is not preferable because it does not increase compared to that.
最も好ましくは１５容量％以下である。Most preferably it is 15% by volume or less.
緩衝液のイオン強度は、１Ｍ付近が好ましいがごく低い
値でもよく、この値に限定されるものではない。The ionic strength of the buffer solution is preferably around 1M, but may be a very low value, and is not limited to this value.
［実施例］
本実施例中における還元糖生成量は、Ｓ ｏｍｏｇｙ−
Ｎｅｌｓｏｎ法、または高速液体クロマトグラフィー（
カラム： ５ｈｏｄｅｘ Ｓｕｇａｒ ＳＰＩ Ｏ１０
、カラム温度＝７０℃）により測定した。[Example] The amount of reducing sugar produced in this example is Somogy-
Nelson method or high performance liquid chromatography (
Column: 5hodex Sugar SPI O10
, column temperature = 70°C).
実施例１、比較例１
イオン強度ＬＭ、ｐＨ５，０の酢酸−酢酸ナトリウムＭ
ｍ液１５ｍ１にＡｓｐｅｒｇｉｌｌｕｓ ｎｉｇｅｒ由
来のセルラーゼ（シグマ社製）を０．４ｇの割合で加え
、撹拌溶解して酵素水溶液を作製した。Example 1, Comparative Example 1 Ionic strength LM, pH 5.0 acetic acid-sodium acetate M
Cellulase derived from Aspergillus niger (manufactured by Sigma) was added at a rate of 0.4 g to 15 ml of the m solution, and dissolved with stirring to prepare an enzyme aqueous solution.
ヘキサン１３５ｍ１にアビセル（結晶性セルロース、シ
グマ社製）５０ｍｇを加えて分散した後、上記酵素水溶
液１５ｍ１を添加し、４０℃、６００ｒｐｍの条件で所
定時間撹拌処理した〈実施例１）＃
ヘキサン１３５ｍ１の代わりに上記の緩衝液１３５ｍ１
を加えた他は上記と同様にして実験した（比較例１）。After adding and dispersing 50 mg of Avicel (crystalline cellulose, manufactured by Sigma) in 135 ml of hexane, 15 ml of the above enzyme aqueous solution was added and stirred for a predetermined time at 40°C and 600 rpm (Example 1) # 135 ml of hexane 135 ml of the above buffer instead
The experiment was carried out in the same manner as above except that the following was added (Comparative Example 1).
各処理時間後の還元糖生成量を第１図に示す。Figure 1 shows the amount of reducing sugar produced after each treatment time.
第１図中、標準化還元糖濃度はＭ薄液中におけるアビセ
ルおよび酵素の濃度を同一水準で比較するため、次のよ
うにして算出した値である。In FIG. 1, the standardized reducing sugar concentration is a value calculated as follows in order to compare the concentrations of Avicel and enzyme in the M thin solution at the same level.
ヘキサン（水分ｉ１０容量％）から分離した緩衝液中の
還元糖濃度を基準とし、緩衝液（水分量１００容量％）
中における還元糖濃度を１０倍した。ヘキサン−Ｍ１！
’ｊ液中における還元糖の生成量は、緩衝液系のそれよ
りも優れている。Based on the reducing sugar concentration in the buffer solution separated from hexane (water content 10% by volume), the buffer solution (water content 100% by volume)
The reducing sugar concentration in the medium was increased 10 times. Hexane-M1!
The amount of reducing sugar produced in the 'j solution is superior to that in the buffer system.
実施例２
実施例１で使用した酢酸−酢酸ナトリウム）！’Ｉ！ｉ
液のヘキサンに対する添加量を変え（各１Ｊｆａｆ液中
のセルラーゼ含有旦は０．４ｇ）、処理温度２０℃、撹
拌処理時間を１００時間とした以外は実施例１と同様に
操作して、緩衝液添加量の相違による還元糖の生成量を
調べた。結果を第２図に示す。Example 2 Acetic acid-sodium acetate used in Example 1)! 'I! i
The buffer solution was prepared in the same manner as in Example 1, except that the amount of the solution added to hexane was changed (cellulase content in each JFAF solution was 0.4 g), the treatment temperature was 20°C, and the stirring treatment time was 100 hours. The amount of reducing sugar produced was investigated depending on the amount added. The results are shown in Figure 2.
緩衝液の添加Ｉが２０容量％以下であるときに優れた還
元糖生成１を示した。なおＭ衝液の添加量はＭｆＲ液と
ヘキサンとの合計に対する緩衝液の割合を示す。Excellent reducing sugar production 1 was shown when the buffer addition I was less than 20% by volume. Note that the amount of the M buffer solution added indicates the ratio of the buffer solution to the total of the MfR solution and hexane.
実施例３
実施例１で使用したヘキサンを炭素数５〜１２の飽和炭
化水素に変え、撹拌処理時間を２２０時間とした以外は
、実施例１と同様に処理して還元糖の生成Ｉを調べた。Example 3 The hexane used in Example 1 was replaced with a saturated hydrocarbon having 5 to 12 carbon atoms and the stirring treatment time was changed to 220 hours, but the same procedure as in Example 1 was conducted to investigate the formation of reducing sugars I. Ta.
結果を第３図に示す、炭素数７の飽和炭化水素による還
元糖の生成量が最も少ないことを示している。The results are shown in FIG. 3 and show that the amount of reducing sugar produced by the saturated hydrocarbon having 7 carbon atoms was the smallest.
［発明の効果］
本発明によれば、爆砕処理、蒸煮処理等の前処理を不要
とするセルロース含有材料の加水分解方法が提供される
。[Effects of the Invention] According to the present invention, a method for hydrolyzing a cellulose-containing material that does not require pretreatment such as blasting treatment or steaming treatment is provided.
さらに本発明によれば、１ｊｉｆＲ液系における還元糖
の生成量に比して極めて優れた還元糖生成量が得られる
セルロース含有材料の加水分解方法が提供される。Further, according to the present invention, there is provided a method for hydrolyzing cellulose-containing materials that yields an extremely superior amount of reducing sugar produced compared to the amount of reducing sugar produced in the 1jifR liquid system.
第１図は、実施例１および比較例１で得られた還元糖の
標準化濃度を示す図面である。第２図は、実施例２で得
られたヘキサン中における！！街液量の相違による還元
糖濃度の相違を示す図面である。
第３図は、実施例３で得られた飽和炭化水素の炭素数を
変えたとき功還元糖濃度の相違を示す図面である。
１：実施例１で得られた還元糖の濃度
２：比較例１で得られた還元糖の濃度FIG. 1 is a drawing showing the standardized concentrations of reducing sugars obtained in Example 1 and Comparative Example 1. Figure 2 shows the ! in hexane obtained in Example 2! ! It is a drawing showing the difference in reducing sugar concentration due to the difference in the amount of street liquid. FIG. 3 is a diagram showing the difference in effective reducing sugar concentration when the carbon number of the saturated hydrocarbon obtained in Example 3 is changed. 1: Concentration of reducing sugar obtained in Example 1 2: Concentration of reducing sugar obtained in Comparative Example 1
Claims (1)
、緩衝液に溶解させたセルロース加水分解酵素を添加し
撹拌することからなるセルロース含有材料の酵素加水分
解方法。A method for enzymatic hydrolysis of a cellulose-containing material, which comprises adding a cellulose hydrolase dissolved in a buffer solution to a saturated hydrocarbon solution in which the cellulose-containing material is dispersed and stirring.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4425289A JP2689161B2 (en) | 1989-02-23 | 1989-02-23 | Method for enzymatic hydrolysis of cellulose-containing material |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP4425289A JP2689161B2 (en) | 1989-02-23 | 1989-02-23 | Method for enzymatic hydrolysis of cellulose-containing material |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH02222693A true JPH02222693A (en) | 1990-09-05 |
| JP2689161B2 JP2689161B2 (en) | 1997-12-10 |
Family
ID=12686339
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP4425289A Expired - Lifetime JP2689161B2 (en) | 1989-02-23 | 1989-02-23 | Method for enzymatic hydrolysis of cellulose-containing material |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP2689161B2 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2011205933A (en) * | 2010-03-29 | 2011-10-20 | Aichi Prefecture | Method for producing high-concentration saccharified liquid |
-
1989
- 1989-02-23 JP JP4425289A patent/JP2689161B2/en not_active Expired - Lifetime
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2011205933A (en) * | 2010-03-29 | 2011-10-20 | Aichi Prefecture | Method for producing high-concentration saccharified liquid |
Also Published As
| Publication number | Publication date |
|---|---|
| JP2689161B2 (en) | 1997-12-10 |
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