JPH02200196A - Production of low molecular weight chitosan - Google Patents
Production of low molecular weight chitosanInfo
- Publication number
- JPH02200196A JPH02200196A JP1945489A JP1945489A JPH02200196A JP H02200196 A JPH02200196 A JP H02200196A JP 1945489 A JP1945489 A JP 1945489A JP 1945489 A JP1945489 A JP 1945489A JP H02200196 A JPH02200196 A JP H02200196A
- Authority
- JP
- Japan
- Prior art keywords
- chitosan
- molecular weight
- low molecular
- reaction solution
- lipase
- 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
- 229920001661 Chitosan Polymers 0.000 title claims abstract description 72
- 238000004519 manufacturing process Methods 0.000 title claims description 5
- 102000035092 Neutral proteases Human genes 0.000 claims abstract description 10
- 108091005507 Neutral proteases Proteins 0.000 claims abstract description 10
- 108090001060 Lipase Proteins 0.000 claims abstract description 9
- 102000004882 Lipase Human genes 0.000 claims abstract description 9
- 239000004367 Lipase Substances 0.000 claims abstract description 9
- 235000019421 lipase Nutrition 0.000 claims abstract description 9
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 abstract description 12
- 108090000145 Bacillolysin Proteins 0.000 abstract description 8
- 101000968491 Pseudomonas sp. (strain 109) Triacylglycerol lipase Proteins 0.000 abstract description 5
- 229920002101 Chitin Polymers 0.000 abstract description 3
- 239000000126 substance Substances 0.000 abstract description 3
- 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 abstract description 2
- 239000003456 ion exchange resin Substances 0.000 abstract description 2
- 229920003303 ion-exchange polymer Polymers 0.000 abstract description 2
- 241000238565 lobster Species 0.000 abstract 1
- 238000001556 precipitation Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 43
- 238000006243 chemical reaction Methods 0.000 description 35
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 25
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 24
- 238000000034 method Methods 0.000 description 19
- 238000003756 stirring Methods 0.000 description 19
- 239000002244 precipitate Substances 0.000 description 7
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- 101710180012 Protease 7 Proteins 0.000 description 5
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 101710098556 Lipase A Proteins 0.000 description 3
- 101710099648 Lysosomal acid lipase/cholesteryl ester hydrolase Proteins 0.000 description 3
- 102100026001 Lysosomal acid lipase/cholesteryl ester hydrolase Human genes 0.000 description 3
- 238000000354 decomposition reaction Methods 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 235000011121 sodium hydroxide Nutrition 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- KZBUYRJDOAKODT-UHFFFAOYSA-N Chlorine Chemical compound ClCl KZBUYRJDOAKODT-UHFFFAOYSA-N 0.000 description 2
- 235000011054 acetic acid Nutrition 0.000 description 2
- 108010089807 chitosanase Proteins 0.000 description 2
- XBDQKXXYIPTUBI-UHFFFAOYSA-N dimethylselenoniopropionate Natural products CCC(O)=O XBDQKXXYIPTUBI-UHFFFAOYSA-N 0.000 description 2
- 238000000909 electrodialysis Methods 0.000 description 2
- 238000000855 fermentation Methods 0.000 description 2
- 230000004151 fermentation Effects 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000007935 neutral effect Effects 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000007921 spray Substances 0.000 description 2
- 235000020138 yakult Nutrition 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- 241000238557 Decapoda Species 0.000 description 1
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 101710177166 Phosphoprotein Proteins 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 239000004373 Pullulan Substances 0.000 description 1
- 229920001218 Pullulan Polymers 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- 230000000397 acetylating effect Effects 0.000 description 1
- 150000008044 alkali metal hydroxides Chemical class 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000009833 condensation Methods 0.000 description 1
- 230000005494 condensation Effects 0.000 description 1
- 239000002537 cosmetic Substances 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 230000000850 deacetylating effect Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000000593 degrading effect Effects 0.000 description 1
- 238000011033 desalting Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000003925 fat Substances 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 150000007522 mineralic acids Chemical class 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229910017604 nitric acid Inorganic materials 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 235000019260 propionic acid Nutrition 0.000 description 1
- 235000019423 pullulan Nutrition 0.000 description 1
- IUVKMZGDUIUOCP-BTNSXGMBSA-N quinbolone Chemical compound O([C@H]1CC[C@H]2[C@H]3[C@@H]([C@]4(C=CC(=O)C=C4CC3)C)CC[C@@]21C)C1=CCCC1 IUVKMZGDUIUOCP-BTNSXGMBSA-N 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 238000006722 reduction reaction Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 239000013585 weight reducing agent Substances 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、自然界のキチンを脱アセチル化して得られる
高分子量のキトサンから、任意の分子量を有する低分子
量キトサンを、簡単な操作で安全に製造する方法に関す
る。[Detailed Description of the Invention] [Industrial Application Field] The present invention is a method for safely producing low-molecular-weight chitosan with a desired molecular weight from high-molecular-weight chitosan obtained by deacetylating chitin in nature by a simple operation. Relating to a method of manufacturing.
近年、化粧品、医薬品、抗菌剤など、各種の分野で低分
子量のキトサンの需要が高まっている。In recent years, demand for low-molecular-weight chitosan has been increasing in various fields such as cosmetics, pharmaceuticals, and antibacterial agents.
従来、低分子量キトサンを製造する方法としては、微生
物由来のキトサナーゼを用いる方法、高濃度の塩酸水溶
液中で加熱処理する方法、塩素ガスを用いる方法、過酸
化水素を用いる方法などが知られている。Conventionally, known methods for producing low-molecular-weight chitosan include a method using microbial-derived chitosanase, a method using heat treatment in a highly concentrated aqueous hydrochloric acid solution, a method using chlorine gas, and a method using hydrogen peroxide. .
これらの方法はキトサナーゼの安全性の未確認、塩酸法
の後処理の煩雑さ、塩素ガスの有毒性、残留過酸化水素
の問題など、安全性及び経済性に難点があった０本発明
者らは種々検討の結果、安全にしかも安価に、任意の分
子量を有する低分子量キトサンを得る方法を見出し、本
発明を完成するに至った。These methods had drawbacks in terms of safety and economy, such as the unconfirmed safety of chitosanase, the complexity of post-treatment with the hydrochloric acid method, the toxicity of chlorine gas, and the problem of residual hydrogen peroxide. As a result of various studies, we have found a method for safely and inexpensively obtaining low molecular weight chitosan having a desired molecular weight, and have completed the present invention.
本発明はキトサンをキトサン分解能を有する中性プロテ
アーゼ及びリパーゼの１種以上で処理することによって
低分子化することを特徴とする低分子量キトサンの製造
方法に関する。The present invention relates to a method for producing low-molecular-weight chitosan, which is characterized in that the chitosan is reduced in molecular weight by treating chitosan with one or more types of neutral protease and lipase that have the ability to decompose chitosan.
中性プロテアーゼは食品加工、醸造・発酵、医薬品、飼
料等で使用されている安全性の十分ｖａ認されている市
販品が利用できる。キトサン分解能を有する中性プロテ
アーゼの例として、プロチンＰ（大和化成株式会社製造
）パンチダーゼＮＰ−２（株式会社ヤクルト本社製造）
、プロテアーゼＡ「アマノ」 （天野製薬株式会社製造
）等が上げられる。As the neutral protease, commercially available products that are used in food processing, brewing/fermentation, pharmaceuticals, feed, etc. and have been fully approved for safety can be used. Examples of neutral proteases capable of degrading chitosan include Protin P (manufactured by Daiwa Kasei Co., Ltd.) and Punchdase NP-2 (manufactured by Yakult Honsha Co., Ltd.).
and protease A "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.).
リパーゼは油脂の分解・改質、乳製品の改質、醸造・醗
酵、医薬品等で使用されている安全性の十分ｎ！認され
ている市販品が利用できる。キトサン分解能を有するリ
パーゼの例として、リパーゼＰ「アマノ」リパーゼＡ「
アマノ」いずれも（天野製薬株式会社製造）等が上げら
れる。Lipase is used in the decomposition and modification of fats and oils, the modification of dairy products, brewing and fermentation, pharmaceuticals, etc., and is sufficiently safe! Approved commercially available products are available. Examples of lipases having the ability to decompose chitosan include Lipase P "Amano" and Lipase A "
Amano'' (manufactured by Amano Pharmaceutical Co., Ltd.), etc.
本発明で使用するキトサンは、エビ、カニ等から得られ
る天然のキチンを、例えば苛性ソーダ等で説アセチル化
したものが好ましい１通常分子量１００万程度以上のも
のを使用するが、低分子化したいキトサンであればこれ
以下の分子量のものでも使用しうる。The chitosan used in the present invention is preferably obtained by acetylating natural chitin obtained from shrimp, crab, etc. with caustic soda, etc. 1 Usually, chitosan with a molecular weight of about 1 million or more is used, but chitosan desired to have a lower molecular weight is used. If so, those having a molecular weight lower than this can also be used.
キトサンは塩酸又は硝酸等の無機酸及びギ酸、酢酸、プ
ロピオン酸、乳酸等の有機酸を用いて水溶液とすること
ができる。Chitosan can be made into an aqueous solution using an inorganic acid such as hydrochloric acid or nitric acid and an organic acid such as formic acid, acetic acid, propionic acid, or lactic acid.
キトサン溶液に対して中性プロテアーゼ又はリパーゼの
１種以上を作用せさることにより低分子量キトサンが得
られる。Low molecular weight chitosan can be obtained by allowing one or more types of neutral protease or lipase to act on a chitosan solution.
低分子化反応の条件は本発明では特に限定されないが、
キトサン溶液の濃度は１〜２０％、好ましくは３〜１０
％である。キトサン溶液のＰＨは３〜６が好ましい。中
性プロテアーゼ及びリパーゼの使用量は任意の分子量の
低分子量キトサンを得るため、特に限定されないが、通
常キトサンに対し０．００１ ＝１０ｗ／ｗ％程度使用
される。又、反応温度は常温〜約５５℃、好ましくは４
０〜５０℃、反応時間は１−１００時間程度がよい。The conditions for the molecular weight reduction reaction are not particularly limited in the present invention, but
The concentration of chitosan solution is 1-20%, preferably 3-10%
%. The pH of the chitosan solution is preferably 3-6. The amount of neutral protease and lipase to be used is not particularly limited in order to obtain low molecular weight chitosan of arbitrary molecular weight, but is usually used in an amount of about 0.001 = 10 w/w% based on chitosan. Further, the reaction temperature is room temperature to about 55°C, preferably 4°C.
The reaction time is preferably about 1 to 100 hours at 0 to 50°C.
分解反応後の低分子量キトサン溶液は、先ず不純物をろ
過し、次いでアルカリ金属の水酸化物溶液でＰＨを約８
に調整するか、又はエタノール、アセトン等の溶媒を用
いて沈殿を生成させ、沈殿物をろ過することにより低分
子量キトサンを得ることができる。あるいは分解反応後
の低分子量キトサン溶液を濾過し、ＰＨを約８に調整し
た後、生じた沈殿を除去し、次いで生成した塩をイオン
交換樹脂、電気透析、透析膜あるいは他の膜を使う方法
、等により脱塩した後、ａ縮乾固、凍結乾燥又はスプレ
ードライヤー等で乾燥することにより水溶性の低分子量
キトサンを得ることができる。After the decomposition reaction, the low molecular weight chitosan solution is first filtered to remove impurities, and then the pH is adjusted to about 8 with an alkali metal hydroxide solution.
Low-molecular-weight chitosan can be obtained by adjusting the amount of chitosan, or by generating a precipitate using a solvent such as ethanol or acetone, and filtering the precipitate. Alternatively, the low molecular weight chitosan solution after the decomposition reaction is filtered, the pH is adjusted to about 8, the resulting precipitate is removed, and the resulting salt is removed using an ion exchange resin, electrodialysis, dialysis membrane, or other membrane. , etc., and then drying by condensation to dryness, freeze drying, spray dryer, etc., water-soluble low molecular weight chitosan can be obtained.
低分子量キトサンの分子量は目的とする性質により種々
異なるが、通常１０万以下、水溶性を付与する場合は数
百−数千程度である。しかし目的によっては原料キトサ
ンの分子量以下であればｌＯ万以上の場合もある。The molecular weight of low molecular weight chitosan varies depending on the desired properties, but is usually 100,000 or less, and in the case of imparting water solubility, it is about several hundred to several thousand. However, depending on the purpose, the molecular weight may be 10,000 or more if it is less than the molecular weight of the raw material chitosan.
以下に実施例で詳細に説明する。This will be explained in detail in Examples below.
〔実施例１〕
キトサン１０ｇを５００−の水に分散し濃塩酸４６２ｇ
を加え、攪拌熔解した後、市販の中性プロテアーゼのパ
ンチダーゼＰＮ−２（株式会社ヤクルト本社製造） ０
．５ｇを１Ｏ１１の水に熔解し、反応液に添加した０反
応液の温度を４５″Ｃに保持し、１２時間攪拌を続けた
。この時の反応液のｐＨは５．０であった。[Example 1] Disperse 10 g of chitosan in 500-g of water and add 462 g of concentrated hydrochloric acid.
After stirring and dissolving, add commercially available neutral protease Panchidase PN-2 (manufactured by Yakult Honsha Co., Ltd.) 0
．． 5 g was dissolved in 1O11 water and added to the reaction solution.The temperature of the 0 reaction solution was maintained at 45''C and stirring was continued for 12 hours.The pH of the reaction solution at this time was 5.0.
キトサンの分子量はプルランを標準物質として東ソー株
式会社製のＴＳＫｇｅｌ ＧＭＰＷｘＬカラムを用いて
ＧＰＣ法により測定した０反応前の分子量は１３２万で
あったが、反応後のキトサンの分子量は７０．０００ま
で低下した。The molecular weight of chitosan was measured by GPC method using a TSKgel GMPWxL column manufactured by Tosoh Corporation using pullulan as a standard substance.The molecular weight before reaction was 1.32 million, but after reaction the molecular weight of chitosan decreased to 70,000. did.
反応液は９０°Ｃで３０分の加熱処理を行い、−旦、ろ
過して後、５％のＮａＯＨを用いてｐＨを８に調整し沈
殿物を得た。これをろ過、水洗、乾燥させて低分子量キ
トサン９．５ｇを得た。The reaction solution was heated at 90° C. for 30 minutes, filtered, and adjusted to pH 8 using 5% NaOH to obtain a precipitate. This was filtered, washed with water, and dried to obtain 9.5 g of low molecular weight chitosan.
〔実施例２〕
キトサン１０ｇを５００ ｄの水に分散し、濃塩酸４．
２８を加え、攪拌溶解した後、市販のプロチンＰＣＩＯ
（大和化成株式株式会社製造） ０．５ｇを１０ｍ１の
水に溶解し、反応液に添加した０反応液の温度を５０°
Ｃに保持し、１２時間攪拌を続けた。[Example 2] 10 g of chitosan was dispersed in 500 d of water, and 4.0 g of concentrated hydrochloric acid was added.
After adding 28 and stirring to dissolve, commercially available protin PCIO
(Manufactured by Daiwa Kasei Co., Ltd.) 0.5g was dissolved in 10ml of water and added to the reaction solution.The temperature of the reaction solution was adjusted to 50°C.
C. and continued stirring for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は４６，０００であった。The molecular weight of the reaction solution was measured using the same GPC method as in Example 1, and the result was 46,000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例３〕
キトサン１０ｇを５００ ｍの水に分散し、濃塩酸４．
２ｇを加え、攪拌溶解した後、プロテアーゼＡ「アマノ
」 （天野製薬株式会社製造） ０．５ｇを１０ｉｄの
水に溶解し、反応液に添加した０反応液の温度を４５°
Ｃに保持し、１２時間攪拌を続けた。[Example 3] 10 g of chitosan was dispersed in 500 m of water, and 4.0 g of concentrated hydrochloric acid was added.
After adding 2 g and stirring to dissolve, 0.5 g of protease A "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.) was dissolved in 10 id of water, and the temperature of the 0 reaction solution added to the reaction solution was adjusted to 45°.
C. and continued stirring for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果３０．０００であった。The molecular weight of the reaction solution was measured by the same GPC method as in Example 1, and the result was 30.000.
この後、実施例Ｉと同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example I to obtain low molecular weight chitosan.
〔実施例４〕
キトサンｌｏｇを５００ ａｌｌ！の水に分散し、試薬
特級の酢酸３，２ｇを加え、撹拌溶解した後にプロテア
ーゼＡ「アマノＪ 、０．５ｇを１０ｄの水に溶解し、
反応液に添加した０反応液の温度を４５°Ｃに保持し１
２時間攪拌を続けた。[Example 4] Chitosan log 500 all! of water, add 3.2 g of reagent grade acetic acid, stir and dissolve, then dissolve 0.5 g of protease A "Amano J" in 10 d of water.
0 The temperature of the reaction solution added to the reaction solution was maintained at 45 °C, and 1
Stirring was continued for 2 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は２５．０００であった。The molecular weight of the reaction solution was measured using the same GPC method as in Example 1, and the result was 25,000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例５〕
キトサン１０ｇを５００ｄの水に分散し濃塩酸４．２ｇ
を加え、攪拌熔解した後、プロテアーゼＡ「アマノ、
０．５ｇを１０ａｅの水に溶解し、反応液に添加した。[Example 5] Disperse 10 g of chitosan in 500 d of water and add 4.2 g of concentrated hydrochloric acid.
was added, stirred and dissolved, and protease A "Amano,"
0.5 g was dissolved in 10 ae of water and added to the reaction solution.
反応液の温度を４５℃に保持し、４８時間攪拌を続けた
。The temperature of the reaction solution was maintained at 45° C., and stirring was continued for 48 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は１ 、５００であった。The molecular weight of the reaction solution was measured by the same GPC method as in Example 1, and the result was 1,500.
反応液は９０℃で３０分の加熱処理を行い、５％のＮａ
ＯＨを用いてＪ）Ｈを８に調整し、生じた沈殿を除去し
た後、無水エタノール１５００ｄを加え沈殿物を得た。The reaction solution was heated at 90°C for 30 minutes, and 5% Na
After adjusting J)H to 8 using OH and removing the resulting precipitate, 1500 d of absolute ethanol was added to obtain a precipitate.
これをろ過、乾燥して低分子量キトサン６ｇを得た。This was filtered and dried to obtain 6 g of low molecular weight chitosan.
得られた低分子量キトサンは中性の水に溶解した。The obtained low molecular weight chitosan was dissolved in neutral water.
〔実施例６〕 実施例５と同一の方法で低分子量キトサン溶液を得た。[Example 6] A low molecular weight chitosan solution was obtained in the same manner as in Example 5.
この溶液を９０゛Ｃで３０分の加熱処理を行い、５％の
ＮａＯＨを用いてＰＨを８に調整し、生じた沈殿を除去
した後、電気透析で脱塩を行った。その溶液中のキトサ
ン濃度を２倍になるように濃縮した後、スプレードライ
ヤーにて乾燥し微粉末の低分子量キトサン５ｇを得た。This solution was heated at 90° C. for 30 minutes, the pH was adjusted to 8 using 5% NaOH, and the resulting precipitate was removed, followed by desalting by electrodialysis. After concentrating the solution so that the chitosan concentration in the solution was doubled, it was dried with a spray dryer to obtain 5 g of low molecular weight chitosan in the form of a fine powder.
得られた低分子量キトサンは中性の水に溶解した。The obtained low molecular weight chitosan was dissolved in neutral water.
〔実施例７〕
キトサンｌＯｇを５００ｄの水に分散し濃塩酸４．２ｇ
を加え、攪拌熔解した後、市販のリパーゼＰ「アマノ」
（天野製薬株式会社製造） ０．５ｇを１０−の水に
溶解し、反応液に添加した０反応液の濃度を５０℃に保
持し、１２時間攪拌を続けた。[Example 7] Disperse 10g of chitosan in 500d of water and add 4.2g of concentrated hydrochloric acid.
After adding and stirring to dissolve, commercially available lipase P "Amano"
(Manufactured by Amano Pharmaceutical Co., Ltd.) 0.5 g was dissolved in 10-water, and the concentration of the 0-reaction solution added to the reaction solution was maintained at 50° C., and stirring was continued for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は６２．０００であった。The molecular weight of the reaction solution was measured by the same GPC method as in Example 1, and the result was 62.000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例８〕
キトサン１０ｇを５００ ｄの水に分散し濃塩酸４．２
ｇを加え、攪拌溶解した後、市販のリパーゼＡ「アマノ
」 （天野製薬株式会社製ａ ） ０．５ｇを１０ｄの
水に溶解し、反応液に添加した。反応液の温度を４５°
Ｃに保持し、１２時間攪拌を続けた。[Example 8] Disperse 10 g of chitosan in 500 d of water and add 4.2 g of concentrated hydrochloric acid.
After stirring and dissolving, 0.5 g of commercially available lipase A "Amano" (manufactured by Amano Pharmaceutical Co., Ltd.) was dissolved in 10 d of water and added to the reaction solution. The temperature of the reaction solution was set to 45°.
C. and continued stirring for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は６４，０００であった。The molecular weight of the reaction solution was measured using the same GPC method as in Example 1, and the result was 64,000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例９〕
キトサン１０ｇを５００ ｍの水に溶解し、濃塩酸４゜
２ｇを加え、攪拌溶解した後、プロテアーゼＡｒアマノ
」０．２５ｇとリパーゼＡ「アマノＪ 、０．２５ｇを
１ＯＩｄの水に溶解し、反応液に添加した。反応液の温
度を４５°Ｃに保持し、１２時間撹拌を続けた。[Example 9] Dissolve 10 g of chitosan in 500 m of water, add 4.2 g of concentrated hydrochloric acid, stir and dissolve, and then add 0.25 g of protease Ar Amano and 0.25 g of lipase A Amano J to 1 OId of water. The mixture was dissolved in water and added to the reaction solution.The temperature of the reaction solution was maintained at 45°C and stirring was continued for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は３８，０００であった。The molecular weight of the reaction solution was measured using the same GPC method as in Example 1, and the result was 38,000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例１０〕
キトサン１０ｇを５００ｄの水に分散し、濃塩酸４゜２
１を加え、攪拌溶解した後、プロチンＰ ＣＩＯ，０゜
２５ｇとリパーゼＰ「アマノＪ Ｓｏ、２５ｇを１０Ｒ
１の水に溶解し、反応液に添加した６反応液の温度を４
５°Ｃに保持し１２時間撹拌を続けた。[Example 10] 10 g of chitosan was dispersed in 500 d of water, and 4.2 g of concentrated hydrochloric acid was added.
Add 1 and stir to dissolve, then add 25 g of Protein P CIO, 0° and 25 g of Lipase P “Amano J So” to 10R.
The temperature of the reaction solution 6 dissolved in water and added to the reaction solution 4
The temperature was maintained at 5°C and stirring continued for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は４６，０００であった。The molecular weight of the reaction solution was measured using the same GPC method as in Example 1, and the result was 46,000.
この後、実施例１と同様に処理して低分子量キトサンを
得た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔実施例１１）
キトサンＩＯｇを５００ ｄの水に分散し、濃塩酸４゜
２ｇを加え、攪拌熔解した後、リパーゼＰ「アマノ」０
．１ｇ、プロテアーゼＡ「アマノＪ 、０．２ｇ、プロ
チンｐｃｔｏ、０．２ｇを１０ｍの水に溶解し、反応液
に添加した。[Example 11] Chitosan IOg was dispersed in 500 d of water, 4.2 g of concentrated hydrochloric acid was added, and after stirring and melting, Lipase P "Amano" 0
．． 1 g of protease A, 0.2 g of Amano J, and 0.2 g of protin pcto were dissolved in 10 m of water and added to the reaction solution.
反応液の温度を５０″Ｃに保持し、１２時間攪拌を続け
た。The temperature of the reaction solution was maintained at 50''C and stirring was continued for 12 hours.
反応液について実施例１と同一のＧＰＣ法による分子量
の測定の結果は３５．０００であった。The molecular weight of the reaction solution was measured by the same GPC method as in Example 1, and the result was 35,000.
この後、実施例１同様に処理して低分子量キトサンを得
た。Thereafter, it was treated in the same manner as in Example 1 to obtain low molecular weight chitosan.
〔効果）
実施例の結果からも明らかなように、本発明の低分子量
キトサンの製造法は、キトサンを溶解し中性プロテアー
ゼ及びリパーゼの１種以上で処理することにより、効率
良くまた安全な、低分子量キトサンが得られる。[Effects] As is clear from the results of the examples, the method for producing low molecular weight chitosan of the present invention can efficiently and safely produce chitosan by dissolving chitosan and treating it with one or more of neutral protease and lipase. Low molecular weight chitosan is obtained.
特許出願人 日本化薬株式会社Patent applicant: Nippon Kayaku Co., Ltd.
Claims (1)
はリパーゼの１種以上で処理することによって低分子化
することを特徴とする、低分子量キトサンの製造方法。A method for producing low molecular weight chitosan, which comprises reducing the molecular weight of chitosan by treating it with one or more types of neutral proteases or lipases having the ability to decompose chitosan.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP1019454A JP2732107B2 (en) | 1989-01-31 | 1989-01-31 | Method for producing low molecular weight chitosan |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP1019454A JP2732107B2 (en) | 1989-01-31 | 1989-01-31 | Method for producing low molecular weight chitosan |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH02200196A true JPH02200196A (en) | 1990-08-08 |
| JP2732107B2 JP2732107B2 (en) | 1998-03-25 |
Family
ID=11999768
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP1019454A Expired - Fee Related JP2732107B2 (en) | 1989-01-31 | 1989-01-31 | Method for producing low molecular weight chitosan |
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| Country | Link |
|---|---|
| JP (1) | JP2732107B2 (en) |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100370929B1 (en) * | 2000-03-28 | 2003-02-05 | 조훈형 | Preparing Methode for Aqueous Chitosan |
| US6693188B2 (en) | 2001-08-08 | 2004-02-17 | Cargill Incorporated | N-acetyl-D-glucosamine and process for producing N-acetyl-D-glucosamine |
| KR100441270B1 (en) * | 2001-09-25 | 2004-07-22 | 나재운 | The Method for Preparation of Water Soluble Free Amine Chitosan |
| KR100506710B1 (en) * | 2003-03-24 | 2005-08-05 | 주식회사 건풍바이오 | Chitosan oligosaccharide ascorbic acid salt having anti-diabetic effect |
| US6972284B2 (en) | 2000-03-15 | 2005-12-06 | Cargill, Incorporated | Chitosan and method of preparing chitosan |
| CN1320123C (en) * | 2004-05-08 | 2007-06-06 | 合肥学院 | Production technology of low molecular weight chitin |
| US7488812B2 (en) | 2002-04-02 | 2009-02-10 | Cargill, Incorporated | Chitosan production |
| US7816514B2 (en) | 2001-02-16 | 2010-10-19 | Cargill, Incorporated | Glucosamine and method of making glucosamine from microbial biomass |
| US7923437B2 (en) | 2001-02-16 | 2011-04-12 | Cargill, Incorporated | Water soluble β-glucan, glucosamine, and N-acetylglucosamine compositions and methods for making the same |
| US8222232B2 (en) | 2001-02-16 | 2012-07-17 | Cargill, Incorporated | Glucosamine and N-acetylglucosamine compositions and methods of making the same fungal biomass |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102673886B1 (en) * | 2023-10-06 | 2024-06-10 | 농업회사법인(주)건강애 | Manufacturing method of germinated brown rice containing chitosan |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS6363388A (en) * | 1986-09-04 | 1988-03-19 | Nitta Zerachin Kk | Production of low molecular weight chitosan |
-
1989
- 1989-01-31 JP JP1019454A patent/JP2732107B2/en not_active Expired - Fee Related
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS6363388A (en) * | 1986-09-04 | 1988-03-19 | Nitta Zerachin Kk | Production of low molecular weight chitosan |
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6972284B2 (en) | 2000-03-15 | 2005-12-06 | Cargill, Incorporated | Chitosan and method of preparing chitosan |
| US7413881B2 (en) | 2000-03-15 | 2008-08-19 | Cargill, Incorporated | Chitosan and method of preparing chitosan |
| KR100370929B1 (en) * | 2000-03-28 | 2003-02-05 | 조훈형 | Preparing Methode for Aqueous Chitosan |
| US7816514B2 (en) | 2001-02-16 | 2010-10-19 | Cargill, Incorporated | Glucosamine and method of making glucosamine from microbial biomass |
| US7923437B2 (en) | 2001-02-16 | 2011-04-12 | Cargill, Incorporated | Water soluble β-glucan, glucosamine, and N-acetylglucosamine compositions and methods for making the same |
| US8034925B2 (en) | 2001-02-16 | 2011-10-11 | Cargill, Incorporated | Glucosamine and method of making glucosamine from microbial biomass |
| US8222232B2 (en) | 2001-02-16 | 2012-07-17 | Cargill, Incorporated | Glucosamine and N-acetylglucosamine compositions and methods of making the same fungal biomass |
| US6693188B2 (en) | 2001-08-08 | 2004-02-17 | Cargill Incorporated | N-acetyl-D-glucosamine and process for producing N-acetyl-D-glucosamine |
| KR100441270B1 (en) * | 2001-09-25 | 2004-07-22 | 나재운 | The Method for Preparation of Water Soluble Free Amine Chitosan |
| US7488812B2 (en) | 2002-04-02 | 2009-02-10 | Cargill, Incorporated | Chitosan production |
| KR100506710B1 (en) * | 2003-03-24 | 2005-08-05 | 주식회사 건풍바이오 | Chitosan oligosaccharide ascorbic acid salt having anti-diabetic effect |
| CN1320123C (en) * | 2004-05-08 | 2007-06-06 | 合肥学院 | Production technology of low molecular weight chitin |
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| Publication number | Publication date |
|---|---|
| JP2732107B2 (en) | 1998-03-25 |
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