JP3924612B2 - Complex of silk protein and chitosan and method for producing the same - Google Patents

Description

【００４８】
（表2）

【００４９】
(6)橋かけ構造の導入の確認
フーリエ変換赤外分光光度計((株)日本分光、FT-IR 430)を用いて、上記橋かけキトサンNo.1〜3と原料キトサンFM80のIR差スペクトルを測定し、反応を確認した。また、5vol%ギ酸および5vol%酢酸に橋かけキトサンNo.1〜3を加え、その溶解性の変化から橋かけの存在を確認した。
【００５０】
(7)塩縮絹タンパク質とキトサンとの複合体に対する酸性染料の吸着実験
未処理絹糸、塩縮処理絹糸および塩縮処理した絹糸(絹タンパク質)とキトサンとの複合体をそれぞれ、0.01g精秤し、pH4の0.1mol・dm^-3の酢酸緩衝溶液に酸性染料(OR）を溶解した溶液200ml中に入れ、40℃で、72hrs撹拌した。吸着後の染料溶液を紫外・可視分光光度計((株)日立製作所製1100型)を用い、λ=485nmにおいて比色して遊離染料濃度を求めた。また、染色絹糸から染料をホルムアミドで3回、60℃、30minにて完全に抽出し、50mlに定容した溶液を同様に比色して、絹糸による吸着染料量を算出した。
【００５１】
(8)塩縮率
野蚕絹糸を加熱した濃厚な中性塩水溶液中に浸漬すると、野蚕絹糸は繊維長方向に任意に収縮する。次式を用いて塩縮率(%)を求めた。
塩縮率(%) = [(Li-Lf)/Li]×100
上式において、Liは収縮前の試料長を、また、Lfは収縮後の試料長を示す。
(9)機械的特性
機械的特性として、試料の降伏点の伸度(%)、切断時の試料の強度と伸度(%)を測定した。測定条件は、試料長50mm、引張り速度10mm/min、チャートスケール250gであり、島津製インストロン(オートグラフAGS-5D)を使用して測定した。
【００５２】
(10)アルカリ溶解度(Asol)
65℃の0.1M水酸化ナトリウム水溶液に試料を1時間浸漬処理し、処理前後の重量変化からアルカリ溶解度を評価した。
Asol = (Mb-Ma)/Mb×100
Mb：アルカリ処理前のサンプル重量(105℃で1時間30分乾燥)
Ma：アルカリ処理後のサンプル重量(105℃で1時間30分乾燥)
【００５３】
(11)金属イオンの吸着および定量方法
被測定試料を硝酸カリウムを含む0.5mMの硝酸銀水溶液あるいは硝酸銅水溶液(アンモニア水を加えてpHを11.4に調整)に室温で30時間浸漬することで金属イオンを吸着させた。
被測定試料に吸着した金属イオンを、パーキンエルマー社製のプラズマ原子吸光スペクトロメーター(ICP-AES)を用いて分析した。5〜10mgの試料をミクロウェーブ加水分解炉(MDS-81DCCEM)を用いて2mLの65%硝酸で完全に加水分解し、実験時にさらに10mLの水を加え、ICP-AES分析を行った。金属イオンの吸着量は、試料重量あたりの金属イオン量をmmolで表示した。
【００５４】
上記実験で明らかとなった実施例を下記に示す。
(実施例1)絹糸の塩縮処理
60℃の50wt%硝酸カルシウム四水和物水溶液に家蚕絹糸(浴比：硝酸カルシウム水溶液の体積／絹糸の重量＝100とした)を5min浸漬することで絹糸を長さ方向に収縮させ、更に塩縮後、蒸留水にて3回洗浄し、風乾・真空乾燥して塩縮絹糸を得た。この塩縮率は、14%であった。
なお、塩縮絹糸をキトサンの酢酸水溶液に浸漬する際は、上記のようにして得られた塩縮絹糸を蒸留水で洗浄後、乾燥せずに蒸留水に浸漬したものを塩縮試料として用いた。
【００５５】
(実施例2)キトサン水溶液中での浸漬処理
実施例１にしたがって家蚕絹糸を塩縮処理後、蒸留水に浸漬し、膨潤しておいた塩縮絹糸をセパラブルフラスコに入れ、これに、5vol%酢酸水溶液500mlにキトサン10gを溶解した溶液を加え、30℃、24hrs浸漬処理することでキトサン含有絹糸を製造した。
【００５６】
(実施例3)キトサン含有絹糸への架橋結合の導入
実施例2で製造したキトサン含有絹糸への架橋結合導入は次のように行った。0.7gのキトサン含有絹糸を水250mlに入れ、EGDGE 0.44gを加え、70℃で3hrs静かに撹拌することで架橋反応を進めた。反応終了後、ろ過し、ジエチルエーテル、アセトンで数回洗浄後、風乾、真空乾燥した。このようにして架橋結合を導入したキトサン含有絹糸は、酢酸に浸漬して長時間静置しても、重量減少が起こらず、キトサンが絹糸から離脱しなかった。
【００５７】
(実施例4)絹糸へのキトサン導入量の定量
実施例1で得られた塩縮絹糸(塩縮率14%)をキトサンの酢酸水溶液に浸漬する処理で、量的にどの程度のキトサンが絹糸に導入されるかをニンヒドリン試験を行って評価した。すなわち、塩縮処理のみの絹糸と塩縮・キトサン処理絹糸との両方について、ニンヒドリン試験を行った。このとき、ニンヒドリンとの反応にあずかるのは、塩縮処理絹糸の場合は、絹糸を構成している塩基性アミノ酸の塩基性基と末端アミノ基であり、塩縮・キトサン処理絹糸の場合は、これらの基に加えて、導入されたキトサンの中の未反応のアミノ基である。また、基質中の塩基性基1当量とニンヒドリン1当量(1モル)とが反応するので、検量線を作成する場合の標準物質としてキトサンの構成単位(繰り返し)であるD-グルコサミンの塩酸塩を選んだ。
【００５８】
測定により得たグルコサミン濃度と吸光度との関係(検量線)を図1に示した。この検量線の回帰直線式は下式で示される。
A=0.1527C (式1)
上式中、Aは吸光度(単位を持たない)を、Cはグルコーサミン濃度(mol・dm^-3 )を意味する。
次に、塩縮処理絹糸および絹タンパク質とキトサンとの複合体について、ニンヒドリン反応で得られた吸光度を式1に代入し、グルコサミン濃度(mol・dm^-3 )として求め、アミノ基量を算出した。得られた各絹糸試料のアミノ基量を表3に示した。
【００５９】
（表3）

なお、表3中の導入キトサン量は、塩縮・キトサン処理で絹糸に新たに付加したアミノ基の導入量に対応する。
表3から分かるように、塩縮絹糸へのキトサン導入量は0.44×10^-5 mol・g^-1であった。キトサン導入量を更に増やすには、キトサンを溶解量の限界まで溶かした溶液を用いればよい。
なお、上記で用いた塩縮絹糸(塩縮率14%)の代わりに塩縮率の度合いを上げた塩縮絹糸を用いて、上記と同様にキトサンの酢酸水溶液で処理し、ニンヒドリン試験を行って導入キトサン量を評価した。塩縮率が上がるにつれて、導入キトサン量が増加した。しかし、塩縮率が上がるにつれて塩縮絹糸の機械的性質は劣化するので、所望の結果を達成するには、塩縮率を30%以下にコントロールすることが必要である。
【００６０】
(実施例5)
未処理絹糸(A)、塩縮処理(塩縮率15%)・キトサン処理絹糸(B)、塩縮処理(塩縮率15%)・キトサン処理絹糸にエポキシ試薬で架橋結合を導入した絹糸(C)を切断するまで引きのばし、切断時の強度と伸度とを測定することで機械的特性を調べた。得られた結果を表4に示す。
【００６１】
(表4)

注：強度430±18の意味は、強度が430gf、標準偏差が18であることを意味する。
表4から明らかなように、塩縮率が15%程度であると、塩縮処理後の絹糸(B)の強度、伸度は、未処理絹糸(A)の場合と比べて大きな変化はない。しかし、塩縮率が30%を超えると、塩縮処理絹糸の強度、伸度も急激に低下してしまう。また、架橋結合を導入した絹糸(C)の機械的特性も本来の絹糸の特性を保持しており、加工時の機械特性の劣化がない。
【００６２】
(実施例6)酸性染料による染色
絹タンパク質とキトサンとの複合体(架橋結合を導入してキトサンを固定化したもの。以下、固定化絹糸と称す。)の染色挙動を次のようにして調べた。
代表的な酸性染料であるC.I.Acid Orange 7(Orange II：以下ORと略記)で絹タンパク質とキトサンとの複合体を染色した。すなわち、酸性染料をpH 4の0.1 mol・dm^-3の酢酸緩衝溶液に溶解させた染色溶液200mLに被検試料を入れ、40℃で72hrs撹拌し、試料に染料を吸着させた。染料を吸着した後の染色浴を紫外・可視分光光度計((株)日立製作所製1100型)を用い、λ=485 nmにおいて比色して遊離染料濃度(×10^-5mol・dm^-3)を求めた。また、吸着量は次のようにして求めた。染色絹糸から、染料をホルムアミドで60℃、30 minの抽出処理を3回行って完全に抽出し、50mlに定容した溶液を同様に比色して、絹糸による吸着染料量(×10^-5mol・g^-3)を算出した。 なお、未処理絹糸、塩縮処理のみの絹糸(以下、塩縮絹糸と称す。)の染料吸着測定も併せて行った。得られた吸着等温線(pH=4,40℃)を図2に示す。
【００６３】
図2から分かるように、低濃度においては、ORの吸着量にほとんど差がみられなかったが、比較的高い染料濃度においては、絹タンパク質とキトサンとの複合体(図2中の固定化処理)に対する吸着量は塩縮絹糸(図2中の塩縮処理)および未処理絹糸(図2中の未処理)に対する吸着量を大きく上回っており、キトサン固定化の効果は明白である。
【００６４】
キトサン固定化絹糸へのORの吸着機構想定図を図3に示す。図3に示したように、例えば、絹分子のアミノ基にORが直接イオン結合したり、絹分子にEGDGEを介して結合したキトサン(Chitosan)にORが結合したり、また、絹糸に入り込んだキトサン同士がEGDGEを介して共有結合した場合の各キトサンにORが結合するものと考えると、本発明の絹タンパク質とキトサンとの複合体への染料の吸着量が増加する現象は矛盾無く説明できる。
また、図2のデータを基にScatchard plotにより算出した各絹糸の染料の結合座席数を表5に記した。この結果より、絹タンパク質とキトサンとの複合体(固定化絹糸)は未処理絹糸の1.4倍の結合座席を有していることになる。
【００６５】
(表5)

【００６６】
(実施例7)生成ポリマーの橋かけの確認
表1に示した橋かけキトサンNo.2と甲陽ケミカル株式会社製キトサンFM80(脱アセチル化度85%)とのFT-IRスペクトルを測定し、それらの差スペクトルを図4に示す。この差スペクトル中の2959cm^-1と1472cm^-1に新しいピークが見られる。これは、EGDGEの導入に伴ったC-C結合の増加により生じた。また、1600cm^-1付近のアミノ基(-NH₂)のNHの変角振動のピークが減少している。架橋剤の仕込比の異なる橋かけキトサンNo.1, No.3に関しても、同様のピークが検出された。以上のことより、キトサンとEGDGEの反応を確認できた。
【００６７】
また、橋かけキトサンNo.1〜3について、5%酢酸、5%ギ酸による溶解試験を行ったところ、いずれも不溶であったことから、橋かけの存在が認められた。
上記のことから、キトサンが橋かけされていることが確認された。キトサンとキトサンとに架橋結合が導入されたことから、同様に絹とキトサンとの分子間にも架橋結合が導入されていると判断できる。キトサンおよび絹に対する反応染料の染着挙動が類似していることから、絹中の塩基性基のEGDGEに対する反応性とキトサンのアミノ基のそれも似ていると考えられる。
【００６８】
また、キトサン固定化絹糸に関し、キトサンが絹糸内で固定化していることは以下のことからも間接的に証明できる。すなわち、実施例6に示した染色は、pH 4の0.1 mol dm^-3の酢酸緩衝溶液中で、40℃、72hrsもの長時間行われていることから、もし、キトサンが固定化されていなければ、このような条件の下では絹中からキトサンが溶出してしまうと考えられるからである。
【００６９】
(実施例8)アルカリ溶解度
未処理絹糸、および架橋結合を導入した絹タンパク質とキトサンとの複合体のアルカリ溶解度を評価した。
絹糸単独のアルカリ溶解度が42%であるのに対して、架橋結合を導入した絹タンパク質とキトサンとの複合体のアルカリ溶解度は30%となった。このことから、未処理絹糸の場合のアルカリ水溶液に溶解しやすいという欠点は、キトサンと複合したものに架橋結合を導入することで解決できることが分かる。
【００７０】
(実施例9)金属イオンの吸着
上記金属イオンの吸着方法に従って調製した金属塩水溶液として、0.5mMの硝酸銀(AgNO₃)水溶液および硝酸銅(Cu(NO₃)₂)水溶液を用いた。それぞれの水溶液に、未処理家蚕絹糸、塩縮処理家蚕絹糸、塩縮・キトサン処理家蚕絹糸、および塩縮・キトサン処理後架橋結合を導入して不溶化した家蚕絹糸の試料をそれぞれ、金属イオンの水溶液に25℃で30分間浸漬し、銀イオン、銅イオンを吸着させて、各試料に吸着した金属イオン量を測定した。得られた結果を表6に示す
【００７１】
（表6）

表6から分かるように、未処理家蚕絹糸の場合、金属イオン水溶液に浸漬するとAg⁺, Cu²⁺ともにわずかに吸着するが、この吸着金属イオン量は塩縮処理によりわずか増加する。塩縮絹糸にキトサンを含浸すると金属イオンの吸着率は更に増加し、キトサン含浸試料をエポキシ化合物で不溶化したものは更に多くの金属イオンを吸着することが確かめられた。
【００７２】
【発明の効果】
本発明の絹タンパク質とキトサンとの複合体によれば、キトサン同士の架橋結合に加えて、キトサンと絹タンパク質との間に架橋結合が導入されているので、単独の各素材の特性に加えて、優れた染色性、耐薬品性等の機能を付与することができる。
本発明の複合体では、絹タンパク質として、中性塩水溶液中で塩縮処理により前処理した衣料用汎用繊維である絹糸を用いることができるので、絹糸の優れた機械的特性に加えて、様々な生化学特性を持つキトサンの特性を兼ね備えたキトサン含有の絹糸を提供できる。
【００７３】
本発明の複合体によれば、架橋結合の導入が、好ましくは二官能性、三官能性、多官能性のエポキシ化合物からなる架橋剤を用いて行われるので、キトサンの酢酸水溶液中で他の触媒を用いることなく架橋結合がスムーズに導入される。
また、本発明の複合体の製造方法によれば、衣料用汎用繊維である絹糸を好ましくは前処理として塩縮処理し、この塩縮絹糸にキトサン水溶液、好ましくはキトサンを溶かした酢酸水溶液を含浸させてキトサン含有絹糸を製造し、このキトサン含有絹糸に架橋剤としてエポキシ化合物を作用させて、絹タンパク質とキトサンとの間及び絹糸に充填されたキトサン同士に架橋結合を導入している。かくして、絹糸に含まれるキトサンが脱離することはないし、調製できる絹タンパク質とキトサンとの複合体を水中に長時間浸漬しても架橋結合がはずれることはない。その結果、絹タンパク質単独の特徴とキトサン単独の特徴を併せ持つ複合材料の提供が可能となった。
【図面の簡単な説明】
【図１】 グルコサミン濃度と吸光度との関係(検量線)を示すグラフ。
【図２】 吸着等温線を示すグラフ。
【図３】 キトサン固定化絹糸へのORの吸着機構想定図。
【図４】 橋かけキトサンNo.2とキトサンFM80のFT-IRスペクトルの差スペクトルを示すグラフ。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a complex of silk protein and chitosan and a method for producing the same.
[0002]
[Prior art]
Silk yarn has been used mainly as a material in the Japanese clothing field because it has excellent practical properties as a clothing material in terms of moisture absorption, dyeability, and texture. Recently, the amount of cheap silk products imported has increased, and silk products have been widely used in the Western clothing field. Silk has long been used as a woven material, but it has not been used as a knitted material because silk has no crimp. If silk thread can be used as a material for knitting, it will be possible to increase the demand for silk thread.
[0003]
Therefore, conventionally, attempts have been made to impart a specific function to the silk thread by subjecting the silk thread to salt shrinkage. That is, when the silk thread is immersed in a heated aqueous neutral salt solution, the silk thread swells in a short period of time from several tens of seconds to several minutes after immersion, and contracts significantly in the fiber length direction (this is called salt crimping, which is called crimping). Also). As a neutral salt for salinating silk thread, calcium nitrate is well known, and a method of treating it with an aqueous solution is common (for example, see Non-Patent Document 1). In this case, the density is between 1.410 and 1.420 g / cm^ThreeWhen the silk thread is immersed in the heated calcium nitrate aqueous solution, the salt contraction rate reaches 40% in the salt contraction process in which it is immersed in the neutral salt aqueous solution for about 1 minute. When a silkworm silk thread is immersed in an aqueous neutral salt solution, the silkworm silk swells and the fiber length shrinks in a short immersion time of about 20 seconds, and as a result, the silk thread surface becomes irregular and crimps. By such salt shrinkage treatment, the silk thread becomes soft, and the bulk of the silk thread increases, resulting in a light and soft touch. Bulky materials are soft and contain a lot of air, so they are favorably used as clothing materials such as knit materials.
[0004]
Chitosan, a natural polymer, is obtained by a “deacetylation reaction” that chemically removes the acetyl group of chitin contained in the crustacean shells such as krill and crabs, and is widely used as various industrial materials. They are used and their applications are diverse. In addition to its use as a fiber itself, it can also be used as a processing agent for clothing. Thus, crustacean-derived chitin and chitosan are high-value-added materials in various fields.
Although chitosan has excellent protein adsorption ability, when powdered or flaky chitosan is used as it is, its application is limited because of its poor moldability and physical strength, and it gives practical value. It was difficult to take a usage form. When chitosan is dissolved in an organic acid such as lactic acid, formic acid, acetic acid, methanesulfonic acid, etc., if the acidity of the organic acid is strong, chitosan's original high molecular weight decreases, and the resulting molded product has poor moldability. turn into.
[0005]
Therefore, since chitosan is well dissolved in an acetic acid aqueous solution, attempts have been made to form shaped articles such as fibers and membranes by using an aqueous acetic acid solution of chitosan as a starting material. The properties of the fiber and membrane of the chitosan molded product are due to the sugar chain of chitosan, that is, the chemical structure of chitosan, and it is desirable to make use of this property. However, the mechanical properties such as strength and elongation of the fiber that is a chitosan molded product differ depending on whether the molded product is in a dry state or in a wet state, and particularly in a wet state, the properties are significantly reduced. It has become a practical problem.
Focusing on the various functions of chitosan as described above, techniques for attaching chitosan to fibers have been developed. For example, it is known that by pretreating cotton fibers with chitosan, the direct dye is efficiently adsorbed on the fiber material and the amount of dyeing is increased (see, for example, Non-Patent Document 2).
[0006]
There are also attempts to introduce chitosan into silk thread. For example, it has been reported that by attaching chitosan to the surface of a silk thread, the dyeability with a direct dye is improved and the dyeing speed is increased (for example, see Non-Patent Document 3). The chitosan immersion treatment described in Non-Patent Document 3 is to immerse the silk thread in an acetic acid aqueous solution of chitosan and heat-treat, and the chitosan diffuses into the silk thread only by adhering to the surface of the silk thread. They are not connected. That is, in order to attach chitosan to the surface of the silk thread, the silkworm silk thread is simply dipped in an aqueous acetic acid solution of chitosan and then dried. That is, the silk thread is padded (2 dips, 2 nips) in an aqueous acetic acid solution of chitosan so as to cover the surface of the silk thread with chitosan (see, for example, Non-Patent Document 4).
[0007]
In the methods described in Non-Patent Documents 3 and 4 described above, there is a problem that the thin film of chitosan only physically adheres to the surface of the silk thread, and the durability is low because chitosan does not enter the silk thread. In addition, no special chemical bond is introduced between the adhering chitosan and the silk thread, and even if the materials prepared by these conventional methods are dyed with an acid dye or an acid mordant dye, the amount of dyeing increases slightly. The dyeing rate does not increase remarkably (see, for example, Non-Patent Document 5).
[0008]
[Non-Patent Document 1]
Hiroshi Kato, Journal of Japanese Society of Sericultural Science, 59, 271-279, 280-287, 341-349 (1990)
[Non-Patent Document 2]
J.A.Rippon, J.Soc.Dyers and Colorists, 100, 298-303 (1984)
[Non-Patent Document 3]
Takeshi Kako and Akira Katayama, Japanese Journal of Sericultural Science, 68, 429-431 (1999)
[Non-Patent Document 4]
Takeshi Kako and Akira Katayama, Japanese Journal of Sericultural Science, 57, 31-37 (1988)
[Non-Patent Document 5]
Takeshi Kako and Akira Katayama, Japanese Journal of Sericultural Science, 58, 374-379 (1989)
[0009]
[Problems to be solved by the invention]
The chitosan-treated silk thread produced by the above-described conventional technique has a drawback that no improvement in the dyeability exceeding the dyeability of the silk thread not subjected to chitosan treatment is observed (Non-Patent Documents 3 to 5). The disadvantages are that (1) when silk is pad-dried with an aqueous chitosan solution, the amount of chitosan attached to the surface of the silk is extremely small, and (2) chitosan is only physically attached to the surface of the silk. This is due to the fact that chitosan is easily detached from the surface of the silk thread during a reaction process such as a dyeing process. In the general dyeing process, the temperature of the dyeing bath must be raised during dyeing, and the silk thread is repeatedly subjected to washing with water and hot water treatment. Unbound chitosan was easy to desorb.
[0010]
If the chitosan or chitosan molded body can be compensated for and the function of biopolymers such as silk protein with high chemical reactivity can be newly added to chitosan, a material with more diversified functions of chitosan can be prepared. can do. For this reason, there has been a strong demand for the appearance of a composite material that can compensate for the disadvantages of both the fibrous silk protein, which is a natural protein, without deteriorating the excellent function of chitosan alone.
[0011]
In order to solve such conventional problems, it was necessary to introduce a strong bond between chitosan and silk to bond chitosan with silk protein and chitosan and to completely insolubilize the material.
An object of the present invention is to solve the above-mentioned problems of the prior art, and silk protein that does not detach chitosan attached to the surface of the silk thread or contained in the silk thread even in a process such as dyeing. It is an object of the present invention to provide a novel complex of chitosan and a method for producing the same.
[0012]
[Means for Solving the Problems]
The present inventors target silkworm silk, wild silk silk thread, etc., in the first stage as pre-treatment, soaking the silk thread by neutralization with a neutral salt aqueous solution, and the silk microstructure becomes coarse After soaking, the salt-shrinked silk thread is immersed in chitosan acetic acid aqueous solution, and the silk thread is impregnated with chitosan to produce silk protein containing chitosan. Then, in the second stage, the silk protein containing chitosan is cross-linked. A new material is created by introducing chemical bonds between (1) silk protein and chitosan, and (2) chitosan filled in silk thread, and making the silk fibroin silk protein and chitosan complex water-insoluble. Has been found to be able to be produced, and the present invention has been completed.
[0013]
  The complex of the present invention comprisesSilk yarn salt-treated in neutral salt solutionAnd a complex of chitosan,The chitosan is bonded to the silk thread through a cross-linking agent, and chitosans that have entered the silk thread are covalently bonded to each other through the cross-linking agent. The cross-linking agent is glycerol polyglycidyl ether, resorcinol diglycidyl ether, neopentyl Glycol diglycidyl ether, 1,6 A bifunctional or trifunctional epoxy compound selected from hexanediol diglycidyl ether, ethylene glycol diglycidyl ether, and polyethylene glycol diglycidyl ether, or sorbitol polyglycidyl ether, sorbitan polyglycidyl ether, and penta It is a polyfunctional epoxy compound selected from erythritol polyglycidyl ether.
[0014]
  In the method for producing a composite of the present invention, an insect-derived silk thread is subjected to salt-shrinkage treatment in an aqueous solution of a neutral salt selected from lithium bromide, lithium nitrate, lithium chloride, lithium thiocyanate, calcium nitrate, and calcium chloride. Then, after the salt shrinkage treatment, without causing water removal or drying, the silk yarn subjected to the salt shrinkage treatment is immersed in a chitosan-containing acetic acid aqueous solution to produce a chitosan-containing silk yarn.A bifunctional epoxy compound selected from glycerol polyglycidyl ether, resorcinol diglycidyl ether, neopentyl glycol diglycidyl ether, 1,6-hexanediol diglycidyl ether, ethylene glycol diglycidyl ether, and polyethylene glycol diglycidyl ether, or Trifunctional epoxy compound or polyfunctional epoxy compound selected from sorbitol polyglycidyl ether, sorbitan polyglycidyl ether, and pentaerythritol polyglycidyl etherThe cross-linking agent is allowed to act at 40 to 80 ° C. in the presence of water or an organic solvent, so that the chitosan is bonded to the silk thread via the cross-linking agent, and the chitosans that have entered the silk thread are interlinked via the cross-linking agent. Thus, a cross-linking bond is introduced so as to be covalently bonded to obtain a composite of the silk thread and chitosan. If the temperature of the cross-linking reaction is less than 40 ° C., the reaction efficiency is low and the reaction takes a long time. In addition, if the temperature of the crosslinking reaction exceeds 90 ° C., it is not economically efficient from the viewpoint of energy efficiency..
[0015]
  Immersion of the salt-shrinked silk thread in a chitosan-containing aqueous solution is performed by immersing in an aqueous acetic acid solution having a concentration of 5 to 15% at 10 to 50 ° C. for 1 to 24 hours.It is preferable.
[0016]
  The introduction of the cross-linking bond to the chitosan-containing silk is preferably performed in water containing a cross-linking agent or in a tetrachloroethane solvent at a temperature of 40 to 90 ° C. The amount of the cross-linking agent added is chitosan and silk It is preferably 5 to 100% of the total number of moles of total basic groups contained therein.
[0017]
DETAILED DESCRIPTION OF THE INVENTION
Examples of the object to be treated in the present invention include natural protein fibers that are silk protein fibers such as rabbit silk thread and wild silk thread. The form of the object to be treated may be a fiber, a fiber product, or a fiber assembly thereof.
[0018]
Hereinafter, an embodiment of the present invention will be described by taking silk thread as an example.
In the present invention, as a pretreatment, the silk thread is subjected to salt shrinkage using a neutral salt aqueous solution, and then the salt crimped silk thread is immersed in an aqueous acetic acid solution of chitosan to introduce chitosan into the silk thread, and then introduced into the silk thread in the next step. The present invention also relates to a composite made of chitosan-containing silk thread obtained by chemically bonding chitosan and silk thread to each other with a cross-linking agent. According to the present invention, the salt shrinkage rate is suppressed to a certain value or less in the salt shrinkage treatment so as not to deteriorate the original mechanical properties of the silk thread. In this salt shrinkage process, the fine structure of the silk thread is roughened, and when this silk thread is immersed in an aqueous acetic acid solution of chitosan, the chitosan is easily diffused and introduced into the silk thread. According to the present invention, the original mechanical properties of silk thread are not deteriorated in the process of producing the “complex of silk protein and chitosan”, and the original properties of silk can be maintained. Moreover, the characteristics, such as the outstanding dyeing | staining property which chitosan has, can also be provided to a silk thread.
[0019]
Thus, by introducing chitosan into the fibrous silk thread, a silk thread containing chitosan is obtained, and cross-linking is introduced between chitosan and silk protein and between chitosan and chitosan filled in the silk thread. Both are definitely water-insoluble.
The complex of silk protein and chitosan according to the present invention can be used as a complex of various forms such as a fiber, a woven, a cotton, a nonwoven, a powder, and a film.
Since the dyeability of this chitosan-containing silk composite is higher in the dyeing rate than in the case of untreated silk, the range of use as various industrial materials including clothing materials is expanded.
[0020]
According to the present invention, as the neutral salt used when performing the salt shrinkage processing as the pretreatment, for example, as described above, lithium bromide, lithium nitrate, lithium chloride, lithium thiocyanate, calcium nitrate, or Calcium chloride and the like can be mentioned, and these neutral salts may be anhydrous or hydrated. The heat treatment in the salt shrinkage processing is generally performed at 60 to 95 ° C, preferably 75 to 95 ° C, more preferably 80 to 90 ° C. If the temperature is less than 60 ° C, salt shrinkage hardly occurs. If the temperature exceeds 95 ° C, salt shrinkage occurs, but swelling of rabbit silk thread or the like proceeds so much that it becomes difficult to control the salt shrinkage rate. Further, the salt shrinkage treatment time is related to the type and concentration of the neutral salt, but may be adjusted as appropriate while observing the contraction state of the silkworm silk. Generally, it is 1 minute to 30 minutes, preferably 5 to 30 minutes. In particular, about 20 minutes is convenient for showing a stable salt shrinkage effect.
[0021]
In the salt shrinkage processing, a high-concentration neutral salt aqueous solution is used, and when treated at a high temperature for a long time, a silk thread having a high salt shrinkage rate can be obtained. The salinity rate is preferably 30% or less, and if it exceeds 30%, the original mechanical properties of the silk thread will deteriorate. Further, when the salinity is less than 5%, the amount of chitosan introduced is reduced, and the intended purpose cannot be achieved. Therefore, in order to obtain a silk thread that maintains desired properties without reducing strength or elongation, the salt shrinkage rate is desirably suppressed to 30% or less. Generally, the salt shrinkage rate is 5 to 30%, preferably 10%. ~ 20%.
As described above, in order to impregnate even a little amount of chitosan into the salt-crimped silk thread, it is desirable to use a silk thread with a high salt-shrinkage ratio, but the higher the salt-shrinkage ratio, the worse the mechanical properties of the silk thread. It is desirable to perform predetermined processing in consideration of characteristics. Therefore, depending on the purpose, it is necessary to take into consideration the balance between the salinity ratio and the chitosan content, and use silk threads within the above-mentioned range of salinity ratio.
[0022]
The chitosan that can be used in the present invention may be a known chitosan or a chitosan prepared by N-deacetylation of chitin by a known method as described below.
Chitosan can be obtained, for example, by N-deacetylation by treating chitin derived from crustaceans in a 40-50% sodium hydroxide solution at 80-120 ° C. for several hours. In this case, conventionally known conditions can be applied as the operating conditions such as the drug used for N-deacetylation and pH. For example, the conditions described in Brine et al., Comp. Biochem. Physiol., 69B, 283 (1983) are applicable.
The outline of known deacetylation for removing the acetyl group of chitin is described below. When deacetylation is performed in a homogeneous system using the fact that chitin is dissolved in concentrated alkali, partially deacetylated chitin becomes soluble in water only when the degree of deacetylation is in the range of 40 to 60%. The standard preparation method is as follows.
[0023]
3.0 g of powdered chitin (80 mesh) is suspended in 75 g of 40 wt% NaOH and left under vacuum at 25 ° C. for 3 hours. While mechanically stirring the produced alkaline chitin, 255 g of crushed ice is added to prepare a solution containing 1% chitin and 10% alkali. Next, 300 g of this solution is allowed to stand at 25 ° C. for 77 hours, and then cooled to 5 ° C. or lower. In this case, 210 g of crushed ice is added to prevent temperature rise due to heat of neutralization, and concentrated HCl and then dilute HCl are added until the pH is 8.7. The obtained solution is dropped into 5 L of an acetone / water mixture (acetone: water = 7: 1) at 0 ° C. (in order to keep the acetone concentration substantially constant, cold acetone is also dropped simultaneously). The produced white fibrous precipitate is washed with an acetone / water mixture having the same blending ratio as described above until NaCl is eliminated, and further washed with acetone and then dried over phosphorus pentoxide. Thus, chitosan which is deacetylated chitin having a predetermined degree of deacetylation is obtained.
[0024]
When obtaining chitosan, which is partially deacetylated chitin with different degree of deacetylation using chitin as a starting material, the degree of deacetylation should be 70% to 100%, and solubilization in acetic acid aqueous solution should be 90% or more Is particularly preferable since it becomes easier. Silk yarn dipped in an aqueous acetic acid solution of chitosan having a degree of deacetylation lower than 70% is dyed with a typical acid dye, for example, CI Acid Orange 7 (Orange II) (hereinafter this dye may be abbreviated as OR). Then, the higher the degree of acetylation, the lower the amount of dye adsorbed. This is because there are few amino groups which are adsorption seats of dye. If the degree of deacetylation is lower than 70%, it is difficult to dissolve in an aqueous acetic acid solution. As a result, even when treated with an epoxy compound, crosslinking between silk fibroin (silk protein) and chitosan hardly occurs.
[0025]
The chitosan thus obtained can be dissolved in an organic acid such as lactic acid, formic acid, acetic acid, and methanesulfonic acid. Therefore, in order to impregnate the above-mentioned salted silk yarn with chitosan, the organic acid aqueous solution containing chitosan is used. What is necessary is just to immerse the salt crimped silk thread for a fixed time. However, when high molecular weight chitosan is hydrolyzed to lower the molecular weight, the characteristic of chitosan having good moldability is lost. A preferred organic acid that does not reduce the molecular weight inherent in chitosan is acetic acid among the above organic acids. The theoretical basis is as follows.
[0026]
Pka (= -log ka) is obtained from an acid dissociation constant ka (a larger value means a stronger acid) which is a measure of the strength of the acid. The smaller this value, the stronger the acid. Among the above organic acids, the pka values of acetic acid, formic acid, and lactic acid can be estimated as 4.76, 3.75, and 3.86, respectively. Therefore, the most preferable for dissolving chitosan is acetic acid, which has a large pka value, is inexpensive in price, and has low toxicity to the living body. A large pka value means that when chitosan is dissolved, the molecular weight of chitosan is not lowered significantly because it acts gently on chitosan and dissolves it. Among the acids mentioned above, methanesulfonic acid has the highest risk of lowering the molecular weight of chitosan when it dissolves chitosan, so it must be used with caution.
[0027]
When chitosan is contained in the salt-shrunk silk thread, it is preferable to use, for example, a solution in which chitosan is dissolved in an aqueous acetic acid solution having a concentration of 5 to 15%. If the acetic acid concentration is less than 5%, the chitosan is not sufficiently dissolved, and if it exceeds 15%, the chitosan may be slightly hydrolyzed. In order to immerse the silk thread that has been salt-shrinked in an acetic acid aqueous solution, the sample may be allowed to stand, the immersion temperature is 10 to 50 ° C., preferably 25 to 45 ° C., and the immersion time may be 1 to 24 hours. The dyeing effect of the composite of silk thread and chitosan thus obtained is higher as the amount of chitosan introduced into the silk thread is larger. The amount of chitosan introduced is determined by the amount (volume) of the amorphous region that is increased by salt-shrinking the silk thread.
[0028]
The “complex of silk protein and chitosan” in which chitosan is impregnated into the silk thread obtained as described above is then subjected to a crosslinking treatment to introduce a crosslinking bond. Examples of the crosslinking agent that can be used to introduce this crosslinking bond include known protein crosslinking agents such as epoxy compounds, aldehyde compounds such as glyoxal and glutaraldehyde, isocyanate compounds such as hexamethylene diisocyanate, and epichlorohydride. It is conceivable to use an epoxy-chloroether compound such as phosphorus, triacryloyl hexahydrotriacin (TAF), or the like. Among the above-mentioned crosslinking agents, an epoxy compound is preferably used because the crosslinking reaction of chitosan can proceed smoothly without using another catalyst in an aqueous acetic acid solution of chitosan.
[0029]
Examples of the epoxy compound include bifunctional, trifunctional, or polyfunctional epoxy compounds as described below.
For example, a bifunctional epoxy selected from glycerol polyglycidyl ether, resorcinol diglycidyl ether, neopentyl glycol diglycidyl ether, 1,6-hexanediol diglycidyl ether, ethylene glycol diglycidyl ether, and polyethylene glycol diglycidyl ether It is preferable to use a compound or a trifunctional epoxy compound or a polyfunctional epoxy compound selected from sorbitol polyglycidyl ether, sorbitan polyglycidyl ether, and pentaerythritol polyglycidyl ether.
[0030]
Among these various epoxy compounds, bifunctional epoxy compounds such as ethylene glycol diglycidyl ether (hereinafter abbreviated as EGDGE) are (1) a solution-free homogeneous system containing no solids and without catalyst. It is most preferable because it can be crosslinked, (2) has high reactivity, and (3) its molecular weight is not so high. EGDGE, a bifunctional epoxy compound, has a relatively low molecular weight and is susceptible to crosslinking reactions in homogeneous systems. In addition, since the molecular weight is small, EGDGE is sufficiently diffused into the chitosan portion in the fiber and serves as a crosslinking agent.
[0031]
When a crosslink is introduced into the chitosan-containing silk thread by the epoxy compound, the chitosan-containing silk thread is treated in water containing an epoxy compound at a predetermined concentration or in an organic solvent such as tetrachloroethane. In this case, the reaction temperature is about 40 to 90 ° C, preferably about 60 to 80 ° C. A reaction temperature of less than 40 ° C. is not preferable because the reaction efficiency is low and the reaction takes a long time. Usually, the cross-linking reaction of chitosan is carried out in an aqueous solution diluted with methanol. Therefore, the reaction is preferably carried out at as low a temperature as possible and at a temperature with good reaction efficiency. For example, when the reaction is performed at about 70 ° C., the reaction time is usually 2 hours or longer, preferably about 3 hours. When the reaction system is carried out with an organic solvent such as tetrachloroethane described above instead of water, the reaction efficiency is remarkably increased. However, from the viewpoint of ease of work and environmental conservation, the reaction in the aqueous system is preferred.
[0032]
When using EGDGE as a crosslinking agent, the amount of epoxy compound added is usually about 5 to 100%, preferably about 5 to 50% of the total number of moles of total basic groups contained in the chitosan and silk protein used. Good. If the amount of EGDGE is less than 5% of the total number of moles of total basic groups, the crosslinking effect is weakened, and chitosan and silk protein impregnated in silk cannot be sufficiently water-insolubilized. In addition, when the amount of EGDGE exceeds 100%, the substitution rate of the epoxy compound with respect to the basic group is increased, and a two-dimensional network is excessively formed, so that the dye penetrates into the sample or the dye penetrates into the sample. Since the seat (basic group in the substrate) serving as a reaction base is reduced, the amount of dyeing is reduced. In order to increase the amount of dyeing to the sample as much as possible, it is desirable to control the degree of reduction of the basic group accompanying the crosslinking reaction so as not to be excessive.
[0033]
A monofunctional epoxy compound having one epoxy group reacts with silk or chitosan, but cannot be used because it cannot crosslink between molecules due to its monofunctionality. The trifunctional or polyfunctional epoxy compound reacts with silk or chitosan to form a three-dimensional network, and the complex of silk and chitosan is insolubilized. As a result, the chemical resistance is improved, but the crosslinking density is too high and the dye is difficult to diffuse, which is not preferable from the viewpoint of improving the dyeability. Therefore, when using these epoxy compounds, it is necessary to control the formation of a three-dimensional network as desired.
[0034]
Aldehyde compounds are the most commonly used reagents for protein cross-linking, but as a result of the reaction of aldehyde compounds, -CH = N- bonds are introduced into the protein, causing protein to color, and even at room temperature, hydrochloric acid solution There is a disadvantage that it will be disassembled. This is due to the property of the protein that when an aldehyde compound introduces a cross-linking bond to the protein to insolubilize it, it is easily hydrolyzed with an aqueous hydrochloric acid solution. Therefore, when using an aldehyde compound, the -CH = N site introduced into the protein as a result of the reaction of the aldehyde compound is reduced, and -CH₂It is necessary to reduce the degree of hydrolysis with hydrochloric acid solution in the form of -NH-. In general, an isocyanate compound reacts rapidly in the presence of water, and it is difficult to control the amount of crosslinking introduced, and therefore it is preferable to carry out the reaction in an anhydrous organic solvent system. Therefore, in the present invention, when the crosslinking reaction is carried out in an aqueous system, an epoxy compound is preferable to an isocyanate compound.
[0035]
As described above, both chitosan and silkworm-derived silk protein, which are natural polymers, are promising materials as natural biopolymers, and both contain many active groups that serve as bases for chemical reactions. By subjecting these materials to chemical modification, functions such as dyeability and chemical resistance can be imparted in addition to the properties of a single material. In addition, silk protein containing chitosan can produce chitosan-containing silk yarn having the characteristics of chitosan having various biochemical properties in addition to the excellent mechanical properties of silk yarn.
[0036]
The composite of the present invention is made by impregnating silk thread, which is a general-purpose fiber for clothing, with an acetic acid aqueous solution in which chitosan is dissolved to infiltrate chitosan into silk thread, and then allowing an epoxy compound to act as a crosslinking agent to produce silk protein and chitosan. And a cross-linked bond between chitosans filled in silk thread. For this reason, chitosan contained in the silk thread is not detached, and even if the prepared “complex of silk protein and chitosan” is immersed in water for a long time, the cross-linking bond is not released. As a result, it has become possible to produce a composite material that combines the characteristics of silk protein alone and chitosan alone.
Chitosan fiber made of chitosan is a practical problem that it shows low strength and elongation when wet, but the material that can be prepared by introducing chitosan into silk thread and introducing cross-linking with cross-linking agent, While taking advantage of the strength of silk thread, it has excellent characteristics that strength and elongation do not decrease even when wet. When dyed with an acid dye, the dyeability of the chitosan-containing silk thread was clearly improved as compared with the silk thread alone, and the effect of introducing chitosan was recognized.
[0037]
The dye used for dyeing the composite of silk fibroin and chitosan introduced with the cross-linking of the present invention includes a cationic dye, a disperse dye, and an anionic dye such as a reactive dye, a direct dye and an acid dye. Any vat dye can be used. In particular, it can be satisfactorily dyed with known acid dyes, and in general, the degree of dyeing of cationic and nonionic dyes is not as high as that of acid dyes. Examples of acid dyes include CI Acid Orange 7, CI Acid Red 88, CI Acid Red 138, CI Acid Red 1, CI Acid Blue 9, CI Acid Blue 15, CI Acid Blue 22, and CI Acid Orange 7. Can do. Since silk is a hydrophilic fiber and silk chitosan complex is also hydrophilic and has a basic group that serves as a dyeing seat, -SO₃H or -SO_ThreeAn acid dye that is Na-containing and easily soluble in water is preferably used. According to the present invention, in addition to the amino acid residue involved in the adsorption of the dye contained in the silkworm silk, it is also affected by the active group of the introduced chitosan, so that the basic serving as an adsorption seat for the complex of silk protein and chitosan Since the total number of groups, acidic groups, hydroxyl groups, and nonpolar groups is larger than that of silk alone, the complex of silk protein and chitosan of the present invention can be effectively dyed with various dyes.
[0038]
Since the complex of the present invention has properties of both silk protein and chitosan which are rich in chemical reactivity, it can be used as an adsorbent for metal ions and surfactants in waste liquids, or a carrier for immobilizing enzymes.
Chitosan can be used as an antibacterial food retention agent. Since chitosan and its degradation products show growth inhibitory activity against phytopathogenic bacteria, chitosan-containing silk protein can be used as a soil conditioner and a natural pesticide. Chitosan is easily degradable against natural microorganisms. Since chitosan has a wound healing promoting effect and a microphage activity ability, chitosan-containing silk thread can be used for artificial skin, sutures, shampoos, and as a sustained-release drug carrier with immunopotentiating ability.
[0039]
In the present invention, the amount of chitosan introduced into the silk thread can be freely increased by changing the degree of salt shrinkage of the silk thread. Compared with chitosan alone, the material with chitosan introduced into silk thread exhibits the superior mechanical properties of silk alone as a synergistic effect, so it is possible to produce high-strength fibers by producing a composite of silk and chitosan. It is possible to manufacture. If desired, the chitosan contained in the complex can be acetylated into chitin to be used as a silk fibroin / chitin complex. In this case, chitin and chitosan not only cover the silk surface, but also diffuse in the silk fibroin matrix and are chemically bonded to the silk fibroin through a cross-linking reaction. Even if a chemical or chemical action is added, it does not leave.
[0040]
In the present invention, if a large amount of chitosan is introduced into ultra-fine silk thread, it has the antibacterial property, cell adhesion proliferation property, wound healing promotion effect, microphage activity ability and immunity enhancement ability, taking advantage of the properties of hard silk thread like steel New materials with the characteristics of chitosan can be prepared.
According to the present invention, since chitosan is included in the silk thread, if desired, the chitosan included in the silk thread is converted into chitin by acetylating the included chitosan by a conventionally known method, and the novel has the characteristics of fine chitin fibers. It can be used as a material. Thus, in addition to the conventionally used silk thread, a new material having both chitin and chitosan applications can be prepared.
[0041]
【Example】
EXAMPLES Next, although an Example demonstrates this invention in detail, this invention is not limited to these examples. Before describing specific examples, first, samples, measurement conditions, methods for synthesizing crosslinked chitosan, conditions for adsorbing acid dyes, and the like in the following examples will be described.
[0042]
(1) Sample
Chitosan FM80 (degree of deacetylation 85%) manufactured by Koyo Chemical Co., Ltd. was used as chitosan. Ethylene Glycol Diglycidyl Ether (ethylene glycol diglycidyl ether, hereinafter abbreviated as EGDGE) from Aldrich Chemical Company, Inc. was used as a crosslinking agent.
As the neutral salt calcium nitrate tetrahydrate used for the salt shrinkage treatment, a commercially available reagent-grade product manufactured by Wako Pure Chemical Industries, Ltd. was used.
As reagents for the ninhydrin reaction, hydrochloric acid-D-glucosamine (Wako first grade), ninhydrin (for automatic amino acid analysis), tin (II) chloride dihydrate (for automatic amino acid analysis), methyl, manufactured by Wako Pure Chemical Industries, Ltd. Cellosolve (reagent grade), acetic acid (for precision analysis), and sodium acetate trihydrate (for automatic amino acid analysis) were used.
As a dye, C. I. Acid Orange 7 (Orange II) (hereinafter, this dye is sometimes abbreviated as OR) was synthesized and purified before use.
[0043]
(2) Determination of amino groups in salt-crimped silk and salt-fried chitosan-treated silk
The amount of chitosan combined with silk fibroin (silk) was evaluated as follows. Focusing on the correspondence between 1 equivalent of amino group of chitosan crosslinked with silk fibroin and 1 equivalent of glucosamine, we evaluated the chemical reactivity of hydrochloric acid-D-glucosamine with a known structure as a compound that reacts with ninhydrin.
[0044]
First, a pH 5.5 buffer solution [4 mol / l- (CH_ThreeCOOH + CH_ThreeCOONa)] was prepared. Next, 0.04313 g of hydrochloric acid-D-glucosamine (2.000 × 10^-Fourmol) was dissolved in a 4 mol / l acetate buffer solution, and the volume was adjusted to 100 ml to prepare a hydrochloric acid-D-glucosamine solution. Also, 5 g of ninhydrin was dissolved in 187.5 ml of methyl cellosolve (ethylene glycol monomethyl ether), 0.10 g of stannous chloride was added, and the volume was adjusted to 250 ml with a 4 mol / l acetic acid buffer solution to prepare a ninhydrin solution. A small amount of the thus prepared hydrochloric acid-D-glucosamine solution and ninhydrin solution was mixed in a test tube in a certain amount and heated in a boiling water bath with stirring for 1.5 hrs. After heating, the mixture was cooled with cold water, made up to 100 ml with distilled water, and colorimetrically measured at λ = 565 nm using an ultraviolet / visible spectrophotometer (1100 model, manufactured by Hitachi, Ltd.). A calibration curve for amino group quantification was prepared with the vertical axis representing the value obtained by subtracting the absorbance obtained as a result of the blank test and the horizontal axis representing the hydrochloric acid-D-glucosamine concentration.
[0045]
(3) Ninhydrin reaction
Put 0.01g of salt-treated and chitosan-treated silk or untreated chitosan-treated silk into a test tube, add 2ml of ninhydrin solution, 2ml of 4mol / l acetate buffer solution, and stir in a water bath for 1.5hrs Heated. After heating, the mixture was cooled with water, filtered through a glass filter, and the volume was adjusted to 100 ml with distilled water. The solution was colorimetrically measured at λ = 565 nm, and the amino groups of these silk threads were quantified based on the calibration curve.
(4) Immobilization of chitosan on silk thread
Salt shrinkage treatment-0.44 g of EGDGE was added dropwise to 250 ml of water containing 0.7 g of chitosan-treated silk thread, and the mixture was reacted at 70 ° C with stirring for 3 hrs. After the reaction, the mixture was filtered, washed several times with diethyl ether and acetone, then air dried and vacuum dried.
[0046]
(5) Synthesis of bridged chitosan (using EGDGE)
EGDGE (1) 0.44g, (2) 2.20g, (3) 4.4g as a cross-linking agent was added dropwise to 250ml aqueous solution containing 2.5g of chitosan, respectively, and stirred at 70 ° C for 3hrs, reacted to react with 3 types of crosslinking Chitosan (No. 1-3) was obtained. After the reaction, the mixture was filtered through a No. 3 glass filter, washed several times with diethyl ether and acetone, then air dried and vacuum dried. In addition, the elemental analysis results of the crosslinked chitosan Nos. 1 to 3 measured using an elemental analyzer (manufactured by PerkinElmer) PE-2400 CHN are shown in Table 1, and the reaction rate of the crosslinked chitosan obtained from the analytical values ( Table 2 shows the reaction rate of amino groups, the number of moles of amino groups, and the charge ratio of the crosslinking agent (epoxy groups / amino groups).
[0047]
(table 1)

[0048]
(Table 2)

[0049]
(6) Confirmation of introduction of bridge structure
Using a Fourier transform infrared spectrophotometer (JASCO Corporation, FT-IR 430), the IR difference spectra of the bridged chitosan Nos. 1 to 3 and the raw material chitosan FM80 were measured to confirm the reaction. Moreover, bridge | crosslinking chitosan No. 1-3 was added to 5 vol% formic acid and 5 vol% acetic acid, and the presence of bridge | crosslinking was confirmed from the solubility change.
[0050]
(7) Adsorption experiment of acid dye on the complex of salt-shrinked silk protein and chitosan
Weigh each 0.01g of untreated silk, salt-shrunk silk and salt-shrunk silk (silk protein) and chitosan complex, 0.1 mol · dm, pH 4^-3In an acetic acid buffer solution in 200 ml of an acid dye (OR) dissolved therein and stirred at 40 ° C. for 72 hours. The dye solution after adsorption was subjected to colorimetry at λ = 485 nm using an ultraviolet / visible spectrophotometer (Hitachi, Ltd. Model 1100) to determine the free dye concentration. In addition, the dye was completely extracted from the dyed silk with formamide three times at 60 ° C. for 30 minutes, and the amount of the dye adsorbed by the silk was calculated in the same manner by comparing the volume of the solution in 50 ml.
[0051]
(8) Salinity rate
When the wild silk silk is immersed in a heated concentrated neutral salt solution, the wild silk silk is arbitrarily contracted in the fiber length direction. The salinity percentage (%) was calculated using the following equation.
Salinity rate (%) = [(Li-Lf) / Li] x 100
In the above equation, Li represents the sample length before shrinkage, and Lf represents the sample length after shrinkage.
(9) Mechanical properties
As mechanical properties, the elongation (%) of the yield point of the sample and the strength and elongation (%) of the sample at the time of cutting were measured. The measurement conditions were a sample length of 50 mm, a tensile speed of 10 mm / min, and a chart scale of 250 g, and the measurement was performed using an Shimadzu Instron (Autograph AGS-5D).
[0052]
(10) Alkali solubility (Asol)
The sample was immersed in an aqueous 0.1M sodium hydroxide solution at 65 ° C. for 1 hour, and the alkali solubility was evaluated from the change in weight before and after the treatment.
Asol = (Mb-Ma) / Mb × 100
Mb: Sample weight before alkali treatment (dried at 105 ° C for 1 hour 30 minutes)
Ma: Sample weight after alkali treatment (dried at 105 ° C for 1 hour 30 minutes)
[0053]
(11) Metal ion adsorption and quantification method
Metal ions were adsorbed by immersing the sample to be measured in a 0.5 mM silver nitrate aqueous solution or potassium nitrate aqueous solution containing potassium nitrate (pH adjusted to 11.4 by adding aqueous ammonia) for 30 hours at room temperature.
The metal ions adsorbed on the sample to be measured were analyzed using a plasma atomic absorption spectrometer (ICP-AES) manufactured by PerkinElmer. A 5-10 mg sample was completely hydrolyzed with 2 mL of 65% nitric acid using a microwave hydrolysis furnace (MDS-81DCCEM), an additional 10 mL of water was added during the experiment, and ICP-AES analysis was performed. The amount of metal ions adsorbed was expressed in mmol as the amount of metal ions per sample weight.
[0054]
Examples clarified in the above experiment are shown below.
(Example 1) Salt shrinkage treatment of silk thread
Immerse silkworm silk thread (bath ratio: calcium nitrate aqueous solution volume / silk weight = 100) into a 50wt% calcium nitrate tetrahydrate aqueous solution at 60 ° C for 5min to shrink the silk yarn in the length direction and further salt After shrinking, it was washed 3 times with distilled water, air-dried and vacuum-dried to obtain a salt-crimped silk thread. This salinity rate was 14%.
When soaking salt-shrunk silk in an aqueous acetic acid solution of chitosan, the salt-shrunk silk obtained as described above was washed with distilled water and then immersed in distilled water without drying. It was.
[0055]
(Example 2) Immersion treatment in aqueous chitosan solution
Rabbit silk is salt-shrunk according to Example 1 and immersed in distilled water. The swollen salt-shrunk silk is placed in a separable flask, and a solution obtained by dissolving 10 g of chitosan in 500 ml of 5 vol% acetic acid aqueous solution is added thereto. In addition, a chitosan-containing silk thread was produced by immersion treatment at 30 ° C. for 24 hours.
[0056]
(Example 3) Introduction of cross-linking to chitosan-containing silk thread
Cross-linking was introduced into the chitosan-containing silk thread produced in Example 2 as follows. 0.7 g of chitosan-containing silk thread was put into 250 ml of water, 0.44 g of EGDGE was added, and the crosslinking reaction was advanced by gently stirring at 70 ° C. for 3 hours. After completion of the reaction, the mixture was filtered, washed several times with diethyl ether and acetone, then air dried and vacuum dried. Thus, even if the chitosan-containing silk thread into which the cross-linked bond was introduced was immersed in acetic acid and allowed to stand for a long time, the weight did not decrease and the chitosan did not detach from the silk thread.
[0057]
(Example 4) Quantification of chitosan introduction amount into silk thread
In the treatment of immersing the salt-crimped silk thread obtained in Example 1 (salt shrinkage ratio 14%) in an acetic acid aqueous solution of chitosan, the amount of chitosan introduced quantitatively was evaluated by performing a ninhydrin test. . That is, the ninhydrin test was performed on both the salt-reduced silk yarn and the salt-reduced / chitosan-treated silk yarn. At this time, it is the basic group and terminal amino group of the basic amino acid that constitutes the silk thread in the case of the salt-shrinkage treated silk, which is involved in the reaction with ninhydrin. In addition to these groups, it is an unreacted amino group in the introduced chitosan. In addition, 1 equivalent of basic group in the substrate reacts with 1 equivalent (1 mol) of ninhydrin, so as a standard substance for creating a calibration curve, the hydrochloride of D-glucosamine, which is a constituent unit (repeat) of chitosan, is used. I chose.
[0058]
The relationship (calibration curve) between the glucosamine concentration and the absorbance obtained by the measurement is shown in FIG. The regression line equation of this calibration curve is shown by the following equation.
A = 0.1527C (Formula 1)
In the above formula, A is absorbance (no unit), C is glucosamine concentration (mol^-3).
Next, for the salt-shrinkage silk and the complex of silk protein and chitosan, the absorbance obtained by the ninhydrin reaction was substituted into Equation 1, and the glucosamine concentration (mol · dm^-3) And the amino group content was calculated. Table 3 shows the amino group content of each obtained silk thread sample.
[0059]
(Table 3)

In addition, the amount of chitosan introduced in Table 3 corresponds to the amount of amino groups newly added to the silk thread by salt shrinkage / chitosan treatment.
As can be seen from Table 3, the amount of chitosan introduced into the salted silk thread is 0.44 × 10^-Fivemol ・ g^-1Met. In order to further increase the introduction amount of chitosan, a solution in which chitosan is dissolved to the limit of the dissolution amount may be used.
In place of the salt crimped silk yarn (salt shrinkage rate 14%) used above, the salt crimped silk yarn having an increased degree of salt crimp rate was treated with an aqueous acetic acid solution of chitosan in the same manner as described above, and a ninhydrin test was performed. The amount of chitosan introduced was evaluated. As the salinity rate increased, the amount of chitosan introduced increased. However, since the mechanical properties of the salt-shrunk silk yarn deteriorate as the salt-shrinkage rate increases, it is necessary to control the salt-shrinkage rate to 30% or less in order to achieve the desired result.
[0060]
(Example 5)
Untreated silk (A), salt shrinkage treatment (salt shrinkage rate 15%), chitosan-treated silk yarn (B), salt shrinkage treatment (salt shrinkage rate 15%) The mechanical properties were examined by stretching C) until it was cut and measuring the strength and elongation at the time of cutting. The results obtained are shown in Table 4.
[0061]
(Table 4)

Note: Strength 430 ± 18 means strength is 430gf and standard deviation is 18.
As is clear from Table 4, when the salt shrinkage rate is about 15%, the strength and elongation of the silk thread (B) after the salt shrinkage treatment are not significantly changed compared to the case of the untreated silk thread (A). . However, when the salt shrinkage rate exceeds 30%, the strength and elongation of the salt shrinkage-treated silk yarn are also rapidly decreased. In addition, the mechanical properties of the silk thread (C) into which the cross-linked bond is introduced retain the original characteristics of the silk thread, and there is no deterioration of the mechanical characteristics during processing.
[0062]
(Example 6) Dyeing with acid dye
The dyeing behavior of a complex of silk protein and chitosan (immobilized chitosan by introducing a cross-linking bond, hereinafter referred to as immobilized silk thread) was examined as follows.
A complex of silk protein and chitosan was stained with C.I. Acid Orange 7 (Orange II: hereinafter abbreviated as OR), which is a typical acid dye. That is, 0.1 mol · dm of acidic dye with pH 4^-3A test sample was placed in 200 mL of a staining solution dissolved in an acetic acid buffer solution and stirred at 40 ° C. for 72 hours to adsorb the dye to the sample. Using a UV / visible spectrophotometer (1100 model, manufactured by Hitachi, Ltd.), the dyeing bath after adsorbing the dye was colorimetrically measured at λ = 485 nm and the free dye concentration (× 10^-Fivemol ・ dm^-3) Further, the adsorption amount was determined as follows. From the dyed silk, the dye was completely extracted with formamide at 60 ° C for 30 min three times, and the solution was adjusted to a volume of 50 ml.^-Fivemol ・ g^-3) Was calculated. In addition, the dye adsorption measurement of the untreated silk yarn and the silk yarn treated only with salt crimp (hereinafter referred to as salt crimped silk yarn) was also performed. The obtained adsorption isotherm (pH = 4, 40 ° C.) is shown in FIG.
[0063]
As can be seen from FIG. 2, there was little difference in the amount of OR adsorbed at low concentrations, but at a relatively high dye concentration, a complex of silk protein and chitosan (immobilization treatment in FIG. 2). The amount of adsorbed to salt-silk silk (salt-shrunk treatment in FIG. 2) and untreated silk yarn (untreated in FIG. 2) is greatly exceeded, and the effect of chitosan immobilization is obvious.
[0064]
Fig. 3 shows the assumed OR adsorption mechanism on chitosan-immobilized silk. As shown in FIG. 3, for example, OR is directly ion-bonded to the amino group of the silk molecule, OR is bonded to chitosan bonded to the silk molecule via EGDGE, and also enters the silk thread. When chitosan is covalently bonded to each other via EGDGE, the phenomenon that the amount of dye adsorbed to the complex of silk protein and chitosan of the present invention increases can be explained without contradiction. .
In addition, Table 5 shows the number of dye binding sites of each silk thread calculated by Scatchard plot based on the data of FIG. From this result, the complex of silk protein and chitosan (immobilized silk thread) has 1.4 times the binding seat of the untreated silk thread.
[0065]
(Table 5)

[0066]
(Example 7) Confirmation of crosslinking of produced polymer
FT-IR spectra of the bridged chitosan No. 2 shown in Table 1 and chitosan FM80 (degree of deacetylation 85%) manufactured by Koyo Chemical Co., Ltd. were measured, and the difference spectrum thereof is shown in FIG. 2959cm in this difference spectrum^-1And 1472cm^-1A new peak can be seen. This was caused by an increase in C—C bonds with the introduction of EGDGE. Also 1600cm^-1Nearby amino group (-NH₂) NH's peak in bending vibration is reduced. Similar peaks were also detected for crosslinked chitosan No. 1 and No. 3 with different charge ratios of cross-linking agents. From the above, the reaction between chitosan and EGDGE could be confirmed.
[0067]
Moreover, about the bridge | crosslinking chitosan No. 1-3, when the dissolution test by 5% acetic acid and 5% formic acid was conducted, since all were insoluble, presence of bridge | crosslinking was recognized.
From the above, it was confirmed that chitosan was bridged. Since cross-linking was introduced into chitosan and chitosan, it can be determined that cross-linking is also introduced between the silk and chitosan molecules. Since the dyeing behavior of reactive dyes on chitosan and silk is similar, the reactivity of basic groups in silk to EGDGE is considered to be similar to that of amino groups in chitosan.
[0068]
Moreover, regarding chitosan-immobilized silk thread, it can be indirectly proved from the following that chitosan is immobilized in the silk thread. That is, the staining shown in Example 6 is 0.1 mol dm at pH 4.^-3In the acetate buffer solution, it is carried out at 40 ° C for 72 hours, so if chitosan is not immobilized, chitosan will be eluted from silk under these conditions Because it is.
[0069]
(Example 8) Alkali solubility
The alkali solubility of the untreated silk and the complex of silk protein and chitosan into which a cross-linked bond was introduced was evaluated.
The alkali solubility of the silk thread alone was 42%, whereas the alkali solubility of the complex of silk protein and chitosan into which a crosslink was introduced was 30%. From this, it can be seen that the disadvantage of being easily dissolved in an alkaline aqueous solution in the case of an untreated silk thread can be solved by introducing a cross-linking bond into a composite with chitosan.
[0070]
(Example 9) Adsorption of metal ions
As an aqueous metal salt solution prepared according to the above metal ion adsorption method, 0.5 mM silver nitrate (AgNO_Three) Aqueous solution and copper nitrate (Cu (NO_Three)₂) Aqueous solution was used. Samples of untreated rabbit silk yarn, salt-crimped silkworm silk yarn, salt-fried silkworm-chitosan-treated silkworm silk yarn, and salt-fried silkworm silk yarn insolubilized after introduction of cross-linking after salt-fried silktochitosan treatment were added to each aqueous solution. Was immersed for 30 minutes at 25 ° C. to adsorb silver ions and copper ions, and the amount of metal ions adsorbed on each sample was measured. The results obtained are shown in Table 6.
[0071]
(Table 6)

As can be seen from Table 6, in the case of untreated rabbit silk thread, when immersed in an aqueous metal ion solution, Ag⁺, Cu²⁺Both adsorb slightly, but the amount of adsorbed metal ions increases slightly by salt condensation treatment. It was confirmed that when the salt-condensed silk yarn was impregnated with chitosan, the adsorption rate of metal ions was further increased, and the chitosan-impregnated sample insolubilized with an epoxy compound adsorbed more metal ions.
[0072]
【The invention's effect】
According to the complex of silk protein and chitosan of the present invention, in addition to the cross-linking bond between chitosans, a cross-linking bond is introduced between chitosan and silk protein. It is possible to impart functions such as excellent dyeability and chemical resistance.
In the composite of the present invention, since silk protein, which is a general-purpose fiber for clothing pretreated by salt shrinkage treatment in a neutral salt aqueous solution, can be used as the silk protein, in addition to the excellent mechanical properties of silk yarn, various A chitosan-containing silk thread having the characteristics of chitosan with excellent biochemical characteristics can be provided.
[0073]
According to the composite of the present invention, since the introduction of the cross-linking is preferably performed using a cross-linking agent composed of a bifunctional, trifunctional, or polyfunctional epoxy compound, Crosslinks are introduced smoothly without the use of a catalyst.
Further, according to the method for producing a composite of the present invention, the silk yarn, which is a general-purpose fiber for clothing, is preferably subjected to a salt crimping treatment as a pretreatment, and the salt crimped silk yarn is impregnated with an aqueous chitosan solution, preferably an acetic acid aqueous solution in which chitosan is dissolved. Thus, a chitosan-containing silk thread is produced, and an epoxy compound is allowed to act on the chitosan-containing silk thread as a cross-linking agent to introduce a cross-linking bond between the silk protein and chitosan and between the chitosans filled in the silk thread. Thus, the chitosan contained in the silk thread is not detached, and the cross-linking bond is not released even if the complex of silk protein and chitosan that can be prepared is immersed in water for a long time. As a result, it has become possible to provide a composite material that combines the characteristics of silk protein alone and chitosan alone.
[Brief description of the drawings]
FIG. 1 is a graph showing the relationship (calibration curve) between glucosamine concentration and absorbance.
FIG. 2 is a graph showing an adsorption isotherm.
FIG. 3 is a schematic diagram of an OR adsorption mechanism on chitosan-immobilized silk thread.
FIG. 4 is a graph showing a difference spectrum between FT-IR spectra of bridged chitosan No. 2 and chitosan FM80.

Claims (5)

A complex of silk and chitosan salt-treated in a neutral salt aqueous solution , wherein the chitosan is bonded to the silk thread via a crosslinking agent , and chitosans that have entered the silk thread are bonded to each other via the crosslinking agent. Covalently bonded and the cross-linking agent is selected from glycerol polyglycidyl ether, resorcinol diglycidyl ether, neopentyl glycol diglycidyl ether, 1,6 -hexanediol diglycidyl ether, ethylene glycol diglycidyl ether, and polyethylene glycol diglycidyl ether A selected bifunctional epoxy compound or trifunctional epoxy compound, or a polyfunctional epoxy compound selected from sorbitol polyglycidyl ether, sorbitan polyglycidyl ether, and pentaerythritol polyglycidyl ether Complex characterized in that. Insect-derived silk is subjected to salt shrinkage treatment in an aqueous solution of a neutral salt selected from lithium bromide, lithium nitrate, lithium chloride, lithium thiocyanate, calcium nitrate, and calcium chloride. Without drying, the salt-shrunk silk thread was immersed in an aqueous acetic acid solution containing chitosan to produce a chitosan-containing silk thread. glycidyl ether, 1,6 - hexanediol diglycidyl ether, ethylene glycol diglycidyl ether, and bifunctional epoxy compound selected from polyethylene glycol diglycidyl ether or trifunctional epoxy compound or sorbitol polyglycidyl ether, , A polyfunctional epoxy compound selected from sorbitan polyglycidyl ether and pentaerythritol polyglycidyl ether is allowed to act at 40 to 80 ° C. in the presence of water or an organic solvent so that the chitosan becomes a cross-linking agent. A composite of the silk thread and chitosan is obtained by introducing a cross-linking bond so that chitosans entering the silk thread are covalently bonded via a cross-linking agent. Body manufacturing method . Claims immersion in the chitosan-containing aqueous solution of the Shiochijimi treated silk is present in 5 to 15% strength aqueous acetic acid, at 10 to 50 ° C., characterized by being conducted by immersing for 1 to 24 hours 2. A method for producing the composite according to 2 . The method for producing a composite according to claim 2 or 3, wherein the introduction of a cross-linking bond into the chitosan-containing silk yarn is performed in water or a tetrachloroethane solvent containing a cross-linking agent at a temperature of 40 to 90 ° C. . The amount of the cross-linking agent added is 5 to 100% of the total number of moles of total basic groups contained in chitosan and silk thread, The composite according to any one of claims 2 to 4 Method.

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