セルロースミクロフィブリルの高次構造解析への電子回折法の導入

Abstract

セルロースの結晶構造およびセルロースミクロフィブリルの配向はこれまで詳しく研究されてきた。しかし両者の間を橋渡しするセルロースの高次構造については未だ不明の点が多い。そこで電子回折法をセルロースの高次構造解析に応用するために, 試料作製と回折法の両面から改善を加え, その有効性を検討した。まず, バロニア, ラミー, 仮道管壁のS_2層, ポプラのG層から, 細胞壁本来の構造を維持するために薄層を剥離した。そして電子回折につきまとう試料の電子線損傷を軽減するために, 無用な電子線照射を極力減らすことを工夫した。その結果, 多数の回折スポットを持つ鮮明な回折パターンを得ることに成功し, セルロースの高次構造解明の端緒を得ることができた。さらに酢酸ウラニル染色することによりミクロフィブリルの人為的移動を防止し, また間隙のコントラストを増大させ, その広角および小角領域の回折強度を解析することで, ミクロフィブリルの周期的分布などが評価できるようになった。

The cellulose microfibrils are generally recognized to influence both physical and chemical properties of plant cell walls, constituting their framework. Although the crystal structure of cellulose in molecular level and its angular distribution in macrolevel have been investigated, the intermediate higher structure between two dimensions still remains to be exploited. The electron diffraction method, which has been widely applied to the ultimately selected area to obtain crystallographic data, was modified to evaluate the higher order structure with some precautions in specimen preparations. In order to reduce unavoidable electron damages, the irradiation area was adjusted by a condenser lens to 2-5μmφ, and the diffraction patterns were recorded only from a fresh area without the focussing process. When this modified method was applied to thin layers stripped from Valonia, ramie, and S2 and G layer of some woods, the diffraction patterns with a large number of diffraction spots were obtained. The uranyl-acetate staining was introduced to maintain the original higher order structure of the microfibrils, as well as to enhance the contrast between intermicrofibrillar space and the microfibrils. In the former an artificial movements of the microfibrils were well prevented. Periodicity of microfibril distribution in their cross section was able to be precisely evaluated with small angle scattering pattern.