Abstract
In order to obtain more conclusive results on tissue degradation by pectolytic enzyme, the present investigation was conducted to confirm the properties of pectin-like polysaccharides liberated from cell wall-materials by the action of endo-PTE. Three hundred mg of radish fiber was incubated with the purified enzyme (2.0 unit) at 30C for 90 minutes in 0.033M tris-HCl buffer, pH8.0. The maceration mixture was fractionated into 6 subfractions of cell wall-materials according to the method of McColloch22).
In contrast to non-macerated radish fiber, dil. HCl-soluble carbohydrate subfraction (F-III) was remarkably decreased with a visible increase of water-soluble subfraction (F-I) during the enzymatic maceration. It is concluded, therefore, most of water-soluble carbohydrates in macerating mixture were mainly liberated from cell wall-materials extractable with dil. HCl solution, corresponding to so-called protopectin.
Both F-I and F-III subfractions were heterogeneous polysaccharide and separated further into 6 fractions by DEAE-cellulose column chromatography. When each fraction of F-I which originated from macerating mixture was hydrolyzed with 1N H2SO4 at 100C for 20 hours, neutral sugars such as galactose, arabinose, rhamnose and xylose were also detected on paper chromatograms, in addition to galacturonic acid. Major components of F-I subfraction, which eluted from the column with 0.2, 0.3 and 0.4M of acetate buffer (pH6.0), were obviously increased with the enzyme action as compared with these components of non-macerated control. On the other hand, 3 major components of F-III subfraction, which eluted with 0.3 and 0.4M acetate duffer anb 0.1N NaOH, were remarkably decreased during the enzymatic maceration. These visible changes of major components in F-I and F-III were mainly due to the fluctuations of uronide substances, and no visible change was observed in neutral carbohydrate contents. In F-I subfraction prepared from macerating mixture, both 0.2M and 0.3M fractions abounding with uronide components also showed relatively higher contents of unsaturated compounds and methoxyl groups. From the reasons mentioned above, it is supposed that both fractions were released from water-insoluble cell wall-polysaccharides by trans-eliminative reaction of the enzyme used.
