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The apparent average molar masses (M w,app ), apparent average radii of gyration (R g,app ), diffusion co-efficients (D T ), and hydrodynamic radii (R h ) of normal corn (maize) starch and fractions were determined using asymmetrical flow field-flow fractionation coupled with multiangle light scattering and refractive index detectors (AF4/MALS/RI). AM-type (Fraction A) and AP-type (Fraction B) were chemically separated from normal corn starch. Normal corn starch and Fractions (A-B) were dissolved in 1 M KSCN using a high pressure microwave vessel. The effect of varying cross flow rates at a fixed channel flow rate upon the M w,app and R g,app distributions of normal corn starch and Fractions (A-B) were investigated. The average M w,app of normal corn starch, Fractions (A) and Fraction (B) were 41 Â 10 6 , 1.4 Â 10 6 and 39 Â 10 6 g/mol, respectively, with R g,app values of 129, 60 and 129 nm, respectively.

The apparent average molar masses (Mw,app), apparent average radii of gyration (Rg,app), of native sago starch and fractions were determined using asymmetrical flow field‐flow fractionation coupled with multi‐angle light scattering and refractive index detectors (AF4/MALS/RI). Amylose‐type (Fraction A) and amylopectin‐type (Fraction B) were chemically separated from native sago starch. Native sago starch and Fractions (A–B) were dissolved in 1M KSCN using a high pressure microwave vessel. The effect of varying cross flow rates at a fixed channel flow rate upon the Mw,app and Rg,app distributions of native sago starch and Fractions (A–B) were investigated. The average Mw,app values of native sago starch, Fraction (A) and Fraction (B) were 60 × 106, 1.5 × 106 and 60 × 106 g/mol, respectively, with average Rg,app values of 142, 75 and 127 nm, respectively. The sphere‐equivalent hydrodynamic radii (Rh) values for native sago starch and fractions were determined from AF4 experimental parameters.

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The enzyme levanase encoded by the sacC gene from Bacillus subtilis was overexpressed in Escherichia coli with the strong, inducible tac promoter. The enzyme was purified from crude E. coli cell lysates by salting out with ammonium sulfate and chromatography on DEAE-Sepharose CL-6B, S-Sepharose, and MonoQ-Sepharose. The purified protein had an apparent molecular mass of 75,000 Da in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which is in agreement with that expected from the nucleotide sequence. Levanase was active on levan, inulin, and sucrose with K m values of 1.2 M, 6.8 mM, and 65 mM, respectively. The pH optimum of the enzyme acting on inulin was 5.5, and the temperature optimum was 55؇C. Levanase was rapidly inactivated at 60؇C, but activity could be retained for longer times by adding fructose or glycerol. The enzyme activity was completely inactivated by Ag ؉ and Hg 2؉ ions, indicating that a sulfhydryl group is involved. A ratio of sucrase to inulinase activity of 1.2 was found for the purified enzyme with substrate concentrations of 50 mg/ml. The mechanism of enzyme action was investigated. No liberation of fructo-oligomers from inulin and levan could be observed by thin-layer chromatography and size exclusion chromatography-low-angle laser light scattering-interferometric differential refractive index techniques. This indicates that levanase is an exoenzyme acting by the single-chain mode. Enzymes involved in the hydrolysis of polyfructans are of interest both for fundamental studies and for industrial applications. Especially inulin is of growing interest as a renewable carbohydrate raw material for biotechnology. Two aspects are of main importance: (i) production of pure fructose syrups, so-called high-fructose inulin syrups, by enzymatic hydrolysis of inulin (43) and (ii) direct fermentation of inulin by employing inulinase-producing microbes in order to synthesize various products such as ethanol or aceton-butanol (20, 21). ␤-D-Fructofuranosidases are usually classified upon their ability to hydrolyze levan (levanases), inulin (inulinases), and also the disaccharide sucrose (sucrases and invertases). However, many of these enzymes are capable of hydrolyzing more than one type of these substrates. Inulinases (or inulases) which are specific for inulin have been isolated only from Jerusalem artichoke tubers (7, 12), whereas levanases which are specific for levan have been isolated from bacteria only. Examples are the levanases of Streptococcus salivarius KTA-19 (38) and Actinomyces viscosus ATCC 19246 (16). Conversely, a variety of nonspecific ␤-D-fructofuranosidases have been found in bacteria, yeasts, and fungi. For example, inulinases and levanases which are capable of hydrolyzing inulin, levan, and sucrose have been isolated from Bacillus subtilis (17), Actinomyces viscosus ATCC 15987 (24), Streptococcus mutans (5), Kluyveromyces fragilis (35), Chrysosporium pannorum (46, 47) and Penicillium sp. strain (27). Enzymes active on inulin and sucrose but not on levan have been found in f...