Seinosuke Ueda

Fungal glucoamylases and raw starch digestion

Abstract Mold glucoamylase which has a high debranching activity and is active in raw starch digestion, may be used successfully for the production of industrial alcohol from starch, without steaming, in order to save energy.

Behaviour of Endomycopsis fibuligera glucoamylase towards raw starch

Abstract An extracellular glucoamylase [exo-1,4-α- d -glucosidase, 1,4-α- d -glucan glucohydrolase, EC 3.2.1.3] of Endomycopsis fibuligera has been purified and some of its properties studied. It had a very high debranching activity (0.63). The enzyme was completely adsorbed onto raw starch at all the pH values tested (pH 2.0–7.6). Amylase inhibitor from Streptomyces sp. did not prevent the adsorption of glucoamylase onto raw starch although the enzyme did not digest raw starch in the presence of amylase inhibitor. Sodium borate (0.1 m ) eluted only 35% of the adsorbed enzyme from raw starch. The optimum pH for raw starch digestion was 4.5 whereas that of boiled soluble starch hydrolysis was 5.5. Waxy starches were more easily digested than non-waxy starches, and root starches were slowly digested by this enzyme.

Solubilization of a novel isoflavone glycoside-hydrolyzing β-glucosidase from Lactobacillus casei subsp. rhamnosus

A novel isoflavone glycoside-hydrolyzing β-glucosidase produced by Lactobacillus casei subsp. rhamnosus IFO 3425 was solubilized by ultrasonic disruption of the cells in the presence of 2-mercaptoethanol and sorbitol as stabilizer. The β-glucosidase from L. casei subsp. rhamnosus specifically hydrolyzed soybean isoflavone glycosides, namely, daidzin and genistin, converting them to daidzein and genistein, respectively. By contrast, a commercial preparation of almond emulsin β-glucosidase could not hydrolyze these soybean isoflavone glycosides. The undesirably bitter and astringent isoflavone glycosides in soybean were decomposed for the first time with this novel β-glucosidase, an enzyme which has hitherto been considered difficult to solubilize, produced by a lactic acid bacterium.

A Study on the Mechanism of DNA Excretion from P. aeruginosa KYU-1—Effect of Mitomycin C on Extracellular DNA Production—

The mechanism of excretion of DNA was investigated using strain KYU-1. This strain produced a considerable amount of DNA extracellularly (7.8mg/ml), and the amount of extracellular DNA per ml of the culture was scores of times more compared to that of intracellular DNA per ml of the culture. DNA synthesis was repressed by the addition of inhibitors (nalidixic acid, 5-fluoro uracil or mitomycin C), their inhibition rates were 35 to 54%. With the addition of 100 μg/ml of mitomycin C at the middle of the logarithmic growth phase, only 0.89 mg of extracellular DNA per ml of the culture was obtained, the inhibition rate was 95%. However, the amount of organic phosphate synthesized in the culture was almost the same no matter whether with or without an inhibitor.Furthermore, the excretion of DNA from the cell surface of strain KYU-1 was observed with on electron microscope and the DNA molecules excreted were found to be double-stranded.

Hydrolysis of isoflavones in soybean cooked syrup by Lactobacillus casei subsp. rhamnosus IFO 3425

Abstract A novel cheese-like fermented food, designated soybean cooked syrup cheese (SCSC), was produced by lactic acid fermentation of soybean cooked syrup (SCS) generated during production of natto (a Japanese fermented soybean preparation). SCS contains two characteristic compounds, daidzin (35.1 mg/100 g SCS) and genistin (40.5 mg/100 g SCS), which originate in soybean seeds and have an unpleasant taste. These two isoflavone glycosides were properly decomposed during fermentation by every lactic acid bacterium tested. In particular, Lactobacillus casei subsp. rhamnosus IFO 3425, which produces β-glucosidase, showed significant hydrolytic activity against the isoflavone glycosides in the SCS. Thus, a useless industrial byproduct was converted to a cheese-like fermented food by an economical and uncomplicated procedure.

Preparation of Koji from corn hulls for alcoholic fermentation without cooking

Abstract Corn hulls, the outer peel covering the corn grain, were used for preparation of koji , which was applied to alcoholic fermentation without cooking of raw strachy materials. Corn hull koji had lower saccharifying power, α-amylase, CMCase and xylanase than wheat bran koji , but higher protease and pectinase activities, Its alcoholic fermentations of cassava starch and sweet potato were also superior to those of wheat bran koji : corn hull koji gave 10.3% (v/v) of alcohol with 93% yield from 20 g of cassava starch, while wheat bran koji gave 9.4% (v/v) of alcohol with 90.4% yeild; and corn hull koji gave 9.1% (v/v) of alcohol with 92.6% yielded from 50 g of sweet potato, while wheatbran koji gave 8.1 (v/v) of alcohol with 88.6% yield.

The DNA sequence of γ-glutamyltranspeptidase gene of Bacillus subtilis (natto) plasmid pUH1

SummaryThe γ-glutamyltranspeptidase (γ-GTP) gene of Bacillus subtilis (natto) plasmid designated pUH1, which is responsible for polyglutamate production, has been cloned and the nucleotide sequence determined. The sequence contains a single open-reading frame stretching for 1260 bp with a relative molecular mass of 49356. Putative -35 and -10 sequences, TTCAAA and TATTAT, were observed as the consensus sequence for the promoter recognized by the σ43 RNA polymerase of B. subtilis, and the ribosome binding site, the sequence of which was AACGAG, was complementary to the binding sequence of B. subtilis 16S rRNA except for one base. The amino acid sequence of the gene with the segment of putative protein C403 of staphylococcal plasmid pE194 indicates homology, whereas that with Escherichia coli and mammalian γ-GTPs does not show any similarity at all.

Batchwise ethanol fermentation with shochu distillery waste.

Abstract A novel vacuum distillation procedure at low temperature (35–40°C) was applied to shochu distillation to produce a shochu with a mild aroma. The resulting kome (rice) shochu distillery waste contained a large amount of viable yeast (7 × 10 6 cells/ml), glucoamylase activity (70 U/ml), acid protease activity (720 U/ml), and neutral protease activity (420 U/ml). Secondary ethanol fermentation of the mash composed only of glucose as a carbon source and kome or mugi (barley) shochu distillery waste was properly effected. About 10% of ethanol (v/v) was produced in the fermented mash at 30°C within 3 d. In contrast, distillery waste discharged by conventional distillation at high temperature (55–60°C) could not be used for secondary ethanol fermentation at all. The filtrate of secondarily-fermented distillery waste, containing ethanol and possessing a fine aroma fortified with higher alcohols and volatile esters during ethanol fermentation, could be useful for the production of a mirin -like liquor for cooking, Akazake , a characteristic red-colored, sweet alcoholic beverage produced in Kumamoto prefecture only, and a bath additive containing ethanol, a fine aroma, and enzymes. The solid cake of the fermented mash could be used in the same way as conventional sake-kasu, sake -cake. Here, a novel system of utilizing shochu distillery waste, conventionally treated as wastewater, was established economically using a simple vacuum distillation procedure without a need of any complicated process.

Design of microbial processes and manufactures based on the specialities and traditions of a region: A Kumamoto case

Efforts have been made to produce two types of ancient and/or novel alcoholic beverages as follows. (i) Sprouting-rice wine, a legendary and mysterious alcoholic beverage that dates back to antiquity in Japan, was successfully reproduced using sprouting rice as a saccharifying agent. (ii) Red-rice wine, a novel alcoholic beverage, was made from red-rice by ethanol fermentation without cooking. Attempts were also made to utilize by-products of the brewing process and from food industries as starting materials for new products. (i) Shochu distillery waste was successfully used for production of various alcoholic beverages with red-rice bran and chrysanthemum flowers (Higo-giku) and for semicontinuous ethanol fermentation. (ii) Soybean cooked-syrup, discharged from a natto factory, was successfully used for production of a cheese-like food and a yogurt-like drink. Characteristic alcoholic beverages and foods closely related to the brewing and food industries in Kumamoto Prefecture were produced from an example of cooperation of bioindustry, public research institution and educational institution. Attempts were also made to utilize industrial by-products and waste as materials for new products with the intent of developing regional biorecycling systems for the brewing and food industries.

Production and characteristics of inhibitory factor of raw starch digestion from Aspergillus niger

SummaryThe glucoamylase preparation of Aspergillus niger 19 inhibited the raw starch digestion by it at high enzyme concentration. The inhibitory factor (IF) was isolated from the glucoamylase preparation by heat treatment and purified by DEAE-Sephadex A-25 column chromatography, an initial Sephadex G-50 gel filtration followed by SP-Sephadex C-25 column chromatography (twice) and then second Sephadex G-50 gel filtration. The IF thus purified was homogenous in polyacrylamide gel electrophories. The inhibitory activity of IF increased with the increasing IF concentration but decreased with an increasing quantity of raw starch or enzyme concentration. The IF had no effect on the hydrolysis of boiled soluble starch. It was completely adsorbed onto raw starch. The IF had a molecular weight of about 10,500. It was abundant in hydroxy amino acids such as threonine and serine. Xylose, mannose, glucose, galactose, and galacturonic acid were present in it.