Koji Extract Research Articles

米麹中の糖化力の新規測定法

A new method for measuring glucose-forming activity in rice koji is proposed. In this method, G 2-PNP (4-nitrophenyl O-α-D glucopyranosyl-(1-4)-β-D- glucopyranoside) is used as the substrate. Because both glucoamylase and a-glucosidase can hydrolyze the substrate, the results showed the sum total of these enzymes activities (that is, glucose-forming activity) in rice koji extract. The substrateis hydrolyzed by these enzymes and the resulted product (PNP) is measured to determine the amount of glucose-forming activity quantitatively. This method has the following advantages:(i) The presence of glucose in the sample does not influence the results of the measurement, and therefore there is no need to dialyze the rice koji extract prior to analysis.(ii)α-Amylase of Aspergillus oryzae in the sample does not affect the results of measurement because if cannot hydrolyze the substrate.(iii) The assay procedure is simple, and can be easily accomplished in a short time. The proposed method showed good correlation with the method generally used for glucoamylase assay when purified glucoamylase is used as a sample. The averages of recovery of glucoamylase and a-glucosidase added were 99.6% and 101.1%, respectively. Coefficients of variation in within-day and day-to-day experiments were within 1% and 2.8.%, respectively.

秋田流・花酵母「AK-1」による清酒実地醸造試験

The sake yeast, Akita Ryu-Hana yeast (AK-1) with high ethyl caproate productivity and low acidproductivity, could be isolated by using koji extract medium supplemented with alcohol dehydratedkoji. Brewing tests for Gin jo-shu (specially made sake) were carried out by using this strain on pilot scale (total rice 100kg) and in 9 factories (total rice 690-2, 000kg) throughout Akita Prefecture. The brewed sake contained a large amount of ethyl caproate, a small amount of total acid, and had good quality. In moromi, the activities of esterase and alcohol acyltransferase of yeast were constant and higher than those of Kyokai Sake Yeast No.9. High ethyl-caproate productivity and non-foaming ability of this yeast were stable after 10 times of periodic transfer.

アルコール脱水麹を用いる培地による優良酵母の分離とその性状

Mutants of sake-yeast having high ability to produce isoamyl acetate and ethyl caproate could be isolated at high frequency by using koji extract medium supplemented with alcohol dehydrated koji. One yeast strain (No.1-9) having high ability to produce ethyl caproate was selected from 925 strains collected. In order to enhance the ability of fermentation, non-foaming mutant was obtained from strain No.1-9 and used for sake brewing. The brewed sake contained a large amount of ethyl caproate, a small amount of total acid, and had good quality.

Purification and some properties of acid-stable .ALPHA.-amylases from shochu koji (Aspergillus kawachii).

Aspergillus kawachii α-amylase [EC 3.2.1.1] 1 and II were purified from shochu koji extract by DEAE Bio-Gel A ion exchange chromatography, Sephacryl S-300 gel chromatography (pH 3.6), ω-amino dodecyl agarose column chromatography and Sephacryl S-200 gel chromatography. By gel chromatography on a Sephacryl S-300 column, the molecular weights of the purified α-amylase I and II were estimated to be 104, 000 and 66, 000, respectively. The isoelectric points of α-amylase I and II were 4.25 and 4.20, respectively. The optimal pH range of α-amylase I was 4.0 to 5.0, and the optimum pH of α-amylase 'II was 5.0. The optimum temperatures of both α-amylases were around 70°C at pH 5.0. Both α-amylases were stable from pH 2.5 to 6.0 and up to 55°C, retaining more than 90% of the original activities. Heavy metal ions such as Hg2+ and Pb2+ were potent inhibitors for both α-amylases.

Biochemical Studies on Oligo- and Polysaccharides

The topics from the results of the studies conducted by the author during the past three decades are reviewed. The studies are mainly related to oligo- and polysaccharides and include the following projects: 1. Glucobioses in Japanese rice wine (Sake) and Koji extract. 2. Chemical synthesis of kojibiose and nigerose. 3. Acetolysis of gluco-oligosaccharides and glucans. 4. Structural studies of Konjac glucomannan. 5. Biochemical studies of dextrans. 6. Biosynthesis of fungal glycogen. 7. Structural studies of cell wall polysaccharides. 8. Biochemical studies of plant cell wall polysaccharides. 9. Some polysaccharide degrading enzymes including an α-1, 3- glucanase of a Streptomyces species, a dextranase capable of splitting α-1, 2-linkage at the branch point, and a /3-glucosidase of Pyricularia rr yzae.

Effect of Rhizopus koji extract on the conception rate of infertile heifers and cows

Rhizopus麹抽出物(R.E.)を宮城県玉造郡岩出山地方および古川市郊外の農家において,卵巣疾患(嚢腫を除く)で不受胎の50頭の乳牛を用い,R. E.3~5gを30日間経口投与し,投与開始から90日間の受胎に及ぼす影響を調べて次の結果を得た。1. 試験期間中の人工授精により37頭の牛が受胎した(受胎率74%)。2.受胎に至る授精回数は,25頭のものが1回,11頭のものが2回,1頭が3回であった。3.1.の結果を年令,産次別に分けると,特に2~3才の未経産牛に対して好成績を得た。4.R. E.投与前長期に亘り無発情であった牛20頭では試験中に18頭のものが発情を回帰し,そのうち4頭が受胎した。

Biologically active impurities in reagent glucose

Two mutants, EA3-867 and N2-78, with high cellulase yields were obtained from wild strains of Trichoderma pseudokoningii Rifai, 1096 and Mo3, respectively, by mutagenic treatments with a linear accelerator, 60Co, u.v., nitrosoguanidine (NTG) and diethylsulphate (DTS). The mutants grew slowly to produce small colonies on agar plates with synthetic medium. On agar plates of peptone-yeast extract, the small colonies were as large as those of wild strains. The cellulase activities of these mutants in Koji extracts, shake flask culture filtrates, and enzyme preparations were markedly higher than those of their parents. The mutant N2-78 reached quite high cellulase activity level when cultured for 60 h in shake flasks in a simple medium containing milled straw, wheat bran, mineral salts plus waste glucose molasses. The cellulase saccharifying activities on CMC, filter paper and cotton, were 255, 8.2 and 13.4 mg glucose/ml enzyme, respectively, or 11, 4.3 and 6 times more than those of its parent Mo3.The cellulase synthesis of EA3-867 and N2-78 was strongly induced by sophorose, isolated from pods of Sophora japonica L., and was inhibited by glucose, sugar phosphates, glycerol and organic acids. We conclude that cellulase synthesis of the mutants is regulated by catabolite repression as well as by induction. The increase in cellulase production by both mutants results from changes in the regulatory systems for cellulase synthesis, i.e. the mutants showed higher sensitivity to inducer and lower susceptibility to catabolite repression than did the wild types.A cellulase preparation of Trichoderma pseudokoningii Rifai N2-78 induced by sophorose was fractionated by DEAE-Sephadex A-50 and Sephadex G-100 column chromatography, selective inactivation and polyacrylamide gel electrophoresis. The components C1(exo-β1,4-glucanase), Cx(endo-β1,4-glucanase) and β-glucosidase were separated, and their molecular weights were estimated to be 67 000, 62 000 and 42 000 respectively. The homogeneity of C1 was verified by polyacrylamide gel electrophoresis, immunoelectrophoresis and ultracentrifugal analysis. It is a glycoprotein and is rich in glycine, aspartic acid, threonine, serine and glutamic acid. The C1 showed a strong synergistic action with Cx in the degradation of cotton, Avicel and Walseth cellulose.A poly(A)-RNA, induced by sophorose in N2-78 mycelium, was isolated by oligo(dT)-cellulose affinity chromatography.

Kojibiose (2-O-α-D-Glucopyranosyl-D-Glucose): Isolation and Structure: Isolation from Hydrol

IT has previously been reported that sake and koji extract produced from rice by the application of Aspergillus oryzae contains two new disaccharides1, one of which has been confirmed to be 1 : 3-α-linked gluco-biose (kojibiose)3,4 by its conversion into glucose with acid, R F in paper chromatography, M G in ionophoresis in borate buffer, its resistance to almond β-glucosidase, lack of formation of osazone and the similarity of its stain (aniline hydrogen phthalate spray) to that of sophorose. Peat et al. 4 have also described the isolation of a gluco-biose (octaacetate; melting point 166°, [α]D + 153°), believed to contain the 1 : 2-α-linkage, from a mixture of disaccharides synthesized from glucose by Aspergillus niger. One of us (K. A.) has detected kojibiose in the acid hydrolysate (hydrol) of sweet potato starch which is known to have undergone marked reversion5. We now report the isolation of this sugar from hydrol.

麹汁及び清酒中の二糖類

Both α, α- and α, β-trehalose were isolated from Koji extract as their crystalline octaacetates, and the latter was identified by the mixed melting point with the synthetic specimen prepared by the method described by MICHEEL et al. Separation of these sugars from isomaltose was performed by removing of the latter as its water insoluble phenylosazone. Kojibiose also gives no phenylosazone and is presumed to be 1, 2-α linked disaccharide; neither the free sugar nor its octaacetate could be isolated in the crystalline form.

醤油酵母の研究

(1) The author studied on methods for determining viable counts of yeasts in the presence of molds such as Koji-molds. (2) As it was difficult to differentiate yeast cells from the spores of the molds under micros-cope, plate culture method was adopted. (3) By anaerobic plate culture (duplicate dishes method) growth of molds could be inhibited and growth of yeasts, especially pellicle forming ones, was also inhibited to some extent. Moreover, this method was shown to be inconvenient to count the exact number of yeasts owing to the cracking of the media by the production of gas. (4) Antifungal agents which do not inhibit the growth of yeasts were investigated. And sodium propionate was found to be comparatively effective for that purpose. Its antifungal activity increased with the lowering of the pH of the medium in a certain extent. The plate culture using Koji extract agar added with sodium propionate by 0.2% and pH 4.6 was revealed to be most suitable for determining the viable counts of soy yeasts.

清酒酵母の発育に及ぼすビタミンの影響に就て

The effect of vitamins was studied on the growth of sake yeasts of the Society of Brewing on the basis of Burkholder's medium; the results are as follows:(1) The amounts of inositol (10, 000 γ/l) and of nicotinic acid (200 γ/l) as given in the prescription were too large, and the most suitable amounts were 25-50 γ/1 for both.(1) The amounts of inositol (10, 000 γ/l) and of nicotinic acid (200 γ/l) as given in the prescription were too large, and the most suitable amounts were 25-50 γ/l for both.(3) However, this syntehtic medium was yet inferior to the natural koji extract as the medium of sake yeasts.

麹の研究 (IV)

Aspergilli for “Amasake” brewing were selected by practilal niannfacturing trials under controlled conditions. Suitable Aspergillif or “Sake” brewing were selected from these strains suitable for “Amasake” brewing considering the low production of coloring substances in the Koji extract. Two strains of Aspergilli available for the submerged culture were obtained after practical culture urials. somewere suitable for amylase. production, and the other were for protease production.

麹菌に關する研究

Conidia of a selected strain of Aspergillus sojae (No. 260) were exposed to ultraviolet radiation, and random isolations were made from the colonies resulting from irradiated cells. Approximately two hundred of these were cultivated upon Czapek's agar solution. The majority of the isolates remained stable with normal appearance and similarity to cultural mutants. Eleven types of mutations were recognized. One of these resulted from restricted colony. Another was unable to utilize nitrate nitrogen-but developed normally if supplied with Koji extracts. Other mutations showed a total or partial loss of color without apparent alterations in their physiological properties. These eleven types are as follow : 1. Normal, 2. Restricted, 3. Extremely, restricted 4. Albino, 5. Olive, 6. Yellow, 7. Brown, 8. Sterile, 9. Floccose, 10. Nitratophobe, and 11. Nitrate non assimilative.