Deproteinized waste water from a potato starch factory was treated by a membrane-combined two-phase methane fermentation system for 100 days in the operating season. The system was composed of an acidogenic reactor, membrane modules and a methanogenic reactor. At the start, methanogenesis was seriously inhibited by hydrogen sulfide, which either derived from membrane filtrate fed into the methanogenic reactor or was produced as a result of sulfate reduction in the methanogenic reactor. To prevent this inhibition, the following treatments were carried out: The membrane filtrate was bubbled with desulfurized biogas generated in the acidogenic reactor to remove hydrogen sulfide and then diluted three times by the addition of tap water. In addition, the pH of the methanogenic reactor was maintained between 7.3 and 7.6 by sodium hydroxide. After those, the system could be operated satisfactorily and the following results were obtained: The system was operated at 5.2 kg/m3 day of TOC loading, BOD and SS concentration of effluent in the system was 160 mg/l and 100 mg/l, respectively. From the methanogenic reactor, about 7200 kcal/m3 biogas containing 84% methane was recovered, which amounted to 9 times the volume of waste water. From the acidogenic reactor, the biogas yield was 3 times as much as waste water and contained less than 10% methane. The conversion rate to sludge was 8% to TOC of influent in the acidogenic reactor and was 1.5% to that in the methanogenic reactor, so it was not necessary to remove excess sludge from each reactor during this operation period. In this operation, degradation of high molecular weight substances and acidogenesis occurred dominantly in the acidogenic reactor due to serious inhibition of methanogenesis by sulfide, and acidogenesis and methanogenesis occurred in the methanogenic reactor. Accordingly, it was concluded that the acidogenesis phase was almost completely separated from the methanogenesis phase in the anaerobic degradation process.

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Two varieties of potato (Solanum tuberosum L., varieties: Danshaku and Mayqueen) were harvested 5 times in total during the period February to July, 1988. The properties of starch granules prepared from the potato tubers were examined. The results obtained were as follows: 1) The average granular size of Danshaku and Mayqueen starches were in the range of 12.2 to 26.9 um and 15.0 to 38.7 um, respectively, and increased with increasing weights of tubers. The average granular size of starch from the same weight range of potato tubers tended to be higher in the later stages of develoument than in the earlier stage. 2) The amylose contents of Danshaku and Mayqueen starches determined by amperometric titration method were in the range of 20.7-23.0% and 19.8-23.4%, respectively, and the amylose contents of Mayqueen starches tended to increase in the early stages of tuber develoument (from May 25 to June 6). 3) The initiation temperature for gelatinization of Danshaku and Mayqueen starches determined by photo-pastegraphy were in the range of 64-67°C and 62-67°C, respectively, and it remained essentially unchanged throughout tuber develoument. 4) X-ray diffractography of the potato starches showed B-type patterns throughout tuber develoument. 5) The susceptibility of the potato starch granules to porcine pancreatin was higher in small granular size harvested in the early stage of tuber develoument than in large granular size. 6) By scanning electron microscopy (SEM), a stripe structure on the potato starch granules in the earlier stage was observed after attack by pancreatin.

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The static linear viscoelastic behavior (creep compliance) of starch gels at the earlier retrogradation stage has been investigated. Wheat starch, corn starch, and non-glutinous rice starch were used. Starch gels (starch content 15.2-35.0%) were stored at 2°C up to 36 hr. Creep compliance curves of wheat starch gels and corn starch gels could be represented by a Voigt-type four-element model, which was combined of a elastic element, a Voigt element, and a viscous element in series array. In case of rice starch gels, creep behavior was represented by a six-element mechanical model combining a Voigt-type four-element model and a Voigt element in series array. Changes in creep compliance (J(300)), which were measured under uniaxial compression for 300 s, of wheat starch gels with storage period showed four regions of gel-hardening: 1) in region I, from 0 to several hours, J(300) decreased exponentially with increasing storage period, 2) in region II, the compliance increased with increasing storage time for several hours, 3) in region III, the slope of J(300) versus the storage period curve continued to decrease exponentially from the initial point until an inflection point, and 4) in region IV, the compliance continued with decreasing slowly. The occurence of region II was first confirmed by the present study. The gel-hardening process of corn starch gels also separated into the four regions. On the other hand, the process of retrogradation of rice starch was monomorphic. The values of initial creep compliance, J0, of 35.0% starch gels without storage or with shorter storage period, increased in the following order: rice starch>corn starch>wheat starch. The values of the hardening rate constant in region I, k1, of wheat starch gel and corn starch gel were increased about 2 times and 7 times higher than that of rice starch gel, respectively.

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This study was conducted to investingate the molecular structures and the properties of starches prepared from six different kinds of rice (Oryza sativa L. japonica) cultivars produced in Japan. These six kinds were Yukihikari, Kirara397, Koshihikari, Nipponbare, Koganebare and Reiho. The results can be summarized as follows: 1) Analysis of RVA viscogram revealed that three kinds of cultivars, Yukihikari, Kirara397 and Koshihikari, reached peak viscosity in a shorter period of time and showed higher break-down values and lower set-back values compared to the other three kinds. The former tended to gelatinize more quickly and to be more resistant to retrogradation than the latter. 2) An examination of endothermal temperature (To, Tp, Tc) showed that Koshihikari and Nipponbare had the highest value among the six kinds tested. For all the cultivars, a negative correlation was found between the blue values of starches and the levels of To and Tp. A similar relationship was also found between the enthalpy levels of the cultivars and the blue values of amylopectins. 3) Through X-ray diffractometry experiments, starches of Yukihikari, Kirara397, Koshihikari and Nipponbare turned out to have the A-type pattern. Starches of the other two had a similar pattern, but they also exhibited some characteristics of the C-type pattern. 4) Analysis of iodine affinities of starch, amylose and amylopectin revealed that all the rice starches experimented, except for Koshihikari, contained 1617% of amyloses, whereas Koshihikari starch contained only 14.6%. 5) The d.p.n of amyloses from the six kinds of cultivars ranged from 630 to 980, and these amyloses were found to form some branches. The d.p. of larger amylose molecules varied in weight in a wider range than that of smaller molecules. Particularly, Koganebare amylose contained the largest d.p.w molecules, while its c.l. was short and the n.c. large. 6) An experiment of β-amylolysis demonstrated that Koganebare amylopectin had the highest β-amylolysis limit. The c.l. of Koganebare amylose stood at 14 by the rapid SMITH degradation method as opposed to 28 by isoamylolysis. This difference appears to suggest that this cultivar is likely to have a different structure from the others. 7) The DSC data lead us to conclude that the length and the amount of the linear region of amyloses and amylopectins may influence the gelatinization properties of starches.

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[in Japanese]

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Isomalto-dextranase [EC 3.2.1.94] from Arthrobacter globiformis catalyzes the hydrolysis of α-1, 6 linkages of dextran and isomalto-oligosaccharides to liberate isomaltose from the non-reducing ends of those sugars. A new method for the preparation of isomaltose was developed by using the isomaltodextranase and an acid-treated dextran. When dextran T2000 was used as a substrate, the maximal degree of hydrolysis by the isomalto-dextranase was less than 40%. For this reason, the enzyme reaction was thought to be hindered at the branch points (α-1, 2, α-1, 3 and α-1, 4-glucosidic linkages) of dextran. The branch points of dextran were presumed to be selectively hydrolyzed by an appropriate mild acid pretreatment, because theα-1, 6-glucosidic linkage constructing main chain of dextran is less acid-labile than other α-1, 2, α-1, 3, α-1, 4-glucosidic linkages. Therefore, dextran was incubated at 95°C for 4 hr in 0.2 N hydrochloric acid. When the dextran treated with 0.2 N hydrochloric acid was acted on by the enzyme, the maximal degree of hydrolysis went up to over 90%. Consequently, the branch points were considered to be hydrolyzed selectively. In order to enable the continuous preparation of isomaltose, the acid-treated dextran and isomalto-dextranase were confined in the DIAFLOTM cell with the ultrafiltration membrane. The products were taken out through the membrane of the apparatus. By this system, 6 g of isomaltose was obtained from 200 ml of 5% acid-treated dextran.

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An α-amylase was purified from germinating cotyledons of Phaseolus vulgaris L. cv Toramame by precipitation with ammonium sulfate, chromatographies on β-cyclodextrin Sepharose 4B and Bio-Gel P-200 columns. This enzyme migrated as a single band on polyacrylamide gel electrophoresis. The molecular weight was estimated to be about 46, 000 by SDS-disc gel electrophoresis and the optimum pH was 6.0. The rate parameters for the hydrolysis of a series of maltooligosaccharides were estimated by the Lineweaver-Burk plots. The ratios of the V values for malto-triose, -tetraose, -pentaose, -hexaose, -heptaose, and -octaose, and maltodextrin (d.p.=18) were estimated to be 5.9: 9.2:10:14: 36:108:100, and the Km values for these substrates were 143, 88, 77, 20, 5.5, 4.8, and 3.3 mM, respectively. Based on the these rate parameters and on the action mode of this enzyme toward each maltooligosaccharide, the Subsite structure was assumed to be made up of eight subsites. Those affinities (A1 s) in the active site of the enzyme were evaluated to be 1.0, 0.15, 4.7 and 0.58 (or 0.77) kcal/mol for Subsites 3, 4, 5 and 8, respectively, and the sum of the affinities of Subsites 1 and 2 was 2.2 kcal/mol, and that of Subsites 6 and 7 was -2.8 kcal/mol.

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The effects of an exocellular polysaccharide (EPS-B) produced by Lactobacillus helveticus TN-4 on the proliferative response of murine spleen cells stimulated by several mitogens were examined by the MTT method. As a result, EPS-B inhibited the proliferative responses of spleen cells stimulated by B-cell mitogens, however EPS-B showed little inhibitory effect on those of spleen cells stimulated by T-cell mitogens. These results were confirmed by measuring the number of viable cells by flow cytometric analysis. When spleen cells were cultured with several mitogens in the presence of EPS-B, the number of viable B cells decreased, but the number of viable T cells showed little decrease compared with those in the absence of EPS-B. These results suggest that EPS-B has an immunosuppressive activity on murine B cells.

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Reaction condition of trehalose formation was investigated when maltooligosyl trehalose synthase (MTSase) and maltooligosyl trehalose trehalohydrolase (MTHase) from Arthrobacter sp. Q36 and isoamylase were acted on starch. The optimum temperature was 35-40°C and the optimum pH was 5.6-6.4. The lower hydrolysis of substrate, the higher yield of trehalose. Among five kinds of starch (potato, sweet potato, corn, wheat, tapioca) as substrate, tapioca starch gave the highest yield. The maximum yield of trehalose was 85.3% when the reaction was done under the optimum conditions using liquefied tapioca starch of 0.3% hydrolysis as substrate.

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The effects of water-soluble α-glucosyl rutin (G-rutin) on some rheological properties of wheat flour dough, an interaction between gluten and starch in dough and on loaf volume were studied. The addition of G-rutin (200 ppm) increased the loaf volume significantly. Apparently, the addition of G-rutin decreased the development and stability times by farinograph test. The dough containing a higher amount (1000 ppm) of G-rutin increased the modulus of elasticity, viscosity coefficient and relaxation time to a greater extent than a lower amount (50 ppm) or the control. Combined additions of G-rutin and L-ascorbic acid (AsA, 50 ppm) apparently gave higher values in all parameters tested. Scanning electron microphotography of Boughs showed that G-rutin-added Boughs changed somewhat in viscous appearance compared with the controls. There were no changes in either the gelatinization temperature and the enthalpy of starch in dough. The SH content in dough treated with G-rutin was higher than that of the control. The size of gas cells of crumb baked with G-rutin and AsA decreased slightly. Furthermore, the addition of G-rutin alone and in combination with AsA improved the staleness of bread crumb during storage.

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The gene for cycloisomaltooligosaccharide glucanotransferase from Bacillus circulars T-3040 was cloned and expressed in the recombinant plasmid pCI429 in Escherichia coli. The enzyme gene consisted of a unique open reading frame of 2916 bp. Comparison of the DNA sequence data with the N-terminal, C-terminal and 12 internal amino acid sequences of the purified enzyme secreted from B. circulars T-3040 suggested the enzyme was translated from mRNA as a secretory precursor with a signal peptide of 38 amino acid residues. The deduced amino acid sequence of the mature enzyme contained 934 residues, resulting in a polypeptide with a molecular mass of 103, 103 Da. It had no common consensus region which is characteristic of the α-amylase family. The cycloisomaltooligosaccharide glucanotransferase activity, cyclization activity in transformant was about 3.0 mU/ml which was the same as that from B. circulars T-3040.

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