Isolation and characterization of a derepressed mutant of Schwanniomyces castellii for amylase production

Summary The production of -amylase and amyloglucosidase activity in the yeast Schwanniomyces castellii strain CBS 2863 is repressed in the presence of glucose. Mutants displaying increased amylase activity were obtained after treatment with UV light and screening for resistance to 2-deoxy-glucose. One mutant was found to exhibit derepressed amylase activity. Biosynthesis and the rate of excretion did not appear to be as highly sensitive to dissolved oxygen, pH and dilution rate as in the parental strain.     

Fermentation of wheat grain by Schwanniomyces castellii

Fermentation of flour and wheat grain, without enzymatic hydrolysis has been demonstrated using a strain of Schwanniomyces castellii. Ethanol production yields were fairly good although starch was not totally degraded. The composition of the fermentation residues was compared to that of distillers dried grains.     

Kinetics of potato starch fermentation by Schwanniomyces castellii

The growth behaviour of Schwanniomyces castellii in slurry fermentation systems using untreated potato starch as substrate was studied in order to asses the eventual effect of the initial concentration of substrate (S_o) on cell growth rate. By applying the elementary balance method in combination with a Monod-type kinetic equation it was possible to formulate not only an unstructured model, but also the stoichiometry for such a yeast fermentation process. From a kinetic viewpoint, the Monod model was found to be redundant with respect to the pseudo-first order one, it being impossible to discriminate the contribution of v _M and K _S on the overall fermentation kinetics. Whereas the main yield coefficients appeared to be independent of S _O, the pseudo-first order rate constant was found to be inversely proportional to S _O. Therefore, cell growth appears to be controlled by the initial amount of amylolytic enzymes, that is to some extent proportional to the inoculum size, instead of the initial concentration of potato starch, at least within the experimental range of 3 to 30 g dm³.     

Behaviour of Endomycopsis fibuligera glucoamylase towards raw starch

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.     

Glucose transport and catabolite repression in Endomycopsis fibuligera yeasts

The role of systems for glucose transport in the manifestation of carbon catabolite repression of glucoamylase synthesis was studied in the yeast Endomycopsis fibuligera. Experimentas were conducted with its mutant AB-192 defective in the system of transport universal for glucose and 2-deoxy-D-glucose (2-DG). The nature of the mutation was established from the following data: (1) transport of labeled glucose into the mutant cells was twice as low in comparison with the parent culture 20-9; (2) transport of labeled 2-DG was suppressed almost entirely; (3) no competition was found between glucose and 2-DG for penetration into the mutant cells. Glucoamylase synthesis in the mutant AB-192 was not sensitive to catabolite repression by glucose. This was confirmed by the resistance of the AB-192 cells to the inhibition by glucose and their complete resistance to the repression by 2-DG. Moreover, an addition of cAMP did not stimulate glucoamylase synthesis by the mutant culture in the presence of glucose and 2-DG. It can be concluded therefore that the resistance of the yeast to catabolite repression by the glucose is caused by the mutation in the system for carbohydrate transport. The results suggest that the system of glucose transport plays an important role in the manifestation of carbon catabolite repression in the yeast Endomycopsis fibuligera.     

Characterization of Schwanniomyces castellii mutants with increased productivity of amylases

Summary The production of -amylase activity in the yeast Schwanniomyces castellii strain 1402 is repressed in the presence of the non-metabolizable glucose analogue, 2-deoxy-glucose. Selection for resistance to 2-deoxy-glucose after treatment with ethyl methane sulphonate (EMS) or UV light has yielded mutants displaing increased -amylase activities. One such mutant, S. castellii strain 1436, was found to exhibit constitutive -amylase activity in glucose-containing medium. This constitutive enzyme activity was also observed under pilot scale fermentation conditions when the pH was maintained constant at 5.5±0.1. The disaccharide maltose served as a stronger inducer of -amylase activity than the natural substrate starch in both the wild type (1402) and mutant (1436) strains.     

Optimization of single-cell-protein production from cassava starch using Schwanniomyces castellii

Schwanniomyces castellii B5285 grew faster and produced greater biomass and higher protein yield than either S. alluvius ATCC 26074 or S. alluvius 81Y when these amylolytic yeasts were grown with 2% (w/v) cassava starch as sole C source. With 0.5% (w/v) glutamate as N source, S. castellii reached 7.12 g cell dry mass/l, with a protein yield of 6.4 g/100 g starch. The optimal agitation speed, aeration rate and pH for growth of this yeast in a fermenter were 400 rev/min, 1.67 vol./vol.min. and 5.0, respectively. Tween 80 at 0.1% increased cell dry mass to 8.90 g/l, cell yield to 44 g/100 g starch and protein yield to 7.4 g/100 g starch.The authors are with the Department of industrial Biotechnology, Faculty of Agro-Industry, Prince of Songkla University, Hat Yai 90110, Thailand     

Influence of culture conditions on the biosynthesis of Schwanniomyces castellii phytase

Summary The influence of several factors on the biosynthesis of Schwanniomyces costellii phytase was studied in continuous culture. The level of phytase production increased with pH and dilution rate. It decreased when the phytic acid or phosphate content increased.     

D-Arabitol Production by Endomycopsis chodati

Endomycopsis chodati in an aerated fermentation produced d-arabitol in yields of 35 to 40% of the sugar supplied. Glucose, mannose, and sucrose were suitable substrates. A synthetic medium was developed for the fermentation that showed that nitrogen in the medium must be limiting to obtain high yields of arabitol. Excess phosphate also tended to lower arabitol yields, although the effect was not so great as with nitrogen. Pilot plant-size fermentations were made in which all the nutrients were supplied by blackstrap molasses and urea. Arabitol yields in these fermentations were about 40% of the sugar supplied.     

Effect of calcium carbonate on conversion of starch to ethanol by Schwanniomyces castellii

Conversion of starch to ethanol without its prior hydrolysis is of special interest in the treatment of the starch containing waste-waters. In this work the effect of calcium carbonate on the growth and ethanol production from soluble starch by Schwanniomyces castellii NRC 2676 was studied. When the medium was supplemented with 0.1% calcium carbonate the biomass yield and the total starch consumption were increased by 20% and 22%, respectively, while the ethanol yield decreased for 28% in comparison with the control. The rate of biomass production, measured for the period of time between 0 and 24 h, was the highest in the medium with 0.06% calcium carbonate while the rates of ethanol production and starch consumption decreased with and increase in the concentration of that salt; the media with 0.5% and 0.7% calcium carbonate had 3.5 and 5.5 times lower ethanol production rates, and 2.3 and 3.8 times lower rates of starch consumption than the control medium which was not supplemented with calcium carbonate, respectively. The obtained results also showed that the percentage of starch consumed was higher in the media supplemented with calcium carbonate than in the control. Empirical mathematical expressions are given for the relationship of the biomass and ethanol yields to the concentration of calcium carbonate in the medium.     

Isolation and characterization of a mutant of Schwanniomyces castellii with altered respiration

We have tried to isolate respiratory deficient mutants of the amylolytic yeast Schwanniomyces castellii CBS 2863 after mutagenesis with acriflavine. One of the mutants called DR 12 has been studied in more detail. Pasteur effect present in the wild-type is lost in the mutant, on the contrast an obvious Crabtree effect was observed: fermentation was almost as active in aerobiosis as in anaerobiosis. Moreover, the rate of anaerobic fermentation of the mutant was almost twice that of the wild type. This mutant was cytrochrome b-deficient while the amount of the other cytochromes was larger than in the wild-type. Moreover, the level of these remaining cytochromes in the mutant was higher on non-repressive medium than on glucose medium. However, the fact that the mutant DR 12 retained a cyanide-sensitive respiration and that it was able to grow on ethanol as a non-fermentable substrate is noteworthy.     

Ethanol production from native cassava starch by a mixed culture of Endomycopsis fibuligera and Zymomonas mobilis

The conversion of starch from unhydrolyzed cassava flour to ethanol by a pure culture of Endomycopsis fibuligera and by a co-culture of this amylolytic yeast and the bacterium Zymomonas mobilis was studied. The best overall results were obtained using the mixed culture. After 96 h of fermentation of a medium containing 150 g/l initial cassava starch, an ethanol concentration of 31.4 g/l, a productivity of 0.33 g ethanol/l × h and a yield of 0.21 g ethanol/g initial starch were reached. The highest yield (0.37 g/g) was obtained after 48 h when using a medium containing 50 g/l initial starch.     

Effect of medium composition on excretion and biosynthesis of the amylases of Schwanniomyces castellii

Summary The influence of different salts on the excretion of amylases has been studied sodium and potassium phosphates were added to the basal YNB soluble starch medium. The effects of sodium and potassium (from 0.005 M to 0.2 M) and the two surfactants, Tween 80 and Triton X were tested. The effect of the excretion of the amylolytic enzymes on the biomass yield and growth rate are discussed.     

Influence of culture conditions on the cell yield and amylases biosynthesis in continuous culture by Schwanniomyces castellii

Schwanniomyces castellii excreted -amylase and amyloglucosidase into the medium in the presence of starch. The biosynthesis and the rate of excretion were influenced by dissolved oxygen (specially for -amylase), pH of the culture and dilution rate. The cell yield observed (0.59) remained constant up to D=0.35h^-1 with starch as substrate. But in the case of growth on glucose, the yield observed was equal to 0.62 up to a dilution rate of D=0.18 h^-1. Beyond this value Y x/s decreased and ethanol was produced. The onset of fermentation dependend partly on the nature of the substrate and not only on the environment in particular on the quantity of dissolved oxygen present.