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     

Glucose metabolism in the yeast Schwanniomyces castellii: role of phosphorylation site I and an alternative respiratory pathway.

Glucose metabolism in a Crabtree-negative yeast, Schwanniomyces castellii, and a cytochrome b-deficient mutant of this strain was investigated in chemostat culture. The wild-type and mutant strains exhibited the same behavior. Oxidative metabolism was observed when the substrate uptake rate (qS) was low. Fermentative metabolites were excreted when the qS value was higher than 0.40 g.g-1.h-1, indicating the occurrence of a respirofermentative metabolism; however, the respiratory quotient (RQ) remained near 1. When fermentation occurred, the cytochrome pathway was repressed but not the salicylhydroxamic acid (SHAM)-sensitive pathway. The presence of an alternative SHAM-sensitive respiratory pathway and the presence of phosphorylation site I in all metabolic conditions explained the RQ value of 1 and accounted for high biomass yields in oxidative metabolism conditions (0.62 g.g-1 for the wild-type strain and 0.31 g.g-1 for the cytochrome b-deficient mutant 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.     

New Blastomycetes isolated from soils of Spain I: Schwanniomyces castellii nov. spec.

Summary A new species of Schwanniomyces is described; it was isolated from a soil taken in Spain. This species is named Schwanniomyces castellii nov. spec. in honour of prof. Tommaso Castelli (director of Istituto di Microbiologia Agraria e Tecnica della Universita di Perugia).     

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.     

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.     

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.     

Production of amylase(s) by Schwanniomyces castellii and Endomycopsis fibuligera

Schwanniomyces castellii and Endomycopsis fibuligera Produced extracellular amylase(s) when grown on various carbon sources and at different pH values. Both yeast species showed significant amylase synthesis in the presence of either maltose or soluble starch. On the other substrates tested (glucose, cellobiose, sucrose, trehalose, melezitose, raffinose, ethanol, glycerol) differences were found regarding growth and amylase production. Free glucose in the culture medium apparently inhibited enzyme synthesis. The pH range allowing maximal growth and amylase production was 4.5–6.0 for E. fibuligera and 5.5–7.0 for S. castellii.     

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.     

Effects of oligomycins on adenosine triphosphatase activity of mitochondria isolated from the yeasts Saccharomyces cerevisiae and Schwanniomyces castellii

Functional mitochondria with respiratory control were isolated from the yeasts Saccharomyces cerevisiae and Schwanniomyces castellii. The presence of site I in Schw. castellii was indicated by higher ADP/O ratio than in S. cerevisiae where this site is absent. The ATPase Vmax was higher in S. cerevisiae than in Schw. castellii mitochondria. The latter was increased by the DR12 nuclear mutation. Nevertheless, the stimulation by heat and the inhibition profile of oligomycins on mitochondrial F1-F0 ATPase activities were similar in all three tested strains. In S. cerevisiae and Schw. castelli wild type or mutant mitochondria, the well-known inhibition of F1-F0 ATPase activity by low concentrations of oligomycins is abolished at high inhibitor concentrations near 60microg/ml suggesting uncoupling of F1 activity. At still higher oligomycin concentration the ATPase activity of both species and mutant is again strongly inhibited, suggesting an inhibitory effect on yeast F1 activity not detected so far.     

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.     

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.     

Characterization of alternative respiratory pathways in the yeast Schwanniomyces castellii by the study of mutants deficient in cytochromes a+a3 and/or b.

After a general review of the proposed mechanisms and physiological roles of the alternative respiratory pathways found in various organisms, the studies are focussed on the amylolytic yeast Schwaniomyces castellii. In addition to the cytochrome chain, the wild type presents two alternative pathways insensitive to antimycin A. One is salicylhydroxamic acid (SHAM)-sensitive and azide-insensitive; the other is SHAM-insensitive and sensitive to high azide concentration. Conditions for mutagenesis and screening are described, which allow isolation of mutants deficient in cytochromes a+a3 and/or b in this yeast previously classified as petite negative. The relative proportions of the alternative respiratory pathways are compared in the wild type and mutant strains following inhibition by SHAM and azide at optimal concentration as determined by iso-inhibition curves. The growth of the cytochrome deficient mutants on citrate, a non-fermentable carbon source, and the ability of the wild type to grow on citrate+antimycin A, after a lag of about 10 h, indicate an involvement of the alternative pathway(s) in energy production. Rotenone sensitivity of respiration and ATP level confirm the presence of a functional phosphorylation site 1. The role of each alternative respiratory pathway in energy production is discussed.     

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.     

Importance of medium pH in solid state fermentation for growth of Schwanniomyces castellii

Utilization of soluble starch by Schwanniomyces castellii in a solid state fermentation system was highest in unbuffered medium when initial and final pH of the medium were 6.5–7.0 and 4.0–4.6, respectively. An economic strategy involving the use of urea as a sole nitrogen source in medium with initial pH of 6.5 allowed maximum substrate utilization in the absence of buffer and without any contamination in column fermenter.     

Single-cell protein production from potato starch by the yeast Schwanniomyces alluvius

Fully aerated cultures of Schwanniomyces alluvius grew on 4% soluble potato starch in a defined minimal medium at a doubling time of 1.5 h at 30°C. A recovery of 51% (0.51 g of dried biomass from 1 g of starch) was obtained. Yields and growth rates of cultures on starch were similar to those on glucose.