高浓度盐对耐高渗酵母产多元醇的影响

假丝酵母OS－300菌株在含有30％葡萄糖的高浓度基质中能产三种多元醇,甘油,阿拉伯醇和赤藓糖醇,但是在含18％NaCl的高浓度基质中产甘油,。同时,还发现几种原来不产甘油的耐高渗酵母在含有9％NaCl的高浓度培养液中也能形成大量的甘油,该现象表明：产多元醇耐高渗酵母的代谢途径可以在高盐浓度下被明显地改变。     

Studies on Osmophilic Yeasts

Various conditions were studied which gave influences on polyalcohol production by Pichia miso. Pichia miso gave an excellent yield of polyalcohol showing good growth in the vitamin-free medium. Unlike Zygosaccharomyces high concentrations of phosphate such as 2% as KH₂PO₄ in the medium showed no detrimental effect on polyalcohol production. Remarkable reduction in polyalcohol yield was observed in the medium of high concentration of nitrogen sources, among which yeast extract showed the most striking effect. Polyalcohol fermentation was provoked in the medium of 0.1% yeast extract while the metabolic activity distinctly converted to ethanol fermentation when the organisms were incubated in the medium of 4.0% yeast extract. The fact that a large amount of ethanol more than 6% in the medium was produced aerobically by genus Pichia of oxidative dissimilation type seemed to be very interesting and noticeable. For the purpose of industrial production, it was shown that polyalcohol production in jar-fermenter scale was achieved with as good yield as that in shaking flask culture.     

Studies on Osmophilic Yeasts

In a medium containing high concentration of yeast extract was observed aerobic formation of a large amount of ethanol by various yeasts of non-fermenting or extremely poor fermenting type such as Torulopsis famata, Candida polymorpha and Pichia membranaefaciens etc. Acetaldehyde formed as a metabolic intermediate leading to ethanol formation was trapped by the addition of sulfite and was identified as 2,4-dinitrophenyl-hydrazone.     

Determination of Growth and Glycerol Production Kinetics of a Wine Yeast Strain Saccharomyces cerevisiae Kalecik 1 in Different Substrate Media

Summary Glycerol has been known as an important by-product of wine fermentations improving the sensory quality of wine. This study was carried out with an endogenic wine yeast strain Saccharomyces cerevisiae Kalecik 1. The kinetics of growth and glycerol biosynthesis were analysed at various initial concentrations of glucose, fructose, and sucrose in a batch system. Depending on the determined values of Monod constants, glucose (K_s = 28.09 g/l) was found as the most suitable substrate for the yeast growth. Initial glucose, fructose and sucrose concentrations necessary for maximum specific yeast growth rate were determined as 175 g, 100 l, and 200 g/l, respectively. The yeast produced glycerol at very high concentrations in fructose medium. Fructose was determined as the most suitable substrate for glycerol production while the strain showed low tendency to use it for growth. S. cerevisiae Kalecik 1 could not produce glycerol below 200 g/l initial sucrose concentration. When natural white grape juice was used as fermentation medium, maximum glycerol concentration and dry weight of the yeast were determined as 9.3 g/l and 11.8 g/l, respectively.     

Studies on Osmophilic Yeasts

Assimilation of galactose and maltose by Saccharomyces rouxii, which is a typical salt-tolerant yeast playing an important role in soy-brewing, was negligible or extremely poor in the medium containing 18% NaCl, although the assimilation in the ordinary medium was vigorous. The yeasts which were able to assimilate and ferment galactose, maltose and/or saccharose in the high-saline medium were limited to a few strains. From the studies on balances of fermentation products, it was revealed that ethanol yield based on sugar consumed in the 18% NaCl-medium was lower than that in the ordinary medium, suggesting that other fermentation products than ethanol should be accumulated. Analytical results of the fermented broths showed much polyalcohol production in the high-saline medium.     

Influence of invertase activity and glycerol synthesis and retention on fermentation of media with a high sugar concentration by Saccharomyces cerevisiae.

In the past, the fermentation activity of Saccharomyces cerevisiae in substrates with a high concentration of sucrose (HSuc), such as sweet bread doughs, has been linked inversely to invertase activity of yeast strains. The present work defines the limits of the relationship between invertase activity and fermentation in hyperosmotic HSuc medium. Fourteen polyploid, wild-type strains of S. cerevisiae with different invertase levels gave a similar ranking of fermentation activity in HSuc and in medium in which glucose and fructose replaced sucrose (HGF medium). Thus, invertase is unlikely to be the most important determinant of fermentation in sweet doughs. Yeasts produce the compatible solute-osmoprotective compound glycerol when exposed to hyperosmotic environments. Under low sugar concentrations (and nonstressing osmotic pressure), there was no correlation between glycerol and fermentation activities. However, there was a strong correlation between the ability of yeasts to ferment in HSuc or HGF medium and their capacity to produce and retain glycerol intracellularly. There was also a strong correlation between intracellular glycerol and fermentation activity of yeasts in a medium in which the nonfermentable sugar alcohol sorbitol replaced most of the sugars (HSor), but the ability to produce and retain glycerol was greater when yeasts were incubated in HGF medium under the same osmotic pressure. The difference between the amounts of glycerol produced and retained in HSor and in HGF media varied with strains. This implies that high fermentable sugar concentrations cause physiological conditions that allow for enhanced glycerol production and retention, the degree of which is strain dependent. In conclusion, one important prerequisite for yeast strains to ferment media with high concentrations of sugar is the ability to synthesize glycerol and especially to retain it.     

Cassava as a Cheap Source of Carbon for Rhizobial Inoculant Production using an Amylase-producing Fungus and a Glycerol-producing Yeast

Summary The aim of this research was to develop methods to use low-cost carbon compounds for rhizobial inoculant production. Five raw starch materials; steamed cassava, sticky rice, fresh corn, dry corn and sorghum were tested for sugar production by an amylase-producing fungus. Streamed cassava produced the highest amount of reducing sugar after fermentation. Bradyrhizobium japonicum USDA110, Azorhizobium caulinodans IRBG23, Rhizobium phaseoli TAL1383, Sinorhizobium fredii HH103, and Mesorhizobium ciceri USDA2429 were tested on minimal medium supplemented with reducing sugar obtained from cassava fermentation. All strains, except B. japonicum USDA110, could grow in medium containing cassava sugar derived from 100 g steamed cassava per litre, and the growth rates for these strains were similar to those in medium containing 0.5 (w/v) mannitol. The sugar derived from steamed cassava was further used for production of glycerol using yeast. After 1 day of yeast fermentation, the culture containing glycerol and heat-killed yeast cells, was used to formulate media for culturing bradyrhizobia. A formulation medium, FM4, with a glycerol concentration of 0.6 g/l and yeast cells (OD₆₀₀ = 0.1) supported growth of B. japonicum USDA110 up to 3.61 × 10⁹ c.f.u./ml in 7 days. These results demonstrate that steamed cassava could be used to provide cheap and effective carbon sources for rhizobial inoculant production.     

Hyperosmotic stress response by strains of bakers' yeasts in high sugar concentration medium

Four strains of bakers' yeast were analysed for their hyperosmotic responses when in media that mimic conditions occurring in bread doughs. Two of the strains produced strong fermentative activity in medium with low osmotic stress, but produced considerably less ethanol in high sucrose concentration medium. Two other strains produced more similar fermentation activities across the range of media tested. The strains that were inhibited by high sucrose concentration were unable to produce significant amounts of glycerol under hyperosmotic conditions. By contrast, the yeasts that were not inhibited significantly by high sucrose produced a considerable amount of glycerol. The strains that produced significant glycerol exhibited efficient expression of the glycerol-3-phosphate dehydrogenase gene GPD1. These novel data on the molecular responses of industrially relevant strains of bakers' yeasts are prerequisite to designing strategies for improving the performance of industrial yeasts in high sugar concentration media.     

Polyalcohol Production by Pichia miso in a Jar Fermentor

Cultural conditions for polyalcohol production by Pichia miso were examined in Waldhof type 20 liter-fermentor scale. The best result was obtained under conditions where the aeration rate was 1 volume per volume of the medium per minute with the stirring rate of 500r.p.m., (Kd=5×l0^-6 [g-mol of O₂/atm. min. ml.]); in 5 days incubation, Pichia miso completely dissimilated the glucose of a high concentration, 30%, and produced glycerol, D-arabitol and erythritol in a very high yield, 50% of sugar consumed. The greatest advantage compared with the shake flask culture is that the required fermentation time is shortened to half.     

Quantitative production of xylitol from D-xylose by a high-xylitol producing yeast mutant Candida tropicalis HXP2

Summary Xylitol was produced as a metabolic by-product by a number of yeasts when grown on medium containing D-xylose as carbon and energy sources. Among the yeast strains tested, a mutant strain of Candida tropicalis (HXP2) was found to produce xylitol from D-xylose with a high yield (>90%). Ethanol was also produced by HXP2 when D-glucose, D-fructose, or sucrose were used as substrates. The high-xylitol-producing yeast mutant is a good organism for the production of xylitol from biomass that contains D-xylose.     

Inhibition of Cathepsin B1 by E-64, a Thiol Proteinase Inhibitor,and Its Derivatives

Four formaldehyde-resistant yeasts were isolated from soil. Three were tentatively identified as Debaryomyces vanriji and one as Trichosporon penicillatum. These yeasts almost completely consumed formaldehyde at 0.15 to 0.55% in growth medium containing glucose as carbon source, but the carbon of formaldehyde was not incorporated into the cell constituents. In formaldehyde-containing medium, yeast growth occurred after formaldehyde consumption. The yeasts showed relatively high activities of formaldehyde dehydrogenase, S-formylglutathione hydrolase and formate dehydrogenase. The resistance to formaldehyde is attributed to detoxification by oxidation.     

Combined effects of endo- and exogenous trehalose on stress tolerance and biocontrol efficacy of two antagonistic yeasts

Effects of endo- and exogenous trehalose on viability of two antagonistic yeasts, Cryptococcus laurentii (Kuffer.) Skinner and Rhodotorula glutinis (Fresen.) Harrison, were investigated after being treated with rapid-freezing, slow-freezing and freeze-drying, respectively. The accumulation of intracellular trehalose in the two yeasts was induced by culturing the yeast cells in trehalose-containing medium, which significantly enhanced viabilities of both yeasts in the slow-freezing test. Trehalose, as an exogenous protectant, at the concentration of 5% or 10% could markedly increase survivals of the two yeasts when subjected to freeze-drying. When combined with exogenous trehalose as a protective substance, the yeasts containing high intracellular trehalose level showed higher viabilities as compared to those containing low levels under both freezing and freeze-drying stresses. The highest survival of C. laurentii and R. glutinis were 90% and 97% after freeze-drying, respectively, compared to 63% and 28% for the yeasts with lower intracellular trehalose levels. These results may be due to the fact that a combined effect occurred between endo- and exogenous trehalose of yeast cells. The combined effect on C. laurentii and R. glutinis also resulted in the highest level of biocontrol efficacy against blue mold in apple fruit caused by Penicillium expansum Link, and reduced the disease indexes to 45 and 56, respectively, compared to 94 and 81 in the untreated control. Meanwhile, the combination of endo- and exogenous trehalose significantly increased population of both yeasts in apple wounds, especially at the first 48 h after inoculation, which might explain the reason of the improvement in biocontrol effects of the two yeasts.     

Ethanol-sensitive mutants of Saccharomyces cerevisiae

Saccharomyces cerevisiae mutants unable to grow at ethanol concentrations at which the wild type strain S288C does grow, have been isolated. Some of them show additional phenotypic alterations in colony size, temperature sensitivity and viability in ethanol, which cosegregate with the growth sensitivity in ethanol. 21 selected monogenic ethanol-sensitive mutants define 20 complementation groups, denominated ETA1 to ETA20, which indicates that there is a high number of genes involved in the ethanol tolerance/sensitivity mechanism.Out of 21 selected monogenic mutants, 20 are not altered in the glycolytic pathway since, when maintained in glucosesupplemented medium, they can produce as much ethanol as the wild type and at about the same velocity. Nor do any of the mutants seem to be altered in the lipid biosynthetic pathway since, whether grown in the absence or in the presence of ethanol, their concentration of fatty acids and ergosterol is similar to that of the wild type under the same conditions. Therefore growth sensitivity to ethanol does not seem necessarily to be related to carbohydrate or lipid metabolism.Non-common abbreviations YP yeast extract peptone medium - YPD yeast extract peptone dextrose agar or medium - YPG yeast extract peptone glycerol agar - YPDE yeast extract peptone dextrose ethanol agar or medium - SD yeast nitrogen base dextrose agar - SPO yeast extract potassium acetate glucose agar - PD parental ditype - NPD non-parental ditype - TT tetratype     

Characteristics of the Aroma Constitution Found in Native China Black Teas

The distribution of glutathione S-transferase (GST) (EC 2.5.1.18) in yeasts was investigated. High enzyme activity was found in some strains of Issatchenkia and Candida. Of 168 strains tested, Iss. orientalis showed the highest activity. The enzyme activity exists constitutively in the yeast cells but it increased with the addition of an enzyme substrate, o-dinitrobenzene, to the culture medium. Moreover, the addition of l-cysteine and glycine to the medium also increased the enzyme activity. This enzyme was so unstable that it lost almost all its activity on ammonium sulfate precipitation and 93% of its activity was lost when it was stored at 4°C for two weeks in a soluble state. We found that it was stabilized considerably in a solution containing 20% glycerol, 1 mm EDTA, 2 mm DTT and 10 mm sodium sulfite.     

Phenotypic expression of Kluyveromyces lactis killer toxin against Saccharomyces spp.

The secretion of killer toxins by some strains of yeasts is a phenomenon of significant industrial importance. The activity of a recently discovered Kluyveromyces lactis killer strain against a sensitive Saccharomyces cerevisiae strain was determined on peptone-yeast extract-nutrient agar plates containing as the carbon source glucose, fructose, galactose, maltose, or glycerol at pH 4.5 or 6.5. Enhanced activity (50 to 90% increase) was found at pH 6.5, particularly on the plates containing galactose, maltose, or glycerol, although production of the toxin in liquid medium was not significantly different with either glucose or galactose as the carbon source. Results indicated that the action of the K. lactis toxin was not mediated by catabolite repression in the sensitive strain. Sensitivities of different haploid and polyploid Saccharomyces yeasts to the two different killer yeasts S. cerevisiae (RNA-plasmid-coded toxin) and K. lactis (DNA-plasmid-coded toxin) were tested. Three industrial polyploid yeasts sensitive to the S. cerevisiae killer yeast were resistant to the K. lactis killer yeast. The S. cerevisiae killer strain itself, however, was sensitive to the K. lactis killer yeast.     

Phenotypic expression of Kluyveromyces lactis killer toxin against Saccharomyces spp

The secretion of killer toxins by some strains of yeasts is a phenomenon of significant industrial importance. The activity of a recently discovered Kluyveromyces lactis killer strain against a sensitive Saccharomyces cerevisiae strain was determined on peptone-yeast extract-nutrient agar plates containing as the carbon source glucose, fructose, galactose, maltose, or glycerol at pH 4.5 or 6.5. Enhanced activity (50 to 90% increase) was found at pH 6.5, particularly on the plates containing galactose, maltose, or glycerol, although production of the toxin in liquid medium was not significantly different with either glucose or galactose as the carbon source. Results indicated that the action of the K. lactis toxin was not mediated by catabolite repression in the sensitive strain. Sensitivities of different haploid and polyploid Saccharomyces yeasts to the two different killer yeasts S. cerevisiae (RNA-plasmid-coded toxin) and K. lactis (DNA-plasmid-coded toxin) were tested. Three industrial polyploid yeasts sensitive to the S. cerevisiae killer yeast were resistant to the K. lactis killer yeast. The S. cerevisiae killer strain itself, however, was sensitive to the K. lactis killer yeast.     

The ability of Pichia kudriavzevii to tolerate multiple stresses makes it promising for developing improved bioethanol production processes

Thermotolerant ethanol fermenting yeasts have been extensively used in industrial bioethanol production. However, little is known about yeast physiology under stress during bioethanol processing. This study investigated the physiological characteristics of the thermotolerant yeast Pichia kudriavzevii, strains NUNS-4, NUNS-5 and NUNS-6, under the multiple stresses of heat, ethanol and sodium chloride. Results showed that NUNS-4, NUNS-5 and NUNS-6 displayed higher growth rates under each stress condition than the reference strain, Saccharomyces cerevisiae TISTR5606. Maximum specific growth rates under stresses of heat (45°C), 15% v/v ethanol and 1·0 M sodium chloride were 0·23 ± 0·04 (NUNS-4), 0·11 ± 0·01 (NUNS-5) and 0·15 ± 0·01 h^–1 (NUNS-5), respectively. Morphological features of all yeast studied changed distinctly with the production of granules and vacuoles when exposed to ethanol, and cells were elongated under increased sodium chloride concentration. This study suggests that the three P. kudriavzevii strains are potential candidates to use in industrial–scale fermentation due to a high specific growth rate under multiple stress conditions. Multiple stress-tolerant P. kudriavzevii NUNS strains have received much attention not only for improving large-scale fuel ethanol production, but also for utilizing these strains in other biotechnological industries.     

Enrichment of a continuous culture of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> with the yeast <Emphasis Type="Italic">Issatchenkia orientalis</Emphasis> in the production of ethanol at increasing temperatures

A fermentation system was continuously fed with sugar-cane syrup and operated with recycling of Saccharomyces cerevisiae cells at temperatures varying from 30 to 47°C. The aim of the present work was to obtain and study the colonies of isolates showing elongated cells of yeasts which were sporadically observed at the end of this continuous process. Based on a sequence of assays involving methods of classical taxonomy and RAPD-PCR, two groups of isolates showing characteristics of non-Saccharomyces yeasts were identified in the yeast population where S. cerevisiae was the dominant yeast. The largest group of non-Saccharomyces yeasts, resulting from a slow proliferation over the 2 months, reached a final level of 29.6% at the end of the process. RAPD-PCR profiles obtained for the isolates of this dominant non-Saccharomyces yeast indicated that they were isolates of Issatchenkia orientalis. Pichia membranifaciens was the only species of non-Saccharomyces yeast detected together with I. orientalis but at a very low frequency. The optimum temperature for ethanol formation shown by the isolate 195B of I. orientalis was 42°C. This strain also showed a faster ethanol formation and biomass accumulation than the thermotolerant strain of S. cerevisiae used as the starter of this fermentation process. Some isolates of I. orientalis were also able to grow better at 40°C than at 30°C on plates containing glycerol as carbon source. Yeasts able to grow and produce ethanol at high temperatures can extend the fermentation process beyond the temperature limits tolerated by S. cerevisiae.     

不同氯化钠浓度对Candida etchellsii CICIM Y0600生长和生物特性的影响研究

在酱油发酵过程中增香酵母的代谢产物能显著提高酱油产品的风味和品质.为了提高增香酵母菌在酱醪中的适应性,通过逐步提高氯化钠浓度的方法对酵母菌的耐盐度进行驯化培养.通过分析表明,经过5个批次的驯化,增香酵母菌的耐氯化钠浓度由180 g/L提高到240 g/L.对驯化后的三株耐不同氯化钠浓度的增香酵母菌株进行生物特性研究.结果表明,在200 g/L氯化钠浓度的培养基中,Candida etchellsii CICIM Y0600产游离氨基酸和酸解氨基酸总量分别达到2.01g/L和7.00 g/L.和其他不同氯化钠浓度条件下比较,其产有机酸种类没有变化,挥发性酯类物质含量最高,为0.80 g/L.耐盐度的提高使增香酵母菌更好地适应了在酱油发酵中的高盐度环境.