Production of citric acid from sugars present in wood hemicellulose usingAspergillus niger andSaccharomycopsis lipolytica

Summary One strain each of the fungus,Aspergillus niger, and the yeast,Saccharomycopsis lipolytica, were investigated for their ability to produce citric acid from the sugars present in hemicellulose hydrolysates.S. lipolytica produced citric acid as efficiently from mannose as from glucose, but failed to assimilate xylose, arabinose or galactose.A. niger readily assimilated mannose, xylose and arabinose, and produced citric acid from these sugars although the yields were lower than from glucose. A possible inhibitory effect of arabinose on citric acid production from other sugars was observed usingA. niger.     

2,3-Butanediol production from xylose and other hemicellulosic components by Bacillus polymyxa

In the xylose fermentation of Bacillus polymyxa strain 9035, best 2,3-butanediol yields were obtained with 1.0 % yeast extract, 4–6 % xylose, shaking at 125 rpm and incubation at 30°C. Under these conditions, mannose, galactose, L-arabinose, cellobiose, starch and glucose were readily metabolized and yielded significant amounts of diol. Diol production from xylan was also demonstrated. In addition, the screening of a number of B. polymyxa strains on xylose revealed that only strains 9031-1 and 9035 used xylose extensively and produced significant amounts of diol. The latter strain proved best under scaled-up conditions.NRCC #22775     

Characterization of extracellular substrate specific glucose and xylose isomerases of Chainia

Summary Specific glucose and xylose isomerases have been identified in cell-free culture filtrates of a Chainia species. Treatment with DEAE-cellulose selectively adsorbed xylose isomerase activity while only the glucose isomerase was adsorbed on CM-cellulose. Glucose isomerase was completely inhibited by xylose at 1.3 × 10^-4 M concentration. The differential identity of the extracellular glucose and xylose isomerases, unique to Chainia, is discussed.(NCL Communication 3562)     

Use of elephant grass (Pennisetum purpureum) acid hydrolysate for microbial oil production by Trichosporon cutaneum

Elephant grass (Pennisetum purpureum) dilute acid hydrolysate contains 34.6?g/L total sugars. The potential of lipid production by oleaginous yeast Trichosporon cutaneum grown on elephant grass acid hydrolysate was investigated for the first time. During the fermentation process on the elephant grass acid hydrolysate, glucose, xylose, and arabinose could be well utilized as carbon sources by T. cutaneum. Interestingly, xylose was almost no use before glucose was consumed completely. This illustrated that simultaneous saccharification of xylose and glucose by T. cutaneum did not occur on elephant grass acid hydrolysate. The highest biomass, lipid content, lipid yield, and lipid coefficient of T. cutaneum were measured after the sixth day of fermentation and were 22.76?g/L, 24.0%, 5.46?g/L, and 16.1%, respectively. Therefore, elephant grass is a promising raw material for microbial oil production by T. cutaneum.     

Identification of oleaginous yeast strains able to accumulate high intracellular lipids when cultivated in alkaline pretreated corn stover

Microbial oil is a potential alternative to food/plant-derived biodiesel fuel. Our previous screening studies identified a wide range of oleaginous yeast species, using a defined laboratory medium known to stimulate lipid accumulation. In this study, the ability of these yeasts to grow and accumulate lipids was further investigated in synthetic hydrolysate (SynH) and authentic ammonia fiber expansion (AFEX?)-pretreated corn stover hydrolysate (ACSH). Most yeast strains tested were able to accumulate lipids in SynH, but only a few were able to grow and accumulate lipids in ACSH medium. Cryptococcus humicola UCDFST 10-1004 was able to accumulate as high as 15.5 g/L lipids, out of a total of 36 g/L cellular biomass when grown in ACSH, with a cellular lipid content of 40 % of cell dry weight. This lipid production is among the highest reported values for oleaginous yeasts grown in authentic hydrolysate. Preculturing in SynH media with xylose as sole carbon source enabled yeasts to assimilate both glucose and xylose more efficiently in the subsequent hydrolysate medium. This study demonstrates that ACSH is a suitable medium for certain oleaginous yeasts to convert lignocellullosic sugars to triacylglycerols for production of biodiesel and other valuable oleochemicals.     

D-Xylose fermentation to ethanol bySchizosaccharomyces pombe cloned with xylose isomerase gene

Summary Schizosaccharomyces pombe cloned with the xylose isomerase gene from E. coli is able to grow on YNB and YMP broths containing xylose as the sole carbon source. This yeast can ferment D-xylose to ethanol directly; however, the ethanol production rate and the yield were dependent on the nitrogen source. With the YMP broth as a nitrogen source, the final ethanol concentration can reach 3.7% (w/v), and the ethanol yield was 80% of the theoretical value based on the amount of xylose that was metabolized. The ethanol production is slow, and the xylitol production is still very active; apparently, the limiting step is the isomerization of xylose to xylulose.     

Molecular cloning of theEscherichia coli gene encoding xylose isomerase

Summary A 2.4 Kb DNA fragment restricted from a Clarke-Carbon ColEl plasmid, pLC32-9, containing the xylose isomerase gene has been inserted into the PstI site of pDB248, a shuttle plasmid between the bacteriumE. coli and the fission yeast,Schizosaccharomyces pombe. This recombinant plasmid, pDB248-XI, can genetically complement xylose isomerase deficientE. coli strains and xylose isomerase gene can be expressed inSchizosaccharomyces pombe.     

Characterization of a bacterial xylanase resistant to repression by glucose and xylose

Summary Bacillus subtilis CD4, when grown in nutrient broth or minimal medium in presence of xylan, produced extracellular xylanase that hydrolyzed xylan optimally at pH 5. The enzyme was induced by xylan, xylose and glucose. Addition of xylose or glucose in xylan containing medium did not affect enzyme production. The structural gene encoding xylanase was cloned and expressed in E. coli. The recombinant enzyme exhibited similar properties like that of native enzyme including resistance to repression by xylose and glucose.     

High density cell culture ofRhodotorula glutinis using oxygen-enriched air

Summary Fed-batch culture ofRhodotorula glutinis, an oleaginous yeast, was carried out to obtain high biomass concentration. With air, final concentration of biomass was 100–110 g/L, whereas with oxygen-enriched air, 185 g/L was obtained. Obligate aerobic metabolism ofR. glutinis under oxygen limitation and low oxygen requirement under lipidaccumulating condition were found to be advantageous for the high density culture of this microbe.     

糠醛对粘红酵母生长与油脂积累的影响

为了探究纤维素水解液中常见的发酵抑制物糠醛对粘红酵母Rhodotorula glutinis生长与油脂积累的影响,对比了不同的糠醛浓度(0.1、0.4、0.6、1.5 g/L)下粘红酵母的生物量和油脂积累情况,并探究了1.0 g/L的糠醛对粘红酵母不同碳源(葡萄糖和木糖)利用的影响。研究表明,当糠醛浓度达1.5 g/L时,粘红酵母的延迟期延长至96 h,残糖高达17.7 g/L,生物量最高6.6 g/L,仅为正常积累量的47%,油脂含量也减少了约50%;以木糖为碳源时,糠醛对粘红酵母的抑制程度小于葡萄糖为碳源时的情况;在糠醛存在的逆境中,粘红酵母倾向于生成更多的18碳脂肪酸或18碳不饱和脂肪酸。     

Bioemulsifier production by an oleaginous yeastRhodotorula glutinis IIP-30

Summary A locally isolated oleaginous strain ofRhodotorula glutinis strain IIP-30 produced a growth associated extracellular emulsifying agent while utilizing glucose during fed batch fermentation under nitrogen limitation at 30°C and pH 4. 0. Similar optimum conditions were also noted for intracellular lipid accumulation.     

Monitoring growth and lipid production of new isolated oleaginous yeast Cryptococcus aerius UIMC65 on glucose and xylose cultures

Process monitoring is one of the most important factors affecting production efficiency at industrial scale bioprocesses. In the present work, Flow-cytometric analysis has been employed to monitor and determine neutral lipid cell droplets, granularity and size of the cells of the new oleaginous yeast, Cryptococcus aerius UIMC65. It has been shown that, differences of fluorescent intensity as well as side and forward scatter light properties have close correlations with the differences in lipid production by these yeast cells. The lipid content-related fluorescent intensity versus forward scatter parameter has been used to monitor and compare different subpopulations during growth phases on both glucose and xylose in batch cultures. Flow cytometric results have revealed that the observed differences in the proportion of each subpopulation were related to the specific growth phase and lipid content of the cells. The highest lipid content and lipid productivity were attained at 82.62%, 4.47 g/L (at 72 h) and 78.41%, 6.21 g/L (at 60 h) on glucose and xylose growth cultures, respectively. The highest biomass, lipid yield and biomass yield were found to be 7.92 g/L (on glucose culture, at 60 h), 20.92% (on glucose culture, at 48 h) and 50.71% (on glucose culture, at 24 h), respectively.     

High‐yield lipid production from lignocellulosic biomass using engineered xylose‐utilizing Yarrowia lipolytica

Lignocellulosic biomass shows high potential as a renewable feedstock for use in biodiesel production via microbial fermentation. Yarrowia lipolytica, an emerging oleaginous yeast, has been engineered to efficiently convert xylose, the second most abundant sugar in lignocellulosic biomass, into lipids for lignocellulosic biodiesel production. Yet, the lipid yield from xylose or lignocellulosic biomass remains far lower than that from glucose. Here we developed an efficient xylose‐utilizing Y. lipolytica strain, expressing an isomerase‐based pathway, to achieve high‐yield lipid production from lignocellulosic biomass. The newly developed xylose‐utilizing Y. lipolytica, YSXID, produced 12.01 g/L lipids with a maximum yield of 0.16 g/g, the highest ever reported, from lignocellulosic hydrolysates. Consequently, this study shows the potential of isomerase‐based xylose‐utilizing Y. lipolytica for economical and sustainable production of biodiesel and oleochemicals from lignocellulosic biomass.     

Xylose isomerase improves growth and ethanol production rates from biomass sugars for both Saccharomyces pastorianus and Saccharomyces cerevisiae

The demand for biofuel ethanol made from clean, renewable nonfood sources is growing. Cellulosic biomass, such as switch grass (Panicum virgatum L.), is an alternative feedstock for ethanol production; however, cellulosic feedstock hydrolysates contain high levels of xylose, which needs to be converted to ethanol to meet economic feasibility. In this study, the effects of xylose isomerase on cell growth and ethanol production from biomass sugars representative of switch grass were investigated using low cell density cultures. The lager yeast species Saccharomyces pastorianus was grown with immobilized xylose isomerase in the fermentation step to determine the impact of the glucose and xylose concentrations on the ethanol production rates. Ethanol production rates were improved due to xylose isomerase; however, the positive effect was not due solely to the conversion of xylose to xylulose. Xylose isomerase also has glucose isomerase activity, so to better understand the impact of the xylose isomerase on S. pastorianus, growth and ethanol production were examined in cultures provided fructose as the sole carbon. It was observed that growth and ethanol production rates were higher for the fructose cultures with xylose isomerase even in the absence of xylose. To determine whether the positive effects of xylose isomerase extended to other yeast species, a side-by-side comparison of S. pastorianus and Saccharomyces cerevisiae was conducted. These comparisons demonstrated that the xylose isomerase increased ethanol productivity for both the yeast species by increasing the glucose consumption rate. These results suggest that xylose isomerase can contribute to improved ethanol productivity, even without significant xylose conversion.     

Optimization of C16 and C18 fatty alcohol production by an engineered strain of <Emphasis Type="Italic">Lipomyces starkeyi</Emphasis>

The oleaginous yeast Lipomyces starkeyi was engineered for the production of long-chain fatty alcohols by expressing a fatty acyl-CoA reductase, mFAR1, from Mus musculus. The optimal conditions for production of fatty alcohols by this strain were investigated. Increased carbon-to-nitrogen ratios led to efficient C16 and C18 fatty alcohol production from glucose, xylose and glycerol. Batch cultivation resulted in a titer of 1.7 g/L fatty alcohol from glucose which represents a yield of 28 mg of fatty alcohols per gram of glucose. This relatively high level of production with minimal genetic modification indicates that L. starkeyi may be an excellent host for the bioconversion of carbon-rich waste streams, particularly lignocellulosic waste, to C16 and C18 fatty alcohols.     

Butanol production of Clostridium acetobutylicum grown on sugars found in hemicellulose hydrolysates

Summary Clostridium acetobutylicum was grown on a variety of different media and assayed for enhanced butanol production. Butanol values of about 90% of theoretical were obtained when glucose was used as the substrate. Five other sugars were assayed and the order of butanol production using these substrates was cellobiose>mannose>arabinose>xylose >galactose. The addition of calcium carbonate to the media to enhance xylose utilized was beneficial although lower butanol values were obtained.     

High activity extracellular glucose (xylose) isomerase from a Chainia species

Summary A sclerotia-forming actinomycete of the genus Chainia secreted high levels of glucose (xylose) isomerase when grown in submerged culture on a wheat bran - yeast extract medium. Maximum activity (4 units/ml) was obtained after 3–4 days when the cell bound activity was 0.19 units/ml. The two enzymes differed significantly in pH optima (extracellular, 9.5; cell-bound, 7.0) and in their adsorption behaviour on CM and DEAE celluloses. Both Mg⁺⁺ and Co⁺⁺ are required by the cell-bound enzyme for its optimum activity while either Mg⁺⁺ or Co⁺⁺ is necessary for the extracellular enzyme.NCL Communication 3320     

NADPH-specific and NADH-specific xylose reduction is catalyzed by two separate enzymes in Pachysolen tannophilus

Summary Pachysolen tannophilus contains — in addition to an NADPH-linked xylose reductase — a separate NADH-linked one, in this respect differing from the yeast Pichia stipitis. Both enzyme proteins can conveniently be separated from each other by either ion exchange chromatography or chromatofocusing.     

Draft Genome Assembly and Annotation for Cutaneotrichosporon dermatis NICC30027, an Oleaginous Yeast Capable of Simultaneous Glucose and Xylose Assimilation

The identification of oleaginous yeast species capable of simultaneously utilizing xylose and glucose as substrates to generate value-added biological products is an area of key economic interest. We have previously demonstrated that the Cutaneotrichosporon dermatis NICC30027 yeast strain is capable of simultaneously assimilating both xylose and glucose, resulting in considerable lipid accumulation. However, as no high-quality genome sequencing data or associated annotations for this strain are available at present, it remains challenging to study the metabolic mechanisms underlying this phenotype. Herein, we report a 39,305,439 bp draft genome assembly for C. dermatis NICC30027 comprised of 37 scaffolds, with 60.15% GC content. Within this genome, we identified 524 tRNAs, 142 sRNAs, 53 miRNAs, 28 snRNAs, and eight rRNA clusters. Moreover, repeat sequences totaling 1,032,129 bp in length were identified (2.63% of the genome), as were 14,238 unigenes that were 1,789.35 bp in length on average (64.82% of the genome). The NCBI non-redundant protein sequences (NR) database was employed to successfully annotate 11,795 of these unigenes, while 3,621 and 11,902 were annotated with the Swiss-Prot and TrEMBL databases, respectively. Unigenes were additionally subjected to pathway enrichment analyses using the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Cluster of Orthologous Groups of proteins (COG), Clusters of orthologous groups for eukaryotic complete genomes (KOG), and Non-supervised Orthologous Groups (eggNOG) databases. Together, these results provide a foundation for future studies aimed at clarifying the mechanistic basis for the ability of C. dermatis NICC30027 to simultaneously utilize glucose and xylose to synthesize lipids.