Sugar Consumption Rate Research Articles

Efficient Acetone–Butanol–Ethanol Production from Corncob with a New Pretreatment Technology—Wet Disk Milling

Acetone–butanol–ethanol (ABE) production from corncob was achieved using an integrated process combining wet disk milling (WDM) pretreatment with enzymatic hydrolysis and fermentation by Clostridium acetobutylicum SE-1. Sugar yields of 71.3 % for glucose and 39.1 % for xylose from pretreated corncob were observed after enzymatic hydrolysis. The relationship between sugar yields and particle size of the pretreated corncob was investigated, suggesting a smaller particle size benefits enzymatic hydrolysis with the WDM pretreatment approach. Analysis of the correlation between parameters representing particle size and efficiency of enzymatic hydrolysis predicted that frequency 90 % is the best parameter representing particle size for the indication of the readiness of the material for enzymatic hydrolysis. ABE production from corncob was carried out with both separate hydrolysis and fermentation (SHF) and simultaneous saccharification and fermentation (SSF) processes using C. acetobutylicum SE-1. Interestingly, when considering the time for fermentation as the time for ABE production, a comparable rate of sugar consumption and ABE production in the SHF process (0.55 g/l·h sugar consumption and 0.20 g/l·h ABE production) could be observed when glucose (0.50 g/l·h sugar consumption and 0.17 g/l·h ABE production) or a mixture of glucose and xylose (0.68 g/l·h sugar consumption and 0.22 g/l·h ABE production) mimicking the corncob hydrolysate was used as the substrate for fermentation. This result suggested that the WDM is a suitable pretreatment method for ABE production from corncob owing to the mild conditions. A higher ABE production rate could be observed with the SSF process (0.15 g/l·h) comparing with SHF process (0.12 g/l·h) when combining the time for saccharification and fermentation and consider it as the time for ABE production. This is possibly a result of low sustained sugar level during fermentation. These investigations lead to the suggestion that this new WDM pretreatment method has the potentials to be exploited for efficient ABE production from corncob.

Simultaneous utilization of glucose, xylose and arabinose in the presence of acetate by a consortium of Escherichia coli strains

BackgroundThe efficient microbial utilization of lignocellulosic hydrolysates has remained challenging because this material is composed of multiple sugars and also contains growth inhibitors such as acetic acid (acetate). Using an engineered consortium of strains derived from Escherichia coli C and a synthetic medium containing acetate, glucose, xylose and arabinose, we report on both the microbial removal of acetate and the subsequent simultaneous utilization of the sugars.ResultsIn a first stage, a strain unable to utilize glucose, xylose and arabinose (ALS1392, strain E. coli C ptsG manZ glk crr xylA araA) removed 3 g/L acetate within 30 hours. In a subsequent second stage, three E. coli strains (ALS1370, ALS1371, ALS1391), which are each engineered to utilize only one sugar, together simultaneously utilized glucose, xylose and arabinose. The effect of non-metabolizable sugars on the metabolism of the target sugar was minimal. Additionally the deletions necessary to prevent the consumption of one sugar only minimally affected the consumption of a desired sugar. For example, the crr deletion necessary to prevent glucose consumption reduced xylose and arabinose utilization by less than 15% compared to the wild-type. Similarly, the araA deletion used to exclude arabinose consumption did not affect xylose- and glucose-consumption.ConclusionsDespite the modest reduction in the overall rate of sugar consumption due to the various deletions that were required to generate the consortium of strains, the approach constitutes a significant improvement in any single-organism approach to utilize sugars found in lignocellulosic hydrolysate in the presence of acetate.

A molecular transporter engineering approach to improving xylose catabolism in Saccharomyces cerevisiae

Traditional metabolic pathway engineering rarely considers the influence of molecular transport. Here, we describe the directed evolution of two heterologous transporters, Candida intermedia GXS1 and Scheffersomyces stipitis XUT3. Growth rate on xylose was improved up to 70% by mutant transporter expression. Most mutants were found to exhibit vastly improved Vmax values and display an increase in high cell density sugar consumption rates. Mixed glucose and xylose fermentations reveal that mutant transporters can alter the diauxic shift dynamics and the simultaneous sugar utilization capacity of the host strain. Analysis of mutations highlights several important residues influencing transporter function including point mutations at F40 of C. intermedia GXS1 and at E538 of S. stipitis XUT3. This work is the first to demonstrate that molecular transporter proteins can be improved for biotechnological applications through directed evolution in yeast.

Effects of Wort Gravity and Nitrogen Level on Fermentation Performance of Brewer’s Yeast and the Formation of Flavor Volatiles

Normal gravity wort and high gravity wort with different nitrogen levels were used to examine their effects on the fermentation performance of brewer's yeast and the formation of flavor volatiles. Results showed that both the wort gravity and nitrogen level had significant impacts on the growth rate, viability, flocculation, and gene expression of brewer's yeast and the levels of flavor volatiles. The sugar (glucose, maltose, and maltotriose) consumption rates and net cell growth decreased when high gravity worts were used, while these increased with increasing nitrogen level. Moreover, high gravity resulted in lower expression levels of ATF1, BAP2, BAT1, HSP12, and TDH, whereas the higher nitrogen level caused higher expression levels for these genes. Furthermore, the lower nitrogen level resulted in increases in the levels of higher alcohols and esters at high wort gravity. All these results demonstrated that yeast physiology and flavor balance during beer brewing were significantly affected by the wort gravity and nitrogen level.

Fermentation kinetics and ethanol production from different sweet sorghum varieties.

Abstract: In recent years bioethanol fuels derived from agricultural biomass resources or waste have been considered the cleanest liquid fuel alternative to fossil fuels. ?Ethanol consumption is expected to reach 11.2 billion gallons by 2012. ?Sweet sorghum, grown in the Southeast and Midwest states in US, contains 14%-20% fermentable sugar and is an ideal feedstock, owing to its ease of fermentation by yeast. ?The objectives of this research were to perform kinetic studies to determine the factors that may affect the rates of sugar consumption and ethanol production during fermentation of two varieties of sweet sorghum juice, Variety 1 (V-1) and Variety 2 (V-2), and to optimize the fermentation efficiency and ethanol production by varying strategies to process the juice before fermentation. ?Kinetic parameters of fermentation provide total sugar consumption rate of 3.4 g/(L

Fed-batch fermentation for n-butanol production from cassava bagasse hydrolysate in a fibrous bed bioreactor with continuous gas stripping

Concentrated cassava bagasse hydrolysate (CBH) containing 584.4g/L glucose was studied for acetone–butanol–ethanol (ABE) fermentation with a hyper-butanol-producing Clostridiumacetobutylicum strain in a fibrous bed bioreactor with gas stripping for continuous butanol recovery. With periodical nutrient supplementation, stable production of n-butanol from glucose in the CBH was maintained in the fed-batch fermentation over 263h with an average sugar consumption rate of 1.28g/Lh and butanol productivity of 0.32±0.03g/Lh. A total of 108.5g/L ABE (butanol: 76.4g/L, acetone: 27.0g/L, ethanol: 5.1g/L) was produced, with an overall yield of 0.32±0.03g/g glucose for ABE and 0.23±0.01g/g glucose for butanol. The gas stripping process generated a product containing 10–16% (w/v) of butanol, ∼4% (w/v) of acetone, a small amount of ethanol (<0.8%) and almost no acids, resulting in a highly concentrated butanol solution of ∼64% (w/v) after phase separation.

The effect of sulfide and ammonia on cassava fermentation for ethanol production in an ethanol–methane coupled system

An ethanol-methane coupled system was proposed to resolve wastewater pollution in cassava ethanol production. The wastewater originated from ethanol distillation is treated with two-stage anaerobic digestion and then recycled for medium preparation for the next batch ethanol fermentation, thus eliminating wastewater discharge and saving fresh water. The constituents of the two-stage anaerobic digestion effluent were complex which influenced the ethanol fermentation performance. This paper aimed to study the effect of two constituents in the effluent, i.e. sulfide and ammonia, on cassava-based ethanol fermentation performance. It was found that sulfide reduced the consumption rate of total sugar by significantly inhibiting the growth of Saccharomyces cerevisiae, but the total consumption amount of total sugar at the end of fermentation was not influenced. S. cerevisiae produced more glycerol and less ethanol at the end of fermentation containing higher concentration of sodium sulfide. Ethanol fermentation performance could be hardly influenced by the sulfide in the two-stage effluent because of the very low concentration. More glycerol was produced while final ethanol concentration was reduced when the level of ammonia in the two-stage effluent was higher.

Effect of Co‐Inoculation of Saccharomyces cerevisiae and Lactobacillus fermentum on the Quality of the Distilled Sugar Cane Beverage Cachaça

The effect of Lactobacillus fermentum in co-inoculation with Saccharomyces cerevisiae UFLA CA11 on the quality of cachaça (sugar cane spirit) was evaluated. The co-inoculation was first evaluated in flask fermentation, and subsequently, yeast and bacteria were co-cultured at approximately 10(5) CFU/mL and 10(8) CFU/mL, respectively, in 4 consecutive batches. L. fermentum did not affect the growth or activity (sugar consumption and fermentation rate) of S. cerevisiae UFLA CA11 during fermentation. The physicochemical analysis revealed a higher concentration (Tukey test) of aldehydes (22.07 mg/100 mL anhydrous alcohol) in cachaça produced by co-inoculation. Analysis of volatile compounds using GC-FID demonstrated that cachaça produced by co-inoculation had higher concentrations of acetaldehyde (25.15 mg/L), ethyl acetate (30.17 mg/L), and 2,3-butanedione (170.39 μ/L), while cachaça produced by UFLA CA11 contained higher concentrations of ethyl lactate (1205.98 μ/L), propionic acid (127.97 μ/L), butyric acid (2335.57 μ/L), and 1-pentanol (469.23 μ/L). The lowest concentration of acetic acid measured by HPLC was found in cachaça obtained with UFLA CA11. The sensory analysis, performed using the Mann-Whitney test, revealed that cachaça produced by co-inoculation differed from that produced by UFLA CA11 in taste and aroma. This study reports on the use of a mixed culture of Saccharomyces cerevisae and Lactobacillus fermentum to produce cachaça and shows the influence of co-inoculation of yeast and bacteria on the quality of this beverage.

Effect of inhibitors formed during wheat straw pretreatment on ethanol fermentation by Pichia stipitis

The inhibitory effect of the main inhibitors (acetic acid, furfural and 5-hydroxymethylfurfural) formed during steam explosion of wheat straw was studied through ethanol fermentations of model substrates and hydrolysates from wheat straw by Pichia stipitis. Experimental results showed that an increase in acetic acid concentration led to a reduction in ethanol productivity and complete inhibition was observed at 3.5g/L. Furfural produced a delay on sugar consumption rates with increasing concentration and HMF did not exert a significant effect. Fermentations of the whole slurry from steam exploded wheat straw were completely inhibited by a synergistic effect due to the presence of 1.5g/L acetic acid, 0.15g/L furfural and 0.05g/L HMF together with solid fraction. When using only the solid fraction from steam explosion, hydrolysates presented 0.5g/L of acetic acid, whose fermentations have submitted promising results, providing an ethanol yield of 0.45g ethanol/g sugars and the final ethanol concentration reached was 12.2g/L (10.9g ethanol/100g DM).

PHB Biosynthesis in Catabolite Repression Mutant of Burkholderia sacchari

Due to the effect of catabolite repression, sugar mixtures cannot be metabolized in a rapid and efficient way implicating in lower productivity in bioprocesses using lignocellulosic hydrolysates. In gram-negative bacteria, this mechanism is mediated by the phosphotransferase system (PTS), which concomitantly internalizes and phosphorylates sugars. In this study, we isolated a UV mutant of Burkholderia sacchari, called LFM828, which transports hexoses and pentoses by a non-PTS uptake system. This mutant presented released glucose catabolite repression over the pentoses. In mixtures of glucose, xylose, and arabinose, specific growth rates and the specific sugar consumption rates were, respectively, 10 and 23% higher in LFM828, resulting in a reduced time to exhaust all sugars in the medium. However, in polyhydroxybutyrate (PHB) biosynthesis experiments it was necessary the supplementation of yeast extract to maintain higher values of growth rate and sugar consumption rate. The deficient growth in mineral medium was partially recovered by replacing the ammonium nitrogen source by glutamate. It was demonstrated that the ammonium metabolism is not defective in LFM828, differently from ammonium, glutamate can also be used as carbon and energy allowing an improvement on the carbohydrates utilization for PHB production in LFM828. In contrast, higher rates of ammonia consumption and CO(2) production in LFM828 indicate altered fluxes through the central metabolism in LFM828 and the parental. In conclusion, PTS plays an important role in cell physiology and the elimination of its components has a significant impact on catabolite repression, carbon flux distribution, and PHB biosynthesis in B. sacchari.

Effects of Aldehydes on the Growth and Lipid Accumulation of Oleaginous Yeast Trichosporon fermentans

The effects of five representative aldehydes in lignocellulosic hydrolysates on the growth and the lipid accumulation of oleaginous yeast Trichosporon fermentans were investigated for the first time. There was no relationship between the hydrophobicity and the toxicity of aldehyde, and 5-hydroxymethylfurfural was less toxic than aromatic aldehydes and furfural. Binary combination of aromatic aldehydes caused a synergistic inhibitory effect, but combination of furan and aromatic aldehydes reduced the inhibition instead. A longer lag phase was found due to the presence of aldehydes and the decrease of sugar consumption rate, but more xylose was utilized by T. fermentans in the presence of aldehydes, especially at their low concentrations. The variation of malic enzyme activity was not related to the delay of lipid accumulation. Furthermore, the inhibition of aldehydes on cell growth was more dependent on inoculum size, temperature, and initial pH than that on lipid content.

Identification of succinate exporter in Corynebacterium glutamicum and its physiological roles under anaerobic conditions

Corynebacterium glutamicum produces succinate from glucose via the reductive tricarboxylic acid cycle under microaerobic and anaerobic conditions. We identified a NCgl2130 gene of C. glutamicum as a novel succinate exporter that functions in succinate production, and designated sucE1. sucE1 expression levels were higher under microaerobic conditions than aerobic conditions, and overexpression or disruption of sucE1 respectively increased or decreased succinate productivity during fermentation. Under microaerobic conditions, the sucE1 disruptant sucE1Δ showed 30% less succinate productivity and a lower sugar-consumption rate than the parental strain. Under anaerobic conditions, succinate production by sucE1Δ ceased. The intracellular succinate and fructose-1,6-bisphosphate levels of sucE1Δ under microaerobic conditions were respectively 1.7-fold and 1.6-fold higher than those of the parental strain, suggesting that loss of SucE1 function caused a failure of succinate removal from the cells, leading to intracellular accumulation that inhibited upstream sugar metabolism. Homology and transmembrane helix searches identified SucE1 as a membrane protein belonging to the aspartate:alanine exchanger (AAE) family. Partially purified 6x-histidine-tagged SucE1 (SucE1-[His] 6) reconstituted in succinate-loaded liposomes clearly demonstrated counterflow and self-exchange activities for succinate. Together, these findings suggest that sucE1 encodes a novel succinate exporter that is induced under microaerobic conditions, and is important for succinate production under both microaerobic and anaerobic conditions.

Cellulolytic Enzymes Production by Solid State Culture

Problem Statement: Great interest in the use of lignocellulosic biomass is increasing in order to diminish the accumulation of residues, such as pecan nut shells. One of the alternatives is the fungal degradation of these residues. Approach: The capacity of Trichoderma (coded as T1, T2 and T3) strains to produce cellulase and xylonite was evaluated. Results: Pecan nut shell fibers were used as sole carbon source. The fiber characterization study showed that cellulose levels were of 0.1% while hemicellulose was up to 25 %. Three Trichoderma strains were used on solid fungal cultures using the fibers as sole carbon and inductor source for the production of cellulolytic enzymes. The behavior of the sugars liberated by the fungi showed that the strain T2 is able to accumulate more monomeric reducing sugars than the other two strains, this could be attributed at this strain has a higher sugar liberation rate and slower sugar consumption rate. This strain also expressed more cellulase and xylanase activity. The low quantity of cellulose registered in the fibers can still be used to induce cellulase activity. Conclusion: The T2 strain had the highest level of enzymatic activity both cellulase and xylanase.

Identification of mannose uptake and catabolism genes in Corynebacterium glutamicum and genetic engineering for simultaneous utilization of mannose and glucose

Here, focus is on Corynebacterium glutamicum mannose metabolic genes with the aim to improve this industrially important microorganism's ability to ferment mannose present in mixed sugar substrates. cgR_0857 encodes C. glutamicum's protein with 36% amino acid sequence identity to mannose 6-phosphate isomerase encoded by manA of Escherichia coli. Its deletion mutant did not grow on mannose and exhibited noticeably reduced growth on glucose as sole carbon sources. In effect, C. glutamicum manA is not only essential for growth on mannose but also important in glucose metabolism. A double deletion mutant of genes encoding glucose and fructose permeases (ptsG and ptsF, respectively) of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was not able to grow on mannose unlike the respective single deletion mutants with mannose utilization ability. A mutant deficient in ptsH, a general PTS gene, did not utilize mannose. These indicate that the glucose-PTS and fructose-PTS are responsible for mannose uptake in C. glutamicum. When cultured with a glucose and mannose mixture, mannose utilization of manA-overexpressing strain CRM1 was significantly higher than that of its wild-type counterpart, but with a strong preference for glucose. ptsF-overexpressing strain CRM2 co-utilized mannose and glucose, but at a total sugar consumption rate much lower than that of the wild-type strain and CRM1. Strain CRM3 overexpressing both manA and ptsF efficiently co-utilized mannose and glucose. Under oxygen-deprived conditions, high volumetric productivity of organic acids concomitant with the simultaneous consumption of the mixed sugars was achieved by the densely packed growth-arrested CRM3 cells.

Increased biomass production of industrial bakers' yeasts by overexpression of Hap4 gene

HAP4 encodes a transcriptional activator of respiration-related genes and so, redirection from fermentation to respiration flux should give rise to an increase in biomass production in Saccharomyces cerevisiae transformants that overexpress HAP4. With this aim, three bakers' yeasts, that is, V1 used for lean doughs, its 2-deoxy-D-glucose resistant derivative DOG21, and V3 employed for sweet doughs, were transformed with integrative cassettes that carried HAP4 gene under the control of constitutive promoter pTEF2; in addition VTH, DTH and 3TH transformants were selected and characterized. Transformants showed increased expression of HAP4 and respiration-related genes such as QCR7 and QCR8 with regard to parental, and similar expression of SUC2 and MAL12; these genes are relevant in bakers' industry. Invertase (Suc2p) and maltase (Mal12p) activities, growth and sugar consumption rates in laboratory (YPD) or industrial media (MAB) were also comparable in bakers' strains and their transformants, but VTH, DTH and 3TH increased their final biomass production by 9.5, 5.0 and 5.0% respectively as compared to their parentals in MAB. Furthermore, V1 and its transformant VTH had comparable capacity to ferment lean doughs (volume increase rate and final volume) while V3 and its transformant 3TH fermented sweet doughs in a similar manner. Therefore transformants possessed increased biomass yield and appropriate characteristics to be employed in bakers' industry because they lacked drug resistant markers and bacterial DNA, and were genetically stable.

Efficient production of recombinant IgG by metabolic control and co-expression with GLUT5 in a fructose-based medium

A fructose-based cell culture is suitable for the process control of protein production because of slow sugar consumption rate and low lactate accumulation. The fructose transporter, GLUT5, mediates its incorporation into cells and is required for the fructose-based culture. In order to produce efficiently recombinant IgG by metabolic control and co-expression with GLUT5 in a fructose-based medium, an IgG and GLUT5 co-expression vector was constructed and transfected into the human myeloma derived cell line, SC-01MFP, which produced stably recombinant proteins. The cell proliferation in the fructose-based medium was improved by the GLUT5 gene transfection. The recombinant IgG production of the cells cultured in the fructose-based medium exhibited about two-fold increase of that in the glucose-based medium. Flow cytometoric analysis indicated that the GLUT5 protein expression level in cell surface was increased in the fructose-based medium. An exogenous but not endogenous GLUT5 transcription activator remarkably raised IgG productivity in the fructose-based medium when compared to that in the glucose-based medium, suggesting that exogenous GLUT5 expression may be involved in it. The GLUT5 co-expression system may be useful for efficient production of recombinant proteins by the fructose-based cell culture.

Regulation of meilingmycin in Streptomyces nanchangensis: Effect of ammonium ion

Effects of different ammonium sulfate concentrations on meilingmycin biosynthesis were studied in this research. The results show that a lower concentration of ammonium ions stimulates the biosynthesis of meilingmycin, while a concentration higher than 5 mmol/L inhibits the mycelial growth and the biosynthesis of the products. However, increased sugar consumption rate with the elevated concentration of ammonium sulfate was observed during the fermentation process. On this basis, six enzymes, which are responsible for the meilingmycin biosynthesis and the glucose metabolism, were measured and analyzed during the bioprocess. The results suggest that glucose-6-phosphate dehydrogenase, citrate synthase, succinate dehydrogenase and fatty acid synthase are stimulated by higher concentration of ammonium ions, while valine dehydrogenase and methylmalonyl-CoA carboxyltransferase are inhibited. From the results it follows that the precursor supply was restricted with the higher concentration of ammonium ions, which results in the lower production of meilingmycin. Thus, the strategy of maintaining a low level of FAS activity is a critical factor for high yield of meilingmycin.

Simultaneous utilization of d-cellobiose, d-glucose, and d-xylose by recombinant Corynebacterium glutamicum under oxygen-deprived conditions

Corynebacterium glutamicum R was metabolically engineered to broaden its sugar utilization range to D-xylose and D-cellobiose contained in lignocellulose hydrolysates. The resultant recombinants expressed Escherichia coli xylA and xylB genes, encoding D-xylose isomerase and xylulokinase, respectively, for D-xylose utilization and expressed C. glutamicum R bglF317A and bglA genes, encoding phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) beta-glucoside-specific enzyme IIBCA component and phospho-beta-glucosidase, respectively, for D-cellobiose utilization. The genes were fused to the non-essential genomic regions distributed around the C. glutamicum R chromosome and were under the control of their respective constitutive promoter trc and tac that permitted their expression even in the presence of D-glucose. The enzyme activities of resulting recombinants increased with the increase in the number of respective integrated genes. Maximal sugar utilization was realized with strain X5C1 harboring five xylA-xylB clusters and one bglF317A-bglA cluster. In both D-cellobiose and D-xylose utilization, the sugar consumption rates by genomic DNA-integrated strain were faster than those by plasmid-bearing strain, respectively. In mineral medium containing 40 g l(-1) D-glucose, 20 g l(-1) D-xylose, and 10 g l(-1) D-cellobiose, strain X5C1 simultaneously and completely consumed these sugars within 12 h and produced predominantly lactic and succinic acids under growth-arrested conditions.

Expression of a heterologous xylose transporter in a Saccharomyces cerevisiae strain engineered to utilize xylose improves aerobic xylose consumption

The goal of this investigation was to determine the effect of a xylose transport system on glucose and xylose co-consumption as well as total xylose consumption in Saccharomyces cerevisiae. We expressed two heterologous transporters from Arabidopsis thaliana in recombinant xylose-utilizing S. cerevisiae cells. Strains expressing the heterologous transporters were grown on glucose and xylose mixtures. Sugar consumption rates and ethanol concentrations were determined and compared to an isogenic control strain lacking the A. thaliana transporters. Expression of the transporters increased xylose uptake and xylose consumption up to 46% and 40%, respectively. Xylose co-consumption rates (prior to glucose depletion) were also increased by up to 2.5-fold compared to the control strain. Increased xylose consumption correlated with increased ethanol concentration and productivity. During the xylose/glucose co-consumption phase, strains expressing the transporters had up to a 70% increase in ethanol production rate. It was concluded that in these strains, xylose transport was a limiting factor for xylose utilization and that increasing xylose/glucose co-consumption is a viable strategy for improving xylose fermentation.

Cellulosic ethanol production using the naturally occurring xylose-fermenting yeast, Pichia stipitis

Rising crude oil prices and environmental concerns have renewed interest in renewable energy. Cellulosic ethanol promises to deliver a renewable fuel from non-food feedstocks. One technical challenge producing cellulosic ethanol economically is a robust organism to utilize the different sugars present in cellulosic biomass. Unlike starch where glucose is the only sugar present, cellulosic biomass has other sugars such as xylose and arabinose, usually called C5 sugars. This review examines the most promising naturally occurring C5 fermenting organism, Pichia stipitis. In this work, the properties that make P. stipitis unique from other organisms, its physiology and fermentation results on lignocellulosic substrates have been reviewed. P. stipitis can produce 41 g ethanol/l with a potential to cleanup some of the most concentrated toxins. These results coupled with the less stringent nutritional requirements, great resistance to contamination and its thick cell walls makes P. stipitis a viable organism for scale-up. However, P. stipitis has a slower sugar consumption rate compared to Saccharomyces cerevisiae and requires microaerophilic condition for ethanol production. Finally, future studies to enhance fermentation capabilities of this yeast have been discussed.