Improving the performance of industrial ethanol‐producing yeast by expressing the aspartyl protease on the cell surface

The yeasts used in fuel ethanol manufacture are unable to metabolize soluble proteins. The PEP4 gene, encoding a vacuolar aspartyl protease in Saccharomyces cerevisiae, was either secretively or cell‐surface anchored expressed in industrial ethanol‐producing S. cerevisiae. The obtained recombinant strains APA (expressing the protease secretively) and APB (expressing the protease on the cell wall) were studied under ethanol fermentation conditions in feed barley cultures. The effects of expression of the protease on product formation, growth and cell protein content were measured. The biomass yield of the wild‐type was clearly lower than that of the recombinant strains (0.578 ± 0.12 g biomass/g glucose for APA and 0.582 ± 0.08 g biomass/g glucose for APB). In addition, nearly 98–99% of the theoretical maximum level of ethanol yield was achieved (relative to the amount of substrate consumed) for the recombinant strains, while limiting the nitrogen source resulted in dissatisfactory fermentation for the wild‐type and more than 30 g/l residual sugar was detected at the end of fermentation. In addition, higher growth rate, viability and lower yields of byproducts such as glycerol and pyruvic acid for recombinant strains were observed. Expressing acid protease can be expected to lead to a significant increase in ethanol productivity. Copyright © 2010 John Wiley & Sons, Ltd.     

The glycerol and ethanol production kinetics in low-temperature wine fermentation using Saccharomyces cerevisiae yeast strains

Increasing glycerol production in low-temperature wine fermentation is of concern for winemakers to improve the quality of wines. The objective of this study was to investigate the effect of 10 different Saccharomyces cerevisiae on the kinetics of production of glycerol, ethanol and the activities of glycerol-3-phosphate dehydrogenase (GPD) and alcohol dehydrogenase (ADH) in low-temperature fermentation. Ethanol production was influenced by temperature, and it was slightly higher at 13 °C than at 25 °C. Glycerol yields were significantly affected by both temperature and strains. More glycerol was produced at 25 °C than at 13 °C because the activity of GPD was higher at 25 °C than at 13 °C. Glycerol production of the different yeast strains was up to 3.19 and 3.18 g L⁻¹ at 25 and 13 °C, respectively. Therefore, isolating the yeast strains with high glycerol production and adaptation to low-temperature fermentation is still the best method in winemaking.     

Modulation of Glycerol and Ethanol Yields During Alcoholic Fermentation in Saccharomyces cerevisiae Strains Overexpressed or Disrupted for GPD1 Encoding Glycerol 3-Phosphate Dehydrogenase

The possibility of the diversion of carbon flux from ethanol towards glycerol in Saccharomyces cerevisiae during alcoholic fermentation was investigated. Variations in the glycerol 3-phosphate dehydrogenase (GPDH) level and similar trends for alcohol dehydrogenase (ADH), pyruvate decarboxylase and glycerol-3-phosphatase were found when low and high glycerol-forming wine yeast strains were compared. GPDH is thus a limiting enzyme for glycerol production. Wine yeast strains with modulated GPD1 (encoding one of the two GPDH isoenzymes) expression were constructed and characterized during fermentation on glucose-rich medium. Engineered strains fermented glucose with a strongly modified [glycerol] : [ethanol] ratio. gpd1Δ mutants exhibited a 50% decrease in glycerol production and increased ethanol yield. Overexpression of GPD1 on synthetic must (200 g/l glucose) resulted in a substantial increase in glycerol production (×4) at the expense of ethanol. Acetaldehyde accumulated through the competitive regeneration of NADH via GPDH. Accumulation of by-products such as pyruvate, acetate, acetoin, 2,3 butane-diol and succinate was observed, with a marked increase in acetoin production. © 1997 John Wiley & Sons, Ltd.     

Fermentation properties of a wine yeast over-expressing the Saccharomyces cerevisiae glycerol 3-phosphate dehydrogenase gene (GPD2)

Wine yeasts efficiently convert sugar into ethanol. The possibility of diverting some of the sugar into compounds other than ethanol by using molecular genetic methods was tested. Over-expression of the yeast glycerol 3-phosphate dehydrogenase gene ( GPD2 ) in a laboratory strain of Saccharomyces cerevisiae led to an approximate two-fold increase in the extracellular glycerol concentration. In the medium fermented with the modified strain, acetic acid concentration also increased approximately two-fold when respiration was blocked. A strain deleted for the GPD2 gene had the opposite phenotype, producing lower amounts of glycerol and acetic acid, with the latter compound only reduced during non-respiratory growth. A commercial wine yeast over-expressing GPD2 produced 16.5 g/L glycerol in a wine fermentation, compared to 7.9 g/L obtained with the parent strain. As seen for the laboratory strain, acetic acid concentrations were also increased when using the genetically modified wine yeast. A panel of wine judges confirmed the increase in volatile acidity of these wines. The altered glycerol biosynthetic pathway sequestered carbon from glycolysis and reduced the production of ethanol by 6 g/L.     

The importance of the glycerol 3-phosphate shuttle during aerobic growth of Saccharomyces cerevisiae

Maintenance of a cytoplasmic redox balance is a necessity for sustained cellular metabolism. Glycerol formation is the only way by which Saccharomyces cerevisiae can maintain this balance under anaerobic conditions. Aerobically, on the other hand, several different redox adjustment mechanisms exist, one of these being the glycerol 3-phosphate (G3P) shuttle. We have studied the importance of this shuttle under aerobic conditions by comparing growth properties and glycerol formation of a wild-type strain with that of gut2Δ mutants, lacking the FAD-dependent glycerol 3-phosphate dehydrogenase, assuming that the consequent blocking of G3P oxidation is forcing the cells to produce glycerol from G3P. To impose different demands on the redox adjustment capability we used various carbon sources having different degrees of reduction. The results showed that the shuttle was used extensively with reduced substrate such as ethanol, whereas the more oxidized substrates lactate and pyruvate, did not provoke any activity of the shuttle. However, the absence of a functional G3P shuttle did not affect the growth rate or growth yield of the cells, not even during growth on ethanol. Presumably, there must be alternative systems for maintaining a cytoplasmic redox balance, e.g. the so-called external NADH dehydrogenase, located on the outer side of the inner mitochondrial membrane. By comparing the performance of the external NADH dehydrogenase and the G3P shuttle in isolated mitochondria, it was found that the former resulted in high respiratory rates but a comparably low P/O ratio of 1·2, whereas the shuttle gave low rates but a high P/O ratio of 1·7. Our results also demonstrated that of the two isoforms of NAD-dependent glycerol 3-phosphate dehydrogenase, only the enzyme encoded by GPD1 appeared important for the shuttle, since the enhanced glycerol production that occurs in a gut2Δ strain proved dependent on GPD1 but not on GPD2. © 1998 John Wiley & Sons, Ltd.     

Reduced pyruvate decarboxylase and increased glycerol-3-phosphate dehydrogenase [NAD+] levels enhance glycerol production in Saccharomyces cerevisiae

This investigation deals with factors affecting the production of glycerol in Saccharomyces cerevisiae. In particular, the impact of reduced pyruvate-decarboxylase (PDC) and increased NAD-dependent glycerol-3-phosphate dehydrogenase (GPD) levels was studied. The glycerol yield was 4·7 times (a pdc mutant exhibiting 19% of normal PDC activity) and 6·5 times (a strain exhibiting 20-fold increased GPD activity resulting from overexpression of GPD1 gene) that of the wild type. In the strain carrying both enzyme activity alterations, the glycerol yield was 8·1 times higher than that of the wild type. In all cases, the substantial increase in glycerol yield was associated with a reduction in ethanol yield and a higher by-product formation. The rate of glycerol formation in the pdc mutant was, due to a slower rate of glucose catabolism, only twice that of the wild type, and was increased by GPD1 overexpression to three times that of the wild-type level. Overexpression of GPD1 in the wild-type background, however, led to a six- to seven-fold increase in the rate of glycerol formation. The experimental work clearly demonstrates the rate-limiting role of GPD in glycerol formation in S. cerevisiae.     

Using crude glycerol and thin stillage for the production of microbial lipids through the cultivation of Rhodotorula glutinis

Single cell oils (SCO) produced from oleaginous microorganisms are a potential alternative oil feedstock for biodiesel production. The worldwide production of glycerol, a 10% (w/w) byproduct produced in the transesterfication process of oils converted to biodiesel, is increasing as more biodiesel is being produced. For the purposes of cost reduction, crude glycerol was regarded as a suitable carbon source for the cultivation of Rhodotorula glutinis. In addition to using renewable crude glycerol, waste solution collected from the brewing company (called thin stillage) was adopted as a substitute to replace a costly nitrogen source used in the medium. The results of using mixture of crude glycerol and thin stillage indicated about a 27% increase in total biomass as compared to that of using crude glycerol with a standard medium. Using glycerol instead of glucose as the carbon source could also alter the lipid profile, resulting in an increase in linolenic acid (C18:2) to comprise over 20% of the total lipid. Successfully using renewable crude glycerol and thin stillage for the cultivation of oleaginous microorganisms could greatly enhance the economic competition of biodiesel produced from SCO.     

Effect of dilution rate and nutrients addition on the fermentative capability and synthesis of aromatic compounds of two indigenous strains of Saccharomyces cerevisiae in continuous cultures fed with Agave tequilana juice

Knowledge of physiological behavior of indigenous tequila yeast used in fermentation process is still limited. Yeasts have significant impact on the productivity fermentation process as well as the sensorial characteristics of the alcoholic beverage. For these reasons a better knowledge of the physiological and metabolic features of these yeasts is required. The effects of dilution rate, nitrogen and phosphorus source addition and micro-aeration on growth, fermentation and synthesis of volatile compounds of two native Saccharomyces cerevisiae strains, cultured in continuous fed with Agave tequilana juice were studied. For S1 and S2 strains, maximal concentrations of biomass, ethanol, consumed sugars, alcohols and esters were obtained at 0.04 h⁻¹. Those concentrations quickly decreased as D increased. For S. cerevisiae S1 cultures (at D = 0.08 h⁻¹) supplemented with ammonium phosphate (AP) from 1 to 4 g/L, concentrations of residual sugars decreased from 29.42 to 17.60 g/L and ethanol increased from 29.63 to 40.08 g/L, respectively. The S1 culture supplemented with AP was then micro-aerated from 0 to 0.02 vvm, improving all the kinetics parameters: biomass, ethanol and glycerol concentrations increased from 5.66, 40.08 and 3.11 g/L to 8.04, 45.91 and 4.88 g/L; residual sugars decreased from 17.67 g/L to 4.48 g/L; and rates of productions of biomass and ethanol, and consumption of sugars increased from 0.45, 3.21 and 7.33 g/L·h to 0.64, 3.67 and 8.38 g/L·h, respectively. Concentrations of volatile compounds were also influenced by the micro-aeration rate. Ester and alcohol concentrations were higher, in none aerated and in aerated cultures respectively.     

Effects of altering fermentation parameters on glycerol and bioethanol production from cane molasses

Current bioethanol production processes do not produce more than approximately 13 g l^?1 glycerol at a purity of 11% total solids in the fermented mash. Such quantities are not sufficiently high for economic exploitation unless very‐large‐capacity plants are involved. It was envisaged that the ratio of glycerol to bioethanol could be altered in favour of glycerol by adjusting such fermentation parameters as osmotic pressure (water activity), pH, temperature and yeast cell inoculum. At 39.9 ° brix, pH 8.7, 38 °C, 90.7 atm and 120 × 10⁶ yeast cells ml^?1, glycerol formation in fermented mash increased to 56 g l^?1 at a purity of 14% total solids. The potential for high glycerol production, hence higher revenue, has been demonstrated. Adoption of this technology could have a significant positive economic impact on existing or new biomass‐to‐ethanol plants. © 2002 Society of Chemical Industry     

Xylitol production on sugarcane biomass hydrolysate by newly identified Candida tropicalis JA2 strain

Xylitol is a building block for a variety of chemical commodities, besides being widely used as a sugar substitute in the food and pharmaceutical industries. The aim of this work was to develop a microbial process for xylitol production using sugarcane bagasse hydrolysate as substrate. In this context, 218 non-Saccharomyces yeast strains were screened by growth on steam-exploded sugarcane bagasse hydrolysate containing a high concentration of acetic acid (8.0 g/L). Seven new Candida tropicalis strains were selected and identified, and their ability to produce xylitol on hydrolysate at low pH (4.6) under aerobic conditions was evaluated. The most efficient strain, designated C. tropicalis JA2, was capable of producing xylitol with a yield of 0.47 g/g of consumed xylose. To improve xylitol production by C. tropicalis JA2, a series of experimental procedures were employed to optimize pH and temperature conditions, as well as nutrient source, and initial xylose and inoculum concentrations. C. tropicalis JA2 was able to produce 109.5 g/L of xylitol with a yield of 0.86 g/g of consumed xylose, and with a productivity of 2.81 g·L·h, on sugarcane bagasse hydrolysate containing 8.0 g/L acetic acid and177 g/L xylose, supplemented with 2.0 g/L yeast nitrogen base and 4.0 g/L urea. Thus, it was possible to identify a new C. tropicalis strain and to optimize the xylitol production process using sugarcane bagasse hydrolysate as a substrate. The xylitol yield on biomass hydrolysate containing a high concentration of acetic acidobtained in here is among the best reported in the literature.     

碳源对圆酵母(Torula sp.)B84512发酵合成赤藓糖醇及副产物甘油的影响

研究了圆酵母(Torula sp.)B84512以不同碳源发酵产赤藓糖醇过程中副产物甘油的生成与消耗情况。发现该菌株在以任何碳源为底物发酵过程中均会产生甘油,且在发酵中后期甘油逐渐被消耗。以甘油为唯一碳源时该菌株合成赤藓糖醇的速率及产率均低于葡萄糖。葡萄糖为圆酵母B84512发酵产赤藓糖醇的最佳碳源。采用分批补料的方式提高赤藓糖醇的产率并期望能抑制甘油的生成,实验结果表明补料至总糖浓度为50%时赤藓糖醇产量最高为253 g/L,产率为1.03 g/(L.h)。但甘油产量与葡萄糖的浓度呈正相关,分批补料并不能有效抑制甘油的生成,反而导致发酵周期大大延长,对于工业化生产极其不利。通过对甘油的生成及消耗过程中关键酶胞浆3-磷酸甘油脱氢酶(ctGPD)、3-磷酸甘油酯酶(GPP)、线粒体3-磷酸甘油脱氢酶(mtGPD)酶活测定,确定胞浆3-磷酸甘油脱氢酶为甘油合成途径的关键酶,为以后对圆酵母B84512中甘油代谢途径的基因工程改造选育奠定了基础。     

Effects of the addition of direct-fed microbials and glycerol to the diet of lactating dairy cows on milk yield and apparent efficiency of yield

A study was conducted to evaluate the effects of a direct-fed microbial (M) and dietary glycerol (G) on milk yield, efficiency of yield, and nutrient digestibility during hot weather. Sixty Holstein cows averaging 120 d in milk (DIM) and 36.2 kg/d of milk were used in a 12-wk 2 × 2 factorial design trial from June through September 2008. Cows were fed a common diet during the 2-wk standardization period and were blocked by milk yield, DIM, parity, and dry matter intake. Diets were based on corn and ryegrass silages and balanced to be isocaloric and isonitrogenous. Treatments included a negative control (M− or G−), 4 × 10⁹ cfu/head of a combination of Lactobacillus acidophilus NP51 and Propionibacterium freudenreichii NP24 (M+), control plus 400 g/h per day of 99% pure food-grade glycerol (G+), and 4 × 10⁹ cfu/h per day of a combination of Lactobacillus acidophilus NP51 and Propionibacterium freudenreichii NP24 plus 400 g/h per day of 99% pure food-grade glycerol (MG++). No interactions were observed between direct-fed microbials and dietary glycerol in the study except on apparent nutrient digestibility. No differences were observed in dry matter intake, which averaged 22.7, 23.1, 23.4, and 22.9 for M−, G−, M+, and G+, respectively. Milk yield was increased for M+ compared with M− at 34.1 and 31.7 kg/d, but G+ had no effect on yield. No treatment effect was noted for milk fat percentage or milk protein percentage among diets. Milk protein yield was higher for M+ compared with M− at 0.93 versus 0.87 kg/d. Energy-corrected milk was improved for the M+ versus M− groups at 33.5 and 31.6 kg/d, respectively. No differences in respiratory rate, skin temperature, body temperature, or concentrations of serum glucose or urea N were observed among treatments. Improvement in apparent digestibility was observed with M+ and G+ compared with M−/G− in this experiment. The addition of a direct-fed microbial alone improved milk and protein yield, energy-corrected milk, and apparent digestibility of crude protein, neutral detergent fiber, and acid detergent fiber, and the inclusion of glycerol (G+) had a positive effect on apparent dry matter and acid detergent fiber digestibility compared with M−/G−. The addition of a direct-fed microbial and dietary glycerol may improve yield and digestibility for cows subject to heat stress.     

Modulation of the glycerol and ethanol syntheses in the yeast Saccharomyces kudriavzevii differs from that exhibited by Saccharomyces cerevisiae and their hybrid

In the last years there is an increasing demand to produce wines with higher glycerol levels and lower ethanol contents. The production of these compounds by yeasts is influenced by many environmental variables, and could be controlled by the choice of optimized cultivation conditions. The present work studies, in a wine model system, the effects of temperature, pH and sugar concentration on the glycerol and ethanol syntheses by yeasts Saccharomyces cerevisiae T73, the type strain of Saccharomyces kudriavzevii IFO 1802^T, and an interspecific hybrid between both species (W27), which was accomplished by the application of response surface methodology based in a central composite circumscribed design. Results show that carbon flux could be especially directed towards glycerol synthesis instead of ethanol at low pH, high sugar concentrations and low temperatures. In general, the non-wine yeast S. kudriavzevii produced higher glycerol levels and lower ethanol content than wine strains S. cerevisiae T73 and the hybrid W27, with specific and different glycerol production profiles as a function of temperature and pH. These results were congruent with the higher glycerol-3-phosphate dehydrogenase activities estimated for this species, chiefly at low temperatures (14 °C), which could explain why S. kudriavzevii is a cryotolerant yeast compared to S. cerevisiae.     

Identification and characterization of cytosolic Hansenula polymorpha proteins belonging to the Hsp70 protein family

We have isolated two members of the Hsp70 protein family from the yeast Hansenula polymorpha using affinity chromatography. Both proteins were located in the cytoplasm. One of these, designated Hsp72, was inducible in nature (e.g. by heat shock). The second protein (designated Hsc74) was constitutively present. Peptides derived from both Hsp72 and Hsc74 showed sequence homology to the cytosolic Saccharomyces cerevisiae Hsp70s, Ssa1p and Ssa2p. The gene encoding Hsp72 (designated HSA1) was cloned, sequenced and used to construct HSA1 disruption and HSA1 overexpression strains. Comparison of the stress tolerances of these strains with those of wild-type H. polymorpha revealed that HSA1 overexpression negatively affected the tolerance of the cells to killing effects of temperature or ethanol, but enhanced the tolerance to copper and cadmium. The tolerance for other chemicals (arsenite, arsenate, H₂O₂) or to high osmolarity was unaffected by either deletion or overexpression of HSA1. The nucleotide sequence of HSA1 was submitted to the EMBL data library and given the Accession Number Z29379.     

Green recovery of phenolic compounds from rice byproduct (rice bran) using glycerol based on viscosity,conductivity and density

Rice bran, the industry byproduct, contains high levels of phenolic compounds. Based on glycerol, a green recovery of phenolics from rice bran was proposed. Extraction temperature (X₁), glycerol concentration (X₂) and liquid-to-solid ratio (X₃) were optimised by response surface methodology. Basic physical properties, that is, viscosity, conductivity, density were analysed and UPLC-Triple-TOF/MS method was applied for phenolic identification. Results showed the optimum conditions were X₁ = 66.76 °C, X₂ = 19.47% and X₃ = 32.92 mL g⁻¹. Under these conditions, the recovery yield in total polyphenols was 708.58 ± 12.36 mg GAE/100 g DW. Moreover, the recovery capacity of phenolic compounds was significantly correlated with viscosity, conductivity and density of glycerol solution. Under the optimal conditions, totals of thirteen phenolic compounds were identified and phenolic acids (4-hydroxybenzoic acid, vanillic acid, syringic acid, 4-coumaric acid, ferulic acid and salicylic acid) were quantified. These results demonstrate glycerol can serve as an ideal green solvent to recover phenolic compounds from rice bran.     

Fermentative behavior of Saccharomyces strains during microvinification of raspberry juice (Rubus idaeus L.)

Sixteen different strains of Saccharomyces cerevisiae and Saccharomyces bayanus were evaluated in the production of raspberry fruit wine. Raspberry juice sugar concentrations were adjusted to 16°Brix with a sucrose solution, and batch fermentations were performed at 22 °C. Various kinetic parameters, such as the conversion factors of the substrates into ethanol (Y_p/s), biomass (Y_x/s), glycerol (Y_g/s) and acetic acid (Y_ac/s), the volumetric productivity of ethanol (Q_p), the biomass productivity (P_x), and the fermentation efficiency (E_f) were calculated. Volatile compounds (alcohols, ethyl esters, acetates of higher alcohols and volatile fatty acids) were determined by gas chromatography (GC-FID). The highest values for the E_f, Y_p/s, Y_g/s, and Y_x/s parameters were obtained when strains commonly used in the fuel ethanol industry (S. cerevisiae PE-2, BG, SA, CAT-1, and VR-1) were used to ferment raspberry juice. S. cerevisiae strain UFLA FW 15, isolated from fruit, displayed similar results. Twenty-one volatile compounds were identified in raspberry wines. The highest concentrations of total volatile compounds were found in wines produced with S. cerevisiae strains UFLA FW 15 (87,435 μg/L), CAT-1 (80,317.01 μg/L), VR-1 (67,573.99 μg/L) and S. bayanus CBS 1505 (71,660.32 μg/L). The highest concentrations of ethyl esters were 454.33 μg/L, 440.33 μg/L and 438 μg/L for S. cerevisiae strains UFLA FW 15, VR-1 and BG, respectively. Similar to concentrations of ethyl esters, the highest concentrations of acetates (1927.67 μg/L) and higher alcohols (83,996.33 μg/L) were produced in raspberry wine from S. cerevisiae UFLA FW 15. The maximum concentration of volatile fatty acids was found in raspberry wine produced by S. cerevisiae strain VR-1. We conclude that S. cerevisiae strain UFLA FW 15 fermented raspberry juice and produced a fruit wine with low concentrations of acids and high concentrations of acetates, higher alcohols and ethyl esters.     

A pilot-scale evaluation of using gaseous chlorine dioxide for decontamination of foodborne pathogens on produce and low-moisture foods

Small-scale studies have shown that chlorine dioxide gas, ClO_2(g), was effective for decontamination of produce, nuts, and spices. This study conducted a pilot-scale evaluation to identify effective ClO_2(g) treatment parameters for commercial-scale applications. The gas was produced by a generator utilizing sodium chlorite and chlorine gas for decontamination of Shiga toxin-producing Escherichia coli (STEC), Listeria monocytogenes, and Salmonella inoculated on tomatoes, blueberries, baby-cut carrots, almonds, and peppercorns. Inoculated samples and 45 kg tomatoes in a 1,246-L treatment chamber were exposed to various ClO_2(g) concentrations (mg/L) and times 9 (hr) at 70–95% RH to determine the treatment effects on the pathogen reductions. Results showed that the treatment caused higher reductions on produce. A ClO_2(g) treatment of 1 mg/L-3 hr at 70% RH reduced 4.9–6.8, 5.1–5.6, and 4.2–6.3 log CFU/g of STEC, L. monocytogenes, and Salmonella, respectively, on produce, with the highest reductions on baby-cut carrots. For almonds and peppercorns, ClO_2(g) treatments under higher RH caused higher reductions. The treatment of 2 mg/L-9 hr or 3 mg/L-4 hr at 95% RH reduced >4.0 log of STEC and Salmonella on almonds, and 1 mg/L-5 hr at 85% RH achieved >5.0 log reductions on peppercorns. Applying moisture to the surfaces of almonds caused >4.0 log reductions using 1 mg/L-5 hr at 95% RH. This study identified effective ClO_2(g) treatment parameters for achieving >4.0 log reductions of common pathogens on tomatoes, blueberries, baby-cut carrots, almonds, and peppercorns and showed that ClO_2(g) generator is suitable for large-scale decontamination. These findings can be used for pilot-scale ClO_2(g) decontamination of these products and for testing using ClO_2(g) for commercial-scale decontamination trials.     

Comparison of bioethanol and beta‐galactosidase production by Kluyveromyces and Saccharomyces strains grown in cheese whey

The production of ethanol and beta‐galactosidase by Kluyveromyces marxianus and Saccharomyces fragilis strains grown in cheese whey was evaluated. The conditions for fermentation in 50 g/L (3.3% lactose) and 150 g/L (8.8% lactose) cheese whey were 100 rpm for 24 h at 30, 35 and 40 °C. Saccharomyces fragilis IZ 275 in 8.8% lactose at 40 °C resulted in 3.90% ethanol. Kluyveromyces marxianus CCT 3172 showed higher beta‐galactosidase, 1.10 U/mg, at 30 °C. Therefore, the choice of cultivation conditions and the most suitable species is important for obtaining high yields of the products of interest.     

Fermentation profiles of the yeast Brettanomyces bruxellensis in d-xylose and l-arabinose aiming its application as a second-generation ethanol producer

The yeast Brettanomyces bruxellensis is able to ferment the main sugars used in first-generation ethanol production. However, its employment in this industry is prohibitive because the ethanol productivity reached is significantly lower than the observed for Saccharomyces cerevisiae. On the other hand, a possible application of B. bruxellensis in the second-generation ethanol production has been suggested because this yeast is also able to use d -xylose and l -arabinose, the major pentoses released from lignocellulosic material. Although the latter application seems to be reasonable, it has been poorly explored. Therefore, we aimed to evaluate whether or not different industrial strains of B. bruxellensis are able to ferment d -xylose and l -arabinose, both in aerobiosis and oxygen-limited conditions. Three out of nine tested strains were able to assimilate those sugars. When in aerobiosis, B. bruxellensis cells exclusively used them to support biomass formation, and no ethanol was produced. Moreover, whereas l -arabinose was not consumed under oxygen limitation, d -xylose was only slightly used, which resulted in low ethanol yield and productivity. In conclusion, our results showed that d -xylose and l -arabinose are not efficiently converted to ethanol by B. bruxellensis, most likely due to a redox imbalance in the assimilatory pathways of these sugars. Therefore, despite presenting other industrially relevant traits, the employment of B. bruxellensis in second-generation ethanol production depends on the development of genetic engineering strategies to overcome this metabolic bottleneck.     

Amino acid and fatty acid compositions and nutritional quality of muscle in the pomfret, Pampus punctatissimus

The pomfret, Pampus punctatissimus, is an important fisheries resource in China, but little is known about its amino acid and fatty acid compositions. Pomfret muscle contained 18.6% crude protein and 4.95% crude fat. Pomfret protein has a well-balanced amino acid composition, with high amounts of glutamic acid (114 mg/g), lysine (82.8 mg/g), leucine (76.7 mg/g), and aspartic acid (76.0 mg/g). Twenty two fatty acids were found in pomfret oil and saturated fatty acids were the most abundant (48.3%). Palmitic acid (16:0) was the dominant fatty acid, followed by oleic acid (18:1), DHA (22:6n-3), myristic acid (14:0) and stearic acid (18:0), with percentages of 30.5, 26.3, 12.2, 7.37 and 6.86, respectively. The ratio of n-3/n-6 polyunsaturated fatty acids (PUFAs) was 8.04; thus, pomfret muscle is rich in n-3 PUFA.