Influence of malting procedure on the isoflavonoid content of soybeans.

The goal of this study was to analyse, whether malting technique (consisting of seed hydration, germination and drying) can be used to modify concentration of various isoflavonoids in soybean seeds. Seeds of three soybean varieties were germinated by different lengths of time (from 24 to 120 h) and dried by two different methods, typically used to produce so-called 'light' and 'caramel' malts. It was determined, that malting decreases concentration of 7-O-ß-D-glucosides such as daidzin, genisitin and glycitin, while at the same time increasing concentration of aglycones (daidzein, genistein and glycitein). Increasing time of the germination period increased concentration of aglycones. 'Caramel' type malts were characterised with higher concentration of most of the isoflavonoids (daidzin, daidzein, genistin, genistein and glycitein) than 'light' type malts. Results of this study suggest that soybean malts can be an interesting substrate in the production of various food products with increased aglycone content.

The Reaction of the Yeast Saccharomyces cerevisiae to Contamination of the Medium with Aflatoxins B2 and G1, Ochratoxin A and Zearalenone in Aerobic Cultures.

The mechanisms by which yeast cells respond to environmental stress include the production of heat shock proteins (HSPs) and the reduction of oxidative stress. The response of yeast exposed to aflatoxins B2+G1 (AFB2+G1), ochratoxin A (OTA), and zearalenone (ZEA) in aerobic conditions was studied. After 72 h of yeast cultivation in media contaminated with mycotoxins, the growth of yeast biomass, the level of malondialdehyde, and the activity of superoxide dismutase, glutathione S-transferase and glutathione peroxidase were examined; the expression profile of the following heat shock proteins was also determined: HSP31, HSP40, HSP60, HSP70, and HSP104. It was demonstrated that at the tested concentrations, both AFB2+G1 and ZEA inhibited yeast biomass growth. OTA at a concentration of 8.4 [µg/L] raised the MDA level. Intensified lipoperoxidation and increased activity of SOD and GPx were observed, regardless of the level of contamination with ZEA (300 µg/L or 900 µg/L). Increased contamination with AFB2+G1 and OTA caused an increase in the production of most HSPs tested (HSP31, HSP40, HSP70, HSP104). ZEA contamination in the used concentration ranges reduced the production of HSP31. The response of yeast cells to the presence of mycotoxin as a stressor resulted in the expression of certain HSPs, but the response was not systematic, which was manifested in different profiles of protein expression depending on the mycotoxin used. The tested mycotoxins influenced the induction of oxidative stress in yeast cells to varying degrees, which resulted in the activation of mainly SOD without GST mobilization or with a small involvement of GPx.

High-pressure microwave-assisted pretreatment of softwood, hardwood and non-wood biomass using different solvents in the production of cellulosic ethanol.

BACKGROUND: Pretreatment is an indispensable stage of the preparation of lignocellulosic biomass with key significance for the effectiveness of hydrolysis and the efficiency of the production of cellulosic ethanol. A significant increase in the susceptibility of the raw material to further degradation can be attained as a result of effective delignification in high-pressure conditions. With this in mind, a method of high-pressure pretreatment using microwave radiation and various solvents (water, 40% w/v NaCS, 1% v/v H2SO4, 1% w/v NaOH or 60% v/v EtOH with an addition of 1% v/v H2SO4) was developed, enabling the acquisition of biomass with an increased susceptibility to the process of enzymatic hydrolysis. The medium obtained in this way can be used for the production of cellulosic ethanol via high-gravity technology (lignocellulosic media containing from 15 to 20% dry weight of biomass). For every type of biomass (pine chips, beech chips and wheat straw), a solvent was selected to be used during the pretreatment, guaranteeing the acquisition of a medium highly susceptible to the process of enzymatic hydrolysis. RESULTS: The highest efficiency of the hydrolysis of biomass, amounting to 71.14 ± 0.97% (glucose concentration 109.26 ± 3.49 g/L) was achieved for wheat straw subjected to microwave-assisted pretreatment using 40% w/v NaCS. Fermentation of this medium produced ethanol concentration at the level of 53.84 ± 1.25 g/L. A slightly lower effectiveness of enzymatic hydrolysis (62.21 ± 0.62%) was achieved after high-pressure microwave-assisted pretreatment of beech chips using 1% w/v NaOH. The hydrolysate contained glucose in the concentration of 91.78 ± 1.91 g/L, and the acquired concentration of ethanol after fermentation amounted to 49.07 ± 2.06 g/L. In the case of pine chips, the most effective delignification was achieved using 60% v/v EtOH with the addition of 1% v/v H2SO4, but after enzymatic hydrolysis, the concentration of glucose in hydrolysate was lower than in the other raw materials and amounted to 39.15 ± 1.62 g/L (the concentration of ethanol after fermentation was ca. 19.67 ± 0.98 g/L). The presence of xylose and galactose was also determined in the obtained fermentation media. The highest initial concentration of these carbohydrates (21.39 ± 1.44 g/L) was observed in beech chips media after microwave-assisted pretreatment using NaOH. The use of wheat straw after pretreatment using EtOH with an addition of 1% v/v H2SO4 for the preparation of fermentation medium, results in the generation of the initial concentration of galactose and xylose at the level of 19.03 ± 0.38 g/L. CONCLUSION: The achieved results indicate a high effectiveness of the enzymatic hydrolysis of the biomass subjected to high-pressure microwave-assisted pretreatment. The final effect depends on the combined use of correctly selected solvents for the different sources of lignocellulosic biomass. On the basis of the achieved results, we can say that the presented method indicates a very high potential in the area of its use for the production of cellulosic ethanol involving high-gravity technology.

Changes in the raffinose family oligosaccharides content in the lentil and common bean seeds during malting and mashing processes.

Raffinose family oligosaccharides (RFOs) are sugars, which are considered anti-nutritional substances, which are not digestible by human gastric enzymes and can lead to flatulence. Legume seeds are often rich in these compounds, which can be cumbersome for many people, such as vegetarians or the population of developing countries, whose diets consists of large amounts of these food products. In this study, simple procedures used around the world in the brewing industry (malting and mashing) were used to determine, whether these processes could be applied to popular legume seeds (lentil and bean) to reduce the RFOs content. Acquired malts and worts were characterised by radically decreased concentration (up to 90%) of most ubiquitous RFOs, such as raffinose and stachyose.

Cellulosic Ethanol Production Using Waste Wheat Stillage after Microwave-Assisted Hydrotropic Pretreatment.

One of the key elements influencing the efficiency of cellulosic ethanol production is the effective pretreatment of lignocellulosic biomass. The aim of the study was to evaluate the effect of microwave-assisted pretreatment of wheat stillage in the presence of sodium cumene sulphonate (NaCS) hydrotrope used for the production of second-generation bioethanol. As a result of microwave pretreatment, the composition of the wheat stillage biomass changed significantly when compared with the raw material used, before treatment. Microwave-assisted pretreatment with NaCS effectively reduced the lignin content and hemicellulose, making cellulose the dominant component of biomass, which accounted for 42.91 ± 0.10%. In post pretreatment, changes in biomass composition were also visible on FTIR spectra. The peaks of functional groups and bonds characteristic of lignins (C-O vibration in the syringyl ring, asymmetric bending in CH3, and aromatic skeleton C-C stretching) decreased. The pretreatment of the analyzed lignocellulosic raw material with NaCS resulted in the complete conversion of glucose to ethanol after 48 h of the process, with yield (in relation to the theoretical one) of above 91%. The highest observed concentration of ethanol, 23.57 ± 0.10 g/L, indicated the high effectiveness of the method used for the pretreatment of wheat stillage that did not require additional nutrient supplementation.

Delignification efficiency of various types of biomass using microwave-assisted hydrotropic pretreatment.

The use of a method of an effective delignification of lignocellulosic biomass is a key stage of designing processes of its microbiological conversion e.g. for the purposes of the production of cellulosic ethanol. The study was aimed at evaluating the effectiveness of microwave-assisted hydrotropic pretreatment using sodium cumene sulfonate (NaCS) for the delignification of pine and beech chips and wheat straw. Research results presenting the impact of process parameters of microwave-assisted hydrotropic delignification confirm a high effectiveness of this method of pretreatment of lignocellulosic biomass. The observed effects included changes in the composition of the biomass and an increased susceptibility of cellulose to the subsequent enzymatic hydrolysis. The use of microwave heating combined with an addition of hydrotrope of 40% w/v NaCS and 117 PSI for 60 min enabled a reduction of the absolute concentration of lignins by 36.58% in pine chips, by 57.68% in beech chips, and by 74.08% in wheat straw. After enzymatic hydrolysis was conducted, the highest concentration of glucose: 463.27 ± 11.25 mg glucose/g (hydrolysis yield 46.76 ± 1.14%) was obtained from the wheat straw, while 327.70 ± 22.15 mg glucose/g (hydrolysis yield 35.13 ± 2.37%) was acquired from the beech chips, and only 50.77 ± 0.75 mg glucose/g (hydrolysis yield 6.63 ± 0.10%) was obtained from the pine chips. Microwave-assisted hydrotropic delignification in the optimum process conditions additionally allows a complete removal of hemicellulose from biomass, which improves the effectiveness of enzymatic hydrolysis. Due to a significant reduction of lignin and hemicellulose concentration in biomass, cellulose-which is susceptible to enzymatic hydrolysis and a source of carbon in biosynthesis processes-becomes the main biomass component.

Application of white grape pomace in the brewing technology and its impact on the concentration of esters and alcohols, physicochemical parameteres and antioxidative properties of the beer.

Main by-product of white wine production is white grape pomace (WGP). It has attracted attention of food scientists, because it possesses high concentration of nutrients and bioactive substances. In this study, WGP was added to the beer after primary fermentation in two different concentrations (10% w/w and 20% w/w) and two different pretreatments (pasteurised and unpasteurised) to determine, whether the most abundant waste from white wine industry could be used to modify the volatilome and phenolic content of the beer. The addition of white grape pomace increased the concentration of phenolic compounds in all of the tested beers (from 321.584 mg gallic acid equivalent (GAE)/dm3 to 501.459 mg GAE/dm3). Antioxidant activity of the beers with addition of WGP (tested with the ABTS+•, DPPH• and FRAP assays) also increased. The composition of volatiles in beers changed as WGP was added. The most significant difference was in the concentration of acetaldehyde - beers with WGP added had 4-7 times lower acetaldehyde content (17.425-31.425 mg/dm3) than the control sample (134.050 mg/dm3).

Pyrazines Biosynthesis by Bacillus Strains Isolated from Natto Fermented Soybean.

Pyrazines are organic compounds with a varied, intense aroma of roasted nuts, occasionally with hints of baked potatoes, almonds, and others. As a result, they are used in the food industry as food flavorings. Biosynthesis of pyrazines using microorganisms in environmentally friendly conditions is an alternative to chemical synthesis. However, screening is required to isolate efficient producer strains for efficient biosynthesis of this compound. The study's goal was to assess the ability of Bacillus subtilis cultures isolated from natto (fermented soybeans) to biosynthesize a broad range of alkylpyrazines. B. subtilis isolated cultures were found to be capable of producing 2-methylpyrazine, 2,3-dimethylpyrazine, 2,5-dimethylpyrazine, 2,6-dimethylpyrazine, 2,3,5-trimethylpyrazine, and 2,3,5,6-tetramethylpyrazine. As a result of the screening, two cultures of B. subtilis capable of producing alkylpyrazines were isolated. At a total concentration of 3261 µg/L, the BcP4 strain primarily produced 2-methylpyrazine (690 µg/L), 2,3-dimethylpyrazine (680 µg/L), and 2,6-dimethylpyrazine (1891 µg/L). At a total concentration of 558 mg/L, the BcP21 strain produced 2,5-dimethylpyrazine (4.5 mg/L), 2,3,5-trimethylpyrazine (52.6 mg/L), and 2,3,5,6-tetramethylpyrazine (501.1 mg/L). The results show that different B. subtilis strains are predisposed to produce different alkylpyrazines.

Influence of the Medium Composition and the Culture Conditions on Surfactin Biosynthesis by a Native Bacillus subtilis natto BS19 Strain.

An effective microbial synthesis of surfactin depends on the composition of the culture medium, the culture conditions and the genetic potential of the producer strain. The aim of this study was to evaluate the suitability of various medium components for the surfactin producing strain and to determine the impact of the culture conditions on the biosynthesis of surfactin isoforms by the newly isolated native strain Bacillus subtilis natto BS19. The efficiency of surfactin biosynthesis was determined by measuring the surface tension of the medium before and after submerged culture (SmF) and by qualitative and quantitative analysis of the obtained compound by high performance liquid chromatography. The highest efficiency of surfactin biosynthesis was achieved using starch as the carbon source and yeast extract as the nitrogen source at pH 7.0 and 37 °C. Potato peelings were selected as an effective waste substrate. It was shown that the increase in the percentage of peel extract in the culture medium enhanced the biosynthesis of surfactin (mg/L) (2-30.9%; 4-46.0% and 6-58.2%), while reducing surface tension of the medium by about 50%. The obtained results constitute a promising basis for further research on biosynthesis of surfactin using potato peelings as a cheap alternative to synthetic medium components.

Impact of Lignocellulose Pretreatment By-Products on S. cerevisiae Strain Ethanol Red Metabolism during Aerobic and An-aerobic Growth.

Understanding the specific response of yeast cells to environmental stress factors is the starting point for selecting the conditions of adaptive culture in order to obtain a yeast line with increased resistance to a given stress factor. The aim of the study was to evaluate the specific cellular response of Saccharomyces cerevisiae strain Ethanol Red to stress caused by toxic by-products generated during the pretreatment of lignocellulose, such as levulinic acid, 5-hydroxymethylfurfural, furfural, ferulic acid, syringaldehyde and vanillin. The presence of 5-hydroxymethylfurfural at the highest analyzed concentration (5704.8 ± 249.3 mg/L) under aerobic conditions induced the overproduction of ergosterol and trehalose. On the other hand, under anaerobic conditions (during the alcoholic fermentation), a decrease in the biosynthesis of these environmental stress indicators was observed. The tested yeast strain was able to completely metabolize 5-hydroxymethylfurfural, furfural, syringaldehyde and vanillin, both under aerobic and anaerobic conditions. Yeast cells reacted to the presence of furan aldehydes by overproducing Hsp60 involved in the control of intracellular protein folding. The results may be helpful in optimizing the process parameters of second-generation ethanol production, in order to reduce the formation and toxic effects of fermentation inhibitors.

Hydrotropic pretreatment on distillery stillage for efficient cellulosic ethanol production.

Effectiveness of hydrotropic delignification using sodium cumene sulfonate for pretreatment of rye, wheat and maize stillage for further use in the production of bioethanol was evaluated. The highest stillage biomass extractives was obtained for a biomass particle size <1.0 mm, when exposed to 131 °C for 1 h at 20% v/v hydrotrope concentration. It has been shown that hydrotropic treatment causes changes in the stillage biomass structure (increase in porosity) and reduces the lignin content in biomass by 7-17%. Delignification with a hydrotrope also increased the concentration of fermentable sugars in the media prepared with stillage biomass, which led to a higher final ethanol concentration (up to ca. 3.5 g/L). Hydrotropic treatment is an effective way of pretreatment of stillage biomass. It provides a high degree of biomass bioconversion and creates the prospect of integrating the 1st and 2nd generation ethanol production process to more fully utilize the raw material.

Evaluation of various methods of selection of B. subtilis strains capable of secreting surface-active compounds.

The aim of the study was the evaluation of a three-step method for the selection of bacterial strains capable of producing surfactin. The procedure consisted of the following steps: 1.blood agar test, 2. measurement of the surface tension (ST) of the medium using the du Nouy method before and after submerged culture, 3. qualitative and quantitative assessment of surfactin by HPLC. Forty five Bacillus subtilis natto strains producing haemolysis zones (≥3mm) were selected. Nineten of them reduced ST of the medium to ≤ 40 mN/m; in six cases, the reduction was as much as 50%. All indicated strains produced surfactin. Positive correlations (p <0.5) between the percentage reduction of ST of the medium and surfactin concentration (r = 0.44), indicate that this parameter is determinant of the ability to synthesize this compound. The blood agar test has been shown to be useful only as a pre-selection criterion for surfactin producers (18 strains selected by this method reduced ST by only ≤30%). The proposed selection strategy proved effective and made it possible to select the BS15 strain that reduced the ST of the medium to 30.56 ± 0.15 mN/m and simultaneously provided a high concentration of surfactin compared to other strains.

Extracellular Phytase Production by the Wine Yeast S. cerevisiae (Finarome Strain) during Submerged Fermentation.

One of the key steps in the production of phytases of microbial origin is selection of culture parameters, followed by isolation of the enzyme and evaluation of its catalytic activity. It was found that conditions for S. cerevisiae yeast culture, strain Finarome, giving the reduction in phytic acid concentration of more than 98% within 24 h of incubation were as follows: pH 5.5, 32 °C, continuous stirring at 80 rpm, the use of mannose as a carbon source and aspartic acid as a source of nitrogen. The highest catalytic activity of the isolated phytase was observed at 37 °C, pH 4.0 and using phytate as substrate at concentration of 5.0 mM. The presence of ethanol in the medium at a concentration of 12% v/v reduces the catalytic activity to above 60%. Properties of phytase derived from S. cerevisiae yeast culture, strain Finarome, indicate the possibility of its application in the form of a cell's free crude protein isolate for the hydrolysis of phytic acid to improve the efficiency of alcoholic fermentation processes. Our results also suggest a possibility to use the strain under study to obtain a fusant derived with specialized distillery strains, capable of carrying out a highly efficient fermentation process combined with the utilization of phytates.

Evaluation of phytic acid utilization by S. cerevisiae strains used in fermentation processes and biomass production.

Saccharomyces cerevisiae is a well-studied yeast species used mainly in fermentation processes, bakery, and for SCP (Single Cell Protein) acquisition. The aim of the study was to analyze the possibility of phytic acid utilization as one of the hydrolysis processes carried out by yeast. The analysis of 30 yeast strains used in fermentation and for biomass production, that were grown in media containing phytic acid, revealed a high variability in the biomass production rate and the capability to hydrolyze phytates. No correlation between a high biomass concentration and a high level of phytate hydrolysis was found. Only four analyzed strains (Bayanus IOC Efficience, Sano, PINK EXCEL, FINAROME) were able to reduce the phytic acid concentration by more than 33.5%, from the initial concentration 103.0 ± 2.1 µg/ml to the level below 70 µg/ml. The presented results suggest that the selected wine and fodder yeast can be used as in situ source of phosphohydrolases in fermentation processes, and especially in the production of fodder proteins. However, further studies aimed at the optimization of growing parameters, such as the maximization of phytase secretion, and a comprehensive analysis of the catalytic activity of the isolated phosphohydrolases, are necessary.

Changes in the Concentration of Carbonyl Compounds during the Alcoholic Fermentation Process Carried out with Saccharomyces cerevisiae Yeast.

The aim of the study was to determine the influence of the source material and the applied S. cerevisiae strain on the concentrations of carbonyl fractions in raw spirits. Acetaldehyde was the most common aldehyde found, as it accounted for 88-92% of the total amount of aldehydes. The concentration of acetaldehyde in maize, rye and amaranth mashes was highly correlated with fermentation productivity at a given phase of the process, and reached its highest value of 193.5 mg/l EtOH in the first hours of the fermentation, regardless of the yeast strain applied. The acetaldehyde concentration decreased over the time with the decreasing productivity, reaching its lowest value at the 72nd hour of the process. The final concentration of acetaldehyde depended on the raw material used (ca 28.0 mg/l EtOH for maize mashes, 40.3 mg/l EtOH for rye mashes, and 74.4 mg/l EtOH for amaranth mashes). The effect of the used yeast strain was negligible. The overall concentration of the analyzed aldehydes was only slightly higher: ca 30.3 mg/l EtOH for maize mashes, 47.8 mg/l EtOH for rye mashes, and 83.1 mg/l EtOH for amaranth mashes.

The effect of raw material contamination with mycotoxins on the composition of alcoholic fermentation volatile by-products in raw spirits.

The effects of the mycotoxins, aflatoxin B(1), B(2), G(1), G(2) (AF), ochratoxin A, (OTA), zearalenone (ZEA), deoxynivalenol (DON), and fumonisin B(1) (FB(1)) added to corn grain mashes on the composition of fermentation volatile by-products in raw spirits were determined. Except for FB(1), the mycotoxins increased acetaldehyde concentration in the obtained spirits from about 30% to 100% in relation to the control set (30.9+/-1.0mg of acetaldehyde/L EtOH). The largest effect was observed for OTA and AF contaminations (65.9+/-5.9 and 62.4+/-5.0mg/L EtOH, respectively). At the concentrations used (ppb): FB(1), 1875; FB(2), 609; FB(3), 195; DON, 2274; ZEA, 352; AFB(1), 11.65; AFB(2), 12.6; AFG(1), 12.34; AFG(2), 12.04; OTA, 177.5, the mycotoxins did not have a significant effect on the total level of higher alcohols in distillates. As compared to the control, contamination with OTA and FB(1) decreased the 3-methyl-1-butanol concentration by 11.2% and 12.6% respectively, whereas AF decreased the 2-methyl-1-butanol concentration by 14.9%. The mycotoxins AF, ZEA, FB(1), had no significant effect on the concentration of total esters. Whereas OTA caused twofold higher esters concentration in the distillates, DON lowered esters concentration by 32% as compared to control. Presented results show that quantitative changes in composition of volatile fermentation by-products in raw spirits can be related to the presence of increased level of mycotoxins in raw material, especially in the absence of other identifiable factors disturbing the normal course of process.

Characterisation of fermentation of high-gravity maize mashes with the application of pullulanase, proteolytic enzymes and enzymes degrading non-starch polysaccharides.

The aim of the research was to assess the possibility of the fermentation productivity rising through the increase in corn mashes extract from 16-17 to 20-21 degrees Balling, yet keeping a 3-day fermentation period. The second goal was to obtain the highest possible utilization of starch in the raw material through deep enzymatic degradation and utilization of available sugars and simultaneous maintenance of high quality spirit. It was found that fulfilling the above during the 3-day fermentation period was possible with the application of pullulanase as an additional amylolytic enzyme. Adding pullulanase resulted in the acceleration of the starch hydrolysis degree, which led to lower amounts of unhydrolyzed dextrins and higher ethanol yield. When the supportive enzymes complex (pullulanase, protease and cellulase) was used, the final ethanol concentration reached 10.86+/-0.04% v/v, with ethanol yield at 68.41+/-0.23 dm(3) of absolute ethanol (A(100)) per 100 kg of starch, which was 95.25+/-0.32% at the theoretical value. The acceleration of starch enzymatic degradation and the application of a proteolytic preparation visibly shortened both initial and main fermentation phases. This in turn increased the time of the final fermentation phase and resulted in more extensive utilization of substrates by yeasts with simultaneous reduction of the final concentration of acetaldehyde (26.0+/-0.5 mg/dm(3)A(100)) and diethyl acetal of acetaldehyde (2.5+/-0.5 mg/dm(3)A(100)). The quality of spirit obtained was positively verified also in terms of relatively low concentration of higher alcohol (3912.2+/-9.8 mg/dm(3)A(100)). Preliminary analysis of costs (without raw-material) of 1 l distillate production indicated the possibility to reduce the costs by 18-20%.

The influence of raw material contamination with mycotoxins on alcoholic fermentation indicators.

The aim of the research was to describe the influence of selected mycotoxins on major factors (alcohol concentration, productivity, yield and energy) that are characteristic of the fermentation process of maize mashes. Indicators of the alcoholic fermentation of mashes made from raw material with low contaminations levels were compared with mashes obtained from raw material that was selectively contaminated with mycotoxins on the following concentrations: aflatoxin B(1)-11.65 ppb, B(2)-12.60 ppb, G(1)-12.34 ppb, G(2)-12.04 ppb; ochratoxin A-177.5 ppb; zearalenone-352 ppb; deoxynivalenol-2274 ppb; fumonisin B(1)-1875 ppb, B(2)-609 ppb, B(3)-195 ppb. It was found that, apart from fumonisin, all mycotoxins substantially affected the course of subsequent fermentation phases, in particular the first and the main fermentation phases. The highest drop in alcohol concentration at the main stage of the process amounted to 1% v/v and it was achieved by contamination with zearalenone. The statistically significant drop in the final fermentation yield was observed; this was caused by raw material contaminated with all studied mycotoxins, except for fumonisin. The decrease in ethanol yield in reference to the control variant ranged from 1.42 to 3.20 dm(3) of absolute alcohol out of 100 kg of starch, depending on a toxin.