Evolution of volatile byproducts during wine fermentations using immobilized cells on grape skins

A biocatalyst was prepared by immobilization of Saccharomyces cerevisiae cells on grape skins. Repeated batch fermentations were conducted using this biocatalyst as well as free cells, at 25, 20, 15, and 10 degrees C. Solid phase microextraction (SPME) was used in monitoring the evolution of volatile byproducts. The effect of immobilization and temperature on evolution patterns of volatiles was obvious. The major part of esters was formed after consumption of 40-50% of the sugars. Similar processes were observed for amyl alcohols and 2-phenylethanol, whereas 1-propanol and 2-methyl-1-propanol were formed during the whole alcoholic fermentation period at an almost constant formation rate. Acetaldehyde and acetoin were synthesized in the early stages of fermentation. Afterward, their amount decreased. In most cases, immobilized cells exhibited higher formation rates of volatiles than free cells. The final concentration of esters was higher in wines produced by immobilized biocatalyst. Their amount increased with temperature decrease. The opposite was observed for higher alcohols.     

Volatile compounds of wines produced by cells immobilized on grape skins

A biocatalyst was prepared by immobilization of Saccharomyces cerevisiae cells on grape skins. Repeated batch fermentations were conducted using the immobilized biocatalyst as well as the free yeast cells at 25, 20, 15, and 10 degrees C. The major volatile byproducts were determined by GC, whereas the minor volatile constituents were extracted in dichloromethane and analyzed by HRGC-MS. The qualitative profiles of the wines produced were similar in every case. Immobilized cells gave wines with higher contents of ethyl and acetate esters that increased with temperature decreases from 25 to 15 degrees C. The amount of volatile alcohols was more pronounced in wines produced by free cells and decreased dramatically at low fermentation temperatures (10 degrees C).     

Investigation of volatiles evolution during the alcoholic fermentation of grape must using free and immobilized cells with the help of solid phase microextraction (SPME) headspace sampling

A biocatalyst was prepared by immobilization of Saccharomyces cerevisiae strain AXAZ-1 on delignified cellulosic material (DCM). Repeated batch fermentations were conducted using these biocatalysts and free cells, separately, at temperatures of 20, 15, and 10 degrees C. Solid phase microextraction (SPME) was used in monitoring the formation of volatile alcohols, acetate esters, and ethyl esters of fatty acids. The kinetics of volatile production were similar for free and immobilized cells. In all cases immobilized cells showed a better rate of volatile production, which was directly connected to sugar consumption. The main difference observed was in propanol production, which increased with temperature decrease for the immobilized cells, whereas it remained constant for the free ones. In the case of immobilized cells significant amounts of esters were also produced. It is well-known that esters contribute to the fruity aroma of wine. It was also established that SPME is a very sensitive, accurate, and reliable technique and can be used without any reservation in the characterization of volatile constituents of wine.     

Low-temperature brewing using yeast immobilized on dried figs

Dried figs, following exhaustive extraction of their residual sugars with water, were used for immobilization of Saccharomyces cerevisiae AXAZ-1. The immobilized biocatalyst was used in repeated batch fermentations of glucose at 30 degrees C, where significant reduction of the fermentation time was observed, falling from 65 h in the first batch to 7 h after the sixth batch. Repeated fermentations of wort at room and low temperatures resulted in fermentation times that fell from 26 to 20 h and from 27 to 24 days at 18 and 3 degrees C, respectively. Ethanol and beer productivities were high, showing suitability of the biocatalyst for low-temperature brewing. Diacetyl concentrations were low (0.3-0.5 mg/L), and polyphenols were lower than in commercial products and decreased as the fermentation temperature was decreased (126-50 mg/L). Ethyl acetate concentrations increased from 53 to 88 mg/L as the temperature was decreased, while the concentration of amyl alcohols at 3 degrees C (58 mg/L) was lower than half of that at 18 degrees C (125 mg/L). The beers produced at the end of the main fermentation had a fine clarity and a special fruity figlike aroma and taste, distinct from commercial products and more intense than beers produced by cells immobilized on other food-grade supports (gluten pellets or delignified cellulosic materials). GC-MS analysis did not show significant differences in the qualitative composition of the aroma compounds of the beers produced by immobilized and free cells.     

Evaluation of the thermally dried immobilized cells of Lactobacillus delbrueckii subsp. bulgaricus on apple pieces as a potent starter culture

The aim of the present study was to evaluate the impact of thermal drying of immobilized Lactobacillus delbrueckii subsp. bulgaricus on apple pieces on the use of the derived biocatalyst in whey fermentation. The thermally dried immobilized biocatalyst was compared to wet and freeze-dried immobilized cells, in respect to maintenance of cell viability and fermentation efficiency. The thermal drying process appeared to be more efficient on survival rate as an 84% of the cells used for immobilization survived the process, while the freeze-drying process led to a 78% rate. The thermally dried immobilized biocatalyst was used in 12 repeated batch fermentations of synthetic lactose medium and whey at 37, 45, and 50 degrees C in order to evaluate its metabolic activity. The high number of repeated batch fermentations showed a tendency for high operational stability. Fermentations continued for up to 2 months without any significant loss of metabolic activity. SPME GC/MS analysis of aroma-related compounds revealed the distinctive character of fermented whey produced by the thermally dried immobilized bacterium cells. The effect of storage at 4-6 degrees C for up to 165 days of the biocatalyst, held directly after drying and after repeated batch fermentations, on fermentation activity was also studied. After storage, reactivation in whey was immediate, and the immobilized biocatalyst was able to produce up to 51.7 g/L lactic acid at 37 degrees C. The potential of thermally dried immobilized L. delbrueckii as a starter culture for food production was subsequently evaluated.     

Wine production using yeast immobilized on apple pieces at low and room temperatures

A biocatalyst was prepared by immobilization of Saccharomyces cerevisiae strain AXAZ-1 on apple pieces. It was examined by electron microscope and studied during the fermentation of grape must for batch wine-making. The immobilized yeast showed an important operational stability without any decrease of its activity even at low temperatures (1-12 degrees C). Specifically, at 6 degrees C the biocatalyst favored wine production within 8 days, which is less time than is required for the natural fermentation of grape must. At 1 degrees C wine production was effected in 1 month. This speeding up of the fermentation could be accepted and adopted by the industry for scaling up the wine-making process. Total and volatile acidities were similar to those found in dry wines. The concentrations of higher alcohols (propanol-1 and isobutyl alcohol) were low. The presence of amyl alcohols proved to be temperature dependent and decreased with the temperature decrease. The values of ethyl acetate concentrations were relatively high, up to 154 mg/L. This probably contributes to the fruity aroma and excellent taste of the produced wines. There was no indication of vinegar odor in the product given that the volatile acidity was <0.47 g of acetic acid/L. From the GC-MS analysis it was concluded that cell immobilization did not create serious changes in the product (wine) with regard to the qualitative composition of the aroma compounds.     

Red wine making by immobilized cells and influence on volatile composition

Red wine making using yeast cells immobilized in two types of raisin berries, at various temperatures (6-30 degrees C), was studied. A modification of the batch bioreactor was used to separate the grape skins used for color extraction from the biocatalyst and the fermenting grape must. The evaluation of the immobilized biocatalysts was made on terms of productivity and organoleptic quality, including color intensity and formation of volatiles. The immobilized cells were found capable of low-temperature wine making, producing red wines containing more than 11% v/v alcohol in 8 days at 6 degrees C. The quality of wines was examined by gas chromatography (GC) and GC-MS analysis and sensory evaluation. Higher alcohol concentrations were decreased, and ethyl acetate concentrations increased by the drop of temperature. Many esters, alcohols, carbonyls, and miscellaneous compounds were identified in wines produced by immobilized cells, revealing no significant qualitative differences as compared to wines produced by free cells. The sensory evaluation showed that the best red wine was produced at 6 degrees C.     

Low-temperature brewing by freeze-dried immobilized cells on gluten pellets

A biocatalyst, prepared by the immobilization of a cryotolerant strain of Saccharomyces cerevisiae on gluten pellets, was freeze-dried without any protecting medium and used for repeated batch fermentations of wort for each of the temperatures 15, 10, 5, and 0 degrees C. The fermentation time for freeze-dried immobilized cells was about 2-fold that of the corresponding time for wet immobilized cells on gluten pellets, and lower than the corresponding time for freeze-dried free cells, especially at 5 and 0 degrees C. Beers produced by freeze-dried immobilized cells contained alcohol levels in the range of 5.0-5.5% v/v, diacetyl concentrations lower than 0.5 mg/L, polyphenol concentrations lower than 145.5 mg/L, and free cell concentrations lower than 3 g/L. As a result, they had a very good clarity after the end of primary fermentation. The amounts of amyl alcohols were lower than 129.1 mg/L and reduced as the temperature was decreased. Ethyl acetate concentrations were found in the range of 22.1-29.2 mg/L, giving a very good aroma and taste in the produced beers.     

Storage of immobilized yeast cells for use in wine-making at ambient temperature

A comparative study of the storage and reuse of immobilized yeast cells on apple pieces, kissiris, and gamma-alumina was carried out. The immobilized biocatalysts were allowed to remain in the fermented alcoholic liquid after the end of each fermentation batch for extended periods at 30 degrees C before reactivation in batch fermentation for wine-making. The results showed that the biocatalysts were able to reactivate and ferment after successively increased periods of storage compared to free cell systems both on glucose medium and on grape must. In glucose medium, apple-, kissiris-, and gamma-alumina-supported biocatalysts reactivated after 120, 80, and 83 days, respectively. Possible storage periods for grape must were lower but remained high. Immobilized yeast biocatalyst on apple pieces produced wines with an improved volatiles composition compared to kissiris- and gamma-alumina-supported biocatalysts. There were no significant negative effects on the fermentation activity and volatile byproduct composition.     

Food additive lactic acid production by immobilized cells of Lactobacillus brevis on delignified cellulosic material

Improvements in yield and productivity in lactic acid fermentation by Lactobaccilus brevis cells immobilized on delignified cellulosic (DC) material are reported. The system proved to be more efficient in comparison with the work reported by other workers. Yields of 80 and 100% conversion using glucose were obtained at 30 degrees C in 1 day of fermentation time. Lactic acid fermentation using whey as substrate was obtained at 30 degrees C in 1-1.5 days, resulting in 70% yield, whereas the remaining lactose in whey was converted to alcohol byproduct, leading to a 90% lactose exploitation and 100% conversion. Cell immobilization of L. brevis on DC material was proved by its reuses in repeated batch fermentations and through electron microscopy. A series of 10 repeated batch fermentations without any loss in cell activity showed a tendency for high operational stability. The presence of DC material resulted in a drastic drop of the fermentation time from 48 to 13 h.     

Delignified cellulosic material supported biocatalyst as freeze-dried product in alcoholic fermentation

Freeze-dried delignified cellulosic (DC) material supported biocatalyst is proposed as a suitable form of biocatalyst to be preserved. The alcoholic fermentation of glucose using freeze-dried immobilized cells is reported. Freeze-dried immobilized baker's yeast cells on DC material do not need any protective medium during freeze-drying. The effect of initial glucose concentration and temperature on the alcoholic fermentation kinetic parameters is reported in the present study. It was found that the freeze-dried immobilized cells ferment more quickly than free freeze-dried cells and have a lower fermentation rate as compared with wet immobilized cells. However, repeated batch fermentations showed freeze-dried immobilized cells to ferment at about the same fermentation rate as wet immobilized cells. The results indicate that the freeze-dried immobilized cells must be further studied to establish a process for the preservation of immobilized cells.     

Whey liquid waste of the dairy industry as raw material for potable alcohol production by kefir granules

Kefir granular biomass was used in the fermentation of sweet whey and proved to be more effective compared to single-cell biomass of kefir yeast. The operational stability of the biocatalyst was assessed by carrying out 20 repeated batch fermentations. Levels of ethanol productivity reached 2.57 g L(-1) h(-1)), whereas the yield was 0.45 g/g. The fermentation time was only 8 h. Mixtures of sweet whey with molasses were fermented at initial densities ranging from 4.2 to 10.2 degrees Be and resulted in ethanol yield factors between 0.36 and 0.48 g of ethanol/g of utilized sugar. Lower degrees Be values led to an increase of percentages of ethyl acetate on total volatiles determined and a reduction of amyl alcohols. The addition of 1% black raisin extract to whey appears to promote whey fermentation, whereas the same was not observed in the case of white sultana extract addition. It was finally established that it is preferable to ferment mixtures of whey-molasses by adding molasses in whey after the completion of whey fermentation.     

Volatile and color composition of young and model-aged Shiraz wines as affected by diammonium phosphate supplementation before alcoholic fermentation

A Shiraz must with low yeast assimilable nitrogen (YAN) was supplemented with two concentrations of diammonium phosphate (DAP) and then fermented with maceration on grape skins. The nonvolatile, volatile, and color composition of the final wines were investigated. Ethanol and residual sugars were not affected by DAP supplementation, while glycerol, SO 2, and residual YAN increased and acetic acid decreased. DAP-supplemented treatments gave rise to higher concentrations of acetates, fatty acids, and fatty acid ethyl esters but lower concentrations of branched-chain fatty acids and their ethyl esters. No major difference between treatments was observed for higher alcohols, monoterpenes, norisoprenoids, and low-molecular-weight sulfur compounds. DAP-supplemented fermentations resulted in wines with higher concentrations of malvidin-3-glucoside, higher color intensity, and altered color tonality. Model aging studies indicated that higher concentrations of esters are still present in wines from the DAP-treated fermentations after aging. DAP supplementation also resulted in increased concentrations of dimethyl sulfide after model aging. It can be concluded that DAP treatment of a low YAN must fermented by maceration on skins can significantly affect wine color, aroma, and flavor.     

Experimental Investigation and Artificial Neural Network-Based Modeling of Batch Reduction of Hexavalent Chromium by Immobilized Cells of Newly Isolated Strain of Chromium-Resistant Bacteria

The batch bioreduction of Cr(VI) by the cells of newly isolated chromium-resistant Acinetobacter sp. bacteria, immobilized on glass beads and Ca-alginate beads, was investigated. The rate of reduction and percentage reduction of Cr(VI) decrease with the increase in initial Cr(VI) concentration, indicating the inhibitory effect of Cr(VI). Efficiency of bioreduction can be improved by increasing the bioparticle loading or the initial biomass loading. Glass bioparticles have shown better performance as compared to Ca-alginate bioparticles in terms of batch Cr(VI) reduction achieved and the rate of reduction. Glass beads may be considered as better cell carrier particles for immobilization as compared to Ca-alginate beads. Around 90% reduction of 80 ppm Cr(VI) could be achieved after 24 h with initial biomass loading of 14.6 mg on glass beads. Artificial neural network-based models are developed for prediction of batch Cr(VI) bioreduction using the cells immobilized on glass and Ca-alginate beads.     

Low-temperature alcoholic fermentation by delignified cellulosic material supported cells of kefir yeast.

A novel system for low-temperature alcoholic fermentation of glucose is described. This system consists of kefir yeast immobilized on delignified cellulosic materials. Batch fermentations were carried out at various pH values, and the effect of temperature on kinetic parameters, in the range of 5-30 degrees C, was examined. At pH 4.7 the shortest fermentation time was obtained. The formation of volatiles indicates that the concentration of amyl alcohols (total content of 2-methylbutanol-1 and 3-methylbutanol-1) is reduced as the temperature becomes lower. Propanol-1 and isobutyl alcohol formation drops significantly below 15 degrees C. The percentage of ethyl acetate increases as the temperature is diminished. At 5 degrees C the content of total volatiles in the product was only 38% of the volatiles formed during fermentation at 30 degrees C.     

Production of hard-type cheese using free or immobilized freeze-dried kefir cells as a starter culture

This study provides a contribution to hard-type cheese starter culture production through the use of a freeze-dried culture in the ripening of hard-type cheeses. The effect of initial cell concentration, ripening temperature, and cell immobilization of kefir on the degree of openness, mold spoilage, microbial associations, physicochemical characteristics, and aroma-related compounds was studied. Use of kefir starter cultures resulted in cheese with an increased shelf life and resistance to spoilage as compared to control cheeses without kefir inoculants. Furthermore, the freeze-dried kefir culture improved aroma, taste, and texture characteristics while increasing the degree of openness in comparison to traditional hard-type cheese products. The kefir culture resulted in an increase in counts of total aerobic bacteria, yeasts and molds, lactococci, and lactobacilli until the 15th day of ripening. From then on, only lactobacilli counts increased, reaching levels up to 9.17 log CFU/g in cheeses ripened at 5 degrees C using freeze-dried kefir cells immobilized on casein. SPME-GC/MS analysis revealed major differences in volatile composition, especially with regard to alcohols (up to 75%), carbonyl compounds (up to 75%), and esters (up to 64%) between cheeses made with kefir cells and cheeses made without kefir inoculants.     

Malolactic fermentation in wine with Lactobacillus casei cells immobilized on Delignified cellulosic material

In this work Lactobacillus casei ATCC 393 cells immobilized on delignified cellulosic material (DCM) were used for malolactic fermentation (MLF) of wine. Wine was produced using yeast cells immobilized on DCM at 20 degrees C, and after alcoholic fermentation, MLF at 27 degrees C followed using immobilized L. casei ATCC 393 cells. A total of 11 repeated alcoholic and subsequent MLF batches were performed within a period of 1 month. As the repeated MLF batches proceeded, the MLF activity of the immobilized biocatalyst was reduced. Malic acid degradation was reduced from 80 to 2%, pH was reduced by 0.5-0.1 unit, acetic acid concentrations were slightly reduced or remained stable (0.002 g/L), the higher alcohols 1-propanol, isobutyl alcohol, and amyl alcohol were decreased by 84, 23, and 11%, respectively, and ethyl acetate concentration was increased by approximately 56%. Wine samples were analyzed by GC-MS before and after MLF, revealing some qualitative differences.     

Immobilization of recombinant invertase (re-INVB) from Zymomonas mobilis on D-sorbitol cinnamic ester for production of invert sugar

The recombinant invertase (re-INVB) from Zymomonas mobilis was immobilized by adsorption onto the totally cinnamoylated derivative of D-sorbitol. The polymerization and cross-linking of the derivative initially obtained was achieved by irradiation in the ultraviolet region, where this prepolymer shows maximum sensitivity. Immobilization of re-INVB on this support involves a process of physical adsorption and intense hydrophobic interactions between the cinnamoyl groups of the support and related groups of the enzyme. Enzyme concentration, immobilization time, and irradiation time were important parameters affecting the immobilization efficiency. The optimum reaction pH of immobilized enzyme was 5, and the optimal reaction temperature was 40 degrees C. The apparent Michaelis constant and the apparent catalytic constant of re-INVB immobilized on the SOTCN derivative acting on sucrose was 78+/-5 mM and 5x10(4)+/-3x10(2) s(-1), respectively, while for the free enzyme, it was 98.0+/-4 mM and 1.2x10(4)+/-2.5x10(2) s(-1), respectively, suggesting a better apparent affinity of the enzyme for the substrate and a better hydrolysis rate when immobilized than when in solution. Immobilized re-INVB also showed good thermal stability and good operational stability (40% of the initial activity remaining after 45 cyles of 1 min duration and 90.6 mg of sucrose being hydrolyzed in 45 min per 2.5 mg of immobilized protein). The results showed that cinnamic carbohydrate esters of D-sorbitol are an appropriate support for re-INVB immobilization and the production of invert sugar.     

Removal of Pentachlorophenol by Adsorption on Magnetite-immobilized Chitin

The application of magnetite-immobilized chitin in pentachlorophenol (PCP) removal was demonstrated in this study. The physicochemical parameters for immobilization of chitin by magnetite, and for PCP adsorption using magnetite-immobilized chitin were optimized. For chitin immobilization, the optimized conditions were: magnetite to chitin (m:c) ratio at 1:2, initial pH 6, 25°C, 200 rpm and 60 min in batch system. The immobilization efficiency (IE) was 99.4% and immobilization capacity (IC) was 2.0 mg chitin mg^?1 magnetite. High initial pH (pH?>?11) and temperature (>30°C) lowered the IE and IC. For PCP (10 mg l^?1) adsorption, the optimized conditions were: 1,500 mg l^?1 immobilized chitin, initial pH 6, 25°C, 200 rpm and 60 min in batch system. The removal efficiency (RE) was 57.9% and removal capacity (RC) was 5.4 mg g^?1. The adsorption ability of immobilized chitin decreased with pH and temperature increased. However, increasing the amount of immobilized chitin (24,000 mg l^?1) can increase the RE up to 92%. Both chitin immobilization and PCP adsorption exhibited Langmuir and Freundlich adsorption isotherms. Results in this study indicated that magnetite-immobilized chitin was a cost-effective and environmental friendly adsorbent to remove environmental pollutants such as PCP.     

Fermentation of white wines in the presence of wood chips of American and French oak

Must obtained from Airén grapes was fermented in the presence of wood chips (4 and 7 g/L) of either French oak (from Vosges, central France, and Allier zones) or American oak. Fermentation yields were higher than in the control fermentations carried out in the absence of wood, and production of volatile substances during fermentation (alcohols, esters, and acetates) was also higher. The volatile substances that leached out of the wood were analyzed by GC-MS-SIR. The results showed that their concentrations depended on the type and amount of the oak; some of these substances were consumed in part by the yeasts during fermentation. A taste panel favorably assessed the wines produced by fermentation in the presence of oak chips, which retained part of the must original fruity aroma.