Continuous whey fermentation using kefir yeast immobilized on delignified cellulosic material

Delignified cellulosic-supported biocatalyst, prepared by immobilization of kefir yeast on delignified cellulosic material (DCM), was found to be suitable for continuous, modified whey fermentation. The modified whey contained 1% raisin extract and molasses. Ethanol productivities ranged from 3.6 to 8.3 g L(-1)day(-1), whereas parameters such as ethanol concentration, residual sugars, and daily fermented whey productivity were acceptable for the production of potable alcohol and alcoholic drinks in industrial fermentations. The continuous fermentation bioreactor was operated for 39 days, stored for 18 days at 4 degrees C, and operated again for another 15 days without any diminution of the ethanol productivity. The concentrations of higher alcohols (propanol-1, isobutyl alcohol, and amyl alcohols) were low. The main volatile byproducts formed in the continuous process were similar to those observed in alcoholic beverages, and the fermented whey had a good aroma. The concentrations of higher alcohols were very low when compared to that of ethyl acetate, therefore resulting in a quality product. The possibility of using such a process for the production of potable alcohol or a novel, low-alcohol content drink is proposed.     

Effect of carbohydrate substrate on fermentation by kefir yeast supported on delignified cellulosic materials

The suitability of delignified cellulosic (DC) material supported kefir yeast to ferment raw materials that contain various single carbohydrates, for the production of potable alcohol and alcoholic drinks, is examined in this investigation. Results are reported of fermentations carried out with sucrose, fructose, and glucose in synthetic media. Repeated batch fermentations at various initial sugar concentrations of sucrose, fructose, and glucose were performed at 30 degrees C in the presence of the aforementioned biocatalyst. The results clearly show feasible yields in the range of 0.38-0.41 g/g, alcohol concentrations of 7.6-8.2% v/v, fermentation time of 90-115 h, and conversion of 92-96%. DC material supported kefir fermented 11-fold more rapidly than free cells and 9-fold more rapidly in comparison to kissiris supported kefir. The main volatile byproducts such as amyl alcohols (mixture of 2-methyl-1-butanol and 3-methyl-1-butanol), ethanal, and ethyl acetate were formed in all sugar fermentation products. The formation of 65-110 ppm of ethyl acetate is as high and even higher than that obtained with traditional wine yeasts. The increase of the initial concentration of sugar in the fermentation media resulted in an increase in contents of volatiles. The fine aroma that was obtained in the product of fructose could be attributed to the high percentage of ethyl acetate on total volatiles. The efficiency of DC material supported kefir was the same for the fermentations of individual sugars or a mixture of fructose, sucrose, and glucose. When whey with raisin extracts was fermented, lower yields were obtained but the aroma of the product was even better.     

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.     

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.     

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.     

生物传感器在纤维素乙醇测定中的应用

为探讨生物传感器在纤维素乙醇测定中的应用,采用乙醇生物传感器分析仪对发酵液中的纤维素乙醇含量进行测定,分析其特点、规律。结果：乙醇生物传感分析仪线性关系良好,线性范围为0.2～20.0 mg/100 mL,回归方程为：Y=5.00225X+0.72273,R＝0.99976;对乙醇的检测快速而准确,检测时间为20 s,回收率为99.3%～100.4 %;与气相色谱方法、酒精计法相比较,差异不显著（P>0.05）;专一性强,且仪器重现性和稳定性较好,检测灵敏度高,检测下限为0.2mg/100mL。乙醇生物传感分析仪用于纤维素乙醇检测,具有检测精度高、速度快（20s）、稳定性好、使用成本低等优势,有望代替酒精计、气相色谱仪等,成为检测纤维素乙醇含量的新方法、新手段。     

Extremely low temperature fermentations of grape must by potato-supported yeast, strain AXAZ-1. A contribution is performed for catalysis of alcoholic fermentation

This investigation announces the use of potato pieces as a suitable support for cell immobilization resulting in extremely low temperature wine making. The results showed an increase of the total esters by immobilized cells and reduction of higher alcohols. Likewise, percentages of total esters on total volatiles were increased by the drop in temperature, while percentages of higher alcohols were reduced in wines. Kinetics experiments at different temperatures allowed the calculation of activation energy (Ea) and showed reduction in the case of immobilized cells as compared with free cells. These results may lead to the conclusion that the increased productivities that are obtained by immobilized cells, can be attributed to the catalytic activity by the support to enzymes, which are involved in the process. Biocatalysts were prepared by immobilization of Saccharomyces cerevisiae, strain AXAZ-1, on whole potatoes and potato pieces, and their efficiency for alcoholic repeated batch fermentations of glucose and grape must in the range 2-30 degrees C was examined. To study the operational stability of biocatalyst, 35 repeated batch fermentations of grape must were performed without any significant reduction of the fermentation activity. Wines were analyzed for volatile byproducts determination by GC and GC-MS.     

Production of organically bound selenium yeast by continuous fermentation

This paper describes a protocol for incorporation of sodium selenite or sodium selenate into Saccharomyces cerevisiae biomass by continuous fermentation in a medium with minimal sulfur and methionine concentrations. Selenium incorporation was followed by atomic absorption analysis and methylene blue reduction time (MBRT). Continuous fermentation at 0.2 h(-1) dilution rate and sodium selenite addition gradient up to 0.69 g/L of Na(2)SeO(3) yielded 1. 89 g/L of biomass with 1904 microg of selenium/g of dry biomass. However, MBRT was 0.1 min, which indicated that the majority of selenium was in the inorganic form. On the other hand, continuous fermentation at 0.2 h(-1) dilution rate and sodium selenate gradient up to 0.28 g/L of Na(2)SeO(4) yielded 0.76 g/L of dry biomass with 687 microg of selenium/g of dry biomass, and MBRT was 26 min, which indicated a high concentration of organically bound selenium. Overall, the results indicate a Se/S ratio of 3.9:1 and a dry biomass/Se ratio of 5.5:1 as optimal for continuous production of organically bound selenium.     

Enhanced organically bound selenium yeast production by fed-batch fermentation

This study describes a fed-batch fermentation protocol for enhanced production of Saccharomyces cerevisiae containing organically bound selenium. Two levels of sodium selenate concentrations were applied as either a single dose or continuous addition. Fermentations with high sodium selenate (63.2 g/L in cane molasses feeding medium) demonstrated 24 g/L of biomass with 1382 microg of selenium/g of dry biomass for single-dose addition and 40 g/L of biomass with 1491 microg of selenium/g of dry biomass for continuous addition. Low selenium concentration (31.6 g/L in cane molasses feeding medium) demonstrated higher biomass concentration with higher selenium level; 37 g/L of biomass with 2846 microg of selenium/g of dry biomass and 45 g/L of biomass with 2495 microg of selenium/g of dry biomass for single-dose and continuous addition, respectively. Also, two adapted S. cerevisiae strains were evaluated in fed-batch fermentation. A single dose of low concentration demonstrated >3000 microg of selenium/g of dry biomass, but biomass concentration was lower (< or =32 g/L) for these adapted strains.     

Comparison of biotransformation of inorganic selenium by Lactobacillus and Saccharomyces in lactic fermentation process of yogurt and kefir

The aim of the present study was to characterize, quantify, and compare the different selenium species that are produced when lactic fermentation with two different types of microorganisms, bacteria (Lactobacillus) and yeast (Saccharomyces), take place to produce yogurt and kefir, respectively, and to study the transformation process of these species as a function of time. These two dairy products were chosen for the study because they are highly consumed in different cultures. Moreover, the microorganisms present in the fermentation processes are different. While the bacteria Lactobacillus is the one responsible for yogurt fermentation, a partnership between bacteria and the yeast Saccharomyces causes kefir fermentation. A comparative study has been carried out by fermenting Se(IV) enriched milk in the presence of both types of microorganisms, where the concentration range studied was from 0.5 to 20 microg g (-1). Enzymatic extraction enabled selenium speciation profiles, obtained by anionic exchange and ion-pairing reversed phase high performance liquid chromatography (IP-RP-HPLC) with inductively coupled plasma mass spectrometry (ICP-MS) detection. Scanning electron microscopy (SEM) applied to the enriched samples showed segregated Se (0), at added concentrations higher than 5 microg g (-1). The main Se species formed depended on the type of microorganism involved in the fermentation process, SeCys 2 and MeSeCys being the main species generated in yogurt and SeMet in kefir. The results obtained are different for both kinds of samples. Lactic fermentation for yogurt produced an increment in selenocystine (SeCys 2) and Se-methylselenocysteine (MeSeCys), while fermentation to produce kefir also incremented the selenomethionine (SeMet) concentration. The Se species are stable for at least 10 and 15 days for kefir and yogurt, respectively. After this period, selenocystine concentration decreased, and the concentration of Se-methylselenocysteine was found to significantly increase.     

甜高粱茎秆汁液固定化酵母酒精发酵的研究

本文应用一元线性回归分析法得出了甜高粱茎秆汁液锤度和可溶性总糖含量之间的关系。以甜高粱茎秆汁液为原料,在摇床及流化床反应器上进行了固定化酵母酒精发酵的试验研究。结果表明,以沈农甜杂2号甜高粱为原料,在摇床上作营养盐配比的正交试验,试验得出对提高酒精得率影响因素的主次顺序为：(NH₄)₂SO₄,K₂HPO₄,MgSO₄。最优组合为：0.125% K₂HPO₄,0.200%(NH₄)₂SO₄,0.050% MgSO₄。加营养盐的酒精得率比不加任何营养盐的酒精得率可提高4.0%～8.0%。K₂HPO₄,(NH₄)₂SO₄的配比对试验结果影响极显著,MgSO₄的配比对试验结果影响显著。在流化床反应器上试验,4种较优营养盐组合酒精得率近乎相等,比不加营养盐空白试验酒精得率高。从降低成本角度来讲,选择0.125% K₂HPO₄,0.200%(NH₄)₂SO₄,0.010% MgSO₄作为投产试验。本文为燃料乙醇的发展提供了科学的依据。     

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.     

Stability of patulin to sulfur dioxide and to yeast fermentation.

The affinity of patulin for sulfur dioxide (SO2) is much less than was previously reported and is of little significance at the SO2 concentrations (below 200 ppm) used in the processing of apple juice and cider. However, at concentrations of 2000 ppm SO2 and 15 ppm patulin, combination was 90% complete in 2 days. Removal of SO2 liberated only part of the patulin, which suggests that 2 mechanisms are involved: one reversible (opening the hemiacetal ring) and one irreversible (SO2 addition at the double bond). Test with 2 yeasts used in English commercial cider making confirmed that patulin is effectively removed during yeast fermentation.     

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.     

Monitoring of alcoholic fermentation of onion juice by NIR spectroscopy: valorization of worthless onions

The valorization of vegetable byproducts is one of the main objectives of industry today. The project on which this study is based examined the potential usefulness of worthless onions (Allium cepa L. sp.) and overproduction to obtain several functional products with different applications in the food industry. Near-infrared (NIR) spectroscopy, combined with multivariate calibration, has been used to monitor the alcoholic fermentation of onion juice. Good results were obtained, revealing the suitability of NIR spectroscopy for controlling and optimizing this process in real time.     

基于电导率酒精发酵过程在线监测的研究

本试验对酒精发酵过程中电导率与还原糖、酒精度、pH的变化规律以及氯化钙和硫酸铵等盐类对电导率的影响进行了研究,进而探讨电导率在线监测酒精发酵过程的适用性。结果表明：电导率与还原糖和酒精度之间存在着一定的逻辑关系,利用电导率的变化能间接地反映还原糖和酒精度的变化;pH值与电导率之间存在着一定的线性关系(发酵60 h前变化规律相同);氯化钙和硫酸铵两种盐类在一定范围内对电导率的变化不产生影响。电导率达到最低点时,还原糖和酒精度均符合酒精发酵终点的参数指标,因此,该研究结果可用于发酵过程的在线监测和发酵终点的判定。     

Adsorption of anthocyanins by yeast cell walls during the fermentation of red wines

This paper reports the anthocyanin adsorption profiles of the cell walls of different Saccharomyces strains isolated from grapes collected in the Spanish appellation controlée regions of La Rioja, Navarra, and Ribera del Duero. These strains are habitually used in red wine-making. The acyl derivatives of anthocyanins (acetyl and p-coumaryl compounds) were more strongly adsorbed than nonacyl derivatives. Peonidin-3G was also strongly adsorbed, as were its acyl derivatives. The greater presence of acetyl derivatives in the cell wall adsorbate leads to an increase in yellow color and a reduction in blue color with respect to the corresponding wine.     

L-(-)-malic acid production by Saccharomyces spp. during the alcoholic fermentation of wine (1)

In an attempt to increase the acidity of wine by biological means, malate-producing yeasts were selected from a collection of 282 strains isolated in different parts of Spain. Only 4% of these strains (all of which belonged to Saccharomyces cerevisiae) produced l-(-)-malic acid in the range of 0.5-1 g/L. This was formed between days 2 and 6 of alcoholic fermentation, reaching a maximum on days 3 and 4; the concentration remained stable from day 7. Malic acid production was favored by temperatures in the 18-25 degrees C range and by musts with a high pH and low concentrations of sugar, initial malic acid, and yeast-assimilable nitrogen. Oxaloacetic acid, a precursor of malic acid, had no influence on malate production. The precursors pyruvic and fumaric acid did, however, have a significant effect on the production of this acid in some strains. No direct relation between pyruvate and malate metabolism was observed.     

Behavior of a fenhexamid photoproduct during the alcoholic fermentation of Saccharomyces cerevisiae

The fungicide fenhexamid [N-(2,3-dichloro-4-hydroxyphenyl)-1-methylcyclohexanecarboxamide] degraded rapidly by UV or sunlight irradiation, yielding 7-chloro-6-hydroxy-2-(1-methylcyclohexyl)-1,3-benzoxazole (CHB) as a main photoproduct. CHB was isolated, and its effect on alcoholic fermentation of Saccharomyces cerevisiae was studied. The results indicate that the presence of CHB does not affect the extent of alcohol production. After 12 days, the amount of CHB in the fermentation medium decreased by ca. 65%. Only 25% of the missing CHB was recovered unchanged from yeasts, most likely because it was adsorbed on the yeast wall cell. The remaining part degraded during the fermentation process. Glucan and chitin, two potential adsorbents, which constitute yeast cell walls, exhibited affinity for CHB.