Methylene blue dye test for rapid qualitative detection of hydroxyl radicals formed in a Fenton's reaction aqueous solution

A new procedure, the methylene blue dye test, qualitatively indicates the presence of hydroxyl radicals through the immediate, distinct bleaching of methylene blue dye on a paper test strip. This method employs a simple procedure requiring inexpensive materials, without the addition of competitive probe chemicals that potentially can interfere with the reaction. A Fenton's reaction with an Fe2+:H2O2 molar ratio of 1:20 generated hydroxyl radicals in Milli-Q water. The presence and absence of hydroxyl radicals were determined prior to and following quenching of the Fenton's reaction with 10% sodium sulfite, respectively. Bleaching of methylene blue dye, due to the presence of hydroxyl radicals in a sample,was indicated by a discoloration from a dark blue color to an almost white color, concentrated at the point of application, with a dark blue outline. A lack of bleaching indicated the absence of hydroxyl radicals in the sample. The presence of hydroxyl radicals was verified by benzoic acid chemical probe experiments with thin-layer chromatography (TLC) and spectrophotometric wavelength scans. The presence of hydroxyl radicals was indirectly determined by detection of hydroxylated benzoic acids on TLC plates and a violet solution color with a peak absorbance at a wavelength close to 520 nm.     

Next‐generation biofuels: a new challenge for yeast

Economic growth depends strongly on the availability and price of fuels. There are various reasons in different parts of the world for efforts to decrease the consumption of fossil fuels, but biofuels are one of the main solutions considered towards achieving this aim globally. As the major bioethanol producer, the yeast Saccharomyces cerevisiae has a central position among biofuel‐producing organisms. However, unprecedented challenges for yeast biotechnology lie ahead, as future biofuels will have to be produced on a large scale from sustainable feedstocks that do not interfere with food production, and which are generally not the traditional carbon source for S. cerevisiae. Additionally, the current trend in the development of biofuels is to synthesize molecules that can be used as drop‐in fuels for existing engines. Their properties should therefore be more similar to those of oil‐derived fuels than those of ethanol. Recent developments and challenges lying ahead for cost‐effective production of such designed biofuels, using S. cerevisiae‐based cell factories, are presented in this review. Copyright © 2015 John Wiley & Sons, Ltd.     

Oxidation of amino groups by hydroxyl radicals in relation to the oxidation degree of the alpha-carbon

Nitrogen organic compounds constitute a large class of aqueous pollutants. These compounds include not only azoic structures, nitrogen heterocycles, and nitrous groups but also amides and amines. This work consisted in studying the OH* induced oxidation of simple primary amines in dilute aqueous solution with special attention to mineralization of the nitrogen group as a function of the nature of the alpha-carbon. H2O2/UV and gamma-irradiation processes were used for the production of OH* radicals, and the molecules studied were one alpha-amino acid i.e., glycine (HOOCCH2NH2), and two primary amides i.e., acetamide (CH3CONH2) and oxamic acid (HOOCCONH2). It was shown that the oxidation of glycine leads to the formation of ammonia, whereas the acetamide molecule is first oxidized into oxamic acid ending in complete mineralization with production of nitrates. Reaction mechanisms are proposed which account for the observed inorganic nitrogen end product depending on the oxidation degree of the carbon atoms of the molecules. It follows that the present study will allow for prediction of the fate of nitrogen resulting from the oxidation of primary amino groups by OH* radicals.     

流加培养生产玉米酵母工艺研究

通过研究以玉米粉作为培养基流加培养生产酵母的工艺,得出:流加培养生产玉米酵母最佳接种量10%,溶氧量为50%,同时添加营养盐和微量营养素,最适宜酵母细胞的生长温度为30℃,流加培养的最佳时间是20～22h,采用植物油为消泡剂,发酵过程中减少CO2,鲜酵母的产量达到了99.8g/L。     

Long-lived yeast as a model for ageing research

Yeast has essentially two lifespans: a replicative lifespan (the number of daughters produced by each dividing mother cell) and a chronological lifespan (the capacity of stationary (G0) cultures to maintain viability over time). There is a tendency now to label every investigation that addresses these lifespans as ageing research. It is, though, analyses of the longest lifespans that will be most informative about the determinants of longevity and yield results most relevant to ageing in more complex systems. This review addresses these issues and describes the ongoing studies that are now attempting to address ageing in yeast cells of maximal replicative or chronological longevity.     

Roles of hydroxyl radicals in electrolyzed oxidizing water (EOW) for the inactivation of Escherichia coli

The food industry has recognized electrolyzed oxidizing water (EOW) as a promising alternative decontamination technique. However, there is not a consensus about the sanitizing mechanism of EOW. In this study, we evaluated the disinfection efficacy of different types of EOW on Escherichia coli. Based on the hypothesis of hydroxyl radicals existing in EOW, in the present study, the hydroxyl radicals existed in slightly acidic electrolyzed water (SAEW) and acidic electrolyzed water (AEW) diluted to different levels were detected quantitatively. An ultraviolet (UV) spectrophotometer was used to scan EOW with different pH values. Accounting for the results of UV scanning to EOW with different pH value and the disinfection efficacy of different types of EOW, it can be concluded that considering the lower chlorine concentration of EOW compared with traditional chlorine disinfectants, the existing form of chlorine compounds rather than the hydroxyl radicals played important role in the disinfection efficacy of EOW.     

不同麦汁浓度及酵母代数的电导率在线监测

采用电导率测量的方法对啤酒发酵过程进行了在线监测，并且对不同麦汁浓度、不同酵母代数进行了研究。结果表明：对于不同浓度的麦汁，其发酵过程中电导率的变化呈现明显的规律性，酵母代数对电导率的影响较大，1代酵母要比0代酵母提前约50min出现第1个拐点，即结束迟滞期；提前3h出现第2个拐点。这为研究啤酒发酵提供了一种有效的方法。     

Budding yeast as a model organism for population genetics

Population genetics is a highly theoretical field in which many models and theories of broad significance have received little experimental testing. Microbes are well-suited for empirical population genetics since populations of almost any size may be studied genetically, and because many have easily controlled life cycles. Saccharomyces cerevisiae is almost ideal for such studies as the growing body of knowledge and techniques that have made it the best characterized eukaryote genome also allow the experimental manipulation and analysis of its population genetics. In experiments to date, the evolution of laboratory yeast populations has been observed for up to 1000 generations. In several cases, adaptation has occurred by gene duplications. The interaction between mutation, selection and genetic drift at varying population sizes is a major area of theoretical study in which yeast experiments can provide particularly valuable data. Conflicts between gene-level and among-cell selection, and co-evolution between genes within a genome, are additional topics in which a population genetics perspective may be particularly helpful. The growing field of genomics is increasingly complementary with that of population genetics. The characterization of the yeast genome presents unprecedented opportunities for the detailed study of evolutionary and population genetics. Conversely, the redundancy of the yeast genome means that, for many open reading frames, deletion has only a quantitative effect that is most readily observed in competitions with a wild-type strain.     

浓盐法提取啤酒废酵母核糖核酸的研究

以抽提率和收率为指标,通过单因素试验,确定了盐法提取啤酒废酵母中核糖核酸的参数,然后通过正交试验获得了最佳提取工艺,结果为:提取时间5 h,提取温度100℃,氯化钠浓度8%,酵母浓度8%.在此条件下RNA抽提率为7.12%,收率为99.30%,OD260/OD280为2.01.     

酵母RNA超声辅助提取工艺研究

为提高酵母RNA提取效果,采用超声辅助提取工艺,在单因素实验的基础上,采用超声处理时间、超声功率和提取时间进行三因素三水平响应面分析实验以优化此工艺条件.结果表明,酵母RNA超声辅助提取的最佳工艺条件为超声处理时间10min,超声功率300W,提取时间20min,在此工艺条件下,酵母RNA实际得率为9.56%,与模型预测值之间具有较好的拟合性.在3个因素中,超声处理时间、超声功率对酵母RNA得率的影响极显著,提取时间影响不显著,且相互之间无交互效应.     

INT reduction as a basis for determining bakers' yeast viability

Two rapid and simple methods for determining the reduction of 2-( p -iodophenyl)-3-( p -nitrophenyl)-5-phenyltetrazolium chloride (INT) to INT-formazan by bakers' yeast were compared. The first method involves counting with a light microscope the number of yeast cells containing INT-formazan crystals. In the second method, INT-formazan is extracted from cells retained on membrane filters, using dimethyl sulphoxide. The INT-formazan is then quantitated by absorption spectrophotometry. Several brands of bakers' yeast of various ages were tested. Results obtained with the two methods were closely correlated. The extraction method is the more rapid and convenient of the two assays investigated and may be used to follow change in activity with time, compare various yeast brands or investigate the effect of various environmental conditions on yeast INT-dehydrogenase activity.     

Detection of specific RNA sequences in yeast by in situ colony hybridization

Recently a convenient method for detection of specific RNA sequences in bacteria has been developed but the original protocol was inapplicable to microorganisms with a rigid cell wall. Here we report a modification of the RNA colony hybridization for use with yeast. The modified method includes the following consecutive procedures: a) treatment of the yeast colonies on the membrane filter with 10% SDS at 65% C for 30 min; b) treatment of the same filter with 3 x SSC, 10% formaldehyde at 65 degrees C for 30 min; c) hybridization with 32P-labelled oligonucleotide (or DNA) specific for the RNA sequence of interest. The intensity of the radioactive signals thus obtained is comparable with that of the E. coli colonies.     

Experimental Design as a Tool for the Development of Citric Acid Production by Yarrowia lipolytica Using Glycerol as a Feedstock

Due to the scarcity of fossil fuels in the world, there is increasing interest in the commercial production of biodiesel, which leads to obtaining large amounts of glycerol as a byproduct. If not disposed of properly, glycerol can generate environmental impact. One of the promises, the application of the crude glycerol is the production of citric acid by microbial fermentation. Citric acid is industrially produced by a submerged fermentation process with Aspergillus niger, using sucrose as carbon source, but due to increased demand for citric acid, alternative processes using renewable sources or waste materials as substrates and the cultivation of yeast strains are being studied. The aim of the study was to determine the best culture condition for maximum citric acid synthesis and lower isocitric acid production from crude glycerol through experimental design tool. For this purpose, the yeast strain Yarrowia lipolytica IMUFRJ-50682 was cultivated in nitrogen-limited glycerol-based media. Therefore, glycerol and yeast extract concentrations and agitation speed were evaluated as independent variables. With pure glycerol, the highest citric acid production achieved was 16.5 g/L with an isocitric acid production of 7.7% （in relation to citric acid）. With crude glycerol, citric acid production reduced to 6.7 g/L because of higher biomass yield. Therefore, an increase in the initial carbon to nitrogen molar ratio from 714 to 1,561 was necessary to increase citric acid production to 9.2 g/L, reducing isocitric acid production and to achieve a yield of 0.41 g of citric acid per glycerol consumed. In this condition, less nitrogen source was used, reducing production costs.     

Breeding of brewer's yeast by hybridization between a top-fermenting yeast Saccharomyces cerevisiae and a cryophilic yeast Saccharomyces bayanus

To improve the fermentability of a top-fermenting yeast at low-temperature, we performed hybridization trials between four top-fermenting Saccharomyces cerevisiae strains and a cryophilic yeast Saccharomyces bayanus YM84 with good fermentability at low-temperature. The hybrids selected using 5-bromo-4-chloro-3-indolyl-alpha-D-galactopyranoside were checked with pulsed-field gel electrophoresis and their brewing performance at the low-temperature of 10.5 degrees C was observed using small-scale (2 l) fermentation trials.     

Pseudomonas aeruginosa lectin PA-IIL as a powerful probe for human and bovine milk analysis

Milk composition exhibits species-specific differences depending on genetic, evolutionary, and environmental factors. In addition, commercial milk preparations are also changed by industrial manipulations, including severe heat processing. Cow milk, used as human food, provides important nutrients but lacks some essential components that are present in raw human milk. The present study, which was aimed at comparing infant breastfeeding to cow-based formula nourishment, shows major differences between the human and the commercial cow milk glycans detectable by the lectins PA-IL (galactose-binding) and PA-IIL (fucose and mannose-binding) isolated from the cells of human pathogen Pseudomonas aeruginosa. More than 40 human milk samples, several cow milks, and bovine milk-based infant formulas, were examined using these two lectins. For purposes of comparison, the plant lectins Concanavalin A (Con A), which binds mannose, and Ulex europaeus 1st lectin (UEA-I), which binds fucose, were also used. The most prominent difference was revealed using PA-IIL, which displayed a unique high sensitivity to the human milk fucosylated compounds. PA-IL and UEA-I also exhibited preferential sensitivity to the human milk but considerably lower than that of PA-IIL. Con A was inhibited by human and the other milk preparations examined to the same extent. These findings indicate the superb applicability of PA-IIL for rapid and reliable comparative investigation of milk glycans from human and cow, indicating which glycans could be added to infant formulas in order to enrich them, as well as for verification and quality control of otherwise improved bovine milk-based infant formulas.     

Induction of autolytic breakdown of RNA in yeast by addition of ethanol and by drying/rehydration.

Autolytic degradation of RNA in suspensions of yeast at 50°C has been further studied as a means of preparing single-cell protein (SCP) suitable for human consumption. Membrane disorganisation with initiation of autolysis was promoted by addition of ethanol (5 % v/v) or, more effectively, by first air-drying yeast in a fluid-bed dryer and rehydrating it at 4°C, or at 50°C, before incubation at 50°C. Water was a poor suspension medium, but nearly complete breakdown of RNA within 1.5 h was achieved with a buffer, or with a 0.5 m solution of NaCl. Salt in high concentration possibly destroyed ribosomal structure. With pressed commercial baker 's yeast, a consideration of its composition suggested that an ideal procedure for removing RNA would result in the loss of about 24% of dry matter, with the production of SCP containing 54% protein. With careful timing, SCP with 48% protein could be obtained from air-dried yeast on extraction with 0.5 M-NaCl, with a loss of 32% of the dry matter. Active dried baker 's yeast also autolysed rapidly at 50°C, and had a relatively low proteinase activity. After extensive autolysis of baker 's yeast, the residual nucleic acid (about 5 % of that initially present) was characterised by a relatively high ratio of adenine/guanine.