Sodium ion interaction with psyllium husk (Plantago sp.).

The nature of and factors effecting sodium interactions with psyllium were investigated in vitro. In a batch extraction system, psyllium mucilage gel retained at least 50% of sodium across a range of concentrations (5-300 mg sodium per g psyllium) and pH (2-10) environments. FTIR and Na NMR analyses of psyllium gels indicated that binding was complex with non-specific multi-site interactions. The potential use of psyllium husk as a binding agent for the reduction of bioavailable sodium was therefore evaluated. The binding of sodium at physiologically relevant conditions (pH 1.2 (stomach) and 6.8 (intestine)) was studied in a gastrointestinal tract (GIT) pH simulated model. Results show consistently high sodium retention (∼50%) across the GIT model and less than 20% loss of bound sodium under the simulated intestinal pH conditions after repeated washings.

The phenotypic characterization of yeast strains to stresses inherent to wine fermentation in warm climates.

AIMS: Climate change is exerting an increasingly profound effect on grape composition, microbiology, chemistry and the sensory aspects of wine. Identification of autochthonous yeasts tolerant to stress could help to alleviate this effect. METHODS AND RESULTS: Tolerance to osmotic pressure, ethanol and pH of 94 Saccharomyces cerevisiae strains and 29 strains non-Saccharomyces from the warm climate region DO 'Vinos de Madrid' (Spain) using phenotypic microarray and their fermentative behaviour were studied. The screening highlighted 12 strains of S. cerevisiae isolated from organic cellars with improved tolerance to osmotic stress, high ethanol concentrations and suitable fermentative properties. Screening of non-Saccharomyces spp. such as Lanchacea thermotolerans, Torulaspora delbrueckii, Schizosaccharomyces pombe and Mestchnikowia pulcherrima also highlighted tolerance to these stress conditions. CONCLUSIONS: This study confirmed the adaptation of native strains to the climatic conditions in each area of production and correlated these adaptations with good fermentation properties. Screening has revealed that identifying yeast strains adapted to fermentation stresses is an important approach for making quality wines in very warm areas. SIGNIFICANCE AND IMPACT OF THE STUDY: The results have special relevance because it is a pioneering study that has approached the problem of climate change for wines from a microbiological aspect and has analysed the situation in situ in wineries from a warm climate zone. Resistant strains were found with good biological properties; studying these strains for their stress response mechanisms during fermentation will be of interest to the wine making industry.

Impact of nitrogen flushing and oil choice on the progression of lipid oxidation in unwashed fried sliced potato crisps.

Unwashed, sliced, batch-fried potato crisps have a unique texture and are growing in popularity in the UK/EU premium snack food market. In this study, the storage stability of unwashed sliced (high surface starch) potatoes (crisps) fried in regular sunflower oil (SO) or in high oleic sunflower oil (HOSO) was compared over accelerated shelf life testing (45°C, 6 weeks); with and without nitrogen gas flushing. Primary oxidation products (lipid hydroperoxides) were measured with a ferrous oxidation-xylenol orange (FOX) assay and volatile secondary oxidation products (hexanal) were quantified by using solid phase micro-extraction gas chromatography mass spectrometry (HS-SPME-GC/MS). Results revealed that crisps fried in SO were the least stable. Flushing the stored crisps with nitrogen gas proved to be effective in slowing down the oxidation rate after frying with sunflower oil, significantly stabilizing the crisps. However, crisps fried in HOSO were the most stable, with the lowest rate of development of oxidation markers, and this has previously not been shown for crisps with a high free starch content.

Expression of RCK2 MAPKAP (MAPK-activated protein kinase) rescues yeast cells sensitivity to osmotic stress.

BACKGROUND: Saccharomyces cerevisiae is the micro-organism of choice for the conversion of fermentable sugars during beverage or bioethanol fermentations. These fermentations are characterised by high osmotic stress on a yeast cell, with selected brewing fermentations beginning at 20-25% fermentable sugars and bioethanol fermentations at 13% fermentable sugars. RESULTS: RCK2 encodes for a MAPKAP (MAPK-activated protein kinase) enzyme and was identified on a locus by QTL analysis in yeast cells under osmotic stress, RCK2 expression was placed under a tetracycline regulatable vector and rescued glucose, sorbitol or glycerol induced osmotic stress in an rck2 null strain. A strain overexpressing RCK2 had significantly faster fermentation rates when compared with the empty vector control strain. CONCLUSIONS: Presence of RCK2 increased rates of glucose utilisation (~40 g glucose in first 8 h) during a 15% glucose fermentation and concurrent production of ethanol when compared with empty vector controls. Tolerance to osmotic stress using the tetracycline regulatable vectors could be turned off with the addition of tetracycline returning a rck2 null strain back to osmotic sensitivity.

Phenotypic characterisation of Saccharomyces spp. for tolerance to 1-butanol.

Biofuels are expected to play a role in replacing crude oil as a liquid transportation fuel, and research into butanol has highlighted the importance of this alcohol as a fuel. Butanol has a higher energy density than ethanol, butanol-gasoline blends do not separate in the presence of water, and butanol is miscible with gasoline (Szulczyk, Int J Energy Environ 1(1):2876-2895, 40). Saccharomyces cerevisiae has been used as a fermentative organism in the biofuel industry producing ethanol from glucose derived from starchy plant material; however, it typically cannot tolerate butanol concentrations greater than 2 % (Luong, Biotechnol Bioeng 29 (2):242-248, 27). 90 Saccharomyces spp. strains were screened for tolerance to 1-butanol via a phenotypic microarray assay and we observed significant variation in response with the most tolerant strains (S. cerevisiae DBVPG1788, S. cerevisiae DBVPG6044 and S. cerevisiae YPS128) exhibiting tolerance to 4 % 1-butanol compared with S. uvarum and S. castelli strains, which were sensitive to 3 % 1-butanol. Response to butanol was confirmed using traditional yeast methodologies such as growth; it was observed that fermentations in the presence of butanol, when using strains with a tolerant background, were significantly faster. Assessing for genetic rationale for tolerance, it was observed that 1-butanol-tolerant strains, when compared with 1-butanol-sensitive strains, had an up-regulation of RPN4, a transcription factor which regulates proteasome genes. Analysing for the importance of RPN4, we observed that a Δrpn4 strain displayed a reduced rate of fermentation in the presence of 1-butanol when compared with the BY4741 background strain. This data will aid the development of breeding programmes to produce better strains for future bio-butanol production.

Development of a phenotypic assay for characterisation of ethanologenic yeast strain sensitivity to inhibitors released from lignocellulosic feedstocks.

Inhibitors released by the breakdown of plant cell walls prevent efficient conversion of sugar into ethanol. The aim of this study was to develop a fast and reliable inhibitor sensitivity assay for ethanologenic yeast strains. The assay comprised bespoke 96-well plates containing inhibitors in isolation or combination in a format that was compatible with the Phenotypic Microarray Omnilog reader (Biolog, hayward, CA, USA). A redox reporter within the assay permits analysis of inhibitor sensitivity in aerobic and/or anaerobic conditions. Results from the assay were verified using growth on spot plates and tolerance assays in which maintenance of viability was assessed. The assay allows for individual and synergistic effects of inhibitors to be determined. It was observed that the presence of both acetic and formic acid significantly inhibited the yeast strains assessed, although this impact could be partially mitigated by buffering to neutral pH. Scheffersomyces stipitis, Candida spp., and Pichia guilliermondii demonstrated increased sensitivity to short chain weak acids at concentrations typically present in lignocellulosic hydrolysates. S. cerevisiae exhibited robustness to short chain weak acids at these concentrations. However, S. stipitis, Candida spp., and P. guilliermondii displayed increased tolerance to HMF when compared to that observed for S. cerevisiae. The results demonstrate that the phenotypic microarray assay developed in the current study is a valuable tool that can be used to identify yeast strains with desirable resistance to inhibitory compounds found in lignocellulosic hydrolysates.

Presence of proline has a protective effect on weak acid stressed Saccharomyces cerevisiae.

Fermentation of sugars released from lignocellulosic biomass (LCMs) is a sustainable option for the production of bioethanol. LCMs release fermentable hexose sugars and the currently non-fermentable pentose sugars; ethanol yield from lignocellulosic residues is dependent on the efficient conversion of available sugars to ethanol, a side-product of the process is acetic acid production. Presence of acetic acid reduced metabolic output and growth when compared with controls; however, it was observed that incubation with proline had a protective effect, which was proline specific and concentration dependent; the protective effect did not extend to furan or phenolic stressed yeast cells. Proline accumulating strains displayed tolerance to acetic acid when compared with background strains, whereas, strains with a compromised proline metabolism displayed sensitivity. Sensitivity to weak acids appears to be reduced with the addition of proline; proline is an imino acid freely available as a nitrogen source in the aerobic phase of fermentations. Yeast strains with higher intracellular proline concentrations would be desirable for industrial bioethanol fermentations.

An old activity in the cytochrome P450 superfamily (CYP51) and a new story of drugs and resistance.

Cytochrome P450 51 (CYP51) is sterol 14alpha-demethylase, known also as Erg11p in yeast. First studied in yeast, where it is one of three CYPs in the genome, it has subsequently gained attention as the only CYP found so far in different kingdoms of life. As such it is central to considerations of CYP evolution. Recent use of CYP51-inhibiting antifungal drugs, such as fluconazole, has also been associated with dramatic CYP51 evolution to numerous resistant forms in fungal pathogens. CYP51 has also been discovered in mycobacteria where antifungal azoles have effect and might be of value against tuberculosis. Evolutionary and therapeutic aspects of CYP51 studies are discussed.