Ethanol tolerance of thermotolerant yeasts cultivated on mixtures of sucrose and ethanol

The final levels of ethanol (levels of ethanol produced plus that added initially to the media) reached by the thermotolerant yeasts were highest (16.5-20.3%, v/v) at 8% initial ethanol. The thermotolerant yeasts were found to have the following characteristics: constant levels of ethanol formation (10.5-12.3%, v/v), fog additions of external ethanol within the range 2-8% (v/v) of initial ethanol; constant values of product coefficients when initial ethanol was in the range of 2-6%, which increased or decreased, depending on the strain, when initial ethanol exceeded 6%; growth activity was inhibited at different levels of addition of external ethanol when final biomass and specific rate of growth were compared; significant differences among the yeast strains in the amount of external ethanol capable of reducing biomass formation by one half. In addition, the viability of the strains (early stationary phase) varied with the amount of external ethanol, the lowest viabilities occurring at concentrations of initial ethanol ranging from 4 to 7% and the highest in the range of 7 to 8% (v/v). The relative levels of trehalose (with/without 7% ethanol added initially) in the yeast strains (the stationary phase) ranged from 1.03 to 1.75, suggesting that the effect of produced ethanol on trehalose accumulation was stronger than that of external ethanol. The levels of final ethanol shown by the yeast strains were also correlated with the cellular levels of glycerol-3-phosphate dehydrogenase (increase in enzyme levels with decrease in final ethanol) for cells harvested at the stationary phase.

Homogenous implant in rat tibias of matrix preserved in 98% glycerin: histomorphologic study.

As a chemical medium for preservation of tissues, glycerin has shown good results because it maintains the cellular integrity despite the tissue dehydration it causes. Taking advantage of the osteoinducing properties of the osseous matrix and glycerin as a proper medium for tissue preservation, osseous matrix was implanted in rat tibias. Twenty-four rats were used, each receiving two surgical wounds. In one of the wounds an osseous matrix preserved in 98% glycerin was implanted and the other received a matrix without preservatives. Six animals were sacrificed on days 10, 20, 30 and 60 post-implant. After routine histological processing, the specimens were stained in hematoxylin-eosin and Masson's trichrome. The results showed that the matrixes preserved in glycerin presented faster resorption with replacement by newly formed tissue.

Distribution of acid phosphatases in the hypopharyngeal glands from workers, queens and males of a Brazilian stingless bee Scaptotrigona postica Latreille: An ultrastructural cytochemical study

The present study reports the localization of acid phosphatase in the hypopharyngeal gland cells from workers (newly-emerged, nurse and forager), queens (newly-emerged and laying) and males (newly-emerged and mature for mating) of the Brazilian stingless bee, Scaptotrigona postica. The phosphatase activity varied in intensity and localization depending on the individual class, physiological age and the substrate used. In newly-emerged workers, the phosphatase-positive sites suggest the involvement of the enzyme with cellular differentiation that occurs in the presecretory phase, in nurse workers with protein synthesis and in forager workers with changes in cellular activity or glandular regression. In males mature for mating and laying queens, the positive sites are related to secretory activity, showing that the gland maintains some activity in spite of the regressive aspect. Of the substrates used, β-glycerophosphate gave the least specific localization.

Evidence for involvement of Saccharomyces cerevisiae protein kinase C in glucose induction of HXT genes and derepression of SUC2

The PKC1 gene in the yeast Saccharomyces cerevisiae encodes protein kinase C that is known to control a mitogen-activated protein (MAP) kinase cascade consisting of Bck1, Mkk1 and Mkk2, and Mpk1. This cascade affects the cell wall integrity but the phenotype of Pkc1 mutants suggests additional targets which have not yet been identified. We show that a pkc1Δ mutant, as opposed to mutants in the MAP kinase cascade, displays two major defects in the control of carbon metabolism. It shows a delay in the initiation of fermentation upon addition of glucose and a defect in derepression of SUC2 gene after exhaustion of glucose from the medium. After addition of glucose the production of both ethanol and glycerol started very slowly. The V max of glucose transport dropped considerably and Northern blot analysis showed that induction of the HXT1, HXT2 and HXT4 genes was strongly reduced. Growth of the pkc1Δ mutant was absent on glycerol and poor on galactose and raffinose. Oxygen uptake was barely present. Derepression of invertase activity and SUC2 transcription upon transfer of cells from glucose to raffinose was deficient in the pkc1Δ mutant as opposed to the wild-type. Our results suggest an involvement of Pkc1p in the control of carbon metabolism which is not shared by the downstream MAP kinase cascade. © 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

Effects of low-intensity pulsed ultrasound on wound healing in corneas of dogs following keratoplasty

The effects of low-intensity pulsed ultrasound on wound healing were evaluated at the graft-cornea transition in dogs following lamellar keratoplasty using tunica vaginalis preserved in 98% glycerin. Twenty-one dogs were subdivided into three groups of seven animals. The first group (W/US) received daily treatment of low-intensity pulsed ultrasound (20 mW/cm 2) for 15 min for the first 10 days post surgery. The second group (N/US) was submitted to the same procedure but with the ultrasound apparatus turned off. The third group, the control (CO), underwent the surgical procedure only. The animals were clinically evaluated during the initial (1-15 days), intermediate (16-30 days) and late (31-120 days) postoperative period. The corneas were evaluated by light microscopy at 1, 3, 7, 15, 30, 60 and 120 days after surgery. Clinically, there were no differences which would promote an advantage to any of the treatments. Light microscopy, however, revealed more extensive vascularization and more advanced wound healing in the W/US group, as well as a tendency towards early graft incorporation. Based on the present results, low-intensity pulsed ultrasound shows advantages, especially in situations where trophic support is a mandatory condition, facilitating better graft incorporation and rapid recovery of stromal organization.

Eucalyptus ESTs associated with resistance to herbicide inhibitors of aromatic and branched-chain amino acid synthesis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Solubilization of proteins from human lymph node tissue and two-dimensional gel storage.

In the present study, we compared six different solubilization buffers and optimized two-dimensional electrophoresis (2-DE) conditions for human lymph node proteins. In addition, we developed a simple protocol for 2-D gel storage. Efficient solubilization was obtained with lysis buffers containing (a) 8 M urea, 4% CHAPS (3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate), 40 mM Tris base, 65 mM DTT (dithiothreitol) and 0.2% carrier ampholytes; (b) 5 M urea, 2 M thiourea, 2% CHAPS, 2% SB 3-10 (N-decyl-N,N-dimethyl-3-ammonio-1-propanesulfonate), 40 mM Tris base, 65 mM DTT and 0.2% carrier ampholytes or (c) 7 M urea, 2 M thiourea, 4% CHAPS, 65 mM DTT and 0.2% carrier ampholytes. The optimal protocol for isoelectric focusing (IEF) was accumulated voltage of 16,500 Vh and 0.6% DTT in the rehydration solution. In the experiments conducted for the sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), best results were obtained with a doubled concentration (50 mM Tris, 384 mM glycine, 0.2% SDS) of the SDS electrophoresis buffer in the cathodic reservoir as compared to the concentration in the anodic reservoir (25 mM Tris, 192 mM glycine, 0.1% SDS). Among the five protocols tested for gel storing, success was attained when the gels were stored in plastic bags with 50% glycerol. This is the first report describing the successful solubilization and 2D-electrophoresis of proteins from human lymph node tissue and a 2-D gel storage protocol for easy gel handling before mass spectrometry (MS) analysis.

Aspectos toxicológicos do bissulfito e outros agentes tóxicos sobre a síntese de glicerol-3-fosfato desidrogenase citoplasmá tica de levedura de panificação

The synthesis of intracellular glycerol-3-phosphate dehydrogenase (EC 1.1.1.8) in baker's yeast was investigated in submerged culture supplied with glucose or glycerol as sole carbon sources. Inhibitors of the glycolytic pathway, Krebs cycle and respiratory chain did not stimulate glycerol-3-phosphate dehydrogenase synthesis when added in low concentrations in up 7.5 × 10 -5 mol/L. The repression exercised by glucose on the synthesis of glycerol-3-phosphate dehydrogenase in YP-glucose medium was reduced by the addition of fermentation products and of sodium bisulfite. Synthesis of the enzyme was raised 22-110%. However, in YP-glycerol medium, the addition of 0.06% (w/v) sodium bisulfite reduced (29%) the synthesis of the enzyme, while 0.012% (v/v) acetaldehyde stimulated the synthesis of glycerol-3-phosphate dehydrogenase by 12%.

Dispersed lipid liquid crystalline phases stabilized by a hydrophobically modified cellulose

Aqueous dispersions of monoolein (MO) with a commercial hydrophobically modified ethyl hydroxyethyl cellulose ether (HMEHEC) have been investigated with respect to the morphologies of the liquid crystalline nanoparticles. Only very low proportions of HMEHEC are accepted in the cubic and lamellar phases of the monoolein-water system. Due to the broad variation of composition and size of the commercial polymer, no other single-phase regions were found in the quasi-ternary system. Interactions of MO with different fractions of the HMEHEC sample induced the formation of lamellar and reversed hexagonal phases, identified from SAXD, polarization microscopy, and cryogenic TEM examinations. In excess water (more than 90 wt %) coarse dispersions are formed more or less spontaneously, containing particles of cubic phase from a size visible by the naked eye to small particles observed by cryoTEM. At high polymer/MO ratios, vesicles were frequently observed, often oligo-lamellar with inter-lamellar connections. After homogenization of the coarse dispersions in a microfluidizer, the large particles disappeared, apparently replaced by smaller cubic particles, often with vesicular attachments on the surfaces, and by vesicles or vesicular particles with a disordered interior. At the largest polymer contents no proper cubic particles were found directly after homogenization but mainly single-walled defected vesicles with a peculiar edgy appearance. During storage for 2 weeks, the dispersed particles changed toward more well-shaped cubic particles, even in dispersions with the highest polymer contents. In some of the samples with low polymer/MO ratio, dispersed particles of the reversed hexagonal type were found. A few of the homogenized samples were freeze-dried and rehydrated. Particles of essentially the same types, but with a less well-developed cubic character, were found after this treatment. © 2007 American Chemical Society.

Fatty acid production by four strains of Mucor hiemalis grown in plant oil and soluble carbohydrates

Four Mucor hiemalis strains (M1, M2, M3 and M4), isolated from soil at a depth of 0 - 15 cm in the Juréia-Itatins Ecology Station (JIES), in the state of São Paulo, Brazil and were evaluated for the production of γ-linolenic (GLA) and other unsaturated fatty acids. Five growth variables (temperature, pH, carbon source, nitrogen source, and vegetable oils) were studied. Liquid media containing 2% vegetable oil (palm oil, canola oil, soybean oil, sesame oil, or sunflower oil) or 2% carbohydrate (fructose, galactose, glycerol, glucose, lactose, maltose, sucrose, sorbitol or xylose) and 1% yeast extract as a nitrogen source were used. The greatest biomass production was observed with M3 and M4 strains in palm oil (91.5 g l -1) and sunflower oil (68.3 g l -1) media, respectively. Strain M4 produced greater quantities of polyunsaturated acids in medium containing glucose. The GLA production in the M4 biomass was 1,132.2 mg l -1 in glucose medium. Plant oils were inhibitors of fatty acid production by these strains. © 2007 Academic Journals.