Design and techno-economic evaluation of microbial oil production as a renewable resource for biodiesel and oleochemical production

Experimental results from the open literature have been employed for the design and techno-economic evaluation of four process flowsheets for the production of microbial oil or biodiesel. The fermentation of glucose-based media using the yeast strain Rhodosporidium toruloides has been considered. Biodiesel production was based on the exploitation of either direct transesterification (without extraction of lipids from microbial biomass) or indirect transesterifaction of extracted microbial oil. When glucose-based renewable resources are used as carbon source for an annual production capacity of 10,000 t microbial oil and zero cost of glucose (assuming development of integrated biorefineries in existing industries utilising waste or by-product streams) the estimated unitary cost of purified microbial oil is $3.4/kg. Biodiesel production via indirect transesterification of extracted microbial oil proved more cost-competitive process compared to the direct conversion of dried yeast cells. For a price of glucose of $400/t oil production cost and biodiesel production cost are estimated to be $5.5/kg oil and $5.9/kg biodiesel, correspondingly. Industrial implementation of microbial oil production from oleaginous yeast is strongly dependent on the feedstock used and on the fermentation stage where significantly higher productivities and final microbial oil concentrations should be achieved.

Spatio-temporal variability of polynya dynamics and ice production in the Laptev Sea between the winters of 1979/80 and 2007/08

Polynyas in the Laptev Sea are examined with respect to recurrence and interannual wintertime ice production.We use a polynya classification method based on passive microwave satellite data to derive daily polynya area from long-term sea-ice concentrations. This provides insight into the spatial and temporal variability of open-water and thin-ice regions on the Laptev Sea Shelf. Using thermal infrared satellite data to derive an empirical thin-ice distribution within the thickness range from 0 to 20 cm, we calculate daily average surface heat loss and the resulting wintertime ice formation within the Laptev Sea polynyas between 1979 and 2008 using reanalysis data supplied by the National Centers for Environmental Prediction, USA, as atmospheric forcing. Results indicate that previous studies significantly overestimate the contribution of polynyas to the ice production in the Laptev Sea. Average wintertime ice production in polynyas amounts to approximately 55 km39 27% and is mostly determined by the polynya area, wind speed and associated large-scale circulation patterns. No trend in ice production could be detected in the period from 1979/80 to 2007/08.

Rhodosporidium toruloides cultivated in NaCl-enriched glucose-based media: adaptation dynamics and lipid production

In the present report and for the first time in the international literature, the impact of the addition of NaCl upon growth and lipid production on the oleaginous yeast Rhodosporidium toruloides was studied. Moreover, equally for first time, lipid production by R. toruloides was performed under non-aseptic conditions. Therefore, the potentiality of R. toruloides DSM 4444 to produce lipid in media containing several initial concentrations of NaCl with glucose employed as carbon source was studied. Preliminary batch-flask trials with increasing amounts of NaCl revealed the tolerance of the strain against NaCl content up to 6.0% (w/v). However, 4.0% (w/v) of NaCl stimulated lipid accumulation for this strain, by enhancing lipid production up to 71.3% (w/w) per dry cell weight. The same amount of NaCl was employed in pasteurized batch-flask cultures in order to investigate the role of the salt as bacterial inhibiting agent. The combination of NaCl and high glucose concentrations was found to satisfactorily suppress bacterial contamination of R. toruloides cultures under these conditions. Batch-bioreactor trials of the yeast in the same media with high glucose content (up to 150 g/L) resulted in satisfactory substrate assimilation, with almost linear kinetic profile for lipid production, regardless of the initial glucose concentration imposed. Finally, fed-batch bioreactor cultures led to the production of 37.2 g/L of biomass, accompanied by 64.5% (w/w) of lipid yield. Lipid yield per unit of glucose consumed received the very satisfactory value of 0.21 g/g, a value amongst the highest ones in the literature. The yeast lipid produced contained mainly oleic acid and to lesser extent palmitic and stearic acids, thus constituting a perfect starting material for “second generation” biodiesel

Emission of polycyclic aromatic hydrocarbons from gasohol and ethanol vehicles

The exhaust emission of the polycyclic aromatic hydrocarbons (PAHs) considered toxic to human health were investigated on two spark ignition light duty vehicles, one being gasohol (Gasohol, in Brazil, is the generic denomination for mixtures of pure gasoline plus 20-25% of anhydrous ethyl alcohol fuel (AEAF).)-fuelled and the other a flexible-fuel vehicle fuelled with hydrated ethanol. The influence of fuel type and quality, aged lubricant oil type and use of fuel additives on the formation of these compounds was tested using standardized tests identical to US FTP-75 cycle. PAH sampling and chemical analysis followed the basic recommendations of method TO-13 (United States. Environmental Protection Agency, 1999. Compendium Method TO-13A - Determination of polycyclic Aromatic hydrocarbons (PAH) in Ambient Air Using Gas Chromatography/Mass Spectrometry (CG/MS). Center for environmental research information, Cincinnati, p. 78), with the necessary modification for this particular application. Results showed that the total PAH emission factor varied from 41.9 mu g km(-1) to 612 mu g km(-1) in the gasohol vehicle, and from 11.7 mu g km(-1) to 27.4 mu g km(-1) in the ethanol-fuelled vehicle, a significant difference in favor of the ethanol vehicle. Generally, emission of light molecular weight PAHs was predominant, while high molecular weights PAHs were not detected. In terms of benzo(a)pyrene toxicity equivalence, emission factors varied from 0.00984 mu g TEQ km(-1) to 4.61 mu g TEQ km(-1) for the gasohol vehicle and from 0.0117 mu g TEQ km(-1) to 0.0218 mu g TEQ km(-1) in the ethanol vehicle. For the gasohol vehicle, results showed that the use of fuel additive causes a significant increase in the emission of naphthalene and phenanthrene at a confidence level of 90% or higher; the use of rubber solvent on gasohol showed a reduction in the emission of naphthalene and phenanthrene at the same confidence level; the use of synthetic oil instead of mineral oil also contributed significantly to a decrease in the emission of naphthalene and fluorene. In relation to the ethanol vehicle, the same factors were tested and showed no statistically significant influence on PAH emission. (c) 2008 Elsevier Ltd. All rights reserved.

Human-animal interaction, stress, and embryo production in Bos indicus embryo donors under tropical conditions

This study investigated the effect of human-animal interaction (HAI) and the stress response on the quality of embryo production in superovulated Nelore (Bos indicus) cattle, under tropical conditions. Thirty-two females underwent a superovulation protocol for 5 days. Cortisol concentrations were determined in blood plasma collected on days 0, 4, and 5. Artificial insemination was performed on days 4 and 5, and nonsurgical embryo flushing on day 11. Embryo production and viability were determined. Human stimulation, animal behaviors, accidents, and handling time were recorded to assess HAI. Cattle age was negatively correlated with accidents, frequency of aversive behaviors, and negative stimuli by stockperson during transit through corral compartments to receive superovulation treatments. The factor analysis revealed two distinct groups. The first group was called stressed and had higher cortisol concentration than the nonstressed group, 16.0 +/- 2.1 and 12.5 +/- 1.0 ng/mL, respectively. Comparisons between these groups showed that the frequency of voice emissions by the stockperson and the number of accidents were higher in the stressed group, and also, the mean handling time was longer in the stressed group than for the nonstressed. As a result, viability rate of the embryos was 19% lower in the stressed group (P < 0.05). This indicates that intensive negative HAI is likely related to stress, which affects embryo production in a superovulation program.

Ethylene and polyamine production patterns during in vitro shoot organogenesis of two passion fruit species as affected by polyamines and their inhibitor

Levels of ethylene and polyamines (PAs) were measured during organogenesis of hypocotyl explants of two species of passion fruit (Passiflora cincinnata Masters and Passiflora edulis Sims f. flavicarpa Degener `FB-100`) to better understand the relationships of these regulators and their influence on cell differentiation and morphogenesis. Moreover, histological investigation of shoot ontogenesis was conducted to characterize the different events involved in cell redifferentiation and regulation of PA and ethylene levels. A delay was observed in morphogenic responses of P. edulis f. flavicarpa as compared to P. cincinnata, and these changes coincided with production of elevated levels of polyamine and ethylene levels. During differentiation, cells showed high rates of expansion and elongation, and high ethylene levels were associated with high PA levels, suggesting that the two biosynthesis pathways were highly regulated. Moreover, their interaction might be an important factor for determining cell differentiation. The addition of PAs to the culture medium did not promote organogenesis; however, the incorporation of the PA inhibitor methylglyoxal bisguanylhydrazone in the culture medium reduced shoot bud differentiation, suggesting the need to maintaining a minimum level of PAs for morphogenic events to take place.

Up-regulation of NADPH oxidase components and increased production of interferon-gamma by leukocytes from sickle cell disease patients

We have previously demonstrated that mononuclear leukocytes from patients with sickle cell disease (SCD) release higher amounts of superoxide compared with normal controls. The aim of this study was to further study the NADPH oxidase system in these patients by investigating gene expression of NADPH oxidase components, phosphorylation of p47(phox) component, and the release of cytokines related to NADPH oxidase activation in mononuclear leukocytes from patients with SCD. gp91(phox) gene expression was significantly higher in monocytes from SCD patients compared with normal controls (P = 0.036). Monocytes from SCD patients showed higher levels of p47 phox phosphorylation compared with normal controls. INF-gamma release by lymphocytes from SCD patients was significantly higher compared with normal controls, after 48 h culture with phytohemagglutinin (P = 0.02). The release of TNF-alpha by monocytes from SCD patients and normal controls was similar after 24 and 48 h culture with lipopolysaccharide (P > 0.05). We conclude that monocytes from SCD patients show higher levels of gp91(phox) gene expression and p47(phox) phosphorylation, along with increased IFN-gamma release by SCD lymphocytes. These findings help to explain our previous observation showing the increased respiratory burst activity of mononuclear leukocytes from SCD patients and may contribute to inflammation and tissue damage in these patients.

Diapocynin versus apocynin as pretranscriptional inhibitors of NADPH oxidase and cytokine production by peripheral blood mononuclear cells

Apocynin has been extensively used as an inhibitor of NADPH oxidase (NOX) in many experimental models using phagocytic and non-phagocytic cells. Currently, there is some controversy about the efficacy of apocynin in non-phagocytic cells, but in phagocytes the reported results are consistent, which could be due to the presence of myeloperoxidase in these cells. This enzyme has been proposed as responsible for activating apocynin by generating its dimer, diapocynin, which is supposed to be the active compound that prevents NADPH oxidase complex assembly and activation. Here, we synthesized diapocynin and studied its effect on inhibition of gp91(phox) RNA expression. We found that diapocynin strongly inhibited the expression of gp91(phox)mRNA in peripheral blood mononuclear cells (PBMC). Only at a higher concentration, apocynin was able to exert the same effect. We also compared the apocynin and diapocynin efficacy as inhibitors of tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) production in response to lipopolysaccharide (LPS)-activated PBMC. Although apocynin did inhibit TNF-alpha production, diapocynin had a much more pronounced effect, on both TNF-alpha and IL-10 production. In conclusion, these findings suggest that the bioconversion of apocynin to diapocynin is an important issue not limited to enzymatic activity inhibition, but also for other biological effects as gp91(phox) mRNA expression and cytokine production. Hence, as diapocynin can be easily prepared from apocynin, a one-step synthesis, we recommend its use in studies where the biological effects of apocynin are searched. (C) 2010 Elsevier Inc. All rights reserved.

Neurodegeneration and Increased Production of Nitrotyrosine, Nitric Oxide Synthase, IFN-gamma and S100 beta Protein in the Spinal Cord of IL-12p40-Deficient Mice Infected with Trypanosoma cruzi

Background/Aim: Chagas` disease is caused by Trypanosoma cruzi and occurs in most Latin American countries. The protozoan may colonize the central nervous system (CNS) of immune-compromised human hosts, thus causing neuronal disorders. Systemic control of the intracellular forms of the parasite greatly depends on the establishment of a TH1 response and subsequent nitric oxide (NO) release. At the CNS, it is known that low concentrations of NO promote neuronal survival and growth, while high concentrations exert toxic effects and neuron death. Accounting for NO production by astrocytes is the glia-derived factor S100 beta, which is overproduced in some neurodegenerative diseases. In the current work, we studied the expression of NO, interferon (IFN)-gamma and S100 beta in the spinal cord tissue of IL-12p40KO mice infected with T. cruzi, a model of neurodegenerative process. Methods: IL-12p40KO and wild-type (WT) female mice infected with T. cruzi Sylvio X10/4 (10(5) trypomastigotes, intraperitoneally) were euthanized when IL-12p40KO individuals presented limb paralysis. Spinal cord sections were submitted to immunohistochemical procedures for localization of neurofilament, laminin, nitrotyrosine, NO synthases (NOS), IFN-gamma and S100 beta. The total number of neurons was estimated by stereological analysis and the area and intensity of immunoreactivities were assessed by microdensitometric/morphometric image analysis. Results: No lesion was found in the spinal cord sections of WT mice, while morphological disarrangements, many inflammatory foci, enlarged vessels, amastigote nests and dying neurons were seen at various levels of IL-12p40KO spinal cord. Compared to WT mice, IL-12p40KO mice presented a decrement on total number of neurons (46.4%, p<0.05) and showed increased values of immunoreactive area for nitrotyrosine (239%, p<0.01) and NOS (544%, p<0.001). Moreover, the intensity of nitrotyrosine (16%, p<0.01), NOS (38%, p<0.05) and S100 beta (21%, p<0.001) immunoreactivities were also augmented. No IFN-gamma labeled cells were seen in WT spinal cord tissue, contrary to IL-12p40KO tissue that displayed inflammatory infiltrating cells and also some parenchymal cells positively labeled.Conclusion: We suggest that overproduction of NO may account for neuronal death at the spinal cord of T. cruzi-infected IL-12p40KO mice and that IFN-gamma and S100 beta may contribute to NOS activation in the absence of IL-12. Copyright (C) 2009 S. Karger AG, Basel

Relative populations and toxin production by Aspergillus flavus and Fusarium verticillioides in artificially inoculated corn at various stages of development under field conditions

The presence, development and production of mycotoxins by Aspergillus flavus and Fusarium verticillioides were studied in corn ears under field conditions after artificial contamination of corn silks. The planted area was divided into five treatments: T1, inoculated with A.flavus solution containing 1 x 10(8) spores, ears covered; T2, inoculated with F. verticillioides solution containing 1 X 10(8) spores, ears covered; T3, inoculated with E verticillioides plus A. flavus solution containing 1 x 10(8) spores of each, ears covered; T4, sprayed with sterile phosphate-buffered saline, ears covered; TS, non-sprayed silks, uncovered ears. Soil and air samples were also collected and analysed for the occurrence of fungi. Water activity, relative air humidity, rainfall and temperature were determined to assess the correlation between abiotic factors and the presence of fungi in the samples. Contamination with the inoculated fungus predominated in T1 and T2. In the other treatments, F. verticillioides was the most frequently isolated contaminant irrespective of treatment. Considering the production of mycotoxins, a positive relation between the production of fumonisins B-1 and B-2 and the frequency of F. verticillioides was statistically verified in all treatments. (C) 2007 Society of Chemical Industry.

Influence of macro- and micronutrient fertilization on fungal contamination and fumonisin production in corn grains

The aim of this study was to investigate the effects of nutrients (nitrogen, zinc and boron) on fungal growth and fumonisins production in corn samples obtained at the beginning of grain formation and at harvest. Three nitrogen doses were applied to the corn plants through soil in combination with three zinc doses and two boron doses during sowing. Mycological analysis of grains, using Dichloran Rose-Bengal Chloramphenicol Agar, collected at the beginning of formation demonstrated a fungal population predominantly of yeasts. Analysis of freshly harvested corn revealed a higher frequency of Penicillium spp. (72%) and F verticillioides (27%). High Performance Liquid Chromatography analysis revealed that 100% of grains were contaminated with fumonisins B, at levels ranging from 0.3 to 24.3 mg/kg and 93% contaminated with fumonisin B(2) at levels ranging from 0.05 to 5.42 mg/kg. Nitrogen (50 kg/ha) in combination with boron (0.5 kg/ha) resulted in an increased fumonisin B2 production. (c) 2007 Elsevier Ltd. All rights reserved.

Molecular characterization and fumonisin production by Fusarium verticillioides isolated from corn grains of different geographic origins in Brazil

Gibberella moniliformis is most commonly associated with maize worldwide and produces high levels of fumonisins, some of the most agriculturally important mycotoxins. Studies demonstrate that molecular methods can be helpful for a rapid identification of Fusarium species and their levels of toxin production. The purpose of this research was to apply molecular methods (AFLP, TEF-1 alpha partial gene sequencing and PCR based on MAT alleles) for the identification of Fusarium species isolated from Brazilian corn and to verify if real time RT-PCR technique based on FUM1 and FUM19 genes is appropriated to estimate fumonisins B(1) and B(2) production levels. Among the isolated strains, 96 were identified as Fusarium verricillioides, and four as other Fusarium species. Concordant phylogenies were obtained by AFLP and TEF-1 alpha sequencing, permitting the classification of the different species into distinct clades. Concerning MAT alleles, 70% of the F. verricillioides isolates carried the MAT-1 and 30% MAT-2. A significant correlation was observed between the expression of the genes and toxin production r=0.95 and r=0.79 (correlation of FUM1 with FB(1) and FB(2), respectively, P < 0.0001): r=0.93 and r =0.78 (correlation of FUM19 with FB(1) and FB(2). respectively, P < 0.0001). Molecular methods used in this study were found to be useful for the rapid identification of Fusarium species. The high and significant correlation between FUM1 and FUM19 expression and fumonisins production suggests that real time RT-PCR is suitable for studies considering the influence of abiotic and biotic factors on expression of these genes. This is the first report concerning the expression of fumonisin biosynthetic genes in Fusarium strains isolated from Brazilian agricultural commodity. (c) 2010 Elsevier B.V. All rights reserved.

Use of L-Proline and ATP Production by Trypanosoma cruzi Metacyclic Forms as Requirements for Host Cell Invasion

The process of host cell invasion by Trypanosoma cruzi depends on parasite energy. What source of energy is used for that event is not known. To address this and other questions related to T. cruzi energy requirements and cell invasion, we analyzed metacyclic trypomastigote forms of the phylogenetically distant CL and G strains. For both strains, the nutritional stress experienced by cells starved for 24, 36, or 48 h in phosphate-buffered saline reduced the ATP content and the ability of the parasite to invade HeLa cells proportionally to the starvation time. Inhibition of ATP production by treating parasites with rotenone plus antimycin A also diminished the infectivity. Nutrient depletion did not alter the expression of gp82, the surface molecule that mediates CL strain internalization, but increased the expression of gp90, the negative regulator of cell invasion, in the G strain. When L-proline was given to metacyclic forms starved for 36 h, the ATP levels were restored to those of nonstarved controls for both strains. Glucose had no such effect, although this carbohydrate and L-proline were transported in similar fashions. Recovery of infectivity promoted by L-proline treatment of starved parasites was restricted to the CL strain. The profile of restoration of ATP content and gp82-mediated invasion capacity by L-proline treatment of starved Y-strain parasites was similar to that of the CL strain, whereas the Dm28 and Dm30 strains, whose infectivity is downregulated by gp90, behaved like the G strain. L-Proline was also found to increase the ability of the CL strain to traverse a gastric mucin layer, a property important for the establishment of T. cruzi infection by the oral route. Efficient translocation of parasites through gastric mucin toward the target epithelial cells in the stomach mucosa is an essential requirement for subsequent cell invasion. By relying on these closely associated ATP-driven processes, the metacyclic trypomastigotes effectively accomplish their internalization.

Calculations of vibrational frequencies, Raman activities and degrees of depolarization for complexes involving water, methanol and ethanol

Raman activities and degrees of depolarization are reported for 14 complexes involving methanol, ethanol and water using the MP2/aug-cc-pVDZ model. For ethanol both trans and gauche isomers are considered. The red-shifts of the OH stretching and the blue shifts of the bending tau(CO-OH) mode were analyzed for the proton-donor molecules upon hydrogen bond. The shift of the nu(CO) stretching mode of the alcohol molecules are also analyzed and found to be specific giving characterization of the amphoteric relation, being positive for the proton-acceptor and negative for the proton-donor molecule. (c) 2008 Elsevier B.V. All rights reserved.

Culture and exopolysaccharide production from sugarcane molasses by Gordonia polyisoprenivorans CCT 7137, isolated from contaminated groundwater in Brazil

Gordonia polyisoprenivorans CCT 7137 was isolated from groundwater contaminated with leachate in an old controlled landfill (Sauo Paulo, Brazil), and cultured in GYM medium at different concentrations of sugarcane molasses (2%, 6%, and 10%). The strain growth was analyzed by monitoring the viable cell counts (c.f.u. mL(-1)) and optical density and EPS production was evaluated at the end of the exponential phase and 24 h after it. The analysis of the viable cell counts showed that the medium that most favored bacterial growth was not the one that favored EPS production. The control medium (GYM) was the one that most favored the strain growth, at the maximum specific growth rate of 0.232 h(-1). Differences in bacterial growth when cultured at three different concentrations of molasses were not observed. Production of EPS, in all culture media used, began during the exponential phase and continued during the growth stationary phase. The highest total EPS production, after 24 h of stationary phase, was observed in 6% molasses medium (172.86 g L(-1)) and 10% (139.47 g L(-1)) and the specific total EPS production was higher in 10% molasses medium (39.03 x10(-11)g c.f.u.(-1)). After the exponential phase, in 2%, 6%, and 10% molasses media, a higher percentage of free exopolysaccharides (EPS) was observed, representing 88.4%, 62.4%, and 64.2% of the total, respectively. A different result was observed in pattern medium, which presented EPS made up of higher percentage of capsular EPS (66.4% of the total). This work is the first study on EPS production by G. polyisoprenivorans strain in GYM medium and in medium utilizing sugarcane molasses as the sole nutrient source and suggests its potential use for EPS production by G. polyisoprenivorans CCT 7137 aiming at application in biotechnological processes.