Browsing by Author "Teixeira, A. V."
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- Flow cytometric method for cell viability evaluation of Gordonia alkanivorans strain 1B in fossil fuels biodesulfurization processesPublication . Teixeira, A. V.; Silva, Tiago; Silva, Teresa Lopes da; Paixão, Susana M.; Alves, LuísThis work reports the development of a rapid flow cytometric method for the viability assessment of Gordonia alkanivorans strain 1B, a bacterium used in the biodesulfurization process. To demonstrate that it is possible to monitor by flow cytometric analysis changes in this bacterium physiological state, positive controls using the 5(6)-carboxyfluorescein diacetate (CFDA) and propidium iodide (PI) staining mixture were set. The loss of viability of G. alkanivorans resting cells in the presence of different concentrations of 2-hydroxybiphenyl, a very toxic end product of the dibenzotiophene desulfurization process, was assessed over a period of time. The results here reported demonstrate the potential of this technique for the biodesulfurization process monitoring and consequent enhancement.
- Flow cytometric method for cell viability evaluation of Gordonia alkanivorans strain 1B in fossil fuels biodesulfurization processesPublication . Teixeira, A. V.; Silva, Tiago; Silva, Teresa Lopes da; Paixão, Susana M.; Alves, LuísThe most commonly method used for sulfur removal from fossil fuels is hydrodesulfurization, a physico-chemical process at very high temperatures and pressures. An alternative to this process is biodedesulfurization (BDS), a microbiological process that works at atmospheric pressure and temperature making it easier to work with and less expensive. It also as the advantage of easily desulfurizing recalcitrant sulfur compounds which are hard to remove by hydrodesulfurization [1]. Several bacteria species, such as Gordonia alkanivorans strain1B [2], are able to desulfurize dibenzothiophene, a model compound used commonly in BDS studies, to 2-hydroxybiphenyl (2-HBP) via the 4S pathway without destroying the carbon structure [3], therefore maintaining the fuel potential energy. BDS limitations are related with process parameters and with the cost of maintaining bacterial cultures so to enhance the BDS process, it is necessary to monitor how changes in the experimental system affect the microbial cells viability and consequently the process efficiency. An alternative method to conventional microbial techniques to determine cell viability is flow cytometry. This method provides a fast and accurate quantitative method for measurement of thousands of individual cells, based on scattered light and fluorescence emitted by specific dyes. The goal of this study was to develop a rapid method for viability assessment of G. alkanivorans cells using flow cytometry for further application to monitor and optimize BDS processes.
- Influence of the carbon source on Gordonia alkanivorans strain 1B resistance to 2-hydroxybiphenyl toxicityPublication . Teixeira, A. V.; Paixão, Susana M.; Silva, Teresa Lopes da; Alves, LuísThe viability of bacteria plays a critical role in the enhancement of fossil fuels biodesulfurization efficiency since cells are exposed to toxic compounds such as 2-hydroxybiphenyl (2-HBP), the end product of dibenzothiophene (DBT) biodesulfurization. The goal of this work was to study the influence of the carbon source on the resistance of Gordonia alkanivorans strain 1B to 2-HBP. The physiological response of this bacterium, pregrown in glucose or fructose, to 2-HBP was evaluated using two approaches: a growth inhibition toxicity test and flow cytometry. The results obtained from the growth inhibition bioassays showed that the carbon source has an influence on the sensitivity of strain 1B growing cells to 2-HBP. The highest IC50 value was obtained for the assay using fructose as carbon source in both inoculum growth and test medium (IC50-48 h=0.464 mM). Relatively to the evaluation of 2-HBP effect on the physiological state of resting cells by flow cytometry, the results showed that concentrations of 2-HBP >1 mM generated significant loss of cell viability. The higher the 2-HBP concentration, the higher the toxicity effect on cells and the faster the loss of cell viability. In overall, the flow cytometry results highlighted that strain 1B resting cells grown in glucose-SO4 or glucose-DBT are physiologically less resistant to 2-HBP than resting cells grown in fructose-SO4 or fructose-DBT, respectively.
- Optimization of low sulfur carob pulp liquor as carbon source for fossil fuels biodesulfurizationPublication . Silva, Tiago; Paixão, Susana M.; Teixeira, A. V.; Roseiro, J. Carlos; Alves, LuísBackground:Biodesulfurization (BDS) is a complementary technology to hydrodesulfurization since it allows the removal of recalcitrant sulfur compounds present in fossil fuels. The cost of culture medium to produce the biocatalysts is still one limitation for BDS application. Carob pulp, as an alternative carbon source, can reduce this cost. However, the presence of sulfates is critical, since BDS is inhibited at very low concentrations. Thus, the goal of this work was to optimize the process of sulfur precipitation on carob pulp liquor. Result:The effect of BaCl2 concentration (0–0.5%) and exposure time (6–36 h) on sulfate removal from carob pulp liquor was studied according to a statistical design following the Doehlert distribution for two factors. This experimental design determined that 0.5% BaCl2 concentration for 21 h were adequate conditions for sulfate removal from carob pulp liquor using BDS.Conclusion:These results demonstrate that it is possible to use alternative carbon sources derived from agro-industrial wastes for BDS, even those with high sulfur levels. For future industrial application, an inexpensive culture medium would have to be employed in a large-scale process and carob pulp liquor could be the carbon source.
- Screening of novel yeast inulinases and further application to bioprocessesPublication . Paixão, Susana M.; Teixeira, Pedro D.; Silva, Tiago; Teixeira, A. V.; Alves, LuísInulin is a carbohydrate composed of linear chains of ß-2,1-linked D-fructofuranose molecules terminated by a glucose residue through a sucrose-type linkage at the reducing end. Jerusalem artichoke (JA) is one of the most interesting materials among unconventional and renewable raw materials, with levels of inulin reaching 50–80% of dry matter. Inulin or inulin-rich materials can be actively hydrolyzed by microbial inulinases to produce glucose and fructose syrups that can be used in bioprocesses. In this study, several microbial strains were isolated and their ability to inulinase biosynthesis was evaluated. The novel yeast strain Talf1, identified as Zygosaccharomyces bailii, was the best inulinase producer, attaining 8.67 U/ml of inulinase activity when JA juice was used as the inducer substrate. Z. bailii strain Talf1 and/or its enzymatic crude extract were further applied for bioethanol production and biodesulfurization (BDS) processes, using inulin and JA juice as carbon source. In a consolidated bioprocessing for ethanol production from 200 g/l inulin, Z. bailii strain Talf1 was able to produce 67 g/l of ethanol. This ethanol yield was improved in a simultaneous saccharification and fermentation (SSF) process, with the ethanologenic yeast Saccharomyces cerevisiae CCMI 885 and the Talf1 inulinases, achieving a production of 78 g/l ethanol. However, the highest ethanol yield (~48%) was obtained in a SSF process from JA juice (~130 g/l fermentable sugars), where the S. cerevisiae produced 63 g/l ethanol. Relatively to the dibenzothiophene BDS tests, the Gordonia alkanivorans strain 1B achieved a desulfurization rate of 4.8 µM/h within a SSF process using Talf1 inulinases and JA juice, highlighting the potential of JA as a less expensive alternative carbon source. These results showed the high potential of Z. bailii strain Talf1 inulinases as a versatile tool for bioprocesses using inulin-rich materials.
- Zygosaccharomyces bailii strain talf1 inulinases: a versatile tool for bioprocessesPublication . Paixão, Susana M.; Teixeira, Pedro D.; Silva, Tiago; Teixeira, A. V.; Alves, LuísFructans are one of the most abundant non-structural polysaccharides found in a wide range of plants. Inulin is a polydisperse fructan polymer composed by linear chains of b-2, 1-linked D-fructofuranose molecules terminated by a glucose residue through a sucrose-type linkage at the reducing end. Inulin or inulin-rich materials can be actively hydrolyzed into fermentable sugars (glucose and fructose) using inulinases and then further used within bioprocesses.