Susana M. Paixão

Acute toxicity evaluation of olive oil mill wastewaters: A comparative study of three aquatic organisms

Acute toxicity of olive mill wastewaters (traditional and continuous processes) collected from different regions of Portugal was evaluated using three test species (Vibrio fischeri formerly Photobacterium phosphoreum, Thamnocephalus platyurus, and Daphnia magna) and correlated with several physical and chemical parameters. Acute toxicity of these effluents, expressed in LC50 or EC50, ranged from: 0.16 to 1.24% in Microtox test, 0.73 to 12.54% in Thamnotoxkit F test, and 1.08 to 6.83% in Daphnia test. These values reflect the high toxicity of the olive mill wastewaters to all test species. Statistical analysis of the results shows a high correlation between the two microcrustacean bioassays. Microtox test did not correlate significantly with the other bioassays used. A significative correlation (p≤0.05) could also be established between L(E)C50 obtained in the microcrustacean tests and some physicochemical parameters of the effluent.  ©1999 John Wiley & Sons, Inc. Environ Toxicol 14: 263–269, 1999

Ecotoxicity tests in the environmental analysis of wastewater treatment plants: case study in Portugal.

A global evaluation of wastewaters should include ecotoxicological tests to complement the chemical characterization, with advantages especially in the case of complex wastewaters. A European project developed in Trancão River Basin (Portugal), integrated the ecotoxicological and physicochemical studies of wastewater samples from two municipal sewer networks and respective wastewater treatment plants. Wastewater samples were analysed for physicochemical parameters, ecotoxicological acute and chronic tests performed and the potential for endocrine disruption evaluated. Organic load parameters and total suspended solids showed significant correlations with Microtox and ThamnoToxKit test results. Data analysis showed that treated treatment plant effluent samples are associated with less organic contamination and less toxicity in ThamnoToxKit test. Chronic toxicity test and endocrine disruption assay of treatment plant effluent samples indicated that, in a long term, potential population effects could arise in the receiving waters. A test battery to monitor this type of wastewaters is proposed, including tests with a bacterium, an alga and a crustacean. In a screening phase the most sensitive test, Microtox, can be used. The use of an ecotoxicological approach can have added value to hazard and risk assessment of discharges to the receiving waters and can contribute to the environmental management of the treatment plant.

Screening of novel yeast inulinases and further application to bioprocesses.

Inulin is a carbohydrate composed of linear chains of β-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.

Ecotoxicological assessment of industrial wastewaters in Trancão River Basin (Portugal)

It is important to assess the toxicity of complex effluents, since chemical evaluation alone is insufficient to protect the environment. Direct Toxicity Assessment is valuable in the decision process regarding the final disposal of complex wastewaters as it measures the total effects of the discharge, because of its known and unknown chemicals, additionally having some degree of ecological relevance. In Portugal, ecotoxicity tests are not used on a regular basis to control wastewaters. So, an integrated ecotoxicological, physical, and chemical study of wastewaters from 17 industries, in the Trancão River Basin, was carried out viewing proposing a test battery to be used in wastewater evaluation. An approach which does not include an ecotoxicological characterization may not properly evaluate the potential risks of effluent discharges, especially when they are complex. From the study carried out the use of a battery of assays to apply in the monitoring of complex wastewaters was proposed, including Microtox test, Daphnia test, and an algal test. Moreover, the added value of the ecotoxicological assessment of industrial wastewaters was demonstrated and could support the implementation of EU Directives (e.g. IPPC, WFD) within the Portuguese situation.

Toxicity evaluation of 2-hydroxybiphenyl and other compounds involved in studies of fossil fuels biodesulphurisation

The acute toxicity of some compounds used in fossil fuels biodesulphurisation studies, on the respiration activity, was evaluated by Gordonia alkanivorans and Rhodococcus erythropolis. Moreover, the effect of 2-hydroxybiphenyl on cell growth of both strains was also determined, using batch (chronic bioassays) and continuous cultures. The IC₅₀ values obtained showed the toxicity of all the compounds tested to both strains, specially the high toxicity of 2-HBP. These results were confirmed by the chronic toxicity data. The toxicity data sets highlight for a higher sensitivity to the toxicant by the strain presenting a lower growth rate, due to a lower cells number in contact with the toxicant. Thus, microorganisms exhibiting faster generation times could be more resistant to 2-HBP accumulation during a BDS process. The physiological response of both strains to 2-HBP pulse in a steady-state continuous culture shows their potential to be used in a future fossil fuel BDS process.

Biodesulphurization of fossil fuels: energy, emissions and cost analysis

In order to achieve stringent environmental and safety requirements, refineries are in search of “green” and cost-effective methods for crude oil desulphurization. Combined desulphurization technologies are being studied, including bioprocessing to upgrade fossil fuels. Using biodesulphurization (BDS), which is a biochemical process mediated by specific microorganisms, it is possible to desulphurize most of the hydrodesulphurization (HDS) recalcitrant sulphur compounds under mild operating conditions, making it a simple and eco-friendly process. In this study, two BDS process designs are compared, in terms of energy consumption, greenhouse gas emissions and operational costs by following a life cycle assessment (LCA) and life cycle cost (LCC) based methodology. The industrial HDS process is used as the reference technology for sulphur removal from fossil fuels. Different theoretical scenarios were considered and the best BDS results are scaled-up to evaluate a case study of providing ultra low sulphur diesel to an urban taxi fleet. This study exploits the potential of BDS as a cost-effective and eco-friendly alternative or complementary technology to the commonly HDS towards ultra low sulphur fuels.

High-pressure carbon dioxide/water pre-treatment of sugarcane bagasse and elephant grass: Assessment of the effect of biomass composition on process efficiency

The performance of two lignocellulosic biomasses was studied in high-pressure carbon dioxide/water pre-treatment. Sugarcane bagasse and elephant grass were used to produce C5-sugars from hemicellulose and, simultaneously, to promote cellulose digestibility for enzymatic saccharification. Different pre-treatment conditions, with combined severity factor ranging from -1.17 to -0.04, were evaluated and maximal total xylan to xylose yields of 59.2wt.% (34.4wt.% xylooligomers) and 46.4wt.% (34.9wt.% xylooligomers) were attained for sugarcane bagasse and elephant grass, respectively. Furthermore, pre-treated biomasses were highly digestible, with glucan to glucose yields of 77.2mol% and 72.4mol% for sugarcane bagasse and elephant grass, respectively. High-pressure carbon dioxide/water pre-treatment provides high total C5-sugars and glucose recovery from both lignocellulosic biomasses; however it is highly influenced by composition and intrinsic features of each biomass. The obtained results confirm this approach as an effective and greener alternative to conventional pre-treatment processes.

Fructophilic behaviour of Gordonia alkanivorans strain 1B during dibenzothiophene desulfurization process

Biodesulfurization (BDS) aims at the removal of recalcitrant sulfur from fossil fuels at mild operating conditions with the aid of microorganisms. These microorganisms can remove sulfur from dibenzothiphene (DBT), a model compound, or other polycyclic aromatic used as sulfur source, making BDS an easy and environmental friendly process. Gordonia alkanivorans strain 1B has been described as a desulfurizing bacterium, able to desulfurize DBT to 2-hydroxybiphenyl (2-HBP), the final product of the 4S pathway, using d-glucose as carbon source. However, both cell growth and desulfurization can be largely affected by the nutrient composition of the growth medium, due to cofactor requirements of many enzymes involved in the BDS biochemical pathway. In this study, the main goal was to investigate the influence of several sugars, as carbon source, on the growth and DBT desulfurization ability of G. alkanivorans strain 1B. The results of desulfurization tests showed that the lowest values for the growth rate (0.025 hour(-1)) and for the overall 2-HBP production rate (1.80 μm/hour) by the strain 1B were obtained in glucose grown cultures. When using sucrose, the growth rate increase exhibited by strain 1B led to a higher biomass productivity, which induced a slightly increase in the 2-HBP production rate (1.91 μm/hour), conversely in terms of 2-HBP specific production rate (q2-HBP) the value obtained was markedly lower (0.718 μmol/g/hour in sucrose versus 1.22 μmol/g/hour in glucose). When a mixture of glucose and fructose was used as carbon source, strain 1B reached a value of q2-HBP=1.90 μmol/g/hour, close to that in fructose (q2-HBP=2.12 μmol/g/hour). The highest values for both cell growth (μ=0.091 hour(-1)) and 2-HPB production (9.29μm/hour) were obtained when strain 1B was desulfurizing DBT in the presence of fructose as the only carbon source, indicating a fructophilic behaviour by this bacterium. This fact is in agreement with the highest value of biomass productivity by strain 1B be in fructose, which resulted in a higher amount cells fulfilling the DBT-desulfurization. The greater number of functional cells conducted to a more effectiveness BDS process by strain 1B, as they attained a q2-HBP about 74% higher than in glucose grown cultures. Moreover, this significant BDS enhancement can better be observed in terms of the overall 2-HBP production rate, which increased over 5-fold, from 1.80 μm/hour (in glucose) to 9.29 μm/hour (in fructose).

Ecotoxicological evaluation of wastewater in a municipal WWTP in Lisbon area (Portugal)

Abstract Wastewater management has a central role in sustainable development, and, in this context, an integrated management of wastewater treatment plants (WWTP) can be important. WWTP discharge complex effluents and for a new strategy in environmental protection ecotoxicological evaluation should complement the usual chemical evaluation. The EU project WW4Environment was set up for a WWTP located in Lisbon area and discharging into Tagus estuary (Portugal). One of the main objectives of the project is to optimize the management of the WWTP in terms of environmental impact. A battery of toxicity tests with organisms bearing different functions at the ecosystem level (the bacterium Vibrio fischeri, the alga Pseudokirchneriella subcapitata, the crustaceans Thamnocephalus platyurus and Daphnia magna, and the plant Lemna minor) was used to characterize the wastewater in the different treatment phases. V. fischeri, test organism for Microtox test, was the most sensitive species in WWTP samples evaluation. Mic...

A multi-integrated approach on toxicity effects of engineered TiO2 nanoparticles

The new properties of engineered nanoparticles drive the need for new knowledge on the safety, fate, behavior and biologic effects of these particles on organisms and ecosystems. Titanium dioxide nanoparticles have been used extensively for a wide range of applications, e.g, self-cleaning surface coatings, solar cells, water treatment agents, topical sunscreens. Within this scenario increased environmental exposure can be expected but data on the ecotoxicological evaluation of nanoparticles are still scarce. The main purpose of this work was the evaluation of effects of TiO2 nanoparticles in several organisms, covering different trophic levels, using a battery of aquatic assays. Using fish as a vertebrate model organism tissue histological and ultrastructural observations and the stress enzyme activity were also studied. TiO2 nanoparticles (Aeroxide® P25), two phase composition of anatase (65%) and rutile (35%) with an average particle size value of 27.6±11 nm were used. Results on the EC50 for the tested aquatic organisms showed toxicity for the bacteria, the algae and the crustacean, being the algae the most sensitive tested organism. The aquatic plant Lemna minor showed no effect on growth. The fish Carassius auratus showed no effect on a 21 day survival test, though at a biochemical level the cytosolic Glutathione-S-Transferase total activity, in intestines, showed a general significant decrease (p<0.05) after 14 days of exposure for all tested concentrations. The presence of TiO2 nanoparticles aggregates were observed in the intestine lumen but their internalization by intestine cells could not be confirmed.