Catalytic Activity of Rare Earth Elements (REEs) in Advanced Oxidation Processes of Wastewater Pollutants: A Review.

In recent years, sewage treatment plants did not effectively remove emerging water pollutants, leaving potential threats to human health and the environment. Advanced oxidation processes (AOPs) have emerged as a promising technology for the treatment of contaminated wastewater, and the addition of catalysts such as heavy metals has been shown to enhance their effectiveness. This review focuses on the use of rare earth elements (REEs) as catalysts in the AOP process for the degradation of organic pollutants. Cerium and La are the most studied REEs, and their mechanism of action is based on the oxygen vacancies and REE ion concentration in the catalysts. Metal oxide surfaces improve the decomposition of hydrogen peroxide to form hydroxide species, which degrade the organics. The review discusses the targets of AOPs, including pharmaceuticals, dyes, and other molecules such as alkaloids, herbicides, and phenols. The current state-of-the-art advances of REEs-based AOPs, including Fenton-like oxidation and photocatalytic oxidation, are also discussed, with an emphasis on their catalytic performance and mechanism. Additionally, factors affecting water chemistry, such as pH, temperature, dissolved oxygen, inorganic species, and natural organic matter, are analyzed. REEs have great potential for enhancing the removal of dangerous organics from aqueous solutions, and further research is needed to explore the photoFenton-like activity of REEs and their ideal implementation for wastewater treatment.

Hormetic Effects of Cerium, Lanthanum and Their Combination at Sub-micromolar Concentrations in Sea Urchin Sperm.

Rare earth elements (REEs) cerium (Ce) and lanthanum (La) and their combination were tested across a concentration range, from toxic (10-4 to 10-5 M) to lower concentrations (10-6 to 10-8 M) for their effects on sea urchin (Sphaerechinus granularis) sperm. A significantly decreased fertilization rate (FR) was found for sperm exposed to 10-5 M Ce, La and their combination, opposed to a significant increase of FR following 10-7 and 10-8 M REE sperm exposure. The offspring of REE-exposed sperm showed significantly increased developmental defects following sperm exposure to 10-5 M REEs vs. untreated controls, while exposure to 10-7 and 10-8 M REEs resulted in significantly decreased rates of developmental defects. Both of observed effects-on sperm fertilization success and on offspring quality-were closely exerted by Ce or La or their combination.

Hygienic assessment of digestate from a high solids anaerobic co-digestion of sewage sludge with biowaste by testing Salmonella Typhimurium, Escherichia coli and SARS-CoV-2.

Anaerobic digestion is a consolidated technology to convert sewage sludge and other organic wastes into biogas and a nutrient-rich fertilizer (i.e. digestate). The origin of sewage sludge does not exclude the potential presence of pathogens (e.g. Salmonella spp. and SARS-CoV-2) in mature digestate that hence could represent a source of sanitary concerns when it is spread on soil for agriculture purpose. Therefore, an experimental study aimed at proving the sanitizing effect of a full scale thermophilic high solids anaerobic digestion process was conducted by monitoring the hygienic characteristics of mature digestate. Although Salmonella spp. was detected in the sewage sludge fed to the full scale plant, the anaerobic digestion treatment demonstrated sanitization capacity since the monitored pathogens were never found in the mature digestate over the entire duration of the monitoring survey. Furthermore, tests on the regrowth of Salmonella Typhimurium and Escherichia coli, artificially inoculated on mature digestate, were also conducted under both anaerobic and aerobic conditions with the aim to assess the effectiveness of mature digestate as microbial growth medium. Concentrations of Salmonella Typhimurium and Escherichia coli were drastically reduced after a short time of incubation under anaerobic process and the two microorganisms already resulted undetectable after 24-48 h, whereas, under aerobic conditions, two microorganisms' concentrations were stably high for longer than 10 days. The combination of no free oxygen, high temperature, anaerobic metabolites (e.g. total ammonium nitrogen, and volatile fatty acids) production, bacteria competition and lack of nutritional elements in mature digestate considerably reduced in 24-48 h the sanitary risks associated to accidently contaminated digestate. Furthermore, a SARS-CoV-2 monitoring survey on mature digestate during 13 months, resulted in the absence of the virus RNA in the analyzed digestate.

Octocrylene: From Sunscreens to the Degradation Pathway during Chlorination Processes: Formation of Byproducts and Their Ecotoxicity Assessment.

Octocrylene is an organic sunscreen whose main action is to absorb UVB radiation and short UVA wavelengths; it is used in various cosmetic products in order to provide an adequate sun-protection factor or to protect the cosmetic formulations themselves from UV radiation. This filter is believed to be a possible endocrine disruptor and is also questioned due to its allergic and/or photoallergic potential. However, it continues to be widely used, and it has been found in various environments, not least those of swimming pools, where it is evidently released by consumers, to the point that it is now considered an emerging micropollutant. The present investigation presents the possible chemical fate of octocrylene in the typical chlorination conditions of wastewater or swimming pools. A total of 11 disinfection byproducts were identified, and 6 were identified for the first time, and separated by HPLC. These products were identified through careful mass spectrometry studies and 1D and 2D NMR experiments. A formation mechanism has been proposed that justifies the chemical structures of all of the compounds identified. The ecotoxicological assessment of octocrylene and their products was carried out by employing Phaeodactylum tricornutum, Brachionus plicatilis and Aliivibrio fischeri as bioindicators. The ecotoxicity results reveal that toxic byproducts might be generated during the oxidation process, increasing the potential risk to the marine environment.

Ecotoxicity and photodegradation of Montelukast (a drug to treat asthma) in water.

The present work focuses on the ecotoxicological effects of montelukast sodium (MTL) and its photoproducts, obtained under environmentally-like conditions. Despite of the potential presence in surface waters and the common use of MTL as asthma drug, limited data has been published for its photodegradation, while no information is available for its ecotoxicity. Light-induced degradation is an effective way for drugs to degrade in aquatic environments, and MTL is highly photosensitive, even by exposure to sunlight. In this study, solar-simulated irradiation of the drug in water was investigated. The drug was quickly converted into a series of photoproducts that were spectroscopically characterized. The possible photoreaction pathways were proposed. Ecotoxicity tests were performed on parent compound and mixture of photoproducts towards two bioindicators (Raphidocelis subcapitata and Daphnia magna). Results evidenced that effects of MTL on D. magna (EC50 = 16.4 mg/L) were greater than effects on R. subcapitata (EC50 = 195.7 mg/L). Microscopy observations revealed that MTL had mainly accumulated in the gut of daphnia. Toxicity data on photolysed solutions highlighted the presence of residual toxicity in all samples, evidencing that no complete mineralization occurred. Future research should focus on monitoring of MTL concentrations in the environment and study its effects in bioaccumulation tests.

Effects of commercial formulations of glyphosate on marine crustaceans and implications for risk assessment under temperature changes.

Glyphosate-based formulations are the most commonly used herbicides worldwide with the risk of potential contamination of aquatic bodies. The present study assessed the response of four marine crustaceans to three different brands of herbicides Roundup®Platinum, Efesto® and Taifun® MK CL.T, under two selected temperatures of 20 °C and 30 °C. The harpacticoid copepod Tigriopus fulvus, the anostracan Artemia franciscana, the amphipod Corophium insidiosum and the isopod Sphaeroma serratum were chosen as testing organisms. Effects of herbicides and temperatures were assessed by estimating lethal concentrations. The results showed that the high temperature rises the toxicity of glyphosate with an increase of mortality of all the tested species. This is an important aspect for future risk assessments of pesticides under global climate change scenarios. Efesto® resulted the most toxic brand, showing C. insidiosum the most sensitive with 96 h-LC50 values of 3.25 mg/L acid equivalent (a.e.) at 30 °C and 7.94 mg/L a.e. at 20 °C followed by T. fulvus while A. franciscana and S. serratum were the less sensitive. This study provides important information for assessing the toxic effects of three different brands of glyphosate-based herbicides on non-target marine organisms suggesting that they should be carefully managed to minimize any negative impact on marine organisms.

Sub-Chronic Effects of Slight PAH- and PCB-Contaminated Mesocosms in Paracentrotus lividus Lmk: A Multi-Endpoint Approach and De Novo Transcriptomic.

Sediment pollution is a major issue in coastal areas, potentially endangering human health and the marine environments. We investigated the short-term sublethal effects of sediments contaminated with polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) on the sea urchin Paracentrotus lividus for two months. Spiking occurred at concentrations below threshold limit values permitted by the law (TLVPAHs = 900 µg/L, TLVPCBs = 8 µg/L, Legislative Italian Decree 173/2016). A multi-endpoint approach was adopted, considering both adults (mortality, bioaccumulation and gonadal index) and embryos (embryotoxicity, genotoxicity and de novo transcriptome assembly). The slight concentrations of PAHs and PCBs added to the mesocosms were observed to readily compartmentalize in adults, resulting below the detection limits just one week after their addition. Reconstructed sediment and seawater, as negative controls, did not affect sea urchins. PAH- and PCB-spiked mesocosms were observed to impair P. lividus at various endpoints, including bioaccumulation and embryo development (mainly PAHs) and genotoxicity (PAHs and PCBs). In particular, genotoxicity tests revealed that PAHs and PCBs affected the development of P. lividus embryos deriving from exposed adults. Negative effects were also detected by generating a de novo transcriptome assembly and its annotation, as well as by real-time qPCR performed to identify genes differentially expressed in adults exposed to the two contaminants. The effects on sea urchins (both adults and embryos) at background concentrations of PAHs and PCBs below TLV suggest a need for further investigations on the impact of slight concentrations of such contaminants on marine biota.

PAHs and PCBs Affect Functionally Intercorrelated Genes in the Sea Urchin Paracentrotus lividus Embryos.

Polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) represent the most common pollutants in the marine sediments. Previous investigations demonstrated short-term sublethal effects of sediments polluted with both contaminants on the sea urchin Paracentrotus lividus after 2 months of exposure in mesocosms. In particular, morphological malformations observed in P. lividus embryos deriving from adults exposed to PAHs and PCBs were explained at molecular levels by de novo transcriptome assembly and real-time qPCR, leading to the identification of several differentially expressed genes involved in key physiological processes. Here, we extensively explored the genes involved in the response of the sea urchin P. lividus to PAHs and PCBs. Firstly, 25 new genes were identified and interactomic analysis revealed that they were functionally connected among them and to several genes previously defined as molecular targets of response to the two pollutants under analysis. The expression levels of these 25 genes were followed by Real Time qPCR, showing that almost all genes analyzed were affected by PAHs and PCBs. These findings represent an important further step in defining the impacts of slight concentrations of such contaminants on sea urchins and, more in general, on marine biota, increasing our knowledge of molecular targets involved in responses to environmental stressors.

LC and NMR Studies for Identification and Characterization of Degradation Byproducts of Olmesartan Acid, Elucidation of Their Degradation Pathway and Ecotoxicity Assessment.

The discovery of various sartans, which are among the most used antihypertensive drugs in the world, is increasingly frequent not only in wastewater but also in surface water and, in some cases, even in drinking or groundwater. In this paper, the degradation pathway of olmesartan acid, one of the most used sartans, was investigated by simulating the chlorination process normally used in a wastewater treatment plant to reduce similar emerging pollutants. The structures of nine isolated degradation byproducts (DPs), eight of which were isolated for the first time, were separated via chromatography column and HPLC methods, identified by combining nuclear magnetic resonance and mass spectrometry, and justified by a proposed mechanism of formation beginning from the parent drug. Ecotoxicity tests on olmesartan acid and its nine DPs showed that 50% of the investigated byproducts inhibited the target species Aliivibrio fischeri and Raphidocelis subcapitata, causing functional decreases of 18% and 53%, respectively.

Secondary Effects of Hypochlorite Treatment on the Emerging Pollutant Candesartan: The Formation of Degradation Byproducts and Their Toxicological Profiles.

In recent years, many studies have reported the frequent detection of antihypertensive agents such as sartans (olmesartan, valsartan, irbesartan and candesartan) in the influents and effluents of wastewater treatment plants (WWTPs) and in the superficial waters of rivers and lakes in both Europe and North America. In this paper, the degradation pathway for candesartan (CAN) was investigated by simulating the chlorination process that is normally used to reduce microbial contamination in a WWTP. Twelve isolated degradation byproducts (DPs), four of which were isolated for the first time, were separated on a C-18 column by employing a gradient HPLC method, and their structures were identified by combining nuclear magnetic resonance and mass spectrometry and comparing the results with commercial standards. On the basis of these results, a mechanism of formation starting from the parent drug is proposed. The ecotoxicity of CAN and its DPs was studied by conducting a battery of ecotoxicity tests; bioassays were performed using Aliivibrio fischeri (bacterium), Daphnia magna (planktonic crustacean) and Raphidocelis subcapitata (alga). The ecotoxicity results shed new light on the increased toxicity of DPs compared with the parent compound.

Eobania vermiculata as a potential indicator of nitrate contamination in soil.

The effects of nitrates were analysed on the land snail Eobania vermiculata, a good bioindicator to assess the effects of certain pollutants in soil. It is known that the molluscs are very sensitive to contamination substances and can be used as sentinel organism for environmental pollution assessment. The nitrates are present in fertilizers and in food additives and their excess can not only be harmful to the environment but also dangerous for the humans. Indeed, in the mammals the nitrates are converted into nitrites and can cause a series of complications as the formation of methaemoglobin and cancers. In this study, adult organisms of E. vermiculata were exposed to soil containing 2000 mg/L of nitrates for 30 days to evaluate the stool microbiome and the histological changes at the level of the foot. Eggs of these snails were similarly treated to observe their hatching, survival and development. Histological changes were observed at level of the foot of adult snails exposed to nitrate and in their stools was evident an increase of bacteria, especially those that have a high ability to exploit nitrates and nitrogen as nutrients. Instead, the treated eggs showed changes in hatching, hypopigmentation of newborn snails and a decrease of their survival in time. The overall information obtained from these endpoints can provide important information regarding the quality of the environment. In addition, they also showed that the invertebrate organism E. vermiculata despite being a simple organism is very useful and efficient for ecotoxicological studies.

Combined Effects of Diatom-Derived Oxylipins on the Sea Urchin Paracentrotus lividus.

Oxylipins are diatom-derived secondary metabolites, deriving from the oxidation of polyunsatured fatty acids that are released from cell membranes after cell damage or senescence of these single-celled algae. Previous results revealed harmful toxic effects of polyunsaturated aldehydes (PUAs) and hydroxyacids (HEPEs) on sea urchin Paracentrotus lividus embryonic development by testing individual compounds and mixtures of the same chemical group. Here, we investigated the combined effects of these compounds on sea urchin development at the morphological and molecular level for the first time. Our results demonstrated that oxylipin mixtures had stronger effects on sea urchin embryos compared with individual compounds, confirming that PUAs induce malformations and HEPEs cause developmental delay. This harmful effect was also confirmed by molecular analysis. Twelve new genes, involved in stress response and embryonic developmental processes, were isolated from the sea urchin P. lividus; these genes were found to be functionally interconnected with 11 genes already identified as a stress response of P. lividus embryos to single oxylipins. The expression levels of most of the analyzed genes targeted by oxylipin mixtures were involved in stress, skeletogenesis, development/differentiation, and detoxification processes. This work has important ecological implications, considering that PUAs and HEPEs represent the most abundant oxylipins in bloom-forming diatoms, opening new perspectives in understanding the molecular pathways activated by sea urchins exposed to diatom oxylipins.

Molecular and Morphological Toxicity of Diatom-Derived Hydroxyacid Mixtures to Sea Urchin Paracentrotus lividus Embryos.

Oxylipins such as polyunsaturated aldehydes (PUAs) and hydroxyacids (HEPEs) are signaling molecules derived from the oxidation of polyunsaturated fatty acids. They are common in diatoms that constitute a major group of microalgae in freshwater and oceanic ecosystems. Although HEPEs represent the most common oxylipins produced by diatoms, little information is available on their effects on marine invertebrates, and most of the information has been obtained by testing individual HEPEs. Our previous studies reported that four hydroxyacids, i.e., 5-, 9-, 11-, and 15-HEPE, were able to induce malformations and a marked developmental delay in sea urchin Paracentrotus lividus embryos, which had not been reported for other oxylipins. Here, we tested a mixture of 5-, 9-, 11-, and 15-HEPE at different concentrations for the first time. The results showed that mixtures of HEPEs have synergistic effects that are much more severe compared to those of individual HEPEs: The HEPE mixtures induced malformations in sea urchin embryos at lower concentrations. Increasing HEPE mixture concentrations induced a marked increase in the number of delayed embryos, until all embryos were delayed at the highest concentration tested. At the molecular level, the HEPE mixtures induced variations in the expression of 50 genes involved in different functional processes, mainly down-regulating these genes at the earliest stages of embryonic development. These findings are ecologically significant, considering that during diatom blooms, sea urchins could accumulate HEPEs in concentrations comparable to those tested in the present study.

Antibiotic effects on seed germination and root development of tomato (Solanum lycopersicum L.).

Antibiotics are emerging pollutants released into the environment through wastewater and manure or effluents from livestock plants. Compared to the wide literature on the effects of antibiotics on the development of drug-resistant bacteria and on the adverse effects on animals and human beings, the effects on plants are less investigated. Here we evaluated the effects of four antibiotics (cloramphenicol: CAP, spiramycin: SPR, spectinomycin: SPT, vancomycin: VAN) belonging to different chemical groups, on seed germination and root development of tomato (Solanum lycopersicum L. cv. San Marzano). Specifically, seed germination and root elongation kinetics, as well as the number of mithotic figures in root apical meristem, were studied in relation to different concentrations of each antibiotic (0, 0.1, 1, 10, 100, 1000mgL-1) for 10 and 7 days, respectively. Results showed that seed germination was not affected, but root development (root elongation kinetics and cell division) was impaired at concentrations from 10mgL-1 (SPT) and 100mgL-1 (CAP) to 1000mgL-1 (SPR and VAN).

Assessing the potential phytotoxicity of digestate from winery wastes.

In this study, digestate from winery wastes was investigated focusing on phytotoxicity using macrophytes and evaluating the potential contribution of ammonium and copper. Spreading of digestate on soil could represent a suitable approach to recycle nutrients and organic matter, creating an on site circular economy. In this study, digestate quality was evaluated considering both chemical-physical characteristics and biological toxicity applying germination test. The effluent did not meet the entire amendment quality standard defined by Italian law (Decree 75/2010 germination index > 60% with solution of 30% v/v of digestate), but bio-stimulation was observed at low doses (3.15-6.25% v/v) for S. alba and S. saccharatum. The beneficial concentration agreed with Nitrate Directive dose and suggested that limited addition of digestate could have several positive effects on soil characteristics and on crop growth. Specific test using ammonium and copper solutions showed that these pollutants were not directly correlated to observed phytotoxicity.

Assessment of metal pollution in the Lambro Creek (Italy).

This study assessed the effect of metal pollution in the Lambro Creek (Southern Italy). Water, sediment and biota were collected at six sampling sites (June) for metal concentration assessment (Cr, Cu, Ni, Pb and Zn). Sequential extraction was performed to determine the distribution of metals in different geochemical sediment fractions. The influence of pH and leaching time on the release of metals from sediment to the water column was investigated via remobilization tests. A battery of toxicity tests (Vibrio fischeri, Raphidocelis subcapitata, Phaeodactylum tricornutum, and Daphnia magna) with multi-endpoints (bioluminescence, growth inhibition, and immobilization) was used to determine the overall toxicity in sediment water extracts. The results showed that metals did not exceed the probable effect concentration levels, with Cr concentration exceeding the threshold effect concentration level at all sampling points except for the one closer to the source of the creek, suggesting potential negative effect on the biota. Considering the cumulative criterion unit, sediment contamination was moderate at all sampling sites, except for L3 and L5 where biota was exposed to a very high risk. With respect to sequential analysis, the most readily available fraction of metal can be generalised as Ni > Cr > Cu > Zn > Pb. For better understanding the fate of metals in the water-sediment environment, their biogeochemical cycles should also be investigated in small creeks including both fresh (watercourse) and saltwater (river mouth) sediments.

Biomonitoring of nutrient and toxic element concentrations in the Sarno River through aquatic plants.

The Sarno River is considered the most polluted river in Europe and one of the ten most polluted rivers in the world. So far, its quality has been usually evaluated by water and sediment analyses of either inorganic or organic pollutants. However, a biomonitoring approach would be of paramount importance in the evaluation of river quality, since it integrates pollutant temporal fluctuations, as in the case of discontinuous inputs from urban, industrial and agricultural activities. To this end, a passive biomonitoring study of the Sarno River was carried out, using two native aquatic plants accumulators of inorganic pollutants. The spring area was monitored analysing the roots of the semi-submerged Apium nodiflorum, whereas the whole river course was monitored analysing the shoots of the submerged Potamogeton pectinatus. The information on the four macronutrient (Ca, K, Mg, P), the six micronutrient (Cu, Fe, Mn, Na, Ni, Zn) and the four toxic element (Cd, Cr, Pb, V) concentrations were separately combined in the Nemerow Pollution Index. Results evidenced a severe pollution degree of the Sarno River, attributable to toxic elements > micronutrients > macronutrients. In particular, the spring area showed high K concentrations, as well as high concentrations of several micronutrients and toxic elements. A generalized Zn contamination and a progressive macronutrient (above all Ca and P), micronutrient (above all Ni, Cu and Fe) and toxic element (above all Cr and Pb) accumulation toward the mouth was related to pollution from agricultural and urban activities. Industrial sources, especially tanneries along the Solofrana tributary, accounted for high Mn concentrations, whereas the volcanic origin of the substrate accounted for a generalized V contamination.

Combined effects of arsenic, salinity and temperature on Crassostrea gigas embryotoxicity.

The combined effects of different salinity and temperature levels on the toxicity of Arsenic (As) were studied on the embryonic development of the oyster Crassostrea gigas. A standardized embryotoxicity test was performed to assess the interactive effects of these stressors, in a full factorial design experiment including a range of salinities (15, 19, 24, 28 and 32), temperatures (16, 20, 24, 28 and 32°C) and As concentrations (100, 300, 600, 1200, 2400µgL-1). The embryotoxicity endpoint was about the determination of normal larvae development rates at various conditions, and median effect concentration (EC50) determination for each As exposure condition. Results showed that toxicity induced by As was characterized by retardation of embryonic development observing toxic effects at lower concentrations than previously reported studies. The presence of As in seawater resulted in a narrower range of tolerance to both salinity and temperature. These findings bring new insights on the impacts of a common contaminant on an important shellfish species having a planktonic early life stage development, with potential implications for population survival and ecosystem functioning in a changing environment.

Toxicity Effects of Functionalized Quantum Dots, Gold and Polystyrene Nanoparticles on Target Aquatic Biological Models: A Review.

Nano-based products are widespread in several sectors, including textiles, medical-products, cosmetics, paints and plastics. Nanosafety and safe-by-design are driving nanoparticle (NP) production and applications through NP functionalization (@NPs). Indeed, @NPs frequently present biological effects that differ from the parent material. This paper reviews the impact of quantum dots (QDs), gold nanoparticles (AuNPs), and polystyrene-cored NPs (PSNPs), evidencing the role of NP functionalization in toxicity definition. Key biological models were taken into consideration for NP evaluation: Saccharomyces cerevisiae, fresh- (F) and saltwater (S) microalgae (Raphidocelis subcapitata (F), Scenedesmus obliquus (F) and Chlorella spp. (F), and Phaeodactylum tricornutum (S)), Daphnia magna, and Xenopus laevis. QDs are quite widespread in technological devices, and they are known to induce genotoxicity and oxidative stress that can drastically change according to the coating employed. For example, AuNPs are frequently functionalized with antimicrobial peptides, which is shown to both increase their activity and decrease the relative environmental toxicity. P-NPs are frequently coated with NH2- for cationic and COOH- for anionic surfaces, but when positively charged toxicity effects can be observed. Careful assessment of functionalized and non-functionalized NPs is compulsory to also understand their potential direct and indirect effects when the coating is removed or degraded.

Photocatalytic degradation of the antibiotic chloramphenicol and effluent toxicity effects.

Chloramphenicol sodium succinate (CAP, C15H15Cl2N2 Na2O8) is a broad-spectrum antibiotic exhibiting activity against both Gram-positive and Gram-negative bacteria as well as other groups of microorganisms only partially removed by conventional activated sludge wastewater treatment plants. Thus, CAP and its metabolites can be found in effluents. The present work deals with the photocatalytic degradation of CAP using TiO2 as photocatalyst. We investigated the optimization of reaction contact time and concentration of TiO2 considering CAP and its by-products removal as well as effluent ecotoxicity elimination. Considering a CAP real concentration of 25mgL(-1), kinetic degradation curves were determined at 0.1, 0.2, 0.4, 0.8, 1.6 and 3.2gL(-1) TiO2 after 5, 10, 30, 60 and 120min reaction time. Treated samples were checked for the presence of by-products and residual toxicity (V. fischeri, P. subcapitata, L. sativum and D. magna). Results evidenced that the best combination for CAP and its by-products removal could be set at 1.6gL(-1) of TiO2 for 120min with an average residual toxicity of approximately 10%, that is the threshold set for negative controls in most toxicity tests for blank and general toxicity test acceptability.