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Sevilla, Spain

Moreno I.M.,Area of Toxicology | Pichardo S.,Area of Toxicology | Moyano R.,University of Cordoba, Spain | Camean A.M.,Area of Toxicology
International Journal of Environmental Analytical Chemistry | Year: 2010

A new HPLC-DAD method has been developed to identify and quantify free microcystins (MC) in biological samples from fish (intestine and liver). The toxins were extracted from 500 mg sample with a mixture of methanol-water (85:15, v/v) and the extracts obtained were purified employing immunoaffinity columns (IAC). The purification step was optimised by a full factorial 3 design. MC were separated using conventional C18 column and an acetonitrile-acidified water (pH 3) gradient. Detection and quantification limits resulted equal for the two toxins assayed (MC-RR and MC-LR) and were 0.15 and 0.5 μg_1, respectively. The accuracy for each MC in liver samples were 96% (range 80-113%) for MC-RR and 101% (range 93-118%) for MC-LR. The results were slightly lower for intestine samples, with recoveries ranging between 85% (75-93%) for MC-RR and 88% (80-97%) for MC-LR. The proposed method was applied for the determination of free MC in fish intoxicated with these toxins, in order to determine its utility to evaluate the potential risks for human health if MC-contaminated fish are consumed. The results showed the transference of MC-LR from cyanobacterial cells to fish tissues. © 2010 Taylor & Francis.

Guzman-Guillen R.,Area of Toxicology | Prieto Ortega A.I.,Area of Toxicology | Gutierrez-Praena D.,Area of Toxicology | Moreno I.M.,Area of Toxicology | And 3 more authors.
Environmental Toxicology | Year: 2015

Cylindrospermopsin (CYN) is a cyanotoxin frequently involved in blooms with a predominantly extracellular availability, which makes it easily taken up by a variety of aquatic organisms. CYN is a potent protein and glutathione synthesis inhibitor, and also induces genotoxicity, oxidative stress and several histopathological lesions. The present study investigates the protective role of a vitamin E pretreatment (700 mg vit E/kg fish bw/day, for 7 days) on the histopathological alterations induced in different organs of tilapia (Oreochromis niloticus) acutely exposed to a single oral dose of 400 μg pure CYN/kg bw fish. The major histological changes observed were degenerative glucogenic process and loss of the hepatic structure in the liver, glomerulopathy and tubular tumefaction in the kidney, myofibrolysis and edema in the heart, catarrhal enteritis and necrosis in the gastrointestinal tract, hyperemic processes in the gill lamellae, and high basophilia, degeneration and tumefaction of granular neurons in the brain. Vitamin E pretreatment was effective in preventing or ameliorating the abovementioned alterations induced by CYN. In addition, a morphometric study indicated that the average nuclear diameter of hepatocytes, and cross-sections of proximal and distal convoluted tubules, together with the cardiac fiber and capillaries diameters represent a useful tool to evaluate the damage induced by CYN. This is the first study reporting vitamin E prevention of histopathological damage in tissues (liver, kidney, heart, gastrointestinal tract, gills and brain) of fish intoxicated with CYN. Therefore, vitamin E can be considered a useful chemoprotectant in the treatment of histopathological changes induced in CYN-intoxicated fish. © 2015 Wiley Periodicals, Inc.

Gutierrez-Praena D.,Area of Toxicology | Jos A.,Area of Toxicology | Pichardo S.,Area of Toxicology | Camean A.M.,Area of Toxicology
Aquatic Toxicology | Year: 2011

Cylindrospermopsin (CYN) is a toxin produced by various cyanobacterial species that are increasingly being found in freshwater systems. Although CYN can have toxic effects in humans, domestic animals and wildlife, it has been subject to very little investigation (particularly in fish). It has been reported to deplete the cellular glutathione content but the role of oxidative stress in the pathogenicity of CYN in fish is unknown. For this reason tilapia fish were exposed to 200. μg/kg pure CYN through two different exposure routes-gavage and intraperitoneal injection-and sacrificed after 24 h and 5 days. The results showed an increase in NADPH oxidase activity (a biomarker of reactive oxygen species formation), lipid peroxidation (LPO) and protein oxidation; no changes in DNA oxidation; and a reduction in glutathione levels (GSH) and γ-glutamylcysteine synthetase (GCS) activity, the limiting enzyme in glutathione synthesis. The time of sacrifice had a bigger influence on the results than the exposure route because after 5 days some of the biomarkers assayed had recovered their pre-intoxication levels, which was not the case after 24 h. © 2011 Elsevier B.V.

Gutierrez-Praena D.,Area of Toxicology | Jos A.,Area of Toxicology | Pichardo S.,Area of Toxicology | Puerto M.,Area of Toxicology | Camean A.M.,Area of Toxicology
Chemosphere | Year: 2013

Cylindrospermopsin is a cyanobacterial toxin frequently implicated in cyanobacterial blooms that is approaching an almost cosmopolitan distribution pattern. Moreover, the predominant extracellular availability of this cyanotoxin makes it particularly likely to be taken up by a variety of aquatic organisms including fish. Recently, Cylindrospermopsin has shown to alter the activity and gene expression of some of the glutathione related enzymes in tilapias (Oreochromis niloticus), but little is known about the influence of the route of exposure and the time of sacrifice after a single exposure to Cylindrospermopsin on these biomarkers. With this aim, tilapias were exposed by gavage or by intraperitoneal injection to a single dose of 200μgkg-1 bw of pure Cylindrospermopsin and after 24h or 5d they were sacrificed. The activity and relative mRNA expression by real-time PCR of antioxidant enzymes glutathione peroxidase and soluble glutathione-S-transferases (sGST) and the sGST protein abundance by Western blot analysis were evaluated in liver and kidney. Results showed differential responses in dependence on the variables considered with a higher toxicity with the intraperitoneal exposure and with 5d as time of sacrifice. © 2012 Elsevier Ltd.

Puerto M.,Area of Toxicology | Gutierrez-Praena D.,Area of Toxicology | Prieto A.I.,Area of Toxicology | Pichardo S.,Area of Toxicology | And 4 more authors.
Ecotoxicology | Year: 2011

The increasing occurrence of toxic cyanobacterial blooms in eutrophic water bodies is nowadays of worldwide concern due to their ability to produce toxins such as microcystins (MCs). These cyanobacterial toxins have been shown to affect aquatic organisms such as fish, resulting in oxidative stress. Among the antioxidant enzymes, glutathione peroxidase (GPx) and soluble glutathione-S-transferases (sGST) play an important role in the detoxification of MCs. In the present work tilapia (Oreochromis niloticus) were orally exposed to cyanobacterial cells containing MCs and non-containing MCs for 21 days. The activity and relative mRNA expression by real-time PCR of both enzymes and the GST protein abundance by Western blot analysis were evaluated in liver and kidney. Also the induction of lipid peroxidation (LPO) was assayed. MCs containing cyanobacterial cells induced an increase of LPO products in both organs, and MCs containing and MCs non-containing cyanobacterial cells altered the activity, gene expression and protein abundance of the enzymes, indicating the importance of GPx and sGST in MCs detoxification. Moreover, liver, the main organ involved in biodegradation and biotransformation, experienced an adaptative response to the toxic insult. These results show for the first time that the subchronic exposure to cyanobacterial cells causes changes in antioxidant and detoxification enzymes and that GPx and GST gene expression are good markers of these alterations in tilapia. © 2011 Springer Science+Business Media, LLC.

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