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Agriculture has gained increasing importance in response to the continuous growth of the world population and constant need for food. To avoid production losses, farmers commonly use pesticides. Mancozeb is a fungicide used in agriculture as this compound is effective in combating fungi that harm crops. However, this fungicide may also produce damage to non-target organisms present in soil and water. Therefore, this study aimed to investigate the influence of exposure to mancozeb on survival rate, locomotor activity, behavior, and oxidative status utilizing adult zebrafish (Danio rerio) as a model following exposure to environmentally relevant concentrations of this pesticide. The experimental groups were negative control, positive control, and mancozeb (0.3; 1.02; 3.47; 11.8 or 40 µg/L). Zebrafish were exposed to the respective treatments for 96 hr. Exposure to mancozeb did not markedly alter survival rate and oxidative status of Danio rerio. At a concentration of 11.8 µg/L, the fungicide initiated changes in locomotor pattern of the animals. The results obtained suggest that the presence of mancozeb in the environment might produce locomotor alterations in adult zebrafish, which subsequently disrupt the animals' innate defense mechanisms. In nature, this effect attributed to mancozeb on non-target organisms might result in adverse population impacts and ecological imbalance.

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Studies have indicated that glyphosate induces endocrine disruption and may adversely affect the male reproductive system. However, evidence of its effects on ovarian function is poorly understood so far, making further studies necessary on the mechanisms of the glyphosate toxicity in the female reproductive system. The aim of this work was to evaluate the effect of a subacute exposure (28 days) to the glyphosate-based formulation Roundup® (1.05, 10.5 and 105 µg/kg b.w. of glyphosate) on steroidogenesis, oxidative stress, systems involved in cell redox control and histopathological parameters in rat ovaries. Hence we quantify plasma estradiol and progesterone by chemiluminescence; non-protein thiol levels, TBARS, superoxide dismutase and catalase activity by spectrophotometry; gene expression of steroidogenic enzymes and redox systems by real-time PCR; and ovarian follicles by optical microscopy. Our results demonstrated that oral exposure increased progesterone levels and the mRNA expression of 3ß-hydroxysteroid dehydrogenase. Histopathological analysis revealed a decrease in the number of primary follicles and an increase in the number of corpus luteum in rats exposed to Roundup®. An imbalance of the oxidative status was also evidenced by decreasing the catalase activity at all groups exposed to the herbicide. Increased lipid peroxidation and gene expression of glutarredoxin and decreased of glutathione reductase were also observed. Our results indicate that Roundup® causes endocrine disruption of hormones related to female fertility and reproduction and changes the oxidative status by altering antioxidant activity, inducing lipid peroxidation, as well as changing the gene expression of the glutathione-glutarredoxin system in rat ovaries.

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Recycled polyvinyl chloride (PVC) microplastics have been detected in the aquatic environment. These recycled microparticles contain chemicals that are released into the environment reaching different organisms. Although the problem of the presence of recycled PVC microparticles in the environment is evident, the toxicological consequences of this contaminant to exposed organisms remains to be better determined. The aim of this study was to investigate the toxicity attributed to exposure to environmentally relevant concentrations of recycled PVC microplastics in adult zebrafish (Danio rerio). The experimental groups were: negative control, vehicle control, positive control, and recycled microplastics (20 ± 5 µm) at 5, 10 or 20 µg/L. Zebrafish (D. rerio) were exposed to respective treatments for 96 hr. Locomotion and oxidative status parameters were measured and mortality recorded. The positive control group presented increased mortality rates and decreased locomotor activity. Animals from the vehicle group did not show marked differences. Finally, no significant disturbances were found in survival rate, locomotion pattern and oxidative status of animals exposed to recycled PVC microparticles at 5, 10 or 20 µg/L. Taken together our results suggest that recycled PVC microplastics in this particle size range do not appear to exert harmful effects on exposed adult D. rerio. However, these results need to be carefully observed due to limitations including size of particle and duration of exposure parameters that might affect ecological consequences. It is suggested that additional studies applying other particles sizes and chronic exposure are needed to more comprehensively verify the toxicity of the contaminant investigated here.

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Ochratoxin A (OTA) is a mycotoxin produced by species of filamentous fungi widely found as a contaminant in food and with high toxic potential. Studies have shown that this toxin causes kidney and liver damage; however, data on the central nervous system effects of exposure to OTA are still scarce. Thus, this study aimed to investigate the effects of exposure to OTA on behavioral and neurochemical parameters in adult zebrafish. The animals were treated with different doses of OTA (1.38, 2.77, and 5.53 mg/kg) with intraperitoneal injections and submitted to behavioral evaluations in the open tank and social interaction tests. Subsequently, they were euthanized, and the brains were used to assess markers associated with oxidative status. In the open tank test, OTA altered distance traveled, absolute turn angle, mean speed, and freezing time. However, no significant effects were observed in the social interaction test. Moreover, OTA also increased glutathione peroxidase (GPx), glutathione-S-transferase (GST), and glutathione reductase (GR) levels and decreased non-protein thiols (NPSH) levels in the zebrafish brain. This study showed that OTA can affect behavior and neurochemical levels in zebrafish.

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BACKGROUND: Curcumin, a polyphenol extracted from the rhizome of Curcuma longa L. (Zingiberaceae), presents neuroprotective properties and can modulate neuronal pathways related to mental disorders. However, curcumin has low bioavailability, which can compromise its use. The micronization process can reduce mean particle diameter and improve this compound's bioavailability and therapeutic potential. METHODS: We compared the behavioral (open tank test, OTT) and neurochemical (thiobarbituric acid reactive substances (TBARS) and non-protein thiols (NPSH) levels) effects of non-micronized curcumin (CUR, 10 mg/kg, ip) and micronized curcumin (MC, 10 mg/kg, ip) in adult zebrafish subjected to a 90-min acute restraint stress (ARS) protocol. RESULTS: ARS increased the time spent in the central area and the number of crossings and decreased the immobility time of the animals in the OTT. These results suggest an increase in locomotor activity and a decrease in thigmotaxis behavior. Both CUR and MC were not able to prevent these effects. Furthermore, ARS also induced oxidative damage by increasing TBARS and decreasing NPSH levels. Both CUR and MC did not prevent these effects. CONCLUSION: ARS-induced behavioral and biochemical effects were not blocked by any curcumin preparation. Therefore, we conclude that curcumin does not have acute anti-stress effects in zebrafish.

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Zebrafish larvae have been widely used in neuroscience and drug research and development. In the larval stage, zebrafish present a broad behavioral repertoire and physiological responses similar to adults. Curcumin (CUR), a major component of Curcuma longa L. (Zingiberaceae), has demonstrated the ability to modulate several neurobiological processes relevant to mental disorders in animal models. However, the low bioavailability of this compound can compromise its in vivo biological potential. Interestingly, it has been shown that micronization can increase the biological effects of several compounds. Thus, in this study, we compared the effects of acute exposure for 30 min to the following solutions: water (control), 0.1% DMSO (vehicle), 1 µM CUR, or 1 µM micronized curcumin (MC) in zebrafish larvae 7 days post-fertilization (dpf). We analyzed locomotor activity (open tank test), anxiety (light/dark test), and avoidance behavior (aversive stimulus test). Moreover, we evaluated parameters of oxidative status (thiobarbituric acid reactive substances and non-protein thiols levels). MC increased the total distance traveled and absolute turn angle in the open tank test. There were no significant differences in the other behavioral or neurochemical outcomes. The increase in locomotion induced by MC may be associated with a stimulant effect on the central nervous system, which was evidenced by the micronization process.

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Schizophrenia pathophysiology has been associated with dopaminergic hyperactivity, NMDA receptor hypofunction, and redox dysregulation. Most behavioral assays and animal models to study this condition were developed in rodents, leaving room for species-specific biases that could be avoided by cross-species approaches. As MK-801 and amphetamine are largely used in mice and rats to mimic schizophrenia features, this study aimed to compare the effects of these drugs in several zebrafish (Danio rerio) behavioral assays. Male and female adult zebrafish were exposed to MK-801 (1, 5, and 10 µM) or amphetamine (0.625, 2.5, and 10 mg/L) and observed in paradigms of locomotor activity and social behavior. Oxidative parameters were quantified in brain tissue. Our results demonstrate that MK-801 disrupted social interaction, an effect that resembles the negative symptoms of schizophrenia. It also altered locomotion in a context-dependent manner, with hyperactivity when fish were tested in the presence of social cues and hypoactivity when tested alone. On the other hand, exposure to amphetamine was devoid of effects on locomotion and social behavior, while it increased lipid peroxidation in the brain. Key outcomes induced by MK-801 in rodents, such as social interaction deficit and locomotor alterations, were replicated in zebrafish, corroborating previous studies and reinforcing the use of zebrafish to study schizophrenia-related endophenotypes. More studies are necessary to assess the predictive validity of preclinical paradigms with this species and ultimately optimize the screening of potential novel treatments.

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2,4-Dichlorophenoxyacetic acid (2,4-D) is one of the most commonly used herbicides worldwide. While the effects of 2,4-D in target organisms are well known, its consequences in nontarget organisms are not fully explained. Therefore, the purpose of this study was to investigate the effects of the herbicide on mitochondrial energy metabolism, oxidative status, and exploratory behavior in adult zebrafish. Animal exposure to 2,4-D increased cytochrome c oxidase and catalase activities and reduced SOD/CAT ratio, moreover, increased the total distance traveled and the number of crossings. Finally, animals exposed to 2,4-D spent more time in the upper zone of the tank and traveled a long distance in the upper zone. Overall, our results indicate the 2,4-D can provoke disabling effects in nontarget organisms. The obtained data showed that exposure to 2,4-D at environmentally relevant concentrations alters mitochondrial metabolism and antioxidant status and disturbs the zebrafish innate behavior.

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Different veterinary drugs have been widely found in surface and groundwater, affecting non-target organisms. Ractopamine (RAC) is one of these drugs found in water bodies. It is a ß-adrenergic agonist used as a feed additive to modulate the metabolism, redirect nutrients from the adipose tissue towards muscles, and increase protein synthesis in swine, cattle, and turkeys. RAC shows toxicological potential, but there is no data about its impacts on the development of non-target organisms, such as zebrafish (Danio rerio). In this study, we evaluated the effect of the exposure to this feed additive on critical parameters (hatching, survival, spontaneous movement, heart rate, and exploratory and locomotor behavior) in zebrafish embryos and larvae. The animals were exposed to RAC hydrochloride at 0.1, 0.2, 0.85, 8.5, and 85 µg/L. Zebrafish exposed to the drug showed increased heart rate at all tested concentrations and alterations on locomotion and exploratory behavior at 85 µg/L. No changes were observed in the survival, hatching rate and spontaneous movement. Our results suggest that RAC present in the environment can induce disabling effects on non-target organisms and elicit an ecological imbalance by increasing the animals' vulnerability to predation due to greater visibility.

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Anxiety disorders are highly prevalent and a leading cause of disability worldwide. Their etiology is related to stress, an adaptive response of the organism to restore homeostasis, in which oxidative stress and glutamatergic hyperactivity are involved. N-Acetylcysteine (NAC) is a multitarget approved drug proved to be beneficial in the treatment of various mental disorders. Nevertheless, NAC has low membrane permeability and poor bioavailability and its limited delivery to the brain may explain inconsistencies in the literature. N-Acetylcysteine amide (AD4) is a synthetic derivative of NAC in which the carboxyl group was modified to an amide. The amidation of AD4 improved lipophilicity and blood-brain barrier permeability and enhanced its antioxidant properties. The purpose of this study was to investigate the effects of AD4 on behavioral and biochemical parameters in zebrafish anxiety models. Neither AD4 nor NAC induced effects on locomotion and anxiety-related parameters in the novel tank test. However, in the light/dark test, AD4 (0.001 mg/L) increased the time spent in the lit side in a concentration 100 times lower than NAC (0.1 mg/L). In the acute restraint stress protocol, NAC and AD4 (0.001 mg/L) showed anxiolytic properties without meaningful effects on oxidative status. The study suggests that AD4 has anxiolytic effects in zebrafish with higher potency than the parent compound. Additional studies are warranted to characterize the anxiolytic profile of AD4 and its potential in the management of anxiety disorders.

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Pesticide commercial mixtures, including the insecticide fipronil and the fungicides pyraclostrobin and methyl-thiophanate, have been used in concomitant pest control, facilitating agricultural management. Their widespread use can lead to soil and water contamination and potentially induce damages in the ecosystem, producing toxic effects in non-target organisms. Despite their toxicological potential, their effects on behavioral and biochemical parameters are not well understood. Here we investigated the effects of the mixture of fipronil and fungicides (MFF) pyraclostrobin and methyl- thiophanate on behavioral and biochemical parameters of oxidative stress in adult zebrafish. Animals exposed to the highest MFF tested concentration showed a decrease in the total distance traveled and in the number of crossings in the different zones of the tank. Furthermore, animals exposed to highest MFF tested concentration spent more time in water surface. In addition, our data showed that the exposure to this preparation promoted a decrease in non-protein thiol content as well as in catalase activity. Finally, pesticide exposure induced an increase in the superoxide dismutase/catalase ratio. Our results indicate that alterations in behavioral and oxidative parameters are involved in MFF toxicity in zebrafish. The antioxidant mechanisms analyzed were altered in concentrations that did not affect zebrafish behavior. Therefore, the assessment of oxidative stress parameters in zebrafish brains could be very useful to detect the early effects of environmental exposure to the MFF.

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The use of pesticides has been growing along with the demand for agricultural products. These compounds, however, are not restricted to the field, spreading easily through the soil, contaminating groundwater and reaching urban centers. Propiconazole is a triazole fungicide that has been increasingly used in agriculture. However, there are few data about its effects on non-target organisms. This study aimed to evaluate the effects of propiconazole in zebrafish. The animals were exposed for 96 h to different concentrations of propiconazole (425, 850, 1700, 8500 ng/L), then submitted to the novel tank test for behavioral analyses. The brains were collected for evaluation of oxidative stress parameters. Exposure to propiconazole (1700 and 8500 ng/L) decreased the number of crossings, entries, and time spent in the top, and increased the time spent in the bottom area of the tank. We also observed an increase in the activities of superoxide dismutase and catalase in zebrafish brain exposed to propiconazole at 425, 850, and 1700 ng/L. We conclude that propiconazole alters normal fish behavior and disrupts oxidative status. More studies are necessary to elucidate the exact mechanism underlying the effects of propiconazole on non-target-organisms.

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Iron (Fe) and manganese (Mn) are metals commonly found at high concentrations in underground water. These metals are essential for the good functioning of living organisms, but high concentrations lead to imbalance, potentiating the appearance of pathologies. This study aimed to evaluate the effect of exposure to naturally occurring metals in groundwater, using zebrafish (Danio rerio) as an experimental model. Thus, zebrafish were exposed to Fe (0.8 and 1.3 mg/L), Mn (0.2 and 0.4 mg/L), and groundwater collected from deep tube wells with Fe and Mn (Fe 0.8/Mn 0.2 mg/L and Fe 1.3/Mn 0.4 mg/L) for 30 days. Bioaccumulation of these metals has been demonstrated in the livers and muscles of zebrafish. Acetylcholinesterase activity changed only in zebrafish muscles in all groups. Sulfhydryl levels changed mainly in the group Mn 0.4. SOD/CAT ratio decreased in the groups Fe 0.8 and 1.3, Mn 0.4, and Fe 0.8/Mn 0.4. An increase in the frequency of micronucleus in all groups was shown as a consequence of these changes. Behavioral parameters (time and distance traveled, mean speed, turn angle, latency, and number of crossings between compartments) have also changed, mainly in the groups Fe 1.3, Mn 0.4, and Fe 1.3/Mn 0.4. Therefore, long-term exposure to Fe and Mn, even at not so high concentrations, may cause biochemical, genotoxic, and behavioral changes in zebrafish.

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There is accumulating evidence on the use of N-acetylcysteine (NAC) in the treatment of patients with neuropsychiatric disorders. As a multi-target drug and a glutathione precursor, NAC is a promising molecule in the management of stress-related disorders, for which there is an expanding field of research investigating novel therapies targeting oxidative pathways. The deleterious effects of chronic stress in the central nervous system are a result of glutamatergic hyperactivation, glutathione (GSH) depletion, oxidative stress, and increased inflammatory response, among others. The aim of this study was to investigate the effects of NAC in zebrafish submitted to unpredictable chronic stress (UCS). Animals were initially stressed or not for 7 days, followed by treatment with NAC (1 mg/L, 10 min) or vehicle for 7 days. UCS decreased the number of entries and time spent in the top area in the novel tank test, which indicate increased anxiety levels. It also increased reactive oxygen species (ROS) levels and lipid peroxidation (TBARS) while decreased non-protein thiols (NPSH) and superoxide dismutase (SOD) activity. NAC reversed the anxiety-like behavior and oxidative damage observed in stressed animals. Additional studies are needed to investigate the effects of this agent on glutamatergic modulation and inflammatory markers related to stress. Nevertheless, our study adds to the existing body of evidence supporting the clinical evaluation of NAC in mood disorders, anxiety, post-traumatic stress disorder, and other conditions associated with stress.

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Jaboticaba Plinia peruviana (Poir.) Govaerts is a Brazilian berry that presents high levels of polyphenols, which may play a key role in preventing cytotoxic and genotoxic effects of harmful agents. Although copper is an essential micronutrient that plays an important role in organisms, high copper concentrations may trigger toxicity to animals and plants. Here, we investigated whether Plinia peruviana hydroalcoholic extract prevents copper-induced cytotoxicity in Allium cepa root cells. Five different anthocyanins and phenolic compounds were identified in Plinia peruviana extract. Importantly, the exposure to 1.53 mg/L copper for 24 h impaired mitotic index, as well as increased mitosis disturbances and triggered DNA damage. Pre-incubation with Plinia peruviana extract (0.25 g/L and 0.75 g/L) for 3 h prevented copper-induced changes in the mitotic index and reduced the number of abnormal cells. In conclusion, we suggest that Plinia peruviana peel extract has protective effects against cellular and genetic disturbances induced by copper.

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BACKGROUND: The enriched environment (EE) is a laboratory housing model that emerged from efforts to minimize the impact of environmental conditions on laboratory animals. Recently, we showed that EE promoted positive effects on behavior and cortisol levels in zebrafish submitted to the unpredictable chronic stress (UCS) protocol. Here, we expanded the characterization of the effects of UCS protocol by assessing parameters of oxidative status in the zebrafish brain and reveal that EE protects against the oxidative stress induced by chronic stress. METHODS: Zebrafish were exposed to EE (21 or 28 days) or standard housing conditions and subjected to the UCS protocol for seven days. Oxidative stress parameters (lipid peroxidation (TBARS), reactive oxygen species (ROS) levels, non-protein thiol (NPSH) and total thiol (SH) levels, superoxide dismutase (SOD) and catalase (CAT) activities were measured in brain homogenate. RESULTS: Our results revealed that UCS increased lipid peroxidation and ROS levels, while decreased NPSH levels and SOD activity, suggesting oxidative damage. EE for 28 days prevented all changes induced by the UCS protocol, and EE for 21 days prevented the alterations on NPSH levels, lipid peroxidation and ROS levels. Both EE for 21 or 28 days increased CAT activity. DISCUSSION: Our findings reinforce the idea that EE exerts neuromodulatory effects in the zebrafish brain. EE promoted positive effects as it helped maintain the redox homeostasis, which may reduce the susceptibility to stress and its oxidative impact.

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The occurrence of ractopamine (RAC) hydrochloride in water bodies is of significant concern due to its ecological impacts and toxicity to humans. RAC hydrochloride is a ß-adrenergic agonist drug used as a feed additive to (1) improve feed efficiency, (2) rate of weight gain, and (3) increase carcass leanness in animals raised for their meat. This drug is excreted by animals in urine and introduced into the environment affecting nontarget organisms including fish. In wastewater released from farms, RAC concentrations were detected from 0.124 µg/L to 30.1 µg/L, and in levels ranging from 1.3 × 10-5 to 5.4 × 10-4 µg/L in watersheds. The aim of this study was to examine the effects of exposure to RAC at 0.1, 0.2, 0.85, 8.5, or 85 µg/L dissolved in water on behavior and oxidative status in adult zebrafish. At 0.85 µg/L, RAC treatment increased exploratory behavior of zebrafish; while at 8.5 µg/L, decreased locomotor and exploratory activities were noted. With respect to oxidative stress biomarkers, results showed that RAC at 0.2 µg/L induced lipid peroxidation and elevated total thiol content in zebrafish brain. All drug tested concentrations produced a fall in nonprotein thiol content. Finally, RAC at 0.85, 8.5, or 85 µg/L increased catalase enzyme activity. Our results demonstrated that the exposure to RAC induced behavioral alterations and oxidative stress in zebrafish.

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ABSTRACT Celtis iguanaea (Jacq.) Sarg., Cannabaceae, is popularly used in the treatment of diabetes mellitus. However, chemical and pharmacological investigations are lacking. In this study, we investigated the effects of the hydroalcoholic extract from C. iguanaea on markers of cardiovascular diseases and the glucose metabolism in cholesterol-fed rats. Therefore, hypercholesterolemic rats (1% cholesterol) were orally treated with C. iguanaea extract (C-150, CI-300, or CI-600 mg/kg) or simvastatin (4 mg/kg) (n = 6) once a day (30 days) with a hypercholesterolemic diet. A control group (C) was given saline. C. iguanaea extract showed significant decreases in serum levels of total cholesterol, LDL-cholesterol, HMG-CoA-reductase, interleukin-1 and 6, TNF-α and IFN-γ when compared to group C (p < 0.001). Hypoglycemic effects were observed along with a decrease of the activity of sucrase (CI-600), maltase (CI-150, CI-300), and an increase in muscle glycogen levels (CI-300). Antioxidant effects were observed in plasma by the decrease of TBARS and increase of nonprotein thiols levels (CI-600). The histopathological analysis showed a significant decrease in the liver fat area for C. iguanaea extract compared to group C (p < 0.001). Our results suggest that the biological effects of C. iguanaea extract could be related to the flavonoids that possibly exert antioxidant, enzymatic inhibitory, and insulin-mimetic effects.

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ABSTRACT Cynara scolymus L., Asteraceae, are traditionally used to treat dyspepsia. This study evaluated the hypolipidemic and antiatherogenic effects of an aqueous extract prepared from the leaves of C. scolymus in rat's model. Hypercholesterolemic rats (1% cholesterol and 0.5% cholic acid for 15 days) were treated (0.5 ml/200 g) with extract of C. scolymus (150, 300, or 600 mg/kg p.o.; n = 6) or simvastatin (4 mg/kg p.o.; n = 6) once per day for 30 days along with hypercaloric diet. A control group (C) was given water (0.5 ml/200 g; n = 6). A high-cholesterol diet was maintained throughout the treatment period. Rats treated with extract of C. scolymus (150, 300, or 600 mg/kg) and simvastatin showed significant decreases in serum levels of total cholesterol (−46.9%, −51.9%, −44%, and −41.9%, respectively) and low-density lipoprotein-cholesterol (LDL-C; −52.1%, −54.8%, −51.9%, and −46.7%, respectively), compared with group C (p < 0.005). Biochemical analyses revealed significant decrease in the concentration of IL-1, IL-6, TNF-α, IFN-γ, C-reactive protein, oxidized-LDL, and antioxidized-LDL in rats treated with extract of C. scolymus (150, 300, or 600 mg/kg). There were no differences in serum ALT enzyme activity between the groups. Our results suggest that hypolipidemic and antiatherogenic effects could be related with the presence of polar substances present in aqueous extract of C. scolymus.

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The jaboticaba tree, Plinia trunciflora (O. Berg) Kausel, is popularly named "jabuticabeira" in Brazil and is used in folk medicine to treat diabetes and chronic inflammation of the tonsils, but studies evaluating the central effects of this species are limited. This study evaluated the antidepressant-like and antioxidant effects of P. trunciflora (PT) aqueous extract, in which five different anthocyanins were identified. PT showed significant ferric-reduction power and DPPH radical scavenging activity in vitro and reduced lipid peroxidation both in vitro and ex vivo. At the behavioural level, PT (400 and 800 mg/kg, i.p.) dose-dependently reduced immobility time in the tail suspension test in Swiss male mice. The identification of bioactive compounds accompanied by the in vitro and ex vivo antioxidant activity of PT suggests that these activities might be related to the antidepressant-like activity of P. trunciflora.