Among the most important worldwide environmental problems that civilization has recently faced is aquatic pollution [1]. Many of the nation's rivers are dealing with complex pollution issues as a result of industry and the spontaneous growth that characterizes the nation's major cities [2,3]. They are responsible for the dangerous contamination and deterioration of the ecology, especially in the watery habitat [4]. In living things, enzymes are necessary for food utilization and metabolic [5]. Herbicides are a major source of pollution in aquatic environments that can harm living organisms in the short or long term. Fish face many difficulties while interacting with their aquatic environment. One typical physiological reaction to external stresses is the generation of reactive oxygen species (ROS), which can result in oxidative stress [6]. An imbalance in the prooxidant-antioxidant equilibrium that favours prooxidants and causes cellular damage is what defines oxidative stress [7]. An increase in ROS levels, a breakdown in antioxidant defense systems, or an incapacity to repair oxidative damage could all be the cause of this imbalance [8]. Lipid peroxidation, DNA damage, and changes in the activity of antioxidant enzymes including catalase, superoxide dismutase, glutathione reductase, reduced glutathione, and glutathione peroxidase can all result from interactions between elevated ROS and biological macromolecules [9]. Numerous illnesses, including as cancer, respiratory ailments, and neurological conditions, have been linked to oxidative stress.

It is common practice to use aquatic species to manage aquaculture environments and conduct toxicity research [10]. Aquatic pollution has significantly increased in recent years as a result of the widespread and reckless use of primary fertilizers like nitrogen and phosphorus. Aquatic habitats now contain higher concentrations of pollutants, such as industrial chemicals, heavy metals, and pesticides, as a result of human activity. As organic contaminants build up in aquatic environments, oxygen levels drop, increasing mortality rates and disrupting exposed aquatic creatures' ability to operate normally [11]. Understanding the detrimental effects on aquatic creatures' physicochemical conditions, survival rates, and health variations requires research and accurate data collecting [12]. Fish, plants, invertebrates, and vertebrates are examples of aquatic species that are important natural resources that assist lessen the extreme stress brought on by human activity [13]. According to studies, the main causes of pollution in aquatic ecosystems are residential activities, agricultural runoff, and industrial effluents [14]. Aquatic animals have suffered to varied degrees as a result of the growing release of toxins from homes, businesses, and agriculture into aquatic habitats [15]. The use of pesticides in agriculture is a major source of concern for environmental and health professionals since some of these chemicals can negatively impact aquatic systems even when they are sprayed in approved regions, especially when there is rain or flooding. Particularly susceptible to these effects are fish [16]. Furthermore, significant accumulation and an elevated risk of aquatic life mortality might result from pesticides' high solubility, frequent application, unintentional spills, discharge from untreated effluents, and spray drift [17].The objective of this study was to evaluate the level of antioxidant enzyme activity in the plasma of Sarotherodon melanotheron following exposure to varying concentrations of butachlor in the laboratory.

Experimental Site

The study was conducted at the African Regional Aquaculture Center, an outpost of the Nigerian Institute for Oceanography and Marine Research, situated in Buguma, Rivers State, Nigeria. Over the course of seven days, 180 S.melanotheron fish were acclimated, 90 of which were juvenile and adult fish collected from ponds at low tide. The fish were transported to the laboratory in six 50-liter, perforated plastic containers.

Preparation of Test Solutions and Exposure of Fish

The herbicide Butachlor, which belongs to the chloroacetanilide class, is well-known for its pre-emergent ability to kill weeds. It is recommended, especially in rice crops, for the pre-emergence control of certain broad-leaved weeds and annual grasses. It is also widely used as a post-emergence herbicide in rice when it is in the form of granules. The herbicide was purchased in Port Harcourt, Nigeria, from a retailer. S.melanotheron  were exposed to the chemical  in triplicates at doses of 0.05, 0.10, 0.15, and 0.20, mg/L, with 0.00 mg/L serving as the control. Five fish were randomly selected for each test tank. The fifteen-day trial continued. Fresh water was added to the tanks each day. Twice daily, the fish received commercial feed at a rate of 3% of their body weight.

Analytical Procedure

Two millilitres of fresh blood were extracted at the end of each experimental session using a tiny needle to perform a caudal puncture, and the blood was then transferred into heparinized sample containers. Blood samples were centrifuged right away for 15 minutes at 5000 rpm. Plasma samples were pipetted into eppendorf tubes, separated, and stored in a refrigerator at -20°C before to analysis [15]. The results were read using a Jen way visible spectrophotometer (Model 6405) equipped with a universal micro plate reader. Following spectrophotometric analysis, the antioxidant activity in centrifuged plasma was evaluated following the methodology of Beechey et al. [18]. The water quality parameters were also ascertained using the APHA methodology [19].

Statistical Analysis

The mean and standard deviation of the mean were used to express all the data. The data analysis was conducted using SPSS Version 22, a statistical software. Two-way ANOVA was used to separate the means, and at 5% (P<0.05), the two means were deemed significant.

All of the water quality metrics (Table 1) fell within the same range, with the exception of DO, which showed decreasing values as herbicide concentrations rose. Table 2 shows how the antioxidants in the plasma of juvenile S. melanotheron are affected by butachlor. It was shown that when the concentration of herbicide increased, the levels of SOD and GSH decreased. On the other hand, in comparison to the control levels, CAT and LPO increased considerably. Adult fish exposed to the pesticide had antioxidant levels seen in Table 3. SOD and GSH levels dropped as the pesticide concentration increased. On the other hand, in comparison to the control levels, CAT and LPO increased considerably.

Table1: Physico-Chemical Parameters of Water in Experimental Tanks of S.melanotheron Exposed To Butachlor.

Means within the same column with different super scripts are significantly different (P<0.05)

Table 2: Changes in Antioxidants Activities in Juvenile Sizes of S.melanotheron Exposed to Butachlor.

Means within the same column with different super scripts are significantly different (P<0.05)

Key: CAT-Catalase; SOD-Superoxide dismutase; LPO- Lipid peroxidase; GSH-Glutathione

Table 3: Changes in Antioxidants Activities in Adult Sizes of S.melanotheron Exposed to Butachlor.

Means within the same column with different super scripts are significantly different (P<0.05)

Key: CAT- Catalase; SOD- Superoxide dismutase; LPO- Lipid peroxidase; GSH-Glutathione

Oxidative stress is brought on by reactive oxygen metabolites (ROS), which are produced by glyphosate's toxicity. The generation of ROS may interact with cell lipid membranes, causing lipid peroxidation, DNA damage, and disturbance of physiological processes [20]. Butachlor has been connected to oxidative stress and cytoplasmic membrane toxicity, both of which might affect how cells function [19]. Glutathione (GSH), an antioxidant molecule, is activated when there is an imbalance in the generation and elimination of ROS, balancing the MDA concentration [21]. The plasma of the fish used in this investigation showed elevated levels of GSH activity. Superoxide dismutases (SODs) are the first line of defense against free radicals because they can catalyze the conversion of superoxide radicals into hydrogen peroxide and molecular oxygen. Both sizes of S. melanotheron were suppressed by butachlor treatment, in contrast to the control fish in this experiment. According to the study's findings, the treated fish's plasma contained lower amounts of SOD, which suggests that the tissues' ability to fend off oxygen-releasing free radicals has diminished. Similar effects on decreased SOD have been seen in Oreochromis niloticus tissues exposed to heavy metal ingestion [22].

The two S.melanotheron sizes used in this study demonstrated dose- and time-dependent increases in LPO. This may have to do with the capacity of butachlor formulations to generate reactive oxygen species (ROS), which may then interact with fish macromolecules to alter antioxidant levels and induce cell damage. The current study's findings, which show that butachlor raised LPO and induced oxidative stress, are consistent with those of Faheem et al. [23], who discovered that LPO rose in grass carp that had been exposed to zinc and mercury. Similar increases in LPO that result in oxidative stress have been seen following butachlor therapy in the species Rana ridibunda [24]. High quantities of LPO were found in the plasma of C. gariepinus when it was exposed to sub-lethal doses of deltamethrin [25]. Carassius carassius treated with endosulfan exhibited a substantial LPO amplification in its blood, per Taweed et al. [25]. The lipid membrane is one of the targets of ROS generated by lipid peroxidation (LPO). In order to ascertain whether contaminants are generating oxidative stress in aquatic species, LPO measurement has proven to be a useful technique. As bioindicators of oxidative stress, fish exposed to various toxins may show increased activity of antioxidant enzymes and LPO [26].

The study's findings demonstrated that the plasma of S. melanotheron treated with butachlor had a notably higher CAT activity following a 15-day exposure period. The rise in CAT seen in this study may be a physiological reaction to the decrease in ROS production. Similar results have been observed when tilapia (O. niloticus) are exposed to pesticides in the laboratory [27]. Because they protect aquatic species from free radicals, which can result in oxidative stress, antioxidant defense enzymes like SOD and CAT are extremely beneficial. Based on the information at hand, SOD activity often decreased as CAT activity increased. It was also shown that CAT was the most sensitive antioxidant enzyme when compared to the other antioxidant enzymes. The increase in CAT activity may be a reaction to the increased oxidative stress caused by chemical exposures. Other fish species showed comparable increases in CAT activity after being exposed to pesticides [28]. The sensitivity of CAT activity to herbicide exposures was also corroborated by our previous research [29]. SOD activity reduction could indicate oxidative stress brought on by pesticides damaging the antioxidant systems [30].

The results of the present investigation demonstrate that butachlor induced oxidative stress, as evidenced by reductions in SOD and GSH levels and increases in CAT and LPO levels. Regulatory agencies may find it helpful to combine the use of oxidative stress biomarkers with a fish model when assessing the danger of contaminants in aquatic ecosystems. These traits can be used as indicators to assess a pesticide's toxicity in watery environments. Further studies are needed to evaluate the pesticide's residual effects in different fish body tissues in order to better understand the physiological implications of the methyl parathion status in natural populations.

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