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Full Research Article

Pulicaria odora as a Natural Antioxidant Remedy for Metribuzin-induced Toxicity

A
A. Benmahammed1
https://orcid.org/0009-0007-7461-1401
A
A. Basli1
https://orcid.org/0000-0001-7535-7029
F
F. Bouleknafet1
https://orcid.org/0000-0002-0676-930X
F
F.A. Al-Mekhlafi2,*
https://orcid.org/0000-0002-8190-7901
N
N. Abutaha2
https://orcid.org/0000-0002-1887-7789
H
H.M. Alharbi3
https://orcid.org/0009-0008-8185-9111
M
M.A. Wadaan2
https://orcid.org/0000-0002-8330-6990
S
S. Lambiase4
https://orcid.org/0000-0003-0846-1162
1Laboratory of Interaction Research, Biodiversity, Ecosystems and Biotechnology. Faculty of Sciences, University of August 20, 1955 Skikda, Skikda, Algeria.
2Department of Zoology, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia.
3Department of Biology, College of Science, Princess Nourah bint Abdulrahman University, P.O. Box 84428, Riyadh, 11671, Saudi Arabia.
4Department of Public Health, Experimental and Forensic Medicine, University of Pavia, Pavia, Italy.

ABSTRACT

Background: Pesticides like metribuzin are environmental pollutants that induce toxicity and oxidative stress in organisms.

Methods: This study evaluates the protective role of Pulicaria odora extract against metribuzin-induced toxicity in Wistar rats. The leaves of P. odora were extracted using methanol-water (70:30, v/v), yielding phenolic (2.77 mg GAE/g) and flavonoid (0.77 mg QE/g) contents. Its antioxidant capacity, assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays.

Result: Rats treated with metribuzin (13.2 mg/kg for 36 days) exhibited significant activity, including weight loss, altered hematological parameters and elevated hepatic (GPT, GOT) and renal (urea, creatinine) biomarkers. Co-administration of P. odora extract (34.2 mg/kg) significantly reversed these effects, restoring biomarkers and alleviating hepatic and renal toxicity by reducing transaminases, urea, creatinine, cholesterol and triglycerides. The extract’s antioxidant properties, attributed to its polyphenolic content, mitigated oxidative stress and stabilized cellular functions without adverse behavioral effects. This study highlights P. odora potential as a natural agent against herbicide-induced toxicity. Its bioactive compounds and mechanisms warrant further investigation, alongside exploring therapeutic applications in other Pulicaria species.

INTRODUCTION

The Asteraceae family comprises over 100 species, with Pulicaria ranking as the third-largest genus, widely distributed across Africa, Asia and Europe (Naqvi et al., 2020). Traditionally, Pulicaria species have been used to treat musculoskeletal pain, inflammation, menstrual cramps, digestive disorders, dysentery and diarrhea (Liu et al., 2010). Recent studies highlight their diverse bioactive properties, including the cytotoxic effects of P. undulata (Hegazy et al., 2021), anti-inflammatory potential of P. jaubertii (Hussain et al., 2023), antibacterial efficacy of P. odora (Touati et al., 2018) and antioxidant and hepatoprotective effects of P. crispa and P. incisa (El-Sabagh et al., 2021).
       
The extensive use of herbicides like metribuzin poses significant environmental and public health risks, particularly through their persistence, non-target exposure, and potential adverse effects on animals and humans (Kumar et al., 2024). Metribuzin, a selective systemic herbicide, is widely applied in agricultural and domestic settings, as well as for pest control in residential and commercial facilities and in veterinary medicine for managing ectoparasites (Gitsopoulos et al., 2024; Close and Banasiak, 2022). Studies have shown that metribuzin accumulates in the kidneys, liver, heart, adrenal glands and body fat of mammals and fish (Chiali et al., 2013; Pelić et al., 2020; Sousa et al., 2020), leading to oxidative stress and the formation of reactive oxygen species (ROS), which contribute to its toxicity and pose risks to human health (Gupta et al., 1999). Conversely, increased consumption of antioxidant-rich plant products has been shown to reduce environmental toxicant levels in the body by scavenging free radicals (Djordjević et al., 2024).
       
P. odora, locally known as “ouden halouf,” is an annual plant that can grow up to 100 cm tall and produces yellow flowers in humid conditions. It is traditionally valued for treating stomach ulcers, wounds, gastric disorders and abdominal pain (Djilali, 2020). Phytochemical studies on Pulicaria species have identified various bioactive compounds, including monoterpene glucosides (El-Ghaly, 2016), diterpenes, sesquiterpenes, flavonoids, polyphenols, triterpenes and steroids (De la Luz Cádiz Gurrea et al., 2019).
       
This study is driven by the limited information regarding the phytochemical composition and bioactive properties of P. odora. The aim is to investigate the plant’s biological efficacy in mitigating the harmful effects of agricultural toxins, particularly metribuzin and to evaluate its antioxidant and anti-inflammatory properties. Through in vivo studies on rats, this research seeks to demonstrate P. odora’s potential as a protective or supportive treatment to reduce tissue damage caused by exposure to hazardous substances.

MATERIALS AND METHODS

Pulicaria odora was collected in May 2022 at peak flowering from the Ain Kechra region in Skikda, northeastern Algeria, at an altitude of 282 meters (36°46′03.4"N, 6°26′52.8"E). The plant was identified and authenticated by Dr. Sahraoui, a botanist from the Faculty of Sciences, University of August 20, 1955, Skikda. After collection, the stems were separated and the leaves were thoroughly rinsed with tap water to remove surface impurities. The cleaned material was air-dried in a shaded, room-temperature environment for approximately 15 days. Once dried, the plant material was ground into a fine powder using a laboratory mill in preparation for extraction and subsequent analyses.
 
Extraction of plant
 
Fifty grams of powdered P. odora leaf material was immersed in 500 mL of a methanol-water solution (70:30, v/v) and continuously stirred using a magnetic stirrer for 24 hours at room temperature (20°C) to facilitate thorough extraction. Following the maceration process, the solution was filtered through Whatman No. 1 filter paper. The filtrate was then concentrated by vacuum evaporation at 40°C to remove the solvent, resulting in a concentrated extract. The dried extracts were carefully transferred into glass containers and stored at 4°C for subsequent analysis.
 
Determination of total phenolic content
 
To determine the total phenolic content, 500 µL of 10% Folin-Ciocalteu reagent was added to a solution containing 100 µL of the extract diluted in pure methanol. After a reaction time of 5 minutes, 400 µL of 7% sodium carbonate (Na‚ COƒ ) solution was added to the mixture. The solution was then incubated at room temperature for 30 minutes in a low-light environment to prevent photodegradation. Following incubation, the absorbance of the solution was measured at 375 nm using a Shimadzu UV-1601 UV/Vis spectrophotometer (Germany). The results were expressed as milligrams of gallic acid equivalents per gram of dry extract (mg GAE/g), quantitatively measuring the total phenolic content.
 
Quantification of flavonoid content
 
The flavonoid content of P. odora extract was quantified using the aluminum chloride colorimetric method, as Hegazy and Ibrahium (2012) described. In this procedure, 1 mL of the extract was mixed with 1 mL of a 2% aluminum chloride (AlClƒ ) solution. The mixture was then incubated for 10 minutes at room temperature to allow for the formation of a stable complex. After incubation, the absorbance was measured at 430 nm using a Shimadzu UV-1601 UV/Vis spectrophotometer (Germany). The total flavonoid content was expressed in micrograms of quercetin equivalents per gram of dry weight (µg QE/g DW).
 
DPPH radical scavenging activity
 
The free radical scavenging activity of P. odora leaf extract was assessed using the DPPH assay described by Jeong et al. (2004). This method mixed 100 µL of diluted leaf extract with 1 mL of a freshly prepared 0.05 mM DPPH solution in methanol. The mixture was incubated for 30 minutes at room temperature in a dark environment to prevent light-induced degradation of the DPPH radical. After incubation, the absorbance was measured at 517 nm using a spectrophotometer to quantify the decrease in DPPH concentration. The scavenging activity was expressed as percent inhibition of DPPH, calculated using the following equation:

 
Where,
A0= The absorbance of the control (DPPH solution without extract).
A1= The absorbance with the extract.
 
FRAP assay for ferric reducing antioxidant capacity
 
The ferric reducing antioxidant power (FRAP) of P. odora leaf extract was assessed following the protocol by Hemma et al., (2018). This assay evaluates the reduction power of the sample by measuring its ability to reduce Fe3+ to Fe2+. In this procedure, 400 µL of various concentrations of leaf extract or a standard solution were combined with 400 µL of phosphate buffer (0.2 M, pH 6.6) and 400 µL of potassium ferricyanide solution (1% aqueous K3Fe(CN)6). The mixture was incubated at 50°C for 20 minutes. After incubation, 400 µL of trichloroacetic acid solution (10% in water) was added, followed by centrifugation at 3000 rpm for 10 minutes to separate the phases. A portion of the supernatant (400 µL) was then diluted with 400 µL of distilled water and 80 µL of FeCl3  solution (0.1% in water) was added to the mixture. The absorbance of the final solution was measured at 700 nm to determine the reduction power. Ascorbic acid (AA) was used as a positive control in the assay.
 
Animal study
 
Thirty-six male albino Wistar rats, six weeks old and weighing between 150 and 180 grams, were obtained from the Pasteur Institute of Algeria. The study was conducted in the animal laboratory of the Science Department on August 20, 1955, at the University of Skikda, following institutional and national guidelines on laboratory animal care and the World Medical Association’s ethical guidelines (2016). The rats were housed in polyethylene cages lined with sterilized softwood shavings under controlled conditions of 25±2°C, 45-55% humidity and a 12-hour light/dark cycle. They had ad libitum access to food and water. Before the experiment, the rats were acclimatized to laboratory conditions for two weeks. All animal handling procedures adhered to the European Ethics Committee’s guidelines (06 EC/DCMB/FNSL/EU2021).
 
Animal treatment and experimental design
 
Following an acclimatization period, the rats were randomly allocated into four experimental groups, each comprising nine animals. Group GT, designated as the negative control, received mineral water orally. Group GP was administered an oral dose of P. odora extract at 34.2 mg/kg body weight daily. Group GM received metribuzin at a daily dose of 13.2 mg/kg body weight. Finally, Group GMP was treated with a combined daily regimen of metribuzin (13.2 mg/kg body weight) and P. odora extract (34.2 mg/kg body weight). After 36 days of treatment, the rats were euthanized and blood samples were collected for biochemical and hematological analysis. The serum was separated from the collected blood immediately post-euthanasia.
 
Hematological analysis
 
Hematological parameters were evaluated using whole blood samples and analyzed with a fully automated blood cell counter (Mindray BC 31). The parameters measured included red and white blood cell counts, hemoglobin (Hb) concentration, hematocrit, mean corpuscular values, lymphocyte counts and platelet levels.
 
Biochemical analysis
 
Serum samples were analyzed for clinical biochemistry using the Mindray BS-330E automated instrument. The biochemical parameters measured included glucose, urea, creatinine, triglycerides (TG), cholesterol, glutamic-oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT).
 
Statistical analysis
 
All data were analyzed in triplicate and the results are presented as mean ± standard deviation (SD). IC50 values for antioxidant assays were determined using linear regression analysis. Statistical analyses were performed using GraphPad Prism version 9.5.1. Differences between groups were assessed using ANOVA followed by Tukey’s test, with statistical significance set at P<0.05, P<0.01 and P<0.001.

RESULTS AND DISCUSSION

Extraction yields
 
The use of hydro-methanol as the solvent in maceration extraction enabled the calculation of the extraction rate. The total polyphenol yield obtained from P. odora leaves was 17.1% (w/w dry plant material). In comparison, methanolic crude extracts of P. crispa and P. undulata, wild species from Sudan, yielded 22.6% and 23%, respectively (Hegazy et al., 2021). This slight variation in yield percentage may be attributed to various factors, such as the extraction method, particle size, duration of extraction, solvent type and solvent volume (De la Luz Cádiz Gurrea et al., 2019).
 
Total flavonoids and total phenolic contents
 
To standardize the hydro-methanolic extract of P. odora leaves, the total phenolic content was measured using the folin-ciocalteu method and the UV-Vis colorimetric method was employed to quantify the total flavonoid content. Results showed a flavonoid concentration of 0.77 mg quercetin equivalent per gram of dry extract and a phenolic content of 2.77 mg gallic acid equivalent. Compared to these findings, Touati et al., (2018) reported lower total polyphenol and flavonoid levels in the methanolic extract of P. odora leaves, at 90±0.63 µg GAE/g DWE and 11.34±3.15 µg QE/g DWE, respectively. Additionally, the authors observed that P. odora leaves contain significantly higher levels of flavonoids and total phenolics than the roots. Prior studies also indicate that plants within the Pulicaria genus typically contain high levels of phenolics and flavonoids. For example, Senhaji et al., (2017) identified total phenolics and flavonoids in the ethyl acetate extract of P. murritanica at 72.88±0.21 mg GAE/g DWE and 38.95±0.75 mg QE/g DWE, respectively. Similarly, Malarz et al., (2023) reported that hydroalcoholic extracts of P. inuloides contain 43.81±2.36 mg GAE/g DWE of total polyphenols. Various factors may explain these differences, including genetic variation, plant maturity, storage duration and environmental conditions such as high temperatures, intense sunlight, drought and salinity, which are known to enhance secondary metabolite production, particularly polyphenols (Astill et al., 2001; Martínez et al., 2021; Turkmen et al., 2009).
 
Antioxidant activity
 
Two methods were employed to assess the antioxidant effects of P. odora cured extracts: DPPH radical scavenging activity and ferric ion (FeCl3) reducing power. The hydro-methanolic leaf extract of P. odora demonstrated dose-dependent DPPH radical scavenging, with inhibition rates of 44.02%, 48%, 48.6%, 50.27%, 54.27%, 59.24% and 63.4% at concentrations of 80, 100, 120, 140, 160, 180 and 200 µg/mL, respectively (Fig 1). A strong positive correlation was observed between extract concentration and radical inhibition, indicating increased antioxidant activity with higher concentrations. The IC50 for ascorbic acid was 103.33 µg/mL, while the IC50 for the P. odora extract was 123.69 µg/mL.

Fig 1: DPPH radicals scavenging activity of P. odora.


       
To further validate antioxidant activity, the reducing power test was conducted. Results showed a direct correlation between absorbance at 700 nm and extract concentration, where increased absorbance reflected the reduction of ferric ions by extract components. The inhibition percentages for the crude extract were 11.33%, 14.62%, 32.52%, 48.44%, 75.34% and 90.2%, compared to ascorbic acid’s 34.66%, 45.66%, 57.03%, 69.62%, 79.83% and 97.21% at 15.62, 31.25, 62.5, 125, 250 and 500 µg/mL, respectively (Fig 2). The IC50 values were 42.56 µg/mL for ascorbic acid and 192.24 µg/mL for the crude extract. Similar antioxidant responses have been reported for medicinal plant extracts, where phenolic-rich extracts demonstrated substantial DPPH and FRAP activities and improved antioxidant status in experimental rats (Azyu et al., 2025).

Fig 2: Antioxidant activity of methanol extract from leaves of P.odora using FRAP assay.


 
Sub-acute toxicity study
 
The current study investigates the potential detoxifying effects of P. odora on metribuzin-induced toxicity in Wistar albino rats. A comprehensive evaluation of physiological parameters, including body weight, hematological indices and biochemical markers, was conducted to elucidate the impact of both P. odora and metribuzin. Behavioral observations indicated that the oral administration of P. odora extract did not result in mortality or exhibit any toxicological symptoms, thereby underscoring the extract safety profile and potential therapeutic efficacy.
 
Growth parameters
 
Fig 3 displays rat body weight data across the experimental groups, highlighting differences among the four groups. At the beginning of the study, the initial body weight of the experimental rats averaged 170±10.2 g. A significant reduction in average body weight was observed in the GM group, which received metribuzin alone, compared to the control group (p<0.001). This weight loss is likely due to decreased food intake during the study period. These results align with findings by Samir and Asma (2018), who reported similar weight reduction effects following metribuzin exposure in rabbits over 6 to 18 days. In contrast, co-administration of P. odora leaf extract with metribuzin in the GPM group effectively prevented weight loss, showing no statistically significant difference from the control group. The phenolic compounds in P. odora extract are likely instrumental in counteracting rat metribuzin-induced oxidative stress (Zeng et al., 2021).

Fig 3: Average body weight of male albino wistar rats (n = 9) over a 36-day treatment period, across four groups.


 
Hematological and immune response to metribuzin and P. odora extract in rats
 
Fig 4 illustrates the hematological responses to metribuzin and P. odora extract treatments, highlighting notable changes in these parameters. In the GM group, which received metribuzin alone, significant elevations were observed in red blood cell (RBC) counts and hematocrit levels (P<0.01), along with a more pronounced increase in white blood cell (WBC) counts, lymphocytes, hemoglobin (Hb), mean corpuscular volume (MCV) and platelets (PLTs) (P<0.001) compared to the control. In contrast, co-treatment with P. odora extract in the GMP group led to a marked reduction in platelets, Hb, WBC, RBC and MCV levels relative to the GM group, while lymphocyte and hematocrit levels remained unaffected. Rats administered only P. odora extract (GP) showed a significant increase in RBC counts (P<0.01) and a minor but significant decrease in MCV (P<0.05) when compared to the control group, with other hematological parameters showing no significant differences.

Fig 4: The hematological profile of rats intoxicated with metribuzin and after treatment with methanolic extract of P. odora.


       
The observed elevation in RBC counts, MCV, hematocrit and Hb levels in the GM group may reflect a physiological stress response to metribuzin exposure, potentially due to compensatory mechanisms aimed at countering herbicide-induced oxidative stress. This response likely involves increased oxygen transport to cells, evidenced by the rise in RBC counts, possibly through enhanced release or synthesis of erythrocytes from hematopoietic tissues. Similar responses have been reported by Lutnicka et al., (2019) in C. carpio exposed to linuron. Furthermore, (Kadeche et al., 2016) and Samir et al. (2020) observed declines in RBC, Hb and hematocrit levels and reduced erythrocyte counts in metribuzin-treated rats, particularly with prolonged or high-dose exposure, attributing these effects to oxidative damage and hematotoxicity.
       
Metribuzin-treated rats in the present study exhibited significantly increased lymphocyte, platelet and WBC counts (P<0.001), likely indicating an immune response to herbicide-induced toxicity. Elevated WBC and lymphocyte levels may suggest an immune mobilization against tissue damage, consistent with findings from Samir et al. (2020), who reported similar leukocyte increases in response to metribuzin’s cytotoxic effects. The rise in platelet count may stem from metribuzin-related tissue damage, as platelets contribute to inflammatory responses by releasing pro-inflammatory mediators to recruit immune cells (Chen et al., 2020). Additionally, metribuzin’s oxidative stress-inducing properties (Almeida et al., 2019) may further explain the increase in platelet levels as a compensatory response to RBC suppression.
       
Treatment with P. odora extract alone induced minor changes in RBC counts and MCV relative to the GP and control groups, similar to findings by Mansouri et al. (2015) in studies with Foeniculum vulgare extracts. This effect may result from erythropoietin stimulation by the liver and kidneys in response to P. odora. The phenolic constituents of P. odora likely contribute antioxidant effects, stabilizing cell membranes and protecting against free radical damage (Koren et al., 2010). Although changes in platelet, MCV and RBC levels were not statistically significant, they suggest a potential protective effect of P. odora. Moreover, Hb and WBC levels in the GM group showed a significant reduction (P<0.001) relative to controls, while these levels remained elevated in the GP group, further supporting P. odora’s stabilizing role.
       
Collectively, these results imply that oxidative stress plays a central role in metribuzin toxicity, with P. odora extract, rich in polyphenols and flavonoids, offering significant antioxidant and free radical-scavenging properties that may counteract metribuzin-induced damage. This aligns with findings by Kadeche et al. (2017) on vanillin’s protective effect against metribuzin-induced hematological alterations, underscoring P. odora’s potential as a therapeutic agent against herbicide toxicity.
 
Biochemical effects of metribuzin and P. odora extract in rats
 
In this study, exposure to metribuzin (GM group) significantly increased (P<0.001) several biochemical markers, including blood glucose, total cholesterol, triglycerides, glutamate pyruvate transaminase (GPT), glutamate oxaloacetate transaminase (GOT), urea and creatinine, relative to the control group (Fig 5). These elevations suggest a toxic impact on liver and kidney function, as evidenced by raised transaminase levels and markers of renal function. Interestingly, the co-treatment group (GMP), which received P. odora extract alongside metribuzin, exhibited significant reductions in GPT, GOT and creatinine (P<0.001), urea (P<0.01), as well as total cholesterol and triglycerides (P< 0.05), compared to the GM group. This indicates that P. odora may confer a protective effect, mitigating metribuzin-induced hepatic and renal toxicity. However, co-treatment did not significantly affect blood glucose levels, suggesting limited influence on glucose metabolism. Additionally, the group receiving only P. odora extract (GP group) showed a mild, statistically insignificant increase in GPT, GOT and triglycerides compared to the control group, with no significant changes in other biochemical parameters, indicating that P. odora alone does not markedly disrupt biochemical homeostasis. Changes in hematological indices and serum biomarkers of hepatic and renal function are commonly used to evaluate the systemic effects and safety of bioactive plant extracts in Wistar rats (Al-Ghamdi et al., 2026). Collectively, these findings support the potential protective role of P. odora against metribuzin-induced toxicity.

Fig 5: Intoxicated rats’ biochemical parameters with metribuzin and after treatment with a methanolic extract of P. odora.


       
This study also noted a marked increase in blood glucose levels in rats exposed to metribuzin (13.2 mg/mL over 36 days), consistent with previous findings indicating that chronic low-level metribuzin exposure elevates plasma glucose, possibly due to reduced glucose uptake and impaired tissue functionality (Chiali et al., 2013). Similar glucose elevations were reported in Cyprinus carpio after atrazine exposure (Blahova et al., 2014), suggesting that pesticides may disrupt carbohydrate metabolism. Potential mechanisms include increased glycogenolysis, elevated adrenocorticotropic and glucagon hormones, or insulin dysfunction (Mehra et al., 2014). Conversely, studies on long-term, low-dose metribuzin exposure in rats (1/20 to 1/5 of LD50, twice weekly) reported decreased glucose levels, possibly due to depleted carbohydrate reserves or inhibition of glucose release from hepatic tissue (Maksymiv et al., 2015; Samir et al., 2020). Metribuzin exposure also increased lipid parameters, particularly triglycerides and total cholesterol, potentially due to enhanced cholesterol synthesis in the liver or compromised liver cell membrane integrity. Elevated triglycerides may result from decreased lipase activity and increased mobilization of adipose stores (Chiali et al., 2013; El-Demerdash and Nasr 2014). Some studies, however, have reported reduced plasma cholesterol under metribuzin exposure, potentially due to diminished cholesterol synthesis or tissue accumulation.
       
Increased urea and creatinine levels in metribuzin-treated rats indicate renal impairment and reduced glomerular filtration rate, consistent with findings in albino rats exposed to metribuzin (Khozimy et al., 2017). Elevated liver biomarkers GPT and GOT indicate hepatotoxicity, aligning with Samir and Asma (2018), who observed increased transaminase activity in metribuzin-exposed rabbits and fetuses. The herbicide’s residual accumulation in the liver, as confirmed by HPLC analysis, underscores hepatic vulnerability to metribuzin toxicity. Similarly, Sena et al. (2021) found heightened GPT and GOT in Xenopus laevis exposed to atrazine. Rats treated with P. odora extract alone (GP group) exhibited slight, non-significant increases in triglycerides, GOT and GPT, potentially indicating activation of lipid metabolism-related enzymes. However, no studies to date have specifically examined the biochemical effects of P. odora, underscoring the need for further investigation.
       
Importantly, co-treatment with P. odora extract and metribuzin mitigated the metabolic disturbances induced by metribuzin, restoring levels of GPT, GOT, urea, creatinine, cholesterol and triglycerides to near normal. This protective effect is likely due to the natural antioxidants, particularly polyphenols, in P. odora, which may counteract metribuzin-induced biochemical imbalances. These findings align with previous studies on P. crispa extract, which lowered cholesterol, urea and creatinine in diabetic rats (Daradka et al., 2021) and support evidence of the benefits of Pulicaria genus on liver and kidney health (Bakr et al., 2021; Alharthi et al., 2023). Similar antioxidant compounds, such as vanillin and quercetin, have demonstrated efficacy in ameliorating metribuzin-induced biochemical changes (Abarikwu 2014; Kadeche et al., 2017), suggesting that P. odora’s quercetin content may contribute to its protective properties.

CONCLUSION

The findings of this study demonstrate that P. odora extract possesses significant protective effects against metribuzin-induced toxicity in Wistar rats. The extract’s antioxidant properties, attributed to its phenolic and flavonoid content, played a key role in reducing oxidative stress and mitigating the physiological and biochemical alterations induced by metribuzin. Co-administration of P. odora extract resulted in a marked improvement in hematological and biochemical parameters, including liver and kidney function markers. The absence of adverse effects on behavioural assessments further supports the safety and potential therapeutic value of P. odora. These results suggest that P. odora may serve as a promising natural agent for combating herbicide-induced toxicity. Further research is warranted to explore its bioactive compounds, mechanisms of action and the therapeutic potential of other species within the Pulicaria genus.

ACKNOWLEDGEMENT

The authors would like to thank the Algerian Ministry of Higher Education and Scientific Research for providing the necessary facilities. They also extend their heartfelt gratitude to the Princess Nourah bint Abdulrahman University Researchers Supporting Project number (PNURSP2026R931), Princess Nourah bint Abdulrahman University, Riyadh,Saudi Arabia.
 
Disclaimers
 
The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
 
Authors’ contributions
 
Conceptualization: A. Benmahammed and ABA, Data curation: ABA and F. Bouleknafet, Formal analysis: F.A. Al-Mekhlafi and N. Abutaha, Funding acquisition: M.A. Wadaan and H.M. Alharbi Investigation: M.A. Wadaan, Methodology: A. Benmahammed and ABA, Project administration: F. Bouleknafet and S. Lambiase Resources: F.A. Al-Mekhlafi. Supervision: ABA. Writing-original draft: ABA, F.A. Al-Mekhlafi and F. Bouleknafet Writing-review and editing: N. Abutaha, F.A. Al-Mekhlafi and H.M. Alharbi.
 
Funding
 
This project was funded by Princess Nourah bint Abdulrahman University Researchers Supporting Project number (PNURSP2026R931), Princess Nourah bint Abdulrahman University, Riyadh,Saudi Arabia.
 
Data availability statement
 
All the data is available within the manuscript.
 
Informed consent
 
All animal procedures for experiments were approved by the Committee of Experimental Animal care and handling techniques were approved by the University of Animal Care Committee.

CONFLICT OF INTEREST

The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

REFERENCES


  1. Abarikwu, S.O. (2014). Protective effect of quercetin on atrazine- induced oxidative stress in the liver, kidney, brain and heart of adult Wistar rats. Toxicology International. 21: 148-155.

  2. Al-Ghamdi, N.A., Omer, S.A., Alterary, S.S., El-Tohamy, M.F., Virk, P., El-Nagar, D.M. and Mohammed, O.B. (2026). Hepatorenal toxicity with hematological and biochemical changes in wistar rats fed the ethanolic extract of the pulp of Citrullus colocynthis from two regions in Saudi Arabia. Indian Journal of Animal Research. 60(6): 1030-1037. doi: 10.18805/IJAR.BF-2006.

  3. Alharthi, A.S., Alruwaili, N.W., Al-Baadani, H.H., Al-Garadi, M.A., Shamlan, G., Alhidary, I. A. (2023). Investigating the effect of Pulicaria jaubertii as a natural feed additive on the growth performance, blood biochemistry, immunological response and cecal microbiota of broiler chickens. Animals. 13: 1116.

  4. Almeida, A.C., Gomes, T., Langford, K., Thomas, K.V. and Tollefsen, K.E. (2019). Oxidative stress potential of the herbicides bifenox and metribuzin in the microalgae Chlamydomonas reinhardtii. Aquatic Toxicology. 210: 117-128.

  5. Astill, C., Birch, M.R., Dacombe, C., Humphrey, P.G. and Martin, P.T. (2001). Factors affecting the caffeine and polyphenol contents of black and green tea infusions. Journal of Agricultural and Food Chemistry. 49: 5340-5347.

  6. Azyu, R.Z., Ali Ayub, M., Gali Mohanarao, J., Behera, P., Das, H., Mayengbam, P. and Devi, I.L. (2025). Effect of ethanolic extract of Scurrula parasitica L. on in vivo antioxidant status of diabetic rats. Indian Journal of Animal Research. doi: 10.18805/IJAR.B-5462.

  7. Bakr, R.O., Shahat, E.A., Elissawy, A.E., Fayez, A.M. and Eldahshan, O.A. (2021). Evaluation of the hepatoprotective activity of Pulicaria incisa subspecies candolleana and in silico screening of its isolated phenolics. Journal of Ethnopharmacology. 271: 113767.

  8. Blahova, J., Modra, H., Sevcikova, M., Marsalek, P., Zelnickova, L., Skoric, M. and Svobodova, Z. (2014). Evaluation of biochemical, haematological and histopathological responses and recovery ability of common carp (Cyprinus carpio L.) after acute exposure to atrazine herbicide. Biomedical Research International. 2014: 980948.

  9. Chen, Y., Zhong, H., Zhao, Y., Luo, X., Gao, W. (2020). Role of platelet biomarkers in inflammatory response. Biomarker Research. 8: 1-7.

  10. Chiali, F., Merzouk, H., Merzouk, S., Medjdoub, A. and Narce, M. (2013). Chronic low-level metribuzin exposure induces metabolic alterations in rats. Pesticide Biochemistry and Physiology. 106: 38-44.

  11. Close, M. and Banasiak, L. (2022). National Survey of Pesticides in Groundwater 2022.

  12. Daradka, H.M., Aldhilan, A.M., Eskandrani, A.A., Bataineh, Y. and Alzoubi, K.H. (2021). The effect of Pulicaria crispa ethanolic extract on haematological and biochemical parameters in alloxan-induced diabetic rats. Advances in Traditional Medicine. 21: 65-72.

  13. De la Luz Cádiz-Gurrea, M., Zengin, G., Kayacik, O., Lobine, D., Mahomoodally, M.F., Leyva-Jiménez, F.J. and Segura- Carretero, A. (2019). Innovative perspectives on Pulicaria dysenterica extracts: Phyto-pharmaceutical properties, chemical characterization and multivariate analysis. Journal of the Science of Food and Agriculture. 99: 6001-6010.

  14. Djilali, A.B., Benseddik, A., Bouacem, K., Allaf, K. and Nabiev, M. (2020). Functional properties of Pulicaria odora L. leaves pre-coated in gel-based Ziziphus jujuba Mill. peel powder. Nature Products in Agro-Industrial Processing. 6: 1-4.

  15. Djordjević, M., Djordjević, M., Starowicz, M. and Krupa-Kozak, U. (2024). Plant-based antioxidants in gluten-free bread production: Sources, technological and sensory aspects, enhancing strategies and constraints. Antioxidants. 13: 142.

  16. El-Demerdash, F.M and Nasr, H.M. (2014). Antioxidant effect of selenium on lipid peroxidation, hyperlipidemia and biochemical parameters in rats exposed to diazinon. Journal of Trace Elements in Medicine and Biology. 28: 89-93.

  17. El-Ghaly, E.S.M., Shaheen, U., Ragab, E., El-Hila, A.A. and Abd- Allah, M.R. (2016). Bioactive constituents of Pulicaria jaubertii: A promising antihypertensive activity. Pharmacognosy Journal. 8: 81-86.

  18. El-Sabagh, O.A., El-Toumy, S.A., Mounir, R., Farag, M.A. and Mahrous, E.A. (2021). Metabolite profiles of Pulicaria crispa and P. incisa in relation to their in vitro/in vivo antioxidant activity and hepatoprotective effect: A comparative mass spectrometry-based metabolomics. Journal of Pharmaceutical and Biomedical Analysis. 194: 113804.

  19. Gitsopoulos, T., Georgoulas, I., Vazanelli, E. and Botsoglu, D. (2024). Selectivity of the premixtures flufenacet, diflufenican and flufenacet, diflufenican, metribuzin on bread wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) and efficacy on ALS/ACCase-resistant populations of lolium rigidum gaudin. Agronomy. 14: 949.

  20. Gupta, H.V., Sorooshian, S. and Yapo, P.O. (1999). Status of automatic calibration for hydrologic models: Comparison with multilevel expert calibration. Journal of Hydrologic Engineering. 4: 135-143.

  21. Hegazy, A.E. and Ibrahium, M.I. (2012). Antioxidant activities of orange peel extracts. World Applied Sciences Journal. 18: 684-688.

  22. Hegazy, M.E.F., Dawood, M., Mahmoud, N., Elbadawi, M., Sugimoto, Y., Klauck, S.M., Mohamed, N. and Efferth, T. (2021). 2α- Hydroxyalantolactone from Pulicaria undulata: Activity against multidrug-resistant tumor cells and modes of action. Phytomedicine. 81: 153409.

  23. Hemma, R., Belhadj, S., Ouahchia, C. and Saidi, F. (2018). Antioxidant activity of Pistacia lentiscus methanolic extracts. Revista de Agrobiología. 8: 845-852.

  24. Hussain, S., Sagar, T., Kaur, S., Nipuntha, K., Kapoor, N. and Mahajan, R. (2023). Omics Approaches to Study the Biosynthesis of Bioactive Compounds in Medicinal and Aromatic Plants. In Biosynthesis of Bioactive Compounds in Medicinal and Aromatic Plants: Manipulation by Conventional and Biotechnological Approaches. Springer. (pp. 41-64).

  25. Jeong, S.M., Kim, S.Y., Kim, D.R., Jo, S.C., Nam, K., Ahn, D. and Lee, S.C. (2004). Effect of heat treatment on the antioxidant activity of extracts from citrus peels. Journal of Agricultural and Food Chemistry. 52: 3389-3393.

  26. Kadeche, L., Bourogaa, E., Boumendjel, A., Djeffal, A., El Feki, A. and Messarah, M. (2016). Quercetin attenuates metribuzin- induced biochemical and hematological toxicity in adult rats. International Journal of Pharmaceutical Sciences Review and Research. 40: 38-46.

  27. Kadeche, L., Bourogaa, E., Saoudi, M., Boumendjel, A., Djeffal, A., El Feki, A. and Messarah, M. (2017). Ameliorative effects of vanillin against metribuzin-induced oxidative stress and toxicity in rats. International Journal of Pharmaceutical and Pharmaceutical Sciences. 9: 56-62.

  28. Khozimy, A.M., Ramadan, M.F., Shaldam, H.A. and Aly, T.A. (2017). Effect of storage temperature on degradation of two herbicides and their sub-acute toxicity on albino rats. Journal of Applied Plant Protection, Suez Canal University. 6: 1.

  29. Koren, E., Kohen, R. and Ginsburg, I. (2010). Polyphenols enhance total oxidant-scavenging capacities of human blood by binding to red blood cells. Experimental Biology and Medicine. 235: 689-699.

  30. Kumar, J.S., Monica, S.S., Vinothkumar, B., Suganthi, A. and Paramasivam, M. (2024). Impact of pesticide exposure on environment and biodiversity: A review. Agricultural Reviews. 45(1): 01-12. doi: 10.18805/ag.R-2325.

  31. Liu, L.L., Yang, J.L. and Shi, Y.P. (2010). Phytochemicals and biological activities of Pulicaria species. Chemistry Biodiversity. 7: 327-349.

  32. Lutnicka, H., Bojarski, B., Witeska, M., Tombarkiewicz, B. and Formicki, G. (2019). Exposure to herbicide linuron results in alterations in hematological profile and stress biomarkers of common carp (Cyprinus carpio). Ecotoxicology. 28: 69-75.

  33. Maksymiv, I.V., Husak, V.V., Mosiichuk, N.M., Matviishyn, T.M., Sluchyk, I.Y., Storey, J.M., Storey, K.B. and Lushchak, V.I. (2015). Hepatotoxicity of herbicide Sencor in goldfish may result from induction of mild oxidative stress. Pesticide Biochemistry and Physiology. 122: 67-75.

  34. Malarz, J., Michalska, K., Galanty, A., Kiss, A.K. and Stojakowska, A. (2023). Constituents of Pulicaria inuloides and cytotoxic activities of two methoxylated flavonols. Molecules. 28: 480.

  35. Mansouri, E., Kooti, W., Bazvand, M., Boroon, M.G., Amirzargar, A., Afrisham, R., Afzalzadeh, M.R., Ashtary-Larky, D. and Jalali, N. (2015). The effect of hydro-alcoholic extract of Foeniculum vulgare Mill on leukocytes and hematological tests in male rats. Jundishapur Journal of Natural Pharmaceutical Products. 10(1).

  36. Martínez, C., Valenzuela, J.L. and Jamilena, M. (2021). Genetic and pre-and postharvest factors influencing the content of antioxidants in cucurbit crops. Antioxidants. 10(6): 894.

  37. Mehra, B., Sharma, P., Kaushik, U. and Joshi, S. (2014). Effect of fytolan on haematology and serum parameters of male albino rats. World Journal of Pharmaceutical Sciences. 3: 817-829.

  38. Naqvi, S.A.R., Shah, S.M.A., Kanwal, L., Saeed, M., Atta-ul-Haq, Nisar, J., Nisar, Z. and Akram, M. (2020). Antimicrobial and antihypercholesterolemic activities of Pulicaria gnaphalodes. Dose-Response. 18(1): 1559325820904858.

  39. Pelić, M., Kartalović, B., Živkov-Baloš, M., Mirolić, M., Đorđević, M., Teodorović, V., Ćirković, M., Ljubojević-Pelić, D. (2020). Health risks associated with residual pesticide levels in fish reared in purified wastewater from slaughterhouse. Journal of the Hellenic Veterinary Medical Society. 71(1): 1991-1996.

  40. Samir, D and Asma, S. (2018). Triazinone herbicide metribuzin induced acute liver injury: A study of animal model. Journal of Acute Diseases. 7(4): 152-157.

  41. Samir, D., Selma, R.M.O. and Asma, S. (2020). The effect of herbicide metribuzin on environment and human: A systematic review. Pharmaceutical Biosciences Journal. pp 10-15.

  42. Sena, L., Asouzu Johnson, J., Nkomozepi, P. and Mbajiorgu, E.F. (2021). Atrazine-induced hepato-renal toxicity in adult male Xenopus laevis frogs. Applied Science. 11(24): 11776.

  43. Senhaji, B., Chebli, B., Mayad, E., Hamdouch, A., Heimeur, N., Chahid, A. and Ferji, Z. (2017). Phytochemical screening, quantitative analysis and antioxidant activity of Asteriscus imbricatus and Pulicaria mauritanica organic extracts. International Food Research Journal. 24(6): 2482-2489.

  44. Sousa, S., Maia, M., Correira-Sá, L., Fernandes, V., Delerue-Matos, C., Calhau, C. and Domingues, V. (2020). Chemistry and toxicology behind insecticides and herbicides. In Controlled Release of Pesticides for Sustainable Agriculture. pp 59-109.

  45. Touati, N., Saidani, K., Boudries, H., Hammiche, H., Ouazene, N. and Bedjou, F. (2018). Antibacterial activity of phenolic compounds of Pulicaria odora, wild plant in northern Algeria. International Food Research Journal. 25(5): 2021-2030.

  46. Turkmen, N., Sari, F. and Velioglu, Y.S. (2009). Factors affecting polyphenol content and composition of fresh and processed tea leaves. Akademik Gida. 7(6): 29-40.

  47. Zeng, X., Du, Z., Ding, X. and Jiang, W. (2021). Protective effects of dietary flavonoids against pesticide-induced toxicity: A review. Trends in Food Science and Technology. 109: 271-279.
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    Full Research Article

    Pulicaria odora as a Natural Antioxidant Remedy for Metribuzin-induced Toxicity

    A
    A. Benmahammed1
    https://orcid.org/0009-0007-7461-1401
    A
    A. Basli1
    https://orcid.org/0000-0001-7535-7029
    F
    F. Bouleknafet1
    https://orcid.org/0000-0002-0676-930X
    F
    F.A. Al-Mekhlafi2,*
    https://orcid.org/0000-0002-8190-7901
    N
    N. Abutaha2
    https://orcid.org/0000-0002-1887-7789
    H
    H.M. Alharbi3
    https://orcid.org/0009-0008-8185-9111
    M
    M.A. Wadaan2
    https://orcid.org/0000-0002-8330-6990
    S
    S. Lambiase4
    https://orcid.org/0000-0003-0846-1162
    1Laboratory of Interaction Research, Biodiversity, Ecosystems and Biotechnology. Faculty of Sciences, University of August 20, 1955 Skikda, Skikda, Algeria.
    2Department of Zoology, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia.
    3Department of Biology, College of Science, Princess Nourah bint Abdulrahman University, P.O. Box 84428, Riyadh, 11671, Saudi Arabia.
    4Department of Public Health, Experimental and Forensic Medicine, University of Pavia, Pavia, Italy.

    ABSTRACT

    Background: Pesticides like metribuzin are environmental pollutants that induce toxicity and oxidative stress in organisms.

    Methods: This study evaluates the protective role of Pulicaria odora extract against metribuzin-induced toxicity in Wistar rats. The leaves of P. odora were extracted using methanol-water (70:30, v/v), yielding phenolic (2.77 mg GAE/g) and flavonoid (0.77 mg QE/g) contents. Its antioxidant capacity, assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric reducing antioxidant power (FRAP) assays.

    Result: Rats treated with metribuzin (13.2 mg/kg for 36 days) exhibited significant activity, including weight loss, altered hematological parameters and elevated hepatic (GPT, GOT) and renal (urea, creatinine) biomarkers. Co-administration of P. odora extract (34.2 mg/kg) significantly reversed these effects, restoring biomarkers and alleviating hepatic and renal toxicity by reducing transaminases, urea, creatinine, cholesterol and triglycerides. The extract’s antioxidant properties, attributed to its polyphenolic content, mitigated oxidative stress and stabilized cellular functions without adverse behavioral effects. This study highlights P. odora potential as a natural agent against herbicide-induced toxicity. Its bioactive compounds and mechanisms warrant further investigation, alongside exploring therapeutic applications in other Pulicaria species.

    INTRODUCTION

    The Asteraceae family comprises over 100 species, with Pulicaria ranking as the third-largest genus, widely distributed across Africa, Asia and Europe (Naqvi et al., 2020). Traditionally, Pulicaria species have been used to treat musculoskeletal pain, inflammation, menstrual cramps, digestive disorders, dysentery and diarrhea (Liu et al., 2010). Recent studies highlight their diverse bioactive properties, including the cytotoxic effects of P. undulata (Hegazy et al., 2021), anti-inflammatory potential of P. jaubertii (Hussain et al., 2023), antibacterial efficacy of P. odora (Touati et al., 2018) and antioxidant and hepatoprotective effects of P. crispa and P. incisa (El-Sabagh et al., 2021).
           
    The extensive use of herbicides like metribuzin poses significant environmental and public health risks, particularly through their persistence, non-target exposure, and potential adverse effects on animals and humans (Kumar et al., 2024). Metribuzin, a selective systemic herbicide, is widely applied in agricultural and domestic settings, as well as for pest control in residential and commercial facilities and in veterinary medicine for managing ectoparasites (Gitsopoulos et al., 2024; Close and Banasiak, 2022). Studies have shown that metribuzin accumulates in the kidneys, liver, heart, adrenal glands and body fat of mammals and fish (Chiali et al., 2013; Pelić et al., 2020; Sousa et al., 2020), leading to oxidative stress and the formation of reactive oxygen species (ROS), which contribute to its toxicity and pose risks to human health (Gupta et al., 1999). Conversely, increased consumption of antioxidant-rich plant products has been shown to reduce environmental toxicant levels in the body by scavenging free radicals (Djordjević et al., 2024).
           
    P. odora    , locally known as “ouden halouf,” is an annual plant that can grow up to 100 cm tall and produces yellow flowers in humid conditions. It is traditionally valued for treating stomach ulcers, wounds, gastric disorders and abdominal pain (Djilali, 2020). Phytochemical studies on Pulicaria species have identified various bioactive compounds, including monoterpene glucosides (El-Ghaly, 2016), diterpenes, sesquiterpenes, flavonoids, polyphenols, triterpenes and steroids (De la Luz Cádiz Gurrea et al., 2019).
           
    This study is driven by the limited information regarding the phytochemical composition and bioactive properties of P. odora. The aim is to investigate the plant’s biological efficacy in mitigating the harmful effects of agricultural toxins, particularly metribuzin and to evaluate its antioxidant and anti-inflammatory properties. Through in vivo studies on rats, this research seeks to demonstrate P. odora’s potential as a protective or supportive treatment to reduce tissue damage caused by exposure to hazardous substances.

    MATERIALS AND METHODS

    Pulicaria odora was collected in May 2022 at peak flowering from the Ain Kechra region in Skikda, northeastern Algeria, at an altitude of 282 meters (36°46′03.4"N, 6°26′52.8"E). The plant was identified and authenticated by Dr. Sahraoui, a botanist from the Faculty of Sciences, University of August 20, 1955, Skikda. After collection, the stems were separated and the leaves were thoroughly rinsed with tap water to remove surface impurities. The cleaned material was air-dried in a shaded, room-temperature environment for approximately 15 days. Once dried, the plant material was ground into a fine powder using a laboratory mill in preparation for extraction and subsequent analyses.
     
    Extraction of plant
     
    Fifty grams of powdered P. odora leaf material was immersed in 500 mL of a methanol-water solution (70:30, v/v) and continuously stirred using a magnetic stirrer for 24 hours at room temperature (20°C) to facilitate thorough extraction. Following the maceration process, the solution was filtered through Whatman No. 1 filter paper. The filtrate was then concentrated by vacuum evaporation at 40°C to remove the solvent, resulting in a concentrated extract. The dried extracts were carefully transferred into glass containers and stored at 4°C for subsequent analysis.
     
    Determination of total phenolic content
     
    To determine the total phenolic content, 500 µL of 10% Folin-Ciocalteu reagent was added to a solution containing 100 µL of the extract diluted in pure methanol. After a reaction time of 5 minutes, 400 µL of 7% sodium carbonate (Na‚ COƒ ) solution was added to the mixture. The solution was then incubated at room temperature for 30 minutes in a low-light environment to prevent photodegradation. Following incubation, the absorbance of the solution was measured at 375 nm using a Shimadzu UV-1601 UV/Vis spectrophotometer (Germany). The results were expressed as milligrams of gallic acid equivalents per gram of dry extract (mg GAE/g), quantitatively measuring the total phenolic content.
     
    Quantification of flavonoid content
     
    The flavonoid content of P. odora extract was quantified using the aluminum chloride colorimetric method, as Hegazy and Ibrahium (2012) described. In this procedure, 1 mL of the extract was mixed with 1 mL of a 2% aluminum chloride (AlClƒ ) solution. The mixture was then incubated for 10 minutes at room temperature to allow for the formation of a stable complex. After incubation, the absorbance was measured at 430 nm using a Shimadzu UV-1601 UV/Vis spectrophotometer (Germany). The total flavonoid content was expressed in micrograms of quercetin equivalents per gram of dry weight (µg QE/g DW).
     
    DPPH radical scavenging activity
     
    The free radical scavenging activity of P. odora leaf extract was assessed using the DPPH assay described by Jeong et al. (2004). This method mixed 100 µL of diluted leaf extract with 1 mL of a freshly prepared 0.05 mM DPPH solution in methanol. The mixture was incubated for 30 minutes at room temperature in a dark environment to prevent light-induced degradation of the DPPH radical. After incubation, the absorbance was measured at 517 nm using a spectrophotometer to quantify the decrease in DPPH concentration. The scavenging activity was expressed as percent inhibition of DPPH, calculated using the following equation:

     
    Where,
    A0= The absorbance of the control (DPPH solution without extract).
    A1= The absorbance with the extract.
     
    FRAP assay for ferric reducing antioxidant capacity
     
    The ferric reducing antioxidant power (FRAP) of P. odora leaf extract was assessed following the protocol by Hemma et al., (2018). This assay evaluates the reduction power of the sample by measuring its ability to reduce Fe3+ to Fe2+. In this procedure, 400 µL of various concentrations of leaf extract or a standard solution were combined with 400 µL of phosphate buffer (0.2 M, pH 6.6) and 400 µL of potassium ferricyanide solution (1% aqueous K3Fe(CN)6). The mixture was incubated at 50°C for 20 minutes. After incubation, 400 µL of trichloroacetic acid solution (10% in water) was added, followed by centrifugation at 3000 rpm for 10 minutes to separate the phases. A portion of the supernatant (400 µL) was then diluted with 400 µL of distilled water and 80 µL of FeCl3  solution (0.1% in water) was added to the mixture. The absorbance of the final solution was measured at 700 nm to determine the reduction power. Ascorbic acid (AA) was used as a positive control in the assay.
     
    Animal study
     
    Thirty-six male albino Wistar rats, six weeks old and weighing between 150 and 180 grams, were obtained from the Pasteur Institute of Algeria. The study was conducted in the animal laboratory of the Science Department on August 20, 1955, at the University of Skikda, following institutional and national guidelines on laboratory animal care and the World Medical Association’s ethical guidelines (2016). The rats were housed in polyethylene cages lined with sterilized softwood shavings under controlled conditions of 25±2°C, 45-55% humidity and a 12-hour light/dark cycle. They had ad libitum access to food and water. Before the experiment, the rats were acclimatized to laboratory conditions for two weeks. All animal handling procedures adhered to the European Ethics Committee’s guidelines (06 EC/DCMB/FNSL/EU2021).
     
    Animal treatment and experimental design
     
    Following an acclimatization period, the rats were randomly allocated into four experimental groups, each comprising nine animals. Group GT, designated as the negative control, received mineral water orally. Group GP was administered an oral dose of P. odora extract at 34.2 mg/kg body weight daily. Group GM received metribuzin at a daily dose of 13.2 mg/kg body weight. Finally, Group GMP was treated with a combined daily regimen of metribuzin (13.2 mg/kg body weight) and P. odora extract (34.2 mg/kg body weight). After 36 days of treatment, the rats were euthanized and blood samples were collected for biochemical and hematological analysis. The serum was separated from the collected blood immediately post-euthanasia.
     
    Hematological analysis
     
    Hematological parameters were evaluated using whole blood samples and analyzed with a fully automated blood cell counter (Mindray BC 31). The parameters measured included red and white blood cell counts, hemoglobin (Hb) concentration, hematocrit, mean corpuscular values, lymphocyte counts and platelet levels.
     
    Biochemical analysis
     
    Serum samples were analyzed for clinical biochemistry using the Mindray BS-330E automated instrument. The biochemical parameters measured included glucose, urea, creatinine, triglycerides (TG), cholesterol, glutamic-oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT).
     
    Statistical analysis
     
    All data were analyzed in triplicate and the results are presented as mean ± standard deviation (SD). IC50 values for antioxidant assays were determined using linear regression analysis. Statistical analyses were performed using GraphPad Prism version 9.5.1. Differences between groups were assessed using ANOVA followed by Tukey’s test, with statistical significance set at P<0.05, P<0.01 and P<0.001.

    RESULTS AND DISCUSSION

    Extraction yields
     
    The use of hydro-methanol as the solvent in maceration extraction enabled the calculation of the extraction rate. The total polyphenol yield obtained from P. odora leaves was 17.1% (w/w dry plant material). In comparison, methanolic crude extracts of P. crispa and P. undulata, wild species from Sudan, yielded 22.6% and 23%, respectively (Hegazy et al., 2021). This slight variation in yield percentage may be attributed to various factors, such as the extraction method, particle size, duration of extraction, solvent type and solvent volume (De la Luz Cádiz Gurrea et al., 2019).
     
    Total flavonoids and total phenolic contents
     
    To standardize the hydro-methanolic extract of P. odora leaves, the total phenolic content was measured using the folin-ciocalteu method and the UV-Vis colorimetric method was employed to quantify the total flavonoid content. Results showed a flavonoid concentration of 0.77 mg quercetin equivalent per gram of dry extract and a phenolic content of 2.77 mg gallic acid equivalent. Compared to these findings, Touati et al., (2018) reported lower total polyphenol and flavonoid levels in the methanolic extract of P. odora leaves, at 90±0.63 µg GAE/g DWE and 11.34±3.15 µg QE/g DWE, respectively. Additionally, the authors observed that P. odora leaves contain significantly higher levels of flavonoids and total phenolics than the roots. Prior studies also indicate that plants within the Pulicaria genus typically contain high levels of phenolics and flavonoids. For example, Senhaji et al., (2017) identified total phenolics and flavonoids in the ethyl acetate extract of P. murritanica at 72.88±0.21 mg GAE/g DWE and 38.95±0.75 mg QE/g DWE, respectively. Similarly, Malarz et al., (2023) reported that hydroalcoholic extracts of P. inuloides contain 43.81±2.36 mg GAE/g DWE of total polyphenols. Various factors may explain these differences, including genetic variation, plant maturity, storage duration and environmental conditions such as high temperatures, intense sunlight, drought and salinity, which are known to enhance secondary metabolite production, particularly polyphenols (Astill et al., 2001; Martínez et al., 2021; Turkmen et al., 2009).
     
    Antioxidant activity
     
    Two methods were employed to assess the antioxidant effects of P. odora cured extracts: DPPH radical scavenging activity and ferric ion (FeCl3) reducing power. The hydro-methanolic leaf extract of P. odora demonstrated dose-dependent DPPH radical scavenging, with inhibition rates of 44.02%, 48%, 48.6%, 50.27%, 54.27%, 59.24% and 63.4% at concentrations of 80, 100, 120, 140, 160, 180 and 200 µg/mL, respectively (Fig 1). A strong positive correlation was observed between extract concentration and radical inhibition, indicating increased antioxidant activity with higher concentrations. The IC50 for ascorbic acid was 103.33 µg/mL, while the IC50 for the P. odora extract was 123.69 µg/mL.

    Fig 1: DPPH radicals scavenging activity of P. odora.


           
    To further validate antioxidant activity, the reducing power test was conducted. Results showed a direct correlation between absorbance at 700 nm and extract concentration, where increased absorbance reflected the reduction of ferric ions by extract components. The inhibition percentages for the crude extract were 11.33%, 14.62%, 32.52%, 48.44%, 75.34% and 90.2%, compared to ascorbic acid’s 34.66%, 45.66%, 57.03%, 69.62%, 79.83% and 97.21% at 15.62, 31.25, 62.5, 125, 250 and 500 µg/mL, respectively (Fig 2). The IC50 values were 42.56 µg/mL for ascorbic acid and 192.24 µg/mL for the crude extract. Similar antioxidant responses have been reported for medicinal plant extracts, where phenolic-rich extracts demonstrated substantial DPPH and FRAP activities and improved antioxidant status in experimental rats (Azyu et al., 2025).

    Fig 2: Antioxidant activity of methanol extract from leaves of P.odora using FRAP assay.


     
    Sub-acute toxicity study
     
    The current study investigates the potential detoxifying effects of P. odora on metribuzin-induced toxicity in Wistar albino rats. A comprehensive evaluation of physiological parameters, including body weight, hematological indices and biochemical markers, was conducted to elucidate the impact of both P. odora and metribuzin. Behavioral observations indicated that the oral administration of P. odora extract did not result in mortality or exhibit any toxicological symptoms, thereby underscoring the extract safety profile and potential therapeutic efficacy.
     
    Growth parameters
     
    Fig 3 displays rat body weight data across the experimental groups, highlighting differences among the four groups. At the beginning of the study, the initial body weight of the experimental rats averaged 170±10.2 g. A significant reduction in average body weight was observed in the GM group, which received metribuzin alone, compared to the control group (p<0.001). This weight loss is likely due to decreased food intake during the study period. These results align with findings by Samir and Asma (2018), who reported similar weight reduction effects following metribuzin exposure in rabbits over 6 to 18 days. In contrast, co-administration of P. odora leaf extract with metribuzin in the GPM group effectively prevented weight loss, showing no statistically significant difference from the control group. The phenolic compounds in P. odora extract are likely instrumental in counteracting rat metribuzin-induced oxidative stress (Zeng et al., 2021).

    Fig 3: Average body weight of male albino wistar rats (n = 9) over a 36-day treatment period, across four groups.


     
    Hematological and immune response to metribuzin and P. odora extract in rats
     
    Fig 4 illustrates the hematological responses to metribuzin and P. odora extract treatments, highlighting notable changes in these parameters. In the GM group, which received metribuzin alone, significant elevations were observed in red blood cell (RBC) counts and hematocrit levels (P<0.01), along with a more pronounced increase in white blood cell (WBC) counts, lymphocytes, hemoglobin (Hb), mean corpuscular volume (MCV) and platelets (PLTs) (P<0.001) compared to the control. In contrast, co-treatment with P. odora extract in the GMP group led to a marked reduction in platelets, Hb, WBC, RBC and MCV levels relative to the GM group, while lymphocyte and hematocrit levels remained unaffected. Rats administered only P. odora extract (GP) showed a significant increase in RBC counts (P<0.01) and a minor but significant decrease in MCV (P<0.05) when compared to the control group, with other hematological parameters showing no significant differences.

    Fig 4: The hematological profile of rats intoxicated with metribuzin and after treatment with methanolic extract of P. odora.


           
    The observed elevation in RBC counts, MCV, hematocrit and Hb levels in the GM group may reflect a physiological stress response to metribuzin exposure, potentially due to compensatory mechanisms aimed at countering herbicide-induced oxidative stress. This response likely involves increased oxygen transport to cells, evidenced by the rise in RBC counts, possibly through enhanced release or synthesis of erythrocytes from hematopoietic tissues. Similar responses have been reported by Lutnicka et al., (2019) in C. carpio exposed to linuron. Furthermore, (Kadeche et al., 2016) and Samir et al. (2020) observed declines in RBC, Hb and hematocrit levels and reduced erythrocyte counts in metribuzin-treated rats, particularly with prolonged or high-dose exposure, attributing these effects to oxidative damage and hematotoxicity.
           
    Metribuzin-treated rats in the present study exhibited significantly increased lymphocyte, platelet and WBC counts (P<0.001), likely indicating an immune response to herbicide-induced toxicity. Elevated WBC and lymphocyte levels may suggest an immune mobilization against tissue damage, consistent with findings from Samir et al. (2020), who reported similar leukocyte increases in response to metribuzin’s cytotoxic effects. The rise in platelet count may stem from metribuzin-related tissue damage, as platelets contribute to inflammatory responses by releasing pro-inflammatory mediators to recruit immune cells (Chen et al., 2020). Additionally, metribuzin’s oxidative stress-inducing properties (Almeida et al., 2019) may further explain the increase in platelet levels as a compensatory response to RBC suppression.
           
    Treatment with P. odora extract alone induced minor changes in RBC counts and MCV relative to the GP and control groups, similar to findings by Mansouri et al. (2015) in studies with Foeniculum vulgare extracts. This effect may result from erythropoietin stimulation by the liver and kidneys in response to P. odora. The phenolic constituents of P. odora likely contribute antioxidant effects, stabilizing cell membranes and protecting against free radical damage (Koren et al., 2010). Although changes in platelet, MCV and RBC levels were not statistically significant, they suggest a potential protective effect of P. odora. Moreover, Hb and WBC levels in the GM group showed a significant reduction (P<0.001) relative to controls, while these levels remained elevated in the GP group, further supporting P. odora’s stabilizing role.
           
    Collectively, these results imply that oxidative stress plays a central role in metribuzin toxicity, with P. odora extract, rich in polyphenols and flavonoids, offering significant antioxidant and free radical-scavenging properties that may counteract metribuzin-induced damage. This aligns with findings by Kadeche et al. (2017) on vanillin’s protective effect against metribuzin-induced hematological alterations, underscoring P. odora’s potential as a therapeutic agent against herbicide toxicity.
     
    Biochemical effects of metribuzin and P. odora extract in rats
     
    In this study, exposure to metribuzin (GM group) significantly increased (P<0.001) several biochemical markers, including blood glucose, total cholesterol, triglycerides, glutamate pyruvate transaminase (GPT), glutamate oxaloacetate transaminase (GOT), urea and creatinine, relative to the control group (Fig 5). These elevations suggest a toxic impact on liver and kidney function, as evidenced by raised transaminase levels and markers of renal function. Interestingly, the co-treatment group (GMP), which received P. odora extract alongside metribuzin, exhibited significant reductions in GPT, GOT and creatinine (P<0.001), urea (P<0.01), as well as total cholesterol and triglycerides (P< 0.05), compared to the GM group. This indicates that P. odora may confer a protective effect, mitigating metribuzin-induced hepatic and renal toxicity. However, co-treatment did not significantly affect blood glucose levels, suggesting limited influence on glucose metabolism. Additionally, the group receiving only P. odora extract (GP group) showed a mild, statistically insignificant increase in GPT, GOT and triglycerides compared to the control group, with no significant changes in other biochemical parameters, indicating that P. odora alone does not markedly disrupt biochemical homeostasis. Changes in hematological indices and serum biomarkers of hepatic and renal function are commonly used to evaluate the systemic effects and safety of bioactive plant extracts in Wistar rats (Al-Ghamdi et al., 2026). Collectively, these findings support the potential protective role of P. odora against metribuzin-induced toxicity.

    Fig 5: Intoxicated rats’ biochemical parameters with metribuzin and after treatment with a methanolic extract of P. odora.


           
    This study also noted a marked increase in blood glucose levels in rats exposed to metribuzin (13.2 mg/mL over 36 days), consistent with previous findings indicating that chronic low-level metribuzin exposure elevates plasma glucose, possibly due to reduced glucose uptake and impaired tissue functionality (Chiali et al., 2013). Similar glucose elevations were reported in Cyprinus carpio after atrazine exposure (Blahova et al., 2014), suggesting that pesticides may disrupt carbohydrate metabolism. Potential mechanisms include increased glycogenolysis, elevated adrenocorticotropic and glucagon hormones, or insulin dysfunction (Mehra et al., 2014). Conversely, studies on long-term, low-dose metribuzin exposure in rats (1/20 to 1/5 of LD50, twice weekly) reported decreased glucose levels, possibly due to depleted carbohydrate reserves or inhibition of glucose release from hepatic tissue (Maksymiv et al., 2015; Samir et al., 2020). Metribuzin exposure also increased lipid parameters, particularly triglycerides and total cholesterol, potentially due to enhanced cholesterol synthesis in the liver or compromised liver cell membrane integrity. Elevated triglycerides may result from decreased lipase activity and increased mobilization of adipose stores (Chiali et al., 2013; El-Demerdash and Nasr 2014). Some studies, however, have reported reduced plasma cholesterol under metribuzin exposure, potentially due to diminished cholesterol synthesis or tissue accumulation.
           
    Increased urea and creatinine levels in metribuzin-treated rats indicate renal impairment and reduced glomerular filtration rate, consistent with findings in albino rats exposed to metribuzin (Khozimy et al., 2017). Elevated liver biomarkers GPT and GOT indicate hepatotoxicity, aligning with Samir and Asma (2018), who observed increased transaminase activity in metribuzin-exposed rabbits and fetuses. The herbicide’s residual accumulation in the liver, as confirmed by HPLC analysis, underscores hepatic vulnerability to metribuzin toxicity. Similarly, Sena et al. (2021) found heightened GPT and GOT in Xenopus laevis exposed to atrazine. Rats treated with P. odora extract alone (GP group) exhibited slight, non-significant increases in triglycerides, GOT and GPT, potentially indicating activation of lipid metabolism-related enzymes. However, no studies to date have specifically examined the biochemical effects of P. odora, underscoring the need for further investigation.
           
    Importantly, co-treatment with P. odora extract and metribuzin mitigated the metabolic disturbances induced by metribuzin, restoring levels of GPT, GOT, urea, creatinine, cholesterol and triglycerides to near normal. This protective effect is likely due to the natural antioxidants, particularly polyphenols, in P. odora, which may counteract metribuzin-induced biochemical imbalances. These findings align with previous studies on P. crispa extract, which lowered cholesterol, urea and creatinine in diabetic rats (Daradka et al., 2021) and support evidence of the benefits of Pulicaria genus on liver and kidney health (Bakr et al., 2021; Alharthi et al., 2023). Similar antioxidant compounds, such as vanillin and quercetin, have demonstrated efficacy in ameliorating metribuzin-induced biochemical changes (Abarikwu 2014; Kadeche et al., 2017), suggesting that P. odora’s quercetin content may contribute to its protective properties.

    CONCLUSION

    The findings of this study demonstrate that P. odora extract possesses significant protective effects against metribuzin-induced toxicity in Wistar rats. The extract’s antioxidant properties, attributed to its phenolic and flavonoid content, played a key role in reducing oxidative stress and mitigating the physiological and biochemical alterations induced by metribuzin. Co-administration of P. odora extract resulted in a marked improvement in hematological and biochemical parameters, including liver and kidney function markers. The absence of adverse effects on behavioural assessments further supports the safety and potential therapeutic value of P. odora. These results suggest that P. odora may serve as a promising natural agent for combating herbicide-induced toxicity. Further research is warranted to explore its bioactive compounds, mechanisms of action and the therapeutic potential of other species within the Pulicaria genus.

    ACKNOWLEDGEMENT

    The authors would like to thank the Algerian Ministry of Higher Education and Scientific Research for providing the necessary facilities. They also extend their heartfelt gratitude to the Princess Nourah bint Abdulrahman University Researchers Supporting Project number (PNURSP2026R931), Princess Nourah bint Abdulrahman University, Riyadh,Saudi Arabia.
     
    Disclaimers
     
    The views and conclusions expressed in this article are solely those of the authors and do not necessarily represent the views of their affiliated institutions. The authors are responsible for the accuracy and completeness of the information provided, but do not accept any liability for any direct or indirect losses resulting from the use of this content.
     
    Authors’ contributions
     
    Conceptualization: A. Benmahammed and ABA, Data curation: ABA and F. Bouleknafet, Formal analysis: F.A. Al-Mekhlafi and N. Abutaha, Funding acquisition: M.A. Wadaan and H.M. Alharbi Investigation: M.A. Wadaan, Methodology: A. Benmahammed and ABA, Project administration: F. Bouleknafet and S. Lambiase Resources: F.A. Al-Mekhlafi. Supervision: ABA. Writing-original draft: ABA, F.A. Al-Mekhlafi and F. Bouleknafet Writing-review and editing: N. Abutaha, F.A. Al-Mekhlafi and H.M. Alharbi.
     
    Funding
     
    This project was funded by Princess Nourah bint Abdulrahman University Researchers Supporting Project number (PNURSP2026R931), Princess Nourah bint Abdulrahman University, Riyadh,Saudi Arabia.
     
    Data availability statement
     
    All the data is available within the manuscript.
     
    Informed consent
     
    All animal procedures for experiments were approved by the Committee of Experimental Animal care and handling techniques were approved by the University of Animal Care Committee.

    CONFLICT OF INTEREST

    The authors declare that there are no conflicts of interest regarding the publication of this article. No funding or sponsorship influenced the design of the study, data collection, analysis, decision to publish, or preparation of the manuscript.

    REFERENCES


    1. Abarikwu, S.O. (2014). Protective effect of quercetin on atrazine- induced oxidative stress in the liver, kidney, brain and heart of adult Wistar rats. Toxicology International. 21: 148-155.

    2. Al-Ghamdi, N.A., Omer, S.A., Alterary, S.S., El-Tohamy, M.F., Virk, P., El-Nagar, D.M. and Mohammed, O.B. (2026). Hepatorenal toxicity with hematological and biochemical changes in wistar rats fed the ethanolic extract of the pulp of Citrullus colocynthis from two regions in Saudi Arabia. Indian Journal of Animal Research. 60(6): 1030-1037. doi: 10.18805/IJAR.BF-2006.

    3. Alharthi, A.S., Alruwaili, N.W., Al-Baadani, H.H., Al-Garadi, M.A., Shamlan, G., Alhidary, I. A. (2023). Investigating the effect of Pulicaria jaubertii as a natural feed additive on the growth performance, blood biochemistry, immunological response and cecal microbiota of broiler chickens. Animals. 13: 1116.

    4. Almeida, A.C., Gomes, T., Langford, K., Thomas, K.V. and Tollefsen, K.E. (2019). Oxidative stress potential of the herbicides bifenox and metribuzin in the microalgae Chlamydomonas reinhardtii. Aquatic Toxicology. 210: 117-128.

    5. Astill, C., Birch, M.R., Dacombe, C., Humphrey, P.G. and Martin, P.T. (2001). Factors affecting the caffeine and polyphenol contents of black and green tea infusions. Journal of Agricultural and Food Chemistry. 49: 5340-5347.

    6. Azyu, R.Z., Ali Ayub, M., Gali Mohanarao, J., Behera, P., Das, H., Mayengbam, P. and Devi, I.L. (2025). Effect of ethanolic extract of Scurrula parasitica L. on in vivo antioxidant status of diabetic rats. Indian Journal of Animal Research. doi: 10.18805/IJAR.B-5462.

    7. Bakr, R.O., Shahat, E.A., Elissawy, A.E., Fayez, A.M. and Eldahshan, O.A. (2021). Evaluation of the hepatoprotective activity of Pulicaria incisa subspecies candolleana and in silico screening of its isolated phenolics. Journal of Ethnopharmacology. 271: 113767.

    8. Blahova, J., Modra, H., Sevcikova, M., Marsalek, P., Zelnickova, L., Skoric, M. and Svobodova, Z. (2014). Evaluation of biochemical, haematological and histopathological responses and recovery ability of common carp (Cyprinus carpio L.) after acute exposure to atrazine herbicide. Biomedical Research International. 2014: 980948.

    9. Chen, Y., Zhong, H., Zhao, Y., Luo, X., Gao, W. (2020). Role of platelet biomarkers in inflammatory response. Biomarker Research. 8: 1-7.

    10. Chiali, F., Merzouk, H., Merzouk, S., Medjdoub, A. and Narce, M. (2013). Chronic low-level metribuzin exposure induces metabolic alterations in rats. Pesticide Biochemistry and Physiology. 106: 38-44.

    11. Close, M. and Banasiak, L. (2022). National Survey of Pesticides in Groundwater 2022.

    12. Daradka, H.M., Aldhilan, A.M., Eskandrani, A.A., Bataineh, Y. and Alzoubi, K.H. (2021). The effect of Pulicaria crispa ethanolic extract on haematological and biochemical parameters in alloxan-induced diabetic rats. Advances in Traditional Medicine. 21: 65-72.

    13. De la Luz Cádiz-Gurrea, M., Zengin, G., Kayacik, O., Lobine, D., Mahomoodally, M.F., Leyva-Jiménez, F.J. and Segura- Carretero, A. (2019). Innovative perspectives on Pulicaria dysenterica extracts: Phyto-pharmaceutical properties, chemical characterization and multivariate analysis. Journal of the Science of Food and Agriculture. 99: 6001-6010.

    14. Djilali, A.B., Benseddik, A., Bouacem, K., Allaf, K. and Nabiev, M. (2020). Functional properties of Pulicaria odora L. leaves pre-coated in gel-based Ziziphus jujuba Mill. peel powder. Nature Products in Agro-Industrial Processing. 6: 1-4.

    15. Djordjević, M., Djordjević, M., Starowicz, M. and Krupa-Kozak, U. (2024). Plant-based antioxidants in gluten-free bread production: Sources, technological and sensory aspects, enhancing strategies and constraints. Antioxidants. 13: 142.

    16. El-Demerdash, F.M and Nasr, H.M. (2014). Antioxidant effect of selenium on lipid peroxidation, hyperlipidemia and biochemical parameters in rats exposed to diazinon. Journal of Trace Elements in Medicine and Biology. 28: 89-93.

    17. El-Ghaly, E.S.M., Shaheen, U., Ragab, E., El-Hila, A.A. and Abd- Allah, M.R. (2016). Bioactive constituents of Pulicaria jaubertii: A promising antihypertensive activity. Pharmacognosy Journal. 8: 81-86.

    18. El-Sabagh, O.A., El-Toumy, S.A., Mounir, R., Farag, M.A. and Mahrous, E.A. (2021). Metabolite profiles of Pulicaria crispa and P. incisa in relation to their in vitro/in vivo antioxidant activity and hepatoprotective effect: A comparative mass spectrometry-based metabolomics. Journal of Pharmaceutical and Biomedical Analysis. 194: 113804.

    19. Gitsopoulos, T., Georgoulas, I., Vazanelli, E. and Botsoglu, D. (2024). Selectivity of the premixtures flufenacet, diflufenican and flufenacet, diflufenican, metribuzin on bread wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) and efficacy on ALS/ACCase-resistant populations of lolium rigidum gaudin. Agronomy. 14: 949.

    20. Gupta, H.V., Sorooshian, S. and Yapo, P.O. (1999). Status of automatic calibration for hydrologic models: Comparison with multilevel expert calibration. Journal of Hydrologic Engineering. 4: 135-143.

    21. Hegazy, A.E. and Ibrahium, M.I. (2012). Antioxidant activities of orange peel extracts. World Applied Sciences Journal. 18: 684-688.

    22. Hegazy, M.E.F., Dawood, M., Mahmoud, N., Elbadawi, M., Sugimoto, Y., Klauck, S.M., Mohamed, N. and Efferth, T. (2021). 2α- Hydroxyalantolactone from Pulicaria undulata: Activity against multidrug-resistant tumor cells and modes of action. Phytomedicine. 81: 153409.

    23. Hemma, R., Belhadj, S., Ouahchia, C. and Saidi, F. (2018). Antioxidant activity of Pistacia lentiscus methanolic extracts. Revista de Agrobiología. 8: 845-852.

    24. Hussain, S., Sagar, T., Kaur, S., Nipuntha, K., Kapoor, N. and Mahajan, R. (2023). Omics Approaches to Study the Biosynthesis of Bioactive Compounds in Medicinal and Aromatic Plants. In Biosynthesis of Bioactive Compounds in Medicinal and Aromatic Plants: Manipulation by Conventional and Biotechnological Approaches. Springer. (pp. 41-64).

    25. Jeong, S.M., Kim, S.Y., Kim, D.R., Jo, S.C., Nam, K., Ahn, D. and Lee, S.C. (2004). Effect of heat treatment on the antioxidant activity of extracts from citrus peels. Journal of Agricultural and Food Chemistry. 52: 3389-3393.

    26. Kadeche, L., Bourogaa, E., Boumendjel, A., Djeffal, A., El Feki, A. and Messarah, M. (2016). Quercetin attenuates metribuzin- induced biochemical and hematological toxicity in adult rats. International Journal of Pharmaceutical Sciences Review and Research. 40: 38-46.

    27. Kadeche, L., Bourogaa, E., Saoudi, M., Boumendjel, A., Djeffal, A., El Feki, A. and Messarah, M. (2017). Ameliorative effects of vanillin against metribuzin-induced oxidative stress and toxicity in rats. International Journal of Pharmaceutical and Pharmaceutical Sciences. 9: 56-62.

    28. Khozimy, A.M., Ramadan, M.F., Shaldam, H.A. and Aly, T.A. (2017). Effect of storage temperature on degradation of two herbicides and their sub-acute toxicity on albino rats. Journal of Applied Plant Protection, Suez Canal University. 6: 1.

    29. Koren, E., Kohen, R. and Ginsburg, I. (2010). Polyphenols enhance total oxidant-scavenging capacities of human blood by binding to red blood cells. Experimental Biology and Medicine. 235: 689-699.

    30. Kumar, J.S., Monica, S.S., Vinothkumar, B., Suganthi, A. and Paramasivam, M. (2024). Impact of pesticide exposure on environment and biodiversity: A review. Agricultural Reviews. 45(1): 01-12. doi: 10.18805/ag.R-2325.

    31. Liu, L.L., Yang, J.L. and Shi, Y.P. (2010). Phytochemicals and biological activities of Pulicaria species. Chemistry Biodiversity. 7: 327-349.

    32. Lutnicka, H., Bojarski, B., Witeska, M., Tombarkiewicz, B. and Formicki, G. (2019). Exposure to herbicide linuron results in alterations in hematological profile and stress biomarkers of common carp (Cyprinus carpio). Ecotoxicology. 28: 69-75.

    33. Maksymiv, I.V., Husak, V.V., Mosiichuk, N.M., Matviishyn, T.M., Sluchyk, I.Y., Storey, J.M., Storey, K.B. and Lushchak, V.I. (2015). Hepatotoxicity of herbicide Sencor in goldfish may result from induction of mild oxidative stress. Pesticide Biochemistry and Physiology. 122: 67-75.

    34. Malarz, J., Michalska, K., Galanty, A., Kiss, A.K. and Stojakowska, A. (2023). Constituents of Pulicaria inuloides and cytotoxic activities of two methoxylated flavonols. Molecules. 28: 480.

    35. Mansouri, E., Kooti, W., Bazvand, M., Boroon, M.G., Amirzargar, A., Afrisham, R., Afzalzadeh, M.R., Ashtary-Larky, D. and Jalali, N. (2015). The effect of hydro-alcoholic extract of Foeniculum vulgare Mill on leukocytes and hematological tests in male rats. Jundishapur Journal of Natural Pharmaceutical Products. 10(1).

    36. Martínez, C., Valenzuela, J.L. and Jamilena, M. (2021). Genetic and pre-and postharvest factors influencing the content of antioxidants in cucurbit crops. Antioxidants. 10(6): 894.

    37. Mehra, B., Sharma, P., Kaushik, U. and Joshi, S. (2014). Effect of fytolan on haematology and serum parameters of male albino rats. World Journal of Pharmaceutical Sciences. 3: 817-829.

    38. Naqvi, S.A.R., Shah, S.M.A., Kanwal, L., Saeed, M., Atta-ul-Haq, Nisar, J., Nisar, Z. and Akram, M. (2020). Antimicrobial and antihypercholesterolemic activities of Pulicaria gnaphalodes. Dose-Response. 18(1): 1559325820904858.

    39. Pelić, M., Kartalović, B., Živkov-Baloš, M., Mirolić, M., Đorđević, M., Teodorović, V., Ćirković, M., Ljubojević-Pelić, D. (2020). Health risks associated with residual pesticide levels in fish reared in purified wastewater from slaughterhouse. Journal of the Hellenic Veterinary Medical Society. 71(1): 1991-1996.

    40. Samir, D and Asma, S. (2018). Triazinone herbicide metribuzin induced acute liver injury: A study of animal model. Journal of Acute Diseases. 7(4): 152-157.

    41. Samir, D., Selma, R.M.O. and Asma, S. (2020). The effect of herbicide metribuzin on environment and human: A systematic review. Pharmaceutical Biosciences Journal. pp 10-15.

    42. Sena, L., Asouzu Johnson, J., Nkomozepi, P. and Mbajiorgu, E.F. (2021). Atrazine-induced hepato-renal toxicity in adult male Xenopus laevis frogs. Applied Science. 11(24): 11776.

    43. Senhaji, B., Chebli, B., Mayad, E., Hamdouch, A., Heimeur, N., Chahid, A. and Ferji, Z. (2017). Phytochemical screening, quantitative analysis and antioxidant activity of Asteriscus imbricatus and Pulicaria mauritanica organic extracts. International Food Research Journal. 24(6): 2482-2489.

    44. Sousa, S., Maia, M., Correira-Sá, L., Fernandes, V., Delerue-Matos, C., Calhau, C. and Domingues, V. (2020). Chemistry and toxicology behind insecticides and herbicides. In Controlled Release of Pesticides for Sustainable Agriculture. pp 59-109.

    45. Touati, N., Saidani, K., Boudries, H., Hammiche, H., Ouazene, N. and Bedjou, F. (2018). Antibacterial activity of phenolic compounds of Pulicaria odora, wild plant in northern Algeria. International Food Research Journal. 25(5): 2021-2030.

    46. Turkmen, N., Sari, F. and Velioglu, Y.S. (2009). Factors affecting polyphenol content and composition of fresh and processed tea leaves. Akademik Gida. 7(6): 29-40.

    47. Zeng, X., Du, Z., Ding, X. and Jiang, W. (2021). Protective effects of dietary flavonoids against pesticide-induced toxicity: A review. Trends in Food Science and Technology. 109: 271-279.
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