In vitro study of anti-coccidial activity of Ocimum gratissimum and Vernonia amygdalina leaves extracts against Eimeria magna and Eimeria media

In vitro study of anti-coccidial activity of Ocimum gratissimum and Vernonia amygdalina leaves extracts against Eimeria magna and Eimeria media | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article In vitro study of anti-coccidial activity of Ocimum gratissimum and Vernonia amygdalina leaves extracts against Eimeria magna and Eimeria media Basile Konmy, Cocou Christian Dansou, Lissette Dègla, Rodrigue Towanou, and 9 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6198606/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Background Ocimum gratissimum and Vernonia amygdalina are used in traditional medicine for their various biological properties, but few studies have assessed their antioxidant and anticoccidial capacities and toxicity. The aim of this study was to evaluate the antioxidant capacity, anticoccidial activity (against Eimeria magna and Eimeria media ), phytochemical content and toxicity of leaves extracts from two plants. Methodology: Aqueous and acetonic extracts of the leaves of both plants were prepared and analysed for their polyphenol, flavonoid and tannin contents. Antioxidant activity was measured via the DPPH method. The anticoccidial effects of increasing concentrations of extracts were tested on Eimeria oocysts and sporozoites. Toxicity was assessed by mortality tests on Artemia salina larvae and clinical tests on rats. Result The results show that extracts of V. amygdalina leaves have greater antioxidant and anticoccidial capacities than those of O. gratissimum . The acetone extracts of the two plants presented lower EC 50 values (2.4025 ± 0.0385) for O. gratissimum and 3.541 ± 0.112 for V. amygdalina ) than did the aqueous extracts. With respect to anti-ocyst activity, acetone extracts of V. amygdalina inhibited sporulation of E. magna oocysts by up to 91% at 40 mg/mL (p˂0.05). Toxicity tests revealed that the extracts were not toxic to Artemia salina . No major histological changes were observed. O. gratissimum and V. amygdalina extracts have significant antioxidant and anticoccidial properties and are nontoxic for antioxidant and anticoccidial applications. Conclusion This study shows that extracts of O. gratissimum and V. amygdalina possess antioxidant and anticoccidial properties, with superior efficacy for V. amygdalina . These extracts have potential as natural alternatives for the management of coccidiosis in farm animals, helping to reduce the use of synthetic antiparasitics and promote sustainable agricultural practices. Further studies on their long-term effects and their application in livestock farming are required. Natural coccidiostat Oxidative stress Medicinal plants Bioactive substances Biocompatibility Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction Coccidial infections caused by protozoa of the genus Eimeria are a major threat to the health of farm animals, particularly rabbits. These infections can cause serious gastrointestinal disorders and affect the growth, reproduction and productivity of animals [ 1 , 2 ]. E. magna and E. media are two species of coccidia commonly observed in rabbits and are responsible for considerable economic losses on farms due to clinical symptoms such as diarrhea, dehydration and reduced production [ 3 ]. Although anticoccidial drugs are available to treat these infections, their overuse has led to the emergence of resistance, reducing their efficacy [ 4 ]. In addition, growing concerns about food safety and antibiotic resistance have encouraged the search for alternative treatments, including plant extracts with therapeutic properties. Medicinal plants play a central role in traditional medicine in many parts of the world, particularly in Africa and Asia, where they are used to treat a variety of conditions, including parasitic infections. Among these plants, O. gratissimum and V. amygdalina have attracted particular interest because of their antioxidant, antimicrobial and antiparasitic properties [ 5 , 6 ]. These plants are distinguished by their exceptional phytochemical composition, including flavonoids, tannins, alkaloids, saponins and essential oils [ 7 , 8 ]. O. gratissimum is known for its high content of phenolic compounds, particularly eugenol, a bioactive phenol with antioxidant, anti-inflammatory and antimicrobial properties [ 9 ]. O. gratissimum , which is rich in essential oils, has antioxidant and antimicrobial activities, as confirmed by several studies [ 10 , 11 ]. These properties indicate its therapeutic potential in the management of intestinal infections and metabolic disorders linked to oxidative stress, which are frequently observed in parasitic infections. In addition, its antimicrobial and antiparasitic effects suggest promising activity against protozoa, although few studies have explored its anticoccidial potential. V. amygdalina , also known as the “bitter leaf”, is rich in saponins, alkaloids and flavonoids. These compounds are known for their antiparasitic, immunomodulatory and antioxidant activities [ 8 , 12 ]. Saponins, in particular, have demonstrated significant antiparasitic activity by disrupting parasite membranes or modulating the host immune response [ 13 ]. In addition, V. amygdalina leaves are traditionally used to treat gastrointestinal disorders and intestinal infections, reinforcing their relevance in the context of coccidiosis [ 14 ]. These two plants have complementary properties. O. gratissimum has been studied for its antimicrobial and antioxidant properties, whereas V. amygdalina is better known for its immunomodulatory and antiparasitic abilities. Together, they offer synergistic potential in the fight against E. magna and E. media , two species of interest because of their economic and health impacts. However, despite these promising observations, scientific evidence concerning the efficacy of these plants against Eimeria in rabbits, as well as an understanding of their mechanisms of action, remains limited. However, to consider their practical application in zootechnics, it is crucial to assess not only their efficacy but also their safety. Although plant extracts are natural, they may contain toxic compounds, and their potential toxicity at high doses or with prolonged use needs to be rigorously studied [ 9 ]. Indeed, the potential cytotoxic effects of these plants on parasite larvae, as well as their systemic toxicity in animal hosts, remain poorly explored. These plants contain various secondary metabolites, such as saponins, flavonoids, alkaloids and essential oils, which can have powerful biological effects but are sometimes toxic [ 15 , 16 ]. Assessment of larval cytotoxicity is essential for determining whether these extracts effectively target parasites without causing collateral damage to host tissues. Moreover, assessing acute toxicity in animals makes it possible to establish safe doses and avoid side effects such as liver or kidney damage, metabolic imbalances or impaired growth [ 17 ]. Given this lack of data, this study proposes an in-depth evaluation of the antioxidant capacities and in vitro anticoccidial activity of O. gratissimum and V. amygdalin extracts against E. magna and E. media strains, as well as a toxicity study. This study also examined the phytochemical content of these plants to identify the bioactive compounds potentially responsible for their therapeutic effects. This research aims to answer the following questions: do extracts of O. gratissimum and V. amygdalina exert anticoccidial activity in vitro against E. magna and E. media , and what is their possible mode of action? What is the larval cytotoxicity and acute toxicity of extracts of V. amygdalina and O. gratissimum ? The hypotheses of this study are that extracts of O. gratissimum and V. amygdalina display significant antioxidant and anticoccidial activity in vitro against E. magna and E. media without any toxicity and that this activity is associated with the contents of bioactive compounds such as flavonoids, alkaloids and tannins. This study is motivated by the need to develop safer and more sustainable alternative therapeutic solutions for coccidiosis in rabbits. The use of plant extracts could not only offer a natural alternative to chemical treatments but also help to preserve biodiversity and reduce the risks associated with antibiotics. In addition, this study could pave the way for the use of O. gratissimum and V. amygdalina in integrated pest management protocols, thereby contributing to a more sustainable approach to animal disease management. The aim of this study is therefore twofold: (i) to assess the in vitro antioxidant and anticoccidial activity of extracts of O. gratissimum and V. amygdalina against E. magna and E. media ; (ii) to identify the bioactive compounds present in these plants that are likely to play a key role in these activities; and (iii) to assess the larval and acute oral toxicity of the leaves of both plants. In doing so, this study aims to provide a sound scientific basis for the use of these plants as alternative treatments for the control of parasitic infections in rabbits. The ultimate aim of this research is to offer a natural and effective alternative to chemical treatments while helping to combat the increase in drug resistance in rabbit farms. Materials and Methods Harvesting O. gratissimum and V. amygdalina The present study used the leaves of O. gratissimum and V. amygdalina . O. gratissimum and V. amygdalina leaves were harvested in April 2019 from market gardeners at the University of Abomey-Calavi (UAC) in Benin. These plants were subsequently grown on organic fertilizers containing compost. These plants were then authenticated with voucher numbers YH524/HNB and YH523/HNB, respectively, at the herbarium of UAC. The harvested leaves were washed in clean water and then dried in the laboratory at 18°C for 14 days. Once dry and crunchy, the leaves were ground into a powder and stored in hermetically sealed glass jars for later use. Preparation of aqueous and acetonic extracts of O. gratissimum and V. amygdalina leaves The leaves powders of both plants were used as materials for the extractions. Fifty grams of plant powder was dissolved in 500 mL of solvent S1, which was composed of 70% acetone and 30% distilled water. After 72 h of continuous maceration, the mixture was filtered through Whatman No. 1 paper, and the residue was recovered several times until exhaustion. The filtrate was evaporated under vacuum at 47°C using a Rotavapor (BUCHI RII). The acetonic extract was then dried in an oven at 40°C for 7 days. The aqueous extract was obtained by dissolution via a previously described procedure. This time, solvent S1 was replaced with distilled water. The filtrate was evaporated under vacuum at 65°C using a Rotavapor (BUCHI RII). The aqueous extract was then dried in an oven at 50°C for 7 days. Phytochemical screening of powder from O. gratissimum and V. amygdalina leaves The qualitative determination of the major groups of chemical compounds contained in the leaves of O. gratissimum and V. amygdalina was carried out according to the classical method of Houghton and Raman [ 18 ], which was reviewed and adapted to the conditions of the Laboratory of Biotechnologies and Animal Improvement of the Faculty of Agronomic Sciences of UAC. Total Phenol Assay The quantitative determination of total phenols was performed according to the classical method of Singleton, et al. [ 19 ]. Starting from an aqueous stock solution of gallic acid with a mass concentration of 10 mg/mL, a standard range of aqueous solutions obtained by successive dilution was prepared. A total of 625 µL of Folin-Ciocalteu reagent (FCR) was added to 125 µL of the prepared extract at 1 mg/mL in distilled water. The mixture was incubated for 5 minutes, after which 500 µL of sodium carbonate Na 2 CO3 at 75 mg/mL and 4.75 mL of distilled water were added. After 30 minutes of incubation in the dark, the optical density was read at 760 nm via a Bio Mate 5 spectrophotometer against a blank. The total phenol content was calculated via the calibration curve of 10 mg/mL gallic acid prepared under the same conditions. Total Flavonoid Assay Five hundred microliters of the extract at 1 mg/mL in methanol was mixed with 500 µL of 2% AlCl3 and 3 mL of methanol. The mixture was vigorously shaken. After 10 minutes of incubation in the dark, the optical density was read at 415 nm against a blank via a BioMate3 spectrophotometer. The total flavonoid content was determined via the calibration curve of 10 mg/mL rutin according to the methods described by Kim, et al. [ 20 ] and Zhishen, et al. [ 21 ]. Total Condensed Tannin Assay Five hundred microliters of methanolic extract at 1 mg/mL was mixed with 3 mL of 4% vanillin. To this mixture, 2 mL of methanol and 1.5 mL of fuming hydrochloric acid were added. After 15 min of incubation, the optical density was read at 500 nm via a BioMate5 spectrophotometer against a blank according to the method described by Heimler, et al. [ 22 ]. The standard used to determine the condensed tannin content was prepared under the same conditions with catechin Antioxidant activity of O. gratissimum and V. amygdalina leaves extracts Diphenyl-1-picrylhydrazyl Assay The evaluation of radical scavenging activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals in Crataegus fruit extracts was conducted spectrophotometrically. The method, which was originally introduced by Chokki, et al. [ 23 ] and further developed by Dah-Nouvlessounon, et al. [ 24 ], was utilized. The principle of the assay relies on the color transformation of the DPPH solution, which shifts from purple to yellow as the radical is neutralized by the antioxidant. One hundred microliters of methanolic hawthorn fruit extract was mixed with 1.9 mL of 410–3 mM DPPH in methanol, resulting in a total volume of 2 mL. The ability to scavenge free radicals via the DPPH radical was assessed by monitoring the decrease in absorbance at 517 nm every 2 min until the reaction reached equilibrium. Further dilution was necessary if the measured DPPH value exceeded the linear range of the standard curve [ 25 ]. The inhibition activity (I%) was then calculated as follows: $$\:I\left(\%\right)=\frac{Abs\:contol-Abs\:compound}{Abs\:control}*100$$ The extract concentration providing 50% inhibition concentration (IC50) was calculated from the graph plotting the percentage of inhibition against the extract concentration. In vitro anticoccidial activities of O. gratissimum and V. amygdalina leaves extracts Preparation of E. media and E. magna cultures Potassium dichromate (K 2 Cr 2 O 7 ): 2.5% potassium dichromate was prepared by dissolving 2.5 g of potassium dichromate in 100 ml of distilled water. This culture medium was stored and used to prepare the concentrations of the plant extracts. Preparation of sporulated oocysts: Field isolates of Eimeria magna and Eimeria media were collected from the small intestine (jejunum and ileum) of three naturally infected rabbits, confirmed by the flotation technique. These rabbits were purchased from independent local breeders. The oocysts were then washed and concentrated using the flotation method. Sporulated oocysts were stored in a 2.5% potassium dichromate solution at 4°C until experimental use. Field isolates of E. magna and E. media were maintained by periodic passage in healthy young albino New Zealand rabbits from the Laboratory of Biotechnology and Animal Improvement of University of Abomey-Calavi, Benin. In vitro oocysticidal effects of the extracts Petri dishes were used to assess in vitro antioocyst activities. Each well contained a total volume of 2 ml of each extract (2.5, 5, 10, 20, or 40 mg/ml) inoculated with an equal number of nonsporulated oocysts and incubated at 28°C. For comparison, phenol was used as the reference solution. The preparation was examined after 24 h and 48 h. The number of sporulated and nonsporulated oocysts was counted, and the percentage of sporulation was estimated by counting the number of sporulated oocysts out of a total of 100 oocysts. The percentage inhibition of sporulation was calculated as follows. $$\:Sporulation\:\left(sp\right)inhibition\:percentage\:\left(\%\right)=\frac{sp\:\%\:of\:control-sp\:\%\:of\:extraact\:}{Sp\:of\:control}x\:100$$ In vitro anti-sporozoidal effect of extracts The oocysts stored in K 2 Cr 2 O 7 were washed several times with HBSS (pH 7.2) until the K 2 Cr 2 O 7 was completely eliminated. The oocysts were then incubated in a water bath at 41°C and shaken for 60 minutes. The suspension was centrifuged at 3,000–5,000 × g for 10 min and resuspended in HBSS. The released sporozoites were washed with HBSS. The sporozoites were counted in McMaster cells. Petri dishes were used to evaluate the in vitro sporocidal activity. Each well contained a total volume of 2 ml of each extract concentration (250, 500, 750, and 1000 µg/ml) and was inoculated with an equal number of sporozoites. For comparison, amprocox was used as the reference drug. The setup was examined after 12 h and 24 h. The number of viable and nonviable sporozoites was counted, and the percentage of viability was estimated by counting the number of viable sporozoites out of a total of 100 sporozoites [ 26 ]. The percentage inhibition of viability was calculated as follows. $$\:Viability\:\:\left(Vi\right)\:inhibition\:\left(\%\right)=\frac{Vi\:\%\:of\:control-Vi\:\%\:of\:extrac}{Vi\:\%\:of\:control}x100$$ Larval cytotoxicity test of aqueous and acetonic extracts of O. gratissimum and V. amygdalina leaves Artemia salina eggs were used for the larval cytotoxicity test. The cytotoxic effect of the hydroethanol extract of the plants studied was assessed on brine shrimp larvae via a preliminary nonclinical toxicity test as described by Adoho, et al. [ 27 ]. Artemia salina larvae were obtained by incubating 10 mg of Artemia salina eggs in 1 L of seawater under continuous agitation for 48 hours. Serial dilutions of the extract were prepared from a stock solution (20 mg/mL) to obtain increasing concentrations. Each diluted solution (1 ml) was mixed with 1 ml of seawater containing 16 live larvae. A control solution, without extract, was prepared simultaneously under the same conditions. After incubation for 24 hours under continuous agitation, the number of dead larvae in each solution was determined via optical microscopy. These data were used to establish a dose‒response curve (the number of surviving larvae as a function of extract concentration). The concentrations were then log-transformed, and the median lethal concentration (LC 50 ) was calculated. Larval toxicity was interpreted on the basis of the correlation grid proposed by Ugwah-Oguejiofor, et al. [ 28 ], which associates the degree of toxicity with the LC 50 value. According to this grid: LC 50 ≥ 0.1 mg/mL ◊ Nontoxic extract 0.1 mg/mL > LC 50 ≥ 0.050 mg/mL ◊ Low toxicity 0.050 mg/mL > LC 50 ≥ 0.0 mg/mL ◊Medium toxicity LC 50 < 0.01 mg/mL ◊ High toxicity Acute oral toxicity of acetonic extracts of O. gratissimum and V. amygdalina leaves Animal materials Nine female albino Wistar rats weighing between 150 and 200 g and at least three months of age were used for the acute oral toxicity test. These nulliparous, nonpregnant female rats were supplied by the Institute of Applied Biomedical Sciences at the University of Abomey-Calavi, Benin. The rats were then randomly housed in groups of 3 in cages fitted with stainless steel lids to allow 14 days of acclimatization at the animal house of the Zootechnical Research and Livestock System Unit (URZoSE) at the National University of Agriculture. The rats had access to water and food ad libitum . The daily temperature was 23 ± 2°C, and the relative humidity was 60 ± 10%, with a 12-hour light/dark cycle. Study design The toxicity of the acetonic extracts of O. gratissimum and V. amygdalina was evaluated according to the guidelines of the Organization for Economic Cooperation and Development (OECD) No. 423 for chemical testing [ 29 ]. This method allows the determination of the dose of extract tolerated by the animal subjects. The 9 rats were divided into 3 groups of 3 rats each, thus forming groups of similar average weights. Group 1 (control), group 2 (rats receiving the acetone extract of O. gratissimum leaves), and group 3 (rats receiving the acetone extract of V. amygdalina leaves) were included. Before the extracts were administered, all the rats underwent a 12-hour fast during which they had access only to water. The individual weight of each rat was determined. The rats in groups 2 (n = 3) and 3 (n = 3) received a single dose of acetone extracts of O. gratissimum and V. amygdalina dissolved in 1 ml of distilled water by oral gavage. Group 1 (n = 3) received 1 ml of distilled water by oral gavage. The rats were observed individually for the first thirty minutes and regularly for the first 24 h and every day for 14 days to observe any signs of macroscopic toxicity (convulsions, agitation, a moribund state, severe distress, or animal mortality) and possible mortality. Animal weights were measured on days 1, 7 and 14 of the experiment. At the end of the 14-day observation period, the animals were anaesthetized with thiopental and then euthanized by cervical dislocation. The major organs—the brain, liver, spleen, heart and kidneys — were removed, weighed and macroscopically observed for lesions and abnormalities. The relative weight of each organ was calculated by relating its weight to the body weight of the corresponding animal. In addition, samples of these organs were fixed in 10% neutral buffered formalin for further histological analysis. All animal handling, including oral administration and posttreatment observations, was performed by qualified personnel specifically trained in rodent care and handling in accordance with institutional and regulatory guidelines. Biochemical and haematological analysis On day 14 of treatment, a blood sample (1 ml) was taken from each experimental animal in a dry tube and in a tube with EDTA via puncture of the retro-orbital plexus via heparinized capillary tubes. The blood in the dry tube was centrifuged at 2,500 rpm for 5 minutes at 4°C. Serum samples collected in this way were subjected to biochemical analysis via specific commercial kits for the quantification of various biomarkers: blood glucose (G O D - P AP, Biolabo. #Ref 7409), creatine kinetics (creatinine, Biolabo. #Ref 0107), urea (Uree, Biolabo. #Ref 0221), aspartate aminotransferase (AST, Biolabo. #Ref 0025), alanine aminotransferase (ALT, Biolabo. #Ref 0027), alkaline phosphatase (ALP, Biolabo. #Ref 80014) and gamma-glutamyl transferase (γ-GT, Biolabo. #Ref 81310). Readings were taken on an SP-350-BIO spectrometer, COLE-PARMER® formerly GenovaPlus from JENWAY®. The blood samples collected in EDTA-coated tubes were used to analyse various hematological parameters, including red blood cell (RBC) count, white blood cell (WBC) count, hemoglobin (Hb) concentration, packed cell volume (PCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), and blood platelet (BP) count. These parameters were measured via an automated Beckman Coulter Ac hematology analyser [ 30 ]. Histological examination The rats were sacrificed, and tissue samples from the liver and kidneys were removed and processed into paraffin blocks via labelled tissue processing cassettes. The samples were dehydrated with increasing concentrations of alcohol (70%, 80%, 90% and absolute alcohol). Further dehydration was carried out via two changes of absolute alcohol, followed by three changes of xylene. The tissues were then infiltrated and embedded in paraffin. Sections (4 µm thick) were cut from each paraffin block, mounted on microscope slides and stained with hematoxylin and eosin (H&E) as described previously Ugwah-Oguejiofor, et al. [ 28 ]. The stained sections were examined under an Olympus microscope for morphological changes, and representative images were captured. Statistical analysis The experimental data were systematically organized and analyzed using the Excel 2019 database. The data were analyzed using one-way analysis of variance (ANOVA) and presented as the mean ± standard deviation (SD) of 3 replicates. Significance levels were determined using the Waller-Duncan test, with p-values < 0.05 considered statistically significant. Statistical analysis was performed using the R software (version 4.4.2). The IC50 values of the extracts and their main compounds, which showed high anticoccidial efficacy, and the LC50 values, which showed the safety of the extracts, were evaluated by regression analysis. For graphical representation and further statistical analysis, GraphPad Prism® software (version 10.4.1) was utilized Results Phytochemical screening Several chemical groups are present in the leaves of both plants, suggesting that they share similar biological properties. Common groups include tannins (catechic and gallic), flavonoids, quinone derivatives, coumarins, reducing compounds, O-heterosides (reduced and unreduced), C-heterosides and alkaloids. There are also some notable differences, such as leucoanthocyanins, which are absent in the leaves of O. gratissimum but present in the leaves of V. amygdalina . Saponosides are absent in the leaves of O. gratissimum but are present at relatively high concentrations in the leaves of V. amygdalina (lather > 1 cm). Free anthracenics and cyanogenic derivatives are absent in the leaves of both plants. Table 1. Qualitative screening of O. gratissimum and V. amygdalina powder leaves. O. gratissimum V. amygdalina Tannins + + Catechin tannins + + Gallic tannins + + Leucoanthocyanins - + Flavonoids + + Quinone derivatives + + Saponosides - + > 1 cm Mucilages + + Coumarins + + Reducing compounds + + Free anthracenics - - O-heterosides + + Reduced genine O-heterosides + + C-heterosides + + Alkaloids + + Cyanogenic derivatives - - “+” = presence; “-’’ = absence; >1 cm represents the height of the lather created. Polyphenol and flavonoid contents of O. gratissimum and V. amygdalina powder leaves The acetone extract of V. amygdalina leaves (VaAc) had the highest total phenol content (736.27 ± 0.97 mg EAG/g ES), followed by the acetone extract of O. gratissimum leaves (OgAc), 549.85 ± 1.55 mg EAG/g ES (p˂0.05). The aqueous extracts of V. amygdalina (VaAq) and O. gratissimum leaves (OgAq) presented the lowest total phenol contents, with values of 420.35 ± 1.94 mg EAG/g ES and 374.48 ± 0.58 mg EAG/g ES, respectively (p˂0.05). The acetone extract of O. gratissimum had the highest flavonoid content (37.06 ± 0.31 mg ER/g ES), followed by the acetone extract of V. amygdalina (30.71 ± 0.68 mg ER/g ES). However, the aqueous extracts of O. gratissimum (18.89 ± 0.48 mg ER/g ES) and V. amygdalina (16.74 ± 0.19 mg ER/g ES) presented the lowest levels (p˂0.05). Condensed tannins were present in high amounts in the acetone extract of V. amygdalina (5.94 ± 0.42 mg EC/g ES), followed by the aqueous extract of V. amygdalina (3.53 ± 0.52 mg EC/g ES). The aqueous and acetone extracts of O. gratissimum presented the lowest concentrations (p˂0.05), 2.83 ± 0.13 mg EC/g ES and 1.53 ± 0.12 mg EC/g ES, respectively (Table 2). Table 2. Contents of total phenols, flavonoids and condensed tannins in O. gratissimum and V. amygdalina leaves extracts. O. gratissimum V. amygdalina Equations R 2 Pvalue Extraits OgAq OgAc VaAq VaAc Total phenols (mg EAG/g ES) 374.48±0.58a 549.85±1.55d 420.35±1.94c 736.27±0.97f y= 0.115x + 0.003 0.998 2.E-16 Flavonoïds (mg ER/g ES) 18.89±0.48b 37.06±0.31e 16.74±0.19a 30.71±0.68c y = 0.160x + 0.006 0.9993 6.36E-15 Condensed tannins (mg EC/g ES) 1.53±0.12a 2.83±0.13a 3.53±0.52b 5.94±0.42d y = 0.129x + 0.005 0.9953 9.56E-12 The values bearing the different letters a, b, c, d, and e in the same line are significantly different at the 5% level (p < 0.05) for the same metabolite. OgAq: Aqueous extracts of O. gratissimum , OgAc: Acetonic extract of O. gratissimum , VaAq: Aqueous extract of V. amygdalina , VaAc: Acetonic extract of V. amygdalin , EAG: gallic acid equivalent, ER: rutin equivalent, EC: catechin equivalent, ES: dry extract. Antioxidant activity Ascorbic acid (Acid Asc) has a relatively low effective concentration 50 (EC 50 ) value (2.802 ± 0.024), with R 2 =0.9747. This finding indicates high efficacy in producing an effect (standard reference). The acetone extract of V. amygdalina leaves (VaAc) had a slightly higher EC 50 (3.541 ± 0.112) than ascorbic acid did, with an R 2 = 0.986, suggesting that it was less effective than the standard. The acetone extract of O. gratissimum (OgAc) had an EC 50 (2.4025 ± 0.0385) even lower than that of ascorbic acid, with an R 2 = 0.9457, indicating high efficacy. The aqueous extract of V. amygdalina (VaAq) presented the highest EC 50 value (4.0305 ± 0.101), with R 2 = 0.9439, indicating that it was the least effective of the extracts tested. The aqueous extract of O. gratissimum (OgAq) presented a moderate EC 50 (3.467 ± 0.022) with R 2 = 0.9856, which was lower than that of V. amygdalina (VaAq) but higher than those of the acetonic extracts and the standard (Table 3). In summary, the acetone extract of O. gratissimum (OgAc) had the lowest EC 50 (2.4025 ± 0.0385), followed by ascorbic acid (AcidAsc), which was used as a reference. These findings suggest that this extract has a high capacity for action at relatively low concentrations. The aqueous extract of V. amygdalina (VaAq) had the highest EC 50 (4.0305 ± 0.101), indicating lower efficacy. Acetonic extracts had a lower EC 50 (better efficiency) than aqueous extracts, probably due to better extraction of bioactive compounds (Table 3). Table 3. Free radical scavenging activity via the DPPH method EC 50 CI R square Acid Asc 2.802±0.024 1.930 to 4.055 0.9747 VaAc 3.541±0.112 2.229 to 5.359 0.986 OgAc 2.4025±0.0385 1.154 to 4.315 0.9457 VaAq 4.0305±0.101 2.587 to 8.024 0.9439 OgAq 3.467±0.022 2.385 to 5.283 0.9856 In vitro oocysticidal activities of O. gratissimum and V. amygdalina leaves extracts. At low concentrations (2.5 mg/mL), the sporulation inhibition efficacy was generally low for all the extracts. The acetonioic extract of V. amygdalina leaves had the greatest activity, with inhibition percentages reaching 35.33% for E. magna at 24 h and 44.67% for E. media at 48 h. The other extracts produced significantly lower results (p˂0.05). At an intermediate concentration (5–10 mg/mL), inhibition increased in proportion to the concentration. The V. amygdalina and O. gratissimum leaves acetone extracts remained significantly more effective (p˂0.05), reaching nearly 62.67% ( E. magna , 24 h) for the V. amygdalina leaves acetone extracts at 5 mg/mL and up to 82.33% ( E. magna , 24 h) at 10 mg/mL for the O. gratissimum leaves acetone extract. At high concentrations (20–40 mg/mL), inhibition was maximal, particularly for the acetone extract of V. amygdalina leaves at 40 mg/mL. The percentage of inhibition reached 91% at 24 h and 100% at 48 h for E. magna with the acetone extract of V. amygdalina . Similar results were recorded for E. media (Table 4). The aqueous extracts of the two plants were less effective overall than the acetonic extracts were. The positive control (5%) shows almost complete inhibition (97.67% to 100%) for both species, confirming that the extracts tested do not exceed this efficacy. The inhibition of the negative control (K₂Cr₂O₇ alone or with DMSO) was very low (6–9%), indicating that the effect observed with the extracts was due mainly to the bioactive compounds present in the plants. Acetone extracts, particularly from V. amygdalina leaves, were the most effective against E. magna and E. media at all concentrations tested. These values almost reached the values of the positive control at high concentrations (40 mg/mL). The aqueous extracts showed moderate efficacy, with the aqueous extract of V. amygdalina leaves being slightly superior to the aqueous extract of O. gratissimum leaves. The inhibition of both Eimeria species increased with increasing extract concentration. At high concentrations (20–40 mg/mL), the effect was close to or equal to that of the positive control (Table 4). Table 4. Sporulation percentages of O. gratissimum and V. amygdalina in different d’ E. magna and E. media . Concentration (mg/ml) Extracts Incubation times for Eimeria strains E. magna E. media 24 hours 48 hours 24 hours 48 hours 2.5 OgAq 13.67±4.73 a 28.33±3.21 a 12.00±3.00 a 25.67±8.96 a VaAq 18.00±5.29 ab 36.33±4.16 abc 14.33±2.89 a 37.33±6.43 ab OgAc 23.67±1.53 b 30.00±4.00 a 19.33±1.53 b 34.67±9.24 ab VaAc 35.33±4.51 c 43.33±3.79 c 22.33±0.58 b 44.67±4.51 b 5 OgAq 33.33±4.16 a 58.00±2.65 a 33.00±4.00 a 54.00±9.54 a VaAq 46.00±5.29 b 66.00±5.57 ab 40.00±3.61 a 67.33±7.02 ab OgAc 49.67±3.51 b 60.33±4.73 a 39.33±2.89 a 63.33±11.55 ab VaAc 62.67±10.21 c 71.00±2.65 b 51.00±1.73 b 73.33±3.79 b 10 OgAq 51.00±1.00 a 78.33±10.21 ab 49.00±6.00 a 75.33±11.85 a VaAq 64.33±6.81 b 85.33±4.04 ab 64.00±1.00 bc 86.33±5.69 ab OgAc 69.00±6.93 b 81.33±4.73 ab 60.00±2.65 b 85.00±8.72 ab VaAc 82.33±10.02 c 90.00±2.00 b 70.33±0.58 bc 91.67±5.13 b 20 OgAq 55.00±5.00 a 83.00±8.89 a 54.00±7.00 a 80.67±9.61 a VaAq 69.00±4.58 b 90.33±3.21 ab 68.33±1.53 bc 92.33±1.53 b OgAc 73.67±7.77 b 85.33±6.66 ab 63.67±3.51 ab 87.33±9.29 ab VaAc 88.00±7.21 c 94.67±1.53 c 78.00±6.08 c 96.67±4.16 b 40 OgAq 62.00±6.56 a 87.00±8.72 a 57.33±6.51 a 84.00±8.89 a VaAq 71.67±3.79 ab 93.00±4.58 ab 73.00±1.00 b 95.00±2.65 b OgAc 78.00±8.19 b 91.00±6.24 b 68.33±3.51 b 90.67±7.77 ab VaAc 91.00±2.65 c 100.00±0.00 b 84.33±5.13 c 100.00±0.00 b Nagative controle K 2 Cr 2 O 7 +DMSO 6±1.73 6.33±1.53 7±2.65 5.66±152 K 2 Cr 2 O 7 9±2 9.67±0.58 9±1 9.00±2.00 Positivve controle 5% 97.67±2.52 100.00±0.00 97.00±3.00 100.00±0.00 The values bearing the different letters a, b, c, d, and e in the same column are significantly different at the 5% level (p < 0.05) for the same metabolite. OgAq: Aqueous extracts of O. gratissimum , OgAc: Acetonic extract of O. gratissimum , VaAq: Aqueous extract of V. amygdalina , VaAq: Acetonic extract of V. amygdalin , In vitro anti-sporozoidal activities of O. gratissimum and V. amygdalina leaves extracts The efficacy of the extracts increased significantly (p˂0.05) with concentration, regardless of the type of extract or Eimeria species. At a dose of 125 µg/ml, the percentages of viability inhibition of E. magna and E. media strains were relatively low, with greater efficacy observed for the acetone extract of V. amygdalina (29.00 ± 2.65% on E. magna at 12 h and 47.33 ± 6.03% at 24 h). At a dose of 250 µg/ml, a clear increase in the percentage of inhibition was observed for all extracts, with the acetone extract of V. amygdalina retaining its superiority (47.00 ± 2.00% at 12 h and 65.67 ± 1.15% at 24 h over E. magna ). At a dose of 500 µg/ml, the efficacy of the aqueous extracts of the two plants was similar to that of the acetone extracts, although the percentage inhibition of the acetone extract of V. amygdalina remained greater (91.00 ± 1.73% in E. media at 24 h). At a dose of 1000 µg/ml, the extracts reached efficacy levels close to or equal to 100%, particularly for the acetone extracts of V. amygdalina and O. gratissimum . The effect was dose dependent (Table 5). E. magna responded slightly more sensitively to the extracts, particularly the acetone extracts of V. amygdalina leaves, at 24 hours. At 1000 µg/ml, the acetone extracts of V. amygdalina leaves achieved 100% inhibition, which was identical to that of the positive control. E. media showed a similar trend to that of E. magna , but the inhibition values were often slightly lower (Table 5). Table 5. Percent inhibition of viability of E. magna and E. media strains in contact with O. gratissimum and V. amygdalin extracts Concentration µg/ml Extracts Incubation times for Eimeria strains E. magna E. media 12 hours 24 hours 12 hours 24 hours 125 OgAq 5.67±1.15 c 14.67±3.79 a 6.33±1.53 c 13.33±3.21 a VaAq 11.67±2.52 bc 23.67±6.51 a 13.33±2.08 b 24.33±8.14 ab OgAc 16.67±5.69 ba 33.33±7.02 b 16.67±2.52 b 32.00±10.39 bc VaAc 29.00±2.65 a 47.33±6.03 c 31.00±4.58 d 46.00±1.73 d 250 OgAq 23.33±7.02 a 39.00±2.65 e 23.00±3.61 a 38.67±8.62 a VaAq 36.00±5.57 b 51.33±11.06 cd 32.67±3.21 ab 51.33±6.43 b OgAc 35.67±4.04 b 44.33±0.58 de 35.00±6.56 abc 43.67±1.15 ab VaAc 47.00±2.00 c 65.67±1.15 a 46.33±4.93 c 67.00±3.00 c 500 OgAq 50.67±4.62 d 58.67±4.73 a 49.67±6.66 b 57.67±7.37 c VaAq 63.00±6.24 bc 88.33±11.72 c 63.67±8.50 a 85.00±5.29 ab OgAc 52.33±8.33 cd 72.67±11.50 b 48.33±9.45 b 73.00±8.19 b VaAc 77.00±5.20 a 89.67±0.58 c 78.00±10.54 a 91.00±1.73 a 1000 OgAq 65.00±4.58 a 77.67±2.52 a 66.67±9.07 c 78.67±6.66 c VaAq 80.67±7.64 bc 93.67±1.53 c 83.67±9.50 ab 97.00±3.00 ab OgAc 74.67±7.57 ab 88.67±4.16 b 73.00±8.89 bc 92.67±5.51 b VaAc 91.67±2.08 c 100.00±0.00 d 90.67±11.15 a 100.00±0.00 a Negative control DMSO 0.00±0.00 0.00±0.00 0.00±0.00 0.00±0.00 HBSS 0.00±0.00 0.00±0.00 0.00±0.00 0.00±0.00 Positive control 50 µg/ml 88.67±3.51 100.00±0.00 89.00±1.73 100.00±0.00 OgAq: Aqueous extract of O. gratissimum , OgAc: Acetonic extract of O. gratissimum , VaAq: Aqueous extract of V. amygdalina , VaAc: Acetonic extract of V. amygdalina , DMSO: Dimethyl sulfoxide, HBSS: Hank's buffered salt solution. The results are presented as the means and standard deviations of triplicate in vitro tests after 12 h and 24 h of incubation at room temperature. For the same column and concentration, values with the same superscript letter are not significantly different at p ≥ 0.05 (Duncan's test) The curves show an inverse relationship between the compound concentration and oocyst sporulation. As the concentration increased, sporulation decreased, which is typical of antiparasitic substances. The IC₅₀ values indicate the concentrations required to inhibit 50% of sporulation. The IC 50 values of the extracts from the two plants were therefore examined in more detail via regression analysis (Figure 1). Aqueous and acetone extracts of O. gratissimum leaves inhibited the sporulation of 50% of E. media coccidia oocysts at a concentration of 3.466 mg/ml (IC₅₀ = 3. 466 mg/ml) and 4.413 mg/ml (IC₅₀ = 4.413 mg/ml), with coefficients of determination R² = 0.9749 and R² = 0.9873, respectively (Figure 1A and 1B). In contrast, aqueous and acetone extracts of V. amygdalina leaves inhibited the sporulation of 50% of E. media oocysts at a concentration of 2. 920 mg/ml (IC₅₀ = 2.920 mg/ml) and 3.415 mg/ml (IC₅₀ = 3.415 mg/ml), with coefficients of determination R² = 0.9873 and R² = 0.9817, respectively (Figure 1C and 1D). The logarithmic equation had the highest coefficient of determination (R 2 ) (Figure 1C and 1D), and the IC 50 values were 2.92 and 3.41 mg/ml for the acetone and aqueous extracts of V. amygdalina leaves, respectively (Figure 1). This figure illustrates the inhibitory effects of different leaves extracts on the sporulation of E. magna oocysts. The IC 50 (median inhibitory concentration) and R 2 values were used to assess the power and goodness of fit of the dose‒response curves. The high R 2 values (>0.97) for all the graphs confirm a strong correlation between the extract concentrations and the reduction in the percentage of sporulated oocysts (Figure 2). The results show that extracts of V. amygdalina (Figure 2 C and D) are more effective than those of O. gratissimum (Figure 2A and D) in inhibiting the sporulation of E. magna oocysts. The lower IC 50 values for V. amygdalina (3.097 mg/mL for the acetone extract and 3.408 mg/mL for the aqueous extract) suggest better anticoccidial activity, regardless of the extraction method. In comparison, O. gratissimum had slightly higher IC 50 values (4.066 mg/mL for the acetone extract and 4.091 mg/mL for the aqueous extract), indicating moderate efficacy (Figure 2). Larval Toxicity of Extracts. The larval toxicity results revealed that the plant extracts studied were not toxic to Artemia salina larvae, with notable differences between the aqueous and acetonic extracts. The aqueous extract of V. amygdalina (Figure 3A), with an LC 50 of 0.269 mg/mL (R² = 0.93), was classified as a nontoxic extract. Similarly, the aqueous extract of O. gratissimum (Figure 3C) had an LC 50 of 0.168 mg/mL (R² = 0.95), which also placed it in the category of nontoxic extracts, indicating better accuracy in model fitting. In contrast, the acetone extract of V. amygdalina has an LC 50 of 0.978 mg/mL (R² = 0.92), which also classifies it as nontoxic (Figure 3B), whereas the acetone extract of O. gratissimum , with an LC 50 of 0.137 mg/mL, is also considered nontoxic, although the R² of 0.76 suggests some variability in the model fit (Figure 3D). In summary, both plants produced mainly nontoxic extracts, with LC 50 variations influenced by the type of solvent used (Figure 3). Acute oral toxicity Mortality and clinical signs No mortality was observed in the rats treated with the acetone extracts of the three plants during the acute toxicity tests. No signs or abnormalities were recorded on the skin, coat, eyes or mucous membranes. There was also a complete absence of hyperactivity, eye twitches, catalepsy, convulsions, tremors and catatonia following treatment with the acetonic fractions of the plants. Administration of the acetonic extracts produced no signs of erythema or edema until after 48 hours of observation. Food consumption and body weight of the rats There were no significant differences in food consumption between the test group and the control group. The weights of the subjects in the treated batches differed significantly (p < 0.05) from those in the control batches at D7 and D14 (Figure 4). Hematological parameters of the rats The results in the table show several significant differences in haematological parameters among the three groups. White blood cell (WBC) counts were significantly lower in the OgAc group (12.7 ± 0.9) than in the control group (13.98 ± 0.38) and the VaAc group (13.25 ± 0.05), with a p value of 0.008. Compared with the control group (7.35 ± 0.55) and the OgAc group (7.755 ± 0.015), the VaAc group presented a significant reduction in red blood cell count (6.63 ± 0.1). Haemoglobin (Hb) levels were significantly lower in the VaAc group (13.35 ± 0.05) than in the control (14.425 ± 0.425) and OgAc (14.5 ± 0) groups. Red blood cell (RBC) volume was significantly greater (p=0.017) in the OgAc (48.4 ± 0.2) and VaAc (47.2 ± 1.8) groups than in the control group (43.35 ± 0.35). The mean corpuscular volume (MCV) was significantly greater (p ˂0.001) in the VaAc group (71.15 ± 1.65) than in the control group (62.55 ± 1.05) and the OgAc group (62.4 ± 0.1). The mean amount of haemoglobin per red blood cell (MCH) was significantly lower (p ˂0.001) in the OgAc group (18.7 ± 0) than in the Control (23 ± 1) and VaAc (20.15 ± 0.35) groups. The hemoglobin concentration per red blood cell volume (MCHC) was significantly lower (p ˂0.001) in the OgAc (29.95 ± 0.15) and VaAc (28.3 ± 1.2) groups than in the control group (34.85 ± 1.15). Finally, the platelet count (PCR) was significantly lower (p ˂0.001) in the OgAc group (578 ± 13) than in the control group (745.5 ± 5.5) and the VaAc group (793.5 ± 3.5). These results suggest that treatments with extracts of O. gratissimum (OgAc) and V. amygdalina (VaAc) modify several blood parameters, particularly white blood cells, red blood cells, haemoglobin and platelets (Table 6). Table 6. Hematological parameters of the rats. Control OgAc VaAc P value WBC (10 3 /mm 3 ) 13.98±0.38 a 12.7±0.9 b 13.25 ±0.05 a 0.008 RBC (10 6 /mm 3 ) 7.35±0.55 7.755±0.015 6.63±0.1 0.031 Hb (g/dL) 14.425±0.425 14.5±0 13.35±0.05 0.029 PVC (%) 43.35±0.35 48.4±0.2 47.2±1.8 0.017 MCV (fL) 62.55±1.05 62.4±0.1 71.15 ±1.65 <0.001 MCH (pg) 23±1 18.7±0 20.15±0.35 <0.001 MCHC (g/dL) 34.85±1.15 29.95±0.15 28.3±1.2 <0.001 BP (10 3 /mm 3 ) 745.5±5.5 578±13 793.5±3.5 <0.001 RBC=Red blood cells. PCV=Packed cell volume. MCV=mean corpuscular volume. MCH=mean corpuscular haemoglobin. MCHC=mean corpuscular haemoglobin concentration. WBC=white blood cell. OgAc: Acetonic extract of O. gratissimum . VaAc: Acetonic extract of V. amygdalina Biochemical parameters of the rats The results show that there was no significant difference in blood glucose levels between the three groups, with similar values for the Control (0.72 ± 0.05), OgAc (0.67 ± 0.03) and VaAc (0.66 ± 0.01) groups. However, the OgAc group had a significantly higher urea concentration (0.33 ± 0.08) compared with the Control group (0.20 ± 0.01), while the VaAc group showed no difference compared with the Control group. No significant difference was observed for creatinine between groups. ALT levels were significantly higher in the OgAc (92.08 ± 1.01) and VaAc (98.66 ± 0.74) groups than in the control group (66.58 ± 2.02). The VaAc group also had a significantly higher AST concentration (148.38 ± 0.96) than the control group (124.33 ± 5.09). Although there was a significant difference for ALP, the values were not radically different between the groups, with the OgAc group having a higher value (477.43 ± 31.75). Finally, the VaAc group showed a higher concentration of γ‐GT (4.52 ± 0.67) than the control group (3.85 ± 0.08), with a significant difference (p = 0.006), (Table 7). Table 7. Biochemical parameters of the rats Control OgAc VaAc pvalue Glycemia (mg/dL) 0.72±0.05 0.67±0.03 0.66±0.01 0.143 Urea (g/L) 0.20±0.01 0.33±0.08* 0.24±0.04 0.007 Creatinine (g/L) 10.12±1.85 11.51±0.48 12.46±2.48 0.304 ALT (U/L) 66.58±2.02 92.08±1.01** 98.66±0.74** 0.000 AST (U/L) 124.33±5.09 121.13±2.13 148.38±0.96** 0.000 ALP (U/L) 446.83±2.35 477.43±31.75 419.82±28.20 0.001 γ‐GT (U/L) 3.85±0.08 3.47±0.42 4.52±0.67 0.006 * = p < 0.05; ** = p < 0.01. AST=aspartate amino transaminase. ALT=alanine amino transaminase. ALP=alkaline phosphate, γ‐GT= Gamma‐Glutamyl Transferase Relative organ weights and necroscopy of the rats Relative organ weight was calculated via the formula (organ weight/body weight) × 100%. Relative vital organ weights and microscopic changes were studied in comparison with those in the control group, and no significant changes (p>0.05) were observed. Histopathological examination of different organs of rats Histological analysis of livers from rats fed acetone extracts of O. gratissimum leaves revealed a generally preserved hepatic architecture, with well-defined hepatocytes but diffuse cytoplasmic vacuolation, suggesting potential lipid accumulation or mild hydropic degeneration. The hepatic sinusoids appear slightly dilated in some areas, with no major disruption to the radial organization of the hepatocytes. The portal areas and central veins showed no evidence of inflammation, fibrosis or abnormal cellular infiltration, indicating the absence of severe liver toxicity. Although the uneven quality of the staining may have introduced artefacts, the observations suggest a slight adaptive metabolic response to the extract (Figure 5A). Histological analysis of liver sections from rats fed an acetone extract of V. amygdalina leaves revealed a generally preserved lobular architecture, with hepatocytes organized radially around the central veins. Hepatic cells showed diffuse cytoplasmic vacuolation, suggesting lipid accumulation or hydropic degeneration, possibly in the metabolic response to the extract. The hepatic sinusoids appeared mildly dilated, whereas the portal areas and central veins showed no evidence of inflammatory infiltration or fibrosis. No obvious hepatocellular damage, such as necrosis or apoptosis, was detected. A few artefacts related to section preparation and staining were noted, but these did not alter the overall interpretation of the observations. These results suggest reasonable hepatic tolerance to the extract (Figure 5B), as do the liver structures of the control rats (Figure 5C). Histological analysis of kidney sections from rats fed an acetone extract of O. gratissimum leaves revealed well-preserved overall nephron architecture, including glomeruli and renal tubules. The glomeruli generally appear intact, with normal vascularisation and no obvious signs of sclerosis or narrowing. Bowman's spaces are homogeneous and show no signs of edema or inflammation. However, subtle alterations were observed in the proximal tubules, with evidence of diffuse cytoplasmic vacuolation, probably related to reversible degeneration or a metabolic response to the extract compounds. The distal tubules appeared normal, with no signs of atrophy or obstruction. No obvious inflammatory infiltrates or necrosis were identified in the renal interstices. Minor artefacts attributed to sample preparation and staining were noted but had no impact on the interpretation of the results. These observations suggest satisfactory renal tolerance to the extract (Figure 5D). Histological observation of renal sections from rats fed acetone extract from the leaves of V. amygdalina revealed an intact overall organization of the glomeruli and renal tubules. The glomeruli appear well defined, with an intact capillary structure, indicating unimpaired glomerular filtration. The proximal and distal tubules have slightly granulated cytoplasm, probably due to normal metabolic activity or a slight adaptive response to the extract. Some areas presented with cytoplasmic vacuoles and slight disorganization, which were interpreted as artefacts of fixation or sectioning. The absence of necrosis, significant inflammatory infiltration or alterations to the basement membrane suggested that the V. amygdalin extract did not cause apparent renal toxicity at the administered dose (Figure 5E). The structures of these kidneys are similar to those of control rat kidneys (Figure 5F). Discussion The results obtained for O. gratissimum and V. amygdalina show that these plants are rich in bioactive compounds such as tannins, flavonoids, saponosides, mucilages, alkaloids and heterosides, which are of great interest for animal production and health [ 31 ]. Tannins and flavonoids are well known for their antioxidant and anti-inflammatory properties and play key roles in reducing oxidative stress and managing inflammatory diseases in animals. This activity could be particularly beneficial in animal production systems where animals are exposed to high levels of stress, which can affect their immunity and productivity. For example, flavonoid-rich extracts could be used to prevent infectious diseases by reducing the bacterial load and modulating immune responses [ 32 ]. Saponosides, which are present only in V. amygdalina , are also of major interest in animal production because of their antimicrobial and immunomodulatory properties. Their ability to improve resistance to bacterial and parasitic infections could be used to reduce the use of antibiotics in livestock farming, thereby promoting a more sustainable and ethical approach to animal production. Moreover, mucilage can be used to improve digestive health. The results show that acetone extracts of O. gratissimum and V. amygdalina have significantly higher concentrations of bioactive compounds, particularly total phenols, flavonoids and condensed tannins, than aqueous extracts do. V. amygdalina , in particular, contains relatively high levels of phenols (736.27 mg EAG/g ES) and condensed tannins (5.94 mg EC/g ES) in its acetone extract, suggesting greater antioxidant and antimicrobial potential [ 5 ]. Similarly, flavonoids, mainly the acetone extract of O. gratissimum (37.06 mg ER/g ES), are known for their anti-inflammatory and cardioprotective effects, which enhance intestinal and immune health [ 33 , 34 ]. These results indicate that acetone extracts from both plants could play a key role in the management of oxidative stress and infections in animal production. These bioactive compounds, particularly acetonic extracts, have significant potential for animal health, offering natural alternatives to antibiotics and other chemicals. Their incorporation into animal diets could improve resistance to infection, promote digestive health and support the immune system [ 35 ]. In addition, the antimicrobial effects of tannins and flavonoids suggest that they could be used to treat digestive disorders and reduce infectious diseases in animal production systems. The study compared the EC50 values and confidence intervals of antioxidant activity for different extracts of V. amygdalina (Va) and O. gratissimum (Og). The acetone extract of O. gratissimum (Og Ac) exhibited the lowest EC50 value (2.40), indicating higher efficacy at a lower concentration compared to other extracts. The confidence interval for OgAc (1.15–4.31) was narrow, reflecting good precision in its efficacy estimation. In contrast, the aqueous extract of V. amygdalina (VaAq) had a higher EC 50 value (4.03), indicating the need for a higher concentration for a similar effect. Its wider confidence interval (2.59–8.02) suggests more variability in its efficacy. These findings highlight important differences that could impact animal health and productivity management [ 5 ]. The acetone extract of V. amygdalina (VaAc) has a moderate EC 50 value (3.54) and an R 2 of 0.986, indicating that this extract has a strong ability to explain the variation in biological effects as a function of concentration. In comparison, O. gratissimum has a slightly lower R 2 for its acetone extract (0.9457), suggesting a less precise relationship between the concentration and the observed effect; however, this difference is relatively small and could be considered acceptable for practical applications [ 36 – 38 ]. The acetonic extracts of O. gratissimum and V. amygdalina , with low EC50 values (2.40 and 3.54 respectively), could serve as effective natural alternatives for managing oxidative stress and infectious diseases in farm animals. Their low EC50 values indicate that these extracts are potent at low concentrations, which may reduce costs and minimize the risk of side effects. In intensive breeding systems, incorporating these extracts could enhance animals' resistance to oxidative stress, reduce infectious diseases, and support growth [ 39 , 40 ]. The use of O. gratissimum and V. amygdalina extracts could reduce the reliance on antiparasitics, particularly for preventing digestive and respiratory infections, by acting as natural antimicrobial agents. This would support sustainable animal husbandry while enhancing animal health and productivity. The in vitro results showed significant inhibition of sporulation of E. magna and E. media oocysts, with acetone extracts being more effective than aqueous extracts due to their higher content of lipophilic compounds such as flavonoids, condensed tannins, and alkaloids. This observation is consistent with the work of Dagnino, et al. [ 41 ] and Okoye, et al. [ 42 ], who reported that fat-soluble bioactive compounds extracted from medicinal plants often exhibit better antiparasitic activity in vitro . At high concentrations (40 mg/ml), the acetone extract of V. amygdalina achieved 100% inhibition of oocyst sporulation after 48 hours, rivalling the efficacy of commercial coccidiostats. These in vitro results highlight the potential of O. gratissimum and V. amygdalin extracts as natural alternatives for the management of coccidiosis in cattle. In vitro tests provide a valuable first indication, but they do not consider the complex factors present in the body, such as bioavailability, metabolism of bioactive compounds and potential interactions with the intestinal microflora [ 43 , 44 ]. The progressive increase in efficacy with concentration and incubation time reflects a dose‒dependent relationship, which is often observed in vitro pharmacological tests. The results also confirmed that aqueous extracts are less effective than acetonic extracts are, probably due to the low solubility of lipophilic compounds in water. These observations are consistent with previous work, such as that of [ 45 ], which demonstrated that organic solvent-based extracts generally exhibit better antiparasitic activity against coccidial strains. The efficacy of extracts may also be attributed to their ability to disrupt the cell membrane of oocysts or interfere with their energy metabolism, as suggested by studies on the properties of tannins and flavonoids against protozoa [ 46 ]. V. amygdalina and O. gratissimum extracts have antiparasitic effects on their bioactive compounds. Flavonoids and tannins, which are present in large quantities, are known to alter the membranes of oocysts and inhibit enzymes essential for their sporulation. Alkaloids, on the other hand, can disrupt protozoan replication mechanisms by targeting their energy metabolism. These mechanisms of action explain the high inhibition observed at increasing concentrations. The differences in efficacy between the plant species studied could be attributed to the variability in the chemical composition of the extracts, as noted by Kebede, et al. [ 47 ], who reported that pest control properties vary depending on the relative concentrations of flavonoids and tannins in the plant extracts. In addition, the acetone extract of V. amygdalina , which has shown the best efficacy, seems to contain particularly potent bioactive compounds or is better extracted in organic solvents. Extracts of O. gratissimum and V. amygdalina dose-dependently inhibited the viability of oocysts in E. magna and E. media , with a maximum performance for acetone extracts, particularly V. amygdalina leaves acetone extract (VaAc). At 1000 µg/ml, after 24 hours, VaAc achieved 100% inhibition, matching the positive control, whereas the aqueous extracts showed less, but significant, efficacy. These results indicate the importance of lipophilic metabolites, such as flavonoids and alkaloids, which are better extracted by acetone, for antiparasitic activity [ 48 – 50 ]. The phenolic and tannic compounds present in these extracts could interfere with oocyst membranes, leading to osmotic imbalances and metabolic inhibition [ 51 ]. Alkaloids, in particular, may target enzymes essential for oocyst development, a hypothesis supported by similar studies on other plant extracts [ 49 , 52 , 53 ]. These data reinforce interest in the use of plant extracts as natural coccidiostats in poultry and rabbit farming, where coccidiosis remains a major cause of economic losses [ 54 – 56 ]. The increased efficacy of acetone extracts compared to aqueous extracts can be attributed to their ability to extract more fat-soluble active compounds, such as apolar flavonoids and lignans, which are less effectively extracted in aqueous solutions. This emphasizes the importance of solvent selection to optimize the antiparasitic activity of plant extracts. These findings are especially relevant in rabbit production, where coccidiosis poses significant health and economic challenges. The inhibition of E. media oocyst sporulation suggests that these extracts could help limit the environmental spread of infectious oocysts, reducing infectious pressure on farms and potentially replacing chemical coccidiostats, addressing concerns about drug resistance and residues. The toxicity tests on Artemia salina larvae showed no toxicity, with LC50 values well above the toxic threshold, indicating that both aqueous and acetonic extracts of V. amygdalina and O. gratissimum are safe for use. These results are consistent with those of previous studies that have shown that extracts of these plants can have little or no biological effects on certain larval species, depending on the concentration used [ 30 , 57 , 58 ]. The differences in the LC 50 values observed between the aqueous and acetonic extracts can be explained by the nature of the compounds extracted by each solvent. The aqueous extracts of V. amygdalina and O. gratissimum contain mainly polyphenols and flavonoids, which are known for their antioxidant effects and low toxicity [ 59 , 60 ]. In contrast, acetone extracts, which are rich in lipophilic compounds such as terpenes and alkaloids, could theoretically exhibit greater biological activity, but here, they are also classified as nontoxic, suggesting that the bioavailability of secondary metabolites in these extracts is not high enough to result in significant toxicity to Artemia salina larvae [ 61 , 62 ]. Although the extracts were classified as nontoxic in this study, these results cannot be generalized to other species or long-term applications. Plant extracts can have toxic effects at relatively high concentrations or after prolonged exposure. Furthermore, these extracts have shown pharmacological potential in other contexts, notably in the treatment of human diseases, but their ecological impact must be taken into account before any application in the natural environment [ 63 , 64 ]. In summary, these results indicate that aqueous and acetonic extracts of V. amygdalina and O. gratissimum have little or no toxicity to Artemia salina larvae. The results of histological analyses of liver and kidney tissues from rats fed acetone extracts of O. gratissimum and V. amygdalina leaves provide essential information on the potential metabolic and toxicological effects of these phytochemicals. Both extracts showed a general preservation of histoarchitecture in liver and kidney tissues, suggesting their relative safety at the doses administered. The diffuse cytoplasmic vacuolation observed in hepatocytes from both experimental groups indicates potential lipid accumulation or mild hydropic degeneration. These changes may reflect adaptive responses to the bioactive constituents present in O. gratissimum and V. amygdalina . Similar observations have been reported in studies evaluating the hepatoprotective and hepatomodulatory properties of phytochemicals, suggesting that mild vacuolization may represent a transient metabolic adjustment to increased lipid metabolism or oxidative stress [ 65 – 68 ]. Importantly, the absence of fibrosis, necrosis or inflammatory infiltration in portal and central veins supports the idea that these extracts do not induce overt liver toxicity under the conditions studied. This finding is consistent with previous findings indicating that O. gratissimum possesses antioxidant and hepatoprotective properties, mainly attributed to its high polyphenol and flavonoid contents [ 69 ]. Renal histological analysis revealed preserved glomerular and tubular architecture, with no evidence of necrosis, inflammation or basement membrane rupture in either group. Subtle cytoplasmic vacuolation in proximal tubules may suggest reversible metabolic adjustments or mild degenerative changes related to biotransformation of plant extract constituents. These observations are consistent with the literature, which reports mild renal changes in response to bioactive compounds without significant functional alterations [ 70 – 72 ]. The absence of glomerular sclerosis or interstitial fibrosis further supports the renal safety of these extracts at the doses tested. Notably, V. amygdalina has been reported to exert nephroprotective effects in models of oxidative stress, probably due to its richness in saponins, tannins and flavonoids [ 73 – 75 ]. Although the two extracts produced similar histological results, slight differences in vacuolar changes and sinusoidal dilatation between the groups suggest variations in the bioavailability and metabolic processing of their respective phytochemical profiles. These differences could stem from the distinct chemical compositions of O. gratissimum and V. amygdalina , which include eugenol, terpenes and flavonoids for the former and sesquiterpene lactones and saponins for the latter [ 76 ]. Limits However, these observations remain limited to in vitro conditions, where complex host–environment interactions are absent. It would be crucial to confirm these effects in in vivo assays while assessing the bioavailability, safety and impact of extracts on zootechnical performance. In addition, thorough characterization of active compounds and optimization of formulations are needed to maximize efficacy and minimize the required doses. Conclusion This study evaluated the antioxidant and anticoccidial properties of aqueous and acetonic extracts of V. amygdalina and O. gratissimum for their potential to manage coccidial infections in rabbits. Antioxidant analyses revealed significant activity attributed to the presence of bioactive compounds such as flavonoids and phenols, which were particularly concentrated in the acetonic extracts. The acetone extracts also markedly inhibited the sporulation of E. magna and E. media oocysts, suggesting greater efficacy than aqueous extracts. In vitro and in vivo toxicity tests confirmed the relative safety of the extracts, with no evidence of larval toxicity or acute oral toxicity. Histological examination of the target organs revealed no toxicity at a dose of 2000 mg/kg. Among the plants studied, V. amygdalina stood out for its superior anticoccidial properties. These results highlight the potential of these plants as natural and sustainable alternatives to conventional treatments for coccidiosis while offering additional benefits linked to their antioxidant properties. Abbreviations % Percentage µg/ml Micrograms per milliliter AcidAsc Ascorbic Acid ALP alkaline phosphatase ALT Alanine aminotransaminase AST Aspartate Aminotransaminase BP Blood Patelets CI Confidence Interval DMSO Dimethyl sulfoxide DPPH 2,2-diphenyl-1-picrylhydrazyl E.magna Eimeria magna E.media Eimeria media EAG Gallic Acid Equivalent EC Catechin Equivalent EC 50 Half maximal effective concentration ER Rutin Equivalent ES Dry Extract FCR Folin-Ciocalteu reagent fL femtoliter g/dL Grams per deciliter g/L Grams per liter h Hour HBSS Hank's buffered salt solution IC 50 half-maximal inhibitory concentration K2Cr2O 7 Potassium dichromate LaBAA Laboratory of Biotechnology and Animal Improvement LC 50 Lethal concentration 50% MCH Mean Corpuscular Haemoglobin MCHC Mean Corpuscular Haemoglobin Concentration MCV Mean Corpuscular Volume mg/dL Milligrams per deciliter mg/ml Milligram per millilitre mm 3 Cubic millimetres Na 2 CO 3 Sodium carbonate O. gratissimum Ocimum gratissimum OgAc Acetonic extract of Ocimum gratissimum OgAq Aqueous extract of Ocimum gratissimum PCV Packed Cell Volume pg Picograms R 2 Rsquare RBC Red blood cells U/L Units per liter V.amygdalina Vernonia amygdalina VaAc Acetonic extract of Vernonia amygdalina VaAq Aqueous extract of Vernonia amygdalina WBC White Blood Cells γ‐GT Gamma‐Glutamyl Transferase Declarations Ethics approval and consent to participate The study was approved by the Research and Ethics Committee of the National Agricultural University of Benin (N° 143-2018/President-CER/SA). Throughout the study, measures were taken to ensure that animal welfare was respected during experimentation. The experiments were carried out by trained and certified researchers in compliance with current regulations on the use of animals for scientific purposes (European Directive 2010/63/EU/IACUC). Consent for publication Not applicable. Availability of data and materials The raw data for the current study are available from the corresponding author upon reasonable request. Competing Interests The authors declare that no competing interests exist. Funding This paper received no funding. Authors' contributions B.K. Conceptualization, Formal analysis, Investigation, Methodology, Software, Writing – original draft, Writing – review & editing; C.C.D. Investigation, Methodology, Writing – review & editing; L.D. Investigation, Resources , Writing – review & editing; R.T. Investigation, Writing – review & editing; C.H. Investigation, Resources , Writing – review & editing; F.L.M.A. D. Writing – review & editing; G.G.A. Investigation, Writing – review & editing; E.A. Conceptualization, Resources, Supervision, Validation, Writing – original draft; Writing – review & editing; S-y.D.A. Conceptualization, Funding acquisition, Project administration, Resources, Supervision, Validation, Visualization, Writing – original draft; Writing – review & editing; S.H.A-H. Writing – review & editing; L.B.M. Resources, Supervision, Validation, Visualization, Writing – original draft; Writing – review & editing; P.A.O. Project administration Conceptualization, Methodology, Project administration, Funding acquisition, Resources, Software, Supervision, Validation, Visualization, Writing – original draft; Writing – review & editing. Acknowledgements We would like to thank the entire team of the Laboratory of Biotechnology and Animal Improvement (LaBAA) of the Faculty of Agronomic Sciences of the University of Abomey-Calavi. 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Azando","email":"","orcid":"","institution":"Université de Parakou","correspondingAuthor":false,"prefix":"","firstName":"Erick","middleName":"V.B.","lastName":"Azando","suffix":""},{"id":431558773,"identity":"c0f280c5-3977-48d8-9319-8e9dd9716261","order_by":9,"name":"Sanni-yo Doko Allou","email":"","orcid":"","institution":"Université d'Agriculture de Kétou","correspondingAuthor":false,"prefix":"","firstName":"Sanni-yo","middleName":"Doko","lastName":"Allou","suffix":""},{"id":431558774,"identity":"78729350-ac3e-4739-9995-4dc3f59fd75f","order_by":10,"name":"Sylvie H. Adoté-Hounzangbé","email":"","orcid":"","institution":"Université d'Abomey-Calavi","correspondingAuthor":false,"prefix":"","firstName":"Sylvie","middleName":"H.","lastName":"Adoté-Hounzangbé","suffix":""},{"id":431558775,"identity":"0ce9775c-295c-4fb6-b7be-31ba37e53b61","order_by":11,"name":"Lamine Baba Moussa","email":"","orcid":"","institution":"Université d'Abomey-Calavi","correspondingAuthor":false,"prefix":"","firstName":"Lamine","middleName":"Baba","lastName":"Moussa","suffix":""},{"id":431558776,"identity":"9242c81c-8cde-4ca0-aae6-6d306115f0de","order_by":12,"name":"Pascal Abiodoun Olounladé","email":"","orcid":"","institution":"Université d'Agriculture de Kétou","correspondingAuthor":false,"prefix":"","firstName":"Pascal","middleName":"Abiodoun","lastName":"Olounladé","suffix":""}],"badges":[],"createdAt":"2025-03-10 21:38:12","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6198606/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6198606/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":79089902,"identity":"ac273ebd-84bd-43bd-9a07-dcf3828c1129","added_by":"auto","created_at":"2025-03-24 09:47:18","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":69260,"visible":true,"origin":"","legend":"\u003cp\u003eThe logarithmic correlation between the percentage of sporulated oocysts in \u003cem\u003eE. media\u003c/em\u003e and the concentrations of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves acetonic extract (1a), \u003cem\u003eO. gratissimum\u003c/em\u003e leaves aqueous extract (1b), \u003cem\u003eV. amygdalina\u003c/em\u003e leaves acetonic extract (1c), and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves aqueous extract (1d).\u003c/p\u003e","description":"","filename":"Picture1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/1354498653cacba1595e40dd.jpg"},{"id":79091245,"identity":"e16f99dc-d738-4ec7-acda-e054fc521855","added_by":"auto","created_at":"2025-03-24 10:03:18","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":65133,"visible":true,"origin":"","legend":"\u003cp\u003eThe logarithmic correlation between the percentage of sporulated oocysts of E. \u003cem\u003emagna\u003c/em\u003e and the concentrations of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves acetonic extract (2A), \u003cem\u003eO. gratissimum\u003c/em\u003e leaves aqueous extract (2B), \u003cem\u003eV. amygdalina\u003c/em\u003e leaves acetonic extract (2C), and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves aqueous extract (2D).\u003c/p\u003e","description":"","filename":"Picture2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/a52944fdedab6ad6ec3ca8ab.jpg"},{"id":79089903,"identity":"e0d4e28c-9d53-43d2-a8f3-bf7ad0c64e3a","added_by":"auto","created_at":"2025-03-24 09:47:18","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":73409,"visible":true,"origin":"","legend":"\u003cp\u003eThe logarithmic correlation between the percentage of death of \u003cem\u003eArtemia salina\u003c/em\u003e larvae and the concentration of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves aqueous extract (3A). \u003cem\u003eV. amygdalin\u003c/em\u003e leaves acetonic extract (3B), \u003cem\u003eO. gratissimum\u003c/em\u003e leaves aqueous extract (3C), and \u003cem\u003eO. gratissimum\u003c/em\u003e leaves acetonic extract (3D).\u003c/p\u003e","description":"","filename":"Picture3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/c8a70ed4bb8b4d60affb03f0.jpg"},{"id":79091029,"identity":"6d3f2d80-4567-45fb-ac9d-a90c65540289","added_by":"auto","created_at":"2025-03-24 09:55:18","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":34809,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of the extracts of the three plants on the weight growth of the rats\u003c/p\u003e","description":"","filename":"Picture4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/4b00f17b8172f7fd8f6b99e2.jpg"},{"id":79089911,"identity":"f007b45c-5c6b-4183-9b68-2a0c85073654","added_by":"auto","created_at":"2025-03-24 09:47:18","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":184467,"visible":true,"origin":"","legend":"\u003cp\u003eHistological sections of the internal organs of rats fed different acetone extracts of the two plants by single gavage observation at 400× magnification: (A) liver of rats fed with \u003cem\u003eO. gratissimum\u003c/em\u003e extract; (B) liver of rats fed with \u003cem\u003eV. amygdalin\u003c/em\u003e extract; (C) kidney of rats fed with \u003cem\u003eO. gratissimum\u003c/em\u003e extract; (D) kidney of rats fed with \u003cem\u003eV. amygdalin\u003c/em\u003eextract; (E) liver of control rats; (F) kidney of control rats. (V) Centrilobular vein; (G) glomerulus; (RT) renal tubules.\u003c/p\u003e","description":"","filename":"Picture5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/c500e3894b9293889cd3caf7.jpg"},{"id":82502593,"identity":"3597e00b-b10c-4cca-8ee3-c7f5313ac9d2","added_by":"auto","created_at":"2025-05-12 09:02:17","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2365794,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6198606/v1/cf3b9399-c29e-4517-a854-4372b96083d7.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"In vitro study of anti-coccidial activity of Ocimum gratissimum and Vernonia amygdalina leaves extracts against Eimeria magna and Eimeria media","fulltext":[{"header":"Introduction","content":"\u003cp\u003eCoccidial infections caused by protozoa of the genus Eimeria are a major threat to the health of farm animals, particularly rabbits. These infections can cause serious gastrointestinal disorders and affect the growth, reproduction and productivity of animals [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e are two species of coccidia commonly observed in rabbits and are responsible for considerable economic losses on farms due to clinical symptoms such as diarrhea, dehydration and reduced production [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. Although anticoccidial drugs are available to treat these infections, their overuse has led to the emergence of resistance, reducing their efficacy [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. In addition, growing concerns about food safety and antibiotic resistance have encouraged the search for alternative treatments, including plant extracts with therapeutic properties.\u003c/p\u003e \u003cp\u003eMedicinal plants play a central role in traditional medicine in many parts of the world, particularly in Africa and Asia, where they are used to treat a variety of conditions, including parasitic infections. Among these plants, \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e have attracted particular interest because of their antioxidant, antimicrobial and antiparasitic properties [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. These plants are distinguished by their exceptional phytochemical composition, including flavonoids, tannins, alkaloids, saponins and essential oils [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. \u003cem\u003eO. gratissimum\u003c/em\u003e is known for its high content of phenolic compounds, particularly eugenol, a bioactive phenol with antioxidant, anti-inflammatory and antimicrobial properties [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. \u003cem\u003eO. gratissimum\u003c/em\u003e, which is rich in essential oils, has antioxidant and antimicrobial activities, as confirmed by several studies [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. These properties indicate its therapeutic potential in the management of intestinal infections and metabolic disorders linked to oxidative stress, which are frequently observed in parasitic infections. In addition, its antimicrobial and antiparasitic effects suggest promising activity against protozoa, although few studies have explored its anticoccidial potential. \u003cem\u003eV. amygdalina\u003c/em\u003e, also known as the \u0026ldquo;bitter leaf\u0026rdquo;, is rich in saponins, alkaloids and flavonoids. These compounds are known for their antiparasitic, immunomodulatory and antioxidant activities [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. Saponins, in particular, have demonstrated significant antiparasitic activity by disrupting parasite membranes or modulating the host immune response [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. In addition, \u003cem\u003eV. amygdalina\u003c/em\u003e leaves are traditionally used to treat gastrointestinal disorders and intestinal infections, reinforcing their relevance in the context of coccidiosis [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThese two plants have complementary properties. \u003cem\u003eO. gratissimum\u003c/em\u003e has been studied for its antimicrobial and antioxidant properties, whereas \u003cem\u003eV. amygdalina\u003c/em\u003e is better known for its immunomodulatory and antiparasitic abilities. Together, they offer synergistic potential in the fight against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e, two species of interest because of their economic and health impacts. However, despite these promising observations, scientific evidence concerning the efficacy of these plants against Eimeria in rabbits, as well as an understanding of their mechanisms of action, remains limited. However, to consider their practical application in zootechnics, it is crucial to assess not only their efficacy but also their safety. Although plant extracts are natural, they may contain toxic compounds, and their potential toxicity at high doses or with prolonged use needs to be rigorously studied [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Indeed, the potential cytotoxic effects of these plants on parasite larvae, as well as their systemic toxicity in animal hosts, remain poorly explored. These plants contain various secondary metabolites, such as saponins, flavonoids, alkaloids and essential oils, which can have powerful biological effects but are sometimes toxic [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Assessment of larval cytotoxicity is essential for determining whether these extracts effectively target parasites without causing collateral damage to host tissues. Moreover, assessing acute toxicity in animals makes it possible to establish safe doses and avoid side effects such as liver or kidney damage, metabolic imbalances or impaired growth [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eGiven this lack of data, this study proposes an in-depth evaluation of the antioxidant capacities and \u003cem\u003ein vitro\u003c/em\u003e anticoccidial activity of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalin\u003c/em\u003e extracts against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e strains, as well as a toxicity study. This study also examined the phytochemical content of these plants to identify the bioactive compounds potentially responsible for their therapeutic effects. This research aims to answer the following questions: do extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e exert anticoccidial activity \u003cem\u003ein vitro\u003c/em\u003e against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e, and what is their possible mode of action? What is the larval cytotoxicity and acute toxicity of extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e? The hypotheses of this study are that extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e display significant antioxidant and anticoccidial activity \u003cem\u003ein vitro\u003c/em\u003e against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e without any toxicity and that this activity is associated with the contents of bioactive compounds such as flavonoids, alkaloids and tannins.\u003c/p\u003e \u003cp\u003eThis study is motivated by the need to develop safer and more sustainable alternative therapeutic solutions for coccidiosis in rabbits. The use of plant extracts could not only offer a natural alternative to chemical treatments but also help to preserve biodiversity and reduce the risks associated with antibiotics. In addition, this study could pave the way for the use of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e in integrated pest management protocols, thereby contributing to a more sustainable approach to animal disease management. The aim of this study is therefore twofold: (i) to assess the \u003cem\u003ein vitro\u003c/em\u003e antioxidant and anticoccidial activity of extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e; (ii) to identify the bioactive compounds present in these plants that are likely to play a key role in these activities; and (iii) to assess the larval and acute oral toxicity of the leaves of both plants. In doing so, this study aims to provide a sound scientific basis for the use of these plants as alternative treatments for the control of parasitic infections in rabbits.\u003c/p\u003e \u003cp\u003eThe ultimate aim of this research is to offer a natural and effective alternative to chemical treatments while helping to combat the increase in drug resistance in rabbit farms.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e \u003cb\u003eHarvesting\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe present study used the leaves of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e. \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves were harvested in April 2019 from market gardeners at the University of Abomey-Calavi (UAC) in Benin. These plants were subsequently grown on organic fertilizers containing compost. These plants were then authenticated with voucher numbers YH524/HNB and YH523/HNB, respectively, at the herbarium of UAC. The harvested leaves were washed in clean water and then dried in the laboratory at 18\u0026deg;C for 14 days. Once dry and crunchy, the leaves were ground into a powder and stored in hermetically sealed glass jars for later use.\u003c/p\u003e \u003cp\u003e \u003cb\u003ePreparation of aqueous and acetonic extracts of\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe leaves powders of both plants were used as materials for the extractions. Fifty grams of plant powder was dissolved in 500 mL of solvent S1, which was composed of 70% acetone and 30% distilled water. After 72 h of continuous maceration, the mixture was filtered through Whatman No. 1 paper, and the residue was recovered several times until exhaustion. The filtrate was evaporated under vacuum at 47\u0026deg;C using a Rotavapor (BUCHI RII). The acetonic extract was then dried in an oven at 40\u0026deg;C for 7 days.\u003c/p\u003e \u003cp\u003eThe aqueous extract was obtained by dissolution via a previously described procedure. This time, solvent S1 was replaced with distilled water. The filtrate was evaporated under vacuum at 65\u0026deg;C using a Rotavapor (BUCHI RII). The aqueous extract was then dried in an oven at 50\u0026deg;C for 7 days.\u003c/p\u003e \u003cp\u003e \u003cb\u003ePhytochemical screening of powder from\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe qualitative determination of the major groups of chemical compounds contained in the leaves of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e was carried out according to the classical method of Houghton and Raman [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], which was reviewed and adapted to the conditions of the Laboratory of Biotechnologies and Animal Improvement of the Faculty of Agronomic Sciences of UAC.\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eTotal Phenol Assay\u003c/h2\u003e \u003cp\u003eThe quantitative determination of total phenols was performed according to the classical method of Singleton, et al. [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Starting from an aqueous stock solution of gallic acid with a mass concentration of 10 mg/mL, a standard range of aqueous solutions obtained by successive dilution was prepared. A total of 625 \u0026micro;L of Folin-Ciocalteu reagent (FCR) was added to 125 \u0026micro;L of the prepared extract at 1 mg/mL in distilled water. The mixture was incubated for 5 minutes, after which 500 \u0026micro;L of sodium carbonate Na\u003csub\u003e2\u003c/sub\u003eCO3 at 75 mg/mL and 4.75 mL of distilled water were added. After 30 minutes of incubation in the dark, the optical density was read at 760 nm via a Bio Mate 5 spectrophotometer against a blank. The total phenol content was calculated via the calibration curve of 10 mg/mL gallic acid prepared under the same conditions.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eTotal Flavonoid Assay\u003c/h3\u003e\n\u003cp\u003eFive hundred microliters of the extract at 1 mg/mL in methanol was mixed with 500 \u0026micro;L of 2% AlCl3 and 3 mL of methanol. The mixture was vigorously shaken. After 10 minutes of incubation in the dark, the optical density was read at 415 nm against a blank via a BioMate3 spectrophotometer. The total flavonoid content was determined via the calibration curve of 10 mg/mL rutin according to the methods described by Kim, et al. [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e] and Zhishen, et al. [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e].\u003c/p\u003e\n\u003ch3\u003eTotal Condensed Tannin Assay\u003c/h3\u003e\n\u003cp\u003eFive hundred microliters of methanolic extract at 1 mg/mL was mixed with 3 mL of 4% vanillin. To this mixture, 2 mL of methanol and 1.5 mL of fuming hydrochloric acid were added. After 15 min of incubation, the optical density was read at 500 nm via a BioMate5 spectrophotometer against a blank according to the method described by Heimler, et al. [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e]. The standard used to determine the condensed tannin content was prepared under the same conditions with catechin\u003c/p\u003e \u003cp\u003e \u003cb\u003eAntioxidant activity of\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves extracts\u003c/b\u003e\u003c/p\u003e\n\u003ch3\u003eDiphenyl-1-picrylhydrazyl Assay\u003c/h3\u003e\n\u003cp\u003eThe evaluation of radical scavenging activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals in Crataegus fruit extracts was conducted spectrophotometrically. The method, which was originally introduced by Chokki, et al. [\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e] and further developed by Dah-Nouvlessounon, et al. [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e], was utilized. The principle of the assay relies on the color transformation of the DPPH solution, which shifts from purple to yellow as the radical is neutralized by the antioxidant. One hundred microliters of methanolic hawthorn fruit extract was mixed with 1.9 mL of 410\u0026ndash;3 mM DPPH in methanol, resulting in a total volume of 2 mL. The ability to scavenge free radicals via the DPPH radical was assessed by monitoring the decrease in absorbance at 517 nm every 2 min until the reaction reached equilibrium. Further dilution was necessary if the measured DPPH value exceeded the linear range of the standard curve [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. The inhibition activity (I%) was then calculated as follows:\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$\\:I\\left(\\%\\right)=\\frac{Abs\\:contol-Abs\\:compound}{Abs\\:control}*100$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eThe extract concentration providing 50% inhibition concentration (IC50) was calculated from the graph plotting the percentage of inhibition against the extract concentration.\u003c/p\u003e \u003cp\u003e \u003cb\u003eIn vitro\u003c/b\u003e \u003cb\u003eanticoccidial activities of\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves extracts\u003c/b\u003e\u003c/p\u003e \u003cp\u003e \u003cb\u003ePreparation of\u003c/b\u003e \u003cb\u003eE. media\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eE. magna\u003c/b\u003e \u003cb\u003ecultures\u003c/b\u003e\u003c/p\u003e \u003cp\u003ePotassium dichromate (K\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e): 2.5% potassium dichromate was prepared by dissolving 2.5 g of potassium dichromate in 100 ml of distilled water. This culture medium was stored and used to prepare the concentrations of the plant extracts.\u003c/p\u003e\n\u003ch3\u003ePreparation of sporulated oocysts:\u003c/h3\u003e\n\u003cp\u003eField isolates of \u003cem\u003eEimeria magna\u003c/em\u003e and \u003cem\u003eEimeria media\u003c/em\u003e were collected from the small intestine (jejunum and ileum) of three naturally infected rabbits, confirmed by the flotation technique. These rabbits were purchased from independent local breeders. The oocysts were then washed and concentrated using the flotation method. Sporulated oocysts were stored in a 2.5% potassium dichromate solution at 4\u0026deg;C until experimental use. Field isolates of \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e were maintained by periodic passage in healthy young albino New Zealand rabbits from the Laboratory of Biotechnology and Animal Improvement of University of Abomey-Calavi, Benin.\u003c/p\u003e \u003cp\u003e \u003cb\u003eIn vitro\u003c/b\u003e \u003cb\u003eoocysticidal effects of the extracts\u003c/b\u003e\u003c/p\u003e \u003cp\u003ePetri dishes were used to assess \u003cem\u003ein vitro\u003c/em\u003e antioocyst activities. Each well contained a total volume of 2 ml of each extract (2.5, 5, 10, 20, or 40 mg/ml) inoculated with an equal number of nonsporulated oocysts and incubated at 28\u0026deg;C. For comparison, phenol was used as the reference solution. The preparation was examined after 24 h and 48 h. The number of sporulated and nonsporulated oocysts was counted, and the percentage of sporulation was estimated by counting the number of sporulated oocysts out of a total of 100 oocysts. The percentage inhibition of sporulation was calculated as follows.\u003cdiv id=\"Equb\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equb\" name=\"EquationSource\"\u003e\n$$\\:Sporulation\\:\\left(sp\\right)inhibition\\:percentage\\:\\left(\\%\\right)=\\frac{sp\\:\\%\\:of\\:control-sp\\:\\%\\:of\\:extraact\\:}{Sp\\:of\\:control}x\\:100$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003e \u003cb\u003eIn vitro\u003c/b\u003e \u003cb\u003eanti-sporozoidal effect of extracts\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe oocysts stored in K\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e were washed several times with HBSS (pH 7.2) until the K\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e was completely eliminated. The oocysts were then incubated in a water bath at 41\u0026deg;C and shaken for 60 minutes. The suspension was centrifuged at 3,000\u0026ndash;5,000 \u0026times; g for 10 min and resuspended in HBSS. The released sporozoites were washed with HBSS. The sporozoites were counted in McMaster cells.\u003c/p\u003e \u003cp\u003ePetri dishes were used to evaluate the \u003cem\u003ein vitro\u003c/em\u003e sporocidal activity. Each well contained a total volume of 2 ml of each extract concentration (250, 500, 750, and 1000 \u0026micro;g/ml) and was inoculated with an equal number of sporozoites. For comparison, amprocox was used as the reference drug. The setup was examined after 12 h and 24 h. The number of viable and nonviable sporozoites was counted, and the percentage of viability was estimated by counting the number of viable sporozoites out of a total of 100 sporozoites [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe percentage inhibition of viability was calculated as follows.\u003cdiv id=\"Equc\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equc\" name=\"EquationSource\"\u003e\n$$\\:Viability\\:\\:\\left(Vi\\right)\\:inhibition\\:\\left(\\%\\right)=\\frac{Vi\\:\\%\\:of\\:control-Vi\\:\\%\\:of\\:extrac}{Vi\\:\\%\\:of\\:control}x100$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003e \u003cb\u003eLarval cytotoxicity test of aqueous and acetonic extracts of\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves\u003c/b\u003e\u003c/p\u003e \u003cp\u003e \u003cem\u003eArtemia salina\u003c/em\u003e eggs were used for the larval cytotoxicity test. The cytotoxic effect of the hydroethanol extract of the plants studied was assessed on brine shrimp larvae via a preliminary nonclinical toxicity test as described by Adoho, et al. [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. \u003cem\u003eArtemia salina\u003c/em\u003e larvae were obtained by incubating 10 mg of \u003cem\u003eArtemia salina\u003c/em\u003e eggs in 1 L of seawater under continuous agitation for 48 hours.\u003c/p\u003e \u003cp\u003eSerial dilutions of the extract were prepared from a stock solution (20 mg/mL) to obtain increasing concentrations. Each diluted solution (1 ml) was mixed with 1 ml of seawater containing 16 live larvae. A control solution, without extract, was prepared simultaneously under the same conditions. After incubation for 24 hours under continuous agitation, the number of dead larvae in each solution was determined via optical microscopy. These data were used to establish a dose‒response curve (the number of surviving larvae as a function of extract concentration).\u003c/p\u003e \u003cp\u003eThe concentrations were then log-transformed, and the median lethal concentration (LC\u003csub\u003e50\u003c/sub\u003e) was calculated. Larval toxicity was interpreted on the basis of the correlation grid proposed by Ugwah-Oguejiofor, et al. [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e], which associates the degree of toxicity with the LC\u003csub\u003e50\u003c/sub\u003e value. According to this grid:\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"No\" id=\"Taba\" border=\"1\"\u003e \u003ccolgroup cols=\"1\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026ge;\u0026thinsp;0.1 mg/mL \u0026loz; Nontoxic extract\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0.1 mg/mL\u0026thinsp;\u0026gt;\u0026thinsp;LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026ge;\u0026thinsp;0.050 mg/mL \u0026loz; Low toxicity\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e0.050 mg/mL\u0026thinsp;\u0026gt;\u0026thinsp;LC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026ge;\u0026thinsp;0.0 mg/mL \u0026loz;Medium toxicity\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.01 mg/mL \u0026loz; High toxicity\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eAcute oral toxicity of acetonic extracts of\u003c/b\u003e \u003cb\u003eO. gratissimum\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eV. amygdalina\u003c/b\u003e \u003cb\u003eleaves\u003c/b\u003e\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eAnimal materials\u003c/h2\u003e \u003cp\u003eNine female albino Wistar rats weighing between 150 and 200 g and at least three months of age were used for the acute oral toxicity test. These nulliparous, nonpregnant female rats were supplied by the Institute of Applied Biomedical Sciences at the University of Abomey-Calavi, Benin. The rats were then randomly housed in groups of 3 in cages fitted with stainless steel lids to allow 14 days of acclimatization at the animal house of the Zootechnical Research and Livestock System Unit (URZoSE) at the National University of Agriculture. The rats had access to water and food \u003cem\u003ead libitum\u003c/em\u003e. The daily temperature was 23\u0026thinsp;\u0026plusmn;\u0026thinsp;2\u0026deg;C, and the relative humidity was 60\u0026thinsp;\u0026plusmn;\u0026thinsp;10%, with a 12-hour light/dark cycle.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eStudy design\u003c/h3\u003e\n\u003cp\u003eThe toxicity of the acetonic extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e was evaluated according to the guidelines of the Organization for Economic Cooperation and Development (OECD) No. 423 for chemical testing [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. This method allows the determination of the dose of extract tolerated by the animal subjects. The 9 rats were divided into 3 groups of 3 rats each, thus forming groups of similar average weights. Group 1 (control), group 2 (rats receiving the acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves), and group 3 (rats receiving the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves) were included. Before the extracts were administered, all the rats underwent a 12-hour fast during which they had access only to water. The individual weight of each rat was determined. The rats in groups 2 (n\u0026thinsp;=\u0026thinsp;3) and 3 (n\u0026thinsp;=\u0026thinsp;3) received a single dose of acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e dissolved in 1 ml of distilled water by oral gavage. Group 1 (n\u0026thinsp;=\u0026thinsp;3) received 1 ml of distilled water by oral gavage. The rats were observed individually for the first thirty minutes and regularly for the first 24 h and every day for 14 days to observe any signs of macroscopic toxicity (convulsions, agitation, a moribund state, severe distress, or animal mortality) and possible mortality. Animal weights were measured on days 1, 7 and 14 of the experiment. At the end of the 14-day observation period, the animals were anaesthetized with thiopental and then euthanized by cervical dislocation. The major organs\u0026mdash;the brain, liver, spleen, heart and kidneys \u0026mdash; were removed, weighed and macroscopically observed for lesions and abnormalities. The relative weight of each organ was calculated by relating its weight to the body weight of the corresponding animal. In addition, samples of these organs were fixed in 10% neutral buffered formalin for further histological analysis.\u003c/p\u003e \u003cp\u003e All animal handling, including oral administration and posttreatment observations, was performed by qualified personnel specifically trained in rodent care and handling in accordance with institutional and regulatory guidelines.\u003c/p\u003e\n\u003ch3\u003eBiochemical and haematological analysis\u003c/h3\u003e\n\u003cp\u003eOn day 14 of treatment, a blood sample (1 ml) was taken from each experimental animal in a dry tube and in a tube with EDTA via puncture of the retro-orbital plexus via heparinized capillary tubes. The blood in the dry tube was centrifuged at 2,500 rpm for 5 minutes at 4\u0026deg;C. Serum samples collected in this way were subjected to biochemical analysis via specific commercial kits for the quantification of various biomarkers: blood glucose (G O D - P AP, Biolabo. #Ref 7409), creatine kinetics (creatinine, Biolabo. #Ref 0107), urea (Uree, Biolabo. #Ref 0221), aspartate aminotransferase (AST, Biolabo. #Ref 0025), alanine aminotransferase (ALT, Biolabo. #Ref 0027), alkaline phosphatase (ALP, Biolabo. #Ref 80014) and gamma-glutamyl transferase (γ-GT, Biolabo. #Ref 81310). Readings were taken on an SP-350-BIO spectrometer, COLE-PARMER\u0026reg; formerly GenovaPlus from JENWAY\u0026reg;. The blood samples collected in EDTA-coated tubes were used to analyse various hematological parameters, including red blood cell (RBC) count, white blood cell (WBC) count, hemoglobin (Hb) concentration, packed cell volume (PCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), and blood platelet (BP) count. These parameters were measured via an automated Beckman Coulter Ac hematology analyser [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e].\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eHistological examination\u003c/h2\u003e \u003cp\u003eThe rats were sacrificed, and tissue samples from the liver and kidneys were removed and processed into paraffin blocks via labelled tissue processing cassettes. The samples were dehydrated with increasing concentrations of alcohol (70%, 80%, 90% and absolute alcohol). Further dehydration was carried out via two changes of absolute alcohol, followed by three changes of xylene. The tissues were then infiltrated and embedded in paraffin. Sections (4 \u0026micro;m thick) were cut from each paraffin block, mounted on microscope slides and stained with hematoxylin and eosin (H\u0026amp;E) as described previously Ugwah-Oguejiofor, et al. [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. The stained sections were examined under an Olympus microscope for morphological changes, and representative images were captured.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eThe experimental data were systematically organized and analyzed using the Excel 2019 database. The data were analyzed using one-way analysis of variance (ANOVA) and presented as the mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (SD) of 3 replicates. Significance levels were determined using the Waller-Duncan test, with p-values\u0026thinsp;\u0026lt;\u0026thinsp;0.05 considered statistically significant. Statistical analysis was performed using the R software (version 4.4.2). The IC50 values of the extracts and their main compounds, which showed high anticoccidial efficacy, and the LC50 values, which showed the safety of the extracts, were evaluated by regression analysis. For graphical representation and further statistical analysis, GraphPad Prism\u0026reg; software (version 10.4.1) was utilized\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003ch2\u003ePhytochemical screening\u003c/h2\u003e\n\u003cp\u003eSeveral chemical groups are present in the leaves of both plants, suggesting that they share similar biological properties. Common groups include tannins (catechic and gallic), flavonoids, quinone derivatives, coumarins, reducing compounds, O-heterosides (reduced and unreduced), C-heterosides and alkaloids. There are also some notable differences, such as leucoanthocyanins, which are absent in the leaves of \u003cem\u003eO. gratissimum\u003c/em\u003e but present in the leaves of \u003cem\u003eV. amygdalina\u003c/em\u003e. Saponosides are absent in the leaves of \u003cem\u003eO. gratissimum\u003c/em\u003e but are present at relatively high concentrations in the leaves of \u003cem\u003eV. amygdalina\u003c/em\u003e (lather \u0026gt; 1 cm). Free anthracenics and cyanogenic derivatives are absent in the leaves of both plants.\u003c/p\u003e\n\u003cp\u003eTable 1. Qualitative screening of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u0026nbsp;\u003c/em\u003epowder leaves.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"575\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e\u003cem\u003eO. gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e\u003cem\u003eV.\u003c/em\u003e\u003cem\u003e\u0026nbsp;amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eTannins\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eCatechin tannins\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eGallic tannins\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eLeucoanthocyanins\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eFlavonoids\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eQuinone derivatives\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eSaponosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+ \u0026gt; 1\u0026nbsp;cm\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eMucilages\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eCoumarins\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eReducing compounds\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eFree anthracenics\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eO-heterosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eReduced genine O-heterosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eC-heterosides\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eAlkaloids\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 257px;\"\u003e\n \u003cp\u003eCyanogenic derivatives\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 167px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 151px;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"3\" valign=\"top\" style=\"width: 575px;\"\u003e\n \u003cp\u003e\u0026ldquo;+\u0026rdquo; = presence; \u0026ldquo;-\u0026rsquo;\u0026rsquo; = absence; \u0026gt;1 cm represents the height of the lather created.\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch2\u003ePolyphenol and flavonoid contents of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u0026nbsp;\u003c/em\u003epowder leaves\u003c/h2\u003e\n\u003cp\u003eThe acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves (VaAc) had the highest total phenol content (736.27 \u0026plusmn; 0.97 mg EAG/g ES), followed by the acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves (OgAc), 549.85 \u0026plusmn; 1.55 mg EAG/g ES (p˂0.05). The aqueous extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAq) and \u003cem\u003eO. gratissimum\u003c/em\u003e leaves (OgAq) presented the lowest total phenol contents, with values of 420.35 \u0026plusmn; 1.94 mg EAG/g ES and 374.48 \u0026plusmn; 0.58 mg EAG/g ES, respectively (p˂0.05). The acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e had the highest flavonoid content (37.06 \u0026plusmn; 0.31 mg ER/g ES), followed by the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (30.71 \u0026plusmn; 0.68 mg ER/g ES). However, the aqueous extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e (18.89 \u0026plusmn; 0.48 mg ER/g ES) and \u003cem\u003eV. amygdalina\u003c/em\u003e (16.74 \u0026plusmn; 0.19 mg ER/g ES) presented the lowest levels (p˂0.05). Condensed tannins were present in high amounts in the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (5.94 \u0026plusmn; 0.42 mg EC/g ES), followed by the aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (3.53 \u0026plusmn; 0.52 mg EC/g ES). The aqueous and acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e presented the lowest concentrations (p˂0.05), 2.83 \u0026plusmn; 0.13 mg EC/g ES and 1.53 \u0026plusmn; 0.12 mg EC/g ES, respectively (Table 2).\u003c/p\u003e\n\u003cp\u003eTable 2. Contents of total phenols, flavonoids and condensed tannins in \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV.\u0026nbsp;\u003c/em\u003eamygdalina leaves extracts.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"769\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 137px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" style=\"width: 193px;\"\u003e\n \u003cp\u003e\u003cem\u003eO. gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"top\" style=\"width: 194px;\"\u003e\n \u003cp\u003e\u003cem\u003eV. amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 122px;\"\u003e\n \u003cp\u003eEquations\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 55px;\"\u003e\n \u003cp\u003eR\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 67px;\"\u003e\n \u003cp\u003ePvalue\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 137px;\"\u003e\n \u003cp\u003eExtraits\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 122px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 55px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 67px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 137px;\"\u003e\n \u003cp\u003eTotal phenols (mg EAG/g ES)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e374.48\u0026plusmn;0.58a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e549.85\u0026plusmn;1.55d\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e420.35\u0026plusmn;1.94c\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e736.27\u0026plusmn;0.97f\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 122px;\"\u003e\n \u003cp\u003ey= 0.115x + 0.003\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 55px;\"\u003e\n \u003cp\u003e0.998\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 67px;\"\u003e\n \u003cp\u003e2.E-16\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 137px;\"\u003e\n \u003cp\u003eFlavono\u0026iuml;ds (mg ER/g ES)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e18.89\u0026plusmn;0.48b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e37.06\u0026plusmn;0.31e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e16.74\u0026plusmn;0.19a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e30.71\u0026plusmn;0.68c\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 122px;\"\u003e\n \u003cp\u003ey = 0.160x + 0.006\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 55px;\"\u003e\n \u003cp\u003e0.9993\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 67px;\"\u003e\n \u003cp\u003e6.36E-15\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 137px;\"\u003e\n \u003cp\u003eCondensed tannins (mg EC/g ES)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e1.53\u0026plusmn;0.12a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e2.83\u0026plusmn;0.13a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 96px;\"\u003e\n \u003cp\u003e3.53\u0026plusmn;0.52b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 97px;\"\u003e\n \u003cp\u003e5.94\u0026plusmn;0.42d\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 122px;\"\u003e\n \u003cp\u003ey = 0.129x + 0.005\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 55px;\"\u003e\n \u003cp\u003e0.9953\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 67px;\"\u003e\n \u003cp\u003e9.56E-12\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"8\" valign=\"top\" style=\"width: 769px;\"\u003e\n \u003cp\u003eThe values bearing the different letters a, b, c, d, and e in the same line are significantly different at the 5% level (p \u0026lt; 0.05) for the same metabolite. OgAq: Aqueous extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e, OgAc: Acetonic extract of \u003cem\u003eO. gratissimum\u003c/em\u003e, VaAq: Aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e, VaAc: Acetonic extract of \u003cem\u003eV. amygdalin\u003c/em\u003e, EAG: gallic acid equivalent, ER: rutin equivalent, EC: catechin equivalent, ES: dry extract.\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch2\u003eAntioxidant activity\u003c/h2\u003e\n\u003cp\u003eAscorbic acid (Acid Asc) has a relatively low effective concentration 50 (EC\u003csub\u003e50\u003c/sub\u003e) value (2.802 \u0026plusmn; 0.024), with R\u003csup\u003e2\u003c/sup\u003e=0.9747. This finding indicates high efficacy in producing an effect (standard reference). The acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves (VaAc) had a slightly higher EC\u003csub\u003e50\u003c/sub\u003e (3.541 \u0026plusmn; 0.112) than ascorbic acid did, with an R\u003csup\u003e2\u003c/sup\u003e = 0.986, suggesting that it was less effective than the standard. The acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (OgAc) had an EC\u003csub\u003e50\u003c/sub\u003e (2.4025 \u0026plusmn; 0.0385) even lower than that of ascorbic acid, with an R\u003csup\u003e2\u003c/sup\u003e = 0.9457, indicating high efficacy. The aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAq) presented the highest EC\u003csub\u003e50\u003c/sub\u003e value (4.0305 \u0026plusmn; 0.101), with R\u003csup\u003e2\u003c/sup\u003e= 0.9439, indicating that it was the least effective of the extracts tested. The aqueous extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (OgAq) presented a moderate EC\u003csub\u003e50\u003c/sub\u003e (3.467 \u0026plusmn; 0.022) with R\u003csup\u003e2\u003c/sup\u003e = 0.9856, which was lower than that of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAq) but higher than those of the acetonic extracts and the standard (Table 3).\u003c/p\u003e\n\u003cp\u003eIn summary, the acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (OgAc) had the lowest EC\u003csub\u003e50\u003c/sub\u003e (2.4025 \u0026plusmn; 0.0385), followed by ascorbic acid (AcidAsc), which was used as a reference. These findings suggest that this extract has a high capacity for action at relatively low concentrations. The aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAq) had the highest EC\u003csub\u003e50\u003c/sub\u003e (4.0305 \u0026plusmn; 0.101), indicating lower efficacy. Acetonic extracts had a lower EC\u003csub\u003e50\u003c/sub\u003e (better efficiency) than aqueous extracts, probably due to better extraction of bioactive compounds (Table 3).\u003c/p\u003e\n\u003cp\u003eTable 3. Free radical scavenging activity via the DPPH method\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"402\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003eEC\u003csub\u003e50\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eCI\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003eR square\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\n \u003cp\u003eAcid Asc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003e2.802\u0026plusmn;0.024\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e1.930 to 4.055\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e0.9747\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003e3.541\u0026plusmn;0.112\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e2.229 to 5.359\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e0.986\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003e2.4025\u0026plusmn;0.0385\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e1.154 to 4.315\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e0.9457\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003e4.0305\u0026plusmn;0.101\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e2.587 to 8.024\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e0.9439\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 82px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 119px;\"\u003e\n \u003cp\u003e3.467\u0026plusmn;0.022\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e2.385 to 5.283\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e0.9856\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch2\u003e\u003cem\u003eIn vitro\u003c/em\u003e oocysticidal activities of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves extracts.\u003c/h2\u003e\n\u003cp\u003eAt low concentrations (2.5 mg/mL), the sporulation inhibition efficacy was generally low for all the extracts. The acetonioic extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves had the greatest activity, with inhibition percentages reaching 35.33% for \u003cem\u003eE. magna\u003c/em\u003e at 24 h and 44.67% for \u003cem\u003eE. media\u003c/em\u003e at 48 h. The other extracts produced significantly lower results (p˂0.05). At an intermediate concentration (5\u0026ndash;10 mg/mL), inhibition increased in proportion to the concentration. The \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e leaves acetone extracts remained significantly more effective (p˂0.05), reaching nearly 62.67% (\u003cem\u003eE. magna\u003c/em\u003e, 24 h) for the \u003cem\u003eV. amygdalina\u003c/em\u003e leaves acetone extracts at 5 mg/mL and up to 82.33% (\u003cem\u003eE. magna\u003c/em\u003e, 24 h) at 10 mg/mL for the \u003cem\u003eO. gratissimum\u003c/em\u003e leaves acetone extract. At high concentrations (20\u0026ndash;40 mg/mL), inhibition was maximal, particularly for the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves at 40 mg/mL. The percentage of inhibition reached 91% at 24 h and 100% at 48 h for \u003cem\u003eE. magna\u003c/em\u003e with the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e. Similar results were recorded for \u003cem\u003eE. media\u003c/em\u003e (Table 4).\u003c/p\u003e\n\u003cp\u003eThe aqueous extracts of the two plants were less effective overall than the acetonic extracts were. The positive control (5%) shows almost complete inhibition (97.67% to 100%) for both species, confirming that the extracts tested do not exceed this efficacy. The inhibition of the negative control (K₂Cr₂O₇ alone or with DMSO) was very low (6\u0026ndash;9%), indicating that the effect observed with the extracts was due mainly to the bioactive compounds present in the plants. Acetone extracts, particularly from \u003cem\u003eV. amygdalina\u003c/em\u003e leaves, were the most effective against \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e at all concentrations tested. These values almost reached the values of the positive control at high concentrations (40 mg/mL). The aqueous extracts showed moderate efficacy, with the aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves being slightly superior to the aqueous extract of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves. The inhibition of both Eimeria species increased with increasing extract concentration. At high concentrations (20\u0026ndash;40 mg/mL), the effect was close to or equal to that of the positive control (Table 4).\u003c/p\u003e\n\u003cp\u003eTable 4. Sporulation percentages of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e in different d\u0026rsquo;\u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e.\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"606\"\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 118px;\"\u003e\n \u003cp\u003eConcentration\u003c/p\u003e\n \u003cp\u003e(mg/ml)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 115px;\"\u003e\n \u003cp\u003eExtracts\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" style=\"width: 373px;\"\u003e\n \u003cp\u003eIncubation times for Eimeria strains\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"2\" style=\"width: 176px;\"\u003e\n \u003cp\u003e\u003cem\u003eE. magna\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 197px;\"\u003e\n \u003cp\u003e\u003cem\u003eE. media\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 86px;\"\u003e\n \u003cp\u003e\u003cem\u003e24 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 90px;\"\u003e\n \u003cp\u003e\u003cem\u003e\u0026nbsp;48 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 87px;\"\u003e\n \u003cp\u003e\u003cem\u003e\u0026nbsp;24 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 110px;\"\u003e\n \u003cp\u003e\u003cem\u003e\u0026nbsp;48 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e2.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e13.67\u0026plusmn;4.73\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e28.33\u0026plusmn;3.21\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e12.00\u0026plusmn;3.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e25.67\u0026plusmn;8.96\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e18.00\u0026plusmn;5.29\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e36.33\u0026plusmn;4.16\u003csup\u003eabc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e14.33\u0026plusmn;2.89\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e37.33\u0026plusmn;6.43\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e23.67\u0026plusmn;1.53\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e30.00\u0026plusmn;4.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e19.33\u0026plusmn;1.53\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e34.67\u0026plusmn;9.24\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e35.33\u0026plusmn;4.51\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e43.33\u0026plusmn;3.79\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e22.33\u0026plusmn;0.58\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e44.67\u0026plusmn;4.51\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e33.33\u0026plusmn;4.16\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e58.00\u0026plusmn;2.65\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e33.00\u0026plusmn;4.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e54.00\u0026plusmn;9.54\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e46.00\u0026plusmn;5.29\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e66.00\u0026plusmn;5.57\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e40.00\u0026plusmn;3.61\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e67.33\u0026plusmn;7.02\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e49.67\u0026plusmn;3.51\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e60.33\u0026plusmn;4.73\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e39.33\u0026plusmn;2.89\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e63.33\u0026plusmn;11.55\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e62.67\u0026plusmn;10.21\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e71.00\u0026plusmn;2.65\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e51.00\u0026plusmn;1.73\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e73.33\u0026plusmn;3.79\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e51.00\u0026plusmn;1.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e78.33\u0026plusmn;10.21\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e49.00\u0026plusmn;6.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e75.33\u0026plusmn;11.85\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e64.33\u0026plusmn;6.81\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e85.33\u0026plusmn;4.04\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e64.00\u0026plusmn;1.00\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e86.33\u0026plusmn;5.69\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e69.00\u0026plusmn;6.93\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e81.33\u0026plusmn;4.73\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e60.00\u0026plusmn;2.65\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e85.00\u0026plusmn;8.72\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e82.33\u0026plusmn;10.02\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e90.00\u0026plusmn;2.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e70.33\u0026plusmn;0.58\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e91.67\u0026plusmn;5.13\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e55.00\u0026plusmn;5.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e83.00\u0026plusmn;8.89\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e54.00\u0026plusmn;7.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e80.67\u0026plusmn;9.61\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e69.00\u0026plusmn;4.58\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e90.33\u0026plusmn;3.21\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e68.33\u0026plusmn;1.53\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e92.33\u0026plusmn;1.53\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e73.67\u0026plusmn;7.77\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e85.33\u0026plusmn;6.66\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e63.67\u0026plusmn;3.51\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e87.33\u0026plusmn;9.29\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e88.00\u0026plusmn;7.21\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e94.67\u0026plusmn;1.53\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e78.00\u0026plusmn;6.08\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e96.67\u0026plusmn;4.16\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e62.00\u0026plusmn;6.56\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e87.00\u0026plusmn;8.72\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e57.33\u0026plusmn;6.51\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e84.00\u0026plusmn;8.89\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e71.67\u0026plusmn;3.79\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e93.00\u0026plusmn;4.58\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e73.00\u0026plusmn;1.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e95.00\u0026plusmn;2.65\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e78.00\u0026plusmn;8.19\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e91.00\u0026plusmn;6.24\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e68.33\u0026plusmn;3.51\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e90.67\u0026plusmn;7.77\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e91.00\u0026plusmn;2.65\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e84.33\u0026plusmn;5.13\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\u0026nbsp;\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003eNagative controle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eK\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u0026nbsp;\u003c/sub\u003e+DMSO\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e6\u0026plusmn;1.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e6.33\u0026plusmn;1.53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e7\u0026plusmn;2.65\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e5.66\u0026plusmn;152\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003eK\u003csub\u003e2\u003c/sub\u003eCr\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e7\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e9\u0026plusmn;2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e9.67\u0026plusmn;0.58\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e9\u0026plusmn;1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e9.00\u0026plusmn;2.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 118px;\"\u003e\n \u003cp\u003ePositivve controle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 115px;\"\u003e\n \u003cp\u003e5%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 86px;\"\u003e\n \u003cp\u003e97.67\u0026plusmn;2.52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 90px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 87px;\"\u003e\n \u003cp\u003e97.00\u0026plusmn;3.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 110px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\" valign=\"top\" style=\"width: 606px;\"\u003e\n \u003cp\u003eThe values bearing the different letters a, b, c, d, and e in the same column are significantly different at the 5% level (p \u0026lt; 0.05) for the same metabolite. OgAq: Aqueous extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e, OgAc: Acetonic extract of \u003cem\u003eO. gratissimum\u003c/em\u003e, VaAq: Aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e, VaAq: Acetonic extract of \u003cem\u003eV. amygdalin\u003c/em\u003e,\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch2\u003e\u003cem\u003eIn vitro\u003c/em\u003e anti-sporozoidal activities of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves extracts\u003c/h2\u003e\n\u003cp\u003eThe efficacy of the extracts increased significantly (p˂0.05) with concentration, regardless of the type of extract or Eimeria species. At a dose of 125 \u0026micro;g/ml, the percentages of viability inhibition of \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e strains were relatively low, with greater efficacy observed for the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (29.00 \u0026plusmn; 2.65% on \u003cem\u003eE. magna\u003c/em\u003e at 12 h and 47.33 \u0026plusmn; 6.03% at 24 h). At a dose of 250 \u0026micro;g/ml, a clear increase in the percentage of inhibition was observed for all extracts, with the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e retaining its superiority (47.00 \u0026plusmn; 2.00% at 12 h and 65.67 \u0026plusmn; 1.15% at 24 h over \u003cem\u003eE. magna\u003c/em\u003e). At a dose of 500 \u0026micro;g/ml, the efficacy of the aqueous extracts of the two plants was similar to that of the acetone extracts, although the percentage inhibition of the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e remained greater (91.00 \u0026plusmn; 1.73% in \u003cem\u003eE. media\u003c/em\u003e at 24 h). At a dose of 1000 \u0026micro;g/ml, the extracts reached efficacy levels close to or equal to 100%, particularly for the acetone extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e. The effect was dose dependent (Table 5).\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eE. magna\u003c/em\u003e responded slightly more sensitively to the extracts, particularly the acetone extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves, at 24 hours. At 1000 \u0026micro;g/ml, the acetone extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves achieved 100% inhibition, which was identical to that of the positive control. \u003cem\u003eE. media\u003c/em\u003e showed a similar trend to that of \u003cem\u003eE. magna\u003c/em\u003e, but the inhibition values were often slightly lower (Table 5).\u003c/p\u003e\n\u003cp\u003eTable 5. Percent inhibition of viability of \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u0026nbsp;\u003c/em\u003estrains in contact with \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalin\u003c/em\u003e extracts\u003c/p\u003e\n\u003ctable width=\"572\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 110px;\"\u003e\n \u003cp\u003eConcentration\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u0026micro;g/ml\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 76px;\"\u003e\n \u003cp\u003eExtracts\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"4\" style=\"width: 385px;\"\u003e\n \u003cp\u003eIncubation times for Eimeria strains\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"2\" style=\"width: 177px;\"\u003e\n \u003cp\u003e\u003cem\u003eE. magna\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 208px;\"\u003e\n \u003cp\u003e\u003cem\u003eE. media\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e\u003cem\u003e12 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u003cem\u003e24 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e\u003cem\u003e12 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e\u003cem\u003e24 hours\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" style=\"width: 110px;\"\u003e\n \u003cp\u003e125\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e5.67\u0026plusmn;1.15\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e14.67\u0026plusmn;3.79\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e6.33\u0026plusmn;1.53\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e13.33\u0026plusmn;3.21\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e11.67\u0026plusmn;2.52\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e23.67\u0026plusmn;6.51\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e13.33\u0026plusmn;2.08\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e24.33\u0026plusmn;8.14\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e16.67\u0026plusmn;5.69\u003csup\u003eba\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e33.33\u0026plusmn;7.02\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e16.67\u0026plusmn;2.52\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e32.00\u0026plusmn;10.39\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e29.00\u0026plusmn;2.65\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e47.33\u0026plusmn;6.03\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e31.00\u0026plusmn;4.58\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e46.00\u0026plusmn;1.73\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 110px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" style=\"width: 110px;\"\u003e\n \u003cp\u003e250\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e23.33\u0026plusmn;7.02\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e39.00\u0026plusmn;2.65\u003csup\u003ee\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e23.00\u0026plusmn;3.61\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e38.67\u0026plusmn;8.62\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e36.00\u0026plusmn;5.57\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e51.33\u0026plusmn;11.06\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e32.67\u0026plusmn;3.21\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e51.33\u0026plusmn;6.43\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e35.67\u0026plusmn;4.04\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e44.33\u0026plusmn;0.58\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e35.00\u0026plusmn;6.56\u003csup\u003eabc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e43.67\u0026plusmn;1.15\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e47.00\u0026plusmn;2.00\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e65.67\u0026plusmn;1.15\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e46.33\u0026plusmn;4.93\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e67.00\u0026plusmn;3.00\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 110px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" style=\"width: 110px;\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e50.67\u0026plusmn;4.62\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e58.67\u0026plusmn;4.73\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e49.67\u0026plusmn;6.66\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e57.67\u0026plusmn;7.37\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e63.00\u0026plusmn;6.24\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e88.33\u0026plusmn;11.72\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e63.67\u0026plusmn;8.50\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e85.00\u0026plusmn;5.29\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e52.33\u0026plusmn;8.33\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e72.67\u0026plusmn;11.50\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e48.33\u0026plusmn;9.45\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e73.00\u0026plusmn;8.19\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e77.00\u0026plusmn;5.20\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e89.67\u0026plusmn;0.58\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e78.00\u0026plusmn;10.54\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e91.00\u0026plusmn;1.73\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 110px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"4\" style=\"width: 110px;\"\u003e\n \u003cp\u003e1000\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e65.00\u0026plusmn;4.58\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e77.67\u0026plusmn;2.52\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e66.67\u0026plusmn;9.07\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e78.67\u0026plusmn;6.66\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e80.67\u0026plusmn;7.64\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e93.67\u0026plusmn;1.53\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e83.67\u0026plusmn;9.50\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e97.00\u0026plusmn;3.00\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e74.67\u0026plusmn;7.57\u003csup\u003eab\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e88.67\u0026plusmn;4.16\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e73.00\u0026plusmn;8.89\u003csup\u003ebc\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e92.67\u0026plusmn;5.51\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e91.67\u0026plusmn;2.08\u003csup\u003ec\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003csup\u003ed\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e90.67\u0026plusmn;11.15\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 110px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 110px;\"\u003e\n \u003cp\u003eNegative control\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eDMSO\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003eHBSS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e0.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 110px;\"\u003e\n \u003cp\u003ePositive control\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 76px;\"\u003e\n \u003cp\u003e50 \u0026micro;g/ml\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 85px;\"\u003e\n \u003cp\u003e88.67\u0026plusmn;3.51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 92px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 95px;\"\u003e\n \u003cp\u003e89.00\u0026plusmn;1.73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 113px;\"\u003e\n \u003cp\u003e100.00\u0026plusmn;0.00\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"6\" style=\"width: 572px;\"\u003e\n \u003cp\u003eOgAq: Aqueous extract of \u003cem\u003eO. gratissimum\u003c/em\u003e, OgAc: Acetonic extract of \u003cem\u003eO. gratissimum\u003c/em\u003e, VaAq: Aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e, VaAc: Acetonic extract of \u003cem\u003eV. amygdalina\u003c/em\u003e, DMSO: Dimethyl sulfoxide, HBSS: Hank\u0026apos;s buffered salt solution. The results are presented as the means and standard deviations of triplicate \u003cem\u003ein vitro\u003c/em\u003e tests after 12 h and 24 h of incubation at room temperature. For the same column and concentration, values with the same superscript letter are not significantly different at p \u0026ge; 0.05 (Duncan\u0026apos;s test)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eThe curves show an inverse relationship between the compound concentration and oocyst sporulation. As the concentration increased, sporulation decreased, which is typical of antiparasitic substances. The IC₅₀ values indicate the concentrations required to inhibit 50% of sporulation. The IC\u003csub\u003e50\u003c/sub\u003e values of the extracts from the two plants were therefore examined in more detail via regression analysis (Figure 1). Aqueous and acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves inhibited the sporulation of 50% of \u003cem\u003eE. media\u003c/em\u003e coccidia oocysts at a concentration of 3.466 mg/ml (IC₅₀ = 3. 466 mg/ml) and 4.413 mg/ml (IC₅₀ = 4.413 mg/ml), with coefficients of determination R\u0026sup2; = 0.9749 and R\u0026sup2; = 0.9873, respectively (Figure 1A and 1B). In contrast, aqueous and acetone extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves inhibited the sporulation of 50% of \u003cem\u003eE. media\u003c/em\u003e oocysts at a concentration of 2. 920 mg/ml (IC₅₀ = 2.920 mg/ml) and 3.415 mg/ml (IC₅₀ = 3.415 mg/ml), with coefficients of determination R\u0026sup2; = 0.9873 and R\u0026sup2; = 0.9817, respectively (Figure 1C and 1D). The logarithmic equation had the highest coefficient of determination (R\u003csup\u003e2\u003c/sup\u003e) (Figure 1C and 1D), and the IC\u003csub\u003e50\u003c/sub\u003e values were 2.92 and 3.41 mg/ml for the acetone and aqueous extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves, respectively (Figure 1).\u003c/p\u003e\n\u003cp\u003eThis figure illustrates the inhibitory effects of different leaves extracts on the sporulation of \u003cem\u003eE. magna\u003c/em\u003e oocysts. The IC\u003csub\u003e50\u003c/sub\u003e (median inhibitory concentration) and R\u003csup\u003e2\u003c/sup\u003e values were used to assess the power and goodness of fit of the dose‒response curves. The high R\u003csup\u003e2\u003c/sup\u003e values (\u0026gt;0.97) for all the graphs confirm a strong correlation between the extract concentrations and the reduction in the percentage of sporulated oocysts (Figure 2). The results show that extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e (Figure 2 C and D) are more effective than those of \u003cem\u003eO. gratissimum\u003c/em\u003e (Figure 2A and D) in inhibiting the sporulation of \u003cem\u003eE. magna\u003c/em\u003e oocysts. The lower IC\u003csub\u003e50\u003c/sub\u003e values for \u003cem\u003eV. amygdalina\u003c/em\u003e (3.097 mg/mL for the acetone extract and 3.408 mg/mL for the aqueous extract) suggest better anticoccidial activity, regardless of the extraction method. In comparison, \u003cem\u003eO. gratissimum\u003c/em\u003e had slightly higher IC\u003csub\u003e50\u003c/sub\u003e values (4.066 mg/mL for the acetone extract and 4.091 mg/mL for the aqueous extract), indicating moderate efficacy (Figure 2).\u003c/p\u003e\n\u003ch2\u003eLarval Toxicity of Extracts.\u003c/h2\u003e\n\u003cp\u003eThe larval toxicity results revealed that the plant extracts studied were not toxic to \u003cem\u003eArtemia salina\u003c/em\u003e larvae, with notable differences between the aqueous and acetonic extracts. The aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (Figure 3A), with an LC\u003csub\u003e50\u003c/sub\u003e of 0.269 mg/mL (R\u0026sup2; = 0.93), was classified as a nontoxic extract. Similarly, the aqueous extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (Figure 3C) had an LC\u003csub\u003e50\u003c/sub\u003e of 0.168 mg/mL (R\u0026sup2; = 0.95), which also placed it in the category of nontoxic extracts, indicating better accuracy in model fitting. In contrast, the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e has an LC\u003csub\u003e50\u003c/sub\u003e of 0.978 mg/mL (R\u0026sup2; = 0.92), which also classifies it as nontoxic (Figure 3B), whereas the acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e, with an LC\u003csub\u003e50\u003c/sub\u003e of 0.137 mg/mL, is also considered nontoxic, although the R\u0026sup2; of 0.76 suggests some variability in the model fit (Figure 3D). In summary, both plants produced mainly nontoxic extracts, with LC\u003csub\u003e50\u003c/sub\u003e variations influenced by the type of solvent used (Figure 3).\u003c/p\u003e\n\u003ch2\u003eAcute oral toxicity\u003c/h2\u003e\n\u003ch3\u003eMortality and clinical signs\u003c/h3\u003e\n\u003cp\u003eNo mortality was observed in the rats treated with the acetone extracts of the three plants during the acute toxicity tests. No signs or abnormalities were recorded on the skin, coat, eyes or mucous membranes. There was also a complete absence of hyperactivity, eye twitches, catalepsy, convulsions, tremors and catatonia following treatment with the acetonic fractions of the plants. Administration of the acetonic extracts produced no signs of erythema or edema until after 48 hours of observation.\u003c/p\u003e\n\u003ch3\u003eFood consumption and body weight of the rats\u003c/h3\u003e\n\u003cp\u003eThere were no significant differences in food consumption between the test group and the control group. The weights of the subjects in the treated batches differed significantly (p \u0026lt; 0.05) from those in the control batches at D7 and D14 (Figure 4).\u003c/p\u003e\n\u003ch3\u003eHematological parameters of the rats\u003c/h3\u003e\n\u003cp\u003eThe results in the table show several significant differences in haematological parameters among the three groups. White blood cell (WBC) counts were significantly lower in the OgAc group (12.7 \u0026plusmn; 0.9) than in the control group (13.98 \u0026plusmn; 0.38) and the VaAc group (13.25 \u0026plusmn; 0.05), with a p value of 0.008. Compared with the control group (7.35 \u0026plusmn; 0.55) and the OgAc group (7.755 \u0026plusmn; 0.015), the VaAc group presented a significant reduction in red blood cell count (6.63 \u0026plusmn; 0.1). Haemoglobin (Hb) levels were significantly lower in the VaAc group (13.35 \u0026plusmn; 0.05) than in the control (14.425 \u0026plusmn; 0.425) and OgAc (14.5 \u0026plusmn; 0) groups. Red blood cell (RBC) volume was significantly greater (p=0.017) in the OgAc (48.4 \u0026plusmn; 0.2) and VaAc (47.2 \u0026plusmn; 1.8) groups than in the control group (43.35 \u0026plusmn; 0.35). The mean corpuscular volume (MCV) was significantly greater (p ˂0.001) in the VaAc group (71.15 \u0026plusmn; 1.65) than in the control group (62.55 \u0026plusmn; 1.05) and the OgAc group (62.4 \u0026plusmn; 0.1). The mean amount of haemoglobin per red blood cell (MCH) was significantly lower (p ˂0.001) in the OgAc group (18.7 \u0026plusmn; 0) than in the Control (23 \u0026plusmn; 1) and VaAc (20.15 \u0026plusmn; 0.35) groups. The hemoglobin concentration per red blood cell volume (MCHC) was significantly lower (p ˂0.001) in the OgAc (29.95 \u0026plusmn; 0.15) and VaAc (28.3 \u0026plusmn; 1.2) groups than in the control group (34.85 \u0026plusmn; 1.15). Finally, the platelet count (PCR) was significantly lower (p ˂0.001) in the OgAc group (578 \u0026plusmn; 13) than in the control group (745.5 \u0026plusmn; 5.5) and the VaAc group (793.5 \u0026plusmn; 3.5). These results suggest that treatments with extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e (OgAc) and \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAc) modify several blood parameters, particularly white blood cells, red blood cells, haemoglobin and platelets (Table 6).\u003c/p\u003e\n\u003cp\u003eTable 6. Hematological parameters of the rats.\u003c/p\u003e\n\u003ctable width=\"523\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003eControl\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003eP value\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eWBC (10\u003csup\u003e3\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e13.98\u0026plusmn;0.38 a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e12.7\u0026plusmn;0.9\u0026nbsp;b\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e13.25 \u0026plusmn;0.05 a\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e0.008\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eRBC (10\u003csup\u003e6\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e7.35\u0026plusmn;0.55\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e7.755\u0026plusmn;0.015\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e6.63\u0026plusmn;0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e0.031\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eHb (g/dL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e14.425\u0026plusmn;0.425\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e14.5\u0026plusmn;0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e13.35\u0026plusmn;0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e0.029\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003ePVC (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e43.35\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e48.4\u0026plusmn;0.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e47.2\u0026plusmn;1.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e0.017\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eMCV (fL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e62.55\u0026plusmn;1.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e62.4\u0026plusmn;0.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e71.15 \u0026plusmn;1.65\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eMCH (pg)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e23\u0026plusmn;1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e18.7\u0026plusmn;0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e20.15\u0026plusmn;0.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eMCHC (g/dL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e34.85\u0026plusmn;1.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e29.95\u0026plusmn;0.15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e28.3\u0026plusmn;1.2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 120px;\"\u003e\n \u003cp\u003eBP (10\u003csup\u003e3\u003c/sup\u003e/mm\u003csup\u003e3\u003c/sup\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 107px;\"\u003e\n \u003cp\u003e745.5\u0026plusmn;5.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 103px;\"\u003e\n \u003cp\u003e578\u0026plusmn;13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 94px;\"\u003e\n \u003cp\u003e793.5\u0026plusmn;3.5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 99px;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\" style=\"width: 523px;\"\u003e\n \u003cp\u003eRBC=Red blood cells. PCV=Packed cell volume. MCV=mean corpuscular volume. MCH=mean corpuscular haemoglobin. MCHC=mean corpuscular haemoglobin concentration. WBC=white blood cell. OgAc: Acetonic extract of \u003cem\u003eO. gratissimum\u003c/em\u003e. VaAc: Acetonic extract of \u003cem\u003eV. amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch3\u003eBiochemical parameters of\u0026nbsp;the rats\u003c/h3\u003e\n\u003cp\u003eThe results show that there was no significant difference in blood glucose levels between the three groups, with similar values for the Control (0.72 \u0026plusmn; 0.05), OgAc (0.67 \u0026plusmn; 0.03) and VaAc (0.66 \u0026plusmn; 0.01) groups. However, the OgAc group had a significantly higher urea concentration (0.33 \u0026plusmn; 0.08) compared with the Control group (0.20 \u0026plusmn; 0.01), while the VaAc group showed no difference compared with the Control group. No significant difference was observed for creatinine between groups. ALT levels were significantly higher in the OgAc (92.08 \u0026plusmn; 1.01) and VaAc (98.66 \u0026plusmn; 0.74) groups than in the control group (66.58 \u0026plusmn; 2.02). The VaAc group also had a significantly higher AST concentration (148.38 \u0026plusmn; 0.96) than the control group (124.33 \u0026plusmn; 5.09). Although there was a significant difference for ALP, the values were not radically different between the groups, with the OgAc group having a higher value (477.43 \u0026plusmn; 31.75). Finally, the VaAc group showed a higher concentration of\u0026nbsp;\u0026gamma;‐GT\u0026nbsp;(4.52 \u0026plusmn; 0.67) than the control group (3.85 \u0026plusmn; 0.08), with a significant difference (p = 0.006), (Table 7).\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;Table 7. Biochemical parameters of the rats\u003c/p\u003e\n\u003ctable width=\"568\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003eControl\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003epvalue\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eGlycemia (mg/dL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e0.72\u0026plusmn;0.05\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e0.67\u0026plusmn;0.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e0.66\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.143\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eUrea (g/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e0.20\u0026plusmn;0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e0.33\u0026plusmn;0.08*\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e0.24\u0026plusmn;0.04\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.007\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eCreatinine (g/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e10.12\u0026plusmn;1.85\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e11.51\u0026plusmn;0.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e12.46\u0026plusmn;2.48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.304\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eALT (U/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e66.58\u0026plusmn;2.02\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e92.08\u0026plusmn;1.01**\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e98.66\u0026plusmn;0.74**\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.000\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eAST (U/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e124.33\u0026plusmn;5.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e121.13\u0026plusmn;2.13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e148.38\u0026plusmn;0.96**\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.000\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003eALP (U/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e446.83\u0026plusmn;2.35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e477.43\u0026plusmn;31.75\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e419.82\u0026plusmn;28.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 136px;\"\u003e\n \u003cp\u003e\u0026gamma;‐GT (U/L)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 96px;\"\u003e\n \u003cp\u003e3.85\u0026plusmn;0.08\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003e3.47\u0026plusmn;0.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 111px;\"\u003e\n \u003cp\u003e4.52\u0026plusmn;0.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 109px;\"\u003e\n \u003cp\u003e0.006\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"5\" style=\"width: 568px;\"\u003e\n \u003cp\u003e* = p \u0026lt; 0.05; ** = p \u0026lt; 0.01. AST=aspartate amino transaminase. ALT=alanine amino transaminase. ALP=alkaline phosphate, \u0026gamma;‐GT= Gamma‐Glutamyl Transferase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003ch3\u003eRelative organ weights and necroscopy of the rats\u003c/h3\u003e\n\u003cp\u003eRelative organ weight was calculated via the formula (organ weight/body weight) \u0026times; 100%. Relative vital organ weights and microscopic changes were studied in comparison with those in the control group, and no significant changes (p\u0026gt;0.05) were observed.\u003c/p\u003e\n\u003ch3\u003eHistopathological examination of different organs of rats\u003c/h3\u003e\n\u003cp\u003eHistological analysis of livers from rats fed acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves revealed a generally preserved hepatic architecture, with well-defined hepatocytes but diffuse cytoplasmic vacuolation, suggesting potential lipid accumulation or mild hydropic degeneration. The hepatic sinusoids appear slightly dilated in some areas, with no major disruption to the radial organization of the hepatocytes. The portal areas and central veins showed no evidence of inflammation, fibrosis or abnormal cellular infiltration, indicating the absence of severe liver toxicity. Although the uneven quality of the staining may have introduced artefacts, the observations suggest a slight adaptive metabolic response to the extract (Figure 5A). Histological analysis of liver sections from rats fed an acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves revealed a generally preserved lobular architecture, with hepatocytes organized radially around the central veins. Hepatic cells showed diffuse cytoplasmic vacuolation, suggesting lipid accumulation or hydropic degeneration, possibly in the metabolic response to the extract. The hepatic sinusoids appeared mildly dilated, whereas the portal areas and central veins showed no evidence of inflammatory infiltration or fibrosis. No obvious hepatocellular damage, such as necrosis or apoptosis, was detected. A few artefacts related to section preparation and staining were noted, but these did not alter the overall interpretation of the observations. These results suggest reasonable hepatic tolerance to the extract (Figure 5B), as do the liver structures of the control rats (Figure 5C). Histological analysis of kidney sections from rats fed an acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e leaves revealed well-preserved overall nephron architecture, including glomeruli and renal tubules. The glomeruli generally appear intact, with normal vascularisation and no obvious signs of sclerosis or narrowing. Bowman\u0026apos;s spaces are homogeneous and show no signs of edema or inflammation. However, subtle alterations were observed in the proximal tubules, with evidence of diffuse cytoplasmic vacuolation, probably related to reversible degeneration or a metabolic response to the extract compounds. The distal tubules appeared normal, with no signs of atrophy or obstruction. No obvious inflammatory infiltrates or necrosis were identified in the renal interstices. Minor artefacts attributed to sample preparation and staining were noted but had no impact on the interpretation of the results. These observations suggest satisfactory renal tolerance to the extract (Figure 5D). Histological observation of renal sections from rats fed acetone extract from the leaves of \u003cem\u003eV. amygdalina\u003c/em\u003e revealed an intact overall organization of the glomeruli and renal tubules. The glomeruli appear well defined, with an intact capillary structure, indicating unimpaired glomerular filtration. The proximal and distal tubules have slightly granulated cytoplasm, probably due to normal metabolic activity or a slight adaptive response to the extract. Some areas presented with cytoplasmic vacuoles and slight disorganization, which were interpreted as artefacts of fixation or sectioning. The absence of necrosis, significant inflammatory infiltration or alterations to the basement membrane suggested that the \u003cem\u003eV. amygdalin\u003c/em\u003e extract did not cause apparent renal toxicity at the administered dose (Figure 5E). The structures of these kidneys are similar to those of control rat kidneys (Figure 5F).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe results obtained for \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e show that these plants are rich in bioactive compounds such as tannins, flavonoids, saponosides, mucilages, alkaloids and heterosides, which are of great interest for animal production and health [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. Tannins and flavonoids are well known for their antioxidant and anti-inflammatory properties and play key roles in reducing oxidative stress and managing inflammatory diseases in animals. This activity could be particularly beneficial in animal production systems where animals are exposed to high levels of stress, which can affect their immunity and productivity. For example, flavonoid-rich extracts could be used to prevent infectious diseases by reducing the bacterial load and modulating immune responses [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e]. Saponosides, which are present only in \u003cem\u003eV. amygdalina\u003c/em\u003e, are also of major interest in animal production because of their antimicrobial and immunomodulatory properties. Their ability to improve resistance to bacterial and parasitic infections could be used to reduce the use of antibiotics in livestock farming, thereby promoting a more sustainable and ethical approach to animal production. Moreover, mucilage can be used to improve digestive health.\u003c/p\u003e \u003cp\u003eThe results show that acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e have significantly higher concentrations of bioactive compounds, particularly total phenols, flavonoids and condensed tannins, than aqueous extracts do. \u003cem\u003eV. amygdalina\u003c/em\u003e, in particular, contains relatively high levels of phenols (736.27 mg EAG/g ES) and condensed tannins (5.94 mg EC/g ES) in its acetone extract, suggesting greater antioxidant and antimicrobial potential [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Similarly, flavonoids, mainly the acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (37.06 mg ER/g ES), are known for their anti-inflammatory and cardioprotective effects, which enhance intestinal and immune health [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e, \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. These results indicate that acetone extracts from both plants could play a key role in the management of oxidative stress and infections in animal production. These bioactive compounds, particularly acetonic extracts, have significant potential for animal health, offering natural alternatives to antibiotics and other chemicals. Their incorporation into animal diets could improve resistance to infection, promote digestive health and support the immune system [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. In addition, the antimicrobial effects of tannins and flavonoids suggest that they could be used to treat digestive disorders and reduce infectious diseases in animal production systems.\u003c/p\u003e \u003cp\u003eThe study compared the EC50 values and confidence intervals of antioxidant activity for different extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e (Va) and \u003cem\u003eO. gratissimum\u003c/em\u003e (Og). The acetone extract of \u003cem\u003eO. gratissimum\u003c/em\u003e (Og Ac) exhibited the lowest EC50 value (2.40), indicating higher efficacy at a lower concentration compared to other extracts. The confidence interval for OgAc (1.15\u0026ndash;4.31) was narrow, reflecting good precision in its efficacy estimation. In contrast, the aqueous extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAq) had a higher EC\u003csub\u003e50\u003c/sub\u003e value (4.03), indicating the need for a higher concentration for a similar effect. Its wider confidence interval (2.59\u0026ndash;8.02) suggests more variability in its efficacy. These findings highlight important differences that could impact animal health and productivity management [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. The acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e (VaAc) has a moderate EC\u003csub\u003e50\u003c/sub\u003e value (3.54) and an R\u003csup\u003e2\u003c/sup\u003e of 0.986, indicating that this extract has a strong ability to explain the variation in biological effects as a function of concentration. In comparison, \u003cem\u003eO. gratissimum\u003c/em\u003e has a slightly lower R\u003csup\u003e2\u003c/sup\u003e for its acetone extract (0.9457), suggesting a less precise relationship between the concentration and the observed effect; however, this difference is relatively small and could be considered acceptable for practical applications [\u003cspan additionalcitationids=\"CR37\" citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe acetonic extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e, with low EC50 values (2.40 and 3.54 respectively), could serve as effective natural alternatives for managing oxidative stress and infectious diseases in farm animals. Their low EC50 values indicate that these extracts are potent at low concentrations, which may reduce costs and minimize the risk of side effects. In intensive breeding systems, incorporating these extracts could enhance animals' resistance to oxidative stress, reduce infectious diseases, and support growth [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. The use of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e extracts could reduce the reliance on antiparasitics, particularly for preventing digestive and respiratory infections, by acting as natural antimicrobial agents. This would support sustainable animal husbandry while enhancing animal health and productivity. The \u003cem\u003ein vitro\u003c/em\u003e results showed significant inhibition of sporulation of \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e oocysts, with acetone extracts being more effective than aqueous extracts due to their higher content of lipophilic compounds such as flavonoids, condensed tannins, and alkaloids. This observation is consistent with the work of Dagnino, et al. [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e] and Okoye, et al. [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e], who reported that fat-soluble bioactive compounds extracted from medicinal plants often exhibit better antiparasitic activity \u003cem\u003ein vitro\u003c/em\u003e. At high concentrations (40 mg/ml), the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e achieved 100% inhibition of oocyst sporulation after 48 hours, rivalling the efficacy of commercial coccidiostats.\u003c/p\u003e \u003cp\u003eThese \u003cem\u003ein vitro\u003c/em\u003e results highlight the potential of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalin\u003c/em\u003e extracts as natural alternatives for the management of coccidiosis in cattle. \u003cem\u003eIn vitro\u003c/em\u003e tests provide a valuable first indication, but they do not consider the complex factors present in the body, such as bioavailability, metabolism of bioactive compounds and potential interactions with the intestinal microflora [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e]. The progressive increase in efficacy with concentration and incubation time reflects a dose‒dependent relationship, which is often observed \u003cem\u003ein vitro\u003c/em\u003e pharmacological tests. The results also confirmed that aqueous extracts are less effective than acetonic extracts are, probably due to the low solubility of lipophilic compounds in water. These observations are consistent with previous work, such as that of [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e], which demonstrated that organic solvent-based extracts generally exhibit better antiparasitic activity against coccidial strains. The efficacy of extracts may also be attributed to their ability to disrupt the cell membrane of oocysts or interfere with their energy metabolism, as suggested by studies on the properties of tannins and flavonoids against protozoa [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e].\u003c/p\u003e \u003cp\u003e \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e extracts have antiparasitic effects on their bioactive compounds. Flavonoids and tannins, which are present in large quantities, are known to alter the membranes of oocysts and inhibit enzymes essential for their sporulation. Alkaloids, on the other hand, can disrupt protozoan replication mechanisms by targeting their energy metabolism. These mechanisms of action explain the high inhibition observed at increasing concentrations. The differences in efficacy between the plant species studied could be attributed to the variability in the chemical composition of the extracts, as noted by Kebede, et al. [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e], who reported that pest control properties vary depending on the relative concentrations of flavonoids and tannins in the plant extracts. In addition, the acetone extract of \u003cem\u003eV. amygdalina\u003c/em\u003e, which has shown the best efficacy, seems to contain particularly potent bioactive compounds or is better extracted in organic solvents.\u003c/p\u003e \u003cp\u003eExtracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e dose-dependently inhibited the viability of oocysts in \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e, with a maximum performance for acetone extracts, particularly \u003cem\u003eV. amygdalina\u003c/em\u003e leaves acetone extract (VaAc). At 1000 \u0026micro;g/ml, after 24 hours, VaAc achieved 100% inhibition, matching the positive control, whereas the aqueous extracts showed less, but significant, efficacy. These results indicate the importance of lipophilic metabolites, such as flavonoids and alkaloids, which are better extracted by acetone, for antiparasitic activity [\u003cspan additionalcitationids=\"CR49\" citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. The phenolic and tannic compounds present in these extracts could interfere with oocyst membranes, leading to osmotic imbalances and metabolic inhibition [\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e]. Alkaloids, in particular, may target enzymes essential for oocyst development, a hypothesis supported by similar studies on other plant extracts [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e, \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e, \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e]. These data reinforce interest in the use of plant extracts as natural coccidiostats in poultry and rabbit farming, where coccidiosis remains a major cause of economic losses [\u003cspan additionalcitationids=\"CR55\" citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e]. The increased efficacy of acetone extracts compared to aqueous extracts can be attributed to their ability to extract more fat-soluble active compounds, such as apolar flavonoids and lignans, which are less effectively extracted in aqueous solutions. This emphasizes the importance of solvent selection to optimize the antiparasitic activity of plant extracts. These findings are especially relevant in rabbit production, where coccidiosis poses significant health and economic challenges. The inhibition of \u003cem\u003eE. media\u003c/em\u003e oocyst sporulation suggests that these extracts could help limit the environmental spread of infectious oocysts, reducing infectious pressure on farms and potentially replacing chemical coccidiostats, addressing concerns about drug resistance and residues. The toxicity tests on Artemia salina larvae showed no toxicity, with LC50 values well above the toxic threshold, indicating that both aqueous and acetonic extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e are safe for use.\u003c/p\u003e \u003cp\u003eThese results are consistent with those of previous studies that have shown that extracts of these plants can have little or no biological effects on certain larval species, depending on the concentration used [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e, \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e, \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e]. The differences in the LC\u003csub\u003e50\u003c/sub\u003e values observed between the aqueous and acetonic extracts can be explained by the nature of the compounds extracted by each solvent. The aqueous extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e contain mainly polyphenols and flavonoids, which are known for their antioxidant effects and low toxicity [\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e, \u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e]. In contrast, acetone extracts, which are rich in lipophilic compounds such as terpenes and alkaloids, could theoretically exhibit greater biological activity, but here, they are also classified as nontoxic, suggesting that the bioavailability of secondary metabolites in these extracts is not high enough to result in significant toxicity to \u003cem\u003eArtemia salina\u003c/em\u003e larvae [\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e, \u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e]. Although the extracts were classified as nontoxic in this study, these results cannot be generalized to other species or long-term applications. Plant extracts can have toxic effects at relatively high concentrations or after prolonged exposure. Furthermore, these extracts have shown pharmacological potential in other contexts, notably in the treatment of human diseases, but their ecological impact must be taken into account before any application in the natural environment [\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e, \u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e]. In summary, these results indicate that aqueous and acetonic extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e have little or no toxicity to \u003cem\u003eArtemia salina\u003c/em\u003e larvae.\u003c/p\u003e \u003cp\u003eThe results of histological analyses of liver and kidney tissues from rats fed acetone extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e leaves provide essential information on the potential metabolic and toxicological effects of these phytochemicals. Both extracts showed a general preservation of histoarchitecture in liver and kidney tissues, suggesting their relative safety at the doses administered. The diffuse cytoplasmic vacuolation observed in hepatocytes from both experimental groups indicates potential lipid accumulation or mild hydropic degeneration. These changes may reflect adaptive responses to the bioactive constituents present in \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e. Similar observations have been reported in studies evaluating the hepatoprotective and hepatomodulatory properties of phytochemicals, suggesting that mild vacuolization may represent a transient metabolic adjustment to increased lipid metabolism or oxidative stress [\u003cspan additionalcitationids=\"CR66 CR67\" citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e]. Importantly, the absence of fibrosis, necrosis or inflammatory infiltration in portal and central veins supports the idea that these extracts do not induce overt liver toxicity under the conditions studied. This finding is consistent with previous findings indicating that \u003cem\u003eO. gratissimum\u003c/em\u003e possesses antioxidant and hepatoprotective properties, mainly attributed to its high polyphenol and flavonoid contents [\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e]. Renal histological analysis revealed preserved glomerular and tubular architecture, with no evidence of necrosis, inflammation or basement membrane rupture in either group. Subtle cytoplasmic vacuolation in proximal tubules may suggest reversible metabolic adjustments or mild degenerative changes related to biotransformation of plant extract constituents. These observations are consistent with the literature, which reports mild renal changes in response to bioactive compounds without significant functional alterations [\u003cspan additionalcitationids=\"CR71\" citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e]. The absence of glomerular sclerosis or interstitial fibrosis further supports the renal safety of these extracts at the doses tested. Notably, \u003cem\u003eV. amygdalina\u003c/em\u003e has been reported to exert nephroprotective effects in models of oxidative stress, probably due to its richness in saponins, tannins and flavonoids [\u003cspan additionalcitationids=\"CR74\" citationid=\"CR73\" class=\"CitationRef\"\u003e73\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e]. Although the two extracts produced similar histological results, slight differences in vacuolar changes and sinusoidal dilatation between the groups suggest variations in the bioavailability and metabolic processing of their respective phytochemical profiles. These differences could stem from the distinct chemical compositions of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e, which include eugenol, terpenes and flavonoids for the former and sesquiterpene lactones and saponins for the latter [\u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e76\u003c/span\u003e].\u003c/p\u003e \u003cdiv id=\"Sec24\" class=\"Section2\"\u003e \u003ch2\u003eLimits\u003c/h2\u003e \u003cp\u003eHowever, these observations remain limited to \u003cem\u003ein vitro\u003c/em\u003e conditions, where complex host\u0026ndash;environment interactions are absent. It would be crucial to confirm these effects in in vivo assays while assessing the bioavailability, safety and impact of extracts on zootechnical performance. In addition, thorough characterization of active compounds and optimization of formulations are needed to maximize efficacy and minimize the required doses.\u003c/p\u003e \u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThis study evaluated the antioxidant and anticoccidial properties of aqueous and acetonic extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e and \u003cem\u003eO. gratissimum\u003c/em\u003e for their potential to manage coccidial infections in rabbits. Antioxidant analyses revealed significant activity attributed to the presence of bioactive compounds such as flavonoids and phenols, which were particularly concentrated in the acetonic extracts. The acetone extracts also markedly inhibited the sporulation of \u003cem\u003eE. magna\u003c/em\u003e and \u003cem\u003eE. media\u003c/em\u003e oocysts, suggesting greater efficacy than aqueous extracts. \u003cem\u003eIn vitro\u003c/em\u003e and in vivo toxicity tests confirmed the relative safety of the extracts, with no evidence of larval toxicity or acute oral toxicity. Histological examination of the target organs revealed no toxicity at a dose of 2000 mg/kg. Among the plants studied, \u003cem\u003eV. amygdalina\u003c/em\u003e stood out for its superior anticoccidial properties. These results highlight the potential of these plants as natural and sustainable alternatives to conventional treatments for coccidiosis while offering additional benefits linked to their antioxidant properties.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e%\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ePercentage\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u0026micro;g/ml\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMicrograms per milliliter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eAcidAsc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAscorbic Acid\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eALP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ealkaline phosphatase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eALT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAlanine aminotransaminase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eAST\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAspartate Aminotransaminase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eBP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eBlood Patelets\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eCI\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eConfidence Interval\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eDMSO\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eDimethyl sulfoxide\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eDPPH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003e2,2-diphenyl-1-picrylhydrazyl\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u003cem\u003eE.magna\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003e\u003cem\u003eEimeria magna\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u003cem\u003eE.media\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003e\u003cem\u003eEimeria media\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eEAG\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eGallic Acid Equivalent\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eEC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eCatechin Equivalent\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eEC\u003csub\u003e50\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eHalf maximal effective concentration\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eER\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eRutin Equivalent\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eES\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eDry Extract\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eFCR\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eFolin-Ciocalteu reagent\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003efL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003efemtoliter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eg/dL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eGrams per deciliter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eg/L\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eGrams per liter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eh\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eHour\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eHBSS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eHank\u0026apos;s buffered salt solution\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eIC\u003csub\u003e50\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ehalf-maximal inhibitory concentration\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eK2Cr2O\u003csub\u003e7\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ePotassium dichromate\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eLaBAA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eLaboratory of Biotechnology and Animal Improvement\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eLC\u003csub\u003e50\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eLethal concentration 50%\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eMCH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMean Corpuscular Haemoglobin\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eMCHC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMean Corpuscular Haemoglobin Concentration\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eMCV\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMean Corpuscular Volume\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003emg/dL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMilligrams per deciliter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003emg/ml\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eMilligram per millilitre\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003emm\u003csup\u003e3\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eCubic millimetres\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eNa\u003csub\u003e2\u003c/sub\u003eCO\u003csub\u003e3\u003c/sub\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eSodium carbonate\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u003cem\u003eO. gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003e\u003cem\u003eOcimum gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eOgAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAcetonic extract of \u003cem\u003eOcimum gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eOgAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAqueous extract of \u003cem\u003eOcimum gratissimum\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003ePCV\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ePacked Cell Volume\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003epg\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003ePicograms\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eR\u003csup\u003e2\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eRsquare\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eRBC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eRed blood cells\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eU/L\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eUnits per liter\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u003cem\u003eV.amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003e\u003cem\u003eVernonia amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eVaAc\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAcetonic extract of \u003cem\u003eVernonia amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eVaAq\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eAqueous extract of \u003cem\u003eVernonia amygdalina\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003eWBC\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eWhite Blood Cells\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 145px;\"\u003e\n \u003cp\u003e\u0026gamma;‐GT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 392px;\"\u003e\n \u003cp\u003eGamma‐Glutamyl Transferase\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003cp\u003eEthics approval and consent to participate\u003c/p\u003e\n\u003cp\u003eThe study was approved by the Research and Ethics Committee of the National Agricultural University of Benin (N\u0026deg; 143-2018/President-CER/SA). Throughout the study, measures were taken to ensure that animal welfare was respected during experimentation. The experiments were carried out by trained and certified researchers in compliance with current regulations on the use of animals for scientific purposes (European Directive 2010/63/EU/IACUC).\u003c/p\u003e\n\u003cp\u003eConsent for publication\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003eAvailability of data and materials\u003c/p\u003e\n\u003cp\u003eThe raw data for the current study are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003eCompeting Interests\u003c/p\u003e\n\u003cp\u003eThe authors declare that no competing interests exist.\u003c/p\u003e\n\u003cp\u003eFunding\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThis paper received no funding.\u003c/p\u003e\n\u003cp\u003eAuthors\u0026apos; contributions\u003c/p\u003e\n\u003cp\u003eB.K. Conceptualization, Formal analysis, Investigation, Methodology, Software, Writing \u0026ndash; original draft,\u0026nbsp;Writing \u0026ndash; review \u0026amp; editing; C.C.D. Investigation, Methodology, Writing \u0026ndash; review \u0026amp; editing; L.D. Investigation, Resources , Writing \u0026ndash; review \u0026amp; editing; R.T. Investigation, Writing \u0026ndash; review \u0026amp; editing; C.H. Investigation, Resources , Writing \u0026ndash; review \u0026amp; editing; F.L.M.A. D. Writing \u0026ndash; review \u0026amp; editing; G.G.A. Investigation, Writing \u0026ndash; review \u0026amp; editing; E.A. Conceptualization, Resources, Supervision, Validation, Writing \u0026ndash; original draft; Writing \u0026ndash; review \u0026amp; editing; S-y.D.A. Conceptualization, Funding acquisition, Project administration, Resources, Supervision, Validation, Visualization, Writing \u0026ndash; original draft; Writing \u0026ndash; review \u0026amp; editing; S.H.A-H. Writing \u0026ndash; review \u0026amp; editing; L.B.M. Resources, Supervision, Validation, Visualization, Writing \u0026ndash; original draft; Writing \u0026ndash; review \u0026amp; editing; P.A.O. Project administration Conceptualization, Methodology, Project administration, Funding acquisition, Resources, Software, Supervision, Validation, Visualization, Writing \u0026ndash; original draft; Writing \u0026ndash; review \u0026amp; editing.\u003c/p\u003e\n\u003cp\u003eAcknowledgements\u003c/p\u003e\n\u003cp\u003eWe would like to thank the entire team of the Laboratory of Biotechnology and Animal Improvement (LaBAA) of the Faculty of Agronomic Sciences of the University of Abomey-Calavi.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eOkumu PO, Gathumbi PK, Karanja DN, Mande JD, Wanyoike MM, Gachuiri CK, Kiarie N, Mwanza RN, Borter DK. 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Egypt J Basic Appl Sci. 2024;11(1):469\u0026ndash;78.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFarombi EO, Owoeye O. Antioxidative and Chemopreventive Properties of \u003cem\u003eVernonia amygdalina\u003c/em\u003e and \u003cem\u003eGarcinia biflavonoid\u003c/em\u003e. Int J Environ Res Public Health. 2011;8(6):2533\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAsante D-B, Wiafe GA. Therapeutic Benefit of \u003cem\u003eVernonia amygdalina\u003c/em\u003e in the Treatment of Diabetes and Its Associated Complications in Preclinical Studies. J Diabetes Res. 2023;2023(1):3159352.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMgbemena NM, Amako NF. Comparative Analysis of the Phytochemicals, Proximate and Mineral Compositions of Scent Leaf (\u003cem\u003eOcimum gratissimum\u003c/em\u003e) and Bitter Leaf (\u003cem\u003eVernonia amygdalina\u003c/em\u003e) Leaves. Int J Biochem Res Rev. 2020;29(7):1\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Natural coccidiostat, Oxidative stress, Medicinal plants, Bioactive substances, Biocompatibility","lastPublishedDoi":"10.21203/rs.3.rs-6198606/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6198606/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003e \u003cem\u003eOcimum gratissimum\u003c/em\u003e and \u003cem\u003eVernonia amygdalina\u003c/em\u003e are used in traditional medicine for their various biological properties, but few studies have assessed their antioxidant and anticoccidial capacities and toxicity. The aim of this study was to evaluate the antioxidant capacity, anticoccidial activity (against \u003cem\u003eEimeria magna\u003c/em\u003e and \u003cem\u003eEimeria media\u003c/em\u003e), phytochemical content and toxicity of leaves extracts from two plants.\u003c/p\u003e\u003ch2\u003eMethodology:\u003c/h2\u003e \u003cp\u003eAqueous and acetonic extracts of the leaves of both plants were prepared and analysed for their polyphenol, flavonoid and tannin contents. Antioxidant activity was measured via the DPPH method. The anticoccidial effects of increasing concentrations of extracts were tested on Eimeria oocysts and sporozoites. Toxicity was assessed by mortality tests on \u003cem\u003eArtemia salina\u003c/em\u003e larvae and clinical tests on rats.\u003c/p\u003e\u003ch2\u003eResult\u003c/h2\u003e \u003cp\u003eThe results show that extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e leaves have greater antioxidant and anticoccidial capacities than those of \u003cem\u003eO. gratissimum\u003c/em\u003e. The acetone extracts of the two plants presented lower EC\u003csub\u003e50\u003c/sub\u003e values (2.4025\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0385) for \u003cem\u003eO. gratissimum\u003c/em\u003e and 3.541\u0026thinsp;\u0026plusmn;\u0026thinsp;0.112 for \u003cem\u003eV. amygdalina\u003c/em\u003e) than did the aqueous extracts. With respect to anti-ocyst activity, acetone extracts of \u003cem\u003eV. amygdalina\u003c/em\u003e inhibited sporulation of \u003cem\u003eE. magna\u003c/em\u003e oocysts by up to 91% at 40 mg/mL (p˂0.05). Toxicity tests revealed that the extracts were not toxic to \u003cem\u003eArtemia salina\u003c/em\u003e. No major histological changes were observed. \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e extracts have significant antioxidant and anticoccidial properties and are nontoxic for antioxidant and anticoccidial applications.\u003c/p\u003e\u003ch2\u003eConclusion\u003c/h2\u003e \u003cp\u003eThis study shows that extracts of \u003cem\u003eO. gratissimum\u003c/em\u003e and \u003cem\u003eV. amygdalina\u003c/em\u003e possess antioxidant and anticoccidial properties, with superior efficacy for \u003cem\u003eV. amygdalina\u003c/em\u003e. These extracts have potential as natural alternatives for the management of coccidiosis in farm animals, helping to reduce the use of synthetic antiparasitics and promote sustainable agricultural practices. Further studies on their long-term effects and their application in livestock farming are required.\u003c/p\u003e","manuscriptTitle":"In vitro study of anti-coccidial activity of Ocimum gratissimum and Vernonia amygdalina leaves extracts against Eimeria magna and Eimeria media","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-03-24 09:47:13","doi":"10.21203/rs.3.rs-6198606/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"e3922575-6ff5-4293-bf20-7266656f9f45","owner":[],"postedDate":"March 24th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-05-12T08:54:01+00:00","versionOfRecord":[],"versionCreatedAt":"2025-03-24 09:47:13","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6198606","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6198606","identity":"rs-6198606","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}