In vitro and in silico anticoccidial activity of the seeds of Lepidium sativum red and black varieties

Abstract Coccidiosis is a parasitic disease of major economic importance in the poultry industry. The exploration of natural alternative strategies such as phytochemicals is becoming significant in controlling coccidiosis. Therefore, in the present study, we investigate the in vitro anticoccidial activity of the seeds of red and black Lepidium sativum varieties that have multiple traditional medicinal uses and identify their major active components. The seeds of red and black L. sativum varieties were extracted with 80% methanol and tested for their in vitro anticoccidial activity using a sporulation inhibition assay. The 80% methanol extract of the red variety was solvent partitioned into four fractions, and each was tested for activity. Then, column chromatography of the active fractions was carried out to isolate compounds. The chemical structures of the compounds were determined by spectroscopic methods (1H, 13C and DEPT NMR). The 80 methanol extract of both the black and red varieties inhibited sporulation of oocysts by 83.71% and 98.27% at concentrations of 1000 mg/ml, respectively. Among all tested substances, the chloroform fraction, followed by the hexane, was found to be the most active one with sporulation inhibition values of 93.5% and 98.91% at 500 and 1000 mg/ml doses, respectively. Moreover, linoleic acid, alpha-linolenic acid and methyl sinapate were identified and characterized as the major components of the active fractions. Furthermore, molecular docking and conformational similarity scanning study of the compounds and anticoccidial drugs revealed that methyl sinapate had a strong binding affinity and conformational alignment to dihydrofolate reductase (HYDE score, -41.6 KJ/mol) and trimethoprim (FlexS scores, 0.84), respectively. In addition, to find further chemically similar analogs of methyl sinapate, we navigated the chemical space library of billions of commercially available synthetic compounds using the bioinformatics tool infiniSee 5.0.1 and identified 100 molecules with FlexS scores in the range of 0.67–0.78. These findings along with the safety profile derived from the acute oral toxicity tests of the extract suggest L. sativum as an alternative source of treatment for coccidiosis.