BackgroundDue to its rich content of active phytochemicals, milk thistle is regarded as a promising nutritional supplement for quails, particularly in regions with limited financial resources. Thus, our study aimed to evaluate the possible beneficial impact of aqueous extract of milk thistle seeds (MTS) at two graded concentrations (10 and 20 ml/L of drinking water) on male Japanese quails during their reproductive period.ResultsThe phytochemical analysis revealed the presence of 29 active compounds, including nine flavonoids and ten phenolic compounds. The supplemented groups showed significant improvements in body weight gain. MTS1 group exhibited a notable decrease in daily feed consumption, while MTS2 group showed a significant increase in daily water consumption. There was a dose-dependent increase in cecum length. The total count of intestinal bacteria decreased in a dose-dependent manner. Incorporating aqueous extract of MTS at concentration of 10 ml /L resulted in a significant increase in total protein and packed cell volume. Similar increases in globulin and decreases in the albumin/globulin ratio and aspartate aminotransferase (AST) were observed with both doses of supplementation. A significant decrease in total cholesterol and AST was observed in the high-dose group. Significantly higher plasma testosterone and triiodothyronine levels were observed only in the high-dose group, while plasma thyroxine levels were similarly increased in both supplemented groups. Intervention with MTS resulted in dose-dependent increases in cloacal gland index and cloacal foam production. Both supplemented groups showed significant increases in the diameter of seminiferous tubules and the number of Sertoli cells.ConclusionMarked growth-promoting, antibacterial, and reproductive-enhancing effects were observed when incorporating aqueous extract of MTS into the quails’ drinking water, particularly at a dosage of 20 ml/L.