This study aimed to compare the efficiency of different green extraction methods for obtaining bioactive compounds from the roots of Rheum cordatum Losinsk and to evaluate their antioxidant and antimicrobial properties. The presence of some important phytochemicals in the extracts obtained using ultrasound-assisted extraction (UAE), subcritical ethanol extraction (Sbc-EtOH), and supercritical CO₂ (ScCO₂) extraction was determined by LC-MS/MS, and their antioxidant and antimicrobial properties were examined against Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa, and Escherichia coli. The goal was to determine the optimal extraction conditions that maximize the yield of bioactive compounds while preserving their biological properties. Different pressures (100 bar and 400 bar) were tested in UAE extraction, different solvents and times were tested in Sbc-EtOH extraction, and different pressures were tested in ScCO₂ extraction. Most of the 53 important phenolic compounds have been extracted using the ScCO₂ extraction method, either exclusively or in the highest amounts. It has been observed that more and higher amounts of phenolic compounds were extracted at lower pressure. The highest antioxidant activity was exhibited by the ScCO₂ extracts. Additionally, the ScCO₂-100 extract obtained at 100 bar showed strong antimicrobial activity, with a minimum inhibitory concentration (MIC) ranging from 31.25 to 250 μg/mL. Gallic acid, epicatechin gallate, epigallocatechin gallate, and catechin were found in extracts. Additionally, molecular docking studies against the 1QWZ, 2ANQ, 3H77, and 6QXS proteins revealed that epicatechin exhibited docking scores of −6.127, −9.479, −5.836, and −7.067 kcal/mol, respectively.

Rheum cordatum Losinsk is a plant species distributed in Kazakhstan but remains relatively understudied despite its promising biological potential. The present study aimed to explore leaf extracts of R. cordatum by utilizing advanced green extraction technologies including supercritical CO₂ (ScCO₂), subcritical ethanol (Sc) and ultrasound-assisted extraction (UAE) to characterize their phytochemical composition and evaluate their antioxidant and antimicrobial activities. A total of 53 phytochemical compounds were identified, with gallic acid (30.71 µg/mg UAE-EtOH-4h), rutin (21.93 µg/mg ScCO₂-150) and hesperidin (14.98 µg/mg ScCO₂-150) being notably abundant. Among the tested extracts, ScCO₂ extraction at 150 bar (ScCO₂-150) demonstrated the highest antioxidant activity, exhibiting IC₅₀ values of 0.0132 mg/mL (DPPH) and 0.0462 mg/mL (ABTS), coupled with the highest total phenolic content (140 mg GAE/g). Moreover, the ScCO₂-150 extract showed pronounced antimicrobial efficacy, particularly against Bacillus subtilis Pseudomonas aeruginosa and Staphylococcus aureus, with minimum inhibitory concentrations (MIC) ranging from 125 to 250 µg/mL. These findings highlight the considerable potential of R. cordatum leaves as a valuable, abundant and sustainable source of natural antioxidants and antimicrobial agents, with supercritical CO₂ extraction presenting substantial advantages in selectively obtaining bioactive phytochemicals.