Identification of reactive compounds in Salvia officinalis L. extract by high-performance liquid chromatography (HPLC) in presence of 2,2′-azobis(2-amidinopropane) dihydrochloride (AAPH) radicals

In this study, a high-performance liquid chromatography (HPLC) method based on 2,2′-azobis(2-amidinopropane) dihydrochloride (AAPH) radicals was applied to investigate the scavenging activity of phenolic compounds in sage (Salvia officinalis L.) extract. This approach was selected due to its ability to simulate oxidative stress conditions that resemble those found in biological systems. To this purpose, 34 compounds were identified by HPLC-HRMS. The extract was then incubated with AAPH, which released ROO· radicals under physiological conditions (37°C, pH 7.4). Significant peak reduction rates (> 80%) in HPLC chromatograms were observed for compounds with antioxidant activity following the incubation with AAPH. Namely, rosmarinic acid hydrated (83.2±0.5%), salvianolic acid K (91.9±0.7%), rosmarinic acid (97.2±0.8%), rosmarinic acid isomer (99.7±0.1%), 6-hydroxyluteolin 7-O-glucuronide (99.8±0.1%), carnosol (95.4±0.6%), and carnosic acid (100±0.0%) reported the highest peroxyl radicals scavenging activity. Moreover, rosmarinic acid detected in the extract reported a significantly lower peak reduction rate compared to the analytical standard reacting at the same concentration (7.5±0.5 mg/g). These results suggest that other compounds present in the extract, such as phenolics or terpenes, may potentially exert a protective effect on rosmarinic acid reducing its peak reduction rate. Overall, the post-incubation HPLC-MS analysis following exposure to AAPH-derived radicals enabled the assessment of compound-specific antioxidant reactivity.