Two-step sequential supercritical fluid extracts from rosemary with enhanced anti-proliferative activity

Previously, carnosic acid and carnosol have demonstrated anti-proliferative activity against different types of cancer. To obtain extracts enriched in these two key phenolic compounds, two different processes have been developed in the present work based on the use of two-step sequential supercritical fluid extraction (SFE). By removing the interfering, less active fractions in a first step (150 or 300 bar, 40°C, neat CO2, 60 min), suitable enrichment is achieved in the second step (150 bar, 40°C, CO2 + 7% ethanol, 120 min), and this leads to carnosic acid concentrations in the extract as high as 40% of total dry weight, which are among the highest concentrations that have been described with this type of process. The enriched extracts were tested against the HT-29 human adenocarcinoma cell line, showing enhancement of their antiproliferative activity by approximately 3-fold compared to previously reported SFE rosemary extracts and higher inhibitory effects at lower concentrations (30 µg mL−1 of extract). Thus, the proposed two-step SFE process effectively improves the carnosic acid and carnosol recovery in shorter processing times (180 min vs. 300 min). Moreover, the obtained extracts possess higher anti-proliferative activity and consume less solvent.