Copaifera langsdorffii supercritical fluid extraction: Chemical and functional characterization by LC/MS and in vitro assays

Copaifera langsdorffii is a native Brazilian plant containing important amounts of polyphenols and diterpenes with biological activity. To obtain extracts enriched in these two types of compounds, supercritical fluid extraction (SFE) conditions have been optimized by using a response surface methodology (RSM) and considering as factors: extraction pressure and temperature and percentage of co-solvent (ethanol). The response variables selected were the concentration of the two most representative compounds in C. langsdorffii, kaurenoic acid (diterpenic acid) and quercitrin (glycosylated flavonol), total flavonoids content, and antioxidant activity (measured by DPPH and TEAC assays). According to the statistical analysis of the experimental design, extraction temperature and ethanol percentage were the main factors influencing the selectivity towards the extraction of the target compounds. Two optimized extracts were obtained: (1) containing high amount of total flavonoids, high concentration of quercitrin and low EC50 for the DPPH assay, named HQE - High Quercitrin Extract (100 bar, 70 °C and 50% of ethanol, in the region of gas-expanded liquids, GXLs) and (2) containing high value of kaurenoic acid extract, named HKAE-High Kaurenoic Acid Extract, (390 bar, 70 °C and pure CO2). Chemical characterization by LC/MS of HQE showed the presence of galloylquinic acids and flavonoids while, on the other hand, HKAE presented only nonpolar compounds such as kaurenoic acid. Moreover, a solid-liquid extraction (SLE) with methanol was carried out as a benchmark protocol for comparative purposes. SFE presented approximately 10-fold higher content of total flavonoids than the extracts obtained by SLE.