An integrated process for the recovery of high added-value compounds from olive oil using solid support free liquid-liquid extraction and chromatography techniques

- Two different extraction methods were developed on CPE using EVOO as mobile phase.
- The continuous strategy based on co-current elution led to high productivity of TPF.
- TPF was successfully fractionated using stepwise gradient elution CPC.
- Direct recovery of oleocanthal, oleacein and hydroxytyrosol was achieved.
- One new secoiridoid was isolated by combining orthogonal chromatographic techniques.

An integrated extraction and purification process for the direct recovery of high added value compounds from extra virgin olive oil (EVOO) is proposed by using solid support free liquid-liquid extraction and chromatography techniques. Two different extraction methods were developed on a laboratory-scale Centrifugal Partition Extractor (CPE): a sequential strategy consisting of several “extraction-recovery” cycles and a continuous strategy based on stationary phase co-current elution. In both cases, EVOO was used as mobile phase diluted in food grade n-hexane (feed mobile phase) and the required biphasic system was obtained by adding ethanol and water as polar solvents. For the sequential process, 17.5 L of feed EVOO containing organic phase (i.e. 7 L of EVOO treated) were extracted yielding 9.5 g of total phenolic fraction corresponding to a productivity of 5.8 g/h/L of CPE column. Regarding the second approach, the co-current process, 2 L of the feed oil phase (containing to 0.8 L of EVOO) were treated at 100 mL/min yielding 1.03 g of total phenolic fraction corresponding to a productivity of 8.9 g/h/L of CPE column. The total phenolic fraction was then fractionated by using stepwise gradient elution Centrifugal Partition Chromatography (CPC). The biphasic solvent systems were composed of n-hexane, ethyl acetate, ethanol and water in different proportions (X/Y/2/3, v/v). In a single run of 4 h on a column with a capacity of 1L, 910 mg of oleocanthal, 882 mg of oleacein, 104 mg of hydroxytyrosol were successfully recovered from 5 g of phenolic extract with purities of 85%, 92% and 90%, respectively. CPC fractions were then submitted to orthogonal chromatographic steps (adsorption on silica gel or size exclusion chromatography) leading to the isolation of additional eleven compounds belonging to triterpens, phenolic compounds and secoiridoids. Among them, elenolic acid ethylester was found to be new compound. Thin Layer Chromatography (TLC), Nuclear magnetic Resonance (NMR) and High Performance Liquid Chromatography - Diode Array Detector (HPLC-DAD) were used for monitoring and evaluation purposes throughout the entire procedure.