Angelis Apostolis, Hamzaoui Mahmoud, Aligiannis Nektarios, Nikou Theodora, Michailidis Dimitris, Gerolimatos Panagiotis, Termentzi Aikaterini, Hubert Jane, Halabalaki Maria, Renault Jean-Hugues, Skaltsounis Alexios-Léandros
Department of Pharmacognosy and Natural Products Chemistry, Faculty of Pharmacy, University of Athens, Panepistimiopolis Zografou, 15771, Athens, Greece; UMR CNRS 7312, Université de Reims Champagne-Ardenne, Bât. 18, Moulin de la Housse, BP 1039, 51687 Reims, Cedex 2, France.
Department of Pharmacognosy and Natural Products Chemistry, Faculty of Pharmacy, University of Athens, Panepistimiopolis Zografou, 15771, Athens, Greece.
J Chromatogr A. 2017 Mar 31;1491:126-136. doi: 10.1016/j.chroma.2017.02.046. Epub 2017 Feb 22.
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, 17.5L of feed EVOO containing organic phase (i.e. 7L of EVOO treated) were extracted yielding 9.5g of total phenolic fraction corresponding to a productivity of 5.8g/h/L of CPE column. Regarding the second approach, the co-current process, 2L of the feed oil phase (containing to 0.8L of EVOO) were treated at 100mL/min yielding 1.03g of total phenolic fraction corresponding to a productivity of 8.9g/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 4h on a column with a capacity of 1L, 910mg of oleocanthal, 882mg of oleacein, 104mg of hydroxytyrosol were successfully recovered from 5g 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.
本文提出了一种集成提取与纯化工艺，通过无固相支持的液-液萃取和色谱技术，直接从特级初榨橄榄油（EVOO）中回收高附加值化合物。在实验室规模的离心分配萃取器（CPE）上开发了两种不同的萃取方法：一种是由几个“萃取-回收”循环组成的连续策略，另一种是基于固定相并流洗脱的连续策略。在这两种情况下，EVOO均用作稀释于食品级正己烷中的流动相（进料流动相），通过添加乙醇和水作为极性溶剂获得所需的双相体系。对于连续工艺，萃取了17.5L含有有机相的进料EVOO（即处理了7L EVOO），得到9.5g总酚类馏分，对应CPE柱的生产率为5.8g/h/L。对于第二种方法，即并流工艺，以100mL/min的流速处理2L进料油相（含0.8L EVOO），得到1.03g总酚类馏分，对应CPE柱的生产率为8.9g/h/L。然后使用逐步梯度洗脱离心分配色谱（CPC）对总酚类馏分进行分离。双相溶剂体系由不同比例（X/Y/2/3，v/v）的正己烷、乙酸乙酯、乙醇和水组成。在容量为1L的柱上进行单次4小时的运行，从5g酚类提取物中成功回收了910mg油橄榄苦醛、882mg橄榄苦苷、104mg羟基酪醇，纯度分别为85%、92%和90%。然后将CPC馏分进行正交色谱步骤（硅胶吸附或尺寸排阻色谱），从而分离出另外11种属于三萜类、酚类化合物和裂环烯醚萜类的化合物。其中，乙基亚油酸酯被发现是一种新化合物。在整个过程中，使用薄层色谱（TLC）、核磁共振（NMR）和高效液相色谱-二极管阵列检测器（HPLC-DAD）进行监测和评估。
