Development of a Simple Green Extraction Procedure and HPLC Method for Determination of Oleuropein in Olive Leaf Extract Applied to a Multi-Source Comparative Study

A simple, green and inexpensive water-based procedure was developed to extract oleuropein from olive leaf samples. Extraction was optimized in terms of the solvent, pH of the solvent, temperature and time of extraction. The experimental results revealed that deionised water adjusted to pH 3, at 60 °C for 4 h had the highest extraction efficiency. Samples were lyophilized and stored in tight containers at -18 °C until the analysis. Oleuropein was identified by comparing the retention time of the extracts with standard compound using UV detector at 280 nm. The method of analysis was based on RP-HPLC, with a mobile phase of water (adjusted to pH 3):acetonitrile (80:20 v/v) with a flow rate of 1 ml min-1. Linear dynamic range and limit of detection were found to be 50-900 mg ml-1 and 9.5 mg ml-1, respectively. Intra- and inter-day precision of the method were calculated as RSD% of 1.2 and 5.7, respectively. Olive leaves gathered from various cultivars of Iran were analyzed by the method and results showed that olive leaves from Shiraz had the highest oleuropein content (about 13 mg g-1). The developed method can be used in industry for proper mass production of this compound that is of great significance in medicine as well as food and cosmetics industry.