Automated accelerated salting-out solvent extraction (A-ASASE) of stilbenoids from Vitis vinifera L. co-products: fool proof or a proof of concept?

Stilbenoids are a family of naturally occurring phenolic compounds with many health benefits including antiviral, and anti-inflammatory activities. Extraction of these compounds is quite challenging as conventional extraction techniques take long time and aren’t easy to apply. In this context, we describe the first demonstration of a fully automated ASE coupled to salting-out assisted liquid-liquid extraction (SALLE). The latter is considered as an extraction and a purification technique which depends on adding a salt into a miscible mixture of two solvents usually water and an organic solvent (e.g. acetonitrile, ACN),
inducing a two phases separation (organic phase from the bulk aqueous phase). This method was given the name Automated Accelerated Salting-out Assisted Solvent Extraction (A ASASE) which makes a full use of the advantages of both techniques by combining the extraction at high temperature/pressure and an enrichment step using eco-friendly solvents (e.g., water and ethanol) in a fully integrated and simple way that requires only low volumes of solvents. The SALLE methodology was first optimised by testing different mixtures of ethanol/water at different ratios and dipotassium hydrogen phosphate (K2PO4) as the salting out agent. For each ratio, different amounts of K2HPO4 ranging from 0.5 to 10 % (m/v) were tested. A full factorial design using Minitab 19.1 program followed by statistical tests with the help of the Statgraphics Centurion 19 program were used to obtain the optimal conditions. The optimised manual SALLE methodology was evaluated for the extraction of three major stilbenes namely E-resveratrol, E-ε-viniferin and E-vitisin B from two Vitis Vinifera grape vine co-products: branches and roots. Stilbenes were efficiently extracted in the upper (mostly organic) phase which was directly lyophilised and reconstituted in a mixture of methanol/water, 50/50 (v/v). The solution was analysed by using HPLC coupled to photodiode array (DAD) detector. The optimised SALLE methodology was then, coupled to ASE by adding the salt to the extraction cell containing plant material (i.e. vine co-products). Different ASE conditions were also tested and optimised including number of cycles, number of complete run and temperature. The extract obtained was treated by the same way and analysed by HPLC coupled to DAD detector. The extraction efficiency of stilbenes was enhanced compared to conventional techniques confirming the applicability of the proposed methodology.

Authors: Ahmed Omar1,2, Marti-Mestres Gilberte3, Weber Jean-Frédéric1 and Waffo-Téguo Pierre1

1Oenology Research Unit, Molecules of Biological Interest Axis
2Department of Analytical Chemistry, Faculty of Pharmacy, Misr University for Science and Technology (MUST)

3University of Montpellier, IBMM, UMR5247, 15 Av. Charles Flahault, 34093 Montpellier, France 

*corresponding author:

Keywords: Accelerated solvent extraction (ASE), Salting-out assisted liquid–liquid extraction (SALLE), Stilbenes, Vine by-products, Eco-extraction

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