title: Use of an enzyme-assisted method to improve protein extraction from olive leaves.
abstract: The improvement of protein extraction from olive leaves using an enzyme-assisted protocol has been investigated. Using a cellulase enzyme (Celluclast® 1.5L), different parameters that affect the extraction process, such as the influence and amount of organic solvent, enzyme amount, pH and extraction temperature and time, were optimised. The influence of these factors was examined using the standard Bradford assay and the extracted proteins were characterised by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The optimum extraction parameters were: 30% acetonitrile, 5% (v/v) Celluclast® 1.5L at pH 5.0 and 55°C for 15min. Under these conditions, several protein extracts from olive leaves of different genetic variety (with a total protein amount comprised between 1.87 and 6.64mgg(-1)) were analysed and compared by SDS-PAGE, showing differences in their electrophoretic protein profiles. The developed enzyme-assisted extraction method has shown a faster extraction, higher recovery and reduced solvent usage with respect to the use of the non-enzymatic methods described in literature.
authors.affiliation: Department of Analytical Chemistry, University of Valencia, C. Doctor Moliner 50, E-46100 Burjassot, Valencia, Spain.
authors.forename: M
authors.initials: M
authors.lastname: Vergara-Barberán
authors.affiliation: Department of Analytical Chemistry, University of Valencia, C. Doctor Moliner 50, E-46100 Burjassot, Valencia, Spain.
authors.forename: M J
authors.initials: MJ
authors.lastname: Lerma-García
authors.affiliation: Department of Analytical Chemistry, University of Valencia, C. Doctor Moliner 50, E-46100 Burjassot, Valencia, Spain.
authors.forename: J M
authors.initials: JM
authors.lastname: Herrero-Martínez
authors.affiliation: Department of Analytical Chemistry, University of Valencia, C. Doctor Moliner 50, E-46100 Burjassot, Valencia, Spain. Electronic address: ernesto.simo@uv.es.
authors.forename: E F
authors.initials: EF
authors.lastname: Simó-Alfonso
mesh_terms: D002480:Cellulase / Q000737:chemistry*; D005591:Chemical Fractionation / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D031658:Olea / Q000737:chemistry*; D010936:Plant Extracts / Q000737:chemistry / Q000302:isolation & purification*; D018515:Plant Leaves / Q000737:chemistry*; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D013696:Temperature
chemical_list: D010936:Plant Extracts; D010940:Plant Proteins; D002480:Cellulase
keywords: Enzyme-assisted extraction; Genetic variety; Olive leaf protein profiles; SDS–PAGE